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 lipase enzyme from Geobacillus stearothermophilus AH22 strain

Bu çalışmada, yeni izole edilmiş ve termofilik bir bakteri olan Geobacillus stearothermophilus AH22 suşundan endüstriyel bir enzim olan lipaz, iyon değişim kromatografisi ve jel filtrasyon kromatografisi kullanılarak kısmi olarak saflaştırıldı ve karakterize edildi. Lipaz enzimi %19.7 verimle 18.3 kat saflaştırılarak elde edildi. Kısmi saflaştırılmış lipaz enziminin moleküler ağırlığı SDS-PAGE kullanılarak yaklaşık 26 kDa olarak belirlendi. Enzim p-nitrofenil asetat, p-nitrofenil bütirat, p-nitrofenil oktaonat ve p-nitrofenil laurat substratları varlığında karakterize edildi. Enzimin Km ve Vmaks değerleri p-nitrofenil asetat, p-nitrofenil bütirat, p- nitrofenil oktaonat ve p- nitrofenil laurat substratları varlığında Lineweaver-Burk eğrisi yardımıyla belirlendi. Enzimin Km değerleri sırasıyla 0.16, 0.02, 0.19 ve 0.55 mM olarak, Vmaks değerleri ise sırasıyla 0.52 EU mg-1, 1.03 EU mg-1, 0.72 EU mg-1 ve 0.15 EU mg-1 olarak hesaplandı. Enzimin kullanılan substratlar varlığında en yüksek aktiviteyi 50 ile 60 oC' de ve pH 8 ile 9' da gösterdiği tespit edildi. Enzimin 50 ile 60 °C arasında termal kararlığının, +4 °C' de pH 8.0-10.0 aralığında ise pH karalılığının oldukça iyi olduğu gözlendi. Lipaz aktivitesine bazı metal tuzlarının ve bileşiklerin etkisi incelendi. Enzim aktivitesini en fazla Hg2+ iyonunun inhibe ettiği tespit edildi. Bununla birlikte orlistat, kateşin, propil paraben, p-kumarik asit ve 3,4- dihidroksi hidrosinamik asit inhibitörlerinin IC50 değerleri sırasıyla 0.0085, 0.06, 0.5, 1.25 ve 1.7 mM olarak belirlendi. Bu sonuçlar, Geobacillus stearothermophilus AH22 lipazının ısıl ve pH kararlılığı ve diğer özellikleri açısından endüstriyel uygulamalar için oldukça elverişli özelliklere sahip olduğunu göstermektedir. Anahtar kelimeler: Geobacillus stearothermophilus AH22, Lipaz, Saflaştırma,Karekterizasyon In this study, the lipase, as an industrial enzyme, was partially purified by ion exchange and gel filtration column chromatographies and was characterised from Geobacillus stearothermophilus AH22 strain. The lipase was purified 18.3-folds with 19.7% recovery. Protein samples obtained from the purification steps were migrated on SDS-PAGE and molecular weight of enzyme was found to be about 26 kDa. The lipase activity was determined by using p-nitrophenly acetate, p-nitrophenly butyrate, p-nitrophenly octaonate and p-nitrophenly laurate 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. Also Vmax values were found 0.52 EU mg-1, 1.03 EU mg-1, 0.72 EU mg-1and 0.15 EU mg-1. The enzyme showed maximum activity at 50 °C and pH ranging for in 8.0-9.0. It was observed that the enzyme was quite stable at pH range of 8.0-10.0 at +4 °C and thermal stability between 50 and 60 °C. The effect of some metal salts and compounds were investigated on the lipase activity. It was found that Hg2+ is the best inhibitor for this enzyme activity. The inhibitory effect of some chemicals including as orlistat, catechin, propyl paraben, p-coumaric acid, 3,4-dihydroxy hydro cinnamic acid, PMSF was examined and their IC50 values was calculated as 0.0085, 0.06, 0.5, 1.25, 1.7 and 1.6 mM, respectively. These results suggest that Geobacillus stearothermophilus AH22 lipase presents very suitable proporties for industrial applications. Keywords: Geobacillus stearothermophilus AH22, Lipase, Purification, Characterizatio

Partial purification and characterization of lipase from Geobacillus stearothermophilus AH22

Adiguzel, Ahmet/0000-0001-8848-6647WOS: 000368719800018PubMed: 25798692The 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 (C-2-C-12) as substrates. the K-m values of the enzyme for these substrates were found as 0.16, 0.02, 0.19 and 0.55mM, respectively, while V-max values were 0.52, 1.03, 0.72 and 0.15 Umg(-1). the enzyme showed maximum activity at 50 degrees 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 degrees C. It was found that the best inhibitory effect of the enzyme activity was of Hg2+. 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.Research Fund of Recep Tayyip Erdogan UniversityRecep Tayyip Erdogan University [2010.102.02.3]This work was financially supported by Research Fund of Recep Tayyip Erdogan University (Project No: 2010.102.02.3)