Carbohydrate-derived bicyclic selenazolines as new dual inhibitors (cholinesterases/OGA) against Alzheimer's disease.

Concerned by the urgent need to explore new approaches for the treatment of Alzheimer's disease, we herein describe the synthesis and evaluation of new multitarget molecules. In particular, we have focused our attention on modulating the activity of cholinesterases (AChE, BuChE) in order to restore the levels of the neurotransmitter acetylcholine, and of O-GlcNAcase (OGA), which is associated with hyperphosphorylation of tau protein, in turn related to the formation of neurofibrillary tangles in the brain. Specifically, we considered the possibility of using carbohydrate-fused 1,3-selenazolines, decorated with a 2-alkylamino or 2-alkoxy moieties. On the one hand, the presence of a selenium atom might be useful in modulating the intrinsic oxidative stress in AD. On the other hand, such bicyclic structure might behave as a transition state analogue of OGA hydrolysis. Moreover, upon protonation, it could mimic the ammonium cation of acetylcholine. The lead compound, bearing a propylamino moiety on C-2 position of the selenazoline motif, proved to be a good candidate against AD; it turned out to be a strong inhibitor of BuChE (IC50 = 0.46 µM), the most prevalent cholinesterase in advanced disease stages, with a roughly 4.8 selectivity index in connection to AChE (IC50 = 2.2 µM). This compound exhibited a roughly 12-fold increase in activity compared to galantamine, one of the currently marketed drugs against AD, and a selective AChE inhibitor, and virtually the same activity as rivastigmine, a selective BuChE inhibitor. Furthermore, it was also endowed with a strong inhibitory activity against human OGA, within the nanomolar range (IC50 = 0.053 µM for hOGA, >100 µM for hHexB), and, thus, with an outstanding selectivity (IC50(hHexB)/IC50(hOGA) > 1887). The title compounds also exhibited an excellent selectivity against a panel of glycosidases and a negligible cytotoxicity against tumor and non-tumor cell lines. Docking simulations performed on the three target enzymes (AChE, BuChE, and OGA) revealed the key interactions to rationalize the biological data.

Structural variation of the 3-acetamido-4,5,6-trihydroxyazepane iminosugar through epimerization and C-alkylation leads to low micromolar HexAB and NagZ inhibitors.

We report the synthesis of seven-membered iminosugars derived from a 3S-acetamido-4R,5R,6S-trihydroxyazepane scaffold and their evaluation as inhibitors of functionally related exo-N-acetylhexosaminidases including human O-GlcNAcase (OGA), human lysosomal ß-hexosaminidase (HexAB), and Escherichia coli NagZ. Capitalizing on the flexibility of azepanes and the active site tolerances of hexosaminidases, we explore the effects of epimerization of stereocenters at C-3, C-5 and C-6 and C-alkylation at the C-2 or C-7 positions. Accordingly, epimerization at C-6 (L-ido) and at C-5 (D-galacto) led to selective HexAB inhibitors whereas introduction of a propyl group at C-7 on the C-3 epimer furnished a potent NagZ inhibitor.

O-GlcNAcase inhibitor has protective effects in intracerebral hemorrhage by suppressing the inflammatory response.

BACKGROUND: Intracerebral hemorrhage (ICH) is aggravated by immune cells that participate in the inflammatory response from the blood-brain barrier (BBB). O-Glycosylation has been reported to regulate the inflammatory response in the central nervous system but its cerebral protective effects remain unknown. Therefore, this study was carried out to investigate the protective effects of O-GlcNAcylation in a murine model of ICH and the possible mechanisms involved. METHODS: The effects of O-GlcNAcylation on hematoma and edema formation were tested using pathological and dry/wet weight methods, whereas its effects on neural function were determined using neurologic tests. The effect of O-GlcNAcylation on BBB integrity was determined by Evans blue dye extrusion. Flow cytometry was used to quantify the immune cells in the central nervous system. Immunofluorescence was used to detect the protective effect of O-GlcNAcylation in ICH. RESULTS: The hematoma volume was significantly lower in the prevention and treatment groups than in the control group after ICH induction, indicating that O-GlcNAcylation had reduced the formation of cerebral hematoma in ICH. In the prevention and treatment groups, the modified neurological severity score, corner turn test and rotating rod test results were improved and the BBB integrity was better than that in the control group. O-GlcNAcylation also regulated the microglia, neutrophils and other central nervous system immune cells after ICH, effectively reducing the inflammatory response. CONCLUSIONS: O-GlcNAcylation played an important role in suppressing the inflammatory response, enhancing the BBB integrity and reducing edema after ICH.

