Introduction Bioactive compounds with unique functional properties derived from marine bivalves have been gaining increasing attention. Marcia hiantina is a bivalve clam found in many coastal regions of the Philippines but is underutilized despite its nutritional value. The study aimed to isolate bioactive compounds from M. hiantina using a mass spectrometry-guided technique to separate target analytes and characterize their biological activities. Methods Bioactive fractions were detected by combining Liquid Chromatography-Mass Spectrometry (LC-MS) and High-Performance Liquid Chromatography (HPLC) with biological assays. The bioactive compounds were subsequently identified using Ultra-High-Performance Liquid Chromatography-Elevated Energy Mass Spectrometry (UHPLC-MSE). Results A UHPLC-MSE analysis of the isolate revealed polymeric Tryptophan (Trp) and its metabolites. The M. hiantina-derived peptide exhibited inhibitory effects on the proliferation of human breast cancer cells (MCF-7), with an IC50 value of 95.20 +/- 0.11 mu g/mL, measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Moreover, the peptide also inhibited the growth of both Gram-positive and Gram-negative bacterial strains and demonstrated strong antioxidant potential as a 2,2-diphenyl-1-picrylhydrazyl (DPPH) scavenger (73.54% at 2.5 mg/mL). Discussion The presence of Trp metabolites, including indole and indole-3-propionic acid in M. hiantina may result from the host-microbe interactions, or be influenced by environmental stress, as Trp requirements in clams increase under oxidative conditions, reflecting their adaptation to stressors like intermittent hypoxia and pollutants. Conclusion This study revealed that M. hiantina is a new source of bioactive compounds, and can be a promising novel ingredient in functional foods promoting health and well-being.

Carotenoids are a diverse class of biologically active compounds that contribute significantly to human health, serving vital functions in nutrition and overall well-being. Magallana bilineata, a commercially important oyster species, yields a shelf-stable powder residue possessing bioactivities with unknown specific compounds. Carotenoids are key marine bioactive compounds, but their presence in oysters remains underexplored. The present study aimed to identify the bioactive compounds from oyster powder residue through mass spectrometry for optimum utilization and value creation as a biomedical resource. The study employed solvent extraction of oyster powder residue, followed by fractionation using octadecylsilyl (ODS) column chromatography, liquid chromatography-mass spectrometry (LC-MS)-guided profiling, and ultra-high-performance liquid chromatography-ultra-high-performance liquid chromatography-elevated energy mass spectrometry-elevated energy mass spectrometry (UHPLC-MSE) analysis to identify carotenoids, with bioactivity assays conducted to assess the cytotoxic, antimicrobial, antioxidant, and anti-inflammatory properties. The study obtained two HPLC fractions and enabled the identification of carotenoid compounds based on retention times and UHPLC-MSE, with elemental compositions inferred from the observed mass-to-charge ratios. The bioactivities of the two HPLC fractions, identified as zeaxanthin in fraction 1 and a zeaxanthin/lutein isomeric mixture in fraction 2, were assessed. The zeaxanthin/lutein isomeric mixture exhibited higher effectiveness in MCF-7 cancer cell inhibition (IC50 = 93.29 ± 0.07 μg/mL) than cisplatin, but both HPLC fractions showed strong antibacterial activity against Klebsiella pneumoniae and Escherichia coli. Moreover, notable antioxidant activity was observed in both fractions for 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) radical scavenging activity assays, while zeaxanthin demonstrated anti-inflammatory activity (43.68 ± 0.11%) comparable to aspirin (43.49 ± 0.17%). These findings suggest that the observed biological activities of the HPLC fractions may be the consequences of the adaptive response and filter-feeding behaviors of oysters, which result in the accumulation of bioactive carotenoids. This study offers a promising perspective on applying mass spectrometry techniques for advanced compound extraction and identification, and on utilizing oyster powder residue as a sustainable approach to waste valorization and as a functional ingredient for biomedical applications.

The culturable endophytic bacteria from the weeds Cleome rutidosperma of the family Cleomaceae and Digitaria sanguinalis of the family Poaceae obtained from a previous dumpsite in Pampanga, Philippines have been assessed for their anti-methicillin-resistant Staphylococcus aureus (MRSA) activity, and the analytes with such activity should be identified. However, due to the limited amounts collected from the isolation process, 1.8 mg yield of compound 1 from the endophyte of C. rutidosperma and 1.2 mg of a mixture from the endophyte of D. sanguinalis were selected for LC-MSE analysis. The production of compounds from the culturable endophytic bacteria Pseudomonas aeruginosa- determined by gene-sequencing, an untargeted and data-independent analysis (DIA) by ultra-high performance liquid chromatography-high resolution-elevated energy mass spectrometry (UHPLC-HR-MSE) technique was employed to profile the metabolites present in the two high-performance liquid chromatography (HPLC) fractions. The analytes present from P. aeruginosa detected by UHPLC-HR-MSE isolated from C. rutidosperma was phenazine-1-carboxylic acid (1), and for D. sanguinalis were chamigrenal (2), dialkyl resorcinol (3), and a pyoverdine elicitor (4). This study proves that UHPLC-HR-MSE could identify the anti-MRSA constituents in P. aeruginosa from commensal weeds C. rutidosperma and D. sanguinalis. The UHPLC-HR-MSE could help strengthen metabolomics antibacterial research and its related applications from a future perspective. Application of metabolomics research using UHPLC-HR-MSE could enhance the rehabilitation of dumpsites by the microbial community present.

