Inhibition of membrane Na(+)-K+ Atpase of the brain, liver and RBC in rats administered di(2-ethyl hexyl) phthalate (DEHP) a plasticizer used in polyvinyl chloride (PVC) blood storage bags.

Significant amounts of di(2-ethylhexyl) phthalate (DEHP) leach out into blood stored in DEHP plasticized polyvinyl chloride (PVC) bags resulting in the exposure of recipients of blood transfusion to this compound. The aim of this study was to find out whether DEHP at these low levels has any effect on the activity of membrane Na(+)-K+ ATPase, since a decrease in this enzyme activity has been reported to take place in a number of disorders like neurodegenerative and psychiatric disorders, coronary artery disease and stroke, syndrome-X, tumours etc. DEHP was administered (ip) at a low dose of 750 microg/100 g body weight to rats and the activity of membrane Na(+)-K+ ATPase in liver, brain and RBC was estimated. Histopathology of brain, activity of HMG CoA reductase (a major rate limiting enzyme in the isoprenoid pathway of which digoxin, the physiological inhibitor of Na(+)-K+ ATPase is a product), intracellular concentration of Ca2+ and Mg2+ in RBC (which is altered as a result of inhibition of Na(+)-K+ ATPase) were also studied. (In the light of the observation of increase of intracellular Ca2+ load and intracellular depletion of Mg2+ when Na(+)-K+ ATPase is inhibited). Histopathology of brain revealed areas of degeneration in the rats administered DEHP. There was significant inhibition of membrane Na(+)-K+ ATPase in brain, liver and RBC. Intracellular Ca2+ increased in the RBC while intracellular Mg2+ decreased. However activity of hepatic HMG CoA reductase decreased. Activity of Na(+)-K+ ATPase and HMG CoA reductase, however returned to normal levels within 7 days of stopping administration of DEHP. The inhibition of membrane Na(+)-K+ ATPase activity by DEHP may indicate the possibility of predisposing recipients of transfusion of blood or hemodialysis to the various disorders mentioned above. However since this effect is reversed when DEHP administration is stopped, it may not be a serious problem in the case of a few transfusion; but in patients receiving repeated blood transfusion as in thalassemia patients or patients undergoing hemodialysis, possibility of this risk has to be considered. This inhibition is a direct effect of DEHP or its metabolites, since activity of HMG CoA reductase, (an enzyme which catalyses a major rate limiting step in the isoprenoid pathway by which digoxin, the physiological inhibitor of Na(+)-K+ ATPase is synthesized) showed a decrease.

Tryptophan and tyrosine catabolic pattern in neuropsychiatric disorders.

Catabolism of tryptophan and tyrosine in relation to the isoprenoid pathway was studied in neurological and psychiatric disorders. The concentration of trytophan, quinolinic acid, kynurenic acid, serotonin and 5-hydroxyindoleacetic acid was found to be higher in the plasma of patients with all these disorders; while that of tyrosine, dopamine, epinephrine and norepinephrine was lower. There was increase in free fatty acids and decrease in albumin (factors modulating tryptophan transport) in the plasma of these patients. Concentration of digoxin, a modulator of amino acid transport, and the activity of HMG CoA reductase, which synthesizes digoxin, were higher in these patients; while RBC membrane Na+-K+ ATPase activity showed a decrease. Concentration of plasma ubiquinone (part of which is synthesised from tyrosine) and magnesium was also lower in these patients. No morphine could be detected in the plasma of these patients except in MS. On the other hand, strychnine and nicotine were detectable. These results indicate hypercatabolism of tryptophan and hypocatabolism of tyrosine in these disorders, which could be a consequence of the modulating effect of hypothalamic digoxin on amino acid transport.

Changes in the composition of erythrocyte membrane during storage of blood in di-(2-ethyl hexyl) phthalate [DEHP] plasticized poly vinyl chloride (PVC) blood storage bags.

