Leptochelins A-C, Cytotoxic Metallophores Produced by Geographically Dispersed Leptothoe Strains of Marine Cyanobacteria.

Metals are important cofactors in the metabolic processes of cyanobacteria, including photosynthesis, cellular respiration, DNA replication, and the biosynthesis of primary and secondary metabolites. In adaptation to the marine environment, cyanobacteria use metallophores to acquire trace metals when necessary as well as to reduce potential toxicity from excessive metal concentrations. Leptochelins A-C were identified as structurally novel metallophores from three geographically dispersed cyanobacteria of the genus Leptothoe. Determination of the complex structures of these metabolites presented numerous challenges, but they were ultimately solved using integrated data from NMR, mass spectrometry and deductions from the biosynthetic gene cluster. The leptochelins are comprised of halogenated linear NRPS-PKS hybrid products with multiple heterocycles that have potential for hexadentate and tetradentate coordination with metal ions. The genomes of the three leptochelin producers were sequenced, and retrobiosynthetic analysis revealed one candidate biosynthetic gene cluster (BGC) consistent with the structure of leptochelin. The putative BGC is highly homologous in all three Leptothoe strains, and all possess genetic signatures associated with metallophores. Postcolumn infusion of metals using an LC-MS metabolomics workflow performed with leptochelins A and B revealed promiscuous binding of iron, copper, cobalt, and zinc, with greatest preference for copper. Iron depletion and copper toxicity experiments support the hypothesis that leptochelin metallophores may play key ecological roles in iron acquisition and in copper detoxification. In addition, the leptochelins possess significant cytotoxicity against several cancer cell lines.

What Is Authentic Maple Water? A Twelve-Month Shelf-Life Study of the Chemical Composition of Maple Water and Its Biological Activities.

Maple water (maple sap) products are produced from sap tapped directly from maple trees, but there is confusion and lack of industry consensus and consumer knowledge as to what constitutes 'authentic' maple water. With an immense potential for growth in the multi-billion dollar functional beverage market, the market promotion of maple water products hinges on establishing standards of identity (SI), which are currently lacking. Herein, we aim to provide publishable SI and compositional chemistry findings of maple water. The chemical composition (including polyphenols, sugars, amino acids, and organic acids) of a pasteurized maple water was monitored over a 12-month (at 0, 4, 8, and 12 months) shelf-life. Furthermore, LC-MS/MS and molecular networking-based methods were developed to identify the phytochemical profile of a maple water extract (MWX) and to compare it to a previously chemically characterized phenolic-enriched maple syrup extract (MSX). Both MSX and MWX have similar phytochemical profiles and chemical characteristics. In addition, MSX and MWX showed moderate antioxidant capacity (in free radical scavenging and anti-tyrosinase assays) and anti-inflammatory effects (in soluble epoxide hydrolase and cyclooxygenase-2 inhibition assays). Our findings provide critical information on the SI and stability (in chemical composition) of maple water, which will help define, authenticate, and distinguish it from other functional beverages, thereby positioning the maple industry for promotion and growth in this market sector.

Isolation of Isotrichophycin C and Trichophycins G-I from a Collection of Trichodesmium thiebautii.

The trichophycin family of compounds are chlorinated polyketides first discovered from environmental collections of a bloom-forming Trichodesmium sp. cyanobacterium. In an effort to fully capture the chemical space of this group of metabolites, the utilization of MS/MS-based molecular networking of a Trichodesmium thiebautii extract revealed a metabolome replete with halogenated compounds. Subsequent MS-guided isolation resulted in the characterization of isotrichophycin C and trichophycins G-I (1-4). These new metabolites had intriguing structural variations from those trichophycins previously characterized, which allowed for a comparative study to examine structural features that are associated with toxicity to murine neuroblastoma cells. Additionally, we propose the absolute configuration of the previously characterized trichophycin A (5). Overall, the metabolome of the Trichodesmium bloom is hallmarked by an unprecedented amount of chlorinated molecules, many of which remain to be structurally characterized.

A joint molecular networking study of a Smenospongia sponge and a cyanobacterial bloom revealed new antiproliferative chlorinated polyketides.

