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 hydrocarbons into membrane phospholipids (and their constituent individual fatty acids and polar headgroup) of putatively bioremediative bacteria. Bacteria are grown on minimal media containing a specified carbon source (in this case, C₁₆ and C₁₈ alkanes), either natural abundance (99% ¹²C) or enriched (99% ¹³C). After extraction (but no other prior separation) of the membrane lipids, electrospray ionization yields a negative-ion FT-ICR mass spectrum containing prominent phospholipid parent ions. If ¹³C-enriched hydrocarbon incorporation is complete, then the mass of the parent ion will increase by n Da, in which n is the number of its constituent carbon atoms; moreover, the ¹³C isotopic distribution pattern will be reversed. The identities of the constituent fatty acids and polar headgroup are obtained by collisional dissociation (MS/MS), and their extent of ¹³C incorporation determined individually. The method is demonstrated for Rhodecoccus rhodochrous (ATCC 53968), for which all 44 carbons of a representative phosphatidylinositol are shown to derive from the hydrocarbon source. Interestingly, although only C₁₆ and C₁₈ alkanes are provided in the growth medium, the bacteria synthesize uniformly enriched C16:0 and C19:0 fatty acids.