Inhibitory effects of Ginkgo biloba extract on renal inflammation in diabetic nephropathy model mice and its mechanism / 中国药房

OBJECTIVE To investigate the inhibitory effects of Ginkgo biloba extract （GBE） on renal inflammation in diabetic nephropathy （DN） model mice， and its potential mechanism. METHODS KK/Ay mice were fed with high fat and high sugar to induce DN model. They were divided into model group， positive control group [metformin 200 mg/（kg·d）]， GBE low-dose and high-dose groups [100， 200 mg/（kg·d）]， with 6 mice in each group. Six C57BL/6J mice were fed with a regular diet as the control group. Administration groups were given relevant liquid intragastrically， control group and model group were given constant volume of normal saline intragastrically， once a day， for 8 consecutive weeks. The body weight， fasting blood glucose， 24-hour food intake， 24-hour urine output， monocyte chemoattractant protein-1 （MCP-1）， interleukin-12 （IL-12）， IL-10， advanced glycation end products （AGEs）， blood urea nitrogen （BUN） and serum creatinine （Scr） of mice were measured， and the ratio of bilateral kidneys to body weight was also calculated. The pathological injury and fibrotic changes of the renal cortex were observed， and the expressions of macrophage polarization marker proteins [type M1： inducible nitric oxide synthase （iNOS）； type M2： arginase-1 （Arg-1）] and AGEs-the receptor of advanced glycation end products （RAGE）/Ras homolog gene pharm_chenjing@163.com family member A （RhoA）/Rho-associated coiled-coil forming protein kinase （ROCK） signaling pathway-related proteins were determined in renal cortex. RESULTS Compared with the model group， the symptoms such as renal cortical hyperplasia， vacuoles， infiltration of inflammatory cells， and renal cortical fibrosis had been improved in GBE low-dose and high-dose groups； body weight， serum level of IL-10， the expression of Arg-1 in the renal cortex were significantly higher than model group （P＜ 0.01）； fasting blood glucose， 24-hour food intake， 24-hour urine output， serum levels of MCP-1， IL-12， BUN， Scr and AGEs， the ratio of bilateral kidneys to body weight， renal injury score， the proportion of renal interstitial fibrosis， the protein expressions of iNOS， RAGE， RhoA and ROCK1 （except for GBE low-dose group） in renal cortex were significantly lower than model group （P＜0.01）. CONCLUSIONS GBE could improve kidney damage and alleviate inflammatory response in DN model mice， the mechanism of which may be related to inhibiting the AGEs-RAGE/RhoA/ROCK signaling pathway and regulating macrophage polarization.

Data-driven engineering framework with AI algorithm of Ginkgo Folium tablets manufacturing

Smart manufacturing still remains critical challenges for pharmaceutical manufacturing. Here, an original data-driven engineering framework was proposed to tackle the challenges. Firstly, from sporadic indicators to five kinds of systematic quality characteristics, nearly 2,000,000 real-world data points were successively characterized from Ginkgo Folium tablet manufacturing. Then, from simplex to the multivariate system, the digital process capability diagnosis strategy was proposed by multivariate Cpk integrated Bootstrap-t. The Cpk of Ginkgo Folium extracts, granules, and tablets were discovered, which was 0.59, 0.42, and 0.78, respectively, indicating a relatively weak process capability, especially in granulating. Furthermore, the quality traceability was discovered from unit to end-to-end analysis, which decreased from 2.17 to 1.73. This further proved that attention should be paid to granulating to improve the quality characteristic. In conclusion, this paper provided a data-driven engineering strategy empowering industrial innovation to face the challenge of smart pharmaceutical manufacturing.

Permitted daily exposure from preclinical studies of Ginkgo biloba L. dry extract

Abstract Resolution 658/2022 of the Brazilian Regulatory Agency requires the determination of the permitted daily exposure (PDE) of pharmaceutical agents. Ginkgo biloba L. is used therapeutically to treat memory deficits and other brain diseases. However, published results indicate that more studies are needed to confirm the safety of Ginkgo biloba. This study aimed to evaluate the dry extract of Ginkgo biloba L. leaves PDE as an ingredient in an oral pharmaceutical product in preclinical studies using mice. Acute oral toxicity and repeated dose experiments were performed based on OECD guidelines, as well as genotoxicity tests. The results indicate that Ginkgo biloba L. has low acute toxicity, no liver toxicity, and does not alter blood glucose levels. No changes in weight gain were observed, but food intake decreased in males during the first week of treatment at the highest dose. Hematological parameters were not altered in males, whereas females presented lower leukocyte and lymphocyte counts and higher neutrophil counts at the highest dose. The lipid profile was not altered in males, whereas total cholesterol was increased in females. The estimated PDE was 0.1 mg/day and, when related to the maximum residual concentration, indicates that the cleaning process used is safe and does not require reassessment.

