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Background Pisum sativum: (PS) is a universal legume plant utilized for both human and animal consumption, particularly its seeds, known as green peas. The processing of PS in food industries and households produces a significant amount of waste that needs to be valorized. Methods: In this study, the metabolite profiles of the 70% ethanolic extracts of PS wastes, namely peels (PSP) and a combination of leaves and stems (PSLS), were investigated by liquid chromatography-electrospray ionization-quadrupole time-of-flight tandem mass spectrometry (LC-ESI-QTOF-MS/MS) followed by molecular networking. Results: Different classes of metabolites were identified, being flavonoids and their derivatives, along with phenolic acids, the most abundant categories. Additionally, a comprehensive network pharmacology strategy was applied to elucidate potentially active metabolites, key targets, and the pathways involved in cytotoxic activity against breast cancer. This cytotoxic activity was investigated in MCF-7 and MCF-10a cell lines. Results revealed that PSLS extract exhibited a potent cytotoxic activity with a good selectivity index (IC50 = 17.67 and selectivity index of 3.51), compared to the reference drug doxorubicin (IC50 = 2.69 µg/mL and selectivity index of 5.28). Whereas PSP extract appeared to be less potent and selective (IC50 = 32.92 µg/mL and selectivity index of 1.62). A similar performance was also observed for several polyphenolics isolated from the PSLS extract, including methyl cis p-coumarate, trans p-coumaric acid, and liquiritigenin/ 7-methyl liquiritigenin mixture. Methyl cis p-coumarate showed the most potent cytotoxic activity against MCF-7 cell line and the highest selectivity (IC50 = 1.18 µg/mL (6.91 µM) and selectivity index of 27.42). The network pharmacology study revealed that the isolated compounds could interact with several breast cancer-associated protein targets including carbonic anhydrases 1, 2, 4, 9, and 12, as well as aldo-keto reductase family 1 member B1, adenosine A3 receptor, protein tyrosine phosphatase non-receptor type 1, and estrogen receptor 2. Conclusion: The uncovered therapeutic potential of PSLS and its metabolite constituents pave the way for an efficient and mindful PS waste valorization, calling for further in-vitro and in-vivo research. [ABSTRACT FROM AUTHOR]

Streptomyces sp. VB1, an actinomycete isolated from marine sediments in Valparaíso Bay, Chile, synthesizes antimicrobial and antiproliferative compounds. This study presents comprehensive metabolomics and comparative genomics analyses of strain VB1. LC-HRMS dereplication and Molecular Networking analysis of crude extracts identified antibiotics such as globomycin and daunorubicin, along with known and potentially novel members of the arylomycin family. These compounds exhibit activity against a range of clinically relevant bacterial and cancer cell lines. Phylogenomic analysis underscores the uniqueness of strain VB1, suggesting it represents a novel taxon. Such uniqueness is further supported by its Biosynthetic Novelty Index (BiNI) and BiG-SCAPE analysis of Gene Cluster Families (GCFs). Notably, two Biosynthetic Gene Clusters (BGCs) were found to be unique to VB1 compared to closely related strains: BGC #15, which encodes potentially novel anthracycline compounds with cancer cell growth inhibition properties, and BGC #28, which features a non-canonical configuration combining arylomycin, globomycin, and siamycin BGCs. This supercluster, the first described to consist of more than two adjacent and functional BGCs, co-produces at least three antimicrobial compounds from different antibiotic families. These findings highlight Streptomyces sp. VB1’s potential for discovering new bioactive molecules, positioning it as a promising candidate for further research. [ABSTRACT FROM AUTHOR]

Currently, clinically available cancer chemopreventive drug options are limited to mostly tamoxifen and its derivatives, such as raloxifene, and approved specifically for breast cancer. Thus, the availability of chemopreventive drug molecules for other types of malignant cancers would be desirable. In previous reports, the arils of Myristica fragrans (mace) have been found to exhibit cancer chemopreventive activity. Therefore, the purpose of the present study was to identify a natural product from this species with potential chemopreventive activity guided by chemoinformatic sample analysis via Global Natural Products Social (GNPS) molecular networking and molecular docking. The neolignan licarin A (1) was identified as a potential chemopreventive constituent, and subsequently submitted to several in vitro bioassays and a zebrafish toxicity evaluation. In this work, 1 afforded superior phosphoNF-κBp65 phosphorylation activity in DU-145 prostate cancer cells compared to isoliquiritigenin (2), which was used as a natural product chemopreventive control. Both 1 and 2 showed a longer-lasting reduction in cellular stress in a cell oxidative stress real-time dose–response assay than the positive control using Hepa1c1c7 mouse hepatoma cells. In addition, 1 displayed similar activities to 2, while also being less toxic to zebrafish (Danio rerio) than both this chalcone and the clinically used chemopreventive drug tamoxifen. [ABSTRACT FROM AUTHOR]

Ectopic pregnancy (EP) is the leading cause of maternal morbidity and mortality in the first trimester. Using an untargeted metabolomic approach, we sought to identify putative plasma biomarkers using tandem liquid chromatography–mass spectrometry for the detection of tubal EP. This case-control study included the prospective recruitment of 50 tubal EP cases and 50 early intrauterine pregnancy controls. To avoid over-fitting, logistic regression models were developed in a randomly selected discovery group (30 cases vs. 30 controls) and validated in the test group (20 cases vs. 20 controls). In total, 585 mass spectral features were detected, of which 221 molecular features were significantly altered in EP plasma (p < 0.05). Molecular networking and metabolite identification was employed using the Global Natural Products Social Molecular Networking (GNPS) database, which identified 97 metabolites at a high confidence level. Top significant metabolites include subclasses of sphingolipids, carnitines, glycerophosphocholines, and tryptophan metabolism. The top regression model, consisting of D-erythro-sphingosine and oleoyl-carnitine, was validated in a test group and achieved an area under receiving operating curve (AUC) (95% CI) = 0.962 (0.910–1) with a sensitivity of 100% and specificity of 95.9%. Metabolite alterations indicate alterations related to inflammation and abnormal placentation in EP. The validation of these metabolite biomarkers in the future could potentially result in improved early diagnosis. [ABSTRACT FROM AUTHOR]

The building blocks-based molecular network (BBMN) strategy was applied to the phytochemical investigation of Cleistocalyx operculatus , leading to the targeted isolation of eighteen novel cinnamoylphloroglucinol-terpene adducts (CPTAs) with diverse skeleton types (cleistoperones A–R, 1 – 18). Their structures including absolute configurations were determined by extensive spectroscopic methods, quantum chemical calculations, and single-crystal X-ray crystallographic experiments. Cleistoperone A (1), consisting of a cinnamoylphloroglucinol motif and two linear monoterpene moieties, represents an unprecedented macrocyclic CPTA, whose densely functionalized tricyclo[15.3.1.03,8]heneicosane bridge ring skeleton contains an enolizable β , β ′-triketone system and two different kinds of stereogenic elements (including five point and three planar chiralities). Cleistoperones B and C (2 and 3) are two new skeletal CPTAs with an unusual coupling pattern between the (nor)monoterpene moiety and the cinnamoyl chain of the cinnamoylphloroglucinol unit. Cleistoperone D (4) possesses an unprecedented cage-like 6/6/6/4/6-fused heteropentacyclic scaffold. The plausible biosynthetic pathways for 1 – 18 were also proposed. Notably, compounds 1 , 4 , 7 , 8 , and 18 exhibited significant antiviral activity against respiratory syncytial virus (RSV). The most potent one, cleistoperone A (1) with IC 50 value of 1.71 ± 0.61 μmol/L, could effectively inhibit virus replication via affecting the Akt/mTOR/p70S6K signaling pathway. The BBMN strategy led to the isolation of eighteen unprecedented cinnamoylphloroglucinol-terpene adducts from Cleistocalyx operculatus. Cleistoperone A (1) showed potent anti-RSV activity via affecting the Akt/mTOR/p70S6K signaling pathway. [Display omitted] [ABSTRACT FROM AUTHOR]

Background/Objectives: Recently, antimicrobial-resistant pathogens and cancers have emerged as serious global health problems, highlighting the immediate need for novel therapeutics. Consequently, we aimed to isolate and characterize endophytic Streptomyces strains from the rhizospheres of the Himalayan region of Nepal and identify specialized metabolites with antibacterial, antifungal, and cytotoxic potential. Methods: To isolate Streptomyces sp., we collected two soil samples and cultured them on an ISP4 medium after pretreatment. We isolated and identified the strains PY108 and PY109 using a combination of morphological observations and 16S rRNA gene sequencing. Results: The BLAST results showed that PY108 and PY109 resembled Streptomyces hundungensis PSB170 and Streptomyces sp. Ed-065 with 99.28% and 99.36% nucleotide similarity, respectively. Antibacterial assays of ethyl acetate (EA) extracts from both isolates PY108 and PY109 in a tryptic soy broth (TSB) medium were conducted against four pathogenic bacteria. They showed significant antibacterial potential against Staphylococcus aureus and Klebsiella pneumoniae. Similarly, these extracts exhibited moderate antifungal activities against Saccharomyces cerevisiae and Aspergillus niger. Cytotoxicity assays on cervical cancer cells (HeLa) and breast cancer cells (MCF-7) revealed significant potential for both extracts. LC-MS/MS profiling of the EA extracts identified 27 specialized metabolites, including diketopiperazine derivatives, aureolic acid derivatives such as chromomycin A, and lipopeptide derivatives. In comparison, GC-MS analysis detected 34 metabolites, including actinomycin D and γ-sitosterol. Furthermore, a global natural product social molecular networking (GNPS)-based molecular networking analysis dereplicated 24 metabolites in both extracts. Conclusions: These findings underscore the potential of endophytic Streptomyces sp. PY108 and PY109 to develop new therapeutics in the future. [ABSTRACT FROM AUTHOR]

