Synthesis of δ-MnO2 via ozonation routine for low temperature formaldehyde removal.

Nowadays, it is still a challenge to prepared high efficiency and low cost formaldehyde (HCHO) removal catalysts in order to tackle the long-living indoor air pollution. Herein, δ-MnO2 is successfully synthesized by a facile ozonation strategy, where Mn2+ is oxidized by ozone (O3) bubble in an alkaline solution. It presents one of the best catalytic properties with a low 100% conversion temperature of 85°C for 50 ppm of HCHO under a GHSV of 48,000 mL/(g·hr). As a comparison, more than 6 times far longer oxidation time is needed if O3 is replaced by O2. Characterizations show that ozonation process generates a different intermediate of tetragonal ß-HMnO2, which would favor the quick transformation into the final product δ-MnO2, as compared with the relatively more thermodynamically stable monoclinic γ-HMnO2 in the O2 process. Finally, HCHO is found to be decomposed into CO2 via formate, dioxymethylene and carbonate species as identified by room temperature in-situ diffuse reflectance infrared fourier transform spectroscopy. All these results show great potency of this facile ozonation routine for the highly active δ-MnO2 synthesis in order to remove the HCHO contamination.

Quantitative and qualitative assessment of airborne microorganisms during gross anatomical class and the bacterial and fungal load on formalin-embalmed corpses.

Mold growth on body donations remains an underreported yet serious issue in anatomical teaching. Bacterial and fungal growth pose health risks to lecturers and students, alongside with ethical and aesthetic concerns. However, limited information exists on the presence of bacteria and fungi on body donations and their underlying causes. To investigate the potential impact of airborne germs on body donation contamination, we conducted indoor air measurements before, during, and after our anatomical dissection course, with outdoor measurements serving as a control. Tissue samples from the dissected body donations were collected to assess the germ load, with qualitative and quantitative microbiological analyses. Air samples from the dissection hall contained no fungi, but various fungal species were identified in the adjacent stairways and outdoors which implies that fungal occurrence in the dissection hall air was independent of lecturers' and students' presence. Moreover, our results indicate that adequate ventilation filters can effectively reduce indoor fungal germs during courses, while the bacterial load in room air appears to increase, likely due to the presence of lecturers and students. Additionally, the tissue samples revealed no bacterial or fungal germs which implies that our ethanol-formalin-based embalming solution demonstrates an effective long-term antimicrobial preservation of corpses.

Quantitative Comparison of Deparaffinization, Rehydration, and Extraction Methods for FFPE Tissue Proteomics and Phosphoproteomics.

Formalin-fixed paraffin-embedded (FFPE) tissues are suitable for proteomic and phosphoproteomic biomarker studies by data-independent acquisition mass spectrometry. The choice of the sample preparation method influences the number, intensity, and reproducibility of identifications. By comparing four deparaffinization and rehydration methods, including heptane, histolene, SubX, and xylene, we found that heptane and methanol produced the lowest coefficients of variation (CVs). Using this, five extraction methods from the literature were modified and evaluated for their performance using kidney, leg muscle, lung, and testicular rat organs. All methods performed well, except for SP3 due to insufficient tissue lysis. Heat n' Beat was the fastest and most reproducible method with the highest digestion efficiency and lowest CVs. S-Trap produced the highest peptide yield, while TFE produced the best phosphopeptide enrichment efficiency. The quantitation of FFPE-derived peptides remains an ongoing challenge with bias in UV and fluorescence assays across methods, most notably in SPEED. Functional enrichment analysis demonstrated that each method favored extracting some gene ontology cellular components over others including chromosome, cytoplasmic, cytoskeleton, endoplasmic reticulum, membrane, mitochondrion, and nucleoplasm protein groups. The outcome is a set of recommendations for choosing the most appropriate method for different settings.

Impact of formaldehyde on ozone formation in Central China: Important role of biogenic emission in forest region.

