Characterization of Crude and Processed Pulp Cell Walls of Three Selected Mamey Accessions (Mammea americana L.).

Mamey (Mammea americana L.) is a tropical fleshy fruit native from the West Indies and northern South America. It is very appreciated for its flavor and color but has been little described. The present study investigates the composition and histochemistry of the pulp cell walls of three mamey accessions readily available in Martinique. The impact of pulp processing into puree on cell wall composition is evaluated. The histology and rheology of mamey puree are assessed considering these characterizations. Mamey pulp cell wall composition is dominated by highly methyl-esterified pectins (DM: 66.2-76.7%) of high molecular weight, and show few hemicelluloses, mainly xyloglucans. Processing reduced methyl-esterified uronic acid contents and gave purees with significantly different viscosities. Mamey puree was composed of polydisperse particles (20-2343 µm), which size distributions were different depending on the accession: Ti Jacques was dominated by smaller particles (50% had approximated diameters lower than 160 µm), Sonson's by larger particles (50% had approximated diameters higher than 900 µm), and Galion's had an intermediate profile. This new knowledge on mamey pulp is valuable for future works on mamey processing into new food products, even more so for those including cell wall polysaccharide-degrading enzymes.

In vivo antimalarial effect of 1-hydroxy-5,6,7-trimethoxyxanthone isolated from Mammea siamensis T. Anders. flowers: pharmacokinetic and acute toxicity studies.

BACKGROUND: The potent antiplasmodial activity of 1-hydroxy-5,6,7-trimethoxyxanthone (HTX), isolated from Mammea siamensis T. Anders. flowers, has previously been demonstrated in vitro. However, its in vivo activity has not been reported. Therefore, this study aimed to investigate the antimalarial activity and acute toxicity of HTX in a mouse model and to evaluate the pharmacokinetic profile of HTX following a single intraperitoneal administration. METHODS: The in vivo antimalarial activity of HTX was evaluated using a 4-day suppressive test. Mice were intraperitoneally injected with Plasmodium berghei ANKA strain and given HTX daily for 4 days. To detect acute toxicity, mice received a single dose of HTX and were observed for 14 days. Additionally, the biochemical parameters of the liver and kidney functions as well as the histopathology of liver and kidney tissues were examined. HTX pharmacokinetics after intraperitoneal administration was also investigated in a mouse model. Liquid chromatography triple quadrupole mass spectrometry was used to quantify plasma HTX and calculate pharmacokinetic parameters with the PKSolver software. RESULTS: HTX at 10 mg/kg body weight significantly suppressed parasitemia in malaria-infected mice by 74.26%. Mice treated with 3 mg/kg HTX showed 46.88% suppression, whereas mice treated with 1 mg/kg displayed 34.56% suppression. Additionally, no symptoms of acute toxicity were observed in the HTX-treated groups. There were no significant alterations in the biochemical parameters of the liver and kidney functions and no histological changes in liver or kidney tissues. Following intraperitoneal HTX administration, the pharmacokinetic profile exhibited a maximum concentration (Cmax) of 94.02 ng/mL, time to attain Cmax (Tmax) of 0.5 h, mean resident time of 14.80 h, and elimination half-life of 13.88 h. CONCLUSIONS: HTX has in vivo antimalarial properties against P. berghei infection. Acute toxicity studies of HTX did not show behavioral changes or mortality. The median lethal dose was greater than 50 mg/kg body weight. Pharmacokinetic studies showed that HTX has a long elimination half-life; hence, shortening the duration of malaria treatment may be required to minimize toxicity.

Antibacterial Effect of Ethanolic Extracts of Dodonaea viscosa L. Jacq. and Mammea americana L. against Staphylococci Isolated from Skin Lesions.

