Some metals inhibit the glutathione S-transferase from Van Lake fish gills.

Glutathione S-transferases (GSTs) are the superfamily of multifunctional detoxification isoenzymes and play important role cellular signaling. The present article focuses on the role of Cd2+ , Cu2+ , Zn2+ , and Ag+ in vitro inhibition of GST. For this purpose, GST was purified from Van Lake fish (Chalcalburnus tarichii Pallas) gills with 110.664 EU mg-1 specific activity and 79.6% yield using GSH-agarose affinity chromatographic method. The metal ions were tested at various concentrations on in vitro GST activity. IC50 values were found for Cd+2 , Cu+2 , Zn+2 , Ag+ as 450.32, 320.25, 1510.13, and 16.43 µM, respectively. Ki constants were calculated as 197.05 ± 105.23, 333.10 ± 152.76, 1670.21 ± 665.43, and 0.433 ± 0.251 µM, respectively. Ag+ showed better inhibitory effect compared with the other metal ions. The inhibition mechanisms of Cd2+ and Cu2+ were non-competitive, whereas Zn2+ and Ag+ were competitive. Co2+ , Cr2+ , Pb2+ , and Fe3+ had no inhibitory activity on GST.

Synthesis and bioactivity of several new hetaryl sulfonamides.

1-(4-Methylsulfonyl)-2-thione-4-aryl-5-Z-6-methyl and oxyalkyl-imidazoles were synthesized from different tetrahydropyrimidinethiones and aryl sulfonyl chloride. These compunds were tested for metal chelating effects and to determine the phrase in which inhibition occured between two physiologically pertinent compunds and carbonic anhydrase (CA) isozymes I and II (hCA I and II), butyrylcholinesterase (BChE) and acetylcholinesterase (AChE). AChE was detected in high concentrations in the brain and red blood cells. BChE is another enzymes that is abundant available in the liver and released into the blood in a soluble form. Newly synthesized hetaryl sulfonamides exhibited impressive inhibition profiles with Ki values in the range of 1.42-6.58 nM against hCA I, 1.72-7.41 nM against hCA II, 0.20-1.14 nM against AChE and 1.55-5.92 nM against BChE. Moreover, acetazolamide showed Ki values of 43.69 ± 6.44 nM against hCA I and 31.67 ± 8.39 nM against hCA II. Additionally, tacrine showed Ki values of 25.75 ± 3.39 nM and 37.82 ± 2.08 against AChE and BChE, respectively.

Purification of glutathione S-transferase from Van Lake fish (Chalcalburnus tarichii Pallas) muscle and investigation of some metal ions effect on enzyme activity.

Glutathione S-transferases (GSTs) are an important enzyme family which play a critical role in detoxification system. In our study, GST was purified from muscle tissue of Chalcalburnus tarichii Pallas with 301.5-fold purification and 19.07% recovery by glutathione agarose affinity chromatography. The purity of enzyme was checked by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, showing a two band, because of having heterodimer structure. KM values were 1.59 and 0.53 mM for 1-chloro-2,4-dinitrobenzene (CDNB) and glutathione (GSH), respectively. Vmax values for CDNB and GSH were also determined as 5.58 and 1.88 EU/mL, respectively. In addition, inhibition effects of Ag(+), Cu(2+), Cd(2+), Fe(3+), Pb(2+), Cr(2+), Co(2+) and Zn(2+) metal ions were investigated on the enzyme activity and IC50, Ki values were calculated for these metal ions.

Heavy metal ion inhibition studies of human, sheep and fish α-carbonic anhydrases.

Carbonic anhydrases (CAs, EC 4.2.1.1) were purified from sheep kidney (sCA IV), from the liver of the teleost fish Dicentrarchus labrax (dCA) and from human erythrocytes (hCA I and hCA II). The purification procedure consisted of a single step affinity chromatography on Sepharose 4B-tyrosine-sulfanilamide. The kinetic parameters of these enzymes were determined for their esterase activity with 4-nitrophenyl acetate as substrate. The following metal ions, Pb(2+), Co(2+), Hg(2+), Cd(2+), Zn(2+), Se(2+), Cu(2+), Al(3+) and Mn(3+) showed inhibitory effects on these enzymes. The tested metal ions inhibited these CAs competitively in the low milimolar/submillimolar range. The susceptibility to various cations inhibitors differs significantly between these vertebrate α-CAs and is probably due to their binding to His64 or the histidine cluster.

