This study discusses the development and validation of a universal microwell spectrophotometric assay for TKIs, regardless of the diversity in their chemical structures. The assay depends on directly ...measuring the native ultraviolet light (UV) absorption of TKIs. The assay was carried out using UV-transparent 96-microwell plates and the absorbance signals were measured by a microplate reader at 230 nm, at which all TKIs had light absorption. Beer's law correlating the absorbances of TKIs with their corresponding concentrations was obeyed in the range of 2-160 µg mL
with excellent correlation coefficients (0.9991-0.9997). The limits of detection and limits quantitation were in the ranges of 0.56-5.21 and 1.69-15.78 µg mL
, respectively. The proposed assay showed high precision as the values of the relative standard deviations for the intra- and inter-assay precisions did not exceed 2.03 and 2.14%, respectively. The accuracy of the assay was proven as the recovery values were in the range of 97.8-102.9% (±0.8-2.4%). The proposed assay was successfully applied to the quantitation of all TKIs in their pharmaceutical formulations (tablets) with reliable results in terms of high accuracy and precision. The assay greenness was evaluated, and the results proved that the assay fulfils the requirements of green analytical approach. The proposed assay is the first assay that can analyse all TKIs on a single assay system without chemical derivatization or modifications in the detection wavelength. In addition, the simple and simultaneous handling of a large number of samples as a batch using micro-volumes of samples gave the assay the advantage of high throughput analysis, which is a serious demand in the pharmaceutical industry.
In this study, a new green microwell spectrofluorimetric assay (MW-SFA) with high throughput was developed and validated, for the first time, for the determination of three selective serotonin ...reuptake inhibitors (SSRIs) in pharmaceutical dosage forms and plasma. These SSRIs were fluoxetine (FLX), fluvoxamine (FXM), and paroxetine (PXT), which are commonly prescribed drugs for depression treatment. The MW-SFA is based on the condensation reaction of SSRIs with 4-chloro-7-nitrobenzo-2-oxa-1,3-diazole (NBD-Cl) in alkaline media to form highly fluorescent derivatives. The MW-SFA procedures were conducted in 96-microwell white opaque assay plates with a flat bottom and the fluorescence signals were measured using a microplate reader at their maximum excitation and emission wavelengths. The calibration curves were generated with good correlation coefficients (0.9992-0.9995) between the relative fluorescence intensity (RFI) and the SSRI concentrations in the range of 35-800 ng/mL. The limits of detection were in the range of 11-25 ng/mL, and the precision and accuracy were satisfactory. The proposed MW-SFA was successfully applied to the analysis of the SSRIs in their pharmaceutical dosage forms. The statistical analysis for the comparison between the MW-SFA assay results and those of pharmacopeial assays showed no significant differences between the assays in terms of their accuracy and precision. The application of the proposed MW-SFA was extended to successfully analyze SSRIs in plasma samples. The greenness of the assay was confirmed using three different metric tools. The assay was characterized with high throughput properties, enabling the sensitive simultaneous analysis of many samples in a short time. This assay is valuable for rapid routine applications in pharmaceutical quality control units and clinical laboratories for the determination of SSRIs.
This study describes the development and validation of a new green and high-throughput microwell spectrophotometric assay (MW-SPA) for the determination of three selective serotonin reuptake ...inhibitors (SSRIs) in their pharmaceutical dosage forms. These SSRIs are fluoxetine, fluvoxamine, and paroxetine, the most prescribed drugs for the treatment of depression. The proposed assay was based on the formation of orange-colored
-substituted naphthoquinone derivatives upon the reaction of SSRIs with 1,2-naphthoquinone-4-sulphonate (NQS) in alkaline media. The assay was conducted in 96-microwell assay plates, and the absorbances of the reaction products were measured by a microplate reader at their maximum absorbance wavelengths. The optimum conditions of the reaction were refined and established. Under these conditions, calibration curves were generated, and linear regression equations were computed. The linear relations between the absorbances and drug concentrations were linear with good correlation coefficients (0.9992-0.9997) in the range of 2-80 µg/mL. The assay limits of detection were in the range of 1.5-4.2 µg/mL. The precision was satisfactory as the values of relative standard deviation did not exceed 1.70%. The accuracy of the assay was ≥98.2%. The proposed MW-SPA was successfully applied to the analysis of the SSRIs in their pharmaceutical dosage forms with acceptable accuracy and precision; the label claims were 99.2-100.5% (±0.96-1.35%). The results of the proposed MW-SPA were compared with those of the official/pre-validated assays by statistical analysis with respect to the accuracy (by
-test) and precision (by F-test). No significant differences were found between the calculated and theoretical values of the t- and F-tests at the 95% confidence level, proving similar accuracy and precision in the determination of SSRIs by both assays. The greenness of the proposed assay was confirmed by two metric tools. In addition, the assay is characterized with a high-throughput property which enables the simultaneous analysis of many samples in a short time. Therefore, the assay is a valuable tool for rapid routine application in pharmaceutical quality control units for the determination of the investigated SSRIs.
