Thesis (M.Sc. (Animal Production)) -- University of Limpopo, 2018 / The study was conducted to determine the macroscopic and microscopic raw semen characteristics of Bapedi rams, to evaluate the effect of different egg yolk (EY) concentrations in Tris-based extenders on cryopreservation of Bapedi ram semen and to determine the effect of supplementing different phosphatidylcholine (PC) concentrations in Tris-based extenders with or without egg yolk on cryopreservation of Bapedi ram semen. Semen ejaculates were collected from four matured Bapedi rams aged 2-4 years using artificial vagina (AV) and pooled to eliminate individual differences. The first experiment was performed to characterise Bapedi ram semen parameters immediately after semen collection. The macroscopic semen parameters such as volume, pH and concentration and microscopic semen parameters such as motility, viability and morphology, membrane integrity and acrosome integrity were evaluated. The experiment was replicated 8 times and the data was subjected to descriptive statistics. The second experiment evaluated the effect of Tris-based extenders with five different EY concentrations (0, 5, 10, 15 and 20 %) on the microscopic quality of cryopreserved Bapedi ram semen. The treatments were subjected to a Completely Randomized Design (CRD) and replicated 4 times. The third experiment evaluated the effects of different PC concentrations supplemented to Tris-based extenders with or without 10% EY and the PC was added as liposomes. The experiment was a 2 x 4 factorial design in a CRD with two concentrations of EY: 0 and 10 %, and four concentrations of PC: 0, 0.25, 0.50, 0.75 mg/ml in Tris-based extenders. Pooled semen samples were divided into 5 and 8 aliquots to comply with objective 2 and objective 3, respectively. The semen aliquots were diluted with Tris-based extenders and equilibrated in a refrigerator at 5°C for another 4 hours. The semen was frozen using a programmable freezer and plunged into liquid nitrogen tank (-196°C).The volume, sperm concentration and pH of Bapedi ram semen ranged between 0.4-1.5 ml, 0.52-8.84 × 109 sperm/ml, and 5-7, respectively. The average total motility (TM), progressive motility (PM) and rapid motility (RM) characteristics were 85.95±2.58 %, 29.33±2.11 % and 39.47±4.99 %, respectively. The results for the mean percentage live spermatozoa, abnormalities, intact membrane and intact acrosome were 70.19±2.29 %, 2.50±1.34 %, 72.39±1.71 % and 75.37±5.39 %, respectively. There was a general decrease trends in frozen-thawed motility characteristics such as TM, PM and RM as compared to raw semen (p<0.05). The frozen-thawed semen in Tris-based extenders with 10, 15 and 20% EY concentrations resulted in significantly (p<0.05) higher TM, PM and RM motility characteristics compared to 0 and 5%. The percentage of live spermatozoa, membrane and acrosome integrities were found higher in raw semen than in frozen–thawed semen of respective extenders (p<0.05). The supplementation of PC in extenders either with or without EY did not improve the TM, PM and RM parameters (p>0.05). The membrane integrity in extenders either with or without EY were not influenced by the supplementation PC after freezing and thawing (p>0.05). The supplementation of PC in treatments with EY did not improve the acrosome integrity (p>0.05). Interestingly, the supplementation 0.75 mg/ml PC resulted in acrosome integrity that was not significantly different (P>0.05) to treatments with EY. In conclusion, the macroscopic and microscopic semen parameters of raw Bapedi ram semen were characterized. The use of 10% EY concentration resulted in higher motility parameters and membrane integrity of frozen-thawed Bapedi ram semen. However, 20% EY resulted in higher acrosome integrity of frozen-thawed Bapedi ram semen. The supplementation of PC in extenders in extenders with or without EY did not improve the motility parameters, percentage live spermatozoa and membrane integrity. However, the acrosome integrity was improved in extenders without EY supplemented with 0.75 mg/ml PC / Agricultural research council professional development programme (ARC-PDP)

Thesis (Ph. D.)--University of Wisconsin--Madison, 1969. / Typescript. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references.

