Content: Unhairing process brought serious pollution, and enzyme application for replacing polluting chemicals in unhairing process attracted much attention in recent years. However, the unhairing enzymes haven’t been accepted widely in actual production due to low purity, complex composition and poor stability. To solve these problems, unhairing enzyme is suggested to be improved by genetic modification in this research. The High-keratinase-producing gene (KerT), which was extracted from B. amyloliquefaciens TCCC11319, was introduced into the B.subtilis WB600 by heterologous expression. Because Bacillus subtilis WB600 is deficient in six extracellular proteases, this process successfully reduced the collagenolytic protease content in crude broth as well as improved the keratinase content. Meantime, the recombinant KerT produced by B.subtilis WB600 had the obviously unhairing effect to remove hairs. The results showed that the collagen degradability of recombinant KerT was slightly and it did not cause any adverse effects on the hide quality. This research will contribute to the development of unhairing enzyme, and the novel unhairing enzyme might be applied as the key factor for the advanced cleaning biotechnology in leather production process. Take-Away: 1. The keratinase gene KerT was firstly reported and analyzed which was extracted from B. amyloliquefaciens TCCC11319. 2. The collagenolytic protease activity of unhairing enzyme was successfully inhibited by heterologous expression of kerT. 3. The unhairing effect of this novel unhairing enzyme was similar to current Sulphur-lime method without damaging hide structure.

Les travaux de recherche réalisés, dans le cadre de la préparation de cette thèse ont pour objectif l’élaboration de nouveaux réseaux thermoréversibles biodégradables à base de polyglycolide (PGA) et d’hydrogels à base de polylactide (PLA) modifiés chimiquement au moyen du noyau furanique et du cycle maléimide. La réticulation du PGA, en suivant deux stratégies, en vue d’élaborer des réseaux thermoréversibles et biodégradables via la réaction de Diels-Alder a fait l’objet de la première partie de ce travail. Le but était entre autres de valoriser le PGA en conduisant à des structures réticulées avec des propriétés mécaniques requises tout en améliorant ses propriétés de stabilité. Des réseaux hybrides de PLA/PEG et PLA/PHEMA ont été ensuite synthétisés en phase fondu en adoptant la réaction de Diels-Alder comme mécanisme de réticulation. Une fois obtenus, leur mise au contact de l’eau conduit à la formation d’hydrogels. Les analyses structurales menées par RMN ont permis de confirmer la formation des structures attendues. La thermoréversibilité des réseaux obtenus a été montrée par des analyses rhéologiques. La morphologie des gels avant et après gonflement a été analysée par Microscopie Electronique à balayage. La dégradabilité des réseaux préparés a été examinée selon deux modes : hydrolytique et par les microorganismes en milieu aérobie. / The research conducted for the preparation of this thesis aims to develop new thermoreversible and biodegradable polyglycolic-acid (PGA) based networks and polylactic-acid (PLA) based hydrogels, from polymers chemically modified by means of furanic, and maleimide cycle. The cross-linking of PGA to develop thermoreversible and biodegradable networks via the Diels-Alder reaction has been done by following two strategies and was the first part of this work. The aim was, among other things, to enhance the PGA by leading to reticulated structures with required mechanical properties while improving its stability properties. Hybrid networks of PLA / PEG and PLA / PHEMA were then synthesized in the melt by adopting the Diels-Alder reaction as a crosslinking mechanism. Once obtained, their contact with water leads to the formation of hydrogels. NMR structural analyzes confirmed the formation of expected structures. The thermoreversibility of the obtained networks has been shown by rheological analyzes. The morphology of the gels before and after swelling was analyzed by Scanning Electron Microscopy. The degradability of prepared networks was examined in two modes: hydrolytic and aerobic by microorganisms.

Bovine somatotropin (bST) is a protein synthesized at the base of the brain and released by the pituitary gland into the circulatory system. BST is transported by the circulatory system and absorbed only by cells of target organs that possess cell surface receptors for the protein (11, 41). Its effect is initiated via a protein receptor initiation and cyclic AMP cascade . This effect on the cell continues well past the degradation of the bST molecule. BST is likely transported into the cell, where it is degraded . Its constituent blocks, amino acids, are used to synthesize new proteins or converted to other metabolites such as sugars (1).

Twenty-four early to midlactation Holstein cows were allocated to one of three treatments. Treatments consisted of: 1) basal ration plus 60 g wheat bran per day per head (control); 2) basal ration plus 57 g yeast culture per day per head; 3) basal ration plus 57 g yeast culture plus 3 g Aspergillus oryzae per day per head. Feed intake and milk yield were recoreded daily and milk composition and body weights were recorded weekly. Feed and fecal samples were recorded weekly. Feed and fecal samples were collected to determine apparent nutrient digestibility. No statistical difference was observed in milk yield among treatments. Percent fat, protein, and solid non fat of milk samples for the control fed group was significantly higher than the other treatment groups. Apparent crude protein, acid detergent fiber, and neutral detergent fiber digestibilities for cows fed the fungal culture treatments were significantly higher. Six rumen-fistulated Holstein cows were randomly assigned to one of three treatments in a 3x3 replicated Latin square design. Rumen pH was significantly lower for cows fed treatment two. Total volatile fatty acid and ammonia nitrogen concentration were higher for cows fed treatment two. Three rumen-fistulated Holstein cows were assigned one of three treatments in a 3x3 Latin square design for an in situ study. Approximately 15 g of grass hay, low quality alfalfa, and high quality alfalfa were sealed in nylon bags and put into the rumen. Samples were analyzed for dry matter, acid detergent fiber, neutral detergent fiber, and crude protein. Regardless of substrate used, dry matter disappearance was lower for control. Regardless of treatment ration fed, high quality alfalfa as a substrate was significantly higher in dry matter disappearance.

