The microstructure of acrylonitrile–butadiene rubber (NBR) was shown to be dependent on the polymerization conditions. The NBR investigated in the current study was obtained via radical copolymerization under azeotropic conditions (AN/BD = 38/62) in organic solution in the presence of either a conventional chain transfer agent or a reversible addition fragmentation chain transfer (RAFT) agent. The variation in the polymer microstructure was proven to originate from different radical environments during the polymerizations with initial radical initiator concentrations in the polymerizations studied ranging from 1.0 mM to 34.1 mM. The variation of the polymer microstructure was evidenced by triple SEC measurements, making use of the simultaneous determination of molecular weights with two independent methods, namely on-line viscometry and on-line light scattering. It is additionally evidenced that the microstructure shows a gradual variation during the course of a polymerization, a behaviour observed when the polymerizations were performed in the presence of an elevated initial radical initiator concentration. Furthermore, experimental evidence for the variation of the NBR microstructure during RAFT polymerizations is provided. At low conversions, a rather uniform polymer is obtained. With increasing conversion, a loss of the controlled character is observed and the microstructures approach those of nitrile rubber obtained by conventionally controlled free radical copolymerizations employing mercaptane transfer agents. Despite the differences in the polymer microstructure, it is possible to report a single set of MHKS parameters for the prepared NBR with azeotropic composition. A linear regression of the Mark–Houwink plots of the samples polymerized under different conditions gives values of K = (49.5 ± 5.5) × 10−5 dL g−1 and α = 0.689 ± 0.010 with a low error margin for SEC separation in THF at 25 °C. Weight average molecular weights of the investigated NBR samples were in the range of 40000 to 155000 g mol−1. The molecular weights of the copolymers determined via universal calibration with the MHKS parameters presented in the current study show a good agreement with molecular weights obtained from light scattering, underpinning the veracity of the obtained parameters.