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THE ANALYSTTHE ANALYTICAL JOURNAL OF THE ROYAL SOCIETY OF CHEMISTRYADVISORY BOARD'Chairman: J. M. Ottaway (Glasgow, U.K.)'L. S. Bark (Salford, U.K.)E. Bishop (Exeter, U.K.)W. L. Budde (U.S.A.)D. T. Burns (Belfast, U.K.)L. R. P. Butler (South Africa)H. J. Cluley (Wembley, U.K.)E. A. M. F. Dahmen (The Netherlands)L. de Galan (The Netherlands)A. C. Docherty (Billingham, U.K.)D. Dyrssen (Sweden)G. Ghersini (Italy)J. Hoste (Be!gium)A. Hulanicki (Poland)'G. W. Kirby (Glasgow, U.K.)W. S. Lyon (U.S.A.)H. V. Malmstadt (U.S.A.)G. W. C. Milner (Harwell, U.K.)*A. C. Moffat (Aldermaston, U.K.)E. J. Newman (Poole, U.K.)H. W. Nurnberg (West Germany)"T. B. Pierce (Harwell, U.K.)E. Pungor (Hungary)P. H. Scholes (Middlesbrough, U.K.)D. Simpson (Thorpe-le-Soken, U.K.)"J.M. Skinner (Billingham, U.K.)"J. D. R. Thomas (Cardiff, U.K.)K. C. Thompson (Sheffield, U.K.)"A. M. Ure (Aberdeen, U.K.)A. Walsh, K.B. (Australia)G. Werner (German Democratic Republic)T. S. West (Aberdeen, U.K.)"P. C. Weston (London, U.K.)'J. Whitehead (Stockton-on-Tees, U.K.)J. D. Winefordner (U.S.A.)P. Zuman (U.S.A.)'G. J. Dickes (Bristol, U.K.)'Members of the Board serving on the Analytical Editorial BoardEditor: P. C. WestonSenior Assistant Editor: R. A. YoungAssistant Editors: Mrs. J. Brew, Miss D. ChevinREG I ONAL ADVlSO RY EDIT0 RSDr. J. Aggett, Department of chemistry, University of Auckland, Private Bag, Auckland, NEW ZEALAND.Professor L. Gierst, Universit6 Libre de Bruxelles, Facult6 des Sciences, Avenue F.-D.Roosevelt 50,Professor H. M. N. H. Irving, Department of Theoretical Chemistry, University of Cape Town, Ronde-Professor W. A. E. McBryde, Faculty of Science, University of Waterloo, Waterloo, Ontario, CANADA.Dr. 0. Osibanjo, Department of Chemistry, University of Ibadan, Ibadan, NIGERIA.Dr. G. Rossi, Chemistry Division, Spectroscopy Sector, CEC Joint Research Centre, EURATOM, lspraDr. 1. RubeGka, Geological Survey of Czechoslovakia, Malostranskh 19, 1 1 8 21 Prague 1, CZECHO-Professor J. R&iEka, Chemistry Department A, Technical University of Denmark, 2800 Lyngby,Professor K. Saito, Department of Chemistry, Tohoku University, Sendai, JAPAN.Professor L. E. Smythe, Department of Chemistry, University of New South Wales, P.O.Box 1,Professor P. C. Uden, Department of Chemistry, University of Massachusetts, Amherst, MA 01 003,Editorial: Editor, The Analyst, The Royal Society of Chemistry, Burlington House,Piccadilly, London, W1 V OBN. Telephone 01 -734 9864. Telex No. 268001Advertisements: Advertisement Department, The Royal Society of Chemistry, Burlington House,Piccadilly, London, WIV OBN. Telephone 01 -734 9864. Telex No. 268001The Analyst (ISSN 0003-2654) is published monthly by The Royal Society of Chemistry, BurlingtonHouse, London W1V OBN, England. All orders accompanied with payment should be sent directly toThe Royal Society of Chemistry, The Distribution Centre, Blackhorse Road, Letchworth, Herts. SG6 1 HN,England. 1983 Annual subscription rate UK f93.50, Rest of World f99.00, USA $201 .OO.Purchased withAnalytical Abstracts UK f226.50, Rest of World f238.50, USA $487.00. Purchased with AnalyticalAbstracts plus Analytical Proceedings UK f251 .OO, Rest of World f265.00, USA $539.00. Purchasedwith Analytical Proceedings UK f 1 1 7.50, Rest of World f 124.50, USA $253.00. Air freight and mailingin the USA by Publications Expediting Inc., 200 Meacham Avenue, Elmont, NY 11003.USA Postmaster: Send address changes to: The Analyst, Publications Expediting Inc., 200 MeachamAvenue, Elmont, NY 11003. Second class postage paid a t Jamaica, NY 11431. All otherdespatches outside the UK by Bulk Airmail within Europe, Accelerated Surface Post outside Europe.PRINTED IN THE UK.Q The Royal Society of Chemistry, 1983. All rights reserved. No part of this publication may be reproduced,stored in a retrieval system, or transmitted in any form, or by any means, electronic, mechanical,photographic recording, or otherwise, without the prior permission of the publishers.Bruxelles, BELGIUM.bosch 7700, SOUTH AFRICA.Establishment, 21 020 lspra (Varese), ITALY.SLOVAKIA.DEN MARK.Kensington, N.S.W. 2033, AUSTRALIA.U.S.A

ISSN:0003-2654    

DOI:10.1039/AN98308FX025

出版商:RSC    

年代:1983

数据来源: RSC

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ANALAO 108 (1 288) 777-904 (1 983) July 1983THE ANALYST77780881 382182783584084785185786487088088688989389 5899THE ANALYTICAL JOURNAL OF THE ROYAL SOCIETY OF CHEMISTRYCONTENTSThe Forensic Analysis of Drugs o f Abuse. A Review-Peter B. Baker and Geoffrey F.Phi II i psDesign and Performance o f an Unenclosed Tantalum Atomiser f o r Atomic-absorptionS pectrosco py-J. AggettDetermination of Lead in Blood by Atomic-absorption Spectroscopy with ElectrothermalAtomisation-Denis J. Hodges and (the late) Douglas SkeldingSelective Spectrophotometric Determination o f Trace Amounts o f Iron w i t h Di(2-pyridyl)-NN-di[(8-quinolyl)amino]methane: Determination o f Iron i n BloodSerum-R. Escobar and J. M. Can0 PavonDirect Determination o f Inorganic Mercury i n Biological Materials after Alkali Digestionand Amalgamation-Tetsuro Konishi and Hitoshi TakahashiDirect Determination o f Selenium(1V) in Biological Samples by Cathodic-strippingVoltammetry-Rahmalan bin Ahmad, John 0.Hill and Robert J. MageeOn-line Bromimetric Determination of Phenol, Aniline, Aspirin and lsoniazid Using FlowInjection Voltammetry-Arnold G. Fogg, Md. Ashraf Ali and Mohamed A. AbdallaPolarographic Determination o f Adrenaline from the Catalysed Wave o f Germanium(1V)-Juan Miguel L6pez Fonseca and Maria del Carmen ArredondoStability o f Procaine in Aqueous Systems-Da-Peng WangSemi-automatic Catalytic Titration of Aminopolycarboxylic Acids and Metal Ions withthe 4,4'-Dihydroxybenzophenone Thiosemicarbatone - Hydrogen Peroxide -Copper(l1) System-T.Raya-Saro and D. PBrez-BenditoExtraction and Spectrophotometric Determination o f Cobalt in Coal Fly Ashes Using2-[2-(3,5-Dibromopyridyl)azo]-5-dimethylaminobenzoic Acid-Takeo Katarni,Tornokuni Hayakawa, Masamichi Furukawa and Shozo ShibataDetermination o f Iron(ll1) and Copper(l1) by Zeroth, First and Second Derivative Spectro-photometry w i t h 2-Thiobarbituric Acid (4,6-Dihydroxy-2-mercaptopyrimidine) asReagent-Basilio MorelliSelective Cation-exchange Separation o f Lead on Tin( 11) Hexacyanoferrate( 111) Gel-Ajay K. Jain, Raj Pal Singh and Chand BalaSHORT PAPERSColorimetric, Spectrophotometric and Fluorimetric Determination o f PrenalterolHydrochloride-Abdel-Aziz M. Wahbi, Mohamed E.Mohamed, El Rasheed A. Gad Kariemand Hassan Y. Aboul-EneinSpecific Spot Test f o r Gold(ll1) with Primaquin-Shen Nai-Kui, Zhao Xiang-Cong and ChuWen-TienCO M M U NICATIO NSAutomatic Graphite Probe Sample Introduction f o r Electrothermal Atomic-absorptionSpectrometry-David Littlejohn, John Marshall, John Carroll, William Cormack andJohn M. OttawayImportance o f Selectivity in the Determination o f N-Nitroso Compounds as a Group-P. L. R. Smith, Clifford L. Walters and P. I. ReedBOOK REVIEWSSummaries of Papers in this issue-Pages iv, v, vi, vii, viii, ix,Printed by Heffers Printers Ltd Cambridge EnglandEntered as Second Class at New York, USA, Post OfficThe Periodic Table ofthe ElementsThe Royal Society of Chemistry has produced acolourful wall chart measuring 125cm x 75cmcovering the first 105 elements as they existtoday.Each group is pictured against the same tintedbackground and each element, where possiblephotographed in colour and discussed withregard to its position in the hierarchy of matter.Additional information for each element includeschemical symbol, atomic number, atomic weightand orbits of electrons.The chart is particularly useful for both teachersand students and would make a worthwhileaddition to any establishment.Price f2.20 ($4.50) RSC Members f 1 .OOTeacher Members f4.60 for 10Prices subject to VAT in the UKRSC members should send their orders to: TheRoyal Society of Chemistry, The MembershipOfficer, 30 Russell Square, London WClB 5DT.Non-RSC members should send their orders to:The Royal Society of Chemistry, DistributionCentre, Blackhorse Road, Letchworth, HertsSG6 1HN.BUREAU OF ANALYSEDSAMPLES LTDannounce the availability ofTWO NEWHIGH MANGANESE STEELSPECTROSCOPICSTANDARD CERTIFIEDREFERENCE MATERIALSSS-CRM 491/1 & 494/1For full details of these, and for copiesof new catalogue of all CRMs suppliedby BAS, write, telephone or telex to:BAS Ltd., Newham Hall, Newby,Middlesbrough, Cleveland, TS8 9EATelephone: Middlesbrough 31 721 6Telex: 587765 BASRID9201 for further information.See page x.~~ ~~ ~Annual Reports on AnalyticalAtomic Spectroscopy Vol. 11Edited by M. S. Cresser and 8. L. SharpThis volume reports on current developmentsin all branches of analytical atomic emission,absorption and fluorescence spectroscopywith references to papers published andlectures presented during 1981.Much of theinformation is i n tabular form for ease ofreference.Brief ContentsAtomization and ExcitationArcs, Sparks, Lasers and Low-pressure Dis-charges; Plasmas; Flames; ElectrothermalAtomization; Vapour Generation;InstrumentationLight Sources; Optics; Detector Systems;Instrument Automation; Complete Instru-ments; New Commercial Instruments;MethodologyNew Methods; Detection Limits, Precision andAccuracy; Standards and Standardization;ApplicationsChemicals; Metals; Refractories and MetalOxides, Ceramics, Slags, Cements; Minerals;Air Analysis; Water Analysis; Soils, Plants andFertilizers, Foods and Beverages; Body Tissuesand Fluids;Hardcover 388pp 0 85186 707 3f48.00 ($88.00) RSC Members f32.00RSC members should send their orders to: The Royal Society of Chemistry, The Membership Officer,30 Russell Square, London WClB 5DT. Non-RSC members should send their orders to: The RoyalThe Royal Society of ChemistryBurlington HouseLondon W1V OBNistry, Distribution Centre, Blackhorse Road, Letchworth, Herts SG6 1 HN

ISSN:0003-2654    

DOI:10.1039/AN98308BX027

出版商:RSC    

年代:1983

数据来源: RSC

摘要:

iV SUMMARIES OF PAPERS I N THIS ISSUE July, 1983Summaries of Papers in this IssueThe Forensic Analysis of Drugs of AbuseA ReviewSummary of ContentsIntroduction and scopeScreening techniquesIntroductionPhysical characteristics of drug samplesChemical spot testsField testsUltraviolet spectrometryInfrared spectrometryMicrocrystal testsChromatographic techniquesNuclear magnetic resonance spectrometryMass spectrometryMiscellaneous techniquesCannabis productsOpium and poppy strawNarcoticsCoca leafCocaineLSD and indolic hallucinogensAmphetamines and structurally related drugsPhencyclidine and analoguesMethaqualoneBarbituratesConcluding remarksKeywords : Review ; forensic analysis ; drug analysisPETER B. BAKER and GEOFFREY F.PHILLIPSLaboratory of the Government Chemist, Cornwall House, Stamford Street, London,SE19NQ.Analyst, 1983, 108, 777-807.Design and Performance of an Unenclosed Tantalum Atomiserfor Atomic-absorption SpectroscopyAn unenclosed tantalum-strip atomiser for atomic-absorption spectroscopyis described. Its performance for the determination of a number of elementshas been evaluated and several applications are reported.Keywords : Tantalum-strip atomiser ; atomic-absorption spectroscopyJ. AGGETTChemistry Department, University of Auckland, Auckland, New Zealand.Analyst, 1983, 108, 808-812July, 1983 SUMMARIES OF PAPERS I N THIS ISSUEDetermination of Lead in Blood by Atomic-absorptionSpectroscopy with Electrothermal AtomisationA method is described for the determination of lead in blood by atomic-absorption spectroscopy with electrothermal atomisation.The addition oforthophosphoric acid together with pre-coating the graphite tube withmolybdenum is effective in minimising matrix interference and in promotingstable routine operation. No special oxidative treatment is necessary toprevent loss of lead during the drying and ashing steps of the procedure.The range of the method is 5-50 p g dl-1.Keywords ; Lead determination ; blood ; electrothermal atomisation atomic-absorption spectroscopy ; matrix modificationDENIS J. HODGES and (the late) DOUGLAS SKELDINGResearch and Development Department, The Associated Octel Company Limited,P.O. Box 17, Oil Sites Road, Ellesmere Port, South Wirral, L65 4HF.Analyst, 1983, 108, 813-820.Selective Spectrophotometric Determination of Trace Amounts ofIron with Di(2-pyridyl) -NN- di[( 8- quinolyl)amino]methane :Determination of Iron in Blood SerumDi(2-pyridyl)-NN-di[(8-quinolyl)amino]methane (DPQAM) has been exam-ined to evaluate its usefulness as a sensitive and selective spectrophotometricreagent for iron.A green complex is formed, which is extracted into chloro-form at pH 2.9-5.4 in the presence of perchlorate and ascorbic acid (the molarabsorptivity, E = 1.14 x lo* 1 mol-1 cm-1 at 693 nm). The distributionratio of this extraction is very favourable; so that iron can be determined inthe range 0.25-2.50 p.p.m. ( E = 1.71 x 1051mol-1cm-1). The effect offoreign ions has been studied and the method has been applied to the deter-mination of iron in blood serum with good results.Keywords : Iron determination ; spectrophotometry ; di (8-pyridyZ)-NN-di-[ (8-quinolyl)amino]methane ; blood semmR.ESCOBARDepartment of Analytical Chemistry, Faculty of Chemistry, University of Seville,Seville-4, Spain.and J. M. CAN0 PAVONDepartment of Analytical Chemistry, Faculty of Sciences, University of Malaga,Malaga-4, Spain.Analyst, 1983, 108, 821-826.vi SUMMARIES OF PAPERS IN THIS ISSUEDirect Determination of Inorganic Mercury in BiologicalMaterials after Alkali Digestion and AmalgamationJztly, 1983A reliable method is described for the determination of inorganic mercury inbiological materials in the presence of organic mercury.This is principallybased on the fact that hydrogen peroxide oxidatively liberates inorganicmercury from organic substances in strong alkali and reduces i t to the metallicstate without decomposing organic mercurials concomitantly present. Themetallic mercury, vaporised with a nitrogen stream, is trapped by gold amalga-mation and then released for electrothermal atomisation atomic-absorptionspectrometry. The detection limit is 1 ng of inorganic mercury and the co-efficient of variation for 40 ng of inorganic mercury is 2.8%.Keywords : Inorganic mercury determination ; biological materials ; hydrogenperoxide reduction ; gold amalgamation ; atomic-absorption spectrometryTETSURO KONISHIKumamoto Municipal Institute of Public Health, 269 Tainoshima, Tamukaemachi,Kumamoto 862, Japan.and HITOSHI TAKAHASHIDepartment of Pharmacology, Institute for Medical Immunology, Kumamoto Uni-versity Medical School, Kumamoto 860, Japan.Analyst, 1983, 108, 827-834.Direct Determination of Selenium( IV) in Biological Samplesby Cathodic-stripping VoltammetryThe use of a commercially available digesting solvent, Lumatom, for thedetermination of selenium (IV) in biological samples using cathodic-strippingvoltammetry is reported.I t was found possible to determine seleniumdirectly in biological samples containing metals such as arsenic(III), copper(II),lead(II), iron(II1) and zinc(I1). The results for selenium in bovine liver andoyster tissue are reported.Keywords : Selenium determination ; biological samples ; cathodic-strippingvoltammetryRAHMALAN BIN AHMAD, JOHN 0.HILL and ROBERT J. MAGEEDepartment of Inorganic and Analytical Chemistry, La Trobe University, Melbourne,Victoria, Australia 3083.Analyst, 1983, 108, 835-839.On-line Bromimetric Determination of Phenol, Aniline, Aspirinand Isoniazid Using Flow Injection VoltammetryBromine is produced on-line in a flow injection system by injection of acidinto neutral or slightly alkaline bromate - bromide eluent and is determinedvoltammetrically at a glassy carbon electrode. Organic compounds that canbe brominated or oxidised by bromine are determined by injecting them, inacidic solution, into the bromate - bromide eluent and noting the decreasein the bromine signal. Determinations can also be made by injecting pre-reacted solutions of bromine and the organic compounds in order to monitorthe excess of bromine.Keywords : Flow injection analysis ; voltammetry ; bromination reactionsARNOLD G. FOGG, MD. ASHRAF ALI and MOHAMED A. ABDALLAChemistry Department, Loughborough University of Technology, Loughborough,Leicestershire, LE11 3TU.Analyst, 1983, 108, 840-846

ISSN:0003-2654    

DOI:10.1039/AN98308FP069

出版商:RSC    

年代:1983

数据来源: RSC

摘要:

July, 1983 SUMMARIES OF PAPERS I N THIS ISSUEPolarographic Determination of Adrenaline from theCatalysed Wave of Germanium(1V)Adrenaline catalyses the reduction of germanium(1V) a t the dropping-mercury electrode in perchloric acid media. The catalytic polarographicwave obtained has been studied in detail and a mechanism is proposed forthe electrode process, which involves the formation of a catalytic complexbetween protonated H,GeO, and adrenaline adsorbed on to the surface ofthe electrode, followed by electrochemical reduction of the complex andregeneration of the catalytic ligand. For the determination of adrenalinethe best system has been found to be 1.5 x lo-, M germanium(1V) - 1 Mperchloric acid with ascorbic acid as antioxidant a t a concentration below1 x The calibration graph is linear over the range of adrenalineconcentrations 3 x 10-5-70 x l o - 5 ~ .An application of the method toinjectable solutions is described.Keywords : Adrenaline determination ; d.c. polarogvaphy ; germanium(I V )JUAN MIGUEL LOPEZ FONSECA and MARIA DEL CARMEN ARRE-DONDODepartamento de Fisico-Quimica Aplicada, Facultad de Farmacia, Universidad deGalicia, Santiago de Compostela, Spain.Analyst, 1983, 108, 847-850.catalysed waveStability of Procaine in Aqueous SystemsA stability-indicating assay for procaine, based on high-performance liquidchromatography (HPLC), is described. A mixture of methanol and 1%acetic acid (40 + 60) was employed as the mobile phase on a pBondapakC,, column for HPLC. This method is simple, accurate and requires noextraction procedure.It was used to study procaine stability in variousaqueous solutions. Stability tests were conducted both a t room tempera-ture and a t high temperatures (70-90 "C) a t various p H values and withdifferent vehicles. Solutions of procaine in mixed aqueous solution (propyleneglycol - ethanol - water) can enhance the stability relative to aqueous solu-tions.Keywovds : Stability assay ; procaine ; aqueous systems ; high-performanceliquid chromatogvaphyDA-PENG WANGDepartment of Pharmacy, National Defence Medical Centre, Taipei 107, Republic ofChina.Analyst, 1983, 108, 851-856.Semi- automatic Catalytic Titration of Aminopolycarboxylic Acidsand Metal Ions with the 4,4'-DihydroxybenzophenoneThiosemicarbazone - Hydrogen Peroxide - Copper( 11) SystemA semi-automatic spectrophotometric method is described for the catalytictitration of EDTA and ethylene glycol bis( p-aminoethyl ether)-"'-tetra-acetic acid (EGTA), based on their inhibitory effect on the copper(I1)-catalysed oxidation of 4,4'-dihydroxybenzophenone thiosemicarbazone byhydrogen peroxide.Amounts of EDTA and EGTA in the 370-1100 and380-1 140 pg ranges, respectively, can be determined with a relative standarddeviation of about 0.5% (n = 11). Methods are also described for the indirectcatalytic titrations of copper(II), nickel(I1) and manganese(II), in the range10-120 pg, with a relative standard deviation of about 0.60/, (n = 11).Keywovds : EDTA and EGTA ; coppev, nickel and manganese detevwination;spectvophotometric catalytic titvation; 4,4'-dihydvoxybenzophenone thio-semicarbazoneT.RAYA-SARO and D. PBREZ-BENDITODepartment of Analytical Chemistry, Faculty of Sciences, University of Chrdoba,Cbrdoba, Spain.Analyst, 1983, 108, 857-863.viviii SUMMARIES OF PAPERS I N THIS ISSUEExtraction and Spectrophotometric Determination of Cobalt inCoal Fly Ashes Using 2- [2- (3,5-Dibromopyridyl)azo] -5-dimethyl-aminobenzoic AcidJuly, 1983The dibromo derivative of 2-(2-pyridylazo) -5-dimethylaminobenzoic acid wassynthesised and evaluated as a chromogenic reagent for the determination oftrace amounts of cobalt. The reagent was very sensitive and reacted withcobalt(II1) to form a 1:2 (Co:reagent) stable green complex, which wasextracted into dichloromethane.The apparent molar absorptivity of thecobalt(II1) complex was 1.55 x lO51mo1-lcm-l at 673nm in dichloro-methane. The reagent was used in the determination of trace amounts ofcobalt in coal fly ashes and the results were compared with those obtainedusing the nitroso-R method.Keywords : Cobalt determination ; coal ~ 7 y ash analysis ; spectrophotometry ;2-[2-( 3,5-dibromo~yridyl)azo]-5-dimethylaminobenzoic acidTAKE0 KATAMI and TOMOKUNI HAYAKAWAGifu Prefectural Research Institute for Environmental Pollution, 58-8, Yabuta, Gifu-shi, 500 Japan.MASAMICHI FURUKAWA and SHOZO SHIBATAGovernment Industrial Research Institute, Nagoya, Kita-ku, Nagoya-shi, 462 Japan.Analyst, 1983, 108, 864-869.Determination of Iron(II1) and Copper(I1) by Zeroth, First andSecond Derivative Spectrophotometry with 2-Thiobarbituric acid(4,6-Dihydroxy-2-mercaptopyrimidine) as ReagentSensitive methods for the determination of iron(II1) alone and of iron(II1)and copper(I1) in mixtures, using 4,6-dihydroxy-2-mercaptopyrimidine(2-thiobarbituric acid), by normal and derivative spectrophotometry havebeen studied and compared experimentally. Fundamental conditions forthe determination and the composition of the iron(II1) - 2-thiobarbituricacid complex are discussed.A statistical analysis of the results is reported.Keywords : Derivative spectrophotometry ; iron(III) determination ; co$$er(II)determination ; 2-thiobarbituric acidBASIL10 MORELLIIstituto di Chimica Analitica, Universitk di Bari, Via Amendola 173, 70126-Bari,Analyst, 1983, 108, 870-879.Italy.Selective Cation-exchange Separation of Lead on Tin(I1)Hexacyanoferrate( 111) GelTin(I1) hexacyanoferrate(II1) gel, prepared by adding tin(I1) chlaride topotassium hexacyanoferrate(II1) solution and heating at 80 "C, has beenfound to be stable in acids and salt solutions.Distribution coefficients,determined for various metal ions, show that the exchanger has a highaffinity for lead. Binary separations of lead from a number of other metalions at different concentrations were achieved on the column of the exchangerand a ternary separation of Pb2+ - Cu2+ - Zn2+ was also carried out.Keywords : Tin(II) hexacyanoferrate(III) ; ion-exchange col.umn ; static anddynamic sorption of leadAJAY K. JAIN, RAJ PAL SINGH and CHAND BALADepartment of Chemistry, University of Roorkee, Roorkee-247672, India.Analyst, 1983, 108, 880-885Juuly, 1983 SUMMARIES OF PAPERS I N THIS ISSUEColorimetric, Spectrophotometric and Fluorimetric Determinationof Prenalterol HydrochlorideShort PaperKeywords : Prenalterol hydrochloride ; colorimetry ; fluorimetry ; spectrophoto-metry ; tabletsABDEL-AZIZ M.WAHBI, MOHAMED E. MOHAMED, EL RASHEED A.GAD KARIEM and HASSAN Y. ABOUL-ENEINDepartment of Pharmaceutical Chemistry, College of Pharmacy, King Saud Uni-versity, Riyadh, Saudi Arabia.Analyst, 1983, 108, 886-889.Specific Spot Test for Gold(II1) with PrimaquinShort PaperKeywords : Spot test ; gold ; primaquinSHEN NAI-KUI, ZHAO XIANG-GONG and CHU WEN-TIENDepartment of Chemistry, Bengbu Medical College, Bengbu, Anhui Province] China.Analyst, 1983, 108, 889-892.Automatic Graphite Probe Sample Introduction for ElectrothermalAtomic-absorption SpectrometryCommunicationKeywords : A tomic-absorption spectrometry ; electrothermal atomisation ;graphite probe sample introduction ; automatic probe mechanismDAVID LITTLEJOHN, JOHN MARSHALL, JOHN CARROLL, WILLIAMCORMACK and JOHN M.OTTAWAYDepartment of Pure and Applied Chemistry, University of Strathclyde, CathedralStreet, Glasgow, G1 1XL.Analyst, 1983, 108, 893-896.Importance of Selectivity in the Determination ofN-Nitroso Compounds as a GroupCommunicationKeywords : N-Nitroso compounds ; denitrosation ; nitrite ; nitrate ; nitrogenoxideP.L. R. SMITH and CLIFFORD L. WALTERSLeatherhead Food Research Association, Leatherhead, Surrey, KT22 7RY.and P. I. REEDGastrointestinal Unit, Wexham Park Hospital, Slough, Berkshire.ixAnalyst, 1983, 108, 896-898TUCK IN UNDER FLAP ATHE ANALYST July, 1983READER ENQUIRY SERVICE1 H l l(Please use BLOCK CAPITALS)NAM E ......................................................................................................................................................................................... iOCCUPATION ................................................................................................................................................................. iADDRESS ............................................................................................................................................................................. iSECOND FOLDPostagewill bePaid byLicenseeDo not affix Postage Stamps if posted inGt. Britain, Channel islands or N. IrelandI BUSINESS REPLY SERVICE I Licence No. W.D. 106Reader Enquiry ServiceThe AnalystThe Royal Society of Chemistry2 I :Burlington HousePiccadilly London W1 E 6WFENGLANDTHIRD FOLD- n * 5 ;y ig ! " Iin i. i .* .. * . . :

