Anti-malarial activity of the alkaloid, heptaphylline, and the furanocoumarin, imperatorin, from Clausena anisata against human Plasmodium falciparum malaria parasites: ex vivo trophozoitocidal, schizonticidal and gametocytocidal approach.

BACKGROUND: The erythrocytic stage of the life cycle of the malaria parasite, Plasmodium falciparum, consists of trophozoite, schizont and gametocyte stages in humans. Various anti-malarial agents target different stages of the parasite to produce treatment outcomes. This study reports on the stage-specific anti-malarial activity of heptaphylline and imperatorin against human P. falciparum in addition to their cytotoxicity and selectivity indices (SI). METHODS: The compounds were isolated from Clausena anisata using column chromatography and their structures elucidated using NMR spectroscopy. The anti-malarial activity was determined by measuring the trophozoitocidal, schizonticidal and gametocytocidal activities of the compounds using the SYBR green assay. Cytotoxicity was evaluated using the tetrazolium-based colorimetric assay. RESULTS: Heptaphylline and imperatorin produced trophozoitocidal, schizonticidal and gametocytocidal activities with IC50s of 1.57 (0.2317)-26.92 (0.3144) µM with those of artesunate (the standard drug) being 0.00024 (0.0036)-0.0070 (0.0013) µM. In the cytotoxicity assay, the compounds produced CC50S greater than 350 µM and SI of 13.76-235.90. Also, the trophozoitocidal and schizonticidal activities of the compounds were more pronounced than their gametocytocidal activity. Imperatorin was 42.04% more trophozoitocidal than hepthaphyline. However, hepthaphyline has more schizonticidal and gametocytocidal properties than imperatorin. CONCLUSION: Heptaphylline and imperatorin are promising anti-malarial agents, since they possess potent anti-malarial activity with weak cytotoxicity on RBCs. However, imperatorin is a better anti-malarial prophylactic agent whereas heptaphylline is a better malaria treatment agent.

Panaxindole, a novel indole alkaloid N-glucoside from the leaves of Panax vietnamensis Ha et Grushv. (Vietnamese ginseng).

Vietnamese ginseng (Panax vietnamensis Ha and Grushv., Araliaceae) is indigenous in the central highlands of Vietnam and the southernmost distribution in the Panax genus. Like other ginseng, Vietnamese ginseng is well known has been used as a tonic and for management of certain diseases in the traditional medicine. Nevertheless, it is noteworthy that in respect to the long history in use and systematic studied on Korean ginseng (P. ginseng), American ginseng (P. quinquefolius), Japanese ginseng (P. japonicus), and Chinese ginseng (P. notoginseng), the up-to-date published database on Vietnamese ginseng is relatively much less extensive. In our ongoing research on the promising Vietnamese medicinal plants, the present phytochemical investigation of the ethanol extract of the leaves of Panax vietnamensis led to the isolation of three compounds (1-3), including a new indole alkaloid N-glycoside (1) and two known compounds. Their structures were elucidated based on extensive physiochemical and chemical methods, especially the interpretation of NMR and MS spectra. The absolute configuration of 1 was determined based on the comparison of its experimental and theoretical ECD spectra along with NMR calculation. Compound 1 is naturally isolated N-glycoside, which is rarely found in natural products. The isolated compounds showed weak or no inhibitory activity against acetylcholinesterase enzyme (AChE).

Triterpenoid esters from Capparis erythrocarpos (Isert), Capparaceae, root bark ameliorates CFA-induced arthritis.

