Abstract Purpose The purpose of the study was to investigate the relationship between gas challenge–blood oxygen level-dependent (GC-BOLD) response angiogenesis and tumor size in rat Novikoff hepatoma model. Materials and Methods Twenty adult male Sprague–Dawley rats (weighting 301–325 g) were used for our Animal Care and Use Committee-approved experiments. N1-S1 Novikoff hepatomas were grown in 14 rats with sizes ranging from 0.42 to 2.81 cm. All experiments were performed at 3.0 T using a custom-built rodent receiver coil. A multiple gradient-echo sequence was used for R2 ⁎ measurements, first during room air (78% N 2/20% O 2) breathing and then after 10 min of carbogen (95% O 2/5% CO 2) breathing. After image acquisition, rats were euthanized, and the tumors were harvested for histological evaluation. Results The R2 ⁎ change between air and carbogen breathing for small hepatomas was positive; R2 ⁎ changes changed to negative values for larger hepatomas. We found a significant positive correlation between tumor R2 ⁎ change and tumor microvessel density (MVD) ( r=0.798, P=.001) and a significant inverse correlation between tumor R2 ⁎ change and tumor size ( r=−0.840, P<.0001). Conclusions GC-BOLD magnetic resonance imaging measurements are well correlated to MVD levels and tumor size in the N1-S1 Novikoff hepatoma model; GC-BOLD measurements may serve as noninvasive biomarkers for evaluating angiogenesis and disease progression and/or therapy response.