Search Results (7,871)

The crude metabolic extract from plant biocontrol bacteria plays a very important role in sustainable agricultural production. These extracts help maintain healthy plants and have very important application prospects in biotechnology related to alleviating apple replant disease (ARD). In this study, Bacillus velezensis XC1 (T1), Bacillus amyloliquefaciens QSB-6 (T2), and Lactobacillus reuteri LBR (T3) were examined to characterize the ability of their crude metabolic extracts to alleviate ARD. The high-throughput sequencing data of the soil microbial community structure were analyzed in relation to LBR crude metabolic extracts, and an extensive untargeted metabolomic analysis of UHPLC-Qex active components was performed. Active LC-MS/MS revealed that the main secondary metabolites involved in the biological control exerted by L. reuteri included 3-hydroxypropionaldehyde, extracellular polysaccharides (EPS), p-hydroxybenzoic acid, and azelaic acid. These crude metabolic extracts significantly inhibited the growth of soil pathogenic fungi, reduced the abundance of Fusarium, promoted the abundance of beneficial bacteria such as Pseudomonas, and optimized the soil microbial community structure. Improved modern extraction and purification technologies will be able to offer additional insights into the mechanism of action of these secondary metabolites and enable them to be used in biological preparations to prevent and control ARD in the future, as well as to allow harmful chemical fumigants to be discontinued. Full article

Infectious diseases, including vector-borne and antibiotic-resistant infections, present significant global health challenges, necessitating the exploration of natural alternatives for disease control. In this study, we investigated the essential oil (EO) profile as well as larvicidal and antibacterial properties of two wild Apiaceae species used in Algeria: Daucus carota L. (DCEO) and Foeniculum vulgare Mill. (FVEO). EO was extracted from the aerial parts by steam distillation and analyzed using Gas Chromatography Mass Spectrometry (GC/MS). Major constituents identified in DCEO were geranyl acetate (50.07%) and elemicin (10.77%), while FVEO contained estragole (24.93%), fenchone (20.20%), and α-phellandrene (17.96%). Both EOs were highly effective towards Culex pipiens larvae, with low LC₅₀ values of 30.6 ± 1.06 ppm for DCEO and 34.7 ± 1.06 ppm for FVEO, indicating their potential as bioinsecticides due to their bioactivity and bioavailability. Additionally, the two Eos demonstrated moderate antibacterial efficacy against gram-positive bacteria, Staphylococcus aureus ATCC 25923 and Staphylococcus aureus MRSA ATCC 43300, respectively, with DCEO showing MIC values of 10 and 20 mg/mL, respectively, and FVEO exhibiting MIC values > 20 mg/mL. However, both EOs showed limited effectiveness against gram-negative bacteria, Escherichia coli ATCC 25922 and Klebsiella pneumonia ATCC 700603. These results highlight the potential applications of DCEO and FVEO as natural bioinsecticides and antibacterial agents, offering promising avenues for further research and development in pest control and food preservation. Full article

Glioblastoma (GBM) is the most aggressive cancer originating in the brain, but unfortunately combination treatments with resection, radiation, and chemotherapy are relatively ineffective. Therefore, novel methods of adjuvant therapy are critically needed. Cyclotides are plant-derived circular peptides that chemosensitize drug-resistant breast cancer to doxorubicin. We analyzed naturally occurring and synthetic cyclotides (Cycloviolacin O3, Cycloviolacin O19, natural Kalata B1, synthetic Kalata B1, and Vitri E) alone and in co-exposure treatments with the drug temozolomide (TMZ) in human glioblastoma cells. The cyclotides were identified by UPLC-PDA and HPLC-UV. The synthetic Kalata B1 sequence was verified with orbitrap LC-MS, and structural confirmation was provided by NMR spectroscopy. The cyclotides displayed dose-dependent cytotoxicity (IC₅₀ values 2.4–21.1 µM) both alone and as chemosensitizers of U-87 MG and T 98 cells to TMZ. In fact, a 16-fold lower concentration of TMZ (100 µM) was needed for significant cytotoxicity in U-87 MG cells co-exposed to synthetic Kalata B (0.5 µM). Similarly, a 15-fold lower concentration of TMZ (75 µM) was required for a significant reduction in cell viability in T 98 cells co-exposed to synthetic Kalata B1 (0.25 µM). Kalata B1 remained stable in human serum stability assays. The data support the assertion that cyclotides may chemosensitize glioblastoma cells to TMZ. Full article

