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Microbial Cell Factories Sep 2023Methanol, synthesized from CO, is a potentially sustainable one-carbon (C1) resource for biomanufacturing. The use of methanol as a feedstock to produce single cell...
BACKGROUND
Methanol, synthesized from CO, is a potentially sustainable one-carbon (C1) resource for biomanufacturing. The use of methanol as a feedstock to produce single cell protein (SCP) has been investigated for decades as an alternative to alleviate the high global demand for animal-derived proteins. The methylotrophic yeast Pichia pastoris is an ideal host for methanol-based SCP synthesis due to its natural methanol assimilation ability. However, improving methanol utilization, tolerance to higher temperature, and the protein content of P. pastoris are also current challenges, which are of great significance to the economical industrial application using methanol as a feedstock for SCP production.
RESULTS
In the present work, adaptive laboratory evolution (ALE) has been employed to overcome the low methanol utilization efficiency and intolerance to a higher temperature of 33 °C in P. pastoris, associated with reduced carbon loss due to the lessened detoxification of intracellular formaldehyde through the dissimilation pathway and cell wall rearrangement to temperature stress resistance following long-term evolution as revealed by transcriptomic and phenotypic analysis. By strengthening nitrogen metabolism and impairing cell wall synthesis, metabolic engineering further increased protein content. Finally, the engineered strain via multi-strategy produced high levels of SCP from methanol in a pilot-scale fed-batch culture at 33 °C with a biomass of 63.37 g DCW/L, methanol conversion rate of 0.43 g DCW/g, and protein content of 0.506 g/g DCW. SCP obtained from P. pastoris contains a higher percentage of protein compared to conventional foods like soy, fish, meat, whole milk, and is a source of essential amino acids, including methionine, lysine, and branched-chain amino acids (BCAAs: valine, isoleucine, leucine).
CONCLUSIONS
This study clarified the unique mechanism of P. pastoris for efficient methanol utilization, higher temperature resistance, and high protein synthesis, providing a P. pastoris cell factory for SCP production with environmental, economic, and nutritional benefits.
Topics: Animals; Methanol; Pichia; Carbon; Recombinant Proteins
PubMed: 37770920
DOI: 10.1186/s12934-023-02198-9 -
Diseases (Basel, Switzerland) Dec 2023The application of single-cell RNA sequencing has greatly improved our understanding of various cellular and molecular mechanisms involved in physiological and...
INTRODUCTION
The application of single-cell RNA sequencing has greatly improved our understanding of various cellular and molecular mechanisms involved in physiological and pathophysiological processes. However, obtaining living cells for this technique can be difficult under certain conditions. To solve this problem, the methanol fixation method appeared as a promising alternative for routine clinical use.
MATERIALS AND METHODS
In this study, we selected two AML samples that had been fixed in methanol for 12-18 months. Once the cells were rehydrated, these samples were subjected to single-cell RNA sequencing. We then compared the results obtained from these samples with those obtained from the same samples cryopreserved in DMSO.
RESULTS
We used a previously validated methanol fixation protocol to perform scRNA-seq on DMSO cryopreserved cells and cells fixed in methanol for more than one year. Preliminary results show that methanol fixation induces some genetic and transcriptional modification compared with DMSO cryopreservation but remains a valuable method for single-cell analysis of primary human leukemia cells.
CONCLUSIONS
The initial findings from this study highlight certain resemblances in methanol fixation over a 12-month period and cryopreservation with DMSO, along with associated transcriptional level modifications. However, we observed genetic degradation in the fixation condition when extending beyond one year. Despite certain study limitations, it is evident that short-term methanol fixation can be effectively used for leukemia blast samples. Its ease of implementation holds the potential to simplify the integration of this technique into routine clinical practice.
PubMed: 38275564
DOI: 10.3390/diseases12010001 -
Heliyon Sep 2023The current study sought to examine the pharmacological potentials of crude methanolic extracts of and , as well as their various solvent fractionates, with a focus on...
