Prunin from Poncirus trifoliata (L.) Rafin Inhibits Aldose Reductase and Glucose-Fructose-Mediated Protein Glycation and Oxidation of Human Serum Albumin.

Diabetes complications are associated with aldose reductase (AR) and advanced glycation end products (AGEs). Using bioassay-guided isolation by column chromatography, 10 flavonoids and one coumarin were isolated from Poncirus trifoliata Rafin and tested in vitro for an inhibitory effect against human recombinant AR (HRAR) and rat lens AR (RLAR). Prunin, narirutin, and naringin inhibited RLAR (IC50 0.48-2.84 µM) and HRAR (IC50 0.68-4.88 µM). Docking simulations predicted negative binding energies and interactions with the RLAR and HRAR binding pocket residues. Prunin (0.1 and 12.5 µM) prevented the formation of fluorescent AGEs and nonfluorescent Nε-(carboxymethyl) lysine (CML), as well as the fructose-glucose-mediated protein glycation and oxidation of human serum albumin (HSA). Prunin suppressed the formation of the ß-cross-amyloid structure of HSA. These results indicate that prunin inhibits oxidation-dependent protein damage, AGE formation, and AR, which may help prevent diabetes complications.

Discovery of Potent Angiotensin-Converting Enzyme Inhibitors in Pomegranate as a Treatment for Hypertension.

Pomegranate (Punica granatum L.) is associated with numerous health benefits due to its high levels of antioxidant polyphenolic substances. Since pomegranate extract has been shown to inhibit angiotensin-converting enzyme (ACE), the potential inhibitory effect of most of its main constituents against ACE is unknown. Therefore, we tested the activities of 24 major compounds, the majority of which significantly inhibited ACE. Notably, pedunculagin, punicalin, and gallagic acid were the most effective ACE inhibitors with IC50 values of 0.91, 1.12, and 1.77 µM, respectively. As demonstrated in molecular docking studies, compounds block ACE by forming multiple hydrogen bonds and hydrophobic interactions with catalytic residues and zinc ions in ACE's C- and N-domains, consequently inhibiting ACE's catalytic activity. Also, the most active pedunculagin stimulated nitric oxide (NO) production, activated the endothelial nitric oxide synthase enzyme (eNOS), and significantly increased eNOS protein expression levels up to 5.3-fold in EA.hy926 cells. Furthermore, pedunculagin increased in cellular calcium (Ca2+) concentration promoted eNOS enzyme activation and reduced the production of reactive oxygen species (ROS). In addition, the active compounds improved glucose uptake in insulin-resistant C2C12 skeletal muscle cells in a dose-dependent manner. The results of these computational, in vitro, and cellular experiments provide further evidence to the traditional medicine that involves using pomegranates to treat cardiovascular diseases like hypertension.

Icariside II, a Prenyl-Flavonol, Alleviates Inflammatory and Neuropathic Pain by Inhibiting T-Type Calcium Channels and USP5-Cav3.2 Interactions.

Cav3.2 channels play an important role in the afferent nociceptive pathway, which is responsible for both physiological and pathological pain transmission. Cav3.2 channels are upregulated during neuropathic pain or peripheral inflammation in part due to an increased association with the deubiquitinase USP5. In this study, we investigated nine naturally occurring flavonoid derivatives which we tested for their abilities to inhibit transiently expressed Cav3.2 channels and their interactions with USP5. Icariside II (ICA-II), one of the flavonols studied, inhibited the biochemical interactions between USP5 and Cav3.2 and concomitantly and effectively blocked Cav3.2 channels. Molecular docking analysis predicts that ICA-II binds to the cUBP domain and the Cav3.2 interaction region. In addition, ICA-II was predicted to interact with residues in close proximity to the Cav3.2 channel's fenestrations, thus accounting for the observed blocking activity. In mice with inflammatory and neuropathic pain, ICA-II inhibited both phases of the formalin-induced nocifensive responses and abolished thermal hyperalgesia induced by injection of complete Freund's adjuvant (CFA) into the hind paw. Furthermore, ICA-II produced significant and long-lasting thermal anti-hyperalgesia in female mice, whereas Cav3.2 null mice were resistant to the action of ICA-II. Altogether, our data show that ICA-II has analgesic activity via an action on Cav3.2 channels.