Background: The resistance to antibiotics shown by some dermatological pathogenic microorganisms has increased the interest of pharmaceutical and cosmetic industries in developing natural products that possess different biological activities, including antimicrobial effects. Methods: In the present investigation, the antibacterial activity of ethanolic extracts of Dodonaea viscosa aerial part and Mammea americana leaves and seed was evaluated against resistant strains of Staphylococcus isolated from skin lesions and against S. aureus ATCC 25923 (reference strain). Column chromatography (CC) and preparative thin-layer chromatography (PTLC) were used to obtain separate fractions of the seed extract of M. americana. We also determined the antimicrobial resistance of the strains against antibiotics using the agar disc diffusion assay. In addition, phytochemical screening was performed by colorimetric standard techniques. Results: M. americana seed extract showed the highest antibacterial activity with MBC from 2.3 µg/mL to 19.5 µg/mL without differences with gentamicin (p = 0.998). The isolated strain S. epidermidis I showed the highest antimicrobial resistance against the tested antibiotics. PTLC-fractions of M. americana seed extract showed MBC from 3.2 µg/mL to 40.7 µg/mL against S. epidermidis I and S. aureus 25923 (reference), respectively, which suggests a synergistic effect of the secondary metabolites present in the crude ethanolic extract compared to its active PTLC-fractions, where only coumarins and compounds with lactone groups were detected in the phytochemical screening. Conclusion: M. americana seed extract has promising effects that should be considered in further studies as an alternative or adjuvant in treating skin infections caused by staphylococci.

Natural Compounds with BMI1 Promoter Inhibitory Activity from Mammea siamensis and Andrographis paniculata.

A new coumarin derivative (1) and 30 known compounds were isolated from Mammea siamensis and Andrographis paniculata, guided by B cell-specific Moloney murine leukemia virus insertion region 1 (BMI1) promoter inhibitory activity. Among the isolated compounds, 15 compounds showed BMI1 promoter inhibitory activity, and five compounds were found to be cytotoxic. 14-Deoxy-11,12-dehydroandrographolide (18) was highly cytotoxic to DU145 cells with an IC50 value of 25.4 µM. Western blotting analysis of compound 18 in DU145 cells suggested that compound 18 suppresses BMI1 expression.

Phytochemicals with Chemopreventive Activity Obtained from the Thai Medicinal Plant Mammea siamensis (Miq.) T. Anders.: Isolation and Structure Determination of New Prenylcoumarins with Inhibitory Activity against Aromatase.

With the aim of searching for phytochemicals with aromatase inhibitory activity, five new prenylcoumarins, mammeasins K (1), L (2), M (3), N (4), and O (5), were isolated from the methanolic extract of Mammea siamensis (Miq.) T. Anders. flowers (fam. Calophyllaceae), originating in Thailand. The stereostructures of 1-5 were elucidated based on their spectroscopic properties. Among the new compounds, 1 (IC50 = 7.6 µM) and 5 (9.1 µM) possessed relatively strong inhibitory activity against aromatase, which is a target of drugs already used in clinical practice for the treatment and prevention of estrogen-dependent breast cancer. The analysis through Lineweaver-Burk plots showed that they competitively inhibit aromatase (1, Ki = 3.4 µM and 5, 2.3 µM). Additionally, the most potent coumarin constituent, mammea B/AB cyclo D (31, Ki = 0.84 µM), had a competitive inhibitory activity equivalent to that of aminoglutethimide (0.84 µM), an aromatase inhibitor used in therapeutics.

Antimalarial potential of compounds isolated from Mammea siamensis T. Anders. flowers: in vitro and molecular docking studies.

