Nahit Gencer

New saccharin derivatives as tyrosinase inhibitors

A newly series of 6-(phenylurenyl/thiourenyl) saccharin (6a-y) derivatives were synthesized and their inhibitory effects on the diphenolase activity of banana tyrosinase were evaluated. A 70-fold purification of the enzyme with 6.85% yield was achieved by using a Sepharose 4B-l-tyrosine-p-amino benzoic acid affinity column. The result showed that all the synthesized compounds inhibited the tyrosinase enzyme activity. Among the compounds synthesized, 6-(3-iodophenylthiourenyl) saccharin (6s) was found to be most active one (K(i)=3.95 μM) and the inhibition kinetics analyzed by Lineweaver-Burk double reciprocal plots revealed that compound 6s was a competitive inhibitor. Structure-activity relationships study showed that generally, most of the 6-(phenylthiourenyl) saccharin derivatives (6m-y) exhibited higher inhibitory activity than 6-(phenylurenyl) saccharin derivatives (6a-l). An electron-withdrawing group at 3-position of phenylurenyl-ring increased in activity and the halogen series at 3-position of phenylthiourenyl-ring showed a qualitative relationship for higher inhibitory activity with increasing size and polarizability. We also calculated HOMO-LUMO energy levels and dipole moments of some selected the synthesized compounds (6a, 6h, 6m and 6s) using Gaussian software.

Purification human PON1Q192 and PON1R192 isoenzymes by hydrophobic interaction chromatography and investigation of the inhibition by metals

In this study, a new purification strategy for human PON1 enzyme was developed using two-step procedures, namely ammonium sulfate precipitation and sepharose-4B-L-tyrosine-9-aminophenantrene hydrophobic interaction chromatography. SDS polyacrylamide gel electrophoresis of the enzyme indicates a single band with an apparent MW of 43kDa. Overall purification rate of our method was found 901-fold for R isoenzyme and 453-fold for Q isoenzyme. The V(max) and K(M) of the purified enzyme were determined for Q isoenzyme 55 EU and 0.599 mM and for R isoenzyme 50 EU and 0.492 mM, respectively. The in vitro effects of some heavy metals (Hg, Cd, Cu, Mn and Ni) were investigated on the purified human serum PON1Q and R isoenzyme, using paraoxon as substrate. Metals were more effective inhibitors on purified human serum PON1(R192) activity than PON1(Q192) activity. The kinetics of interaction of metals with the purified human serum PON1(R192) and PON1(Q192) indicated a different inhibition pattern. Kinetic constants K(M), V(max), and inhibition type were determined.

Synthesis and carbonic anhydrase inhibitory properties of novel coumarin derivatives

A newly series of water-soluble 1-alkyl-3-(4-methyl-7, 8-dihydroxy-2H-chromen-2-one) benzimidazolium chloride salts (3a-j) were synthesized and their inhibitory effects on the activity of purified human carbonic anhydrase (hCA) I and II were evaluated. hCA I and II from human erythrocytes were purified by a simple one step procedure by using Sepharose 4B-L-tyrosine-sulphanilamide affinity column. The result showed that all the synthesized compounds were inhibited the CA isoenzymes activity. Among them, 3g and 3j were found to be most active (IC50 = 22.09 µM and 20.33 µM) for hCA I and hCA II, respectively.

Evaluation of in vitro effects of some analgesic drugs on erythrocyte and recombinant carbonic anhydrase I and II

The in vitro effects of the injectable form of analgesic drugs, dexketoprofen trometamol, dexamethasone sodium phosphate, metamizole sodium, diclofenac sodium, thiocolchicoside, on the activity of purified human carbonic anhydrase I and II were evaluated. The effect of these drugs on erythrocyte hCA I and hCA II was compared to recombinant hCA I and hCA II expressed in Ecoli. IC50 values of the drugs that caused inhibition were determined by means of activity percentage diagrams. The IC50 concentrations of dexketoprofen trometamol and dexamethasone sodium phosphate on hCA I were 683 μM and 4250 μM and for hCA II 950 μM and 6200 μM respectively. Conversely, the enzyme activity was increased by diflofenac sodium. In addition, thiocolchicoside has not any affect on hCA I and hCA II. The effect of these drugs on erythrocyte hCA I and hCA II were consistent with the inhibition of recombinant enzymes.

In vitro effects of some anabolic compounds on erythrocyte carbonic anhydrase I and II

The in vitro effects of the anabolic compounds, zeranol, 17 β-estradiol, diethylstilbestrol (DES), and trenbolone, on the activity of purified human carbonic anhydrase I and II were evaluated. In vitro CA enzyme activity was determined colorimetrically using the CO2 hydration method of Maren. IC50 values of the compounds that caused inhibition were determined by means of activity percentage diagrams. The IC50 concentrations of zeranol, 17 β-estradiol, DES and trenbolone on hCA I were 94, 55, 10, 898 µM and for hCA II 89, 159, 439 and 101 μM, respectively.

