Development of urinary assay methods for estimation of paracetamol glucuronide and paracetamol sulphate in preterm neonates with patent ductus arteriosus.

Aims: This study aimed to develop a high-performance liquid chromatography (HPLC) technique for estimating paracetamol glucuronide and paracetamol sulphate in the urine samples of preterm neonates. Background: Validated methods exist for estimating the principal metabolites of paracetamol in older children and those with liver disease. Here, we have developed and validated a simple technique for estimating the same in urine samples of preterm neonates. Objective: The study aims to develop and validate a simple, reliable, and accurate HPLC technique for estimating urinary paracetamol glucuronide and paracetamol sulphate metabolites. Methods: Preterm neonates of either sex diagnosed with patent ductus arteriosus (PDA) receiving paracetamol intravenously at the dose of 15 mg/kg every six hours were recruited. We ran the samples under standardized chromatographic conditions and using various dilutions of the calibration standards. Measures of assay selectivity, linearity, accuracy, and precision were estimated. Results: We observed that the peaks for paracetamol glucuronide and paracetamol sulphate were distinguished from those of the drug-free urine samples. The results for both metabolites revealed good reproducibility, with a percent coefficient of variation (% CV) of 4.3 and 4.9 for the slope for paracetamol glucuronide and paracetamol sulphate, respectively. Similarly, we observed good linearity, as indicated by the correlation coefficients of 0.99 for the metabolites. The validation assays revealed that the method is linear, accurate, and precise over the defined concentration ranges. Conclusion: We demonstrated that HPLC has good accuracy, reliability, and precision, and it can be used for estimating the principal metabolites from urine samples in neonates for defining the ontogeny of conjugation enzymes and in paracetamol overdose.

Rapid quantification of Psilocybin with reversed-phase HPLC and single-wavelength detection

The alkaloid psilocybin (4-phosphoryloxy-N,N-dimethyltryptamine) and the neurologically active psilocin (4-hydroxy-N,N-dimethyltryptamine) are the foremost compounds of pharmaceutical interest in Psilocybe mushrooms. As these compounds are infrequently analyzed in analytical labs, validated methods for rapid purity analysis are lacking. Newfound therapeutic use has invigorated academic and commercial interests in the molecules and new methods of production and available products are expanding. As a result, high-throughput methods of analysis for psilocybin must be improved to promptly determine chemical differences between mushroom genera or other sources of psilocybin and psilocin, as well as refined product purity. To address this, we developed an inexpensive HPLC technique for the efficient quantification of psilocybin and psilocin by using readily available equipment and dilute reagents. Aqueous ammonium formate (0.143 mM) was found to be preferable over techniques with much higher buffer concentrations or stronger acids for controlling psilocybin Zwitterion resolution. The chromatographic run time satisfied high-throughput analytical requirements with an efficient total runtime under 2 minutes. A standard octadecyl silica (C18) column provided excellent resolution between psilocybin and psilocin signals. The quality of the method was validated using certified analytical reference standards and was found to be accurate (3.5% bias, Psilocybin), reliable (0.32% RSD), and efficient (Psilocybin k’ = 1.78).

Analysis of Biogenic Amines Using Immunoassays, HPLC, and a Newly Developed IC-MS/MS Technique in Fish Products—A Comparative Study

In this study, comparative analyses were carried out with ion chromatography mass-spectrometry (IC-MS/MS) which has no derivatization step, high-performance liquid chromatography (HPLC) technique, as well as two quantitative and two semi-quantitative immunoassays. The results demonstrated that HPLC and quantitative immunoassay methods were well-correlated with IC-MS/MS in determining histamine in various types of fish products. The best correlation was observed with the HistaSure ELISA Fast Track kit (R2 = 0.9903). More than half of the values (68%) obtained by two methods were also statistically similar. The results of semi-quantitative test kits also supported histamine values estimated by quantitative methods, with some exceptions. The best results were found for HistaSure Lateral Flow in supporting the quantitative techniques. Therefore, these methods are found suitable for monitoring histamine in fish products in terms of food safety. Good correlations were also observed HPLC and IC-MS/MS in determining cadaverine, putrescine, and tyramine with the highest value observed for tyramine as R2 = 0.9785. However, no correlation was observed for other biogenic amines, and the majority of the results were significantly different from each other for these amines (p < 0.05). The differences may be caused by the drawbacks reported previously for HPLC. However, further studies are required to confirm the possible effects. This study provides a comparative evaluation of several methods in terms of their suitability in determining biogenic amines in fish products for both monitoring and regulatory purposes.

