Evaluation of ALA-capped silver, copper, and silver-copper nanoparticles for controlling fungal plant pathogens.

Phytopathogenic fungi significantly threaten global food security, causing substantial yield and quality losses. Sustainable solutions are urgently needed to combat these agricultural pathogens. This study explored the potential of silver (Ag), copper (Cu), and combined Ag/Cu nanoparticles capped with aminolevulinic acid (ALA) as antifungal agents. The nanoparticles (ALAAg, ALACu, and ALAAgCu) were synthesized via photoreduction and characterized using various techniques (UV-Vis, TEM, XRD, Zeta potential). Their antifungal activity against four key plant pathogens (Alternaria grandis, Colletotrichum truncatum, Corynespora cassiicola, and Fusarium oxysporum) was evaluated using poisoned food techniques. Notably, ALAAgCuNPs demonstrated superior antifungal activity compared to a conventional fungicide against two fungal strains. Even at lower concentrations, ALAAgCuNPs exhibited fungistatic effects comparable to those of the control. These promising results suggest the potential of ALAAgCu NPs as a broad-spectrum, potentially eco-friendly alternative for fungal control in plants and seeds. This approach is crucial for ensuring crop health, harvest quality, and food safety.

Long-term ayahuasca use is associated with preserved global cognitive function and improved memory: a cross-sectional study with ritual users.

Although several studies have been conducted to elucidate the relationship between psychedelic consumption and cognition, few have focused on understanding the long-term use influence of these substances on these variables, especially in ritualistic contexts.  To verify the influence of ritualistic ayahuasca consumption on the cognition of experienced ayahuasca religious users (> 20 years) and beginners (< 3 years), which participated in rituals of the Centro Luz Divina (CLD), a Santo Daime church in Brazil. Observational, descriptive, and cross-sectional study was carried out in which 48 people participated divided into three groups: (a) experienced ayahuasca users (n = 16), (b) beginner ayahuasca users (n = 16) and (c) control group (n = 16). All groups were matched by sex, age, and education and contained 8 women and 8 men. Cognition was assessed with the WASI (intelligence quotient), Digit Span (verbal working memory), Corsi Block-Tapping Task (visuospatial-related and working memory), Rey-Osterrieth Complex Figure test (visual perception, immediate memory), and Wisconsin Card Sorting and Five Digit Test (executive functions). Groups were homogenous in terms of sociodemographic characteristics, with participants presenting average intellectual performance. There was no evidence of cognitive decline amongst ayahuasca users. The experienced group showed higher scores compared to the less experienced group in the Digit Span and Corsi Block-Tapping tasks, which assess working verbal and visuospatial memories respectively. We confirmed the botanical identities of Psychotria viridis and Banisteriopsis caapi and the presence of the alkaloids both in the plants and in the brew. Short and long-term ayahuasca consumption does not seem to alter human cognition, while long-term use seems to be associated with improvements in aspects of working memory when compared with short-term use.

Discovering New Natural Products Using Metabolomics-Based Approaches.

The incessant search for new natural molecules with biological activities has forced researchers in the field of chemistry of natural products to seek different approaches for their prospection studies. In particular, researchers around the world are turning to approaches in metabolomics to avoid high rates of re-isolation of certain compounds, something recurrent in this branch of science. Thanks to the development of new technologies in the analytical instrumentation of spectroscopic and spectrometric techniques, as well as the advance in the computational processing modes of the results, metabolomics has been gaining more and more space in studies that involve the prospection of natural products. Thus, this chapter summarizes the precepts and good practices in the metabolomics of microbial natural products using mass spectrometry and nuclear magnetic resonance spectroscopy, and also summarizes several examples where this approach has been applied in the discovery of bioactive molecules.

Karmitoxin: An Amine-Containing Polyhydroxy-Polyene Toxin from the Marine Dinoflagellate Karlodinium armiger.

Marine algae from the genus Karlodinium are known to be involved in fish-killing events worldwide. Here we report for the first time the chemistry and bioactivity of a natural product from the newly described mixotrophic dinoflagellate Karlodinium armiger. Our work describes the isolation and structural characterization of a new polyhydroxy-polyene named karmitoxin. The structure elucidation work was facilitated by use of 13C enrichment and high-field 2D NMR spectroscopy, where 1H-13C long-range correlations turned out to be very informative. Karmitoxin is structurally related to amphidinols and karlotoxins; however it differs by containing the longest carbon-carbon backbone discovered for this class of compounds, as well as a primary amino group. Karmitoxin showed potent nanomolar cytotoxic activity in an RTgill-W1 cell assay as well as rapid immobilization and eventual mortality of the copepod Acartia tonsa, a natural grazer of K. armiger.

HPLC-HRMS Quantification of the Ichthyotoxin Karmitoxin from Karlodinium armiger.

