Biochemical medicine最新文献

Seven aromatic metabolites of phenylalanine were determined in plasma of 20 patients with classical phenylketonuria by means of capillary gas chromatography. The results obtained showed good correlation with plasma phenylalanine levels. Plasma aromatic acid levels may prove useful in the diagnosis and management of phenylketonuria, as well as in research of this disorder.

The effect of isatin (indole-2,3-dione) on d-glucose uptake has been studied in rat intestine. Isatin at 6 mm concentration significantly inhibited both the sugar uptake and transmural (mucosal to serosal side) transport in the intestine. The suppression of glucose uptake by isatin was irreversible. Similar to the action of various SH-group-reacting agents, isatin inhibited the sugar uptake, presumably by binding to membrane sulfhydryl groups through a covalent linkage. Isatin-induced reduction in glucose uptake was unaffected by pH (between 5.5 and 8.4) and by DTT addition to incubation medium. Inhibition of sugar uptake by isatin and harmaline was additive in nature; this suggested that these compounds interact at different sites on the microvillus membrane surface.

Human placental β-glucocerebrosidase modified by covalent attachment of N²-{N²,N⁶-bis[3-(α-d-mannopyranosylthio)propionyl]-l-lysyl}-N⁶-[3-(α-d-mannopyr-anosylthio) propionyl]-l-lysine was administered to rats by intravenous injection. Comparison of enzyme distribution in isolated liver cell populations indicates an increase in enzyme-specific activity of 18-fold in nonparenchymal cells and only 1.5-fold to hepatocytes compared to uninjected control animals. This macrophage-specific delivery of an active lysosomal enzyme has potential for application in enzyme replacement trials.

Three different isocratic systems for the separation by reversed-phase high-performance liquid chromatography (HPLC) of different species of insulin have been investigated. The effect of different solvent compositions and temperatures on elution time and resolution have been studied. These studies have been used to devise a method for reversed-phase liquid chromatographic separation of bovine, porcine, and human insulin, as well as the A and B chains of bovine insulin. The method can also be used for the separation of the various products of the iodination of porcine insulin. ¹²⁵I-A14 tyrosine-labeled porcine insulin can be readily separated from nonlabeled porcine insulin and from other idoinated constitutents of the mixture. A flow-though gamma-counting system that was designed for this work is described.

In five of six patients with symptomatic Wilson's disease (WD) with increased hepatic copper content, increased renal copper excretion, and decreased serum concentrations of ceruloplasmin, significantly low levels of hepatic reduced glutathione (GSH) were found. Three of these patients showed increased levels of oxidized glutathione which in part could account for the missing GSH. These changes may result from increased lipid peroxidation due to the rise of intracellular copper concentration. Furthermore, WD patients showed a 50% decrease in the activity of hepatic GSH S-transferases.

From these results we conclude that the disturbance in the hepatic glutathione system of patients with symptomatic WD may contribute to the perpetuation of liver damage. These patients, additionally, may be predisposed to an increased sensitivity to drugs interacting with glutathione.

We measured the activity of aromatic l-amino acid decarboxylase with l-dihydroxyphenylalanine as a substrate (DOPA decarboxylase) in normal lung tissues and lung tumors obtained fresh at surgery. The activity in control human lung tissues was low and variable: 3.50 ± 0.42 pmole/min/mg protein (n = 56, mean ± SE, range 0.01–15), indicating the wide individual variations. Most of small cell carcinoma specimens showed very high activity, as compared with both control lung tissues and with other types of non-SCC lung cancers. Similar results were also obtained in the athymic mice heterotransplants of SCC. High activity was also observed using 5-l-hydroxytryptophan as a substrate (5-HTP decarboxylase) in nine SCC samples. Serotonin was not detected in any control lung tissues, but was detected in all the nine SCC samples, but dopamine was detected only in three out of nine SCC samples.

Manganese causes a significant rise in hepatic glucokinase and hexokinase in 16-day-old suckling rats, and has an insulinomimetic effect in producing a precocious emergence of glucokinase (EC 2.7.1.2) and a rise in the low K_m, kexokinases (EC 2.7.1.1) activities. These enzyme changes occur within 6 hr of manganese administration and there are accompanying increases in plasma insulin and hepatic cyclic GMP. That the effect of manganese is at a site other than, or in addition to, insulin secretion is suggested by the significant increases in glucokinase and hexokinase in 16-day-old streptozotocin-diabetic rats; in this group there is also an increase in hepatic cGMP similar in time scale to that of the normal-manganese-treated group. The effects of manganese and insulin were not additive. It is proposed that one site of action of manganese may be at the level of cyclic GMP systems. The results are also discussed in relation to the known action of manganese at the level of the protein phosphatases.

Human spleen dihydroorotate dehydrogenase is associated with the mitochondrial membrane and is linked to the respiratory chain via ubiquinone. The enzyme activity was unaffected by pyridine nucleotides. The product of the reaction, orotate, was a potent inhibitor. However, a range of other naturally occurring pyrimidines or purines had no significant effect on the activity. No evidence for the involvement of a complexed metal ion or for an active sulfhydryl group was obtained.

Purification of the enzyme was achieved by preparation of an acetone powder and extraction with Triton X-100, followed by preparative polyacrylamide gel electrophoresis. Activity was observed by the addition of the artificial electron acceptors, ubiquinone 50 or PMS. Purification resulted in alteration of the pH optimum and of other kinetic characteristics. Two molecular-weight species, of molecular weight 88,000 and 98,000, were consistently observed.

The properties of the human spleen enzyme were similar in principle to those for the rat liver enzyme. Differences in the mode of linkage to the respiratory chain for the mitochondrially bound enzyme, and in the characteristics of the purified enzyme, were observed.

Effect of depot medroxyprogesterone acetate on the hepatic drug-metabolizing enzymes was studied in female protein-deficient and normal pair-fed rats. Treatment with this drug did not cause any change in organ weight, microsomal protein, and soluble protein yield per gram of tissue in both groups. MPA administration resulted in significant increases in the content of cytochrome P-450 and b₅, and activities of benzo[a]pyrene hydroxylase, UDP-glucuronosyltransferase, and NADPH-Cyt c reductase in both pair-fed control and protein-deficient rats. However, the content of glutathione and activity of glutathione-S-transferase were not affected appreciably. The present study suggests that MPA treatment induces drug-metabolizing enzymes in liver to almost the same extent in both protein-deficient and normal pair-fed rats.

Seventy-one pyrimidine analogs have been tested as possible inhibitors of human spleen mitochondrial dihydroorotate dehydrogenase. Of these nine were demonstrated to be effective inhibitors of the enzymic activity. Two compounds, dihydro-5-azaorotate and 6-thiobarbiturate appeared to be specific inhibitors of the DHO-DHase. In addition, three compounds, 5-azaorotate, 5-bromoorotate, and barbiturate were also inhibitory against the two subsequent enzymes of the pathway, orotate phosphoribosyltransferase and orotidylate decarboxylase, so that they could act against three enzymes of the mammalian pyrimidine de novo biosynthetic pathway.