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Journal of Chromatography. A May 2024Birds are excellent bioindicators of environmental pollution, and blood provides information on contaminant exposure, although its analysis is challenging because of the...
Optimization and uncertainty assessment of a gas chromatography coupled to Orbitrap mass spectrometry method to determine organic contaminants in blood: A case study of an endangered seabird.
Birds are excellent bioindicators of environmental pollution, and blood provides information on contaminant exposure, although its analysis is challenging because of the low volumes that can be sampled. The objective of the present study was to optimize and validate a miniaturized and functional extraction and analytical method based on gas chromatography coupled to Orbitrap mass spectrometry (GCOrbitrap-MS) for the trace analysis of contaminants in avian blood. Studied compounds included 25 organochlorine pesticides (OCPs), 6 polychlorinated biphenyls (PCBs), 8 polybrominated diphenyl ethers (PBDEs) and 15 polycyclic aromatic hydrocarbons (PAHs). Four extraction and clean-up conditions were optimized and compared in terms of efficiency, accuracy, and uncertainty assessment. Extraction with hexane:dichloromethane and miniaturized Florisil pipette clean-up was the most adequate considering precision and accuracy, time, and costs, and was thereafter used to analyse 20 blood samples of a pelagic seabird, namely the Bermuda petrel (Pterodroma cahow). This species, endemic to the Northwest Atlantic, is among the most endangered seabirds of the region that in the '60 faced a decrease in the breeding success likely linked to a consistent exposure to dichloro-diphenyl-trichloroethane (DDT). Indeed, p,p'-DDE, the main DDT metabolite, was detected in all samples and ranged bewteen 1.13 and 6.87 ng/g wet weight. Other ubiquitous compounds were PCBs (ranging from 0.13 to 6.76 ng/g ww), hexachlorobenzene, and mirex, while PAHs were sporadically detected at low concentrations, and PBDEs were not present. Overall, the extraction method herein proposed allowed analysing very small blood volumes (∼ 100 µL), thus respecting ethical principles prioritising the application of less-invasive sampling protocols, fundamental when studying threatened avian species.
Topics: Animals; Gas Chromatography-Mass Spectrometry; Pesticides; Birds; Hydrocarbons, Chlorinated; Polychlorinated Biphenyls; Halogenated Diphenyl Ethers; Polycyclic Aromatic Hydrocarbons; Endangered Species; Environmental Monitoring; Environmental Pollutants
PubMed: 38604058
DOI: 10.1016/j.chroma.2024.464870 -
Frontiers in Microbiology 2023Swab pooling may allow for more efficient use of point-of-care assays for SARS-CoV-2 detection in settings where widespread testing is warranted, but the effects of...
INTRODUCTION
Swab pooling may allow for more efficient use of point-of-care assays for SARS-CoV-2 detection in settings where widespread testing is warranted, but the effects of pooling on assay performance are not well described.
METHODS
We tested the Thermo-Fisher Accula rapid point-of-care RT-PCR platform with contrived pooled nasal swab specimens.
RESULTS
We observed a higher limit of detection of 3,750 copies/swab in pooled specimens compared to 2,250 copies/swab in individual specimens. Assay performance appeared worse in a specimen with visible nasal mucous and debris, although performance was improved when using a standard laboratory mechanical pipette compared to the transfer pipette included in the assay kit.
CONCLUSION
Clinicians and public health officials overseeing mass testing efforts must understand limitations and benefits of swab or sample pooling, including reduced assay performance from pooled specimens. We conclude that the Accula RT-PCR platform remains an attractive candidate assay for pooling strategies owing to the superior analytical sensitivity compared to most home use and point-of-care tests despite the inhibitory effects of pooled specimens we characterized.
PubMed: 37608952
DOI: 10.3389/fmicb.2023.1219214 -
Talanta Sep 2023On-site specific capture is a critical step in accurate analysis of trace Pb(II) in environmental waters. In this connection, a new Pb(II)-imprinted polymer-based...
