Quantitative Determination Research Articles

A-083 Performance evaluation of Sentinel Diagnostics’ ACE assay on the Abbott Alinity c system and ARCHITECT c16000 System

Abstract Background The analytical performances of the Angiotensin Converting Enzyme (ACE) assay for quantitative determination of ACE activity in human serum and plasma have been evaluated on the Abbott Alinity c system and ARCHITECT c16000 System. ACE (dipeptidyl carboxypeptidase) is a glycoprotein peptidyldipeptide hydrolase that cleaves histidylleucine dipeptide from angiotensin I, a relatively inactive decapeptide. The latter is converted to the potent vasoconstrictor, angiotensin II. ACE also inactivates bradykinin. Elevated levels of ACE activity occur in serum of patients with active sarcoidosis, and occasionally in premature infants with respiratory distress syndrome, in adults with tuberculosis, Gaucher’s disease, leprosy, and in many other pathologic conditions involving lung and liver diseases. Methods ACE hydrolyses urylacryloylphenylalanine-glycylglycine (FAPGG) to furylacryloylphenylalanine (FAP) and glycylglycine. Hydrolysis of FAPGG results in a decrease in absorbance at 340 nm. The ACE activity in the sample is determined by comparing the sample reaction rate to that obtained with the Sentinel ACE calibrator. Performances evaluation have been done following the latest CLSI guidelines protocols available. Results Summary of the analytical performances for the tested assay is reported in table below. Conclusions Analytical performances of Sentinel Diagnostics’ ACE assay demonstrate that quantitative determination of ACE activity in human serum and plasma are suitable for the routine measurement of the analyte on Abbott Alinity c system and ARCHITECT c16000 System.

B-357 Siemens CA 72-4: an assay for measurement of Tumor Associated Glycoprotein (TAG) 72

Abstract Background The tumor associated glycoprotein (TAG) 72, also known as CA 72-4 is a mucin protein of high molecular weight. CA 72-4 is found on the surface of many cancer cells, including stomach (gastric), ovary, breast, colon and pancreatic cells. CA 72-4 is currently one of the primary markers for the diagnosis of gastric cancer and monitoring the course and efficacy of gastric cancer. In ovarian cancer, it has a higher sensitivity for mucin- type ovarian cancer, and its combination with CA125 for detection can significantly improve the clinical sensitivity. In colorectal cancer, CA 72-4 combined with CEA detection was also found to significantly improve the sensitivity of the initial diagnosis. An assay for CA 72-4 on Siemens Atellica IM (Siemens CA 72-4) is being developed and analytical performance is being presented. Methods The Siemens CA 72-4 assay (in development) is a one-step chemiluminescent microparticle immunoassay (CMIA) for the quantitative determination of CA 72-4 in human serum and plasma. The specimen is incubated with paramagnetic microparticles coated with the monoclonal antibody (mAb) B72.3 and acridinium-labeled mAb CC49 conjugate on the Siemens Atellica IM® System. After incubation and washing, Acid and Base reagents are added. The resulting relative light units (RLUs) are directly proportional to the amount of CA 72-4 in the sample allowing for the quantitative determination of CA 72-4 in the specimen. Results The following performance were determined using 3 unique lots of reagents and calibrators with the calibration range for the assay being 0.00 to 200.00 U/mL. The limit of blank (LoB), limit of detection (LoD), and limit of quantitation at 20% CV (LoQ) were determined and met the goal of ≤ 1.00 U/mL. Linearity according to CLSI EP06 Ed2 was demonstrated for a range of 1.00 through 200.00 U/mL. A Precision study of 2 controls and 5 panels spanning the range of the assay demonstrated a total %CV ≤ 6% for samples ≥ 5.00 U/mL and ≤ 0.25 SD for samples <5.00 U/mL. In the sample tube type study, a matrix comparison of 23 matched serum and plasma samples were evaluated. Passing-Bablock slopes of < 8% and r > 0.99 were observed when evaluating samples within the measurement range for plasma (LiHep, LiHep separator, K2EDTA and K3EDTA) tubes and serum separator tubes (SST) compared to the Red Top serum samples. Five endogenous substances were evaluated for interference in the CA 72-4 assay. The average percent difference between test and control samples for all endogenous interferents was < 10%. Percent difference in the presence of 28 potentially interfering drugs was ≤ 4.0%. Conclusions The Siemens CA 72-4 assay currently in development is a sensitive and precise assay for the quantitative determination of CA 72-4 in human serum and plasma.

Advanced Electrochemical Monitoring of Carbendazim Fungicide in Foods Using Interfacial Superassembly of NRPC/NiMn Frameworks.

