Reduction of nonpolar amino acids to amino alcohols to enhance volatility for high-precision isotopic analysis

Amino acids are routinely derivatized using carbon-containing groups prior to gas chromatography continuous-now isotope ratio mass spectrometry (GCC-I RMS). Derivative C contaminates analyte C because the entire derivatized co mpound is combusted to CO2, Correction procedures are required to extract t he analyte isotope ratio. We present a method for reduction of six nonpolar amino acids to their corresponding amino alcohols, demonstrate a GC strate gy to produce acceptable peak shapes from the resulting strongly H-bonding analytes, and present isotopic analysis for amino acids and their correspon ding amino alcohols to evaluate any possible isotopic fractionation, Alanin e, valine, leucine, isoleucine, methionine, or phenylalanine was reduced us ing NaBH4 in THF with I-2 as an electrophile, Reactions were performed with 2 g of analyte to permit isotopic analysis by conventional elemental analy sis-IRMS, All reactions were quantitative as assessed by IR spectra, meltin g points, and GC, Recovery from the reaction mixture was 60-84%, GC separat ion of a mixture of the six amino alcohols was achieved using a thick stati onary-phase (5 pm) capillary column to avoid tailing due to hydrogen bondin g to the walls of the fused-silica capillary, The reproducibility of GCC-IR MS determinations of amino alcohols averaged SD(delta C-13) = 0.25 +/- 0.9 parts per thousand. The absolute differences between delta C-13 Of amino ac ids measured by an elemental analyzer coupled to IRMS and amino alcohols me asured by GCC-IRMS was Delta delta C-13 = 0.14 parts per thousand and showe d no general trend. Reactions performed with 2 mg of analyte yielded equiva lent chromatograms. These data indicate that the reduction method does not induce isotopic fractionation and can be used for continuous-flow isotopic analysis to avoid addition of contaminating carbon.