Exploring the Bottom-up and Top-down Techniques of Nanosuspension for Enhanced Solubility and Drug Loading of Atovaquone

Karishma Patil; Rushikesh Ghongane; Tejas Nakte; Harsha Kathpalia

doi:10.26650/IstanbulJPharm.2025.1570740

Exploring the Bottom-up and Top-down Techniques of Nanosuspension for Enhanced Solubility and Drug Loading of Atovaquone

Abstract

Background and Aims: Atovaquone is a Class II drug under the Biopharmaceutical Classification System. It is one of the most commonly used drugs to treat pneumonia and malaria. It shows anti-protozoal and anti-pneumocystic activity, but its low water solubility results in poor bioavailability. This research aimed to prepare and analyse atovaquone nanosuspension using a combination of bottom-up and top-down techniques. Methods: The study explored the efficiency of two bottom-up methods, namely, pH-based and antisolvent- based precipitation, to achieve high drug loading in the nanosuspension. Tetrahydrofuran was used as the solvent to dissolve atovaquone in the presence of sodium hydroxide and povidone as solubilizers. The poloxamer was used as the wetting agent, and Phospholipon 90H as the stabilising agent. Citric acid was used as the acidifying agent for the precipitation. Results: High-pressure homogenisation in the top-down step after precipitation produced particles with a size of 195.2 nm. The nanosuspension showed 6% drug loading. The formulation was freeze-dried to improve stability and restrict crystal growth, resulting in a particle size of 343±20 nm. The aqueous solubility of atovaquone in the nanosuspension was 13 times higher, and the dissolution rate was increased by seven-fold compared with the unprocessed drug. The atovaquone nanosuspension prepared by this combination method was stable for 3 months. Conclusion: The study successfully demonstrated the potential of combination methods to obtain a nanosuspension with high drug loading and small particle size.

Keywords

References

Afifi, S. A., Hassan, M. A., Abdelhameed, A. S., & Elkhodairy, K. A. (2015). Nanosus-pension: an emerging trend for bioavailability enhancement of etodolac. International Journal of Polymer Science, 2015, 1–16. https://doi.org/10.1155/ 2015/938594 google scholar
Ahmadi Tehrani, A., Omranpoor, M. M., Vatanara, A., Seyedabadi, M., & Ramezani, V. (2019). Formation of nanosuspensions in bottom-up approach: theories and optimization. Daru : Journal of Faculty of Pharmacy, Tehran University of Medical Sciences, 27(1), 451–473. https://doi.org/10.1007/s40199-018-00235-2 google scholar
Al-Kassas, R., Bansal, M., & Shaw, J. (2017). Nanosizing techniques for improving bioavailability of drugs. Journal of Controlled Release, 260, 202–212. https://doi.org/10.1016/j.jconrel.2017.06.003 google scholar
Dearn, A. R. (2003). Atovaquone pharmaceutical compositions (U.S. Patent No. 6,649,659). U.S. Patent and Trademark Office google scholar
Baggish, A. L., & Hill, D. R. (2002). Antiparasitic Agent Atovaquone. Antimicrobial Agents and Chemotherapy, 46(5), 1163–1173. https://doi.org/10.1128/AAC.46.5.1163-1173.2002 google scholar
Bettinetti G. P., Sorrenti, M., Rossi, S., Ferrari, F., Mura, P., & Faucci, M. T. (2002). Assess-ment of solid-state interactions of naproxen with amorphous cyclodextrin derivatives by DSC. Journal of Pharmaceutical and Biomedical Analysis, 30(4), 1173–1179. https://doi.org/10.1016/S0731-7085(02)00421-1google scholar
Bilgili, E., & Guner, G. (2020). Mechanistic modelling of wet stirred media milling for production of drug nanosuspensions. AAPS PharmSciTech, 22(1), 2. https://doi. org/10.1208/s12249-020-01876-w google scholar
Borhade, V., Pathak, S., Sharma, S., & Patravale, V. (2014). Formulation and charac-terization of atovaquone nanosuspension for improved oral delivery in the treatment of malaria. Nanomedicine, 9(5), 649–666. https://doi.org/10.2217/ nnm.13.61 google scholar

