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Metabolomics based on liquid chromatography coupled with mass spectrometry (LC-MS) was used to study the initiation and development of diabetes in rats, and the ability of Ginkgo biloba extract (GBE) to ameliorate this pathology. Diabetes mellitus (DM) was induced by intra-peritoneal injection of streptozotocin. The rats were randomly divided into a normal control group treated with drug-free solution (NC), a normal control group treated with GBE (N-GBE), a DM group treated with drug-free solution (DM), and a DM group treated with GBE (D-GBE); rats were maintained on this protocol for 9 weeks. Rat plasma was collected from the sixth week to the ninth week and then analyzed with LC-MS. Animal experimentation was approved by the Committee on the Ethics of Animal Experiments of Xuzhou Medical University. Twelve plasma metabolites with continuous differentiation were monitored to indicate dysfunction of metabolic pathways including fatty acid metabolism, phospholipid metabolism, amino acid metabolism, tricarboxylic acid cycle activity, bile acid metabolism, and purine metabolism to confirm the occurrence and development of DM. Treatment with GBE partially reversed the changes seen in five metabolites in DM rats, indicating that GBE could prevent the occurrence and development of DM by acting on fatty acid metabolism, phospholipid metabolism, amino acid metabolism, and the tricarboxylic acid cycle.
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Abstract Ginkgo biloba is a traditional Chinese herbal medicine containing multiple components that contribute to its notable bioactivities. Variations of seasonal, meteorological and planting soil on the phytochemicals contents in G. biloba leaves due to the effects of growth meteorological and soil parameters were investigated in this study. The leaves of G. biloba were collected from different months and place in Zhejiang province, the contents of flavones (quercetin, kaempferol and isorhamnetin) and terpene lactones (bilobalide, ginkgolides A, B and C) were quantified by high performance liquid chromatography (HPLC) and the evaporative light scattering detector (ELSD) method. The established methods were validated with good linearity, precision, repeatability, stability, and recovery. Comprehensive analysis suggested the proper harvest time for G. biloba was in October of Zhejiang province. The result of correlation analysis with meteorological factors shows that the temperature and precipitation have non-significant effect on the main components of G. biloba. In addition, the type and content (Mn and Zn) of the soil showed significantly effect on the content of flavonoids and terpene lactones. This study enriched the knowledge on the development and utilization value of the G. biloba leaves and was useful for determining the optimal harvest time and growing condition.
Resumo Ginkgo biloba é um fitoterápico tradicional da medicina chinesa que contém vários componentes que contribuem para suas notáveis bioatividades. Variações sazonais, meteorológicos e de plantio do solo sobre os teores fitoquímicos em folhas de G. biloba, devido aos efeitos do crescimento meteorológico e parâmetros do solo, foram investigadas neste estudo. As folhas de G. biloba foram coletadas em diferentes meses e na província de Zhejiang, os teores de flavonas (quercetina, kaempferol e isorhamnetina) e lactonas terpênicas (bilobalida, ginkgolídeos A, B e C) foram quantificados por Cromatografia Líquida de Alta Eficiência (CLAE) e pelo método do detector de dispersão da luz por evaporação. Os métodos estabelecidos foram validados com boa linearidade, precisão, repetibilidade, estabilidade e recuperação. Uma análise abrangente sugeriu que o tempo de colheita adequado para G. biloba foi em outubro na província de Zhejiang. O resultado da análise de correlação com fatores meteorológicos mostra que a temperatura e a precipitação não têm efeito significativo sobre os principais componentes de G. biloba. Além disso, o tipo e composição (Mn e Zn) do solo apresentaram efeito significativo sobre o teor de flavonoides e lactonas terpênicas. Este estudo enriqueceu o conhecimento sobre o valor de desenvolvimento e utilização das folhas de G. biloba e foi útil para determinar o melhor tempo de colheita e condição de crescimento.
