title: Targeted arginine metabolomics: A rapid, simple UPLC-QToF-MS(E) based approach for assessing the involvement of arginine metabolism in human disease.
abstract: BACKGROUND
  Nitric oxide synthase (NOS) mediated conversion of arginine (ARG) to citrulline (CIT) is a key pathway for nitric oxide synthesis. ARG is also metabolised by alternate pathways to ornithine (ORN), homoarginine (HMA), N(G)-monomethyl-L-arginine (MMA), N(G),N(G)-dimethyl-L-arginine (ADMA) and N(G),N(G)'-dimethyl-L-arginine (SDMA), all of which have the capacity to alter NOS activity. Simultaneous assessment of these analytes, when assessing the impact of arginine metabolism in human disease states, is desirable.
  
  
  METHODS
  Analytes (ARG, ADMA, SDMA, MMA, HMA, CIT and ORN) were isolated from human plasma by solvent extraction, evaporated and reconstituted. Ultra-performance liquid chromatography (UPLC) was performed on a 150mm×2.1mm T3 HSS column using a gradient mobile phase comprising ammonium formate (10mM, pH3.8) in methanol (1% to 63%). Analytes were detected by time-of-flight mass spectrometry (Q-ToF-MS) in positive ion mode with electrospray ionisation (ESI+). Data were collected using MS(E).
  
  
  RESULTS
  Solvent extraction provided high recovery (>95%). UPLC-QToF-MS(E) facilitated the separation and quantification of the 7 analytes in an analysis time of 6min. The approach has high sensitivity; LOQ range from 0.005μM (NMMA) to 0.25μM (ARG and ORN), and good precision; intra- and inter-day %RSD are <6% for all analytes.
  
  
  CONCLUSIONS
  This approach provides the capacity to quantify 7 key compounds involved in ARG metabolism in a small sample volume, with a short total analysis time. These characteristics make this approach ideal for undertaking a comprehensive characterisation of this pathway in large data sets (e.g. population studies).
authors.affiliation: Department of Clinical Pharmacology, School of Medicine, Flinders University, Adelaide, Australia.
authors.forename: Madelé
authors.initials: M
authors.lastname: van Dyk
authors.affiliation: Department of Clinical Pharmacology, School of Medicine, Flinders University, Adelaide, Australia.
authors.forename: Arduino A
authors.initials: AA
authors.lastname: Mangoni
authors.affiliation: Centre for Clinical Epidemiology & Biostatistics, Hunter Medical Research Institute, University of Newcastle, Callaghan, New South Wales 2308, Australia.
authors.forename: Mark
authors.initials: M
authors.lastname: McEvoy
authors.affiliation: Centre for Clinical Epidemiology & Biostatistics, Hunter Medical Research Institute, University of Newcastle, Callaghan, New South Wales 2308, Australia.
authors.forename: John R
authors.initials: JR
authors.lastname: Attia
authors.affiliation: Department of Clinical Pharmacology, School of Medicine, Flinders University, Adelaide, Australia.
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Sorich
authors.affiliation: Department of Clinical Pharmacology, School of Medicine, Flinders University, Adelaide, Australia. Electronic address: andrew.rowland@flinders.edu.au.
authors.forename: Andrew
authors.initials: A
authors.lastname: Rowland
mesh_terms: D000368:Aged; D000369:Aged, 80 and over; D001120:Arginine / Q000097:blood / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D005260:Female; D064368:Healthy Volunteers; D006801:Humans; D008297:Male; D013058:Mass Spectrometry / Q000379:methods*; D055432:Metabolomics / Q000379:methods*; D008875:Middle Aged; D015203:Reproducibility of Results; D013997:Time Factors
chemical_list: D001120:Arginine
keywords: Ageing; Arginine metabolism; Chronic disease; Time-of-flight mass spectrometry; Ultra-performance liquid chromatography