LC-MS based untargeted metabolomics studies of the metabolic response of Ginkgo biloba extract on arsenism patients.

Arsenic is an environmentally ubiquitous toxic metalloid. Chronic exposure to arsenic may lead to arsenicosis, while no specific therapeutic strategies are available for the arsenism patients. And Ginkgo biloba extract (GBE) exhibited protective effect in our previous study. However, the mechanisms by which GBE protects the arsenism patients remain poorly understood. A liquid chromatography-mass spectrometry (LC-MS) based untargeted metabolomics analysis was used to study metabolic response in arsenism patients upon GBE intervention. In total, 39 coal-burning type of arsenism patients and 50 healthy residents were enrolled from Guizhou province of China. The intervention group (n = 39) were arsenism patients orally administered with GBE (three times per day) for continuous 90 days. Plasma samples from 50 healthy controls (HC) and 39 arsenism patients before and after GBE intervention were collected and analyzed by established LC-MS method. Statistical analysis was performed by MetaboAnalyst 5.0 to identify differential metabolites. Multivariate analysis revealed a separation in arsenism patients between before (BG) and after GBE intervention (AG) group. It was observed that 35 differential metabolites were identified between BG and AG group, and 30 of them were completely or partially reversed by GBE intervention, with 14 differential metabolites significantly up-regulated and 16 differential metabolites considerably down-regulated. These metabolites were involved in promoting immune response and anti-inflammatory functions, and alleviating oxidative stress. Taken together, these findings indicate that the GBE intervention could probably exert its protective effects by reversing disordered metabolites modulating these functions in arsenism patients, and provide insights into further exploration of mechanistic studies.

Ginkgo biloba L. exocarp petroleum ether extract inhibits methicillin-resistant Staphylococcus aureus by modulating ion transport, virulence, and biofilm formation in vitro and in vivo.

ETHNOPHARMACOLOGICAL RELEVANCE: As reported in the Ancient Chinese Medicinal Books, Ginkgo biloba L. fruit has been used as a traditional Chinese medicine for the treatment asthma and cough or as a disinfectant. Our previous study demonstrated that G. biloba exocarp extract (GBEE), an extract of a traditional Chinese herb, inhibits the formation of methicillin-resistant Staphylococcus aureus (MRSA) biofilms. However, GBEE is a crude extract that contains many components, and the underlying mechanisms of purified GBEE fractions extracted with solvents of different polarities are unknown. AIM OF THE STUDY: This study aimed to investigate the different components in GBEE fractions extracted with solvents of different polarities and their antibacterial effects and mechanisms against MRSA and Staphylococcus haemolyticus biofilms both in vitro and in vivo. METHODS: The components in different fractions were detected by high-performance liquid chromatography-high resolution mass spectrometry (HPLC-HRMS). Microbroth dilution assays and time growth curves were used to determine the antibacterial effects of the fractions on 15 clinical bacterial isolates. Crystal violet staining, scanning electron microscopy (SEM) and transmission electron microscopy (TEM) were utilized to identify the fractions that affected bacterial biofilm formation. The potential MRSA targets of the GBEE fraction obtained with petroleum ether (PE), denoted GBEE-PE, were screened by transcriptome sequencing, and the gene expression profile was verified by quantitative polymerase chain reaction (qPCR). RESULTS: HPLC-HRMS analysis revealed that the four GBEE fractions (extracted with petroleum ether, ethyl acetate, n-butanol, and water) contained different ginkgo components, and the antibacterial effects decreased as the polarity of the extraction solvent increased. The antibacterial activity of GBEE-PE was greater than that of the GBEE fraction extracted with ethyl acetate (EA). GBEE-PE improved H. illucens survival and reduced MRSA colonization in model mouse organs. Crystal violet staining and SEM and TEM analyses revealed that GBEE-PE inhibited MRSA and S. haemolyticus biofilm formation. Transcriptional analysis revealed that GBEE-PE inhibits MRSA biofilms by altering ion transport, cell wall metabolism and virulence-related gene expression. In addition, the LO2 cell viability and H. illucens toxicity assay data showed that GBEE-PE at 20 mg/kg was nontoxic. CONCLUSION: The GBEE fractions contained different components, and their antibacterial effects decreased with increases in the polarity of the extraction solvent. GBEE-PE limited MRSA growth and biofilm formation by affecting ion transport, cell wall synthesis, and virulence-related pathways. This research provides a more detailed overview of the mechanism by which GBEE-PE inhibits MRSA both in vitro and in vivo and suggests that GBEE-PE is a new prospective antimicrobial with the potential to be used in MRSA therapeutics in the future.

