Horse Plasma Research Articles

Equine angiotensin converting enzyme: a zinc metalloenzyme.

1. Angiotensin I converting enzyme from horse plasma has been extensively purified and shown to be homogeneous by disc-gel electrophoresis. 2. The metal ion involved in the catalytic reaction of the enzyme has been identified for the first time as zinc by atomic absorption spectrometry. 3. A number of other physicochemical properties of the enzyme are described and compared with results obtained by other investigators. The molecular weight was determined by gel filtration to be 113 000 daltons. The pH maximum was found to be 7-4. The chloride activation of the enzyme appears to act by facilitation of substrate binding to the enzyme. 4. By use of enzyme inhibitors, tyrosine has been implicated as a functional residue at the active site of the enzyme. 5. The enzyme shows a fairly high degree of specificity towards its substrates.

Hydrolysis of histones by horse urinary kallikreins.

Horse urinary kallikrein was shown to hydrolyze histones from calf thymus and chicken nuclei erythrocytes. The hydrolysis is inhibited by benzamidine and hyposulfite but not by soy-bean trypsin inhibitor; horse plasma kallikrein also produces this hydrolysis.

A Fluorometric Enzyme Inhibition Detector for Carbamate Pesticide Analysis by High Speed Liquid Chromatography

Abstract A fluorometric enzyme inhibition detector has been developed for the detection of carbamate pesticides after separation using high speed liquid chromatography. Housefly head cholinesterase was found to have the greatest sensitivity toward the carbamates, together with horse plasma and bovine erythrocyte cholinesterases. The substrate N-methyl indoxyl acetate is used for monitoring of enzyme activity.

The pharmacology of anaphylaxis in the chicken intestine.

1 The Schultz-Dale phenomenon has been demonstrated in several circular smooth muscle strips of oesophagus, crop, duodenum, jejunum and ileum taken from young and adult domestic fowl sensitized actively to crystalline bovine albumin or horse plasma. 2 The ileal strips contract to acetylcholine, histamine, 5-hydroxytryptamine (5-HT), prostaglandins E1, E2, F2alpha, bradykinin and bovine slow reacting substance of anaphylaxis (SRS-A). Marked seasonal and individual variations in the responsiveness of gut tissues to these exogenous agonists were noted. 3 Antagonism of contractions to histamine by mepyramine suggests the existence of H1-histamine receptors in chicken ileum. Blockade of 5-HT-induced contractions by methysergide shows the preponderance of 'D'-musculotropic tryptamine receptors. 4 Failure of selective receptor antagonists of acetylcholine, histamine and 5-HT to modify the Schultz-Dale reaction suggests the nonparticipation of aminergic mechanisms in this reaction. 5 Partial to complete blockade of the Schultz-Dale reaction by a prostaglandin receptor antagonist (polyphloretin phosphate, PPP); prostaglandin synthetase inhibitors (sodium meclofenamate and phenylbutazone); inhibitors of synthesis and release of histamine and SRS-A (PR-D-92-EA, M & B 22948, diethylcarbamazine citrate, and PPP) and an inhibitor of proteinases (aprotinin) strongly suggests the involvement of vasoactive lipids and polypeptides in the anaphylactic response of chicken ileum to specific antigen.

Pharmacological experiments as a basis for the administration of digoxin in the horse

It is shown that the concentration of ouabain necessary for 50 per cent inhibition of the Na+K activated membrane ATPase of red cells is similar in man and horse. This is taken to indicate that the two species have similar sensitivity towards cardiac glycosides in general. In five adult healthy horses plasma digoxin concentration was measured with a radioimmunoassay technique after a single intravenous injection of 1 mg/100 kg body weight digoxin. The half time of elimination was 23 h and the apparent volume of distribution 7.3 litres/kg. An approximate estimate of plasma protein binding of digoxin was obtained by measuring digoxin with a fluorimetric assay in the ultrafiltrate from horse plasma containing 2-20 mug/ml. At concentrations below 10(-5)(g/ml 20 to 40 per cent of digoxin present in horse plasma is bound to protein. With this information and by using effective digoxin concentration measured in humans an average daily maintenance dose of 0-5-0-75 mg/100 kg body weight was calculated which may serve as a guideline in the treatment of congestive heart failure with digoxin in the horse.

