GB 28317-2012Food additives -- Indigo (English PDF)

1 Scope

This standard applies to indigo as raw material, the sulfonation, refined and made of food additives indigo.

2 Chemical name, molecular formula, structural formula, relative molecular mass

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 dark purple to dark purple brown State powder or particles Take the appropriate amount of sample placed in a clean, dry white porcelain dish, in the natural light, Observe its 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 Indigo content, w /% >= 85.0 Appendix A, A.3 Dry reduction, chloride (in terms of NaCl) and sulfate (In terms of Na2SO4), w /% <= 15.0 Appendix A A.4 Water insoluble matter, w /% <= 0.20 Appendix A, A.5 Sub-dye, w /% <= 1.0 Appendix A A.6 Total arsenic (in As)/(mg/kg) <= 1 Appendix A A.7 Lead (Pb)/(mg/kg) <= 10 Appendix A A.8 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 Sulfuric acid solution. 1 20.

A.2.1.2 Sodium hydroxide solution. 2.5 g/L.

A.2.1.3 Ammonium acetate solution. 1.5 g/L.

A.2.2 Instruments and equipment A.2.2.1 Spectrophotometer.

A.2.2.2 Cuvette. 10mm.

A.2.3 Analysis steps A.2.3.1 Color response A.2.3.1.1 Weigh about 0.1g sample, add 10mL sulfuric acid solution, shake from time to time, the solution was dark purple, after cooling, take 2 to 3 drops, add 5mL water, the solution was blue and purple.

A.2.3.1.2 measure the blue-purple indigo-sulfuric acid solution 5mL, add 1mL sodium hydroxide solution, slightly shaking, was yellow-green.

A.2.3.2 Maximum absorption wavelength Weigh about 0.1g sample, add 100mL water, shake from time to time, dissolved into purple blue solution. Measure the solution 1mL, add ammonium acetate solution with to 100 mL. The maximum absorption wavelength of this solution is 612 nm ± 2 nm.

A.3 Determination of indigo content A.3.1 Titanium trichloride titration (Arbitration) A.3.1.1 Method Summary in the acidic medium, the amino group in the dye structure is reduced to the amino compound by titanium trichloride, and the solution is titrated according to the standard of titanium trichloride Consumption, calculate its content.

A.3.1.2 Reagents and materials A.3.1.2.1 Sodium hydrogen tartrate.

A.3.1.2.2 Titanium trichloride standard titration solution. c (TiCl3) = 0.1mol/L (now with the use of the preparation method see Appendix B).

A.3.1.2.3 Carbon dioxide.

A.3.1.3 Instruments and equipment See Figure A.1.

A - conical flask (500mL);

B - brown burette (50mL);

C --- black paper with black paper bottle (2000mL);

D - a mixture of ammonium carbonate and ferrous sulfate (5000 mL);

E --- piston

F - empty bottle;

G --- water washing gas bottle.

Figure A.1 Titanium trichloride titration device

A.3.1.4 Analysis steps Approximately 0.5 g of the sample was weighed to 0.0001 g, placed in a 500 mL Erlenmeyer flask, dissolved in 50 mL of freshly boiled and cooled to room temperature Water, add 15 g of sodium hydrogen tartrate and 150 mL of freshly boiled and cooled to room temperature, dissolve and dissolve, install the instrument as shown in Figure A.1, Liquid under the surface of carbon dioxide at the same time, heated to boiling, and titration with titanium titanate titration to its inherent color disappeared as the end.

A.3.1.5 Calculation of results the indigo content is expressed in terms of mass fraction w1 and the value in%, calculated according to formula (A.1) w1 = (V/1000) x c (M/2) m1 × 100% (A.1) where.

V --- titration sample consumption of titanium trichloride standard titration solution volume of the exact value in milliliters (mL);

1000 --- volume conversion factor;

c - Titanium trichloride standard titration solution concentration of the exact value in moles per liter (mol/L);

M - the molar mass of indigo in grams per mole (g/mol) [M (C16H8N2Na2O8S2) = 466.36];

2 --- concentration conversion factor;

m1 --- the value of the sample quality, in grams (g).

The result of the calculation indicates one bit after the decimal point.

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 value should not exceed 1.0% of the arithmetic mean.

A.3.2 Spectrophotometric colorimetric method A.3.2.1 Methodological Summary the sample and the known content of indigo standard samples were dissolved in water, respectively, at the maximum absorption wavelength, respectively, measured its absorbance value, and then Calculate its content.

