α-Amylase from Trichoderma harzianum was covalently immobilized on activated wool by cyanuric chloride. Immobilized α-amylase exhibited 75% of its initial activity after 10 runs. The soluble and immobilized α-amylases exhibited maximum activity at pH values 6.0 and 6.5, respectively. The immobilized enzyme was more thermally stable than the soluble one. Various substrates were hydrolyzed by immobilized α-amylase with high efficiencies compared to those of soluble α-amylase. The inhibition of the immobilized α-amylase by metal ions was low as compared with soluble enzyme. On the basis of the results obtained, immobilized α-amylase could be employed in the saccharification of starch processing.

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The aim of this study was to develop a chemical method for demythylation of curcumin. The methoxy groups reduced solubility and low bioavailability of curcumin. The treatment of curcumin with hydrogen bromide or choline chloride increased cucumin water solubility from 1 mg/mL to 30 or 25 mg/mL, respectively. 1 HNMR spectra showed that the chemical shift of O-methoxy groups at 3.9 ppm disappeared upon chemical treatment of curcumin, and indicated that these groups were removed especially after hydrogen bromide treatment. The antioxidant activity of treated curcumin and untreated curcumin was measured using different in vitro assays (i.e., 1,1-diphenyl-2-picrylhydrazyl and 2,2'-azinobis (3-ethylbenzo-thiazoline-6-sulfonic acid) radical scavenging, and phosphomolybdenum complex formation). A remarkable increase in 1,1-diphenyl-2-picrylhydrazyl and 2,2'-azino-bis (3-ethylbenzo-thiazoline-6-sulfonic acid) radical scavenging was observed as curcumin-hydrogen bromide > curcumin-Choline chloride > curcumin. The formation of phosphomolybdenum complex was found to increase in the order of curcumin-choline cloride > curcumin-hydrogen bromide > curcumin with EC 50 30, 41, and 114 µg/mL, respectively. In conclusion, hydrogen bromide-treated curcumin could be used as potential antioxidant in new functional foods.

A 1.2 kb BamHI fragment from pDK30 [Robinson, Kenan, Sweeney & Donachie (1986) J. Bacteriol. 167, 809-817] was cloned in pDOC55 [O'Connor & Timmis (1987) J. Bacteriol. 169, 4457-4482] to give two constructs, pDOC89 and pDOC87, in which the Escherichia coli D-alanine:D-alanine ligase (EC 6.3.2.4) gene (ddl) was placed under the control of the lac and lambda PL promoters respectively. Both constructs, when used to transform E. coli M72, gave similar levels of expression of the ddl gene. The expressed enzyme was purified to homogeneity and the amino acid sequence of its N-terminal region was found to be consistent with that predicted from the gene sequence, except that the N-terminal methionine was not present in the mature protein. [1(S)-Aminoethyl][(2RS)2-carboxy-1-octyl]phosphinic acid (I), previously shown to bind tightly to Enterococcus faecalis and Salmonella typhimurium D-alanine:D-alanine ligases following phosphorylation Parsons, Patchett, Bull, Schoen, Taub, Davidson, Combs, Springer, Gadebusch, Weissberger, Valiant, Mellin & Busch (1988) J. Med. Chem. 31, 1772-1778; Duncan & Walsh (1988) Biochemistry 27, 3709-3714], was found to be a classical slow-binding inhibitor of the E. coli ligase.

Background: Diabetic patients with hyperglycaemia show oxidative stress and increased formation of advanced glycation endproducts (AGEs) which increases their susceptibility to chronic complications. Aged garlic extract has antioxidant properties and prevents the formation of AGEs in vitro. This study investigated the effects of dietary intervention with Kyolic® aged garlic extract on glycaemia, lipidaemia and oxidative stress in diabetic patients. Methods: Blood samples were collected from 48 diabetic patients on recruitment, after one month and then monthly following an intake of 3000 mg of aged garlic extract daily over a period of 3-months. Samples were analysed for glucose, glycated haemoglobin and lipid profile using automated analyses. Low molecular weight AGEs were measured using a fluorometric method. Lipid hydroperoxides and total antioxidant status were determined using colorimetric kit methods. Results: Intervention with aged garlic extract did not affect blood glucose, glycated haemoglobin or the lipid profile but serum triacylglycerol concentrations declined after 3-months of intervention (P< 0.05). Aged garlic extract intake did reduce levels of serum AGEs although this was not significant. Lipid hydroperoxide, an indicator of oxidative stress, was significantly reduced following intake of aged garlic extract (P<0.05). Conclusion: Aged garlic extract may therefore protect against the harmful effects of AGEs and oxidative stress thus further investigations are needed to fully evaluate the benefits of long-term consumption of aged garlic extract, in particular its effects on tissue AGEs and oxidative stress.

