Water-deficit stress induces prenylated stilbenoid production and affects biomass in peanut hairy roots: Exploring the role of stilbenoid prenyltransferase downregulation.

The peanut plant is one of the most economically important crops around the world. Abiotic stress, such as drought, causes over five hundred million dollars in losses in peanut production per year. Peanuts are known to produce prenylated stilbenoids to counteract biotic stress. However, their role in abiotic stress tolerance has not been elucidated. To address this issue, hairy roots with the capacity to produce prenylated stilbenoids were established. An RNA-interference (RNAi) molecular construct targeting the stilbenoid-specific prenyltransferase AhR4DT-1 was designed and expressed via Agrobacterium rhizogenes-mediated transformation in hairy roots of peanut cultivar Georgia Green. Two transgenic hairy roots with the RNAi molecular construct were established, and the downregulation of AhR4DT-1 was validated using reverse transcriptase quantitative PCR. To determine the efficacy of the RNAi-approach in modifying the levels of prenylated stilbenoids, the hairy roots were co-treated with methyl jasmonate, hydrogen peroxide, cyclodextrin, and magnesium chloride to induce the production of stilbenoids and then the stilbenoids were analyzed in extracts of the culture medium. Highly reduced levels of prenylated stilbenoids were observed in the RNAi hairy roots. Furthermore, the hairy roots were evaluated in a polyethylene glycol (PEG) assay to assess the role of prenylated stilbenoids on water-deficit stress. Upon PEG treatment, stilbenoids were induced and secreted into the culture medium of RNAi and wild-type hairy roots. Additionally, the biomass of the RNAi hairy roots decreased by a higher amount as compared to the wild-type hairy roots suggesting that prenylated stilbenoids might play a role against water-deficit stress.

Effect of peanut stilbenoids, arachidin-1 and arachidin-3, on Streptococcus mutans growth and acid production.

In this study we evaluated the effect of prenylated peanut stilbenoids on the growth, biofilm accumulation and acid production of the dental caries pathogen Streptococcus mutans. Prior research with the non-prenylated stilbenes, resveratrol and piceatannol, has shown that these molecules are active against S. mutans. Here we sought to determine if the addition of a prenyl group to the stilbene backbone increased anti-S. mutans activities. Two prenylated stilbenes, arachidin-1 and arachidin-3, were produced using a peanut hairy root production system. Compared to resveratrol and piceatannol, both arachidin-1 and arachidin-3 led to greater inhibition of S. mutans planktonic growth. This effect also led to reduced biofilm formation, by inhibiting growth, instead of a specific action against biofilm cells. Lastly, sub-MIC concentrations of arachidin-3 reduced the acid production of S. mutans above the 'critical pH' that leads to tooth enamel erosion. In summary, stilbenoids have anti-S. mutans activity, and prenylation enhances this activity.

Arachidin-1, a Prenylated Stilbenoid from Peanut, Enhances the Anticancer Effects of Paclitaxel in Triple-Negative Breast Cancer Cells.

Triple-negative breast cancer (TNBC) is one of the deadliest forms of breast cancer. Investigating alternative therapies to increase survival rates for this disease is essential. To this end, the cytotoxic effects of the prenylated stilbenoids arachidin-1 (A-1) and arachidin-3 (A-3), and non-prenylated resveratrol (RES) were evaluated in human TNBC cell lines as potential adjuvants for paclitaxel (Pac). A-1, alone or in combination with Pac, showed the highest cytotoxicity in TNBC cells. Apoptosis was further evaluated by measuring key apoptosis marker proteins, cell cycle arrest, and intracellular reactive oxygen species (ROS) generation. Furthermore, the cytotoxic effect of A-1 combined with Pac was also evaluated in a 3D spheroid TNBC model. The results showed that A-1 decreased the Pac IC50 approximately 2-fold in TNBC cells. The synergistic combination of A-1 and Pac arrested cells in G2/M phase and activated p53 expression. In addition, the combined treatment increased intracellular ROS generation and induced apoptosis. Importantly, the combination of A-1 with Pac inhibited TNBC spheroid growth. Our results demonstrated that A-1 in combination with Pac inhibited cell proliferation, induced apoptosis through mitochondrial oxidative stress, and reduced TNBC spheroid growth. These findings underscore the impactful effects of the prenylated stilbenoid A-1 as a novel adjuvant for Pac chemotherapy in TNBC treatment.