The root bark of Capparis erythrocarpos (CERB) is employed to treat rheumatoid arthritis (RA) in Africa, particularly in Ghana. However, there was no isolation and characterization of the bioactive constituents responsible for the pharmacological actions of this plant. The aim of this study is to isolate, characterize and evaluate the anti-arthritic activity of the constituents of CERB. CERB was soxhleted and partitioned into various fractions. The constituents were isolated using column chromatography and characterized by 1D and 2D NMR spectroscopy. The precise carboxylic acid residues of the esters were determined using saponification, derivatization and GC-MS analysis. Anti-arthritic activity was evaluated in the CFA-induced arthritic model. Two triterpenoid esters namely, sitosterol 3-hexadecanoate or sitosterol 3-palmatate (1) and sitosterol 3-tetradecanoate or sitosterol 3-myristate (2) in addition to beta sitosterol (3) were isolated and characterized. Compounds 1 and 2 administered at 3 µmol/kg (p.o.) produced anti-inflammatory activity (P < 0.0001) of 31.02 and 39.14% respectively, in addition to arthritic score index (P < 0.0001) of 1.600 ± 0.2449 and 1.400 ± 0.2449 against CFA-induced arthritis which are equivalent to those of the standard drug, diclofenac sodium (DS), 3 µmol/kg (p.o.), (30.79% anti-inflammatory activity and 1.800 ± 0.3742 arthritic score index). The compounds produced similar anti-inflammatory effects as DS. Also, radiographical and histopathological studies showed that, the compounds and DS protected against bone destruction, inflammatory cells invasion into interstitial spaces and synovial liner hyperplasia of the joints. This is the first study to report the characterization of the constituents of C. erythrocarpos in addition to anti-arthritic activity of sitosterol 3-palmatate and sitosterol 3-myristate. These results provide the missing link between the chemistry and the pharmacological activities of C. erythrocarpos. The isolates also offer a different class of molecule which could provide alternative treatment for RA.

Iridoids from Morinda lucida, (Benth.) Rubiaceae, produced analgesic and anti-inflammatory activities via agonism at the kappa and delta opioid receptors, inhibition of COX-2 besides elevation of CAT and SOD activities.

ETHNOPHARMACOLOGICAL RELEVANCE: Pain and inflammation are the major symptoms of almost every human disease. Herbal preparations from Morinda lucida are used to treat pain and inflammation in traditional medicine. However, the analgesic and anti-inflammatory activities of some of the plant's chemical constituents are not known. AIM OF THE STUDY: The aim of this study is to evaluate the analgesic and anti-inflammatory activities and possible mechanisms of these activities of iridoids from Morinda lucida. MATERIAL AND METHODS: The compounds were isolated using column chromatography and characterized by NMR spectroscopy and LC-MS. Anti-inflammatory activity was evaluated using carrageenan-induced paw edema. Whereas, the analgesic activity was assessed in the hot plate and acetic acid-induced writhing assays. Mechanistic studies were conducted using pharmacological blockers, determination of antioxidant enzymes, lipid peroxidation, and docking studies. RESULTS: The iridoid, ML2-2 exhibited inverse dose-dependent anti-inflammatory activity (42.62% maximum at 2 mg/kg p. o). ML2-3 produced dose-dependent anti-inflammatory activity (64.52% maximum at 10 mg/kg p. o.). Anti-inflammatory activity of diclofenac sodium was 58.60% at 10 mg/kg p. o. Furthermore, ML2-2 and ML2-3 produced analgesic activity (P < 0.01) of 44.44 ± 5.84 and 54.18 ± 19.01%. at 10 mg/kg p. o. respectively in the hot plate assay and 64.88 and 67.44% in the writhing assay. ML2-2 significantly elevated catalase activity. However, ML2-3 elevated SOD and catalase activity significantly. In the docking studies, both iridoids formed stable crystal complexes with delta and kappa opioid receptors, and the COX-2 enzyme with very low free binding energies (ΔG) from -11.2 to -14.0 kcal/mol. However, they did not bind with the mu opioid receptor. The lower bound RMSD of most of the poses were found to be ≤ 2. Several amino acids were involved in the interactions through various inter molecular forces. CONCLUSION: These results indicate that ML2-2 and ML2-3 possessed very significant analgesic and anti-inflammatory activities via acting as both delta and kappa opioid receptor agonist, elevation of anti-oxidant activity and inhibition of COX-2.

Microorganisms for Ginsenosides Biosynthesis: Recent Progress, Challenges, and Perspectives.

