Polyketide Derivatives from the Mangrove-Derived Fungus Penicillium sp. HDN15-312.

Four new polyketides, namely furantides A-B (1-2), talamin E (3) and arugosinacid A (4), and two known polyketides were obtained from the mangrove-derived fungus Penicillium sp. HDN15-312 using the One Strain Many Compounds (OSMAC) strategy. Their chemical structures, including configurations, were elucidated by detailed analysis of extensive NMR spectra, HRESIMS and ECD. The DPPH radicals scavenging activity of 3, with an IC50 value of 6.79 µM, was better than vitamin C.

The Impact of the Culture Regime on the Metabolome and Anti-Phytopathogenic Activity of Marine Fungal Co-Cultures.

Co-cultivation, coupled with the OSMAC approach, is considered an efficient method for expanding microbial chemical diversity through the activation of cryptic biosynthetic gene clusters (BGCs). As part of our project aiming to discover new fungal metabolites for crop protection, we previously reported five polyketides, the macrolides dendrodolides E (1) and N (2), the azaphilones spiciferinone (3) and 8α-hydroxy-spiciferinone (4), and the bis-naphtho-γ-pyrone cephalochromin (5) from the solid Potato Dextrose Agar (PDA) co-culture of two marine sediment-derived fungi, Plenodomus influorescens and Pyrenochaeta nobilis. However, some of the purified metabolites could not be tested due to their minute quantities. Here we cultivated these fungi (both axenic and co-cultures) in liquid regime using three different media, Potato Dextrose Broth (PDB), Sabouraud Dextrose Broth (SDB), and Czapek-Dox Broth (CDB), with or without shaking. The aim was to determine the most ideal co-cultivation conditions to enhance the titers of the previously isolated compounds and to produce extracts with stronger anti-phytopathogenic activity as a basis for future upscaled fermentation. Comparative metabolomics by UPLC-MS/MS-based molecular networking and manual dereplication was employed for chemical profiling and compound annotations. Liquid co-cultivation in PDB under shaking led to the strongest activity against the phytopathogen Phytophthora infestans. Except for compound 1, all target compounds were detected in the co-culture in PDB. Compounds 2 and 5 were produced in lower titers, whereas the azaphilones (3 and 4) were overexpressed in PDB compared to PDA. Notably, liquid PDB co-cultures contained meroterpenoids and depside clusters that were absent in the solid PDA co-cultures. This study demonstrates the importance of culture regime in BGC regulation and chemical diversity of fungal strains in co-culture studies.

Mycobolome of Phialomyces macrosporus across OSMAC-based assorted culture media.

The fungus Phialomyces macrosporus was cultured using the One Strain Many Compounds (OSMAC) strategies to evaluate its metabolome. Variations in the nutrient culture media, culture regime, and cultivation parameters can significantly influence fungal extract quantity and chemical diversity. This study aimed to explore the mycobolome of P. macrosporus in five different culture media and two different cultivation conditions using NMR-based metabolomics. Principal component analysis (PCA) of 1H NMR spectra revealed clear differentiation between these samples, highlighting the rice dextrose agar medium (RDA) and potato dextrose broth (PDB) as standard complex media for conducting a fungal metabolite screening program.

Exploring Culture Media Diversity to Produce Fungal Secondary Metabolites and Cyborg Cells.

Fungi are microorganisms of significant biotechnological importance due to their ability to provide food and produce several value-added secondary metabolites and enzymes. Its products move billions of dollars in the pharmaceutical, cosmetics, and additives sectors. These microorganisms also play a notable role in bionanotechnology, leading to the production of hybrid biological-inorganic materials (such as cyborg cells) and the use of their enzyme complex in the biosynthesis of nanoparticles. In this sense, optimizing the fungal growth process is necessary, with selecting the cultivation medium as one of the essential factors for the microorganism to reach its maximum metabolic expression. The culture medium's composition can also impact the nanomaterial's stability and prevent the incorporation of nanoparticles into fungal cells. Therefore, our main objectives are the following: (1) compile and discuss the most commonly employed culture media for the production of fungal secondary metabolites and the formation of cyborg cells, accompanied by preparation methods; (2) provide a six-step guide to investigating the fungal metabolomic profile and (3) discuss the main procedures of microbial cultivation to produce fungal cyborg cells.

A new secondary metabolite from the marine-derived fungus Phomopsis lithocarpus FS508.

