Anti-gout Effect Of Extracts From Compound Selaginella Moellendorffii And Its Pharmaceutical Study

Gout belongs to the areas of Chinese arthralgia syndrome and the main therapeutic method was heat-clearing and diuretic detoxification.The compound Selaginella moellendorffii consist of Selaginella moellendorffii Herba,Smilacis glabrae Rhizoma and Plantaginis Semen.Selaginella moellendorffii has many clinical efficacies such as removing stasis,clearing heat,excreting damp and stopping bleeding and it also has significantly effect of inhibiting Xanthine oxidase and uric acid production.It has been used to treat gouty arthritis in folk.Smilacis glabrae Rhizoma is an effective medicine which can turbid toxic,thus it has the effect of detoxication,dehumidification and easing joint movement.Plantaginis Semen has the role in clearing heat,excreting damp,dredging stranguria and diuresing,hence,it can promote the excretion of uric acid by the diuretic.Three-drug compatibility could HSXZ,enhancing efficacy,improving symptoms and lowering serum uric acid concentration.This paper was aimed to study pharmacologocal function of Selaginella moellendorffii compound,the possible mechanism and its material basis.In the meantime,we developed the study of its preparation and molding process so as to provide the basis for the development of a historical preparation used to treat HUA and acute gouty arthtitis.Objective: To study the pharmacologocal function and its possible mechanism of compound Selaginella moellendorffii on Hyperuricemia and Acute gouty arthritis;to study the process of water extraction and alcohol precipitation and preparations molding process of compound of Selaginella moellendorffii based on the efficacy study.Methods: Kunming mice as experimenal subjects,the experimental model of HUA was induced by potassium oxonate with intraperitoneal injection and adenine;uric acid(UA),creatinine(Cr)and blood urea nitrogen(Bun)were as indexes to research the effect of SME on HUA mice by biochemistry.SD rats as experimental subjects,the experimental model of acute gouty arthristis was induced by injecting monosodium urate(MSU)crystals to the right hind paw;MPO,SOD and MDA were as indexes to research the effect of SME on SD rats by biochemistry;TNF-? and IL-1? were as indexes to study the anti-inflammatory action of SME on AGA rats by elisa assay;the injuries of joint were evaluated by histopathologic sections;HPLC was applied to analyze the main chemical ingredient of the aqueous extracts of Selaginella moellendorffii;Kunming mice as experimenal subjects,the experimental model of HUA was induced by potassium oxonate with intraperitoneal injection,UA as index to assess the effect of CSME on HUA mice;SD rats as experimental subjects,the experimental model of acute gouty arthristis was induced by injecting monosodium urate(MSU)crystals to the right hind paw;NO was applied toassess the effect of CSME on SD rats by biochemistry;the serum levels of PGE-2,IL-1?,IL-6 and IL-8 were applied to study the effect of CSME on AGA rats by elisa assay;the injuries of joint were also evaluated by histopathologic sections;Kunming mice as experimenal subjects,the clinical medication safety of Selaginella moellendorffii and its compound was evaluated by gavage with the maximum dosage;Experiment on the aqueous extracts of compound Selaginella moellendorffii,the extraction technology of compound Selaginella moellendorffii was optimized by multi-index comprehensive analysis using 6-C-?-Dglucopyranosul-8-C-?-D-xylopyranosylapigonin(Y ?),astilbin and extracting rate as indexes and the extracting time,times and the amount of wateras inspecting factors;astilbin,extracting rate and 6-C-?-D-glucopyranosul-8-C-?-Dxylopyranosylapigonin(Y?)as indexes and the solution density,final ethanol concentration,alcohol precipitation time andalcohol precipitation temperature as inspecting factors were used to optimize the purification process of CSM by multi-index comprehensive analysis;the kind of excipient and the ratio of excipient and wetting agent were optimized with granulation,water content,liquidity and hygroscopicity index.Results:(1)Anti-hyperuricemic effects of SM extract in hyperuricemic mice: compared to normal control group,serum urate,Cr and Bun levels were significantly increased(P<0.01).Compared to hyperuricemic control group,Allopurinol presented a significant reduction on urate level but an increase on Cr and Bun levels of hyperuricemic mice,however,Treatments with low,medium and high dose SM extracts were able to reduce serum urate levels and reduce the Cr and Bun levels at the same time,with a dose-depenfent manner.