Sample Pretreatment Research Articles

Electrochemical sensor based on a glassy carbon electrode modified with a 3D carbon nanoporous composite for the detection of paracetamol in pharmaceutical samples

With the continued appearance of new drugs, there is a growing interest in the development of new analytical tools to detect and quantify drugs using a simple and reliable way. In this work, we describe the development of an electrochemical sensor based on glassy carbon electrodes modified with a carbon tubular nanocomposite for the detection of paracetamol in pharmaceutical samples. Paracetamol shows a strong adsorption on the electrode surface, which makes possible its determination by adsorptive differential pulse voltammetry in 0.1 mol/L citrate–phosphate pH 5 buffer solution. Optimal conditions for dispersion of carbonaceous material and best conditions for surface adsorption of paracetamol were obtained from experimental designs based on response surface methodology. The high adsorption capacity of paracetamol on the 3D carbon nanoporous composite and the sensitivity of the electrochemical technique employed showed a synergistic effect that allowed reaching a detection limit of 30 nmol L− 1 (4.5 ppb). In addition, the reproducibility and the repeatability were 7 and 5 %, respectively. The paracetamol determination from pharmaceutical samples was performed without pre-treatment of samples. The values of paracetamol founded for different pharmaceutical samples were in a very good agreement with those values obtained from the same samples using HPLC. Therefore, the developed electrochemical sensor is a promising, inexpensive, and easy-to-use tool for the detection and quantification of paracetamol in pharmaceutical samples.

Continuous Biosensor Based on Particle Motion: How Does the Concentration Measurement Precision Depend on Time Scale?

Continuous biosensors measure concentration-time profiles of biomolecular substances in order to allow for comparisons of measurement data over long periods of time. To make meaningful comparisons of time-dependent data, it is essential to understand how measurement imprecision depends on the time interval between two evaluation points, as the applicable imprecision determines the significance of measured concentration differences. Here, we define a set of measurement imprecisions that relate to different sources of variation and different time scales, ranging from minutes to weeks, and study these using statistical analyses of measurement data. The methodology is exemplified for Biosensing by Particle Motion (BPM), a continuous, affinity-based sensing technology with single-particle and single-molecule resolution. The studied BPM sensor measures specific small molecules (glycoalkaloids) in an industrial food matrix (potato fruit juice). Measurements were performed over several months at two different locations, on nearly 50 sensor cartridges with in total more than 1000 fluid injections. Statistical analyses of the measured signals and concentrations show that the relative residuals are normally distributed, allowing extraction and comparisons of the proposed imprecision parameters. The results indicate that sensor noise is the most important source of variation followed by sample pretreatment. Variations caused by fluidic transport, changes of the sensor during use (drift), and variations due to different sensor cartridges and cartridge replacements appear to be small. The imprecision due to sensor noise is recorded over few-minute time scales and is attributed to stochastic fluctuations of the single-molecule measurement principle, false-positive signals in the signal processing, and nonspecific interactions. The developed methodology elucidates both time-dependent and time-independent factors in the measurement imprecision, providing essential knowledge for interpreting concentration-time profiles as well as for further development of continuous biosensing technologies.

Development and validation of multiresidue analysis method for biomonitoring of pesticides and metabolites in human blood and urine by LC-QToF-MS

The widespread use of pesticides and their consequential presence in the environment is a growing concern due to the harmful health effects associated with pesticide exposure. For clinical and toxicology laboratories, a method for simultaneously determining these compounds and their metabolic products in body fluids, such as blood and urine, is important. In the present study, a rapid, sensitive and simultaneous LC-QToF-MS method for detecting multiclass pesticides and metabolites in blood and urine samples has been developed and validated. Four sample preparation procedures, protein precipitation and three different variants of QuEChERS-based extraction were evaluated to find a suitable, simple, and effective sample pretreatment technique. The final optimized sample preparation method (acetonitrile; 400 μl, MgSO4; 40 mg and NaCl; 10 mg) was validated for accuracy, precision, matrix effect, recovery, stability, carryover, and dilution integrity. Analyte recoveries ranged from 75.40 to 113.54 % while accuracy was evaluated in the range of 71.41–108.26 % and precision (%RSD) in the range of 0.01 %–16.85 %. The limit of quantification (LOQ) for all compounds was established in the range of 0.82–7.05 ng mL-1. The developed reliable, robust, and sensitive method was successfully applied for the quantification of target pesticides and metabolites in human blood and urine samples. Evaluated samples resulted in detection of eleven analytes (seven pesticides and four metabolites).

