Supernatant Fraction Of Liver Homogenate Research Articles

Pharmacokinetics of clarithromycin in rats with acute renal failure induced by uranyl nitrate

In rats pretreated with dexamethasone (an inducer of CYP3A1/2 in rats) and troleandomycin (an inhibitor of CYP3A1/2 in rats), the area under the plasma concentration-time curve from time zero to time infinity (AUC) values of clarithromycin were significantly smaller (365 compared with 600 micro g min/ml) and greater (1410 compared with 581 micro g min/ml), respectively, than those in control rats. This indicated that clarithromycin was metabolized via CYP3A1/2 in rats. The expression of CYP3A1(23) increased in rats with acute renal failure induced by uranyl nitrate (rats with U-ARF). Hence, it could be expected that AUC of clarithromycin could be smaller in rats with U-ARF. However, after intravenous administration of clarithromycin at a dose of 20mg/kg, the AUC and time-averaged total body (Cl) and nonrenal (Cl(nr)) clearance values were comparable between the two groups of rats. The 9000 x g supernatant fraction of liver homogenates in rats with U-ARF had comparable metabolic activities for clarithromycin compared with those in control rats, suggesting that the CYP3A isozyme responsible for metabolism of clarithromycin seemed not to be expressed considerably in the rats. This could explain the comparable AUC, Cl and Cl(nr) values of clarithromycin between the two groups of rats.

Actions of quercetin on gluconeogenesis and glycolysis in rat liver

1. The action of quercetin on glucose catabolism and production was investigated in the perfused rat liver. 2. Quercetin inhibited lactate production from glucose: 80% inhibition was found at a quercetin concentration of 100 µM, and at higher concentrations inhibition was complete. 3. Pyruvate production from glucose presented a complex pattern, but stimulation was evident at 100 and 300 µM quercetin. Oxygen uptake tended to be increased. 4. Glucose synthesis from lactate and pyruvate was inhibited. Inhibition was already evident at 50 µM quercetin and almost complete at 300 µM. Concomitantly, the increment in oxygen uptake caused by lactate plus pyruvate was stimulated by 50 µM quercetin, but clearly inhibited by higher concentrations (100-500 µM). 5. Glucose phosphorylation in the high-speed supernatant fractions of liver homogenates was inhibited by quercetin, but only at concentrations above 150 µM. 6. It is concluded that quercetin can inhibit both glucose degradation and production and increase the cytosolic NAD + /NADH ratio. 7. These effects are likely to arise from many causes. Reduction of oxidative phosphorylation, inhibition of Na + -K + -ATPase, inhibition of glucokinase and inhibition of glucose 6-phosphatase could all contribute to the overall action of quercetin.

Glucose and glycogen catabolism in perfused livers of Walker-256 tumor-bearing rats and the response to hormones

The alterations in hepatic glucose and glycogen catabolism were evaluated in rats bearing the Walker-256 tumor. Food intake was monitored concomitantly with measurements of the in vivo hepatic glycogen levels. Glycogenolysis, glycolysis and oxygen uptake were measured in the isolated perfused liver. The hepatic glucose phosphorylating capacity was measured in the high-speed supernatant fraction of liver homogenates. Food intake was 21.4% reduced in tumor-bearing rats; the glycogen levels were decreased by 63.6%. Initial basal rates of glucose release (glycogenolysis) and lactate+pyruvate production from endogenous glycogen (glycolysis) in the perfused liver were not changed by the tumor-bearing state, resulting in a higher relative rate of glycogen breakdown (% of glycogen degradation per unit time). In absolute terms stimulation of glycogen mobilization by glucagon or norepinephrine was smaller in the tumor-bearing state. The percentage of extra glycogen degradation per unit time caused by both hormones, however, was practically the same in the control and in the tumor-bearing state. The hepatic glucose phosphorylating capacity was reduced from 3.92±0.39 nmol min −1 (mg protein) −1 in normal rats to 2.61±0.23 nmol min −1 (mg protein) −1 in livers from tumor-bearing rats. Glycolysis from exogenous glucose (20 mM) in perfused livers was diminished from 0.136±0.023 μmol min −1 (g liver) −1 in normal rats to 0.046±0.008 μmol min −1 (g liver) −1 in tumor-bearing rats. It can be concluded that livers from rats bearing the Walker-256 tumor are less able to transform glucose and accumulate glycogen while possessing a greater tendency of releasing glucose from the glycogen stores.