Ginsenosides are major bioactive compounds present in the Panax species. Ginsenosides exhibit various pharmaceutical properties, including anticancer, anti-inflammatory, antimetastatic, hypertension, and neurodegenerative disorder activities. Although several commercial products have been presented on the market, most of the current chemical processes have an unfriendly environment and a high cost of downstream processing. Compared to plant extraction, microbial production exhibits high efficiency, high selectivity, and saves time for the manufacturing of industrial products. To reach the full potential of the pharmaceutical resource of ginsenoside, a suitable microorganism has been developed as a novel approach. In this review, cell biological mechanisms in anticancer activities and the present state of research on the production of ginsenosides are summarized. Microbial hosts, including native endophytes and engineered microbes, have been used as novel and promising approaches. Furthermore, the present challenges and perspectives of using microbial hosts to produce ginsenosides have been discussed.

New cycloartane coronalyl acetate and other terpenoids with anti-inflammatory activity from the leaves of Vietnamese Gardenia philastrei.

Phytochemical study on the methanolic extract of the leaves of Vietnamese plant Gardenia philastrei Pierre ex Pit. has led to the isolation of a new cycloartane coronalyl acetate (1) together with six known ones, coronlolide methyl ester (2), sootepin D (3), coronalolide (4), coronalolic acid (5), sootepin G (6) and 23-deoxojessic acid (7). Their structures were elucidated by a combination of 2 D NMR and HR-ESI-MS spectroscopies. These compounds (1-7) were tested for their anti-inflammatory activity. The result showed that six compounds (1-6) inhibit LPS-induced nitric oxide production in RAW264.7 macrophages with their IC50 values ranging from 3.76 - 75.47 µg/mL. This is the first report on the chemical constituents and anti-inflammatory activity of the G. philastrei.

Hirsutanone Isolated from the Bark of Alnus japonica Attenuates Melanogenesis via Dual Inhibition of Tyrosinase Activity and Expression of Melanogenic Proteins.

Hirsutanone (Hir) and oregonin (Ore) are diarylheptanoids isolated from the bark of Alnus japonica. In this study, we investigated the anti-melanogenic activity of Hir and Ore in B16-F1 murine melanoma and normal human epidermal melanocytes (HEMn-DP) and elucidated the mechanisms of action. In B16-F1 cells, Hir and Ore suppressed melanin synthesis induced by α-melanocyte-stimulating hormone (α-MSH) without cytotoxicity. The inhibitory effect of Hir on melanin synthesis was much stronger than that of Ore. In addition, Hir reduced melanin content in HEMn-DP cells. As tyrosinase is a key enzyme in melanin synthesis, the effect of Hir on tyrosinase activity was assessed. The results demonstrated that Hir partially decreased tyrosinase activity and intracellular tyrosinase activity. Moreover, Hir suppressed the protein expression of melanogenic enzymes, including tyrosinase, tyrosinase-related protein (TRP)-1, and TRP-2, leading to reduced melanin biosynthesis. Hir also led to the suppression of cAMP response element-binding protein (CREB) phosphorylation and microphthalmia-associated transcription factor (MITF) expression, which control the expression of melanogenic enzymes. These results suggest that Hir suppressed melanin synthesis by dual inhibition of tyrosinase activity and the CREB/MITF pathway leading to the expression of melanogenic enzymes and may be a potent cosmetic and therapeutic agent for hyperpigmentation disorders.

(+)-Magnolin Enhances Melanogenesis in Melanoma Cells and Three-Dimensional Human Skin Equivalent; Involvement of PKA and p38 MAPK Signaling Pathways.