Manually operated microtube automatic capper/decapper system for clinical laboratory and biological laboratory personnel

Polymerase chain reaction (PCR) is an effective method for diagnosing infectious diseases and has been the primary method throughout the novel coronavirus disease (COVID-19) pandemic. PCR tests (from specimen collection to result acquisition) involve sample pretreatment, nucleic acid extraction, and PCR procedure. Automating the pretreatment process is crucial to mitigate the risk of infection for workers and to reduce the likelihood of sample contamination-triggered misdiagnosis, particularly when handling centrifuge tubes, cryopreservation tubes, and microtubes. Robotic systems have been engineered to automate cell culture and PCR-based diagnosis, predominantly designed for use with screw-capped containers. However, this leaves a notable gap in automation solutions for microtubes equipped with press-type caps. To address this gap, we developed a versatile microtube capper/decapper system. On the other hand, many tasks of manual operation using microtubes, which are routinely conducted in clinical tests and biological experiments, are performed. Compared to screw-type caps for centrifuge and cryopreservation tubes, press-type caps for microtubes present a considerably higher risk of the worker's fingers contacting the inside of the cap and/or generating airborne droplets. Despite the risks of contamination and infection derived from the manual handling of microtube caps, which can compromise diagnosis/experiment accuracy and worker safety, devices for manually opening and closing microtube caps without direct contact remain lacking. Therefore, leveraging the technology from the developed versatile microtube capper/decapper system for laboratory automation, we created a manually operated microtube equipped with an automatic capper/decapper system tailored for personnel in clinical and biological laboratories.In this study, we first examined the required specifications and prerequisites for a manual microtube capper/decapper and clarified the operating methods, operating procedures, operation environment, device size, accompanying functions, etc. Based on the required specifications and preconditions, we proceeded with the mechanical and control design of the conceptual model, manufactured a prototype, and confirmed its basic functions and performance. The compliant to the required specifications and preconditions and the usefulness of the proposed manual microtube capper/decapper were validated through various experiments and demonstrations. Using the proposed microtube capper/decapper, even small-scale operations, which are challenging to streamline, can be performed nearly as efficiently as full manual operations. Although operation time was not reduced, the ability to open and close microtubes without manual contact is crucial for improving diagnostic and experimental accuracy and for reducing the burden on and enhancing the safety of laboratory personnel. Because microtubes are used in various clinical tests and biological experiments, we believe that the proposed system can markedly reduce the workload for personnel across numerous clinical and biological laboratories.

Development of an Activity-Based Ratiometric Electrochemical Switch for Direct, Real-Time Sensing of Pantetheinase in Live Cells, Blood, and Urine Samples.

Pantetheinase is a key biomarker for the diagnosis of acute kidney injury and the monitoring of malaria progression. Currently, existing methods for sensing pantetheinase, also known as Vanin-1, show considerable potential but come with certain limitations, including their inability to directly sense analytes in turbid biofluid samples without tedious sample pretreatment. Here, we describe the first activity-based electrochemical probe, termed VaninLP, for convenient and specific direct targeting of pantetheinase activity in turbid liquid biopsy samples. The probe was designed such that cleavage of the pantetheinase amide linkage, triggered by a self-immolative reaction, simultaneously ejects an amino ferrocene reporter. Among the distinctive properties of the VaninLP probe for sensing pantetheinase are its high selectivity, sensitivity, and enzyme affinity, a wide linear concentration range (8-300 ng/mL), and low limit of detection (2.47 ng/mL). The designed probe precisely targeted pantetheinase and was free of interference by other electroactive biological species. We further successfully applied the VaninLP probe to monitor and quantify the activity of pantetheinase on the surfaces of HepG2 tumor cells, blood, and urine samples. Collectively, our findings indicate that VaninLP holds significant promise as a point-of-care tool for diagnosing early-stage kidney injury, as well as monitoring the progression of malaria.

Phosphate Ion-Selective Electrode Based on Electrochemically Modified Iron.