Very little information is available on the changes in the erythrocyte membrane composition during storage of blood at 4 degrees C, particularly with respect to the glycosaminoglycans and glycoproteins. In view of this, a detailed study was carried out on the changes in the membrane proteins, glycosaminoglycans (GAG), carbohydrate components of glycoproteins, cholesterol, phospholipids and vitamin E in blood stored in glass bottles and a di-(2-ethyl hexyl) phthalate (DEHP) plasticized PVC bag (Penpol blood bag). Blood was collected in CPDA solution in glass bottles and in Penpol blood bags and kept at 4 +/- 1 degrees C. Analysis was made immediately after blood collection and after 28 and 42 days. Significant increase in the total protein in the erythrocyte membrane was observed during storage of whole blood in glass bottles and Penpol blood bag at 4 degrees C. This increase was progressively more with increase in storage time. Significant changes were also observed in GAG, carbohydrate components of glycoproteins, cholesterol, phospholipids and vitamin E in the erythrocyte membrane under these conditions. The protein:GAG ratio, protein:carbohydrate ratio, cholesterol:phospholipid ratio as well as protein:lipid ratio showed significant increase in the membrane. The extent of these changes was lower in the Penpol bag, indicating the stabilizing effect of DEHP on the erythrocyte membrane.

Synthesis of NAD+ in erythrocytes incubated with nicotinic acid and the effect of di-(2-ethyl hexyl) phthalate (DEHP).

Synthesis of NAD+ from nicotinic acid by erythrocytes incubated in SAGM phosphate solution and effect of di-[2-ethyl hexyl] phthalate, a plasticizer commonly used in PVC blood/component storage bags, on this synthesis was studied. Erythrocytes are able to synthesise NAD+ in SAGM (sodium chloride, adenine, glucose, mannitol) phosphate solution and this synthesis was more in the presence of added nicotinic acid (optimum concentration 1 mM). The level of NAD+ decreased when the incubation period was increased from 24 to 48 hr. Glutamine had a deleterious effect on this synthesis, possibly due to the decrease in pH. Di-[2-ethyl hexyl] phthalate had an inhibitory effect on NAD+ synthesis when the cells were incubated in SAGM phosphate solution, either alone or in the presence of added nicotinic acid. There was significant decrease in the release of potassium and haemoglobin from the cells in the presence of nicotinic acid, indicating increased red cell stability.

Effect of DEHP [di-(2-ethyl hexyl) phthalate] on lipid peroxidation in liver in rats and in primary cultures of rat hepatocytes.

The effect of DEHP [di-(2-ethly hexyl) phthalate] on lipid peroxidation in the liver in rats and in primary cultures of rat hepatocytes incubated with it was studied. The doses of DEHP used in this study corresponded to the low levels of this substance leaching into blood stored in DEHP plasticised PVC bags. Increased activity of superoxide dismutase (SOD) and catalase, increased concentration of malondialdehyde (MDA) and conjugated dienes and decrease in the concentration of glutathione and vitamin E have been observed in the liver of rats administered DEHP. Primary cultures of rat hepatocytes incubated with DEHP also showed increase in the activity of these enzymes, increase in the concentration of MDA and decrease in vitamin E. These results indicate that DEHP promotes lipid peroxidation. Incorporation of vitamin E along with DEHP into the culture medium containing hepatocytes counteracted these effects.

Toxic effect of systemic administration of low doses of the plasticizer di-(2-ethyl hexyl) phthalate [DEHP] in rats.

DEHP [di-(2 ethyl hexyl) phthalate], a widely used plasticizer in blood storage bags, leaches out in appreciable amounts into blood (about 10 mg/100 ml) resulting in exposure of recipients of blood transfusion to this compound. Various reports indicate the toxicity of DEHP, particularly in liver and reproductive organs but all these studies used large doses (up to 2 g or more/Kg body weight) and oral route of administration which are not relevant to the intravenous administration during blood transfusion or the low amounts present in blood. We have studied changes in the activity of some important enzymes-gamma-GT, ALT, CPK, LDH, alkaline phosphatase, acid phosphatase, beta-glucuronidase and few other parameters like vitamin E, glutathione, serum albumin etc in rats administered low doses of DEHP (corresponding to transfusion of 2, 4, 6 and 10 units of blood). Histopathology of the organs has also been carried out. The results obtained indicate no serious toxic effects for DEHP at the level present in blood stored in DEHP plasticized blood bags as evidenced by the lack of any significant alteration in most of the biochemical parameters studied. Even in those cases where there was alteration (for e.g., decrease in the level of vitamin E) 24 hr after administration of DEHP, it returned to near normal level with in 72 hr to 7 days. No histopathological changes were observed in any of the organs at these levels of DEHP. It is concluded that DEHP did not cause any serious toxic effect even at doses corresponding to transfusion of several units of blood in a recipient.