ISSN:0003-2654    

DOI:10.1039/AN98308BP073

出版商:RSC    

年代:1983

数据来源: RSC

摘要:

July 1983 Vol. 108 No. 1288 The Analyst The Forensic Analysis of Drugs of A Review Peter B. Baker and Geoffrey F. Phillips Abuse Laboratory of the Government Chemist Cornwall House Stamford Street London SEI 9NQ Summary of Contents Introduction and scope Screening techniques Introduction Physical characteristics of drug samples Chemical spot tests Field tests Ultraviolet spectrometry Infrared spectrometry Microcrystal tests Chromatographic techniques Nuclear magnetic resonance spectrometry Mass spectrometry Miscellaneous techniques Cannabis products Opium and poppy straw Narcotics Coca leaf Cocaine LSD and indolic hallucinogens Amphetamines and structurally related drugs Phencyclidine and analogues Methaqualone Barbiturates Concluding remarks Keywords Review ; forensic analysis ; drug analysis Introduction and Scope The principal aim of this review is to aid forensic drug analysts by collating information on different analytical methods under the heading of the drug or group of drugs to which such techniques have been usefully applied.A secondary purpose is to bring together background information on drug analysis that the scientist may find helpful when called upon to present evidence in Courts-of-Law or Courts-Martial. The reviewers do not propose to recommend specific methods of analysis for particular drugs but to suggest approaches that may help the scientist to select a technique appropriate to the problem in hand. The scope of this review has deliberately not been extended to cover the analytical tech-niques that particularly apply to the analysis of drugs from physiological specimens.The reviewers consider simple separation and unequivocal identification of the very small amounts of drugs found in such samples to be a specialised analytical field which merits discussion as a separate topic. The reviewers have drawn on the experience of their colleagues past and present at the Laboratory of the Government Chemist and other forensic science laboratories in the UK and overseas in their selection of the drugs and drug classes covered in this review ; it is therefore restricted to those substances that a practising forensic drug analyst is likely to encounter albeit rarely in some instances. Few details of actual methods are presented Crown Copyright.77 778 BAKER AND PHILLIPS THE FORENSIC Analyst Vol. 108 as the reader is referred to the original literature throughout this review. No topic as wide as forensic drug analysis can be comprehensively covered in one review; however about 1300 original papers books and other documents have been read by the reviewers to provide the background information that has been selected and where necessary critically reviewed. Literature available to the reviewers up to the end of 1981 has been surveyed. Screening Techniques Introduction There is not only a wide range but also many different presentations of drugs that may be submitted to the forensic analyst for identification. Many thousands of drugs are currently available licitly either over-the-counter or upon presentation of a prescription from a medical practitioner.Considerable numbers of drugs with potential for abuse or misuse are further controlled by law. New products appear continually often in a variety of presentations. The analyst may be asked to examine many products ranging from tablets and capsules bearing easily identifiable manufacturers’ markings to illicit products of poor quality and unknown composition. Amounts may vary from traces in clothing motor vehicles syringes or smuggling concealments to seizures of many tons. Samples may consist of synthetic drugs or vegetable matter or a mixture of the two. Purity may vary from pharmaceutical-grade drug substance to fractions of a per cent. in samples found at “street” level which may be adulterated (“cut”) with physiologically inactive materials or mixed with other drugs.Drugs in solution may be encountered in or from ampoules or concealed within licit media such as wines and spirits. They may be present either as a salt or the free base or acid. It may be necessary for the analyst to consider the stereoisomeric composition of the material. However although the list of drugs for analysis may be potentially large, only a relatively small number of drugs are commonly encountered. The experienced analyst may swiftly be able tentatively to identify many drugs from their physical appearance, but in laboratories where the caseload is low or where analysts are lacking in experience, identification must begin with simple screening tests. Physical Characteristics of Drug Samples Solid drug samples submitted to the analyst may either be licit preparations (tablets, capsules or ampoules) or illicitly manufactured materials.The latter may be either in raw (undivided) form or as an illicit preparation that is tabletted or otherwise divided into crude dosage forms. The majority of legally manufactured tablets most of which do not contain drugs controlled by the Misuse of Drugs Act 1971,l may be tentatively identified from readily available guides covering the UK2-4 and many other parts of the world (e.g. references5-*). A new tablet and capsule identification system (Tablident) has recently been proposed to cover UK preparationsg; the system is based on measurement of size and the colour of the sample as in current guides but uses a computer for storing background data.Confirmation of the active constituents of a licit preparation may then be carried out by a simple technique such as ultraviolet (UV) or infrared (IR) spectrometry or by chromato-graphy. Many forensic science laboratories have large collections of licit drug presentations, which are useful for comparison with submitted samples. In addition the reviewers have invariably found drug manufacturers to be extremely helpful in identification of licit presenta-tions bearing a maker’s mark or code but which are unfamiliar to the laboratory. Further confirmatory tests on materials identified in this way are mandatory in view of the number of forged presentations on the illicit market many of which may be of high quality and bear manufacturers’ markings perhaps from stolen or forged tablet punches.Where marks are absent on tablets or capsules screening is of course necessary and identification of the actual drug present is made by an appropriate analytical technique. Considerable useful evidence concerning manufacture may be deduced from a detailed physical examination of illicit tablets; this has been discussed by Gomm et al.1° Simulations of licit tablets have been encountered that contain drugs not present in authentic material. The reviewers advise confirmation of the identity of a supposedly licit preparation by comparison of its properties with an authentic sample. Further studies on the analysis of dyes used in illicit tablets have also been made.11J2 Individual illicit drugs are discussed under appropriate headings in this review J d y 1983 ANALYSIS OF DRUGS OF ABUSE 779 Few botanical species are controlled by the Misuse of Drugs Act 1971l those which are so controlled will be discussed under the appropriate drug class heading.Der Marderosian and Chao13 presented a brief review of the problems involved in the identification of botanical material. Jolliffe and Jolliff el4 described a computer-aided identification programme for powdered vegetable drugs which they considered would enable a relatively inexperienced microscopist to identify such materials. However in the reviewers’ opinion the recognition of botanical material solely by microscopy should not be undertaken without professional training in botany. Chemical Spot Tests Many reagents have been suggested for use in spot tests for particular drugs; for example, more than 50 have been put forward for qualitative and quantitative reaction with opium alkaloids,15 and it is beyond the scope of this review to discuss their uses and limitations more than briefly.Before the advent of more sophisticated and readily available equipment colour tests were often employed for identification; even as recently as 1971 Hider16 described a system for the rapid identification of unknown drugs using colour and microcrystal tests. Despite this author’s emphasis on the use of authentic standards at all stages the reviewers assert that the use of simple tests alone to identify drugs is an unsafe procedure; the wide range of mixtures encountered for example in illicit narcotics and the numerous closely related isomers of stimulants make this an inherently unreliable method of identification.Gupta et a1.l’ considered that identification using only colour tests was likely to lead to mis-identification in many instances. Velapoldi and Wicks15 and Johns et aZ.la have published valuable reports on the use of colour tests for analysing drugs of abuse. Both these groups considered that the primary purpose of colour tests was to narrow the list of substances possibly present in an unknown sample. The former group considered the main problems associated with colour tests in general and spot test kits in particular were the interpretation of the colours and the lack of specificity of the tests. Both groups discussed multiple reagent testing schemes using up to seven reagents and numerical codes for possible identification.Velapoldi and Wicks15 emphasised that final identification should always be made by skilled laboratory personnel and that colour tests should never be used as the sole evidence of identifi-cation. The reviewers agree strongly with both these opinions. Considerable data on colour test results are given in the publications discussed above and by Masoud19; Stevens20 has discussed many colour tests and Clarke has presented a comprehensive list of the more important results.21s22 Colour tests for individual groups of compounds (such as narcotics) and individual drugs are discussed in the relevant sections of the review. Simple precipita-tion tests such as addition of silver nitrate or barium chloride (to detect chloride or sulphate ions) may be used to indicate whether a drug is present as a salt or a free base.This may be confirmed by more sophisticated techniques such as IR or nuclear magnetic resonance (NMR) spectrometry. The application of these techniques to salt identification is discussed in more detail under the appropriate drug class. Field Tests Police and Customs investigators frequently need a rapid sorting test for the detection of controlled drugs at the time and place of seizure. Such a test should where negative, minimise the risk of unnecessary detention of a person or goods and where positive justify professional analysis. An account has been given23 of the development of a composite field test kit difficulties encountered with packaging and storage under various climatic conditions and the philosophy of testing.In the reviewers’ experience the introduction of field kits for investigators can dramatically increase the proportion of positively identified drugs of abuse.24 In some instances the demonstration of a successful field test in the presence of a suspect may elicit an admission of guilt. These tests and others requiring greater discrimination or involving simple chromatographic procedures can assist scientists attending the scene of crime or screening substances discovered in clandestine laboratories and generally when working in accommodation without regular analytical equipment. There are numerous schemes wherein selected spot tests are adapted as preliminary colour tests undertaken with minimum equipment remote from laboratory facilities.In this revie 780 BAKER AND PHILLIPS THE FORENSIC Analyst VoL 108 particular examples are quoted under specific classes of drugs. A United Nations working group collected information available up to 1974 on field tests for drugs of abuse commonly found in illicit traffic25; their selection was based on the relative merits of stability specificity, simplicity and safety. A range of colour tests convenient for field use for major classes of controlled drugs as well as the use of portable UV lamps and thin-layer chromatography (TLC) kits were described by Phillips.26 Commercial pre-coated polyester glass and aluminium sheets with a propellent cartridge and chromogen reservoirs provide a better facility for a TLC field kit26 than hand-coated microscope slides2’ Odour tests generally are at best only grounds for suspicion [e.g.acetic acid vapour associated with degraded heroin (or aspirin or vinegar!)]. Even a more distinctive field test such as the transesterification of cocaine to yield the characteristic odour of methyl benzoate (see under Cocaine) is better left to scientific evaluation. For searching vehicles furniture and clothing for traces of drugs a modified vacuum cleaner attachment that collects debris into a Soxhlet thimble has been recommended.28 However while field tests may be useful for screening small amounts caution should be exercised in not consuming a significant part or contaminating the remainder of the visual evidence. For trace amounts of drug material the reviewers’ strongly recommend removing for subsequent examination all debris thus recovered.Many kits are commercially available for field use either by trained analysts or by lay personnel. Although almost all such tests lack specificity the proportion of misleading results in preliminary sorting tests can be markedly reduced by the use of a “logical tree” sequence.29 One such procedure has been patented30s31 and used in a commercially available kit. However the warning15 already given in connection with colour tests intended for laboratory use applies afortiori; field tests taken alone should not be relied upon to be more than presumptive evidence requiring further action and appropriate confirmation. Ultraviolet Spectrometry The use of UV spectrometry as a screening test suffers from similar limitations to colour tests.Rarely is it possible to identify a drug more than tentatively by this technique but useful indications as to the structure of an unknown may be obtained from absorption maxima and minima or from the effects of varying the pH of the solution under examination. It is of very limited value for quantitation except for pure drug substances. Useful discussions on the use of UV spectrometry for tentatively identifying drugs have been p ~ b l i s h e d ~ ~ s ~ ~ and considerable data have been assembled by Clarke.21s22 Major collections of UV data covering a wide range of compounds have been published (for example The Sadtler Catal~gue~~). UV data on specific compounds and groups are discussed in this review.Extinction co-efficients may vary by several orders of magnitude and therefore a small proportion of a strongly absorbing impurity could totally mask the spectrum of a weakly absorbing major component giving rise to erroneous identification and quantitation. Illicitly prepared drugs containing many structurally related impurities with similar or identical spectra may also be wrongly identified and quantified. The technique is useful in confirming the identity of the active ingredient in licit preparations but even with these apparently simple samples it is often necessary to confirm identity by using an alternative technique which may involve chromatographic separation. Infrared Spectrometry In contrast to UV spectrometry which can only rarely be applied to the identification of a particular compound IR spectrometry finds wide application because with few exceptions, every compound produces a different spectrum.Even when samples are very impure it is often possible to identify at least one component from an IR spectrum. The recognition of characteristic spectra is a routine matter for experienced forensic analysts particularly as many laboratories maintain collections of spectra of commonly encountered drug presenta-tions. However when drugs may be present in small amounts relative to other materials present some form of initial clean-up procedure must be employed prior to identification of any drug in the sample. Particular problems are encountered both with tablets whether licit or illicit as large amounts of excipient may be present and in diluted samples of drugs encountered at “street” level.Many different methods of sample clean-up have bee July 1983 ANALYSIS OF DRUGS OF ABUSE 781 described35; simple and rapid clean-up on Celite columns has been commonly ~ s e d ~ ~ p ~ ~ and minor variations may be used to separate for example neutral and basic drugs. Simple acid - base extraction may also achieve similar ends and many procedures have been described by de Faubert Maunder.38 Identification of an unknown drug by IR spectrometry relies on comparison of the spectrum of the unknown with that of an authentic specimen; spectra of many of the more common drugs were illustrated by Clarke,21p22 but the reproductions though clear are very small. De Faubert Maunder38 discussed Clarke's system in detail drawing attention to the problems of polymorphism and sample matrix interactions especially ion exchange.Major collections df IR spectra are also available (e.g. the Sadtler Catalogue34) and many of these are updated annually; these are of particular use when totally unknown samples are submitted. The identity of a purported medicinal substance may be checked if it is one of 358 for which British Pharmacopoeia Infrared Reference Spectra have been p~blished.~S Computer-based systems are also becoming available and these have been listed by Moss et aLgO; these authors also discussed the use of IR spectra for classification of drugs into groups. The reviewers consider that IR spectrometry is a most useful screening technique which can often lead to the identification of an actual drug substance; in their own laboratory they have found that one of the most useful spectra collections is that of drug presentations encountered in the course of routine analysis.For rapid and accurate identification of drugs during screening it is most helpful if both the unknown spectrum and any reference collection spectra are on chart paper of the same size and preferably run on the same instrument, enabling comparison to be made by direct superimposition using a light box. De Faubert Maunder38 gave many useful practical hints on the use of IR spectrometry by forensic drug analysts. Microcrystal Tests Crystal or microcrystal tests find little current application in the forensic analysis of drugs of abuse at least in the UK.Clarkeg1 considered that although useful for supporting a provisional diagnosis being simple rapid and specific such tests were unsuitable as a primary means of identification as they did not lend themselves to forming the basis of a scheme of identification. Der Marderosian and Chao13 recognised that although some tests were very specific others were more general in application,and that there was a tendency for related compounds to give similar crystals with the same reagent. Short discusisions of the tech-nique have been published with useful referen~es.l~,~~ Clarkeg1 has discussed the topic more fully and described tests on many individual drugs. Fultong3 has discussed the whole range of microcrystal tests for drugs in considerable detail. In view of the lack of data on closely related compounds (for example amphetamine isomers) and the lack of a systematic identification system the reviewers do not recommend microcrystal tests as a means of identification of illicit drugs without confirmation by a more specific technique.However microcrystal tests may still have a role to play in drug screen-ing but considerable time and skill is necessary to acquire the relevant expertise.13 Chromatographic Techniques full up to July 1982 by Gough and Baker.44 The application of chromatography in the forensic analysis of drugs has been reviewed in Nuclear Magnetic Resonance Spectrometry Until recently NMR spectrometry has found only limited application in the forensic analysis of drugs. However the increasing popularity of 13C NMR spectrometry particularly in pharmaceutical analysis suggests that this technique may play a significant role in the future.Examples are cited under individual drugs. Mass Spectrometry Lawsong5 considered that the high cost and complexity of mass spectrometry (MS) equip-ment was a considerable drawback to its routine use. Very few laboratories have the resources to use MS as a screening tool and the reviewers believe that the principal use o 5 82 BAKER AND PHILLIPS THE FORENSIC Analyst Vol. 108 the technique a t least in the analysis of drugs above trace levels is in the confirmation of the identity of a substance tentatively identified by some other technique. Klein46 has reviewed the development of MS as a tool in forensic drug analysis. Finkle and co-workers47~48 have published data on both electron impact (EI) and chemical ionisation (CI) MS of drugs: these authors emphasised the importance of matching all data and analytical information prior to the acceptance of a positive identification.Mass spectral data collections which include drugs have been p~blished.~~ The use of MS in drug analysis has been discussed by Scaplehorn .50 The reviewers consider that trace amounts of drugs found for example in motor vehicles or clothing and tentatively identified perhaps by chromatographic techniques should always be confirmed by MS where this provides unequivocal identification. Miscellaneous Techniques Folen51 presented X-ray powder diffraction (XRD) data for some drugs excipients and adulterants in illicit samples and considered that this technique could be applied to the identification of drugs in illicit samples without prior clean-up.The reviewers have not found this technique to be widely applied in routine drug analysis; however in their experi-ence it has proved useful for the identification of particular salts in complex mixtures of illicit narcotics. Bowen et considered the use of circular dichroism as an alternative method of drug analysis. However in the reviewers’ opinion the technique is limited to samples containing a single active ingredient in a pure state; the numerous optically active impurities in illicit preparations of lysergide (LSD) for example are likely to lead to erroneous conclusions. Cannabis Products Introduction In the UK products from plants of the genus Cannabis are listed in separate parts of Schedule 2 of the Misuse of Drugs Act 1971l; cannabinol (CBN) and cannabinol derivatives (i.e.chemical substances) are included with many other drugs in Part I (Class A drugs), whereas cannabis and cannabis resin are listed in Part I1 (Class B drugs). Cannabinol derivatives are defined as “the following substances except where contained in cannabis or cannabis resin namely tetrahydro derivatives of cannabinol and 3-alkyl homologues of cannabinol or of its tetrahydro derivatives.” Thus are controlled tetrahydrocannabinol (THC) and synthetic variants thereof in which a larger 3-alkyl group is included (for example, “synhexyl”). Cannabis was redefined in the Criminal Law Act 197753 as “(except in the expression ‘cannabis resin’) any plant of the genus Cannabis or any part of any such plant (by whatever name designated) except that it does not include cannabis resin or any of the following products after separation from the rest of the plant namely-(a) mature stalk of any such plant (b) fibre produced from mature stalk of any such plant and (c) seed of any such plant.” Cannabis resin is explicitly definedl as “the separated resin whether crude or purified obtained from any plant of the genus Cannabis.” Thus in the UK no question need arise as to the species of Cannabis a problem not infrequently encountered by analysts in the USA.54-56 Recent case law (R.vs. Best and Others5’) has determined on appeal that alternative charging of cannabis and cannabis resin does not constitute “duplicity” for a single act of possession but it is still important to differentiate between these substances if there is more than one exhibit.Phillips58 has discussed difficulties arising under previous legislation including traces identified in smoking residues or clothing. Liquid cannabis (hash oil) is no longer explicitly controlled by UK legislation and although Extract and Tincture of Cannabis were both named specifically in succeeding Dangerous Drugs Acts,68 they were not specified as such in the Act of 1971.l Consequently it falls to the analyst to demonstrate the precise legal status of this substance in order that correct charges may be laid. Forensic analysis of products from plants of the genus Cannabis may be subdivided into four (overlapping) operations screening for the presence of a Cannabis product in the sample, identification of the product quantitation of that product and rarely precise quantitation of a specific cannabinoid in the sample.The products of the Cannabis plant differ con-siderably in their macroscopic appearance from simple plant material through resins t July 1983 ANALYSIS OF DRUGS OF ABUSE 783 extract~.~9s~~ In view of the number of samples submitted to forensic science laboratories, and their varied presentations identification procedures for Cannabis products have been very widely studied and in general are simple and rapid techniques.61 Procedures fall into three groups microscopic and macroscopic appearance colorimetric (field) tests and chro-matographic methods primarily TLC.Microscopy The microscopic features of Cannabis have been fully described62; where these are present in a sample unequivocal identification of the presence of cannabis may be made by a trained micros~opist,~~ but where distinctive morphology is missing or damaged other supporting techniques must be employed.64 In the reviewers’ opinion this limits the general application of microscopy to plant material alone because in the majority of cannabis resin samples many characteristic features have been destroyed65 and in liquids are absent. Residues or traces such as in smoking utensils or clothing are also rarely amenable to simple micro-scopic examination owing either to combustion or gross contamination by other materials. Numerous authors have discussed the microscopic appearance of Cannabis and the criteria necessary for unequivocal identification by this t e ~ h n i q u e .