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The pathogenetic sequence of reactions mediated by endotoxin (LPS) leading to the production of sepsis involves the oxygen radicals or reactive oxygen species, which has been evaluated in the present review. Among reactive oxygen species hydroxyl radical either singly or in combination with peroxynitrite, produces tissue damage often observed during septic injury. Inactivation of these damaging radicals by antioxidants or nitric oxide inhibitor(s) may be helpful for protecting sepsis mediated derangements but the application of these agents as drugs in humans has not been fully successful. Transcription factor NF-kappa B is reported to be the oxygen sensor in LPS induced endotoxemia. Polyphenols, especially the catechin group of compounds, are important therapeutic agents, which may be used for the treatment of endotoxin mediated sepsis.

administration of isoproterenol to mice at a dose of 30 mg/100 g body weight for 3 consecutive days at an interval of 24 h induced lipid peroxidation in cardiac tissue and exhibited a significantly elevated serum glutamate oxaloacetate transaminase (SGOT) level. Increased superoxide dismutase (SOD) activity with a concomitant decrease in catalase activity has also been observed in cardiac tissue with isoproterenol treatment. Quinidine, a class I antiarrhythmic agent has been found to exhibit a protective role in isoproterenol induced myocardial ischaemia. Cardiac tissue of quinidine treated mice showed reduction of lipid peroxidation reaction. In addition, quinidine treatment is found to influence the cardiac antioxidant enzymes – catalase and SOD. The decrease of SOD activity and increase of catalase activity suggests that quinidine also exerts an ‘indirect antioxidant’ effect in protecting the myocardial tissue from reactive oxygen species. Furthermore, our current in vitro studies with quinidine have clearly shown in this work that it possesses a very convincing hydroxyl radical scavenging potential with almost no ability to scavenge superoxide anion and hydrogen peroxide (H2O2) in vitro. Thus, our present investigation suggests that quinidine, when administered to mice, strengthens the antioxidant defense system to resist the free radical induced damage brought about by isoproterenol induced ischaemic condition.

The effect of cadmium (Cd2+), mercury (Hg2+) and copper (Cu2+) was studied with partially purified flavokinase (ATP:riboflavin 5′-phosphotransferase EC 2.7.1.26) from rat liver. All the divalent heavy metal cations inhibited flavokinase activity in a concentration-dependent manner. The inhibitory effect of cadmium on the enzyme was completely reversed by increasing concentration, of Zinc (Zn2+) indicating a competition between Zn2+ and Cd2+ for binding with the enzyme. A competition between riboflavin and Cd2+ is also evident from the present investigation. These observations hint at the possibility that Zn2+ and Cd2+ probably compete for the same site on the enzyme where riboflavin binds. However, inhibition of flavokinase by Hg2+ could not be reversed by Zn2+. Our studies further reveal that hepatic flavokinase appears to contain an essential, accessible and functional thiol group(s) which is evident from a concentration dependent inhibition of activity by sulfhydryl reagent s like parachloromercuribenzoate (PCMB), 5,5′-dithiobis (2-nitrobenzoic acid)(DTNB), and N-ethylmaleimide (NEM). Inhibition of flavokinase by sulfhydryl reagents were protected, except in case of NEM inhibition, when the enzyme was incubated with thiol protectors like glutathione (GSH) and dithiothreitol (DTT). Furthermore, the enzyme could also be protected from the inhibitory effect of Cd2+ and Hg2+ by GSH and DTT suggesting that Cd2+ probably interacts with a reactive thiol group at or near the active site of enzyme in bringing about its inhibitory effect. (Mol Cell Biochem 167: 73-80, 1997)

The effect of cadmium on levels of hepatic riboflavin, flavin mononucleotide (FMN), flavin adenine dinucleotide (FAD), as well as the activities of flavokinase and FMN-phosphatase were studied following injection of cadmium sulfate solution subcutaneously to male. Sprague-Dawley rats at a dose of 0.44 mg/kg body weight, every alternate day for 15 days. The body weights and wet weights of liver were found to increase significantly following cadmium treatment ( P P P P P > 0.05, respectively). A significant decrease in activities of hepatic flavokinase and FMN-phosphatase were observed both in in vivo and in vitro experiments, although the degree of inhibition was found to be more pronounced in case of flavokinase than FMN-phosphase ( P P

