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GB 5009.308-2025: National food safety standard - Determination of ascorbyl palmitate in food
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National food safety standard - Determination of ascorbyl palmitate in food
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GB 5009.308-2025
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Basic data | Standard ID | GB 5009.308-2025 (GB5009.308-2025) | | Description (Translated English) | National food safety standard - Determination of ascorbyl palmitate in food | | Sector / Industry | National Standard | | Classification of Chinese Standard | X04 | | Word Count Estimation | 7,736 | | Date of Issue | 2025-09-02 | | Issuing agency(ies) | National Health Commission; State Administration for Market Regulation | GB 5009.308-2025: National food safety standard - Determination of ascorbyl palmitate in food---This is an excerpt. Full copy of true-PDF in English version (including equations, symbols, images, flow-chart, tables, and figures etc.), auto-downloaded/delivered in 9 seconds, can be purchased online: https://www.ChineseStandard.net/PDF.aspx/GB5009.308-2025
National Standards of the People's Republic of China
National Food Safety Standards
Determination of ascorbate palmitate in food
Published on 2025-09-02
Implemented on 2026-03-02
National Health Commission of the People's Republic of China
State Administration for Market Regulation issued
National Food Safety Standards
Determination of ascorbate palmitate in food
1.Scope
This standard specifies a liquid chromatography method for the determination of ascorbyl palmitate in food.
This standard applies to the determination of ascorbyl palmitate in food.
2.Principle
Ascorbate palmitate in the sample was extracted with citric acid-isoascorbic acid methanol solution and then separated by reversed-phase liquid chromatography.
Detection is performed using an array detector or ultraviolet detector, and quantification is performed using the external standard method.
3.Reagents and Materials
Unless otherwise stated, all reagents used in this method are of analytical grade, and the water is Grade I water as specified in GB/T 6682.
3.1 Reagents
3.1.1 Methanol (CH4O). chromatographic grade.
3.1.2 Acetonitrile (C2H3N). chromatographic grade.
3.1.3 Phosphoric acid (H3PO4).
3.1.4 Citric acid (C6H8O7).
3.1.5 Isoascorbic acid (C6H8O6).
3.2 Reagent Preparation
3.2.1 Citric acid-isoascorbic acid methanol solution. Weigh 1.0 g of citric acid and 1.0 g of isoascorbic acid, dissolve and dilute with methanol to a final concentration.
1000mL.
3.2.2 Phosphoric acid solution (0.1%). Measure 1.0 mL of phosphoric acid, dilute with water to 1000 mL, mix well and set aside.
3.2.3 Methanol-acetonitrile solution (50.50). Mix methanol and acetonitrile in a volume ratio of 50.50 and set aside.
3.3 Standard Products
3.3.1 Ascorbate palmitate standard (C22H38O7, CAS No.. 137-66-6). purity ≥98%, or certified by the state and granted a standard.
Standard substances for quasi-material certificates.
3.4 Preparation of Standard Solutions
3.4.1 Ascorbate palmitate standard solution (1.00 mg/mL). Accurately weigh 25 mg (accurate to 0.1 mg) of the standard, and dilute with citric acid-
Dissolve isoascorbic acid in methanol and bring the volume to 25 mL, then mix well. Prepare fresh before use.
3.4.2 Ascorbate Palmitate Standard Intermediate Solution (0.100 mg/mL). Accurately pipette 1.00 mL of the 1.00 mg/mL standard solution and use...
Dilute the citric acid-isoascorbic acid methanol solution to a final volume of 10 mL and mix well. Prepare fresh before use.
3.4.3 Ascorbate Palmitate Standard Working Solution. Accurately pipette 0.10 mL, 0.20 mL, 0.50 mL, 1.0 mL,...
5.0 mL was diluted with citric acid-isoascorbic acid methanol solution and brought to a final volume of 10 mL. The solution was mixed thoroughly to obtain concentrations of 1.00 μg/mL.
Standard working solutions of 2.00 μg/mL, 5.00 μg/mL, 10.0 μg/mL, and 50.0 μg/mL. Prepare fresh before use.
Note. The concentration point of the standard working solution can be adjusted appropriately within the linear range.
3.5 Materials
Organic microporous filter membrane. 0.22μm.
4.Instruments and Equipment
4.1 Liquid Chromatograph. Equipped with a diode array detector or an ultraviolet detector.
4.2 Electronic balance. sensitivity is 0.01mg and 0.1mg respectively.
4.3 Tissue grinder.
4.4 Horizontal Oscillator.
4.5 Centrifuge. speed not less than 8000 r/min.
5.Analysis Steps
5.1 Sample Preparation
Liquid samples, such as peanut oil and fruit and vegetable juice beverages, should be shaken well before testing; powdery or paste-like samples with uniform texture, such as milk powder and fruit puree, should be thoroughly mixed before testing.
