Mutagenic Activity In Urine Research Articles

Analysis of urinary mutagens produced by cigarette-smoking baboons

Urine concentrates from 17 cigarette-smoking baboons and 12 sham puffers were analyzed for mutagenic activity in S. typhimurium tester strain TA1538. Both the proportion of animals exhibiting measurable mutagenic mutagenic activity in urine and the mean level of mutagenic activity present were significantly greater in cigarette smoking baboons ( P < 0.05). Mutagenic activity in the urine of male and female cigarette-smoking baboons was not significantly different. Age and smoking history did not, but mean blood carboxyhemoglobin did, correlate with mutagenic activity of the urine concentrate from individual animals. Fractionation of the urine concentrates on silicic acid separated the concentrate into fractions that were more active in TA100 and others that were more active in TA1538. Further fractionation was accomplished by HPLC.

Occupational handling of cytostatic drugs.

The bacterial fluctuation test and measurement of the frequency of sister chromatid exchanges were used for evaluation of the exposure of different groups of hospital personnel to cytostatic drugs. Increased mutagenic activity in the urine was detected only in personnel working with inadequate safety precautions, e.g., lack of a ventilated safety cabin for preparation of parenteral solutions. Although such a safety cabin was used within the hospital pharmacy, increased mutagenic activity was detected in the urine of prescriptionists preparing parenteral cytostatic drugs. After a change of glove material and improvement of ventilation in the safety cabin, no work-related increase in urinary mutagenic activity was seen. None of the different groups tested, showed any increase in the frequency of sister chromatid exchanges. It is therefore concluded that handling of cytostatic drugs according to the issued safety recommendations including working in a well ventilated safety cabin, will not result in any enhancement of mutagenic activity in the urine related to work.

A simple modification of the Salmonella liquid-incubation assay Increased sensitivity for detecting mutagens in human urine

A simple modification of the Salmonella/microsome liquid-incubation procedure improves the sensitivity of the assay for detecting mutagens in human urine. Extracts from cigarette smokers' urine were used as a model complex mutagenic mixture for validation of the assay. The modification consists of adding increased numbers of bacterial cells (approximately 10 9) in a concentrated suspension to liver homogenate mix and urine extract, all in 0.2-ml volume. After 90 min incubation at 37°C, the mixture is processed according to the standard Ames test protocol. This procedure is 20 times more sensitive than the standard plate-incorporation test and 13 times more sensitive than a previously reported liquid-incubation protocol. The number of spontaneous revertants did not increase under these conditions and, compared to the plate-incorporation test, 10-fold less liver homogenate and 5-fold less enzymatic cofactors were needed per plate. The procedure was approximately 14 times more sensitive in detecting the mutagenic activity of benzo[ a]pyrene. We also used the modification to determine mutagenic activity in urine from a group of non-smokers. The method may be generally useful for investigations of mutagenic activity in human urine samples.

Urinary mercapturic acid in chemical workers and in control subjects.

Mercapturic acid urinary excretion was followed in a control group and in chemical workers exposed to toxic and mutagenic compounds. Chemical workers had a significantly lower excretion of mercapturic acid when compared with controls. Smoking effects were not significant statistically. No correlation was found between urinary mutagenic activity and mercapturic acid urinary excretion.

Detection of occupational exposure to genotoxic agents with a urinary mutagen assay

The levels of mutagenic activity in the urine of 198 smoking and non-smoking chemical workers, coke plant workers and controls were measured using a urinary mutagen assay based on the Ames Salmonella/microsome test. Urine samples were concentrated using the non-polar resin XAD-2 eluted with methylene chloride and acetone. Activity towards Salmonella strains TA1538 and TA100 with and without microsomal preparation S9 was measured. Urinary mutagenic activity was calculated from linear regression models fitted to the data from the methylene chloride and acetone extracts. Both the non-smoking chemical workers and non-smoking coke plant workers had significantly higher mutagenic activity on strain TA1538 with S9 than did a group of office and laboratory worker controls. However, urinary mutagenic activity from non-smoking chemical and coke plant workers on both strains TA1538 and TA100 without S9, and on strain TA100 with S9, was not significantly different from the mutagenic activity of this control group. Regardless of occupational exposure, smokers had considerably higher urinary mutagenic activity than non-smokers on strain TA1538 with S9, and to a lesser extent on strain TA100 with S9. Estimated tar exposure, calculated from reported cigarette consumption rate and reported cigarette brand, was closely correlated with urinary mutagenic activity on strain TA1538 with S9 ( r = 0.73, P < 0.001 in the Spearman rank order correlation test). d-Glucaric acid concentrations were determined in 150 of the urine samples as a measure of mixed-function oxydase activity. No significant differences were found between the exposed and control groups or between smokers and non-smokers.

