The methaemoglobin forming and GSH depleting effects of dapsone and monoacetyl dapsone hydroxylamines in human diabetic and non-diabetic erythrocytes in vitro

The respective methaemoglobin forming and GSH depleting capabilities of monoacetyl dapsone hydroxylamine (MADDS-NHOH) and dapsone hydroxylamine (DDS-NHOH) were compared in human diabetic and non-diabetic erythrocytes in vitro with a view to select the most potent agent for future oxidative stress and antioxidant evaluation studies. Administration of both metabolites to non-diabetic erythrocytes over the 20min period of the study resulted in significantly more methaemoglobin formation at all four time points compared with the diabetic erythrocytes (P<0.0001). At all four time points, significantly more methaemoglobin was formed in response to MADDS-NHOH in non-diabetic cells compared with the effects of DDS-NHOH on diabetic erythrocytes (P<0.0001). At the 5 and 10min time points, significantly more methaemglobin was formed in non-diabetic cells in the presence of MADDS-NHOH compared with DDS-NHOH (P<0.05). At the 5min time point only, significantly more methaemoglobin was formed in the presence of MADDS-NHOH in diabetic cells compared with that of DDS-NHOH (P<0.01). However, compared with diabetic control GSH levels, the presence of DDS-NHOH caused a significant depletion in GSH at 5, 10 and 20min time points in diabetic cells (P<0.001). In addition, the presence of DDS-NHOH caused a significant reduction in GSH levels in diabetic cells in comparison with those of non-diabetics at the 5, 10 and 20min, (P<0.005). DDS-NHOH was also associated with a significant depletion of GSH levels in diabetic cells compared with those of non-diabetic control erythrocytes (P<0.0001). The presence of MADDS-NHOH in diabetic erythrocytes led to a significant reduction in GSH levels at the 20min time point compared with those of non-diabetics (P<0.001), but there were no significant differences at the 5, 10 and 15min points. Due to its greater GSH-depleting action, DDS-NHOH will be selected for future use in the oxidative stress assessment in diabetic erythrocytes.     

Studies on the differential sensitivity between diabetic and non-diabetic human erythrocytes to monoacetyl dapsone hydroxylamine-mediated methaemoglobin formation in vitro

Methaemoglobin generation by monoacetyl dapsone hydroxylamine in non-diabetic and diabetic erythrocytes was investigated in vitro. Methaemoglobin formation in purified haemoglobin isolated from both types of erythrocytes as well as haemolysates from both diabetic and non-diabetic erythrocytes did not differ. Prior to 18 h incubation with 10 and 20 mM glucose diabetic erythrocytes were significantly less sensitive to monoacetyl dapsone-induced methaemoglobinaemia. After pre-incubation the differential was lost although significant change in glutathione concentrations could not be shown between the two cell types. NADH-diaphorase levels measured in diabetics and non-diabetics did not significantly differ. It is possible that diabetic cells display reduced hydroxylamine-mediated methaemoglobin generation due to differences in glutathione metabolism.     

A preliminary evaluation of a novel method to monitor a triple antioxidant combination (vitamins E, C and α-lipoic acid) in diabetic volunteers using in vitro methaemoglobin formation