Formaldehyde (HCHO) is an important source for driving tropospheric ozone (O3) formation. This study investigated the combined effects of anthropogenic and biogenic emission on O3 formation in the Guanzhong Basin (GZB), Central China, providing useful information into the mechanisms of O3 formation due to the interaction between anthropogenic and biogenic volatile organic compounds (VOCs). A severe O3 pollution episode in summer of 2017 was simulated using the Weather Research and Forecasting model coupled with Chemistry (WRF-Chem) to examine the impacts of ambient HCHO on ground-level O3. Results showed secondary HCHO dominated ambient levels, peaking in the afternoon (up to 86 %), while primary emissions contributed 14 % on average. This enhanced O3 production by 7.7 % during the morning rush hour and 24.3 % in the afternoon. In addition, HCHO concentration peaked before that of O3, suggesting it plays significant role in O3 formation. Biogenic emission oxidation contributed 3.1 µg m-3 (53.1 %) of HCHO and 5.2 pptv (40.1 %) of hydroperoxyl radicals (HO2) in average urban areas, where the downwind regions of the forests had high nitrogen oxides (NOx) levels and favorable conditions for O3 production (17.3 µg m-3, 20.5 %). In forested regions, sustained isoprene oxidation led to elevated oxidized VOCs including HCHO and acetaldehyde downwind, which practiced further photolysis of O3 formation with anthropogenic NOx in urban areas. Sensitivity experiments recommend controlling industrial and traffic NOx emissions, with regional joint prevention and regulation, which are essential to reduce O3 pollution.

New vaccination approach using formalin-killed Streptococcus pyogenes vaccine on the liver of Oreochromis niloticus fingerlings.

Newly synthesized vaccines prepared from formalin-killed bacteria Streptococcus pyogenes were investigated in the current study to evaluate the effectiveness of the newly synthesized vaccine as well as their safety by injected intraperitoneal. The study involved several steps 1st step is the preparation of the vaccine followed by the 2nd step: Evaluate the effectiveness and vaccine safety against pathogenic S. pyogenes through 4 different groups including control (Group I). Group II (Bacterial, infected group), Group III (Vaccine), and the Last group was the challenged group after the vaccination (Vacc + Bac). Different Immunological and biochemical parameters were measured in addition to hematological and histopathological examinations. For example, oxidative/antioxidants, inflammatory biomarkers, fragmentation and cell damage, and finally the histopathological study. The current study showed an increase in all oxidative, inflammatory, and cell damage (DNA fragmentation assays), additionally markedly elevation in histopathological cell damage in the infected group (Group II) compared with the control group. The vaccine and challenged after vaccination group (vaccine + Bacteria), showed great improvement in oxidative biomarkers (LPO) and an increase in antioxidants biomarkers (GSH, SOD, GST, DPPH, ABTS, GR and GPx), Also the inflammation and histopathological examination. The newly synthesized vaccine improved the resistance of Oreochromis niloticus and can be used as a preventive therapy agent for pathogenic bacteria S. pyogenes.

Proximity ligation-based sequencing for the identification of human papillomavirus genomic integration sites in formalin-fixed paraffin embedded oropharyngeal squamous cell carcinomas.

