ISSN:0144-557X    

DOI:10.1039/AP98825FX013

出版商:RSC    

年代:1988

数据来源: RSC

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ANPRDI 25(4) 105-140 (1988) April 1988 Analytical Proceedings Proceedings of the Analytical Division of The Royal Society of Chemistry CONTENTS 105 Annual General Meeting of the Analytical Division 105 New Member of Council 106 New Director of Metropolitan Police Laboratory 107 Correspondence 108 Ronald Belcher Memorial Lecture: 'Nylon Mesh and Platinum Wire Enzyme Amperometric Electrodes' by G. S. Sanghera 111 Silver Medal Lecture: 'A Judicious Mixture of Reaction Chemistry and Instrumentation' by Julian F. Tyson 'Chemical Typing of Cereal Grains' by R. Cook 'Sampling of Soils and Plants for Trace Element Analysis' by M. L. Berrow 'Action from Analysis: Challenges for Agricultural Analysis' by D. Atkinson 'Design of a Scottish Soil Fertility Information System' by A.H. Sinclair, G. A. Reaves 115 SUMMARIES OF PAPERS 115 Analysis in Agricultural Sciences: 115 116 118 120 and K. W. M. Brown 'The Use of Rapid Tests for Measurement of Plant Nutrient Status' by I. G. Burns 'Approaches to the Analysis of Animal Feedstuffs' by A. Smith 122 124 127 The Quest for Sensitivity, Specificity and Reliability of Diagnostic Reagents-a Biological 127 128 129 Answer: 'Chlamydia Detection with Monoclonal Antibodies' by Caroline Ash 'Cloned Nucleic Acid Probes for Detection and Identification of Legionellae' by N. A. Saunders, N. Kachwalla, T. G. Harrison and A. G. Taylor 'Development of a Diagnostic Reagent for the Detection of Adenovirus Antigen by Immunofluorescence' by I. R. Sharp and A. Bailey 132 Equipment News 136 Conferences and Meetings 136 Courses 137 Publications Received 139 Analytical Division Diary Typeset and printed by Black Bear Press Limited, Cambridge, England

ISSN:0144-557X    

DOI:10.1039/AP98825BX015

出版商:RSC    

年代:1988

数据来源: RSC

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ANALYTICAL PROCEEDINGS, APRIL 1988, VOL 25 New Member of Council A. M. C. (Tony) Davies joined the Council in November, 1986, as the Chairman of the East Anglia Region. He was born in Norwich in 1938 and attended several schools before being educated at the King Edward V1 School, Norwich. On leaving school he entered the Royal Air Force College, Cranwell, but left the RAF after failing to persuade any instructor to allow him to fly solo twice! He joined Glaxo Laboratories at Greenford in 1959, where he received expert analytical tuition while his formal chemical education was acquired at Brunel (in its CAT period). Part of his experience at Glaxo was to work with the third Pye Argon Chromato- graph which was the beginning of twenty years’ labour in chromatography. On gaining his HNC he started work in the food industry with the HP group as works chemist in a canning factory.Two and half years later he was attracted to the idea of research and joined the Research Depart- ment of J.& J. Colman (Colman Foods) and worked on particle size analysis. gas chromatography and amino-acid analysis. He was also given the time to take an HNC Endorsement course in Advanced Analytical Chemistry which resulted in his first paper [on the analysis of the (then) new artificial sweetener Cycla- mate]. It was through ion-exchange chro- matography that he was able to exchange industrial research for government funded research, because in 1972 he joined the protein chemistry group at the Food Research Institute to work for Dr. (now Professor) D .S . Robinson and Professor R. L. M. Synge. After the departure and retirement of his eminent tutors he took on responsibility for NMR and AAS in an Analytical Services Group in the reorgan- ised FRI under Professor R. F. Curtis. Part of the remit of the group was to investigate new methods of analysis and, after brief flirtations with automatic de- vices and isotachophoresis, he became involved with near infrared (NIR) analy- sis. This became such an obsession that it took over the whole group and he has been associated with much of the recent interest in NIR analysis which has made106 ANALYTICAL PROCEEDINGS, APRIL 1988, VOL 25 him a regular visitor to the USA. Cuts in government funding coupled with new management at the Institute of Food Research, Norwich, Laboratory have recently closed his group.Following his ”voluntary” redundancy Mr. Davies joined Oxford Analytical Instruments in December, 1987, as a consultant in food analysis. He was recruited to the RSC/AD East Anglia region by Tony Croft and has served as Assistant Secretary, Secretary and Treasurer and Vice-chairman before election to his current office. He was granted his MRSC in 1981 and his FRSC in 1988. He was invited to join the inaugural committee of the Division‘s Molecular Spectroscopy Group and is the (nominated) Vice-chairman. He was Chairman of the organising committee for the “spectroscopy Across the Spectrum” (SAS) Conference and is a member of the SAC ‘89 organising committee. Outside the RSC, he has been associated with the international development of NIR analy- sis and is the Honorary Secretary and Treasurer of the International Committee for Near Infrared Spectroscopy, whose first conference formed part of the SAS conference. Tony married his Norwich-born Welsh wife, Megan, in 1960 and their daughters. Siin and Rhiannon. are legally adults but are in fact still dependent graduate and undergraduate students. His spare time activities include the Norwich Welsh Society, the Norwich Branch of the Inland Waterways Association and holidaying on canals or in France (or both). He also claims to play squash but his increasing weight suggests that this is not a regular activity .

ISSN:0144-557X    

DOI:10.1039/AP988250105b

出版商:RSC    

年代:1988

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106 ANALYTICAL PROCEEDINGS, APRIL 1988, VOL 25 New Director of Metropolitan Police Laboratory The new Director of the Metropolitan Police Forensic Science Laboratory is Dr. Brian Sheard. He will head a team of 210 scientists who deal with 26 000 investiga- tions a year from their headquarters in South London. Dr. Sheard, aged 46, joins the labora- tory after 14 years with ICI’s Pharmaceut- ical Division at Alderley Park, Cheshire, where he was a research manager. After taking a degree in chemistry and gaining a PhD in biophysics from Oxford, Dr. Sheard worked for Unilever Research at Port Sunlight in Merseyside. He then spent five and a half years in the United States of America working on research for the Bell Telephone Laboratories and later working as Assistant Professor at the Albert Einstein College of Medicine in New York.Until recently. he was Chair- man of a joint research venture between four major industrial companies and the Science and Engineering Research Coun- cil. providing funding for research in politan Police in the front rank of forensic science and finds the biggest excitement in forensic science at the moment to be DNA profiling. a technique that his staff are currently introducing to the Labora- tory. In 1980 Dr. Sheard appeared with friends as a contributing finalist in the BBC’s Mick Burke amateur film series after filming their expedition to remote canyons on the Navaho Indian Reserva- universities. tion. He is married with two daughters. Dr. Sheard hopes to keep the Metro- by T.P. Coultate, Polytechnic ofthe South Bank.This book ives a detailed account of the chemistry of the principlal substances of which our food is composed. Both the macro-components, the carbohydrate, lipids and proteins, which can be classified by their chemical structures, and the micro-components, the colours, flavours, vitamins and preservatives, which are classified in terms of function, are considered. Throughout the book, Dr. Coultate’s theme is the relationship between the chemical structure of a substance and its contribution to the properties and behaviour of foodstuffs - whether observed in the laboratory, the factory, the kitchen o r the dining room. Contents: Introduction; Carbohydrates; Lipids: Proteins; Colours; Flavours; Vitamins: Presewatives; EEC Numbers for Food Additives; Subject Index. This book will be of particular benefit to students and teachers of food science and related courses in universities, colleges of further education and schools. RSC Paperback (1984) Softcover 202pp ISBN 0 85 186 483 X Price E5.95 ($1 1.00) ORDERING: RSC Members should send their orders to: The Royal Society of Chemistry, Membership Manager, 30 Russell Square, London WC 1B 5DT, UK. Non-RSC Members should send their orders to: The Royal Society of Chemistry, Distribution Centre, Blackhorse Road, Letchworth, Herts SG6 lHN, UK. Information Services

ISSN:0144-557X    

DOI:10.1039/AP9882500106

出版商:RSC    

年代:1988

数据来源: RSC

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ANALYTICAL PROCEEDINGS, APRIL 1988, VOL 25 107 Correspondence Correspondence is accepted on all matters of interest to analytical chemists. Letters should be addressed to the Editor, Analytical Proceedings, The Royal Society of Chemistry, Burlington House, London, W1V OBN. Determination of Oil or Fat in Feeds and Food Sir, The challenge made in my letter of August, 1986 (accompan- ied by a diagram of the Manley and Wood apparatus) not having been answered during the subsequent eighteen months I submit that it is correct to conclude that 2 h is indeed the maximum time (instead of the lengthy official 6 h ) in which to determine oil in feeding stuffs. or for that matter, fat in cocoa. A former deputy, later himself an official agricultural analyst. writing to me on the subject stated that it was incomprehens- ible that the Manley and Wood method had not been officially adopted.Agricultural analysts may well wonder why the position of the framers of the present Amendment Regulations is that apparently in their view the old (Soxhlet) method is to be preferred and time saving counts for little or nothing. Three or four years ago, to a MAFF official, I advocated a collaborative trial to place the case on a sound basis, but to date there is no indication that such has ever been carried out. References 1. 2. Manley, C. H.. and Wood. E. G.. A n r r l ~ ~ t , 1945. 70. 173. The Feeding Stuffs (Sampling and Analysis) (Amendment) Regulations 1985. HM Stationery Office, London. 1985. C. H. Manley "Ifflej. " 3 Great Brockeridge, Westhirrj-on- Trym, Bristol BS9 3 T Y Sir, I refer to C.H. Manley's letter in Analytical Proceedings in August, 1986, and the correspondence published above, to which I have been asked to reply. For those who have not had the privilege of meeting him, Mr. Manley is a much respected nonagenarian. former Public Analyst to the City of Leeds, with an unquenchable interest in analytical chemistry. The issue concerns the failure on the part of the EEC to recognise the potential application of the Manley and Wood extraction apparatus for the rapid determination of oil or fat in animal feed or foods by direct solvent extraction (Manley. C. H . , and Wood, E. G., Analyst, 1945, 70, 173). From enquiries I have made it is generally recognised that "hot" solvent extraction techniques such as the Manley and Wood and other "straight through" extractors are generally faster and equally as efficient as "cold processes," such as the Soxhlet apparatus, where direct extraction is appropriate.The Feeding Stuffs (Sampling and Analysis) (Amendment) Regulations 1985 were introduced in order to honour UK obligations as a member of the EEC. The principal change incorporates a pre-extraction acid-hydrolysis phase essential for certain feeds to ensure that certain fatty constituents, which would not be extracted directly by solvent alone, are deter- mined. The direct determination of oil and fat. where applicable, is an empirical method which requires for Statutory purposes the method to be specified using standard, readily available equipment. The Soxhlet apparatus has been recog- nised for this purpose for many years.It is a pity that alternative methods, where equivalent efficiency can be shown, may not also be officially recognised. Anthony J. Harrison Chairmiin A n alv tical Methods Com rn ittee ROYAL SOCIETY OF CHEMISTRY: ANALYTICAL DIVISION EAST ANGLIA REGION A meeting on LABORATORY INFORMATION MANAGEMENT SYSTEMS (LIMS) will be held at Smith, Kline and French Laboratories Ltd., Welwyn Garden City, Hertfordshire on May 5th, 1988 The speakers at this meeting will be: Dr. B. McDowaII (SK and F Research Ltd., Welwyn), "The Requirement Specifications for LIMS"; Mr. P. Snowdon (Schering, Saffron Walden)," LlMS (Beckman) in the Pesticide Residue Laboratory"; Dr. A. Wagland (Roche, Welwyn Garden City), "Current and Future Impact of LIMS (Perkin Elmer) in Quality Control"; Mr. I. Smith (Glaxo, Ware), "HP LABSAM in an Integrated System"; and Mr. A. D. Henderson (SK and F Laboratories Ltd., Welwyn Garden City), "LlMS (Beckman) for the End User." For further details contact Mr. A. D. Henderson, Analytical QC, Smith Kline and French Laboratories Ltd., Mundells, Welwyn Garden City, Hertfordshire AL7 1 EY.

