BS EN 12136-1998 Fruit and vegetable juices - Determination of total carotenoid content and individual carotenoid fractions《水果汁和蔬菜汁 总类胡萝卜素含量及各分类胡萝卜素比例的测定 》.pdf

《BS EN 12136-1998 Fruit and vegetable juices - Determination of total carotenoid content and individual carotenoid fractions《水果汁和蔬菜汁 总类胡萝卜素含量及各分类胡萝卜素比例的测定 》.pdf》由会员分享，可在线阅读，更多相关《BS EN 12136-1998 Fruit and vegetable juices - Determination of total carotenoid content and individual carotenoid fractions《水果汁和蔬菜汁 总类胡萝卜素含量及各分类胡萝卜素比例的测定 》.pdf（12页珍藏版）》请在麦多课文档分享上搜索。

1、| | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | BRITISH STANDARD BS EN 12136 : 1998 The Eu

2、ropean Standard EN 12136 : 1997 has the status of a British Standard ICS 67.160.20 NO COPYING WITHOUT BSI PERMISSION EXCEPT AS PERMITTED BY COPYRIGHT LAW Fruit and vegetable juices Determination of total carotenoid content and individual carotenoid fractionsThis British Standard, having been prepare

3、d under the direction of the Consumer Products and Services Sector Board, was published under the authority of the Standards Board and comes into effect on 15 February 1998 BSI 1998 ISBN 0 580 28957 5 BS EN 12136 : 1998 Amendments issued since publication Amd. No. Date Text affected National forewor

4、d This British Standard is the English language version of EN 12136 : 1997. The UK participation in its preparation was entrusted to Technical Committee AW/21, Fruit and vegetable juices, which has the responsibility to: aid enquirers to understand the text; present to the responsible European commi

5、ttee any enquiries on the interpretation, or proposals for change, and keep the UK interests informed; monitor related international and European developments and promulgate them in the UK. A list of organizations represented on this committee can be obtained on request to its secretary. ISO 5725 :

6、1986, to which informative reference is made in the text, has been superseded by ISO 5725-1 : 1994, ISO 5725-2 : 1994, ISO 5725-3 : 1994, ISO 5725-4 : 1994 and ISO 5725-6 : 1994 which are identical with BS ISO 5725 Accuracy (trueness and precision) of measurement methods and results, BS ISO 5725-1 :

7、 1994 General principles and definitions, BS ISO 5725-2 : 1994 Basic method for the determination of repeatability and reproducibility of a standard measurement method, BS ISO 5725-3 : 1994 Intermediate measures of the precision of a standard measurement method, BS ISO 5725-4 : 1994 Basic method for

8、 the determination of the trueness of a standard measurement method, and BS ISO 5725-6 : 1994 Use in practice of accuracy values. Cross-references The British Standards which implement international or European publications referred to in this document may be found in the BSI Standards Catalogue und

9、er the section entitled International Standards Correspondence Index, or by using the Find facility of the BSI Standards Electronic Catalogue. Compliance with a British Standard does not of itself confer immunity from legal obligations. Summary of pages This document comprises a front cover, an insi

10、de front cover, the EN title page, pages 2 to 10, an inside back cover and a back cover.CEN European Committee for Standardization Comite Europe en de Normalisation Europa isches Komitee fu r Normung Central Secretariat: rue de Stassart 36, B-1050 Brussels 1997 CEN All rights of exploitation in any

11、form and by any means reserved worldwide for CEN national Members. Ref. No. EN 12136 : 1997 E EUROPEAN STANDARD EN 12136 NORME EUROPE ENNE EUROPA ISCHE NORM September 1997 ICS 67.160.20 Descriptors: Fruit and vegetable juices, chemical analysis, determination of content, carotenoids, procedure Engli

12、sh version Fruit and vegetable juices Determination of total carotenoid content and individual carotenoid fractions Jus de fruits et de le gumes De termination de la teneur en carote noides totaux et en fractions individuelles de carote noides Frucht- und Gemu sesa fte Bestimmung des Gesamtcarotinoi

13、dgehaltes sowie enzelner Carotinoid-Fraktionen This European Standard was approved by CEN on 6 September 1997. CEN members are bound to comply with the CEN/CENELEC Internal Regulations which stipulate the conditions for giving this European Standard the status of a national standard without any alte

14、ration. Up-to-date lists and bibliographical references concerning such national standards may be obtained on application to the Central Secretariat or to any CEN member. This European Standard exists in three official versions (English, French, German). A version in any other language made by trans

15、lation under the responsibility of a CEN member into its own language and notified to the Central Secretariat has the same status as the official versions. CEN members are the national standards bodies of Austria, Belgium, Czech Republic, Denmark, Finland, France, Germany, Greece, Iceland, Ireland,

16、Italy, Luxembourg, Netherlands, Norway, Portugal, Spain, Sweden, Switzerland and United Kingdom.Page 2 EN 12136 : 1997 BSI 1998 Foreword This European Standard has been prepared by Technical Committee CEN/TC 174, Fruit and vegetable juices Methods of analysis, the secretariat of which is held by AFN