The significance of the detection of phosphate ions is immense in the realms of chemistry, biology, medicine, environment, and industry. The detection of phosphate ions is currently mainly reliant on blue molybdenum colorimetry, which is accurate but requires sample pretreatment, intricate operation, and a high price tag. Consequently, it is essential to create a sensor with superior efficiency, precision, straightforward functioning, and instantaneous online detection. This study has designed and created an electrochemical modification based on an iron metal electrode for this purpose. Cyclic voltammetry was used to initially ascertain the potential (-0.57 V) necessary for constant potential electrolysis. Employing a constant potential electrolysis method, the iron oxide and its phosphate were modified onto the surface of the iron electrode to enable reaction with phosphate ions. Scanning electron microscopy and energy dispersive X-ray spectroscopy were used to characterize and analyze the morphology and elemental composition of Fe-PME, elucidating how it responds to phosphate ionation. The two-electrode system was then utilized for the evaluation of the phosphate ion response of Fe-PME at pH 4. Fe-PME's response to phosphate ions is demonstrated by the results, ranging from 10-5 to 0.1 M and with a slope of -52.8 mV dec-1. Fe-PME exhibited satisfactory results when compared to the conventional blue colorimetry of molybdenum.

Rapid measurement of ethyl carbamate in Chinese liquor by fast gas chromatography photoionization-induced chemical ionization mass spectrometry

As a common by-product during the production of alcoholic beverages, such as Chinese liquor, ethyl carbamate (EC) poses potential genotoxicity and is associated with the risk of various cancers. Hence, rapidly and accurately measuring the content of EC in liquor is critical to assess the product quality and risks of mass samples during the production process. In this study, a feasible method based on fast gas chromatography photoionization-induced chemical ionization mass spectrometry (FastGC-PICI-TOFMS) was developed for the analysis of EC in Chinese liquor. The rapid separation of EC in Chinese liquor was conducted using FastGC based on a thermostatic column set at 150 °C to eliminate the interferences of matrix effects. The PICI-TOFMS could realize accurate quantification of EC without any sample pre-treatment due to the efficient ionization of EC by the PICI source. As a result, the total analysis time for EC in Chinese liquor was less than 4 min. The limit of detection (LOD) for EC was 4.4 μg L−1. And the intra-day and inter-day precision were 3.2%–3.7 % and 1.6 %, respectively. Finally, the ability of the proposed method was preliminarily proved by high-throughput and accurate measurement of EC in four different flavors of Chinese liquors.

Simultaneous determination of florfenicol, its metabolite and three fluoroquinolone residues in poultry muscles via high-performance liquid chromatography with fluorescence detection

A high-performance liquid chromatography with fluorescence detection (HPLC–FLD) procedure was developed and refined to detect florfenicol (FF), its metabolite florfenicol amine (FFA), and three fluoroquinolone (FQ) residues present in poultry muscles concurrently. Sample pretreatment was conducted through liquid–liquid extraction (LLE). Ethylenediaminetetraacetic acid (EDTA), 90 % acetonitrile and acetonitrile were used as extraction solvents. The stationary phase employed was an XBridge BEH C18 column (5 μm, 4.6 mm × 150 mm), and the mobile phase consisted of a 0.01 mol/L solution of sodium dihydrogen phosphate and acetonitrile (65:35, V/V); the former contained 0.005 mol/L sodium dodecyl sulfate and 0.1 % triethylamine (pH adjusted to 4.8 with phosphoric acid). The limits of detection (LODs) and limits of quantification (LOQs) were 0.03–1.5 µg/kg and 0.1–5.0 μg/kg, respectively. The linear relationships were satisfactory within the corresponding concentration ranges, with determination coefficients (R2) ≥ 0.9997. The mean recoveries ranged from 70.34 to 94.98 %, and the intraday relative standard deviations (RSDs) and interday RSDs were 1.46–6.24 % and 1.60–7.31 %, respectively. This verified method is applicable and reliable for analyzing actual samples.