Human papillomavirus (HPV) infections are an increasing cause of oropharyngeal squamous cell carcinomas (OPSCC). Integration of the viral genome into the host genome is suggested to affect carcinogenesis, however, the correlation with OPSCC patient prognosis is still unclear. Research on HPV integration is hampered by current integration detection technologies and their unsuitability for formalin-fixed paraffin-embedded (FFPE) tissues. This study aims to develop and validate a novel targeted proximity-ligation based sequencing method (targeted locus amplification/capture [TLA/TLC]) for HPV integration detection in cell lines and FFPE OPSCCs. For the identification of HPV integrations, TLA/TLC was applied to 7 cell lines and 27 FFPE OPSCCs. Following preprocessing steps, a polymerase chain reaction (PCR)-based HPV enrichment was performed on the cell lines and a capture-based HPV enrichment was performed on the FFPE tissues before paired-end sequencing. TLA was able to sequence up to hundreds of kb around the target, detecting exact HPV integration loci, structural variants, and chromosomal rearrangements. In all cell lines, one or more integration sites were identified, in accordance with detection of integrated papillomavirus sequences PCR data and the literature. TLC detected integrated HPV in 15/27 FFPE OPSCCs and identified simple and complex integration patterns. In general, TLA/TLC confirmed PCR data and detected additional integration sites. In conclusion TLA/TLC reliably and robustly detects HPV integration in cell lines and FFPE OPSCCs, enabling large, population-based studies on the clinical relevance of HPV integration. Furthermore, this approach might be valuable for clonality assessment of HPV-related tumors in clinical diagnostics.

Orofacial antinociceptive activity of codeine-associated geraniol in mice: a controlled triple-blind study.

This is a nonclinical, controlled, and triple-blind study to investigate the effects of codeine-associated geraniol on the modulation of orofacial nociception and its potential central nervous system depressing effect in an animal model. The orofacial antinociceptive activity of geraniol in combination with codeine was assessed through the following tests: (i) formalin-induced pain, (ii) glutamate-induced pain, and (iii) capsaicin-induced pain. Six animals were equally distributed into six groups and received the following treatments, given intraperitoneally (i.p.) 30 minutes before the experiments: a) geraniol/codeine 50/30 mg/kg; b) geraniol/codeine 50/15 mg/kg; c) geraniol/codeine 50/7.5 mg/kg; d) geraniol 50 mg/kg; e) codeine 30 mg/kg (positive control); or f) 0.9% sodium chloride (negative control). We performed pain behavior analysis after the injection of formalin (20 µL, 20%), glutamate (20 µL, 25 µM), and capsaicin (20 µL, 2.5 µg) into the paranasal region. Rubbing time of the paranasal region by the hind or front paw was used as a parameter. In the neurogenic phase of the formalin test, the geraniol/codeine at 50/7.5 mg/kg was able to promote the maximum antinociceptive effect, reducing nociception by 71.9% (p < 0.0001). In the inflammatory phase of the formalin test, geraniol/codeine at 50/30 mg/kg significantly reduced orofacial nociception (p < 0.005). In the glutamate test, geraniol/codeine at 50/30 mg/kg reduced the rubbing time by 54.2% and reduced nociception in the capsaicin test by 66.7% (p < 0.005). Geraniol alone or in combination does not promote nonspecific depressing effects on the central nervous system. Based on our findings, we suggest the possible synergy between geraniol and codeine in the modulation of orofacial pain.

Methods for Fixing Biofilms of Staphylococcus aureus and Salmonella enterica for Microscopic Examination.

Different methods for fixing biofilms of Staphylococcus aureus and Salmonella enterica for light and electron microscopy were compared. Paraformaldehyde fixation did not preserve biofilm integrity during dehydration; Ito-Karnovsky fixation revealed cell morphology, but did not preserve the matrix. Ruthenium red combined with aldehydes allowed the matrix to be preserved and visualized. An analysis of the ultrastructure of S. aureus and S. enterica cells in biofilms and suspensions at various fixations is presented. The ultrastructure of the biofilm matrix has been described.

Systematic review and feasibility study on pre-analytical factors and genomic analyses on archival formalin-fixed paraffin-embedded breast cancer tissue.

