HPLC Method Research Articles

Two-dimensional hydrophilic imprinted resin–graphene oxide composite for selective extraction and rapid determination of gibberellin traces in licorice samples

This study presents novel methodologies and materials for selectively and sensitively determining gibberellin traces in licorice to address food safety concerns. A novel hydrophilic imprinted resin–graphene oxide composite (HMIR–GO) was developed with fast mass transfer, high adsorption capacity, and exceptional aqueous recognition performance for gibberellin. Leveraging the advantages of molecular imprinting, hydrophilic resin synthesis, and rapid mass transfer characteristics of GO, HMIR–GO was employed as an adsorbent, showing resistance to matrix interference. Coupled with HPLC, a rapid and selective method for determining gibberellin was established. Under optimal conditions, the method exhibited a wide linear range (0.02–5.00 μg g−1, r = 0.9999), low detection limits (3.3 ng g−1), and satisfactory recoveries (92.0–98.4%), enabling the accurate and rapid detection of gibberellin in licorice. This study introduces a pioneering strategy for the selective extraction and determination of trace gibberellin levels, offering insights for similar applications in functional foods.

Development of Attenuated Total Reflectance Fourier Transform Infrared Spectroscopy Coupled with Multivariate Classification Chemometric Model for Routine Screening of Paracetamol, Ibuprofen, and Aspirin Adulteration in Herbal Products

Objective: The objective of this study is to develop and validate a routine screening test for the determination of three common antipyretic-analgesic synthetic drugs (paracetamol, ibuprofen, and aspirin) adulteration in herbal products using Attenuated Total Reflectance Fourier Transform Infrared (ATR-FTIR) coupled with chemometric method. Method: ATR-FTIR spectra of sixteen testing sets of herbal product samples for pain and fever indications were used for multivariate chemometrics model construction. Linear Discriminant Analysis (LDA) was selected as a method for model construction with IBM SPSS for statistical analysis. Model development employed feature selection, such as the stepwise method for variable selection. The model with a high %correct classification and cross-validation was selected and was then validated with an independent testing data set with an auto-prediction test, confusion matrix, and Receiver Operating Characteristic (ROC) curve. To validate the developed test for routine use, the result from ATR-FTIR method was compared with the standard HPLC and TLC analyses used for adulteration screening. objective: Creating validated screening tools for herbal products adulterated with three common antipyretic-analgesic synthetic drugs (Paracetamol, Ibuprofen, and Aspirin) using ATR-FTIR couple with chemometric method Results: The selected model's overall %correct classification result was 97.7%, with a cross-validation of 93.8% rate in training set samples. External validation with an independent testing dataset gave an overall correct classification of 93.8%, with an area under the curve of ROC at 0.979. Comparative testing revealed that model performance was comparable with the HPLC and TLC methods, which routinely detect the presence of paracetamol, aspirin, and ibuprofen. The results of testing set samples classification were consistent with training set samples. Conclusion: Against the standard chromatographic methods, the multivariate chemometric model based on ATR-FTIR demonstrates comparable detection capability to determine adulteration of paracetamol, ibuprofen, and aspirin in herbal products.

Two chromatographic methods for determination of cytarabine and dexamethazone; two co-administered drugs for the treatment of leukemia: green analytical perspective

Two sensitive, straightforward and repeatable chromatographic techniques were developed for the determination of Cytarabine HCl and Dexamethazone in their pure form and spiked human plasma without prior separation. The drugs are used co-administered for the treatment of Leukemia, a certain type of blood cancer. Method (A) is an isocratic chromatographic HPLC method; separation was accomplished on C18 column using the eluting mixture of 6.9 g/L Monobasic Sodium Phosphate pH 3: methanol (70:30, v/v) and detection was at 275 nm. Concentrations were in the range of 0.2–15 μg/mL for both CYT and DEX. Method (B) is a HPTLC method in which separation was attained on HPTLC F254 plates using methanol: ethyl acetate: ammonia, (7.8:2:0.2, by volume) as eluting solvents and detection was at 275 nm. Concentrations were in the range of 0.1–4 μg/band for both CYT and DEX. The parameters for system suitability testing were evaluated to determine the effectiveness of the developed chromatographic procedures in terms of performance. The recently developed techniques were applied for the determination of the drugs under investigation in spiked human plasma. Validation parameters were examined in accordance with US-FDA criteria. All results were found to be within the acceptable ranges. To evaluate the greenness characters of the proposed methods to the environment; three greenness assessment tools including eco-scale assessments (ESA), green analytical procedure index (GAPI), and Analytical Greenness calculator (AGREE) were used. Acceptable and satisfying results that demonstrated the greenness characteristics of the suggested methods were attained.

