| ObjectiveJupi Decoction is the Classical prescriptions in synopsis of Golden Chamber,composed of ginger and citrus peel according to the proportion of 1:2,the actual application often replace the ginger with radix aconiti laterlis.At present,a lot of the clinical application of Jupi Decoction on the basis of the party to add or subtract in one application,the market has yet to be seen on the basis of the party of the Chinese patent medicine or health food listed,therefore failed to maximizing the function of the prescription.In addition to the prescriptions of modern pharmacological research also needs to be strengthened.On the basis of a large number of previous experimental study,we have Jupi Decoction(pericarpium citri reticulatae and radix aconiti laterlis)for secondary development,combined with modern preparation process,development of anti-oxidant,anti-fatigue and improve cardiovascular function and various prepearation-Chenpijiang soft capsule.The main ingredients of pericarpium citri reticulatae and pericarpium citri reticulatae 6-Gingerol,Hesperidin and Nobiletin,pharmacokinetic studies,such as to clarify its Plasma concentration in the body change and relevant pharmacokinetic parameters,at the same time,research on correlation between the effect of metabolic enzymes and drug interactions.For the development of Jupi Decoction and its experimental basis for clinical reasonable application.Methodspericarpium citri reticulatae and radix aconiti laterlis effective parts extraction research:(1)pericarpium citri reticulatae essential oil extraction was studied.According to the factors influencing the extraction of essential oil,selected soaking time,extracting time,add water to examine factors,essential oil yield as index,orthogonal experiment design method to screen the optimum process parameters.(2)radix aconiti laterlis,supercritical fluid extraction of essential oil extraction was studied.By the method of single factor investigation combined with uniform design,the main factors influencing the supercritical extraction such as extraction pressure,extraction temperature,parsing,the analytical temperature,pressure,etc,with extraction rate as the index,screening the optimum process conditions.Chenpijiang soft capsule development and quality control in the study:Preparation formulation screening and soft capsule preparation;Thin layer chromatography(TLC)is adopted to establish the preparation the identification method of radix aconiti laterlis,high performance liquid chromatography(HPLC)method is adopted to establish the preparation of the method for the determination of the content of 6-gingerol.Chenpijiang soft capsule anti-oxidant function in vivo and in vitro experimental study:Determination by ultraviolet spectrophotometry,different concentrations of finished product of ability of scavenging DPPH radical,evaluate its in vitro anti-oxidant effect;Aging mice by MDA level can be divided into aging control group and low,medium and high dose group.Continuous lavage after15,30 days,Serum MDA levels,superoxide dismutase(SOD)activity and glutathione peroxidase(GSH-PX)activity were measured.Pericarpium citri reticulatae was evaluated by high performance liquid chromatography(HPLC)method(Hesperidin and Nobiletin)and radix aconiti laterlis(6-Gingerol)main components in rats Plasma concentration,and study its pharmacokinetics.Hesperidin.Nobiletin and drug interactions between 6-Gingerol:Use of human liver microsomal(HLM,mixed components)and specific inhibitors to investigate a phase transformation way of hesperidin,nobiletin,6-gingerol,infer what metabolic enzymes involved in the transformation way;Use of HLM and probe the substrate to investigate inhibition effects of hesperidin,nobiletin,6-gingerol to CYP enzymes.ResultsPericarpium citri reticulatae essential oil optimum extraction conditions for:Soak for 2 hours,the extracting time 10 hours,10 times the amount of water added;radix aconiti laterlis essential oils of supercritical extraction conditions for optimum:radix aconiti laterlis medicinal particle size 30 mu,extraction pressure,28 Mpa.48 ℃ temperature of extraction,parsing Ⅰ pressure 6 Mpa,parsing I temperature 35 ℃ parsing Ⅱ pressure 6 Mpa.parsing Ⅱ temperature 30 ℃,extraction time 3 h.The development of the Chenpijiang