On-site specific capture is a critical step in accurate analysis of trace Pb(II) in environmental waters. In this connection, a new Pb(II)-imprinted polymer-based adsorbent (LIPA) was in-situ prepared in pipette tip and used as the extraction medium of laboratory-made portable three channels in-tip microextraction apparatus (TIMA). Density function theory was employed to verify the selection of functional monomers for the preparation of LIPA. The physical and chemical properties of the prepared LIPA were inspected with various characterization techniques. Under the beneficial preparation parameters, the LIPA presented satisfactory specific recognition performance towards Pb(II). Selectivity coefficients of LIPA towards Pb(II)/Cu(II) and Pb(II)/Cd(II) were 6.82 and 3.27 times higher than that of non-imprinted polymer-based adsorbent, respectively, and the adsorption capacity towards Pb(II) was as high as 36.8 mg/g. Freundlich isotherm model fitted well with the adsorption data, revealing that the adsorption of Pb(II) on LIPA was a multilayer process. After optimizing the extraction conditions, the developed LIPA/TIMA was employed to field selectively separate and enrich trace Pb(II) in various environmental waters followed by quantification with atomic absorption spectrometry. The enhancement factor, linear range, limit of detection and RSDs for precision were 183, 0.50-10000 ng/L, 0.14 ng/L and 3.2-8.4%, respectively. Accuracy of the developed approach was inspected by means of spiked recovery and confirmation experiments. Achieved results reveal that the developed LIPA/TIMA technique is good for field selective separation and preconcentration of Pb(II) and the introduced approach can be used to measure ultra-trace Pb(II) in a variety of waters.
PubMed: 37220687
DOI: 10.1016/j.talanta.2023.124676 -
Analytica Chimica Acta Jul 2023Aflatoxin B (AFB) is a highly toxic and carcinogenic chemical substance that endangers food safety and human health. Magnetic relaxation switching (MRS) immunosensors...
Aflatoxin B (AFB) is a highly toxic and carcinogenic chemical substance that endangers food safety and human health. Magnetic relaxation switching (MRS) immunosensors are utilized in a variety of applications in food analysis due to its resistance to matrix interferences, but they often suffer from magnetic separation-based multi-washing steps and low sensitivity. Herein, we propose novel MRS strategy for the sensitive detection of AFB using "Limited-Magnitude" size particles: a single millimeter sized polystyrene spheres (PS) and 150 nm superparamagnetic nanoparticles (MNP). Only a single PS is used as the microreactor to enhance all of the magnetic signal on its surface in high concentration by an immune competitive response, successfully preventing signal dilution, which can be transferred by pipette, simplifying the process of separation and washing. The established single polystyrene sphere magnetic relaxation switch biosensor (SMRS) was able to quantify AFB from 0.02 to 200 ng/mL with a detection limit of 14.3 pg/mL. SMRS biosensor has been successfully used for the detection of AFB in wheat and maize samples, and the results in agreement with high performance liquid chromatography-tandem mass spectrometry (HPLC-MS). Benefiting from high sensitivity and convenient operation, the simple and enzyme-free method is promising in trace small molecules applications.
Topics: Humans; Immunoassay; Aflatoxin B1; Biosensing Techniques; Polystyrenes; Magnetic Phenomena; Limit of Detection
PubMed: 37244666
DOI: 10.1016/j.aca.2023.341329 -
Biotechnology Journal Feb 2024Three-dimensional (3D) cell cultures have garnered significant attention in biomedical research due to their ability to mimic the in vivo cellular environment more...
Three-dimensional (3D) cell cultures have garnered significant attention in biomedical research due to their ability to mimic the in vivo cellular environment more accurately. The formation of 3D cell spheroids using hanging drops has emerged as a cost-effective and crucial method for generating uniformly-sized spheroids. This study aimed to validate the potential of a tip-refill wafer (TrW), a disposable laboratory item used to hold pipette tips, in facilitating 3D cell culture. The TrW allows for easy generation of hanging drops by pipetting the solution into the holes of the wafer. The mechanical stability of the hanging drops is ensured by the surface wettability and thickness of the TrW. Hanging drops containing 60-µL of solution remained securely attached to the TrW even when subjected to orbital shaking at 210 rpm. The exceptional resistance to mechanical shaking enabled the use of inertial focusing to facilitate spheroid formation. This was demonstrated through live/dead cell staining, quantitative polymerase chain reaction (qPCR) analysis, and cytoskeleton staining, which revealed that horizontal orbiting at 60 rpm for 15 min promoted cell aggregation and ultimately led to the formation of 3D spheroids. The spheroid harvest rate is 96.1% ± 3.5% across three TrWs, each containing 60 hanging drops. In addition to generating mono-culture 3D spheroids, the TrW-based hanging drop platform also enables the formation of multicellular spheroids, and on-demand pairing and fusion of spheroids. The TrW is a disposable item that does not require any fabrication or surface modification procedures, further enhancing its application potential in conventional biological laboratories.
