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Objective: Melinjo (Gnetum gnemon L.) seeds are known to contain resveratrol, which are classified as a phenolic compound of the stilbenoid. Melinjoseeds have high water content, making them unstable to be stored for prolonged periods with open packaging at room temperature. The present studyaimed to explore the use of ultrasonic-assisted extraction with a betaine-based natural deep eutectic solvent (NADES) for resveratrol extraction fromirradiated melinjo seeds.Methods: The best betaine-based NADES component was identified among betaine-urea, betaine-lactic acid, and betaine-malic acid. Optimization ofextraction methods was performed using the best NADES and extraction variables such as time of extraction, water percentage, and sample: solventratio. The outcome of extraction was evaluated by measuring resveratrol content using high-performance liquid chromatography and the results wereanalyzed using response surface methodology.Results: The best betaine-based NADES was found to be betaine-lactic acid, yielding a resveratrol content of 0.3344 mg/g powder. The optimumextraction was achieved in 10 min with 60% water and a sample: solvent ratio of 1:10, yielding a resveratrol content of 0.227 mg/g powder.Conclusion: Betaine-based NADES can be purposed as an alternative solvent for resveratrol extraction from irradiated melinjo seeds.

Resveratrol has been extensively studied to investigate its biological effects, including its chemopreventive potential against cancer. Over the past decade, various resveratrol oligomers, both naturally occurring and synthetic, have been described. These resveratrol oligomers result from the polymerization of two or more resveratrol units to form dimers, trimers, tetramers, or even more complex derivatives. Some oligomers appear to have antitumor activities that are similar or superior to monomeric resveratrol. In this review, we discuss resveratrol oligomers with anticancer potential, with emphasis on well-characterized compounds, such as the dimer gnetin-C and other oligomers from Gnetum gnemon, whose safety, pharmacokinetic, and biological activities have been studied in humans.

In this study, antioxidant activities and identification of the bioactive substances in Gnetum gnemon L. (Gg) seed hard shell were evaluated. The seed of Gnetum gnemon L., an Indonesian native plant, is commonly consumed as a vegetable or further processed as cracker. Isolated substances from Gnetum gnemon seed are mainly stilbenoid derivatives which show potent antioxidant, tyrosinase inhibitor, and antimicrobial activities. Nevertheless, the antioxidant activity of its crude extract is still considered weak. In this study, an effort was made to improve antioxidant potency by fractionation using macroporous adsorptive resin (MAR). This fractionation successfully enhanced antioxidant activity of red Gg seed hard shell extract with efficient adsorption contact time within 30 min. Antioxidant activity of fractions 25–75% v/v ethanol increased three- to sevenfold as compared to crude extract and more importantly resulted in dry product which was easier for further processes. Identification of bioactive compounds in Gg seed hard shell extract with different degrees of ripeness was also performed by HPLC and confirmed the presence of Gnetin C, resveratrol, and other stilbenoid derivatives. These other stilbenoid derivatives could be the main substances contributing in antioxidant action with lower IC50 as compared to both Gnetin C and resveratrol. In summary, fractionation process using MAR HPD-600 reduced unnecessary sugar molecules from red Gg seed hard shell extract hence resulting to fraction with strong antioxidant activity.

Gnetum gnemon is an arboreal dioecious plant that is cultivated in Indonesia. The seeds of this species mainly contain dimeric stilbenoid compounds [gnetin C (1), gnemonoside A (2), and gnemonoside D (3)] along with trans-resveratrol (4). trans-Resveratrol has been reported to have antiaging, anticancer, and antidiabetic effects, as well as being a calorie restriction mimetic. SIRT1 exerts a protective effect against vascular senescence. In this study, the effects of these four main stilbenoid derivatives of a G. gnemon seed endosperm ethanolic extract on endothelial senescence were investigated. In streptozotocin-induced diabetic mice, administration of the G. gnemon ethanolic extract increased SIRT1 and decreased endothelial senescence. The concentration of 1 in blood plasma was 6-fold higher than 4 in these mice. Next, the in vitro effects of the four main stilbenoid derivatives of G. gnemon seeds were investigated. Senescent human umbilical vein endothelial cells were induced by hydrogen peroxide. Endothelial senescence was inhibited by 4, which increased the expression of endothelial nitric oxide synthase and SIRT1, whereas 1-3 had no effect. These results indicated that the ethanolic extract of G. gnemon seeds inhibits endothelial senescence, suggesting that 4 plays a critical role in the prevention of endothelial senescence.

