GB/T 5009.35-2003 Determination of synthetic colorants in food

The method for the determination of synthetic colorants in food has been determined. This standard is applicable to the determination of synthetic colorants in food. The detection limits of this method are: new red 5ng, lemon yellow 4ng, amaranth red 6ng, cochineal red 8ng, sunset yellow 7ng, erythrosine 18ng, brilliant blue 26ng. When the injection volume is equivalent to 0.025g, the detection concentrations are 0.2mg/g; 0.16mg/g; 0.24mg/g; 0.32mg/g; 0.28mg/g; 0.72mg/g; 1.04mg/g. GB/T 5009.35-2003 Determination of synthetic colorants in food GB/T5009.35-2003 standard download decompression password: www.bzxz.net

ICs: 67.040
National Standard of the People's Republic of China
CB/T 5009. 35--2003
Replacement of GR/E005.35-16
Determination of synthetic colorants in food
Determination of svnthetic cnlnur in Fds2003-08-11PromulgatedbzxZ.net
Ministry of Health of the People's Republic of China
Standardization Administration of the People's Republic of China
Implemented on 2004-01-01
GB/T5009.35-2003
This standard replaces CB/T5009.351996. Determination of colorants in food containing packaging materials 3. Compared with GH/T5009.35-1SSG ±The following are the main rectifications: the Chinese name of the standard has been changed. The Chinese name of the standard has been changed to "Determination of synthetic colorants in food 3: According to the (H/20CC1.1-2Uu1 standard writing rules Part 4: Chemical analysis methods 3) of the original standard structure; the oscillographic addition method has been added as the first version, and the Ministry of Health of the People's Republic of China has put forward this standard. The standard is drafted by the Tianjin Food Hygiene Supervision and Inspection Institute, the Ningxia Food Dust Supervision and Inspection Institute, the Ningxia Hui Autonomous Region Health and Epidemic Prevention Station, and the Xi'an Health and Epidemic Prevention Station. This standard is first issued in 193 and revised for the first time in 1996. This is the second revision. This standard specifies the determination of synthetic colorants in food. Determination of synthetic ions. This standard applies to the determination of synthetic ions in food, GB/T5009.35-2003. The limits of this standard are: new red 5g, lemon yellow 2g, broad red, carmine Ag, sunset yellow 7g, east 18g, and complete 26ng. When the intake temperature is 0.02sg, the reported pressures are n2mg/kg: 0.16mg/kg0.24mg/kg: C.32nag/kg51. 28 mp/hg10. 7? mx/kg;1. t4 mg/kg.The first method is commercial liquid chromatography
2 Principle
1 The coloring agent in food is condensed by cumulative adsorption method to make a drop solution, injected into commercial liquid chromatography instrument: lightly beat the package spectrum, and quantified according to the retention time and the barrier area ratio. 3 Reagents
3.2 Hydrochloric acid.
3.3 Acetic acid.
3. 4 Medium: Filter through 0. Mm full membrane
Tight powder (ratio 6): Pass through 200 pieces of sieve: 3.6 Acetic acid environment drop solution (0.02mol/1.), take 1, i4R ethanol section, water 1000mL rate dance, pass through 0.45Am membrane.3.73
Study water: Quantitative ammonia? ml., add water to 1nL, mix well. 3.8
water-acetic acid solution (0.32ma1/1.) weigh 0.ml of ammonia, acetic acid solution (0.02ol/L to 100umI, mix well.
3. 9 methanol-formic acid [i-4] 60 nL, formic acid 4U ml., cage, 3.1 C treatment acid solution: weigh 20g of sodium sulfoxide (CTL, ·H 60>. Add water to 100m, float and filter without wood ammonia water (712+1) solution: ethanol 7Gm1. Hydrogenated water 20ml. 101ml. Very column 3. 11
3.12 Add 5mL of trioctylamine to 100L of butyl alcohol and mix. 3.13 Add 1% ethyl acetate to 1% ethanol.
3.14 Sulfur absorption line (2≤/L).
3,15 Add water to control the pH value of E6. 3.15, Standard colorant: Accurately weigh 0.00% of lemon yellow, sunset yellow, amaranth red, lipstick red, new red, red, light vegetable, sugar, each in a 1mL bottle, H6 aqueous emulsion, and 1% bismuth. mg/ml.)
3.17 Synthetic colorant standard is the same as F1 above (or 3.16) diluted 20 times with water, passed through 0.45μm, and then matched with the appropriate synthetic colorant.
