High Content Samples Research Articles

Evaluation of a Highly Efficient Multidrug Biochip Array Technology for a Simultaneous and High-Throughput Urine Drug Screening in Clinical and Toxicological Settings.

A high-throughput and highly efficient analytical platform for urine drug screening is critical in both clinical and forensic settings. Mass spectrometry (MS) has better sensitivity and specificity than conventional immunoassays (IA); however, not all laboratories have the necessary resources and workforce to operate MS. The goal of this study was to evaluate a multidrug biochip with 20 discrete testing regions (DTRs) for high-throughput urine drug screening (UDS). The Randox DOA Ultra Urine (DOAULT URN) biochip employs chemiluminescent IA to detect various analytes, including stimulants, hallucinogens, sedatives, narcotics, and dextromethorphan. The verification included the evaluation of the limits of detection (LOD), stability of calibrators and controls, cross-reactivity, carryover, interference, and overall performance. LODs < quality control low for each DTR. The reconstituted calibrators were stable for up to 2 weeks at -20°C. Controls were stable for 4-6 hours at 22-25°C, with <20% within-day and ≤23% between-day imprecision. The accuracy of the controls (%bias) was within ±20% of the target concentration, except for dextromethorphan at -23.8%. No interference was observed with common over-the-counter medications. No carryover was detected in the high-concentration samples. Satisfactory cross-reactivity (≥50%) with known analytes produced presumptive positive results, with readings higher than the proposed decision points. The overall biochip performance of 165 confirmed samples showed 98.0% sensitivity, 96.9% specificity, and 97.5% efficiency. The DOAULT URN biochip is a multidrug analyte IA capable of detecting dozens of parent drugs and their metabolites in urine. It offers clinical and forensic laboratories an alternative UDS tool with LODs comparable to those of MS.

Commutability Assessment of Processed Human Plasma Samples for Normetanephrine and Metanephrine Measurements Based on the Candidate Reference Measurement Procedure.

BackgroundTo identify candidate external quality assessment (EQA) materials for normetanephrine and metanephrine measurements, we assessed the commutability of eight processed human plasma samples. The agreement between routine assays and the candidate reference measurement procedure (cRMP) was also evaluated.MethodsFifty-three clinical samples and eight processed plasma samples were prepared. The processed samples included pooled and individual plasma samples spiked with pure normetanephrine and metanephrine and non-spiked pooled and individual plasma samples. The clinical and processed samples were subjected to four routine isotope dilution tandem mass spectrometry assays and cRMP. Commutability was assessed based on two approaches recommended by the CLSI and International Federation of Clinical Chemistry (IFCC). Passing–Bablok regression and Bland–Altman analysis were used to evaluate the agreement between the routine assays and cRMP.ResultsThe commutability results of the CLSI approach were better than those of the IFCC approach. For the CLSI approach, spiked individual plasma samples and spiked high-concentration pooled plasma samples were commutable for all routine assays for both analytes. The non-spiked pooled plasma sample was commutable for two out of four routine assays for metanephrine and three out of four routine assays for normetanephrine. The agreement between the routine assays and the cRMP was satisfactory, except for one routine assay showing significant bias.ConclusionsHigh-concentration spiked pooled plasma samples and spiked individual plasma samples are candidate EQA materials for normetanephrine and metanephrine measurements.

