Progression of the disability of multiple sclerosis (MS) is thought to be related to axonal damage that is seen even in normal appearing white matter (NAWM) of patients' brains. Interferon beta (IFNβ) treatment for MS reduces the frequency of clinical exacerbations and the appearance of new inflammatory demyelinating lesions on magnetic resonance imaging (MRI). However, the effect of IFNβ treatment on axonal damage in MS is not known. The present study aimed to assess the effect of IFNβ on axonal function by comparing the data from localised single voxel proton MR spectroscopy (MRS) performed on eight MS patients given IFNβ-1b treatment and 11 untreated patients. Brain MRI and MRS were performed before the initiation of IFNβ-1b treatment and 24 months after treatment initiation. Levels of N-acetylaspartate (NAA), choline-containing compounds (Cho) and creatine (Cr) were calculated as the areas under their peaks shown on MRS and the ratios of NAA/Cr and Cho/Cr were calculated. In the patients treated with IFNβ-1b there was a significant increase in the ratio of NAA/Cr in NAWM (p=0.028) at 24 months after the initiation of treatment. In contrast, there was no significant change in the NAA/Cr ratio in the untreated patients. These results suggest that IFNβ-1b treatment might recover axonal function in NAWM of MS patients.