Abstract: Objective To evaluate the gene mutations caused by the potential genotoxicity of monanthone with emodin type by means of the miniAmes test and in vitro Pig-a gene mutation test. Methods The miniAmes test involved the control group (DMSO), monanthone with emodin type (0.6, 1.1, 2.3, 4.5, 9 μg/well), and positive group. Salmonella typhimurium TA97, TA98, TA100, TA102, TA1535, TA1537 bacterial reverse mutation assay based on 6-well plate culture was carried out in E. coli WP2 uvrA under the conditions of -S₉ and S₉, respectively. The number of mutant colonies was counted after 48 h. The In vitro Pig-a gene mutation test involved L5178Y cells that were used as the system, and metabolic conditions without S₉: control group (1% DMSO), positive agent (EMS 500 μg/mL), and monanthone with emodin type (0.2 μg/mL,0.39 μg/mL,0.78 μg/mL,1.56 μg/mL).After the test object was treated for 4 h, the test substance and culture were removed. After 24 h, the cells were counted and expressed for 8 days. The cell density during the expression period was maintained at 1×10⁶~2×10⁶ cells/mL. After the expression was terminated, antibody incubation flow detection was performed. Metabolic conditions with S₉: control group (1% DMSO), positive agent (B(a)P 5 μg/mL), and monoterpene ketone (0.2 μg/mL,0.39 μg/mL,0.78 μg/mL,1.56 μg/mL). After the test object was treated for 4 h, the test material was removed and cultured, and counted 24 hours later for 8 days. The cell density during the expression period was 1×10⁶~2×10⁶ cells/mL. After the expression was over, the antibody incubation flow assay was performed. Results In the Ames test, the number of colonies of TA97 and TA1537 reverting mutants induced by monanthone with emodin type increased compared with the negative control group under the metabolic activation conditions without S₉. Upon metabolic activation with S₉, the number of colonies of TA1537 reverting mutants induced by monanthone with emodin type increased compared with the negative control group. The rate of increase was more than two or three times that of the negative control group, and there was dose-related change in the above results. In vitro Pig-a gene mutation test showed that under metabolic activation conditions without S₉, the concentration of monanthone with emodin type exceeded 0.78 μg/mL, and the mutation frequency of Pig-a gene was significantly different from that of the vehicle control group (P <0.01) with a dose correlation. Conclusion Under the conditions described in this study, monanthone with emodin type has the risk of gene mutation.

Key words: Polygonummultiflorum Thunb, monanthone with emodin type, gene mutation, genotoxicity, Ames test, in vitro Pig-a gene mutation