Harnessing alkaline-pH regulatable promoters for efficient methanol-free expression of enzymes of industrial interest in Komagataella Phaffii.

BACKGROUND: The yeast Komagataella phaffii has become a very popular host for heterologous protein expression, very often based on the use of the AOX1 promoter, which becomes activated when cells are grown with methanol as a carbon source. However, the use of methanol in industrial settings is not devoid of problems, and therefore, the search for alternative expression methods has become a priority in the last few years. RESULTS: We recently reported that moderate alkalinization of the medium triggers a fast and wide transcriptional response in K. phaffii. Here, we present the utilization of three alkaline pH-responsive promoters (pTSA1, pHSP12 and pPHO89) to drive the expression of a secreted phytase enzyme by simply shifting the pH of the medium to 8.0. These promoters offer a wide range of strengths, and the production of phytase could be modulated by adjusting the pH to specific values. The TSA1 and PHO89 promoters offered exquisite regulation, with virtually no enzyme production at acidic pH, while limitation of Pi in the medium further potentiated alkaline pH-driven phytase expression from the PHO89 promoter. An evolved strain based on this promoter was able to produce twice as much phytase as the reference pAOX1-based strain. Functional mapping of the TSA1 and HSP12 promoters suggests that both contain at least two alkaline pH-sensitive regulatory regions. CONCLUSIONS: Our work shows that the use of alkaline pH-regulatable promoters could be a useful alternative to methanol-based expression systems, offering advantages in terms of simplicity, safety and economy.

Antioxidant activity and Inhibitory effects of Cydonia oblonga Miller. leaves extracts against calcium oxalate stones.

Urinary stone disease is a widespread health problem in both adults and children, and its prevalence has been increasing worldwide. Various plants preparations have already been used since ancient times in order to treat urolithiasis. The aim of this study is to evaluate the antioxidant capacity and litholytic effect on kidney stones of Cydonia oblonga Miller. leaves. The infusion, methanol and acetone extracts were made from Cydonia oblonga Miller. leaf at different concentration. Estimation of mass fractions of total polyphenol, flavonoid, and flavonol contents, as well as the in vitro radical scavenging potential on 2,2'-diphenyl-1-picrylhydrazyl radical (DPPH·) of the investigated extracts was carried out using colorimetric methods. The litholytic property of the extracts was performed by an in-vitro model using experimentally prepared kidney stones- calcium oxalate. As results, the quince leaf extracts revealed stronger antioxidant properties in the DPPH assay, which was proved by the semi-maximal inhibitory concentration values, being about 36.06 ± 3.55, 74.15 ± 6.29, and 142.35 ± 5.09 µg/ml for methanol, acetone and infusion extracts respectively. Furthermore, the tested extracts were found to be more effective in dissolving calcium oxalate stones compared to the control solutions, the mass loss is about 15.13 ± 1.10% with methanol extract, while it is 14.77 ± 1.74% and 11.14 ± 2.86% for acetone and infusion extracts respectively. These findings confirm the quince leaf's richness in phyto-components, offering anti-oxidant property and being able to be used as a remedy for the management of kidney stones by dissolving calcium oxalate stones in the kidneys.

Employing Piper longum extract for eco-friendly fabrication of PtPd alloy nanoclusters: advancing electrolytic performance of formic acid and methanol oxidation.

Advancement in bioinspired alloy nanomaterials has a crucial impact on fuel cell applications. Here, we report the synthesis of PtPd alloy nanoclusters via the hydrothermal method using Piper longum extract, representing a novel and environmentally friendly approach. Physicochemical characteristics of the synthesized nanoclusters were investigated using various instrumentation techniques, including X-ray photoelectron spectroscopy, X-ray diffraction, and High-Resolution Transmission electron microscopy. The electrocatalytic activity of the biogenic PtPd nanoclusters towards the oxidation of formic acid and methanol was evaluated chronoamperometry and cyclic voltammetry studies. The surface area of the electrocatalyst was determined to be 36.6 m2g-1 by Electrochemical Surface Area (ECSA) analysis. The biologically inspired PtPd alloy nanoclusters exhibited significantly higher electrocatalytic activity compared to commercial Pt/C, with specific current responses of 0.24 mA cm - 2 and 0.17 mA cm - 2 at synthesis temperatures of 180 °C and 200 °C, respectively, representing approximately four times higher oxidation current after 120 min. This innovative synthesis approach offers a promising pathway for the development of PtPd alloy nanoclusters with enhanced electrocatalytic activity, thereby advancing fuel cell technology towards a sustainable energy solution.

