Chromatographic Method For Analysis Research Articles

Validation of Reverse Phase High-performance Liquid Chromatography Analytical Method for Osimertinib Mesylate and its Degradation Products for Osimertinib Mesylate Tablets

Validation of Reverse Phase High-performance Liquid Chromatography Analytical Method for Osimertinib Mesylate and its Degradation Products for Osimertinib Mesylate Tablets

Recent trends in analysis of mycotoxins in food using carbon-based nanomaterials

Mycotoxins (MYTs), a class of low molecular weight secondary metabolites produced by filamentous fungi in food and feed, pose serious global threat to both human health and world economy. Due to their mutagenic, teratogenic, carcinogenic and immunosuppressive effects, the International Agency for Research on Cancer has classified various MYTs under Group 1 to 3 category with aflatoxins being designated under Group 1 category (carcinogenic to humans). Also, the presence of MYTs in trace amounts in diverse food matrices necessitates exploration of highly sensitive methods for onsite analysis. Although conventional chromatographic methods are highly sensitive, they are expensive, tedious and cannot be applied for rapid onsite analysis. In recent years the application of nanomaterials especially carbon-based nanomaterials (CNMs) in the fabrication of low-cost and miniaturized electrochemical and optical sensors has enabled rapid onsite analysis of MYTs with high sensitivity and specificity. Moreover, the CNMs are employed as effective solid phase extraction (SPE) adsorbents possessing high specific surface area for effective enrichment of MYTs to improve the sensitivity of chromatographic methods for MYT analysis in food. This article aims to overview the recent trends in the application of CNMs as SPE adsorbents for sample pretreatment in chromatographic methods as well as in the fabrication of highly sensitive electrochemical and optical sensors for rapid analysis of MYTs in food. Initially, the efficiency of various functionalized CNMs developed recently as adsorbent in packed SPE cartridges and dispersive SPE adsorbent/purification powder is discussed. Then, their application in the development of various electrochemical immunosensors involving functionalized carbon nanotubes/nanofibers, graphene oxide, reduced graphene oxide and graphene quantum dots is summarized. In addition, the recent trends in the use of CNMs for fabrication of electrochemical and fluorescence aptasensors as well as some other colorimetry, fluorometry, surface-enhanced Raman spectroscopy and electrochemical based sensors are compared and tabulated. Collectively, this review article can provide a research update on analysis of MYTs by carbon-based nanomaterials paving a way for identifying future perspectives.

Relationship between the UPLC Fingerprints of Citrus reticulata "Chachi" Leaves and Their Antioxidant Activities.

Citrus reticulata "Chachi" (CRC) leaves contain abundant flavonoids, indicating that they possess good nutritional/pharmacological research and development potential. This study aims to explore chemical antioxidant quality markers based on the spectrum-effect relationship and quality control strategy of CRC leaves. The ultrahigh performance liquid chromatography (UPLC) system was used to establish chromatographic fingerprints of Citrus reticulata "Chachi" leaves. Simultaneously, they were evaluated by using similarity analysis (SA), hierarchical cluster analysis (HCA), and principal component analysis (PCA). Afterwards, the DPPH assay was adopted to study the antioxidant effects. The spectrum-effect relationship between UPLC fingerprints and DPPH radical-scavenging activities was studied with grey relational analysis (GRA). Analysis results indicated that there were twenty-one common peaks of fourteen batches of CRC leaves which were from different regions of Guangdong province, and their similarities ranged from 0.648 to 0.997. HCA results showed that fourteen batches of samples of CRC leaves could be divided into six classes at Euclidean distance of 5. The results from GRA showed that tangeretin and hesperidin were the main flavonoids responsible for the antioxidant activity in CRC leaves. In conclusion, this research established a chromatographic analysis method suitable for CRC leaves and demonstrated that chromatographic fingerprints analysis combined with the antioxidant activity could be used to evaluate the material basis of CRC leaves and may provide a reference to establish a quality standard.

