Triggered Release from Thermoresponsive Polymersomes with Superparamagnetic Membranes

Magnetic polymersomes were prepared by self-assembly of the amphiphilic block copolymer poly(isoprene-b-N-isopropylacrylamide) with monodisperse hydrophobic superparamagnetic iron oxide nanoparticles (SPION). The specifically designed thermoresponsive block copolymer allowed for efficient incorporation of the hydrophobic nanoparticles in the membrane core and encapsulation of the water soluble dye calcein in the lumen of the vesicles. Magnetic heating of the embedded SPIONs led to increased bilayer permeability through dehydration of the thermoresponsive PNIPAM block. The entrapped calcein could therefore be released in controlled doses solely through exposure to pulses of an alternating magnetic field. This hybrid SPION-polymersome system demonstrates a possible direction for release applications that merges rational polymersome design with addressed external magnetic field-triggered release through embedded nanomaterials.


Reagents
All chemicals were purchased from Sigma-Aldrich (Austria) and used without further purification except otherwise noted.

Spectroscopic Analysis
Figures S1-S3 show additional characterization of the synthesized block copolymer BCP 2, demonstrating purity of the product.

Characterization of Residual THF
THF remaining at high concentration in the sample could affect stability over time and lead to toxicity in biological applications. However, THF is a high vapor pressure solvent and is readily evaporated under continuous nitrogen flow until a homogeneous suspension of vesicles containing SPIONs is achieved. The amount of residual THF for the preparation of liposomes through the same protocol for solvent inversion was previously quantified by us by NMR to be 0.05 ‰ or 50 ppm of its initial value ( Figure S4) [1]. Also after 3 h under continuous nitrogen evaporation it is negligible. Such minimal traces of THF retained are far below any toxic level and suitable for biological and medical applications [2].  Figure S5 shows additional TEM micrographs of calcein-loaded, extruded PI-b-PNIPAM polymersomes with 20% w/w 3.5 nm hydrophobic SPION input. Figure S5. TEM micrographs of calcein-loaded, extruded PI-b-PNIPAM polymersomes at 1 mg/mL with 20% w/w 3.5 nm hydrophobic SPION input. Samples were prepared by THF solvent inversion into 5 mg/mL calcein solution to form polydisperse, large polymersomes and subsequent extrusion through 100nm track-etched polycarbonate membranes after evaporation of the organic solvent.

Thermogravimetric Analysis
For TGA determination of the effective SPION content of the polymersome membranes the lyophilized samples were burnt under oxidative conditions (synthetic air) to yield near complete combustion. Yet a considerable residue (~11% w/w) remained even in the case of polymersomes containing no SPIONs. The reported final SPION loading content therefore refers to the non-combusted material at 650 °C in excess of the residue for samples not containing nanoparticles, which amounts to approximately 9% w/w for extruded SPION loaded samples.

Optical Density Determination
Optical density (OD) values at 350 nm (OD 350 ) were used for spectroscopic quantification of the SPION embedding efficiency. The OD 350 values were obtained by dilution of the respective suspensions to match the amide absorptions at 208 nm. Background spectra of the plain extruded PI-b-PNIPAM vesicles were recorded to account for vesicular scattering. The OD 350 value of the initial SPION loaded suspension was assigned to the input SPION weight fraction (20%) and the final loading content was determined by evaluating the OD 350 decrease upon extrusion. In this way we estimate an effective loading content of around 9% w/w which agrees with the results obtained by TGA. The slight difference between the two methods is likely to be caused by remote mass loading and use of augmenting relations in TGA while the choice of evaluation wavelength and variations in lamellarity cause deviations in UV/VIS. Figure S6A shows the increase in bulk temperature resulting from magnetic heating and Figure S6B shows the long-term passive release of calcein at room temperature when no actuation is applied. Figures S7 and S8 show the effect of temperature on polymersome size from bulk heating and from localized heating by application of an alternating magnetic field.