title: Effects of acidification, lipid removal and mathematical normalization on carbon and nitrogen stable isotope compositions in beaked whale (Ziphiidae) bone.
abstract: BACKGROUND
  The analysis of stable isotopes in tissues such as teeth and bones has been used to study long-term trophic ecology and habitat use in marine mammals. However, carbon isotope ratios (δ(13) C values) can be altered by the presence of (12) C-rich lipids and carbonates. Lipid extraction and acidification are common treatments used to remove these compounds. The impact of lipids and carbonates on carbon and nitrogen isotope ratios (δ(15) N values), however, varies among tissues and/or species, requiring taxon-specific protocols to be developed.
  
  
  METHODS
  The effects of lipid extraction and acidification and their interaction on carbon and nitrogen isotope values were studied for beaked whale (Ziphiidae) bone samples. δ(13) C and δ(15) N values were determined in quadruplicate samples: control, lipid-extracted, acidified and lipid-extracted followed by acidification. Samples were analyzed by means of elemental analysis isotope ratio mass spectrometry. Furthermore, the efficiency of five mathematical models developed for estimating lipid-normalized δ(13) C values from untreated δ(13) C values was tested.
  
  
  RESULTS
  Significant increases in δ(13) C values were observed after lipid extraction. No significant changes in δ(13) C values were found in acidified samples. An interaction between both treatments was demonstrated for δ(13) C but not for δ(15) N values. No change was observed in δ(15) N values for lipid-extracted and/or acidified samples. Although all tested models presented good predictive power to estimate lipid-free δ(13) C values, linear models performed best.
  
  
  CONCLUSIONS
  Given the observed changes in δ(13) C values after lipid extraction, we recommend a priori lipid extraction or a posteriori lipid normalization, through simple linear models, for beaked whale bones. Furthermore, acidification seems to be an unnecessary step before stable isotope analysis, at least for bone samples of ziphiids. Copyright © 2015 John Wiley & Sons, Ltd.
authors.affiliation: Programa de Pós-Graduação em Oceanografia Biológica, Instituto de Oceanografia, Universidade Federal do Rio Grande, 96203-900, Rio Grande, RS, Brazil.|Laboratório de Ecologia e Conservação da Megafauna Marinha - ECOMEGA, Instituto de Oceanografia, Universidade Federal do Rio Grande, 96203-900, Rio Grande, RS, Brazil.
authors.forename: Ana Carolina C
authors.initials: AC
authors.lastname: Tatsch
authors.affiliation: Programa de Pós-Graduação em Oceanografia Biológica, Instituto de Oceanografia, Universidade Federal do Rio Grande, 96203-900, Rio Grande, RS, Brazil.|Laboratório de Ecologia e Conservação da Megafauna Marinha - ECOMEGA, Instituto de Oceanografia, Universidade Federal do Rio Grande, 96203-900, Rio Grande, RS, Brazil.
authors.forename: Eduardo R
authors.initials: ER
authors.lastname: Secchi
authors.affiliation: Programa de Pós-Graduação em Oceanografia Biológica, Instituto de Oceanografia, Universidade Federal do Rio Grande, 96203-900, Rio Grande, RS, Brazil.|Laboratório de Ecologia e Conservação da Megafauna Marinha - ECOMEGA, Instituto de Oceanografia, Universidade Federal do Rio Grande, 96203-900, Rio Grande, RS, Brazil.
authors.forename: Silvina
authors.initials: S
authors.lastname: Botta
mesh_terms: D053000:Analytic Sample Preparation Methods / Q000379:methods* / Q000592:standards; D000818:Animals; D001842:Bone and Bones / Q000737:chemistry* / Q000378:metabolism; D002247:Carbon Isotopes / Q000032:analysis* / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D008055:Lipids / Q000737:chemistry / Q000302:isolation & purification*; D013058:Mass Spectrometry; D008962:Models, Theoretical; D009587:Nitrogen Isotopes / Q000032:analysis* / Q000378:metabolism; D014907:Whales / Q000378:metabolism*
chemical_list: D002247:Carbon Isotopes; D008055:Lipids; D009587:Nitrogen Isotopes