Nanoparticle mediated drug delivery approaches provide potential opportunities for targeting and killing of intracellular bacteria. Among them, the porous silica nanoparticles deserve special attention due to their multifunctional properties such as high drug loading, controlled drug release and targeting of organs/cells. A review of the functional requirements of an ideal drug delivery system is provided. A general comparison between different drug delivery carriers and key issues to be addressed for intracellular drug delivery is discussed. Acid catalyzed and acid-base catalyzed, sol-gel derived, silica xerogel systems were investigated for sustained release of an aminoglycosides antimicrobial against salmonella infection in a mouse model. The release of gentamicin from the inner hollow part of the carrier is delayed. Further, the higher porosity of the acid–base catalyzed silica xerogel allows for high drug loading compared to the acid catalyzed silica xerogel system. Efficacy of these particles in killing intracellular bacteria (salmonella) was determined by administering three doses of porous silica loaded gentamicin. This proved to be useful in reducing the salmonella in the liver and spleen of infected mice. Furthermore, the presence of silanol groups provides the ability to functionalize the silica xerogel system with organic groups, poly (ethylene glycol) (PEG), to further increase the hydrophilicity of the silica xerogel matrix and to modify the drug release properties. Increase in the hydrophilicity of the matrix allows for faster drug release rate. In order to facilitate controlled drug release, magnetic porous silica xerogels were fabricated by incorporating iron particles within the porous silica. The particles were fabricated using an acid-base catalyzed sol-gel technique. The in-vitro drug release studies confirm that the release rate can be changed by the magnetic field "ON-OFF" mechanism. This novel drug release methodology combined with the property of high drug loading capacity proves to be influential in treating salmonella intracellular bacteria. The potential application of any drug delivery carrier relies on the ability to deliver the requisite drug without adversely affecting the cells over the long term. We have developed silica/calcium nanocomposites and evaluated their solubility behavior. The solubility of particles was characterized by particle size measurements for different periods of time. It was found that the solubility behaviour of the silica/calcium particles was dependent on their calcium content. The results obtained demonstrate the potential to use mesoporous silica/calcium nano-composites for drug delivery applications. The significant contribution of this research to drug delivery technology is on design and development of the novel porous core-shell silica nano-structures. This new core-shell nano-structure combines all the above mentioned properties (high drug loading, magnetic field controlled drug release, and solubility). The main aim of preparing these porous core-shell particles is to have a control over the solubility and drug release property, which is a significant phenomenon, which has not been achieved in any other drug delivery systems. The shell layer acts as a capping agent which dissolves at a controllable rate. The rate at which the shell layer dissolves depends on the composition of the particles. This shell prevents the drug "leakage" from the particles before reaching the target site. The core layer drug loading and release rate was modified by application of a magnetic field. Additionally, inclusion of the calcium ions in the core layer destabilizes the silica network and allows the particles to dissolve at an appropriate rate (which can be controlled by the concentration of the calcium ions). / Ph. D.

