Vol. 74, No. 6, 1997 727 ENZYMES Optimization of Temperature, Time, and Lactic Acid Concentration to Inactivate Lipoxygenase and Lipase and Preserve Phytase Activity in Barley (cv. Blenheim) During Soaking A. RUTGERSSON, 1,3 E.-L. BERGMAN, 2 H. LINGNERT, 1 and A.-S. SANDBERG 2 ABSTRACT Cereal Chem. 74(6):727–732 A barley-soaking process was studied to find conditions that inactivate the prooxidative enzyme lipoxygenase and the lipolytic enzyme lipase but preserve phytase activity to develop possible procedures for production of barley products with potentially high mineral bioavailability and good oxidative stability. Lactic acid concentration, temperature, and soaking time were studied. The study was done using a multivariate experimental design. Lactic acid concentration varied between 0 and 1%, temperature varied between 45 and 70°C, and soaking time varied between 30 and 120 min. Although conditions under which lipoxygenase was inactivated were found, total inactivation of lipase was not obtained. Total lipoxy- genase inactivation with <20% remaining lipase activity and >60% remain- ing phytase activity was reached after soaking in 1% lactic acid at suit- able time-temperature combinations of 70–110 min and 53–58°C. Barley has a high starch, dietary fiber, vitamin, and mineral con- tent and, therefore, fits well with present dietary guidelines (Sundberg 1995). Human studies have shown that consuming barley products reduces postprandial glucose, insulin, and serum lipid levels. Due to its higher β-glucan content, barley contains as much total dietary fiber and soluble dietary fiber as oat (Ranhotra et al 1991). However, barley, like all other cereals, contains phytic acid (myo- inositol hexaphosphoric acid), which is an antinutrient that nat- urally occurs in foods and is present in large quantities in cereals (Lolas et al 1976). The antinutritive effect of phytic acid is caused by its ability to form chelates (phytates) with minerals such as zinc (Lönnerdal et al 1988, Sandberg 1991, Rossander et al 1992, Sandström and Sandberg 1992), calcium (Heaney et al 1991), and iron (Hallberg et al 1989, Brune et al 1992, Rossander et al 1992), making them unavailable for absorption in the human intestine. Cereals are staple foods in a large part of the world and are a major source of minerals; therefore, it is important that the min- erals from these products have a high bioavailability. The intake of large amounts of foods rich in phytate may cause mineral deficiency symptoms (Maga 1982, Torre et al 1991). However, the bioavailability of the minerals in cereals improves if phytate is degraded during food processing (Nävert et al 1985, Brune et al 1992). Dietary phytase can degrade phytate in the stomach and small intestine, which also improves mineral bioavailability (Sandberg et al 1987, Sandberg and Andersson 1988). By activating the nat- urally occurring enzyme phytase in cereals during food processing (soaking and malting), it is possible to degrade phytate to lower inositol phosphates, inorganic phosphate, and, in some cases, free myo-inositol (Cosgrove 1980). Phytase activity differs among cereals: rye has the highest activity, followed by wheat and barley, and oat has the lowest activity (Peers 1953, Bartnik and Szafranska 1987). The prooxidative enzyme lipoxygenase and the lipolytic enzyme lipase, which occur in barley, also may be active during process- ing, which limits the shelf-life of the final cereal product. To improve the oxidative stability of cereal products, it is important to min- imize the action of the lipoxygenase enzymes. Lipid oxidation during food processing may promote further oxidation during subsequent storage (Lingnert 1992). Yabuuchi (1976) hypothesized that lipoxy- genase is the main factor in off-flavor production in malt. Lipoxy- genase (EC 1.13.11.21) catalyzes the addition of molecular oxygen to the cis, cis-1,4-pentadiene system in polyunsaturated fatty acids, forming hydroperoxides. The hydroperoxides are decomposed or transformed into a variety of aldehydes and oxygenated fatty acids (Gardner 1988) that give rise to off-flavors. The undesirable flavors and hydroperoxides generated by lipoxygenase activity also are believed to interact with proteins, causing changes in food texture. Cereal lipids are rich in polyunsaturated fatty acids, making them good substrates for lipoxygenase action (Gardner 1988). Linoleic acid constitutes ≈57% and linolenic acid ≈5% of the fatty acid composition in barley (Morrison 1978). Lipase (EC 3.1.1.3) acts at the oil-water interface to release fatty acids from triglycerides (Brockenhoff and Jensen 1974), providing substrates for lipoxygenase enzymes that require the free acids as substrate. Degradation of fat, beginning with hydrolysis and followed by oxi- dation, leads to rancidification, which limits storage and handling of cereal products. Therefore, low lipase activity is desirable. The aim of this study was to find soaking conditions for barley cv. Blenheim that inactivate lipoxygenase and lipase but preserve phytase activity to develop possible procedures for production of barley products with potentially high mineral bioavailability and good oxidative stability. The influence of lactic acid concentration, temperature, and time on inactivation of enzyme activities was studied to optimize a soaking process. MATERIALS AND METHODS Barley Seed Whole barley kernels (cv. Blenheim) from the 1994 crop were provided by Skånska Lantmännen (Malmö, Sweden). Experimental Design A central composite face-centered design (CCF) was used with three variables and three replicates at the center point, for a total of 17 experiments. The three process variables studied included the lactic acid (La) concentration of the soaking agent (%, v/v), the temperature during soaking (T), and the soaking duration (t, min). Experimental conditions at the center point were La = 0.5%, T = 57°C, and t = 75 min. The experimental number, scaled values, and real values are given in Table I. The scaled values were x 1 = (La – 1 SIK, The Swedish Institute for Food and Biotechnology-AB, P.O. Box 5401, S-402 29 Göteborg, Sweden. 2 Chalmers University of Technology, Department of Food Science, P.O. Box 5401, S-402 29 Göteborg, Sweden. 3 Corresponding author. Phone: +46 31 35 56 07. Fax: +46 31 83 37 82. E-mail: Annika.Rutgersson@sik.se. Publication no. C-1997-1025-07R. © 1997 by the American Association of Cereal Chemists, Inc.