BACKGROUND: The emergence of antimalarial drug resistance encourages the search for new antimalarial agents. Mammea siamensis belongs to the Calophyllaceae family, which is a medicinal plant that is used in traditional Thai preparations. The hexane and dichloromethane extracts of this plant were found to have potent antimalarial activity. Therefore, this study aimed to isolate active compounds from M. siamensis flowers and evaluate their antimalarial potential and their interactions with Plasmodium falciparum lactate dehydrogenase (PfLDH). METHODS: The compounds from M. siamensis flowers were isolated by chromatographic techniques and evaluated for their antimalarial activity against chloroquine (CQ)-resistant P. falciparum (K1) strains using a parasite lactate dehydrogenase (pLDH) assay. Interactions between the isolated compounds and the PfLDH enzyme were investigated using a molecular docking method. RESULTS: The isolation produced the following thirteen compounds: two terpenoids, lupeol (1) and a mixture of ß-sitosterol and stigmasterol (5); two mammea coumarins, mammea A/AA cyclo D (6) and mammea A/AA cyclo F (7); and nine xanthones, 4,5-dihydroxy-3-methoxyxanthone (2), 4-hydroxyxanthone (3), 1,7-dihydroxyxanthone (4), 1,6-dihydroxyxanthone (8), 1-hydroxy-5,6,7-trimethoxyxanthone (9), 3,4,5-trihydroxyxanthone (10), 5-hydroxy-1-methoxyxanthone (11), 2-hydroxyxanthone (12), and 1,5-dihydroxy-6-methoxyxanthone (13). Compound 9 exhibited the most potent antimalarial activity with an IC50 value of 9.57 µM, followed by 10, 1, 2 and 13 with IC50 values of 15.48, 18.78, 20.96 and 22.27 µM, respectively. The molecular docking results indicated that 9, which exhibited the most potent activity, also had the best binding affinity to the PfLDH enzyme in terms of its low binding energy (-7.35 kcal/mol) and formed interactions with ARG109, ASN140, and ARG171. CONCLUSION: These findings revealed that isolated compounds from M. siamensis flowers exhibited antimalarial activity. The result suggests that 1-hydroxy-5,6,7-trimethoxyxanthone is a possible lead structure as a potent inhibitor of the PfLDH enzyme.

Anti-proliferative activities of coumarins from the Thai medicinal plant Mammea siamensis (Miq.) T. Anders. against human digestive tract carcinoma cell lines.

Geranylated coumarins named mammeasins G-J (1-4) were isolated from the methanol extract of the flowers of Mammea siamensis (Miq.) T. Anders. (Calophyllaceae) originating in Thailand. Their structures were established based on detailed spectroscopic analyses. The isolates, including previously reported coumarin constituents (5-28), exhibited anti-proliferative activities against human carcinoma cell lines HSC-2, HSC-4, MKN-45, and MCF-7. Mammeasin A (7, IC50 = 13.6 µM) and surangin B (15, 15.2 µM), both consisting of the geranyl group, were found to show relatively strong activities against HSC-4 cells and their mechanisms of action were found to involve apoptotic cell death.

Mammea type coumarins isolated from Calophyllum brasiliense induced apoptotic cell death of Trypanosoma cruzi through mitochondrial dysfunction, ROS production and cell cycle alterations.

Chagas Disease is a neglected tropical disease caused by the protozoan parasite Trypanosoma cruzi which affects 6-8 million people, mostly in Latin America. The medical treatment is based on two nitroimidazole compounds, which have limited effectiveness in the chronic phase of the disease and produce several adverse effects; consequently, there is an urgent need to develop new, safe, and effective drugs. Previous reports had shown that natural coumarins, especially mammea A/BA isolated from the tropical tree Calophyllum brasiliense, is a promissory molecule for developing new drugs, due to its potent activity, higher than benznidazole, selectivity, and its low toxicity in mice. However, its mode of action is still unknown. In the present work, we evaluated the mechanism of action of the coumarin mammea A/BA (93.6%), isolated from the tropical tree C. brasiliense on Querétaro strain (Tc1) of T. cruzi. This compound was tested in vitro on epimastigotes and trypomastigotes of T. cruzi for intracellular esterase activity, plasma membrane integrity, phosphatidylserine exposure, ROS production, mitochondrial membrane potential, caspase-like activity, DNA integrity, cell cycle and autophagy. Mammea A/BA showed a 50% lethal concentration (LC50) of 85.8 and 36.9 µM for epimastigotes and trypomastigotes respectively. It affected intracellular esterase activity, produced important plasma membrane damage and induced phosphatidylserine exposure. An increase in reactive oxygen species (ROS) and decrease in mitochondrial membrane potential were detected. Caspase-like activity was present in both parasite forms producing DNA integrity damage. This compound also induced a cell cycle arrest in the G1 phase and the presence of autophagy vacuoles. The above data suggest that mammea A/BA induce cell death of T. cruzi by autophagy and apoptosis-like phenomena and support our suggestion that mammea A/BA could be a promising molecule for the development of new drugs to treat Chagas Disease.