An Arylbenzofuran, Stilbene Dimers, and Prenylated Diels-Alder Adducts as Potent Diabetic Inhibitors from Morus bombycis Leaves.

Morus bombycis has a long history of usage as a treatment for metabolic diseases, especially, diabetes mellitus (DM). Thus, we aimed to isolate and evaluate bioactive constituents derived from M. bombycis leaves for the treatment of DM. According to bioassay-guided isolation by column chromatography, eight compounds were obtained from M. bombycis leaves: two phenolic compounds, p-coumaric acid (1) and chlorogenic acid methyl ester (2), one stilbene, oxyresveratrol (3), two stilbene dimers, macrourin B (4) and austrafuran C (6), one 2-arylbenzofuran, moracin M (5), and two Diels-Alder type adducts, mulberrofuran F (7) and chalcomoracin (8). Among the eight isolated compounds, the anti-DM activity of 3-8 (which possess chemotaxonomic significance in Morus species) was evaluated by inhibition of α-glucosidase, protein tyrosine phosphatase 1B (PTP1B), human recombinant aldose reductase (HRAR), and advanced glycation end-product (AGE) formation as well as by scavenging peroxynitrite (ONOO-), which are crucial therapeutic targets of DM and its complications. Compounds 4 and 6-8 significantly inhibited α-glucosidase, PTP1B, and HRAR enzymes with mixed-type and non-competitive-type inhibition modes. Furthermore, the four compounds had low negative binding energies in both enzymes according to molecular docking simulation, and compounds 3-8 exhibited strong antioxidant capacity by inhibiting AGE formation and ONOO- scavenging. Overall results suggested that the most active stilbene-dimer-type compounds (4 and 6) along with Diels-Alder type adducts (7 and 8) could be promising therapeutic and preventive resources against DM and have the potential to be used as antioxidants, anti-diabetic agents, and anti-diabetic complication agents.

Ursonic acid from Artemisia montana exerts anti-diabetic effects through anti-glycating properties, and by inhibiting PTP1B and activating the PI3K/Akt signaling pathway in insulin-resistant C2C12 cells.

Artemisia is one of the largest genera in the plant family Asteraceae and has long been used in traditional medicine for its antitussive, analgesic, antihypertensive, antitoxic, antiviral, antimalarial, and anti-inflammatory properties. However, the anti-diabetic activity of Artemisia montana has not been broadly studied. The goal of this study was to determine whether extracts of the aerial parts of A. montana and its main constituents inhibit protein tyrosine phosphatase 1B (PTP1B) and α-glucosidase activities. We isolated nine compounds from A. montana including ursonic acid (UNA) and ursolic acid (ULA), which significantly inhibited PTP1B with IC50 values of 11.68 and 8.73 µM, respectively. In addition, UNA showed potent inhibitory activity against α-glucosidase (IC50 = 61.85 µM). Kinetic analysis of PTP1B and α-glucosidase inhibition revealed that UNA was a non-competitive inhibitor of both enzymes. Docking simulations of UNA demonstrated negative binding energies and close proximity to residues in the binding pockets of PTP1B and α-glucosidase. Molecular docking simulations between UNA and human serum albumin (HSA) revealed that UNA binds tightly to all three domains of HSA. Furthermore, UNA significantly inhibited fluorescent AGE formation (IC50 = 4.16 µM) in a glucose-fructose-induced HSA glycation model over the course of four weeks. Additionally, we investigated the molecular mechanisms underlying the anti-diabetic effects of UNA in insulin-resistant C2C12 skeletal muscle cells and discovered that UNA significantly increased glucose uptake and decreased PTP1B expression. Further, UNA increased GLUT-4 expression level by activating the IRS-1/PI3K/Akt/GSK-3 signaling pathway. These findings clearly demonstrate that UNA from A. montana shows great potential for treatment of diabetes and its complications.