~ ~ - ~ ~ Although some authors in the forensic science field describe identification of Cannabis products by microscopy the majority of reports rely on a combination of tests that may include microscopy. Mitosinka et aZ.73 examined Cannabis and other plant materials using the scanning electron microscope and the use of this instrument in the preparation of evidence for presentation in Courts-of-Law has been discussed.74 Gross Morphology The gross morphological appearance of different illicit Cannabis products has been d e s ~ r i b e d ~ ~ s ~ ~ ; it is clear from both of the publications cited that there is a very wide variation in the appearance of Cannabis products from different geographical origins.Colour (Field) Tests Prior to the elucidation of the structures of the major cannabinoids and the development of chromatography chemical identification of Cannabis products was made by simple colour tests. Of the many tests for presumptive identification those which have found widest discussion in the literature are those based on an original test described by Duquenois and M u ~ t a p h a . ~ ~ ~ ~ ~ Generally described as the Duquenois or Duquenois - Negm test there is considerable confusion in the literature over the many variations of the original test and it is rarely clear which test has been used. However the modification that is most widely used is that generally referred to as the Duquenois - Levine test.77 Thornton and N a k a m ~ r a ~ ~ and Pitt et aZ.7s have discussed the mechanism of this test in detail in the light of more recent discoveries of the structures of the major cannabinoids.The latter authors in a discussion of the specificity of the Duquenois - Levine test concluded that if the criteria for a positive test were rigorously adhered to and if botanical evidence was also available then this test was a reliable screen for cannabinoids. However (they say) if botanical evidence is not available then the test must be supplemented with chromatographic evidence. This con-clusion is substantiated by a report that certain brands of coffee give a positive Duquenois -Levine test.79 Baileyso discussed the value of the Duquenois test in great detail and found that the Duquenois - Levine test77 was the most specific of all the modifications of the original test.The Beam tests1 and a modifications2 have also been widely used but although reported to be more specific these were found less sensitives3 than the Duquenois - Levine test and have also been shown to be ~nreliable.8~98~ It has been suggested that this reagent reacts with cannabidiol (CBD) and cannabigerol or their acidss5 but not with THC.86 Consequently, it is not surprising that the test has proved unreliable in view of the absence of CBD from some samples of The rapid field test88 (and the improved procedure89) described by de Faubert Maunder as a reliable indicator of the absence of Cannabis products within samples has been used in the Laboratory of the Government Chemist for over 14 years and has been incorporated in a commercially available Drug Test Kit.9o A positive test is presumptive evidence for the This test does not appear to be in wide current use 784 BAKER AND PHILLIPS THE FORENSIC Analyst Vol.108 presence of a Cannabis product within the sample; only nutmeg and mace are reported to give false positive reactions.88 Many other colour tests have been described; these include the Ghamravy test,91 the furfural testg2vg3 and a number of other^^^-^^; these tests do not appear to have found wide-spread usage. There is little evidence that many laboratories are identifying Cannabis products on the basis of chemical or colour testing alone; Mechoulam et al. in a major review ( 1976),97 considered that “by their nature colour tests are non-specific and should be confined to field work and screening while other techniques should be used for identification.” The reviewers concur with this opinion and emphasise that such screening tests should be limited to bulk seizures and should not be applied to trace amounts or smoking residues where evidence may be destroyed by such testing.Further it is also the reviewers’ opinion that criminal prosecutions relating to Cannabis products where the only evidence is a positive field test are inherently unsafe and should not be pursued. There seems to be little to choose between the use of the Duquenois - Levine test7’ for the presumptive identification of Cannabis products and the rapid field test for their presumptive absence described by de Faubert Maundersg; however in the reviewers’ experience the latter is more convenient to use in the field.The reviewers consider that the identification of cannabis and cannabis resin can be achieved unequivocally in most instances by combining one of the above two colour tests, correctly carried out with a careful consideration of the morphology of the sample. Where both macro- and microscopic features are absent or damaged as in some cannabis resins or extracts and in trace amounts or smoking residues colour tests alone cannot be relied upon and the identification of the presence of a product from Cannabis must be made by a second technique. Most commonly used is TLC and the many systems in use have been discussed by Gough and Baker.44 Mechoulam et al.,97 although of the opinion that gas - liquid chromatography (GLC) has superseded TLC for many routine analyses felt that the use of TLC would persist where requirements are not stringent and where speed and convenience are of greater importance.In an important paper,98 Hughes and Warner noted that no mixture of components previously reported had the same TLC characteristics as Cannabis products. The possible confusion between nutmeg or mace and cannabis that might arise using the rapid field testsg may be resolved using TLCeg9 In 1973 MechoulamlOO reported that cannabinoids had not been isolated from any plant or animal except Cannabis and the reviewers are not aware of any more recent reports which indicate otherwise. Fenselau et a.Z.lo1 examined hop extracts (genus Humulus) by MS and found that cannabinoids were not present (detection level less than 10 ng g-l).Hz~mulus and Cannabis are the only genera in the family Cannabinaceae. In a valuable study Coutts and JonesS1 suggested that only if the “three-parameter approach” (morphology colour tests and TLC) could be shown to be in error should further tests be considered a necessary part of the identification of Cannabis products. These authors re-examined 100 seizures using GLC combined with MS (GLC - XIS). Agreement was found with all but two analyses; these two errors arose from the Duquenois - Levine test77 giving erroneous negative results on smoking residues. TLC indicated the presence of THC and CBN. These authors concluded that TLC identification of THC and one other cannabinoid in an extract was sufficient to show that the source of the extract was a Cannabis product and that the “three-parameter approach” was unequivocal in identifying Cannabis products.It is the reviewers’ opinion that authentic standards of cannabinoids whether pure or in a standard tincture should always be used when TLC is applied to the identification of cannabinoids. Some authors have suggested that GLC should be used for the identification of a Cannabis product102-104 but in the reviewers’ opinion TLC is sufficient. Macro- and microscopic sorting may be helpful in determining the proportion of the Cawabis product in the sample. Normally all that a Court requires in the United Kingdom is the mass of Cannabis product. With trace amounts and smoking residues it may be necessary to take into account the detection limit of the analytical technique used.Gough and Baker44 have discussed TLC detection limits of several chromogens used for cannabinoids. It is always necessary to identify the actual drug present,57 whether it is (or was) cannabis or cannabis resin.58 Samples of cannabis resin have always been found to contain CBD,60 as do some samples of cannabis from the “resin belt,”59 whereas many samples of cannabis are devoid of CBD.59*60~s7 Conse-quently if TLC indicates the absence of CBD in a sample where no morphology is apparent, the analyst can decide with a considerable degree of certainty that the cannabinoids presen July 1983 ANALYSIS OF DRUGS OF ABUSE 785 derive from cannabis rather than from cannabis resin.The converse is of course not so. Similar considerations apply to samples of liquid cannabis; those devoid of CBD may be considered to have been made by extraction of cannabis whereas those containing CBD could have been prepared from either cannabis resin or from Cannabis grown in a resin-producing area. Although Cannabis is grown in many parts of the world there are only a small number of countries from which illicit Cannabis products reach the UK. It has been considered104@5 that a means of identifying the geographical origin of illicit Cannabis products would be extremely useful in criminal investigation and in international control. Baker et aE.eO have summarised available data on cannabinoid content in relation to country of origin and have studied the physical and chemical features of Cannabis products illicitly imported into the UK from known geographical origins.On the basis of their study of the many thousands of specimens of Cannabis products from all over the world maintained as a reference collection at the Laboratory of the Government Chemist these workers60 consider that a combination of careful visual inspection of a sample of unknown provenance and comparative TLC enables an analyst to offer an opinion as to its geographical origin. In order to validate further such an opinion these same workers made studies of the physical and chromatographic properties of Cannabis plants grown in the UK from seeds of known origin.106s107 The gross physical appearance and cannabinoid patterns of many of the UK-produced specimens were in general, closely related to those of their parents but some exceptions were recorded.In comparison with imported material,lo8 cannabis produced in the UK had higher tetrahydrocannabinolic acid (THCA) to THC ratios than imported material. In the first study,lo6 the authors also reviewed previously published data on Cannabis plants grown under controlled conditions (although not necessarily from seeds of known provenance). It is often necessary to compare two samples of a Cannabis product in order to establish or refute a common origin. Methods for comparison are primarily chromatographic and have been reviewed by Gough and Baker.44 It is rarely necessary to quantify a particular cannabinoid for forensic purposes although it may be necessary to measure levels in particular samples for example the THC content of Thai cannabislog or that of oils or extracts when so required by Courts-of-Law.Deter-mination of THC content is normally made by GLC.44 Such determinations give the total THC content i.e. THC + THCA the latter being decarboxylated on injection into the chroma tograph. Opium and Poppy Straw In the UK the Misuse of Drugs Act 1971l controls three natural products of the opium poppy Papaver somniferum under Part I of Schedule 2 opium (whether raw prepared or medicinal) poppy straw and concentrate of poppy straw. Opium otherwise known as raw opium is the latex obtained by incision of the unripe seed capsule of Papaver somniferum L., dried or partly dried by heat or spontaneous evaporation.l1° Part IV of Schedule 2 of the Act includes powdered or granulated opium within this definition.Prepared opium is defined in Paragraph 37 of the Act as “opium prepared for smoking and includes dross or any other residues remaining after opium has been smoked.” Medicinal opium is defined in Part IV of Schedule 2 as “raw opium which has undergone the process necessary to adapt it for medicinal use in accordance with the requirements of the British Pharmacopoeia, whether it is in the form of powder or is granulated or is in any other form and whether it is or is not mixed with neutral substances.” Certain low dosages of medicinal opium are excluded from control under the Act by the Misuse of Drugs Regulations 1973.ll1 Poppy straw and concentrate of poppy straw are defined in Part IV of Schedule 2 as “all parts, except the seeds of the opium poppy after mowing” and “the material produced when poppy straw has entered into a process for the concentration of its alkaloids,” respectively.It is of interest that whereas cultivation of Cannabis is specifically prohibited by the Act,l cultivation of Papaver somniferum is not so controlled. In contrast opium smoking equip-ment intended for use is controlled even if unused whereas no such control is exercised over unused equipment specifically designed for the smoking of Cannabis products 786 BAKER AND PHILLIPS THE FORENSIC Analyst VoE. 108 Opium Opium contains 25-30 alkaloids,l12 of which morphine averaging 10% by mass in Indian opium,ll3 codeine thebaine noscapine papaverine and narceine are the most important .I12 In a detailed and valuable paper de Faubert Maunder114 described field and laboratory tests for raw and prepared opium; a comprehensive account was given of the reactions of opium to common colour tests and the author presented a general analytical scheme for these materials.Lim and Kwok115 discussed opium usage in Singapore and based part of their analytical procedure on de Faubert Maunder's scheme.ll* Unless samples were very small or in a dry powdered form they found little difficulty in successfully identifying different types of opium. Final confirmation of the identity of an opium sample may be made by TLC1l4-ll6 using authentic opium as a standard. Smithll' recommended GC - MS for the forensic identification of opium but the reviewers consider that this is unnecessary unless only very small samples are available for analysis.Antipyrine a synthetic drug has been reported in samples of Iranian opium118 and mixtures of opium with cannabis resin have been seized,'O but no other mixtures of illicit opium are known to the reviewers. The Misuse of Drugs Act 1971l does not specify any particular alkaloid levels in opium and this review does not therefore cover analytical techniques for such determinations as they are not forensically necessary. Poppy Straw Samples may be identified by the presence of opium within the seed capsules using appropriate techniques. Although Fairbairnllg considered morphine and codeine to be of very limited occurrence in other species of Papaver Bentley120 reported the occurrence of morphine in Fructus papaveris (blue poppy) The reviewers therefore consider that in addition to chemical evidence for the presence of opium alkaloids botanical evidence should always be obtained prior to final identification of poppy straw.Concentrate of poppy straw may be identified from its alkaloidal constituents and any botanical debris present. The reviewers consider the borderline between concentrate of poppy straw and crude or impure morphine to be ill-defined. Concentrate of poppy straw is not clearly defined in the Explanatory Notes to the Customs' Co-operation Council although it is listed for tariff purposes.121 Such cases should they arise may have to rely on the expert opinion of the analyst as to the appropriate charge in Law.In such cases the reviewers consider that a morphine content determination is imperative in order that the Courts may gauge the seriousness of any charge. Narcotics Introduction Part 1 of Schedule 2 of the Misuse of Drugs Act 1971l consists mainly of a list of narcotic analgesics. Most of these drugs have been synthesised for experimental therapeutic purposes and few have any current use. Only a small number are misused to any considerable extent. The major misused narcotics are diamorphine (heroin ; 3,6-diacetylmorphine) and morphine. A small number including codeine (controlled by Part I1 of Schedule 2)) dipipanone pethidine and methadone are occasionally encountered and the rest scarcely at all. Stereoisomers, esters ethers salts and preparations or products containing these substances are also con-trolled.Certain preparations containing low levels of morphine and codeine are excluded from control under the Misuse of Drugs Act Regulations 1973,ll1 but all their preparations designed for administration by injection are controlled (codeine preparations designed for injection are controlled under Part 1 of the Schedule). In view of the predominance of heroin and morphine in the field of abused narcotics the reviewers do not propose to discuss in any detail other narcotics but mention will be made of them where relevant. Although small amounts of licitly manufactured preparations containing narcotics are occasionally seized most narcotics analysed in forensic science laboratories are illicitly manu-factured. Large seizures have been made of licitly manufactured morphine tablets (15 or 30 mg of active ingredient) after illegal importation from the Indian sub-continent but this is the sole major exception to this generalisation July 1983 ANALYSIS OF DRUGS OF ABUSE 787 No licit preparations of heroin are known to the reviewers to be widely available or mis-used although small amounts reach “street level,” mainly resulting from thefts from pharmacists’ premises or hospitals.Such preparations may be tentatively identified from codes or manufacturers’ markings but such occurrences are rare. Licit preparations of some other controlled narcotics are encountered at street level and therefore guides and charts may find application to their tentative identification. Illicit preparations vary from essentially pure heroin hydrochloride (intended for injection) to crude and impure materials often containing heroin base and probably intended for smoking.Johnson and Gunn122 discussed in detail the diluents and adulterants in “street level” heroin seizures and Baker and G o ~ g h l ~ ~ listed many of the compounds found in heroin at importation. Heroin is manu-factured by acetylation of morphine and the major impurity is 6-acetylmorphine as well as acetylcodeine and other opium alkaloids. Major adulterants with physiological effects include caffeine and strychnine (in products from South-East Asia) and procaine (from South-West Asia). The appearence of illicit heroin varies from pure white crystalline samples of heroin hydrochloride through amorphous beige and brown powders to dark brown lumpy material.It is the reviewers’ experience that there is very little correlation between colour and heroin (as hydrochloride or base) content. Clark and Miller124 studied dyes added to illicit heroin samples and found that their use was widespread. The appearance of Chinese No. 3 heroin which contains caffeine is characteristically granular and beige to pale brown in colour often with a distinct odour of acetic acid. Some samples of Middle-Eastern heroin have an odour of opium and may be prepared by acetylation of opium extracts rather than from crude morphine. Two useful studies of the specificity of analytical techniques for the identification of heroin have been p r e ~ e n t e d l ~ ~ J ~ ~ and these will be discussed in detail below.Colour and Microcrystal Tests Until the advent of routine and relatively cheap analytical instrumentation identification of heroin was based on colour and microcrystal tests.127 The Marquis reagent128 (formaldehyde and concentrated sulphuric acid) gives a characteristic purple colour with heroin morphine and structurally related compoundsls and is widely used as a field test. L e r n e ~ - l ~ ~ found the test to be sensitive to as little as 1 pg of pure heroin. He used concentrated nitric acid as a second colour test for heroin the colour change from yellow to pale green being in his opinion, specific for heroin but considerably less sensitive. However Manura et aZ.126 showed in a study of the colour reactions of heroin and 56 structurally related compounds that neither the Marquis nor the nitric acid test,129 nor the two tests in combinaticn were in any way specific for heroin.Clark125 showed that 17 compounds structurally related to heroin gave similar colours with the Marquis128 and Mecke reagents.130 Engelke and Vincent131 found some improvement in the specificity of colour reagents if colour reference charts were used, but data were only presented on pure alkaloids. Clarke21y22 listed the colours given by the principal narcotics with the Marquis reagent128 and some other common colour tests. The reviewers consider that although the Marquis test128 is non-specific it is still useful for screening at point-of-seizure particularly if part of a “logical tree” s ~ h e m e ~ ~ ~ ~ ~ and it is currently included in many commercial field test kits.Tetracycline a widely used anti-biotic may be confused with illicit narcotics as it gives an instant purple with this reagent, but the colour changes rapidly to yellow - brown which distinguishes if from the narcotics, where the purple colour is stable for many minutes. In addition tetracycline preparations are yellow an unusual colour for illicit narcotics. Fult0nl3~ described in detail several microcrystal tests for heroin and related compounds, but only pure drugs were used. Lerner and Mills133 found problems in microcrystal formation with illicit heroin samples which they attributed to the presence of 6-acetylmorphine. Clark125 found no correlation between molecular structure and crystal shape in a study of heroin and 17 related compounds and in addition found difficulty in describing crystal shapes.Manura et aZ.,126 in a study of heroin and 56 related compounds found that microcrystal descriptions were subjective and encountered considerable problems with impure samples. Clarke21@ has described common microcrystal tests for the principal narcotics. The reviewers consider that microcrystal tests have little value in modern forensic analysis of narcotics. Important diluents include sugars and quinine 788 BAKER AND PHILLIPS THE FORENSIC Analyst Vol. 108 Nuclear Magnetic Resonance Spectrometry NMR spectrometry is little used in the forensic analysis of narcotics.126 Clark,125 in a study of heroin and structurally related compounds found that although each of their NMR spectra were different fairly pure samples were needed for unambiguous identification.Ultraviolet Spectrometry UV spectrometry has been found to be far from specific in the analysis of illicit nar-c o t i c ~ . ~ ~ ~ * ~ ~ ~ Although the technique may be useful for the screening of licitly manufactured materials the reviewers consider that the use of this technique is very limited either for identification or for quantitation of illicitly manufactured material. UV spectrometric absorption at selected wavelengths is however very widely and satisfactorily used as a detection system in the analysis of narcotics after their separation by high-performance liquid chromatography (HPLC) .44 Infrared Spectrometry Curry and Patterson134 used IR spectrometry as the first technique in a procedure for the analysis of illicit heroin samples.In a study of the IR spectra of many samples all were found to have at least six major absorption maxima in their IR spectra. Caffeine was also identified in many samples. These authors considered that the presence of heroin in illicit samples could be virtually confirmed on the basis of their IR spectra. They also consider that although heroin might appear to be absent from suspect samples this absence should be confirmed by chromatography. Clark,125 in a study of heroin and structurally related com-pounds and Manura et a1.,126 in a larger study found that all the individual compounds and their salts were each distinguishable by their IR spectra. The latter authors considered IR spectrometry to be the most specific technique for the identification of heroin.Shaler and J e r ~ e l ~ ~ studied illicit heroin samples by GLC coupled with IR spectrometry and considered this a useful and very specific technique for the unambiguous identification of heroin. In the reviewers’ experience IR spectra identify the presence of heroin in illicit heroin samples with little difficulty and it is their opinion that as an identification technique it is of great value. The spectra of some samples of illicit heroin containing appreciable levels of 6-acetyl-morphine and/or morphine are however difficult to interpret. Morphine codeine and other narcotics may also be identified from their IR spectra.21,22 Chromatographic Techniques The identification and quantitation of heroin may be achieved by the principal chromato-graphic techniques widely used in forensic science laboratories.126 Most common illicit narcotics may be analysed by GLC or HPLC often after screening of the sample by TLC.Chromatographic techniques for the analysis of major narcotics have been reviewed by Gough and Baker.44 The reviewers consider that identification should always be based on two different quantitative chromatographic techniques if IR spectrometry is unavailable or an ambiguous spectrum is obtained. Mass Spectrometry Saferstein et al.136 presented CI MS data on morphine codeine heroin the two acetyl-morphines and acetylcodeine. Nakamura et al.137 used GC - MS to identify heroin in “street” samples at the 5-20?/ level. Herman and Kan138 found P-chloromorphide in a sample of partially acetylated morphine and presented mass spectral data to support this identification.Clark125 studied GC - MS data from heroin and related compounds and found that each was unambiguously identifiable. This author considered that this was a particularly useful identification technique as preliminary clean-up was not required. Jerpe et al.139 used single ion monitoring as a quantitative technique for the identification and assay of heroin and found it valuable particularly in the quantitation of heroin in small seizures. MS has been little used in the identification of heroin and other major narcotics in illicit seizures as identification can usually be made by IR spectrometry. However we believe its use to confirm the presence of narcotics in trace amounts particularly from syringes and from scenes-of-crime to be mandatory.In addition it is valuable in identifying minor constituents of larger seizures particularly when combined with prior GLC analysis July 1983 ANALYSIS OF DRUGS OF ABUSE Coca Leaf 789 Introduction In the UK coca leaf is controlled under Part I of Schedule 2 of the Misuse of Drugs Act 1971l and is defined in Part IV of the same Schedule to that Act as “the leaf of any plant of the genus Erythroxylon from whose leaves cocaine can be extracted either directly or by chemical transformation.” Archer and HawkeslgO described other alkaloids structurally related to cocaine that occur in the leaves of plants of the genus Erythroxylon. Aynilian et ~ 1 . ~ ~ ~ briefly discussed the distribution of cocaine within the different species of that genus.Seizures of coca leaf in the UK are rare and usually small. Most of the Coca species are native to South America mainly from Peru Bolivia and C01umbia.l~~ Identification The dry uncurled coca leaf can be recognised by its tea-like odour and by two longitudinal indentations on either side of the midrib. These are more conspicuous on the grey - green underside than on the dark green upper side.lg3 Powdered coca leaf may be identified by microscopic examination.lg4 Because the control of coca leaf under the Misuse of Drugs Act 1971l places emphasis on the extractability of cocaine from the leaf the reviewers con-sider that identification of this material in the forensic laboratory should always be confirmed by a chemical analysis which demonstrates the presence of cocaine in the leaf material presented for identification.After extraction of the alkaloids from the leaf,141s145 detection of cocaine together with other related alkaloids is usually made by chromatographic tech-niques. This aspect of coca leaf analysis has been reviewed by Gough and Baker.44 Cocaine Introduction Cocaine has been used throughout recorded history for its local anaesthetic activity.lgB However the stimulant and euphoric properties of cocaine induce a high level of psycho-logical dependence and hence its widespread illicit use.147 A detailed discussion of the historical aspects of cocaine use and abuse has been presented by Aldrich and Barker.lg6 As a result of the potential for abuse control of cocaine is now almost worldwide.In the UK cocaine together with its stereoisomers esters ethers salts and any preparation or product containing cocaine or any of these derivatives are controlled under Part I of Schedule 2 of the Misuse of Drugs Act 1971.l Certain preparations of cocaine containing very small dosages are excluded from control by the Misuse of Drugs Regulations 1973.ll1 The same Part of the Schedule also controls ecgonine and any derivative of ecgonine which is con-vertible to ecgonine or cocaine. This latter group includes acyl esters of the 2-hydroxyl (e.g. benzoylecgonine) and alkyl esters of the 3-carboxyl such as methyl ecgonine; both series can be converted into cocaine by an esterification step. A third important derivative of ecgonine is cinnamoylcocaine (that is cinnamoylecgonine methyl ester) a compound naturally occurring in coca leaf which can be converted into ecgonine or cocaine using simple tech-niques.The significance of the occurrence of this compound in seizures of cocaine has been discussed by Moore.148 The physicall49 and chemical140 properties of cocaine and its principal derivatives have been described. Seizures of cocaine made in the UK by Officers of Her Majesty’s Customs and Excise originate principally from Bolivia Columbia and Peru.150 Such seizures vary from being essentially pure to highly adulterated often with non-controlled topical anaesthetics. It is not uncommon in the reviewers’ laboratory to analyse a sample that consists wholly of one or more of the latter compounds.The adulteration of cocaine samples has been discussed by Baker and G 0 ~ g h . l ~ ~ Colour Tests The majority of colour tests for cocaine depend on the production of a blue colour when cocaine reacts with cobalt (11) thiocyanate in acidic media.151 Alliston et aZ.152 examined several cobalt thiocyanate-based reagents as part of a preliminary sorting sequence29 prior to full laboratory analysis. Although the specificity of any single reagent was not good mos 790 BAKER AND PHILLIPS THE FORENSIC Analyst Vol. 108 reagents also reacting in a positive fashion to methadone methaqualone pethidine and some local anaesthetics these workers achieved some measure of discrimination by disregarding responses not apparent in less than 5 s. When used as an integral part of a logical test kit29g90 and in particular in conjunction with a test reagent based on 4-dimethylamino-benzaldehyde (DMAB) ,153J54 some improvement in specificity was obtained.This sequence of testing was used by Baker and Gough150 in preliminary tests on mixtures of cocaine with local anaesthetics. These authors warn that a positive response to these tests is by no means specific and reiterate that field tests are only for preliminary sorting. S ~ o t t 1 ~ ~ developed a cobalt thiocyanate-based reagent that was described as sensitive specific and difficult to misinterpret. Only tropacocaine a local anaesthetic rarely encountered in seizures gave a false positive reaction to this reagent. Phencyclidine and a number of local anaesthetics gave negative reactions. Winek and E a ~ t l y l ~ ~ evaluated the test and considered it useful for pure materials suspected of being cocaine but found false positive reactions with some drug mixtures that did not contain cocaine.These authors recommended that con-firmation of drug identity should always be sought by an alternative (chromatographic) technique. Grant et aZ.15' considered specific cobalt thiocyanate-based test reagents were unlikely to be developed and suggested that the characteristic odour of methyl benzoate, generated when cocaine is transesterified by heating with methanolic alkali was both more sensitive and more specific than the more commonly used field test reagents. Most drugs were found to give no odour under the conditions of this test and further the smell was persistent and could be distinguished from for example methyl acetate arising from the testing of aspirin (acetylsalicyclic acid).Bastos and Hoffman,42 in a brief review of field testing emphasised that all field tests should be confirmed in the laboratory a point which the reviewers have emphasised. Microcrystal Tests Bastos and Hoffmang2 considered that microcrystal tests were the best physico-chemical techniques for the identification and confirmation of small amounts of cocaine. These authors have briefly reviewed this to pi^.