Based on the One Strain-Many Compounds (OSMAC) strategy, the secondary metabolites of Phomopsis lithocarpus FS508 were investigated. As a result, a new secondary metabolite, 4-methoxy-3-[4-(acetyloxy)-3-methyl-2-butenyl]benzoic acid (1) as well as eleven known compounds were isolated from the fermentation product of the strain FS508. Their structures were determined by NMR, IR, UV, and MS spectroscopic data analyses. All the isolated compounds were evaluated for cytotoxic and anti-inflammatory activities. Among them, compounds 3 and 9 displayed potent cytotoxicity against HepG-2 cell line, and compounds 2, 3 and 12 showed significant anti-inflammatory activities.

A Close View of the Production of Bioactive Fungal Metabolites Mediated by Chromatin Modifiers.

Secondary metabolites produced by fungi are well known for their biological properties, which play important roles in medicine. These metabolites aid in managing infections and treating chronic illnesses, thereby contributing substantially to human health improvement. Despite this extensive knowledge, the vast biodiversity and biosynthetic potential of fungi is still largely unexplored, highlighting the need for further research in natural products. In this review, several secondary metabolites of fungal origin are described, emphasizing novel structures and skeletons. The detection and characterization of these metabolites have been significantly facilitated by advancements in analytical systems, particularly modern hyphenated liquid chromatography/mass spectrometry. These improvements have primarily enhanced sensitivity, resolution, and analysis flow velocity. Since the in vitro production of novel metabolites is often lower than the re-isolation of known metabolites, understanding chromatin-based alterations in fungal gene expression can elucidate potential pathways for discovering new metabolites. Several protocols for inducing metabolite production from different strains are discussed, demonstrating the need for uniformity in experimental procedures to achieve consistent biosynthetic activation.

OSMAC strategy integrated with molecular networking discovery peniciacetals A-I, nine new meroterpenoids from the mangrove-derived fungus Penicillium sp. HLLG-122.

Nine new highly oxygenated meroterpenoids, peniciacetals A-I (1-9), along with five known analogues (10-14) were isolated from the mangrove-derived fungus Penicillium sp. HLLG-122 based on the guidance of molecular networking and OSMAC approach. Peniciacetals A-B (1-2) were characterized with a unique 6/6/6/6/5 pentacyclic system possessing an unusual 4,6-dimethyl-2,5-dioxohexahydro-6-carboxy-4H-furo[2,3-b]pyran moiety. Peniciacetals C-D (3-4) possessed an uncommon 3,6-dimethyldihydro-4H-furo[2,3-b]pyran-2,5-dione unit with 6/6/6/5/6 fused pentacyclic skeleton. The structures and absolute configurations of new compounds were elucidated by HR-ESI-MS, 1D and 2D NMR spectroscopic data, X-ray diffraction analysis, and quantum chemical electronic circular dichroism (ECD) calculation. The plausible biosynthetic pathway of 1-9 were also proposed. Compound 14 showed good cytotoxicity against HepG2, MCF-7, HL-60, A549, HCT116 and H929 cell with IC50 values of 6.6, 14.8, 3.2, 5.7, 6.9 and 3.0 µM, respectively. Further research showed that the compound 14 induced necrosis or late apoptosis contributes to the HL-60 cells toxicity.

Induction of New Aromatic Polyketides from the Marine Actinobacterium Streptomyces griseorubiginosus through an OSMAC Approach.

Using the OSMAC (One Strain Many Compounds) approach, the actinobacterium Streptomyces griseorubiginosus, derived from an unidentified cnidarian collected from a reef near Pointe de Bellevue in Réunion Island (France), was subjected to cultivation under diverse conditions. This endeavour yielded the isolation of a repertoire of 23 secondary metabolites (1-23), wherein five compounds were unprecedented as natural products (19-23). Specifically, compounds 19 and 20 showcased novel anthrone backbones, while compound 23 displayed a distinctive tetralone structure. Additionally, compounds 21 and 22 presented an unusual naphtho [2,3-c]furan-4(9H)-one chromophore. Interestingly, the detection of all these novel compounds (19-23) was exclusively achieved when the bacterium was cultured in FA-1 liquid medium supplemented with the epigenetic modifier γ-butyrolactone. The elucidation of the structural features of the newfound compounds was accomplished through a combination of HRESIMS, 1D and 2D NMR spectroscopy, as well as QM-NMR (Quantum Mechanical-Nuclear Magnetic Resonance) methods and by comparison with existing literature. Moreover, the determination of the relative configuration of compound 23 was facilitated by employing the mix-J-DP4 computational approach.