O-GlcNAcase inhibitors as potential therapeutics for the treatment of Alzheimer's disease and related tauopathies: analysis of the patent literature.

Introduction: O-GlcNAcylation is a highly abundant post-translational modification of multiple proteins, including the microtubule-binding protein tau, governed by just two enzymes' concerted action O-GlcNAc transferase OGT and the hydrolase OGA. It is an approach to reduce abnormal tau hyperphosphorylation and aggregation in Alzheimer's disease (AD) and related tauopathies based on the ability of O-GlcNAcylation competing with tau phosphorylation, thus minimizing aggregation. The preclinical validation confirmed OGA inhibitors' efficacy in different transgenic tau mice models. Only three other OGA inhibitors have advanced into clinical trials thus far.Areas covered: 2008-2020 patent literature on OGA inhibitors.Expert opinion: Neurodegenerative disorders and AD specifically represent an enormous challenge since no effective treatments are available. Promising preclinical data has prompted considerable interest in searching for OGA inhibitors as a potential treatment for neurodegenerative disorders. Efforts from different companies have yielded a diverse set of chemotypes. OGA is a highly ubiquitous enzyme with many client proteins, generated data confirms a promising benign profile for OGA inhibition in healthy volunteers. Additionally, OGA PET tracers' existence will be critical for proper dose selection for future PoC Phase II studies, which will proof the true potential of OGA inhibition for the treatment of AD and other tauopathies.

Thiamme2-G, a Novel O-GlcNAcase Inhibitor, Reduces Tau Hyperphosphorylation and Rescues Cognitive Impairment in Mice.

BACKGROUND: Abnormal hyperphosphorylation of microtubule-associated protein tau plays a pivotal role in Alzheimer's disease (AD). We previously found that O-GlcNAcylation inversely correlates to hyperphosphorylation of tau in AD brain, and downregulation of brain O-GlcNAcylation promotes tau hyperphosphorylation and AD-like neurodegeneration in mice. OBJECTIVE: Herein we investigated the effect of increasing O-GlcNAcylation by using intermittent dosing with low doses of a potent novel O-GlcNAcase (OGA) inhibitor on AD-like brain changes and cognitive function in a mouse model of sporadic AD (sAD) induced by intracerebroventricular (ICV) injection of streptozotocin (STZ). METHODS: STZ was injected into the lateral ventricle of C57BL/6J mice. From the second day, Thiamme2-G (TM2G) or saline, as a vehicle control, was orally administered to the ICV-STZ mice three times per week for five weeks. A separate group of ICV-saline mice treated with saline was used as a baseline control. Behavioral tests, including open field and novel object recognition, were conducted three weeks after the first dose of the TM2G or saline. Protein O-GlcNAcylation, tau hyperphosphorylation, synaptic proteins, and neuroinflammation in the mouse brain were assessed by western blotting. RESULTS: ICV-STZ caused decreased protein O-GlcNAcylation. Enhancement of O-GlcNAcylation to moderate levels by using low-dose OGA inhibitor in ICV-STZ mice prevented STZ-induced body weight loss, rescued cognitive impairments, and restored AD-like pathologies, including hyperphosphorylation of tau and abnormalities in synaptic proteins and neuroinflammation. CONCLUSION: These findings suggest that moderately increasing protein O-GlcNAcylation by using low doses of OGA inhibitor may be a suitable therapeutic strategy for sAD.