The Ethnopharmacological Uses, Metabolite Diversity, and Bioactivity of Rhaponticum uniflorum (Leuzea uniflora): A Comprehensive Review.

Rhaponticum uniflorum (L.) DC. (syn. Leuzea uniflora (L.) Holub) is a plant species of the Compositae (Asteraceae) family that is widely used in Asian traditional medicines in China, Siberia, and Mongolia as an anti-inflammatory and stimulant remedy. Currently, R. uniflorum is of scientific interest to chemists, biologists, and pharmacologists, and this review includes information from the scientific literature from 1991 to 2022. The study of the chemodiversity of R. uniflorum revealed the presence of 225 compounds, including sesquiterpenes, ecdysteroids, triterpenes, sterols, thiophenes, hydroxycinnamates, flavonoids, lignans, nucleosides and vitamins, alkanes, fatty acids, and carbohydrates. The most studied groups of substances are phenolics (76 compounds) and triterpenoids (69 compounds). Information on the methods of chromatographic analysis of selected compounds, as well as on the quantitative content of some components in various organs of R. uniflorum, is summarized in this work. It has been shown that the extracts and some compounds of R. uniflorum have a wide range of biological activities, including anti-inflammatory, antitumor, immunostimulatory, anxiolytic, stress-protective, actoprotective, antihypoxic, anabolic, hepatoprotective, inhibition of PPARγ receptors, anti-atherosclerotic, and hypolipidemic. Published research on the metabolites and bioactivity of R. uniflorum does not include clinical studies of extracts and pure compounds; therefore, an accurate study of this traditional medicinal plant is needed.

Stability indicating validated high‐performance liquid chromatography method for simultaneous estimation of chlorin e6 and curcumin in bulk and drug‐loaded lipidic nanoformulation

AbstractChlorin e6 and curcumin are often used in the treatment of various cancers, including breast, skin, colon, and lung cancer. The present objective of this study is to develop a simple, sensitive, precise, and selective reversed‐phase high‐performance liquid chromatography analytical method for the estimation of curcumin and chlorin e6. Curcumin and chlorin e6 are both estimated simultaneously in bulk and on nanocarriers based formulations. The efficient chromatographic separation of chlorine e6 and curcumin was obtained using SUPELCO C‐18 column with a mobile phase containing ACN and water (pH 2.6 adjusted with trifluoracetic acid 0.1%) in an isocratic mode at a detection wavelength of 412 nm. The flow rate and column oven temperature were kept at 1 ml/min and 30°C, respectively. The retention times for curcumin and chlorin e6 were found to be 11.7 and 13.5 min, respectively. The simultaneous technique was used to estimate the %drug entrapment efficiency, drug loading, as well as the cumulative percentage of drug release of chlorin e6 and curcumin in‐house designed nanoformulations. %Drug loading and entrapment efficiency were found to be 2.22% and 79.92% for curcumin, while 2.43% and 89.47% for chlorin e6, respectively.

Phosphorothioate oligonucleotides separation in ion-pairing reversed-phase liquid chromatography: effect of temperature

Analysis of diastereomers of phosphorothioate oligonucleotides in ion-pairing reversed-phase liquid chromatography is affected not only by the character and concentration of ion-pairing system, but also by the separation temperature. In this work, eight ion-pairing systems at two concentrations buffered with acetic acid were used with octadecyl column to investigate the effects of temperature (in the range from 20 °C to 90 °C) on retention, diastereomeric separation, resolution of mers of different length and resolution of oligonucleotides with different number of phosphorothioate linkages. It was observed that elevated temperature suppresses the diastereomeric separation and oligonucleotide peaks become narrower. This improves the resolution of n and n-1 mers at elevated temperature. Plots of ln k (k = retention factor) versus reciprocal absolute temperature show that for 100 mM ion-pairing systems the increase in temperature does not lead to simple decrease in oligonucleotides retention as generally observed in reversed-phase liquid chromatography. The aim of this work is to improve chromatographic method for analysis of phosphorothioate oligonucleotides.