Magnoliae Flos is a traditional herbal medicine used to treat nasal congestion associated with headache, empyema, and allergic rhinitis. In our preliminary screening of crude drugs used in Japanese Kampo formulas for melanin synthesis, the methanol extract of Magnoliae Flos was found to exhibit strong melanin synthesis activity. However, there have been no studies evaluating the effects of Magnoliae Flos or its constituents on melanogenesis. The present study aimed to isolate the active compounds from Magnoliae Flos that activate melanin synthesis in melanoma cells and three-dimensional human skin equivalent, and to investigate the molecular mechanism underlying melanin induction. The methanol extract of Magnoliae Flos induced an increase of melanin content in both B16-F1 and HMV-II cells. A comparison of melanin induction by three fractions prepared from the extract showed that the ethyl acetate fraction markedly induced melanin synthesis. Bioassay-guided separation of the ethyl acetate fraction resulted in the isolation of seven lignans (1:  - 7: ). Among them, (+)-magnolin (5: ) strongly induced melanin synthesis and intracellular tyrosinase activity. Furthermore, the ethyl acetate fraction and 5: clearly induced melanin content in a three-dimensional human skin equivalent. Molecular analysis revealed that 5: triggered the protein expression of tyrosinase, tyrosinase-related protein-1, and tyrosinase-related protein-2. Further analysis of transcriptional factors and signaling pathways demonstrated that 5: induces the protein expression of tyrosinase, tyrosinase-related protein-1, and tyrosinase-related protein-2 activated by the protein kinase A- and p38 mitogen-activated protein kinase-dependent pathways, leading to cAMP-responsive element-binding protein phosphorylation and microphthalmia-associated transcription factor expression. These findings demonstrate the potential of 5: as a potent therapeutic agent for hypopigmentation.

Majonoside-R2 extracted from Vietnamese ginseng protects H9C2 cells against hypoxia/reoxygenation injury via modulating mitochondrial function and biogenesis.

Vietnamese ginseng has a therapeutic effect on various diseases; however its bioactivity against cardiac hypoxia/reoxygenation (HR) injury remains unclear. In this study, we evaluated the protective roles of total saponin extract (TSE) and majonoside-R2 (MR2) targeting mitochondria in HR-induced rat cardiomyocyte H9C2 cells. The results showed that both TSE and MR2 effectively protected the cells from HR damage. Particularly, 9 µM of MR2 significantly increased the viability of HR-induced cells (p < 0.05). Interestingly, MR2 treatment markedly prevented the loss of mitochondrial membrane potential and cardiolipin content, and an increase in reactive oxygen species production in HR-treated H9C2 cells. Moreover, MR2 treatment altered the mRNA expression of genes involved in mitochondrial biogenesis under HR conditions. The present study documented for the first time the cardioprotective effects of MR2 against HR injury by maintaining mitochondrial function and modulating mitochondrial biogenesis.

Preventing Dementia Using Saffron, The Kampo Medicine, Kamiuntanto, and Their Combination, Kamiuntantokabankoka.

The objective of this review is to evaluate the anti-dementia activities of saffron and its combination with Kampo medicine. The Kampo formula Kamiuntanto composed of 13 crude drugs is well known for its anti-dementia activity. A significant increase in choline acetyltransferase activity and mRNA levels were observed. Polygala radix was identified as the most essential component drug in Kamiuntanto, probably due to the saponins, tenuifolin, and sinapinic acid. Ginseng was also identified as an essential Kamiuntanto component in terms of its synergistic functions with Polygala radix. Saffron, which was recommended in the Bencao Gangmu for memory and dementia, and is used as an anti-spasmodic, anti-catarrhal, and sedative herbal drug. Saffron and its major constituent, crocin were shown to enhance learning-memory, non-rapid eye movement (rem) sleep, and inhibit depression and neuronal cell death due to strong anti-oxidant and anti-inflammation activities. In addition based on the epidemiological studies such as the treatment of sleeping disorders and the clinical trials of saffron for Alzheimer patients, we demonstrated the indirect and direct anti-dementia activities of crocin and saffron.

The old pharmaceutical oleoresin labdanum of Cistus creticus L. exerts pronounced in vitro anti-dengue virus activity.

