Search Results (55)

Nicotine is a highly used addictive substance that has recently also become available through electronic cigarettes. Here we present a study of nicotine from e-cigarette liquids through reversed-phase (RP) and hydrophilic interaction (HILIC) liquid chromatography. Multiple aqueous mobile-phase additives are considered for the RP mechanism, focusing on chaotropic agents, mobile-phase concentrations and mixing ratios, and column temperature. Sample preparation was conducted by toluene liquid–liquid extraction of e-cigarette liquids diluted with aqueous 25 mM NaHCO₃/Na₂CO₃. Optimal RP results for retention and peak symmetry were obtained using aqueous 0.1% formic acid and 20 mM ammonium hexafluorophosphate with 0.1% formic acid in acetonitrile, using a gradient profile with a C18 column, exploited at 40 °C and a 1.5 mL/min flow rate. A dilute-and-shoot alternative with automated flow reversal after isocratic elution is presented. For HILIC, aqueous 100 mM ammonium formate and 0.1% formic acid in acetonitrile were used as mobile-phase components, using a gradient profile, on a Thermo Scientific™ Acclaim™ Mixed-Mode HILIC-1 column, operated at 25 °C with a 1 mL/min flow rate. UV detection was at 260 nm. Absolute limits of quantitation in the 1 μg/mL range were obtained for all tested alternatives, with 1 μL injection volumes. Full article

To enhance the signal intensity of kynurenines, which are present at trace concentrations in biological fluids, a novel analytical approach was developed, combining pressure-assisted electrokinetic injection (PAEKI) with a mixed micelle system based on sodium dodecyl sulfate (SDS) and Brij-35. The method was applied to key compounds of the kynurenine pathway, including L-tryptophan, kynurenine, 3-hydroxykynurenine, and kynurenic acid, as well as to the aromatic amino acids (AAs) L-tyrosine and L-phenylalanine. PAEKI was performed by electrokinetic injection for 2 min at −6.5 kV (reversed polarity) and 0.5 psi (3.45 kPa) using a fused silica capillary (50 cm in length, 50 µm inner diameter). The background electrolyte (BGE) consisted of 20 mM Na₂B₄O₇ (pH 9.2), 2 mM Brij-35, 20 mM SDS, and 20% (v/v) methanol (MeOH). The limit of detection (LOD) using a diode array detector (DAD) was 1.2 ng/mL for kynurenine and ranged from 1.5 to 3.0 ng/mL for the other analytes. The application of PAEKI in conjunction with micellar electrokinetic capillary chromatography (MEKC) and solid-phase extraction (SPE) of artificial urine samples resulted in a 146-fold increase in signal intensity for kynurenines compared to that observed using the hydrodynamic injection (HDI) mode. The developed method demonstrates strong potential for determining kynurenine pathway metabolites in complex biological matrices. Full article

The antihypertensive drugs indapamide, atenolol, metoprolol, propranolol and bisoprolol were considered in this research. Because they have structures that are affected by pH, developing a chromatographic method was challenging. Based on the speciation diagram of these compounds versus pH scale, a mixed-mode stationary phase (hydrophobic stationary phase, C18 and strong cation exchanger (SCX)) was our first choice. Design of Experiments (DoE) was used to estimate how various factors such as pH, mobile phase composition and flow rate influenced chromatographic performance. As a result, the separation was achieved in 24 min using an aqueous phosphate buffer phase (pH 7.20) and 20 mM triethylamine, with methanol being used as organic modifier (30%). Their retention mechanism was investigated. The new method was validated in term of linearity, limits of detection and quantification, precision, accuracy, and robustness. The method was applied to river water samples, and good results were obtained. Full article

Rice husk (RH), a globally abundant agri-food waste, presents a promising renewable silicon source for producing SBA-15 mesoporous silica-based materials. This study aimed to synthesize and bifunctionalize SBA-15 using RH as a silica precursor, incorporating sulfonic and octadecyl groups to create a mixed-mode sorbent, RH-SBA-15-SO₃H-C18, with reversed-phase and cation exchange properties. The material’s structure and properties were characterized using advanced techniques, including X-ray diffraction, infrared spectroscopy, N₂ adsorption–desorption isotherms, nuclear magnetic resonance, and electron microscopy. These analyses confirmed an ordered mesoporous structure with a high specific surface area of 238 m²/g, pore volume of 0.45 cm³/g, pore diameter of 32 Å, and uniform pore distribution, highlighting its exceptional textural qualities. This sorbent was effectively utilized in solid-phase extraction to purify 29 alkaloids from three families—tropane, pyrrolizidine, and opium—followed by an analysis using ultra-high performance liquid chromatography coupled to ion-trap tandem mass spectrometry. The developed analytical method was validated and applied to gluten-free bread samples, revealing tropane and opium alkaloids, some at concentrations exceeding regulatory limits. These findings demonstrate that RH-derived RH-SBA-15-SO₃H-C18 is a viable, efficient alternative to commercial sorbents for monitoring natural toxins in food, offering a sustainable solution for repurposing agri-food waste while addressing food safety challenges. Full article