Cauchetier, E., Deniau, M., Fessi, H., Astier, A., & Paul, M. (2003). Atovaquone-loaded nanocapsules: Influence of the nature of the polymer on their in vitro charac-teristics. International Journal of Pharmaceutics, 250(1), 273–281. https://doi. org/10.1016/S0378-5173(02)00556-2 google scholar
Cauchetier, E., Paul, M., Rivollet, D., Fessi, H., Astier, A., & Deniau, M. (2000). Therapeutic evaluation of free and liposome-encapsulated atovaquone in the treatment of murine leishmaniasis. International Journal for Parasitology, 30(6), 777–783. https://doi.org/10.1016/S0020-7519(00)00053-9 google scholar
Chogale, M., Ghodake, V., & Patravale, V. (2016). Performance Parameters and Charac-terizations of Nanocrystals: A Brief Review. Pharmaceutics, 8(3), 26. https:// doi.org/10.3390/pharmaceutics8030026google scholar
Darade, A., Pathak, S., Sharma, S., & Patravale, V. (2018). Atovaquone oral bioavail-ability enhancement using electrospraying technology. European Journal of Pharmaceutical Sciences, 111, 195–204. google scholar
Deng, J., Huang, L., & Liu, F. (2010). Understanding the Structure and Stability of Paclitaxel Nanocrystals. International Journal of Pharmaceutics, 390(2), 242-249. https://doi.org/10.1016/j.ijpharm.2010.02.013 google scholar
Dolenc, A., Kristl, J., Baumgartner, S., & Planinšek, O. (2009). Advantages of celecoxib nanosuspension formulation and transformation into tablets. International Journal of Pharmaceutics, 376(1–2), 204–212. https://doi.org/10.1016/j.ijpharm.2009.04.038google scholar
El Hage, S., Ane, M., Stigliani, J.-L., Marjorie, M., Vial, H., Baziard-Mouysset, G., & Payard, M. (2009). Synthesis and antimalarial activity of new atovaquone derivatives. European Journal of Medicinal Chemistry, 44(11), 4778–4782. https://doi.org/10. 1016/j.ejmech.2009.07.021 google scholar
Essa, E. A., Elmarakby, A. O., Donia, A. M. A., & El Maghraby, G. M. (2017). Controlled precipitation for enhanced dissolution rate of flurbiprofen: development of rapidly disintegrating tablets. Drug Development and Industrial Pharmacy, 43(9), 1430–1439. https://doi.org/10.1080/03639045.2017.1318905google scholar
Fontana, F., Figueiredo, P., Zhang, P., Hirvonen, J. T., Liu, D., & Santos, H. A. (2018). Production of pure drug nanocrystals and nano co-crystals by confinement methods. Advanced Drug Delivery Reviews, 131, 3–21. https://doi.org/10.1016/j. addr.2018.05.002 google scholar
Freitas, C., & Müller, R. H. (1998). Effect of light and temperature on zeta potential and physical stability in solid lipid nanoparticle (SLNTM) dispersions. International Journal of Pharmaceutics, 168(2), 221–229. https://doi.org/10.1016/S0378-5173(98)00092-1google scholar
Gao, L., Zhang, D., & Chen, M. (2008). Drug nanocrystals for the formulation of poorly soluble drugs and its application as a potential drug delivery system. Journal of Nanoparticle Research, 10(5), 845–862. https://doi.org/10.1007/s11051-008-9357-4 google scholar
Ghosh, I., Bose, S., Vippagunta, R., & Harmon, F. (2011). Nanosuspension for improving the bioavailability of a poorly soluble drug and screening of stabilizing agents to inhibit crystal growth. International Journal of Pharmaceutics, 409(1), 260-268. https://doi.org/10.1016/j.ijpharm.2011.02.051 google scholar
