HPTLC Method For Simultaneous Estimation Research Articles

DoE based development and validation of HPTLC method for simultaneous estimation of Curcumin and Naringin in topical gel formulation

DoE based development and validation of HPTLC method for simultaneous estimation of Curcumin and Naringin in topical gel formulation

Analytical Quality Risk Assessment and Design of Experiments to Green HPTLC Method for Simultaneous Estimation of Sildenafil Citrate and Dapoxetine Hydrochloride.

A green and robust high-performance thin-layer chromatographic method has been developed for the simultaneous estimation of sildenafil citrate and dapoxetine hydrochloride. A fractional factorial design was applied for analytical quality risk assessment of potential analytical risk factors. The identified critical analytical risk factors were optimized using the design of experiment-based response surface analysis by full factorial design. The analytical design space was navigated for the optimization of the method and the control strategy was framed for low-risk life-cycle management of the chromatographic method. The chromatographic analysis of sildenafil and dapoxetine was carried out on a TLC plate coated with silica gel G60 F254 using n-butanol:ethyl acetate:ethanol (8.0 + 2.0 + 0.5, v/v) as mobile phase. The chromatographic peaks of sildenafil and dapoxetine were found to be at Rf 0.29 and 0.69, respectively. The method was found to be accurate, precise, robust, specific and sensitive. The fixed-dose combinations of sildenafil and dapoxetine were assayed and results were found in compliance with their labeled claim. The present method was developed using safe and eco-friendly organic solvents for the safety of analysts and the protection of the environment. The greenness profiles of developed and reported methods were evaluated using the NEMI scale and AGREE software.

Development and Validation of HPTLC Method for Simultaneous Estimation of Azelnidipine and Telmisartan

Development and Validation of HPTLC Method for Simultaneous Estimation of Azelnidipine and Telmisartan

Development and Validation of HPTLC Method for Simultaneous Estimation of Piperine and Scopoletin in <i>Ajmodadi churna</i>

In this recent study, several trials were made to develop a HPTLC method for quantification of scopoletin and piperine in the Ajmodadi formulation. HPTLC was done on pre-coated silica gel 60 F254 plates with a mobile phase of toluene:ethyl acetate:methanol:formic acid (3.9:3.9:0.3:1.7 v/v/v/v). The retardation factors for scopoletin 0.75 and for piperine was 0.86 and found a good and defined resolution peak. Densitometer analysis of scopoletin and piperine was carried out at 335 wavelength (nm). The developed method was validated as reported in ICH guidelines (Q2R1). A linearity study shows that scopoletin and piperine was linear, and the recovery studies revealed a recovery in between 98-102 % as per guideline. This method was established to be rapid, delicate, accurate and specific, thus, these methods may be used in the quantification of the drug in any polyherbal formulation.

Analytical Method Development and Validation of Hptlc Method for Simultaneous Estimation of Telmisartan and Azelnidipine in Tablet Dosage Form

Background: A specific, accurate, precise, robust and cost effective HPTLC method was developed and validated for quantitative analysis of Telmisartan and Azelnidipine in fixed dose combination. The separation was carried out on pre-coated TLC plates with Silica gel 60 F254 as stationary phase using Chloroform: Ethyl acetate: Methanol (7.5:2:0.5 v/v/v) as developing system followed by densitometry measurement of bands at 280nm. Result: The separation of Telmisartan and Azelnidipine was found to be at the following RF values 0.3 and 0.63 respectively. Calibration curves were found to be linear over the concentration ranges 400-4000ng/band, 80-800ng/band for Telmisartan and Azelnidipine with correlation coefficient 0.9957 and 0.9966 respectively. Accuracy was found between 98.03% - 102% and 98.72% - 102 % for Telmisartan and Azelnidipine respectively. LOD was found to be 1.04% and 1.6% and LOQ was found to be 3.49% and 5.57% for Telmisartan and Azelnidipine respectively Conclusion: The results showed that the proposed HPTLC method is reliable for routine analysis for simultaneous determination of Telmisartan and Azelnidipine without any interference.

