The current work represents a comparative study of the phenolic profiles of three under-explored Onosma (Boraginaceae) species from Greece—Onosma leptantha (OL), Onosma erecta (OE), and Onosma graeca (OG). Although Onosma spp. have ethnopharmacological significance, previous phytochemical studies have focused primarily on roots. Methanolic extracts of the aerial parts were analyzed using qualitative LC-MS enhanced by molecular networking-based dereplication, annotating 94 phenolics categorized into hydroxybenzoic acids (7), hydroxycinnamic acids (24), lignans (14), neolignans (14), stilbenes (4), coumarins (5), and flavonoids (26). OG exhibited the broadest distribution of flavonoid glycosides. OL contained the greatest number of hydroxycinnamic and neolignan derivatives, and OE was notably abundant in lignans. Total phenolic (TPC) and total flavonoid (TFC) contents were quantified, and the antioxidant capacity and enzyme inhibition against cholinesterases, α-amylase, and α-glucosidase were assessed. OL showed a high TPC (69.03 mg GAE/g extract) and strong antioxidant activity, while OG exhibited a high TFC (45.80 mg RE/g extract). All extracts demonstrated stronger AChE inhibition than BChE, with OG showing the highest AChE inhibition (2.35 mg GALAE/g). Additionally, OL was the most active against both α-glucosidase (5.69 mmol ACAE/g) and α-amylase (0.48 mmol ACAE/g). This study improved our understanding of the chemical diversity within these species, providing a more comprehensive insight into their longstanding ethnopharmacological potential.

(1) Background: Species of the anamorphic genus Cladobotryum, are known for their fungicolous lifestyle, making them important mycopathogens in fungiculture. Many morphological, ecological, and molecular phylogenetic studies of the genus have been done to date, but taxonomic uncertainties and challenges still remain. Fungal secondary metabolites, being vastly diverse, are utilised as an extra tool in fungal systematics. Despite being studied for their potentially bioactive compounds, Cladobotryum species are insufficiently investigated regarding metabolomics. (2) Methods: The aim of this study is the identification of Greek strains of Cladobotryum by integrating morphological data, ITS-based phylogeny, and ¹H NMR-based metabolomics into a polyphasic approach. (3) Results: Twenty-three strains, isolated from sporophores of macromycetes inhabiting diverse Greek ecosystems, were morphologically identified as Cladobotryum apiculatum, C. fungicola, C. mycophilum, C. varium, C. verticillatum, and Hypomyces rosellus (anamorph C. dendroides), whereas seven strains, which produced red-pigmented metabolites, presented an ambiguous taxonomic position at the species level. Molecular phylogenetics and metabolomics corroborated the morphological findings. (4) Conclusions: Thorough morphological study, ITS region-based phylogeny, and NMR-based metabolomics contribute complementarily to resolving the genus Cladobotryum systematics.

Actinobacteria are a rich secondary metabolite source, accounting for nearly half of known bioactive microbial compounds, thus representing promising targets for novel bioactive molecule discovery. To explore potential antiaging compounds, we screened extracts from 980 actinobacterial strains isolated from diverse Greek ecosystems. Extracts were evaluated for elastase and tyrosinase inhibition in vitro, followed by toxicity and efficacy assessments in human cell lines. One Amycolatopsis and two Streptomyces strains exhibited significant tyrosinase inhibition, and one showed elastase inhibition, prompting further investigation. Culture optimization and fractionation of one of the most promising Streptomyces extracts resulted in the isolation of the six most bioactive and least toxic molecules, namely, Cyclo (L-proline-L-tyrosine) (1), Cyclo (Pro-Phe) (2), Lumichrome (3), P-(acetylamino) benzoic acid (4), Daidzein (5), and Uracil (6). These were tested for elastase and tyrosinase inhibition as well as antioxidant activity, and the activation of the autophagy-lysosome and the ubiquitin-proteasome system in cell lines and in Drosophila melanogaster. Molecules (1) and (4) demonstrated moderate elastase inhibition, while molecules (2), (3), (5), and (6) reduced reactive oxygen species under certain conditions. None activated the proteasome but all increased lysosomal activity in cell lines. Molecules (1), (2), (4), (6) were selected for study on Drosophila. Molecules (1) and (2) increased the activity of proteasome and molecules (1), (2), (4) increased the activity of lysosomes. All four molecules triggered antioxidant responses in Drosophila. This study highlights the potential of Greek actinobacterial biodiversity as a valuable resource for developing novel antiaging compounds with significant therapeutic implications for skin aging.