^^^^^^ They recommend gold chloride in phosphoric acid as particularly useful because local anaesthetics are poorly precipitated by this medium. F ~ l t o n l ~ ~ suggested that gold bromide in a mixture of acetic and sulphuric acids was also useful when local anaesthetics were likely to be present.Valanju et aZ.l59 described micro-crystal tests for ecgonine and benzoylecgonine. The reviewers consider that microcrystal tests are of little value in the identification of cocaine. Ultraviolet Spectrometry Bastos and H ~ f f m a n ~ ~ ~ ~ ~ ~ in reviewing UV and colorimetric methods for the analysis of cocaine and related compounds concluded that although cocaine benzoylecgonine and ecgonine could be detected and .quantified by UV spectrometry,l60 this method was neither specific nor sufficiently sensitive for use in the analysis of drugs of abuse. Moorelg8 observed that cinnamoylcocaine could be detected in cocaine seizures by UV spectrometry and that the presence or absence of this compound might be a useful pointer to the method of manu-facture of the cocaine.This author emphasised that the presence of cinnamoylcocaine should be confirmed. Infrared Spectrometry O'Brien and Sullivanl6l found that IR spectrometry could distinguish between cocaine and 14 local anaesthetics. Bastos and Hoffman149 considered that the technique was very useful in the identification of cocaine. Moore148 established the presence of cinnamoylcocaine in illicit cocaine seizures using IR spectrometry. Trinler and Reulandls2 considered that IR spectrometry was the best method of unequivocally identifying cocaine in mixtures in view of the high cost of routine MS. These authors used a preliminary HPLC separation of cocaine from local anaesthetics prior to identification by IR spectrometry.Chromatography The forensic analyst not only must unequivocally identify the drug (or drugs) present in July 1983 ANALYSIS OF DRUGS OF ABUSE 791 sample but also the amount of that drug must be quantified accurately. The most common technique used is GLC. Chromatographic methods for the analysis of cocaine and related compounds have been reviewed by Gough and Baker.44 Mass Spectrometry Mass spectrometric data for cocaine and some derivatives mainly after GLC have been p r e ~ e n t e d . ~ ~ ~ s ~ ~ ~ s ~ ~ ~ - ~ ~ ~ The reviewers do not consider that routine MS of cocaine in bulk seizures is necessary to confirm identity as chromatographic and IR spectrometric techniques provide sufficient evidence. However when sufficient sample is not available for structural confirmation by the usual methods or when the sample is highly contaminated such as in the examination of trace amounts from clothing or vehicles confirmation of identify by MS is, in the reviewers’ opinion mandatory.In a valuable paper Clark166 discussed in detail the identification and quantitation (by addition of deuterium-labelled cocaine) of cocaine by MS. No difference in accuracy (at the 95% confidence level) was observed in quantitative results from this method as compared with gas chromatography.167 Optical Isomers and Stereoisomers It is probable that all currently available illicit cocaine is prepared from coca leaves and therefore consists of the natural isomer (-)-cocaine.l68 However as the occurrence of synthetic (&)-cocaine or (+)-cocaine is possible it may be necessary for the forensic analyst to distinguish between the optical isomers and the racemic mixture.In the USA Federal and most state laws subsume cocaine within the following definition “coca leaves any salt, compound derivative or preparation of coca leaves and any salt compound derivative or preparation thereof which is chemically equivalent or identical with any of these substances,” wording that Siege1 and CormieP9 consider imprecise. Indeed some authorities assert that, in the USA only (-)-cocaine is controlled the other optical isomers of cocaine and its stereoisomers pseudococaine allococaine and pseudoallococaine being not controlled. Useful discussions of the legal position in the USA have been presented in several p a p e r ~ . l ~ ~ - l ~ ~ In the UK all stereoisomeric forms of cocaine are subsumed by Paragraph 2 of Part I of Schedule 2 of the Misuse of Drugs Act 1971l; thus the question as to whether a substance identified as “cocaine” may not be a controlled drug cannot arise under UK legislation.However it may be of forensic importance for the analyst to determine which isomer (or isomers) is present in a sample. Such information may be useful in determining the method of manufacture of the sample. Chromatographic techniques for the separation of optical and stereoisomers have been reviewed by Gough and Baker.44 Si.egel and CormieP9 described the preparation of (+) -pseudococaine from (-)-cocaine ; the former was charac-terised by commonly used chromatographic and physical techniques.The authors concluded that the use of these (routine) tests would confirm or refute pseudococaine as a substance purporting to be cocaine. Allen et aZ.,l72 in an important paper described a logical sequence for the identification of the stereoisomer present (by IR NMR and MS) and then determina-tion of chirality by crystal tests IR spectrometry melting-points or optical rotation. KrolP70 determined the enantiomorphic composition of cocaine using the chiral lanthanide shift reagent europium tris-d-trifluoroacetyl camphorate ; 2 mg of cocaine sample could be analysed (less if Fourier transform NMR spectrometry was used) with an isomer detection limit of 5%. Other Forensic Aspects In illicit seizures of cocaine the drug is more commonly present as the hydrochloride than as other salts or the free base.Anions may usually be confirmed with common precipitation reagents but greater certainty is possible using IR spectrometryl6l or X-ray diffra~ti0n.l~~ Moore14s has suggested that cocaine samples may be compared on the basis of impurities and gave spectral data on cinnamoylcocaine. Lukaszewski and Jeff ery1G4 suggested that NMR spectroscopy could be used for the comparison of cocaine samples by studying the presence and amount of cinnamoylcocaine. These authors further suggested that the presence of this compound in a cocaine sample indicated that the material was derived from Coca rather than prepared synthetically. The converse is however not true as the sample could have been prepared by hydrolysis of all ecgonine esters to ecgonine and re-esterification 792 BAKER AND PHILLIPS THE FORENSIC Analyst Vol.108 Samples that do not contain cinnamoylcocaine cannot therefore be unequivocally identified as synthetic. LSD and Indolic Hallucinogens Introduction In the UK the Misuse of Drugs Act 1971,l as originally promulgated controlled in Part I of Schedule 2 only the limited number of hallucinogens specified in previous legislation. These include bufotenine lysergamide (lysergic acid amide) lysergide (lysergic acid diethyl-amide LSD) and other N-alkyl derivatives of lysergamide psilocin NN-dimethyltryptamine (DMT) and NN-diethyltryptamine (DET). In addition stereoisomers esters ethers salts and preparations or other products containing these substances are controlled. Psilocybin, the phosphate ester of psilocin therefore falls into this category.A subsequent Modification Order174 added a further group of compounds derived from tryptamine to Part I of Schedule 2 (where not controlled under the 1971 Act1) describing them generically as “any compound structurally derived from tryptamine or from a ring-hydroxy tryptamine by substitution at the nitrogen atom of the side-chain with one or more alkyl substituents but no other substituent .” but many hallucinogens are substances naturally occurring in plants (and for bufotenine also in a species of toad). For many centuries people in whose area these plants grew exploited their hallucinogenic nature ; several discussions and reviews of the occurrence of hallucinogenic plants have been pre~ented.l7~-l’~ The history of the use of these materials and their occurrence has been described in detail by Emboden.lSo The occurrence of hallucinogens in Psilocybe species,lal Argyreia nemosa (Hawaiian Baby Wood Rose) ,lS2 Virola per~viana,l8~ Rivea corymbosa and Ipomoea violacea (Morning Glory)la4 and in ergot fungusls5 have been described.Whereas cannabis coca leaf and poppy straw are controlled by the Misuse of Drugs Act 1971 ,l no plants containing controlled hallucinogens are explicitly named. Indeed it is implicit in the House of Lords’ ruling in the case of DPP us G o o d ~ h i l d ~ ~ ~ ~ ~ ~ ~ that drugs while still contained within a plant (or animal) (i.e. in their natural state) do not fall within the scope of the Act. Despite lack of control it may be necessary to identify such materials in the UK for example in relation to military prosecutions and in cases of poisoning.Despite the occurrence of numerous hallucinogenic plants from which a large number of chemicals could be extracted the chief source of hallucinogens for misuse is illicit labora-tories.ls8 Although there have been occasional reports principally from the USA of misuse of DMT and DET and an upsurge of interest in Psilocybe mushrooms in some ~ o u n t r i e s ~ ~ ~ - - l ~ LSD is the most commonly abused hall~cinogenl9~J93 and almost all analytical data on hallucinogens concern this compound. LSD is active in man at doses as low as 1 pg kg-l body mass.lg4 Illicit dosage units contain very variable amounts of LSDlg5 and impurities and excipients are often present in an excess.l88 Precursors of LSD are present in many plants, one of the most important being lysergamide in Morning Glory seeds.ls4 Brief reviews of identification methods for hallucinogens have been p ~ b l i s h e d .~ ~ ~ ~ ~ ~ 6 - ~ ~ ~ LSD is not as far as is known a naturally occurring However in many countries control of botanicals is explicit. Physical Characteristics Many of the more obscure hallucinogenic plants are not described microscopically in readily available literature and the reviewers recommend that any identification is made by expert botanists. Descriptions of the gross morphology of many plants containing hallucinogens are given by Ernboden.ls0 Heim et aZ.ls1 have described the microscopic appearance of Psilocybe genus. Lophophora williamsii (peyote) was described by Kapadia and Fayez201 DMT and other tryptamines are inactive orally200 and are usually inhaled as siiuffls0 or smoked,200 and thus may be encountered as finely powdered material or mixed with tobacco.Few of the hallucinogens either within natural products extracted or synthetic have been encountered in large amounts with the single exception of LSD. LSD is encountered in many presentations in solution on paper often with elaborate patterns added (in solution) to other tablets on sugar lumps in refilled capsules as illicitly prepared tablets (“microdots” or “domes”) and occasionally mixed with other drUgs,202 although in the reviewers’ experienc July 1983 ANALYSIS OF DRUGS OF ABUSE 793 this is rare. Once encountered many of these presentations are easily recognisable by the forensic analyst but the occurrence of outright fakes is higher than with many drugs and the analyst should be on his guard.Many forensic drug laboratories maintain collections of LSD presentations and the matching of punch marks and tablet sizes may be indicative of a common origin of supply.l0 Dyestuffs in tablets may also indicate to the analyst a common origin of samples.llJ2 Colour Tests Although some presentations of LSD and related indolic hallucinogens may be detected by observing their fluorescence under UV light,203 caffeine and quinine which are common adulterants in drugs,122 and certain detergents153 may respond in like manner.26 Dimethyl-aminobenzaldehyde (DMAB) reacts with many a- or p-unsubstituted indoles in the presence of air and strong acid to form violet or blue compounds.This reaction has been utilised universally as a field test for LSD and other hallucinogen^.^^^ Look205 prepared test papers by saturating filter-paper in DMAB solution and air drying them. A portion of the suspect material was placed on this prepared paper followed by one drop of concentrated hydrochloric acid; a positive response indicated by a violet - red or violet - blue colour indicated that LSD or an indolic hallucinogen might be present. The test was sensitive to 1 pg of LDS. This test was subjected to collaborative study175 and found to be valuable in the preliminary examination of suspected hallucinogens. Alliston et aZ.153 devised an improved test avoiding the prepared test papers whereby one drop of a mixture of DMAB methanol and concentrated hydrochloric acid was added to the sample on a filter-paper.Violet or purple striations on the filter-paper indicated the possible presence of an indolic hallucinogen. This test was found to be more specific and more sensitive than fluorescence. In a further improvement,159 orthophosphoric acid was substituted for hydrochloric acid resulting in a stronger final response and a sensitivity of 10 ng for LSD. This reagent is used as part of a logical tree sequence field test kitg0 issued to Officers of HM Customs and Excise and has been found to remain satisfactory after prolonged storage. In earlier reagents the vapour of concentrated hydrochloric acid tended to destroy labels and instructions and acidify other test reagents in the kit.Some local anaesthetics give orange colours with this reagent which may be helpful in the preliminary screening of suspect cocaine samples where these are common adulter-a n t ~ . ~ ~ ~ The reviewers also consider that the careful use of this test reagent may con-siderably reduce the hazards inherent in the investigation of illicit laboratories. Such laboratories may contain large amounts of prepared drugs precursors and impure specimens and preliminary screening of all suspect materials found may prevent accidental ingestion of drugs during scenes-of-crime investigations such as fingerprint searches. This reagent has been found by the reviewers to give a positive response to many natural products containing tryptamines which may help in the sorting of botanicals.Again we emphasise the impor-tance of laboratory confirmation of all field test results. Ultraviolet and Fluorescence Spectrometry Although field tests may be carried out with advantage on raw botanicals and crude preparations containing hallucinogens the small dosages used makes clean-up by solvent e x t r a c t i ~ n ~ ~ ~ s ~ ~ ~ or column ~hromatographyl~~s~o6 a necessary preliminary to the use of either UV or fluorescence spectrometry. Although LSD in the pure state may be accurately quantified by UV ~ p e c t r o m e t r y l ~ ~ ~ ~ ~ ~ s ~ ~ ~ the impurities likely to be present in illicit prepara-tions make this a technique of limited value and sensitivity.lg6 Bailey ef ~ 1 . ~ ~ ~ showed that UV spectrometry did not distinguish lysergic and isolysergic acid derivatives.DeZan et d 2 0 9 have compiled fluorescence data for a number of hallucinogens including LSD bufotenine DMT (and DET and NN-dipropyltryptamine) psilocin and psilocybin. The inherent fluorescence of many hallucinogens has made fluorescence spectrometry a method of choice for the detection and quantitation of LSD.193s200s207s210 The technique is not specific and is of limited use with impure specimens. However when fluorescence detection is coupled with a separative technique such as TLC or HPLC it is both sensitive and specific and is now widely used for identification and quantitation of LSD and related compounds.4 AnaLryst VoZ. 108 Infrared Spectrometry Although identification of LSD by IR spectrometry may be difficult in view of the small amounts available in individual doses,200 the current generation of instruments has con-siderably reduced this problem.Samples of pure LSD or its salts can be identified by direct IR spectrometry,206 but the presence of impurities may make interpretation of IR data difficult.ls8 Mesley and Evanslg8 made a detailed study of the IR spectra of LSD and its tartrate salts and drew attention to the variability of the spectra and the appreciable changes observed when using potassium bromide discs. The LSD spectra were however distinctive and useful for identification. The same authors211 also published a detailed survey of the IR spectra of tryptamines and their salts and noted the occurrence of polymorphism in many salts and some bases. Detailed discussion was presented on the characteristic absorptions of these compounds which the authors considered would permit recognition of compounds of this type even in the absence of reference materials or spectra.Cromp and Turney212 studied the IR spectra of LSD and related compounds and considered that specific identifica-tion of these compounds was possible by this technique. Martin and A l e ~ a n d e r ~ ~ ~ y ~ ~ ~ reported that they could distinguish LSD and isoLSD but no IR spectra were published. Although the spectra were similar pure samples of ten lysergic and isolysergic acid dialkylamides were reported by Bailey et aL2W to be distinguishable by IR spectrometry. 794 BAKER AND PHILLIPS THE FORENSIC Chromatographic Techniques TLC and HPLC are widely used for both preliminary and final identification and quantifica-tion of LSD and other hallucinogens.LSD is degraded by UV light to lumi-derivatives213 and this property has been used to confirm the identity of LSD by TLC. The chromato-graphy of hallucinogens has been reviewed by Gough and Baker.44 Mass Spectrometry GLC analysis of many hallucinogens is unsatisfactory as many of these compounds are thermally labile.44 Consequently most mass spectral data have been obtained using direct insertion by either EI or CI techniques. Ardrey and Moffat214 presented both EI and CI MS data on 19 hallucinogens and concluded that the identity of specific ergot alkaloids and LSD could be unambiguously confirmed (usually after TLC) by this technique. Psilocin and psilocybin cannot be distinguished by MS1889215 and preliminary identification by chromato-graphic techniques44 must be initially made.Trimethylsilyl (TMS) derivatives have been prepared from some tryptamines prior to MS.216 Psilocin and psilocybin may be distinguished by MS if TMS derivatives are first prepared. Repke et identified and distinguished between these compounds after extraction from a sample of PsiZocybe cubensis. Mass spectral data on LSD218 and related derivativesl8*@9 have been presented and White215 postulated from mass spectral evidence that a third hallucinogen 4-phosphoryl-N-methyltryptamine (baeocystin) originally isolated from P. baeocystis may be present in P. semilanceata. Amphetamines and Structurally Related Drugs Introduction Numerous drugs structurally related to amphetamine (d,Z-a-methylphenethylamine) are controlled by the Misuse of Drugs Act 1971l or by subsequent Modification Orders.174y219 Such drugs may be stimulants and anorectics structurally derived from amphetamine or may be hallucinogens of little medical use derived by ring substitution.Three substances of the latter type named explicitly in Part I of Schedule 2 of the Act are mescaline 2,5-dime t hox y-a ,4-dime t h ylphenet h ylamine (“ STP ” ) and 4-bromo-2,5-dime t hox y-a-me t hyl-phenethylamine (“bromo-STP”) .219 Further compounds of this type are controlled generi-~allyl’~ as follows “any compound [not being methoxyphenamine or a compound for the time being specified in sub-paragraph (a) above] (this being Part I of Schedule 2 of the Act and compounds added by Modification Orders) structurally derived from phenethylamine, any N-alkylphenethylamine a-methylphenethylamine any N-alkyl-a-methylphenethyl-amine a-ethylphenethylamine or any N-alkyl-a-ethylphenethylamine by substitution in the ring to any extent with alkyl alkoxy alkylenedioxy or halide substituents whether or not further substituted in the ring by one or more univalent substituents.” Although numerou July 1983 ANALYSIS OF DRUGS OF ABUSE 795 compounds are subsumed under this paragraph few have been encountered on the illicit market.The following drugs have however been seized by law enforcement officers either in the UK or in the USA 4-methylamphetamine 4-methoxyamphetamine (PMA) 2,5-dimethoxyamphet amine (DMA) 2,4,5-t rimet hox yamphetamine (TMA) 3,4-met hylene-dioxyamphetamine (MDA) 2- and 5-methoxy-3,4-methylenedioxyamphetamine (MMDAs) and 3,4-methylenedioxy-N-methylamphetamine (MDMA) .The stimulant drugs explicitly controlled in Schedule 2 include amphetamine dexamphetamine methylamphetamine, methylphenidate and phenmetrazine (in Part 11) and benzphetamine chlorphentermine, mephentermine and phendimetrazine (in Part 111). Stereoisomers salts and products con-taining the above drugs are also controlled by the Act as are ethers and esters of those drugs specified in Part I of the Schedule. As dexamphetamine and amphetamine are listed separately by the Act it is necessary for the forensic analyst to identify which optical isomer is present in seized material. In view of the greater stimulatory effect of dexamphetamine on the central nervous system220 there may also be medical reasons for discrimination between these two substances in overdose situations.In the UK amphetamine dexamphetamine and methylamphetamine are the most commonly encountered members of the group; the others are rarely found. Amphetamine dexamphetamine and methylamphetamine although widely used in the past to treat a variety of medical conditions have been controlled in the UK since 1964221 when their misuse had become prevalent.222 These drugs have a high potential for abuse and are now more widely used as stimulants than as appetite suppressants,223 originally a principal therapeutic use. and the structure -activity relationships225 of the drugs controlled generically have been discussed.Although some licitly manufactured preparations of amphetamine and structurally related drugs are occasionally examined in the reviewers’ laboratory a result of almost world-wide control is that the majority of amphetamine seizures are manufactured in illicit laboratories.226-228 The hallucinogenic and stimulatory Colour Tests A preliminary identification of licitly manufactured tablets may be made by reference to charts and guide^^-^ or to tablet collections held within laboratories. However the occur-rence of such presentations is decreasing as manufacturers withdraw these materials from the market. Preliminary classification of a sample into this class may be made using commonly available colour tests but the close structural relationships between many of these compounds limits the ability of simple reagents to discriminate between them in many instances.Although Masoudlg described amphetamine and methylamphetamine as giving a similar colour with the Marquis reagent to the opiates (ie. purple) it is the reviewers’ experience that a yellow -orange colour is produced. Clarke21s22 listed the colours obtained when the principal amphetamines were tested with the Marquis reagent. Microcrystal Tests A ~ e r b a c h ~ ~ ~ distinguished d- and d,Z-methylamphetamine and dexamphetamine and amphetamine by volatilisation of the sample into a hanging drop of gold chloride. By simple direct examination of crystals formed between amphetamine optical isomers on a microscope slide Clark230 was able to distinguish d- and I-isomers.Julian and Plein231 formed the 5-nitrobarbituric acid derivatives of the amine drugs found in illicit preparations in the USA (“mini-bennies” or “white cross” tablets); these authors used the same descrip-tions of morphological forms as Clarke41 and presented microcrystallographic data on optical isomers of amphetamine methylamphetamine and some related drugs. Caffeine and theo-phylline which may be present in many illicit drug presentations were found not to interfere with this test. Genest and L o ~ r y ~ 3 ~ have described microcrystalloptic tests for STP. Bastos and Hoffman42 considered that microcrystal tests were the simplest technique for the differentiation of optical isomers of amphetamine and methylamphetamine. These authors also summarised the important reagents used for microcrystal tests.The reviewers consider that great care should be taken in the interpretation of results from these tests in view of the close structural relationships and the lack of data on microcrystalline derivatives of many amphetamines 796 BAKER AND PHILLIPS THE FORENSIC Analyst Vol. 108 Fluorescence Ultraviolet and Visible Spectrometry The majority of illicit amphetamine presentations especially tablets contain only a small proportion of stimulant often mixed with other drugs such as barbiturates. Prior to analysis by any spectrometric method most of these presentations will require a simple clean-up by either column c h r o m a t ~ g r a p h y ~ ~ ? ~ ~ or solvent e ~ t r a c t i o n ~ ~ to remove excipients and any neutral or acidic drugs.Bastos and Hoff man42 briefly reviewed the many fluorescent derivatives that may be prepared from amphetamine. Most are derived from aldehydes or polycarbonyl compounds the resulting fluorophores often being of unknown structure. Clark223 considered that such derivatives were tedious to prepare and non-specific. It is the reviewers’ opinion that although the sensitivity of fluorescence may be helpful in the analysis of trace samples the lack of specificity of the technique taken with the unknown nature of the derivatives renders this unsafe in the forensic analysis of amphetamines. UV spectrometry is widely used in amphetamine analysis but tedious clean-up is needed, as samples are often dilute and low in drug content and amphetamine has low specific ab~orptivity.