Determination of plasma pipecolic acid by an easy and rapid liquid chromatography-tandem mass spectrometry method.

Abstract

Pipecolic acid (PA) is an important biochemical marker for the diagnosis of peroxisomal disorders. PA is also a factor responsible for hepatic encephalopathy and a possible biomarker for pyridoxine-dependent seizures. We developed an easy and rapid PA quantification method, by high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS), requiring no derivatization and applicable to small sample volumes. Plasma (100 μl) is extracted with 500 μl acetonitrile (ACN) containing 2 μmol/l [(2)H5]-phenylalanine as internal standard, vortexed and centrifuged. The supernatant is analyzed by HPLC-MS/MS in positive-ion mode using multiple reaction monitoring scan type. HPLC column is a Luna HILIC (150×3.0mm; 3 μ 200A): Buffer A: ammonium formate 5 mmol/l; Buffer B: ACN/H20 90:10 containing ammonium formate 5 mmol/l. PA retention time is 4.86 min. Recovery was 93.8%, linearity was assessed between 0.05 and 50 μmol/l (R(2)=0.998), lower limit of detection was 0.010 μmol/l and lower limit of quantification was 0.050 μmol/l. Coefficient of variation was 3.2% intra-assay and 3.4% inter-assay, respectively. Clinical validation was obtained by comparing PA plasma values from 5 patients affected by peroxisomal disorders (mean, 23.38 μmol/l; range, 11.20-37.1 μmol/l) to 24 ages related healthy subjects (mean, 1.711 μmol/l; range, 0.517-3.580 μmol/l).

• Authors: Michela Semeraro, Maurizio Muraca, Giulio Catesini, Rita Inglese, Francesca Iacovone, Gloria Maria Barraco, Melania Manco, Sara Boenzi, Carlo Dionisi-Vici, Cristiano Rizzo
• Journal: Clinica chimica acta; international journal of clinical chemistry (Clin Chim Acta) Vol. 440 Pg. 108-12 (Feb 02 2015) ISSN: 1873-3492 [Electronic] Netherlands
• PMID: 25447702 (Publication Type: Journal Article)
• Copyright: Copyright © 2014 Elsevier B.V. All rights reserved.

Chemical References

• Biomarkers
• Isonipecotic Acids
• Nipecotic Acids
• Pipecolic Acids
• nipecotic acid
• pipecolic acid

Topics

• Adolescent
• Adult
• Biomarkers (blood)
• Calibration
• Child
• Child, Preschool
• Chromatography, High Pressure Liquid (methods)
• Epilepsy (blood, diagnosis)
• Female
• Humans
• Infant
• Infant, Newborn
• Isonipecotic Acids (isolation & purification)
• Limit of Detection
• Male
• Nipecotic Acids (isolation & purification)
• Peroxisomal Disorders (blood, diagnosis)
• Pipecolic Acids (blood, isolation & purification)
• Reference Values
• Reproducibility of Results
• Tandem Mass Spectrometry (methods)
• Young Adult