Investigation of bacterial neuraminidase inhibition of xanthones bearing geranyl and prenyl groups from Cratoxylum cochinchinense.

Introduction: The root of Cratoxylum cochinchinense has been widely used as Chinese folk medicine to cure fevers, burns, and abdominal complications because it contains various bioactive metabolites such as xanthones, triterpenes, and flavonoids. In this study, we estimated bacterial neuraminidase inhibition with a series of xanthones from C. cochinchinense. BNA has connected to various biological functions such as pathogenic bacteria infection inflammatory process after infection and biofilm formation. Methods: The identification of xanthones (1-6) bearing geranyl and prenyl groups was established by spectroscopic data using UV, IR, NMR, and HREIMS. BNA inhibitory modes of isolated xanthones were investigated by Double-reciprocal plots. Moreover, the competitive inhibitor was evaluated the additional kinetic modes determined by kinetic parameters (k 3, k 4, and K i app). The molecular docking (MD) and molecular dynamics simulations (MDS) studies also provided the critical information regarding the role of the geranyl and prenyl groups against BNA inhibition. Results: A series of xanthones (1-6) appended prenyl and geranyl groups on the A-ring were isolated, and compounds 1-3 were shown to be new xanthones. The analogues within this series were highly inhibited with excellent affinity against bacterial neuraminidase (BNA). A subtle change in the prenyl or geranyl motif affected the inhibitory potency and behavior significantly. For example, the inhibitory potency and binding affinity resulting from the geranyl group on C4: xanthone 1 (IC50 = 0.38 µM, KA = 2.4434 × 105 L·mol-1) were 100-fold different from those of xanthone 3 (IC50 = 35.8 µM, KA = 0.0002 × 105 L·mol-1). The most potent compound 1 was identified as a competitive inhibitor which interacted with BNA under reversible slow-binding inhibition: K i app = 0.1440 µM, k 3 = 0.1410 µM-1s-1, and k 4 = 0.0203 min-1. The inhibitory potencies (IC50) were doubly confirmed by the binding affinities (KA). Discussion: This study suggests the potential of xanthones derived from C. cochinchinense as promising candidates for developing novel BNA inhibitors. Further research and exploration of these xanthones may contribute to the development of effective treatments for bacterial infections and inflammatory processes associated with BNA activity.

Inhibitory Potential of Quercetin Derivatives Isolated from the Aerial Parts of Siegesbeckia pubescens Makino against Bacterial Neuraminidase.

This study aimed to isolate bacterial neuraminidase (BNA) inhibitory O-methylated quercetin derivatives from the aerial parts of S. pubescens. All the isolated compounds were identified as O-methylated quercetin (1-4), which were exhibited to be noncompetitive inhibitors against BNA, with IC50 ranging from 14.0 to 84.1 µM. The responsible compounds (1-4) showed a significant correlation between BNA inhibitory effects and the number of O-methyl groups on quercetin; mono (1, IC50 = 14.0 µM) > di (2 and 3, IC50 = 24.3 and 25.8 µM) > tri (4, IC50 = 84.1 µM). In addition, the binding affinities between BNA and inhibitors (1-4) were also examined by fluorescence quenching effect with the related constants (KSV, KA, and n). The most active inhibitor 1 possessed a KSV with 0.0252 × 105 L mol-1. Furthermore, the relative distribution of BNA inhibitory O-methylated quercetins (1-4) in S. pubescens extract was evaluated using LC-Q-TOF/MS analysis.

1-Aminocyclopropane-1-carboxylic Acid Enhances Phytoestrogen Accumulation in Soy Plants (Glycine max L.) by Its Acceleration of the Isoflavone Biosynthetic Pathway.