A simple, sensitive and reliable sensing system based on nitrogen-rich porous carbon (NRPC) and transition metals, NRPC/Ni, NRPC/Mn and NRPC/NiMn was developed and successfully applied as electrode materials for the quantitative determination of carbendazim (CBZ). The synergistic effect of NRPC and bimetals with acceptable pore structure together with flower-like morphology resulted in producing a highly conductive and interconnected network in NRPC/NiMn@GCE, which significantly enhanced the detection performance of CBZ. The electrochemical behavior investigated by cyclic voltammetry (CV) showed improved CBZ detection for NRPC/NiMn, due to the controlled adsorption/diffusion process of CBZ by the NRPC/NiMn@GCE electrode. The influences of various factors such as pH, NRPC/NiMn concentration, CBZ concentration and scan rate were studied. Under optimal conditions, 0.1 M phosphate-buffered saline (PBS) with a pH of 7.0 containing 30 µg/mL NRPC/NiMn, a favourable linear range detection of CBZ from 5 to 50 µM was obtained. Moreover, a chronoamperometric analysis showed excellent repeatability, reproducibility and anti-interfering ability of the fabricated NRPC/NiMn@GCE sensor. Furthermore, the sensor showed satisfactory results for CBZ detection in real samples with acceptable recoveries of 96.40-104.98% and low RSD values of 0.25-3.45%.

B-356 Siemens CYFRA 21-1: an assay for measurement of cytokeratin fragment 21-1

Abstract Background Cytokeratins are structural proteins forming the subunits of epithelial intermediary filaments. The soluble fragment of cytokeratin 19, cytokeratin fragment 21-1 (CYFRA 21-1), can be found in the blood of lung cancer patients, primarily those with non-small cell lung cancer (NSCLC). CYFRA 21-1 is the preferred biomarker in the management of NSCLC, particularly squamous cell and large cell carcinoma subtypes. An assay for CYFRA 21-1 on Siemens ADVIA Centaur® XPT and Siemens Atellica® IM (Siemens CYFRA) is being developed and analytical performance is being presented. Methods The Siemens CYFRA 21-1 assay (in development) is a one-step chemiluminescent microparticle immunoassay (CMIA) for the quantitative determination of CYFRA 21-1 in human serum and plasma. The specimen is incubated with paramagnetic microparticles coated with the monoclonal antibody (mAb) BM19.21 and acridinium-labeled mAb KS19.1 conjugate on the Siemens ADVIA Centaur XPT System and Siemens Atellica IM System. After incubation and washing, Acid and Base reagents are added. The resulting relative light units (RLUs) are directly proportional to the amount of CYFRA 21-1 in the sample allowing for the quantitative determination of CYFRA 21-1 in the specimen. Results The following performance were determined using 3 unique lots of reagents and calibrators with the calibration range for the assay being 0.00 to 100.00 ng/mL. For the ADVIA Centaur XPT and Atellica IM systems were shown to be equivalent within 4%. The limit of blank (LoB), limit of detection (LoD), and limit of quantitation at 20% CV (LoQ) were determined and met the goal of ≤ 1.00 ng/mL. Linearity according to CLSI EP06 Ed2 was demonstrated for a range of 0.66 through 100.00 ng/mL. A Precision study of 2 controls and 5 panels spanning the range of the assay demonstrated a total %CV ≤ 8% at all levels. In the sample tube type study, a matrix comparison of 61 matched serum and plasma samples were evaluated. Passing-Bablock slopes of < 5% and r > 0.99 were observed when evaluating samples within the measurement range for plasma (LiHep, LiHep separator, K2EDTA and K3EDTA) tubes and serum separator tubes (SST) compared to the Red Top serum samples. Six endogenous substances were evaluated for interference in the CYFRA 21-1 assay. The average percent difference between test and control samples for all endogenous interferents was < 10%. Percent difference in the presence of 20 potentially interfering drugs was ≤ 4.0%. Conclusions The Siemens CYFRA 21-1 assay currently in development is a sensitive and precise assay for the quantitative determination of CYFRA 21-1 in human serum and plasma.

B-358 Siemens NSE: an assay for measurement of human Neuron-Specific Enolase

Abstract Background Neuron-specific enolase (NSE), a glycolytic enzyme (2-phospho-D-Glycerate hydrolase) can be found in a variety of non-neuroendocrine cells. Determination of NSE is used for monitoring response to therapy and detection of recurrent disease, such as SCLC and neuroendocrine tumors. An assay for NSE on Siemens ADVIA Centaur® XPT System and Siemens Atellica® IM system (Siemens NSE) is being developed and analytical performance is being presented. Methods The Siemens NSE assay (in development) is a one-step chemiluminescent microparticle immunoassay (CMIA) for the quantitative determination of NSE in human serum. The specimen is incubated with paramagnetic microparticles coated with the monoclonal antibody (mAb) NSE21, and acridinium-labeled mAb NSE17 conjugate on the Siemens ADVIA Centaur XPT system and Siemens Atellica IM system. After incubation and washing, Acid and Base reagents are added. The resulting relative light units (RLUs) are directly proportional to the amount of NSE in the sample allowing for the quantitative determination of NSE in serum. Results The following performance were determined using 3 unique lots of reagents and calibrators with the calibration range for the assay being 0.0 to 400.0 ng/mL. For the ADVIA Centaur XPT and Atellica IM systems were shown to be equivalent within 2%. The limit of blank (LoB), limit of detection (LoD), and limit of quantitation at 20% CV (LoQ) were determined and met the goal of ≤ 1.6 ng/mL. Linearity according to CLSI EP06 Ed2 was demonstrated for a range of 1.6 through 400.0 ng/mL. A Precision study of 2 controls and 6 panels spanning the range of the assay demonstrated a total %CV ≤ 7% at all levels. Comparison of 50 specimens collected in Red Top serum and Serum Separator Tubes (SST) had a Passing-Bablock slope of < 2% and r of > 0.99 when evaluating samples within the measurement range. Five endogenous substances were evaluated for interference in the Siemens NSE assay. The average percent difference between test and control samples for all endogenous interferents was < 10%. Percent difference in the presence of 38 potentially interfering drugs was ≤ 10.0%. The potential cross-reactant αα enolase was evaluated at concentration of 900 ng/mL and the percent cross-reactivity was ≤ 2%. Conclusions The Siemens NSE assay currently in development is a sensitive and precise assay for the quantitative determination of NSE in human serum.