Quantum transport evidence of boundary states and Lifshitz transition in Bi4Br4

The quasi-one-dimensional van der Waals compound ${\mathrm{Bi}}_{4}{\mathrm{Br}}_{4}$ was recently found to be a promising high-order topological insulator with exotic electronic states. In this paper, we study the electrical transport properties of ${\mathrm{Bi}}_{4}{\mathrm{Br}}_{4}$ bulk crystals. Two electron-type samples with different electron concentrations are investigated. Both samples exhibit a saturation behavior of resistivity in low temperature. In the low-concentration sample, two-dimensional quantum oscillations are clearly observed in the magnetoresistance measurements, which are attributed to the band-bending-induced surface state on the (001) surface. In the high-concentration sample, the angular magnetoresistance exhibits two pairs of symmetrical sharp valleys with an angular difference close to the angle between the crystal planes (001) and (100). The additional valley can be explained by the contribution of the boundary states on the (100) facet. Besides, Hall measurements at low temperatures reveal an anomalous decrease of electron concentration with increasing temperature, which can be explained by the temperature-induced Lifshitz transition. These results shed light on the abundant boundary state transport signals and the temperature-induced Lifshitz transition in ${\mathrm{Bi}}_{4}{\mathrm{Br}}_{4}$.

Green Synthesis of Bioactive Hydroxyapatite/Cellulose Composites from Food Industrial Wastes

This work aimed at conversion of worthless indurtial wastes to valuable product. Herein, bioactive composites based on bacterial cellulose (BC) and eggshell or eggshell-derived hydroxyapatite (HAp) were prepared by a green method using Gluconacetobacter xylinum bacteria. The effect of addition of eggshell (BC/Eg) and eggshell-derived HAp (BC/HAp-Eg) on the bacterial cellulose yield, biodegradation and biocompatibility was studied. For comparison, HAp derived from chemical precursors was synthesized (BC/HAp-ch). The resultant composites were characterized by XRD, FTIR, and SEM/EDX. Furthermore, the biodegradation and bioactivity were assessed in SBF, and the cell viability was studied against oral normal cells. The results showed that the productivity of BC applied HAp-derived eggshell (1.83 g/L) was higher than that of using (1.37 g/L). Interestingly, the eggshell was converted to Ca3(PO4)2 during incubation in the bacterial culture medium, while Ca3(PO4)2 was formed as a secondary phase when using either eggshell-derived HAp or chemically-derived. The in vitro bioactivity test in SBF showed that all composites were induced the formation of a bone-like apatite layer on their surface with Ca/P ratio, 1.49, 1.35, and 1.41 for BC/Eg, BC/HAp-ch, and BC/HAp-Eg, respectively, near to the ratio in the natural HAp. Finally, the in vitro cell viability test was confirmed good biocompatibility against the composites. However, at high sample concentration (250 µg/mL), BC/HA-Eg showed the higher cell viability (95.2%) than that of BC/Eg (80.5%) and BC/HA-ch (86.2%). In conclusion, eggshell waste could be used directly with bacterial cellulose to produce bioactive composites without the need to convert it to HAp which reduced the cost of production and thus has a higher economic return. Obiviously, eggshell waste can act as calcium, organic matter source, pH preservation, nuterilizing agent along with potential instead of costly buffering agent in the BC culture medium and further for increased the BC production.

Evaluation of analytical performances of a serum acetaminophen assay designed for round-the-clock use

Acetaminophen (paracetamol) voluntary (or not) intoxications are a major cause of life-threatening liver damage. Most of available assays on the market show an upper range measurement level that is close to thresholds considered as toxic levels. Therefore, high concentration samples must be further diluted in to accurately measure paracetamol levels. This study aims at assessing the performances of a Cambridge life sciences (CLS) test for acetaminophen determination in human serum, which solves this issue. Indeed, the particularities of this test consist in (i) an extended announced upper limit of quantitation, and (ii) in the use of a genetically unique enzyme (Aryl acylamidase). This evaluation encompasses comparison of results with those obtained using the acetaminophen Thermofisher test currently used in the University Hospital of Lille, and here after referred to as “reference method”. Analytical performance was evaluated on a Beckman Coulter AU480 chemistry analyzer. Within-run precision was determined by assaying 3 serum samples at 3 concentration levels (low, medium, high) in 10 replicates and between-run precision in running 5 samples on 6 days, twice a day. Linearity was assessed against a set of standards covering the entire claimed linearity range. Fifty-two fresh serum samples form patients were prospectively assessed for correlation to the reference method. The within-run and between-run precision (%CV) was respectively < 1% and < 3%. The assay showed good linearity with very low CV < 2% at concentrations up to 859 mg/L, consistent with the analytical range announced by the manufacturer. The regression equation against the reference method was Y = 1.133 x + 0.145, r2 = 0.99988. However, the lower limit of quantification (LLOQ) appears to be slightly higher than the manufacturer's stated (3 mg/L). Comparison to reference method showed that values given by the CLS assay were overestimated especially in the low range of concentration values (< 50 mg/L). The CLS acetaminophen assay has proven to have good analytical performance. Non-correlation to reference method does not affect the ability of the test to detect and to guide the management of paracetamol poisoning. The major advantage of CLS acetaminophen assay is the broad linearity range covering high acetaminophen concentrations. The use of this assay eliminates the need for dilution of highly positive samples and is particularly designed for round-the-clock use.