New phenylbutenoids and terpene glycosides from Ginkgo biloba leaves.

Our continued works on the chemical constituents of Ginkgo biloba (G. biloba) leaves has led to the isolation of two novel phenylbutenoids (1, 2), along with five previously unidentified terpene glycosides (3-7). Among them, compounds 1 and 2 represent unique (Z)-phenylbutenoids, 3-6 are megastigmane glycosides, and 7 is identified as a rare bilobanone glycoside (Fig. 1). This study marks the first reported isolation of phenylbutenoid and bilobanone glycoside from G. biloba. The chemical structures of these compounds were elucidated through extensive spectroscopic analysis, including HR-ESI-MS and various 1D and 2D NMR experiments. Furthermore, the absolute configurations of these molecules were determined using Mosher's method, ECD experiments, and Cu-Kα X-ray crystallographic analyses.

The molecular mechanisms of ginkgo (Ginkgo biloba) activity in signaling pathways: A comprehensive review.

BACKGROUND: One of the most unique plants that have ever grown on the planet is Ginkgo biloba L., a member of the Ginkgoaceae family with no close living relatives. The existence of several differently structured components of G. biloba has increased the chemical variety of herbal therapy. Numerous studies that investigated the biochemical characteristics of G. biloba suggest this plant as a potential treatment for many illnesses. PURPOSE: Review the molecular mechanisms involved in the signaling pathways of G. biloba activity in varied circumstances and its potential as a novel treatment for various illnesses. METHODS: Studies focusing on the molecular processes and signaling pathways of compounds and extracts of G. biloba were found and summarized using the proper keywords and operators from Google Scholar, PubMed, Web of Science, and Scopus without time restrictions. RESULTS: G. biloba exerts its effects through its anti-inflammatory, anti-apoptotic, anti-cancer, neuroprotective, cardioprotective, hepatoprotective, antiviral, antibacterial, pulmoprotective, renoprotective, anti-osteoporosis, anti-melanogenic, retinoprotective, otoprotective, adipogenic, and anti-adipogenic properties. The most important mechanisms involved in these actions are altering the elevation of ROS formation, inhibiting NADPH oxidases activation, altering the expression of antioxidant enzymes, downregulating MAPKs (p38 MAPK and ERK, and JNK) and AP-1, increasing cAMP, inactivating Stat5, activating the AMPK signaling pathway, affecting Stat3/JAK2, NF-κB, Nrf-2, mTOR, HGF/c-Met, Wnt/ß-catenin and BMP signaling pathways, and changing the mitochondrial transmembrane potential, the Bax/Bcl-2 ratio, the release of Cyc from mitochondria to cytosol, the protein cleavage of caspases 3, 7, 8, 9, and 12, poly (ADP-ribose) polymerase, and MMPs levels. CONCLUSIONS: G. biloba and its components have gained attention in recent years for their therapeutic benefits, such as their anti-inflammatory, antioxidant, anti-apoptotic, and apoptotic effects. By understanding their molecular mechanisms and signaling pathways, potential novel medicines might be developed in response to the rising public desire for new therapies.

Study on network pharmacology of Ginkgo biloba extract against ischaemic stroke mechanism and establishment of UPLC-MS/MS methods for simultaneous determination of 19 main active components.