Mutagenicity studies with nitrofurans: II. Mutagenicity of nitrofurylacrylic acid for Salmonella typhimurium

The mutagenicity of 3-(5-nitro-2-furyl)acrylic acid (5-NFA) for Salmonella typhimurium tester strains TA100, TA98, TA1535 and TA1950 was studied by using the spot test, liquid-incubation test, microsomal assay, host-mediated assay and mutagenic analysis of blood and urine from treated mice. 5-NFA was dissolved in physiological saline or 10% ethanol. In the spot test, mutagenicity was found with TA100 and TA98 tester strains. A dose-related increase in mutation frequency was observed in the liquid-incubation test (at concentrations ranging from 1 to 50 μg 5-NFA/ml) with TA100 and TA1535; TA100 was the more sensitive. The use of ethanol reduced bacterial cell survival and increased the frequency of revertants. The Salmonella/microsomal assay showed no difference in mutagenic activity between the 5-NFA samples in tests with and without metabolic activation in vitro. In the host-mediated assay, the mutagenicity of 5-NFA was detected after its application in 3 fractionated doses. Subcutaneously applied 5-NFA was mutagenic after a total dose of 15 mg/kg, and after intragastric application of a total dose of 150 mg/kg. When blood samples from treated mice were analysed, the mutagenicity of 5-NFA at intragastric doses of 75, 150 and 300 mg/kg was observed for the tester strains TA100, TA98 and TA1950. The highest frequency of revertants was detected 15 or 30 min after administration of 5-NFA, the control level being reached, according to the dose applied and strain used, after 1–2 h and confirmed in the course of 48 h. At the dose of 10 mg/kg, no mutagenic change was observed. Both tests on blood samples and host-mediated assay yielded identical results regardless of whether 5-NFA was dissolved in physiological saline or in 10% ethanol. Experiments in which mutagenic activity in urine was analysed demonstrated the mutagenicity of 5-NFA at the dose of 150 mg/kg with maxima after 24 and 48 h.

Detection of mutagenic activity in human urine using mutant strains of Salmonella typhimurium.

Histidine-requiring mutants of Salmonella typhimurium that can be reverted to prototrophy by a variety of mutagens were used mutagenic activity in the urine of patients receiving chemotherapeutic agents. Patients given cyclophosphamide and BCNU had detectable urinary mutagenic activity over a 24-hour period, with maximal levels occurring 12 to 21 hours after drug injection. Whereas native cyclophosphamide required the presence of a rat liver extract to be mutagenic in the test system, the cyclophosphamide metabolites in the urine were fully active in the absence of added liver extract. Mutagenic activity was detected in only the first voided urine specimen of patients receiving fluorouracil. Patients receiving Adriamycin, methotrexate, Mitomycin C, and low doses or oral melphalan did not have detectable mutagenic activity in their urines. One thousand and ten random urine speciments were screened for mutagenic activity. Only eight had greater than 26 revertant colonies per plate. Four of the eight had received metronidazole (Flagyl) for vaginitis while two others had received chemotherapeutic drugs. We were unable to detect increased mutagenic metabolites in the urine of 43 patients with known malignancies, using the standard assay conditions.

Mutagenic activity in urine concentrates from rats orally dosed with a chemical which itself is inactive in the Ames test with microsomal activation

The published results on 60 chemicals and X-rays investigated in the mouse spot test were compared with data on the same chemicals tested in the bacterial mutation assay (Ames test) and lifetime rodent bioassays. The performance of the spot test as an in vivo complementary assay to the in vitro bacterial mutagenesis test reveals that of 60 agents, 38 were positive in both systems, 6 were positive only in the spot test, 10 were positive only in the bacterial test and 6 were negative in both assays.The spot test was also considered as a predictor of carcinogenesis; 45 chemicals were carcinogenic of which 35 were detected as positive by the spot test and 3 out of 6 non-carcinogens were correctly identified as negative.If the results are regarded in sequence, i.e. that a positive result in a bacterial mutagenicity test reveals potential that may or may not be realized in vivo, then 48 chemicals were mutagenic in the bacterial mutation assay of which 38 were active in the spot test and 31 were confirmed as carcinogens in bioassays. 12 chemicals were non-mutagenic to bacteria of which 6 gave positive responses in the spot test and 5 were confirmed as carcinogens.These results provide strong evidence that the mouse coat spot test is an effective complementary test to the bacterial mutagenesis assay for the detection of genotoxic chemicals and as a confirmatory test for the identification of carcinogens. The main deficiency at present is the paucity of data from the testing of non-carcinogens.With further development and improvement of the test it is probable that the predictive performance of the assay in identifying carcinogens should improve, since many of the false negative responses may be due to inadequate testing.