A-033 Comparison of the Roche hCG STAT and hCG+β Assays

Abstract Background Roche Diagnostics offers hCG STAT and Elecsys hCG+β for the quantitative determination of human chorionic gonadotropin (hCG) in serum. While hCG STAT detects intact hCG after a 9-minute incubation, hCG +β; detects intact hCG, nicked hCG forms, the β-core fragment and the free β-subunit after 18 minutes. The manufacturer reported a strong correlation (R = 0.999) between the two assays using 81 samples with hCG concentrations ranging from 6 - 7675 mIU/mL. However, hCG concentrations during early pregnancy are low. This study aims to determine the correlation between the two assays with samples of < 50 mIU/mL hCG concentration. Additionally, we aim to determine the qualitative concordance between the two assays assuming the upper 95% confidence limit reported by the manufacturer as the positive cutoff value. Methods We compared hCG results from a cohort of remnant serum samples (N = 65; hCG 2 - 49 mIU/mL) previously analyzed using the Elecsys hCG + β assay on Roche cobas e 801 analyzers against the Elecsys HCG STAT assay on the cobas e 601 analyzers. A quantitative method comparison was also performed on a subset (N =33) of samples with hCG < 5 mIU/mL. Regression analysis was conducted using EP Evaluator software (version 12). A qualitative concordance between the two assays was performed assuming the upper 95% confidence limit in healthy, non-pregnant premenopausal women as the positive cutoff value (5.3 mIU/mL and 4.9 mIU/mL for the hCG+β and hCG STAT assay, respectively). Results Quantitative comparison using a wider range revealed an R value of 0.968, however, at hCG < 5mIU/ml concentration, the R value was 0.755. Qualitative comparison using each assay’s cutoff value identified seven discrepancies. Three samples that would be classified as positive with the hCG+β assay would be considered negative with hCG STAT assay, and four samples that would be considered negative with the hCG+β assay would be classified as positive with the hCG STAT assay. When a 4.9 mIU/mL cutoff value is used for both assays, eight samples that would be positive with the hCG+β assay would be considered negative with the hCG STAT assay, and one sample goes the other way. Conversely, if 5.3 mIU/mL is used as a cutoff value for both assays, three samples that would be classified as positive with the hCG+β assay would be considered negative with the hCG STAT assay, and one sample tested the other way. Conclusions The correlation between the two assays is strongest when comparison is made using a wide range of hCG values, consistent with the manufacturer's claims. However, the correlation weakens when comparing samples with low hCG values. Qualitative comparison of the assays using their respective cutoff values resulted in 89.2% overall agreement, 84.6% when a lower cutoff value of 4.9mIU/ml is used and 93.8% overall agreement when 5.3mIU/ml cutoff value used. We concluded that while the assays are comparable, minor differences were observed at low hCG values. Further study using a larger sample size and clinical correlation is needed to reinforce the conclusions.

B-044 Development and Evaluation of an Assay for the Detection of Human Total IgE in Serum on the Fully Automated Chemiluminescence Immunoassay Platform - Autolumo® 2000 Plus Analyzer