Antimicrobial activity of endophytic fungi from the medicinal plants Mammea americana (Calophyllaceae) and Moringa oleifera (Moringaceae) / [Actividad antimicrobiana de hongos endófitos de las plantas medicinales Mammea americana (Calophyllaceae) y Moringa oleifera (Moringaceae)].

Introduction: Infectious diseases represent one of the leading causes of death worldwide. Considering the growing global challenge of antimicrobial resistance, research into new sources of potentially effective antimicrobial agents from natural origins is of great importance for world health. Objective: To evaluate the antimicrobial activity of endophytic fungi from Mammea americana and Moringa oleifera upon Staphylococcus aureus (ATCC 29213), S. aureus (resistant strain USb003), Escherichia coli (ATCC 25922), and E. coli (resistant strain USb007). Materials and methods: We isolated endophytic fungi from the leaves, seeds, and stems of the two plants under study. We evaluated their antimicrobial activity through the formation of sensitivity haloes in dual tests in vitro, as well as in trials using crude ethanolic extracts from the endophytes. The minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC), and cytotoxicity o the substances were analyzed. Results: Three ethanolic extracts of Penicillium sp., Cladosporium (001), and Cladosporium (002) exhibited the greatest inhibition halos in sensitive and resistant strains of E. coli and S. aureus. The MIC and CBM found were statistically significant (p≤0.05) compared with the gentamicin control. Furthermore, the cytotoxicity test results of CC50>1,000 demonstrated that the endophytic fungi studied exhibit bactericidal characteristics without causing unintended damage. Conclusion: The endophytic fungi M. oleifera and M. americana represent a source of active secondary metabolites with antimicrobial and non-toxic properties. In light of these findings, further research should proceed with chemical identification of the compounds and the study of their mechanisms of action, especially given the paucity of current scientific knowledge concerning the isolation of endophytes in these plants.

Introducción. Las enfermedades infecciosas son una causa importante de muertes en el mundo. La resistencia antimicrobiana es un problema global, por lo que es conveniente la investigación de nuevas fuentes de agentes antimicrobianos de origen natural potencialmente efectivos. Objetivo. Evaluar la actividad antimicrobiana de hongos endófitos de Mammea americana y Moringa oleifera en la cepa sensible (ATCC 29213) y en la cepa resistente (USb003) de Staphylococcus aureus, así como en la cepa sensible (ATCC 25922) y la cepa resistente (USb007) de Escherichia coli. Materiales y métodos. Se aislaron 14 hongos endófitos de las hojas, semillas y tallos de las dos plantas en estudio. Se evaluó su actividad antimicrobiana mediante la formación de halos de sensibilidad por ensayo dual in vitro y pruebas con extractos etanólicos crudos provenientes de los endófitos a los que se les evaluó la concentración mínima inhibitoria (CMI), la concentración bactericida mínima (CBM) y la citotoxicidad. Resultados. Tres extractos etanólicos de Penicillium sp., Cladosporium sp. (001) y Cladosporium sp. (002) presentaron mayores halos de inhibición en cepas sensibles y resistentes de E. coli y S. aureus. La CMI y la CBM halladas fueron estadísticamente significativas (p≤0,05), comparadas con el control de gentamicina. Las pruebas de citotoxicidad (concentración citotóxica, CC50>1.000) demostraron que los hongos endófitos poseen características bactericidas y no ocasionan daño alguno. Conclusión. Se halló una fuente de metabolitos secundarios activos con propiedades antimicrobianas y no tóxicas en los hongos endófitos de M. oleifera y M. americana; estos hallazgos son importantes para continuar con la identificación química de los compuestos y el estudio de sus mecanismos de acción en estas plantas en las que el aislamiento de endófitos ha sido escaso.

Metabolite Profiling by UPLC-MSE, NMR, and Antioxidant Properties of Amazonian Fruits: Mamey Apple (Mammea Americana), Camapu (Physalis Angulata), and Uxi (Endopleura Uchi).

