Betaine increases the butyrylcholinesterase activity in rat plasma.

The physiological function of butyrylcholinesterase (EC 3.1.1.8, BChE) is not clearly understood, but a role was suggested in the fat utilization process, resulting in positive correlation between plasma triglyceride (TG) levels and BChE activity. Consequently we tested the hypothesis that regular intake of betaine, a natural compound intervening in the liver TG metabolism could influence the BChE activity. The BChE activity was estimated spectrophotometrically in plasma of rats fed with betaine enriched standard (B) or high-fat diet (HFB). The results confirmed decreased TG plasma levels after betaine treatment independently on the type of diet (0.15+/-0.03 (B) vs. 0.27+/-0.08 (control) mmol/l; p=0.003 and 0.13+/-0.03 (HFB) vs. 0.27+/-0.08 (control) mmol/l; p=0.005). The BChE activity increased significantly with betaine administration, however the change was more distinct in the HFB group (0.84+/-0.34 (HFB) vs. 0.22+/-0.04 (control) O.D./min/mg; p<0.001 and 0.41+/-0.11 (B) vs. 0.22+/-0.04 (control) O.D./min/mg; p=0.001). In conclusion, betaine intake led to elevated BChE activity in plasma and this effect was potentiated by the HF diet. Since betaine is in general used as a supplement in the treatment of liver diseases accompanied by TG overload, its impact on the BChE activity in the role of the liver function marker should be taken into account.

The effect of respiratory diseases on serum lactate dehydrogenase and its isoenzyme patterns in calves.

In this study we examined the serum activity of lactate dehydrogenase (LDH) and its isoenzyme patterns in 28 calves of a lowland black spotted breed and its crossbreeds at the age of 2-6 months suffering from clinically noticeable manifested respiratory diseases--bronchopneumonia (BRD Group). As a control group we used 35 clinically healthy calves of the same age, breed and nutrition (Healthy Group). The sick calves did not show clinical signs or pathological lesions on other organ systems. The results found in sick calves showed a significantly higher total activity of LDH than in clinically healthy animals (P < 0.01). The mean activity of LDH was 2012 U/I in healthy calves and in calves with respiratory diseases 2529 U/1. The differences in all LDH isoenzyme patterns between both groups of animals were significant (P < 0.001) and in calves with respiratory diseases are characterized by a marked increase of the LDH 1 fraction and a decrease in the proportion of the other four LDH isoenzymes. Our results differ from those observed and presented in respiratory diseases in human medicine or in sheep. The explanation for the obtained results in calves and the determination of their diagnostic significance needs further studies and investigations using more animals with various severity of clinical signs and pathological changes, including analysis and determination of lactate dehydrogenase isoenzyme patterns in healthy and affected cattle lung tissue.

Influence of lipid imbalance on butyrylcholinesterase activity and biotransformation efficiency.

Butyrylcholinesterase (EC 3.1.1.8, BChE) is highly active in plasma, skin and lung, the tissues that first contact xenobiotics, supporting a role for BChE in detoxication of xenobiotics including medicaments. A possible involvement of BChE in lipid metabolism has been suggested. Elevated BChE activity in obese individuals correlates with some parameters of lipid metabolism including increased levels of triacylglycerols (TAG) and cholesterol. The aim of this study was to estimate the BChE activity in rats on subcellular and inter-organ levels under the conditions of untreated and treated primary hypertriacylglycerolemia with the TAG lowering agent fenofibrate. No changes in BChE activity were observed in obese animals. However fenofibrate administration led to significant increase of BChE activity in all examined tissues (plasma, liver, white adipose tissue). The impact of lipid metabolic imbalance on BChE biotransformation ability was tested by measuring the rate of hydrolysis of 0,1 to 8 mM concentrations of the antimicrobial agent N-(2-benzoyloxyethyl)-ethyldimethylammonium bromide (BCH2). The results revealed a complete shift in the BChE kinetics in all studied models. In animals with hypertriacylglycerolemia the Km value of liver BChE rised 4,6-fold, but the total enzyme efficiency expressed as Vmax/Km dropped 40% comparing to control. In contrast, in animals treated with fenofibrate the BChE efficiency increased in liver 1,6-fold. We conclude here that BChE detoxification capacity is essentially altered under conditions of disturbed lipid metabolism. Clinically, this knowledge could be important in a view of xenobiotic elimination, especially when routinely prescribed medicaments are concerned.

Enzymes cooperating in the hydrolyzing process of benzoylcholines: subcellular and inter-tissue comparison.