Formalin-fixed paraffin-embedded (FFPE) tissue represents a valuable source for translational cancer research. However, the widespread application of various downstream methods remains challenging. Here, we aimed to assess the feasibility of a genomic and gene expression analysis workflow using FFPE breast cancer (BC) tissue. We conducted a systematic literature review for the assessment of concordance between FFPE and fresh-frozen matched tissue samples derived from patients with BC for DNA and RNA downstream applications. The analytical performance of three different nucleic acid extraction kits on FFPE BC clinical samples was compared. We also applied a newly developed targeted DNA Next-Generation Sequencing (NGS) 370-gene panel and the nCounter BC360® platform on simultaneously extracted DNA and RNA, respectively, using FFPE tissue from a phase II clinical trial. Of the 3701 initial search results, 40 articles were included in the systematic review. High degree of concordance was observed in various downstream application platforms. Moreover, the performance of simultaneous DNA/RNA extraction kit was demonstrated with targeted DNA NGS and gene expression profiling. Exclusion of variants below 5% variant allele frequency was essential to overcome FFPE-induced artefacts. Targeted genomic analyses were feasible in simultaneously extracted DNA/RNA from FFPE material, providing insights for their implementation in clinical trials/cohorts.

Targeted proteomic approach for quantification of collagen type I and type III in formalin-fixed paraffin-embedded tissue.

Collagen is the most abundant protein in mammals and a major structural component of the extracellular matrix (ECM). Changes to ECM composition occur as a result of numerous physiological and pathophysiological causes, and a common means to evaluate these changes is the collagen 3 (Col3) to collagen 1 (Col1) ratio. Current methods to measure the Col3/1 ratio suffer from a lack of specificity and often under- or over-estimate collagen composition and quantity. This manuscript presents a targeted liquid chromatography tandem mass spectrometry (LC-MS/MS) method for quantification of Col3 and Col1 in FFPE tissues. Using surrogate peptides to generate calibration curves, Col3 and Col1 are readily quantified in FFPE tissue sections with high accuracy and precision. The method is applied to several tissue types from both human and reindeer sources, demonstrating its generalizability. In addition, the targeted LC-MS/MS method permits quantitation of the hydroxyprolinated form of Col3, which has significant implications for understanding not only the quantity of Col3 in tissue, but also understanding of the pathophysiology underlying many causes of ECM changes. This manuscript presents a straightforward, accurate, precise, and generalizable method for quantifying the Col3/1 ratio in a variety of tissue types and organisms.

A Fast-Tracking Sample Preparation Protocol for Proteomics of Formalin-Fixed Paraffin-Embedded Tumor Tissues.

Archived tumor specimens are routinely preserved by formalin fixation and paraffin embedding. Despite the conventional wisdom that proteomics might be ineffective due to the cross-linking and pre-analytical variables, these samples have utility for both discovery and targeted proteomics. Building on this capability, proteomics approaches can be used to maximize our understanding of cancer biology and clinical relevance by studying preserved tumor tissues annotated with the patients' medical histories. Proteomics of formalin-fixed paraffin-embedded (FFPE) tissues also integrates with histological evaluation and molecular pathology strategies, so that additional collection of research biopsies or resected tumor aliquots is not needed. The acquisition of data from the same tumor sample also overcomes concerns about biological variation between samples due to intratumoral heterogeneity. However, the protein extraction and proteomics sample preparation from FFPE samples can be onerous, particularly for small (i.e., limited or precious) samples. Therefore, we provide a protocol for a recently introduced kit-based EasyPep method with benchmarking against a modified version of the well-established filter-aided sample preparation strategy using laser-capture microdissected lung adenocarcinoma tissues from a genetically engineered mouse model. This model system allows control over the tumor preparation and pre-analytical variables while also supporting the development of methods for spatial proteomics to examine intratumoral heterogeneity. Data are posted in ProteomeXchange (PXD045879).

Robust Detection of Gene Amplification in Formalin-Fixed Paraffin-Embedded Samples by Fluorescence in situ Hybridization.