ISSN:0144-557X    

DOI:10.1039/AP9882500107

出版商:RSC    

年代:1988

数据来源: RSC

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1 OH ANALYTICAL PROCEEDINGS, APRIL 1988, VOL 25 Ronald Belcher Memorial Lecture ~ ~~~~~ The fourth Ronald Belcher Memorial Lecture was given by Dr. G. S. Sanghera at the Research and Development Topics in Analytical Chemistry meeting of the Division held on July 8th and 9th, 1987, at The University of Strathclyde, Glasgow. Nylon Mesh and Platinum Wire Enzyme Amperometric Electrodes G. S. Sanghera" Department of Applied Chemistry, Redwood Building, This paper is concerned with the immobilisation of enzymes for use with amperometric electrodes set up in a flow injection analysis (FIA) system. The first system described consists of single-enzyme electrodes for glucose and cholesterol based on glucose oxidase and cholesterol oxidase , respectively, immobi- lised on nylon mesh' and held over a platinum electrode.The anodic decomposition of hydrogen peroxide (produced in the enzymic reaction) was monitored at a platinum working electrode. The second approach involves the use of bi-enzyme elec- trodes, wherein glucose oxidase - peroxidase and xanthine oxidase - peroxidase are again immobilised on nylon mesh for the determination of glucose and xanthine - hypoxanthine, respectively. The enzymically generated hydrogen peroxide was monitored amperometrically after its peroxidase-catalysed reaction with hexacyanoferrate( 11): (1) and the generated hexacyanoferrate(II1) is reduced at a platinum electrode at a low applied potential of -100 mV vs. Fe(CN)63- + e- --j Fe(CN)6A- . . . . (2) The electrodes were tested for the determination of glucose in serum and hypoxanthine in fish meat.The nylon mesh membrane enzjme electrodes described were used in an FIA system incorporating a Stelte cell modified2 for FIA. The modified platinum wire approach involves the direct covalent attachment of glucose oxidase to an activated platinum surface to form a "micro" glucose enzyme electrode. Its response characteristics were determined in an FIA system incorporating a laboratory-built three-electrode amperometric cell, by monitoring the anodic decomposition of hydrogen peroxide. The surface activation of platinum and the sub- sequent immobilisation of glucose oxidase are described. HzOz + 2Fe(CN)6J- + 2H+ + 2H20 + 2Fe(CN)6-+ . . Ag - AgCl: Experimental Immobilisation of Enzyme Glucose oxidase was immobilised on nylon mesh by a modification of the method devised by Hornby and Morris.' Nylon mesh was activated with dimethyl sulphate and reacted with lysine and glutaraldehyde, as previously described.' Enzyme was attached to the modified nylon by dipping the mesh in the appropriate enzyme solution.For single-enzyme membranes the modified nylon was dipped in enzyme (10 mg cm-3) in phosphate buffer (0.1 M, pH 7.0). For bienzyme membranes the modified nylon was dipped in glucose oxidase or xanthine oxidase as appropriate (10 mg cm-3) in phosphate 7_ Present address: Inorganic Chemistry Laboratory. University of Oxford. South Parks Road, Oxford OX1 3QR. UWIST, P.O. Box 13, Cardiff CF7 3XF buffer (0.1 M, pH 7.0) containing peroxidase ( 5 mg cm-j) wherein the oxidase : peroxidase ratio is optimised at 2 : 1 as previously reported.' The final structure may be simply shown as: Nylon mesh 1 + HN=C-NHCOO- H N=CH-(CH2)3-C I/ Enzyme-N Immobilisation of enzyme directly on to platinum wire was based on a simplification of the chemical modification outlined by Yao,J followed by the attachment of glucose oxidase via the bifunctional reagent glutaraldehyde using a procedure similar to that described for controlled porosity glass (CPG) by Masoom and Townshend.5 Thus, platinum wire (length 2.5 cm, diameter 0.1 cm) was activated by successive 10-min immer- sions in hot concentrated nitric, chromic and hydrochloric acids and then electrochemically treated in sulphuric acid (0.5 M) by cycling between 0 and +1.3 V (vs.Ag - AgC1) for 2 h or alternatively overnight." The activated platinum wire was anodised at +2.50 V (vs.Ag - AgCl) in sulphuric acid (0.1 M) for 5 min. The activated - anodised wire was then reacted with 3-aminopropyltriethoxysilane and treated with glutar- aldehyde. After thorough washing, the wire was placed in glucose oxidase (10 mg cm--7) in phosphate buffer (0.1 M , pH 7.0) to obtain an enzyme entity as shown: CH I1 Enzyme-N Apparatus Electrode potentials were controlled and currents monitored with a potentiostat (Metrohm VA, detector E611). A Linear y - t chart recorder (Model 500) was used to record the flow injection analysis signals. The sample was propelled with a four-channel peristaltic pump (Ismatec Model IP4) with sample injections being made with an electrically activated valve (Valco Instruments).All connecting tubing was PTFEANALYTICAL PROCEEDINGS, APRIL 1988, VOL 25 109 (nominal i.d. 1.27 mm). Pump pulsation was reduced with a suppressor placed immediately after the pump. For the nylon mesh enzyme electrodes the detector was a three-electrode modified Stelte cell, with a platinum working electrode, a glassy carbon auxiliary electrode and a silver - silver chloride reference electrode. The working electrode chamber of the detector cell was modified to produce a wall jet for the enzyme electrode as previously described.2 For the platinum wire enzyme electrode a laboratory-made flow- through cell was used. The cell design was such that the reference (silver wire - silver chloride) and the auxiliary (platinum wire) electrodes were placed in a stationary solution of saturated potassium chloride and contacted with the flowing stream by means of a T-junction. A slight back-pressure from a potassium chloride reservoir maintained a stationary phase for the reference and auxiliary electrodes while the working electrode was positioned in the flowing stream.Results Single-enzyme Electrodes Glucose oxidase immobilised on nylon mesh served as a model for the development and optimisation of an FIA system for nylon mesh enzyme electrodes. The effect of pH over the range 5-8 on the glucose response exhibited a maximum between 6.8 and 7.2. Similarly, the effect of flow-rate on glucose response was investigated over the range 0.5-5 cm3 min-1 and the optimum region was found to be between 1.5 and 3.0 cm-' min-I.The optimisation of response for the glucose nylon mesh enzyme electrode has been reported previously.3 The optimised FIA system was calibrated with a set of glucose standards exhibiting linearity over the range 1 x 10-5-3 x 10-3 M glucose with response and wash times each around The FIA system developed for glucose was extended, with limited success. to cholesterol using cholesterol oxidase immo- bilised nylon mesh. The main difficulty was the availability of suitable aqueous cholesterol standards for response character- isation and calibration of the cholesterol enzyme electrode. Several methods were investigated' for the production of suitable aqueous standards, but that adopted involved the use of Triton X-100 surfactant in phosphate buffer (0.1 M , pH 7.0).Surfactant concentrations in excess of 1% V/V suppressed the cholesterol signal and so limited the linearity of the cholesterol enzyme electrode calibration, namely, 10 VM-0.75 mM choles- terol. The linearity may be extended beyond 0.75 mM cholesterol when suitable standards of higher concentration become available. 45-60 S . Bi-enzyme Electrodes Xanthine oxidase exhibits twice the activity towards xanthine compared with hypoxanthine and consequently the optimum response to xanthine was also investigated with the xanthine oxidase - peroxidase bi-enzyme electrode. The effect of pH on bi-enzyme electrodes for glucose and xanthine - hypoxanthine exhibited an optimum pH range between 6.75 and 7.25. The flow-rate vs. peak height profile exhibited an optimum region between 2.75 and 3.25 cm3 min-1.The concentration of the mediator [hexacyanoferrate(II)] was optimised at 1.5 mM for the glucose bi-enzyme electrode and 2.5 mM for the xanthine - hypoxanthine bi-enzyme electrode. The bi-enzyme electrode for glucose exhibited linearity over the range 0.02-3 mM glucose. Analysis of glucose in blood serum gave good agreement with a soluble enzyme test kits according to with a correlation coefficient of 0.997. The xanthine oxidase - peroxidase bi-enzyme electrode exhibited linearity to both xanthine and hypoxanthine over the range 2-100 VM. The deterioration of fish meat quality was [G1~~0~e]electrode = [Glucoselwlub~c tc\t kit -0.219 reflected by increases in hypoxanthine levels that occurred during storage at room temperature. The tests were carried out on four different kinds of fish, namely, rainbow trout.herring. hake and plaice, and the results were compared with those of the spectrophotometric-based method recommended by the Analytical Methods Committee.9 Thus, for the sets of data relating to fresh fish and stored fish together, ( H y p o ~ a n t h i n e ] , ~ ~ ~ ~ , ~ ~ ~ ~ = 1.01 [HypoxanthineIAMc + 4 x l O b . 3 with a correlation coefficient of 0.998. Modified Platinum Wire Prior to calibration of the platinum wire enzyme electrode, its optimum pH range and the effect of flow-rate were investi- gated. The effect of pH on glucose response exhibited an optimum between 5.8 and 6.5. The peak height versrts flow-rate profile increased almost linearly until a limiting value was reached for flow-rates above 3.2 cm3 min-1.A more detailed investigation over the flow-rate range 2-4 cm3 min-l confirmed a limiting value above 3.2 cm3 min-1, with little change for higher flow-rates up to 8 cm3 min- I . The platinum wire glucose electrode was calibrated over the range 0.1-30 mM glucose and a typical chart recorder output and corresponding calibration are shown in Figs. 1 and 2. The calibration was linear over the range 0.1-10 mM glucose with excellent response times (<25 s) and wash times (<30 s). The linear portion of the calibration graph corresponds to Log(current/A) = 0.992 log([glucose]/mM) -3.94 with a correlation coefficient of 0.999. 25 fTlM T 2 . 5 m ~ 200 nA 1 10 mM M c- min Scan Fig. 1. Typical recorder output for the platinum wire enzyme electrode Discussion Effect of pH Enzyme immobilisation will produce a shift in the optimum pH of a given enzyme in a soluble form, as a direct result of constitutional changes imposed on the enzyme.The immobil- isation of enzyme(s) on platinum and nylon mesh yielded110 ANALYTICAL PROCEEDINGS, APRIL 1988, VOL 25 optimum pH values of 6.0 and 7.0, respectively. Studies with solubilised glucose oxidase and xanthine oxidase showed a maximum at pH 5.5 and 4.6, respectively. Immobilisation on nylon mesh produces a shift to a more alkaline pH optimum of 7.0. The effect of pH on glucose oxidase immobilised on platinum wire shows a shift of only 0.5 pH unit to pH 6.0. The shifts in both instances indicate the way in which changes in the microenvironment of an enzyme affect its pH optimum.This example illustrates the pH dependence on the immobilisation procedure and the nature of the support material. Effect of Flow-rate Each of the three described enzyme electrode systems exhibits a slightly different peak height - flow-rate profile. Single- enzyme nylon mesh electrodes exhibit a great sensitivity to flow-rate on either side of the optimum region (1.5-3.0 cm3 min-I). In this region the diffusion of substrate to, and products away from, the enzyme electrode is at an optimum. For bi-enzyme electrodes an optimum response is observed over the range 2.75-3.25 cm3 min-1. The use of a redox mediator reduces the over-all sensitivity of the bi-enzyme electrode to flow-rate compared with single-enzyme elec- trodes. The slight increase in the optimum flow-rate region is due to the measurement of hydrogen peroxide indirectly via a secondary reaction.For the platinum wire an increase in response with increasing flow-rate up to 3.3 cm3 min- * is unexpected and can arise from the proximity of the enzyme to the platinum surface, which may exert steric hindrance on the enzyme. Hence, faster flow-rates and consequently increased substrate diffusion can produce the observed increasing response. I I - 1 0 1 Log ((glucoselim~) Lifetime The lifetime, durability and storage stability are important parameters when considering immobilised enzyme systems. Single-enzyme nylon mesh electrodes exhibit a response to glucose for up to 4 months of intermittent use when stored in buffer (4 "C). Bi-enzyme electrodes have a lifetime of 3 months for intermittent use with storage in buffer (4 "C) for glucose oxidase - peroxidase electrodes.The shorter lifetimes ex- perienced here compared with single-enzyme electrodes may be attributed to the fact that the second enzyme, peroxidase, reduces the surface area taken up by the primary glucose oxidase. The xanthine oxidase - peroxidase electrode had a working lifetime of 2 months under the above-mentioned conditions. The reduced electrode lifetime in comparison with the glucose electrode is probably due to the low activity of enzyme immobilised, namely 1-2 units mg-* for xanthine oxidase compared with 100 units mg-1 for glucose oxidase. The platinum wire enzyme electrode exhibited a response to glucose for daily use with flow injection samples over a period of 10 d.The relatively short over-all lifetimes compared with electrodes based on nylon mesh may relate to the small surface area of the platinum wire and/or the nature of the Pt-0 bonding. Masoom and Townshends reported glucose oxidase activity on CPG of up to 1 year, owing to the much greater surface area of the CPG. The modified platinum wire carries a thin layer of immobilised enzyme and therefore its small size offers the prospect for development as an implantable glucose sensor. The author thanks the Department of Trade and Industry (Laboratory of the Government Chemist) for financial sup- port. Thanks are also extended to his supervisors, Dr. J. D. R. Thomas and Dr. G. J. Moody, for their sustained help and counselling throughout the course of this work.1. 2. 3. 4. 5. 6. References Hornby, W. E., and Morris, D. L., in Weetall, H. H., Editor. "Immobilized Enzymes, Antigens, Antibodies and Peptides." Volume 1, Marcel Dekker, New York, 1975, p. 141. Moody, G. J . , Sanghera, G. S . , and Thomas, J . D. R., Analyst, 1986, 111, 605. Moody, G . J . , Sanghera, G. S., and Thomas, J . D. R.. Anulwr, 1987, 112, 65. Yao, T., Anal. Chim. Acta, 1983, 148. 27. Masoom, M., and Townshend, A , , Anal. Chim. Acta, 1984. 166, 111. Moody, G. J . , Sanghera, G. S . , and Thomas, J. D. R., Analyst. 1986, 111. 1235. Fig. 2. Calibration graph for the platinurn wire enzyme electrode. Numbers above the line indicate response times (s) and those below the line indicate wash times (s) 7. 8. 9. Sanghera, G. S., PhD Thesis, University of Wales. 1987. Trinder, P., J . Clin. Puthol.. 1969. 22, 246. Analytical Methods Committee, Analyst, 1979, 104, 434. NUCLEAR AND RADIOCHEMISTRY July 11th-lfith, 1988, Brighton The Second International Conference on Nuclear and Radiochemistry will be held at the Brighton Metropole Hotel. The programme will consist of seventeen scientific sessions supplemented by nine plenary lectures. A social programme will also be provided. Full delegates registering before June 1st will be charged f220 (compared with f275 after June 1st) exclusive of accommodation. For further information, contact Mr. M. A. Crook, 17 Broughton Road, Otford, Kent TN14 5LY.

ISSN:0144-557X    

DOI:10.1039/AP9882500108

出版商:RSC    

年代:1988

数据来源: RSC

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ANALYTICAL PROCEEDINGS, APRIL 1988, VOL 25 introduction 1 1 1 Silver Medal Lecture The following is the Silver Medal lecture delivered by Dr. J. F. Tyson, the Fourteenth SAC Silver Medallist, at the Research and Development Topics in Analytical Chemistry Meeting of the Division held on July 8th and 9th, 1987, at the University of Strathclyde, Glasgow. A Judicious Mixture of Reaction Chemistry and instrumentation Julian F. Tyson Department of Chemistry, Loughborough University of Technology, Loughborough, Leicestershire, LEI7 3TU I chose a general title for this lecture for several reasons. Firstly. as about 13 o r 14 scientific meetings in 1987 already featured my name amongst the list of speakers and there were. no doubt, several members of the audience who must have felt they had attended all of them, I wished t o say something different in this lecture.Secondly, as atomic spectroscopists have featured prominently as Silver Metal lecturers. I wished t o avoid atomic spectroscopy. Thirdly, as it is well known in educational circles that the attention span of an audience is about 20 min, I wished to present three 20-min lectures on somewhat different topics. However, they are linked by the common underlying theme of “time and the philosophy of analytical chemistry.” The title of the lecture is, in fact, a quotation,’ which in full reads, “The study of chemical reactions is necessary for the future chemist t o learn what chemistry is, in the same manner as learning about laboratory instruments shows him how to apply them. The use of properly chosen reactions in conjunc- tion with an appropriate instrument gives perhaps an ideal connection of theory with practice.For these reasons, modern teaching of analytical chemistry should contain a judicious mixture of reaction chemistry and instrumentation.” Anyone aspiring t o the title of analytical chemist should read reference 1, as it contains much thought-provoking material. In this paper I wish to extend the definition given and argue that all of analytical chemistry-the teaching of, research in, and practice of-can be summarised as a “judicious mixture of reaction chemistry and instrumentation.” Analytical Chemistry-The Present The entire philosophy of analytical chemistry is summarised in Fig. 1, which shows the processes involved in providing etc. The teaching of analytical chemistry involves the processes whereby students become conversant with the various pro- cedures available t o progress from “bulk material” to “deci- sion” and acquire the ability t o evaluate critically the merits of such procedures.