Comparing methanol and glycerol as carbon sources for mainstream partial denitrification/anammox in an IFAS process.

This study explored the implementation of mainstream partial denitrification with anammox (PdNA) in the second anoxic zone of a wastewater treatment process in an integrated fixed film activated sludge (IFAS) configuration. A pilot study was conducted to compare the use of methanol and glycerol as external carbon sources for an IFAS PdNA startup, with a goal to optimize nitrogen removal while minimizing carbon usage. The study also investigated the establishment of anammox bacteria on virgin carriers in IFAS reactors without the use of seeding, and it is the first IFAS PdNA startup to use methanol as an external carbon source. The establishment of anammox bacteria was confirmed in both reactors 102 days after startup. Although the glycerol-fed reactor achieved a higher steady-state maximum ammonia removal rate because of anammox bacteria (1.6 ± 0.3 g/m2/day) in comparison with the methanol-fed reactor (1.2 ± 0.2 g/m2/day), both the glycerol- and methanol-fed reactors achieved similar average in situ ammonia removal rates of 0.39 ± 0.2 g/m2/day and 0.40 ± 0.2 g/m2/day, respectively. Additionally, when the upstream ammonia versus NOx (AvN) control system maintained an ideal ratio of 0.40-0.50 g/g, the methanol-fed reactor attained a lower average effluent TIN concentration (3.50 ± 1.2 mg/L) than the glycerol-fed reactor (4.43 ± 1.6 mg/L), which was prone to elevated nitrite concentrations in the effluent. Overall, this research highlights the potential for PdNA in IFAS configurations as an efficient and cost-saving method for wastewater treatment, with methanol as a viable carbon source for the establishment of anammox bacteria. PRACTITIONER POINTS: Methanol is an effective external carbon source for an anammox startup that avoids the need for costly alternative carbon sources. The methanol-fed reactor demonstrated higher TIN removal compared with the glycerol-fed reactor because of less overproduction of nitrite. Anammox bacteria was established in an IFAS reactor without seeding and used internally stored carbon to reduce external carbon addition. Controlling the influent ammonia versus NOx (AvN) ratio between 0.40 and 0.50 g/g allowed for low and stable TIN effluent conditions.

Phytochemical characterization, antimicrobial and antioxidant activities of Terminalia catappa methanol and aqueous extracts.

BACKGROUND: A study carried out by World Health Organization revealed that around 80% of individuals globally depends on herbal forms of medication with 40% of pharmaceutical products being sourced from medicinal plants. The study objective was to evaluate the phytochemicals composition, in vitro antimicrobial and antioxidant properties of the leaves of Terminalia catappa L. aqueous and methanolic extracts. METHODS: Antimicrobial activity was analyzed by disk diffusion, the minimum inhibitory concentration in-vitro assays with ciprofloxacin as the standard for antibacterial assay while nystatin for antifungal assay. Ferric reducing antioxidant power and 2,2-diphenyl-1-picryl-hydrazyl-hydrate assays were used for the evaluation of antioxidant properties of the crude extracts while the groups responsible for this activity identified using Fourier transform infrared spectrophotometer. RESULTS: The study found that the leaves of Terminalia catappa contained alkaloids, tannins, steroids, cardiac glycosides, flavonoids, phenols, saponins, and coumarins, but terpenoids were absent. Presence of functional groups associated with this class of compounds such as OH vibrational frequencies were observed in IR spectrum of the crude extracts. Methanolic extract from Terminalia catappa exhibited greater antibacterial properties against Pseudomonas aeruginosa, Escherichia coli and Staphylococcus aureus, whereas aqueous extract displayed greater antibacterial activity against Bacillus subtilis for all concentrations tested. The amount of the sample that scavenged 50 percent of DPPH (IC50) was found to be 8.723, 13.42 and 13.04 µg/mL for L-ascorbic acid, Terminalia catappa L. methanolic and aqueous extracts respectively. The antimicrobial and antioxidant activities varied with the extract concentration and solvent used in extractions. CONCLUSION: Terminalia catappa L. leaves are prospective for use as a source of therapeutic agents that could lead to the advancement of new antimicrobial and antioxidant products.

