Preparation Of Icaritin-loaded Micelles And Evaluation Of The Protective Effects On Cerebral Ischemia-reperfusion Injury

Icaritin is an isoprene flavonoid extracted from traditional Chinese medicineEpimedium.It has a variety of pharmacological and biological activities,including promoting cardiomyocyte differentiation,inhibiting tumor cell growth,and promoting fracture healing.Recent studies have found that icaritin has neuroprotective effects on cerebral ischemia-reperfusion injury.However,icaritin is extremely difficult to dissolve in water,its solubility is less than 1.0μg/mL,and its low bioavailability limits its clinical application.Polymeric micelles have multiple advantages as a drug delivery system,including increasing solubility of poorly soluble drugs,improved drug stability,delayed release,controlled release,improved drug efficacy,reduced toxicity and targeting.We synthesized amphiphilic polyethylene glycol monomethyl ether-polylactic acid（mPEG-PLA）copolymer by ring-opening polymerization,and prepared injectable icaritin drug-loaded micelles by membrane hydration method.In order to improve the drug-loading effect of icaritin-loaded micelles,we optimized the preparation process of polymer materials and icaritin-loaded micelles.The best prescription process is:weighing icaritin 10 mg,mPEG-PLA（5:6）80 mg,adding 60 mL of methanol to ultrasonically disperse evenly,and removing the methanol by heating in a 55°C water bath.Add 70 mL of ultra-pure water of the same temperature.The particle size,potential,encapsulation efficiency,drug loading,critical micelle concentration and solubility of icaritin-loaded micelles were（64.25±0.21）nm,（-1.37±0.31）mV,83.96%,9.33%,2.24μg/mL and 2.0mg/mL,respectively,shows a better encapsulation effect on icaritin,greatly improving the solubility of the drug.In vivo plasma kinetics results showed that icaritin-loaded micelles significantlyslowed the metabolism of icaritin in vivo,and the AUC and t_1/2z values were 4.3-fold and1.1-fold higher than those of the icaritin.Apparent volume of distribution Vz and the clearance rate CLz are reduced,increasing the plasma concentration of the drug.The results of tissue distribution showed that the distribution of icaritin-loaded micelles in the brain was 8-10 times that of icaritin,showing a better tissue distribution effect.Pharmacodynamic experiments showed that icaritin-loaded micelles have a good neuroprotective effect on mice with cerebral ischemia-reperfusion injury.The neurological scores of the icaritin group and the mPEG-PLA/icaritin micelles group were1.9±0.88 and 1.2±0.79,cerebral edema were 79.82±1.04%and 78.88±0.77%,respectively.Compared with the icaritin group,the cerebral infarction volume of mPEG-PLA/icaritin micelles reduced significantly,which were 22.49±3.71%and 15.8±1.98%,respectively,p<0.01.Icaritin-loaded micelles not only significantly improve the solubility,plasmaconcentration and the tissue distribution of icaritin,but also have a good neuroprotective effect on mice with cerebral ischemia-reperfusion injury,which have broad prospects for the treatment of ischemic stroke.