Chemical investigation of the cyanobacterium Dolichospermum sp. NIES-1697 afforded nostosin G (1), a linear tripeptide, spiroidesin B (2), and two known compounds, anabaenopeptins I (3) and J (4). Planar structures and absolute configurations for 1 and 2 were determined by 2D NMR, HRMS, Marfey's methodology, chiral-phase HPLC, and enzymatic degradation. Nostosin G (1) is a unique example of a linear peptide containing three subunits, 4-hydroxyphenyllactic acid (Hpla), homotyrosine (Hty), and argininal, with potent trypsin inhibitory properties. The biosynthetic gene clusters for nostosin G (1) and spiroidesin B (2) were investigated based on the genome sequence of Dolichospermum sp. NIES-1697.

Strain B-9, which has a 99% similarity to Sphingosinicella microcystinivorans strain Y2, is a Gram-negative bacterium with potential for use in the degradation of microcystin-related compounds and nodularin. We attempted to extend the application area of strain B-9 and applied it to mycotoxins produced by fungi. Among the tested mycotoxins, only ochratoxin A was completely hydrolyzed to provide the constituents ochratoxin α and l-phenylalanine, and levels of fumonisin B1 gradually decreased after 96 h. However, although drugs including antibiotics released into the aquatic environment were applied for microbial degradation using strain B-9, no degradation occurred. These results suggest that strain B-9 can only degrade amino acid-containing compounds. As expected, the tested compounds with amide and ester bonds, such as 3,4-dimethyl hippuric acid and 4-benzyl aspartate, were readily hydrolyzed by strain B-9, although the sulfonamides remained unchanged. The ester compounds were characteristically and rapidly hydrolyzed as soon as they came into contact with strain B-9. Furthermore, the degradation of amide and ester compounds with amino acids was not inhibited by the addition of ethylenediaminetetraacetic acid (EDTA), indicating that the responsible enzyme was not MlrC. These results suggest that strain B-9 possesses an additional hydrolytic enzyme that should be designated as MlrE, as well as an esterase.

The rise of bleeding and bleeding complications caused by oral anticoagulant use are serious problems nowadays. Strategies that block the initiation step in blood coagulation involving activated factor VII-tissue factor (fVIIa-TF) have been considered. This study explores toxic Microcystis aeruginosa K-139, from Lake Kasumigaura, Ibaraki, Japan, as a promising cyanobacterium for isolation of fVIIa-sTF inhibitors. M. aeruginosa K-139 underwent reversed-phase solid-phase extraction (ODS-SPE) from 20% MeOH to MeOH elution with 40%-MeOH increments, which afforded aeruginosin K-139 in the 60% MeOH fraction; micropeptin K-139 and microviridin B in the MeOH fraction. Aeruginosin K-139 displayed an fVIIa-sTF inhibitory activity of ~166 µM, within a 95% confidence interval. Micropeptin K-139 inhibited fVIIa-sTF with EC50 10.62 µM, which was more efficient than thrombin inhibition of EC50 26.94 µM. The thrombin/fVIIa-sTF ratio of 2.54 in micropeptin K-139 is higher than those in 4-amidinophenylmethane sulfonyl fluoride (APMSF) and leupeptin, when used as positive controls. This study proves that M. aeruginosa K-139 is a new source of fVIIa-sTF inhibitors. It also opens a new avenue for micropeptin K-139 and related depsipeptides as fVIIa-sTF inhibitors.

Mass spectrometry (MS) imaging is a useful tool for direct and simultaneous visualization of specific molecules. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) is used to evaluate the abundance of molecules in tissues using sample homogenates. To date, however, LC-MS/MS has not been utilized as an imaging tool because spatial information is lost during sample preparation. Here we report a new approach for LC-MS/MS imaging using a thermal film-based laser microdissection (LMD) technique. To isolate tissue spots, our LMD system uses a 808-nm near infrared laser, the diameter of which can be freely changed from 2.7 to 500 μm; for imaging purposes in this study, the diameter was fixed at 40 μm, allowing acquisition of LC-MS/MS images at a 40-μm resolution. The isolated spots are arranged on a thermal film at 4.5-mm intervals, corresponding to the well spacing on a 384-well plate. Each tissue spot is handled on the film in such a manner as to maintain its spatial information, allowing it to be extracted separately in its individual well. Using analytical LC-MS/MS in combination with the spatial information of each sample, we can reconstruct LC-MS/MS images. With this imaging technique, we successfully obtained the distributions of pilocarpine, glutamate, γ-aminobutyric acid, acetylcholine, and choline in a cross-section of mouse hippocampus. The protocol we established in this study is applicable to revealing the neurochemistry of pilocarpine model of epilepsy. Our system has a wide range of uses in fields such as biology, pharmacology, pathology, and neuroscience. Graphical abstract Schematic Indication of LMD-LC-MS/MS imaging.

