TY - GEN
T1 - Spectroscopic characterization of the binding mechanism of fluorescein and carboxyfluorescein in human serum albumin
AU - Sulaiman, Saba A.J.
AU - Kulathunga, H. Udani
AU - Abou-Zied, Osama K.
N1 - Publisher Copyright:
© 2015 SPIE.
PY - 2015
Y1 - 2015
N2 - Fluorescein (FL) and some of its precursors have proven to be effective fluorescent tracers in pharmaceutical and medical applications owing to their high quantum yield of fluorescence in physiological conditions and their high membrane permeability. In order to protect FL from metabolic effects during the process of its delivery, human serum albumin (HSA) has been used as a carrier because of its compatibility with the human body. In the present work, we used spectroscopic methods to characterize the binding mechanisms of FL and one of its derivatives, 5(6)- carboxyfluorescein (CFL), in the HSA protein. The absorbance change of the two ligands (FL and CFL) was quantified as a function of the HSA concentration and the results indicate a moderate binding strength for the two ligands inside HSA (1.00 ± 0.12 × 104 M-1). The quenching effect of FL(CFL) on the fluorescence intensity of W214 (the sole tryptophan in HSA) indicates that FL and CFL occupy Site I in the protein which is known to bind several hydrophobic drugs. By performing site-competitive experiments, the location of the ligands is determined to be similar to that of the anticoagulant drug warfarin. At higher ratios of [ligand]/[HSA], we observed an upward curvature in the Stern-Volmer plots which indicates that the ligands occupy more pockets in Site I, close to W214. Our results indicate that both ligands bind in HSA with a moderate strength that should not affect their release when used as fluorescent reporters. The chemical and physical identities of the two ligands are also preserved inside the HSA binding sites.
AB - Fluorescein (FL) and some of its precursors have proven to be effective fluorescent tracers in pharmaceutical and medical applications owing to their high quantum yield of fluorescence in physiological conditions and their high membrane permeability. In order to protect FL from metabolic effects during the process of its delivery, human serum albumin (HSA) has been used as a carrier because of its compatibility with the human body. In the present work, we used spectroscopic methods to characterize the binding mechanisms of FL and one of its derivatives, 5(6)- carboxyfluorescein (CFL), in the HSA protein. The absorbance change of the two ligands (FL and CFL) was quantified as a function of the HSA concentration and the results indicate a moderate binding strength for the two ligands inside HSA (1.00 ± 0.12 × 104 M-1). The quenching effect of FL(CFL) on the fluorescence intensity of W214 (the sole tryptophan in HSA) indicates that FL and CFL occupy Site I in the protein which is known to bind several hydrophobic drugs. By performing site-competitive experiments, the location of the ligands is determined to be similar to that of the anticoagulant drug warfarin. At higher ratios of [ligand]/[HSA], we observed an upward curvature in the Stern-Volmer plots which indicates that the ligands occupy more pockets in Site I, close to W214. Our results indicate that both ligands bind in HSA with a moderate strength that should not affect their release when used as fluorescent reporters. The chemical and physical identities of the two ligands are also preserved inside the HSA binding sites.
KW - Carboxyfluorescein
KW - Drug binding sites
KW - Fluorescein
KW - Fluorescent probes
KW - Fluorescent tracers in medicine
KW - Human serum albumin
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U2 - 10.1117/12.2077520
DO - 10.1117/12.2077520
M3 - Conference contribution
AN - SCOPUS:84931843491
T3 - Progress in Biomedical Optics and Imaging - Proceedings of SPIE
BT - Reporters, Markers, Dyes, Nanoparticles, and Molecular Probes for Biomedical Applications VII
A2 - Raghavachari, Ramesh
A2 - Achilefu, Samuel
PB - SPIE
T2 - Reporters, Markers, Dyes, Nanoparticles, and Molecular Probes for Biomedical Applications VII
Y2 - 8 February 2015 through 10 February 2015
ER -