Process optimization using response surface design and pilot plant production of dietary diacylglycerols by lipase-catalyzed glycerolysis

Two approaches to shift the acylglycerol equilibrium were tested as follows: addition of monoacylglycerols and lowering of the temperature. None of these approaches were able to shift the equilibrium toward higher diacylglycerol (DAG) contents. The glycerolysis reaction was optimized with five factors using response surface methodology. Evaluation of the resulting model enabled the determination of optimal reaction conditions for glycerolysis aiming at high DAG yield. However, verification of the model showed that the model was unable to take the molecular equilibrium into account but it provided good insight in how process settings can be chosen to, for example, minimize production costs. Optimal conditions were found to be the following: no extra water, low content of glycerol (molar ratio of 2), temperature of 60-65 degrees C, 4-5 h reaction time, and only 5 wt % lipases. Up scaling of the glycerolysis process was performed and revealed that scale-up to a 20 kg production in a pilot plant batch reactor was possible with a similar DAG yield (60 wt %) as in lab scale. Purification of DAG oil using batch deodorization and short path distillation yielded 93 wt % pure DAG oil.     

In vitro antioxidant activities of edible artichoke (Cynara scolymus L.) and effect on biomarkers of antioxidants in rats

Artichoke (Cynara scolymus L.), an edible vegetable from the Mediterranean area, is a good source of natural antioxidants such as vitamin C, hydroxycinnamic acids, and flavones. The antioxidant activity of aqueous-organic extracts of artichoke were determined using three methods: (a) free radical 2,2-diphenyl-1-picrylhydrazyl (DPPH(*)) scavenging, (b) ferric-reducing antioxidant power (FRAP), and (c) inhibition of copper(II)-catalyzed in vitro human low-density lipoprotein (LDL) oxidation. In addition, the present study was performed to investigate the ability of the edible portion of artichoke to alter in vivo antioxidative defense in male rats using selected biomarkers of antioxidant status. One gram (dry matter) had a DPPH(*) activity and a FRAP value in vitro equivalent to those of 29.2 and 62.6 mg of vitamin C and to those of 77.9 and 159 mg of vitamin E, respectively. Artichoke extracts showed good efficiency in the inhibition in vitro of LDL oxidation. Neither ferric-reducing ability nor 2,2'-azinobis(3-ethylbenzothiazolin-6-sulfonate) radical scavenging activity was modified in the plasma of the artichoke group with respect to the control group. Among different antioxidant enzymes measured (superoxide dismutase, gluthatione peroxidase, gluthatione reductase, and catalase) in erythrocytes, only gluthatione peroxidase activity was elevated in the artichoke group compared to the control group. 2-Aminoadipic semialdehyde, a protein oxidation biomarker, was decreased in plasma proteins and hemoglobin in the artichoke-fed group versus the control group. In conclusion, the in vitro protective activity of artichoke was confirmed in a rat model.     

Mitral valve prolapse in 3-year-old healthy Cavalier King Charles Spaniels. An echocardiographic study.

Clinical studies have shown that Cavalier King Charles Spaniels (CKCS) have a high prevalence of mitral valvular insufficiency (MVI). Echocardiography has the potential to disclose early valvular changes, and the present prospective study was designed to investigate the occurrence of mitral valve prolapse (MVP) in young CKCS without heart murmurs, and to correlate the degree of MVP with the clinical status of the dogs by including CKCS with MVI as well. The study was based on blinded evaluations of echocardiographic recordings of mitral valves from 34 CKCS and 30 control dogs. Thirteen (87%) of 15 three-year-old CKCS without heart murmurs had MVP (2 total and 11 partial), as compared with 1 (7%) of 15 three-year-old normal Beagle dogs (P < 0.0001), and none of 15 three-year-old normal Medium Size Poodles (P < 0.0001). Of 19 CKCS with MVI, MVP was found in 84% of the entire group and in 100% of dogs with pulmonary congestion or edema. The occurrence of total MVP tended to be higher in the group with MVI (47%, 9/19), when compared with the younger CKCS without heart murmurs (13%, 2/15, P = 0.06). MVP was positively associated with excessive heart rate variability (P = 0.003). The radius of curvature of the anterior mitral valve leaflet in systole was significantly reduced in dogs with MVP when compared with those without (P < 0.0001). In conclusion, this study shows that CKCS at an early age have a high occurrence of MVP. This suggests: 1) A genetic predisposition of CKCS to MVP; and 2) That MVP is a pathogenetic factor in the development of mitral valvular insufficiency. Follow up studies may add further support to these proposals, and clarify whether echocardiography may be an aid in selecting CKCS for future breeding.     

The response of normal and iron anemic calves to nasal infection with an attenuated strain of parainfluenza-3 virus.

Calves with experimentally induced iron deficiency anemia and normal calves, both groups deprived of colostrum, were exposed to intranasal instillation of an attenuated parainfluenza-3 virus strain.The calves became infected, but there was no difference in the clinical picture between the 2 groups of calves. Neither was there a difference in the humoral or local immune response to parainfluenza-3 virus.     

Cytology in veterinary practice. A review of clinical cytology--body fluids, lymph nodes and skin neoplasms

A review over the most applicable areas of veterinary clinical cytology is given. The areas described are body fluids, lymph nodes and selected skin neoplasms. The review includes tables that survey general and specific cytologic criteria and is supplemented by photomicrographs. A classification of effusions according to cytologic criteria is discussed and there is a short discussion of 62 cases which also included histopathology.     