Eight otherwise healthy diabetic volunteers took a daily antioxidant supplement consisting of vitamin E (200 IU), vitamin C (250 mg) and α-lipoic acid (90 mg) for a period of 6 weeks. Diabetic dapsone hydroxylamine-mediated methaemoglobin formation and resistance to erythrocytic thiol depletion was compared with age and sex-matched non-diabetic subjects. At time zero, methaemoglobin formation in the non-diabetic subjects was greater at all four time points compared with that of the diabetic subjects. Resistance to glutathione depletion was initially greater in non-diabetic compared with diabetic samples. Half-way through the study (3 weeks), there were no differences between the two groups in methaemoglobin formation and thiol depletion in the diabetic samples was now lower than the non-diabetic samples at 10 and 20 min. At 6 weeks, diabetic erythrocytic thiol levels remained greater than those of non-diabetics. HbA(1c) values were significantly reduced in the diabetic subjects at 6 weeks compared with time zero values. At 10 weeks, 4 weeks after the end of supplementation, the diabetic HbA1(c) values significantly increased to the point where they were not significantly different from the time zero values. Total antioxidant status measurement (TAS) indicated that diabetic plasma antioxidant capacity was significantly improved during antioxidant supplementation. Conversion of α-lipoic acid to dihydrolipoic acid (DHLA) in vivo led to potent interference in a standard fructosamine assay kit, negating its use in this study. This report suggests that triple antioxidant therapy in diabetic volunteers attenuates the in vitro experimental oxidative stress of methaemoglobin formation and reduces haemoglobin glycation in vivo.     

The use of xenobiotic-mediated methaemoglobin formation to assess the effects of thyroid hormones on diabetic and non-diabetic human erythrocytic oxidant defence mechanisms in vitro

Diabetes is associated with an abnormal incidence of hypothyroidism, which exacerbates hyperglycaemia, so further damaging already compromised erythrocytic defence mechanisms. Methaemoglobin formation is a useful measure of the health of these mechanisms, as it determines the resistance of diabetic erythrocytes to sustained oxidative stress. The effect of l-tri-iodothyronine (T(3)) was, therefore, studied on nitrite and monoacetyldapsone hydroxylamine (MADDS-NHOH) mediated methaemoglobin formation in diabetic and non-diabetic human erythrocytes. Diabetic erythrocytes showed less sensitivity compared with non-diabetics to methaemoglobin formation mediated by both compounds. A 30 min pre-incubation with T(3) at 3 and 30 nM did not affect nitrite-mediated methaemoglobin formation compared with control observations in both cell types. In diabetic erythrocytes incubated with T(3) at 30 nM, there were significant increases in MADDS-NHOH-mediated methaemoglobin formation compared with control in both diabetic and non-diabetic cells. Studies comparing blood isolated from diabetic patients stabilised on thyroxine (T(4); 50 μG/day), T(4)-free diabetics and non-diabetics, showed that T(4) supplementation significantly increased MADDS-NHOH-mediated methaemoglobin formation compared with T(4)-free diabetic cells so that for two time points, T(4)-treated diabetic erythrocytic methaemoglobin formation was indistinguishable from that of non-diabetics. These studies indicate that T(4) supplementation improves some erythrocytic oxidant defence mechanisms in a time dependent manner.     

Methaemoglobin formation due to nitrite, disulfiram, 4-aminophenol and monoacetyldapsone hydroxylamine in diabetic and non-diabetic human erythrocytes in vitro

Nitrite, monoacetyl dapsone hydroxylamine, 4-aminophenol and disulfiram-mediated methaemoglobin formation was studied in human diabetic and non-diabetic erythrocytes in vitro. Diabetic intact erythrocytes were significantly less sensitive compared with those of non-diabetics to haemoglobin oxidation caused by the hydroxylamine, nitrite and 4-aminophenol, but not disulfiram. In haemolysates, differential sensitivity did occur with disulfiram and was partially retained with 4-aminophenol and nitrite. The differences were lost with 4-aminophenol, nitrite and disulfiram in the presence of haemoglobin purified from the respective erythrocyte types. Diethyl maleate reduced methaemoglobin formation in non-diabetic intact erythrocytes with 4-aminophenol, the hydroxylamine and disulfiram, but not with nitrite. Overall, the differential sensitivity to methaemoglobin formation seen in diabetic compared with non-diabetic erythrocytes, is probably linked to differences in the respective cells' cytosolic anti-oxidant systems.     

