Serpa Pinto was born at the castle of Poldras in Portugal, on the river Douro. He joined Colégio Militar at age 10. There he became the first student Battalion Commander in 1864, when he joined the Portuguese army and was sent to Mozambique. In 1869 he took part in suppressing tribes in revolt around the lower Zambezi.
In the same year, Pinto went to eastern Africa on an exploration of the Zambezi River. Eight years later he led an expedition from Benguela, Angola, into the basins of the Congo and Zambezi rivers. The town of Menongue was named Serpa Pinto, after him, up to 1975.
In 1877, he and Portuguese naval captains Capelo and Ivens were sent to explore the southern African interior. They left Benguela in November. Soon after their departure, however, they parted company, Capello and Ivens turning northward whilst Serpa Pinto continued eastward. He crossed the Cuando (Kwando) river in June 1878 and in August reached Lealui, the Barotse capital on the Zambezi. There he received assistance from the missionary François Coillard, enabling him to continue his journey along the Zambezi to the Victoria Falls. He then turned south and arrived at Pretoria in northern South Africa on February 12, 1879.
Serpa Pinto was the fourth explorer to cross Africa from west to east and the first to lay down a reasonably accurate route between Bié (in present-day Angola) and Lealui. In 1881 the Royal Geographical Society awarded him their Founder's Medal, "for his journey across Africa ... during which he explored five hundred miles of new country". The account of his travels appeared in English in two volumes entitled How I crossed Africa (London, 1881).
In 1884, Serpa Pinto undertook a less successful exploration of the regions between Mozambique and Lake Nyasa. He was appointed governor of Mozambique in 1889 and organized an expedition to secure the Shire Highlands for Portugal, but was frustrated by the British agents John Buchanan and H. H. Johnston. Shortly afterward he returned to Portugal where he was promoted to the rank of colonel.