Introduction: Compound annotation is always a challenging step in metabolomics studies. The molecular networking strategy has been developed recently to organize the relationship between compounds as a network based on their tandem mass (MS2) spectra similarity, which can be used to improve compound annotation in metabolomics analysis. Objective: This study used Bupleuri Radix from different geographic areas to evaluate the performance of molecular networking strategy for compound annotation in liquid chromatography‐mass spectrometry (LC–MS)–based metabolomics. Methodology: The Bupleuri Radix extract was analyzed by LC‐quadrupole time‐of‐flight MS under MSe acquisition mode. After raw data preprocessing, the resulting dataset was used for statistical analysis, including principal component analysis (PCA) and partial least squares discriminant analysis (PLS‐DA). The chemical makers related to the sample growth place were selected using variable importance in projection (VIP) > 2, fold change (FC) > 2, and p < 0.05. The molecular networking analysis was applied to conduct the compound annotation. Results: The score plots of PCA showed that the samples were classified into two clusters depending on their growth place. Then, the PLS‐DA model was constructed to explore the chemical changes of the samples further. Sixteen compounds were selected as chemical makers and tentatively annotated by the feature‐based molecular networking (FBMN) analysis. Conclusion: The results showed that the molecular networking method fully exploits the MS information and is a promising tool for facilitating compound annotation in metabolomics studies. However, the software used for feature extraction influenced the results of library searching and molecular network construction, which need to be taken into account in future studies. The molecular networking strategy could organize the relationship between compounds as a network based on their tandem mass spectra similarity, which can be used to improve compound annotation in metabolomics analysis. However, the software used for feature extraction influenced the results of library searching and molecular network construction, which need to be taken into account in future studies. [ABSTRACT FROM AUTHOR]

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• Metabolite screening was achieved by using standard qualitative tests. • Bioactive metabolites were separated and identified by De-replication approach, coupling HPLC-ESI-QTOF-MS with MZmine 3, global natural products social molecular networking (GNPS) and METLIN Gen2. • Microwave digestion was used to extract metals for inductively coupled plasma optical emission spectroscopy (ICP-OES) analysis. • Health risk assessment was achieved by determining estimated daily intake (EDI), hazard quotients (HQ), and carcinogenic risk (CR). The bark of Entandrophragma delevoyi De Wild (E. delevoyi) is widely used in the Zambian traditional health care system as a very effective medicinal plant. Nevertheless, little is known about the phytochemicals affiliated with the ethno-pharmacological uses of the plant. Further, the heavy metals safety of consuming the plant for medicinal purposes remains unknown. In this study, metabolite profiling of the methanolic stem bark extract of E. delevoyi was performed using a de-replication approach by coupling HPLC-ESI-QTOF-MS with MZmine 3 based two molecular networking approaches, that is, GNPS (global natural products social molecular networking) and METLIN Gen2. Varieties of metabolites were tentatively identified for the first time in E. delevoyi. Health risk assessment was achieved by analysing the presence and concentration of heavy metals such as Ag, Al, As, Cd, Co, Cr, Cu, Fe, Mn, Ni, Pb, Se, Si, Sn, U and Zn using inductively coupled plasma optical emission spectroscopy (ICP-OES). Further, estimated daily intake (EDI), hazard quotients (HQ), and carcinogenic risk were determined. The utilization of qualitative tests to screen metabolite classes and the molecular networking tools revealed a detailed phytochemical profile of this medicinal plant species. Qualitative metabolite screening tests revealed the presence of flavonoids, alkaloids, phenolics, saponins, anthraquinones, terpenoids, steroids, cardiac glycosides, anthocyanins, and tannins in the methanol extract. LCMS2 coupled to mzmine3.6 revealed the presence of thirty-five metabolites from MoNA database. In addition, LCMS2-GNPS revealed the presence of twenty-six metabolites, and LCMS-METLIN Gen2 revealed a further sixteen compounds. Furthermore, total polyphenolic content and antioxidant (1,1-diphenyl-2-picryl-hydrazylhydrate - DPPH) capacities were assayed. Total phenolic content was 169.46 ± 0.71 mg GAE/g and total flavonoid content was 53.14 ± 1.78 mg QE/g. The crude extract had significant total polyphenolic content and antioxidant potential (IC 50 = 48.91 µg/mL) compared to the standard, ascorbic acid (IC 50 = 74.47 µg/mL). This plant, full of bioactive compounds and significant antioxidant properties, offers unique health solutions and robust possibilities for future drug development and pharmacology research. Although the bark of E. delevoyi possesses many health-promoting bioactive metabolites, the heavy metals evaluated in the bark of E. delevoyi in this study have carcinogenic risk (Al, 0.096; Cd, 0.002; Cr, 1.271; Ni, 0.024). Based on the non-carcinogenic health risk evaluation, E. delevoyi bark may be safe. However, its safety can only be determined if the plant's health risks are fully described in extensive chronic toxicity tests. E. delevoyi should therefore be used with caution in medicinal preparations. [ABSTRACT FROM AUTHOR]

Lake Avernus is a volcanic lake located in southern Italy. Since ancient times, it has inspired numerous myths and legends due to the occurrence of singular phenomena, such as coloring events. Only recently has an explanation been found for them, i.e., the recurring color change over time is due to the alternation of cyanobacterial blooms that are a consequence of natural nutrient inputs as well as pollution resulting from human activities. This current report specifically describes the red coloring event that occurred on Lake Avernus in March 2022, the springtime season in this region of Italy. Our innovative multidisciplinary approach, the Fast Detection Strategy (FDS), was devised to monitor cyanobacterial blooms and their toxins. It integrates remote sensing data from satellites and drones, on-site sampling, and analytical/bioinformatics analyses into a cohesive information flow. Thanks to FDS, we determined that the red color was attributable to a bloom of Planktothrix rubescens, a toxin-producing cyanobacterium. Here, we report the detection and identification of 14 anabenopeptins from this P. rubescens strain, seven of which are known and seven are newly reported herein. Moreover, we explored the mechanisms and causes behind this cyclic phenomenon, confirming cyanobacterias role as reliable indicators of environmental changes. This investigation further validates FDSs effectiveness in detecting and characterizing cyanobacterial blooms and their associated toxins, expanding its potential applications.

Abstract Background Pisum sativum (PS) is a universal legume plant utilized for both human and animal consumption, particularly its seeds, known as green peas. The processing of PS in food industries and households produces a significant amount of waste that needs to be valorized. Methods In this study, the metabolite profiles of the 70% ethanolic extracts of PS wastes, namely peels (PSP) and a combination of leaves and stems (PSLS), were investigated by liquid chromatography-electrospray ionization-quadrupole time-of-flight tandem mass spectrometry (LC-ESI-QTOF-MS/MS) followed by molecular networking. Results Different classes of metabolites were identified, being flavonoids and their derivatives, along with phenolic acids, the most abundant categories. Additionally, a comprehensive network pharmacology strategy was applied to elucidate potentially active metabolites, key targets, and the pathways involved in cytotoxic activity against breast cancer. This cytotoxic activity was investigated in MCF-7 and MCF-10a cell lines. Results revealed that PSLS extract exhibited a potent cytotoxic activity with a good selectivity index (IC50 = 17.67 and selectivity index of 3.51), compared to the reference drug doxorubicin (IC50 = 2.69 µg/mL and selectivity index of 5.28). Whereas PSP extract appeared to be less potent and selective (IC50 = 32.92 µg/mL and selectivity index of 1.62). A similar performance was also observed for several polyphenolics isolated from the PSLS extract, including methyl cis p-coumarate, trans p-coumaric acid, and liquiritigenin/ 7-methyl liquiritigenin mixture. Methyl cis p-coumarate showed the most potent cytotoxic activity against MCF-7 cell line and the highest selectivity (IC50 = 1.18 µg/mL (6.91 µM) and selectivity index of 27.42). The network pharmacology study revealed that the isolated compounds could interact with several breast cancer-associated protein targets including carbonic anhydrases 1, 2, 4, 9, and 12, as well as aldo-keto reductase family 1 member B1, adenosine A3 receptor, protein tyrosine phosphatase non-receptor type 1, and estrogen receptor 2. Conclusion The uncovered therapeutic potential of PSLS and its metabolite constituents pave the way for an efficient and mindful PS waste valorization, calling for further in-vitro and in-vivo research. Graphical Abstract

