Extraction and determination of bioactive flavonoids from Abelmoschus manihot (Linn.) Medicus flowers using deep eutectic solvents coupled with high‐performance liquid chromatography

A highly efficient and ecofriendly extraction method using deep eutectic solvents was developed to extract bioactive flavonoids from Abelmoschus manihot (Linn.) Medicus flowers. First, a series of deep eutectic solvents using choline chloride as hydrogen bond acceptor with different hydrogen bond donors was successfully synthesized. Then, the types of deep eutectic solvents and the extraction conditions for bioactive flavonoids (hyperoside, isoquercitrin, and myricetin) were optimized based on the flavonoids extraction efficiencies. The optimized deep eutectic solvent for hyperoside and isoquercitrin extraction was composed of choline chloride and acetic acid with a molar ratio of 1:2. The optimized deep eutectic solvent for myricetin extraction was composed of one mole of choline chloride and two moles of methacrylic acid. The optimal extraction conditions were set as: solid to solvent ratio, 35:1 (mg/mL); extraction time, 30 min; extraction temperature, 30°C. Qualitative and quantitative analysis were performed using ultra high performance liquid chromatography with tandem mass spectrometry and high‐performance liquid chromatography. And the extraction efficiencies of hyperoside, isoquercitrin, and myricetin under optimal extraction conditions were calculated as 11.57, 5.64, and 1.11 mg/g, much higher than those extracted by traditional extraction solvents. Therefore, the prepared deep eutectic solvents can be selected as alternative solvent to extract bioactive flavonoids.     

A New Flavone Glycoside from Isodon enanderianus

Isodon enanderianus (Hand.-Mazz.) H. W. Li, a perennial shrub plant of Labiatae family, is widely distributed in the southern part of Yunnan province. It has long been used as folk medicine to diminish inflammation and detoxify1. The Isodon genus is known to be rich in ent-kaurane diterpenoids, a series of new ent-kaurane diterpenoids have been isolated from the dried leaves of I. enanderianus2-4. During the course on a re-investigation of the chemical constituents of I. enanderianus, four…     

Antioxidant product analysis of Folium Hibisci Mutabilis

Folium Hibisci Mutabilis, a new member of Chinese Pharmacopoeia, can treat some diseases induced by reactive oxygen species. The study prepared a lyophilized aqueous extract of Folium Hibisci Mutabilis (LAFHM). LAFHM was found to enrich eight flavonoids (i.e., quercetin, luteolin, hyperoside, isoquercitrin, rutin, kaempferol, tiliroside, and vitexin) by HPLC analysis. These flavonoids were further compared using antioxidant assays, where triliroside and vitexin always exhibited higher IC₅₀ values than the others. In ultra-performance liquid chromatography coupled with electrospray ionization quadrupole time-of-flight tandem mass spectrometry analysis, these flavonoids could basically give two characteristic m/z values (226 and 196) and their corresponding double m/z values (i.e., 602, 570, 926, 926, 570, 1186, and 862), when treated by 1,1-diphenyl-2-picryl-hydrazl radical (DPPH^?). Finally, the coupling products of DPPH^?-treated triliroside were investigated using computational chemistry. It was found that the –OH in para-coumaroyl moiety to have the lowest bond disassociation energy among all phenolic -OHs in the triliroside. In conclusion, Folium Hibisci Mutabilis contains the above eight antioxidant flavonoids. Despite of the different antioxidant levels, they can generally produce flavonoid-radical coupling product and flavonoid-flavonoid homodimer during antioxidant process. Especially, tiliroside uses para-coumaroyl as linker to construct a tiliroside-radical coupling product at the meta-carbon atom.     

HPLC/DAD,Antibacterial and Antioxidant Activities of Plectranthus Species (Lamiaceae) Combined with the Chemometric Calculations

