Jump to Main Content
A new BODIPY-based long-wavelength fluorescent probe for chromatographic analysis of low-molecular-weight thiols
- Zhang, Li-Yun, Tu, Feng-Qin, Guo, Xiao-Feng, Wang, Hong, Wang, Peng, Zhang, Hua-Shan
- Analytical and bioanalytical chemistry 2014 v.406 no.26 pp. 6723-6733
- acetylcysteine, blood, cysteine, detection limit, fluorescence, glutathione, homocysteine, mice, penicillamine, reversed-phase high performance liquid chromatography, thiols, wavelengths
- A new long-wavelength fluorescent probe, 1,7-dimethyl-3,5-distyryl-8-phenyl-(4′-iodoacetamido)difluoroboradiaza-s-indacene (DMDSPAB-I), was designed and synthesized for thiol labeling in high-performance liquid chromatography (HPLC). The excitation and emission wavelengths of DMDSPAB-I are 620 and 630 nm, respectively, with a high fluorescence quantum yield of 0.557, which is advantageous in preventing interference of intrinsic fluorescence from complex biological matrices and enabling high sensitivity HPLC. Based on DMDSPAB-I, a reversed-phase HPLC method was developed for measuring low-molecular-weight thiols including glutathione, cysteine, homocysteine, N-acetylcysteine, cysteinylglycine, and penicillamine. After the specific reaction of DMDSPAB-I with thiols, baseline separation of all six stable derivatives was achieved through isocratic elution on a C₁₈column within 25 min, with the limits of detection (signal-to-noise ratio = 3) from 0.24 nmol L⁻¹for glutathione to 0.72 nmol L⁻¹for penicillamine. The proposed method was validated in part by measuring thiols in blood samples from mice, with recoveries of 95.3–104.3 %.