A.3.2.2 Reagents and materials A.3.2.2.1 Ammonium acetate solution. 1.5 g/L.

A.3.2.2.2 Indigo standard sample. content >=85.0% (mass fraction, measured in A.3.1).

A.3.2.3 Instruments and equipment A.3.2.3.1 Spectrophotometer.

A.3.2.3.2 Cuvette. 10mm.

A.3.2.4 Analysis steps A.3.2.4.1 Preparation of indigo standard sample solution Weigh about 0.25g indigo standard sample, accurate to 0.0001g, dissolved in water, into the 1000mL volumetric flask, diluted with water to the scale, shake. Accurately absorb 10mL, into the 500mL volumetric flask, add ammonium acetate solution diluted to the mark, shake, spare.

A.3.2.4.2 Preparation of indigo sample solution Weighing and handling methods are prepared with indigo standard sample solutions.

A.3.2.4.3 Determination the indigo standard sample solution and indigo sample solution were placed in 10mm cuvette, respectively, with the maximum absorption wavelength (612nm ± 2nm) with a spectrophotometer to determine the respective absorbance values to ammonium acetate solution as the reference solution.

A.3.2.5 Calculation of results the indigo content is expressed in terms of mass fraction w2 and the value in%, calculated according to formula (A.2) w2 = A1 x m0A0 × m × w0 (A.2) where.

A1 --- Indigo sample solution absorbance value;

m0 --- Indigo standard sample quality value in grams (g);

A0 --- Indigo standard sample solution absorbance value;

m - the mass of the sample, in grams (g);

w0 --- indigo standard sample content of the value,%.

The result of the calculation indicates one bit after the decimal point.

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 value should not exceed 1.0% of the arithmetic mean.

A.4 Determination of dry reduction, total amount of chloride (in terms of NaCl) and sulfate (calculated as Na2SO4) A.4.1 Determination of dry reduction A.4.1.1 Analysis steps Weigh about 2g sample, accurate to 0.0001g, placed in 135 °C ± 2 °C constant temperature oven constant volume of the bottle, the 135 °C ± 2 °C constant temperature oven bake to constant volume.

A.4.1.2 Calculation of results the drying loss is expressed in terms of mass fraction w3 and the value in%, calculated according to formula (A.3) w3 = m2-m3m2 × 100% (A.3) where.

m2 --- the value of the sample before drying, in grams (g);

m3 --- the value of the sample after drying to constant volume, in grams (g).

The result of the calculation indicates one bit after the decimal point.

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 value should not exceed 0.2% of the arithmetic mean.

A.4.2 Determination of chloride (in terms of NaCl) A.4.2.1 Reagents and materials A.4.2.1.1 Nitrobenzene.

A.4.2.1.2 Nitric acid solution. 1 1.

A.4.2.1.3 Silver nitrate solution. c (AgNO3) = 0.1 mol/L.

A.4.2.1.4 ammonium ferric sulfate test solution. Weigh 14g ammonium ferric sulfate, dissolved in 100mL water, filtered, add nitric acid 10mL, stored in brown Bottle.

A.4.2.1.5 Ammonium thiocyanate standard titration solution. c (NH4SCN) = 0.1 mol/L.

A.4.2.1.6 Activated carbon.

A.4.2.2 Analysis steps A.4.2.2.1 Preparation of sample solution Weigh about 2g sample, accurate to 0.0001g, dissolved in 150mL water, add about 15g activated carbon, gently boil 2min ~ 3min.

Cooled to room temperature, add nitric acid solution 1mL, continue to shake evenly, placed 30min (during which time shaking). Filter with dry filter paper. Such as the filtrate is colored, then add activated carbon 5g, from time to time shaking under 1h, and then dry filter paper filter (such as still color is replaced by activated carbon repeated exercise Make the filtrate colorless). Each time with water 10mL wash activated carbon three times, the filtrate combined to move to.200mL volumetric flask, add water to the mark, shake.

For the determination of chloride and sulfate content.

A.4.2.2.2 Determination Remove the 50 mL sample solution, place in a 500 mL Erlenmeyer flask, add 2 mL of nitric acid solution and 10 mL of silver nitrate solution (chloride containing the amount of time to add more) and 5mL nitrobenzene, shake to silver chloride condensation, add 1mL ammonium ferric sulfate test solution, with ammonium thiocyanate standard Titration solution titrated excess silver nitrate to the end and hold for 1 min, while the same method to do a blank test.