The low levels of bioactive metabolites in target plants present a bottleneck for the functional food industry. The major disadvantage of soy leaves is their low phytoestrogen content despite the fact that these leaves are an enriched source of flavonols. Our study demonstrated that simple foliar spraying with 1-aminocyclopropane-1-carboxylic acid (ACC) significantly enhanced the phytoestrogen contents of the whole soy plant, including its leaves (27-fold), stalks (3-fold), and roots (4-fold). In particular, ACC continued to accelerate the biosynthesis pathway of isoflavones in the leaves for up to 3 days after treatment, from 580 to 15,439 µg/g. The detailed changes in the levels of this metabolite in soy leaves are disclosed by quantitative and metabolomic analyses based on HPLC and UPLC-ESI-TOF/MS. The PLS-DA score plot, S-plot, and heatmap provide comprehensive evidence to clearly distinguish the effect of ACC treatment. ACC was also proved to activate a series of structural genes (CHS, CHR, CHI, IFS, HID, IF7GT, and IF7MaT) along the isoflavone biosynthesis pathway time-dependently. In particular, ACC oxidase genes were turned on 12 h after ACC treatment, which was rationalized to start activating the synthetic pathway of isoflavones.

Isoflavone-Enriched Soybean Leaves (Glycine Max) Alleviate Cognitive Impairment Induced by Ovariectomy and Modulate PI3K/Akt Signaling in the Hippocampus of C57BL6 Mice.

(1) Background: The estrogen decline during perimenopause can induce various disorders, including cognitive impairment. Phytoestrogens, such as isoflavones, lignans, and coumestans, have been tried as a popular alternative to avoid the side effects of conventional hormone replacement therapy, but their exact mechanisms and risk are not fully elucidated. In this study, we investigated the effects of isoflavone-enriched soybean leaves (IESLs) on the cognitive impairment induced by ovariectomy in female mice. (2) Methods: Ovariectomy was performed at 9 weeks of age to mimic menopausal women, and the behavior tests for cognition were conducted 15 weeks after the first administration. IESLs were administered for 18 weeks. (3) Results: The present study showed the effects of IESLs on the cognitive function in the OVX (ovariectomized) mice. Ovariectomy markedly increased the body weight and fat accumulation in the liver and perirenal fat, but IESL treatment significantly inhibited them. In the behavioral tests, ovariectomy impaired cognitive functions, but administration of IESLs restored it. In addition, in the OVX mice, administration of IESLs restored decreased estrogen receptor (ER) ß and PI3K/Akt expression in the hippocampus. (4) Conclusions: The positive effects of IESLs on cognitive functions may be closely related to the ER-mediated PI3/Akt signaling pathway in the hippocampus.

Amorphigenin from Amorpha fruticosa L. Root Extract Induces Autophagy-Mediated Melanosome Degradation in mTOR-Independent- and AMPK-Dependent Manner.

In this study, we investigated the depigmentation effect of Amorpha fruticosa L. root extract (RE), an herbal medicine. A. fruticosa RE significantly induced depigmentation in α-MSH-treated B16F10 cells at noncytotoxic concentrations. Further, the RE decreased the protein levels of the melanosomal proteins Tyr and Pmel without decreasing their transcript levels. We found that MG132, a proteasome complex inhibitor, was unable to rescue the protein levels, but PepA/E-64D (a lysosomal enzyme inhibitor), 3-MA (a representative autophagy inhibitor), and ATG5 knockdown effectively rescued the protein levels and inhibited the depigmentation effect following RE treatment. Among rotenoids, amorphigenin composed in the RE was identified as a functional chemical that could induce depigmentation; whereas rapamycin, an mTOR inhibitor and a nonselective autophagy inducer, could not induce depigmentation, and amorphigenin effectively induced depigmentation through the degradation of melanosomal proteins. Amorphigenin activated AMPK without affecting mTOR, and knockdown of AMPK offset the whitening effect through degradation of melanosome proteins by amorphigenin. Results from this study suggested that amorphigenin can induce degradation of the melanosome through an AMPK-dependent autophagy process, and has the potential to be used as a depigmentation agent for the treatment of hyperpigmentation.

Soybean phytochemicals responsible for bacterial neuraminidase inhibition and their characterization by UPLC-ESI-TOF/MS.

