Benzyl alcohol, a preservative commonly added to multidose therapeutic protein formulations, can accelerate aggregation of recombinant human interleukin‐1 receptor antagonist (rhIL‐1ra). To investigate the interactions between benzyl alcohol and rhIL‐1ra, we used nuclear magnetic resonance to observe the effect of benzyl alcohol on the chemical shifts of amide resonances of rhIL‐1ra and to measure hydrogen–deuterium exchange rates of individual rhIL‐1ra residues. Addition of 0.9% benzyl alcohol caused significant chemical shifts of amide resonances for residues 90–97, suggesting that these solvent‐exposed residues participate in the binding of benzyl alcohol. In contrast, little perturbation of exchange rates was observed in the presence of either sucrose or benzyl alcohol. © 2011 Wiley‐Liss, Inc. and the American Pharmacists Association J Pharm Sci 100:4215–4224, 2011