A.4.2.3 Calculation of results Chloride (in terms of NaCl) in terms of mass fraction w4, expressed in%, calculated according to formula (A.4).

[(V1-V0)/1000] × c1 × M1 m4 × (50/200) × 100% (A.4) where.

V1 --- titration blank solution consumption of ammonium thiocyanate standard titration solution volume value in milliliters (mL);

V0 --- titration sample solution consumption of ammonium thiocyanate standard titration solution volume value in milliliters (mL);

1000 --- volume conversion factor;

c1 --- ammonium thiocyanate standard titration solution concentration value, the unit is the mole per liter (mol/L);

M1 - sodium chloride molar mass in grams per mole (g/mol) (M (NaCl) = 58.4);

m4 --- the value of the sample quality, in grams (g);

50/200 --- dilution factor.

The result of the calculation indicates one bit after the decimal point.

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 value should not exceed 0.3% of the arithmetic mean.

A.4.3 Determination of Sulfate (calculated as Na2SO4) A.4.3.1 Reagents and materials A.4.3.1.1 Sodium hydroxide solution. 0.2 g/L.

A.4.3.1.2 Hydrochloric acid solution. 1 99.

A.4.3.1.3 barium chloride standard titration solution. c (1/2BaCl2) = 0.1mol/L (see Appendix C for the preparation method).

A.4.3.1.4 phenolphthalein indicator solution. 10g/L.

A.4.3.1.5 Sodium Roseate Indicator. Weigh 0.1 g of sodium rosin sodium and dissolve in 10 mL of water (now available).

A.4.3.2 Analysis steps Absorb 25mL sample solution (A.4.2.2.1), placed in 250mL Erlenmeyer flask, add 1 drop of phenolphthalein indicator solution, add sodium hydroxide solution Liquid was pink, and then dropping hydrochloric acid solution to pink disappeared, shake, dissolved in the constant shaking with barium chloride standard titration solution titration, with sodium alanate indicator liquid for the external indicator solution, the reaction solution and indicator liquid on the filter paper at the intersection of rose red spots and keep 2min Fade for the end. While the same method to do blank test.

A.4.3.3 Calculation of results the content of sulfate (in terms of Na2SO4) is expressed in terms of mass fraction w5, expressed in%, calculated according to formula (A.5) w5 = [(V2-V3)/1000] x2 (M2/2) m5 × (25/200) × 100% (A.5) where.

V2 --- titration blank solution consumption of barium chloride standard titration solution volume value in milliliters (mL);

V3 --- titration sample solution consumption of barium chloride standard titration solution volume value in milliliters (mL);

1000 --- volume conversion factor;

c2 --- barium chloride standard titration solution concentration value, the unit is mol per liter (mol/L);

M2 --- sodium sulfate molar mass in grams per mole (g/mol) [M (Na2SO4) = 142];

2 --- concentration conversion factor;

m5 --- the mass of the sample, in grams (g);

25/200 --- dilution factor.

The result of the calculation indicates one bit after the decimal point.

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 value should not exceed 0.2% of the arithmetic mean.

A.4.4 Calculation of dry reduction, chloride (calculated as NaCl) and sulfate (calculated as Na2SO4) the total amount of dry reduction, chloride (in terms of NaCl) and sulfate (calculated as Na2SO4) is expressed in terms of mass fraction w6, expressed as% Calculated according to formula (A.6).

w6 = w3 w4 w5 (A.6)

Where.

w3 --- dry reduction,%;

w4 --- chloride (in terms of NaCl) content,%;

w5 --- sulfate (in Na2SO4 dollars) content,%.

The result of the calculation indicates one bit after the decimal point.

A.5 Determination of water insoluble matter A.5.1 Instruments and equipment A.5.1.1 Glass sand core crucible. G4, pore size of 5µm ~ 15µm.

A.5.1.2 Constant temperature oven.

A.5.2 Analysis steps Weigh about 3g sample, accurate to 0.001g, placed in 500mL beaker, add 50 °C ~ 60 °C water 250mL, so that dissolved, with a glass sand core crucible which has been baked at 135 ° C ± 2 ° C until constant and washed with hot water until the washing solution is colorless, at 135 ° C ± 2 ° C Constant oven in the oven.

A.5.3 Calculation of results Water insoluble matter in mass fraction w7, the value in%, according to formula (A.7).

w7 = m6m7 ×

100% (A.7)

Where.

m6 --- the value of the water insoluble matter after drying, in grams (g);

m7 --- the value of the sample quality, in grams (g).

