Culture Extracts Research Articles

In vitro, in vivo and in silico anticancer activity and toxicity of Usnic acid extracted from the mycobiont culture of Usneabaileyi.

Lichen has widespread uses in indigenous medicinal traditions, and it has been used to treat various conditions. Among the various active compounds in the lichen, Usnic acid has unique biological activities. However, the toxicology and the mechanism of action of Usnic acid need to be examined in detail to the extent of clinical trials. Hence, the present research is aimed to isolate usnic acid from the mycobiont culture of Usnea baileyi lichen and explore its toxicological properties and those of pharmacological importance. The methanolic extract of these mycobiont cultures has shown the presence of Usnic acid via column chromatography. The Usnic acid was computationally evaluated using molecular docking studies (GLIDE docking) and MMGBSA energy calculations. The results obtained from TLC, LC-MS, UV-VIS spectrophotometer, NMR spectroscopy (1H and 13c), and FT-IR indicated the presence of Usnic acid. An acute toxicity analysis by an in vivo study using a zebrafish animal model showed acute toxicity. The standard disc diffusion and DPPH assay suggested an antimicrobial and antioxidant activity with this lichen compound. The in vitro analysis of Usnic acid in G361 skin cancer cell lines demonstrated its anticancer potential against malignant melanoma. Furthermore, a molecular in silico study returned a good docking score of -7.977, and the MMGBSA free energy was measured as -18 and -30. Therefore, further extensive research on Usnic acid might lead to the development of novel anticancer drugs from the natural sources.

In vitro toxicity of microalgae species of the phyla chlorophyta and ochrophyta in CHO-k1 and HEP G2 cells for potential use in human nutrition

Microalgae are a promising component to enhance human nutrition, but to date only a few species have been authorized to be used in human nutrition. In this study, the in vitro toxicity of eight novel microalgae strains (Botryococcus braunii, Chlorococcum novae-angliae, Microchloropsis salina, Myrmecia bisecta, Stichococcus sp. Tetraselmis suecica, Tetradesmus obliquus and Spongiochloris minor) selected for their potential for human nutrition was investigated. N-hexane, acetone, ethanol and aqueous extracts of the lyophilized biomass were tested in the CHO-k1 and HEP G2 cell lines at concentrations of up to 1.81 mg ml-1 extracted biomass per well. None of the tested microalgae extracts reached values defined as a significant cytotoxic effect (IC50 < 0.02 mg ml-1). The highest cytotoxic effects were measured for Stichococcus sp. in both cell lines with IC50 values of 0.17 mg ml-1 for the acetone extract in CHO-k1 cell culture and 0.18 mg ml-1 for the acetone extract in HEP G2 cell culture. Most cytotoxic effects occurred with the acetone and ethanol extracts, while the water and n-hexane extracts showed almost no measurable cytotoxic effects. Only Tetraselmis suecica showed no cytotoxic effect under the chosen conditions in both tested cell lines, marking this microalgae as particularly interesting for further investigations into its use in human nutrition.

Chemical composition of anti-microbially active fractions derived from extract of filamentous fungus Keratinophyton Lemmensii including three novel bioactive compounds

Screening for new bioactive microbial metabolites, we found a novel okaramine derivative, for which we propose the trivial name lemmokaramine, as well as two already known okaramine congeners – okaramine H and okaramine J - responsible for antimicrobial activity of the recently described microscopic filamentous fungus, Keratinophyton lemmensii BiMM-F76 (= CCF 6359). In addition, two novel substances, a new cyclohexyl denominated lemmensihexol and a new tetrahydroxypyrane denominated lemmensipyrane, were purified and characterized. The compounds were isolated from the culture extract of the fungus grown on modified yeast extract sucrose medium by means of flash chromatography followed by preparative HPLC. The chemical structures were elucidated by NMR and LC-MS. The new okaramine (lemmokaramine) exerted antimicrobial activity against Gram-positive and Gram-negative bacteria, yeasts and fungi and anticancer activity against different mammalian cell lines (Caco-2, HCT116, HT29, SW480, MCM G1, and MCM DLN). Furthermore, we found a significant antioxidant effect of lemmokaramine following H2O2 treatment indicated by activation of the Nrf2 pathway. This is the first report describing analysis and structural elucidation of bioactive metabolites for the onygenalean genus Keratinophyton.

