Monolithic column-based reversed-phase liquid chromatography separation for amino acid assay in microdialysates and cerebral spinal fluid.
The development of a HPLC method using a monolithic C18 column is described using fluorescence detection for the assay of 21 amino acids and related substances with derivatisation using ortho-phthaldialdehyde (OPA) in the presence of 3-mercaptopropionic acid (3-MPA). The method employs a tertiary gradient and has a run time of 24 min. Linearity (r2) for each amino acid was found to be greater than 0.99 up to a 10 microM concentration; reproducibility across all analyses (relative standard deviation (R.S.D.)) was between 0.97 and 6.7% and limit of detection (LOD) between 30 and 300 fmol on column. This method has been applied to the analysis of amino acids in both spinal microdialysis and cerebral spinal fluid samples.
Devall
A J
AJ
Pain Signalling Group, Department of Pharmacology, Division of Neuroscience, The Medical School, University of Birmingham, Edgbaston, Birmingham B15 2TT, UK. ajd069@bham.ac.uk
Blake
R
R
Langman
N
N
Smith
C G S
CG
Richards
D A
DA
Whitehead
K J
KJ
D015097:3-Mercaptopropionic Acid / Q000737:chemistry; D000596:Amino Acids / Q000032:analysis* / Q000134:cerebrospinal fluid / Q000737:chemistry; D000818:Animals; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D015314:Dialysis Solutions / Q000032:analysis* / Q000737:chemistry; D017551:Microdialysis; D015394:Molecular Structure; D051381:Rats; D015203:Reproducibility of Results; D009764:o-Phthalaldehyde / Q000737:chemistry
D000596:Amino Acids; D015314:Dialysis Solutions; D009764:o-Phthalaldehyde; D015097:3-Mercaptopropionic Acid