Abstract Background The escalating prevalence of allergic diseases on a global scale, affecting approximately 20% of the world’s population, has become a significant health concern. Clinical determination of total IgE in serum and plasma is used as an aid to the diagnosis of allergic diseases. The tIgE CLIA Microparticles Kit is a chemiluminescent microparticle immunoassay for the quantitative determination of total IgE concentrations in human serum. This kit, integrated with the AutoLumo® 2000 Plus Analyzer, offers an automated, cutting-edge platform for total IgE detection. This streamlined combination provides clinicians with a reliable tool for comprehensive and efficient allergic diseases diagnosis. Methods The performance of tIgE CLIA Microparticles Kit was evaluated on AutoLumo® A2000 Plus automated chemiluminescence analyzer and compared to a well-characterized platform (Elecsys IgE II on Cobas e601, Roche). This assay is based upon the two-step sandwich method. During detection, the patient sample and the anti-IgE coated microparticles solution were added sequentially. Unbound components were washed off after incubation. The formation of a complex occurred with the subsequent addition of a HRP conjugated anti-IgE antibody solution. The catalysis of chemiluminescent substrates by this complex generated a measurable light signal recorded as RLUs (Relative Light Units). A 2-point calibrator was used to analyze the tIgE antibody concentrations in the serum sample. The limit of blank (LoB), limit of detection (LoD), limit of quantitation (LoQ), accuracy, repeatability, linearity, specificity, on-board stability and real-time stability of the assay kit were evaluated. Serum samples (n=220) that had been previously well characterized were used for method comparisons. Results The tIgE CLIA Microparticles assay demonstrated a measurement range of 5-2500 IU/mL, with LoB, LoD, and LoQ values of ≤3 IU/mL, ≤4 IU/mL, and ≤5 IU/mL, respectively. The accuracy of the assay kit was assessed by trueness and precision following CLSI EP15-A2 and EP5-A2 guidelines. The trueness was evaluated using WHO IRP 11/234 as the reference standard, the estimate %bias between the expected and measured mean value was ≤5%. The average %CV of the intra-run and inter-run precision were 5.00% and 3.24% respectively, with a linearity of R≥0.990. The specificity to tIgE is evident, with no cross-reactivity to IgA, IgG, and IgM (≤100 μg/mL), and minimal interference from rheumatoid factor (RF) and anti-nuclear antibodies (ANA) (<10%). Furthermore, based on the CLSI EP7-A2 guideline, it showed no endogenous interference with bilirubin (30 mg/dL), hemoglobin (250 mg/dL), and triglycerides (3000 mg/dL) and no exogenous interferences with 14 commonly used allergy treatment drugs except Omalizumab (360μg/mL). Notably, the assay remains stable on-board for 28 days and exhibits real-time stability for up to 12 months at 2-8ᵒC. Clinical evaluation, involving 220 samples covering the linearity range of 5.0-2500 IU/mL, reveals a strong correlation (R>0.990) and the bias was within 15% when compared to a reference kit according to the CLSI EP9-A2 guideline. Conclusions Autolumo® Analyzers, in conjunction with the tIgE CLIA Microparticles Assay, presents a fully automated chemiluminescence immunoassay platform for accurate and cost-effective screening to aid allergic diseases diagnostics with excellent specificity, precision, and on-board stability.

Quantitative determination by UHPLC-MS/MS of 18 common drugs of abuse and metabolites, including THC and OH-THC, in volumetric dried blood spots: a sustainable method with minimally invasive sampling

The increased use of drugs of abuse urges forensic toxicologists to create quick, simple, minimally invasive sampling techniques for biological fluids combined with analytical methods assuring accurate results. To this purpose, a method was developed aimed at quantifying 18 drugs of abuse and metabolites in DBS. Validation of the method was conducted by spiking blank whole blood with the analytes on Capitainer® B cards. The extracts were analyzed by a targeted UHPLC-MS/MS method. Linear calibration was achieved in the range of 1–100 ng/mL for: amphetamine, MDA, MDMA, methamphetamine, cocaine, codeine, benzoylecgonine, cocaethylene, morphine, 6-MAM, buprenorphine, methadone, EDDP, ketamine, norbuprenorphine norketamine, THC, and OH-THC. Experimental LOD was found at 0.5 ng/mL for all analytes except for norbuprenorphine, THC and THC-OH which yielded a LOD of 1 ng/mL. Intra- and inter-day accuracy was satisfactory with bias% resulting within 5%. Evaluation of intra- and inter-day precision yielded CV% values within 20%, for all compounds except EDDP. Average extraction recovery calculated at low (2 ng/mL) and high (75 ng/mL) concentration levels was 63% while average matrix effect determined at the same levels was found to be within 85% − 115% for all analytes except from codeine (70%) and MDMA (131%). The method was applied to authentic blood samples spotted onto the DBS card and the minimum value detected was 1.3 ng/mL. HPLC-MS/MS proved capable to identify all the targeted analytes at low concentrations in the small blood volumes obtainable from DBS cards, which in turn confirmed to be effective and sustainable micro-sampling devices.

Quantitative assessment of corneal biomechanical changes in vivo after photorefractive intrastromal corneal cross-linking using optical coherence elastography.