Glucagon increased the activities of alanine amino transferase (AAT), fructose-1:6-bisphosphatase (fru-P2ase) and glucose-6-phosphatase (G-6-Pase) in goat brain tissue by about 100%, 150% and 50% respectively. These increase in activities were reversed by beta-antagonists propranolol. Well known alpha-agonist and antagonist like phenylephrine and phenoxybenzamine also increased AAT and G-6-Pase activities and these increased activities were reversed by propranolol. Phenylephrine and phenoxybenzamine however did not increase brain Fru-P2ase activity. However the most interesting finding is that cerebral cortical slices could produce glucose from alanine and this glucose production was enhanced by glucagon, phenylephrine and phenoxybenzamine. Propranolol reversed the effects of these agonists and antagonist to a great extent. From all these experiments we suggest brain to be a gluconeogenic organ although much less efficient than liver.

The effect of erythropoietin (Ep), a glycoprotein hormone, has been studied on lipid peroxidation induced by Cu2+ and ascorbate in vitro, Mg2+ ATPase activity and spectrin of RBC membrane. Our present investigation reveals that Cu2+ and ascorbic acid increases lipid peroxidation of RBC membrane significantly. It has further been observed that under the same experimental condition spectrin, a major cytoskeleton membrane protein, and Mg(2+)-ATPase activity of RBC membrane decrease significantly. However, exogenous administration of Ep completely restores lipid peroxidation and Mg(2+)-ATPase activity and partially recovers spectrin of RBC membrane.

The oxidative stress status of the transfusion-dependent Ebeta- and beta-thalassemia patients were studied before and after treatment with vitamin E for a period of four weeks. The level of cellular vitamin antioxidants viz. ascorbic acid and vitamin E in the thalassemia patients were found to be considerably lower compared to normal subjects. The activities of enzymatic antioxidants viz. catalase, glutathione peroxidase and glutathione reductase were found to be drastically reduced in untreated Ebeta- and beta-thalassemic patients when compared to normal subjects. However, the activity of superoxide dis-mutase was found to be increased in both types of untreated thalassemic patients when compared to normal individuals. An increase in superoxide dismutase and a decrease in catalase activity reflects the presence of a severe oxidative stress situation in the erythrocytes of the untreated transfusion dependent Ebeta- and beta-thalassemia patients. Changes in erythrocyte membrane protein pattern in untreated Ebeta- and beta-thalassemia patients when compared to normal erythrocyte further confirm the presence of continued oxidative stress in the ailing thalassemic erythrocytes. All these changes in the antioxidant status as well as the changes in the erythrocyte membrane proteins are ameliorated to considerable extent when the transfusion-dependent Ebeta- and beta-thalassemia patients were treated with vitamin E at a dose of 10 mg/kg/day for a period of four weeks. The patients during the treatment period did not exhibit any side effects and gained in body weight indicating a healthy status. The present study reveals that the lipophilic antioxidant vitamin E could be useful in the management of transfusion-dependant Ebeta- and beta-thalassemia patients.

Administration of isoproterenol to mice at a dose of 30 mg/100 g body weight for 3 consecutive days at an interval of 24 h induced lipid peroxidation in cardiac tissue and exhibited a significantly elevated serum glutamate oxaloacetate transaminase (SGOT) level. Increased superoxide dismutase (SOD) activity with a concomitant decrease in catalase activity has also been observed in cardiac tissue with isoproterenol treatment. Quinidine, a class I antiarrhythmic agent has been found to exhibit a protective role in isoproterenol induced myocardial ischaemia. Cardiac tissue of quinidine treated mice showed reduction of lipid peroxidation reaction. In addition, quinidine treatment is found to influence the cardiac antioxidant enzymes - catalase and SOD. The decrease of SOD activity and increase of catalase activity suggests that quinidine also exerts an 'indirect antioxidant' effect in protecting the myocardial tissue from reactive oxygen species. Furthermore, our current in vitro studies with quinidine have clearly shown in this work that it possesses a very convincing hydroxyl radical scavenging potential with almost no ability to scavenge superoxide anion and hydrogen peroxide (H2O2) in vitro. Thus, our present investigation suggests that quinidine, when administered to mice, strengthens the antioxidant defense system to resist the free radical induced damage brought about by isoproterenol induced ischaemic condition.