INTRODUCTION: Ginkgo biloba extract (GBE) is an effective substance from traditional Chinese medicine (TCM) G. biloba for treating ischaemic stroke (IS). However, its active ingredients and mechanism of action remain unclear. OBJECTIVES: This study aimed to reveal the potential active component group and possible anti-IS mechanism of GBE. MATERIALS AND METHODS: The network pharmacology method was used to reveal the possible anti-IS mechanism of these active ingredients in GBE. An ultra-high-performance liquid chromatography triple quadrupole electrospray tandem mass spectrometry (UPLC-MS/MS) method was established for the simultaneous detection of the active ingredients of GBE. RESULTS: The active components of GBE anti-IS were screened by literature integration. Network pharmacology results showed that the anti-IS effect of GBE is achieved through key active components such as protocatechuic acid, bilobalide, ginkgolide A, and so on. Gene Ontology (GO) function and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis showed that the possible anti-IS mechanism of GBE is regulating the PI3K-Akt signalling pathway and other signal pathways closely related to inflammatory response and apoptosis regulation combined with AKT1, MAPK, TNF, ALB, CASP3, and other protein targets. Nineteen main constituents in seven batches of GBE were successfully analysed using the established UPLC-MS/MS method, and the results showed that the content of protocatechuic acid, gallic acid, ginkgolide A, and so forth was relatively high, which was consistent with network pharmacology results, indicating that these ingredients may be the key active anti-IS ingredients of GBE. CONCLUSION: This study revealed the key active components and the anti-IS mechanism of GBE. It also provided a simple and sensitive method for the quality control of related preparations.

Effects of Zinc Oxide Nanoparticles on Growth, Development, and Flavonoid Synthesis in Ginkgo biloba.

Ginkgo biloba is a highly valuable medicinal plant known for its rich secondary metabolites, including flavonoids. Zinc oxide nanoparticles (ZnO-NPs) can be used as nanofertilizers and nano-growth regulators to promote plant growth and development. However, little is known about the effects of ZnO-NPs on flavonoids in G. biloba. In this study, G. biloba was treated with different concentrations of ZnO-NPs (25, 50, 100 mg/L), and it was found that 25 mg/L of ZnO-NPs enhanced G. biloba fresh weight, dry weight, zinc content, and flavonoids, while 50 and 100 mg/L had an inhibitory effect on plant growth. Furthermore, quantitative reverse transcription (qRT)-PCR revealed that the increased total flavonoids and flavonols were mainly due to the promotion of the expression of flavonol structural genes such as GbF3H, GbF3'H, and GbFLS. Additionally, when the GbF3H gene was overexpressed in tobacco and G. biloba calli, an increase in total flavonoid content was observed. These findings indicate that 25 mg/L of ZnO-NPs play a crucial role in G. biloba growth and the accumulation of flavonoids, which can potentially promote the yield and quality of G. biloba in production.

Uncovering the anti-obesity constituents in Ginkgo biloba extract and deciphering their synergistic effects.

Obesity has been recognized as a key risk factor for multiple metabolic disorders, including diabetes, cardiovascular diseases and many types of cancer. Herbal medicines have been frequently used for preventing and treating obesity in many countries, but in most cases, the key anti-obesity constituents in herbs and their anti-obesity mechanisms are poorly understood. This study demonstrated a case study for uncovering the anti-obesity constituents in an anti-obesity herbal medicine (Ginkgo biloba extract) and deciphering their synergistic effects via targeting human pancreatic lipase (hPL). Following screening the anti-hPL effects of eighty herbal medicines, Ginkgo biloba extract (GBE50) was found with the most potent anti-hPL activity. Global chemical profiling of herbal constituents coupling with hPL inhibition assay revealed that the bioflavonoids and several flavonoids in GBE50 were key anti-hPL constituents. Among all tested thirty-eight constituents, sciadopitysin, bilobetin, quercetin, isoginkgetin, and ginkgetin showed potent anti-hPL effects (IC50 values <2.5 µM). Inhibition kinetic analyses suggested that sciadopitysin, bilobetin, quercetin, isoginkgetin, and ginkgetin acted as non-competitive inhibitors of hPL, with the Ki values were <2 µM. Docking simulations revealed that four bioflavonoids (sciadopitysin, bilobetin, isoginkgetin, and ginkgetin) could tightly bind on hPL at cavity 2, which it is different from the binding cavity of quercetin on hPL. Further investigations demonstrated that the combinations of quercetin and one bioflavonoid-type hPL inhibitor (sciadopitysin or bilobetin) showed synergistic anti-hPL effects, suggesting that the multi-components in GBE50 may generate more potent anti-hPL effect. Collectively, our findings uncovered the anti-obesity constituents in GBE50, and explored their anti-hPL mechanisms as well as synergistic effects at molecular levels, which will be very helpful for further understanding the anti-obesity mechanisms of Ginkgo biloba.