Glucose phosphorylation capacity and glycolysis in the liver of arthritic rats.

Glycolysis and the glucose phosphorylation capacity of livers from arthritic rats were studied because alterations in these parameters are suggested by some studies. Arthritis was induced in male albino rats (Wistar; 180-220 g). The animals were injected with 100 microl heat inactivated Mycobacterium tuberculosis suspended in mineral oil at a concentration of 0.5% (w/v). Animals showing lesions after 14 to 21 days were selected. Glucose phosphorylation was measured in the high speed supernatant fraction of liver homogenates and glycolysis in the isolated perfused liver. The glucose concentration for half-maximal rates was reduced from 18.32+/-5.69 in normal to 9.84+/-3.15 mM in arthritic rats (p = 0.024). Vmax was increased from 8.77+/-0.27 in normal to 11.49+/-0.29 nmol min(-1) mg protein(-1) in arthritic rats (p = 0.001). Perfused livers from arthritic rats showed a 2.43-fold higher rate of glycolysis. Livers from arthritic rats present a higher glucose phosphorylation capacity. Possibly this phenomenon is caused by circulating inflammatory mediators produced during adjuvant-induced arthritis.

Effect of phenobarbital, 3-methylcholanthrene, and chloramphenicol pretreatment on the pharmacokinetics and pharmacodynamics of azosemide in rats.

The effects of pretreatment with the enzyme inducers phenobarbital (PB) and 3-methylcholanthrene (3-MC) and the enzyme inhibitor chloramphenicol (CM) on the pharmacokinetic and pharmacodynamic parameters of azosemide were examined after intravenous (i.v.) administration of azosemide, 10 mg kg-1, to rats. The nonrenal clearance (1.63 versus 3.30 mL min-1 kg-1) of azosemide increased significantly in 3-MC pretreated rats. This suggested that the nonrenal metabolism of azosemide increased by pretreatment with 3-MC. This relationship was supported by the significant decrease in 24 h urinary excretion of unchanged azosemide in 3-MC pretreated rats (54.1 versus 41.1% of i.v. dose). This relationship was also supported at least in part by the results of a liver homogenate study; the amount of azosemide remaining per gram of liver decreased significantly (48.2 versus 43.0 micrograms) and the amount of M1 formed increased significantly (4.88 versus 6.66 micrograms when expressed in terms of azosemide) in 3-MC pretreated rats after 30 min incubation of 50 micrograms azosemide in 9000 g supernatant fractions of liver homogenates. The content of hepatic cytochrome P-450 (0.751 versus 1.57 nmol/mg protein) and the weight of liver (3.53 versus 4.20% of body weight) increased significantly in 3-MC pretreated rats, suggesting that the metabolizing enzyme(s) for azosemide seemed to be induced by pretreatment with 3-MC. The 8 h urine output (29.2 versus 18.1 mL) and 8 h urinary excretion of sodium (4.02 versus 2.39 mmol) and chloride (4.01 versus 2.73 mmol) per 100 g body weight decreased significantly in 3-MC pretreated rats. However, the diuretic, natriuretic, kaluretic, and chloruretic efficiencies were not significantly different between the control and 3-MC pretreated rats. The pharmacokinetic and pharmacodynamic parameters of azosemide were not significantly different between the control and PB pretreated rats, and similar results were also obtained from the control and CM pretreated rats. The above data indicate that the metabolizing enzyme(s) for azosemide seem(s) to be neither induced by PB pretreatment nor inhibited by CM pretreatment. However, the content of hepatic cytochrome P-450 and the weight of liver increased significantly in PB pretreated rats, while the values were not significantly different between the control and CM pretreated rats.

