Doe-based Rp-hplc Method Development and Validation for Estimation of Preservative Phenoxyethanol in the Presence of Adapalene in an Anti-acne Formulation Using Principles of White Analytical Chemistry
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Abstract
Introduction: A simple and economical reversed-phase high-performance liquid chromatography (RP-HPLC) method was developed and validated as per ICH guidelines to determine Phenoxyethanol in the presence of Adapalene. This study aimed to create a robust and environment friendly method for simultaneous analysis of these drugs in pharmaceutical formulations. Methods: The method employed C18 column as a stationary phase, with Acetonitrile: Water as a mobile phase at a flow rate of 1.2 mL/min and a detection wavelength of 272 nm. Validation parameters included linearity, sensitivity, accuracy, and precision. A Design of Experiments (DoE) approach, specifically the Box-Behnken design, was used to evaluate the effect of Critical Method Variables (CMVs) flow rate, wavelength, and acetonitrile proportion on Critical Analytical Attributes (CAAs) retention time, tailing factor, and theoretical plates. The Red, Green, Blue (RGB) model used for White analytical Chemistry (WAC) assessment of published and proposed method. Results: The method demonstrated excellent linearity and sensitivity for both compounds, with high accuracy and precision under varied conditions. Robustness was confirmed through the DoE approach, minimal variability observed in key chromatographic parameters. The WAC score was calculated for published method (72.33) and proposed method (88.33). Conclusion: The developed RP-HPLC method was simple, economical, and robust, suitable for routine quality control analysis of Adapalene and Phenoxyethanol. Its usability, validity, cost and time effectiveness evaluated by WAC approach enhances its applicability as a sustainable method for pharmaceutical quality assurance.
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Waugh J, Noble S, Scott L. Adapalene: a review of its use in the treatment of acne vulgaris. Drugs. 2004;64(13):1465–78.
Kosmadaki M, Katsambas A. Topical treatments for acne. Clin Dermatol. 2017;35(2):173–8. doi:10.1016/j.clindermatol.2016.10.010.
PubChem (Internet). AD (cited 2025 Jan 29). Available from: https://pubchem.ncbi.nlm.nih.gov/compound/AD.
Tripathi KD. Essentials of Medical Pharmacology. 6th ed. New Delhi: Jaypee Brothers Medical Publishers; 2008. p. 853.
U.S. Pharmacopeia National Formulary (USP 42-NF 37). Volume I. Rockville (MD): The United States Pharmacopeial Convention; 2019. p. 95–7 and 567–72.
British Pharmacopoeia. Volume I. London: Medicine and Healthcare Products Regulatory Agency (MHRA); 2019. p. 74–5.
Modi PB. Novel stability-indicating RP-HPLC method for the simultaneous estimation of clindamycin phosphate and AD along with preservatives in topical gel formulations. Scientia Pharmaceutica. 2014;82(4):799–813. doi:10.3797/scipharm.1404-01.
Bajaj A, John C, Tripathi RM, Choudhury J. Validated RP-capillary HPLC-DAD method for simultaneous analysis of some paraben preservatives in pharmaceutical and cosmetics/personal care formulations. Asian J Pharm Ana. 2017;7(4):229–34. doi:10.5958/2231-5675.2017.00037.
Głaz P, Rosińska A, Woźniak S, Boguszewska-Czubara A, Biernasiuk A, Matosiuk D. Effect of commonly used cosmetic preservatives on healthy human skin cells. Cells. 2023 Apr 3;12(7):1076. doi:10.3390/cells12071076.
Adhikari L, Jagadev S, Sahu S, Moitra SK, Murthy PN. Derivative spectrophotometric determination of AD in its bulk and pharmaceutical dosage form. Asian Journal of Chemistry. 2012;24:1094–6.
Doyle RM, Kline J. LC-MS-MS quantitative analysis of 12 retinoids, derivatives, and metabolites in serum for clinical research use. Thermo Fisher Scientific Inc. (Internet). Available from: PO-65009-LC-MS-VitA-Serum-ASMS2017-PO65009-EN.pdf.
Dobricic V, Pajic N, Markovic B, Vladimirov S, Savic S, Vuleta G. Development and validation of an LC-MS/MS method for the determination of AD in pharmaceutical forms for skin application. Journal of the Serbian Chemical Society. 2016;81:1171–81.
Thummar K, Tilva K, Dudhatra B, Mardia R, Sheth N. Validated stability-indicating HPTLC method for the estimation of AD in drugs and the LC-MS identification of its degradation products. Journal of Planar Chromatography. 2020;1–10. doi:10.1007/s00764-020-00042-z.
Roy C. Validated stability-indicating RP-HPLC method for estimation of degradation behaviour of organic peroxide and third-generation synthetic retinoids in topical pharmaceutical dosage formulation. Scientia Pharmaceutica. 2015;83:321–38.
Mailvelan R, Selvamani P, Rameshkumar T, Raviraj T. HPLC method development and validation for the estimation of AD in pharmaceutical formulations. Asian J Pharm Anal Med Chem. 2013;1:166–71.
Mudasir M, Tabassum N, Ali J, Jan R. Estimation of AD through isocratic HPLC method in pharmaceutical gel formulations. Analytical Chemistry an Indian Journal. 2018;7:801–6.
Martins LA, Meneghini LZ, Junqueira CA, Ceni DC, Bergold AM. A simple HPLC-DAD method for determination of AD in topical gel formulation. J Chromatogr Sci. 2011;49:796–800.
Jamkhandi C, Disouza J, Dashwant V, Pisal S. Development of a conductometric method of estimation for AD gel in dosage forms. Asian J Biochem Pharm Res. 2013;3:136–9.
