Purification and characterization of hepatic triacylglycerol lipase isolated from rainbow trout,Oncorhynchus mykiss

Trialcylglycerol (TG) lipase was isolated and partially purified from rainbow trout liver. Triacylglycerol lipase activity was assayed by measuring¹⁴C-oleic acid release from¹⁴C-triolein.¹⁴C-oleic acid release was linear for up to two hours. Optimal activity occurred at pH 7.0 and 15°C. Most of the lipase activity was recovered in the cytosolic fraction. A 27,000-fold purification was achieved after Sepharose (Bio-gel A 0.5 M, 200–400 mesh) chromatography of a resuspended 20% ammonium sulfate fraction. The molecular weight of the trout hepatic lipase as determined by size-exclusion chromatography and by SDS-polyacrylamide gel electrophoresis was 40–43 kD. Lipase-mediated hydrolysis of TG resulted in the production of diacylglycerols, monoacylglycerols, and fatty acids. Kinetic analysis indicated that V_max=0.016 nmol/h/mg protein and that K_m=0.28 mM triolein. Lipolytic activity was enhanced in the presence of cAMP/ATP-Mg²⁺. These results suggest that the liver of trout possesses a neutral TG lipase that is responsible for mobilizing stored TG and is catalytically activated by phosphorylation.A part of this work was presented at the Annual Meeting of the American Society of Zoologists, December 26–30, 1990, San Antonio, TX.     

Plasma cholesterol and lipoprotein concentrations in the dog: The effects of age, breed, gender and endocrine disease

A combined ultracentrifugation and precipitation technique was used to quantify the plasma lipoprotein concentrations of control dogs (n=33) and dogs with diabetes mellitus (n=11), hyper-adrenocorticism (n=14), hypothyroidism (n=10) and obesity (n=20). In addition, the effect of breed type, age and gender on the lipoprotein phenotype was assessed. Breed type and age were found to have no effect upon cholesterol and lipoprotein concentrations but the high density lipoprotein cholesterol (HDL-C) concentration was greater in intact females than intact males. Cholesterol concentrations were significantly higher than those of the control group in dogs with diabetes mellitus (P<0·01), hyper-adrenocorticism (P<0·01) and hypothyroidism (P<0·001). In dogs with diabetes mellitus this was due to increased concentrations of very low density lipoprotein cholesterol (VLDL-C) (P<0·01) and HDL-C (P<0·05). The concentrations of low density lipoprotein cholesterol (LDL-C) (P<0·05) were significantly increased in dogs with hyperadrenocorticism, while in the hypothyroid dogs, VLDL-C (P<0·05), LDL-C (P<0·001) and HDL-C (P<0·05) were significantly higher than the control group. The cholesterol and lipoprotein concentrations in the obese population were not significantly different from control dogs.     

Surgical treatment of lameness.

Digital diseases are commonly seen in cattle. Cattle affected by digital disorders do not always respond to conservative therapy and require surgery. Surgical procedures for the common digital disorders are described, with emphasis on the different approaches to the distal interphalangeal joint.     

Quantitation of adenine nucleotides in equine colonic mucosal tissue using high performance liquid chromatography.

The objectives were to use high performance liquid chromatography (HPLC) to validate an established method for adenine nucleotide separation in equine colonic mucosal tissue, to determine the inherent variability in the tissue and extraction method, and to determine the stability of ATP, ADP, and AMP in the tissue with time. Equine colonic mucosal tissue obtained from a single horse was immediately submersed in liquid nitrogen, and stored at -70 degrees C. Samples were lyophilized, extracted, and separated by HPLC. The limit of quantitation was 0.05 microg/mL. The coefficient of variation for the instrument was less than 10% for all nucleotides measured. When the tissue was not homogenized prior to sampling, there were significant differences in adenine nucleotide content between samples. However, when the tissue was homogenized prior to analysis, these differences were no longer significant. There was no significant decrease in ATP, ADP, or AMP content over a 54-day analysis period.     

Research note: Postnatal development of electrolyte transport in calf rumen as affected by weaning time.