4 Instrument
High-efficiency micro-phase elimination, with ultraviolet detector, 2F1rm wavelength. 277
GB/T5009.35·-2J03|| tt||5 Analysis steps
5.1 Test mixture
5.1.1 Orange juice, fruit water, fruit dew, etc.: weigh 3U.0~4V.: put 10mL into a small cup and heat to remove oxidation.
5.1.2 Prepare wine: weigh 20.0g~-40.Cg and put 10CmL of sieve, heat to remove oxidation. 5.1.3 Mix well, weigh 5.20R~10.(0 For the crushed sample, place it in a 200mL small cup, add 3U of water, and warm it to a temperature of 1000 °C. If the concentration of the sample is higher than the commercial value, use a flame to melt the sample until it reaches the value of 2I. 5.1.4 Chocolate and other color tree clothing products: Take 1.M~1.) and place it in a 1mT small cup, wash with water, and stop the sample without any inclusions. Combine the washed samples and make the sample solution. 5.2 Color capsule extraction||tt| |5.2.1 Polymerization deamination method: the liquid is not removed by acid wave adjustment! 5. Heat to 6U℃. 6 Polymerization is cut into small pieces, poured into the sample drop, burned, extracted with G3 melting funnel, washed with iodine = 1-5 times, then washed with alcohol methyl methacrylate solution 3 times ~ 3 times (containing red peak test peak, 2. "method treatment") and then washed with water. , add ethanol-ammonia water and decompose the solution again for 3 to 5 times, 5al each time. Collect the solution. Neutralize with acid and decompose to nearly 10ml, decompose with water: vacuo to 3ml. Filter through 5.5 filter membrane. Enter into commercial phase (5.2.2): the prepared liquid-liquid containing 10ml is suitable for the test of commercial phase: put the crushed liquid into Doushan, grab 2m. Take the organic phase: the cut off rate is 1/23m. Extract again until the organic phase is free of 2m, combine the organic phases, change the saturated acid to 2m, take the organic phase 4m each time, transfer to a heating rack in a water-filled oven for 1m, add hexane or water, extract evenly for 1 to 6 times, combine the aqueous hydrogen peroxide layer (containing water for absorption) and wash it with water for 10m each time, and adjust the volume to 50%. Filter at 0:r, take 1. Pass HPLC and chromatograph. 5.3 HPLC reference conditions
5.3.1 Column, Ywciclrm not transparent example *4.emmi,dx.2aumm5.3.2 Full gram phase: hand/2 press (pH=4,2.02mol/1)5.3.3 Elution, methanol: 20~6.3%/io:35%-98%.9%/min; 98% line 6i5.3.4 Flow rate: 1 n1./mir.
5.3.5 Use an ultraviolet balance, 27 source length,
Take the same volume of sample concentration and the combined or old color standard liquid and put them into the high-efficiency phase chromatograph, and make the plate according to the retention time, and the external standard method is as follows:
5.5 Calculation of results
The content of the color flash in the total sample is calculated according to formula (1): 1
X=:0×10g
Wherein:
The content of the aromatic colorant in the sample is grams per point (s/kg); the content of the colorant in the liquid is in micrograms. —Annual production unit (liters/L)
V, —Total volume of test sample, in milliliters (test sample, where the digit is grams).
Calculation result shall not exceed two valid digits,
5.6 Precision
GB/T 5009.35-2003
The absolute difference between the two values ​​obtained under the condition of repeated operation shall not exceed half of the mean. 5.7 Others
-Industry news;
-English vegetable:
-Blue:
:--Red,
Provision:
Figure: Chromatographic separation of eight kinds of colorants
Method 2 Thin ink chromatography
e Principle
If the colorant is adsorbed by the water under the condition of alkaline solution, it will be desorbed under the condition of alkali, and then separated by paper chromatography or thin layer chromatography, and then compared with the standard for qualitative and quantitative analysis, the maximum detection point is 50. The limit of the method is 1 and the limit of the method is 30mg/kp. 7 Trial
7.1 No oleic acid. Boiling range 60℃.~92.
7.2 Methanol
7.3. Dyeing rubber powder tilong e: 2co H
7.4 push rubber G.
7.5 system: 110).
7.6 alcohol formic acid solution: (G=4),
7.7 hydrogen peroxide (302/1.)
