Intra-day Precision Research Articles

Simultaneous quantification of 14 glucosinolates in rapeseeds by ultra high performance liquid chromatography – Tandem mass spectrometry

Glucosinolates' species and content serve as crucial indicators for Brassicaceae plants and their products. However, endogenous enzymes in Brassicaceae can hydrolyze glucosinolates, influencing their quantitative detection. Our study utilized rapeseed, a vital Brassicaceae oil crop, to establish a novel UHPLC-MS/MS method for glucosinolate determination, incorporating a pretreatment approach with enzyme inhibition. This enabled intact glucosinolate determination at room temperature. The straightforward process involved homogenizing samples with saturated sodium chloride and methanol, followed by vortex extraction, centrifugation, and filtration. The method exhibited a strong linear relationship (R2 > 0.990), with LODs and LOQs ranging from 0.1 to 1.1 mg/kg and 0.3–3.7 mg/kg, respectively. Intra-day precision (7 repetitions) ranged from 3.21 to 10.47 %, while inter-day precision (3 days) ranged from 3.13 to 15.97 %.

Application of deep eutectic solvent based dispersive liquid–liquid microextraction method followed by HPLC-DAD for quantification of selected neonicotinoid pesticides in vegetable oils

The use of deep eutectic solvents (DES) as alternative green solvents in various analytical chemistry fields has garnered attention of researchers globally. Therefore, this study presents environmentally friendly DES for extraction of selected neonicotinoids (acetamiprid, imidacloprid, thiacloprid and thiamethoxam) prior to their determination using high-performance liquid chromatography- diode array detector (HPLC-DAD). The DES mixture that resulted in the highest extraction recoveries of neonicotinoids (NEOs) was choline chloride with ethylene glycol (CCl:EG) and nuclear magnetic resonance and Fourier transform infrared confirmed the successful synthesized CCl:EG. Thereafter, the CCl:EG was applied in dispersive liquid–liquid microextraction (DLLME) for the preconcentration and extraction of acetamiprid, imidacloprid, thiacloprid and thiamethoxam. The optimum factors affecting extraction efficiency of the DES-DLLME procedure were 0.8 mL (DES volume), 4.5 min (extraction time), 0.5 mL (eluent volume), 7 pH and 75 s (eluent time). The optimized DES-DLLME achieved limit of detection (LOD) and quantification (LOD) range of 0.4–4.95 ng.µL−1 and 1.43–9.70 ng.µL−1 respectively. Additionally, the extraction method achieved good accuracy (79.0–119.6 %), preconcentration factors (39.3–118), interday (0.61–1.62 %) and intraday (0.1–0.98 %) precisions. The greenness assessment of the proposed DES-DLLME method using AGREE, NEMI and AES tools revealed that the method was green. Finally, the developed DES-DLLME method was applied to real vegetable oils and the concentrations were below detection limits.

Development & validation of a simple, sensitive, robust, reproducible & economical spectrofluorimetric method for estimation of taurine in novel chewable gel

BackgroundTaurine, known as 2-aminoethanesulphonic acid, is a sulfur-containing β-amino acid not incorporated into proteins. However, it takes part in biochemical reactions of significant importance, such as conjugation with bile acids to form bile salts essential for fat absorption, cell membrane stabilization, antioxidation, detoxification, osmoregulation, neuromodulation, and brain and retinal development. ObjectiveTo increase its dissolution and absorption rate for early onset of action, we developed a medicated gel formulation of taurine. Chewable gels are suitable dosage forms for pediatric, geriatric, and dysphagic patients. The current research aims to develop a simple, sensitive, and reproducible spectrofluorometric method for determining taurine in chewable gel. MethodA spectrofluorimetric method was developed, and the determination of taurine was based on Hantzsch condensation synthesis, and the fluorescence intensity was measured at emission wavelength (λem) of 474 nm after excitation (λex) at 416 nm. The spectrofluorimetry method for taurine was validated according to the Q2B(R1) ICH guideline for method validation and validated for linearity, LOD, LOQ, interday and intraday precision, accuracy, and robustness. ResultsThe calibration curve was linear in a 1.5–50 µg/ml concentration range. The correlation coefficient (r2) was found to be 0.994. The % RSD values of intra-day precision lie between 0.11 and 0.09; for inter-day precision, it was found to be between 0.09 and 0.11; in both cases, it is less than 2 %. The percentage accuracy study was between 91.81 and 99.93 with a % RSD value of 0.078, less than 2 %. LOD & LOQ values have been mathematically calculated to 4.03 µg/ml and 12.23 µg/ml, respectively. The method was precise, accurate, and robust. ConclusionAll validation parameters were found to be within their acceptable limits. Thus, the developed spectrofluorimetric method of taurine is economical, fast, and robust. Further, this method was utilized to estimate taurine content uniformity in chewable gel for in-vitro release studies.