Development of new Validated HPTLC Method for simultaneous estimation of Canagliflozin and Metformin in Tablet Formulation

A new HPTLC method for simultaneous determination of Canagliflozin and Metformin was developed, validated and compared with the reported one. An excellent resolution with Rf values 0.21±0.02 and 0.50±0.03 for Metformin and Canagliflozin respectively was achieved by using optimized chromatographic conditions. Toluene: Methanol: Triethyl amine: Glacial Acetic acid (7:2.6:0.2:0.2, v/v/v/v) was chosen as a mobile phase. Detection was done at 254nm. When marketed formulation was analyzed by developed method, the % drug contents were found to be 98.20±0.24 and 99.33±1.80% w/w for Metformin and Canagliflozin respectively. Developed method was validated for linearity and range, DL, QL, accuracy, precision, robustness and specificity. The method was found to be linear, accurate, precise, robust and sensitive. The method was found to be linear in a range of 75-750ng/band of Canagliflozin with R2 value 0.9981 and 250-2500ng/band of Metformin with R2 value 0.9963. Detection and quantitation limits were found to be 8.01 and 24.28ng/band for Metformin and 8.27 and 25.07ng/band for Canagliflozin respectively. The % drug recovery was calculated as a measure of accuracy. The % drug recovery was found to be near to 100 %w/w confirmed the accuracy of the method. In case of precision studies, %RSD values were found to be less than two indicated, the method is precise. The % RSD values of deliberate variations like amount of mobile phase, saturation time were found to be less than two showed method is robust. The developed method was compared with the reported method and found to be more linear, accurate, precise and sensitive.

Validated HPTLC Method for Simultaneous Estimation of Dihydroartemisinin and Piperaquine Phosphate in Pharmaceutical Dosage Form

Validated HPTLC Method for Simultaneous Estimation of Dihydroartemisinin and Piperaquine Phosphate in Pharmaceutical Dosage Form

Developed and validated method for the simultaneous estimation of ferulic acid and chlorogenic acid in four Indian ananas comosus merrill cultivars through HPTLC

Ananas comosus (L) Merrill fruit is used for the treatment of urinary tract infection, and jaundice. The fruit has anti-obesity activity and used as antihelmithic agent for the treatment of intestinal worms. The Ananas comosus fruit contains many phytoconstituents; among them are Ferulic acid and Chlorogenic acid exhibiting various pharmacological activities. To ensure the content of uniformity of biomarkers, the aim of the present study was to develop a HPTLC method for simultaneous estimation of Ferulic acid and Chlorogenic acid in Ananas comosus fruit. The standard marker compounds and four different fruit extract solutions were applied on the HPTLC plate (20 mm × 10 mm dimension) with the help of Linomat 5 applicator by using WinCATS software. The solvent system toluene, ethyl acetate, methanol, formic acid (4:4:1.2:0.8, v/v) was used as mobile phase. The λmax was set for Ferulic acid and Chlorogenic acid at 280 nm. The calibration range of Ferulic acid was found to be 300-900 ng/ml, with the linear equation Y= 338.485+6.238x with coefficient of determinants (r2) of 0.992 and the calibration range of Chlorogenic acid was found to be 200-800 ng/spot with the linear equation Y= 282.512+ 4.932x with coefficient of determinants (r2) of 0.996. The limit of detection and limit of quantification were found to be for Ferulic acid 14.35, 40.12 ng/spot, and 10.26, 29.51 ng/spot, for Chlorogenic acid respectively. The relative standard deviation of precision and recovery of Ferulic acid and Chlorogenic acid was < 2.0%. The developed method was accurate, specific, precise and reproducible. Keywords: Ananas comosus cultivars, Ferulic acid, Chlorogenic acid, Simultaneous estimation, Method Validation.

Chemometric and DoE-Based Analytical Quality Risk Management to HPTLC Method for Simultaneous Estimation of Metronidazole and Norfloxacin.

According to the upcoming ICH Q14 guideline, the development of an analytical method by the implementation of the AQbD approach based on analytical quality risk management and design of experiments will become a regulatory requirement for the registration of new drug substances and products. In literature, the HPTLC method has not been reported yet for simultaneous estimation of metronidazole and norfloxacin. Hence, the robust HPTLC method has been developed and validated for simultaneous estimation of metronidazole and norfloxacin using QRM and the DoE-based enhanced AQbD approach. The principal component analysis was applied for chemometric-based risk assessment of method risk parameters. The high-risk method parameters were optimized by a DoE-based full-factorial design. The MODR and control strategy was estimated for quality risk management throughout the lifecycle of the HPTLC method. The HPTLC method was developed using silica gel 60F254 as stationary phase and acetonitrile-methanol-formic acid-ammonia (9.5 + 0.5 + 0.5 + 0.3, v/v) as mobile phase. The developed method was validated as per ICH Q2 (R1) guideline. The developed method was applied for the assay of combined pharmaceutical dosage forms of metronidazole and norfloxacin and results were found in compliance with their respective labeled claim.