Discovery of a Novel and Brain-Penetrant O-GlcNAcase Inhibitor via Virtual Screening, Structure-Based Analysis, and Rational Lead Optimization.

O-GlcNAcase (OGA) has received increasing attention as an attractive therapeutic target for tau-mediated neurodegenerative disorders; however, its role in these pathologies remains unclear. Therefore, potent chemical tools with favorable pharmacokinetic profiles are desirable to characterize this enzyme. Herein, we report the discovery of a potent and novel OGA inhibitor, compound 5i, comprising an aminopyrimidine scaffold, identified by virtual screening based on multiple methodologies combining structure-based and ligand-based approaches, followed by sequential optimization with a focus on ligand lipophilicity efficiency. This compound was observed to increase the level of O-GlcNAcylated protein in cells and display suitable pharmacokinetic properties and brain permeability. Crystallographic analysis revealed that the chemical series bind to OGA via characteristic hydrophobic interactions, which resulted in a high affinity for OGA with moderate lipophilicity. Compound 5i could serve as a useful chemical probe to help establish a proof-of-concept of OGA inhibition as a therapeutic target for the treatment of tauopathies.

Emerging roles of protein O-GlcNAcylation in cardiovascular diseases: Insights and novel therapeutic targets.

Cardiovascular diseases (CVDs) are the leading cause of death globally. While the major focus of pharmacological and non-pharmacological interventions has been on targeting disease pathophysiology and limiting predisposing factors, our understanding of the cellular and molecular mechanisms underlying the pathogenesis of CVDs remains incomplete. One mechanism that has recently emerged is protein O-GlcNAcylation. This is a dynamic, site-specific reversible post-translational modification of serine and threonine residues on target proteins and is controlled by two enzymes: O-linked ß-N-acetylglucosamine transferase (OGT) and O-linked ß-N-acetylglucosaminidase (OGA). Protein O-GlcNAcylation alters the cellular functions of these target proteins which play vital roles in pathways that modulate vascular homeostasis and cardiac function. Through this review, we aim to give insights on the role of protein O-GlcNAcylation in cardiovascular diseases and identify potential therapeutic targets in this pathway for development of more effective medicines to improve patient outcomes.

OGA Inhibition Alters Energetics and Nutrient Sensing in Alzheimer's Disease Cytoplasmic Hybrids.

BACKGROUND: Alzheimer's disease (AD) features reductions in key bioenergetic fluxes and perturbed mitochondrial function. Cytoplasmic hybrids (cybrids) generated through the transfer of AD subject mitochondria to mtDNA-depleted SH-SY5Y neuroblastoma cells recapitulate some of these features in an in vitro setting. OBJECTIVE: For this study, we used the AD cybrid model to assess the impact of a nutrient-excess like-state via increasing O-GlcNAcylation on whole cell and mitochondrial homeostasis. METHODS: We induced increased O-GlcNAc by treating AD and control cybrid cell lines with Thiamet G (TMG), an inhibitor of the O-GlcNAcase enzyme that mediates removal of the nutrient-dependent O-GlcNAc modification. RESULTS: Relative to control cybrid cell lines, AD cybrid lines showed a blunted response to TMG-induced O-GlcNAcylation. At baseline, AD cybrid cell line mitochondria showed partial activation of several proteins that help maintain bioenergetic homeostasis such as AMP-Regulated Kinase suggesting that AD mitochondria initiate a state of nutrient stress promoting energetic compensation; however, this compensation reduces the capacity of cells to respond to additional nutrient-related stresses such as TMG treatment. Also, TMG caused disruptions in acetylation and Sirtuin 3 expression, while lowing total energetic output of the cell. CONCLUSION: Together, these findings suggest that modulation of O-GlcNAc is essential for proper energetic function of the mitochondria, and AD mitochondrial capacity to handle nutrient-excess is limited.