Genome-resolved metagenomics identifies novel active microbes in biogeochemical cycling within methanol-enriched soil.

Metagenome assembled genomes (MAGs), generated from sequenced 13C-labelled DNA from 13C-methanol enriched soils, were binned using an ensemble approach. This method produced a significantly larger number of higher-quality MAGs compared to direct binning approaches. These MAGs represent both the primary methanol utilizers and the secondary utilizers labelled via cross-feeding and predation on the labelled methylotrophs, including numerous uncultivated taxa. Analysis of these MAGs enabled the identification of multiple metabolic pathways within these active taxa that have climatic relevance relating to nitrogen, sulfur and trace gas metabolism. This includes denitrification, dissimilatory nitrate reduction to ammonium, ammonia oxidation and metabolism of organic sulfur species. The binning of viral sequence data also yielded extensive viral MAGs, identifying active viral replication by both lytic and lysogenic phages within the methanol-enriched soils. These MAGs represent a valuable resource for characterizing biogeochemical cycling within terrestrial environments.

Neuroprotective effects of methanolic extract from Chuanxiong Rhizoma in mice with middle cerebral artery occlusion-induced ischemic stroke: suppression of astrocyte- and microglia-related inflammatory response.

BACKGROUND: In traditional Asian medicine, dried rhizomes of Ligusticum chuanxiong Hort. (Chuanxiong Rhizoma [CR]) have long been used to treat pain disorders that affect the head and face such as headaches. Furthermore, they have been used primarily for blood circulation improvement or as an analgesic and anti-inflammatory medicine. This study aimed to investigate the neuroprotective effects of a methanol extract of CR (CRex) on ischemic stroke in mice caused by middle cerebral artery occlusion (MCAO). METHODS: C57BL/6 mice were given a 1.5-h transient MCAO (MCAO control and CRex groups); CRex was administered in the mice of the CRex group at 1,000-3,000 mg/kg either once (single dose) or twice (twice dose) before MCAO. The mechanism behind the neuroprotective effects of CRex was examined using the following techniques: brain infarction volume, edema, neurological deficit, novel object recognition test (NORT), forepaw grip strength, and immuno-fluorescence staining. RESULTS: Pretreating the mice with CRex once at 1,000 or 3,000 mg/kg and twice at 1,000 mg/kg 1 h before MCAO, brought about a significantly decrease in the infarction volumes. Furthermore, pretreating mice with CRex once at 3,000 mg/kg 1 h before MCAO significantly suppressed the reduction of forepaw grip strength of MCAO-induced mice. In the MCAO-induced group, preadministration of CRex inhibited the reduction in the discrimination ratio brought on by MCAO in a similar manner. CRex exhibited these effects by suppressing the activation of astrocytes and microglia, which regulated the inflammatory response. CONCLUSIONS: This study proposes a novel development for the treatment of ischemic stroke and provides evidence favoring the use of L. chuanxiong rhizomes against ischemic stroke.

Biogenic Zinc Oxide Nanoparticles synthesized from Tinospora Cordifolia induce oxidative stress, mitochondrial damage and apoptosis in Colorectal Cancer.