Mitogen stimulation of lymphocytes in pigs with hereditary vitamin C deficiency

Recently an inherited vitamin G deficiency in the pigs presumably based on an autosomal recessive gene was decribed* Homozygotes are in contrast to heterozygotes and normal pigs unable to synthesize ascorbic acid. In an experiment comprising 3 littermate pigs, 2 homozygous and 1 heterozygous for the vitamin C deficiency gene, the influence of ascorbic acid depletion, and repletion on mitogen stimulation of peripheral blood lymphocytes was studied. Ascorbic acid depletion of the vitamin C dependent pigs resulted in a rapid decline in plasma ascorbic acid. Response of lymphocytes to stimular tion with Concanavalin A (Con A) and phytohemagglutinin M (PHA) decreased more slowly reaching a minimum, which coincidedi with the occurrence of the first clinical symptoms of scurvy. Following resupplementation with vitamin C the plasma content of ascorbic acid rapidly returned to normal, while the lymphocyte response to Con A and PHA stimulation only gradually approached the initial values. The repletion with ascorbic acid caused a transitory increase in the response to pokeweed mitogen (PWM) stimulation. The significance of these findings in relation to the cellular immune system in normal pigs is discussed.     

Analysis of the in vitro activation and proliferation process in lymphocytes deriving from canine thymus and mesenteric lymphnode

Lymphocytes from dog thymus and mesenteric lymphnodes have been stimulated in vitro with 3 different mitogens. Culture medium was enriched with either autologous plasma, fetal calf serum or a newly described defined serum substitute. In such cultures the number of surviving and activated cells was quantified by cytofluorometry and the proliferation was assessed by thymidine incorporation. Results obtained with the 3 media were very similar. A significant number of lymphocytes died during the first 42 hours of incubation. There was a tendency toward more surviving cells with fetal calf serum and the serum substitute, whereas more activated (G₁) cells and higher thymidine incorporation could be observed with autologous plasma. Furthermore, when results obtained with various mitogen concentrations and from individual dogs were analyzed, a high correlation between “highly activated (G_1b) cells and thymidine incorporation was found, i.e. r=0.84 for thymocytes and 0.68 for lymphnode cells. The correlation between all G₁ (G_1a+b) cells and thymidine incorporation was lower or absent (r=0.02 and 0.55, respectively). It is concluded from these results that the population of G_1b cells have received all required signals necessary for proliferation whereas the total G₁ cell population also include activated cells, which are not obligatorily undergoing subsequent proliferation     

Canine lymphocyte cultures in vitro: Evaluation of peripheral blood lymphocyte response to mitogens

Lymphocytes from dog peripheral blood have been stimulated in vitro with 3 different mitogens (Con A, PHA and PWM). Culture medium was RPMI 1640 enriched with either autologous plasma, fetal calf serum or a newly described defined serum substitute. In such cultures the number of surviving and activated cells was measured by cytofluorometry and the proliferation was assessed by thymidine incorporation. In unstimulated cultures, up to 70% of all cells had disappeared (died) during the first 42 hours of incubation, whereas the number of viable cells was reduced to 50–60% in mitogen stimulated cultures. Of the surviving lymphocytes, between 25–40% of the cells appeared to have an elevated RNA-content (activated or G₁ cells). By comparison between thymidine incorporation and number of mitogen induced G₁ cells, a very high correlation was found (r=0.92). However, the slope of the regression line was much lower than expected. The low thymidine incorporation per activated cell was primarily related to the high cell death and a resulting dilution of tritiated thymidine. Indeed, preliminary results suggested that the same thymidine incorporation per G_1b cells could be obtained if peripheral blood lymphocytes were washed immediately before pulsing as could be obtained with lymphnode cells without washing.     

Exudative epidermitis in pigs caused by toxigenic Staphylococcus chromogenes

Staphylococcus chromogenes is closely related to Staphylococcus hyicus, which is recognised as the causative agent of exudative epidermitis (EE) in pigs. S. chromogenes is part of the normal skin flora of pigs, cattle and poultry and has so far been considered non-pathogenic to pigs. A strain of S. chromogenes producing exfoliative toxin type B, ExhB, was identified by the use of a multiplex PCR specific for the exfoliative toxins from S. hyicus. The exfoliative toxin from S. chromogenes reacted in immunoblot analysis with polyclonal and monoclonal antibodies specific to ExhB from S. hyicus and had an apparent molecular weight of 30 kDa. Sequencing the gene encoding the exfoliative toxin from S. chromogenes revealed that the molecular weight of the toxin with the signal peptide and the mature toxin was 30,553 and 26,694 Da, respectively. Comparison of the exhB genes from S. chromogenes strain VA654 and S. hyicus strain 1289D-88 showed differences in seven base pairs of the DNA sequences and in two amino acid residues in the deduced amino acid sequences. Pigs were experimentally inoculated with S. chromogenes strain VA654. By clinical observations and histopathological evaluation of the skin alterations, all pigs revealed development of generalized exudative epidermitis. No toxin producing S. hyicus was isolated from the pigs and all ExhB-positive bacterial isolates were identified as S. chromogenes. This confirmed that the disease-causing agent was the inoculated S. chromogenes strain VA654. The results of this study show that S. chromogenes may cause exudative epidermitis in pigs.