| Coixol is a colorless needle-like crystal extracted from plants such as coix,imperata,and scoparia dulcis,and it is also one of the main active ingredients in the 'Kanglaite' injection.At present,there are many clinical reports about the anti-cancer effect and the reduction of cancer pain of 'Kanglaite' injection.It also reports that coixol has analgesic,anti-inflammatory,antibacterial,and hypoglycemic effects.The research and development of coixol formulations can meet the needs of clinical drugs,which has the value of practical applications.Due to the poor water-solubility of coixol,hydroxypropyl-?-cyclodextrin(HP-?-CD)was used for inclusion of coixol in this experiment.Based on the successful preparation of its clathrates,the release situation in vitro and stability were investigated.Additionally,the method for the determination of coixol in biological samples of HPLC was established.After intragastric administration of coixol and its clathrates,the drug concentrations in rat plasma and tissues were examined to study its pharmacokinetic characteristics and tissue distribution in vivo,providing an experimental basis for the research and development of new oral formulations of coixol.The main research contents and results are as follows:1 Preparation and characterization of coixol-HP-?-CD clathratesThe clathrates of coixol were prepared by the solvent-stirring and freeze-drying method.The clathrates were characterized by SEM,FT-IR,DSC and XRD.A HPLC method was used to determine the content of coixol in its clathrates,and methodological investigations were carried out to evaluate the method.The results showed that the peak area of coixol had a good linear relationship.The regression equation of coixol is:y=108096x-48927(R2=0.9995).The specificity,precision,stability and recovery rate of coixol all meet the requirement which showed this method can be applicable to the determination of coixol in its clathrates.2 Optimization of preparation process of coixol-HP-?-CD clathratesWith inclusion rate and the inclusion complex yield rate as indexes,star points were selected for three factors:HP-?-CD and coixol feed ratio,inclusion time,and inclusion temperature by the single-factor investigation.The design-response surface experiment was conducted to determine the optimum process for preparation of clathrates:the feed ratio of HP-?-CD:coixol was 3:1,the inclusion temperature was 60 ?,and the inclusion time was 5 h.3 Release test in vitro and stability testWe used dialysis bags(molecular weight cutoff of 8000?14000)for the release experiments in vitro.The results showed that the cumulative release rate of coixol-HP-?-CD clathrates was 2.55 times higher than that of coixol,which shows that the solubility of coixol-HP-?-CD clathrates is far greater than that of coixol.The results of stability test showed that the coixol-HP-?-CD clathrates have good stability under high temperature and light conditions,but it is easily hygroscopic and deliquescent in a high-humidity environment.Therefore,it is as possible as to keep it sealed and dry during the production,storage and transportation process.4 Study on pharmacokinetics and tissue distributionAHPLC method for the determination of coixol in biological samples was established and the methodology was investigated In order to more effectively avoid the interference of complex components in biological tissues,an internal standard method was used for quantitative analysis.It was found that coixol peaks and internal standard peaks were well-shaped and completely separated.In the plasma samples,the coixol had good linearity in the concentration range of 0.16?25.00 ?g/mL.In each tissue sample,the coixol had good linearity in the concentration range of 0.03?3.13 ?g/mL.The precision,the stability and the recovery rate are all good,which is in accordance with the methodological requirements of detection of biological samples in vivo.The pharmacokinetics of coixol and its clathrates in rats were studied using plasma concentration assay.After intragastric administration to rats at a dose of 100 mg/kg,data were processed using a two-compartment model to determine pharmacokinetic parameters.The results showed that the Cmax value of coixol(14.9±2.82 ?g/mL)was lower than that of its clathrates(17.48 ±2.33 ?g/mL),and the t1/2Z of coixol and its clathrates were(0.83±0.44)h?(1.97±0.63)h,respectively.The mean retention time of coixol and its clathrates were(1.00±0.13)h?(2.37±0.23)h,respectively.Compared with coixol,the elimination half-life and the mean retention time of coixol-HP-?-CD clathrates was significantly prolonged(p<0.01).The AUC(0?t0)of coixol-HP-?-CD clathrates in rats was significantly increased(p<0.01),which was 232.83%of coixol.This shows that clathrates can increase the absorption capacity of drugs,and can delay the elimination of coixol in the body to achieve long-lasting effect and increase its oral bioavailability.Rats were administered coixol and its clathrates by 100 mg/kg,HPLC determination of coixol concentration in rats(heart,liver,spleen,lung,kidney and brain)at 0.25?0.5?1?3?6?10 h.The results showed that the concentration of the clathrates group was lower than that of the coixol group at 0.25 hours.The concentrations of the clathrates group in the tissues began to be higher than those of the coixol group at 0.5 hours.The concentrations of the clathrates group in each tissue was higher than those in the coixol group at 1 hours.At 6 hours,the concentration of drugs in the brain,heart,spleen,lung were lower than the detection limit in the coixol group,while the concentration of drugs in the tissues was detected in the clathrates group.In addition,the concentration of the drug in the coixol group was the highest at 0.25 hours,while the concentration of the coixol group in the tissues(except kidney)was the highest at the 0.5 hours.The results can be indicated that HP-?-CD can increase the concentration of coixol in tissues and release the coixol slowly to have a long-lasting effect at the same time.The content of the clathrates group in the tissues decreased at a slower rate than that of the coixol group,which indicated that the removal of coixol in vivo could be delayed after the clathrates were made.Generally,the content of coixol in liver and kidney was significantly larger than that in other tissues,and the concentration of coixol in the organs was as follows:kidney>liver>spleen>heart>lung>brain.Compared with the coixol group,the content of coixol in the clathrates group in the kidney and liver decreased slowly,indicating that the clathrates can delay the elimination of drugs in vivo.It is noteworthy that the brain can also detect the content of coixol,which suggested that coixol can use the blood-brain barrier and coixol may play an analgesic role in the brain's central nervous center.The content of coixol-HP-?-CD clathrates decreased slowly in the brain with prolonging of time,indicating that the clathrates could slow down the drug's elimination rate in the brain,and increase the retention time in the brain.It is suggested that the analgesic effect of 'Kanglaite' injection on cancer patients is likely to be caused by coixol. |
PDF Full Text Request