Topical delivery of clofazimine, artemisone and decoquinate utilizing vesicles as carrier system

Abstract

Artemisone, clofazimine and decoquinate are part of the MALTBRedox MRC South African University Flagship Projects, which focus on oxidant-redox drug combinations for the treatment of tuberculosis and a few other diseases. These active pharmaceutical ingredients (APIs) were chosen as a possible treatment of cutaneous tuberculosis (CTB), an uncommon and undefined disease that is often misdiagnosed (Abdelmalek et al., 2013; Baig et al., 2014; Fader et al., 2010). Currently CTB is only treated with regular oral anti-tuberculous medication, with occasional invasive procedures such as skin grafts (Yates, 2010). Artemisone, clofazimine and decoquinate have a log P of 2.49, 7.7 and 7.8, respectively (Biamonte et al., 2013; Dunay et al., 2009; Nagelschmitz et al., 2008; Steyn et al., 2011). A high log P-value indicates that the API is highly lipophilic and therefore a delivery system, namely vesicles, was chosen to improve skin permeation. Many vesicles are currently being investigated all over the world as carriers for APIs in topical delivery, though for this study liposomes, niosomes and transferosomes were selected. Dispersions containing a single API, a combination of all three APIs, as well as no API, were prepared for all three types of vesicles. Characterisation of dispersions containing 0.2%, 0.4% and 1% API was performed. Isothermal calorimetry indicated that no incompatibility occurred in the 1% API combination dispersions, except the niosome dispersion, which indicated a probable incompatibility. Encapsulation efficiency was above 85% for all 1% API dispersions. The empty vesicles depicted an average size of 154 nm, 167.5 nm and 106.3 nm for liposomes, niosomes and transferosomes, respectively. Vesicle sizes increased with increase in API concentration, whereas stability decreased. Clofazimine was found to have the most significant impact on vesicle size and stability when added as 1%, increasing the average niosome size to 2 461 nm. Viscosity was below 2 mPa.s for all 1% API dispersions, ensuring even spreadability when applied to the skin. The pH of all the dispersions were between 5—6, thus limiting skin irritation. In vitro transdermal diffusion studies were conducted on black skin, using dispersions containing 1% of all three APIs. No APIs could be detected in the receptor phase. Artemisone was not detected in the skin by means of HPLC analysis, which might be due to the fact that the concentration was below the limit of detection (LOD). The LOD for artemisone was determined at 4.42 μg/ml, whereas it was 0.042 μg/ml for clofazimine and 0.703 μg/ml for decoquinate. Higher API concentrations were present in the stratum corneum-epidermis (SCE), compared to in the epidermis-dermis (ED) for all the dispersions. Transferosomes delivered the highest concentration clofazimine into the SCE and ED, as well as the highest concentration decoquinate into the ED. The highest concentration decoquinate in the SCE, however, was obtained by the niosome dispersion. Efficacy against tuberculosis of the APIs (1%) encapsulated in vesicles was tested on strain H37Rv. All dispersions were found to be effective to some degree against the tuberculosis strain tested, with clofazimine in niosomes being the most effective with 52% growth inhibition. The least effective was decoquinate in niosomes, with only 8% inhibition. The combination dispersions delivered inhibitions of 42%, 38% and 12% for liposomes, niosomes and transferosomes, respectively. Surprisingly, it was found that the vesicle dispersions containing no APIs also presented some efficacy against the tuberculosis strain tested. New knowledge contributed to pharmaceutics by this study includes encapsulating the three APIs in liposomes, niosomes and transferosomes and successfully delivering them into the skin as proved by transdermal diffusion studies. Developing an HPLC method for the concurrent analysis of the three APIs and determining the activity of the vesicle dispersion against the specific tuberculosis strain tested also contributed new knowledge. Results indicated that decoquinate, an API never before considered for tuberculosis, does have anti-tuberculous activity. No significant increase in efficacy against the tuberculosis strain was noted when combining the three APIs in a vesicle dispersion, compared to when the APIs were incorporated separately into the vesicles, though the blank vesicles had surprisingly high activity against the specific tuberculosis strain tested.