EFFECT OF PHENOBARBITAL, 3-METHYLCHOLANTHRENE, AND CHLORAMPHENICOL PRETREATMENT ON THE PHARMACOKINETICS AND PHARMACODYNAMICS OF AZOSEMIDE IN RATS

The effects of pretreatment with the enzyme inducers phenobarbital (PB) and 3-methylcholanthrene (3-MC) and the enzyme inhibitor chloramphenicol (CM) on the pharmacokinetic and pharmacodynamic parameters of azosemide were examined after intravenous (IV) administration of azosemide, 10 mg kg−1, to rats. The nonrenal clearance (1·63 versus 3·30 mL min−1 kg−1) of azosemide increased significantly in 3-MC pretreated rats. This suggested that the nonrenal metabolism of azosemide increased by pretreatment with 3-MC. This relationship was supported by the significant decrease in 24 h urinary excretion of unchanged azosemide in 3-MC pretreated rats (54·1 versus 41·1% of IV dose). This relationship was also supported at least in part by the results of a liver homogenate study; the amount of azosemide remaining per gram of liver decreased significantly (48·2 versus 43·0 μg) and the amount of M1 formed increased significantly (4.88 versus 6.66 μg when expressed in terms of azosemide) in 3-MC pretreated rats after 30 min incubation of 50 μg azosemide in 9000 g supernatant fractions of liver homogenates. The content of hepatic cytochrome P-450 (0·751 versus 1·57 nmol/mg protein) and the weight of liver (3.53 versus 4·20% of body weight) increased significantly in 3-MC pretreated rats, suggesting that the metabolizing enzyme(s) for azosemide seemed to be induced by pretreatment with 3-MC. The 8 h urine output (29·2 versus 18·1 mL) and 8 h urinary excretion of sodium (4·02 versus 2·39 mmol) and chloride (4·01 versus 2·73 mmol) per 100 g body weight decreased significantly in 3-MC pretreated rats. However, the diuretic, natriuretic, kaluretic, and chloruretic efficiencies were not significantly different between the control and 3-MC pretreated rats. The pharmacokinetic and pharmacodynamic parameters of azosemide were not significantly different between the control and PB pretreated rats, and similar results were also obtained from the control and CM pretreated rats. The above data indicate that the metabolizing enzyme(s) for azosemide seem(s) to be neither induced by PB pretreatment nor inhibited by CM pretreatment. However, the content of hepatic cytochrome P-450 and the weight of liver increased significantly in PB pretreated rats, while the values were not significantly different between the control and CM pretreated rats. © 1997 John Wiley & Sons, Ltd.

Pharmacokinetics and pharmacodynamics of furosemide after intravenous and oral administration to spontaneously hypertensive rats and DOCA‐salt‐induced hypertensive rats

The pharmacokinetics and pharmacodynamics of furosemide were investigated after intravenous (i.v.), 1 mg/100 g body weight, and oral administration, 2 mg per 100 g body weight, to spontaneously hypertensive rats (SHRs) and deoxycorticosterone acetate-salt-induced hypertensive rats (DOCA-salt rats). After i.v. administration, the 8 h urinary excretion of furosemide/g kidney (397 versus 572 micrograms) was significantly lower and the non-renal clearance (5.78 versus 3.94 ml min-1 kg-1) was significantly faster in SHRs of 16 weeks of age than in age-matched control Wistar rats. This suggested that the non-renal metabolism of furosemide could be faster in SHRs of 16 weeks of age than in age-matched control Wistar rats, and this could be supported by the significantly greater amount of 4-chloro-5-sulphamoyl anthranilic acid, a metabolite of furosemide, excreted in 8 h urine as expressed in terms of furosemide (11.1 versus 4.79% of the i.v. dose) in SHRs. It could also be supported at least in part by a study of liver homogenate; the amount of furosemide remaining per gram of liver after 30 min incubation of 50 micrograms of furosemide with the 9000g supernatant fraction of liver homogenate was significantly smaller (40.4 versus 43.7 micrograms) in SHRs of 16 weeks of age than in age-matched Wistar rats. The greater metabolic activity of furosemide in liver may also be supported by the result that the amount of hepatic cytochrome P-450 (0.7013 versus 0.5186 nmol/mg protein) and the weights of liver (3.52 versus 2.93% of body weight) were significantly greater in SHRs of 16 weeks of age than in age-matched Wistar rats. After i.v. administration of furosemide, the 8 h urine output (9.93 versus 16.5 ml) and 8 h urinary excretion of sodium (1.21 versus 2.05 mmol) and chloride (1.37 versus 2.17 mmol) per gram of kidney in SHRs of 16 weeks of age were lower than those in age-matched Wistar rats, this could be due to the significantly smaller amount of furosemide excreted in 8 h urine per gram of kidney. After oral administration, the pharmacokinetics and pharmacodynamics of furosemide were not significantly different between SHRs and the control Wistar rats of 16 weeks of age. After i.v. and oral administration of furosemide, there were no significant differences in the pharmacokinetics and pharmacodynamics between DOCA-salt rats and control SD rats of 16 weeks of age except for the significantly lower urinary excretion of potassium per gram of kidney in DOCA-salt rats.(ABSTRACT TRUNCATED AT 400 WORDS)