The bloom-forming cyanobacteria Trichodesmium sp. have been recently shown to produce some of the chlorinated peptides/polyketides previously isolated from the marine sponge Smenospongia aurea. A comparative analysis of extracts from S. aurea and Trichodesmium sp. was performed using tandem mass spectrometry-based molecular networking. The analysis, specifically targeted to chlorinated metabolites, showed that many of them are common to the two organisms, but also that some general differences exist between the two metabolomes. Following this analysis, six new chlorinated metabolites were isolated and their structures elucidated: four polyketides, smenolactones A-D (1-4) from S. aurea, and two new conulothiazole analogues, isoconulothiazole B (5) and conulothiazole C (6) from Trichodesmium sp. The absolute configuration of smenolactone C (3) was determined by taking advantage of the conformational rigidity of open 1,3-disubstituted alkyl chains. The antiproliferative activity of smenolactones was evaluated on three tumor cell lines, and they were active at low-micromolar or sub-micromolar concentrations.

Polyphenol Microbial Metabolites Exhibit Gut and Blood⁻Brain Barrier Permeability and Protect Murine Microglia against LPS-Induced Inflammation.

Increasing evidence supports the beneficial effects of polyphenol-rich diets, including the traditional Mediterranean diet, for the management of cardiovascular disease, obesity and neurodegenerative diseases. However, a common concern when discussing the protective effects of polyphenol-rich diets against diseases is whether these compounds are present in systemic circulation in their intact/parent forms in order to exert their beneficial effects in vivo. Here, we explore two common classes of dietary polyphenols, namely isoflavones and lignans, and their gut microbial-derived metabolites for gut and blood-brain barrier predicted permeability, as well as protection against neuroinflammatory stimuli in murine BV-2 microglia. Polyphenol microbial metabolites (PMMs) generally showed greater permeability through artificial gut and blood-brain barriers compared to their parent compounds. The parent polyphenols and their corresponding PMMs were evaluated for protective effects against lipopolysaccharide-induced inflammation in BV-2 microglia. The lignan-derived PMMs, equol and enterolactone, exhibited protective effects against nitric oxide production, as well as against pro-inflammatory cytokines (IL-6 and TNF-α) in BV-2 microglia. Therefore, PMMs may contribute, in large part, to the beneficial effects attributed to polyphenol-rich diets, further supporting the important role of gut microbiota in human health and disease prevention.

Trichophycins B-F, Chlorovinylidene-Containing Polyketides Isolated from a Cyanobacterial Bloom.

NMR-guided isolation (based on 1D 1H and 13C NMR resonances consistent with a chlorovinylidene moiety) resulted in the characterization of five new highly functionalized polyketides, trichophycins B-F (1-5), and one nonchlorinated metabolite tricholactone (6) from a collection of Trichodesmium bloom material from the Gulf of Mexico. The planar structures of 1-6 were determined using 1D and 2D NMR spectroscopy, mass spectrometry, and complementary spectroscopic procedures. Absolute configuration analysis of 1 and 2 were carried out by 1H NMR analysis of diastereomeric Mosher esters in addition to ECD spectroscopy, J-based configuration analysis, and DFT calculations. The absolute configurations of 3-6 were proposed on the basis of comparative analysis of 13C NMR chemical shifts, relative configurations, and optical rotation values to compounds 1 and 2. Compounds 1-5 represent new additions to the trichophycin family and are hallmarked by a chlorovinylidene moiety. These new trichophycins and tricholactone (1-6) feature intriguing variations with respect to putative biosynthetic starting units, halogenation, and terminations, and trichophycin E (4) features a rare alkynyl bromide functionality. The phenyl-containing trichophycins showed low cytotoxicity to neuro-2A cells, while the alkyne-containing trichophycins showed no toxicity.

Microcystins Containing Doubly Homologated Tyrosine Residues from a Microcystis aeruginosa Bloom: Structures and Cytotoxicity.

Four new microcystin congeners are described including the first three examples of microcystins containing the rare doubly homologated tyrosine residue 2-amino-5-(4-hydroxyphenyl)pentanoic acid (Ahppa) (1-4). Large-scale harvesting and biomass processing allowed the isolation of substantial quantities of these compounds, thus enabling complete structure determination by NMR as well as cytotoxicity evaluation against selected cancer cell lines. The new Ahppa-toxins all incorporate Ahppa residues at the 2-position, and one of these also has a second Ahppa at position 4. The two most lipophilic Ahppa-containing microcystins showed 10-fold greater cytotoxic potency against human tumor cell lines (A549 and HCT-116) compared to microcystin-LR (5). The presence of an Ahppa residue in microcystin congeners is difficult to ascertain by MS methods alone, due to the lack of characteristic fragment ions derived from the doubly homologated side chain. Owing to their unexpected cytotoxic potency, the potential impact of the compounds on human health should be further evaluated.

