GB 5009.33-2025 National food safety standard – Method for determination of nitrite and nitrate in foods
1 Scope
This standard specifies the determination method of nitrite and nitrate in food.
Method I of this standard is applicable to the determination of nitrite and nitrate in food (except formula foods for special medical purposes, such as partially hydrolyzed milk protein formula, deeply hydrolyzed milk protein formula, amino acid formula, and amino acid metabolism disorder formula).
Methods II and III of this standard are applicable to the determination of nitrite and nitrate in food.
Method IV of this standard is applicable to the determination of nitrate in fresh vegetables and fruits.
Method I Ion Chromatography
2 Principle
After protein precipitation and fat removal, the sample is extracted and purified, eluted with eluent potassium hydroxide solution or sodium hydroxide solution, separated by anion exchange column, and detected by conductivity detector or ultraviolet detector. Qualitative analysis is performed based on retention time, and quantitative analysis is performed using the external standard method.
3 Reagents and materials
Unless otherwise specified, analytically-pure reagents and Grade I water (as specified in GB/T 6682) are adopted for the purpose of this method.
3.1 Reagent
3.1.1 Acetic acid (CH3COOH): chromatographically pure.
3.1.2 Potassium hydroxide (KOH).
3.1.3 Sodium hydroxide (NaOH).
3.1.4 α-amylase: with an enzyme activity ≥ 1.5 U/mg.
3.2 Preparation of reagents
3.2.1 Acetic acid solution (3%): Measure out 3 mL of acetic acid into a 100 mL volumetric flask, dilute to volume with water, and mix well.
3.2.2 Potassium hydroxide solution (1 mol/L): Weigh out 5.6 g of potassium hydroxide, add freshly-boiled cold water to dissolve, dilute to 100 mL, and mix well.
3.2.3 Sodium hydroxide solution (1 mol/L): Weigh out 4.0 g of sodium hydroxide, add freshly-boiled cold water to dissolve, dilute to 100 mL, and mix well.
3.3 Standard substance
3.3.1 Sodium nitrite standard substance (NaNO2, CAS No.: 7632-00-0): purity ≥ 99%. Or nitrite standard substances certified by the state and granted a reference material certificate.
3.3.2 Sodium nitrate standard substance (NaNO3, CAS No.: 7631-99-4): purity ≥ 99%. Or nitrate standard substances certified by the state and granted a reference material certificate.
3.4 Preparation of standard solutions
3.4.1 Sodium nitrite standard stock solution (100 mg/L): Accurately weigh out 0.1000 g of sodium nitrite dried to constant weight at 110 ℃ ~ 120 ℃, dissolve and dilute to 1,000 mL with water, mix well, and store it in a refrigerator at 2 ℃ ~ 6 ℃. The solution is valid within 3 months.
3.4.2 Sodium nitrate standard stock solution (100 mg/L): Accurately weigh out 0.1000 g of sodium nitrate dried to constant weight at 110 ℃ ~ 120 ℃, dissolve and dilute to 1,000 mL with water, mix well, and store it in a refrigerator at 2 ℃ ~ 6 ℃. The solution is valid within 3 months.
3.4.3 Sodium nitrite and sodium nitrate mixed standard intermediate solution (2.00 mg/L): Pipette accurately 2.00 mL of each of the standard stock solutions of sodium nitrite and sodium nitrate into a 100 mL volumetric flask and dilute to volume with water. This solution shall be prepared immediately before use.
3.4.4 Sodium nitrite and sodium nitrate mixed standard using solution: Pipette a proper amount of sodium nitrite and sodium nitrate mixed standard intermediate solution, add water and dilute it level by level, and make a series of standard solutions with mass concentrations of 0 mg/L, 0.0100 mg/L, 0.0200 mg/L, 0.0500 mg/L, 0.100 mg/L, 0.200 mg/L, 0.500 mg/L, 1.00 mg/L and 2.00 mg/L. This solution shall be prepared immediately before use.
Note: The range of the standard curve may be adjusted to include at least 5 concentration levels (excluding zero points) according to the content of the target analyte in the sample.
3.5 Material
3.5.1 Filter membrane syringe filter: aqueous, 0.22 μm.
3.5.2 C18 purification column: 1 mL, or a column with equivalent performance. It shall be activated with 10 mL of methanol and 15 mL of water in turn before use.
3.5.3 Ag purification column: with a column capacity of 2.0 ion equivalent ~ 2.2 ion equivalent, or a column with equivalent performance. It shall be activated with 10 mL of water before use.
3.5.4 Na purification column: with a column capacity of 2.0 ion equivalent ~ 2.2 ion equivalent, or a column with equivalent performance. It shall be activated with 10 mL of water before use.
