Assessing spatial impacts of historical pulp mill effluent on trophic dynamics in a coastal marine ecosystem using stable isotope (δ13C and δ15N) analysis.

Boat Harbour, Nova Scotia was a tidal estuary that was converted into a wastewater treatment facility for pulp mill effluent in 1967. Treated effluent from Boat Harbour was discharged into the coastal Northumberland Strait, contributing significant nutrient and freshwater inputs into the coastal environment, potentially impacting local biogeochemistry and ecosystem structure. This study used stable isotope analysis of carbon (δ13C) and nitrogen (δ15N) of representative taxa to assess spatial variability in nutrient sources and trophic dynamics. Results identified stable isotope variation with depleted δ13C and δ15N values in taxa near Boat Harbour. Blue mussel (Mytilus edulis) and mummichog (Fundulus heteroclitus) were the most suitable bioindicators for identifying variation in nutrient sources. Stable isotope signatures in this study may be reflective of residual pulp mill effluent-derived nutrients, differences in marine versus terrestrial nutrient sources, and a pronounced coastal salinity gradient. The present study defined the baseline nutrient conditions of the Northumberland Strait and will be useful in assessing the effectiveness of remediation activities.

Mechanistic Insights into Myofibrillar Protein Oxidation by Fenton Chemistry Regulated by Gallic Acid.

Gallic acid (GA, 3,4,5-trihydroxybenzoic acid) is a widely used natural food additive of interest to food chemistry researchers, especially regarding its effects on myofibrillar protein (MP) oxidation. However, existing studies regarding MP oxidation by GA-combined with Fenton reagents are inconsistent, and the detailed mechanisms have not been fully elucidated. This work validated hydroxyl radical (HO·) as the primary oxidant for MP carbonylation; in addition, it revealed three functions of GA in the Fenton oxidation of MP. By coordination with Fe(III), GA reduces Fe(III) to generate Fe(II), which is the critical reagent for HO· generation; meanwhile, the coordination improves the availability and reactivity of Fe(III) under weakly acidic and near-neutral pH, i.e., pH 4-6. Second, the intermediates formed during GA oxidation, including semiquinone and quinone, promoted Fenton reactivity by accelerating Fe catalytic cycling. Finally, GA can scavenge HO· radicals, thus exhibiting a certain degree of antioxidant property. All three functions contribute to MP oxidation as observed in GA-containing meat.

Baseline monitoring of contaminant concentrations in American lobster (Homarus americanus) tissues from coastal Northumberland Strait, Nova Scotia, Canada.

A baseline survey was conducted in 2018 to characterize contaminants in American lobsters, Homarus americanus in the Northumberland Strait, Canada. Sampling included three age classes of lobsters at sites 4, 20, and 70 km from the Boat Harbour estuary, a historically contaminated site set to undergo remediation. Lobster tissues were measured for metal(loids), methylmercury, polycyclic aromatic hydrocarbons, and polychlorinated dibenzo-p-dioxins and polychlorinated dibenzo-p-furans. Contaminant concentrations were generally below the guidelines set by the Canadian Council of Ministers of the Environment and the Canadian Food Inspection Agency, except for arsenic which was elevated in all age classes from all sites (4.8-12.68 mg kg-1). Mercury and methylmercury (both ~0.04 mg kg-1) minimally exceeded one guideline in some age-classes and sites. There was also no consistent pattern of contaminant accumulation across either age classes or at particular sites. This study serves as a baseline for future monitoring following remediation of Boat Harbour.

A universal monoclonal antibody-aptamer conjugation strategy for selective non-invasive bioparticle isolation from blood on a regenerative microfluidic platform.