Treatment of Wastewaters Containing Sulfonylurea Herbicides by Electroflotation: Chemical and Ecotoxicological Efficacy

Electroflotation (EF) is an electroanalytical technique applied for separating substances suspended in phase and is reported to be efficient for effluents and wastewater treatments. To date, there are no studies employing EF for the treatment of wastewater containing toxic organic compounds. In the present study, EF was tested as an electro-oxidative process to decontaminate wastewater containing the sulfonylurea herbicide chlorimuron-ethyl. To evaluate the value of EF, both the chemical and ecotoxicological efficacies were assessed in non-treated and treated wastewaters; the former monitored the herbicide and degradation products in the reaction medium through analytical chromatographic methods, whereas the latter comprised testing the 72 h growth of the green microalga Raphidocelis subcapitata and the 48 h lethality of the cladoceran Daphnia magna. Analysis by HPLC-UV allowed the monitoring of the electrochemical reaction, and a degradation mechanism based on gas chromatography–mass spectrometry was proposed. Despite the compromised herbicide structure, non-treated and treated wastewaters were similarly toxic for the microalgae Raphidocelis subcapitata and the invertebrate Daphnia magna. Even though EF did not remove the wastewater toxicity, the results indicate that toxic organic compounds are potentially oxidized by EF while signaling the need to combine chemical and ecotoxicological approaches to gauge the environmental sustainability of EF.

A Validated Reversed-phase High-performance Liquid Chromatography Analytical Method for the Analysis of Methylcobalamin in Bulk Drugs and T-Dosage Formulation

>The aim of the existing work is to develop and validate a reversed-phase high-performance liquid chromatography (RP-HPLC) technique for the quantitative determination of methylcobalamin in bulk and t formulations that is easy, rapid, precise, accurate, affordable, and sensitive. Proceeding a princeton (C18) column with dimensions (250 x 4.6 mm, 5 μ), the isocratic elution technique was used. The mobile buffer phase comprised water (pH 6.5, adjusted with sodium chloride) and methanol in the relation (55:45) v/v. It was shown that the methylcobalamin retention time was 2.022 minutes under ideal circumstances. The correlation coefficient (r2) for the 900–2400 mcg/mL methylcobalamin concentration range, which was used to verify the method’s linearity, was 0.9994. Methylcobalamin had a recovery rate of 99.98–100.01% and RSD of 2%. The marketed t formulation test was successfully completed with 99.86%. The proposed and validated approach underwent the accuracy, precision, specificity, linearity, and system suitability testing advised by the ICH. RP-HPLC was used in the market formulation.

Quality Evaluation of Crude and Salt-Processed Cuscutae Semen through Qualitative and Quantitative Analysis of Multiple Components Using HPLC Combined with Chemometrics

Cuscutae Semen (CS; Chinese common name “Tusizi”) is one of the most common traditional herbal medicines used to treat liver and kidney diseases in China. Although it is most commonly used as a processed product, little progress has been made on the quality control of CS and salt-processed Cuscutae Semen (PCS). The purpose of this study was to establish a comprehensive strategy integrating chromatographic analysis and chemometric methods for quality evaluation and discrimination of CS and PCS. An accurate and reliable HPLC method was established for the simultaneous quantification of 12 analyte compounds in CS and PCS. The results showed that this method exhibited desirable sensitivity, precision, stability, and repeatability. Multivariate chemometric methods were applied to analyze the obtained HPLC fingerprints, including hierarchical cluster analysis (HCA), principle component analysis (PCA), and partial least squares discriminant analysis (PLS-DA). The results indicated that CS and PCS samples showed a clear classification of the two groups, and three chemical markers with great contributions to the differentiation were screened out. Collectively, the chemometrics combined with the quantitative analysis based on HPLC fingerprint results indicated that salt processing may change the contents and types of components in Cuscutae Semen.