Effects of some metal ions on rainbow trout erythrocytes glutathione S-transferase enzyme: an in vitro study.

Glutathione S-transferase enzyme (GST) (EC 2.5.1.18) was purified from rainbow trout erythrocytes, and some characteristics of the enzyme and effects of some metal ions on enzyme activity were investigated. For this purpose, erythrocyte glutathione S-transferase enzyme which has 16.54 EU/mg protein specific activities was purified 11,026-fold by glutathione-agarose affinity chromatography with a yield of 59%. Temperature was kept under control (+4°C) during purification. Enzyme purification was checked by performing SDS-PAGE. Optimal pH, stable pH, optimal temperature, and K(M) and Vmax values for GSH and 1-chloro-2, 4-dinitrobenzene (CDNB) were also determined for the enzyme. In addition, IC50 values, Ki constants and the type of inhibition were determined by means of Line-Weaver-Burk graphs obtained for such inhibitors as Ag(+); Cd(2+), Cr(2+) and Mg(2+).

Kinetic and docking studies of phenol-based inhibitors of carbonic anhydrase isoforms I, II, IX and XII evidence a new binding mode within the enzyme active site.

Carbonic anhydrases (CAs, EC 4.2.1.1) are inhibited by sulfonamides, inorganic anions, phenols, coumarins (acting as prodrugs) and polyamines. A novel class of CA inhibitors (CAIs), interacting with the CA isozymes I, II (cytosolic) and IX, XII (transmembrane, tumor-associated) in a different manner, is reported here. Kinetic measurements allowed us to identify hydroxy-/methoxy-substituted benzoic acids as well as di-/tri-methoxy benzenes as submicromolar-low micromolar inhibitors of the four CA isozymes. Molecular docking studies of a set of such inhibitors within CA I and II allowed us to understand the inhibition mechanism. This new class of inhibitors binds differently compared to all other classes of inhibitors known to date: they were found between the phenol-binding site and the coumarin-binding site, filling thus the middle of the enzyme cavity. They exploit different interactions with amino acid residues and water molecules from the CA active site compared to other classes of inhibitors, offering the possibility to design CAIs with an interesting inhibition profile compared to the clinically used sulfonamides/sulfamates.

In Vitro inhibition of human carbonic anhydrase I and II isozymes with natural phenolic compounds.

Inhibition of two human cytosolic carbonic anhydrase (hCA, EC 4.2.1.1) isozymes I and II with some natural phenolic derivatives was investigated using the esterase assay with 4-nitrophenyl acetate as substrate. Resveratrol, catechin, silymarin, dobutamin, and curcumin showed K(I) values in the range of 4.47-9.47 mm for hCA I and of 2.86-7.44 µm against hCA II, respectively. These natural product phenols were generally competitive inhibitors with 4-nitrophenylacetate as substrate. Some natural phenols investigated here showed effective hCA II inhibitory effects, in the same range as the clinically used sulfonamide acetazolamide, and might be used as leads for generating enzyme inhibitors possibly targeting other CA isoforms that have not been yet assayed for their interactions with such agents.

Carbonic anhydrase inhibitors. Inhibition of human erythrocyte isozymes I and II with a series of antioxidant phenols.

The inhibition of two human cytosolic carbonic anhydrase (hCA, EC 4.2.1.1) isozymes I and II, with a series of phenol derivatives was investigated by using the esterase assay, with 4-nitrophenyl acetate as substrate. 2,6-Dimethylphenol, 2,6-diisopropylphenol (propofol), 2,6-di-t-butylphenol, butylated hydroxytoluene, butylated hydroxyanisole, vanillin, guaiacol, di(2,6-dimethylphenol), di(2,6-diisopropylphenol), di(2,6-di-t-butylphenol), and acetazolamide showed K(I) values in the range of 37.5-274.5 microM for hCA I and of 0.29-113.5 microM against hCA II, respectively. All these phenols were non-competitive inhibitors with 4-nitrophenylacetate as substrate. Some antioxidant phenol derivatives investigated here showed effective hCA II inhibitory effects, in the same range as the clinically used sulfonamide acetazolamide, and might be used as leads for generating enzyme inhibitors possibly targeting other CA isoforms which have not been yet assayed for their interactions with such agents.

In vitro inhibition of salicylic acid derivatives on human cytosolic carbonic anhydrase isozymes I and II.

