摘要:

Proceedings of the Society for Analytical Chemistry Analytical Division Chemical Society Proc. SOC. Analyt. Chem. Vol. 10 No. 8 Pages 189-226 CONTENTS Appointment of Managing Editor 189 Report of Meeting . . . . 189 The East Anglia Region . . . . 190 Summaries of Papers Annual Chemical Congress- Analytical Symposia A. “Physical Methods for An- B. “Newer Methods for En- vironmental Pollution An- Characterisation and Quality of Food and Agricultural Products With Special Reference t o EEC Requirements . . . 206 “Original Papers in Pharmaceuti- cal Analysis” . . . . . . 208 “Aspects of Chromatography” . . 210 alysis in the 70s” . . 191 alysis“ . . . . . 197 Obituary . . . . . . . 220 Silver Medal . . . . I 221 SAC Centenary Celebrations . . 222 Notices . . . . . . . 222 CS Autumn Meeting-Analytical Division Symposium .. . . 224 Papers Accepted for The Analyst 224 Forthcoming Meetings Back Cover August 1973 PAYCAL Vol. 10 No. 8 August 1973 PROCEEDINGS THE SOCIETY FOR ANALYTICAL CHEMISTRY ANALYTICAL DIVISION CHEMICAL SOCIETY OF Hon. Secretary W. H. C. Shaw Officers of The Society for Analytical Chemistry and the Analytical Division of The Chemical Society President G. W. C. Milner Hon. Treasurer Hon. Assistant Secretaries J. K. Foreman D. I. Coomber O.B.E.; D. W. Wilson Secretary Miss P. E. Hutchinson 9/10 SAVILE ROW LONDON WIX I A F Telephone 01 -734 9864 Editor J. B. Attrill Proceedings is published by The Society for Analytical Chemistry. Members subscriptions must be sent to The Chemical Society Non-members can be supplied with Proceedings only as part of a combined subscription with The Analyst and Analytical Abstracts Single copies can be obtained direct from The Chemical Society Publications Sales Office Blackhorse Road Letchworth Herts.SG6 1 HN (NOT through Trade Agents) price 30p post free. Remittances MUST accompany orders 0 The Society for Analytical Chemistry Assistant Editor P. C. Weston ANNUAL REPORTS ON ANALYTICAL ATOMIC SPECTROSCOPY Volume 2 1972 This comprehensive and critical report of developments in ana.;tical atomic spectroscopy has been compiled from more than 1000 reports received from world-wide correspondents who are internationally recognised authori- ties in the field and who constitute the Editorial Board. In addition to surveying developments throughout the world published in national or international journals a particular aim has been to include less widely accessible reports from local national and international symposia and conferences concerned with atomic spectroscopy. Volume 2 covers the year 1972 Published July 1973 21 6 pages Price f5.00 ISBN 0 85990 252 8 Obtainable from The Society for Analytical Chemistry (Book Department) 9/10 Savile Row London W1 X 1 AF Members of The Chemical Society may buy personal copies at the special price of €3.00

ISSN:0037-9697    

DOI:10.1039/SA97310FX027

出版商:RSC    

年代:1973

数据来源: RSC

摘要:

SOCIETY FOR .484LYTICAL CHEMISTRY ASALYTICAL DIVISION CHEMICAL SOCIETY Forthcoming Meetings September Monday 3rd to KADIocHEiIIcAL METHODS GROUP Symposium on “Liquid Scintillation Count- Thursday 6th ing.” BRIGHTOX Hotel Netropole Brighton. IVednesday 5th PARTICLE Size A N A L v s i s GKOITP on “l’articlc S i x .\iial!.sis in tlii Sub-inicru- “Size .Analysis of Pigments,” by \V. Carr. “Particle Size Rkasureiiient of Kon-uniform Laticcs I>>. ’1~url)idiinetr “Scanning and Transmission Electron Microscopes,” by J . RI. Creasy. “The IUPAC/SCIlXPL Surface Xrca Standards,” by I<. \ \ I s o n . The Polytechnic Hatfield; 11 a.m. MIDLASDS REGIOX on “Environmental Pollution.” Speakers K. I. Beynon P. B. Smith J . L). Butler and Al. I’. Smith. The University Edgbaston Birmingham; 11 a.m.HATFIELD mctre Range”. G. C. S. Cheesman. Tuesday 1 lth BIRMIXCHAM \Vednesday 12th ELECTROANALYTICAL GROUP Participation in Joint Anglo - Czcch Syniposium to Friday 14th on “Electrochemistry.” LOKDON Fviday 14th- Discussion on “Polarography 50 Tears On.” Speakers A. -4. I’lEek J . Yolke IT. F. Smyth G. C . Barker J . Tcnygl and R. Chelsea College ilIanresa Road London S.\V.3. JOINT YHAR>iAcErTIcAL X~ALYSIS GROL-P on “Cliallengc of 1:urmulation to Speakers A. Holbrook K. Lces and E. B. Reynolds. School of Pharmacy Brunswick Square London \TC 1 5 1AX. CHROW~TOGRAPHT AND ELECTROPHORESIS GROUP Discussion on “Chroiuato- Imperial Collegc Senior Chmmon Room Princes Gardens South Side I,ontlon Fleet. lhursday 13th LONDON the Pharmaceutical ~4nalyst.” Tuesday 18th LOXDOX graphic Analysis of Pyrolysates,” to he introduced b y TI.F. G. Pusey. London S.JV.7; 6.15 p.m. SCOTTISH KEGION jointly i,~itA the Scottish Region of thc Industrial Division of the CS on “The Importance of the Analyst to Managenlent (ivith Special Reference t o Changes Due to YI‘K Entry into the EEC).” Thursday 20th GLASGOI~ Speakers t o include C. IVhalley J . I<. Foreman and D. R. Deans. University of Strathclyde Glasgow ; 10 a.m. Thursday 20th ~IICROCHE~TICAL METHODS X T o i i I c SPECTROSCOPY .\UTOMATIC ~IIXHODS and Friday RADIOCHEXIICAL METHODS ELECTROAKALYTICAL and EULTCATIOS AKD 21st TRAIKISG GROUPS and the S o R T H \VEST REC;IOs on “Trace Analysis- SALFORD Different Approaches.” Speakers to includc L. S. Bark H. J . 11. Bowen I<. X . Chlmcrs J . B. Clegg 1:. T. Ewart J . H. S. Green I. I’raser J. Hislop I<. 1). Jee 11 ill. Poole -\. Ton-nshencl C. J . Little R. J . JlcCulloch J . 31. Ottau.ay T. B. Pierce B. Yeoman and J . D. \5’ilson. The University Salford. 3Iontlay 24th t o CS .Autumn Meeting. Symposiuin organisetl 1))- S.\C . \ I ) on “’111~ Charac- Thursday 27th SORWICH terisation of Materials.” The programmc appears 1111 1).224 of this issue. University of East Xnglia Sorwicli. Printed b y Heffers Printers Ltd Cambridge England

ISSN:0037-9697    

DOI:10.1039/SA97310BX029

出版商:RSC    

年代:1973

数据来源: RSC

摘要:

August 1973 Report of Meeting Vol. 10. No. 8 ORDINARY MEETING AN Ordinary Meeting of the SACjAD was held on Thursday and Friday July 12th and 1311 1973 in the Chemistry Department The University Sheffield. The Chair was taken by Mr. F. C. Shenton \’ice-President. The Meeting was devoted to short papers on “Research Topics in Analytical Chemistry,” and the following papers were presented and discussed “The Determination of Lead in Steel by Anodic Stripping Voltammetry,” by B. Jletters and B. G. Cooksey; “The Determina- tion of Nanograms of Indium Lead and Drugs of Forensic Importance by Enzymic Inhibi- tion,” by R. A. Sheikh; “Estimation of Mercury (both Inorganic and Organic Forms),” by P. Jones and G. Nickless; ‘Ton-dispersive Atomic Fluorescence Spectrometry,” by R. RI. Dagnall R. Mounce B. L. Sharp and T. S. \Vest; “Thin-layer Phosphorimetrp,” by L. G. Gifford ; “The Solution Chemistry of Simple Phenol-based Complexones,” by G. T. Murray and hl. A. Leonard; “Solvent Effects in the Catalytic Thermometric Titration of Weak Acids,” by R. Hargitt and E. J. Greenhow; “Polarography of 2-Ethylanthraquinone in Sulphuric Acid,” by S. R. Porter; “Digital Pulse Coulometry,” by D. R. \\:right; “Computer Analysis of Photoelectron Spectra,” by Rlrs. M. A. L$‘illiams; “Time Resolved Son-dispersive Atomic Fluorescence on a Carbon Rod,” by A. I;. King and T. S. \\yest; “Some Studies in Complexing Ion-exchange Resins,” by H. Eccles and 1;. Yernon; “Studies in the Solvent Extraction and Atomic Absorption Spectroscopy of the Xoble Metals,” by 11. A. Ashy. 189

ISSN:0037-9697    

DOI:10.1039/SA973100189b

出版商:RSC    

年代:1973

数据来源: RSC

摘要:

190 THE BAST ASGLIA REGION The East Anglia Region [Proc. SOC. Analyt. Chem. THE formation of an East Anglia Region (or Section as it was known then) had been suggested from time to time in the past and active steps towards this end were taken early in 2966. As a preliminary a circular was sent to all members of the Society for Anxlytical Chemistry residing in the 1l:ast Anglia area in order to ascertain the extent to which they might support a Section of their own-as at that time tlie area was included in the Midlands Section. More than half of those who were approached either expressed interest or promised support. At that time the boundaries of the Section were represented by a line drawn from Spalding to Peterborough down the A1 to Stevenage and finally curving round to Southend to include Hcrtford and Ware.The number of members residing in this area was ninety-three. The fields of activity included drugs foods fertilisers animal feeding stuffs plastics and clinical biological and chemical research. As a result of the positive response to the preliminary circular an Acting Committee was appointed and two trial meetings were held in order to assess the support-one meeting was held in Ipswich and the other in Cambridge. The response was sufficiently good to justify a formal application to Council for the formation of the Section and the application was approved unanimously at the meeting which took place on June 14tli 1967. The Inaugural Meeting was held on November 7th 1967 at tlie University of Essex Wivenlioe Park Colcliester. After the Section had been inaugurated formally by the Presi- dent o f the Society Mr.A. G. Jones talks were given by Dr. A. A. Srnales and Professor T. S. West. The first Chairman of tlie Section was IIr. C. E. Waterhouse the second Chairman Mr. N. R. Jones and the present Chairman is Mr. J. S. Leahy. The Honorary Secrctary/Treasurer -from tlie inauguration to tlic present day-is Mrs. I). Simpson. An inaugural dinner was held in the evening at Wivenlioe House. Mrs. 1). Simfison MY. J . S . Lealay (Honorary Secvetavy and Treasurer) (Chairman) MY. A. W . Hartley ( Vice-Clzaivman) Meetings on topics of interest are held three or four times a year in different parts of East Anglia. They usually take the form of afternoon or one-day meetings rather than evening meetings as travelling across East Anglia is not always easy particularly in the winter months. For this reason the Annual General Meeting is held in October rather than November December or January. Areas where East Anglia Kegion meetings have been held or are expected to be held include Colchester Huntingdon Ipswich Nonvich Cambridge Saffron Walden Hoddesdon Manningtree Hatfield and Bedford. The present membership which has increased since the formation of the new Chemical Society is now between 600 and 700.

ISSN:0037-9697    

DOI:10.1039/SA9731000190

出版商:RSC    

年代:1973

数据来源: RSC

摘要:

August 19731 PHYSICAL METHODS 1~012 ANALYSIS I N THE 70s 191 ANNUAL CHEMICAL CONGRESS-ANALYTICAL SYMPOSIA SWANSEA MARCH 2 7 ~ ~ TO ~ O T H 1973 THE Second Joint Annual Congress of tlie Chemical Society and the Royal Institutc of Cheni- istry was held at Swansea from March 27th to 30th 1973. Two Symposia were orgaiiised by the SA4C/Analytical Division and summaries of all of the papcrs are presented b e l o x i Analytical Symposium “A”-Physical Methods for Analysis in the 70s. ilrialytical Symposium “B”-Newer Methods for Environmental Pollution Analysis. Physical Methods for Analysis in the 70s Tlie following are summaries of the five papers presented at Analytical Symposium “A,” held on March ZYth 1973 during the Annual C.hemica1 Congress in Swansea. Fourier Transform Infrared Spectroscopy BY PETER R.GRIFFITHS (Depccrtiiieiit of Chernistvy Ohio Uwiueysit;y Athens Ohio 45701 US.A .) IKFRAKEI) spectrometers that use a monochromator for dispersing and arialysing radiation are inefficient because of the total energy that is emitted from thc sourcc on157 thcsmall resolution element that passes through the exit slit of the monochromator rcaclics the detector. The priiicipal advantage of using a Michclson interferornctcr for mcasuriiig infrared spectra is that all of the radiation that enters the aperture of the instrument is detected at all times during the measurement. Spectra can therefore be measured by an interfcrometer more quickly than if a monochromator is used by a factor equal to the number of resolution elcrncnts undcr study (Fellgctt’s advantage). The use of signal-averaging techniques allows the signal to noise ratio of interferonietrically measured spectra to be increased in proportion to the square root of the number of scans being averaged.The main disadvantage of interferometry is that the output from the detector is not the spectrum but a signal (called the interferogram) which is rclated to the spectrum by its Fourier transform. Hence rcsults have to bc collected in cligital forin and the spectrum is calculated with a computer. Modern Fourier transform spectrometers generally have a mini-computer (together with some forin of mass memory such as a disc) as an integral part of the instrument. In view of Fellgett’s advantage interferograms can be measured very rapidly and high- quality mid-infrared spectra can be obtained from interferograms measured in less than 1 s.This property has led to the interface of rapid-scanning interferometers with gas chromato- graphs for the rapid on-line identification of gas chromatographic peaks. Currently the limiting sample size for such measurements is about 10 pg for most compounds although i t is felt that the sensitivity will be increased in the near future. The detection limits can of coursc be reduced if the sample is trapped in the heated light-pipe gas cell through which the effluent gas from the gas chromatograph usually flows. At one time it was believed that the use of Fourier transform spectroscopy would allow the spectra of extremely small amounts of materials dispersed in potassium bromide micro-discs to be measured. However unless extremely non-volatile materials are being studied the sensitivity appears to be limited by the volatility of the sample rather than by the instru- mental signal to noise ratio.Two applications of Fourier transform spectroscopy in pollution analysis appear to be particularly promising. The first of these involves measuring the high-rcsolution infrared absorption spectrum of gases over very long path lengths (e.g. 400 m). The use of high resolution allows discrimination against absorption features due to coinmon atmospheric components such as water and carbon dioxide. When high-resolution spectra are measured with a grating spectrometer the slit width of the monochromator must be narrow so that the signal to noise ratio of thc spectra is low. I n Fourier transform spectroscopy Fellgett’s 192 [Pvoc. SOC.Annlyt. Chew. advantage enables high-resolution spectra to be rapidly measured over the complete mid- infrared spectrum with a sufficiently low noise level that absorption bands due to parts per lo9 amounts of pollutants and sometimes cvcn less can be rccogniscd. The second application is the measurement of the discrete infrared emission spectra of heated gaseous species as they emerge from smoke stacks. These gases have a low tempera- ture so that the amount of emitted energy collected at any measurement device is low. The use of Fourier transform spectroscopy will enable the composition of the effluent to bc nioni- tored remotely with the only restriction on the typc of molcculcs that can be detected being that they should have at least one infrared active vibration. In summary the use of Fourier transform spectroscopy (and possibly other similar multiplexing methods) should herald an increase in the use of infrared spectroscopy as an analytical technique.PHYSICAL METHODS FOR ANALYSIS I N THE 70s The Impact of Automation on Clinical Chemistry BY F. L. MITCHELL (Division of Clinical Chemistvy Cli$tical Research Cepzztve Hawow Middlesex H A 1 3 L,’J) ANALYTICAL chemistry covers a very wide area and niechanisation must be creeping into every branch but in none perhaps so much as in clinical chemistry where the necessity of circum- stance has made it imperative that we should mechanise and automate as quickly and com- pletely as possible. Modern medicine is fundarnentally based almost entirely on chemistry and it is not sur- prising therefore that with the recent and rapid increase in knowledge in the subject there has been almost an explosion in the demands made or1 hospital clinical chemistry departments.Since 1945 their output has becn doubling approximately every 4 years. Initially the increase in load was covered by simple expansion but prohibitive costs limited this solution and since the mid-1950s almost all of the increase has becn covcrcd by improved efficiency. Considerable improvements were possible by introducing work study and ergonomics together with aids such as automatic dispensers diluters and colorinieters with automatic sampling and printing of results directly in units of concentration. The major effect however came with the introduction of the Tcchnicon Mark 1 AutoAnalyzer in the late 1950s. In this instrument reagents are added and all reactions take place while the diluted specimen is flowing con- tinuously along a tube ending in the flow cell of a colorimeter.Later several streams were operated together using different assay techniques and the twelve-channel SMA 12/60 machine is now in fairly widespread use. It is planned next year to introduce a revolutionary forty- channel instrument again using continuous flow but computer-controlled and requiring only 600 pl of serum for all channels. Nost other companies interested in the field have concentratcd upon an alternative approach in which reactions take place in separate tubes. These machines tend to be more complex but have the advantage of more readily combining speed with precision. A wide range of small machines in which this principle is used is now available but the most sophisticated is perhaps the Amcrican Monitor Corporation’s Prograniachem 1040.This instrument has many refinements which have led it to be described as a “chemistry slot machine.” The insertion of a punched card instructs the instrument as to which of twenty available assays is required and supplies data for the assay such as which reagents to select and what incubation time is required. The machine then performs entirely auto- matically and finally prints the results. The larger discrete machines are typificd by the Vickers MC-300 which is the fastcst multi- channel instrument yet developed accepting specimens at the rate of 300 per hour and carrying out up to twenty assays on each. It is completely under the control of a PDP 8 computer. The measurement of enzymes is an increasing requirement and is best achieved kinetic- ally.Machines for this purpose were not easy to design but several are now available. Most of them usc relatively conventional techniques but the GeMS4EC system developed in the Oak Ridge National Laboratory Tennessee is unique. In machines in which this principle is used the reaction components are placed separately in tubes radiating in line from the centre of a centrifuge rotor each line of tubes constituting one channel. On rotation all of the components move by centrifugal force to a cuvette situated at the periphery of the rotor August 19731 PHYSICAL RlETHODS FOR ASALYSIS I?; THE 70s 193 where mixing takes place and the reaction commences. The optical absorbance is followed by light passing through each cuvette during rotation and on to a suitable detector the output of which is transmitted to a computer which calculates the reaction rate for each channel.Instruments in which this system is used have very high precision and very fast reactions can be followed but drawbacks at present lie in the cost of the sophisticated electronics required and the time necessary to stop reload and start the machine. Medical research has made enormous strides in recent years but unfortunately patients throughout the world who are in a position to take full advantage of this are few. The hospital laboratory forms one of the narrow channels whereby the fruits of this research can reach the patient ; modern technology can broaden this channel and although the techniques for achiev- ing this at present differ greatly between developed and underdeveloped countries it is the duty of all those concerned to ensure that this important aspect of the advancement of medi- cine is not neglected.Electron Spectroscopy for Chemical Analysis BY DAVID 31 HERCULES (Depnitnient of Ckcnzistvy ZInivevsity of Gtovgia Athens Geovgia 30602 l:.S.A .) ELECTROK spectroscopy for chemical analysis or ESCA as it has become known is a technique devised by Professor Iiai Siegbahn and his group at the University of Uppsala which is currently making a major impact on chemistry. ESC-4 shows considerable promise in its application to thc study of surfaces and catalysts and also as a tool for studying chemical structure and bonding. ESCA utilises soft X-rays to photo-eject electrons from a sample and the kinetic energy of these electrons is analysed under high resolution.By measuring the energy of photo-ejected electrons the binding energy of the electrons in the atoms can be determined. The utility of ESCX is derived from the chemical shifts observed as the electron density in the parent atoms is changed. For inorganic compounds this can be indicative of changes in oxidation state and for organic compounds the shift is a composite of changes in electron density and inductive effects of nearby groups. The instrumentation for studying ESCA utilises both electrostatic and magnetic kinetic energy analysing devices. -4 variety of commercial electrostatic instru- ments is available. A primary utilisation of ESCA by the analytical chemist will be in the study of species on surfaces.The sampling depth of photo-ejected electrons is approximately 2 nni and therefore ESCX lends itself well to surface studies. The detection of a small amount of tellurium oxide on otherwise highly purified elemental tellurium has been achieved. Although the elemental tellurium was over 99.9 per cent. pure the surface layer was contaminated with approximately 20 per cent. of tellurium oxide. Similarly adhesives can be applied to aluminium surfaces and stripped and the question arises whether or not under such circumstances a layer of adhesive remains. The adhesive studied contained silicon; after removal of the adhesive la!;er strong silicon peaks were observed in addition to the normal aluminium peaks which means that the adhesive layer is thin as both its peaks and the underlying aluminium surface were detected.It is also possible to follow reactions on glass surfaces that have organic functional groups attached. \T;hen organic groups are placed on a glass surface by silylating reagents these groups can be subsequently reacted and ESCA provides a convenient technique for following such reactions. For example we added an amino functional group to a glass surface and followed the resulting sulphonation by observing the appearance of the sulphur 2p peak. The classical structure of the bis(tripheny1phosphine)iminium cation (PPN) has been given as (C,H,)3P=N+=P(C,H,), containing charged nitrogen and uncharged phosphorus atoms. ESCA studies showed that the phosphorus atoms in these cations have binding energies identical with those of a series of phosphonium salts and that the nitrogen has a binding energy indicative of an electron density greater than that for an albylamine.These results are incompatible with the classical structure but can be explained on the basis of the “islands of charge” structure often proposed for phosphonitrilic trimers. On the basis of ESCA studies the structure (C,H,),-P+-I\’-P+(C,H,) has been proposed. Both primary and Auger electrons are observed in ESC,2. No adhesive layer can be detected by microscopy. ESCA is useful for studying charge distributions in chemical bonds. 194 [Proc. Sac. Andyt. Chem. A linear calibration graph based on calcium carbonate and silica mixtures has been used to monitor the calcium concentration on a glass surface and changes observed due to the diffusion of calcium to the surface when the glass is heated.Similarly we have studied the quantitative determination of molybdenum oxide mixtures (Moo - Moo,) ranging between 0 and 100 per cent. This is a particularly interesting problem as the MOO surface is contaminated with MOO,. Precision was good at Moo concentrations greater than 15 per cent. We have also shown that ESCA can be used for trace-metal analysis by coating fibre-glass surfaces with chelating agents and using these to scavenge trace-metal ions from solution. The use of these “chelating glasses” has shown a sensitivity to lead of about 20 parts per lo9 using a 100-in1 sample. Calibration of ESCA spectra both to determine binding energies and to compensate for surface-charging effects is absolutely essential. Traditionally the gold standard has been used.However we have observed that vacuuni-deposited gold on potassium cyanide sodium chloride and copper phthalocyanine gave rise to chemical reactions that caused shifts in tlie gold spectrum depending upon the temperature and the history of the sample. Hence it is imperative to USC more than one calibrant when measuring BSCA spectra. PHYSICAL METHODS FOR ANALYSIS I N THE 70s We have investigatcd the use of ESCA in quantitative analysis. Auger Spectroscopy BY C. R. BRUXDLE THE auger process is one of two mechanisms by which a positive ion in a highly excited state ( i ~ with an electron vacancy in one of its inner-core atomic-like orbitals) may decay to a less excited state. In the Auger process the inner-core hole is filled by the transition to it of an electron from one of the less tightly bound orbitals.This transition releases an amount of energy approximately cqual to the difference in energy levels of the two orbitals concerned which is sufficient to eject another electron (the Auger electron) from one of tlie less tightly bound orbitals with some excess kinetic cnergy; the resultant state is therefore a doubly charged ion. The phenomenon was first experimentally observed by Pierre Auger1 in 1925. As the kinetic energy of the Auger electron depends only on the energies of the levels involved in the transitions which are in turn characteristic of the atom involved the recording of the electron kinetic energy spectrum from a sample (Auger spectroscopy) provides a direct identification of the atoms present. There will in general be several Auger processes and hence ejected electron kinetic energies for an atom the number increasing as the atomic number and hence the number of filled atomic levels increases.Those which involve the valence molecular orbitals as the less tightly bound orbitals provide in theory at least the additional possibility of a molecular identification as the precise energy pattern of the molecular orbitals is characteristic of the molecule in its entirety and not an individual atom. (The initial inner-core hole is still an atomic property however so tlie general kinetic energy region of the Auger electron is still characteristic of the atom.) In practice for gaseous samples the complexity of the Auger spectra involving the molecular orbitals turns such a molecular analysis into a “fingerprinting” process.As all molecular orbitals fall within approximately thc samc binding energy span (I) to 30 eV) there is almost complete overlap of the “fingerprint” from the same atom in different molecules,2 thus making the use of the technique for quantitative analytical identi- fication in gaseous mixtures unlikely except possibly for very simple molecules. Its use mainly for quantitativc atomic analysis in conjunction perhaps with a gas chromatograph should be investigated further. Sensitivity is likely to be greater than for ESCA since a high concentration of initial core holes can be made by impact with high-intensity high-energy (above 3 kV) electron beams rather than the monochromatic X-rays neccssary for For solids the molecular orbitals largely coalesce into band structure and molecular identification becomes even more remote.The vast majority of published work in Auger spectroscopy concerns its use in semi-quantitative atomic identification of species present at or near tlie surface of solids (a problem which is tremendously difficult by any technique) mainly for metals metal oxides and semiconductors because when applied to solids the technique is highly surfacc sensitive the Auger electrons escaping only from the first few atomic laycrs of This effect is accentuated for metals for which the mean free path (School of Studies in Chemistry Unioersity of Bradjord Bradford Yoikshive BD7 IDP) The other more generally known mechanism is X-ray fluorescence. August 19731 PHYSICAL METHODS FOR AYALYSIS IN THE 70s 195 lengths for inelastic scattering of the electrons is very short (a few Angstroms).The controlling factor in deciding just what depth is examined is the energy of the escaping Auger electron (the mean free path length varies with energy) a factor which is equally true for ESCA.4 Auger spectroscopy has been used extensively for surface analysis while ESCA has not (until recently&+) because of (a) the interests of the people developing each technique and ( b ) the greater speed of Auger spectroscopy over ESCA for monitoring small surface concentrations because of the higher electron beam currents available. Accurate figures are hard to obtain and depend critically on the sophistication of the instrumentation used. The limits of detect- ion using commercially available cylindrical mirror analysers’ seem to be ol the order of 0.1 per cent.of a monolayer of carbon or oxygen atoms in recording times of about 20 s per atomic species. Oscilloscope techniquess can be used when much greater coverages (or distribution through the first few monolayers) are concerned allowing the continuous monitoring of surface reactions. Examples of the use of the technique include the evaluation of chemical and in. i~~c,z.co cleaning procedures for thin films single crystals etc. ; the study of photoemissive surfaces9 (which combination of elements in what proportions give the best results) ; numer- ous applications in metallurgy relating to grain-boundary segregation and fracturing8 ; electroplating arid the strength of deposited metal - metal bonds1”; semiconductor technology in which surface control may be critical; chernisorption studies; and basic studies of hetero- geneous catalysis.ll The technique of stripping off surface layers by ion bombardment while rnonitoring the surface composition by Auger spectroscopy adds considerably to the infor- mation by providing a depth profile of the elements present.As the core levels of atoms which undergo small “chemical shifts” in their energies on change of their bonding environment (cf. ESCk3) are involved chemical shifts are to be expected in Auger spectroscopy. These shifts may help identify the bonding characteristics of the atoms concerned. They are not nearly so often observed or studied as in ESCA because (a) the emphasis has been on rapid atomic identification ( b ) the line widths are often large compared with the shifts expected (c) interpretation may be difficult when three energy levels are involved (cf.m e l e d for ESCA) and ( d ) inany of the transitions in the surface-sensitive electron cncrgy range (20 to 1000 eV)4 involve valence-band electrons which do not undergo true chemical shifts.4 Several examples of the use of chemical shifts in surface analysis have been reported however,l2 arid in a few instances13 the shifts observed are far larger than the corresponding ESCA shifts a fact which may be very useful analytically e.g. copper(I1) oside and copper zinc oxide and zinc phases can easily bc distinguished when present together.13 REFEREXCES 1 . > -. 3. 4. ,). f i . 8. 9. 10. 11. 12. 13. r .\uger P. .T. Phys. Radium 192.5 6 205. 11odtleman. 1.T‘. E. Csrlson T. A . Krause M. (I. Pnllen R. P. Hull R’. E.and Schweitzer G. I<. 1. Chem. Phvs. 1971 55 2317. HGcules D. P&K. Soc. Aftalyt. Chem. 1973 10 193. Brundle C. R. aa Roberts M. W. and Thomas J . lI. Editoi*s “Surface and Defect Properties o1 Solids,” \*olnnie 1 Chemical Society Specialist Periodical Report Chemical Society London 1972. Urundlc C. R. and Rolierts ILL. W. Pvoc. R. Soc. d 1972 331 383. ~~ Chem. Phys. Lett. 1973 18 380. Bishop H. E. Coad. J. F. and Riviere J . C. J . Elrctr. Spec. 1973. 1 389. Palmherg P. W. Bohn G. I<. and Tracy- J. C. Appl. Phys. L r t t . 1969 15 254. Wcber H. E. Solid St. Tmiznol. 1970 December 49. Marcus H. I.. Waldrop J . R. Schuler F. T. and Cain E. F. C. J . EZeclroclit~t,z. Soc. 1972 119 1 :148 _.__I Uonrel H. P. Suvface Sci. 1971 27 387. Haas T. W. Grant J. T. and Uooley G. J. J .Appl. Phys. 1972 43 1853 Schiin G. J. Electv. Spec. to be published. Scanning Electron Microscopy BY R. W. WILLIAMS THE scanning electron microscope is an analytical tool and imaging device that has developed out of the needs and limitations found in light optical microscopy transmission electron microscopy and electron-probe microanalysis. Its primary- function is to provide higli-quality (UP Reseuvch Centw Sainbzir~,~oir-Thnnzes Middlesex) 196 [Proc. SOC. Annlyt. Chew,. images of solid specimens and commercial instruments in general have a magnification range of 20 to 100 000 x with a resolution dependent on the sample and mode of operation of 10 to 200 nm. The advantages of the scanning electron microscope are ease of operation and sample preparation ; large specimen chamber which facilitates dynamic experiments ; the means of direct observation of surface detail on bulk specimens with the ability to zoom in on fine detail; and a large depth of focus so that images in general display the three- dimensional nature of the specimen.Tlie images are produced by scanning an area of the specimen with a very fine electron beam which can have a spot diameter as small as 10 nm at the specimen surface. The high- energy electron beam interacts with the specimen to give rise to several simultaneously occurring physical phenomena any of which can be used to produce a signal on a cathode-ray display tube scanning in synchronism with tlie electron beam. There is thus an exact point-to-point correspondence between the raster on the specimen surlace and the raster on tlie cathode-ray tube and the brightness of each point on the display tube is a direct measure of the intensity of the signal obtained from the corresponding point on the specimen surface.Consequently the picture build-up on the display tube is some kind of image of the specimen surface dependent on the nature of the physical phcnomeria used to produce the signal. The magnification is determined by the ratio of the raster length on the clisplay tube which is constant to that on the specimen surface which can be varied. For example if the image on the display screen is 10 cm across then a useful range of magnifications between 20 and 26 000 x is obtained corresponding to scanned areas of side 5000 pm and 4 p i square on the specimen. A further reduction of the scanned area to obtain greater magnification although possible will not generally reveal finer detail owing to the limit imposed by resolution.There are three main prerequisites for specimens to be examined in the scaririirig electron microscope they must be electrically conductive or rendered conductive by coating with a thin layer of metal or carbon; they must withstand a vacuuni of 10-5 nini of mercury or lower; and they must be rigidly supported so as to minimise vibrations. The physical phenomena that result from the interaction of the incident electron bcam and the specimen and which are used to provide the image signal are elastically scattered incident electrons secondary electron emission X-ray emission cathodoluminescence and beam-induced currents and voltages. The variety of information signals available on the scanning electron microscope has increased its scope such that its applications are too numerous to detail here. For a detailed account ol the scanning electron microscope and its applications readers are therefore referred to the “Proceedings of tlie Symposium on the Scanning Electron Microscope The Instrument and its Applications,” and reference may also be made to several books.’-8 PHYSICAL METHODS FOR ANALYSIS IN THE 70s REFERENCES 1. Oatley C. \V. “The Scanning Electron Microscope. Part 1. The Instrument,” Cambridge L7niver- 3. Thornton Y. lL “Scanning Elcctron Microscopy (Application to Materials and Dcvice Science) ,” 3. Hcarle J. \\’. S. Sparrow J . T. and Cross P. M. “Thc Usc of thc Scanning Electron BIicroscope,” sity Press Cambridge 1972. Chapman and Hall London 1‘368. Pergainon Yress Oxford 1972.