Treatment of red blood cells with the copper (II) ascorbate system causes increased lipid peroxidation, increased membrane microviscosity, and phospholipid translocation with a concurrent decrease in cytosolic catalase and glutathione peroxidase activities. All these changes are prevented if the cells are treated with erythropoietin prior to the exposure to copper (II) ascorbate. The present investigation further indicates that the oxidative damage brought about by copper (II) ascorbate is due to generation of hydroxyl radical and that erythropoietin plays a unique role in protecting the membrane from oxidative damage.

The effect of cadmium (Cd2+) was studied in vitro on the flavokinase (ATP: riboflavin 5′-phosphotransferase, EC 2.7.1.26) activity purified from rat liver. Cadmium inhibited flavokinase activity in a concentration-dependent manner and the effect was completely reversed by increasing concentration of zinc (Zn2+), indicating a competition between Zn2+ and Cd2+ for binding with the enzyme. Further, a competition between riboflavin and Cd2+ hints at the possibility that Zn2+ and Cd2+ probably compete for the same site on the enzyme where riboflavin binds. Our studies further reveal that hepatic flavokinase contains essential, accessible and functional thiol group(s) as evidenced by a concentration-dependent inhibition by sulfhydryl reagents and protection by thiol protectors like glutathione or dithiothreitol. Furthermore, the enzyme could also be protected from the inhibitory effect of Cd2+ and Hg2+ by glutathione and dithiothreitol suggesting that Cd2+ probably interacts with reactive thiol group at or near the active site of the enzyme to cause inhibition.

Using rat hepatocytes we confirmed our previous results that glucagon and beta-adrenergic agonists increased the enzyme activity of alanine aminotransferase (AAT) and propranolol abolished their effects. Only the enzyme activity was measured and other parameters like quantity of the enzyme or activation due to modification were not looked for. As in perfusion experiment phenylephrine and phenoxybenzamine (alpha-agonist and alpha-antagonist respectively) also alpha-antagonist respectively) also increased the AAT activity in isolated rat hepatocytes and propranolol reversed these effects. The additive effect of glucagon and phenoxybenzamine on AAT was also persistent in hepatocyte system. Fructose-1:6-bisphosphatase (Fru-P2-ase), another key enzyme in gluconeogenic pathway, was elevated by glucagon and other beta-adrenergic agonists both in liver perfusion and isolated hepatocyte experiments and was brought back to the normal level by propranolol. In this case also only the enzyme activity was measured and no other parameters were looked for. Unlike AAT this enzyme was not stimulated by phenylephrine or phenoxybenzamine. But AAT and Fru-P2-ase activities were increased significantly by adenylate cyclase activators like fluoride or forskolin. Thus, it appears that the regulation of fru-P2-ase by glucagon is purely a b-receptor mediated process whereas AAT activation shows a mixed type of regulation where some well known alpha-agonist and antagonists are behaving as beta-agonists. Results further indicate the presence of phosphodiesterase in hepatocyte membrane which was stimulated by glucagon and brought back to the normal level by propranolol. The different adrenergic compounds stated above, not only modified the activity of the above two enzymes but also stimulated glucose production by hepatocytes from alanine which was in turn abolished by propranolol as well as amino oxyacetate (AOA), a highly specified inhibitor of AAT. This confirm the participation of AAT in gluconeogenesis from alanine in liver. Forskolin and fluoride also increased the glucose production from alanine and showed additive effects with glucagon, phenylephrine and phenoxybenzamine.

Glucagon increased alanine amino transferase (AAT) activity in perfused rat liver by about 90% over control. Propranolol, the beta receptor antagonist, abolished the effect of glucagon on this enzyme. Well known beta receptor agonists like isoproterenol, norepinephrine and epinephrine also increased the enzyme activity under identical condition and the enhancement was similarly abolished by propranolol. These experiments suggest that the effect of glucagon on AAT was mediated through beta adrenergic receptor. However, the interesting observation was that phenylephrine, alpha receptor agonist and phenoxybenzamine and tolazoline, two alpha receptor antagonists, increased the AAT activity like glucagon in perfusion experiments and the effects of all these three agents were also abolished by propranolol. Glucagon, when perfused with phenoxybenzamine showed some additive effect. From all these results we are proposing that in our system phenoxybenzamine is acting as beta agonist although it is known to be an alpha antagonist.