Preparation and functional characteristics of protein from Ginkgo endophytic Pseudomonas R6 and Ginkgo seed.

Ginkgo seed protein (GSP) has excellent processing characteristics and antioxidant properties. In this study, Gingko endophytic protein (GEP) was synthesized by Ginkgo endophytic Pseudomonas R6. SDS-PAGE analysis indicated that the molecular weights of GSP and GEP were mainly distributed at 17 KDa and 48 KDa, respectively. FTIR showed that GEP and GSP exhibited characteristic absorption in the amide I, II, and III bands, and absorption in amide A and B indicated the presence of hydrogen bonding. HPLC analysis showed that both proteins had 17 amino acids, but their relative abundance was different, with GSP having the highest Ser content (74.713 mg/g) and GEP having the highest Val content (35.905 mg/g). Stomata were observed on the surface of both proteins by SEM, and there were lamellar and some spherical structures on GEP, while the opposite was observed on GSP. GEP had superior solubility, OHC, FC and EC, while GSP showed good WHC. Both proteins exhibited antioxidant activities, with GSP exhibiting stronger hydroxyl radical scavenging ability than GEP, with IC50 of 0.46 mg/mL and 1.54 mg/mL, respectively. This work demonstrates the antioxidant potential of GEP as an alternative to GSP in the food industry.

Sodium alginate coating of Ginkgo biloba leaves extract containing phenylpropanoids as an ecofriendly preserving agent to maintain the quality of peach fruit.

Plant constituents are of great interest in the food processing industry as potential natural preservative agents for controlling foodborne pathogens. In this study, the 95% EtOH/H2 O extract of Ginkgo biloba leaves was separated using polarity extraction solvents with petroleum ether (PE), ethyl acetate (EA), n-butanol (nB), and water (W) by the principle of similarity and compatibility. Through TLC and NMR analysis of these extracts, it can be concluded that the main component of PE extract were organic acids, for EA extract were flavonoids, for nB extract were phenylpropanoids, and water extract were oligosaccharides. Twelve monomer compounds were separated from the extracts to verify the composition of each extraction stage. Results of morphological and molecular identification revealed that Monilinia fructicola and Rhizopus stolonifer were the main fungi causing peach rot. After evaluating the antifungal activity and peach quality of the four extract/sodium alginate coatings, it was found that the n-butanol extract/sodium alginate coating containing phenylpropanoids had the lowest decay index and the best preservation effect, providing a sustainable alternative to reduce the harm to the environment of synthetic preservatives. PRACTICAL APPLICATION: The abuse of synthetic preservatives poses a threat to the ecological environment and physical health. Therefore, this study developed sodium alginate coating of Ginkgo biloba leaves extract containing phenylpropanoids, which has good effects on the preservation of peaches. The agent is a promising environmentally friendly alternative for synthetic preservatives.

The Potential of Ginkgo biloba as a Source of Biologically Active Compounds-A Review of the Recent Literature and Patents.

