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GB 28301-2012: Food additives -- Riboflavin 5-sodium phosphate
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Food additives -- Riboflavin 5-sodium phosphate
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GB 28301-2012
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Basic data | Standard ID | GB 28301-2012 (GB28301-2012) | | Description (Translated English) | Food additives -- Riboflavin 5-sodium phosphate | | Sector / Industry | National Standard | | Classification of Chinese Standard | C54;X40 | | Classification of International Standard | 67.220.20 | | Word Count Estimation | 10,132 | | Regulation (derived from) | Ministry of Health Bulletin 2012 No. 7 | | Issuing agency(ies) | Ministry of Health of the People's Republic of China | | Summary | This Chinese standard applies to riboflavin as raw material, through phosphorylation, neutralization and purification steps such as the production of food additives riboflavin 5 ' phosphate. | GB 28301-2012: Food additives -- Riboflavin 5-sodium phosphate---This is a DRAFT version for illustration, not a final translation. Full copy of true-PDF in English version (including equations, symbols, images, flow-chart, tables, and figures etc.) will be manually/carefully translated upon your order.
Food additives.Riboflavin 5-sodium phosphate
National Standards of People's Republic of China
National standards for food safety
Food Additives Riboflavin 5'-Phosphate
2012-04-25 release
2012-06-25 Implementation
Issued by the Ministry of Health of the People's Republic of China
National standards for food safety
Food Additives Riboflavin 5'-Phosphate
1 Scope
This standard applies to riboflavin as raw material, phosphorylation, and purification, refining and other steps to produce food additives riboflavin 5'-phosphate.
2 molecular formula and relative molecular mass
2.1 Molecular formula
C17H20N4NaO9P · 2H2O
2.2 Relative molecular mass
514.36 (according to the.2007 International Relative Atomic Quality)
3 technical requirements
3.1 sensory requirements. should be consistent with the provisions of Table 1.
Table 1 sensory requirements
The project requires a test method
Color orange yellow
State crystalline powder
Take appropriate sample in a clean, dry white porcelain dish
Natural light, observe the color and state
3.2 Physical and chemical indicators. should be consistent with the provisions of Table 2.
Table 2 Physical and chemical indicators
Item Index Test Method
Riboflavin (C17H20N4O6) content, w /% 73.0 ~ 79.0 Appendix A, A.3
Specific rotation αm (25 ℃, D)/[(°) · dm2 · kg-1] 37.0 ~ 42.0 Appendix A A.4
pH 5.0 to 6.5 Appendix A, A.5
Dry reduction, w /% ≤ 7.5 Appendix A, A.6
Residue on ignition, w /% ≤ 25.0 Appendix A A.7
Free phosphoric acid, w /% ≤ 1.0 Appendix A A.8
Free riboflavin, w /% ≤ 6.0 Appendix A A.9
Riboflavin diphosphate, w /% ≤ 6.0 Appendix A A.9
Lead (Pb)/(mg/kg) ≤ 2.0 GB 5009.12
Appendix A
Testing method
A.1 General provisions
The reagents and water used in this standard, when not specified in other requirements, refer to the analysis of pure reagents and GB/T 6682-2008 in the provisions of the three
Water level. Standard titration solution used in the test, the standard solution for the determination of impurities, preparations and products, without any other requirements,
GB/T 601, GB/T 602, GB/T 603. The solution used in the test refers to water when it is not specified with the formulation of the solvent
Solution.
A.2 Identification test
A.2.1 Reagents and materials
A.2.1.1 hydrochloric acid solution. take hydrochloric acid 1mL, diluted with water to 3mL.
A.2.1.2 sodium hydroxide solution. take sodium hydroxide 1g, dissolved in water and diluted to 25mL.
A.2.2 Identification method
Weigh about 1.5mg sample, add 100mL water dissolved, this is the sample solution. The sample solution was light yellowish green under transmission light and was strong
Yellow-green fluorescent. Take the sample solution 25mL, add 0.5mL hydrochloric acid solution, fluorescence disappeared; another sample solution 25mL, add 0.1mL hydrogen
Sodium oxide solution, fluorescence disappeared.