Thesis (MScAgric)--Stellenbosch University, 2012. / ENGLISH ABSTRACT: Grinding of feedstuffs prior to in sacco incubation results in fine particles that could escape from dacron bags without being degraded. The objective of this study was to determine the effect of the removal of these fine particles on protein degradation parameters. The feedstuffs that were used were protein feedstuffs that are commonly used in dairy cattle diets in South Africa. Feedstuffs were soybean oilcake, sunflower oilcake, maize gluten 20, maize gluten 60 and fish meal. Treatments were; 1. grinding through a 2 mm screen with no subsequent sieving, 2. grinding through a 2 mm screen followed by sieving through a 106 μm mesh, and 3. grinding through a 2 mm screen followed by sieving through a 150 μm mesh. In the first trial, the material that was lost through sieving and the chemical composition of the different treatments were determined. Protein solubility was also determined. Between 20 and 60% of the material dry matter was lost with the sieving. The chemical composition for the soybean and sunflower oilcake and maize gluten 60 was similar between the three different treatments (or fractions). The CP content of fish meal and maize gluten 20 differed somewhat between the treatments. The 106 μm mesh seemed to be most suitable for fish meal, but a suitable mesh size could not be found for maize gluten 20. In the second trial, the degradability parameters were determined according to the in sacco degradation procedure. Three lactating Holstein cows that were fitted with rumen cannulae were used. The cows received a commercial lactation diet and oat hay that was supplied ad libitum. Samples of all the protein sources were placed in dacron bags and incubated in the rumen. The following removal times were used: 0, 2, 4, 8, 12, 24 or 48 hours. Dry matter and CP disappearances were determined, and the values were used to estimate DM and CP degradability parameters using a non-linear model. Effective CP degradability was also determined. The a-values were affected most of all. On average, the a-values were 39.4 and 40.3% higher for the un-sieved treatments than for the sieved treatments, for DM and CP, respectively. The effective CP degradability was also, on average, 43% higher for the un-sieved treatments. Grinding without the subsequent sieving of samples appears to result in an overestimation of DM and CP degradation in the rumen. It is therefore recommended that after grinding, feedstuffs should be sieved through at least a 106 μm mesh in preparation for in sacco incubations. / AFRIKAANSE OPSOMMING: Die maal van grondstowwe ter voorbereiding van in sacco inkubasies lei tot ‘n potensiële verlies van klein partikels uit die dakronsakkies sonder dat hulle gedegradeer word. Die doel van hierdie studie was om die invloed van die verwydering van fynmateriaal op proteïen-degradeerbaarheidsparameters te bepaal. Die grondstowwe wat in hierdie studie gebruik is, is proteïengrondstowwe wat algemeen in melkbeesdiëte in Suid-Afrika gebruik word. Die grondstowwe was soja-oliekoek, sonneblom-oliekoek, mielie gluten 20, mielie gluten 60 en vismeel. Behandelings was; 1. maal deur ‘n 2 mm sif sonder verdere sifting, 2. maal deur ‘n 2 mm sif gevolg deur sifting deur ‘n 106 μm sif en 3. maal deur ‘n 2 mm sif gevolg deur sifting deur ‘n 150 μm sif. In die eerste proef is die hoeveelheid materiaal wat verlore gaan as gevolg van sifting van die grondstowwe bepaal en die chemiese samestelling van die verskillende behandelings. Proteïenoplosbaarheid is ook bepaal. Tussen 20 en 60% van die materiaal het verlore gegaan as gevolg van sifting. Die chemiese samestelling van die soja- en sonneblom oliekoek, asook dié van die mielie gluten 60 was soortgelyk vir al drie die behandelings. Die ruproteïeninhoud (RP) van die vismeel en mielie gluten 20 het verskil tussen die drie behandelings. Dit wil voorkom asof die 106 μm sif die mees geskikte is vir vismeel, maar ‘n geskikte sif kon nie vir mielie gluten 20 gevind word nie. In die tweede proef is die degradeerbaarheidsparameters bepaal met behulp van die in sacco-metode. Drie lakterende Holsteinkoeie met rumen kannulas is gebruik. Die koeie het ‘n kommersiële melkbeesdieët ontvang en hawerhooi ad libitum. Die monsters is in dakronsakkies in die rumen geïnkubeer. Die sakkies is na die volgende inkubasietye verwyder: 0, 2, 4, 8, 12, 24 of 48 uur. Die DM- en RP-verdwyning is bereken en die waardes is gebruik om die DM- en RP- degradeerbaarheidsparameters te bereken met behulp van ‘n nie-lineêre model. Effektiewe RP- degradeerbaarheid is ook bereken. Die waardes wat die meeste beïnvloed is, is die a-waardes. Die a-waardes was gemiddeld 39.4 en 40.3% hoër vir die ongesifte behandelings as vir die gesifte behandelings, vir DM en RP, onderskeidelik. Die effektiewe RP-degradeerbaarheid was ook gemiddeld 43% hoër vir die ongesifte behandelings. Dit wil voorkom asof DM- en RP-degradeerbaarheid oorskat word wanneer voermonsters slegs gemaal word. Dit word aanbeveel dat grondstowwe ten minste deur ‘n 106 μm sif gesif word ter voorbereiding vir in sacco-studies.