Ethanol extract of soybean (Glycine max (L.) Merr.) showed good inhibitory activity against bacterial neuraminidase (BNA), which plays a pivotal role in the pathogenesis of a number of microbial diseases. The saponin portion fractionated through preparative HPLC (IC50 = 2.25 µg mL-1) was found to be responsible for the observed BNA inhibition. Estimation of the inhibitory effects by individual compounds showed that the soyasaponins of group B (Ba, Bb, Bb', Bc, and Bd) exhibited extremely high inhibitions (IC50 = 0.25-0.48 µM), whereas group A (Aa, Ab, and Ac) was almost inactive. Kinetic studies determined that group B soyasaponins were noncompetitive inhibitors. Furthermore, molecular docking experiments confirmed that soyasaponin Ba (group B) could undergo binding interactions with various residues in the binding pocket. In contrast, soyasaponin Aa (group A) failed to enter the binding pocket due to its extra scaffold structure of oligosaccharides bonded to the 22-hydroxyl position. The metabolites in the soybean extract were fully characterized using UPLC-ESI-TOF/MS.

New dihydrobenzoxanthone derivatives with bacterial neuraminidase inhibitory activity isolated from Artocarpus elasticus.

Artocarpus elasticus is a popular fruit tree in the tropical regions. Primary screenings of methanol extracts of the root bark confirmed its potent inhibition of bacterial neuraminidase (BNA), which plays an essential role in the pathogenesis of many microbial diseases. Assessments of the responsible phytochemicals were conducted by isolating eight compounds (1-8) and two of them (6 and 8) were identified as new compounds. Among the isolates, the dihydrobenzoxanthones attained the highest BNA inhibition with IC50 values of 0.5 âˆ¼ 3.9 µM. Further investigation of the inhibitory mechanism by Lineweaver-Burk plots revealed the phytochemicals to function as reversible noncompetitive inhibitors. Fluorescence quenching showed their binding affinities were highly correlated with their inhibitory potential dose-dependently. Molecular docking experiments suggested the dihydrobenzoxanthones (4 and 6) as noncompetitive inhibitors of BNA with unique interaction with Tyr435 of BNA in comparison with the mother flavonoid (7).

Iridal-Type Triterpenoids Displaying Human Neutrophil Elastase Inhibition and Anti-Inflammatory Effects from Belamcanda chinensis.

The aim of this study is to explore anti-inflammatory phytochemicals from B. chinensis based on the inhibition of pro-inflammatory enzyme, human neutrophil elastase (HNE) and anti-inflammatory activities in lipopolysaccharide (LPS)-stimulated RAW264.7 macrophage. Three stereoisomers of iridal-type triterpenoids (1-3) were isolated from the roots of B. chinensis and their stereochemistries were completely identified by NOESY spectra. These compounds were confirmed as reversible noncompetitive inhibitors against HNE with IC50 values of 6.8-27.0 µM. The binding affinity experiment proved that iridal-type triterpenoids had only a single binding site to the HNE enzyme. Among them, isoiridogermanal (1) and iridobelamal A (2) displayed significant anti-inflammatory effects by suppressing the expressions of pro-inflammatory cytokines, such as iNOS, IL-1ß, and TNF-α through the NF-κB pathway in LPS-stimulated RAW264.7 cells. This is the first report that iridal-type triterpenoids are considered responsible phytochemicals for anti-inflammatory effects of B. chinensis.

Inhibitory mechanism of O-methylated quercetins, highly potent ß-secretase inhibitors isolated from Caragana balchaschensis (Kom.) Pojark.