Polychlorinated Biphenyls in the Snow Cover of South-Eastern Kazakhstan

The presence of large sources of environmental pollution due to persistent organic pollutants (POPs), in particular, polychlorinated biphenyls (PCBs), in Kazakhstan necessitates the assessment of pollution as a result of these toxicants. For this purpose, we chose snow cover as an indicator for assessing pollution status in the study area. An assessment of the PCB accumulation level included in the list of POPs was carried out for a snow cover (SC) study in south-east Kazakhstan. The content of PCBs with a wide congener composition was determined using the chromatographic analysis method. During the winter periods of 2014, 2015, 2018–2020 and 2021, the SC pollution of the study area from up to 25 individual PCB congeners was identified. These congeners included highly toxic dioxin-like congener PCBs 105; 108; 114; 118 and “marker” PCBs 52; 101; 138; 153. These congeners were mainly found in snow samples with a wide range of PCB congener compositions. The main PCB pollution sources were indicated. The analysis of the obtained results and structure of the congener composition of PCBs show that the SC contamination in this territory occurs under the influence of local and regional sources.

Development and validation of stability indicating RP-HPLC method for the estimation of azilsartan in Bulk

Angiotensin-receptor blockers are often considered adequately efficacious in reducing blood pressure. By accepting the current regulatory requirements for an analytical method development, a reversed phase high performance liquid chromatographic method for routine analysis of Azilsartan has been developed. The optimized method was achieved using unisol C-18 (3μm, 4.6×150mm) coloumn with mobile phase consisting of mixture acetonitrile and methonal (90:10v/v) with a flow rate of 0.8ml/min at 249nm. The Linear concentration range is 2-10μg/ml. The detection and quantitation limit was found to be 0.45μg/ml. And the detection limit was found to be 0.149μg/ml. There are no interfering peaks under performed degradation conditions. The optimized method was validated according to ICH guidelines.

Review of new technologies used for meat identification

The present article represents an analysis of trends in development of test-systems for identification of meat. These test systems are commonly used in food production and research laboratories. The relevance of development of methods for identification of meat kinds is related not only to the food restrictions, which are practiced in some religions and related to consumption of certain types of meat, but also with the hygienic aspects of food production. Also, this research is inspired also by the acute issue of food products adulteration and the replacement of one type of meat with another one. The article considers the trends in the develop ment of microanalysis method that use immunochromatic research, i. e. methods based on molecular biology. Also this article considers the devices that do not use chromatographic methods of analysis. Examples of the development of test systems based on various methods of analysis for the identification of meat are given below. Attention is focused on the prospects of combining these methods, including colorimetric methods for identification of meat. It is also specified that the emergence of new dyes and new enzyme systems, suitable for use in enzyme-immunoassay, can enhance the sensitivity of these test systems. It is also noted that the development of technologies associated with sorbents can contribute to a better separation of the test substrates and this way to increase the sensitivity of the test in case of small amounts of test substrate. It is also noted that the use of various types of iso thermal amplification can reduce the analysis time necessary for meat identification. Various schemes of devices for microanalysis are given; their advantages and disadvantages are listed. An example of proteomes application for meat identification is given. It is shown that this method can also be applied in the heat treatment of meat. The prospects for the development of such devices are analyzed. It is concluded that the development of systems for microanalysis in the form of quick tests is quite relevant and promis ing. It is indicated that theoretically in the future such analytical systems, due to the use of microfluidic technologies, will be able to combine several methods. The authors proposed to use machine-aided cognition methods to analyze data obtained from similar test systems in order to increase their sensitivity.

Design of experiment techniques for the optimization of chromatographic analysis conditions: A review.