The inhibition of two human cytosolic carbonic anhydrase (hCA, EC 4.2.1.1) isozymes, hCA I and II, with a series of salicylic acid derivatives was investigated by using the esterase method with 4-nitrophenyl acetate as substrate. IC(50) values for sulfasalazine, diflunisal, 5-chlorosalicylic acid, dinitrosalicylic acid, 4-aminosalicylic acid, 4-sulfosalicylic acid, 5-sulfosalicylic acid, salicylic acid, acetylsalicylic acid (aspirin) and 3-metylsalicylic acid were of 3.04 microM, 3.38 microM, 4.07 microM, 7.64 microM, 0.13 mM, 0.29 mM, 0.42 mM, 0.56 mM, 2.71 mM and 3.07 mM for hCA I and of 4.49 microM, 2.70 microM, 0.72 microM, 2.80 microM, 0.75 mM, 0.72 mM, 0.29 mM, 0.68 mM, 1.16 mM and 4.70 mM for hCA II, respectively. Lineweaver-Burk plots were also used for the determination of the inhibition mechanism of these substituted phenols, most of which were noncompetitive inhibitors with this substrate. Some salicylic acid derivatives investigated here showed effective hCA I and II inhibitory activity, and might be used as leads for generating enzyme inhibitors eventually targeting other isoforms which have not been assayed yet for their interactions with such agents.

Amide derivatives with pyrazole carboxylic acids of 5-amino-1,3,4-thiadiazole 2-sulfonamide as new carbonic anhydrase inhibitors: synthesis and investigation of inhibitory effects.

Pyrazole carboxylic acid amides of 5-amino-1,3,4-thiadiazole-2-sulfonamide were synthesized from 4-benzoyl-1,5-diphenyl-1H-pyrazole-3-carbonyl chloride and 4-benzoyl-1-(3-nitrophenyl)-5-phenyl-1H-pyrazole-3-carbonyl chloride. Carbonic anhydrase isoenzymes (hCA-I and hCA-II) were purified from human erythrocyte cells by the affinity chromatography method. The inhibitory effects of 5-amino-1,3,4-thiadiazole-2-sulfonamide 1, acetazolamide 2 and new synthesized amides on these isozymes have been studied in vitro. The I(50) concentrations (the concentration of inhibitor producing a 50% inhibition of CA activity) against hydratase activity ranged from 1.2 to 2.2 nM for hCA-I and from 0.4 to 2 nM for hCA-II. The I(50) values against esterase activity ranged from 1.4 to 8 nM for hCA-I and from 1.3 to 6 nM for hCA-II. The K(i) values were observed between 8.2 x 10(- 5) to 6.2 x 10(- 4) M for hCA-I and between 2.9 x 10(- 4) to 8.2 x 10(- 4) M for hCA-II. The comparison of new synthesized amides to 5-amino-1,3,4-thiadiazole-2-sulfonamide 1, acetazolamide 2 indicated that the new synthesized compounds (18-23) inhibit CA activity more potently than the parent compounds.

Effects of some antibiotics on human erythrocyte glutathione reductase: an in vitro study.

Inhibitory effects of some antibiotics on purified human erythrocyte glutathione reductase were investigated. Human erythrocyte glutathione reductase was purified 2800-fold (29% yield) at 4 degrees C using 2', 5'-ADP Sepharose 4B affinity gel and Sephadex G-200 gel filtration chromatography. SDS polyacrylamide gel electrophoresis showed a single band for the enzyme. Imipenem, rifamycin, sulfanylacetamide, ceftazidime, chloramphenicol, seftriaxon, vancomycin, cefuroxime and ornidazole exhibited inhibitory effects but clindamycin, lincomycin, amoxicillin, amikacin exhibited activatory effects on the enzyme in vitro. The IC(50) values of imipenem, rifamycin, sulfanylacetamide, ceftazidime, chloramphenicol, seftriaxon, vancomycin, cefuroxime and ornidazole were 0.030, 0.146, 0.59, 2.476, 2.36, 2.88, 4.83, 15.43 and 19.632 mM, respectively, and the K(i) constants were 0.06 +/- 0.01, 0.275 +/- 0.10, 0.85 +/- 0.05, 3.59 +/- 0.51, 3.85 +/- 0.40, 3.71 +/- 0.60, 15.11 +/- 2.50, 23.50 +/- 2.94 and 28.49 +/- 6.50 mM, respectively. While imipenem, rifamycin, sulfanylacetamide, ceftazidime, chloramphenicol and seftriaxon cefuroxime and ornidazole showed competitive inhibition, vankomycine displayed noncompetitive inhibition.

