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GB 5413.20-2022 PDF English

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GB 5413.20-2022: National food safety standard - Determination of Choline in foods, milk and milk products for infants and young children
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GB 5413.20: Historical versions

Standard IDUSDBUY PDFDeliveryStandard Title (Description)Status
GB 5413.20-2022215 Add to Cart Auto, 9 seconds. National food safety standard - Determination of Choline in foods, milk and milk products for infants and young children Valid
GB 5413.20-201370 Add to Cart Auto, 9 seconds. National Food Safety Standard Determination of choline in infant foods and dairy products Obsolete
GB/T 5413.20-1997199 Add to Cart 2 days Milk powder and formula foods for infant and young children--Determination of choline Obsolete

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GB 5413.20-2022: National food safety standard - Determination of Choline in foods, milk and milk products for infants and young children


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NATIONAL STANDARD OF THE PEOPLE’S REPUBLIC OF CHINA National Food Safety Standard – Determination of Choline in Foods, Milk and Milk Products for Infants and Young Children 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 Contents

Foreword... 3 1 Scope... 4 2 Principle... 4 3 Reagents and Materials... 4 4 Apparatus... 6 5 Analysis Procedures... 6 6 Presentation of the Analysis Results... 7 7 Precision... 8 8 Others... 8 9 Principle... 9 10 Reagents and Materials... 9 11 Apparatus... 10 12 Analytical Procedures... 10 13 Presentation of the Analysis Results... 12 14 Precision... 12 15 Others... 13 16 Principle... 13 17 Reagents and Materials... 13 18 Apparatus... 15 19 Analytical Procedures... 15 20 Presentation of Analysis Results... 18 21 Precision... 18 22 Others... 18 Appendix A Ion Chromatogram of Choline Standard Solution... 19 Appendix B Choline Standard Solution Mass Spectrum Scanning Diagram, Standard Solution and Internal Standard Multi-Reaction Monitoring (MRM) Diagram... 21

Foreword

This Standard replaced GB 5413.20-2013 National Food Safety Standard - Determination of Choline in Infant Foods and Dairy Products. Compared with GB 5413.20-2013, the major changes of this Standard are as follows. --- Delete the second method of Ray's salt spectrophotometry; and add ion chromatography as the second method; --- Add liquid chromatography-tandem mass spectrometry as the third method. National Food Safety Standard – Determination of Choline in Foods, Milk and Milk Products for Infants and Young Children

1 Scope

This Standard specifies the method for the determination of choline in food, milk and milk products for infants and young children. This Standard applies to the determination of choline in food, milk and milk products for infants and young children. The First Method – Enzymic Colorimetric Method

2 Principle

The specimen is hydrolyzed by acid; and then reacts with the color developer to form colored substances after enzymatic action. The alkali content is proportional to the external standard method. Within a certain concentration range, the color depth is proportional to the choline content, and the external standard method is used for quantification.

3 Reagents and Materials

Unless otherwise specified, the reagents used in this method are all analytically pure; and the water is Class-3 water specified in GB/T 6682. 3.1 Reagents 3.1.1 Tris(hydroxymethyl)aminomethane [(CH2OH)3CNH2]. 3.1.2 Phenol (C6H5OH). 3.1.3 Concentrated hydrochloric acid (HCl). 3.1.4 Sodium hydroxide (NaOH). 3.1.5 Choline oxidase. ≥10U/mg; store at -20 ℃. 3.1.6 Peroxidase. ≥250U/mg; store at 2℃～8℃. 3.1.7 4-Aminoantipyrine (C11H13N3O). 3.1.8 Phospholipase D. ≥60U/mg, store at -20°C. 3.2 Preparation of reagents 3.3 Standard product Choline bitartrate standard product (C9H19NO7, relative molecular mass. 253.25, CAS number. 87-67-2). purity ≥ 99%, or the reference material certified by the state and awarded the reference material certificate. 3.4 Preparation of standard solution 3.5 Materials

4 Apparatus

4.1 Analytical balance. The sensitivities are 0.1 mg and 0.001 g, respectively. 4.3 pH meter. Accuracy 0.01. 4.4 Spectrophotometer.