Cancer chemotherapy remains a serious challenge, and new approaches to therapy are urgently needed to build novel treatment regimens. The methanol extract of the stem of Tinospora Cordifolia was used to synthesize biogenic zinc oxide nanoparticles (ZnO-NPs) that display anticancer activities against colorectal cancer. Biogenic ZnO-NPs synthesized from methanol extract of Tinospora Cordifolia stem (ZnO-NPs TM) were tested against HCT-116 cell lines to assess anticancer activity. UV-Vis, FTIR, XRD, SEM, and TEM analysis characterized the biogenic ZnO-NPs. To see how well biogenic ZnO-NPs fight cancer, cytotoxicity, AO/EtBr staining, Annexin V/PI staining, mitochondrial membrane potential (MMP), generation of reactive oxygen species (ROS) analysis, and caspase cascade activity analysis were performed to assess the anticancer efficacy of biogenic ZnO-NPs. The IC50 values of biogenic ZnO-NPs treated cells (HCT-116 and Caco-2) were 31.419 ± 0.682µg/ml and 36.675 ± 0.916µg/ml, respectively. qRT-PCR analysis showed that cells treated with biogenic ZnO-NPs Bax and P53 mRNA levels increased significantly (p ≤ 0.001). It showed to have impaired MMP and increased ROS generation. In a corollary, our in vivo study showed that biogenic ZnO-NPs have an anti-tumour effect. Biogenic ZnO-NPs TM showed both in vitro and in vivo anticancer effects that could be employed as anticancer drugs.

In Silico and Chromatographic Methods for Analysis of Biotransformation of Prospective Neuroprotective Pyrrole-Based Hydrazone in Isolated Rat Hepatocytes.

In the current study, chromatographic and in silico techniques were applied to investigate the biotransformation of ethyl 5-(4-bromophenyl)-1-(2-(2-(2-hydroxybenzylidene) hydrazinyl)-2-oxoethyl)-2-methyl-1H-pyrrole-3-carboxylate (11b) in hepatocytic media. The initial chromatographic procedure was based on the employment of the conventional octadecyl stationary phase method for estimation of the chemical stability. Subsequently, a novel and rapid chromatographic approach based on a phenyl-hexyl column was developed, aiming to separate the possible metabolites. Both methods were performed on a Dionex 3000 ThermoScientific (ACM 2, Sofia, Bulgaria) device equipped with a diode array detector set up at 272 and 279 nm for analytes detection. An acetonitrile: phosphate buffer of pH 3.5: methanol (17:30:53 v/v/v) was eluted isocratically as a mobile phase with a 1 mL/min flow rate. A preliminary purification from the biological media was achieved by protein precipitation with methanol. A validation procedure was carried out, where the method was found to correspond to all ICH (Q2) and M10 set criteria. Additionally, an in silico-based approach with the online server BioTransformer 3.0 was applied in an attempt to predict the possible metabolites of the title compound 11b. It was hypothesized that four CYP450 isoforms (1A2, 2C9, 3A4, and 2C8) were involved in the phase I metabolism, resulting in the formation of 12 metabolites. Moreover, docking studies were conducted to evaluate the formation of stable complexes between 11b and the aforementioned isoforms. The obtained data indicated three metabolites as the most probable products, two of which (M9_11b and M10_11b) were synthesized by a classical approach for verification. Finally, liquid chromatography with a mass detector was implemented for comprehensive and summarized analysis, and the obtained results revealed that the metabolism of the 11b proceeds possibly with the formation of glucuronide and glycine conjugate of M11_11b.

Development and Validation of a High-Performance Liquid Chromatography Method to Quantify Marker Compounds in Lysimachia vulgaris var. davurica and Its Effects in Diarrhea-Predominant Irritable Bowel Syndrome.

Irritable bowel syndrome (IBS), a common gastrointestinal disorder worldwide, is characterized by chronic abdominal pain, bloating, and disordered defecation. IBS is associated with several factors, including visceral hypersensitivity, gut motility, and gut-brain interaction disorders. Because currently available pharmacological treatments cannot adequately improve symptoms and may cause adverse effects, the use of herbal therapies for managing IBS is increasing. Lysimachia vulgaris var. davurica (LV) is a medicinal plant used in traditional medicine to treat diarrhea. However, information on whether LV can effectively improve diarrhea-predominant IBS (IBS-D) remains limited. In this study, using an experimental mouse model of IBS-D, we elucidated the effects of the LV extract. The methanol extract of LV decreased fecal pellet output in the restraint stress- or 5-hydroxytryptamine (5-HT)-induced IBS mouse model and inhibited 5-HT-mediated [Ca2+]i increase in a dose-dependent manner. Furthermore, we developed and validated a high-performance liquid chromatography method using two marker compounds, namely, chlorogenic acid and rutin, for quality control analysis. Our study results suggest the feasibility of the methanol extract of LV for developing therapeutic agents to treat IBS-D by acting as a 5-HT3 receptor antagonist.