ETHNOPHARMACOLOGICAL RELEVANCE: The haemorrhagic dengue fever affects up to 500 million patients, annually causing 20.000 deaths, with no chemotherapeutic agent available. The oleoresin labdanum of Cistus creticus L. has been established as an anti-infective agent since antiquity in Mediterranean ethnopharmacology. MATERIALS AND METHODS: We tested several extracts and fractions of labdanum - standardised on labdane-type diterpenes via GC-MS - on their activity against the dengue virus (DENV-2 strain 00st-22A) in in vitro Vero cell cultures (96-well-plates, 5 days). Preliminary experiments with a labdanum diethyl ether raw-extract did not yield measureable results due to cytotoxic effects against Vero cells. In all following experiments, cell viability was constantly checked using the MTT-test. Fractionation of this raw-extract by liquid-liquid-extraction and column-chromatography on silica-gel (gradient elution with hexane, EtOAc, CHCl3, MeOH) succeeded in separating the anti-viral activity of labdanum from its cytotoxic effect. RESULTS: In the most active fraction GS5 at 30 µg/ml, dengue virus proliferation was 100% suppressed and cell viability was over 90%. Structural elucidation of major constituents of GS5 is currently ongoing, but thin-layer chromatography showed that this fraction is mainly dominated by manoyloxides, a class of labdane-type diterpenes with known antimicrobial activity. Claims concerning the antiviral activity of above ground parts of C. creticus have been made previously, but these generally ascribe this activity to hot water soluble polyphenols and propose an unspecific tanning effect of the viral surface proteins as the mechanism of action. However, the water soluble fraction enhanced viral proliferation. CONCLUSION: We therefore describe a direct, pharmacological, antiviral activity of a diethyl ether extract of labdanum against a virulent haemorrhagic fever like dengue for the first time.

Establishment of a quantitative and qualitative analysis and isolation method for tetracyclic iridoids from Morinda lucida Bentham leaves.

A new high performance liquid chromatography (HPLC) method has been established for quantitative and qualitative analysis of three tetracyclic iridoids: ML-2-3 (1), molucidin (2), and ML-F52 (3), which are responsible for anti-trypanosomal and anti-leishmanial activities of Morinda lucida Bentham leaves. Separation of 1-3 from dried 80% aqueous (aq.) ethanol extract was achieved on a reversed-phase cholester column packed with cholesteryl-bonded silica using an acetonitrile-0.1% aq. formic acid mobile phase system. Ultraviolet-visible (UV-VIS) spectroscopy was employed for detection of compounds, and their contents were determined by measuring absorbance at 254 nm. Depending on the above system, several factors potentially affecting the concentration of tetracyclic iridoids were evaluated resulting in several variation on plant organs, seasonality, variation between individual trees, and branch positions within the trees. Moreover, we developed a simple, quick, and effective method for tetracyclic iridoid isolation from M. lucida leaves that consisted of extraction by sonication into 80% aq. ethanol, basic hydrolysis, acid neutralization, liquid-liquid extraction into an organic solvent, and reverse phase open column chromatography. Employing this method, we have succeeded to obtain 1 as a colorless crystal yielding of 0.23%, which was 28 times higher than that of previous isolation method. Setting up methodology in this paper may be important for future in vitro and in vivo studies of tetracyclic iridoids and moreover for their applications in new drug design and development.

In vitro antiprotozoan activity and mechanisms of action of selected Ghanaian medicinal plants against Trypanosoma, Leishmania, and Plasmodium parasites.

Trypanosomiasis, leishmaniasis, and malaria are protozoan infections of public health importance with thousands of new cases recorded annually. Control of these infection(s) with existing chemotherapy is limited by drug toxicity, lengthy parenteral treatment, affordability, and/or the emergence of resistant strains. Medicinal plants on the other hand are used in the treatment of various infectious diseases although their chemical properties are not fully evaluated. In this study, we screened 112 crude extracts from 72 selected Ghanaian medicinal plants for anti-Trypanosoma, anti-Leishmania, and anti-Plasmodium activities in vitro and investigated their mechanisms of action. Twenty-three extracts from 20 plants showed significant antiprotozoan activity against at least 1 of 3 protozoan parasites screened with IC50 values less than 20 µg/ml. Eleven extracts showed high anti-Trypanosoma activity with Bidens pilosa whole plant and Morinda lucida leaf extracts recording the highest activities. Their IC50 (selectivity index [SI]) values were 5.51 µg/ml (35.00) and 5.96 µg/ml (13.09), respectively. Nine extracts had high anti-Leishmania activity with Annona senegalensis and Cassia alata leaf extracts as the most active. Their IC50 (SI) values were 10.8 µg/ml (1.50) and 10.1 µg/ml (0.37), respectively. Six extracts had high anti-Plasmodium activity with the leaf and stem-bark extracts of Terminalia ivorensis recording the highest activity. Their IC50 (SI) values were 7.26 µg/ml (129.36) and 17.45 µg/ml (17.17), respectively. Only M. lucida at 25 µg/ml induced significant apoptosis-like cell death in Trypanosoma parasites. Anti-Leishmania active extracts induced varying morphological changes in Leishmania parasites such as multiple nuclei and/or kinetoplast, incomplete flagella division, or nuclear fragmentation. Active extracts may be potential sources for developing new chemotherapy against these infections.

