Study On The In Vivo/in Vitro Metabolism And Pharmacokinetics Of The Xinjiang Pomegranate Peel Polyphenols’ Purification Main Components Of Anti-tumor

Objective: In this study,the metabolic and pharmacokinetics of punicalagin and ellagic acid in pomegranate peel were studied in vivo and in vivo,and the pharmacokinetics of antitumor activity.Methods:1.The in vitro metabolic model of intestinal microflora in rats was established.The metabolites of punicalagin in the intestinal flora incubation system were identified and analyzed by HPLC-ESI-TQ-MS.2.To establish the metabolic model of liver microsome and liver S9 in vitro,the activity of esterase in rat liver and guinea pig liver microsomes and rat liver S9 fragments were characterized and compared by UPLC-ESI-QTRAP-MS.The UPLC-ESI-Q TOF-MS was used to identify and analyze the metabolites of punicalagin and ellagic acid in rat liver microsomes Ⅰ phase CYP450 enzyme,esterase and Ⅱ phase UGT enzyme.And the metabolic pathways of punicalagin and ellagic acid were analyzed.3.The plasma,urine,feces and bile samples were collected after the administration of the rats.The metabolites of each biological sample were identified and analyzed by UPLC-ESI-QTOF-MS.4.Punicalagin using rat oral administration route,ellagic acid using rat oral and intravenous injection route,at different times in the eye canthus blood.Punicalagin and ellagic acid were analyzed by HPLC and UPLC-ESI-QTRAP-MS.The pharmacokinetic parameters were analyzed by PK solution 2.0TM.Results: 1.HPLC and LC-MS/MS detection methods were established.After being incubated by HPLC,the punicalagin was almost completely metabolized after incubation with the intestinal flora of rats,and M1,M2 and M3 were metabolized.According to the peak The time was initially speculated and verified that M3 was ellagic acid.By LC-MS/MS,it was further confirmed that M3 was ellagic acid and speculated that M2 might be Gallagic acid.2.The liver metabolites of liver microsomes and liver S9 fragments were established.UPLC-ESI-QTRAP-MS was used to determine and analyze the metabolites produced by esterase catalyzed by rat liver and guinea pig liver microsomes and rat liver S9 fragments,The results showed that both had esterase activity,the activity of esterase in guinea pig liver microsomes was significantly different from that in rat liver microsomes and rat liver S9 fragments,but the activity of esterase in rat liver microsomes was correlated with the activity of esterase no significant difference.The UPLC / ESI-QTOF-MS was used to identify and analyze the phase I and Ⅱ metabolites of punicalagin and ellagic acid in rat liver microsomes,the results showed that 6 metabolites of M1-M6 were detected by punicalagin,and the product M6 was ellagic acid compound,and the metabolites of phase II and phase I were identical.Ellagic acid detected a metabolite,and the metabolite may be a glucuronide metabolite.3.In addition to the detection of two metabolites in the plasma sample and verified that the punicalin and ellagic acid compounds were tested,no punicalagin-related metabolites were detected in the urine,feces and bile samples by UPLC-ESI-QTOF-MS detection technology analysis.No ellagic acid-related metabolites were detected in plasma,urine,feces and bile by UPLC-ESI-QTOF-MS detection technology analysis.4.HPLC and UPLC-ESI-QTRAP-MS quantitative detection methods were established.After the administration of punicalagin(300mg/kg),the plasma samples were detected by HPLC method.The plasma concentration-time curve of punicalagin in rat plasma was drawn.The main pharmacokinetic parameters were:AUC(0-t)(243.40±111.62)ng·h·m L-1、T1/2a(42.03±0.8)h、T1/2d(2.68±1.76)h、T1/2e(18.87±17.92)h、Tmax(1.67±0.52)h、Cmax(26.33±5.65)ng·m L-1、Ka(0.42±0.25)h-1;The plasma samples were detected by UPLC-ESI-QTRAP-MS after oral administration(90.0 mg / kg)and intravenous injection(1.0 mg / kg).The plasma concentration-time curves of ellagic acid in plasma of rats were given orally and intravenously.The main pharmacokinetic parameters of oral administration were: AUC(0-t)(71.89±32.02)ng·h·m L-1、T1/2a(2.52±2.96)h、T1/2d(1.89±1.14)h、T1/2e(17.99±11.20)h、Tmax(1.11±1.30)h、Cmax(8.86±4.65)ng·m L-1、Ka(0.57±0.45)h-1,the main pharmacokinetics of intravenous administration were: AUC(0-t)6(362.15±126.8)ng·h·m L-1、T1/2a(0.09±0.01)h、T1/2d(0.65±0.21)h、T1/2e(7.24±5.96)h、Tmax(0.03±0.00)h、Cmax(2512.29±640.13)ng·m L-1、Ka(7.64±0.83)h-1.In addition,the absolute bioavailability of ellagic acid was 24.49%.Conclusion: Through the data integration,the punicalagin produced 9 metabolites in vitro and in vivo,and ellagic acid produced a metabolite in vitro and in vivo.The metabolic process was an open two-compartment model with obvious absorption phase,distribution phase and elimination phase;ellagic acid oral administration is also consistent with the two-compartment model;ellagic acid intravenous injection of plasma concentration-time curve in the elimination phase showed a sharp downward trend,which can be rapidly metabolized;ellagic acid absolute bioavailability low,may be related to its strong lipophilicity and liver first effect.