Enhanced Molecular Networking Shows Microbacterium sp. V1 as a Factory of Antioxidant Proline-Rich Peptides.

Two linear proline-rich peptides (1-2), bearing an N-terminal pyroglutamate, were isolated from the marine bacterium Microbacterium sp. V1, associated with the marine sponge Petrosia ficiformis, collected in the volcanic CO2 vents in Ischia Island (South Italy). Peptide production was triggered at low temperature following the one strain many compounds (OSMAC) method. Both peptides were detected together with other peptides (3-8) via an integrated, untargeted MS/MS-based molecular networking and cheminformatic approach. The planar structure of the peptides was determined by extensive 1D and 2D NMR and HR-MS analysis, and the stereochemistry of the aminoacyl residues was inferred by Marfey's analysis. Peptides 1-8 are likely to arise from Microbacterium V1 tailor-made proteolysis of tryptone. Peptides 1 and 2 were shown to display antioxidant properties in the ferric-reducing antioxidant power (FRAP) assay.

OSMAC-Based Discovery and Biosynthetic Gene Clusters Analysis of Secondary Metabolites from Marine-Derived Streptomyces globisporus SCSIO LCY30.

The one strain many compounds (OSMAC) strategy is an effective method for activating silent gene clusters by cultivating microorganisms under various conditions. The whole genome sequence of the marine-derived strain Streptomyces globisporus SCSIO LCY30 revealed that it contains 30 biosynthetic gene clusters (BGCs). By using the OSMAC strategy, three types of secondary metabolites were activated and identified, including three angucyclines, mayamycin A (1), mayamycin B (2), and rabolemycin (3); two streptophenazines (streptophenazin O (4) and M (5)); and a macrolide dimeric dinactin (6), respectively. The biosynthetic pathways of the secondary metabolites in these three families were proposed based on the gene function prediction and structural information. The bioactivity assays showed that angucycline compounds 1-3 exhibited potent antitumor activities against 11 human cancer cell lines and antibacterial activities against a series of Gram-positive bacteria. Mayamycin (1) selectively exhibited potent cytotoxicity activity against triple-negative breast cancer (TNBC) cell lines such as MDA-MB-231, MDA-MB-468, and Bt-549, with IC50 values of 0.60-2.22 µM.

New Eremophilane-Type Sesquiterpenes from the Marine Sediment-Derived Fungus Emericellopsis maritima BC17 and Their Cytotoxic and Antimicrobial Activities.

The fungal strain BC17 was isolated from sediments collected in the intertidal zone of the inner Bay of Cadiz and characterized as Emericellopsis maritima. On the basis of the one strain-many compounds (OSMAC) approach, four new eremophilane-type sesquiterpenes (1-4), together with thirteen known derivatives (5-17) and two reported diketopiperazines (18, 19), were isolated from this strain. The chemical structures and absolute configurations of the new compounds were determined through extensive NMR and HRESIMS spectroscopic studies and ECD calculation. Thirteen of the isolated eremophilanes were examined for cytotoxic and antimicrobial activities. PR toxin (16) exhibited cytotoxic activity against HepG2, MCF-7, A549, A2058, and Mia PaCa-2 human cancer cell lines with IC50 values ranging from 3.75 to 33.44 µM. (+)-Aristolochene (10) exhibited selective activity against the fungal strains Aspergillus fumigatus ATCC46645 and Candida albicans ATCC64124 at 471 µM.

OSMAC Method to Assess Impact of Culture Parameters on Metabolomic Diversity and Biological Activity of Marine-Derived Actinobacteria.