7.B sea sand: use hydrochloric acid (1-10) 15nm water to medium rot, then use reduced sodium solution <50g/1. boiled 15min. water to neutral, then dry in ICS: the energy of the ground in the military end: 7.97 53%.
7.10 Alcohol-ammonia solution: Take mL of oxygenated water, add alcohol (?%) to 1mL279
CQ/T 5009.35-2003
7.1p6 water, add lemongrass (20%) to pH7.12 Hydrochloric acid (1+=0).
7.13 Bamboo acid solution (200/1.)
7.14 Grain (100/T.)
7.15 Confirmation method is the same as 7.8
7.16 and the following:
butanol-anhydrous alcohol-ammonia water (1%) (6+2+3), for paper color, 7. 16. 2
certification: J original pyridine-water (1※++4) for paper color. Methyl ethyl ketone propylene glycol (7133): used for paper color. 7. 16.3
Methylenediamine-hydrogen water (1u-102), for full-layer color. 7. 16. 4
And-hydrogen water-ethanol (-1-10): for color grinding 7. 16. 5
7.16.6 Sodium citrate solution (25g/1.)-hydrogen water-ethanol (H-1+2) for thin layer chromatography 7.17 Synthetic colorant standard solution: According to 3.16, prepare the colorant standard solution with a concentration of 1/10 equivalent (TLE). Colorant standard solution: Before use, take 5.0mL of the color standard solution and dilute it to 10ml volumetric flasks. Add 16g 7.18
water until solid. This solution is dried and scanned with 0.0% color. 8.1 The spectrophotometer can be used.
8.2 Slightly inject or absorb the color,
8.3 Spread it out to 25cm×6cm×4cm. || tt||8.4 Analytical cylinder,
8.5 Key paper, medium speed filter paper, for paper chromatography. 8.6 Thin layer, 5cm×2n(r
8.7 Electric blower.
8.8 Water.
9 Analysis steps
9.1 Sample treatment
1.1 Fruit water, fruit water, carbonated water, weigh 50.0% sample in 100ml beaker, carbonated water needs to be heated to remove the alcohol, 9.1.2 Prepared wine: weigh 10% sample, 1:1% sample in 100ml beaker. Heat the broken porcelain pieces to remove the alcohol, 9.1.3 Stretch, starch, starch: weigh 0.5g or 0.8g sample, stop 3cm3 water thermal decomposition, the right liquid H value is higher. 3.1.4 Toffees: weigh 10.Ug of the sample, grind it evenly, add 30mL of ethanol-hydrogen solution, and reduce it to 2) rl., then add 1.0m] of acid (110) [acid can be melted (1K/1, until it is self-refined, and then swell with a little simmering, and collect the filtrate. 9.1.5 Cakes: Take 5.Cg of the sample, add sea sand and mix, blow dry with hot air (the hand model has been diffused): add 3mL of petroleum ether. Let it sit for a while, and the oiliness will come out frequently. Repeat this process three times to remove all the fat, blow dry and grind it into powder. All the samples are re-melted in a bucket or a bucket, and then use Add ethanol-nitrogen solution to remove color, until the colorant is completely removed, the following steps are as follows: S.1.4: "The concentration of the solution in the bath is about 2 ml.", 9.2: Separation of adsorbents, heat the solution obtained after treatment to 40 °C, add 0.5 g\-1.0 g of anhydride powder and stir thoroughly, filter with citric acid (2C9R/,) until the colorant is completely adsorbed. If there is still color in the solution, remove it with an acyl filter. Transfer all the adsorbed anhydride to a filter (filter). If it is filtered with a vertical filter, it can be slowly drawn out with water. If it contains natural colorant, wash it with 2 ml of water. Wash it with a neutral solution 1 to 3 times, each time until it is colorless. Use 7CT water to neutralize the effluent. Stir well during washing, then dissolve in ethanol for 1-2 hours, and then desorb the effluent in water. If it is monochromatic, dilute it accurately with water to 50 ml and measure it by spectrophotometer. If more than one kind of solvent is separated, select paper chromatography or thin film chromatography for separation and then measure, that is, concentrate the above solution to 2mL with water and transfer it to a 100ml volumetric flask, rinse with 60% 70% ethanol, fill it up and add it to the scale in the volumetric flask.
