人参与林蛙油配伍的保健食品体外致突变试验研究

目的体外观察人参与林蛙油配伍的保健食品能否诱发鼠伤寒沙门氏菌组氨酸营养缺陷型突变株的回复突变。方法 Ames试验用TA97、TA98、TA100、TA102菌株并分加与不加肝微粒体酶活化系统（＋/-S9）试验。结果人参与林蛙油配伍的保健食品对4个菌株各剂量组的回复菌落数都未超过自然回复菌落数,Ames试验结果为阴性。结论在5000-8μg/皿剂量下未见人参与林蛙油配伍的保健食品致突变性。     

中药提取物的抗突变作用

近来,已对一些常用中药的抗突变作用进行了研究.本文报道从日本第10版药典中选出的一些中药,用Ames试验对其抗突变作用进行评价.选出的药材为甘草根、芍药根、柴胡根、茯苓、桔梗和人参根.结果表明,芍药根和茯苓对TA98和TA100两种菌株有相似的作用,其存活菌落数不受提取物浓度的影响,但回复变异数随浓度的增加而明显减少.说明这两种药材的提取物可能含有对抗苯并芘致突变的成分.在柴胡根的实验中,在TA98菌株中显示了抗突变作用,而在TA100菌株中回复变异菌落     

受试物中组氨酸含量对Ames试验自发回变菌落数的影响

目的探索试验样品组氨酸含量对Ames试验中测试菌株TA97 a、TA98、TA100、TA102和TA1535自发回变菌落数的影响。方法使用0.8、0.4、0.2、0.1和0.05 mg/ml 5个浓度的组氨酸水溶液作为受试物按平板掺入法进行Ames试验。结果在一定浓度范围内,随着组氨酸剂量的增高,5个菌株的回变菌落数均逐渐增多。TA98和TA1535的回变菌落数分别在0.8和0.2 mg/ml时超过阴性对照组的2倍。所有测试菌株在0.2 mg/ml时均出现了与对照组间的差异有统计学意义。结论在进行Ames试验前,对于受试物中游离组氨酸含量的判定十分必要;当受试物中组氨酸含量高于0.2 mg/ml时,有可能造成"假阳性"结果的产生,建议采用以基因突变为检测终点的其他试验进行替代。     

微型细菌回复突变试验方法的建立及验证

目的 建立高通量评价药物遗传毒性的微型细菌回复突变（Ames）试验。方法 采用平皿掺入法进行标准Ames试验,采用6孔板掺入法进行微型Ames试验,两试验均设置阴性对照组和阳性对照组,2组又分别分为代谢活化和非活化2个亚组,各组培养基中分别加入菌液（鼠伤寒沙门菌组氨酸缺陷型菌株：TA97、TA98、TA100、TA102和TA1535）,活化组加大鼠肝微粒体酶（S9）混合液,阳性对照组再加入阳性诱变剂：Dexon（-S9）,应用于TA97、TA98和TA102;NaN₃（-S9）,应用于TA100和TA1535;2-AA（+S9）,应用于TA97、TA98、TA100、TA102和TA1535。凝固后37℃温箱培养约48 h,计数各组回变菌落数。结果 在标准Ames试验中,阴性对照组各个菌株回变菌落数均在本实验室自发回变菌落数正常值范围内。在两试验中,阳性对照组所诱导的回变菌落数均是阴性对照组的2倍以上,标准试验细菌回变菌落数约为微型试验的5倍。结论 在6孔板上进行微型细菌回复突变试验,大大减少了受试物的用量,提高了筛选速度。本实验室建立的微型细菌回复突变试验背景数据,用于遗传毒理的检测是高效可靠的。     