Mechanism of inhibition in vitro of mammalian acetylcholinesterase and cholinesterase in solutions of O, O-dimethyl 2,2,2-trichloro-1-hydroxyethyl phosphonate (trichlorphon)

The rate of decomposition of trichlorphon into DDVP was measured polarographically at pH 7.4. The first order rate constants of decomposition at 25° are 7.27 × 10 −4 and 6.05 × 10 −4 min −1 for trichlorphon concentrations of 0.150 and 15.0 mM respectively; at 37° the corresponding rate constants are 53.1 × 10 −4 and 37.1 × 10 −4 min −1. The rate of decomposition of trichlorphon was also calculated from the kinetics of inhibition of acetylcholinesterase (EC 3.1.1.7) and cholinesterase (EC 3.1.1.8) in trichlorphon solutions at 25° and 37° (pH 7.4). The following enzyme sources were used: bovine erythrocytes and rat brain acetylcholinesterase, and human, horse and rat plasma cholinesterase. The rate of decomposition of trichlorphon was calculated by assuming that only DDVP formed from trichlorphon is the enzyme inhibitor, while trichlorphon itself does not act as an inhibitor. The calculated rate constants for the decomposition of trichlorphon are lower or just within the range of the rate constants obtained by the polarographic method. This agreement was taken as kinetic evidence that trichlorphon is not an inhibitor of mammalian cholinesterases. The effect of pH on enzyme inhibition supports this conclusion. The rate of inhibition of bovine erythrocyte acetylcholinesterase by DDVP is the same at pH 7.4 and pH 6.0 (37°). However, the rate of enzyme inhibition in trichlorphon solutions is 30 times faster at pH 7.4 than at pH 6.0, and this agrees with the greater stability of trichlorphon at the lower pH value. The rate of spontaneous reactivation of the enzyme was measured (37°, pH 7.4) after inhibition in trichlorphon solutions of acetylcholinesterase (human and bovine erythrocytes) and cholinesterase (human plasma). For all three enzyme preparations, the rate of spontaneous reactivation was the same as that obtained after inhibition by DDVP. All results point to the conclusion that trichlorphon in vitro is not an inhibitor of mammalian cholinesterases.

Cholinesterase inhibition by phenothiazine and nonphenothiazine antihistaminics: Analysis of its postulated role in synergizing organophosphate toxicity

Several antihistaminic compounds inhibit in vitro pseudocholinesterase (ChE) from several sources (human, rat, or horse plasma), pI50 ranging from 5.0 to 6.0. This inhibition is competitive with acetylcholine (ACh) and is reversible. Acetylcholinesterase (AChE) from rat brain was nonsensitive to antihistaminics even at a concentration of 4 m m. Antihistaminics do not compete with either organophosphates or quaternary ammonium compounds in inhibiting ChE. Inhibition of ChE by antihistaminics is more pronounced at alkaline pH values than at acidic or neutral pH. Administration of promethazine ip to rats at a dose of 50 mg/kg resulted in a 90% inhibition of ChE 1 hr later, while the AChE was totally nonsensitive to the inhibitor even when the dose of the antihistamine was doubled. Results were analyzed in relation to the postulated role of ChE inhibition in antihistaminic potentiation of organophosphorus intoxication.