Clinical presentation and management of methanol poisoning outbreaks in Riyadh, Saudi Arabia: a retrospective analysis.

BACKGROUND: Acute methanol intoxication, whether unintentional or deliberate, necessitates prompt intervention to prevent severe morbidity and mortality. Homemade alcoholic beverages are a frequent source of such poisoning. This retrospective analysis examined two outbreaks of methanol intoxication in Saudi Arabia. It investigated the clinical presentation, implemented management strategies, and any lasting complications (sequelae) associated with these cases. The aim was to assess the potential impact of different treatment modalities and the timeliness of their initiation on patient outcomes. METHODS: This was a retrospective case series of methanol poisoning cases which presented to the adult emergency department (ED) at King Abdulaziz Medical City (KAMC) in Riyadh, Saudi Arabia. There were two separate outbreaks in the city, the first one was from September 1 to September 10, 2020 and the second one was from May 14 to May 20, 2021. Electronic charts were reviewed, and data were extracted to previously prepared data extraction sheets. RESULT: From the 22 patients who arrived in the ED alive, the most common complaints were nausea or vomiting followed by altered level of consciousness. About 9% from the patient were hypotensive, 36% were tachycardic, 41% were tachypneic and 4% were having SpO2 < 94%. Brain CT was abnormal in 6 patients. Vision impairment was the most common sequalae of methanol poisoning (7 out of 12 patients who were assessed by ophthalmologist, 58%). When the patients were divided based on severity (mild, moderate, severe), nausea or vomiting and loss of consciousness were the most common complaints among the moderate group while loss of consciousness predominated in the severe group. Two patients presented with low blood pressure and were in the sever group. The severe group had a mean Glasgow Coma Scale (GCS) of 8. Most of the patients in the severity groups underwent the same management apart from those who died or deposited. Eight patients in the severe group had to be intubated. CONCLUSION: This study demonstrates the multifaceted clinical presentation of methanol poisoning, culminating in a 17.4% mortality rate. Notably, our findings emphasize the critical role of prompt diagnosis and swift initiation of combined fomepizole therapy and hemodialysis in mitigating mortality and minimizing the potential for chronic visual sequelae associated with methanol poisoning.

Biodiesel Production by Methanolysis of Rapeseed Oil-Influence of SiO2/Al2O3 Ratio in BEA Zeolite Structure on Physicochemical and Catalytic Properties of Zeolite Systems with Alkaline Earth Oxides (MgO, CaO, SrO).

Alkaline earth metal oxide (MgO, CaO, SrO) catalysts supported on BEA zeolite were prepared by a wet impregnation method and tested in the transesterification reaction of rapeseed oil with methanol towards the formation of biodiesel (FAMEs-fatty acid methyl esters). To assess the influence of the SiO2/Al2O3 ratio on the catalytic activity in the tested reaction, a BEA zeolite carrier material with different Si/Al ratios was used. The prepared catalysts were tested in the transesterification reaction at temperatures of 180 °C and 220 °C using a molar ratio of methanol/oil reagents of 9:1. The transesterification process was carried out for 2 h with the catalyst mass of 0.5 g. The oil conversion value and efficiency towards FAME formation were determined using the HPLC technique. The physicochemical properties of the catalysts were determined using the following research techniques: CO2-TPD, XRD, BET, FTIR, and SEM-EDS. The results of the catalytic activity showed that higher activity in the tested process was confirmed for the catalysts supported on the BEA zeolite characterized by the highest silica/alumina ratio for the reaction carried out at a temperature of 220 °C. The most active zeolite catalyst was the 10% CaO/BEA system (Si/Al = 300), which showed the highest triglyceride (TG) conversion of 90.5% and the second highest FAME yield of 94.6% in the transesterification reaction carried out at 220 °C. The high activity of this system is associated with its alkalinity, high value of the specific surface area, the size of the active phase crystallites, and its characteristic sorption properties in relation to methanol.