In vitro inhibitory effects of some heavy metals on human erythrocyte carbonic anhydrases.

The inhibition of two human carbonic anhydrase (HCA, EC 4.2.1.1) isozymes, the cytosolic HCA I and II, with heavy metal salts of Pb(II), Co(II) and Hg(II) has been investigated. Human erythrocyte CA-I isozyme was purified with a specific activity of 920 EUmg(-1) and a yield of 30% and CA-II isozyme was purified with a specific activity of 8000 EUmg(-1) and a yield of 40% using Sepharose-4B-L tyrosine-sulfanilamide affinity gel chromatography. The overall purification was approximately 104-fold for HCA-I and 900-fold for HCA-II. The inhibitory effects of different heavy metals (lead, cobalt and mercury) on CA activity were determined at low concentrations using the esterase method under in vitro conditions. Ki values for these metals were calculated from Lineweaver-Burk graphs as 1.0, 3.22 and 1.45 mM for HCA-I and 0.059, 1.382 and 0.32 mM for HCA-II respectively. Lead was a noncompetitive inhibitor for HCA-I and competitive for HCA-II, cobalt was competitive for HCA-I and noncompetitive for HCA-II and mercury was uncompetitive for both HCA-I and HCA-II. Lead was the best inhibitor for both HCA-I and HCA-II.

Effects of some metal ions on human erythrocyte glutathione reductase: an in vitro study.

In this study, we investigated inhibitory effects of some metal ions on human erythrocyte glutathione reductase. For this purpose, initially human erythrocyte glutathione reductase was purified 1051-fold in a yield of 41% by using 2', 5'-ADP Sepharose 4B affinity gel and Sephadex G-200 gel filtration chromatography. SDS polyacrylamide gel electrophoresis was done in order to control the purification of enzyme. SDS polyacrylamide gel electrophoresis showed a single band for enzyme. A constant temperature (4 degrees C) was maintained during the purification process. Enzyme activity was determined with the Beutler method by using a spectrophotometer at 340 nm. Hg(2+), Cd(2+), Pb(2+), Cu(2+), Fe(3+) and Al3+ exhibited inhibitory effects on the enzyme in vitro. K(i) constants and IC(50) values for metal ions were determined by Lineweaver-Burk graphs and plotting activity % vs. [I]. IC(50) values of Pb(2+), Hg(2+), Cu(2+), Cd(2+), Fe(3+) and Al(3+) were 0.011, 0.020, 0.0252, 0.0373, 0.209 and 0.229 mM, and the Ki constants 0.0254+/-0.0027, 0.0378+/-0.0043, 0.0409+/-0.0048, 0.0558+/-0.0083, 0.403+/-0.043 and 1.137+/-0.2 mM, respectively. While Pb(2+), Hg(2+), Cd(2+) and Fe(3+) showed competitive inhibition, others displayed noncompetitive inhibition.

Effects of some drugs on purified human erythrocyte CuZnSOD and in vitro inhibitiory effect of 5-fluorouracil on leukocyte total SOD activity.

The inhibition and activation effects of some drugs on the activities of superoxide dismutase enzymes (SOD) in human erythrocyte and leukocyte cells was investigated. Firstly, CuZnSOD enzyme was purified 837-fold and 12% efficiency from human erythrocytes by ethanol-chloroform treatment to remove hemoglobin and then ion exchange chromatography (DEAE-Sepharose) and copper chelate affinity chromatography techniques. Inhibition or activation effects of fourteen drugs on CuZnSOD was investigated. None of the studied drugs except for 5-fluorouracil showed any effects on the enzyme. 5-fluorouracil showed activation effects on CuZnSOD at 3.33mg/ml and 4mg/ml concentrations with 33% and 32% activation, respectively. Leukocytes were isolated from healthy human blood, lysed in liquid nitrogen and the effect of 5-fluorouracil on the lysate SOD activity investigated. 5-Fluorouracil showed inhibition effects on total SOD activity of human leukocytes at 2 mg/ml and 4 mg/ml concentrations with 42% and 62% inhibition, respectively.