Gora, S., Mustafa, G., Sahni, J. K., Ali, J., & Baboota, S. (2016). Nanosizing of valsartan by high pressure homogenization to produce dissolution enhanced nanosuspen-sion: pharmacokinetics and pharmacodyanamic study. Drug Delivery, 23(3), 930–940. https://doi.org/10.1071/7544.2014.923066google scholar
Hao, J., Gao, Y., Zhao, J., Zhang, J., Li, Q., Zhao, Z., & Liu, J. (2015). Preparation and optimization of resveratrol nanosuspensions by antisolvent precipitation using box-behnken design. AAPS PharmSciTech, 16(1), 118–128. https://doi.org/ 10.1208/s12249-014-0211-y google scholar
ICH_Q3C(R9)_Guideline_MinorRevision_2024_2024_Approved.pdf. (n.d.). Retrieved March 14, 2025, from https://database.ich.org/sites/default/files/ICH_Q3C(R9)_Guideline_MinorRevision_2024_2024_Approved.pdfgoogle scholar
Jacob, S., Nair, A. B., & Shah, J. (2020). Emerging role of nanosuspensions in drug delivery systems. Biomaterials Research, 24(1). https://doi.org/10.1186/s40824-020-0184-8 google scholar
Jakubowska E., Bielejewski, M., Milanowski, B., & Lulek, J. (2022). Freeze-drying of drug nanosuspension– study of formulation and processing factors for the opti-mization and characterization of redispersible cilostazol nanocrystals. Journal of Drug Delivery Science and Technology, 74, 103528. https://doi.org/10.1016/j. jddst.2022.103528 google scholar
Kakad, S. P., Gangurde, T. D., Kshirsagar, S. J., & Mundhe, V. G. (2022). Nose to brain delivery of nanosuspensions with first line antiviral agents is alternative treatment option to Neuro-AIDS treatment. Heliyon, 8(7), e09925. https://doi. org/10.1016/j.heliyon.2022.e09925 google scholar
Kakade, P., Gite, S., & Patravale, V.B. (2019). Development of Atovaquone Nanosuspen-sion: Quality by Design Approach. Current drug delivery. 17(2), 112–125. https:// doi.org/10.2174/1567201817666191227095019 google scholar
Kathpalia, H., Das, S., & Shidhaye, S. (2021). Formulation and optimization of atovaquone micronized suspension by top-down method. Indian Journal of Pharmaceutical Education and Research, 55(1), 77–85. https://doi.org/10.5530/ ijper.55.1.11 google scholar
Kathpalia, H., Juvekar, S., Mohanraj, K., Apsingekar, M., & Shidhaye, S. (2020). Inves-tigation of pre-clinical pharmacokinetic parameters of atovaquone nanosus-pension prepared using a pH-based precipitation method and its pharmaco-dynamic properties in a novel artemisinin combination. Journal of Global Antimicrobial Resistance, 22, 248–256. https://doi.org/10.1016/j.jgar.2020.02.018 google scholar
Kathpalia, H., Juvekar, S., & Shidhaye, S. (2019). Design and In Vitro Evaluation of Atovaquone Nanosuspension Prepared by pH Based and Anti-solvent Based Precipitation Method. Colloids and Interface Science Communications, 29, 26-32. https://doi.org/10.1016/j.colcom.2019.01.002 google scholar
Koutsoviti, M., Siamidi, A., Pavlou, P., & Vlachou, M. (2021). Recent advances in the excipients used for modified ocular drug delivery. Materials, 14(15), 4290. https://doi.org/10.3390/ma14154290 google scholar
Kumar, A., Mathur, P., Samantaray, P., & Higgins, W. (2016). (12) United States Patent(45) Date of Patent: (54) (71) (72) (73) (*) (21) (22) (86) (87) (65) (30) Foreign Application Priority Data2013.01) (58) Field of Classification Search Foreign Patent Documents.google scholar