Diazaspirononane Nonsaccharide Inhibitors of O-GlcNAcase (OGA) for the Treatment of Neurodegenerative Disorders.

O-GlcNAcylation is a post-translational modification of tau understood to lower the speed and yield of its aggregation, a pathological hallmark of Alzheimer's disease (AD). O-GlcNAcase (OGA) is the only enzyme that removes O-linked N-acetyl-d-glucosamine (O-GlcNAc) from target proteins. Therefore, inhibition of OGA represents a potential approach for the treatment of AD by preserving the O-GlcNAcylated tau protein. Herein, we report the multifactorial optimization of high-throughput screening hit 8 to a potent, metabolically stable, and orally bioavailable diazaspirononane OGA inhibitor (+)-56. The human OGA X-ray crystal structure has been recently solved, but bacterial hydrolases are still widely used as structural homologues. For the first time, we reveal how a nonsaccharide series of inhibitors binds bacterial OGA and discuss the suitability of two different bacterial orthologues as surrogates for human OGA. These breakthroughs enabled structure-activity relationships to be understood and provided context and boundaries for the optimization of druglike properties.

Synthesis of ureido thioglycosides as novel insect ß­N­acetylhexosaminidase OfHex1 inhibitors.

The insect ß-N-acetylhexosaminidase OfHex1 from Ostrinia furnacalis (one of the most destructive agricultural pests) has been considered as a promising pesticide target. In this study, a series of novel and readily available ureido thioglycosides were designed and synthesized based on the catalytic mechanism and the co-crystal structures of OfHex1 with substrates. After evaluation via enzyme inhibition experiments, thioglycosides 11c and 15k were found to have inhibitory activities against OfHex1 with the Ki values of 25.6 µM and 53.8 µM, respectively. In addition, all these ureido thioglycosides exhibited high selectivity toward OfHex1 over hOGA and HsHexB (Ki > 100 µM). Furthermore, to investigate the inhibitory mechanism, the possible binding modes of 11c and 15k with OfHex1 were deduced based on molecular docking analysis. This work may provide useful structural starting points for further rational design of potent inhibitors of OfHex1.

MK-8719, a Novel and Selective O-GlcNAcase Inhibitor That Reduces the Formation of Pathological Tau and Ameliorates Neurodegeneration in a Mouse Model of Tauopathy.

Deposition of hyperphosphorylated and aggregated tau protein in the central nervous system is characteristic of Alzheimer disease and other tauopathies. Tau is subject to O-linked N-acetylglucosamine (O-GlcNAc) modification, and O-GlcNAcylation of tau has been shown to influence tau phosphorylation and aggregation. Inhibition of O-GlcNAcase (OGA), the enzyme that removes O-GlcNAc moieties, is a novel strategy to attenuate the formation of pathologic tau. Here we described the in vitro and in vivo pharmacological properties of a novel and selective OGA inhibitor, MK-8719. In vitro, this compound is a potent inhibitor of the human OGA enzyme with comparable activity against the corresponding enzymes from mouse, rat, and dog. In vivo, oral administration of MK-8719 elevates brain and peripheral blood mononuclear cell O-GlcNAc levels in a dose-dependent manner. In addition, positron emission tomography imaging studies demonstrate robust target engagement of MK-8719 in the brains of rats and rTg4510 mice. In the rTg4510 mouse model of human tauopathy, MK-8719 significantly increases brain O-GlcNAc levels and reduces pathologic tau. The reduction in tau pathology in rTg4510 mice is accompanied by attenuation of brain atrophy, including reduction of forebrain volume loss as revealed by volumetric magnetic resonance imaging analysis. These findings suggest that OGA inhibition may reduce tau pathology in tauopathies. However, since hundreds of O-GlcNAcylated proteins may be influenced by OGA inhibition, it will be critical to understand the physiologic and toxicological consequences of chronic O-GlcNAc elevation in vivo. SIGNIFICANCE STATEMENT: MK-8719 is a novel, selective, and potent O-linked N-acetylglucosamine (O-GlcNAc)-ase (OGA) inhibitor that inhibits OGA enzyme activity across multiple species with comparable in vitro potency. In vivo, MK-8719 elevates brain O-GlcNAc levels, reduces pathological tau, and ameliorates brain atrophy in the rTg4510 mouse model of tauopathy. These findings indicate that OGA inhibition may be a promising therapeutic strategy for the treatment of Alzheimer disease and other tauopathies.

