## 3 Results and Discussion
### Permeation  
The permeation of drug across biologic membranes is influenced by various factors which include physicochemical properties of drug, physiological properties of the barrier and its composition, as well as transport route selected for the drug permeation [4; 12]. The release results obtained using cellophane membrane not always fits to those results obtained using natural skin. The skin composition with hair and sweat glands and its thickness is greatly affecting the drug diffusion [12]. Moreover, the skin interacts with the drug vehicles that could disrupt its barrier function and enhance drug diffusion. Figure 7 shows the order of insulin permeation across the rat skin which was found highly influenced by the vehicle used as follows: insulin niosome emulgel \(>\) insulin niosome gel \(>\) insulin emulgel \(>\) insulin niosomes \(>\) insulin solution. However, comparison of permeation data with release data (Figure 4) signifies that there is no correlation between them. Insulin permeated across the rat skin was significantly lower compared with that released using cellulose membrane. This may be probably due to the lipophilic barrier of the skin that hindered insulin diffusion [37]. These data also suggest that the insulin diffusion happens mainly from the carriers. Careful examination of Figures 4 and 7 shows that prepared niosomes could enhance the insulin permeability by only 1.45-fold (Table 6). The SSTF of insulin from niosome preparation was only 13.17 \(\mathrm{\SIUnitSymbolMicro g}\mathrm{/}\mathrm{c}\mathrm{m}^{2}\mathrm{/}\mathrm{h}\) compared to 9 \(\mathrm{\SIUnitSymbolMicro g}\mathrm{/}\mathrm{c}\mathrm{m}^{2}\mathrm{/}\mathrm{h}\) from insulin solution. The possible reasons for greater permeation by niosomes could be due to its potential to extract skin lipids or disruption of corneocytes [38]. On the other hand, gel prepared using sodium carboxymethyl cellulose gave SSTF of 18.39 \(\mathrm{\SIUnitSymbolMicro g}\mathrm{/}\mathrm{c}\mathrm{m}^{2}\mathrm{/}\mathrm{h}\) with 2-fold enhancement ratio which could be due to the colloidal properties of the gel and its enhanced surface area contact to the skin [17]. Insulin emulgel on the other hand showed 3.17-fold enhancements for drug permeability and is likely due to its surfactant contents that act as permeability enhancer in addition to its colloidal properties [28]. In addition, the in vitro permeability study showed superior enhancement for insulin diffusion across the rat skin for both niosome emulgel and niosome gel by 10 and 5.86-fold, respectively. Greater flux observed with niosome emulgel could be attributed to the dual effect of surfactants present in niosome formulation and emulsion preparation, which has the ability to diffuse through narrow pores [6], hence giving 10-fold increase in permeability. \begin{table}
\begin{tabular}{c c c c c} \hline \hline \multirow{2}{*}{**Formulation**} & **Zero Order** & **First-Order** & **Higuchi Model** & \multirow{2}{*}{**Order**} \\  & **(R2)** & **(R2)** & **(R2)** & **(R2)** \\ \hline Insulin solution & 0.9819 & 0.7157 & 0.9130 & Zero \\ Insulin gel & 0.9242 & 0.4780 & 0.9896 & Higuchi \\ Insulin niosome & 0.9282 & 0.5246 & 0.9850 & Higuchi \\ Insulin niosome gel & 0.9421 & 0.5786 & 0.9866 & Higuchi \\ Insulin emulgel & 0.9482 & 0.6694 & 0.9838 & Higuchi \\ Insulin niosome emulgel & 0.9236 & 0.6288 & 0.9918 & Higuchi \\ \hline \hline \end{tabular}
\end{table} 
Table 5: Release kinetics of insulin formulations across cellulose membrane. However, ribosomes only interact with skin by enhancing drug permeability via surfactant effect [38] so it gives lower permeability compared to niosome emulgels. The designed batches were also evaluated for the amount of insulin permeated through the skin membrane from prepared niosome emulgels and the effect of three independent variables on in vitro permeation of insulin is shown in Figure 8 in the form of 3D surface response plot and contour plot.