Qualitative and Quantitative Analysis Method of Recombinant Collagen in Complex Matrix Based on HPLC-MS/MS

The purpose of this study is to achieve the quantitative detection of recombinant type III collagen (rh-COL-III) in dressings with complex matrix. First of all, the marker peptide (GEAGIPGVPGAK) of rhCOL-III was identified with HPLC-MS/MS. Then, a qualitative and quantitative method based on marker peptides was established and validated. In order to obtain higher sensitivity, a pretreatment method of liquid, gel, and ointment dressings was optimized. The reference material for quantification was combined using rhCOL-III and blank matrix of each dressing. The results indicated that the relative standard deviation (RSD) of the quantitative method was 2.77%, and the RSD of intraday and interday precision was 2.76% and 2.31%, respectively. The spiking recovery rate was between 80% and 90%. The optimal pretreatment method was Tris-HCl solvent replacement. The optimal trypsin concentration for the dressing solution was 20 μg in 500 μL. The method of preparing standard substances with a blank matrix can effectively eliminate the influence of the matrix effect on the quantitative results. The average spiking recovery rates of 50 μg/mL, 100 μg/mL, and 200 μg/mL in three different dressings ranged from 80% to 120%. The quantitative detection of limit (LOD) of rhCOL-III was 1 ng/mL, 2 ng/g, and 1 ng/g in liquid, ointment, and gel dressings.

Rapid determination of caine-based anesthetics and their metabolite residues in fish using a modified QuEChERS method coupled with UPLC-MS/MS

Caine-based anesthetics are frequently used in the production and transportation of aquatic products, but residues in fish threaten human health. A rapid, sensitive, and effective method was developed for detecting caine-based anesthetics and their metabolite residues in fish using ultra-high performance liquid chromatography-tandem mass spectrometry coupled with a modified QuEChERS (quick, easy, cheap, effective, rugged, and safe) method. The sample was extracted with 0.05 % (v/v) formic acid-80 % acetonitrile, 4 g NaCl was added for liquid partitioning, and 50 mg C18 was used for purification. Matrix-matched calibrations showed good correlation coefficients with R2 ≥ 0.9942. The limits of detection were 0.5–4.4 ng/g. The recoveries ranged from 71.4 %–115.8 %, and the relative standard deviation for intra-day and inter-day precision was less than 8.5 % and 9.2 %, respectively. This method effectively analyzed the residues of caine-based anesthetics and their metabolites in fish and could be applied to aquatic product anesthetic regulation.

Concurrent estimation of Pregabalin and Etoricoxib by new stability indicating RP-UPLC approach: application in assay of commercial tablets

