Evaluation of the genotoxicity of (-)-hydroxycitric acid (HCA-SX) isolated from Garcinia cambogia

(-)-Hydroxycitric acid (HCA) is widely used as an ingredient for nutritional supplements aimed at reducing food intake, appetite, and body weight. In this study, the genotoxicity of HCA was evaluated using three tests: a bacterial reverse mutation assay (Ames test), an in vitro chromosomal aberration (CA) test, and an in vivo micronucleus (MN) test. HCA was negative by the Ames test in the presence or absence of a microsomal metabolizing system. HCA did not induce mutagenic activity in the Ames test, and no significant mutagenic potency was indicated by CA tests. However, HCA significantly and dose-dependently increased the number of MNPCEs (micronucleated polychromatic erythrocytes/1000 polychromatic erythrocytes) and PCE/(PCE + NCE) ratios according to the MN test. These results suggest that HCA preferentially induce micronuclei.     

Genotoxicity evaluation of Isaria sinclairii (ISE) extract

The mutagenic potential Isaria sinclairii, a traditional Chinese medicine composed of the fruiting bodies of I. sinclairii and its parasitic host larva, was evaluated using short-term genotoxicity tests, namely, the Ames test, chromosome aberration (CA), and micronuclei (MN) tests. In a Salmonella typhimurium assay, I. sinclairii extract (ISE) did not produce any mutagenic response in the absence or presence of 59 mix with TA98, TA100, TA1535, and TA1537. In the chromosome aberration (CA) test, ISE induced no significant effect on Chinese hamster ovary (CHO) cells compared with control. In the MN test, no significant change in the occurrence of micronucleated polychromatic erythrocytes was observed in male ICR mice intraperitoneally administered ISE at doses of 15, 150, or 1500 mg/kg. These results indicate that ISE has no mutagenic potential in these in vitro and in vivo systems.     

Study of mutagenicities of phthalic acid and terephthalic acid using in vitro and in vivo genotoxicity tests

Genotoxicities of phthalic acid (PA) and terephthalic acid (TPA) were examined using three mutagenicity tests: Ames, chromosome aberration (CA), and micronucleus (MN). In the Ames test, these two agents did not produce any mutagenic responses in the absence or presence of S9 mix on the Salmonella typhimurium strains TA98, TA100, TA102, TA1535, or TA1537. The CA test also showed that PA and TPA exerted no significant cytogenetic effect on Chinese hamster ovary (CHO) cells. In the mouse MN test, no significant alteration in occurrence of micronucleated polychromatic erythrocytes was observed in ICR male mice ip administered any of these agents at doses of 0, 20, 100, 500, 2500 or 12,500 microM/kg. These results indicate that PA and TPA produced no mutagenic effects using these in vitro and in vivo mutagenic test systems.     

Toxicological evaluation of an apicidin derivative, histone deacetylase inhibitor SD-2007 in mice

SD-2007 is a new derivative of apicidin, an anti-parasitic agent and a histone deacetylase (HDAC) inhibitor. A subacute toxicological evaluation of SD-2007 was investigated for 2 weeks in ICR mice. After oral administration of SD-2007 (0, 0.2, 1, 5 or 25 mg/mouse), the clinical signs, mortalities, body weight changes, blood biochemical parameters, absolute and relative organ weights were examined. One day after the administration of SD-2007, excretion of soft feces in 1 and 5 mg/head groups, and one male in 25 mg/mouse group developed diarrhea, but theses complications were disappeared two days after administration. No mortalities were observed in animals up to 25 mg/mouse (LD₅₀, >25 mg/kg), but absolute and relative weights of testes were significantly lower at the highest dose group (25 mg/mouse) and serum LDH and glucose levels were elevated in male mice. In addition, alanine aminotransferase (ALT), aspartate aminotransferase (AST), and alkaline phosphatase (ALP) activities were reduced in female mice at all dosages. These data suggest that SD- 2007 may be sex specific and be toxic to the male reproductive organ, and thus our findings require further investigation and in particular chronic toxicological investigations should be investigated.     

