Development and Validation of an HPLC Method for Simultaneous Determination of Miconazole Nitrate and Chlorhexidine Digluconate in Chitosan-Based Gel Formulations

Year 2023, Volume: 48 Issue: 1, 75 - 90, 01.03.2023

Ece Türkmen Selin Parmaksız Mustafa Çelebier Sevda Şenel

https://doi.org/10.55262/fabadeczacilik.1133276

Cited By: 1

Abstract

Miconazole nitrate (MN) and chlorhexidine digluconate (CHX) are the commonly used antimicrobials for topical treatment of dermal infections. Combination of antimicrobials has been investigated to enhance the efficacy of the treatment. Gel formulations based on bioadhesive polymers are preferred for delivery of these drugs. Chitosan is a promising bioadhesive polymer due to its penetration enhancing, antimicrobial and tissue healing properties. Yet, most of the gel-based formulations present analytical challenges during testing the drug content. It was aimed to develop an HPLC method for simultaneous determination of MN and CHX in chitosan-based gel formulations. Different solvent combinations were investigated for extraction of drugs from the gels. HPLC conditions such as mobile phase, flow rate, run time, column temperature and wavelength were explored. The method was validated according to ICH guideline Q2(R1). MN and CHX were extracted in solvent composition same with the mobile phase. The method was employed on ACE-C8 column at 40°C by isocratic elution using the mobile phase consisting of methanol:phosphate (75:25 v/v) buffer (containing triethylamine). Flow rate was 1 mL/min. The drugs were detected at 254 nm (CHX) and 230 nm (MN). Linearity was obtained between 5 to 80 μg/mL for both drugs. LOD and LOQ obtained for CHX was 1.61 and 1.06, for MN: 4.87 and 3.21 µg/mL, respectively. A new validated HPLC method was developed for simultaneous determination of CHX and MN in chitosan-based gels, with 98 to 102% recovery, without any interference with the excipients.

Keywords

HPLC method, simultaneous analysis, miconazole nitrate, chlorhexidine digluconate, chitosan gel, validation

Supporting Institution

Hacettepe Üniversitesi Bilimsel Araştırma Projeleri Koordinasyon Birimi

Project Number

THD-2020-18516

Thanks

Authors are thankful to IE Ulagay–Menarini, İstanbul, Turkey for providing Miconazole nitrate and Koyo Company, Japan for providing chitosan as a generous gift. This work was supported by Hacettepe University Scientific Research Coordination Unit, Project number: THD-2020-18516