Lappe, S., Mulac, D., & Langer, K. (2017). Polymeric nanoparticles – Influence of the glass transition temperature on drug release. International Journal of Phar-maceutics, 517(1–2), 338–347. https://doi.org/10.1016/j.ijpharm.2016.12.025 google scholar
Li, Z., Gong, Z., Zhang, B., & Nawaz, A. (2024). Investigation of the influence of anti-solvent precipitation parameters on the physical stability of amorphous solids. Molecules, 29(6), 1275. https://doi.org/10.3390/molecules29061275 google scholar
Liu, D., Zhang, H., Cito, S., Fan, J., Mäkilä, E., Salonen, J., Hirvonen, J., Sikanen, T. M., Weitz, D. A., & Santos, H. A. (2017). Core/Shell Nanocomposites Produced by Superfast Sequential Microfluidic Nanoprecipitation. Nano Letters, 17(2), 606–614. https://doi.org/10.1021/acs.nanolett.6b03251 google scholar
Medarević, D., Djuriš, J., Ibrić, S., Mitrić, M., & Kachrimanis, K. (2018). Optimization of formulation and process parameters for the production of carvedilol nanosuspension by wet media milling. International Journal of Pharmaceu-tics, 540(1–2), 150–161. https://doi.org/10.1016/j.ijpharm.2018.02.011 google scholar
Mohtar, N., Khan, N. A. K., & Darwis, Y. (2015). Solid lipid nanoparticles of atovaquone based on 24full-factorial design. Iranian Journal of Pharmaceutical Research, 14(4), 989–1000. https://doi.org/10.22037/ijpr.2015.1766google scholar
Möschwitzer, J., Achleitner, G., Pomper, H., & Müller, R. H. (2004). Development of an intravenously injectable chemically stable aqueous omeprazole formulation using nanosuspension technology. European Journal of Pharmaceutics and Biopharmaceutics, 58(3), 615–619. https://doi.org/10.1016/j.ejpb.2004.03.022 google scholar
Nixon, G. L., Moss, D. M., Shone, A. E., Lalloo, D. G., Fisher, N., O’Neill, P. M., Ward, S. A., & Biagini, G. A. (2013). Antimalarial pharmacology and therapeutics of atovaquone. Journal of Antimicrobial Chemotherapy, 68(5), 977–985. https:// doi.org/10.1093/jac/dks504google scholar
Oktay A. N., Ilbasmis-Tamer, S., Karakucuk, A., & Celebi, N. (2020). Screening of stabi-lizing agents to optimize flurbiprofen nanosuspensions using experimental design. Journal of Drug Delivery Science and Technology, 57, 101690. https:// doi.org/10.1016/j.jddst.2020.101690 google scholar
Ozsoysal, S., & Bilgili, E. (2024). Non-Traditional Natural Stabilizers in Drug Nanosus-pensions. Journal of Pharmaceutical and BioTech Industry, 1(1), Article 1. https://doi.org/10.3390/jpbi1010005 google scholar
Patel, C. M., Chakraborty, M., & Murthy, Z. V. P. (2014). Preparation of fenofibrate nanoparticles by combined stirred media milling and ultrasonication method. Ultrasonics Sonochemistry, 21(3), 1100–1107. https://doi.org/10.1016/j.ultsonch.2013.12.001 google scholar
Patel, D., Zode, S. S., & Bansal, A. K. (2020). Formulation aspects of intra-venous nanosuspensions. International Journal of Pharmaceutics, 586, 119555. https://doi.org/10.1016/j.ijpharm.2020.119555 google scholar
Patravale, V. B., Date, A. A., & Kulkarni, R. M. (2010a). Nanosuspensions: a promising drug delivery strategy. Journal of Pharmacy and Pharmacology, 56(7), 827–840. https://doi.org/10.1211/0022357023691 google scholar
Patravale, V. B., Date, A. A., & Kulkarni, R. M. (2010b). Nanosuspensions: a promising drug delivery strategy. Journal of Pharmacy and Pharmacology, 56(7), 827–840. https://doi.org/10.1211/0022357023691 google scholar
