ABSTRACT
OBJECTIVE To establish the fingerprint of total saponins from Mussaenda pubescens， and to study the spectrum- effect relationship of its hepatoprotective activity. METHODS Ten batches of total saponins from M. pubescens from different origins were prepared using 75% ethanol as solvent. High-performance liquid chromatography （HPLC） and the Similarity Evaluation System for Traditional Chinese Medicine Chromatographic Fingerprints （2012 edition） were used to draw the fingerprints of 10 batches of total saponins from M. pubescens. The similarity evaluation and identification of common peaks were conducted. The same HPLC method was adopted to determine the contents of five triterpenoid saponins （mussaendoside H， mussaendoside U， mussaglaoside C， mussaendoside G and mussaendoside O）. The hepatoprotective effect of total saponins from M. pubescens was investigated by establishing carbon tetrachloride-induced acute liver injury model mice， and the spectrum-effect relationship was studied by using grey correlation analysis. RESULTS There were 11 common peaks in 10 batches of total saponins from M. pubescens， 5 of which were identified， i.e. mussaendoside H （peak 3）， mussaendoside U （peak 7）， mussaglaoside C （peak 8）， mussaendoside G （peak 9） and mussaendoside O （peak 11）； the similarities of 10 batches of samples ranged 0.940- 0.991. Average contents of mussaendoside H， mussaendoside U， mussaglaoside C， mussaendoside G， mussaendoside O were 0.01- 0.05， 0.10-0.21， 0.10-0.18， 0.03-0.08， 0.20-0.40 mg/g， respectively. Ten batches of total saponins from M. pubescens could generally reduce the contents of alanine aminotransferase （ALT） and aspartate aminotransferase （AST） in serum， and tumor necrosis factor-α （TNF-α）， interleukin-6 （IL-6） and IL-1β in liver tissue of model mice （P＜0.05 or P＜0.01）. The E-mail：13878195336@139.com correlation between the common peak areas and the contents of ALT， AST， TNF-α， IL-6 and IL-1β were 0.602-0.757， 0.585-0.833， 0.593-0.795， 0.618-0.820， 0.607-0.804， respectively； the peaks with high correlation were peaks 11， 9 and 8 in order. CONCLUSIONS Ten batches of total saponins from M. pubescens have similar components， and the average contents of mussaendoside H， mussaendoside U， mussaglaoside C， mussaendoside G and mussaendoside O are different. The batches of samples have a certain degree of hepatoprotective effect； mussaendoside O， mussaendoside G and mussaglaoside C may be its main active components.
ABSTRACT
ObjectiveTo explore the effect of Gelsemium elegans combined with Mussaenda pubescens on efflux transporter breast cancer resistance protein （BCRP） and cytochrome P450 3A11 （CYP3A11） and their attenuation mechanism， and to investigate whether the nuclear receptors were involved in such regulation by intervening it with nuclear receptor activators. MethodC57BL/6 mice were divided into the blank group， G. elegans （GE， 0.25 g·kg-1）group， GE + M. pubescens （MP） （0.25 g·kg-1+10 g·kg-1） group， GE + pregnane X receptor （PXR） activator （rifampicin）（GE + Rif，0.25 g·kg-1+50 mg·kg-1） group， GE + MP + Rif （0.25 g·kg-1+10 g·kg-1+50 mg·kg-1） group， GE + constitutive androstane receptor （CAR） activator （1，4-Bis ［2-（3，5-Dichloropyridyloxy）］ benzene， TCPOBOP）（GE + TCP， 0.25 g·kg-1+0.5 mg·kg-1） group， and GE + MP + TCP （0.25 g·kg-1+10 g·kg-1+0.5 mg·kg-1） group. The medication lasted for 14 successive days. One hour after the last administration， the mice were sacrificed by cervical dislocation and the liver tissue was harvested. The left liver tissue was stained with hematoxylin- eosin （HE） for observing the pathological changes. The right liver tissue was used for BCRP and CYP3A11 mRNA and protein expression detection by real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） and Western blot. ResultThe survival rates of mice in the GE + Rif group， GE group， and GE + MP group were 25% （the lowest）， 40%， and 80%， respectively， and no death was observed in the other groups. Compared with the obvious lesions in the liver cells of the GE group， the pathological changes in liver cells of the GE + MP group were alleviated， while those in the GE + Rif group were worsened. Compared with the GE + Rif group， the GE + MP + Rif group exhibited relieved pathological changes in liver cells. Both the GE + TCP group and the GE + MP + TCP group showed mild liver lesions. The comparison with the GE + MP group revealed that the pathological changes in the GE + MP + TCP group were slightly relieved. Compared with the blank group， the expression of BCRP protein and mRNA in GE group were significantly decreased （P<0.05，P<0.01）.The expression of CYP3A11 protein in GE group were significantly decreased （P<0.01）. Compared with the GE group， the GE + MP group displayed remarkably up-regulated BCRP protein and mRNA expression （P<0.05，P<0.01） and CYP3A11 protein expression （P<0.05）， but slightly up-regulated CYP3A11 mRNA expression. Compared with the GE group， the GE + Rif group exhibited down-regulated BCRP protein expression （P < 0.05）. The protein and mRNA expression levels of BCRP were lower in the GE + MP + Rif group than in the GE + MP group （P<0.05，P<0.01）. The PXR activator rifampicin regulated BCRP before and after the combination of G. elegans with M. pubescens. The CYP3A11 protein and mRNA expression levels in the GE + TCP group were higher than those in the GE group （P<0.05，P<0.01）. Compared with the GE + MP group， the GE + MP + TCP group showed up-regulated CYP3A11 protein and mRNA expression （P<0.05，P<0.01）. CAR activator TCPOBOP also had a regulatory effect on CYP3A11 before and after the compatibility of G. elegans with M. pubescens. ConclusionThe attenuated toxin after the combination of G. elegans with M. pubescens is closely related to the efflux transporter BCRP and the drug-metabolizing enzyme CYP3A11.
ABSTRACT
This article was aimed to study the pharmacokinetics of mussaenoside in rats. Gardenoside was used as the internal standard in the content determination of mussaenoside in rat plasma with HPLC. The chromatographic conditions were Diamonsil C18 (2) column (150 mm í 4.6 mm, 5 μm), mobile phase of methanol:0.5% glacial acetic acid (30:70), flow amount of 1.0 ml/min, detection wavelength at 238 nm, column temperature at 30℃. Compartment model fitting of average plasma concentration in rats - time data had been conducted after mussaenoside was given to mice through single intravenous injection with the help of DAS 2.0 pharmacokinetics intelligence analysis software. The results showed that the plasma concentration using 2-comparment model intravenous injection calculation method, which corresponded well to that of actually measured (weighting of 1), was proved superior to other models. The distribution half-life time (t1/2α) was 9.892 min and the elimination half-life time (t1/2β) was 55.384 min. It was concluded that the established method was convenient, accurate and stable, which can be used for the study on phar-macokinetics of mussaenoside. Mussaenoside was distributed rapidly in the body of rats. The main process of mus-saenoside in the body of rats was elimination. The metabolic rate was at moderate speed.