References

• Abdelrahman, M. M., Naguib, I., Nagieb, H. M., & Zaazaa, H. E. (2017). Different spectrophotometric methods for determination of miconazole nitrate and hydrocortisone in bulk and in topical pharmaceutical preparation without prior separation. Chem Res J, 2(2), 56-65.
• Abdelrahman, M. M., Naguib, I. A., Elsayed, M. A., & Zaazaa, H. A. (2016). Spectrophotometric Methods for Quantitative Determination of Chlorhexidine Gluconate and its Major Impurity, Metabolite and Degradation Product: Para-chloro-aniline. Anal Chem Lett, 6(3), 232-248. doi:https://doi.org/10.1080/22297928.2016.1196148
• Abtheen, K. S. S. A., Maheswari, R., Shanmugasundaram, P., & Vijeyaanandhi, M. (2008). Simultaneous estimation of chlorhexidine gluconate, metronidazole, lignocaine hydrochloride and triamcinolone acetonide in combined dosage form by RP-HPLC. Asian J Chem, 20(2), 1130-1136. Retrieved from https://www.scopus.com/inward/record.uri?eid=2-s2.0-53949098973&partnerID=40&md5=63af943945eaed54876a38068fa49367
• Al-Badr, A. A. (2005). Miconazole Nitrate: Comprehensive Profile. In H. G. Brittain (Ed.), Profiles of Drug Substances, Excipients and Related Methodology (pp. 3-65): Academic Press.
• Ammara, S., Syed, H. K., Sajid, A., Muhammad, I., Ikram, u. K., Akhtar, R., . . . Khizar, A. (2018). Miconazole Nitrate Microemulsion: Preparation, Characterization and Evaluation for Enhancement of Antifungal Activity. Lat Am J Pharm, 37, 1578-1586.
• Atiyeh, B. S., Dibo, S. A., & Hayek, S. N. (2009). Wound cleansing, topical antiseptics and wound healing. Int Wound J, 6(6), 420-430. doi:https://doi.org/10.1111/j.1742-481X.2009.00639.x
• Belal, T. S., & Haggag, R. S. (2012). Gradient HPLC-DAD stability indicating determination of miconazole nitrate and lidocaine hydrochloride in their combined oral gel dosage form. J Chromatogr Sci, 50(5), 401-409. doi:https://doi.org/10.1093/chromsci/bms019
• Borissova, R., & Mandjukova, S. (1997). Titrimetric and spectrophotometric determination of chlorhexidine digluconate in tooth pastes. Fresenius J Anal Chem, 357(7), 977-980. doi:https://doi.org/10.1007/s002160050285
• Bouckaert, S., & Remon, J. P. (1993). In-vitro bioadhesion of a buccal, miconazole slow-release tablet. J Pharm Pharmacol, 45(6), 504-507. doi:https://doi.org/10.1111/j.2042-7158.1993.tb05588.x
• Brookes, Z. L. S., Bescos, R., Belfield, L. A., Ali, K., & Roberts, A. (2020). Current uses of chlorhexidine for management of oral disease: a narrative review. J Dent, 103, 103497. doi:https://doi.org/10.1016/j.jdent.2020.103497
• Cartagena, A. F., Lyra, A. M., Kapuchczinski, A. C., Urban, A. M., Esmerino, L. A., Klein, T., . . . Campanha, N. H. (2017). Miconazole Nitrate-loaded Microparticles For Buccal Use: Immediate Drug Release and Antifungal Effect. Curr Drug Deliv, 14(8), 1144-1153. doi:https://doi.org/10.2174/1567201813666161006115041
• Chiapetta, S., Oliveira, É., Olivier, B., Mercante, L., Henriques, D., & Pereira Netto, A. (2011). Intralaboratory Validation, Comparison and Application of HPLC-UV-DAD Methods for Simultaneous Determination of Benzalkonium Chloride, Chlorexidine Digluconate and Triclosan. J Braz Chem Soc, 22, 1913-1920. doi:https://doi.org/10.1590/S0103-50532011001000012
• Chlorhexidine. (2016). In J. K. Aronson (Ed.), Meyler's Side Effects of Drugs (Sixteenth Edition) (pp. 239-248). Oxford: Elsevier.
• Denton, G. W. (2001). Chapter 15 - Chlorhexidine. In S. S. Block (Ed.), Disinfection, sterilization, and preservation (5th ed., pp. 321-333). Philadelphia: Lippincott Williams & Wilkins.