Actinobacteria are known for their production of bioactive specialized metabolites, but they are still under-exploited. This study uses the "One Strain Many Compounds" (OSMAC) method to explore the potential of three preselected marine-derived actinobacteria: Salinispora arenicola (SH-78) and two Micromonospora sp. strains (SH-82 and SH-57). Various parameters, including the duration of the culture and the nature of the growth medium, were modified to assess their impact on the production of specialized metabolites. This approach involved a characterization based on chemical analysis completed with the construction of molecular networks and biological testing to evaluate cytotoxic and antiplasmodial activities. The results indicated that the influence of culture parameters depended on the studied species and also varied in relation with the microbial metabolites targeted. However, common favorable parameters could be observed for all strains such as an increase in the duration of the culture or the use of the A1 medium. For Micromonospora sp. SH-82, the solid A1 medium culture over 21 days favored a greater chemical diversity. A rise in the antiplasmodial activity was observed with this culture duration, with a IC50 twice as low as for the 14-day culture. Micromonospora sp. SH-57 produced more diverse natural products in liquid culture, with approximately 54% of nodes from the molecular network specifically linked to the type of culture support. Enhanced biological activities were also observed with specific sets of parameters. Finally, for Salinispora arenicola SH-78, liquid culture allowed a greater diversity of metabolites, but intensity variations were specifically observed for some metabolites under other conditions. Notably, compounds related to staurosporine were more abundant in solid culture. Consequently, in the range of the chosen parameters, optimal conditions to enhance metabolic diversity and biological activities in these three marine-derived actinobacteria were identified, paving the way for future isolation works.

Rapid Discovery of Substances with Anticancer Potential from Marine Fungi Based on a One Strain-Many Compounds Strategy and UPLC-QTOF-MS.

The secondary metabolites of marine fungi with rich chemical diversity and biological activity are an important and exciting target for natural product research. This study aimed to investigate the fungal community in Quanzhou Bay, Fujian, and identified 28 strains of marine fungi. A total of 28 strains of marine fungi were screened for small-scale fermentation by the OSMAC (One Strain-Many Compounds) strategy, and 77 EtOAc crude extracts were obtained and assayed for cancer cell inhibition rate. A total of six strains of marine fungi (P-WZ-2, P-WZ-3-2, P-WZ-4, P-WZ-5, P56, and P341) with significant changes in cancer cell inhibition induced by the OSMAC strategy were analysed by UPLC-QTOF-MS. The ACD/MS Structure ID Suite software was used to predict the possible structures with inhibitory effects on cancer cells. A total of 23 compounds were identified, of which 10 compounds have been reported to have potential anticancer activity or cytotoxicity. In this study, the OSMAC strategy was combined with an untargeted metabolomics approach based on UPLC-QTOF-MS to efficiently analyse the effect of changes in culture conditions on anticancer potentials and to rapidly find active substances that inhibit cancer cell growth.

Bioactivity and Metabolome Mining of Deep-Sea Sediment-Derived Microorganisms Reveal New Hybrid PKS-NRPS Macrolactone from Aspergillus versicolor PS108-62.

Despite low temperatures, poor nutrient levels and high pressure, microorganisms thrive in deep-sea environments of polar regions. The adaptability to such extreme environments renders deep-sea microorganisms an encouraging source of novel, bioactive secondary metabolites. In this study, we isolated 77 microorganisms collected by a remotely operated vehicle from the seafloor in the Fram Strait, Arctic Ocean (depth of 2454 m). Thirty-two bacteria and six fungal strains that represented the phylogenetic diversity of the isolates were cultured using an One-Strain-Many-Compounds (OSMAC) approach. The crude EtOAc extracts were tested for antimicrobial and anticancer activities. While antibacterial activity against methicillin-resistant Staphylococcus aureus (MRSA) and Enterococcus faecium was common for many isolates, only two bacteria displayed anticancer activity, and two fungi inhibited the pathogenic yeast Candida albicans. Due to bioactivity against C. albicans and rich chemical diversity based on molecular network-based untargeted metabolomics, Aspergillus versicolor PS108-62 was selected for an in-depth chemical investigation. A chemical work-up of the SPE-fractions of its dichloromethane subextract led to the isolation of a new PKS-NRPS hybrid macrolactone, versicolide A (1), a new quinazoline (-)-isoversicomide A (3), as well as three known compounds, burnettramic acid A (2), cyclopenol (4) and cyclopenin (5). Their structures were elucidated by a combination of HRMS, NMR, [α]D, FT-IR spectroscopy and computational approaches. Due to the low amounts obtained, only compounds 2 and 4 could be tested for bioactivity, with 2 inhibiting the growth of C. albicans (IC50 7.2 µg/mL). These findings highlight, on the one hand, the vast potential of the genus Aspergillus to produce novel chemistry, particularly from underexplored ecological niches such as the Arctic deep sea, and on the other, the importance of untargeted metabolomics for selection of marine extracts for downstream chemical investigations.

