GB 1886.174-2024 National food safety standard-Food additives Enzyme preparations for food industry
1 Scope
This standard is applicable to the enzyme preparations used in for food industry permitted by GB 2760 National food safety standard - Standard for the use of food additives and related announcements.
2 Terms and definitions
2.1 Enzyme preparations for food industry
preparations which are used in food industry and have special catalytic activity, directly extracted from edible or inedible parts of animals or plants, or obtained from fermented and extracted microorganisms (including but not limited to bacteria, actinomycetes and fungi) by traditional breeding or gene recombination technology, or by further purification, preparation and other processes (which may contain one or more active enzyme components)
2.2 Enzyme activity
indicator of enzyme to catalyze a specific reaction under certain conditions
2.3 Antibacterial activity
characteristic to inhibit or kill microorganisms
2.4 Accessory materials for enzyme preparations
food raw materials and food additives added for the purposes of activity preservation, circulating storage and standardized use of enzyme preparations
2.5 Immobilized enzyme preparations
preparations that immobilize enzyme on the carrier by physical and/or chemical methods, and effect in insoluble state during food production
2.6 Immobilized carrier
materials used to immobilize enzymes in immobilized enzyme preparations
3 Product classification
They can be classified into solid (including immobilized) and liquid preparations according to the form.
4 Technical requirements
4.1 Requirements for raw and accessory materials
4.1.1 Raw and accessory materials used for the production of enzyme preparations shall meet the relevant requirements, and under the specified conditions of use, there shall be no harmful residual contamination to the final food.
4.1.2 Animal tissues used to extract enzyme preparations shall meet the requirements of meat quarantine.
4.1.3 Plant tissues used to extract enzyme preparations shall not be mildewed.
4.1.4 Taxonomic and/or genetic identification shall be performed on microbial production strains, which shall conform to GB 2760 National food safety standard - Standard for the use of food additives and related announcements. The preservation methods and conditions of strains shall ensure the stability and repeatability between fermentation batches.
4.1.5 For the active preservation, circulated storage and standardized use of enzyme preparations, necessary accessory ingredients are allowed to be added into commercial enzyme preparations. The accessory materials for enzyme preparations shall not play a functional role in the final food, but shall generally be used in appropriate amount according to the production needs, and the varieties and amount used shall be minimized under the expected purpose. See Annex A for the list of accessory materials permitted in enzyme preparations.
4.1.6 The immobilized carrier used in immobilized enzyme preparations shall conform to GB 9685 National food safety standard - Standard for the use of additives for food contact materials and products, GB 2760 National food safety standard - Standard for the use of food additives or other provisions in the list of raw materials allowed to be used in food.
4.2 Physical and chemical indexes
The enzyme activity of the product shall conform to the claim.
Note: See Annex B for the reference method of enzyme activity determination of some enzyme preparations. Enterprises may comply with the methods given in the corresponding standards or stipulated by enterprises.
4.3 Contaminant limit
The limits of contaminants shall meet the requirements of Table 1.
Table 1 Limits of contaminants
4.4 Limits of microorganisms
The limits of microorganisms shall meet the requirements of Table 2. The enzyme preparations produced by microorganisms obtained by gene recombination technology shall not be detected with producing strain.
Table 2 Limits of microorganisms
4.5 Antibacterial activity
Antibacterial activity of enzyme preparations from microorganism shall not be detected, and the antibacterial activity shall comply with GB 4789.43 National food safety standard - Food microbiological examination - Determination of antibacterial activity of enzyme preparations from microbial sources.
Annex A
List of accessory materials for enzyme preparations
For the list of accessory materials for enzyme preparations, see Table A.1.
Table A.1 List of accessory materials for enzyme preparations
Annex B
Reference method for the determination of enzyme activity of some enzyme preparations
B.1 Reference method for the determination of enzyme activity of some enzyme preparations
For the reference method for the determination of enzyme activity of some enzyme preparations, see Table B.1.
Table B.1 Reference method for the determination of enzyme activity of some enzyme preparations
B.2 Determination of α-acetolactate decarboxylase activity
B.2.1 α-acetolactate decarboxylase
enzyme that can react with α-acetolactic acid for decarboxylation and convert the carboxyl group of α-acetolactic acid into 3- hydroxy -2- butanone.
B.2.2 α-acetolactate decarboxylase activity
At 30℃ and pH 6.0, 1g or 1mL of enzyme sample reacts with α-acetolactic acid, generating 1μmol of 3- hydroxy -2- butanone (acetoin) every minute, which is a unit of enzyme activity, expressed in U/g (or U/mL).
B.2.3 Spectrophotometer
B.2.3.1 Scope
This method specifies the determination method of α-acetolactate decarboxylase activity.
This method is applicable to the determination of the enzyme activity of α-acetolactate decarboxylase in α-acetolactate decarboxylase preparation by spectrophotometer. This method is not applicable to the determination of the enzyme activity of α-acetolactate decarboxylase in beer and other alcoholic products.
The existence of 3- hydroxy -2- butanone (acetoin) and/or diacetyl in the sample will make the test results larger.
Acetoin is likely to form dimer during storage, which will affect the test results. However, this method can still be used if the measures described in the preparation step are followed to prevent it.
B.2.3.2 Principle
α-acetolactate decarboxylase reacts with α-acetolactate to decarboxylate to produce acetoin. Acetoin reacts with the mixture of naphthol and creatine under alkaline conditions to produce red products. By measuring the absorbance of the solution at 522nm, the amount of acetoin produced by the reaction can be obtained from the standard curve of acetoin, and then the enzyme activity of α-acetolactate decarboxylase can be calculated.
B.2.3.3 Reagents and materials
Unless otherwise specified, all the reagents used are analytically pure, and all the water is secondary water meeting GB/T 6682.
B.2.3.3.1 Acetoin (3- hydroxy -2- butanone) [CH3COCH(OH)CH3, CAS No.: 513-86-0], with a purity of ≥ 95%, or a standard certified by the nation and awarded the certificate of reference material.
B.2.3.3.2 MES(9.76g/L)- sodium chloride (35.064g/L)- polyoxyethylene dodecyl ether (1.52mL/L) buffer: weigh 48.80g of 2-N-morpholino] ethanesulfonic acid and 175.32g of sodium chloride respectively into a beaker, dissolve the mixture with about 4.5L of water, then add into 7.60mL of 15% polyoxyethylene dodecyl ether solution and stir it even. Adjust the pH to 6.00±0.05 with about 1mol/L sodium hydroxide solution. Then transfer the solution to a 5000mL volumetric flask and dilute it to the scale with water, mix well. The solution is valid for 1 week at normal temperature (15℃~20℃).
B.2.3.3.3 α-acetolactic acid substrate (2.00mL/L): absorb 100μL of ethyl -2- acetoxy -2- methylacetoacetic acid into a 50mL volumetric flask, add 6.0mL of sodium hydroxide solution of about 0.50mol/L, shake for 20min, and then add buffer to about 40.0mL; adjust the pH of the solution to 6.00±0.05 with about 1mol/L hydrochloric acid. Then, use the buffer to dilute it to the scale. This solution shall be prepared right before use.
B.2.3.3.4 Naphthol (10.0g/L)/ Creatine (1.0g/L) developer: weigh 5.0g of 1- Naphthol and 0.5g of Creatine respectively, put them into a 500mL volumetric flask, and dissolve them with about 1mol/L sodium hydroxide solution, and dilute it to the scale. This solution shall be prepared right before use. Avoid light when preparing, and take an ice bath.