In this work, two novel (-)-menthol-based hydrophobic natural eutectic solvents with vanillin and cinnamic acid were prepared and applied as extraction solvents. In this regard, 12 endocrine disruptors, including phenol, 2,4-dimethylphenol, 2,3,6-trimethylphenol, 4-tert-butylphenol, 4-sec-butylphenol, 4-tert-amylphenol, 4-n-hexylphenol, 4-tert-octylphenol, 4-n-heptylphenol, 4-n-octylphenol, and 4-n-nonylphenol and bisphenol A, were studied in a green tea drink. A temperature-controlled liquid–liquid microextraction was used as the extraction method, and nano-liquid chromatography–ultraviolet detection was used as the separation and determination system. Different parameters affecting the compatibility of the non-ionic eutectic solvents with water-polar organic solvent mixtures and chromatographic and detection systems were optimized, including injection/dilution solvent, injection mode, mobile phase composition, and step gradient. With the same purpose, two stationary phases were tested, including XBridge^® C₁₈ and a mixed-phase Cogent C₃₀-XBridge^® C₁₈. Finally, the greenness and blueness of the methodology were assessed to evaluate the environmental profile and usability of the procedure. Full article

MonoTrap^TM was introduced in 2009 as a novel miniaturized configuration for sorptive sampling. The method for the characterization of volatile organic compound (VOC) emission profiles from hot mix asphalt (HMA) consisted of a two-step procedure: the analytes, initially adsorbed into the coating in no vacuum- or vacuum-assistance mode, were then analyzed following an automated thermal desorption (TD) step. We took advantage of the theoretical formulation to reach some conclusions on the relationship between the physical characteristics of the monolithic material and uptake rates. A total of 35 odor-active volatile compounds, determined by gas chromatography-mass spectrometry/olfactometry analysis, contributed as key odor compounds for HMA, consisting mainly of aldehydes, alcohols, and ketones. Chemometric analysis revealed that MonoTrap^TM RGC18-TD was the better coating in terms of peak area and equilibrium time. A comparison of performance showed that Vac/no-Vac ratios increased, about an order of magnitude, as the boiling point of target analytes increased. The innovative hybrid adsorbent of silica and graphite carbon monolith technology, having a large surface area bonded with octadecylsilane, showed effective adsorption capability, especially to polar compounds. Full article

Host cell proteins (HCPs) are one of the process-related impurities that need to be well characterized and controlled throughout biomanufacturing processes to assure the quality, safety, and efficacy of monoclonal antibodies (mAbs) and other protein-based biopharmaceuticals. Although ELISA remains the gold standard method for quantification of total HCPs, it lacks the specificity and coverage to identify and quantify individual HCPs. As a complementary method to ELISA, the LC-MS/MS method has emerged as a powerful tool to identify and profile individual HCPs during the downstream purification process. In this study, we developed a sensitive, robust, and reproducible analytical flow ultra-high-pressure LC (UHPLC)-high-resolution accurate mass (HRAM) data-dependent MS/MS method for HCP identification and monitoring using an Orbitrap Ascend BioPharma Tribrid mass spectrometer. As a case study, the developed method was applied to an in-house trastuzumab product to assess HCP clearance efficiency of the newly introduced POROS™ Caprylate Mixed-Mode Cation Exchange Chromatography resin (POROS Caprylate mixed-mode resin) by monitoring individual HCP changes between the trastuzumab sample collected from the Protein A pool (purified by Protein A chromatography) and polish pool (purified by Protein A first and then further purified by POROS Caprylate mixed-mode resin). The new method successfully identified the total number of individual HCPs in both samples and quantified the abundance changes in the remaining HCPs in the polish purification sample. Full article

The adsorption of l-tryptophan (l-Trp) was studied based on the hydrophobic interaction/ion exchange/ion exclusion mixed-mode adsorption resin D301. Firstly, the interaction mode between l-Trp and resin was analyzed by studying the influence of pH variation on the adsorption capability and the dissociation state of l-Trp. Secondly, the adsorption mechanism was illuminated by studying the adsorption equilibrium and kinetic behaviors. The adsorption equilibrium and a kinetics model were constructed. The augmentation of pH gradually elicited an enhancement in the adsorption capacity of l-Trp. l-Trp existing in varied dissociation states could be adsorbed by the resin, and the interaction mode relied upon the pH of the solution. An integrated adsorption equilibrium model with the coadsorption of different dissociation states of l-Trp was developed and could predict the adsorption isotherms at various pH levels satisfactorily. Both external mass transfer and intra-particle diffusion collectively imposed constraints on the mass transfer process of l-Trp onto the resin. An improved liquid film linear driving force model (ILM) was constructed, and the model provided a satisfactory fit for the adsorption kinetics curves of l-Trp at various pH levels. l-Trp molecules had a high mass transfer rate at a relatively low solution pH. Full article

