The influences of poorly water-soluble anthracene on ester-linked phospholipid fatty acid (PLFA) and glycolipid fatty acid (GLFA) profiles of Mycobacterium sp. LB501T were studied. Bacteria were cultivated on either anthracene or glucose (one culture with successively amended small doses of this substrate and one with excess concentrations) to distinguish between influences of the chemical structure and the bioavailability of the growth substrate. Results revealed that GLFA and PLFA profiles of M . sp. LB501T depended on the availability and the structure of the carbon source. Fatty acid profiles obtained with anthracene differed from those obtained with excess glucose. They were interpreted as a specific adaptation to this poorly bioavailable polycyclic aromatic hydrocarbon (PAH). In contrast, profiles obtained with low glucose concentrations showed clear signs of starvation stress. Stable carbon isotopic ratios (delta(13) C) of GLFA and PLFA of M. sp. LB501T were analysed to characterize the (13) C-fractionation during the biosynthesis of individual fatty acids and to evaluate their value as markers for substrate usage. Although the delta(13) C values of PLFA and GLFA showed differential isotope fractionation during anthracene- and glucose-degradation, they were sufficiently distinct to be used as signatures of bacterial substrate usage.