A.3 Determination of riboflavin (C17H2ON4O6) content
A.3.1 Reagents and materials
A.3.1.1 Glacial acetic acid.
A.3.1.2 Sodium acetate solution. 14 g/L.
A.3.2 Instruments and equipment
Spectrophotometer.
A.3.3 Analysis steps
Avoid light operation. Weigh about 100mg sample, accurate to 0.0001g, placed in 500mL volumetric flask, add 1mL glacial acetic acid and
75mL water. After the sample is dissolved, dilute to the mark with water and shake it. Accurately measure 10mL, placed in 100mL volumetric flask, add
7mL sodium acetate solution, diluted with water to the mark, shake. To the water as a blank, with a 1cm caliber cup, measured at 444nm wavelength
The absorbance of the sample solution. The absorption coefficient (E1 m) of riboflavin is calculated as 328.
A.3.4 Calculation of results
Riboflavin content of riboflavin to the mass fraction w0, the value expressed in%, according to formula (A.1) calculation.
w0 = 50 × A328 × m × (1 - w1) ×
100% (A.1)
Where.
50 --- dilution factor;
A - the absorbance of the sample solution;
m - the value of the sample quality, in grams (g);
w1 --- Measured sample dry reduction value,%.
The experimental results are based on the arithmetic mean of the parallel measurement results. The absolute difference between the two independent determinations obtained under repeatability conditions
The ratio of the value to the arithmetic mean is not more than 1.0%.
A.4 Determination of specific rotation
A.4.1 Reagents and materials
Hydrochloric acid solution. take hydrochloric acid 27.0mL, diluted with water, constant volume to 50mL.
A.4.2 Instruments and equipment
Polarimeter, accuracy ± 0.01 °.
A.4.3 Analysis steps
Avoid light operation. Weigh 0.75g (in anhydrous terms) sample, accurate to 0.0001g, placed in 50mL volumetric flask, add hydrochloric acid solution solution
Solution and dilution to the scale, in 15min, 25 ℃ ± 1 ℃ conditions with a polarimeter to determine the specific rotation.
A.4.4 Calculation of results
The value of the specific rotation αm (25 ° C, D) is expressed by (°) · dm2 · kg-1, calculated according to the formula (A.2)
αm (25 ° C, D) = α lρα
(A.2)
Where.
α - the measured angle of rotation, in degrees (°);
l - the length of the rotating tube in dm (dm);
ρα --- The concentration of the sample solution in grams per milliliter (g/mL).
The experimental results are based on the arithmetic mean of the parallel measurement results. The absolute difference between the two independent determinations obtained under repeatability conditions
The ratio of the value to the arithmetic mean is not more than 1.0%.
A.5 Determination of pH
A.5.1 Instruments and equipment
Acidity meter, accuracy ± 0.01.
A.5.2 Analysis steps
Weigh 1.0g sample, accurate to 0.0001g, placed in.200mL clean and dry beaker, add 100mL of water dissolved at 20 ℃
The pH was measured with an acidity meter.
A.5.3 Calculation of results
The experimental results are based on the arithmetic mean of the parallel measurement results. The absolute difference between the two independent determinations obtained under repeatability conditions
The ratio of the value to the arithmetic mean is not more than 1.0%.
A.6 Determination of dry reduction
A.6.1 Instruments and equipment
Constant temperature vacuum oven.
A.6.2 Analysis steps
Weigh about 2g sample, accurate to 0.0001g, placed in the constant weight to the weighing bottle, at 100 ℃ ± 1 ℃, 0.01MPa
Dried for 5 h, then placed in a desiccator to cool to room temperature and weighed.
A.6.3 Calculation of results
The drying loss is expressed in terms of mass fraction w1 and the value in%, calculated according to formula (A.3)
w1 = m1 - m2m3 ×
100% (A.3)
Where.
m1 --- the value of the sample before the dry and the quality of the weighing bottle, in grams (g);
m2 --- the value of the sample after drying and the mass of the weighing bottle, in grams (g);
m3 --- the value of the sample quality before drying, in grams (g).