Ginkgo biloba is a relict tree species showing high resistance to adverse biotic and abiotic environmental factors. Its fruits and leaves have high medicinal value due to the presence of flavonoids, terpene trilactones and phenolic compounds. However, ginkgo seeds contain toxic and allergenic alkylphenols. The publication revises the latest research results (mainly from 2018-2022) regarding the chemical composition of extracts obtained from this plant and provides information on the use of extracts or their selected ingredients in medicine and food production. A very important section of the publication is the part in which the results of the review of patents concerning the use of Ginkgo biloba and its selected ingredients in food production are presented. Despite the constantly growing number of studies on its toxicity and interactions with synthetic drugs, its health-promoting properties are the reason for the interest of scientists and motivation to create new food products.

Effect of Ginkgo biloba extract on pharmacology and pharmacokinetics of atorvastatin in rats with hyperlipidaemia.

Ginkgo biloba extract (GBE) is a common dietary supplement used by people with dyslipidaemia worldwide to reduce the risk of cardiovascular disease. Many studies have found that GBE itself has a variety of pharmacological activities. However, the role of GBE as an adjunct to conventional therapy with chemical drugs remains controversial. Therefore, this study explored the additional benefits of GBE in the treatment of hyperlipidaemia with statins in terms of both pharmacodynamics and pharmacokinetics. A hyperlipidaemia model was established by feeding rats a high-fat diet for a long time. The animals were treated with atorvastatin only, GBE only, or a combination of atorvastatin and GBE. The results showed that statins combined with GBE could significantly improve the blood lipid parameters, reduce the liver fat content, and reduce the size of adipocytes in abdominal fat. The effect was superior to statin therapy alone. In addition, the combination has shown additional liver protection against possible pathological liver injury or statin-induced liver injury. A lipidomic study showed that GBE could regulate the abnormal lipid metabolism of the liver in hyperlipemia. When statins are combined with GBE, this callback effect introduced by GBE on endogenous metabolism has important implications for resistance to disease progression and statin resistance. Finally, in the presence of GBE, there was a significant increase in plasma statin exposure. These results all confirmed that GBE has incremental benefits as a dietary supplement of statin therapy for dyslipidaemia.

Efficacy and safety of Ginkgo biloba extract as an adjuvant in the treatment of Chinese patients with sudden hearing loss: a meta-analysis.

CONTEXT: Ginkgo biloba Linn (Ginkgoaceae) [leaves extract (GBE)] is authorized for the treatment of sudden hearing loss (SHL); however, its clinical feasibility in SHL has not been thoroughly investigated. OBJECTIVE: To evaluate the efficacy and safety of adjuvant GBE in the treatment of SHL. MATERIALS AND METHODS: We used PubMed, EMBASE, Web of Science, Cochrane Library, China National Knowledge Infrastructure, Wanfang, Chinese Scientific Journal Database, China Biomedical Database for literature research, starting from inception to 30 June 2022. The key terms: Ginkgo biloba extract, Sudden Sensorineural Deafness. This meta-analysis contained randomized controlled trials that compared the safety and efficacy of the combination of GBE and general treatments (GT) with GT alone for SHL. The extracted data were analyzed using Revman5.4 software with risk ratio (RR), 95% confidence intervals (CI) and mean difference (MD). RESULTS: Our meta-analysis included 27 articles with a total of 2623 patients. The results revealed that the effects of GBE adjuvant therapy was superior than GT (total effective rate: RR = 1.22, 95% CI: 1.18-1.26, p < 0.00001), the pure tone hearing threshold (MD = 12.29, 95% CI: 11.74-12.85, p < 0.00001) and hemorheology indexes (whole blood high shear viscosity: MD = 1.46, 95% CI: 0.47-2.44, p = 0.004) after treatment were significantly improved compared to non-treatment, while there was no significant difference as for hematocrit (red blood cells) (MD = 4.15, 95% CI: -7.15-15.45, p = 0.47). CONCLUSION: The efficacy of GBE + GT for the treatment of SHL may be more promising than GT alone.

Polyprenols in Ginkgo biloba; a review of their chemistry (synthesis of polyprenols and their derivatives), extraction, purification, and bioactivities.