Changes of RAGE/p38MAPK/NF-κB signaling pathway in brain tissue of subchronic fluorosis rats and protective effects of Ginkgo biloba extract and RAGE antagonist / 环境与职业医学

Background Fluorine accumulates in the brain tissue after long-term excessive intake and subsequently cause nerve damage and decline of learning and memory ability. Receptor of advanced glycation end-products (RAGE)/p38 mitogen-activated protein kinase (p38MAPK)/nuclear factor kappa-B (NF-κB) signaling pathway is considered to be involved in the associated mechanism. Objective To study the changes of RAGE/ p38MAPK/ NF-κB signaling pathway in rats with subchronic fluorosis, and to explore the protective effects of extract of Ginkgo biloba 761 (EGb761) and RAGE antagonist (FPS-ZM1) on neuromemory ability. Methods Ninety male clean SD rats were divided into 9 groups with 10 rats in each group. The modeling period was 6 months. Control group (C group): free drinking tap water (fluoride content <0.5 mg·L−1), low- and high-dose fluoride groups (LF group, HF group): free drinking tap water with 10 or 50 mg·L−1 fluoride; intervention group of Ginkgo biloba extract (CE, LFE, and HFE groups): on the basis of the C group, LF group, and HF group, 100 mg·kg−1·d−1 EGb761 was given daily via intragastric administration; FPS-ZM1 intervention groups (CF, LFF, and HFF groups): 7 d before the end of modeling, 1 mg·kg−1·d−1 FPS-ZM1 was injected intraperitoneally daily on the basis of the C group, LF group, and HF group. The contents of fluoride in brain and blood of each group were detected. The learning and memory ability was tested by water maze experiment. The histopathologic changes of the hippocampus were detected by Nissl staining. The protein expression levels of RAGE and its ligand high mobility group protein B1 (HMGB1), NF-κB, p38MAPK, phospho-p38MAPK (p-p38MAPK), interleukin-6 (IL-6), and tumour necrosis factor-α (TNF-α) in brain tissue were detected by Western blotting. The mRNA expression levels of RAGE, HMGB1, and p38MAPK were detected by quantitative real-time PCR. Results Compared with the C group, the contents of blood fluoride and brain fluoride in the LF and the HF groups were increased (P<0.05). The results of the water maze experiment showed that, compared with the C group, the escape latency time of the LF group and the HF group was longer and the crossing times were reduced; compared with the HF group, the escape latency time of the HFE group and the HFF group was shortened, and the crossing times were increased (P<0.05). The Nissl staining results showed that the number of Nissl body in the HF group decreased compared with the C group; compared with the HF group, the number of Nissl body in the HFE group and the HFF group increased. The Western blotting results showed that compared with the relative protein expression levels of RAGE, HMGB1, NF-κB, p38MAPK, p-p38MAPK, IL-6, and TNF-α in the C group , the levels of above indicators in the HF group and the levels of RAGE, HMGB1, NF-κB, p-p38MAPK, and IL-6 in the LF group were up-regulated (P<0.05); compared with the HF group, the levels of above indicators in the HFE group and the HFF group were all down-regulated (P<0.05); compared with the relative protein expression levels of RAGE and HMGB1 in the LF group, the levels in the LFE group and the LFF group were all down-regulated (P<0.05). The quantitative real-time PCR results showed that compared with the C group, the mRNA expression levels of RAGE and HMGB1 in the LF group and the HF group were up-regulated; compared with the LF group, the mRNA expression levels of RAGE in the LFE group and the LFF group were down-regulated ; compared with the HF group, the mRNA expression levels of RAGE and HMGB1 in the HFE group and the HFF group were down-regulated (P<0.05). Conclusion The central nervous system injury caused by subchronic fluorosis may be related to the activation of RAGE/p38-MAPK/NF-κB signaling pathway, which can impair the learning and memory ability of rats, while EGb761 and FPS-ZM1 may have certain protective effects on the nerve injury.

A simulating method for dripping process of Ginkgo biloba leaf dripping pills based on computational fluid dynamics technology / 药学学报

A simulating method for dripping process of Ginkgo biloba leaf dripping pills based on computational fluid dynamics was constructed. Ginkgo biloba leaf dripping pills was explored as the experimental subject to simulate the dripping process based on FLOW-3D software. The dripping process was simulated through the derivation of the governing equations, the selection of the models, and simulation parameters. Firstly, the droplet morphologies and drop speeds under different liquid viscosity were simulated. It was found that with the increase of the liquid viscosity, the drop speed decreased and the difficulty of droplet preparation gradually increased. The simulation results were consistent with the experiment results. Secondly, the droplet morphologies at different drop speeds were investigated and verified by experiments. It was found that the simulation results had a good correlation with the experiment results. The results shown that the viscosity of the liquid was the critical material attribute, and the drop speed was the critical process parameter, according to the droplet morphology. The establishment of the simulation method can deepen the understanding of the dripping process and provide a reference for the selection of raw materials and process parameters.

Effects of Ginkgo biloba on arsenic-induced lung injury in rats based on HMGB1/RAGE pathway / 中华地方病学杂志