Synthesis of Pyrrolidine Monocyclic Analogues of Pochonicine and Its Stereoisomers: Pursuit ofSimplified Structures and Potent ß-N-Acetylhexosaminidase Inhibition.

Ten pairs of pyrrolidine analogues of pochonicine and its stereoisomers have been synthesized from four enantiomeric pairs of polyhydroxylated cyclic nitrones. Among the ten N-acetylamino pyrrolidine analogues, only compounds with 2,5-dideoxy-2,5-imino-d-mannitol (DMDP) and pochonicine (1) configurations showed potent inhibition of ß-N-acetylhexosaminidases (ß-HexNAcases); while 1-amino analogues lost almost all their inhibitions towards the tested enzymes. The assay results reveal the importance of the N-acetylamino group and the possible right configurations of pyrrolidine ring required for this type of inhibitors.

Evidence of a compensatory regulation of colonic O-GlcNAc transferase and O-GlcNAcase expression in response to disruption of O-GlcNAc homeostasis.

O-GlcNAcylation is a post-translational modification of thousands of intracellular proteins that dynamically regulates many fundamental cellular processes. Cellular O-GlcNAcylation levels are regulated by a unique couple of enzymes: O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA), which adds and removes the GlcNAc residue, respectively. Maintenance of O-GlcNAc homeostasis is essential to ensure optimal cellular function and disruption of this homeostasis has been linked to the etiology of several human diseases including cancer. The mechanisms through which the cell maintains O-GlcNAc homeostasis are not fully understood but several studies have suggested that a reciprocal regulation of OGT and OGA expression could be one of them. In this study, we investigated the putative regulation of OGT and OGA expression in response to disruption in O-GlcNAc homeostasis in colon. We provide in vitro and in vivo evidences that in colon cells, modulation of O-GlcNAcylation levels leads to a compensatory regulation of OGT and OGA expression in an attempt to restore basal O-GlcNAcylation levels. Our results also suggests that the regulation of colonic OGA expression in response to changes in O-GlcNAc homeostasis occurs mostly at the transcriptional level whereas OGT regulation seems to rely mainly on post-transcriptional mechanisms.

O-GlcNAc stimulation: A new metabolic approach to treat septic shock.

Septic shock is a systemic inflammation associated with cell metabolism disorders and cardiovascular dysfunction. Increases in O-GlcNAcylation have shown beneficial cardiovascular effects in acute pathologies. We used two different rat models to evaluate the beneficial effects of O-GlcNAc stimulation at the early phase of septic shock. Rats received lipopolysaccharide (LPS) to induce endotoxemic shock or saline (control) and fluid resuscitation (R) with or without O-GlcNAc stimulation (NButGT-10 mg/kg) 1 hour after shock induction. For the second model, rats received cecal ligature and puncture (CLP) surgery and fluid therapy with or without NButGT. Cardiovascular function was evaluated and heart and blood samples were collected and analysed. NButGT treatment efficiently increased total O-GlcNAc without modification of HBP enzyme expression.Treatment improved circulating parameters and cardiovascular function in both models, and restored SERCA2a expression levels. NButGT treatment also reduced animal mortality. In this study, we demonstrate that in septic shock O-GlcNAc stimulation improves global animal and cardiovascular function outcomes associated with a restoration of SERCA2a levels. This pre-clinical study opens avenues for a potential therapy of early-stage septic shock.

