Effects of ginkgo flavone aglycone on atherosclerosis based on network pharmacology,molecular docking,and in vitro experiments

Background:Ginkgo flavone aglycones(GA),a Ginkgo(Ginkgo biloba)extract,has been proven to have good biological activity in atherosclerosis(AS)treatment.Moreover,its active compounds and the corresponding mechanism for the treatment of AS remain unclear.Methods:To evaluate and identify the potential pharmacological mechanisms of GA in AS treatment,the program Cytoscape was used to generate network mappings of the GA-AS-potential target gene.GO and KEGG enrichment analyses were performed to further investigate the potential mechanism of AS and the pharmacological properties of GA.A molecular docking approach was utilized to determine the GA components that interact with Akt.In vitro experiments were carried out to identify the anti-atherosclerotic effects of GA by targeting Akt.Results:Network pharmacological research determined that the active components of GA(quercetin,kaempferol,and isorhamnetin)correlated with AS target genes such as AKT1,EGFR,SRC,ESR1,PTGS2,MMP9,KDR,GSK3B,APP,and MMP2,respectively.GO enrichment and KEGG analysis showed that PI3K-Akt signaling may play an important role in GA treatment.Molecular docking experiments indicated that quercetin,kaempferol,and isorhamnetin integrate into the binding pockets of the most potentially beneficial GA-AS target protein(Akt).Consequently,cell experiments were conducted to support the anti-atherosclerotic activity of GA on AS by inhibiting the phosphorylation of AKT1 and its downstream signaling molecules,which regulated the proliferation of HASMCs.Conclusion:Our results detailed GA's active ingredients,potential targets,and molecular basis against AS.GA may exert anti-atherosclerotic effects by suppressing Akt phosphorylation and inhibiting the proliferation of HASMCs.It also proposed a viable approach to determining the scientific foundation and therapeutic mechanism of Chinese herbal medicine extracts in disease therapy.

高浓度的staurosporine aglycone激活大鼠肺动脉平滑肌细胞中的ERK1／2

目的有报道表明staurosporine作为一种蛋白激酶C（PKC）的抑制剂，可以在多种细胞系内调节细胞外信号调节激酶（ERK1／2）的活性，但是它的衍生物staurosporineagly—COfle（SA）是否具有相同的作用还不清楚，本次实验旨在阐明其对ERK1／2活性的调节作用。方法培养至4～8代的大鼠肺动脉平滑肌细胞，经过SA处理后提取细胞蛋白，应用Westernblot方法检测磷酸化ERK1／2的含量，观察不同浓度和时间点的SA对ERK1／2活性的影响。结果在纳摩尔浓度水平的SA可以降低大鼠肺动脉平滑肌细胞中ERK1／2的磷酸化水平，但是30μmol·L-1的SA可以激活ERK1／2，这种激活作用可以被丝裂素活化蛋白激酶的激酶（MEK）的抑制剂PD98059或蛋白激酶A（PKA）的激活剂异丙肾上腺素（isoproteren01）所抑制。结论sA对肺动脉平滑肌细胞中的ERK1／2活性有双向调节作用，这种作用是通过PKC和（或）PKA通路产生的。

Minor secoiridoid aglycones from the low-polarity part of the traditional Chinese herb: Swertia mileensis

Eleven new secoiridoid aglycones involving unusual C9-skeleton:swerimilegenins A-F(1-6);bis-C9-skeleton:swerimilegenin G(7);and C_(10)-skeleton:swerimilegenins H−K(8−11),as well as six known ones,were isolated from the low-polarity part of the traditional Chinese herb medicine Swertia mileensis.Their structures were determined by extensive spectroscopic data and X-ray diffraction.Biogenetically,swerimilegenin A(1)belonged to 10-nor-secoiridoid,and swerimilegenins B-F(2-6)were 1-nor-secoiridoids.Erythrocentaurin(12)and gentiogenal(15)showed moderate anti-HBV activity on HepG 2.2.15 cell line in vitro.

Synthesis of New 13-O-Acylavermectin Bl Aglycones

Six new 13-O-acylavermectin Bl aglycones(3-8) were synthesized from avermectin B1 aglycone and their bioactivities were evaluated against Spodoptera exigua, Spodoptera eridania, Tetranychus urticae and Aphis fabae.

Insignin A, A Novel C_21-Steroidal Aglycone from Biondia insignis

Insignin A, a new C21-steroidal aglycone having the rare 15,16-seco pregnane skeleton was isolated from the acidic hydrolysis part of the 95% EtOH extract of Biondia insignis. It's structure was identified to be 15R,16-epoxy-3?,14?,16?-trihydroxy-15,16-secopregn-5-ene-20-one based on the spectral data.

