The dissociation constant and the second-order-rate constant for the slow-tight-binding inhibition of human α-thrombin with r-hirudin are 1.9 ± 0.4 pM and (7.14 ± 0.07) x 10⁷ M^-1 s^-1 at Na⁺ = I = 0.19 M (I = ionic strength) in pH 8.06, 0.05 M barbital buffer, and 0.56 ± 0.07 pM and (1.50 ± 0.06 ) x 10⁸ M^-1 s^-1 at Na⁺ = I = 0.31 M in pH 8.12, 0.02 M Tris buffer at 25.0 ± 0.1 C. These parameters are similar at Na⁺ = 0.03 M and in the identically prepared deuterium oxide buffered at pD 8.6. Hirudin is the most potent thrombin inhibitor and interacts non-covalently at the exosite of the enzyme. The binding efficiency of r-hirudin is contingent upon a Na⁺-dependent conformational adjustment of thrombin to the "fast" form, which is predominant in hemostasis. A non-covalent, slow-binding, active-site inhibitor, α-(2-naphthyl-sulfonyl-glycyl)-DL-p-amidinophenyl-alanylpiperidine, (NAPAP), inhibits thrombin with a dissociation constant of 4.8 ± 0.8 nM and a second-order rate constant (1.1 ± 0.2) x 10⁶ M^-1 s^-1 when Na⁺ = I = 0.19 M at the pH optimum, 8.0 to 8.3, in 0.05 M barbital buffer, and 25.0 ± 0.1 C. The deuterium solvent isotope effect for the association of enzyme and inhibitor is unity, which is consistent with the non-covalent nature of the process. The determination of the buffer-independent effects of sodium and deuterium oxide on the inhibition constants are now in progress.