Abstract
Environmental contaminant is one of several problems harming people and wildlife. An example of current emerging contaminants are antibiotics residues that can present in water and food. ...Although antibiotics are intended to treat or prevent human and animal infections, antibiotics have also been used as animal food supplements for their ability to promote growth and feed efficiency. This overuse of antibacterial has resulted in the accumulation of antibiotics residues in food products which are eventually consumed by human. The continuous unnecessary exposure of human to antibiotics through the direct animals meet or milk, or indirectly through plants or soil can increase the chance of the emergence of multi drug resistance bacteria and consequently adversely affecting human health. New regulations have been imposed regarding antibiotics utilization. Due to the scarce of data regarding antibiotics residue conditions in different types of food intended for human consumption in Saudi Arabia, this study proposed an optimized chromatographic method (HPLC-DAD) followed by an immunoassay approach for specifically detecting tetracyclines antibiotics in animal milk samples. The method was carried out using an RP-C18 column with a mobile phase consisting of 0.01 M KH
2
PO
4
: acetonitrile:methanol (70:20:10, v/v/v) adjusted to pH 4. Improvements were observed in the method in terms of resolution and sensitivity. The protein precipitation method used for extraction demonstrated high percent recoveries of 85–101%. The method was validated according to the guidelines of the International Conference for Harmonization (ICH). It is evidently clear from these findings that the presence of tetracycline and oxytetracycline antibiotics residues in milk products from the Saudi market are below the maximum residual limits (MRLs).
Ceritinib (CER) is a potent drug that has been recently approved by the Food and Drug Administration for the treatment of patients with non-small cell lung cancer harboring the anaplastic lymphoma ...kinase mutation gene. The existing methods for the quality control of CER are very limited and suffer from limited analytical throughput and do not meet the requirements of the green analytical principles. This study presented the first-ever development and validation of three innovative green and high-throughput microwell spectrophotometric methods (MW-SPMs) for the quality control of CER in its dosage form (Zykadia® capsules). These MW-SPMs were based on the formation of colored N-vinylamino-substituted haloquinone derivatives of CER upon its reactions with each of chloranil, bromanil, and 2,3-dichloro-1,4-naphthoquinone in the presence of acetaldehyde. The optimized procedures of the MW-SPMs were established, and their analytical performances were validated according to the ICH. The linear range of the MW-SPMs was 5–150 µg/mL, with limits of quantitation of 5.3–7.6 µg/mL. The accuracy and precision of the MW-SPMs were proved, as the average recovery values were 99.9–101.0%, and the relative standard deviations did not exceed 1.8%. The three methods were applied to the determination of CER content in Zykadia® capsules and drug content uniformity testing. The greenness of the MW-SPMs was proved using three different metric tools. In addition, these methods encompassed the advantage of high-throughput analysis. In conclusion, the three methods are valuable tools for convenient and reliable application in the pharmaceutical quality control units for CER-containing capsules.
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•The fluorescence of loratadine was enhanced by dual strategy.•The fluorescence was enhanced by PET blocking and micellization.•The enhanced native fluorescence of loratidine was used ...in developing a microwell assay.•The assay was applied to the quantitation of loratidine in dosage forms and urine.•The assay meets the requirements of green analytical approaches.
