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High-performance thin-layer chromatography (HPTLC) is a simple but reliable, sensitive, economical, and environmentally benign analytical technique for pharmaceuticals and herbal samples. Its advantages towards quantitative analysis include accurate sample application, quicker and efficient resolution of mixtures, minimum solvent usage and reduced sample size. The ability of HPTLC to examine numerous samples of varying complexity simultaneously and in parallel sets it apart from HPLC. This allows for a high sample throughput and the consumption of solvent and energy is low per sample. Furthermore, compared to HPLC, the use of HPTLC can conserve environmental resources because it requires little to no sample pre-treatments such as liquid–liquid extraction or solid-phase extraction. Several research studies on analytical method development using HPTLC now focus on the use of green solvents in lieu of traditional organic solvents, with the aim to reduce environmental pollution without compromising on analytical performance. In addition, advancements in assessment tools for determining “greenness”, such as AGREE (Analytical GREEnness metric approach and software), GAPI, NEMI, facilitate the evaluation of greenness for specific analysis. The green methods continue to offer rapid, sensitive, and environmentally friendly methodologies, complying with international guidelines of analytical techniques. Overall, the integration of green solvent-based HPTLC methods into analytical workflows represents a significant step towards sustainable and environmentally conscious analytical practices.

Graphical Abstract

The high solubility of ethanolamines in the aqueous solutions is a big challenge for extracting these compounds. To overcome this problem, in this research, an easy sample preparation method based on salting-out micro-liquid–liquid extraction has been introduced to extract ethanolamines from aqueous samples, and determination has been done by gas chromatography-flame ionization detector (GC-FID). The solvent 1-propanol demonstrated significantly greater extraction efficiency than other solvents, including tetrahydrofuran, acetonitrile, and diethyl carbonate. Different variables affecting the extraction and derivatization of the target compounds were also investigated and optimized. The highest response was achieved with NaCl at a concentration of 35%, using 1000 µL of 1-propanol, a sonication power of 30 W for 20 s, a derivatization agent volume of 25 µL, and a derivatization time of 5 min. The analytical performances of the method were evaluated in optimal conditions. The linear ranges varied from 2 to 4000 mg/L, depending on the compound. The method demonstrated good repeatability, with relative standard deviations ranging from 2.4 to 10.3%. Limit of detections (LODs) were in the range of 0.5–2.3 mg/L. The method was finally used to determine ethanolamines in the commercial sweetening solutions. The relative recoveries (RR%) were achieved between 89 and 111%.

During the registration procedure of a drug substance, the actual and potential impurities must be summarized in a legal documentation. Diketene is a reactive chemical compound which is used for the introduction of an acetoacetyl structural unit into organic compounds. Since this material commonly used as a reagent in the pharmaceutical industry, it is a potential impurity of certain drug substances. The aim of this work was to develop and validate a suitable method for the analysis of diketene by means of decomposition headspace gas chromatography. The product of the derivatization reaction is acetone, which can be easily measured by gas chromatography. Various parameters have been integrated into the elaborated limit test validation, such as specificity, detection limit (LD), quantification limit (LQ), system precision, recovery at limit level, robustness and solution stability. The limits of detection and quantification are 60 and 200 ppm, respectively. Using the above-mentioned new analytical method, the concentration of diketene can be determined accurately. These precise measurements are necessary for the appropriate characterization of the drug substance, which is a prerequisite step of a marketing authorization for the finished product.

This study presents a stability-indicating method for teriflunomide, a drug administered to adults for the treatment of multiple sclerosis (MS) in its various forms, including clinically isolated syndrome, relapsing–remitting, and active secondary progressive disease. The method, which has been validated comprehensively for both impurities and the assay, plays a key role in determining teriflunomide in tablet formulations. It successfully separates two specified degradation impurities in line with the European Pharmacopeia (EP) standards. To assess the stability of teriflunomide, the method subjects it to stress conditions such as acid/base hydrolysis, oxidation, photodegradation, and thermal degradation. Using a C18 column at 30 ℃, with a mobile phase of pH 3.4 solutions and acetonitrile at 0.8 mL/min, and UV detection at 248 nm, the method ensures the resolution of degradation products from the primary teriflunomide peak. The teriflunomide peak remained pure under all stress conditions, confirming the specificity and stability-indicating nature of the method. The validation followed FDA and ICH guidelines, with results for linearity, accuracy, precision, specificity, robustness, limit of detection, limit of quantification, and system suitability all meeting established acceptance criteria.

This study describes a simple, rapid, sensitive and selective RP-HPLC method for the quantification of daprodustat in bulk and tablet dosage forms. In the study, the stationary phase consisted of a Zorbax SB-C18 (150 mm × 4.6 mm, 5 μm particle size) column, and the mobile phase consisted of a mixture of acetonitrile:distilled water (80:20, v/v) containing 0.1% formic acid. The flow rate was 1.2 mL min⁻¹, the injection volume was 10 μL and the column temperature was set at 30 °C. The linearity range of the method was 10–150 μg mL⁻¹ and acceptable precision and accuracy values were obtained for the values in this range. After the developed method was validated according to ICH Q2 (R1) guideline, it was successfully applied to bulk and tablet dosage form preparations of daprodustat. The method has also been successfully applied to forced degradation studies with exposure to various stress conditions, including acid, base, oxidative stress and thermal degradation following ICH Q1B guidelines.

