Quantification Of Acids Research Articles

Behavioral comorbidities treatment by fecal microbiota transplantation in canine epilepsy: a pilot study of a novel therapeutic approach.

Anxiety and cognitive dysfunction are frequent, difficult to treat and burdensome comorbidities in human and canine epilepsy. Fecal microbiota transplantation (FMT) has been shown to modulate behavior in rodent models by altering the gastrointestinal microbiota (GIM). This study aims to investigate the beneficial effects of FMT on behavioral comorbidities in a canine translational model of epilepsy. Nine dogs with drug-resistant epilepsy (DRE) and behavioral comorbidities were recruited. The fecal donor had epilepsy with unremarkable behavior, which exhibited a complete response to phenobarbital, resulting in it being seizure-free long term. FMTs were performed three times, two weeks apart, and the dogs had follow-up visits at three and six months after FMTs. Comprehensive behavioral analysis, including formerly validated questionnaires and behavioral tests for attention deficit hyperactivity disorder (ADHD)- and fear- and anxiety-like behavior, as well as cognitive dysfunction, were conducted, followed by objective computational analysis. Blood samples were taken for the analysis of antiseizure drug (ASD) concentrations, hematology, and biochemistry. Urine neurotransmitter concentrations were measured. Fecal samples were subjected to analysis using shallow DNA shotgun sequencing, real-time polymerase chain reaction (qPCR)-based Dysbiosis Index (DI) assessment, and short-chain fatty acid (SCFA) quantification. Following FMT, the patients showed improvement in ADHD-like behavior, fear- and anxiety-like behavior, and quality of life. The excitatory neurotransmitters aspartate and glutamate were decreased, while the inhibitory neurotransmitter gamma-aminobutyric acid (GABA) and GABA/glutamate ratio were increased compared to baseline. Only minor taxonomic changes were observed, with a decrease in Firmicutes and a Blautia_A species, while a Ruminococcus species increased. Functional gene analysis, SCFA concentration, blood parameters, and ASD concentrations remained unchanged. Behavioral comorbidities in canine IE could be alleviated by FMT. This study highlights FMT's potential as a novel approach to improving behavioral comorbidities and enhancing the quality of life in canine patients with epilepsy.

One-step microwave-assisted synthesis of fluorescent carbon quantum dots for determination of ascorbic acid and riboflavin in vitamin supplements

The synthesis of carbon quantum dots (CQDs) using chemical precursors with different organic groups is a strategy to improve optical properties and expand applications in several fields of research such as Analytical Chemistry. Ascorbic acid and riboflavin are widely used in human food supplementation, making quality monitoring of these vitamin supplements relevant and necessary. In this work, disodium ethylenediaminetetraacetic, sodium thiosulfate and urea were applied to obtain CQDs through a single-step microwave-assisted synthesis. The CQDs were characterized by transmission electron microscopy, X-ray photoelectron spectroscopy, X-ray powder diffraction, infrared spectroscopy, zeta potential measurements, ultraviolet–visible spectroscopy and photoluminescence spectroscopy. The synthesized nanoparticles exhibited satisfactory and stable optical properties with luminescence at 430 nm, water solubility, and fluorescence quantum yield of 8.9 %. They were applied in the quantification of ascorbic acid and riboflavin in vitamin supplements. The fluorescence mechanisms observed were dynamic quenching for the CQDs/Cr(VI) sensor, followed by a return of fluorescence in the presence of ascorbic acid, and static quenching and inner filter effect in the interaction with riboflavin. Factorial designs 23 and 24 were used to optimize the analytical parameters. The CQDs/Cr(VI) sensor used in the determination of ascorbic acid, employing an on–off-on strategy, resulted in a linear range of 0.5 to 50 µg mL−1 and a limit of detection of 0.15 µg mL−1. The ratiometric fluorescence used in the determination of riboflavin resulted in a linear range of 0.1 to 7 µg mL−1 and a limit of detection of 0.09 µg mL−1. The analytical results for ascorbic acid were compared to the reference method of the Brazilian pharmacopeia, showing accuracy and precision according to the Brazilian Health Regulation Agency. Therefore, the synthesized CQDs were used to determine ascorbic acid and riboflavin in vitamin supplements, and the application of this nanomaterial can be expanded to different analytes and matrices, using simple and low-cost analysis techniques.