The building blocks-based molecular network (BBMN) strategy was applied to the phytochemical investigation of Cleistocalyx operculatus, leading to the targeted isolation of eighteen novel cinnamoylphloroglucinol-terpene adducts (CPTAs) with diverse skeleton types (cleistoperones A–R, 1–18). Their structures including absolute configurations were determined by extensive spectroscopic methods, quantum chemical calculations, and single-crystal X-ray crystallographic experiments. Cleistoperone A (1), consisting of a cinnamoylphloroglucinol motif and two linear monoterpene moieties, represents an unprecedented macrocyclic CPTA, whose densely functionalized tricyclo[15.3.1.03,8]heneicosane bridge ring skeleton contains an enolizable β,β′-triketone system and two different kinds of stereogenic elements (including five point and three planar chiralities). Cleistoperones B and C (2 and 3) are two new skeletal CPTAs with an unusual coupling pattern between the (nor)monoterpene moiety and the cinnamoyl chain of the cinnamoylphloroglucinol unit. Cleistoperone D (4) possesses an unprecedented cage-like 6/6/6/4/6-fused heteropentacyclic scaffold. The plausible biosynthetic pathways for 1–18 were also proposed. Notably, compounds 1, 4, 7, 8, and 18 exhibited significant antiviral activity against respiratory syncytial virus (RSV). The most potent one, cleistoperone A (1) with IC50 value of 1.71 ± 0.61 μmol/L, could effectively inhibit virus replication via affecting the Akt/mTOR/p70S6K signaling pathway.

Abstract Several Pelargonium species are cultivated mainly to produce essential oils used in perfume industry and for ornamental purposes. Although the chemical composition and biological activities of their essential oils were extensively investigated, there is limited information about the chemical composition of their non-volatile constituents. In this study, we report an Ultra-Performance Liquid Chromatography-Mass Spectrometry (UPLC-MS)-based metabolomics approach for the annotation and analysis of various metabolites in three species; P. graveolens, P. denticulatum, and P. fragrans utilizing The Global Natural Product Social Molecular Networking (GNPS) and multivariate data analyses for clustering of the metabolites. A total of 154 metabolites belonging to different classes were annotated. The three species are good sources of coumarins, benzoic acid derivatives, organic acids, fatty acids, and phospholipids. However, the highest level of flavonols (mono- and di-O-glycosides) and cinnamic acid derivatives was found in P. graveolens and P. denticulatum, whereas tannins and flavone C-glycosides were abundant in P. fragrans. The metabolic profiles clarified here provide comprehensive information on the non-volatile constituents of the three Pelargonium species and can be employed for their authentication and possible therapeutic applications.

To investigate the metabolite profiles of typical Chinese brick dark tea and the relationship between the metabolite types of brick dark tea and fungi isolated from it, this study used non-targeted metabolomics based on liquid chromatography-mass spectrometry (LC-MS) and feature-based molecular networking (FBMN) for metabolomic analysis of typical brick dark tea in China (Hubei Qingzhuan brick tea, Sichuan Kangzhuan brick tea, dark brick tea, Huazhuan brick tea and Fuzhuan brick tea from Hunan, and Fuzhuan brick tea from Shaanxi), fresh tea leaves and various fungal strains isolated from dark tea. The results indicated that the non-volatile metabolite profiles of brick dark tea tended to be classified into two groups according to production regions: one group included brick dark tea from Hunan and Shaanxi, and the other group included brick dark tea from Sichuan and Hubei. The major factor determining the metabolite profile was the stalk percentage of brick tea, and the stalk percentage of brick dark tea from Sichuan and Hubei was significantly higher than that of the other group. The major reason for the difference in metabolite profile between the two groups was the higher contents of flavan-3-ols as well as most amino acids and their derivatives in brick dark tea from Hunan and Shaanxi. The molecular network analysis of the metabolites of brick dark tea and fungi showed that linear and cyclic dipeptides derived from fungi were widely distributed in brick dark tea, especially Kangzhuan brick tea. Isopentenyl diketopiperazine compounds of diverse structural types were found to be abundant in Fuzhuan brick tea, which were derived from Eurotium cristatum. Therefore, fungi contribute to the rich variety of metabolites in brick dark tea. This research can provide a theoretical reference for a comprehensive understanding and knowledge of the chemical basis of quality in typical brick dark tea in China.

Context Dayuanyin decoction is a traditional Chinese medicine formulation that is commonly used in modern clinical practice to treat viral infections such as viral pneumonia, viral fever, influenza, and hepatitis. Although the usage rate of Dayuanyin decoction is gradually increasing in clinical practice, its pharmacological constituents are still unclear.Objective This study comprehensively characterized the chemical constituents in Dayuanyin decoction using ultra-performance liquid chromatography quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS) and molecular networking.Materials and methods The overall strategy involved retrieving structural information, such as fragment ions and precursor ion masses, from self-built databases to identify the target constituents of the Dayuanyin decoction extract. The identification of non-targeted constituents was achieved by analyzing different categories, fragment pathways, mass spectrometry data, and the relationships between clusters of structures in molecular networking. Unannotated constituents were inferred from secondary mass spectrometry similarity and molecular weight differences and annotated constituents in the same constituent cluster. A few predicted constituents were selected and validated by comparing them to reference standards under identical mass spectrometry conditions.Results This study preliminarily identified 216 constituents, including flavonoids, amino acids, alkaloids, triterpenes, steroidal saponins, phenylpropanoids, and other constituents.Conclusions This integrated strategy using UPLC-QTOF-MS and molecular networking lays the foundation for clinical research on pharmacologically active substances in Dayuanyin decoction and could be popularized for the comprehensive profiling of chemical constituents of other traditional Chinese medicines.

ABSTRACTAdvanced mass spectrometry and molecular networking techniques have led to an increase in the discovery of novel metabolites from bacteria and fungi. However, a systematic approach to exploring the metabolite profiles of microalgae in response to stress has not been performed. Unlocking the chemical potential of microalgae could provide further biotechnology applications in nutraceutical, biofuel, and cosmetic industries. This study explored the changes in metabolite production of strains of the three microalgae Dunaliella primolecta, Nannochloropsis oculata, and Phaeodactylum tricornutum cultured under varying nitrate, NaCl, salinity and pH conditions. A total of 2284 metabolites were detected across all strains and conditions, with 49% of those metabolites specific to cultures grown under stress (i.e., not present in the control). From comparison with 33 libraries of mass spectral data, only five metabolites were identified, stressing the need for more open-access natural product databases specifically focused on microalgae.

Objective: The ancient classical Chinese formulation, Xie-Bai-San (XBS), was studied qualitatively and quantitatively, and its main components were characterized. Materials and Methods: The chemical compounds in XBS were identified using a combination of ultra-performance liquid chromatography-quadrupole-Orbitrap mass spectrometry and molecular networking (MN) analysis. In addition, a multiple reaction monitoring (MRM) quantification method was established using ultra-high-performance liquid chromatography–triple quadrupole mass spectrometry. Mulberroside A, kukoamine B, liquiritin, and glycyrrhizic acid were quantitatively analyzed in ten batches of XBS using this method. Results: Using clusters of MN and databases, 91 compounds were rapidly identified, including stilbene glycosides, saponins, flavonoids, and alkaloids. Of these, 30, 33, and 28 compounds were from Mori Cortex, Lycii Cortex, and Glycyrrhizae Radix et Rhizoma, respectively. The MRM scanning mode was used for the simultaneous sensitive quantitative determination of multiple components in a wide range of complex samples. The four main components were quantified using this method, which has good linearity. Conclusions: This study provides a comprehensive and reliable method for the identification and quantitative analysis of XBS.

Abstract Clematis graveolens Lindl., an indigenous climbing plant found in the Himalayan areas, is used by local communities for the treatment of neck tumors. The objective of this work is to examine the comprehensive metabolomic profile, antioxidant capability, in vitro and in silico anti-glioma effects on U-87 human glioma cell lines of the crude extract and fractions from C. graveolens. Liquid chromatography coupled with mass spectroscopy (LC–MS/MS) was used to establish detailed metabolite profiling of C. graveolens. The assessment of cell cytotoxicity was conducted using MTT cell viability assay on U-87 and BHK-21. Through molecular docking studies, the mode of inhibition and binding interaction between identified compounds and target proteins were also determined to evaluate the in vitro results. The use of LC–MS/MS-based global natural products social (GNPS) molecular networking analysis resulted in the identification of 27 compounds. The crude extract, ethyl acetate fraction, and chloroform fraction exhibited significant inhibitory activity against the U-87 cell lines, with IC50 values of 112.0, 138.1, and 142.7 µg/mL, respectively. The ethyl acetate fraction exhibited significant inhibitory concentration for 2,2’-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) activity, 2,2-diphenyl-1-picrylhydrazyl (DPPH) activity and the metal chelation activity with IC50 value of 39.50 µg/mL, 32.27 µg/mL, and 53.46 µg/mL, respectively. The crude extract showed maximum total phenolic, and total flavonoid concentration measuring 338.7 µg GAE/mg, and 177.04 µg QE/mg, respectively. The findings of this study indicate that C. graveolens consists of a diverse range of active phytoconstituents that possess antioxidant and anti-glioma properties.

