The Study On Absorption Mechanism Of Evodia Rutaecarpa With Schisandra As Prescription Compatibility

ObjectiveSchisandra is a commonly used clinical adjuvant in TCM which is widely used in a variety of prescriptions compatibility of traditional Chinese medicine. This research selected Schisandra powder as research object to study the effect of Schisandra compatibility on the intestinal absorption of Evodia Rutaecarpa by establishing the rat in situ intestinal pursion model. To study the effects of Schisandra on the expression of intestinal mucosa transporter through RT-PCR technology and Western Blot, in order to reveal the prescription compatibility mechanism of adjuvant of Schisandra from the perspectives of intestinal absorption kinetics.Methods1. Prepare alcohol extractive and water extractive of Evodia rutaecarpa with the method of ethanol-backflow extraction and water decoction. Establish the detection method of Evodiamine (Evo) and Rutaecarpine (Rut) by high performance liquid chromatography(HPLC).2. Establish the rats in vivo intestinal circulation to examine the effects of Schisandra compatibility on the intestinal absorption of major constituent of alcohol extractive of Evodia Rutaecarpa. To calculate the constant (Ka) and opparent permeability coefficient (Papp) of the two alkaloid on the basis of the Evodia Rutaecarpa and Evodiamine, then compare with the effects of P-gp inhibitor, namely, verapamil(VP).3. Establish the rats single pass intestinal perfusion model to study intestinal absorption characters of different proportions of Schisandra and Evodia Rutaecarpa water decoction. To calculate the constant (Ka) and opparent permeability coefficient (Papp) of the two alkaloid of Evodia Rutaecarpa, then compare with the effects of P-gp inhibitor, namely, verapamil(VP).4. The animal models established after14days’animal gastric perfusion with medicines of different experimental groups, then take the intestinal tissue of rats to examine the effects of Schisandra on the expression of intestinal mucosa transporter, P-gp, MRP2, and Nrf-2through RT-PCR technology and Western Blot. To probe the proposed mechanism of the effect of Schisandra on the expression of intestinal mucosa transporter.Results1. Chromatographic method to simultaneously test the Evodiamine(Evo)and Rutaecarpine(Rut) has been determined:acetonitrile-water (49:51) as the mobile phase; flow rate of lml·min-1; dual wavelength detection, Evodiamine (Evo)225nm, Rutaecarpine (Rut)345nm; sample size,20μ1; column temperature,25℃. The Evodiamine(Evo) measured in the alcohol extractive of Evodia Rutaecarpa is0.427%, Rutaecarpine(Rut) is0.405%; The Evodiamine(Evo) contained in Evodia Rutaecarpa water decoction is3.62%, Rutaecarpine (Rut) is3.27%.2. After the compatibility of Schisandra, both the constant(Ka) and opparent permeability coefficient of Evodiamine(Evo) and Rutaecarpine(Rut) were significantly increased (P<0.05). The inhibitor verapami1(VP) of Positive drug P-gp has also significantly increased the constant(Ka) and opparent permeability coefficient of two alkali (P<0.05). However, when take the concentration range of Schisandra into consideration, the constant (Ka) of Evodiamine(Evo) and Rutaecarpine(Rut) did not change obviously.3. Conduct the intestinal absorption experimentation with Evodiamine (Evo) and Rutaecarpine (Rut) as index components, compared the water decoction with different proportions of Schisandra and Evodia Rutaecarpa with the single decoction of just Evodia Rutaecarpa, both constant(Ka) and opparent permeability coefficient of Evodiamine(Evo) and Rutaecarpine(Rut) have significantly decreased (P<0.05). Moreover, compared the group with the ratio1:2of Schisandra and Evodia Rutaecarpa with the ratio1:1group, both constant(Ka) and opparent permeability coefficient also have significantly decreased. The addition of inhibitors of verapamil(VP) have little effect on the intestinal absorption of principal components of the Evodia Rutaecarpa water decoction.4. Low concentration Schisandra can up regulate the expression of the-mRNA and protein of P-glycoprotein, and high concentration Schisandra down regulate the expression;Low concentration Schisandra can down regulate the expression of Nrf2, and high concentrations Schisandra up regulate the expression; Schisandra play the same role in the expression of MRP2. The Nrf2agonist t-BHQ can induce the expression of three gene. While the P-gp agonist bifendate only induce the expression of P-glycoprotein and inhibit the expression of the other two genes Nrf2and MRP2.ConclusionIn the experiment of intestinal absorption of ethanol extract, the compatibility of Schisandra and Evodia Rutaecarpa can increase intestinal absorption of principal components of Evodia Rutaecarpa. The absorption enhancing mechanism may be explained by that high concentration Schisandra down regulated the expression of P-glycoprotein, thereby inhibiting the exocytosis of P-glycoprotein drug pump on drugs. But in the experiment of intestinal absorption of water decoction of different proportions of Schisandra and Evodia Rutaecarpa, Low concentration Schisandra up regulate the expression of P-glycoprotein, thereby enhancing the exocytosis of P-glycoprotein drug pump on drugs, result in inhibiting the absorption of drugs.