Surface amphiphilic hybrid porous polymers based on cage-like organosiloxanes for pipette tip solid-phase extraction of microcystins in water

The occurrence of microcystins (MCs) during harmful algal blooms (HABs) represents a major threat to freshwater environments. In this work, a novel surface amphiphilic hybrid porous polymers based on cage-like organosiloxanes (PCSs) was prepared for the enrichment of MCs. The copolymerization of bifunctional amphiphilic monomers, 2-methacryloyloxyethyl phosphorylcholine (MPC) and N-benzylquininium chloride (BQN), with the cross-linker methacryl substituted polyhedral oligomeric silsesquioxane (POSS) was achieved in an ionic liquid-based porogenic medium. The hierarchical porous structure, a variety of surface functional groups and weak hydrophilicity were well characterized on the prepared materials using scanning electron microscopy, nitrogen adsorption/desorption analysis, Fourier transform infrared spectroscopy, X-ray photoelectron spectroscopy, zeta potential analysis and water contact angle testing, respectively. The as-prepared surface amphiphilic PCSs was used as an adsorbent for pipette tip solid-phase extraction (PT-SPE) to enrich microcystins (MCs) from surface waters before their analysis by capillary electrochromatography (CEC) and liquid chromatography-mass spectrometry (LC-MS). Under the optimal conditions, the established PT-SPE-LC-MS method exhibited a wide linear range (10–10,000 ng L−1), low limits of detection (4.0–8.0 ng L−1) and satisfactory recoveries (89.5–102.8 %) for MCs. An adsorption mechanism involving electrostatic interactions, hydrogen bonding, hydrophilic interactions, and π-π stacking has been proposed. The findings suggest that the use of surface amphiphilic PCSs materials as adsorbents in the PT-SPE platform facilitates efficient enrichment of MCs for subsequent chromatographic analysis. These investigations offer a new perspective on the simple and uncomplicated pretreatment of complex environmental samples.

Analysis of rebaudioside A in probiotic-fermented milk via quantitative nuclear magnetic resonance spectroscopy

Rebaudioside A is used in the food and beverage industries owing to its sweetness, even in extremely small amounts, as well as its natural origin. Liquid chromatography is commonly used to quantify rebaudioside A; however, it suffers from complicated sample pretreatment and time consumption. External-calibration quantitative nuclear magnetic resonance (qNMR) analysis of probiotic-fermented milk has thus far not been reported. Moreover, it involves more factors contributing to errors than internal standard approaches used in the official methods, such as those recommended by the International Organization for Standardization. In this study, we developed a method for quantifying rebaudioside A in probiotic-fermented milk using internal-calibration qNMR. This method entails a simple pretreatment step with dilution and solid-phase extraction, enabling rebaudioside A determination in five types of beverages including probiotic-fermented milk. The results revealed a recovery rate and relative standard deviation of 85.4–104 % and 1.57–4.09 %, respectively, satisfying the international guidelines. The working range of qNMR was 0.80–26.7 mg/100 g of product, and the analysis time was 10–20 min. Therefore, the proposed internal-calibration qNMR method facilitates easy, rapid, and accurate quantification of rebaudioside A in probiotic-fermented milk and is applicable for analyzing various other beverages and foods.

A super convenient and specific CRISPR/Cas12a diagnostic platform for toxigenic Burkholderia gladioli based on virulence genes

In recent years, food poisoning cases caused by toxigenic Burkholderia gladioli pv. cocovenenans (BGC) have been frequently reported in Asian countries like China and Indonesia, which seriously threaten food safety and public health. BGC is the only subspecies within the Burkholderia gladioli species that can cause food poisoning in humans, but there is a high degree of sequence homology between the different subspecies, making detection and differentiation difficult. In this study, a specific DNA target was screened out from the bongkrekic acid biosynthentic gene cluster. A rapid, sensitive, and highly specific CRISPR/Cas12a molecular diagnostic platform was developed for the first time to differentiate between toxigenic and non-toxigenic subspecies. Genomic DNA was extracted from the samples after a simple boiling process. The sensitivity was evaluated from three different levels, the genomic DNA sensitivity was determined to be 10−4 ng/μL, the target DNA sensitivity was determined to be 14.9 copies/μL, and the bacterial suspension sensitivity was confirmed to be 1.52 CFU/mL. The detection limit of BGC in artificially contaminated coconut water samples was determined to be 1.50 CFU/mL without any microbial enrichment. Including sample pretreatment and DNA extraction process, the whole detection of BGC in samples can be completed within 60 min under the assistance of some small and portable devices. This super convenient diagnostic platform is suitable for food field testing and provides a theoretical basis for the specific identification of BGC.

