The Identification and Heterologous Expression of the Biosynthetic Gene Cluster Encoding the Antibiotic and Anticancer Agent Marinomycin.

With the rise in antimicrobial resistance, there is an urgent need for new classes of antibiotic with which to treat infectious disease. Marinomycin, a polyene antibiotic from a marine microbe, has been shown capable of killing methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant Enterococcus faecium (VREF), as well as having promising activity against melanoma. An attractive solution to the photoprotection of this antibiotic has been demonstrated. Here, we report the identification and analysis of the marinomycin biosynthetic gene cluster (BGC), and the biosynthetic assembly of the macrolide. The marinomycin BGC presents a challenge in heterologous expression due to its large size and high GC content, rendering the cluster prone to rearrangement. We demonstrate the transformation of Streptomyces lividans using a construct containing the cluster, and the heterologous expression of the encoded biosynthetic machinery and production of marinomycin B.

A natural solution to photoprotection and isolation of the potent polyene antibiotic, marinomycin A.

The photoprotection and isolation of marinomycin A using sporopollenin exine capsules (SpECs) derived from the spores of the plant Lycopodium clavatum is described. The marinomycins have a particularly short half-life in natural light, which severely impacts their potential biological utility given that they display potent antibiotic and anticancer activity. The SpEC encapsulation of the marinomycin A dramatically increases the half-life of the polyene macrodiolide to the direct exposure to UV radiation by several orders of magnitude, thereby making this a potentially useful strategy for other light sensitive bioactive agents. In addition, we report that the SpECs can also be used to selectively extract culture broths that contain the marinomycins, which provides a significantly higher recovery than with conventional XAD resins and provides concomitant photoprotection.

Antiviral activity of Salidroside from the leaves of Nigerian mistletoe (Loranthus micranthus Linn) parasitic on Hevea brasiliensis against respiratory syncytial virus.

Isolated Salidroside from the leaves of Nigerian mistletoe (Loranthus micranthus Linn) parasitic on Hevea brasiliensis was evaluated for its antiviral activity against respiratory syncytial virus. Semi- preparative HPLC separation of the ethyl acetate fraction of the leave extract of Loranthus micranthus Linn parasitic on Hevea brasiliensis led to the isolation of a polyphenol. Using spectroscopic methods (1D and 2D NMR and mass spectroscopic data) as well as by comparison with literature data the structure of the compound was determined as 6-O-galloyl salidroside. The antiviral activity of the isolated compound was evaluated against the respiratory syncytial virus. The isolated Salidroside showed potent inhibition towards a recombinant straining respiratory syncytial virus with Inhibitory Concentration (IC 50) value of 10.3±1.50 µg/mL. The result indicates that Salidroside is an efficient antiviral agent against RSV infection and might be useful for the management of RSV pathogenesis.

Modulation of intracellular expression of IFNγ and IL-2 in culture of splenic T lymphocytes by some flavonoid glycosides of Alchornea floribunda.

Context Alchornea floribunda Müll. Arg. (Euphorbiaceae) leaves are widely used in ethnomedicine for the management of rheumatism, arthritis and toothache. Objective In this study, flavonoid glycosides isolated from Alchornea floribunda were screened for their effect on the intracellular expression of interferon-gamma (IFNγ) and interleukin-2 (IL-2) type-1 cytokines. Materials and methods Chromatographic purification of the ethyl acetate fraction of the methanol leaf extract led to the isolation of seven flavonoid glycosides (1-7). Their structures were elucidated by 1D and 2D nuclear magnetic resonance and mass spectrometry. Splenocytes were treated with graded concentrations of the compounds (6.25-25 µg/mL) and incubated for 24 h. Thereafter, their effect on the expression of IFNγ and IL-2 by CD4(+ )and CD8(+ )T-lymphocytes was evaluated using intracellular cytokine staining and FACS analysis. Results Compounds 1-7 (6.25-25 µg/mL) caused the up-regulation of activated CD8(+ )(57.85-72.45% versus 57.85% for untreated control) and, to a lesser extent, activated CD4(+ )(3.21-7.21% versus 2.75% for the untreated control) T-lymphocytes that were both largely interferon-gamma-releasing in treated mouse T lymphocytes relative to untreated control. FACS data analysis showed that stimulation with all the compounds increased the proportion of CD8(+)/IFNγ(+ )and CD4(+)/IFNγ(+ )T lymphocytes up to two-fold when compared with the cells in untreated control wells. Intracellular IL-2 secretion by treated T cells was not detected. Conclusion This recorded T-lymphocyte-specific immune-modulatory property may contribute to explain in part the dynamics associated with the ethnomedicine of Alchornea floribunda, and may find relevance as a necessary cellular immune response precursor to infection-associated disease management.

New neolignan glycoside and an unusual benzoyl malic acid derivative from Maytenus senegalensis leaves.

