Human serum albumin (HSA), the most prominent protein in plasma, is best known for its exceptional ligand (i.e., drug) binding capacity. Here, values of the dissociation equilibrium constant (Kd) for the binding of HIV protease and reverse transcriptase inhibitors to HSA are reported. The binding of abacavir, atazanavir, didanosine, efavirenz, emtricitabine, lamivudine, nelfinavir, nevirapine, ritonavir, saquinavir, stavudine, zalcitabine, and zidovudine to the Sudlow site I (i.e., the warfarin cleft) located in the subdomain IIA involves the alteration of the HSA structure around Trp214 and induces intrinsic tryptophan fluorescence quenching. Accordingly, ibuprofen that primarily binds to the Sudlow site II located in the subdomain IIIA does not affect the HSA intrinsic tryptophan fluorescence and the binding of anti‐HIV drugs to the Sudlow site I. Accounting for the physiological concentration of HSA (= 7.0 × 10‐4 M), the average anti‐HIV drug concentration in plasma (= 1.0 × 10‐4 M), and Kd values for the binding of anti‐HIV drugs to HSA (ranging between 4.4 × 10‐5 M and 3.8 × 10‐4 M), it appears that the fraction of HIV protease and reverse transcriptase inhibitors bound to HSA ranges between 63% and 91%. This represents a significant drawback in the anti‐HIV therapy and management, the anti‐HIV drug concentration required to achieve 90% protease and reverse transcriptase inhibition in the presence of plasma proteins appears to be at least one order of magnitude higher than that required in their absence. IUBMB Life, 56: 609‐614, 2004