Objective:To investigate the antagonistic and therapeutic effects of Ginkgo biloba on arsenic-induced lung injury in rats and its mechanism.Methods:A total of 42 healthy clean grade Wistar rats, half male and half female, weighing 120 - 130 g, were randomly divided into 7 groups with 6 rats in each group. Two intervention models of Ginkgo biloba antagonism and treatment were established, respectively. The specific treatments were as follows: (1) Experimental study on the antagonism of Ginkgo biloba (4 groups): the control A group was given deionized water; the Ginkgo biloba control (GBE) group was given Ginkgo biloba solution (50 mg·kg -1·bw); the arsenic-treated (As) group was given sodium arsenite solution (10 mg·kg -1·bw); the Ginkgo biloba antagonistic (As + GBE) group was treated with sodium arsenite solution (10 mg·kg -1·bw) and Ginkgo biloba solution (50 mg·kg -1·bw), and the above administration was by gavage for 6 days/week, for 4 months. (2) Experimental study on the treatment of Ginkgo biloba (3 groups): the control B group was given deionized water for 5.5 months; in the arsenism natural recovery (recovery) group, sodium arsenite solution (10 mg·kg -1·bw) was administered by gavage for 4.0 months and deionized water for 1.5 months; the Ginkgo biloba treatment (treatment) group was given sodium arsenite solution (10 mg·kg -1·bw) by gavage for 4.0 months and Ginkgo biloba solution (50 mg·kg -1·bw) for 1.5 months, and the above administration was for 6 days/week. Masson staining was used to evaluate collagen fiber deposition in lung tissue. Western blotting was used to detect the expression level of related proteins in lung tissue homogenates, including inflammatory cytokines matrix metalloproteinase-9 (MMP-9), interleukin (IL)-1β, IL-18; high mobility group box 1 (HMGB1) and receptor for advanced glycation end-products (RAGE) of the HMGB1/RAGE pathway; phosphatidylinositol-4, 5-bisphosphate 3-kinase (PI3K), protein kinase B (AKT), phosphorylated AKT (p-AKT) of the PI3K/AKT pathway; transforming growth factor (TGF)-β1, SMAD2, p-SMAD2, SMAD3, p-SMAD3 and SMAD4 of the TGF-β1/SMAD pathway. Results:(1) Antagonistic effect of Ginkgo biloba: compared with the control A group, there was no significant change in protein expression and collagen fiber deposition in the lung tissue of GBE group ( P > 0.05); the levels of MMP-9, IL-1β and IL-18 protein expression and collagen fiber deposition in the lung tissue of As group were significantly increased ( P < 0.05); and the levels of HMGB1, RAGE, PI3K, p-AKT, TGF-β1, p-SMAD2, p-SMAD3 and SMAD4 protein expression were significantly increased ( P < 0.05). Compared with As group, the levels of MMP-9, IL-1β and IL-18 protein expression and collagen fiber deposition were significantly decreased in As + GBE group ( P < 0.05); and levels of HMGB1, RAGE, PI3K, p-AKT, TGF-β1, p-SMAD2, and p-SMAD3 protein expression were significantly decreased ( P < 0.05). (2) Therapeutic effect of Ginkgo biloba: compared with control B group, the levels of MMP-9, IL-1β, IL-18 protein expression and collagen fiber deposition were significantly increased in recovery group ( P < 0.05); and the levels of HMGB1, RAGE, PI3K, p-AKT, TGF-β1, p-SMAD2, p-SMAD3 and SMAD4 protein expression were significantly increased ( P < 0.05). Compared with recovery group, the levels of MMP-9, IL-1β, IL-18, HMGB1, RAGE, PI3K and p-AKT protein expression were significantly decreased in treatment group ( P < 0.05); and there was no significant change in collagen fiber deposition and TGF-β1, p-SMAD2, p-SMAD3 and SMAD4 protein expression levels in lung tissue ( P > 0.05). In both experiments, there was no significant difference in the protein expression levels of AKT, SMAD2 and SMAD3 between the groups ( P > 0.05). Conclusion:Ginkgo biloba intervention has ameliorated inflammatory injury and collagen fiber deposition in lung tissue of arsenic-treated rats possibly by inhibiting the expression levels of HMGB1/RAGE pathway-related proteins.

Network pharmacology analysis of ginkgo leaves extract against diabetic retinopathy / 中华实验眼科杂志

Objective:To explore the key targets of ginkgo leaves extract against diabetic retinopathy (DR), and the underlying pharmacological mechanism by network pharmacology.Methods:Potential targets of active components in ginkgo folium were searched from the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCMSP) and Swiss Target Prediction database.Therapeutic targets highly related to DR were retrieved from the GeneCards and DisGENET databases.The intersection targets were subjected to Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis to construct the active components-targets network.Based on the degree of network nodes, key active components and key targets were obtained for molecular docking verification.Results:A total of 27 active components and 34 potential therapeutic targets of ginkgo leaves extract in the treatment of DR were screened out.GO enrichment analysis indicated that these therapeutic targets were mainly enriched in the processes of inflammation, oxidative stress, angiogenesis and hypoxic damage.KEGG pathway enrichment analysis revealed that the advanced glycation end product-receptor for advanced glycation end product, mitogen active protein kinase, phosphatidylinositol-3-kinase-protein kinase B, hypoxia inducible factor-1, tumor necrosis factor (TNF) and interleukin (IL)-17 signaling pathways were mainly enriched.According to degree value calculated from the active components-targets network, the top 4 key active components with higher degrees were quercetin, kaempferol, luteolin and isorhamnetin, and the top 8 key targets were serine/threonine protein kinase 1, vascular endothelial growth factor A, IL-6, TNF, nitric oxide synthase 3, peroxisome proliferator activated receptor-γ, IL-10 and matrix metalloproteinase-9.The binding activity between key active components and key targets was good.Conclusions:A variety of active compounds contained in ginkgo leaves extract can target various therapeutic targets related to DR, and regulate multiple signal pathways, thereby exerting a therapeutic effect on DR.