Chlorinated drinking water is mutagenic and causes 3-methylcholanthrene type induction of hepatic monooxygenase

Acid/neutral fractions of 4 chlorinated drinking water samples were tested for mutagenicity in the Ames' test and injected intraperitoneally to 10- and 20-day-old Wistar rats at doses of 200 and 100 liters of water/kg body weight. Cytochrome P-450 mediated enzyme activities of ethylmorphine-N-demethylase (EMND), 7-ethoxycoumarin-O-deethylase (ECOD), 7-ethoxyresorufin-O-deethylase (EROD) and 7-pentoxyresorufin-O-dealkylase (PEROD) were determined in the 9000 g supernatant fraction of liver homogenate. EROD was introduced by the concentrates. The induction was related to the mutagenic activity. About 4-fold increase in activity was observed with the most mutagenic sample. PEROD was also slightly enhanced. EMND and ECOD activities were not affected by the lower dose, but the higher dose caused inhibition of 30–40%. Although the extracts were not toxic to bacteria, they were unexpectedly toxic to rats. It is concluded that the samples contained 3-methylcholanthrene (3-MC) type inducer(s).

Inhibitors of cholesterol biosynthesis. Further studies of the metabolism of 5α-cholest-8(14)-en-3β-ol-15-one in rat liver preparations

5α-Cholest-8(14)-en-3β-ol-15-one is a potent inhibitor of sterol biosynthesis in mammalian cells in culture and has significant hypocholesterolemic activity upon oral administration to rodents and non-human primates. The conversion of the 15-ketosterol to cholesterol upon incubation with the 10,000 × g supernatant fraction of rat liver homogenate preparations under aerobic conditions has been reported (D.J. Monger, E.J. Parish and G.J. Schroepfer, Jr. (1980) J. Biol. Chem. 255, 11122–11129). Presented herein are results of studies of the metabolism of [2,4- 3H]5α-cholest-8(14)-en-3β-ol-15-one obtained upon incubation with the microsomal, cytosolic and the 10,000 × g supernatant fractions of liver homogenates of female rats under a variety of conditions. The results of these studies indicated metabolism of the 15-ketosterol to materials with the chromatographic properties of fatty acid esters of the 15-ketosterol, fatty acid esters of C 27-monohydroxysterols, a component similar to the 15-ketosterol (possibly an isomer of the Δ 8(14)-15-ketosterol), and a polar component. Detailed studies of the C 27-monohydroxysterols obtained from incubation of the 15-ketosterol under anaerobic conditions indicated the formation of labeled 5α-cholesta-8, 14-dien-3β-ol and a 5α-cholest-7-en-3β-ol which were characterized by their behavior on silicic acid column chromatography, by the behavior of their acetate derivatives on medium pressure liquid chromatography on alumina-AgNO 3 columns, and by co-crystallization of the labeled sterols with authentic unlabeled standards. The identification of 5α-cholesta-8, 14-dien-3β-ol and 5α-cholest-7-en-3β-ol as metabolites of the 15-ketesterol, coupled with previous studies of the metabolism of 5α-cholesta-8, 14-dien-3β-ol and of 5α-cholest-8(14)-ene-3β, 15α-diol and 5α-cholest-8(14)-ene-3β, 15β-diol has permitted the formulation of a scheme for the overall metabolism of the 15-ketosterol to cholesterol.

