GB 5009.285-2022 PDF EnglishUS$290.00 · In stock · Download in 9 seconds
GB 5009.285-2022: National food safety standard - Determination of Vitamin B12 in foods Delivery: 9 seconds. True-PDF full-copy in English & invoice will be downloaded + auto-delivered via email. See step-by-step procedure Status: Valid
Similar standardsGB 5009.285-2022: National food safety standard - Determination of Vitamin B12 in foods---This is an excerpt. Full copy of true-PDF in English version (including equations, symbols, images, flow-chart, tables, and figures etc.), auto-downloaded/delivered in 9 seconds, can be purchased online: https://www.ChineseStandard.net/PDF.aspx/GB5009.285-2022NATIONAL STANDARD OF THE PEOPLE’S REPUBLIC OF CHINA National food safety standard - Determination of Vitamin B12 in foods Issued on. JUNE 30, 2022 Implemented on. DECEMBER 30, 2022 Issued by. National Health Commission of the People’s Republic of China; State Administration for Market Regulation. Table of ContentsForeword... 3 1 Scope... 4 2 Principle... 4 3 Reagents and materials... 4 4 Instruments and apparatuses... 6 5 Analysis steps... 6 6 Description of the analysis results... 8 7 Precision... 9 8 Others... 9 9 Principle... 9 10 Reagents and materials... 9 11 Instruments and apparatuses... 11 12 Analysis steps... 12 13 Description of the analysis results... 15 14 Precision... 15 15 Others... 15 16 Principle... 16 17 Reagents and materials... 16 18 Instruments and equipment... 18 19 Test procedures... 19 20 Description of the analysis results... 21 21 Precision... 22 22 Others... 22 Appendix A Immunoaffinity column reference verification method... 23 Appendix B Liquid chromatogram... 24 Appendix C Liquid chromatography - mass spectrum... 25 Appendix D Medium preparation method... 26ForewordThis Standard replaces GB 5413.14-2010 National food safety standard - Determination of vitamin B12 in foods for infants and young children, milk and milk products. Compared with GB 5413.14-2010, the major changes of this Standard are as follows. -- Add Method I Liquid chromatography; -- Add Method II Liquid chromatography - mass spectrometry; -- Remove normative references; -- Modify the original microbiological method to method III. National food safety standard - Determination of Vitamin B12 in foods1 ScopeThis Standard specifies the determination method of vitamin B12 in foods. Method I of liquid chromatography applies to the determination of vitamin B12 in foods for infants and young children, milk and milk products, meat and meat products. Method II of liquid chromatography - mass spectrometry applies to the determination of vitamin B12 in foods for infants and young children, milk and milk products, meat and meat products, ready-to-eat cereals, baked foods, jelly, and beverages. Method III of microbiological method applies to the determination of vitamin B12 in foods for infants and young children, milk and milk products. Method I – Liquid chromatography2 PrincipleAfter the sample is enzymatically hydrolyzed, use potassium cyanide (or sodium cyanide) solution to convert cobalamin isomers (hydroxocobalamin, methylcobalamin and 5-deoxyadenosylcobalamin, etc.) to cyanocobalamin. After the sample solution is purified and concentrated by the immunoaffinity column, use the reversed-phase liquid chromatography column for separation, the ultraviolet detector for detection, and the external standard method for quantification.3 Reagents and materialsUnless otherwise specified, all the reagents are analytical reagents, and the water is grade-1 water which is specified by GB/T 6682. 3.1 Reagents 3.1.1 Anhydrous sodium acetate (CH3COONa). 3.1.2 Acetic acid (CH3COOH). 3.1.3 Methanol (CH3OH). chromatographic grade. 3.1.4 Acetonitrile (CH3CN). chromatographic grade. 3.1.5 Trifluoroacetic acid (CF3COOH). chromatographic grade. 3.1.6 Potassium cyanide or sodium cyanide (KCN/NaCN). 3.1.7 Pepsin (CAS number. 9001-75-6, activity ≥ 400 U/mg). 3.1.8 Amylase (activity ≥ 50 U/mg). 3.1.9 Ethanol (C2H6O). 3.2 Preparation of reagents 3.3 Standard Vitamin B12 (cyanocobalamin) standard substance (C63H88CoN14O14P, CAS number. 68-19-9). purity ≥99%, or standard substance certified by the nation and granted a certificate of reference material. 