BACKGROUND: Plasma amino acids are usually analyzed by ion-exchange chromatography (IEC), a reproducible but time consuming method. Here, we test whether plasma amino acids can be analyzed using reverse-phase high performance liquid chromatography (HPLC). METHODS: Filtered plasma, with S-carboxymethyl-l-cysteine as the internal standard, was derivatized and analyzed by an Agilent 1100 HPLC system. Primary amino acids were derivatized with o-phthalaldehyde 3-mercaptopropionic acid (OPA) and detected by a diode array detector. Secondary amino acids were derivatized with 9-fluorenylmethyl chloroformate (FMOC) and detected fluorometrically. Chromatographic separation is achieved by two gradient elutions (two injections per sample), starting at different pHs, on a reverse phase Agilent Zorbax Eclipse C(18) column AAA (4.6 x 150 mm). RESULTS: The HPLC method evaluated correlated well with IEC (0.89</=r</=1.00) with linearity up to 2500 mumol/l. The between- and within-run CVs were <6.0%. In addition, this method is able to separate argininosuccinic acid, homocystine and allo-isoleucine, rare but clinically significant amino acids. CONCLUSION: This HPLC method was comparable to IEC and could represent an alternative for amino acid analysis.
BACKGROUND: Plasma amino acids are usually analyzed by ion-exchange chromatography (IEC), a reproducible but time consuming method. Here, we test whether plasma amino acids can be analyzed using reverse-phase high performance liquid chromatography (HPLC). METHODS: Filtered plasma, with S-carboxymethyl-l-cysteine as the internal standard, was derivatized and analyzed by an Agilent 1100 HPLC system. Primary amino acids were derivatized with o-phthalaldehyde3-mercaptopropionic acid (OPA) and detected by a diode array detector. Secondary amino acids were derivatized with 9-fluorenylmethyl chloroformate (FMOC) and detected fluorometrically. Chromatographic separation is achieved by two gradient elutions (two injections per sample), starting at different pHs, on a reverse phase Agilent Zorbax Eclipse C(18) column AAA (4.6 x 150 mm). RESULTS: The HPLC method evaluated correlated well with IEC (0.89</=r</=1.00) with linearity up to 2500 mumol/l. The between- and within-run CVs were <6.0%. In addition, this method is able to separate argininosuccinic acid, homocystine and allo-isoleucine, rare but clinically significant amino acids. CONCLUSION: This HPLC method was comparable to IEC and could represent an alternative for amino acid analysis.
Authors: Tobias Eisenberg; Mahmoud Abdellatif; Sabrina Schroeder; Uwe Primessnig; Slaven Stekovic; Tobias Pendl; Alexandra Harger; Julia Schipke; Andreas Zimmermann; Albrecht Schmidt; Mingming Tong; Christoph Ruckenstuhl; Christopher Dammbrueck; Angelina S Gross; Viktoria Herbst; Christoph Magnes; Gert Trausinger; Sophie Narath; Andreas Meinitzer; Zehan Hu; Alexander Kirsch; Kathrin Eller; Didac Carmona-Gutierrez; Sabrina Büttner; Federico Pietrocola; Oskar Knittelfelder; Emilie Schrepfer; Patrick Rockenfeller; Corinna Simonini; Alexandros Rahn; Marion Horsch; Kristin Moreth; Johannes Beckers; Helmut Fuchs; Valerie Gailus-Durner; Frauke Neff; Dirk Janik; Birgit Rathkolb; Jan Rozman; Martin Hrabe de Angelis; Tarek Moustafa; Guenter Haemmerle; Manuel Mayr; Peter Willeit; Marion von Frieling-Salewsky; Burkert Pieske; Luca Scorrano; Thomas Pieber; Raimund Pechlaner; Johann Willeit; Stephan J Sigrist; Wolfgang A Linke; Christian Mühlfeld; Junichi Sadoshima; Joern Dengjel; Stefan Kiechl; Guido Kroemer; Simon Sedej; Frank Madeo Journal: Nat Med Date: 2016-11-14 Impact factor: 53.440
Authors: Norbert J Tripolt; Felix Aberer; Regina Riedl; Barbara Hutz; Jasmin Url; Gudrun Dimsity; Andreas Meinitzer; Tatjana Stojakovic; Ronald Hödl; Marianne Brodmann; Franz Hafner; Harald Sourij Journal: Trials Date: 2016-10-13 Impact factor: 2.279
Authors: Andreas Baranyi; Omid Amouzadeh-Ghadikolai; Hans-Bernd Rothenhäusler; Simon Theokas; Christoph Robier; Maria Baranyi; Michael Koppitz; Gerhard Reicht; Peter Hlade; Andreas Meinitzer Journal: PLoS One Date: 2015-11-18 Impact factor: 3.240