<p>A common modification of starch is esterfication with anhydrous octenyl succinic acid (OSA).  The modification makes the polymer surface active and it also incorporates a carboxyl group to the starch, which can be negatively charged. The characteristics of OSA starch make it interesting for usage in combination with egg yolk proteins in food emulsions. It is not only the individual ingredients that affect the product; interactions between ingredients and ingredient-dispersion medium have a great impact on factors such as structure and stability. Knowledge about how the interactions affect emulsion properties would make it possible to predict the behavior of an emulsion, which would be a great advantage in the formulation of food emulsions. Therefore, this is a subject of interest.</p><p>The purpose of this master thesis was to further investigate the interactions between OSA starch and α – β-livetin in solutions and in emulsions. First, the charges of the macromolecules were studied by titration. Interactions in solution were then analyzed through turbidity and solubility measurements. The adsorption of OSA starch onto livetin and the interfacial rheology were also studied. Finally, an emulsion stability experiment was made.</p><p>Strong interactions between the two macromolecules were observed in solutions at pH 4.0. This was probably due to hydrophobic interaction; however it could also be explained by electrostatic interaction. In the emulsions the adsorption of starch onto livetin was highest at pH 4.5, and then decreased with increasing pH values. The absence of OSA starch adsorption at pH 4.0, despite the strong interaction in solution, could be explained by complex formation immediately in solution. Less starch would then be able to reach the interface and adsorb. In the interfacial rheology experiments, an indication of decreased complex dilational modulus of the interfacial layer, caused by OSA starch addition was seen at low pH values. This could be due to aggregation of the proteins and formation of an uneven interfacial layer. OSA starch would then be able to adsorb and disturb the elasticity. Some differences in the stability of an emulsion only containing livetin, and an emulsion with both livetin and OSA starch could be observed. However, more investigations are needed to be made to understand the underlying mechanisms.</p>

A common modification of starch is esterfication with anhydrous octenyl succinic acid (OSA).  The modification makes the polymer surface active and it also incorporates a carboxyl group to the starch, which can be negatively charged. The characteristics of OSA starch make it interesting for usage in combination with egg yolk proteins in food emulsions. It is not only the individual ingredients that affect the product; interactions between ingredients and ingredient-dispersion medium have a great impact on factors such as structure and stability. Knowledge about how the interactions affect emulsion properties would make it possible to predict the behavior of an emulsion, which would be a great advantage in the formulation of food emulsions. Therefore, this is a subject of interest. The purpose of this master thesis was to further investigate the interactions between OSA starch and α – β-livetin in solutions and in emulsions. First, the charges of the macromolecules were studied by titration. Interactions in solution were then analyzed through turbidity and solubility measurements. The adsorption of OSA starch onto livetin and the interfacial rheology were also studied. Finally, an emulsion stability experiment was made. Strong interactions between the two macromolecules were observed in solutions at pH 4.0. This was probably due to hydrophobic interaction; however it could also be explained by electrostatic interaction. In the emulsions the adsorption of starch onto livetin was highest at pH 4.5, and then decreased with increasing pH values. The absence of OSA starch adsorption at pH 4.0, despite the strong interaction in solution, could be explained by complex formation immediately in solution. Less starch would then be able to reach the interface and adsorb. In the interfacial rheology experiments, an indication of decreased complex dilational modulus of the interfacial layer, caused by OSA starch addition was seen at low pH values. This could be due to aggregation of the proteins and formation of an uneven interfacial layer. OSA starch would then be able to adsorb and disturb the elasticity. Some differences in the stability of an emulsion only containing livetin, and an emulsion with both livetin and OSA starch could be observed. However, more investigations are needed to be made to understand the underlying mechanisms.