Photorefractive intrastromal corneal cross-linking (PiXL) treatment corrects myopia by enhancing localized central corneal biomechanics. However, the dose-effect relationship between the changes in corneal biomechanics and alterations in corneal curvature resulting from this treatment remain unclear. We therefore developed an acoustic radiation force optical coherence elastography (ARF-OCE) technique to investigate the dose-effect relationship in PiXL. ARF-OCE measurements and corneal topography were performed 3 days before and 1 week after PiXL treatment. Depth-resolved Young's modulus images of the in vivo corneas were obtained based on the phase velocity of the Lamb wave. PiXL treatments with five ultraviolet-A (UVA) energy doses (5.4, 15, 25, 35, and 45 J/cm2) were administered to rabbit corneas in vivo (n=15). The percentage change in Young's modulus (ΔE%) of the cornea increased from 0.26 to 1.71 as the UVA energy dose increased from group I (5.4 J/cm2) to group V (45 J/cm2). Meanwhile, the change in the mean keratometry (ΔKm ) of the cornea increased from 0.40 to 2.10 diopters (D) as the UVA energy dose increased from group I to group IV (35 J/cm2). Furthermore, a statistically significant positive correlation was observed between ΔE% and ΔKm in groups I to IV. With increasing UVA energy dose, the corneal Young's modulus significantly increased. Given the observed correlation, ΔE% holds promise as a new quantitative biomechanical parameter for determining the dose-effect relationship in PiXL treatment. It should be emphasized that there may be an inflection point of ΔE%, at which corneal keratometry ceases to flatten and begins to increase. The ARF-OCE system has demonstrated its efficacy in quantitatively assessing changes in corneal biomechanics in vivo following PiXL treatment. This technique has great potential in facilitating the quantitative determination of the dose-effect relationship in PiXL treatment.

An integrated three-signal biosensor based on phage multifunctional probe for simultaneous and ultrasensitive detection of live /dead Listeria monocytogenes

The simultaneous and sensitive detection of live/dead bacteria in food samples is crucial for the control and prevention of foodborne pathogen contamination. Herein, a novel three-signal biosensor for simultaneous and ultrasensitive detection of live/dead Listeria monocytogenes (L.m) was developed. Concretely, a phage/prussian blue nanoparticles (PBNPs) and phage/polyethyleneimine@magnetic nanoparticles (PEI@MNPs) multifunctional probes were prepared. In the presence of L.m, a sandwich complex of phage/PBNPs-L.m-phage/PEI@MNPs was formed and separated by magnetic field. With the lytic activity of phages, the bioluminescent signal generated by adenosine triphosphate (ATP) was utilized for the quantification of live L.m. Photothermal and colorimetric signals triggered by the PBNPs could quantitative determination of total L.m. The combined results of total/live bacteria detection facilitate the calculation of dead bacteria. Bioluminescent signal allows for the precise detection of live L.m with a limit of detection (LOD) of 1 CFU/mL, while the photothermal and colorimetric signals enable on-site measurement of total bacteria with a LOD of 5 CFU/mL. This biosensor exhibits the multiple signals, realizing the diversification of bacterial information output. Meanwhile, the three signals can realize the mutually support, thereby improving the accuracy of detection results. This work opens a new avenue for the simple, ultrasensitive and simultaneous quantification of both live and dead bacteria.

Stand-off hazardous materials identification based on near-infrared hyperspectral imaging combined with convolutional neural network

Near-infrared (NIR) hyperspectral imaging enables rapid, non-contact imaging of hazardous materials in a non-destructive manner, allowing for analysis based on spectral reflection information. However, using traditional methods, it is challenging to identify hazardous materials with less distinct spectral reflection features. This study utilizes a self-built NIR hyperspectral imaging system and proposes a new approach. Using a convolutional neural network (CNN), This allows for the rapid completion of high-throughput spectral screening, marking suspicious spectra at spatial points. we sophisticatedly classified six hazardous material types, generating impactful warning images. The optimized CNN demonstrated superior performance (accuracy 91.08 %, recall 91.15 %, specificity 91.62 %, precision 90.17 %, and 0.924 F1 score) compared to SVM and KNN methods. Our study included multitask validation tests, revealing a sensitive detection of 10 mg/cm2 for ammonium nitrate and trinitrotoluene, capable of identifying over 100 targets simultaneously. By simulating real-world scenarios, we successfully detected hazardous chemicals scattered on the ground and accurately identified these hazardous materials in glass and thin plastic products. Even in situations where clothing obstructed the view, we could still correctly identify hazardous chemicals and generate corresponding warning images. Our system demonstrated precise identification capabilities even amidst complex backgrounds. This method provides an accurate and rapid solution for identifying and locating hazardous chemicals, laying a strong foundation for the next steps in non-contact, long-distance quantitative determination of chemical concentrations. This study highlights the effective application of CNN in non-contact, long-distance classification, and recognition of hazardous materials, paving the way for further scientific and engineering applications.

