Using a keratinase to degrade chicken feathers for improved extraction of glucocorticoids

Stress in animals is a concern in conservation breeding programs and livestock production facilities. The biological stress response is mediated by the release of glucocorticoids, which can suppress reproduction, growth, and immunity if recurrently activated. Feathers can be used to extract and monitor concentrations of corticosterone, a primary glucocorticoid in birds. However, current techniques for extracting feather corticosterone present challenges, including difficulty assessing extraction efficiency or hormone recovery, inconsistent extraction across feather lengths or pieces, and several uncertainties regarding the mechanisms of hormone deposition into feathers. To overcome such challenges and to provide tools useful for facilitating conservation breeding and livestock production, we developed and validated an alternative procedure for extracting feather glucocorticoids. We first developed a protocol to enzymatically digest the protein matrix of feathers using a keratinase, such that non-protein analytes could be isolated by organic extraction. We then developed an extraction protocol and evaluated techniques by measuring extraction efficiency and by testing parallelism and hormone recovery (accuracy) using radioimmunoassay. Our results demonstrated high and consistent extraction efficiency, as well as high accuracy and reliable parallelism to a standard curve upon measurement of corticosterone concentrations from extracts. By dissolving feather material into solution prior to extraction, we were able to replicate hormone deposition into the feather matrix and ensure consistent extraction across feathers. This work provides additional support for the validity and practicality of extracting glucocorticoids from feathers. Our extraction protocol is likely to extend to other applications as well, including the isolation of numerous non-protein analytes from various keratinized tissues.