Study On The Preparation And Characterization Of Brucine Stealth Liposomes

Brucine, the major active alkaloid constituent extracted from traditional Chinese herbal medicine Nux vomica, had been found to possess remarkable antitumor, analgesic and anti-inflammatory activities. Unfortunately, the potential use of brucine was severely limited due to its violent toxicity. Stealth liposomes are novel drug delivery carriers. After encapsulated into stealth liposomes, the pharmacokinetic characteristics of the drug were significantly improved and the therapeutic index was also improved. In the present study, stealth liposomes containing brucine were prepared. Furthermore, the in vitro and in vivo properties of the stealth liposomes were also investigated.To control the quality of prepared brucine liposomes, the methods for the determination of brucine content in liposomes and the encapsulation efficiency (EE) of liposomes were established. Then brucine liposomes were prepared by ammonium sulfate transmembrane gradients and the influence of various factors on the entrapment efficiencies were investigated and optimized. The EE was enhanced with increased ammonium sulfate concentration, ethanol volume and phospholipid concentration. To obtain the brucine with the highest EE, soy phosphatidylcholine (SPC) and cholesterol at a weight ratio of6:1were dissolved in8mL ethanol and then injected into the solution of10mL0.2M ammonium sulfate. The concentration of the prepared liposomes was0.9mg/mL and the EE was80.48%.Brucine stealth liposomes and conventional liposomes were both prepared by ammonium sulfate transmembrane gradients. The encapsulation efficiency, particle size, in vitro release profiles and stability were compared. The encapsulation efficiency of brucine stealth liposomes and conventional liposomes were80.7±0.5%and80.5±0.3%, respectively. The mean paricle sizes were82.8nm and164.9nm, respectively. Whether rat plasma was added or not, the release rate and degree of brucine stealth liposomes were significantly lower than those of conventional liposomes. After centrifuge, brucine stealth liposomes and conventional liposomes were both stable. After stored at4℃for70days, the EE of the stealth liposomes and conventional liposomes was94.9and3.2%, respectively. As the antitumor durg delivery system, the in vitro characteristics of brucine stealth liposomes were more satisfactory than the corresponding conventional liposomes.KM mice received inoculation of murine hep2carcinoma cells (H22). The H22mice were randomly administered with brucine solution, brucine conventional liposomes or brucine stealth liposome (3.23mg/kg, ip) for8days. The effectiveness was assessed based on tumor inhibition rate. Meanwhile, the mice body weight and immune organ weight were also compared. The brucine solution, buricne conventional liposomes and brucine stealth liposomes in the same dose showed an inhibition rate of28.64%,57.96%and71.36%, respectively. Obvious immunosuppressive effect was not found in the brucine stealth liposome group. Compared with conventional liposomes, stealth liposomes, as the carrier of brucine, could improve the antitumor effect.The pharmacokinetic characteristics in rats were evaluated by comparison with brucine solution. Brucine plasma concentration was determined by HPLC. The3p97program was adopted to simulate the compartment model and calculate the pharmacokinetic parameters.The concentration-time profiles of brucine stealth liposome was in accordance with three-compartment model, while brucine solution with two-compartment model. Compared with brucine solution, pharmacokinetic parameters such as AUC and CL of brucine stealth liposome were increased significantly. For example, MRT of brucine in the stealth liposome was2.87-fold of that of solution. Therefor, brucine stealth liposome had more satisfactory pharmacokinetic properties than solution, which could account for its improved antitumor effect.HPLC method was developed to measure the brucine concentration in plasma, liver and brain of mice. After intravenous injection of brucine solution and brucine stealth liposomes, the tissue distribution of brucine was compared. The results revealed that brucine concentrations in plasma after administration of brucine stealth liposome were significantly increased while the concentrations in liver and brain were not increased compared with brucine solution.