Gingko biloba extract improves the lipid profile, inflammatory markers, leptin level and the antioxidant status of T2DM patients poorly responding to metformin: A double- blind, randomized, placebo-controlled trial

Abstract The present study aims to evaluate the effects of Ginkgo biloba (GKB) extract as "add- on" therapy with metformin on the lipid profile, inflammatory markers, leptin and the total antioxidant capacity (TAOC) of patients with type 2 diabetes mellitus (T2DM). It is a multi- center, randomized, placebo-controlled double-blinded clinical study. Sixty patients were allocated into two groups: control and treatment groups; they received orally either 120 mg starch/capsule or 120mg GKB/capsule, respectively as an adjuvant with metformin for 90 days. Blood samples were obtained at zero time and after 90 days. The blood was utilized for analysis of the lipid profile, inflammatory markers, leptin, and TAOC. The GKB extract produced a significant decrease in the levels of TG, LDL-c, and CRP, with a significant increase in HDL-c compared to baseline values. There were no significant changes reported in the placebo-treated group. It also produced a significant decrease in the concentrations of IL-6, TNF-α, and leptin compared to baseline values and placebo-treated groups with a significant increase in TAOC compared to baseline values. In conclusion, GKB extract, as an adjuvant with metformin, decreases inflammatory mediators, leptin level and improves the antioxidant status and lipid profile of T2DM patients improperly managed with metformin

Effects of ginkgo biloba tablets on liver injury in patients with coal-burning-borne arsenism based on DNA damage and repair inhibition / 中华地方病学杂志

Objective:To investigate the role of DNA damage and repair inhibition in the effect of ginkgo biloba on liver injury in patients with coal-burning-borne arsenism.Methods:In March 2017, the investigation was conducted in Jiaole village arsenic poisoning area in Yuzhang Town, Xingren County, Guizhou Province. According to the "Diagnosis of Endemic Arsenicosis" (WS/T 211-2015) and the "Diagnostic Criteria of Occupational Toxic Hepatopathy" (GBZ 59-2010), 52 patients with arsenism were selected as the ginkgo biloba intervention group, and 49 cases of arsenism patients as intervention control group. Ginkgo biloba tablets were given orally for 3 months (1 tablet/time, 3 times/d) according to the commonly used clinical methods, and no other drugs were given to all subjects during the intervention period. The intervention control group was given placebo in the same way as that of ginkgo biloba intervention group. A total of 41 residents who did not burn high arsenic coal 12 km away with no abnormal liver function were selected as normal control group. Physical examinations were performed before the intervention and at the end of the intervention at 3 months. After receiving signed informed consent, morning urine and peripheral venous blood samples were collected to detect urinary arsenic content by inductively coupled plasma mass spectrometry (ICP-MS); liver function biochemical indexes [albumin (ALB), albumin/globulin (A/G), cholinesterase (CHE), total bile acid (TBA)] were determined by automatic biochemical analyzer, DNA damage by single-cell gel electrophoresis assay, and the expression of miR-145 (repair inhibition index) by qRT-PCR.Results:There were 116 subjects, 41 in normal control group, 39 in ginkgo biloba intervention group and 36 in intervention control group. In ginkgo biloba and intervention and intervention control groups, there was no significant difference in age, gender, smoking habits and drinking compared with normal control group ( P > 0.05). Urinary arsenic content, TBA level, DNA damage degree [comet tail DNA percentage (TailDNA%) and olive tail moment (OTM)] and plasma miR-145 expression level [(38.75 ± 19.09) μg/g Cr, (11.13 ± 1.55) μmol/L, 8.50 ± 0.88, 7.43 ± 0.68, 5.78 ± 0.75, respectively] in ginkgo biloba intervention group patients before intervention were higher than those in normal control group [(11.62 ± 5.33) μg/g Cr, (5.36 ± 0.87) μmol/L, 5.24 ± 0.33, 4.71 ± 0.29, 2.05 ± 0.27, respectively], the differences were statistically significant ( P < 0.05); the levels of ALB, A/G and CHE were significantly lower than those in normal control group ( P < 0.05). After the intervention of ginkgo biloba, urinary arsenic content, TBA level, DNA damage degree (TailDNA% and OTM) and plasma miR-145 expression level in patients were significantly lower than those before the intervention ( P < 0.05); the levels of ALB, A/G and CHE were significantly higher than those before the intervention ( P < 0.05). There was no significant difference in the above indexes before and after intervention in the intervention control group ( P > 0.05). The results of correlation analysis between DNA damage degree, miR-145 and liver function indexes after the intervention of ginkgo biloba showed that, DNA damage degree (TailDNA% and OTM) was negatively correlated with the levels of ALB, A/G and CHE ( r = - 0.34, - 0.33, - 0.48, - 0.31, - 0.31, - 0.42, P < 0.05), and positively correlated with the level of TBA ( r = 0.49, 0.48, P < 0.05); miR-145 was negatively correlated with the levels of ALB, A/G and CHE ( r = - 0.26, - 0.23, - 0.38, P < 0.05), which was positively correlated with the level of TBA ( r = 0.32, P < 0.05); and DNA damage degree was positively correlated with the expression of miR-145 ( r = 0.65, 0.52, P < 0.05). Conclusion:Ginkgo biloba tablets can alleviate the liver damage caused by arsenic through coal burning, and the mechanism of this process is related to its inhibition of miR-145 expression and reduction of DNA damage.