Thesis (MScAgric (Animal Sciences))--Stellenbosch University, 2010. / ENGLISH ABSTRACT: South Africa is well-known for periodic dry periods and uncertain rainfall. Ensiling of crops is a method of preserving forage and ensures feed availability during periods when the supply of good quality forage is low. Cereal-based silages, especially in the Western Cape, South Africa, represent a significant proportion of feed consumed by ruminant animals, particularly high-producyion dairy cattle. However, farmers are still concerned about the technical challenges of ensiling cereal crops. Previous research done on lactic acid bacteria (LAB) inoculants used on cereal based silage has indicated a potential for improving silage fermentation, stability and degradability, thus enhancing feed conversion and production by ruminants. Two experiments were conducted to determine the effects of inoculating whole-crop oat silage with Lalsil® Cereal Lactobacilli (Lactobacillus buchneri (NCIMB 40788) and Pediococcus acidilactici (CNCM MA 18/5M)) LAB on (1) silage fermentation, (2) aerobic stability and (3) nutritional value of silage ensiled under a. micro-silos conditions and b. in a bunker under outdoor conditions of a Mediterranean summer. Oats (Avena sativa, cv SSH 405) were planted on 60 ha under dryland conditions at Elsenburg in the Western Cape province, South Africa. Whole-crop oats were harvested at the soft dough stage and length of the chopped material was 9 mm (Day of ensiling – Day 0). Chopped whole-crop oats were sampled, mixed thoroughly and divided into two portions. The Inoculant (Lalsil® Cereal) was applied to one portion to provide 5.79 x 109 colony forming units (CFU) of LAB per gram of fresh material. In the first experiment twenty - four glass silos (1.5 L glass jars) (WECK, GmbH u.Co., Wehr-Ofligen, W. Germany) were filled for each of the control and inoculant treatments. The glass silos were stored in a dark room in the laboratory at ambient temperature. Three glass silos were opened for each treatment on days 1, 2, 4, 8, 15, 30, 60 and 102 post-ensiling to determine fermentation dynamics. A parallel study was done with the same chopped whole-crop oats using the buried bag technique in a bunker silo. Whole-crop oats were ensiled in six net bags per treatment buried in a bunker filled with the same untreated whole crop oats. Bags, attached with nylon lines (3 m lengths) for easy retrieval were buried at 1m and 2 m depths in the same bunker. The net bags in the bunker were retrieved after 186 days of ensiling. Dry matter (DM), organic matter (OM), neutral detergent fibre (NDF), crude protein (CP), lactic acid levels, pH, water soluble carbohydrates (WSC) and in vitro organic matter degradability (IVOMD) for both studies were determined. Silage of both experiments was exposed to aerobic conditions for ten days to determine aerobic stability. It is concluded that the inoculant Lalsil® Cereal had the effect of reducing the rate of consumption of WSC during the anaerobic phase and aerobic exposure for both experiments. Silage spoilage due to yeasts and moulds was however more evident with the inoculated silage due the presence of sugars in the micro-silos experiment. / AFRIKAANSE OPSOMMING: Suid-Afrika is bekend vir droë periodes en wisselvallige reënval. Die inkuiling van gewasse is ‘n goeie manier om ruvoer te voorsien in tye van droogtes of tekorte. Kleingraan kuilvoer is veral bekend in die Wes-Kaap, Suid-Afrika en maak ‘n groot deel uit van die melkkoei se rantsoen. Landbouers is nog steeds bekommerd oor die tegniese aspekte wanneer dit kom by die inkuil van gewasse. Vorige navorsing het getoon dat die gebruik van ‘n melksuurbakteriese inokulant saam met die inkuiling van gewasse moontlik die potensiaal het om fermentasie, stabiliteit en degradering te verbeter en sodoende voeromset te verbeter. Twee eksperimente is uitgevoer om die effek van die inkuiling van hawerkuilvoer met Lalsil® Cereal Lactobacilli (Lactobacillus buchneri (NCIMB 40788) en Pediococcus acidilactici (CNCM MA 18/5M)) LAB te bepaal op (1) kuilvoer fermentasie, (2) aërobiese stabiliteit en (3) nutriëntwaarde van die kuilvoer ingekuil in a. mikrosilo’s en b. in ‘n bunker in die buitelug. Hawer (Avena sativa, cv SSH 405) is op 60 ha droë land geplant op Elsenburg in die Wes-Kaap, Suid-Afrika. Die hawer is ingekuil tydens die sagte deeg stadium en die gekapte materiaal was ongeveer 9 mm lank. Gekapte material was deeglik gemeng en in twee gedeel. Die inokulant (Lalsil® Cereal) is op die een gedeelte gesproei om 5.79 x 109 kolonie-vormende eenhede (KVE) melksuurbakterieë per gram vars materiaal te voorsien. Tydens die eerste eksperiment is 24 mikrosilo’s (1.5 L glas silo) (WECK, GmbH u.Co., Wehr-Ofligen, W. Duitsland) vir elke behandeling vol kuilvoer gemaak. Hierdie mikrosilo’s is gestoor in ‘n donker kamer teen kamertemperatuur. Drie mikrosilo’s is per behandeling oopgemaak op dag 1, 4, 8, 15, 30, 60 en 102 na inkuiling om die fermentasie-dinamika te bepaal. ‘n Parallelle studie is gedoen met dieselfde materiaal ingekuil in netsakke binne die bunker. Die materiaal was ingekuil in ses netsakke vir elke behandeling. Nylon toue (3 m) is aan die sakke vasgemaak om die uithaal daarvan op latere stadium te vergemaklik. Hierdie sakke is ingekuil op verskillende dieptes, 1 m en 2 m in dieselfde bunker. Die sakke is na 186 dae weer