Allelopathy and Identification of Five Allelochemicals in the Leaves of the Aromatic Medicinal Tree Aegle marmelos (L.) Correa.

Aegle marmelos (L.) Correa is an economically and therapeutically valuable tree. It is cultivated as a fruit plant in southeast Asian countries. In this research, we investigated the allelopathy and possible allelochemicals in the leaves of A. marmelos. Aqueous methanol extracts of A. marmelos exhibited significant inhibitory effects against the growth of Lepidium sativum, Lactuca sativa, Medicago sativa, Echinochloa crusgalli, Lolium multiflorum, and Phleum pratense. Bioassay-directed chromatographic purification of the A. marmelos extracts resulted in identifying five active compounds: umbelliferone (1), trans-ferulic acid (2), (E)-4-hydroxycinnamic acid methyl ester (3), trans-cinnamic acid (4), and methyl (E)-3'-hydroxyl-4'-methoxycinnamate (5). The hypocotyl and root growth of L. sativum were considerably suppressed by these compounds. Methyl (E)-3'-hydroxyl-4'-methoxycinnamate also suppressed the coleoptile and root growth of E. crusgalli. The concentrations of these compounds, causing 50% growth reduction (I50) of L. sativum, were in the range of 74.19-785.4 µM. The findings suggest that these isolated compounds might function in the allelopathy of A. marmelos.

Isolation and Identification of Allelopathic Substances from Forsythia suspensa Leaves, and Their Metabolism and Activity.

The fruit of Forsythia suspensa (Thunb.) Vahl has been used in traditional Chinese medicine as "Forsythiae fructus". The species is also grown in parks and gardens, and on streets and building lots, as an ornamental plant, but it requires pruning. In this study, the allelopathic activity and allelopathic substances in the leaves of pruned branches of F. suspensa were investigated to determine any potential application. The leaf extracts of F. suspensa showed growth inhibitory activity against three weed species; Echinochloa crus-galli, Lolium multiflorum, and Vulpia myuros. Two allelopathic substances in the extracts were isolated through the bioassay-guided purification process, and identified as (-)-matairesinol and (-)-arctigenin. (-)-Matairesinol and (-)-arctigenin, which showed significant growth inhibitory activity at concentrations greater than 0.3 mM in vitro. The inhibitory activity of (-)-arctigenin was greater than that of (-)-matairesinol. However, both compounds were more active than (+)-pinolesinol which is their precursor in the biosynthetic pathway. The investigation suggests that F. suspensa leaves are allelopathic, and (-)-matairesinol and (-)-arctigenin may contribute to the growth inhibitory activities. Therefore, the leaves of the pruned branches can be applied as a weed management strategy in some agricultural practices such as using the leaf extracts in a foliar spray and the leaves in a soil mixture, thereby reducing the dependency on synthetic herbicides in the crop cultivation and contributing to developing eco-friendly agriculture.

Synergistic effect of secondary metabolites isolated from Pestalotiopsis sp. FKR-0115 in overcoming ß-lactam resistance in MRSA.

Six aromatic secondary metabolites, pestalone (1), emodin (2), phomopsilactone (3), pestalachlorides B (4), C (5), and D (6), were isolated from Pestalotiopsis sp. FKR-0115, a filamentous fungus collected from white moulds growing on dead branches in Minami Daito Island. The efficacy of these secondary metabolites against methicillin-resistant Staphylococcus aureus (MRSA) with and without meropenem (ß-lactam antibiotic) was evaluated using the paper disc method and broth microdilution method. The chemical structures of the isolated compounds (1-6) were characterised using spectroscopic methods, including nuclear magnetic resonance and mass spectrometry. All six isolated compounds exhibited synergistic activity with meropenem against MRSA. Among the six secondary metabolites, pestalone (1) overcame bacterial resistance in MRSA to the greatest extent.

Allelopathic Activity of a Novel Compound, Two Known Sesquiterpenes, and a C13Nor-Isopenoid from the Leave of Croton oblongifolius Roxb. for Weed Control.