Amino acids (AAs) and their metabolites are important building blocks, energy sources, and signaling molecules associated with various pathological phenotypes. The quantification of AA and tryptophan (TRP) metabolites in human serum and plasma is therefore of great diagnostic interest. Therefore, robust, reproducible sample extraction and processing workflows as well as rapid, sensitive absolute quantification are required to identify candidate biomarkers and to improve screening methods. We developed a validated semi-automated robotic liquid extraction and processing workflow and a rapid method for absolute quantification of 20 free, underivatized AAs and six TRP metabolites using dual-column U(H)PLC-MRM-MS. The extraction and sample preparation workflow in a 96-well plate was optimized for robust, reproducible high sample throughput allowing for transfer of samples to the U(H)PLC autosampler directly without additional cleanup steps. The U(H)PLC-MRM-MS method, using a mixed-mode reversed-phase anion exchange column with formic acid and a high-strength silica reversed-phase column with difluoro-acetic acid as mobile phase additive, provided absolute quantification with nanomolar lower limits of quantification within 7.9 min. The semi-automated extraction workflow and dual-column U(H)PLC-MRM-MS method was applied to a human prostate cancer study and was shown to discriminate between treatment regimens and to identify metabolites responsible for discriminating between healthy controls and patients on active surveillance. Full article

There is great concern in equine sport over the potential use of pharmaceutical agents capable of editing the genome or modifying the expression of gene products. Synthetic oligonucleotides are short, single-stranded polynucleotides that represent a class of agents capable of modifying gene expression products with a high potential for abuse in horseracing. As these substances are not covered by most routine anti-doping analytical approaches, they represent an entire class of compounds that are not readily detectable. The nucleotide sequence for each oligonucleotide is highly specific, which makes targeted analysis for these agents problematic. Accordingly, we have developed a non-targeted approach to detect the presence of specific product ions that are not naturally present in ribonucleic acids. Briefly, serum samples were extracted using solid-phase extraction with a mixed-mode cartridge following the disruption of protein interactions to isolate the oligonucleotides. Following the elution and concentration steps, chromatographic separation was achieved utilizing reversed-phase liquid chromatography. Following an introduction to a Thermo Q Exactive HF mass spectrometer using electrospray ionization, analytes were detected utilizing a combination of full-scan, parallel reaction monitoring and all ion fragmentation scan modes. The limits of detection were determined along with the accuracy, precision, stability, recovery, and matrix effects using a representative 13mer oligonucleotide. Following method optimization using the 13mer oligonucleotide, the method was applied to successfully detect the presence of specific product ions in three unique oligonucleotide sequences targeting equine-specific transcripts. Full article

The aim of this study was to develop and validate a fast and sensitive bioanalytical method for the accurate quantification of fosfomycin concentrations in human prostatic tissue. The sample preparation method only required milligrams of tissue sample. Each sample was mixed with two times its weight of water and homogenized. A methanol solution that was three times the volume of the internal standard (fosfomycin-13C3) was added, followed by vortex mixing and centrifugation. After its extraction from the homogenized prostatic tissue, fosfomycin was quantified by means of a liquid chromatography–tandem mass spectrometry (LC-MS/MS) triple quadrupole system operating in negative electrospray ionization and multiple reaction monitoring detection mode. The analytical procedure was successfully validated in terms of specificity, sensitivity, linearity, precision, accuracy, matrix effect, extraction recovery, limit of quantification, and stability, according to EMA guidelines. The validation results, relative to three QC levels, were 9.9% for both the within-day and inter-day accuracy (BIAS%); 9.8% for within-day precision; and 9.9 for between-day precision. A marked matrix effect was observed in the measurements but was corrected by normalization with the internal standard. The average total recovery was high (approximatively 97% at the three control levels). The dynamic range of the method was 0.1–20 μg/g (R² of 0.999). Negligible carry-over was observed after the injection of highly concentrated samples. F in the sample homogenate extracts was stable at 10 °C and 4 °C for at least 24 h. In the tissue sample freeze–thaw experiments, a significant decrease in F concentrations was observed after only two cycles from −80 °C to room temperature. The novel method was successfully applied to measure fosfomycin in prostatic tissue samples collected from 105 patients undergoing prostatectomy. Full article