5 Analysis Procedures

5.1 Sample preparation 5.1.1 Sample pretreatment Accurately take 20g (accurate to 0.001g) of the mixed liquid specimen into a 100mL conical flask; add 10mL of 3mol/L hydrochloric acid solution; stopper it and mix well. Accurately take 5g (accurate to 0.001g) of the mixed semi-solid or solid specimen into a 100mL conical flask; add 30mL of 1mol/L hydrochloric acid solution; stopper it and mix well. 5.1.3 Filtration Filter the hydrolyzate with filter paper; if the filtrate is not clear; filter again with a 0.45μm aqueous filter membrane syringe filter. The filtrate is collected for testing. 5.2 Determination 5.2.1 Drawing of standard curve Respectively pipette 2.00mL, 4.00mL, 6.00mL, 8.00mL, and 10.00mL of choline standard working solution (250mg/L) into a 10mL volumetric flask; make constant volume to the mark with water; mix well; and prepare the standard series working solutions with concentrations of 50.0 mg/L, 100 mg/L, 150 mg/L, 200 mg/L and 250 mg/L, respectively. Prepare 6 colorimetric tubes; one colorimetric tube is used as reagent blank (A); add 100μL of water; add 100μL of standard series working solutions to the other 5 colorimetric tubes, respectively; and then add 3.00 mL of color developer, respectively; mix well; the colorimetric tube is placed in a water bath at 37°C±2°C for 15 min for thermal-insulation reaction.

6 Presentation of the Analysis Results

6.1 Calculation of net absorbance value Usually formulated reagents shall produce a slight color; and the filtrate is not colorless due to hydrolysis. In order to remove these interference factors, The respective blank values (colorimetric tubes A and B) must be subtracted from the total absorbance value. The net absorbance value of the specimen is calculated according to Formula (1). 6.2 Calculation of choline content The choline content (by choline hydroxide) in the specimen shall be calculated according to Formula (2).

7 Precision

The absolute difference between the results of two independent determinations obtained under repeatability conditions shall not exceed 10% of the arithmetic mean.

8 Others

When the solid or semi-solid specimen weighs 5g, the detection limit of the method is 1mg/100g, and the limit of quantification is 3mg/100g.

9 Principle

The specimen is hydrolyzed by acid, purified by solid phase extraction column, then separated by ion chromatography; detected by conductivity detector, and quantified by external standard method.

10 Reagents and Materials

Unless otherwise specified, the reagents used in this method are all analytically pure; and the water is the Class-1 water specified in GB/T 6682. 10.1 Reagents 10.1.1 Concentrated hydrochloric acid (HCl). 10.1.2 Methanesulfonic acid (CH4O3S). chromatographically pure. 10.2 Preparation of reagents 10.2.1 Hydrochloric acid solution (1 mol/L). Pipette 85 mL of concentrated hydrochloric acid and inject it into about 900 mL of water; and dilute to 1000mL. 10.2.2 Hydrochloric acid solution (1.7 mol/L). Pipette 145 mL of concentrated hydrochloric acid and inject it into about 800 mL of water; and dilute to 1000mL. 10.3 Standard products Choline bitartrate standard product (C9H19NO7, relative molecular mass. 253.25, CAS number. 87-67-2). purity ≥ 99%; or a standard substance certified by the state and granted a standard substance certificate. 10.4 Preparation of standard solutions 10.5 Materials 10.5.1 0.45μm aqueous filter membrane syringe filter. 10.5.2 Purification column. C18 solid phase extraction cartridge 1.0mL or equivalent. 10.5.3 Syringe. 5mL or equivalent.

11 Apparatus

11.1 Ion chromatograph (IC). Equipped with a conductive detector. 11.2 Analytical balance. The sensitivity is 0.1mg and 0.001g, respectively. 11.3 Electrothermal constant temperature water bath device. The temperature can be controlled at 70℃±2℃. 11.4 Vortex mixer.

12 Analytical Procedures

12.1 Sample preparation 12.1.1 Specimen extraction 12.1.2 Specimen purification C18 solid phase extraction cartridge (1.0mL) is passed through successively with 10mL of methanol and 15mL of water before use; and stand for activation for 30min. Dilute the extraction solution 50 times (the dilution ratio can be adjusted appropriately according to the concentration of choline in the specimen, and it shall be no less than 10 times) with water; take about 15 mL of the diluted solution; pass through a 0.45μm aqueous filter membrane and a C18 solid phase extraction cartridge (1.0mL); discard the first 3mL; and collect the later eluate for testing. ......
Source: Above contents are excerpted from the full-copy PDF -- translated/reviewed by: www.ChineseStandard.net / Wayne Zheng et al.
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