ETHNOPHARMACOLOGICAL RELEVANCE: Caragana has a standing history of implementation in Traditional Chinese Medicine (TCM). Most species of this genus have been explored for multi-functional purposes, such as promoting blood circulation and curing neuralgia, fatigue, migraine, arthritis, and vascular hypertension (Meng et al., 2009). Among them, the well-known species C. sinica showed the most promising potential to increase the expression of ADAM10 among 313 tested medicinal plants, which is one of the promising approach for the treatment of Alzheimer's disease (AD). (Schuck et al., 2015). AIM OF THIS STUDY: The aim of this work is to explore ß-secretase inhibitory activity of compounds isolated from the aerial part of endemic Caragana balchaschensis (Kom.) Pojark. We provided a full characterization of their inhibitory mechanisms, binding affinities, and binding modes. MATERIALS AND METHODS: The isolation of quercetin derivatives was accomplished by various chromatographical approaches and their structures were annotated by spectroscopic analysis. The detailed kinetic behavior of ß-secretase inhibitors was determined by estimation of kinetic parameters (Km, Vmax, KI, and KIS). Binding affinities (KSV) and binding modes of inhibitors were elucidated by fluorescence quenching and molecular docking studies, respectively. RESULTS: O-methylated quercetins (2-7) were significantly effective in ß-secretase inhibition with IC50 ranging from 1.2 to 6.5 µM. The most active one (6) was 20-fold effective than the mother skeleton, quercetin. The O-methyl motif was a critical factor in ß-secretase inhibition: tri-O-methylated (1.2 µM) > di-O-methylated (3.5 µM) > mono-O-methylated (6.5 µM) > quercetin (25.2 µM). In the kinetic study, all quercetins (1-7) showed a noncompetitive inhibition, but glucoside ones (8 and 9) were mixed type I inhibitors. The binding affinities (KSV) were agreed with inhibitory potencies. The O-methylated quercetins were annotated as the most natural abundant metabolites in the aerial part by LC-ESI-TOF/MS. Binding modes of inhibitors to enzyme were elucidated by molecular docking experiments. CONCLUSION: This study disclosed that most of the major phenolic metabolites of the aerial part of C. balchaschensis are O-methylated quercetins, which have a significant inhibitory effect on ß-secretase, which is a critical factor for AD.

Phytochemical analysis of aerial part of Ikonnikovia kaufmanniana and their protection of DNA damage.

Ikonnikovia kaufmanniana is an endemic plant of Kazakhstan of which phytochemical analysis has not been reported. The present study found out that this species enriched with antioxidant chemicals. Isolation and structural identification processes reveal twelve phenolic compounds (1-12) having dihydroflavanonol, flavonol, isoflavone and flavanol skeletons. The annotation of individual components in the extract was carried out by LC-ESI-MS/MS to represent a chemotaxonomic marker of the target plant. The antioxidant activities of all compounds were screened using three different radical sources (DPPH, ORAC, and hydroxyl radicals). Most compounds (1-11) had significant antioxidant activity against three radical sources, and their efficacies were found to differ by their functionality and skeleton. The potential of the isolated compounds in preventing oxidative damage of DNA was evaluated with pBR322 plasmid DNA. Compounds (1, 5, 7, and 8) had protective effects on DNA damaged with 80% efficacy at 60 µM concentration.

Xanthine Oxidase Inhibition and Anti-LDL Oxidation by Prenylated Isoflavones from Flemingia philippinensis Root.

Xanthine oxidase is a frontier enzyme to produce oxidants, which leads to inflammation in the blood. Prenylated isoflavones from Flemingia philippinensis were found to display potent inhibition against xanthine oxidase (XO). All isolates (1-9) inhibited XO enzyme with IC50 ranging 7.8~36.4 µM. The most active isoflavones (2-5, IC50 = 7.8~14.8 µM) have the structural feature of a catechol motif in B-ring. Inhibitory behaviors were disclosed as a mixed type I mode of inhibition with KI < KIS. Binding affinities to XO enzyme were evaluated. Fluorescence quenching effects agreed with inhibitory potencies (IC50s). The compounds (2-5) also showed potent anti-LDL oxidation effects in the thiobarbituric acid-reactive substances (TBARS) assay, the lag time of conjugated diene formation, relative electrophoretic mobility (REM), and fragmentation of apoB-100 on copper-mediated LDL oxidation. The compound 4 protected LDL oxidation with 0.7 µM in TBARS assay, which was 40-fold more active than genistein (IC50 = 30.4 µM).