- 4 1 ~ 0 , research in analytical chemistry involves the generation of improvements in one, some or all of the stages involved or, indeed, the generation of suitable over-all procedures for analytical problems which have, hitherto, remained unsolved. In order to appreciate fully the role of chemistry in the processes shown in Fig. 1, the role of the instrument in a procedure involving a quantitative measurement is examined in more detail in Fig. 2. It can thus be seen that a variety of chemical processes take place inside the instrument (e.g..an atomic absorption spectrometer with a graphite furnace electrothermal atomiser o r a high-performance liquid chromat- ograph). in addition t o a variety of physical and electronic processes. It is important that the analytical chemist under- stands each of these processes and how an over-all uncertainty in the chemical information is produced from the uncertainties in each of the stages involved both inside and outside the instrument. Internal Processed E I ect rica I sample sample signal t 1 I I I physics I I information Pre-treated sample L 1 I I Fig. 2. Taking the lid off the black box Client +m ANALYTICAL CHEMIST t t t t Constraints] IExpertence) 1 Skills I I Research 1 Fig. 1. The analytical chemist at work information about the chemical composition of a bulk material.together with an indication of the quality of that information (the k term to go with a quantitative result). The analytical chemist uses the resources available t o decide on the most appropriate way of building up an over-all method given the constraints imposed by the client. availability of equipment, Analytical Chemistry-The Future Many areas of scientific endeavour (including medical diag- noses, forensic science and industrial manufacture) are underpinned by analytical chemistry and therefore a continued supply of analytical chemists is needed. However, the major source of such analytical chemists is under threat. For some years now, analytical chemistry within universities has been in decline: a decline that has accelerated since the University Grants Committee’s (UGC) cuts of 1981, which have resulted in the non-replacement of staff who have left or retired from Chemistry Departments which were traditionally strong in analytical chemistry (such as Aberdeen.112 I Minor modifications (chemometrics) instruments I I New information from existing data to existing ANALYTICAL PROCEEDINGS, APRIL 1988, VOL 25 I I Major modifications to existing instruments Birmingham, Chelsea, Exeter, Imperial College and Salford).The untimely death of senior staff at Strathclyde and UMIST and, as yet, their non-replacement has been a further blow. Although the emergence of one or two strong schools of analytical chemistry in polytechnics (notably Sheffield and Plymouth) and the establishment of groups at Hull University and Birkbeck College may have partly off-set the general decline, the national picture is still one of diminished activity compared with the situation in the early 1970s.The effects of the most recent UGC cuts have yet to be fully felt, but the news will not be good for analytical chemistry as the revised levels of funding to universities took account of the results of the UGC’s “selectivity in research” exercise. Of the chemistry depart- ments currently research active in analytical chemistry, only one was judged “above average,“ two were “average” and six were “below average.” On top of all this, an even more serious crisis for analytical chemistry may be imminent. The UGC has selected chemistry2 as the next subject to undergo an “Oxburgh” style review.3“ Before this can get under way, the Advisory Board for Research Councils (ABRC) has proposed that instead of conducting such reviews subject by subject, whole institutions should be placed into one of three categories.4 The top 15 would be fully funded for research, the same number in the next category would be doing some high-level research in less costly areas and the remainder would be teaching at undergrad- uate and master’s level with little real research. At this early stage it is not clear what criteria are to be applied in deciding into which category a Department or University (depending on whether the UGC or ABRC review is implemented) will be placed, but it is almost certain that the UGC’s own selectivity in research ranking list(s) will be involved. The future for analytical chemistry can hardly be said to be bright; and when, in the not too distant future, (a) the supply of graduates with analytical skills has decreased, (b) the numbers of higher graduates with MScs and PhDs in analytical chemistry has dwindled and (c) there are no active research groups to provide consultancy or short- or long-term research facilities for industry and government, don’t say you weren’t warned! New phenomena New chemistry instrumentation with existing I se‘rrs 1 and instriments I Fig.3. A spectrum o f analytical chemistry research Professor R. Oxburgh is the Chairman of a UGC group reviewing Earth Sciences in Universities. The group’s recommendations are to divide the existing 51 departments into three categories.The top group, of 12-15 institutions, would be funded to carry out teaching and research to doctoral level and beyond. The middle group. 1S+. would be able t o teach at master’s level and the remainder would only offer undergraduate courses. This third group would be recommended to stop maintaining separate departments and to run courses through service teaching from other departments. Diffusion-controlled Reactions At an electrode surface When an electroactive material arrives at a plane electrode under conditions of semi-infinite linear diffusion, the current which flows is proportional to the number of moles diffusing per unit time per unit electrode area, which, in turn, is proportional to the concentration gradient at the electrode surface, i.e. , where i is the current flowing at time r , n is the number of electrons involved in the electrode reaction, F is the Faraday constant, a is the electrode area, D, is the diffusion coefficient of the oxidised species (i.e., the species diffusing towards the electrode) and (dC/dx)o. is the concentration gradient at zero distance from the electrode surface at time t. As where Ck is the bulk concentration of the oxidised species, then Taking equation (3) as a starting point, it can be shown5 that if the reduced species is monitored by a light beam passing normally through the electrode, the absorbance, A . is given by where ER is the molar absorptivity of the reduced species (it is assumed that E,, the molar absorptivity of the oxidised species, is zero at the wavelength in question).When equation (4) is compared with the Beer’s law relationship A = E ~ C . . . . . . . . ( 5 ) it can be seen that the behaviour at the electrode surface may be modelled by the movement of a boundary out into the solution to give a fath length b of 2(D,,rln)+ up to which the concentration is C,, and beyond which it is zero. Although equation (4) provides, in principle, the quantitative basis of an analytical technique, a simple calculation shows that for typical values of D,, (10-5 cm’ s- 1) and reasonable time scales the path lengths are too short to be analytically useful. However, if the light beam is rotated through 90” and passed at grazing incidence along the electrode surface, as shown in Fig. 4, the path length term becomes 2(D,,t/m)%/h, i.e., the path length is increased by a factor blh.where b is the length of the electrode surface and h is the thickness of the light beam. c = Cob r------ -b- Fig. 4. Grazing incidence spectroelectrochemical configuration Values of blh of several tens are possible and the sensitivity enhancement gives the configuration of grazing incidence the basis for useful analytical measurements. The full equation is given by A = ( E ~ -q,) [2(D,,rlm)5 blh] C,”, . where E,, is the molar absorptivity of the oxidised species.ANALYTICAL PROCEEDINGS, APRIL 1988, VOL 25 113 Not only is absorbance directly proportional to concentra- tion (measured at a fixed time after the start of the experi- ment), but At-$ is also proportional to concentration, so the slope of the plot of A versus fi can be used as a quantitative analytical parameter.Thus, the working range of the technique may be increased as for more concentrated solutions, if the absorbance value measured goes off-scale, there will still be sufficient data available to calculate a value of At-+. It has been found6 that a linear relationship exists for both quantitative parameters, not only for simple homogeneous redox reactions (such as the reduction of polyphosphomolybdate) but also when a chemical reaction following the electrochemical reac- tion is monitored (such as the reaction of starch with iodine produced by the oxidation of iodide). Although the technique removes some of the kinetic restrictions often associated with conventional spectrophoto- metric methods, allows the working range to be extended and offers interesting possibilities in the design of chemical and electrochemical reactions for analytical purposes, there is no improvement in detection limit compared with the conven- tional technique (the concentration in the light beam cannot be greater than the bulk concentration).Also, the technique is intrinsically slow, as the absorbance - time characteristics are governed by the diffusion of the electroactive species up to the electrode surface (diffusion in liquids is a slow process). The sensitivity of the technique can be improved by introducing a pre-concentration stage whereby a metal is electrodeposited on the electrode surface before stripping into a solution containing a colorimetric reagent.Silver has been determined6 by anodic dissolution into an o-phenanthroline - tetrabromofluorescein reagent after pre-concentration for 15 min. The characteristic concentration (concentration for ab- sorbance 0.0044) obtained was 0.38 p.p.b., which compares favourably with a typical value for flame atomic absorption spectrometry of 50 p.p.b. To speed up the production of an analytical signal based on a diffusion-controlled process, the concentration gradient responsible for the diffusion must be made steeper. This is thc basis of the increased sensitivity of pulsed voltammetric techniques (detection limits are improved as greater discrimi- nation between Faradaic and non-Faradaic processes may be achieved). This approach has yet to be tried for the spectro- electrochemical studies outlined here.Combination with laminar flow If the experiment is changed so that the concentration gradients generated by laminar flow in a narrow circular tube are used as the driving force, diffusion-controlled chemistry on short time scales may be exploited for analytical purposes. This concept is the basis of the single-line flow injection manifold, in which a discrete volume of sample is injected into a continu- ously flowing reagent carrier stream and the reaction product is monitored at a downstream detector. The early stages of the experiment are shown in Fig. 5 . There is difficulty in giving an accurate representation of a flow injection experiment in diagrams of this type because, firstly, if the typical values of tube diameter and volume injected of 0.5 mm and 50 1-11, respectively, are taken, whatever scale is used in the diagram the length of the injected slug should be 500 times the diameter of the tube (i.e., the sample occupies a length of 25 cm).Secondly, at the typical flow-rate of 1 ml min-1 (33 vl s-1) the central stream line (which in laminar flow is moving at twice the average linear velocity) is moving at 170 mm s-1, so the lower diagram in Fig. 5 shows the shape of the boundary approxi- mately 0.009 s after injection. Molecules on the leading boundary diffuse from faster to slower moving stream-lines, as shown by the arrows, whereas molecules on the trailing boundary diffuse from slower to faster moving stream-lines. Hence the entire sample zone is transported down the tube to the detector, during which time the zone is mixing with the carrier stream.Under the conditions normally employed in 0.5 mm i.d. tube; 50 pi injected; 1 ml min-’ 4 500 d * t u = 1 m min-1 7 Y 4 - A 3 Elapsed time = 0.009 s Fig. 5 . Initial stages of dispersion by laminar flow in a straight tube flow injection analysis, no analytical solution of the diffusion - convection equations is possible7 and one approach to predict- ing the shapes of the peaks produced is to model the dispersion process. One very simple model is the single well stirred tank (WST) model, the basis of which is that all dispersion is modelled by the passage of a discrete slug through a single well stirred tank. This model produces exponential-shaped peaks and the concentrations at the peak of sample (injected), C& and reagent, CF, (in the carrier stream) are given by = C: [I - exp(-V,/V)] CF = C: exp(-V,/V) .respectively, where Cs and C R are the steady-state concentra- tions of sample and reagent, respectively, and Vi and Vare the volume injected and tank volume, respectively. The extent of dispersion is normally quantified by the dispersion coefficient, D , at the peak, where D is given by D = C,/C, . . . . . . . . Transforming equations (7) and (8) by the appropriate substitutions for D and DR (sample and reagent dispersion. respectively) produces the relationship D = DR/(DR -1) . . . . . . (10) It can be shown that this relationship holds at all points o n the rise and fall curves and indeed is independent of the dispersion model used.8 If only physical dispersion is considered (see upper diagrams in Fig.6), the peak width, At, at any concentration C’ of the injected sample material is, on the basis of the WST model, given by At = (V/u)ln[(C&/C’)-lJ - (V/u)ln(D - 1) . . (11) where u is the volumetric flow-rate. Equation ( 1 1) indicates that peak-width measurement has potential as the basis of a quantitative method as At is a logarithmic function of the injected concentration. However, the presence of C’ in the logarithmic expression means that to use the relationship, when in practice instrument response, H , as a function of time is measured, the concentration corresponding to the measure- ment level must be found from a separate experiment. If the approximation Cz/C’ >> 1 is made.S equation (1 1) reduces to At = (V/u)ln Ck- (V/u)ln C ’ ( 0 - 1) .. (12) This equation is valid regardless of the relationship between the response of the instrument; all that is necessary is that the response is stable with time. Thus, the working range of the instrument may be extended, as has been shown‘’ for flame atomic absorption spectrometry, by several orders of magni- tude.114 BTB ANALYTICAL PROCEEDINGS, APRIL 1988, VOL 25 - - 52 x 1.5 mm When chemical reaction occurs and the reaction product is monitored, two possible situations arise as shown in Fig. 6. If the dispersion characteristics are such that the reagent is always in excess (left-hand diagrams in Fig. 6), the product profile will match the sample profile and equation (1 1) will describe the peak width.However, if the dispersion characteristics are such that the sample is in excess in the profile centre (right-hand diagrams in Fig. 6) then a doublet product peak is formed, For a 1 : 1 reaction (the situation illustrated in Fig. 6), the points at r I ---) A t +- t- -Ateq- t - t- -A req- t- Physical dispersion and chemical reaction in a single-line flow Fig. 6. injection manifold which the sample and reagent profiles intersect are the equivalence points to the flow injection “titration” on the leading edges and on the trailing edges of the dispersed profiles. If the concentration at the equivalence point is C,,, substitution for the appropriate values of D and DR at these points into equation (10) gives C,, = C#2&/(Cg + Ck) . . .. (13) and substitution of C,, given by equation (13) for C‘ in equation (11) gives At,, = (V/u)ln Cz - (V/u) In CE ( D - 1) . . (14) showing that the time interval between the equivalence points, At,,, is a simple logarithmic function of the injected concentra- tion. The use of equation (14) is illustrated in Fig. 7 , which shows the calibration obtained for the determination of OH- over the range 5 x 10-5 to 5 M. The reagent was an acidified solution of bromothymol blue and good approximations to exponential peaks were obtained from a “gradient tube” 52 mm x 1.5 mm i.d. As the response of the detector only has to be stable with time, it is suggested that an inexpensive detector could be constructed for such measurements based on light-emitting diodes and photodiodes and making use of simple digital logic circuitry to measure time (via a clock chip) between two peaks.As this is peering into the future to some extent, perhaps such an instrument, making quantitative measurements in the time domain, should be known as an analytical chemistry time machine. 110 100 90 2 80 a U 70 60 -6 -5 - 4 - 3 - 2 - 1 0 1 Log pH- ( 5 x 10 - 5 M) Fig. 7. Determination of OH- by a peak-width flow injection method Conclusion In acknowledging the help and guidance of several groups of people, I would like to recall the opening theme of this paper as I was undoubtedly influenced to pursue a career in analytical chemistry by my experiences as an undergraduate at Aberdeen University. Thus Bob Chalmers along with Professor Tom West, my PhD Supervisor, and my present colleagues at Loughborough University, particularly Professor Jim Miller, must take some responsibility for my contributions to ana- lytical chemistry and I thank many other members of the Analytical Division for being such supportive professional colleagues. Several overseas governments have sponsored research workers, for which I am grateful, as I am to both the SERC and the Trustees of the Analytical Division Trust Fund for financial support over several years. 1. 2. 3. 4. 5. 6. 7. 8. 9. References Baiulescu, G. E., Patroescu. G., and Chalmers, R. A,. ”Education and Teaching in Analytical Chemistry.” Ellis Horwood, Chichester, 1982. p. 54. Williams. E . , Times Higher Educ. Suppl.. June 12th. 1987. Turner. J . , Times Higher Educ. Suppl.. June 19th. 1987. Turner, J.. and Williams, E., Times Higher Educ. Suppl.. June 19th, 1987. Tyson. J. F.. and West, T. S . . Talanta. 1980. 27. 335. Tyson.’J. F., Talanta. 1986, 33, 51. Stone, D. C., and Tyson. J . F.. Analvsr, 1987. 112. 515. Tyson. J . F.. Anal Chim. Acta, 1986, 179. 131. Bysouth, S. R.. and Tyson. J. F.. Anal. Proc.. 1986. 23. 412.