Glycohydrolases in diabetes: characterization of acid α-glucosidase from liver and neutral α-glucosidase from sera of diabetic patients and controls

Acid alpha-glucosidase activity measured in the supernatant fraction of liver homogenates obtained from adult-onset diabetic patients is significantly decreased when compared to controls (2.86 +/- 1.18 and 5.79 +/- 1.82 nmoles/min/mg protein +/- S.D., respectively). The biochemical properties (Km values, thermostability, pH optimum, isoelectric focusing profiles) of acid alpha-glucosidase obtained from the livers of diabetic patients were similar to those of acid alpha-glucosidase obtained from the livers of controls. Mixing studies gave additivity of acid alpha-glucosidase activity suggesting that neither inhibitors nor activators are present (in diabetic and control livers, respectively). Neutral alpha-glucosidase activity measured in the sera of diabetic patients was significantly increased when compared to controls (4.35 +/- 1.82 and 2.44 +/- 1.05 nmoles/h/ml +/- S.D., respectively). Neutral alpha-glucosidase in the sera of diabetic and control patients has a similar pH optimum but the enzyme in the serum of diabetics has a slightly lower apparent Km value for the 4-methylumbelliferyl substrate (0.6 vs. 0.9 mmol/L) and slightly increased thermostability. Experiments involving dialysis of patient serum, addition of glucose to patient serum and mixing of control and diabetic patient sera all suggest that glucose exhibits only slight inhibition of serum neutral alpha-glucosidase activity. Isoelectric focusing indicates that neutral alpha-glucosidase activity in the sera of diabetic patients is consistently different from the enzyme in the sera of control patients in that a significantly smaller percentage of activity is found in the acidic region with pI values less than 4.8.

Amphibian ornithine aminotransferase: some general biological and chemical characteristics

1. 1. The specific activities of ornithine aminotransferase (OAT) in the livers of adult Rana pipiens and pre- and postmetamorphic R. catesbeiana were measured. 2. 2. There were no significant differences between specific activities of liver OAT from any of the organisms examined. 3. 3. Cell fractionation studies indicated most of the OAT activity was present in the supernatant fraction of liver homogenates from R. catesbeiana and Bufo woodhousii, while liver homogenates from R. pipiens exhibited most of the OAT activity in the mitochondrial fraction.

Sterol synthesis: studies of the metabolism of 14 alpha-methyl-5 alpha-cholest-7-en-3 beta-ol.

[3 alpha-3H]14 alpha-Methyl-5 alpha-cholest-7-en-3 beta-ol has been prepared by chemical synthesis. The metabolism of this compound has been studied in the 10,000 g supernatant fraction of liver homogenates of female rats. Efficient conversion to cholesterol was observed. Other labeled compounds recovered after incubation of [3 alpha-3H]14 alpha-methyl-5 alpha-cholest-7-en-3 beta-ol with the enzyme preparations include the unreacted substrate, 5 alpha-cholesta-7,14-dien-3 beta-ol, 5 alpha-cholesta-8,14-dien-3 beta-ol, cholesta-5,7-dien-3 beta-ol, 5 alpha-cholest-8(14)-en-3 beta-ol, 5 alpha-cholest-8-en-3 beta-ol, and 5 alpha-cholest-7-en-3 beta-ol. In addition, significant amounts of incubated radioactivity were recovered in steryl esters. The steroidal components of these esters were found to contain labeled 14 alpha-methyl-5 alpha-cholest-7-en-3 beta-ol, 5 alpha-cholesta-8,14-dien-3 beta-ol, 5 alpha-cholesta-7,14-dien-3 beta-ol, 5 alpha-cholest-8-en-3 beta-ol, 5 alpha-cholest-7-en-3 beta-ol, and cholesterol.