Production of Phenyldiazene Derivatives Using the Biosynthetic Pathway of an Aromatic Diazo Group-Containing Natural Product from an Actinomycete.

The diazo group is an important functional group in organic synthesis because it confers high reactivity to the compounds and has been applied in various chemical reactions, such as the Sandmeyer reaction, Wolff rearrangement, cyclopropanation, and C-N bond formation with active methylene compounds. Previously, we revealed that 3-diazoavenalumic acid (3-DAA), which is potentially produced by several actinomycete species and contains an aromatic diazo group, is a biosynthetic intermediate of avenalumic acid. In this study, we aimed to construct a production system for phenyldiazene derivatives by adding several active methylene compounds to the culture of a 3-DAA-producing recombinant actinomycete. First, acetoacetanilide and its derivatives, which have an active methylene and are raw materials for arylide yellow dyes, were individually added to the culture of a 3-DAA-producing actinomycete. When their metabolites were analyzed, each expected compound with a phenyldiazenyl moiety was detected in the culture extract. Moreover, we established a one-pot in vitro enzymatic production system for the same phenyldiazene derivatives using a highly reactive diazotase, CmaA6. These results showed that the diazo group of natural products is an attractive tool for expanding the structural diversity of natural products both in vivo and in vitro.

Влияние биологически активных веществ на тепловой и окислительный стресс модельных объектов Caenorhabditis elegans

Modern medicine strives to prevent age-related diseases. Oxidative stress is associated with development and progression of various diseases. Reactive oxygen species are part of vital physiological processes. High levels of reactive oxygen lead to stress and pathology whereas low ones are associated with healthy physiology. Plant-derived adaptogens demonstrate good results in stress tolerance and homeostasis. Plant materials are a pharmacologically optimal source of chemical compounds to treat various diseases, including those caused by oxidative stress. The research featured biologically active substances isolated from extracts of callus, suspension, and root cultures of medicinal plants. Baicalin and trans-cinnamic acid were obtained from Scutellaria baicalensis while ursolic acid came from Thymus vulgaris. The biologically active substances were tested for neuroprotective properties, as well as for the impact on the expression of SOD-3 and HSP-16.2. Caenorhabditis elegans served as a model to study the accumulation of carbonylated proteins and lipofuscin. The neuroprotective activity of all tested substances decreased as their concentration fell from 200 to 10 μmol/L. C. elegans proved more resistant to thermal stress if pretreated with the biologically active substances. In response to thermal stress, nematodes expressed SOD-3 more actively than HSP-16.2. At 100 μmol/L, the biologically active substances could reduce the level of carbonylated proteins. Ursolic acid was especially effective against protein carbonylation and lipofuscin accumulation in all concentrations. Baicalin, trans-cinnamic acid, and ursolic acid made it possible to reduce oxidative and thermal stress, thus demonstrating good prospects for further studies as part of adaptogenic prepa rations.

Biological active compounds from edible mushroom: Effect on hela cells

Background: Mushrooms are unique organisms rich in proteins, carbohydrates, vitamins, and minerals, which allows them to be used not only as a source of nutrition but also in cosmetics and medicine. The fungus P. ostreatus possesses two strong enzymatic complexes – lignin-decaying and cellulose-decaying. Objective: The research aimed to study the antitumor activity of treated mm-wave mycelial extracts of mushrooms on the HeLa cell line. Research on edible mushrooms against tumors is very actual and important for the field of functional food science and bioactive compounds field. Methods: The effect of the fungal culture extract on the cyto-photometric and morphometric indices of the HeLa cell line culture was studied according to the nature of proliferation and transcription of malignant tissues. Using high-resolution liquid chromatography, the amino acid composition of acid-soluble proteins in mycelial extracts of a mushroom culture treated with mm waves was studied. Results: Extracts of P. ostreatus mycelial culture significantly suppressed the growth of proliferative activity of human tumor cultures and completely suppressed the mitotic activity of continuous HeLa cell cultures to 72 hours. Our study revealed a significant increase in glutamic and aspartic acids in mycelial culture extracts treated by mm-waves. Conclusion: Based on HPLC analysis, we suggest polyfunctional peroxidase as the candidate for the role of biologically active compound in the composition of aqueous extracts of the mycelial culture. So, the polyfunctional peroxidase plays an active role in the therapeutic properties of the oyster mushroom. Keywords: anticancer activity, ant-inflammation activity, mm-waves, mushroom`s, mycelial extracts, HPLC of acid-soluble proteins, HeLa cell line.

