The most popular and convenient route of drug administration remains the oral route, however, protein and peptide drugs such as insulin have poor membrane permeability and stability in the gastrointestinal tract. Absorption enhancers can be added to drug delivery systems to overcome the epithelial cell membrane permeability problem. Although previous studies have shown that aloe leaf materials improve the transport of drugs across intestinal epithelia, their performance in solid oral dosage forms has not yet been investigated. Beads containing insulin and each of the selected absorption enhancers (i.e. Aloe ferox, Aloe marlothii and Aloe vera gel materials) were produced by extrusion-spheronisation, using a full factorial design to optimise the formulations based on transepithelial electrical resistance (TEER) reduction of Caco-2 cell monolayers as response. The optimum bead formulations were evaluated in terms of friability, mass variation, particle surface texture, shape, size and dissolution. The transport of insulin across excised pig intestinal tissue from the optimised bead formulations was determined over a 2 h period. The samples obtained from the transport studies were analysed for insulin content by means of high-performance liquid chromatography (HPLC). The results showed that the TEER reduction, as an indication of tight junction modulation, obtained for the bead formulations containing aloe materials was concentration dependent. Furthermore, inclusion of croscarmellose sodium (Ac-di-sol®) as a disintegrant showed an enhanced TEER reduction effect in combination with the aloe gel materials. Dissolution profiles indicated that the beads containing aloe leaf materials in conjunction with insulin, released the insulin within an hour. In accordance with the TEER reduction results, the A. marlothii and A. vera materials containing beads showed similar increased insulin delivery across excised pig intestinal tissue, which was pronouncedly higher than that of the control group (insulin alone). It can be concluded that beads containing aloe leaf materials have high potential as effective delivery systems for protein therapeutics such as insulin via the oral route of administration. / MSc (Pharmaceutics), North-West University, Potchefstroom Campus, 2015

The most popular and convenient route of drug administration remains the oral route, however, protein and peptide drugs such as insulin have poor membrane permeability and stability in the gastrointestinal tract. Absorption enhancers can be added to drug delivery systems to overcome the epithelial cell membrane permeability problem. Although previous studies have shown that aloe leaf materials improve the transport of drugs across intestinal epithelia, their performance in solid oral dosage forms has not yet been investigated. Beads containing insulin and each of the selected absorption enhancers (i.e. Aloe ferox, Aloe marlothii and Aloe vera gel materials) were produced by extrusion-spheronisation, using a full factorial design to optimise the formulations based on transepithelial electrical resistance (TEER) reduction of Caco-2 cell monolayers as response. The optimum bead formulations were evaluated in terms of friability, mass variation, particle surface texture, shape, size and dissolution. The transport of insulin across excised pig intestinal tissue from the optimised bead formulations was determined over a 2 h period. The samples obtained from the transport studies were analysed for insulin content by means of high-performance liquid chromatography (HPLC). The results showed that the TEER reduction, as an indication of tight junction modulation, obtained for the bead formulations containing aloe materials was concentration dependent. Furthermore, inclusion of croscarmellose sodium (Ac-di-sol®) as a disintegrant showed an enhanced TEER reduction effect in combination with the aloe gel materials. Dissolution profiles indicated that the beads containing aloe leaf materials in conjunction with insulin, released the insulin within an hour. In accordance with the TEER reduction results, the A. marlothii and A. vera materials containing beads showed similar increased insulin delivery across excised pig intestinal tissue, which was pronouncedly higher than that of the control group (insulin alone). It can be concluded that beads containing aloe leaf materials have high potential as effective delivery systems for protein therapeutics such as insulin via the oral route of administration. / MSc (Pharmaceutics), North-West University, Potchefstroom Campus, 2015