Tyrosinase is the key enzyme involved in melanogenesis. The aim of this study was to investigate the in vitro inhibitory effects of gnetin C, a resveratrol dimer isolated from melinjo (Gnetum gnemon) seeds, on tyrosinase activity and melanin biosynthesis in murine B16 cells. The inhibitory activities of gnetin C and resveratrol were shown to be almost equal against tyrosinase and melanin biosynthesis in the cells. The IC(50) values of gnetin C activity against tyrosinase and melanin biosynthesis were 7.0 and 7.6 µM, respectively, whereas resveratrol demonstrated IC(50) values of 7.2 and 7.3 µM, respectively. These results indicated that gnetin C inhibited melanogenesis, in a manner similar to that of resveratrol, by inhibiting tyrosinase and may therefore function as a new skin-whitening agent. However, the direct effects of gnetin C and resveratrol on murine tyrosinase activities were not equal. The IC(50) value of resveratrol was 10.1 µM, while gnetin C only exhibited a 25.2% enzyme inhibition at 16 µM. The IC(25) values for gnetin C and resveratrol were 15.5 and 4.0 µM, respectively. Therefore, it is suggested that the effects of gnetin C may be due to mechanisms other than the direct inhibition of tyrosinase activity.

Melinjo fruit ( Gnetum gnemon L.) has been used as a food in Southeast Asia. To investigate if this fruit has regulatory actions on ileal immune responses, we measured T-helper (Th) cytokine production, i.e., interleukin-2 (IL-2), IL-4, IL-5, and interferon-gamma (IFN- γ), in cultured Peyer's patch (PP) cells from mice orally treated with a methanol extract of melinjo fruit. Oral administration of the 50 % ethanol extract at 100 mg/kg/day significantly enhanced the production of the Th1 cytokines IL-2 and IFN- γ irrespective of concanavalin-A stimulation, whereas the production of the Th2 cytokines IL-4 and IL-5 was not affected. We also isolated seven active constituents accompanied with two new stilbenoids from the ethylacetate fraction of the extracts. The structure of the new stilbene glucosides gnemonoside L (5) and gnemonoside M (7) was determined by 1D and 2D NMR and MS analyses. Five known stilbenoids were identified as resveratrol (1), isorhapontigenin (2), gnemonoside D (4), gnetins C (3) and E (6). Among these tested compounds, only new stilbenoid 7 strongly enhanced Th1 cytokine production in cultured PP cells at 10 mg/kg/day. These results indicated that this melinjo extract and its active constituent 7 potentiated T-cell-dependent immune responses in the ileal mucosa.

Objective: This study aimed to determine the inhibitory activity of 1-butyl-3-methylimidazolium bromide ([bmim]Br) extracts of melinjo seeds(Gnetum gnemon) on dipeptidyl peptidase-4 (DPP-4).Methods: Melinjo seeds were extracted by a [bmim]Br microwave-assisted method using various extraction parameters and the inhibitory activityof DPP-4 of all extracts was determined in 96-well microplates using Cayman inhibitor screening assay. Determination of trans-resveratrol contentwas conducted using a reverse-phase high-performance liquid chromatography method. The total phenolic content was determined using a 96-wellmicroplate reader. Data analysis for the determination of the optimal extraction conditions was developed by response surface methodology.Results: The extract obtained from the third run showed the highest inhibition (28.73%) against DPP-4 activity with the total phenolic content of1.96 mg gallic acid equivalent/g the seed powder.Conclusion: The analytical results revealed the following optimal conditions: Solvent concentration 1.5 M, liquid-solid ratio 23:1, and extraction time15 min.

The identification of fruit-bark of Melinjo was carried out using fractionation column chromatography resulted in 3 isolates A-C with presumption of containing five stilbene derivatives compounds (isorhapontigenin, resveratrol, gnetin D, gnetifolin K, gnetol) and one lignan compound ((+)-lirioresinol B) based on the results of the characterization of UV-Vis, FTIR, and LC-ESI-MS. Isolate C that containing gnetol and (+)-lirioresinol B was reacted through prenylation process with prenyl bromide as a source of prenyl group and K2CO3 as catalyst with reflux system at 60 °C for 24 h. Characterization using LC-ESI-MS showed that gnetol and (+)-lirioresinol B was successfully prenylated with one additional prenyl group. The prenylated stilbene showed comparable antioxidant effect and was slightly lower than the nature stilbene structure, as the impact of the attachment of prenyl moieties at their hydroxyl groups in stilbene, and showed moderate activity against P-388 murine leukemia b cells. The observed inhibition of the bioactivity test provides a reasonable mechanism for the potent cancer chemo preventive activity of prenylated stilbene compounds and may pose this compound as a valuable agent for the treatment of diseases.