Investigation of Antimicrobial Compounds Produced by Endolichenic Fungi in Different Culture Media.

Continuous use of synthetic fungicides has led to explosive emergence of fungicide-resistant microbes. Therefore, there are urgent needs for environmentally friendly antimicrobial agents with novel modes of action. This study investigated endolichenic fungi (ELF) as a source of antimicrobial compounds against various plant pathogens. We utilized an One Strain MAny Compounds (OSMAC) approach to enhance the chemical diversity of fourteen ELF. This involved cultivation of ELF in four growth media and subsequently assessing antimicrobial activities of culture extracts. Nearly half of the culture extracts exhibited antimicrobial activity against a Gram-positive bacterium, but showed minimal activity against Gram-negative bacteria tested. Notably, culture extracts from two ELF, Chaetomium globosum and Nodulisporium sp., demonstrated significant inhibitory effects against plant pathogenic fungi. LC-MS/MS-based metabolome profiling confirmed the presence of known bioactive compounds like cyclic dipeptides and chaetoglobosins. These findings highlight the effectiveness of combining OSMAC and metabolomics for identifying antimicrobial agents for agricultural use.

Exploring Antimicrobial Potency, ADMET, and Optimal Drug Target of a Non-ribosomal Peptide Sevadicin from Bacillus pumilus, through In Vitro Assay and Molecular Dynamics Simulation.

The current study was designed to explore the biosynthetic potential of sevadicin in Bacillus pumilus species and its interaction with bacterial drug target molecules. The non-ribosomal peptide (NRP) cluster in B. pumilus SF-4 was preliminarily confirmed using PCR-based screening, and the bioactivity of strain SF-4 culture extract was assessed against a set of human pathogenic strains. The susceptibility assay showed that strain SF-4 extract had higher inhibitory concentrations (312-375µg/mL) than ciprofloxacin. Genome mining of B. pumilus strains (n = 22) using AntiSMASH and BAGEL identified sevadicin coding biosynthetic gene cluster only in strain SF-4, constitutes of two core biosynthetic genes, three additional biosynthetic genes, two transport-related genes, and one regulatory gene. The molecular docking of sevadicin with various putative bacterial drug targets such as dihydropteroate, muramyl ligase E, topoisomerase, penicillin-binding protein, and in vitro safety analyses were conducted with detailed ADMET screening. The results showed that sevadicin makes hydrophobic interaction with MurE (PDB ID: 1E8C and 4C13) via hydrogen bonding, suggesting bacterial growth inhibition by disrupting the cell wall synthesis pathway and exhibiting a secure biosafety profile. The stability and compactness of sevadicin/MurE complexes were assessed via molecular dynamic simulation using RMSD, RMSF, and Rg. The simulation results revealed the binding stability of sevadicin/MurE complexes and indicated that the complexes can't be easily deformed. In conclusion, the current study explored the biosynthesis of sevadicin in B. pumilus for the first time and found that sevadicin inhibits bacterial growth by inhibiting cell wall synthesis via targeting the MurE enzyme and exhibits no toxicity.

Antibacterial and Anticancer Properties of Endophenazines from Streptomyces prasinus ZO16, an Endophyte in Zingiber officinale Rosc.