soft capsule:According to pharmacopoeia the maximum dosage of Chinese medicinal materials and the optimum proportion of effective parts in accordance with the requirements of the suppression of 0.4g/grain of soft capsule,to determine the dosage of soybean oil,evenly mixing raw materials,and suppress the 10000 grains of Chenpijiang soft capsule;identification of radix aconiti laterlis in the finished product in the chromatogram of the test sample,with radix aconiti laterlis contrast medicinal materials and 6-gingerol reference substance chromatography show the same spot on the corresponding position;According to the high performance liquid chromatography(HPLC)method was determination of 6-gingerol in finished product content of 3.58%(14.31 mg/grain).Chenpijiang soft capsule in vitro anti-oxidant experiment shows that its half the clearance rate of DPPH IC50 was 0.55mg/mL;In vivo experiment of aged mice after 15 days of treatment,the high dose group of MDA content was significantly decreased compared with control group,SOD and GSH-PX activity increased significantly,lower dose antioxidant drug group is not obvious;30 days after administration of each dose group of MDA content was significantly decreased compared with control group,SOD and GSH-PX activity increased significantly obviously,antioxidation.Set up a method that high performance liquid chromatography determination of hesperidin,nobiletin,6-gingerol in rat plasma,methodology to investigate all conform to the requirements of biological sample analysis.Hesperidin and nobiletin,6-gingerol t1/2(β)value respectively was 11.55±2.11,15.07± 2.94,15.40 ± 2.14;Cmax is:1.88 ±0.37,2.55±0.42,0.98± 0.26;AUC0-∞ are:16.59 ± 2.20,16.90 ± 3.91,3.56 ± 0.96.Hesperidin is incubation in the reaction system of human liver microsomal with no apparent metabolic trend,shows that hesperidin in the human body’s metabolism is not made of CYP or FMO need NADPH coenzyme metabolic enzyme mediated.While nobiletin and 6-gingerol shown could soon be metabolic in the reaction system of liver microsomal.nobiletin to 1A2 have strong inhibition(IC50=3.1μM),and 6-gingerol has strong inhibition on 2C19(IC50=9.9μM).In addition,nobiletin to 2C8,2C9 and 2C19 has certain inhibition,and 6-gingerol for CYP1A2,2E1 and 3A4 has certain inhibitory effect.Nobiletin to metabolism of 6-gingerol has obvious inhibitory effect,and the inhibition is time dependent inhibition(TDI),high concentration(10μM)can increase 6-gingerol in the reaction system of human liver microsomal incubation after 30 minutes the residual rate of 6.4 times(from 14%to 93%),in low concentration(1μM),can also be increased 2.6 times(from 14%to 38%).Hesperidin on 6-gingerol metabolism and hesperidin/6-gingerol to nobiletin has little effect on the metabolism.ConclusionThis paper established the optimum process parameters of radix aconiti laterlis supercritical extraction and pericarpium citri reticulatae essential oil extraction,amplified by pilot experiment,good reproducibility,suggests that the optimum process conditions stable and feasible.Chenpijiang soft capsule each index of preparation all conform to the requirements,set up by the thin layer identification and content determination method is accurate,reliable,simple,specific,which can effectively control the quality of the products.In this article,Chenpijiang soft capsule in vitro anti-oxidant test results show that the anti-oxidant effects quite BHT,antioxidant effect is stronger.In vivo experiments show that it can significantly reduce the content of MDA in aged rats,and significantly increase the activity of SOD and GSH-PX.This article studies the main ingredient in pericarpium citri reticulatae and radix aconiti laterlis,hesperidin,nobiletin,6-gingerol inhibition of metabolic enzymes and metabolic interaction between each other,according to a study found that nobiletin has obvious inhibitory effect of 6-gingerol metabolism,due to the nobiletin for CYP 1A2 have stronger inhibition,speculated that I phase metabolism of 6-gingerol mainly mediated by this CYP1A2,therefore,when nobiletin used in combination with 6-gingerol,because inhibition of this CYP1A2 metabolic pathway,to reduce the body remove for 6-gingerol,increase the efficacy of the role. |
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