Monitoring diabetic antioxidant status: a role for in vitro methaemoglobin formation

Diabetes leads to premature organ and system failure and considerably shortens lifespan. Careful control of glucose levels may not be enough to prevent the onset of complications in most diabetics. Compared with non-diabetics, diabetic tissues must not only resist a much greater long-term threat from hyperglycaemia-mediated reactive species but also defend themselves with compromised antioxidant systems. Although antioxidant therapy is a logical step in the prevention of oxidant and carbonyl stresses in the face of intermittent hyperglycaemia, this approach is not yet universally accepted to be effective in preventing complications. Although there are many biochemical indices of oxidant stress, piecemeal elevations of individual markers may not necessarily reflect true diabetic cellular antioxidant status. A dynamic process such as in vitro methaemoglobin generation may provide an opportunity to compare the response of a diabetic erythrocyte with that of a non-diabetic before and after corrective antioxidant therapy. Due to compromised cellular antioxidant capacity, diabetic cells generate less methaemoglobin in the presence of aromatic amine hydroxylamines, 4-aminophenol and nitrite compared with non-diabetics. Agents such as dihydrolipoic acid have been shown to correct methaemoglobin formation-mediated thiol deficits during in vitro studies. It is hoped that the progress of antioxidant supplementation studies in diabetics can be monitored with the aid of in vitro methaemoglobin generation using agents such as hydroxylamines, 4-aminophenol and nitrite. The most appropriate antioxidants and dosages can thus be recommended to diabetics worldwide to attenuate the development of complications.     

Effects of oxidised alpha-lipoic acid and alpha-tocopherol on xenobiotic-mediated methaemoglobin formation in diabetic and non-diabetic human erythrocytes in-vitro

The effects of oxidised alpha-lipoic acid and alpha-tocopherol were investigated on a human erythrocytic in vitro model of diabetic metabolic stress. Preincubation of non-diabetic and diabetic erythrocytes with oxidised alpha-lipoic acid or alpha-tocopherol resulted in marked increases in nitrite-mediated methaemoglobin formation. In contrast, oxidised alpha-lipoic acid resulted in considerable reductions in 4-aminophenol-mediated methaemoglobin formation in both diabetic and non-diabetic cells. alpha-Tocopherol showed an increase only in diabetic cells, at one time point. Monoacetyl dapsone hydroxylamine (MADDS-NHOH)-mediated methaemoglobin formation was reduced by oxidised alpha-lipoic acid in non-diabetic and diabetic cells at all three time points, although alpha-tocopherol had no effect with MADDS-NHOH. In diabetic cells only, alpha-tocopherol incubation caused a reduction in GSH levels compared with non-diabetic cells. As the agents showed pro- as well as anti-oxidant effects in this study, further studies are required to demonstrate potential diabetic benefit from alpha-lipoic acid adminstration.     

Studies on the mechanisms of oxidation in the erythrocyte by metabolites of primaquine

The interaction of certain metabolites of the 8-aminoquinoline antimalarial primaquine with both normal and glucose-6-phosphate dehydrogenase (G6PD)-deficient erythrocytes and with haemoglobin preparations was studied in an attempt to elucidate the mechanisms of methaemoglobin formation and haemolytic anaemia associated with the use of primaquine. Studies using erythrocytes revealed that oxidation of haemoglobin and reduced glutathione (GSH) was due to the metabolites rather than the parent drug. Incubation of free haemoglobin with 5-hydroxylated metabolites of primaquine also led to oxidation of oxyhaemoglobin and GSH. Oxidation of GSH also occurred in the absence of oxyhaemoglobin. The results suggest a dual mechanism for these oxidative effects, involving autoxidation of the 5-hydroxy-8-aminoquinolines and their coupled oxidation with oxyhaemoglobin. The initial products of these processes would be drug metabolite free radicals, superoxide radical anions, hydrogen peroxide and methaemoglobin. Further free radical reactions would lead to oxidation of GSH, more haemoglobin and probably other cellular constituents. NADPH had no effect on the oxidative effects of the primaquine metabolites in these experiments. In the G6PD-deficient erythrocyte, the oxidation of haemoglobin and GSH leads to Heinz body formation and eventually to haemolysis, the mechanisms of which are as yet unclear. The possible role of oxygen free radicals in the mode of action of 8-aminoquinolines against the malaria parasite is also briefly discussed.     

