Enzyme-linked immunoassay (ELISA) is highly specific and selective towards target molecules and is convenient for on-site detection. However, in many cases, lack of high sensitivity makes it hard to ...reveal a significant colorimetric signal for detecting a trace amount of target molecules. Thus, analytical instruments are required for detection, which limits the application of ELISA for on-site detection. In the present study, a highly sensitive and naked-eyed detectable colorimetric biosensor for chloramphenicol (CAP) was prepared by incorporating ELISA onto surfaces of microporous and nanofibrous membranes. The high specific surface areas of the nanofibers significantly increased the number of antibodies covalently linked onto the fiber surfaces and binding capacity of the sensor with antigens present in a sample. With such an integration, the sensitivity of the ELISA sensor was dramatically increased, and a trace number of targets could reveal a naked-eye detectable color. The immunoassay sensor exhibited a significant naked-eye distinguishable color to chloramphenicol (CAP) at 0.3 ng/mL. The successful design and fabrication of the nanofibrous membrane immunoassay sensor provide new paths towards the development of on-site inspection sensors without the assistance from any instrument.
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•Naked eye detectable colorimetric sensor.•ELISA on nanofibrous membranes.•Rapid detection of antibiotics without instrument.
•Antibiotic recovery and matrix effects were tested in pigmented and non-pigmented salmon.•Extraction recoveries with the QuEChERS method was low with pigmented salmon.•Matrix effects were more ...evident in pigmented compared to non-pigmented salmon.•Accuracy was minimally impacted.•Salmon pigmentation causes matrix effects and reduced recoveries without impacting accuracy.
Carotenoid pigmentation in salmon may interfere with the accuracy of antibiotic analysis with ultra-high pressure liquid chromatography coupled to tandem spectrometry (UPLC-MS/MS) by causing matrix effects or affecting the recovery of compounds during extraction. In the present study, we used both pigmented and non-pigmented salmon to understand the role pigments play on antibiotic analysis, and tested whether clean-up of the extract with dispersive solid phase extraction (dSPE) or hydrophilic-lipophilic balance (HLB) SPE clean-up reduces matrix effects. Thirty antibiotics and their respective class-specific surrogate standards were measured in Sockeye (pigmented), King (pigmented) and Ivory King (non-pigmented) salmon extracted using the QUEChERS method, or a modified QUEChERS method involving dSPE or HLB SPE clean-up (for Sockeye salmon only). Significant matrix effects and lower percent recoveries of spiked antibiotics were observed in pigmented versus non-pigmented salmon extracted with the QUEChERS method. Dispersive SPE clean-up did not improve extraction recoveries or matrix effects. However, SPE clean-up with HLB columns improved matrix effects for several antibiotics but reduced the percent recovery to < 30%. Across all types of salmon analyzed, the accuracy of quantitation was minimally impacted, likely due to similar behavior of the surrogate standards tagged to each antibiotic class during extraction. Our results demonstrate that carotenoids in salmon are associated with significant matrix effects and low extraction recoveries, but do not impact accuracy.
Increased intake of omega-6 rich plant oils such as soybean and corn oil over the past few decades has inadvertently tripled the amount of n-6 linoleic acid (LA, 18:2n-6) in the diet. Although LA is ...nutritionally "essential", very little is known about how it affects the brain when present in excess. This review provides an overview on the metabolism of LA by the brain and the effects of excess dietary LA intake on brain function. Pre-clinical evidence suggests that excess dietary LA increases the brain's vulnerability to inflammation and likely acts via its oxidized metabolites. In humans, excess maternal LA intake has been linked to a typical neurodevelopment, but underlying mechanisms are unknown. It is concluded that excess dietary LA may adversely affect the brain. The potential neuroprotective role of reducing dietary LA merits clinical evaluation in future studies.