Topics: Spheroids, Cellular; Cell Culture Techniques
PubMed: 38403449
DOI: 10.1002/biot.202300427 -
Plant Disease Apr 2024Anthracnose fruit rot affecting field peppers ( L.) has been reported in Ontario, Canada, leading to significant crop losses of up to 80% over the past three years. Ten...
Anthracnose fruit rot affecting field peppers ( L.) has been reported in Ontario, Canada, leading to significant crop losses of up to 80% over the past three years. Ten symptomatic fruits per field, exhibiting one or more soft, sunken lesions covered with salmon-colored spore masses (Fig. S1), were collected from one and two Banana pepper fields in August 2022 and 2023, respectively, all located in southwestern Ontario. Small sections of diseased tissue (0.5 cm in length) from lesion edges underwent surface sterilization and plated on 2% potato dextrose agar (PDA, Difco) supplemented with kanamycin (50 mg liter), neomycin sulfate (12 mg liter) and streptomycin sulfate (100 mg liter), and incubated at 22°C for 7 days in the dark. Fifteen fungal colonies were isolated and purified using the hyphal tipping method. All fungal isolates showed a pale gray colony morphology with a faint salmon tint on PDA (Fig. S1). Conidia, produced on PDA after incubating the 15 isolates at 22°C for 17 days in the dark, were hyaline, aseptate, smooth-walled, cylindrical with obtuse ends (Fig. S1), and measured 9.4 to 15.0 × 2.7 to 4.8 µm (mean ± standard deviation of 145 conidia = 11.3 ± 1.2 μm × 3.7 ± 0.5 μm), the typical morphology of species (Damm et al. 2012). Internal transcribed spacer (ITS), actin (ACT), chitin synthase 1 (CHS-1), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), glutamine synthetase (GS), histone H3 (HIS3) and beta-tubulin 2 (TUB2) gene regions of all isolates were amplified and sequenced with primers ITS1/ITS4, ACT-512F/ACT-783R, CHS-79F/CHS-345R, GDF1/GDR1, GSF1/GSR1, CYLH3F/CYLH3R and Bt2a/Bt2b and deposited in GenBank (Accession Nos. ITS: PP060584 to PP060596; ACT, CHS-1, GAPDH, GS, HIS3 and TUB2: PP085919 to PP086005), respectively. The sequences were 100% identical to Colletotrichum scovillei strains from different hosts and countries (ITS: PP079643; ACT: MN718468; CHS-1: MN718466, GAPDH: MN718465.1, HIS3: MT592502, TUB2: MK462971). The maximum likelihood-based phylogenetic analysis of ITS, ACT, CHS-1, GAPDH, GS, HIS3, and TUB2 concatenated sequences was conducted using IQ-TREE 2.2.2.7 (Minh et al. 2020). All isolates from this study were grouped with high bootstrap support values with the holotype CBS 126529 (Fig. S2). Living cultures of these isolates were deposited in the Canadian Collection of Fungal Cultures (DAOMC 252833 to 252847). Pathogenicity was tested by inoculating 4 Banana (cv. Jumbo Stuff) and 4 Bell (cv. Archimedes) pepper fruits with 10 μl droplet of a 1 × 10 conidia ml suspension of each isolate onto a wound made with a sterile pipette tip. Eight control fruits were mock-inoculated with sterilized water. Nine days post-inoculation, necrotic lesions measuring 24.7 ± 0.3 mm on Bell and 27.9 ± 0.2 mm on Banana peppers were observed. was re-isolated from all symptomatic fruits, and its species identity was confirmed through morphology, fulfilling Koch's postulates. Control fruits remained symptom-free, and no fungi were isolated from them. This is the first report of in Canada. Previously identified as a pathogen causing anthracnose on peppers in eastern Asia, the United States, Brazil, and Kosovo (Farr and Rossman 2024; Xhemali et al. 2023), its emergence in Ontario raises significant concerns for pepper crops. Additional research is essential to better understand the epidemiology of the disease and develop effective phytosanitary strategies for control.