The experimental results are based on the arithmetic mean of the parallel measurement results. The absolute difference between the two independent determinations obtained under repeatability conditions
The ratio of the value to the arithmetic mean is not more than 1.0%.
A.7 Determination of burning residue
A.7.1 Instruments and equipment
High temperature furnace.
A.7.2 Analysis steps
Weigh about 1g sample, accurate to 0.001g, placed in the 600 ℃ ~ 700 ℃ was burning to constant weight of the crucible, slowly heated to the same
The product is fully carbonized. The carbonized sample was cooled, the residue was wetted with 0.5 mL of sulfuric acid, heated to sulfuric acid to escape,
600 ℃ ~ 700 ℃ high temperature furnace burning to constant weight.
A.7.3 Calculation of results
The ignition residue is expressed in terms of mass fraction w2 and the value in%, calculated according to formula (A.4)
w2 = m4-m5m6 x
100% (A.4)
Where.
m4 - the value of the mass of the residue and the empty crucible, in grams (g);
m5 --- the value of the quality of the empty crucible, in grams (g);
m6 --- the value of the sample quality, in grams (g).
The experimental results are based on the arithmetic mean of the parallel measurement results. The absolute difference between the two independent determinations obtained under repeatability conditions
The ratio of the value to the arithmetic mean is not more than 1.0%.
A.8 Determination of free phosphoric acid
A.8.1 Reagents and materials
A.8.1.1 Standard Phosphate Solution. 0.22 g of potassium dihydrogen phosphate was dried at 105 ° C for 2 h, placed in a 1000 mL volumetric flask,
Add the appropriate amount of water to dissolve, and diluted to the mark, shake, temporary use and then diluted 5 times.
A.8.1.2 Ammonium molybdate [(NH4) 6Mo7O24 · 4H2O] solution. 70 g/L.
A.8.1.3 sulfuric acid solution. 3.75 mol/L.
A.8.1.4 Acid Ammonium Molybdate Solution. 25 mL of ammonium molybdate solution was diluted with water to.200 mL and then slowly added 25 mL of sulfuric acid solution
Liquid, shake.
A.8.1.5 ferrous sulfate solution. accurately weighed 10.0g of ferrous sulfate, placed in 100mL volumetric flask, add the appropriate amount of water and 2mL
Sulfuric acid solution dissolved, and diluted to the mark, shake.
A.8.2 Instruments and equipment
UV-visible spectrophotometer.
A.8.3 Analysis steps
A.8.3.1 Preparation of sample solution
Accurately weighed 300.0mg samples (according to anhydrous items), placed in 100mL volumetric flask, add the appropriate amount of water dissolved and diluted to the mark,
Shake well.
A.8.3.2 Determination
Accurately measure the standard phosphate solution and sample solution of 10.0mL, were placed in 50mL Erlenmeyer flask, and then to the conical flask
By adding 10.0 mL of acidic molybdate solution and 5 mL of ferrous sulfate solution, mixing in 10.0 mL of water, 10.0 mL of acidic molybdate
Liquid and 5 mL of ferrous sulfate solution as a blank, with an optical path of 1cm cuvette, measured at 700nm wavelength of the cone
The absorbance of the solution in the bottle. The absorbance of the sample solution should not be greater than the absorbance of the standard solution.
A.8.4 Calculation of results
The experimental results are based on the arithmetic mean of the parallel measurement results. The absolute difference between the two independent determinations obtained under repeatability conditions
The ratio of the value to the arithmetic mean is not more than 1.5%.
A.9 Determination of free riboflavin and riboflavin diphosphate
A.9.1 Reagents and materials
A.9.1.1 KH2PO4 solution. 0.054 mol/L.
A.9.1.2 Methanol. Chromatographic Purification.
A.9.1.3 riboflavin standard. riboflavin content of not less than 99.0% (mass fraction).
A.9.1.4 riboflavin 5'-sodium phosphate standard. riboflavin content of not less than 74.0% (mass fraction).
