Cinnamon essential oil is commonly used in daily life, production, and health. This study introduces a straightforward, reliable, and highly accessible HPLC method for the quality assessment of cinnamon essential oil. Three selected markers: trans-cinnamaldehyde, cinnamic acid, as the main components, are used as components in the process of quantifying cinnamon components, and coumarin is of interest due to concerns about the risk of liver toxicity. These three components are assayed in cinnamon bark and leaves, the raw materials for distilling cinnamon essential oil. The sample was diluted with a 7:3 methanol:water ratio, analyzed by HPLC-DAD using Phenomenex Gemini C18 column (250 & times; 4.6 mm, 5 & micro;m), a gradient mobile phase program consisting of acetonitrile and 0.05% phosphoric acid, and a detection wavelength of 280 nm. The validated method had suitable specificity, a linear range from 0.51 to 385.16 & micro;g/mL for coumarin, 0.48-199.60 & micro;g/mL for cinnamic acid, and 38.18-458.16 & micro;g/mL for trans-cinnamaldehyde, good repeatability with RSD from 0.7-1.1%, and the precision of coumarin, cinnamic acid, and trans-cinnamaldehyde were 100.3%, 98.8%, and 100.5%, respectively, meeting the requirements of AOAC. This method was used to quantify seventeen cinnamon essential oil samples, including essential oil from leaves and bark of Cinnamomum cassia, commercial samples, and smallholders' products.
A stability-indicating reverse-phase high-performance liquid chromatography method with charged aerosol detection was developed and validated for the quantitative determination of bempedoic acid and six related impurities. Due to weak or absent UV chromophores in the analyte and its impurities, the Charged aerosol detector was selected for its near-universal detection capability, enabling consistent and sensitive quantification. Method development followed an Analytical Quality by Design approach using a design-of-experiments strategy to evaluate the effects of mobile-phase pH, flow rate, and column temperature on critical analytical attributes, ensuring method robustness. Chromatographic separation was achieved on an XBridge BEH C18 XP column (150 & times; 4.6 mm, 2.5 & micro;m) using gradient elution with 10 mM ammonium formate buffer (pH 4.0) and acetonitrile at a flow rate of 1.00 mL/min and column temperature of 40 degrees C. The method was validated according to ICH guidelines. Forced degradation studies confirmed its stability-indicating capability by effectively separating degradation products from the analyte. Greenness assessment using established green analytical chemistry metrics demonstrated favorable environmental performance with reduced solvent consumption and waste generation. The developed method is suitable for routine quality control and stability testing of bempedoic acid drug substance.
This work provides the establishment of a simple, affordable procedure to check the folic acid content of locally sourced pharmaceutical samples in rural areas. This procedure is designed to identify adulteration/deterioration in inadequately stored medicines. Folic acid was separated with pH 8.0 phosphate buffer and methanol by solvent extraction before analysis. The developed method was then validated for evaluating the pharmaceutical properties of different brands of folic acid in combination with IPC available on the local market, following ICH guidelines. The mobile phase of pH 7.2 was prepared by a mixture of potassium dihydrogen phosphate and sodium perchlorate buffer with methanol in a ratio of 185:15. Results indicated their suitability to be adopted on commercial scale as their linearity over a concentration range of 3.5 to 28 & micro;g/mL with an R2 value of 0.9995 and RSD: <= 1%. This method can be used in quality of raw materials and finished products.