Protein recovery and quality of alfalfa extracts obtained by acid precipitation and fermentation

Alfalfa has a high potential as a novel food protein and can contribute to meet the increasing food demand. To facilitate food grade quality, protein extraction is essential. In this study, different extractions with isoelectric focusing the protein were compared; acid precipitation (citric and lactic acids) and fermentation (endogenous and Lactobacillus salivarius). Acid precipitation resulted in the highest protein recovery (30 %) compared with L. salivarius and endogenous fermentation. The protein ribulose-1.5-bisphosphate carboxylase/oxygenase (RuBisCO) was found intact and in high concentrations in acid precipitated samples. With L. salivarius, a higher content of peptides and possible cross-linked proteins were discovered. However, for both extractions, the amino acid composition showed a promising distribution meeting the recommendation of the Food and Agriculture Organization (FAO). Overall, these results indicate that protein extracts obtained with acid precipitation might be more appropriate as a food ingredient, whereas fermented products might have potential as food supplement.

Toward protein NMR at physiological concentrations by hyperpolarized water-Finding and mapping uncharted conformational spaces.

Nuclear magnetic resonance (NMR) spectroscopy is a key method for determining the structural dynamics of proteins in their native solution state. However, the low sensitivity of NMR typically necessitates nonphysiologically high sample concentrations, which often limit the relevance of the recorded data. We show how to use hyperpolarized water by dissolution dynamic nuclear polarization (DDNP) to acquire protein spectra at concentrations of 1 μM within seconds and with a high signal-to-noise ratio. The importance of approaching physiological concentrations is demonstrated for the vital MYC-associated factor X, which we show to switch conformations when diluted. While in vitro conditions lead to a population of the well-documented dimer, concentrations lowered by more than two orders of magnitude entail dimer dissociation and formation of a globularly folded monomer. We identified this structure by integrating DDNP with computational techniques to overcome the often-encountered constraint of DDNP of limited structural information provided by the typically detected one-dimensional spectra.

Real-Time Underway Mapping of Nutrient Concentrations of Surface Seawater Using an Autonomous Flow Analyzer.

High-frequency field nutrient analyzers offer a promising technology to solve time-consuming and laborious sampling problems in dynamic and complex river-estuarine-coastal ecosystems. However, few studies on the simultaneous underway analysis of five key nutrients (ammonium, nitrite, nitrate, phosphate, and silicate) in seawaters are available because of the limitations of the technique. In this study, a state-of-the-art autonomous portable analyzer for the shipboard analysis of nutrients in the environment of varied salinities and concentration ranges was reported. The analyzer consisted of compact hardware that was well suited for shipboard deployment with minimal maintenance. Moreover, a novel LabVIEW-based software program was developed, containing additional functions such as automated calibration curve generation, autodilution of high-concentration samples, and a user-friendly interface for multiparameter analysis using a single instrument. After the optimization of chemical reactions and work flow chart, the analyzer exhibited low limits of detection, a large linear range with automated dilution, and relative standard deviations of less than 2% (n = 11). Compared to other flow-based techniques, this analyzer is more portable and consumes less reagent with an autonomous data processing function and applicability within a broad salinity range (0-35). The analyzer was successfully applied for real-time analysis in the Jiulong River Estuary-Xiamen Bay with excellent on-site accuracy and applicability. The relationship between high spatial resolution nutrient concentrations and salinities showed very different patterns in estuarine and coastal areas, indicating the benefit of using an underway automated analyzer for chemical mapping in a dynamic environment.