Accurate determination of three new antihypertensive drugs illegally added to food using ultra-high performance liquid chromatography-tandem mass spectrometry

Effective strategies are required to address food safety issues related to the illegal addition of antihypertensive drugs to food and claims of antihypertensive function. In this study, a novel ultra-high performance liquid chromatography-triple-quadrupole mass spectrometry (UHPLC-MS/MS) method was developed for the simultaneous determination of three antihypertensive drugs (azilsartan, candesartan cilexetil, and lacidipine) in 12 food matrices (pressed candies, solid beverages, alternative teas, tea drinks, biscuits, jellies, mixed liquors, oral liquids, medicinal teas, tablets, hard capsules, and soft capsules). Initially, mass spectrometry parameters, such as the collision energies of the three antihypertensive drugs, were optimized. Subsequently, the response intensities and chromatographic separation conditions of the three drugs in different mobile phases were compared. In addition, to enhance the recoveries, various extraction solvents and purification methods, including solid-phase extraction (SPE) columns and the QuEChERS technique, were investigated. In the developed method, sample determination involved three steps. First, the sample was extracted using 0.2% (v/v) formic acid in acetonitrile and then filtered using high-speed centrifugation, in addition, the extracted solution of alternative tea and medicinal tea was purified using the QuEChERS technique. Second, the supernatant was diluted with water, and filtered through a 0.22 μm polytetrafluoroethylene (PTFE) membrane. Finally, the analytes were separated on an Agilent Eclipse Plus RRHD C18 column (50 mm×2.1 mm, 1.8 μm) using a 5 mmol/L ammonium formate aqueous solution and acetonitrile as the mobile phases under gradient elution conditions and then detected using UHPLC-MS/MS with positive electrospray ionization (ESI) in the multiple reaction monitoring (MRM) mode. Quantitative analysis was performed using a matrix-matched external standard method. Methodological validation showed good linear relationships for all three antihypertensive drugs in the investigated concentration ranges, with correlation coefficients (r2) greater than 0.996. The limit of detection (LOD) and limit of quantification (LOQ) of lacidipine were 0.02 mg/kg and 0.04 mg/kg, respectively, whereas those of the other two drugs were 0.01 mg/kg and 0.02 mg/kg, respectively. In the 12 food matrices, the average recoveries of the drugs ranged from 86.6% to 107.5% with relative standard deviations (RSDs) of 1.1%-10.9% (n=6) at low, medium, and high spiked levels. Furthermore, this method was successfully applied to the analysis of real food samples. Overall, the newly developed method is simple, rapid, sensitive, accurate, and suitable for the qualitative and quantitative determination of antihypertensive drugs in different food matrices. This work could provide technical support for food safety agencies in implementing measures against the illegal addition of antihypertensive drugs to food.

An ultrasensitive liquid crystal aptasensing chip assisted by three-way junction DNA pockets for acrylamide detection in food samples

Acrylamide, an unsaturated amide found in heat-processed foods, poses serious risks to human health due to its neurotoxicity, carcinogenicity, and genotoxicity. This highlights the importance of quantitative determination of acrylamide in foods and the environments to ensure public health safety. Therefore, there is an urgent need for simple, rapid, and highly sensitive methods to accurately quantify acrylamide. In the present study, a user-friendly aptasensor was designed to quantify ultra-low levels of acrylamide in nuts for the first time. This innovative approach utilizes chemical engineering of a glass slide as a portable sensing platform, which incorporates liquid crystal (LC) molecules and a three-way junction (TWJ) DNA pocket. The immobilized TWJ pocket can disrupt the vertical alignment of LCs, turning the dark polarized background of the aptasensor to a colorful state. The binding of the specific aptamer to acrylamide disrupts the TWJ structure, enabling the LCs to return to their homotropic alignment. This structural change restores the dark polarized view of the sensing platform. The TWJ-engineered LC aptasensor effectively detects ultra-low concentrations of acrylamide in the range of 0.0005 to 50 fmol/L, with a detection limit of 0.106 amol/L. The aptasensor was successfully applied to real roasted nut samples, including peanut, almond, pistachio, and hazelnut, achieving recovery values ranging from 96.84 % to 99.61 %. With its simplicity, portability, ease of use, and cost-effectiveness, this aptasensor is a powerful sensing device for food safety monitoring.

Determination of three dechloranes in environmental water by magnetic solid-phase extraction-gas chromatography-negative chemical ionization mass spectrometry based on the covalent organic framework material