In vitro and in vivo studies on the potential mutagenicity of alclofenac, dihydroxyalclofenac and alclofenac epoxide

Alclofenac (A) and two of its metabolites, dihydroxyalclofenac (DHA) and alclofenac epoxide (AE), were tested for their mutagenic potential. Alclofenac and DHA showed no mutagenic, transforming or clastogenic potential in any in vitro experiment. The addition of a supplementary metabolic activation system did not change the response of those two compounds in any test procedures in vitro. AE, an intermediary metabolite between A and DHA, was mutagenic by itself in the Ames test, but in the presence of a liver post-mitochondrial fraction its activity of Salmonella typhimurium was greatly decreased. Dominant lethal mutations were not induced in male rats given alclofenac, and AE had no effect in the micronucleus test. In human volunteers given alclofenac at therapeutic dose levels, no mutagenic activity was found in the urine and no significant increase in the incidence of structural chromosome aberrations was observed in peripheral blood lymphocytes.     

Antigenotoxicity properties of Copaifera multijuga oleoresin and its chemical marker,the diterpene (−)-copalic acid

In view of the biological activities and growing therapeutic interest in oleoresin obtained from Copaifera multijuga, this study aimed to determine the genotoxic and antigenotoxic potential of this oleoresin (CMO) and its chemical marker, diterpene (?)-copalic acid (CA). The micronucleus (MN) assay in V79 cell cultures and the Ames test were used for in vitro analyses, as well as MN and comet assays in Swiss mice for in vivo analyses. The in vitro genotoxicity/mutagenicity results showed that either CMO (30, 60, or 120 µg/ml-MN assay; 0.39–3.12 mg/plate-Ames test) or CA (2.42; 4.84, or 9.7 µg/ml-MN assay; 0.39–3.12 mg/plate-Ames test) did not induce a significant effect on the frequency of MN and number of revertants, demonstrating an absence of genotoxic and mutagenic activities, respectively, in vitro. In contrast, these natural products significantly reduced the frequency of MN induced by methyl methanesulfonate (MMS), and exerted a marked inhibitory effect against indirect-acting mutagens in the Ames test. In the in vivo test system, animals treated with CMO (6.25 mg/kg b.w.) exhibited a significant decrease in rate of MN occurrence compared to those treated only with MMS. An antigenotoxic effect of CA was noted in the MN test (1 and 2 mg/kg b.w.) and the comet assay (0.5 mg/kg b.w.). Data suggest that the chemical marker of the genus Copaifera, CA, may partially be responsible for the observed chemopreventive effect attributed to CMO exposure.

Abbreviations: 2-AA, 2-anthramine; 2-AF, 2-aminofluorene; AFB₁, aflatoxin B₁; B[a]P, benzo[a]pyrene; BOD, biological oxygen demand; BPDE, benzo[a]pyrene-7,8-diol-9,10-epoxide; CA, (?)-copalic acid; CMO, oleoresin of Copaifera multijuga, DMEM, Dulbecco`s Modified Eagles`s Medium; DMSO, dimethylsulfoxide; EMBRAPA, Brazilian agricultural research corporation; GC–MS, gas chromatography–mass spectrometry; HAM-F10, nutrient mixture F-10 Ham; HPLC, high performance liquid chromatography; LC–MS, liquid chromatography–mass spectrometry; MI, mutagenic index; MMC, mitomycin C; MMS, methyl methanesulfonate; MN, micronucleus; MNPCE, micronucleated polychromatic erythrocyte; NCE, normochromatic erythrocyte; NDI, nuclear division index; NMR, nuclear magnetic resonance; NPD, 4-nitro-o-phenylenediamine; PBS, phosphate-buffered saline; PCE, polychromatic erythrocyte; SA, sodium azide; V79, Chinese hamster lung fibroblast.     

Assessment of combinations of antiandrogenic compounds vinclozolin and flutamide in a yeast based reporter assay

Glycyrrhiza glabra Linn. (licorice) is widespread throughout the Mediterranean region and certain areas of Asia. Historically, the dried rhizome and root of the plant were used by the Chinese, Egyptian, Greek, Indian, and Roman civilizations as expectorant and carminative. In the modern medicinal system, licorice is used to treat liver ailments, dyspepsia, bronchitis, rheumatoid arthritis etc. Despite the extensive pharmacological applications, the genotoxic potential of G. glabra extract (GutGard™) has not been evaluated. Hence, this study was conducted to investigate the genotoxic potential of GutGard™ using battery of in vitro test systems: bacterial reverse mutation test (Ames II™), chromosome aberration (CA) and micronucleus (MN) tests. GutGard™ did not show significant increase in number of revertant colonies in Salmonella typhimurium strains (TA98 and TAMix) with/without S9 fraction. In CA and MN studies, GutGard™ did not show clastogenic effect at 4 and 18 h treatments with and without S9 fraction. Results indicated that GutGard™ is not mutagenic in a battery of genotoxicity tests.     