Differential regulation of Cav 3.2 and Cav 2.2 calcium channels by CB1 receptors and cannabidiol.

BACKGROUND AND PURPOSE: Cannabinoids are a promising therapeutic avenue for chronic pain. However, clinical trials often fail to report analgesic efficacy of cannabinoids. Inhibition of voltage gate calcium (Cav ) channels is one mechanism through which cannabinoids may produce analgesia. We hypothesized that cannabinoids and cannabinoid receptor agonists target different types of Cav channels through distinct mechanisms. EXPERIMENTAL APPROACH: Electrophysiological recordings from tsA-201 cells expressing either Cav 3.2 or Cav 2.2 were used to assess inhibition by HU-210 or cannabidiol (CBD) in the absence and presence of the CB1 receptor. Homology modelling assessed potential interaction sites for CBD in both Cav 2.2 and Cav 3.2. Analgesic effects of CBD were assessed in mouse models of inflammatory and neuropathic pain. KEY RESULTS: HU-210 (1 µM) inhibited Cav 2.2 function in the presence of CB1 receptor but had no effect on Cav 3.2 regardless of co-expression of CB1 receptor. By contrast, CBD (3 µM) produced no inhibition of Cav 2.2 and instead inhibited Cav 3.2 independently of CB1 receptors. Homology modelling supported these findings, indicating that CBD binds to and occludes the pore of Cav 3.2, but not Cav 2.2. Intrathecal CBD alleviated thermal and mechanical hypersensitivity in both male and female mice, and this effect was absent in Cav 3.2 null mice. CONCLUSION AND IMPLICATIONS: Our findings reveal differential modulation of Cav 2.2 and Cav 3.2 channels by CB1 receptors and CBD. This advances our understanding of how different cannabinoids produce analgesia through action at different voltage-gated calcium channels and could influence the development of novel cannabinoid-based therapeutics for treatment of chronic pain.

6-Formyl Umbelliferone, a Furanocoumarin from Angelica decursiva L., Inhibits Key Diabetes-Related Enzymes and Advanced Glycation End-Product Formation.

Over the years, great attention has been paid to coumarin derivatives, a set of versatile molecules that exhibit a wide variety of biological activities and have few toxic side effects. In this study, we investigated the antidiabetic potential of 6-formyl umbelliferone (6-FU), a novel furanocoumarin isolated from Angelica decursiva. Numerous pharmacological activities of 6-FU have been previously reported; however, the mechanism of its antidiabetic activity is unknown. Therefore, we examined the action of 6-FU on a few candidate-signaling molecules that may underlie its antidiabetic activity, including its inhibition of protein tyrosine phosphatase 1B (PTP1B), α-glucosidase, human recombinant aldose reductase (HRAR), and advanced glycation end-product (AGE) formation (IC50 = 1.13 ± 0.12, 58.36 ± 1.02, 5.11 ± 0.21, and 2.15 ± 0.13 µM, respectively). A kinetic study showed that 6-FU exhibited mixed-type inhibition against α-glucosidase and HRAR and competitive inhibition of PTP1B. Docking simulations of 6-FU demonstrated negative binding energies and close proximity to residues in the binding pockets of those enzymes. We also investigated the molecular mechanisms underlying 6-FU's antidiabetic effects. 6-FU significantly increased glucose uptake and decreased PTP1B expression in insulin-resistant C2C12 skeletal muscle cells. Moreover, 6-FU (0.8-100 µM) remarkably inhibited the formation of fluorescent AGEs in glucose-fructose-induced human serum albumin glycation over the course of 4 weeks. The findings clearly indicate that 6-FU will be useful in the development of multiple target-oriented therapeutic modalities for the treatment of diabetes and diabetes-related complications.