Dechloranes are additive-type chlorine flame retardants that are widely used in processing industrial products, such as electronic equipment and textiles. Dechloranes, which can enter the human body through various routes, pose significant health risks because of their toxicity, persistence, and bioaccumulation. In 2023, dechlorane plus was listed in the Stockholm Convention on Persistent Organic Pollutants. In the same year, China recognized this compound as a priority-controlled substance. Dechloranes are commonly found at trace levels in water, which is extremely harmful to the environment and human health. Therefore, the development of detection methods for dechloranes is crucial. Magnetic solid-phase extraction (MSPE) has attracted considerable attention because of its low organic solvent consumption, simplicity of adsorbent separation, and ease of operation. In general, the selectivity and efficiency of MSPE depend on the characteristics of the adsorbent. Covalent organic frameworks (COFs) have regular porosity, structural predictability and stability, high specific surface areas, and adjustable pore sizes, which are advantageous for a wide range of separation and analysis applications. In this study, Fe3O4 magnetic nanoparticles and a COF material (TpBD) were combined to prepare Fe3O4@TpBD as an adsorbent for dechloranes. Subsequently, an effective method for analyzing dechlorane in environmental water was established by coupling MSPE with gas chromatography-negative chemical ionization mass spectrometry (GC-NCI/MS). The successful synthesis of Fe3O4@TpBD was confirmed using transmission electron microscopy, scanning electron microscopy, Fourier transform infrared spectroscopy, X-ray diffraction, and vibrating sample magnetometry. A single-factor method was used to optimize the extraction conditions, including the Fe3O4@TpBD dosage, pH of water sample, elution solvent type and volume, extraction time, elution time, and ionic strength. The target analytes were separated on a TG-5SILMS column (30 m×0.25 mm×0.25 μm) and quantified using the external standard method in the selected-ion monitoring (SIM) mode. Under the optimal extraction conditions, the method validation results showed a linear range of 2-1000 ng/L. The limits of detection (LODs) and quantification (LOQs) were 0.18-0.27 ng/L and 0.60-0.92 ng/L, respectively, for the three analytes. The intra-day and inter-day precisions at three spiked levels were 4.2%-16.2% and 6.9%-15.7%, respectively. This method was successfully applied to the determination of dechloranes in environmental water samples (laboratory tap water, reservoir water, wastewater treatment plant effluent, and landfill leachate treatment effluent). The recoveries of the three dechloranes at different spiked levels ranged from 77.8% to 113.3% with relative standard deviations (RSDs) of 2.5%-16.3% (n=3). With the advantages of operational simplicity, high sensitivity, and good reproducibility, the proposed method is suitable for the qualitative and quantitative determination of dechloranes in environmental water.

Structure-Reactivity Relationships in a Small Library of Imine-Type Dynamic Covalent Materials: Determination of Rate and Equilibrium Constants Enables Model Prediction and Validation of a Unique Mechanical Softening in Dynamic Hydrogels.

The development of next generation soft and recyclable materials prominently features dynamic (reversible) chemistries such as host-guest, supramolecular, and dynamic covalent. Dynamic systems enable injectability, reprocessability, and time-dependent mechanical properties. These properties arise from the inherent relationship between the rate and equilibrium constants (RECs) of molecular junctions (cross-links) and the resulting macroscopic behavior of dynamic networks. However, few examples explicitly measure RECs while exploring this connection between molecular and material properties, particularly for polymeric hydrogel systems. Here we use dynamic covalent imine formation to study how single-point compositional changes in NH2-terminated nucleophiles affect binding constants and resulting hydrogel mechanical properties. We explored both model small molecule studies and model polymeric macromers, and found >3-decade change in RECs. Leveraging established relationships in the literature, we then developed a simple model to describe the cross-linking equilibrium and predict changes in hydrogel mechanical properties. Interestingly, we observed that a narrow ≈2-decade range of Keq's determine the bound fraction of imines. Our model allowed us to uncover a regime where adding cross-linker before saturation can decrease the cross-link density of a hydrogel. We then demonstrated the veracity of this predicted behavior experimentally. Notably this emergent behavior is not accounted for in covalent hydrogel theory. This study expands upon structure-reactivity relationships for imine formation, highlighting how quantitative determination of RECs facilitates predicting macroscopic behavior. Furthermore, while the present study focuses on dynamic covalent imine formation, the underlying principles of this work are applicable to the general bottom-up design of soft and recyclable dynamic materials.

Experimental determination of the sulfur K-shell fundamental parameters employing the holistic approach

Abstract Sulfur and its compounds are both very abundant in the environment and very important for technological applications such as batteries. X-ray spectroscopic characterizations of sulfur compounds for a quantitative determination of the present amount of sulfur often requires a good knowledge on the atomic fundamental parameters involved. These quantitatively describe the processes of photoionization and X-ray fluorescence emission, making them crucial for X-ray spectroscopy-based quantifications. 
By employing the recently demonstrated holistic approach, the atomic fundamental parameters of the sulfur K-shell were experimentally determined using the radiometrically calibrated instrumentation of the Physikalisch-Technische Bundesanstalt (PTB). The transition probabilities of the main K-shell fluorescence lines, the K-shell fluorescence yield, the K-shell Auger yield, the subshell photoionization cross section and fluorescence production cross section were determined by means of photon energy dependent X-ray fluorescence and transmission measurements on a thin InS coated silicon nitride membrane. The results are also downloadable from Zenodo as plain text.