Quantitative Analysis of Swertiamarin Content from Fagraea fragrans Leaf Extract using HPLC Technique and its Correlation to Antibacterial Activity

The rise in drug resistance poses escalating challenges for antibacterial medications, leading to an urgent demand for the exploration and innovation of new antibacterial drugs. Fagraea fragrans Roxb., belonging to the Gentianaceae family, is one of the common herbal medicines which can be found abundantly in Southeast Asia. The secoiridoid glucoside swertiamarin, one of the major compounds in F. fragrans leaf, exhibits antimicrobial effects. To guarantee the medicinal effectiveness of F. fragrans leaves, it is essential to identify a standardized analytical method for quantifying the active compound. In this study, the optimized HPLC method following ICH guideline was validated for the quantitative analysis of swertiamarin content in F. fragrans leaf in terms of linearity (y = 5733.5x - 369.1; R2 = 0.9999), accuracy (93.57-96.39% recovery), precision (0.91% RSD for repeatability precision; 1.19% RSD for intermediate precision), limit of detection (0.73 µg/mL), limit of quantitation (2.23 µg/mL), specificity (peak purity index = 0.999995) and robustness (% RSD <1).The maximum wavelength of swertiamarin was found to be at 238 nm. The amount of swertiamarin content in F. fragrans leaf extract conducted from the validated HPLC method was found to be 0.0259 ± 0.0005 g/100 g crude drug. The leaf extract exhibited antimicrobial activity against clinical isolates of Enterobacter cloacae, Klebsiella pneumonia, and Escherichia coli at 0.125 mg/mL, and Pseudomonas aeruginosa at 0.5, showing minimum inhibitory concentration (MIC) values. Whereas swertiamarin exhibited even lower MIC values. The developed HPLC analysis effectively determines swertiamarin content as a chemical marker to ensure the antimicrobial potential of F. fragrans leaves.

A stability-indicating HPLC assay of ten different vitamins in a food supplement: appraisal of the method’s greenness, whiteness, and blueness

Due to their susceptibility to degradation, vitamin levels in food formulations may differ from those found in the finished product. Vitamin levels can be impacted by processing and storage. In this work, the ingredients of Strong B50 ® film-coated tablets were estimated simultaneously using simple efficient stability indicating HPLC method. Strong B50 ® film-coated tablets contain thiamine (VB1), riboflavin (VB2), calcium pantothenate (VB5), pyridoxine (VB6), vitamin C (VC), folic acid (FA), biotin (BT), inositol (IS), niacin (NC), para-aminobenzoic acid (PB), cyanocobalamine (B12), choline bitartarate, and iron gluconate. Hypersil BDS C18 column was used for achieving reasonable separation. Mobile phases (A) and (B) were utilized, the mobile phase (A) consisted of 0.015M Hexane sulfonic acid sodium salt + 0.1% Triethylamine and orthophosphoric acid was used to adjust the pH to (2.9) while (B) system consisted of acetonitrile. Validation of the method was assessed using International Conference of Harmonization (ICH) parameters, where linearity, accuracy, selectivity, and robustness of the method were investigated. Correlations were above 0.99, accuracy results ranged from 97.6 to 102.8 % and limits for detection and quantitation (LOD and LOQ) values were determined for each vitamin in μg/mL except for FA and BT in ng/mL. LOD values were between 0.006 and 15.08 μg/mL while LOQ values ranged from 0.031 to 49.77 μg/mL. Stability studies were conducted under stressed conditions and degradation percentages were computed. Where, VB5, VB6, FA and PB, VC, and NC were the most degradable vitamins. Whiteness evaluation using the modern RGB 12 algorithm compared our method and the old reported one by Sasaki et.al, 2020. The comparison favored our newly developed method in terms of analytical performance, practical applicability and greenness. Besides, AGREE and GAPI soft wares were used to assess the greenness of the method. It was clear that the results of colored pictograms confirm low hazardous impact and that the new method is greener with AGREE score of 0.66. Furthermore, the functionality and applicability of the novel HPLC approach were concluded via the Blue Applicability Grade Index (BAGI) tool with a final score of 82.5.

