In aquariums and zoos across the world marine mammals are among the most popular exhibits. Caretakers strive to provide the least stressful and natural environments for the animals. Analysis of (steroid) hormones in excrement has proven to be an effective, non-invasive method. It allows animal keepers to monitor physiological health associated with reproduction and the stress response in captive animals. Prolactin (PRL) is a non-steroidal, non-tropic peptide hormone most commonly associated with milk production in nursing females. However, it has a multitude of normal physiological functions such as: regulation of the immune system, osmotic balance, angiogenesis, an indicator of parental care (in both males and females), sexual maturity in males, and near-term pregnancy in females. Chronically high or low levels of PRL have been associated with a number of ailments. For instance, impotence reduces the secretion of estrogens and testosterone. To date, no studies have been documented on prolactin PRL levels in captive northern fur seals, (Callorhinus ursinus), harbor seals (Phoca vitulina), or sea otters (Enhydra lutris). Using an enzyme-immunoassay (EIA) kit, previously tested only in human and elephant serum analysis, we investigated PRL levels of several samples collected from captive marine mammals (Northern fur seal blood serum, harbor seal saliva, and sea otter urine/fecal samples). Due to the helical structure of PRL, fecal samples were not expected to survive intact after transit through the digestive tract; therefore, were not expected to produce positive results. Here we report preliminary results of the EIA PRL assay on several different types of samples, as well as several longitudinal samples from the same individual, to show significant changes in this hormone over time. If detectable in urine, fecal matter, and/or saliva, this assay could prove to be a useful non-invasive method for determining health in captive marine mammals.