Cyclosporine A-induced changes to erythrocyte redox balance is time course-dependent

Cyclosporine A-treated transplant recipients develop pronounced cardiovascular disease and have increased oxidative stress and altered antioxidant capacity in erythrocytes and plasma. These experiments investigated the time-course of cyclosporine A-induced changes to redox balance in plasma and erythrocytes. Rats were randomly assigned to either a control or cyclosporine A-treated group. Treatment animals received 25 mg/kg of cyclosporine A via intraperitoneal injection for either 7 days or a single dose. Control rats were injected with the same volume of the vehicle. Three hours after the final injections, plasma was analysed for total antioxidant status, alpha-tocopherol, malondialdehyde, and creatinine. Erythrocytes were analysed for reduced glutathione (GSH), alpha-tocopherol, methaemoglobin, malondialdehyde, and the activities of superoxide dismutase, catalase, GSH peroxidase, and glucose-6-phosphate dehydrogenase (G6PD). Cyclosporine A administration for 7 days resulted in a significant increase (P<0.05) in plasma malondialdehyde, methaemoglobin, and superoxide dismutase and catalase activities. There was a significant decrease (P<0.05) in erythrocyte GSH concentration and G6PD activity in cyclosporine A animals. There were no significant differences (P>0.05) between groups following a single dose of cyclosporine A in any of the measures. In summary, cyclosporine A alters erythrocyte redox balance after 7 days administration, but not after a single dose.     

Preventive effects of Laminaria japonica aqueous extract on the oxidative stress and xanthine oxidase activity in streptozotocin-induced diabetic rat liver

Increasing evidence in both experimental and clinical studies suggests that oxidative stress plays a major role in the pathogenesis of diabetes mellitus type 1 and type 2. Xanthine oxidase (XO) has been proposed as one of the sources of free radical formation in diabetes. We therefore investigated the preventive effects of Laminaria japonica aqueous extract (LJE) on alterations in the activity of hepatic XO and oxidative stress in the streptozotocin-induced experimental diabetes. We found that lipid peroxide levels and xanthine oxidase activity were increased, whereas glutathione (GSH), GSH reductase and GSH peroxidase were decreased in the liver of streptozotocin-induced diabetic rats. Pretreatment with LJE of 100 mg/kg orally for 5 d significantly reduced blood glucose levels and hepatic lipid peroxidation in the diabetic rats. In addition, the content of glutathione was restored to the control level by LJE pretreatment. Furthermore, LJE significantly suppressed the increased activity of XO and type conversion of the xanthine dehydrogenase to XO in diabetic rat liver. The results suggest that Laminaria japonica would be of great value in preventing hyperglycemia in diabetes mellitus as a dietary supplement possibly, through its antioxidant activity.     

Protective effect of N-acetylcysteine on reduced glutathione, reduced glutathione-related enzymes and lipid peroxidation in methanol intoxication

The primary metabolic appropriation of methanol is oxidation to formaldehyde and then to formate. These processes are accompanied by formation of superoxide anion and hydrogen peroxide. This paper reports data on the effect of N-acetylcysteine (NAC) on reduced glutathione (GSH) and on activity of some GSH-metabolising enzymes in the liver, erythrocytes and serum of rats intoxicated with methanol (3 g/kg b.w.) during 7 days after intoxication. Methanol administration, increasing concentration of the lipid peroxidation products, decreased the liver glutathione-peroxidase and glutathione reductase (GSSG-R) activities, GSH concentration and total antioxidant status (TAS). The use of NAC after methanol ingestion apparently diminished lipid peroxidation, elevated the GSH level in the liver and erythrocytes, and increased activity of GSH-related enzymes in the serum, erythrocytes and in the liver. These results suggest that NAC exerts its protective effect by acting as a precursor for glutathione, the main low molecular antioxidant and as a free radical scavenger.     