Mix thoroughly; other solid and semi-solid samples such as canned goods, bread, oatmeal, and candy should be pulverized (for gum-based candies, chopping or freezing may be necessary).
The sample is crushed, sieved to a particle size of less than 2 mm, and mixed thoroughly. It is then stored in a light-proof, airtight container under cold conditions.
5.2 Sample preparation
5.2.1 Sample Extraction
Weigh 5g of the sample (accurate to 0.001g), place it in a 50mL centrifuge tube, and add 30mL of citric acid-isoascorbic acid methanol solution.
Tighten the cap, place the tube on a horizontal shaker, and extract for 10 minutes at a shaking rate of at least 250 times/min. Transfer the entire extract to a 50mL volumetric flask.
In the middle, dilute to the mark with citric acid-isoascorbic acid methanol solution and mix well.
5.2.2 Purification
The above sample extract was filtered through filter paper (discarding the initial filtrate) or centrifuged at 8000 r/min for 1 min to obtain the supernatant, and then filtered through a 0.22 μm filter.
The sample is filtered through a microporous membrane into a brown sample vial for analysis by high-performance liquid chromatography (HPLC).
Note. If necessary, the sample solution can be diluted with citric acid-isoascorbic acid methanol solution to ensure that the concentration of ascorbate palmitate in the sample solution is within the range of the standard working solution.
Within the liquid concentration range; the sample solution should be tested within 12 hours.
5.3 Blank Test
Except for the absence of a sample, all other steps are performed simultaneously with the sample as per section 5.2.
5.4 Instrument Reference Conditions
5.4.1 Chromatographic column. C18 column (4.6 mm × 150 mm, 5 μm) or a column with equivalent performance.
5.4.2 Detection wavelength. 245nm.
5.4.3 Mobile phase. A is methanol-acetonitrile solution (50.50), B is phosphoric acid solution (0.1%).
5.4.4 Flow rate. 1.0 mL/min.
5.4.5 Injection volume. 10 μL.
5.4.6 Column temperature. 30℃.
5.4.7 The gradient elution procedure is shown in Table 1.
Table 1 Gradient elution program
Time/min Mobile phase A/% Mobile phase B/%
0.0 40 60
1.0 40 60
5.0 60 40
12.0 95 5
19.0 95 5
20.0 40 60
25.0 40 60
5.5 Construction of Standard Curve
The standard series of working solutions were injected into the high-performance liquid chromatograph in order of increasing concentration, according to the mass of ascorbate palmitate.
A standard curve was constructed with concentration on the x-axis and the measured peak area on the y-axis. The chromatogram of the standard solution is shown in Figure A.1 of Appendix A.
5.6 Determination of Sample Solution
The sample solution was injected into a high-performance liquid chromatograph to obtain the corresponding peak area. The ascorbic acid content in the sample solution was then determined based on the standard curve.
Mass concentration of palmitic acid ester.
6.Presentation of Analysis Results
The content of ascorbate palmitate in the sample is calculated according to formula (1).
X=
(ρ-ρ0)×V×f×1000
m×1000
(1)
In the formula.
X --- The content of ascorbyl palmitate in the sample, expressed in milligrams per kilogram (mg/kg);
ρ --- The concentration of ascorbate palmitate in the sample solution obtained from the standard curve, in micrograms per milliliter (μg/mL);
ρ0 --- The concentration of ascorbate palmitate in the blank test solution obtained from the standard curve, in micrograms per milliliter (μg/mL);
V --- The volume of the sample solution after dilution, in milliliters (mL);
f --- Dilution factor of the sample solution;
m --- Mass of the sample, in grams (g);
1000---Conversion factor.
The calculation result should be rounded to 3 significant figures.
Note. When the result is expressed in terms of ascorbic acid, the ascorbate palmitate content in the sample is multiplied by a conversion factor of 0.425 to obtain the ascorbic acid content; if the result needs to be expressed in terms of...
Ascorbic acid content in oils and fats is determined according to GB 5009.6.
7 Precision
The absolute difference between two independent measurements obtained under repeatability conditions shall not exceed 10% of the arithmetic mean.
8 Other
When the sample weight is 5g and the volume is 50mL, the detection limit of this method is 5mg/kg and the quantitation limit is 10mg/kg.
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