Regulatory agencies prioritize the safety and efficacy of pharmaceuticals. In this context, our research aims to develop a simple, sensitive, and eco-friendly LCMS-compatible high-performance liquid chromatography stability indicating analytical method (SIAM) technique to identify impurities in Acalabrutinib (ACB) under various stress conditions. This method is designed in compliance with ICH QIA (R2) and Q3, incorporating Green Chemistry principles and Quality by Design (QbD). To optimize the method, an ideal split-plot design was used to evaluate critical method parameters (CMPs), followed by a central composite design (CCD) to refine the conditions. Statistical analysis revealed p values < 0.001 for the model and 0.05 for lack of fit, indicating the most suitable statistical model for the evaluated responses (peak resolutions R1–R4). The optimized method attributes include an ACN:ammonium acetate buffer (pH 5) ratio of 50:50 (v/v), a flow rate of 0.8 mL/min, and a column oven temperature of 35 °C, derived from CCD. An isocratic elution method using the Waters e2998 HPLC Kromasil 100-5-C18 (250 × 4.6 mm; 5 µm) analytical column enabled superior separation of components, with a run time of 35 min and a wavelength of 254 nm. Stress studies revealed that ACB is sensitive to hydrolysis and photolytic conditions, with ACN identified as the most suitable diluent. Method validation was conducted according to ICH Q2 guidelines, and showed a correlation coefficient (r²) exceeding 0.992, with RSD values (n = 6) ranging from 0.76 to 1.93% across the LOQ-150% range. Specificity studies confirmed no interference between impurities and active analytes. Through stress testing, 28 degradation products were identified, and the method was assessed for eco-friendliness using seven different tools, confirming its sustainable nature for pharmaceutical applications.

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A novel HPLC system utilizing a phase-separation multiphase flow as the eluent has been developed, referred to as phase-separation mode. This research explores the influence of the porous structure in an octadecyl-modified silica (ODS) column (with a pore diameter of 12 nm) on chromatographic outcomes under the phase-separation mode in HPLC. The chromatograms obtained from the porous ODS column were compared to those generated with a non-porous ODS column. In preliminary experiments, twenty-four mixed solutions, comprising combinations of water/acetonitrile/ethyl acetate and water/acetonitrile, were introduced as eluents at a column temperature of 20 °C. A model mixture of 2,6-naphthalenedisulfonic acid (2,6-NDS) and 1-naphthol (1-NA) was injected into the system, with separation achieved in most solutions except for some highly organic solvent-rich solutions where 2,6-NDS eluted faster than 1-NA, indicating reverse-phase mode operation. Subsequently, the separation of the model mixture was assessed at 0 °C, and four specific ternary mixtures were analyzed in detail at both 20 °C and 0 °C. These ternary mixtures, defined by their volume ratios, exhibited a two-phase separation, establishing a phase-separation multiphase flow. Consequently, the solution flow was homogeneous at 20 °C and heterogeneous at 0 °C. For instance, solutions with water/acetonitrile/ethyl acetate ratios of 20:60:20 (organic solvent-rich) and 70:23:7 (water-rich) were introduced as eluents at both 20 °C and 0 °C. At 0 °C in the organic solvent-rich eluent, 1-NA eluted faster than 2,6-NDS, characteristic of the phase-separation mode. In contrast, the water-rich eluent resulted in faster elution of 2,6-NDS at both temperatures. The porous ODS column displayed improved separation efficiency at 0 °C compared to the non-porous column, which can be attributed to the porous effect under phase-separation conditions.

Combination nasal spray products containing fluticasone furoate and oxymetazoline hydrochloride are commonly used to treat asthma, chronic pulmonary diseases, and perennial allergic rhinitis. However, testing for impurities in pharmaceutical products is a challenging task due to the similarity in properties between the impurities and the drugs. It is important to strictly maintain the product safety and quality to ensure the regulatory compliance. An attempt has been made to develop and validate a new simple, single-run RP-HPLC method for the simultaneous estimation of seven known and several unknown impurities from fluticasone furoate and oxymetazoline hydrochloride nasal spray products. The separation of multiple impurity peaks was achieved by using a 5 µm, Inertsil ODS 3 V, 250 × 4.6 mm column. Dual-wavelength UV detector (220 nm and 240 nm) was used for the detection of oxymetazoline hydrochloride, fluticasone furoate and their impurities. The relative response factor was calculated for all the known impurities to make the method accurate, simple and economical. The developed method was validated following the guidelines outlined in the ICH Q2(R2). The forced degradation study results confirmed the stability-indicating nature of the method. The developed impurity profiling methodology will greatly help the quality control labs in maintaining the safety and efficacy of finished products and to comply with the regulatory requirements of various countries. Overall, the proposed method offers unparalleled precision, accuracy, sensitivity, and cost-effectiveness making it a valuable addition to the pharmaceutical industry.