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Biomedical subjects

E Roets

Publications and source records attributed to E Roets.

At least 91 records · Page 5Linked to original sources

Classification of newly calved cows into moderate and severe responders to experimentally induced Escherichia coli mastitis.

In the present study newly calved cows were tentatively classified as moderate and severe responders to experimentally induced Escherichia coli mastitis based upon the reactive oxygen species (ROS)-generating capacity of their blood neutrophils before infection. The groups differed in blood and milk composition prior to infection. This initial classification was supported by the corresponding variation in clinical symptoms and in the changes in milk production and composition measured during mastitis. Responses of newly calved cows to Esch. coli challenge varied from mild to severe symptoms of inflammation in infected glands and differed in the intensity of systemic disturbances and general illness. Losses in milk yield and compositional changes were most pronounced in inflamed glands and in severe responders. In inflamed glands milk yield and composition did not return to preinfection level in either moderate or severe responders. The yields of lactose, alpha-lactalbumin, casein and fat followed the same pattern as milk yield. It is concluded that the severe and long lasting systemic disturbances observed in severe responders can be ascribed to absorption of endotoxin from infected glands into circulation, indicating the important role of endotoxin in the pathology of coliform mastitis in periparturient cows. Evaluation of the ROS-generating capacity of blood neutrophils and blood and milk composition before infection might help to predict the cow's sensitivity to Esch. coli mastitis.

Animals↗

Effect of clenbuterol on beta-adrenoceptors and adenylate cyclase activity in smooth muscle and epithelium of the trachea of calves.

A potent beta-agonist (clenbuterol) was administered perorally to young calves for 50 days. After this period the animals were slaughtered and beta-adrenoceptor density, ligand affinity, and basal and stimulated adenylate cyclase activities were studied in smooth muscle and epithelium of the trachea. Although the density of lung beta-adrenoceptors was down regulated by clenbuterol, cAMP production remained constant (epithelium) or even increased (smooth muscle). Therefore desensitization of beta-adrenoceptors in the trachea was not observed. This might be a reason for the effectiveness of long-term treatment with beta-agonists.

Adenylyl Cyclases↗

Muscarinic receptor subtypes, beta-adrenoceptors and cAMP in the tracheal smooth muscle of conventional and double-muscled calves.

The total muscarinic (M1 + M2 + M3) and beta-adrenergic receptors in the tracheal smooth muscle of conventional and double-muscled calves were identified and characterized with the non-specific antagonists [3H]quinuclidinyl benzilate ([3H]QNB) and [3H]dihydroalprenolol ([3H]DHA) respectively. Although the quantity of beta-adrenoceptors in double-muscled calves was 25% lower (p < 0.05) than in conventional calves (Bmax = 327 +/- 89 fmol/mg protein), adenylate cyclase assays indicated that the basal adenylate cyclase activity and the (-)-isopropylnoradrenaline (ISO)- and sodium fluoride (NaF)-stimulated values were not significantly different between these calves. However, the density of muscarinic receptors in double-muscled calves was 40% higher (p < 0.01) than in conventional calves (Bmax = 2955 +/- 625 fmol/mg protein). Subtypes of muscarinic receptors were studied with [3H]telenzepine (M1-receptors), [3H]AF-DX 384 (M2-receptors) and [3H]4DAMP (M1 and M3-receptors). It was found that in both double-muscled and conventional calves about 40% of the receptors were of the M3-subtype, the remaining 60% being M2-receptors. From these results, it is suggested that inflammation of the respiratory tract in double-muscled calves may be complicated by an imbalance between the cholinergic bronchoconstrictor and the beta-adrenergic bronchodilator components of the autonomic nervous system.

Animals↗

Determination of metacycline and related substances by column liquid chromatography on poly(styrene-divinylbenzene).

Isocratic column liquid chromatography on poly(styrene-divinylbenzene) copolymer allowed complete separation of metacycline, 4-epimetacycline, oxytetracycline, doxycycline and 6-epidoxycycline. 2-Acetyl-2-decarboxamidometacycline was eluted on the tail of metacycline. The mobile phase was 2-methyl-2-propanol-0.2 M phosphate buffer (pH 9.0)-0.01 M sodium ethylenediaminetetraacetate (pH 9.0)-water (2.5:10:10:77.5, m/v/v/v). The flow-rate was 1.0 ml/min and detection was performed at 254 nm. Official standards were compared and a number of commercial bulk samples and specialties were analysed. 2-Acetyl-2 decarboxamidometacycline, 6-epidoxycycline and doxycycline were the main impurities, while 4-epimetacycline and oxytetracycline were minor impurities.

