对氧磷酶脂质体的制备及其在大鼠体内的药代动力学

Preparation of paraoxonase liposomes and their pharmacokinetics in rats

目的 制备对氧磷酶1（PON1）脂质体,观察普通PON1脂质体（L-PON1）与经聚乙二醇修饰的PON1长循环脂质体（PEG-PON1-LCL）在大鼠体内的药代动力学.方法 采用薄膜分散法制备兔L-PON1和PEG-PON1-LCL,测定其包封率、粒径和表面电位,并评价其稳定性.将36只Wistar大鼠按随机数字表法分为3组,每组12只,分别静脉注射PON1、L-PON1和PEG-PON1-LCL 700 U/kg.采用乙酸苯酯法测定大鼠血清PON1活性,以注射后不同时间点PON1活性与注射前PON1活性差值作为外源性PON1活性值,并绘制酶活性-时间曲线;用DAS 2.0药代动力学软件及SPSS 17.0统计软件处理和分析数据,计算药代动力学参数.结果 L-PON1和PEG-PON1-LCL两种脂质体的包封率均〉85%,平均粒径为126 nm左右,Zeta电位为-14.35 mV;避光保存2周后上述指标无明显变化,说明脂质体稳定性较好,制剂学性质基本稳定.与单纯注射PON1比较,注射L-PON1或PEG-PON1-LCL两种脂质体后,大鼠体内酶活性明显提高,半衰期明显延长〔分布半衰期（T1/2α,h）：0.142±0.018、0.147±0.021比0.126±0.022,清除半衰期（T1/2β,h）：3.877±1.010、4.520±1.117比1.226±0.422〕,酶活性-时间曲线下面积（AUC）明显增加〔0-24 h的AUC（AUC0-24,U·h^-1·L^-1）：499.305±64.710、563.576±70.450比18.053±2.190,注射即刻至酶活性消失的AUC（AUC0 -∞,U·h^-1·L^-1）：516.256±60.940、587.801±76.210比21.044±3.250〕,表观分布容积（Vd）和清除率（CL）明显降低〔Vd（L）：0.140±0.065、0.144±0.064比0.493±0.032,CL（L/h）： 0.039±0.008、0.034±0.006比0.952±0.082,均P〈0.05〕;但两种脂质体间药代动力学参数差异无统计学意义.结论 采用薄膜分散法制备的PON1脂质体包封率较高,粒径较小,且稳定性好;两种PON1脂质体均提高了PON1的体内活性,同时改变了PON1的药代动力学,从而延长了其在血液循环中的时间,弥补了PON1在体内半衰期短的缺点.

Objective To prepare paraoxonase 1 （PON1） liposomes, and investigate pharmacokinetics of common PON1 liposomes （L-PON1） and polyethylene glycol-modified PON1 long circulating liposomes （PEG-PON1-LCL） in rats after intravenous administration. Methods L-PON1 and PEG-PON1-LCL were prepared by film dispersion method. The entrapment efficiency, mean diameter and Zeta potential of the liposomes were measured, and the stability was evaluated. Thirty-six Wistar rats were divided into three groups according to random number table, with 12 rats in each group. The rats were intravenously administrated with PON1, L-PON1 or PEG-PON1-LCL 700 U/kg, respectively. The activity of PON1 in serum was determined by phenyl acetate method, the activity of PON1 at different time points after drug administration was compared with that before drug administration, and the difference value was considered as the activity of exogenous PON1, and PON1 activity-time curve was plotted. The pharmacokinetic parameters were calculated and analyzed by DAS 2.0 pharmacokinetic program and SPSS 17.0. Results The entrapment efficiencies of L-PON1 and PEG-PON1-LCL were above 85%, the mean diameter was about 126 nm, and Zeta potential was -14.35 mV. After 2 weeks of preservation, the above parameters showed no obvious change, indicating that liposomes had good stability and the properties of preparations were basically stable. Compared with purified PON1 administration, after L-PON1 and PEG-PON1-LCL administration, the activity of PON1 was increased, the half-life of PON1 activity in rats was significantly prolonged [the half-life of distribution （T1/2α, hours）： 0.142±0.018, 0.147±0.021 vs. 0.126±0.022; the half-life of clearance （T1/2β, hours）： 3.877±1.010, 4.520±1.117 vs. 1.226±0.422], the area under PON1 activity-time curve （AUC） was significantly increased [AUC from 0 hour to 24 hours （AUC0-24, U·h^-1·L^-1）： 499.305±64.710, 563.576±70.450 vs. 18.053±2.190; AUC from the immediate injection to the disappearance of PON1 activity （AUC0-∞, U·h^-1·L^-1）： 516.256±60.940, 587.801±76.210 vs. 21.044±3.250], the apparent volume of distribution （Vd） and clearance （CL） were significantly decreased [Vd （L）： 0.140±0.065, 0.144±0.064 vs. 0.493±0.032, CL （L/h）：0.039±0.008, 0.034±0.006 vs. 0.952±0.082, all P 〈 0.05]. There was no significant difference in pharmacokinetics between L-PON1 and PEG-PON1-LCL. Conclusions The film dispersion method prepared PON1 liposomes have high entrapment efficiency and small particle size with a good stability. Both liposomes can raise PON1 activity in vivo, change the pharmacokinetics of PON1 in vivo, prolong the resident time of PON1 in the blood circulating system, and compensate for the short half-life of PON1 in vivo.