Inhibition of Aldose Reductase by Ginsenoside Derivatives via a Specific Structure Activity Relationship with Kinetics Mechanism and Molecular Docking Study.

This present work is designed to evaluate the anti-diabetic potential of 22 ginsenosides via the inhibition against rat lens aldose reductase (RLAR), and human recombinant aldose reductase (HRAR), using DL-glyceraldehyde as a substrate. Among the ginsenosides tested, ginsenoside Rh2, (20S) ginsenoside Rg3, (20R) ginsenoside Rg3, and ginsenoside Rh1 inhibited RLAR significantly, with IC50 values of 0.67, 1.25, 4.28, and 7.28 µM, respectively. Moreover, protopanaxadiol, protopanaxatriol, compound K, and ginsenoside Rh1 were potent inhibitors of HRAR, with IC50 values of 0.36, 1.43, 2.23, and 4.66 µM, respectively. The relationship of structure-activity exposed that the existence of hydroxyl groups, linkages, and their stereo-structure, as well as the sugar moieties of the ginsenoside skeleton, represented a significant role in the inhibition of HRAR and RLAR. Additional, various modes of ginsenoside inhibition and molecular docking simulation indicated negative binding energies. It was also indicated that it has a strong capacity and high affinity to bind the active sites of enzymes. Further, active ginsenosides suppressed sorbitol accumulation in rat lenses under high-glucose conditions, demonstrating their potential to prevent sorbitol accumulation ex vivo. The findings of the present study suggest the potential of ginsenoside derivatives for use in the development of therapeutic or preventive agents for diabetic complications.

A Synthetically Accessible Small-Molecule Inhibitor of USP5-Cav3.2 Calcium Channel Interactions with Analgesic Properties.

Cav3.2 calcium channels are important mediators of nociceptive signaling in the primary afferent pain pathway, and their expression is increased in various rodent models of chronic pain. Previous work from our laboratory has shown that this is in part mediated by an aberrant expression of deubiquitinase USP5, which associates with these channels and increases their stability. Here, we report on a novel bioactive rhodanine compound (II-1), which was identified in compound library screens. II-1 inhibits biochemical interactions between USP5 and the Cav3.2 domain III-IV linker in a dose-dependent manner, without affecting the enzymatic activity of USP5. Molecular docking analysis reveals two potential binding pockets at the USP5-Cav3.2 interface that are distinct from the binding site of the deubiquitinase inhibitor WP1130 (a.k.a. degrasyn). With an understanding of the ability of some rhodanines to produce false positives in high-throughput screening, we have conducted several orthogonal assays to confirm the validity of this hit, including in vivo experiments. Intrathecal delivery of II-1 inhibited both phases of formalin-induced nocifensive behaviors in mice, as well as abolished thermal hyperalgesia induced by the delivery of complete Freund's adjuvant (CFA) to the hind paw. The latter effects were abolished in Cav3.2 null mice, thus confirming that Cav3.2 is required for the action of II-1. II-1 also mediated a robust inhibition of mechanical allodynia induced by injury to the sciatic nerve. Altogether, our data uncover a novel class of analgesics─well suited to rapid structure-activity relationship studies─that target the Cav3.2/USP5 interface.

Structural Bases for Hesperetin Derivatives: Inhibition of Protein Tyrosine Phosphatase 1B, Kinetics Mechanism and Molecular Docking Study.