The metabolite profiling associated with the antioxidant potential of Amazonian fruits represents an important step to the bioactive compound's characterization due to the large biodiversity in this region. The comprehensive bioactive compounds profile and antioxidant capacities of mamey apple (Mammea americana), camapu (Physalis angulata), and uxi (Endopleura uchi) was determined for the first time. Bioactive compounds were characterized by ultra-performance liquid chromatography coupled to high resolution mass spectrometry (UPLC-MSE) in aqueous and ethanolic extracts. Globally, a total of 293 metabolites were tentatively identified in mamey apple, campau, and uxi extracts. The main classes of compounds in the three species were terpenoids (61), phenolic acids (58), and flavonoids (53). Ethanolic extracts of fruits showed higher antioxidant activity and total ion abundance of bioactive compounds than aqueous. Uxi had the highest values of phenolic content (701.84 mg GAE/100 g), ABTS (1602.7 µmol Trolox g-1), and ORAC (15.04 µmol Trolox g-1). Mamey apple had the highest results for DPPH (1168.42 µmol TE g-1) and FRAP (1381.13 µmol FSE g-1). Nuclear magnetic resonance (NMR) spectroscopy results showed that sugars and lipids were the substances with the highest amounts in mamey apple and camapu. Data referring to chemical characteristics and antioxidant capacity of these fruits can contribute to their economic exploitation.

EFFECT OF THAI SARAPHI FLOWER EXTRACTS ON WT1 AND BCR/ABL PROTEIN EXPRESSION IN LEUKEMIC CELL LINES.

BACKGROUND: Saraphi (Mammea siamensis) is a Thai traditional herb. In this study, the cytotoxic effects of crude ethanolic and fractional extracts including hexane, ethyl acetate, and methanol fractions from M. siamensis flowers were investigated in order to determine their effect on WT1 expression in Molt4 and K562 cells and Bcr/Abl expression in K562 cells. MATERIALS AND METHODS: The flowers of M. siamensis were extracted using ethanol. The ethanol flower extract was further fractionated with hexane, ethyl acetate, and methanol. Cytotoxic effects were measured by the MTT assay. Bcr/Abl and WT1 protein levels after treatments were determined by Western blotting. The total cell number was determined via the typan blue exclusion method. RESULTS: The hexane fraction showed the strongest cytotoxic activity on Molt4 and K562 cells, with IC50 values of 2.6 and 77.6 µg/ml, respectively. The hexane extract decreased Bcr/Abl protein expression in K562 cells by 74.6% and WT1 protein expressions in Molt4 and K562 cells by 68.4 and 72.1%, respectively. Total cell numbers were decreased by 66.2 and 48.7% in Molt4 and K562 cells, respectively. Mammea E/BB (main active compound) significantly decreased both Bcr/Abl and WTlprotein expressions by 75 and 49.5%, respectively when compared to vehicle control. CONCLUSION: The hexane fraction from M. siamensis flowers inhibited cell proliferation via the suppression of WT1 expression in Molt4 and K562 cells and Bcr/Abl expression in K562 cells. The active compound may be mammea E/BB. Extracts from M. siamensis flowers show promise as naturally occurring anti-cancer drugs.

Mammea longifolia Planch. and Triana Fruit Extract Induces Cell Death in the Human Colon Cancer Cell Line, SW480, via Mitochondria-Related Apoptosis and Activation of p53.

The methanol extract of Mammea longifolia Planch. and Triana (M. longifolia) fruit was studied for anticancer and apoptotic effects in the SW480 colon cancer cell line. The apoptotic and necrotic effects of M. longifolia were detected by 3-(4,5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2H tetrazolium bromide (MTT) and lactate dehydrogenase assays, respectively. One hundred µg/mL of the extract killed ∼82.4% of the cells; however, 2% of the death was related to necrosis. The morphological changes in M. longifolia-stimulated SW480 cells were observed directly by light microscopy. DNA fragmentation assay was employed to analyze the apoptosis induction. M. longifolia-treated SW480 cells promoted the expression of Bax, Bad, cleaved-poly-ADP-ribose polymerase (PARP), and p53 proteins and decreased the protein expression of pro-caspases Bcl-2 and Bcl-XL. The ratios of Bax/Bcl-2 and cleaved-PARP/PARP, predictive markers of apoptotic stimuli in cancer, increased and may play an important role in regulating the progression of apoptosis. The results suggested that M. longifolia induces cell death via mitochondrial-related apoptosis in SW480 cells.

