GB 5009.31-2025 National Food Safety Standard - Determination of Hydroxybenzoate Compounds in FoodsBy the National Health Commission of the People's Republic of China and the State Administration for Market RegulationReleased on March 16th, 2025And inFormally implemented on September 16, 2025.

1、 Scope of application and core detection substances

（1） Scope of application

Suitable for use in foodHydroxybenzoate preservativesThe content determination is based on the standard, but there are differences in the scope of application of the three specific detection methods (first method, second method, third method) in the food.

1. First method: Gas chromatography (GC)

(1) Applicable food:Other foods except for heat cured egg products, fermented sauces, and seasoning sauces;

(2) Main featuresThis is a traditional method adopted from the 2016 version of the standard, but its scope of application is limited.

2. Second method: High performance liquid chromatography (HPLC)

(1) Applicable food:All foods (with no special exceptions);

(2) Main features:This is a newly added method in the 2025 version, with a wide range of applications, which solves the problem of gas chromatography not being applicable to some complex matrix samples (such as sauces).

3. Third method: Liquid chromatography tandem mass spectrometry (LC-MS/MS)

(1) Applicable food:All foods (with no special exceptions);

(2) Main features:This is also a newly added method in the 2025 version, with high sensitivity and strong anti-interference ability, especially suitable for trace analysis, complex matrices, or detection scenarios that require high certainty results.

（2） Core testing substances

Methyl p-hydroxybenzoate

Hydroxybenzoic acid ester

Propyl hydroxybenzoate

Isopropyl p-hydroxybenzoate (newly added in the 2025 version)

Butyl p-hydroxybenzoate

Isobutyl p-hydroxybenzoate (newly added in the 2025 version)

Heptyl paraben (newly added in the 2025 version)

2、 Operation process

（1） Method 1: Gas Chromatography

This method is applicable to foods other than heat cured egg products, fermented sauces, and seasoning sauces.

1.sample preparation：

(1) Weigh about 2g of uniform sample;

(2) For complex matrices containing oil or protein, such as meat products and pastries, it is necessary to add an appropriate amount of sodium chloride and perform homogeneous extraction.

2. Extraction and purification：

(1) Mainly using methanol water solution for ultrasonic extraction;

(2) The extraction solution needs to be purified and enriched through HLB solid-phase extraction columns or similar devices, which is a key step to ensure accurate results.

3. Instrument analysis：

(1) Use a gas chromatograph equipped with a hydrogen flame ionization detector for analysis;

(2) It is recommended to use capillary columns with similar polarity, such as HP-5, for chromatographic columns;

(3) The program heating conditions need to be set according to standards to achieve effective separation of 7 ester compounds.

（2） Method 2: High performance liquid chromatography

This method is applicable to all food matrices.

1. Sample preparation:

(1) Solid or semi-solid samples need to be crushed evenly;

(2) Unlike the first method, this method typically uses methanol or acetonitrile water solution for extraction.

2. Extraction and Purification:

(1) The extraction method is a combination of vortex oscillation and ultrasound;

(2) For samples with high oil content (such as edible oil), low-temperature freezing is required after extraction to remove fat interference.

3. Instrument analysis:

(1) Use a high-performance liquid chromatograph equipped with a UV detector or diode array detector;

(2) The chromatographic column is usually a C18 reverse phase chromatographic column;

(3) The mobile phase is a mixture of methanol (or acetonitrile) and a specific concentration of ammonium acetate aqueous solution, using gradient elution.

（3） Third method: liquid chromatography tandem mass spectrometry

This method is the most accurate way to confirm and quantify trace target substances, applicable to all food matrices.

1. Sample preparation:

The process is similar to the second method, but requires higher purity of the reagents.

2. Extraction and Purification:

(1) The extraction solvent is usually acetonitrile;

(2) The purification process may be more stringent, sometimes requiring the use of techniques such as dispersed solid-phase extraction to remove matrix interference to a large extent.

3. Instrument analysis:

(1) Use a liquid chromatography tandem mass spectrometer;

(2) In the mass spectrometry part, the electrospray ion source is used to scan in the negative ion mode. Qualitative and quantitative analysis is carried out by monitoring specific parent ion and sub ion pairs, with strong anti-interference ability.

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