Comparison of DeNovix, NanoDrop and Qubit for DNA quantification and impurity detection of bacterial DNA extracts.

Accurate DNA quantification is key for downstream application including library preparations for whole genome sequencing (WGS) and the quantification of standards for quantitative PCR. Two commonly used technologies for nucleic acid quantification are based on spectrometry, such as NanoDrop, and fluorometry, such as Qubit. The DS-11+ Series spectrophotometer/fluorometer (DeNovix) is a UV spectrophotometry-based instrument and is a relatively new spectrophotometric method but has not yet been compared to established platforms. Here, we compared three DNA quantification platforms, including two UV spectrophotometry-based techniques (DeNovix and NanoDrop) and one fluorometry-based approach (Qubit). We used genomic prokaryotic DNA extracted from Streptococcus pneumoniae using a Roche DNA extraction kit. We also evaluated purity assessment and effect of a single freeze-thaw cycle. Spectrophotometry-based methods reported 3 to 4-fold higher mean DNA concentrations compared to Qubit, both before and after freezing. The ratio of DNA concentrations assessed by spectrophotometry on the one hand, and Qubit on the other hand, was function of the A260/280. In case DNA was pure (A260/280 between 1.7 and 2.0), the ratio DeNovix or Nanodrop vs. Qubit was close or equal to 2, while this ratio showed an incline for DNA with increasing A260/280 values > 2.0. The A260/280 and A260/230 purity ratios exhibited negligible variation across spectrophotometric methods and freezing conditions. The comparison of DNA concentrations from before and after freezing revealed no statistically significant disparities for each technique. DeNovix exhibited the highest Spearman correlation coefficient (0.999), followed by NanoDrop (0.81), and Qubit (0.77). In summary, there is no difference between DeNovix and NanoDrop in estimated gDNA concentrations of S. pneumoniae, and the spectrophotometry methods estimated close or equal to 2 times higher concentrations compared to Qubit for pure DNA.

Bioinspired amino acid-functionalized cobalt ferrite nanocomposite: A nanozyme-based colorimetric sensor for sensitive and selective quantification of phenolic compounds and ascorbic acid antioxidant capacity

The escalating oxidative stress has heightened the daily human demand for diverse antioxidants. Therefore, development of the novel approaches to assess the total antioxidant capacity (TAC) of various nutrients is essential. In this study, drawing inspiration from the active site of native peroxidase enzymes, a novel peroxidase (POD)-like nanozyme was developed based on the cobalt ferrite (CoFe2O4) nanoparticles functionalized with different catalytic amino acids. Based on the TMB/H2O2 colorimetric system, the most substantial enhancement in POD-like activity was obtained by the glutamic acid coating among different charged amino acids studied, with more than 74% increase in specific activity compared to the bare CoFe2O4. A signal-off colorimetric sensing platform based on the obtained nanobiocatalyst was developed for the accurate quantification of the antioxidant capacity of phenolic compounds and vitamin C. The sensitive and selective quantification of ascorbic acid, tannic acid, gallic acid, cyanidin-3-glucoside, and quercetin was obtained by this colorimetric method.

Novel and Extremely Sensitive NiAl2O4-NiO Nanostructures on an ITO Sensing Electrode for Enhanced Detection of Ascorbic Acid.

The current study focused on the design of an extremely sensitive electrochemical sensor of ascorbic acid based on a mixture of NiAl2O4-NiO nanoparticles that, produced in a single step using the sol-gel method, on an ITO electrode. This new sensing platform is useful for the detection of ascorbic acid with a wide range of concentrations extending from the attomolar to the molar. SEM micrographs show the porous structure of the NiAl2O4-NiO sample, with a high specific surface area, which is beneficial for the catalytic performance of the nanocomposite. An XRD diffractogram confirmed the existence of two phases, NiAl2O4 and NiO, both corresponding to the face-centred cubic crystal structure. The performances of the modified electrode, as a biomolecule, in the detection of ascorbic acid was evaluated electrochemically by cyclic voltammetry and chronoamperometry. The sensor exhibited a sensitive electrocatalytic response at a working potential of E = +0.3 V vs. Ag/Ag Cl, reaching a steady-state current within 30 s after each addition of ascorbic acid solution with a wide dynamic range of concentrations extending from attolevels (10-18 M) to molar (10 mM) and limits of detection and quantification of 1.2 × 10-18 M and 3.96 × 10-18 M, respectively. This detection device was tested for the quantification of ascorbic acid in a 500 mg vitamin C commercialized tablet that was not pre-treated.