Abstract Background A growing number of studies have demonstrated that the polar regions have the potential to be a significant repository of microbial resources and a potential source of active ingredients. Genome mining strategy plays a key role in the discovery of bioactive secondary metabolites (SMs) from microorganisms. This work highlighted deciphering the biosynthetic potential of an Arctic marine-derived strain Aspergillus sydowii MNP-2 by a combination of whole genome analysis and antiSMASH as well as feature-based molecular networking (MN) in the Global Natural Products Social Molecular Networking (GNPS). Results In this study, a high-quality whole genome sequence of an Arctic marine strain MNP-2, with a size of 34.9 Mb was successfully obtained. Its total number of genes predicted by BRAKER software was 13,218, and that of non-coding RNAs (rRNA, sRNA, snRNA, and tRNA) predicted by using INFERNAL software was 204. AntiSMASH results indicated that strain MNP-2 harbors 56 biosynthetic gene clusters (BGCs), including 18 NRPS/NRPS-like gene clusters, 10 PKS/PKS-like gene clusters, 8 terpene synthse gene clusters, 5 indole synthase gene clusters, 10 hybrid gene clusters, and 5 fungal-RiPP gene clusters. Metabolic analyses of strain MNP-2 grown on various media using GNPS networking revealed its great potential for the biosynthesis of bioactive SMs containing a variety of heterocyclic and bridge-ring structures. For example, compound G-8 exhibited a potent anti-HIV effect with an IC50 value of 7.2 nM and an EC50 value of 0.9 nM. Compound G-6 had excellent in vitro cytotoxicities against the K562, MCF-7, Hela, DU145, U1975, SGC-7901, A549, MOLT-4, and HL60 cell lines, with IC50 values ranging from 0.10 to 3.3 µM, and showed significant anti-viral (H1N1 and H3N2) activities with IC50 values of 15.9 and 30.0 µM, respectively. Conclusions These findings definitely improve our knowledge about the molecular biology of genus A. sydowii and would effectively unveil the biosynthetic potential of strain MNP-2 using genomics and metabolomics techniques.

Abstract The untargeted metabolomics-based molecular networking approach combined with multivariate analysis, proves to be an effective strategy for distinguishing raw materials in herbal medicine according to specific criteria. It exhibits the correlations between chemical constituents and the geographical habitats of plants, providing a valuable tool for ensuring quality control in mass production within the industry. In this study, we conducted a comprehensive investigation of the chemical compositions of Epimedium koreanum Nakai and performed comparative analyses on four extracts collected from distinct regions in South Korea using untargeted metabolomics tools. Through the comprehensive use of UPLC-QTOF/MS analysis and advanced statistical techniques, we elucidated the chemical composition, leading to the identification of key chemical markers. Additionally, the molecular networking analysis revealed distinct clusters of flavonoids and phenolic acids, highlighting the influence of regional factors on the metabolite profiles. These findings offer a promising avenue for enhancing quality control and traceability in the herbal medicine industry, underscoring the important role of geographical variation in the chemical profiles of herbal products.

Tribulus terrestris L. is a cosmopolitan annual herb that grows under various ecological conditions, reflecting the variation in its morphological, chemical, and biological features. T. terrestris was collected from three different populations (I, II, III) in Egypt and subjected to comparative investigation to evaluate the morphological, metabolomic, and genetic variations among these populations. Additionally, the antioxidant activity of the collected specimens was assessed. Significant variations in vegetative morphology were observed among the three populations. Detailed microscopic examination revealed unique traits in the foliar epidermal and pollen grains characteristics of each population. LC-ESI-MS/MS analysis of aqueous methanol extracts led to the tentative identification of 101 compounds across 17 different chemical classes from which flavonoids are predominant and represented by 28 compounds; Polymethoxylated flavones were reported for the first time in the three investigated extracts. GC/FID analysis of diethyl ether extracts identified 12 saturated and 8 unsaturated fatty acids, with linolenic acid being the major component, constituting 52.21 %, 65.71 %, and 49.52 % in populations I, II and III, respectively. The DNA fingerprinting showed variations with 54.54 % polymorphism in ISSR markers, confirming the existence of two distinct genotypes among the studied T. terrestris plants. Furthermore, the extracts from the three populations exhibited weak antioxidant activity against DPPH. The obtained data revealed considerable morphological, metabolomic, and genetic diversity among the three populations of T. terrestris. These findings suggest treating T. terrestris (I) as a distinct variety of T. terrestris and enhancing the understanding of the plant's diversity, which is essential for its effective application in traditional and modern medicinal practices. [Display omitted] • T. terrestris : morphological, chemical & molecular traits analyzed comprehensively. • LC-ESI-MS-MS with GNPS identified a new chemical class; polymethoxylated flavones. • DNA fingerprinting confirmed polymorphism indicating a distinct variety. [ABSTRACT FROM AUTHOR]

Gabon has a rich flora, many species of which are used in traditional medicine. However, little research has been carried out on this wealth. An ethnopharmacological survey in the Fang language was carried out among traditional practitioners to collect antimicrobial medicinal plants. Phytochemical profiling of ethanolic and methanolic extracts from Erismadelphus exsul Mildbr leaves was carried out using HPLC-ESI-Q/TOF and a molecular network approach. Antibacterial activity was assessed with disk diffusion and microdilution methods, antioxidant activity via DPPH and FRAP methods, and in vitro cell viability via Cell Counting Kit-8. A total of 21 medicinal plants were collected, grouped into 10 families, of which the Fabaceae is the most represented. Erismadelphus exsul was chosen for chemical and biological studies due to its citation frequency (RCF = 0.59) and the absence of previous phytochemical studies. These studies revealed 4 major families of natural compounds and annotated 19 compounds for the first time. The crude leaf extract showed significant antioxidant and antibacterial activity. Cytotoxicity studies showed that the leaves were not cytotoxic, unlike the bark. This study underlines the importance of preserving the ancestral knowledge of the Fang populations, while showing promising results for Erismadelphus exsul. [ABSTRACT FROM AUTHOR]

The present study is focused on evaluating acaricidal activity and chemical compositions of Erigeron acer root, which was identified as a promising candidate among fifteen Mongolian plant extracts tested for acaricidal activity. The acaricidal effect was evaluated against Haemaphysalis longicornis, assessed for toxicity to normal human skin fibroblast, and analyzed for its chemical constituents. The acetone extract of E. acer root showed significant activity against H. longicornis, with a lethal concentration (LC50) of 5.31 mg/mL and low toxicity, evidenced by a cytotoxic concentration (CC50) of 267.00 µg/mL. Using liquid chromatography-tandem mass spectrometry and molecular networking, thirteen natural compounds were identified, including pyrrolidines, alkaloids, fatty acids, and flavonoids, highlighting the efficacy of E. acer root extract as an effective acaricide against H. longicornis and offering insights for developing new tick control solutions. [ABSTRACT FROM AUTHOR]

Clematis graveolens Lindl., an indigenous climbing plant found in the Himalayan areas, is used by local communities for the treatment of neck tumors. The objective of this work is to examine the comprehensive metabolomic profile, antioxidant capability, in vitro and in silico anti-glioma effects on U-87 human glioma cell lines of the crude extract and fractions from C. graveolens. Liquid chromatography coupled with mass spectroscopy (LC–MS/MS) was used to establish detailed metabolite profiling of C. graveolens. The assessment of cell cytotoxicity was conducted using MTT cell viability assay on U-87 and BHK-21. Through molecular docking studies, the mode of inhibition and binding interaction between identified compounds and target proteins were also determined to evaluate the in vitro results. The use of LC–MS/MS-based global natural products social (GNPS) molecular networking analysis resulted in the identification of 27 compounds. The crude extract, ethyl acetate fraction, and chloroform fraction exhibited significant inhibitory activity against the U-87 cell lines, with IC50 values of 112.0, 138.1, and 142.7 µg/mL, respectively. The ethyl acetate fraction exhibited significant inhibitory concentration for 2,2'-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) activity, 2,2-diphenyl-1-picrylhydrazyl (DPPH) activity and the metal chelation activity with IC50 value of 39.50 µg/mL, 32.27 µg/mL, and 53.46 µg/mL, respectively. The crude extract showed maximum total phenolic, and total flavonoid concentration measuring 338.7 µg GAE/mg, and 177.04 µg QE/mg, respectively. The findings of this study indicate that C. graveolens consists of a diverse range of active phytoconstituents that possess antioxidant and anti-glioma properties. [ABSTRACT FROM AUTHOR]