Reactivity of isolated bovine mesenteric and hepatic veins to vasoactive agents and specific antigen

In an attempt to investigate the possible contribution of the mesenteric and hepatic vasculature to the haemodynamic changes occuring during bovine systemic anaphylaxis, the pharmacological reactivity of isolated bovine mesenteric and hepatic veins was studied. Both mesenteric and hepatic veins contracted in the presence of histamine, 5-hydroxytryptamine (5HT), acetylcholine, dopamine and noradrenaline, and relaxed in the presence of isoprenaline. The mesenteric veins contracted in the presence of concentrations of isoprenaline greater than 1 μg/ml. By the use of antagonists, the receptor populations of bovine mesenteric and hepatic veins were shown to be similar to those of many other vascular preparations. Hepatic veins isolated from calves previously sensitised to horse serum contracted forcefully in the presence of horse plasma (the Schultz-Dale phenomenon), whereas mesenteric veins isolated from the same animals did not. Hepatic veins may be implicated in the haemodynamics of bovine systemic anaphylaxis.

A steady-state kinetic model of butyrylcholinesterase from horse plasma.

The steady-state kinetics of the butyrylcholinesterase-catalysed hydrolysis of butyrylthiocholine and thiophenyl acetate were shown to deviate from Michaelis-Menten kinetics. The ;best' empirical rate law was selected by fitting different rate equations to the experimental data by non-linear regression methods. The results were analysed in view of two alternative interpretations: (1) the reaction is catalysed by a mixture of enzymes, or (2) the activity is due to a single enzyme displaying deviations from Michaelis-Menten kinetics. It was concluded that the second alternative applies, and this conclusion was further supported by experiments involving simultaneous hydrolysis of alternative thiol ester substrates (butyrylthiocholine/thiophenyl acetate) as well as alternative thiol ester and oxygen ester substrates (butyrylthiocholine/phenyl acetate; thiophenyl acetate/butyrylcholine; acetylthiocholine/phenyl acetate). On the basis of the conclusion that a single enzyme is responsible for the activity, a molecular model is proposed. This model involves an acylated enzyme, and implies binding to the enzyme of one acyl group and one ester molecule, but not two ester molecules at the same time. Thus butyrylcholinesterase, which is structurally a tetramer, behaves functionally as a co-operative dimer, an interpretation in accordance with available data from active-site titrations.

Comparative study of plasma protein binding of diphenylhydantoin

1. Plasma protein binding of 14C-labelled diphenylhydantoin (DPH) was determined in vitro by equilibrium dialysis (37° C.) and liquid scintillation spectroscopy. 2. Binding was studied in the goat, sheep, ox, horse, pony, swine, dog, cat, rabbit, opossum, and human plasmas. Extensive binding was observed in all species. 3. The percentage DPH bound decreased biphasically with increase in drug concentra-(2.4–40.0 μg. per ml.). 4. Highly significant interspecies variation was found in the unbound fraction of DPH. 5. Nephrectomy caused a significant decrease in DPH binding in goats. 6. Lower binding capacity may be due to a qualitative change in drug-binding proteins.

Plasma Protein Binding of Digitoxin and Digoxin in Several Mammalian Species

The extents of protein binding of 3H-digitoxin and 3H-digoxin in plasma of different species were determined in vitro by the equilibrium dialysis technique and liquid scintillation counting. Independence of drug concentration and the percentage of drug that was protein bound was observed for both digitoxin (0.01 to 1.00 μg/ml) and digoxin (0.0025 to 1.00 μg/ml). The extents of binding of 3H-digitoxin (50 ng/ml) and 3H-digoxin (10 ng/ml) were measured in goat, sheep, ox, horse, pony, swine, dog, cat, human, monkey, rat, rabbit and opossum plasmas. Digitoxin was more than 80% bound in all species except opossum (72%). There was a significant difference in extent of digitoxin binding among species (F-test, p

Improved Measurement of Corticosteroids in Plasma and Urine by Competitive Protein-Binding Radioassay