Within benzoylcholine biotransformation, butyrylcholinesterase (3.1.1.8, BuChE) appears to be the key enzyme in the hydrolyzing process. Except for BuChE in the process of benzoylcholine hydrolysis, carboxylesterase (3.1.1.1, CE) could play a role in the splitting of the ester bond. The aim of this work was to clarify the interaction between BuChE and CE in the hydrolyzing process of a homologic row of benzolycholines on subcellular and inter-tissue level. Two fractions, microsomes and cytosol of rabbit lung and liver were investigated. Participation of the enzyme activities was determined on the base of kinetic inhibitory studies, using eserine as cholinesterase inhibitor. Despite the fact that in all studied fractions of both organs BuChE and CE were confirmed, only in lung microsomes exclusive BuChE activity in benzoylcholine hydrolyzing process was observed, without substrate specifity. In the other fractions studied interaction of both enzymes were recorded, whereas the benzoylcholine structure played an important role. It seems that, the portion of CE depends predominantly on substrate structure and elevates with bulk of alcoholic part of benzoylcholines. Despite the same enzyme equipment in all tissue fractions studied, the affinity of hydrolyzing enzymes interestingly differs. This might be as a result of distinct subcellular pattern of CE activity localization in lung and liver.

[Antilipoxygenase activity and the trace elements content of Aloe vera in relation to the therapeutical effect]. / Antilipoxygenázová aktivita a obsah stopových prvkov Aloe vera vo vzt'ahu k terapeutickému úcinku.

Aloe vera is a rich source of many natural-health-promoting substances. The results of contemporary research on animal models indicate that the extracts have an antiinflammatory property. In this work the results of some in vitro experiments are shown: determination of the inhibitory effect of the Aloe vera extracts on the activity of partially purified lipoxygenase from the rat lung cytosol fraction, and quantitative determination of the trace elements presented in the extract (Mn, Fe, Cu, Zn) carried out by using the x-ray fluorescence analysis. The findings could explain the inhibitory effect (antilipoxygenase activity) of the Aloe vera extract in the acute inflammation process, expecially in the topical application for healing of minor burns and skin ulcers.

[Catalytic activity of butyrylcholinesterase in biodegradation of organic ammonium salts in vitro]. / Katalytická aktivita butyrylcholínesterázy v biodegradácii organických amóniových solí in vitro.

Organic ammonium salts of N-(2-benzoyloxyethyl)-alkyldimethylammonium bromide (BCHn-1) type are formed by the homological series Ar-COO(CH2)2-N+(CH3)2CnH2a + 1.Br-, whose structure contains a biodegradably labile ester bond, on the basis of which they rank among disinfectants and antiseptics of soft character. They are preferentially biotransformed hydrolytically to produce benzoic acid and substituted choline. The rapidity of enzymatic hydrolysis depends on the chemical structure (the length of the aliphatic chain on the ammonium nitrogen), it increases up to the number of 10 nitrogens of the aliphatic chain, and it rapidly decreases with further prolongation. The paper aimed to demonstrate the catalytic activity of butyrylcholinesterase on the enzymatic hydrolysis of selected organic ammonium salts in the medium of the microsomal fraction of the rat liver on the basis of inhibitory kinetic studies with physostigmine, a cholinesterase inhibitor. The product of enzymatic hydrolysis of BCHn-1, benzoic acid, was determined after extraction with chloroform from the acid medium by means of HPLC analysis with the use of the internal standard p-iodobenzoic acid at the wavelength of 228 nm. Kinetic parameters K(M) and VMAX were evaluated following Lineweaver-Burke using the method of linear regression analysis. The specific activity of butyrylcholinesterase (E.C.3.1.1.8) in the enzymatic hydrolytic process of BCHn-1 was significantly influenced by the presence of physostigmine, which was manifested by increased K(M), KI, and IC50 values in the investigated enzymatic process of selected substrates of the homological series BCHn-1, and by decreased VMAX and rate constants.

[Inter-organ variability in enzymatic hydrolysis of ammonium salts in rats in vitro]. / Medziorgánová variabilita enzýmovej hydrolýzy organických amóniových solí u potkana in vitro.

