BS 4585-18-1996 Methods of test for spices and condiments - Determination of total capsaicinoid content of chillies and chilli oleoresins (high-performance liquid chromatographic m.pdf

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1、BRITISH STANDARD BS 4585-18: 1996 Methods of test for Spices and condiments Part 18: Determination of total capsaicinoid content of chillies and chilli oleoresins (high-performance liquid chromatographic method)BS4585-18:1996 This British Standard, having been prepared under the directionof the Cons

2、umer Products and Services Sector Board, was published under the authority of the Standards Board and comes into effect on 15February1996 BSI08-1999 The following BSI references relate to the work on this standard: Committee reference AW/7 Draft for comment 95/500122 DC ISBN 0 580 25186 1 Committees

3、 responsible for this British Standard The preparation of this British Standard was entrusted to Technical Committee AW/7, Spices and condiments, upon which the following bodies were represented: Association of Public Analysts Consumer Policy Committee of BSI Department of Trade and Industry (Labora

4、tory of the Government Chemist) International General Produce Association Leatherhead Food Research Association Ministry of Agriculture, Fisheries and Food Natural Resources Institute Seasoning and Spice Association Amendments issued since publication Amd. No. Date CommentsBS4585-18:1996 BSI 08-1999

5、 i Contents Page Committees responsible Inside front cover Foreword ii 1 Scope 1 2 References 1 3 Principle 1 4 Reagents 1 5 Apparatus 1 6 Sampling 1 7 Preparation of test sample 1 8 Procedure 2 9 Expression of results 2 10 Test report 2 List of references Inside back coverBS4585-18:1996 ii BSI 08-1

6、999 Foreword This Part of BS 4585 has been prepared by Technical Committee AW/7. The International Standard ISO7543-2:1993 was taken into account during the preparation of this Part of BS4585. However, it was decided that the method described in ISO7543-2, which involves the use of tetrahydrofuran,

7、should not be adopted as a British Standard. This British Standard calls for the use of substances that may be injurious to health if adequate precautions are not taken. It refers only to technical suitability and does not absolve the user from legal obligations relating to health and safety at any

8、stage. A British Standard does not purport to include all the necessary provisions of a contract. Users of British Standards are responsible for their correct application. Compliance with a British Standard does not of itself confer immunity from legal obligations. Summary of pages This document com

9、prises a front cover, an inside front cover, pagesi andii, pages1and2, an inside back cover and a back cover. This standard has been updated (see copyright date) and may have had amendments incorporated. This will be indicated in the amendment table on the inside front cover.BS4585-18:1996 BSI 08-19

10、99 1 1 Scope This Part of BS4585 describes a method for the determination, by high-performance liquid chromatography (HPLC), of the total capsaicinoid content of whole and powdered chillies and their oleoresins. 2 References 2.1 Normative references This Part of BS4585 incorporates, by dated or unda

11、ted reference, provisions from other publications. These normative references are made at the appropriate places in the text and the cited publications are listed on the inside back cover. For dated references, only the edition cited applies; any subsequent amendments to or revisions of the cited pu

12、blication apply to this Part of BS4585 only when incorporated in the reference by amendment or revision. For undated references, the latest edition of the cited publication applies, together with any amendments. 2.2 Informative references This Part of BS 4585 refers to other publications that provid

13、e information or guidance. Editions of these publications current at the time of issue of this standard are listed on the inside back cover, but reference should be made to the latest editions. 3 Principle Extraction of the capsaicinoids with methanol and determination by HPLC at a wavelength of280n

14、m. 4 Reagents 4.1 General. Unless otherwise stated, use reagents of recognized analytical grade and water conforming to grade3 of BSENISO3696. NOTEReference should be made to the safety data sheets available with each reagent. 4.2 Methanol, HPLC grade. 4.3 Standard solution of capsaicinoids, prepare

15、d as follows. Weigh accurately 0.100g of capsaicin (8-methyl-n-vanillyl-6-nonenamide) (98% (m/m) crystalline) into a100ml one-mark volumetric flask containing about50ml of methanol. Stopper the flask, shake to dissolve, dilute to the mark with the methanol (4.2) and mix. WARNING. Capsaicin should be

16、 handled with extreme care as it is toxic by inhalation, in contact with skin and if swallowed. It causes severe irritation. 4.4 Working standard solution, prepared as follows. Transfer by safety pipette5ml of the standard solution (4.3) to a50ml one-mark volumetric flask. Dilute to the mark with th

17、e methanol (4.2) and mix. 4.5 Elution solvent, comprising0.5% (V/V) glacial acetic acid,49.5% (V/V) water, and50.0% (V/V) acetonitrile, prepared as follows. Add 10 ml of glacial acetic acid to990ml of water. Add1000ml of acetonitrile. De-gas before use. 4.6 Acid-washed sand 5 Apparatus 5.1 Usual lab

18、oratory apparatus 5.2 Continuous extraction apparatus (Soxhlet type), fitted with a100ml flask. 5.3 Extraction thimbles 5.4 Membrane filters, with a pore size of0.024m. 5.5 Disposable syringe, of1ml capacity. 5.6 High-performance liquid chromatograph, equipped with a detector allowing measurements t