17、OR. This European Standard shall be given the status of a national standard, either by publication of an identical text or by endorsement, at the latest by March 1998, and conflicting national standards shall be withdrawn at the latest by March 1998. According to the CEN/CENELEC Internal Regulations

18、, the national standards organizations of the following countries are bound to implement this European Standard: Austria, Belgium, Czech Republic, Denmark, Finland, France, Germany, Greece, Iceland, Ireland, Italy, Luxembourg, Netherlands, Norway, Portugal, Spain, Sweden, Switzerland and the United

19、Kingdom. Contents Page Foreword 2 1 Scope 3 2 Normative references 3 3 Definitions and symbols 3 4 Principle 3 5 Reagents 3 6 Apparatus 4 7 Procedure 4 8 Calculation 5 9 Precision 6 10 Test report 6 Annexes A (informative) Bibliography 7 B (informative) Statistical results of the inter-laboratory te

20、st 7Page 3 EN 12136 : 1997 BSI 1998 1 Scope This European Standard specifies a method for the determination of total carotenoid content and individual carotenoid fractions of fruit and vegetable juices and related products. 2 Normative references This European Standard incorporates by dated or undat

21、ed reference, provisions from other publications. These normative references are cited at the appropriate places in the text and the publications are listed hereafter. For dated references, subsequent amendments to or revisions of any of these publications apply to this European Standard only when i

22、ncorporated in it by amendment or revision. For undated references the latest edition of the publication referred to applies. EN ISO 3696 : 1995 Water for analytical laboratory use Specification and test methods (ISO 3696 : 1987) ISO 5725 : 1986 Precision of test methods Determination of repeatabili

23、ty and reproducibility for a standard test method by inter-laboratory tests. 3 Definitions and symbols 3.1 Definitions For the purposes of this standard, the following definitions apply. 3.1.1 carotenoid fraction I (mainly hydrocarbons) Material eluted with petroleum ether, as described in 7.3.2. 3.

24、1.2 carotenoid fraction II (crypotxanthin esters) Material eluted with elution mixture A (5.7), as described in 7.3.2. 3.1.3 carotenoid fraction III (xantophyll esters) Material eluted with elution mixture B (5.8), as described in 7.3.2. 3.1.4 carotenoid fraction IV Material eluted with acetone (5.3

25、), as described in 7.3.2. 3.2 Symbols For the purposes of this standard, the following symbol applies. g acceleration due to gravity at the surface of the earth (9,81 m/s 2 ). 4 Principle Quantitative adsorptive precipitation of carotenoids by zinc hexacyanoferrate (II) by treatment with Carrez I an

26、d Carrez II solutions. Extraction of carotenoids from the precipitate using acetone and transfer of the solution to petroleum ether. Determination of the total carotenoid content by spectrometry. Fractionation of the carotenoids using chromatography column on aluminium oxide. Individual determinatio

27、n of the single carotenoid fractions spectrometrically. Calculation of the total carotenoid content and the single fractions as b-carotene, expression of the content of the individual carotenoid fractions as a percentage of total carotenoid content. 5 Reagents Use only reagents of recognized analyti

28、cal grade and only water in accordance with at least grade 3 of EN ISO 3696 : 1995. 5.1 Carrez I solution Dissolve 15,0 g of potassium hexacyanoferrate (II) trihydrate (K 4 Fe(CN) 6 .3H 2 O) in water and dilute to 100 ml with water. 5.2 Carrez II solution Dissolve 30 g of zinc sulfate heptahydrate (

29、Zn SO 4 .7H 2 O) in water and dilute to 100 ml with water. 5.3 Acetone (CH 3 COCH 3 ) 5.4 Petroleum ether, boiling range 40 C to 60 C. Clean the petroleum ether on a column filled with aluminium oxide (Al 2 O 3 )( 5.9). 5.5 Sodium sulfate (Na 2 SO 4 ), anhydrous. 5.6 Toluene (C 6 H 5 CH 3 ) 5.7 Elut

30、ion mixture A Mix four volumes of petroleum ether (5.4) with one volume of toluene (5.6). 5.8 Elution mixture B Mix two volumes of petroleum ether (5.4) with one volume of toluene (5.6). 5.9 Aluminium oxide (Al 2 O 3 ), activated, neutral, acitivity grade I, for column chromatography and adsorptive

31、filtration. 5.10 Aluminium oxide (Al 2 O 3 ), deactivated partly by suspension in water. Transfer 100 g of aluminium oxide (5.9) into a flask with a ground glass joint, add 12 ml of water and mix thoroughly until a fine uniform suspension is obtained. Keep the flask well stoppered. The uniform suspe

32、nsion obtained should stand for 2 h. NOTE. The deactivated aluminium oxide represents the activity level IV-V. Since the activity changes after long periods of storage, the required quantities should be used between 2 h and 24 h after preparation.Page 4 EN 12136 : 1997 BSI 1998 6 Apparatus WARNING.