The result of the calculation indicates one bit after the decimal point.

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 value should not exceed 0.2% of the arithmetic mean.

A.6 Determination of secondary dyes A.6.1 Methodological summary the components were separated, eluted and then quantitated by spectrophotometry.

A.6.2 Reagents and materials A.6.2.1 2-butanone.

A.6.2.2 Acetone.

A.6.2.3 Acetone solution. 1 1.

A.6.2.4 Hydrochloric acid solution. 1.200.

A.6.3 Instruments and equipment A.6.3.1 Spectrophotometer.

A.6.3.2 chromatography filter paper. No. 1 medium speed, 150mm × 250mm.

A.6.3.3 chromatography cylinder. phi240mm × 300mm.

A.6.3.4 Micro Sampler. 100 µL.

A.6.3.5 Nessler's colorimetric tube. 50mL with a glass grinder.

A.6.3.6 glass sand core funnel. G3, aperture of 15µm ~ 40µm.

A.6.3.7 50mm cuvette.

A.6.3.8 10mm cuvette.

A.6.4 Analysis steps A.6.4.1 Paper chromatography conditions A.6.4.1.1 Expanding agent. 2-butanone acetone water = 7 3 3.

A.6.4.1.2 Temperature. 20 ° C to 25 ° C.

A.6.4.2 Preparation of sample solution Weigh about 1g sample, accurate to 0.001g, placed in a beaker, add the appropriate amount of water dissolved, into the 100mL volumetric flask, diluted to Scale, shake back, the sample solution concentration of 1%.

A.6.4.3 Preparation of sample eluate Use a micro-injector to draw 100 µL of the sample solution, evenly on a bottom line of 25 mm from the bottom of the filter paper, its width in the filter paper does not exceed 5mm, length of 130mm, with a hair dryer to dry. Place the filter paper in a pre-prepared deployment the bottom of the filter paper is immersed in the developer surface under the surface of 10mm, to be developed before the agent rose to 150mm or until the sub-dyeing Material separation satisfaction. Remove the filter paper, dry with cold air.

With the blank filter paper under the same conditions to expand, the blank filter should be used with the above steps to start with the filter paper in the same filter on the adjacent parts Cut.

The chromatogram of the secondary dye paper is shown in Figure A.2.

Figure A.2 Schematic diagram of secondary dye chromatography

After the expansion of the various sub-dyes (except for isomers) and in the blank filter paper and the corresponding parts of the corresponding part of the filter paper by the same large Small cut, and cut into about 5mm × 15mm thin strips, were placed in 50mL Na colorimetric tube, accurately add 5mL acetone solution, Shake 3min ~ 5min, and then accurately add 20mL hydrochloric acid solution, shake, and then in the G3 glass sand core funnel naturally Filter, the filtrate should be clear, no suspended solids. Respectively, to obtain the secondary dye and blank wash solution. At the maximum absorption wavelength of the respective sub-dyes 50mm cuvette, the respective dyes of the eluent on the spectrophotometer to determine the respective absorbance value.

When the absorbance was measured on a spectrophotometer, a 5 ml solution of acetone and 20 mL of hydrochloric acid was used as a reference solution.

A.6.4.4 Preparation of standard solutions Draw 2mL sample solution into the 100mL volumetric flask, dilute to the mark, shake. The solution is a standard solution.

A.6.4.5 Preparation of standard elution solution Use a micro-injector to draw 100 µL of standard solution, evenly point on the bottom of the filter paper 25mm on a baseline, with a hair dryer blowing dry. The filter paper into a pre-prepared with a good agent into the chromatography cylinder to start, before the agent to be raised 40mm, take out with cold Dry the cut and cut all the expanded dyes and perform the extraction as described in A.6.4.3 to obtain a standard wash solution. With a 10mm ratio the colorimeter measures the absorbance value at the maximum absorption wavelength.

At the same time with the blank filter paper under the same conditions to expand, according to the same method after the test wash the absorbance value.

A.6.4.6 Calculation of results the content of the secondary dye is expressed in terms of mass fraction w8 and the value in%, calculated according to the formula (A.8) w8 = [(A1-b1) (An-bn)]/5 (AS-bS) (100/2) x ws (A.8) where.