4 Apparatus
4.1 Ion chromatograph: equipped with a conductance detector and a suppressor, or an ultraviolet detector.
4.2 Food crusher.
4.3 Ultrasonic cleaner.
4.4 Electronic balance: with a sensitivity of 0.1 mg and 0.01 g.
4.5 Centrifuge: with a rotating speed of ≥10,000 r/min.
5 Analytical procedures
5.1 Preparation and storage of specimens
5.1.1 Preparation of specimens
5.1.1.1 Liquid or semi-solid sample: Take an appropriate amount of the sample, and mix the sample thoroughly by stirring or shaking.
5.1.1.2 Solid powder sample: Take an appropriate amount of the sample and mix the sample thoroughly by shaking and other means.
5.1.1.3 Cheese: Take an appropriate amount of the sample and grind it into uniform paste. To avoid moisture loss, avoid generating excessive heat during grinding.
5.1.1.4 Solid samples of other kinds: Take an appropriate amount of the sample and crush it to uniform powder or make it uniform paste for later use.
5.1.2 Storage of specimens
Contents
Foreword i
1 Scope
Method I Ion Chromatography
2 Principle
3 Reagents and materials
4 Apparatus
5 Analytical procedures
6 Calculation (expression) of analytic result
7 Precision
8 Others
Method II Spectrophotometry
9 Principle
10 Reagents and materials
11 Apparatus
12 Analytical procedures
13 Calculation (expression) of analytic result
14 Precision
15 Others
Method III Continuous flow analysis-spectrophotometry
16 Principle
17 Reagents and materials
18 Apparatus
19 Analytical procedures
20 Calculation (expression) of analytic result
21 Precision
22 Others
Method IV Determination of nitrate in vegetables and fruits
23 Principle
24 Reagents and materials
25 Apparatus
26 Analytical procedures
27 Calculation of results
28 Precision
29 Others
Annex A Chromatograms of nitrite and nitrate ions
Annex B Preparation of cadmium particles
Annex C Determination of cadmium column reduction efficiency
Standard
GB 5009.33-2025 National food safety standard - Determination of nitrite and nitrate in foods (English Version)
Standard No.
GB 5009.33-2025
Status
to be valid
Language
English
File Format
PDF
Word Count
10500 words
Price(USD)
315.0
Implemented on
2026-3-2
Delivery
via email in 1 business day
Detail of GB 5009.33-2025
Standard No.
GB 5009.33-2025
English Name
National food safety standard - Determination of nitrite and nitrate in foods
Chinese Name
食品安全国家标准 食品中亚硝酸盐与硝酸盐的测定
Chinese Classification
Professional Classification
GB
ICS Classification
Issued by
National Health Commission and State Administration for Market Regulation
GB 5009.33-2025 National food safety standard – Method for determination of nitrite and nitrate in foods
1 Scope
This standard specifies the determination method of nitrite and nitrate in food.
Method I of this standard is applicable to the determination of nitrite and nitrate in food (except formula foods for special medical purposes, such as partially hydrolyzed milk protein formula, deeply hydrolyzed milk protein formula, amino acid formula, and amino acid metabolism disorder formula).
Methods II and III of this standard are applicable to the determination of nitrite and nitrate in food.
Method IV of this standard is applicable to the determination of nitrate in fresh vegetables and fruits.
Method I Ion Chromatography
2 Principle
After protein precipitation and fat removal, the sample is extracted and purified, eluted with eluent potassium hydroxide solution or sodium hydroxide solution, separated by anion exchange column, and detected by conductivity detector or ultraviolet detector. Qualitative analysis is performed based on retention time, and quantitative analysis is performed using the external standard method.
3 Reagents and materials
Unless otherwise specified, analytically-pure reagents and Grade I water (as specified in GB/T 6682) are adopted for the purpose of this method.
3.1 Reagent
3.1.1 Acetic acid (CH3COOH): chromatographically pure.
3.1.2 Potassium hydroxide (KOH).
3.1.3 Sodium hydroxide (NaOH).
3.1.4 α-amylase: with an enzyme activity ≥ 1.5 U/mg.
3.2 Preparation of reagents
3.2.1 Acetic acid solution (3%): Measure out 3 mL of acetic acid into a 100 mL volumetric flask, dilute to volume with water, and mix well.
3.2.2 Potassium hydroxide solution (1 mol/L): Weigh out 5.6 g of potassium hydroxide, add freshly-boiled cold water to dissolve, dilute to 100 mL, and mix well.
3.2.3 Sodium hydroxide solution (1 mol/L): Weigh out 4.0 g of sodium hydroxide, add freshly-boiled cold water to dissolve, dilute to 100 mL, and mix well.