The effect of Ep on different ATPases and acetylcholinesterase of rat RBC membrane was studied. Starvation caused a slight decrease in Mg2+-, Ca2+-, and Na+ + K+-ATPases. However, these enzyme activities were markedly increased on Ep treatment of starved rats. Specific activities of all three ATPases increased linearly with increasing concentration of Ep. Under identical conditions the hormone failed to stimulate the ATPase activity of liver plasma membrane. Desensitization by fluoride of allosteric inhibition of erythrocyte membrane-bound Na+ + K+-ATPase was observed under starvation which showed a return to normal n values on Ep administration. The enzyme from normal animals was inhibited almost completely at 0.1 mm fluoride whereas enzyme from starved and Ep-treated animals showed only about 50% inhibition at that fluoride concentration. Ep increased the acetylcholinesterase activity of normal RBC membrane to a small extent whereas the stimulation was much higher under starvation. The fluoride inhibition curve of this enzyme changed from sigmoidal to hyperbolic under starvation which again changed to allosteric on administration of Ep. These changes were closely correlated to n values. Red blood cells of Ep-treated animals became more susceptible to osmotic shock under the experimental conditions.

Effect of Ep on [14C]acetate incorporation into different lipid fractions of RBC membranes in starved and phenylhydrazine-treated rats was studied. The incorporation was increased into both neutral and phospholipid fractions on Ep treatment to starved or phenylhydrazine-treated rats. A slight decrease in the ratio of neutral lipid to phospholipid was observed under the influence of Ep in starved rats (23%) or in phenylhydrazine-treated rats (36%). Incorporation of radioactivities into different phospholipid fractions of RBC membrane increased on Ep treatment to starved rats, whereas, the relative percentages of these phospholipids (except LPC) remained more or less unchanged under similar conditions. Phenylhydrazine treatment increased the relative percentage of PC and concomitantly decreased the percentage of Sph. Percentage composition of both these two phospholipids showed a tendency to return to their normal levels on administration of Ep to phenylhydrazine-treated rats. Ep decreased the sigma saturated/sigma unsaturated ratio of fatty acids in PE, PS, and PC of RBC membrane in starved rats. On the other hand, no significant change was observed in this ratio of fatty acids in the phospholipids except Sph of RBC membrane in the presence of phenylhydrazine and Ep. In Sph, the ratio went down under similar conditions.

EDTA (4 mM) blocks the oxidation of iodide to I-3 (increase of extinction at 353 nm) by H2O2 catalyzed by horseradish peroxidase, which is reversed by the addition of an equimolar concentration of Zn2+. Addition of suboptimal concentration of EDTA (2 mM) not only decreases the rate of forward reaction of I-3 formation but also causes loss of extinction of the same when I-3 is generated. The loss of extinction of I-3 is proportional to the enzyme concentration and is blocked by azide, the inhibitor of the peroxidase. EDTA also causes bleaching of nonenzymatically formed I-3 (from iodide and H2O2) only in the presence of horseradish peroxidase, and the effect is reversed by the equimolar concentration of Zn2+. Both the bleaching of I-3 by EDTA and reversal of EDTA effect by Zn2+ are sensitive to azide. The decrease of extinction of I-3 (formed by dissolving iodine in KI solution) is dependent on EDTA, H2O2, and horseradish peroxidase. Molecular iodine is also bleached but at a slower rate than I-3. Evidence is presented to show that this bleaching of I-3 is due to enzymatic conversion of I-3 to iodide in presence of EDTA and H2O2 and this involves pseudocatalatic degradation of H2O2 to O2.

Cell-free extracts of submaxillary glands from rat and goat iodinate exogenously added 3,5-diiodothyronine to form 3,5,3’-triidothyronine and thyroxine. This capacity was elevated after either thyroidectomy or exposure of rats to cold (3 ±1 ‡C) for 5 h. However, there was no further increase of iodination of 3,5-diiodothyronine when the thyroidectomized rats were exposed to cold stress. The submaxillary extracts have the ability to incorporate radioactive iodide into 3,5-diiodothyronine, 3,5,3’-triiodothyronine and thyroxine. The presence of 3,5-diiodothyronine was also detected in the soluble supernatant of submaxillary extract.

In anin vitro rat liver slice incubation system, the synthesis of fibrinogen, when measured by immunoprecipitation technique was stimulated in the presence of epinephrine. An increase in poly (A)+ RNA content of the liver slice was also observed after epinephrine treatment. Thisin vitro experiment demonstrated that epinephrine stimulatedde novo synthesis of fibrinogen by acting directly on the liver.

