title: Fluorescence quenching properties and chemiluminescence responses of alpha-ketothiols derivatized with o-phthalaldehyde and primary amino compounds.
abstract: Fluorescence and chemiluminescence responses of thiols derivatized with o-phthalaldehyde (OPA) and primary amines have been studied. Among thiols tested, those having an alpha-ketothiol structure [R-CH(SH)-CO-] were found to give extremely low fluorescence intensities when derivatized with OPA and L-valine. The relative fluorescence intensities of alpha-ketothiols, such as mercaptoacetic acid (MAA), 2-mercaptopropionic acid (2-MPA), mercaptosuccinic acid (MSA) and N-(2-mercaptopropionyl)glycine (MPG), were only below 3% of that of N-acetyl-L-cysteine (NAC), having no alpha-ketothiol structure. The addition of detergents such as sodium dodecylsulphate had no effect on the fluorescence and an aprotic solvent (dimethylsulphoxide) showed little effect, whereas these thiols chemiluminesced intensely when derivatized with OPA and N-(4-aminobutyl)-N-ethylisoluminol as a primary amine and successively detected by high-performance liquid chromatography (HPLC) with postcolumn chemiluminescence reactions using hydrogen peroxide and haematin. The relative chemiluminescence intensities of MSA, 2-MPA and MPG to NAC were 41.0%, 27.8% and 117%, respectively. MPG was estimated to be detectable at a concentration level of at least 10 fmol per injection. In conclusion, the weak fluorescence of the alpha-ketothiols derivatized with OPA and primary amines is probably due to the low fluorescence quantum yields of the derivatives, and HPLC with a chemiluminescence detection system is useful for the sensitive detection of thiols.
authors.affiliation: Department of Analytical Chemistry, Faculty of Pharmaceutical Sciences, Science University of Tokyo, 12 Ichigaya-funagawara-machi, Shinjuku-ku, Tokyo 162-0826, Japan.
authors.forename: A
authors.initials: A
authors.lastname: Sano
authors.forename: H
authors.initials: H
authors.lastname: Nakamura
mesh_terms: D000588:Amines; D002851:Chromatography, High Pressure Liquid; D005453:Fluorescence; D008163:Luminescent Measurements; D013050:Spectrometry, Fluorescence; D013438:Sulfhydryl Compounds / Q000032:analysis* / Q000737:chemistry*; D009764:o-Phthalaldehyde
chemical_list: D000588:Amines; D013438:Sulfhydryl Compounds; D009764:o-Phthalaldehyde