Sensitive Method Research Articles

Near-Infrared Fluorescent Probe Reveals Elevated Mitochondrial Viscosity during Acute Alcoholic Liver Injury.

Acute alcoholic liver injury (AALI) has become an important cause of liver disease worldwide, and there is an urgent need to develop noninvasive and sensitive methods to detect and evaluate AALI. We report herein three novel but readily available mitochondrial targeting fluorescence probes (ICR, ICJ, and ICQ) for AALI detection. These probes contain different electron-donating groups, among which ICQ exhibits NIR fluorescence (740 nm), a large Stokes shift (110 nm), and a sensitive response to viscosity (73-fold enhancement in fluorescence from water to glycerol), making it suitable for in vivo imaging. ICQ also exhibits an excellent ability to image mitochondrial viscosity changes in cells. More importantly, ICQ can target the liver selectively and image the viscosity changes in the liver noninvasively. Through establishing an AALI mouse model, ICQ was successfully applied to the in situ imaging changes in liver viscosity during the AALI process. The results showed a significant increase in liver viscosity in AALI mice, indicating that viscosity can serve as a marker for AALI, and ICQ is a promising noninvasive and sensitive tool for detecting and evaluating AALI.

Direct competitive immunoassay method for sensitive detection of the histamine in foods based on a MI-Cu-GMP nanozyme marker and molecularly imprinted biomimetic antibody.

Histamine may lead to low blood pressure, skin flushing and edema when it accumulates in large amounts in the body. Therefore, establishing sensitive methods for the detection of histamine in foods is extremely important to ensure food safety and human health. The MI-Cu-GMP NPs (2-methylimidazole-copper-guanosine monophosphate nanozymes) with high laccase-like activity were synthesized. Using the prepared molecular imprinted membrane as biomimetic antibody and MI-Cu-GMP NPs as marker, a sensitive direct competitive biomimetic enzyme-linked immunoassay (BELISA) method for rapid detection of the histamine in foods was developed. Under optimal conditions, the limit of detection (LOD, IC15) and sensitivity (IC50) of the BELISA method for histamine was 0.05 mg L-1 and 1.22 mg L-1, respectively. The liquor samples spiked with histamine was detected by the BELISA method with satisfactory recoveries ranging from 90.00% to 116.00%. Further, the level of histamine in three samples (cooking wine, rice vinegar and soy sauce) was tested by the BELISA and high-performance liquid chromatography (HPLC), with no significant difference found between the two methods. Given the advantages, the established BELISA method is expected to provide practical guidance for the monitoring of histamine in food and provides a foundation for the detection of other food hazards. © 2024 Society of Chemical Industry.

Fluorescence/colorimetric dual-signal sensor based on carbon dots and gold nanoparticles for visual quantification of Cr3.

A convenient and sensitive dual-signal visualization method is constructed for detection of trivalent chromium ions (Cr3+) based on fluorescent carbon dots (CDs) and glutathione-modified gold nanoparticles (GSH-Au NPs). The fluorescence of CDs can be quenched by GSH-Au NPs due to the inner filter effect. Cr3+ induces aggregation of GSH-Au NPs because of the coordination with GSH on the surface of Au NPs, leading to the red shift of surface plasmon resonance absorption of Au NPs that provides a "turn-on" fluorescence and colorimetric assay for Cr3+. The fluorescence/colorimetric dual signal detection shows high sensitivity for Cr3+ with wide detection linear ranges (0.5-70μM for fluorescence detection and 2-50μM for colorimetric detection) and low detection limits (0.31μM for fluorescence detection and 0.30μM for colorimetric detection). Besides, the method has high selectivity for Cr3+ and can be used for detection of Cr3+ in lake water and tap water samples, showing great potential for visual detection of environmental Cr3+.

Pharmacokinetic study of iptacopan and its two acyl glucuronide metabolites in monkey plasma by liquid chromatography combined with electrospray ionization tandem mass spectrometry.