Correction: Gajurel et al. Production and Secretion of Isowighteone in Hairy Root Cultures of Pigeon Pea (Cajanus cajan) Co-Treated with Multiple Elicitors. Plants 2022, 11, 834.

In the original publication [...].

Induction of the Prenylated Stilbenoids Arachidin-1 and Arachidin-3 and Their Semi-Preparative Separation and Purification from Hairy Root Cultures of Peanut (Arachis hypogaea L.).

Prenylated stilbenoids such as arachidin-1 and arachidin-3 are stilbene derivatives that exhibit multiple pharmacological activities. We report an elicitation strategy using different combinations of cyclodextrin, hydrogen peroxide, methyl jasmonate and magnesium chloride to increase arachidin-1 and arachidin-3 production in peanut hairy root cultures. The treatment of hairy root cultures with cyclodextrin with hydrogen peroxide selectively enhanced arachidin-1 yield (132.6 ± 20.4 mg/L), which was 1.8-fold higher than arachidin-3. Similarly, cyclodextrin combined with methyl jasmonate selectively enhanced arachidin-3 yield (178.2 ± 6.8 mg/L), which was 5.5-fold higher than arachidin-1. Re-elicitation of the hairy root cultures further increased the levels of arachidin-1 and arachidin-3 by 24% and 42%, respectively. The ethyl acetate extract of the culture medium was consecutively fractionated by normal- and reversed-phase column chromatography, followed by semi-preparative HPLC purification on a C18 column to yield arachidin-1 with a recovery rate of 32% and arachidin-3 with a recovery rate of 39%, both at higher than 95% purity. This study provided a sustainable strategy to produce high-purity arachidin-1 and arachidin-3 using hairy root cultures of peanuts combined with column chromatography and semi-preparative HPLC.

Production and Secretion of Isowighteone in Hairy Root Cultures of Pigeon Pea (Cajanus cajan) Co-Treated with Multiple Elicitors.

Isowighteone (3'-isoprenyl genistein) is a prenylated stilbenoid derivative that exhibits neuroprotective, antibacterial, and anti-inflammatory properties. To establish a bioproduction system for this bioactive compound, hairy root cultures of pigeon pea (Cajanus cajan (L.) Millsp.) were developed via Agrobacterium rhizogenes-mediated transformation. The cultures were co-treated with methyl jasmonate, cyclodextrin, hydrogen peroxide, and magnesium chloride to enhance the production of isowighteone. The amount of isowighteone that accumulated in the culture medium upon elicitation varied with the period of elicitation. Isowighteone was purified from extracts of the culture medium by semi-preparative HPLC, and its identity was confirmed by tandem mass spectrometry. After 144 h of elicitation in 12-day-old hairy root culture, the total yield of isowighteone was 8058.618 ± 445.78 µg/g DW, of which approximately 96% was found in the culture medium. The yield of isowighteone in the elicited hairy root culture was approximately 277-fold higher than in the non-elicited root culture. The difference between the phenotypes of the elicited and non-elicited pigeon pea hairy roots was studied using scanning electron microscopy. The non-elicited hairy roots had uniform surfaces whereas the elicited roots had non-uniform shapes. Pigeon pea hairy roots provide a sustainable platform for producing and studying the biosynthesis of isowighteone.

Arachidin-1, a Prenylated Stilbenoid from Peanut, Induces Apoptosis in Triple-Negative Breast Cancer Cells.

