The preservative effects of different types of polyhydroxyl compounds on the stability of biocatalysts, β-amylase, pyruvate kinase and heat shock proteins, under dehydration stress were investigated. Polyhydroxyl compounds used in the present study were trehalose, sucrose, raffinose, maltose, lactose, glucose, glycerol, and dimethyl sulfoxide. The enzymatic activity of β-amylase was lost following both freeze-drying and air-drying; that of pyruvate kinase was lost only after air-drying. Non-reducing sugars, including trehalose, sucrose, and raffinose had marked preservative effects on the stability of these two biocatalysts. In contrast, glycerol caused the deterioration of their enzymatic activities. To our surprise, however, glycerol effectively retarded the inactivation of heat shock proteins, GroEL and Hsc73, caused by short-term dehydration. These findings are of considerable significance for understanding the interplay between heat shock proteins and the stress-induced accumulation of glycerol in anhydrobiotic organisms. Moreover, the utilization of non-reducing sugars as well as glycerol for storage of food materials and certain specific biocatalysts has been elucidated. Keywords IntroductionDehydration causes drastic changes in the stability of biomolecules such as enzymes and lipids, leading to massive cell damage in food storage and preservation. Water is usually indispensable to the maintenance of the integrity of biological membranes, lipid bilayers and many soluble proteins by the formation of hydrogen bonds. Upon drying, intracellular proteins and membranes compensate for the loss of hydrogen bonds with water by bonding with other molecules (Prestrelski et al., 1993;Dong et al., 1995). In the case of proteins, the hydrogen bonds with water can be replaced by protein-protein interactions that lead to irreversible conformational changes, and especially in enzymes, to a loss of activity (Hanafusa, 1969). The removal of water may cause a transition of membranes from the biologically active fluid phase to the gel phase (Crowe et al., 1992(Crowe et al., , 1997.Anhydrobiotic organisms, including certain nematodes, the brine shrimp embryo Artemia salina, the 'resurrection' we found the biological activities of GroEL and Hsc73 to be markedly preserved even in the presence of glycerol under short-term dehydration stress. Materials and MethodsMaterials β-Amylase purified from soy bean was kindly provided by Dr. Mikami (Kyoto University). Pyruvate kinase and lactate dehydrogenase derived from rabbit muscle were purchased from Wako Pure Chemical Industries. Hsc73 purified from bovine brain was obtained from StressGen Biotechnologies Corp. (Victoria BC, Canada). GroEL was prepared as described by Tani et al. (2004). Dihydrofolate reductase (DHFR; EC 1.5.1.3) was obtained from Sigma Chemical Co. (St. Louis, MO). ADP and NADH were purchased from Oriental Yeast Co., Ltd. (Japan). Sugars, polyhydroxy compounds, 3,5-dinitrosalicylic acid (DNSA), soluble starch, phosphoenolpyruvic acid trisodium salt hydrate, and oth...