A Validated Reversed-Phase HPLC Analytical Method for the Analysis of Fenofibrate in Bulk Drug and Tablet Dosage Formulation

Aims: A accurate, precise, and stability-indicating Reversed-Phase HPLC technique has been established for the estimation of fenofibrate in tablet formulation. Study Design: Experimental study. Place and Duration of Study: Department of Pharmaceutical Sciences, RTM Nagpur University, Nagpur-440033, Maharashtra, India between June 2019 and March 2020. Methodology: The chromatographic separation was attained on RP Princeton column (C18) (250 mm x 4.6 mm, 5 µ) with mobile solvent system as a mixture of water (pH 3.0 along o-phosphoric acid) and acetonitrile in the proportion (40:60) v/v, flow rate 1.0 ml per minute, at 240 nm. The retention time of fenofibrate was 3.905 minutes. Results: The method demonstrated linearity in the concentration range of 87-232 µg/ml with a coefficient of correlation (r2) of 0.9994. The % RSD was ˂2% and percentage recovery was found to be 99.13-100.74%. The assay of marketed tablet formulations was found to be 99.98%. Conclusion: The developed and validated technique as per ICH rules for specificity, accuracy, precision, linearity, and system suitability. Reverse Phase-HPLC technique was utilized to the market formulation.

Development and Validation of a Stability-Indicating HPLC Technique for Measuring Temozolomide in its Pharmaceutical Dose Form

Using RP-HPLC, an accurate and precise technique for the measurement of Temozolomide in its pharmaceutical dose form was developed and validated. Chromatographic separation was achieved on an X Terra RP 18(250mm x 4.6mm), 5 µ column using a mobile phase consisting of methanol and buffer in the ratio of 10:990v/v. The flow rate was 1mL/min with the detection wavelength of 254 nm and retention time was found to be 20 min. The developed method was validated according to ICH guidelines. With a correlation coefficient of 0.9990, linearity was observed in the range of 50-150 percent. The %RSD of the developed method for method precision and Intermediate precision was found to be 0.65 % and 0.59 % respectively. With a percent recovery of 99.82 ±0.045, the approach was confirmed to be reliable. All of the validation parameters yielded results that were within acceptable limits. It was discovered that the procedure was accurate, exact, specific, rugged, and durable. As a result, the newly discovered approach can be used for finish product of quality control and stability testing on a regular basis and it has been confirmed to be stable for Temozolomide is available in both pure and pharmaceutical dose forms.

Identification and Quantification of Soil Pesticides in Coastal Lakshmipur District of Bangladesh

This study was carried out to determine the presence and quantity of some selected pesticides from soil sediments collected from some ponds and canals located in the Lakshmipur district of Bangladesh. The high performance liquid chromatography (HPLC) technique was used to determine the concentration of pesticide residues. Some soil samples were found to be contaminated with carbamate (carbofuran and carbaryl) and organophosphorus (diazinon) pesticides. The concentration of carbofuran pesticide ranged from 0.303 μg/kg to 1.851 μg/kg. The highest concentration of carbofuran pesticide was found in SSP6 (1.851 μg/kg) and the lowest concentration was found in SSP9 (0.303 μg/kg). Carbaryl pesticide was found to be present in the sediment of only one pond, the concentration being 1.047 μg/kg. Organophosphorus (diazinon) pesticide was found in soil samples and the concentrations ranged from 0.147 μg/kg to 0.759 μg/kg, which were higher than the EEC-recommended limit of 0.1 μg/kg. Asiat. Soc. Bangladesh, Sci. 46(2): 191-200, December 2020