3.4 Preparation of standard solutions 3.5 Materials4 Instruments and apparatuses4.1 Liquid chromatograph. equipped with a UV detector. 4.2 Balance. sensitivity 0.01 g, 0.001 g and 0.000 01 g. 4.3 pH meter. accuracy 0.01. 4.5 Ultrasonic cleaner. 4.6 Centrifuge. speed ≥ 10 000 r/min.5 Analysis stepsNote. Avoid ultraviolet light during the operation, and operate as far away from light as possible. 5.1 Sample pretreatment 5.1.1 Sample preparation Crush and grind the solid sample, or use a meat grinder to make it into chyme; homogenize and mix. Shake and mix the liquid sample before testing. 5.1.2 Sample extraction Weigh 5 g ~ 10 g of the sample (accurate to 0.01 g) into a 150 mL conical flask with stopper; add 30 mL of sodium acetate buffer, 0.2 g of pepsin, 0.05 g of amylase and 2 mL of potassium cyanide (or sodium cyanide) solution in turn; mix well. Put the sample solution into a water bath constant temperature oscillator; at 37 °C, carry out enzymatic hydrolysis for 30 minutes (for meat samples, carry out enzymatic hydrolysis for 10 h ~ 16 h). 5.1.3 Purification Connect the immunoaffinity column to the solid-phase extraction device. After discarding the buffer in the immunoaffinity column, pipette an appropriate amount of the above filtrate (containing 10 ng ~ 500 ng of vitamin B12) to pass through the column; adjust the column passing speed to 2 mL/min ~ 3 mL/min. After the sample solution has completely passed through the column, use 10 mL of water to rinse the immunoaffinity column at a steady flow rate and drain. Place a 10 mL glass test tube under the immunoaffinity column; use 3 mL of methanol to elute in three times; collect all the eluates; use nitrogen flow to blow slowly below 60 °C until nearly dry; use 0.04% trifluoroacetic acid solution to fix the volume to 1.0 mL; vortex for 30 s to dissolve the residue; filter through a 0.22 μm filter; test. 5.2 Liquid chromatography reference conditions 5.2.1 Chromatographic column. C18 column (column length 150 mm, column inner diameter 4.6 mm, packing particle size 2.5 μm), or equivalent. 5.2.2 Mobile phase. phase A, 0.04% trifluoroacetic acid solution; phase B, acetonitrile. 5.2.3 Gradient elution. 0 min ~ 6.0 min, 90% A; 6.0 min ~ 8.5 min, 90% ~ 0% A; 8.5 min ~ 14.0 min, 90% A. 5.2.4 Flow velocity. 0.8 mL/min. 5.3 Preparation of the standard curve Inject the standard series solutions into the liquid chromatograph from low concentration to high concentration in turn; measure the corresponding chromatographic peak areas. 5.4 Sample determination Inject the to-be-tested sample solution into the liquid chromatograph, to obtain the peak area of vitamin B12 in the to-be-tested solution; measure the concentration of vitamin B12 in the test solution according to the standard curve. The response value of vitamin B12 in the to-be-tested sample solution shall be within the linear range of the standard curve. If it exceeds the linear range, the solution on the machine shall be diluted, or the sampling amount shall be adjusted and the analysis shall be carried out again after processing according to the sample analysis steps. 5.5 Blank test Do not weigh the sample; operate according to the sample analysis steps. Substances that interfere with the to-be-measured components shall not be contained.6 Description of the analysis resultsThe content of vitamin B12 (calculated as cyanocobalamin) in the sample is calculated according to Formula (1).7 PrecisionThe absolute difference of two independent test results obtained under repeatability cannot exceed 15% of the arithmetic mean value.8 OthersWhen the sampling amount is 5.00 g, the detection limit of foods for infants and young children, dairy products, meat and meat products is 0.2 μg/100 g, and the quantification limit is 0.5 μg/100 g.9 PrincipleAfter the sample is enzymatically hydrolyzed, use potassium cyanide (or sodium cyanide) solution to convert cobalamin isomers (hydroxocobalamin, methylcobalamin and 5-deoxyadenosylcobalamin, etc.) to cyanocobalamin. After the sample solution is purified and concentrated by the immunoaffinity column, use the reversed-phase liquid chromatography column for separation, the tandem mass spectrometry for detection, and the isotope internal standard method for quantification.10 Reagents and materialsUnless otherwise specified, all the reagents are analytical reagents, and the water is grade-1 water which is specified by GB/T 6682. 