ISSN:0144-557X    

DOI:10.1039/AP9882500111

出版商:RSC    

年代:1988

数据来源: RSC

摘要:

ANALYTICAL PROCEEDINGS, APRIL 1988. VOL 25 115 Analysis in Agricultural Sciences The following are summaries of six of the papers presented at a Joint Meeting of the Scottish and East Anglia Regions and the Agricultural Group of the Industrial Division held on June 29th-30th, 1987, in the University of Aberdeen. Chemical Typing of Cereal Grains R. Cook United Distillers plc, Glenochil Research Station, Menstrie, Clackmannanshire FK 1 1 7ES Over the past few years, the number of commercially available cereal varieties listed annually by the National Institute of Agricultural Botany has steadily increased. 1 In these lists varieties are recommended on the basis of field performance in terms of yield. disease susceptibility. etc., but only a few barley varieties are deemed suitable for malting.brewing and distilling. For reasons of quality control in these applications it is important that reliable identification methods are readily available for both barley and malt. Grain Morphology Only a few morphological features can be uged reliably to aid the identification of cereal grains.' For barley, these include "twisting" in grains derived from &row varieties, "bib"-type lodicules, which occur in a minority of grains, "woolly-haired" rachillae and distinguishing hairs. which are sometimes present in the ventral furrow. Other features such as rachilla length and the presence of surface spicules are less reliable indicators because of their dependence on environmental factors. Threshing and handling alter many grain characters and others are destroyed during malting and kilning.Chemical Staining Blue anthocyanin pigments. contained in the aleurone layer of some barley varieties, are revealed by acid de-husking. Pigmentation can be chemically enhanced (to red) by treating the de-husked grains with methanolic hydrochloric acid. Low-powered microscopy is used to define the pigmented areas.3 Samples of malted barley that have been de-husked by hand can also be examined in this manner. Protein Composition On the basis of solubility, cereal proteins can be divided into four groups. Barley albumins and globulins. which together account for 20% of the total grain nitrogen, are soluble in water and dilute salt solutions, respectively. Alcohol-soluble prolam- ins (hordeins) make up 40% of the nitrogen and the remaining protein, which is soluble in dilute acid or alkali.is classified as glutelin. Barley hordeins are further sub-divided into groups A, B. C and D according to their relative molecular mass and amino-acid composition. Groups B, C and D vary in their polypeptide compositional patterns between varieties. This characteristic appears to be genetically stable and thus, by using suitable analytical techniques, constituent polypeptides can be separated to yield reproducible "fingerprint" patterns for each variety. Hordein Separation by Reversed-phase HPLC Extraction and separation procedures were based on those described in the published 1iterature.J Alcohol extracts of the grain were separated on columns of Hypersil WP 300 Octyl and were eluted by means of a computer-generated gradient based on a mixture of water and acetonitrile.Acetophenone was run as an internal standard and polypeptides were detected photometrically at 210 nm. The average running time was 80 min. Group "A" hordeins, which showed a poor resolution, were eluted first. These were followed by group "C" hordeins, the sulphur-rich "B" proteins being the last to leave the column, Apart from the "A" region, characteristic and highly reprodu- cible patterns were obtained for most of the varieties examined and these patterns were still discernible even after 9 days' germination, thus providing adequate information for identifi- cation of the corresponding malts. Extracts prepared from fine grists were used to examine mixtures of varieties. Although changes in the relative proportions of each component variety were readily detectable, quantitative assessment of the mix- tures was complicated by the observation that some peak heights.but not their positions. were influenced by the nitrogen contents of the grains. As corn-by-corn examination is precluded by time requirements. the usefulness of HPLC for resolving varietal mixtures is reduced. The detection of mixtures can, however, still be carried out quite rapidly by comparing the elution profile of the suspected sample, based on a grist extract. with a reference profile derived from a single-corn extract. Because of the relatively short over-all time requirement for the HPLC technique. the high degree of reproducibility and the reduced exposure of the operator to the effects of potentially hazardous chemicals, the technique remains invaluable as a confirmatory test for single varieties.Polyacrylarnide Gel Electrophoresis (PAGE) Early techniques were based upon complex formation between the hordein polypeptides and sodium dodecyl sulphate (SDS). Subsequent separation took place in polyacrylamide gels according to the apparent relative molecular mass of the individual polypeptide fractions. However, the method required highly-purified SDS, took up to 7 days to complete and was not particularly suited to the examination of malted barleys. It was therefore abandoned in favour of simpler procedures in which the separation of uncomplexed poly- peptides took place in acid gels. Typical extraction procedures are described in the litera- ture.5 Useful results were obtained by extracting either single corns or a fine grist with an aqueous solution containing 3 M urea, 1 "/o mercaptoethanol and 20% chloroethanol. The total running time was 3 hours, during which the polypeptides were resolved into bands which progressed in the order A-D from anode to cathode.Bands were fixed by trichloroacetic acid, then stained with Page Blue G-90. Gels can be stored wet in sealed polythene bags or they can be scanned by means of a116 ANALYTICAL PROCEEDINGS, APRIL 1988, VOL 25 high resolution densitometer to produce a profile consisting of a series of peaks. The nitrogen content of the grain influences peak height but not position, so that varietal band patterns remain reproducible. Because 20-40 individual corns can be run simultaneously it is possible to investigate the composition of varietal mixtures.The choice of extractant is important and, particularly for malted barley, reference samples must be run for comparison. Isoelectric focusing techniques can be used if simple separa- tion procedures are inadequate for complete identification. Likewise, gradient gel methods or 2-dimensional procedures can often supply additional taxonomic information. Alterna- tively, gels containing separated protein bands, usually extrac- ted with dilute salt solution, can be incubated with a suitable substrate and reaction products from a selected enzymic activity demonstrated by the use of histochemical stains. Most enzymes in barley are represented by sets of isoenzymes and often isoenzyme patterns are very different for pairs of varieties that are not easily resolved by PAGE of their proteins. Conclusions Most barley varieties can be identified by combining results from chromatographic techniques with those derived from chemical staining and examination of grain morphology.Reversed phase HPLC techniques are rapid and precise, but sample throughput is limited. With PAGE, many samples can be analysed simultaneously, making the technique more suitable for the resolution of mixtures. Both chromatographic methods can be applied to malted barley and similar techniques are applicable, not only to other cereals, but also to a wide range of seeds of agricultural importance. References 1 . National Institute o f Agricultural Botany."Recommended Varieties of Cereals 1987," Farmers Leaflet No. 8. National Institute of Agricultural Botany, Cambridge. National Institute of Agricultural Botany. "Detailed Descrip- tions of Varieties of Wheat. Barley. Oats. Rye and Triticale." 1987. National Institute of Agricultural Botany. Cambridge. Day, K. L.. J . Narl Znsr. Agric. Bor.. 1979, 15, 51. Marchylo. B. A.. and Kruger. J . E.. J. Am. Soc. Brchv'ng Chem.. 1985. 43. 29. Marchylo. B. A . . and Laberge. D. E.. Gun. J. Plcinr Sci.. 1980. 60. 1343. 2. 3 . 1. 5 . Sampling of Soils and Plants for Trace Element Analysis M. L. Berrow Department of Soectrochemistrv, Macaulav Land Use Research Institute, Craigiebuckler, Aberdeen A69 2QJ Soil is a complex mixture of primary and secondary minerals, living organisms, their decomposed residues and anthro- pogenic inputs. It is 3-dimensional and varies both vertically.as in the soil profile, and laterally across an area. Soil variability is a factor which has to be borne in mind in designing any sampling procedure, and sound sampling is central to the accurate estimation of soil properties in areas of land of any size. Sampling must be efficient in that it involves no more replication than is necessary to obtain the desired precision. Increasing concern over the contamination of soils and the requirement to assess soil quality is likely to demand an increasingly quantitative approach. The recent CEC Directive, containing agreed maximum permissible limits of potentially toxic elements in soils, will be related to sampling depths and procedures and these will therefore come under critical examination.The sampling of plants poses its own problems because the plant is a living, dynamic organism. The chemical composition of its constituent parts can change considerably over a matter of days as it develops, particularly during phases of rapid growth and dry matter production. A factor that has to be borne in mind in the sampling of both soils and plants for trace elements is contamination, which can arise at any stage during the sampling or preparation of the sample for analysis. It is impossible in a short paper to cover all aspects of the sampling of soils and plants, but further details can be obtained from other publications. 1-2 This paper attempts to describe some well tested procedures and to present some results obtained using them.Soil Sampling Procedure Soils are sampled for many purposes, which include assess- ments of nutrient status, inorganic and organic pollutants. pathogenic organisms, soil-borne pests and diseases, physical parameters and soil mapping. Samples should be placed in thick gauge polyethylene bags and clearly labelled. The use of metal staples. particularly copper-plated staples, to close bags is best avoided. Paper bags should not be used if boron or zinc are to be determined because they can contain substantial amounts of these elements. Bags with a bitumen-impregnated layer should also be avoided as the ash from these can contain over 1 % of vanadium.4 Cloth bags are unsatisfactory because they are neither waterproof nor dustproof.Soil samples should always be packed in a container separate from plant samples. Soil Profile Sampling For soil survey purposes, including mapping, a soil profile or vertical column of soil is exposed to a depth of about 1 m and samples from the various soil horizons identified are taken. The profile is essentially 3-dimensional and has to be large enough in volume and lateral dimensions to evaluate soil properties at a particular place. Representative samples from each specific horizon can be taken, avoiding diffuse boundaries between horizons. This procedure is often used for the characterisation and mapping of soils. Where an assessment of the depth of penetration of surface applied nutrients or contamination is required, the complete profile is best sampled in successive layers of, say, 10 cm in thickness.Care is needed at all times to avoid the use of sampling equipment. which may introduce trace element contamination such as stainless steel. Topsoil Sampling In order to obtain a representative sample of a topsoil a number of soil cores (sub-samples) are mixed to provide a composite sample for analysis. Such a composite can be described as representative if there is a high probability that a similar sample obtained in the same way would give similar results. It has been established that at least 25 cores or sub-samples taken randomly will constitute a representative sample from an area of land that has received unifortn treatment. Large fields or extensive unenclosed areas, even if apparently uniform.should be divided into smaller samplingANALYTICAL PROCEEDINGS, APRIL 1988, VOL 25 117 units each preferably of not more than 4 ha in size. Sampling along two diagonals of a field or unit introduces a serious bias to the centre of an area, whilst sampling in a W-pattern also has disadvantages. Sub-samples should be taken by traversing the area in a zigzag manner along different sections of the area until the whole area is covered on a random basis as recommended by Scott et ul.5 Sub-samples should be taken with a soil auger or corer to a uniform depth and cross section. The sampling of arable land should normally be to plough depth (15 or 20 cm). but on established pasture, a shallower depth (7.5 or 10 cm) is usually adequate to include the main root system of the herbage. The objective is to provide at least 500 g of air-dried.<2 mm sieved soil for analysis. Sampling immediately following lime or fertiliser applications is undesir- able, and any obvious patches resulting from lime dumps, fires, storage pits, etc., should be avoided. as should access areas and end-rigs. Samples for copper or zinc analyses should avoid material in the vicinity of electric power cables or galvanised fences, respectively. Where a problem arises only on a specific area of a field, samples from affected and non-affected areas should be taken. This applies to crop samples as well because directly comparable analytical findings can facilitate diagnostic interpretation. Sampling of Field Plots The results for 0.05 M EDTA-extractable copper in field plot soils sampled three times in three successive years are shown in Table 1.The soils were freely drained (FD). imperfectly drained (ID). poorly drained (PD) and very poorly drained (VPD) soils of the Tarves association. The r.s.d. values of 7.7. 8 and 13°/0 obtained are comparable with those of 11, 6. 4 and 12% obtained at another comparable site on a different soil parent material. The r.s.d. values for 0.43 M acetic acid extractable cobalt averaged 17 and 11 YO, respectively, for the same two sites. The over-all mean r.s.d. values obtained for 28 different field plots on a variety of Toil types were 12% for copper and 15"l0 for cobalt." These results are similar to those obtained by Aichberger er nl. ,7 who found r.s.d. values of 17, 17, 15 and 15 for copper.zinc. nickel and lead, respectively, in sampling studies on a 1 ha field plot. Table 1. EDTA (0.05 M) extractable copper in soils sampled 3 times in 3 successive years as mg kg I in dry soil. Tarves Association soils 1069 1970 1971 r.s.d.. June Aug. Oct. June Aug. Oct. June Aug. Oct. (YO FD 4.6 4.1 3.9 4.3 4.8 4.3 4.5 4.0 4.7 7 ID 4.5 4.4 4.1 5.2 4.8 4.5 1.5 4.5 4.7 7 PD 6.X 5.6 6.4 7.0 7.0 7.5 7.5 6.8 6.8 X VPD 7.0 6.5 7.7 8.0 7.5 9.2 9.9 X.9 8.5 13 At another field plot experiment site screw auger samples were taken most years between 1956 and 1982. often at different times of the year. Two plots which had been treated with 6 kg ha-' of copper and sampled 45 times in all produced the following results: range. 1.06-2.19; mean. 1.60 ug g-1; standard deviation.0.289 pg g-1; r.s.d.. 18%. For a plot receiving 30 kg ha-1 of copper and sampled 22 times in all the results were: range. 3.5G8.12; mean, 6.06 pg g-1; standard deviation. 1.329 ug g--I: r.s.d. 22%. At two other field sites where copper had been applied in 1964 and 1972. both screw auger and soil profile pit samples were taken in 1984 and 1981. respectively. to a depth of &20 cm on each plot. Six control and six copper treated plots were sampled at each site and the agreement between the auger and profile sample pairs was good. The 12 analyses obtained for all four treatments produced r.s.d. values ranging between 13 and 15%. In another investigation, extractable copper and cobalt were determined in soils from 16 different sites at locations ranging from Aberdeenshire to Berwickshire.Fenced field plots were sampled using a screw auger and at least 25 sub-samples composited to obtain a "plot" soil sample. The field around the plot was also sampled in the same manner to obtain a "field" sample. If the sampling technique is valid the two samples from each site should produce the same result. The regression equation obtained by plotting y = log (plot) value against x = log (field) value was v = -0.025 + 1 . 0 7 ~ . showing good agreement between the two sets of results. For cobalt the agreement was not quite as good and the corresponding regression equation obtained was y = 0.014 + 0.997~. Plant Sampling Procedure In sampling plant materials it is necessary to ensure that the composite sample is representative of the population of plants being studied and that it is free from soil contamination.The principal form of contamination commonly derives from the soil and from the sampling operation. Contamination of leaves by soil, foliar sprays and atmospheric deposition can be a major problem. Washing of leaves with a 0.1 % solution of a non-ionic detergent is claimed to remove soil contamination but not metals added as a fungicide or foliar spray..' Washing of herbage is better avoided, if possible. but large. smooth leaves can be wiped clean with a damp cloth. The extent of any soil contamination depends upon the format of the plant and on the weather conditions prevailing immediately before sampling. Pasture herbage being grazed should not be sampled because of trampling and at least 2 weeks should elapse between the cessation of grazing and sampling.Table 2 shows soil : plant ratios based on average total soil contents taken from a recent reviews and of plants on a dry matter basis.4 For the major elements, and boron, copper. molybdenum and zinc, with ratios up to about 5 soil contamina- tion can be disregarded. For elements with ratios above 100, however, serious problems arise. The ratios can also provide some measure of the extent of soil contamination. Titanium is the best element for this purpose, with a ratio of 5000 to 1 . Titanium contents of clean herbage sampled during the growing season normally fall in the range 0.5-2.0 pg g-I and herbage containing more than 10 pg g-I can be considered contaminated. Titanium is about 10 times more sensitive than aluminium and 20 times more sensitive than iron as an indicator of soil contamination.Cobalt. chromium and vanad- ium results can be seriously affected by soil contamination. For example, an apparent uptake of chromium by beet and lettuce leaves grown on a sewage sludge treated soil has been shown by the close linear relationship between chromium and titanium values to be a result of soil contamination and not of plant uptake.9 Table 2. Soil : plant ratios based on average total contents o f soil and plant dry matter P 0.2 S 0.4 K 1 Ca 1 B 2 Mo 2 Zn 2 cu 3 Hg 5 Mg 4 Mn 8 co 120 Na 1 0 Fe 300 Sr 10 Al 700 Cd 1 0 Cr 1000 As 20 V 1000 Se 20 Si 1000 Pb 25 Ni 35 Ti SO00 The growing plant is a living organism. and its chemical composition changes throughout its life.Factors to be borne in mind in sampling plants are firstly, differences in the composi- tion of different species, even when growing on the same soil, secondly, differences in plant part composition, and thirdly,118 ANALYTICAL PROCEEDINGS, APRIL 1988, VOL 25 effects of the stage of growth on plant cornposition.'".'' All of these factors have to be taken into consideration when sampling plants for trace element analysis. Trace element contamination during the sampling of plants can be almost entirely eliminated if the following precautions are taken. Plants should be held in one hand and cut with clean, steel, sheep shears at least 3 cm above soil level. The plants must not be plucked or pulled during sampling.A composite of at least 25 sub-samples should be taken. and the mass of fresh material should be not less than 1 kg. Leaves that are obviously dust contaminated. damaged by insects. mechanically or chemically injured, diseased or in temperature or moisture stress condi- tion are better avoided. Each sub-sample should be placed immediately into an adequately large polyethylene bag. Herbage samples should be firmly compressed to expel as much air as possible and tightly packed in double polyethylene bags with labels placed between the two bags. The neck of the outer bag should be securely tied and a tie-on label attached. The bags should be kept cool and transported to the laboratory with as little delay as possible and transferred to a cold room on arrival.With root crops it is impossible to avoid soil contamina- tion of the original samples. These should be transferred to the laboratory in polyethylene bags. then thoroughly washed before sub-sampling by coring or sectioning in soil-free condition. As the leaves. stems and flowering heads of plants contain different amounts of trace elements it is essential to compare like with like when examining samples of plants taken from different parts of the same field. The samples analysed should consist of comparable parts of the plant depending upon the objective, e.g., leaves, grain, fruit, roots or whole plants excluding the root system. Samples for comparison should, in addition, be at the same stage of growth, e . g . , flowering. as it is seldom justifiable to compare directly samples taken at different times during the growing season.1. 1 L. 3. 4. 3. 6. 7. 8. 9. 10. 1 1 . References Ministry o l Agiculture. Fisheries and Food. Technical Book- let No. 2082. "Sampling of Soils. Soil-less Growing Media. Crop Plants and Miscellaneous Substances for Chemical Analysis." HM Stationery Office 1979. pp. 1-44. Calder. A. B . . and Voss. R. C . . "The Sampling of Hill Soils and Herbage with Particular Reference t o the Determination of Trace Elements." Bulletin 1. Macaulay Institute for Soil Research. Aberdeen. 1957. pp. 1-52. Gomez. A.. Leschber. R.. and L'Hermite. P.. Edirors. "Sampling Problems for the Chemical Analysis of Sludge. Soils and Plants." Elsevier. Amsterdam. London and New York. 1986. Mitchell. R. L.. J . Sci. Food Agric..1960. 10. 553. Scott. R. 0.. Mitchell. R. L.. Purves. D., and Voss. R. C.. "Spectrochrmical Methods lor the Analysis of Soils. Plants and 0 the r Agr i c u I t u r a I Mate r i a 1 s . " B u I I e t i n 2 . Ma ca u I a y I n s t i t u t e for Soil Research, Aberdeen. 1971. p. 8. Berrow. M. L.. Burridge. J . C.. and Reith. J . W. S.. J . Sci. Food Agric., 1983. 34. 53. Aichberger. K . . Eibelhuber. A . . and Hofer. G.. iri Gomez. A.. Leschber. R.. and L'Hermite. P.. "Sampling Problems for the Chemical Analysis o f Sludge. Soils and Plants." Elsevier. Amsterdam. London and New York. 1986. pp. 38-43. Ure. A. M.. and Berrow. M. L.. in Bowen. H. J . M.. Ediror. "Environmental Chemistry." Volume 2. Royal Society of Chemistry. London. 1982, pp. 94-203. Berrow. M.L.. and Burridge. J . C.. in "Inorganic Pollution and Agriculture." MAFF Reference Book 326. HM Stationery Office. London. 1980. pp. 159-183. Burridge. J . C.. Reith. J . W. S., and Berrow. M. L.. in Suttle. N. F.. Gunn. R. G . . Allen. W. M.. Linklater. K. A.. and Wiener. G . . Editors. "Trace Elements in Animal Production an d Vet e r i n a r y Practice . '- Occasion a I Pub I i ca t ion N o . 7. British Society of Animal Production, 1983. pp. 77-85. Mitchell. R. L.. and Burridge. J . C.. Phil. Traris. R. Soc. h t l d O t 7 . Ser. A, 1979. 288. 