糖尿病大鼠肾脏抗氧化防御系统机能的改变

目的：探讨糖尿病对肾脏抗氧化防御机能的影响。方法：观察12周糖尿病大鼠肾皮质丙二醛(MDA)及谷胱甘肽(GSH)水平,以及超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-PX)、谷胱甘肽S转移酶(GSH-ST)和过氧化氢酶(CAT)活性的变化。结果：糖尿病大鼠肾组织中SOD、CAT活性下降;GSH含量显著降低;MDA没有变化;GSH-PX活性却明显增强。结论：糖尿病大鼠肾组织抗氧化防御机能明显下降。     

Resistance to glutathione depletion in diabetic and non-diabetic human erythrocytes in-vitro

We have investigated the resistance of erythrocytes from diabetics and non-diabetics to glutathione depletion caused by p-benzoquinone, 1-chloro-2,4-dinitrobenzene (CDNB), diethyl maleate and 4-aminophenol. Incubation of erythrocytes with 4-aminophenol (2 mM) caused a precipitous reduction (>80%) in cellular glutathione levels although there was no significant difference between 4-aminophenol-mediated glutathione depletion in the diabetic and non-diabetic cells. p-Benzoquinone and CDNB were both associated with a less severe initial reduction in glutathione levels (>50% at 30 min) although p-benzoquinone caused greater depletion (P < 0.001) at 4.5 h (21.1 +/- 3.1%, non-diabetic; 20.0 +/- 1.0%, diabetic) compared with CDNB (49.2 +/- 2.2%, non-diabetic; 51.3 +/- 1.1% diabetic). Although there was no significant difference between the two types of cell in terms of level of depletion, administration of diethyl maleate caused a significant reduction in glutathione levels at 30 min (P < 0.0005), 3.5 h (P < 0.05) and 4.5 h (P < 0.05) in erythrocytes from diabetic man compared with those from non-diabetic man. Co-administration of buthionine sulphoximine (20 mM) and 4-aminophenol (1 mM) also led to a significant reduction in glutathione levels in diabetic cells at 30 min (P < 0.05), 3.5 h (P < 0.02) and 4.5 h (P < 0.007) compared with those in non-diabetic cells. The observations that diabetic red cells' resistance to depletion was similar to that of nondiabetic cells for three of the four depletors, and that the combination of 4-aminophenol and buthionine sulphoximine-mediated inhibition of glutathione synthesis was required to illustrate differences suggests that diabetic complications might be a result of the long-term effect of small deficiencies in oxidative self-defence mechanisms such as glutathione.     

Restoration of antioxidants by ethanolic Tinospora cordifolia in alloxan-induced diabetic Wistar rats

The present study investigates the effect of oral administration of an alcoholic extract of Tinospora cordifolia roots on antioxidant defence in alloxan-induced diabetes in rats. A significant increase in the concentration of thiobarbituric acid reactive substances (TBARS) in brain along with a decrease in heart was observed in diabetic rats. Decreased concentration of glutathione (GSH) and decreased activities of superoxide dismutase (SOD), catalase and glutathione peroxidase (GPx) in heart and brain of diabetic rats were also noted. Alcoholic Tinospora cordifolia root extract (TCREt) administered at a dose of 100 mg/kg to diabetic rats orally for six weeks normalized the antioxidant status of heart and brain. The effect of T. cordifolia root extract was more prominent than glibenclamide (600 microg/kg). Insulin (6 units/kg) restored all the parameters to normal status.     