~~3 The UV spectra of common non-ring-substituted amphetamines and structurally related compounds are generally similar,233 rendering the technique of little value in identification but it is useful for screening purposes.In view of the low absorptivity of amphetamines and the occurrence of considerable levels of impurities in many amphetamine samples UV spectrometry is not in the reviewers’ opinion a reliable method of quantitation of amphetamines. Fontani and M ~ r a n d i n i ~ ~ ~ described a colorimetric method for the determination of amphetamine in solid dosage forms. Derivatisation was with DMAB after heating in 2,5-dimethoxytetrahydrofuran ; secondary and tertiary amines did not interfere with quantitative measurements. Clark223 considered colorimetric determinations of amphetamine to be inherently non-specific a point with which the reviewers agree and consequently do not recommend for forensic analysis.Unreacted amphetamine is only weakly fluorescent .223 Infrared Spectrometry IR spectrometry is an extremely valuable technique for the identification of this class of compound but must almost invariably be preceded by a simple clean-up. In an important paper Mesley and Evans211 reviewed the published spectra of many amphetamines and assigned characteristic absorptions. These authors discussed the occurrence of polymorphism and found it rare in this class of compound although more frequent in salts than bases. Dexamphetamine sulphate and levamphetamine sulphate were found to have identical IR spectra which were different from that of (racemic) amphetamine sulphate.The same was found for the hydrochloride salts. The optical isomers of the free bases were not distin-guishable from the racemic base all being liquids. &Methylamphetamine hydrochloride and d,l-methylamphetamine hydrochloride were not distinguishable by IR spectrometry. Warren et aZ.233 confirmed this finding. distinguished all three optical forms of amphetamine by reaction with d-mandelic acid followed by IR spectrometry of the mandelates. This method has been examined at the reviewers’ laboratory using authentic standards of the three optical forms (checked by polarimetry); it was found that the spectra identifying the optical isomers in the published paper235 were mis-~aptioned,~~~ the spectrum identified as that of the I-amphetamine d-mandelate in fact being that of &amphetamine d-mandelate and vice zleysa.Once this had been ascertained the reviewers found it to be a useful tech-nique. Chromatographic Techniques drugs may be quantified by GLC. isomers of amphetamines may also be separated and quantified by GLC. graphy of these drugs has been reviewed by Gough and Baker.44 Nuclear Magnetic Resonance Spectrometry Clark2lSz2 has presented the spectra of the principal stimulants subject to control. TLC has been widely used for the preliminary identification of amphetamines. These Optical The chromato-HPLC has found little application in this field. Warren et al.233 have presented NMR spectral data on amphetamine and methyl July 1983 ANALYSIS OF DRUGS OF ABUSE 797 amphetamine.Wainer et ~ 1 . ~ ~ ~ differentiated between and quantified d- d,Z- and Z-amphet-amine by proton NMR in the presence of a europium chiral NMR shift reagent; a plot of molar fraction of d-isomer against molar fraction of 1-isomer in synthetic mixtures was linear over the range O-lOO% with a minimum detection limit of 5% of either isomer. Liu et ~ 1 . 2 3 8 used a similar technique to determine the ratios of methylamphetamine enantiomers in mixtures. The reviewers consider that these are valuable techniques as special derivative preparation is not required a problem with GLC quantitation of isomer mi~tures.4~ Bailey and L e g a ~ l t ~ ~ ~ studied the 13C NMR spectrum of amphetamine and found it to be distinctive and suitable for identification.This technique also distinguished salts from bases and the authors considered this an excellent means of distinction identification and confirmation of structural authenticity. Proton NMR has been little used for amphetamine identification as IR spectrometry provides a simpler unequivocal means of identification. Mass Spectrometry MS has been little used with the amphetamine group as the majority of the compounds give weak spectra with little or no molecular ion. Reisch et ~ 1 . ~ ~ ~ gave data on the mass spectra of 18 phenethylamines including amphetamine methylamphetamine chlorphenter-mine and methylphenidate. BellmanlsS reported that both mescaline and STP gave weak molecular ions. Coutts et ~ 1 . ~ ~ ~ prepared N-trimethylsilyl derivatives of amphetamine prior to mass spectral analysis and found that characterisation of this compound using this deriva-tive was possible.Hallucinogenic Amphetamines Although numerous hallu~inogenic~~~ drugs structurally related to amphetamine are controlled by the Misuse of Drugs Act 1971l and its subsequent modification^,^^^^^^^ few have been encountered in the UK although rather more have been seized in the USA. 2,5-Dimethoxy-a-4-dimethylphenethylamine (STP) a hallucinogenic amphetamine,242 was studied by Phillips and M e ~ l e y ~ ~ ~ who used NMR UV IR XRD and MS; these techniques un-equivocally identified the drug. XRD and IR spectrometry showed the base to be poly-morphic. distinguished 3- and 4-methoxyamphetamine by IR and NMR spectro-metry. Bailey et ~ 1 .3 ~ studied ring-monomethylated and ring-monomethoxylated amphet-amines and showed that UV NMR and MS data distinguished the two series but that IR spectrometry was necessary to differentiate between the isomers and distinguish salts from bases. Only weak molecular ions were recorded by MS and the authors considered this technique to be useful only in providing corroborative evidence. The UV spectra of the methoxy compounds closely resembled those of the dimethoxy compounds. showed that it was possible to characterise the monomethoxyamphetamines from the mass spectra of N-trimethylsilyl derivatives. Direct insertion was necessary as these compounds were thermally labile and therefore could not be subjected to GLC. Bailey and L e g a ~ l t ~ ~ ~ showed that the 13C NMR spectra of individual ring-monomethoxyamphetamines were distinguishable and useful for identification and authentication.Salts were also distin-guishable from bases by this technique. Bailey et u Z . ~ ~ ~ in a study of the six isomeric ring-substituted dimethylamphetamines, found that MS analysis gave weak molecular ions and was of little value in distinguishing isomers. NMR spectrometry was found to identify individual compounds but only with difficulty. However IR spectrometry allowed the bases to be distinguished provided that samples were free from impurities and excipients. Shaler and Padden245 identified 2,5-dimethoxyamphetamine from NMR MS and IR data. Bailey246 showed by studying the mass spectra of the dimethoxyamphetamines that although the spectra were weak they were distinguishable and characteristic but required careful assessment.He considered that mass spectra alone were probably not reliable for the unequivocal identification of individual isomers especially if small amounts of other sub-stances with strong mass spectra were present. In a subsequent study Bailey et uE.247 used proton NMR IR and MS and showed that positive identification of each isomer could be made. IR and NMR spectrometry were shown to distinguish and identify each isomer and also to distinguish the monomethoxyamphetamines from dimethoxyamphetamines. Bailey Warren et Coutts e 798 BAKER AND PHILLIPS THE FORENSIC Analyst Vol. 108 and L e g a ~ l t ~ ~ ~ found the 13C NMR spectra of these six compounds were distinctive suitable for identification and authentication purposes and would distinguish salts from bases.The same authors also found that this technique would identify each of the six trimethoxy-amphetamines.248 Bailey et aZ.224 identified 2- 3- and 4-methoxy-N-methylamphetamines 3-methoxy-4,5-methylenedioxyamphetamine and 3,4-methylenedioxy-N-me t hylamphe t amine by proton NMR and IR spectrometry. Proton NMR spectrometry also distinguished the salt of each compound from the base and from the corresponding non-N-methylated compound.37 Other Forensic Aspects Although Lophophora williamsii (peyote) the cactus containing mescaline is not itself controlled by the Misuse of Drugs Act 1971 ,l it may be necessary for example under military law to identify this botanical. Chao249 has discussed the legal status of peyote in the USA.Kapadia and Fayez2o1 and Emb~den~~O have described the physical appearance of the plant and the former authors have discussed its active constituents.201 Barron et aL251 discussed the significance of impurities in illicit methylamphetamine samples. A knowledge of contaminants and by-products in such samples may help the forensic analyst to decide whether the drug was manufactured licitly or illicitly and upon its method of preparation Comparison of different seizures may be possible using fingerprint patterns of impurities and knowledge of actual impurities may help to reduce interferences in quantitative analysis. Many of the contaminants in illicitly manufactured materials are more toxic than the intended drug and knowledge of these is therefore important from a toxicity standpoint.Impurities in amphetamines have been studied by a variety of techniques including UV,251 IR,251-253 NMR251-258 and MS.252-254J599260. Lukas~ewski~~~ studied the UV IR NMR and mass spectra of a number of possible precursors intermediates and impurities in samples of MDA that might interfere with analyses. A review with 40 references on the comparison of illicit drugs has been published.228 Phencyclidine and Analogues Introduction 1-( 1-Phenylcyclohexy1)piperidine (phencyclidine PCP) is controlled in the UK by a Modification Order261 made under the Misuse of Drugs Act 1971.l Although illicit importa-tion and misuse of this drug has not been widely reported in the UK it is widely abused in the USA262 and Canada.263 The ready availability of this drug is due largely to its com-paratively simple synthesis from cheap and readily available starting material^.^^^^^^^ It was at one time available as an anaesthetic for both human and veterinary purposes but these applications were discontinued because of adverse effects on patient^.^^^^^^^ Further, the therapeutic dose was found to be close to the toxic dose and many deaths have resulted from the misuse of this Two principal problems confront the forensic analyst when examining samples of suspected PCP the presence of contaminants resulting from careless illicit synthesis and the large number of structurally related analogues that have been manufactured in illicit laboratories.The toxic266 precursor 1-piperidinocyclohexanecarbonitrile (PCC) is controlled in the USA and a number of analogues controlled in both the USA and are candidates for international control.These include the thienyl analogue tenocyclidine (known as TCP), rolicyclidine (PHP or PCPy in which pyrrolidine replaces piperidine) and eticyclidine (PCE , where ethylamine has replaced piperidine). PCC has been found to be present in many samples of illicit PCP.262 PCP and related preparations may be taken orally or smoked when mixed with spices or cannabis268 ; forensic analysts should therefore be alert to this when analysing apparently innocuous botanical materials. A major problem at least in the UK is the general un-availability of authentic standards of PCP and related substances ; the identification of PCP should therefore be made by the most rigorous techniques.Colour Tests improved cocaine test of Alliston et PCP reacts positively to the cobalt(I1) thiocyanate field test for cocainelg and in the in a field test kit.g0 PCE has been reported t July 1983 ANALYSIS OF DRUGS OF ABUSE 799 react similarly.269 PCP and PCE give no colour with the Marquis reagent.26g Although data are lacking the reviewers would expect the majority of PCP analogues to give similar results with these field tests. Ultraviolet and Infrared Spectrometry Column chroma tog rap hi^^^^ or solvent extraction264 techniques have been described for the clean-up of illicit samples of PCP; in view of the probability of adulterants and impurities in such samples the reviewers consider that this is an essential first step prior to spectrometric analysis.UV spectra have been found to differentiate between groups of structurally related PCP analogues271 but were found not to be useful for identifying particular compound~,~67 for which IR spectrometry is recommended. Bailey et aZ.,271 in a study of PCP and five analogues, found that the IR spectra of both the free bases and the hydrochloride salts were clearly distinguishable. In a related study of PCE and its analogues Bailey and L e g a ~ l t ~ ~ ' found the IR spectra of the bases were similar to but distinguishable from those of the PCP analogues whereas the IR spectra of the hydrochloride salts were suitable for identification purposes. Nuclear Magnetic Resonance Spectrometry NMR spectrometry has been found to be useful for distinguishing PCP and related com-pounds.Proton NMR spectra have been used to distinguish PCP from TCP its thienyl analogue,271 from PCE and its anal0gues~~7 and in the identification of PCE.26s 13C NMR spectra have been reported to be suitable for the identification of PCP PCE and their analogues.272 Brine et aZ.273 studied the hydrochloride salts of PCP and 16 related substances by 13C NMR and considered that they could be easily differentiated from each other using this technique but that differentiation by proton NMR was more difficult. Bailey and L e g a ~ l t ~ ' ~ studied the 13C NMR spectra of 20 possible aminocyclohexanecarbonitrile pre-cursors of PCP and related compounds and found that such spectra could be used to identify these compounds when reference materials were available and would provide a preliminary identification of unknown materials.Chromatographic Techniques and related compounds. possibility of misidentification of this compound when GLC was used for its analysis. Gough and Baker44 have discussed in detail the TLC GLC and HPLC analyses of PCP These authors drew attention to the instability of PCC and the Mass Spectrometry Bailey et aZ.271 analysed PCP and some analogues by MS and found it a valuable technique which not only would distinguish the compounds used in this particular study but would also help to elucidate the structure of uncharacterised compounds structurally related to PCP. Molecular ions were found to be variable in intensity but to be assignable. Freeman and Martin26s found that MS using single ion monitoring would identify phenylcyclohex-1-ene (a product resulting from the smoking of PCP) in the presence of PCP.Bailey and L e g a ~ l t ~ ~ ~ found MS (with prior GLC) suitable for the identification of PCP PCE and their analogues. In two studies Cone and c o - ~ o r k e r s ~ ~ ~ ~ ~ ~ ~ analysed PCP precursors analogues and metabolites by CI MS; these authors considered this technique to be the most specific means of identifying these compounds. In view of the structural similarities between PCP and its analogues and the rarity of their occurrence (at least in the UK) the reviewers consider that both IR and MS data should be used before a conclusion is reached as to the identity of a suspected phencyclidine sample.Mass spectral data on PCP have been presented by Lindgren et Methaqualone Methaqualone is at present controlled in the UK by being specified in Part I11 of Schedule 2 of the Misuse of Drugs Act 1971.l Preliminary identification of licit preparations may be made by reference to charts and guide^.^-^ However the recent large numbers of well manufactured fake preparations some containing methaqualone and many more containin 800 BAKER AND PHILLIPS THE FORENSIC Analyst “01. 108 other drugs encountered in North America and Europe make it imperative to confirm the presence of methaqualone by rigorous analysis. Methaqualone reacts positively to the cocaine field test of Alliston et aZ.,152 which although by no means specific is useful as a sorting test because no licit or illicit tabletted preparation of cocaine has been encountered, in the reviewers’ recent experience, The relatively large dose of methaqualone (150 mg or more) in many licit preparations allows the characteristic UV spectrum of methaqualone277 to be used for identificati~n,~?~ particularly as its absorption maxima vary with pH.However in view of the occurrence of fakes and structurally related usually illicitly manufactured compounds such as meclo-qualone more specific techniques should in the reviewers’ opinion be applied. Identifica-tion by IR spectrometry is possible and has been described.279 Chromatographic techniques for the analysis of methaqualone are included in reviews by Gough and Baker.44 MS data on methaqualone have been p r e ~ e n t e d . ~ ~ ~ ~ Dal Cason et a1.280 presented the IR NMR and MS data on methaqualone and 15 analogues and isomers including mecloqualone; using data from these techniques these authors could differentiate between all the compounds studied.Dal Cason et aL281 have discussed the illicit synthesis of mecloqualone. This drug is a candi-date for international control and its forensic identification may become more important. Barbiturates Introduction Barbiturates have a high dependence and addiction potential even in “normal” clinical usage.282 Barbiturates are not currently controlled in the UK by the Misuse of Drugs Act 1971l but many countries do exercise control over this class of drugs. A number of these substances are specifically listed in the Schedules to the Convention on Psychotropic Sub-stances 1971,283 to which the UK is not currently a signatory.Schedule I11 includes amylobarbitone cyclobarbitone pentobarbitone and quinalbarbitone whereas barbitone, methylphenobarbitone and phenobarbitone appear in Schedule IV. Over 2 500 barbiturates have been synthesised and about 50 have found clinical acceptance2B4; 11 barbiturates (including thiopentone sodium) are currently available in 23 preparations on prescription in the UK285 compared with 61 preparations of 14 barbiturates in December 1979. Three possible systems of control in the UK have been publically discussed (a) all 5,5-disubstituted barbituric acid derivatives with or without addition of methylphenobarbitone ; or (b) all 5,5-disubstituted barbituric acid derivatives exclztding phenobarbitone ; or ( c ) five explicit medium acting barbiturates that are frequently abused namely amylobarbitone, butobarbitone cyclobarbitone pentobarbitone and quinalbarbitone.Stead et nl.284 have discussed these proposals in detail. Analytical schemes for the analysis of suspect barbiturate-containing samples will vary depending on the form of control finally adopted. Whatever that might be the reviewers believe that the forensic analyst will still be required to identify the particular barbiturate present. The case of Nuir V S . Smith,57 although concerning Cannabis products implies that the Courts require explicit identification of controlled drugs. In addition the identification of barbiturate preparations specifically in overdose situations, is perhaps of greater importance as the physiological and toxicological properties of these drugs vary considerably.286 Colour Tests Whilst licit barbiturate preparations remain widely available many samples of tablets or capsules may be tentatively identified by reference to the appropriate guide^.^-^ Such identification must be confirmed by laboratory tests.The classic chemical field test for barbiturates is the modification by Dille and K ~ p p a n y i ~ ~ ? of the Zwikker test288; colours are developed by the addition of an amine to an anhydrous cobalt (11) acetate solution of a barbiturate. Many conimercial field test kits contain reagents based on this reaction. De Faubert Maunder289 considered that the original test lacked sensitivity and specificity and developed an improved version.A single solution of cobalt(I1) thiocyanate in methanol containing 2,6-dimetliylniorpholine is added to a small amount of the suspect sample on a filter-paper a violet or purple colour on the paper is interpreted as a positive reaction. Detection limits of 0.2-0.5 pg were obtained with five common barbi-turates ; similar colours were given by some hydantoin hypnotics. The author advised31 tha July 1983 ANALYSIS OF DRUGS OF ABUSE 801 the test be used as part of a logical tree field test procedure and such a test is included in a commercial field test kit.g0 Following a positive field test the presence of a barbiturate must be confirmed in the laboratory. Ultraviolet Spectrometry This technique has been very widely used for the determination of barbiturates using methods devised by B r o u g h t ~ n .~ ~ ~ 5,5-Disubstituted barbituric acid derivatives are weak acids with two pK values initial ionisation is at pH 7-8 and the second at pH 12-13. The mono-anions exhibit UV absorption at around 240nm and the di-anions absorb at about 255 nm.284 Stead et al.284 found that UV spectrometry alone would reliably establish the presence of a 5,5-disubstituted barbiturate in a sample. These authors also presented a useful discussion on the use of UV spectra in barbiturate analysis; D a g l i ~ h ~ ~ l and Higgins and Leach292 have also described the applications of UV spectrometry to barbiturate analysis. If only specific barbiturates [control system ( c ) ] were to become controlled in the UK or if phenobarbitone is excluded [system ( b ) ] application of UV spectrometry to barbiturate analysis becomes more limited.Problems may also occur when two barbiturates are mixed as occurs even in some licit preparations. Synthesis of barbiturates from readily obtainable chemicals is a simple process and should barbiturates become controlled there seems little reason why their manufacture in illicit laboratories should not become common (as is so with amphetamines). Further there is no reason why such laboratories should limit their opera-tions to currently common barbiturates. If such illicit operations become widespread, impurities and uncommon preparations may limit even further the use of UV spectrometry in the screening of samples for 5,5-disubstituted barbiturates. Post-column ionisation techniques utilising the increased UV absorption of ionised barbiturates over the free acids, have been used to detect these compounds after separation by HPLC.293 Nevertheless the reviewers consider that in spite of the drawbacks outlined above UV spectroscopy of barbiturates could play a useful part in a fuller analytical scheme for the identification of these compounds as a class whichever method of control is finally adopted in the UK.Infrared Spectrometry Most IR studies of barbiturates have been made on pure compounds; in view of their structural similarities preliminary clean-up of samples is likely to be necessary prior to use of IR spectrometry. Mesley and clement^^^^ discussed the IR spectra of barbiturates in considerable detail in a study of 12 such compounds and drew attention to the widespread occurrence of polymorphism in this class of compounds (up to 13 forms of phen~barbitone~~~).However they found that although different forms of the same material might not always be distinguishable from their IR spectra they could be differentiated from other barbiturates. Stead et al.284 reported that only cyclobarbitone and heptabarbitone could not be distinguished from 28 other barbiturates. Heptabarbitone was not studied by Mesley and clement^.^^^ Discussions of the IR spectra of barbiturates have also been presented by Chapman and MOSS.^^^^^^ However UV spectrometry does not identify the actual drug present. Nuclear Magnetic Resonance Spectrometry Although this technique has not been widely used in barbiturate analysis Stead et aLm4 considered it most promising and reported that all the major barbiturates could be distin-guished.However samples of good purity were considered necessary and these authors reported problems in interpretation of the NMR spectra of mixtures. Chromatographic Techniques Although IR NMR and some MS techniques may be used to identify pure or purified samples of barbiturates the reviewers consider that chromatography will be a necessary part of any analytical scheme to identify these compounds either prior to IR spectrometry (in order for example to distinguish cyclobarbitone and he~tabarbitone28~) or prior to IR or MS in order to obtain pure samples. It may also be necessary to use a chromatographic screening technique in order to reduce the final identification problem.Confirmation of final identification by chromatography using an authentic standard may also be desirable 802 BAKER AND PHILLIPS THE FORENSIC Analyst Vol. 108 TLC GLC (often with prior derivative formation) and more recently HPLC have been widely used for the analysis of barbiturates; these topics have been reviewed by Gough and Baker.44 Mass Spectrometry EI MS has been found to be of limited value for barbiturate identification as few of these compounds give characteristic molecular ions.284g297-298 Fales et aZ.299 used both EI and CI MS in a study of eight important barbiturates and showed that they could be distinguished. GC - MS of the alkyl derivatives of barbiturates has been shown to provide considerably more data for the identification of b a r b i t u r a t e ~ ~ ~ J ~ ~ ~ ~ ~ ~ ; however some barbiturates give the same derivative on methylation and prior screening by an alternative technique may be necessary.44 Jones and Whitehou~e~~~ observe that CI MS was not specific for isomeric barbiturates and studied the anion mass spectra under CI conditions of 30 barbiturates and obtained simple spectra that distinguished all the compounds except butalbarbitone and allylbutylbarbitone.Concluding Remarks Forensic drug identification has progressed in the last three decades from the application of simple testing procedures to the almost routine use of the most powerful instruments available in modern chemistry. This is simply a reflection of the ever increasing pressure on the analyst unequivocally and rapidly to identify the drugs in any sample presented.The reviewers consider this trend likely to continue and it behoves analysts not only to keep up-to-date with the scientific literature but also to keep abreast of developments in analytical instrument at ion. Most common drugs can be simply and rapidly identified by existing techniques but with the misuse of drugs increasing world-wide continual method development is necessary if forensic laboratories already in many instances overstretched are to be able to keep up with a workload of increasing amount and complexity. However certainty of identification must not be compromised in a search for analytical speed. It is outside the scope of this review to discuss presentation of evidence but it is clearly necessary for the expert to be totally familiar with the legal status of the substances identified.The haphazard growth of legislative control of drugs and the pitfalls encountered in the UK up to implementation of the comprehensive Misuse of Drugs Act 1971 have been extensively re~iewed.30~ For the forensic analyst it is salutary and necessary to remember that the outcome of nearly every positive identification of a controlled drug is a sworn witness statement which may require oral presentation of evidence in Court. It is then that the expertise possessed by the analyst is truly tested. Useful discussions on the role of the expert witness from both the UK303-306 and the USA,3071308 have been presented. 1. 2. 3. 4. 5. 6. 7. 8. 9.10. 11. 12. 13. 14. 15. 16. 17. 18. 19. 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Moffat A.C. and “Monthly Index of Medical Specialities” (MIMS) Medical Publications London March 1983. Skinner R. F. Gallaher E. G. and Predmore D. B. Anal. Chem. 1973 45 574. Dille J. M. and Koppanyi T. J Am. Pharm. Assoc. 1934 23 1079. Zwikker J. J . L. Pharm. Weekbl. 1931 68 975. De Faubert Maunder M. J. Analyst 1975 100 878. Broughton P. M. G. Biochem. J. 1956 63 207. Daglish C. in Clarke E. G. C. (assisted by Berle J.) Editors “Isolation and Identification of Drugs,” Pharmaceutical Press London 1969 p. 103. Higgins G. and Leach H. in Clarke E. G. C. (assisted by Lang M. and Marriott K. G.) Editors, “Isolation and Identification of Drugs,” Volume 11 Pharmaceutical Press London 1975 p. 873. Clark C. R. and Chan J. L. Anal. Chem. 1978 50 635. Mesley R. J. and Clements R. L. J . Pharm. Pharmacol. 1968 20 341. Mesley R. J. Clements R. L. Flaherty B. and Goodhead K. J . Pharm. Pharmacol. 1968 20 329. Chapman D. I. and Moss M. S. in Clarke E. G. C. (assisted by Lang M. and Marriott K. G.), Editors “Isolation and Identification of Drugs,” Volume 11 Pharmaceutical Press London 1975, 51 154. Don M. C. H. J . Forensic Scz. Soc. 1981 21 41. p. 935. Jones L. V. and Whitehouse M. J. Biomed. Mass Spectrom. 1981 8 231. Coutts R. T. and Locock R. A. J . Pharm. Sci. 1968 57 2096. Fales H. M. Milne G. W. A. and Axenrod T. Anal. Chem. 1970 42 1432. Menez J. F. Berthou F. Picart D. Bardon L. and Floch H. H. J . Chromatogr. 1976 129 155. Gilbert J. N. T. Millard B. J. and Powell J . W. J . Pharm. Pharmacol. 1970 22 897. Phillips G. F. Chem. Br. 1972 8 123. Samuels A. Med. Sci. Law 1974 14 17. Coleman R. F. and Walls H. J. Crim. Law Rev. 1974 276. Turner-Samuels D. Chem. Ind. (London) 1975 857. Howard A. J. Chem. Ind. (London) 1975 858. Phillips K. A. J . Forensic Sci. 1977 22 457. Kogan J. D. J Forensic Sci. 1978 23 190. Received December 3rd 1982 Accepted January 14th 198