Chelidonine Induces Caspase-Dependent and Caspase-Independent Cell Death through G2/M Arrest in the T98G Human Glioblastoma Cell Line.

Chelidonium majus L. (family Papaveraceae), commonly known as greater celandine or tetterwort, has been reported to have antibacterial and anticancer effects and chelidonine is known as a functional metabolite extracted from C.

Human neutrophil elastase (HNE) inhibitory polyprenylated acylphloroglucinols from the flowers of Hypericum ascyron.

In the course of an investigation of human neutrophil elastase (HNE) associated with inflammation, the extract of the flower parts of Hypericum ascyron showed a significant influence to HNE. The responsible metabolites to HNE inhibition were found to be eight polyprenylated acylphloroglucinols, PPAPs (1-8) which showed IC50 ranges between 2.4 and 19.9 µM. This is the first report to demonstrate that PPAP skeleton exhibits potent HNE inhibition. The compounds 1-3 were characterized and newly named as ascyronone E (IC50 = 4.3 µM), ascyronone F (IC50 = 19.9 µM), ascyronone G (IC50 = 4.5 µM) based on 2D-NMR spectroscopic data. In the kinetic analysis of double reciprocal plots, all the compounds showed noncompetitive behaviors to HNE enzyme with the remaining of Km and the increase of Vmax. The binding affinity levels (KSV) by using fluorescence were sufficient to be able to prove that PPAPs (1-8) had compliant interaction with inhibitory potencies.

Tyrosinase inhibitory study of flavonolignans from the seeds of Silybum marianum (Milk thistle).

Anti-melanogenesis effects of silymarin from milk thistle have been reported recently, but detailed tyrosinase inhibition properties of individual components have not been investigated. This study purported to substantiate tyrosinase inhibition and its mechanism based on a single metabolite. The responsible components for tyrosinase inhibition of target source were found out as flavonolignans which consist of isosilybin A (1), isosilybin B (2), silydianin (3), 2,3-dihydrosilychristin (4), silychristin A (5), silychristin B (6) and silybin (7), respectively. The isolated flavonolignans (1-7) inhibited both monophenolase (IC50 = 1.7-7.6 µM) and diphenolase (IC50 = 12.1-44.9 µM) of tyrosinase significantly. Their inhibitions were 10-fold effective in comparison with their mother skeletons (8-10). Inhibitory functions were also proved by HPLC analysis using N-acetyl-l-tyrosine as substrate. The predominant formation of Emet·I was confirmed from a long prolongation of lag time and a decrease of the static state activity of the enzyme. All tested compounds had a significant binding affinity to tyrosinase with KSV values of 0.06-0.27 × 104 L·mol-1, which are well correlated with IC50s. In kinetic study, all flavonolignan (1-7) were mixed type I (KI < KIS) inhibitors, whereas their mother skeletons (8-10) were competitive ones. The UPLC-ESI-TOF/MS analysis showed that the isolated inhibitors are the most abundant metabolites in the target plant.

Effectiveness of Prenyl Group on Flavonoids from Epimedium koreanum Nakai on Bacterial Neuraminidase Inhibition.

In this study, the inhibitory potential of bacterial neuraminidase (NA) was observed on the leaves of Epimedium koreanum Nakai, which is a popular ingredient in traditional herbal medicine. This study attempted to isolate the relevant, responsible metabolites and elucidate their inhibition mechanism. The methanol extraction process yielded eight flavonoids (1⁻8), of which compounds 7 and 8 were new compounds named koreanoside F and koreanoside G, respectively. All the compounds (1⁻8) showed a significant inhibition to bacterial NA with IC50 values of 0.17⁻106.3 µM. In particular, the prenyl group on the flavonoids played a critical role in bacterial NA inhibition. Epimedokoreanin B (compound 1, IC50 = 0.17 µM) with two prenyl groups on C8 and C5' of luteolin was 500 times more effective than luteolin (IC50 = 85.6 µM). A similar trend was observed on compound 2 (IC50 = 0.68 µM) versus dihydrokaempferol (IC50 = 500.4 µM) and compound 3 (IC50 = 12.6 µM) versus apigenin (IC50 = 107.5 µM). Kinetic parameters (Km, Vmax, and Kik/Kiv) evaluated that all the compounds apart from compound 5 showed noncompetitive inhibition. Compound 5 was proven to be a mixed type inhibitor. In an enzyme binding affinity experiment using fluorescence, affinity constants (KSV) were tightly related to inhibitory activities.