Investigation of allelopathic substances from herbal plants may lead to the development of allelochemical-based natural herbicides. Croton oblongifolius (Roxb.) is a well-known herbal plant with a long history of being used for traditional medicines and for being the source of a diverse range of bioactive compounds. This plant has been reported to have allelopathic potential; however, its allelopathic-related substances have not yet been described. Therefore, we conducted this investigation to explore the allelopathic substances from the leaves of C. oblongifolius. Aqueous methanol extracts of C. oblongifolius leaves exhibited significant growth inhibitory potential against four test plants (monocot barnyard grass and timothy, and dicot cress and lettuce). The leaf extracts were purified in various chromatographic steps and yielded four active compounds identified as (3R,6R,7E)-3-hydroxy-4-7-megastigmadien-9-one (I), 2-hydroxy alpinolide (a novel compound) (II), alpinolide (III), and epialpinolide (IV) via an analysis of the spectral data. These identified compounds significantly restricted the seedling growth of cress. The concentration necessary for 50% growth reduction of the cress seedlings varied from 0.15 to 0.24 mM for (3R,6R,7E)-3-hydroxy-4-7-megastigmadien-9-one, 0.04 to 0.11 mM for 2-hydroxy alpinolide, 0.07 to 0.12 mM for alpinolide, and 0.09 to 0.16 mM for epialpinolide. Therefore, the leaf extracts of C. oblongifolius and the characterized compounds have the potential to be used as weed-suppressive resources for natural weed control.

Phytotoxicity and Phytotoxic Substances in Calamus tenuis Roxb.

Calamus tenuis is a shrub species distributed across South Asia. It grows well in diversified habitats and tends to dominate plants in the surrounding environment. The phytotoxicity of C. tenuis and the action of its phytochemicals against other plant species could explain its dominant behavior. Compounds with phytotoxic activity are in high demand as prospective sources of ecofriendly bioherbicides. Therefore, we investigated the phytotoxicity of C. tenuis. Aqueous methanol extracts of this plant species significantly limited the growth of four test plant species, two monocots (barnyard grass and timothy), and two dicots (alfalfa and cress), in a dose- and species-dependent manner. Bio-directed chromatographic isolation of the C. tenuis extracts yielded two major active substances: a novel compound, calamulactone {(S)-methyl 8-(5-oxo-2,5-dihydrofuran-2-yl) octanoate}, and 3-oxo-α-ionone. Both of the identified compounds exerted strong growth inhibitory effects on cress and timothy seedlings. The concentrations of 3-oxo-α-ionone and calamulactone required to limit the growth of the cress seedlings by 50% (I50) were 281.6-199.5 and 141.1-105.5 µM, respectively, indicating that the effect of calamulactone was stronger with lower I50 values. Similarly, the seedlings of timothy also showed a considerably higher sensitivity to calamulactone (I50: 40.5-84.4 µM) than to 3-oxo-α-ionone (I50: 107.8-144.7 µM). The findings indicated that the leaves of C. tenuis have marked growth-inhibitory potential, and could affect surrounding plants to exert dominance over the surrounding plant community. Moreover, the two identified phytotoxic substances might play a key role in the phytotoxicity of C. tenuis, and could be a template for bioherbicide development. This paper was the first to report calamulactone and its phytotoxicity.

A novel aromatic compound from the fungus Synnemellisia sp. FKR-0921.

The filamentous fungus Synnemellisia sp. strain FKR-0921 was obtained from soil collected on Kume Island, Okinawa. The MeOH extract of FKR-0921 cultured on a solid rice medium yielded a new aromatic compound, synnemellisitriol A (1). The structure, including the absolute configuration, was elucidated by spectroscopic analysis (FT-IR, NMR, and HR-ESI-MS), and the absolute configuration at C-9 of 1 was determined using the modified Mosher's method. Additionally, 1 was evaluated for its biological activities, including metallo-ß-lactamase inhibitory activity, type III secretion system inhibitory activity, antimicrobial activity, antimalarial activity, and cytotoxicity.

Allelopathic Activity of the Invasive Plant Polygonum chinense Linn. and Its Allelopathic Substances.