Antibacterial Activities of Methanolic Seeds Extract of Black pepper (Piper nigrum L.) against Gram Positive Staphylococcus aureus & Gram-Negative Escherichia coli.

Evaluation of the in vitro antibacterial activity of Methanolic extracts isolated from Black pepper seeds (Piper nigrum L.) against two infection causing pathogens, Gram-positive Staphylococcus aureus and Gram-negative Escherichia coli. Between July 2022 and June 2023, this experimental study was conducted at the Mymensingh Medical College's Department of Pharmacology and Therapeutics in conjunction with the Department of Microbiology. Using the disc diffusion and broth dilution methods, the antibacterial activity of methanolic extract of black pepper seeds (MBPE) was evaluated at various doses. The solvents Methanol and 10.0% Di Methyl Sulfoxide (DMSO) were used to make the extract. Using the broth dilution procedure, the conventional antibiotic Ciprofloxacin was utilized and the outcome was contrasted with that of Methanol extracts. Methanolic extract of black pepper seeds (MBPE) at seven distinct concentrations (100, 80, 60, 40, 20, 10 and 5 mg/ml) were utilized, then later in chosen concentrations as needed to confirm the extracts' more precise margin of antimicrobial sensitivity. At 80 mg/ml and above doses of the MBPE, it had an inhibitory impact against the aforementioned microorganisms. For Staphylococcus aureus and Escherichia coli the MIC were 60 and 75 mg/ml in MBPE respectively. As of the MIC of Ciprofloxacin was 1µg/ml against Staphylococcus aureus and Escherichia coli. In comparison to MICs of MBPE for the test organisms, the MIC of Ciprofloxacin was the lowest. This study clearly shows that Staphylococcus aureus and Escherichia coli are sensitive to the methanolic extract of black pepper seeds' antibacterial properties.

In vitro anticancer activity of Hirudinaria manillensis methanolic extract and its validation using in silico molecular docking approach.

Cancer has emerged as a potentially lethal illness, which recently upsurged in the mortality rate. Animal-derived compounds could be promising targets with higher efficacy and low toxicity in anticancer therapy. The present study aimed to explore the presence of anticancer potential compounds in Hirudinaria manillensis methanolic extract and their anticancer potential against various cancer cell types and target identification by Auto dock method. Initially, the identification of bioactive compounds was achieved by GC-MS analysis followed by the anticancer activity by MTT assay against A549, HeLa, MDA-MB-231, MG-63, and MOLT-4. Further, the effect of a lead compound on the cancer cell target was analyzed by the Auto dock method. GC-MS analysis results revealed the presence of 25 different bioactive compounds including anticancer potential compounds, such as Lupeol, Carvacrol, and Demecolcine. Interestingly, MTT assay results demonstrated the anticancer potential of Hirudinaria manillensis extract (LE) against various cancer cell lines, such as A549 (54.60 µg/ml), HeLa (19.93 µg/ml), MDA-MB-231 (20.23 µg/ml), MG-63 (20.04 µg/ml), and MOLT-4 (171.8 µg/ml), respectively. Among these cell types, the maximum inhibition was observed against HeLa with the IC50 concentration of 19.93 µg/ml. Furthermore, Demecolcine compound was docked with the EGFR tyrosine kinase showed the binding affinity of the docked complex was predicted to be - 6.2 kcal/mol. Thus, we conclude that H. manillensis has a significant anticancer effect on human cancer cell lines and could be used as a natural target which paves the way for further studies on biomedical applications in cancer therapeutics.

Emission and optical characteristics of brown carbon in size-segregated particles from three types of Chinese ships.