Simultaneous isolation of various circulating tumor cell (CTC) subtypes from whole blood is useful in cancer diagnosis and prognosis. Microfluidic affinity separation devices are promising for CTC separation because of their high throughput capacity and automatability. However, current affinity agents, such as antibodies (mAbs) and aptamers (Apts) alone, are still suboptimal for efficient, consistent, and versatile cell analysis. By introducing a hybrid affinity agent, i.e., an aptamer-antibody (Apt-mAb) conjugate, we developed a universal and regenerative microchip with high efficiency and non-invasiveness in the separation and profiling of various CTCs from blood. The Apt-mAb conjugate consists of a monoclonal antibody that specifically binds the target cell receptor and a surface-bound aptamer that recognizes the conserved Fc region of the mAb. The aptamer then indirectly links the surface functionalization of the microfluidic channels to the mAbs. This hybrid affinity agent and the microchip platform may be widely useful for various bio-particle separations in different biological matrices. Further, the regeneration capability of the microchip improves data consistency between multiple uses and minimizes plastic waste while promoting environmental sustainability. STATEMENT OF SIGNIFICANCE: A hybrid affinity agent, Apt-mAb, consisting of a universal aptamer (Apt) that binds the conserved Fc region of monoclonal antibodies (mAbs) was developed. The invented nano-biomaterial combines the strengths and overcomes the weakness of both Apts and mAbs, thus changing the paradigm of affinity separation of cell subtypes. When Apt-mAb was used to fabricate microfluidic chips using a "universal screwdriver" approach, the microchip could be easily tuned to bind any cell type, exhibiting great universality. Besides high sensitivity and selectivity, the superior regenerative capacity of the microchips makes them reusable, which provides improved consistency and repeatability in cell profiling and opens a new approach towards in vitro diagnostic point-of-care testing devices with environmental sustainability and cost-effectiveness.

Exacerbated Protein Oxidation and Tyrosine Nitration through Nitrite-Enhanced Fenton Chemistry.

Nitrite is a common additive used during meat curing to prevent microbial contamination and retain an attractive red color in the product. However, the effects of nitrite on Fenton reactions catalyzed by free iron in meat products are not well understood, although such processes can induce protein oxidation and nitration, affecting the nutritional and aesthetic quality of meat products. This contribution reveals the mechanism through which nitrite affects Fenton reactions that generate reactive nitrogen and oxygen species by increasing the availability of Fe3+, facilitating its reduction and stabilizing Fe2+, and accelerating Fe3+/Fe2+ cycling, leading to exacerbated oxidative and nitrosative stress on proteins, with implications not only for meat processing but also in many biological and environmental processes due to the ubiquitous presence of iron, hydrogen peroxide, and nitrite in nature.

Regenerative nanobots based on magnetic layered double hydroxide for azo dye removal and degradation.

A highly regenerative multifunctional nanobot system, using Fe3O4@SiO2@MgFe-LDH nanoparticles, is developed for efficient removal of waterborne azo dyes and pharmaceuticals. Efficient capture of pollutants, powerful Fenton degradation, and superior materials regeneration lead to a simple and cost-effective wastewater remediation solution.

Chemisorption Mechanism of DNA on Mg/Fe Layered Double Hydroxide Nanoparticles: Insights into Engineering Effective SiRNA Delivery Systems.

Layered double hydroxide nanoparticles (LDH NPs) have attracted interest as an effective gene delivery vehicle in biomedicine. Recent advances in clinic trials have demonstrated the efficacy of Mg/Fe LDHs for hyperphosphatemia treatment, but their feasibility for gene delivery has not been systematically evaluated. As a starting point, we aimed to study the interaction between oligo-DNA and Mg/Fe LDH NPs. Our investigation revealed the chemisorption mechanism of DNA on Mg/Fe LDH surfaces, wherein the phosphate backbone of the DNA polymer coordinates with the metal cations of the LDH lattice via the ligand-exchange process. This mechanistic insight may facilitate future gene delivery applications using Mg/Fe LDH NPs.

Chloride accelerated Fenton chemistry for the ultrasensitive and selective colorimetric detection of copper.

A highly selective, ultrasensitive (visual and instrumental detection limits of 40 nM and 0.1 nM, respectively), environmentally-friendly, simple and rapid colorimetric sensor was developed for the detection of copper(II) in water. This sensor is based on a novel signal-amplification mechanism involving reactive halide species (RHSs) including chlorides or bromides, which accelerate copper Fenton reactions oxidizing the chromogenic substrate to develop colour. The results of this study expand our understanding of copper-based Fenton chemistry.

Adsorption of doxorubicin on citrate-capped gold nanoparticles: insights into engineering potent chemotherapeutic delivery systems.