Being able to quantify ichthyotoxic metabolites from microalgae allows for the determination of ecologically-relevant concentrations that can be simulated in laboratory experiments, as well as to investigate bioaccumulation and degradation. Here, the ichthyotoxin karmitoxin, produced by Karlodinium armiger, was quantified in laboratory-grown cultures using high-performance liquid chromatography (HPLC) coupled to electrospray ionisation high-resolution time-of-flight mass spectrometry (HRMS). Prior to the quantification of karmitoxin, a standard of karmitoxin was purified from K. armiger cultures (80 L). The standard was quantified by fluorescent derivatisation using Waters AccQ-Fluor reagent and derivatised fumonisin B1 and fumonisin B2 as standards, as each contain a primary amine. Various sample preparation methods for whole culture samples were assessed, including six different solid phase extraction substrates. During analysis of culture samples, MS source conditions were monitored with chloramphenicol and valinomycin as external standards over prolonged injection sequences (>12 h) and karmitoxin concentrations were determined using the response factor of a closely eluting iturin A2 internal standard. Using this method the limit of quantification was 0.11 µg·mL-1, and the limit of detection was found to be 0.03 µg·mL-1. Matrix effects were determined with the use of K. armiger cultures grown with 13C-labelled bicarbonate as the primary carbon source.

Dichlorinated and Brominated Rugulovasines, Ergot Alkaloids Produced by Talaromyces wortmannii.

UHPLC-DAD-HRMS based dereplication guided the detection of new halogenated alkaloids co-produced by Talaromyces wortmannii. From the fungal growth in large scale, the epimers 2,8-dichlororugulovasines A and B were purified and further identified by means of a HPLC-SPE/NMR hyphenated system. Brominated rugulovasines were also detected when the microbial incubation medium was supplemented with bromine sources. Studies from 1D/2D NMR and HRMS spectroscopy data allowed the structural elucidation of the dichlorinated compounds, while tandem MS/HRMS data analysis supported the rationalization of brominated congeners. Preliminary genetic studies revealed evidence that FADH2 dependent halogenase can be involved in the biosynthesis of the produced halocompounds.

Unveiling new myrsinoic acids and AChE ligands from Myrsine guianensis (Aubl.) Kuntze.

Molecular dereplication and drug-like discovery are important tools for exploring the chemical profile of metabolites in a complex mixture. In order to establish a workflow for discovering novel acetylcholinesterase (AChE) ligands, we performed the chemical study of Myrsine guianensis (Aubl.) Kuntze (Primulaceae). To carry out the bioprospection, nine extracts were obtained from different parts of the plant. Through the dereplication approaches, seventeen metabolites were annotated. In order to confirm the putative inferences, a HPLC preparative method was developed to isolate three known myrsinoic acids, A(1), B(2) and C(3). Along with, we are reporting the obtention of two new congeners, G(5) and H(6), which their structures were elucidated by NMR and HRMS data. Besides that, two extracts were submitted to affinity assays to accelerate the discovery of AChE ligands. Desorbates were analyzed through LC-HRMS for calculating the affinity ratio (AR). Thus, (1) presented AR = 4.59, therefore was considered a potential ligand.

plantMASST - Community-driven chemotaxonomic digitization of plants.

Understanding the distribution of hundreds of thousands of plant metabolites across the plant kingdom presents a challenge. To address this, we curated publicly available LC-MS/MS data from 19,075 plant extracts and developed the plantMASST reference database encompassing 246 botanical families, 1,469 genera, and 2,793 species. This taxonomically focused database facilitates the exploration of plant-derived molecules using tandem mass spectrometry (MS/MS) spectra. This tool will aid in drug discovery, biosynthesis, (chemo)taxonomy, and the evolutionary ecology of herbivore interactions.

Chemodiversity and Anti-Leukemia Effect of Metabolites from Penicillium setosum CMLD 18.

Penicillium setosum represents a Penicillium species recently described, with little up-to-date information about its metabolic and biological potential. Due to this scenario, we performed chemical and biological studies of P. setosum CMLD18, a strain isolated from Swinglea glutinosa (Rutaceae). HRMS-MS guided dereplication strategies and anti-leukemia assays conducted the isolation and characterization of six compounds after several chromatographic procedures: 2-chloroemodic acid (2), 2-chloro-1,3,8-trihydroxy-6- (hydroxymethyl)-anthraquinone (7), 7-chloroemodin (8), bisdethiobis(methylthio)acetylaranotine (9), fellutanine C (10), and 4-methyl-5,6-diihydro-2H-pyran-2-one (15). From the assayed metabolites, (10) induced cellular death against Kasumi-1, a human leukemia cell line, as well as good selectivity for it, displaying promising cytotoxic activity. Here, the correct NMR signal assignments for (9) are also described. Therefore, this work highlights more detailed knowledge about the P. setosum chemical profile as well as its biological potential, offering prospects for obtaining natural products with anti-leukemia capabilities.

Penicillium genus as a source for anti-leukemia compounds: an overview from 1984 to 2020.

Penicillium is a widely explored genus due to its chemical diversity and associated biological properties; in addition, it represents an important source for cytotoxic compounds with good application perspectives. Based on these aspects, in this review, Penicillium compounds that presented activity against human leukemia cell lines are being listed and discussed. For this, a careful bibliographic survey was carried out in the main electronic databases, i.e. Scopus, SciFinder, Web of Science and Pubmed. Between 1984 and 2020, thirty seven original papers were selected, when using the search terms Penicillium and leukemia. The occurrence of l-asparaginase produced by some Penicillium spp. was also highlighted since this enzyme is being employed for acute lymphoblastic leukemia and lymphosarcoma therapies. Therefore, this overview aims to demonstrate the potential of metabolites biosynthesized by Penicillium fungi which can be applied in human leukemia therapies and opportunities for designing new lead compounds.