Pharmacokinetics of tildipirosin in estuarine (Crocodylus porosus) and freshwater (Crocodylus siamensis) crocodiles

Tildipirosin is a macrolide antimicrobial. It is authorised for the treatment and prevention of respiratory disease in cattle and pigs. There are no data on its administration in crocodiles. Therefore, this study evaluated the disposition kinetics of tildipirosin after intravenous (dose: 2mg/kg) and intramuscular (doses: 2 and 4mg/kg) administration in two crocodilian species (estuarine and freshwater; (n = 5). Tildipirosin plasma concentrations were quantified by a validated HPLC method. Plasma concentrations obtained at each extraction time were analysed by non-compartmental methods.In the estuarine and freshwater crocodiles, the apparent volumes of distribution of tildipirosin after intravenous administration were 0.36 ± 0.10 and 1.48 ± 0.26L/kg, respectively. These values, suggesting poorer tissue distribution, were much lower than those obtained in mammals. There was complete bioavailability of tildipirosin after intramuscular route at a dose of 2mg/kg; however, at a dose of 4mg/kg the bioavailability decreased by about 20–25%. Furthermore, the pharmacokinetics of tildipirosin were markedly different in the two crocodilian species. Considering a MIC of 0.5µg/mL, the surrogate marker AUC0→24/MIC indicates that TD would greatly exceed the value of 65h for both crocodile species and dose levels tested. This suggests that both doses (2 and 4mg/kg) may provide a bactericidal effect. Therefore, based on the absence of adverse reactions following the administration of tildipirosin in both crocodilian species, and considering its favourable pharmacokinetic properties, tildipirosin may be useful in treating infections in these reptiles.

Evaluation and comparison of two methods for quantification of HbA1c with the reference HPLC method

Evaluation and comparison of two methods for quantification of HbA1c with the reference HPLC method

An electrochemical sensing potential of cobalt oxide nanoparticles towards citric acid integrated with computational approach in food and biological media

Although citric acid (CA) has antioxidant, antibacterial, and acidulating properties, chronic ingestion of CA can cause urolithiasis, hypocalcemia, and duodenal cancer, emphasizing the need for early detection. There are very few documented electrochemical-based sensing methods for CA detection due to the challenging behavior of electrode fouling caused by reactive oxidation products. In this study, a novel, non-enzymatic, and economical electrochemical sensor based on cobalt oxide nanoparticles (CoOxNPs) is successfully reported for detection CA. The CoOxNPs were synthesized through a simple thermal decomposition method and characterized by SEM, FT-IR, EDX, and XRD techniques. The proposed sensing platform was optimized by various parameters, including pH (7.0), time (15 min), concentration of nanoparticles (100 mM), and analyte, etc. In a linear range of 0.05–2500 μM, a low detection limit (LOD) of 0.13 μM was achieved. Theoretical calculations (ΔRT), confirmed hydrogen bonding and electrostatic interactions between CoOxNPs and CA. The detection method exhibited high selectivity in real media like food and biological samples, with good recovery values when compared favorably to the HPLC method. To facilitate effective on-site investigation, such a sensing platform can be assembled into a portable device.

Nanolevel of detection of ascorbic acid using horse-radish peroxidase inhibition assay

Ascorbic acid plays a significant role in regulation of various bodily functions with high concentrations in immune cells and being involved in connective tissue maintenance. Commonly it is detected through various colorimetric methods. In this study, we propose a one-step simple method based on the inhibitory activity of ascorbic acid on horseradish peroxidase and hydrogen peroxide. The detection is observed by colorimetric changes to TMB (3,3′,5,5′ tetramethylbenzidine). The enzyme inhibition unit was optimized with a high level of linearity (r2 = 0.9999) and the level of detection and level of quantification were found to be 1.35 nM and 4.08 nM, respectively with higher sensitive compared to the HPLC method (11 μM). Both intra and inter-assays showed high correlations at different AA concentrations. (r2 > 0.9999). Similar results were also observed for vitamin C tablets, ascorbate salts, fruits, and market products (r2 = 0.999). There was negligible effect of interference by citric acid, lactic acid, tartaric acids, and glucose with high recoveries (>98%) at 1 mg/mL to 0.0078 mg/mL concentration ranges. The recovery error (RE%) was found to be less than 10%. Our detection method is distinguished by its simplicity, nano-level of detection, reproducibility, and potential application and adaptability as a point-of-use test.