uitgehaal. Droë materiaal (DM), organiese materiaal (OM), neutraal bestande vesel (NBV), ruproteïen (RP), melksuurvlakke, pH, water oplosbare koolhidrate (WOK) en in vitro organiese materiaal verteerbaarheid (IVOMV) vir beide studies is bepaal. Kuilvoer van beide eksperimente is ook blootgestel aan aërobiese toestande vir 10 dae aan aërobiese toestande blootgestel om aërobiese stabiliteit te bepaal. Daar is bepaal dat die inokulant Lalsil® Cereal het die tempo van WOK verbruik verminder gedurende die anaërobies fase sowel as die aërobiese fase vir beide eksperimente. Kuilvoer wat bederf het as gevolg van giste en swamme was meer sigbaar by die inokulant behandelde kuilvoer as gevolg van die teenwoordigheid van suikers in die mikrosilo’s. (Sleutelwoorde: hawerkuilvoer, inokulant, mikrosilo’s, water oplosbare koolhidrate (WOK), in vitro organiese materiaal verteerbaarheid (IVOMV))

Thesis (MScAgric (Animal Sciences))--University of Stellenbosch, 2010. / ENGLISH ABSTRACT: The objective of this study was to evaluate Dohne Merino sheep and Boer goats in terms of the degradable parameters of a high-fibre diet, a low-fibre diet and two vegetable protein sources commonly used in South Africa. Differences between species were evaluated following the potential differences within species. The feedstuffs used were those for the following diets: low-fibre diet (LF); high-fibre diet (HF); sunflower meal (SFM) oilcake; and soybean meal (SBM) oilcake. In the first trial, the digestible characteristics of the HF and LF diets were determined by means of a digestibility study. A 6 × 6 Latin square design was used to determine whether Dohne Merino sheep or Boer goat wethers differ regarding the digestibility characteristics of low- and high-fibre diets. The diets were fed once daily at 1.24 kg to all the wethers, which had ad libitum access to fresh water. Each period consisted of 10 days of adaptation and seven days of faecal and urinary sampling. The results indicated that the intake and digestibility characteristics of nutrients did not differ between sheep and goats. However, the different diets differed in terms of the nutrient intake and digestibility range of sheep and goats. The second trial was an in sacco degradability trial to determine the dry matter (DM) and crude protein (CP) degradability of the LF, HF, SBM and SFM diets. Six Dohne Merino and six Boer goat wethers were fitted with rumen cannulae so that they could be used in the trial. All wethers received the same basal diet. The samples were incubated in the rumen in polyester Dacron bags, with the bags being removed at intervals of 0h, 3h, 9h, 12h, 24h, 48h, 72h, and 96h for the LF and HF diets. All the oilcake was removed at intervals of 0h, 2h, 4h, 8h, 12h, 16h, 24h, 36h and 48h. The sheep and goats were found not to differ from one another in terms of effective degradability of any of the feedstuffs concerned. However, within species differences were observed. To establish a fully integrated outcome of degradability, the study described in the current thesis was structured in such a way that the in vitro trial ran parallel with the in sacco trial, being performed with the aid of a Daisy Incubator (ANKOM Technology Corp., Fairport, NY). Such a procedure was only adopted in relation to the SFM and SBM diets in order to evaluate their in vitro data in relation to the in sacco data. The same oilcake was tested in the case of both trials, with the composite sample of rumen liquid of four sheep or goats, which was used in the in sacco trial, also being used in the in vitro study. In the study, DM disappearance values were determined and fitted to a single-compartment model by means of an iterative least-square procedure in order to determine the DM and CP degradability parameters. The DM used in vitro or in sacco was compared, using the actual values obtained after 8h incubation, due to only a limited amount of residue being left after incubation. In the study, the in vitro method overestimated the digestibility of SBM by 37% to 39% and the digestibility of SFM by 17% to 20% compared with that found to occur in the in sacco method. In vitro DM disappearance values for all SBM samples were found to be higher than those that were detected in the SFM samples. The percentage of in vitro true digestibility parameters was also calculated. No significant differences were found between species for effective degradability, though differences were observed within species between the two substrates concerned. In conclusion, the sheep and goats used in the study were not found to differ in terms of digestion parameters when they were compared on different types of roughage or protein sources. However, within species differences were, indeed, found to occur. Sheep and goats digested the SBM better than they did the SFM. / AFRIKAANSE OPSOMMING: Die doel van hierdie studie was om te bepaal of Dohne Merino skape verskil van Boerbokke in terme van degradeerbaarheidsparameters van ‘n hoë vesel-, ‘n lae veseldieët en twee plantaardige proteïenbronne wat algemeen in Suid-Afrika gebruik word. Die verskille tussen spesies is ge-evalueer en daarna die potensiële verskille binne spesies. Die volgende grondstowwe is geëvalueer: ‘n laevesel-dieët (LF), ‘n hoëvesel-dieët (HF), sonneblom-oliekoekmeel (SFM) en sojaboon-oliekoekmeel (SBM). In die eerste proef is die