Trade-offs between selected and nonselected environments are often assumed to exist during adaptation. This phenomenon is prevalent in microbial metabolism, where many organisms have come to specialize on a narrow breadth of substrates. One well-studied example is methylotrophic bacteria that can use single-carbon (C(1)) compounds as their sole source of carbon and energy, but generally use few, if any, multi-C compounds. Here, we use adaptation of experimental populations of the model methylotroph, Methylobacterium extorquens AM1, to C(1) (methanol) or multi-C (succinate) compounds to investigate specialization and trade-offs between these two metabolic lifestyles. We found a general trend toward trade-offs during adaptation to succinate, but this was neither universal nor showed a quantitative relationship with the extent of adaptation. After 1500 generations, succinate-evolved strains had a remarkably bimodal distribution of fitness values on methanol: either an improvement comparable to the strains adapted on methanol or the complete loss of the ability to grow on C(1) compounds. In contrast, adaptation to methanol resulted in no such trade-offs. Based on the substantial, asymmetric loss of C(1) growth during growth on succinate, we suggest that the long-term maintenance of C(1) metabolism across the genus Methylobacterium requires relatively frequent use of C(1) compounds to prevent rapid loss.