Discovery of MK-8719, a Potent O-GlcNAcase Inhibitor as a Potential Treatment for Tauopathies.

Inhibition of O-GlcNAcase (OGA) has emerged as a promising therapeutic approach to treat tau pathology in neurodegenerative diseases such as Alzheimer's disease and progressive supranuclear palsy. Beginning with carbohydrate-based lead molecules, we pursued an optimization strategy of reducing polar surface area to align the desired drug-like properties of potency, selectivity, high central nervous system (CNS) exposure, metabolic stability, favorable pharmacokinetics, and robust in vivo pharmacodynamic response. Herein, we describe the medicinal chemistry and pharmacological studies that led to the identification of (3aR,5S,6S,7R,7aR)-5-(difluoromethyl)-2-(ethylamino)-3a,6,7,7a-tetrahydro-5H-pyrano[3,2-d]thiazole-6,7-diol 42 (MK-8719), a highly potent and selective OGA inhibitor with excellent CNS penetration that has been advanced to first-in-human phase I clinical trials.

Discovery of Novel Inhibitors Targeting Human O-GlcNAcase: Docking-Based Virtual Screening, Biological Evaluation, Structural Modification, and Molecular Dynamics Simulation.

ß-N-Acetylhexosaminidases have emerged as promising targets for drug and pesticide discovery due to their critical physiological functions in various cellular processes. In particular, human O-GlcNAcase (hOGA) from the glycoside hydrolase family 84 (GH84) has gained significant attention. This enzyme was found to be linked to various diseases such as diabetes, cancer, and Alzheimer's disease (AD). In this study, to develop novel hOGA inhibitors with suitable pharmaceutical properties, virtual screening of the Drugbank database was performed using a docking-based approach targeting hOGA. Chlorhexidine (4, Ki = 4.0 µM) was identified as a potent hOGA inhibitor with excellent selectivity (Ki > 200 µM against human ß-N-acetylhexosaminidase B) and subjected to structural modifications and SAR studies. Furthermore, molecular dynamics simulations as well as binding free energy and free energy decomposition calculations were carried out to investigate the basis for the efficiency of potent inhibitors against hOGA. This present work revealed the new application of the disinfectant chlorhexidine and provided useful information for the future design of hOGA inhibitors.

Antiadhesive Nanosomes Facilitate Targeting of the Lysosomal GlcNAc Salvage Pathway through Derailed Cancer Endocytosis.

Activated endocytosis of extracellular macromolecules and their intracellular trafficking to lysosomes is an essential metabolic mechanism in cancer cells during their rapid proliferation. Cancer cells reuse a vast amount of N-acetylglucosamine (GlcNAc) supplied from the GlcNAc salvage pathway for the accelerated synthesis of a pivotal uridine diphosphate (UDP)-GlcNAc. A method to inactivate key glycosidases in lysosomes could critically contribute to the development of potent anticancer therapy. Here we demonstrate that "nanosomes" made of core metals covered by an antiadhesive mixed self-assembled monolayer allow for avoiding nonspecific surface protein corona and targeted molecular delivery through activated endocytosis. Nanosomes carrying suicide substrates showed that lysosomal glycosidases such as ß-hexosaminidase and ß-galactosidase in cancer cells are promising targets for novel anticancer therapeutic nanomedicine that induce apoptotic cell death through lysosomal membrane permeabilization. The advantage of this method is evident because multivalent surface loading by antiadhesive nanosomes makes it possible to highlight "weak interactions" such as carbohydrate-lectin interactions independent of surface protein corona.

Grifola frondosa extract and ergosterol reduce allergic reactions in an allergy mouse model by suppressing the degranulation of mast cells.