Triple-negative breast cancer (TNBC) is unresponsive to typical hormonal treatments, causing it to be one of the deadliest forms of breast cancer. Investigating alternative therapies to increase survival rates for this disease is essential. The goal of this study was to assess cytotoxicity and apoptosis mechanisms of prenylated stilbenoids in TNBC cells. The prenylated stilbenoids arachidin-1 (A-1) and arachidin-3 (A-3) are analogs of resveratrol (RES) produced in peanut upon biotic stress. The anticancer activity of A-1 and A-3 isolated from peanut hairy root cultures was determined in TNBC cell lines MDA-MB-231 and MDA-MB-436. After 24 h of treatment, A-1 exhibited higher cytotoxicity than A-3 and RES with approximately 11-fold and six-fold lower IC50, respectively, in MDA-MB-231 cells, and nine-fold and eight-fold lower IC50, respectively, in MDA-MB-436 cells. A-1 did not show significant cytotoxicity in the non-cancerous cell line MCF-10A. While A-1 blocked cell division in G2-M phases in the TNBC cells, it did not affect cell division in MCF-10A cells. Furthermore, A-1 induced caspase-dependent apoptosis through the intrinsic pathway by activating caspase-9 and PARP cleavage, and inhibiting survivin. In conclusion, A-1 merits further research as a potential lead molecule for the treatment of TNBC.

Elicitation of Stilbenes and Benzofuran Derivatives in Hairy Root Cultures of White Mulberry (Morus alba).

Stilbene and benzofuran derivatives isolated from the root of white mulberry (Morus alba) have shown various biological activities, including anti-inflammatory, antioxidant, and antimicrobial properties. The objectives of this study were to develop hairy root cultures and assess the effect of multiple elicitors combinations including (I) methyl-ß-cyclodextrin (CD), MgCl2, methyl jasmonate (MeJA), and H2O2, (II) CD, MgCl2, and MeJA and (III) CD, MgCl2, and H2O2, on the production of these bioactive compounds. The highest yields of stilbenes and benzofurans were obtained upon co-treatment with 18 g/L CD, 3 mM H2O2 and 1 mM MgCl2. The stilbenes oxyresveratrol, resveratrol, and 3'-prenylresveratrol accumulated up to 6.27, 0.61, and 5.00 mg/g DW root, respectively. Meanwhile, the aryl benzofurans moracin M and moracin C accumulated up to 7.82 and 1.82 mg/g DW root, respectively. These stilbenes and benzofurans accumulated in the culture medium of the elicited hairy root cultures. They were not detected in the root tissue. However, the oxyresveratrol diglucoside mulberroside A was only detected in the root tissue with yields up to 10.01 mg/g DW. The results demonstrated that co-treatment of white mulberry hairy root cultures with multiple elicitors can significantly enhance production and secretion of stilbenes and benzofurans in this controlled and sustainable axenic culture system.

Antioxidant Assessment of Prenylated Stilbenoid-Rich Extracts from Elicited Hairy Root Cultures of Three Cultivars of Peanut (Arachis hypogaea).

Peanut produces prenylated stilbenoids upon biotic stress. However, the role of these compounds against oxidative stress have not been thoroughly elucidated. To this end, the antioxidant capacity of extracts enriched in prenylated stilbenoids and derivatives was studied. To produce these extracts, hairy root cultures of peanut cultivars Hull, Tifrunner, and Georgia Green were co-treated with methyl jasmonate, cyclodextrin, hydrogen peroxide, and magnesium chloride and then the stilbenoids were extracted from the culture medium. Among the three cultivars, higher levels of the stilbenoid derivatives arachidin-1 and arachidin-6 were detected in cultivar Tifrunner. Upon reaction with 2,2-diphenyl-1picrylhydrazyl, extracts from cultivar Tifrunner showed the highest antioxidant capacity with an IC50 of 6.004 µg/mL. Furthermore, these extracts had significantly higher antioxidant capacity at 6.25 µg/mL and 3.125 µg/mL when compared to extracts from cultivars Hull and Georgia Green. The stilbenoid-rich extracts from peanut hairy roots show high antioxidant capacity and merit further study as potential nutraceuticals to promote human health.

