MLX-NLC gel stored at different temperature and humidity conditions (Table 1) were evaluated for any changes in particle size, PI, zeta potential, and drug content at 30, 60, 90 days time points to assess the effect of storage conditions on the stability as a function of time.
The particle size was monitored at regular time intervals during storage to assess particle aggregation. The nanoparticles are thermodynamically unstable system and for their stability, a zeta potential value between –30 mV and –60 mV is desirable to avoid aggregation of particles (Muller et al., 2000). Thus, zeta potential measurement gives us the assessment of the storage stability of nanoparticles. In addition, Tween 80 could improve the stability of the system due to its steric effect that increases the repulsion between particles and prevents their aggregation (Lim and Kim 2002). Stability of MLX-NLC gel was evaluated in terms of their MLX entrapment efficiency. Log (E.E %) was plotted against time and the slopes (m) were calculated by linear regression. The slopes (m) were then substituted into the following equation for the determination of k values:
k "= m × 2.303"
Shelf life values (the time for 10% loss, i.e. t90) were then calculated by the following equation:
t90 "= 0.105/k"
In case of MLX-NLC gel, no significant (p>0.05) change of particle size, PI, zeta potential, drug entrapment efficiency was observed at 4±2°C over the period of 90 days (Table 3). An increasing trend of particle size and PI, decreasing trend of zeta potential and drug entrapment efficiency were observed with storage time at 25±2°C/60%±5% RH and 40±2°C/75±5% RH. The augmentation of particle size and polydispersity index, and diminution of zeta potential and drug entrapment efficiency were comparatively higher at 40±2°C/75%±5% RH. This demonstrated the physical stability of the MLX-NLC gel at 4±2°C over the period of 90 days.
The results are in agreement with previous results (Junyaprasert et al., 2009) suggesting the good stability of MLX-NLC gel. Upon storage at high temperature, the change in the particle size in MLX-NLC gel can be explained by the fact that the increase in the kinetic energy of system at high temperature may accelerate the collision between particles, and consequently can increase the chances of aggregation of particles (Freits and Muller, 1998).
This decrease in entrapment efficiency at high temperature could also be attributed to some degree of lipid transformation of the solid lipid used in NLC over time, leading to the formation of a highly ordered lipid structure resulting in drug expulsion.
Shelf life values i.e. the time at which the drug concentration is lost by 10% were calculated. A good chemical stability of MLX-NLC gel (t90, 281.90days) was observed when stored at 4±2°C It is evident from the data MLX-NLC gel had good long-term physical and chemical stability at 4±2°C and the recommended storage temperature is 4±2°C.
3.6 Toxicity study