Gold nanomaterials have received great interest for their use in cancer theranostic applications over the past two decades. Many gold nanoparticle-based drug delivery system designs rely on adsorbed ligands such as DNA or cleavable linkers to load therapeutic cargo. The heightened research interest was recently demonstrated in the simple design of nanoparticle-drug conjugates wherein drug molecules are directly adsorbed onto the as-synthesized nanoparticle surface. The potent chemotherapeutic, doxorubicin often serves as a model drug for gold nanoparticle-based delivery platforms; however, the specific interaction facilitating adsorption in this system remains understudied. Here, for the first time, we propose empirical and theoretical evidence suggestive of the main adsorption process where (1) hydrophobic forces drive doxorubicin towards the gold nanoparticle surface before (2) cation-π interactions and gold-carbonyl coordination between the drug molecule and the cations on AuNP surface facilitate DOX adsorption. In addition, biologically relevant compounds, such as serum albumin and glutathione, were shown to enhance desorption of loaded drug molecules from AuNP at physiologically relevant concentrations, providing insight into the drug release and in vivo stability of such drug conjugates.

Robust Ag nanoplate ink for flexible electronics packaging.

Nanoinks are currently a topic of heightened interest with respect to low temperature bonding processes and printable electronics. We have developed an innovative polyvinylpyrrolidone (PVP)-stabilized Ag nanoplate ink amenable to very strong low temperature packaging, and investigated the relationship between bonding strength and electrical conductivity post-bonding. PVP shell plastic deformations observed in failure microcracks with the formation of PVP nanofibers, revealed bonding strength at low temperatures (<250 °C) was primarily due to adhesive bonding. It is found that, utilizing photonic sintering, ∼ 70 °C reduction of transformation temperature from adhesive to metallic bonding was achieved compared to that of thermal sintering. A numerical simulation was developed to better understand the influences of the light-induced heat generation, which demonstrated near-infrared light can facilitate sintering. Bonding strengths of 27 MPa were achieved at room temperatures, and 29.4 MPa at 210 °C with photonic sintering. Moreover, the anisotropic resistivity was observed with different thermal dependences. These results demonstrate Ag nanoplate inks have potential for low temperature 3D interconnections in lead-free microcircuits, flexible electronic packaging, and diverse sensing applications.

Direct writing on paper of foldable capacitive touch pads with silver nanowire inks.

Paper-based capacitive touch pads can be fabricated utilizing high-concentration silver nanowire inks needle-printed directly onto paper substrates through a 2D programmable platform. Post deposition, silver nanowire tracks can be photonically sintered using a camera flash to reduce sheet resistance similar to thermal sintering approaches. Touch pad sensors on a variety of paper substrates can be achieved with optimized silver nanowire tracks. Rolling and folding trials, which yielded only modest changes in capacitance and no loss of function, coupled with touch pad functionality on curved surfaces, suggest sufficient flexibility and durability for paper substrate touch pads to be used in diverse applications. A simplified model to predict touch pad capacitance variation ranges with differing touch conditions was developed, with good agreement against experimental results. Such paper-based touch pads have the advantage of simple structure, easy fabrication, and fast sintering, which holds promise for numerous commercial applications including low-cost portable devices where ultrathin and lightweight features, coupled with reliable bending stability are desirable.

Occurrence and degree of intersex (testis-ova) in darters (Etheostoma SPP.) across an urban gradient in the Grand River, Ontario, Canada.

The variability and extent of the intersex condition (oocytes in testes, or testis-ova) was documented in fish along an urban gradient in the Grand River, Ontario, Canada, that included major wastewater treatment plant outfalls. A method for rapid enumeration of testis-ova was developed and applied that increased the capacity to quantify intersex prevalence and severity. Male rainbow darters (Etheostoma caeruleum) sampled downstream of the first major wastewater outfall (Waterloo) had a significant increase, relative to 4 upstream reference sites, in the mean proportion of fish with at least 1 testis-oocyte per lobe of testes (9-20% proportion with ≤ 1 testis-oocyte/lobe vs 32-53% and >1.4 testis-oocyte/lobe). A much higher mean incidence of intersex proportion and degree was observed immediately downstream of the second wastewater outfall (Kitchener; 73-100% and 8-70 testis-oocyte/lobe); but only 6.3 km downstream of the Kitchener outfall, the occurrence of intersex dropped to those of the reference sites. In contrast, downstream of a tertiary treated wastewater outfall on a small tributary, intersex was similar to reference sites. Estrogenicity, measured using a yeast estrogen screen, followed a similar pattern, increasing from 0.81 ± 0.02 ng/L 17b-estradiol equivalents (EEq) (Guelph), to 4.32 ± 0.07 ng/L (Waterloo), and 16.99 ± 0.40 ng/L (Kitchener). Female rainbow darter downstream of the Kitchener outfall showed significant decreases in gonadosomatic index and liver somatic index, and increases in condition factor (k) relative to corresponding reference sites. The prevalence of intersex and alterations in somatic indices suggest that exposure to municipal wastewater effluent discharges can impact endocrine function, energy use, and energy storage in wild fish.