In this study, a simple and sensitive liquid chromatography tandem mass spectrometric method was developed and validated for the determination of iptacopan and two acyl glucuronidation metabolites in monkey plasma. The plasma sample was precipitated with acetonitrile and then separated on an Acquity UPLC BEH C18 column (2.1 × 100 mm, 1.7 μm) using 0.1% formic acid and 5 mM ammonium acetate in water and acetonitrile as the mobile phase. The mass spectrometry (MS) detection was performed in positive multiple reactions monitoring (MRM) mode with precursor-to-production transitions. The developed assay was validated over the range of 1-2000 ng/mL for three analytes with correlation coefficient (r) more than 0.99. The validation parameters including accuracy, precision, carryover effect, matrix effect, recovery, and stability were all within the acceptable limits. The validated method has been applied to investigate the pharmacokinetics of iptacopan and its two acyl glucuronidation metabolites in monkey plasma. After intravenous administration, iptacopan showed low clearance (2.75 mL/min/kg) in monkey plasma. After oral administration, the bioavailability was 55.43%. The exposure (AUC0-t) of direct acyl glucuronide (AG) of iptacopan accounts for 9.73% of the iptacopan plasma exposure. The AUC0-t of AG of dealkylated metabolite of iptacopan was present at a lower level, accounting for 0.5% of the iptacopan plasma exposure.

A sensitive and robust analytical method for the determination of enramycin residues in swine tissues using UHPLC–MS/MS

Enramycin, a common growth promoter utilized in chickens and pigs, is sensitive against Gram-positive bacteria, and the maximum residue limit (MRL) of enramycin set up by is 30 μg/kg. However, the methods have been reported for detecting enramycin have failed to meet the accuracy requirements, with the required limit of quantification being higher than the MRL. To address this issue, we developed a high-sensitive and robust analytical method based on ultrahigh-performance liquid chromatography coupled with mass spectrometry (UHPLC–MS/MS), to determine enramycin residues in swine tissues, including liver, kidney, pork, and fat. The ENV cartridge was selected to cleanup and enrich analytes after being extracted using a mixture of 55% methanol containing 0.2 M hydrochloric acid. With comprehensively validation, this established method was found great linearity of enramycin in each tissue, with a coefficient of variation above 0.99. Satisfactory recoveries from four different spiking levels were acquired (70.99–101.40%) while the relative standard deviations were all below 9%. The limit of quantification of enramycin in the present study is 5 μg/kg in fat and 10 μg/kg in other tissues, meeting the requirements for conducting the corresponding safety evaluation study. This method was demonstrated with excellent specificity, stability, and high sensitivity. To conclude, this novel approach is sufficiently sensitive and robust for the safety evaluation of enramycin in food products.

2D/0D Heterojunction Fluorescent Probe with Schottky Barrier Based on Ti3C2TX MXene Loaded Graphene Quantum Dots for Detection of H2S During Food Spoilage

AbstractHydrogen sulfide (H2S) contamination of food has raised widespread public health concerns, leading to substantial medical and economic burdens. Herein, a 2D/0D heterojunction fluorescent probe (TCTG) with Schottky barriers (SB) is designed and synthesized, utilizing Ti3C2Tx MXene‐loaded graphene quantum dots (GQDs), for the detection of H2S during food spoilage. The microstructures observed through SEM and TEM reveal that uniformly sized GQDs are evenly attached to the surface of a monolayer Ti3C2Tx. The chemisorption between GQDs and Ti3C2Tx facilitates charge transfer and the formation of SB, resulting in intramolecular charge transfer (ICT) effects. With the introduction of H2S, TCTG(50%) exhibits the highest sensitivity, selectivity, and anti‐interference properties, with ultra‐fast fluorescence transient reaction (3s) and remarkably low detection limit of 41.82 ppb as well as noticeable color change. When TCTG(50%) reacted with H2S, the ICT effects are inhibited, leading to the recovery of photoinduced electron transfer (PET) and fluorescence quenching. Notably, probe TCTG is effectively utilized to detect changes in H2S levels in raw foods to assess their quality. Overall, the significance of this study is its potential to revolutionize food spoilage detection, offering a fast, reliable, and sensitive method to ensure food safety and reduce associated health and economic burdens.