HPLC with chiral stationary phase for separation and kinetics study of aspartic acid epimerization in Peroxiredoxin 2 active site peptide

Amino acid epimerization, a process of converting L-amino acids to D-amino acids, will lead to modification in the protein structure and, subsequently, its biological function. This modification causes no change in protein m/z and may be overlooked during protein analysis. Aspartic Acid Epimerization (AAE) is faster than other amino acids and could be accelerated by free radicals and peroxides. In this work, a novel and site-specific HPLC method using a chiral stationary phase for determining the AAE in the active site model peptide (AP) of Peroxiredoxin 2 has been developed and validated. The developed method showed good linearity (1 – 200 μg/mL) and recoveries of the limit of quantification (LOQ), low, medium, and high concentrations were between 85% and 115%. The Kinetics of AAE in AP were studied using the developed method, and the results showed that when ascorbic acid and Cu2+ coexisted, the AP epimerized rapidly. The AAE extent increased with time and was positively correlated with hydrogen peroxide generation.

Effect of chemical penetration enhancers on the transdermal delivery of olanzapine in human skin in vitro

This research study involves the development of an olanzapine (OLZ) formulation using various chemical penetration enhancers (CPEs) for transdermal delivery. The aim of this study was to obtain the initial data needed about the effects of various CPEs on the skin permeation of OLZ. The effects of the selected CPEs were examined by studying the permeation profiles of OLZ from formulations applied to human cadaver skin samples. A control formulation of OLZ in propylene glycol (PG) was prepared and compared against formulations containing chemical penetration enhancers. Five different CPEs (oleic acid (OA), cineole (Cin), isopropyl alcohol (IPA), Tween 80 (T80), and N-methyl pyrrolidone (NMP)) at 5% w/w were individually added to the formulation containing OLZ in PG. The in vitro permeation study was carried out using vertical Franz diffusion cells mounted with human cadaver skin. Samples from the receptor compartment of the cell were collected at 2 h, 4 h, 8 h, 12 h, and 24 h at room temperature. The amount (µg/cm2) of permeated drug (OLZ) was measured using a validated HPLC method, and the percentage (%) of OLZ permeated was calculated. Based on the data obtained, different CPEs were found to have a significant impact on OLZ permeability compared to the control formulation. The most effective chemical penetration enhancer was shown to be 5% w/w OA with a 3.3-fold increase in enhancement ratio (ER). The rank of order for the highest concentration of OLZ permeated from each of CPE containing formulation was as follows: OA > Cin > IPA > T80 > NMP. The most effective chemical penetration enhancer was OA but the cytotoxic study using human fibroblast cells suggests that OA may not be safe due to its cytotoxic effects.

Rapid determination of hippuric acid as an exposure biomarker of toluene using a colorimetric assay and comparison with high-performance liquid chromatography

Occupational exposure to toluene is associated with health risks that require reliable monitoring methods. Hippuric acid (HA), a urinary metabolite of toluene, serves as a valuable biomarker for such exposure. Colorimetric methods for the quantitative determination of HA have gained prominence due to their simplicity, cost-effectiveness, and suitability for field application. In the present study, a simple colorimetric technique was optimized for the determination of HA in the urine sample, and compared with a usual HPLC technique. The central composite design (CCD) was applied to examine the effective parameters on the colorimetric determination of HA. The calibration curve for HA was established within the concentration range of 6 to 100 mg L−1 with R2 = 0.97. The detection limit (LOD) and quantification limit (LOQ) were determined to be 1.8 mg L−1 and 6 mg L−1 respectively. The relative standard deviation (RSD%) was less than 5%, and the recovery% (R%) was 90.5–100.1. The overall results showed good agreement between the colorimetric and HPLC results. There was a significant relationship between the results obtained from HPLC and colorimetric methods especially for higher concentration levels of HA (≥ 500 mg/g creatinine). In conclusion, our optimized colorimetric method is a simple, cost-effective, and rapid method for determination of HA in occupational exposure, which is comparable with the HPLC technique.