3.3 Standard substance
3.3.1 Sodium nitrite standard substance (NaNO2, CAS No.: 7632-00-0): purity ≥ 99%. Or nitrite standard substances certified by the state and granted a reference material certificate.
3.3.2 Sodium nitrate standard substance (NaNO3, CAS No.: 7631-99-4): purity ≥ 99%. Or nitrate standard substances certified by the state and granted a reference material certificate.
3.4 Preparation of standard solutions
3.4.1 Sodium nitrite standard stock solution (100 mg/L): Accurately weigh out 0.1000 g of sodium nitrite dried to constant weight at 110 ℃ ~ 120 ℃, dissolve and dilute to 1,000 mL with water, mix well, and store it in a refrigerator at 2 ℃ ~ 6 ℃. The solution is valid within 3 months.
3.4.2 Sodium nitrate standard stock solution (100 mg/L): Accurately weigh out 0.1000 g of sodium nitrate dried to constant weight at 110 ℃ ~ 120 ℃, dissolve and dilute to 1,000 mL with water, mix well, and store it in a refrigerator at 2 ℃ ~ 6 ℃. The solution is valid within 3 months.
3.4.3 Sodium nitrite and sodium nitrate mixed standard intermediate solution (2.00 mg/L): Pipette accurately 2.00 mL of each of the standard stock solutions of sodium nitrite and sodium nitrate into a 100 mL volumetric flask and dilute to volume with water. This solution shall be prepared immediately before use.
3.4.4 Sodium nitrite and sodium nitrate mixed standard using solution: Pipette a proper amount of sodium nitrite and sodium nitrate mixed standard intermediate solution, add water and dilute it level by level, and make a series of standard solutions with mass concentrations of 0 mg/L, 0.0100 mg/L, 0.0200 mg/L, 0.0500 mg/L, 0.100 mg/L, 0.200 mg/L, 0.500 mg/L, 1.00 mg/L and 2.00 mg/L. This solution shall be prepared immediately before use.
Note: The range of the standard curve may be adjusted to include at least 5 concentration levels (excluding zero points) according to the content of the target analyte in the sample.
3.5 Material
3.5.1 Filter membrane syringe filter: aqueous, 0.22 μm.
3.5.2 C18 purification column: 1 mL, or a column with equivalent performance. It shall be activated with 10 mL of methanol and 15 mL of water in turn before use.
3.5.3 Ag purification column: with a column capacity of 2.0 ion equivalent ~ 2.2 ion equivalent, or a column with equivalent performance. It shall be activated with 10 mL of water before use.
3.5.4 Na purification column: with a column capacity of 2.0 ion equivalent ~ 2.2 ion equivalent, or a column with equivalent performance. It shall be activated with 10 mL of water before use.
4 Apparatus
4.1 Ion chromatograph: equipped with a conductance detector and a suppressor, or an ultraviolet detector.
4.2 Food crusher.
4.3 Ultrasonic cleaner.
4.4 Electronic balance: with a sensitivity of 0.1 mg and 0.01 g.
4.5 Centrifuge: with a rotating speed of ≥10,000 r/min.
5 Analytical procedures
5.1 Preparation and storage of specimens
5.1.1 Preparation of specimens
5.1.1.1 Liquid or semi-solid sample: Take an appropriate amount of the sample, and mix the sample thoroughly by stirring or shaking.
5.1.1.2 Solid powder sample: Take an appropriate amount of the sample and mix the sample thoroughly by shaking and other means.
5.1.1.3 Cheese: Take an appropriate amount of the sample and grind it into uniform paste. To avoid moisture loss, avoid generating excessive heat during grinding.
5.1.1.4 Solid samples of other kinds: Take an appropriate amount of the sample and crush it to uniform powder or make it uniform paste for later use.
5.1.2 Storage of specimens
Contents of GB 5009.33-2025
Contents
Foreword i
1 Scope
Method I Ion Chromatography
2 Principle
3 Reagents and materials
4 Apparatus
5 Analytical procedures
6 Calculation (expression) of analytic result
7 Precision
8 Others
Method II Spectrophotometry
9 Principle
10 Reagents and materials
11 Apparatus
12 Analytical procedures
13 Calculation (expression) of analytic result
14 Precision
15 Others
Method III Continuous flow analysis-spectrophotometry
16 Principle
17 Reagents and materials
18 Apparatus
19 Analytical procedures
20 Calculation (expression) of analytic result
21 Precision
22 Others
Method IV Determination of nitrate in vegetables and fruits
23 Principle
24 Reagents and materials
25 Apparatus
26 Analytical procedures
27 Calculation of results
28 Precision
29 Others
Annex A Chromatograms of nitrite and nitrate ions
Annex B Preparation of cadmium particles
Annex C Determination of cadmium column reduction efficiency