The present research aims to establish new stability indicating RP-UPLC method for concurrently estimating Pregabalin and Etoricoxib in blended powder and their combined tablet formulation. The effective separation of Pregabalin and Etoricoxib achieved with HSS column C18 (150 × 2.1 mm, 1.8 µm), 0.1% orthophosphoric acid: acetonitrile in 65:35 v/v) at a flow rate of 0.3 mL/min, and isocratic elution at 228 nm. The elution of PRB and ETB was noticed at 1.56 and 2.01 min, with good resolution and system suitability with the developed approach. PRB and ETB have shown linear responses from 18.75 to 112.5 µg/mL and 15 to 90 µg/mL, respectively. The range of the % RSD for intraday and inter-day precision was 0.33 to 0.81. The LOD and LOQ of Pregabalin and Etoricoxib were computed to be 0.07 µg/mL, 0.21 µg/mL, and 0.01 µg/mL, 0.04 µg/mL, respectively, by standard deviation method. The validation method was carried out using ICH standards. The stability- indicating the feature of the method was confirmed by the forced degradation studies where degradants generated by stress testing were clearly distinguished from the peaks of analytes. The shorter elution period and superior sensitivity of both analytes with this method are appropriate for regular analysis of Pregabalin and Etoricoxib.Graphical

Probe electrospray ionization coupled to a quadrupole time-Of-flight: A feasibility study for the detection of cocaine and its derivatives in oral fluid

Probe Electrospray Ionization (PESI) is an atmospheric pressure ionization method that can be directly coupled with a mass spectrometer to allow ultrafast analyses without chromatographic separation and with minimal sample preparation. Using the particular case of cocaine and its metabolites in human oral fluid, the main objective of the present study was to test the feasibility of a new hybrid system combining a PESI source and a quadrupole time-of-flight (QTOF). The best results were obtained for a sample preparation with a simple dilution of 100 µL of oral fluids in an ethanol / 10 mM ammonium formate buffer (50/50) and 10 µL deposited on a dedicated sample plate and introduced into the PESI source. For HRMS acquisition, an approach consisting in a full-mass scan (“untargeted approach” from 100 to 500 m/z; MS1) followed by a targeted scheduled MSMS acquisition (precursor ions of the 3 molecules of interest and their 3 internal standards; MS2) gave the best signals. The total time of analysis was 0.45 min and the method was validated according to ISO15189 standard for a 5–100 ng/mL range, including accuracy and precision (inter-day and intra-day precision and bias values were lower than 15 %), matrix effect, carryover and specificity (no interference with a mixture of 119 psychotropic drugs spiked at 1 mg/L). The LLOD values were 1 ng/mL for the 3 cocaine derivatives and 83 Driving Under the Influence of Drug (DUID) cases sent to our Lab for the determination of illicit drugs in oral fluid were analyzed using the PESI-QTOF method and compared to a LC-MS/MS method. A perfect agreement was observed between the 2 methods, whether the cases were positive or negative. Cocaine was detected in 51 out of these 83 real cases (61.6 %). BZE and EME were also simultaneously detected in 50 of them (98.5 %). This feasibility study reports the first analytical method based on a coupling of a PESI source to a QTOF mass spectrometer. Adding the major advantage of high specificity through HRMS is a step forward for PESI technology.

Plant-Derived and Synthetic Nicotine in E-Cigarettes: Is Differentiation with NMR Spectroscopy Possible?

To circumvent regulatory frameworks, many producers start to substitute plant-derived nicotine (tobacco-derived nicotine, TDN) by synthetic nicotine (tobacco-free nicotine, TFN) in e-cigarette products. Due to the higher costs of enantiomeric synthesis and purification of TFN, there is a need to develop an analytical method that clearly distinguishes between the two sources. To trace nicotine's origin, its enantiomeric purity can be postulated by 1H NMR spectroscopy using (R)-(-)-1,1'-binaphthyl-2,2'-diyl hydrogen phosphate (BNPPA) as a chiral complexing agent. Low-field (LF) NMR conditions were optimized for this purpose even using a small amount of e-liquid sample (limit of quantification 8 mg/mL nicotine). All investigated products were found to contain one isomer (most likely (S)-(-)-nicotine). A direct 13C NMR method at natural abundance has been validated to differentiate (S)-TDN and (S)-TFN in e-cigarettes produced using nicotine of different origin. The method is based on calculation of the relative 13C content of 10 C-positions of the nicotine molecule with intraday and interday precisions below <0.2%. The method was applied to 12 commercial e-cigarette products labeled as containing TDN and TFN. Principal component analysis (PCA) was applied to the relative peak areas to visualize the difference between studied products. The LF 1H NMR method is a good alternative to expensive high-field NMR to differentiate between a racemate mixture and single optical isomers, whereas only high-precision 13C NMR can be used to distinguish (S)-TDN and (S)-TFN in e-cigarettes after appropriate sample extraction.