A new and rapid HPLC method for the determination of phenoxyethanol in topical formulation

A new and rapid HPLC method for the determination of phenoxyethanol in topical formulation

Rapid Quantitation of Non-chromophoric Vigabatrin and Gabapentin by a Validated qNMR Method in Bulk Drug and Marketed Formulations

Background: Vigabatrin and gabapentin, commonly used antiepileptic drugs in clinics, lack a UV active chromophore and, therefore, require cumbersome derivatization methods for analysis by HPLC using fluorescence detection. This study demonstrated the use of NMR for their quantitative determination in pure form and their pharmaceutical formulations. Objective: To develop a validated qNMR method for non-chromophoric drugs Vigabatrin and Gabapentin. Methods: The signal of methine proton of vigabatrin at 3.67 ppm relative to the signal of maleic acid at 6.17 ppm and the methylene signal of gabapentin at 2.88 ppm relative to the signal of caffeine at 7.75 ppm was used for qNMR. The developed method was validated with respect to linearity, limits of detection and quantitation, accuracy, precision, specificity and solution state stability. Results: Linearity range and r2 were found to be from 2.66 to 42.11 mg/mL and 0.9999. The limit of detection and quantification were 0.0129 mg/mL and 0.0391 mg/mL, respectively, for vigabatrin. This method was found to be linear (0.9998) and specific within the gabapentin concentration range from 1.07 to 34.24 mg/mL of D2O. The limits of detection and quantification were 0.0248 mg/mL and 0.0751 mg/mL, respectively. Conclusion: Both methods were highly precise, with a calculated RSD of 0.60% and 0.76%, respectively. The robustness of the methods was revealed by changing pre and post-processing NMR parameters. The developed methods provide a simple and straight approach for the absolute determination of gabapentin and vigabatrin in bulk drugs and their marketed formulations without any pre-procedures.

Development and validation of a quantitative minilab system for quality evaluation of selected medicines: Albendazole, arthemether-lumefantrine combination (Co-artem®), and mebendazole finished pharmaceutical products

AbstractThe current technologies for substandard and counterfeit drug detection are either too expensive for low-resource settings or only provide qualitative or semi-quantitative results. GPHF minilab™ is one of them based on thin layer chromatography(TLC) principles with a semi-quantitative capability by visual observation of the spot area and intensity for medicine quality analysis. Thus, its use as a quality control tool for pharmaceutical products has limitations as spot area and intensity visual observation by the naked eye highly varies from analyst to analyst. As such, in this study, the semi-quantitative technique has been transferred to a quantitative approach by capturing the developed TLC plate image using an Android-based mobile phone inside a simple carton box. Then, the spot area was quantified using justTLC software. The quantitative results were compared with the-high performance liquid chromatography (HPLC) method as the golden standard. Accordingly, linearity was observed in the assayed range (80–120% label claim), and the correlation coefficients found were (R2 = 0.958, 0.997, 0.941, and 0.956 for Albendazole, Mebendazole, Artemether, and Lumefantrine, respectively.). The values are satisfactory. The %RSDs found were less than 2% for all drugs [intraday (n = 6) (RSD = 1.17, 1.61, 1.87, and 1.64), and interday (n = 18) (RSD = 1.16, 0.72, 1.12, and 1.18) for Artemether, Lumefantrine, Mebendazole, and Albendazole, respectively]. Moreover, comparisons of results obtained from the sophisticated CAMAG UV cabinet (R2 =0.991, 0.971, 0.946, and 0.967) and the developed simple carton box (R2 = 0.958, 0.997, 0.941, and 0.956) for Albendazole, Mebendazole, Artemether, and Lumefantrine, respectively. The values are comparable and reveal the accuracy of the method. Robustness testings' that were performed under different altered conditions revealed the robustness of the method (RSD less than 2% for all factors). Additionally the deviations from the golden HPLC results were on average −8.62% for albendazole, −3.79% for artemether, and −4.52% for lumefantrine samples. The developed method shows a satisfactory performance capability to utilize the GPHF minilab™ as a quantitative technique for medicine quality control purposes. It will be a very useful tool in a resource-limited setting. The target method profile, which encompasses a simple, low-cost, linear, precise, robust, accurate, and quantitative GPHF minilab™ system, was obtained for Albendazole, Mebendazole, and Arthemeter lumefantine combinations (Co-artem). The proposed method was successfully applied to analyze the content of the marketed medicines in the above mentioned tablets and offered acceptable deviations from the golden HPLC method. Automation of quantitative GPHF minilab™ was highly recommended to enhance the appropriateness and use of this system.