Extraction of Anacardium occidentale: a kinetic, thermodynamic, phytochemical identification, and antibacterial study

Cashew tree biomass, Anacardium occidentale, represents an underutilized part of the plant and could have great potential for application in several areas, such as bioengineering, food science, and chemical. The stirred bed process was optimized to produce extracts from Anacardium occidentale stem bark (SBAo) and leaves (LAo) for bactericide purposes. Variables as alcohol content, stirring speed (SS), particle size (PS), solid-to-solvent ratio (S/S), temperature, and extraction time were evaluated. Subsequently, kinetic, thermodynamic and diffusivity fittings of the data were performed using different nonlinear models (Peleg, 2nd order, Elovich, power law, and Fick’s 2nd law) applied to the yield curves. Extracts were characterized by Fourier-transform infrared (FTIR) spectroscopy, total phenolic content (TPC), total flavonoid content (TFC), trolox equivalent antioxidant capacity (TEAC), and ultra-high resolution mass spectrometry (UHRMS) techniques for qualitative and quantitative determination of the phytochemicals. The optimized operating conditions obtained were ethanol in solvent at 70%, PS < 45 μm, 1200 rpm SS, 1/10 S/S ratio, and extraction time of 1 h at 50 ºC. The 2nd order kinetic model was the one that presented the best fits: SBAo (R2 > 0.9893 and RMSE < 3.4293) and LAo (R2 > 0.9985 and RMSE < 3.1117). Both A. occidentale extracts showed high levels of polyphenols and flavonoids, such as anacardic acid. Finally, the optimized extracts showed antibacterial susceptibility, with larger halos against S. aureus, 0.88± 0.07 mm (SBAo) and 0.45 ± 0.03 mm (LAo), in relation to E. coli, 0.80 ± 0.10 mm (SBAo) and 0.69 ± 0.08 mm (LAo).

Fluorescently probing anaerobic digester sludge: Measuring single-cell anabolic activity in methanogens (Methanosarcina and Methanothermobacter) with deuterium-labeled Raman analysis

This study addresses the challenge of obtaining in situ information on substrate utilization rates for individual microbial species in complex microbial communities such as anaerobic digester sludge. To overcome this hurdle, a novel approach combining doubly-labelled deuterium, fluorescence in situ hybridization (FISH) and Raman microspectroscopy was developed. The method enables quantitative determination of anabolic heavy hydrogen incorporation into FISH-targeted, exemplified by methanogenic cells from the genera Methanosarcina and Methanothermobacter.The deuterium incorporation rates ascertained by Raman red-shifting of C-Hx vibrational region to C-Dx vibrations, quantified through Raman peak area ratios, were compared for different carbon sources. Methanosarcina exhibited highest kinetic rates with acetate and propionate, while Methanothermobacter demonstrated faster incorporation under acetate and methanol supplementation.This groundbreaking study demonstrates the feasibility of obtaining quantitative metabolic rate information at a single-cell level using deuterium, FISH probes, and Raman microspectroscopy.

Determination of αs1-Casein and αs2-Casein in hypoallergenic infant formula by extended specific peptide -isotope-labeled internal standard dilution-LC-MS/MS

The desensitization effect of the hypoallergenic infant formula products is critical for production quality, and establishment of sensitive and accurate determination method of allergens, such as αs1-casein and αs2-casein, remains challengeable in the field of infant formula industry. Therefore, the objective of this study was to identify specific peptides (YLGYLEQLLR and FALPQYLK) from αs1-casein and αs2-casein in infant formula by orbitrap high resolution mass spectrometry (Orbitrap MS), then establish an analytical method for the qualitative and quantitative determination of αs1-casein and αs2-casein based on extended specific peptide-isotope-labeled internal standard dilution-LC-MS/MS. Specific peptides obtained by trypsin enzymolysis of hypoallergenic infant formula were separated by Waters Acquity LC HSS T3 column, and analyzed under ESI positive ion mode using multi-reaction monitoring (MRM). The specific peptides of αs1-casein and αs2-casein exhibited linear relationships within the concentration range of 0.0025–1.5 μmol/L and correlation coefficients were above 0.99. The detection limits of αs1-casein and αs2-casein were 0.3 mg/kg, with quantification limits of 1.0 mg/kg, respectively. The average recoveries were 83.5%–87.3% (αs1-casein) and 80.1%–84.4% (αs2-casein), with RSDs of 2.3%–3.7% (αs1-casein) and 2.5%–5.0% (αs2-casein), respectively. Our results demonstrated that this method is capable of analyzing the content of αs1-casein and αs2-casein in hypoallergenic infant formula with excellent sensitivity and specificity.

POD-like nanozyme constructed from perspective of charge transfer engineering for biosensing of magnetic separation treated Listeria monocytogenes

For foodborne pathogens pose a serious threat to public health, a magnetic separation strategy and a nanozyme-based biosensor are proposed for biosensing of Listeria monocytogenes (L. monocytogenes). In this work, doripenem is selected as a recognized molecule for the modification of magnetic beads to capture L.monocytogenes in food and environmental samples. Furthermore, the POD-like MXene-Hemin-Au is constructed from perspective of charge transfer engineering which provides a vivid example to rational design of nanozymes. Finally, the captured L.monocytogenes is labeled with MXene-Hemin-Au@mAb, forming the sandwich complexes for quantitative determination. The current signals that generated by the complexes exhibit a good linear relationship with a limit of detection of 2.3 × 101 CFU/mL. The biosensor shows a satisfactory applicability in real samples with recoveries of 91.19% to 102.98%. Overall, the biosensor with integrated magnetic separation strategy presents a potential approach for high sensitivity biosensing of foodborne pathogens.