Multivariate Optimization of a Green Microwave-Assisted Hydrogen Peroxide Digestion of Vegetable Oils and Subsequent Elemental Determination Via Inductively Coupled Plasma–Optical Emission Spectrometry

AbstractA microwave-assisted digestion technique based on dilute hydrogen peroxide (MW-AHPD) was developed for multielemental determination in vegetable oils. The determination of ten trace elements (As, Cd, Cr, Cu, Fe, Pb, Ni, Sn, V, and Zn) was conducted via inductively coupled plasma optical emission spectroscopy (ICP-OES) after digestion. The most influential parameters were investigated by using multivariate optimization tools (two-level full factorial and central composite design) with percent recovery as the chemometric response. The optimum conditions were 2.0 mol L−1 (H2O2]), 156 °C (digestion temperature), 0.1 g (sample mass) and 50 min (digestion time). Under the optimized conditions, the efficiency of digestion was evaluated based on the residual carbon content (RCC) of the final digests. The RCC values were very low, ranging from 0.84 to 1.60% (m/m). The greenness of the technique was evaluated using the Analytical Eco-scale, and the proposed method was considered an excellent green analysis method with a final score of 90. The accuracy of the optimized MW-AHPD was evaluated by spiking sunflower, olive, and peanut oils at concentrations of 2.5 and 5.0 μg L−1, and excellent recoveries between 90.3 and 107.3% were reported. The accuracy of the MW-AHPD method was compared with that of microwave-assisted digestion using concentrated HNO3, and there was no significant difference between the two methods. The limits of detection ranged from 0.026 to 14.6 μg L−1. On the other hand, the interday and intraday precisions were less than 6.67 and 6.96%, respectively. The method was successfully applied to determine the concentrations of trace elements in 5 vegetable oils on the South African market. Thus, MW-AHPD-ICP–OES is applicable for the determination of trace elements in vegetable oils.

Hydrophobic deep eutectic solvent-ferrofluid microextraction followed by liquid chromatography-mass spectrometry for the enantioselective determination of chiral agrochemicals in natural waters.

The increasing use of chiral agrochemicals sold as racemic formulations raises concern for the negative impacts that inactive enantiomers can have on aquatic life and human health. The present work just focuses on the determination of ten chiral pesticides in river water samples by applying a ferrofluid-based microextraction followed by their stereoselective liquid chromatography analysis. To develop the ferrofluid, magnetite nanoparticles were prepared and coated with oleic acid and then dispersed in a hydrophobic natural deep eutectic solvent (NaDES), composed ofL-menthol and thymol (1:1). The stable colloidal dispersion was characterised by scanning electron microscopy, thermogravimetric analysis, energy-dispersive X-ray spectroscopy and attenuated total reflection Fourier transform infrared spectroscopy. The analyte microextraction from 5ml river samples was performed using 50µl of ferrofluid, while acidified acetonitrile (150µl) was used to break down the ferrofluid and solubilise the NaDES containing the analytes. All the extracts were analysed by high-performance liquid chromatography-tandem mass spectrometry. For each analyte, the baseline separation of isomers was achieved on a Lux i-Amylose-3 column (amylose tris(3-chloro-5-methylphenylcarbamate) working in reversed-phase mode; the combination with mass spectrometry detection allows the overall separation of 24 isomers (ten chiral analytes among which eight containing a single (one) chiral centre, one with two chiral centres and the last one existing in four stereoisomeric forms, due to the presence of two regioisomers with a chiral carbon) within 37min. The method showed very good figures of merit in terms of recoveries (77.7-97.5%), intra-day and inter-day precision (2.7-7.7% and 6.9-14.9%, respectively), limit of detection (0.01-0.35µg/L), limit of quantitation (0.03-1.20µg/L), linear dynamic range, and intra-day and inter-day accuracy (1.2-14.8% and 1.8-15.0%, respectively). The presented method was able to detect 14 out of 24 isomers at the preventive limit established by the Italian legislation for single pesticide (that for a chiral pesticide is the sum of all its isomers) in surface waters, set at 0.1µg/l. Finally, the method was evaluated using AGREEprep and ComplexGAPI metrics, compared with other ferrofluid-based methods, and applied to the analysis of water samples from two Italian rivers (the Nera River and the Tiber River), providing to be sustainable and reliable for the application to real river matrices.