An external quality assessment for the molecular testing of the SARS-CoV-2 virus genome in Zhejiang Province, China

The COVID-19 pandemic has necessitated the rapid expansion of laboratories that conduct SARS-CoV-2 tests. A provincial external quality assessment (EQA) scheme on SARS-CoV-2 tests was organized by Zhejiang Provincial CDC to assess the accuracy of the tests in individual CDC municipal and county laboratories in Zhejiang Province, China. Three positive samples in high, medium, and low concentrations, respectively, were prepared using the serial dilutions from the culture with the viral titer concentration of 1×106.3 TCID50/mL, and one negative sample were included. A total of 93 laboratories participated, contributing results from 36 distinct combinations of nucleic acid extraction methods and PCR reagents. There was 100% concordance among all laboratories for all EQA samples, and no false-positive or false-negative results were observed. The EQA survey provides confidence in the identification of infected individuals or asymptomatic populations and assurance for clinical and public health decision-making based on test results.

Development of magnetic molecularly imprinted solid-phase extraction and ultra-high performance liquid chromatography tandem mass spectrometry for rapid and selective determination of urinary diphenyl phosphate of college students

Organophosphate esters (OPEs), known as novel alternative flame retardants, are a class of environmental endocrine disruptors. Long-term exposure to OPEs may bring a non-negligible health risk to human. Urinary OPE metabolites (mOPEs) are generally used as biomarkers to evaluate the internal exposure to OPEs. Diphenyl phosphate (DPHP), the main metabolite of aryl-OPEs, exhibited high detection rates and concentrations in urine samples. To establish a selective and simple analytical method for biomonitoring urinary DPHP, a specific magnetic molecular imprinted polymer (MMIP) was fabricated via a sol-gel method. Under optimum magnetic solid-phase extraction (MSPE) conditions, the resultant MMIP exhibited selective recognition ability, ideal adsorption capacity and good reusability on urinary DPHP enrichment. The developed MSPE method coupled with ultra-high performance liquid chromatography tandem mass spectrometry (U-HPLC-MS/MS) exhibited good precision and accuracy (spiked recoveries of 85.8%-109% with relative standard deviations (RSDs) ranged from 5.1%-13%), low detection limit of 0.035 ng/mL, and negligible matrix inhibition. Then we used this proposed method to detect urinary DPHP levels of recruited 30 college students and investigate the time variability and potential determinants. All urine samples revealed the presence of DPHP at a median concentration of 0.56 μg/g Creatinine (Cr). Moderate reproducibility of DPHP level was observed in first morning urine samples (ICC>0.40). Significant correlations were found between urinary DPHP levels and gender (β=0.72; 95% CI: 0.48∼0.96), sampling time (β=0.36; 95% CI: 0.08∼0.65) as well as the frequency for take-out food (β=0.45; 95% CI: 0.07∼0.74) (p< 0.05). Hence, a fast and sensitive MSPE-U-HPLC-MS/MS method was successfully built to quantify urinary DPHP.

Occurrence and distribution of triclosan and its transformation products in Taihu Lake, China.