Polyunsaturated fatty acids (PUFAs) are structural components of membrane phospholipids, and influence cellular function via effects on membrane properties, and also by acting as a precursor pool for ...lipid mediators. These lipid mediators are formed via activation of pathways involving at least one step of dioxygen-dependent oxidation, and are consequently called oxylipins. Their biosynthesis can be either enzymatically-dependent, utilising the promiscuous cyclooxygenase, lipoxygenase, or cytochrome P450 mixed function oxidase pathways, or nonenzymatic via free radical-catalyzed pathways. The oxylipins include the classical eicosanoids, comprising prostaglandins, thromboxanes, and leukotrienes, and also more recently identified lipid mediators. With the advent of new technologies there is growing interest in identifying these different lipid mediators and characterising their roles in health and disease. This review brings together contributions from some of those at the forefront of research into lipid mediators, who provide brief introductions and summaries of current understanding of the structure and functions of the main classes of nonclassical oxylipins. The topics covered include omega-3 and omega-6 PUFA biosynthesis pathways, focusing on the roles of the different fatty acid desaturase enzymes, oxidized linoleic acid metabolites, omega-3 PUFA-derived specialized pro-resolving mediators, elovanoids, nonenzymatically oxidized PUFAs, and fatty acid esters of hydroxy fatty acids.
Fish and algae oil supplements are enriched with eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), which are precursors to oxidized fatty acids, known as oxylipins. Here, we optimized a ...base hydrolysis method for measuring oxylipins in oil with ultrahigh-performance liquid chromatography coupled to tandem mass-spectrometry (UPLC-MS/MS) and quantified them in fish and algae oil supplements. Hydrolysis of 2 μL of oil with sodium carbonate resulted in greater oxylipin concentrations and minimal matrix effects, compared to higher oil volumes (10, 20, and 30 μL). Oxylipin yield was higher when oil was hydrolyzed in methanol containing 0.1% acetic acid and 0.1% butylated hydroxytoluene, compared to no methanol, and using sodium hydroxide versus sodium carbonate. Oxylipins extracted from 2 μL of oil using sodium hydroxide in solvent showed that EPA-derived oxylipins were most abundant in fish oil (84–87%), whereas DHA-oxylipins were abundant in algae oil (83%). This study shows that fish and algae oils are direct sources of EPA- and DHA-derived oxylipins.
Infant milk formula was used as a model food to compare the sensitivity of thiobarbituric acid reactive substances (TBARS) and hydroperoxide methods to UPLC‐MS/MS oxylipin analysis for detecting ...early lipid oxidation. Two different infant milk formulas were tested during 21 days of storage at 4°C. Formulas 1 and 2 contained canola oil and canola oil + 1% docosahexaenoic acid ethyl ester, respectively. Formulas were sampled up to 21 days of storage. Formula 2 had higher peroxide values than Formula 1 across all time points. However, no significant differences over time in TBARS and peroxide values in either formula were observed. Several oxylipins increased in both formulas starting on day 7 (linoleic acid and alpha‐linolenic acid‐derived oxylipins in Formula 1 and DHA‐derived oxylipins in Formula 2). These results indicate that free oxylipins are effective in detecting early lipid oxidation and distinguishing between formulations containing different fatty acids.
Practical Application
We have recently shown that primary oxidation products known as oxylipins can be measured in their free form by ultra‐high pressure liquid chromatography‐tandem mass spectrometry (UPLC‐MS/MS) to detect early lipid oxidation. However, a head‐to‐head comparison of the sensitivity of this approach to conventional spectrophotometric methods has not been evaluated. Our results indicate that free oxylipin measurements are better than conventional methods in detecting early lipid oxidation in milk infant formula distinguishing between different formulations.
Circulating oxidized linoleic acid (LA) metabolites (OXLAMs) are increased in patients with nonalcoholic steatohepatitis (NASH) and their levels correlate with disease severity. However, the ...mechanisms by which OXLAMs contribute to NASH development are incompletely understood. We tested the hypothesis that LA or OXLAMs provided directly through the diet are involved in the development of hepatic injury. C57BL/6 mice were fed an isocaloric high-fat diet containing low LA, high LA, or OXLAMs for 8 weeks. The livers of OXLAM-fed mice showed lower triglyceride concentrations, but higher FA oxidation and lipid peroxidation in association with increased oxidative stress. OXLAM-induced mitochondrial dysfunction was associated with reduced Complex I protein and hepatic ATP levels, as well as increased mitochondrial biogenesis and cytoplasmic mitochondrial DNA. Oxidative stress increased thioredoxin-interacting protein (TXNIP) in the liver and stimulated the activation of mitochondrial apoptosis signal-regulating kinase 1 (ASK1) leading to apoptosis. We also found increased levels of NOD-like receptor protein 3 (NLRP3) inflammasome components and Caspase-1 activation in the livers of OXLAM-fed mice. In vitro, OXLAMs induced hepatocyte cell death, which was partly dependent on Caspase-1 activation. This study identified key mechanisms by which dietary OXLAMs contribute to NASH development, including mitochondrial dysfunction, hepatocyte cell death, and NLRP3 inflammasome activation.