The increase in antibiotic resistance and the emergence of new bacterial infections have intensified the research for natural products from plants with associated therapy. This study aimed to verify the antibacterial and antioxidant activity of crude extracts of the genus Plectranthus species, being the first report on the modulation of aminoglycosides antibiotic activity by Plectranthus amboinicus extracts. The chemical composition was obtained by chemical prospecting and High-Performance Liquid Chromatography with diode arrangement detector (HPLC/DAD). The antibacterial activities of the extracts alone or in association with aminoglycosides were analyzed using the microdilution test. The antioxidant activity was evaluated by 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical scavenging. The phytochemical prospection allowed the flavonoids, saponins, tannins and triterpenoids to be identified. Quercetin, rutin, gallic acid, chlorogenic acid, caffeic acid, catechin, kaempferol, glycosylated kaempferol, quercitrin, and isoquercitrin were identified and quantified. The principal component analysis (PCA) observed the influence of flavonoids and phenolic acids from Plectranthus species on studied activities. Phytochemical tests with the extracts indicated, especially, the presence of flavonoids, confirmed by quantitative analysis by HPLC. The results revealed antibacterial activities, and synergistic effects combined with aminoglycosides, as well as antioxidant potential, especially for P. ornatus species, with IC₅₀ of 32.21 µg/mL. Multivariate analyzes show that the inclusion of data from the antioxidant and antibacterial activity suggests that the antioxidant effect of these species presents a significant contribution to the synergistic effect of phytoconstituents, especially based on the flavonoid contents. The results of this study suggest the antibacterial activity of Plectranthus extracts, as well as their potential in modifying the resistance of the analyzed aminoglycosides.     

Antioxidant, diuretic activities and polyphenol content of Stereospermum kunthianum Cham. (Bignoniaceae)

Stereospermum kunthianum was used for biological and phytochemical investigations. In biological studies, antioxidant activities were investigated with water, methanol and aqueous acetone extracts. Furthermore, the xanthine oxidase inhibitory activity and the diuretic activity of an aqueous acetone extract were evaluated. In the phytochemical investigations, the flavonoids and polyphenols were quantified spectrophotometrically in all extracts followed by high-performance liquid chromatography-mass spectrometry (HPLC-MS) analysis of an aqueous acetone extract. The 2,2-diphenyl-1-picrylhydrazyl (DPPH), ferric reducing/antioxidant power (FRAP) and 2,2'-azinobis (3-ethylbenzoline-6-sulphonate (ABTS) methods have shown that the aqueous acetone extract presents the best antioxidant activities. This aqueous acetone extract was further proven to have interesting xanthine oxidase inhibitory activity, but only a weak diuretic activity. This aqueous acetone extract also possessed the highest phenolic and flavonoid contents. HPLC-MS analysis allowed identifying and quantifying, rutin, isoquercitrin, quercetin, hyperoside, quercitrin and luteolin and the glycosides of ferulic, sinapic p-coumaric acids and kaempferol, apigenin in aqueous-acetone extract.     

Study of antioxidant properties of flavonoids by voltammetry

Flavonoids are a large group of natural phenolic compounds contained in high concentrations in vegetables, fruits, etc. Antioxidant and redox properties of some flavonoids such as catechin, quercetin, dihydroquercetin, and rutin were investigated in this work. Optimal concentration and time of action of flavonoids were obtained. To determine the more effective range of antioxidant activity, mathematical models and the response surfaces of investigated flavonoids were determined using methods of experiment design. Oxidation potentials of the compounds were also obtained, E?=?0.3?÷?0.4?V. Moreover, the antioxidant activity of flavonoids depends on the redox properties and the structure of the flavonoids. The antioxidant activity of flavonoids, which is correlated to reversible potentials for this compound is good. Finally, the use of these substances as antioxidants with therapeutic effects has been recommended in human diet.     

Development of an HPLC post-column antioxidant assay for Solidago canadensis radical scavengers

The aim of this work was to modify and validate the post-column high-performance liquid chromatography (HPLC)-ABTS and DPPH methods for evaluating the antioxidant activity of the methanolic extracts of Solidago canadensis (Canadian goldenrod) leaves and flowers. Separation of the analytes was performed via the HPLC-PDA method on a YMC analytical column using a gradient elution program. Three compounds with antioxidant properties – chlorogenic acid, rutin and isoquercitrin – and two unidentified antioxidants were established. The research showed that the coil temperature regimes and loop length combinations influence the optimised post-column assay method for detecting the antioxidant activity of goldenrod radical scavengers. Investigations established that the temperature in the reaction coil was a substantial factor contributing to the signal strength of the analytes after reacting with the DPPH and ABTS radicals.     