(2)Anti-inflammatory effects of SM extract in SD rats: Compared to the normal control group,the ankle circumference of rats and the level of MPO,IL-1?,TNF-? and MDA in MSU group were found to be significantly increased(P<0.01).Yet,a decrease of SOD level was also found;Compared with the MSU group,the ankle circumference and the level of MPO,IL-1?,TNF-? and MDA were decreased but the level of SOD was increased with a dose-depenfent manner when Treated with low,medium and high dose SM extracts.Among that the curative effect of high dose group of SM extract was the most notable(P<0.01),which was equal to colchicine.Seen from histopathological sections,the cartilage and synovium in normal control group were resulted in slight inflammation after intra-articular injection of sterile endotoxin-free saline.In the MSU group,a mass of neutrophil infiltration was observed.Yet,treatments with colchicine and SM extracts were able to attenuate the neutrophil infiltration.(3)HPLC chromatogram was applied for examining constituents from aqueous extract.Four flavone C-glycosides and one flavone O-glycosides have been isolated and identified from SM in our research group.As reported in literature,the constituents were flavone-C-glycosides,6,8-di-C-?-Dglucopyranosylapigenin(?),6-C-?-D-glucopyranosul-8-C-?-Dxylopyranosylapigonin(?),6-C-?-D-xylopyranosyl-8-C-?-Dglucopyranosulapigonin(?),5-carboxymethy-4?-hydroxyflavone-7-O-?-D glucopyranoside(?)and amentoflavone.(4)Anti-hyperuricemic effects of CSM extract in hyperuricemic mice: compared to normal control group,the serum urate level was significantly increased in hyperuricemic control group(P<0.01).Compared to hyperuricemic control group,administrations with allopurinol and CSM extract could reduce the UA level in plasm,among that CSM-3 group showed better low-urate level than CSM-2.And it was also superior to the group of RSG and SP used alone.(5)Anti-inflammatory effects of CSM extract in SD rats: Compared to the normal control group,the ankle circumference of rats and the level of NO,PGE-2,IL-8 and IL-1 ? in MSU group were found to be significantly increased(P<0.01);meanwhile,the rats in CSM,RSG and SP extract groups had a decrease in ankle circumference and the NO,PGE-2,IL-8,IL-1? levels Compared to the MSU group.Taken all index results together,the effect of CSM-3 was best in all extract groups(P<0.01)and it has no significant difference with colchicine(P>0.05).It is remarkable that the effect of CSM-3 was equal to or even better than colchicine group on some indicators.Seen from histopathological sections,the cartilage and synovium in normal control group were resulted in slight inflammation after intra-articular injection of sterile endotoxin-free saline.In the MSU group,a mass of neutrophil infiltration was observed.Yet,treatment with colchicine inhibited the neutrophil infiltration.Furthermore,administration with TCM extract could prevent the MSU crystalinduced injury,especially the CSM-3 group.(6)Acute toxicity test of SM and its compound: SME and CSME were gavaged at maximum dose,which was 640 and 480 times that of adults respectively,and mice in each group had no death and obvious symptoms of poisoning.Also,Liver and kidney tissue sections of each mice has no obvious lesions,which can be considered clinical safe drug-use.(7)The optimization of the extraction technology of compound Selaginella moellendorffii: the optimal extraction technology was as follows: decocted 3 times with 12 times the amount of water for 1 hour each time;the optimal depuration technology was as follows: the relative density of liquor 0.5g·mL-1,adding alcohol until the concentration of ethanol at 60%,stilled 36 h at 15?.(8)The preparation process of CSM: The dosage form is granules;the granulating process condition: ratio of raw materials: mannitol: dextrin was 1:0.375: 1.125,wetting agent was 95% ethanol and its amount was 0.4mL·g-1.Conclusion:(1)Selaginella moellendorffii were able to prevent the increase of serum uric acid and the renal damage on HUA mice.The aqueous extracts of Selaginella moellendorffii could improve the swell of joints and the pathological change of synovial membrane and surrounding tissues on acute gouty arthritis,and its mechanism was probablly related to the effect of antioxidant and the inhibition of serum TNF-? and IL-1? levels.(2)The aqueous extract of compound Selaginella moellendorffii has the effects of lowering-urate acid in HUA mice and treating the acute gouty arthritis.Among that,the ratio of 3:1:1 was the best.The mechanism was likely to inbibit the synthesis and release of inflammatory cytokines.(3)Selaginella moellendorffii and its compound were administrated by the dose of equivalent to 640 times and 480 times of the adult respectively,yet they were no toxicity and good safety for clinical treatment.(4)The optimization of water extraction and alcohol precipitation technology was proven to be stable,reasonable and feasible,and these provide a basis for the development and application.(5)In this study,the selected dosage and accessories were reasonable,and the molding process was simple and feasible.These provide an experimental basis for the industrial production of compound Selaginella moellendorffii.