Polygonum chinense Linn., belonging to the Polygonaceae family, is distributed mostly in northern temperate climates. This species is a high-risk invasive plant and is thought to possess allelopathic potential. This study aimed to isolate and identify the allelopathic substances from P. chinense. Aqueous methanol extracts of P. chinense significantly inhibited the growth of alfalfa and Italian ryegrass seedlings in a species- and concentration-dependent manner. Activity-guided fractionation led to the isolation of two active compounds: dehydrovomifoliol and loliolide. A cress bioassay was used to determine the biological activity of dehydrovomifoliol, and cress, alfalfa, and Italian ryegrass were used to determine loliolide. Dehydrovomifoliol significantly suppressed the seedling growth of cress at the concentration of 1 mM, and the concentrations necessary for 50% growth inhibition (I50 values) of the roots and shoots were 1.2 and 2 mM, respectively. Loliolide significantly suppressed the shoot growth of cress, alfalfa, and Italian ryegrass at the concentration of 1 mM, and the concentrations necessary for I50 values of the shoots and roots were 0.15 to 2.33 and 0.33 to 2.23 mM, respectively. The findings of our study suggest the extracts of P. chinense might have growth-inhibitory potential and that dehydrovomifoliol and loliolide might contribute as allelopathic agents.

Differentiation-Promoting Effects of Okeaniamides A and B from an Okeania sp. Marine Cyanobacterium on Preadipocytes.

The linear lipopeptides okeaniamide A (1) and okeaniamide B (2) were isolated from an Okeania sp. marine cyanobacterium collected in Okinawa. The structures of these compounds were established by spectroscopic analyses, and the absolute configurations were elucidated based on a combination of chemical degradations, Marfey's analysis, and derivatization reactions. Okeaniamide A (1) and okeaniamide B (2) dose-dependently promoted the differentiation of mouse 3T3-L1 preadipocytes in the presence of insulin.

Studies on the Catechin Constituents of Bark of Cinnamomum sieboldii.

Screening for bioactivity related to anti-infective, anti-methicillin-resistant Staphylococcus aureus (MRSA) and anti-viral activity, led us to identify active compounds from a methanol extract of Litsea japonica (Thub.) Juss. and the hot water extract of bark of Cinnamomum sieboldii Meisn (also known as Karaki or Okinawa cinnamon). The two main components in these extracts were identified as the catechin trimers (+)-cinnamtannin B1 and pavetannin B5. Moreover, these extracts exhibited anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) activity. The structures of these catechin trimers were previously determined by chemical and spectroscopic methods. Pavetanin B5 has never been reported to be isolated as a pure form and has been obtained as a mixture with another component. Although other groups have reported the putative structure of pavetannin B5, preparation of the methylated derivative of pavetannin B5 in this study allowed us to obtain the pure form for the first time as the undecamethyl derivative and confirm its exact structure. Commercially available (+)-cinnamtannin B1 and aesculitannin B (C2'-epimer of cinnamtannin B1) both of which contained pavetannin B5 as a minor component, and C. sieboldii bark extract (approx. 5/2 mixture of (+)-cinnamtannin B1/pavetannin B5) were assessed for anti-SARS-CoV-2 activity. Both C. sieboldii bark extract and commercially available aesculitannin B showed viral growth inhibitory activity.

The Allelopathic Effects of Trewia nudiflora Leaf Extracts and Its Identified Substances.