Development of a 3D printed chemiluminescence smartphone detector for high performance liquid chromatography

A novel chemiluminescence detector for high-performance liquid chromatography (HPLC) was designed that incorporates a smartphone as photodetector, signal processor, and data storage unit. All detector components, including flow cell, flow cell holder, mixers, and housing, were fabricated by 3D printing and integration of the smartphone and 3D printed components provides a portable, low-cost, and user-friendly device. The detector was applied in the determination of carbamazepine using HPLC through a chemiluminescence reaction with tris(2,2′-bipyridyl)ruthenium(II) and cerium sulfate. Design and fabrication of the flow cell using two fabrication methods—3D printing and laser cutting—along with the integration of a mixer and the optimization of smartphone parameters were investigated. To expand the applicability of the detector for other analytes and different chemiluminescence reactions, detection of three piperazine derivatives by reaction with tris(2,2′-bipyridyl)ruthenium(II) and four phenethylamine compounds via reaction with acidic potassium permanganate reagent was studied. The linear dynamic range, limit of detection, and limit of quantification of the detector for the determination of carbamazepine were measured as 3.0–30.0 mg/L, 1.0 mg/L, and 3.0 mg/L, respectively, using the Samsung S20 smartphone device. Intraday and interday precision was evaluated for carbamazepine, with relative standard deviation (RSD) values for intraday precision ranging from 1.7 % to 6.2 % (n = 3) and interday precision RSD values ranging from 3.2 % to 4.8 % (n = 9).

Absolute Quantification of Phenylbutanoids in Zingiber cassumunar Roxb. Rhizome by Quantitative 1H NMR.

Quantitative determination of pharmacologically active constituents in medicinal plants is critical for quality control. Due to the chemical complexity of the crude plant extracts, the presence of interfering compounds is often problematic for the unambiguous quantitation of the designed bioactive compounds. Considering the method of quantification, quantitative NMR spectroscopy (qNMR) has gained substantial popularity as a powerful and effective technique for both qualitative and quantitative analyses of natural products. The aim of this study is to develop a quantitative NMR method for quantifying the bioactive phenylbutanoids in Zingiber cassumunar rhizome crude extract. Quantitative 1H NMR (qHNMR) measurements were performed on a 600 MHz NMR spectrometer using an internal standard for the determination of the absolute quantities of four phenylbutanoids in Z. cassumunar rhizome crude extract. The direct quantification of four characteristic phenylbutanoids, i.e., (E)-1-(3',4'-dimethoxyphenyl)butadiene (DMPBD), (E)-1-(2',4',5'-trimethoxyphenyl)butadiene (TMPBD), (E)-4-(3',4'-dimethoxyphenyl)but-3-en-1-ol, and (E)-4-(3',4'-dimethoxyphenyl)but-3-en-1-yl acetate, in crude extract by qHNMR using an internal standard was achieved, with high specificity and sensitivity. The selected 1H NMR signals could unambiguously be assigned and did not overlap with other resonances, including the highly similar compounds DMPBD and TMPBD. The method is linear in the concentration range of 0.70-14.52 mg/mL, with a limit of quantification of 0.46-0.68 mg/mL. The RSD values of intraday and interday precisions are in the range of 0.23%-0.74% and 0.29%-0.52%, respectively. The average recoveries are 99.54%-100.18%. A rapid, accurate, and precise method using 1H NMR for the simultaneous quantitation of four phenylbutanoids in the crude extract of Z. cassumunar rhizome was developed and validated.