Tolba MM, El-Gamal RM. Determination of AD in gel formulation by conventional and derivative synchronous fluorimetric approaches. Application to stability studies and in vitro diffusion test. Chem Cent J. 2016;10:33. doi:10.1186/s13065-016-0181-0.
Vichare V, Neve T, Choudhary N, Dhole S, Choudhari V. Development and validation of a high-performance thin-layer chromatography method for the quantification of adapalene and preservative phenoxyethanol in gel formulation—application to stability studies. Journal of Planar Chromatography - Modern TLC. 2024;37. doi:10.1007/s00764-024-00285-0.
Zhang C, Zhao Y, Han C, Guo X. Simultaneous determination of AD, 2-PE, and methyl-4-hydroxybenzoate in AD gels using high-performance liquid chromatography. Chinese Journal of Chromatography. 2008;26:640–2.
New Jersey Department of Health and Senior Services. Hazardous Substance Fact Sheet (Internet). Available from: https://Anyflip.Com/Xflkp/Xikt.
Safety Data Sheet: Tetrahydrofuran (Internet). Available from: https://www.scribd.com/document/673083704/401757.
Material Safety Data Sheet: Tetrahydrofuran, 99.5+%, for spectroscopy (Internet). Available from: https://fscimage.fishersci.com/msds/23011.html.
Gupta PC. Method validation of analytical procedures. PharmaTutor. 2015;3:32–9.
International Conference on Harmonization (ICH). Q2(R1) Validation of analytical procedures: text and methodology. Nov 2005.
International Conference on Harmonization (ICH). Q1A(R2): Stability testing of new drug substances and products (revision 2). 2003.
International Conference on Harmonization (ICH). Q8(R2) Pharmaceutical development. 2009.
Thomas A. Design of experiments for analytical method development and validation. BioPharm International. 2014;27:40–5.
Ferreira SLC, Bruns RE, Ferreira HS, Matos GD, David JM, Brandão GC, et al. Box-Behnken design: An alternative for the optimization of analytical methods. Analytica Chimica Acta. 2007;597(2):179–86. doi:10.1016/j.aca.2007.07.011.
Garg NK, Sharma G, Singh B, Nirbhavane P, Katare OP. Quality by design (QbD)-based development and optimization of a simple, robust RP-HPLC method for the estimation of methotrexate. Journal of Liquid Chromatography and Related Technologies. 2015;38:1629–37. doi:10.1080/10826076.2015.1087409.
Mutalik SP, Mullick P, Pandey A, Kulkarni SS, Mutalik S. Box–Behnken design-aided optimization and validation of developed reverse phase HPLC analytical method for simultaneous quantification of dolutegravir sodium and lamivudine co‐loaded in nano‐liposomes. Journal of Separation Science. 2021;44:2917–31. doi:10.1002/jssc.202100152.
Pena-Pereira F, Wojnowski W, Tobiszewski M. AGREE—analytical greenness metric approach and software. Anal Chem. 2020;92:10076–82. doi:10.1021/acs.analchem.0c01887.
Darji H, Dedania Z. Simultaneous estimation of azelnidipine and metoprolol succinate with greenness assessment using HPLC and UV-spectrophotometric methods. Green Analytical Chemistry. 2023;7:100079. doi:10.1016/j.greeac.2023.100079.
Akabari AH, Patel SK, Shah KV, et al. Analytical quality by design (AQbD) methodology for concurrent determination of perindopril erbumine and moxonidine hydrochloride using RP-HPLC with an eco-friendly evaluation. Discov Chem. 2025;2(1):1. https://doi.org/10.1007/s44371-024-00067-x
Akabari SH, Gajiwala H, Patel S, Surati J, Solanki D, Shah KV, et al. Stability-indicating TLC-densitometric and HPLC methods for simultaneous determination of teneligliptin and pioglitazone in pharmaceutical dosage forms with eco-friendly assessment. J Chromatogr Sci. 2025;63(2):bmae038. https://doi.org/10.1093/chromsci/bmae038
Prajapati P, Shahi A, Acharya A, Pulusu VS, Shah S. Implementation of white analytical chemistry–assisted analytical quality by design approach to green liquid chromatographic method for concomitant analysis of anti-hypertensive drugs in human plasma. J Chromatogr Sci. 2024;62(10):938–52. doi: 10.1093/chromsci/bmad054.
Prajapati P, Rana B, Pulusu VS, Shah S et al. Multipurpose RP-HPLC method for simultaneous estimation of fixed-dose combinations of anti-diabetic drugs: integrating green, economical, and robust approaches with design of experiments and white analytical chemistry. Chem Afr. 2024;7:1385–400. doi: 10.1007/s42250-023-00835-9.
Prajapati P, Salunkhe M, Pulusu VS et al. Integrated approach of white analytical chemistry and analytical quality by design to multipurpose RP-HPLC method for synchronous estimation of multiple fixed-dose combinations of paracetamol. Chem Afr. 2024;7:1353–71. doi: 10.1007/s42250-023-00819-9.
Prajapati P, Rana B, Pulusu VS, Shah S. Simultaneous chromatographic estimation of vildagliptin and dapagliflozin using hybrid principles of white analytical chemistry and analytical quality by design. J AOAC Int. 2024;107(1):212–22. doi: 10.1093/jaoacint/qsad108.
Prajapati P, Shahi A, Acharya A, Pulusu VS, Shah S. Robust method operable design region for economical and eco-friendly chromatographic analysis of azilsartan medoxomil and cilnidipine by incorporating a hybrid approach of green analytical chemistry and analytical quality by design. Smart Sci Chem Pharm. 2023;6(11):2300111. doi: 10.1002/sscp.202300111.