The blood coagulation cascade involves the human coagulation factors thrombin and an activated factor VII (fVIIa). Thrombin and fVIIa are vitamin-K-dependent clotting factors associated with bleeding, bleeding complications and disorders. Thrombin and fVIIa cause excessive bleeding when treated with vitamin-K antagonists. In this research, we explored different strains of toxic Microcystis aeruginosa and cyanobacteria blooms for the probable fVIIa-soluble Tissue Factor (fVIIa-sTF) inhibitors. The algal cells were subjected to acidification, and reverse phase (ODS) chromatography-solid phase extraction eluted by water to 100% MeOH with 20%-MeOH increments except for M. aeruginosa NIES-89, from the National Institute for Environmental Studies (NIES), which was eluted with 5%-MeOH increments as an isolation procedure to separate aeruginosins 89A and B from co-eluting microcystins. The 40%-80% MeOH fractions of the cyanobacterial extract are active against fVIIa-sTF. The fVIIa-sTF active fractions from cultured cyanobacteria and cyanobacteria blooms were subjected to liquid chromatography-mass spectrometry (LC-MS). The 60% MeOH fraction of M. aeruginosa K139 exhibited an m/z 603 [M + H]⁺ attributed to aeruginosin K139, and the 40% MeOH fraction of M. aeruginosa NIES-89 displayed ions with m/z 617 [M - SO3 + H]⁺ and m/z [M + H]⁺ 717, which attributed to aeruginosin 89. Aeruginosins 102A/B and 298A/B were also observed from other toxic strains of M. aeruginosa with positive fVIIa-sTF inhibitory activity. The active fractions contained cyanobacterial peptides of the aeruginosin class as fVIIa-sTF inhibitors detected by LC-MS.

Serine proteases are involved in the blood coagulation cascade-composed of intrinsic, extrinsic, and common pathways affecting coagulation responses. Among the three pathways of the blood coagulation cascade involving serine proteases, this review focuses on serine protease inhibitors of the understudied extrinsic pathway, which is responsible for bleeding and bleeding complications targeting the (activated factor VII-Tissue Factor) fVIIa-TF complex. In lieu of our search for fVIIa-soluble TF (fVIIa-sTF) inhibitors or scaffolds, we review the serine protease inhibitors present as synthetic and natural products, which are involved in the blood coagulation cascade, and compare them with the serine protease peptide inhibitors found in toxic Microcystis cyanobacteria. Some scaffolds for the extrinsic pathway as inhibitors from the toxic Microcystis cyanobacteria will be identified in the review.

P hilippine marine biodiversity ranks among the richest in the world and is still largely unexplored. Such biodiversity can have profound applications in the search for compounds with anticancer properties. Our increased understanding of the molecular mechanisms in cancer has enabled us to demonstrate exactly how these compounds work. In the past two decades, compounds with diverse structures from Philippine marine organisms, ascidians, and associated microorganisms were reported. These compounds are cytotoxic to a variety of cancer cell lines and act on specific molecular targets in major cell signaling pathways implicated in cancer. To address the challenges associated with developing marine natural products for the pharmaceutical drug pipeline, future directions will focus on optimizing the culture of marine organisms and symbionts, utilizing biosynthetic genes in microbial hosts, and developing the concept of treating cancer with combinatorial therapy.

Bioassay-guided investigation of the cyanobacterium Anabaena compacta extracts afforded spumigin J (1) and the known thrombin inhibitor spumigin A (2). The absolute configuration of 1 was analyzed by advanced Marfey's methodology. Compounds 1 and 2 inhibited thrombin with EC(50) values of 4.9 and 2.1 μM, and 0.7 and 0.2 μM in the cathepsin B inhibitory assay, respectively. The MM-GBSA methodology predicted spumigin A with 2S-4-methylproline as the better thrombin inhibitor.

The bark of Ziziphus talanai (Blanco) Merrill, a tree endemic to the Philippines, is traditionally used in Antique to cure kidney problems such as urinary tract infections (UTI), and skin diseases such as scabies and ringworm. Its antimicrobial activity was screened against several microorganisms to have a more comprehensive anti-infective profile of the plant. The MeOH extract of Z. talanai bark exhibited antimicrobial activity against Gram-positive Mycobacterium phlei, S. aureus and B. subtilis at 20,000 mu g and 2,000 mu g but was inactive against Gram-negative E. coli, P aeruginosa, C. albicans and S. cerevisiae. It has minimal antifungal activity against T mentagrophytes. The MeOH extract of Z. talanai is therefore inactive against uncomplicated UTI, which is mostly caused by E. coli. Further bioassay-guided isolation and subsequent spectral analysis yielded ceanothic acid, whose activity is specific to M. phlei at 100 mu g. M. phlei causes chronic ambulatory peritoneal dialysis-associated peritonitis.