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Abstract Summary Recent research has demonstrated that using antioxidants to overcome the occurrence of skin pathologies is a very promising approach. Several delivery systems have been exploited for such purpose, among which are lipid nanoparticles as solid lipid nanoparticles (SLNs), nanostructured lipid carriers (NLCs), as well as vesicular systems as liposomes, niosomes, transfersomes and ethosomes. Nanostructured lipid carriers (NLCs) are lipid nanoparticles consisting of mixtures of solid and fluid lipids, in which the fluid lipid phase is embedded into the solid lipid matrix. On the other hand, vesicular systems as aspasomes consist of a bilayer of ascorbyl palmitate and cholesterol enclosing an aqueous environment. NLCs are widely used in cosmetics enabling drug penetration into epidermal barrier compared to other delivery systems to prevent skin aging. Melatonin was reported to be an effective anti-oxidant. It acts as a free radical scavenger and it has been reported to inhibit the lipid peroxidation in skin. Therefore, the aim of this work was to prepare a highly antioxidant lipid based/vesicular nanosystems for the treatment of androgenic alopecia caused by oxidative stress. The work in this thesis is divided into three chapters: Chapter I: Preparation and characterization of highly anti-oxidant lipid nano particulate system of melatonin (NLCs) This chapter deals with the formulation, and characterization of melatonin- NLCs. These NLCs were prepared by high shear homogenization method followed by solvent diffusion, and characterized for their entrapment efficiency, morphology using transmission electron microscopy, particle size, Summary 160 zeta potential, in vitro permeation, anti-oxidant potential and ex-vivo deposition. Stability of melatonin- NLCs stored at 2-8oC was tested through monitoring the changes in particle size and zeta potential. The work in this chapter included the following: 1- Preparation of melatonin-NLCs by combined solvent evaporation/high shear homogenization. Four different oils (liquid lipids) namely evening primrose oil, olive oil, soybean oil and almond oil were used. Investigation of the factors influencing the EE%, particle size and the anti-oxidant activity of melatonin-NLCs namely, the effect of the type of solid lipid used and the type of liquid lipid (oils) used was performed. 2- characterization of the prepared NLCs through the following studies: a) Particle size, PDI and zeta potential analysis of freshly prepared melatonin NLCs formulations were carried out using zetasizer. b) Determination of melatonin entrapment efficiency EE% in NLCs by using exhaustive dialysis method. c) Measurement of the anti-oxidant potential of NLCs formulations was carried out using the DPPH free radical assay. d) Physical stability study on all melatonin NLCs formulations was conducted over a storage period of three months at 2-8° C by the assessment of the effect of storage on the particle size, charge and PDI of the nanoparticles. e) In vitro permeation experiments were carried out over 6 hours on the selected formulae. Summary 161 f) selected NLC formulation was examined for morphology by transmission electron microscopy. g) Ex vivo deposition/permeation of the selected NLC formula on rat skin was carried out using Franz diffusion apparatus. The results of this work revealed the following: 1- Melatonin NLCs were successfully prepared using combined high shear homogenization/ solvent diffusion method. 2- The EE% of melatonin in NLCs ranged from 36 to 76% owing to its hydrophobicity, suggesting its successful incorporation within the liquid lipid matrix. 3- NLCs particle size ranged from 303-783 nm, depending on the type of solid lipid and liquid lipid used. 4- All of the prepared NLCs were charged, with charges ranging from (-11.8 to -19.7 mV). 5- Melatonin NLC formulae were supposed to be a successful anti-oxidant system. 6- The selected melatonin NLCs formulations showed sustained release of melatonin for 6 hours, and 100% release was achieved after this period of time. 7- Melatonin NLCs displayed good storage properties as manifested by the slight changes in particle size, polydispersity index and zeta potential values. 