<b>Background and Objective:</b> This study investigated a bacterial strain, ZO16, isolated from ginger (<i>Zingiber officinale</i>) roots. Analysis of its 16S ribosomal DNA (rDNA), along with chemical and physical properties, revealed it to be <i>Streptomyces prasinus</i>. This study aimed to isolate and characterize the main bioactive compounds from ZO16, evaluating their antibacterial and anticancer properties. <b>Materials and Methods:</b> Techniques like column chromatography and thin-layer chromatography (TLC) were used to purify the key compounds from ZO16's culture extract. Nuclear Magnetic Resonance (NMR) spectroscopy and mass spectrometry were utilized to confirm the identities of the purified compounds as endophenazine A (compound 1) and endophenazine B (compound 2). The antibacterial and anticancer properties of these compounds were then evaluated. <b>Results:</b> The isolated compounds displayed antibacterial activity against <i>Staphylococcus aureus</i> ATCC 25923 and Methicillin-Resistant <i>Staphylococcus aureus</i> (MRSA). The minimum inhibitory concentration (MIC) of the isolated compounds against bacteria ranged from 8 to 32 μg/mL, while the minimum bactericidal concentration (MBC) was between 32 and 128 μg/mL. These compounds exhibited effectiveness against tested cancer cells with IC<sub>50</sub> values ranging from 30.40 to 32.51 μg/mL for cervical cancer (HeLa), 78.32 to 86.45 μg/mL for liver cancer (HepG2) and 23.41 to 28.26 μg/mL for breast cancer (MDA-MB-231) cells. However, these compounds also showed moderate toxicity towards non-cancerous Vero cells (IC<sub>50</sub> = 317.44-328.63 μg/mL). <b>Conclusion:</b> The findings of this study suggest that <i>Streptomyces prasinus</i> strain ZO16 produces compounds with antibacterial and anticancer properties. Further investigation of these compounds has the potential to contribute to the development of improved methods for controlling and treating bacterial infections and some cancers.

Developing endophytic Penicillium oxalicum as a source of lignocellulolytic enzymes for enhanced hydrolysis of biorefinery relevant pretreated rice straw.

Endophytic fungi, as plant symbionts, produce an elaborate array of enzymes for efficient disintegration of lignocellulosic biomass into constituent monomeric sugars, making them novel source of lignocellulolytic CAZymes with immense potential in future biorefineries. The present study reports lignocellulolytic enzymes production potential of an endophytic halotolerant Penicillium oxalicum strain isolated from Citrus limon, under submerged and solid-state fermentation (SmF & SSF, respectively), in the presence and absence of salt (1M NaCl). The comparative QTOF-LC/MS-based exoproteome analysis of the culture extracts unveiled differential expression of CAZymes, with the higher abundance of GH6 and GH7 family cellobiohydrolase in the presence of 1M salt. The strain improvement program, employing cyclic mutagenesis and diploidization, was utilized to develop hyper-cellulase producing mutant strains of P. oxalicum. The enzyme production of the developed strain (POx-M35) was further enhanced through statistical optimization of the culture conditions utilizing glucose mix disaccharides (GMDs) as an inducer. This optimization process resulted in the lignocellulolytic cocktail that contained high titers (U/mL) of endoglucanase (EG) (146.16), cellobiohydrolase (CBHI) (6.99), β-glucosidase (β-G) (26.21), xylanase (336.05) and FPase (2.02 U/mL), which were 5.47-, 5.54-, 8.55-, 4.96-, and 4.39-fold higher when compared to the enzyme titers obtained in wild HP1, respectively. Furthermore, the lignocellulolytic cocktails designed by blending secretome produced by mutant POx-M35 with xylanases (GH10 and GH11) derived from Malbranchea cinnamomea resulted in efficient hydrolysis of unwashed acid pretreated (UWAP) rice straw slurry and mild alkali deacetylated (MAD) rice straw. This study underscores the potential of bioprospecting novel fungus and developing an improved strain for optimized production and constitution of lignocellulolytic cocktails that can be an important determinant in advancing biomass conversion technologies.

Wychimicins E and F from a rare actinomycete of the genus Cryptosporangium.

Two new spirotetronate class compounds designated wychimicins E (1) and F (2) were isolated from the culture extract of an actinomycete Cryptosporangium sp. RD061707. Their structures were determined through extensive NMR analysis in comparison with wychimicin C. Both compounds lack one hydroxy group in the decalin moiety, and the relative configuration of the remaining hydroxy group was assigned by NMR analysis of triacetylwychimicin E (3). Compounds 1 and 2 showed potent antimicrobial activity against Kocuria rhizophila and Staphylococcus aureus. These compounds were also modestly cytotoxic against P388 murine leukemia cells.

Sporangimicins A-D, acylated maltose derivatives from a rare actinomycete of the genus Pseudosporangium.