Genotoxicity of aspartame

In the present study, the genotoxic effects of the low-calorie sweetener aspartame (ASP), which is a dipeptide derivative, was investigated using chromosome aberration (CA) test, sister chromatid exchange (SCE) test, micronucleus test in human lymphocytes and also Ames/Salmonella/ microsome test. ASP induced CAs at all concentrations (500, 1000 and 2000 microg/ml) and treatment periods (24 and 48 h) dose-dependently, while it did not induce SCEs. On the other hand, ASP decreased the replication index (RI) only at the highest concentration for 48 h treatment period. However, ASP decreased the mitotic index (MI) at all concentrations and treatment periods dose-dependently. In addition, ASP induced micronuclei at the highest concentrations only. This induction was also dose-dependent for 48 hours treatment period. ASP was not mutagenic for Salmonella typhimurium TA98 and TA100 strains in the absence and presence of S9 mix.     

Evaluation of the mutagenic and genotoxic activities of anti-hepatitis B analogs of beta-L-adenosine by the Ames test and the Comet assay

beta-L-2'-deoxyadenosine (beta-L-dA), beta-L-2',3'-dideoxyadenosine (beta-L-ddA) and its two bis (S-acyl-2-thioethyl; SATE) phosphotriester derivatives, beta-L-2',3'-dideoxyadenosine-5'-monophosphate-bis(MeSATE) and beta-L-2',3'-dideoxyadenosine-5'-monophosphate-bis(tButylSATE) have been previously shown to exhibit potent and selective anti-hepatitis B activity in vitro. None of the four compounds was mutagenic up to 100 microg in the Ames test (microtechnique) using Salmonella typhimurium strains TA 97a, TA 98, TA 100 and TA 102, with and without metabolic activation. In addition, the genotoxicity of beta-LdA and the three other compounds was evaluated in human lymphocytes using the Comet assay, at doses up to 5 microg with or without the addition of a microsomal S9 fraction. None of the four compounds induced DNA strand breakage with and without metabolic activation. In summary, the data clearly demonstrate that the purine nucleoside beta-L-dA, beta-L-ddA and the two prodrugs, beta-L-ddAMP-bis(MeSATE) and beta-L-ddAMP-bis(tButylSATE) are not mutagenic in the Ames test and do not induce DNA damage in human lymphocytes, as assessed by the Comet assay.     

Tinospora cordifolia, a safety evaluation

Tinospora cordifolia is one of the indispensable medicinal plants used in veterinary folk medicine/Ayurvedic system of medicine for the treatment of diverse diseases and recommended for improving the immune system by means of body resistance. In the current study, we evaluated the genotoxic risk of the aqueous extract of T. cordifolia (TC) in a battery of four different genotoxicity tests viz., Ames, in vitro chromosome aberration (CA), rodent bone marrow micronucleus (MN), and Comet assay. Experimental results confirmed that in Ames test up to 5000 μg/plate of TC did not exhibit any mutagenic effect in Salmonella typhimurium mutant strains (TA97a, TA98, TA100, TA102, and TA1535). In CA assay, TC was not clastogenic to human peripheral blood lymphocytes up to a concentration of 3000 μg/ml. In MN and Comet assays, TC was pre-treated for 7 days at three dose levels (150, 200 and 250 mg/kg body weight) orally to male Balb/c mice. The results showed that TC treatment did not display clastogenicity and DNA damaging effect in bone marrow erythrocytes and peripheral blood lymphocytes respectively.     

Genotoxicity of Aspartame

In the present study, the genotoxic effects of the low‐calorie sweetener aspartame (ASP), which is a dipeptide derivative, was investigated using chromosome aberration (CA) test, sister chromatid exchange (SCE) test, micronucleus test in human lymphocytes and also Ames/Salmonella/ microsome test. ASP induced CAs at all concentrations (500, 1000 and 2000 µg/ml) and treatment periods (24 and 48 h) dose‐dependently, while it did not induce SCEs. On the other hand, ASP decreased the replication index (RI) only at the highest concentration for 48 h treatment period. However, ASP decreased the mitotic index (MI) at all concentrations and treatment periods dose‐dependently. In addition, ASP induced micronuclei at the highest concentrations only. This induction was also dose‐dependent for 48 hours treatment period. ASP was not mutagenic for Salmonella typhimurium TA98 and TA100 strains in the absence and presence of S9 mix.     