Anti-Bacterial Agents↗

Isolation of doxycycline, 6-epidoxycycline and 2-acetyl-2- decarboxamidometacycline from commercial metacycline by preparative column liquid chromatography on silica gel.

Isolation of doxycycline, 6-epidoxycycline and 2-acetyl-2-decaboxamidometacycline from commercial metacycline was achieved by preparative column liquid chromatography on silica gel, previously impregnated with edetate (EDTA). Careful control of the pH of EDTA allowed fine tuning of the separation. The mobile phases were composed of dichloromethane, methanol and .1 mM EDTA at pH 9.0 or 6.0. Structures were confirmed with nuclear magnetic resonance spectroscopy. The presence of doxycycline and its 6-epimer in commercial metacycline has not previously been described. The presence of the 2-acetyl derivative was not surprising since analogous 2-acetyl derivatives have been identified in other tetracyclines.

Chromatography, Liquid↗

Quantitative analysis of erythromycin by reversed-phase liquid chromatography using column-switching.

A column-switching technique is described for LC of erythromycin. The method allows, in about 1 h, the separation of erythromycin A from all its known potential impurities, except erythromycin D, which is a minor impurity. The switching technique combines two columns (7.5 cm x 4.6 mm and 25.0 cm x 4.6 mm) both packed with RSil C 18 LL 10 microns. The mobile phase is acetonitrile-tetrabutylammonium sulphate (0.2 M, pH 6.0)-ammonium phosphate buffer (0.2 M, pH 6.0)-water (24:5:5:66, v/v/v/v). Temperature was 35 degrees C, flow rate was 1.5 ml min-1, detection was by UV at 210 nm. Results for a number of commercial samples of various origin are reported.

Acetonitriles↗

Assay and purity control of metacycline by thin-layer chromatography combined with UV and fluorescence densitometry--a comparison with liquid chromatography.

A thin-layer chromatographic (TLC) method involving UV and fluorescence densitometry is described for the assay and purity control of metacycline. With a mobile phase dichloromethane-methanol-water (58:35:7, v/v/v) and a silica gel thin-layer, previously sprayed with 10% sodium edetate solution adjusted to pH 9.0, all the potential impurities of metacycline were well separated from the main component and from each other. Results obtained with UV densitometry (TLC-UV) and fluorescence densitometry (TLC-F) were compared with those obtained by a liquid chromatography (LC) method using a poly(styrene-divinylbenzene) stationary phase. The correlation coefficients (r) for TLC-UV and LC or TLC-F and LC were better than 0.9999. For TLC-UV the relative standard deviation (RSD) for the assay of the main component was less than 2%, for TLC-F less than 3.0% and for LC less than 1.0%.

Anti-Bacterial Agents↗

Separation of keto-enol tautomers of chlortetracycline and 4-epichlortetracycline by liquid chromatography on poly(styrene-divinylbenzene)copolymer.

Keto and enol tautomers of chlortetracycline and 4-epichlortetracycline were successfully separated by low temperature high-performance liquid chromatography on poly(styrene-divinylbenzene)copolymer. The keto-enol tautomerism occurs between C-11a and C-12. The elucidation of the chemical structure of the tautomers was realized with the help of on-line and off-line UV-spectrophotometry and of 13C and 1H NMR spectrometry. The kinetics of the equilibrium reaction were investigated.

Buffers↗

Assay and purity control of oxytetracycline and doxycycline by thin-layer chromatography--a comparison with liquid chromatography.

A thin-layer chromatographic (TLC) method using densitometry is described for the assay and purity control of oxytetracycline and doxycycline. With a mobile phase of dichloromethane-methanol-water (59:35:6, v/v/v) and a silica gel thin-layer, previously sprayed with 10% sodium edetate solution adjusted to pH 9.0, all the potential impurities of oxytetracycline or doxycycline are well separated from the main components and from each other. Results obtained with TLC are compared with those obtained by previously established liquid chromatography (LC) methods using poly(styrene-divinylbenzene) stationary phases. A good correlation was obtained (r greater than 0.9999). For TLC the relative standard deviation (RSD) for the assay of the main component was less than 2%, for LC the RSD was less than 1%.

