Synthesis,characterization and biological screening studies of mixed ligand complexes using flavonoids as precursors

Flavonoids are a group of plant phenolics that provide various health benefits through cell signalling pathways and antioxidant effects. In the present study, a new series of mixed ligand complexes of Co(II), Ni(II), Cu(II) and Zn(II) were synthesized by incorporating curcumin and quercetin flavonoid precursors. The structural features of the synthesized complexes were explored using elemental analysis, thermogravimetric analysis, UV–visible, infrared, NMR, mass and electron paramagnetic resonance spectral analyses and conductivity measurements. These data support an octahedral geometry of the synthesized complexes. In silico biological activity score for the ligand was predicted using PASS online software. ADMET properties were studied using VLS3D online software. Anti‐inflammatory and antioxidant activities were experimentally validated which prove that theoretical predictions are in agreement with the experimental results. Interestingly the synthesized complexes interact with calf thymus DNA through groove binding mode. Moreover, they have good potential to cleave pUC19 DNA. Minimum inhibitory concentration values of the synthesized complexes reveal that they have better antimicrobial efficacy than the ligands.     

Binding of Androgen- and Estrogen-Like Flavonoids to Their Cognate (Non)Nuclear Receptors: A Comparison by Computational Prediction

Flavonoids are plant bioactives that are recognized as hormone-like polyphenols because of their similarity to the endogenous sex steroids 17β-estradiol and testosterone, and to their estrogen- and androgen-like activity. Most efforts to verify flavonoid binding to nuclear receptors (NRs) and explain their action have been focused on ERα, while less attention has been paid to other nuclear and non-nuclear membrane androgen and estrogen receptors. Here, we investigate six flavonoids (apigenin, genistein, luteolin, naringenin, quercetin, and resveratrol) that are widely present in fruits and vegetables, and often used as replacement therapy in menopause. We performed comparative computational docking simulations to predict their capability of binding nuclear receptors ERα, ERβ, ERRβ, ERRγ, androgen receptor (AR), and its variant AR^T877A and membrane receptors for androgens, i.e., ZIP9, GPRC6A, OXER1, TRPM8, and estrogens, i.e., G Protein-Coupled Estrogen Receptor (GPER). In agreement with data reported in literature, our results suggest that these flavonoids show a relevant degree of complementarity with both estrogen and androgen NR binding sites, likely triggering genomic-mediated effects. It is noteworthy that reliable protein–ligand complexes and estimated interaction energies were also obtained for some suggested estrogen and androgen membrane receptors, indicating that flavonoids could also exert non-genomic actions. Further investigations are needed to clarify flavonoid multiple genomic and non-genomic effects. Caution in their administration could be necessary, until the safe assumption of these natural molecules that are largely present in food is assured.     

Identification of Flavonoids as Putative ROS-1 Kinase Inhibitors Using Pharmacophore Modeling for NSCLC Therapeutics

Non-small cell lung cancer (NSCLC) is a lethal non-immunogenic malignancy and proto-oncogene ROS-1 tyrosine kinase is one of its clinically relevant oncogenic markers. The ROS-1 inhibitor, crizotinib, demonstrated resistance due to the Gly2032Arg mutation. To curtail this resistance, researchers developed lorlatinib against the mutated kinase. In the present study, a receptor-ligand pharmacophore model exploiting the key features of lorlatinib binding with ROS-1 was exploited to identify inhibitors against the wild-type (WT) and the mutant (MT) kinase domain. The developed model was utilized to virtually screen the TimTec flavonoids database and the retrieved drug-like hits were subjected for docking with the WT and MT ROS-1 kinase. A total of 10 flavonoids displayed higher docking scores than lorlatinib. Subsequent molecular dynamics simulations of the acquired flavonoids with WT and MT ROS-1 revealed no steric clashes with the Arg2032 (MT ROS-1). The binding free energy calculations computed via molecular mechanics/Poisson-Boltzmann surface area (MM/PBSA) demonstrated one flavonoid (Hit) with better energy than lorlatinib in binding with WT and MT ROS-1. The Hit compound was observed to bind in the ROS-1 selectivity pocket comprised of residues from the β-3 sheet and DFG-motif. The identified Hit from this investigation could act as a potent WT and MT ROS-1 inhibitor.     

Strategies for differentiation of isobaric flavonoids using liquid chromatography coupled to electrospray ionization mass spectrometry

Flavonoids are a class of secondary plant metabolites existing in great variety in nature. Due to this variety, identification can be difficult, especially as overlapping compounds in both chromatographic separations and mass spectrometric detection are common. Methods for distinguishing isobaric flavonoids using MS² and MS³ have been developed. Chromatographic separation of various plant extracts was done with RP‐HPLC and detected with positive ESI‐MS operated in information‐dependent acquisition (IDA) mode. Two methods for the determination of flavonoid identity and substitution pattern, both featuring IDA criteria, were used together with the HPLC equipment. A third method where the collision energy was ramped utilized direct infusion. With the developed strategies, it is possible to differentiate between many isobaric flavonoids. Various classes of flavonoids were found in all of the plant extracts, in the red onion extract 45 components were detected and for 29 of them the aglycone was characterized, while the substituents were tentatively identified for 31 of them. For the strawberry extract, those numbers were 66, 30 and 60, and for the cherry extract 99, 56 and 71. The great variety of flavonoids, several of them isobaric, found in each of the extracts highlights the need for reliable methods for flavonoid characterization. Methods capable of differentiating between most of the isobars analyzed have been developed. Copyright © 2014 John Wiley & Sons, Ltd.     