Genotoxicity of quinocetone,cyadox and olaquindox in vitro and in vivo

Quinocetone (QCT) and Cyadox (CYA) are important derivative of heterocyclic N-oxide quinoxaline (QdNO), used actively as antimicrobial feed additives in China. Here, we tested and compared the genotoxic potential of QCT and CYA with olaquindox (OLA) in Ames test, HGPRT gene mutation (HGM) test in V79 cells, unscheduled DNA synthesis (UDS) assay in human peripheral lymphocytes, chromosome aberration (CA) test, and micronucleus (MN) test in mice bone marrow. OLA was found genotoxic in all 5 assays. In Ames test, QCT produced His⁺ mutants at 6.9 μg/plate in Salmonella typhimurium TA 97, at 18.2 μg/plate in TA 100, TA 1535, TA 1537, and at 50 μg/plate in TA 98. CYA produced His⁺ mutants at 18.2 μg/plate in TA 97, TA 1535, and at 50 μg/plate in TA 98, TA 100 and TA 1537. QCT was found positive in HGM and UDS assay at concentrations ⩾10 μg/ml while negative results were reported in CA test and MN test. Collectively, we found that OLA was more genotoxic than QCT and CYA. Genotoxicity of QCT was found at higher concentration levels in Ames test, HGM and UDS assays while CYA showed weak mutagenic potential to bacterial cells in Ames test.     

Antiarrhythmic and electrophysiological effects of SD-3212, a novel Na+ and Ca++ channel blocker,in anaesthetized dogs with myocardial infarction in comparison with its stereoisomer (SD-3211) and bepridil

Summary Antiarrhythmic and electrophysiological effects of SD-3212, a novel antiarrhythmic agent, which has both Na⁺ channel and Ca⁺⁺ channel blocking activites, were compared with those of its (+)-stereoisomer, SD-3211, which has only a Ca⁺⁺ channel blocking activity, and bepridil, a known Ca⁺⁺ channel blocker with additional Na⁺ channel blocking activity, using the two-stage coronary ligation induced arrhythmia (24h after the ligation of the left anterior descending coronary artery) and 7 day-old myocardial infarcted hearts in anaesthetized dogs.SD-3212 showed a dose-dependent antiarrhythmic effect on the two-stage coronary ligation induced arrhythmia. SD-3212 at a dose of 3 mg/kg reduced the arrhythmic ratio, i.e. ectopic beats per min divided by the sum of ectopic beats and sinus beats per min, significantly from 1 up to 12 min after the administration. Neither bepridil (1–6 mg/kg) nor SD-3211 (1 mg/kg) had an antiarrhythmic effect. SD-3212 (0.3–3 mg/kg) prolonged both the conduction time in the normal myocardium and the delayed potential in the infarcted myocardium in the 7 day-old myocardial infarcted hearts in anaesthetized dogs in a dose-dependent manner. This effect of SD-3212 was shown at coupling intervals of 150–1000 ms increasing with decreasing interval. In this respect, SD-3212 is similar to drugs which show fast recovery of V_max from use-dependent block such as lidocaine. Bepidril (1–6 mg/kg) also prolonged these parameters in a dose-dependent manner, however, the prolongation induced by bedripil was limited to shorter coupling intervals as compared with that induced by SD-3212. SD-3212 (0.1–1 mg/kg) did not show this prolonging effect. SD-3212 increased the refractory period in the infarcted zone to a small extent (not significantly) at all strengths tested between 0.5–4 mA and also in the normal zone between 0.2–1 mA to an even lesser extent than in the infarcted zone. Bepridil produced a significant increase of refractory period in the infarcted zone. In the normal zone, bepridil produced a non-significant, but greater increase of the refractory period as compared with SD-3212. SD-3211 did not affect the refractory period in the infarcted zone or in the normal zone. None of the three drugs produced a significant change in the excitation threshold.Thus, SD-3212 showed electrophysiological properties of a drug with fast recovery kinetics without producing a significant increase of refractory period, and these properties are very similar to those of class Ib antiarrhythmic agents such as lidocaine. The present study suggests that there might be a possibility of SD-3212 to become a safe and unique antiarrhythmic agent with suppresses both Na⁺ and Ca⁺⁺ inward current.Send offprint requests to S. Nagashima at the above address     