Surfactants are molecules that contain a water-soluble and afat-soluble part. They have important functions in productssuch as detergents, cosmetics, pharmaceuticals and foods aswell as in many industrial processes. Surfactants are used onvery large scale, which makes it important to decrease theirimpact on the environment. This can be done by starting withnatural materials, by improving the synthetic methods and byreducing the use of limited resources such as energy andorganic solvents. This thesis focuses on lipase-catalyzed synthesis ofsurfactants based on natural products. It also includesfunctional studies of the produced surfactants; as antioxidantsin oils, or as surfactants to solubilize pharmaceuticals. Unsaturated fatty acid esters of ascorbic acid weresynthesized with catalysis by Candida antarctica lipase B int-amyl alcohol and in ionic liquids. High yields ofascorbyl oleate were obtained in an ionic liquid that wasdesigned to improve the solubility of the fatty acid, when thereaction was performed under vacuum. Ascorbyl oleate wasamorphous and was a better antioxidant than ascorbyl palmitatein rapeseed oil. Polyethylene glycol (PEG) stearate, PEG 12-hydroxystearateand a series of PEG 12-acyloxy-stearates were synthesized in avacuum-driven, solvent-free system usingC. antarcticalipase B as catalyst. Critical micelleconcentration and solubilization capacity were determined forthe PEG 12-acyloxy-stearates. Their effects on living cellswere evaluated in studies of hemolysis and transepithelialelectrical resistance. Several PEG1500 12-acyloxy-stearateswere excellent solubilizers for pharmaceutical use and hadnegligible negative effects on living cells even at highconcentrations. Enzymatic and chemo-enzymatic methods offer uniquepossibilities to synthesize surfactants of high purity. Pureand well-defined surfactants enable new applications and areimportant for the understanding of surfactantstructure-function relationships.

<p>Surfactants are molecules that contain a water-soluble and afat-soluble part. They have important functions in productssuch as detergents, cosmetics, pharmaceuticals and foods aswell as in many industrial processes. Surfactants are used onvery large scale, which makes it important to decrease theirimpact on the environment. This can be done by starting withnatural materials, by improving the synthetic methods and byreducing the use of limited resources such as energy andorganic solvents.</p><p>This thesis focuses on lipase-catalyzed synthesis ofsurfactants based on natural products. It also includesfunctional studies of the produced surfactants; as antioxidantsin oils, or as surfactants to solubilize pharmaceuticals.</p><p>Unsaturated fatty acid esters of ascorbic acid weresynthesized with catalysis by Candida antarctica lipase B in<i>t</i>-amyl alcohol and in ionic liquids. High yields ofascorbyl oleate were obtained in an ionic liquid that wasdesigned to improve the solubility of the fatty acid, when thereaction was performed under vacuum. Ascorbyl oleate wasamorphous and was a better antioxidant than ascorbyl palmitatein rapeseed oil.</p><p>Polyethylene glycol (PEG) stearate, PEG 12-hydroxystearateand a series of PEG 12-acyloxy-stearates were synthesized in avacuum-driven, solvent-free system using<i>C. antarctica</i>lipase B as catalyst. Critical micelleconcentration and solubilization capacity were determined forthe PEG 12-acyloxy-stearates. Their effects on living cellswere evaluated in studies of hemolysis and transepithelialelectrical resistance. Several PEG1500 12-acyloxy-stearateswere excellent solubilizers for pharmaceutical use and hadnegligible negative effects on living cells even at highconcentrations.</p><p>Enzymatic and chemo-enzymatic methods offer uniquepossibilities to synthesize surfactants of high purity. Pureand well-defined surfactants enable new applications and areimportant for the understanding of surfactantstructure-function relationships.</p>

