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Preparation of NLCs containing vit A VitA loaded NLCs were prepared by hot hemogenation method in which particle size is reduced by cavitation, high shear forces and particle collision in and after leaving the homogenizing gap (Figure 1).13 For this purpose, vitA was dissolved in liquid oil (Octyloctanoat) and the mixture was added into melted solid lipid (Precirol). Then, the hot aqueous surfactant solution (at the same temperature with melted lipids) was added gradually into the lipid phase under homogenization (Silent crusher M, Heidolph, Nuremberg, Germany) at 20000 rpm for 45 minutes. The produced hot o/w nanoemulsion was cold down in the ambient or lower temperature resulting in the lipid phase recrystallization, and finally the NLC was formed. In this study, optimal formulation for vitA loaded NLC was investigated by changing concentrations of the aqueous surfactant [http://en.wikipedia.org/wiki/MRIP MRIP] as listed in Table 1. Figure 1 Table 1 Composition of VitA palmitate-loaded nanostructured lipid carriers Particle size analysis The average diameter and Span value of the formulations were determined using particle size analyzer (Wing SALD 2101, Shimadzo, Japan), at 25��C. The [http://www.selleckchem.com/products/Temsirolimus.html Temsirolimus] NLC dispersion was diluted with distilled water until suitable obscuration to prevent multiple scattering phenomena because of inter particle interactions. The average particle size was calculated according to the average volume diameter or DeBroukere mean in the equation below: D?[4,3]=��nidi4��nidi3 The Span value is an index helpful to evaluate the particle size distribution and is calculated applying the following equation:14 Span?=?D90%???D10%D50% Encapsulation Efficiency [http://www.selleckchem.com/products/PD-173074.html PD173074 concentration] (EE) Estimation of the loaded amounts of VitA into NLCs was carried out using HPLC (Knauer, Germany) equipped with a UV detector. Detection was carried out at 325 nm. Column used was C-18 (10?m 25mm��4.6 mm). The mobile phase consisted of Acetonitrile-methanol (70:30%, v/v) and the flow rate was set at 2.0 ml/min. HPLC linearity was calculated using various concentrations of vitA (0.5, 1, 2, 3, 4 ?l) in each ml of the mobile phase. To extract lipid nanodispersion solution for injecting into the HPLC machine, first the samples were gently centrifuged (600 rpm, 10min) and then 0.5 ml of chloroform was added to one ml of transparent lipid nanodispersion, and finally the product was shaken for 15 min. Then the chloroform phase including encapsulated vitamin was extracted and dried using nitrogen gas. In the last stage, 0.5 ml of the mobile phase was added and the EE was calculated using the following formula: EE?(%)?=?Amount?of?vitA?incorporated?into?NLCAmount?of?total?vitA?��?100 Fourier transform infrared spectroscopy (FTIR) The infrared spectra were scanned on a FTIR spectrophotometer (Shimadzo, Japan), at 4 cm-1 resolution in frequency range between 4000 and 400 cm-1 using KBr Pellet method with the sample:KBr ratio of 1:10.