Antioxidant Activities of Phenolic Metabolites from Flemingia philippinensis Merr. et Rolfe and Their Application to DNA Damage Protection.

F. philippinensis Merr. et Rolfe has been cultivated on a large scale and is widely consumed by local inhabitants as an important nutraceutical, especially against rheumatism which has a deep connection with antioxidants. In this study, a total of 18 different phenolic metabolite compounds in F. philippinensis were isolated and identified, and evaluated for their antioxidant and DNA damage protection potential. The antioxidant activity of the 18 identified compounds was screened using DPPH, ORAC, hydroxyl and superoxide radical scavenging assays. The antioxidant potential of the compounds was found to differ by functionality and skeleton. However, most compounds showed a good antioxidant potential. In particular, seven of the identified compounds, namely, compounds 2, 3, 6, 10, 11, 15 and 16, showed significant protective effects on pBR322 plasmid DNA against the mutagenic and toxic effects of Fenton's reaction. The most active compound, compound 2, displayed a dose-dependent DNA damage protection potential in the range of 7.5~60.0 µM. The DNA damage protective effect of the identified compounds was significantly correlated with the hydroxyl radical scavenging activity. Compounds that exhibited effective (IC50 = 5.4~12.5 µg/mL) hydroxyl radical scavenging activity were found to be the ones with higher DNA damage protection potential.

A Multi-Omics Analysis of Glycine max Leaves Reveals Alteration in Flavonoid and Isoflavonoid Metabolism Upon Ethylene and Abscisic Acid Treatment.

Phytohormones are central to plant growth and development. Despite the advancement in our knowledge of hormone signaling, downstream targets, and their interactions upon hormones action remain largely fragmented, especially at the protein and metabolite levels. With an aim to get new insight into the effects of two hormones, ethylene (ET) and abscisic acid (ABA), this study utilizes an integrated proteomics and metabolomics approach to investigate their individual and combined (ABA+ET) signaling in soybean leaves. Targeting low-abundance proteins, our previously established protamine sulfate precipitation method was applied, followed by label-free quantification of identified proteins. A total of 4129 unique protein groups including 1083 differentially modulated in one (individual) or other (combined) treatments were discerned. Functional annotation of the identified proteins showed an increased abundance of proteins related to the flavonoid and isoflavonoid biosynthesis and MAPK signaling pathway in response to ET treatment. HPLC analysis showed an accumulation of isoflavones (genistin, daidzein, and genistein) upon ET treatment, in agreement with the proteomics results. A metabolome analysis assigned 79 metabolites and further confirmed the accumulation of flavonoids and isoflavonoids in response to ET. A potential cross-talk between ET and MAPK signaling, leading to the accumulation of flavonoids and isoflavonoids in soybean leaves is suggested.

Auriculasin-induced ROS causes prostate cancer cell death via induction of apoptosis.

Recent studies have demonstrated that natural agents targeting the accumulation of reactive oxygen species (ROS) that selectively kill, leaving normal cells undamaged, can suppress prostate cancer. Here, we show that auriculasin, derived from Flemingia philippinensis, induces significant cell death and apoptosis via ROS generation in prostate cancer cells. Auriculasin treatment resulted in selective apoptotic cell death in LNCaP prostate cancer cells, characterized by DNA fragmentation, accumulation of sub-G1 cell population, cleavage of poly (ADP-ribose) polymerase (PARP), regulation of Bax/Bcl-2 ratio, increase of cytosolic apoptosis-inducing factor (AIF) and endonuclease G (EndoG), in addition to inhibiting tumor growth in a xenograft mouse model. Interestingly, auriculasin-induced apoptosis did not result in caspase-3, -8, and -9 activations. We found that auriculasin treatment decreased phosphorylation of AKT/mTOR/p70s6k in a dose- and time-dependent manner. Further, cellular ROS levels increased in LNCaP cells treated with auriculasin and blocking ROS accumulation with ROS scavengers resulted in inhibition of auriculasin-induced PARP cleavage, AIF increase, upregulation of Bax/Bcl-2 ratio, and decrease in AKT/mTOR phosphorylation. Taken together, these data suggest that auriculasin targets ROS-mediated caspase-independent pathways and suppresses PI3K/AKT/mTOR signaling, which leads to apoptosis and decreased tumor growth.

