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Objective: To study the antioxidant chemical constituents and antioxidant activity of Patrinia villosa. Methods: The 70% ethanol-water extract of the herb was separated by silica gel column chromatography, ODS column chromatography and sephadex column chromatography. Then, the compound were further purified and extracted by semi-preparative HPLC. Their structures were elucidated by physiochemical property and spectral analysis. DPPH and ABTS methods were used to determine the antioxidant bioactivities of the isolated compounds. Results: A total of ten compounds were isolated and synthesized, including chlorogenic acid butyl ester (1), 3,4-di-O-caffeoyl quinic acid methyl ester (2), luteolin-7-O-rutinoside (3), 1β-O-β-D-glucopyranosy- 15-O-(p-hydroxylphenylacetate)-5α,6βH-eudesma-3,11(13)-dien-12,6α-olide (4), 3,4-di-O-caffeoyl quinic acid ethyl ester (5), 4,5-di-O-caffeoyl quinic acid methyl ester (6), 4,5-di-O-caffeoyl quinic acid n-butyl ester (7), luteolin-7-O-β-D-glucuronide methyl ester (8), luteolin-7-O-β-D-glucuronide ethyl ester (9), and apigenin-7-O-β-D-glucuronide methyl ester (10). The DPPH radical scavenging IC50 of compounds 3, 8, and 9 were (23.95 ± 0.71), (73.09 ± 0.33), and (25.06 ± 0.65) μmol/L, respectively. The ABTS radical scavenging IC50 was (7.13 ± 0.07), (11.48 ± 0.21), (5.15 ± 0.08) mol/L, respectively. Conclusion: Eight compounds except compounds 3 and 8 are obtained from this species for the first time. Compounds 3, 8, and 9 had significant antioxidant activity.
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OBJECTIVE To investigate the protective effect and mechanisms of luteolin-7-O-β-d-glucuronide (LGU) on oxygen glucose deprivation (OGD)-induced H9C2 cardiomyocytes injury. METH-ODS The protective effect of LGU on OGD-induced H9C2 cardiomyocytes death were investigated by MTT assay. The microfilament change of H9C2 cardiomyocytes was detected by phalloidin staining and the lactate dehydrogenase (LDH) leakage rate was also detected by LDH kit. In order to explore the possible mechanisms of LGU, ATP content, intracellular Ca2+fluorescent intensity and concentra-tion, mitochondrial membrane potential (MMP)and the expressions of apoptosis-related proteins were detected by ATP kit,CLSM(Fluo-3/AM probe),Ca2+kit,CLSM(JC-1 probe)and western blotting meth-od, respectively. RESULTS The inhibition of H9C2 cardiomyocyte survival rate inducedby OGD was improvedby pretreated with LGU in a concentrationdependent manner. The microfilaments injury as well as the increase of LDH leakage rate were also improvedby pretreated with LGU.The ATP content was significantly decreased,intracellular Ca2+fluorescent intensity and concentration were significantly increased and the MMP was significantly decreased 4 hafter OGD. LGU significantly reversed the de-crease of intracellular ATP content,the increase of Ca2+fluorescent intensity and concentration and the decrease of MMP.The release of cytochrome C,the expressionsof caspase-9 and caspase-3 in H9C2 cardiomyocytes were increased 16 h after OGD.LGUsignificantly inhibited the changes of these apop-tosis-related proteins. CONCLUSION LGU has a significant protective effect against OGD-induced H9C2 cardiomyocytes injury through inhibiting calcium overload,increasing ATP content,improving mi-tochondrial function and inhibiting apoptosis.
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OBJECTIVE To investigate the neuroprotective effect and possible mechanisms of lute-olin-7-O-β-D-glucuronide (LGU) against focalcerebral ischemic injury. METHODS The focal cerebral ischemic injury model was established by middle cerebral artery occlusion (MCAO). Male Sprague Dawley rats were randomly divided into sham group,model group(MCAO),LGU group(0.24,0.72 and 2.16 mg·kg-1)and positive control group(Edaravone at 5 mg·kg-1).LGU was injected intravenously 30 min after MCAO.Neurological severity score,infarct volume and brain water content were detected 24 h after MCAO and the levels of Na+-K+ATPase,Ca2+ATPase,TNF-α and IL-1β were detected to explore the possible mechanisms.For the therapeutic time window test,LGU(0.72 mg·kg-1)was injected intrave-nously 0.5, 2, 4, 6, 8, 10 and 12 h respectively after MCAO. To evaluate motion behavior, LGU were injected intravenously 30 min after MCAO and once per day during detection period. The changes of motor coordination were detected by rotating rod method and grip strength analysis, and the changes of gaits were detected using DigiGait Imaging System. RESULTS LGU improved the neurological severity score, infarct volume ratio and brain water content. The therapeutic time window of LGU for cerebral infarction and brain edema was at least 6 h and for neurological dysfunction was 12 h.LGU also prolonged the latency on rotarod, increased the forelimb tension and improved 8 gait parameters, including stance duration,stride length,stance width,paw area,paw area variability,gait symmetry,ataxia coefficient and tau propulsion.Furthermore,LGU increased Na+-K+-ATPase and Ca2+-ATPase levels in the cortex and hippocampus in the ischemic side,reduced the levels of TNF-α and IL-1β in the serum. CONCLUSION LGU has a significant neuroprotective effect against cerebral ischemic injury via improving energy metabolism and reducing inflammation.