ISSN:0003-2654    

DOI:10.1039/AN9830800777

出版商:RSC    

年代:1983

数据来源: RSC

摘要:

808 Analyst July 1983 Vol. 108 pp. 808-812 Design and Performance of an Unenclosed Tantalum Atomiser for Atomic-absorption Spectroscopy J. Aggett Chemistry Department University of Auckland Auckland New Zealand An unenclosed tantalum-strip atomiser for atomic-absorption spectroscopy is described. Its performance for the determination of a number of elements has been evaluated and several applications are reported. Keywords Tantahm-styip atomiser ; atomic-absorption spectroscofiy Although the major thrust in the development of electrothermal atomisation for atomic-absorption spectroscopy has been associated with graphite furnaces there has been occasional interest in the use of metal surfaces in the form of wires,l boat^^-^ and tubes6s7 for this purpose. Examination of the earlier publications on metal atomisers shows that in general these devices were particularly effective for atomisation of the more volatile elements and in this respect they compare very favourably with graphite atomisers.Their performance with less volatile ele-ments was seldom reported possibly because of the limitations imposed on their operating temperatures by the melting-points of the metals. More recently the use of metals in com-bination with graphite furnaces either as furnace or impregnated saltslOJl has been shown to be beneficial not only in terms of sensitivity but also in terms of precision and the lifetime of the furnace. Of particular interest in this respect is the observation of L’Vov and Pelievag that the sensitivities of several relatively involatile elements are enhanced by the use of a tantalum furnace lining in a graphite furnace.This suggests that limitations of the earlier metal atomisation devices may be related to the temperature above the volatilisation surface, and hence to the design rather than to the nature of the material itself. It therefore seems that metal atomisers may eventually prove to be more useful than is the current general belief. This is not meant to imply that tantalum and other metals will prove to be better than graphite for the atomisation of all elements as it seems clear from currently available informa-tion that the relative benefits of metals and graphite for atomisation of any particular element are associated with the mechanism of atom formation of that element. This is illustrated by the findings of L’Vov and Pelievag and Gregoire and Chakrabarti,12 both of whom have reported enhanced sensitivities from tantalum for elements known to form carbides.On the other hand the latter have concluded that the decreases in sensitivities that they observed when vanadium molybdenum and nickel were atomised from tantalum were due to the formation of non-volatile intermetallic compounds between these elements and tantalum. In this laboratory an “unenclosed” tantalum-strip atomiser was originally developed in association with a study on the mechanism of atom formation in electrothermal atomisers.13 The design of this model overcame the need to enclose completely the tantalum in order to protect it from reaction with nitrogen or oxygen at high temperature and thus made it more simple to use than previous metal atomisers.A modified version of this apparatus was re-ported at the Fifth International Conference on Atomic Spectro~copyl~ in 1975. From that time this atomiser has been used in a number of studies involving trace elements. This paper reports details of the construction of the atomiser its performance potential and a selection of applications for which it has been used. Atomiser Design I t consists of two water-cooled brass terminal blocks separated by a mica sheet and is mounted on a poly(viny1 chloride) base the dimensions of which can be altered to fit the burner apertures on different commercial atomic-absorption spectrophotometers. One of the terminal blocks contains the two apertures used to limit the cross-section of the optical path through the tantalum strip.They are machined so that the diameter increases from 1.5 mm at the inside of the block to 3 mm at the outside. The tantalum strips are formed on a jig into the shape shown in Fig. 2 from ribbon 6 mm wide x 0.25 mm thick x 35 mm long. The jig consists of four parts (1) the main body; The design of the atomiser is illustrated in Fig. 1 AGGETT 809 (2) a removable rod of diameter 2.3 mm; and (3) a slide that can be positioned and secured by (4 and 5) the positioning and locking screws. Shaping of a tantalum strip is accomplished by inserting it into the gap between the rod and the main body so that it protrudes an equal distance on each side. The ends of the tantalum strip are then bent upwards until they meet.The slide is moved upwards until the top is 0.5 mm below the top of the rod and is secured in that position by the locking screw; reproducibility in this action is assisted by use of the positioning screw. The ends of the tantalum strip are then bent downwards over the top of the slide and the rod is removed from the main body to release the tantalum strip. Finally the gap at the top is adjusted to approximately 1 mm by hand pressure. These shaped strips are supported on shoulders in the terminal blocks and positioned so that light from the hollow-cathode lamp passes immediately above the lower internal surface of the loop in the tantalum. This alignment is readily made with the aid of a stainless-steel syringe needle inserted through the light-guide apertures.The position of the tantalum strip is then fixed from above by tightening the screws through the clamping blocks. These tantalum strips can accommodate up to 20 pl of sample. Fig. 1. Exterior view of atomiser showing 1 power connectors; 2 fila-ment clamping blocks; 3 light guide aperture; and 4 cooling water circula-tors. -1 I- 50 m m l 3 1 4 Fig. 2. The tantalum strip and the jig on which it is formed: 1 main body; 2 removable rod; 3 slide; 4 positioning screw; and 5 locking screw. Samples for analysis are inserted into the loop of the tantalum strip through the access hole at the top of the assembly. Although the strip is accessible in this way it is protected from the atmosphere by a stream of argon supplied from below through the gas-mixing chamber of the flame atomiser via the dinitrogen oxide line and the fuel inlet.The nebuliser assembly is removed and the front of the mixing chamber sealed with a bung. It is necessary to use argon for this purpose as nitrogen has been found to destroy the tantalum strips very quickly, probably through formation of tantalum nitride. An important feature of the design is the provision of shaped projections on the end of each clamping block. These projections overhang the ends of the clamping blocks by 1 mm and have curved lower surfaces so that argon is directed over the top of the tantalum strips in the vicinity of the terminals in order to prevent condensation of solvents and in particular acids that have otherwise been found to destroy the tantalum strips in this region.The atomiser temperature is controlled by a four-stage programmable digital current -timer unit designed to supply a current of <300 A for up to 100 s in each stage from a 12 V supply. As with other devices for electrothermal atomisation the current - time sequence requlres precise control for acceptable performance. The atomiser has normally been used with a Varian 1000 spectrophotometer. Lamp currents have usually been set at 10 mA and the result obtained in the peak-height mode using a recorder with response time 0.5 s full-scale deflection 810 AGGETT DESIGN AND PERFORMANCE OF AN UNENCLOSED AnaZyst VoZ. 108 The apparatus has been modified to accommodate 4.5 mm diameter West-type graphite rods by machining recesses into the shoulders of the terminal blocks.Graphite supplied by Johnson Matthey (Specpure Grade 11) was chosen for this purpose as its heating characteristics were found to be compatible with the power supply designed for use with tantalum. How-ever the graphite rods have been used infrequently as tantalum strips have been adequate for laboratory requirements. Results and Discussion The general level of performance of the atomiser is illustrated by the detection limit data in Table I. For most of the elements in Table I these detection limits compare reasonably well with those reported for commercial graphite furnaces an indication that this tantalum atom-iser is capable of a satisfactory performance for a number of elements insofar as sensitivity is concerned. There is cause to believe however that by increasing the diameter of the aper-tures in the terminal block and using the atomiser with a more sophisticated spectrophoto-meter than the Varian 1000 these detection limits can be improved e.g.Townshend and Shahidullah15 reported a sensitivity of 10-l2 g for lead when this model of the tantalum atom-iser was used with a Varian-Techtron AA-5 spectrophotometer. Comparison between signals obtained with atomisation of chromium from tantalum and from a West-type graphite rod showed that the former gave higher sensitivity. I t is possible that the lower sensitivity observed when chromium was atomised from graphite is associated with the formation of non-volatile carbides because Veillon et aZ.l* have demonstrated that about 35% of chromium is not volatilised from the surface of graphite atomisers.Attempts to determine iron and nickel with the tantalum atomiser have been unsuccessful. For both elements the blank signals obtained with new strips were very high and these per-sisted through several atomisation cycles sufficient to remove trace amounts of other impuri-ties in the tantalum. Nickel however was found to be detectable when atomised from a molybdenum strip the detection limit being lop9 g which is inferior to that generally quoted for graphite furnaces. Molybdenum also appeared more useful for the determination of cobalt the detection limit being better by a factor of approximately two over that obtained with tantalum. These observations with cobalt and nickel are similar to those reported by McIntyre et However in our over-all experience molybdenum was found to be less ductile than tantalum and it was much more readily attacked by trace amounts of air at high tempera-tures.The reproducibility is adequate e.g. 20 successive analyses of chromium (25 ng ml-l) gave a mean absorbance reading of 0.243 with a relative standard deviation of 2.4%. The lifetime of the tantalum strips was found to be of the order of 150-200 atomisation cycles when used for the analysis of the elements in Table I. In one test run the absorbance For these reasons tantalum has been the metal of our choice whenever possible. TABLE I ATOMISATION CONDITIONS AND DETECTION LIMITS Atomisation conditions Element ' Time/s Current/A' Detection limit*/g Arsenict . . 3 70 5 x 10-10 Barium 2 75 1 x 10-11 Cadmium .. 3 40 3 x 10-18 Calcium . . . . 3 70 <5 x 10-12 Chromium . . . I 3 55 5 x 10-12 Cobalt 3 80 3 x 10-10 3 50 1 x 10-11 3 70 5 x 10-1' Lead (283 nmi Magnesium . . 3 70 <5 x 10-12 Manganese 3 45 1 x 10-12 Strontium . . 3 65 2 x 10-12 Thallium 2 48 2 x 10-1' mean absorbance of the blank plus three standard deviations in the blank. . . . 3 45 1 x 10-11 ZEer * Defined as the concentration that gives an absorbance that is equal to the t In the presence of nickel. Results limited by purity of water J d y 1983 TANTALUM ATOMSIER FOR ATOMIC-ABSORPTION SPECTROSCOPY 81 1 of a solution of copper was measured at regular intervals during the use of a tantalum strip. The mean absorbance obtained during atomisation cycles 100-105 was 4% lower than the mean of the first 10 atomisations.Through atomisation cycles 150-155 the mean absorbance had fallen by a further 3% and the strip still appeared to be in good condition and shape. The minor change in sensitivity during the life of this strip was probably caused by very small movements of the strip with repeated thermal cycling rather than by deterioration of the tantalum surface within the loop. These observations suggest that in terms of long-term stability tantalum is superior to graphite a result that is in line with that of L’Vov and P e l i e ~ a . ~ No extensive studies have been made on interferences. However it has been established that the atomiser does not cope with undigested biological materials as these tend to char and remain unoxidised in the loop after ashing and atomising.With digested materials the prob-lem of interferences is essentially reduced to one of the interaction of inorganic species. One such study on the interference of sodium potassium magnesium and calcium chlorides on the analysis of chromium (in the form of dichromate) showed that of these only calcium chloride, which is the least volatile of the chlorides still caused loss of signal after heating at elevated temperature during the “ashing” step. At the 2000 ng ml-l level potassium did not interfere and sodium reduced the signal by 8%. One possible consequence of the use of tantalum or other metallic surfaces for atom forma-tion is that the mechanism of atomisation may be altered from that which occurs when graph-ite is used and this may result in a different appearance temperature for the atoms of a given element and a different pattern of interferences.Although this was first demonstrated for a limited number of examples in 1974,13 and the postulated atom formation mechanisms were subsequently confirmed by several separate studies,17 there do not appear to have been any further extensive studies on the effect of the atomisation surface on interferences. The atomiser has been used in a number of studies that required the determination of trace elements. The determination of strontium concentrations has been used to indicate whether moisture in a container originated from condensation or from leakage of seawater. Strontium analysis has also been used in studies of estuarine waters.Organotin fungicide residues have been determined in kiwi fruit (Actinidia chinensis). Copper lead and zinc have been deter-mined in seawater in the course of an extensive study of the distribution of these elements in the Waitemata and Manukau harbours in Auckland.18 In this study matrix problems were avoided by extracting the analytes into organic solvents prior to analysis. These pre-concentration procedures also improved the precision of the analysis. The atomiser has also been used for the determination of arsenic in a fundamental study of the formation of arsine, methylarsine and dimethylarsine by conventional hydride generation methods in atomic-absorption spectroscopy . l9 Data from this study are shown in Fig. 3. 1 10 100 lnterferent elemental concentration/pg ml-’ Fig.3. Interference of magnesium chloride and calcium chloride on chromium (0.1 p g ml-l) 1 Cr in presence of Mg without preliminary heating; 2 Cr in presence of Mg with preliminary heating for 30 s a t 28 V; 3 Cr in presence of Ca without preliminary heating; and 4 Cr in presence of Ca with preliminary heating for 30 s a t 28 V 812 AGGETT Experience with this atomiser has shown it to possess four significant positive features: (1) high sensitivity with volatile and moderately volatile elements ; (2) relatively high precision, resulting at least in part from the fact that the sample cell is specifically designed to contain the volume of sample normally used; (3) reasonable long-term stability; and (4) it provides an inexpensive and easily operated accessory for occasional users of the technique.Simplicity of operation has also proved advantageous for teaching purposes. Its more important limitations appear to be the inability to handle undigested organic matrixes and to produce successfully atoms of less volatile elements. However in the light of the observations of L’Vov and Pelievag the latter disadvantage seems most likely to be associ-ated with the form of the filament rather than its chemical nature. 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. 14. 15. 16. 17. 18. 19. References Williams M. and Piepmeier E. H. Anal. Chem. 1972 44 1342. Donega H. M. and Burgess T. E. Anal. Chem. 1970 42 1521. Hwang J. W. Mokeler C. J. and Ullucci P.A. Anal. Chem. 1972 44 2018. McIntyre M. S. Cook M. G. and Boase D. G. Anal. Chem. 1974 46 1983. Schrenk W. G. and Everson R. T. Appl. Spectrosc. 1975 29 41. Ohta K. and Suzuki M. Talanta 1975 22 465. Sychra V. Kolihovb D. VyskoEilovA O. and HlavaC R. Anal. Chim. Acta 1979 105 263. Wall C. D. Talanta 1977 24 755. L’Vov B. V. and Pelieva L. A. Can. J . Spectrosc. 1978 23 1. Zatka V. J. Anal. Chem. 1978 50 538. Manning D. C. and Slavin W. Anal. Chem. 1978 50 1235. Gregoire D. C. and Chakrabarti C. E. Spectrochim. Acta Part B 1982 37 611. Aggett J. and Sprott A. J. Anal. Chim. Acta 1974 72 49. Aggett J . Fifth International Conference on Atomic Spectroscopy Melbourne 25-29 August, Townshend A. and Shahidullah M. personal communication 1979. Veillon C. Guthrie B. E. and Wolf W. R. Anal. Chem. 1980 52 457. Matousek J. P. Prog. Anal. A t . Spectrosc. 1981 4 247. Simpson J. D. MSc Thesis University of Auckland 1978. O’Brien G. A. PhD Thesis University of Auckland 1983. 1975 Paper A2. Received November 8th 1982 Accepted February 7th 198

ISSN:0003-2654    

DOI:10.1039/AN9830800808

出版商:RSC    

年代:1983

数据来源: RSC

摘要:

Analyst July 1983 Vol. 108 $9. 813-520 813 Determination of Lead in Blood by Atomic-absorption Spectroscopy with Electrothermal Atomisation Denis J. Hodges and (the late) Douglas Skelding Research and Development Department The Associated Octel Company Limited P.O. Box 17 Oil Sites Road, Ellesmere Port South Wirral L65 4HF A method is described for the determination of lead in blood by atomic-absorption spectroscopy with electrothermal atomisation. The addition of orthophosphoric acid together with pre-coating the graphite tube with molybdenum is effective in minimising matrix interference and in promoting stable routine operation. No special oxidative treatment is necessary to prevent loss of lead during the drying and ashing steps of the procedure. The range of the method is 5-50 pg dl-l.Keywords Lead determination ; blood ; electrothermal atomisation atomic-absorption spectroscopy ; matrix modification The development of a rapid method for the determination of lead in urine by electrothermal atomisation atomic-absorption spectrometry (ETA - AAS) has previously been reported.lF2 Alongside this method for physiological monitoring of personnel engaged in the manufacture of alkyl lead compounds the more widely used method of monitoring exposure by determination of lead in blood is carried out. In this determination there was also a requirement for a rapid method of analysis in order to provide greater flexibility in instrumentation and laboratory resources. The method used previously in these laboratories for many years is that described by Bambach and Burkey3 in which lead is determined spectrophotometrically using dithizone after wet ashing of the blood sample by digestion with nitric acid.The method although very reliable has the disadvantages of both a large sample requirement and a long processing time. Some involve simple dilution of the blood normally with a haemolysing agent and then introduction into the carbon furnace for ashing and subsequent atomisati~n.~s~ A problem encountered in the application of electrothermal atomisation to volatile analytes in samples containing appreciable amounts of organic matter is obtaining satisfactory separation of the signals due to ashing and atomisation. This separation is necessary to some extent even when using automatic background correction if reliable results are to be obtained.To overcome this problem it has been found necessary to apply various comparatively time-consuming techniques to remove organic matter either by pre-ashing6 or de-proteini~ing.~ In the matrix modification molybdenum tube coating procedure developed and adopted for the determination of lead in urine1s2 it was found that the addition of orthophosphoric acid was effective in raising the atomisation temperature of lead thereby allowing greater time resolu-tion of the ashing and atomising processes. The substitution of more “volatile” anions by orthophosphate or pyrophosphates which occurs in the furnace after the phosphoric acid treatment was also of benefit in minimising if not eliminating the interference of halide and sulphate salts.The effect of phosphoric acid on the atomisation temperature has been studied for a number of metak8 For lead the lowest temperature at which atomisation is detectable is increased from 630 to 840 “C in the presence of orthophosphoric acid. The role of molyb-denum as a coating for carbon tubes is not clear. Initially it was postulatedl that the coating played some part in modifying the matrix so that background absorption due to phosphate salts was reduced. However benefits in the use of molybdenum and other metal coated tubes have also been observed when using matrix modifiers other than phosphoric a~id.~-ll The over-all effect of metal-coated tubes appears to be in allowing more reproducible analyte absorption signals to be obtained from a wide variety of sample matrices.Many ETA - AAS methods for small volume blood samples have been published 814 HODGES AND SKELDING DETERMINATION OF LEAD IN Experimental Analyst VOZ. 108 In initial trials a simple 1 + 10 dilution of the blood with a 0.1% Triton X-100 solution was used.12 This gave a reasonably stable haemolysed suspension. However on analysis using standard instrumental conditions for lead repeatability was poor. An examination of back-ground absorption showed a high background peak which indicated that ashing was incom-plete before atomisation of the lead occurred. To improve ashing efficiency nitric acid was included in the Triton X-100 diluent. It was found that comparatively low concentrations of nitric acid caused precipitation in the diluted blood but a level of 1 yo could be tolerated.Fig. 1 shows absorbance veysus time charts for blood with the nitric acid - Triton X-100 diluent mixture. Furnace conditions were as follows dry 60 s at 95 "C; ash 20 s at 450 "C; and atomise 5 s at 1800 "C. A pyrolytic carbon tube was used. Although the background was lower than for blood with Triton X-100 alone there was still a sharp background peak of about 50% of the total signal. In formulating the diluent for urine1 a concentration of 2% orthophosphoric acid was chosen to give a lOOyo excess over normal levels of salts present in urine. By the same considerations for blood a concentration of 1% was accepted. No precipitation of the blood occurred with this mixture provided that the Triton X-100 was retained. I I I I I I I I 1 I I I -J 'I Ash Atomise Ash Atomise Time -+ Fig.1. Total and background absorbance signals obtained for blood diluted with 1 yo nitric acid - Triton X-100 diluent using conditions as given in the text. (A) Deuterium lamp; and (B) hollow-cathode lamp. Ash Atomise Ash Atomis' Time --+ Fig. 2. TotaI and background absorbance signals obtained for blood diluted with the phosphoric acid - Triton X-100 diluent using conditions as in the appended method. (A) Deuterium lamp; and (B) hollow-cathode lamp. Fig. 2 shows the absorption peaks obtained for blood diluted with the phosphoric acid - Triton X-100 diluent ; conditions were as given in the appended method. A molybdenum-coated tube was used. The background peak occurring during atomisation is much lower than that shown in Fig.1 for nitric acid - Triton X-100 diluent. TABLE I EFFECT OF NITRIC ACID- AND PHOSPHORIC ACID-BASED DILUENTS ON The ashing peak is much sharper and is complete well before atomisation. PRECISION AND SENSITIVITY WHEN USED WITH BLOOD OF KNOWN LEAD LEVELS Sensitivity for 1 yo absorption/ Precision standard deviation pg dl-l per pg dl-l A Level of lead in blood/ I \ r A 3 pg dl-l HNO H3P04 HNO %PO* 11 0.73 1.42 0.78 1.05 17 1.05 1.33 1.8 1.1 26 1.07 1.32 2.0 0.6 59 1.30 1.26 8.2 2.2 Mean 1.04 1.33 Relative s.d. 22.6% 4.95 July 1983 BLOOD BY AAS WITH ELECTROTHERMAL ATOMISATION 815 The effect of the two diluents on precision and sensitivity is shown in Table I. None of the blood samples used contained added lead.The lead values given in the table were the accepted means of analyses carried out in our own laboratory and also externally. The phosphoric acid treatment produces an improved performance with regard to both precision and consistency of sensi t ivi t y . It should be noted that as each blood sample was from a different physiological source they may be sufficiently different constitutionally so as to produce an effect that is not necessarily related to lead levels. These differences can give rise to two main effects changes in sensitiv-ity because of different proportions of constituents and changes in reproducibility due to variations in the total amount of matrix present. The second variable is probablymore important in determining the effectiveness of background compensation.Although the above treatment is an oversimplification and interaction of the two conditions with other factors obviously occurs their separate consideration is helpful in the interpretation of the results. Preservatives formation of microclots which are a potential source of error in blood lead analysis. of the sample is of additional benefit in ensuring complete haemolysis. We have found EDTA to be the best preservative. It is more effective in preventing the Freezing Graphite Tubes Our experience is limited to the use of pyrolytically coated tubes apart from an occasion when uncoated tubes were supplied by mistake and rather erratic and unreliable results were obtained. Dodecamolybdophosphoric acid was convenient to use for tube conditioning be-cause of its high solubility in aqueous solution.However ammonium molybdate was found to have a lower lead blank and was therefore used in the diluent to replace any molybdenum lost during analysis cycles. Calibration In our laboratory the following quality control system is in operation This could supple-ment or even take the place of the standard additions calibration given in the appended met hod. A stock of internal quality control standards is built up from analysed unadulterated samples. These samples must be analysed at least ten times within the laboratory over a period of days and preferably also analysed by exchange with external laboratories to give accepted mean levels. From this stock three levels are chosen from which to construct a calibration graph (or to calculate a least-squares regression line) making at least duplicate injections.This calibra-tion is carried out at the beginning of a batch of samples. After each fifth sample a control (internal or external) is analysed. The result obtained should not be in error by more than an amount given by Error = & [1.54444 - (0.004444 x mean level)] which was derived from the acceptable instrument difference in replication of 0.002 absorbance unit. If a control is outside these limits then the five sample results prior to this control must be rejected the source of the error corrected and then the five sample determinations repeated. Any series of analyses should end with a control sample. Loss of Organically Bound Lead In our lead in urine studies2 it was considered necessary to include an oxidative treatment with iodine in order to prevent loss of organically bound lead.It was felt that it would be difficult to incorporate a similar step into the lead in the blood method owing to the hetero-geneous nature of the sample which would make the procedure unnecessarily complicated. Lead in the blood is predominately associated with the cell fraction rather than theplasma unless cell rupture has taken place.13 Heard et aZ.14 have shown in alkyllead exposure experi-ments that for a period after exposure a significant fraction of the total lead is found in the plasma probably as the water-soluble metabolite trialkyllead. In in vitro experiments we have found that 75435% of the added lead is present in the plasma after equilibrating trialkyl 816 HODGES AND SKELDING DETERMINATION OF LEAD IN Analyst VoZ.108 lead chloride with whole blood for 3 h. The behaviour of added inorganic lead is different; Clarkson and Kench13 found virtually complete transfer of added lead chloride to erythrocytes. A set of samples from exposed workers was analysed by the proposed method for lead in whole blood and after centrifuging for lead in plasma. The results are shown in Table 11. TABLE I1 COMPARISON OF LEAD IN WHOLE BLOOD AND LEAD IN PLASMA Sample Packed cell 1 47 2 38 3 45 4 42 6 41 6 42 7 42 8 48 Mean 43 No. volume (Couler) Lead contentlpg dl-l - Whole blood Blood plasma 32 2.4 31 1.2 29 1.4 29 0.9 42 1.5 42 1.3 36 1.4 36 1.6 36 1.45 Lead in plasma % 7.5 3.9 4.8 2.3 3.6 3.1 4.0 4.2 4.2 Plasma samples 1 4 inclusive were also analysed by the method we used for urine which involves treatment with iodine.The results obtained were virtually identical with those given in Table I1 obtained by the proposed method. The low proportion of lead found in the plasma is in agreement with other studiesl59l6 where no obvious potential exposure to alkyllead existed. It was therefore concluded that no special treatment to prevent possible loss of volatile organically bound lead was necessary. It may also be tentatively inferred that exposure to alkyllead for the subjects examined was low. In the studies of the distribution of metals between erythrocytes and plasma it is important to consider the nature of any anticoagulant or preservative that may be present in the blood sample.Table I11 shows the results of an experiment in which a blood sample after being taken in a syringe containing no anticoagulant was split into three portions. To one portion was added the preservative commonly used for blood transfusion which contains citric acid, sodium citrate and dextrose. The second and third portions were treated with EDTA (potassium salt) and heparin respectively. After mixing on a roller mixer for periods of 6 and 24 h samples were taken and analysed for total and plasma lead. There is a marked effect on the lead distribution with both EDTA and citrate if the plasma separation is delayed. Heparin does not appear to have too serious an effect and its presence could be tolerated.Although K,EDTA was used for our plasma lead study its effect on lead distribution was minimised by rapid sample processing. TABLE I11 EFFECT OF VARIOUS BLOOD PRESERVATIVES ON THE LEAD CONTENT OF BLOOD PLASMA Lead contentlpg dl-1 6 h mixing 24 h mixing -A Preservative added Whole blood Plasma Whole blood Plasma Citrate 35 5 36 7 K,EDTA . . 36 9 36 16 Heparin . . 35 1 36 2 Results The proposed method was compared with that of Bambach and Burkey.2 The comparative study extended over a period of 3 months and involved 210 samples taken in the course of rou-tine medical examination of workers engaged in the manufacture of lead alkyls. The atomic-absorption method was carried out using Instrumentation Laboratory equipment with manua July 1983 BLOOD BY AAS WITH ELECTROTHERMAL ATOMISATION 817 injection of samples.The correlation coefficient obtained was 0.950 and the linear regression equation was BB = 0.932AAS + 2.64 where BB is the result obtained by Bambach and Burkey and AAS that obtained by the pro-posed atomic-absorption method. The instrument used for determination of lead in urine in our laboratories is the Varian 175B with the CRA 90 carbon tube furnace and the ASD 53 automatic injection system. To ensure flexibility in instrumentation in the event of breakdown or changes in work load it was desirable that the Varian and Instrumentation Laboratory equipment should be interchange-able. To this end a comparison was carried out between the two instrument systems using the proposed method.The correlation coefficient was 0.973 and the linear regression equation was This comparison study involved 151 samples. V = 0.9981L + 0.564 where I L is the result obtained with Instrumentation Laboratory equipment and V that obtained with Varian equipment. The repeatability of the method was tested by analysing nine blood samples using nine separate aliquots of each. Varian instrumentation was used and the results are given in Table IV. TABLE IV METHOD REPEATABILITY DATA Sample No. 1 2 3 4 5 6 7 8 9 Mean lead content/ pg dl-1 34.0 17.9 10.1 46.8 39.4 24.9 19.7 7.5 48.7 Standard deviation/ pg dl-l (n = 9) 1.63 0.93 0.66 2.06 2.39 1.17 1.03 0.47 1.16 From 1976 we have taken part in the UK National External Quality Assessment Scheme (NEQAS) for blood lead.Table V shows a random selection of results obtained by the appended method together with the mean result of the other laboratories participating in the scheme . TABLE V ANALYSIS OF NEQAS SAMPLES BY THE PROPOSED METHOD Lead contentlpg dl-l Mean of all This laboratory laboratories A I \ 34 34 47 50 74 67 66 52 63 63 56 52 36 36 28 30 10 12 42 40 62 62 Lead contentlpg dl-1 Mean of all This laboratory laboratories 84 82 66 66 48 63 82 82 68 66 31 32 60 63 61 63 46 46 A t \ Mean 52.1 52.0 Standard deviation 18.6 17. 818 HODGES AND SKELDING DETERMINATION OF LEAD IN An@hSf VOl. 108 The range of blood lead levels covered by the NEQAS samples is higher than we normally encounter.The data given in Fig. 3 more typically illustrate the range of blood lead values for samples handled in our laboratory during 1980. Fig. 3. Histogram showing levels of lead in blood. Samples taken over a period of one year during routine medical examination of employees. Conclusions The phosphoric acid molybdenum matrix modification technique previously reported for the determination of lead in urine has been successfully applied to the determination of lead in blood. In use over an extended period the method described had proved to be reliable and has exhibited consistently good performance as regards both precision and accuracy. The applicability of the method to samples from the work force in our particular industry has been established.Acknowledgements are made to Mr. F. G. Noden for helpful discussions and to The Associ-ated Octel Company Limited for permission to publish this paper. Appendix Sample Pre-treatment Clean the skin thoroughly. Take the sample in a plain syringe (Sabre 10-ml Luer) that contains no anticoagulant reagent. Transfer immediately into a sample tube (KE/4 Seward) containing EDTA close tightly and mix well (10 min on a roller mixer). If possible keep the sample tube at a temperature of approximately -10 "C for at least 2 h before analysis. Apparatus The following were used atomic-absorption spectrophotometer fitted with background correction and a carbon tube furnace (for the development work both an Instrumentation Laboratory IL 151 fitted with an IL 455 carbon tube and a Varian AA 175B fitted with CRA 90 tube furnace and ASD 53 auto sampler dispenser were used) ; a Denley Spiramix 5 roller mixer ; Gilson adjustable-volume and Eppendorf fixed-volume disposable-tip pipettes for transfer of samples and reagents; and 2-ml polystyrene sample cups with polythene over-caps for sample preparation.Clean prior to use in batches of approximately 200 as follows shake the cups and caps with 2.5% Decon 90 and then leave to soak overnight. After draining rinse with de-ionised water and wash with dilute nitric acid (1 + 4). Finally wash with de-ionised water until free from acid and dry in a drying cabinet. All glassware was of borosilicate glass. Clean with nitric acid before use. Reagents Aristar grade (BDH Chemicals).All reagents should be of at least AnalaR quality but the orthophosphoric acid should be o July 1983 BLOOD BY AAS WITH ELECTROTHERMAL ATOMISATION 819 Dissolve 1 g of dodecamolybdophosphoric acid in about 60 ml of de-ionised water transfer quantitatively into a 100-ml calibrated flask and make up to vol-ume with de-ionised water. Dissolve 0.25 g of ammonium molybdate in approximately 60 ml of de-ionised water then add 2.5 ml of orthophosphoric acid mix and transfer quantitatively into a 500-ml calibrated flask add 0.5 ml of Triton X-100 and make up to volume with de-ionised water. Dissolve 0.1599 g of lead(I1) nitrate in approximately 80 ml of de-ionised water; add 1 ml of nitric acid mix and transfer quantitatively into a 100-ml calibrated flask make up to volume with de-ionised water and shake to mix.This stock solution contains 1 mg ml-l of lead. Dilute standard lead solution. Transfer 125 pl of the stock solution into a 25-ml calibrated flask containing approximately 20 ml of de-ionised water. Make up to volume with de-ionised water and shake thoroughly to mix. This solution contains 5 pg ml-l of lead and should be made up daily. T%be conditioning solution. Blood diluent. Standard lead solution. Procedure Using a pyrolytically coated carbon tube and a lead hollow-cathode lamp set up the instru-mentation according to the manufacturer's instructions to give the following parameters. Spectrophotometer wavelength 283.3 nm ; background correction on; and scale expansion, x 5. Carbon tube furnace dry stage 60 s at 100 "C; ashing stage 30 s at 750 "C; and atomise 2-5 s at 2500 "C.With the Varian CRA 90 a threaded pyrolytically coated tube together with a 10-pl sample cam in the ASD 53 was used. Tube Conditioning solution. Carry out at least four injections and furnace heating cycles using 10 pl of tube conditioning Analysis of the Sample To one of two 2-ml sample cups transfer using a microlitre pipette 5 p1 of the dilute standard lead solution. This represents an addition of 25 pg dl-l of blood. Then to each cup add 100 p1 of the sample of well mixed whole blood and finally 1.0 ml of blood diluent. Close the cups with polythene caps and allow to mix on the Denley mixer for 10 min. Inject 10 pl of the diluted sample into the carbon tube furnace and initiate the furnace heating programme.Record at least dupli-cate readings of both the blood sample and the standard addition. Carry out a blank deter-mination on the diluent alone again recording at least duplicate readings. When a batch of sampies is being analysed it has been found that one calibration for every ten samples is sufficient. Allow the sample to thaw completely whilst mixing on the Denley mixer. NOTES-1. 2. 3. The amount of lead added should be changed from day to day within the range 25-30 pg dl-l to When the lead content of a sample is found to be greater than 50 pg dl-l a second determination Dilute 50 pi of blood with 1.0 ml of blood diluent and make a standard addition of 5 pl of dilute indicate changes in sensitivity and pipette errors. should be carried out a t greater dilution i.e.as follows: standard lead solution representing an addition of 50 pg dl-' of blood. 1. 2. 3. 4. 5. 6. 7. 8. 9. References Hodges D. J. Analyst 1977 103 66. Hodges D. J. and Skelding D. Analyst 1981 106 299. Bambach K. and Burkey R. E. Ind. Eng. Chem. 1942 14 904. Kubasik N. P. Volosin M. T. and Murray M. H. Clilz. Chem. 1972 18 410. Fernandez F. J. Clin. Chem. 1975 21 558. Garnys V. P. and Smythe L. E. Talanta 1975 22 881. Stoeppler M. Brandt K. and Rains T. C. Analyst 1978 103 714. Czobik E. J. and Matousek J. P. Talanta 1977 24 573. Manning D. C. and Slavin W. Anal. Chem. 1978 50 1234 820 HODGES AND SKELDING Poldoski J. E. Anal. Chem. 1980 52 1147. Halliday M. C. Houghton C. and Ottaway J. M. Anal. Chim. Acta 1980 119 67. “Clinical Applications of Atomic Absorption/Emission Spectroscopy,” Instrumentation Laboratory Clarkson T. W. and Kench J. E. Biochem. J. 1958 69 432. Heard M. J. Wells A. C. Newton D. and Chamberlain A. C. Proc. Int. Conf. Manugement and Control of Heavy Metals in the Environment London Sept. 1979 CEP Consultants Edinburgh 1980, p. 103. Copeland T. R. Christie J. H. Osteryoung R. A. and Skogerboe R. K. Anal. Chem. 1973 45, 2171. Cavalleri A. Minoia C. Pozzoli L. and Baruffini A. BY. J . I n d . Med. 1978 35 21. Inc. Wilmington MA 1972. Received March 31st 1982 Accepted February 1 lth 1983 10. 11. 12. 13. 14. 15. 16