ISSN:0037-9697    

DOI:10.1039/SA9731000191

出版商:RSC    

年代:1973

数据来源: RSC

摘要:

ALlgUSt 19731 XEWER METHODS FOR ENVIRONMEKTAL POLLUTION ANALYSIS 197 Newer Methods for Environmental Pollution Analysis The following are summaries of the ten papers presented a t Analytical Symposium “B,” held on March 28th 1973 during the Annual Chemical Congress in Swansea. A Microwave Plasma Detector System for the Measurement of Trace Levels of Carbon Monoxide in Air BY J. hfOOftr; AND T. s. \\’EST (Deflavtna~nt of Chemistvy Iwzperial College /.on,do7z SW7 2.4 Y ) AND R. M. DAGNALL (Huntiizgdon Research Centre Hzintingdon PIS18 GES) A RIETEI~D was described for the adaptation of the classical iodine pentoxide method for the deterinination of oxygen in organic compounds in order to permit tlie liberated iodine to be determined in a microwave emissive detector system by its atomic emission at 206.1 nni and thus provide a procedure for the determination of carbon monoxide- The niicrowave plasma is a non-LTE (local thermodynamic equilibrium) system exhibiting high (5500 OK) electronic and low (1300 ”I<) thermal temperaturcs.Power requirements for such a system are low about 15 to 200 W when it is coupled via a &wave resonant cavity. This system was explored because other methods lacked either the sensitivity or selectivity required. The optimal input microwave power was found to be 7<5 \V when the cavity and 2 mni i d . quartz tubing containing the plasma were not cooled. However the use of nitrogen as a coolant gas enabled input powers of 100 W to be used effectively. The iodine pentoxidc contained in the same length of quartz tubing as the plasma was heated to 165 “C.The plasma was supported in nitrogen for nitrogen - carbon niorioxide mixtures and thus operated under vacuum. The pressure drop between the sampling valve and the plasma was 15 to 30 torr. A liquid nitrogen cold trap was placed just before the plasma in order to remove chemical interferents particularly hydrocarbons. For a one-shot system a straight-line calibration graph was obtained in the concentration range 0.1 to 1.0 per cent. For a continuously operating system the quartz tubing containing the iodine pentoxide was increased in size to 8 mni i.d. and molecular sieve material was included in order to absorb the simultaneously evolved carbon dioxide which has a large electronic cross-section and would quench the iodine signal. At high concentrations (041 per cent.of carbon monoxide) the reagent appeared to undergo a two-stage reaction involving saturation of the reagent. However a t lower concentrations this effect disappeared and straight-line calibration graphs were obtained in the concentration range 1 to 80 v.p.m. This method should therefore be useful in the continuous on-line analysis of low concen- trations of carbon monoxide mixtures. We are grateful to the British Oxygen Company Limited for providing gas samples and a research grant for the project. .. .. (1) 5 C 0 + 1 2 0 + 5COZ $- 1 .. Methods of Analysis for Traffic Pollutants BY R. G. DERWENT AND H. N. M. STEWART As in all other fields of analytical chemistry the measurement of the local concentration of traffic pollutants in the air we breathe is moving from manual to instrumental methods.4 trace component of a gas can usually be measured by selective absorption in the liquid or solid phase but this approach suffers from a number of drawbacks for traffic pollution studies. Sampling is carried out over a finite period and whcn the inherent sensitivity is low this period might be extended over several hours until tlie detail of the behaviour of the pollu- tant build-up and decay is lost in this integrated sample. If the measurement in solution (Wavren Spring Laboratory Gunnels Wood Road Stcoenage Ilrvtfordshive 2 G l 2 B S ) 198 NEWER METHODS FOR ENVIRONRlENTA4L POLLUTIOK -4NALYSIS I Y T O C . SOC. /t?dJf. ChelPZ. is completed manually numbers of values are limited and the manpower has to be available on schedule. Determinations in solution of gaseous pollutants can be made autoinatically on either an integrated or a continuous basis e.g.the coulometric determination of sulphur dioxide and the colorimetric determination of nitrogen dioxide have been carried out automatically. These methods liave been widely used in the U.S.A. They are suitable for a long-term study and the detail they provide is dependent on the time constant of the system as a whole but this can bc almost instantaneous. In general it is difficult to make such methods specific for one pollutant. Dependencc on basicity or acidity or on oxidising or reducing propcrtics of a pollutant must inevitably lead to adverse effects from pollutants of the same or possibly opposing characteristics. Most satislactory are methods that are based on a specific physical property or gas-phase reaction of the pollutant.Fortunately a number of such methods are available which are rapid and specific and which will provide a continuous output to any data acquisition system chosen. The best established of these methods is non-dispersive irilrared analysis ol air for carbon monoxide. The analysis of air for total hydrocarbon content is best achieved with a flame-ionisation detector. The natural background concentration of nietliaiie ( 1 4 p.p.ni.) can be allowed for. For a more specific study a chromatographic separation is appropriate and carbon monoxide can be determined simultaneously. For determining sulphur dioxide which is produced chiefly by diesel vehicles insofar as traffic is concerned the most satisfactory method is flame photometry. We have carried out a comparison of the acid titration method with the coulometric method and the flame photometric method on the same site.For the determination of nitric oxide and nitrogen dioxide the method of Saltzmaii has found widespread acceptance but unfortunately this method is subject to practical difficulties. The absorption of nitrogen dioxide is not easily carried out efficiently. The production of nitrite is not stoicheiometric and the conversion of nitric oxide into nitrogen dioxide is not quantitative. This method has now been superseded by the clieniiluiiiiiiescerit method for determining oxides of nitrogen as the method of choice. The cherniluminesceIit Irietliod for determining nitric oxide is based on the reaction be- tween ozone and nitric oxide to produce nitrogen dioxide.Light at a waveleiigth of about 500 nni is emitted which is quantitatively related i o the nitric oxide concentration. The light emitted is measured photometrically. The method is specific for nitric oxide and by prior reduction nitrogen dioxide can also be dcterniined in the same instrument. For the calibration of such an instrument the Saltzman method was used in an indirect method which avoids the problems of inefficient absorption and oxidation of nitric oxide to nitrogen dioxide. For the analysis of ozone which is a naturally occurring oxidant but an important product of photochemical rcactions between pollutants another ~chemiluminescent instrument has replaced older non-specific methods. The ozone instrument which also measures light photo- metrically is based on the rcaction between ethylene and ozone.For the non-gaseous pollutants from vehicles smoke and metallic compounds chiefly lead compounds the shift to instruments has not been so marked. It is necessary t o collect samples on filter-paper for subsequent analysis by reflectance pliotonictry for smoke and by X-ray fluorescence for lead. These methods are not satisfactory and it is desirable to replace them with fully automatic and continuous methods when they become available. The Dynamic Reflectometer BY 13. UILES AND C. RROWN j,TIedical Rescavch Council A ir Pollutiom Unit St. h'avtholowieids Hospital .lfrdical Coliege Cirarterhozkse Square Tmzdon E C l l I GBQ) AND T. NASH (illicvobiological Research Establiskment Povtow Dozw Tl'iltsh i v e ) THE latest version of the instrument made in the MRC workshop was exhibited.It is intended for the measurement of very low concentrations of smoke or some trace gases in the atmosphere. August 19731 KEWER METHODS FOR ENVIRONMEXTAL POLLC:TIOP; ANALYSIS 199 The concentration of particulate pollution in the atmosphere can be determined by drawing tlie air through a white filter until a visible stain is produced. The reflectance of the stain can be nieasured as a percentage of the reflectance of a control and used as an estimate of the degree of pollution of the air. However standard “static” reflectonieters cannot deal with rural or maritime air in general because tlie stain is too weak to measure. The dynamic rekctometer increases the sensitivity about 100-fold by rapidly scanning stained and unstained portions of the filter as follows.A circular pattern of spots is produced on a 19-mm membrane filter disc by placing i t on a perforated plate and drawing the polluted air through the perfor- ations. The disc is then removed and placed on a rotating holder on the shaft of a synchro- nous motor. The disc spinning in its own plane is viewed by a lens - photocell system such that the image of only one spot falls on to the photocell at a time. The resulting a.c. signal is amplified rectified and displayed on a meter using an amplifier tuned to the spot frequency. The signal is linear over a wide range of stain intensity including stains of 99.9 to 90 per cent. reflectance. By using membrane filters impregnated with silver nitrate - glycerol the concentration of hydrogen sulpliide in ambient air at Porton could be measured in about 20 minutes arid was typically 0.2 p.p.b.in agreement with independent estimates of tlie global average. Sub-millimetre Spectroscopy Applied to Atmospheric Pollution Problems BY N. \fr. B. STOKE AXU J. E. HARRIES THE multiplex advantage of using an interferometer as a spectroscopic instrument was pointed out by Felgett at the Ohio State Symposium in 1962 and the energy throughput advantage by Jacquinot. The method of Fourier transform spectroscopy was niade practirable by the advent of the fast digital computer and since 1957 the technique has been progressively developed and refined. Early work was pioneered in France by the Comes in America by Mertz and in England by Gebbie working in the far-infrared spectral region. Recent import- ant advances have been the use of phase modulation of the interferometer1 and the develop- ment of helium-cooled indium antimonide and germanium detectors for routine use.These trcliniques have enabled us to obtain very high signal to noise atmospheric spectra between 10 and 40 cni-l. The investigation of the composition of the stratosphere has attracted increasing interest during the last few years in particular regarding thc amount of water vapour present. The dry model of the stratosphere has now been established by several workers including Harries and Uurroughs2 at the Kational Physical Laboratory working both froin aircraft and balloons by 31astenbrook3 in the U.S.A. McKinnon and hlorewood in Canada4 and by other workers. However tlie stratospheric problem has received great publicity during the last year since Johnston5 a t the University of California predicted that the amount of ozone in the middle stratosphere might become significantly depleted by the injection of large amounts of other pollutant gases (notably nitrogen oxides) in this region by large nunibers of supersonic trans- port aircraft.The gases of immediate interest are water vapour ozone nitric acid and the oxides of nitrogen. Other gases such as sulphur dioxide may also turn out to be ol importance. All of these gases are light molecules that have well populated rotational energy levels at temper- atures below 300 “K. ,4s they are dipolar it is in principle possible to measure their con- centrations in the atmosphere by recording their emission spectra. At lower altitudes the intense spectrum of water vapour normally masks emission from other species but in the stratosphere thc pressure line-broadening is small as is the water content and a great deal of information can be obtained regarding other trace gases.In the National Physical J>aboratory the stratospheric emission spectrum is observed by using a phase-modulated sub-millimetre wave Michelson interferometer directed toward tlie liorizon (zenith angle 90”). Work is also planned in which various other zenith angles will be used in order to obtain concentration profile information. Equipment has been mounted both in aircraft and balloon experiments although so far high-resolution spectra have been ob- tained only froin aircraft. Spectra have been recorded to a maximum resolution of 0.05 cin by using equipment mounted in the British prototype Concorde 002 flying at altitudes (Ihpartiizeiit o.f ?’lade and Iwdustry National Physical I-aboraloJy l‘eddi?zgforz ddlP.trx TR-I 1 Or- IT”) It is this problem which is now being investigated in depth.200 NEWER METHODS FOR ENVIROKMEKTAL POLLUTION ANALYSIS [Proc. SOC. A ~snZt;t. C k e m up to 50 000 feet. Observations were made between London arid Sydney during tlie far- eastern sales tour in June 1972 and irom Loridon to Capetown in January and Fehruary 1973. So far we have been able to measure water ozone nitric acid and nitrous oxide with confidence and Ineasuremcnts of nitrogen dioxide and sulphur dioxide are being steadily improved. More data are needed at higher resolution and work directed to this end is already under way. One stroke ol good fortune in this work has becn that oxygen of which the distribution is reasonably well established in the atmosphere while having zero electric dipole moment does possess a magnetic dipole.times the strength of electric dipole transition moments) but because of the high mixing ratio of oxygen present one therefore observes a strong rotational oxygen spectrum with which to compare the line intensities of other species. This fact combined with extensive laboratory work on the spectra of all of the atmospheric gases of interest provides adequate information on which to base the concentration measurements. REFEREXES Magnetic dipole transitions are very weak (typically I. 2. 3. 4. 5. Charnherlnin J. IMfhZnved P~.JJSZCS 1971 11 25. Harries J . E. and Rurroughs W. J . Q. J I R. AWt,t. SOC. 1971 97 519 Mastcnbrook H.J . -7. Atmos. Sci. 1971 28 1495. Mcliinnon D. and Morcwood 11. \\’. Ibid. 1970 27 485. Johnston H. Science N.Y. 1971 173 517. Air Pollution Measurement as a Tool for Practical Policy and Control Strategy BY L. A. CLAREXBUKG BECAUSE the production of certain goods such as food is inevitably accompanied by air- water and soil pollution today the population living in the immediate vicinity ol the yro- duction sites has to accept a certain degree of cliernisation of the erivironrnent. In environ mental care one tries to achieve pollution lcvcls as low as possible at least to maintain or to bring levels below the levels specified in standards. One of the reasons for sampling the quality of the air is to establish reliable data on which these standards can be based.To this end it is imperative that different monitoring and analytical tecliriiqucs yield at least directly intercomparable results. The present standards however deal with one contaminant orily although it is common knowledge that the combination of two or more contaminants inay aggravate the action ol the uthcrs. In practice tlie population living in industrial areas is always subjcctcd to more or less complicated mixtures of contaminants and it is desirable to have analytical techniques that are capable of analysing quantitatively the composition of the mixture. Safeguarding the physical health of the population requires “real time” monitoring of the pollutant concentrations. To develop a long-term policy it is necessary to derive from the data an expectation of the most probable development of the pollutjon levels 5 to 10 years ahead; only if rcliable data are available can such an analysis be made.Two reasons for nionitoring the quality of the air can be added related to pre- ventive cliecks the first is stack emission sampling and the second involves lorccasting air pollution cpisodes. The “real time” output of a monitoring system can be transformed into a parameter indicating whcther or not the actual meteorological situation gives rise t o pollution levels above “normal.” Cnfortunately monitoring techniques depend largely on the scale ol the problein e.g. in the lower stratosphcrc an excess of watcr vapour may be considered as a contaminant whereas the presence of a sufficiently high level of ozone is required in order to maintain the radiation balance around the earth; a t ground level water vapour is a normal constituent of the atmosphere whereas levels of ozone exceeding the natural background arc considered as a contamination.On a global scale attention must be focused on carbon dioxide water vapour ozone and perhaps oxides of nitrogen. One of the problems encountered is to dis- tinguish man-made pollution from a high fluctuating natural background. On a regional scale (e.g. North West Europe) the dominant problems are related to the conversion of sulphur dioxide and oxides of nitrogen into sulphuric acid arid nitric acid respectively. On a local scale very many pollutants must be considered. (Rijizwond Autiiurily Rottevdaili. Vastdaiad 96/101 T h e ;\‘ctheida~tds) i%UgUst 19731 NEWER METHODS FOR ENVIRONMENTAL POLLUTION AKALYSIS 201 In general one can say that the pollutants of concern depend largely 011 the scale of tlic problem.As a consequence the requirements to be met by monitors depend on thc scale and on the type of the problem; monitors should be capable of distinguishing a signal from a vari- able background the sensitivity should be adequate and the specificity high. The possible use of the data for deriving practical policieswas demonstratcd by giving two examples. The first example dealt with the “peak shaving” of odorous pollution. By using sulphur dioxide as a tracer for the behaviour of odorous pollution under adverse meteoro- logical conditions “real time” monitoring yields an early warning of the accumulation of pollution. If concentrations become significantly higher than “normal,” an alert is issued to the joint industries in the Rijnmond area.Each individual industry has prepared measures to prevent to a inaximuin the emissions of odorous pollution under such conditions. The second example dealt with the expectation of the sulphur dioxide trend in a city in the Rijnmond arm until 1980. We have entered a decade during which i t is very likely that far-reaching decisions will have to be made in order to prevent further deterioration of the environment arid eventually to improve the quality of our existence. These decisions which arc very often difficult and contradictory will have to be made at every level-the locality the region the globe. Good reliable data will facili- tate their achievement and will convince the opposition. In this whole process the analytical chemist has a vital r8lc to establish methods suitable for incasuring a t high altitudes for measuring the composition of mixtures of pollutants “real time,” for separating effects from variahle backgrounds and for many other reasons.In conclusion the demand for good reliable data was stressed. Analysis of Waste Waters and Interpretation of the Results BY A. (1. DOCHERTY [ I ii?pt?ial Chemical Industries Limited dgricultural Diziision P’om~cv aiid Sewices I17nrk.q E’fflzimf a ~ i d Analysis Group Billingham Z’eesside l‘S23 1LE) 1.u the whole pollution field industry has been overtaken by events and is trying hard to catch up. Watcr pollution is particularly iniportant bcc.ause i t is so difficult to avoid and industry is devoting a great deal of attention to this aspect of the problem.sampling of waste waters however they arise must be carried out with great care because a high proportion of the polluting material is normally present in the suspended solids and the samplc therefore must contain the appropriate proportion of thesc solids. Auto- inatic sampling is always t o be preferred if possible and there are proprietary sampling devices that can be used to take samples at regular intervals if continuous sampling is not practicable. Some are clockwork or battery-operated which is convenient as the ends of effluent pipes are often in remote locations. Fish inay be killed by substances that are poisonous to them but if several toxic sub- stances are present in different effluents reaching the same water-course complications can arise. For example cyanide is highly toxic but if some iron is also present the hexacyano- ferrate(I1) ion may be formed which is rnucli less toxic.On the other hand although phenol and hypochlorite ion arc both toxic the chlorinated phenol formed by their interaction is more toxic more odorous and more persistent. Again ammonium salts in acidic solution are not toxic to fish but if the pH of the water moves towards alkalinity an increasing proportion of the arnirioriium ion is converted to un-ionised ammonia which is highly toxic to fish. The separate toxicities of most toxic substances are already known and as a practical approach we have assumed that these toxicities are additive on the basis that complcxing effccts are likely to bc balanced by synergistic effects. This has proved to be a useful working hypothesis.The need to make calculations of this kind arises because at any given time there is only a limitecl amount of money available for effluent improvement purposes and it is wise t o see that this money is spent in such a way as to effect the greatest improvement. Once the back of the problem has been broken other questions will no doubt arise such as whether it is better to eliminate cyanide which although very toxic when diluted is readily degraded by natural biological processes to harmless substances or to eliminate zinc which although much less toxic is a conserved species that remains unchanged in the environment howev& much it is diluted. 202 NEWER METHODS FOR EXVIRONMESTBL POLLUTION ANALYSIS :Pt'OC. S O C . 11 I'LdJ~t. ChelPt. Fish may also be killed by being deprived of oxygen and therefore the biochemical oxygen demand (B.O.D.) of an effluent is an important characteristic of it.The conventional B.O.D. test used by the River Authorities takes 5 days to complete; it is entirely appropriate for monitoring the condition of an effluent from a domestic sewage treatment plant which normally changes only very slowly with time but it is unsuitable for an industrial effluent. For example if as is hoped there may in the future be fish in the River Tees a t Billingham it would be unacceptable to have to wait 5 days to know that a change had taken place in the effluent that was de-oxygenating the river because by that time the dead fish would be only too much in evidence. For this reason much effort has been exerted to find an alternative and the \Vater Research Association and Imperial Chemical Industries Limited Agricultural Division have jointly devised an apparatus that will measure the total organic carbon in an effluent on a continuous basis.The equipment that is now marketed commercially by Phase Separations Limited is based on a much earlier design which the \I'ater Research Association developed with Imperial Chemical Industries Limited Organics Division. The basic principle is to oxidise the carbon over copper oxide to carbon dioxide and then to reduce the carbon dioxide with hydrogen over a nickel catalyst to methane which in its turn can be detected and measured with a flame-ionisation detector which was developed for use in gas chromato- graphy and is highly sensitive to methane. The total organic carbon in an effluent is a different parameter from the B.O.D.but it has been found that for a given effluent the ratio between the two is usually constant. To conclude on a more hopeful note the pollution of the Tees Estuary is now not only not becoming worse it is actually improving which is not unexpected because industry on Teesside gave an undertaking to the Sorthumbrian River Authority just over a year ago that they would halve their effluent load to the estuary by 1955 and this process is now well under way. Industry and the Local Authority on Teesside are committeed to the target to induce fish back into the Tees and it is now beginning to look as if this target is indeed a real possi- bility. The Identification of Sources of Oil Spills BY D. 11. GREEK ASD J. ROBURN (Labovatory of the Government Chewiist ComvNall House Slawford Stieet Loizdoii SE1 9 s Q ) THE application of gas chromatography to the identification of the sources of oil spills began in the late 1960s and accelerated the analysis and opened the way to more sophisticated approaches.The technique used is temperature-programmed gas - liquid chromatography with a packed column non-polar stationary phase and flame-ionisation detection. The hydrocarbon profiles indicate the type of oil i.e. crude oil fuel oil or tanker sludge. However these profiles are often inadequate to define unequivocally a particular source and further tests are then necessary. From a survey of organic geochemistry it appeared that the isoprenoid alkanes might provide suitable markers in pollution studies. Just over 2 years ago a technique was de- veloped for the separation of the principal isoprenoid alkanes phytane pristane and the C, homologue.The first step is gel filtration by column chromatography on Sephadcx LH-20 which separates the oil into a number of fractions. Al specific fraction is treatedwith silicagel and 5A molecular sieve in order to remove unwanted components. The final extract when subjected to gas - liquid chromatography gives an envelope of other unresolved hydrocarbons with the isoprenoid peaks superimposed. Inspection of the chromatograms reirealed that weathered pollution samples had sustained some loss of isoprenoids; even the ratio of adjzcent peaks pristane to phytane was affected by weathering. The next ratio considered was phytane $Zzis C, homologue to pristane and this ratio was found to be much more stable the maximum variation for a highly weathered sample being 12 per cent.while for most pollution samples it was seldom more than 5 per cent. A few crude oils are difficult to resolve by these ratios particularly after weathering so that another parameter is necessary. The parameter chosen was the vanadium and nickel content and these elements have been determined directly on the oil by both X-ray fluores- cence and atomic-absorption spectroscopy. August 19731 SEWER METHODS FOR EXVIROSMENTAL POLLUTIOT ANALYSIS 203 If further supporting evidence is required another fraction from the Sephadex column can be examined. This fraction gives distinctive gas - liquid chromatographic profiles for many crude oils and its detailed examination by mass spectrometry has sho~vn the presence of two groups of compounds viz.C, to Clj dibenzothiophenes and C, to C, naphthalenes. The former group is more useful because the high boiling-points limit losses by volatilisation. These dibenzothiophene peaks often give useful confirmatory evidence but are not regarded as sufficiently diagnostic in themselves. It is interesting to note that recent publications on the gas - liquid chromatographic detection of sulphur compounds in oil by means of a capillary column and flame photometric detector give similar profiles. Recent evidence both published and from our own work has shown that capillary gas chromatography can effect sufficient resolution without the previous separation of the iso- prenoids. Unfortunately the reproducibility of peak ratios was inferior to that in the packed column method.Furthermore because it is advisable to distil pollution residues before injecting them on to a capillary column this becomes impracticable with some pollution samples which can be as small as 2 to 3 mg. This identification of crude oils is usually possible in those instances when a suspect sample is available. \Then such reference samples are not available more reliance has to be placed on circumstantial evidence. Fuel oils on the other hand are more difficult to identify owing to the possible variation in composition and in these instances a suspect sample is essential. Analysis can thus greatly contribute to the reduction of the oil pollution of coasts and inland waters by enabling existing laws to be more effectively enforced.The deliberate pollution of the open sea is a more difficult problem but even in this area analysis is making an increasing contribution. Gas - Liquid Chromatographic Detectors and Derivative-forming Procedures for the Selective and Sensitive Analysis of Pesticides BY K. I. BEYSON (Shell R e s e n d Limited Tunstnll Labovntov31 Bvoad Oak Road Sittingbotrvuzr Iiiizt I1IES 8.4G) THE analysis for pesticide residues involves sampling; sample storage; sample division ; extraction; storage of the extract; purification of the extract; quantitative determination; and confirmation of identity. An error in any one of these stages can lead to an erroneous result. The purification of the extract is frequently the most difficult step and the confirmation of identity the most neglected.Procedures for the quantitative determination of residues include elemental analysis bioassay ultraviolet fluorimetric and infrared spectrometry colorimetry paper and thin- layer chromatography polarography enzyme inhibition and gas - liquid chromatography. Most residue analysis is now carried out by gas - liquid chromatography although interest in colorimetric and spectrometric procedures may well increase as they are more amenable to complete automation. The widespread application of gas - liquid chromatograph)- is due to the availability of a range of sensitive and selective detectors including electron-capture flame-photometric micro-coulometric conductivity halogen-leak emission-spectrometric and plasma s!-stems together with mass spectrometry operated in the selective ion mode.The scope of these q-stems was discussed and recent developments in electron-capture detectors were outlined including the improvements in their linear range and the use of two detectors in series as a means of confirming identification and for providing a coulometric and absolute detector. Many compounds are not sufficiently volatile for gas - liquid chromatography or if volatile do not contain functional groups that can be detected with high sensitivity. Pro- cedures were described for the conversion of such compounds into derivatives that could be analysed by gas - liquid chromatography and using reactions which proceeded quantitatively at the sub-microgram level in the presence of naturally occurring co-extractives. Following a discussion of procedures of esterification silylation acetplation and phos- phorylation in general terms some specific applications of derivative-forming reactions were described for pesticides including .4zodrin DD and Frescon or their metabolites.204 SEWER METHODS FOR ENVIROKRIEKTAL POLLUTIOK .1SALYSIS [ P T O C . SOC. .4 I Z d J J t . ChCWZ. It was emphasised that current proccdures are capable of detecting trace amounts of pesticides in a wide range of substrates. There are considerable difficulties in applying these sensitive procedures but it is possible that our ability to detect the trace residues has outstrip- ped our ability to determine the significance of the results. Neutron-activation Analysis of Air Pollutants BY I. RI. DALE" ASD C. MCDOKALD (Chmistvy Defiai,tineizt T h e L7izivwsity Glasgow G12 SQQ) IT is well known that many elements exhibit toxic effects on biological systems at very low concentrations1 and accordingly analytical chemists involved in the investigation of environ- mental pollution should use the most sensitive and accurate methods available.These requirements are of special importance in the examination of air particulates because in most instances the elements are present in microgram to nanogram amounts. Seutron-activation analysis has been shown to be effective for the siinultaiieous determination of a number of elements in atmospheric dust In this paper the application of neutron activation to the analysis of dust samples collected from Glasgow's atmosphere in the period August 1972 to January 1973 is presented. The principles of activation analysis have been adequately discussed by a number of workers"1G and need not be considered further.In recent years the introduction of high- resolution solid-state gamma-ray detectors and multi-channel analysers that have from 1000 to more than 4000 channels has obviated the necessity for many of the chemical separations previously used. About twenty to twenty-five elements can be determined routinely and non- destructively in a single sample of atmospheric dust by using such a Samples of air were obtained by using a modification of the IYarren Spring air sampler. The pump is capable of sampling 20 to 30 1113 of air every 2 days. The dust particles are Collected on 2.5-cm diameter Nillipore filters with a pore diameter of 0.5 pni. The sampling funnel is situated about 15 feet above ground-level outside the Chemistry Department of Glasgow University approximately 50 yards from the nearest main road.After the required volume of air has been sampled the filter is removed and heat sealed in polythene sheeting. The first neutron irradiation is carried out in the reactor of the Scottish Research Reactor Centre East Kilbride about 10 miles from Glasgow. The dust samples together with suitable elemental standards are irradiated for 5 minutes at a thermal neutron flux of 3 x 1012 neutrons cm-2 s-l. Two hours after the irradiation the samples are returned to the l-niversity where they are counted by using a gamma-ray spectrometer (Suclear Enterprises 60 cm3 lithium-drifted germanium detector and Suclear Data 4400 computer-based multi-channel analyser) . The characteristic gamma-rays from sodium-24 (t+ 15 hours) manganese-56 (ti 2.58 hours) and bromine-82 (ti 35.3 hours) are measured and the concentrations are obtained from the peak areas.The samples are usually counted for 6 to 10 minutes and the standards for 1 minute. The samples are then irradiated in the DIDO Reactor Harwell where they are exposed to a neutron flux of 6 x 1012 neutrons cm-* s-l for 96 hours. After allowing short-lived activi- ties to decay for approximately 10 days the long-lived gamma-emitting isotopes are counted for 1 to 2 hours. X large number of gamma-ray peaks are measured these being characteristic of scandium-46 (t+ 84 days) chromium-61 (ti 27.8 days) iron-59 (ti 4.5 days) cobalt-60 (t 5.26 years) zinc-65 (t+ 245 days) selenium-i5 (ti 121 days) silver-llbz ( t i 270 days) anti- mony-124 (ti 60 days) caesium-134 (t 2.05 years) cerium-141 (t; 33 days) europium-154 (t+ 16 years) hafnium-181 (ti 42.8 days) gold-198 (ti 2.70 days) mercury-203 (ti 17 days) and protactinium-233 (from thorium-232) (t+ 27.0 days).The raw data from the spectrometer i n the form of punched tape are processed by a KDF9 computer aiid peak areas standard deviations and energy calibrations are calculated. A selection of the results obtained for the concentration of elements in dust particulates from Glasgow's atmosphere is presented in Table I ; a more comprehensive compilation of the * Present address \Vestern Regional Hospital Board Department of Clinical Physics and Bio-Engin- eering 11 \Vest Graham Street Glasgow G4 9LF. August 1973- SEWER METHO1)S FOR ENVIRONMENTAT. T’OLLIJTIOS ANALYSIS 205 analyses is being prepared for publication.The accuracy of the analyses estimated from counting statistics and reproducibility of replicate analyses for sodium manganese bromine and zinc is & 10 per cent. and for the otlier elements i.25 per cent. The values ohtaincd for Glasgow’s atmosphere are comparable with results quoted for similar industrial cities in the United and Great Britain.4 TABLE I cONCENTR?\TIOSS OF ELEMENTS IN GLASGOW’S ATMOSPHERE AUGGST TO T\TO\’EMBER 1972 Element Range/ng 1n-3 Meanjng 111-3 Na * 90 -’ 980 3.50 Mn* 1 ~ 290 29 Br* 25 - 300 76 s c 0.007 - 0.31 0.1 F e 900 - 18 000 5900 r 5.6 ~- 92 15.5 o 0.3 .- 2.1 0.9 Zn 25 - 1740 300 Se 0.4 - 12.5 3.3 Sb 1.5 - 29 8.7 -% 0.4- 13 3 ’ 3 * August 1972 to Jannary 1973; 7.5 sainplcs. I t is possible to correlate many- of the variations in the concentrations of tliese elements with the weather conditions prevailing at the time of collection.For example periods o f fog produced greatly increased concentrations whereas periods following raidall and at the weekends showed decreased levels. The exact relationship between Incteorological conditions and the levels of the various elements in the atniospliere is the subject of a separate study. Plans are in hand t o extend the number of sampling sites in tlie Glasgow area in order to investigate in detail local fluctuations of metal concentration in the atmosphere. \Ve are pleased to express our gratitude to the staff of the Glasgow Weather Centre Glasgow’s Smoke Officer Prof. H. W. Wilson of thc Scottish Research Reactor Centre and t o a private Scottish trust fund for a grant to purchase the counting equipment.One of us (C.M.) acknowlcdges with thanks an S.R.C. Research Studentship. REFERENCES 1. 2 . 4. 4. .i. A. de Sorte U. Gijhels R. and IIoste J. “Xeutron Activation A4nnl?;sis,” m’ilev-InterscieIlce U~iderwood E. J “Trace Elements in Human and Animal Nutrition,” Acadcmic Press London Zollcr. W. H. and Gordon G. E. 14naZyt. Chevn.. 1970 42 237. Dams R. Robbins J. h. Rahn I<. A, and Winchester J. W. Ihid. 1970 42 861. Peirson D. II. Cawsc P. A,. Salmon L. and Cnrubrxy K. S . Natuve L o m ~ . 1973 241 232. Krnger P. “Principles of Activation Axdysis,” Wile).-Interscience IAondon 197 1 . 1971. London 1972. A Mobile Laboratory for Monitoring Atmospheric Pollution BY P. A. HOLLIKGDALE-SMITH AND L. C. THOMAS (Illiwistvy of Defmice Cheiviical UcfeNce Establish.inc?zt Porton Down Salisbury TlyiZtsliivr) A RIORITX laboratory was described that has been equipped to identify and measurc all atmos- pheric pollutants rather than specific pollutants as is the normal practice.The laboratory contains an infrared spectrophotorneter fitted with long-path (40 m) gas cells and a quadrupole mass analyser. These instrumcnts will measure directly pollutant concentrations down t o 1 part in 10’. For the measurement of lower concentrations long-term sarnpling equipment is included for both vapours arid particles. Analysis of particulate pollutants is at present restricted to heavy-metal analysis. In addition a gas chromatograph fitted with flame-ionisat ion tliermionic and electron- capture detectors is used to monitor organochlorine and organopliosphorus pollutants.206 [Piroc. SOC. Afialyt. Chew. Portable air-sampling equipment for use in atmospheres inaccessible to the laboratory is also carried. A mobile generator is carried to enable the laboratory to operate a t sites where mains electric power is not available. These included the application of the portable sampling equipment to studies of submarine atmoaplieres and the variations in pollutant concentrations as a function of wind direction w n d velocity and time. The importance of a knowledge of the long-distance air travel as well as the local wind direction in assessing sources of pollution was also discussed. During the period of the Chemical Congress in Swansea the mobile laboratory was work- ing on a site adjacent to the Chemistry Department and the equipment and the results being obtained were discussed with a large number of visitors. CHARACTERISATIOS OF FOOD ASD AGRICULTURAL PRODUCTS Examples of the results obtained with this equipment were discussed.

