title: Comparison of three pretreatment methods for the analysis of distributions and hydrogen isotopic compositions of leaf wax n-fatty acids in natural samples.
abstract: BACKGROUND
  The hydrogen isotopic compositions (δ(2)H values) of leaf wax n-fatty acids (n-FAs) are widely used as powerful biomarkers in biogeochemical and paleoenvironmental studies. However, the use of a variety of pretreatment methods to isolate n-FAs from natural samples for the measurement of δ(2)H values potentially causes discrepancies in the measured δ(2)H values, thereby confounding their use as a biomarker.
  
  
  METHODS
  We tested three typical pretreatment methods for the analysis of the distributions and δ(2)H values of n-FAs from terrestrial higher plants (Carex siderosticta Hance and Ilex chinensis Sim), surface soil, and lacustrine sediment. Method 1 employed an aminopropyl-bonded solid phase to selectively separate free n-FAs from extractable lipids, while Methods 2 and 3 involved alkali- and acid-catalysed chemical reactions in extractable lipids prior to isolation of the n-FAs.
  
  
  RESULTS
  Methods 2 and 3 yielded higher amounts of C16-C24 n-FAs and of certain long-chain n-FAs than Method 1 due to the release of the corresponding ester-bound n-FAs by saponification and transesterification reactions, respectively. Although additional ester-bound n-FAs were recovered by Methods 2 and 3, there were no significant differences in the δ(2)H results for all homologous n-FAs from the two plants and for mid- and long-chain n-FAs from the soil and sediment among the three pretreatment methods.
  
  
  CONCLUSIONS
  Our study indicates that the three different pretreatment methods might not seriously influence the interpretations of δ(2)H values of sedimentary long-chain n-FAs as a terrestrial paleohydrological proxy; however, the release of ester-bound n-FAs by alkali- and acid-catalysed chemical reactions means that users should be cautious when interpreting and comparing distribution data of n-FAs from natural samples with different pretreatment protocols.
authors.affiliation: School of Human Settlements and Civil Engineering, Xi'an Jiaotong University, Xi'an, 710049, China.|State Key Laboratory of Loess and Quaternary Geology, Institute of Earth Environment, Chinese Academy of Sciences, Xi'an, 710061, China.
authors.forename: Yuan
authors.initials: Y
authors.lastname: Yao
authors.affiliation: School of Human Settlements and Civil Engineering, Xi'an Jiaotong University, Xi'an, 710049, China.|State Key Laboratory of Loess and Quaternary Geology, Institute of Earth Environment, Chinese Academy of Sciences, Xi'an, 710061, China.
authors.forename: Weiguo
authors.initials: W
authors.lastname: Liu
mesh_terms: D031255:Carex Plant / Q000737:chemistry; D003903:Deuterium / Q000032:analysis*; D005227:Fatty Acids / Q000032:analysis* / Q000737:chemistry; D008401:Gas Chromatography-Mass Spectrometry; D030017:Ilex / Q000737:chemistry; D018515:Plant Leaves / Q000737:chemistry*; D012987:Soil / Q000737:chemistry; D014885:Waxes / Q000032:analysis* / Q000737:chemistry
chemical_list: D005227:Fatty Acids; D012987:Soil; D014885:Waxes; D003903:Deuterium