The constitutions of terpenoids and hormones of leaves in red-flesh citrus mutants and their wild types

To study the profiles and relationships of terpenoids and phytohormones in leaves of red-flesh citrus,we examined four red-flesh mutants that accumulate lycopene in the fruit and their corresponding wild types.We compared their contents of monoterpenoid and sesquiterpenoid volatiles,triterpenoid limonoid aglycones,and tetraterpenoid carotenoids.The types and concentrations of these terpenoids in the leaves varied among genotypes,and the leaf terpenoid profiles differed between red-fleshmutants and theirwild types.However,lycopenewas not detected in citrus leaves of all eight varieties,including the fourmutants.According to phytohormone accumulation in the leaves,the citrus varieties could be classified into a low-phytohormone group(‘Red Anliu’sweet orange,‘Anliu’sweet orange,and‘Cara Cara’navel orange),a high-phytohormone group(‘Seike’navel orange),and a high-jasmonic acid group(‘Red-flesh Guanxi’pummelo,‘Guanxi’pummelo,‘Chuhong’pummelo,and‘Feicui’pummelo).The contents of terpenoid volatiles were very low in leaves of‘Red-flesh Guanxi’and‘Guanxi’pummelo;therefore,they are ideal materials for verifying the functions of genes related to terpenoid volatiles in overexpression analyses.Sesquiterpenoid volatiles were positively correlated with phytohormones of abscisic acid,jasmonic acid,and salicylic acid.Taken together,lycopene was undetected in the leaves of redflesh citrus mutants with abundant lycopene in their juice sacs,which suggested a tissue-specific accumulation pattern of lycopene in citrus red-flesh mutants.Furthermore,leaf volatile profiles could be suitable to develop fingerprint chromatograms of citrus resources,since leaf volatile profiles were dominated by several compounds that varied among genotypes in their combinations and concentrations.

Phytoestrogen Enriched Tofu from Soybean Meal

Isoflavone, a group of phytoestrogen, reduces postmenopausal symptoms and the risk of osteoporosis of women. Glycosidic forms of isoflavones are presented in non-fermented soyfoods such as tofu and they are less bioavailable than the aglycone isoflavones. Aglycone forms of isoflavones or more bioavailable forms can be increased by acid hydrolysis during tofu processing. The present study investigated the possibility of increasing the aglycone forms of isoflavones by acid hydrolysis. We used five types of tofu in this study: soybean tofu with hydrolysis, soybean meal tofu with hydrolysis, soybean tofu in general process, soybean meal tofu in general process, and commercial tofu. Defatted soybean meal was used as the major ingredient in the tofu which was made by using the new method—acid hydrolysis. To identify the isoflavone quantities in all five types of tofu, high performance liquid chromatography with diode array detection (HPLC-DAD) analysis was employed. The genistein ratio between hydrolyzed tofu and standard tofu was 1:1-8, and the daidzein ratio between hydrolyzed tofu and standard tofu was 1:6-12. The five types of tofu were analyzed for the crude protein and micronutrients such as calcium (Ca), magnesium (Mg), potassium (K), iron (Fe), zinc (Zn), and selenium (Se) by the modified Kjeldahl method and inductively coupled plasma emission spectroscopy (ICP-ES), respectively. The mean crude protein concentration of hydrolyzed tofu from soybean meal was 40.8%. In addition, especially the hydrolyzed soybean meal tofu showed the higher concentration of Ca (27,307 mg/kg) and K (25,553 mg/kg). By and large, soybean meal tofu with acid hydrolysis is a rich source of isoflavone aglycone compared with other types of tofu.

Synthesis analogues of milbemycin and their bioactivity evaluation

Eight new 13-O-aminocarbonylivermectin aglycones and 4＇-O-aminocarbonylivermectin monosaccharide were synthesized from ivermectin agiycone and ivermectin monosaccharide by the selective protection of C5-OH group.Their bioactivities were evaluated against spider mites（Tetranychus cinnabarimts）,aphid（Aphis fabae） and oriental armyworm（Mytliimma sepatara）. Their structures were confirmed by ^1H NMR.MS.