The Ginkgo biloba L. (GB) contains high bioactive compounds. To date, flavonoids and terpene trilactone have received the majority of attention in GB studies, and the GB has been utilized globally in functional food and pharmacological firms, with sales > $10 billion since 2017, while the other active components, for instance, polyprenols (a natural lipid) with various bioactivities have received less attention. Hence, this review focused on polyprenols' chemistry (synthesis of polyprenols and their derivatives) extraction, purification, and bioactivities from GB for the first time. The various extractions and purification methods (nano silica-based adsorbent, bulk ionic liquid membrane, etc.) were delved into, and their advantages and limitations were discussed. Besides, numerous bioactivities of the extracted Ginkgo biloba polyprenols (GBP) were reviewed. The review showed that GB contains some polyprenols in acetic esters' form. Prenylacetic esters are free of adverse effects. Besides, the polyprenols from GB have numerous bioactivities such as anti-bacterial, anti-cancer, anti-viral activity, etc. The application of GBPs in the food, cosmetics, and drugs industries such as micelles, liposomes, and nano-emulsions was delved into. Finally, the toxicity of polyprenol was reviewed, and it was concluded that GBP was not carcinogenic, teratogenic, or mutagenic, giving a theoretical justification for using GBP as a raw material for functional foods. This article will aid researchers to better understand the need to explore GBP usage.

Effects of processing methods on quality, antioxidant capacity, and cytotoxicity of Ginkgo biloba leaf tea product.

BACKGROUND: Ginkgo biloba leaves contain beneficial flavonoids, bilobalide (BB), and ginkgolides. However, the toxic ginkgolic acid (GA) limit its application. In this study, various traditional processing methods were used to prepare G. biloba leaf tea (GBLT), including white tea, black tea, dark tea, green tea, and freeze-dried as control, followed by investigations of their effects on quality, antioxidant capacity, bioactive components, and cytotoxicity of the tea products. RESULTS: Results showed that different processing methods significantly impact the tea products' quality indexes and the principal component analysis (PCA) and hierarchical cluster analysis (HCA) corroborated it. White tea had the highest total sugar (TS) and GA content and the most potent cytotoxicity on HepG2 cells. However, TS and GA content and the cytotoxicity of GBLT markedly decreased during fermentation and fixation. Moreover, white tea possessed higher total phenolic content (TPC), total flavonoid content (TFC), and more vigorous antioxidant activities than green tea, black tea, and dark tea. Terpene trilactones value was stable, but different catechins contents fluctuated according to the manufacturing process of different GBLTs. Among the four GBLTs, dark tea combining fixation and fermentation had the lowest GA content and cytotoxicity, less bioactive components reduction, appropriate quality, and stronger flavor. CONCLUSION: These findings demonstrate that fixation and fermentation help reduce GAs during the manufacturing of GBLT. However, their ability to retain bioactive substances needs further optimization in future studies. © 2023 Society of Chemical Industry.

Antifungal and antifeedant terpenoids from Paraphaeosphaeria sp. cultured by extract of host Ginkgo biloba.

Five undescribed terpenoids including a polyketide-terpenoid hybrid paraphaone, and four eremophilane sesquiterpenoids, paraphaterpenes A-D, as well as two known compounds were isolated from the endophytic fungus Paraphaeosphaeria sp. cultured by extract of host Ginkgo biloba L. The structures were established by spectroscopic analyses, and the single-crystal X-ray diffraction. The antifungal activity of Paraphaeosphaeria sp. cultured by extract of G. biloba against the plant pathogen Alternaria alternata was significant and higher than that of PDB medium. Tested compounds indicated antifeedant activities against silkworms with feeding deterrence index at 10-70%. Paraphaone, paraphaterpenes A, C, D and alternariol methyl ether showed significant antifungal activities against the phytopathogens A. alternata, Aspergillus fumigatus, and entomopathogen Beauveria bassiana with MICs ≤4 µg/mL. And the preliminary structure-activity relationship of eremophilane sesquiterpenoids was exhibited. The culture of Paraphaeosphaeria sp. by host G. biloba medium afforded agricultural antibiotics.

