Study On A Novel Nano-Delivery System Of Emodin Based On Intestinal Flora Target

At present,the incidence rate of diabetes worldwide is increasing year by year,and the proportion of type II diabetes has exceeded 90%.Type II diabetes has the characteristics of intestinal flora disorder,and intestinal flora is considered as a potential target for the treatment of type II diabetes.Emodin has the effect of treating type II diabetes and regulating intestinal flora,but there is no research on using emodin to treat type II diabetes with intestinal flora as the target,and the bioavailability of emodin in vivo is low.Therefore,in this study,emodin was prepared as a microbiota type oral colon targeted drug delivery system to improve drug bioavailability and target intestinal flora to treat type II diabetes.Objective:To prepare an emodin microbiota type oral colon targeting nanoparticles,in order to achieve the intestinal microbiota targeting in the colon,and provide a reference for the development of drug preparations targeting the intestinal microbiota for the treatment of type II diabetes.Methods:The polylactic acid-glycolic acid copolymer（PLGA）loaded emodin（EM）nanoparticles（EM-PLGA NPs）were prepared by emulsion solvent evaporation method,and the formulation was optimized by single factor examination and Box-Behnken effect surface method.Pectin and chitosan-coated PLGA nanoparticles（EM-PLGA-CS-PEC NPs）were prepared after chitosan（CS）and pectin（PEC）were successively coated on the surface of EM-PLGA NPs by layer-by-layer adsorption.EM-PLGA-CS-PEC NPs was evaluated for their appearance and dispersibility,morphology,encapsulation efficiency,drug loading,particle size and potential,Fourier transform infrared spectroscopy and in vitro release quality.In addition,the pharmacokinetic differences between EM-PLGA-CS-PEC NPs and EM APIs in vivo were compared by pharmacokinetic method;The distribution and residence time of EM-PLGA-CS-PEC NPs in vivo were investigated by using mouse in vivo imaging;The hypoglycemic effect of EM-PLGA-CS-PEC NPs was analyzed by measuring the blood glucose values at different time points after administration;Finally,the effect of EM-PLGA-CS-PEC NPs on intestinal flora of type II diabetes rats was analyzed by high-throughput sequencing.Results:（1）The best process of EM-PLGA-CS-PEC NPs:2.00 mg emodin and 20.28 mg PLGA,dissolved with 2 m L acetone,slowly injected into 1.11%polyvinyl alcohol solution after complete dissolution,stirred for 3.39 h at 400 r/min after ultrasonic emulsification for 10min,and finally obtained EM-PLGA NPs by ultrasonic probe at 550 W power for 10min;EM-PLGA NPs solution is dropped into an equal volume of 0.1 mg/m L chitosan solution,stirred at300 r/min for 1 h,and then stood for 0.5 h,and then dropped into an equal volume of 0.5 mg/m L pectin solution,stirred at 300 r/min for 1 h to obtain EM-PLGA-CS-PEC NPs;（2）Quality evaluation:The morphology of EM-PLGA-CS-PEC NPs is spherical or quasi-spherical;The encapsulation rate was（85.31±0.20）%,and the drug loading was（2.20±0.01）%;The particle size is（438.89±1.26）nm,with uniform distribution;The potential is（-20.73±0.82）m V;Fourier transform infrared spectroscopy showed that emodin was successfully encapsulated on the surface of nanoparticles,and chitosan and pectin were also adsorbed on the surface of nanoparticles in turn;The release of EM-PLGA-CS-PEC NPs in the medium of p H1.2 and p H 6.8 is low,and the release of EM-PLGA-CS-PEC NPs in the medium of p H 7.8 is high.The best fitting equation for the release results in vitro is Weibull equation;（3）Pharmacokinetic study:Based on the pharmacokinetic parameters of EM raw materials administered orally to rats,the relative bioavailability of EM-PLGA-CS-PEC NPs was205.68%;Compared with EM bulk drug,the AUC_（0-t）of EM-PLGA-CS-PEC NPs after intragastric administration was（19.97±5.17）mg/L*h,an increase of 105.66%,indicating that EM-PLGA-CS-PEC NPs can significantly increase the bioavailability of EM in vivo;MRT_（0-t）and t_1/2 were（34.26±2.99）h and（45.76±33.23）h,increased by 80.79%and 104.60%respectively,indicating that EM-PLGA-CS-PEC NPs had a slow release effect;In addition,the apparent distribution volume parameter V_z/F and clearance parameter CL_z/F of EM-PLGA-CS-PEC NPs group decreased by 49.06%and 63.27%,respectively,which to some extent showed that the distribution range of EM-PLGA-CS-PEC NPs in vivo is narrower than that of EM raw materials;（4）In vivo imaging:EM-PLGA-CS-PEC NPs can prolong the retention time of drugs in vivo,and the fluorescence signal is mainly concentrated in the colon;（5）Hypoglycemic effect:Compared with the crude drug of emodin by gavage,the hypoglycemic effect of EM-PLGA-CS-PEC NPs by gavage is more lasting and can be maintained for 24 hours;（6）Intestinal flora analysis:The intestinal flora of type II diabetes model rats appears disorder to a certain extent;At 24 hours after the treatment of EM-PLGA-CS-PEC NPs,the composition and abundance of intestinal microflora at the level of phylum,family and genus and the analysis index of intestinal microflora diversity were the closest to those of the normal group,and the number of shared OTUs with the normal group was the largest,and the number of shared OTUs with the T2D group was the smallest,indicating that EM-PLGA-CS-PEC NPs had the function of targeting and regulating intestinal microflora,and its effect was stronger than that of emodin crude drug.Conclusion:Emodin microbiota type oral colon targeted drug delivery nanoparticles were successfully prepared,which can reach the colon and target the intestinal flora,and treat type II diabetes by regulating the intestinal flora.