Metabolite Profile of Lactic Acid-Fermented Soymilk

Isoflavone aglycone-rich soymilk fermented by Lactobacillus delbrueckii subsp. delbrueckii, strain TUA-4408L (F4408) has stronger lipid metabolism-modulating effects than isoflavone aglycone-poor soymilk fermented by Lactobacillus delbrueckii subsp. delbrueckii, strain TUA-4404L (F4404). As such, fermentation products other than isoflavone aglycones may also exert lipid metabolism-modulating effects. Therefore, the present study aimed to compare the metabolites in soymilk (SM), F4404, and F4408 via a metabolomics approach. Various aglycones including isoflavones, flavones, flavonols, and flavanones increased in both F4404 and F4408 compared with SM. The increases in isoflavone and flavanone aglycones were greater in F4408 than in F4404. Some types of dipeptides and free amino acids, especially ornithine, increased in both fermented soymilks. The increase in free amino acids, especially ornithine, was higher in F4408 than in F4404. Thus, F4408 exhibited stronger glycosidase and protease activities than F4404. Functional components produced by lactic acid fermentation are known to improve lipid metabolism. Therefore, it is suggested that not only isoflavone aglycones but also other functional components exert lipid metabolism-modulating effects in fermented soymilks, especially soymilk fermented using TUA-4408L.

Hydrolysis of soy isoflavone conjugates using enzyme may underestimate isoflavone concentrations in tissue

Objective:To investigate the differences of using enzymatic hydrolysis and acid hydrolysis for identification and quantification of isoflavone aglycones from biomatrices. Methods: β-glucuronidase/sulfatase isolated from Helix pomatia for routine enzymatic hydrolysis or 6N HCl was used to release glucuronide and sulfate conjugates in the serum, urine and tissue samples. Profiles of soy isoflavones after enzymatic hydrolysis or acid hydrolysis in several tissues of rat fed with diets containing soy protein isolate were also compared using LC/MS and HPLC-ECD. Results: Acid hydrolysis released more aglycone than enzymatic digestion (P<0.05) in liver tissue. The total genistein, daidzein and other metabolites were 20% to 60% lower in samples from enzymatic hydrolysis than in acid hydrolysis. Conclusion: These results indicated that unknown factors in tissues reduced the enzymatic hydrolytic efficiency for releasing isoflavone aglycones even in optimized condition. This would underestimate isoflavone tissue concentrations up to 60%.

Glycosidically Bound Volatile Compounds of Satureja montana L.,S.cuneifolia Ten.,S.subspicata Vis.and Endemic S.visianii SiliC

In this paper, the glycoconjugated volatile compounds of four Croatian Satureja species (Satureja montana L., S. cuneifolia Ten., S. subspicata Vis. and endemic S. visianii Silic) were investigated. Content and composition of these compounds were examined depending on the stage of plant development. GC and GC–MS analysis of volatile aglycones revealed twenty-one compounds. Thymoquinone, geraniol and carvacrol were detected in all vegetative phases of the investigated plants. Other quantitatively important aglycones were eugenol and thymol of S. montana, phenyl ethyl alcohol, benzene acetaldehyde, borneol, α-terpineol, thymol and eugenol of S. cuneifolia, phenyl ethyl alcohol, benzene acet-aldehyde, terpinen-4-ol, α-terpineol and β-ionone of S. subspicata and camphor, thymol and 8a-acetoxylemolol of S. visianii. Moderate similarity in the chemical composition of essential oils and volatile aglycones of investigated plant species indicate that many biologically active compounds are glycosylated and accumulate as non-volatile glycosides.

Synthesis of pennogenin utilizing the intact skeleton of diosgenin

The first synthesis of pennogenin, an aglycone of bioactive components of Chinese traditional medicine named Chonglou(Paris), starting from diosgenin, has been reported, which displays a new strategy of utilizing the resource compounds. According to this new strategy, the full and rational utilization of the intact skeleton and functional groups of starting material has been realized in the conversion of diosgenin to pennogenin. The key step for synthesis of pen-nogenin is the regioselective transformation of cholest-5-en-16,22-dion-3,26-diol to cholest- 5,16-dien-22-on-3,26-diol, which can be used to synthesize other bioactive steroids such as cephalostatin and OSW-1.

Virtual Evaluation on Activities of Flavonoids from Scutellaria baicalensis

Objective To explore the investigation method of complicated and profound traditional Chinese herbal medicine,the potential action mechanisms of flavonoids from Scutellaria baicalensis were studied by docking calculation.Methods In total,eight flavonoids(aglycones and their glicosides) from S.baicalensis were selected as ligands.The crystalline structures of targets related to common diseases were used as the receptors for calculation.The calculations were conducted with Schrdinger software package.The grading standard of selectivity was developed according to G-score between ligands and receptors.Results Twenty-six pharmacologic actions have been reported.Among all effects in literature,nine of them can be deduced from the docking calculation of aglycone.From glycosides with grade ++,25 reported effects can be estimated by calculation.Apparently,the target selectivity of aglycones and their glycosides are different form the virtual evaluation.The virtual evaluation results of glycosides were closer to the reported effects.Conclusion Our proposed virtual evaluation method seems an effective way to investigate the complicated system of traditional Chinese herbal medicine.It suggests that aglycones may be effective as the form of glucoside in vivo,and metabolism is a very important factor for virtual evaluation.