Effects of some antibiotics on activity of glucose-6-phosphate dehydrogenase from human erythrocytes in vitro and effect of isepamicin sulfate on activities of antioxidant enzymes in rat erythrocytes.

The purpose of this study was to investigate effects of some antibiotics on glucose-6-phosphate dehydrogenase (G6PD), antioxidant enzymes, and malondialdehyde (MDA). Initially, for in vitro studies, G6PD was purified from human erythrocyte, 9811-fold in a yield of 42.4% by using ammonium sulfate precipitation and 2',5' ADP-Sepharose 4B affinity gel. The purified enzyme showed a single band on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The effects of four different antibiotics (isepamicin sulfate, meropenem, chloramphenicol, and thiamphenicol glisinat hydrochloride) were investigated on the purified enzyme. K(i) value and type of inhibition were determined by means of Lineweaver-Burk graphs and regression analysis graphs. Isepamicin sulfate inhibited the enzyme activity (I(50) value, 2.1 mM; K(i) value, 1.7 mM), whereas thiamphenicol glisinat hydrochloride activated the G6PD dose dependently. Other drugs showed no inhibition and activation effect. In addition, the effects of isepamicin sulfate on the activities of G6PD, glutathione reductase (GR), superoxide dismutases (SOD), glutathione peroxidase (GPx), catalase (CAT), and glutathione S-transferase (GST) and MDA concentrations were examined in Sprague-Dawley rat erythrocytes in vivo. A marked alteration in the activities of these enzymes and MDA levels may be the result of oxidative stress in the rats receiving isepamicin sulfate.

Purification and characterization of polyphenol oxidase from nettle (Urtica dioica L.) and inhibitory effects of some chemicals on enzyme activity.

Polyphenol oxidase (PPO) of nettle (Urtica dioica L.) was extracted and purified through (NH4)2SO4 precipitation, dialysis, and CM-Sephadex ion-exchange chromatography and was used for its characterization. The PPO showed activity to catechol, 4-methylcatechol, L-3,4-dihydroxyphenylalanine (L-DOPA), L-tyrosine, p-cresol, pyrogallol, catechin and trans-cinnamic acid. For each of these eight substrates, optimum conditions such as pH and temperature were determined and L-tyrosine was found to be one of the most suitable substrates. Optimum pH and temperature were found at pH 4.5 and 30 degrees C respectively and Km and Vmax values were 7.90 x 10(-4) M, and 11290 EU/mL for with L-tyrosine as substrate. The inhibitory effect of several inhibitors, L-cysteine chloride, sodium azide, sodium cyanide, benzoic acid, salicylic acid, L-ascorbic acid, glutathione, thiourea, sodium diethyl dithiocarbamate, beta-mercaptoethanol and sodium metabisulfite were tested. The most effective was found to be sodium diethyl dithiocarbamate which acted as a competitive inhibitor with a Ki value of 1.79 x 10(-9)M. In addition one isoenzyme of PPO was detected by native polacrylamide slab gel electrophoresis.

Effects of low molecular weight plasma inhibitors of rainbow trout (Oncorhynchus mykiss) on human erythrocyte carbonic anhydrase-II isozyme activity in vitro and rat erythrocytes in vivo.

The effects of low molecular weight plasma inhibitors from rainbow trout (Oncorhynchus mykiss) (RT) were investigated on the carbonic anhydrase enzyme (CA) activities in in vitro human and in in vivo Sprague-Dawley rat erythrocytes. The RT blood was used as extracellular fluid (plasma) source and plasma inhibitors were obtained by dialysis of the plasma. For the in vitro study, human carbonic anhydrase-II (HCA-II) isozyme was obtained by Sepharose 4B-L-tyrosine-sulfanylamide affinity chromatography with an overall purification of about 646-fold. The enzyme (specific activity of 7750 EU/mg protein) was obtained with a yield of 71.1% and SDS-PAGE showed a single band. From in vitro studies, the I50 value for RT plasma inhibitors obtained was 0.37 mg/ml. From in vivo studies on rat erythrocytes, CA activity was significantly inhibited by the inhibitors from the extracellular fluid of RT for up to 3 h (p < 0.05) following intraperitoneal administration.

Investigation of the mutation points and effects of some drugs on glucose-6-phosphate dehydrogenase-deficient people in the Erzurum region.