In the present study, we investigated the structure-activity relationship of naturally occurring hesperetin derivatives, as well as the effects of their glycosylation on the inhibition of diabetes-related enzyme systems, protein tyrosine phosphatase 1B (PTP1B) and α-glycosidase. Among the tested hesperetin derivatives, hesperetin 5-O-glucoside, a single-glucose-containing flavanone glycoside, significantly inhibited PTP1B with an IC50 value of 37.14 ± 0.07 µM. Hesperetin, which lacks a sugar molecule, was the weakest inhibitor compared to the reference compound, ursolic acid (IC50 = 9.65 ± 0.01 µM). The most active flavanone hesperetin 5-O-glucoside suggested that the position of a sugar moiety at the C-5-position influences the PTP1B inhibition. It was observed that the ability to inhibit PTP1B is dependent on the nature, position, and number of sugar moieties in the flavonoid structure, as well as conjugation. In the kinetic study of PTP1B enzyme inhibition, hesperetin 5-O-glucoside led to mixed-type inhibition. Molecular docking studies revealed that hesperetin 5-O-glucoside had a higher binding affinity with key amino residues, suggesting that this molecule best fits the PTP1B allosteric site cavity. The data reported here support hesperetin 5-O-glucoside as a hit for the design of more potent and selective inhibitors against PTP1B in the search for a new anti-diabetic treatment.

Phytochemistry, Ethnopharmacological Uses, Biological Activities, and Therapeutic Applications of Cassia obtusifolia L.: A Comprehensive Review.

Cassia obtusifolia L., of the Leguminosae family, is used as a diuretic, laxative, tonic, purgative, and natural remedy for treating headache, dizziness, constipation, tophobia, and lacrimation and for improving eyesight. It is commonly used in tea in Korea. Various anthraquinone derivatives make up its main chemical constituents: emodin, chrysophanol, physcion, obtusifolin, obtusin, au rantio-obtusin, chryso-obtusin, alaternin, questin, aloe-emodin, gluco-aurantio-obtusin, gluco-obtusifolin, naphthopyrone glycosides, toralactone-9-ß-gentiobioside, toralactone gentiobioside, and cassiaside. C. obtusifolia L. possesses a wide range of pharmacological properties (e.g., antidiabetic, antimicrobial, anti-inflammatory, hepatoprotective, and neuroprotective properties) and may be used to treat Alzheimer's disease, Parkinson's disease, and cancer. In addition, C. obtusifolia L. contributes to histamine release and antiplatelet aggregation. This review summarizes the botanical, phytochemical, and pharmacological features of C. obtusifolia and its therapeutic uses.

Inhibition of Angiotensin-I Converting Enzyme by Ginsenosides: Structure-Activity Relationships and Inhibitory Mechanism.

Ginseng (Panax ginseng C. A. Meyer) extract has been reported to inhibit the angiotensin converting enzyme (ACE); however, the possible inhibitory action of most of its constituents (ginsenosides) against ACE remains unknown. Thus, in this study, we investigated ginsenoside derivatives' inhibitory effect on ACE. We assessed the activities of 22 ginsenosides, most of which inhibited ACE significantly. Notably, protopanaxatriol, protopanaxadiol, and ginsenoside Rh2 exhibited the most potent ACE inhibitory potential, with IC50 values of 1.57, 2.22, and 5.60 µM, respectively. Further, a kinetic study revealed different modes of inhibition against ACE. Molecular docking studies have confirmed that ginsenosides inhibit ACE via many hydrogen bonds and hydrophobic interactions with catalytic residues and zinc ion of C- and N-domain ACE that block the catalytic activity of ACE. In addition, we found that the active ginsenosides stimulated glucose uptake in insulin-resistant C2C12 skeletal muscle cells in a dose-dependent manner. Moreover, the most active ginsenosides' reactive oxygen species (ROS) and peroxynitrite (ONOO-) scavenging properties were evaluated, in which IC50 values ranged from 1.44-43.83 to 2.36-39.56 µM in ONOO- and ROS, respectively. The results derived from these computational and in vitro experiments provide additional scientific support for the anecdotal use of ginseng in traditional medicine to treat cardiovascular diseases such as hypertension.

Insulin-Mimetic Dihydroxanthyletin-Type Coumarins from Angelica decursiva with Protein Tyrosine Phosphatase 1B and α-Glucosidase Inhibitory Activities and Docking Studies of Their Molecular Mechanisms.