Fish community responses to multiple municipal wastewater inputs in a watershed.

Municipalities utilize aquatic environments to assimilate their domestic effluent resulting in eutrophication, anoxia, toxicity and endocrine disruption of aquatic biota. The objective of this study was to assess the potential cumulative impacts of municipal wastewater effluent (MWWE) discharges in the Grand River on the health status of a sentinel species and the fish community downstream of 2 MWWE discharges. The fish communities downstream of the MWWE outfalls demonstrated differences in the abundance and diversity, species and family richness, % tolerance and % vulnerability when compared to the fish community upstream or further downstream of these points of effluent discharge. In both years studied, the fish community exposed to MWWE in the riffle-run habitats demonstrated reductions in the proportion of the most prominent fish (Rainbow Darter, Ethoestoma caeruleum) downstream of the outfalls, and a significant increase in the proportion of large mobile, tolerant-omnivorous fish species such as suckers and sunfish. There was less variability in the responses of the fish community to MWWE in the same season between years than between seasons within the same year. An examination of how impaired health of a sentinel species exposed to MWWE discharges parallels changes in the fish community is also conducted. This study successfully demonstrates the cumulative impact of urban development, including multiple outfalls of treated wastewater effluents on fish populations and communities. Municipalities are the major source of nutrients and pharmaceuticals and personal care products to aquatic systems, and they need to consider their impacts carefully with increasing urban population growth and ageing demographics.

Development and evaluation of a new in vivo solid-phase microextraction sampler.

The use of solid-phase microextraction (SPME) as a nonlethal technique for in vivo sampling of pharmaceutical residue in fish tissue has been documented in the literature. However, there is need to improve its simplicity and robustness for wider applications in the laboratory and field. The objective of this research is to develop and improve the SPME device for sampling of pharmaceuticals in fish tissue. The practical application of the new device was demonstrated in the field where some wild fish (Esox masquinongy) were caught in the river and sampled by the device. The samples were analyzed using LC coupled with MS/MS (LC-MS/MS). The new in vivo SPME device with a PDMS extraction phase (sorbent) was demonstrated to a robust tool by both experts and nonexpert of the method and it is simpler than the traditional device. The detection limit of the method in gel and fish tissue was 0.01-0.26 ng/g. The interday reproducibility in gel and fish homogenized fish tissue was 8-16% RSD. This study demonstrates that the new device will provide a platform or opportunity for rapid sampling of carbamazepine, diazepam, and nordiazepam in fish muscle with acceptable precision.

Determination of selected pharmaceutical residues in wastewater using an automated open bed solid phase microextraction system.

The detection of trace levels of pharmaceuticals in environmental matrices requires an analyte pre-concentration procedure to obtain the required sensitivity for quantitative determination. This research aims to develop a simple automated analytical method based on C(18) thin film solid phase microextraction (TF-SPME) for the simultaneous extraction of pharmaceutical compounds detected in surface waters. As a sample preparation method, solid phase microextraction, is a rapid, environmentally friendly, and a sensitive analytical technique which isolates and pre-concentrates trace organic pollutants from environmental water samples in a single step. High throughput analysis was achieved with the use of a robotic auto sampler which enabled parallel analyte extraction in a 96-well plate format. Application of the method was demonstrated using wastewater from pilot-scale municipal treatment plants and environmental water samples from wastewater-dominated reaches of the Grand River (adjacent Waterloo, ON) which were analysed using a liquid chromatography-mass spectrometry (LC-ESI-MS/MS) technique. The proposed method successfully determined concentrations of carbamazepine, fluoxetine, sertraline, and paroxetine in treated effluent at concentrations ranging from 240 to 3820 ng/L with a method detection limit of 2-13 ng/L with a relative standard deviation of less than 16%. Matrix effect was not observed with this method; therefore internal standards are not necessary for quantification of target compounds. The results suggest that this method is capable of detecting and quantifying many compounds present in both wastewater and wastewater-influenced surface water from multiple municipal sources. In this study, automated TF-SPME system is demonstrated as a simple and fast alternative method for high throughput analysis of pharmaceutical contaminants in environmental matrices.