Four prenylated indole alkaloids (1−4) were targeted isolated from the mangrove rhizosphere soil-derived fungus Penicillium janthinellum HK1-6 by using molecular networking strategies. Among them, the planar structure and relative configuration of notoamide X (1) were elucidated by detailed analysis of the spectroscopic data especially the NOESY spectrum for the first time and its absolute configuration was determined by ECD spectrum. Furthermore, curated molecular networks of MS/MS data were generated with GNPS which allowed highlighting six prenylated indole alkaloids (5, 6, 8, 9, 11, 12) that had not previously been identified in this fungus and two (7, 10) that had never been observed in any fungus. The MS/MS fragmentation pathway of these prenylated indole alkaloids was summarized. [ABSTRACT FROM AUTHOR]

Due to the emergence of drug-resistant microorganisms, the search for broad-spectrum antimicrobial compounds has become extremely crucial. Natural sources like plants and soils have been explored for diverse metabolites with antimicrobial properties. This study aimed to identify microorganisms from agricultural soils exhibiting antimicrobial effects against known human pathogens, and to highlight the chemical space of the responsible compounds through the computational metabolomics-based bioprospecting approach. Herein, bacteria were extracted from soil samples and their antimicrobial potential was measured via the agar well diffusion method. Methanolic extracts from the active bacteria were analyzed using the liquid chromatography quadrupole time-of-flight mass spectrometry (LC-QTOF-MS) technique, and the subsequent data was further analyzed through molecular networking approach which aided in identification of potential anti-microbial compounds. Furthermore, 16S rRNA gene sequencing enabled identification of the active bacterial isolates, where isolate 1 and 2 were identified as strains of Bacillus pumilus, whilst isolate 3 was found to be Bacillus subtilis. Interestingly, isolate 3 (Bacillus subtilis) displayed wide-ranging antimicrobial activity against the tested human pathogens. Molecular networking revealed the presence of Diketopiperazine compounds such as cyclo (D-Pro-D-Leu), cyclo (L-Tyr-L-Pro), cyclo (L-Pro-D-Phe), and cyclo (L-Pro-L-Val), alongside Surfactin C, Surfactin B, Pumilacidin E, and Isarrin D in the Bacillus strains as the main anti-microbial compounds. The application of the molecular networking approach represents an innovation in the field of bio-guided bioprospection of microorganisms and has proved to be an effective and feasible towards unearthing potent antimicrobial compounds. Additionally, the (computational metabolomics-based) approach accelerates the discovery of bioactive compounds and isolation of strains which offer a promising avenue for discovering new clinical antimicrobials. Finally, soil microbial flora could serve an alternative source of anti-microbial compounds which can assist in the fight against emergence of multi-drug resistance bacterial pathogens. [ABSTRACT FROM AUTHOR]

Background: A growing number of studies have demonstrated that the polar regions have the potential to be a significant repository of microbial resources and a potential source of active ingredients. Genome mining strategy plays a key role in the discovery of bioactive secondary metabolites (SMs) from microorganisms. This work highlighted deciphering the biosynthetic potential of an Arctic marine-derived strain Aspergillus sydowii MNP-2 by a combination of whole genome analysis and antiSMASH as well as feature-based molecular networking (MN) in the Global Natural Products Social Molecular Networking (GNPS). Results: In this study, a high-quality whole genome sequence of an Arctic marine strain MNP-2, with a size of 34.9 Mb was successfully obtained. Its total number of genes predicted by BRAKER software was 13,218, and that of non-coding RNAs (rRNA, sRNA, snRNA, and tRNA) predicted by using INFERNAL software was 204. AntiSMASH results indicated that strain MNP-2 harbors 56 biosynthetic gene clusters (BGCs), including 18 NRPS/NRPS-like gene clusters, 10 PKS/PKS-like gene clusters, 8 terpene synthse gene clusters, 5 indole synthase gene clusters, 10 hybrid gene clusters, and 5 fungal-RiPP gene clusters. Metabolic analyses of strain MNP-2 grown on various media using GNPS networking revealed its great potential for the biosynthesis of bioactive SMs containing a variety of heterocyclic and bridge-ring structures. For example, compound G-8 exhibited a potent anti-HIV effect with an IC50 value of 7.2 nM and an EC50 value of 0.9 nM. Compound G-6 had excellent in vitro cytotoxicities against the K562, MCF-7, Hela, DU145, U1975, SGC-7901, A549, MOLT-4, and HL60 cell lines, with IC50 values ranging from 0.10 to 3.3 µM, and showed significant anti-viral (H1N1 and H3N2) activities with IC50 values of 15.9 and 30.0 µM, respectively. Conclusions: These findings definitely improve our knowledge about the molecular biology of genus A. sydowii and would effectively unveil the biosynthetic potential of strain MNP-2 using genomics and metabolomics techniques. [ABSTRACT FROM AUTHOR]

The untargeted metabolomics-based molecular networking approach combined with multivariate analysis, proves to be an effective strategy for distinguishing raw materials in herbal medicine according to specific criteria. It exhibits the correlations between chemical constituents and the geographical habitats of plants, providing a valuable tool for ensuring quality control in mass production within the industry. In this study, we conducted a comprehensive investigation of the chemical compositions of Epimedium koreanum Nakai and performed comparative analyses on four extracts collected from distinct regions in South Korea using untargeted metabolomics tools. Through the comprehensive use of UPLC-QTOF/MS analysis and advanced statistical techniques, we elucidated the chemical composition, leading to the identification of key chemical markers. Additionally, the molecular networking analysis revealed distinct clusters of flavonoids and phenolic acids, highlighting the influence of regional factors on the metabolite profiles. These findings offer a promising avenue for enhancing quality control and traceability in the herbal medicine industry, underscoring the important role of geographical variation in the chemical profiles of herbal products. [ABSTRACT FROM AUTHOR]

Weeds are one of the biotic factors that cause crop productivity losses worldwide. Due to the consequences to human health and the environment of the indiscriminate use of synthetic herbicides, alternative methods involving the use of the allelopathy phenomenon have been gaining prominence. Here, we explore the allelopathic effect of Jatropha gossypiifolia L. on the weed Bidens bipinnata L. and investigate its potential herbicidal allelochemicals. In vitro bioassays demonstrated that the use of J. gossypiifolia leaf powder was able to inhibit seed germination and early growth of B. bipinnata seedlings, obtaining significant reductions with increasing concentration. Bioguided fractionation of the aqueous extract indicated that the hexane and ethyl acetate fractions were bioactive in inhibiting weed growth. Metabolomics based on mass spectrometry and molecular networks was used to annotate the allelochemicals of the bioactive fractions, generating the dereplication of metabolites from the classes of alkaloids, phenolics, fatty acids, steroids, and terpenoids, which may be associated with herbicidal activity. These results point to the allelopathic effect of the J. gossypiifolia leaf powder and its putative herbicide allelochemicals, providing subsidies for future studies on the application of this species in alternative weed management strategies. [ABSTRACT FROM AUTHOR]

Breonadia salicina (Vahl) Hepper & J.R.I. Wood is widely distributed throughout Africa. It is used ethnobotanically to treat various diseases. However, the metabolic profile of the Breonadia species is not well characterized and the metabolites that are responsible for the bioactivity of this plant remain unknown. Therefore, there is a need to determine the phytochemical and bioactivity profile to identify metabolites that contribute to the antidiabetic, anti-inflammatory and antiproliferation activity, including the genotoxicity and cytotoxic effects, of Breonadia salicina. The study is aimed at exploring the metabolomic profile antidiabetic, anti-inflammatory and antiproliferation activity, as well as the genotoxicity and cytotoxicity effects, of constituents of B. salicina. The compounds in the B. salicina extract were analyzed by ultra-performance liquid chromatography with quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS), and the resultant data were further analyzed using a molecular networking approach. The crude stem bark and root extracts showed the highest antidiabetic activity against α-amylase at the lowest test concentration of 62.5 µg/mL, with 74.53 ± 0.74% and 79.1 ± 1.5% inhibition, respectively. However, the crude stem bark and root extracts showed the highest antidiabetic activity against α-glucosidase at the lowest test concentration of 31.3 µg/mL, with 98.20 ± 0.15% and 97.98 ± 0.22% inhibition, respectively. The crude methanol leaf extract showed a decrease in the nitrite concentration at the highest concentration of 200 µg/mL, with cell viability of 90.34 ± 2.21%, thus showing anti-inflammatory activity. No samples showed significant cytotoxic effects at a concentration of 10 µg/mL against HeLa cells. Furthermore, a molecular network of Breonadia species using UPLC-QTOF-MS with negative mode electrospray ionization showed the presence of organic oxygen compounds, lipids, benzenoids, phenylpropanoids and polyketides. These compound classes were differentially distributed in the three different plant parts, indicating the chemical differences between the stem bark, root and leaf extracts of B. salicina. Therefore, the identified compounds may contribute to the antidiabetic and anti-inflammatory activity of Breonadia salicina. The stem bark, root and leaf extracts of B. salicina yielded thirteen compounds identified for the first time in this plant, offering a promising avenue for the discovery of new lead drugs for the treatment of diabetes and inflammation. The use of molecular networking produced a detailed phytochemical overview of this Breonadia species. The results reported in this study show the importance of searching for bioactive compounds from Breonadia salicina and provide new insights into the phytochemical characterization and bioactivity of different plant parts of Breonadia salicina. [ABSTRACT FROM AUTHOR]