A1, and An - absorbance values measured by 50 mm optical path length of each sub-dye wash solution;

b1, bn - the absorbance values measured by the 50 mm optical path length of each of the sub-dye control blank wash solutions;

AS --- standard sample washout solution to 10mm optical path length measured absorbance value;

bS --- standard sample control blank wash solution to 10mm optical path length measured absorbance value;

5 --- converted into a 10mm optical path length ratio;

100/2 --- standard sample elution solution converted into 1% sample solution ratio;

ws - the mass fraction of indigo in the sample,%.

The result of the calculation indicates one bit after the decimal point.

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 value should not exceed 0.2% of the arithmetic mean.

A.7 Determination of arsenic A.7.1 Methodological Summary Indigo was digested by wet method and prepared as sample solution, and the content of arsenic was determined by atomic absorption spectrometry.

A.7.2 Reagents and materials A.7.2.1 Nitric acid.

A.7.2.2 Sulfuric acid solution. 1 1.

A.7.2.3 Nitric acid - Perchloric acid mixed solution. 3 1.

A.7.2.4 arsenic (As) standard solution. according to GB/T 602 preparation and calibration, and then according to the use of equipment required dilution prepared with arsenic the corresponding concentration of the three standard solutions.

A.7.2.5 Sodium hydroxide solution. 1 g/L.

A.7.2.6 Sodium borohydride solution. 8 g/L (solvent 1 g/L sodium hydroxide solution).

A.7.2.7 Hydrochloric acid solution. 1 10.

A.7.2.8 Potassium iodide solution..200 g/L.

A.7.3 Instruments and equipment Atomic Absorption Spectrometer, the following conditions should be met.

a) Instrument reference conditions. arsenic hollow cathode lamp analysis line wavelength. 193.7nm; slit. 0.5nm ~ 1.0nm; lamp current. 6mA ~ 10mA;

b) Carrier gas flow rate. argon 250 mL/min;

c) Atomizer temperature. 900 ° C.

A.7.4 Analysis steps A.7.4.1 Sample digestion Approximately 1.0 g of the sample was weighed, accurate to 0.001 g, placed in a 250 mL conical or round bottom flask with 10 mL to 15 mL of nitric acid and 2mL sulfuric acid solution, shake and heat with a small fire out of the nitrogen dioxide gas, the solution becomes brown, stop heating, let cool after adding 5mL of nitrate Acid - perchloric acid mixture, strong fire to the solution to the transparent colorless or slightly yellow, such as still opaque, put cold and then add 5mL nitric acid - perchloric acid Mix the solution, continue to heat the solution to clarify the colorless or yellowish and produce white smoke (to avoid burning carbonization phenomenon), stop heating, let cool Add water 5mL heated to boiling, remove the residual nitric acid - perchloric acid (if necessary, add water to boil again), continue to heat to the occurrence of white smoke, keep 10min, put cold into the 100mL volumetric flask (if the solution appears turbidity, precipitation or mechanical impurities to be filtered), diluted with hydrochloric acid solution volume.

At the same time, the blank solution was prepared in the same manner.

A.7.4.2 Determination Measure 25mL of the digested sample solution to 50mL volumetric flask, add 5mL potassium iodide solution, diluted with hydrochloric acid solution volume, shake Uniform, static 15min.

The blank test solution was prepared in the same manner as a blank solution.

Open the instrument, wait until the instrument and the arsenic hollow cathode lamp are fully warmed, and after the baseline is stabilized, the sodium borohydride solution is used as the hydride reduction agent to Standard blank, standard solution, sample blank test solution and the order of the sample solution, according to the computer instructions were injected. The computer is automatically finished after the test Generate the working curve and subtract the arsenic concentration from the sample solution after the sample blank, and input the sample information (name, sample volume, dilution volume, etc.) Calculate the arsenic content in the sample.

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 Value of not more than 0.1mg/kg.

A.8 Determination of lead A.8.1 Methodological Summary Indigo was digested by wet method and prepared as sample solution, and the content of lead was determined by atomic absorption spectrometry.

A.8.2 Reagents and materials A.8.2.1 lead (Pb) standard solution. according to GB/T 602 preparation and calibration, and then according to the requirements of the instrument used to prepare diluted with lead the corresponding concentration of the three standard solutions.

A.8.2.2 Sodium hydroxide solution. 1 g/L.

A.8.2.3 sodium borohydride solution. 8g/L (solvent 1g/L sodium hydroxide solution).

A.8.2.4 Hydrochloric acid solution. 1 10.

A.8.3 Instruments and equipment Atomic Absorption Spectrometer, the instrument reference conditions should meet the requirements of Flame ...