As a traditional medicine, Angelica decursiva has been used for the treatment of many diseases. The goal of this study was to evaluate the potential of four natural major dihydroxanthyletin-type coumarins-(+)-trans-decursidinol, Pd-C-I, Pd-C-II, and Pd-C-III-to inhibit the enzymes, protein tyrosine phosphatase 1B (PTP1B) and α-glucosidase. In the kinetic study of the PTP1B enzyme's inhibition, we found that (+)-trans-decursidinol, Pd-C-I, and Pd-C-II led to competitive inhibition, while Pd-C-III displayed mixed-type inhibition. Moreover, (+)-trans-decursidinol exhibited competitive-type, and Pd-C-I and Pd-C-II mixed-type, while Pd-C-III showed non-competitive type inhibition of α-glucosidase. Docking simulations of these coumarins showed negative binding energies and a similar proximity to residues in the PTP1B and α-glucosidase binding pocket, which means they are closely connected and strongly binding with the active enzyme site. In addition, dihydroxanthyletin-type coumarins are up to 40 µM non-toxic in HepG2 cells and have substantially increased glucose uptake and decreased expression of PTP1B in insulin-resistant HepG2 cells. Further, coumarins inhibited ONOO--mediated albumin nitration and scavenged peroxynitrite (ONOO-), and reactive oxygen species (ROS). Our overall findings showed that dihydroxanthyletin-type coumarins derived from A. decursiva is used as a dual inhibitor for enzymes, such as PTP1B and α-glucosidase, as well as for insulin susceptibility.

Effect of Manure Application on Net Nitrification Rates, Heavy Metal Concentrations and Nitrifying Archaea/Bacteria in Soils.

In this study, we determined the effect of manure application on net nitrification rates (NNRs), heavy metal concentrations (HMCs), and abundance of ammonia-oxidizing archaea (AOA)/bacteria (AOB), and nitrite-oxidizing bacteria (NOB) in soil. HMCs were measured by atomic absorption spectroscopy. Abundance of AOA, AOB, and NOB was enumerated by q-PCR. NNRs ranged from 2.8 to 14.7 mg kg-1 h-1 and were significantly (p < 0.05) increased in manure soils as compared to control soils. NNRs were affected by pH 7 and temperature 30°C. Cd, Fe and Pb concentrations were classified as excessively polluted, moderate contamination and slight pollution, respectively, in the manure soils. NNRs and concentrations of Fe and Pb were significantly (p < 0.00) positive correlated, but Cu and Cd were significantly (p < 0.00) negative correlated with NNRs. Application of manure significantly (p < 0.05) increased HMCs (Fe, Cu, and Pb), which have indirect and direct effects on NNRs and nitrifying bacteria.

Angiotensin-I-Converting Enzyme Inhibitory Activity of Coumarins from Angelica decursiva.

The bioactivity of ten traditional Korean Angelica species were screened by angiotensin-converting enzyme (ACE) assay in vitro. Among the crude extracts, the methanol extract of Angelica decursiva whole plants exhibited potent inhibitory effects against ACE. In addition, the ACE inhibitory activity of coumarins 1-5, 8-18 was evaluated, along with two phenolic acids (6, 7) obtained from A. decursiva. Among profound coumarins, 11-18 were determined to manifest marked inhibitory activity against ACE with IC50 values of 4.68-20.04 µM. Compounds 12, 13, and 15 displayed competitive inhibition against ACE. Molecular docking studies confirmed that coumarins inhibited ACE via many hydrogen bond and hydrophobic interactions with catalytic residues and zinc ion of C- and N-domain ACE that blocked the catalytic activity of ACE. The results derived from these computational and in vitro experiments give additional scientific support to the anecdotal use of A. decursiva in traditional medicine to treat cardiovascular diseases such as hypertension.