Chemical Composition,Antioxidant and Anticancer Activities of Leptocarpha rivularis DC Flower Extracts

An evaluation of antioxidant and anticancer activity was screened in Leptocarpha rivularis DC flower extracts using four solvents (n-hexane (Hex), dichloromethane (DCM), ethyl acetate (AcOEt), and ethanol (EtOH)). Extracts were compared for total extract flavonoids and phenol contents, antioxidant activity (2,2-diphenyl-1-picryl-hydrazyl-hydrate (DPPH), ferric reducing antioxidant potential (FRAP), total reactive antioxidant properties (TRAP) and oxygen radical absorbance capacity (ORAC)) across a determined value of reduced/oxidized glutathione (GSH/GSSG), and cell viability (the sulforhodamine B (SRB) assay). The most active extracts were analyzed by chromatographic analysis (GC/MS) and tested for apoptotic pathways. Extracts from Hex, DCM and AcOEt reduced cell viability, caused changes in cell morphology, affected mitochondrial membrane permeability, and induced caspase activation in tumor cell lines HT-29, PC-3, and MCF-7. These effects were generally less pronounced in the HEK-293 cell line (nontumor cells), indicating clear selectivity towards tumor cell lines. We attribute likely extract activity to the presence of sesquiterpene lactones, in combination with other components like steroids and flavonoids.     

Chemical composition and antioxidant activity of Borago officinalis L. leaf extract growing in Algeria

The aqueous and hydroalcoholic extracts of borage (Borago officinalis) leaves from Annaba region (Algeria) were preliminary analyzed for their phenolic profile (total phenolics, total flavonoids, total flavonols, total tannins and total anthocyanins). These extracts were evaluated for their antioxidant properties by different methods such as DPPH radical scavenging, test NBT and total antioxidant activity. The two extracts have exhibited a high antiradical capacity. Indeed, the ethanolic extract showed the lower IC50 values and the highest amount of phenolics (94.09 ± 1.72 mg gallic acid/g dry extract). Using LC-MS/MS analysis, it was possible to identify phenolic acids, flavonoids, sterol and for the first time oleuropein was identified in the aqueous extract of the plant. The obtained results have demonstrated that phenolic compounds are the major contributor to the antioxidant activity of plants.     

4种黄酮小分子对DPPH自由基的清除作用及构效关系研究

通过紫外可见光谱测定了4种黄酮小分子芦丁、牡荆素、山奈素、金丝桃苷对DPPH自由基的清除率、稳定性及半抑制浓度(IC50),并以常用的天然抗氧化剂抗坏血酸作为对照,考察了其抗氧化效果,探讨了黄酮类化合物的抗氧化性与结构的关系。结果表明：不同的抗氧化剂清除DPPH自由基达到平衡的时间不同,芦丁所需时间最长。4种黄酮小分子及抗坏血酸均对DPPH自由基有清除效果,并存在一定的量效关系。对DPPH自由基的清除能力从大到小依次为金丝桃苷、抗坏血酸、芦丁、山奈素、牡荆素。结构分析表明,B环邻二酚羟基是黄酮类化合物抗氧化所必需的基团,其羟甲基化及A环羟基糖苷化不利于黄酮类化合物的抗氧化活性。而C环3-OH的糖苷化对抗氧化活性有利,且单糖苷优于双糖苷。     

Effects of <Emphasis Type="Italic">A.marina</Emphasis>-Derived Isoquercitrin on TNF-Related Apoptosis-Inducing Ligand Receptor (TRAIL-R) Expression and Apoptosis Induction in Cervical Cancer Cells

TNF-related apoptosis-inducing ligand (TRAIL) is an anticancer agent, which has greater apoptosis inducing capacity, but most of the cancer cells become resistant to TRAIL-induced apoptosis. The combined treatment of TRAIL with natural products could restore the cancer cell sensitivity to recombinant human TRAIL (rhTRAIL) protein and might enhance the TNF-related apoptosis-inducing ligand receptor (TRAIL-R) expression. This investigation was aimed to isolate flavonoids from leaves of Avicennia marina and evaluate their potential for sensitization of rhTRAIL in human cervical cancer cells (SiHa). The methanolic extract of A.marina leaves were purified and structure was elucidated as isoquercitrin by NMR and LC-MS analysis. Isolated isoquercitrin showed cytotoxicity against SiHa cell line at IC₅₀ of 980 μM. Messenger RNA (mRNA) expression of TRAIL-Rs was quantified by qRT-PCR, combination of isoquercitrin, and/or rhTRAIL increased TRAIL-R1 and TRAIL-R2 gene expression by 7 folds and 4 folds, respectively. Also, FACS assay revealed that combined treatment has increased the early apoptosis up to 7.24%. In the present study, we found that isoquercitrin enhances the mRNA expression of TRAIL-Rs, but the percentage of apoptosis was meager, possibly due to the influence of other anti-apoptotic proteins.     

