Starting four decades ago, studies have examined the ecology and evolutionary dynamics of populations and species using short mitochondrial DNA fragments and stable isotopes. Through technological and analytical advances, the methods and biomolecules at our disposal have increased significantly to now include lipids, whole genomes, proteomes, and even epigenomes. At an unprecedented resolution, the study of ancient biomolecules has made it possible for us to disentangle the complex processes that shaped the ancient faunal diversity across millennia, with the potential to aid in implicating probable causes of species extinction and how humans impacted the genetics and ecology of wild and domestic species. However, even now, few studies explore interdisciplinary biomolecular approaches to reveal ancient faunal diversity dynamics in relation to environmental and anthropogenic impact. This review will approach how biomolecules have been implemented in a broad variety of topics and species, from the extinct Pleistocene megafauna to ancient wild and domestic stocks, as well as how their future use has the potential to offer an enhanced understanding of drivers of past faunal diversity on Earth.