degradeerbaarheidsparameters van die HF dieët en die LF dieët met behulp van ‘n verteerbaarheidstudie bepaal. Dohne Merino hamels of Boerbok kapaters was gebruik om te bepaal of skape en bokke verskil in terme van inname en degradeerbaarheid van voedingstowwe wanneer hul hoë- en lae vesel voere gevoer word. Al die hamels en kapaters het ad libitum toegang tot vars water gehad en hul was een keer per dag (1.24 kg) gevoer. Elke periode het bestaan uit ‘n 10 dag aanpassingsperiode en ‘n toegelate 7 dae vir mis- en urienmonster versameling. Die resultate het aangedui dat die inname- en degradeerbaarheidsparameters van nutriënte beinvloed word deur verskillende diëte binne spesies. Geen verskille is gevind tussen spesies wanneer daar hoë- en lae kwaliteit voere gevoer is nie. Die tweede proef was ‘n in sacco-degradeerbaarheidsstudie om te bepaal wat die droë materiaal (DM) en ruproteïen (RP) verteerbaarheidsparameters van die HF dieët, die LF dieët, die SBM en die SFM is. Ses Dohne Merino’s en ses Boer bokke met rumen kanullas is in die studie gebruik en al die diere het dieselfde basale dieët ontvang. Die monsters is in die rumen geïnkubeer in poliester dakronsakkies en die sakkies is verwyder na onderskeidelik 0 uur, 3 uur, 9 uur, 12 uur, 24 uur, 48 uur, 72 uur en 96 uur intervalle. Laasgenoemde intervalle was geldig vir die lae vesel- en hoëveseldieët. Die oliekoeke se intervalle het verskil en is verwyder na 0 uur, 2 uur, 4 uur, 8 uur, 12 uur, 16 uur, 24 uur, 36 uur en 48 uur. Daar was geen verskille tussen spesies in effektiewe degradeerbaarheid nie, alhoewel verskille voorgekom het binne spesies. Skape verteer veselagtige grondstowwe meer effektief terwyl bokke weer hoë proteïn bevattende grondstowwe beter verteer. Om ‘n volkome geïntegreede uitkoms van degradeerbaarheid te bewerkstellig is die in vitro proef en die in sacco proef gelyktydig gedoen. Die in vitro-degradeerbareheidstudie is met behulp van ‘n ANKOM Daisy Inkubeerder uitgevoer (ANKOM Tegnologie Korp., Fairport, NY) vir net die oliekoek behandelings. Gedurende die studie is dieselfde oliekoeke gebruik. ‘n Saamgestelde monster van die rumenvloeistof van vier van die skape of bokke wat vir die in sacco-studie gebruik was, is gebruik vir die in vitro-inkubasie van die monsters. DM verdwyningparameters is bereken en dan met ‘n interaktiewe kleinste kwadraat prosedure op ‘n een-kompartement model gepas om die in sacco DMdegradeerbaarheidsparameters te bepaal. Die DM verdwyning, na 8h inkubasie, was gebruik om die in vitro en die in sacco metodes met mekaar te vergelyk, weens ‘n beperkte residu na die afloop van die elke inkubasiestudie. Tydens die studie het die in vitro metode degradering oorskat in vergelyking met die in sacco metode. DM verdwyningswaardes vir al die SBM monsters was hoër in vitro as die SFM monsters. In die studie is die persentasie in vitro ware degradeerbaarheidswaardes bereken. Geen verskille is opgemerk tussen spesies vir effektiewe degradeerbaarheid nie. Daar was wel verskille binne spesies. Om af te sluit het dit voorgekom dat skape en bokke nie verskil aan degradeerbaarheidswaardes wanneer daar ‘n vergelyking was tussen verskillende vesels- en proteϊenbronne nie, alhoewel verskille voorgekom het binne spesies. Skape en bokke het SBM effektief beter verteer as SFM.

La nanomédecine, appliquée en particulier au traitement du cancer, suscite depuis une quinzaine d’années un intérêt grandissant, développant des stratégies innovantes pour le ciblage spécifique de tissues malades. De nouveaux progrès en la matière sont encore à venir, mais nécessitent de nouveaux matériaux offrant une grande flexibilité en termes de synthèse, ainsi que la possibilité de fonctionnalités et de propriétés physicochimiques variées ; avantages tous présentés par l’utilisation des techniques de polymérisation radicalaire contrôlée (CRP). Ces techniques de polymérisation ont déjà démontré leur fort potentiel à travers différents systèmes nanoparticulaires à base de prodrogues de polymère, mais aucun d’entre eux ne s’avère dégradable ce qui pourrait empêcher à l’avenir leur utilisation et leur développement.Anticipant les besoins, ce projet a eu pour but la synthèse de polymères PEGylés dégradables par la technique de polymérisation contrôlée par les nitroxydes (NMP), travail très en amont de l’habituel procédé d’élaboration d’un nouveau nanomédicament. Pour ce faire, la NMP du méthacrylate de méthyl éther oligo(éthylène glycol) (MeOEGMA) a été combinée à la polymérisation radicalaire par ouverte de cycle (rROP) des acétals de cétène cyclique (CKAs), connus comme précurseurs de fonctions esters.Parmi trois CKAs étudiés, le 2-méthylène-4-phényl-1,3-dioxolane (MPDL) a montré une capacité unique à copolymériser avec les dérivés de méthacrylates, grâce à sa structure ouverte de type « styrènique » permettant son utilisation en NMP. A travers une étude approfondie des propriétés de contrôle et de caractère vivant de ces copolymères, le MPDL s’est également révélé être le premier comonomère de contrôle des méthacrylates à être dégradable. Un lien direct entre dégradabilité et quantité de MPDL insérée a été démontré, permettant jusqu’à l’hydrolyse complète des matériaux. Ces copolymères n’ont montré aucune cytotoxicité, et ce sur trois types de cellules différents (fibroblastes, cellules endothéliales et macrophages), et une étude similaire sur la toxicité