Trewia nudiflora Linn. is a woody plant of the Euphorbiaceae family. It is well known for its use as a folk remedy, but its potential for phytotoxicity has not been explored. Therefore, this study investigated the allelopathic potential and the allelopathic substances in T. nudiflora leaves. The aqueous methanol extract of T. nudiflora was found to have a toxic effect on the plants used in the experiment. The shoot and root development of lettuce (Lactuca sativa L.) and foxtail fescue (Vulpia myuros L.) were significantly (p ≤ 0.05) reduced by the T. nudiflora extracts. The growth inhibition by the T. nudiflora extracts was proportional to the extract concentration and varied with the test plant species. The chromatographic separation of the extracts resulted in the isolation of two substances, identified as loliolide and 6,7,8-trimethoxycoumarin based on their respective spectral analyses. Both substances significantly inhibited lettuce growth at a concentration of 0.01 mM. To inhibit 50% of the growth of the lettuce, the required concentration of loliolide was 0.043 to 0.128 mM, while that of 6,7,8-trimethoxycoumarin was 0.028 to 0.032 mM. Comparing these values, the lettuce growth was more sensitive to 6,7,8-trimethoxycoumarin than loliolide, suggesting that 6,7,8-trimethoxycoumarin was more effective than loliolide. Therefore, the inhibition of the growth of the lettuce and foxtail fescue suggests that loliolide and 6,7,8-trimethoxycoumarin are responsible for the phytotoxicity of the T. nudiflora leaf extracts. Thus, the growth-inhibitory effectiveness of the T. nudiflora extracts and the identified loliolide and 6,7,8-trimethoxycoumarin may be used to develop bioherbicides that restrict the growth of weeds.

Identification of four allelopathic compounds including a novel compound from Elaeocarpus floribundus Blume and determination of their allelopathic activity.

Allelopathic compounds can play a vital role in protecting the environment from pollution by synthetic herbicides. Compounds isolated from plant species with allelopathic potential can be used as natural herbicides to control weeds and help reduce environmental pollution. Elaeocarpus floribundus has been reported to contain allelopathic compounds. Aqueous methanolic extracts of the leaves of this plant showed strong growth inhibitory potential against two test species (monocotyledonous Italian ryegrass and dicotyledonous alfalfa) in plants- and dose-dependent technique. Several extensive chromatographic separations of the E. floribundus leaf extracts yielded four active compounds 1, 2, 3, and 4 (novel compound). All the identified compounds showed strong growth inhibitory potential against cress. The concentrations caused for 50% growth limitation (I50 values) of the cress seedlings were in the range 500.4-1913.1 µM. The findings indicate that the identified compounds might play a pivotal function in the allelopathic potential of E. floribundus tree. This report is the first on elaeocarpunone and its allelopathic potential.

A new isomaneonene derivative from the red alga Laurencia cf. mariannensis.

Determining the structures of new natural products from marine species not only enriches our understanding of the diverse chemistry of these species, but can also lead to the discovery of compounds with novel and/or important biological activities. Herein, we describe the isolation of isomaneonene C (1), a new halogenated C15 acetogenin, and three known compounds, α-snyderol (2), cis-maneonene D (3), and isomaneonene B (4), from the organic extract obtained from the red alga Laurencia cf. mariannensis collected from Iheya Island, Okinawa, Japan. The structures of these secondary metabolites were elucidated spectroscopically. All compounds were inactive at 30 µg/disc against methicillin-resistant Staphylococcus aureus (MRSA) in combination treatment with a ß-lactam drug, meropenem.

Koshidacins A and B, Antiplasmodial Cyclic Tetrapeptides from the Okinawan Fungus Pochonia boninensis FKR-0564.

Two new antiplasmodial peptides, named koshidacins A (1) and B (2), were discovered from the culture broth of the Okinawan fungus Pochonia boninensis FKR-0564. Their structures, including absolute configurations, were elucidated by a combination of spectroscopic methods and chemical derivatization. Both compounds showed moderate in vitro antiplasmodial activity against Plasmodium falciparum strains, with IC50 values ranging from 17.1 to 0.83 µM. In addition, compound 2 suppressed 41% of malaria parasites in vivo when administered intraperitoneally at a dose of 30 mg/kg/day for 4 days.

Isolation of Caldorazole, a Thiazole-Containing Polyketide with Selective Cytotoxicity under Glucose-Restricted Conditions.