Development and Validation of HPTLC Method for Simultaneous Estimation of Berberine, Gallic Acid and Ursolic Acid in a Polyherbal Blend

A sensitive high-performance thin-layer chromatography method was developed for simultaneous estimation of berberine, gallic acid and ursolic acid in a polyherbal blend and validated as per ICH guidelines. Polyherb-al blend was prepared using widely recommended herbal plants for platelet augmentation activity viz. Carica papaya, Berberis aristata, Ocimum Sanctum, and Tinospora Cordifolia. The optimized separation was ob-tained with TLC aluminum plates pre-coated with silica gel G60 F254 as a stationary phase and a solvent sys-tem containing Toluene : Ethyl acetate : Methanol : Formic acid (3:3:0.2:0.1 v/v/v/v). Berberine and gallic ac-id were found to demonstrate linearity in the range of 1μg/band – 6μg/band and ursolic acid in the range of 20 μg/band – 100 μg/band with the regression coefficient in acceptable limits. The method was also found to be specific and precise one. The accuracy of the developed method at 80 %, 100 % and 120 % levels was found to be within limits and % RSD was found to be less than 2. LOD and LOQ of all three standards were also determined. Blend was also quantified for the amount of berberine, gallic acid and ursolic acid presence and was found to be 37.8 μg, 46.1 μg and 108 μg per mg, respectively.

Development and validation of HPTLC method for simultaneous estimation of bioactive components in combined extracts of three hepatoprotective plants

Picroside-I, andrographolide and silybin exhibit manifold biological activities, present as bioactive components in Picrorhiza kurroa (roots), Andrographis paniculata (aerial parts) and Silybum marianum (seeds), respectively. A validated HPTLC method was developed for the simultaneous determination of picroside-I, andrographolide and silybin in the combined hydroalcoholic extract of the plants as per ICH guidelines. On silica gel F254 HPTLC plates, the combined hydroalcoholic extract of plants and standard solutions were applied. The plates were run in twin chamber consisting n-butanol: glacial acetic acid: water (6:1:3, v/v/v) as a mobile phase and documented at 254 and 366 nm using Camag TLC scanner III with WinCATS software. The analysis revealed that a good linear relationship was established between response (peak area) and amount in the range of 60–600 ng/spot. The LOD (ng/spot) and LOQ (ng/spot) for estimation of picroside-I, andrographolide and silybin were found to be 15.089, 21.969, 25.702 and 45.725, 66.573, 77.885, respectively. Recovery experiments showed 97.75–103.74%, 99.75–101.12% and 98.96–101.69% for picroside-I, andrographolide and silybin, respectively. The HPTLC method showed good linearity, recovery and high precision of all the three bioactive components, which can be applied for the routine analysis of polyherbal formulations containing these plants as their ingredients.

Stability Indicating HPTLC Method for Sofosbuvir and Daclatasvir in Combination

Direct acting antiviral agents represent the major advance in treatment of hepatitis C virus (HCV) infection. Daclatasvir with sofosbuvir that are co-administrated once per day oral dose has been reported to achieve a high rate of virological response in patients with HCV genotype 1, 2 or 3. So, the basic objective of a research involved development and validation of stability indicating HPTLC Method for simultaneous estimation of Sofosbuvir and Daclatasvir available in market in the form of combination tablet. Samples were applied on HPTLC aluminium plates precoated with silica gel 60 F254 (250μm thickness). Mobile phase consists of ethyl acetate: isopropanol in the ratio 9:1v/v. A good resolution was observed between peaks of both the drugs. The retention factor for Sofosbuvir is about 0.51 ±0.02. And for Daclatasvir is about 0.30 ± 0.02. Deuterium lamp as a source of radiation at the wavelength of 260 and 318 nm for analysis of Sofosbuvir and Daclatasvir, respectively. The proposed method was validated according to ICH guidelines and the results were acceptable for linearity and range, accuracy, precision, robustness, detection limit and quantitation limit. The calibration curves were linear over a wide range of 200-1000 ng/band (r2= 0.991) for Sofosbuvir and 45-225 ng/band (r2=0.990) for Daclatasvir. The limit of detection was found to be 21.17 ng/band and 4.38 ng/band for SOF and DAC and limit of quantitation was 64.18 ng/band and 13.28 ng/band for SOF and DAC respectively. During stress degradation study, it was observed that the Daclatasvir is degraded less in thermal and photolytic condition but more in basic hydrolysis condition. Sofosbuvir was found to be sensitive to all stress conditions except the fluorescent light. The suggested method was successfully applied for analysis of both drugs and excellent recovery results were obtained. Being simple, fast, robust, and economic, the method could be applied to the quality control and routine stability monitoring of Sofosbuvir and Daclatasvir.