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OBJECTIVE: To study the chemical constituents of Inula cappa. METHODS: Chromatographic techniques were employed for isolation and purification of the constituents and their structures were determined by spectral analysis and chemical evidence. RESULTS: Seventeen compounds were obtained and identified as friedelin(1), epifriedelanol(2), α-amyrin(3), β-amyrin(4), benzyl 2-O-β-D-glucopyranosy-2, 6-dihydroxybenzoate(5), scopoletin(6), luteolin-7-O-β-D-glucuronide ethyl ester(7), benzyl alcohol glucoside(8), ophiopogonoside A(9), apigenin-7-O-β-D-glucoside(10), luteolin-7-O-β-D-rutinoside(11), hydnocarpin-D(12), luteolin(13), luteolin-7-O-β-D-glucoside (14), luteolin-4'-O-β-D-glucoside (15), quercetin-3-O-β-D-glucoside (16), and juglans cerebroside A(17). CONCLUSION: Compounds 5, 7, 9, 11, 12, and 17 are for the first time obtained from the genus Inula and compounds 8, 10, 14, and 16 are isolated from Inula cappa for the first time.
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Objective: To investigate the technology for the separation and purification of extract in Dracocephalum moldevica (EDM) by macroporous resin. Methods: Static and dynamic adsorption-desorption were used to select the best one from seven different type macroporous resins; With the content of total flavonoids, tilianin, luteolin-7-O-β-D-glucuronide, and rosmarinic acid as indexes, the purification technology parameters of EDM were optimized. Results: HPD600 resin showed the best purifying profile, its optimum technology conditions were as follows: The optimum concentration of the sample liquid was 0.08 g/mL equivalent to raw material, the resin column diameter-height ratio was 1:9, the amount of used adsorption was 0.32 g dried medicinal herb/mL resin, sample flow rate was 1.5 BV/h, and adsorption time was 12 h. In the course of elution, the resin column chromatography was eluted with 6 BV of 70% ethanol after removing impurities with 4 BV of water by flow rate of 1.5 BV/h. The contents of total flavonoids, tilianin, luteolin-7-O-β-D-glucuronide, and rosmarinic acid were more than 53%, 5.5%, 4.7%, and 2.5%. Conclusion: Macroporous resin HPD600 is suitable to separate and purify EDM.
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OBJECTIVE: To establish an HPLC method for simultaneous detemination the contents of rosmarinic acid (phenylpropanoids), luteolin-7-O-β-D-glucuronide and tilianin (flavonoids) in Dracocephalum moldavica L. Extract. METHODS: The samples were separated on Purospher® STARLP RP-18 endcapped column(4.6 mm × 250 mm, 5 μm) by gradient elution with acetonitrile (A)-0.5% formic acid solution(B) (0-28 min, 20% A; 28-55 min, 20%→28% A; 55-80 min, 28%→30% A)as the mobile phase at a flow ratio of 1.0 mL · min-1. The detection wavelength was set at 330 nm and the column temperature was maintained at 35℃. RESULTS: The calibration curves of rosmarinic acid, luteolin-7-O-β-D-glucuronide, tilianin were in good linearity over the ranges of 2.184-21.84 μg · mL-1 (r =0.999 6), 3.14-31.84 μg · mL-1(r =0.999 6), 7.9-39.5 μg · mL-1(r =0.999 5) respectively. The average recoveries were 99.62%, 99.64% and 100.12% with RSD values of 1.27%, 1.05% and 1.09%. CONCLUSION: The method is reliable, simple, and accurate, and can be used for the comprehensive quality control of Dracocephalum moldavica L. extract.