This study tested different concentrations of egg yolk in canine freezing extender void of glycerol, a commonly used cryoprotectant, by examining the motility and morphology throughout the freezing process: initial (baseline after extender added), post-cool (after three hours at 5"C) and post-thaw (after freezing.)  Initial values of pH, osmolarity, motility and morphology were obtained for comparison of the samples.  Spermatozoa from six normal dogs as determined by progressive linear motility > 70% and normal morphology > 60% was used. Semen was collected and pooled for five freezing trials.  The concentrations of egg yolk used in the extender were: 0%, 10%, 20%, 30% and 40%. Assessment of each sample was blinded to the treatments until all results were obtained and statistics had been analyzed. Based on this study a 20% egg yolk concentration is slightly superior to a 30% egg yolk concentration when assessing post-thaw motility, morphology and longevity and significantly superior to a 0%, 10% or 40% egg yolk concentration. The study also showed motility and normal post-cool and post-thaw sperm morphology did not always correlate.  Utilization of 0% and 10% concentrations of egg yolk has negative effects on semen quality as measured by the motility and/or morphology.  Results confirm freezing does not affect secondary sperm abnormalities, abnormalities of the tail and distal section of the middle piece, during cooling or freezing.  Primary abnormalities, abnormalities of the head and midpiece, increased in the 0% extender during cooling and all extenders during freezing. The pH of the extenders before the addition of sperm was significantly different. Once sperm was added to the extenders, there was no longer a significant difference in pH.  There was a positive correlation for both motility and normal morphology percentages post-cool and post-thaw for the extenders with similar osmolarity to the semen. / Master of Science

Cryopreservation of germplasm is widely used in agriculture, biotechnology, conservation of threatened species and human reproductive medicine. There is a need however to improve the reproductive efficiency of breeding with cryopreserved semen, which may involve increasing the post-thaw quality of sperm through improvements in cryopreservation extenders. Extenders including egg yolk from chickens are successfully used worldwide for cryopreservation of bovine semen, whereas the protective agent in the egg yolk is believed to be the low-density lipoprotein (LDL) fraction. Egg yolks of different avian species vary in their cholesterol, phospholipid and polyunsaturated fatty acid content which have been shown to have important effects on sperm’s freezing capability. The purpose of this study was to determine the cryoprotective effect of clarified egg yolk and LDLs extracted from different egg yolk sources (chicken, chicken omega-3, pigeon, quail and turkey) on bovine sperm. Semen from six bulls was collected four times each by electroejaculation, split and diluted with the 10 following extenders: chicken clarified (Ccl), chicken omega-3 clarified (O3cl), pigeon clarified (Pcl), quail clarified (Qcl), turkey clarified (Tcl), chicken LDL (CLDL), chicken omega-3 LDL (O3LDL), pigeon LDL (PLDL), quail LDL (QLDL) and turkey LDL (TLDL). The extended semen was evaluated, cryopreserved and examined directly after thawing (0h) and after two hours at 37 ˚C (2h). Computer assisted sperm analysis (CASA) was used to determine total sperm motility (TM), progressive motility (PM), straight line velocity (VSL), curvilinear velocity (VCL) and average path velocity (VAP). Intact plasma membrane (IPM) and intact acrosomes (IA) were measured by flow cytometry. The percentage change (loss; Δ%) of each sperm characteristic was calculated and used to compare the effect of the extenders. From extending to 0h post-thaw, the pigeon LDL extender lead to greater losses in sperm total and progressive motility, as well as of intact acrosomes, than the other nine extenders tested (P < 0.05). During 0h to 2h post-thaw, the sperm in PLDL extender experienced greater losses in total and progressive motility (P < 0.0001), as well as in curvilinear velocity (P < 0.05), than in all the other nine extenders. Sperm in turkey clarified extender had a greater loss in the velocity parameters (VSL, VAP, VCL) than sperm in several of the other extenders such as O3cl, CLDL, O3LDL, QLDL and TLDL from 0h to 2h (P < 0.05). Concomitantly, sperm in the Tcl extender had a greater loss in the velocity parameters and of intact acrosomes compared to sperm in its counterpart, the turkey LDL extender, from 0h to 2h post-thaw (P < 0.05). The differences produced in post-thaw quality of cryopreserved bovine sperm in the pigeon LDL and turkey clarified extenders were attributed to methodological differences in these egg yolk preparations compared with the other eight extenders. Importantly, the results demonstrate that with most egg yolk preparations derived from a variety of species, there are equivalent cryoprotective effects afforded by the use of omega-3 chicken, pigeon, quail, or conventional chicken egg yolk in a clarified form in freezing extenders for bovine semen. We further proved that the freezing capabilities of bovine semen extenders containing the low-density lipoprotein fraction of omega-3 chicken, quail, turkey and conventional chicken egg yolk were similar.