Antiproliferative activity and apoptosis induction by trijuganone C isolated from the root of Salvia miltiorrhiza Bunge (Danshen).

In this study, we found that the hexane fraction of Danshen, the dried root of Salvia miltiorrhiza (Lamiaceae), exerted antiproliferative effects on human leukemia cells. Phytochemical investigation of the hexane fraction achieved the isolation of the tanshinone diterpenes: dihydrotanshinone I (1), trijuganone C (2), trijuganone B (3), cryptotanshinone (4), tanshinone IIA (5), and tanshinone I (6). Compound 2 showed significant antiproliferative activities against human leukemia cells HL-60, Jurkat, and U937. The antiproliferative activities of 2 against human cancer and normal cells indicated that 2 exhibited potent antiproliferative activities with IC50 values less than 10 µM against HL-60 and Jurkat cells as well as on the colon cancer cells DLD-1, COLO 205, and Caco-2. Compound 2 induced chromatin condensation, DNA fragmentation, activation of caspase-3, -8, and -9, and the cleavage of poly (ADP-ribose) polymerase (PARP) in HL-60 cells. Moreover, 2 activated Bid and Bax, leading to the loss of mitochondrial membrane potential, and 2 induced the cytochrome c release from mitochondria into cytosol. In contrast, Bcl-2 and Bcl-xL were unaffected by 2. These results suggest that 2 exerts antiproliferative effects via apoptosis induction mediated by mitochondrial dysfunction and caspase activation. Compound 2 may serve as a candidate of potential chemotherapeutic agent for human leukemia.

Quassinoids from the Root of Eurycoma longifolia and Their Antiproliferative Activity on Human Cancer Cell Lines.

BACKGROUND: The roots of Eurycoma longifolia Jack have traditionally been used as an aphrodisiac tonic besides the other remedies for boils, fever, bleeding gums, and wound ulcer. Recently, the antiproliferative activity of E. longifolia has been reported and remained attractive to natural chemists. OBJECTIVE: The objective of this study was to study on antiproliferative compounds from the root of E. longifolia. MATERIALS AND METHODS: Column chromatography was used to separate individual compounds, spectroscopic data including nuclear magnetic resonances and mass spectrometry were analyzed to determine the chemical structure of the isolates and for biological testing, antiproliferative activity of compounds was tested on seven human cancer cell lines (KATO III, HCT-15, Colo205, HepG2, PC-3, Jurkat, HL-60) by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide method. RESULTS: Nine quassinoids including a new C19 longilactone-type quassinoid glycoside were characterized from the roots of the title plant. Among them, the major quassinoid eurycomanone exhibited selectively potential antiproliferative activities on the leukemia cell lines (HL-60 and Jurkat) and had very low toxic effects on normal skin fibroblast cell line (NB1RGB). CONCLUSION: The current study reveals one new quassinoid glycoside and the potential active component (eurycomanone) from E. longifolia for the leukemia treatment. SUMMARY: Nine quassinoids (1-9) including one new quassinoid glycoside (9) and eight known ones were isolated from the roots of Eurycoma longifoliaThe structure of the new quassinoid 9 was determined by extensive chemical and spectroscopic analysesThe major quassinoid, eurycomanone (3), exhibited selectively potential antiproliferative activities on both Jurkat and HL-60 leukemia cells and had very low toxic effects on normal skin fibroblast cell line (NB1RGB). Abbreviations used: COSY: Correlation spectroscopy; HMBC: Heteronuclear multiple-bond correlation; HMQC: Heteronuclear multiple quantum correlation; NMR: Nuclear magnetic resonance; NOESY: Nuclear Overhauser enhancement spectroscopy; TLC: Thin layer chromatography.

