As part of his undergraduate course at Yale, Chittenden, who was born in New Haven, Connecticut, was asked to investigate why scallops taste sweeter when reheated from a previous meal than when freshly cooked. This project led to his discovery of glycogen and glycine in the muscle tissue – the first demonstration of the free occurrence of glycine (or glycocoll as it was then known) in nature. The work attracted the attention of Willy Kühne at Heidelberg who invited Chittenden to his laboratory. Later collaboration between Chittenden (at New Haven) and Kühne (in Heidelberg) provided a strong foundation for studies in enzymology.

Chittenden also did important work in toxicology and on the protein requirements of man, showing that the so-called Voit standard, which recommended 118 grams of protein per day, was a vast overestimate, and that good health could be maintained on 50 grams a day. He played a major part in the establishment of physiological chemistry (biochemistry) as a science in its own right.