The current study sought to examine the pharmacological potentials of crude methanolic extracts of and , as well as their various solvent fractionates, with a focus on cytotoxic, thrombolytic, membrane stabilizing, antioxidant, and antibacterial activities via and approaches. The extensive chromatographic and spectroscopic analyses confirmed and characterized two compounds as (±)-licarin B () and stigmasterol () from and , respectively. Petroleum ether soluble fraction of and the aqueous soluble fraction of showed the lowest 50% lethal concentrations (1.41 and 1.94 μg/mL, respectively) in brine shrimp bioassay. Likewise, petroleum ether soluble fraction of and aqueous soluble fraction of showed the highest thrombolytic activity with 46.66% and 50.10% lyses of the clot, respectively. The methanol and dichloromethane soluble fractions of reduced erythrocyte hemolysis by 64.03% and 37.08%, respectively, under hypotonic and heat-induced conditions, compared to 81.97% and 42.12% for standard acetylsalicylic acid. In antioxidant activity test, aqueous soluble fraction (IC = 7.22 μg/mL) revealed promising antioxidant potentialities in comparison to standard butylated hydroxytoluene (IC = 21.20 μg/mL). In antibacterial screening, chloroform, and dichloromethane soluble fractions of showed a mild antibacterial activity compared with the standard drug ciprofloxacin. Additionally, the molecular docking study corroborated the current findings, and the isolated two constituents had higher binding affinities toward epidermal growth factor receptor, tissue plasminogen activator, vFLIP-IKK gamma stapled peptide dimer, glutathione reductase, and dihydrofolate reductase enzyme than their corresponding standard drugs. In addition, the both isolated compounds exerted favorable pharmacokinetics (absorption, distribution, metabolism, excretion) and toxicological profiles with drug-like qualities in computational-based ADMET and drug likeliness analyses. The current research suggests that both plants have potential as a natural treatment for treating thrombosis, inflammation, and oxidative stress. However, more thorough research is required to thoroughly screen for phytochemicals and pinpoint the precise mechanisms of action of the bioactive metabolites derived from these plants against a broad range of molecular targets.
PubMed: 37809757
DOI: 10.1016/j.heliyon.2023.e20100 -
ACS Catalysis Aug 2023Low temperature and high pressure are thermodynamically more favorable conditions to achieve high conversion and high methanol selectivity in CO hydrogenation. However,...
Low temperature and high pressure are thermodynamically more favorable conditions to achieve high conversion and high methanol selectivity in CO hydrogenation. However, low-temperature activity is generally very poor due to the sluggish kinetics, and thus, designing highly selective catalysts active below 200 °C is a great challenge in CO-to-methanol conversion. Recently, Re/TiO has been reported as a promising catalyst. We show that Re/TiO is indeed more active in continuous and high-pressure (56 and 331 bar) operations at 125-200 °C compared to an industrial Cu/ZnO/AlO catalyst, which suffers from the formation of methyl formate and its decomposition to carbon monoxide. At lower temperatures, precise understanding and control over the active surface intermediates are crucial to boosting conversion kinetics. This work aims at elucidating the nature of active sites and active species by means of X-ray absorption spectroscopy, Raman spectroscopy, ambient-pressure X-ray photoelectron spectroscopy (AP-XPS), and diffuse reflectance infrared Fourier transform spectroscopy (DRIFTS). Transient DRIFTS studies uncover the activation of CO to form active formate intermediates leading to methanol formation and also active rhenium carbonyl intermediates leading to methane over cationic Re single atoms characterized by rhenium tricarbonyl complexes. The transient techniques enable us to differentiate the active species from the spectator one on TiO support, such as less reactive formate originating from spillover and methoxy from methanol adsorption. The AP-XPS supports the fact that metallic Re species act as H activators, leading to H-spillover and importantly to hydrogenation of the active formate intermediate present over cationic Re species. The origin of the unique reactivity of Re/TiO was suggested as the coexistence of cationic highly dispersed Re including single atoms, driving the formation of monodentate formate, and metallic Re clusters in the vicinity, activating the hydrogenation of the formate to methanol.
PubMed: 37614518
DOI: 10.1021/acscatal.3c01599 -
International Journal of Analytical... 2023Alcohol beverages have been widely consumed in several parts of the world. In this study, volatile organic compounds in alcoholic beverages including acetaldehyde, ethyl...
Alcohol beverages have been widely consumed in several parts of the world. In this study, volatile organic compounds in alcoholic beverages including acetaldehyde, ethyl acetate, methanol, and higher ethanol were investigated and evaluated using a headspace gas chromatograph equipped with a mass spectrometer. This study evaluated the suitability of the chromatographic system, linearity, limit of detection, and limit of quantification, accuracy, and precision of the single and simultaneous determination of acetaldehyde, ethyl acetate, and methanol in thousand folds of ethanol. Results showed that the acetaldehyde concentration in local beer samples and local manual product liqueur samples ranged from 4.65 to 13.22 mg/L and from 5.55 to 75.96 mg/L, respectively, but in local industrial product liqueur samples, acetaldehyde was not detected. Methanol was only detected in a few local beer samples and locally manually produced liqueur samples within low concentrations. Ethyl acetate was only detected in all local beer samples, but it was not present in local industrial product liqueur samples.