Abstract Acinetobacter baumannii is without a doubt one of the most problematic bacteria causing hospital-acquired nosocomial infections in today's healthcare system. To solve the high prevalence of multi-drug resistant (MDR) in A. baumannii, we investigated one of the medicinal plants traditionally used as antibacterial agent; namely Murraya koenigii (L.) Sprengel. The total methanolic extracts of seeds and pericarps were prepared and their anti-bacterial activity was assessed using the agar diffusion method and minimum inhibitory concentration (MIC) was then calculated as compared to tigecycline. Then, an in-vivo murine model was established which confirmed the promising activity of M. koenigii seeds in demonstrating anti-bacterial and anti-inflammatory actions. The histopathological study of lungs, scoring of pulmonary lesions, counting of bacterial loads after infection by multi-drug resistant A. baumannii all provided evidence to support these findings. LC–MS/MS profiling coupled to molecular networking and chemometrics detected the presence of carbazole alkaloids, and coumarins as dominate metabolites of the active seed extracts. Positively correlated metabolites to antibacterial potential were 6-(2ʹ,3ʹ-dihydroxy-3-methylbutyl)-8-prenylumbelliferone, scopoline, and 5-methoxymurrayatin. An in-silico study was also performed on the crystal structure of MurF from A. baumannii (PDB ID: 4QF5), the studied structures of the mentioned extracts revealed good docking interaction at the active site suggestive of competition with the ATP ligand. These collective findings suggest that extracts of Murraya koenigii (L.) Sprengel seed is a novel prospective for the discovery of drug candidates against infections caused by MDR A. baumannii.

The past decades have marked the rise of metabolomics and lipidomics as the -omics sciences which reflect the most phenotypes in living systems. Mass spectrometry–based approaches are acknowledged for both quantification and identification of molecular signatures, the latter relying primarily on fragmentation spectra interpretation. However, the high structural diversity of biological small molecules poses a considerable challenge in compound annotation. Feature-based molecular networking (FBMN) combined with database searches currently sets the gold standard for annotation of large datasets. Nevertheless, FBMN is usually based on collision-induced dissociation (CID) data, which may lead to unsatisfying information. The use of alternative fragmentation methods, such as electron-activated dissociation (EAD), is undergoing a re-evaluation for the annotation of small molecules, as it gives access to additional fragmentation routes. In this study, we apply the performances of data-dependent acquisition mass spectrometry (DDA-MS) under CID and EAD fragmentation along with FBMN construction, to perform extensive compound annotation in the crude extracts of the freshwater sentinel organism Gammarus fossarum. We discuss the analytical aspects of the use of the two fragmentation modes, perform a general comparison of the information delivered, and compare the CID and EAD fragmentation pathways for specific classes of compounds, including previously unstudied species. In addition, we discuss the potential use of FBMN constructed with EAD fragmentation spectra to improve lipid annotation, compared to the classic CID-based networks. Our approach has enabled higher confidence annotations and finer structure characterization of 823 features, including both metabolites and lipids detected in G. fossarum extracts. [ABSTRACT FROM AUTHOR]

N-Benzylphenethylamine derivatives are 5-HT2A receptor agonists with hallucinogenic properties, including NBOMe (N-(2-methoxybenzyl)-2-(3,4,5-trimethoxyphenyl)ethan-1-amine) and NBOH (2-(((2,5-dimethoxyphenethyl)amino)methyl)phenol). We reported here the case of a 23-year-old man who presented a serotoninergic syndrome and a loss of consciousness following the consumption of a powder labelled as 25I-NBOH. Toxicological analyses of biological samples were carried out using a liquid chromatography high-resolution mass spectrometry. Two new psychoactive substances were identified and confirmed with certified reference materials: 25E-NBOH (2-(((4-ethyl-2,5-dimethoxyphenethyl)amino)methyl)phenol) and MDPHP (1-(benzo[d][1,3]dioxol-5-yl)-2-(pyrrolidin-1-yl)hexan-1-one). Pharmaceuticals administered to the patient during his medical care were found in plasma and urine. 25E-NBOH and MDPHP concentrations were respectively at 2.3 ng/mL and 3.4 ng/mL in plasma, and 25.7 ng/mL and 30.5 ng/mL in urine. 25I-NBOH (2-(((4-iodo-2,5-dimethoxyphenethyl)amino)methyl)phenol) was specifically searched in both samples and was not detected. These results are discussed along with a literature review on human cases of exposure to N-benzylphenethylamine derivatives. Using molecular networking approach, we propose the first 25E-NBOH metabolism study using authentic biological samples (plasma and urine). We described seven metabolites (M1 to M7), including two phase I (m/z 330.172; m/z 288.160) and five phase II metabolites (m/z 464.191, m/z 478.207, m/z 492.223, m/z 508.218; m/z 396.156). The M6 (m/z 492.223) was the most intense ion detected in plasma and urine and could be proposed as a relevant 25E-NBOH consumption marker. Overall, we described an original case of 25E-NBOH poisoning and identified metabolites that could potentially be used as consumption markers to detect 25E-NBOH intoxications with a higher confidence level and probably a longer detection window. [ABSTRACT FROM AUTHOR]

The Streptomyces strain G222, isolated from a Vietnamese marine sediment, was confidently identified by 16S rRNA gene sequencing. Its AcOEt crude extract was successfully analyzed using non-targeted LC-MS/MS analysis, and molecular networking, leading to a putative annotation of its chemical diversity thanks to spectral libraries from GNPS and in silico metabolite structure prediction obtained from SIRIUS combined with the bioinformatics tool conCISE (Consensus Annotation Propagation of in silico Elucidations). This dereplication strategy allowed the identification of an interesting cluster of a series of putative cyclic and linear lipopeptides of the lichenysin and surfactin families. Lichenysins (3–7) were isolated from the sub-fraction, which showed significant anti-biofilm activity against Pseudomonas aeruginosa MUC-N1. Their structures were confirmed by detailed 1D and 2D NMR spectroscopy (COSY, HSQC, HMBC, TOCSY, ROESY) recorded in CD3OH, and their absolute configurations were determined using the modified Marfey's method. The isolated lichenysins showed anti-biofilm activity at a minimum concentration of 100 µM. When evaluated for antibacterial activity against a panel of Gram-positive and Gram-negative strains, two isolated lichenysins exhibited selective activity against the MRSA strain without affecting its growth curve and without membranotropic activity. This study highlights the power of the MS/MS spectral similarity strategy using computational methods to obtain a cross-validation of the annotated molecules from the complex metabolic profile of a marine sediment-derived Streptomyces extract. This work provides the first report from a Streptomyces strain of combined cyclic and linear lichenysins and surfactins, known to be characteristic compounds of the genus Bacillus. [ABSTRACT FROM AUTHOR]

Due to dengue fever's swift global expansion and the lack of effective antiviral remedies, it is crucial to discover and develop new antiviral drugs. The study aimed to assess the antiviral potential of the extract and fractions from the leaves of Goniothalamus lanceolatus Miq. (Annonaceae). New Guinea C strain DENV-2, at a multiplicity of infection of 0.4, was used to infect Vero cells. The assessment of antiviral effectiveness was conducted through the plaque reduction assay. Deep metabolome analysis of the active fraction identified nine styryllactones based on reference standards and eighteen styryllactones were further annotated through a molecular database search. One new 2 H -tetrahydropyran derivative, 3- epi -goniothalesdiol A (19), together with seven known styryllactones (8 , 11 , 14 , 16 , 17 , 18 , and 22) were isolated from the leaves of G. lanceolatus. The active bis-styryllactone goniolanceolatin A (22) with the highest selectivity index (SI) underwent further evaluation using quantitative reverse transcription qRT-PCR to determine the viral RNA level. The qRT-PCR data showed that the IC 50 value for the active compound was 5.07 µg/mL, and its corresponding SI value was 5.30. Following comprehensive docking studies, the bis-styryllactone derivative 22 showed potential binding interactions with crucial amino acids of the Envelope (E) of DENV proteins comparable to those of ribavirin. [Display omitted] • In vitro evaluation of Goniothalamus lanceolatus leaf dichloromethane (GLLD) extract showed the strongest anti-dengue activity. • One new 2H-tetrahydropyran derivative was successfully isolated from the leaves of G. lanceolatus during a bioactivity-directed. • In vitro and in silico evaluation of bis-styryl lactone isolated from the most active fraction of leaves of G. lanceolatus showed the most potential anti-dengue activity. [ABSTRACT FROM AUTHOR]

Marigold oleoresin is an oil-soluble natural colorant mainly extracted from marigold flowers. Xinjiang of China, India, and Zambia of Africa are the three main production areas of marigold flowers. Therefore, this study utilized ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS/MS) technology, combined with Global Natural Products Social Molecular Networking (GNPS) and multivariate statistical analysis, for the qualitative and discriminant analysis of marigold oleoresin obtained from three different regions. Firstly, 83 compounds were identified in these marigold oleoresin samples. Furthermore, the results of a principal component analysis (PCA) and orthogonal partial least squares discriminant analysis (OPLS-DA) indicated significant differences in the chemical compositions of the marigold oleoresin samples from different regions. Finally, 12, 23, and 38 differential metabolites were, respectively, identified by comparing the marigold oleoresin from Africa with Xinjiang, Africa with India, and Xinjiang with India. In summary, these results can be used to distinguish marigold oleoresin samples from different regions, laying a solid foundation for further quality control and providing a theoretical basis for assessing its safety and nutritional aspects. [ABSTRACT FROM AUTHOR]

