Transcriptomic Analysis Of Anti-diarrhea Effect And Extraction Process Optimization Of Functional Component In Punica Granatum L.

Pericarpium Granati are dry peels of pomegranate in the Punicaceae family.It tastes gentle,sour,pucker and converges to stomach and large intestine channel.There are many benficial effect like astring intesetine to correct diarrhea,expell insect and stop the bleeding.It's usually used in diarrhea,dysentery,abdominal pain and hemostasis.It has been proved that Ethyl acetate fraction is active part of water extract of pomegranate peels and has been identified that punicalagin,punicalin and ellagic acid are effective components in herb by previous study,whereas its antidiarrhea mechanism is not clear.Study on the anti-diarrhea effect of pomegranate peel is helpful for its advantagement and development.Also,it's the basis of the optimization of extraction process.Therefore,firstly,talking about possible anti-diarrhea molecular mechanism using RNA-seq at the level of genes.Secondly,optimizing the extraction of punicalagin,punicalin and ellagic acid from pomegranate peels by Taguchi.Lastly,testing its anti-diarrhea effect optimized by Taguchi.BALB/c mice are randomly devided into balnk group,castor oil group,punicalagin & castoroil group,punicalin & castor-oil group and ellagic acid & castor-oil group.All mice are given intraperitoneal injection once a day for seven days then execute and collect jejunums.Analysis transcriptome data.Compare differentially expressed genes among groups and analysis GO enrichment and KEGG pathway.The results show that there are 711 differentially expressed genes between castor-oil group and blank group with 553 genes go up and 158 genes go down.There are 214 differentially expressed genes between punicalagin & castor-oil group and castor oil group with 104 genes go up and 110 genes go down.There are 259 differentially expressed genes between punicalin & castor-oil group and castor oil group with 130 genes go up and 129 genes go down.There are 273 differentially expressed genes between ellagic acid & castor-oil group and castor oil group with 160 genes go up and 113 genes go down.KEGG pathway analyzed with these differentially expressed genes are: PPAR signaling pathway,vitamin digestion and absorbtion,retinol metabolism and so on.Punicalagin,punicalin and ellagic acid can decrease the impact of castor oil on vitamin digestion and absorbtion and retinol metabolism by intervening genes referring to RNA-seq results and literature report.On the basis of single-factor experiment,optimizing extraction process of punicalagin,punicalin and ellagic acid from pomegranate peels by Taguchi.Optimized conditions are: ethanol concentration of 20%,extraction temperature of 70?,extraction time of 2 h,liquid-solid ratio of 50 m L/g,particle size of 0.125 mm and extraction twice.Extracting three times in the optimized conditions.The yield of punicalagin,punicalin and ellagic acid was 498.66 ± 2.13,17.89 ± 0.27,7.30 ± 0.05 mg/g,respectively,which is close to predicted value.Testing anti-diarrhea effect of pomegranate peel extract optimized in the former step in Mice diarrhea model induced by castor oil and compared with water ectract and ethyl acetate fraction of pomegranate peel.The results show that all three extracts in the dose of 200 mg/kg can prolong first time of diarrhea,reduce the number of loose stools,reduce propulsion rate of carbon powder,reduce fluid collection in the small intestine.Anti-diarrhea effect of pomegranate peel extract optimized by Taguchi is weaker than ethyl acetate fraction but stronger than water extract of pomegranate peel.Pomegranate peel extract optimized by Taguchi is more effective in anti-diarrhea and optimization of extraction technology is valuable in application.The researah study on anti-diarrhea mechanism of active components in pomegranate peel: punicalagin,punicalin and ellagic acid by transcriptome for the first time.Also,the study optimizes extraction technology of three active components in pomegranate peel by Taguchi and prove that optimized extract is better than bfore optimize in anti-diarrhea.The study provides a certain scientific evidence for exploitation and utilization of pomegranate peel resource in anti-diarrhea.Also,this is the basis on industrial product of pomegranate peel in traditional Chinese medicine extraction.