The hemipteran insect Rhodnius prolixus ingests large amount of vertebrate blood in a single meal and the hydrolysis of hemoglobin inside its digestive tract releases huge amounts of free heme, a pro-oxidant molecule that can iniciate an oxidative stress situation through an increase in the production of reactive oxygen species (ROS). Hematophagous insects deal with this situation by means of several antioxidant defenses. Besides the toxic effects of ROS it has been shown that these molecules have important physiological roles in a wide range of processes like cellular proliferation, inflammation and microbial killing. Little is known about the role of ROS in insect physiology, although a few recent papers have demonstrated that theses molecules might be involved in immunity towards pathogens. Here we studied the production of ROS in Rhodnius prolixus gut. Adult Rhodnius females were fed with rabbit blood and 3 days or 21 days after the blood meal (ABM) both anterior and posterior midguts were dissected and their contents were washed out with cold saline. Tissues were homogenized in PBS and centrifuged for 1 min at room temperature. Midgut epithelium hydrogen peroxide (H₂O₂) content was measured using scopoletin-horseradish peroxidase method. Three days ABM insects showed approximately 17,5 nmol of H₂O₂ in the posterior midgut, compared to 4 nmol in the anterior midgut. The insects dissected 21-days ABM showed 2,6 nmol H₂O₂ in the posterior midgut and 2,7 nmol in the anterior midgut. These findings suggest that the blood digestion inside the gut increase ROS production. The large pool of free heme that is created upon hemoglobin hydrolysis, potentially could interact with midgut-derived ROS, thus representing an extra source of oxidative stress.