Development of a liposomal nanodelivery system for nevirapine

Abstract

The treatment of AIDS remains a serious challenge owing to high genetic variation of Human Immunodeficiency Virus type 1 (HIV-1). The use of different antiretroviral drugs (ARV) is significantly limited by severe side-effects that further compromise the quality of life of the AIDS patient. In the present study, we have evaluated a liposome system for the delivery of nevirapine, a hydrophobic non-nucleoside reverse transcriptase inhibitor. Liposomes were prepared from egg phospholipids using thin film hydration. The parameters of the process were optimized to obtain spherical liposomes below 200 nm with a narrow polydispersity. The encapsulation efficiency of the liposomes was optimized at different ratios of egg phospholipid to cholesterol as well as drug to total lipid. The data demonstrate that encapsulation efficiency of 78.14% and 76.25% were obtained at egg phospholipid to cholesterol ratio of 9:1 and drug to lipid ratio of 1:5, respectively. We further observed that the size of the liposomes and the encapsulation efficiency of the drug increased concomitantly with the increasing ratio of drug and lipid and that maximum stability was observed at the physiological pH. Thermal analysis of the drug encapsulated liposomes indicated the formation of a homogenous drug-lipid system. The magnitude of drug release from the liposomes was examined under different experimental conditions including in phosphate buffered saline (PBS), Dulbecco's Modified Eagle's Medium (DMEM) supplemented with 10% fetal bovine serum or in the presence of an external stimulus such as low frequency ultrasound. Within the first 20 minutes 40, 60 and 100% of the drug was released when placed in PBS, DMEM or when ultrasound was applied, respectively. We propose that nevirapine-loaded liposomal formulations reported here could improve targeted delivery of the anti-retroviral drugs to select compartments and cells and alleviate systemic toxic side effects as a consequence.

Figure 1

SEM and TEM micrographs of the lyophilized liposomes. (A) The sample was imaged s performed at a magnification of 43,000. Inset shows the magnified image of spherical liposome (B) TEM micrographs of liposome at a magnification of 60,000.

Figure 2

Effect of pH on colloidal stability of liposome.

Figure 3

Effect of Cholesterol on the encapsulation efficiency of nevirapine. Statistical data infers that each group is significantly different (p < 0.05).

Figure 4

Correlation between the mean particle size (white circle) and percent encapsulation efficiency (Black square) at various drug-lipid ratios.

Figure 5

SC thermograms of plain (filled line) and nevirapine-loaded (dashed line) liposomes.

Figure 6

Release profile of nevirapine from liposomes at various pH containing phospholipid to cholesterol in ratio 9:1 (* < 0.05).

Figure 7

In vitro release profile of nevirapine in PBS (pH 7.4) release medium at various conditions. (Black square) liposomes with phospholipid to cholesterol ratio 10:00 in PBS, (Circle with cross) Liposomes with phospholipid to cholesterol ratio 9:1 in PBS, (Black triangle) liposomes with Phospholipid to cholesterol ratio 10:00 in PBS subjected to low frequency Ultrasound and (Inverted triangle with partial shading) liposomes with phospholipid to cholesterol ratio 10:00 in DMEM cell culture medium. PBS represents the phosphate buffered saline; DMEM refers to Dulbecco's Modified Eagle's medium.

Figure 8

In vitro release profile of nevirapine in PBS (pH 7.4) subjected to ultrasound (US) under various conditions. (Grey square) liposomes with phospholipid to cholesterol ratio 9:1 in DMEM, (white square) liposomes with phospholipid to cholesterol ratio 10:0 in DMEM, (Black square) liposomes with phospholipid to cholesterol ratio 10:0 in PBS. PBS refers the phosphate buffered saline; DMEM refers to Dulbecco's Modified Eagle's medium. Statistical data shows that a < 0.05 vs 5 minutes, b < 0.05 vs 20 minutes, c < 0.05 vs 10 minutes and d < 0.05 vs 20 minutes.