15. Action from Analysis : Challenges for Agricultural Analysis D. Atkinson The Macaulay Land Use Research Institute, Craigiebuckler, Aberdeen AB9 2QJ Soil analysis is used as a means of identifying the probable needs of a crop for fertilizers.This practice depends upon a number of basic tenets of which the more important are: firstly, that the amount of soil nutrient available to plants can be measured; secondly. that soil nutrient status influences plant nutrient content: and thirdly. that "fertilizer need'' can be related to soil test values. These ideas are conceptualised in Fig. 1. Routine soil analysis in Scotland currently includes tests for pH (and lime requirement) and available phosphorus, potas- sium and magnesium. Nitrogen is estimated from crop and fertilizer history. Estimates of the needs for fertilizer phospho- rus were established from trials linking the response of swedes to phosphorus additions with available phosphorus test val- ues.' A similar logic was followed for potassium and magne- sium.Thus. general. widely applicable, relationships are used to estimate the fertilizer needs for economic cropping on particular soils. Test values can be adjusted systematically for other factors. e.g., for a Tarves series soil 20 mg dm-3 of phosphorus is the upper boundary of a low status soil, while for a Countesswells soil the limit is 35 mg of phosphorus.' Other adjustments are made on a more subjective basis. This system has worked well over a number of decades. In an era of agricultural surpluses, and in the light of current technological developments and of changed aims for, and restraints upon. farming, it may now need to be amended. Current Problems Difficulties occurring with the current analytical system are discussed in this section.The first is inaccuracies in characteris- ing the available soil nutrient supply. especially for nitrogen, for which there is no reliable soil test. Nitrogen is the nutrient added to soils in the largest amounts. If it is not used efficiently by a crop it has the potential to become a pollutant. lp Scotland, between 1983 and 1985,' average nitrogen rate, for all crops and grass, increased from 124 to 130 kg ha-1. As imbalances between the nitrogen removed by the crop and that added as a fertilizer can be substantial (for winter wheat 215 kg are added while 150 kg ha-1 are removedj) and as the residual value of unused nitrogen is ni1,5 then any excess of fertilizer added must be at risk (Fig. 1). Correlations between soil tests and crop responses may be poor.For phosphorus, Reith er al. 1 found correlations with the standard acetic acid method to vary from 0.35 non-significant for a Foudland soil, to 0.67 (P 3 0.01) for a Countesswells soil. For the acetic method their model correctly assigned soils into a fertilizer phosphorus category on 63% of occasions. Recent studies using a mixed resin improved the above correlations to 0.46 (P 2 0.05) and 0.70 (P 2 0.001). respectively. In addition, resin copes better with previous fertilizer residues, e.g.. rock phosphate. the availability of which is over-estimated by the use of acetic acid." Some of these variable responses ofANALYTICAL PROCEEDINGS. APRIL 1988, VOL 25 119 different soils may relate to effects on root system develop- ment. The recommendations made to farmers are often based on generalised (not local) predictive models and on samples derived from extensive areas.It has been suggested that samples should not represent areas >4 ha.' Even this size of sample can cause problems. In a survey of the soils of IHR, East Malling, White'; found the p1-I to vary across the whole farm from 4.0 to 8.5. with some individual fields. as small as 0.45 ha. varying from 4.5 to 7.9. Here. liming to an average of 6.2 would still leave 43% of the field at a pH of 5.7 and so with a high risk of crop damage. Additionally. samples for analysis are normally taken from bulk soil although. for micronutrients at least, Linehan et ctl.') showed bulk soil to differ from that around the root. Atmosphere Saleable Waste t \ I Plant biomass I I SU 'face +:e rt i IT r Product If I water Fig.1. A conceptual diagram of the soil-plant nutrient supplv svstern. Boxed areas represent those influencing current advisory soil analytical considerations General recommendations make little allowance for local effects (except, perhaps, for soil type a s detailed above and informal modifications made by local advisors). Even where general correlations between soil factors (such as pII) and plant nutrient levels (such as leaf manganese) are poor. they have a predictive value on individual sites. In addition. soil factors may interact. so changing nutrient requirements. At pH 5.4 a 60 kg ha-1 phosphorus application gave a barley yield of approximately 6 x 10' kg ha-', while at pH 5.8 a similar yield was achieved with 20 kg ha-' of phosphorus.1" Similarly.a sulphur application to oilseed rape of 40 kg ha - 1 resulted in the uptake of 31 kg ha-1 of sulphur with a 100 kg nitrogen treatment. With 200 kg of nitrogen. 49 kg was absorbed." This type of phenomenon could and should be forrnalised in recommendations. Soil or Tissue Analysis: a Choice Soil analysis characterises a soil's potential to supply nutrients. Where other factors. e.g.. impedance to root system develop- ment. restrict nutrient uptake. or where other sources of nutrients are significant, e.g.. stored nutrients in perennial crops. then correlations between soil test and plant perfor- mance will be poor and plant tissue tests may be more sensitive. Additionally. nutrients sampled by an extractant solution may be chemically unavailable to the plant (see above on resins) or made unavailable by factors such as drought.The availability of selenium, which is not currently assessed by soil analysis. depends upon the extent of leaching of the selenate ion and its "mineralisation" from organic forms and from the relatively unavailable selenite ion, and may be better characterised by its concentration in plant tissue. For tree fruits. where much of the nutrient in current growth or fruit is derived from reserves, foliar analysis. which often correlates better with the previous than the current year's soil data, may give a better guide to nutrient requirements. Implications of Current Technological Advances Changes of agricultural importance include the following. Firstly, improved methods of vegetation assessment.By using radiometric methods, vegetation ground cover and biomass can he assessed." Nutrient demand is driven by growth so the greater the growth the higher the demand for soil nutrients. Ingestad" developed from this relationship his concept of the relative addition rate (under field conditions ..the nutrient flux density"). Nutrients are supplied at a rate corresponding to the plant's consumptive potential (allowing for natural nutrient supply in the field). A constant internal nutrient concentration (0.6'% fresh weight in birch) is maintained regardless of the actual growth rate, which will be affected by a range of external factors. Increasing amounts of nutrient are needed by the increasing growth. A constant rate of nutrient supply or the decreasing rate of nutrient supply which results from a single annual fertilizer addition will both result in reduced internal nutrient concentrations and. as a result, growth decreasing.In the field the practical application of this concept requires the ability to monitor vegetation growth (radiometry) and soil nutrient content (in sitii sensors). Technical advances seem to mean that shortly this monitoring should be feasible. In this system there is no over-supply. hence almost no risk of pollution. Secondly, improved analytical methods. The potential value of resin extractions which remove only those nutrients from the soil solution and the labile nutrient in equilibrium with it has been discussed above for phosphorus. A mixed anion - cation resin may shortly be in use to determine the range of elements covered by current analysis.Thirdly. improved information handling systems. Powerful microcomputers are now available on many farms. Availability will increase over the next decade and so allow local (e.g., soil distributions within a field) and marginal factors (e.g.. the influence of pH - phosphorus interactions) to be formally used in the interpretation of soil analytical data. New Challenges for the Future Adjustments to respond to changes in farming practice could include the following. Firstly. improved conservation of nutrients. Ceiling rates of nitrogen tend to be related to the absence of an economic yield response, although with the higher rates both the recovery and the unit response tend to be less than with lower rates (Fig.1). In an era of agricultural surpluses and increased environmental awareness excessive nutrient losses are unacceptable. Both production systems and advisory recommendations must now consider the prevention of nitrogen losses as essential rather than desirable. The Ingestad relative addition rate system would aid this conserva- tion. Links between nutrient additions and losses are not simple and so an improved understanding of the fate of added nitrogen is desirable. For example. studies at this Institute" showed that following the harvest of a spring barley crop 20 kg ha-' of nitrate remained available in the soil but that little of this derived from the current year's fertilizer. More biologically stable production systems may need rather different interpret- ations of soil tests.Secondly. improved product quality. The quality of the final product is now as significant an agricultural target as maximum production and quality can respond differently to inputs. Increasing the nitrogen supply to potatoes beyond the point for the maximum biomass production continued to increase their protein content. 13 Similarly. applications of sulphur to grass can change its acceptability to stock even in situations where effects on growth are limited. The quality targets must be incorporated into recommendations. Thirdly, the use of multi-crop systems. Advisory systems are normally geared to the production of a single commodity. Where two economic crops are being produced simul- taneously, e.g., in a silvopastoral system.where quality timber120 ANALYTICAL PROCEEDINGS. APRIL 1988, VOL 25 and grass for sheep are grown together, recommendations and the related analysis must reflect differences in nutrient needs and perhaps the different periodicities of need of the com- ponent species. Conclusions Current advisory methods are geared to the maximum produc- tion of single crops with only secondary concern for the efficiency of fertilizer use. An era of agricultural surpluses, increased awareness of adverse environmental impact and more complex cropping systems requires analytical methods giving a better targetted nutrient supply. Integration of a wider range of variables. and especially the incorporation of more on-farm information into current systems, should help to achieve this supply.References 1. Reith. J . W. S . . Inkson. R. H . E.. Scott. N. M.. Caldwell. K. S . , Ross. J . A. M.. and Simpson. W. E.. Ferflizer Resecrrch. 1987, 11. 123. "Advisory Soil Analysis and Interpretation." Bulletin 1 of the Macaulay Institute for Soil Research and the Scottish Agricul- tural Colleges Liaison Group. Macaulay Land Use Research Institute. Aberdeen. 1985. 2. 3. 3 . 5 . 6 . 7. 8. 9. 10. 1 1 . 12. 13. "Survey of Fertilizer Practice. Scotland. 1985." Rothamsted Experimental Station. Harpenden. 1986. "Fertilizer Recommendations." Publication 160. The Scottish Agricultural Colleges. Perth. 1985. Scottish Standing Committee. -'Residual Values of Fertilizers and Feeding Stuffs." 38th Report. HM Stationery Office.London, 1986. Atkinson. D., "Soil Fertility." i r i "Annual Report o f Macaulay Institute for Soil Research for 1986." Macaulay 1,and Use Research Institute. Aberdeen. 1987. "Sampling Soil for Analysis." MAFF Leaflet h55. HM Sta- tionery Office, London. 1980. White. G. C.. "A survey o f the pH of the soils of East Malling Research Station. in "Report o f East Malling Research Station for 1960." East Malling Research Station. East Malling. 1961. Linchan. D. J . . Sinclair. A. H.. and Mitchell. M. C.. Plurr(Soi1. 1985. 86. 147. Sinclair. A . H.. and Edwards. A. C.. "Soil Acidity and its Interaction with Phosphorus and Micronutrients .'* Technical Note 7. Macaulay Institute for Soil Research. Aberdeen 19x6. Birnie. R. V.. Millard. P.. Adams. M. J . . and Wright. G .G.. Res. Dev. Agric.. 1087. 4. 3.7. Ingestad, T.. Plant. Cell Eni?roti.. 1982. 5 . 443. Millard. P.. and Mackie. L. A , . "The Influence of Nitrogen Supply on Potato Yield and Quality." Technical Note 8. Macaulay Institute for Soil Research. Aberdeen. 1986. pp. 77-80. Design of a Scottish Soil Fertility Information System A. H. Sinclair Crop Husbandry Division, School of Agriculture, 581 King Street, Aberdeen A59 1 UD G. A. Reaves Macaulay Land Use Research Institute, Craigiebuckler, Aberdeen A59 2QJ K. W. M. Brown Robert Gordon's Institute of Technology, St. Andrews Street, Aberdeen Greater benefit could be derived from soil analysis if more site information were to be collected at the time of sampling soils. Computer storage of data on farming systems, crop rotation. fertiliser usage, grid reference, soil type and climate, along with soil analytical data.would allow the factors influencing the fertility of such an important national resource as the soil to be identified. A better record of temporal trends in soil nutrient status and fertiliser inputs would be obtained for different agricultural practices. Introduction The Scottish Soil Fertility Information System (SSFIS) is based on data from soils that are analysed for advisory purposes by the East of Scotland College of Agriculture, Edinburgh (all analysis after April lst, 1987), The West of Scotland Agri- cultural College. Ayr, and the Macaulay Land Use Research Institute (MLURI) for the North of Scotland College of Agriculture, Aberdeen. Samples are analysed as a result of requests from individual farmers and not as a result of systematic sampling. Farm and field data are collected at the time of sampling and are stored in a computer along with the analytical results.The essential features of SSFIS are described below and examples of the types of information which can be generated are shown (Tables 2,3 and 4). The data used in these examples were obtained for soil samples collected from the North of Scotland College of Agriculture area during 1985. 1986 and early 1987. Data Storage and Processing Data for individual college areas are stored locally on the corresponding college computer or. for the North College. on the MLURI computer. Data from the East and West Colleges are transferred to the Data General Eclipse C/ISO computer at the MLURI.where Data General's INFOS I1 database management system is used. Although a queryireport writer program is supplied by Data General and has been used to extract data for some purposes. the program employed to generate the information presented here was writfen "in- house. '' At the time of soil sampling. data on such features as the farm type, soil series,' cropping. and lime and fertiliser use are coded' together with the eight character grid reference for each soil sample. The complete list of site variables is given in Table I . The number of chemical determinations is dependent on the customer's request. However. soil pH. lime requirement, extractable phosphorus. potassium and magnesium are re- corded for each sample. Soil extractants' were chosen-on the basis of crop trial work under Scottish conditions and- may differ from those of other laboratories.Each sample is represented in the database by a single record. A record consists of 41 fields. each field corresponding to one variable. but in none of the records are data for 211 41 variables stored. Currently something in excess of 32 000 records are stored. The complete set of variables consisting ofANALYTICAL PROCEEDINGS, APRIL 1988, VOL 25 I21 23 field data and 18 analytical results are described more fully elsewhere .? Table 1. Site variables Farm variables Farm name Farm type College Area Office and District Average long-term rainfall Advisory routine. crop problem. etc. College or non-College sampled Individual field variables Area sampled Depth of sampling Grid reference Soil series Texture Last crop Next o r current crop Soil N N .P. K fertiliser for last crop Additional P fertiliser for last Year. type and rate of last lime Type and frequency of organic crop applied manure additions between 1940 and 1978 from north Scotland has been reported4 but no comparison has previously been made for different farming systems (Table 3). Table 3. Soil phosphorus versus farm type. Percentages of samples within farm types Years in five that fields are in grass Extractable P/ P mgl-1 status < 1 1-2 2-5’ 2-St. Hill <9 VL 4 4 7 12 32 >9 s 2 5 L 1 1 16 26 32 38 >25 a 7 5 M 59 58 53 47 23 > 75 H 26 21 14 9 6 No. of samples 1426 6335 3576 3473 298 * Nitrogen fertiliser rate greater than 125 kg ha 1 .+ Nitrogen fertiliser rate less than 125 kg ha- I . Results and Discussion Soil pH One of the most important requirements of good soil manage- ment is the maintenance of the correct balance between soil acidity and alkalinity. The frequency distribution of soil pH differs from one farm type to another (Table 2). Few samples analysed from continuous cropping farms had low or very low phosphorus status, whereas a considerable proportion had high phosphorus status. There appears to be scope for reducing the amount of phosphate fertiliser applied to a number of arable fields. In contrast, samples from grass farms showed an over-all poorer phosphorus fertility, with a further decline on farms of lower nitrogen inputs and hill farms. Table 2. Soil pH Venus farm type.Percentages of simples within farm types Years in five that fields are in grass pH range < 1 1-2 2-5’ 2-St Hill s 5 . s 7 1 1 14 36 66 >5.5 s 6 . 0 49 53 59 33 23 >6.0 S6.5 41 33 25 16 7 X . 5 3 1 I 4 3 No.otwmples 1334 6377 3591 3493 302 7 Nitrogen fertiliser rate greater than 125 kg ha I + Nitrogen fertiliser rate less than 125 kg ha- I. The pH values of 40% of the samples analysed from continuous cropping farms were in the range 6.1-6.5, which is suitable for most arable crops. However, the risk of certain micronutrient problems in crops and stock and of common scab in potatoes is increased at the higher end of this pH range, and is even greater in the 3% of samples with pH values greater than 6.5. At the other extreme. 7% of fields had pH values less than 5.6 and in this range there is a possibility of failure of many arable crops.On farms where grass is the main crop at least 14% of the samples analysed had pH values less than 5.6. and the persistence and vigour of clover and rye grass would have been reduced, especially in mineral soils. The soil acidity levels of these grass farms appeared to be related to nitrogen use. Farms with higher nitrogen inputs (greater than 125 kg ha-’) were found to have higher percentages of samples with satisfactory pH levels compared with those on which less nitrogen had been used. Thus, it appears that the acidifying effect of nitrogen fertiliser is being more than compensated by the use of lime in fields given higher nitrogen dressings. compared with lower nitrogen input farms where liming appears to be neglected to a greater extent.Extractable Phosphorus Most of the soils in north Scotland are naturally deficient in phosphorus and have high phosphorus retention capacities. The phosphorus status of soils (very low. VL: low. L; moderate. M: or high. H) collected as part of advisorv testing Extractable Copper Copper analysis was requested as part of a regular soil testing programme more frequently than any other micronutrient. Soil analysis for copper is often useful to confirm visual diagnosis of copper deficiency in cereals, and it also enables preventative treatment to be carried out where the soil copper status is very low (VL) or low (L). Table 4. Soil copper status Percentage of Extractable copper/ Copper sample\ within mgkg I status each status a 0 .9 VL 10 >0.9 a 1 . 6 L 21 >1.6 s8.5 M 64 >8.S H 6 No. of samples 1391 Conclusions The availability of farm and field data, along with soil analytical results, has allowed comparisons to be made of the effect of farming systems, fertiliser use and soil type on soil fertility. These comparisons have only recently become pos- sible on such a large number of fields with the design and implementation of the Scottish Soil Fertility Information System. We thank farmers. Scottish Agriculture College advisers, soil samplers and analysts, without whose contribution the Scottish Soil Fertility Information System would not exist. Particular gratitude is expressed to Dr. S. Cooke for computer work at an early stage. References 1 .MLURI. “Soil Survey of Scotland Publications List“ (issued annually). The Macaulay Land Use Research Institutc. Aberdeen. MLURIlSAC Scottish Soil Fertility Information System No. 1 . North of Scotland College o f Agriculture. Aberdeen. 1987. in the press. 2 .122 ANALYTICAL PROCEEDINGS, APRIL 1988, VOL 25 3 . MLURUSAC. “Advisory Soil Analysis and Interpretation.” The Macaulay Land Use Research Institute and Scottish Agricultural Colleges. Bulletin 1. 1985. pp. 1-13. Reith. J . W, S . . ”Trends in the Nutrient Status on Scottish Soils. Proceedings of the Third Study Conference o f the Scottish Agricultural Colleges. 1980.” The West of Scotland Agricultural College. Ayr. 1980. 4. The Use of Rapid Tests for Measurement of Plant Nutrient Status 1. G. Burns Institute of Horticultural Research, Wellesbourne, Warwick CV35 9EF The maintenance of an adequate nutrient supply to roots is essential for maximum growth and quality of crops.The supply of nutrients can become inadequate at any stage of growth, but young plants are particularly vulnerable because their poorly developed root systems limit the amount of soil that can be exploited and make them more susceptible to changes in nutrient availability due to drought or leaching. As plants grow larger they also require increasing amounts of nutrients and, in extreme circumstances, may even deplete the soil completely. This problem can be particularly acute with crops raised or grown under glass, as they invariably rely on regular feeding to maintain growth. Although conventional analyses of plant dry matter can be used to keep a check on nutrient status, there is often a considerable delay before the results are available and, recently, interest has been renewed in the use of rapid tests as an alternative. These have the important advantage of provid- ing an immediate result which might be used as a basis for rectifying certain deficiencies as they occur.Rapid tests have been used for many years as a simple “rough and ready” means of monitoring crop nutrient concen- trations for diagnosing deficiencies. Most of the early methods involved do-it-yourself spot tests on filter-paper using sap squeezed from a selected leaf petiole, but these tests were often unreliable and of little value other than for characterising nutrient status into one of three different categories.More recently, a range of test papers and test kits has become available which gives much more reproducible results and considerable attention has been paid to devising and evaluating procedures for the use of these kits with plant sap. With the increased accuracy and precision of these new methods, it has also become important to have more reliable information about the concentrations of nutrients required in plants for maximum growth, in order to interpret the results. New improved techniques for studying the relationships between plant nutrient concentration and growth have recently been devised and these are now being used to provide more accurate measurements of critical concentration for some of the major nutrients in plants.The object of this paper is to re-evaluate the potential of rapid tests for the determination of plant nitrate, phosphate and potassium in the light of these advances and to attempt to pinpoint some of the remaining problems with their use and interpretation. Analytical Procedures The potential use of commercially available test papers and test kits for the measurement of nutrients in plants was first recognised by Scaife and Bray,’ who found that the Mercko- quant nitrate test strips could be adapted for the determination of nitrate in sap. Any nitrate present in sap squeezed on to the colourless active zone of these strips is first reduced to nitrite, which then initiates a diazotisation and coupling reaction, creating a purple colour that is allowed to develop for 2 min.This method is used to determine nitrate concentrations up to 500 mg 1-1 directly, either by comparison with a calibrated scale or, for added precision, by use of a simple inexpensive reflectometer,z commercial versions of which are just becom- ing available. Concentrations above 500 mg 1 - 1 do not produce any darker colouration, but can still be determined. with reduced precision, from the time taken for the colour generated to match that of the top standard by using a specially prepared calibration curve.z.3 Although there is some evidence that the test strips underestimate nitrate concentrations in plant sap’ (see also Table I ) , they are now widely used to assess nitrogen status and to diagnose nitrogen deficiency in crops all over the world.‘ Recently, however, an alternative solution technique has also been evaluated.This is based on an adaptation of the Hach nitrate test procedure. in which the colour developed by the addition of a reagent to a diluted sample of sap is measured in a simple comparator.’ The method has the dual advantage of using a common procedure for the whole range of nitrate concentrations found in plants and of providing results of greater accuracy and precision than the test strip method (see Table 1 ) . Table 1. Comparison of the rapid nitrate tests with a standard HPLC method using sap samples. Concentration range: 0-2200 mg 1 I of nitrate Regression line Variance Test accounted method DF Slope S.E. for. o/o Merck 199 0.687 0.020H 75.6 Hach 199 0.964 0.0 140 92.4 Merckoquant test strips have also been used for the determination of potassium in sap.6 These strips are supplied with a brightly coloured active zone containing dipicrylamine.part of which becomes chemically “fixed” when sap containing potassium is squeezed on to it. Excess reagent is removed by immersing the strip in dilute acid. allowing potassium concen- trations (up to 2000 mg 1 - 1 ) to be determined directly by use of a coloured scale. The presence of ammonium interferes with this reaction, but its concentration in sap is normally too small to be significant. However, the test was found to be sensitive to low relative molecular mass amines, which accumulate in K-deficient plants. This, together with the associated increase in colouration of the sap from such plants.causes some overestimation of potassium concentration below 1000 mg 1-1.6 No equivalent test strips are available for phosphate, but accurate determinations of its concentration in sap have been obtained by a molybdenum blue method based o n an adapta- tion of the Hach orthophosphate test using a measured drop of sap.h The method follows a similar procedure to that used for the determination of nitrate (above). Sampling Procedures When plants are small, representative samples of sap can conveniently be expressed from the whole shoot by using disposable plastic syringes. The resultant sample is often coloured but its clarity can be improved. either by prior freezing of the plant material or by passing the sap through a microfilter before analysis.ANALYTICAL PROCEEDINGS, APRIL 1988, VOL 2.5 2.5 ._ 2 I 2.0 F m E 1.5 1.0 " 0.5 a, 0 123 - - - - - Where large plants are involved, whole-shoot sampling is impracticable and a suitable sub-sample must be selected from which to express the sap.A young mature leaf is normally recommended as being the most appropriate part of the plant to sample-7 with the sap expressed from its petiole to reduce the risk of colour interference. However, experience has shown that it can be difficult to recognise just which is a young mature leaf, especially in annual crops, where there is often a gradation of leaf sizes over the whole plant. Trends in the natural distribution of nutrients between leaves (particularly for nitrate and potassium) are often quite large in the "mature" region (lower third) of the shoot.so a small error in leaf selection could have an important effect on the interpretation of the result. In addition, the variability in nutrient concentra- tions appears to be greater when measurements are made on older leaves (see Table 2), which suggests that mature leaves may not be the most appropriate choice for analysis. Table 2. Average percentage coefficients of variation of individual measurements (based on data from 4 replicates) for plants of medium to high nutrient status Organ sampled Nitrate" Potassium Phosphate Young leaf sap 19.2 12.1 17.3 Middle leaf sap 1.5.3 14.9 22.7 Old leaf sap 38.0 18.7 27.1 Total shoot/dm 8.3t 6.1 9.9t * Merckoquant test strip method only 'i Data for total N or P in shoot. In annual crops, the young leaves seem to be most sensitive to changes in the nutrient supply.Comparisons of the distributions of nitrate, phosphate and potassium in both non-deficient plants and those in the early stages of deficiency have shown that the biggest changes occur in younger leaves, with those in the middle and older ones occurring progressively more slowly (see Fig. 1). When these nutrients are re-supplied after a period of starvation, they are preferentially directed to the young leaves at the expense of the older ones.8 However. these leaves also seem to be more sensitive to the presence or absence of other (non-specific) ions, such as sodium or chloride, which can build up when potassium or nitrate are in short supply (see below). This may make it too difficult to obtain reliable determinations of critical concentration for young leaves and, at present, the best compromise appears to be to select leaves from the middle part of the plant.Only limited information is available about the number of leaves required in each sample for analysis. Best estimates from tissue analyses of leaf dry matter suggest that up to 80 plants should be sampled, with the leaves split into four equal groups.7 Similar numbers are likely to be needed for sap analysis, but our experience has suggested that representative samples of sap cannot be obtained when this number of petioles are pressed simultaneously (even when they have been finely chopped). We have found it better to sub-divide the petioles into groups of about five before pressing and to combine the resultant samples of sap into larger batches for analysis.If the data in Table 2 are assumed to be typical. sampling as many as 80 plants would reduce the coefficient of variation of measurement on young and middle leaves to less than 5% in all instances. By comparison, a maximum of only 16 samples would be needed to obtain the same precision for the corresponding elemental analyses of the total shoot dry matter. Critical Concentration and the Use of Rapid Tests in Practice Practical use of rapid tests can only be made if the minimum concentrations of nutrients required in plants are already known. In order to determine these minima each crop is, by tradition, grown at a range of fertilizer rates and the final yields r 1 1.25 1 .oo 0.75 0.50 0.25 m 0 1 2 3 4 5 6 Leaf number 3.5 r p 3.0 1 Potassium \ u t 1 I 1 1 1 1 Leaf number 0 1 2 3 4 5 6 7 8 9 I A .'../"p..'".. - 1 0 0 1 20 401 1 1 1 1 1 1 I " I J Leaf number o 1 2 3 4 5 6 7 8 Fig. 1. Distributions o f nitrate. potassium and phosphate between individual leaves of non-deficient plants (0) and deficient plants (A 1. Leaf 1 is the oldest lead sampled plotted against the internal nutrient concentrations measured at any early stage of growth.7 The critical concentration is then defined as that which corresponds to 90% of maximum yield, on the grounds that if the concentration is maintained above this level, yields will not be significantly restricted. Experience has shown that critical concentrations defined in this way can be unreliable,y as they depend on when the concentration is measured and do not take account of changes in nutrient availability (e.g., due to leaching or drought) after the measurement has been made.More consistent results can often be obtained if the critical concentration is determined from measurements of yield and nutrient concentration made at the same stage of growth. However, it is not always appreciated that errors are still liable to occur if these measurements are made using this method. The problems arise because, although the yields and concen- trations are measured simultaneously, the growth effects are actually generated over the whole of the previous lifetime of the crop and so the measured concentrations are more likely to be the result rather than the cause of the reductions in growth.The concentrations which led to the growth effects would normally have been higher than those recorded on the sampling day. Recently. an alternative method, which overcomes these difficulties, has been developed. This is based on the measure- ment of the reductions in growth of non-deficient plants following interruption of their nutrient supply. 1 0 Young plants of the required size are transferred to a deficient treatment from which a chosen nutrient is missing and the changes in their mass and nutrient concentration are measured over a series of harvests at about 1- or 2-d intervals as the deficiency develops.I24 ANALYTICAL PROCEEDINGS, APRIL 1988, VOL 25 The growth of the crop (expressed as a fraction of the growth of an unstressed control treatment) is plotted against its nutrient concentration in order to obtain curves of similar shape to those of the traditional method.from which the critical concentration can be obtained (see Fig. 2). The advantage of this method originates from the short intervals between harvests, and means that the concentrations measured are more directly related to the changes in growth. In addition, because growth is regularly monitored, critical concentrations based on changes in growth rate can also be determined. 1 .o 0.9 0.8 0.7 !!! 0.4 0.3 0.2 0.1 a / I I I I I I I I I I I I I I I I I I I I I I Critical I y n c e n t r a t i o n I \J I I I I I I I 1 1 1 I I I 0 10 20 30 40 50 60 70 80 Concentration/mmol 1-1 Fig. 2. middle leaf in the presence (0) and absence (A) o f sodium Relation between growth and potassium concentration of The use of this more controlled method has enabled critical concentrations to be measured more accurately and, at the same time, allowed other inter-related factors, such as non- specific ion effects, to be reliably determined (see Fig.2). Such effects are believed to originate from the ability of ions such as sodium and chloride to replace potassium and nitrate in their osmoregulatory roles within the plant, causing changes in critical concentrations which are most pronounced in rapidly expanding young tissues. 1 1 Conclusions The development of simple new techniques for the rapid measurement of sap nutrient concentrations has improved the reliability and ease of use of the analytical procedures involved.This momentum will continue as the new generation of sensors (including reflectometers and photometers for the measure- ment of colour on both test papers and in solution) now being developed becomes available. These are likely to be multi- purpose devices, which can be programmed to allow the operator to select one of a range of different tests, to provide step by step procedural instructions. to give warnings when errors occur and to output the results directly in units of concentration. Similar improvements in the techniques for characterising critical nutrient concentrations in plants have recently been made and these are likely to lead to more accurate determina- tions of the minimum target levels required for maximum growth, particularly for small plants, where sap can be expressed from whole shoots.However, these developments will only result in more reliable tests for larger plants if greater attention is paid to the choice of the best part of the shoot to select for analysis. The present trend towards analysing mature leaves for arable crops seems to be ill founded. These leaves are less sensitive to changes in nutrient supply and have a larger error of measurement than young expanding or middle leaves. However, preliminary evidence suggests that the younger the leaf. the more susceptible it is to non-specific ion effects. and more work is required to determine the practical significance of this before clear-cut recommendations of the best sampling procedure can be made. I . 3. 3 -. 4. 5 . 6. 7. 8. 9.10. 1 1 . References Scaife. M. A.. and Bray. B. G..ADAS (2. Re\,.. 1977.27. 137. Schaefer. N . L.. Soil Sci. Hutit Ajiul.. 1986. 17. 037. Scaife. A . . and Stevens. K. L.. Soil Sci. Plarit Atiul.. 1983. 14. 761. Prasad. M.. and Spiers. T. M.. SoilSci. Plunt And.. 1984. 15. 673. Burns. I. G.. and Griffin. D. A.. i~ "Report of the National Vegetable Research Station for 1986/87." National Vegetable Research Station, Wellesbourne. 1987. p. 90. Burns, I. G.. and Hutsby. W . . Soil Sci. Plutif A d . . 1984. 15. 1463. Ulrich. A.. in Reisenauer. H. M.. Ediror. "Soil and Plant- Tissue Testing in California." Bulletin 1879 (revised). Univer- sity of California. USA. 1978. pp. 1-4. Burns. I. G . . and Hutsby. W . . J. Sci. Food Apric.. 1986. 37. 11s. Scaife. A.. and Turner.M. K. iii "Proccedings of the 6th International Colloquium for Optimization of Plant Nutrition. Montpellier. 2-8 September 1984.'. Pierre Martin-Prevel. Montpellier. 1984. pp. 617-624. Burns. I. G.. Plum arid Soil. 1986. 94. 301. Burns. I. G.. i r i "Report of the National Vegetable Research Station for 1986/87." National Vegetable Research Station. Wellesbourne. 1987. p. 89. Approaches to the Analysis of Animal Feedstuffs A. Smith Rowett Research Institute, Greenburn Road, Bucksburn, Aberdeen AB2 9% Animal feedstuffs have been analysed traditionally for their chemical composition. The analysis of not only commodities but also of by-products and waste products. which can be incorporated into animal diets, has now become more sophisti- cated and detailed with a view to gaining the maximum nutritional advantage from the ingredients used in the formula- tion of diets.Particular attention has been paid to the dietary content of protein, fat, fibre, sugars and inorganic elements in addition to anti-nutritive factors which occur in many of the commodities used in the preparation of diets. This paper reviews current methods used in the determination of these key parameters. Probably the element most analysed in feedstuffs is nitrogen. the concentration of which can be related to the protein content. Although there are a number of methods for estimating nitrogen, the three most applicable to feedstuffs relate to the Dumas and Kjeldahl methods and near-infrared reflectance. 1 The classical Dumas method involves the catalytic oxidation of the sample to effect the total conversion of nitrogenous material to nitrogen oxides, which are then reduced to nitrogen and other products of combustion such as carbon dioxide.water and inorganic acids. This method has been incorporated into an automatic elemental analyser in which the elementalANALYTICAL PROCEEDINGS, APRIL 1988, VOL 25 125 nitrogen is separated from other incidental combustion gases by gas chromatography prior to thermal conductivity detec- tion. The standard method of estimating nitrogen in most biological samples involves the Kjeldahl method with detection of the nitrogen after digestion of the sample either titrimetric- ally or spectrophotometrically; the spectrophotometric method usually involves the indophenol blue (Bertholet) reaction. Automated systems are available commercially for digestion and determination of the nitrogen by titration.The technique of near-infrared reflectance is predictive and depends on the construction of calibration graphs to predict the nitrogen content of feedstuffs; the calibration graph of a commodity is peculiar to that commodity and is constructed from data obtained by chemical analysis. Although the major use of the near-infrared reflectance technique relates to the prediction of the nitrogen content of (particularly) cereals. it is used to a limited extent in the determination of nitrogen in formulated diets. The importance of amino acids in animal nutrition may be gauged from the plethora of methods which have been developed for this type of determination.Amino acids are released from proteins by acid hydrolysis. by performate oxidation prior to acid hydrolysis or by alkaline hydrolysis. The released amino acids are determined after separation by cation-exchange chromatography followed by detection with ninhydrin according to the Moore and Stein method.' However, the advent of high-performance liquid chromato- graphy (HPLC) has introduced an additional element into the determination of amino acids. Reversed-phase HPLC columns have been used in conjunction with pre-column derivatisation methods to separate and quantify most of the amino acids in protein hydrolysates. One of the earlier derivatives used in this approach was that using o-phthalaldehyde with detection effected fluorimetrically.3 The disadvantage with this reagent is that it does not form fluorescing derivatives with secondary amino acids. Phenylisothiocyanate (PITC).which was originally intro- duced by Edman for the determination of amino acid sequences in proteins and peptides. is another reagent which has been used in the HPLC determination of amino acids.4 PITC reacts with the amine group of all common amino acids and the resulting derivatives are detected by ultraviolet spectrophotometry. The yields are good for the common amino acids but the products tend to be unstable and, because detection is by ultraviolet spectrophotometry after reversed- phase HPLC separation, the sensitivity of the method is not as high as that with derivatives which can be detected fluorime- trically. A recent derivative used in the determination of amino acids is 9-fluorenylmethyl chloroformate.which was developed originally as a protective group in peptide synthesis.5 The reagent forms derivatives with secondary amino acids but it is necessary to extract the derivatives to eliminate interfering components before HPLC. Numerous attempts have been made to use gas - liquid chromatography for the routine determination of amino acids. Derivatives involving N-acetylation of the amino group and alkylation of the carboxyl group have been used but. despite the use of conventionally packed columns and high resolution capillary columns. it has been difficult to obtain a derivative which is suitable for all amino acids. However. r-butyldimethyl- silylation of both the amino and carboxyl groups yield stable derivatives of all the amino acids.h The derivatives are amenable to gas - liquid chromatography and they may prove to be the derivatives which will allow gas - liquid chromato- graphy to be used on a routine basis for amino acid determination.Although ion-exchange chromatography is the accepted standard method for amino acid determination, it is likely that HPLC, particularlv with the continuing development of reagents suitable for the formation of derivatives, may well become the analytical tool whereby amino acids can be determined with high sensitivity and with fast throughput of samples. The value for the content of fat in feedstuffs relates to that fraction of lipid material which is extracted with a non-polar solvent such as petroleum spirit or diethyl ether.Lipid thus extracted does not include polar material such as phospholipids nor that component of the feed which may be in the form of a complex or as fatty acid soaps. This disadvantage is overcome by an acid hydrolysis procedure prior to solvent extraction and the values thus obtained give a good indication of the availability of total lipid for digestion by the animal. Fatty acid profiles are an integral part of the assessment of a diet for essential fatty acid status and these profiles are normally obtained by subjecting the crude fat extract to hydrolysis and methylation of the released fatty acids prior to determination by gas chromatography. Substantial errors can arise by this method in that the polyunsaturated fatty acids are very susceptible to oxidation during the removal of the solvent and the values obtained are significantly lower than the true concentration of these acids in the feed.To overcome this problem it is essential that the solvent from the crude lipid extract is removed at a low temperature and under nitrogen. Fibre has long been recognised as an indigestible and, to some extent. an undesirable component of animal feed, although in some forms it may be required for lipogenesis. The analysis of fibre has received much attention and a crude fibre value is often used as an index of the fibre content of a diet. Crude fibre is defined as the loss on ignition of the dried residue remaining after digestion of the sample with acid followed by alkali under specific conditions.The hemicellulose, cellulose and lignin content of a diet can be obtained by treatment with a neutral detergent solution, which extracts soluble material to leave the so-called neutral detergent fibre (NDF), and with an acid detergent solution resulting in an acid detergent fibre (ADF) value. The NDF fraction contains hemicellulose. cellulose and lignin whereas the ADF fraction consists of cellulose and lignin. A value for the cellulose content may be obtained from the loss due to ashing of the permanganate- treated ADF fraction, whereas lignin can be determined as the loss due to permanganate treatment of the ADF residue. Apart from the nutritionally active factors in feedstuffs for animals, anti-nutritive factors also occur. One of these factors. which has received much attention recently.is the presence of glucosinolates in rapeseed meal. a common ingredient in formulated diets in that it has a high protein content and a balanced amino acid composition. On hydrolysis by the naturally occurring enzyme, myrosinase, glucosinolates yield a variety of compounds, among which are isothiocyanates, thiocyanates. nitriles and oxazolidine-2-thiones. Some of these compounds can damage the liver and others can initiate the onset of goitre. Most of the analyses for glucosinolates in rapeseed meal involve the release of glucose which can be determined either enzymatically or spectrophotometrically and which can be related directly to the glucosinolate concen- tration.' Other methods for determining glucosinolate concen- trations include gas - liquid chromatography either singly or in combination with mass spectrometry.# mass spectrometry using various ionisation modesY--l" and HPLC. Each method has its advantages but it is likely that HPLC.because of its versatility and sensitivity, and particularly as plant breeders can now produce rapeseed with very low or indeed negligible levels of glucosinolates. will become the accepted technique whereby glucosinolates can be determined in a routine manner in feed-trade laboratories. I n terms of instrumentation which may be used in future in laboratories which are concerned with the routine analysis of animals feed, it is likely that instruments which are highly cost effective in terms of sample throughput will be required. Included among those are likely to be near-infrared reflectance126 ANALYTICAL PROCEEDINGS, APRIL 1988, VOL 25 instruments for moisture, protein and fibre determination, flow injection analysers for cation and anion determination and inductively coupled plasma atomic emission spectrometers for inorganic elements. References 1. Norris. K . H . , and Butler. W. L . . IRE Trtms. Biomed. Electron.. 1961. 8. 153. 2. Moore, S . , Spackman. D. H.. and Stein. W. H.. Anal. Chenz., 1958, 30. 1185. 3. Lindroth. P.. and Mopper. K., Anal. Chetii.. 1979. 5 1 . 1667. 4. 5. 6. 7. Cohen. S . A . . Tarvin, T. L.. and Bidlingmeyer. H . A , . Am. Lab.. 1984. August. 48. Carpino. L. A.. and Ham. G. Y . . J . Org. Chem.. 1O72.37.3404. Goh. C.-J.. Craven, K. G . . Lepock. J . R.. and Dumbroff. E. B.. And. Biochem.. 1987. 163, 175. Heaney. R. K.. and Fenwick. F. R.. it? "Methods of Enzymatic Analysis." Third Edition. Verlag Chemie. Weinheim. 1981. p . 208. Spencer, G. F.. and Daxenbichler. M. E., J . Sci. Fuod Agrrc.. 1980, 31. 359. Eagles. J . . Fenwick. G. R . . and Heaney. R. K.. Riomed. iMuss Specfrom., 1981. 8. 278. Fenwick. G. R.. Eagles. J . . and Self. R.. Org. Mass Spectroni.. 1982. 17. 544. 8. 9. 10. TWO NEW BOOKS FROM THE AMERICAN CHEMICAL SOCIETY Controlled-Release Technology: Pharmaceutical Applications ACS Symposium No. 348 Edited by: Ping I. Lee and William R. Good. Ciba-Giegy Corporation Controlled-release technology is a rapidly emerging, interdisciplinary science that offers novel approaches to the delively of bioactive agents. This excellent new publication examines recent advances in controlled -release technology, with emphasis on pharmaceutical applications. The 25 chapters are divided into the following more general categories and are preceeded by an overview of controlled release delivery. Fundamental aspects Characterization methodologies * Hydrophilic polymers and hydrogels * Transdermal and transmucosal delivery systems * Other applications. Essential reading to a l l those interested in controlled-release technology. ISBN: 0 84 12 14 13 1 Hardbound 353 pp. Price: €62.00 The Chemistr of Acid Rain: Processes Sources and A f mospheric ACS Symposium No. 349 Edited by: Russel W. Johnson, Signal Research Center. Glen E. Gordon, University ofMaryland During the past decade, significant progress has been made in our understanding of acid deposition, although large gaps in our knowledge still exist. For instance, methods for collection and analysis of wet deposition are well established, but understanding of dry deposition remains poor. And, although the importance of OH radicals is clear, no reliable, portable instrumentation for real- time measurement of their concentrations a t low altitude has been developed. This book is timely due to growing concern for both the environment and 'acid rain.' It describes our growing understanding of the sources and chemistry of acidic species in the environment. ISBN: 0 8412 1414 X Hardbound 337 pp. Price: g53.00 ORDERING: RSC members should send their orders to: The Royal Society of Chemistry. Membership Manager, 30 Russell Square. London WClB 5DT. UK. Nonmembers should send their orders to: The Royal Society of Chemistry, Distribution Centre. Blackhorse Road, Letchworth, Herts SG6 1HN. UK. - ROYAL Information Services