Further investigation of the methanol leaf extract of Maytenus senegalensis led to the isolation of six compounds, including mayselignoside (1) and an unusual benzoyl malic acid derivative, benzoyl R-(+)-malic acid (2). Two known lignan derivatives (+)-lyoniresinol (3) and (-)-isolariciresinol (4), a known neolignan derivative dihydrodehydrodiconiferyl alcohol (5) and the triterpenoid, ß-amyrin (6) were also isolated. The structures of these compounds were elucidated by a combination of 1D and 2D NMR and mass spectroscopy. All compounds were tested for cytotoxicity against mouse lymphoma cell line (L5178Y) and for antimicrobial activity against strains of bacteria and fungi. None of the compounds showed promising cytotoxic and/or antimicrobial activities.

Isolation and structure elucidaton of polyphenols from Loranthus micranthus Linn. parasitic on Hevea brasiliensis with antiinflammatory property.

The present study was carried out to evaluate the anti-inflammatory activities of polyphenols isolated from the leaves of mistletoe (Loranthus micranthus Linn.) parasitic on Hevea brasiliensis. The anti-inflammatory properties of the isolated compounds were evaluated on the basis of their ability to inhibit the production of nitric oxide (NO) and tumuor necrosis factor-α (TNF-α) in lipopolysaccharide (LPS) activated RAW 264.7 mouse macrophages. Semi-preparative HPLC separation of the ethyl acetate (EtOAc) and butanol (n-BuOH) fractions of the leaves of mistletoe (Loranthus micranthus Linn) parasitic on Hevea brasiliensis led to the isolation of four polyphenols: 3-O-(3,4,5-trimethoxybenzoyl)-(-)-epicatechin (TMECG) (1); (-)-epicatechin-3-O-(3″-O-methyl)-gallate (ECG3″Me) (2); rutin (3) and peltatoside (4). Compounds 1-4 were isolated for the first time from this plant while 1 was isolated for the first time in nature. These compounds (1-4) were readily identified by comparison of their spectroscopic data with those reported in the literature. The polyphenols proved to have anti-inflammatory activity as evidenced by the suppression of inducible nitric oxide (iNO) and cytokine (TNF-α) levels in the culture supernatant of lipopolysaccharide (LPS)-stimulated RAW 264.7 murine macrophages. However, the study showed that the quercetin diglycosides showed stronger inhibition of proinflammatory mediators than the epicatechin derivates. These data provide evidence that polyphenolic compounds isolated from the mistletoe parasitic on Hevea brasiliensis may contribute to its anti-inflammatory properties by inhibiting the expression of inducible nitric oxide and proinflammatory cytokines such as tumour necrosis factor-α.

Antioxidative polyphenols from Nigerian mistletoe Loranthus micranthus (Linn.) parasitizing on Hevea brasiliensis.

Two new phenolic glycosides, linamarin gallate (1) and walsuraside B (2), together with nine known compounds, catechin (3), epicatechin (4), epicatechin 3-O-gallate (5), epicatechin 3-O-(3-O-methyl)gallate (6), epicatechin 3-O-(3,5-O-dimethyl)gallate (7), epicatechin 3-O-(3,4,5-O-trimethyl)gallate (8), quercetin 3-O-ß-d-glucopyranoside (9), rutin (10), and peltatoside (11), were isolated from the leafy twigs of Nigerian mistletoe Loranthus micranthus (Linn.) parasitic on Hevea brasiliensis. Compound 1 was characterized as an unusual cyanogenic glycoside, while compound 8 was isolated for the first time from a natural source. This is the first report of a cyanogenic glycoside from mistletoes. The structures of the new compounds were unambiguously elucidated by 1D ((1)H, (13)C), 2D NMR (COSY, HSQC, and HMBC) and by mass spectroscopy. The antioxidant activities of the isolated compounds (1-11) were evaluated using the 2, 2-diphenyl-1-picrylhydrazyl (DPPH) assay.

Antimotility effects of extracts and fractions of Eastern Nigeria mistletoe (Loranthus micranthus Linn).

OBJECTIVE: To evaluate the antimotility activity of Eastern Nigerian mistletoe[Loranthus micranthus (L. micranthus) Linn] parasitic on six different host trees viz. Baphia nitida, Persia americana, Kola accuminata, Irvingia gabonensis, Citrus simensis and Pentacletra macrophylla (P. mycrophylla). METHODS: The antimotility of the methanol extracts and solvent fractions were evaluated in castor oil induced diarrheoa in rats. RESULTS: The methanol extracts (200 mg/kg, i.p.) inhibited defeacation significantly (P < 0.05) 4 h after administration (75.73% to 93.33%) more than that of atropine sulphate (2 mg/kg, i.p.) which inhibited defeacation by 80.0%. The methanol extract (200 mg/kg, i.p.) of L. micranthus parasitic on P. mycrophylla exhibited significant (P<0.05) inhibition in gastrointestinal transit (67.6%) more than that of atropine sulphate (2 mg/kg, i.p.) which inhibited gastrointestinal transit by 26.4%. The solvent fractions of L. micranthus parasitic on P. mycrophylla at dose levels of 150 mg/kg inhibited significantly the gastrointestinal transit of mice. Fraction F(5) exhibited inhibitory activity which was comparable to loperamide (73.3%). CONCLUSION: The methanol extract of L. micranthus parasitic on P. macrophylla exhibits higher antimotility activity that other extracts. The solvent fractions could serve as source of novel antimotility agents.