Parallel-disposable pipette extraction (Pa-DPX) using cork as a green high-throughput method for the determination of four pesticides in environmental water samples with quantification by HPLC-DAD.

This study utilized cork powder as a green alternative for extractor phase using the parallel-disposable pipette extraction (Pa-DPX) method for the determination of pesticides such as atrazine, carbofuran, diuron, and simazine from aqueous environmental samples. The separation/detection was performed by HPLC-DAD. Optimization experimental conditions was achieved through univariate and multivariate designs, resulting in the following parameters: 15mg of cork as extractor phase, 7 extraction cycles with 7.5% (m/v) NaCl in the sample, and 10s were fixed per extraction cycle. For the desorption step, 300 µL of acetonitrile was used as solvent with 1 desorption cycle. The limit of detection and limit of quantification values were 7.5µg L-1 and 25µg L-1 for atrazine, carbofuran, and diuron, and 15.1µg L-1 and 50µg L-1 for simazine, respectively. Linear working range varied from 25 to 400µg L-1 with coefficients of determination (R2) ≥ 0.983. The values of intraday precision varied from 4.9 to 18.1%, and those of interday precision varied from 21.3 to 32.2%. The accuracy of the method was determined through relative recoveries and the values ranged from 63.4 to 119.9%. The water environmental samples were collected at two different locations in Florianópolis, Santa Catarina, Brazil, and showed no detectable pesticides. The method presented here stands out due to the low consumption of sample and reagents, as well as the use of a renewable and biodegradable extraction phase (cork). The method is also a high-throughput alternative, due to the use of the Pa-DPX apparatus, which allows the preparation of up to five samples simultaneously.

Direct aqueous injection LC−ESI-MS/MS analysis for the determination of 23 EU watch list substances in whole water samples

The Watch List (WL) monitoring program in the European Union (EU) aims to identify and prioritize emerging pollutants in the aquatic environment that may pose risks to human health and the environment. Thus, reliable methods are needed to determine these substances. This study presents the development of a simple, fast, and green method for analysing 23 WL substances (4 antibiotics, 9 azole compounds, 6 other pesticides, and 4 sunscreens), covering a range of medium to very low polarity (log Kow 2.2–6.9), in whole environmental water samples using direct aqueous injection (DAI) in combination with liquid chromatography–electrospray ionization tandem mass spectrometry (LC–ESI-MS/MS). To process the samples, 0.5 mL of acetonitrile is added to 1 mL of raw water, followed by brief shaking. The mixture is then centrifuged to remove sedimented suspended solids before being injected into the LC instrument. A rigorous analytical greenness metric for sample preparation (AGREEprep) assigned this procedure a green score of 0.61. The method showed satisfactory performance characteristics for all analytes under investigation, including limits of quantification meeting the WL requirements, recoveries in the range of 79–120%, and intra-day precision with relative standard deviations in the range of 1.9–20%. The method is suitable for screening and monitoring purposes, as confirmed by the analysis of real samples.

An Estimation of Baricitinib by AQbD-driven UV Spectrophotometry Development and Validation Process

