Effect of colonic distension on exocrine pancreatic secretion of the conscious rat

In the conscious rat the increasing distension of the colon (from 2 to 6 ml) induced a slight decrease of exocrine pancreatic secretion which was more important for protein output than for volume. Nevertheless this inhibitory effect was never significant even with the important distension of 6 ml which induced a visible uneasiness in the animal.     

Reversed-phase high-performance liquid chromatography separation of dimethylaminoazobenzene sulfonyl- and dimethylaminoazobenzene thiohydantoin-amino acid derivatives for amino acid analysis and microsequencing studies at the picomole level

A simple and fast reversed-phase high-performance liquid chromatographic method has been developed for the complete separation of 35 dimethylaminoazobenzene sulfonyl (DABS)-amino acids and by-products. This method allows simultaneous determination of primary and secondary amino acids which can be present in protein and peptide hydrolysates and also detects the presence of cysteic acid, S-sulfocysteine, hydroxyproline, taurine, norleucine, cystine, and delta-hydroxylysine. The precolumn derivatization of amino acids with dimethylaminoazobenzene sulfonyl chloride (DABS-Cl) is simple and quick (10 min at 70 degrees C) and allows the complete reaction of primary and secondary amino acids. The separation of the compounds under investigation is achieved in 25 min using a reversed-phase 3-microns Supelcosil LC-18 column at room temperature. The versatility of the proposed method is documented by amino acid determination on protein samples obtained using different hydrolysis techniques (HCl, methane-sulfonic acid, and NaOH), with attention given to the detection of tryptophan in protein samples with high sugar concentration. Furthermore, we have reported the experimental conditions necessary to apply this method to the amino acid analysis of very low amount of proteins (1 to 5 micrograms) electroeluted from a stained band after sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The stability of DABS-derivatives, the short time of analysis, the high reproducibility and sensitivity of the system, and the complete resolution of all compounds of interest make this method suitable for routine analysis. Furthermore, we have also developed a fast reversed-phase high-performance liquid chromatographic method for the complete separation of dimethylaminoazobenzene thiohydantoin (DABTH)-amino acids. The separation of the compounds under investigation is obtained, at room temperature, in less than 18 min using a reversed-phase Supelcosil LC-18 DB column, 3-micron particles, and also allows the complete separation of DABTH-Ile, DABTH-Leu, and DABTH-Norleu. The short time of analysis, together with the high reproducibility of the system and its sensitivity at picomole levels, make this method very suitable for the identification of DABTH-amino acids released during microsequencing studies of proteins and peptides with the dimethylaminoazobenzene isothiocyanate reagent. In addition, we have shown that it is possible to obtain complete separation of DABTH-amino acids also under isocratic conditions.(ABSTRACT TRUNCATED AT 400 WORDS)     

Identification and purification of the tricarboxylate carrier from rat liver mitochondria

The tricarboxylate carrier from rat liver mitochondria was solubilized with Triton X-100 and purified by chromatography on hydroxyapatite and celite. SDS-gel electrophoresis of the purified fraction showed a single polypeptide band with an apparent Mr of 30,000. When reconstituted into liposomes, the tricarboxylate transport protein catalyzed a 1,2,3-benzenetricarboxylate-sensitive citrate/citrate exchange. We obtained a 1070-fold purification with respect to the mitochondrial extract, the recovery was 22% and the protein yield 0.02%. The properties of the reconstituted carrier, i.e., requirement for a counteranion, substrate specificity and inhibitor sensitivity, were similar to those of the tricarboxylate transport system as characterized in intact mitochondria.     

