Interaction of pirprofen enantiomers with human serum albumin

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Abstract
The interaction of pirprofen enantiomers with human serum albumin (HSA) was investigated by means of high‐performance liquid chromatography (HPLC), circular dichroism (CD), and 1H NMR spectroscopy. HPLC experiments indicated that both pirprofen enantiomers were bound to one class of high‐affinity binding sites (n(+) = 1.91 ± 0.13, K(+) = (4.09 ± 0.64) × 105 M−1, n(‐) = 2.07 ± 0.13, K(‐) = (6.56 ± 1.35) × 105 M−1) together with nonspecific binding (n′K′(+) = (1.51 ± 0.21) × 104 M−1, n′K′(‐) = (0.88 ± 0.13) × 104 M−1). Slight stereoselectivity in specific binding was demonstrated by the difference in product n(+)K(+) = (0.77 ± 0.08) × 106 M−1 vs. n(‐)K(‐) = (1.30 ± 0.21) × 106 M−1, i.e., the ratio n(‐)K(‐)/n(+)K(+) = 1.7. CD measurements showed changes in the binding sites located on the aromatic amino acid side chains (a small positive band at 315 nm and a pronounced negative extrinsic Cotton effect in the region 250–280 nm). The protein remains, however, in its predominantly alpha‐helical conformation. The 1H NMR difference spectra confirmed that both pirprofen enantiomers interacted with HSA specifically, most probably with site II on the albumin molecule.