(T1330-04-21) Lipid Core Nanocapsules for Drug Delivery of Antimicrobials

Purpose: Over the past years, the development of biodegradable nanoparticulate systems for drug delivery has aroused great interest. Substantial advances happened in the development of functional nanomaterials towards biomedical materials, mainly for targeted therapies and drug delivery. Lipid core nanocapsules (LCN), a class of nanocapsules in which a polymeric shell is arranged around an inner core and is formed by a dispersion of a liquid lipid and a solid lipid, have gained attention since they can vehicle actives soluble in their lipidic core or polymeric shell [1]. PCL (poly-E-caprolactone), a biocompatible and biodegradable polymer, has been used to encapsulate a wide range of drugs, achieving targeted and controlled drug delivery [2, 3]. Fluconazole, a bis-triazole antifungal [4], and rifamycin SV, an ansamycin antibiotic [5], could benefit from encapsulation. Our objective was to encapsulate fluconazole and rifamycin SV in LCN, aiming to improve existing treatments by controlling their release.

Methods: Empty-LCN (unloaded), Flu-LCN (LCN loaded with fluconazole) and Rif-LCN (LCN loaded with lidocaine) were prepared as previously described by Oliveira, et al. [6]. Mean diameter and polydispersity index were determined by dynamic light scattering and zeta potential by electrophoretic mobility, using ZetaSizer NanoZS instrument. Total drug concentration was determined by HPLC/UV, as well as encapsulation efficiency after separating non-encapsulated drug by ultrafiltration-centrifugation, and drug distribution in nanocapsule [6]. In vitro antimicrobial activity of free drugs and LCN formulations was determinate using minimal inhibitory concentration (MIC) method against S. aureus, P. aeruginosa and C. albicans microorganisms [7].

Results: Empty-LCN, Flu-LCN and Rif-LCN characterization by dynamic light scattering is presented in Figure 1. All nanocapsules showed negative zeta potential, which is due to the chemical structure of poly(ε-caprolactone) which has a negative charge density in the carboxyl functions in the presence of water [8]. For all formulations, significant differences (p < 0.05) were observed regarding the average diameter when comparing the types of liquid oil used, and in all cases the particles with baru oil had a smaller diameter, but no significant differences were observed. in the polydispersity index. Fluconazole presented a type II distribution in LCN, with encapsulation efficiency calculated as 8.0%, drug content of 798 µg/mL and drug loading of 0.17%. On the other hand, rifamycin SV presented a type III distribution, with encapsulation efficiency of 66.2%, drug content of 743 µg/mL and drug loading of 1.28%. Drugs release profile are shown in Figure 2. Around 53% of fluconazole was released in the first 7h, reaching 77% in 48h; while less than 5% of rifamycin SV was released in the first 8h, reaching 21% in 48h. Empty-LCN did not show any antimicrobial activity. Free-Flu and Flu-LCN did not showed activity in the tested concentrations for S. aureus and P. aeruginosa, and a MIC of 0.39 µg/mL for C. albicans. Free-Rif and Rif-LCN showed MIC of 3.1x10^-3 µg/mL for S. aureus, 100 ug/mL for P. aeruginosa, and no activity against C. albicans.

Conclusion: LCN formulations with fluconazole and rifamycin SV were successfully produced. Depending on drug characteristics, higher encapsulation efficiency and drug loading was achieved, with a more prolonged release. Antimicrobial activity of fluconazole and rifamycin SV was maintained after encapsulation in LCN formulations, and no activity was reported for empty-LCN.

References: [1] C.G. Venturini, E. Jäger, C.P. Oliveira, A. Bernardi, A.M.O. Battastini, S.S. Guterres, A.R. Pohlmann, Formulation of lipid core nanocapsules, Colloids and Surfaces A: Physicochemical and Engineering Aspects 375(1) (2011) 200-208.
[2] V.R. Sinha, K. Bansal, R. Kaushik, R. Kumria, A. Trehan, Poly-ϵ-caprolactone microspheres and nanospheres: an overview, International Journal of Pharmaceutics 278(1) (2004) 1-23.
[3] A.N. Zelikin, C. Ehrhardt, A.M. Healy, Materials and methods for delivery of biological drugs, Nature Chemistry 8(11) (2016) 997-1007.
[4] F.-X. Mathy, D. Ntivunwa, R.K. Verbeeck, V. Préat, Fluconazole Distribution in Rat Dermis Following Intravenous and Topical Application: A Microdialysis Study, Journal of Pharmaceutical Sciences 94(4) (2005) 770-780.
[5] Floss, H. G.; Yu, T. W. Rifamycin-mode of action, resistance, and biosynthesis. Chem. Rev. 2005, 105, 621– 632, DOI: 10.1021/cr030112j
[6] C.P. Oliveira, C.G. Venturini, B. Donida, F.S. Poletto, S.S. Guterres, A.R. Pohlmann, An algorithm to determine the mechanism of drug distribution in lipid-core nanocapsule formulations, Soft Matter 9(4) (2013) 1141-1150.
[7] Methods for dilution antimicrobial susceptibility tests for bacteria that grow aerobically; approved standard-sixth edition. Wayne, PA: CLSI - Clinical and Laboratory Standards Institute 2003.
[8] Woodruff, M. A.; Hutmacher, D. W. The return of a forgotten polymer—polycaprolactone in the 21st century. Progress in Polymer Science, 35, n. 10, p. 1217-1256, 2010.

Acknowledgments: Authors do not have any conflict of interest to disclaimer. Authors acknowledge the financial support from FAPESP (process number 2018/07707-6, 2018/06475-4 and 2021/12452-0).

Figure 1. Lipid core nanocapsules characterization by DLS according to average size (A), polydispersity index (B) and zeta potential (C). Results are presented as average ± SD.



Figure 2. Drug release profile from Flu-LCN and Rif-LCN in 48 hours. Results are presented as average ± SD (n=6).