Design of experiment (DoE) techniques have been widely used in the field of chromatographic parameters optimization as a valuable tool. A systematic literature review of the available DoE techniques applied to the development of a chromatographic analysis method is presented in this paper. First, the most common available designs and the implementation steps of DoE are comprehensively introduced. Then the studies in recent 10 years for the application of DoE techniques in various chromatographic techniques are discussed, such as capillary electrophoresis, liquid chromatography, gas chromatography, thin-layer chromatography, and high-speed countercurrent chromatography. Current problems and future outlooks are finally given to provide a certain inspiration of research in the application of DoE techniques to the different chromatographic techniques field. This review contributes to a better understanding of the DoE techniques for the efficient optimization of chromatographic analysis conditions, especially for the analysis of complex systems, such as multicomponent drugs and natural products.

Chemometric-Based AQbD and Green Chemistry Approaches to Chromatographic Analysis of Remogliflozin Etabonate and Vildagliptin.

According to the green chemistry approach, during method development, the usage of toxic and carcinogenic organic solvents should be avoided or minimized for the safety of the environment and analysts. The chromatographic methods such as reverse-phase high-pressure liquid chromatography (RP-HPLC) and high-performance thin-layer chromatography (HPTLC) include the usage of class 2 organic solvents as per the International Council for Harmonization (ICH) Q3C (R6) guideline. The chromatographic analysis by HPTLC requires less organic solvent compared to the RP-HPLC method. Hence, HPTLC-based chromatographic analysis of vildagliptin (VIL) and remogliflozin etabonate (RMG) has been carried out using green chemistry and analytical quality by design (AQbD) approaches. The principal component analysis (PCA)-based chemometric analysis was applied for the identification of critical method variables (CMV) for the development of the method. The design of experiments (DoE)-based Box-Behnken design (BBD) was applied for response surface modeling (RSM) and optimization of CMV. The analytical design space (ADS) and analytical control point were navigated for the development of the HPTLC method as per the quality target analytical profile. The chromatographic analysis of VIL and RMG was carried out using silica gel G60 F254 as the stationary phase and acetone-ethyl acetate-water-triethylamine (7.0 + 2.5 + 0.3 + 0.2, v/v) as the mobile phase. The HPTLC method was validated as per the ICH Q2 (R1) guideline. The HPTLC method was applied for the assay of fixed-dose combinations (FDCs) of VIL and RMG, and the results were found to comply with their labeled claim. The developed method included the usage of organic solvents that belong to the class 3 category as per the ICH Q3C (R6) guideline. Hence, the developed method can be used as an eco-friendly alternative to published chromatographic methods for quality control and routine analysis of FDCs of VIL and RMG in the pharmaceutical industry. Chromatographic analysis of VIL and RMG using green chemistry and AQbD approaches. Application of the method for assay of drugs in their combined pharmaceutical dosage forms.

Principal Component Analysis and DoE-Based AQbD Approach to Multipurpose HPTLC Method for Synchronous Estimation of Multiple FDCs of Metformin HCl, Repaglinide, Glibenclamide and Pioglitazone HCl.

Metformin HCl (MET) is a mostly used antidiabetic drug and several fixed-dose combinations are available in the market with other antidiabetic drugs for the treatment of insulin-independent diabetes. Numerous chromatographic methods have been reported for the estimation of multiple fixed dose combination (FDC) of MET but each FDC needs a separate and dedicated chromatographic method for analysis. No chromatographic method has been reported yet in the literature which promotes synchronous estimation of multiple FDCs of MET to minimize organic solvent consumption, time and cost of analysis. Hence, the multipurpose-high performance thin layer chromatography (HPTLC) method was developed for the synchronous estimation of FDCs of MET by the implementation of an enhanced analytical quality by design approach. Principal component analysis was applied for the identification of critical method risk parameters. Box-Behnken design was applied for response surface analysis. The method operable design ranges was navigated and the control strategy was framed. The developed method was applied for the analysis of multiple FDCs of metformin. The developed method can synchronously analyze multiple FDCs of MET which required several chromatographic methods. Hence, the present method is multipurpose-HPTLC for the synchronous estimation of multiple FDCs of MET which required minimum organic solvent, time and cost of analysis.