Purification and characterization of prophenoloxidase from Galleria mellonella L.

Abstract Prophenoloxidase (PPO) was purified from Galleria mellonella L. A 67-fold purification of the proenzyme with 352% yield was achieved by using a Sepharose 4B-L-tyrosine-p-amino benzoic acid affinity column. The purified enzyme was migrated as a single band on SDS-polyacrylamide gel electrophoresis. Km and Vmax values were 0.017 M and 1430.45 EU for catechol. Inhibition of PPO was investigated with inhibitors such as p-aminobenzoic acid, etyleneglycol, and ascorbic acid. Among them, ascorbic acid showed the strongest inhibitory activity with IC50 value of 2.94 μM. The current paper represents new strategies for the biological control of the Galleria mellonella L. insect.

Synthesis and carbonic anhydrase inhibitory properties of novel 1,4-dihydropyrimidinone substituted diarylureas

Abstract A new series of 1,4-dihydropyrimidinone (DHPM) substituted diaryl urea and thiourea derivatives were synthesized and their inhibitory effects on the activity of purified human carbonic anhydrase (hCA) I and II were evaluated. 4-Nitrophenyl-1,4-DHPM was prepared with dimedone, nitrobenzaldehyde and urea or thiourea and nitro group was reduced to amine derivative. The compound was reacted with isocyanates and isothiocyanates to get the final products. The results showed that all the synthesized compounds inhibited the carbonic anhydrase isoenzyme activity; 4c (IC50 = 66.23 µM for hCA I) and 4f (IC50 = 63.09 µM for hCA II) have the most inhibitory effect. The synthesized compounds are very bulky to be able to bind near the zinc ion and they much more probably bind as the coumarins and activators.

Effects of some metals on paraoxonase activity from shark Scyliorhinus canicula

Paraoxonase (PON) is an organophosphate hydrolyser enzyme which also has antioxidant properties in metabolism. Due to its crucial functions, the inhibition of the enzyme is undesirable and very dangerous. PON enzyme activity should not be altered in any case. Inhibitory investigations of this enzyme are therefore important and useful. Metal toxicology of enzymes has become popular in the recent years. Here, we report the in vitro inhibitory effects of some metal ions, including Ni2+, Cd2+, Cu2+ and Hg2+, on the activity of shark serum PON (SPON). For this purpose, we first purified the enzyme from shark Scyliorhinus canicula (LINNAEUS, 1758) serum and analysed the alterations in the enzyme activity in the presence of metal ions. The KM and Vmax is 0.227 mM and 454.545 U/mL, respectively. The results show that metal ions exhibit inhibitory effects on SPON1 at low concentrations with IC50 values ranging from 0.29 to 2.00 mM. Copper was determined to be the most effective inhibitor with IC50 of 0.29 mM.

In vitro efficacy of some cattle drugs on bovine serum paraoxonase 1 (PON1) activity

Serum paraoxonase 1 (EC 3.1.8.1, PON1), a calcium-associated enzyme, has an ability to hydrolyze organophosphate compounds. Related to this property, PON1 has a critical role in antioxidant mechanisms. It is well-known that the enzyme protects LDL from oxidation. In this study we investigated the in vitro inhibitory effects of some drugs. These drugs are oxytocin, dexamethasone, atropine sulphate, gentamicin sulphate, sulfadoxine-trimethoprim, furosemid, metamizole sodium and toldimfos sodium. The IC50 values obtained varied markedly from 0.014 to 507.72 mg/mL. According to our findings, most potent and significant inhibition was displayed by dexamethasone, atropine sulphate and furosemid.

In vitro inhibition effect and structure–activity relationships of some saccharin derivatives on erythrocyte carbonic anhydrase I and II

Abstract In this study, in vitro inhibitory effects of some saccharin derivatives on purified carbonic anhydrase I and II were investigated using CO2 as a substrate. The results showed that all compounds inhibited the hCA I and hCA II enzyme activities. Among the compounds, 6-(p-tolylthiourenyl) saccharin (6m) was found to be the most active one for hCA I activity (IC50 = 13.67 μM) and 6-(m-methoxyphenylurenyl) saccharin (6b) was found to be the most active one for hCA II activity (IC50 = 6.54 μM). Structure–activity relationships (SARs) study showed that, generally, thiourea derivatives (6l--v) inhibited more hCA I and hCA II than urea derivatives (6a–k). All compounds (excluding 6c and 6r) have higher inhibitory activity on hCA II than on hCA I.