ACFs-NH2 developed for dispersive solid phase extraction combined with Py-GC/MS for nanoplastic analysis in ambient water samples

Accurate determination of nanoplastics (NPs) in aquatic ecosystems constitutes a challenge for which highly sensitive analytical methods are necessitated. Herein, a sample pretreatment based on self-made amino-functionalized activated carbon fibers (ACFs-NH2) dispersive solid-phase extraction (DSPE) allows for high-recovery, followed by high-sensitivity detection of NPs by pyrolysis-gas chromatography/mass spectrometry (Py-GC/MS). The developed methodology allowed low detection limits (20–100 μg/L) to be achieved quickly in a few steps. Under optimal conditions, ACFs-NH2 (12.5 mg) was able to recover ≥98.45 % of polystyrene (PS) nanoplastics at high concentration (100 mg/L) in 10 mL seawater. Based on the high adsorption performance of materials, the adsorption dynamics and isotherms were determined to infer the interaction mechanism of PSNPs on ACFs-NH2. After adsorption, the target on the surface of the adsorbent can be directly pyrolyzed, which can simplify the operation steps and avoid the elution of organic solvents, making the process more environmentally friendly. This strategy is feasible for the analysis of trace NPs in water systems.

Boronic acid ester-based hydrogel as surface-enhanced Raman scattering substrates for separation, enrichment, hydrolysis and detection of hydrogen peroxide residue in dairy product all-in-one

Rapid and selective separation, enrichment and detection of trace residue all-in-one in complex samples is a major challenge. Hydrogels with molecular sieve properties can selectively separate and enrich target analytes, and the combination with high sensitivity detection of surface-enhanced Raman scattering (SERS) is expected to achieve the above all-in-one detection. Herein, the core-shell structured Au@poly(N-isopropylacrylamide)-phenylboronic acid hydrogel (Au@PNIP-VBA) with boronic acid ester groups was prepared by thermally initiated polymerization. The boronic acid ester groups in hydrogel are selectively hydrolyzed by hydrogen peroxide (H2O2) to hydroxyl structures, leading to a reduction in SERS signals. The Au@PNIP-VBA hydrogel has molecular sieve properties and high SERS activity, making it suitable for separation, enrichment, hydrolysis and detection of H2O2 all-in-one. A rapid SERS method was developed for analysis of H2O2 based on the Au@PNIP-VBA hydrogel with the linear range of 8.5 × 10−2-6.8 mg L−1 and the detection limit of 33 μg L−1. The method was successfully applied to the determination of H2O2 residue in fresh milk, pure milk, yogurt and camel milk, with the recoveries were in the range of 82.2%–109.3% and the relative standard deviations were 2.8%–8.3%. This efficient all-in-one strategy has the advantages of simple sample pre-treatment, rapid analysis (30 min) and high sensitivity, making it highly promising for food quality and safety analysis.

Development of an affordable light emitting diode spectrophotometer paired with a Python program for calibration and linearity testing and the measurement of uranium(VI).

Uranium (U) is a radiologically and chemically toxic element that occurs naturally in water, soil, and rock at generally low levels. However, anthropogenic uranium can also leach into groundwater sources due to mining, ore refining, and improper nuclear waste management. Over the last few decades, various methods for measuring uranium have emerged; however, most of these techniques require skilled scientists to run samples on expensive instrumentation for detection or require the pretreatment of samples in complex procedures. In this work, a Schiff base ligand (P1) is used to develop a simple spectrophotometric method for measuring the concentration of uranium (VI) with an accurate and affordable light-emitting diode (LED) spectrophotometer. A test for a higher-order polynomial relationship was used to objectively determine the calibration data's linearity. This test was done with a Python program on a Raspberry Pi computer that captured the spectrophotometer's calibration and sample measurement data.