Analytical quality risk management and DoE based development of robust chromatographic method for simultaneous estimation of tizanidine hydrochloride and nimesulide in their combined pharmaceutical dosage forms

Analytical quality risk management (ICH Q9 guideline) concept based robust high performance thin layer chromatographic method has been developed with help of design of experiment (DoE) tool for simultaneous estimation of tizanidine hydrochloride and nimesulide in their combined pharmaceutical dosage forms. Risk identification and assessment were done with brainstorming process with help of ishikawa diagram and experimental results based risk factor priority number (RPN). Critical risk factors which having RPN number more than sixty were further analysed for their criticality in method development by DoE based Taguchi screening design. From seven critical risk factors, volume of methanol in mobile phase and migration distance were identified as highly risky factors for development of HPTLC method. DoE based central composite design was applied for risk factors analysis and mitigation by optimisation of identified high risk factors. After implementation of risk minimisation operable design region and control strategy were set for development of robust HPTLC method for simultaneous estimation of tizanidine hydrochloride and nimesulide. Developed analytical method was validated for specificity, linearity, accuracy, precision and LOD-LOQ as per ICH guideline Q2R1. Validated HPTLC method was applied for assay of combined marketed pharmaceutical dosage forms of both drugs and results were found in good agreement with labelled claim of dosage forms.

Study of Intrinsic Stability of Mometasone Furoate in Presence of Salicylic Acid by HPTLC and Characterization, Cytotoxicity Testing of Major Degradation Product of Mometasone Furoate

Background: A successful attempt has been done to develop and validate a simple stability indicating HPTLC method for the estimation of Mometasone furoate (MF) and its degradation product in the presence of Salicylic acid (SA). The degradation product was isolated, characterized and tested for cytotoxicity. Introduction: Mometasone furoate (MF) is chemically 9,21-Dichloro-17α-[(2-furanylcarbonyl) oxy]- 11β-hydroxy-16-α-methylpregna-1,4-diene-3,20-dione, a high potency glucocorticoid. Salicylic acid (SA) has antiseptic, antifungal and keratolytic properties. Combination of MF and SA is available in the market as an ointment and is used for the treatment of skin inflammation, skin diseases, acne, skin redness and other conditions. Till now, there is no scientific documentation on HPTLC method for simultaneous estimation of MF and SA in the topical formulation; stress testing of drugs and determination of degradation products. Methods: Combination of Toluene: Ethyl Acetate: Methanol: Ammonia (6.4:1.5:2.0:0.1) was selected as the mobile phase. Detection was done by UV absorbance mode at wavelength 250 nm. Topical formulation containing MF and SA was analyzed by the developed method. The developed method was validated as per ICH guidelines. The standard drugs were subjected to stress testing like hydrolysis, oxidative, thermal and photolytic degradation. Results: Good separation with Rf values 0.61 ± 0.02 (MF) and 0.21 ± 0.02 (SA) was achieved by optimized chromatographic conditions. The % drug content was found to be 97.41±1.15 and 99.43 ± 0.73 for MF and SA, respectively in a topical formulation. From the results of validation parameters, the developed method was found to be specific, accurate, precise, sensitive and robust. After stress testing, SA was found to be stable under different stress conditions. Whereas, MF was found to be base sensitive and single degradation product was observed and isolated by preparative TLC. It was characterized by LC-MS and LC-MS/MS studies. Isolated degradation product was subjected to cytotoxicity testing on A549 and SiHa cell lines. Conclusion: A simple stability indicating HPTLC method was developed and validated for the estimation of MF and its degradation product in presence of SA. Probable structure of degradation product of MF and probable pathway of degradation was interpreted. Results of cytotoxicity testing showed that the degradation product was more cytotoxic as compared to MF against both the cell lines.

Validated HPTLC method for simultaneous estimation of metoprolol succinate and ramipril in bulk drug and marketed formulation

Validated HPTLC method for simultaneous estimation of metoprolol succinate and ramipril in bulk drug and marketed formulation

Validated HPTLC method for simultaneous estimation of metoprolol succinate and ramipril in bulk drug and marketed formulation

Validated HPTLC method for simultaneous estimation of metoprolol succinate and ramipril in bulk drug and marketed formulation

ANALYTICAL METHOD DEVELOPMENT AND ITS VALIDATION FOR EVALUATION OF POLYHERBAL FORMULATION

Talekt capsule is a branded polyherbal formulation used for enhancing the immune response to dermal infections and also for treating skin disorders. Curcumin and embelin, which are the active constituents of Haridra and Vidanga, respectively were used as marker compounds for developing a simple, accurate, precise and robust HPTLC method for simultaneous estimation. The mobile phase of toluene: ethyl acetate: formic acid (6.5: 3: 0.2 v/v/v) was used for separation. 381nm was used as wavelength for analysis. The Rf value obtained for curcumin, and embelin was found to be 0.51 ± 0.2 and 0.33 ± 0.2, respectively. The developed method was validated on the basis of ICH Q2 (R1) guidelines.