Focal gene amplification, such as extrachromosomal DNA (ecDNA), plays an important role in cancer development and therapy resistance. While sequencing-based methodologies enable an unbiased identification of ecDNA, cytogenetic-based techniques, such as fluorescence in situ hybridization (FISH), remain time and cost-effective for identifying ecDNA in clinical specimens. The application of FISH in formalin-fixed paraffin-embedded (FFPE) tissue samples offers a unique avenue for detecting amplified genes, particularly when viable specimens are not available for karyotype examination. However, there is a lack of consensus procedures for this technique. This protocol provides comprehensive, fully optimized, step-by-step instructions for conducting FISH to detect gene amplification, including ecDNA, in FFPE tissue samples which present unique challenges that this protocol aims to overcome and standardize. By following this protocol, researchers can reproducibly acquire high-quality imaging data to assess gene amplification.

Nanoscale intracellular ultrastructures affected by osmotic pressure using small-angle X-ray scattering.

Although intracellular ultrastructures have typically been studied using microscopic techniques, it is difficult to observe ultrastructures at the submicron scale of living cells due to spatial resolution (fluorescence microscopy) or high vacuum environment (electron microscopy). We investigate the nanometer scale intracellular ultrastructures of living CHO cells in various osmolality using small-angle X-ray scattering (SAXS), and especially the structures of ribosomes, DNA double helix, and plasma membranes in-cell environment are observed. Ribosomes expand and contract in response to osmotic pressure, and the inter-ribosomal correlation occurs under isotonic and hyperosmolality. The DNA double helix is not dependent on the osmotic pressure. Under high osmotic pressure, the plasma membrane folds into form a multilamellar structure with a periodic length of about 6 nm. We also study the ultrastructural changes caused by formaldehyde fixation, freezing and heating.

DropBlot: single-cell western blotting of chemically fixed cancer cells.

Archived patient-derived tissue specimens play a central role in understanding disease and developing therapies. To address specificity and sensitivity shortcomings of existing single-cell resolution proteoform analysis tools, we introduce a hybrid microfluidic platform (DropBlot) designed for proteoform analyses in chemically fixed single cells. DropBlot serially integrates droplet-based encapsulation and lysis of single fixed cells, with on-chip microwell-based antigen retrieval, with single-cell western blotting of target antigens. A water-in-oil droplet formulation withstands the harsh chemical (SDS, 6 M urea) and thermal conditions (98 °C, 1-2 hr) required for effective antigen retrieval, and supports analysis of retrieved protein targets by single-cell electrophoresis. We demonstrate protein-target retrieval from unfixed, paraformaldehyde-fixed (PFA), and methanol-fixed cells. Key protein targets (HER2, GAPDH, EpCAM, Vimentin) retrieved from PFA-fixed cells were resolved and immunoreactive. Relevant to biorepositories, DropBlot profiled targets retrieved from human-derived breast tumor specimens archived for six years, offering a workflow for single-cell protein-biomarker analysis of sparing biospecimens.

Design, construction and optimization of formaldehyde growth biosensors with broad application in biotechnology.

Formaldehyde is a key metabolite in natural and synthetic one-carbon metabolism. To facilitate the engineering of formaldehyde-producing enzymes, the development of sensitive, user-friendly, and cost-effective detection methods is required. In this study, we engineered Escherichia coli to serve as a cellular biosensor capable of detecting a broad range of formaldehyde concentrations. Using both natural and promiscuous formaldehyde assimilation enzymes, we designed three distinct E. coli growth biosensor strains that depend on formaldehyde for cell growth. These strains were engineered to be auxotrophic for one or several essential metabolites that could be produced through formaldehyde assimilation. The respective assimilating enzyme was expressed from the genome to compensate the auxotrophy in the presence of formaldehyde. We first predicted the formaldehyde dependency of the biosensors by flux balance analysis and then analysed it experimentally. Subsequent to strain engineering, we enhanced the formaldehyde sensitivity of two biosensors either through adaptive laboratory evolution or modifications at metabolic branch points. The final set of biosensors demonstrated the ability to detect formaldehyde concentrations ranging approximately from 30 µM to 13 mM. We demonstrated the application of the biosensors by assaying the in vivo activity of different methanol dehydrogenases in the most sensitive strain. The fully genomic nature of the biosensors allows them to be deployed as "plug-and-play" devices for high-throughput screenings of extensive enzyme libraries. The formaldehyde growth biosensors developed in this study hold significant promise for advancing the field of enzyme engineering, thereby supporting the establishment of a sustainable one-carbon bioeconomy.