Establishment and clinical application of a candidate reference measurement procedure for quantification of urinary vanillylmandelic acid and homovanillic acid using ID-LC-MS/MS method

Neuroblastoma (NB), an embryonic tumor of the autonomous nervous system, poses a significant threat to the health and lives of children. Accurate measurement of vanillylmandelic acid (VMA) and homovanillic acid (HVA) in human urine is crucial for screening and diagnosis of NB. Although various laboratories have developed liquid chromatography tandem mass spectrometry (LC-MS/MS) method to detect VMA and HVA, the comparability between the results obtained from different laboratories and methods was poor. The absence of reference method for VMA and HVA hinders the standardization of their measurements. Therefore, a candidate reference measurement procedure (cRMP) based on isotope dilution LC-MS/MS (ID-LC-MS/MS) for the detection of VMA and HVA in human urine was established. Urine samples were spiked with VMA-d3 and HVA-d5 as internal standards and extracted using a protein precipitation method. The cRMP exhibited desirable precision with the total imprecision below 5 %. The accuracy of this cRMP was demonstrated by the high analytical recovery (98.64 % - 102.22 % and 98.41 % - 100.97 % for VMA and HVA, respectively), and comparability between different reference systems. The limit of detection for HVA and VMA were 15.625 ng/mL and 3.906 ng/mL, respectively; the quantification limits were 62.5 ng/mL and 7.813 ng/mL, respectively, which can meet the clinical detection requirements. The linear range was from 78.125 ng/mL to 20 μg/mL. Specificity evaluations showed no corresponding interference from structurally similar analogs. In conclusion, we have established a cRMP based on ID-LC-MS/MS for the measurement of VMA and HVA in urine samples, demonstrating well-defined method performance including accuracy, precision, and specificity. This newly established cRMP is suitable for routine assay standardization and evaluation of clinical samples. Furthermore, this method has the potential to significantly enhance the diagnostic accuracy for neuroblastoma.

In vitro evaluation of algae and their effect as dietary ingredient on growth, chemical composition and intestinal functionality in juvenile turbot (Scophthalmus maximus)

The objectives of this study were to evaluate the in vitro protein hydrolysis of five algae (Arthrospira sp., Chlorella sp., Tisochrysis sp., Microchloropsis sp. and Ulva rigida) by the digestive proteases of Scophthalmus maximus, and to assess the effect of feeding turbot juveniles on diets supplemented with these algae in a 60-day feeding trial. The in vitro results revealed that algal protein was hydrolysed by turbot digestive proteases. Besides, the quantification of total amino acids released during the in vitro hydrolysis evidenced noticeable amino acid bioavailability, mainly in the case of Arthrospira sp. with up to 42.7% of the amino acids released after 120 min of in vitro hydrolysis. However, the structural characteristics of cell wall of each species influenced the protein accessibility as evidenced by the lower values obtained in Microchloropsis sp. For the in vivo feeding trial, 255 specimens of turbot, with an average initial weight of 26.4 ± 7.6 g, were used, and were fed with diets including 5% and 10% of a blend of the same microalgae tested in vitro (M5 and M10) or U. rigida (U5 and U10), compared to a control feed without algal biomass (CT). At the end of the feeding trial, growth performance and nutrient utilization did not show differences among the experimental groups. In addition, the fish fed with the diets supplemented with algae showed lower lipid oxidation in the muscle, and an improvement in digestive functionality, as the activity of pancreatic and intestinal enzymes increased. The ultrastructural study of the intestinal mucosa revealed an increase in the length of the microvilli and the absorptive surface in fish fed with the algae-supplemented diets, as well as decreased thickness of the lamina propria and the number of goblet cells. These results confirmed that the supplementation of turbot juvenile diets with low percentage of microalgae or U. rigida is appropriate for improving digestive functionality, as well as reducing the oxidation of lipids in the muscle.

