Neutering is a common veterinary recommendation and is often associated with obesity development. Thus, the aim of the present study was to evaluate the effects of two different amounts of protein intake by neutered dogs regarding maintenance energy requirement (MER), body composition, and biochemical and hormonal parameters. A total of fourteen healthy adult dogs were fed either a diet containing 59·7 g protein/1000 kcal (4184 kJ) (P60) or a diet with 94·0 g protein/1000 kcal (4184 kJ) (P94) for 26 weeks after neutering to maintain their body weight prior to neutering. A mixed model was fitted to verify diet, time and diet × time interaction effects on biochemical parameters, serum concentrations of insulin, glucagon, leptin and insulin-like growth factor-1 (IGF-1). MER and the body composition data were evaluated within diets (paired t test) and within times (unpaired t test). A time effect was found for fructosamine, TAG, total lipids and IGF-1 serum concentrations. The diet × time interaction was significant for glucagon (P < 0·05). No differences between diets in the MER within each time were found. However, there was a reduction in the MER of dogs fed the P60 diet 26 weeks after neutering (P = 0·042). The fat body mass of dogs fed the P60 diet increased (P < 0·05) after neutering, even without a body-weight change. Some of the biochemical parameters changed over time, but all remained within the normal range. For the period evaluated in the present study, a diet with 94·0 g of protein/1000 kcal (4184 kJ) metabolisable energy seems to be a beneficial nutritional strategy to maintain the MER and the body composition of dogs after neutering.