Nitrile-derivatized amino acids have recently been proposed to serve as infrared (IR) probes of local environment of proteins. In this Letter, we presented results regarding the temperature dependence of the Cequivalent toN stretching vibration of a nitrile-derivatized phenylalanine, Phe(CN), in water. Two-dimensional (2D) infrared correlation analysis revealed that at least three species, corresponding to Phe(CN) Molecules with either free or hydrogen-bonded CN groups, exist. This microheterogeheity was further studied by a global curve-fitting procedure, which yielded: DeltaH = -4.7 kcal mol(-1), AS = -15 cal K-1 mol(-1), and DeltaC(p)= -79 cal K-1 mol(-1), for the transition from the free to hydrogen-bonded species. (C) 2003 Elsevier Science B.V. All rights reserved.