Umbelliferone derivatives exert neuroprotective effects by inhibiting monoamine oxidase A, self-amyloidß aggregation, and lipid peroxidation.

Umbelliferone has been demonstrated to have a wide range of biological activities. However, the effect of incorporating a formyl moiety in the umbelliferone scaffold has not been investigated. In this paper, we investigated the inhibitory activity of six coumarins, namely umbelliferone (1), 6-formyl umbelliferone (2), 8-formyl umbelliferone (3), umbelliferone-6-carboxylic acid (4), esculetin (5), and scopoletin (6) against human monoamine oxidases (hMAOs), self-amyloid ß (Aß) aggregation, and lipid peroxidation. We found that all compounds had high selectivity for hMAO-A in comparison with hMAO-B. Among the compounds, 2 exhibited the highest hMAO inhibitory activity with an IC50 value of 3.23 µM for hMAO-A and 15.31 µM for hMAO-B. Enzyme kinetic analysis showed that 2 and 3 were competitive hMAO inhibitors. In silico hydrated molecular docking simulations revealed that the coumarins interacted with substrate-binding site residues of the enzymes and the isoalloxazine ring of FAD. In addition, formyl coumarins 2 and 3 significantly inhibited lipid peroxidation in rat brain homogenates and self-Aß25-35 aggregation compared to other derivatives. These represent the first experimental and modelling data for hMAO-A/B inhibition by umbelliferone derivatives. Together, the data suggest that introduction of a formyl moiety in the 7-hydroxycoumarin scaffold, especially at the 6 position, plays an important role in the inhibition of hMAOs, Aß self-aggregation, and lipid peroxidation. Umbelliferone derivative 2 is a promising therapeutic lead scaffold for developing anti-neuropsychiatric disorder drugs that function via selective hMAO-A inhibition.

Flavanone glycosides inhibit ß-site amyloid precursor protein cleaving enzyme 1 and cholinesterase and reduce Aß aggregation in the amyloidogenic pathway.

Alzheimer's disease (AD) is a slow but progressive neurodegenerative disease. One of the pathological hallmarks of AD is the progressive accumulation of ß-amyloid (Aß) in the form of senile plaques, and Aß insult to neuronal cells has been identified as one of the major causes of AD onset. In the present study, we investigated the anti-AD potential of four flavonoids, naringenin, didymin, prunin, and poncirin, by evaluating their ability to inhibit acetylcholinesterase (AChE), butyrylcholinesterase (BChE), and ß-site amyloid precursor protein cleaving enzyme 1 (BACE1). All four flavonoids displayed promising inhibitory activity against AChE, BChE, and BACE1. Structure-activity relationships suggested that glycosylation of naringenin at sugar moieties, and at different positions of the glycosidic linkage, might be closely associated with anti-AD potential. Kinetic and docking studies showed the lowest binding energy and highest affinity for the mixed, competitive, and non-competitive type inhibitors didymin, prunin, and poncirin. Hydrophobic interactions and the number of hydrogen bonds determined the strength of the protein-inhibitor interaction. We also examined the neuroprotective mechanisms by which flavonoids act against Aß25-35-induced toxicity in PC12 cells. Exposure of PC12 cells to 10 µM Aß25-35 for 24 h resulted in a significant decrease in cell viability. In addition, pretreatment of PC12 cells with different concentrations of flavonoids for 1 h significantly reversed the effects of Aß. Furthermore, treatment with the most active flavonoid, didymin, significantly reduced BACE1, APPsß, and C99 expression levels in a dose-dependent manner, without affecting amyloid precursor protein (APP) levels in the amyloidogenic pathway. Together, our results indicate that flavonoids, and in particular didymin, exhibit inhibitory activity in vitro, and may be useful in the development of therapeutic modalities for the treatment of AD.

Didymin, a dietary citrus flavonoid exhibits anti-diabetic complications and promotes glucose uptake through the activation of PI3K/Akt signaling pathway in insulin-resistant HepG2 cells.