Study of the Synchronization and Transmission of Intracellular Signaling Oscillations in Cells Using Bispectral Analysis

The oscillation synchronization analysis in biological systems will expand our knowledge about the response of living systems to changes in environmental conditions. This knowledge can be used in medicine (diagnosis, therapy, monitoring) and agriculture (increasing productivity, resistance to adverse effects). Currently, the search is underway for an informative, accurate and sensitive method for analyzing the synchronization of oscillatory processes in cell biology. It is especially pronounced in analyzing the concentration oscillations of intracellular signaling molecules in electrically nonexcitable cells. The bispectral analysis method could be applied to assess the characteristics of synchronized oscillations of intracellular mediators. We chose endothelial cells from mouse microvessels as model cells. Concentrations of well-studied calcium and nitric oxide (NO) were selected for study in control conditions and well-described stress: heating to 40 °C and hyperglycemia. The bispectral analysis allows us to accurately evaluate the proportion of synchronized cells, their synchronization degree, and the amplitude and frequency of synchronized calcium and NO oscillations. Heating to 40 °C increased cell synchronization for calcium but decreased for NO oscillations. Hyperglycemia abolished this effect. Heating to 40 °C changed the frequencies and increased the amplitudes of synchronized oscillations of calcium concentration and the NO synthesis rate. The first part of this paper describes the principles of the bispectral analysis method and equations and modifications of the method we propose. In the second part of this paper, specific examples of the application of bispectral analysis to assess the synchronization of living cells in vitro are presented. The discussion compares the capabilities of bispectral analysis with other analytical methods in this field.

Simultaneous Determination of Tobacco Smoke Exposure and Stress Biomarkers in Saliva Using In-Tube SPME and LC-MS/MS for the Analysis of the Association between Passive Smoking and Stress

Passive smoking from environmental tobacco smoke not only increases the risk of lung cancer and cardiovascular disease but may also be a stressor triggering neuropsychiatric and other disorders. To prevent these diseases, understanding the relationship between passive smoking and stress is vital. In this study, we developed a simple and sensitive method to simultaneously measure nicotine (Nic) and cotinine (Cot) as tobacco smoke exposure biomarkers, and cortisol (CRT), serotonin (5-HT), melatonin (MEL), dopamine (DA), and oxytocin (OXT) as stress-related biomarkers. These were extracted and concentrated from saliva by in-tube solid-phase microextraction (IT-SPME) using a Supel-Q PLOT capillary as the extraction device, then separated and detected within 6 min by liquid chromatography–tandem mass spectrometry (LC−MS/MS) using a Kinetex Biphenyl column (Phenomenex Inc., Torrance, CA, USA). Limits of detection (S/N = 3) for Nic, Cot, CRT, 5-HT, MEL, DA, and OXT were 0.22, 0.12, 0.78, 0.39, 0.45, 1.4, and 3.7 pg mL−1, respectively, with linearity of calibration curves in the range of 0.01–25 ng mL−1 using stable isotope-labeled internal standards. Intra- and inter-day reproducibilities were under 7.9% and 14.6% (n = 5) relative standard deviations, and compound recoveries in spiked saliva samples ranged from 82.1 to 106.6%. In thirty nonsmokers, Nic contents positively correlated with CRT contents (R2 = 0.5264, n = 30), while no significant correlation was found with other biomarkers. The standard deviation of intervals between normal beats as the standard measure of heart rate variability analysis negatively correlated with CRT contents (R2 = 0.5041, n = 30). After passive smoke exposure, Nic levels transiently increased, Cot and CRT levels rose over time, and 5-HT, DA, and OXT levels decreased. These results indicate tobacco smoke exposure acts as a stressor in nonsmokers.

Detection and health implications of phthalates in tea beverages in market: Application of novel solid-phase microextraction fibers

Assessment and control of emerging organic pollutants in food have become critical for global food safety and health. The European Union has set standards for certain emerging organic pollutants, such as phthalic acid esters (PAEs) in food. Because of being endocrine disruptors, PAEs are toxic and carcinogenic to humans. Release of PAEs from packaging materials poses a potential risk to human health and causes environmental pollution. In this study, a highly sensitive analytical method for the detection of PAE contents in tea beverages was established using hydroxyl-functionalized covalent organic frameworks (COFs) as solid-phase microextraction (SPME) coating. Results indicate that functionalization with hydroxyl groups enhances the adsorption of PAEs. The proposed method exhibits a wide linear range (1–20,000 ng L−1), low limits of detection (> 0.048 ng L−1), and satisfactory recovery (72.8 %–127.3 %). To investigate the PAE contamination in beverages, contamination levels of six typical PAEs and their health impacts were surveyed across various brands/types/packaging materials of tea beverages sold in China. Results of the hazard quotient and hazard index approaches suggest no or extremely low health concerns regarding PAE levels. We observe that hydroxyl groups functionalized on COFs enhance the adsorption of PAEs. Moreover, an important outcome of this study is development of an efficient and sensitive direct detection method for PAEs in complex tea matrices, providing a reliable approach for the assessment of PAEs in other complex matrices.