The effect of Ep on radioactive glucose and methyl-alpha-D-glucoside transport by rat erythrocytes and bone marrow cells were studied. There is initial linearity followed by saturation kinetics of [14C]glucose transport by the erythrocytes of starved and starved plus Ep-treated rats at different concentrations of glucose. Starvation caused slight inhibition of glucose transport which increased markedly on Ep administration to starved rats. Normal animals failed to show any significant change in glucose transport after Ep treatment. Methyl-alpha-D-glucoside inhibited the Ep-stimulated glucose transport significantly. Ep also stimulated the transport of radioactive methyl-alpha-D-glucoside which was competitively inhibited in presence of D-glucose. Glucose transport in erythrocytes was found to be sensitive to metabolic inhibitors like azide and DNP. A sulfhydryl reagent and ouabain also inhibited the transport process. Ep stimulated glucose and methyl-alpha-D-glucoside transport in the bone marrow cells of starved rats. The sugar analog competitively inhibited the glucose transport in bone marrow cells and vice versa.

Alanine-2-oxoglutarate aminotransferase activity in mouse liver is stimulated by the intravenous injection of glucagon. The stimulation is abolished by pretreatment with actinomycin D indicating that the increased activity is probably due to new enzyme formation. Administration of dibutyryl cyclic AMP, isoproterenol, an activator of adenyl cyclase and theophylline, an inhibitor of phosphodiesterase also increases the enzyme activity suggesting the involvement of cyclic AMP in glucagon-mediated increase of enzyme activity.

Cold exposure of rats for 3 h (6 +/- 2 degrees C) caused an increase in amylase activity of the submaxillary gland. This effect was not observed in other salivary glands i.e. parotid and sublingual or in the pancreas. The increase of amylase activity during cold exposure was completely abolished by the beta-receptor antagonist, propranolol, and the alpha-receptor antagonist, phenoxybenzamine, reduced the effect. Administration of actinomycin D to the cold-exposed rats produced a tremendous increase of enzyme activity instead of abolition of the increase as had been expected.

In vivo administration of short acting or long acting inhibitor of monoamine oxidase elevates plasma fibrinogen level in rats. These inhibitors cannot elevate plasma fibrinogen level in rats pretreated with p-chlorophenyl alanine and alpha-methyl-meta-tyrosine, inhibitors of catecholamine and serotonin formation respectively. However, administration of epinephrine or serotonin under the above experimental condition elevates fibrinogen level indicating rise of biogenic monoamine, even for a short period, is a good stimulus for increase of plasma fibrinogen. Dibutyryl cyclic-AMP (c-AMP) also elevates fibrinogen level and shows additive effect with theophylline or serotonin.

Surgical removal of submaxillary glands in immature female rats caused an increase in size and about 3-fold increase in dry and wet weight of the uterus compared to that of the sham operated animals of the same age group. Histological examination reveal a significant increase in the diameter of the uterus with considerable elongation of the luminal epithelium from cubical to columnar in the experimental group. Biochemical studies showed that the uterine peroxidase, a marker enzyme for oestrogen action, increased (P < 0.01) on submaxilariectomy. Incorporation of [14C] phenylalanine into the nuclear fraction of uterus was also enhanced significantly on removal of submaxillary glands. The elevation of peroxidase activity as well as of [14C]phenylalanine incorporation into the nuclear fraction due to removal of submaxillary glands were abolished in ovariectomized rats suggesting the involvement of ovarian hormones. The results show that oestrogen was responsible for all the above mentioned changes, whereas progesterone had little effect. Results further suggest the existence of a factor in the submaxillary glands through which they exert an effect on the uterus and ovary.

L-Epinephrine, serotonin, and isoproterenol stimulate the incorporation of [14C]leucine into thrombin-induced clottable protein; this stimulation was abolished by actinomycin D. The incorporation of 32P into total RNA of rat liver, the site of fibrinogen synthesis, was stimulated by epinephrine and was highest at 2 h after 32P administration. [14C]Orotic acid incorporation into polysomal RNA of liver was also increased significantly by epinephrine and serotonin. The immunoprecipitation of newly synthesized protein by monospecific antibody raised against pure rat fibrinogen clearly demonstrates that L-epinephrine increased fibrinogen formation in vivo under the experimental condition. Translation of poly (A)-containing RNA from total polysomal RNA clearly indicates that L-epinephrine increased mRNA specific for fibrinogen.