8- Transmission electron microscopy displayed coarsely and somehow spherical shaped NLCs with smooth surface 9- Ex vivo skin deposition experiments demonstrated the high potential of the selected NLC formula (NLC2) in accumulating the drug into the Summary 162 deeper epidermal and dermal layers of the skin, and hence it was selected for further clinical studies. Chapter 2: Preparation and characterization of highly anti-oxidant vesicular system of melatonin (aspasomes) This chapter deals with the formulation and characterization of melatonin aspasomes prepared by thin film hydration technique. characterization of the prepared vesicles was done through the following studies: entrapment efficiency, particle size, zeta potential, morphology using transmission electron microscopy, in vitro permeation, anti-oxidant potential and deposition experiments. Stability study of melatonin-aspasomes stored at 2- 8oC was tested through monitoring the changes in particle size and zeta potential. The work in this chapter included the following: 1- Preparation of melatonin aspasomes by thin film hydration method using ascorbyl palmitate (ASP) and cholesterol in varied molar ratio with a negative charge inducer (dicetyl phosphate). 2- characterization of the prepared aspasomes was done through the following studies: a) Particle size, PDI and zeta potential analysis of freshly prepared melatonin aspasomal formulations were carried out using zetasizer. b) EE% of melatonin in aspasomes. c) Physical stability study on all melatonin aspasomes formulations was conducted over a storage period of three months at 2-8° C by the assessment the effect of storage on the particle size, charge and PDI of the nanoparticles. d) Measurement of the anti-oxidant potential of the selected melatonin aspasome system using DPPH assay. Summary 163 e) In vitro release study on the selected melatonin aspasomal formulae. f) The selected aspasomal formula was examined for morphology by transmission electron microscopy g) Ex vivo deposition/permeation of the selected aspasomal formula on rat skin was carried out using Franz diffusion apparatus. The results of this work revealed the following: 1- Melatonin aspasomes were successfully prepared using the thin film hydration method. 2- The EE% of melatonin in aspasomes ranged from 52.23 to 91.21 % owing to the hydrophobicity of the drug, suggesting its successful incorporation within the lipid bilayers. 3- Aspasomes particle size ranged from 287- 950 nm, depending on the amount of ascorbyl palmitate. 4- All of the prepared aspasomes were charged, with charges ranging from (-37.3 to -63.3) mV. 5- selected melatonin formulae systems were supposed to be a successful anti-oxidant system. 6- selected melatonin aspasomes’ formulations showed sustained release of melatonin for 6 hours. 7- Melatonin aspasomes displayed good storage properties as manifested by the slight changes in particle size, polydispersity index and zeta potential values. 8- Transmission electron microscopy displayed spherical vesicles with smooth surface with an evident bilayer. 9- Ex vivo skin deposition experiments demonstrated the high potential of the selected aspasomal formula (A3) in accumulating the drug Summary 164 into the deeper epidermal and dermal layers of the skin, and hence it was selected for further clinical studies. Chapter 3: Clinical evaluation of the selected NLCs and aspasomes formulae In this study attempts were made to examine the clinical efficacy of the selected melatonin NLCs and aspasomes in treatment of male AGA compared to the conventional melatonin solution. The work in this chapter included the following: The clinical study was conducted on 60 male patients suffering from AGA. Patients were divided into 3 groups: group I (20 patients) were treated with melatonin solution in phosphate buffer saline, pH 7.4, group II (20 patients) were treated with topically applied nanostructured lipid carriers formula NLC2 and group III (20 patients) were treated with topically applied aspasomal formula A3 formula. In each group, all patients were instructed to apply the provided formula once daily on affected scalp areas for a total period of 3 months. Assessment of the clinical response was done at 16th week (i.e. 1 month after the end of treatment) based on demographical and diagnostic data. The results of this work revealed the following: 1- Regarding photographic assessment, group II patients treated with NLCs formulation displayed significantly better improvement scores compared to group III patients treated with aspasomal formulation and group I treated with melatonin solution. 2- Regarding hair pull test assessment, after treatment, the extent of decrease in hair loss was significantly higher in group II patients, followed by group III then group I. Summary 165 3-Regarding hair shaft diameter histometric assessment, after treatment, the extent of hair diameter increase was significantly higher for group II patients, followed by group III patients, followed by group I patients. 4- Regarding dermoscopic evaluation, in post-treated patients of AGA, it was found to be significantly higher for group II patients followed by group III patients and finally group I patients. from the previous results, it can be concluded that melatonin delivery systems adopted in the current thesis (NLCs and aspasomes) are promising topical delivery systems for the delivery of the |