LC–MS/MS method for simultaneous determination of flavonoids and physalins in rat plasma: Application to pharmacokinetic study after oral administration of Physalis alkekengi var. franchetii (Chinese lantern) extract

A rapid and sensitive liquid chromatography with tandem mass spectrometry (LC‐MS/MS) method was developed and validated for the simultaneous determination of luteolin, luteolin‐7‐O ‐β ‐D‐glucopyranoside, physalin A, physalin D and physalin L in rat plasma. Scutellarein and dexamethasone were used as the internal standards (IS). Plasma samples were prepared by liquid‐liquid extraction with ethyl acetate. The five constituents were separated on an Acquity UPLC BEH C₁₈ column (100 mm × 2.1 mm, 1.7 μm). A gradient elution procedure was used with acetonitrile (A)‐0.1% aqueous formic acid (B). Mass spectrometric detection was performed in negative ion multiple reaction monitoring mode with an electrospray ionization (ESI) source. This method showed good linearity (r ² > 0.997) over a concentration range of 2.0–500 ng/mL with a lower limit of quantification of 2.0 ng/mL for all five compounds. The inter‐ and intra‐day accuracy ranged from 91.7 to 104%, and precisions (RSD) were <6.46% for all analytes. The extraction recoveries of all analytes were >85%. This validated method was successfully applied for the first time to the pharmacokinetic study of five ingredients after oral administration of 70% ethanol extract of Chinese lantern in rats.     

Zinc sulfide nanosheets as a novel solid‐phase extraction material for flavonoids

As a novel solid‐phase extraction material, zinc sulfide nanosheets were prepared by a simple method and were used to extract flavonoids. We used scanning electron microscopy to show its nanosheet morphology and energy dispersive X‐ray spectroscopy and powder X‐ray diffraction to confirm its chemical and phase compositions. Coupled to a high‐performance liquid chromatography, the zinc sulfide nanosheets were packed into a microcolumn and were used to extract four model flavonoids to examine their extraction ability. The parameters of sample loading and elution were investigated. Under optimized conditions, the analytical method for flavonoids was established. For the method, wide linearities from 1 to 250 μg/L and low limits of detection from 0.25 to 0.5 μg/L were obtained. The relative standard deviations for single column repeatability and column to column reproducibility were less than 7.7 and 10.4%, respectively. The established method was also used to analyze two real samples and the recoveries from 88.7 to 98.2% further proved the reliability of the method. Moreover, the zinc sulfide nanosheets have good stability and that in one column can be reused for more than 50 times. This work proves that the prepared zinc sulfide nanosheets are a good candidate as the flavonoids sorbent.     

Structure‐oriented UHPLC‐LTQ Orbitrap‐based approach as a dereplication strategy for the identification of isoflavonoids from Amphimas pterocarpoides crude extract

Hyphenated techniques and especially ultra‐high performance liquid chromatography‐mass spectrometry (UHPLC‐MS) are nowadays widely employed in natural products research. However, the complex nature of plant extracts complicates considerably the analysis and the identification of their constituents. Nevertheless, new MS analyzers with increased resolving power and accuracy such as the orbital trap (Orbitrap) could facilitate drastically this process. The objective of this study is the development of a new structure‐oriented approach based on fast UHPLC‐high‐resolution (HR)MS and HRMS/MS methodologies for the identification of isoflavonoids in crude extracts. In addition, aims to assist dereplication procedures, to decrease the laborious isolation steps and orient the focused isolation of compounds of interest. As a proof of concept, the methanol extract of the stem bark of Amphimas pterocarpoides (Leguminosae) was selected. Based on chromatographic (retention time, polarity) and spectrometric features (ultraviolet spectra, accurate m/z, proposed elemental composition, ring double bond equivalent, and relative isotopic abundance) as well as HRMS/MS spectra, several isoflavonoids were identified. In order to verify the proposed structures, 11 isoflavonoids were selectively isolated and unambiguously identified using 1&2D nuclear magnetic resonance techniques. Moreover, the isolated isoflavonoids were studied in HRMS/MS level, employing electrospray ionization and atmospheric pressure chemical ionization sources, in both modes. Useful information regarding their fragmentation patterns was obtained, and characteristic diagnostic ions were defined for the identification of methoxylated isoflavones, dihydroisoflavones and 5‐hydroxylated isoflavonoids. Based on the current results, the proposed dereplication strategy was verified and could comprise a novel approach for the analysis of crude extracts in the future not only for isoflavonoids but also for other chemical classes of natural products. Copyright © 2013 John Wiley & Sons, Ltd.     

明日叶黄酮类化合物清除羟基自由基活性研究

为了研究明日叶黄酮类化合物对羟基自由基的清除作用,以明日叶（主要取叶片）为原料,用体积分数为65％乙醇提取明日叶总黄酮,测定其总黄酮含量.通过Fenton反应体系产生羟基自由基,利用明日叶提取液中的功能成分黄酮类化合物对羟基自由基的清除作用进行研究.结果表明：明日叶提取物总黄酮质量分数为10.18％,且黄酮类化合物对羟基自由基有较强清除效力,当提取物总黄酮浓度在0.1～1.0 mg/mL范围内,其与清除率呈正相关.明日叶中黄酮类化合物对羟基自由基有较强清除效力,作为天然抗氧化产品开发具有一定价值.