1. 1. Complete organization of the cell components was essential for succinate production, since the cell-free extract of Leishmania donovani could not produce succinate from PEP or pyruvate by carboxylation reaction. But it could effectively produce oxaloacetate or malate from PEP or pyruvate and bicarbonate. 2. 2. Studies with a cell-free extract of the organism further showed that at least three different carboxylating systems were present in the preparation of the organism. 3. 3. The activities of NADP-dependent malic enzyme, NADH-dependent malate dehydrogenase and fumarase were shown to be present in the cell-free extract of L. donovani .

The effect of erythropoietin on [14C]acetate incorporation into the lipids of erythrocyte menbranes in starved and in phenylhydrazine-treated rats was studied. The incorporation was increased in both in vivo and in vitro experiments after injection of erythropoietin. However, addition of erythropoietin in the incubation mixture for acetate incorporation did not show any stimulation, suggesting that its effect was manifested only when it was injected in vivo. Erythropoietin showed no effect on the [14C]acetate incorporation into the lipids of leukocytes and a very slight effect on platelets.

A highly active peroxidase (EC 1.11.1.7) has been found to be localized in the mitochondria isolated from the fundic region of mouse stomach. The stomach has also the property of concentrating iodide significantly. Evidence has been presented to show that the peroxidase is orientated outside the mitochondrial membrane. The enzyme is strongly inhibited by antithyroid drugs like methimazole and thiouracil. Azide and cyanide completely inactivate the enzyme. The activity is inhibited by SH-blocking reagents like mersalyl or pchloromercuribenzene sulphonate, but not by N-ethylmaleimide. The enzyme is also sensitive to the action of some proteolytic enzymes. It can catalyse the formation of mono- or diiodotyrosine from tyrosine or monoiodotyrosine as substrate, respectively. The enzyme is capable of synthesizing thyroxine and triiodothyronine on the backbone of a protein, such as thyroglobulin or albumin.

Respiratory studies with the protozoal organism Leishmania donovani have shown that this organism is capable of utilizing ribose as substrate. Studies with cell-free extract of this organism showed that it is rich in some key enzymes of the pentose phosphate pathway. Carbon dioxide production from glucose-1- 14 C and glucose-6- 14 C indicates that in this organism probably the Embden-Meyerhof and Parnas pathway is playing a more important role than the shunt pathway.

During an attempt to explore the mechanism of cobalt and vitamin B 12 effect on the peroxidase and tyrosine iodinase activity, it was observed that cobalt and not vitamin B 12 in vivo elicits a high plasma titer of a humoral factor, erythropoietin, which caused increased 59 Fe incorporation into red blood corpuscles. This erythropoietin was extracted from plasma or kidney and partially purified. This partially purified erythropoietin preparation when injected into starved rat caused increased 59 Fe incorporation into red blood corpuscles. The potency of the partially purified erythropoietin preparation from plasma and kidney was compared. This partially purified erythropoietin, as well as sheep step-I erythropoietin, when administered to mice gave rise to a response almost similar to cobalt administration. It appears that the effect of cobalt on the peroxidase or tyrosine iodinase activity might be mediated through the elevation of the erythropoietin titer of plasma. Moreover, this erythropoietin was found to stimulate both peroxidase and tyrosine iodinase activity of the enzyme preparation of submaxillary gland in experiments in vitro. Erythropoietin, purified further in our laboratory, also produced the same effect. Neuraminidase-treated erythropoietin, in contrast, failed to execute any such stimulatory effect on the iodination of tyrosine.

SUMMARY Subcellular fractions of goat submaxillary gland catalysed the iodination of monoiodotyrosine to form di-iodotyrosine; the microsomal fraction was found to have maximum activity. The enzyme needed copper or hydrogen peroxide for its activity. Thiourea and thiouracil inhibited the enzymic activity. Manganese and ferric ions inhibited the enzymic formation of di-iodotyrosine. The enzyme was solubilized by sodium desoxycholate treatment.