Nitrosamine Drug Substance Related Impurities constitute a distinct and analytically demanding subclass of nitrosamines due to their high molecular weight, active pharmaceutical ingredient specific formation pathways, and pronounced matrix-dependent ionization behavior, differentiating them from small-molecule nitrosamines addressed by multi-analyte screening approaches. Following the recent inclusion of N-Nitroso Dasatinib and N-Nitroso N-Deshydroxyethyl Dasatinib in Annex 1 of the updated EMA guideline, there is a growing need for drug-specific, regulation-driven analytical strategies capable of reliably controlling these impurities at trace levels. In this study, a highly sensitive LC-MS/MS method was developed and fully validated for quantitative determination of dasatinib-related NDSRIs, addressing compound-specific challenges including severe induced ion suppression, complex fragmentation behavior, and potential isobaric interferences. The method demonstrated excellent linearity (R-2 > 0.995), accuracy (recoveries within 80-120%), and precision across concentration ranges of 1.875-75.0 ppb for N-Nitroso Dasatinib and 0.50-20.0 ppb for N-Nitroso N-Deshydroxyethyl Dasatinib, in compliance with ICH Q2(R2) requirements. To ensure unambiguous structural confirmation and regulatory defensibility, high-resolution LC-QTOF-MS was applied as a confirmatory tool, achieving mass accuracies below 2 ppm and consistent fragmentation patterns. This compound-tailored analytical workflow provides a robust, scientifically defensible solution for routine quality control, lifecycle management, and proactive regulatory compliance monitoring of dasatinib-related NDSRIs worldwide.
When quantifying gluten, it is important to ensure that variations of the protein composition in samples due to environmental influences or genetic differences between cultivars do not affect the analytical results. One way to achieve this is to use multiple quantitative markers that cover the majority of gluten proteins in the sample. This study advances our previously developed liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for gluten quantification by introducing an automated workflow for marker peptide selection and data processing. The algorithm identifies optimal combinations of peptides, reducing the time and labor costs associated with manual selection. The evaluation confirmed that algorithm-derived marker sets perform equivalently to those selected by the operator, which was demonstrated by reproducing analytical procedures for gluten determination developed earlier. The method previously used to assess the content of gluten originating from wheat was tested for another gluten source: rye. It showed acceptable precision and recovery, with an overall lower limit of quantification of 10 mg gluten/kg product, demonstrating the applicability of the method to matrices other than wheat. Automated data processing further improves robustness and facilitates routine implementation.
This study investigates the interaction between quercetin (Quer) and cis-diphenylethylene (cis-DPE) using reversed-phase HPLC and Density Functional Theory (DFT). Chromatographic analyses under isocratic conditions revealed notable changes in peak intensities and UV-visible spectra bands upon interaction. While peak intensities increased with increasing concentration, retention times remained unchanged. Upon quercetin-cis-DPE interaction, shifts in the OH (with broadening), C-O, and carbonyl IR bands were observed. Parametric studies showed that peak intensity increased with concentration, while retention slightly increased with temperature, suggesting a spontaneous exothermic process accompanied by increased system disorder under the studied conditions. Additional chromatographic experiments with trans-DPE were performed for peak assignment. The second peak observed in the Quer-trans-DPE system (28.04 min) closely matches the second peak detected in the Quer-cis-DPE system, indicating the formation of a trans-related species via partial isomerization. DFT calculations identified the 4 '-OH group of Quer as the most reactive site. The Quer-cis-DPE complex exhibits a lower electronic energy (-44755.81 eV) than the Quer- trans-DPE complex (-44730.22 eV), indicating higher stability. HOMO/LUMO (Highest Occupied and Lowest Unoccupied Molecular orbitals, NCI (Non Covalent Interactions), and IRI (Interaction Region Indicator) analyses further support hydrogen transfer and intermolecular interactions with a HOMO-LUMO gap of the cis system (3.43 eV) lower than that of the trans one (3.59 eV), suggesting higher reactivity and a more favorable interaction. Computational and experimental results are in agreement regarding the stability of the quercetin-cis/trans-DPE interacting systems.