Sporangimicins A-D (1-4), four anomeric pairs of diacyl disaccharides that represent a new metabolite class, were discovered from the culture extract of an actinomycete Pseudosporangium sp. RD061809. Compounds 1-4 caused peak separation in the HPLC chromatogram and partial duplication of the NMR resonances by anomeric interconversion of a maltose core modified at the two sugar 6-positions with an isobutanoyl and a methyl-branched long-chain dienoyl groups. A highlight of the structure elucidation was application of Ohrui-Akasaka's method to a chromatographically inseparable mixture of 3 and 4, which proved the composition ratio of 3 and 4 to be 82:18 and the R/S ratio at the anteiso-methyl bearing chiral center in 3 to be 66:34. Compounds 1-4 showed antimicrobial activity against Gram-positive bacteria and modest cytotoxicity toward P388 murine leukemia cells.

Investigation of miniature goat cheese produced from Cynara scolymus L. pistil cell suspension culture extracts elicited with melatonin and salicylic acid.

The present study aimed to evaluate the potential of enzymes produced by artichoke (Cynara scolymus L.) flower cell suspension cultures elicited by melatonin (5 μm) and salicylic acid (50 μm) on the production and characteristics of miniature goat cheeses. In this study, five types of fresh miniature goat cheese were produced using whole artichoke flower extract, salicylic acid (50 μm) or melatonin (5 μm) treated artichoke suspension cell culture extracts, a culture extract without elicitor treatment (control) and rennin enzyme. The milk clotting activity values of the enzymes were measured in the range 0.52-0.74. The effect of the enzymes on the titratable acidity, water-soluble nitrogen, ripening index, αs-caseins and β-caseins of goat cheese was significant (P < 0.05). The highest level of casein degradation was observed in the cheese produced with the enzyme containing melatonin, followed by the cheese produced with the enzyme containing salicylic acid. For the enzymes produced by the suspension cell culture method, the addition of melatonin and salicylic acid had a slightly positive effect on the proteolytic activity of the extracts. It was also found that the enzymes obtained from artichokes by the suspension cell culture method did not achieve successful cheese production in terms of chemical, textural and biochemical aspects compared to those obtained from animal enzymes. © 2024 Society of Chemical Industry.

Ether-Diol Ambiguity: An Inconspicuous Issue in the Structure Elucidation of Oxygenated Natural Products.

Tertiary and allylic hydroxyl groups readily eliminate water during positive ion mode mass spectrometry and may show similar NMR spectra to their corresponding ethers. In a routine structure elucidation workflow, these factors can cause researchers to incorrectly assign diol moieties as ethers or vice versa, leading to inaccurate chemical structures. After facing this problem during our work on oxygenated sesquiterpenoids from a Fusarium sp. fungal strain, we became aware of this challenging issue. We examined the literature for oxygenated natural products bearing these functional groups, and with the aid of density functional calculations of NMR chemical shifts, we now report the structures of 15 natural products that should be revised. We further establish that derivatizing sub-micromolar amounts of alcohols to their sulfates can be used to distinguish these from their corresponding ethers using liquid chromatography negative ion mode mass spectrometry. Finally, we isolated lignoren/cyclonerodiol from the Fusarium sp. culture extract and supported its revised identity as cyclonerodiol using this sulfation approach. Our results suggest that ether-diol ambiguity could be a prevalent issue affecting the structure elucidation of oxygenated natural products and highlight the importance of using complementary techniques, such as sulfation with LC-(-)-ESI-MS or density functional calculations of NMR chemical shifts.

Exploring Indonesian actinomycete extracts for anti-tubercular compounds: Integrating inhibition assessment, genomic analysis, and prediction of its target by molecular docking

