GB 5009.259-2023 PDF English
US$260.00 · In stock · Download in 9 secondsGB 5009.259-2023: National food safety standard - Determination of biotin in foods Delivery: 9 seconds. True-PDF full-copy in English & invoice will be downloaded + auto-delivered via email. See step-by-step procedureStatus: Valid GB 5009.259: Evolution and historical versions
| Standard ID | Contents [version] | USD | STEP2 | [PDF] delivery | Name of Chinese Standard | Status |
| GB 5009.259-2023 | English | 260 |
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National food safety standard - Determination of biotin in foods
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| GB 5009.259-2016 | English | 75 |
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National food safety standard -- Determination of free biotin in foods for infants and young children, milk and milk products
| Obsolete |
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GB 5009.259-2023: National food safety standard - Determination of biotin 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/GB5009.259-2023
GB
NATIONAL STANDARD OF THE
PEOPLE’S REPUBLIC OF CHINA
National Food Safety Standard - Determination of Biotin in
Foods
Issued on. SEPTEMBER 6, 2023
Implemented on. MARCH 6, 2024
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
Method I - Liquid Chromatography - Tandem Mass Spectrometry... 4
2 Principle... 4
3 Reagents and Materials... 4
4 Instruments and Equipment... 6
5 Analytical Procedures... 6
6 Expression of Analysis Results... 9
7 Precision... 10
8 Others... 10
Method II - Microbiological Method... 10
9 Principle... 10
10 Reagents and Materials... 10
11 Instruments and Equipment... 12
12 Analytical Procedures... 13
13 Expression of Analysis Results... 18
14 Precision... 19
15 Others... 19
Appendix A Mass Spectrum Scan and MRM Chromatogram of Biotin Standard
Solution... 20
Appendix B Preparation of Culture Medium... 21
Foreword
This Standard serves as a replacement of GB 5009.259-2016 National Food Safety Standard -
Determination of Biotin in Foods.
In comparison with GB 5009.259-2016, the main changes are as follows.
---Method 1 “Liquid Chromatography - Tandem Mass Spectrometry” is added, and the
“Microbiological Method” is adjusted to Method 2;
---The microplate culture method is added to Method 2 - “Microbiological Method”;
---The scope of application, sample category and pre-treatment method of the Standard are
modified;
---The expression of the drawing of standard curve and the preparation of sample
extracting solution is modified;
---The detection limit, quantitation limit and linear range are modified.
National Food Safety Standard - Determination of Biotin in
Foods
1 Scope
This Standard specifies the methods for the determination of biotin in foods.
Method 1 - liquid chromatography - tandem mass spectrometry is applicable to the
determination of biotin in prepared milk powder and special dietary foods.
Method 2 - microbiological method is applicable to the determination of biotin in foods.
Method I - Liquid Chromatography - Tandem Mass
Spectrometry
2 Principle
The specimen is dissolved and extracted, and the starch-containing specimen is subject to
enzymatic hydrolysis by amylase, protein precipitation and centrifugal filtration, and separated
on a C18 reversed-phase chromatography column. Adopt the liquid chromatography - tandem
mass spectrometry multi-ion reaction monitoring mode for detection, and the isotope dilution
internal standard method for quantitative determination.
3 Reagents and Materials
Unless it is otherwise specified, the reagents used in this Method are all analytically pure, and
the water is Grade-1 water specified in GB/T 6682.
3.1 Reagents and Materials
3.1.3 Ethanol (CH3CH2OH). chromatographically pure.
3.1.4 Ammonium formate (HCOONH4). chromatographically pure.
3.1.5 Perchloric acid (HClO4). 70% ~ 72%.
3.1.6 Sodium hydroxide (NaOH). purity 99.9%.
3.1.7 Amylase. Taka-amylase, CAS. 9001-19-8, enzyme activity 100 U/mg.
3.2 Preparation of Reagents
3.3 Reference Material
3.5 Preparation of Isotope Internal Standard Solutions
3.6 Preparation of Standard Series of Working Solutions
Respectively and accurately draw-take an appropriate amount of biotin standard working
solution into 10 mL volumetric flasks, accurately add 150 L of biotin-D4 isotope internal
standard working solution (1 g/mL), use mobile phase A to reach a constant volume to the
scale, so that the mass concentration of the standard series of biotin is respectively. 1.0 ng/mL,
2.0 ng/mL, 5.0 ng/mL, 10 ng/mL, 15 ng/mL, 20 ng/mL and 25 ng/mL. The mass concentration
of isotope in each series of solution is 15 ng/mL. Prepare them right before use.
4 Instruments and Equipment
4.1 Liquid chromatograph - tandem mass spectrometer.
4.2 Balance. with a division value of 0.001 g and 0.01 mg, respectively.
4.4 pH meter. with an accuracy of 0.01.
4.5 Ultrasonic oscillator.
4.6 Vortex mixer. 600 r/min ~ 3,200 r/min.
4.8 2 mm aperture test sieve (optional).
5 Analytical Procedures
5.1 Sample Preparation
5.1.1 Solid samples
The sampling size needs to be greater than 0.5 kg. Use a high-speed pulverizer to pulverize
non-uniform samples, until all of them pass through a 2 mm aperture test sieve. After evenly
mixing, divide the samples to 100 g and store in a wide-mouth bottle. Seal it and reserve it for
testing. For uniform samples, directly evenly mix them and reserve them for testing.