ISSN:0037-9697    

DOI:10.1039/SA9731000197

出版商:RSC    

年代:1973

数据来源: RSC

摘要:

206 CHARACTERISATIOS OF FOOD ASD AGRICULTURAL PRODUCTS [Piroc. SOC. Afialyt. Chew. The Characterisation and Quality of Food and Agricultural Products With Special Reference to EEC Requirements The following is a summary of one of the papers presented at a Joint Meeting of the Rficro- cheniical Methods Group and the JYestern Region held on ,April 5th and Gth 19’73 and first reported in the May issue of Proceediizgs (p. 99). A further report and summaries of sei’en of the papers presented at the Meeting appeared in the July issue of P~octwliiigs (pp. 171-153). The Determination of Pesticide Residues in Food Crops BY 11. J. EDWARDS BEFORE a new pesticide can be marketed it must be shown that the use of the compound in good agricultural practice will not leave harmful chemical residues in the treated food crops.A41so the pesticide must not produce contamination of the environment or undesirable effects upon non-target organisms. It is necessary therefore to develop analytical procedures that are capable of determining the minute amounts of pesticide and its breakdown products that may occur. Lower limits of detection for the pesticide of 0.1 to 0.01 p.p.m. are generally required for food crops and it is necessary to work at the parts per billion le\Tel with milk and water samples. A residue analytical method generally contains the following steps-sampling (including transport sub-sampling and storage) ; extraction ; “clean-up” ; and determination. As in all analytical work the importance of correct sampling and sub-sampling cannot be over-emphasised. At present there are no internationally agreed sampling procedures al- though the Codex Committee on Pesticide Residues has been studying this problem for some important food commodities.However a number of useful papers have been published on the sampling of food crops and on specific sampling problems.lv2 Loss of pesticide residues (by volatilisation and/or breakdown) during transport and during storage before analysis is mini- mised by deep freezing (below -10 “C) ; it should however be proved that losses do not occur even under these low-temperature conditions. The procedure used to extract a pesticide will depend upon its structure and on the nature of the sample and a wide variety of solvents and extraction procedures have been a d ~ p t e d . ~ However for the majority of organo-soluble pesticides mixtures of solvents are generally pre- ferred and usually consist of a polar water-miscible solvent with a non-polar water-immiscible solvent (e.g.acetone - hexane). A clean-up step is included in order to remove co-extracted materials that would interfere in the subsequent determination. Of the many techniques a ~ a i l a b l e ~ ~ liquid - liquid parti- tion and column chromatography are the most widely used. The former separates the pesti- cide from the co-extractives by selective partitioning between two immiscible solvents ; the latter approach uses a chromatographic separation on a suitable adsorbent. Sumerous materials are available including alumina silica gel carbon and ion-exchange resins but Florid (a synthetic magnesium silicate) is the most widely used and tested.Other important clean-up techniques include paper and thin-layer chromatography sweep co-distillation chemical treatment and low-temperature precipitation. (I.C.I. Plant Pvotectzoiz Limited Jealott’s Hill Reseavch Station Bvackiaell Bevksh ive) August 19731 CHARXCTERISATION OF FOOD AKD AGRICULTURAL PRODCCTS -307 The determination of the pesticide in the cleaned-up extract requires sensitive detection systems. In the past spectrophotometric methods including colorimetric and ultraviolet were most frequently used but were not very sensitive or selective. Todai- the most useful technique is gas - liquid chromatography with selective detectors. The successful application of the extremely sensitive electron-capture detector to the analysis of chlorinated hydro- carbon pesticides is now well known.More recently the introduction of organophosphate and carbamate pesticides has prompted the development of phosphorus- and nitrogen-selective thermionic detectors and the phosphorus- and sulphur-selective flame photometric detectors. The flame photometric detector of Brody and Chaneyj is outstandingly simple but extremely selective and sensitive. Compounds that contain phosphorus or sulphur produce escited species such as HPO and S-S when burned in a hydrogen-rich flame. The characteristic radiation emitted is isolated by narrow band-pass filters (526nm for phosphorus; 394nm for sulphur) and detected by a photomultiplier tube. The detector has proved to be stable and essentially trouble-free-we have used one continuously for over 2 years (approximatell- 10 000 individual injections) for the determination of organopliosphate insecticides without the need to dismantle the equipment for cleaning.The response to phosphorus is extremely repeatable and linear over a wide range down to sub-nanogram levels. (Sulphur produces a rather less sensitive non-linear response.) The extreme selectivity of the detector6 allows the direct analysis of many crop extracts without clean-up and often the latter is only necessar!. because of the deleterious effect of co-extractives upon column performance. After each injection the detector flame blows out and there is a simple standard re-ignition procedure- it is possible however to prevent this effect by interchanging the hydrogen and oxygen;air lines to the detector without any apparent effect upon response and noise levels.The selective thermioriic detectors’ are essentially flame-ionisation detectors modified b>- inclusion of an alkaii metal salt (e.g. caesium bromide rubidium sulphate rubidium bromide or potassium chloride) generally as a crystal or coated metal electrode. -in enhanced response is obtained to phosphorus- and nitrogen-containing compounds when these are burned in the Hame. They have been widely used for the detection of organophosphorus compounds down to picogram levels. The determination of nitrogen compounds has been more difficult requir- ing more critical control of the hydrogen gas flow to the detector and of the shape and position of the alkali metal salt electrode. During the past 2 years we have used rubidium bromide detectors (Hewlett Packard Model l5lGlX mounted on an HP 5750 series gas chromatograph) almost continuously for the routine determination of residues of hydroxypyrimidine fungicides and related compounds in grain straw and soil samples.An example is ethirimol the active ingredient of Nilstem fungicide The detectors have been stable and trouble free in operation with a repeatable and linear response down to sub-nanogram amounts of nitrogen (less than 06ng of ethirimol can be detected). The parameters that most affect detector response are detector temperature hydrogen flow-rate and distance between the flame tip and the surface of the rubidium bromide crystal. The response to nitrogen and the noise level increased with increasing detector temperature and the recommended operating temperature of 370 “C gave optimum results.(Transmission of heat from the detector can produce problems when using glass columns with silicone O-rings.) The hydrogen flow-rate is accurately controlled a t its optimum (25 to 26 nil min-l) by a mass flow controller. The accurate placement of the rubidium bromide crystal above the flame to give the optimum response to nitrogen is obtained using adjusting screws. .After several weeks use with silicone columns “bleeding” of the stationary phase causes a deposit of silica on the crystal whicli lowers the detector response and increases the noise level. 208 ORIGINAL PAPERS IN PHARMACEUTICAL AKALYSIS LProc. SOC. il??nl$ Chew. This deposit is removed by scraping it with a sharp blade thus restoring the original response. Helium has been used as the carrier gas and there has been little change in response or in the shape of the crystal after 2 years use.Kitrogen carrier gas can be used but the increased standing current obtained can cause “backing off” problems ; the hotter flame also produces a more rapid loss of rubidium bromide with a consequent change in shape of the crystal face. Recently a modification of the rubidium bromide detector has been made available in which an auxiliary ring electrode is placed around the flame tip. A variable voltage is applied to this electrode to suppress background noise-it is claimed that the sensitivity and stability are increased by a factor of ten over the original detector design. REFERENCES I. 2. 3. Lykken L. hfitchcll L. E. and IVoogcrd S . X Apric. Fd C ~ ~ J H . ~ 1967> 5 601. Lykken L. Residue R e v . 1963 3 19. 3IcCully I<. A, and RIcLeod H. A, i?z “Proceedings of International Symposiuni on Identi- fication and Measurement of Environmental Pollutants,” rational Research Council of Canada Ottawa 1971 p. 224. NcCully K. A zvz Tahori A. S. Editor “hlethods i n Residue .Analysis,” Proceedings of the 2nd International TUP=\C Congress on Pesticide Chemistry 1971 T-olurne 4 p. 31;. Brody S. S. and Chaney J . J . Gas Chvomat. 1966 4 42. Beroza &I. and Bowman 31. C. ziz “Proceedings of International Symposium on Identification and Measurement of Environmental Pollutants,” rational Research Council of Canada Ottaxa 1971 p. 216. 4. 5 . 6 7. Aue 11’. X. A d u . Chem. S e r . 1971 104 39.