9.3 Qualitative analysis
9.3.1 Paper color tone
Take a paper for spectrum analysis and dot 3-10 drops of 12 standard colorants on the starting line of the cut-off point. 7.16.1.7.16.2 Fold the developing agent evenly and develop it by the upward method. When the developing agent has been developed to 1cm, take out the paper and let it air dry for ten minutes. Compare it with the standard for qualitative analysis. 5mL of Zen solution, dot it in strips from left to right on the starting line, dot a drop of colorant on the left side of the paper, develop it according to the method, and after it is dried, dot the qualitative analysis first and then use it for qualitative analysis. Indigo is easy to see under alkaline conditions. You can use 7.16.3 Developing agent, 9.3.2 Chromatography
9.3.2.1 Preparation of the plate
Weigh 1.6 g of indigo powder, 0.4 g of starch and 2 g of gelatin in a suitable base, grind with 15 mL of water, and cut into a 0.mm thick plate in a thickener. After drying at room temperature for 10 minutes, dry for 30 minutes and place in a drying oven for later use. 9.3.2.2 Spotting
At the bottom 2c of the plate, spot 0.mL of sample from left to right in a parallel shape with the edge. The left spot 2L of the sample is the color standard.
9.3.2.3 Development
For red and sunscreen red, use 7.16.4 to develop blue and shadow blue. Use 7.16.5 to develop lemon yellow and other potentials. Use 7.15.5 as the developing agent. Take an appropriate amount of developing agent and place the thin layer plate in the developing rack. If the color agent is obviously absorbed and then recovered, and it is lighter than the standard spot, such as length: the same as the positive and negative:
9.4. 1 Sample determination
Collect the stripe spots on the paper chromatogram, wash with hot water several times, transfer the washing liquid into a 10 ml colorimetric tube, and dilute to the scale with water for colorimetric measurement
Scrape off the diffused part of the thin layer chromatogram with a mouse, transfer it into a funnel, adsorb the colorant with ethanol-nitrogen, add hydrogen several times until the desorption liquid is evaporated, remove nitrogen on water, transfer it into a 10 ml colorimetric tube, add water to the scale for colorimetric measurement.
9.4.2 Preparation of standard solution
Pipette 0, 0.1, 1.0, 2.0, 1.C, 4.0 mL of lipid red, leucovorin, lemon balm, and yellow pigment standards, or 0, 0.30.1, 0.6, 0.5, 1.mL of brilliant blue and indigo blue standard drop solution respectively and place them in 1:19 nm/1 colorimetric tubes, making sure to add water to the mark.
The above samples and standard tubes were respectively used in 1m colorimetric cups with tubes to adjust the zero point, and the absorbance was determined (510ml of steel red, 521ml of rutin red, 100nm of lemon yellow, 432am of chrysanthemum yellow, 627mm of indigo, 520nm of saturated blue). The absorbance was determined, and the standard curve was drawn for comparison or compared with the standard series of measurements.
9. 5 The results were calculated using the formula (2) for the colorant content of sample 1: AX000
X==×VMx1 000
GB/\I50D9.35--2003
X--the content of colorant in the sample, in grams per gram 1/g--the same colorant in the plate, in grams (MF)--the mass of the sample, in grams per milliliter (g or product); V--the total volume of the sample after desorption, in liters (mL)) sample plating point (paper) volume, in milliliters.). The third method is oscillographic polarography
10 principle
The colorant in food can be measured by a certain buffer solution. The higher the value, the higher the change in the colorant. When there is one or more different colors in food, it can be used for qualitative and quantitative separation. 11 reagents || tt || 11.1 Waste liquid A: phosphate grade wash solution (commonly used for red and red composite color), for the determination of amaranth, breast red, glycine yellow, lemon yellow and red color,
Take 13. sodium dihydrogen phosphate: KI and 14.1 sodium dihydrogen phosphate (Na[IP]) or 3. sodium dihydrogen phosphate (Na[IP]) containing product water, 1 HU) and 10.0 chlorine oxide, dissolve in water and adjust to 1. 11.2 Base liquid: acetic acid pad liquid for green color (color), called blue, lemon, the determination of red color agent base liquid:
Take 40.ml ice water, add about 1 ml of water, 20.0 acetic acid acetic acid solution, release rate 11. 11.3 citric acid solution, 2UUg/1-
11.4 ethanol-ammonia solution, take 10 ml of concentrated ammonia water, add acetic acid 11.5 Color standard color reagent 6.10, after decomposition, add water to the scale, this solution has 1.00ml, 11.6 color standard solution, absorb 1.00ml of color standard solution, put it in [001.1. yield bottle, add water to the mark: this mg color reagent || tt || 12 12.1 Spectrometer. 12.2 Commonly used glass fiber. 13 Analysis steps 13.1 Sample preparation 13.1.1 Samples and two types: Take 10,1-5.1> samples, cool them down with 2)g/cm2 of oxygen and aldehyde (1+1) by heat-driving, and then add water to the original volume. 13.1.2 Surface chromatogram: Take samples. .1K~10,0 algae water wash until the color is completely removed: the eluent volume reaches a certain value.