A study of hypoglycemic and antioxidant activity of Aegle marmelos in alloxan induced diabetic rats

The present study was performed to evaluate the hypoglycemic and antioxidant effect of aqueous extract of Aegle marmelos leaves (AML) on diabetic rats. Male albino rats were randomly divided into three groups: Group I: Control; Group II: Diabetic rats; and Group III: Diabetic rats administered AML. Glucose, urea and glutathione-S-transferase (GST) in plasma, glutathione (GSH) and malondialdehyde (MDA) levels in erythrocytes were estimated in all the groups at the end of four weeks. There was a decrease in blood glucose at the end of four weeks in group III animals compared with group II, however it did not reach the control levels. There was an increase in erythrocyte GSH and a decrease in MDA in group III as compared to group II. The plasma GST levels were raised in diabetic rats when compared to controls. In the group III animals, there was a decrease in GST as compared to group II. Owing to hypoglycemic and antioxidant properties, AML may be useful in the long-term management of diabetes.     

Effects of lipoic acid and dihydrolipoic acid on 4-aminophenol-mediated erythrocytic toxicity in vitro

The effects of the antioxidant lipoic acid and its reduced form, dihydrolipoic acid (DHLA), were studied on the process of the erythrocytic toxicity of 4-aminophenol in human erythrocytes in vitro. 4-Aminophenol alone caused a stepwise increase in methaemoglobin formation, along with a commensurate decrease in total thiols. At 10 min., in the presence of lipoic acid alone and the thiol depletor 1-chloro-2,4-dinitrobenzene (CDNB) alone, 4-aminophenol-mediated methaemoglobin formation was significantly increased, whilst thiol levels were significantly reduced compared with the 4-aminophenol alone. At 10 min., with DHLA and CDNB alone, 4-aminophenol was associated with significantly increased methaemoglobin formation. However, thiol levels were not significantly different in the presence of DHLA compared with 4-aminophenol alone, although thiol levels were different compared with control (4-aminophenol alone) in the incubations with CDNB alone. At 15 min., only CDNB/4-aminophenol methaemoglobin formation differed from control, whilst thiol levels were significantly lower in the presence of CDNB alone compared with 4-aminophenol alone. Lipoic acid enhanced the toxicity of 4-aminophenol in terms of increased methaemoglobin formation coupled with increased thiol depletion, whilst DHLA showed increased 4-aminophenol-mediated methaemoglobin formation without thiol depletion. Lipoic acid, and to a lesser extent its reduced derivative DHLA, acted as a prooxidant in the presence of 4-aminophenol, enhancing the oxidative stress effects of the amine in human erythrocytes.     

The role of N-acetylcysteine treatment on anti-oxidative status in patients with type II diabetes mellitus

The development of diabetic complications has usually been attributed to the nonenzymic glycation of tissue proteins. Only recently, however, have researchers examined the possible role on free radicals in the pathogenesis of diabetes. In the present study, glutathione (GSH) and major antioxidant enzyme levels in plasma of patients with type II diabetes mellitus were assessed both before and after 3 months of N-acetylcysteine (NAC) therapy. Thirty-two diabetic patients were examined as well as fifteen healthy controls. Before treatment with NAC, glutathione peroxidase (GPx), catalase (CAT), and (GSH) levels of diabetic patients and control subjects showed no significant differences, whereas glutathione S-transferase (GST) levels were higher in type II diabetic patients. Following 3 months of Following NAC supplementation, GSH, GST, and CAT levels were found to be similar to the levels before treatment. On the other hand, GPx activity was significantly lower compared with the values before treatment. According to this finding, NAC treatment could have a positive effect on GPx values in type II diabetic patients showing abnormally high values.     

Centella asiatica enhances hepatic antioxidant status and regulates hepatic inflammatory cytokines in type 2 diabetic rats

Context: Neutralizing the over-activation of oxidative stress and inflammation remains an important goal in the management of type 2 diabetes mellitus (T2DM). Centella asiatica (L.) Urban (Apiaceae) (CA) has been used in traditional folklore in Africa and Asia to treat various ailments including diabetes.

Objective: We investigated the hepatic antioxidant and anti-inflammatory potential of methanol extract of CA leaves in T2DM.