• Dimopoulou, M., Mourouti, C.-S., Vertzoni, M., Symillides, M., & Reppas, C. (2015). In-vitro evaluation of performance of solid immediate release dosage forms of weak bases in upper gastrointestinal lumen: experience with miconazole and clopidogrel salts. J Pharm Pharmacol, 68(5), 579-587. doi:https://doi.org/10.1111/jphp.12406
• Ei, Z., Pimthon, J., Vajragupta, O., Leanpolchareanchai, J., & Phechkrajang, C. (2016). Development and validation of high-performance liquid chromatography method for determination of miconazole , triamcinolone , methylparaben and propylparaben in cream. Mahidol Univ J Pharm Sci, 43(5), 211-221. doi:https://doi.org/10.14456/mujps.2016.24
• Eticha, T., Kahsay, G., Hailu, T., Gebretsadikan, T., Asefa, F., Gebretsadik, H., & Thangabalan, B. (2018). Development and Validation of an Extractive Spectrophotometric Method for Miconazole Nitrate Assay in Pharmaceutical Formulations. J Anal Methods Chem, 2018, 1-5. doi:https://doi.org/10.1155/2018/2191072
• European Medicines Agency. ICH Topic Q 2 (R1) Validation of Analytical Procedures: Text and Methodology. (1995). Retrieved from https://www.ema.europa.eu/en/documents/scientific-guideline/ich-q-2-r1-validation-analytical-procedures-text-methodology-step-5_en.pdf.
• Facts about Chlorhexidine Gluconate. (2017). Retrieved from https://www.fda.gov/drugs/drug-safety-and-availability/fda-drug-safety-communication-fda-warns-about-rare-serious-allergic-reactions-skin-antiseptic#:~:text=Facts%20about%20Chlorhexidine%20Gluconate,-Chlorhexidine%20gluconate%20is&text=The%20OTC%20products%20are%20available,Pharmaseal%20Scrub%20Care%2C%20and%20Prevantics.
• Final Report on the Safety Assessment of Chlorhexidine/Chlorhexidine Diacetate/Chlorhexidine Dihydrochloride/Chlorhexidine Digluconate (1993). J Am Coll Toxicol, 12(3), 201-223. doi:https://doi.org/10.3109/10915819309140642
• Frymus, T., Gruffydd-Jones, T., Pennisi, M. G., Addie, D., Belák, S., Boucraut-Baralon, C., . . . Horzinek, M. C. (2013). Dermatophytosis in Cats: ABCD guidelines on prevention and management. J Feline Med Surg, 15(7), 598-604. doi:https://doi.org/10.1177/1098612X13489222
• Ghibaudo, G., Santospirito, D., Sala, A., Flisi, S., Taddei, S., Cavirani, S., & Cabassi, C. S. (2016). In vitro antimicrobial activity of a gel containing antimicrobial peptide AMP2041, chlorhexidine digluconate and Tris-EDTA on clinical isolates of Pseudomonas aeruginosa from canine otitis. Vet Dermatol, 27(5), 391-e398. doi: https://doi.org/10.1111/vde.12371
• Greenstein, G., Berman, C., & Jaffin, R. (1986). Chlorhexidine: An Adjunct to Periodontal Therapy. J Periodontol, 57(6), 370-377. doi:https://doi.org/10.1902/jop.1986.57.6.370
• Guaguère, E. (1996). Topical treatment of canine and feline pyoderma. Vet Dermatol, 7, 145-151. doi:https://doi.org/10.1111/j.1365-3164.1996.tb00239.x
• Guardabassi, L., Ghibaudo, G., & Damborg, P. (2010). In vitro antimicrobial activity of a commercial ear antiseptic containing chlorhexidine and Tris-EDTA. Vet Dermatol, 21(3), 282-286. doi:10.1111/j.1365-3164.2009.00812.x
• Havlíková, L., Matysová, L., Nováková, L., Hájková, R., & Solich, P. (2007). HPLC determination of chlorhexidine gluconate and p-chloroaniline in topical ointment. J Pharm Biomed Anal, 43(3), 1169-1173. doi:https://doi.org/10.1016/j.jpba.2006.09.037
• Heneedak, H. M., Salama, I., Mostafa, S., & El-Sadek, M. (2012). HPLC and chemometric methods for the simultaneous determination of miconazole nitrate and nystatin. J Chromatogr Sci, 50(10), 855-861. doi:https://doi.org/10.1093/chromsci/bms127
• Hoang, T. P. N., Ghori, M. U., & Conway, B. R. (2021). Topical Antiseptic Formulations for Skin and Soft Tissue Infections. Pharmaceutics, 13(4), 558. doi:https://doi.org/10.3390/pharmaceutics13040558
