Ali Atasever

One-step purification of lactoperoxidase from bovine milk by affinity chromatography

Sulphanilamide was determined to be a new inhibitor of lactoperoxidase (LPO) with an IC(50) of 0.848.10(-5)M. The K(i) for sulphanilamide was determined to be 3.57.10(-5)M and sulphanilamide showed competitive inhibition, which makes it a suitable ligand for constructing a Sepharose 4B-L-tyrosine affinity matrix. The affinity matrix was synthesised by coupling sulphanilamide as the ligand and L-tyrosine as the spacer arm to a cyanogen bromide (CNBr)-activated-Sepharose 4B matrix. Lactoperoxidase was purified 409-fold from the synthesized affinity matrix in a single step, with a yield of 62.3% and a specific activity of 40.9 EU/mg protein. The enzyme activity was measured using ABTS as a chromogenic substrate (pH 6.0). The degree of LPO purification was monitored by SDS-PAGE and its R(z) (A(412)/A(280)) value. The R(z) value for the purified LPO was found to be 0.7. Maximum binding was achieved and K(m) and V(max) values were determined.

The effects of some avermectins on bovine carbonic anhydrase enzyme

Abstract Avermectins are effective agricultural pesticides and antiparasitic agents that are widely employed in the agricultural, veterinary and medical fields. The aim of this study was to investigate the inhibitory effects of selected avermectins including abamectin, doramectin, emamectin, eprinomectin, ivermectin and moxidectin that are used as drugs against a wide variety of internal and external mammalian parasites, on the carbonic anhydrase enzyme (CA, EC 4.2.1.1.) purified from fresh bovine erythrocyte. CA catalyses the rapid interconversion of carbon dioxide (CO2) and water (H2O) to bicarbonate () and protons (H+) and regulate the acidity of the local tissues. Bovine erythrocyte CA (bCA) enzyme was purified by Sepharose-4B affinity chromatography with a yield of 21.96% and 262.7-fold purification. The inhibition results obtained from this study showed Ki values of 9.73, 17.39, 20.43, 13.39, 16.44 and 17.73 nM for abamectin, doramectin, emamectin, eprinomectin, ivermectin and moxidectin, respectively. However, acetazolamide, well-known clinically established CA inhibitor, possessed a Ki value of 27.68 nM.

Impact of Some Avermectins on Lactoperoxidase in Bovine Milk

Many macrocyclic lactones, including avermectins, are known to be used as a veterinary drug, agricultural pesticides, and insecticides. Lactoperoxidase (EC 1.11.1.7) is one of the peroxidases found in milk. Lactoperoxidase has a natural host defense system against micro-organisms and a natural antimicrobial system. In this study, some macrocyclic lactones, including emamectin-benzoate, doramectin, eprinomectin, abamectin, moxidectin-vetranal, and ivermectin were investigated for in vitro inhibitory effects on the bovine lactoperoxidase enzyme, which was purified using amberlite CG-50 H+ resin and sepharose 4B-L-tyrosine-sulphanamide affinity chromatography 344.6-fold, with a yield of 61.1% and a specific activity of 39.11 EU/mg protein. Emamectin-benzoate, doramectin, eprinomectin, abamectin, moxidectin-vetranal, and ivermectin are also known strong antiparasitary properties. In this study, we demonstrated that avermectins have strong lactoperoxidase inhibitory effects. Of these, the emamectin-benzoate was shown to have the most inhibiting effect against lactoperoxidase with Ki value of 6.82 ± 2.60 µM.

Single-step purification of peroxidase by 4-aminobenzohydrazide from Turkish blackradish and Turnip roots.

Peroxidases (PODs) were purified from the Turkish blackradish (Raphanus sativus L.) (TBR) and Turnip (Brassica rapa L.) using a simple and effective single-step method. An affinity resin was synthesised by coupling the 4-aminobenzohydrazide ligand and the l-tyrosine spacer-arm to CNBr-activated-Sepharose-4B. The purification factors for the TBR-POD and the Turnip-POD were 40.3-fold (with a yield of 10.6%) and 269.3-fold (with a yield of 9%), respectively. The molecular masses of the TBR-POD and Turnip-POD were approximately 67.3 and 65.8kDa, respectively. For guaiacol, the Km and Vmax values were calculated as 24.88mM and 3.23EU/mL, respectively for TBR-POD and as 4.09mM and 0.797EU/mL for the Turnip-POD. For H2O2, the Km and Vmax values were calculated as 3.247mM and 0.799EU/mL, respectively for TBR-POD, and as 12.49mM and 4.055EU/mL, respectively for the Turnip-POD. Furthermore, 4-aminobenzohydrazide was determined to be a non-competitive inhibitor of TBR-POD and Turnip-POD.