Myeloperoxidase inhibitory and radical scavenging activities of flavones from Pterogyne nitens

Two new flavone glucosides, nitensosides A and B (1, 2), together with four known compounds, sorbifolin (3), sorbifolin 6-O-beta-glucopyranoside (4), pedalitin (5), and pedalitin 6-O-beta-glucopyranoside (6) were isolated from Pterogyne nitens. Their structures were elucidated from 1D and 2D NMR analysis, as well as by high resolution mass spectrometry. All the isolated flavones were evaluated for their myeloperoxidase (MPO) inhibitory activity. The most active compound, pedalitin, exhibited IC50 value of 3.75 nM on MPO. Additionally, the radical-scavenging capacity of flavones 1-6 was evaluated towards ABTS and DPPH radicals and compared to standard compounds quercetin and Trolox.     

Evaluation of the redox properties and anti/pro-oxidant effects of selected flavonoids by means of a DNA-based electrochemical biosensor

Quercetin and rutin as well as catechin and epigallocatechin gallate were investigated, as widely distributed representatives of flavonols and flavanols, respectively, regarding their anti/pro-oxidant properties. The flavonoids are irreversibly oxidized at a dsDNA-modified screen-printed electrode within 0.368 to 0.449 V vs. SHE without binding to DNA. Using the DNA biosensor the detection scheme of a DNA prevention/degradation exploits the [Co(phen)(3)](3+) complex as an electrochemical DNA marker. Antioxidant activity of flavonoids was tested in a model cleavage mixture composed of 5 x 10(-7) mol L(-1) [Cu(phen)(2)](2+) as the catalyst, 1 x 10(-3) mol L(-1) ascorbic acid as the chemical reductant and atmospheric oxygen as the natural oxidant where reactive oxygen radicals are generated. The antioxidant activity increases with the concentration of flavonoids reaching a maximum where pro-oxidative behaviour becomes of importance. The pro-oxidant potency of flavonoids depends on the presence of atmospheric oxygen and follows the order quercetin>rutin>epigallocatechin gallate>catechin.     

Electrochemical approach for discriminating and measuring predominant flavonoids and phenolic acids using differential pulse voltammetry: towards an electrochemical index of natural antioxidants

This paper deals with possibilities of employing differential pulse voltammetry (DPV) at a glassy carbon electrode in the analytical characterization of natural phenolic antioxidants in samples containing phenolic classes such as cinnamic acids, flavan-3-ols, and flavonols. A selective electrochemical oxidation of predominant flavonoids at pH 7.5 and an electrochemical oxidation of the total phenolic acids and flavonoids at pH 2.0 was carried out. Our electrochemical approach, based on DPV, has proved to be an easy, fast, and reliable method for discriminating flavonoids and phenolic acids. It has also helped establish the relationship between their voltammetric behavior and their in vitro “antioxidant power”. The analytical possibilities of the proposed method have been explored using both, representative standards and real samples where the predominant flavonoid and phenolic acid classes are present in different concentrations. The new approach has also proved satisfactory for the study of apple and pear juices as well as fresh apples and pears in different matrices such as peels and pulps. In general terms, “high antioxidant power” was always related to predominant phenolic classes (acids and flavan-3-ols) while the “low antioxidant power” was a result of a contribution of predominant phenolics (flavan-3-ols) and fingerprint phenolics: phlorizdin and arbutin. Likewise, the proposed electrochemical strategy showed a satisfactory analytical performance. A reproducibility of oxidation potential (less 5%) as well as in the height of peaks and calibration slopes (with R.S.D.s ranging between 0.3 and 8.3%) were also obtained. Concomitantly, the newly proposed voltammetric method showed recoveries between 83.5 and 108.6%.     

Determination of Polyphenols in Lycium barbarum Leaves by High-Performance Liquid Chromatography–Tandem Mass Spectrometry

High-performance liquid chromatography coupled with high-resolution mass spectrometry was used for the determination of polyphenols in Lycium barbarum leaves. Twenty compounds extracted by methanol–water were tentatively identified that included chlorogenic acids, flavonoids, and phenolic acids. Neochlorogenic acid, chlorogenic acid, cryptochlorogenic acid, isochlorogenic acid B, isochlorogenic acid A, and isochlorogenic acid C were identified in Chinese cultivated L. barbarum leaves for the first time. Caffeic acid and isoquercitrin were also present. The concentrations of these compounds in L. barbarum leaves were determined. The results showed that all analytes had linear calibration relationships with limits of detection from 0.318 to 3.35?ng mL^?1. The polyphenols and flavonoids in L. barbarum leaves provided strong 2,2-diphenyl-1-picrylhydrazyl radical-scavenging capacity (IC₅₀ of 23.1?±?0.4 to 26.0?±?0.4?µg mL^?1). This method is suitable for the determination of polyphenols in L. barbarum leaves, which provide polyphenols with suitable antioxidant activity.     