de leurs produits de dégradation a permis d’aboutir à la même conclusion, soulignant la possible biocompatibilité de ces nouveaux matériaux qui, si confirmée, permettrait leur utilisation pour des applications biomédicales.Parallèlement, un second projet portant sur la mise au point d’une nouvelle alcoxyamine à base du nitroxide SG1 et présentant une fonction azlactone, baptisée AzSG1, a été développé pour la synthèse de polymères fonctionnalisables par NMP. Utilisant l’alcoxyamine AzSG1 comme amorceur, les NMPs du styrène, de l’acrylate de n-butyle et du méthacrylate de méthyle ont été réalisées avec succès, ainsi que le couplage quantitatif de la benzylamine comme preuve de concept de la possible fonctionnalisation. Dans un avenir proche, utiliser cet amorceur fonctionnalisable pour la synthèse de copolymères à base de MeOEGMA et de MPDL pourrait ainsi permettre l’élaboration de copolymères PEGylés, dégradables et fonctionnalisables par NMP, pour des applications dans le domaine de la bioconjugaison et du drug delivery. / Nanomedicine, especially for cancer treatment, has attracted much interest over the last fifteen years, developing innovative strategies for targeting diseased tissues. Further improvements of these approaches will require advanced new materials affording versatility, functionalities and specific physico-chemical properties, all advantages offered by the controlled radical polymerization (CRP) techniques. These radical polymerizations already showed their great potential through various efficient anticancer polymer nanocarriers but all lacking of degradability, which may hinder any actual developments.Anticipating the needs, this project focused on the synthesis of degradable PEG-based polymers by nitroxide-mediated polymerization (NMP), as an early stage in the usual process of nanocarrier design. To do so, NMP of oligo(ethylene glycol) methyl ether methacrylate (MeOEGMA) has been for the first time combined to the radical ring-opening polymerization (rROP) of various cyclic ketene acetals (CKAs), known as ester precursors.Among three CKAs tested, 2-methylene-4-phenyl-1,3-dioxolane (MPDL) has shown a unique ability to copolymerize with methacrylate derivatives, likely due to a styrene-like open structure allowing for its use in NMP. Through a careful study of the control and livingness properties of these copolymers, MPDL was also demonstrated to be the first degradable controlling comonomer for polymethacrylate synthesis. The degradability of the resulting PEG-based copolymers was proven to be proportional to the adjustable amount of MPDL inserted, up to complete degradation. These copolymers showed no cytotoxic effect on various cell types (fibroblasts, endothelial cells and macrophages), and an additional study of the innocuousness of their degradation products led to similar results, underlining their potential biocompatibility which, if confirmed, would allow these materials to be used for biomedical applications.A second project about a new azlactone-functionalized SG1-based alkoxyamine (AzSG1) was also set up, as initiator for the synthesis of functionalizable polymers by NMP. Using the AzSG1 alkoxyamine, the NMP of styrene, n-butyl acrylate and methyl methacrylate were successfully performed, as well as a quantitative coupling of benzylamine as proof of concept. In the near future, making use of this functionalizable initiator for copolymerizing MeOEGMA with MPDL may allow the easy synthesis of functionalized degradable copolymers by NMP, for bioconjugation and drug delivery applications.

A inoculação de forrageiras com fungos lignocelulolíticos é uma alternativa para melhorar a qualidade destas sem adição de substâncias químicas. Este projeto visa melhorar a degradabilidade de seis forrageiras: Brachiaria decumbens cv. Basilisk, Pennisetum purpureum Schum. cv. Napier, Panicum maximum cv. Aruana, Cenchrus ciliares cv. Buffel, bagaço de cana-de-açúcar e cana-de-açúcar picada por meio do cultivo de quatro fungos do gênero Pleurotus. Outrossim, extrato enzimático, nas concentrações de 2, 4 e 6 mL, produzido pelo fungo P. sajor-caju CCB 020 crescido em vinhaça, durante 6º, 12º e 18º dias, foram aplicados às mesmas forrageiras. As avaliações das dietas foram pela técnica de produção de gases, degradabilidade in vitro e análise bromatológica do substrato. As atividades das enzimas Lacase, Peroxidase, Manganês Peroxidase, Endoglucanase, Exoglucanase e Xilanase foram determinadas, por um período de 24 dias. Os resultados foram estatisticamente avaliados, utilizando SAS® (Statistical Analysis System Inst., Cary, North Carolina). Os fungos crescidos nas forrageiras, produziram enzimas hidrolíticas e oxidativas durante o processo de fermentação, que atuaram na transformação das forrageiras. A quantidade de proteínas aumentou significativamente na cana-de-açúcar picada e capim Buffel, inoculados com os fungos P. ostreatus e P. albidus CCB 068. A forrageira Aruana foi a que melhor respondeu ao tratamento com fungo, sendo que com P. sajor-caju, ao 18º dia de incubação, teve a concentração de acetato 1,24 vezes maior, em relação a amostra controle, e as demais concentrações de AGCC, como propionato e butirato, diminuir e a razão C2:C3 aumentou. A cana-de-açúcar incubada com P. albidus CCB 068, ao 18º dia, diminuiu as concentrações de acetato, propionato e butirato e aumentou