An S-adenosyl-L-methionine-dependent O-methyltransferase capable of methylating 2-hydroxy-3-alkylpyrazine (HP) was purified 7,300-fold to apparent homogeneity with an 8.2% overall recovery from Vitis vinifera L. (cv. Cabernet Sauvignon) through a purification procedure including column chromatography on DEAE-Sepharose FF, Ether-5PW, hydroxyapatite, G2000SW(XL), and DEAE-5PW. The relative molecular mass of the native enzyme estimated on gel permeation chromatography was 85 kDa, and the subunit molecular mass was estimated to be 41 kDa on SDS-polyacrylamide gel electrophoresis. The enzyme also methylates caffeic acid. The Vmax for IBHP and caffeic acid were 0.73 and 175 pkatals/mg, respectively, and the respective Km for IBHP and caffeic acid were 0.30 and 0.032 mm. The optimum pH for IBHP (8.5) was different from that for caffeic acid (7.5).