Zirconium-based metal-organic framework composites for solid phase extraction of brevetoxin-A from seawater

Red tides are a type of natural marine disaster caused by harmful algae characterized by a high toxicity, wide distribution, and long duration. Since the concentration of algal toxins in seawater increases with the occurrence of red tides, algal toxins detected in seawater could be used to predict the occurrence and evolution of red tides. Brevetoxin-A (BTX-A) is a secondary metabolite produced by the harmful algae Karenia brevis, whose detection in seawater could form the basis of an accurate warning system for incoming red tides. However, due to the inherent complexity of the seawater matrix and the extremely low levels of BTX-A in seawater, the use of instruments for its direct detection is difficult. Therefore, there is an urgent need to develop a sample pretreatment method for the efficient enrichment of BTX-A in seawater. In this study, a metal-organic backbone material (UiO-66) and its composite with silica microspheres (SiO2@UiO-66) were successfully synthesized using the solvothermal method. The prepared SiO2@UiO-66 exhibited good hydrophilicity, water stability, and large specific surface area. Furthermore, it also exhibited hydrogen bonding and electrostatic interactions with BTX-A, had a strong affinity for BTX-A, and was able to efficiently adsorb BTX-A in complex matrices. Therefore, SiO2@UiO-66 showed potential as a novel packing material for the extraction of BTX-A from solid phase extraction columns. Combined with high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS), a highly sensitive detection method for the determination of BTX-A in marine water was established. The established analytical method had a low detection limit (3.0 pg/mL), a wide linear range (10.0 -200.0 pg/mL), and a good linear relationship (R=0.9992). Combined with the Fujian Province Red Tide Monitoring and Early Warning Information 2021 issued by the Fujian Provincial Oceanic and Fisheries Bureau, the analytical method established herein was successfully applied to analyze and monitor the content of BTX-A in actual seawater samples. This highlights the proposed system's potential for use as an early warning factor in the monitoring of red tides, representing a simple and fast pretreatment methodology for the detection of BTX-A in seawater.

A "signal-off" anodic photoelectrochemical sensor based on ZnIn2S4/TiO2 heterojunction for dopamine detection

Dopamine (DA) is an important neurotransmitter. Abnormal levels of it in human body can increase the risk of many neurological diseases. Thus, developing a simple, sensitive detection method of DA is crucial. In this paper, we reported a "signal-off" anodic PEC sensor based on fluorine-doped tin oxide (FTO) glass modified ZnIn2S4/TiO2 heterojunction (ZnIn2S4/TiO2/FTO) for DA detection. The experimental results show that the ZnIn2S4/TiO2/FTO electrode prepared by two-step hydrothermal method has a good photocurrent response performance under visible light. After incubation with DA, the photocurrent response decreases significantly because DA can rapidly oxidizes to polydopamine (PDA) through the action of superoxide radical (·O2-) and hydroxyl radical (·OH) intermediate species, which are intermediates produced by the ZnIn2S4/TiO2/FTO electrode under visible light irradiation. The constructed PEC sensor has a good linear relationship in the concentration range from 0.5 to 1000.0 μM, and its detection limit is 0.253 μM. In addition, the results of the proposed PEC sensor in real serum samples are satisfactory. The PEC sensor provides a promising platform for DA detection, laying the foundation for future advances in disease diagnosis and prevention.