33、Since the determination involves the use of volatile flammable solvents, electrical apparatus employed has to be in accordance with legislation relating to the hazards in using such solvents. Usual laboratory apparatus and, in particular, the following. 6.1 Spectrometer, capable of measuring at a wa

34、velength of 450 nm. 6.2 Glass or quartz cuvettes, of 10 mm optical path length, and which have no significant absorption at a wavelength of 450 nm. 6.3 Centrifuge, capable of producing a centrifugal acceleration of 2 000 g at the base of the centrifuge tube (6.4). NOTE. The rotational frequency requ

35、ired to give correct centrifugal acceleration can be calculated from the following equation: a = 11,18 r( n /1 000) 2 (1) where: a is the centrifugal acceleration; r is the radius of the centrifuge in centimetres, measured from the mid point (the centrifuge axis) to the bottom of the centrifuge tube

36、 when swung out; n is the rotation frequency per minute. 6.4 Centrifuge tubes, of 60 ml to 100 ml capacity. 6.5 Chromatography tube, having a length of between 250 mm and 300 mm and an inner diameter of 20 mm, equipped with a polytetrafluorethylene (PTFE) stopcock. 6.6 Rotary vacuum evaporator 6.7 S

37、eparating funnel, of 200 ml capacity. 6.8 Volumetric flask, of 100 ml capacity. 6.9 Round bottom flask, of 250 ml capacity. 6.10 Graduated flasks, of suitable capacity. 7 Procedure Carotenoids are sensitive to light and heat. Carry out the analysis in a place protected from direct sunlight or UV exp

38、osure. 7.1 Preparation of the test sample Normally products shall not be pre-treated, however dilution may be necessary and their analysis by this method shall be on a volumetric basis, results being expressed per litre of sample. The analysis of concentrated products may also be carried out on a vo

39、lumetric basis, after dilution to a known relative density. In this case, the relative density shall be indicated. Based on a weighed sample and taking the dilution factor for analysis into account, the results may also be expressed per kilogram of product. In products with a high viscosity and/or a

40、 very high content of cells (for example pulp), a determination on the basis of a weighed test sample is the usual procedure. 7.2 Determination of total carotenoid content Shake the liquid test sample thoroughly and pipette a test portion of 5 ml to 50 ml (V 1 ) into a centrifuge tube (6.4). Dilute

41、to 50 ml with water if necessary and add 1 ml each of Carrez I (5.1) and Carrez II (5.2) solutions. Mix thoroughly, allow to stand for about 2 min and then centrifuge at 2 000 g (6.3) for about 5 min. NOTE 1. The absorbance of the final extract made up to 100 ml (V 2 ) should not exceed 0,5 units. T

42、o allow for this, it is necessary to adjust the amount of sample taken (V 1 ) for the analysis. The following values are given as guidance: 25 ml for orange juice, 50 ml for orange nectar, 5 ml to 20 ml for passion fruit juice, 25 ml for passion fruit nectar, and 20 ml for tangerine juice. Remove th

43、e nearly colourless supernatant solution by decantation. Add 40 ml of acetone (5.3) to the precipitate in the centrifuge tube, mix thoroughly with a glass rod and continue to stir for about 3 min. Centrifuge again for about 5 min and decant the supernatant yellow acetone solution into a separating f

44、unnel (6.7). Add 50 ml of petroleum ether (5.4) using a measuring cylinder (6.10). Shake this mixture thoroughly and allow to stand briefly to allow the organic phase to separate. Discard the aqueous phase of about 10 ml. Repeat the extraction of the precipitate in the centrifuge tube with 20 ml of

45、acetone and proceed as before. After centrifugation, carefully transfer the supernatant organic extract into the separating funnel. Normally one repetition of the extraction is sufficient. If necessary, the extraction can be repeated once more. NOTE 2. If the extraction procedure is carried out more

46、 than twice beware of the formation of an emulsion. If there is the danger of incomplete extraction, it is recommended to start from the beginning of the procedure using a smaller volume of test solution (V 1 ). Wash the organic phase with 50 ml of water by agitating the extract carefully. As soon a

47、s the slight emulsion which may be formed during agitation disappears discard the aqueous phase. Transfer the organic phase into a new centrifuge tube containing 2 g of the anhydrous sodium sulfate (5.5) as a drying agent. Mix briefly with a glass rod and centrifuge. Pour the petroleum ether phase c

48、arefully into a 100 ml volumetric flask (6.8). Resuspend the precipitate in a further 30 ml of petroleum ether, stir with a glass rod and centrifuge again. Add the slightly coloured petroleum ether fraction to the first extract and make the flask up to 100 ml (V 2 ) with petroleum ether. Measure the

49、 absorbance of the solution against a petroleum ether blank at a wavelength of 450 nm, using cuvettes of 10 mm path length (6.2).Page 5 EN 12136 : 1997 BSI 1998 7.3 Determination of individual carotenoid fractions 7.3.1 Preparation of chromatography column Suspend 28 g to 30 g of partly deactivated aluminium oxide (5.10) in about 30 ml of petroleum ether and pour the suspension into a chromatography tube (6.5) that has previously been stoppered with a small plug of glass wool and filled with petroleum ether. Run off the petroleum