ASCARIS SUUM EGG RECOVERY FROM SLUDGE SAMPLES AFTER PHASE EXTRACTION.

Some helminth test methods for sanitation samples include a phase extraction step to reduce lipid content and final pellet size before microscopy. Hydrophilic and lipophilic solutions are used to create 2 phases, with a plug of organic material or debris in between, whilst eggs are supposedly compacted at the bottom of the test tube. We tested 10% formalin, acetoacetic buffer, and acid alcohol as the hydrophilic solutions, and ethyl acetate and diethyl ether as the lipophilic solvents for egg recoverability from water, primary sludge, and fatty sludge. Normally, the supernatant and debris plug are discarded and the sedimented pellet of eggs is microscopically examined. We, however, also collected the entire supernatant plus debris plug to determine where eggs were possibly lost. We found that eggs were lost when samples were extracted with 10% formalin + ethyl acetate, 10% formalin + diethyl ether, acetoacetic buffer + ethyl acetate, and acetoacetic buffer + diethyl ether combinations (<50% egg recovery). Acid alcohol + ethyl acetate resulted in 93.2, 89.8, and 57.3% egg recovery in the pellet of water, primary sludge, and fatty sludge, respectively; however, the size of the final pellet was not reduced, defeating the purpose of the extraction step. We thus recommend that this step be excluded.

Enhancing Bioactivity through the Transfer of the 2-(Hydroxymethoxy)Vinyl Moiety: Application in the Modification of Tyrosol and Hinokitiol.

Utilizing Density Functional Theory (DFT) calculations at the B3LYP/QZVP level and incorporating the Conductor-like Polarizable Continuum Model (C-PCM) for solvation, the thermodynamic and chemical activity properties of 21-(hydroxymethoxy)henicosadecaenal, identified in cultured freshwater pearls from the mollusk Hyriopsis cumingii, have been elucidated. The study demonstrates that this compound releases formaldehyde, a potent antimicrobial agent, through dehydrogenation and deprotonation processes in both hydrophilic and lipophilic environments. Moreover, this polyenal exhibits strong anti-reductant properties, effectively scavenging free radicals. These critical properties classify the pearl-derived ingredient as a natural multi-functional compound, serving as a coloring, antiradical, and antimicrobial agent. The 2-(hydroxymethoxy)vinyl (HMV) moiety responsible for the formaldehyde release can be transferred to other compounds, thereby enhancing their biological activity. For instance, tyrosol (4-(2-hydroxyethyl)phenol) can be modified by substituting the less active 2-hydroxyethyl group with the active HMV one, and hinokitiol (4-isopropylotropolone) can be functionalized by attaching this moiety to the tropolone ring. A new type of meso-carrier, structurally modeled on pearls, with active substances loaded both in the layers and the mineral part, has been proposed.

Retrospective Single Nucleotide Polymorphism Analysis of Host Resistance and Susceptibility to Ovine Johne's Disease Using Restored FFPE DNA.