Tuberculosis (TB) is the foremost cause of infectious fatality globally. The primary global challenge in combatting TB lies in addressing the emergence of drug-resistant variants of the disease. However, the number of newly approved agents for treating TB has remained remarkably low over recent decades. Hence, research endeavors for discovering novel anti-TB agents are always needed. In the present study, we screened over 1,500 culture extracts from actinomycetes isolated in Indonesia for their inhibitory activity against Mycobacterium smegmatis used as a surrogate in the primary screening. The initial screening yielded approximately 6.2 % hit extracts, with a selection criterion of >80 % growth inhibition. The confirmed hit extracts were subsequently subjected to growth inhibition assay against Mycobacterium bovis and Mycobacterium tuberculosis. Approximately 20 % of the hit extracts that showed growth inhibition also exhibited efficacy against M. bovis BCG and M. tuberculosis H37Rv pathogenic strain. An active compound was successfully purified from a large-scale culture of the most potent representative extract by high-performance liquid chromatography and thin-layer chromatography. The structure of the active compound was elucidated by mass spectrometry and nuclear magnetic resonance. This compound displayed structural similarities to actinomycin group and exhibited robust inhibition, with IC50 values of 0.74, 0.02, and 0.07 μg/mL against M. smegmatis, M. bovis, and M. tuberculosis, respectively. The Actinomycetes strain A612, which produced the active compound, was taxonomically classified by phylogenetic analysis of 16s rRNA gene and whole genome sequencing data as Streptomyces parvus. Computational genome analysis utilizing anti-SMASH 7.0 unveiled that S. parvus A612 strain harbors 40 biosynthetic gene clusters with the potential to produce 16 known (with >70 % similarity) and 24 unknown compounds. A non-ribosomal peptide synthesis (NRPS) gene cluster associated with actinomycin D biosynthesis was also identified, boasting an 85 % similarity. Molecular docking analysis of actinomycin D and 21 potential M. tuberculosis targets revealed possible interactions with multiple targets. The purified active compound inhibited recombinant M. tuberculosis shikimate kinase (MtSK), which validated the results obtained from the docking analysis.

Progress in culture technology and active substance research on Nostoc sphaeroides Kützing.

Nostoc sphaeroides Kützing is a freshwater edible cyanobacterium that is rich in active substances such as polysaccharides, proteins and lipids; it has a variety of pharmacological effects such as antioxidant, anti-inflammatory, antitumor and cholesterol-lowering effects; and is often used as a traditional Chinese medicine with many potential applications in food, cosmetics, medical diagnostics and disease treatment. However, to meet the needs of different fields, such as medicine, there is an urgent need for basic research and technological innovation in culture technology, extraction and preparation of active substances, and the pharmacological mechanism of N. sphaeroides. This paper reviews the pharmacological effects of N. sphaeroides active substances, discusses current culture techniques and methods for extracting active components, and outlines the challenges encountered in cultivating and industrializing N. sphaeroides while discussing future development trends. © 2024 Society of Chemical Industry.

Animation Production Contributes to the Innovation and Development of Intangible Oil Paper Umbrella Culture

This study explores using animation to enhance the cultural promotion and innovation of intangible heritage oil paper umbrellas. These umbrellas, rich in history and craftsmanship, face modern challenges like market decline and skill inheritance issues. The research seeks to offer fresh insights and strategies for cultural preservation and innovation through animation. It starts by examining the cultural significance and current challenges of oil paper umbrellas, such as waning market interest and a lack of youth engagement with tradition. The study then discusses the role of animation in promoting intangible culture, highlighting its potential to raise awareness, engage the youth, and showcase traditional Chinese charm. The research suggests innovative promotional ideas like animation series, interactive platforms, and cultural events, focusing on multimedia, storytelling, and technology. It also proposes new concepts like cultural element extraction, cross-industry collaboration, and digital presentation to highlight the umbrella's unique appeal and facilitate its promotion. The conclusion emphasizes that animation is a powerful tool for cultural promotion and preservation, with the potential to revitalize the oil paper umbrella culture in the modern era. It anticipates future developments in market reach, education, cultural exchange, and social engagement, stressing the need for ongoing innovation in cultural inheritance. Overall, the study aims to provide a theoretical foundation and practical guidance for the preservation and innovative development of the oil paper umbrella culture, fostering integration with modern society and promoting sustainable cultural growth.

A biosensor monitoring approach for toxic algae: Construction of calibration curves to infer cell numbers in field material