Didymin is a naturally occurring orally active flavonoid glycoside (isosakuranetin 7-O-rutinoside) found in various citrus fruits, which has been previously reported to possess a wide variety of pharmacological activities including anticancer, antioxidant, antinociceptive, neuroprotective, hepatoprotective, inflammatory, and cardiovascular. However, there have not been any reports concerning its anti-diabetic potential until now. Therefore, we evaluated the anti-diabetic potential of didymin via inhibition of α-glucosidase, protein tyrosine phosphatase 1B (PTP1B), rat lens aldose reductase (RLAR), human recombinant AR (HRAR), and advanced glycation end-product (AGE) formation inhibitory assays. Didymin strongly inhibited PTP1B, α-glucosidase, HRAR, RLAR, and AGE in the corresponding assays. Kinetic study revealed that didymin exhibited a mixed type inhibition against α-glucosidase and HRAR, while it competitively inhibited PTP1B and RLAR. Docking simulations of didymin demonstrated negative binding energies and close proximity to residues in the binding pocket of HRAR, RLAR, PTP1B and α-glucosidase, indicating that didymin have high affinity and tight binding capacity towards the active site of these enzymes. Furthermore, we also examined the molecular mechanisms underlying the anti-diabetic effects of didymin in insulin-resistant HepG2 cells which significantly increased glucose uptake and decreased the expression of PTP1B in insulin-resistant HepG2 cells. In addition, didymin activated insulin receptor substrate (IRS)-1 by increasing phosphorylation at tyrosine 895 and enhanced the phosphorylations of phosphoinositide 3-kinase (PI3K), Akt, and glycogen synthasekinase-3(GSK-3). Interestingly, didymin reduced the expression of phosphoenolpyruvate carboxykinase and glucose 6-phosphatase, two key enzymes involved in the gluconeogenesis and leading to a diminished glucose production. The results of the present study clearly demonstrated that didymin will be useful for developing multiple target-oriented therapeutic modalities for treatment of diabetes, and diabetes-associated complications.

Inhibition of ß-site amyloid precursor protein cleaving enzyme 1 and cholinesterases by pterosins via a specific structure-activity relationship with a strong BBB permeability.

We extracted 15 pterosin derivatives from Pteridium aquilinum that inhibited ß-site amyloid precursor protein cleaving enzyme 1 (BACE1) and cholinesterases involved in the pathogenesis of Alzheimer's disease (AD). (2R)-Pterosin B inhibited BACE1, acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) with an IC50 of 29.6, 16.2 and 48.1 µM, respectively. The Ki values and binding energies (kcal/mol) between pterosins and BACE1, AChE, and BChE corresponded to the respective IC50 values. (2R)-Pterosin B was a noncompetitive inhibitor against human BACE1 and BChE as well as a mixed-type inhibitor against AChE, binding to the active sites of the corresponding enzymes. Molecular docking simulation of mixed-type and noncompetitive inhibitors for BACE1, AChE, and BChE indicated novel binding site-directed inhibition of the enzymes by pterosins and the structure-activity relationship. (2R)-Pterosin B exhibited a strong BBB permeability with an effective permeability (Pe) of 60.3×10-6 cm/s on PAMPA-BBB. (2R)-Pterosin B and (2R,3 R)-pteroside C significantly decreased the secretion of Aß peptides from neuroblastoma cells that overexpressed human ß-amyloid precursor protein at 500 µM. Conclusively, our study suggested that several pterosins are potential scaffolds for multitarget-directed ligands (MTDLs) for AD therapeutics.

Correction to: Dihydroxanthyletin-type coumarins from Angelica decursiva that inhibits the formation of advanced glycation end products and human recombinant aldose reductase.

The author would like to include conflict of interest statement of the online published article. The correct conflict of interest statement should read as: Conflict of interest The authors declare no conflict of interest.