1. 1. A comparative study on some aspects of the catalytic properties of peroxidase and iodinating enzymes of thyroid and submaxillary glands of the mouse has been made. 2. 2. Both the peroxidases are soluble and most of the activity is located in 105,000 g supernatant. However, the particulate fractions also show considerable activities. 3. 3. The iodinating enzyme of the thyroid gland is particle-bound, while that of the submaxillary gland is soluble in character. The 105,000 g supernatant of the thyroid gland cannot catalyse the iodination reaction even after dialysis. 4. 4. Both are true peroxidases, as evidenced by guaiacol peroxidation, however they differ in their relative ability to catalyse the oxidation of different electron donors such as o -dianisidine, guaiacol, and KI. 5. 5. The thyroid and submaxillary peroxidases differ in their optimum p H values and H 2 O 2 , requirements, the latter being markedly different for the oxidation of KI to I 3 − . 6. 6. The iodinating enzymes of the thyroid and the submaxillary glands differ significantly with respect to their p H optima and their efficiency to catalyse the iodination of tyrosine or monoiodotyrosine. 7. 7. The submaxillary iodinating enzyme is unique in catalysing the “iodine exchange reaction”, while the thyroid iodinating enzyme is incapable of catalysing the same.

Erythropoietin stimulated lactoperoxidase (LP) or horseradishperoxidase (HRP) catalyzed iodination of tyrosine, the stimulation being more pronounced in case of LP. The diiodotyrosine (DIT) formation was stimulated more than MIT formation although the total quantity of DIT formed was less than MIT. Both sheep plasma erythropoietin and human urinary erythropoietin exhibited the above phenomenon and the stimulatory effects were directly propertional to their potencies. Preincubation of erythropoietin with neuraminidase abolished its stimulatory effect. Other glycoprotein hormones like thyrotropic hormone (TSH) and human chorionic gonadotrophin (HCG) did not stimulate the above system.

SUMMARY: Erythrose strongly inhibits the growth of Vibrio cholerae and Vibrio eltor. The inhibition can be reversed by washing the cells free from erythrose with sterile normal saline. The respiration of V. cholerae cells in the presence of glucose is markedly inhibited by erythrose but the oxygen uptake of cell-free extract under the same conditions is not affected. From the results of experiments on the uptake of [14C] glucose and [α-14C]methylglucoside, it may be concluded that erythrose inhibits the transport of glucose across the cell membrane and thereby inhibits the growth of the organism.

An iodinating enzyme has been identified and partially purified from the homogenate of goat submaxillary gland. The iodinating enzyme was purified 130-fold and found to be closely associated with iodide peroxidase activity in every step of purification. The finally purified preparation, although not homogeneous, exhibited both iodinase and peroxidase activities. The iodinase and the iodide peroxidase activities have comparable pH curves, hydrogen peroxide requirement and similar reactions towards some common reagents. Partial inactivation by pronase digestion also affects the two activities in a similar manner and fails to dissociate them. These observations led us to suggest that a single enzyme is responsible for the oxidation of I− and incorporation of active iodine to the tyrosine moiety. The enzyme catalyses a production of 31.0 μmol triiodide × min−1× mg−1 and can also catalyse effectively the oxidation of o-dianisidine which is stimulated 2000% in presence of KI. The enzyme also catalyses the iodination of thyroglobulin and albumin, other than free tyrosine, with the formation of iodotyrosines.

Free thyroid follicular cell suspension was prepared from pharyngeal and head kidney thyroid tissue of a teleost by a continuous trypsinization procedure. Single cell suspensions of both the regions retained the capacity of radioiodine uptake and metabolized it into iodoamino acids and thyroxine. Formation of iodotyrosines and thyroxine in free follicular cells was considerably stimulated by the addition of bovine TSH in the incubation medium and this stimulation was abolished by actinomycin D.

Exposure of rats to 6 +/- 2 degrees C for 3 h caused an increase in peroxidase activity (55%) and tyrosine iodinase activity (40%) of the submandibulary glands. Thiocyanate, hydrogen peroxide and formation of triiodide from iodide were also elevated under the same conditions. Administration of alpha-receptor blockers and indomethacin prevented the rise of peroxidase activity during cold stress whereas beta-receptor blockers were less effective.

In vivo administration of epinephrine or serotonin has been shown to stimulate the incorporation of 14C-orotic acid into Poly(A)+ RNA. However, only epinephrine and not serotonin could stimulate DNA dependent RNA polymerase activity of isolated hepatic nuclei in in vitro experiments.