The transformation products of triclosan (5-chloro-2-(2,4-dichlorophenoxy)phenol) may be more persistent and toxic than their parent compound, yet their occurrence in aquatic environments is poorly understood. In this study, we identified three transformation products in sediment samples from Taihu Lake and compared their concentrations with the parent compound triclosan. Triclosan in Taihu Lake was at low level, ranging from 0.086 to 1.1 ng/L in surface water and 0.0058-8.3 ng/g in sediments. The three detected transformation products included methyl triclosan, chlorinated triclosan derivatives, and methyl chlorinated triclosan derivatives. Those transformation products constituted 0.73-87.5% of the total triclosan (total triclosan is the sum of triclosan, methyl triclosan, chlorinated triclosan derivatives, and methyl chlorinated triclosan derivatives on a molar basis), indicating that the ecological risk of transformation products should be considered in addition to the parent compound. Different transformation products had distinct spatial distributions. Chlorinated triclosan derivatives had the highest concentration in samples from the northwest region (0.016-0.21 ng/g) of the lake and were positively correlated with triclosan, which may indicate the possible transformation from triclosan to chlorinated triclosan derivatives. Methyl triclosan and methyl chlorinated triclosan derivatives were generally higher in samples from the center of the lake (0.22-0.28 ng/g for methyl triclosan and 0.017-0.021 ng/g for methyl chlorinated triclosan derivatives, respectively), indicating the possible occurrence of in situ microbial methylation of triclosan and chlorinated triclosan derivatives and the accumulation of those methylated analogues in Taihu Lake.

Risk assessment of pesticide residues in some samples of carrots (&lt;em&gt;Daucus carota&lt;/em&gt;)

Background and Objective: The production of safe foods for consumption is an important issue worldwide. Pesticide application is being used in recent agricultural practices to cultivate crops. Their use can have serious impacts on the health of humans. This research work aims to assess pesticide residues in carrot samples from Kaduna and Plateau States in Northern Nigeria and their risk to human health. Materials and Methods: Ten samples of carrots were collected in two groups from Kaduna and Plateau States and analyzed for pesticide residues. Extraction was performed and analysis was performed using gas chromatography hypherated to a mass spectrophotometer. Risk assessment was carried out by determining the health risk index. Results: Results of the analysis showed that nineteen pesticides were detected. Organochlorines were the highest with Trans-nonachlor appearing in nine of the samples with the highest mean concentration being 0.064±0.003 mg/kg in samples from Plateau State and the lowest being 0.022±0.004 mg/kg in samples from Kaduna State. Organophosphates were also detected with high concentrations in samples from Plateau State with a mean concentration of 0.034±0.003 mg/kg and the lowest being 0.000±0.0001 mg/kg. The risk index was

Antimicrobial, Antioxidant and Phytotoxic Assessment of Agave Americana, Mentha Spicata and Mangifera Indica L. Extract

Purpose: The research study was prototyped to assess pharmacological activities (antimicrobial, anti-oxidant, phytotoxic and haemagglutination) of Agave americana, Mentha spicata and Mangifera indica leaves extracts. The positive outcomes of the study can aid in formulations of novel therapeutic opportunities by using natural, eco-friendly, and economic raw materials having least side effects and aced potency. Method: Leaves extracts of Agave americana, Mentha spicata and Mangifera indica were prepared using analytical grade ethanol, methanol, acetone and n-hexane. Then the extracts were subjected to antibacterial, antifungal, antioxidant, phytotoxic and haemagglutination assay. Well-diffusion antibacterial protocol was followed against pathogenic bacteria. Tube dilution antifungal protocol was followed against pathogenic fungi. DPPH radical scavenging assay at variable sample dilutions was followed for antioxidant assay. Lemna minor lethality assay was followed for phytotoxic assay. Haemagglutination assay were assessed against ABO blood groups. Results: Crude leaves extracts (aqueous, acetone, methanolic and ethanolic) of preferred plants i.e., Agave americana, Mentha spicata and Mangifera indica owned superlative antimicrobial efficacy against all test micro-organisms specifically Escherichia coli ((91%), Pseudomonas aeroginosa (85%), Methicillin resistant Staphylococcus aureus (81%) and mycelium of Fusarium oxysporum (85%) was halted by Agave americana leaves extracts. In comparison to these crude extracts of Mentha spicata terrifically halted growth of Proteus mirabilis (80%) while leaves extracts Mangifera indica halted growth of Streptococcus griseus (90%) respectively. Excellent antioxidant and phytotoxic efficacy were recorded at higher sample concentrations. No haemagglutination efficacy recorded. Conclusion: From the study it was concluded that leaves of Agave americana, Mentha spicata and Mangifera indica possess excellent antimicrobial potentials which can aid to formulation new antibiotics having excellent effects in least doses. These extracts can also be used to treat many maladies by eliminating reactive oxidants and promoting healthy metabolism. These extracts can also be utilized as ecofriendly and user-friendly herbicide.