Genetic evolutionary analysis of a strain of Senecavirus A in Anhui and the establishment of its detection method

BackgroundSenecavirus A (SVA) is the only member of the genus Senecavirus in the family Picornaviridae, and is one of the pathogens of porcine blistering disease. SVA has been reported in the United States, Canada, China, Thailand, and Colombia. MethodsIn this study, positive SVA infection was detected by RT-PCR in sick materials collected from pig farms of different sizes in Anhui Province. ResultsIn this study, a virulent strain of SVA was successfully obtained by viral isolation on BHK21 cells and named SVA-CH-AHAU-1. Meanwhile, a simple, rapid and accurate nano-PCR method for the detection of SVA infection was established in this study, using the recombinant plasmid pClone-SVA-3D as a template. ConclusionsThe complete genome of SVA-CH-AHAU-1 is 7286 bp, including a 5′ non-coding region (UTR), an open reading frame (ORF) of 6546 nucleotides, encoding 2182 amino acids (aa), and a 3’ UTR with Poly(A) features, and phylogenetic analysis showed that this isolate had the highest nucleotide homology (97.9 %) with the US isolate US-15-41901SD. In this study, the virulent strain SVA-CH-AHAU-1 was found to recombine in the ORF region with isolates SVA-CH-SDGT-2017 and SVA/Canada/ON/FMA-2015-0024 T2/2015. The complete genome has been submitted to GeneBank with the accession number OM654411. In addition, our results suggest that the established nano-PCR assay can be used as an economical, reliable and sensitive method for the field diagnosis of SVA method, especially in resource-limited areas.

The importance of determining the concentration of myco- phenolic acid in children with steroid-dependent nephrotic syndrome

Despite the significant effectiveness of steroid therapy, specialists face serious difficulties in managing patients with steroid-dependent nephrotic syndrome, which requires the prescription of immunosuppressive therapy. Among the available adjuvant drugs, mycophenolate mofetil stands out as a preferred option due to its safety profile, good tolerability, and high efficacy. Currently, only a few studies are presented in the literature on the effectiveness of the use of mycophenolic acid in children with nephrotic syndrome, which is explained by the complexity of the pharmacokinetics of the drug. The article presents the results of a study whose purpose was to evaluate the value of determining mycophenolic acid in patients with steroid-dependent nephrotic syndrome to maintain stable clinical and laboratory remission of the disease. The study examined 78 patients aged 1 to 18 years with steroid-dependent nephrotic syndrome. The results demonstrated that determination of mycophenolic acid concentrations in children with steroid-dependent nephrotic syndrome is an effective method for monitoring therapy for nephrotic syndrome in aunts. Determining the concentration of mycophenolic acid at the C0 point is a highly specific and sensitive method for predicting the development of relapses of the disease, while a mycophenolic acid concentration level of more than 3.2 μg/ml can serve as a minimum guideline for monitoring the use of mycophenolic acid in children with steroid-dependent nephrotic syndrome.

A self-driven microfluidic immunosensor for rapid separation, enrichment, and detection of biomarkers in serum.

Biomarkers and their concentration levels are critical indicators of metabolomics for clinical applications. Rapid and sensitive analysis methods are essential for realizing timely and efficient quantitation of those significant biomarkers. In this work, a self-driven microfluidic immunosensor was developed for rapid all-in-one separation, enrichment, and detection of biomarkers. This immunosensor was constructed from a cyclic olefin copolymer (COC) channel layer and a polydimethylsiloxane (PDMS) sensing layer. The COC channel layer was modified through protein adsorption, immobilization, and remaining active site blocking. The obtained hydrophilic microchannels not only reduce the nonspecific adsorption, but also provide stable capillary-driven flow generation with linear velocities up to 20mm/s for aqueous solution auto-injection. The PDMS sensing layer was modified using capture antibodies to accomplish affinity recognition of target biomarkers. Procalcitonin (PCT) and serum amyloid A (SAA) were selected as model biomarkers in the feasibility study on applying the self-driven microfluidic immunosensor to bioassay. The limits of detection of PCT and SAA were 7.9ng/L and 7.6μg/L, respectively. Moreover, the whole process can be accomplished within 60min with excellent selectivity and reproducibility. In clinical serum sample analysis, satisfactory recoveries were achieved for PCT and SAA in the ranges of 85.0-103.0% and 95.5-106.0%, respectively, with relative standard deviations less than 5.3%. The method accuracy was further confirmed by the results of commercial immunoassay kits. This simple and easily operated immunosensor provides a rapid and sensitive biomarker analysis tool, and promotes the further development of automated and easy-to-use microfluidic immunoassays.