The peroxidase (EC 1.11.1.7) and iodinase (EC 1.11.1.8) activities of rat submaxillary gland were found to be increased after thyroidectomy. The enzyme activities were maximal on the seventh day after operation and then decreased slightly. However, the enzyme activities were still more than 100% even 28 days following operation. Administration of thyroxine (10 μg/100 g body weight) prevented the increase. Puromycin, cycloheximide, and actinomycin D, the inhibitors of protein synthesis, as well as thiouracil partially abolished the increase of activities. These results suggest that thyroxine acts as a regulator of the iodinase and peroxidase enzyme(s) of submaxillary gland.

Surgical removal of submaxillary gland in immature rats causes a large increase in size and about three to four fold increase in dry and wet weight of uterus compared to that of the sham operated animals of the same age group. Histological examination reveals a significant increase in the diameter of the uterus with considerable elongation of the luminer epithelium from cubical to columnar in the experimental group. Biochemical studies show that the uterine peroxidase (donor: hydrogen-peroxide oxidoreductase, EC 1.11.1.7), a marker enzyme for uterine growth, increases by ten to fifteen fold on submaxillariectomy and returns almost to the normal level on administration of submaxillary gland extract (105,000 × g supernatant) to the submaxillariectomized animals. Estrogen estimation by radioimmunoassay shows a similar increase of three to four fold on removal of submaxillary glands and decrease almost to the normal value on administration of the submaxillary extract.

A method has been developed for the isolation of cells, high in iodine uptake and peroxidase activity, from the stomach and submaxillary gland of mice. The isolated cells could produce protein-bound monoiodotyrosine, di-iodotyrosine and an unknown iodocompound. The reactions were catalysed by peroxidase and were sensitive to antithyroid drugs and haemoprotein inhibitors but were insensitive to TSH. In-vitro iodination of stomach or submaxillary soluble proteins with the respective peroxidase yielded similar iodocompounds while thyroxine was produced when thyroglobulin was used instead. J. Endocr. (1985) 106, 159–165

Intramuscular administration of iproniazid caused significant elevation of plasma fibrinogen in rats and rabbits. The elevation may be due to accumulation of monoamines, as the drug inhibited plasma monoamine oxidase quite strongly. This possibility was further supported by the fact that the direct administration of 5HT also elevated the plasma fibrinogen level by about 50 per cent. The effect of 5HT was more pronounced (about 75 per cent) when rabbits were pretreated with reserpine. The above observation has also been confirmed by reserpine-treated rats where different biogenic amines like 5HT, tryptamine, tyramine and epinephrine at low doses elevated plasma fibrinogen level. These biogenic monoamines possibly mediated their action through synthesis of protein since the increase of plasma fibrinogen level after 5HT administration was prevented by pretreatment of animals with actinomycin D or cyclohexamide.

Reconstitution of membrane-bound enzymes and transport proteins with the artificial phospholipid bilayer (liposomes) is one of the most useful techniques to study the functional aspects of these proteins. Several biochemical and biophysical parameters related to the expression of the functions of these proteins can be conveniently studied in the reconstituted proteoliposomes. Methods have been described for the preparation of model membranes and emphasis has been given specially to liposomes. Methodologies for the reconstitution of biologically active proteoliposomes using varieties of membrane-bound enzymes and proteins and their assay have been presented in details. The merits and demerits of each method and the subsequent modification of the technique have been indicated. Factors controlling the orientation and functions of the enzymes and carrier proteins in the reconstituted proteoliposomes have also been described. Attempts have been made to include few examples to describe how the reconstituted model membrane is helping us to know the molecular basis of many salient features of the biomembranes. Finally, the possible application of the proteoliposomes for the study of other complex membrane phenomena has been postulated.

Ep enhanced the exchange of unesterified cholesterol from plasma to the RBC membrane and vice versa. Similar to unesterified cholesterol, the exchange of phospholipids from plasma to the RBC membrane and vice versa in starved rats increased on the administration of Ep. But, unlike cholesterol exchange, the hormone favored phospholipid transport from the RBC membrane to plasma more significantly than from plasma to the RBC membrane.

Starved animals having low levels of erythropoietin in blood showed increased MDA, fluorescent pigments, and met-Hb values whereas the hemoglobin concentration decreased significantly on starvation. In vivo and in vitro studies with Ep reversed the effects of starvation and brought these values close to normal. The activities of the enzymes (SOD, catalase, GSH-PX, GR G6PD, and 6PGD) which protect the RBC membrane directly or indirectly from peroxidative threat, decreased on starvation and restored to normal levels after Ep treatment.