Effects of increasing efficacy and decreasing toxicity of ginkgo flavone aglycone on doxorubicin in the treatment of liver cancer / 中国药房

OBJECTIVE To study the effects of increasing efficacy and decreasing toxicity of ginkgo flavone aglycone （GA） on doxorubicin （DOX）in the treatment of liver cancer. METHODS A tumor bearing model was established by inoculating liver cancer cell H 22 into the right axillary skin of ICR mice. The successfully modeled mice were randomly divided into model control group，DOX group （2.5 mg/kg，once every other day ，via tail vein ），GA group （30 mg/kg，once a day ，gavage）and GA+DOX group（the usage was the same as single drug groups ），with 6 mice in each group. The administration cycle was 15 days. The general growth of mice in each group were observed ，body weight and tumor weight were measured ，and the inhibition rate of tumor was calculated. Jin’s formula was used to evaluate the effect of combined medication （Q）. The serum level of alpha-fetal protein（AFP），the pathological changes of tumor tissue ，cell apoptosis and the expression of platelet-endothelial cell adhesion molecule-1（CD31）were detected in each group. The cardiac index，serum levels of B-type natriuretic peptide （BNP）and N-terminal pro-brain natriuretic peptide （NT-pro BNP ），pathological changes of heart and myocardial fibrosis degree were also detected. RESULTS The percentage of body weight change （except for GA group ） and tumor weights of DOX group，GA group and GA + DOX group were all decreased significantly，compared with model control group （P＜0.05 or P＜0.01），while tumor weight of GA+DOX gro up was significantly lower than DOX group （P＜0.01）. Inhibitory rates of tumor in 3 administration groups were 54.29％，42.50％ and 89.29％ respectively，and Q of two-drug combination was 1.21. The tumor tissues of mice in each administration group were necrotic to varying degrees ；the serum level of AFP and the expression of CD31 in tumor tissue were decreased significantly ，compared with model control group （P＜0.05 or P＜0.01）；the percentage of necrosis area of tumor tissue and the positive rate of apoptosis （except for single drug groups ）were significantly increased （P＜0.05 or P＜0.01），while positive rate of apoptosis in GA+DOX group was significantly higher than DOX group （P＜0.05）. Cardiac index of mice in DOX group was significantly lower than model control group （P＜0.01）；serum levels of BNP and NT-pro BNP in DOX group and GA+ DOX group were significantly higher than model control group （P＜0.05 or P＜0.01）；pathological changes of heart and the degree of myocardial fibrosis in GA+DOX group were lower than DOX group. CONCLUSIONS GA combined with DOX show synergistic antitumor effect. GA can strengthen the apoptosis promoting effect of DOX ，and can help to reduce the cardiotoxicity of DOX.

Establishment of determination method for five Ginkgo biflavones in Ginkgo biloba leaves tablets / 中国药房

OBJECTIVE To establish a met hod for the determination of amentoflavone ，bilobetin，ginkgetin，isoginkgetin and sciadopitysin in Ginkgo biloba leaves tablets. METHODS After extracted with methanol ，ultra-performance liquid chromatography （UPLC）was adopted to determine G. biloba leaves tablets. The determination was performed on Waters Acquity UPLC HSS T 3 column with acetonitrile- 0.4％ phosphoric acid as mobile phase （gradient elution ）at the flow rate of 0.4 mL/min. The column temperature was set at 35 ℃，and the detection wavelength was 340 nm. The sample size were 1 μL（substance control ）and 10 μL （test sample ）. The relative correction factors （RCFs）of bilobetin ，ginkgetin，isoginkgetin and sciadopitysin were calculated by quantitative analysis of multicomponents by single marker （QAMS）using amentoflavone as control. The chromatographic peak was located with the relative retention time method. Then the contents of the above components were calculated ，and the results were compared with those of external standard method （ESM）（except for amentoflavone ）. RESULTS The linear ranges of amentoflavone，bilobetin，ginkgetin，isoginkgetin and sciadopitysin were 0.10-8.21，0.24-19.34，0.16-12.98，0.22-17.66，0.06-4.86 ng，respectively（all r＞0.999）. The quantitation limits were 0.10，0.24，0.16，0.22，0.06 ng，respectively. RSDs of precision ， repeatability and stability tests （36 h）were all lower than 3.00％. The average recoveries were 99.77％-102.85％，and RSDs were 1.90％-4.40％（n＝6）. The average RCFs of bilobetin ，ginkgetin，isoginkgetin and sciadopitysin were 0.91，0.93，0.96 and 0.95， respectively. The average relative retention times were 1.08，1.18，1.19 and 1.30，respectively. The relative deviation between the calculation result of QAMS and ESM was within ±3.00％. CONCLUSIONS The established method is accurate and stable ，and can be applied to the determination of Ginkgo biflavones in G. biloba leaves tablets and control the quality.

Quality standard improvement study of ginkgo semen decoction pieces / 药学实践杂志

Objective To establish and improve the quality standard of ginkgo semen decoction pieces. Methods The morphological character for 29 batches of ginkgo gemen and 12 batches of stir-fried ginkgo gemen were observed, and the moisture contents were assayed using the method in Chinese Pharmacopoeia 2015 edition, the first supplement. Results The character of ginkgo gemen and stir-fried ginkgo gemen were consistent in different batches. The moisture content of ginkgo gemen was 8.8% to 12.2%, with an average of 10.5%. The moisture content of stir-fried ginkgo gemen was from 5.4% to 12.3%, with an average of 8.9%. Considering that ginkgo semen decoction pieces are stored for a long time, they are prone to the attack of mildew and insects, and the moisture limit is set to be no more than 10.0% in ginkgo gemen and stir-fried ginkgo. Conclusion Compared with the Chinese Pharmacopoeia 2015 edition, the first supplement, the character identification for ginkgo semen decoction pieces was added and the quality standards were improved. The moisture content of ginkgo semen decoction pieces needs to be strictly controlled under 10.0% to prevent mildew and insects.