Chromatography, Liquid↗

Determination of beta-adrenoceptors and cAMP in muscle from normal and splayleg Belgian Landrace pigs.

beta-Adrenoceptors were identified and characterized by [3H]dihydroalprenolol ([ 3H]DHA) binding experiments in muscle membrane preparations from piglets. The [3H]DHA binding was rapid, reversible and stereoselective. Catecholamines competed for specific binding with a rank order of potency (-)-isopropylnoradrenaline greater than (-)-epinephrine much greater than (-)-norepinephrine, indicating a beta 2-subtype of adrenoceptor. Saturation binding experiments with [3H]DHA showed no significant difference for either the number of binding sites or the equilibrium dissociation constants in normal and splayleg pigs. Adenylate cyclase assays indicated that the basal adenylate cyclase activity and the prostaglandin E1 (PGE1)-, (-)-isopropylnoradrenaline (ISO)-, 5'-guanylyl-imidodiphosphate (GppNHp)- and sodium fluoride (NaF)-stimulated values were not significantly different in normal and splayleg pigs. In both groups, PGE1 did not affect basal activity, whereas ISO and GppNHp stimulated adenylate cyclase activity significantly (p less than 0.001) to about 40% above basal level. NaF induced a significant (p less than 0.001) increase of cAMP in normal and splayleg pigs amounting to 48% and 61% respectively. Significant correlations between absolute adenylate cyclase responses to ISO (r = 0.83***), NaF (r = 0.72**) and GppNHp (r = 0.61*) and basal activity in the splayleg pigs were the most striking findings. In contrast, these correlations could not be detected in the normal pigs. Whether or not this observation reflects an alteration in the signal transduction system needs to be further investigated. To the best of our knowledge this is the first biochemical study which relates an altered beta-adrenoceptor function and porcine splayleg.

Adenylyl Cyclases↗

Quantitative analysis of chlortetracycline and related substances by high-performance liquid chromatography.

Isocratic high-performance liquid chromatography on Zorbax C8 7 microns allows quantitative determination of chlortetracycline, 4-epichlortetracycline, tetracycline, demethylchlortetracycline and isochlortetracycline using a mobile phase containing dimethylsulphoxide, 1 M perchloric acid and water (35:5:60). The minor impurities anhydrochlortetracycline and 4-epianhydrochlortetracycline, which are more strongly retained can be determined using a second isocratic system with a mobile phase containing more organic modifier. The method has been used for the comparison of official standards and for the analysis of a number of commercial samples.

Chlortetracycline↗

Quantitative analysis of demeclocycline by high-performance liquid chromatography.

A high-performance liquid chromatographic (HPLC) method suitable for the quality control of demeclocycline is described. The stationary phase is a poly(styrene-divinylbenzene) copolymer, kept at 60 degrees C. The mobile phase comprises 2-methyl-2-propanol-0.2 M potassium phosphate buffer (pH 9.0)-0.02 M tetrabutylammonium hydrogen sulphate (pH 9.0)-0.01 M sodium edetate (pH 9.0)-water (8:10:15:10:57, m/v/v/v/v). The flow rate is 1 ml min-1 and detection is performed at 254 nm. Official standards are compared and results for the analysis of a number of commercial bulk samples and preparations are presented. 4-Epidemeclocycline and demethyltetracycline are the main impurities. 4-Epidemethyltetracycline and 2-acetyl-2-decarboxamido-demeclocycline can also be present.

Capsules↗

Changes in N-acetyl-beta-D-glucosaminidase activities in relation to other milk components throughout normal lactation in the guinea pig.

Changes in milk production, NAGase activities, Na, Cl, K, lactose, total N, and fat were followed throughout normal lactation in 26 healthy guinea pigs. Appropriate microassays were used and all determinations were performed on the same small skim milk sample (microhematocrit tube). A very sensitive and rapid spectrofluorimetric determination of NAGase in 10-microliters skim milk samples was evaluated. Four days after parturition, daily milk production reached a maximum of about 20 ml. Thereafter milk yield decreased progressively to about 7 ml on d 15. During the entire course of lactation, a progressive increase in NAGase, total N, fat, Na, and Cl was observed. However, K and lactose decreased. Highly significant positive correlations between NAGase and Na, Cl, fat, and total N were evident, whereas correlations were negative between NAGase and K, lactose, and milk production. These changes can be explained on basis of alterations that occur at the level of the blood-milk barrier. The short lactation period of the guinea pig, the easy handling and milking practices, together with the available sensitive micromethods can lead to the establishment of a convenient guinea pig mastitis model.