BackgroundThe miswak (Salvadora persica) is a natural toothbrush. It is well known that very little information has been reported on enzymes in miswak as medicinal plant. Recently, we study peroxidase in miswak. In the present study, the main goal of this work is to purify and characterize α-amylase from miswak. The second goal is to study the storage stability of α-amylase in toothpaste.MethodThe purification method included chromatographaphy of miswak α-amylase on DEAE-Sepharose column and Sephacryl S-200 column. Molecular weight was determined by gel filtration and SDS-PAGE.ResultsFive α-amylases A1, A4a, A4b, A5a and A5b from miswak were purified and they had molecular weights of 14, 74, 16, 30 and 20 kDa, respectively. α-Amylases had optimum pH from 6 to 8. Affinity of the substrates toward all enzymes was studied. Miswak α-amylases A1, A4a, A4b, A5a and A5b had Km values for starch and glycogen of 3.7, 3.7, 7.1, 0.52, 4.3 mg/ml and 5.95, 5.9 4.16, 6.3, 6.49 mg/ml, respectively. The optimum temperature for five enzymes ranged 40°C- 60°C. Miswak α-amylases were stable up to 40°C- 60°C after incubation for 30 min. Ca+2 activated all the miswak α-amylases, while Ni2+, Co+2 and Zn+2 activated or inhibited some of these enzymes. The metal chelators, EDTA, sodium citrate and sodium oxalate had inhibitory effects on miswak α-amylases. PMSF, p-HMB, DTNB and 1,10 phenanthroline caused inhibitory effect on α-amylases. The analysis of hydrolytic products after starch hydrolysis by miswak α-amylases on paper chromatography revealed that glucose, maltose, maltotriose and oligosaccharide were the major products. Crude miswak α-amylase in the toothpaste retained 55% of its original activity after 10 months of storage at room temperature.ConclusionsFrom these findings, α-amylases from miswak can be considered as beneficial enzymes for pharmaceuticals. Therefore, we study the storage stability of the crude α-amylase of miswak, which contained the five α-amylases, in toothpaste. The enzyme in the toothpaste retained 55% of its original activity after 10 months of storage at room temperature.

Nanoparticles have the advantage of a superior surface area to volume ratio, and thus such materials are useful for enzyme immobilization. A silver nanoparticle coated cotton fabric (AgNp-CF) is used to immobilize camel liver catalase in the present work. The effect of loading levels of AgNp inside cotton fabrics on the immobilization of catalase was investigated. The results revealed that a 6 mL loading level of AgNp precursor (silver nitrate, 2 mM) at pH 8 showed the maximum immobilization efficiency (76%). The morphological properties of the cotton fabric (CF), AgNp-CF and AgNp-CF-catalase were characterized by SEM. The reusability of the immobilized enzyme was tested over ten reuses to show a 67% retained function of its initial activity. Compared with the soluble enzyme’s working pH (6.5), a rather broader working pH (6.5–7.0) was observed for the immobilized catalase. Additionally, the optimum working temperature increased from 30 for the soluble enzyme to 40 °C for the immobilized one, indicating thermal stability. The free and immobilized catalase enzyme’s Km values were 22.5 and 25 mM H2O2, respectively, reflecting the enzyme’s effective properties. The inhibitory effect of metal ions on the enzyme activity was higher toward soluble catalase than the immobilized catalase. This work has developed a method for immobilizing catalase to be useful for several applications.

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