The CH2Cl2-MeOH extract of the Mediterranean sponge Crambe crambe was investigated via UHPLC-HRMS/MS employing manual dereplication and in silico mass spectrometry tools. A deconvolution approach was implemented toward the extensive metabolic characterization of the sample, resulting in the annotation of 53 compounds. The analysis of data-dependent HRMS/MS scans was conducted to establish fragmentation patterns characteristic of each crambescin A, B and C sub-families. Among the 39 compounds identified from these groups, 22 analogues were reported for the first time including 4 new homologous series that differed by the ratio of methylene units in the upper (n+2) and lower (m+2) alkyl side chains. More specifically, crambesins presenting m= 5 or 6 and n=5 (compounds 7, 11, 22 and 24) as well as m=5 or 6 and n=4 (compounds 5, 6, 8, 9, 12 and 14) were characterized. Additionally, 4 new crambescidin analogues (compounds 13, 15, 35 and 39) were also reported. The identity of the dereplicated features was further validated by studying crambescins spectral similarities through feature-based molecular networking approach. Overall, this study suggests UHPLC-HRMS/MS—through integration of manual and computational dereplication approaches—as a valuable tool for the investigation and high-throughput characterization of C. crambe metabolome.

Heliotropium procumbens, a Boraginaceae species native to Panama, has remained largely unexplored regarding its nitrogen-containing metabolites, including pyrrolizidine alkaloids (PAs). In the current study, a comprehensive phytochemical investigation of its aerial parts is presented using HPLC-DAD-IT-MS, UHPLC–HRMS, and GC-MS primarily to profile its PA composition. A total of twelve PAs and N-oxides (PANOs) were identified, along with two phenolamides—including N¹, N¹⁰-diferuloylspermidine, which is biosynthetically related to PAs—and the distinctive metabolite heliotropamide. The detected PAs included unsaturated necines, primarily monoesters of retronecine and heliotridine, as well as saturated PAs such as a platynecine-type PA and the less commonly encountered triol necines and their N-oxides. Among these, helifoline-N-oxide was isolated and structurally elucidated by NMR spectroscopy for the first time as a natural product. Comparison with the chemodiversity of PAs within the Heliotropium genus revealed a high degree of diversity in H. procumbens, which can be attributed both to the species’ inherent biosynthetic capacity for chemical variation and to the more comprehensive and extensive studies conducted on it, which naturally enrich the apparent diversity observed. This work expands the phytochemical knowledge of H. procumbens and contributes to a broader understanding of PA diversity in the genus, offering new insights into their potential ecological and toxicological significance.

The CH₂Cl₂-MeOH extract of the Mediterranean sponge Crambe crambe was investigated via UHPLC-HRMS/MS employing manual dereplication and in silico mass spectrometry tools. A deconvolution approach was implemented for the extensive metabolic characterization of the sample, resulting in the annotation of 53 compounds. The analysis of data-dependent HRMS/MS scans was conducted to establish fragmentation patterns characteristic of each crambescin A, B, and C sub-families. Among the 39 compounds identified from these groups, 22 analogues were reported for the first time including 4 new homologous series that differed by the ratio of methylene units in the upper (n + 2) and lower (m + 2) alkyl side chains. More specifically, crambescins presenting m = 5 or 6 and n = 5 (compounds 7, 11, 22 and 24) as well as m = 5 or 6 and n = 4 (compounds 5, 6, 8, 9, 12 and 14) were characterized. Additionally, four new features, potentially corresponding to new crambescidin analogues (compounds 13, 15, 35, and 39), were also reported. The identity of the dereplicated features was further validated by studying crambescins’ spectral similarities through a feature-based molecular networking approach. Overall, this study suggests UHPLC-HRMS/MS—through the integration of manual and computational dereplication approaches—as a valuable tool for the investigation and high-throughput characterization of the C. crambe metabolome.