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The biodegradability under aerobic conditions of volatile hydrocarbons (4-6 carbons) contained in gasoline and consisting of n-alkanes, iso-alkanes, cycloalkanes and alkenes, was investigated. Activated sludge was used as the reference microflora. The biodegradation test involved the degradation of the volatile fraction of gasoline in closed flasks under optimal conditions. The kinetics of biodegradation was monitored by CO2 production. Final degradation was determined by gas chromatographic analysis of all measurable hydrocarbons (12 compounds) in the mixture after sampling the headspace of the flasks. The degradation of individual hydrocarbons was also studied with the same methodology. When incubated individually, all hydrocarbons used as carbon sources, except 2,2-dimethylbutane and 2,3-dimethylbutane, were completely consumed in 30 days or less with different velocities and initial lag periods. When incubated together as constituents of the light gasoline fraction, all hydrocarbons were metabolised, often with higher velocities than for individual compounds. Cometabolism was involved in the degradation of dimethyl isoalkanes.

A bacterial strain (strain IFP 2173) was selected from a gasoline-polluted aquifer on the basis of its capacity to use 2,2, 4-trimethylpentane (isooctane) as a sole carbon and energy source. This isolate, the first isolate with this capacity to be characterized, was identified by 16S ribosomal DNA analysis, and 100% sequence identity with a reference strain of Mycobacterium austroafricanum was found. Mycobacterium sp. strain IFP 2173 used an unusually wide spectrum of hydrocarbons as growth substrates, including n-alkanes and multimethyl-substituted isoalkanes with chains ranging from 5 to 16 carbon atoms long, as well as substituted monoaromatic hydrocarbons. It also attacked ethers, such as methyl t-butyl ether. During growth on gasoline, it degraded 86% of the substrate. Our results indicated that strain IFP 2173 was capable of degrading 3-methyl groups, possibly by a carboxylation and deacetylation mechanism. Evidence that it attacked the quaternary carbon atom structure by an as-yet-undefined mechanism during growth on 2,2,4-trimethylpentane and 2,2-dimethylpentane was also obtained.

A methodology was devised and was found useful for the selection of populations degrading recalcitrant hydrocarbons. The work was part of a programme aiming at developing knowledge of the intrinsic capacities of autochtonous microflorae of the environment for gasoline biodegradation. The methodology involved monitoring the progress of degradation in enrichment liquid cultures on the selected hydrocarbon by gas chromatographic analysis of CO2 production and O2 consumption. Populations degrading in particular o-xylene, 1,2,4-trimethylbenzene, cyclohexane were obtained. Concerning 2,2,4-trimethylpentane (isooctane), one microflora (and a pure strain derived from it) growing on this hydrocarbon were obtained from gasoline-polluted water.

A methodology allowing the detailed assessment of the capacities of microflorae to degrade gasoline in aerobic conditions has been developed. It consisted in the determination of the degradation of a gasoline model mixture in liquid cultures in optimal conditions. The gasoline model mixture contained 23 representative hydrocarbons of gasoline (GM23). The kinetics and extent of biodegradation were evaluated by continuous overall monitoring of CO2 production and final chromatographic analysis (usually after about 30 days) of the consumption of each hydrocarbon. The methodology was used with soil and water samples from polluted and non polluted sites. The experimentation aimed at assessing the distribution of the degradative capacities in the environment and the prospects for natural attenuation of gasoline. Nine microflorae were tested. The intrinsic biodegradability (existence of mechanisms of biodegradation) appeared total for GM23 as shown by the results obtained with several microflorae. The degradative capacities of microflorae from non polluted samples were high (total degradation rates at least 85%). Incomplete degradation was observed essentially for trimethylalkanes (2,2,4-trimethylpentane and 2,3,4-trimethylpentane) and for cyclohexane. In several cases, samples from polluted sites exhibited more extensive degradative capacities, with total degradation of all hydrocarbons being observed for three out of the six samples.

The degradation of gasoline by a microflora from an urban waste water activated sludge was investigated in detail. Degradation kinetics were studied in liquid cultures at 30 degrees C by determination of overall O2 consumption and CO2 production and by chromatographic analysis of all 83 identifiable compounds. In a first fast phase (2 d) of biodegradation, 74% of gasoline, involving mostly aromatic hydrocarbons, was consumed. A further 20%, involving other hydrocarbons, was consumed in a second slow phase (23 d). Undegraded compounds (6% of gasoline) were essentially some branched alkanes with a quaternary carbon or/and alkyl chains on consecutive carbons but cycloalkanes, alkenes and C10- and C11-alkylated benzenes were degraded. The degradation kinetics of individual hydrocarbons, determined in separate incubations, followed patterns similar to those observed in cultures on gasoline. Carbon balance experiments of gasoline degradation were performed. The carbon of degraded gasoline was mainly (61.7%) mineralized into CO2, the remaining carbon being essentially converted into biomass.