This study describes the enhancement of the weak native fluorescence of loratadine (LOR) by dual strategy via photoinduced electron transfer (PET) blocking followed by micellization into sodium dodecyl sulfate micelles. The enhanced fluorescence was employed as a basis for the development of a novel microwell spectrofluorimetric assay (MW-SFA) for the determination of LOR in its pharmaceutical dosage forms and urine samples. The assay was conducted in 96-microwell assay plates, and the enhanced fluorescence signals were measured by a microplate reader at 290 and 435 nm for excitation and emission, respectively. The optimum conditions of the assay were established, calibration curve was generated, and the linear regression equation was computed. The relation between the fluorescence signals and LOR concentrations was linear with good determination coefficients (0.9992) in the range of 10 – 2000 ng mL−1. The assay limits of detection and quantitation were 4.1 and 12.5 ng mL−1, respectively. The precision was satisfactory, with values of relative standard deviation not exceeding 1.68%, and the assay’s accuracy was ≥ 99.1%. The proposed was successfully applied to the analysis of LOR in its pharmaceutical dosage forms with acceptable accuracy and precision. The label claims were 99.3 – 100.5% (±0.95 – 1.59%). Statistical analysis comparing the results of the proposed assay with those obtained by a reported pre-validated assay revealed no significant difference between both methods in terms of the accuracy and precision at the 95% confidence level. The assay was also applied to the analysis of urine samples containing LOR with accuracy ≥ 98.24%. The greenness of the proposed assay was confirmed by three efficient metric tools. In overall conclusion, the proposed assay is characterized by high sensitivity, procedure simplicity, and high throughput, enabling the simultaneous analysis of many samples in a short time. Therefore, it is a valuable tool for rapid routine application in pharmaceutical quality control units and clinical laboratories for the determination of LOR.
Abstract Background Galidesivir (GDV) is a promising new antiviral drug for the potent and safe treatment of a broad spectrum of viral diseases, including COVID-19. In the literature, no analytical ...method exists for the determination of GDV in bulk and dosage form. Objective The aim of this study was the development of versatile green and simple microwell spectrophotometric methods (MW-SPMs) for the determination of GDV in its bulk form and capsules. Methods Three MW-SPMs were developed involving the oxidation of GDV by ammonium metavanadate (AMV), chromium trioxide (CTO), and potassium iodate (PIO) in an acid medium. The reactions were carried out in 96-well plates at room temperature and the absorbances of chromogenic reaction products were measured by an absorbance microplate reader at 780, 595, and 475 nm for AMV, CTO, and PIO, respectively. Variables influencing the reactions were carefully investigated and optimized. Results Linear relations with excellent correlation coefficients (0.9991–0.9997) were found between the absorbances and GDV concentrations in the range of 25–500 µg/mL. The LODs were ≥8.3 µg/mL. The accuracy and precision of the three MW-SPMs were confirmed by recovery and replicate analysis, respectively. The recovery values were 98.6–101.2% and the RSDs were ≤1.02%. The proposed MW-SPMs were successfully applied to the analysis of GDV in bulk drug and capsules with high accuracy and precision. The greenness of the MW-SPMs was confirmed by three comprehensive metric tools. Conclusion The proposed MW-SPMs combined the inherent advantages of microwell-based analysis and the use of common laboratory reagents for the reactions involved. These advantages include high-throughput, ready automation, reduced sample/reagent volume, precise measurements, and versatility. The advantages of the use of common laboratory reagents include availability, consistency, compatibility, safety, and cost-effectiveness. Highlights Overall, the proposed MW-SPMs are versatile valuable tools for the quantitation of GDV during its pharmaceutical manufacturing.