Lentinus sajor-caju has shown potential for the bioconversion of lignocellulosic biomass in various substrates. Here, we evaluated whether L. sajor-caju affects the lignocellulosic biomass, in vitro fermentation and antioxidant properties of Astragalus membranaceus var. mongholicus (AMM) stems. The lignocellulosic biomass content and the antioxidant activity of AMM stems were determined by scanning electron microscopy, chemical component analysis, in vitro fermentation and LC-MS metabolomics after 30 days of fermentation by L. sajor-caju at 25 °C. L. sajor-caju significantly altered the rigid structure of the stems. Compared with the control condition, the lignocellulose contents were significantly reduced (p < 0.001) and improved in vitro digestibility and total volatile fatty acids. In total, 624 differential metabolites, including 201 up-regulated and 423 down-regulated, were identified in unfermented and fermented comparison groups. Correlation analysis indicated that there were strongly correlations of the total phenolic content and total antioxidant capacity. Meanwhile, the differential metabolites were primarily associated with antioxidant activity, with 4(3H)-quinazolinone, dihydrocarvyl acetate and jaceoside being the most representative. In summary, L. sajor-caju altered the composition of the cell wall of AMM stems, thereby enhancing their antioxidant activity. Full article

Triterpenoid saponins from the Astragalus species possess valuable effects (cytotoxic, adjuvant, hepatoprotective, neuroprotective, antiviral, etc.). Some also have immunomodulatory activities. Astragalus glycyphyllos is distributed in Bulgaria and mainly accumulates cycloartane saponins. From the overground parts of the species, a triterpenoid cyloartane-type saponin (AGOS3) was isolated by different chromatographic techniques. A quantitative LC-MS method for the determination of the saponin was developed and validated. Further, the saponin was loaded in copolymeric micelles based on triblock copolymers of polyethylene oxide and polypropylene oxide (Pluronics). The LC-MS method was applied on the developed micelles to determine their loading degrees. Afterwards, the possible pharmacological effects of free and encapsulated in polymeric nanoparticles of triterpenoid saponin (1, 5, 10, 25, 50, and 100 µg/mL) were evaluated in isolated murine macrophages and lymphocytes in vitro. Free AGOS3 stimulated proliferation only at the highest tested concentrations (50–100 µg/mL), and the effect was more evident in isolated macrophages. Interestingly, AGOS3-loaded polymeric micelles caused concentration dependency and statistically significant increases in the proliferation of both isolated lymphocytes and macrophages, even at a lower concentration (10 µg/mL). These results could serve as the basis for further research on the immunomodulatory effect of this saponin. Full article

Background: Historically used as a marker for inherited disorders, the current interest in plasma homocysteine measurement lies in its ability to provide valuable information about the metabolic and nutritional status of patients. Specifically, nitrous oxide (N₂O) abuse can lead to functional vitamin B12 deficiency by oxidation and increase oxidative stress, resulting in elevated plasma homocysteine levels, which mimic neurological conditions such as Guillain–Barré syndrome. Rapid identification of hyperhomocysteinemia is crucial for timely intervention and avoiding costly, unnecessary treatments. Objective: This study evaluates the performance of a rapid immunoassay technique (Snibe) compared to mass spectrometry (LC-MS/MS) for measuring plasma homocysteine levels in patients with nitrous oxide abuse and non-inherited caused of elevated homocysteine, aiming to enhance differential diagnosis related to oxidative stress. Methods: 235 patients from Lille University Hospital were included. EDTA blood samples were collected and analyzed using both rapid immunoassay (Snibe) and LC-MS/MS. Neurological assessment was performed using the peripheral neuropathy disability (PND) score. Results: Firstly, significant elevations in plasma homocysteine levels were observed in patients abusing nitrous oxide measured by LC-MS/MS. Secondly, the immunoassay provided rapid results, essential for early clinical decision-making, but tended to underestimate high values compared to LC-MS/MS. A good correlation was found between the methods for low and moderate values. Conclusion: The immunoassay tended to underestimate high-value samples compared to LC-MS/MS, which is a common problem with the competitive methodology. The rapid immunoassay technique is effective for initial screening and early intervention, aiding in the differential diagnosis of conditions related to oxidative stress. Therefore, it is recommended to use the CLIA method for initial screening and confirm with mass spectrometry if there are abnormal samples. Integrating both techniques can enhance diagnostic accuracy and improve patient outcomes. Full article