Expert consensus on clinical application of GBE50 Dispersible Tablets for ischemic cardiovascular and cerebrovascular diseases / 中国中药杂志

Ginkgo biloba Extract( GBE50) Dispersible Tablets is a new standardized prescription,which is widely used in the treatment of ischemic cardiovascular and cerebrovascular diseases. However,there are still many problems in its clinical application.Rational and safe use of GBE50 Dispersible Tablets is pivotal to the medication safety and clinical prognosis of patients. This consensus has been jointly formulated by clinical experts of traditional Chinese medicine and western medicine in cardiovascular and cerebrovascular diseases and followed the Manual for the Clinical Experts Consensus of Chinese Patent Medicine published by the China Association of Chinese Medicine. The present study identified clinical problems based on clinical investigation,searched the research papers according to PICO clinical problems,carried out evidence evaluation,classification,and recommendation by GRADE system,and reached the expert consensus with nominal group technique. The consensus combines evidence with expert experience. Sufficient evidence of clinical problems corresponds to " recommendations",while insufficient evidence to " suggestions". Safety issues of GBE50 Dispersible Tablets,such as indications,usage and dosage,and medication for special populations,are defined to improve clinical efficacy,promote rational medication,and reduce drug risks. This consensus needs to be revised based on emerging clinical issues and evidencebased updates in practical applications in the future.

A network pharmacology-based strategy for predicting the protective mechanism of Ginkgo biloba on damaged retinal ganglion cells / 中国天然药物

Hallmarks of the pathophysiology of glaucoma are oxidative stress and apoptotic death of retinal ganglion cells (RGCs). Ginkgo biloba extract (EGb) with multi-target, multi-pathway functions has been reported to exert positive pharmacological effects on oxidative stress and damaged RGCs. However, the ingredients and anti-apoptotic targets of EGb in the treatment of open-angle glaucoma (OAG) have not been fully elucidated. Therefore, in-depth analysis is necessary for further research. Ginkgo biloba-related and anti-apoptotic targets were identified and then combined to obtain the intersection, representing the potential anti-apoptotic targets of Ginkgo biloba. In addition, compound-anti-apoptotic target and OAG-target protein-protein interaction network were merged to obtain five core genes and compound-OAG-anti-apoptotic target protein-protein interaction network. Consequently, the active compounds and anti-apoptotic targets of Ginkgo biloba in the treatment of OAG were identified, namely luteolin, β-sitosterol, kaempferol, stigmasterol, quercetin, and p53, Bax, Bcl-2, Caspase-3 and Caspase-9, respectively. For the anti-apoptotic targets of Ginkgo biloba in the treatment of OAG, Gene Ontology (GO) and pathway analysis were executed to confirm the gene functions of Ginkgo biloba in antagonizing apoptosis of RGCs. The pathway enrichment was mainly involved in transcriptional activation of p53 responsive genes, activation of caspases and apoptotic processes. Finally, we confirmed the results of the network analysis by H2O2 treated RGC-5 cells in vitro. The results demonstrated that EGb protection can effectively diminish H2O2-induced apoptosis by inhibiting p53 acetylation, reducing the ratio of Bax/Bcl-2 and suppressing the expression of specific cleavage of Caspase-9 and Caspase-3.

The dietary supplement of Ginkgo biloba: a comprehensive review of its potential interactions based on pre-clinical and clinical evidences / El suplemento dietético de Ginkgo biloba: una revisión exhaustiva de sus posibles interacciones basada en evidencias clínicas y preclínicas

This review present Gingko biloba (GB) interactions, based on clinical and pre-clinical presentations. Literature was retrieved using databases; ScienceDirect, PubMed, Google scholar, Web of Science, Scopus etc. 14/45 interactions were found with clinical presentations. More interactions (80%) were reported with drugs followed by herbs (11.1%), and nutraceuticals (6.7%) with major mechanisms of interaction observed as; inhibition of Cytochrome metabolizing enzymes (44.4%) and platelet-activating factor (PAF) i.e. 15.6%. Major clinical features were; increased bleeding (eye, parietal), hematomas (subdural), and seizures as well as increased blood pressure, priapism, loss of infection/antiviral failure, and coma. Drugs with major interactions belonged to anti-platelet/anti-coagulant and NSAIDs. Synergistic effects were observed for GB vs herbs (except cannabis which showed rhabdomyolysis), foods, and nutraceuticals (except pyridoxine where neurotoxicity was seen). GB use should be monitored and the patient may seek proper advice from a healthcare professional.

Esta revisión presenta las interacciones de Gingko biloba (GB), basadas en presentaciones clínicas y preclínicas. La literatura se recuperó utilizando bases de datos; ScienceDirect, PubMed, Google Scholar, Web of Science, Scopus, etc. Se encontraron 14/45 interacciones con presentaciones clínicas. Se informaron más interacciones (80%) con fármacos seguidos de hierbas (11,1%) y nutracéuticos (6,7%) con los principales mecanismos de interacción observados como; inhibición de las enzimas metabolizadoras del citocromo (44,4%) y factor activador de plaquetas (PAF), es decir, 15,6%. Las principales características clínicas fueron; aumento de sangrado (ojo, parietal), hematomas (subdural) y convulsiones, así como aumento de la presión arterial, priapismo, pérdida de infección / insuficiencia antiviral y coma. Los fármacos con interacciones importantes pertenecían a los antiplaquetarios/anticoagulantes y los AINE. Se observaron efectos sinérgicos para GB frente a hierbas (excepto cannabis que mostró rabdomiólisis), alimentos y nutracéuticos (excepto piridoxina donde se observó neurotoxicidad). Se debe controlar el uso de GB y el paciente puede buscar el asesoramiento adecuado de un profesional de la salud.