Production of Prenylated Stilbenoids in Hairy Root Cultures of Peanut (Arachis hypogaea) and its Wild Relatives A. ipaensis and A. duranensis via an Optimized Elicitation Procedure.

Prenylated stilbenoids are phenolic compounds produced in a small number of plants such as peanut (Arachis hypogaea) to counteract biotic and abiotic stresses. In addition to their role in plant defense, they exhibit biological activities with potential application in human health. Whereas non-prenylated stilbenoids such as resveratrol are commercially available, the availability of prenylated stilbenoids is limited. To this end, hairy root cultures of peanut were developed as an elicitor-controlled bioproduction platform for prenylated stilbenoids. An orthogonal array design approach led to the elucidation of an optimized elicitation procedure consisting of co-treatment of the hairy root cultures with 18 g/L methyl-ß-cyclodextrin, 125 µM methyl jasmonate, 3 mM hydrogen peroxide (H2O2) and medium supplementation with additional 1 mM magnesium chloride. After 168-h of elicitor treatment, the combined yield of the prenylated stilbenoids arachidin-1, arachidin-2, arachidin-3 and arachidin-5 reached approximately 750 mg/L (equivalent to 107 mg/g DW). Moreover, hairy root cultures from the wild Arachis species A. duranensis and A. ipaensis were developed and shown to produce prenylated stilbenoids upon elicitor treatment. These wild Arachis hairy root lines may provide a platform to elucidate the biosynthetic origin of prenylated stilbenoids in peanut.

Stilbenoid prenyltransferases define key steps in the diversification of peanut phytoalexins.

Defense responses of peanut (Arachis hypogaea) to biotic and abiotic stresses include the synthesis of prenylated stilbenoids. Members of this compound class show several protective activities in human disease studies, and the list of potential therapeutic targets continues to expand. Despite their medical and biological importance, the biosynthetic pathways of prenylated stilbenoids remain to be elucidated, and the genes encoding stilbenoid-specific prenyltransferases have yet to be identified in any plant species. In this study, we combined targeted transcriptomic and metabolomic analyses to discover prenyltransferase genes in elicitor-treated peanut hairy root cultures. Transcripts encoding five enzymes were identified, and two of these were functionally characterized in a transient expression system consisting of Agrobacterium-infiltrated leaves of Nicotiana benthamiana We observed that one of these prenyltransferases, AhR4DT-1, catalyzes a key reaction in the biosynthesis of prenylated stilbenoids, in which resveratrol is prenylated at its C-4 position to form arachidin-2, whereas another, AhR3'DT-1, added the prenyl group to C-3' of resveratrol. Each of these prenyltransferases was highly specific for stilbenoid substrates, and we confirmed their subcellular location in the plastid by fluorescence microscopy. Structural analysis of the prenylated stilbenoids suggested that these two prenyltransferase activities represent the first committed steps in the biosynthesis of a large number of prenylated stilbenoids and their derivatives in peanut. In summary, we have identified five candidate prenyltransferases in peanut and confirmed that two of them are stilbenoid-specific, advancing our understanding of this specialized enzyme family and shedding critical light onto the biosynthesis of bioactive stilbenoids.

Crossing kingdoms: Using decellularized plants as perfusable tissue engineering scaffolds.

