In the present work, a new testing strategy was proposed to characterize the effect of a functional group on the interaction between rubber and filler in a rubber compound. In principle, the filler is mixed into a blend of rubber A and rubber B produced by functionalizing the rubber A with a specific chemical group. The phase-selective wetting of the filler is directly related to the effect of the functional group. Based on the new test approach, the impact of the non-rubber components in natural rubber (NR) and its epoxidation on the rubber-filler interaction in carbon black (CB) filled NR compounds was investigated. Accordingly, CB was mixed in three blends consisting of NR/isoprene (IR), deproteinized natural rubber (DPNR)/IR, and NR/epoxidized natural rubber (ENR). The phase selective wetting of CB was quantitatively characterized based on the wetting concept. It was found that proteins and epoxy groups did not affect the affinity between NR and CB, while the linked phospholipids interacted with the CB surface through cation-π bonding. By fitting the experimentally determined selective wetting of CB into the Z-model, the corrected surface tension of NR describing the effect of phospholipids was determined. The proposed test strategy was applied to other NR blends filled with silica and the hybrid CB/silica filler as reference fillers to demonstrate its transferability and verification.