Study On The Intervention Of Volatile Oil Of Atractylodes Chinensis On Diabetic Gastroparesis In Rats And Preparation Of Microcapsules

Objective: Atractylodes chinensis(DC.Koidz)is compositae class annual herb,and its main component has many pharmacological effects However,there are relatively few research on extraction process of Atractylodes chinensis volatile oil(ACVO)and its pharmacological effects and the preparation of dosage forms.In the present study,the extraction technology of volatile oil of rhizoma atractylodis was optimized.The prevention of ACVO on diabetes gastroparesis(DGP)in rats and its mechanism were explored.The preparation of AVCO microcapsule was investigated.Enzymolysis–microwave assisted hydrodistillation(EMAHD)was adopted to extract the volatile oil,and the technology was optimized using a single-factor experiment that incorporated response surface methodology(RSM).The extraction rate of volatile oil by EMAHD,microwave assisted hydrodistillation(MAHD),and hydrodistillation(HD)methods were compared at different times.The ingredients of Atractylodes chinensis volatile oil were analyzed by gas chromatography–mass spectrometry(GC-MS).Scanning electron microscopy(SEM)was used to analyze the microstructural changes in Atractylodes chinensis residue before and after extraction.The inhibition of α-amylase activity was determined.The animals were randomly divided into control group(NCG),diabetic gastroparesis(DGP),ACVO low dose group(ACVO.L),ACVO medium dose group(ACVO.M),ACVO high dose group(ACVO.H),cisapride and metformin hydrochloride mixed administration group(CMG).Rats in all groups except NCG group were injected STZ combined with high sugar high lipid diet to establish DGP model And then,the rats in the ACVO.L,ACVO.M,ACVO.H were given ACVO daily(9.11mg/kg,18.23mg/kg,36.45mg/kg,respectively)by intragastric administration,and those in the CMG were given cisapride(3.5mg/kg)and Metformin Hydrochloride(175mg/kg)once a day for 4weeks.After the experiment,tissue samples were taken from each group.Blood glucose and serum Insulin like growth factor-1(IGF-1)content of rats in each group were determined.Gastric emptying and intestinal propulsion were determined by phenol red assay.Gastric fluid and gastric acid content were determined The protein expression levels of insulin-like growth factor-1(IGF-1R),acetylcholine transferase(Ch AT)and stem cell factor(SCF)in the stomach were determined by Western Blot.The changes of IGF-1R,CHAT and SCF in gastric tissues were detected by immunofluorescence chemical staining.Morphological changes of gastric tissues were detected by hematoxylin eosin staining.The changes of Cajal stromal cells in gastric tissues were detected by immunohistochemistry.The capsular preparation conditions were investigated by Response surface method(RSM).SEM,particle size analyzer and FTIR were used to characterize ACVO microcapsules.Release of ACVO at p H 1.5 and7.4 was measured.The effect of microcapsules on cell viability and the length of rat gastric smooth muscle cells(GSMC),m RNA and protein expression of calmodulin(Ca M)and myosin light chain kinase(MLCK)were investigated.Results: The obtained optimal extraction conditions for EMAHD were as follows:enzyme concentration 1.6%,p H7,enzymolysis time 20 min,enzymolysis temperature50℃,liquid-solid ratio 30:1,microwave power 455 W and microwave time 40 min.The diversity of the main ingredients and the in vitro inhibition of α-amylase activities were higher for ACVO extracted by EMAHD than by HD or MAHD,respcetively.The powder residue of Atractylodes chinensis remaining after EMAHD showed a ruptured and collapsed cell structure,indicating accelerated dissolution of the volatile oil.After treatment with ACVO,body weight increased and blood glucose decreased when compared with rats in DGP group.The gastric emptying and intestinal propulsion accelerated,and gastric acid secretion increased.The serum Insulin-like growth factor-1(IGF-1)level was increased.The protein expressions and positive cells of IGF-1,CHAT and SCF were significantly increased in gastric tissue.Furthermore,the morphology and arrangement of the cells and the number of ICCs in the stomach were restored.When the concentration of chitosan was 1wt%,the mass ratio of chitosan to ACVO was 1,and the curing agent concentration was 1.8wt%,the encapsulation rate encapsuled by chitosan reached 82.19%.The prepared microcapsules had a continuous and spherical surface,which was more in line with the preparation requirements of microcapsules.The stability of ACVO coated with chitosan was significantly improved,and the cumulative release reached 85.32% at p H 7.4,making it more suitable for drug administration in vivo.In addition,ACVO microcapsules showed no cytotoxic at a concentration of 0.02 g/m L and 0.04 g/m L which induced GSMC to contract and improved the m RNA and protein expressions of Ca M and MLCK in GSMC.Conclusion: EMAHD is deemed a method with many advantages for extraction of volatile oil from Atractylodes chinensis.The volatile oil of Atractylodes chinensis is a promising component for treating DGP.The underlying mechanism may be to related to restore the expression of IGF-1 in the stomach,thereby up-regulate the expression of SCF,further restore the number of ICCs,so as to promote gastric movement.Meanwhile,the expression of CHAT in the stomach was restored and gastric movement was increased.The preparation method of volatile oil microcapsule of ACVO using chitosan as capsule material is feasible,simple and harmless,which has a good application prospect.