基于毒理学软件和细菌回复突变试验的大黄素型蒽醌基因突变风险评价

目的 使用毒理学软件和细菌回复突变（Ames）试验评价大黄素型蒽醌类化合物的致突变风险，分析不同取代基及所在位置对大黄素型蒽醌致突变风险的影响。方法 使用Toxtree、Derek Nexus和Sarah Nexus毒性预测软件对大黄素、羟基大黄素、芦荟大黄素、大黄素甲醚、大黄酚、大黄酸、大黄素-8-O-β-D-葡萄糖苷、芦荟大黄素-8-O-β-D-葡萄糖苷、大黄素-1-O-β-D-葡萄糖苷、大黄素甲醚-8-O-β-D-葡萄糖苷的致突变风险进行预测，并使用鼠伤寒沙门氏菌TA97、TA98、TA100、TA102、TA1535和TA1537及大肠杆菌WP2 uvrA开展基于6孔板的Ames试验，评价10种大黄素型蒽醌的致突变性。结果 基于蒽醌母核结构，Toxtree、Derek Nexus和Sarah Nexus毒性预测软件提示所有大黄素型蒽醌均存在致突变风险。在非S9代谢活化状态下，芦荟大黄素导致TA98和WP2 uvrA Ames菌落数增加，大黄酚、大黄酸导致WP2 uvrA Ames菌落数增加。在大鼠肝S9代谢活化状态下，大黄素和大黄酸导致TA98和TA1537 Ames菌落数增加，羟基大黄素导致TA97、TA98、TA1537和WP2 uvrA Ames菌落数增加，芦荟大黄素导致TA98、TA1537和WP2 uvrA Ames菌落数增加，大黄素甲醚导致TA1537 Ames菌落数增加，大黄酚导致TA1537和WP2 uvrA回复菌落突变数增加，大黄素-8-O-β-D-葡萄糖苷可引起TA1537回复突变菌落数增加。结论 大黄素型蒽醌类化合物在大黄素母核的基础引入羟基后其诱变能力显著升高，较大葡萄糖苷基团的引入反而使受试物诱变能力降低。     

微孔板与标准平皿Ames试验比较研究

目的 使用鼠伤寒沙门菌和大肠埃希菌及阳性剂开展基于6孔板、24孔板和标准10 cm平皿的Ames试验，为确立标准化微孔板Ames试验奠定研究基础。方法 6种不同鼠伤寒沙门菌（TA97、TA98、TA100、TA102、TA1535、TA1537）和1种大肠埃希菌（WP2 uvrA）经鉴定及扩增后，分别使用标准平皿、6孔板和24孔板在有无S9代谢活化条件下使用不同浓度的阳性剂开展平板掺入法Ames试验。所有样本置37℃培养约48 h后进行菌落计数。结果 所有试验条件下均出现浓度相关性的回复菌落形成率增加，且最高浓度条件下的菌落数高于对照组3倍。然而微孔板中可出现阴性背景菌落数过少的情况，阳性剂的用量也不能照搬标准平皿中的浓度设置。结论 本研究所用条件可成功开展基于6孔板和24孔板的Ames试验，其试验条件和背景数据需要经过摸索与积累。     

降血糖活性成分Bellidifolin遗传毒性研究

目的研究降血糖活性成分Bellidifolin的遗传毒性。方法整体试验采用小鼠骨髓嗜多染红细胞微核实验；体外试验采用鼠伤寒沙门菌组氨酸营养缺陷型TA97、TA98、TA100、TA102四个菌株，对Bellidifolin进行Ames实验。结果小鼠骨髓嗜多染红细胞微核发生率结果显示Bellidifolin高、中、低剂量组与阴性对照组比较均差异无显著性(均P＞0.05)；Ames实验显示,在实验设置浓度和加S9或不加S9的实验条件下，受试物对各菌株所诱发的回变菌落数,均未超过对照的2倍。结论实验结果为阴性，未见Bellidifolin有致突变性作用。     

灵丹菌质的3次遗传毒性实验研究

目的研究灵丹菌质的遗传毒性,为其开发利用提供依据。方法利用Ames试验、小鼠精子畸形试验以及骨髓细胞微核试验评价灵丹菌质的遗传毒性。Ames试验设立8、40、200、1 000和5 000μg/皿5个剂量组,计算各剂量组在加与不加体外代谢活化系统S9的情况下,对鼠伤寒沙门氏菌TA97、TA98、TA100和TA102的诱发回变菌落数。小鼠精子畸形试验设5、10和20 ml/kg·BW 3个剂量组,计算各剂量组小鼠精子畸形率。小鼠骨髓细胞微核设5、10和20 ml/kg·BW 3个剂量组计算各剂量组微核发生率。结果受试物各剂量组4种菌株的回变菌落数均未超过未处理对照组的2倍;受试物各剂量组的精子畸形率和微核细胞率均未见统计学意义。结论在本研究条件下,灵丹菌质未见遗传毒性。     