ISSN:0037-9697    

DOI:10.1039/SA9731000206

出版商:RSC    

年代:1973

数据来源: RSC

摘要:

208 ORIGINAL PAPERS IN PHARMACEUTICAL AKALYSIS LProc. SOC. il??nl$ Chew. Original Papers in Pharmaceutical Analysis The following is a summary of one of the papers presented at a Xeeting of the Joint Summaries of three of the papers Pharmaceutical Analysis Group held on June 8th 19i2. appeared in the January 1973 issue of Pvoceediiigs (p. 9). The Individual Tablet Assay of Digoxin Tablets BY D. SCL-FFAM (Departiiieizt of Trade and Industry Laboratory o,f the Gozerizvzent Chewzst Covizeall Hozrse Stni~zfoi d Slveet Loiadoiz SE1 9SQ) DIGOXIN is frequently used in the treatment of congestive heart conditions to increase the force of contraction and cardiac tone. Patients placed under digitalis medication undergo two phases of administration the initial course for digitalisation and the second for main- tenance.Both phases require individualised supervision with the dose on digoxin needed to control the patient being accurately quantitated. The test procedure of the currently revised United States Pharmacopoeia is to select thirty tablets and to assay ten of these tablets individually by a suitable modification of the assay procedure for digoxin tablets. The requirements are met if not less than nine tablet results lie between 85 and 115 per cent. of the stated amount and the content of all ten tablets is within liniits of 75 to 125 per cent. If the content of not more than two tablets falls outside the limit of 85 to 115 per cent. the remaining twenty tablets are tested singly and the requirement is met if all twenty have a digoxin content of 85 to 115 per cent. T'arious colorimetric methods have been devised for the determination of digoxin.Alkaline picrate,l xanthydrol,2 3,5-dinitroben~ene,~ acetone phosphoric acid,? iron(II1) chloride,j 2,4-dinitrodiphenylsulphone6 and thiobarbituric acidi have all been used for this purpose. The British Pharmacopoeia favours the use of iron(II1) chloride for all digoxin determinations. The European Pharmacopoeia stipulates the use of alkaline picrate for the assay of the pure drug substance. The United States National Center for Drug Analysis finds the thiobarbituric acid method' suitable for automation although an appropriate correction must be applied for the interference of dextrose. An acid-induced fluorescence method* tried for its very high sensitivity did not give acceptable repr~ducibility.~ The iron(II1) chloride method,j after suitable modification as described was found to give both the accuracy and the reproducibility required for individual tablet assay.ilugust 19731 ORIGINAL I’APERS IN PHARMACEUTICrlL AXALYSIS 209 Tlie tablet is placed in a centrifuge tube and nioistened with methanol then pulverised by shaking it with ten glass balls of 6 mm diameter. An 82 mg 1-1 solution of anhydrous iron(II1) chloride in analytical-reagent grade glacial acetic acid containing 2 per cent. of analytical-reagent grade sulphuric acid is prepared and 25 in1 of this solution are added to the powdered tablet. The tube is well stoppered and vigorously shaken for 110 minutes tlieri centrifuged for 10 minutes and the clear solution is placed in a 4-cm cell and immediately read at 590 nm against a digoxin solution of similar concentration taken through the same process.Light is excluded in all stages following the addition of iron(II1) chloride. Most tablets are pulverised in 5 minutes ; the addition of anti-bumping granules is rcconimended if tablet powder cakes on to the walls of the tube. Tlic ball-mill crushing process is an improvement on techniqucs with pestle and mortar or flanged-rod and provided that the stoppers used are well attached there is no risk of mechanical loss. Investigation of the reagents in successive British Pliarrnacopoeias now shows that both cliloroform and methanol are unnecessary. Although methanol could be retained on solubility grounds and shows no marked effect on the absorbance chloroform intcrfcres and gives a lower absorbance in the presence of lactose.The approximate doubling of the iron( 111) chloride concentration markedly improves the development time and maximum colour is now reached after 1 hour. 1 he maximum absorbance is obtained when the automatic shaker and the spcctrophoto- meter are sited in a dark room and when there is minimal exposure to tungsten light during the actual taking of the reading. In the presence of an authentic tablet excipient containing lactose 53.5 dicalcium phos- phate 19.8 corn starch 144 dextrose 10.0 talc 1.8 and magnesium stearate 0.2 per cent. an average recovery of 99.8 per cent. with a standard deviation oi 0.33 per cent. is demon- strated on ten results. The calibration graph is a straight line for 10 to 500 pg of digoxin iri 25 ml of iron(II1) chloride reagent.Results lor 250-pg tablets supplied by five different manufacturers (Table 1) show the range of content variation of ten tablets from each manufacturer. In all instances the vari- ation in the mass of the sample did not exceed 5 per cent. of the mean tablet mass. Tlie last colunin shows that the B.P. assay obtained by bulking twenty tablets docs not significantly differ from thc single tablet assay average. TABLE I ~. KESCLTS FOR DIGOXIX CONTENT OF TABLETS FROM DIFFERENT MANUFACTURERS Manufacturer Range/& Averagc/pg Found by B.P. mcthod/p.g A 205 to 275 233 237 B 200 to 310 236 241 C 210 to 298 249 250 D 220 to 245 235 232 E 185 to 410 271 268 The proposed content of uniformity on Digoxin Tablets B.P. test requires the individual assay oi ten tablets. The content of each tablet is between 80 and 120 per cent.of the average except that for one tablet the content may be between 75 and 125 per cent. of the average. Further information is required to decide whether the products of manufacturers A B and C pass the proposed U.P. specification although that from manufacturer B fails the U.S.P. XVIII specification. Tablets of manufacturer D pass and of E fail both specifications. The iron(II1) chloride method discussed in this paper is closely similar to the method for uniformity of content included as an Amendment to E.P. 1968 Addendum 1969 official from February lst 1973 and R.P. 1973 to be made official from December lst 1973. REFERENCES 1. 2. 3. 4. 6. 6. 7. S. Baljet H. Schwei?. ApothZtg 1918 56 71 and 84; Chew. Ahstr. 1918 12 1336. Pesez &I. An& Pianvm. Fr. 1952 10 104; CILCI% Abstr. 1952 46 7000c. Tattje D. H. E. J . Pharm. Pharmac. 1957 9 27. Dcqueker R. and Loobuyck M, Ibid. 1966 7 622. James A. E. Laqner 1;. O. arid Mclrityre J. D. J . 4vzev. Pizann. Ass. Scieizt. Edn 1947 36 1 Tattje D. H. E. J . Piravm. Plzavmac. 1958 10 493. Myi-ick J. W. J . Phnraz. Sci. 1969 58 1018. Jilioury A. J. in Knwerau E. Editov “Autoinat.ion in Analytical Chcmistry,” Technicon Sym- posium 1967 Blecliad Inc. New York Voluii~e 1 p. 192.

ISSN:0037-9697    

DOI:10.1039/SA9731000208

出版商:RSC    

年代:1973

数据来源: RSC

摘要:

2 10 ASPECTS OF CHROMATOGRAPHY [P~w. SOC. Analyt. Chein. Aspects of Chromatography The following are summaries of six of the papers presented at a Joint Meeting of the Scottish Region and the Chromatography and Electrophoresis Group held on April 12th and 13th 1973 and reported in the May issue of Proceedings (p. 99). Characterisation of Long-chain Fatty Acids by Gas Chromatography EY G. R. JAMIESON (Chemistry Department The Paisley College OJ Tedmolog.v Paisley Buii/Yeashtve) THE separation of long-chain fatty acids by gas - liquid chromatography was first described by James and Martin in 1952 and some 21 years later although many laboratories have been using the method all the problems have by no means been sol\-ed. Reviews of the subject are still being written1r2; “unusual” fatty acids (I 11) are still being discovered; and fatty acids which have become accepted as “usual” are being found in “unuwal” p l a ~ e j .~ ~ CH,(CH,),CH, H CH,(CH,),CH CH \C&/ H,C=cd \CH,(CI-I,),CO,H H/ \CH,(CH,),CO,H I I1 (marine turtles ; jellyfish) (ocean sunfish) The gas - liquid chromatography of long-chain fatty acids is usually carried out using the methyl esters and more and more inter-esterification is being used to obtain methyl esters from various lipid classes. These inter-esterification methods are used with lipid fractions from column separations and with fractions separated by thin-layer chromatography with or without extraction of the lipids from the layer material. IIetliylatiori techniques have recently been re~iewed.~ STATIONARY PHASE P0LARITX‘- The separation of unsaturated long-chain methyl esters may he discusred with particular reference to two effects London dispersion interactions-these effect separations on a relative molecular mass - solubility - volatility basis.These interactions decrease with an increase in tlic polarity of the stationary phase. The specific molecular attraction between the polarisable olefinic bonds of unsatur- ated methyl esters and the ester groups of a polyester stationary phase. This polar effect specifically retards the unsaturated methyl esters and in most instances outweighs the London interactions which have an opposing effect. The system 1 8 O ; 18:lw9; 18:2wB; 18:3w3 has been likened to a coiled spring which stretches with increasing column polarity until 18 3 w3 attains and then passes the position of 20 0.This simple concept of fatty acid polarity on polar liquid phases is based on a proportionality of retention times among a family of acids. The magnitude of the increase is dependent primarily on the polarity of tlic liquid pliase (i.~. on the actively engaged proportion of oxygen function t o niethylene units). Other effects are secondary and may well work through tlie availability of the oxygen function. Some ol these eflects arc (a) the nature and source of the stationary liquid; ( b ) loading of the liquid pliase and the nature of tlie support; (c) the age of the column; and (d) the column temperature. However all of these effects act in a cuinulative fashion with the primary olefinic polarity of the methyl ester to determine the actual polarity of the system used for gas - liquid chromatographic analysis.The problem of polarity is fundamental in the gas - liquid chromatography of lipids and it must be clearly recognised b y all analysts that every gas - liquid clirornatograpliic colurrin has it own unique separation characteristics and comparison among columns must be strictly limited. August 1973; ASPECTS OF CHROMATOGRAPHY 211 The above simple concept has been elaborated into a promising “universal” approach Literature to the problem of correlating retention data with the polarity of the liquid phase.6 data and original data were processed by computer for the equation where a and b are the computer-derived constants for the methyl ester (x). IDESTIFICATIOS OF A5 ACIDS FROM COSIFER LEAVES~- and were identified as follows -4 series of C, and C, polyolefinic acids with unsaturation were found in conifer leaves (a) application of equation (1) to gas - liquid chromatographic retention data allowed the acids to be placed in “unsaturation” classes and this placing n-as confirmed by argentation thin-layer chromatography; ( b ) application of Type I and Type I1 separation factors allowed the relationships between the acids to be determined; ic) acids obtained from argentation thin-layer chromatographic separations were oaidised by periodate - permanganate and the positions of unsaturation determined; ( d ) esters were examined by the inercuration reaction developed by Gunstone and Inglis* and the nature and amounts of the 1; acids confirmed.SEPARATIOS OF Cis- ASD tYa?ZS-MOXOEXES- It is very easy to report 18 1 as oleic acid but on many gas - liquid chromatographic columns this peak can be a multiple peak due to positional and/or geometric isomers of oleic acid.Separation of cis- and tram-monoenes can be achieved only on very efficient gas - liquid chromatographic columns. These isomers can be separated by argentation thin-layer chroma- tography or they can be separated by gas - liquid chromatography after hydrosylation of the olefinic bond and conversion of the dihydroxy compounds into isopropylidene derivatives. SEPARATIOS OF ASALOGUES OF LOSG-CHAIS ACIDS- Systematic separation factors derived from methyl esters can be applied to alcohols and their acetate trifluoroacetate and trimethylsilyl derivatives and to hydrocarbons. The type I1 separation factors are only slightly affected on going from acetate to trifluoroacetate deriva- tives but trimethylsilyl derivatives gi1.e much lower values for the Type I1 factor.Presum- ably the large trimethylsilyl group reduces the effect of the w3 and w6 structures. T-arious derivatives of glyceryl ethers can be separated by gas - liquid chromatography and the following have been found useful acetate trifluoroacetate trimethg-lsil~rl and cyclic butylboronic esters. Recently we have found that conversion of the glyceryl ether into the corresponding n-propyl ether (CH,CH,CH,-0-R) gives derivatives that have useful chromato- graphic properties. REFERENCES 1 . 2 . 3 . 4. 5 . 0. 1 . S. Jamieson G. R. in Gunstone F. D. E d i t o ~ “Topics in Lipid Chemistry,” I A ~ O S Press London hcliman K. G. Pvog. Chrm. Fats Lipids 1972 12 285. Jamieson G.R. and Reid E. H. Plzytocheivislvy 1969 8 l4S9. -,- Phytochemistry 1951 10 1576. Christie TV. TV, in Gunstone F. D. Editor “Topics in Lipid Chemistry,” I,oqos Prcss London Jamieson G. R. and Reid E. H. J . Clzvonzat. 1969 42 30.2. -,- Pkytocheiaistry 1972 11 269. Gunstone F. D. and Inglis R. P. Chenb. Coiniizuizs. 1972 12. 1970 Volume 1 p. 107. 1972 L-olurne 3 p. 121. 212 ASPECTS OF CHROMATOGRAPHY Gas - Liquid Chromatography of Waxes [Proc. Soc. Annlyt. Chem. BY R. J. HAMILTOK (Depavtment of Chemislvy Liverpool Polytichnic Byrom Stvcet Liuerpool L3 3.4 F ) THE physical appearance of leaf surfaces varies in different plant species as shown by electron micrographs. Part of the difference is due to the surface wax which consists of hydrocarbons esters. free fatty acids and alcohols in almost all plants.In some plants ketones aldehydes and secondary alcohols are also present but a range of rare constituents which may occur in only five or six plant species or families include alkyl acetates alkenes and aromatic hydro- carbons. The separation of most of the wax classes can be accomplished by thin-layer chromato- graphy on silica gel with ether - light petroleum (5 + 95) but the spots of acetates ketones aldehydes and diol plasmalogens tend to overlap. Each of these classes normally consists of a homologous series of compounds which makes their gas - liquid chromatography easier. Xpiezon L is the stationary phase of choice up to 270 "C but above this temperature silicone phases such as OV-1 and Ol7-l7 become more valuable as they are more thermostable.The gas - liquid chromatographic distribution of hydrocarbons can be used as an aid to taxonomy of the plant species as was demonstrated by Herbin working with South African Aloe species. The wax esters of rye grass which range from C, to C,, have been analysed on a 5 foot x t inch column of 017-1 with a pre-heater. The short columns used in triglyceride analysis were not required. The limited number of stationary phases available for high-temperature work have a restricted range of polarity with the result that alkyl acetates have a retention factor similar to that of methyl esters and it is not possible to determine the class of component by its gas - liquid chromatographic retention alone. In rye grass aldehydes were noted by the ready formation of their 2,4-dinitrophenyl- hydrazone derivatives on the thin-layer chromatographic plate but in one plant wax from Cheno$odz'zwz d b w z the aldehydes were mixed with a second component.This second class of constitucnt was identified as alkyl acetates by its infrared absorption band a t 1240 cm-l and by the analysis of the acetic acid liberated on saponification on a Porapak Q column a t 150 "C. Rye grass also contained an unusual constituent that had a retention index of 2150 on Xpiezon L a t 180 "C. This component was shown to be antliraquinone by its ultraviolet absorption a t 252 264 and 253 nm and by its mass spectral breakdown peaks a t q,ie 180 152 and 76. The Analysis of Free Fatty Acids by Gas Chromatography BY G. c. COCHRAXE (The Dzstillers Company Ltd. Glenochzl TecJinzcal Cedi e .Wenstvie Clacki7iaiiizaizsJiire) THE gas-chromatographic analysis of free fatty acids has been limited by the relatively high boiling-points of the acids and by their adsorption in the chromatographic column.Con- sequently gas-chromatographic analysis is generally carried out after the acids have been converted into more volatile and less polar esters such as the methyl ester. However the preparation of methyl esters makes the analytical procedure longer and more tedious and when the lower acids are present in small concentrations the composition of the mixture is altered owing to evaporation losses and the large solvent ~ e a k . l - ~ The use of n - p r ~ p y l ~ n- butyl+ and n-decy16 esters also does not give quantitative results for the lower acids. It was thcrefore desirable to develop a method for analysing the free acids whicli were of interest namely acetic to lauric acid.August 1973 ASPEC,TS OF CHROMATOGRAPHY 213 The following columns were evaluated for the analysis of free fatty acids in which a Pye-L-nicam 104 gas chromatograph fitted with a flame-ionisation detector was used (1) 3 per cent. of orthophosphoric acid on Porapak Q (80 to 100 mesh)6; (2) 6 per cent. of FFAP” on Porapak Q (80 to 100 mesh)’; (3) 10 per cent. of polyethylene glycol 2011 plzis 3 per cent. of orthophosphoric acid on Chromosorb W; (4) 1-2 per cent. of polyethylene glycol 2011 on terephthalic acidg; ( 5 ) 10 per cent. of FFAP on Chromosorb \VO; (6) 10 per cent. of FFAP plus 5 per cent. of isophthalic acid on Chromosorb W; ( 7 ) 25 per cent. of neopentyl glycol adipate on Chromosorb IY11; (8) 10 per cent.of neopentyl glycol adipate on Chromosorb \V. \Yith the exception of the Porapak and terephthalic acid columns Chromosorb Ti‘ (80 to 100 mesh) which had been washed with acid and treated with dimethyldichlorosilane (DRICS) was used as the solid support throughout this work. I t was coated with the station- ary phase with or without an involatile additive in the usual manner. The coated support was packed into glass columns (5 foot x Q inch 0.d.) and fully conditioned at the recom- mended maximum temperature before being evaluated. Although columns (1) to (4) had all been reported in the literature as being suitable for the analysis of free fatty acids it mas found that they became unsatisfactory after a few days owing to either excessive ghosting or deterioration of the column packing.Excellent resolution of the free fatty acids horn acetic to lauric acid was achieved with column ( 5 ) when they were present in concentrations of a t least 1000 p.p.m. When lower concentrations were analysed poor peak shapes were obtained especially for acetic acid and ghosting also occurred. The addition of isophthalic acid to this column (6) gave similar results. As formic acid is more polar than any of the acids to be analysed and does not respond to the flame-ionisation detector it was believed that the addition of formic acid to the solution of acids would result in the formic acid saturating the so-called active sites of the Chromosorb thereby preventing adsorption of the other acids. This approach was initially successful with column (5) but after it had been used for a few days the response of the lower acids began to vary from injection to injection.It was then decided to analyse the fatty acids in a continually polar atmosphere by bubbling the carrier gas through 3x formic acid before it entered the column [column ( 5 ) . This procedure was immediately successful and gave excellent peak shapes for all the acids from acetic to palmitic acid. After 3 to 4 weeks however the column packing began to deteriorate until it could no longer be used. This was repeated twice with the same result. It was then found that Ackman and Burgherll had reported the successful analysis of free fatty acids by passing carrier gas over concentrated formic acid before passing through a column packed with 25 per cent. of neopentyl glycol adipate. Column ( 7 ) was prepared and this procedure was used to give very good results for the acids from acetic to lauric acid and the column began to deteriorate only after having been used for about 10 months.This column was therefore used for the analasis of free fattv acids. Column (8) was more suitable / for higher acids. It was never possible to use an amplification greater than 2 x 10-l0 A (attenuation x 200 on the chromatograph amplifier) as thestanding cirrent of the 1-mV recorder was then greater than 1 niY. This suggests that either the formic acid does in fact have a very small response to the detector or that there is a continual bleed from the column. Fortunately it was unnecessary to use an amplification greater than 5 x 10-lO A (attenuation ~ 6 0 0 ) for any of the samples as peaks representing acids present a t less than 1 p.p.m.were measurable a t this attenuation. The formic acid was contained in a small glass reservoir wliich was sealed by a screw-type metallic top fitted with stainless-steel couplings. These couplings were connected to stainless- steel tubing (1116 inch 0.d.) through which carrier gas passed into the reservoir and then into the column. This glass container can be obtained from Pye-Unicam who use it for holding the sample in their fully automatic preparative-scale gas chromatograph. It is very easily fitted to the 101 gas chromatograph as the chromatograph is supplied with small bolts fitted to the oven unit which allow the container to be held securely to the chromatograph. There was no sign of corrosion of the tubing or of the detector which was constructed of stainless steel after formic acid had been in use for more than 1 year.It was found that the greater the volume of formic acid used the greater waq the standing current of the recorder and the volume finally chosen was 1.5 ml. This volume was just sufficient for 1 day’s operation so that it had to be renewed first thing each day. * Free fatty acid phase. 2 14 ASPECTS OF CHROBI.4TOGRAPHY ~PYOC. SOC. 4iialyt. Clzem. The conditions that were found to give the best results are as follows- Temperature programme . . .. . . 76 "C rising a t 6 "C min-I to 200 "C Injection port temperature . . .. . . 130 "C Gas flow-rates- h'itrogen * . .. .. 120 ml min-I Hydrogen . . .. .. . . .. 50 ml min-l Air . I .. . . .. . 700 ml min-1 Volume of formic acid in reservoir .. . . 1-5 ml (renewed each day) Heptanoic acid was used as the internal standard and the acids were dissolved in a solution containing 20 per cent. of 5 N formic acid and 80 per cent. of acetone. Acetone was found to have an impurity that tended to be eluted on top of n-butyric acid. This impurity was removed by fractional distillation and the formic acid was similaly treated owing to the pre- sence of acetic acid and other impurities. However this treatment did not remove all of the acetic acid with the result that during analysis the value for the acetic acid could be too high. This effect was overcome by using an amplifier attenuation such that the acetic acid peak from the solvent was insignificant. Having achieved a method of determining free fatty acids it was now possible to develop a method for their isolation from solutions that also contain esters and alcohols.Standard solutions made up in 40 per cent. VjV ethanol were used for this work. Before a method could be considered to be satisfactory the following conditions had to be fulfilled (i) hydrolysis of esters must not occur during any stage of the method; (ii) complete removal of the non-acidic components must be achieved in order to reduce the number of components in the final solution; and (iii) the composition of the acids in the final solution must be truly representative of that in the original solution. This involves converting the acids into their sodium salts by the addition of alkali (pH 8 * 7 ) evaporation of the solution to dryness on a boiling water bath followed by dissolution of the residue in acetone - water - formic acid solution.The liberated free acids are then analysed by injecting the solution into the gas chromatograph. I t is possible therefore to concentrate the acids to any desired level depending on the volume of acetone - water - formic acid solution used. I t was found that the ratio of ethanol to water could be critical during the conversion of the free acids into their sodium salts. This arose when too much ethanol was present during the addition of alkali which tended to suppress the ionisation of the free acids with the result that the un-ionised acids are not converted into their sodium salts. Provided that sufficient water was present so that a ratio of at least 1 1 was obtained no difficulties were encountered. When alkali was added to the sample to pH 8.7 no hydrolysis of esters was detected.The analysis of standard solutions containing all the acids from acetic to lauric acid showed that the method was quantitative. This method for analysing the free fatty acids from acetic to lauric acid has been in continuous use for several years and it has been found that the column requires replacing only every 8 to 10 months. It was decided to isolate the free fatty acids by as simple a method as possible. All of the non-acidic components were removed during the evaporation. REFEREKES 1. 2. 3. 4 . 6. 6. 7 . 8. 9. 10. 1 1 . \-orbeck 11. L. Blattick L. R. Lee F. A and Pederson C. S. Analyt. Chew?. 1961 33 1512. Xppleby A. J. and Rlapne J. E. O. J . Gas Chiomat. 19G5 5 211. -,- Ibzd. 1965 5 26G. Cromell E. A and Guymon J .F. A r n e ~ . J. Eizol. Vitic. 1969 20 155. Craig JI. Tulloch A. P. and Marty N. L. J . "Imw Oil C h e w Soc. 1963 40 GI. llahadevan Y. and Stenroos L. A?znZyt. Ckenz. 1967 39 1652. Aevogiupla Res. Sotes 19G5 6. Hindin E. May D. S. Macdonald R. and Dunstan G. H. I'17at. Scwngi. W h s 1964 3 92. Mijake H. IIitsuyuki >I. and Tadashi 11.. .4+znZjd. Ckcw. l9G13 40 113. Baker R. A J. Gas Chroinat. 1966 4 418. .Ickman R . G. and Burgher R. D. A?ialyt. Chew. 1963 35 64i. August 1973 ASPECTS O F CHROMATOGRAPHY Affinity Chromatography BY K. W. WILLIAMS (Pharmacin (G.B.) Limited Eali+zg Londaui 16'6 5.5s) 215 UNLIKE other chromatographic techniques which effect separation between species on the basis of physico-chemical differences such as charge and molecular size affinity chromato- graphy utilises the functional characteristics of species in order to separate them.Many examples of substances that stand complementary to each other in reactivity are known in biochemical systems for example enzyme and inhibitors antigen and antibody and steroid and binding proteins. The principle of affinity chromatography can be illustrated by con- sidering the purification of an enzyme from a complex mixture. The specific inhibitor for the enzyme is insolubilised and packed in a chromatographic column the complex mixture is then run through the column and only the enzyme is retained by virtue of its specific interaction with the inhibitor. -4fter the other components have been eluted the eluting agent is changed so as to split the inhibitor - enzyme complex and the enzyme is then washed out.The insolubilised inhibitor affinity column is ready for re-use. Affinity chromatography is not a new technique and the fundamental principles involved can be found in the literature in 1910l and 1916.2 Its slow development can be attributed mainly to a lack of suitable insolubilisation techniques and the real renaissance can be seen in the middle 1960s. IKSOLUBILISATION TECHNIQVES- Four fundamental methods exist for the insolubilisation of species in the preparation of affinity supports. (a) Adsorption to a physical support. The major difficulty is that the operating limits of this type of material appear to be limited and leaching of the insolubilised material can be high. Further as the matrix is by definition active non-specific interactions with species in sample mixtures can be high.