Mutagenicity testing of seminal fluid: seminal fluid increases the mutagenicity of the precursor mutagen benzo[a]pyrene in the presence of S9 mix.

Small amounts of seminal fluid strongly enhanced the mutagenicity of the precursor mutagen benzo[a]pyrene (BP) in the Salmonella/microsome test. This previously unreported effect was found only in the presence of S9 mix for metabolic activation. The increase far exceeded the additive effect expected from experiments where seminal fluid and BP were tested separately with S9 mix. Testing of the direct-acting mutagen 4-nitro-o-phenylene-diamine (NPD) together with seminal fluid resulted in a lower mutagenic activity than that of NPD alone. Seminal fluid had a bactericidal effect on the Salmonella bacteria, thus only volumes up to 40 microliter could be used per plate. The mutagenic effect of only seminal fluid and S9 mix was slightly increased over controls in a standard Ames test, but was equal to the spontaneous mutation rate with a preincubation test modified according to Kado and coworkers. There were no significant differences between seminal plasma from smokers and non-smokers in any experimental series. Seminal fluid concentrated 20-fold by extraction with the mutagen-removing adsorbant Mutasorb did not have any enhancing effect on the mutagenicity of BP, nor did it exhibit any mutagenic activity in itself with or without S9 mix.

Mutagenicity testing of amniotic fluid from diabetic women, with special reference to their smoking habits.

Amniotic fluid from 16 diabetic and 78 healthy women at term was tested for capacity to cause mutations in Salmonella typhimurium bacterial tester strain TA98 (Ames test). Diabetes as well as heavy smoking increased the mutagenic activity of amniotic fluid. The difference between groups of diabetics and controls was significant in both nonsmokers and women who had smoked more than 5 cigarettes the last 48 h before delivery. It seems plausible that metabolic disturbances, perhaps enhanced by a lowered oxygenation, in some instances could produce mutagenic compounds. Mutagenic activity in amniotic fluid may be one of the factors underlying the increased incidence of congenital malformations in the offspring of diabetic women. Early mutations could cause such developmental errors in the embryos, and possibly also in future generations by damage to germ cells. Heavy smoking alone also caused an increase in mutagenic activity in term amniotic fluid. Our findings reflected an enhancing effect of smoking in diabetes. A pregnant diabetic woman who smoked would thus further endanger her already jeopardized pregnancy.

Mutagenicity testing of human milk from smokers and non-smokers in the Salmonella/microsome test.

Human milk was tested in the Ames plate incorporation test using strain TA98. Pools from smokers and non-smokers respectively, as well as individual samples from 14 smokers and 15 controls were tested. No difference was found between milk from smokers and non-smokers in the concentrations and volumes used. Preliminary to this a cigarette smoke condensate (CSC) was tested. The CSC was then incubated with human milk which was fractionated, concentrated and the different fractions tested for mutagenic activity. 98% of the mutagenic effect added (measured as number of revertants caused by an equivalent volume of CSC) was recovered 52% of the revertant colonies were recovered from the fat fractions, 40% from the skimmed milk and 8% from a precipitate. Breast milk was also incubated with C14-benzo(a)pyrene and then fractionated by density gradient ultracentrifugation. 97% of the radioactivity was found in the lipid fraction.

Mutagenic effect of amniotic fluid from smoking women at term.

Concentrated term amniotic fluid samples from 44 women smokers and 44 controls were investigated with respect to mutagenic effect in the Salmonella/mammalian-microsome mutagenicity test, using tester strains TA98 and TA100. Tests with freeze-dried specimens of term amniotic fluid showed increases in the number of revertant colonies over background values, regardless of smoking status. However, samples from heavy smokers produced a higher number of revertants than did samples from nonsmokers in several experiments with tester strain TA98. The increase was statistically significant, using either total tar content or number of cigarettes smoked to identify heavy smokers. Experimental series with tester strain TA100 also resulted in higher group means for heavy smokers than for nonsmokers, but the difference was not statistically significant with the concentrations used in this assay. We conclude that heavy smokers may expose their unborn children to mutagenic substances.

Study of the amniotic fluid from smokers and non-smokers in the Ames test.

Amniotic fluid from smokers and non-smokers was tested by the Salmonella/mammalian microsome test. Concentrated amniotic fluid from heavy smokers at term showed an increase in the number of revertants with increasing exposure to tar. However, some of the non-smokers had a higher number of revertants than the smokers. No significant differences were found between second-trimester samples from smokers and non-smokers, but the limited volumes available at this stage of pregnancy may be a source of error.