Efficacy of bacterial bioremediation: Demonstration of complete incorporation of hydrocarbons into membrane phospholipids from Rhodococcus hydrocarbon degrading bacteria by electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry

We present a method and example to establish complete incorporation of hydr ocarbons into membrane phospholipids (and their constituent individual fatt y acids and polar headgroup) of putatively bioremediative bacteria. Bacteri a are grown on minimal media containing a specified carbon source (in this case, C-16 and C-18 alkanes), either natural abundance (99% C-12) or enrich ed (99% C-13). After extraction (but no other prior separation) of the memb rane lipids, electrospray ionization yields a negative-ion FT-ICR mass spec trum containing prominent phospholipid parent ions. If C-13-enriched hydroc arbon incorporation is complete, then the mass of the parent ion will incre ase by n Da, in which n is the number of its constituent carbon atoms; more over, the C-13 isotopic distribution pattern Will be reversed. The identiti es of the constituent fatty acids and polar headgroup are obtained by colli sional dissociation (MS/ MS), and their extent of C-13 incorporation determ ined individually. The method is demonstrated for Rhodococcus rhodochrous ( ATCC# 53968), for which all 44 carbons of a representative phosphatidylinos itol are shown to derive from the hydrocarbon source. Interestingly, althou gh only C-16 and C-18 alkanes are provided in the growth medium, the bacter ia synthesize uniformly enriched C16:0 and C19:0 fatty acids.