| Relinqing Granule (RG), a common prescribed preparation based onHerba Polygoni Capitati extract alone, has been clinically used inMiao-nationality herbal medicine system for the treatment of urinary tractinfections and pyelonephritis. RGwas approved for usage with the approvalof the ministry of health production in1970s and included in the Guizhouprovince local standards (Qian D/WS-299-89). RG was selected as one ofthe national protected traditional medicines in2002and further as one ofnational social secutity medicines in2004. RG takes the pricing power alonewith the permission of the national development and reform commission.RG is also the only ‘Miao Medicine’ listed in the Chinese pharmacopoeia(CP, version2010) for the treatment of urinary tract infection andpyelonephritis, and see above syndrome. Given tothe stable curative effect,high safety and reliable quality after many years of application of RG, itenjoys a good reputation and well received by the majority of doctors andpatients.There are, however, not clear of the efficacy material basement of RGin the treatment of urinary tract infectionsandpyelonephritis.The sameambiguity is manifested in the mechanism of its pharmacodynamic effects,biological transformation and metabolic processes in human body. Studieson RG and Herba Polygoni capitati found that they are rich in phenolic acids,flavonoidsand tannins, which possessing anti-oxidative, anti-inflammatory, antivirus and antibacterial activities. Hower, whether these so called ‘activeingredients’ were exactly the ones be absorbed into the bloodstream andexert pharmacodynamic effects is unclear.The amount of Gallic acid (GA) and Protocatechuic acid (PCA) in RGis large, and GA is designated as the index to control the quality of RG in CP.There are, however, few reports of the pharmacokinetics and metabolism ofthe active ingredients of RG in animal or human body.For the purpose ofinvestigating and clarifying the physiological disposition of GA and PCAafter oral administration of RG, systematical study on the absorption,distribution, metabolism, and excretion (ADME) of GA and PCA in ratplasma and other biological fluids were conducted. The results are asfollows:1. Pharmacokinetics of RG in plasmaA sensitive, selective and rapid liquid chromatography-massspectrometry (LC-MS/MS) method was estabilished for simultaneousquantification of GA and PCA in rat plasma.GA, PCA and bergenin (internalstandard, IS) were extracted from plasma and separated on a PhenomenexC18column. The detection of GA and PCA were carried out by multiplereaction monitoring (MRM) on a ThermoTSQUPLC-MS/MS system with anESI interface in the negative ion mode.MRM was performed at unitresolution using the mass transition ion-pairs at m/z169.181'125.268forGA, m/z152.918'109.244for PCA and m/z326.922'192.167for IS. Linearity, specificity, precision, accuracy, recovery, and stability werevalidated and the results show that all validation parameters met therequirements of the relevant guidance criteria.This validated LC-MS/MSmethod could be applied to determine the concentration of GA and PCA inrat plasma precisely.Using the established LC-MS/MS method, the pharmacokinetics of GAand PCA in male rats was studied after different dosage of intragastric (0.36,1.08and2.16g·kg-1) administration of RG, respectively.After oral administration of0.36,1.08and2.16g·kg-1of RG, respectivevalues of pharmacokinetic parameters for GA were: t1/2128.52,93.72, and114.70min; Cmax245.98,477.20, and805.76ng·ml-1;AUC0-∞53776.77,97385.52,204204.47μg·L-1·min-1. Linear pharmacokinetics was establishedbased on high correlation coefficients (γ>0.98) of pharmacokineticparameters.Corresponding values for PCA were: t1/242.81,90.15, and49.80min;Cmax11.90,24.66, and31.04ng·ml-1; AUC0-∞943.88、2943.38、4222.79μg·L-1·min-1. Linear pharmacokinetics was established based on highcorrelation coefficients (γ>0.90) of pharmacokinetic parameters.2. Distribution study of RGSimple and effective LC-MS/MS methods were developed to determinethe concentration of GA and PCA in different tissues.These methods werequick, precise, and reproducible, and could be used to quantify GA and PCA in rat heart, liver, spleen, lung, kidney, and brain after oral administration(1.08g·kg-1). It was the first time to study the tissue distribution of GA andPCA in rats after intragastric administration.The result indicated that1h afteroral administration of RG, GA mainly distributed in kidney, lung, and liverwith the concentration of1218.62,258.08and231.55ng·g-1, respectively;PCA mainly distributed in kidnry and lung with the concentration of43.98and19.48ng·g-1, respectively.3h after oral administration of RG (1.08g·kg-1)，only GA can be detected in kidney tissue with the concentration of143.54ng·g-1.3. Excretion study of RGLC-MS/MS methods were established for the quantification of GA andPCA in rat urine and feces, respectively.These methods were quick, precise,and reproducible. The assays were successfully applied to the excretionstudy of GA and PCA in rat urine and feces.After oral administration of0.36,1.08and2.16g·kg-1RG, thecumulative amount of GA in urine in48h were:108.80、361.44and950.38μg with the excretion rate13.19,14.60, and19.20%, respectively; Thecumulative amount of GA in feces were21.25、93.16and192.10μg with theexcretion rate2.58,3.76, and3.88%, respectively. Less than23.08%prototype of GA was excreted from urine and feces path indicating that GAis extensively metabolized in rat. Corresponding values for the cumulative amount of PCA in urine in48h were:6.76、29.55and60.69μg with the excretion rate10.79,15.72, and16.15%, respectively; The cumulative amount of PCA in feces were3.56、6.74and12.19μg with the excretion rate5.68,3.59, and3.24%, respectively.Less than19.39%prototype of PCA was excreted in total urine and fecespath indicating that PCA is also extensively metabolized in rat.4. Optimization extraction of Herba Polygoni CapitatiResearch on Herba Polygoni capitati found that it is richedin flavonoids,while these flavonoids compounds cannot be extracted efficientlyby water.In this section, the conditions for extraction of quercitrin and total flavonoids(TF) from Herba Polygoni Capitati were optimized by using single-factortest combined with the response surface methodology (RSM). A centralcomposite design (CCD) was adopted to investigate the effects of threeindependent variables including solvent composition (%), solvent-materialratio (mL·g-1), and extraction time (min) on the responses, quercitrin and TFyiedls. The optimized conditions of the extraction are as follows: ultrasonicextraction; ethanol concentration,65.63%; solvent-material ratio,10.55:1(mL·g-1); extraction time,54.33min. As a result, the observed values ofquercitrin and TF could be satisfactorily achieved2.44and106.88mg·g-1,respectively, under this optimized condition. |
PDF Full Text Request