Variability of traits and bioactive compounds in the fruit and pulp of six mamey apple (Mammea americana L.) accessions.

Our objective was to compare fruit morphology, physico-chemistry and bioactive compounds content of the edible pulp of six Mammea americana accessions. The results showed that this fruit was rather big, weighing on average 600 to 1100g depending on the accession, and spherical to oblate-shaped. The pulp represented between 50 and 70% of the weight of the whole fruit. The pulp adhered only partially to the seeds in 5 of the 6 accessions studied, while the last one exhibited full adherence. The fresh pulp was acidic, sweet, succulent and crunchy. The fruits studied had a variety of qualities, providing various opportunities for post-harvest uses: fruit salads, nectar preparation, jams and jellies, or export. We have established for the first time the total phenolic compounds and total flavonoids contents in the pulp of mamey apple fruits. The pulp colour was highly correlated with total phenolic compounds and total carotenoids contents.

Antiviral effect of compounds derived from the seeds of Mammea americana and Tabernaemontana cymosa on Dengue and Chikungunya virus infections.

BACKGROUND: The transmission of Dengue virus (DENV) and Chikungunya virus (CHIKV) has increased worldwide, due in part to the lack of a specific antiviral treatment. For this reason, the search for compounds with antiviral potential, either as licensed drugs or in natural products, is a research priority. The objective of this study was to identify some of the compounds that are present in Mammea americana (M. americana) and Tabernaemontana cymosa (T. cymosa) plants and, subsequently, to evaluate their cytotoxicity in VERO cells and their potential antiviral effects on DENV and CHIKV infections in those same cells. METHODS: Dry ethanolic extracts of M. americana and T. cymosa seeds were subjected to open column chromatographic fractionation, leading to the identification of four compounds: two coumarins, derived from M. americana; and lupeol acetate and voacangine derived from T. cymosa.. The cytotoxicity of each compound was subsequently assessed by the MTT method (at concentrations from 400 to 6.25 µg/mL). Pre- and post-treatment antiviral assays were performed at non-toxic concentrations; the resulting DENV inhibition was evaluated by Real-Time PCR, and the CHIKV inhibition was tested by the plating method. The results were analyzed by means of statistical analysis. RESULTS: The compounds showed low toxicity at concentrations ≤ 200 µg/mL. The compounds coumarin A and coumarin B, which are derived from the M. americana plant, significantly inhibited infection with both viruses during the implementation of the two experimental strategies employed here (post-treatment with inhibition percentages greater than 50%, p < 0.01; and pre-treatment with percentages of inhibition greater than 40%, p < 0.01). However, the lupeol acetate and voacangine compounds, which were derived from the T. cymosa plant, only significantly inhibited the DENV infection during the post-treatment strategy (at inhibition percentages greater than 70%, p < 0.01). CONCLUSION: In vitro, the coumarins are capable of inhibiting infection by DENV and CHIKV (with inhibition percentages above 50% in different experimental strategies), which could indicate that these two compounds are potential antivirals for treating Dengue and Chikungunya fever. Additionally, lupeol acetate and voacangine efficiently inhibit infection with DENV, also turning them into promising antivirals for Dengue fever.

Aromatase Inhibitory Activity of Geranylated Coumarins, Mammeasins C and D, Isolated from the Flowers of Mammea siamensis.

