1、raising standards worldwideNO COPYING WITHOUT BSI PERMISSION EXCEPT AS PERMITTED BY COPYRIGHT LAWBSI Standards PublicationBS EN 16159:2012Animal feeding stuffs Determination of seleniumby hydride generation atomicabsorption spectrometry(HGAAS) after microwavedigestion (digestion with 65 %nitric acid
2、 and 30 % hydrogenperoxide)BS EN 16159:2012 BRITISH STANDARDNational forewordThis British Standard is the UK implementation of EN 16159:2012.The UK participation in its preparation was entrusted to TechnicalCommittee AW/10, Animal feeding stuffs.A list of organizations represented on this committee
3、can beobtained on request to its secretary.This publication does not purport to include all the necessaryprovisions of a contract. Users are responsible for its correctapplication. The British Standards Institution 2012. Published by BSI StandardsLimited 2012ISBN 978 0 580 66997 2ICS 65.120Complianc
4、e with a British Standard cannot confer immunity fromlegal obligations.This British Standard was published under the authority of theStandards Policy and Strategy Committee on 29 February 2012.Amendments issued since publicationDate Text affectedBS EN 16159:2012EUROPEAN STANDARD NORME EUROPENNE EURO
5、PISCHE NORM EN 16159 February 2012 ICS 65.120 English Version Animal feeding stuffs - Determination of selenium by hydride generation atomic absorption spectrometry (HGAAS) after microwave digestion (digestion with 65 % nitric acid and 30 % hydrogen peroxide) Aliments pour animaux - Dosage du slnium
6、 par spectromtrie dabsorption atomique par gnration dhydrures (SAAGH) aprs digestion par micro-ondes (extraction avec de lacide nitrique 65 % et du peroxyde dhydrogne 30 %) Futtermittel - Bestimmung von Selen mit Atomabsorptionsspektrometrie-Hydridtechnik (HD-AAS) nach Mikrowellen-Druckaufschluss (A
7、ufschluss mit 65 % Salpetersure und 30 % Wasserstoffperoxid) This European Standard was approved by CEN on 30 December 2011. 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 wi
8、thout any alteration. Up-to-date lists and bibliographical references concerning such national standards may be obtained on application to the CEN-CENELEC Management Centre or to any CEN member. This European Standard exists in three official versions (English, French, German). A version in any othe
9、r language made by translation under the responsibility of a CEN member into its own language and notified to the CEN-CENELEC Management Centre has the same status as the official versions. CEN members are the national standards bodies of Austria, Belgium, Bulgaria, Croatia, Cyprus, Czech Republic,
10、Denmark, Estonia, Finland, France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway, Poland, Portugal, Romania, Slovakia, Slovenia, Spain, Sweden, Switzerland, Turkey and United Kingdom. EUROPEAN COMMITTEE FOR STANDARDIZATION COMIT EUROPEN
11、DE NORMALISATION EUROPISCHES KOMITEE FR NORMUNG Management Centre: Avenue Marnix 17, B-1000 Brussels 2012 CEN All rights of exploitation in any form and by any means reserved worldwide for CEN national Members. Ref. No. EN 16159:2012: EBS EN 16159:2012EN 16159:2012 (E) 2 Contents Page Foreword 31 Sc
12、ope 42 Normative references 43 Principle 44 Reagents .55 Apparatus and equipment 66 Procedure .76.1 General 76.2 Preparation of the test solution 76.3 Measurement of the test solution 86.3.1 Pre-dilution of the test solution 86.3.2 Pre-reduction of the (pre-diluted) test solution 86.3.3 Spectrometer
13、 settings of the flow-injection hydride generation atomic absorption spectrometer (HGAAS) 96.3.4 HGAAS determination .97 Calculation . 108 Precision 108.1 Introduction . 108.2 General . 118.3 Repeatability 118.4 Reproducibility 119 Test report . 11Annex A (informative) Results of the inter-laborator
14、y test 12Annex B (informative) Flowchart - Determination of selenium by hydride generation atomic absorption spectrometry (HGAAS) after microwave digestion (digestion with 65 % nitric acid and 30 % hydrogen peroxide . 13Annex C (informative) Alternative digestion procedure with the same digestion ef
15、ficiency: Acid digestion with a mixture of 65 % nitric acid and 70 % perchloric acid (7:3 by volume) at atmospheric pressure 14C.1 Preparation of selenium calibration solutions 14C.2 Preparation of the test solution . 14C.3 Measurement of the test solution . 15C.4 Ten rules for automated wet ashing
16、with perchloric acid 15Bibliography . 16BS EN 16159:2012EN 16159:2012 (E) 3 Foreword This document (EN 16159:2012) has been prepared by Technical Committee CEN/TC 327 “Animal feeding stuffs”, the secretariat of which is held by NEN. This European Standard shall be given the status of a national stan
17、dard, either by publication of an identical text or by endorsement, at the latest by August 2012, and conflicting national standards shall be withdrawn at the latest by August 2012. Attention is drawn to the possibility that some of the elements of this document may be the subject of patent rights.