We developed a streamlined analytical method integrating solid-phase supported liquid-liquid extraction (SPS-LLE) with ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) for simultaneous quantification of 35 banned synthetic colorants in preserved and dried fruits. The optimized protocol employs a Poroshell 120 SB-Aq column (2.1 & times; 100 mm, 1.9 mu m) with a 0.1% formic acid-acetonitrile/10 mmol L-1 ammonium acetate gradient, achieving chromatographic separation within 15 min. Key parameters, including phase-separation inducers (ammonium sulfate), elution solvents (10% ammonia-methanol), and reconstitution media (ultrapure water), were systematically optimized to minimize matrix interference. Validation studies demonstrated robust linearity (R-2 > 0.990), sensitivity (LOD: 3 & times; S/N; LOQ: 10 & times; S/N), and precision (RSD <= 7.8%), with recoveries ranging from 80.3% to 117.1%. Application to 20 commercial samples revealed non-compliant additives (e.g., Tartrazine at 39,260.0 mu g kg(-1)) in 40% of products, particularly unregulated street-vended items. This method offers a high-throughput solution for regulatory monitoring of synthetic colorant abuse in complex food matrices.
In this study, 14 Argentinian unconventional crude oil samples from five source rock extraction of renowned hydrocarbon basins were organically and geochemically characterized by GC-FID to obtain the whole oil paraffins fingerprint and GC-MS to classical biomarkers identification. The implemented methodology was rigorously validated using a reference oil and it proved to be highly precise and accurate; a total of seventy-eight diagnostic ratios were obtained to study the differences between formations and to contribute to the systematic knowledge of source rock origin, depositional conditions, maturity and biodegradation of oils: Pristane/phytane (0.89-1.89), Carbon Preference Index (values close to 1), C29 S/(S + R) sterane (0.49-0.58), moretane/hopane ratio (0.0-0.1), and Gammacerane index (values higher than 0.4). These results indicate that these samples are mature oils, formed in a reducing environment, little to no affected by biodegradation or other alteration processes. Along with geochemical interpretation of the results, a chemometric analysis was performed to help interpret differences between basins and formations. Specifically, Principal Component Analysis (PCA), where the three basins were found to be clearly separated. Analysis performed by hierarchical cluster analysis (HCA) produced similar results to PCA in terms of separation and grouping.
Pharmaceutical formulations frequently contain excipients such as glycerol, sorbitol, and sucrose to improve stability and palatability. However, their concentrations are often undeclared, and excessive levels may cause adverse effects. Existing analytical approaches for their quantification are either time-consuming or susceptible to interferences, highlighting the need for a rapid and selective alternative. In this work, a method based on ion chromatography coupled with pulsed amperometric detection (IC-PAD) was developed for the simultaneous quantification of glycerol, sorbitol, and sucrose in pharmaceutical formulations. Using a carbohydrate-specific anion-exchange column, baseline separation was achieved within 8 min under the optimized conditions. Method validation followed ANVISA guideline RDC n degrees 166/2017 and demonstrated excellent linearity (1-100 mg L-1; r2 approximate to 0.999), good precision (RSD <= 0.7%), low limits of detection (3.0-24 & micro;g L-1), and limits of quantification (9.6-74 & micro;g L-1). No significant interference from common excipients was observed, and analysis of commercial products confirmed the method's accuracy and applicability. The greenness of the method was evaluated using the AGREE software, achieving a score of 0.67. Combining short run time, high selectivity, and robustness, the proposed IC-PAD method provides a reliable tool for routine quality control and regulatory analysis of pharmaceutical sirups and related products.
In liquid chromatography simultaneous gradients of solute and temperature can be applied to control adsorption of solute on the stationary phase by simultaneously changing the composition of the mobile phase and the temperature. These parallel variations alter adsorption equilibria and solute migration rates. The work extends a general rate model to incorporate such parallel gradients, enabling the simulation of two-mode gradient elution. The model consists of coupled nonlinear convection-diffusion equations which describe the simultaneous transport of mass, energy, and solvent composition within a chromatographic column. By introducing time-dependent gradients of temperature and solvent composition, it accurately represents advanced elution protocols. The integrated retention model combines the theory of linear solvent strengths with modified van't Hoff behavior, defining Henry's constants and the non-linearity coefficients as functions of solvent composition and temperature. The governing equations are solved with a high-resolution finite-volume scheme, effectively capturing temperature-dependent retention, nonlinear adsorption, and sharp concentration fronts. The effects of positive, negative, and mixed solvent-temperature gradients, as well as important operational parameters, on peak shape and separation efficiency are quantified through numerical simulations. The findings show that, in comparison to single-gradient or isothermal operations, combined gradients achieve faster elution, better peak symmetry, and better separation performance.