PubMed: 37744750
DOI: 10.1155/2023/8851265 -
JACS Au Oct 2023The precise C-C coupling is a challenging goal in C1 chemistry. The conversion of methanol, a cheap and easily available C1 feedstock, into value-added and largely...
The precise C-C coupling is a challenging goal in C1 chemistry. The conversion of methanol, a cheap and easily available C1 feedstock, into value-added and largely demanded olefins has been playing a game-changing role in the production of olefins. The current methanol-to-olefin (MTO) process, however, suffers from limited selectivity to a specific olefin. Here, we present a relay-catalysis route for the high-selective conversion of methanol to ethylene in syngas (H/CO) typically used for methanol synthesis. A bifunctional catalyst composed of selectively dealuminated H-MOR zeolite and ZnO-TiO, which implemented methanol carbonylation with CO to acetic acid and selective acetic acid hydrogenation to ethylene in tandem, offered ethylene selectivity of 85% at complete methanol conversion at 583 K. The selective removal of Brønsted acid sites in the 12-membered ring channel of H-MOR favors the selectivity of acetic acid in CHOH carbonylation. The high capabilities of ZnO-TiO in the adsorption of acetic acid and the activation of H play key roles in selective hydrogenation of acetic acid to ethylene. Our work provides a promising relay-catalysis strategy for precise C-C coupling of C1 to C2 molecules.
PubMed: 37885567
DOI: 10.1021/jacsau.3c00463 -
Nature Communications Sep 2023Weak hydrogen bonds, such as O-H···π and C-H···O, are thought to direct biochemical assembly, molecular recognition, and chemical selectivity but are seldom...
Weak hydrogen bonds, such as O-H···π and C-H···O, are thought to direct biochemical assembly, molecular recognition, and chemical selectivity but are seldom observed in solution. We have used neutron diffraction combined with H/D isotopic substitution to obtain a detailed spatial and orientational picture of the structure of benzene-methanol mixtures. Our analysis reveals that methanol fully solvates and surrounds each benzene molecule. The expected O-H···π interaction is highly localised and directional, with the methanol hydroxyl bond aligned normal to the aromatic plane and the hydrogen at a distance of 2.30 Å from the ring centroid. Simultaneously, the tendency of methanol to form chain and cyclic motifs in the bulk liquid is manifest in a highly templated solvation structure in the plane of the ring. The methanol molecules surround the benzene so that the O-H bonds are coplanar with the aromatic ring while the oxygens interact with C-H groups through simultaneous bifurcated hydrogen bonds. This demonstrates that weak hydrogen bonding can modulate existing stronger interactions to give rise to highly ordered cooperative structural motifs that persist in the liquid phase.
PubMed: 37736749
DOI: 10.1038/s41467-023-41451-y -
Antioxidants (Basel, Switzerland) Nov 2023Spinach methanolic extract (SME) has a hepatoprotective effect due to its polyphenolic antioxidants; however, its action in parenchymal (PQ) and non-parenchymal (nPQ)...