Pınar, S. G., Oktay, A. N., Karaküçük, A. E., & Çelebi, N. (2023). Formulation Strategies of Nanosuspensions for Various Administration Routes. Pharmaceutics, 15(5), Article 5. https://doi.org/10.3390/pharmaceutics15051520 google scholar
Pirincci Tok, Y., Mesut, B., Güngör, S., Sarıkaya, A. O., Aldeniz, E. E., Dude, U., & Özsoy, Y. (2023). Systematic Screening Study for the Selection of Proper Stabilizers to Produce Physically Stable Canagliflozin Nanosuspension by Wet Milling Method. Bioengineering, 10(8), 927. google scholar
Porwal, O. (2022). Box-Behnken Design-based formulation optimization and charac-terization of spray dried rutin loaded nanosuspension: State of the art. South African Journal of Botany, 149, 807–815. https://doi.org/10.1016/j.sajb.2022.04. 028 google scholar
Schöler, N., Krause, K., Kayser, O., Müller, R. H., Borner, K., Hahn, H., & Liesenfeld, O. (2001). Atovaquone Nanosuspensions Show Excellent Therapeutic Effect in a New Murine Model of Reactivated Toxoplasmosis. Antimicrobial Agents and Chemotherapy, 45(6), 1771–1779. https://doi.org/10.1128/AAC.45.6.1771-1779.2001google scholar
Shah, D. A., Murdande, S. B., & Dave, R. H. (2016). A Review: Pharmaceutical and Pharmacokinetic Aspect of Nanocrystalline Suspensions. Journal of Pharma-ceutical Sciences, 105(1), 10–24. https://doi.org/10.1002/jps.24694 google scholar
Shaji, J., & Bhatia, V. (2013). Dissolution enhancement of atovaquone through cyclodextrin complexation and Phospholipid solid dispersion. International Journal of Pharmacy and Pharmaceutical Sciences, 5(3), 642–650. google scholar
Sharma, P., Denny, W. A., & Garg, S. (2009). Effect of wet milling process on the solid state of indomethacin and simvastatin. International Journal of Pharmaceu-tics, 380(1–2), 40–48. https://doi.org/10.1016/j.ijpharm.2009.06.029 google scholar
Srivalli, K. M., & Mishra, B. (2016). Drug nanocrystals: A way toward scale-up. Saudi Pharmaceutical Journal : SPJ : The Official Publication of the Saudi Pharmaceu-tical Society, 24(4), 386–404. https://doi.org/10.1016/j.jsps.2014.04.007 google scholar
Stat-Ease » v23.1 » General Sequence of Analysis » ANOVA Output. (n.d.). Retrieved November 14, 2025, from https://www.statease.com/docs/v23.1/contents/analysis/anova-output/ google scholar
Sutradhar, K. B., Khatun, S., & Luna, I. P. (2013). Increasing Possibilities of Nanosus-pension. Journal of Nanotechnology, 2013, 1–12. https://doi.org/10.1155/2013/ 346581 google scholar
Taneja, S., Shilpi, S., & Khatri, K. (2015). Formulation and optimization of efavirenz nanosuspensions using the precipitation-ultrasonication technique for solubility enhancement. Artificial Cells, Nanomedicine, and Biotechnology, 1–7. https://doi.org/10.3109/21691401.2015.1008505 google scholar
Tuomela, A., Hirvonen, J., & Peltonen, L. (2016). Stabilizing Agents for Drug Nanocrys-tals: Effect on Bioavailability. Pharmaceutics, 8(2), 16. https://doi.org/10.3390/ pharmaceutics8020016 google scholar
Wang, P., Cao, X., Chu, Y., & Wang, P. (2019). Ginkgolides-loaded soybean phospho-lipid-stabilized nanosuspension with improved storage stability and in vivo bioavailability. Colloids and Surfaces B: Biointerfaces, 181, 910–917. https://doi. org/10.1016/j.colsurfb.2019.06.050 google scholar