Despite significant advances in the fabrication of bioengineered scaffolds for tissue engineering, delivery of nutrients in complex engineered human tissues remains a challenge. By taking advantage of the similarities in the vascular structure of plant and animal tissues, we developed decellularized plant tissue as a prevascularized scaffold for tissue engineering applications. Perfusion-based decellularization was modified for different plant species, providing different geometries of scaffolding. After decellularization, plant scaffolds remained patent and able to transport microparticles. Plant scaffolds were recellularized with human endothelial cells that colonized the inner surfaces of plant vasculature. Human mesenchymal stem cells and human pluripotent stem cell derived cardiomyocytes adhered to the outer surfaces of plant scaffolds. Cardiomyocytes demonstrated contractile function and calcium handling capabilities over the course of 21 days. These data demonstrate the potential of decellularized plants as scaffolds for tissue engineering, which could ultimately provide a cost-efficient, "green" technology for regenerating large volume vascularized tissue mass.

A Stilbenoid-Specific Prenyltransferase Utilizes Dimethylallyl Pyrophosphate from the Plastidic Terpenoid Pathway.

Prenylated stilbenoids synthesized in some legumes exhibit plant pathogen defense properties and pharmacological activities with potential benefits to human health. Despite their importance, the biosynthetic pathways of these compounds remain to be elucidated. Peanut (Arachis hypogaea) hairy root cultures produce a diverse array of prenylated stilbenoids upon treatment with elicitors. Using metabolic inhibitors of the plastidic and cytosolic isoprenoid biosynthetic pathways, we demonstrated that the prenyl moiety on the prenylated stilbenoids derives from a plastidic pathway. We further characterized, to our knowledge for the first time, a membrane-bound stilbenoid-specific prenyltransferase activity from the microsomal fraction of peanut hairy roots. This microsomal fraction-derived resveratrol 4-dimethylallyl transferase utilizes 3,3-dimethylallyl pyrophosphate as a prenyl donor and prenylates resveratrol to form arachidin-2. It also prenylates pinosylvin to chiricanine A and piceatannol to arachidin-5, a prenylated stilbenoid identified, to our knowledge, for the first time in this study. This prenyltransferase exhibits strict substrate specificity for stilbenoids and does not prenylate flavanone, flavone, or isoflavone backbones, even though it shares several common features with flavonoid-specific prenyltransferases.

Investigation of Stilbenoids as Potential Therapeutic Agents for Rotavirus Gastroenteritis.

Rotavirus (RV) infections cause severe diarrhea in infants and young children worldwide. Vaccines are available but cost prohibitive for many countries and only reduce severe symptoms. Vaccinated infants continue to shed infectious particles, and studies show decreased efficacy of the RV vaccines in tropical and subtropical countries where they are needed most. Continuing surveillance for new RV strains, assessment of vaccine efficacy, and development of cost effective antiviral drugs remain an important aspect of RV studies. This study was to determine the efficacy of antioxidant and anti-inflammatory stilbenoids to inhibit RV replication. Peanut (A. hypogaea) hairy root cultures were induced to produce stilbenoids, which were purified by high performance countercurrent chromatography (HPCCC) and analyzed by HPLC. HT29.f8 cells were infected with RV in the presence stilbenoids. Cell viability counts showed no cytotoxic effects on HT29.f8 cells. Viral infectivity titers were calculated and comparatively assessed to determine the effects of stilbenoid treatments. Two stilbenoids, trans-arachidin-1 and trans-arachidin-3, show a significant decrease in RV infectivity titers. Western blot analyses performed on the infected cell lysates complemented the infectivity titrations and indicated a significant decrease in viral replication. These studies show the therapeutic potential of the stilbenoids against RV replication.

Enhanced Production of Resveratrol, Piceatannol, Arachidin-1, and Arachidin-3 in Hairy Root Cultures of Peanut Co-treated with Methyl Jasmonate and Cyclodextrin.