• Işık, B. D., & Acar, E. T. (2018). Development and Validation of an HPLC Method for the Simultaneous Determination of Flurbiprofen and Chlorhexidine Gluconate. Chromatographia, 81(4), 699-706. doi:https://doi.org/10.1007/s10337-018-3485-5
• Kampf, G. (2018). Chlorhexidine Digluconate. In G. Kampf (Ed.), Antiseptic Stewardship: Biocide Resistance and Clinical Implications (pp. 429-534). Cham: Springer International Publishing.
• Kenechukwu, F. C., Attama, A. A., Ibezim, E. C., Nnamani, P. O., Umeyor, C. E., Uronnachi, E. M., . . . Akpa, P. A. (2018). Surface-modified mucoadhesive microgels as a controlled release system for miconazole nitrate to improve localized treatment of vulvovaginal candidiasis. Eur J Pharm Sci, 111, 358-375. doi:https://doi.org/10.1016/j.ejps.2017.10.002
• Lakka, N. S., & Kuppan, C. (2019). Principles of Chromatography Method Development. In O.-M. Boldura, C. Baltă, & N. S. Awwad (Eds.), Biochemical Analysis Tools - Methods for Bio-Molecules Studies. London: IntechOpen.
• Main, R. C. (2008). Should chlorhexidine gluconate be used in wound cleansing? J Wound Care, 17(3), 112-114. doi:https://doi.org/10.12968/jowc.2008.17.3.28668
• Martindale: The Complete Drug Reference. (2009). (36th ed.). London: Pharmaceutical Press.
• Másquio Fiorentino, F. A., Corrêa, M. A., & Nunes Salgado, H. R. (2010). Analytical Methods for the Determination of Chlorhexidine: A Review. Crit Rev Anal Chem, 40(2), 89-101. doi:https://doi.org/10.1080/10408340903232020
• Mohammadi, Z. (2008). Chlorhexidine gluconate, its properties and applications in endodontics. Iran Endod J, 2(4), 113-125.
• Moldoveanu, S. C., & David, V. (2017). Chapter 13 - Solvents, Buffers, and Additives Used in the Mobile Phase. In S. C. Moldoveanu & V. David (Eds.), Selection of the HPLC Method in Chemical Analysis (1st ed., pp. 393-450). Boston: Elsevier.
• Mueller, R. S. (2008). Chapter 24 - Topical dermatological therapy. In J. E. Maddison, S. W. Page, & D. B. Church (Eds.), Small Animal Clinical Pharmacology (2nd ed., pp. 546-556). Edinburgh: W.B. Saunders.
• Nenoff, P., Koch, D., Krüger, C., Drechsel, C., & Mayser, P. (2017). New insights on the antibacterial efficacy of miconazole in vitro. Mycoses, 60, 552 —557. doi:https://doi.org/10.1111/myc.12620
• Pasquetti, M., Min, A. R. M., Scacchetti, S., Dogliero, A., & Peano, A. (2017). Infection by Microsporum canis in Paediatric Patients: A Veterinary Perspective. Veterinary sciences, 4(3), 46. doi:10.3390/vetsci4030046
• Paulson, D. S. (2002). Chlorhexidine Gluconate. In D. S. Paulson (Ed.), Handbook of Topical Antimicrobials (1st ed., pp. 117-122). New York: CRC Press.
• Periochip. What Is Periochip. Retrieved from https://www.periochip.com/what-is-periochip/.
• Perrins, N., & Bond, R. (2003). Synergistic inhibition of the growth in vitro of Microsporum canis by miconazole and chlorhexidine. Vet Dermatol, 14(2), 99-102. doi:10.1046/j.1365-3164.2003.00325.x
• Qushawy, M., Nasr, A., Abd-Alhaseeb, M., & Swidan, S. (2018). Design, Optimization and Characterization of a Transfersomal Gel Using Miconazole Nitrate for the Treatment of Candida Skin Infections. Pharmaceutics, 10(1), 26. doi:https://doi.org/10.3390/pharmaceutics10010026
• Rawlings, J. M., Gorrel, C., & Markwell, P. J. (1998). Effect on Canine Oral Health of Adding Chlorhexidine to a Dental Hygiene Chew. J Vet Dent, 15(3), 129-134. doi:https://doi.org/10.1177/089875649801500303
• Rochette, F., Engelen, M., & Vanden Bossche, H. (2003). Antifungal agents of use in animal health--practical applications. J Vet Pharmacol Ther, 26(1), 31-53. doi:https://doi.org/10.1046/j.1365-2885.2003.00457.x