The in‐capillary DPPH‐capillary electrophoresis‐the diode array detector combined with reversed‐electrode polarity stacking mode for screening and quantifying major antioxidants in Cuscuta chinensis Lam

An in‐capillary 2, 2‐diphenyl‐1‐picrylhydrazyl (DPPH)‐CE‐the DAD (in‐capillary DPPH‐CE‐DAD) combined with reversed‐electrode polarity stacking mode has been developed to screen and quantify the active antioxidant components of Cuscuta chinensis Lam. The operation parameters were optimized with regard to the pH and concentration of buffer solution, SDS, β‐CDs, organic modifier, as well as separation voltage and temperature. Six antioxidants including chlorogenic acid, p‐coumaric acid, rutin, hyperin, isoquercitrin, and astragalin were screened and the total antioxidant activity of the complex matrix was successfully evaluated based on the decreased peak area of DPPH by the established DPPH‐CE‐DAD method. Sensitivity was enhanced under reversed‐electrode polarity stacking mode and 10‐ to 31‐fold of magnitude improvement in detection sensitivity for each analyte was attained. The results demonstrated that the newly established in‐capillary DPPH‐CE‐DAD method combined with reversed‐electrode polarity stacking mode could integrate sample concentration, the oxidizing reaction, separation, and detection into one capillary to fully automate the system. It was considered a suitable technique for the separation, screening, and determination of trace antioxidants in natural products.     

Chemical constituents of roots of Epimedium wushanense and evaluation of their biological activities

Seven flavonoids named diphylloside A, epimedoside A, epimedin C, icariin, epimedoside C, icarisoside A, desmethylanhydroicaritin, as well as the oleanolic acid, were isolated from the roots of Epimedium wushanense for the first time. These flavonoids manifested significant antioxidant activity in vitro. Scavenging effects of two flavonoids were comparable to that of Vitamin C. Antibacterial experiment has shown that the diphylloside A, icarisoside A and desmethylanhydroicaritin have significant activity towards Pseudomonas aeruginosa.     

千里光中几种黄酮和酚酸类成分的分离与鉴定

对千里光(Senecio scandens Buch.-Ham.)全草的乙酸乙酯提取部位进一步分离,得到4个黄酮类化合物和2个酚酸类化合物,经理化性质和NMR及MS谱学数据鉴定分别为槲皮素3-O-β-D-葡萄糖苷（Ⅰ）、金丝桃苷（Ⅱ）、槲皮素（Ⅴ）、异鼠李素（Ⅵ）、绿原酸（Ⅲ）和咖啡酸（Ⅳ）。 其中化合物Ⅰ~Ⅵ均为首次从千里光属植物中分离鉴定。     

Enhanced Extraction Efficiency of Flavonoids from Pyrus ussuriensis Leaves with Deep Eutectic Solvents

In this study, deep eutectic solvents (DESs) were synthesized using different ratios of choline chloride (CC) and dicarboxylic acids, and their eutectic temperatures were determined. The DES synthesized using CC and glutaric acid (GA), which showed a higher extraction efficiency than conventional solvents, was used for the extraction of flavonoid components from Pyrus ussuriensis leaves (PUL), and the extraction efficiency was evaluated using the response surface methodology. The flavonoid components rutin, hyperoside, and isoquercitrin were identified through high-performance liquid chromatography (HPLC), equipped with a Waters 2996 PDA detector, and HPLC mass spectrometry (LC-MS/MS) analyses. The optimum extraction was achieved at a temperature of 30 °C using DES in a concentration of 30.85 wt.% at a stirring speed of 1113 rpm and an extraction time of 1 h. The corresponding flavonoid content was 217.56 μg/mL. The results were verified by performing three reproducibility experiments, and a high significance, with a confidence range of 95%, was achieved. In addition, the PUL extracts exhibited appreciable antioxidant activity. The results showed that the extraction process using the DES based on CC and GA in a 1:1 molar ratio could effectively improve the yield of flavonoids from PUL.