~ Pergamon 0146-6380(93)E0029-L Org. Geochem. Vol. 21, No. 6/7, pp. 603-609, 1994 Copyright © 1994 Elsevier Science Ltd Printed in Great Britain. All rights reserved 0146-6380/94 $7.00 + 0.00 Stable carbon isotope compositions of individual amino acid enantiomers in mollusc shell by GC/C/IRMS J. A. SILFER, ! Y. QIAN, 1 S. A. MACKO2 and M. H. ENGEL 1. ISchool of Geology and Geophysics, Energy Center, 100 E Boyd St, The University of Oklahoma, Norman, OK 73019, U.S.A. and 2Department of Environmental Sciences, The University of Virginia, Charlottesville, VA 22903, U.S.A. Abstract--Gas chromatography/combustion/isotope ratio mass spectrometry was used to determine the stable carbon isotope composition of D- and L-enantiomers of amino acids in a fossil Mercenaria shell and in modern samples of Helix pomatia shell that were heated (80°C) for up to 289 days. The similarity in c~ ~3C values for the D- and L-enantiomers of neutral amino acids in Mercenaria implies that these components are indigenous. Whereas differences in the t3C contents of the D- and L-enantiomers of aspartic and glutamic acid in Mercenaria may indicate inputs from exogenous sources, similar discrepancies exist for the acidic amino acids in the heated samples of H. pomatia, suggesting more complicated diagenetic pathways and concomitant isotope effects for the acidic amino acid constituents of shell protein. Key words--enantiomers, amino acids, carbon isotope ratios of amino acids, shell protein, Mercenaria shells, Helix pomatia shells, origin and diagenesis of amino acids INTRODUCTION The stable carbon isotope compositions of individual amino acids reflect the isotope ratios of their precur- sors and the chemical processes or metabolic path- ways responsible for their genesis (e.g. Abelson and Hoering, 1961; Winters, 1971; Macko et al., 1987; Hare et al., 1991). The characteristic molecular signa- tures that arise from source and synthesis variations can be employed to probe the origin and subsequent diagenetic history of amino acids in geological and biological systems (e.g. Hare and Estep, 1983; Serban et al., 1988; Macko and Engel, 1991; Silfer et al., 1992; Qian et al., 1992a). Until recently, the application of compound specific ~ 13C signatures in the earth and biological sciences has been hindered by analytical obstacles. Stable isotope analysis of individual amino acids from complex samples by conventional isotope ratio mass spectrometry requires quantitative isolation by liquid chromatography (LC) and recovery of micro- molar quantities of each pure compound followed by combustion and purification of the resulting carbon dioxide for analysis of individual 13C/12C values (e.g. Hare and Estep, 1983; Macko et al., 1987; Serban et al., 1988). Furthermore, the difficulties encountered in the resolution of a pure D,L-amino acid successfully isolated by HPLC into constituent stereoisomers for carbon and nitrogen isotope analysis are, with respect to geologic materials, substantial (e.g. Serban et al., 1988). The advent of combined gas chromatography/ combustion/isotope ratio mass spectrometry (GC/ *To whom all correspondence should be addressed. C/IRMS) systems has recently stimulated great inter- est in molecular isotope studies of geologic (e.g. Freeman et al., 1990: Hayes et al., 1990; Kennicutt and Brooks, 1990; Rieley et al., 1991; Kohnen et al., 1992; Wakeham et al., 1993) and biologic (Goodman and Brenna, 1992; Yarasheski et al., 1992) systems. The 3 13C analysis of amino acid stereoisomers, in particular, has benefited by this development. The GC/C/IRMS approach eliminates the numerous pre- parative steps needed for conventional carbon iso- tope analysis and provides access to samples with low abundances. However, amino acids are nonvolatile, muitifunctional molecules that require derivatization prior to GC/C/IRMS analysis. The derivatization process introduces additional carbon to the mol- ecules, resulting in modification of the original amino acid isotope compositions. A slightly modified method (after Silfer et al., 1991) is presented here for the determination of amino acid 6 J3C values from GC/C/IRMS analyses of N(O,S)- TFA isopropyl esters. The potential of this method is illustrated by a test of the hypothesis that a compari- son of the carbon isotopic compositions of D- and L-enantiomers of individual amino acids in mollusc shells can be used to appraise their authenticity (Engel and Macko, 1986; Macko and Engel, 1991). Amino acids retain their stable carbon isotopic in- tegrity during racemization (Engel and Macko, 1986; Qian, unpublished results). Differences in the carbon isotopic compositions of D- and L-enantiomers of individual amino acids may, therefore, indicate input from exogenous sources. In this study, samples of modern snail shell were heated to assess whether amino acid constituents of the shell protein retained their carbon isotopic integrity during simulated o~ 2~-~/7--D 603