The article has studied the mechanism of magnetic nanoparticles (MNPs) affecting magnetic field uniformity. The spatial distribution of MNPs in liquid was simulated based on Monte Carlo method. Combined with the induced field of the single MNP, the magnetic field distribution of magnetofluid was obtained. In the simulation, magnetic field uniformity was described by a statistical distribution. As the chemical shift (CS) and full width at half maximum (FWHM) of magnetic resonance (MR) spectrum can reflect the uniformity of magnetic field, the simulation was verified by spectrum experiment. Simulation and experimental results proved that the CS and FWHM of the MR spectrum are basically positively correlated with the concentration of MNPs and negatively correlated with the temperature. The research results can explain how MNPs play a role in MR by affecting the uniform magnetic field, which is of great significance for improving the temperature measurement accuracy of magnetic nanothermometers and the spatial resolution of magnetic particle imaging.