2-Amino-3-cyanopyridine derivatives as carbonic anhydrase inhibitors.

Carbonic anhydrases (CAs, EC 4.2.1.1) are ubiquitous enzymes that catalyze the hydration of CO2 to bicarbonate and protons. Inhibition of CAs has been clinically exploited for the treatment of various classes of diseases for decades, but investigating new classes of inhibitors continues to be important. We have synthesized a series of 2-amino-3-cyano-4-heteroaryl (5a–l) compounds and characterized the structures by NMR, IR and elemental analyses. We tested the ability of these compounds to inhibit two metalloenzyme human carbonic anhydrase (hCA, EC 4.2.1.1) isozymes, hCA I and hCA II. Compounds 5d and 5b showed the best inhibition activity against hCA I (IC50: 33 and 34 µM, respectively), and compound 5d showed the best activity against hCA II (IC50: 56 µM).

Synthesis of 2‐amino‐3‐cyanopyridine derivatives and investigation of their carbonic anhydrase inhibition effects

The conversion of carbon dioxide (CO2) and bicarbonate (HCO3−) to each other is very important for living metabolism. Carbonic anhydrase (CA, E.C.4.2.1.1), a metalloenzyme familly, catalyzes the interconversion of these ions (CO2 and HCO3−) and are very common in living organisms. In this study, a series of novel 2‐amino‐3‐cyanopyridines supported with some functional groups was synthesized and tested as potential inhibition effects against both cytosolic human CA I and II isoenzymes (hCA I and II) using by Sepharose‐4B‐l‐tyrosine‐sulfanilamide affinity chromatography. The structural elucidations of novel 2‐amino‐3‐cyanopyridines were achieved by NMR, IR, and elemental analyses. Ki values of the novel synthesized compounds were found in range of 2.84–112.44 μM against hCA I and 2.56–31.17 μM against hCA II isoenzyme. While compound 7d showed the best inhibition activity against hCA I (Ki: 2.84 μM), the compound 7b demonstrated the best inhibition profile against hCA II isoenzyme (Ki: 2.56 μM).

Katı ve Sıvı Taşıyıcı Kullanılarak Bitki Gelişmesini Teşvik Edici Bakteri Esaslı Formülasyon Geliştirilmesi ve Çay Gelişme Parametreleri ve Enzim Aktivitesine Etkisinin Değerlendirilmesi

Bu arastirma, mineral gubre, bir ticari biyolojik gubre ve organik ve inorganik kaynakli kati (cay artigi, torf, perlit, leonardit, zeolite ve vermikulit) ve sivi tasiyicilarda gelistirilen azot fikseri ve fosfat cozucu mikroorganizma esasli bes farkli biyolojik gubre formulasyonunun (F1: Bacillus atrophaeus 55/6+ Bacillus megaterium 63/1+ Pseudomonas fluorescens 9/7; F2: Bacillus simplex 6/4+ Bacillus subtlis B2/8+ Pseudomonas putida 3/10; F3: Bacillus pumilus 35/6+ Paenibacillus polymyxa 2/2+ Pseudomonas sp . 27/3; F4: Bacillus pumilus RC19+ Paenibacillus polymyxa RC35+ Pseudomonas putida 29/2; F5: Bacillus lentus 29/6+ Paenibacillus polymyxa RC14+ Pseudomona putida RC06), cay gelismesi ve enzim aktivitesi uzerine etkisini degerlendirmek amaciyla Rize’de dogal kosullar altinda iki yil sureyle yurutulmustur. Arastirma, yedi tasiyici, sekiz uygulama ve her bir tekerrurde dort fidan olmak uzere bes tekerrurlu olarak tesaduf parselleri deneme desenine gore duzenlenmistir. Bakteriyel formulasyon etkinligi asilanan strain, tasiyici materyal ve ele alinan parametrelere gore degismistir. Biyolojik gubre formulasyonu Hayrat Turk cay klonunda govde gelismesi, bitki yuksekligi, govde capi, yaprak verimi, klorofil ve antioksidan icerigi, yaprak alani, oksidatif, katalitik, hidrolitik ve antioksidan enzim aktivitesi dahil gelismeyi tesvik etmistir. Arastirmada test edilen bakteri kombinasyonlarinin bitki gelismesini tesvik ettigi, kimyasal gubre gereksinimini azaltabildigi, organik ve surdurulebilir tarim uygulamalarinda biyolojik gubre olarak kullanilabilecek potansiyele sahip oldugu belirlenmistir.