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To evaluate the therapeutic effect of Potentilla discolor on 2,4,6-trinitrobenzensulfonic acid(TNBS)-induced experimental ulcerative colitis(UC) in rats and to determine its therapeutic mechanism through mitochondrial autophagy, immune cells, and cytokines. A rat model of UC was established by TNBS-ethanol enema. Rats were divided into six groups: control, UC model, sulfasalazine(positive drug), and high-dose, moderate-dose, and low-dose ethanol extract groups. After 14-day continuous administration of the corresponding drugs, the disease activity index(DAI) and hematoxylin and eosin(HE) were evaluated. The morphological structure of mitochondria was observed by using transmission electron microscope(TEM), mitophagy-related mRNA expression was detected by using Real-time quantitative polymerase chain reaction(qRT-PCR), immune cell differentiation in rat serum was detected by using flow cytometry(FCM), and cytokine expression in colon tissues of rats was detected by protein microarray. The results showed that compared with the model group, each dose group of P. discolor could significantly reduce the DAI of UC model rats, and decrease the degree of inflammatory cells infiltration in the colon tissue of UC model rats. Meanwhile the expressions of T cells and Th cells in the serum increased significantly, the expression of Tc cells in the serum decreased significantly. Transmission electron microscope found that there was fusion of mitochondria and lysosomes in the colon tissue of the administration group. The expressions of mitochondrial autophagy related genes NF-κB, p62 and parkin were significantly increased in colon tissues. The results of protein chip showed that compared with the model group, the high dose group of P. discolor could significantly regulate the expression of cytokines. In conclusion, these results suggested that P. discolor improved TNBS-induced acute ulcerative colitis in rats by regulating the mitochondrial autophagy and the inflammatory factor expression.
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Animales , Ratas , Autofagia , Colitis Ulcerosa/genética , Colon , Mitocondrias , Potentilla/genéticaRESUMEN
OBJECTIVE:To establish HPLC fingerprints of Potentilla discolor,and to conduct authenticity identification.METHODS:HPLC method was adopted.The determination was performed on InertSustain C18 column with mobile phase consisted of 0.1％ formic acid solution-acetonitrile (gradient elution) at the flow rate of 1.0 mL/min,detection wavelength of 360 nm,colunn temperature of 30 ℃,sample size of 10 μL.Using rutin as reference,HPLC chromatograms of 19 batches of P.discolor and 2 batches of P.chinesis were determined.TCM Fingerprint Similarity Evaluation System (2004) was used for similarity evaluation of 21 batches of samples,and common peak identification of 19 batches of P discolor SPSS 21.0 statisticl software was used for main component analysis and cluster analysis.RESULTS:There were 18 common peaks in HPLC fingerprints of 19 batches of P.discolor,the similarity was higher than 0.9.-HPLC chromatogram was in good agreement with control fingerprint.The similarity of 2 batches of P chinesis was lower than 0.7.The 21 batches of medicinal materials could be grouped into 2 categories,2 batches of P chinesis could be grouped into a category,19 batches of P.discolor could be grouped into a category.P discolor could be grouped into 4 categories.Rutin and quercitrin were main ingredients in 19 batches of P discolor.CONCLUSIONS:Established fingerprint can provide reference for authenticity identification and quality evaluation of P.discolor.
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AIM To prepare the microemulsions loaded with total flavonoids from Potentilla discolor Bunge and to evaluate the therapeutic effect on diabetic mice.METHODS Total flavonoids,the extract from P.discolor by 70％ ethanol,were made into microemulsions after purification with.macroporous resin.With types and amount of oil phase,emulsifier and co-emulsifier,and addition amount of total flavonoids as influencing factors,together with drug loading,encapsulation efficiency,particle size and Zeta potential as evaluation indices,the formulation was optimized by drawing pseudo-ternary phase diagram.By establishing diabetic mouse models induced by streptozotocin (STZ) and high fat diet,the obtained microemulsions' effects on fasting blood glucose,oral glucose tolerance (OGT),insulin,glycosylated serum protein (GSP),triglyceride (TG),total cholesterol (TC),low density lipoprotein (LDL),high density lipoprotein (HDL),superoxide dismutase (SOD) and malondialdehyde (MDA) in mice were investigated.RESULTS The optimal formulation was determined to be 400 mg for isopropyl myristate (oil phase) amount,400 mg for Cremophor RH 40 (emulsifier) amount,200 mg for polyethylene glycol 400 (coemulsifier) amount,and 80 mg for addition amount of total flavonoids.The spherical-like and uniformly-sized microemulsions shared average drug loading of (7.28 ± 0.12)％,encapsulation efficiency of (91.79 ± 1.02)％,particle size of (36.79 ±0.17) nm and Zeta potential of (-15.77 ±2.89) mV.They could significantly reduce fasting blood glucose level,promote insulin secretion,regulate blood lipid,increase SOD activity,and decrease MDA level in diabetic mice.CONCLUSION Once prepared into microemulsions,the total flavonoids from P.discolor demonstrate an obviously enhanced activity in reducing blood glucose.
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OBJECTIVE:To optimize the extraction technology and forming technology of Compound Potentilla discolor gran-ules. METHODS:The extraction technology was optimized by single factor and orthogonal test with the amount of added water, soaking time,decocting time and decocting times as factors using the comprehensive score of the content of kaempferol,quercetin, and total flavonoids as indexes. Validation test was conducted. The type and ratio of excipients in forming technology were opti-mized by single factor test using comprehensive score of molding rate,soluble rate,moisture absorption rate and angle of repose as indexes. RESULTS:The optimal extraction process was 14-fold water,soaking for 0.5 h,decocting for 3 times,1.0 h each time. The RSDs of kaempferol,quercetin and total flavonoids content in validation test were 1.77%,1.76% and 4.62%(n=5). The ex-cipients of forming technology was the mixture of dextrin and soluble starch (2:1);the molding rate,soluble rate,moisture ab-sorption rate and angle of repose were 94.02%,76%,26% and 25.02 °,respectively. CONCLUSIONS:The optimized extraction technology of Compound P. discolor granules is stable and feasible,and prepared granules have good formability and moisture resis-tance.
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Objective To study the genetic diversity of germplasm resources for Potentilla discolor.Methods Twelve species of germplasm resources for P.discolor were analyzed by ISSR molecular markers.To make up the systematic diagram of genetic relationship by POPGENE 1.32 software,cluster by UPGMA method,and establish the dendrogram.Results A total of 128 ISSR bands were scored for 12 primers,among which 101 were polymorphic bands.The average percentage of polymorphic bands was 78.91%.Genetic similarity coefficient was changed from 0.179 2 to 0.632 5.By cluster analysis,the geographical distribution is mutually related to the relationship of germplasm resources for P.discolor and it was also showed some of P.discolor from the same region were in the same group which presented the rule of geographical distribution in the tested materials.Conclusion The diversity level of the different germplasm resources for P.discolor higher and the relationship of P.discolor correlates with the geographical location in some way.