Carbon isotopic fractionation in lipids from methanotrophic bacteria II: The effects of physiology and environmental parameters on the biosynthesis and isotopic signatures of biomarkers

Controls on the carbon isotopic signatures of methanotroph biomarkers have been further explored using cultured organisms. Growth under conditions which select far the membrane-bound particulate form of the methane monooxygenase enzyme (pMMO) leads to a significantly higher isotopic fractionation than does growth based on the soluble isozyme in both RuMP and serine pathway methanotrophs; in an RuMP type the Delta delta(13)C(biomass) equaled -23.9 parts per thousand for pMMO and -12.6 parts per thousand for sMMO. The distribution of biomarker lipids does not appear to be significantly affected by the dominance of one or the other MMO type and their isotopic compositions generally track those of the parent biomass. The C-13 fractionation behaviour of serine pathway methanotrophs is very complex, reflecting the assimilation of both methane and carbon dioxide and concomitant dissimilation of methane-derived carbon. A limitation in CH4 availability leads to the production of biomass which is C-13-enriched with respect to both carbon substrates and this occurs irrespective of MMO type. This startling result indicates that there must be an additional fractionation step downstream from the MMO reaction which leads to incorporation of C-13-enriched carbon at the expense of dissimilation of C-13-depleted CO2. In these organisms, polyisoprenoid lipids are C-13-enriched compared to polymethylenic lipid which is the reverse of that found in the RuMP types. Serine cycle hopanoids, for example, can vary anywhere from 12 parts per thousand depleted to 10 parts per thousand enriched with respect to the CH4 substrate depending on its concentration. Decrease in growth temperature caused an overall increase in isotopic fractionation. In the total biomass, this effect tended to be masked by physiological factors associated with the type of organism and variation in the bulk composition. The effect was, however, clearly evident when monitoring the C-13 signature of total lipid and individual biomarkers. Our results demonstrate that extreme carbon isotopic depletion in field samples and fossil biomarker lipids can be indicative of methanotrophy but the converse is not always true. For example, the hopanoids of a serine cycle methanotroph may be isotopically enriched by more than 10 parts per thousand compared to the substrate methane when the latter is limiting. In other words, hopanoids from some methanotrophs such as M. trichosporium would be indistinguishable from those of cyanobacteria or heterotrophic bacteria on the basis of either chemical structure or carbon isotopic signature. Copyright (C) 1999 Elsevier Science Ltd.