Caldorazole (1) was isolated from the marine cyanobacterium Caldora sp. collected on Ishigaki Island, Okinawa, Japan. Its structure was determined to be a new polyketide that contained two thiazole rings and an O-methylenolpyruvamide moiety. Caldorazole (1) showed strong cytotoxicity toward tumor cells that had been seeded at a high density. Cell death induced by 1 in HeLa and A431 cells was also observed only in the presence of the glycolysis blocker 2-deoxy-d-glucose (2DG). Co-treatment with 1 and 2DG remarkably decreased ATP levels in these cells. Furthermore, 1 selectively inhibited complex I in the mitochondrial respiratory chain. Thus, 1 was demonstrated to exert cytotoxicity toward human tumor cells by blocking mitochondrial respiration.

Odookeanynes A and B, Acetylene-Containing Lipopeptides from an Okeania sp. Marine Cyanobacterium.

Odookeanynes A (1) and B (2), two acetylene-containing lipopeptides, were isolated from an Okeania sp. marine cyanobacterium collected in Okinawa, Japan. Their structures were elucidated by spectroscopic analysis and Marfey's analysis of acid hydrolysates. Odookeanynes A (1) and B (2) dose-dependently promoted the differentiation of mouse 3T3-L1 preadipocytes in the presence of insulin.

Beneficial effects of a nobiletin-rich formulated supplement of Sikwasa (C. depressa) peel on cognitive function in elderly Japanese subjects; A multicenter, randomized, double-blind, placebo-controlled study.

BACKGROUND: Nobiletin exerts beneficial effects on cognitive function in various animal models of Alzheimer's disease. The present study aimed to investigate the benefits and safety of a combination food of nobiletin-rich extract from C. depressa peel for healthy elderly subjects. METHODS: The nobiletin-containing test food (Nobilex®) comprised high-purity nobiletin powder combined with dried root powder of K. parviflora and dried lead powder of P. japonicum and was administered to elderly Japanese subjects once a day for 16 weeks. The Japanese version of the Wechsler Memory Scaled-Revised (WMS-R) was used as a primary evaluation item for the assessment of global memory. Data from a protocol-matched population (Per Protocol Set: PPS) (n = 108) were analyzed. RESULTS: The scores of "general memory" or "visual memory" in the indices of WMS-R were significantly higher in the nobiletin-containing test food group than in the placebo group. The difference in the total WMS-R score was significantly higher in the test-food group (9.0 ± 7.20) than in the placebo group (5.9 ± 7.70). An age-stratified analysis of the WMS-R test showed similar changes in subjects aged ≦74 years to those in the overall subject population. In the stratified analysis involving subjects with an MMSE-J score of between 24 and 28, the "figural memory" subscale score in WMS-R was significantly higher in the test food group than in the placebo group. CONCLUSION: The present results indicate that the nobiletin-containing test food is beneficial for improving memory dysfunction in healthy elderly subjects.

Three Active Phytotoxic Compounds from the Leaves of Albizia richardiana (Voigt.) King and Prain for the Development of Bioherbicides to Control Weeds.

The global population is increasing day by day. To meet the food demand for such a huge number of people, crop production must increase without damaging the environment, and to prevent synthetic chemical herbicides from polluting the environment, controlling weeds using bioherbicides is essential. Accordingly, using phytotoxic substances obtained from plants for biological weed management has attracted attention. The plant Albizia richardiana possesses phytotoxic compounds that have been previously recorded. Hence, we have conducted this research to characterize more phytotoxic compounds in Albizia richardiana. Aqueous methanolic extracts of Albizia richardiana plant significantly restricted the growth of the examined plants lettuce and Italian ryegrass in a species- and concentration-dependent manner. Three active phytotoxic compounds were isolated through various chromatographic methods and identified as compound 1, 2, and 3. Compound 3 exhibited stronger phytotoxic potentials than the other two compounds and significantly suppressed the growth of Lepidium sativum (cress). The concentration of the compounds required for 50% growth reduction (I50 value) of the Lepidium sativum seedlings ranged between 0.0827 to 0.4133 mg/mL. The results suggest that these three phytotoxic compounds might contribute to the allelopathic potential of Albizia richardiana.

Prenylflavonoids from fruit of Macaranga tanarius promote glucose uptake via AMPK activation in L6 myotubes.

