The development of transfersomes containing Eulophia macrobulbon extract

Abstract

Transferosomes were vesicular delivery systems used to carry drugs with wide range of water solubility due to amphiphilic properties. The Eulophia macrobulbon (EM) extract contained the active ingredient called Compound 1 having PDE-5 inhibitor. Therefore, the incorporation of the EM extract in form of transferosomes as topical formulation to improve skin permeability of EM extract was interested. The objectives of this studies were to (1) develop transdermal formulation of EM extract loaded transferosomes; (2) to investigate the skin permeability of EM extract in transferosomes and stability of formulations. The ideals of formulation were small vesicle, high entrapment efficiency, high flexibility and good stability. Transferosomes containing EM extract were prepared by film hydration method. To constructed appropriate transferosomes as drug delivery systems, the affecting parameters on the formulation such as phospholipid types [hydrogenated phosphatidylcholine (HPC, transition temperature (Tm) = 50°C) and soy phosphatidylcholine (SPC, Tm = 50°C)]; ratios of EM extract to phospholipid (1:1, 1:2 and 1:4 w/w); HLBs of edge activator (4.3-15.0); hydration mediums (pH 6.8 phosphate buffer and deionized water) were studied. The efficiency of drug entrapment, drug penetration and drug stability were quantified using high performance liquid chromatography (HPLC). Size distribution, zeta potential, deformability of transferosomes were investigated using ZetaPALS®. Drug permeability through skin was determined using Franz diffusion cell (Strat-M® as membrane). Moreover, the skin permeabilities of EM extract of transferosomes versus solutions were compared. The larger sizes of transferosomes prepared from HPC (1897.50 ± 88.38 nm) than SPC (346.40 ± 19.40 nm) were shown. The appropriate ratio of EM extract to phospholipid was 1:2 w/w. The variation of proportions of edge activators (Span 80 and Tween 80) could provide HLBs of edge activators in the range of 4.3 to 15.0. Increasing HLB of edged activator increased the vesicle size (~350 to ~900 nm), while HLB did not affect size of transferosomes after extrusion.

The transferosome formulations composed of SPC phospholipid; EM extract : SPC ratio as 1:2 w/w; and concentration of edge activator to SPC as 15:85 % w/w with HLBs of edge activator ( 4.3, 9.65 and 15.0) were reduced size using extrusion method. These formulations were extruded pass through 0.4 µm pore size of membrane. The vesicle sizes of all transferosomes were comparable (approximately 200 nm). The deformability determination of transferosomes sized ~200 nm was performed using 0.1 µm pore size of membrane, then transferosomes with vesicle size ~100 nm were obtained. The deformability of these transferosomes with different HLBs (4.3, 9.65 and 15.0) of edge activator were approximately 50%. The deformability of transferosomes with increasing HLB of edge activator were similar. Increasing the concentration of edge activator slightly increased the vesicle size (779.00 ± 15.60 to 957.00 ± 45.30 nm). No effect of hydration mediums on transferosome characteristics was presented. The entrapment efficiency and zeta potential of all formulation were the same (approximately 90% and -5.0 mV, respectively). Interestingly, no effect of HLBs (4.3, 9.65 and 15.0) and transferosome sizes on skin permeability were shown in this study. Transferosomes provided better skin permeability than solutions. The influence of drug loading on skin permeability was presented. Increasing EM loading (5 and 10% w/w) in transferosomes could shorten lag time of drug permeability through the membrane, when compared with 2.5% w/w EM loading transferosomes.

The transferosomes and solutions were physically and chemically stable for 12 weeks in both 4 and 40 °C storage conditions. Transferosomes showed small aggregation, however it became homogenous dispersion after shaking. Size and zeta potential were not change after 12 weeks storage. No change of skin permeability of 10 weeks stored and freshly prepared transferosomes was resulted. In conclusion, EM extract load transferosomes were successfully developed. Formulation parameters could affect the properties and skin permeability of transferosomes. The high transition temperature (Tm) of phospholipid effected the rigidity of vesicle and this probably reduced the skin permeability of transferosomes. An increase of HLB of edge activator enlarged the vesicle size due to more entrapment of other components and hydrophilic part. Transferosomes prepared from SPC provided better skin permeability compared to solutions.

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