Isolation of the Main Biologically Active Substances and Phytochemical Analysis of Ginkgo biloba Callus Culture Extracts.

The work reveals the results of studying the content of biologically active substances in samples of extracts of Ginkgo biloba callus cultures. Callus cultures grown in vitro on liquid nutrient media were the objects of the study. Considering various factors affecting the yield of the target components during extraction, the volume fraction of the organic modifier in the extracting mixture, the temperature factor, and the exposure time were identified as the main ones. The maximum yield of extractive substances (target biologically active substances with a degree of extraction of at least 50%) from the samples of callus culture extracts was detected at a ratio of extragent of 70% ethanol, a temperature of 50 °C, and exposure time of 6 h. Flavonoids, such as luteolin, quercetin, isoramentin, kaempferol, and amentoflavone, were isolated in the extract samples. As a result of column chromatography, fractions of individual biologically active substances (bilobalide, ginkgolide A, B, and C) were determined. The proposed schemes are focused on preserving the nativity while ensuring maximum purification from associated (ballast) components. Sorbents (Sephadex LH-20, poly-amide, silica gel) were used in successive stages of chromatography with rechromatography. The degree of purity of individually isolated substances was at least 95%.

Metabolic interaction between biflavonoids in Ginkgo biloba leaves and tacrolimus.

The aim of this study was to investigate the effect of biflavonoids in Ginkgo biloba leaves on tacrolimus metabolism. First, the inhibitory effects of five main biflavonoids (amentoflavone, sciadopitysin, ginkgetin, isoginkgetin, bilobetin) in G. biloba leaves on tacrolimus metabolism were investigated in vitro in human liver microsomes (HLM), and the concentration-dependent inhibition was further calculated. Then the time-dependent inhibition activities of five biflavonoids were studied and the drug interaction was studied in Sprague-Dawley (SD) rats. Finally, the molecular mechanism of inhibition was explored by molecular docking. The results of in vitro incubation in HLM showed tacrolimus metabolism was strongly inhibited by amentoflavone, ginkgetin, and bilobetin, whose IC50 value was 5.57, 3.16, and 5.03 µM, respectively. The time-dependent inhibition of the three above biflavonoids at 50 µM was 33.47%-50.89%. In the in vivo study in rats, the AUC0-t and Cmax of tacrolimus increased 3.8-fold and 2.5-fold after oral preadministration with amentoflavone. The molecular docking results showed that the inhibitory effect may be related to the formation of hydrogen bonds. The results showed that long-term combination of G. biloba leaves and tacrolimus may cause drug-drug interactions. This study provided theoretical and experimental basis for rational drug use in clinical practice.

Differences in in vivo absorption of flavone glycosides, flavone aglycones and terpene lactones under different dosage forms and physiological conditions.

ETHNOPHARMACOLOGICAL RELEVANCE: Ginkgo biloba L. extract (GBE) oral preparations have been used for many years in the prevention and treatment of cardiovascular and cerebrovascular diseases, and the main active ingredients are flavonoids and terpene lactones. Among them, the oral absorption of the prototype components of flavonoid glycosides into the blood needs to be further clarified, and the differences in the oral absorption of different components in GBE by different dosage forms and physiological conditions are not clear yet. AIM OF THE STUDY: To clarify the oral absorption of the prototype flavonoid glycosides in vivo, and to further explore the differences in the oral absorption of various active compounds under different oral dosage forms and dietary conditions. MATERIALS AND METHODS: Firstly, the target compounds were selected based on the characteristic chromatogram of GBE and literature. Then, the content differences of three different oral GBE preparations were studied, and their pharmacokinetics (PK) were compared. Finally, the PK differences of the preparations with better oral absorption under different dietary conditions were studied. RESULTS: Five flavonoid glycosides, three aglycones and four terpene lactones were selected as the research objects. The content determination results of GBE tablets, guttate pills and tinctures showed that the content of several components especially flavonoid glycosides in the tincture was higher than that of the other two preparations. After oral administration of these three preparations, the PK study showed different results from previous studies. The PK behavior of flavonoid glycosides was also determined at the same time as flavonoid glycosides and terpene lactones. and the bioavailability of flavonoid glycosides in the tincture was higher than that of the other two preparations. PK results of fasting and non-fasting showed that taking GBE tincture on an empty stomach increased the absorption of various compounds, especially flavonoid glycosides. However, due to the existence of food residues in the gastrointestinal tract, the oral bioavailability of flavonoid glycosides was significantly improved. CONCLUSIONS: This study discussed the differences in the content and oral absorption of active compounds in different oral preparations of GBE, clarified the in vivo absorption of flavonoid glycosides prototype, as well as the influence of diet on the PK of active compounds, which has certain guiding significance for the clinical application of GBE oral preparations.