Determination of bioequivalence between generic and reference drugs using laser-induced breakdown spectroscopy

BackgroundIn vitro bioequivalence studies are strictly limited to the comparison of dissolution performance to a reference drug. These studies are performed without considering the chemical similarity between the generic and reference drug formulations. This work has focused on developing a groundbreaking method based on the laser-induced breakdown spectroscopy (LIBS) technique for the in vitro bioequivalence determination of immediate-release solid oral dosage form generic drugs and as an alternative method for establishing the biowaiver of in vivo generic drug studies. ResultsThe novel LIBS-based methodology to determine in vitro bioequivalence is fast, easy to perform, and can be carried out without the requirement of tedious and complicated sample pre-treatment, nor expensive instrumentals and reagents, almost directly on the drug samples. Furthermore, the proposed methodology demonstrated that it is enough to identify the spectrochemical similarity of the formulation between generic drugs to a reference drug through the chemometric study of their LIBS spectra, based on the determination of the differentiation and similarity factors, f1 and f2, respectively, used in the pharmaceutical industry in this purpose. After analysing their LIBS spectra, the generic drugs selected for this work have all been shown to be in vitro bioequivalent, given their f1 values of less than 15 and f2 values greater than 50, according to the technical regulations on which the American and European medicines agencies are based for the approval of registration for generic immediate-release solid oral dosage form drugs. This has been evidenced even for drugs from Class III and Class IV of the biopharmaceutical classification system, whose active principle nominal concentration is very low as 0.1 and 0.25 mg/tablet, respectively. Significancefor the first time the LIBS technique has been successfully used in an advanced application for the pharmaceutical industry. The proposed method constitutes a reliable and specialized methodology for the establishment of formulation similarity between two drugs, without the requirement of separate identification of each of their components, which is a new and potential tool to determine the in vitro bioequivalence for generic immediate-release solid oral dosage form drugs.

Cell Membrane-Camouflaged Biomimetic Magnetic Fluorescent Nanoparticles with Enhanced Stability for High-Performance Lead Drug Discovery.

Biomimetic nanoengineering empowers nanoparticles with enhanced biointerfacial capabilities by directly utilizing cell membranes (CMs) of natural origin. This top-down technique provides a powerful tool for the screening of potentially active compounds in complex matrices. Herein, cartilaginous end plate (CEP) cell membrane biomimetic Nile red (NR)-loaded zeolitic imidazolate frameworks-8 (ZIF-8) modified magnetic graphene oxide (CEP/MGO-ZIF-8-NR) nanocomposites with enhanced stability were accurately prepared by chemical bonding and used as a drug discovery platform for the specific identification and effective extraction of drug leads with anti-intervertebral disc degeneration (IDD) in Yaobitong capsules (YBTCs). The constructed CEP/MGO-ZIF-8-NR exhibited excellent magnetic properties, fluorescence properties, and stability. In addition, drug binding experiments showed that the CEP/MGO-ZIF-8-NR nanocomposites possessed higher adsorption capacity, faster adsorption rate, and superior selectivity compared with uncoated MGO-ZIF-8-NR. Ultimately, four potential bioactive molecules, including ginsenoside Ro, ginsenoside Rg1, astringin, and chikusetsusaponin V methyl ester, were successfully screened and identified in vitro from YBTC. The results of the CCK-8 assay and BrdU ELISA kit showed that the screened compounds promoted CEP cell proliferation in a concentration-dependent manner. Cellular distribution experiments revealed that CEP/MGO-ZIF-8-NR could rapidly escape from lysosomes and into the cytoplasm. And the pharmacological activity of these compounds was further confirmed by real-time cytomorphological imaging of CEP cells by confocal laser scanning microscopy (CLSM). Overall, this surface engineering strategy endows bioaffinity sample pretreatment materials with tremendous versatility, improves drug screening efficiency, and broadens the horizons and methodologies for drug lead discovery.

An Exploration of the Viral Coverage of Mosquito Viromes Using Meta-Viromic Sequencing: A Systematic Review and Meta-Analysis.

The aim of this review was to delve into the extent of mosquito virome coverage (proportion of viral reads) via meta-viromic sequencing and uncover potential factors of heterogeneity that could impact this coverage. Data sources were PubMed, Web of Science, Embase, Scopus, Science-Direct, Google Scholar, and the China National Knowledge Infrastructure. Pooled coverage was estimated using random-effects modeling, and subgroup analyses further reveal potential heterogeneous factors. Within the three mosquito genera studied, Culex exhibited the highest pooled viral coverage of mosquito viromes at 7.09% (95% CI: 3.44-11.91%), followed by Anopheles at 5.28% (95% CI: 0.45-14.93%), and Aedes at 2.11% (95% CI: 0.58-7.66%). Subgroup analyses showed that multiple processing methods significantly affected the viral coverage of mosquito viromes, especially pre-treatment of mosquito samples with saline buffer/medium and antibiotics prior to DNase/RNase treatment and removal of the host genome prior to RNA library construction. In conclusion, the results of this study demonstrate that the viral coverage of mosquito viromes varies between mosquito genera and that pre-treatment of mosquito samples with saline buffer/medium and antibiotics before DNase/RNase treatment and removing host genomes prior to RNA library construction are critical for the detection of RNA viruses in mosquito vectors using meta-viromic sequencing.