The increasing number of patients suffering from allergic diseases is a global health problem. Grifola frondosa is an edible mushroom consumed as a health food in Asia, and has recently been reported to have anti-allergic effects. We previously reported that G. frondosa extract (GFE) and its active components, ergosterol and its derivatives, inhibited the antigen-induced activation of RBL-2H3 cells. Here, we demonstrated that GFE and ergosterol also had an inhibitory effect on the degranulation of bone marrow-derived mast cells (BMMCs) and alleviated anaphylactic cutaneous responses in mice. Using an air pouch-type allergic inflammation mouse model, we confirmed that oral administration of GFE and ergosterol suppressed the degranulation of mast cells in vivo. Our findings suggest that G. frondosa, including ergosterol as its active component, reduces type I allergic reactions by suppressing mast cell degranulation in mice, and might be a novel functional food that prevents allergic diseases.

New insights into mechanism of bisphenol analogue neurotoxicity: implications of inhibition of O-GlcNAcase activity in PC12 cells.

Bisphenol analogues including bisphenol A and its derivatives are ubiquitous environmental contaminants and have been linked to adverse neurodevelopment effects on animals and humans. Most toxicological research focused on estrogen receptor mediated pathways and did not comprehensively clarify the observed toxicity. O-GlcNAcase (OGA), the highest level in brain, plays a critical role in controlling neuronal functions at multi-levels from molecule to animal behaviors. In this work, we intend to investigate the underlying molecular mechanisms for the neurotoxicity of bisphenol analogues by identifying their cellular targets and the resultant effects. The inhibitory actions of seven bisphenol analogues on the OGA activity at molecular level were investigated by our developed electrochemical biosensor. We found that their potency varied with substituent groups, in which tetrabromo bisphenol A (TBBPA) was the strongest. The seven bisphenol analogues (0-100 µM exposure) significantly inhibited OGA activity and up-regulated protein O-GlcNAcylation level in PC12 cells. Inhibition of OGA by bisphenol analogues further induced intracellular calcium, ROS, inflammation, repressed proliferation, interfered with cell cycle, induced apoptosis. And especially, 10 µM tetrabromo bisphenol A (TBBPA) exposure could impair the growth and development of neurite in human neural stem cells (hNSCs). Molecular docking for OGA/bisphenol analogue complexes revealed the hydrophobicity-dominated inhibition potency. OGA, as a new cellular target of bisphenol analogues, would illuminate the molecular mechanism of bisphenol analogues neurotoxicity.

Anti-allergic activity of mung bean (Vigna radiata (L.) Wilczek) protein hydrolysates produced by enzymatic hydrolysis using non-gastrointestinal and gastrointestinal enzymes.

Mung bean seed is a well-known plant protein consumed in Asian countries but the protein is usually retrieved as a waste product during starch production. This study investigated the anti-allergic property of mung bean protein hydrolysates (MBPH) produced by enzymatic hydrolysis using non-gastrointestinal (non-GI), GI and a combination of non-GI+GI enzymes. The hydrolysates were investigated for any anti-allergic property by detecting the amount of ß-hexosaminidase released in RBL-2H3 cells, and complemented with the MTT assay to show cell viability. It was found that MBPH hydrolyzed by a combination of flavourzyme (non-GI enzyme) and pancreatin (GI enzyme) exhibited the highest anti-allergic activity (135.61%), followed by those produced with alcalase, a non-GI enzyme (121.74%) and 80.32% for pancreatin (GI enzyme). Minimal toxicity (<30%) of all hydrolysates on RBL-2H3 cells line was observed. The results suggest that MBPH can potentially serve as a hypoallergenic food ingredient or supplement. PRACTICAL APPLICATIONS: Mung bean (Vigna radiata L. (Wilczek)) is also known as "green gram" and it is an excellent source of protein. The major mung bean storage proteins are the globulin, albumin and legumin, which are also referred to as legume allergens. Our study showed that mung bean peptides obtained after enzymatic hydrolysis influenced ß-hexosaminidase inhibition without any toxic effect on RBL-2H3 cells. This indicates that mung bean allergenicity can be reduced after enzymatic hydrolysis and the protein hydrolysates could be as a hypoallergic food, ingredient, supplement and/or protein substitute in the formulation of food products.