Anaerobic digestion model number 1 applied to the modeling of anaerobic digestion of residues generated in soluble coffee processing

The Anaerobic Digestion Model No. 1 (ADM1) was applied in this study to simulate the anaerobic digestion (AD) of wastewaters from soluble coffee processing in sequential batch reactors. As these substrates present considerable levels of phenolic compounds, modifications were made to the basic structure of the ADM1. Additionally, the Luong inhibition model replaced the Monod model to describe the consumption of soluble organic matter. On the other hand, the removal of phenolic compounds was carried out using the Haldane model, appropriate when the inhibition by excess substrate is present, taking into account in this case the quantification of phenols and benzoic acid as total phenols. The new ADM1-based model was calibrated with experimental data from the AD of post hydrothermal wastewater (PHWW) of spent coffee grounds (SCG) and validated with experimental data of the AD of effluents produced during the soluble coffee processing. The simulation results demonstrated that the new model moderately described the removal of phenolic compounds ( > 90 %), methane production (2.243 kgCODm−3 in calibration and 6.16 kgCODm−3 in validation), and the production and consumption of organic acids over time. However, in future research new metabolic pathways may be included in the model to improve its quality even more.

A new chemical derivatization reagent sulfonyl piperazinyl for the quantification of fatty acids using LC-MS/MS

In our previous study, a chemical derivatization reagent named 5-(dimethylamino) naphthalene-1-sulfonyl piperazine (Dns-PP) was developed to enhance the chromatographic retention and the mass spectrometric response of free fatty acids (FFAs) in reversed-phase liquid chromatography coupled with electrospray ionization-mass spectrometry (RPLC-ESI-MS). However, Dns-PP exhibited strong preferences for long-chain FFAs, with limited improvement for short- or medium-chain FFAs. In this study, a new series of labeling reagents targeting FFAs were designed, synthesized, and evaluated. Among these reagents, Tmt-PP (N2, N2, N4, N4-tetramethyl-6-(4-(piperazin-1-ylsulfonyl) phenyl)-1,3,5-triazine-2,4-diamine) exhibited the best MS response and was selected for further evaluations. We compared Tmt-PP with Dns-PP and four commonly used carboxyl labeling reagents from existing studies, demonstrating the advantages of Tmt-PP. Further comparisons between Tmt-PP and Dns-PP in measuring FFAs from biological samples revealed that Tmt-PP labeling enhanced the MS response for about 80 % (30/38) of the measured FFAs, particularly for short- and medium-chain FFAs. Moreover, Tmt-PP labeling significantly improved the chromatographic retention of short-chain FFAs. To ensure accurate quantification, we developed a stable isotope-labeled Tmt-PP (i.e., d12-Tmt-PP) to react with chemical standards and serve as one-to-one internal standards (IS). The method was validated for accuracy, precision, sensitivity, linearity, stability, extraction efficiency, as well as matrix effect. Overall, this study introduced a new chemical derivatization reagent Tmt-PP (d12-Tmt-PP), providing a sensitive and accurate option for quantifying FFAs in biological samples.

PROMER technology: a new real-time PCR tool enabling multiplex detection of point mutation with high specificity and sensitivity

Abstract Real-time polymerase chain reaction (real-time PCR) is a powerful tool for the precise quantification of nucleic acids in various applications. In cancer management, the monitoring of circulating tumor DNA (ctDNA) from liquid biopsies can provide valuable information for precision care, including treatment selection and monitoring, prognosis, and early detection. However, the rare and heterogeneous nature of ctDNA has made its precise detection and quantification challenging, particularly for ctDNA containing hotspot mutations. We have developed a new real-time PCR tool, PROMER technology, which enables the precise and sensitive detection of ctDNA containing cancer-driven single point mutations. The PROMER functions as both a PRObe and priMER, providing enhanced detection specificity. We validated PROMER technology using synthetic templates with known KRAS point mutations and demonstrated its sensitivity and linearity of quantification. Using genomic DNA from human cancer cells with mutant and wildtype KRAS, we confirmed that PROMER PCR can detect mutant DNA. Furthermore, we demonstrated the ability of PROMER technology to efficiently detect mutation-carrying ctDNA from the plasma of mice with human cancers. Our results suggest that PROMER technology represents a promising new tool for the precise detection and quantification of DNA containing point mutations in the presence of a large excess of wildtype counterpart.