Acinetobacterbaumannii is without a doubt one of the most problematic bacteria causing hospital-acquired nosocomial infections in today's healthcare system. To solve the high prevalence of multi-drug resistant (MDR) in A.baumannii, we investigated one of the medicinal plants traditionally used as antibacterial agent; namely Murrayakoenigii (L.) Sprengel. The total methanolic extracts of seeds and pericarps were prepared and their anti-bacterial activity was assessed using the agar diffusion method and minimum inhibitory concentration (MIC) was then calculated as compared to tigecycline. Then, an in-vivo murine model was established which confirmed the promising activity of M.koenigii seeds in demonstrating anti-bacterial and anti-inflammatory actions. The histopathological study of lungs, scoring of pulmonary lesions, counting of bacterial loads after infection by multi-drug resistant A.baumannii all provided evidence to support these findings. LC–MS/MS profiling coupled to molecular networking and chemometrics detected the presence of carbazole alkaloids, and coumarins as dominate metabolites of the active seed extracts. Positively correlated metabolites to antibacterial potential were 6-(2ʹ,3ʹ-dihydroxy-3-methylbutyl)-8-prenylumbelliferone, scopoline, and 5-methoxymurrayatin. An in-silico study was also performed on the crystal structure of MurF from A.baumannii (PDB ID: 4QF5), the studied structures of the mentioned extracts revealed good docking interaction at the active site suggestive of competition with the ATP ligand. These collective findings suggest that extracts of Murrayakoenigii (L.) Sprengel seed is a novel prospective for the discovery of drug candidates against infections caused by MDR A.baumannii. [ABSTRACT FROM AUTHOR]

Xuetong, the dried stem of Kadsura heteroclita (Roxb.) Craib, is a traditional Tujia medicine extensively used to treat rheumatoid arthritis (RA). All traditional Chinese medicines (TCMs) necessitate a processing stage called "Paozhi" before clinical application. However, there is a dearth of research concerning the processing methods employed for Xuetong. To investigate the impact of vinegar and wine processing on the chemical constituents of Xuetong, this study devised a targeted offline two-dimensional (2D) high-performance liquid chromatography (HPLC) and ultra-high-performance liquid chromatography-orbitrap mass spectrometry (UHPLC-orbitrap-MS) method. By incorporating various MS data-processing techniques, such as molecular networking technology, fragment-ion similarity searching (FISh), online and offline database matching, and fragmentation pattern analysis, a total of 158 components were identified in Xuetong. Among them, 14 were verified by comparison with the reference standards. Notably, aside from triterpenoids and lignans, catechin derivatives were found to be the predominant constituents of Xuetong, and their levels exhibited a significant decrease following processing. This method significantly improved peak capacity and resolution, overcoming the limitations of 1D LC in simultaneously analyzing highly polar catechin derivatives and less polar triterpenoids and lignans. Moreover, the developed method shows promise for Xuetong's quality control. [ABSTRACT FROM AUTHOR]

Due to the emergence of drug-resistant microorganisms, the search for broad-spectrum antimicrobial compounds has become extremely crucial. Natural sources like plants and soils have been explored for diverse metabolites with antimicrobial properties. This study aimed to identify microorganisms from agricultural soils exhibiting antimicrobial effects against known human pathogens, and to highlight the chemical space of the responsible compounds through the computational metabolomics-based bioprospecting approach. Herein, bacteria were extracted from soil samples and their antimicrobial potential was measured via the agar well diffusion method. Methanolic extracts from the active bacteria were analyzed using the liquid chromatography quadrupole time-of-flight mass spectrometry (LC-QTOF-MS) technique, and the subsequent data was further analyzed through molecular networking approach which aided in identification of potential anti-microbial compounds. Furthermore, 16S rRNA gene sequencing enabled identification of the active bacterial isolates, where isolate 1 and 2 were identified as strains of Bacillus pumilus, whilst isolate 3 was found to be Bacillus subtilis. Interestingly, isolate 3 (Bacillus subtilis) displayed wide-ranging antimicrobial activity against the tested human pathogens. Molecular networking revealed the presence of Diketopiperazine compounds such as cyclo (D-Pro-D-Leu), cyclo (L-Tyr-L-Pro), cyclo (L-Pro-D-Phe), and cyclo (L-Pro-L-Val), alongside Surfactin C, Surfactin B, Pumilacidin E, and Isarrin D in the Bacillus strains as the main anti-microbial compounds. The application of the molecular networking approach represents an innovation in the field of bio-guided bioprospection of microorganisms and has proved to be an effective and feasible towards unearthing potent antimicrobial compounds. Additionally, the (computational metabolomics-based) approach accelerates the discovery of bioactive compounds and isolation of strains which offer a promising avenue for discovering new clinical antimicrobials. Finally, soil microbial flora could serve an alternative source of anti-microbial compounds which can assist in the fight against emergence of multi-drug resistance bacterial pathogens.

Diseases, especially degenerative illnesses, can be treated and avoided altogether, and good health can be enhanced by including health-promoting foods in the diet without side effects. Adenia panduriformis (Passifloraceae) is a wild vegetable consumed as part of the everyday diet in some communities in Zambia. The plant is also used in traditional medicine to effectively treat and manage many diseases, including degenerative diseases. Because of significant health benefits and medicinal attributes, this vegetable has very high merit to be investigated for its phytochemicals and bioactivities. Therefore, this study screened phytochemicals using qualitative chemical tests and Liquid chromatography-tandem mass spectrometry (LCMS/MS), identified metabolites using molecular networking tools and evaluated the total polyphenolic content and antioxidant activity of the methanolic leaf extract of the vegetable. Solvent extraction by maceration recovered metabolites for screening by qualitative chemical tests, extraction by QuEChERS (quick, easy, cheap, effective, rugged, and safe) was used to recover metabolites for screening by LCMS/MS, and tentative phytochemical constituents were identified using MZmine3, GNPS (global natural products social molecular networking) and METLIN. Total phenolic content (TPC) determination used the Folin-Ciocalteu (FC) method at 765 nm. Total flavonoid content (TFC) was evaluated using the aluminium chloride colorimetric assay at 510 nm. Antioxidant activity was evaluated using 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical scavenging assay at 517 nm. Phytochemical screening by qualitative chemical tests revealed the presence of many classes of phytocompounds, such as alkaloids, polyphenols, terpenoids, and saponins. TPC was determined to be 122.30 ± 2.00 mg GAE/g, and TFC was 60.17 ± 0.25 mg QE/g. Screening by LCMS/MS revealed the presence of many metabolites, of which 23 phytocompounds were identified using MZmine3, 42 by GNPS, and 21 by METLIN. The crude extract exhibited high DPPH free radical quenching activity with a relatively low IC50 value of 53.48 µg/mL compared to the reference standard ascorbic acid IC50 of 74.47 µg/mL. The significantly lower IC50 value for the wild vegetable than that of ascorbic acid indicates a considerably high efficiency of the metabolites in the vegetable as antioxidants. The presence of many metabolites with health-promoting properties and high antioxidant activity of the leaf extract of A. panduriformis may be the science behind its remarkable health benefits and power to treat and manage a variety of diseases, including degenerative illnesses. This study presents results that indicate the potential of A. panduriformis as a rich source of metabolites with brilliant health-promoting properties and high antioxidant activity, which may open exciting opportunities for further development in the fields of pharmacology and drug development.

Abstract Background Filamentous fungi are prolific producers of bioactive molecules and enzymes with important applications in industry. Yet, the vast majority of fungal species remain undiscovered or uncharacterized. Here we focus our attention to a wild fungal isolate that we identified as Anthostomella pinea. The fungus belongs to a complex polyphyletic genus in the family of Xylariaceae, which is known to comprise endophytic and pathogenic fungi that produce a plethora of interesting secondary metabolites. Despite that, Anthostomella is largely understudied and only two species have been fully sequenced and characterized at a genomic level. Results In this work, we used long-read sequencing to obtain the complete 53.7 Mb genome sequence including the full mitochondrial DNA. We performed extensive structural and functional annotation of coding sequences, including genes encoding enzymes with potential applications in biotechnology. Among others, we found that the genome of A. pinea encodes 91 biosynthetic gene clusters, more than 600 CAZymes, and 164 P450s. Furthermore, untargeted metabolomics and molecular networking analysis of the cultivation extracts revealed a rich secondary metabolism, and in particular an abundance of sesquiterpenoids and sesquiterpene lactones. We also identified the polyketide antibiotic xanthoepocin, to which we attribute the anti–Gram-positive effect of the extracts that we observed in antibacterial plate assays. Conclusions Taken together, our results provide a first glimpse into the potential of Anthstomella pinea to provide new bioactive molecules and biocatalysts and will facilitate future research into these valuable metabolites.