Details

Primary Language

English

Subjects

Pharmaceutical Sciences

Journal Section

Research Article

Authors

Karishma Patil
0009-0001-0696-544X
India

Rushikesh Ghongane
0009-0001-2958-4869
India

Tejas Nakte This is me
0009-0007-7296-828X
India

Harsha Kathpalia ^*
0000-0003-4175-1065
India

Publication Date

January 14, 2026

Submission Date

October 21, 2024

Acceptance Date

November 17, 2025

Published in Issue

Year 2025 Volume: 55 Number: 3

DOI

https://doi.org/10.26650/IstanbulJPharm.2025.1570740

IZ

https://izlik.org/JA43AS33GF

Cite

RIS / Bibtex

APA

Patil, K., Ghongane, R., Nakte, T., & Kathpalia, H. (2026). Exploring the Bottom-up and Top-down Techniques of Nanosuspension for Enhanced Solubility and Drug Loading of Atovaquone. İstanbul Journal of Pharmacy, 55(3), 390-403. https://doi.org/10.26650/IstanbulJPharm.2025.1570740

AMA

1.Patil K, Ghongane R, Nakte T, Kathpalia H. Exploring the Bottom-up and Top-down Techniques of Nanosuspension for Enhanced Solubility and Drug Loading of Atovaquone. iujp. 2026;55(3):390-403. doi:10.26650/IstanbulJPharm.2025.1570740

Chicago

Patil, Karishma, Rushikesh Ghongane, Tejas Nakte, and Harsha Kathpalia. 2026. “Exploring the Bottom-up and Top-down Techniques of Nanosuspension for Enhanced Solubility and Drug Loading of Atovaquone”. İstanbul Journal of Pharmacy 55 (3): 390-403. https://doi.org/10.26650/IstanbulJPharm.2025.1570740.

EndNote

Patil K, Ghongane R, Nakte T, Kathpalia H (January 1, 2026) Exploring the Bottom-up and Top-down Techniques of Nanosuspension for Enhanced Solubility and Drug Loading of Atovaquone. İstanbul Journal of Pharmacy 55 3 390–403.

IEEE

[1]K. Patil, R. Ghongane, T. Nakte, and H. Kathpalia, “Exploring the Bottom-up and Top-down Techniques of Nanosuspension for Enhanced Solubility and Drug Loading of Atovaquone”, iujp, vol. 55, no. 3, pp. 390–403, Jan. 2026, doi: 10.26650/IstanbulJPharm.2025.1570740.

ISNAD

Patil, Karishma - Ghongane, Rushikesh - Nakte, Tejas - Kathpalia, Harsha. “Exploring the Bottom-up and Top-down Techniques of Nanosuspension for Enhanced Solubility and Drug Loading of Atovaquone”. İstanbul Journal of Pharmacy 55/3 (January 1, 2026): 390-403. https://doi.org/10.26650/IstanbulJPharm.2025.1570740.

JAMA

1.Patil K, Ghongane R, Nakte T, Kathpalia H. Exploring the Bottom-up and Top-down Techniques of Nanosuspension for Enhanced Solubility and Drug Loading of Atovaquone. iujp. 2026;55:390–403.

MLA

Patil, Karishma, et al. “Exploring the Bottom-up and Top-down Techniques of Nanosuspension for Enhanced Solubility and Drug Loading of Atovaquone”. İstanbul Journal of Pharmacy, vol. 55, no. 3, Jan. 2026, pp. 390-03, doi:10.26650/IstanbulJPharm.2025.1570740.

Vancouver

1.Karishma Patil, Rushikesh Ghongane, Tejas Nakte, Harsha Kathpalia. Exploring the Bottom-up and Top-down Techniques of Nanosuspension for Enhanced Solubility and Drug Loading of Atovaquone. iujp. 2026 Jan. 1;55(3):390-403. doi:10.26650/IstanbulJPharm.2025.1570740