Therapeutic drug monitoring (TDM) may be beneficial for cyclin-dependent kinase 4/6 inhibitors (CDK4/6is), such as palbociclib, ribociclib, and abemaciclib, due to established exposure–toxicity relationships and the potential for monitoring treatment adherence. Developing a method for quantifying CDK4/6is, abemaciclib metabolites (M2, M20), and letrozole in dried blood spots (DBS) could be useful to enhance the feasibility of TDM. Thus, an optimized LC-MS/MS method was developed using the HemaXis DB10 device for volumetric (10 µL) DBS collection. Chromatographic separation was achieved using a reversed-phase XBridge BEH C18 column. Detection was performed with a triple quadrupole mass spectrometer, utilizing ESI source switching between negative and positive ionization modes and multiple reaction monitoring acquisition. Analytical validation followed FDA, EMA, and IATDMCT guidelines, demonstrating high selectivity, adequate sensitivity (LLOQ S/N ≥ 30), and linearity (r ≥ 0.997). Accuracy and precision met acceptance criteria (between-run: accuracy 95–106%, CV ≤ 10.6%). Haematocrit independence was confirmed (22–55%),with high recovery rates (81–93%) and minimal matrix effects (ME 0.9–1.1%). The stability of analytes under home-sampling conditions was also verified. Clinical validation supports DBS-based TDM as feasible, with conversion models developed for estimating plasma concentrations (the reference for TDM target values) of letrozole, abemaciclib, and its metabolites. Preliminary data for palbociclib and ribociclib are also presented. Full article

Building extensive drug candidate libraries as early in the development pipeline as possible, with high-throughput in vitro absorption, distribution, metabolism, and excretion (ADME) profiling, is crucial for the selection of lead compounds to guide subsequent research and production phases. Traditionally, the analysis of metabolic stability assays heavily relies on high-throughput LC-MS/MS (liquid chromatography tandem mass spectrometry) techniques to meet with the lead profiling demands. Laser-assisted rapid evaporative ionization mass spectrometry (LA-REIMS) is a quick and efficient technique for characterizing complex biological samples without laborious sample preparation. In this study, using an automated LA-REIMS well plate reader, achieving an 8 s per sample measurement time, the oxidative metabolic stability of active drug agents was assessed using biomimetic metalloporphyrin-based oxidative model reactions. The results obtained using the novel LA-REIMS-based protocol were compared to and corroborated by those obtained using conventional HPLC-UV-MS (high performance liquid chromatography with ultra-violet detection coupled with mass spectrometry) measurements. Full article

Rifaximin and rifampicin are good broad-spectrum antimicrobials. The irrational use of antimicrobial drugs in veterinary clinics could threaten public health and food safety. It is necessary to develop a reliable detection method of the residue for enhancing the rational supervision of the use of such drugs, reducing and slowing down the generation of bacterial resistance, and promoting animal food safety and human health. So, this study developed an LC-MS/MS method for the detection of rifaximin and rifampicin residues in animal-origin foods. The residual rifaximin and rifampicin of homogenized test materials were extracted with acetonitrile-dichloromethane solution or acetonitrile in the presence of anhydrous sodium sulfate and vitamin C, purified by dispersible solid phase extraction, determined by LC-MS/MS, and quantified by the internal standard method. The specificity, sensitivity, matrix effect, accuracy, and precision of the method were investigated in the edible tissues of cattle, swine, or chicken. In addition, the stability of the standard stock solution and the standard working solution was also investigated. The method was suitable for the muscle, liver, kidney, fat, milk, and eggs of cattle, swine, or chicken, as well as fish and shrimp. The specificity of the method was good, and the detection of the analytes was not affected by different matrices. Both the LOD and LOQ of the two analytes were 5 μg/kg and 10 μg/kg, respectively. The results of matrix effects in each tissue were in the range of 80–120%; there were no significant matrix effects. The average accuracy of rifaximin and rifampicin in different foodstuffs of animal origin was between 80% and 120%, and the method precision was below 20% (RSD). The proposed method showed good performance for determination, which could be employed for the extraction, purification, and detection of residual rifaximin and rifampicin in edible animal tissues. The pretreatment procedure of tissue samples was simple and feasible. The method was highly specific, stable, reliable, and with high sensitivity, accuracy, and precision, which met the requirements of quantitative detection of veterinary drug residues. Full article