( b ) Entrapment within a polymer mesh3 offers the advantage of minimal conformational changes to insolubilised materials but leaching can be high and there is a possibility of restric- ted access to the insolubilised species. (c) Crosslinking with a bifunctional reagent such as glutaraldehyde4 or ethylmaleic anhydride. The major disadvantages are that the presence of unreacted functional groups can result in a matrix with high non-specific binding properties. Additionally column properties of supports prepared in this way are generally poor. ( d ) Covalent coupling to an insoluble support. This is generally accepted as the best method for preparing affinity materials. CHOICE OF SUPPORT- -4 wide range of materials are available for use as supports and it is possible to choose the most suitable support for a given application.The major requirements of a matrix for use in affinity chromatography of biopolymers can be summarised as (a) hydrophilic ( b ) insoluble; (c) stable under the conditions to be used; (d) have low non-specific adsorption properties; (e) macroreticulate structure ; (f) good column properties and (g) capable of easy modification for derivative formation. Among the many materials used as matrices are glass beads polystyrene beads celluloses polyacrylamides Sephadex bead-form agaroses ( e g . Sepharose) nylon tubes ethylmaleic anhydride copolymers and ultrafilter membranes. With such a wide range of supports available it would be foolish to say that one material is better than another and each applica- tion should be considered separately.However the bead-form agaroses have found very wide application in the biopolymer field and display all of the features considered to be desir- able in an affinity support. COC'PLIXG REACTIOX- materials to the matrix is virtually limitless. As with matrices the choice of the reaction to be used in order t o covalently couple The determining requirements are a stable 216 ASPECTS OF CHROJIATOGRAPHY CPvoc. SOC. dnalj’t. Chew. link minimum loss of functionality of the insolubilised product and ease of reaction. Some of the reactions used in insolubilisation of proteins and ligands are with cyanogen b r ~ n i i d e ~ periodate oxidation,6 carbodiinlide,’ triazine,8 cyclic ~ a r b o n a t e ~ diazotisation,1° hydrazidell and a ~ y l a t i o n .~ The cyanogen bromide reaction developed by Porath and his group has achieved widespread use. The reaction chosen will depend on the nature of matrix to be used and free groups present on the material to be insolubilised. The usual reaction sequence is activation or derivitisation of the gel; coupling of a ligand or protein; and blocking of unreacted groups on the matrix. The amount of material that can be insolubilised by a given coupling reaction depends on a number of factors such as the nature of the species being insolubilised its concentration in the reaction medium and the matrix being used. However it should be realised that high substitution of a support is meaningful only in terms of how much of the functional character of the insolubilised material is retained. Thus Axen and Ernback12 found considerably less activity of bond chymotrypsin towards S-acetyl-L-tryosine methyl ester when Sephadex or cellulose was the matrix than when Sepharose was used although less chymotrypsin was bound to the Sepharose.EXPERIXEKTAL- The choice of column size and type packing of the column sample application and run- ning of the column are governed by the same parameters operating in gel filtration and ade- quately described elsewhere. Unbound species are eluted with a single bed volume of starting buffer but adsorbed materials generally require a different buffer system. In many instances an increase in ionic strength or shift in pH or a combination of the two is sufficient to cause dissociation of the complex and subsequent elution. Gradient elution has been used to separate species on the basis of their affinity for the insolubilised material.Additionally specific eluting agents such as haptens inhibitors and other competing molecules have found application. For example Porath and Sunderberg13 separated chymotrypsin and trypsin from a pancreatic extract by adsorption to a soyabean trypsin inhibitor - Sepharose column a t pH 7.8; chymotrypsin was eluted with 0.013 RI tryptamine at pH 7.8 and trypsin a t pH 3.0. SPACERS- In some instances particuhrly with small ligands loss of activity after insolubilisation occurs. This can be viewed 2 s resulting from steric hindrance due to the proximity of the matrix. In these instances the placing of a hydrocarbon “spacer” arm between the ligand and matrix is necessary. The optimum length for the spacer varies with the ligand type but generally C to C is used.The use of a spacer has the added advantage of forming a deriva- tive of the matrix permitting the use of a wider range of coupling techniques in the insolubilisa- tion process. APPLICATIONS- Affinity chromatography is a selective specific separation procedure particularly suited to the separation of species present in low concentration and complex mixtures. Among the substances purified are enzymes inhibitors steroids antibodies and antigens. The isolation and study of binding sites in membranes has also been performed and affinity chromatography has been used in the resolution of racemic mixtures. Future applications are concentrating on the development of group specific supports such as Con A - Sepharose which is specific for &-D-mannOSyl X-D-ghCOsyl and sterically similar residues.Among other group-specific supports are NAD+ and AMP - matrix materials. I t is also possible to visualise the insolubilisation of complexing and chelating agents for use in an analytical mode by the chemist. Although most work with affinity chromatography has so far been concentrated in the biochemical field it can only be a matter of time before the chemist applies his ingenuity to this area. August 19731 ASPECTS OF CHROMATOGRAPHY 217 OTHER APPLICATIONS- Although not strictly related to affinity chromatography three other areas of application for insolubilised ligands and macromolecules can be mentioned. (1) Radioimmunoassay which has established itself as an extremely accurate and sensitive technique for the determination of potein and steroid levels in fluids.(2) Enzyme reactor systems based on insolubilised enzymes permitting a more manageable reaction mixture and re-use of the enzyme. (3) llacromolecular probes where attachment of an active species to a matrix or macro- molecule allows in some instances better visualisation of processes involved in cellular reactions. 1. 2. 3. 4. 6. 8. 9. 10. 11. 12. 13. 0. 1 REFEREXES Starkenstein E. Biochem. Z. 1910 24 210. Selson J . M. and Griffin E. G. J . Awzer. Chenz. Soc. 1916 38 1109. Jlosbach K . and Larsson P. O. BiotechnoZ. Bioengizg 1970 12 19. Levin Y. Pecht bI. Goldstein L. and Jlatchalski E. Biocizemisfrji 1964 3 1905. Axen R. Porath J. and Ernback S. Nature Loizd. 1967 214 1302. Sanderson C. J . and TS’ilson D. V. Immulzology 1971 20 1061.Cuatrecasas P. .I. Biol. Chem. 1970 245 3069. Surinov B. P. and llanoilov. S. E. Biokhimiya 1966 31 387. Barker S. A. Tun H. C. Doss S. H. Gray H. J . and Kenncdy J . Y. Carbohyd. Res. 1971 Barker S. X. and Somers P. J . Ibid. 1968 8 491. Inman J . K. and Dintzis H. JI. Biochrmistry 1969 8 4074. .\xen R. and Ernback S. Eur. J . Biochem. 1971 18 361. Porath J. and Sunderberg L. in Hare >I.) Editor “The Chemistry of Biosurfaces,” T’olume 2 17 471. 1 9 i 2 . The Potential for High-speed Liquid Chromatography BY JOHK H. KNOX AROLXD 1969 a dramatic advance occurred in the technique of liquid column chromatography that resulted in analyses by this method being carried out approximately 100 times more rapidly than had prcviously been thought possible. The driving force for the advance was undoubtedly an understanding of the mechanism of chromatography which arose from the study of gas chromatography.Although Martin and Synge in 1941 were already aware that increased speed in liquid column chromatography required the “use of very small particles and a high pressure drop across the length of the column,” most practitioners of this tech- nique were unaware of the theoretical background and so were unable to make substantial improvements. In contrast the practitioners of gas chromatography including many chemical engineers and physical chemists developed a thorough understanding of the mech- anism of the process. This understanding confirmed and amplified Martin’s original insight and led to the development by Kirkland and Huber of high-pressure systems that operated a t pressures up to 3000 p.s.i.with particle sizes in the region of 30 p m and which incorporated sensitive post-column detectors analogous to those used in gas chromatography. .Although the mathematical details of the theory of chromatography are complex the broad principles are straightforward. For efficient chromatography band dispersion must be minimised. In liquid column chromatography dispersion arises largely from the finite rate of equilibration or mass transfer between the mobile and stationary phases. This causes the part of the solute band in the mobile phase to move slightly ahead of the part in the stationary phase. The slight displacement that results leads directly to dispersion of the band as it moves along the column. In order to minimise the dispersion the displacement must be minimised.It is easy to show that the displacement is equal to the product of themobile phase velocity and the relaxation time for the equilibration process. Efficient chromato- graphy therefore depends on fast mass transfer between the mobile and stationary phases. (Departknext of Chemistry Unioevsity o j Ediizbtwgh Ediizbzivgh) 218 ASPECTS OF CHROMATOGRAPHY [PYOC. SOC. Analjlt. Chem. This can be encouraged either by using very sniall particles where diffusion distances are small or by using the larger particles that have an impermeable core surrounded by a thin porous layer. It is furthermore important to avoid materials that bear polymeric stationary phases in which diffusion is very slow or materials that possess very fine pores which hinder the diffusion of molecules in and out of the particles.It is interesting to note that the tend- ency to use extremely small particles was first observed in ion-exchange chromatography and was a direct result of the very slow mass-transfer rates in polymeric resins. In the future we are likely to see much faster ion-exchange nlaterials developed in which materials of high surface area are covered with an essentially monomolecular layer of ion-exchanging groups that are readily accessible to the mobile phase. hIodern liquid chromatography in which high pressures small particles and sensitive detectors are used can be carried out in all the classical modes namely liquid - solid adsorption chromatography liquid - liquid partition chromatography liquid - solid solution cliromato- graphy ion-exchange chromatography and exclusion or gel-permeation chromatography.The same equipment can generally be used for all forms and is therefore x'ery versatile. A problem that the modern chromatographer often has to solve is which of the many forms of liquid (and indeed gas) chromatography he should use for any particular separation problem. The first determining property of a solute that should be considered is relative molecular mass or size. Compounds of high relative molecular mass are best separated by exclusion chroma- tography in which a porous material with pores of dimensions close to those of the molecules to be separated is used. Pores are not of course uniform but have complex shapes and exclusion is best considered on the basis that the centres of large molecules cannot approach within one molecular radius of any adjacent surface.In exclusion chromatograph>- the largest molecules emerge first from the chromatographic column. If the molecules are relatively small say with relative molecular masses below 1000 some form of retentive chromatography is indicated. If the substances are readily vaporised without decomposition gas chromatography should be used. If they are salts or can readily form ions for example if they are amines or organic acids ion-exchange chromatography is indicated as the first choice. However ion-exchange chromatography is generally slow and it is always advisable to examine whether or not for an organic substance such as an amine or carboxylic acid adsorption chromatography would be more suitable. In adsorption chromatography the adsorbent surface can be thought of as a highly polar material that will retain polar compounds (that is compounds susceptible to hydrogen bonding) and the eluting agent is usually an organic solvent whose polarity is chosen so as to give a reasonable elution time.An alternative to adsorption chromatography is liquid - liquid partition chromato- graphy in which a liquid stationary phase is supported on a porous material. Partition of the solute then occurs between the eluting agent and the fixed liquid phase. As the two phases must by definition be immiscible one of them must be a relatively polar liquid for example water an alcohol or a cyanoether while the other must be relatively non-polar for example an ether or a hydrocarbon. The stationary phase may be chosen to be either the polar phase (normal liquid - liquid chromatography) or the non-polar phase (reverse-phase liquid - liquid chromatography).Generally the form of liquid - liquid chromatography to be used is selected by choosing a stationary phase in which the solute is moderately soluble and a mobile phase in which it is less soluble. A recent addition to the materials available for high-speed liquid chromatography are the chemically bonded stationary phases. These are prepared by various chemical techniques that link the stationary phase by covalent chemical bonds to the surface of an adsorbent such as silica. In some the layer of bonded stationary phase is monomolecular and such materials are essentially modified adsorbents. In others a polymer for example a silicone is bonded to the surface; here the partitioning process is more akin to liquid - liquid partition and the appropriate name for the chromatographic technique is liquid - solid partition chromato- graphy.Yirtually all compounds can be analysed by liquid chromatograph!. even gases (as solutions in a solvent) but detection may be a problem as the only completely satisfactory detector at the present time is the ultraviolet photometer. This detector can discriminate at the level of 1 part in 109 of solute in the eluting agent if the molar extinction coefficient is about 101. Bulk property detectors such as the refractive index monitor are about 1000 times less sensitive and are generally useful only for the detection of major components in August 19731 ASPECTS OF CHR03IATOGRAPHY 219 mixtures or in exclusion chromatography where sample dilution by the column is much less than in retentive chromatography.The complete range of liquid-chromatographic techniques now at the disposal of the analytical chemist can be carried out a t speeds similar to those obtained in gas chromato- graphy. Although the equipment is currently rather expensive the real cost will undoubtedlj- decline until high-speed liquid-chromatographic equipment is little more expensive than gas- chromatographic equipment of comparable specification. High-speed liquid chromatography is already showing itself to have wide application in organic chemistry and pharmaceutical chemistry and can confidently be expected to have a dramatic impact on biochemistry. Undoubtedly the problems arising in the last field will be much more difficult to solve than those in the analysis of chemical preparations but this simply means that the delay until high-speed liquid chromatography is widely adopted in biochemical laboratories will be greater than in laboratories specialising in organic fine chemicals.One area where high-speed liquid chroma- tography should make an immediate impact is that of synthetic and mechanistic organic chemistry where its ability to distinguish and quantify closely related compounds mill be invaluable. Determination of Isotopically Labelled Lipids by Combined Thin-layer and Gas - Liquid Chromatography BY W. W. CHRISTIE (Ha?z?iah H e s e a i d Institute A y v Scotland) THE wide variety of lipid classes found in animal tissues can be separated by thin-layer chroma- tography and quantified b l r gas chromatography of the methyl ester derivatives of the com- ponent fatty acids with an added internal standard-normally an odd-chain fatty acid such as pentadecanoic or heptadecanoic acid.l It is necessary to add the internal standard a t an early stage in the procedure and to ensure quantitative esterification so that there are no selective losses of fatty acids relative to the internal standard.By applying readily derived factors the masses or molar amounts of each lipid present can be calculated. The procedure is as accurate as others in current use and has the additional merit of allowing the simultaneous determination of lipid and fatty acid composition. If the transesterification reaction is per- formed in the presence of the thin-layer adsorbent the precision of the procedure is improved.z The scope of the method has now been extended to include the analysis of lipids labelled with hydrogen-3 or carbon-14 in the fatty acid moiety by using labelled internal standards.Odd-chain fatty acids labelled with hydrogen-3 or carbon-14 were not available commercially but ( l-14C)-pentadecanoic acid was synthesised by standard procedures3 from tetradecanoic acid via the mesplate of the corresponding alcohol and potassium (14C)-cyanide while (10,ll- 3H)-heptadecanoic acid was synthesised similarly with (9,10-3H)-hexadecanoic acid as the starting material. Thin-layer chromatographic separations were performed as before1p2 but the radioactivity present in the bands was determined by suspending the adsorbent in Unisolve 1 (Koch-Light Laboratories Ltd.) for liquid scintillation counting.In addition two gas - liquid radio-chromatographic systems were available a preparative gas chromatograph that permitted Sb per cent. recovery of labelled fatty acids for liquid scintillation counting and a continuous flow proportional counting system (Panax Ltd.) that allowed the simul- taneous determination of the mass and radioactivity of the samples but in which because of tlir size of the counting cell the measurable activity was only 15 per cent. of that added. The former system was used with low-activitj. samples and the latter with tliose of higher activit!-. The nature of the approach to the problem of analysis of lipids containing fatty acids labelled with carbon-14 then depended on the complexity of the sample and the amount of radioactivity present.Simple in vitvo systems consisting of adipose tissue homogenates4 or liver slices from rats were used to study the incorporation of single fatty acids labelled with carbon-14 into various lipid classes. In the simplest case when the added fatty acid was not modified by chain elongation or desaturation by the iiz aitro system the internal standard labelled with hydrogen-3 was added to each lipid class separated by thin-layer chromatography; 220 OBITUARY [Pvoc. s o c . Anal$. CItenz. the amount of each component was determined as before and the amount of radioactivity corrected for all losses was obtained simply by determining the hydrogen-3 to carbon-14 ratio by liquid scintillation counting of the methyl esters. \T.'hen more than one labelled fatty acid was present in the sample however it was necessary to add the internal standard labelled with carbon-14 and to determine the amount and radioactivity of each component relative to the values for the internal standard by one of the gas-chromatographic procedures.As both the total activity and the specific activity of the standard were known it was possible t o calculate values for the other labelled components. Losses that inevitably occurred during structural analyses of lipids containing fatty acids labelled with carbon-14 were also compensated for by adding prior to the analysis a labelled internal standard. For example in determinations of the positional distributions of labelled fatty acids in triglycerides by stereospecific r n e t h ~ d s ~ ~ ~ tri-( l-14C)-pentadecanoin or tri- (10,l l-3H)-heptadecanoin synthesised from the appropriate fatty acid and glycerol was added as internal standard. The amount and activity of the labelled fatty acid originally present in each position of tlie triglyceride were then determined by relating the activity of tlie acid to that of the internal standard in each of the products of tlie reaction. REFERENCES 1 . 2. 3. 4. 6 6. Christie W. W, Noble R. C. and RIoorc J. H. Aizalyst 1950 95 940. Christie \IT. \V. I b i d . 1972 97 221. Banmann \V. J and Xangold H. I<. J . Lipid Res. 1968 9 285. Angel A and Roncari D. X. K. Biochim. Biophys. Acta 1967 137 464. Christie \Ir. W. and Noore J . H. Ibid. 1969 176 445. ->- Ibid. l9i0 210 46.

