A new BODIPY-based long-wavelength fluorescent probe for chromatographic analysis of low-molecular-weight thiols
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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 C18 column within 25 min, with the limits of detection (signal-to-noise ratio = 3) from 0.24 nmol L−1 for glutathione to 0.72 nmol L−1 for penicillamine. The proposed method was validated in part by measuring thiols in blood samples from mice, with recoveries of 95.3–104.3 %.
Keywords1,7-Dimethyl-3,5-distyryl-8-phenyl-(4′-iodoacetamido)difluoroboradiaza-s-indacene Thiols long-wavelength Fluorescence detection High-performance liquid chromatography
This work was supported by the National Natural Science Foundation of China (No. 20835004, 31170344 and 21105074, Beijing, China) and the Research Projects of General Administration of Quality Supervision, Inspection and Quarantine of China (No. 2013IK159).
- 10.Conlan XA, Stupka N, McDermott GP, Francis PS, Barnett NW (2010) Biomed Chromatogr 24:455–457Google Scholar
- 16.Miller J (2008) In: Resch-Genger U (ed) Standards, standardization and quality assurance in fluorescence measurements, 1st edn. Springer, BerlinGoogle Scholar
- 25.Shindy HA (2012) Basics. Org Chem 9:352–360Google Scholar
- 35.David Cole R, Stein William H, Moore S (1958) J Biol Chem 233:1359–1363Google Scholar
- 39.Iciek M, Chwatko G, Lorenc-Koci E, Bald E, Włodek L (2004) Acta Biochim Pol 51:815–824Google Scholar