Abstract In the assay of corticosteroids in plasma and urine by competitive protein binding, the use of horse serum or plasma as a source of assay protein gives better sensitivity and somewhat better specificity for cortisol than do previously described procedures. Five to 10 µl of plasma unknown or 50 to 100 µl of urine unknown can be used. Precision for the standard curve is 0.12 ng in the range 0-2.00 ng, and accuracy, measured as the amount of added cortisol recovered from plasma, is about 91%. Because of the improved specificity, the need for preliminary purification by chromatography is decreased for some purposes. The same corticosteroid-binding globulin and standard curve can be used for assaying corticosteroids in urine or plasma. The procedure may be useful for unusually small samples or if discriminations within the lower range of physiological concentrations are needed, as in work with infants and neonates or in the study of small adrenal secretory pulses, such as those occurring at the nadir of the circadian cycle.

Hemopoietin-initiated changes in differential retransplantability of mouse femoral marrow-derived colony-forming units (CFU).

SummaryThe spleen colony system was used to test in vivo effects of hemopoietins on the total and differential exocolonizing potential of A/J mouse marrow colony-forming units (CFU) transplanted serially into primary and secondary, supralethally irradiated, isogeneic hosts. The hemopoietins used were GPF (calf renal granulopoietic factor) and ESF (erythrocyte stimulating factor-rich anemic horse plasma).As measured by total and differential primary host spleen colony counts, GPF and ESF evoked a time-dependent increase in a pool of committed donor CFU. These CFU were transplantable from primary into secondary hosts without losing their hemopoietininitiated, cell line-specific commitment.The f-fraction remained within normal limits (0.112-0.125) in all experimental groups when primary host spleen cells were harvested at 24 hr and transplanted into secondary hosts. When the donor CFU were allowed to reside and replicate in the primary host spleen for 8 days before retransplantation into the secondary hosts,...

In vitro commitment by hemopoietins of murine marrow exocolony forming units (CFU)? Preliminary report. 1.

The spleen colony technique was used to test the in vitro effects of GPF (calf renal granulopoietic factor) and ESF (erythrocyte stimulating factor-rich anemic horse plasma) on the survival and differential colonizing potential of A/J mouse marrow cells in suspension culture. The quantitative survival of nucleated cells and transplantable colony forming units remained unaffected by the addition of either hemopoietin. Beginning at 1 hr and increasing progressively, the erythrocytic:granulocytic ratios of cultured marrow-derived spleen colonies shifted markedly in the direction of the hemopoietin-dependent cell line. Alternate theoretical explanations for this phenomenon are discussed.

The determination of phenylbutazone in horse plasma.

The determination of phenylbutazone in horse plasma.

Kallidin (lysylbradykinin), the kinin formed from horse plasma by horse urinary kallikrein

Horse urinary kallikrein when incubated with horse plasma formed kallidin (lysylbradykinin) from the kininogens in the plasma. Horse plasma, like human plasma, was found to contain an aminopeptidase capable of converting kallidin to bradykinin. No evidence, however, could be found that the plasma contained an aminopeptidase capable of converting Met-Lys-bradykinin to kallidin, thus eliminating the possibility that the kallikrein had released Met-Lys-bradykinin which was converted to kallidin during the 1–5 min incubations. The method used for identification of the kinins is rapid, gives a good recovery and requires small amounts of plasma and enzyme.

Water and electrolyte metabolism in skeletal muscle of exercising rats.