Optimization of solid phase microextraction for non-lethal in vivo determination of selected pharmaceuticals in fish muscle using liquid chromatography-mass spectrometry.

A new thin film microextraction (TFME) configuration based on C(18) thin film was optimized to improve solid phase microextraction (SPME) sensitivity and extraction kinetics for in vivo determinations of trace pharmaceuticals in fish tissue. Optimization of SPME involved increasing sampler surface area and volume of extraction phase to improve performance within in vitro applications such as agarose gel matrix and spiked fish tissue, as well as in vivo determinations of pharmaceuticals in fish exposed to municipal wastewater. Rainbow trout (Oncorhynchus mykiss) were exposed for 4d to effluents from a municipal wastewater plant that was treated under three different configurations of a pilot plant. Fathead minnow (Pimephales promelas) were caged for 14 d upstream and downstream of municipal wastewater effluent discharges along Grand River at near Kitchener, ON. TFME and regular C(18) fibers were inserted into the dorsal-epaxial muscle of rainbow trout and fathead minnow, respectively, for 30 min sampling intervals. Sample extracts obtained from fibers were desorbed in methanol:water (3:2) for 90 min under 1500 rpm agitation and analyzed using liquid chromatography coupled with tandem mass spectrometry (LC/MS-MS) to determine pharmaceutical bioconcentration. Most target pharmaceuticals were detected in the wastewater with the exception of norfluoxetine and paroxetine. C(18) TFME phase successfully quantified fluoxetine, venlafaxine, sertraline, paroxetine, and carbamazapine in muscle of living fish at concentrations ranging from 1.7 to 259 ng/g. Reproducibility of the method in spiked fish muscle was 9-18% RSD with limits of detection and quantification ranging from 0.08 to 0.21 ng/g and 0.09 to 0.64 ng/g (respectively) for the analytes examined. Ibuprofen and gemfibrozil were not detected in fish tissues, likely due to rapid excretion leading to low rates of bioconcentration. This study demonstrates improved in vivo SPME sensitivity (recovery) and extraction rates during pre-equilibrium sampling of target pharmaceuticals in fish tissue due to the improved C(18) extraction phase geometry.

Depth-profiling of environmental pharmaceuticals in biological tissue by solid-phase microextraction.

The parallel in vivo measurement of chemicals at various locations in living tissues is an important approach furthering our understanding of biological uptake, transportation, and transformation dynamics. However, from a technical perspective, such measurements are difficult to perform with traditional in vivo sampling techniques, especially in freely moving organisms such as fish. These technical challenges can be well addressed by the proposed depth-profiling solid-phase microextraction (DP-SPME) technique, which utilizes a single soft, flexible fiber with high spatial resolution. The analytical accuracy and depth-profiling capability of DP-SPME was established in vitro within a multilayer gel system and an onion artificially contaminated with pharmaceuticals. In vivo efficacy was demonstrated by monitoring pharmaceutical distribution and accumulation in fish muscle tissue. The DP-SPME method was validated against pre-equilibrium SPME (using multiple small fibers), equilibrium SPME, and liquid extraction methods; results indicated DP-SPME significantly improved precision and data quality due to decreased intersample variation. No significant adverse effects or increases in mortality were observed in comparisons of fish sampled by DP-SPME relative to comparable fish not sampled by this method. Consequently, the simplicity, effectiveness, and improved precision of the technique suggest the potential for widespread application of DP-SPME in the sampling of heterogeneous biotic and abiotic systems.

Temporal changes in stress and tissue-specific metabolic responses to municipal wastewater effluent exposure in rainbow trout.