Development of a Simultaneous Normal-Phase HPLC Analysis of Lignans, Tocopherols, Phytosterols, and Squalene in Sesame Oil Samples.

The objective of this study was to develop a simultaneous analytical method for the determination of lignans, tocols, phytosterols, and squalene using high-performance liquid chromatography coupled with a diode array and fluorescence detector (HPLC-DAD-FLD). The method employed a VertisepTM UPS silica HPLC column (4.6 × 250 mm, 5 µm) with a mobile phase mixture of n-hexane/tetrahydrofuran/2-propanol. This approach enabled the simultaneous analysis of ten compounds within 22 min. The linear correlation (R2) exceeded 0.9901. The limit of detection (LOD) and limit of quantitation (LOQ) were up to 0.43 µg mL-1 for lignans and tocopherols and up to 326.23 µg mL-1 for phytosterol and squalene. The precision and accuracy of the intra-day and inter-day variation were less than 1.09 and 3.32% relative standard deviations (RSDs). Furthermore, the developed method was applied for the analysis of targeted compounds in twenty-eight sesame oil samples (1775-8965 µg g-1 total lignans, 29.7-687.9 µg g-1 total tocopherols, 2640-9500 µg g-1 phytosterol, and 245-4030 µg g-1 squalene). The HPLC method that has been developed was proven to be a reliable and effective tool for the determination of those functional compounds among sesame oil samples.

Development and characterisation of buccal dispersible film, syrup and effervescent granules of extemporaneous antigout remedy

Homemade medicinal remedies used to treat both acute and chronic ailments for centuries need to be transformed into modern evidence-based dosage forms. Therefore, the present study aimed to develop buccal dispersible film, syrup and effervescent granules of an extemporaneous antigout homemade remedy. The dosage forms were prepared using holistic remedy (HR) as an active entity and excipients masking disagreeable features. The dosage forms and HR were subjected to physicochemical evaluation, and standardised by a reversed-phase HPLC method using chlorogenic acid, caffeic acid, vanillin, and ferulic acid as analytical standards. The standardised dosage forms were investigated for antigout activity using an appropriate model. The dosage forms were found to be physically elegant and phytochemically aligned with HR. The developed dosage forms contained the selected markers and possessed antigout activity. Hence, these standardised and evidence-based dosage forms may have wider acceptance in the market compared to HR.

Chemical fingerprinting and multicomponent quantitative analysis for quality control of Cinnamomum tamala collected from Western Himalaya by HPLC-DAD

Cinnamomum tamala, commonly known as “Indian bay leaf” or “Tejpat”, is an economically important plant widely used in medicine, food and cosmetic industries. Growing demand for its leaf and bark in the herbal trade and non-availability of quality materials lead to large-scale species admixture and adulteration in the global market. The present study aims at developing a validated HPLC-DAD (High-performance liquid chromatography coupled with diode array detection) method and multiple markers-based chemical fingerprints for quality evaluation of C. tamala leaf extracts. Five bioactive compounds, viz., coumarin, cinnamyl alcohol, cinnamic acid, cinnamaldehyde and cinnamyl acetate, were identified and quantified in 28 samples collected from the western Himalayan region of India. The chromatographic separation was achieved on Shimadzu Shimpak C18 column (dimension 250 × 4.6 mm, pore size 5 μm) with a gradient elution of mobile phase using acetonitrile and 0.1 percent phosphate buffer and the chromatograms were obtained at a wavelength of 265 nm. The method validation was done by analyzing the linearity, LOD, LOQ, precision, stability, repeatability and recovery rates of standard compounds for quantitative analysis. The values of coefficient of correlation (R2) were found to be close to 1 for linearity and similarity analysis; and standard deviation was less than 3 percent in case of precision, stability, repeatability and recovery rates. The content of target compounds such as coumarin, cinnamyl alcohol, cinnamic acid, cinnamaldehyde and cinnamyl acetate varied in the range of 0–1.09, 0–0.05, 0.07–0.51, 0.39–1.27 and 0–0.27 percent, respectively. In the chemical fingerprint of C. tamala leaves, a total of 13 peaks were assigned as common peaks. The results of the study indicated that the HPLC method now developed combining chemical fingerprint with quantification of analytes could serve as a useful tool for quality evaluation of herbal raw materials of C. tamala and a valuable reference for further study.