茜素型蒽醌基因突变风险评价

目的 使用毒性预测软件及细菌回复突变（Ames）试验评价茜素型蒽醌的基因突变风险。方法 通过毒性软件Toxtree、Derek Nexus和Sarah Nexus对茜素型蒽醌：茜草素、异茜草素、甲基异茜草素、甲基异茜草素-1-甲醚、茜素-1-甲醚、羟基茜草素、光泽汀进行致突变风险预测;每个受试物设置5个给药浓度,分别在有或无S9代谢活化条件下,使用5种鼠伤寒沙门氏菌TA97、TA100、TA102、TA1535和TA1537开展基于6孔板培养的Ames试验,判断该类化合物苯环上不同取代基对致突变性的影响。结果 软件基于蒽醌环的存在预测该类化合物均具有致突变风险。在非S9代谢活化下,异茜草素和羟基茜草素可导致TA1537回复突变菌落数增加;光泽汀可诱导TA97、TA100和TA1537回复突变菌落数增加。在S9代谢活化下,异茜草素可导致TA97、TA100和TA1537回复突变菌落数增加;羟基茜草素可导致TA1537回复突变菌落数增加;光泽汀可导致TA97、TA100和TA1537回复突变菌落数增加;甲基异茜草素可导致TA97、TA100、TA102和TA1537回复突变菌落数大幅增加;甲基异茜草素-1-甲醚可导致TA100回复突变菌落数增加。结论 茜素型蒽醌受试物在有或无S9代谢条件下表现出不同程度、不同菌株的回复突变,开展相关研究评价其毒性风险对该类化合物合理监管具有重要价值。     

葛根素致突,致畸作用评价

<正> 葛根素(puerarinum)为山东省医科院药物研究所从豆科植物野葛根中提取的一种异黄酮类化合物精制成葛根素注射液,是一种β受体拮抗剂.为确保临床应用的安全,进行了药物的短期致突及致畸试验,其结果如下: Ames试验 采用TA97,TA98,TA100,TA102为测试菌株,选取1,5,50,500,5000μg/皿,并设空白,溶剂及阳性对照.结果表明:各实验组与阴性对照组,在加S9和不加S9的实验中,其回变菌落数基本相     

Evaluation of beta-myrcene, alpha-terpinene and (+)- and (-)-alpha-pinene in the Salmonella/microsome assay.

This study was undertaken to investigate the genotoxicity of beta-myrcene, alpha-terpinene and (+) and (-)-alpha-pinene, monoterpenes found in a variety of plant volatile oils. beta-myrcene, alpha-terpinene and alpha-pinene as well as plant oils containing these hydrocarbon monoterpenes have been used as flavoring additives in foods and beverages, as fragrances in cosmetics, and as scent in household products. Mutagenicity was evaluated by the Salmonella/microsome assay (TA100, TA98, TA97a and TA1535 tester strains), without and with addition of an extrinsic metabolic activation system (rat liver S9 fraction induced by Aroclor 1254). Two dose-complementary assays were performed so that a broad range of doses, including a number of regularly-spaced doses in the non-toxic dose interval, were tested. No increase in the number of his+ revertant colonies over the negative control values was observed in any of the four S. typhimurium tester strains. Results from the present study therefore indicated that beta-myrcene, alpha-terpinene, and (+) and (-)-alpha-pinene are not mutagenic in the Ames test.     

Mutagenicity of bovine lactoferrin in reverse mutation test

The mutagenicity of bovine lactoferrin, which is an iron-binding glycoprotein in milk, was evaluated by the Ames mutagenicity test. A total of 5 test strains including 3 base-pair substitution-type strains, Salmonella typhimurium TA100, TA1535 and Escherichia coli WP2uvrA, and 2 frameshift-type strains, TA98 and TA1537, were used in the test. The test was performed by both the direct method and the metabolic activation method with preincubation applied in each instance. The concentration range of the test solution was 0.16 to 5.00 mg/100 microliters (plate). Results of the test revealed that the number of revertant colonies at each concentration of the test solutions was less than 1.4 times that of the control group. In the test system used, bovine lactoferrin did not exhibit mutagenicity.     