Evaluation of enterovirus nucleic acids detection method based on ultra‐fast real‐time fluorescence RT‐PCR technology—A pilot study

The outbreak of COVID-19 epidemic has enabled the establishment and application of various rapid detection methods. It is particularly important to establish a fast and accurate detection method for enterovirus, which will be beneficial for clinical diagnosis, epidemic prevention and control, and timely traceability. Through establishing an ultra-fast reverse transcription-polymerase chain reaction (RT-PCR) equipment, this study aimed to evaluate the sensitivity and specificity of the testing method of enterovirus nucleic acids based on ultra-fast real-time fluorescence RT-PCR technology. A total of 61 cases were sampled, which were then transported and preserved. After the nucleic acid extraction, the nucleic acids of the same sample were tested with the enterovirus nucleic acid detection kit produced by Guangzhou Da An Gene Company and the ultra-fast RT-PCR equipment system established in this study. ABI7500Fast and Ahram biosystems S1 fast equipment were used for amplification detection. If the sample had an S-shaped amplification curve in the FAM channel and the Ct value ≤40.00, the result was positive. The sensitivity, precision, and accuracy of the detection method were then verified. This study established a novel testing method to achieve enterovirus nucleic acid detection within 24 min. The sensitivity detection limit of the method was 1.0 × 102 copies/ml. The coefficients of variation for repeated detection of the high, medium, and low concentration samples were 2.644%, 1.674%, and 4.281%, respectively, with good detection repeatability. In addition, a total of 29 cases were positive by the ultra-fast RT-PCR detection method in 61 suspected samples, which was consistent with the conventional fluorescent RT-PCR method. The established rapid detection method can greatly shorten the time for providing a detection report, which may greatly improve the efficiency of diagnosis and treatment.

Fate of selected pharmaceuticals in hospital and municipal wastewater effluent: occurrence, removal, and environmental risk assessment.

The concentrations and distribution of β-blockers, lipid regulators, and psychiatric and cancer drugs in the influent and effluent of the municipal wastewater treatment plant (WWTP) and the effluent of 16 hospitals that discharge into the wastewater treatment plant mentioned in this study at two sampling dates in summer and winter were examined. The pharmaceutical contribution of hospitals to municipal wastewater was determined. The removal of target pharmaceuticals was evaluated in a WWTP consisting of conventional biological treatment using activated sludge. Additionally, the potential environmental risk for the aquatic receiving environments (salt lake) was assessed. Beta-blockers and psychiatric drugs were detected in high concentrations in the wastewater samples. Atenolol (919 ng/L) from β-blockers and carbamazepine (7008 ng/L) from psychiatric pharmaceuticals were detected at the highest concentrations in hospital wastewater. The total pharmaceutical concentration determined at the WWTP influent and effluent was between 335 and 737 ng/L in summer and between 174 and 226 ng/L in winter. The concentrations detected in hospital effluents are higher than the concentrations detected in WWTP. The total pharmaceutical contributions from hospitals to the WWTP in summer and winter were determined to be 2% and 4%, respectively. Total pharmaceutical removal in the WWTP ranged from 23 to 54%. According to the risk ratios, atenolol could pose a high risk (risk quotient > 10) for fish in summer and winter. There are different reasons for the increase in pharmaceutical consumption in recent years. One of these reasons is the COVID-19 pandemic, which has been going on for 2 years. In particular, hospitals were operated at full capacity during the pandemic, and the occurrence and concentration of pharmaceuticals used for the therapy of COVID-19 patients has increased in hospital effluent. Pandemic conditions have increased the tendency of people to use psychiatric drugs. It is thought that beta-blocker consumption has increased due to cardiovascular diseases caused by COVID-19. Therefore, the environmental risk of pharmaceuticals for aquatic organisms in hospital effluent should be monitored and evaluated.Supplementary InformationThe online version contains supplementary material available at 10.1007/s11356-022-21131-y.