Subject(s)
Soil , Ginkgo biloba , Seasons , Plant Extracts , Chromatography, High Pressure Liquid , Plant LeavesABSTRACT
Objective: To establish an ultra-high performance liquid chromatography coupled with triple quadrupole mass for the determination of five biflavones. Method: Chromatographic separation was carried out on a ACQUITY UPLC BEH C18 column (2.1 mm×100 mm,1.7 μm) with gradient elution of acetonitrile and 0.10%formic acid at a flow rate of 0.3 mL·min-1,and the column temperature was set at 35℃. Result: Amentoflavone,bilobetin,ginkgetin,isoginkgetin,sciadopitysin showed a good linearity within the ranges of 0.02-13.20 mg·L-1(r=0.996 3),0.05-23.60 mg·L-1(r=0.995 5),0.09-18.60 mg·L-1(r=0.992 7),0.10-21.00 mg·L-1(r=0.998 8),0.06-16.00 mg·L-1(r=0.996 7),with average recoveries of 101.50%,98.78%,97.59%,97.24%,101.09%, and RSDs of 2.7%,2.7%,3.1%,2.8%,1.3%. The contents of amentoflavone,bilobetin,ginkgetin,isoginkgetin,sciadopitysin ranged between 121.30-434.74,268.39-847.14,251.80-1 297.10,195.87-691.10,477.48-3 003.90 μg·g-1. The total biflavones ranged between 1 474.45-5 635.40 μg·g-1. It shows a certain regularity that the low-vinegar leaves contain higher total flavonoids,and the total flavonoid content gradually decreases with the increase of tree age. Conclusion: The method was simple, and can be used for qualitative and quantitative analysis of biflavones.
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Abstract Ginkgo biloba is a traditional Chinese herbal medicine containing multiple components that contribute to its notable bioactivities. Variations of seasonal, meteorological and planting soil on the phytochemicals contents in G. biloba leaves due to the effects of growth meteorological and soil parameters were investigated in this study. The leaves of G. biloba were collected from different months and place in Zhejiang province, the contents of flavones (quercetin, kaempferol and isorhamnetin) and terpene lactones (bilobalide, ginkgolides A, B and C) were quantified by high performance liquid chromatography (HPLC) and the evaporative light scattering detector (ELSD) method. The established methods were validated with good linearity, precision, repeatability, stability, and recovery. Comprehensive analysis suggested the proper harvest time for G. biloba was in October of Zhejiang province. The result of correlation analysis with meteorological factors shows that the temperature and precipitation have non-significant effect on the main components of G. biloba. In addition, the type and content (Mn and Zn) of the soil showed significantly effect on the content of flavonoids and terpene lactones. This study enriched the knowledge on the development and utilization value of the G. biloba leaves and was useful for determining the optimal harvest time and growing condition.
Resumo Ginkgo biloba é um fitoterápico tradicional da medicina chinesa que contém vários componentes que contribuem para suas notáveis bioatividades. Variações sazonais, meteorológicos e de plantio do solo sobre os teores fitoquímicos em folhas de G. biloba, devido aos efeitos do crescimento meteorológico e parâmetros do solo, foram investigadas neste estudo. As folhas de G. biloba foram coletadas em diferentes meses e na província de Zhejiang, os teores de flavonas (quercetina, kaempferol e isorhamnetina) e lactonas terpênicas (bilobalida, ginkgolídeos A, B e C) foram quantificados por Cromatografia Líquida de Alta Eficiência (CLAE) e pelo método do detector de dispersão da luz por evaporação. Os métodos estabelecidos foram validados com boa linearidade, precisão, repetibilidade, estabilidade e recuperação. Uma análise abrangente sugeriu que o tempo de colheita adequado para G. biloba foi em outubro na província de Zhejiang. O resultado da análise de correlação com fatores meteorológicos mostra que a temperatura e a precipitação não têm efeito significativo sobre os principais componentes de G. biloba. Além disso, o tipo e composição (Mn e Zn) do solo apresentaram efeito significativo sobre o teor de flavonoides e lactonas terpênicas. Este estudo enriqueceu o conhecimento sobre o valor de desenvolvimento e utilização das folhas de G. biloba e foi útil para determinar o melhor tempo de colheita e condição de crescimento.
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To study Ginkgo biloba leaves in different producing area, we establish an HPLC method for the simultaneously determination of seven flavonoids glycosides and four biflavonoids in G. biloba leaves. The analysis was performed on an Agilent ZORBAX SB-C₁₈ column(4.6 mm×250 mm, 5 μm) wich acetonitrile, and 0.4% phosphoric acid as mobile phase at flow rate of 1 mL•min⁻¹ in a gradient edution, and the detection was carried out at 254 nm.The calibration curves of the seven flavonoids glycosides and four biflavonoids had a good linearitiy with good recoveries. The established HPLC method is simple, rapid, accurate, reliable, and sensitive, and can be applied to the identification and quality control of G. biloba leaves.
