Nanometer size silicon particles for hyperpolarized MRI

Hyperpolarized silicon particles have been shown to exhibit long spin-lattice relaxation times at room temperature, making them interesting as novel MRI probes. Demonstrations of hyperpolarized silicon particle imaging have focused on large micron size particles (average particle size (APS) = 2.2 μm) as they have, to date, demonstrated much larger polarizations than nanoparticles. We show that also much smaller silicon-29 particles (APS = 55 ± 12 nm) can be hyperpolarized with superior properties. A maximum polarization of 12.6% in the solid state is reported with a spin-lattice relaxation time of 42 min at room temperature thereby opening a new window for MRI applications.


Material characterization
The silicon powder was used as obtained from the manufacturer and stored in an air-tight container. No special precautions were taken to prevent air exposure. At the same time, no degradation or change in any of the material properties described in this communication were observed over the period of 10 months.
The average crystalline size and crystal-to-amorphous phase ratio were measured using a Bruker ADVANCE D8 XRD spectrometer. All analysis was made using the manufacturer's dedicated software.
The origin of the paramagnetic properties of the nanoparticles was confirmed with EPR spectroscopy. A CW spectrum was recorded at room temperature using a Bruker ElexSys E580 spectrometer operating at X-band frequency.
Both XRD and EPR spectra confirmed high purity and homogeneity of the silicon particles ( Figure S1). Lack of low-angle dispersion in the XRD spectrum confirmed predominantly crystalline structure of silicon. A single resonance was observed in the EPR spectrum at a gvalue of around ~2.006 which is characteristic for Pb paramagnetic centers 1,2 . Figure S1. A) X-ray diffraction spectroscopy spectrum of powder silicon nanoparticles. The crystal plane assignment was performed according to a well-known silicon standard. B) EPR spectrum recorded in CW dispersion mode.

Particle surface functionalization procedure
All intermediate chemicals were used as purchased. Ethanol (puris) and (3-Aminopropyl)triethoxysilane were sourced from Sigma-Aldrich (Switzerland). An ester of polyethylene glycol in a form of NHS-dPEG4-(m-PEG12)3-ester was obtained from Quanta Biodesign (Ohio, USA). In order to obtain results comparable with previous communications, the same protocol as described in 3 was followed.

Amination
In short, 100 mg of pure silicon nanoparticles were dispersed in 22.5 ml of acidified ethanol (pH~3.5, adjusted with HCl) and sonicated for 10 min. Following that, 0.79 ml of (3-aminopropyl)triethoxysilane (APTES) was added and the solution was stirred for 20 h at 700 rpm on a magnetic stirrer plate. The unreacted APTES was removed but repeated washing with ethanol buffer. Surface amination was confirmed with a fluorescamine test.  nanoparticles. The spectra were shifted in y-direction for better readability.

Size distribution measurement
The total radius of the nanoparticles is expected to increase upon functionalization. The hydrodynamic radius of pure and functionalized material was confirmed by employing dynamic light scattering (DLS). All measurements were done using a Zetasizer Nano Z (Malver) instrument. Less than 1 mg of material was dispersed in 5 ml of distilled water. The measurements were conducted at room temperature with triple repetitions. The size distribution was modeled with a Gaussian distribution ( Figure S3). Although a significant increase in particles radius was observed (by almost a factor of 2), the average diameter of the nanoparticles is still within an acceptable range for theranostics applications 4 (Table S1). Further improvement of the functionalization protocol, especially of the amination time [5][6][7] , in addition to application of high power sonication prior to measurements is expected to minimize effective size of functionalized particles. Furthermore application of zeta potential measurements could help to elucidate detrimental factors affecting particle aggregation. Dextran-coated SPIOs 120-180nm Doxil® Liposomal doxorubicin 80-120nm Table S1. List of FDA approved and clinically used pharmaceutical products based on nanoparticles with their respective size.

Effect of particle surface functionalization
Build-up of 29 Si polarization was compared between pure and functionalized material. No effect on build-up time as well as maximum achievable polarization was found ( Figure S4-A).
In addition, the NMR spectra for both samples were recorded at a 9.4 T imaging system (corresponding to the Larmor frequency ν = 79 MHz). No line broadening due to close proximity of PEG protons was observed ( Figure S4-B), proving that functionalization can be performed without any loss in DNP/NMR properties of silicon nanoparticles. Figure S4. A) Comparison of polarization build-up between pure (black) and functionalized (read) nanoparticles. B) Respective NMR spectra recorded at f=79MHz and at room temperature of pure (black) and functionalized (read) nanoparticles immediately after transfer from the polarizing magnet.

Loss of 29 Si polarization upon dispersion in a solvent
The polarization step was performed on dry silicon powder. Prior to injection, the nanoparticles were dispersed in a solution by manual agitation with a syringe. It has been observed that upon dispersion a substantial loss of 29 Si polarization occurs.
The material was polarized as for previous experiments and transferred to the imaging system. A single free induction decay (FID) signal was acquired with a θ = 20° pulse.
Immediately after the nanoparticles were dispersed in 650 μl of distilled water, a second FID was acquired. The dispersion procedure was carried out at the face of the magnet to avoid any additional relaxation at low magnetic field. As shown in Fig. S5, up to 50% of the initial polarization is lost upon dispersion. At the same time, no change in the line-shape is observed which would be an indication of a rapid motion of silicon nanoparticles in the solution. Such a large loss in polarization could be due to faster relaxation of the nanoparticles caused by a SiNP (-) SINP+APTES+PEG

A) B)
8 temporary boost in rotational motion. As a result, the highly polarized 29 Si nuclei that are closed to surface defects relax instantaneously, leaving only the magnetization from the interior of a particle. Similar results was observed by Cassidy et al. 8 where up to 90% of the initial polarization was lost.