The mechanism by which Ginkgo biloba extract induces the apoptosis of human laryngeal cancer Hep-2 cells by reducing reactive oxygen species level and activating JNK signaling pathway / 中国基层医药

Objective:To investigate the apoptosis-inducing effect of Ginkgo biloba extract (Ginaton) on human laryngeal cancer Hep-2 cells and the underlying molecular mechanism.Methods:Human laryngeal cancer Hep-2 cells were cultured in vitro and human laryngeal cancer Hep-2 cells in the log phase were treated with Ginaton in time and concentration gradients. The cell counting kit-8 (CCK-8) assay was performed to investigate the inhibitory effects of Ginaton on Hep-2 cells. Flow cytometry was performed to detect apoptosis and determine the level of reactive oxygen species (ROS). Western blot assay was performed to detect apoptosis and signaling pathway-related protein expression. Results:Ginaton inhibited the proliferation of Hep-2 cells in a time-dependent and concentration-dependent manner. Malondialdehyde level decreased gradually in a time-dependent manner, and decreased to 2.98 μmol/g after 24 hours of Ginaton treatment. Superoxide dismutase level increased gradually in a time-dependent manner and increased to 90.35 U/g after 24 hours of Ginaton treatment. ROS level decreased gradually in a time-dependent manner and deceased to 18.7% of the level before treatment after 24 hours of Ginaton treatment. There was no significant difference in ROS level between before and after 24 hours of Ginaton treatment ( F = 14.98, 19.65, 11.47, all P < 0.001). After 3, 6, 12 and 24 hours of Ginaton treatment, the expression of phosphorylated N-terminal protein kinase increased to 1.98, 2.57, 2.91 and 3.28 in a time-dependent manner. There was significant difference in the expression of phosphorylated N-terminal protein kinase between before treatment and after 3, 6, 12 and 24 hours of Ginaton treatment ( F = 16.37, P < 0.001). Conclusion:Ginaton can effectively inhibit the proliferation and induce apoptosis of human laryngeal cancer Hep-2 cells in vitro, which may be related to regulating ROS level and activating JNK signaling pathway.

Ginkgo biloba regulate Nrf2-Keap1-ARE signaling pathway to ameliorate liver injury in endemic arsenic poisoning rats induced by coal burning / 中华地方病学杂志

Objective:To explore the effects of Ginkgo biloba on regulating NF-E2-related factor 2 (Nrf2)-Kelch-like ECH-associated protein 1 (Keap1)-antioxidant response element (ARE) signaling pathway in liver injury induced by coal-burning-borne endemic arsenic poisoning in rats.Methods:Group design method was adopted, according to body weight (80-100 g), a total of 30 Wistar rats were divided into 5 groups (6 rats in each group, half males and half females) by random number table method. The normal control group was fed with normal diet ad libitum for 4.5 months; the Ginkgo biloba control group was fed with Ginkgo biloba (25 mg/kg, 6 d/week) for 1.5 months after normal feeding for 3 months; the drinking water arsenic poisoning group and the arsenic contaminated grain group were fed respectively with 100 mg/L arsenic trioxide (As 2O 3) solution and 100 mg/kg arsenic-containing feed for 3 months, and then fed with normal diet for 1.5 months; the Ginkgo biloba treatment group was fed with 100 mg/kg arsenic-containing feed for 3 months, and then was given Ginkgo biloba (25 mg/kg, 6 d/week) for 1.5 months. After sacrificing the animals, the content of malondialdehyde (MDA), the activity of copper zinc superoxide dismutase (SOD1) and the activity of glutathione peroxidase (GPx) in serum were detected by thiobarbituric acid colorimetry, xanthine oxidase method and dimercaptodinitrobenzoic acid reduction method, respectively. The mRNA and protein expressions of indicator genes of Nrf2-Keap1-ARE signaling pathway in liver tissues were detected by quantitative real-time PCR, immunohistochemistry and Western blotting. Correlation between the indexes was analyzed by Pearson. Results:In drinking water arsenic poisoning group, arsenic contaminated grain group and Ginkgo biloba treatment group, the contents of MDA in serum were (3.54±0.51), (3.83±0.87) and (2.93±0.84) μmol/L, respectively, which were higher than that in normal control group [(1.85±0.36) μmol/L, P < 0.05]; and SOD1 activities [(68.21±4.37), (64.53±9.96), (73.09±5.43) U/ml] and GPx activities [(486.41±40.45), (458.24±42.25), (539.79±79.43) U/L] in serum were lower than those in normal control group [(81.47±5.73) U/ml, (747.86±80.33) U/L, P < 0.05]. Compared with the arsenic contaminated grain group, the content of MDA in serum in Ginkgo biloba treatment group was decreased, the activities of SOD1 and GPx in serum were increased ( P < 0.05). Compared with normal control group, the mRNA expressions of SOD1 and GPx1 in the liver tissues in drinking water arsenic poisoning group, arsenic contaminated grain group and Ginkgo biloba treatment group were significantly higher ( P < 0.05). Compared with arsenic contaminated grain group, the mRNA expressions of SOD1 and GPx1 in the liver tissue in Ginkgo biloba treatment group were increased ( P < 0.05). Compared with the normal control group, the protein expression of SOD1 in liver tissue in arsenic contaminated grain group was decreased ( P < 0.05), the protein expressions of GPx1 were decreased in the liver tissues in drinking water arsenic poisoning group, arsenic contaminated grain group and Ginkgo biloba treatment group ( P < 0.05). Compared with the arsenic contaminated grain group, the protein expressions of SOD1 and GPx1 were increased in the liver tissue in Ginkgo biloba treatment group ( P < 0.05). Compared with the normal control group and arsenic contaminated grain group, the protein expression of Keap1 was decreased in the liver tissue in Ginkgo biloba treatment group ( P < 0.05). Compared with the normal control group, the protein expressions of Nrf2 and phosphorylation of Nrf2 (pNrf2) were increased in the cytoplasm in drinking water arsenic poisoning group, arsenic contaminated grain group and Ginkgo biloba treatment group ( P < 0.05). Compared with the arsenic contaminated grain group, the protein expression of pNrf2 was decreased in the cytoplasm in Ginkgo biloba treatment group ( P < 0.05). The protein expressions of Nrf2 and pNrf2 in the nucleus in drinking water arsenic poisoning group, arsenic contaminated grain group and Ginkgo biloba treatment group were also higher than those in normal control group ( P < 0.05). Compared with the arsenic contaminated grain group, the protein expressions of Nrf2 and pNrf2 were increased in the nucleus in Ginkgo biloba treatment group ( P < 0.05). The results of correlation analysis revealed that the protein expressions of Nrf2 and pNrf2 in the nucleus were negatively correlated with Keap1 protein expression ( r=-0.523,-0.401, P < 0.05), and positively correlated with the mRNA expressions of SOD1 and GPx1 ( r=0.658, 0.530, 0.555, 0.603, P < 0.05). In addition, the protein expressions of SOD1 and GPx1 were positively correlated with their enzyme activities ( r=0.472, 0.629, P < 0.05). Conclusions:Arsenic could induce oxidative stress and liver injury. Ginkgo biloba could reduce the protein expression of Keap1, and promote nuclear translocation of Nrf2, which might induce the up-regulation of mRNA expressions of SOD1 and GPx1, and partially reverse the posttranscriptional regulation of arsenic on SOD1 and GPx1, and then increase their protein expressions and enzyme activities, thereby improve arsenic induced oxidative stress and liver injury.

