ABSTRACT
OBJECTIVE：To study the effects of Mahuang xixin fuzi decoction on Toll-like receptors （TLRs）response and cytochrome C oxidase （Cyt-CO）-mediated apoptosis regulation in mice with influenza disease of kidney-yang deficiency. METHODS：Totally 48 male Balb/c mice were randomly divided into normal group （n＝12）and modeling group （n＝36）. The modeling group was intraperitoneally injected with estradiol benzoate solution （8 mg/kg）and intranasally injected with influenza virus H 1N1（20 μL/mice）to establish the influenza disease compound model of kidney-yang deficiency. After modeling ，the mice were randomly divided into model group ，positive drug group （Oseltamivir phosphate capsules ，0.195 g/kg），Mahuang xixin fuzi decoction group （1.802 g/kg，by crude dru g），with 12 mice in each group. Each group was given relevant medicine intragastrically，normal group and model group were given， corresponding volume of normal saline intragastrically 20 mL/kg，once a day ，for consecutive 6 days. During admi-nistration，body weight and anal temperature of mice were mail：xsy407861520@163.com measured daily ；the percentage of initial body weight was calculated. After last medication ，the organ （spleen，thymus and lung ）indexes were calculated ；the pathological changes of lung tissue were observed. The viral load of influenza A virus H 1N1 in lung tissue was detected （reflected by M gene mRNA expression）；mRNA expressions of TLR3，TLR7，myeloid differentiation factor （MyD88）and Caspase- 3 in cardiac tissue as well as the activity of Cyt-CO and the content of cytochrome C （Cyt-C）were also determined. RESULTS ：Compared with normal group，initial body weight percentage and anal temperature of the model group continued to decrease （P＜0.05）；the spleen and thymus indexes were decreased significantly （P＜0.05），while lung index was increased significantly （P＜0.05）；the lung tissue lesions were serious. Viral load in lung tissue ，mRNA expressions of TLR 3，TLR7，MyD88 and Caspase- 3 in cardiac tissue as well as the content of Cyt-C were increased significantly （P＜0.05 or P＜0.01），while the activity of Cyt-CO in cardiac tissue was significantly decreased （P＜0.01）. Compared with model group ，initial body weight percentage and anal temperature of mice in Mahuang xixin fuzi decoction group showed an increasing trend from the fourth day of administration （P＜0.05 or P＜0.01）. The spleen and thymus indexes were increased significantly （P＜0.05），while the lung index was significantly decreased （P＜0.05）；the pathological injury of lung tissue was significantly improved ；viral load in lung tissue ，mRNA expressions of TLR 3 and Caspase- 3 as well as the content of Cyt-C in cardiac tissue were decreased significantly （P＜0.05 or P＜0.01），while the activity of Cyt-CO was increased significantly in cardiac tissue （P＜0.01）. CONCLUSIONS ：Mahuang xixin fuzi decoction can improve influenza disease of kidney-yang deficiency in mice ，the effect may related to inhibit TLRs response and apoptosis regulation pathway mediated by Cyt-CO.
ABSTRACT
OBJECTIVE：To establish a method for simultaneous determination of calycosin glucoside ，ononin，calycosin， formononetin，astragaloside Ⅳ，isoastragaloside Ⅱ，cycloastragenol and isoastragaloside Ⅰ in Astragalus membranaceus before and after bidirectional solid fermentation with Cordyceps kyushuensis ，and to investigate the effects of fermentation on the contents of above 8 components in A. membranaceus . METHODS ：HPLC-DAD-ELSD was adopted. The determination was performed on Agilent 5 TC-C18 column with mobile phase consisted of 0.1％ formic acid aqueous solution-acetonitrile （gradient elution ）at the flow rate of 1 mL/min. The column temperature was set at 30 ℃. DAD detection wavelength was set at 260 nm，ELSD evaporation tube temperature was 100 ℃，atomizer temperature was 80 ℃，carrier gas flow rate was 1.6 L/min；injection volume was 15 μL. RESULTS：The eight components had a good linear relationship within their respective ranges of concentration （all R2＞0.999 0）； RSDs of precision ，stability and repeatability tests were all less than 3％（n＝3 or n＝6）；the recoveries was 97.88％-101.32％， and RSDs were 1.22％-2.39％（n＝6）. Setting the content of components in unfermented A. membranaceus as 100％，after bidirectional solid fermentation with C. kyushuensis ，the change rates of 8 components were －98.51％，－96.41％，－94.74％， －96.40％，289.20％，20.25％，－75.05％，562.46％，respectively. CONCLUSIONS ：After fermentation with C. kyushuensis ，the contents of active components as astragaloside Ⅳ，isoastragaloside Ⅰ and isoastragaloside Ⅱ can be increased significantly in A. membranaceus .