Brown carbon (BrC) is one of the important light absorption substances that have high light absorption ability under short wavelength light. However, limit studies have focused on the BrC emission from ships. In this study, size-segregated particulate matters (PM) were collected from three different types of ships, light absorption characteristics and size distribution of methanol-soluble BrC and water-soluble BrC in PM from ship exhausts were investigated. Results showed that four-stroke low-power diesel fishing boat (4-LDF) had the highest mass concentrations of methanol-soluble organic carbon (MSOC) and water-soluble organic carbon (WSOC), followed by 2-stroke high-power heavy-fuel-oil vessel (2-HHV), and four-stroke high-power marine-diesel vessel (4-HMV). While 2-HHV had obviously higher light absorption coefficients of methanol-soluble BrC (Abs365,M) and water-soluble BrC (Abs365,W) in unit weight of PM than the other two types of ships. The tested ships presented comparable or higher absorption efficiency of BrC in water extracts (MAE365,W) compared with other BrC emission sources. Majority of BrC was concentrated in fine particles, and the particle size distributions of both Abs365,M and Abs365,W showed bimodal patterns, peaking at 0.43-0.65 µm and 4.7-5.8 µm, respectively. However, different particle size distributions were found for MAE365,M between diesel and heavy fuel oil ships. Besides, different wavelength dependence in particles with different size were also detected. Ship exhaust could be confirmed as a non-ignorable BrC emission source, and complex influencing factor could affect the light absorption characteristics of ship emissions. Particle size should also be considered when light absorption ability of BrC was evaluated.

Mosquitocidal Susceptibility and Non-Target Effects of Tricholoma equestre Mushroom (Agaricomycetes) on the Immature Stages of Aedes aegypti, Anopheles stephensi and Culex quinquefasciatus.

The worldwide scientific community is well aware that mosquitoes are the sole agents responsible for transmitting various dreadful diseases and critical illnesses caused by vector-borne pathogens. The primary objective of this current research was to evaluate the effectiveness of methanol extract from Tricholoma equestre mushroom in controlling the early life stages of Culex quinquefasciatus Say, Anopheles stephensi Liston, and Aedes aegypti (Linnaeus in Hasselquist) mosquitoes. The larvae, pupae and eggs of these mosquitoes were exposed to four different concentrations (62.5 to 500 ppm). After 120 h of treatment, the methanol extract of T. equestre exhibited ovicidal activity ranging from 66% to 80% against the eggs of the treated mosquitoes. It also demonstrated promising larvicidal and pupicidal activity with LC50 values of 216-300 and 230-309 ppm against the early life stages of all three mosquito species. Extensive toxicity studies revealed that the methanol extract from T. equestre had no harmful effects on non-target organisms. The suitability index (SI) or predator safety factor (PSF) indicated that the methanol extract did not harm Poecilia reticulata Peters 1859, (predatory fish), Gambusia affinis S. F. Baird & Girard 1853, dragonfly nymph and Diplonychus indicus Venkatesan & Rao 1871 (water-bug). Gas chromatography-mass spectrometry (GCMS) analysis identified key compounds, including 3-butenenitrile, 2-methyl-(25.319%); 1-butanol, 2-nitro-(18.87%) and oxalic acid, heptyl propyl ester (21.82%) which may be responsible for the observed activity. Furthermore, the formulation based on the methanol extract demonstrated similar effectiveness against all treated mosquitoes at the laboratory level and was found to be non-toxic to mosquito predators. This groundbreaking research represents the first confirmation that methanol extract from T. equestre could be effectively employed in preventing mosquito-borne diseases through mosquito population control programs.

Novel optical coherence tomography findings in a patient with methanol toxicity: a case report.

We report a case of acute methanol toxicity with unique optical coherence tomography findings. A 56-year-old man was referred to our ophthalmology clinic with a history of handmade vodka consumption and vision loss. On ophthalmologic examination, his vision was 20/100 in his right eye and 20/200 in his left eye. Bilateral mild optic disk hyperemia was detected on fundus examination. Because of the severity of systemic symptoms in such cases, it is very difficult to include optical coherence tomography in the ophthalmologic examination. However, we managed to perform optical coherence tomography and recorded shallow subretinal fluid and a prominent middle limiting membrane sign as acute retinal structural changes in the patient. The patient was treated with hemodialysis, intravenous ethanol, and sodium bicarbonate. On the fourth day of treatment, visual acuity improved to 20/20 in both eyes. In addition, the prominent middle limiting membrane sign and subretinal fluid disappeared. In this unusual case, retinal pigment epithelium damage and retinal ischemia may have contributed to the prominent middle limiting membrane and subretinal fluid, which are novel optical coherence tomography findings of methanol toxicity.

