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GB 15193.5-2014: National food safety standard - Mammalian Erythrocyte Micronucleus Test Delivery: 9 seconds. True-PDF full-copy in English & invoice will be downloaded + auto-delivered via email. See step-by-step procedure Status: Valid GB 15193.5: Historical versions
Similar standardsGB 15193.5-2014: National food safety standard - Mammalian Erythrocyte Micronucleus Test---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/GB15193.5-2014GB NATIONAL STANDARD OF THE PEOPLE’S REPUBLIC OF CHINA National food safety standard – Mammalian erythrocyte micronucleus test ISSUED ON: DECEMBER 24, 2014 IMPLEMENTED ON: MAY 01, 2015 Issued by: National Health and Family Planning Commission of the People’s Republic of China Table of ContentsForeword ... 3 1 Scope ... 4 2 Terms and definitions ... 4 3 Test purpose and principle ... 5 4 Apparatus and reagents ... 5 5 Test methods ... 6 6 Data processing and result evaluation ... 8 7 Test report ... 9 8 Interpretation of the test ... 9 National food safety standard - Mammalian erythrocyte micronucleus test1 ScopeThis Standard specifies the basic test methods and technical requirements for mammalian erythrocyte micronucleus test. This Standard is applicable to the evaluation of the genotoxicity of test substances.2 Terms and definitions2.1 Micronucleus During the cell mitosis anaphase, when the chromosomes regularly enter the daughter cells to form the cell nucleus, the entire chromatids or acentric fragments or rings of chromosomes that remain in the cytoplasm. In the telophase, one or several regular subnuclei are formed separately and are contained in the cytoplasm of the cell. 2.2 Centromere During the phase of cell division, the region where chromosomes connect to spindle fibers allows the orderly movement of daughter chromosomes to the poles of daughter cells. 2.3 Normochromatic erythrocytes Mature erythrocytes that lack ribosomes and can be distinguished from immature polychromatic erythrocytes by selective ribosome dyes. 2.4 Polychromatic erythrocytes Immature erythrocytes are in the intermediate stage of development and still contain ribosomes, so they can be distinguished from mature normochromatic erythrocytes with selective ribosome dyes. 2.5 Total erythrocytes The sum of normochromatic erythrocytes and polychromatic erythrocytes.5 Test methods5.1 Test substances 5.1.1 Preparation of test substances: The test substances shall be dissolved or suspended in a suitable solvent, where water is the preferred solvent; for water-insoluble test substances, vegetable oils (such as olive oil, corn oil, etc.) can be used; for water- insoluble or oil-insoluble test substances, carboxymethyl cellulose, starch, etc. can also be used to prepare suspensions or pastes. Test substances shall be prepared before use, unless there is data indicating that their solutions or suspensions are storage stable. 5.1.2 Administration route: Oral gavage shall be used. Positive control substances can also be injected intraperitoneally. The gavage volume shall generally not exceed 10 mL/kg body weight. If it is an aqueous solution, the maximum gavage volume can reach 20 mL/kg body weight; if it is an oily liquid, the gavage volume shall not exceed 4 mL/kg body weight; the gavage volume of each group shall be the same. 5.2 Experimental animals 5.2.1 Animal species and strain selection: When using bone marrow, it is recommended to use mice or rats. When using peripheral blood, it is recommended to use mice. If it has been proven that the spleen of a certain strain of animal does not clear polychromatic erythrocytes with micronuclei, or is sufficiently sensitive to detect chemicals that cause structural or numerical aberrations of chromosomes, such animals can be used. Usually, mice aged 7 ~ 12 weeks and weighing 25 g ~ 35 g or rats weighing 200 g ~ 300 g are used. At the beginning of the test, the difference in animal weight shall not exceed ±20% of the average weight of each gender. Use at least 5 animals of each gender in each group. 5.2.2 Animal preparation: Before the test, the animals shall undergo environmental acclimatization and quarantine observation for at least 3 ~ 5 days in the experimental animal room. 5.2.3 Animal feeding: Experimental animal feeding conditions, drinking water, and feed shall comply with national standards and relevant regulations (GB 14925, GB 5749, GB 14924.1, GB 14924.2, GB 14924.3). The animals in each test group shall be housed in separate cages according to gender, with 5 animals in each cage. During the experiment, the experimental animals shall be fed basic feed and have free access to water. 5.3 Dosage The test substances shall be set up in three dose groups. In principle, the maximum dose group is the dose at which animals show severe poisoning and/or individual animals die, which can generally be 1/2 LD50. The low dose groups shall not show toxicity, and 1/4 LD50 and 1/8 LD50 shall be taken as the medium and low doses, respectively. When the animal does not die at the maximum dose (maximum use concentration and maximum gavage capacity) of the test substances in the acute toxicity test and the LD50 cannot be calculated, the high dose groups shall be in the following order: a) 10 g/kg body weight; b) 100 times the possible human intake; c) Design the maximum gavage dose, and then set up medium and low dose groups. Set up a solvent control group. The positive control can be given at cyclophosphamide 40 mg/kg body weight orally or intraperitoneally (oral is preferred). 