a razão C2:C3 em 0,86 - 0,58 - 0,71 - 1,48 vezes, respectivamente. O bagaço inoculado ao 24º dia com P. albidus provocou aumento nas concentrações de acetato, propionato e butirato e diminuição na razão C2:C3, em 1,14 - 1,75 - 1,32 - 0,64 vezes, respectivamente, e com P. ostreatus mostrou o mesmo comportamento. Com os demais tratamentos nenhum efeito significativo foi observado, pelo teste de Tukey a 5%, entretanto ocorre tendência de aumentar a razão C2:C3 com o tratamento com os fungos. Na produção de gases e degradabilidade in vitro das forrageiras, utilizando os extratos enzimáticos, ocorreu aumento com a concentração aplicada. Efeitos significativos foram observados para as forrageiras Brachiaria, Napier e Aruana, já as forrageiras Buffel, bagaço de cana-de-açúcar e cana-de-açúcar picada não mostraram diferenças significativas, pelo teste de Tukey a 5%. Diferenças nas análises de AGCC não foram significativas, pelo teste de Tukey a 5%, dentro das concentrações ou período de incubação do fungo, nas seis forrageiras, entretanto há uma interação com as atividades das enzimas e pela razão C2:C3 a maior concentração testada de 6 mL, do extrato com o fungo, incubado durante 12º dias, possam ser recomendadas para obtenção de ganho nutricional das dietas. Conclui-se que tanto os fungos inoculados nas forrageiras, como seus extratos enzimáticos foram capazes de modificar a composição bromatológica original das forrageiras. A degradabilidade, ganho nutricional e sustentabilidade ambiental, podem variar substancialmente para os diferentes tipos de forrageiras, espécie de fungo e tempo de incubação. Os resultados obtidos sugerem que os Pleurotus, é um fungo apropriado para melhorar o valor nutritivo das forragerias como alimento para ruminates, melhorando a composição bromatoligica, mas também aumentando a degradabilidade / The inoculation of forages with lignocellulolytic fungi is an option for improving quality without adding chemical products. This project aims to improve the degradability of six forages: Brachiaria decumbens cv. Basilisk, Pennisetum purpureum Schum. Cv. Napier, Panicum maximum cv. Aruana, Cenchrus ciliares cv. Buffel, sugarcane bagasse and chopped sugarcane by cultivating four fungi of the genus Pleurotus. In addition, enzymatic extract, at concentrations of 2, 4 and 6 mL, produced by P. sajor-caju CCB 020, developed in vinasse, during 6, 12 and 18 days were applied to the same forages. The evaluations of the diets were by in vitro technique of gas production and bromatological analysis of the substrate. The activities of the enzymes Lacase, Peroxidase, Manganese Peroxidase, Endoglucanase, Exoglucanase and Xylanase were determined, for a period of 24 days. The results were statistically evaluated by SAS® analysis (Statistical Analysis System Inst., Cary, North Carolina). The fungi grown in the forages, produced hydrolytic and oxidative enzymes during the fermentation process that worked in forage transformation. The amount of proteins increased significantly in the chopped sugarcane and Buffel, inoculated with P. ostreatus and P. albidus CCB 068 fungi. The Aruana grass was the one that responded better to fungi treatments. With P. sajor-caju at the 18th day of incubation, the concentration of acetate was 1.24 times higher than the control sample, and the concentrations of other short chain fatty acids (SCFA), such as propionate and butyrate, tended to decrease and the C2:C3 ratio increased. Sugarcane incubated with P. albidus CCB068 at day 18 decreased acetate, propionate and butyrate concentrations and increased the C2:C3 ratio in 0.86, 0.58, and 0.71 - 1.48 fold, respectively. The bagasse inoculated at the 24th day with P. albidus caused an increase in acetate, propionate and butyrate concentrations and a decrease in the C2:C3 ratio in 1.14, 1.75, and 1.32 - 0.64 fold, respectively, and with P. ostreatus showed the same behavior. With the other treatments, no significant effect was observed by the Tukey test. However, there is a tendency to increase the C2:C3 ratio with the fungus treatment. In the production of gases and in vitro degradability of forages, using the enzymatic extracts, an increase with the applied concentrations occurred. Significant effects were observed for the Brachiaria forage at 5%, while forage Buffel, sugarcane bagasse and chopped sugarcane did not show significant differences by Tukey\'s test at 5%. Differences in the SCFA analyzes were not significant, by the Tukey\'s test at 5%, within the concentrations or incubation period of the fungus, in the six forages studied. However there is an interaction with the activities of the enzymes and the C2:C3, the highest concentration tested (6 mL) of the fungi extracts, incubated for 12 days, can be recommended to obtain increase in nutritional value of the diets. It is concluded that both the fungi inoculated on the forage and their enzymatic extracts were able to modify the original bromatological composition of the forage. The degradability, increase of nutritional value and environmental sustainability, can vary substantially for different types of forages, species of fungus and incubation time. The results suggest that Pleurotus is an appropriate fungus for improving the nutritive value of forage crops as feed for ruminates, improving the bromatoligic composition, but also enhancing the degradability