Spinach methanolic extract (SME) has a hepatoprotective effect due to its polyphenolic antioxidants; however, its action in parenchymal (PQ) and non-parenchymal (nPQ) cells remains unknown. This study investigates the hepatoprotective effect of SME on streptozotocin-induced hyperglycemic rats (STZ), focusing on immunohistochemical analyses. : The extract was prepared, and the total polyphenols and antioxidant activity were quantified. Adult male Wistar rats were divided into four groups ( = 8): normoglycemic rats (NG), STZ-induced hyperglycemic (STZ), STZ treated with 400 mg/kg SME (STZ-SME), and NG treated with SME (SME) for 12 weeks. Serum liver transaminases and lipid peroxidation levels in tissue were determined. The distribution pattern and relative levels of markers related to oxidative stress [reactive oxygen species (ROS), superoxide dismutase-1, catalase, and glutathione peroxidase-1], of cytoprotective molecules [nuclear NRF2 and heme oxygenase-1 (HO-1)], of inflammatory mediators [nuclear NF-κB, TNF-α], proliferation (PCNA), and of fibrogenesis markers [TGF-β, Smad2/3, MMP-9, and TIMP1] were evaluated. : SME had antioxidant capacity, and it lowered serum transaminase levels in STZ-SME compared to STZ. It reduced NOX4 staining, and lipid peroxidation levels were related to low formation of ROS. In STZ-SME, the immunostaining for antioxidant enzymes increased in nPQ cells compared to STZ. However, enzymes were also localized in extra and intracellular vesicles in STZ. Nuclear NRF2 staining and HO-1 expression in PQ and nPQ were higher in STZ-SME than in STZ. Inflammatory factors were decreased in STZ-SME and were related to the percentage decrease in NF-κB nuclear staining in nPQ cells. Similarly, TGF-β (in the sinusoids) and MMP-9 (in nPQ) were increased in the STZ-SME group compared to the other groups; however, staining for CTGF, TIMP1, and Smad2/3 was lower. : SME treatment in hyperglycemic rats induced by STZ may have hepatoprotective properties due to its scavenger capacity and the regulation of differential expression of antioxidant enzymes between the PQ and nPQ cells, reducing inflammatory and fibrogenic biomarkers in liver tissue.
PubMed: 38001866
DOI: 10.3390/antiox12112013 -
STAR Protocols Dec 2023Amid a surge in waste volume, the need to achieve sustainable waste treatment has become increasingly important. Here, we present a protocol for the design and...
Amid a surge in waste volume, the need to achieve sustainable waste treatment has become increasingly important. Here, we present a protocol for the design and accelerated optimization of a waste-to-energy system using artificial intelligence tools. We describe steps for waste treatment process advancement as demonstrated by the medical waste-to-methanol conversion and implementing data-driven process optimization. We then detail procedures for streamlining tasks by establishing connectivity between systems such as Aspen Plus and MATLAB. For complete details on the use and execution of this protocol, please refer to Shi et al. (2022) and Fang et al. (2022)..
Topics: Artificial Intelligence; Methanol
PubMed: 37905497
DOI: 10.1016/j.xpro.2023.102685 -
Frontiers in Oncology 2023Early-life osteosarcoma is associated with severe morbidity and mortality, particularly affecting young children and adults. The present cancer treatment regimen is...
INTRODUCTION
Early-life osteosarcoma is associated with severe morbidity and mortality, particularly affecting young children and adults. The present cancer treatment regimen is exceedingly costly, and medications like ifosfamide, doxorubicin, and cisplatin have unneeded negative effects on the body. With the introduction of hyphenated technology to create medications based on plant molecules, the application of ayurvedic medicine as a new dimension (formulation, active ingredients, and nanoparticles) in the modern period is rapidly growing. The primary source of lead compounds for the development of medications for avariety of ailments is plants and their products. Traditionally, (cumin) has been used as medication to treat a variety of illnesses and conditions.
METHODS
The cumin seed was successfully extracted with solvents Hexane, Chloroform, Methanol, Ethanol and Acetone. Following the solvent extraction, the extract residue was assayed in MG63 cells for their anti-proliferative properties.
RESULTS
First, we used the [3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide] (MTT) assay to test the extracted residue's cytotoxicity. The results show that hexane extract Half-maximal inhibitory concentration (IC50 86 µG/mL) effciently inhibits cells by causing programmed cell death. Furthermore, using the Acridine orange/ethidium bromide (AO/EB) staining method, the lactate dehydrogenase assay, and the reactive oxygen species assay using the Dichloro-dihydro-fluorescein diacetate (DCHFDA) staining method, we have demonstrated that the hexane extract causes apoptosis in MG63 cells. Furthermore, flow cytometry research revealed that the hexane extract stops the cell cycle in the S phase. In addition, the hexane extract limits colony formation and the migration potential as shown by the scratch wound healing assay. Furthermore, the extract from cumin seeds exhibits remarkable bactericidal properties against infections that are resistant to drugs. Gas chromatography analysis was used to quantitatively determine the hexane and methanolic extract based on the experimental data. The primary chemical components of the extract are revealed by the study, and these help the malignant cells heal. The present study finds that there is scientific validity in using cumin seeds as a novel method of anticancer therapy after undergoing both intrinsic and extrinsic research.
PubMed: 38125945
DOI: 10.3389/fonc.2023.1322875