This study describes the development of a one-step microwell spectrofluorimetric assay (MW-SFA) with high sensitivity and throughput for the determination of four statins in their pharmaceutical and ...formulations (tablets). These statins were pitavastatin (PIT), fluvastatin (FLU), rosuvastatin (ROS) and atorvastatin (ATO). The MW-SFA involves the measurement of the native fluorescence of the statin aqueous solutions. The assay was conducted in white opaque 96-microwell plates, and the fluorescence intensities of the solutions were measured by using a fluorescence microplate reader. The optimum conditions of the assay were established; under which, linear relationships with good correlation coefficients (0.9991-0.9996) were found between the fluorescence intensity and the concentration of the statin drug in a range of 0.2-200 µg mL
with limits of detection in a range of 0.1-4.1 µg mL
. The proposed MW-SFA showed high precision, as the values of the relative standard deviations did not exceed 2.5%. The accuracy of the assay was proven by recovery studies, as the recovery values were 99.5-101.4% (±1.4-2.1%). The assay was applied to the determination of the investigated statins in their tablets. The results were statistically compared with those obtained by a reference method and the results proved to have comparable accuracy and precision of both methods, as evidenced by the t- and F-tests, respectively. The green and eco-friendly feature of the proposed assay was assessed by four different metric tools, and all the results proved that the assay meets the requirements of green and eco-friendly analytical approaches. In addition, ever-increasing miniaturization as handling of large numbers of micro-volume samples simultaneously in the proposed assay gave it a high-throughput feature. Therefore, the assay is a valuable tool for the rapid routine application in the pharmaceutical quality control units for the determination of statins.
Residues of oxytetracycline, tetracycline and chlortetracycline in seafood products of Saudi Arabia were detected by using a simple, sensitive and rapid method via HPLC-PDA.
The protein precipitation ...method that was used for sample extraction demonstrated high recoveries of OTC, TC and CTC. The limits of detection were 0.015 µg/g and 0.025,0.062 µg/g for all TCs in fish and shellfish, respectively. The limits of quantitation were 0.125 µg/g and 0.175 µg/g for all TCs in fish and shellfish, respectively. The method was precise and accurate since the RSD was less than 2%, while the % recovery was 95–105%. This study determined the occurrence of OTC, TC and CTC in seafood products that are sold in KSA’s markets. The overall occurrence of these three medications in 249 seafood products was 24%(n = 60), while 15%(n = 37) exceeded the MRL. Thus, our recommendations are to enhance the monitoring of food production prior to marketing and to educate people regarding the proper disposal of antibiotics.
This study describes the development of two highly sensitive and selective sensor-assisted fluorescence immunoassays for the trace determination of copper ions, Cu(
ii
) residues, in food samples. ...These assays were the microwell-based fluoroimmuoassay (FIA) and the kinetic exclusion assay (KinExA). FIA and KinExA were assisted by a microplate reader and a KinExA™ 3200 immunosensor, respectively. Both FIA and KinExA were developed utilizing the same antibody, capturing reagent, and fluorescence signal-generating reagent. The antibody was a mouse monoclonal antibody, designated as 8D66, that specifically recognized the Cu(
ii
)-ethylenediaminetetraacetic acid complex (Cu(
ii
)-EDTA) but did not recognize Cu(
ii
)-free EDTA. The capturing reagent was Cu(
ii
)-EDTA covalently linked to bovine serum albumin protein (Cu(
ii
)-EDTA-BSA). The fluorescence-generating reagent was an anti-mouse IgG conjugated with fluorescein isothiocyanate (IgG-FITC). Both FIA and KinExA involved competitive binding reactions between Cu(
ii
)-EDTA complexes, formed in the sample solution, and Cu(
ii
)-EDTA-BSA conjugate which has been immobilized onto microwell fluorescence assay plates (in FIA) or polymethylmethacrylate beads (in KinExA) for a limited quantity of binding sites of 8D66 antibody. The conditions of both FIA and KinExA were investigated, and the optimum procedures were established. Both FIA and KinExA were validated, and all validation parameters were acceptable. Many different metal ions that are commonly encountered in food samples did not interfere with Cu(
ii
) analysis by both FIA and KinExA. Both assays were applied to the determination of Cu(
ii
) in food samples with satisfactory accuracy and precision. Both assays were compared favorably with inductively coupled plasma atomic emission spectroscopy. Comparative evaluation of FIA and KinExA revealed that KinExA had higher sensitivity and better precision than FIA, whereas, both assays had comparable accuracy. Both FIA and KinExA were superior to the existing atomic spectrometric methods for Cu(
ii
). The proposed FIA and KinExA are anticipated to effectively contribute to assessing Cu(
ii
) concentrations and controlling the exposure of humans to its potential toxicities.
This study describes the development of two highly sensitive and selective sensor-assisted fluorescence immunoassays for the trace determination of copper ions, Cu(
ii
) residues, in food samples.