Impact of high-fat diet on ovarian epigenetics: Insights from altered intestinal butyric acid levels

ObjectiveTo investigate the effects of a high-fat diet (HFD) on the gut bacterium Roseburia intestinalis and butyric acid levels, and to assess their impact on ovarian function and epigenetic markers in mice. MethodsA total of 20 female ICR mice aged 4 weeks were randomly assigned to two groups and fed either a control diet (CD) or an HFD for 36 weeks. Post-intervention, ileal contents were analyzed for the quantification of butyric acid using ELISA, while feces were obtained for Roseburia intestinalis expression assessment via qPCR. Histological evaluations of intestinal and ovarian tissues included H&E and Alcian Blue-Periodic Acid Schiff (AB-PAS) staining, alongside immunohistochemical analysis for F4/80, and immunofluorescent detection of Occludin, ZO-1, 5 mC, and H3K36me3. Ovarian health was assessed through follicle counts and morphological evaluations. Statistical analyses were performed using GraphPad Prism 8.0, with P < 0.05 considered significant. ResultsAfter 36 weeks, the HFD group showed significantly higher body weight compared to the CD group (P < 0.01). The HFD led to a decrease in Roseburia intestinalis and butyric acid levels, a reduction in intestinal goblet cells, and an increase in intestinal inflammation. Histological analyses revealed impaired ovarian follicular development and enhanced inflammation in the HFD mice, with immunofluorescent staining showing downregulation of the ovarian epigenetic markers 5 mC and H3K36me3. ConclusionOur study demonstrates that long-term HFD negatively impacts ovarian function and epigenetic regulation. We found decreased levels of the gut bacterium Roseburia intestinalis and its metabolite, butyric acid, which contribute to these adverse effects. Additionally, the associated intestinal inflammation and compromised mucosal barrier may contribute to these adverse outcomes on female reproductive health.

Feeding dogs a high-fat diet induces metabolic changes similar to natural aging, including dyslipidemia, hyperinsulinemia, and peripheral insulin resistance.

The goal of this study was to characterize changes induced by a high-fat diet in body composition, insulin levels and sensitivity, blood lipids, and other key biomarkers also associated with the metabolic dysfunction that occurs with natural aging. 24 male Beagle dogs, 3 to 7 years of age, of mixed castration status. Dogs were randomly assigned to continue twice daily feeding of the commercial adult maintenance diet (n = 12, including 2 intact) that they were previously fed or to a high-fat diet (12, including 2 intact) for 17 weeks between December 1, 2021, and April 28, 2022. Assessments included body composition (weight, body condition score, and adipose mass determined by deuterium enrichment), clinical chemistries, plasma fatty acid quantification, oral glucose tolerance test, and histology of subcutaneous and visceral adipose biopsy samples. The high-fat diet led to increased body weight, body condition score, fat mass and adipocyte size, hyperinsulinemia and peripheral insulin resistance, and elevations in serum lipids, including cholesterol, triglycerides, and several species of free fatty acids. Leptin levels increased in dogs fed a high-fat diet but not in control dogs. There were no significant changes in routine clinical chemistry values in either group. Feeding a high-fat diet for 17 weeks led to potentially deleterious changes in metabolism similar to those seen in natural aging in dogs, including hyperinsulinemia, insulin resistance, and dyslipidemia. A high-fat diet model may provide insights into the similar metabolic dysfunction that occurs during natural aging.

An Optimized Method for LC-MS-Based Quantification of Endogenous Organic Acids: Metabolic Perturbations in Pancreatic Cancer.

Accurate and reliable quantification of organic acids with carboxylic acid functional groups in complex biological samples remains a major analytical challenge in clinical chemistry. Issues such as spontaneous decarboxylation during ionization, poor chromatographic resolution, and retention on a reverse-phase column hinder sensitivity, specificity, and reproducibility in multiple-reaction monitoring (MRM)-based LC-MS assays. We report a targeted metabolomics method using phenylenediamine derivatization for quantifying carboxylic acid-containing metabolites (CCMs). This method achieves accurate and sensitive quantification in various biological matrices, with recovery rates from 90% to 105% and CVs ≤ 10%. It shows linearity from 0.1 ng/mL to 10 µg/mL with linear regression coefficients of 0.99 and LODs as low as 0.01 ng/mL. The library included a wide variety of structurally variant CCMs such as amino acids/conjugates, short- to medium-chain organic acids, di/tri-carboxylic acids/conjugates, fatty acids, and some ring-containing CCMs. Comparing CCM profiles of pancreatic cancer cells to normal pancreatic cells identified potential biomarkers and their correlation with key metabolic pathways. This method enables sensitive, specific, and high-throughput quantification of CCMs from small samples, supporting a wide range of applications in basic, clinical, and translational research.