Inhibitory effect of Ginkgo biloba seeds peptides on methylglyoxal-induced glycations.

The aim of this study was to investigate the antiglycation activity and mechanism of two identified peptides, Valine-Valine-Phenylalanine-Proline-Glycine-Cysteine-Proline-Glutamic acid (VVFPGCPE) and Serine-Valine-Aspartic acid-Aspartic acid-Proline-Arginine-Threonine-Lysine (SVDDPRTL), from Ginkgo biloba seeds protein hydrolysates. Both VVFPGCPE and SVDDPRTL were efficient in bovine serum albumin (BSA)-methylglyoxal (MGO) model to inhibit BSA glycation, while VVFPGCPE showed higher antiglycation activity than SVDDPRTL. In antioxidant assays, VVFPGCPE scavenged more hydroxyl and super anion radicals, and chelated more Fe2+. Moreover, VVFPGCPE was more efficient in alleviating glycoxidation since it retained higher content of tryptophan and reduced dityrosine and kynurenine generation. Compared with SVDDPRTL, VVFPGCPE showed better performance in inhibiting protein aggregation and amyloid-like fibrillation formation. Therefore, VVFPGCPE was selected for further mechanism study. The circular dichroism analysis suggested VVFPGCPE could preserve α-helix structure and stabilize protein structure. The MGO trapping assay indicated VVFPGCPE (5 mg/mL) could capture 66.25% MGO within 24 h, and the mass spectrometry revealed VVFPGCPE could trap MGO by forming VVFPGCPE-mono-MGO adducts. Besides, molecular simulations suggested VVFPGCPE could interact with key glycation residues, arginine and lysine residues, of BSA mainly through van der Waals and hydrogen bonds. This study might supply a theoretical basis for the development of VVFPGCPE as an effective antiglycation agent.

Purification of total flavonoids from Ginkgo biloba flowers with resin column chromatography and evaluation of antioxidant activities in vitro.

Purification of total flavonoids from Ginkgo biloba flowers (GBF) extracts were studied using six resins. Adsorption-desorption experiments indicated that polyamide resin is the most suitable resin. The optimal purification process of total flavonoids of GBF was as follows: a loading concentration of 5.85 mg/mL, a loading volume of 1 bed volume (BV), a loading flow rate of 2 BV/h, a water volume of 2.67 BV, and a desorption solution of 40% ethanol. Under these conditions, the maximum purity of total flavonoids was 37.1 ± 1.1%. The antioxidant activity of purified flavonoids was further evaluated in vitro. It showed that the 40% ethanol purified fraction (Fr. B) group had the strongest antioxidant activity of the 2, 2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity concentration for 50% of maximal effect (EC50, 145.4 ± 13.8 µg/mL) and ferric reducing ability (2.5 ± 0.2 mM FeSO4 equivalent mg-1 Fr. B). In addition, at the concentration of 160 µg/mL, the Fr. B strikingly increased the viability rate of hydrogen peroxide stimulated PC-12 cells to normal levels (***p < 0.001). This method provides a basis for the application and development of GBF resources. It indicated that the purified GBF flavonoids can be used as a source of potential antioxidant.