Peanut (Arachis hypogaea) produces stilbenoids upon exposure to abiotic and biotic stresses. Among these compounds, the prenylated stilbenoids arachidin-1 and arachidin-3 have shown diverse biological activities with potential applications in human health. These compounds exhibit higher or novel biological activities in vitro when compared to their nonprenylated analogues piceatannol and resveratrol, respectively. However, assessment of these bioactivities in vivo has been challenging because of their limited availability. In this study, hairy root cultures of peanut were induced to produce stilbenoids upon treatment with elicitors. Co-treatment with 100 µM methyl jasmonate (MeJA) and 9 g/L methyl-ß-cyclodextrin (CD) led to sustained high levels of resveratrol, piceatannol, arachidin-1, and arachidin-3 in the culture medium when compared to other elicitor treatments. The average yields of arachidin-1 and arachidin-3 were 56 and 148 mg/L, respectively, after co-treatment with MeJA and CD. Furthermore, MeJA and CD had a synergistic effect on resveratrol synthase gene expression, which could explain the higher yield of resveratrol when compared to treatment with either MeJA or CD alone. Peanut hairy root cultures were shown to be a controlled and sustainable axenic system for the production of the diverse types of biologically active stilbenoids.

Effect of light, methyl jasmonate and cyclodextrin on production of phenolic compounds in hairy root cultures of Scutellaria lateriflora.

Scutellaria lateriflora (American skullcap) has been used in traditional medicine to treat several medical conditions including nervous disorders and cancer. Previous studies have associated these medicinal properties to flavones present in roots and leaves of this species. In order to develop a production system and study the biosynthesis of these bioactive compounds, hairy root cultures of S. lateriflora were established and line 4 was selected for further studies based on its growth performance in a modified Murashige and Skoog's medium supplemented with 0.5mg/l indole-3-butyric acid. Scanning electron microscopy of the hairy roots showed a high profusion of hairs along the root. Several phenolic compounds, including verbascoside, and the flavones wogonin, baicalein, scutellarein and their respective glucuronides were identified by high performance liquid chromatography-tandem mass spectrometry in the root tissue, but not in the culture medium. Among these compounds, verbascoside accumulated at the highest levels. Interestingly, cultures incubated under continuous light and treated with 15mM methyl-ß-cyclodextrin for 24h produced significantly higher levels of the aglycones, baicalein and wogonin, but not scutellarein, compared to cultures incubated under continuous darkness. This work demonstrates that hairy root cultures of S. lateriflora have the biosynthetic capacity to produce known Scutellaria flavones and suggest that light may have a selected regulatory effect on the synthesis or accumulation of these phenolic compounds.

Antimalarial evaluation of the chemical constituents of hairy root culture of Bixa orellana L.

Over 216 million malaria cases are reported annually worldwide and about a third of these cases, primarily children under the age of five years old, will not survive the infection. Despite this significant world health impact, only a limited number of therapeutic agents are currently available. The lack of scaffold diversity poses a threat in the event that multi-drug-resistant strains emerge. Terrestrial natural products have provided a major source of chemical diversity for starting materials in many FDA approved drugs over the past century. Bixa orellana L. is a popular plant used in South America for the treatment of malaria. In search of new potential therapeutic agents, the chemical constituents of a selected hairy root culture line of Bixa orellana L. were characterized utilizing NMR and mass spectrometry methods, followed by its biological evaluation against malaria strains 3D7 and K1. The crude extract and its isolated compounds demonstrated EC50 values in the micromolar range. Herein, we report our findings on the chemical constituents of Bixa orellana L. from hairy roots responsible for the observed antimalarial activity.

Differential induction of antioxidant stilbenoids in hairy roots of Vitis rotundifolia treated with methyl jasmonate and hydrogen peroxide.