Mixed-mode high-performance liquid chromatography (HPLC) is increasingly used for the analysis of ionic and highly hydrophilic drugs, which are difficult to separate by conventional single-mode HPLC. In the former case, chromatographic separation is achieved using one of the several commercially available mixed-mode stationary phases, typically combinations of reversed and ion-exchange phases. Moreover, mixed-mode stationary phases can be used as solid-phase extraction (SPE) sorbents. This review focuses on the recent applications of mixed-mode stationary phases in the chromatographic analysis of bioactive compounds, such as drugs, herbicides, and pesticides. Specifically, we briefly summarize HPLC methods utilizing mixed-mode stationary phases and SPE pretreatment procedures utilizing mixed-mode sorbents developed in the last decade, thus providing a reference work for overcoming the difficulties in analyzing ionized or hydrophilic drugs by conventional reversed-phase chromatography. Full article

A novel co-bonded octyl and pyridine silica (OPS) sorbent was prepared and applied for the solid phase extraction (SPE) of cyclopiazonic acid (CPA, a type of mycotoxin) in feed and agricultural products for the first time. A simple mixed-ligand one-pot reaction strategy was employed for OPS sorbent preparation. Nitrogen adsorption–desorption measurements, elemental analysis (EI), thermal gravimetric analysis (TGA), and Fourier transform infrared spectroscopy (FT-IR) analysis demonstrated the successful immobilization of octyl and quaternary ammonium groups onto the surface of silica gel. The large specific surface area, high-density functional groups, and mixed-mode anion-exchange characteristics of these silica particles made them the ideal material for the efficient extraction of CPA. Additionally, the OPS sorbents displayed excellent batch-to-batch reproducibility, satisfactory reusability, and low cost. The SPE parameters were optimized to explore the ionic and hydrophobic interactions between CPA and the functional groups, and the ultra-high performance liquid chromatography coupled with triple-quadrupole tandem mass spectrometry (UPLC-MS/MS) parameters were optimized to obtain a desirable extraction efficiency and high sensitivity to CPA. Meanwhile, the OPS sorbent presented a satisfactory extraction selectivity and low matrix effect. Under the optimized conditions, our developed CPA detection method was used to determine CPA level in rice, wheat flour, corn flour, peanut, and feed samples, exhibiting a lower detection limit, better linearity, higher sensitivity, and satisfactory extraction recovery rate than previously reported methods. Therefore, our method can be preferentially used as a method for the detection of CPA in agricultural products and feeds. Full article

Non-Saccharomyces yeasts, particularly Metschnikowia pulcherrima, are considered alternatives to SO₂ in winemaking, combating specific microorganisms. The sensory profile of the wine is contingent upon the type of yeast, the fermentation conditions, and the concentration and mode of application with Saccharomyces cerevisiae strains (whether pure or used in mixed/sequential co-fermentation). This study assessed the aroma in red wines produced with S. cerevisiae (Sc) and M. pulcherrima (Mp, non-Sc), incorporating variations in the method of addition and the inclusion or exclusion of SO₂. The enological parameters of the wines were slightly affected. Volatile compounds were analysed in the wines through gas chromatography–mass spectrometry (GC-MS) at three moments: at the end of malolactic fermentation (MLF) and after 6 and 9 months of bottle ageing. Sequential fermentation of Sc and Mp reduced the concentration of most identified alcohols and acids, which is favourable, as these compounds can yield undesirable aromas at high concentrations. Regardless of the yeast mixture and Mp dose, a majority of the acetate esters and ethyl esters were quantified at concentrations above their perception thresholds, thus enhancing the sensory quality of the wines. Sensory analysis of wines showed generally positive evaluations. Using non-Saccharomyces as an alternative to SO₂ improves the aromatic profile of wines. Full article

Ellagic acid, known for its various biological activities, is widely used. Ellagic acid from pomegranate peels is safe for consumption, while that from gallnuts is only suitable for external use. However, there is currently no effective method to confirm the source of ellagic acid. Therefore, this study establishes an analysis method using ultra-high-performance liquid chromatography–electrospray ionization–high-resolution mass spectrometry (UHPLC-ESI-HR-MS) to identify the components of crude ellagic acid extracts from pomegranate peels and gallnuts. The analysis revealed that there was a mix of components in the crude extracts, such as ellagic acid, palmitic acid, oleic acid, stearic acid, and 9(10)-EpODE. Furthermore, it could be observed that ellagic acid extracted from gallnuts contained toxic substances such as anacardic acid and ginkgolic acid (15:1). These components could be used to effectively distinguish the origin of ellagic acid from pomegranate peels or gallnuts. Additionally, a rapid quantitative analysis method using UHPLC-ESI-MS with multiple reaction monitoring (MRM) mode was developed for the quality control of ellagic acid products, by quantifying anacardic acid and ginkgolic acid (15:1). It was found that one of three ellagic acid health care products contained ginkgolic acid (C15:1) and anacardic acid at more than 1 ppm. Full article