Development of a method for determining the morpholinium thiazotate using more economic and green GC/MS assay with an fid detector

One of the main steps in the pharmaceutical development of drugs is the choice of quality control methods. The correctness of the method must be confirmed by validation. In addition, manufacturers take into account various economic and environmental factors. It is especially important to determine the above aspects for domestic and promising drugs, such as the morpholinium thiazotate. The aim. During the development of methods for the routine analysis of medicinal products, attention should be paid to efficiency of analysis, budget, as well as their impact to the environment. Because of this reason, not only new methods for routine analysis should be developed. It is important this methods must be environmentally-friendly and cost-efficient. Materials and methods. The determination of the morpholinium thiazotate was carried out by HPLC using the SunFire C18 (150 mm × 4,6 mm, 5,0 μm) and gas chromatography with a flame ionization detector using the Rxi-5 ms (30 m long, 0.25 mm outer diameter and 0.25 μm liquid stationary phase thickness). Results. Various chromatographic methods for the routine quantitative analysis of morpholinium thiazotate were developed. The most suitable conditions for sample preparation were established. Proposed methods were compared to find the most ecological and economic. Conclusions. Proposed methods were accurate and reliable. However, an environmental impact assessment showed that GC-FID is a more environmentally friendly and economical method of analysis. Using 12 Principles of green analytical chemistry, the overall “analytical GREEnness (AGREE)” scale for the proposed analytical approach was computed 0.72, showing the good greener nature of the proposed analytical approach

Green Assessment of Chromatographic Methods Used for the Analysis of Four Methamphetamine Combinations with Commonly Abused Drugs

Numerous agents with anxiolytic or stimulant effects have the potential to be overused, and their misuse is associated with serious side effects. In Saudi Arabia, the estimated percentage of Saudis who abuse drugs is around 7–8% and the age range is 12–22 years. Methamphetamine, captagon, tramadol, heroin, and cannabis/cannabinoids have been proven to be the most commonly abused drugs in Saudi Arabia, with methamphetamine being at the top of the list. The present study focuses on the chromatographic analytical methods used for the analysis of methamphetamine in combination with commonly abused drugs, aiming to point out the greenest among them. These mixtures have been chosen as they are analyzed periodically and frequently in criminal evidence and forensic medicine. Therefore, the chances of hazards for analysts and the environment are high if the mixtures are not handled appropriately. This study aims to compare 23 chromatographic methods used for the analysis of methamphetamine mixtures in four major combinations, and to assess their greenness by using three greenness assessment tools, namely, NEMI, ESA and AGREE, to recommend the greenest analytical method. The NEMI results were proven to have low discriminating abilities and, accordingly, the comparisons are based on ESA and AGREE scores. The analysis results show that the safest methods with the most eco-friendly results (based on ESA and AGREE) are the GC-MS method proposed by Mohammed et al. to analyze methamphetamine and captagon mixtures (ESA = 79 and AGREE = 0.57), the UHPLC–MS-MS method proposed by Busardò et al. to analyze methamphetamine and cannabis/cannabinoid mixtures (ESA = 78 and AGREE = 0.57), the LC-MS method proposed by Herrin et al. to analyze methamphetamine and tramadol mixtures (ESA = 81 and AGREE = 0.56), and the LC-MS method proposed by Postigo-et al to analyze methamphetamine and heroin mixtures (ESA = 76 and AGREE = 0.58).