Currently, clinically available cancer chemopreventive drug options are limited to mostly tamoxifen and its derivatives, such as raloxifene, and approved specifically for breast cancer. Thus, the availability of chemopreventive drug molecules for other types of malignant cancers would be desirable. In previous reports, the arils of Myristica fragrans (mace) have been found to exhibit cancer chemopreventive activity. Therefore, the purpose of the present study was to identify a natural product from this species with potential chemopreventive activity guided by chemoinformatic sample analysis via Global Natural Products Social (GNPS) molecular networking and molecular docking. The neolignan licarin A (1) was identified as a potential chemopreventive constituent, and subsequently submitted to several in vitro bioassays and a zebrafish toxicity evaluation. In this work, 1 afforded superior phosphoNF-κBp65 phosphorylation activity in DU-145 prostate cancer cells compared to isoliquiritigenin (2), which was used as a natural product chemopreventive control. Both 1 and 2 showed a longer-lasting reduction in cellular stress in a cell oxidative stress real-time dose–response assay than the positive control using Hepa1c1c7 mouse hepatoma cells. In addition, 1 displayed similar activities to 2, while also being less toxic to zebrafish (Danio rerio) than both this chalcone and the clinically used chemopreventive drug tamoxifen.

Background/Objectives: Recently, antimicrobial-resistant pathogens and cancers have emerged as serious global health problems, highlighting the immediate need for novel therapeutics. Consequently, we aimed to isolate and characterize endophytic Streptomyces strains from the rhizospheres of the Himalayan region of Nepal and identify specialized metabolites with antibacterial, antifungal, and cytotoxic potential. Methods: To isolate Streptomyces sp., we collected two soil samples and cultured them on an ISP4 medium after pretreatment. We isolated and identified the strains PY108 and PY109 using a combination of morphological observations and 16S rRNA gene sequencing. Results: The BLAST results showed that PY108 and PY109 resembled Streptomyces hundungensis PSB170 and Streptomyces sp. Ed-065 with 99.28% and 99.36% nucleotide similarity, respectively. Antibacterial assays of ethyl acetate (EA) extracts from both isolates PY108 and PY109 in a tryptic soy broth (TSB) medium were conducted against four pathogenic bacteria. They showed significant antibacterial potential against Staphylococcus aureus and Klebsiella pneumoniae. Similarly, these extracts exhibited moderate antifungal activities against Saccharomyces cerevisiae and Aspergillus niger. Cytotoxicity assays on cervical cancer cells (HeLa) and breast cancer cells (MCF-7) revealed significant potential for both extracts. LC-MS/MS profiling of the EA extracts identified 27 specialized metabolites, including diketopiperazine derivatives, aureolic acid derivatives such as chromomycin A, and lipopeptide derivatives. In comparison, GC-MS analysis detected 34 metabolites, including actinomycin D and γ-sitosterol. Furthermore, a global natural product social molecular networking (GNPS)-based molecular networking analysis dereplicated 24 metabolites in both extracts. Conclusions: These findings underscore the potential of endophytic Streptomyces sp. PY108 and PY109 to develop new therapeutics in the future.

Ectopic pregnancy (EP) is the leading cause of maternal morbidity and mortality in the first trimester. Using an untargeted metabolomic approach, we sought to identify putative plasma biomarkers using tandem liquid chromatography–mass spectrometry for the detection of tubal EP. This case-control study included the prospective recruitment of 50 tubal EP cases and 50 early intrauterine pregnancy controls. To avoid over-fitting, logistic regression models were developed in a randomly selected discovery group (30 cases vs. 30 controls) and validated in the test group (20 cases vs. 20 controls). In total, 585 mass spectral features were detected, of which 221 molecular features were significantly altered in EP plasma (p < 0.05). Molecular networking and metabolite identification was employed using the Global Natural Products Social Molecular Networking (GNPS) database, which identified 97 metabolites at a high confidence level. Top significant metabolites include subclasses of sphingolipids, carnitines, glycerophosphocholines, and tryptophan metabolism. The top regression model, consisting of D-erythro-sphingosine and oleoyl-carnitine, was validated in a test group and achieved an area under receiving operating curve (AUC) (95% CI) = 0.962 (0.910–1) with a sensitivity of 100% and specificity of 95.9%. Metabolite alterations indicate alterations related to inflammation and abnormal placentation in EP. The validation of these metabolite biomarkers in the future could potentially result in improved early diagnosis.

This study investigated the potential benefits of the administration of red ginseng (RG) on lipid metabolism and the profiles of individual free fatty acids (FFAs) in healthy horses. Eight healthy horses, raised under similar conditions, were randomly divided into two groups, each comprising four horses. The experimental group received powdered RG (600 mg/kg/day) mixed with a carrier, and the control group received only the carrier. The parameters associated with lipid metabolism and probable adverse effects were evaluated in both groups after 3 weeks. The computational molecular networking (MN) approach was applied to analyze the FFA profiles. The results indicated that RG administration significantly reduced blood triglyceride levels in the experimental group. Analysis of the FFAs using MN revealed significant decreases in specific types of FFAs (C12:0, dodecanoic acid; C14:0, myristric acid; C18:1, oleic acid; C18:2, linoleic acid). RG consumption did not produce significant adverse effects on the renal, hepatic, and immune functions. Thus, RG was found to effectively modulate lipid metabolism and the levels of individual FFAs. The application of the MN for the analysis of FFAs represents a novel approach and can be considered for future research. [ABSTRACT FROM AUTHOR]

The comprehensive and efficient characterization of components in traditional Chinese medicine is crucial for elucidating its active constituents and uncovering its mechanism. Identifying the compounds of the Bushen Huoxue Prescription (BHP) is difficult because of its complex composition and the large difference in concentration among its compounds. In this study, a hydrophilic interaction liquid chromatography coupled with reversed‐phase LC (HILIC × RPLC) offline 2D‐LC tandem high‐resolution mass spectrometry method was established to analyze the total compounds of the BHP. Database screening and molecular networking were performed to identify the compounds. In contrast to conventional 1D chromatography, 2D chromatography increased peak capacity, enriched trace ingredients, and prevented the masking of high‐abundance compounds. A total of 165 compounds were identified, and 14 potential compounds needed to be further identified. This study provided an effective method for comprehensively analyzing the complex system of traditional Chinese medicine compounds. [ABSTRACT FROM AUTHOR]

Marine bacteria living in association with marine sponges have proven to be a reliable source of biologically active secondary metabolites. However, no studies have yet reported natural products from Microbacterium testaceum spp. We herein report the isolation of a M. testaceum strain from the sponge Tedania brasiliensis. Molecular networking analysis of bioactive pre-fractionated extracts from culture media of M. testaceum enabled the discovery of testacosides A–D. Analysis of spectroscopic data and chemical derivatizations allowed the identification of testacosides A–D as glycoglycerolipids bearing a 1-[α-glucopyranosyl-(1 → 3)-(α-mannopyranosyl)]-glycerol moiety connected to 12-methyltetradecanoic acid for testacoside A (1), 14-methylpentadecanoic acid for testacoside B (2), and 14-methylhexadecanoic acid for testacosides C (3) and D (4). The absolute configuration of the monosaccharide residues was determined by 1H-NMR analysis of the respective diastereomeric thiazolidine derivatives. This is the first report of natural products isolated from cultures of M. testaceum. Key points: • The first report of metabolites produced by Microbacterium testaceum. • 1-[α-Glucopyranosyl-(1 → 3)-(α-mannopyranosyl)]-glycerol lipids isolated and identified. • Microbacterium testaceum strain isolated from the sponge Tedania brasiliensis. [ABSTRACT FROM AUTHOR]

Eutylone is a cathinone-derived synthetic amphetamine scheduled by the World Health Organization and European Monitoring Centre for Drugs and Drug Addiction since 2022 due to its growing consumption. We report here an eutylone intoxication involving a 38-year-old man and a 29-year-old woman in a chemsex context. A bag containing a white crystalline powder labelled as a research product was found in their vehicle. Nuclear magnetic resonance and liquid chromatography-high-resolution mass spectrometry (LC-HRMS) analyses identified the powder as eutylone and confirmed purity superior to 99%. LC-HRMS data analysis using molecular networking allowed to propose new eutylone metabolites in blood samples in a graphical manner. We described 16 phase I (e.g. hydroxylated or demethylated) and phase II metabolites (glucuroconjugates and sulfoconjugates). The same metabolites were found both in male and female blood samples. Toxicological analyses measured eutylone concentration in blood samples at 1374 ng/mL and 1536 ng/mL for the man and the woman, respectively. A keto-reduced metabolite (m/z 238.144) was synthesized to permit its quantification at 67 ng/mL and 54 ng/mL in male and female blood samples, respectively. Overall, the identification of these metabolites will increase the knowledge of potential drug consumption markers and allow to implement mass spectrometry databases to better monitor future drug abuse or consumption. [ABSTRACT FROM AUTHOR]