ISSN:0037-9697    

DOI:10.1039/SA9731000210

出版商:RSC    

年代:1973

数据来源: RSC

摘要:

220 OBITUARY Obituary THOMAS EDII’ARD \ITALLIS T. E. \TALLIS D.Sc. Ph.D. F.P.S. F.L.S. F.R.I.C. “Tommy JVallis” to all his friends has died at the age of 96 and one of the most eminent pharmacognosists of all time has gone. He was a member of the Society from 1915 to his death and of the Joint Pharmaceutical Analysis Group from its inception. Dr. IVallis was born in Islington in 1856 and was educated in London attending Birkbeck College from 1893 to 1900. During his qualifying career he obtained several medals in chemistry materia medica and botany. His B.Sc. of London University was Fassed in 1900 and he became a Fellow of the Royal Institute of Chemistry in 1906. His earlier teaching career was in chemistry he then became an assistant from 1913 to 1918 to Thomas Tickle Public Analyst for Devonshire.During his stay there he devised the method of quantitative microscopy by the use of lycopodium spores that has found acceptance by analysts the world over. In 1926 Dr. Wallis was appointed reader in pharmacognosy at London L-niversity and in 1927 he became curator in the museum of the Pharmaceutical Society. President of the Royal Xcroscopical Society in 1951-55 he was also an examiner in pharmacognosy to the Universities of London and \Tales and an external examiner in pharmacy and pharmacognosy to Queen’s L‘niversity Belfast. He was the author of the three well known standard textbooks “Textbook of Pharmacognosy,” “Practical Pharniacognosy” and “Analytical Microscopy.” He was Chairman of the British Pharmaceutical Conference from 1942 to 1944 and he con- tributed a paper t o the Conference as recently as 1967 in his 91st year.He was a member of various British Pharmacopoeia Committees between 1932 and 1953 and he worked on every edition of the British Pharmaceutical Codex from 1930 to 1968. Although his international fame was in pharmacognosy Dr. \Vallis was proud to be considered an analyst which was justified from the exacting standards he required his careful observation and his meticulous attention to detail. It was his ability as a microscopist and his demonstration of the microscope as an important analytical tool for which he will be most remembered by analysts. He had the ability to observe and to describe and moreover the great facility of accurate detailed drawing as those familiar with his textbooks are well aware. He was apprenticed to his father a pharmacist. August 19731 SILVER MEDAL 221 Dr. \ Y a k was a shy most modest and gentle man with a quiet sense of humour who took great pains to help anyone who asked for his assistance. He was active right up to his death and it is difficult to realise that so familiar a figure is no longer with us. D. C. GARRATT

ISSN:0037-9697    

DOI:10.1039/SA9731000220

出版商:RSC    

年代:1973

数据来源: RSC