Simultaneous analysis of plasma thiols by high-performance liquid chromatography with fluorescence detection using a new probe, 1,3,5,7-tetramethyl-8-phenyl-(4-iodoacetamido)difluoroboradiaza-s-indacene.
The design, synthesis and properties of a new derivatizing reagent, 1,3,5,7-tetramethyl-8-phenyl-(4-iodoacetamido)difluoroboradiaza-s-indacene (TMPAB-I), for thiol groups are presented. Using the derivatization of TMPAB-I with thiols, a new high-performance liquid chromatographic method for measuring low-molecular-weight thiol-containing compounds, including coenzyme A (CoA), glutathione, N-acetylcysteine, cysteine, homocysteine (HCys) and 6-mercaptopurine has been developed. The reaction of TMPAB-I with thiols is specific, fast and stable for both TMPAB-I and the derivatives. A baseline separation of all the six derivatives is achieved by isocratic elution on reversed-phase column within 20 min with detection wavelengths of 500 and 510 nm for the excitation and emission, respectively, and the limits of detection (signal-to-noise ratio=3) are from 1.8 fmol (CoA) to 14.0 fmol (HCys), respectively, per 20 microL injection. The utility of the proposed method has been validated by measuring thiol-containing compounds in human plasma samples from healthy persons and patients with hypertension, with recoveries of 94.2-106.8%.
Guo
Xiao-Feng
XF
Department of Chemistry, Wuhan University, Wuhan 430072, China.
Wang
Hong
H
Guo
Yue-Hong
YH
Zhang
Zi-Xing
ZX
Zhang
Hua-Shan
HS
D000081:Acetamides / Q000737:chemistry*; D002851:Chromatography, High Pressure Liquid / Q000295:instrumentation* / Q000379:methods*; D005453:Fluorescence; D006575:Heterocyclic Compounds, 3-Ring / Q000737:chemistry*; D006801:Humans; D013438:Sulfhydryl Compounds / Q000097:blood*
C541287:1,3,5,7-tetramethyl-8-phenyl-(4-iodoacetamido)difluoroboradiaza-s-indacene; D000081:Acetamides; D006575:Heterocyclic Compounds, 3-Ring; D013438:Sulfhydryl Compounds