Bacterial o-methylation of chloroguaiacols: effect of substrate concentration, cell density, and growth conditions.

O-methylation of chloroguaiacols has been examined in a number of gram-positive and gram-negative bacteria to elucidate the effects of substrate concentration, growth conditions, and cell density. Substrate concentrations between 0.1 and 20.0 mg liter were used, and it was found that (i) yields of the O-methylated products were significantly higher at the lowest concentrations and (ii) rates of O-methylation were not linear functions of concentration. With 3,4,5-trichloroguaiacol, the nature of the metabolites also changed with concentration. During growth with a range of substrates, O-methylation of chloroguaiacols also took place. With vanillate, however, de-O-methylation occurred: the chlorocatechol formed from 4,5,6-trichloroguaiacol was successively O-methylated to 3,4,5-trichloroguaiacol and 3,4,5-trichloroveratrole, whereas that produced from 4,5-dichloroguaiacol was degraded without O-methylation. Effective O-methylation in nonproliferating suspensions occurred at cell densities as low as 10 cells ml, although both the yields and the rates were lower than in more dense cultures. By using disk assays, it was shown that, compared with their precursors, all of the O-methylated metabolites were virtually nontoxic to the strains examined. It is therefore proposed that O-methylation functions as a detoxification mechanism for cells exposed to chloroguaiacols and chlorophenols. In detail, significant differences were observed in the response of gram-positive and gram-negative cell strains to chloroguaiacols. It is concluded that bacterial O-methylation is to be expected in the natural environment subjected to discharge of chloroguaiacols.