We have carried out a systematic study of the molecular basis of glucose-6-phosphate dehydrogenase (G6PD) deficiency on three samples of 1,183 children aged 0.5-6 years from Erzurum, in eastern Anatolia. Total genomic DNAs were isolated from the blood samples of a healthy person and the three persons determined with G6PD deficiency by examining the enzyme activity and hemoglobin ratio. Then PCR amplification of the entire coding region in eight fragments was carried out followed by Agarose gel electrophoresis. The 540-bp PCR fragment containing exons VI-VII and the 550bp PCR fragment containing exons XI-XIII were digested with EcoRI and with NIaIII, respectively. SSCP techniques for eight fragments (exons II, III-IV, V, VI-VII, VIII, IX, X, and XI-XIII) were employed to determine the mutations on the exons of the G6PD gene. A mutation occurred on the region of the exons 6 and 7 of one person (person-1) and exon 5 of two G6PD-deficient persons (person 2 and 3) examined. The sequential approach described is fast and efficient and could be applied to other populations. Effects of analgesic drugs on G6PD were studied on the purified enzyme (ammonium fractionation, dialysis and 2',5' ADP-Sepharose 4B affinity chromatography) for the healthy person and G6PD-deficient persons 1, 2 and 3. The effects of remifentanil hydrochloride, fentanyl citrate, alfentanil hydrochloride and pethidine hydrochloride, as analgesic drugs, on G6PD activity were tested. Although remifentanil hydrochloride, fentanyl citrate (I50 values; 1.45mM and 6.1 mM, respectively) inhibited the activity of the enzyme belonging to the healthy person, they did not alter enzyme activity on two of the three persons with G6PD deficiency. Other drugs (alfentanil hydrochloride and pethidine hydrochloride) did not effect the enzyme activity of the healthy or G6PD-deficient children.

Effects of antiemetic drugs on glucose 6-phosphate dehydrogenase and some antioxidant enzymes.

The aim of this study was to investigate effect of antiemetics on G6PD and antioxidant enzymes. Antiemetics are currently being used to reduce or prevent nausea and vomiting in patients. This is the first study to show effect of antiemetics on glucose-6-phosphate dehydrogenase (G6PD) and antioxidant enzyme activities. For in vitro studies, G6PD was purified from human erythrocyte, 10, 26-fold in a yield of 51.3% by using ammonium sulphate precipitation and 2',5'-ADP-Sepharose 4B affinity gel. The purified enzyme showed a single band on sodium dodecyl sulfate-polyacrilamide gel electrophoresis (SDS-PAGE). The effects of four different antiemetics (granisetron hydrochloride, ondansetron hydrochloride, metoclopramide hydrochloride, trimethobenzamide hydrochloride) were investigated on the purified enzyme. Granisetron hydrochloride and ondansetron hydrochloride inhibited the enzyme activity (Ki values; 5.05 mM and 0.034 mM, I50 values; 3.9 mM and 0.036 mM, respectively). Metoclopramide hydrochloride, trimethobenzamide hydrochloride showed no inhibition effects. In addition, in vivo studies, effects of ondansetron hydrochloride on the G6PD, glutathione reductase (GR), glutathione peroxidase (GPx) and catalase (CAT) were examined in the rat erythrocytes. G6PD (49% of control), GR (55% of control), CAT (60% of control) activities in erythrocytes were significantly decreased whereas GPx (183% of control) was significantly increased. A marked alteration in these enzymes may be result of oxidative stress in the rats receiving ondansetron hydrochloride.

Investigation of glucose 6-phosphate dehydrogenase (G6PD) kinetics for normal and G6PD-deficient persons and the effects of some drugs.

In the present study, blood samples from 1183 children aged 0.5-6 years were taken. Three children were found with G6PD deficiency by examining the enzyme activity and hemoglobin ratio. Some kinetic properties of glucose 6-phosphate dehydrogenase enzyme (G6PD) were studied after the purification of the enzyme with ammonium fractionation, dialysis and 2',5' ADP-Sepharose 4B affinity chromatography from a healthy person and from three G6PD-deficient people. The purity of the enzymes was confirmed by SDS-PAGE electrophoresis. The effects of some drugs which are known inhibitors of G6PD activity were studied. Some of the drugs stimulated the activity of the enzyme in two of the three cases with G6PD deficiency. KM values, Vmax values for G6P and NADP+, optimum pH and optimum temperature for the enzyme from the healthy person and the three G6PD-defficient people are reported.