Enzyme-linked immunosorbent assays (ELISA), as one of the most used immunoassays, have been conducted ubiquitously in hospitals, research laboratories, etc. However, the conventional ELISA procedure ...is usually laborious, occupies bulky instruments, consumes lengthy operation time, and relies considerably on the skills of technicians, and such limitations call for innovations to develop a fully automated ELISA platform. In this paper, we have presented a system incorporating a robotic-microfluidic interface (RoMI) and a modular hybrid microfluidic chip that embeds a highly sensitive nanofibrous membrane, referred to as the Robotic ELISA, to achieve human-free sample-to-answer ELISA tests in a fully programmable and automated manner. It carries out multiple bioanalytical procedures to replace the manual steps involved in classic ELISA operations, including the pneumatically driven high-precision pipetting, efficient mixing and enrichment enabled by back-and-forth flows, washing, and integrated machine vision for colorimetric readout. The Robotic ELISA platform has achieved a low limit of detection of 0.1 ng/mL in the detection of a low sample volume (15 μL) of chloramphenicol within 20 min without human intervention, which is significantly faster than that of the conventional ELISA procedure. Benefiting from its modular design and automated operations, the Robotic ELISA platform has great potential to be deployed for a broad range of detections in various resource-limited settings or high-risk environments, where human involvement needs to be minimized while the testing timeliness, consistency, and sensitivity are all desired.
Abstract Background Disturbances in prefrontal cortex phospholipid and fatty acid composition have been reported in patients with schizophrenia (SCZ), often as an incomplete lipid profile or a ...percent of total lipid concentration. In this study, we quantified absolute concentrations (nmol/g wet weight) and fractional concentrations (i.e. percent of total fatty acids) of several lipid classes and their constituent fatty acids in postmortem prefrontal cortex of SCZ patients ( n = 10) and age-matched controls ( n = 10). Methods Lipids were extracted, fractionated with thin layer chromatography and assayed. Results Mean total lipid, phospholipid, individual phospholipids, plasmalogen, triglyceride and cholesteryl ester concentrations did not differ significantly between the groups. Compared to controls, SCZ brains showed significant increases in several monounsaturated and polyunsaturated fatty acid absolute concentrations in cholesteryl ester. Significant increases or decreases occurred in palmitoleic, linoleic, γ-linolenic and n-3 docosapentaenoic acid absolute concentrations in total lipids, triglycerides or phospholipids. Changes in fractional concentrations did not consistently reflect absolute concentration changes. Conclusion These findings suggest disturbed prefrontal cortex fatty acid absolute concentrations, particularly within cholesteryl esters, as a pathological aspect of schizophrenia.
Previous studies have shown that the size of almond particles influences lipid bioaccessibility during digestion. However, the extent of structural breakdown of almond particles during gastric ...digestion and its impact on lipid bioaccessibility is unclear. In this study, in vitro digestion of almond particles was conducted using a dynamic model (Human Gastric Simulator) and a static model (shaking water bath). Structural breakdown of particles during the gastric phase occurred only in the Human Gastric Simulator, as evidenced by a reduction in particle size (15.89 ± 0.68 mm
to 12.19 ± 1.29 mm
, p < 0.05). Fatty acid bioaccessibility at the end of the gastric phase was greater in the Human Gastric Simulator than in the shaking water bath (6.55 ± 0.85% vs. 4.54 ± 0.36%, p < 0.01). Results showed that the in vitro model of digestion which included peristaltic contractions (Human Gastric Simulator) led to breakdown of almond particles during gastric digestion which increased fatty acid bioaccessibility.
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