• Sahoo, D. R., & Jain, S. (2016). A Rapid and Validated RP-HPLC Method for the Simultaneous Quantification of Benzoic Acid, Metronidazole and Miconazole Nitrate in Vaginal Formulations. J Chromatogr Sci, 54(9), 1613-1618. doi: https://doi.org/10.1093/chromsci/bmw113
• Salah, S., Awad, G. E. A., & Makhlouf, A. I. A. (2018). Improved vaginal retention and enhanced antifungal activity of miconazole microsponges gel: Formulation development and in vivo therapeutic efficacy in rats. Eur J Pharm Sci, 114, 255-266. doi:https://doi.org/10.1016/j.ejps.2017.12.023
• Samanidou, V. F. (2015). Basic LC Method Development and Optimization. In J. L. Anderson, A. Berthod, V. Pino, & A. Stalcup (Eds.), Analytical Separation Science (1st ed., pp. 25-42). Weinheim: Wiley-VCH Verlag GmbH & Co. KGaA.
• Sarkiala-Kessel, E. M. (2012). Chapter 3 - Use of antibiotics and antiseptics. In F. J. Verstraete & M. J. Lommer (Eds.), Oral and Maxillofacial Surgery in Dogs and Cats (1st ed., pp. 15-21). Oxford: W.B. Saunders.
• Sawyer, P. R., Brogden, R. N., Pinder, R. M., Speight, T. M., & Avery, G. S. (1975). Miconazole: A Review of its Antifungal Activity and Therapeutic Efficacy. Drugs, 9(6), 406-423. doi:https://doi.org/10.2165/00003495-197509060-00002
• Şenel, S. (2010). Potential applications of chitosan in oral mucosal delivery. J Drug Deliv Sci Technol, 20(1), 23-32. doi:https://doi.org/10.1016/S1773-2247(10)50003-0
• Şenel, S. (2020). Current status and future of chitosan in drug and vaccine delivery. React Funct Polym, 147, 104452. doi:https://doi.org/10.1016/j.reactfunctpolym.2019.104452
• Şenel, S., İkinci, G., Kaş, S., Yousefi-Rad, A., Sargon, M. F., & Hıncal, A. A. (2000). Chitosan films and hydrogels of chlorhexidine gluconate for oral mucosal delivery. Int J Pharm, 193(2), 197-203. doi:https://doi.org/10.1016/S0378-5173(99)00334-8
• Siegel, M. R., Kerkenaar, A., & Sijpesteijn, A. K. (1977). Antifungal activity of the systemic fungicide imazalil. Neth J PI Path, 83(1), 121-133. doi:https://doi.org/10.1007/BF03041427
• Silvestri, D. L., & McEnery-Stonelake, M. (2013). Chlorhexidine: Uses and Adverse Reactions. Dermatitis, 24(3), 112 - 118. doi:https://doi.org/10.1097/DER.0b013e3182905561
• Stojanov, I., Milovanovic, A., RuŽIĆ-MusliĆ, D., Ratajac, R., Baloš, M., Maksimovic, N., & Apić, J. (2018). The application of EDTA-Tris and chlorhexidine in the treatment of endometritis as a replacement for antibiotic therapy in cows. Turk J Vet Anim Sci, 42, 91-96. doi:https://doi.org/10.3906/vet-1703-74
• Tejada, G., Lamas, M. C., Svetaz, L., Salomón, C. J., Alvarez, V. A., & Leonardi, D. (2018). Effect of drug incorporation technique and polymer combination on the performance of biopolymeric antifungal buccal films. Int J Pharm, 548(1), 431-442. doi:https://doi.org/10.1016/j.ijpharm.2018.07.023
• Valkó, K., Snyder, L. R., & Glajch, J. L. (1993). Retention in reversed-phase liquid chromatography as a function of mobile-phase composition. J Chromatogr A, 656(1), 501-520. doi:https://doi.org/10.1016/0021-9673(93)80816-Q
• Velegraki, A., Cafarchia, C., Gaitanis, G., Iatta, R., & Boekhout, T. (2015). Malassezia Infections in Humans and Animals: Pathophysiology, Detection, and Treatment. PLOS Pathogens, 11(1), e1004523. doi:10.1371/journal.ppat.1004523
• Wade, T. R., Jones, H. E., & Chanda, J. J. (1979). Intravenous miconazole therapy of mycotic infections. Arch Intern Med, 139(7), 784-786. doi:https://doi.org/10.1001/archinte.1979.03630440046016
• Zeng, P., Rao, A., Wiedmann, T. S., & Bowles, W. (2009). Solubility Properties of Chlorhexidine Salts. Drug Dev Ind Pharm, 35(2), 172-176. doi:https://doi.org/10.1080/03639040802220318