Site-specific ecological risk assessment of metal-contaminated soils based on the TRIAD approach

Ecological risk assessment based on scientific data is crucial for understanding causal relationships between chemical pollution and environmental risks. Simultaneously, a balance is required between socioeconomic factors and scientific evidence. The TRIAD approach, which incorporates three lines of evidence (LoE)—chemical (Chem-LoE), ecotoxicological (Ecotox-LoE), and ecological (Eco-LoE)—was applied in five sites of an abandoned mine for site-specific soil ecological risk assessment (SERA). In combination, the three LoEs showed that two sites had extremely high risks, one site had moderate risk, and the other site had low risk. At all sites, Chem-LoE exhibited high-integrated risk values. In Ecotox-LoE and Eco-LoE, some species were not affected despite high metal concentrations in the soil samples collected from the sites, indicating that the bioavailability of metals differed according to the physiochemical properties of the soil medium. This study is significant as multiple analyses were performed considering ecosystem structure to reduce uncertainty in SERA. The results provide information to support effective decision-making risk management to protect the soil ecosystem. Moreover, these findings will be useful in establishing policies and priorities for soil risk management.

Development and validation of a sensitive and reliable targeted metabolomics method for the quantification of cardiovascular disease-related biomarkers in plasma using ultrahigh-performance liquid chromatography-tandem mass spectrometry.

Cardiovascular disease, as a multifactorial disease caused by genetics and environment, has emerged as a leading cause of mortality. The discovery of metabolic biomarkers for the clinical diagnosis, early warning and elucidation of the molecular pathogenesis of cardiovascular disease, using metabolomics, has attracted broad interest. Therefore, this work aimed to develop a sensitive and reliable targeted metabolomics method for the quantification of cardiovascular disease-related biomarkers in plasma. The method was developed and validated using ultrahigh-performance liquid chromatography augmented with tandem mass spectrometry (UHPLC/MS/MS). The LC conditions and MS parameters were optimized using selected reaction monitoring scanning mode to high-throughput and sensitive separation, and could detect 20 metabolic biomarkers in a single experiment. And the linearity, selectivity, accuracy, precision, stability and recovery of the developed method were assessed according to the Bioanalytical Method Validation guidelines of the United States Food and Drug Administration. These quantified metabolic biomarkers are involved in pathways such as aromatic amino acid catabolism (e.g. phenylalanine, tryptophan, tyrosine), trimethylamine N-oxide (TMAO) biosynthesis (e.g. TMAO, choline, carnitine, betaine) and histidine metabolism (e.g. histidine), among others. All analytes exhibited excellent linearities with coefficients of determination greater than 0.99. Accuracies deviated by less than 15% for medium- and high-concentration samples and less than 20% for low-concentration samples, with intra- and inter-day precisions of 1.12-14.12% and 0.30-13.74%, respectively. Recoveries and stabilities also met the analysis requirements of biological samples. The targeted metabolomics method was shown to have a powerful ability to accurately analyze metabolic biomarkers, thereby providing valuable information for large-scale biomarker validation and clarifying the potential material basis of cardiovascular disease for clinical diagnosis or early warning.