18、CEN and/or CENELEC shall not be held responsible for identifying any or all such patent rights. This document has been prepared under a mandate given to CEN by the European Commission and the European Free Trade Association. According to the CEN/CENELEC Internal Regulations, the national standards o
19、rganizations of the following countries are bound to implement this European Standard: Austria, Belgium, Bulgaria, Croatia, Cyprus, Czech Republic, Denmark, Estonia, Finland, France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway, Poland,
20、 Portugal, Romania, Slovakia, Slovenia, Spain, Sweden, Switzerland, Turkey and the United Kingdom. BS EN 16159:2012EN 16159:2012 (E) 4 1 Scope This European Standard specifies a method for the determination of selenium in animal feeding stuffs by hydride generation atomic absorption spectrometry (HG
21、AAS) after microwave pressure digestion. The method was successfully tested by an inter-laboratory study of CEN/TC 327/WG 4 in the range of 0,25 mg/kg to 74 mg/kg. The limit of quantification is 0,5 g/l of the test solution which corresponds to the calibration standard 2. Using a test portion of 0,5
22、 g and a volume of the test solution of 25 ml after pressure digestion the limit of quantification is calculated as 0,125 mg/kg in the feed material. NOTE A lower limit of quantification could be achieved each laboratory has to prove it. 2 Normative references The following documents, in whole or in
23、 part, are normatively referenced in this document and are indispensable for its application. For dated references, only the edition cited applies. For undated references, the latest edition of the referenced document (including any amendments) applies. EN ISO 3696, Water for analytical laboratory u
24、se Specification and test methods (ISO 3696) EN ISO 6497, Animal feeding stuffs Sampling (ISO 6497) prEN ISO 6498, Animal feeding stuffs Guidelines for sample preparation (ISO/DIS 6498) 3 Principle Selenium is determined in the test solution by hydride generation atomic absorption spectrometry (fluo
25、rescence hydride generation atomic absorption) after microwave pressure digestion and a pre-reduction step. The homogenised feeding stuff test sample is digested by nitric acid and hydrogen peroxide under pressure and high temperatures in a microwave-heated pressure digestion system. Selenium ions o
26、f the test solution are reduced with hydrochloric acid to selenium (IV) and converted to selenium hydride (SeH2) by sodium borohydride. This selenium hydride is transferred by a gas stream to a heated measurement cell and decomposed. The absorption at the selenium line at 196,0 nm corresponds to the
27、 amount of selenium. NOTE Selenium (VI) is not determined by the hydridisation as described here. It is therefore necessary to adjust the digestion conditions and to exercise a pre-reduction step with hydrochloric acid to yield only selenium (IV). Other digestion procedures with the same digestion e
28、fficiency or other measurement systems like Fl-HGAAS or hydride generation inductively coupled plasma optical emission spectrometry are possible (see Annex C). WARNING The use of this standard can involve hazardous materials, operations and equipment. This standard does not purport to address all th
29、e safety problems associated with its use. It is the responsibility of the user of this standard to establish appropriate safety and health practices and determine the applicability of regulatory limitations prior to use. BS EN 16159:2012EN 16159:2012 (E) 5 4 Reagents The concentration of the trace
30、elements in the reagents and water used shall be low enough not to affect the results of the determination. A blank should be measured simultaneously with the test samples on each day of the analysis to control contamination and carry over with selenium in the reagents and apparatus used. Use only w
31、ater of quality 2 as described in EN ISO 3696. NOTE High purity is essential to avoid potential contamination. Therefore, only use reagents available with high purity or perform an extraction by a sub-boiling distillation for nitric acid (4.1). 4.1 Nitric acid, not less than 65 % (mass fraction), of
32、 approximately (HNO3) = 1,4 g/ml. 4.2 Diluted nitric acid, mix 100 ml nitric acid (4.1) with water to 1 l. 4.3 Hydrogen peroxide, not less than 30 % (mass fraction), of approximately (H2O2) 1,1 g/ml. 4.4 Hydrochloric acid, 30 %, mass concentration of approximately (HCl) = 1,15 g/ml. 4.5 Diluted hydr
33、ochloric acid, e.g. about 3 % (mass fraction), as carrier solution for the use in the flow-injection-procedure. EXAMPLE Dilute approximately 90 ml of hydrochloric acid (4.4) to 1 l with water. 4.6 Sodium borohydride solution, e.g. c = 2 g/l. Dissolve 2 g of sodium hydroxide pellets in water, add 2 g