Stability Indicating HPTLC Method for Simultaneous Estimation of Sacubitril and Valsartan in their Pharmaceutical Dosage Form

A simple, precise, accurate and rapid high performance thin layer chromatographic method has been developed and completely validated for the estimation of Valsartan and Sacubitril in pharmaceutical dosage forms. Quantification of Valsartan and Sacubitril were carried out with precoated silica gel aluminum Plate 60 F254, (20 cm ×10 cm) 100 μm thickness as stationary phase. Toluene: Methanol: Ethyl Acetate: Glacial Acetic acid (8:2:1:1 % v/v/v). The optimized chamber saturation time before chromatographic development was 20 min at room temperature (25ºC ± 2). The length of chromatographic run was 8 cm which took average 15 min to develop. Densitometry scanning was performed using Camag TLC scanner IV with Win CATS software (V 1.4.6.2002, Camag). Optimized chromatogram of Valsartan and Sacubitril having Rf of 0.43 and 0.33, respectively. Linear relationship between peak area and concentration of standard was evaluated over the concentration range expressed in ng/band by making five replicate measurements in the concentrations range of 400-1400 ng/band and 4000-14,000 ng/band, respectively for Valsartan and Sacubitril. Literature reveals that there was no any method developed using HPTLC in simultaneous estimation of Valsartan and Sacubitril in their pharmaceutical dosage form. The proposed method can be successfully applied for the estimation of drug content of different marketed formulations.

Analytical method development and its validation for simultaneous estimation of catechin and curcumin by HPTLC from ancho lean tablets

HPTLC method which is simple, particular and robust has been developed for the simultaneous estimation of Catechin and Curcumin from an Ayurvedic formulation. The method was validated using parameters such as linearity, specificity, and precision, limit of quantification (LOQ), limit of detection (LOD), accuracy and robustness as per ICH guidelines. The present work deals with development of HPTLC method for simultaneous estimation of catechin and curcumin in marketed Ayurvedic formulations. Chromatographic separation of the drugs was performed on Merck TLC aluminium plates pre-coated with silica gel 60F254 as the stationary phase. The mobile phase selected was toluene:ethyl acetate: formic acid (7: 2.5: 0.5 v/v/v). The sample solutions were prepared in methanol and linear ascending development was carried out in twin trough glass chamber and scanned at 269 nm using Camag TLC scanner. The two markers were resolved successfully with Rf values 0.23±0.02 and 0.58±0.02 for catechin and curcumin, respectively. The regression analysis data indicated good linear relationship for the calibration plots for Catechin and Curcumin in the range of 1900-2500 ng/spot and 200-800 ng/spot and regression coefficient was 0.990 and 0.997 respectively. The proposed method can be used for the estimation of these markers in combined Ayurvedic formulation.

Simultaneous estimation of Gallic acid and rutin in Kaishore guggulu and Vatari guggulu by HPTLC

A simple, rapid, robust and reliable HPTLC method has been developed for simultaneous estimation of gallic acid and rutin in Kaishore Guggulu and Vatari Guggulu. Identification and quantification were performed on 20cm x 10cm, layer thickness 0.2mm, aluminium-backed silica gel 60 F254 HPTLC plates previously washed with methanol. Toluene: Acetone: Ethyl Acetate: Formic Acid: Water (2: 3: 2: 1: 2, v/v), was used as a mobile phase. Calibration plots were established showing the dependence of response on the amount chromatographed. The validated calibration range was 400-800ng per spot (R2 = 0.998; 0.996). The spots were scanned at λ = 312 nm. The suitability of this HPTLC method for simultaneous estimation of the marker constituents was proved by validation in accordance with ICH Guidelines. Determination of method accuracy by the standard addition method at three concentration levels returned a mean recovery of 98.92 ± 0.16–101.61 ± 0.24. The developed method has the advantage of being rapid, robust and easy. Hence, it can be applied for routine quality control analysis of gallic acid and rutin in those Ayurvedic formulations.