Study On Piperlongumine Albumin Nanoparticles

Piperlongumine（PL）is a natural alkaloid compound extracted from pepper plant,it has significant cytotoxic effects on colon cancer,breast cancer,liver cancer,ovarian cancer and other malignant tumors,but it has almost no significant effect on normal cells,is a potential antitumor drug.This project uses Nab^TM technology to prepare piperlongumine albumin nanoparticles（PL-BSA-NP）,using albumin as drug carrier to improve the water solubility,and improve bioavailability of PL.Using the single-factor investigation and star-point design experiments to optimize the formulation and preparation process,and filter out the best preparation conditions.And conduct pharmacological evaluation,pharmacokinetic research and in vitro antitumor effect research.Establishing the in-vitro analysis method of PL,taking the encapsulation rate as an index,investigating the drug mass ratio,the type of organic phase,the volume of organic phase and water phase,albumin concentration,p H,homogenization pressure,number of homogenization times,and types of lyophilized protective agents.Finally determined the best preparation conditions:the molar ratio of PL to albumin was 6:1,the ratio of absolute ethanol and methylene chloride was 3.5:1,the volume ratio of organic phase to water phase was 1:8,the albumin concentration was 1.25%,the p H was 8,the homogenization pressure was 80 psi,the number of homogenization times was 9 times,and the sucrose concentration was 2%.The pharmacological evaluation of PL-BSA-NP was carried out,including particle size,drug loading,encapsulation efficiency,shape,physical state,and stability.The morphology of PL-BSA-NP was relatively uniform in size,with an average particle size of about 210 nm,the drug load was 2.1%and the encapsulation rate was 87.6%.In PL-BSA-NP,PL was encapsulated in albumin in an amorphous state.The stability test results show that the PL-BSA-NP can be stable for 4 weeks when stored at 4°C.The release behavior of PL-BSA-NP in vitro has a sustained release effect,the cumulative release amount was 67.24%in about 24 hours.On this basis,we evaluated the pharmacokinetic properties of PL and PL-BSA-NP.The average plasma concentration at each time point of the PL group was lower than the PL-BSA-NP group.The half-life(T_1/2)of the PL-BSA-NP was increased 2.04 times,and the average residence time(MRT_0-∞)was increased 1.61 times,the drug retention time in the body was prolonged.The area under the time curve(AUC_0-∞)of the PL-BSA-NP was 2.20 times of the PL.This shows that PL-BSA-NP can maintain a certain blood drug concentration for a long time,show a certain sustained release,and increased the bioavailability of PL.Finally,we investigated the antitumor activity of PL-BSA-NP in vitro.The results show that PL can significantly inhibit Hep G2 cell proliferation,and its effect is dose-dependent,as the concentration of PL increases,the inhibitory effect gradually increases.At the same time,the inhibitory effect of PL-BSA-NP on Hep G2 cells at the same concentration was significantly stronger than that of PL,indicating that PL-BSA-NP can enhance the inhibitory effect of PL on tumor cell proliferation.In order to investigate the mechanism of killing Hep G2 cells by PL,we measured the changes of reactive oxygen species（ROS）in cells.The results showed that with the increase of the PL concentration,the ROS level gradually increased.At the same time,PL-BSA-NP have higher ROS levels than PL due to their enhanced tumor uptake capacity.In summary,we preliminarily concluded that PL can kill liver cancer cells by increase ROS level.