34、 of sodium borohydride and dilute to 1 000 ml with water into 1 000 ml flask (5.3). Prepare a fresh solution daily and, when necessary, filter before use. When the analysis procedure takes longer, it is recommended to cool the sodium borohydride solution, i.e. with ice around the flask, during its u
35、se in the HGAAS measurement. NOTE 1 The concentration by mass of the sodium borohydride solution may vary with the system and the instructions of the relevant manufacturer shall therefore be observed. NOTE 2 Sodium borohydride, stable aq. solution, 4,4 mol/l in 14 mol/l NaOH is also commercially ava
36、ilable. WARNING It is essential to observe the safety instructions for working with sodium borohydride. Sodium borohydride forms hydrogen with acids and this can result in an explosive air/hydrogen mixture. A permanent extraction system shall be provided at the point where measurements are carried o
37、ut. 4.7 Selenium stock solution, c (Se) = 1 000 mg/l. The stock solution is commercially available. It is advisable to use certified stock solutions. Otherwise, dissolve 1,4053 g of selenium dioxide (SeO2) and 2 g sodium hydroxide in approximately 50 ml water, and dilute to 1 000 ml with water. 4.8
38、Selenium standard solution, c (Se) = 1 mg/l. Dilute e.g. 100 l of the stock solution (4.7) in a 100 ml flask (5.3) to give a concentration of 1 mg/l. The selenium standard solution shall contain an adequate amount of hydrochloric acid, e.g. 2 ml of hydrochloric acid (4.4) per 100 ml. NOTE The standa
39、rd solution is stable for at least three months. 4.9 Selenium calibration solutions. BS EN 16159:2012EN 16159:2012 (E) 6 For the preparation of five calibration solutions, the following procedure is recommended: Take aliquots of 0 l, 50 l, 250 l, 500 l and 1 000 l of the selenium standard solution (
40、4.8) into 100 ml flasks (5.3). After the addition of 20 ml of nitric acid (4.2) and 10 ml of hydrochloric acid (4.4) the calibration solutions are heated for 20 min in a water bath at 80 C (see pre-reduction step 6.3.2). After cooling down to room temperature, the flasks (5.3) are made up to the mar
41、k with water and the calibration solutions are measured. The selenium concentrations of the calibration solutions are: 0 g/l; 0,5 g/l; 2,5 g/l; 5 g/l and 10 g/l (see Table 1). Table 1 Recommended calibration solutions (4.9) for the determination of selenium Selenium (Se) Concentration of calibration
42、 solution (4.9) after pre-reduction procedure g/l Aliquot of selenium standard solution (4.8) transferred in 100 ml flasks (5.3) (pre-reduction step) l Calibration standard 1 0 0 Calibration standard 2 0,5 50 Calibration standard 3 2,5 250 Calibration standard 4 5 500 Calibration standard 5 10 1 000
43、 Choose the concentrations of the calibration solutions so as not to exceed the linear range of the calibration function. It is recommended to use a minimum of five calibration solutions with different concentrations. The calibration solutions are measured from the lowest to the highest concentratio
44、n. In general, the calibration curve should be linear. Using a non-linear calibration function is possible if it is well described. NOTE Prepare fresh calibration solutions (inclusive pre-reduction step) on the day of the analysis. 5 Apparatus and equipment To minimise the contamination, all apparat
45、us which come into direct contact with the sample and the solutions should be carefully pre-treated according to EN 13804. 5.1 Microwave-heated pressure digestion apparatus with inert reaction vessels, i.e. made of polytetrafluoroethylene (PTFE), perfluoroalkoxy (PFA), fluorinated ethylene propylene
46、 (FEP) or quartz, suitable for digestion temperatures of more than 200 C. NOTE 1 The microwave oven should be generally resistant to corrosion and especially the electronics should be protected against corrosion to ensure safe operation. The ventilation should transfer the acid vapours to an extract
47、or hood or a fume cupboard. NOTE 2 The reaction vessels should have a safety valve designed for a pressure of 1 000 kPa. BS EN 16159:2012EN 16159:2012 (E) 7 5.2 Pipettes, volumetric and/or graduated, 100 l, 250 l, 600 l, 1 000 l, 1 500 l, 2 ml, 2,5 ml and 10 ml. 5.3 Volumetric flasks, 25 ml, 50 ml,
48、100 ml, 500 ml and 1 000 ml. 5.4 Flow-injection hydride system, with sample loop, e.g. 500 l. 5.5 Atomic absorption spectrometer (AAS), with measurement recording system, background correction, heated quartz cell and accessories for the hydride procedure. 5.6 Element-specific lamp for selenium. NOTE
49、 An electrodeless discharge lamp would provide a higher sensitivity compared to a hollow-cathode lamp 5.7 Ultrasonic bath and/or water bath. 5.8 Analytical balance, accurate to 0,1 mg. 6 Procedure 6.1 General Sampling and preparation of a test sample is not part of the method. A recommended sampling method and method for sample preparation is given in EN ISO 6497 and prEN ISO 6498. NOTE The use of a stationary or espe
