Fluorescence and chemiluminescence detection of oxazole-labelled amines and thiols

Fluorescence and chemiluminescence analyses of amino acids and thiols derivatized with 2‐fluoro‐4,5‐diphenyloxazole (DIFOX) and 2‐chloro‐4,5‐bis(p‐N,N‐dimethylaminosulphonylphenyl)oxazole (SAOX‐CI) were investigated. Thirteen diphenyloxazole (DIOX)‐derivatized amino acids were separated within 38 min by a linear gradient elution from 100% A [0.05 M phosphate (pH 7.0):CH3CN (75:25)] to 100% B [0.05 M phosphate (pH 7.0):CH3CN (1:1)] over 30 min and an isocratic elution of 100% B for 30 min. The detection limits (S/N = 2) with fluorescence detection were in the range of 19–64 fmol. Thiols derivatized with SAOX‐CI were separated by an isocratic elution using 0.1 M H3PO4:CH3CN (65:35) and detected fluorimetrically. The detection limits (S/N = 2) of reduced glutathione, N‐acetylcysteine, 2‐mercaptopropionylglycine, cysteine, homocysteine and captopril were 1.2, 1.5, 1.9, 5.7, 6.4 and 7.9 fmol, respectively. Peroxyoxalate chemiluminescence (CL) intensities of sulphonyl‐5‐N,N‐dimethylaminonaphthalene (DNS), SAOX and DIOX derivatives were compared using three different oxalate esters (DFPO, TCPO and TDPO) by flow injection analysis. The relative chemiluminescence intensity (RCL) of SAOX‐proline and DIOX‐proline were 76–80% and 19–25% of DNS‐proline (100%), respectively. Other SAOX and DIOX derivatives showed lower CL intensities ( < 12%). Extremely low CL intensities were obtained for the fluorescent tagging reagents (