19、o be made at a wavelength of280nm, fitted to a recorder or integrator. 5.7 Chromatographic column for HPLC, of stainless steel,25cm long and with an inside diameter of0.4cm to0.5cm. Stationary phase: spherical silica with octadecyl functional group C 18and particle size of54m maximum. 5.8 Injection

20、system, comprising an injection valve with a204l loop, or other system giving the same injection accuracy. 5.9 Test sieve, with an aperture size of5004m, conforming to BS410. 5.10 Filter paper, Whatman no.541 or similar. 6 Sampling Sampling shall have been carried out in accordance with BS4540-1. 7

21、Preparation of test sample 7.1 Chillies in powder form 7.1.1 Check that the laboratory sample passes through the test sieve (5.9). If this is not the case, grind the powder in accordance with BS4540-2 until the required particle size is obtained. NOTESufficient material should be prepared, e.g. a to

22、tal of50g for a moisture determination.BS4585-18:1996 2 BSI 08-1999 7.1.2 From the test sample prepared in accordance with7.1.1, weigh, into the extraction thimble (5.3), to the nearest0.001g, between1g and5g, depending on the expected capsaicin content of the sample (record the mass), and mix with

23、an equal volume of the acid-washed sand (4.6). Extract for6h to8h in the continuous extraction apparatus(5.2), using80ml of the methanol (4.2). Cool the extract and filter it through a filter paper(5.10) into a100ml one-mark volumetric flask, washing the flask and the filter with the methanol. Dilut

24、e to the mark with the methanol and mix. 7.2 Whole chillies Grind the laboratory sample in accordance with BS4540-2 and proceed as described in7.1.2. 7.3 Oleoresins of chillies Thoroughly homogenize the laboratory sample. Weigh, to the nearest0.001g,1g of the oleoresin into a100ml one-mark volumetri

25、c flask. Dilute to the mark with the methanol (4.2) and mix. 8 Procedure 8.1 Chilli extract and oleoresin solution Using the disposable syringe (5.5) and a membrane filter (5.4), transfer1ml to2ml of the solution prepared in accordance with7.1.2 or7.3 to a small tube. Stopper the tube to prevent eva

26、poration of the solvent. The test solution shall be clear. If this is not the case, refilter it through a membrane filter (5.4). 8.2 Determination 8.2.1 Set up the chromatograph (5.6) and the column (5.7) in accordance with the manufacturers instructions. Using the injection system (5.8), inject254l

27、 of the working standard solution (4.4) into the column. Commercial capsaicin yields four peaks: nordihydrocapsaicin (C 1 ), capsaicin (C 2 ), dihydrocapsaicin (C 3 ) and homodihydrocapsaicin (C 4 ). Record the results (peak areas). Inject a further254l of the working standard solution (4.4). Record

28、 these results. The second set of results should not deviate from the first by more than5%. 8.2.2 Repeat the operations given in8.2.1 using the methanolic extract of chilli. If the peak areas of the first set of these results deviate by more than5% from those of the second set of results, repeat the

29、 determination. Use the mean of these results for the calculation in clause9. NOTEThe determination should be repeated until the results do not deviate by more than5%. 8.2.3 Determine the moisture content, expressed as a percentage by mass of the test sample, in accordance with BS4585-2. 9 Expressio

30、n of results The total capsaicinoid content, expressed as a percentage by mass of the dry matter, is given by the following formula: where C 1 , C 2 , C 3 , and C 4are the peak areas of nordihydrocapsaicin, capsaicin, dihydrocapsaicin and homodihydrocapsaicin of the test sample solution. c is the co

31、ncentration of the standard solution(4.3) (ing/100ml); S 1 , S 2 , S 3 , and S 4are the peak areas of nordihydrocapsaicin, capsaicin, dihydrocapsaicin and homodihydrocapsaicin of the standard solution (4.4); c 2is the concentration of the test sample solution (ing/100ml) (see7.1.2); H is the moistur

32、e content, expressed as a percentage by mass of the test sample, determined in accordance with BS4585-2. To convert percentage capsaicinoids to Scoville units, use the following formula: 10 Test report The test report shall include the following information: a) a complete identification of the sampl

33、e; b) a reference to this British Standard, i.e.BS4585-18:1996; c) the total capsaicinoid content, expressed as a percentage by mass of the dry matter and/or in Scoville units; d) details of any incidents during the determination which may have influenced the results; e) any operations not included

34、in this British Standard, and any operations regarded as optional. Scoville units % capsaicinoids 15000000 100 - =BS4585-18:1996 BSI 08-1999 List of references (see clause 2) Normative references BSI publications BRITISH STANDARDS INSTITUTION, London BS 410:1986, Specification for test sieves. BS EN

35、 ISO 3696:1995, Water for analytical laboratory use. BS 4540, Specification and test methods. Sampling of spices and condiments. BS 4540-1:1981, Methods of sampling. BS 4540-2:1982, Method for preparation of a ground sample for analysis. BS 4585, Methods of test for spices and condiments. BS 4585-2:

36、1982, Determination of moisture content (entrainment method). Informative references ISO publication INTERNATIONAL ORGANIZATION FOR STANDARDIZATION (ISO), Geneva. (Publication is available from Customer Services, BSI.) ISO 7543-2:1993, Chillies and chilli oleoresins Determination of capsaicinoid con

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