Characterization and inhibition studies of carbonic anhydrase from gill of Russian Sturgeon Fish (Acipenser gueldenstaedtii).

An α-carbonic anhydrase (CA, EC 4.2.1.1) was purified and characterized kinetically from gill of Acipenser gueldenstaedtii as an endangered sturgeon species. The carbonic anhydrase was purified 66-folds with yield 20.7% by Sepharose-4B-l-tyrosine-sulfanilamide affinity column and the specific activity was determined as 222.2 EU/mg protein. Km and Vmax kinetic values for gill carbonic anhydrase were calculated by a Lineweaver-Burk graph using p-nitrophenol acetate (p-NPA) as a substrate, and was defined as 2.5 mM and 5 × 10(6 )µM/min, respectively. It was observed that CA from the sturgeon gill in the presence of the sulfanilamide and acetazolamide as an inhibitor had very low IC50 values such as 13.0 and 0.1 µM, respectively. In addition, it was determined that the enzyme was inhibited by Fe(2+,) Co(2+,) Ni(2+), and Zn(2+)-Ba(2+) with the IC50 values of 0.2, 1.7, 1.2, and 1.1 mM, respectively.

Partial purification and characterization of lipase from Geobacillus stearothermophilus AH22.

The lipase was partially purified by ion exchange chromatography and gel filtration column chromatography, and was characterized from Geobacillus stearothermophilus AH22 strain. The lipase was purified 18.3-folds with 19.7% recovery. The lipase activity was determined by using p-nitrophenyl esters (C2-C12) as substrates. The Km values of the enzyme for these substrates were found as 0.16, 0.02, 0.19 and 0.55 mM, respectively, while Vmax values were 0.52, 1.03, 0.72 and 0.15 U mg(-1). The enzyme showed maximum activity at 50 °C and between pH 8.0 and 9.0. The enzyme was found to be quite stable at pH range of 4.0-10.0, and thermal stability between 50 and 60 °C. It was found that the best inhibitory effect of the enzyme activity was of Hg(2+). The inhibitory effect as orlistat, catechin, propyl paraben, p-coumaric acid, 3,4-dihydroxy hydro-cinnamic acid was examined. These results suggest that G. stearothermophilus AH22 lipase presents very suitable properties for industrial applications.

Protective roles of Cimin grape tissues on oxidative stress markers in the cellular system model.

AIM: To analyze the phenolic composition of the Cimin grape, which is 1 of 2 grape varieties with a protected geographical indication in Turkey and is used locally to treat the symptoms of some disorders such as impotence and cardiovascular diseases, and to investigate its antioxidant potency against oxidant mediators in the models of intra- and extracellular surroundings. MATERIALS AND METHODS: Cimin grape samples were classified into 5 groups according to the grape's tissues and the extraction solvents used. Free radical scavenging (DPPH) and antilipid peroxidation product [thiobarbituric acid reactive substance (TBARS) and conjugated diene (CD)] levels of the grape tissues were extrapolated from the measurement of total phenolic and individual monomeric flavanol contents in each extract. RESULTS: The seed methanolic extract had the highest total amount of flavanols, with the amount of catechin at 4.034 mM. The DPPH activities of the seed extracts were 2- to 10-fold higher than those of the other samples. The seed extract-treated erythrocyte and unfractionated human plasma also showed lower TBARS and CD values. In addition, regeneration of glutathione was more obvious in grape seed extracts than in the rest of the tissues. CONCLUSION: The underlying mechanism of these changes can be related mainly to increased antioxidant status. Cimin grape consumption may have beneficial effects on health maintenance.

Purification and characterization of extracellular α-amylase from a thermophilic Anoxybacillus thermarum A4 strain

ABSTRACT α-Amylase from Anoxybacillus thermarum A4 was purified using ammonium sulphate precipitation and Sephadex G-100 gel filtration chromatography, with 29.8-fold purification and 74.6% yield. A4 amylase showed best performance for soluble potato starch hydrolysis at 70 °C and pH 5.5-10.5. A4 amylase was extremely stable at +4 °C, and the enzyme retained over 65% of its original α-amylase activity at 70 °C and 43% at 90 °C. The enzyme's Km values for soluble starch, amylopectin and amylose substrates were obtained as 0.9, 1.3 and 0.5 mg/mL, respectively. EDTA, Hg2+, B4O7 2-, OH-, CN- , and urea exhibited different inhibition effects; their IC50 values were identified as 8.0, 5.75, 16.5, 15.2, 8.2 and 10.9 mM, respectively. A4 amylase exhibited extreme stability toward some surfactants and perfect match for a wide variety of commercial solid and liquid detergents at 55 °C. So, it may be considered to be potential applications for detergent and other industrial uses.