ISSN:0144-557X    

DOI:10.1039/AP9882500115

出版商:RSC    

年代:1988

数据来源: RSC

摘要:

ANALYTICAL PROCEEDINGS, APRIL 1988, VOL 25 127 The Quest for Sensitivity, Specificity and Reliability of Diagnostic Reagents-a Biological Answer The following are summaries of two papers presented at a Meeting of the Biological Methods Group, held on June 25th, 1987, at the Central Public Health Laboratory, Colindale, London. Chlamydia Detection with Monoclonal Antibodies Caroline Ash IQ(Bio) ltd., Downham House, Downham‘s Lane, Milton Road, Cambridge CB4 IXG The aim of this paper is to describe the background to the development of Chlamydia-IDEIA. a Chlamydia antigen detection test designed, developed and manufactured by IQ(Bio) for distribution by Boots Celltech Diagnostics. Chlamydia trachorziatis. an intracellular Gram-negative bac- terium, is now recognised as the most common sexually- transmitted infectious agent.Over 12 000 cases have been reported in the UK so far this year. It is recognised as a major cause of tuba1 infertility and it has been estimated that up to 50% of women seeking in vitro fertilisation show evidence of past infection with C. trachornatis. 1 To avoid the long-term consequences of infection early diagnosis is vital, but conditions such as pelvic inflammatory disease are difficult to identify and greater facilities for diagnosis are urgently required. There are currently four types of diagnostic aid available to detect Chlumydia: (a) direct detection by immunofluorescence. (b) tissue culture, which is currently used as a “gold standard,” (c) serology, which is only used for confirmation as antibody titres remain persistently high post-infection and (d) enzyme-linked immunosorbent assays (ELISA).The use of ELISA has many advantages over the other three methods; it demonstrates the presence of current infection but the viability of the specimen does not have to be maintained, unlike with tissue culture. because both whole organisms and shed antigen can be detected. In addition. results from ELISA are obtained within hours with the possibility of same-day feedback to patients, and a further advantage is that many specimens can be processed simul- taneously. It is the ELISA method that has been exploited in the Ch lam y dia- I D E I A . Experimental Immunoreaction The antigen detected in IDEIA is a repeated genus-specific epitope of lipopolysaccharide which consists of an immuno- dominant trisaccharide of 3-deoxy-~-mannooctulosonic acid.’ So, theoretically it can also be used to detect the only other Chlamydia species, C.psittaci. The detecting monoclonal antibody was generated by M. Thornley of the Department of Pathology, Addenbrookes Hospital, Cambridge.3 The principle of IDEIA is a sandwich ELISA. The monoclo- nal antibody is coated on to the surface of a polystyrene micro-titre plate, 200 pl of heat-treated specimen is added in a specially formulated transport medium and an incubation step follows for 2 h. during which time any antigen present in the specimen will be captured by the antibody on the plate. Alkaline phosphatasc-labelled antibody is then added to the well and incubated for a further hour to allow binding to the repeating epitope.After thorough washing, the results of the immunoreaction are visualised by IQ’s cycling amplification system. Amplification Amplification is required to boost sensitivity when using highly specific monoclonal antibody detection systems. In IDEIA, after the immunoreaction has been stopped. substrate (NADP+) is added to the plate-well. If alkaline phosphatase is present the NADP+ is dephosphorylated to NAD+. In the second stage of the reaction, when amplifier is added to the well, the first reaction is halted by the presence of phosphate in the buffer which inhibits alkaline phosphatase. NAD+ pro- duced during the primary reaction then catalyses a redox cycle between the secondary amplifier enzymes alcohol dehydro- genase and diaphorase.Diaphorase continually reduces the iodonitrotetrazolium in the amplifer buffer to give an intense red formazan colour (Fig. 1). The effective separation of the two steps maintains linear reaction kinetics. NADP- [Formazan) K NAD+y Ethanol Diaphorase Alcohol dehydrogenase INT ‘NADH! I Acetaldehyde Fig. 1. Principles of IDEIA-Chlamydia assay The constraints on practical applications of enzyme amplifi- cation are considerable. The selection of three compatible enzymes is a major restraint, purity of reagents is vital and as the secondary cycling enzymes have to work in the presence of a large excess of primary substrate this must not act either as a substrate for the secondary enzymes nor must it inhibit them.4 Results By using enzyme amplification the detection of a few thousand molecules of alkaline phosphatase can be demonstrated, but this is seldom achieved in an immunoassay. The sensitivity is128 ANALYTICAL PROCEEDINGS, APRIL 1988.VOL 25 Table 1. Chlamydia IDEIA and tissue culture results from various study populations Culture Sensitivity." Yo Specificity.: % incidence 339/ 18 10 93.3 (70/75) 9 1 .h (342/364) 98.3 (lO7/1OO) 98.5 (1341113612) Male Female Malc Female 92.0 (3121339) 98.4 (144811471) Co-positive by IDEIA and tissue culture Total tissue culture positive Co-negative by IDEIA and tissue culture Total tissue culture negative * Sensitivity = f Specificity = vulnerable to background noise from raw materials. impure reagents and sample matrix effects. However, the ultimate C. psiffaci pneumonia. It has recently been used to detect feline cases of C.psittaci.7 limitation to sensitivity is the efficiency of specimen collection. Results obtained from clinical specimens show that IDEIA performs well in relation to tissue culture. The data shown in Table 1 were collected from a total of six studies on both low and high risk populations.'.f1 &. 1. 3 3. 4. Discussion The major advantages that IDEIA has compared with other Chlamydia detection systems are that it is highly specific, rapid and has good sensitivity. It is particularly valuable for IDEIA has a wide range of potential applications, subject to screening, ocular infections, ear. nose and throat infection, and 5 . 6. 7. processing large numbers of genito-urinary specimens. thorough prior evaluation. such as pre-natal and neo-natal References Editorial. Lancer.April 1986. 892. Brade, H.. Brade. L.. and Nano. F. E.. Proc. Nrrrl. Acurf. Str.. 1987. 84. 2508. Thornley. M.. Thornley. M. J.. Zamze. S. E.. Byrne. M. D.. Lusher, M.. and Evans. R. T.. J. Gen. Micro.. 1985. 131, 7. Johannsson. A , . and Bates. D. L.. rn Kemeny and Challacombe Editors, "Theoretical and Technical A\pects of ELISA and Other Solid-phase Immunoassays." Wiley. New York. 1987. Caul, E. 0.. and Paul. I. D . . Lancer. February 1985. 279. Pugh. S . F.. Slack. R. c'. B.. Caul. E. 0.. Paul. I. D.. Appleton. P. N.. and Gatley. S . . J. Clin. Patiid.. 1985. 38. 1139. Wills. J . M.. Millard. S . (3.. and Howard. P. E.. VPr. Rec.. October 1986. 418. Cloned Nucleic Acid Probes for Detection and lndentification of Legion el I a e N.A. Saunders, N. Kachwalla, T. G. Harrison and A. G. Taylor Legionella Reference Unit, DMROC, CPHL, 61 Colindale Avenue, London NW9 5HT Legionnaires' disease (LD) is an often fatal systemic illness of which the primary symptom is pneumonia. Legionellae, the causative organisms of LD. are ubiquitous, water-borne. Gram-negative bacteria which comprise 23 species belonging to a single genus (Legionella). The majority of cases of LD are associated with Legionella pneumophiia but infection caused by 11 other Legionella species has been demonstrated by culture. Current serological and biochemical indices are inadequate for the identification of strains of all species of Legionella. which in some instances are exclusively defined on the basis of DNA homologies.Furthermore. culture of these organisms is complex and may require from 2 to 10d to complete. We have developed a nucleic acid probe for the recognition of legionellae which offers the possibility of rapid detection and identification of all species of the genus. Experimental Construction of an L. pneumophila Genome Library DNA isolated from the Knoxville-1 strain of L. pnerrrmphila was used to prepare a gene library in the vector hgtWES.hB.1 using methods described by Maniatis et a1.2 The AB. fragment was excised with the restriction endonuclease EcoRl and replaced by EcoRl restriction fragments (2-15 kb) derived by complete digestion of the genomic DNA. Recombinant phage resulting from in vitro packaging were plated out on the E. coli host strain NM531 (supE supF hsdR recA).; Screening of rRNA Genes The library was screened by plaque filter hybridisation' on Hybond-N (Amersham International). The probe was pre- pared from a mixture of 16 and 23s rRNAs from L. pneumophila (Knoxville- 1 strain), which were subjected to polynucleotide kinase labelling with [3'P]-y-ATP following partial alkaline hydrolysis.' Northern Blotting RNA from L.pneiirriophiia Knoxville-1 strain was subjected to agarose gel electrophoresis (1% gel) in the presence of formaldehyde and blotted on to Hybond-N.2 The blot was hybridised to hl42. which had been labelled by nick-translation with [3'P]-(x-dCTP.-' Fragmentation and Selection of rRNA Gene Sequences The EcoRl insert fragment from A142 was purified and digested with the restriction endonuclease Sau3A.The result- ing fragments were ligated into the phosphatase treated BamHl site of the plasmid pT7-1 (Boehringer Mannheim). The recombinant molecules generated were used to transform calcium-shocked E. coli (strain HB 1 0 1 ) . Plasniids from ampi- cillin resistant colonies were extracted by the method of Birnboim and Doly.' Partially purified plasmids were subjec- ted to agarose gel electrophoresis in Tris - borate buffer pH 8.3 (1% gel) and blotted on to Hybond-N.') Membrane4 wereANALYTICAL PROCEEDINGS, APRIL 1988, VOL 25 probed and re-probed as recommended by the manufacturers. The [.3'P]-labelled probes were derived from RNA isolated from L. pneumophila, E. coli or Ps. aeruginoscr by reverse transcription in a reaction mixture containing M-MLV reverse transcriptase.[J'PI-cw-dCTP and random primers.' Dot-blot ting DNA isolated from Legionella and other species by the rapid procedure of Owen and Borman7 was spotted on to Hybond-N (200ng per spot). The filter was probed with [3'P]-labelled RNA transcripts of the insert sequence of pNS9. The transcrip- tion reaction included linear plasmid template (cut with restriction endonuclease Pst l ) , T7 RNA polymerase and [-I'P]-(x-CTP, under conditions recommended by Boehringer Mannheim. Results and Discussion When the L. pneumophila gene library was screened with end-labelled rRNA fragments, approximately 0.5% of the clones gave a strong positive signal. One of these clones (h132) was purified and used to probe a Northern blot of L. pneumophila RNA. Both the 23 and 16s rRNA species hybridised to the probe and gave a signal proportional to the quantity of rRNA sub-units in the RNA preparation.This result suggests that A132 includes at least a high proportion of the complete sequence of both major rRNA molecules and that these genes are clustered. The cloned rRNA gene was fragmented with the restriction endonuclease Sau3A and sub-cloned into the plasmid vector pT7-1, which has the phage T7 promotor $10 adjacent to a polylinker sequence. These recombinant plasmids were screened with complementary DNA (cDNA) prepared from the RNA of L. pneumophila, E. coli and Ps. aeruginosa. The additional species were chosen on the basis of their taxonomic relationship with the genus Legionella and, in the case of fs.aeruginosa, because this species is commonly encountered in specimens suspected of harbouring legionellae. A proportion of the cloned plasmids did not hybridise to any of the cDNA probes. These plasmids probably represent recombinants containing sequences from the A142 insert flanking the rRNA gene. Several plasmids hybridised to the L. pneumophila cDNA under stringent conditions and were therefore identi- fied as having rRNA gene inserts. The strength of the signal was in proportion to the size of the inserts. The levels of hvbridisation of the plasmids to the cDNA probe from Ps. aeruginosa and E. coli were always very similar, however, there were gross differences when the reaction of these cDNAs was compared with that of the homologous cDNA. One plasmid (pNS9), which only gave a signal in the homologous system, was chosen for further study.The size of the insert sequence was judged to be approximately 150 bp by agarose gel electrophoresis. Plasmid pNS9 was converted to linear form, ready for the specific run-off transcription of the insert sequence, by cutting the polylinker distal to the $10 promotor. A major advantage of the RNA transcription system is that the probes produced are virtually free of labelled host and vector sequences, which would otherwise tend to hybridise non- specifically. The [3'P]-labelled RNA probe was hybridised to a dot-blot of purified DNAs which was then washed under conditions of high stringency. The probe hybridised strongly with representative strains of the following Legionella species, L.pneurriophila (serogroups 1-3, 5-1 l), L. bozemanii, L. dumoffii, L. santicrucis. L. cherrii, L . anism, L . longheachae. L. niicdadei, L . jordanis, and L . feeleii. A very weak reaction was obtained with species of other genera, E. coli, Ps. aeruginosa, H. influenzae, N . meningitidis, S. pneumoniae and Listeriu monocytogenes. The weak reaction observed with the non-legionellae did not appear to correspond to the taxonomic relationships between the species involved and the genus Legionella and might be attributable mainly to non-specific reactions. Elimination of these reactions will be an important objective of further studies as the probe will be of most use if the ratio of reaction with legionellae to reaction with non- legionellae can be maximised.It will also be of interest to use the probes derived from pNS9 and similar plasmids as reagents for the detection of legionellae in enriched cultures and in clinical or environmental specimens. References I . Tiemeier. D . , Enquist. L.. and Ledcr. P.. Nafure (London). 1976. 263. 526. 2. Maniatis. T.. Fritsch. E. F.. and Sambrook. J . . "Molecular Cloning: A Laboratory Manual .'* Coldspring Harbor Labora- tory. New York. 1982. Kaiser. K.. and Murray. N. E.. in Gover, D. M.. Ediror. "DNA Cloning." Volume 1, IRL Press. Oxford. 198.5. pp. 1-47. Rigby. P. W. J . , Dieckmann. M.. Rhodes, C.. and Berg. P.. J. ,4401. B i d . 1977. 113. 237. Birnboim. H. C . . and Doly. J . . Ncicl. Acids Res.. 1979. 7, 1523. Owen. R. J . . and Borman, P.. Nuc-1. Acids ReJ.. 1987. 15.3631.3. 4. 5 . 6. Southcrn, E. M.. J . Mol. R i d . 1975. 98. 503. 7. Development of a Diagnostic Reagent for the Detection of Adenovirus Antigen by Immunofluorescence 1. R. Sharp Division of Microbiological Reagents and Quality Control, Public Health Laboratory, Colindale Avenue, London N W9 5HT A. Bailey North Manchester Regional Virus Laboratory, Booth Hall Childrens Hospital, Manchester M9 2AA There are 41 human adenoviruses so far discovered. causing a wide range of diseases. They have a predilection for respira- tory. ocular or gastrointestinal sites. They generally cause mild self-limiting infections but are responsible for a considerable degree of morbidity within the population. They can cause pneumonia and disseminated infections, particularly in the immunosuppressed.Currently, diagnosis relies on isolation in tissue culture. which is time consuming and unreliable. or by electron microscopy. which is a facility not always available to routine laboratories. Immunofluorescence of clinical material or cell cultures. 48 h after infection, is a rapid, easy to perform assay. However, good reagents have not been routinely available. This paper describes the production and characterisation of monoclonal antibodies reactive against the group specific antigen of adenovirus and their development as a diagnostic reagent. Experimental Monoclonal antibodies were raised against adenovirus types 3, 4 and 5 according to the method of Kohler and Milstein,' modified as previously described.'130 ANALYTICAL PROCEEDINGS, APRIL 1988, VOL 25 Immunofluorescent Assay (IFA) Volumes (10 pl) of antibody dropped on smears or infected cell monolayers prepared on multi-spot slides (Flow Laboratories) were incubated at 37°C for 30 min.This was followed by 10-min washes in the indirect assay and a 5-min wash in the direct assay. Preparation of Hexon Antigen for ELISA Briefly, adenovirus type 5 infected HEP 2 cells were harvested into Tris buffer (0.01 M, pH 8. l), disintegrated ultrasonically and treated with fluorocarbon.' The aqueous phase was precipitated in 55% saturated ammonium sulphate solution. dialysed against Bis-tris - propane buffer (30 PM, pH 6.8) and applied to a Mono Q column in a fast protein liquid chromatograph (Pharmacia). A salt gradient in Bis-tris - propane buffer was applied to the column and hexon eluted (at a salt concentration 0.43 M), resolving as a single band in sodium dodecyf sulphate (SDS) polyacrylamide gel electro- phoresis (SDS-PAGE).ELISA Plates were coated overnight at 4 "C with 100 1-11 of hexon, 10 1.18 per well, in hydrogen carbonate buffer at pH 9.6. The plates were quenched with 1% bovine albumin in phosphate buffered saline (PBS). Volumes of 100 1-11 per well of crude ascitic fluid or purified IgG, diluted in PBS containing 0.05% of Tween 20. were incubated at 37°C for 2 h. The plates were washed three times in PBS - Tween 20 and 100 p1 of horseradish peroxidase conjugated antimouse antibody (Tago) was added. After 2 h at 37 "C the plates were washed as before and substrate hydrogen peroxide plus o-phenylenediamine was added.The reaction was stopped with 2 M sulphuric acid after 30 min. Competitive ELISA Fifty microlitres of conjugated antibody plus 50 yl of uncon- jugated antibody were added simultaneously to each cell of a hexon coated plate. The plates were then incubated and substrate added as described above. Fluorescein Isothiocyanate Conjugation This was carried out by using the standard methods.4 For use the conjugate was diluted in Evans Blue (BDH Ltd.), 0.005% final dilution in PBS. Results Forty stable hybridomas were obtained, which reacted strongly in immunofluorescence when tested against their homologous serotype. Altogether, 13 different monoclonal antibodies reacted strongly by IFA against each of the 6 sub-groups, suggesting that they are group specific. They were then analysed by staph A immunoprecipitation' to determine which viral proteins they reacted against.Of the thirteen antibodies twelve produced a single virus specific band in SDS-PAGE, which corresponded to the hexon polypeptide, and one was reactive against penton base. Two of the 12, designated 5.1 and 93.4, were selected as being likely candidates for development as immunofluorescence reagents because of their high titre in IFA coupled with strong positive fluorescence, suggesting that they react against major antigenic epitopes with high affinity. They also react well in ELISA, making further characterisation easier. Sub-class Determination The IgG sub-class of the two monoclonal antibodies was determined in ELISA by using HRPO conjugated antimouse sub-class specific antibody (Miles Scientific).Antibody 5.1 turned out to be an IgG 2a and 93.4 an IgG 1. Competition Assays Fig. 1 shows the results of competition assays between 5.1 and 93.4. Each antibody competes with its homologous conjugated IgG as one would expect. No competition was observed between 5.1 and 93.4, indicating that they react against different epitopes on the group specific region of the hexon polypeptide. 1.4 1. 5.1 v93.4 HRPO 1 l.o - 93.4 v5.1 HRPO z +? 2 0.6 - a 0.4 - 13 0 . 2 p 5 , x - * 1 2 4 8 16 32 64 128 256 Competing antibody dilution x 100 Fig. 1. 93.4 Competition assays between monoclonal antibodies 5.1 and FITC Conjugation Each conjugate was titrated to find its optimum dilution and they were pooled together in proportions related to their titre.Virus specific fluorescence is bright yellow - green, which contrasts well against the red background of the Evans Blue counterstain. Field Trials The reagent was tested against a number of clinical isolates. Respiratory adenovirus isolates (126), encompassing 18 differ- ent serotypes, were tested by IFA after passage for 48 h on HEp2 or Grahame 293 cells. All were detected by IFA. Next, 95 enteric adenoviruses, confirmed by electron microscopy (EM), were examined by IFA after culture on Grahame 293 cells. Of these, 89 were positive on primary isolation and four were positive on further passage. Cell cultures of the two which remained negative by IFA were re-examined by EM and proved to be negative, indicating that the virus had not grown.Then, 44 non-adenovirus clinical isolates. being mainly enteroviruses or members of the herpesvirus group. were examined by IFA. All were negative. Table 1 illustrates the results obtained when clinical material from 75 patients was examined directly by IFA. Adenovirus was isolated from 15 in cell culture. nine of which were IFA positive. The six throat swabs missed by IFA were poor quality smears containing very few cells. Only three of the nine IFA positive smears were positive by indirect IFA using a commer- cially available polyclonal anti-adenovirus serum. Table 1. IFA of clinical material Immunofluorescence + ve No. CPE Source tested + ve Smears Culture Type Throat swab 44 11 5* 1 1 9ofType7 2 of Type 3 NPA 13 2 2 2 lofType2 1 of Type 3 Sputum 3 1 1 1 Type 5 Eye 2 1 1 1 Type 6 Other 13 Neg Neg Neg - * Negatives were poor specimens.see text.ANALYTICAL PROCEEDINGS, APRIL 1988, VOL 25 131 Sensitivity In order to evaluate the sensitivity of the reagent IFA was performed on HEp2 cells 4 h after infection with adenovirus type 3. Fine dots could be seen in the cytoplasm of the infected cells, representing hexon polypeptide beginning to accumulate immediately after its synthesis had commenced. Discussion A reagent for the detection of adenovirus was developed by using a pool of 2 non-competing monoclonal antibodies, both group specific and reactive against the hexon of the viral capsid. The hexon is strongly antigenic and accounts for about 36% of total viral protein. It is therefore an ideal target for an immunofluorescence reagent The use of a pool of two non-competing antibodies reduces the chances of missing a strain which does not possess a group specific antigenic epitope.It also increases the sensitivity of the reagent. In field trials all adenovirus isolates were detected when 48-h tissue cultures were examined and no false positives were observed. Only a few clinical samples have been looked at directly but results suggest that the reagent will perform well when reasonable material is available. However, more direct clinical material does need to be looked at. The ability of the reagent to detect adenovirus antigen only 4 h after infection of cell cultures suggests that it is sensitive enough to detect very small amounts of antigen. It therefore appears that the reagent produced will be a useful diagnostic tool for rapid detection of adenovirus antigen by IFA. References 1. 2. 3. 3 . 5 . Kohler, G.. and Milstein, C.. Nature, London, 1975. 256. 495. Rusdl, W. C., Patel, G.. Previous, B.. Sharp. I.. and Gardner. P. S . , J. Gen. Virol., 1981. 56, 393. Russell, W. C.. Hayashi, K., Sanderson, P. J . . and Pereira, H. G.. J. Gen. Virol. 1967, 1, 395. Hudson, L., and Hay, F. C., "Practical Immunology." Black- well Scientific, Oxford, 1980, pp. 11-12. Ross. S . R., Flint, S. J . , and Levine. A. J., Virology. 1980, 100, 319. I k m volume 10 onwards this series is split into two vnrts: Part A covers organic and Senior Reporter: bio-orgarhc E3.S.R. and Part I3 c';vcrs inorganic and bio-inorganic E.S.R.. Parts A and I3 will be published in altcmatc ycars. Volume 1 1A covers the literature published up to mid 1987. M. C. R. Symons, University of Leicester Brief Contents: Organic 1iadic:als in Solution; 'I'hcoretical Aspects of E.S.H.; Spin Labels: Uiological Membranes; Free Radical Studies in 13iolog-y and Medicine: E.S.K. of the Conformation of 5 and 6 Membered Cyclic Nitroxide (Aminoxyl) Specialist Periodical Report Hardcover 209pp ISBN 0 85186 861 4 Price G69.50 ($139.00) RSC Members Price 3237.50 Radicals. ROYAL SOCIETY OF C H EM I STRY Inforinat ion Swvic F, Ordering: RSC Members should send their orders to: Membership Manager, The Royal Society of Chemistry, 30 Russell Square, London WC1 B 5D'I', UK. Non-RSC Members should send their orders to: The Royal Society of Chemistry, Uistribution Centre, Blackhorse Road, Letchworth, Herts SG6 l H N , UK.

ISSN:0144-557X    

DOI:10.1039/AP9882500127

出版商:RSC    

年代:1988

数据来源: RSC

摘要:

132 ANALYTICAL PROCEEDINGS, APRIL 1988, VOL 25 Equipment News Probe for Fast Atom Bombardment Mass Spectrometry A new dual target probe has been deve- loped to overcome the difficulties of adjusting the optimum concentration ratio between a sample and the internal reference sample; the problem arises because the ionisation efficiency of the sample cannot be known in advance, and the adjustment has to be made by trial and error. The new probe tip has two faces loaded with two different samples, which can be measured by rotating the probe 180". One face is loaded with an unknown sample, the other with a reference sample, and the target face is changed over alternately at each scan. The probe can measure inorganic salts, such as cae- sium iodide, as the reference sample, and it permits mass measurement in a high mass range above miz 1500.The new probe is described in two application papers. Work detailed includes measure- ments on compounds with relative mole- cular masses of 362.2-1857.9; these include angiotensin-1 and gamma-endor- phin, hydrocortisone. erythromycin, alpha-cyclodextrin, bradykinine, bacit- racin A and alpha-MSH. Jeol Ltd., Jeol House, Grove Park, Colindale. London NW9 OJN. X-ray Diffraction Systems Two new on-line systems can analyse crystallographic structure, texture and stress simultaneously. The new diffrac- tometer can read, measure and analyse all angles of diffraction simultaneously by means of a dual, wide angle detector integrated to a high power collimated X-ray source. Applications include qual- ity control of minerals, painting coatings, strip aluminium metals and all general stress analysis.Spectrolab Ltd., P.O. Box 25. New- bury, Berkshire RG16 8BQ. Nuclear Magnetic Resonance Spectrometer The high sensitivity NMR probes intro- duced with the new GSX NMR spec- trometer include a proton probe with a signal to noise ratio of 250: 1. Other features of the GSX include an acquisition processor, which can handle 5 Mwords of 32-bit memory. Matrix shims are fitted as standard, enabling the operator to com- pute and correct fifth and sixth order gradients on installation. using a shim robot system. The system's software offers open or restricted access and allows the operator to control the autosampler and reduces operations such as shimming, phasing and integration to simple one- word commands.The extended range of around fifty probes includes single nucleus, those for dual C-H or dual F-H operation, and those tunable over a range of nuclei; special probes for research applications can also be provided. Jeol Ltd., Jeol House, Grove Park, Colindale, London NW9 OJN. Infrared Spectrophotometer The 1600 Series is a stand-alone, bench- top, Fourier transform instrument which integrates a sealed and desiccated optical system, advanced electronics and power- ful software with interactive graphics and multi-tasking capabilities. It features single-key operation and purge-free start- up. Excellent signal to noise and rapid data acquisition are achieved over the standard instrument range of 7800 to 350 cm-I. The analyst can display up to three spectra simultaneously, pan or zoom in on spectra, expand the ordinate and and abscissa and re-scale or re-range displays.The operator can also scan, manipulate and plot or print data simultaneously. Perkin-Elmer Ltd., Post Office Lane, Beaconsfield, Buckinghamshire HP9 1QA. Software for Ultraviolet - Visible Spectrophotometers A new range of plug-in software is announced for the PU8700 series of instruments. A Multilambda package provides a whole suite of two and three wavelength calculations for sample analy- sis. When more than three wavelengths are needed for sample measurement the operator simply keys in up to eight selected wavelengths and all results are displayed in a data table. For mixture analysis with known components the Mul- ticomponent Analysis software will auto- matically calculate the concentration of each constituent.The PUS700 kinetics software has all the facilities for enzyme characterisation. When analyses require post-run manual recalculation for more specialised techniques the new User Pro- gramming package will provide the answer. The software, provided as a disk for the Makers' analytical data station, enables users to set up a sequence of methods, collect sample data and auto- matically load them into LOTUS 1-2-3. run the spreadsheet macro-calculations and print out the results. In addition to the software packages the makers have intro- duced new hardware, sample handling and result manipulation accessories for the PU8700. Pye Unicam Ltd., York Street, Cam- bridge CB1 2PX. Gas Chromatograph The Shimadzu GC-14A Series is designed for capillary gas chromatography but is also compatible with packed columns.It employs a unique temperature control circuit, which ensures accurate main- tenance of oven temperature. This, coupled with exceptional uniformity of temperature distribution within the oven. delivers the reproducibility of retention times required for capillary GC. Up to four different sample injectors can be fitted; each can be independently temper- ature controlled. The GC-14A can be connected to a Shimadzu Chromatopac data processor via a current loop inter- face, or a PC can be used via the RS 232C interface. If required, five-step, multi- linear temperature programming is pos- sible. Dyson Instruments Ltd., Hetton Lyons Industrial Estate, Hetton, Houghton-le- Spring, DH5 ORH.Gas Chromatograph The Model 8700 offers simultaneous dual- channel operation with a full colour VDU. It features powerful integral data processing, multi-level calibration and multiple reference peak facilities. It also offers dual channel real time or post run graphics, split screen or overlaid, simple graphics manipulation for comparison of sample and reference chromatograms, enhanced software for independent inte- gration of both channels and computer communications for transfer of GC methods, raw and re-integrated data and reports. Fully automated, analytical sequencing software is included as stan- dard for routine applications. Perkin-Elmer Ltd., Post Office Lane, Beaconsfield, Buckinghamshire HP9 1QA. Gas Chromatograph The Model 92 has been designed with special attention to the software con- trolled ovens.The column oven is designed to heat rapidly and to maintain minimal thermal gradients. Rapid cooling is also a feature of the column oven: for example, 250-50°C can be achieved in only 7 min. A minimum column oven temperature of -99°C can be achieved with the incorporation of a sub-ambient facility. Temperature programming with up to four ramps is a standard feature and temperature program profiles are extremely reproducible. The standard injection oven can accommodate up to three injectors and enables a combination of packed and capillary columns to be fitted simultaneously. A second.ANALYTICAL PROCEEDINGS, APRIL 1988, VOL 25 optional, separately controlled injector oven can be fitted.A cold on-column capillary injection system is also avail- able. The Model 92 is fitted as standard with a detector oven into which two ionisation detectors can be mounted. Analytical Measuring Systems, Shire- hill Industrial Estate, Shirehill, Saffron Walden, Essex CBll 3AQ. Preparative HPLC System The Shimadzu LC-8A system is computer controlled and offers a wide flow-rate control range, so that both analytical and preparative HPLC can be performed. sis columns. The Aminex HPX-87N is tailored for sugar analysis in samples with high salt concentrations, such as mol- asses. The Aminex HPX-87K provides improved separation of glucose (Dp 1). maltose (Dp 2) and maltotriose (Dp 3); these carbohydrates are resolved in less than 11 min in samples such as corn syrup and brewing wort.When glucose is the only compound of interest in the sample the Fast Glucose Analysis Column pro- vides sensitive analysis in less than 2 min. Bio-Rad Laboratories Ltd., Caxton Way, Watford Business Park, Watford, Hertfordshire WD1 8RP. Shimadzu LC-8A preparative HPLC system Thus, operational conditions can be accu- rately defined on an analytical column before preparative work commences. The mobile phase flow-rate is adjustable over the range 0.1-150 ml min-1. There is a choice of manual, pumped or automatic sample injection. The flow of the mobile phase can be programmed. The LC-8A system is modular in concept. The com- ponents include a liquid pump, a system controller, ultraviolet and ultraviolet - visible detectors, sample injectors, col- umn holders and valves, a fraction collec- tor and a range of interfaces.When thz LC-8A is linked to a C-R4A Chromato- pac data processor, on-screen processing of chromatograms is possible. Dyson Instruments Ltd., Hetton Lyons Industrial Estate, Hetton, Houghton-le- Spring DH5 ORH. HPLC Column Packings Hypersil spherical silica based materials have narrow particle size distribution and controlled porosity. They are available in bulk in 3, 5 and 10 pm sizes and in pre-packed columns with inverse com- pression fittings in sizes from 50 mm x 4.6 mm up to 250 mm x 7 mm. The latest addition to the Hypersil range is the WP300 series of wide pore materials, especially engineered for separation of biopolymers. This series is available as silica or bonded reverse phase in 5 and 10 pm sizes and in pre-packed columns. Also available is a column packing pump.Shandon Southern Products Ltd., Chadwick Road, Astmoor, Runcorn, Cheshire WA7 1PR. HPLC Columns Three new columns have been added to the Aminex range of carbohydrate analy- HPLC Cells The makers: range of HPLC cells has been extended with the addition of two new units. The high efficiency activated glass cell is packed with high quality glass beads and is capable of achieving 70% efficiency, compared with the 35% or 40% typical of conventional cells. The second new cell is a contamination-free version consisting of a plastic scintillator covered by a very thin PTFE membrane; it is intended for use in checking the purity of radiochemical compounds. Also avail- able are yttrium silicate.glass, liquid mixing, Cerenkov, sodium iodide crystal and X cells. Raytest Instruments Ltd., St. John’s House, 131 Psalter Lane, Sheffield s11 8UX. Fluorescence Detectors A range of three instruments, all featuring a 10 W xenon flash source, are designed for the fast and convenient interchange of flow cells and cuvettes for microbore, analytical and preparative HPLC applica- tions. Measurements of non-flowing samples, such as those taken from fraction collectors, can be taken with a cuvette accessory. The FD-100 is an advanced filter fluorimeter with a wide selection of excitation and emission filters. The FD- 200 features the combination of a high efficiency monochromator for excitation and emission filters common to the FD- 100.The top of the range FD-300 is a dual monochromator instrument with continu- ously variable excitation and emission wavelengths. It is ideal for applications where excitation and emission wavelengths are closely separated. Severn Analytical Ltd.. Unit 2B. St. 133 Francis’ Way, Shefford Industrial Estate, Shefford. Bedfordshire SG17 5DZ. Micro-syringes for Liquid and Gas Chromatography The Exmire range of micro-syringes, made in Japan, for liquid analysis have a pressure resistance of 3.0 kg cm-2 and a superior airtight seal. Calibration is to an accuracy of k 1 O%. Standard models include two micro-syringes, with capaci- ties of 1 and 5 pl, and fixed and inter- changeable needle options with capacities ranging from 5 to 500 pl. Six options are available for gas analysis with capacities from 0.25 to 10 pl.A resistance of 5.0 kg cm-2 prevents gas leakage, and accu- racy is within 22%. A comprehensive range of needles is also available. A catalogue describing the Exmire range is obtainable. Dyson Instruments Ltd., Hetton Lyons Industrial Estate. Hetton, Houghton-le- Spring DH5 ORH. Camera Systems for Electrophoresis The new range of Polaroid camera systems has been selected by Ultra-Violet Products Ltd. for use with their electro- phoresis systems. A choice is offered between the TA40 model and the TA50, which incorporates automatic processing using the new type 331 large format autofilm. The original CU5 is still avail- able. The TA50 has an easy-loading system which removes the danger of accidental film exposure; it also produces slightly larger prints than the CU5.Ultra-Violet Products Ltd., Science Park, Milton Road, Cambridge CB4 4FH. Peptide - Protein Microsequencer The CI 4000. developed by the makers working in conjunction with scientists at Imperial College, London, is economical and offers a flexible approach to protein sequencing. It can be operated either in the gas phase or solid phase to cleave amino acids from the n-terminal end of a protein, making them available for iden- tification by HPLC. The instrument is fully software driven and both large proteins and small peptides can be sequenced with only minor changes in the standard programs supplied. Chelsea Instruments. Biotechnology134 Division, Unit 9, Avon Business Centre, Avonmore Road, London W14 8TS.Electrode The new 1400 Series combined pH/redox electrodes have a robust PVC 'h-in BSP threaded body which houses a platinum and a pH electrode sharing a silver chloride reference electrode, all con- tained in a single complete assembly. The electrode is particularly suitable for the monitoring and control of swimming pool additive levels. It operates over the range 0-14 pH and has an operating tempera- ture range of 0-100°C. Kent Industrial Measurements Ltd., Oldends Lane, Stonehouse, Gloucester- shire GLlO 3TA. pH Indicator Sticks Specially designed for use in weak solu- tions, the three zone and single zone sticks are part of the Dosatest range of pH indicators. The entire pH range is covered by the three zone sticks; one stick covers pH 0.0-6.0, another pH 4.5-10.0 and the third pH 7.0-14.0. A simple comparison of the colour for each zone with that of the reference scale gives the pH of a solution to within 0.5 pH units.For very high definition pH measurements the range of one zone plastic sticks has been designed; they cover a pH range of 0.3-9.5, the single indicator zone recording pH to within 0.2-0.5 units. May and Baker Ltd., Liverpool Road, Eccles, Manchester M30 7RT. Sodium/Potassium Analyser The Orion 1020 fully automated sodium/ potassium analyser produces a result in 60 s; this includes sample measurement, automatic flushing, calibration and needle self cleaning. The test is carried out on 100 1.11 of whole blood or urine. A built-in rechargeable battery makes the instrument ideal for mobile units. Vital Scientific Ltd., Huffwood Trad- ing Estate, Partridge Green, West Sussex RH13 8AU.ANALYTICAL PROCEEDINGS, APRIL 1988, VOL 25 Calibration Dew Point Meter The Series 3020, an enhanced version of the makers' Series 3000DTR, has as its primary role the calibration of other hygrometers and relative humidity instru- ments. Capable of simultaneously regi- stering dewpoint, relative humidity and gas temperature, the Series 3020 is especially suited to applications where a relative humidity readout is required and where the standard RH probe is either too inaccurate or has unacceptable drift. An important feature of the new instrument is its automatic balance compensation cir- cuitry, which periodically and automatic- ally purges the optical cooled-mirror dew point system to obtain a new zero before returning to normal operation.Accuracy is stated as +0.27"C, and resolution 0.1 "C. Michell Instruments Ltd., Unit 9, Nuf- field Close, Nuffield Road, Cambridge CB4 1SS. Moisture Analyser The Computrac TMX provides a safe, simple and fast alternative to Karl Fischer titration or vacuum oven techniques. It requires no glassware or toxic reagents. Because of the large sample capacity (up to 40 g) the instrument provides more representative results than the Karl Fischer technique. Also, more accurate and precise results can be obtained on samples with very low moisture content; this .is because of the shorter sample preparation time. Moisture is extracted from the sample by heating it at a con- stant, programmable temperature. Evol- ved volatiles are driven into an analysis cell, which selectively traps the moisture.The instrument measures the change in weight of the cell as the moisture is collected. Resolution on the display is 0.001% and the instrument will provide moisture specific results down to 0.01%. Arizona Instrument Corporation, P.O. Box 670, Cookham, Maidenhead, Berk- shire SL6 9BJ. Leak Detector Leakseeker 96 is a compact, hand-held instrument, which uses a micro-volume, thermal conductivity detector to locate and identify minute gas leaks in the laboratory. Pulling a trigger initiates a self-check routine, followed by sampling and readout of results. The entire opera- tion takes less than 1 s and any gas with a thermal conductivity different to the ambient surroundings can be detected.Analytical Instruments Ltd., London Road, Pampisford, Cambridge CB2 4EF. Particle Size Analyser The Nicomp 370 sub-micron particle size analyser handles particle sizing from 0.005-3 prn by dynamic light scattering of a laser beam incident on particles suspen- ded in solution. The instrument features a flow-through cell with automatic sample handling capabilities. This allows auto- matic dilution of concentrated samples under computer control resulting in con- stant optimum average scattering inten- sity. A full analysis with reproducible results can be completed well within 20 min. Pacific Scientific Instrument Division, 4 First Avenue, Globe Park, Marlow SL7 1YA. Software for Thermal Analysis Three packages are announced for use with the makers' Delta thermal analysis system: DSC Purity, which allows the purity of most crystalline organics to be measured without the need for a compa- rative standard; DSC Kinetics, to allow those working with thermosetting poly- mers, such as epoxies, to optimise the cure cycle for products or processes; and DSC Partial Areas, which allows over- lapping DSC peaks to be quantified and permits the solid-fat index of fats or waxes to be calculated.Perkin-Elmer Ltd., Post Office Lane, Beaconsfield, Buckinghamshire HP9 1QA. Sampler Scheduler Software This package is an enhancement package for the makers' laboratory information management system, LIMS 2000. It features a spreadsheet format diary, which allows a laboratory to specify, up to a year in advance, when a sample should be taken from which sampling point and which tests are to be performed at the sampling point or in the laboratory.Also featured are collection lists, indicating when and where samples are due for collection, and sample receipt, an auto- scheduling feature which works overnight to search the computer database for all samples expected the next day. Perkin-Elmer Ltd., Post Office Lane, Beaconsfield, Buckinghamshire HP9 1QA. Quality Control Software QCLIMS revision 1.1 is an enhanced version of the QCLIMS system aimed specifically to assist in maintaining pro- duct quality in the pharmaceutical, cos- metic and food and drink industries. The revision contains substantially increased functionality and has been engineered in line with Good Laboratory Practices. Perkin-Elmer Ltd., Post Office Lane, Beaconsfield, Buckinghamshire HP9 1QA.Data Analyser The Graphtec MS5100 is a four channel data analyser, which provides the func- tions of six instruments in one unit while maintaining simple operation by using aninteractive menu display. The six measurement modes are Y - T direct measurement, X - Y direct measurement, direct logging measurement, Y - T memory measurement, X - Y memory measurement and memory event measurement. Environmental Equipments (North- ern) Ltd., Environ House, Welshman’s Lane, Off Welsh Row, Nantwich, Cheshire CWS 6AB. Particle Size Analyser The SediGraph 5100 is a fully automated sedimentation particle size analyser pro- viding size analysis in the range 300-0.1 pm. Up to 6000 sample analyses can be and the measured mass appears instantly on the digital display.Mettler Instrumente AG, CH-8606 Greifensee, Switzerland. Arc Lamps The Cermax range of compact source arc lamps has been extended. The lowest power now available is 75 W and the highest has been increased to 1 kW. Power supplies, lamp holders and acces- sories are available for all sizes. The lamps are small in size. The 300 W lamp is just over 1 in in both diameter and length. In this compact package there is a para- boloidal mirror to generate a highly colli- ANALYTICAL PROCEEDINGS, APRIL 1988, VOL 25 Microrneritics SediGraph 5100 maintained on-disk and over eleven dif- ferent types of graphical reports can be produced in either dot matrix or presenta- tion quality pen-plot formats. Base-line correction and sample cell bubble elimi- nation are automated, as is overlaying up to three plots on to an existing plot.Micromeritics, One Micromeritics Drive, Norcross, GA 30093-1877, USA. Automatic Multi-Tip Pipette The MP20 pipetting instrument is inten- ded for the Biomek 1000 automated workstation. If offers a volume range of 1-20 pm and is equipped with eight nozzles, each using disposable plastic tips to eliminate any possibility of carry-over. Beckman Ltd., Progress Road, Sands Industrial Estate, High Wycombe, Buck- inghamshire. Electronic Scales The Mettler J series of electronic preci- sion scales covers weighing ranges from 0.1 mg to 15000 g. Three models, the PJ360, the PJ3600 and the PJ1220 have the DeltaRange facility, a fine range of ten times the accuracy that can be called up anywhere throughout the weighing range.All scales of the J series can be fitted with an optional data output. The scale is operated with a single control bar mated beam of light which extends from the ultraviolet to the infrared. Laser Lines Ltd., Banbury, Oxford- shire OX16 7TQ. End Adapters for Filters End adapters for Hytrex I1 depth cart- ridge filters are available to fit any com- mercial housing. They include elas- tomeric gasket adapters and all-poly- propylene adapters, including moulded adapters with “222” O-rings, moulded adapters with bayonet-type lock, moul- ded fins, integral self-seal springs, exten- ded core adapters and solid end caps. Osmonics Inc., 5951 Clearwater Drive, Minnetonka, Minnesota 55343, USA. Automatic Filter - Dryer The 75-60 air dryer is designed for use with sensitive high resolution analytical instruments, such as Fourier transform infrared spectrometers, which require gas purge and/or a continuous clean, dry air supply.The 75-60 is capable of removing carbon dioxide to less than 2 p.p.m. over a wide range of inlet concentrations at a pressure dew point of -70 “C. Typically a maximum inlet air pressure of 8.5 bar will produce a maximum dry air flow-rate of 135 6.1 N m3 h-1; the pressure drop at maximum flow-rate is less than 0.5 bar. Balston Ltd., Monckton’s Lane, Maid- stone, Kent ME14 2QB. Clean Room Oven The RCOS-2220 is a clean room oven with high particle-infiltration protection. It provides particle protection to Class 100 level. Four standard sizes are available, each with two temperature ranges.The four oven capacities are 3.2,7.6, 16.2 and 30.5 ft3. Temperature ranges are from +70 to +200°C and +70 to +350°C. Dage (GB) Ltd., Intersem Division, Rabans Lane, Aylesbury, Buckingham- shire HP19 3RG. Molecular Modelling The P-GRAF 1 family of molecular modelling workstations is based on a VAXstation 2000 CPU with 4 MB memory and a 71 or 159 MB disk. The complete system includes a colour moni- tor, tailored VMS environment and the basic set of Chem-X modules. P-GRAF 5 systems, built round a MicroVAX 2000, are available in a variety of configurations and may be incorporated into local area clusters. The integrated high performance raster graphics display is based on a 68020 processor, has a peak performance of 150000 vectors s-1, a pixel resolution of 1024 x 724, and can display 256 simul- taneous colours from a palette of over 16 million.The makers’ Chem-X molecular modelling software can now be controlled entirely by picking items (not commands) from a carefully constructed series of menus using a data tablet or mouse. The new menu-driven interface is combined with ChemGuide, a knowledge based Chem-X module intended to lead novice users towards specified modelling goals. Experienced users may choose not to invoke the menu system; it is still possible to enter commands at the keyboard if required. Chemical Design Ltd., Unit 12, 7 West Way, Oxford OX2 OJB Literature A brochure introduces the new Minibore columns for gas chromatography and GC - MS. Entitled “High Speed Mini- bore Columns,” it offers an 8-page review of new short, small diameter (0.18 mm i.d.) columns specially designed to dec- rease analysis time. Seven chromato- grams are provided to illustrate typical applications, including analyses of naph- tha, cassia oil, pesticides, barbiturates, amphetamines, EPA 625 priority pollu- tants and basic drugs. J and W Scientific, 91 Blue Ravine Road, Folsom, CA 95630, USA. A range of liquid chromatography sup- port literature covers methods develop- ment and the makers’ latest advances in instrumentation. Topics covered include136 ANALYTICAL PROCEEDINGS, APRIL 1988, VOL 25 the analysis of plasma fatty acids, fast LC analysis of the individual bitter com- ponents in hops and the determination of food additives in fruit drinks. Other pap- ers deal with an automated procedure for the ruggedness testing of chromato- graphic conditions in HPLC, the flexibil- ity and capability of the PU4110 ultra- violet - visible detector, and the twin- Company’s range of scintillation cocktails piston pump solvent delivery systems and it includes details of plastic vials. featured in the PU4100 liquid chromato- There is also mention of on-line radioiso- graphs. tope monitoring using the Company’s Pye Unicam Ltd., York Street, Cam- radiochromatography detector system. bridge CBl 2PX. Beckman Ltd., Progress Road, Sands Industrial Estate, High Wycombe, Buck- A brochure gives information on the inghamshire.

ISSN:0144-557X    

DOI:10.1039/AP9882500132

出版商:RSC    

年代:1988

数据来源: RSC