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GB 5009.89-2016 English PDF (GB5009.89-2016)

GB 5009.89-2016 English PDF (GB5009.89-2016)

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GB 5009.89-2016: National Food Safety Standard -- Determination of Niacin and Nicotinamide in Foods

This Standard specifies the determination method for niacin and nicotinamide in foods. In this Standard, method I is microbiological method, which is applicable for the determination of the total content of niacin and nicotinamide in all types of foods including fortified foods using natural foods as the base. Method II is high performance liquid chromatography, which is applicable for the determination of acid and nicotinamide in fortified foods.
GB 5009.89-2016
GB
NATIONAL STANDARD OF THE
PEOPLE REPUBLIC OF CHINA
National food safety standard
Determination of niacin and nicotinamide in foods
ISSUED ON. DECEMBER 23, 2016
IMPLEMENTED ON. JUNE 23, 2017
Issued by. National Health and Family Planning Commission of the
PEOPLE Republic of China;
China Food and Drug Administration.
3. No action is required - Full-copy of this standard will be automatically & immediately delivered to your EMAIL address in 0~60 minutes.
Table of Contents
Foreword ... 3
1 Scope ... 4
2 Principle ... 4
3 Reagents and materials ... 4
4 Instruments and equipment ... 6
5 Analysis procedure ... 6
6 Expression of analysis results ... 9
7 Precision ... 10
8 Others ... 10
9 Principle ... 10
10 Reagents and materials ... 11
11 Instruments and equipment ... 12
12 Analysis procedure ... 12
13 Expression of analysis results ... 14
14 Precision ... 15
15 Others ... 15
Annex A Medium and reagents ... 16
Annex B Standard solution concentration calibration method ... 19
Annex C Liquid chromatogram of niacin and nicotinamide standard solutions ... 20
Foreword
The Standard replaces GB/T 5009.89-2003 ?€?Determination of niacin in foods?€?, GB 5413.15-2010 ?€?National food safety standard - Determination of vitamin niacin and niacinamide in foods for infants and young children, milk and milk products?€? and GB/T 9695.25-2008 ?€?Meat and meat products - Determination of vitamin PP content?€?. Compared with GB 5413.15-2010, the main changes of this Standard are as follows. - The standard name is changed to ?€?National food safety standard - Determination of niacin and nicotinamide in foods?€?;
- ADJUST the reagent sequence and format;
- MODIFY and REFINE the pre-treatment methods applicable to different food types (Method I);
- ADD the standard solution concentration calibration method (Method II); - REASSESS the detection limit, ADD the limit of quantification.
National food safety standard ?€?
Determination of niacin and nicotinamide in foods
1 Scope
This Standard specifies the determination method for niacin and nicotinamide in foods. In this Standard, method I is microbiological method, which is applicable for the determination of the total content of niacin and nicotinamide in all types of foods including fortified foods using natural foods as the base. Method II is high performance liquid chromatography, which is applicable for the determination of acid and nicotinamide in fortified foods.
Method I Microbiological method
2 Principle
Niacin (nicotinamide) is a nutrient necessary for the growth of Lactobacillus plantarum (ATCC 8014). Under certain control conditions, use the specificity of Lactobacillus plantarum to niacin and nicotinamide that forms the optical density in the sample containing niacin and nicotinamide, to determine the content of niacin and nicotinamide. 3 Reagents and materials
Unless otherwise stated, the reagents used in this method are analytical reagents, and the water is Grade 2 water specified in GB/T 6682. The medium can be
commercialized medium that meets the test requirements.
3.1 Strains
Lactobacillus plantarum (ATCC 8014), or other valid standard strains.
3.2 Reagents
3.2.1 Hydrochloric acid (HCl).
3.2.2 Sodium hydroxide (NaOH).
3.2.3 Sodium chloride (NaCl).
stock strains.
The stock strain is inoculated into Lactobacillus agar medium before the test, and cultured in the incubator at 36 ??C ?? 1 ??C for 20 h ~ 24 h to activate the strain for the preparation of inoculation solution. Stock strains that are stored more than a few weeks cannot be used immediately for the preparation of inoculation solution, they shall be continuously inoculated 2 generations to 3 generations before the test to ensure the vitality of the strain.