[Determination of two perfluorinated compounds in urine by liquid chromatography-tandem mass spectrometry].

Objective: To establish a method for the determination of two perfluorinated compounds in urine by liquid chromatography-tandem mass spectrometry. Methods: In November 2022, urine samples were extracted by acidic methanol, purified by WAX solid phase extraction column, and eluted with methanol water, then Waters ACQUITY UPLC BEH C18 column (100 mm×2.1 mm, 1.7 µm) was used with 1.0 mmol/L ammonium acetate solution and methanol as mobile phase. The gradient elution was carried out, the detection was carried out by electrospray negative ion multiple response monitoring (MRM) mode, and the quantitative method was internal standard method. Results: Perfluorooctanoic acid and perfluorooctane sulfonic acid had a good linear relationship in the concentration range of 0.5-50.0 µg/L, and the correlation coefficient was >0.999. The limit of detection was 0.017 µg/L, and the limit of quantitation was 0.005 µg/L. The average recoveries were 96.3% and 101.8%, respectively. Days of precision were 3.5%-6.2% and 3.1%-7.4%, respectively, daytime precision were 4.3%-6.8% and 4.7%-8.1%, respectively. Conclusion: The established method of liquid chromatography-tandem mass spectrometry is high sensitivity and accuracy, and is suitable for the determination of perfluorooctanoic acid and perfluorooctane sulfonic acid in human urine.

LC-MS/MS, GC-MS and molecular docking analysis for phytochemical fingerprint and bioactivity of Beta vulgaris L.

The plants that we consume in our daily diet and use as a risk preventer against many diseases have many biological and pharmacological activities. In this study, the phytochemical fingerprint and biological activities of Beta vulgaris L. leaf extract, which are widely consumed in the Black Sea region, were investigated. The leaf parts of the plant were dried in an oven at 35 °C and then ground into powder. The main constituents in B. vulgaris were identified by LC-MS/MS and GC-MS analyses. Phenolic content, betaxanthin and betacyanin levels were investigated in the extracts obtained using three different solvents. The biological activity of the extract was investigated by anti-microbial, anti-mutagenic, anti-proliferative and anti-diabetic activity tests. Anti-diabetic activity was investigated by in vitro enzyme inhibition and in-silico molecular docking was performed to confirm this activity. In the LC-MS analysis of B. vulgaris extract, a major proportion of p_coumaric acid, vannilin, protecatechuic aldehyde and sesamol were detected, while the major essential oils determined by GC-MS analysis were hexahydrofarnesyl acetone and phytol. Among the solvents used, the highest extraction efficiency of 2.4% was obtained in methanol extraction, and 36.2 mg of GAE/g phenolic substance, 5.1 mg/L betacyanin and 4.05 mg/L betaxanthin were determined in the methanol extract. Beta vulgaris, which exhibited broad-spectrum anti-microbial activity by forming a zone of inhibition against all tested bacteria, exhibited anti-mutagenic activity in the range of 35.9-61.8% against various chromosomal abnormalities. Beta vulgaris extract, which did not exhibit mutagenic, sub-lethal or lethal effects, exhibited anti-proliferative activity by reducing proliferation in Allium root tip cells by 21.7%. 50 mg/mL B. vulgaris extract caused 58.9% and 55.9% inhibition of α-amylase and α-glucosidase activity, respectively. The interactions of coumaric acid, vanniline, hexahydrofarnesyl acetone and phytol, which are major compounds in phytochemical content, with α-amylase and α-glucosidase were investigated by in silico molecular docking and interactions between molecules via various amino acids were determined. Binding energies between the tested compounds and α-amylase were obtained in the range of - 4.3 kcal/mol and - 6.1 kcal/mol, while for α-glucosidase it was obtained in the range of - 3.7 kcal/mol and - 5.7 kcal/mol. The biological activities of B. vulgaris are closely related to the active compounds it contains, and therefore studies investigating the phytochemical contents of plants are very important. Safe and non-toxic plant extracts can help reduce the risk of various diseases, such as diabetes, and serve as an alternative or complement to current pharmaceutical practices.