Stilbenoids are polyphenolic phytoalexins that exhibit potential health applications in humans. Hairy root cultures of muscadine grape (Vitis rotundifolia Michx.) were used to study the biochemical and molecular regulation of stilbenoid biosynthesis upon treatment with 100 µM methyl jasmonate (MeJA) or 10 mM hydrogen peroxide (H2O2) over a 96-h period. Resveratrol, piceid, and ε-viniferin were identified in higher concentrations in the tissue whereas resveratrol was the most abundant stilbenoid in the medium under either treatment. An earlier increase in resveratrol accumulation was observed for the MeJA-treated group showing a maximum at 12 h in the tissue and 18 h in the medium. Furthermore, the antioxidant capacity of extracts from the tissue and medium was determined by the 2,2'-azinobis[3-ethylbenzthiazoline sulfonic acid] (ABTS) and the 2,2-diphenyl-1-picrylhydrazyl (DPPH) assays showing correlation with the stilbenoid content. Fourteen candidate reference genes for qPCR were tested under the described experimental conditions and resulted in the selection of 5 reference genes. Quantitative analyses of transcripts for phenylalanine ammonia-lyase (PAL), resveratrol synthase (RS), and two stilbene synthases (STS and STS2) showed the highest RNA level induction at 3 h for both treatments with a higher induction for the MeJA treatment. In contrast, the flavonoid-related chalcone synthase (CHS) transcripts showed induction and a decrease in expression for MeJA and H2O2 treatments, respectively. The observed responses could be related to an oxidative burst triggered by the exposure to abiotic stressor compounds with signaling function such as MeJA and H2O2 which have been previously related to the synthesis of secondary metabolites.

Antioxidant activity of selected stilbenoids and their bioproduction in hairy root cultures of muscadine grape (Vitis rotundifolia Michx.).

Stilbenoids are polyphenolic phytoalexins with health-related properties in humans. Muscadine grape ( Vitis rotundifolia ) hairy root cultures were established via Agrobacterium rhizogenes -mediated transformation, and the effects of growth regulators (3-indolebutyric acid and 6-benzylaminopurine) and methyl jasmonate (MeJA) on stilbenoid production were studied. Twenty-one-day-old hairy root cultures were treated with 100 µM MeJA for 24 h, and then the stilbenoids were extracted from the medium and tissue with ethyl acetate and analyzed by HPLC. Resveratrol, piceid, and ε-viniferin were observed preferentially in tissue, whereas piceatannol was observed only in medium. Growth regulators did not affect the yield of stilbenoids, whereas higher levels were found upon treatment with MeJA. Stilbenoids identified in the hairy root cultures were analyzed for their radical scavenging capacity showing piceatannol and ε-viniferin as the strongest antioxidants. Muscadine grape hairy root cultures were demonstrated to be amenable systems to study stilbenoid biosynthesis and a sustainable source of these bioactive compounds.

Natural prenylated resveratrol analogs arachidin-1 and -3 demonstrate improved glucuronidation profiles and have affinity for cannabinoid receptors.

RATIONALE: The therapeutic promise of trans-resveratrol (tRes) is limited by poor bioavailability following rapid metabolism. We hypothesise that trans-arachidin-1 (tA1) and trans-arachidin-3 (tA3), peanut hairy root-derived isoprenylated analogs of tRes, will exhibit slower metabolism/enhanced bioavailability and retain biological activity via cannabinoid receptor (CBR) binding relative to their non-prenylated parent compounds trans-piceatannol (tPice) and tRes, respectively. RESULTS: The activities of eight human UDP-glucuronosyltransferases (UGTs) toward these compounds were evaluated. The greatest activity was observed for extrahepatic UGTs 1A10 and 1A7, followed by hepatic UGTs 1A1 and 1A9. Importantly, an additional isoprenyl and/or hydroxyl group in tA1 and tA3 slowed overall glucuronidation. CBR binding studies demonstrated that all analogs bound to CB1Rs with similar affinities (5-18 µM); however, only tA1 and tA3 bound appreciably to CB2Rs. Molecular modelling studies confirmed that the isoprenyl moiety of tA1 and tA3 improved binding affinity to CB2Rs. Finally, although tA3 acted as a competitive CB1R antagonist, tA1 antagonised CB1R agonists by both competitive and non-competitive mechanisms. CONCLUSIONS: Prenylated stilbenoids may be preferable alternatives to tRes due to increased bioavailability via slowed metabolism. Similar structural analogs might be developed as novel CB therapeutics for obesity and/or drug dependency.