Mutagenicity evaluation of riot control agent o-chlorobenzylidene malononitrile (CS) in the Ames Salmonella/microsome test.

o-Chlorobenzylidene malononitrile (CS), a riot control agent, was evaluated for its possible mutagenic activity in the Ames Salmonella/mammalian microsome mutagenicity test. Five histidine-deficient (His-) mutant tester strains of Salmonella typhimurium--TA97a, TA98, TA100, TA102 and TA104--were used. The liquid preincubation procedure was used with metabolic activation (presence of S9 mixture) and without metabolic activation (absence of S9 mixture). For the experiments with metabolic activation, three different concentrations of S9 fraction (supernatant of Aroclor 1254-induced rat liver homogenate at 9000 g)--5%, 15% and 30% in S9 mixture--were used. Along with mutagenic activity, CS was also evaluated for cytotoxic activity in all the five tester strains of Salmonella typhimurium, both in the presence and absence of S9 mixture. The mutagenic and cytotoxic activities of CS were assessed by counting the His+ revertant colonies and by counting the microcolonies (His-, auxotrophs in the background lawn), respectively, and the respective mean values were compared with the relative negative (solvent) control. A dose range of 12.5-800 micrograms plate-1 for CS did not induce a mutagenic response either in the presence or absence of S9 mix. No change in the negative mutagenic response of CS has been observed even in the presence of an elevated level of S9 fraction in the S9 mix. A dose of 200 micrograms plate-1 for CS was found to be cytotoxic by decreasing the surviving cells as well as His+ revertant colonies; however, the effect was reduced in the presence of an elevated level of S9 fraction in the S9 mix.     

Tofisopam--evaluation of mutagenic and genotoxic properties

The mutagenic properties of tofisopam, the member of the 2,3-benzodiazepine family, were evaluated on the basis of Ames test with Salmonella typhimurium TA1537, TA97, TA98, TA100 and TA102 strains. The genotoxic properties of tofisopam were estimated on L929 cell line with the cytokinesis-block technique. Under the experimental conditions, no mutagenic activity of tofisopam in tester bacteria strains was found, and no genotoxic activity was observed.     

Studies of the toxicological potential of tripeptides (L-valyl-L-prolyl-L-proline and L-isoleucyl-L-prolyl-L-proline): IX. Evaluation of the mutagenic potential of synthesized L-valyl-L-prolyl-L-proline in the Salmonella-Escherichia coli/microsome, incorporation assay

The objective of this study was to assess the mutagenic potential of a synthesized tripeptide, L-valyl-L-prolyl-L-proline (VPP), to induce mutational changes in Salmonella typhimurium LT2 strains TA1535, TA1537, TA98, and TA100, and Escherichia coli strain WP2uvrA in the classical Ames test protocol. Bacteria were exposed to plate concentrations of VPP of 0, 156.2, 312.5, 625, 1250, 2500, and 5,000 microg/plate in distilled water, in the presence and absence of Aroclor 1254-induced rat liver homogenate preparation (S9). Positive-control agents included sodium azide (TA100 and TA1535); 2-aminoanthracene (TA98, TA100, TA1535, TA1537, and WP2uvrA); 9-aminoacridine (TA1537); 2-nitrofluorene (TA98); and N-ethyl-N'-nitro-N-nitrosoguanidine (WP2uvrA) in DMSO. Incubations were conducted at 37 degrees C for about 48 h then revertant colonies were counted. All positive-control agents were consistently and unequivocally positive, but there was no evidence that VPP induced increases in the incidences of revertant colonies in any bacterial strain with and without metabolic activation. These findings were replicated in a second, confirmatory test performed with and without S9. The results of the experiments revealed no treatment-associated changes in the incidence of revertant colonies in any bacterial strain tested. These results support a conclusion that, under the experimental conditions described, there is no evidence that VPP possesses mutagenic potential.     