5.1.2 Semi-solid samples
The sampling size needs to be greater than 0.5 kg. At least 3 packages (from the same batch)
need to be collected. After all samples are homogenized and evenly mixed in a container, store
any 100 g of them in a wide-mouth bottle. Seal it and reserve it for testing.
5.1.3 Liquid samples
The sampling size needs to be greater than 0.5 L. At least 3 packages (from the same batch)
need to be collected. After all samples are evenly mixed in a container, store any 100 mL of
them in a wide-mouth bottle. Seal it and reserve it for testing.
5.2 Pre-treatment of Samples
5.3 Reference Conditions of Instrument Determination
5.4 Qualitative Determination
The selected ions all appear, and in the same detection batch, compared with the relative
abundance of the standard solution of an equivalent concentration, the relative abundance of
two daughter ions of biotin in the sample shall have an allowable deviation of not exceeding
the range specified in Table 3.
5.5 Drawing of Standard Curve
In accordance with the concentration from low to high, successively inject the biotin standard
series of working solutions into the liquid chromatograph - tandem mass spectrometer, take the
ratio of biotin concentration to biotin internal standard concentration as the x-coordinate, and
the peak area ratio of biotin to biotin internal standard as the y-coordinate to draw a standard
curve of biotin.
6 Expression of Analysis Results
The content of biotin in the specimen is calculated in accordance with Formula (1).
7 Precision
The absolute difference between the results of two independent determinations obtained under
repeatability conditions shall not exceed 15% of the arithmetic mean.
8 Others
When the sampling size is 5.0 g and the constant volume is 50 mL, the detection limit of this
Method is 0.300 g/100 g, and the quantitation limit is 1.00 g/100 g.
9 Principle
Biotin is an essential nutrient for the growth of Lactiplantibacillus plantarum. In the biotin
determination culture medium, the growth of Lactiplantibacillus plantarum is correlated with
the biotin content.
10 Reagents and Materials
Unless it is otherwise specified, the reagents used in this Method are all analytically pure, and
the water is Grade-2 or Grade-1 water specified in GB/T 6682.
10.1 Strain
Lactiplantibacillus plantarum [the former Lactobacillus plantarum] ATCC 8014, or equivalent strain.
10.2 Culture Media
10.2.1 Lactobacillus agar culture medium. see B.1 in Appendix B.
10.3 Reagents
10.3.1 Absolute ethanol (C2H5OH).
10.3.2 Sulfuric acid (H2SO4). 95% ~ 98%.
10.4 Preparation of Reagents
10.4.1 Ethanol solution (50%). measure-take 500 mL of absolute ethanol, add it to water and
reach a constant volume of 1,000 mL.
10.4.2 0Sulfuric acid solution A (3.0 mol/L). measure-take 163.2 mL of sulfuric acid, add it to
water and reach a constant volume of 1,000 mL.
10.4.6 Sodium hydroxide solution B (0.1 mol/L). draw-take 10 mL of sodium hydroxide
solution A (10 mol/L), add water to reach a constant volume of 1,000 mL.
10.4.7 Sterile physiological saline. weigh-take 8.5 g of sodium chloride and dissolve it in 1,000
mL of distilled water, divide it into stoppered test tubes, with 10 mL in each tube. At 121 C,
perform autoclaved sterilization for 15 min.
10.5 Reference Material
Biotin reference material (C10H16N2O3S). CAS. 58-85-5, purity 98%, or a standard substance
certified by the state and awarded a reference material certificate.
10.6 Preparation of Standard Solutions
10.6.1 Biotin standard stock solution (100 g/mL). place the biotin reference material in a
desiccator containing phosphorus pentoxide and dry it overnight. In accordance with purity,
weigh-take it, so that the biotin content is 50 mg (accurate to 0.1 mg), use ethanol solution (50%)
to dissolve it and reach a constant volume of 500 mL.
11 Instruments and Equipment
Except for the conventional sterilization and culture equipment of microbiology laboratory,
other equipment and materials are as follows.
11.1 Analytical balance. with a division value of 0.001 g and 0.1 mg, respectively.
11.2 Centrifuge. 3,000 r/min ~ 5,000 r/min.
11.3 Vortex mixer.
11.4 pH meter. with an accuracy of 0.01.
11.9 Sterile microplate.
11.10 Quantitative filter paper. with a diameter of 90 mm.
11.12 Volumetric flask. with a capacity of 100 mL, 250 mL and 500 mL.
11.13 One-mark pipette. 1 mL, 5 mL and 10 mL.
11.14 Graduated pipette. 5 mL (with a scale of 0.1 mL).
11.15 Glass funnel. with a diameter of 100 mm.
11.16 Conical flask. with a capacity of 250 mL.
11.19 Micropipette. 1,000 L and 200 L.
11.20 Sterile centrifuge tube. 1.5 mL.
11.21 Syringe filter. with an aperture of 0.22 m.
......
Source: Above contents are excerpted from the full-copy PDF -- translated/reviewed by: www.ChineseStandard.net / Wayne Zheng et al.
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