Impaired drug metabolism in experimental cirrhosis in the rat

Cirrhosis was produced in the rat by chronic administration of phenobarbital and carbon tetrachloride for 10 weeks. The metabolism of three substrates, aminopyrine. hexobarbital and propranolol, has been investigated in a 9000 g supernatant fraction of liver homogenate and in intact hepatocytes in cirrhotic livers and compared to appropriate phenobarbital-treated controls. In the 9000 g supernatant preparation, the maximal velocity of metabolism for each substrate and the cytochrome P450 concentration were reduced significantly in cirrhotic livers, while total protein and DNA concentration remained unchanged. In intact hepatocytes, the V maxfor each substrate was reduced, while the apparent K m remained unchanged. In both in vitro systems, the metabolism of propranolol and of hexobarbital was influenced by cirrhosis to a greater extent than that of aminopyrine. It is concluded that the carbon tetrachloride-phenobarbital model of cirrhosis in the rat is a suitable model in which to study the effects of chronic liver disease on drug disposition.

Irreversible Androgenic Programming at Birth of Microsomal and Soluble Rat Liver Enzymes Active on 4-Androstene-3,17-dione and 5α-Androstane-3α,17β-diol

Abstract The metabolism of 5α-[4-14C]androstane-3α, 17β-diol was studied in the microsomal fraction and the metabolism of 4-[4-14C]androstene-3,17-dione was studied in the microsomal and 105,000 x g supernatant fractions of liver homogenates from male and female rats sham-operated or castrated at birth or castrated at the age of 7, 14, 30, or 43 days. The neonatally castrated male and female rats were treated with a single dose of 1.45 µmoles of testosterone propionate or with vehicle only. One group of neonatally castrated female rats was treated with 0.145 µmole of testosterone propionate. It was found that the hepatic steroid metabolizing enzyme activities irreversibly affected (imprinted or programmed) at birth by testicular androgens could be classified into three categories according to their sensitivity toward (irreversible induction) and negative (irreversible suppression). One category of enzymes (represented by the 5α-reductase active on 4-androstene-3,17-dione) was imprinted by the relatively small dose of testosterone propionate (0.145 µmole) and the imprinting process was complete in rats 7 days of age. A second category of enzymes (represented by the 2α-hydroxylase active on 5α-androstane-3α,17β-diol, the 3β-hydroxysteroid reductase active on 5α-androstane-3,17-dione, and the 17α-hydroxysteroid reductase and 5β-reductase active on 4-androstene-3,17-dione) needed a relatively high dose of testosterone propionate (1.45 µmoles) to be imprinted; the imprinting process was complete or almost complete in rats 7 days of age. The third category of enzymes (represented by the 16α-hydroxylase active on 4-androstene-3,17-dione and the 7α-hydroxylase active on 5α-androstane-3α,17β-diol) encompassed enzymes not completely imprinted in rats 7 days of age; in these cases the imprinting process was not complete until 14 to 30 days after birth. The preputial glands were shown to contain steroid metabolizing enzyme activities that were regulated in quite a different way from corresponding liver enzymes. The activity of preputial gland 5α-reductase, when related to wet tissue weight, was suppressed following postpubertal testectomy, whereas liver 5α-reductase was stimulated following similar treatment. The ratio of 3β- to 3α-hydroxylated 5α-reduced metabolites of 4-androstene-3,17-dione found after incubations with preputial glands was not significantly affected by sex or castration, whereas the same ratio measured after incubations with the liver microsomal fraction was much higher in male than in female rats and was found to be determined both by positive neonatal imprinting and by reversible postpubertal androgenic stimulation of 3β-hydroxysteroid reductase.

Developmental changes in mouse liver alcohol dehydrogenase

Alcohol dehydrogenase activity in the supernatant fraction of liver homogenates was studied in adult and newborn mice. Newborn mice were found to have a lower specific activity than adult animals. K m values were higher in the new born animal. Electrophoresis showed two bands for the adult and only one for the newborn, with a greater intensity seen in adult.

Regulation by dietary choline of hepatic fatty acid synthetase in the rat

Fatty acid synthetase activity was measured in the high-speed supernatant fraction of liver homogenates from rats fed a semisynthetic diet low in lipotropic factors. If choline was omitted from the diet, a significant increase of fatty acid synthetase activity was observed after two feedings of the deficient diet. Compared with controls, the increase of fatty acid synthetase activity was of a magnitude that could account for the amount of triglyceride accumulating in the hepatic floating lipid fraction. Gas-liquid chromatographic analysis of the floating lipid triglycerides showed an increased content of palmitic acid due to choline deficiency; this increase could be predicted from the increased fatty acid synthetase activity and its known characteristic yield of palmitic acid.