Introduction Oral delivery of proteins faces numerous challenges due to their enzymatic susceptibility and instability in the gastrointestinal tract. In recent years, the polyelectrolyte complexes have been explored for their ability to complex protein and protect them against chemical and enzymatic degradation. However, most of the conventional binary polyelectrolyte complexes (PECs) are formed by polycations which are associated with toxicity and non-specific bio-interactions. The aim of this thesis was to prepare a series of ternary polyelectrolyte complexes (APECs) by introduction of a polyanion in the binary complexes to alleviate the aforementioned limitations. Method Eight non-insulin loaded ternary complexes (NIL APECs) were spontaneously formed upon mixing a polycation [polyallylamine (PAH), palmitoyl grafted-PAH (Pa2.5), dimethylamino-1-naphthalenesulfonyl grafted-PAH (Da10) or quaternised palmitoyl-PAH (QPa2.5)] with a polyanion [dextran sulphate (DS) or polyacrylic acid (PAA)] at 2:1 ratio, in the presence of ZnSO4 (4μM). A model protein i.e., insulin was added to a polycation, prior to addition of a polyanion and ZnSO4 to form eight insulin loaded (IL) APECs. PECs were used as a control to compare APECs. The complexes were characterised by dynamic light scattering (DLS) and transmission electron microscope (TEM). In vitro stability of the complexes was investigated at pH (1.2-7.4), temperature (25˚C, 37˚C and 45˚C) and ionic strength (NaCl-68mM, 103mM and 145mM). Insulin complexation efficiency was assessed by using bovine insulin ELISA assay kit. The in vitro cytotoxicity was investigated on CaCo2 and J774 cells by MTT (3-4,5 dimethyl thialzol2,5 diphenyl tetrazolium bromide) assay. All complexes were evaluated for their haemocompatibility by using haemolysis assay, oxidative stress by reactive oxygen species (ROS) assay and immunotoxicity by in vitro and in vivo cytokine generation assay. The potential of the uptake of complexes across CaCo2 cells was determined by flow cytometry and fluorescent microscopy. The underlying mechanism of transport of complexes was determined by TEER measurement, assessment of FITC-Dextran and insulin transport across CaCo2 cells. 15 Results NIL QPa2.5 APECs (except IL QPa2.5-DS) exhibited larger hydrodynamic sizes (228-468nm) than all other APECs, due to the presence of bulky quaternary ammonium moieties. QPa2.5 APECs exhibited lower insulin association efficiency (≤40%) than other APECs (≥55%) due to a competition between the polyanion and insulin for QPa2.5 leading to reduced association of insulin in the complexes. DS based APECs generally offered higher insulin association efficiency (≥75%) than PAA based APECs (≤55%) due to higher molecular weight (6-10kDa) of DS. In comparison to other complexes, Pa2.5 PECs and APECs were more stable at varying temperature, ionic strength and pH due to the presence of long palmitoyl alkyl chain (C16) which reduced the chain flexibility and provided stronger hydrophobic association. The cytotoxicity of polycations on CaCo2 and J774 cells is rated as PAH>Da10=Pa2.5>QPa2.5. The introduction of PAA in Pa2.5 and Da10 brought most significant improvement in IC50 i.e., 14 fold and 16 fold respectively on CaCo2 cells; 9.3 fold and 3.73 fold respectively on J774 cells. In comparison to other complexes, Da10 (8mgml-1) induced higher haemolytic activity (~37%) due to a higher hydrophobic load of 10 percent mole grafting of dansyl pendants. The entire range of APECs displayed ≤12% ROS generation by the CaCo2 cells. The degree of in vitro TNFα production (QPa2.5≥Da10≥Pa2.5=PAH) and in vitro IL-6 generation (QPa2.5≥Pa2.5=PAH≥Da10) by J774 cells established an inverse relationship of cytotoxicity with the cytokine generation. Similar to MTT data, the introduction of PAA in APECs brought more significant reduction in in vitro cytokine secretion than DS based APECs. Pa2.5-PAA brought the most significant reduction in both in vitro and in vivo cytokine generation. All the formulations were able to significantly reduce original TEER, however did not demonstrate appreciable paracellular permeation of a hydrophilic macromolecular tracer of paracellular transport i.e., FITC Dextran. The uptake study revealed internalisation of APECs predominantly by a transcellular route. Transcellular uptake of IL QPa2.5 (≤73%), IL QPa2.5-DS (67%) was higher than their NIL counterparts, whereas the uptake of NIL Pa2.5 (≤89%), NIL Pa2.5-PAA (42%) was higher than their IL counterparts. Conclusion In essence, amphiphilic APECs have shown polyanion dependent ability to reduce polycation associated toxicity and they are able to facilitate transcellular uptake of insulin across CaCo2 cells.