Sub-chronic exposure to municipal wastewater effluent (MWWE) in situ was recently shown to impact the acute response to a secondary stressor in rainbow trout (Oncorhynchus mykiss). However, little is known about whether MWWE exposure in itself is stressful to the animal. To address this, we carried out a laboratory study to examine the organismal and cellular stress responses and tissue-specific metabolic capacity in trout exposed to MWWE. Juvenile rainbow trout were exposed to 0, 20 and 90% MWWE (from a tertiary wastewater treatment plant), that was replenished every 2d, for 14 d. Fish were sampled 2, 8 or 14 d post-exposure. Plasma cortisol, glucose and lactate levels were measured as indicators of organismal stress response, while inducible heat shock protein 70 (hsp70), constitutive heat shock protein 70 (hsc70) and hsp90 expression in the liver were used as markers of cellular stress response. Impact of MWWE on cortisol signaling was ascertained by determining glucocorticoid receptor protein (GR) expression in the liver, brain and, heart, and metabolic capacity was evaluated by measuring liver glycogen content and tissue-specific activities of key enzymes in intermediary metabolism. Plasma glucose and lactate levels were unaffected by exposure to MWWEs, whereas cortisol showed a transient increase in the 20% group at 8d. Liver hsc70 and hsp90, but not hsp70 expression, were higher in the 90% MWWE group after 8d. There was a temporal change in GR expression in the liver and heart, but not brain of trout exposed to MWWE. Liver glycogen content and activities liver gluconeogenic enzyme phosphoenolpyruvate carboxykinase (PEPCK), and alanine aminotransferase (AlaAT) were significantly affected by MWWE exposure. The glycolytic enzymes pyruvate kinase (PK) and hexokinase (HK) activities were significantly higher temporally by MWWE exposure in the gill and heart, but not in the liver and brain. Overall, a 14 d exposure to MWWE evokes a cellular stress response and perturbs the cortisol stress response in rainbow trout. The tissue-specific temporal changes in the metabolic capacity suggest enhanced energy demand in fish exposed to MWWE, which may eventually lead to reduced fitness.

Determination of pharmaceutical residues in fish bile by solid-phase microextraction couple with liquid chromatography-tandem mass spectrometry (LC/MS/MS).

The present study investigates possible uptake and bioconcentration of different classes of pharmaceuticals residues (organic contaminants) in fish bile using a simplified analytical methodology based on solid phase microextration (SPME). The use of solid phase microextraction (SPME), as a simple analytical tool, to screen for target pharmaceuticals in fish bile samples was validated in rainbow trout (Oncorhynchus mykiss) following short-term laboratory exposures to carbamazepine and fluoxetine. While fish bioconcentrated both fluoxetine and carbamazepine from exposure water, fluoxetine accumulated to a greater degree in bile than carbamazepine. Good agreement was obtained for both analytes in bile samples between SPME and traditional liquid (solvent) extraction approaches (R(2) > 0.99). The field application of SPME sampling was further demonstrated in fathead minnow (Pimephales promelas), a small-bodied fish caged upstream and downstream of a local wastewater treatment plant where fluoxetine, atorvastatin, and sertraline were detected in fish bile at the downstream location. SPME is a promising analytical tool for investigating the bioconcentration of trace contaminants in fish bile, facilitating detection of trace environmental contaminants otherwise undetectable due to low concentrations in the environment and biological tissues as well as the complexity of the sample matrices.

PCR-ready human DNA extraction from urine samples using magnetic nanoparticles.

Urine-derived human genomic DNA (gDNA) has wide application in a variety of disciplines including clinical medicine, sports, and forensic science. We describe a novel method for gDNA extraction from urine samples using carboxylated magnetic nanoparticles (CMNPs) as solid-phase adsorbents. Sedimentation associated with freezing of urine samples significantly reduces cell capture by CMNPs. However, the addition of 10 mM EDTA and subsequent pH modification (pH 6.0-7.1) can re-dissolve urine sediments. Purified gDNA ranged from around 0.1 kb to more than 23 kb. PCR using specific primers targeting K-ras, GAPDH, CYP3A4 and GDF5 amplified 100% of varying sized gene fragments, verifying the high quality of the isolated DNA. Successful PCR amplifications using DNA isolated from urine samples as small as 50 µl were demonstrated. Enrichment of urine cells and subsequent adsorption of DNA can be achieved with the same CMNPs, greatly simplifying extraction procedures. The CMNP gDNA extraction technique proved to be simple, rapid, sensitive and environmentally friendly, with application for routine laboratory use and potentially within automated urine extraction platforms.