ArticleWater and electrolyte metabolism in skeletal muscle of exercising rats.P Kõrge, and A ViruP Kõrge, and A ViruPublished Online:01 Jul 1971https://doi.org/10.1152/jappl.1971.31.1.1MoreSectionsPDF (932 KB)Download PDF ToolsExport citationAdd to favoritesGet permissionsTrack citations ShareShare onFacebookTwitterLinkedInWeChat Back to Top Next Download PDF FiguresReferencesRelatedInformation Cited ByEffect of extracellular osmolality on metabolism in contracting mammalian skeletal muscle in vitroApplied Physiology, Nutrition, and Metabolism, Vol. 34, No. 6Elemental composition of muscle at rest and potassium levels in muscle, plasma and sweat of horses exercising at 20°C and 35°C10 June 2010 | Equine Veterinary Journal, Vol. 28, No. S22Hematological and biochemical changes during a short triathlon competition in novice triathletesEuropean Journal of Applied Physiology and Occupational Physiology, Vol. 61, No. 1-2Food deprivation decreases the exertion-induced acid hydrolase response in mouse skeletal muscleEuropean Journal of Applied Physiology and Occupational Physiology, Vol. 57, No. 2Exhaustive physical exercise and acid hydrolase activity in mouse skeletal muscleHistochemistry, Vol. 57, No. 3Work-induced potassium changes in skeletal muscle and effluent venous blood assessed by liquid ion-exchanger microelectrodesPflügers Archiv European Journal of Physiology, Vol. 362, No. 1Analysis of the initial relaxation of mammalian skeletal muscles in ontogenyBulletin of Experimental Biology and Medicine, Vol. 76, No. 3Work-Induced potassium changes in muscle venous effluent blood measured by ion-specific electrodesPflügers Archiv European Journal of Physiology, Vol. 338, No. 2Effects of exercise and cold acclimation on the ventricular and skeletal muscles of white mice (Mus musculus)—I. Succinic dehydrogenase activityComparative Biochemistry and Physiology Part A: Physiology, Vol. 43, No. 4Work-induced increase of extracellular potassium concentration in muscle measured by ion-specific electrodesBrain Research, Vol. 40, No. 2 More from this issue > Volume 31Issue 1July 1971Pages 1-4 https://doi.org/10.1152/jappl.1971.31.1.1PubMed5556956History Published online 1 July 1971 Published in print 1 July 1971 Metrics

Studies on the interaction between inhibitors of drug metabolism and horse plasma cholinesterase

Several inhibitors of drug metabolism were found to inhibit horse plasma cholinesterase activity at concentrations ranging from 0·3 μM to 20 μM. A detailed study of the inhibition was made using 2-diethylaminoethyl 2,2-diphenylvalerate (SKF 525A); 2,4 dichloro-6-phenylphenoxyethyldiethylamine (Lilly 18947) and N-(γ-dimethyl- aminopropyl-iminodibenzyl) (imipramine). The cholinesterase inhibition produced by these three compounds was very fast, competitive and reversible. Only SKF 525A was able to delay the irreversible inhibition of the enzyme by dimethyl-dichlorovinyl-phosphate, indicating that it is probably acting at the esteratic site. The inhibition by these compounds increases from pH 5·5 to pH 7·5–8·0, then decreases. Since the inhibitors are mainly in the basic form at pH 7·5, the results were interpreted as showing that the active portion of the inhibitor is located at the basic nitrogen atom and that this group interacts on a chemical group of the enzyme having a p K a about 8·6–9·0. In agreement with this assumption, changes in basicity of the nitrogen atom of the inhibitors explained very well the differences in inhibitory power between pairs of analog compounds. This hypothesis also explains why bovine erythrocyte acetylcholinesterase exhibited smaller affinity for inhibitors than the horse plasma enzyme, in spite of the similarities in the inhibition at different pH values. Changes in entropy for the inhibition of cholinesterase by Lilly 18947 were also in accordance with this hypothesis.

Carbohydrate metabolism and electrolyte changes in human muscle tissue during heavy work.

Carbohydrate metabolism and electrolyte changes in human muscle tissue during heavy work.

Glass-activated plasma kallikrein

Plasma kallikrein from horse plasma was purified by adsorption on glass-surface followed by chromatography on DEAE-cellulose. It is shown that kallikrein is activated, simultaneously adsorbed and afterwards eluted from glass surface in the same conditions as described by other authors to separate Hageman factor from plasma. Evidence is presented suggesting that glass-activated and acid-activated kallikrein are the same substance.