Ginkgo biloba extract ameliorates streptozotocin-induced diabetes in rats as measured by non-targeted metabolomics / 药学学报

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.

Neuroprotective effects of Ginkgo biloba dropping pills in Parkinson's disease / 药物分析学报

Parkinson's disease(PD)is the second most common neurodegenerative disease in the world;however,it lacks effective and safe treatments.Ginkgo biloba dropping pill(GBDP),a unique Chinese G.biloba leaf extract preparation,exhibits antioxidant and neuroprotective effects and has a potential as an alternative therapy for PD.Thus,the aims of this study were to evaluate the effects of GBDP in in vitro and in vivo PD models and to compare the chemical constituents and pharmacological activities of GBDP and the G.biloba extract EGb 761.Using liquid chromatography tandem-mass spectrometry,46 GBDP constitu-ents were identified.Principal component analysis identified differences in the chemical profiles of GBDP and EGb 761.A quantitative analysis of 12 constituents showed that GBDP had higher levels of several flavonoids and terpene trilactones than EGb 761,whereas EGb 761 had higher levels of organic acids.Moreover,we found that GBDP prevented 6-hydroxydopamine-induced dopaminergic neuron loss in zebrafish and improved cognitive impairment and neuronal damage in methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced PD mice.Although similar effects were observed after EGb 761 treatment,the neuroprotective effects were greater after GBDP treatment on several endpoints.In addition,in vitro results suggested that the Akt/GSK3β pathway may be involved in the neuroprotective effects of GBDP.These findings demonstrated that GBDP have potential neuroprotective effects in the treatment of PD.

Comparative study on the effects of total ginkgo flavonol glycosides and total ginkgolides on neurofunctional recovery in mice with subacute stroke / 药学学报

As a Ginkgo biloba extract preparation, shuxuening injection has a unique advantage in the prevention and treatment of acute and subacute stroke, but its main active ingredient is still unclear. Using a subacute model of stroke in mice constructed earlier, we further explored the contribution and mechanism of the two main components of total ginkgo flavonol glycosides and total ginkgolides in facilitating the neurofunctional recovery in stroke-induced mice. The pharmacodynamics was mainly evaluated by neurobehavioral changes, cerebral infarction volume, blood-brain barrier permeability and brain edema. The pathway and targets were predicted by transcriptome and network pharmacology. Finally, the mechanism was verified at the mRNA and protein levels. The results showed that the beneficial effect of total ginkgolides was greater than that of total ginkgo flavonol glycosides in both the pharmacodynamics and the regulatory mechanism of granulocyte adhesion and diapedesis involving granulocyte colony-stimulating factor (G-CSF), macrophage-1 antigen (MAC-1) and E-selectin. These findings suggest that shuxuening injection may improve the prognosis for mice with subacute stroke by down-regulating G-CSF-mediated granulocyte adhesion and diapedesis pathway mainly through the total ginkgolide components. This finding is expected to provide reference for optimizing prescription and searching for natural drugs for targeting the treatment of ischemic stroke prognosis. The animal experiments in this study followed the regulations of Animal Ethics Committee of Tianjin University of Traditional Chinese Medicine.