Determination of succinic acid and lactic acid in pigs' serum, intestinal contents, and meat by ultrahigh-performance liquid chromatography-tandem mass spectrometry.

Succinic acid and lactic acid have been associated with diarrhea in weaned piglets. The level of succinic acid and lactic acid in serum, meat, and intestinal contents is important to elucidate the mechanism of diarrhea in weaned piglets. A facile method was developed for the quantification of succinic acid and lactic acid in pigs' serum, intestinal contents, and meat using ultrahigh-performance liquid chromatography-tandem mass spectrometry (UHPLC/MS/MS). The serum samples underwent protein precipitation with methanol. The meat and intestinal contents were freeze-dried and homogenized using a tissue grinding apparatus. Methanol-water mixture (80:20, v/v) was used for homogenizing the meat, while water was used for homogenizing the intestinal contents. An additional step of protein precipitation with acetonitrile was required for the intestinal contents. The resulting solution was diluted with water before being analyzed by UHPLC/MS/MS. Separation of succinic acid and lactic acid could be achieved within 3min using a Kinetic XB-C18 column. The coefficients of variation for peak areas of succinic acid and lactic acid were less than 5.0%. The established method demonstrated good linearity as indicated by correlation coefficients exceeding 0.996. Additionally, satisfactory recoveries ranging from 88.58% to 108.8% were obtained. The detection limits (RS/N = 3) for succinic acid and lactic acid were determined to be 0.75 ng/mL and 0.02 μg/mL, respectively. This method exhibited high sensitivity, simplicity in operation, and small sample weight, making it suitable for quantitative determination of succinic acid and lactic acid in pigs' serum, intestinal contents, and meat. The method developed will provide valuable technical support in studying the metabolic mechanisms of succinic acid and lactic acid in pigs.

A look inside the gut as a clue to nutritional status.

Malabsorption and malnutrition are common gastrointestinal manifestations clinicians face, requiring diagnostic workup for effective diagnosis and management of the underlying cause. This review discusses recent advances in diagnostic approaches to malabsorption and maldigestion of macronutrients - lipids, proteins, and carbohydrates. We highlight underrecognized causes, available testing modalities, and ongoing diagnostic unmet needs. Innovations in the diagnostic landscape are enhancing our understanding of malabsorption syndromes. Stool collection and handling is uncomfortable and commonly avoided. The objective quantification of stool lipids, bile acids, and gut enzymes is therefore underused in the diagnosis and management of common disorders such as exocrine pancreatic insufficiency, bile acid diarrhea, protein-losing enteropathy, and more. We review the recent advancements in spot quantification of stool fat and bile acid content, endoscopic imaging techniques such as endocytoscopy, confocal laser endomicroscopy, and optical coherence tomography and the future place in clinical practice. Malabsorption and maldigestion represent significant challenges in clinical nutrition and gastroenterology. Through the integration of advanced diagnostic techniques, clinicians will be better equipped to tailor therapy and monitor treatment response, ultimately improving patient health outcomes. This review underscores the critical role of innovative diagnostic tools in accurately detecting and effectively managing gastrointestinal disorders linked to nutritional status.

Developing centrifugal force real-time digital PCR for detecting extremely low DNA concentration

Digital PCR (dPCR) is a technique for absolute quantification of nucleic acid molecules. To develop a dPCR technique that enables more accurate nucleic acid detection and quantification, we established a novel dPCR apparatus known as centrifugal force real-time dPCR (crdPCR). This system is efficient than other systems with only 2.14% liquid loss by dispensing samples using centrifugal force. Moreover, we applied a technique for analyzing the real-time graph of the each micro-wells and distinguishing true/false positives using artificial intelligence to mitigate the rain, a persistent issue with dPCR. The limits of detection and quantification were 1.38 and 4.19 copies/μL, respectively, showing a two-fold higher sensitivity than that of other comparable devices. With the integration of this new technology, crdPCR will significantly contribute to research on next-generation PCR targeting absolute micro-analysis.