Detection of genotoxicity and mutagenicity of chlorthiophos using micronucleus,chromosome aberration,sister chromatid exchange,and Ames tests

Potential mutagenic and genotoxic effects of Chlorthiophos, an organophosphate pesticide, were evaluated using four standard assays. Five different concentrations of the pesticide were tested by an Ames test using Salmonella typhimurium strains TA97, TA98, TA100, and TA102, with and without S9 metabolic activation. No concentrations of Chlorthiophos showed mutagenic activity on the TA97, TA100, and TA102 strains, with and without S9 fraction, but were all mutagenic to the TA98 strain without S9. Sister chromatid exchange (SCE), chromosome aberration (CA), and micronucleus (MN) tests were used to investigate the genotoxic effects of Chlorthiophos in human peripheral lymphocytes treated with 25, 50, 100, and 200 µg/mL concentrations of Chlorthiophos for 24 and 48 h. The nuclear division index (NDI), replication index (RI), and mitotic index (MI) were also calculated to determine the cytotoxicity of Chlorthiophos. No increase in SCE frequency was seen for any treatment period or concentration, but Chlorthiophos at 200 µg/mL increased the frequency of CAs. Increases in MN formation were only observed at Chlorthiophos concentrations of 200 µg/mL following 24 and 48 h treatments. Chlorthiophos treatment reduced the MI and NDI significantly, but had no effect on the RI. © 2014 Wiley Periodicals, Inc. Environ Toxicol 30: 937–945, 2015.     

Genotoxicity assessment of NIM-76 and its formulation (pessary) in an in vitro Ames Salmonella/microsome assay and in vivo mouse bone marrow micronucleus test

Abstract

The possible genotoxic potential of NIM-76, a volatile fraction obtained from neem oil, having promising contraceptive activity, as well as its formulation product, called pessary (7.5% NIM-76 in polyethylene glycol), were evaluated in the Ames assay and mouse bone marrow micronucleus (MN) assay. Genotoxicity of NIM-76 (0.1–1000?µg/plate) and pessary (0.1–10?000?µg/plate) were studied using the liquid preincubation protocol of the Ames assay both in the presence and absence of S9. Likewise, the ability of NIM-76 [1–1000?mg/kg body weight (b.w.)] and its formulation product (18.75–300?mg/kg b.w.) to induce clastogenic effects were studied in the female mouse bone marrow MN test by using a two-dose intraperitoneal treatment protocol. There was no increase in the number of revertant colonies resulting from NIM-76 or pessary at any of their doses over the respective negative control plates, either in the presence or absence of S9. Similarly, in the MN assay, neither of them showed any clastogenic activity because there was no significant increase in the frequency of micronucleated polychromatic erythrocytes, over the negative control group of animals. The use of this compound in humans is therefore not likely to have mutagenic effects and may be considered as safe with regard to genotoxic potential.     

Genotoxicity assessment of propyl thiosulfinate oxide,an organosulfur compound from Allium extract,intended to food active packaging

Essential oils from onion (Allium cepa L.), garlic (Allium sativum L.), and their main components, such as propyl thiosulfinate oxide (PTSO) are being intended for active packaging with the purpose of maintaining and extending food product quality and shelf life. The present work aims to assess for the first time the potential mutagenicity/genotoxicity of PTSO (0–50 µM) using the following battery of genotoxicity tests: (1) the bacterial reverse-mutation assay in Salmonella typhimurium (Ames test, OECD 471); (2) the micronucleus test (OECD 487) (MN) and (3) the mouse lymphoma thymidine-kinase assay (OECD 476) (MLA) on L5178YTk^+/−, cells; and (4) the comet assay (with and without Endo III and FPG enzymes) on Caco-2 cells. The results revealed that PTSO was not mutagenic in the Ames test, however it was mutagenic in the MLA assay after 24 h of treatment (2.5–20 µM). The parent compound did not induce MN on mammalian cells; however, its metabolites (in the presence S9) produced positive results (from 15 µM). Data from the comet assay indicated that PTSO did not induce DNA breaks or oxidative DNA damage. Further in vivo genotoxicity tests are needed to confirm its safety before it is used as active additive in food packaging.     