10.1 Reagents 10.1.1 Anhydrous sodium acetate (CH3COONa). 10.1.2 Sodium hydroxide (NaOH). 10.1.3 Acetic acid (CH3COOH). 10.1.4 Acetonitrile (CH3CN). chromatographic pure. 10.1.5 Ammonium acetate (CH3COONH4). chromatographic pure. 10.1.6 Potassium cyanide or sodium cyanide (KCN/NaCN). 10.1.7 Pepsin (CAS number. 9001-75-6, activity ≥ 400 U/mg). 10.1.8 Amylase (activity ≥ 50 U/mg). 10.1.9 Ethanol (C2H6O). 10.2 Preparation of reagents 10.3 Standard substance 10.3.1 Vitamin B12 (cyanocobalamin) standard substance (C63H88CoN14O14P, CAS number. 68-19-9). purity ≥99%, or standard substance certified by the nation and granted a certificate of reference material. 10.3.2 Vitamin B12 isotope internal standard solution (13C7-C63H88CoN14O14P). 1 μg/mL methanol solution. 10.4 Preparation of standard solutions 10.4.1 Vitamin B12 standard stock solution (1 mg/mL). Weigh 10 mg (accurate to 0.01 mg) of standard vitamin B12 in a 50 mL beaker; use ethanol solution to dissolve it; then, transfer it to a 10 mL volumetric flask; use ethanol solution to fix the volume to the mark; shake well; transfer to a brown reagent bottle; store at -20 °C in the dark. This solution is valid for 6 months. 10.4.2 Vitamin B12 standard intermediate solution (10 μg/mL). Draw 1.00 mL of vitamin B12 standard stock solution; put it in a 100 mL volumetric flask; use ethanol solution to dilute to the mark. Transfer it to a brown reagent bottle; store at 4 °C in the dark. The validity period is 1 month. 10.5 Materials 10.5.1 Vitamin B12 immunoaffinity column, column capacity ≥800 ng, column recovery ≥85% (see Appendix A for the verification method). 10.5.2 Glass fiber filter paper. 10.5.3 Microporous membrane. water phase, 0.22 μm. ......Source: Above contents are excerpted from the full-copy PDF -- translated/reviewed by: www.ChineseStandard.net / Wayne Zheng et al. Tips & Frequently Asked Questions:Question 1: How long will the true-PDF of English version of GB 5009.285-2022 be delivered?Answer: The full copy PDF of English version of GB 5009.285-2022 can be downloaded in 9 seconds, and it will also be emailed to you in 9 seconds (double mechanisms to ensure the delivery reliably), with PDF-invoice.Question 2: Can I share the purchased PDF of GB 5009.285-2022_English with my colleagues?Answer: Yes. The purchased PDF of GB 5009.285-2022_English will be deemed to be sold to your employer/organization who actually paid for it, including your colleagues and your employer's intranet.Question 3: Does the price include tax/VAT?Answer: Yes. Our tax invoice, downloaded/delivered in 9 seconds, includes all tax/VAT and complies with 100+ countries' tax regulations (tax exempted in 100+ countries) -- See Avoidance of Double Taxation Agreements (DTAs): List of DTAs signed between Singapore and 100+ countriesQuestion 4: Do you accept my currency other than USD?Answer: Yes. www.ChineseStandard.us -- GB 5009.285-2022 -- Click this link and select your country/currency to pay, the exact amount in your currency will be printed on the invoice. Full PDF will also be downloaded/emailed in 9 seconds.How to buy and download a true PDF of English version of GB 5009.285-2022?A step-by-step guide to download PDF of GB 5009.285-2022_EnglishStep 1: Visit website https://www.ChineseStandard.net (Pay in USD), or https://www.ChineseStandard.us (Pay in any currencies such as Euro, KRW, JPY, AUD).Step 2: Search keyword "GB 5009.285-2022". Step 3: Click "Add to Cart". If multiple PDFs are required, repeat steps 2 and 3 to add up to 12 PDFs to cart. Step 4: Select payment option (Via payment agents Stripe or PayPal). Step 5: Customize Tax Invoice -- Fill up your email etc. Step 6: Click "Checkout". Step 7: Make payment by credit card, PayPal, Google Pay etc. After the payment is completed and in 9 seconds, you will receive 2 emails attached with the purchased PDFs and PDF-invoice, respectively. Step 8: Optional -- Go to download PDF. Step 9: Optional -- Click Open/Download PDF to download PDFs and invoice. See screenshots for above steps: Steps 1~3 Steps 4~6 Step 7 Step 8 Step 9 |