Gold nanoparticles decorated kaolinite mineral modified screen-printed electrode: Use for simple, sensitive determination of gallic acid in food samples

The current study offers the nanomolar quantification of gallic acid (GAL) based on gold nanoparticles (AuNps) and kaolinite minerals (KNT) modified on a screen-printed electrode (SPE). The electrochemical behavior of GAL was performed using differential pulse voltammetry (DPV) in Britton Robinson (BR) buffer solution at pH 2.0 as a supporting electrolyte. Under the optimized DPV mode parameters, the oxidation peak current of GAL (at 0.72 V vs Ag/AgCl) increased linearly in the range between 0.002 μmolL−1 and 40.0 μmolL−1 with a detection limit of 0.50 nmolL−1. The effect of common interfering agents was also investigated. Finally, the applicability of the proposed method was verified by quantification analysis of GAL in black tea and pomegranate juice samples.

Digital Sort-Enabled Counting Allows Absolute Electrical Quantification of Target Nucleic Acid.

Quantitative nucleic acid amplification tests are of great importance for diagnostics, but current approaches require complex and costly optical setups that limit their nonlaboratory applications. Herein we describe the implementation of a microfluidics platform that can perform binary DNA-amplification-activated droplet sorting. The digital sort-enabled counting (DISCO) platform enables label-free absolute quantification of the nucleic acid. This is achieved by provoking a pH change in droplets through a loop-mediated isothermal amplification (LAMP) reaction, followed by using sorting by interfacial tension (SIFT) to direct positive and negative droplets to different outlets. With the use of on-chip electrodes at both outlets, we demonstrate that the digital electrical counting of target DNA and RNA can be realized. DISCO is a promising approach for realizing sensitive nucleic acid quantification in point-of-care settings.

High-Performance Liquid Chromatographic Quantification of the Plant Hormone Abscisic Acid at ppb Levels in Plant Samples after a Single Immunoaffinity Column Cleanup.

High-performance liquid chromatography with ultraviolet detection (HPLC-UV) is a common analysis technique due to its high versatility and simple operation. In the present study, HPLC-UV detection was integrated with immunoaffinity cleanup (IAC) of the sample extracts. The matrix effect was greatly reduced, and the limit of detection was as low as 1 ng/g of free abscisic acid (ABA) in fresh plant tissues. A monoclonal antibody 3F1 (mAb 3F1) was developed to specifically recognize free ABA but not ABA analogues. The mAb 3F1-immobilized immunoaffinity column exhibited a capacity of 850 ng/mL and an elution efficiency of 88.8-105% for standards. The extraction recoveries of the column for ABA ranged from 80.4 to 108.9%. ABA content was detected in various plant samples with IAC-HPLC-UV. The results were verified with ultraperformance liquid chromatography-electrospray tandem mass spectrometry. IAC-HPLC-UV can be a sensitive and cost-efficient method for plant hormone analysis.

Optimized quantification of fatty acids in complex media using advanced analytical techniques.

Various medium formulations contain essential fatty acids at concentrations ranging from 10 to 100 mg/L. Accurate and precise lipid measurement in media is crucial for monitoring media quality and conducting studies on lipids in the context of cell culture. This study employed two-dimensional gas chromatography (GC × GC) analyses to offer enhanced resolution, sensitivity, and separation performance compared to GC. Quantification of fatty acid methyl esters (FAMEs) in a medium was conducted using GC × GC combined with a high-resolution mass spectrometer and flame ionization detector, considering potential interference from nonionic surfactant Tween 80, which was precipitated and removed by optimizing the concentration of cobalt thiocyanate (CTA) solution during pretreatment. This advanced analytical approach enabled identification of cis and trans isomers of identical molecular weights and determination of the location and number of double bonds in the same carbon number structure. Our analysis identified 36 FAMEs within the C6-C24 region, and a 5% CTA solution was optimal for efficient removal of Tween 80 during lipid extraction. Additionally, this advanced method minimized FAME contamination and loss during pretreatment, thereby significantly reducing the sample volume required to detect trace levels of FAMEs. This improvement led to a fatty acid recovery rate of 106% while maintaining the average relative standard deviation for the target FAMEs of about 3%. Our research paves the way for future investigation into medium quality control and the role of fatty acids in cell culture. This offers the possibility for economical and effective trace quantification of fatty acids in complex media.