Bioactive Triterpenes from the Root of Salvia miltiorrhiza Bunge.

Danshen (Salvia miltiorrhiza) is a well-known medicinal herb in the oriental medicine. The current study on bioactive triterpenoid in the root of S. miltiorrhiza led to the isolation of a new highly hydroxylated ursane-type triterpene, urs-12-ene-2α,3ß,7ß,16α-tetraol (1) and five known ones including 2ß-hydroxypomolic acid (2), maslinic acid (3), asiatic acid (4), ursolic acid (5), and oleanolic acid (6). Their structures were elucidated on the basis of extensive spectroscopic analyses and comparison with literature data. The antiproliferative testing against HL-60 cells revealed that the new compound 1 and ursolic acid (5) showed weak and moderate activities with IC50 values of 42.2 and 11.7 µM. In addition, compounds 1-3 showed inhibitory effect on ghrelin activity. Copyright © 2017 John Wiley & Sons, Ltd.

Kayeassamin A Isolated from the Flower of Mammea siamensis Triggers Apoptosis by Activating Caspase-3/-8 in HL-60 Human Leukemia Cells.

BACKGROUND: Mammea siamensis (Miq.) T. Anders. is used as a medicinal plant in Thailand and has several traditional therapeutic properties. In a previous study, we isolated eight compounds from the flower of M. siamensis and demonstrated that kayeassamin A (KA) exhibited potent antiproliferative activity against human leukemia and stomach cancer cell lines. OBJECTIVE: In this study, we investigated the effect of KA on cell viability and apoptotic mechanisms in HL-60 human leukemia cells. MATERIALS AND METHODS: Cell viability was measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Nuclear morphology and DNA fragmentation were observed using Hoechst 33258 staining and agarose gel electrophoresis, respectively. The sub-G1 phase of cells was analyzed by flow cytometry after the cellular DNA had been stained with propidium iodide. The protein levels of poly (ADP-ribose) polymerase (PARP) and caspases were determined by Western blotting. RESULTS: KA exhibited a significant cytotoxic effect in a dose- and time-dependent manner, and induced chromatin condensation, DNA fragmentation, and sub-G1 phase DNA content, known as molecular events associated with the induction of apoptosis. In addition, KA strongly induced the activation of PARP and caspase-3 and -8, with weak caspase-9 activation. Furthermore, KA-induced DNA fragmentation was abolished by pretreatment with z-VAD-FMK (a broad caspase inhibitor), z-DEVD-FMK (a caspase-3 inhibitor), and z-IETD-FMK (a caspase-8 inhibitor), but not by z-LEHD-FMK (a caspase-9 inhibitor) pretreatment. CONCLUSION: These results indicate that KA triggers apoptotic cell death by activation of caspase-3 and -8 in HL-60 cells. SUMMARY: Kayeassamin A (KA) isolated from the flower of Mammea siamensis exhibited a significant cytotoxic effect in HL-60 human leukemia cells. KA triggers apoptotic cell death by activating caspase-3/-8. Abbreviations Used: KA: Kayeassamin A; MTT: 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide; PARP: Poly (ADP-ribose) polymerase; PI: Propidium iodide; CA: Corosolic acid.

In vitro anti-Leishmania activity of tetracyclic iridoids from Morinda lucida, benth.