蛞蝓胶囊致畸和致突变实验研究

目的探讨蛞蝓胶囊是否具有致畸和致突变的毒理作用。方法本研究采用大鼠致畸胎、艾姆斯（Ames）试验、小鼠骨髓细胞微核试验、体外细胞染色体畸变试验检测的蛞蝓胶囊致畸胎、致突变性。结果蛞蝓胶囊各剂量对孕鼠体重、胚胎早期发育、胚胎生长发育、以及胎鼠的骨骼发育和内脏器官发育等均无不良影响。无论加与不加S9,各剂量组诱变TA98、TA100种菌落数均未超过自然回变菌落数,与阴性对照组比较均无显著性差异（P〉0．05）。与对照组相比,各剂量组对小鼠的微核率无明显的影响（P〉0．05）。蛞蝓胶囊对培养的哺乳动物体细胞染色体结构无致畸变作用。结论蛞蝓胶囊各剂量均无致畸和致突变的作用,说明在临床应用剂量范围内是安全的。     

Genotoxic evaluation of Halfenprox using the human peripheral lymphocyte micronucleus assay and the Ames test

The genotoxicity and mutagenicity of Halfenprox, a synthetic pyrethroid insecticide and acaricide, was assessed using two standard genotoxicity assays of the Salmonella typhimurium mutagenicity assay (Ames test) and in vitro micronucleus (MN) assay in human peripheral lymphocytes. In the Ames test, Salmonella strains TA98 and TA100 were treated with or without S9 fraction. The doses of Halfenprox were 6.25, 12.5, 25, 50, and 100?μg/plate and test materials were dissolved in DMSO. The concentrations of Halfenprox did not show mutagenic activity on both strains with and without S9 fraction. The MN assay was used to investigate the genotoxic effects of Halfenprox in human peripheral lymphocytes treated with 250, 500, 750, and 1000?μg/ml concentrations of Halfenprox for 24 and 48?h, and at 1000?μg/ml the concentration was significantly increased and the MN formation was compared with the negative control for both treatment periods. In addition, a significant decrease of the nuclear devision index (NDI) values at the higher concentrations of Halfenprox and at both treatment periods was observed.     

Evaluation of the genotoxic potential and acute oral toxicity of standardized extract of Andrographis paniculata (KalmCold™)

Andrographis paniculata is used in the traditional medicine for cold and influenza remedy. The main endeavor in this study was to assess the genotoxicity of the standardized extract of A. paniculata (KalmCold™) through three different in vitro tests: Ames, chromosome aberration (CA), and micronucleus (MN). Ames test was performed at 5000 μg/ml, 1581 μg/ml, 500 μg/ml, 158 μg/ml, 50 μg/ml, 16 μg/ml, while the clastogenicity tests were performed at 80 μg/ml, 26.6 μg/ml, 8.8 μg/ml for short-term treatment without S9; 345 μg/ml, 115 μg/ml, 38.3 μg/ml for short-term treatment with S9; and 46 μg/ml, 15.3 μg/ml, 5.1 μg/ml for long-term without S9 using DMSO as a vehicle control. Results of Ames test confirmed that KalmCold™ did not induce mutations both in the presence and absence of S9 in Salmonella typhimurium mutant strains TA98 and TAMix. In CA and MN, KalmCold™ did not induce clastogenicity in CHO-K1 cells in vitro. Based on our results, it is evident that KalmCold™ is genotoxically safe.     

o-Aminoazotoluene does induce the enzymes of its own mutagenic activation in mouse liver

The objective of this study was to investigate the CYP1A1 and CYP1A2 mRNAs and enzyme activities in mouse liver during induction with o-aminoazotoluene (OAT) as well as the capability of the hepatic S9-fraction from OAT-treated mice to induce its own activation to mutagens in the Ames test using S. typhymurium strain TA98. The data obtained indicate that when used at appropriate doses, OAT is a PAH-type inducer of mouse hepatic microsomal monooxygenases, which activity is not less than that of the known inducer 3,4-benzo[alpha]pyrene. In the absence of S9-fraction enzymes no OAT-mediated mutagenicity was observed in the Ames test. In the presence of the S9-fraction from OAT-pretreated mice, OAT induced as high revertant numbers, as it did in the presence of the S9 fraction from the liver of Aroclor 1254-treated mice. Thus, OAT does induce the enzymes of its own mutagenic activation in mouse liver.