Skeletal muscle is a major tissue of glucose consumption and plays an important role in glucose homeostasis. Prenylflavonoids, a component of Macaranga tanarius fruits, have been reported to have antioxidant, antibacterial, and anticancer effects. However, the effects of these compounds on skeletal muscle glucose metabolism are unclear. Here, we isolated five prenylflavonoids from M. tanarius fruits, and investigated the mechanism of action of these compounds on skeletal muscle cells using L6 myotubes. We found that isonymphaeol B and 3'-geranyl naringenin increased glucose uptake in a dose-dependent manner. Furthermore, both isonymphaeol B and 3'-geranyl naringenin increased AMPK phosphorylation but did not affect PI3K-Akt phosphorylation. Isonymphaeol B and 3'-geranyl naringenin also increased Glut1 mRNA expression and plasma membrane GLUT1 protein levels. These results suggest that isonymphaeol B and 3'-geranyl naringenin have beneficial effects on glucose metabolism through AMPK and GLUT1 pathway. Isonymphaeol B and 3'-geranyl naringenin may be potential lead candidates for antidiabetic drug development.

Characterization of Macroscopic Colony-Forming Filamentous Cyanobacteria from Okinawan Coasts as Potential Sources of Bioactive Compounds.

Marine macroscopic colony-forming filamentous (MMCFF) cyanobacteria are considered as prolific producers of bioactive compounds. Thus, knowledge of the diversity of MMCFF cyanobacteria as related to bioactive compound production has become very important. However, basic taxonomic studies of MMCFF cyanobacteria are lacking. Many cyanobacterial taxa are still misidentified or undescribed. In this study, a total of 32 cyanobacterial colonies from nine coastal regions of Okinawa Prefecture were investigated for a diversity assessment. A polyphasic approach including morphological and molecular studies based on 16S rRNA gene sequences was performed to characterize Okinawan MMCFF cyanobacteria. Both morphological and molecular phylogenetic results showed that MMCFF cyanobacteria from Okinawan coasts are very diverse. We found morphotypes of Lyngbya-like, Phormidium-like, and Leptolyngbya-like groups among Okinawan cyanobacterial samples. Genetically, samples were distributed in various clades in the phylogenetic tree, including within Moorena, Okeania, Caldora, Neolyngbya, Dapis, as well as several unknown clades. In addition, cytotoxic activities of three samples from Kiyan coast were tested against HeLa cells. All three crude extracts of these samples showed strong cytotoxic activity with IC50 < 1 µg/ml.

Effect of pinocembrin isolated from Alpinia zerumbet on osteoblast differentiation.

Bone mass is regulated by osteoblast-mediated bone formation and osteoclast-mediated bone resorption. Osteoporosis is a bone metabolism disorder in which bone mass decreases due to increased bone resorption rather than bone formation. We focused on the traditional plant Alpinia zerumbet in Okinawa, Japan, and searched for promising compounds for the prevention and treatment of osteoporosis. Pinocembrin isolated from the leaves of A. zerumbet showed enhanced alkaline phosphatase (ALP) activity and mineralization and increased mRNA expression of osteoblast-related genes Alp and Osteocalcin (Ocn) in MC3T3-E1 cells. Pinocembrin increased the mRNA expression of Runx2 and Osterix, which are important transcription factors in osteoblast differentiation, and the mRNA expression of Dlx5 and Msx2, which are enhancers of these transcription factors. The bone morphogenetic protein (BMP) antagonist noggin, its receptor kinase inhibitor LDN-193189 and p38 MAPK inhibitor SB203580 attenuated pinocembrin-promoted ALP activity. Pinocembrin increased the mRNA of Bmp-2 and its target gene Id1. In addition, the estrogen receptor (ER) inhibitor ICI182780 suppressed pinocembrin-stimulated ALP activity. Pinocembrin may increase BMP-2 expression via ER. Then, the BMP-2 promotes osteoblast specific genes expression and mineralization through both Smad-dependent and independent pathway following Runx2 and Osterix induction. Our findings suggest that pinocembrin has bone anabolic effects and may be useful for the prevention and treatment of bone metabolic diseases such as osteoporosis.