The study evaluated the rate and kinetics of enzymatic hydrolysis of N-(2-benzoyloxyethyl)-alkyl-dimethylammonium bromides, potential easily biodegradable disinfectants of soft character. The products of enzymatic hydrolysis of the substrates under study, catalysed by microsomal esterase, included substituted substrates choline and benzoic acid which, as a hydrolytic product, was essayed by HPLC. The effect of the length of the alkyl chain of the individual homologues on the rate of enzymatic hydrolysis and their affinity to microsomal esterase of the rat liver and lung in vitro was examined. The structural modification (varying length of the aliphatic chain on the ammonium nitrogen of these compounds) was found to significantly influence the kinetics of enzymatic hydrolysis of the esteric bond. From the viewpoint of the rate of enzymatic hydrolysis no significant inter-organ variability was observed: in the liver as well as the lung the dependence of the rate of enzymatic hydrolysis on the length of the aliphatic chain possesses the shape of a falling hyperbole with the maxima for BCH2 > BCH4 > BCH8 = BCH10. The specific activity of both esterases ranges within 0.2-3.5 nmol.min-1.mg-1. From the viewpoint of affinity, a marked inter-organ difference was manifested by 10 times higher affinity of the substrates to the lung microsomal esterase in comparison with the liver one. The effect of the length of the alkyl of the individual homologues on the affinity is of a non-linear character also in this case. In both organs, a certain correlation was found between the rate of enzymatic hydrolysis and affinity to microsomal esterases.

Lymphocyte blastogenesis to concanavalin A in dogs with localized demodicosis according to duration of clinical disease.

The aim of the study was to determine the degree of Con A induced lymphocyte blastogenesis in dogs with localized demodicosis (LD) within 1-3 and 6-8 weeks from appearance of the clinical signs. Ethidium bromide fluorescence assay was used for evaluation. In observation 9 clinically normal dogs, 6 dogs with LD a 4 dogs with generalized demodicosis (GD) were used. The results showed a statistically significant depression (P < 0.01) of blastogenesis in the LD dogs in comparison with that in the healthy dogs (Fig. 1). Responses to Con A were normal in dogs with LD in 1-3 weeks. However, a significantly depressed response to Con A (P < 0.025-0.001) was demonstrated in the LD dogs in 6-8 weeks (average 6.7 weeks) and it was comparable with that in the GD dogs with the duration of clinical disease on average for 8.7 weeks (Fig. 2; Tab. II). Thus, immunosuppression is not a necessary condition for dogs to develop spontaneous clinical LD and immunosuppression develops with the clinical signs of disease.

Evaluation of canine lymphocyte blastogenesis prior and after in vitro suppression by dog demodicosis serum using ethidium bromide fluorescence assay.

The mitogen induced blastogenic response of lymphocytes from normal dogs and dogs with generalized demodicosis (GD) was measured by the ethidium bromide (EB) fluorescence assay. Serum from GD dogs significantly suppressed the in vitro reactivity to Con A of peripheral blood lymphocytes (PBL) from normal dogs and GD dogs, however with a different percentage of suppression 40.6 and 81.2%, respectively. As a result, the degree of lymphocyte blastogenesis suppression in GD dogs did not parallel the immunosuppressive potency of their serum (Tab. IV). The data indicate that PBL obtained from GD dogs did not respond to Con A as well in the presence of serum from normal dogs as did PBL from normal dogs (Tab. IV). In one third of examined GD dogs a similar situation was described also by Hirsh et al. (1975). The basis for this cellular modified response is unknown. It does not appear that the age or the chronicity of the disease are related to this observation. Further studies are necessary to elucidate this relation. The GD dogs showed not only a significant depression of the lymphocyte response to Con A but also enhancement of the ability of unstimulated cells to proliferate was also observed (Tab. IV). Similar observation was reported by others (Barriga et al., 1992). The meaning of this is not clear at present. This finding is discussed in the light of proposed different effects of the parasite or the host's reactivity to the parasite on different subsets of lymphocytes. No significant difference of PBL responsiveness to Con A between healthy dogs with respect to the age (Tab. III) and the time of examination (compare results in Tabs. I and IV) was observed. Autologous serum showed a better responsiveness of normal canine lymphocytes to Con A than fetal calf serum (FCS). It is suggested that the use of FCS might lead to an erroneous judgement (Tab. I). Both lectins, Con A and PHA induced cell proliferation of healthy dogs in very similar amount (Tab. II). Our results indicated that EB fluorescence assay is a useful method for detection a respondence of canine lymphocyte blastogenesis.

[Metabolic N- and S-oxygenation of dokloxytepin in vitro]. / Metabolická N- a S-oxygenace dokloxytepinu in vitro.

Dokloxytepin in the medium of the induced monooxygenase system of the microsomal fraction of the liver of the rat, rabbit and mice is metabolized into three metabolites: two identical, i.e., N-oxide and 5-sulfoxide, and a third different one. In the rat and rabbit it is the hitherto unknown metabolite M1, and in the mouse S,N-dioxide of dokloxytepin. The metabolites were identified by thin-layer chromatography by comparing with synthetic standards.