A methanol extract of the flowers of Mammea siamensis (Calophyllaceae) was found to inhibit enzymatic activity against aromatase (IC50=16.5 µg/mL). From the extract, two new geranylated coumarins, mammeasins C (1) and D (2), were isolated together with seven coumarins: 8-hydroxy-5-methyl-7-(3,7-dimethyl-octa-2,6-dienyl)-9-(2-methyl-1-oxobutyl)-4,5-dihydropyrano[4,3,2-de]chromen-2-one (9), 8-hydroxy-5-methyl-7-(3,7-dimethyl-octa-2,6-dienyl)-9-(3-methyl-1-oxobutyl)-4,5-dihydropyrano[4,3,2-de]chromen-2-one (10), mammeas A/AA (14), A/AB (15), A/AA cyclo D (18), E/BA (23), and E/BC cyclo D (25). The structures of 1 and 2 were elucidated on the basis of spectroscopic evidence. Among the isolates including 17 previously reported coumarins, 1 (IC50=2.7 µM), 2 (3.6 µM), and mammea B/AB cyclo D (21, 3.1 µM) showed relatively strong inhibitory activities comparable to the activity of the synthetic nonsteroidal aromatase inhibitor aminoglutethimide (2.0 µM).

Down-regulatory mechanism of mammea E/BB from Mammea siamensis seed extract on Wilms' Tumor 1 expression in K562 cells.

BACKGROUND: Wilms' tumor 1 (WT1) is a biological marker for predicting leukemia progression. In this study, mammea E/BB, an active compound from Saraphi (Mammea siamensis) seed extract was examined for its effect on down-regulatory mechanism of WT1 gene expression, WT1 protein and mRNA stability, and cell proliferation in K562 cell line. METHODS: M. siamensis seeds were obtained from the region of Chiang Mai (North of Thailand). Mammea E/BB was extracted from seeds of M. siamensis. WT1 protein expression and stability were evaluated by Western blot analysis. WT1 mRNA stability was assessed by qRT-PCR. WT1-DNA binding and WT1 promoter activity were assayed by ChIP assay and luciferase-reporter assay, respectively. Cell cycle arrest was studied by flow cytometry. RESULTS: Treatment with mammea E/BB led to down-regulation of WT1 expression. The suppression of WT1 expression did not involve protein and mRNA degradation. Rather, WT1 protein was down-regulated through disruption of transcriptional auto-regulation of the WT1 gene. Mammea E/BB inhibited WT1-DNA binding at the WT1 promoter and decreased luciferase activity. It also disrupted c-Fos/AP-1 binding to the WT1 promoter via ERK1/2 signaling pathway and induced S phase cell cycle arrest in K562 cells. CONCLUSION: Mammea E/BB had pleotropic effects on kinase signaling pathways, resulting in inhibition of leukemia cell proliferation.

Inhibitory effect of mammea E/BB from Mammea siamensis seed extract on Wilms' tumour 1 protein expression in a K562 leukaemic cell line.

Mammea siamensis is used in traditional Thai medicine. This study was designed to extract and isolate an active compound from the M. siamensis seeds and to investigate its activity on Wilms' tumour 1 (WT1) protein expression in K562 cells. WT1 is a transcription factor that stimulates cell proliferation. The ethanol saraphi seed (ESS) extract was fractionated using n-hexane, ethyl acetate, n-butanol and water to obtain n-hexane saraphi seed (HSS), ethyl acetate saraphi seed (EASS), n-butanol saraphi seed (BSS), and water saraphi seed (WSS) extracts, respectively. The ESS, HSS and EASS extracts had strong cytotoxic effects on K562 cells in the MTT assay. All three fractions decreased WT1 protein levels and decreased total cell numbers. The HSS extract decreased the WT1 protein levels in a time- and dose-dependent manner. HPLC and NMR analyses indicated that the active compound of HSS was mammea E/BB. M. siamensis seeds are thus identified as a promising source of bioactive compounds for potential inhibition of WT1 protein expression.

Identification of Minor Benzoylated 4-Phenylcoumarins from a Mammea neurophylla Bark Extract.

Through dereplication analysis, seven known Mammea coumarins were identified in a fraction obtained from Mammea neurophylla dichloromethane bark extract selected for its ability to prevent advanced glycation end-product (AGE) formation. Among them, a careful examination of the NMR dataset of pedilanthocoumarin B led to a structural revision. Inspection of LC-DAD-MS(n) chromatograms allowed us to predict the presence of four new compounds, which were further isolated. Using spectroscopic methods (¹H-, (13)C- and 2D-NMR, HRMS, UV), these compounds were identified as new benzoyl substituted 4-phenylcoumarins (iso-pedilanthocoumarin B and neurophyllol C) and 4-(1-acetoxypropyl)coumarins cyclo F (ochrocarpins H and I).