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Objective To investigate the effects of extract of Ginkgo biloba leaves EGb761 on 1-methy-l 4-phenylpyridium (MPP+)-induced injury in human neuroblastoma SH-SY5Y cells; To discuss its mechanism of action.Methods Cell culture method was used and SH-SY5Y cell damage model was induced with different concentrations of MPP+ to build Parkinson’s disease model in vitro. The experiment was divided into control group, MPP+ model group, low-, medium-, and high-dose EGb761 groups. The survival rate was determined by MTT assay, and the apoptotic rate was detected by flow cytometry according to AnnexinV apoptosis detection kit. The cell morphology was observed by inverted microscope. NO content in SH-SY5Y cells was detected by Nitric acid reduction method.Results Compared with the control group, the survival rate of SH-SY5Y cells decreased and the apoptotic rate and NO content increased in the model group (P<0.05); Compared with the model group, the survival rate of SH-SY5Y cells increased and the apoptotic rate and NO content decreased in the low-, medium- and high-dose EGb761 groups (P<0.05).Conclusion EGb761 can protect MPP+-induced SH-SY5Y cell from damage by the inhibition of the content of NO free radical.
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To compare the quality Ginkgo biloba L. leaves collected form various habitats, different tree ages, and different harvesting times and to confirm the source of G. biloba leave used in Xin Mai Capsule. To establish the HPLC fingerprint of G. biloba leaves to determine the fingerprint and content of total flavonol glucoside and terpene lactones in G. biloba leaves collected from various habitats, different tree ages, and different harvesting time, and then to analyze the results. The contents of total flavonol glucoside and terpene lactones and fingerprint similarity in G. biloba leaves from various regions, different tree ages, and collecting times were different. The fingerprint control method has provided the scientific basis for the reasonable collection and quality evaluation of G. biloba leaves. The G. biloba leaves in June to August, 2-3 year-old from Pizhou, Jiangsu province are better, which could be considered as the source of G. biloba leaves used in Xin Mai Capsule.
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Objective To explore the antibacterial activity of extract of Ginkgo biloba leaves on Porphyromonas gingivalis in vitro ,and to provid pharmacological reference for developing a new type of antibacterial drugs in the treatment of periodontal disease.Methods This experiment was divided into negative control group,imipenem control group and different concentrations and forms of extract of Ginkgo biloba leaves groups.Solvent extraction method was used to extract the extract of Ginkgo biloba leaves, punching method and test tube method were performed to detect the antibacterial activity of extract of Ginkgo biloba leaves in anaerobic environment invitro and compared with Staphylococcusaureus and E.coli.By observing the antibacterial ring diameter and determination of the minimum bacteriostasis concentration (MIC),the antibacterial activities of extract of Ginkgo biloba leaves in vitro were measured.Results In the experiment of bacteriostatic ring,Porphyromonas gingivalis was treated with extract of Ginkgo biloba leaves,Ginkgo biloba leaf tablet and Ginkgo biloba soft capsule concentrate and 1∶4 diluent,the bacteriostatic ring diameters were decreased with the decreasing of the concentration.The maximum bacteriostatic diameter of Ginkgo biloba extract was 1 6.5 mm,and the maximum bacteriostasis diameters of Ginkgo biloba leaf tablet and soft capsule were 15.3 and 14.5 mm,respectively;the bacteriostatic diameter of the exact of Ginkgo biloba leaves was bigger than those of Ginkgo biloba leaf tablet and Ginkgo biloba soft capsule (P 0.05);E.coli and Staphylococcusaureus groups get the same results.When the concentration of extract of Ginkgo biloba leaves was more than 1.95 mg·L-1 ,there was no growth of Porphyromonas gingivalis but E. coli and Staphylococcus aureusa still grew;only the concentrations of exact of Girkgo biloba leaves were more than 6.25 and 12.5 mg· L-1 ,E. coli and Staphylococcus aureus didn’t grow;the bacteriostatic effect of extract of Ginkgo biloba on Porphromonas gingivalis was better than E.coli and Staphylococcus aureus . Conclusion Extract of Ginkgo biloba leaves has antibacterial effect on Porphyromonas gingivalis.
