Controlled drug release under a low frequency magnetic field: effect of the citrate coating on magnetoliposomes stability† Silvia Nappini, a Massimo Bonini, a Francesca Baldelli Bombelli, a Francesco Pineider, b Claudio Sangregorio, c Piero Baglioni * a and Bengt Nord en * d Received 8th August 2010, Accepted 26th October 2010 DOI: 10.1039/c0sm00789g The paper describes the effect of a low-frequency alternating magnetic field (LF-AMF) on the permeability and release properties of large (LUVs) and giant (GUVs) unilamellar vesicles loaded with citrate coated cobalt ferrite nanoparticles (NPs). The citrate shell allows a high loading of NPs in lipid vesicles without modifying their magnetic properties. The increase of magnetic LUVs permeability upon exposure to LF-AMF has been evaluated as the fluorescence self-quenching of carboxyfluorescein (CF) entrapped inside the liposome aqueous pool. Liposome leakage has been monitored as a function of field frequency, time exposure and concentration of the citrate coated NPs. Confocal Laser Scanning Microscopy (CLSM) experiments performed on magnetic GUVs labeled with the fluorescent probe DiIC 18 and loaded with Alexa 488-C5-maleimide fluorescent dye provided insights on the release mechanism induced by LF-AMF. The results show that LF-AMF strongly affects vesicles permeability, suggesting the formation of pores in the lipid bilayer due to both hyperthermic effects and nanoparticle oscillations in the vesicles pool at the applied frequency. The behaviour of these magnetic vesicles in the presence of LF-AMF makes this system a good candidate for controlled drug delivery. Introduction Carrier-mediated drug delivery has emerged as one of the most promising methods for biomedical applications of liposomes. The therapeutic index of drugs could be enhanced by increasing the specificity due to the targeting of liposomes to a particular tissue or cell, and the control over the release kinetics. The leakage of the drug can be activated through the destabilization of the carrier system by an external stimulus, which can be of different nature, depending on the system (polymer, gel, lipid vesicles). 1–6 Since their discovery, 7 lipid vesicles have attracted growing interest for their potential application as nanometre-scaled drug delivery vectors. 8–11 The interest is mainly related to their biocompatibility, their flexibility in composition and size, 12 and their ability to encapsulate both hydrophilic and hydrophobic compounds into the aqueous pool 13 or in the lipid bilayer, 14 respectively. Magnetic nanoparticles (NPs) can be efficiently encapsulated inside lipid vesicles and used for targeting drugs to a specific location, i.e. diseased cells, using an external magnetic force. 15,16 Moreover, the presence of magnetic NPs in the aqueous pool or in the lipid bilayer of these structures, called ‘‘magneto- liposomes’’ (MLs), allows to enhance the drug leakage by applying an alternating magnetic field (AMF). 16 A number of investigations has been reported on exploiting super- paramagnetic NPs in targeted and controlled release of drugs. 17,18 In most cases, a high-frequency alternating magnetic field (HF-AMF), 10–400 kHz, was used to promote local heat- ing, the so-called hyperthermia, which can be used to cause the thermal ablation of malignant cells without damaging healthy tissues. 2,19–24 Recently, low-frequency alternating magnetic fields (LF-AMFs), 0.1–5 kHz, have been used to study drug release from magnetic systems in order to minimize the temperature contribution and investigate only the field effect. 5,6,25,26 In a previous work, for the first time, we have demonstrated how magnetoliposomes (MLs) can be used to release a fluores- cent hydrophilic molecule (carboxyfluorescein, CF) entrapped in the aqueous pool upon LF-AMF exposure. 27 The enhancement of MLs’ permeability has been measured as the self-quenching decrease of CF’s fluorescence and the leakage has been moni- tored as a function of field frequency, exposure time and magnetic NP concentration, charge and size. The release results have shown that both hyperthermic effect and nanoparticle motions under LF-AMF effectively affect the bilayer structure promoting CF release. Uncoated cobalt ferrite (CoFe 2 O 4 ) nanoparticles, stabilized by electrostatic repulsions in aqueous solution, were used to load the liposome pool. Although the release results were very promising, the loading of NPs and their encapsulation efficiency were quite low. Furthermore, the stability of the system was strongly affected by the presence of NPs, as shown by the appearance of a black precipitate after few days. The precipitation of aggregates induced by CoFe 2 O 4 NPs is a limit to biomedical applications: in fact, large particles are a Department of Chemistry ‘‘U. Schiff’’ and CSGI, via della Lastruccia 3, 50019 Sesto F.no Florence, Italy. E-mail: baglioni@csgi.unifi.it b INSTM and Department of Chemistry ‘‘U. Schiff’’, via della Lastruccia 3, 50019 Sesto F.no Florence, Italy c CNR-ISTM Milano, via C. Golgi 19, I-20133 Milano, Italy d Department of Chemistry and Bioscience, Chalmers University of Technology, SE-412 96 Gothenburg, Sweden. E-mail: norden@chalmers.se † Electronic supplementary information (ESI) available: Cryo-TEM images of magnetoliposomes, AC-measurements on powder samples of citrate-coated and uncoated Cobalt ferrite nanoparticles, picture of the magnet used to apply the LF-AMF, measurements of the magnetic field anisotropy of the magnet at different positions. See DOI: 10.1039/c0sm00789g This journal is ª The Royal Society of Chemistry 2010 Soft Matter PAPER www.rsc.org/softmatter | Soft Matter Downloaded by Massachusetts Institute of Technology on 26 November 2010 Published on 26 November 2010 on http://pubs.rsc.org | doi:10.1039/C0SM00789G View Online