In this paper, the effect of microstructure of a thick-walled rotor shaft for wind turbines on fracture toughness properties has been investigated. The relevant nodular cast iron grade EN-GJS-600-3 was processed using chill casting technology. Due to different solidification conditions over the wall thickness, heterogeneous microstructures were formed. To illustrate the influence of the microstructure gradient caused by chill casting technology, specimens were taken from different sample positions in the cross section of the casting component. A detailed metallographic analysis revealed essential differences in microstructure. The crack growth resistance under quasi-static loading conditions and the fatigue crack propagation under cyclic loading conditions were measured. The results of the static fracture mechanics investigations revealed that fracture toughness is strongly influenced by the microstructure of this pearlitic ductile iron grade. On the other hand, cyclic fracture mechanics analysis showed that the complex formation of the microstructure has only a minor effect on the fatigue threshold value, but microstructure has a significant effect on the stable crack growth. For the assumed load cases, it was shown that microstructure can be a dominant factor on the mechanical and fracture toughness properties.