LC and LC-MS/MS Studies for Identification and Characterisation of Related Substances and Degradation Products of Abrocitinib

In the pharmaceutical industry, Related Substances (RCs), impurities or Degradation Products (DPs) are associated with the Active Pharmaceutical Ingredient (API) in the final drug product. These compounds must be within permissible limits for safe therapeutic use for consumers. Hence there is a need to quantify these compounds using an appropriate analytical method. No method is reported in the literature for quantification of these compounds in abrocitinib which is a medical drug prescribed to treat severe atopic dermatitis in adults. This study includes the optimisation of stability indicating the HPLC method for resolution and subsequent quantification of abrocitinib RCs and structural characterisation of stressinduced DPs of abrocitinib. The method was optimised by varying mobile phase solvents, pH, flow rate and wavelength of the detector. The finalised conditions were validated and applied for the resolution and evaluation of stress-induced DPs. The stress was induced in abrocitinib pure drug HCl (0.1M), NaOH (0.1M), hydrogen peroxide (3%), 80°C in an air oven and 254nm in a UV chamber. The generated DPs were structurally characterised with the LCMS experiment. Abrocitinib and DPs along with known RCs were resolved on ACE Ultra Core Super C18 250mm column using 0.9mL/min gradient flow of methanol (Solvent A), acetonitrile and buffer (Solvent B). The resolved compounds were detected through a UV detector at 295nm and a mass detector at NMR positive mode. The method identifies 5.85min, 3.13min, 6.60min and 4.38min respectively for abrocitinib, related compounds 1 to 3 respectively with acceptable system suitability. A very high correlate (< 0.999) linear graph was achieved within 5 to 30 μg/mL concentration level for abrocitinib and 0.05 to 0.3μg/ mL for related compounds. Three DPs with molecular mass and formula of C10H13N5 (203.2 g/mol), C11H14N4 (202.2 g/mol) and C13H19N5O2S (309.3 g/mol) in acid-induced stress study and two DPs of C12H21N5O2S (299.3 g/mol) and C11H19N5O2S (285.3 g/mol) in peroxide-induced stress study and one DP of C10H14N4 (190.2 g/mol) in base induced stress study were successfully characterised. The method proposed in this study can effectively resolve unknown degradation products, and known related compounds along with abrocitinib and is hence applicable for quality control analysis of abrocitinib.

High-efficient selection of aptamers by magnetic cross-linking precipitation and development of aptasensor for 1-aminohydantoin detection

Accurate and rapid detection of nitrofurantoin metabolite 1-aminohydantoin (AHD), as one of the tissue-bound toxic metabolites in foods, is crucial. In this study, a fluorescent aptasensor was constructed for the detection of AHD, using anti-AHD aptamers firstly obtained by high-efficiency magnetic crosslinking precipitation (MCP)-SELEX. The AHD-specific aptamers were optimized and identified for their binding affinity and specificity using fluorescence-based assays and molecular dynamics simulations. The truncated aptamer AHD50-1, which exhibited highest affinity (Kd = 86 nM) and an excellent specificity, was applied to the construction of fluorescent aptasensor. Under optimized conditions, the aptasensor showed a low limit of detection (0.2 ng/mL) and a wide dynamic linear range (0.25–150 ng/mL). For the five types of samples (pork, beef, shrimp, fish and honey), the recoveries ranged from 89.4% to 110.04%, with coefficients of variation ranging from 1.43% to 14.46%. These results obtained by the proposed aptasensor corroborate very well with the method of HPLC for detection of AHD, indicating that the developed aptasensor provides an efficient and promising sustainable strategy for monitoring AHD in animal derived food.