Swertia chirayita and Andrographis paniculata are widely used in Ayurveda, Unani, and Homoeopathy but are often adulterated due to morphological similarities and overlapping vernacular names. This study applies a marker-based quality control strategy using high-performance thin-layer chromatography (HPTLC) and ultra-performance liquid chromatography-photodiode array (UPLC-PDA) for determining swertiamarin (SN) in S. chirayita and andrographolide (AD), neoandrographolide (ND), and andrograpanin (AN) in A. paniculata, along with their respective Homoeopathic mother tinctures (HMTs). HPTLC qualitative analysis using optimized solvent systems produced distinct and well-resolved bands, confirming the presence of SN in all S. chirayita samples and the three diterpenoid markers in all A. paniculata samples. UPLC-PDA analysis on a Waters Acquity BEH C18 column (100 & times; 2.1 mm, 1.7 & micro;m) with a binary gradient of acetonitrile and 0.1% formic acid at 0.5 mL/min and 220 nm enabled high-resolution separation of all markers. The method showed excellent linearity (R2 >= 0.9990), with limits of detection (LOD) and limits of quantification (LOQ) values of 1.89-2.03 and 5.71-6.14 & micro;g/mL, respectively, and %RSD below 2% for precision and accuracy studies. An artificial mixture of S. chirayita and A. paniculata yielded distinct UPLC fingerprints, supporting practical quality assessment of raw drugs and related formulations.
Lepidium sativum, a member of the Brassicaceae family, is traditionally used as an edible herb with notable antioxidant, antibacterial, anticancer, and nutritional properties. This study focused on developing and validating a high-performance thin-layer chromatography (HPTLC) method for the quantitative estimation of lepidine in L. sativum seed extract. The HPTLC analysis was performed using pre-coated HPTLC plates 60 F-254 as the stationary phase. The developing mobile phase of n-butanol: water: acetic acid (7:2:1.5 v/v/v) showed good separation of lepidine with an R-f of 0.70 +/- 0.02. Densitometric determination of lepidine was detected at 314 nm using a CAMAG TLC scanner III. The method was validated under ICH guidelines. It demonstrated good linearity between 0.5 and 4 & micro;g/band, with an R-2 of 0.991. The detection limit (LOD) and the quantification limit (LOQ) were 0.162 & micro;g/band and 0.491 & micro;g/band, respectively. The sonication extract of L. sativum showed that the highest lepidine content of 7.8 +/- 0.36 mg/g and exhibited strong antioxidant activity in the DPPH assay, with an IC50 of 14.9 & micro;g/ml. The validated HPTLC method proved to be simple, precise, accurate, cost-effective, and suitable for the rapid quantification of lepidine and supporting the antioxidant potential of L. sativum seed extract. [GRAPHICS]
The use of computers in chromatographic techniques has transformed phytochemical analysis in last decades, particularly in the area of computational intelligence. Several phytochemicals, including carotenoids, polyphenols, polysaccharides, and phytosterols, are vital to the plant and have numerous benefits, including regulating DNA metabolism, exhibiting antioxidant properties, and inducing cell death. Despite its importance, researching phytochemicals presents challenges, particularly in the areas of phenolic extraction, natural ingredient separation, and herbal medicine adulteration. Merging computational intelligence with traditional chromatographic techniques enhances phytochemical analysis by increasing the precision of both qualitative and quantitative compound property predictions. Furthermore to making it easier to identify and characterize phytochemicals, this merging allow a thorough comprehension of them. However, traditional methods provide an excellent foundation to support the analytical process, whereas computational methods provide more accurate and reliable prediction models. However, there are several challenges in developing efficient chromatographic separation and analysis methods due to the complexity and diversity of botanical extracts. The study explores the transformation of chromatographic and computational methods in phytochemical analysis, emphasizing their synergistic potential in improving compound identification. It further discuss the of integration of the both the methods and future prospects for optimizing the phytochemicals profiling for the nutraceutical application.