5.4 Test procedures and observation indicators 5.4.1 Administration of test substances Oral gavage. According to the cell cycle and the characteristics of the action of different substances, a preliminary test can be done first to determine the sampling time. The 30- hour test substance administration method is commonly used. That is, the test substance is administered twice with an interval of 24 hours, and the bone marrow sample is collected 6 hours after the second administration of the test substance. The experiment can also have the following two sampling methods: a) Administer the test substance to the animal once. Collect bone marrow samples at least twice at appropriate intervals, starting no earlier than 24 hours after administration and ending no later than 48 hours after administration. The reason for sampling earlier than 24 hours after administration shall be stated. Collect peripheral blood samples at least twice at appropriate intervals, starting no earlier than 36 hours after administration and ending no later than 72 hours after administration. If a positive result is found at one sampling time, no further sampling is required. b) Administer once a day for 2 or more times (24 h intervals). Bone marrow can be collected once between 18 h and 24 h after the last administration, and peripheral blood can be sampled once between 36 h and 48 h after the last administration. If the rate of micronucleated cells in peripheral blood normochromatic erythrocytes is used as the experimental observation endpoint, the animal shall be administered for more than 4 weeks. 5.4.2 Specimen preparation 5.4.2.1 Bone marrow sample: After killing, take the sternum or femur; use hemostatic forceps to squeeze out the bone marrow fluid and mix it with the calf serum at one end of the slide; make a conventional smear. Alternatively, use calf serum to rinse the femoral bone marrow cavity to make a cell suspension smear; after the smear is naturally dried, put it in methanol to fix it for 5 min ~ 10 min. Store it after fixation on but there is no dose-response relationship, the test shall be repeated. Repeatable results can be considered positive.7 Test report7.1 Test name, test unit name and contact information, and report number. 7.2 Name and contact information of the test entrusting unit, and sample acceptance date. 7.3 Test start and end dates, test item leader, test unit technical leader, and issue date. 7.4 Test summary. 7.5 Test substances: name, batch number, dosage form, status (including sensory, properties, packaging integrity, and labeling), quantity, pretreatment method, and relevant information of the positive control. 7.6 Experimental animals: species, strain, grade, quantity, age, weight, sex, source (supplier name, experimental animal production license number), animal quarantine, adaptation, breeding environment (temperature, relative humidity, experimental animal facility use license number), feed source (supplier name, experimental animal feed production license number). 7.7 Test method: test grouping, number of animals in each group, dose selection basis, test substance administration route and period, sampling time point, specimen preparation method, number of cells observed per animal, statistical methods and judgment standards. 7.8 Test results: Record the number of polychromatic erythrocytes and the number of micronucleated cells observed in each animal, and report the number of polychromatic erythrocytes in each group of animals of different genders, the rate of micronucleated cells and the proportion of polychromatic erythrocytes in total red blood cells, the dose- response relationship, the historical data and range of negative controls in tabular form; state the statistical method for the results. 7.9 Test conclusion: Based on the test results, make a conclusion on whether the test substance can cause an increase in the rate of micronucleated cells in mammalian polychromatic erythrocytes.8 Interpretation of the testA positive result indicates that the test sample can cause an increase in the rate of micronucleated cells in mammalian polychromatic erythrocytes under the test conditions. A negative result indicates that the test sample does not cause an increase in the rate of micronucleated cells in mammalian polychromatic erythrocytes under the ......Source: Above contents are excerpted from the full-copy PDF -- translated/reviewed by: www.ChineseStandard.net / Wayne Zheng et al. Tips & Frequently Asked Questions:Question 1: How long will the true-PDF of English version of GB 15193.5-2014 be delivered?Answer: The full copy PDF of English version of GB 15193.5-2014 can be downloaded in 9 seconds, and it will also be emailed to you in 9 seconds (double mechanisms to ensure the delivery reliably), with PDF-invoice.Question 2: Can I share the purchased PDF of GB 15193.5-2014_English with my colleagues?Answer: Yes. The purchased PDF of GB 15193.5-2014_English will be deemed to be sold to your employer/organization who actually paid for it, including your colleagues and your employer's intranet.Question 3: Does the price include tax/VAT?Answer: Yes. 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