HPTLC – densitometric method of analysis for estimation of efonidipine hydrochloride and telmisartan used in treatment of hypertension

Precise, accurate, and sensitive high performance thin layer chromatographic method has been developed and validated for simultaneous determination of efonidipine hydrochloride (EFD) and telmisartan (TEL) in a tablet dosage form which is used in treatment of hypertension. The proposed method used aluminum plates pre-coated with silica gel 60 F254 as a stationary phase and n-butanol: toluene: acetic acid (3:7:0.2, v/v/v) as a mobile phase for separation purpose. Compact spots of TEL and EFD were obtained at Rf 0.73 and 0.37, respectively. Densitometric detection was carried out at 299 nm in UV. Linear responses were shown by the detector over the range of 200–1200 ng/band for both drugs. The proposed method was validated as per ICH guidelines. The method was successfully applied for estimation of both drugs in tablet formulations. Forced degradation study was carried and efonidipine was found to be susceptible to base hydrolysis and oxidative stress degradation while TEL was stable in acid-base hydrolysis, oxidative stress, dry heat and photo stability testing conditions. The developed method can be used for the analysis of stability samples. Greenness assessment of developed method was performed using AGREE software. The method was found to be greener having greenness score of 0.69.

Rapid and simultaneous determination of ultrashort-, short- and long- chain perfluoroalkyl substances by a novel liquid chromatography mass spectrometry method

Per- and Polyfluoroalkyl Substances (PFAS) are a group of persistent organic pollutants that have received considerable attention from public and regulatory groups. Due to regulations of long-chain PFAS, the use of short-chain and ultrashort-chain PFAS is rapidly growing. Thus, there is an urgent need to develop quantitative methods for determining PFAS with different chain lengths in various environmental matrices. This study introduces an innovative liquid chromatography-mass spectrometry (LC-MS) system combining large volume injection (LVI) and online solid phase extraction (SPE). This system incorporates three columns: a reverse-phase (RP) column, a weak anion exchange (WAX) trap column, and a hybrid HILIC/ion-exchange (HILIC/IE) column, controlled by two valves. With valve switching, ultrashort-chain PFAS that are not retained by the RP column are enriched by the trap column, while other PFAS are separated by the RP column. The trapped ultrashort PFAS are then transferred to the HILIC/IE column for further separation. The LVI significantly enhances the method's sensitivity, allowing for rapid and simultaneous determination of ultrashort-, short- and long- chain PFAS in aqueous samples. The matrix effects from various environmental samples were evaluated, and the results indicate that this unique LC-MS method is suitable for analyzing all chain-length PFAS in various matrices, including surface water, sewage effluent, and seawater. Finally, this novel LC-MS method was applied to quantify PFAS in various water samples.

A novel QbD HPLC approach for the concurrent analysis of amoxicillin, amprolium, and ethopabate in food samples and dosage forms

A simple, rapid, and sensitive reversed-phase HPLC method coupled with photodiode array detection (PDA) was developed for the simultaneous estimation of three co-administered antibacterial drugs, amprolium, ethopabate, and amoxicillin for the first time. An analytical quality-by-design (QbD) approach was adopted to optimize the developed approach. A two-level full factorial design (25 FFD) was implemented to optimize how variable factors affected chromatographic responses. On a Hypersil BDS C18 column, the chromatographic separation was carried out using isocratic elution at 40.1 °C. The mobile phase consisted of methanol: potassium phosphate buffer containing 5.76 mM heptane sulphonate (9.05 mM; pH 3.32, 44.8:55.2 % v/v) pumped at a flow rate of 0.8 mL/min and an injection volume of 20.0 μL. In less than 7 minutes, the suggested method was able to separate the ternary mixture with a challenging ratio of 1: 15.6: 1 for amoxicillin, amprolium, and ethopabate, respectively. The calibration curves showed excellent linearity over the concentration ranges of 0.50-30.0, 0.30-30.0, and 0.05-20.0 μg/mL, respectively. Recovery percentages ranged from 90.30 to 111.13 % with %RSD less than 2% were obtained upon spiking to chicken tissues, liver, and egg samples, indicating the suitability of the proposed method for determining of the studied drugs in complicated food matrices. Furthermore, the proposed method was successfully applied to determine the studied drugs in their commercial formulations. Full validation was carried out according to ICHQ2 (R1) guidelines. Finally, the method's greenness and ecofriendliness were evaluated using various metrics tools, including the Green Analytical Procedure Index (GAPI) and Analytical GREEnness (AGREE).