Acetylglucosaminidase↗

Relationship between numbers of alpha 2- and beta 2-adrenoceptors in teat tissue and blood cells and milkability of primiparous cows.

Milking characteristics were measured on 14 cows at morning milkings. After collection of blood, cows were slaughtered and teats were removed. beta 2-Adrenoceptors on teat tissue and lymphocyte membranes were identified by binding of [3H]dihydroalprenolol. For the determination of alpha 2-adrenoceptors on teat tissue and blood platelet membranes, [3H]rauwolscine was used. In teat tissue and blood cells, except for the amount of milk collected during the 3rd min, all other milking parameters were highly correlated with the ratio of beta 2-adrenoceptors to alpha 2-adrenoceptors. Good milkability appears to be associated with low ratio of beta 2-adrenoceptors to alpha 2-adrenoceptors densities in teat tissue, and a high ratio of beta 2- to alpha 2-adrenoceptors densities in blood cells. We postulated earlier that a low ratio of beta 2- to alpha 2-adrenoceptors in teat tissue of fast milking cows probably reflects changes of mainly prejunctional adrenoceptors. From our results on blood cells, we are now hypothesizing that in teat tissue prejunctional and extra junctional alpha 2- and beta 2-adrenoceptors might have been identified together, whereas alpha 2- and beta 2-adrenoceptors identified on blood platelets and lymphocytes, respectively, are reflecting by preference extra junctional adrenoceptors are activated primarily by circulating epinephrine. The high ratio of beta 2- to alpha 2-adrenoceptors observed in blood cells of fast milking cows might indicate a decreased vasomotor tone in the teat and an increased dilatation of the teat sphincter in these animals.

Animals↗

Thin-layer chromatographic study of the metabolites of erythromycins in the Wistar rat.

The metabolites of erythromycin A, anhydroerythromycin A, N-demethylerythromycin A and erythromycin B in the Wistar rat were studied by thin-layer chromatography. In some experiments germ-free rats, rats with a cannulated bile duct and a gastrectomized rat were used. The erythromycins examined were shown to undergo two principal changes, N-demethylation and acid-catalysed degradation. It was demonstrated that the stomach and the liver are not the sole sites of acid degradation and demethylation of erythromycins, respectively. Erythromycin A gives three principal metabolites, anhydroerythromycin A, anhydro-N-demethylerythromycin A and N-demethylerythromycin A, and erythromycin A enol ether and N-demethylerythromycin A enol ether are present to a minor extent. 5-O-Desosaminylerythronolide A was also identified, suggesting the presence of an erythromycin glycosidase.

Animals↗

Optimization of the separation of erythromycin and related substances by high-performance liquid chromatography.

An improved high-performance liquid chromatographic method for analysis of erythromycin is described. The separation can be performed under mild conditions of pH (6.5) and temperature (35 degrees C) on C8 and C18 silica-based reversed-phase materials of different origins. The mobile phase, with a flow-rate of 1.5 ml/min, contained various amounts of acetonitrile (25-40%, v/v), 5% (v/v) 0.2 M ammonium phosphate buffer pH 6.5, 20% (v/v) 0.2 M tetramethylammonium phosphate and water. UV detection at 215 nm allows quantitation of erythromycins A, B and C, N-demethylerythromycin A, erythromycin A enol ether and anhydroerythromycin A. The column history plays a major role, older columns often giving better separations.

Chromatography, High Pressure Liquid↗

Quantitative analysis of oxytetracycline and related substances by high-performance liquid chromatography.

Isocratic high-performance liquid chromatography on PLRP-S 8-microns poly(styrene-divinylbenzene) copolymer allows complete separation of oxytetracycline, 4-epioxytetracycline, tetracycline, anhydrooxytetracycline, alpha- and beta-apooxytetracycline. The mobile phase was tert.-butanol-0.2 M phosphate buffer pH 8.0-0.02 M tetrabutylammonium sulphate pH 8-0.0001 M sodium ethylenediaminetetraacetate pH 8.0-water (5.9:10:5:10:78.1, m/v/v/v/v). With this isocratic method, 2-acetyl-2-decarboxamidooxytetracycline is only partly resolved from oxytetracycline. The separation and the detection limits can be improved by the use of gradient elution. Gradient elution was used for the comparison of official standards and for the analysis of a number of commercial samples, and to monitor the stability of oxytetracycline hydrochloride during storage in the solid state for about 6 years at various temperatures.

Chemical Phenomena↗