Development of Fermented Rice Water to Improve the Quality of Garaetteok, a Traditional Korean Rice Cake.

In the rice processing industry, wastewater is an inevitable by-product of rice washing. To increase the utilization of washed rice water (WRW), seven types of fermented washed rice water (FWRW) were prepared using lactic acid bacteria (LAB) and carbohydrate hydrolase. The total concentration of small maltooligosaccharides (MOSs) in the amyloglucosidase (AMG) treatment groups was about ten times higher than in the untreated groups. After 6 h of fermentation, six of the seven FWRW samples reached a pH of 4 due to the increased concentration of organic acids and could, therefore, be used as food acidity regulators. To confirm the applicability of FWRW, the traditional Korean rice cake garaetteok was prepared with FWRW and stored at 4 °C for 5 days. A texture profile analysis (TPA) revealed that the hardness of garaetteok treated with FWRW was significantly lower than that of untreated garaetteok following storage. Differential scanning calorimetry (DSC) showed that FWRW retarded the retrogradation of garaetteok during storage. The addition of FWRW using Lactobacillus reuteri with an AMG group was particularly effective for inhibiting microbial activity in garaetteok during storage. These results suggest that FWRW using AMG-added L. reuteri can be used as a novel food additive for improving the quality of traditional Korean starch foods and could also reduce the volume of waste WRW.

Development and applications of enzymatic modified starch with high water solubility providing a continuous supply of glucose.

Amylopectin clusters (APCs) are produced by cyclodextrin glucanotransferase (EC 2.4.1.19). Their solubility rate in aqueous solution was found to be 16.7 %. The weight-average molecular weight of APCs is ∼105 Da, as determined by multiangle laser light scattering analysis. Side chain length analysis indicated that the relative proportions of side chains with a degree of polymerization in the ranges of 2-8 and 25-50 decreased and increased, respectively, during preparation of APCs. In the exercise experiment, the blood glucose level of rats was higher in the APC-treated group than in the groups treated with commercial carbohydrate supplement (CCD) and glucose. In the forced swimming test, the swimming time in the APC and CCD groups increased by 22.6 % and 31.1 %, respectively, compared with the glucose administration group. The insulin levels were also similar between the APC and CCD groups. However, the glycogen levels in the liver and muscles of mice were significantly higher in the APC group than control group. These results suggest that APCs could potentially enhance endurance when added to sports drinks.

Enzymatic formation of cyclic maltooligosaccharides for the application of quercetin inclusion complex.

To improve the applicability of quercetin (QCT), we produced a QCT and cycloamylose (CA-QCT) inclusion complex based on the cyclization activity of cyclodextrin glucanotransferase (CGTase; EC 2.4.1.19). The encapsulated QCT was purified using recycling preparative high-performance liquid chromatography, and its formation was analyzed using matrix-assisted laser desorption/ionization-time of flight mass spectrometry. The water solubility of CA-QCT was 55,000-fold higher than that of QCT. CA-QCT had 97 % stability for one week at pH 8 in a 4 °C water bath. According to a 2,2-diphenyl-1-picrylhydrazyl radical scavenging activity assay, CA-QCT activity in aqueous solution was 24 times higher than that of an equal amount of QCT in aqueous solution. In an anti-inflammatory assay using lipopolysaccharide-induced RAW264.7 macrophages, CA-QCT in aqueous solution decreased nitric oxide production in a similar manner to QCT in dimethyl sulfoxide (DMSO). Additionally, even under aqueous conditions, CA-QCT more effectively inhibited the production of inflammatory mediators, such as interleukin-1ß, interleukin-6, and cyclooxygenase, compared with QCT dissolved in DMSO.

Development of Freeze-Thaw Stable Starch through Enzymatic Modification.

