What is the procedure of toxicological evaluation of food safety? What is the specific operation of Ames experiment?

The toxicological evaluation procedure of food safety includes four stages, namely, acute toxicity test, cumulative toxicity and mutagenicity test, subchronic toxicity (including reproduction and teratogenesis) test and metabolic test, and chronic toxicity (including carcinogenesis) test.

There are two conventional methods of Ames test: spot test and flat doping test, and the specific operations are as follows:

1, spot check;

Suck 0. 1ml of bacterial liquid after enrichment culture of test bacteria, inject it into the upper soft agar at about 45℃ after melting, add 0.3 ml-0.4 ml of S9 mixed liquid for S9 activation, immediately mix it evenly, pour it on the bottom plate, flatten it and concentrate it.

Clamp the edge of sterilized circular filter paper with sterile needle pliers, soak the paper in the test solution, or directly take the solid test substance and stick it on the surface of the upper culture medium. At the same time, make solvent control and positive control, and stick them on the corresponding position on the plate respectively.

The Petri dish was inverted in an incubator at 37℃ for 48 hours. Dense colony circles grow on the periphery of the paper, which is positive; Colony dispersion, density is similar to spontaneous regression, which is negative.

3. Flat doping test:

Add a certain amount of sample solution and 0. 1ml of test bacteria solution to the upper soft agar, and add 0.3 ml-0.4 ml of s9mix for metabolic activation. After stirring, quickly pour it on the bottom plate and spread it for condensation.

At the same time, negative and positive controls were made, and each treatment was 3 parallel. Samples usually have 4 ~ 5 doses. At the beginning, the dose range should be larger. When there are positive or suspected positive results, the dose-response relationship should be determined within a narrow dose range. Training is the same as above.

The ratio of the average number of reverted colonies in each Petri dish with the same dose to the average number of spontaneously reverted colonies in each negative control Petri dish is the mutagenicity ratio (MR). MR value ≥2, and there is a dose-response relationship, the background is normal, and it is judged to be mutagenic positive.

Extended data:

First, the food safety toxicology evaluation experiment project;

The first stage: acute toxicity test, the test items:

1. The oral lethal dose (LD50) was determined by Horn's method, probability unit method or Kohler's method. If the dose of 10g/kg body weight still does not cause animal death, it is not necessary to determine the LD50.

2. If necessary, conduct a seven-day feeding test. The above two projects use mice or rats of two sexes respectively.

The second stage: cumulative toxicity and mutagenicity test, and the test items are:

1. Cumulative coefficient method. Rats or mice of two sexes were used, with 20 rats in each group.

2. Twenty-day test method. Rats or mice of two sexes were used, and there were 10 males and females in each dose group. Choose one of the above two methods.

The third stage: subchronic toxicity (including reproduction and teratogenesis) test and metabolic test. Test items include:

1.90 days feeding experiment.

2. Feeding and breeding experiments.

3. Feeding teratogenic test.

4. Traditional teratogenic test.

The fourth stage: chronic toxicity (including carcinogenesis) test, the test items:

Two years of chronic toxicity test and carcinogenesis test can be combined in one animal test. Male and female rats or mice were used.

2. Experimental requirements for toxicological evaluation of food safety:

1. All new chemicals created in China generally need to go through four stages of testing. In particular, if the chemical structure suggests chronic toxicity or carcinogenicity, the experiment must be carried out in four stages with large output, wide use area and many opportunities for ingestion.

2. Any derivative whose chemical structure is basically the same as that of the known substance (which has passed the safety evaluation and allowed users) can be evaluated by relevant experts according to the results of the first, second and third stage tests to decide whether the fourth stage test is needed.

3. For the chemical substances that are copied in China and have certain toxicity, if most countries have allowed them to be used in food, and there is evidence of safety, or the World Health Organization has announced the allowable intake per person per day (ADI), and the production unit can prove that the physical and chemical properties, purity, impurity composition and content of our products are consistent with those of foreign products, the first and second stage tests can be carried out first.

If the test results are consistent with similar foreign products, the test will not be continued and can be evaluated. If the evaluation results allow it to be used in food, the daily allowable amount will be set. Where the product quality or test results are inconsistent with foreign data or products, the third stage test should be carried out.

Source: Baidu Encyclopedia-Ames Test? Baidu Encyclopedia-Toxicological Evaluation of Food Safety