Metabolism of Drugs by Subfractions of Hepatic Microsomes from Prolonged Ethanol-Treated Rats

In a previous paper, it was reported that in rats treated chronically with ethanol, the side-chain oxidation of hexobarbital, N-demethylation of aminopyrine and /?-hydroxylation of aniline in vitro from either 9,000 g supernatant fraction of liver homogenates or washed microsomes was identical with that of control rats when ethanol was withdrawn and substituted for tap water 24 hr prior to sacrifice. In contrast, the activity of aniline hydroxylase of the rats which continued to ingest ethanol ad libitum up to the time of sacrifice was approx. 1.5-fold increased, compated with that of controls, in spite of no change being detected in hexobarbital oxidase and aminopyrine demethylase activities (1, 2).

The role of L-serine dehydratase in the metabolism of L-serine in the perfused rat liver

1. 1. To determine the role of L-serine dehydratase ( L-serine hydrolyase, deaminating, EC 4.2.1.13) in L-serine metabolism in the rat liver, experiments involving in situ liver perfusion, metabolite measurement in freeze-clamped tissues and enzyme activity determinations were conducted. 2. 2. Results from liver perfusion experiments showed that the maximal rate of gluconeogenesis from 10 mM L-serine was identical to that from equimolar quantity of L-lactate or pyruvate in starved animals. Combining L-serine and L-lactate in the perfusion medium did not produce glucose at a faster rate than with either L-lactate or L-serine alone. Glucose formation from L-serine or L-lactate was inhibited by 4.8 mM quinolinate which was reversible by 4.8 mM MnCl 2. 3. 3. Metabolite measurements revealed that perfusion with L-serine increased liver concentrations of pyruvate, lactate, aspartate, malate, and glutamate. The presence of 4.8 mM quinolinate further elevated the levels of aspartate, malate and glutamate. These metabolite levels dropped after the reversal of quinolinate inhibition by MnCl 2. 4. 4. The increase in glucogenic rate from L-serine by starvation was paralleled by similar increase in serine dehydratase activity in the supernatant fraction of liver homogenate and L-serine disappearance from the perfusion medium. 5. 5. These results strongly suggest that a major portion of L-serine was metabolized by L-serine dehydratase in the perfused rat liver.

Effect of nicotine treatment on the metabolism of nicotine in the mouse liver in vitro.

14C‐nicotine was incubated with a 10000 × g supernatant fraction of liver homogenate from mice pretreated with nicotine for 3, 10 and 17 days respectively. The rate of metabolism was measured by the determination of the oxidative nicotine metabolite cotinine. After intraperitoneal injections of 5 mg/kg of nicotine three times daily for three days a 50% significant decrease in metabolism was observed. A 71% significant decrease in hepatic glycogen was also seen. Giving nicotine 27.8 and 24.5 mg/kg/24 hours in drinking water for 10 and 17 days respectively did not significantly change the metabolism or the hepatic glycogen levels.

Mechanism of stimulation of polyamine synthesis by growth hormone in rat liver

The effect of growth hormone on the synthesis of polyamines, RNA and protein was studied in the normal rat liver. The hormone markedly stimulated the synthesis of spermidine from [ 14C]-methionine. The time-course of this stimulation was close to that observed in RNA and protein. In addition to stimulating spermidine synthesis, growth hormone also caused an early increase in the concentration of hepatic putrescine and markedly stimulated the incorporation of label from [ 14C]ornithine into putrescine. The formation of putrescine from [ 14C]ornithine was also demonstrated in vitro, using the 100 000 × g supernatant fraction of liver homogenate as the source of the enzyme (ornithine decarboxylase). A 4-fold increase in this enzyme activity was observed after growth-hormone treatment. The effects of growth hormone on the ornithine decarboxylase activity, as well as on the accumulation of putrescine and the synthesis of spermidine, could be partially reversed by actinomycin D or puromycin. No change was observed in the capacity of the liver to synthesize spermidine from [ 14C]putrescine after growth-hormone treatment. It is suggested that the stimulation of spermidine synthesis after growth-hormone treatment depends on a primary increase in the concentration of putrescine, a precursor of spermidine.