Method comparison of Particle Enhanced Immunoturbidimetry (PEIT) with High Performance Liquid Chromatography (HPLC) for glycated hemoglobin (HbA1c) analysis

Background and objective High Performance Liquid Chromatography (HPLC) technique is considered as a gold standard for HbA1c analysis however all laboratories cannot adopt it due to certain limitations. Our aim was to compare Particle Enhanced Immunoturbidimetry (PEIT) method with High Performance Liquid Chromatography (HPLC) for HbA1c analysis. Method All blood samples were analyzed by HPLC assay on a Bio-Rad D-10 analyzer and PEIT on an Erba XL-200 analyzer. Precision studies were undertaken and Coefficient of Variation (%CV) calculated. Systemic Error (SE), Random Error (RE) and Total Error (TEcalc) were obtained. The Total Allowable Error (TEa) set by the National Glycohemoglobin Standardization Program (NGSP) for HbA1c is 6%.The acceptable evaluation method is where TEcalc is less than TEa. Results The Precision studies were satisfactory with Coefficient of Variation (%CV) being less than 4% for both techniques. Mean HbA1c levels were slightly higher from HPLC than PEIT 9.07 ± 2.23% and 8.93 ± 2.10% respectively, although the difference was minimal. RE was 1.41%, TEcalc was 1.55%, which was less than TEa set by the NGSP. Both methods strongly correlated with the correlation coefficient (r) 0.9716, p < 0.0001. Conclusion Our study showed HbA1c analysis by PEIT technique is precise, accurate, rapid and convenient and can be employed as an alternative to HPLC technique in countries where cost is a major problem for diagnosis and treatment.

Enrofloxacin and Ciprofloxacin Residues in Table Eggs: Distribution and Heat Treatment Effect

A 500 composite egg samples (2500 eggs-each sample represented by 5 eggs) collected from layer farms and local markets from all over Jordan were studied for presence of antimicrobials using Premi®Test screening test. Positive samples indicated by inhibition of microbial growth represented 12.8% out of total screened egg samples. Positive samples were examined quantitatively using HPLC technique to detect the presence of enrofloxacin and ciprofloxacin. Ciprofloxacin and enrofloxacin residues were detected in 1% and 0.8% of the total samples, respectively, where both drugs were recovered from white but not yolk. The effect of boiling on either drugs concentration in fortified white or yolk was demonstrated by gradual increases in the mean reduction percentages within treatment time with an average of 87% reduction after 15 minutes of boiling for both drugs and egg compartments and 5 minutes of frying at 160°C. The average concentration reduction percentages by the end of four weeks of refrigeration of fortified yolk and white were around 45 and 50% for enrofloxacin and ciprofloxacin respectively. The significance and mechanism of drug deposition and reduction during processing is being highlighted.

Evolution and effectiveness of HPLC Technique for rapid estimation of an Antiallergenic agent Bilastine

A new, simple, accurate, and specific RP-HPLC stability method for determining bilastine was developed and validated. The proposed method was administered using C18 BDS Hypersil thermo column (4.6 × 250mm i.d), 5 µm particle size with a combination of potassium dihydrogen phosphate buffer pH 6.0: acetonitrile: methanol (50:25:25) as the mobile phase at a wavelength of 220nm. The retention time was 3.9 min for bilastine. The calibration plot was linear over the concentration range of 14.4–33.6µg/ml bilastine with LOD and LOQ of 0.04 and 0.11µg/ml, respectively. The technique was validated for linearity, sensitivity, accuracy, precision, and robustness. Percent recoveries were observed to be nearly 100%. The validated method was used for determining bilastine in Pharmabilast(R) tablets. The technique could be appropriate for routine evaluation at laboratories.