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Objective: To purify the biflavonoids from Ginkgo biloba leaves and determine their contents in the leaves collected in different periods. Methods: Dichloromethane fraction of ethanol extract was used to purify biflavonoids through silica gel column and semi-preparative HPLC. The contents of four biflavonoids were simultaneously determined using RP-HPLC. Results: According to the reported 1H-NMR data, four compounds were identified as bilobetin (LB-1), ginkgetin (LB-2), isoginkgetin (LB-3), and sciadopitysin (LB-4). Content determination demonstrated that the biflavonoids accumulation decreased as the leaves collected from the beginning to the end of September, and the content difference can be as large as 30%. Conclusion: RP-HPLC method can be used in the quantitative determination of biflavonoids in G. biloba leaves. The semi-preparative condition can be used in the purification of biflavonoids from G. biloba leaves. The contents of four biflavonoids could be decreased significantly when the leaves turned yellow, which provides the useful information for the collection of G. biloba leaves. However, the mechanism and the accumulation of other kinds of compounds need further study.
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OBJECTIVE: To establish a UPLC fingerprint of Ginkgo biloba leaves and determine the contents of eleven flavonoid glycosides. METHODS: A UPLC method was established for the first time to simultaneously quantify the eleven major flavonoid glycosides in Ginkgo biloba leaves. The analysis was performed on an Agilent ZORBAX Eclipse Plus C18 column (4.6 mm × 50 mm, 1.8 μm), the mobile phase A was acetonitrile, the mobile phase B was 0.4% phosphoric acid, gradient elution was performed at the flow rate of 0.6 mL · min-1, and the detection was carried out at 360 nm. Similarity evaluation system for chromatographic fingerprint of traditional Chinese medicine(2004 A) was used in data analysis. RESULTS: The UPLC fingerprint of Ginkgo biloba leaves was established. Total 21 peaks were selected as the characteristic common peaks and 11 of them were identified. There were some differences in the fingerprint chromatograms between 32 batches of Ginkgo biloba leaf samples. The validation result showed that the 11 flavonoid glycosides had good linearity and recoveries. CONCLUSION: The established UPLC method is simple, rapid, accurate, reliable, and sensitive, and can be applied to the identification and quality control of Ginkgo biloba leaves.
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Objective: To study the chemical constituents of flavonoid glycosides in the extract from Ginkgo biloba leaves used for injection. Methods: The constituents of flavonoid glycosides in the 70% ethanol extract from G. biloba leaves were isolated and purified by chromatography over silica gel, MCI, ODS, Sephadex LH-20 columns and RP-HPLC. The structures were identified on the basis of physicochemical properties and spectral data analyses. Results: Sixteen compounds were isolated and identified as 5, 7-dihydroxyl-4'-methoxylflavonol-3-O-rutinoside (1), quercetin-3-O-(2″, 6″-α-L-dirhamnopyranosyl)-β-D-glucoside (2), quercetin- 3-O-[2″-(6‴-p-coumaroyl)-β-D-glucopyranosyl] -α-L-rhamnopyranosyl-7-O-β-D-glucoside (3), isorhamnetin-3-O-(2″, 6″-α-L-dirhamnopyranosyl)-β-D-glucoside (4), rutin (5), luteolin-7-O-β-D-glucoside (6), quercetin-3-O-β-D-glucoside (7), quercetin- 3-O-(2″-β-D-glucopyranosyl)-α-L-rhamnoside (8), kaempferol-3-O-rutinoside (9), quercetin-3-O-α-L-rhamnoside (10), isorhamnetin-3-O-rutinoside (11), apigenin-7-O-β-D-glucoside (12), syringetin-3-O-rutinoside (13), kaempferol-3-O-(2″- β-D-glucopyranosyl)-α-L-rhamnoside (14), quercetin-3-O-α-L-rhamnopyranosyl-2″- (6‴-p-coumaroyl)-β-D-glucoside (15), and kaempferol-3-O-α-L-rhamnopyranosyl-2″- (6‴-p-coumaroyl)-β-D-glucoside (16). Conclusion: UPLC-PDA shows that all the compounds are flavonoid glycosides in the leaves of G. biloba and are marked in the extract and injection. Compound 1 is isolated from this plant for the first time, named flavonol glycoside K.