Low cost hydrothermal synthesis of optically important graphene/zinc oxide nanocomposites

Graphene oxide (GO)/Zinc Oxide (ZnO) nanocomposites were synthesized by low-cost hydrothermal route under high pressure and temperature. The Samples were characterized by optical, structural, and morphological characterizations. Increase absorbance in the visible region alongwith good luminescence is observed with an increasing ZnO concentration. X-ray diffraction (XRD) studies exhibit much more intense and sharper peaks in higher concentration sample GOZn3 than in GOZn1, confirming the highly crystalline nature of Nanoparticles at a higher amount of ZnO nanoparticles inclusion in GO. The average crystal size was found to be approximately 22 nm and 30 nm in the sample with lower and moderate concentrations GOZn1 and GOZn2 respectively. FTIR spectra analysis for functional groups present in Nanocomposites indicates the presence of aliphatic compounds, hydrocarbon bonding, olefin, hydroxyl, and aliphatic nitro compounds. FESEM micrographs indicate a sheet like the structure of GO and Zinc oxide nanoparticles arranged on a sheet in a regular pattern of hexagonal structures which are in good agreement with XRD results. The size of FESEM and XRD studies is almost the same it varies from 22 nm to 30 nm.

Detection of insulin analogues and large peptides >2kDa in urine.

Insulin analogues and large bioactive peptides may be used by athletes to enhance performance and are banned by the World Anti-Doping Agency (WADA). In addition to insulin analogues, the large peptides include a structurally diverse set of peptides including analogues of growth hormone releasing hormone (GHRH), insulin-like growth factor-1 (IGF-1), and mechano-growth factor (MGF). Detection of this class of peptides is difficult due to their absorptive losses and presence at very low concentrations in urine. In this report, a high throughput method is described that allows sensitive detection of four classes of large peptides in one assay. Sample extraction is performed by ultrafiltration to concentrate the urine followed by solid phase extraction in a 96-well micro-elution plate. Peptides in the urine samples are detected on a triple quadrupole mass spectrometer coupled to standard flow liquid chromatography. The method was validated and evaluated for limit of detection, limit of identification, specificity, precision, carryover, recovery, matrix interference, and post-extraction stability. The limit of detection for insulin analogues is between 5 and 25 pg/ml and between 5 and 50 pg/ml for the other peptide classes. Specificity was good with no detection of interfering peaks in blank urine samples. Carryover from a high concentration sample was not observed and the post-extraction stability was between 77% and 107%. The method was able to detect insulin analogues in three diabetic urine samples. Increased screening for this class of peptides will improve detection and deterrence.

Ecotoxic emissions generated by illegal burning of household waste

The practice of burning household waste including different types of plastic is illegal in Hungary, still an existing problem. As environmental consequences are hardly known, this study attempts to give an initial estimation of the ecotoxicity generated during controlled combustion of different waste types. These samples included polystyrene (PS), polyethylene (PE), polypropylene (PP), polyvinyl chloride (PVC), polyurethane (PU), oriented strand board (OSB) and rag (RAG). Ecotoxicological profiling was completed using the following test battery: Vibrio fischeri bioluminescence inhibition assay, Daphnia magna immobility test and the seedling emergence assay. Also, genotoxicity of plastic waste samples was assessed using the SOS Chromotest. Concerning main pollutants in the samples, the samples could be distinguished as ‘PAH-type’ and ‘heavy metal-type’ samples. PVC, PU and PS samples showed the highest toxicity in the Vibrio and Daphnia assays. The PVC sample was characterized by an extremely high cadmium concentration (22.4 μg/L), PS, PP and PU samples on the contrary had high total PAH content. While Vibrio and Daphnia showed comparable sensitivity, the phytotoxicity assay had no response for any of the samples tested. Samples originating from the controlled burning of different plastic types such as PU, PVC, PS and PP were classified as genotoxic, PS sample showed extremely high genotoxicity. Genotoxicity expressed as SOSIF showed strong correlation with most of the PAHs detected.