PubMed: 38679597
DOI: 10.1094/PDIS-02-24-0373-PDN -
Analytical Sciences : the International... Jul 2023Efficient enzymatic digestion methods are critical for the characterization and identification of glycans. Glycan hydrolysis enzymes are widely utilized for the...
Efficient enzymatic digestion methods are critical for the characterization and identification of glycans. Glycan hydrolysis enzymes are widely utilized for the identification of glycoprotein or glycolipid glycans. The commonly utilized in solution glycan hydrolysis methods require several hours of incubation with enzymes for complete removal of their target monosaccharides. To develop an efficient and simple method for the rapid release of monosaccharides from glycoprotein glycans, we fabricated exoglycosidase-impregnated acrylamide gels in an automatic pipette tip. Our automated enzymatic reactors are based on the simple photochemical copolymerization of monomers comprising acrylamide and methylene-bis-acrylamide-containing enzymes with an azobis compound functioning as the photocatalytic initiator. After filling the tip of the automatic pipette with these acrylamide solutions, polymerization of the acrylamide gel solution was performed by irradiation with a LED. The immobilized enzymes maintained their activities in the pipette tips and their action was completed by fully automatic pipetting for 10 to 30 min. We utilized 8-aminopyrene-1, 3, 6-trisulfonic acid (APTS)-labeled glycans as a substrate and measured by capillary electrophoresis (CE) before and after enzymatic digestion. We demonstrated that this method exhibited quantitative enzymatic and specific cleavage of monosaccharides from glycoprotein glycans.
Topics: Glycoside Hydrolases; Polysaccharides; Glycoproteins; Monosaccharides; Digestion
PubMed: 36917436
DOI: 10.1007/s44211-023-00314-8 -
International Journal of Pharmaceutics Jan 2024Cyclosporine (CsA) is a potent immunosuppressant agent that has been used since 1980 for the treatment of various autoimmune diseases and is extensively used to enhance...
Cyclosporine (CsA) is a potent immunosuppressant agent that has been used since 1980 for the treatment of various autoimmune diseases and is extensively used to enhance the survival rate of patients and grafts following organ transplant surgeries. CsA is a poorly soluble drug with a narrow therapeutic window and inter-subject variability, which can lead to graft rejection, nephrotoxicity and other severe adverse effects. This study explores a novel method that combines solubility enhancement of CsA using SNEDDS formulation and personalized dosage delivery using 3D printing technology. The oil phase was chosen as a combination of caproyl 90 and octanoic acid while the Smix phase was chosen as a combination of cremophore El and PEG 400. The optimized liquid SNEDDS was solidified using PEG 6000. An FDM printer was used to print a capsular shell with an oval base that ascends to form a dome with an opening at the top. This opening is used to fill the molten CsA-loaded SNEDDS formulation using a pipette or syringe. The CsA-loaded SNEDDS formulation was characterized by FTIR, DSC and SEM/EDX. The in-vitro release of CsA showed complete release within sixty minutes and followed Korsmeyer-Peppas release kinetics. The drug release was not affected by either the shell opening size or the amount of the loaded formulation. This novel method is simple and straightforward for personalized dosage delivery of drug-loaded SNEDDS formulations.
Topics: Humans; Drug Delivery Systems; Cyclosporine; Emulsions; Drug Liberation; Solubility; Printing, Three-Dimensional; Particle Size; Nanoparticles; Biological Availability
PubMed: 38101759
DOI: 10.1016/j.ijpharm.2023.123707 -
Analytica Chimica Acta Feb 2024Attentions regarding ordered mesoporous silica materials (OMSs), with large specific surface areas and narrow pore size distribution, which are prepared via...