5.2 Preparation of inoculation solution
One day before the test, MOVE part of the strain from the Lactobacillus agar medium to 10 mL of sterilized Lactobacillus broth medium, and CULTURE in the incubator at 36 ??C ?? 1 ??C for 6 h ~ 18 h. Under aseptic conditions, CENTRIFUGE the culture broth for 15 min and DISCARD the supernatant. ADD 10 mL of sterilized physiological saline to re-disperse the cells; MIX thoroughly on a vortex mixer; CENTRIFUGE for 15 min; DISCARD the supernatant. REPEAT the centrifugation and cleaning steps three times. PIPETTE 1 mL of third-time cell dispersion solution to add to 10 mL of sterilized physiological saline, so that it is mixed evenly into a suspension, for further use. The light transmittance value of the suspension is read with a 721 spectrophotometer at a wavelength of 550 nm and using 0.9 % saline as a reference. The light transmittance value is adjusted with 0.9 % physiological saline or third-time cell dispersion solution to be within 60 % to 80 %. USE immediately.
5.3 Preparation of samples
Potatoes, beans, nuts (shelled) samples shall be crushed, ground and sieved (the aperture of sieve plate is 0.3 mm ~ 0.5 mm); milk powder and rice flour samples shall be mixed well; meat, eggs, fish, animal viscera shall be made into chyme with crushing machine; fruits, vegetables and semi-solid foods samples shall be mixed homogeneously; liquid samples shall be shaken to mix well before use. If the sample cannot be tested immediately, it shall be stored in the refrigerator at 4 ??C. 5.4 Extraction of samples
Accurately weigh niacin samples. WEIGH 2 g ~ 5 g (accurate to 0.01 g) of general dairy, fresh fruit and vegetable sample; WEIGH 0.2 g ~ 1 g (accurate to 0.01 g) of cereals, beans, nuts, viscera, raw meat, dry sample, 5 g of liquid sample; accurately WEIGH 2 g (accurate to 0.01 g) of milk powder, rice flour sample; WEIGH 0.1 g ~ 0.5 g of general nutrient supplement and complex nutrition enhancer; WEIGH 0.2 g ~ 1 g of food; WEIGH 5 g ~ 10 g of liquid beverage or liquid, semi-liquid sample in a 100-mL Erlenmeyer flask, adding sulfuric acid solution with mass 10 times the dry mass of the tested substance. After hydrolysis at 121 ??C for 30 min, cool to room temperature. USE 0.1 mol/L sodium hydroxide solution to adjust the pH to 6.0 ~ 6.5, then USE 0.1 mol/L hydrochloric acid to adjust the pH to 4.5 ?? 0.1, DILUTE with water to 100 mL, FILTER calculate the content of niacin and nicotinamide in the sample.
10 Reagents and materials
Unless otherwise stated, the reagents used in this method are analytical regents and the water is Grade 1 water specified in GB/T 6682.
10.1 Reagents
10.1.1 Hydrochloric acid (HCl). guarantee reagent.
10.1.2 Sodium hydroxide (NaOH). guarantee reagent.
10.1.3 Perchloric acid (HClO4). with volume fraction of 60 %, guarantee reagent. 10.1.4 Methanol (CH3OH). chromatographically pure.
10.1.5 Isopropyl alcohol (C3H8O). chromatographically pure.
10.1.6 Sodium heptanesulfonate (C7H15NaO3S). chromatographically pure.
10.1.7 Amylase. with enzyme activity ??? 1.5 ??/mg.
10.2 Preparation of reagents
10.2.1 Hydrochloric acid (5.0 mol/L). MEASURE 415 mL of hydrochloric acid with a measuring cylinder in a 1000 mL flask; ADD 585 mL of water; MIX well.
10.2.2 Hydrochloric acid (0.1 mol/L). PIPETTE 8.3 mL of hydrochloric acid with a pipette in a 1000 mL flask; ADD 991.7 mL of water; MIX well.
10.2.3 Sodium hydroxide (5.0 mol/L). WEIGH 200 g of sodium hydroxide (3.2.2) in the flask and ADD water to dissolve; TRANSFER to a 1000-mL volumetric flask; ADD water to the constant volume; MIX well.
10.2.4 Sodium hydroxide (0.1 mol/L). WEIGH 4.0 g of sodium hydroxide (3.2.2) in the flask and ADD water to dissolve; TRANSFER to a 1000-mL volumetric flask; ADD water to the constant volume; MIX well.
10.2.5 Mobile phase. 70 mL of methanol, 20 mL of isopropanol and 1 g of sodium heptanesulfonate, that are dissolved with 910 mL of water and mixed well, with the pH adjusted to 2.1 ?? 0.1 with perchloric acid, and filtered through 0.45 ??m membrane. 10.3 Niacin and nicotinamide standard solution
Niacin (C6H5NO2) and nicotinamide (C6H6N2O). with purity > 99 %, or reference materials certified by the state and awarded the reference material certificate. ADD about 25 mL of water at 45 ??C to...

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