Melinjo (Gnetum gnemon L.) seed extracts (MSEs) are rich in resveratrol dimers (gnemonoside A, C, D, gnetin C), trans-resveratrol, and other resveratrol derivatives. trans-Resveratrol is a widely studied caloric restriction mimetic. In mice fed a high-fat diet (HFD), trans-resveratrol protects against obesity, type 2 diabetes, and premature death. Here, treatment of HFD-fed mice with 2.0% MSE significantly reduced body weight gain (p

Four stilbene derivatives, gnemonols K and L (resveratrol trimers), M (isorhapontigenin dimer), and gnemonoside K (glucoside of resveratrol trimer) together with eleven known stilbenoids and a lignan were isolated from the acetone, methanol and 70% methanol soluble parts of the root of Gnetum gnemon (Gnetaceae). The structures of the isolates were determined by spectral analysis. The antioxidant activity of the stilbenoids on lipid peroxide inhibition and super oxide scavenging activity were also investigated.

Objectives: To evaluate the angiotensin converting enzyme (ACE) inhibitory activity of melinjo (Gnetum gnemon) seed extract and to study moleculardocking of stilbene contained in melinjo seeds.Methods: Melinjo seed powders were extracted with n-hexane, dichloromethane, ethyl acetate, methanol, and water successively. The extracts wereevaluated ACE inhibitory activities using ACE kit-Wist and the phenolic content using Folin–Ciocalteu method. The extract demonstrated the highestACE inhibitory activity was subjected to liquid chromatography-mass spectrometry (LC-MS) to know its stilbene constituent. The stilbene constituentsin melinjo seed were performed molecular docking using AutoDock Vina, and ligand-receptor Interactions were processed using Ligand Scout.Results: The ethyl acetate extract demonstrated the highest ACE inhibition activity with inhibitory concentration 50% value of 9.77 × 10−8 μg/mLand the highest total phenolic content (575.9 mg gallic acid equivalent/g). Ultra-performance LC-MS analysis of ethyl acetate extract has detected theexistency of resveratrol, gnetin C, ε-viniferin, and gnemonoside A/B. These compounds displayed similar physiochemical properties to lisinopril (ACEinhibitor), as in silico molecular docking studies demonstrated that they fit into the lisinopril receptors.Conclusion: In vitro analysis ethyl acetate extract from melinjo seeds demonstrated the highest ACE inhibitory activity. Molecular docking analysisindicated that resveratrol dimers, gnetin C and gnemonoside A can be considered ACE inhibitor.Keywords: Angiotensin converting enzyme inhibitor, Gnetum gnemon, Melinjo, Total phenolic, Antihypertension, Molecular docking.

Fruits and seeds of melinjo (Gnetum gnemon L.) are resveratrol derivative-rich materials. Pharmacokinetics of resveratrol derivatives in healthy volunteers after oral administration of 1000 mg of melinjo seed extract (MSE) powder were assessed and compared with those after oral dosing of trans-resveratrol (tRV) powder containing 4.8 mg of tRV only, equivalent to the content in 1000 mg MSE powder. Plasma tRV concentrations with enzymatic hydrolysis were maintained over 24 h, with a tmax of 12 h and a mean residence time (MRT) of 14 h, 5 and 2 times higher than those for tRV powder intake, respectively. Gnetin C, a resveratrol dimer, with hydrolysis was maintained in plasma for >96 h with a 36 h MRT. With repeated doses once daily for 28 days, plasma tRV and gnetin C concentrations with hydrolysis were in good agreement with the theoretical curves. MSE powder was well tolerated up to the oral dosing of 5000 mg with no serious adverse events.

Melinjo (Gnetum gnemonL.) seed extract (MSE) containingtrans-resveratrol (3,5,4′-trihydroxy-trans-stilbene) and other derivatives exerts various beneficial effects. However, its mechanism of action in humans remains unknown. In this study, we aimed to investigate beneficial effects of MSE in healthy adult males. In this double-blind, randomized controlled study, 30 males aged 35–70 years with ≤10% flow-mediated dilatation received placebo or 750 mg MSE powder for 8 weeks, and twenty-nine males (45.1±8.8years old) completed the trial. There was a significant difference in the melinjo and placebo groups. Compared with the placebo control, MSE significantly reduced serum uric acid at 4 weeks and 8 weeks (n=14and 15, resp.). HDL cholesterol was significantly increased in the melinjo group. To clarify the mechanism of MSE for reducing uric acid, we investigated xanthine oxidase inhibitory activity, angiotensin II type 1 (AT1) receptor binding inhibition rate, and agonistic activities for PPARαand PPARγ. MSE,trans-resveratrol, and a resveratrol dimer, gnetin C (GC), significantly inhibit AT1 receptor binding and exhibit mild agonistic activities for PPARαand PPARγ. In conclusion, MSE may decrease serum uric acid regardless of insulin resistance and may improve lipid metabolism by increasing HDL cholesterol.