## 4 Discussion  
Both the colloidal stability and the biocompatibility of the nanoparticles are essential for their in vivo biomedical applications. The FA-PAA-coated MnO nanoparticles exhibited these properties owing to the hydrophilic and biocompatible PAA polymers grafted on their surfaces. The observed high \(\mathrm{r_{1}}\) value and the \(\mathrm{r_{2}}\)/\(\mathrm{r_{1}}\) ratio which was somewhat close to one are attributable to the strong interactions between the many Mn\({}^{2+}\) species on the nanoparticle surfaces and the numerous water molecules around the nanoparticles attracted by PAA (Figure 10a), as confirmed by the large hydrodynamic diameter of the nanoparticles. Figure 8: (**a**) \(1/\mathrm{T_{1}}\) and \(1/\mathrm{T_{2}}\) plots versus the Mn-concentration. The \(\mathrm{r_{1}}\) and \(\mathrm{r_{2}}\) values were estimated from the corresponding slopes, respectively. (**b**) Dose-dependent contrast enhancements in \(\mathrm{R_{1}}\) and \(\mathrm{R_{2}}\) map images. Figure 9: (**a**) \(\mathrm{T_{1}}\) magnetic resonance (MR) images before (pre = 0 h) and 4 h after intravenous administration. The dotted circles indicate the location of the brain cancer. Region-of-interest (ROI) at the brain cancer site and normal brain tissue (labeled with small dots and arrows). (**b**) Plots of SNR-ROI versus time, where SNR is the signal-to-noise ratio. This type of interaction corresponds to the magnetic dipole-dipole interaction between 3d-electron spins (S = 5/2) of Mn\({}^{2+}\) and water proton spins (s = 1/2) [8]. As shown in Figure 10b, the longitudinal water proton spin relaxation from the \(\beta\)-spin (\(-1/2\)) to the \(\alpha\)-spin (1/2) is strongly stimulated by the 3d-electron spins of Mn\({}^{2+}\) since slow electron spin motions match well with the proton spin relaxations [7; 8]. Under such conditions, the T\({}_{1}\) relaxation times of the water proton spins significantly decrease, resulting in a high r\({}_{1}\) value, as observed in various hydrophilic polymer-coated gadolinium oxide nanoparticles [36; 37; 38]. As a comparison, the r\({}_{1}\) values of various MnO nanoparticles and metal-ion chelates are provided in Table 2. As presented in Table 2, the r\({}_{1}\) values of large MnO nanoparticles (d\({}_{\text{avg}}\) > 10 nm) [20; 39; 40; 41] are smaller than those [35] of ultrasmall MnO nanoparticles (d\({}_{\text{avg}}\) < 3 nm), including that in this study. This is due to the fact that ultrasmall nanoparticles possess higher surface-to-volume ratios than large nanoparticles, and thus, they possess a higher number of surface Mn\({}^{2+}\) with S = 5/2 per nanoparticle that interact with water proton spins (s = 1/2) (Figure 10a,b). In addition, the r\({}_{1}\) value of the FA-PAA-coated MnO nanoparticles is 2-3 times higher than those [7; 8] of the molecular Gd\({}^{3+}\)- and Mn\({}^{2+}\)-chelates (Table 2). This property corresponds to the advantage of ultrasmall nanoparticle contrast agents over molecular contrast agents, which likely results from the high density of the Mn\({}^{2+}\) species on the nanoparticle surface that can interact with many water molecules (Figure 10a), whereas only a few water molecules are allowed to interact with the central Gd\({}^{3+}\) or Mn\({}^{2+}\) ion in molecular contrast agents [6; 7; 8]. Herein, the FA-PAA-coated MnO nanoparticles exhibited stronger positive contrast enhancements at the brain cancer site compared to those at the normal brain tissue. This is due to the fact that FA possesses a high binding affinity towards folate receptors, which are over-expressed on U87 MG cancer cells [42]. This result demonstrates the application potential of FA-PAA-coated MnO nanoparticles as a cancer-targeting T\({}_{1}\) MRI contrast agent.