Aralia elata (Miq) Seem Extract Decreases O-GlcNAc Transferase Expression and Retinal Cell Death in Diabetic Mice.

Aralia elata (Miq) Seem (AES) is a medicinal plant used in traditional Chinese and Korean medicine for the treatment of several diseases, including diabetes. This study aimed to investigate the neuroprotective effect of AES extract against high glucose-induced retinal injury in diabetic mice. AES extract (20 and 100 mg/kg body weight) was orally administered to control mice or mice with streptozotocin-induced diabetes. Protein levels of O-linked ß-N-acetylglucosamine (O-GlcNAc) transferase (OGT), carbohydrate-responsive element-binding protein (ChREBP), sterol regulatory element-binding protein (SREBP)-1, thioredoxin-interacting protein (TXNIP), fatty acid synthase (FAS), and acetyl CoA carboxylase (ACC) were analyzed by western blotting. Colocalization of terminal deoxynucleotide transferase-mediated dUTP nicked-end labeling (TUNEL)-positive ganglion cells and OGT, ChREBP, or TXNIP were monitored using double immunofluorescence analysis. Interaction between ChREBP and OGT was assessed using coimmunoprecipitation analysis. AES extract protected the retinas from neuronal injury and decreased levels of OGT, ChREBP, TXNIP, SREBP-1, FAS, and ACC in the diabetic retinas. AES extract reduced colocalization of TUNEL-positive ganglion cells and OGT, ChREBP, or TXNIP in the diabetic retinas. Coimmunoprecipitation analysis indicated that AES extract reduced interaction between ChREBP and OGT and attenuated ganglion cell death in diabetic retinas. Moreover, the ChREBP that colocalized with OGT or the TUNEL signal was significantly decreased in diabetic mice treated with AES extract. These findings show that AES extract can alleviate OGT-, ChREBP-, TXNIP-, or SREBP-1-related retinal injury in diabetic retinopathy.

Phytochemical profile and angiotensin I converting enzyme (ACE) inhibitory activity of Limonium michelsonii Lincz.

Members of the genus Limonium are widely used as medicinal herbs due to their health-promoting effects, such as an ability to improve blood circulation by inhibiting angiotensin I converting enzyme (ACE). While the potential of L. michelsonii Lincz. (a medicinal plant endemic to Kazakhstan) to inhibit ACE has been demonstrated, the inhibitory activities of its secondary metabolites have not been explored. In this work, the principal phenolic compounds (1-20) among these metabolites were isolated to determine the components responsible for ACE inhibition. The natural abundances of the active constituents within the target plant were characterized by UPLC-Q-TOF/MS analysis. All of the isolated compounds except for gallates 10-12 were found to significantly inhibit ACE, with IC50 values of between 7.1 and 138.4 µM. Unexpectedly, the flavonol glycosides 16-20 were observed to be more potent than the corresponding aglycones 4 and 5. For example, quercetin (4) had IC50 = 30.3 µM, whereas its glycosides (16, 17) had IC50 = 10.2 and 14.5 µM, respectively. A similar trend was observed for myricetin (5) and its glycosides (18-20). In a kinetic study, the flavonols 3-5 and 16-20 and the dihydroflavonols 8 and 9 behaved as competitive inhibitors, whereas other flavones (1, 2, 13-15) and flavanones (6, 7) performed noncompetitive inhibition.