Studies On The Angong Niuhuang Nasal Drug Delivery System For Brain Targeting

The prescription of Angong Niuhuang(AN) was a classic formulation in Traditional Chinese Medicine(TCM) for emergency medical treatment,such as high fever,coma,stroke, etc.The common used dosage forms are pills and powder of AN for oral administration,and its simplified prescription of Qingkailing for intravenous(i.v.) administration.However,all these preparations have some limitations in clinical use.Intranasal(i.n.) administration is one of the drug delivery system being given more research in recent years.It has the advantage of rapid absorption,administration convenience,and circumvention of blood brain barrier(BBB) for brain-targeting.In order to fully exert the therapeutic effects of AN for the treatment of central nervous system(CNS) disease,and improve the compliance of the patients,a new nasal formulation of AN was established,on the basis of pre-process of every raw material in the traditional AN prescription.Then,the exist of nose-brain route,formulation factors and dosage form factors affecting nasal absorprion,and the absorption mechanism was studied detailedly after i.n.administration of baicalin,which was the most important ingredient in AN on relieving high fever with definite antipyretic mechanism in CNS.And finally,based on the preparation of baicalin lipid microspheres(BLM),a kind of brain-targeted two-phase drug loading lipid microsphere of AN for nasal use was prepared.At first,eight analytical method for the quality control of baicalin,jasminoidin, curcumine,berberine,cholalic acid,bilirubin and borneol was established.Then,different kinds of pre-process methods were performed with every raw material:jasminoidin and cholalic acid were extracted from gardenia and artificial calculus bovis,respectively;borneol and muscone was solubilized by randomly methylated -/?-cyclodextrin(RAMEB);the inclusion complex of curcumine with RAMEB was prepared and verified by DSC;powder of pearl and buffalo horn was hydrolyzed together;and the solubility of baicalin and berberine in aqueous solution was determinied.According to the content of index ingrediant in pre-processed product and CHP 2005,the formulation and preparation of nasal AN solution (AN-S) were established. The in situ rat nasal perfusion technique was used in the investigation of nasal absorption of index ingredients in the AN formution.The respective absorption study of the index ingredients showed that baiclain,berberine and curcumine had obvious absorption from the nose of rats,except for jasminoidin.The detailed absorption study of baicalin indicated that the absorption of baicalin followed first-order absorption kinetics,and the order of increasing absorption of baicalin caused by the enhancers was 1%sodium deoxycholate ? 0.5% chitosan≈5%borneol≈5%RAMEB >5%HP-β-CD > 5%β-CD>l%Tw-een-80≈0.1% EDTA-Na2≈1%lecithin.A simultaneous blood and brain microdialysis coupled with UPLC-MS/MS method was established for in vivo analysis of baicalin.In vitro recoveries of the probes determined by no net flux method were 19.26%and 18.38%,while in vivo recoveries of the probes determined by retrodialysis were 15.0%and 17.5%for blood and brain,respectively.The pharmacokinetics and brain-targeting study were investigated after i.n.administration (12mg.kg~-1) baicalin solution(BS) of pH5.4(INI),BS of pH 4.8(IN2),BS with 5%RAMEB added(IN3) and BS with 1%borneol added(IN4),and i.v.administration(12mg.kg-~1) of BS (IV1),respectively.Pharmacokinetic calculations were performed on each individual set of animal data using the pharmacokinetic calculation software DAS(drug and statistics) version 2.0 by the statistical moment method.The result showed that baicalin has a slight BBB permeability after i.v administration.The much lower concentration level of baicalin in plasma and delayed Tmax after i.n.administration mean a poor absorption from nasal cavity. The bioavailability of baicalin was 11.7%,12.5%,21.0%and 11.0%in plasma,69.3%,58.2%, 89.0%and 72.5%in CSF,for INI,IN2,IN3 and IN4,repectively.Probably due to the absorption enhancing effect of RAMEB,both bioavailability of IN3 in plasma and CSF were significantly higher than the other three i.n.formulations.Although borneol also has enhancing effect in CSF transport,the effect was not significant.In addition,Drug targeting index(DTI) was 5.86,4.64,4.23,6.55,and direct transport percentage(DTP) to brain was 83.0%,78.6%,76.5%,84.8%,for the four formulations,respectively.And this result supported strongly that there was direct access from nose to brain for baicalin getting into brain through olfactory pathway.The reaction solvents,the ratios of phospholipids were investigated,and the baicalin-phospholipid complex(BPC) was finally prepared with baicalin and soybean lecithin at a ratio of 1:5 in tetrahydrofuran at room for 3h,and then the reaction solvent was evaporated to obtain dry residue.DSC and and X-ray diffraction were also employed to identify the formation of the complex and analyze the possible interaction between clarithormyin and phospholipids.High pressure homogenization was used to prepare baicalin lipid microspheres(BLM). HPLC,dynamic light scattering,electrophoretic light scattering technology,and ultrafiltration were also employed.Taking physical appearance,pH,particle size distribution(PSD), ^-potential,content,entrapment efficiency as index,the final formulation and preparation process for BLM were as follows:as quality percentage,oil phase was composed of 1% baicalin in the form of the complex,20%MCT,1%soybean lecithin;the water phase was composed of 2.5%glycerol,1%F-68,0.06%oleic acid and 0.006%EDTA-Na2;and the oil phase and water phase were both heated to 70°C;the homogenization pressure and cycles were 700bar and 7 times;the pH was adjusted to 5.0 before homogenization.The pharmacokinetic study of BLM(2.4mg.kg(~-1)) after i.n.administration(BLM-IN) to rats showed that the absolute bioavailabilities in plasma and CSF were 55.2 and 227%,which were 5 and 3 times of INI,respectively.By comparing the ratio of AUCcsf/ AUCpiasma between BLM-IN and INI(0.302 vs 0.434),it can be seen that although the absorption of baicalin was greatly improved after i.n.administration of BLM,its brain targeting efficiency became lower.According to the formulation and preparation of BLM,and the AN formulation eatablished in Chapter 1,a kind of AN lipid microsphere(AN-LM) was prepared.The drug loading and entrapment efficiency(EE) were determined based on the content of baicalin.To assess the ciliary toxicity of nasal preparations of AN-S and AN-LM,the hoptoad in situ maxilla mucosa method was used by determining the ciliary movement time.In addition,the method of biochemical index was emploied to evaluate the nasal mucosa toxicity of RAMEB. The result showed that the two dosage forms of AN has obvious ciliary toxicity,and 10%and 20%(w/v) RAMEB has some impact on the physiological change of nasal mucosa of rats.