Leishmaniasis is an infectious disease transmitted by the sand fly. It is caused by over 20 different species of Leishmania and has affected over 14 million people worldwide. One of the main forms of control of leishmaniasis is chemotherapy, but this is limited by the high cost and/or toxicity of available drugs. We previously found three novel compounds with an iridoid tetracyclic skeleton to have activity against trypanosome parasites. In this study, we determined the activity of the three anti-trypanosome compounds against Leishmania using field strain, 010, and the lab strain Leishmania hertigi. The minimum inhibitory concentration (MIC) of the compounds against 010 was determined by microscopy while the IC50 of compounds against L. hertigi was determined by fluorescence-activated cell sorting with Guava viacount analysis. We found two of the three compounds, molucidin and ML-F52, to have anti-Leishmania activity against both strains. The fluor-microscope observation with DAPI stain revealed that both Molucidin and ML-F52 induced abnormal parasites with two sets of nucleus and kinetoplast in a cell, suggesting that compounds might inhibit cytokinesis in Leishmania parasites. Molucidin and ML-F52 might be good lead compounds for the development of new anti-Leishmania chemotherapy.

The Effect of (1S,2S,3E,7E,11E)-3,7,11,15-Cembratetraen-17,2-Olide (LS-1) from Lobophyyum sp. on the Apoptosis Induction of SNU-C5 Human Colorectal Cancer Cells.

(1S,2S,3E,7E,11E)-3,7,11,15-cembratetraen-17,2-olide (LS-1), a marine cembrenolide diterpene, has anticancer activity against colon cancer cells such as HT-29, SNU-C5/5-FU (fluorouracil-resistant SNU-C5) and SNU-C5. However, the action mechanism of LS-1 on SNU-C5 human colon cancer cells has not been fully elucidated. In this study, we investigated whether the anticancer effect of LS-1could result from apoptosis via the modulation of Wnt/ß-catenin and the TGF-ß pathways. When treated with the LS-1, we could observe the apoptotic characteristics such as apoptotic bodies and the increase of sub-G1 hypodiploid cell population, increase of Bax level, decrease of Bcl-2 expression, cleavage of procaspase-3 and cleavage of poly (ADP-ribose) polymerase in SNU-C5 cells. Furthermore, the apoptosis induction of SNU-C5 cells upon LS-1 treatment was also accompanied by the down-regulation of Wnt/ß-catenin signaling pathway via the decrease of GSK-3ß phosphorylation followed by the decrease of ß-catenin level. In addition, the LS-1 induced the activation of TGF-ß signaling pathway with the decrease of carcinoembryonic antigen which leads to decrease of c-Myc, an oncoprotein. These data suggest that the LS-1 could induce the apoptosis via the down-regulation of Wnt/ß-catenin pathway and the activation of TGF-ß pathway in SNU-C5 human colon cancer cells. The results support that the LS-1 might have potential for the treatment of human colon cancer.

Antitrypanosomal Activities and Mechanisms of Action of Novel Tetracyclic Iridoids from Morinda lucida Benth.

Trypanosoma brucei parasites are kinetoplastid protozoa that devastate the health and economic well-being of millions of people in Africa through the disease human African trypanosomiasis (HAT). New chemotherapy has been eagerly awaited due to severe side effects and the drug resistance issues plaguing current drugs. Recently, there has been an emphasis on the use of medicinal plants worldwide. Morinda lucida Benth. is a popular medicinal plant widely distributed in Africa, and several research groups have reported on the antiprotozoal activities of this plant. In this study, we identified three novel tetracyclic iridoids, molucidin, ML-2-3, and ML-F52, from the CHCl3 fraction of M. lucida leaves, which possess activity against the GUTat 3.1 strain of T. brucei brucei The 50% inhibitory concentrations (IC50) of molucidin, ML-2-3, and ML-F52 were 1.27 µM, 3.75 µM, and 0.43 µM, respectively. ML-2-3 and ML-F52 suppressed the expression of paraflagellum rod protein subunit 2, PFR-2, and caused cell cycle alteration, which preceded apoptosis induction in the bloodstream form of Trypanosoma parasites. Novel tetracyclic iridoids may be promising lead compounds for the development of new chemotherapies for African trypanosomal infections in humans and animals.