Glutamate Dehydrogenase in Human Brain: Regional Distribution and Properties

Glutamate dehydrogenase (GDH) activity was studied in 17 regions of six human brains. Duration and conditions of the postmortem period did not affect enzyme activity. Specific activity ranged between 103 and 377 nmoles/min/mg protein at 25 degrees C and it was 10-fold higher than that found in leukocytes. Apart from exclusively white matter regions (corpus callosum and centrum ovale), there was a moderate regional distribution (2.5-fold variation), with highest values in the inferior olive and hypothalamus, and lowest in the cerebellum and lenticular nucleus. With alpha-ketoglutarate (alpha-KG), NADH, or NH4+ as variable substrate, the apparent Km values in human brain were Km alpha-KG = 1.9 X 10(-3) M, KmNADH = 0.21 X 10(-3) M, and KmNH4+ = 28 X 10(-3) M, and in leukocytes they were Km alpha-KG = 1.7 X 10(-3) M, KmNADH = 0.24 X 10(-3) M, and KmNH4+ = 28 X 10(-3) M. The effects of cofactors, inhibitor, and pH were similar in brain and leukocyte GDH.     

Interactions of prostaglandin E1, bradykinin and histamine and the increase of plasma fibrinogen in rats

Considering that tissue injury caused by laparotomy significantly increases the liver synthesis of plasma fibrinogen, and that PGE₁, bradykinini and histamine are released into the injured tissues, the effect of above mentioned inflammatory agents and of adrenal medulla on plasma fibrinogen levels in rats was studied. The subcutaneous administration of PGE₁, bradykinin or histamine does not modify plasma fibrinogen levels acting independently comparing with non-injected animals or injected with the drug vehicle. Bradykinin + histamine did not modify plasma fibrinogen levels either. However the administration of prostaglandin E₁ + bradykinin + histamine reproduced the increase of fibrinogen characteristics of laparotomy. This increase was partially but significantly inhibited in rats that had undergone bilateral removal of the adrenal medulla or administration of PGE₁ + bradykinin + histamine + bupivacaine (a local anesthetic), but it was not modified when the adrenal medullectomy was unilateral. It is concluded that plasma fibrinogen increase is obtained only when PGE₁ acts in presence of bradykinin or histamine and the adrenal medulla should be partially responsible for said increase.     

Comparative study on glucose-6-phosphate dehydrogenase from rabbit tissues

The activity of glucose-6-phosphate dehydrogenase (G6PD) was measured in bone marrow, spleen, lung, liver, kidney, adipose tissue, brain, heart, muscle, and in the erythroid cell line of rabbit. In tissues, the activity ranged from 6.87 to 0.09 U/g wet tissue, found in bone marrow and muscles, respectively, whereas in the erythroid cell line it ranged from 14.3 to 2.4 U/g cells for erythroblasts and erythrocytes, respectively. The electrophoretic patterns of the tissue crude extracts showed an identical set of three activity bands, and the immunotitration curves obtained with rat antirabbit erythrocyte G6PD antibodies shared the same equivalence point. The enzyme, purified to homogeneity from different tissues, showed no significant differences among the Km values for NADP and G6P. The results give a picture of the variability of the G6PD activity in rabbit tissues and suggest the presence of the same enzyme molecule in each tissue.     

Identification of Tuber magnatum Pico DNA markers by RAPD analysis

Summary Different species of truffle were studied in order to identify species-specific markers. The isolation of two Tuber magnatum Pico markers is reported. One of these could be used as a probe in dot blot hybridization, allowing the development of a rapid test able to identify Tuber magnatum species.     

A Chronobiological Approach to Circulating Levels of Renin, Angiotensin-Converting Enzyme, Aldosterone, ACTH, and Cortisol in Addison's Disease

This study deals with a chronobiological approach to the circadian rhythm of the renin-angiotensin-aldosterone system (RAAS) and the ACTH-cor-tisol axis (ACA) in patients with Addison's disease (PAD). The aim is to explore the mechanism(s) for which the circadian rhythmicity of the RAAS and ACA takes place. The study has shown that both the RAAS and ACA are devoid of a circadian rhythm in PAD. The lack of rhythmicity for renin and ACTH provides indirect evidence that their rhythmic secretion is in some way related to the circadian oscillation of aldosterone and cortisol. This implies a new concept: a positive feedback may be included among the mechanisms which chronoregulate the RAAS and ACA.