13.1.3 water and jelly: sampling: 0, hot solution, cool to a certain rate, C, 13.1.4 oil, sampling 5.0 in 50ml. Centrifuge, press oil wash times: about 2ml each time. l.. liter glass stirring spoon, centrifuge, discard the upper elimination level, remove the oil film of the line at low temperature, dissolve in nitrogen solution and mix to 25, uL, island center, take a certain amount of the upper elimination policy, heat with a certain amount of water to dissolve in the element, and Water legal person? L-shaped bottle fixed report, 2
GB/T5009.35—2003
13.1.5 Milk sugar inflammation Take 5.3g of ethyl acetate and shrink to 2.0rl. Centrifuge, take 20.ml of supernatant. Add 20ml of water. Sample about 20mL cooled and adjusted to pH 4 with 2C0/L gel. Then make gel at 29C.5-1.0 yuan for rapid pigment complete adsorption method. Wash the centrifuge with L acidic water, prepare the upper liquid, wash the precipitate with aqueous solution for 3-4 times, and then wash it with appropriate aqueous solution. The chromatogram was eluted with ethanol-ammonia double elution, dried in a water bath, and heated with an appropriate amount of water to degrade the chromatogram. After washing with 11 mT of water, the concentration was measured and the concentration was narrowed. 13.2 Determination 13.2.1 Microspectral conditions: droplet electrophoresis, step effect, three-electrode blocking, scanning rate 19 nm/s, the initial scanning potential of substrate A was 0.05 V, and the full scanning potential was 9 V. The reference potentials were Van der Waals red-42 V, chrysanthemum yellow-5.50 V, lemon yellow-5.50 V, and chrysanthemum red-0.2 V. The initial scanning potential of substrate T was V and the full scanning potential was 1.5 V. Reference potentials (the peak position of the liquid substrate was positive, the peak position was negatively shifted): energy 3.5 V, chrysanthemum yellow-0.32 V, image yellow-C, 15 V, normal temperature-3, 50 V V, 13.2.2 Standard supply line: Collect the old color measurement standard and use the following: 1.5U, 1.2.J.3.0W.4.0mL respectively in the colorimetric solution + add 500l of base liquid and 10.0l of water to determine the silver content respectively 0.0501.00200.3004.00ml. After the determination on the microcomputer instrument, the reagent is automatically reduced. 13.2.3 Test column determination, take the sample and bury 1.6ml. Or quantitative color separation to reduce the electric official, try to reduce the amount). Add 5.m of base liquid and add water to .w. Combine with the standard series wave: 14 Calculate the
sample color reagent estimate according to formula 3 solution: X-
: ×0
xVXVX:30X13
X—the content of the colorant in the sample, in units of gram per milliliter (-/-); Jre
the volume of the sample solution, in units of millimeters: V—the total volume of the test solution, in units of millimolar) The total value of the two values ​​retained is calculated.
15 Abstract Density
The correlation between the results of two independent determinations under vertical conditions shall not exceed the arithmetic mean -,3314 Calculate the colorant content in the sample according to the formula 3: X-
xVXVX:30X13
X—the content of the colorant in the sample, in grams per milliliter (-/-); Jre
The volume of the sample solution in grams, in millimeters: V-the total volume of the test solution, in millimeters) The total value of the two values ​​retained is calculated.
15 Abstract Density
The correlation between the two independent determination results obtained under vertical conditions shall not exceed the arithmetic mean-3314 Calculate the colorant content in the sample according to the formula 3: X-
xVXVX:30X13
X—the content of the colorant in the sample, in grams per milliliter (-/-); Jre
The volume of the sample solution in grams, in millimeters: V-the total volume of the test solution, in millimeters) The total value of the two values ​​retained is calculated.
15 Abstract Density
The correlation between the two independent determination results obtained under vertical conditions shall not exceed the arithmetic mean-33
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