Natural oil nano-emulsion dispersions for the topical delivery of clofazimine, artemisone and decoquinate

Abstract

Tuberculosis (TB), which is caused by Mycobacterium tuberculosis (Mtb) (Godreuil et al., 2007:1), manifests as an extra-pulmonary disease in 3.7 to 8.4% of cases, whereas cutaneous tuberculosis (CTB), which is uncommon, compromises approximately 1.0 to 1.5% of all extrapulmonary manifestations. The active pharmaceutical ingredients (APIs) chosen for this study were clofazimine (CLF), artemisone (ART) and decoquinate (DQ). APIs should possess physicochemical characteristics by containing both hydrophilic properties (to permeate to other hydrophilic skin layers to reach the circulatory system) and lipophilic qualities (to permeate the stratum corneum) (Naik et al., 2000). Penetration enhancers may be used to penetrate barrier properties of the skin to facilitate permeation. Literature suggests by using natural oils (fatty acids), the delivery of both hydrophilic and lipophilic APIs could be promoted (Williams & Barry, 2012:132) as they contain linoleic acid (C18 fatty acid) (Vermaak et al., 2011:920-933). The aim of this study was to investigate whether nano-emulsions containing natural oils would improve the topical delivery of CLF, DQ and ART, separately and in combination. The nanoemulsions were characterised by means of pH, viscosity, drug entrapment efficiency and zetapotential. A novel high performance liquid chromatography (HPLC) method was developed and validated for the simultaneous analysis of CLF, ART and DQ throughout the study during experiments. During membrane and skin diffusion studies, it was evident that no API was released from all eight nano-emulsions within the receptor fluid. The nano-emulsions containing a single API were found within the stratum corneum-epidermis (SCE) and showed similar results in the epidermisdermis (ED), where only CLF and DQ were detected; no ART was observed within the ED. It is evident with the nano-emulsions containing the combination of APIs that CLF and ART was found within the SCE, but no DQ was found within the SCE. In the ED, it was evident that CLF and DQ were detected, but no ART was found, therefore nano-emulsions could deliver APIs topically without entering the blood stream. In vitro cytotoxicity studies showed no cytotoxicity on immortalised human keratinocyte (HaCaT) cells, indicating the nano-emulsions were safe to use on human skin. From the Mtb cell line studies, all the formulations displayed %inhibition on the bacterial tuberculosis (52 – 63% inhibition). During this study, a novel high performance liquid chromatography (HPLC) validation method was developed to determine the concentration of CLF, ART and DQ simultaneously. Subsequently, the validation method was published; in addition, a literature review on nano-emulsions was written and published. Novel contributions have been made to science where the compatibility of the combined use of CLF, ART and DQ were tested during microcalorimetry, differential scanning calorimetry (DSC) and hot stage microscopy (HSM) studies. Nano-emulsions containing natural oils were developed and tested for efficacy against TB strains. In the future, this knowledge can be applied for the possible treatment of CTB in conjunction with systemic treatment. Information gathered during this study serves as basic research, which forms the basis for further studies.