Materials and methods: T2DM was induced in male Sprague-Dawley rats with 10% fructose in drinking water for 14?days followed by a single intraperitoneal injection of streptozotocin (40?mg/kg b.wt). Hepatic oxidant/antioxidant status was assessed by measuring the concentrations of malondialdehyde (MDA), ferric reducing antioxidant power (FRAP), oxygen radical absorbance capacity (ORAC), Trolox equivalent antioxidant capacity (TEAC), reduced glutathione (GSH) and activities of glutathione S-transferase (GST) and glutathione peroxidase (GPX). The concentrations of cytokines IL-1β, IL-4, IL-6, IL-10, MCP-1 and TNF-α in the liver were determined.

Results: Diabetes increased MDA formed (47%) and reduced FRAP (20%), TEAC (15%), GSH levels (32%), significantly; decreased GST and GPX activities in the liver and elevated levels of cytokines studied. Treatment of diabetic rats with 500?mg/kg b.wt CA for 14?days decreased MDA (44%); elevated FRAP (15%) and GSH (131%) levels and increased the activities of GST and GPX by 16%. Hepatic concentrations of IL-1β, MCP-1 and TNF-α in DCA group were reduced to 68%, 75% and 63% of DC values, respectively.

Conclusions: The antioxidant and anti-inflammatory properties of CA may protect tissues such as the liver from diabetes-induced oxidative damage.     

Effects of the antioxidants dihydrolipoic acid (DHLA) and probucol on xenobiotic-mediated methaemoglobin formation in diabetic and non-diabetic human erythrocytes in vitro(1)

The antioxidant effects of dihydrolipoic acid (DHLA) and probucol were investigated in a human erythrocytic in-vitro model of diabetic oxidative stress, where xenobiotics were used to form methaemoglobin. 4-Aminophenol mediated haemoglobin oxidation in non-diabetic erythrocytes was not affected by the presence of either DHLA or probucol. However, with diabetic cells, there were significant increases (P<0.01) in 4-aminophenol-mediated haemoglobin oxidation in the presence of DHLA. Methaemoglobin formed by nitrite in non-diabetic and diabetic cells was not altered by either DHLA or probucol except at one time point in diabetic cells. In non-diabetic as well as diabetic cells, methaemoglobin formed by MADDS-NHOH was significantly reduced at all three time points in the presence of DHLA (P<0.0001) but unaffected by probucol. In the presence of DHLA only, methaemoglobin formed by the products of rat microsomal oxidation of both 4-aminopropiophenone and benzocaine was markedly reduced for both xenobiotics in diabetic and non-diabetic cells (P<0.0001) compared with cells incubated in the absence of DHLA. There were no significant differences between total cellular thiol levels determined between diabetic and non-diabetic erythrocytes, nor did DHLA or probucol affect resting thiol levels. MADDS-NHOH caused a significant thiol depletion in diabetic cells, which was restored in the presence of DHLA. A further study is required to determine how DHLA attenuates the potent REDOX reactions that occur during hydroxylamine-mediated methaemoglobin formation.     

Effects of trichlorfon on malondialdehyde and antioxidant system in human erythrocytes

Organophosphorus insecticides may induce oxidative stress leading to generation of free radicals and alteration in antioxidant system. The aim of this study was to examine the potency of trichlorfon, an organophosphate insecticide, to induce oxidative stress response in human erythrocytes in vitro. For this purpose trichlorfon solutions in different concentrations and erythrocyte solutions were incubated at 37 °C for 60 min. At the end of the incubation time, malondialdehyde (MDA), an end product of lipid peroxidation, total glutathione, reduced glutathione (GSH) levels, activities of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSH-Px) enzymes were determined by spectrophotometric methods. Trichlorfon increased MDA formation depended on the concentration. On the other hand, decreases in the GSH-Px activity, GSH levels and increases in the total glutathione levels, SOD and CAT activities were seen in the studied concentrations. The present findings indicate that the in vitro toxicity of trichlorfon may be associated with oxidative stress.