Folkers was born in Decatur, Illinois, graduated in chemistry from the University of Illinois in 1928, and gained his PhD from the University of Wisconsin in 1931. After postdoctoral work at Yale he joined the pharmaceutical manufacturers Merck and Company in 1934, becoming director of organic and biochemical research in 1945. He was president of the Stanford Research Institute from 1963 to 1968, and then director of the Institute for Biomedical Research at the University of Texas.

In 1948 Folkers' team isolated the antipernicious anemia factor, vitamin B_{1 2} (cyanocobalamin) and they played a major role in the lengthy process of determining the structure of this molecule. Folkers was involved in many investigations of biologically active compounds, especially antibiotics, and the structure of streptomycin was largely determined by his group in 1948.