An efficient strategy integrating cloud point extraction (CPE) and high-speed counter-current chromatography (HSCCC) was developed to separate active flavonoids from Scutellaria baicalensis Georgi. The liquid stationary phase advantage of HSCCC enabled the direct injection of the surfactant-rich phase, preventing the irreversible adsorption of surfactants on solid carriers and avoiding contamination, thereby establishing a rapid and efficient extraction and enrichment methodology. By optimizing the surfactant type and concentration, solid-liquid ratio, extraction pressure, and extraction time, CPE additive type and concentration, equilibrium temperature, and extraction time, the optimal CPE process was obtained. Based on the partition coefficient (KD) values of the target compounds, a solvent system composed of trichloromethane: tetrachloromethane: methanol: water (1:4:3:2, v/v/v/v) was selected from various tested systems, as it provided ideal retention behaviors for the separation. Baicalein (I), wogonin (II), and oroxylin A (III) were successfully isolated from Scutellaria baicalensis with HPLC purities of 96.21%, 94.91%, and 96.68%, respectively. Their chemical structures were further confirmed by NMR and ESI-MS. A rapid and efficient method for enrichment, separation, and preparation of high-purity monomeric compounds was achieved by applying CPE and HSCCC. This comprehensive method provides a new solution for the extraction and separation of natural compounds.
A high-performance liquid chromatography method was developed and validated for quantifying bakuchiol and ten bioactive constituents from Psoralea corylifolia in cosmetics. Sample preparation involved ultrasonic-assisted extraction with methanol, followed by chromatographic separation on a CAPCELL PAK MG III C18 column (250 & times; 4.6 mm, 5 mu m) using gradient elution with methanol and 0.1% aqueous phosphoric acid. External standard calibration demonstrated excellent linearity (r > 0.999) for all analytes, with limits of detection (LOD) and limits of quantitation (LOQ) ranging from 0.6 to 1 and 2 to 4 mg kg(-1), respectively. Method validation showed mean recoveries of 87.9 to 109.9% and relative standard deviations of 0.4 to 4.9% across toner, cream, and gel matrices. The validated method was successfully applied to the analysis of 14 commercial cosmetic products labeled to contain bakuchiol. Quantitative analysis revealed bakuchiol content ranging from 0.0084 to 0.7548% in 13 of the 14 samples, while one sample showed no detectable bakuchiol (below the LOD, 0.6 mg kg(-1)). This method features operational simplicity, precise quantification, and effective separation of structural isomers, thereby enabling robust quality control, reliable label claim verification, and regulatory compliance for P. corylifolia-derived ingredients across diverse cosmetic formulations. [GRAPHICS]
The 2018 U.S. Farm Bill legalized hemp but created regulatory gaps that contributed to the emergence of synthetic THC analogs in consumer products. An UHPLC-DAD method was developed and validated in accordance with ISO/IEC 17025 guidelines for the quantitative analysis of twenty-three neutral cannabinoids, including two epimeric pairs and six Delta(8)/Delta(9) positional isomer pairs of synthetic THC analogs, six naturally occurring neutral cannabinoids, and one common synthetic THC byproduct. The method was applied to fifteen commercial products comprising four tinctures, six vaping oils, and five gummies. Fourteen cannabinoids were detected above their limits of quantification at concentrations ranging from 0.008-79.6%, with triplicate relative standard deviations of 0.8-18.3%. Analytical results revealed mislabeling in ten of the fifteen samples. Real-time recovery was evaluated using spiked abnormal CBD, a synthetic cannabinoid structurally unrelated to the target analytes, yielding recoveries of 95.0-110.3% with RSDs of 1.0-10.5% across all matrices. High-resolution ESI/TOFMS was used for optional confirmation, verifying method specificity, minimal matrix interference, and the presence of several HHC stereoisomers. [GRAPHICS]