BACKGROUND
Attentions regarding ordered mesoporous silica materials (OMSs), with large specific surface areas and narrow pore size distribution, which are prepared via self-assembly techniques, have been raised in sorption, separation, and sample preparation. However, in order to extend and improve their applications, a functionalization step is required. Organic units can be anchored on the inner or outer surface as well as in the silica wall framework by co-condensation-, grafting-, and periodic mesoporous organosilica (PMO) preparation approaches. Apparently, by synthesizing PMO with extensive and flexible organic bridging groups within the mesoporous wall, an efficient extractive phase can be achieved.
RESULTS
We employed tyrosine amino acid to synthesize a PMO-based extractive phase. The FT-IR, H NMR, HR-ESI-MS, Low angle-XRD, TEM, FESEM, BET, and EDX-MAP analyses confirmed the successful synthesis of PMO within the salt-assisted templating method. A comprehensive study on sorption behavior of PMO was performed and its efficiency was evaluated against the grafting and co-condensation methods. Then, it was implemented to the pipette tip-micro solid phase extraction (PT-μ-SPE) of widely used non-steroidal anti-inflammatory drugs (NSAIDs) in water/wastewaters. Limits of detection and quantification were obtained in the range of 0.1-1.5 and 0.3-5 μg L, respectively. The calibration plots are linear in the 1-1000, 3-1000, 10-750, and 3-750 μg L, respectively. The intra-and inter-day precision at 50 and 200 μg L levels are 2.9-7.1 % and 3.5-8%, while recoveries are between 84 and 111 %.
SIGNIFICANCE
High-capacity tyrosine functionalized PMO with 2D hexagonal symmetry silica mesoporous structures found to be highly efficient extractive media. Despite the bulkiness and flexibility of the bridging group within the mesoporous wall, the synthesis condition was optimized in order to load more organic content in the PMO structure. The PMO performance was superior over organically modified ordered mesoporous silica materials prepared by the grafting and co-condensation methods.
Topics: Tyrosine; Spectroscopy, Fourier Transform Infrared; Amino Acids; Anti-Inflammatory Agents, Non-Steroidal; Silicon Dioxide
PubMed: 38246742
DOI: 10.1016/j.aca.2024.342206 -
Journal of Chromatography. A Jan 2024Organic-silica hybrid monolithic materials have attracted considerable attention as potential stationary phases in separation science. These materials combine the... (Review)
Review
Organic-silica hybrid monolithic materials have attracted considerable attention as potential stationary phases in separation science. These materials combine the advantages of organic polymer and silica-based monoliths, including easy preparation, lower back pressure, high permeability, excellent mechanical strength, thermal stability, and tunable surface chemistry with high surface area and selectivity. The outstanding chromatographic efficiency as stationary phase of hybrid monolithic capillary columns for capillary liquid chromatography and capillary electrochromatography has been reported in many papers. Organic-silica hybrid monolithic materials have also been extensively used in the field of sample preparation. Owing to their surface functionalities, these porous sorbents offer unique selectivity for pre-concentration of different analytes in the most complex matrixes by fast dynamic transport. These sorbents not only improve the analytical method sensitivity, but also introduce novelties in terms of extraction devices and instrument coupling strategies. The current review covers the period spanning from 2017 to 2023 and describes the properties of organic-inorganic hybrid monolithic materials, the present status of this technology and summarizes recent developments in their use as innovative sorbents for microextraction sample preparation techniques (solid phase microextraction with pipette tip, offline in-tube SPME, in-tube SPME online with LC, and in-tube SPME directly coupled with mass spectrometry). Aspects such as the synthesis methods (sol-gel process, one-pot approach, and polyhedral oligomeric silsesquioxanes-based procedure), characterization techniques, and strategies to improve extraction efficiency in various applications in different areas (environmental, food, bioanalysis, and proteomics) are also discussed.
Topics: Silicon Dioxide; Chromatography, Liquid; Capillary Electrochromatography; Solid Phase Microextraction; Polymers
PubMed: 38000199
DOI: 10.1016/j.chroma.2023.464518