In vitro cytotoxic activity of Sa-Tri-Lhung-Klod remedy and its herbal ingredients on ovarian and cervical carcinoma cell lines.

BACKGROUND: Sa-Tri-Lhung-Klod is a Thai traditional medicine remedy for postpartum in the lists of The National Drug List ofHerbal Medicine Products AD. It consists ofseventeen herbs and were obtained by maceration and used in the form of liquor for women's health care such as treatment ofamenorrhea, menopause and blood tonic. In addition, it also usedfor postpartum care for being anti-inflammation in postpartum and prevention of cancer in women. OBJECTIVE: To investigate cytotoxic activity ofSa-Tri-Lhung-Klod remedy extracts and its herbal ingredients against human ovarian carcinoma cell line (SKOV-3) and cervical adenocarcinoma (HeLa) cell line. MATERIAL AND METHOD: Sa-Tri-Lhung-Klod remedy and its plant ingredients were extracted by maceration in 95% ethanol and dried using evaporator. All extracts were testedfor cytotoxic activity by sulforhodamine B (SRB) assay. RESULTS: Ethanolic extract ofSa-Tri-Lhung-Klod remedy displayed cytotoxic activity against SKOV-3 and HeLa with IC50 values of 72.84±1.07 and 47.24±2.83 µg/ml, respectively. It was classified as "less-active" according to the NCI guideline. However, Caesalpinia sappan, Mammea siamensis and Curcuma comosa showed high cytotoxic activity against SKOV-3 with IC50 values of 9.55±1.38 13.45±0.82 and 14.21±1.30 µg/ml, respectively. The ethanolic extracts ofCaesalpiniasappan and Mammea siamensis also exhibited cytotoxic activity against HeLa with IC50 values of 6.30±0.06 and 7.72±0.11 µg/ml, respectively. CONCLUSION: These results support the use of Sa-Tri-Lhung-Klod remedy in Thai traditional medicine for preventing of ovarian cancer and cervical cancer Caesalpinia sappan, Curcuma comosa and Mammea siamensis were strikingly active against ovarian and cervical cancer cells. Their extracts have the potential for developing as new anti-cancer drugs for women health.

Advanced glycation inhibition and protection against endothelial dysfunction induced by coumarins and procyanidins from Mammea neurophylla.

Advanced glycation end-products (AGEs) are associated with many pathogenic disorders such as pathogenesis of diabetes or endothelial dysfunction leading to cardiovascular events. Therefore, the identification of new anti-AGE molecules or extracts aims at preventing such pathologies. Many Clusiaceae and Calophyllaceae species are used in traditional medicines to treat arterial hypertension as well as diabetes. Focusing on these plant families, an anti-AGE plant screening allowed us to select Mammea neurophylla for further phytochemical and biological studies. Indeed, both DCM and MeOH stem bark extracts demonstrated in vitro their ability to prevent inflammation in endothelial cells and to reduce vasoconstriction. A bioguided fractionation of these extracts allowed us to point out 4-phenyl- and 4-(1-acetoxypropyl)coumarins and procyanidins as potent inhibitors of AGE formation, potentially preventing endothelial dysfunction. The fractionation steps also led to the isolation of two new compounds, namely neurophyllols A and B from the DCM bark extract together with thirteen known mammea A and E coumarins (mammea A/AA, mammea A/AB, mammea A/BA, mammea A/BB, mammea A/AA cycloD, mammea A/AB cycloD, disparinol B, mammea A/AB cycloE, ochrocarpin A, mammea A/AA cycloF, mammea A/AB cycloF, mammea E/BA, mammea E/BB) as well as δ-tocotrienol, xanthones (1-hydroxy-7-methoxyxanthone, 2-hydroxyxanthone) and triterpenes (friedelin and betulinic acid). During this study, R,S-asperphenamate, previously described from fungal origin was also purified.