The floating freshwater fern Azolla is the only plant that retains an endocyanobiont, Nostoc azollae (aka Anabaena azollae), during its sexual and asexual reproduction. The increased interest in Azolla as a potential source of food and its unique evolutionary history have raised questions about its cyanotoxin content and genome. Cyanotoxins are potent toxins synthesized by cyanobacteria which have an anti-herbivore effect but have also been linked to neurodegenerative disorders including Alzheimer’s and Parkinson’s diseases, liver and kidney failure, muscle paralysis, and other severe health issues. In this study, we investigated 48 accessions of Azolla–Nostoc symbiosis for the presence of genes coding microcystin, nodularin, cylindrospermopsin and saxitoxin, and BLAST analysis for anatoxin-a. We also investigated the presence of the neurotoxin β-N-methylamino-L-alanine (BMAA) in Azolla and N. azollae through LC-MS/MS. The PCR amplification of saxitoxin, cylindrospermospin, microcystin, and nodularin genes showed that Azolla and its cyanobiont N. azollae do not have the genes to synthesize these cyanotoxins. Additionally, the matching of the anatoxin-a gene to the sequenced N. azollae genome does not indicate the presence of the anatoxin-a gene. The LC-MS/MS analysis showed that BMAA and its isomers AEG and DAB are absent from Azolla and Nostoc azollae. Azolla therefore has the potential to safely feed millions of people due to its rapid growth while free-floating on shallow fresh water without the need for nitrogen fertilizers. Full article

The emergence of multidrug-resistant fungi Candida auris is a worldwide health crisis connected with high rates of mortality. There is a critical need to find novel and unique antifungal compounds for treating infections of multidrug-resistant fungi such as C. auris. This study aimed to illustrate that biosynthetic gene clusters in native bacterial isolates are able to produce antifungal compounds against the multidrug-resistant fungus C. auris. It was successfully achieved using large-scale antifungal activity screening, cytotoxicity analysis, and whole genome sequencing integrated with genome mining-guided analysis and liquid chromatography–mass spectrometry (LC/MS). A list of possible gene candidates was initially identified with genome mining methods to predict secondary metabolite gene clusters of antifungal-compound-producing bacteria. Then, gene clusters present in the antifungal-compound-producing bacteria were identified and aligned with the reference genome using comparative genomic approaches. Bacillus halotolerans AQ11M9 was identified through large-scale antifungal activity screening as a natural compound-producer against multidrug-resistant C. auris, while it was nontoxic to normal human skin fibroblast cells (confirmed using a cell viability assay). The genome (4,197,347 bp) of B. halotolerans AQ11M9 with 2931 predicted genes was first mined for detecting and characterizing biosynthetic gene clusters, which revealed 10 candidate regions with antifungal activity. Clusters of AQ11M9 encoded non-ribosomal peptide synthase (NRPS) (bacilysin, bacillibactin, paenibactin, surfactin, plipastin, and fengycin) and polyketide (macrobrevin). The presence of gene clusters with anti-C. auris activity, and surfactin identified through LC/MS, from AQ11M9 suggests the potential of utilizing it as a source for a novel and powerful anti-C. auris compound. Full article