Johne's disease (JD), also known as paratuberculosis, is a chronic, untreatable gastroenteritis of ruminants caused by Mycobacterium avium subsp. paratuberculosis (MAP) infection. Evidence for host genetic resistance to disease progression exists, although it is limited due to the extended incubation period (years) and diagnostic challenges. To overcome this, previously restored formalin-fixed paraffin embedded tissue (FFPE) DNA from archived FFPE tissue cassettes was utilized for a novel retrospective case-control genome-wide association study (GWAS) on ovine JD. Samples from known MAP-infected flocks with ante- and postmortem diagnostic data were used. Cases (N = 9) had evidence of tissue infection, compared to controls (N = 25) without evidence of tissue infection despite positive antemortem diagnostics. A genome-wide efficient mixed model analysis (GEMMA) to conduct a GWAS using restored FFPE DNA SNP results from the Illumina Ovine SNP50 Bead Chip, identified 10 SNPs reaching genome-wide significance of p < 1 × 10-6 on chromosomes 1, 3, 4, 24, and 26. Pathway analysis using PANTHER and the Kyoto Encyclopedia of Genes and Genomes (KEGG) was completed on 45 genes found within 1 Mb of significant SNPs. Our work provides a framework for the novel use of archived FFPE tissues for animal genetic studies in complex diseases and further evidence for a genetic association in JD.

Metabolic engineering of Komagataella phaffii for the efficient utilization of methanol.

BACKGROUND: Komagataella phaffii, a type of methanotrophic yeast, can use methanol, a favorable non-sugar substrate in eco-friendly bio-manufacturing. The dissimilation pathway in K. phaffii leads to the loss of carbon atoms in the form of CO2. However, the ΔFLD strain, engineered to lack formaldehyde dehydrogenase-an essential enzyme in the dissimilation pathway-displayed growth defects when exposed to a methanol-containing medium. RESULTS: Inhibiting the dissimilation pathway triggers an excessive accumulation of formaldehyde and a decline in the intracellular NAD+/NADH ratio. Here, we designed dual-enzyme complex with the alcohol oxidase1/dihydroxyacetone synthase1 (Aox1/Das1), enhancing the regeneration of the formaldehyde receptor xylulose-5-phosphate (Xu5P). This strategy mitigated the harmful effects of formaldehyde accumulation and associated toxicity to cells. Concurrently, we elevated the NAD+/NADH ratio by overexpressing isocitrate dehydrogenase in the TCA cycle, promoting intracellular redox homeostasis. The OD600 of the optimized combination of the above strategies, strain DF02-1, was 4.28 times higher than that of the control strain DF00 (ΔFLD, HIS4+) under 1% methanol. Subsequently, the heterologous expression of methanol oxidase Mox from Hansenula polymorpha in strain DF02-1 resulted in the recombinant strain DF02-4, which displayed a growth at an OD600 4.08 times higher than that the control strain DF00 in medium containing 3% methanol. CONCLUSIONS: The reduction of formaldehyde accumulation, the increase of NAD+/NADH ratio, and the enhancement of methanol oxidation effectively improved the efficient utilization of a high methanol concentration by strain ΔFLD strain lacking formaldehyde dehydrogenase. The modification strategies implemented in this study collectively serve as a foundational framework for advancing the efficient utilization of methanol in K. phaffii.

Isotopologues of potassium 2,2,2-trifluoroethoxide for applications in positron emission tomography and beyond.

The 2,2,2-trifluoroethoxy group increasingly features in drugs and potential tracers for biomedical imaging with positron emission tomography (PET). Herein, we describe a rapid and transition metal-free conversion of fluoroform with paraformaldehyde into highly reactive potassium 2,2,2-trifluoroethoxide (CF3CH2OK) and demonstrate robust applications of this synthon in one-pot, two-stage 2,2,2-trifluoroethoxylations of both aromatic and aliphatic precursors. Moreover, we show that these transformations translate easily to fluoroform that has been labeled with either carbon-11 (t1/2 = 20.4 min) or fluorine-18 (t1/2 = 109.8 min), so allowing the appendage of complex molecules with a no-carrier-added 11C- or 18F- 2,2,2-trifluoroethoxy group. This provides scope to create candidate PET tracers with radioactive and metabolically stable 2,2,2-trifluoroethoxy moieties. We also exemplify syntheses of isotopologues of potassium 2,2,2-trifluoroethoxide and show their utility for stable isotopic labeling which can be of further benefit for drug discovery and development.