Evaluation of Greenness of LC-MS Chromatographic Methods for Simultaneous Analysis of Mixtures of Serotonin, Dopamine, Acetylcholine, GABA and Glutamate: AGREE Tool Application

The analytical GREEnness metric (AGREE) tool is widely used as a reliable greenness assessment method for chromatographic analyses. The AGREE tool has the ability to determine the greenness of analytical methods in terms of both quality and quantity, whereas other commonly used methods assess the greenness either quantitatively or qualitatively. Greenness profiles of six chromatographic methods for simultaneous estimations of serotonin, dopamine, acetylcholine, GABA and glutamate were assessed using AGREE and NEMI tools as a case study. The AGREE assessment tool proved to be user-friendly, and provides a full profile of assessment, hence it can be described as the tool of choice for the assessment of LC-MS chromatographic methods. For optimum application, the weights of 4 of the 12 assessment criteria were set high (weight of four) due to their importance, namely criteria number 7 (waste), number 8 (analysis thruput/number of analytes per run), number 11 (toxicity) and number 12 (operator’s safety). Setting proper weights of the assessment criteria contributed significantly to the discrimination of greenness of the compared methods. The selected greenest method for the analysis of the proposed quinary mixture showed an AGREE tool pictogram with a 0.66 score. Additionally, the selected method allows simultaneous estimation of seven constituents in total. It offers high sensitivity, allowing detection of acetylcholine, serotonin and glutamate at levels as low as 2 pg, and dopamine, norepinephrine, GABA and glycine at levels as low as 10 pg, and finally offers fast analysis where all components can be analyzed within 5 min.

High performance liquid chromatography analytical method for glutaraldehyde determination in disinfectants

Glutaraldehyde (GA) is commonly used to disinfect surfaces and equipment in industries, agriculture, healthcare, laboratories, etc. To evaluate the quality of products containing GA, a HPLC method using 2,4-diphenylhydrazine derivative was developed. The analyte was separated by CN column with isocratic of acetonitrile and phosphoric acid 0,1% (ratio 50 : 50) and detected by PDA detector using a wavelength of 360 nm. The method was validated for specificity, linear curves, precision and accuracy which meet the requirements of AOAC.

Development of HPTLC method for the simultaneous estimation of quercetin, curcumin, and ascorbic acid in herbal formulations

Antioxidants can scavenge free radicals in the human body and have been associated with a protective effect on the human body against a variety of diseases. As a part of normal cellular function, free radical production occurs continuously within the body at a cellular level. Excess-free radical production from endogenous or exogenous sources may result in a variety of diseases. Vitamins (A, C, and E), carotenoids, polyphenols, and flavonoids are some examples of common natural antioxidants. Among the different antioxidants quercetin, curcumin, and ascorbic acids have been extensively studied, and numerous reports on their antioxidant activity are available in the literature. Antioxidants are regarded as a key player in the current pandemic situation, and they play a key role as a preventive and controlling tool. There are a variety of herbal formulations and immune booster therapies available, many of which claim to contain various antioxidants and are effective against covid-19. Numerous herbal formulations and immune booster therapies are available and claimed to have different antioxidants and other constituents. In the present study simple, rapid, accurate, precise, and sensitive high-performance thin-layer chromatographic analytical method was developed for simultaneous estimation of quercetin, curcumin, and ascorbic acid. The developed method can effectively employ for different polyherbal formulations. The developed method was validated according to ICH guidelines. In the present study, an aluminum plate precoated with 60F 254 silica gel was used as a stationary phase whereas chloroform: ethyl acetate: formic acid (6:6:2.5 v/v/v) was used as a mobile phase. Chromatographic detection was performed with a Camag TLC scanner at 265 nm. The linearity range for the proposed method was found to be 500–1000 ng/band for all three analytes. The antioxidant potentiality of selected antioxidants was evaluated by an optimized UV visible spectroscopic-reducing power method. The reducing power method was optimized for the concentration and volume of reagent, incubation temperature, and time. The results of the study revealed that quercetin is having the best antioxidant potential compared to curcumin and ascorbic acid.