Biodegradation of benzo(a)pyrene by Pseudomonas strains, isolated from petroleum refinery effluent: Degradation, inhibition kinetics and metabolic pathway

Benzo(a)pyrene, a five-ring polyaromatic hydrocarbon, originating from coal tar, crude oil, tobacco, grilled foods, car exhaust etc, is highly persistent in the environment. It has been classified as a Group I carcinogen, as on its ingestion in human body, diol epoxide metabolites are generated, which bind to DNA causing mutations and eventual cancer. Among various removal methods, bioremediation is most preferred as it is a sustainable approach resulting in complete mineralization of benzo(a)pyrene. Therefore, in this study, biodegradation of benzo(a)pyrene was performed by two strains of Pseudomonas, i. e WDE11 and WD23, isolated from refinery effluent. Maximum benzo(a)pyrene tolerance was 250 mg/L and 225 mg/L against Pseudomonas sp. WD23 and Pseudomonas sp. WDE11 correspondingly. Degradation rate constants varied between 0.0468 and 0.0513/day at 50 mg/L with half-life values between 13.5 and 14.3 days as per first order kinetics, while for 100 mg/L, the respective values varied between 0.006 and 0.007 L/mg. day and 15.28-16.67 days, as per second order kinetics. The maximum specific growth rate of strains WDE11 and WD23 was 0.3512/day and 0.38/day accordingly, while concentrations over 75 mg/L had an inhibitory effect on growth. Major degradation metabolites were identified as dihydroxy-pyrene, naphthalene-1,2-dicarboxylic acid, salicylic acid, and oxalic acid, indicating benzo(a)pyrene was degraded via pyrene intermediates by salicylate pathway through catechol meta-cleavage. The substantial activity of the catechol 2,3 dioxygenase enzyme was noted during the benzo(a)pyrene metabolism by both strains with minimal catechol 1,2 dioxygenase activity. This study demonstrates the exceptional potential of indigenous Pseudomonas strains in complete metabolism of benzo(a)pyrene.