In a previous study we found a positive correlation between early weaning in calves and morphological parameters which were indicative of ruminal development, i.e. the length and width of the papillae. The objective of the present study was to determine to what extent this observation could be reflected by modulations of absorptive and secretory functions of the rumen mucosa. For this purpose the short-circuit currents (Isc) as a measure of electrogenic net ion fluxes and the transepithelial conductances (G(T)) as a measure of the overall tissue permeability were measured in vitro applying the Ussing-chamber technique. Simultaneously, the unidirectional flux rates of sodium and chloride across rumen wall epithelia were determined in the absence of electrochemical gradients. Under these conditions, significant positive net flux rates (Jnet) clearly indicate active mechanisms for electrolyte absorption. For the experiments 12 male Holstein calves 7 d of age were assigned to three groups of 4 animals each: milk group (I, slaughtered after 6 weeks of age), late weaning group (II, slaughtered after 9 weeks of age) and early weaning group (II, weaned after 6 weeks of age and slaughtered after 9 weeks of age). Whereas G(T) values remained unaffected by different age and feeding, Isc values were significantly affected by early weaning but were not influenced by age. Irrespective of weaning time active absorption of Na+ tended to be higher by about 60% in 9 weeks old animals. Active absorption of chloride was significantly increased in milk fed 9 weeke old calves and this effect was further stimulated by early weaning. In conclusion, the data show an increasing active Na+ absorption with age in calf rumen that could not be influenced by early weaning. Similarly, active Cl- absorption was initially increased during postnatal development and this effect could be stimulated further by early weaning.     

The Effect of Diet on the Biochemical Composition of Juvenile Artemia: Potential Formulations for Rock Lobster Aquaculture

The lipid class and fatty acid (FA) composition of juvenile Artemia fed continuously on four diets—the microalga Tetraselmis suecica , a mix of oat bran-wheat germ-lecithin (OWL), OwL-eicosapentaenoic acid (EPA), and OWL-EPA-arachidonic acid (AA)—were examined over a 9-d experiment in an attempt to approximate the FA profile of phyllosoma larvae of wild southern rock lobster Jasus edwardrii . The main difference in lipid class composition of Artemia fed the four diets was the relative level of polar lipid (PL) and triacylglycerol (TAG). By day 9, the algal-fed Artemia were highest in PL (95% of total lipid) and lowest in TAG (2%), whereas the remaining diets resulted in Artemia with 16–30% PL and 41–82% TAG. After 2 d, the relative FA composition of all Artemia treatments closely reflected those of the diets, with no marked change after further feeding (to day 9). In terms of the content of essential polyunsaturated fatty acids (PUFA), by day 5 Artemia fed: 1) with the algal diet contained 7 mg/g FA dry mass (0.3% DHA, 6.3% EPA, 3.4% AA of total FA); 2) with the OWL diet contained 3 mg/g (0.3% DHA, 0.9% EPA, 0.7% AA); 3) with the OWL-EPA diet contained 55 mg/g (6.2% DHA, 11.6% EPA, 1.1% AA); and 4) with the OWL-EPA-AA contained 83 mg/g (3.8% DHA, 7.5% EPA, 17.4% AA). The PUFA profiles of Artemia using the OWL-oil diets were similar to wild rock lobster phyllmmata, although levels of doco-sahexaenoic acid (DHA) were lower (10% DHA) than in J. edwardsii larvae. On the basis of PUFA composition data alone, the results suggest the suitability of the OWL-oil mixed diets for consideration for feeding to Artemia used in the culture of southern rock lobster larvae, particularly if the level of DHA can be further enhanced.     

Experimental and natural infection with the enzootic leukosis virus of cattle

A trial was performed with heifers at the age of six to seven months. The animals were experimentally infected with the lymphocytes of a virus-productive donor. Infection was produced in all the nine cases, as demonstrated by means of the positive syncytial test. As indicated by the results of the trial, the antibodies to the enzootic bovine leucosis virus (BLV) were produced soon after experimental infection. A high sensitivity of the serum-neutralization test and the ELISA method was demonstrated in this connection: by these methods, the antibodies were identified already two to three weeks after experimental infection whereas by the immunodiffusion test they could be detected only after five weeks. Twenty-four animals were exposed to natural contact infection. Within 270 days of the trial, the disease after contact was recorded only in one heifer out of the four that were in close contact with the experimentally infected animals. In this case, as compared with experimental infection, the antibodies were produced much later--after 85 to 93 days. Leucosis was recorded in none of the remaining animals. The reasons why such a favourable result was obtained were the thorough disinfection of the stables after blood collections and the strict observance of the aseptic conditions. The results of experimental infection in three cows were identical with those obtained in young cattle. In the experimentally infected dairy cows, antibodies in milk were determined by the ELISA method. As found, in milk the antibodies to BLV appear two to three weeks later than they do in serum. The ELISA method of BLV antibody detection can be used for the identification of infected animals in herds where enzootic bovine leucosis occurs.