Development and Validation of RP-HPLC Method for Estimation of Ticagrelor in Pharmaceutical Dosage Form and Force degradation study

Ticagrelor is a selective Adenosine diphosphate (ADP)-receptor antagonist which is prescribed in the form of tablets and acts as an oral antiplatelet for the prevention of further thrombotic events in patients with Acute coronary syndrome (ACS) and those undergoing Percutaneous coronary intervention (PCI). Therefore, accurate and reliable determination of Ticagrelor in bulk and in dosage forms is vital for clinical consideration. The objective of the method is to develop a new, simple, sensitive, accurate, and economical analytical method for the determination of assay of Ticagrelor Tablet and to perform a forced degradation study of Ticagrelor Tablet by RP-HPLC. Chromatographic separation was achieved on an UHPLC equipped with reverse phase C-18 (250 × 4.6mm, 5µ) with a mobile phase composed of acetonitrile and buffer solution (75:25) at a flow rate of 1.10 ml/min. The effluents were detected at a wavelength of 256 nm. The retention time of Ticagrelor was found to be at 3.579 min. The correlation coefficient for Ticagrelor was found to be 0.997. Recovery of Ticagrelor in the formulation was found to be 98%-102%. LOD and LOQ values of Ticagrelor were found to be 1.31 and 3.98, respectively. The method was proven to be precise (%RSD=2%), accurate (>90%), and specific for the measurement of Ticagrelor in tablets. Thus, being simple, accurate, precise, and rapid, the newly developed RP-HPLC method is recommended for the estimation of Ticagrelor in nature. Due to its high sensitivity and specificity, it is a suitable choice for identifying Ticagrelor in drugs and other products and differentiating hid similarities. However, this method can effectively be used for routine assay and stability study, which can help manage cardiovascular diseases since the quality and efficacy of Ticagrelor-containing products are significant to patients’ condition.

Comparison of commercial DNA kits for allergen detection of celery in food matrices

For correct allergen risk management by industry, retail and food safety authorities, sensitive and reliable fast allergen detection methods are required, even more when precautionary allergen labelling based on reference doses will be implemented in legislation.This study aimed to perform a comparative assessment of three commercially available quantitative or qualitative test kits, for DNA analysis of celery in food products. Five product groups, representing different sectors of the AOAC food-matrix triangle, being (plant-based) meat products, snacks, sauces, dried herbs and spices, and smoothies, were identified to potentially contain celery. From each group, blank and incurred (labelled to contain celery) food products were selected, of which the blank food products were additionally spiked with low protein levels of celery prior to qPCR assessment.Results show that the assessed test kits perform according to their specifications, however, a clear influence of the matrix on the detection ability of celery was observed. In addition, quantification of the amount of celery in the different food products showed to be challenging in all food product groups using the two quantification kits.

Novel simultaneous analysis of 18 types of glycosaminoglycan-derived disaccharides using 4-aminobenzoic acid ethyl ester derivatization by HPLC with fluorescence detection.

Glycosaminoglycans (GAGs), including hyaluronic acid (HA), chondroitin sulfate (CS)/dermatan sulfate (DS), heparan sulfate (HS)/heparin (HP), and keratan sulfate (KS), play pivotal roles in living organisms. Generally, GAGs are analyzed after enzymatic digestion into unsaturated or saturated disaccharides. Due to high structural similarity between disaccharides, however, separation during analysis is challenging. Additionally, little is known about the structures of GAGs and their functional relationships. Elucidating the function of GAGs requires highly sensitive quantitative analytical methods. We developed a method for the simultaneous analysis of 18 types of disaccharides derived from HA (1 type), CS/DS (7 types), HS/HP (8 types), and KS (2 types) potentially detectable in analyses of human urine. The simple method involves HPLC separation with fluorescence detection following derivatization of GAG-derived disaccharides using 4-aminobenzoic acid ethyl ester (ABEE) as a pre-labeling agent and 2-picoline borane as a reductant. The ABEE derivatization reaction can be performed under aqueous conditions, and excess derivatization reagents can be easily, rapidly, and safely removed. This method enables highly sensitive simultaneous analysis of the 18 abovementioned types of GAG-derived disaccharides using HPLC with fluorescence detection in small amounts of urine (1mL) in a single run. The versatile method described here could be applied to the analysis of GAGs in other biological samples.