The use of unmodified starch in frozen foods can cause extremely undesirable textural changes after the freeze-thaw process. In this study, using cyclodextrin glucanotransferase (CGTase) and branching enzymes, an amylopectin cluster with high freeze-thaw stability was produced, and was named CBAC. It was found to have a water solubility seven times higher, and a molecular weight 77 times lower, than corn starch. According to the results of a differential scanning calorimetry (DSC) analysis, dough containing 5% CBAC lost 19% less water than a control dough after three freeze-thaw cycles. During storage for 7 days at 4 °C, bread produced using CBAC-treated dough exhibited a 14% smaller retrogradation peak and 37% less hardness than a control dough, suggesting that CBAC could be a potential candidate for clean label starch, providing high-level food stability under repeated freeze-thaw conditions.

Development of an enzymatic encapsulation process for a cycloamylose inclusion complex with resveratrol.

Cyclodextrin glucanotransferase (CGTase; EC 2.4.1.19) produces cycloamyloses (CAs), which are large cyclic glucans, and subsequently transforms them to α-, ß-, and γ-cyclodextrins. We developed a novel encapsulation process based on the cyclization activity of CGTase and applied it to the formation of CA inclusion complexes with resveratrol (RVT), which has limited bioavailability due to its low water solubility. The encapsulated RVT (CA-RVT) was purified using preparative high-performance liquid chromatography. The water solubility of CA-RVT was 6,000-fold higher than that of RVT. CA-RVT in water demonstrated 98% stability for 1 week at 4 °C. According to radical scavenging activity and anti-inflammatory assays, CA-RVT in aqueous solution exhibited similar activities as an equal amount of RVT in dimethyl sulfoxide, suggesting the limited solubility of RVT can be overcome through CA encapsulation by CGTase, thus enhancing its nutraceutical value as a functional ingredient in the food industry.

Properties of recombinant 4-α-glucanotransferase from Bifidobacterium longum subsp. longum JCM 1217 and its application.

To determine the physiochemical properties of the 4-α-glucanotransferase from Bifidobacterium sp., the bllj_0114 gene encoding 4-α-glucanotransferase was cloned from Bifidobacterium longum subsp. longum JCM 1217 and expressed in Escherichia coli. The amino acid sequence alignment indicated that the recombinant protein, named BL-αGTase, belongs to the glycoside hydrolase (GH) family 77. BL-αGTase was purified using nickel-nitrilotriacetic acid affinity chromatography and characterized using various substrates. The enzyme catalyzed the disproportionation activity, which transfers a glucosyl unit from oligosaccharides to acceptor molecules, and had the highest activity at 40 °C and pH 6.0. In the presence of 5 mM metal ions, in particular Cu2+, Zn2+, and Fe2+, BL-αGTase activity was reduced. To determine whether BL-αGTase can be used to generate thermoreversible gels, potato starch was treated with BL-αGTase for various reaction times. The BL-αGTase-treated starches showed sol-gel reversibility and melted at 59.6-75.7 °C.

Characterization of novel α-galactosidase in glycohydrolase family 97 from Bacteroides thetaiotaomicron and its immobilization for industrial application.

Bacteroides thetaiotaomicron (B. thetaiotaomicron), which resides in the human intestinal tract, has a number of carbohydrate enzymes, including glycoside hydrolase (GH) family 97. Only a few GH 97 enzymes have been characterized to date. In this study, a novel α-galactosidase (Bt_3294) was cloned from B. thetaiotaomicron, expressed in Escherichia coli, and purified using affinity chromatography. This novel enzyme showed optimal activity at 60 °C and pH 7.0. Enzyme activity was reduced by 94.4% and 95.7% in the presence of 5 mM Ca2+ and Fe2+, respectively. It is interesting that Bt_3294 specifically hydrolyzed shorter α-galactosyl oligosaccharides, such as melibiose and raffinose. The D-values of Bt_3294 at 40 °C and 50 °C were about 107 and 6 min, respectively. After immobilization of Bt_3294, the D-values at 40 °C and 50 °C were about 37.6 and 29.7 times higher than those of the free enzyme, respectively. As a practical application, the immobilized Bt_3294 was used to hydrolyze raffinose family oligosaccharides (RFOs) in soy milk, decreasing the RFOs by 98.9%.