Background: Baricitinib (BCTB) is a novel Janus Kinase (JAK) 1 and 2 inhibitor used in the therapy of rheumatoid arthritis, approved by the “Food and Drug Administration” in 2018. It has significant dose-dependent effectiveness and severe side effects. Thus, it is crucial to figure out its concentration in developed dosage forms. The literature search revealed that there has only been one UV spectroscopy technique documented up to this point. Methanol was chosen as the detection medium in this approach, which is not comparable with plasma or serum. As a result, the preliminary research suggested developing a UV spectroscopic approach that can estimate BCTB concentration and compare it to its concentration in the plasma or serum. Thus, in the proposed method, 7.4 pH phosphate buffer was selected as a mobile phase. Aim: Using the Analytical Quality by Design (AQbD) methodology, a simple, robust spectrophotometric method for the detection of BCTB in API form and Niosomes drug delivery system is designed and assessed. Methods: In the AQbD approach, a face-centered CCD design of Design Expert 13 software was used to evaluate two critical method variables: scanning speed and sampling interval. The design space suitability was confirmed by standard error and overlay plots. The 2-D contour and 3-D response surface plots were used to forecast the relationship between the response variable and predictor variables. Results: Baricitinib displays an absorption maximum at 249.40 nm in saline phosphate buffer pH 7.4. The distinguished linearity of the method was obtained over a concentration of 5–30 µg/ml with a correlation coefficient (R2 ) value of 0.998. BCTB % assay was found to be near 99 %. Intraday and Interday precision were found to have % RSDs of 0.067–0.488 and 0.146–0.942, respectively. Conclusion: The established spectrophotometric technique was observed to be precise as per ICH revised guidelines ICH Q2 (R1) and Q14 for analytical method validation. Our findings are instructional for the future design and development of safe and reliable therapeutic dosage forms of BCTB for rheumatoid arthritis.

Room-temperature synthesis of fluorinated covalent organic framework coupled with liquid chromatography-mass spectrometry for determination of per- and polyfluoroalkyl substances in drinking water

The routine monitoring of per- and polyfluoroalkyl substances (PFASs) in drinking water has become an important task in the field of public health. In this study, a fluorinated covalent organic framework (COF) was synthesized at room temperature using tetra-(4-aminophenyl) methane (TAM) and 2,3,5, 6-tetrafluoro-terephthalal (TFTA) as building blocks and named as TAM-TFTA-COF. The adsorption characteristics of PFASs on the TAM-TFTA-COF were investigated through adsorption model-fitting and molecular calculation. The TAM-TFTA-COF was served as the solid phase extraction (SPE) cartridge packing for the enrichment of PFASs. Combined with liquid chromatography–tandem mass spectrometry, the proposed method showed good linearity in the range of 1.25–375 ng·L−1, low limits of detection (0.03–0.24 ng·L−1), and excellent intraday and interday precisions with RSD <10.3 %. Furthermore, this analytical method can be utilized for the determination of PFASs in tap water, spring water, and lake water with satisfactory accuracy.

Deep eutectic solution elution assisted ligand affinity assay: A useful tool for the active coumarins screening from Fructus cnidii

BackgroundMany of the ligand affinity analyses are presented in water environment, and the hydrophilic solution such as methanol is used for dissociating the bound compounds. The obtained dissociation solution needs to be concentrated for improving the sensitivity of the assay. However, it is not good for the analysis of hydrophobic and volatile compounds such as coumarins. The solubility of them in water environment is low. And they may be lost in the sample preparation process. In this work, a deep eutectic solvent elution assisted ligand affinity assay for the screening of active coumarins in plants was proposed. ResultsFor the first time, deep eutectic solvent was used as a dissociation solution for dissociating the compounds that bind with acetylcholinesterase immobilized magnetic beads. Analytes could be enriched during the phase separation process of deep eutectic solvent. The concentration of the analytes in sample solution was therefore increased. It is good for the detection of coumarins. The LOD and LOQ of the proposed assay were 0.50 (μ g/mL) and 1.50 (μ g/mL), respectively. The relative recovery of the assay was 99.4 ± 7.2 %. The intra-day and inter-day precision were 7.25 % and 6.80 %. Using the assay as a tool, the active coumarins in Fructus cnidii were analyzed. And two coumarins with acetylcholinesterase inhibition activity such as imperatorin and osthole were found. SignificanceCoumarins have good solubility in the selected deep eutectic solvent, and could be enriched into hydrophobic phase during the phase separation process. The extra concentration process is not needed. Compared with traditional methods, the proposed method has the benefits of simplified operation process, high sensitivity, and is friendly to the volatile and hydrophobic compounds.

Effervescent-assisted Liquid-phase Microextraction Employing Switchable Hydrophilicity Solvent for the Determination of Cortisol and Testosterone in Oral Fluid by High-Performance Liquid Chromatography-Diode Array Detection.