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AimTo study the effect of extracts of ginkgo biloba leaves(GbE) on apoptosis in neuronal cells. Methods Primary cerebral cultures from rat fetus were used to observe the activity of neuronal cells and to determine the release of LDH and DNA gel electrophoresis. Results Gb E enhanced the activity of neuronal cells, decresed the release of LDH,and relieved the structural changes of nucleus and DNA fragmentation. Conclusion GbE can inhibit apoptosis in neuronal cells.
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Aim To study the effect of extracts of ginkgo biloba leaves(GbE) on apoptosis in neuronal cells. Methods Primary cerebral cultures from rat fetus were used to observe the activity of neuronal cells and to determine the release of LDH and DNA gel electrophoresis.Results GbE enhanced the activity of neuronal cells, decresed the release of LDH,and relieved the structural changes of nucleus and DNA fragmentation. Conclusion GbE can inhibit apoptosis in neuronal cells.
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Aim To observe the effect of extracts of ginkgo biloba leaves(GbE) on platelet adhesion and thrombosis. Methods The platelet adhesion was assayed by rotating glass-globe method. The thrombogenesis method in artery-vein bypass was applied to observe thrombosis in vivo. The Chandler method was applied to induce thrombosis in vitro.The tail bleeding time was recorded by shearing tail method.Results GbE significantly decreased platelet adherence rate in rabbits, decreased the weight of thrombus produced in artery-vein bypass in rabbits, shortened the length and reduced dry weight of thrombus of rats in vitro,prolonged the tail bleeding time in mice.Conclusion GbE inhibits the platelet adhesion and thrombosis.
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Objective To investigate the protective effects of Ginkgo Biloba leaves(EGb) preconditioning on liver graft in rat liver transplantation.Methods Male Sprague-Dawley rats were used as donors and(recipients) of orthotopic liver transplantation(OLT).The rats were randomly divided into EGb group,normal saline(NS) control group and sham operation(SO) group.The animals were killed at 2h,6h,24h after graft reperfusion.Plasma samples were collected for ALT and AST test.Liver tissues were collected to detect the expression of TNF-?mRNA and Bcl-2mRNA by RT-PCR.Also,liver tissues were used to detect rat(liver) histological change and apoptosis by TUNEL.Results The serum levels of ALT in EGb group were(significantly) lower than the NS group(P
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Objective:To establish a method for analysis of residual organic solvents (methanol, ethanol, acetone, ethyl acetate) in Ginkgo biloba Leaves for injection.Methods: The contents of residual solvents in Ginkgo biloba Leaves for injection were determined by headspace GC on HP 5 column, with FID detector, high purity nitrogen as the carries gas.Results: There was a good linearity ( r :0.9909～0.9999). The RSD of precision and accuracy was less than 5%, the average recovery rate was in the range of 93.06%～113.78%.Conclusions: The method was simple, quick and accurate and can be used for quality control of Chinese patent medicine.
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Objective: To study the radioprotective effect of flavonoids from Ginkgo biloba leaves(GBF). Methods: Three water extracts of GBF were prepared (low dosage 10 mg/100 ml, medium dosage 20 mg/100 ml and high dosage 100 mg/100 ml) and orally administered to mice . After 10 d, the mice were exposed to 8.5Gy -rays. After another 10 d of oral administration, the survival rates were recorded in 30 d. In another experiment, six groups of mice (three GBF groups, radiation control, normal control and cyclophosphamide group) were arranged. The first three groups were orally administered with low, medium and high dosage of GBF respectively for 11d; the other three groups with distilled water. Then the three GBF groups and radiation group were exposed to 1.0Gy -rays. Then they were orally administered again in the following 7d . Micronucleated polychromatic erythrocytes in bone-marrow and sperms (AFS) in mice were observed on the 21st day after termination of oral administration. Proliferation rates of lymphocyte (PRL) were determined in the three GBF groups and normal control. Results: Low, medium and high dosage of GBF increased the survival rates by 31.7%, 25.3% and 26.5% respectively(P