A New Phenylpropanol Glycoside from Twigs and Leaves of Rhododendron primulaeflorum

Objective To study the constituents in the twigs and leaves of Rhododendron primulaeflorum. Methods The constituents were separated and purified with chromatographic methods. Their structures were elucidated by spectroscopic methods (1D, 2D NMR, IR, and HR-ESI-MS) and chemical analyses. Results One new phenylpropanol glycoside, 4-hydroxyl-5-methoxyl-phenylpropanol-3-O-β-D-glucopyranoside (1), and its aglycone (2) were successfully isolated from the twigs and leaves of R. primulaeflorum. Conclusion Compound 1 is a new phenylpropanol glycoside. Compounds 1 and 2 are isolated from this plant for the first time.

Antihyperuricemic effect of mangiferin aglycon derivative J99745 by inhibiting xanthine oxidase activity and urate transporter 1 expression in mice

A mangiferin aglycon derivative J99745 has been identified as a potent xanthine oxidase(XOD) inhibitor by previous in vitro study. This study aimed to evaluate the hypouricemic effects of J99745 in experimental hyperuricemia mice, and explore the underlying mechanisms. Mice were orally administered 600 mg/kg xanthine once daily for 7 days and intraperitoneally injected 250 mg/kg oxonic acid on the 7 th day to induce hyperuricemia. Meanwhile, J99745(3, 10, and 30 mg/kg), allopurinol(20 mg/kg) or benzbromarone(20 mg/kg) were orally administered to mice for 7 days. On the 7 th day,uric acid and creatinine in serum and urine, blood urea nitrogen(BUN), malondialdehyde(MDA) content and XOD activities in serum and liver were determined. Morphological changes in kidney were observed using hematoxylin and eosin(H&E) staining. Hepatic XOD, renal urate transporter 1(URAT1), glucose transporter type 9(GLUT9), organic anion transporter 1(OAT1) and ATP-binding cassette transporter G2(ABCG2) were detected by Western blot and real time polymerase chain reaction(PCR). The results showed that J99745 at doses of 10 and 30 mg/kg significantly reduced serum urate, and enhanced fractional excretion of uric acid(FEUA). H&E staining confirmed that J99745 provided greater nephroprotective effects than allopurinol and benzbromarone. Moreover, serum and hepatic XOD activities and renal URAT1 expression declined in J99745-treated hyperuricemia mice. In consistence with the ability to inhibit XOD, J99745 lowered serum MDA content in hyperuricemia mice. Our resultssuggest that J99745 exerts urate-lowering effect by inhibiting XOD activity and URAT1 expression, thus representing a promising candidate as an anti-hyperuricemia agent.

Synthesis,bioactivities and 3D-QSAR of novel avermectin B2a aglycon derivatives

Fourteen avermectin B2 a aglycon derivatives were designed and synthesized after removing the oleandrose disaccharide of avermectin B2 a.Their structures were characterized by’H NMR,13 C NMR,HMRS.Preliminary bioassays indicated that these compounds exhibited good insecticidal activity against diamondback moth at 200 mg/L,with mortality no less than 90%.Compounds 10 b,12 a,12 c,17 demonstrated good acaricidal activity against the adult mites,larvae,and good inhibition rate of hatching to mite eggs of Tetranychus cinnabarinus.Compounds 5,10 b,10 c exhibited excellent fungicidal activity against fourteen fungal pathogens in vitro.3 D-QSAR analysis showed that the fungicidal activity of avermectin B2 a aglycon derivatives would be increased when a negatively charged and bulky group was introduced at 13-position,which will be instructive for the further modification of avermectin B2 aaglycon.

Analysis of Flavonoid Aglycons in Rhododendron of Sichuan Liangshan

An aqueous solution of the extracts from Rhododendron leaves from Sichuan Liangshan was extracted with solvents of increasing polarity, petroleum ether, ethyl ether, ethyl acetate and butanol. The main flavonoid aglycons extracted into the ethyl ether phase are the subject of this study. The flavonoid aglycons—mycetin, quercetin, kaempferol and farrerol, were separated and identified by thin layer chromatography (TLC), high performance liquid chromatography (HPLC) and liquid chromatography/mass spectrometer/mass spectrometer (LC/MS/MS). A simple and rapid HPLC method was developed for quantitative determination of quercetin and kaempferol in Rhododendron leaves, ethanol extracts and Jinjuan oral liquid drug (Liquor Jinjuan). The analysis of quercetin is useful for quality control of medicinal materials and Liquor Jinjuan products.