Background/Objectives: Marrubium vulgare L. (M. vulgare), the white horehound, is well known for treating inflammation-related diseases. Methods: In this context, we investigated the efficacy of M. vulgare ingredients in treating Alzheimer’s disease using various in vitro and in silico antioxidant, anti-inflammatory, anti-cholinesterase, and anti-tyrosinase mechanisms. Results: In our results, sixty-one components were tentatively identified using gas and liquid chromatography (GC-MS and LC-MSⁿ) and categorized as hydrocarbons, fatty acids, and polyphenolics. The extract inhibited linoleic oxidation with an IC₅₀ value of 114.72 µg/mL, captured iron (Fe²⁺) ions with an IC₅₀ value of 164.19 µg/mL, and displayed reducing power. In addition, the extract showed radical-scavenging ability towards DPPH^•, NO^•, ABTS^•+, and H₂O₂ assays compared to L-ascorbic acid and butylated hydroxytoluene. The DPPH^• was scavenged by 77.62% at 100 µg/mL, and NO^•, ABTS^•+, and H₂O₂ were scavenged with IC₅₀ values of 531.66, 117.51, and 143.10 µg/mL, respectively. M. vulgare also exhibited discriminating anti-inflammatory potency against cyclooxygenase (COX-2) with IC₅₀ values of 619.15 µg/mL compared to celecoxib (p > 0.05). Notably, three Alzheimer’s biomarkers, acetylcholinesterase (AChE), butyrylcholinesterase (BChE), and tyrosinase were significantly inhibited. The molecular docking study supposed that the phenylethanoid glycosides of samioside and forsythoside B inhibited AChE and tyrosinase enzymes with low binding affinities of −9.969 and −8.804 kcal/mol, respectively. Marruboside was a proper inhibitor of COX and BChE enzymes with a binding score of −10.218 and −10.306 kcal/mol, respectively. Conclusions: M. vulgare extract showed significant inhibitory actions, which suggest that it could have a promising potential as an anti-Alzheimer agent. Full article

Background/Objectives: Human milk is the optimal source of nutrition and protection against infection for infants. If breastfeeding is not possible, standard and hydrolyzed infant formulas (IF) are an alternative. Extensively hydrolyzed IFs (eHFs) contain bioactive peptides, but their activities have rarely been evaluated. The aim of this study was to characterize and compare the bioactive peptide profiles of different eHFs and standard IFs before and after in vitro digestion. Methods: Two forms, liquid and powder, of intact protein formula (iPF) and eHF were subjected to in vitro gastrointestinal digestion, mimicking a young infant’s gut (age 0–4 months) and an older infant’s gut (>6 months). Bioactive peptides of in vitro digested and undigested formulas were analysed with Liquid Chromatography–Mass Spectrometry (LC–MS). Results: In all samples, a variety of peptides with potential bioactive properties were found. Immuno-regulatory peptides, followed by antimicrobial and antioxidative peptides were most frequent, as were peptides promoting wound healing, increasing mucin secretion, regulating cholesterol metabolism, and preventing bacterial infection. Peptides typically found in yoghurt and colostrum were identified in some formula samples. Conclusions: The high amounts of bioactive peptides with various properties in eHFs and iPFs indicate a possible contribution to infection protection, healthy gut microbiomes, and immunological development of infants. eHFs showed similar compositions of bioactive peptides to iPFs, with intermittently increased peptide variety and quantity. Full article

A fast and sample cleanup approach for fluoxetine in human plasma was developed using protein precipitation coupled with LC–MS-MS. Samples were treated with methanol prior to LC–MS-MS analysis. Chromatographic separation was performed on a reverse phase column with an isocratic mobile phase of methanol and 10 mM ammonium formate pH acidified with formic acid (80:20, v/v) at a flow rate of 0.2 mL/min. The run time was 4 min. Mass parameters were optimized to monitor transitions at m/z [M + H]⁺ 310 > > 148 for fluoxetine and m/z [M + H]⁺ 315.1 > > 153 for fluoxetine-d5 as an internal standard. The lower limit of quantification and the dynamic range were 0.25 and 0.25–50 ng/mL, respectively. Linearity was good for intra-day and inter-day validations (R² = 0.999). The matrix effect was acceptable with CV% < 15 and accuracy% < 15. The hemolytic effect was negligible. Fluoxetine was stable in human plasma for 48 h at room temperature (25 °C), for 12 months frozen at −25 °C, for 48 h in an auto-sampler at 6 °C, and for three freeze/thaw cycles. The validated method was applied in a pharmacokinetic study to determine the concentration of fluoxetine in plasma samples. The study provides a fast and simple bioanalytical method for routine analysis and may be particularly useful for bioequivalence studies. The method was successfully applied to a pharmacokinetic study of fixed-dose fluoxetine in nine healthy volunteers. Full article

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. Full article