This thesis describes a study of the aggregational behaviour of egg yolk lecithin (EYL), a natural lecithin, and bile salt mixtures especially with respect to an increase of the ionic strength of the solvent. Mixtures of two amphiphiles with very different spontaneous curvature as EYL lecithin and bile salt form mixed micelles and vesicles in aqueous solution. Their properties have been well-studied under physiological conditions, i.e. 150 mM electrolyte concentration and pH 7- 8, while other conditions are still hardly explored. Upon increasing ionic strength the formed structures and the transitional pathways (micelles, coexistence of micelles and vesicles, and vesicles) change the generated structures completely from those observed under physiological conditions. We quantitatively determined these structures formed in a broad range of electrolyte concentrations with various scattering techniques, x-ray, light and neutron scattering and calorimetry. With calorimetry, phase diagrams in the EYL and bile salt concentration phase plane were determined at various ionic strength ranging from physiological salt concentration to up to 1000 mM. Additionally a new electrochemical approach using functionalised electrodes, i.e. sensitive and selective to bile salt, and thus to control the bile salt concentration in solution (concentrations below the critical micellar concentration (cmc)) was attempted, since bile salt removal or injection drives the micelle-to-vesicle or the vesicle-to-micelle transition, respectively, of the mixed aggregational system of EYL/bile salt. Although this control was not achieved within the framework of this thesis, promising results show directions for future experiments.

No description available.

The effect of several fundamental reproductive 'decisions' upon reproductive success were examined over a three year period at a large, inland gullery in the Pennines. Variations in reproductive parameters in relation to timing of breeding and reproductive success were compared among years. Determinants of the degree of hatching asynchrony were identified. Eggs from two years were taken for yolk lipid analysis, using gas chromatography. Variation in micronutrient content of eggs within clutches and between years was assessed in relation to egg size and yolk size, in order to examine resource allocation decisions of individuals. Timing of breeding of individuals was experimentally manipulated through the exchange of whole clutches of eggs between early and late laying birds, whilst controlling for variation in clutch size and egg-size, in order to assess whether the seasonal decrease in reproductive success was best explained by a decrease in food supply or differences in quality among parents. I experimentally manipulated the within-brood mass hierarchy of gulls, whilst controlling for variation in both chick quality and parental quality, in order to assess the effect of hatching asynchrony per se on chick growth and survival, and whether parents optimized the degree of hatching asynchrony with respect to the prevailing food supply. Brood size was experimentally reduced in order to assess the costs and benefits of the production of supernumary young. This was acheived by comparison of chick growth, feather development and chick survival of unmanipulated three-chick broods with broods where either the a-chick or the c- chick had been removed. I present a general discussion of the results within the context of life-history theory and a model for the evolution of hatching asynchrony in the lesser black-backed gull.

Pseudomonas Aeruginosa is a common gram-negative bacterium present in the environment. It causes severe infections in immunosuppressed patients. Cystic fibrosis patients are especially at risk of being infected with Pseudomonas Aeruginosa. Ongoing studies are preformed to find alternative therapies to antibiotics, due to increased resistance. One new treatment is intake of specific IgY antibodies against Pseudomonas Aeruginosa as an oral therapy. The problem today is that IgY solutions must be kept frozen until consumed.  In this study we examined the possibility to freeze-dry specific IgY antibodies without losing any activity or specificity of the antibodies. This would be more convenient of patients, as well as it makes transportation and storage easier.  The methods used were ELISA for control of activity, western blot analysis and SDS-PAGE gel for control of specificity. Three different batches of the IgY anti-Pseudomonas Aeruginosa solution were tested. The results showed that no loss in activity occurred that would affect clinical outcome or change of specificity in the antibodies after freeze-drying appears. This indicates that it is possible to replace the liquid antibody to a freeze-dried powder.