A variety of shellfish toxin-producing Harmful Algal Blooms (HABs) occur every year in coastal temperate waters worldwide. These toxic HABs may cause lengthy (months) harvesting bans of mussels and other suspension feeding bivalves exposed to their blooms. To safeguard public health and the shellfish industry, European Union regulations request periodic monitoring of potentially toxic microalgae in seawater and phycotoxins in live bivalve molluscs from shellfish production areas. Monitoring of other toxic microalgae, e.g., fish killers, is based solely on cell counts. Morphological identification and quantification of microalgal cells with light microscopy is time-consuming, requires a good expertise, and accurate identification to species level (e.g., Pseudo-nitzschia species) may require electron microscopy. Toxicity varies among morphologically similar species; there are toxic and non-toxic strains of the same species. Molecular techniques using ribosomal DNA sequences offer a possibility to identify and detect precisely the potentially toxic genus/species. In an earlier project (MIDTAL), specific probes against rRNA sequences of all HAB taxa, known at the time of the project, affecting shellfish areas worldwide were designed, and those affecting Europe were tested and calibrated against rRNA extracts of clonal cultures and field samples. Microarray technology was adopted to relate to cell numbers the fluorescence signal from the reaction of all target species probes spotted in the microarray slides with those present in a single sample extract. The EMERTOX project aimed to develop a more automatic “Lab on a chip” (LOC) technology, including a non- (cell) disruptive water concentration system and biosensors for HAB cells detection. Here, calibration curves are presented against toxic microalgae (cultures and field samples) causing endemic and emerging toxicity events in Galicia (NW Spain) and Portugal. Results here relating cell numbers to electrochemical signals will be used in an early warning biosensor for toxic algae.

Metabolic picture of microbial interaction: chemical crosstalk during co-cultivation between three dominant genera of bacteria and fungi in medicinal plants rhizosphere.

Microbial communities affect several aspects of the earth's ecosystem through their metabolic interaction. The dynamics of this interaction emerge from complex multilevel networks of crosstalk. Elucidation of this interaction could help us to maintain the balance for a sustainable future. To investigate the chemical language among highly abundant microbial genera in the rhizospheres of medicinal plants based on the metabolomic analysis at the interaction level. Coculturing experiments involving three microbial species: Aspergillus (A), Trichoderma (T), and Bacillus (B), representing fungi (A, T) and bacteria (B), respectively. These experiments encompassed various interaction levels, including dual cultures (AB, AT, TB) and triple cultures (ATB). Metabolic profiling by LC-QTOFMS revealed the effect of interaction level on the productivity and diversity of microbial specialized metabolites. The ATB interaction had the richest profile, while the bacterial profile in the monoculture condition had the lowest. Two native compounds of the Aspergillus genus, aspergillic acid and the dipeptide asperopiperazine B, exhibited decreased levels in the presence of the AT interaction and were undetectable in the presence of bacteria during the interaction. Trichodermarin N and Trichodermatide D isolated from Trichoderma species exclusively detected during coexistence with bacteria (TB and ATB). These findings indicate that the presence of Bacillus activates cryptic biosynthetic gene clusters in Trichoderma. The antibacterial activity of mixed culture extracts was stronger than that of the monoculture extracts. The TB extract exhibited strong antifungal activity compared to the monoculture extract and other mixed culture treatments. The elucidation of medicinal plant microbiome interaction chemistry and its effect on the environment will also be of great interest in the context of medicinal plant health Additionally, it sheds light on the content of bioactive constituents,and facilitating the discovery of novel antimicrobials.

Antioxidant and Anticancer Activity of Tannins Isolated from Callus Cultures of Achyranthes aspera L.

In the current study, the impact of oxidative stress induced by free radicals generated during metabolic processes was investigated. It was found to be linked to a variety of diseases due to their detrimental effects on nucleic acids and proteins. The primary focus remained on investigating the free radical-scavenging properties and potential anticancer activities of tannins found in callus cultures of A. aspera. It was observed that the induction of callus formation was notably successful when using leaf and root explants, resulting in a callus index as high as 160, as compared to stem explants in the presence of auxins. The extraction yield of callus tannins was the most abundant in chloroform extracts, although the overall antioxidant activity was comparable among chloroform, methanol, and petroleum ether callus extracts. Among these extracts, the chloroform extract from stem callus cultures grown on the MSDN medium showed the highest total antioxidant activity. Notably, tannins extracted from A. aspera leaf callus extracts displayed significant anticancer effects against the Jurket cell line. These effects were evaluated through measures such as cell viability and colony formation. The anticancer activity was notably higher in callus culture extracts as compared to the control. In summary, the results showed that in vitro biomass production can be a valuable approach for augmenting the production of bioactive compounds, such as tannins, from selected medicinal plants including A. aspera. Tannins extracted from A. aspera hold promise for their potential role in the development of new medicines.