Retention/quantitation properties of the o-phthaldialdehyde-3-mercaptopropionic acid and the o-phthaldialdehyde-N-acetyl-L-cysteine amino acid derivatives in reversed-phase high-performance liquid chromatography

摘要: The separation/identification of 25 amino acids as their o-phthaldialdehyde-3-mercaptopropionic acid (OPA/MPA) and o-phthaldialdehyde-N-acetyl-L-cysteine (OPA/NAC) derivatives have been optimized [paying particular attention to those which elute with more than one derivative (glycine, histidine, gamma-aminobutyric acid, beta-alanine, ornithine, lysine) that are expected be present in apples free form]. Optimum separation conditions reported on six reversed-phase columns: Nucleosil 3 5 microm, 150(+20 guard)x4.0 mm; Gromsil 150(+10 Hypersil mm 200(+20 mm. Elutions were followed, simultaneously, photodiode array fluorescence detectors connected line. Optimization studies carried out model solutions a function temperature (30-55 degrees C) eluent flow-rate (0.8-2.5 mL/min) demonstrated optimum resolutions obtained the highest applicable (remaining safe side column pressure << 3500 p.s.i.; 1 p.s.i.=6894.76 Pa) range 30-50 C. Twenty-five acids, eluting 31 separate, characteristic derivatives, determined all columns (the main component, asparagine, overwhelming excess, together minor constituents glutamine, homoserine, homoarginine). case both same type (Hypersil, microm), follows: for OPA/MPA programmed [1.3-2.3 ml/min; column, guard)x4 mm], at 50 C, while, OPA/NAC 2.1 ml/min flow rate, 30 C [column, 40 37 min run times, including equilibration. Responses corresponding proved independent used; reproducibility concentration 6-12,000 pmol, related injected amount was <3.4% RSD (average relative standard deviation percentage). utility protocol quantitation content five apple varieties (Jonagored, Idared, Jonica, Florina, Freedom) various harvesting dates after different storage times. Derivatization pulp performed filtered samples, applying any special isolation processes.