An investigation of the use of nile red as a long-wavelength fluorescent probe for the study of alpha 1-acid glycoprotein-drug interactions.
Spectrofluorimetry in the long-wavelength region of the electromagnetic spectrum (600-1000 nm) is a fairly recent development in photoluminescence spectroscopy, which has numerous advantages over measurements in the more conventional ultraviolet and visible spectral region. 9-Diethylamino-5H-benzophenoxazine-5-one (Nile Red) is an unchanged, hydrophobic molecule, and long-wavelength fluorescence of which is strongly influenced by the polarity of its environment. When Nile Red was added to solutions of alpha 1-acid glycoprotein (Orosomucoid. OMD), it showed an enhancement in fluorescence intensity and a shift to blue in emission wavelength, suggesting it was binding hydrophobically to a non-polar site on the protein. The association constant (12,261,000 +/- 900,000 M-1) and number of binding sites (0.746 +/- 0.044) were calculated for the probe. Upon addition of both acidic and basic drugs, the Nile Red fluorescence reverted to its unbound form, indicating that OMD probably has one high-affinity, wide and flexible binding area for such drugs. Possible enantiomeric selectivity was shown with ephedrine, and the association constant determined for a racemic mixture of propranolol was found to be comparable to other values obtained with alternative, more conventional techniques.
Brown
M B
MB
Dept. of Chemistry, Loughborough University of Technology, Leics., UK.
Miller
J N
JN
Seare
N J
NJ
D002021:Buffers; D005453:Fluorescence; D005456:Fluorescent Dyes / Q000737:chemistry*; D009961:Orosomucoid / Q000737:chemistry*; D010078:Oxazines / Q000737:chemistry*; D011433:Propranolol / Q000032:analysis; D012542:Scattering, Radiation; D013050:Spectrometry, Fluorescence; D013237:Stereoisomerism
D002021:Buffers; D005456:Fluorescent Dyes; D009961:Orosomucoid; D010078:Oxazines; D011433:Propranolol; C044808:nile red