Exploration of the Production of Three Thiodiketopiperazines by an Endophytic Fungal Strain of Cophinforma mamane.

Endophytic fungi possess a versatile metabolism which is related to their ability to live in diverse ecological niches. While culturing under laboratory conditions, their metabolism is mainly influenced by the culture media, time of incubation and other physicochemical factors. In this study, we focused on the production of 3 thiodiketopiperazines (TDKPs) botryosulfuranols A-C produced by an endophytic strain of Cophinforma mamane isolated from the leaves of Bixa orellana L collected in the Peruvian Amazon. We studied the time-course production of botryosulfuranols A-C during 28 days and evaluated the variations in the production of secondary metabolites, including the TDKPs, produced by C. mamane in response to different culture media, light versus dark conditions and different incubation times. We observed a short time-frame production of botryosulfuranol C while its production was significantly affected by the light conditions and nutrients of the culture media. Botryosulfuranols A and B showed a similar production pattern and a similar response to culturing conditions. Molecular networking allowed us to detect three compounds related to TDKPs that will be the focus of future experiments.

Cryptic Metabolites from Marine-Derived Microorganisms Using OSMAC and Epigenetic Approaches.

Marine microorganisms have proven to be a source of new natural products with a wide spectrum of biological activities relevant in different industrial sectors. The ever-increasing number of sequenced microbial genomes has highlighted a discrepancy between the number of gene clusters potentially encoding the production of natural products and the actual number of chemically characterized metabolites for a given microorganism. Homologous and heterologous expression of these biosynthetic genes, which are often silent under experimental laboratory culture conditions, may lead to the discovery of new cryptic natural products of medical and biotechnological interest. Several new genetic and cultivation-based strategies have been developed to meet this challenge. The OSMAC approach (one strain-many compounds), based on modification of growth conditions, has proven to be a powerful strategy for the discovery of new cryptic natural products. As a direct extension of this approach, the addition of chemical elicitors or epigenetic modifiers have also been used to activate silent genes. This review looks at the structures and biological activities of new cryptic metabolites from marine-derived microorganisms obtained using the OSMAC approach, the addition of chemical elicitors, and enzymatic inhibitors and epigenetic modifiers. It covers works published up to June 2021.

New Dibenzo-α-pyrone Derivatives with α-Glucosidase Inhibitory Activities from the Marine-Derived Fungus Alternaria alternata.

Three new dibenzo-α-pyrone derivatives, alternolides A-C (1-3), and seven known congeners (4-10) were isolated from the marine-derived fungus of Alternaria alternata LW37 assisted by the one strain-many compounds (OSMAC) strategy. The structures of 1-3 were established by extensive spectroscopic analyses, and their absolute configurations were determined by modified Snatzke's method and electronic circular dichroism (ECD) calculations. Compounds 6 and 7 showed good 1,1-diphenyl-2-picrylhydrazyl (DPPH) antioxidant scavenging activities with IC50 values of 83.94 ± 4.14 and 23.60 ± 1.23 µM, respectively. Additionally, 2, 3 and 7 exhibited inhibitory effects against α-glucosidase with IC50 values of 725.85 ± 4.75, 451.25 ± 6.95 and 6.27 ± 0.68 µM, respectively. The enzyme kinetics study indicated 2 and 3 were mixed-type inhibitors of α-glucosidase with Ki values of 347.0 and 108.5 µM, respectively. Furthermore, the interactions of 2, 3 and 7 with α-glucosidase were investigated by molecular docking.

Polyketides from the marine-derived fungus Aspergillus falconensis: In silico and in vitro cytotoxicity studies.