Cortisol and testosterone are important biomarkers for diagnosing complex disorders, including polycystic ovary syndrome and Cushing's syndrome, where symptomatology usually overlaps with other prevalent disorders. This work proposes, for the first time, an analytical method based on a switchable hydrophilicity solvent as an extraction phase for the determination of cortisol and testosterone in oral fluid (OF) by high-performance liquid chromatography with diode-array detection. The optimized extraction conditions consisted of 1000µL of OF, 100µL of decanoic acid solution (65mg/mL), 170µL of Na2CO3, 900µL of H2SO4 and 150µL of acetonitrile for dilution. The method was validated, and coefficients of determination higher than 0.9926, the limit of detection of 4.55ng/mL and the limit of quantification of 15.00ng/mL were obtained. Intra-day precision varied from 5.6% to 11.9%, inter-day precision ranged from 6.1% to 13.5%, and relative recoveries ranged from 98.9% to 104.6% for cortisol, and 89.1% to 103.9% for testosterone. This methodology was successfully applied to five OF samples from volunteers. Moreover, the greenness of this methodology was evaluated based on the sample preparation metric of sustainability achieving a global score of 7.37 which can be considered sustainable.

Quantitative analysis of methylmalonic acid in human plasma by LC-MS/MS after derivatization

Methylmalonic acid (MMA) is a reverse biomarker of vitamin B12 that is increasingly utilized in clinical practice. However, its low sensitivity and susceptibility to strong interference from isomer present chromatographic challenges. We have developed a rapid derivatization method for plasma MMA at room temperature, converting it to the corresponding 2,2,2-trifluoroethylamide derivative using N-(3-dimethylaminopropyl)-N’-ethylcarbodiimide hydrochloride (EDC) and 2,2,2-trifluoroethylamine hydrochloride (TFEA). Amidization was completed within 10 min, followed by protein precipitation extraction of the amides with trichloroacetic acid for Liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis. This technique notably enhanced the signal-to-noise ratio of MMA in chromatography. The derivatized MMA exhibited excellent linearity within a concentration range of 42.4–2711.9 nmol/L, with a correlation coefficient (R2) of 0.9990. The intraday and interday precision of replicate measurements ranged from 2.4 % to 4.4 % and 2.6 % to 2.8 %, respectively, while the recovery fell between 97.9 % and 100.1 %.

Development and Validation of an optimized HPLC-UV analytical method for the quantification of 5-Methyltetrahydrofolate (Active Folate) in dietary Supplements: Application to commercial tablets and capsules

5-MTHF is the active form of folic acid, and it is provided as a dietary supplement in the market. Therefore, in this study we provide a new, simple and economic HPLC-UV analysis method to quantify 5-MTHF in dietary supplement. A new HPLC-UV method was developed and validated for the quantification of 5-methyltetrahydrofolate in dietary supplements. The method was optimized in terms of chromatographic conditions, including the mobile phase composition, chromatographic column, and the sample preparation diluent using ascorbic acid. A combination of ammonium acetate buffer with methanol (90:10) was the optimal mobile phase, achieving a retention time of approximately 11.4 min on C18 column with good column performance (N = 6786, Asymmetry = 1.03). The method was validated for selectivity, linearity, precision, accuracy, and robustness. The calibration curve was linear (r2 > 0.999) over the concentration range of 1.20 to 4.80 µg/mL. Precision was demonstrated with RSD% values ranging from 1.38 % to 2.90 % for intra-day precision and 0.96 % to 2.61 % for inter-day precision. Accuracy was confirmed with recovery values between 96.5 % and 110.6 % for spiked samples. To demonstrate the method’s applicability, ten brands of 5-MTHF dietary supplements were collected from local pharmacies. Assay results showed significant variability, with four brands having contents below the acceptable range (90–110 % of labeled contents) and two brands exceeding this range. The stability of 5-MTHF in these products appeared to be formula-dependent, emphasizing the need for regulatory revisions to mandate stability testing for dietary supplements.