Nanozyme-Based Microfluidic Chip System for pH-Regulated Pretreatment and Sensitive Sensing.

Nanozymes, possessing nanomaterial properties and catalytic activities, offer great opportunities to design sensitive analytical detection systems. However, the low interference resistance of nanozymes poses a significant limitation on the precise detection of target substances. Herein, a nanozyme-based microfluidic chip system for pH-regulated pretreatment and sensitive sensing of cysteine (Cys) is reported. The copper metal-organic framework (Cu MOF) exhibits good cysteine oxidase-like activity at pH 7.0, while demonstrating excellent laccase-like activity at pH 8.0. Taking advantage of the pH-regulated enzyme-like activity, the integrated microfluidic device involving the immobilization of Cu MOF eliminates the interference of dopamine (DA) and accurately detects the target Cys. Compared with the untreated reaction system, the developed nanozyme system shows a significantly improved accuracy in detecting Cys, with an R2 value of 0.9914. This work provides an efficient method to enhance the interference resistance of nanozymes and broadens the application in sample pretreatment.

Enhanced Detection of Curcumol in Biological Samples via Acetonitrile‐Anhydrous Sodium Sulfate Salt‐Assisted Liquid‐Liquid Microextraction Coupled With High‐Performance Liquid Chromatography

ABSTRACTCurcumol, a significant bioactive molecule, requires accurate detection methods due to its extensive pharmacological properties. This study develops high‐performance liquid chromatography (HPLC) methods to detect curcumol and constructs sample pretreatment techniques using salt‐assisted liquid‐liquid microextraction (SALLME). The optimization process focuses on conditions for acetonitrile (ACN) anhydrous sodium sulfate‐assisted liquid‐liquid microextraction. By adding excess anhydrous sodium sulfate to an ACN aqueous solution, ACN precipitates, forming a saturated saline solution and resulting in a two‐phase system. Curcumol, being highly soluble in ACN, separates into and enriches within the ACN phase, achieving effective matrix separation. SALLME demonstrated an enrichment factor of up to 12.4 for curcumol. The extracted ACN is then injected into an HPLC instrument for quantitative detection and analysis. The method exhibited a good linear range (0.5–10.0 µg/mL) and a low detection limit (0.05 µg/mL). The results indicate that curcumol can be effectively separated from complex matrices using this technique. By optimizing salt concentration and temperature, the HPLC method for determining curcumol in biological samples was successfully established, achieving recoveries between 86.2% and 137.6% in spiked urine and blood samples. Furthermore, this method offers rapid, efficient, and environmentally friendly sample preparation, promising significant applications in pharmacological and clinical settings.

Fluorescence of Intrinsic Milk Chromophores as a Novel Verification Method of UV-C Treatment of Milk.

The European Food Safety Authority (EFSA) has approved the use of a 1045 J/L UV-C dose as an adjunct to pasteurization to increase the shelf life and vitamin D3 content of milk. However, there are no verification methods analogous to the alkaline phosphatase test for pasteurized milk to ensure that the desired UV-C dose has been correctly applied. The aim is to develop a real-time in-line detector based on fluorescence spectroscopy. In this study, 22 different UV-C doses (ranging from 0 to 2000 J/L) were applied to milk to assess the impact of photooxidation on intrinsic photosensitive chromophores. Fluorescence spectroscopy (90°-angle) was employed as the method of analysis for monitoring the changes in the fluorescence spectra of chromophores in milk without sample pretreatment. Three important chromophore areas (CAs) were identified: CA 1 (riboflavin), CA 3 (vitamin A and dityrosine) and CA 4 (tryptophan), with statistically significant changes at around 1045 J/L and 1500 J/L. The findings of our preliminary study support our hypothesis that the fluorescence of intrinsic chromophores can be used as verification of the applied UV-C dose.