Fermentation of the marine-derived fungus Aspergillus falconensis, isolated from sediment collected from the Red Sea, Egypt on solid rice medium containing 3.5% NaCl yielded a new dibenzoxepin derivative (1) and a new natural isocoumarin (2) along with six known compounds (3-8). Changes in the metabolic profile of the fungus were induced by replacing NaCl with 3.5% (NH4)2SO4 that resulted in the accumulation of three further known compounds (9-11), which were not detected when the fungus was cultivated in the presence of NaCl. The structures of the new compounds were elucidated by HRESIMS and 1D/2D NMR as well as by comparison with the literature. Molecular docking was conducted for all isolated compounds on crucial enzymes involved in the formation, progression and metastasis of cancer which included human cyclin-dependent kinase 2 (CDK-2), human DNA topoisomerase II (TOP-2) and matrix metalloproteinase 13 (MMP-13). Diorcinol (7), sulochrin (9) and monochlorosulochrin (10) displayed notable stability within the active pocket of CDK-2 with free binding energy (ΔG) equals to -25.72, -25.03 and -25.37 Kcal/mol, respectively whereas sulochrin (9) exerted the highest fitting score within MMP-13 active center (ΔG = -33.83 Kcal/mol). In vitro cytotoxic assessment using MTT assay showed that sulochrin (9) exhibited cytotoxic activity versus L5178Y mouse lymphoma cells with an IC50 value of 5.1 µM and inhibition of migration of MDA-MB 231 breast cancer cells at a concentration of 70 µM.

Soybean-associated endophytic fungi as potential source for anti-COVID-19 metabolites supported by docking analysis.

AIMS: To identify the metabolites produced by the endophytic fungus, Aspergillus terreus and to explore the anti-viral activity of the identified metabolites against the pandemic disease COVID-19 in-silico. METHODS AND RESULTS: Herein, we reported the isolation of A. terreus, the endophytic fungus associated with soybean roots, which is then subcultured using OSMAC approach in five different culture media. Analytical analysis of media ethylacetate extracts using liquid chromatography coupled with high-resolution mass spectrometry (LC-HRMS) was carried out. Furthermore, the obtained LC-MS data were statistically processed with MetaboAnalyst 4.0. Molecular docking studies were performed for the dereplicated metabolites against COVID-19 main protease (Mpro ). Metabolomic profiling revealed the presence of 18 compounds belonging to different chemical classes. Quinones, polyketides and isocoumarins were the most abundant classes. Multivariate analysis revealed that potato dextrose broth and modified potato dextrose broth are the optimal media for metabolites production. Molecular docking studies declared that the metabolites, Aspergillide B1 and 3a-Hydroxy-3, 5-dihydromonacolin L showed the highest binding energy scores towards COVID-19 main protease (Mpro ) (-9·473) and (-9·386), respectively, and they interact strongly with the catalytic dyad (His41 and Cys145) amino acid residues of Mpro . CONCLUSIONS: A combination of metabolomics and in-silico approaches have allowed a shorter route to search for anti-COVID-19 natural products in a shorter time. The dereplicated metabolites, aspergillide B1 and 3α-Hydroxy-3, 5-dihydromonacolin L were found to be potent anti-COVID-19 drug candidates in the molecular docking study. SIGNIFICANCE AND IMPACT OF THE STUDY: This study revealed that the endophytic fungus, A. terreus can be considered as a potential source of natural bioactive products. In addition to, the possibility of developing the metabolites, aspergillide B1 and 3α-Hydroxy-3, 5-dihydromonacolin L to be used as phytopharmaceuticals for the management of COVID-19.

Discovery of andrastones from the deep-sea-derived Penicillium allii-sativi MCCC 3A00580 by OSMAC strategy.

Andrastones are unusual 6,6,6,5-tetracyclic meroterpenoids that are rarely found in nature. Previously, three andrastones were obtained from the rice static fermentation extract of the deep-sea-derived fungus Penicillium allii-sativi MCCC 3A00580. Inspired by one strain many compounds (OSMAC) approach, the oat static fermentation on P. allii-sativi was conducted. As a result, 14 andrastones were isolated by UV-guided isolation. The chemical structures of the nine new compounds (1-9) was established by comprehensive analysis of the NMR, MS, ECD, and X-ray crystallography and the five known ones (10-14) were assigned by comparing their NMR, MS, and OR data with those reported in literature. Compound 1 bears a novel hemiketal moiety while 2 is the first example to possess a novel tetrahydrofuran moiety via C-7 and C-15. All isolates were tested for anti-allergic bioactivity. Compound 10, 3-deacetylcitreohybridonol, significantly decreased degranulation with the IC50 value of 14.8 µM, compared to that of 92.5 µM for the positive control, loratadine. Mechanism study indicated 10 could decrease the generation of histamine and TNF-α by reducing the accumulation of Ca2+ in RBL-2H3 cells. These findings indicate andrastones could be potential to discover new anti-allergic candidate drugs.