赤苷脉通注射液的致突变研究

目的探讨中药制剂赤苷脉通注射液的致突变性。方法采用鼠伤寒沙门氏组氨酸营养缺陷型菌株回复突变实验(Ames实验)、中国仓鼠肺成纤维细胞(CHL)染色体畸变实验和小鼠骨髓微核实验来检测赤苷脉通注射液的致突变作用。结果 Ames实验中,赤苷脉通注射液在312.5～5 000μg.皿-1剂量范围内,无论加或不加S9,鼠伤寒沙门氏菌组氨酸缺陷型TA97,TA98,TA100,TA102和TA1535 5株菌的回复突变菌落数均未出现剂量依赖性的增加;染色体畸变实验中,非活化条件或代谢活化条件下,药物质量浓度为1 200,600和300μg.mL-1时,细胞的染色体畸变率均未出现剂量依赖性增加;微核实验中,在1 150,575和287.5mg.kg-1剂量组中均未见骨髓中含微核的嗜多染红细胞数增加。结论在该实验室条件下,Ames实验、CHL细胞染色体畸变实验和小鼠骨髓微核实验结果均为阴性,即中药制剂赤苷脉通注射液无潜在的遗传毒性。     

Mutagenicity study of gadobenate dimeglumine formulation (E7155) (1)--Reverse mutation assays in S. typhimurium and E. coli tester strains

The ability of gadobenate dimeglumine formulation (E7155) to cause gene mutations was assessed in five strains of Salmonella typhimurium (TA100, TA1535, TA98, TA1538, and TA1537) and a strain of Escherichia coli (CM891; WP2, uvrA-, pKM101) using the Ames test (agar plate assay). The results suggest that E7155 is non-mutagenic towards these bacterial tester strains.     

Mutagenicity tests on propiverine hydrochloride

1. The reverse mutation test was carried out on propiverine hydrochloride (P-4) at dose range of 5-500 micrograms/plate use Salmonella typhimurium strains, TA100, TA98, TA1535 and TA1537, and Escherichia coli strain WP2uvrA. As compared with solvent-treated control, no significant increases were observed in the number of revertant colonies in all tester strains in both systems with and without mammalian metabolic activation (S9 Mix). 2. The chromosomal aberration test was also carried out on P-4 using cultured Chinese hamster lung cells (CHL). The cells were treated with P-4 at the doses of 5, 10, 20 and 40 microM without S9 Mix and at 62.5, 125, 250 and 500 microM with S9 Mix. No significant differences were found in the incidence of structural- and numeral-aberrations of chromosomes in both systems with and without S9 Mix. 3. These results indicate that P-4 has no mutagenic activity.     

Evaluation of the safety of the dietary antioxidant ergothioneine using the bacterial reverse mutation assay

The dietary antioxidant l-(+)-ergothioneine was tested for its potential mutagenic activity using the bacterial reverse mutation assay. The experiments were carried out using histidine-requiring auxotrophic strains of Salmonella typhimurium (Salmonella typhimurium TA98, TA100, TA1535 and TA1537), and the tryptophan-requiring auxotrophic strain of Escherichia coli (Escherichia coli WP2 uvrA) in the presence and absence of a post-mitochondrial supernatant (S9) prepared from livers of phenobarbital/β-naphthoflavone-induced rats. The revertant colony numbers of vehicle control plates with and without S9 Mix were within the corresponding historical control data ranges. The reference mutagen treatments (positive controls) showed the expected, biologically relevant increases in induced revertant colonies in all experimental phases in all tester strains. No biologically relevant increases were observed in revertant colony numbers of any of the five test strains following treatment with l-(+)-ergothioneine at any concentration level, either in the presence or absence of metabolic activation (S9 Mix) in the performed experiments. On the basis of the data reported, it can be concluded that l-(+)-ergothioneine did not induce gene mutations by base pair changes or frameshifts in the genome of the strains used. Thus l-(+)-ergothioneine has no mutagenic activity on the applied bacteria tester strains under the test conditions used in this study. Research is continuing to define the role of l-(+)-ergothioneine in disease pathophysiology. Further studies on its safety are suggested.