Testosterone deficiency afflicts approximately 30% of men aged from 40-89 years. It is associated with type 2 diabetes, metabolic syndrome and cardiovascular disease. However, the causality of the relationship between low testosterone and metabolic diseases is unclear. We have analysed plasma samples from thirty healthy male volunteers (19-32 years of age) with pharmacologically-induced testosterone deficiency. Blood samples were collected under three conditions: normal testosterone (baseline), ‘zero’ testosterone and ‘restored’ testosterone. As a proof-of-concept, a sample pooling approach was used to obtain one pool per condition, with each pool consisting of 30 individual samples. The proteomic profiles identified significant functional differences, related to the innate immune system, the regulation of insuline-like growth factors, collagen degradation, glycolysis/gluconeogenesis process, among others.