Tricholides A and B and Unnarmicin D: New Hybrid PKS-NRPS Macrocycles Isolated from an Environmental Collection of Trichodesmium thiebautii.

Bioassay-guided isolation of the lipophilic extract of Trichodesmium thiebautii bloom material led to the purification and structure characterization of two new hybrid polyketide-non-ribosomal peptide (PKS-NRPS) macrocyclic compounds, tricholides A and B (1 and 2). A third macrocyclic compound, unnarmicin D (3), was identified as a new depsipeptide in the unnarmicin family, given its structural similarity to the existing compounds in this group. The planar structures of 1-3 were determined using 1D and 2D NMR spectra and complementary spectroscopic and spectrometric procedures. The absolute configurations of the amino acid components of 1-3 were determined via acid hydrolysis, derivitization with Marfey's reagent and HPLC-UV comparison to authentic amino acid standards. The absolute configuration of the 3-hydroxydodecanoic acid moiety in 3 was determined using a modified Mosher's esterification procedure on a linear derivative of tricharmicin (4) and additionally by a comparison of 13C NMR shifts of 3 to known depsipeptides with ß-hydroxy acid subunits. Tricholide B (2) showed moderate cytotoxicity to Neuro-2A murine neuroblastoma cells (EC50: 14.5 ± 6.2 µM).

Trichophycin A, a Cytotoxic Linear Polyketide Isolated from a Trichodesmium thiebautii Bloom.

In an effort to isolate and characterize bioactive secondary metabolites from Trichodesmium thiebautii blooms, collected cyanobacteria biomass was subjected to bioassay-guided extraction and fractionation using the human colon cancer cell line HCT-116, resulting in the isolation and subsequent structure characterization of a linear polyketide trichophycin A (1). The planar structure of 1 was completed using 1D and 2D NMR spectroscopy and high-resolution electrospray ionization mass spectrometry (HRESIMS). Trichophycin A was moderately toxic against the murine neuroblastoma cell line Neuro-2A (EC50: 6.5 µM) and HCT-116 cells (EC50: 11.7 µM). Trichophycin A was significantly more cytotoxic than the previously isolated polyketides trichotoxin A and trichotoxin B. These cytotoxicity observations suggest that toxicity may be related to the polyol character of these polyketide compounds.

Kalkipyrone B, a marine cyanobacterial γ-pyrone possessing cytotoxic and anti-fungal activities.

Bioassay-guided fractionation of two marine cyanobacterial extracts using the H-460 human lung cancer cell line and the OVC-5 human ovarian cancer cell line led to the isolation of three related α-methoxy-ß, ß'-dimethyl-γ-pyrones each containing a modified alkyl chain, one of which was identified as the previously reported kalkipyrone and designated kalkipyrone A. The second compound was an analog designated kalkipyrone B. The third was identified as the recently reported yoshinone A, also isolated from a marine cyanobacterium. Kalkipyrone A and B were obtained from a field-collection of the cyanobacterium Leptolyngbya sp. from Fagasa Bay, American Samoa, while yoshinone A was isolated from a field-collection of cyanobacteria (cf. Schizothrix sp.) from Panama. One-dimensional and two-dimensional NMR experiments were used to determine the overall structures and relative configurations of the kalkipyrones, and the absolute configuration of kalkipyrone B was determined by (1)H NMR analysis of diastereomeric Mosher's esters. Kalkipyrone A showed good cytotoxicity to H-460 human lung cancer cells (EC50=0.9µM), while kalkipyrone B and yoshinone A were less active (EC50=9.0µM and >10µM, respectively). Both kalkipyrone A and B showed moderate toxicity to Saccharomyces cerevisiae ABC16-Monster strain (IC50=14.6 and 13.4µM, respectively), whereas yoshinone A was of low toxicity to this yeast strain (IC50=63.8µM).

Spongosine production by a Vibrio harveyi strain associated with the sponge Tectitethya crypta.

Spongosine (1), deoxyspongosine (2), spongothymidine (Ara T) (3), and spongouridine (Ara U) were isolated from the Caribbean sponge Tectitethya crypta and given the general name "spongonucleosides". Spongosine, a methoxyadenosine derivative, has demonstrated a diverse bioactivity profile including anti-inflammatory activity and analgesic and vasodilation properties. Investigations into unusual nucleoside production by T. crypta-associated microorganisms using mass spectrometric techniques have identified a spongosine-producing strain of Vibrio harveyi and several structurally related compounds from multiple strains.