ISSN:0003-2654    

DOI:10.1039/AN9830800813

出版商:RSC    

年代:1983

数据来源: RSC

摘要:

Analyst July 1983 Vol. 108 pp. 821-826 Selective Spectrophotometric Determination of 82 1 Trace Amounts of Iron with Di(2=pyridyl)-NN=di[(8= quinolyl)amino]methane Determination of Iron in Blood Serum R. Escobar Department of Analytical Chemistry Faculty of Chemistry University of Seville Seville-4 Spain and J. M. Can0 Pavon" Department of Analytical Chemistry Faculty of Sciences University of Malaga Malaga-4 Spain Di( 2-pyridy1)-NN-di[ (8-quinolyl)amino]methane (DPQAM) has been exami-ned to evaluate its usefulness as a sensitive and selective spectrophotometric reagent for iron. A green complex is formed which is extracted into chloro-form a t pH 2.9-5.4 in the presence of perchlorate and ascorbic acid (the molar absorptivity E = 1.14 x 104 1 mol-l cm-l a t 693 nm).The distribution ratio of this extraction is very favourable so that iron can be determined in the range 0.25-2.50 p.p.m. (E = 1.71 x lo5 1 mol-l cm-l). The effect of foreign ions has been studied and the method has been applied to the deter-mination of iron in blood serum with good results. Keywords Iron determination ; spectrophotometry ; di( Z-pyridyl)-NN-di-[(8-quinolyZ)amino]methane; blood serum DPQAM was first synthesised and studied by us1 DPQAM is obtained easily by the reac-tion of 8-aminoquinoline with 2,2'-dipyridyl ketone. Although Lions and co-workers2-* have reported the formation of Schiff bases in the reaction of 8-aminoquinoline with diverse pyri-dine aldehydes we have demonstrated that in this instance and when pyridine-2-carboxalde-hyde and 6-methylpyridine-2-carboxaldehyde are used the corresponding aminals were obtained instead of the Schiff bases.The spectroscopic features of the aminals formed in these reactions have been described for the elucidation of the molecular structure^.^ DPQAM is a selective reagent for iron(I1) and under suitable conditions the green 1:2 metal - ligand chelate formed can be used for the spectrophotometric determination of iron in aqueous solution and by solvent extraction in the presence of large amounts of foreign metal ions. In this paper a systematic study of the experimental variables involved in the formation of the iron(I1) - DPQAM chelate is reported as well as the extraction of the iron(I1) - DPQAM -perchlorate system into chloroform. A very selective photometric method for determining trace amounts of iron is proposed.The determination of small amounts of iron in blood serum is described. Experimental Apparatus A Pye Unicam SP 8000 spectrophotometer was used for recording spectra in the ultraviolet and visible regions of the spectrum and a Coleman 55 (digital) instrument was used for measure-ments at fixed wavelengths equipped with 1-cm glass or silica cells. A Philips PW 9408 pH meter with glass - calomel electrodes was used for pH measurements. Reagents All solutions were prepared with analytical-reagent grade chemicals. DPQAM reagent solution. * To whom correspondence should be addressed. Dissolve 0.1 g of DPQAM in about 50 ml of chloroform and dilute to 100 ml with chloroform. The solution was prepared fresh daily 822 ESCOBAR AND CANO-PAVON SELECTIVE SPECTROPHOTOMETRIC Analyst VOZ.108 Standard iron(II) solution 3.934 8 g 1-l. Dissolve ammonium iron(I1) sulphate hexahydrate in dilute sulphuric acid dilute to 1 1 (the final concentration of the acid being 0.1 N) and stan-dardise gravimetrically as Fe,O,. Dissolve 100.0 g of NaClO, previously dried at 110 "C in distilled water and dilute to 1 1 to obtain a 10% m/V solution (0.82 M). Dilute this solution as required. water add 25.0 ml of glacial acetic acid and dilute the mixture to 1 1. Procedure Iron determination To 10-150 ml of sample solution in a separating funnel containing between 2.5 and 25 pg of iron(I1) or -(III) add 1 ml of 5% m/V ascorbic acid solution and 5 ml of buffer solution and extract the mixture with one 10-ml volume of DPQAM solution in chloroform.Shake the funnel vigorously for 3 min allow the phases to separate and transfer the lower (organic) layer into a 25-ml flask containing anhydrous sodium sulphate. Measure the absorbance of the green chloroform extracts against water at 693 nm. The calibration graph is prepared by using standard solutions of iron(I1) treated in the same way. Iron determination in blood serum Transfer 2 ml of serum into a 10-ml conical centrifuge tube add 2 drops of thioglycollic acid and mix. Add mixing after each addition 2 ml of hydrochloric acid (2 N) and 0.5 ml of trichloroacetic acid (40%). Stir vigorously with a glass rod for about 45 s and centrifuge for 10 min at 3000 rev min-l. Measure 2 ml of the supernatant fluid by a bulp pipette into a separating funnel add 1 ml of 5% m/V ascorbic acid solution 1 ml of saturated sodium acetate solution 5 ml of buffer solution 3 ml of 10% sodium perchlorate solution and make up to 25 ml with distilled water.Extract the mixture with one 6-ml volume of DPQAM reagent solution in chloroform. Shake for 3 min allow the phases to separate for at least 10 min and dry with anhydrous sodium sulphate. To prepare the standard sample, transfer 2 ml of the standard iron solution containing 1 pg ml-l of iron into a 10-ml conical centrifuge tube and continue as described previously; the absorbance (Astandard) is measured. The amount of iron in 100 ml of blood serum is calculated according to the following equation : Prepare working standards by suitable dilution.Sodium Perchlorate solution 10% m/V. Ascorbic acid solution 5% m/V. Bufler solution 0.84 M pH 4.7. This solution was prepared fresh daily. Dissolve 56.0 g of sodium acetate trihydrate in distilled Measure the absorbance (Asample) at 693 nm. A sample Iron (pg per 100 ml) = 100 x A standard Results and Discussion Absorption Spectra The absorption spectrum of the green complex formed between iron(I1) and DPQAM in aqueous ethanolic medium is shown in Fig. l(a). This spectrum has maximum absorption at 685 nm. The green complex can be extracted in diverse organic solvents but chloroform is the most effective. Voluminous anions are needed for the extraction; the use of perchlorate is the most convenient. A spectrum of the complex extracted into chloroform [Fig.l(b)] shows an absorption maximum at 693 nm; this wavelength was used in all subsequent measurements of absorbance. Effect of pH The effect of pH on the formation of the iron complex and the extraction as the ionpairwith perchlorate has been examined in a series of samples containing 25 pg of iron(II) 1 ml of 0.5% ascorbic acid solution 3 ml of 10% sodium perchlorate solution and various volumes of hydrochloric acid or sodium hydroxide solution of different concentrations. The final volume of the aqueous phase was always 10 ml. A 10-ml volume of 0.1% DPQAM solution in chloro-form was used as the organic phase. Fig. 2 shows that the maximum absorbance was obtained in the pH range 2.9-5.4. A decrease in absorbance below 2.9 can be attributed to incomplete formation of the iron complex owing to protonation of the reagent J d y 1983 DETERMINATION OF TRACE AMOUNTS OF IRON IN BLOOD SERUM 823 0.6 8 2 m * 0.4 0 4 0.2 Wavelengthlnm Fig.1. Absorption spectra of green iron complex. Concentration of iron(II) 25 pg ml-l. A Green complex in aqueous ethanol a t pH 4.75; B green complex extracted into chloroform at pH 4.75; and C reagent blank. Effect of Reagent Concentration and Shaking Time Aliquots of solution in acetate buffer (total volume 10 ml) containing 25 pg of iron(II) were extracted with 10 ml of solutions of 0.0254.2% m/V DPQAM in chloroform. The shaking time was varied from 10 s to 5 min while the other variables were kept constant. The results are shown in Fig. 3. The extraction was quantitative with a reagent concentration of 0.1% m/V and remained constant with increasing concentration.A shaking time of 2 min was necessary for the complete extraction of iron although 3 min was used in all subsequent work. Effect of Perchlorate Concentration The extraction behaviour of iron(I1) on changing the counter anion is shown by plotting the percentage extraction of iron(I1) as a function of the concentration of the anion (Fig. 4). The order of extractability as a measure of half extraction is as follows C104-> I- > SCN-> Br- > NO3-. This sequence suggests that the enthalpy of hydration of the anion is the major factor governing the extractability of the iron(I1) - DPQAM chelate cation as the form of the ion pair. With strongly hydrated anions such as chloride and sulphate ions the iron(I1) chelate is scarcely extracted at all.This result is in good agreement with those obtained by Otomo and Kodamas with 8-picolylideneaminoquinoline and iron(II) by Takamatsu' with the tris( l,l0-phenanthroline)ruthenium(III) cation and by Biswas and Mande18 with surfactant cations. 0.5 0.4 s +! 2 Q 0.2 0.3 0 0.1 I I 2 4 6 8 10 12 PH Fig. 2. Influence of pH on formation of iron complex extracted into chloroform at 693 nm 824 ESCOBAR AND CANO-PAVON SELECTIVE SPECTROPHOTOMETRIC Analyst VoZ. 108 0.5 m r C n n " B - A m ,=I - I 8 0.4 -e n a C 0.3 -0.2 -I I 1 1 2 3 4 5 Ti me/m i n 0.1 Fig. 3. Effect of reagent concentration and shaking time on the formation and extraction of the iron complex a t 693 nm.Reagent concentra-tion A 0.025%; B 0.05%; and C 0.1%. Results obtained with a 0.2% reagent concentration are similar to those a t O.lyo and they are shown. Composition of Extracted Species An attempt was made to determine the composition of the extracted species by the con-tinuous variation technique modified accordingly for the application to a two-phase ~ y s t e m . ~ The results showed a stoicheiometric ratio of iron complex to perchlorate of 1 2 so that the composition of the extracted species is Fe (DPQAM) 2( C10,) 2. Determination of Iron( 11) Based on experimental work a method is proposed for the determination of trace amounts of iron involving the formation of a green complex with DPQAM and its extraction as the ion pair with perchlorate into chloroform.Beer's law is obeyed between 0.25 and 2.50 pg ml-l of iron. The molar absorptivity ( E ) at 693 nm is 1.14 x lo4 1 mol-l cm-l; however as the volume of the aqueous phase can be increased until the ratio Vaq/Vorg = 15 the minimum concentra--5 - 4 -3 -2 -1 0 + I Log [anion] Fig. 4. Extraction of iron(I1) as the DPQAM complex with various anions (25 pg of iron). A Clod-; B I-; C SCN-; D Br-; and E NO,-July 1983 DETERMINATION OF TRACE AMOUNTS OF IRON IN BLOOD SERUM 825 tion of iron that can be determined is 0.01 pg ml-l; the apparent molar absorptivity is there-fore 1.71 x lo5 1 mol-l cm-l. The precision was estimated from 11 results for 10-ml aliquots, with 2.50 pg ml-l of iron(I1). These results give a mean absorbance value of 0.488 with a standard deviation of the mean of 0.001 1 absorbance unit and a relative standard deviation of 0.53%.Effect of Foreign Ions A 10-ml volume of a solution containing various amounts of foreign ions up to 1000 pg and a fixed amount (25 pg) of iron(II1) was treated exactly as under Procedure. The results are given in Table I. This method is selective and the determination of iron is almost free from interferences. TABLE I EFFECT OF VARIOUS IONS ON THE DETERMINATION OF IRON BY THE RECOMMENDED PROCEDURE Iron concentration 2.50 p.p.m. Tolerance, Foreign ion p.p.m. Cd(II) Sb(III) Sn(II) Cr(III) Ni(II) V(V) Pb(I1). Mn(II) Au(III), Al(III) alkali metals alkaline earths chloride bromide iodide oxalate, Cu(II) Zn(I1) . . . . 60 EDTA * .6 thiocyanate tartrate and nitrate . . * . 100 C.o(I1) . . 10 Determination of Iron in Blood Serum In this determina-tion it is necessary to add 1 ml of saturated sodium acetate solution and 5 ml of buffer owing to the increase in acidity by the added acids. A 6-ml volume of reagent is preferable in pathological samples containing small amounts of iron. The results obtained with this reagent have been compared with the procedure of Trinder,lo which uses bathophenanthroline. The results are given in Table 11. Each result is the mean of three separate determinations. Iron in blood serum can be determined with DPQAM with good results. TABLE I1 DETERMINATION OF IRON IN BLOOD SERUM Fe found/pg per 100 ml of serum Sample No. 'With DPQAM With bathophenanthroline' 1 91 91 2 208 209 3 87 87 4 99 100 Conclusion The DPQAM method for iron determination is sensitive and selective owing to the easy extraction of the complex into chloroform and to the favourable distribution ratio in this solvent.It is a good procedure for the determination of small amounts of iron in which classical reagents cannot be used directly except with a previous pre-concentration procedure. This method can be used effectively for the evaluation of iron in blood serum when the con-centration of iron is small as occurs in some diseases such as certain types of anaemia 826 ESCOBAR AND CANO-PAVON 1. 2. 3. 4. 6. 6. 7. 8. 9. 10. References Escobar R. Cano-Pavon J. M. Bellanato J. Galvez E. and Pino F. Talanta 1982 29 135. Dwyer F. P. Gill N. S . Gyarfas E. C. and Lions F. J . Am. Chem. SOC. 1953 75 3834. Lions F. and Martin K. V. J . Am. Chem. SOC. 1957 79 2733. Goodwin H. and Lions F. J . Am. Chem. SOC. 1959 81 6415. Bellanato J . Galvez E. Escobar R. and Cano-Pavon J. M. Can. J . Chem. 1982 60 1775. Otomo M. and Kodama K. Bull. Chem. SOC. Jpn. 1975 48 906. Takamatsu T. Bull. Chem. SOC. Jpn. 1974 47 118. Biswas H. K. and Mandal B. M. Anal. Chem 1972 44 1636. Irving H. and Pierce T. B. J . Chem. SOC. 1959 2565. Trinder P. J. J . Clin. Pathol. 1956 9 170. Received January 17th 1983 Accepted February 21st 198

ISSN:0003-2654    

DOI:10.1039/AN9830800821

出版商:RSC    

年代:1983

数据来源: RSC

摘要:

Analyst J@y 1983 Vol. 108 pp. 827-834 Direct Determination of Inorganic Mercury in 827 Biological Materials after Alkali Digestion and Amalgamation Tetsuro Konishi Kumamoto Municipal Institute of Public Health 269 Tainoshima Tamukaemachi Kumamoto 862 Jaflan and Hitoshi Takahashi Deflartment of Pharmacology Institute for Medical Immunology Kumarnoto University Medical School, Kumamoto 860 Japan A reliable method is described for the determination of inorganic mercury in biological materials in the presence of organic mercury. This is principally based on the fact that hydrogen peroxide oxidatively liberates inorganic mercury from organic substances in strong alkali and reduces it to the metallic state without decomposing organic mercurials concomitantly present. The metallic mercury vaporised with a nitrogen stream is trapped by gold amalga-mation and then released for electrothermal atomisation atomic-absorption spectrometry.The detection limit is 1 ng of inorganic mercury and the co-efficient of variation for 40 ng of inorganic mercury is 2.8%. Keywords Inorganic mercury determination ; biological materials ; hydrogen peroxide reduction ; gold amalgamation ; atomic-absorption spectrometry The separate determination of inorganic from organic mercury compounds in biological materials is required for the study of the biotransformation of organic mercurials and for toxicity evaluation of any mercurials in foodstuffs. Generally the inorganic mercury in biomaterials can be determined by subtracting from the total mercury the amount of organic mercury present in the sample which will be measured gas chromatographica11y.l It is difficult however to determine accurately the amount of inorganic mercury present by this general method especially when the inorganic portion of mercury is relatively small.To date there have been only a few methods available for determining inorganic in the pres-ence of organic mercury. One utilises the 203Hg isotope-exchange reaction in undigested samples2 and the other flameless atomisation atomic-absorption spectrometry with tin( 11) chloride as a reducing reagent in acidic conditions.3-6 The former method however requires handling of the isotope which is not readily available and the latter involves some difficult procedures such as the conversion of organic into inorganic mercury even to an extremely small extent and incomplete volatilisation of mercury from biological samples.The principle of the approach described for mercury determination is based on the finding that in strong alkali hydrogen peroxide reduces inorganic mercury to vaporisable metallic mercury without any decomposition of accompanying organic mercury compounds in the sample. The vaporised mercury is then collected on a gold trap as amalgam under a nitrogen stream. The mercury is thereafter released by rapid heating for electrothermal atomisation atomic-absorption spectrometry (ETA-AAS). This method is as sensitive as other gold amalgam methods7-10 and minimises the influence of interferents. Mercury in the form of mercury(I1) sulphide which is difficult to convert to vaporisable mercury can be determined by this method.Experimental Apparatus and a Hitachi recorder Model 056 were used for mercury determination. determination is shown in Fig. 1. A Hiranuma mercury analyser Model Hg-1 with a 10-cm cylindrical glass cell of 1.8 cm i.d. The device for the The flow-rate of the carrier gas (nitrogen) was controlled a 828 KONISHI AND TAKAHASHI DIRECT DETERMINATION OF Analyst VoZ. 108 Cooling K J Fig. 1. Apparatus for the determination of mercury by electrothermal atomic-absorption spectrometry following gold amalgamation. (A) Carrier gas N,; (B) regulator; (C) reaction flask; (D) cooling tube; (E) gold trap; (F) furnace; (G) absorption cell; (H) flow meter; (I) mercury lamp; (J) detector; (K) recorder; and (L M) cold water.about 1 1 min-l by a pressure regulator. Tygon tubing was used to connect the vessels starting from the reaction flask cooling tube gold trap quartz cell and flow meter successively. A short-necked Kjeldahl flask (200 ml) fitted with a silicone-rubber stopper having two holes with two glass tubes one for the inlet tube with a glass-ball filter at the end and the other for the outlet was used as the reaction vessel. The flask was cooled in a cold water-bath during volatilisation of the mercury. The gold trap was quartz-glass tubing 30 cm in length and 1 cm in diameter in which gold-coated Celite was filled to approximately 1 cm depth with some quartz-wool at both ends. The gold-coated Celite was prepared according to the method by Anderson et aZ.7 The gold trap was surrounded by an electric furnace capable of increasing the temperature to about 700 "C within 30 s.The cooling tube was immersed in a cold water-bath to condense the water in the mercury-containing carrier gas before amalgamation. Reagents All chemicals used were of analytical-reagent grade and glass-distilled water was used for preparing the solutions. Gold-coated Celite. Gold(II1) chloride (2 g) was added to 15 g of acid-washed Celite-545 (30-60 mesh) (Johns-Manville) with sufficient water to make a slurry. The mixture was dried and ignited in a muffle furnace at 600 "C for 1 h. A stock solution of mercury (0.5 mg ml-l) was prepared by dissolving 0.6767 g of mercury(I1) chloride in 1 1 of 0.1 N sulphuric acid. This solution was diluted to a working mercury standard solution (1 pg ml-l) before use.Mercury(I1) sulphide solution (1 pg ml-l) was prepared by passing hydrogen sulphide gas into the working mercury standard solution. Methylmercury chloride ethylmercury chloride and phenyl-mercury acetate were recrystallised twice from methanol ethanol and benzene respectively. The standard solutions of these compounds (each 1 pg ml-1 of Hg) were prepared in the same way as described above. Sodium hydroxide solution 40% m/V. This solution was purged with nitrogen gas to elimin-ate the volatile mercury present. Hydrogen peroxide (mercury free) solution 30% V / V . Potassium cyanide solution 0.5% m/ V . Yeast extract solzttion 1% nz/V. Manganese sulphate solution 0.5% m/V. Silicone anti foam emulsion.Homogenates of biological samples were prepared with water by use of a homogeniser. Mercury(l1) standard solutions. Organic mercury solutions. Procedure In order to accelerate the oxidative liberation of mercury from the biological sample with hydro July 1983 INORGANIC MERCURY IN BIOLOGICAL MATERIALS 829 gen peroxide alkali digestion of the sample with a strong alkali was performed. An aliquot of the homogenate containing less than 0.1 pg of inorganic mercury was mixed in the reaction flask with 10 ml of 40% sodium hydroxide solution and 1 ml of 0.5% potassium cyanide solu-tion and the mixture was allowed to stand for 5 min. After diluting it to about 30 ml with water it was heated for 3 min on a steam-bath and cooled to room temperature by standing.The mixture was then diluted to 90 ml with water and 1 ml of 0.5% manganese sulphate solution 5 ml of 1% yeast extract solution and 1 drop of silicone antifoam emulsion were added. After adding carefully 15 ml of 30% hydrogen peroxide solution to the mixture the reaction flask was immediately connected to the apparatus and purged under stable continuous bubbling of nitrogen so as to vaporise the mercury. After 30 min all of the vaporised mercury was amalgamated on a gold trap; the mercury was then released from the trap by heating it to 700 "C in the furnace. The mercury vaporised again was determined by ETA-AAS at 253.7 nm. The peak height of the mercury reached a maximum within 30 s and a very sharp peak was obtained by pre-concentration on the gold trap.The calibration graph was prepared with 0 25 50 75 and 100 pl of the working mercury(I1) standard solution. In this instance, heating and standing were omitted in the process of alkali digestion to avoid volatilisation of mercury unbound to the protein. Loss of mercury owing to its adsorption on the new silicone-rubber stopper and Tygon tubing was remarkable at the beginning of their use but it became negligible after several assays. Usually the furnace was quickly cooled by air for the next application. Results and Discussion Hydrogen peroxide is a strong oxidising agent ; however it has also been reported that alkali salts of mercury are reduced to the metallic state with hydrogen peroxide.ll This method is based on the reaction in which hydrogen peroxide in strong alkali oxidatively liberates inorganic mercury from organic materials in the biological sample and at the same time reduces the mercury to the metallic state.The reaction depends on the activity of nascent oxygen released by decomposition of hydrogen peroxide. Manganese sulphate used in the reaction accelerates evolution of the oxygen and keeps the rate constant. However this reaction is not directly utilisable for the determination of inorganic mercury because the reduction of inor-ganic mercury with hydrogen peroxide takes a long time. Table I shows the effect of various accelerating reagents on the reduction. Kolbll reported that tartaric acid promotes the reduction but the yeast extract and glutathione tested here were more effective. Fig. 2 shows the effects of yeast extract on the time course of mercury volatilisation from various mercury solutions with hydrogen peroxide in alkali solution using the present appara-tus.The graph with tin(I1) chloride in acidic solution which is very similar to that reported TABLE I EFFECT OF VARIOUS ACCELERATING REAGENTS ON THE REDUCTION OF INORGANIC MERCURY BY HYDROGEN PEROXIDE Accelerating reagent Yeast extract-At . . Bt ,. ct Reduced glutathione . . Oxidised glutathione . . Cysteine . . Tartaric acid . . Fe (as FeSO,) . . . . Zn (as ZnSO,) . . None . . Amount of reagent added/mg 50 50 50 20 20 20 20 0.01 0.1 - No. of determinations 6 5 6 5 6 5 6 6 6 6 Mean absorbance & s.d.* 0.193 f 0.002 0.193 f 0.003 0.191 f 0.003 0.184 f 0.009 0.196 f 0.005 0.067 f 0.002 0.077 f 0.014 0.114 f 0.022 0.053 f 0.004 0.042 f 0.025 * Absorbance at 253.7 nm with 50 ng each of inorganic mercury.t Different commercial products 830 KONISHI AND TAKAHASHI ; DIRECT DETERMINATION OF Analyst Yd. 108 by Magos and Cernik,12 is also shown in Fig. 2(A) indicating complete volatilisation of mercury within a few minutes. With hydrogen peroxide Fig. 2(C) inorganic mercury was slowly vaporised in this system giving about half of the total mercury in 3 h. With yeast extract (B) added to the reaction mixture however almost all of the mercury vaporised within 20 min from the mercury standard solution and within 25 min from the liver homogenate (E). A difference in reactivity between commercial products of yeast extract was not observed.The yeast extract and glutathione probably act as the catalyst for the reducing reaction with hydrogen peroxide. In contrast to inorganic mercury organic mercury compounds with a C-Hg bond such as methylmercury chloride ethylmercury chloride and phenylmercury acetate were not reduced to metallic mercury with hydrogen peroxide (D). This indicates that alkaline hydrogen peroxide does not attack the C-Hg bond although acidic hydrogen peroxide is known to decompose organic to inorganic mercury.l3 Potassium cyanide added in alkali digestion changes the reducing sugar to its cyanohydrin. The reducing sugar glucose or galactose decomposes organic to inorganic mercury in strong alkali. The cyanohydrin of the reducing sugar does not account for this property.The amount of potassium cyanide used is sufficient to avoid the effect of reducing sugars present in ordinary specimens. With the standard mercury solution addition of 8 ml of 30% hydrogen peroxide solution was enough for the complete volatilisation of mercury whereas with the liver homogenate 12 ml of the hydrogen peroxide solution was required. Thus the amount required for actual specimens was decided to be 15 ml for safety. The flow-rate of the carrier gas nitrogen for mercury was set at 1 1 min-l which Nishi et aL9 suggested to be suitable for the gold amalgam method. This rate gave a sufficient reproducibility in the peak height. Table I1 shows absorbance values obtained from the determination of the standard solution containing from 0 to 120 ng of mercury.A good linear relationship was obtained with amounts of mercury below 100 ng; however this relationship tended to be lost above 100 ng. The blank value expressed as amounts of mercury obtained from the carrier gas and all the reagents used in analysis was 3.2 Assuming that the limit of detection is three times the value of the standard deviation of the blank value it is considered to be 1 ng. The results of ten measurements performed on 40 ng of mercury in the standard solution or 0.1 g of the kidney of the rat administered methylmercury chloride are shown in Table 111. The coefficients of variation obtained were 2.8 and 2.7% respectively. Fig. 3 shows effects of various amounts of hydrogen peroxide used in the reaction.0.33 ng. h 0 0 10 30 50 Ti me/m in Fig. 2. Effects of the yeast extract on the time course of mercury volatilisation from various mercury solutions. (A) 40 ng of inorganic mercury SnC1,; (B) 40 ng of inorganic mercury, H,O,; (C) 40ng of inorganic mercury H,O,, without yeast extract; (D) 40ng of organic mercury (methylmercury chloride ethylmercury chloride or phenylmercury acetate) H,O,; and (E) actual specimen 25 ng of inorganic mercury and 0.1 g of liver homogenate H,02. Each point represents separate analysis of inorganic mercury alone. I I I I 0 5 10 15 30% H202/mI Fig. 3. Effect of the amount of 30% hydrogen peroxide solu-tion on volatilisation. (A) 40 ng of inorganic mercury; and (B) 0.1 g of homogenised liver with 25 ng of inorganic mercury added July 1983 INORGANIC MERCURY IN BIOLOGICAL MATERIALS 831 TABLE I1 AVERAGE ABSORBANCE OF INORGANIC MERCURY STANDARDS Mercurylng Absorbance* Mercurylng Absorbance* 0 0.013 80 0.310 20 0.093 100 0.378 40 0.170 120 0.437 60 0.240 * The values are means of three determinations.Table IV shows the effect of interfering elements. Silver which is known to form amalgam with mercury affected volatilisation of mercury; iodide and bromide ions at high concentrations showed some interference. However a sulphur compound which was reported to interfere with the volatilisation of mercury in the tin(I1) chloride reduction did not show any interfer-ence in this method. The interference due to iodide and bromide however was not found with amounts up to 2000 times that of mercury to be determined.Thus it is considered that the interference by the above two ions is negligible for the actual analysis of biological materi-als. TABLE I11 REPRODUCIBILITY OF THE RECOVERY OF INORGANIC MERCURY FROM THE STANDARD SOLUTION AND RAT KIDNEY Absorbance* Mean f s.d. . . Coefficient of variation . . 40 ng of mercury 0.155 0.163 0.162 0.160 0.161 0.160 0.157 0.165 0.166 0.152 0.160 f 0.0044 2.8% 0.1 g of kidneyt . 0.200 0.207 0.208 0.199 0.204 0.199 0.1.95 0.198 0.202 0.190 0.200 f 0.0054 2.7% * All absorbance values were corrected with the blank. The figures given t The kidneys were obtained on the fourth experimental day from the rat were obtained by ten separate determinations on each of the samples.intravenously administered 1 pmol of methylmercury chloride. TABLE IV INFLUENCE OF ADDITION OF VARIOUS ELEMENTS ON INORGANIC MERCURY DETERMINATION Amount of element Interfering element added/mg Ag (as AgSO,) . . Cu (as CuSO,) . . As (as As,O,) Se (as Na,SeO,) . . Mn (as MnSO,) . . S (as Na,S,O,) . . EDTA (as disodium salt) . . ,. Br (as KBr) . . I (as KI) . . 0.1 10 10 10 10 10 10 10 1 0.1 10 1 0.1 Recovery %* 85 100 100 100 100 100 100 27 93 100 10 88 92 * These assays were carried out with 50 ng each of inorganic mercury 832 KONISHI AND TAKAHASHI ; DIRECT DETERMINATION OF AnaZyst VoZ. 108 TABLE V RECOVERY OF RADIOACTIVE MERCURY FROM 203Hg-CONTAINING MATERIALS BY THE WHOLE PROCEDURE OR AFTER ALKALI DIGESTION Recovery yo * f m Samplet Alkali digestion Whole procedure Liver .. 98 99 95 Kidney . . 96 98 93 f 5 (n = 6) * Radioactivity was measured by the use of a well-type scintillation t A rat intravenously injected with 50 pg of 203HgC1 was killed counter Aloka TDC-601. 4 h later. To check the reliability of the method rat tissues containing 203Hg were prepared. A rat injected intravenously with 50 pg of 203HgC12 was killed 4 h later. The liver and kidney were taken out and homogenised in water for sampling. All measurements of radioactivity were performed with a well-type sodium iodide scintillation counter. As it was reported that diluted pure mercury solution under alkaline conditions decreased its concentration slowly by v ~ l a t i l i s a t i o n ~ ~ ~ ~ ~ loss of mercury during alkali digestion was examined.However as shown in Table V almost all of the radioactivity remained in the solution after alkali digestion. Loss of mercury in alkaline solution seemed to be prevented by organic substances present in the sample. Table V also shows the recovery of 203Hg radioactivity through the whole procedure, including alkali digestion where the mercury vaporised from the sample was collected into 0.3% potassium permanganate - 1 N sulphuric acid solution instead of the gold trap. The recovery of radioactivity from the kidney homogenate was 93.5 5y0 in agreement with the following result of the recovery test using cold inorganic mercury. Table VI shows the recovery of mercury added to the specimens such as organs of untreated rats and fish meat.Amounts of 50 or 30 ng of mercury in an inorganic form including mercury(I1) sulphide added to 0.5 g of various tissue specimens were recovered to the extent of 92-103y0 by this method. On the other hand only 0.5% of organic mercury (1 pg of Hg), such as methylmercury chloride ethylmercury chloride or phenylmercury acetate which was added to liver specimens was recovered as inorganic mercury. A very small amount of inorganic mercury obtained may result from impurities in these organic mercurials which still remained after two repeated recrystallisations rather than to decomposition of organic mercury during the assay. In order to confirm that all of the inorganic mercury present in biological materials can be determined the mercury determination was performed on biological samples in which mercury was supposed to be present only as inorganic mercury.The result was compared with that of the total mercury determination conducted by combustion in quartz tubing TABLE VI RECOVERY OF INORGANIC MERCURY FROM BIOLOGICAL SAMPLES Inorganic mercury Biological sample present/ng Brain (0.5 g) . . 6 Liver (0.5 g) . . 13 Kidney (0.5 g) . . 49 Blood (1 ml) . . 4 Fish meat (0.5 g) 10 Liver (0.5 g) . . 13 13 13 13 * Methylmercury chloride. t Ethylmercury chloride. $ Phenylmercury acetate. Mercury f ound/ng 54 43 80 52 56 60 18 18 17 Recovery, % 96 100 103 96 92 94 0.5 0.5 0. Jub 1983 INORGANIC MERCURY IN BIOLOGICAL MATERIALS TABLE VII COMPARISON OF MERCURY VALUES IN RAT TISSUE DETERMINED BY THE PROPOSED METHOD AND COMBUSTION METHOD Mercury found f s.d./pg g-l 833 r 1 Proposed method Combustion method Liver .. . . 1.10 f 0.03 (8)t 1.13 f 0.08 (10) Kidney . . . . 19.66 f 1.01 (8) 19.01 f 0.73 (10) 4 h later. Sample* (inorganic mercury) (total mercury) * A rat was intravenously injected with 0.25 pmol of HgC1 and killed t Number of determinations indicated in parentheses. followed by the gold amalgam m e t h ~ d . ~ The samples were prepared as follows. A rat given no organic mercury was injected with 0.25 pmol of mercury(I1) chloride intravenously and killed 4 h later. The liver and kidney were removed and homogenised for the total and inorganic mercury determination.Table VII shows that the values of inorganic mercury as determined by the proposed method are in good agreement with that of the total mercury, indicating that all of the inorganic mercury in the biological sample can be determined by this met hod. The method was then applied to a study of the biotransformation of methylmercury in rats and inorganic mercury as yielded by biotransformation and present in tissues was directly determined in the presence of the remaining methylmercury. Six male Wistar rats weighing 180 g were administered 1 pmol of methylmercury chloride intravenously and killed on the second or on the tenth day after administration. The organs and faeces were analysed for inorganic and total mercury in order to obtain the ratio of the two.The results by the pro-posed method and that of Norseth and ClarksonlG for the same purpose in which inorganic mercury was determined by the isotope-exchange method are compared in Table VIII. The results from the two methods show reasonable agreement in the ratio of inorganic to total mercury in the faeces liver and kidney. The method using an isotope-exchange reaction has some difficulties because of the limited availability of the isotope and may not be accurate enough for determining small amounts of inorganic mercury formed in vitro from methyl-mercury in various tissues of animals. We believe that this method provides an adequate approach for the accurate determination of small amounts of biotransformed inorganic mercury in the presence of large amounts of organic mercury.The materials such as the brain of the rat injected with methylmercury or incubation mixtures containing methylmercury and various animal tissues could be efficiently analysed by this method. Further the organic portion of mercury in either the alkyl or aryl form, present in biological materials can be readily determined by this method by subtracting the value of inorganic mercury from that of total mercury in the specimens if the organic portion is not extremely small. TABLE VIII RELATIVE AMOUNT OF INORGANIC MERCURY IN ORGANS AND FAECES AFTER INJECTION OF 1 pmol PER RAT OF METHYLMERCURY CHLORIDE Inorganic mercury % * r 1 Proposed methodt Norseth and Clarkson methodl8t A 7- I Organ Second day Tenth day Second day Tenth day 2.5 3.4 0.4-4.0 0.4-4.0 Brain .. Liver . . . . 9.5 21.4 12 27 Kidney . . . . 11.6 36.4 20 41 Faeces. . . . 48.8 51.4 53 51 * Values are the means of determinations on three rats. Rats weighing 180 g were used in both of the experiments 834 KONISHI AND TAKAHASHI The authors are indebted to Emeritus Professor M. Uchida of Kumamoto University Medical School for his help in the preparation of this paper and many useful suggestions for its improve-ment. 1. 2. 3. 4. 5 . 6. 7. 8. 9. 10. 11. 12. 13. 14. 15. 16. References Westoo G. Acta Chem. Scand. 1968 22 2277. Norseth T. and Clarkson T. W. Biochem. Pharmacol. 1970 19 2775. Magos L. Analyst 1971 96 847. Gage J. C. and Warren J. M. Ann. Occup. Hyg. 1970 13 115. Umezaki Y. and Iwamoto K. Bunseki Kagaku 1971 20 173. Kamada T. Hayashi Y. Kumamaru T. and Yamamoto Y. Bunseki Kagaku 1973 22 1481. Anderson D. H. Evans J . H. Murphy J. J. and White W. W. Anal. Chern. 1971 43 1511. Head P. C. and Nicholson R. A. Analyst 1973 98 53. Nishi S. Horimoto Y. and Nakano N. Bunseki Kagaku 1974 23 386. Tanaka K. Fukaya K. and Fukui S. Eisei Kagaku 1974 20 344. Kolb A. Chern. Ztg. 1901 25 21. Magos L. and Cernik A. A. Br. J . Ind. Med. 1969 26 144. Baltisberger R. J. and Knudson C. L. Anal. Chim. Acta 1974 73 266. Shimomura S. Nishihara Y. and Tanase Y . Bunseki Kagaku 1968 17 1148. Toribara T. Y. Shields C. P. and Koval L. Talanta 1970 17 1025. Norseth T. and Clarkson T. W. Arch. Environ. Health 1970 21 717. Received December 21st 1982 Accepted January 26th 198

ISSN:0003-2654    

DOI:10.1039/AN9830800827

出版商:RSC    

年代:1983

数据来源: RSC

摘要:

Analyst July 1983 Vol. 108 $p. 835-839 835 Direct Determination of Selenium( IV) in Biological Samples by Cathodic-stripping Voltammetry Rahmalan bin Ahmad John 0. Hill and Robert J. Magee Defiartment of Inorganic and Analytical Chemistry La Trobe University Melbourne Victoria Australia 3083 The use of a commercially available digesting solvent Lumatom for the determination of selenium( IV) in biological samples using cathodic-stripping voltammetry is reported. It was found possible to determine selenium directly in biological samples containing metals such as arsenic( 111) copper(II), lead(II) iron(II1) and zinc(I1). The results for selenium in bovine liver and oyster tissue are reported. Keywords Selenium determination ; biological samples ; cathodic-stripping voltammetry The determination of selenium in biological samples is usually difficult involving the destruc-tion of organic matrices by acid digestion,l or oxygen plasma combustion,2 followed by separa-tion from interfering metal ions and pre-concentration techniques such as ion e~change,~ solvent extraction4 or hydride generati~n.~ Most of these techniques are laborious and time consuming and as amounts of selenium in biological samples are small there is the possibility of loss during the pre-concentration stage.This paper reports a simple method for digesting the biological samples for the direct determination of selenium( IV) using differential pulse stripping voltammetry. The digesting solvent consists of a tetraalkylammonium hydroxide in organic solvents. The digesting solvent used in the present work was the commercially available Lumatom.The determination of selenium( IV) by polarographic techniques generally involves the use of acidic supporting electrolytes with hydrochloric acid or perchloric acid being preferred.6 Christian et aZ.7 reported that the polarographic reduction of selenium( IV) in hydrochloric acid exhibits two main waves corresponding to the following reactions : Se(1V) + 6e + 2H+ -+ H,Se at E = -0.01 V versus S.C.E. followed by depolarisation of mercury by the reduction product : H,Se + Hg -f HgSe + 2H+ + 2e-The resulting reaction is Se(1V) + 4e + Hg -+ HgSe The second wave at E = -0.54 V versus S.C.E. corresponds to the further reduction of the HgSe to hydrogen selenide : HgSe + 2H+ + 2e -f H2Se + Hg Experimental Apparatus The stripping voltammetric analysis was performed on a PAR 174A Polarographic Analyzer connected to a Houston Omnigraphic 2000 X - Y recorder.A three-electrode system was used throughout the work with a saturated calomel electrode as the reference electrode. A platinum coil was used as auxiliary electrode and a Metrohm type E410 hanging mercury drop electrode as the working electrode 836 AHMAD et al. DIRECT DETERMINATION OF SELENIUM(IV) IN Analyst VoZ. 108 Reagents Distilled de-ionised water obtained from a Millipore filtration system was used in all experiments. Analytical-reagent grade methanol was redistilled in an all-glass apparatus before use. The nitric and hydrochloric acids used were of Aristar grade (BDH Chemicals).Lumatom was obtained from Hans Kurner Neuberg Germany. High-purity nitrogen was passed through a solution of vanadium(I1) chloride and then the supporting electrolyte solu-tion in methanol before reaching the test solution. Procedure About 50-70 mg of the dried biological sample was re-hydrated in a 160 x 16 mm Pyrex glass culture-tube fitted with a screw-cap by adding 150 p1 of de-ionised water for 10 min. Lumatom (1 ml) was then added and the tube capped to prevent solvent loss. The tube was shaken gently and partly submerged in a constant temperature (50 "C) water-bath for several hours or overnight. Occasional shaking facilitates more rapid digestion. When the sample was completely dissolved methanol was added and the solution acidified with hydrochloric acid.The solutions were filtered through a Whatman glass microfibre filter (4.7 cm diameter) and the filtrates collected in 25-cm3 calibrated flasks. Washing with methanol was carried out and the volume made up to 25 cm3. Filtration is required only to separate a white precipitate which separates out with some samples on addition of methanol. The final solution was transferred to the polarographic cell for analysis. The procedure for recording the voltammograms was as follows after de-aeration of the test solution (25 cm3) with pre-purified high-purity nitrogen for 10 min while stirring a mercury drop was dialled on the hanging mercury drop electrode (HMDE) . Pre-electrolysis was carried out at -0.05 V for a given period of time (e.g. 120 s) constant stirring being maintained.This was followed by a rest period of 30 s and the solution was cathodically polarised at a scan rate of 5 mV s-l. Quantification of the voltammograms was accomplished by the standard additions method and measurement of the peak heights of selenium at E = -0.47 V versus S.C.E. was achieved with a reproducibility of & 1 yo by this method. Results and Discussion Preliminary Investigations Lumatom gives a white precipitate with water and an emulsion with organic solvents such as dimethyl sulphox-ide or propylene carbonate. Methanol was found to be a suitable solvent and was used throughout the investigations. On carrying out cathodic-stripping voltammetry with nitric acid as the supporting electrolyte a well developed single peak occurred at E = -0.40 V versus S.C.E.However a plot of peak heights at this peak potential for increasing concentrations of selenium(1V) indicated a linear relationship only over a very small range of concentrations [0-15 p.p.b. (parts per log)]. Further investigations showed that the peak at E = -0.40 V (Fig. 1 peak A) is due to adsorption of the reduction products on to the mercury surface which caused current to flow without involving electron transfer.8 With increasing amounts of selenium(IV) the effect rapidly diminished owing to the formation of a layer of limiting thickness. On increasing the amounts of selenium(1V) and varying the deposition time a second peak was found to appear at E = -0.47 V (Fig. 1, peak B). Plots of the peak heights for various selenium(1V) concentrations indicated that the new peak corresponded to an electron-transfer process i.e.the reduction of selenium. How-ever because of the proximity of this peak to the adsorption peak it was not possible to develop a satisfactory analytical procedure. In contrast the use of hydrochloric acid as supporting electrolyte was found to be very satisfactory. Better resolution of the two peaks at lower concentrations of selenium(1V) was achieved using the peak occurring at E = -0.47 V (Fig. 1). The calibration graph was linear up to a concentration of at least 100 p.p.b. Using a deposition time of 2 min as little as 10 p.p.b. of selenium (IV) could be determined. The use of water as a solvent/diluent for Lumatom was not successful. Several acids were investigated as supporting electrolytes July 1983 BIOLOGICAL SAMPLES BY CATHODIC-STRIPPING VOLTAMMETRY 837 0.8 0.6 f .> 0.4 0.2 0 -0.4 -0.6 -0.8 EN versus S.C.E.Fig. 1. Diff erential-pulse cathodic strip-ping voltammograms of selenium (IV) record-ed a t various concentrations in 0.1 M HCl in methanol + 5% V/V Lumatom containing metal ions Fe(III) 540 p.p.b.; Cu(II) 386 p.p.b.; Zn(II) 260 p.p.b.; Pb(II) 0.6 p.p.b.; Cd(II) 0.6 p.p.b.; Ca(I1) 246 p.p.b.; Mg(II), 1210 p.p.b.; Mo(VI) 6.4 p.p.b.; and Co(II), 0.4 p.p.b. Conditions were as follows: HMDE 4 div. drop-l; temperature 20 "C; modulation amplitude 26 mV; deposition potential -0.05 V versus S.C.E.; salt bridge, 0.1 M HC1; deposition time 120 s with stirring and 30s without stimng; and scan rate 5 mV s-1.Effects of Various Parameters on Peak Heights Scan rate The scan rate had a marked influence on the selenium peak height. As the scan rate was increased from 1 mV s-1 the peak height increased. At 10 mV s-l the peak height was a maximum but the peak shape was distorted with a steep front edge and tailing of the back edge indicating a rapid reduction of selenium(0) to selenium(2-) at E = -0.47 V. A scan rate of 5 mV s-l was therefore selected for the analysis. Modulation amplitude from 5 to 100mV. selenium. The selenium peak height increased exponentially with increasing modulation amplitude A modulation amplitude of 25mV was used for the determination of Deposition potential As the deposition potential was shifted to more negative values from E = +O.lO V the voltammograms showed a region where the peak height was a maximum i.e.-0.05 to -0.15 V versus S.C.E. Shifting the deposition potential to more negative values reduced the peak height to a minimum at -0.30 V and beyond. Temperature The effect of temperature was investigated over the range 1040 "C. With selenium(IV) 838 Analyst VoZ. 108 there is an apparent transformation into an electroinactive state above 20 "C which may be due to increasing solubility of HgSe with temperature. AHMAD et aZ. DIRECT DETERMINATION OF SELENIUM(IV) IN Lumatom concentration Voltammograms were recorded for selenium( IV) after additions of increasing volumes of Lumatom. It was found that increasing amounts of Lumatom have a suppressant effect on the peak height of selenium(1V) up to 1% (V/V) after which the peak height tended to remain constant.Interferences A study was carried out to find metals that might interfere in the determination of selenium. Preliminary investigations indicated that the only elements likely to interfere were arsenic( 111) , copper(II) iron(III) lead(I1) and zinc(I1). Table I shows the results obtained in the inter-ference study. It will be seen that arsenic(II1) and copper(I1) strongly enhance the peak height of selenium while iron(II1) tends to have a suppressant effect. While the method was found to be satisfactory for samples containing small amounts of arsenic copper iron and zinc, it was decided that a better test would be the analysis of samples that contained selenium in the presence of a wide range of other metals.For this purpose two Standard Reference Materials (SRMs) were analysed. TABLE I EFFECT OF SOME METAL IONS ON THE SE(IV) PEAK HEIGHT Concentration ratio of Metal ion Se(1V) to Mn+ As(II1) . . 1 10 Cu(I1) . . 1 10 1 100 Fe(II1) . . 1 100 Pb(I1) . . 1 10 1 100 Zn(I1) . . 1 100 Te(1V) . . 1 10 Peak height recovery % 193 f 3 270 f 10 165 f 5 38 f 5 93 f 11 87 f 0 82 f 4 46 f 3 Determination of Selenium in Bovine Liver and Oyster Tissue The complex mixture of metals present in SRM 1577 bovine liver and SRM 1566 oyster tissue both of which contain selenium is shown in Table 11. The results of six representative individual analyses of the two samples are shown in Table 111. The average values indicate good agreement with the certified values.The use of tetraalkylammonium hydroxide-based digestion solvents such as Lumatom for the solution of biological samples followed by the direct determination of selenium using cathodic stripping voltammetry has proved to be satisfactory even in the analysis of biological samples containing complex mixtures of metals. However in the determination of selenium care TABLE I1 METAL CONTENT OF THE STANDARD REFERENCE MATERIAL BIOLOGICAL SAMPLES Dry masslpg g-' r 3 Metal Bovine liver Oyster tissue As . . 0.005 13.4 f 1.9 cu 193 f 10 63 f 3.5 Fe . . 270 f 20 196 f 34 Pb . . . . 0.34 f 0.08 0.48 f 0.04 Se . . . . 1.1 f 0.1 2.1 f 0.5 Te . . Zn . . 130 f 10 852 f 14 - Jzdy 1983 BIOLOGICAL SAMPLES BY CATHODIC-STRIPPING VOLTAMMETRY TABLE I11 RESULTS FOR THE DETERMINATION OF SELENIUM(IV) SRM 1577 bovine liver r 1 Sample size/mg Selenium found/pg g-l 66.04 1.01 42.82 1.05 69.29 1.34 97.62 1.64 100.57 1.39 152.95 1.24 Average 1.28 f 0.24 Certified value 1.1 f 0.1 SRM 1566 oyster tissue r \ Sample sizelmg Selenium found/pg g-1 A 77.77 2.10 78.73 2.70 70.52 2.03 72.94 2.23 72.64 2.34 55.37 2.17 Average 2.26 f 0.24 Certified value 2.1 f 0.5 839 must be taken to differentiate between adsorption and reduction peaks due to the complex electrochemical reduction mechanism.References 1. 2. 3. 4. 5. 6. 7. 8. Christian G. D. Knoblock E. C. and Purdy W. C . J . Assoc. 08. Anal. Chem. 1965 48 877. Forbes S. Bound G. P. and West T. S. Talanta 1979 26 473. Andrews R. W. and Johnson D. C . Anal. Chem. 1976 48 1056. Blades M. W. Dalziel J. A, and Elson C . M. J . Assoc. Ofl. Anal. Chem. 1976 59 1235. Dennis B. L. Mayers J. L. and Wilson G. S. Anal. Chem. 1976 48 1611. Shafiqul Alam A. M. Vittori O. and Porthault M. Anal. Chim. Acta 1976 87 437. Christian G. D. Knoblock E. C. and Purdy W. C. Anal. Chem. 1967 35 1128. Wopshall R. H. and Shain I. Anal. Chem. 1967 39 1514. Received November 15th 1982 Accepted January 31st 198

ISSN:0003-2654    

DOI:10.1039/AN9830800835

出版商:RSC    

年代:1983

数据来源: RSC