Karmitoxin production by Karlodinium armiger and the effects of K. armiger and karmitoxin towards fish.

The dinoflagellate Karlodinium armiger has a huge impact on wild and caged fish during blooms in coastal waters. Recently, a new toxin, karmitoxin, was chemically characterized from K. armiger and a quantification method was established, thereby allowing investigations of the fish killing mechanism. K. armiger is not able to grow in standard growth media that are based on nitrate as a nitrogen source, and successful cultures of this species have only been achieved in mixotrophic cultures after addition of a prey source. Here we show that addition of ammonium (up to 50 µM) to the growth media is a good alternative, as K. armiger batch cultures achieve growth rates, which are comparable to growth rates reached in mixotrophic cultures. Karmitoxin production (1.9 and 2.9 pg cell-1 d-1) and cellular karmitoxin content (8.72 ± 0.25 pg cell-1 and 7.14 ± 0.29 pg cell-1) were in the same range, though significantly different, in prey-fed cultures and monocultures supplied with ammonium, respectively. Net production of karmitoxin stopped when the K. armiger cultures reached stationary growth phase, indicating no accumulation of karmitoxin in cells or growth media. Toxicity tests towards sheepshead minnow fish larvae indicated rapid death of the fish larvae when exposed to high K. armiger cell concentrations (LT50 of 2.06 h at 44.9 × 103 cells mL-1 cultivated with ammonium). Purified toxins caused the same physical damage to fish larvae as living K. armiger cultures. An exposure of purified karmitoxin to fish larvae and rainbow trout gill cells indicated that the fish larvae were about three times less sensitive than gill cells. When comparing the effect of purified toxins with the effect of whole K. armiger cultures, twice the toxin concentration of the purified toxins was needed to cause the same effect. Although a loss of karmitoxin of twenty percent was observed during the incubation, this could not explain the apparent discrepancy. Other factors, like a direct effect of the K. armiger cells on the fish larvae or other, yet unknown toxins may influence the effect of whole cell cultures. To study the effects of released karmitoxin, fish larvae were exposed to a K. armiger culture that was treated with HP-20 resin, which adsorbs extracellular karmitoxin. The 24 h HP-20 treatment resulted in a K. armiger culture that had 37% less total karmitoxin, without a reduction in cell concentration, and a reduced toxic effect was observed in the HP-20 treated culture, as compared to non-treated controls. Fish larvae that were exposed to HP-20 treated culture were immobilized, but survived during the 12 h exposure, whereas the exposure to non-treated culture led to high mortality of the fish larvae. Direct observations under the microscope revealed no evidence of micropredation of K. armiger on the fish larvae during any of the exposures. Thus, the results presented here, indicate that released karmitoxin is the main cause for fish kills by K. armiger. Finally, we found that juvenile rainbow trout were six times more sensitive than fish larvae towards K. armiger, indicating that juvenile fish are more sensitive to K. armiger in bloom situations than early larval stages.

UHPLC-MS/MS determination of ochratoxin A and fumonisins in coffee using QuEChERS extraction combined with mixed-mode SPE purification.

A method was developed for simultaneous determination of the mycotoxins: ochratoxin A (OTA) and fumonisins B2 (FB2), B4 (FB4), and B6 (FB6) in green, roasted, and instant coffee. Extraction was performed by QuEChERS (quick, easy, cheap, effective, rugged, and safe) under acidic conditions followed by mixed-mode reversed phase-anion exchange solid phase extraction. OTA and FB2 were detected at levels down to 0.5 and 2 µg/kg by UHPLC-MS/MS and quantitated via isotope dilution using U-(13)C-labeled FB2 and OTA as internal standards. Mixing 20% isopropanol in the acetonitrile of the acidic UHPLC gradient system increased the signal intensity by 50% and decreased the ion-suppression with 50-75% in roasted coffee samples. About half of the roasted coffee samples (n = 57, from 9 countries) contained detectable levels of OTA, however, with only 5 samples above the EU regulatory limit of 5 µg/kg and the highest with 21 µg/kg. None of the 25 instant coffee samples contained OTA above the EU regulatory level of 10 µg/kg. Nonetheless, the toxin could be detected in 56% of the analyzed instant coffee samples. Fumonisins were not detected in any of the roasted or instant coffee samples (n = 82). However, in the green coffee samples (n = 18) almost half of the samples were positive with a maximum value of 164 µg/kg (sum of FB2, FB4, and FB6). This discrepancy between green coffee and processed coffees indicated that the fumonisins decompose during the roasting process, which was confirmed in roasting experiments. Here fumonisins could not be detected after roasting of the green, 164 µg/kg coffee, sample. Under the same conditions, OTA was reduced from 2.4 to 0.5 µg/kg.