A.9.2 Instruments and equipment
High performance liquid chromatograph with fluorescence detector (excitation wavelength 440nm; emission wavelength 470nm).
A.9.3 Reference chromatographic conditions
A.9.3.1 Column. C18 column, 3.9 mm x 300 mm, or other equivalent column.
A.9.3.2 mobile phase. according to KH2PO4 solution. methanol = 85.15 (volume ratio) preparation, mixing evenly, with 0.45μm filter over
Filter, ultrasonic degassing standby.
A.9.3.3 Column temperature. room temperature.
A.9.3.4 Mobile phase flow rate. 2.0 mL/min.
A.9.3.5 Injection volume. 100 μL.
A.9.4 Analysis steps
A.9.4.1 Preparation of System Adaptive Solutions
Dissolved 100mg riboflavin 5'-sodium phosphate standard in water, the mass concentration of 2.0mg/mL, adding an equal volume of mobile phase,
Mix well. Absorb 8mL, with the mobile phase diluted to 50mL, mix spare.
A.9.4.2 Preparation of standard solutions
Accurately weighed 60.0mg riboflavin standard in 250mL volumetric flask, carefully add 1mL hydrochloric acid dissolved, diluted with water to the mark,
Mix well. Pipette with 4mL of riboflavin solution in a 100mL volumetric flask, diluted with mobile phase to the scale, mix well.
A.9.4.3 Preparation of sample solution
Accurately weighed 100.0mg sample in 100mL volumetric flask, add 50mL water dissolved, diluted with mobile phase to the scale, mix. Move with
The pipette draws 8 mL of the solution in a 50 mL volumetric flask and is diluted to the scale with the mobile phase and shaken back.
A.9.4.4 System Adaptability Requirements
Take appropriate system into the solution into the chromatograph for chromatographic analysis. Riboflavin 4'-monophosphate and riboflavin 5'-monophosphate
And the relative standard deviation of the repeated injection of riboflavin 5'-monophosphate was not more than 1.5%.
The retention time of riboflavin 5'-monophosphate is about 20min ~ 25min. The approximate relative retention times for each component are given in Table A.1
As shown.
Table A.1 Approximate relative retention times for each component
Component relative retention time
Riboflavin 3 ', 4'-diphosphate 0.23
Riboflavin 3 ', 5'-diphosphate 0.39
Riboflavin 4 ', 5'-diphosphate 0.58
Riboflavin 3'-monophosphate 0.70
Riboflavin 4'-monophosphate 0.87
Riboflavin 5'-monophosphate 1.00
Riboflavin 1.63
A.9.4.5 Determination
An equal volume (about 100 μL) of standard solution, sample solution and system-compatible solution were separately injected into a chromatograph for chromatographic analysis,
Record the chromatograms in the standard solution and the peak response values in the sample solution chromatogram. By comparing the system to the adaptive solution chromatogram
The peak time of the chromatographic peaks is determined by the peak of the sample solution chromatogram.
A.9.5 Calculation of results
Free riboflavin content of free riboflavin by the mass fraction w3, the value expressed in%, according to formula (A.5).
w3 = 625 × ws × PFPS
(A.5)
Where.
625 --- the dilution factor of the sample;
ws - the concentration of riboflavin in the standard solution in milligrams per milliliter (mg/mL);
PF - Peak response of riboflavin in sample solution chromatogram;
PS - peak response of riboflavin in standard solution chromatograms.
Riboflavin diphosphate content of riboflavin diphosphate mass fraction w4, the value in%, calculated according to formula (A.6).
w4 = 625 × ws × PDPS
(A.6)
Where.
625 --- the dilution factor of the sample;
ws - the concentration of riboflavin in the standard solution in milligrams per milliliter (mg/mL);
PS --- peak response of riboflavin in standard solution chromatogram;
PD - the total response of the three riboflavin diphosphate peaks in the sample solution chromatogram.
The experimental results are based on the arithmetic mean of the parallel measurement results. The absolute difference between the two independent determinations obtained under repeatability conditions
The ratio of the value to the arithmetic mean is not more than 1.5%.
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