Born near Hempstead, New York on Long Island, Truxtun had little formal education before joining the crew of the British merchant ship Pitt at the age of twelve. By the time he was twenty, however, his talents had garnered him the command of his own vessel, the Andrew Caldwell. He operated as a privateer during the American Revolution, commanding several ships: Congress, Independence, Mars and St. James. Truxtun was highly successful in capturing enemy ships during this period, not once suffering a defeat.
After the war he returned to the merchant marine, where he remained for twelve years, and in 1786 commanded one of the first American ships to engage in trade with China, the Canton operating from Philadelphia.
Truxtun was appointed a captain in the United States Navy in 1794, and during the Quasi War was in command of the USS Constellation. Previously, he had overseen its construction with Silas Talbot, and after a rank dispute was placed in charge of the ship by President Washington. He was promoted to commodore and met with considerable success. His victories, perhaps most notably that over the French vessel L'Insurgente, made Truxtun something of a media hero at the time.
During this period, however, Truxtun was involved in a dispute over rank with Richard Dale. Truxtun took command of the USS President for a few months in 1800, soon after retiring from the navy and locating first in Perth Amboy, New Jersey and later in Philadelphia. He was offered command during the First Barbary War in 1801 but refused, settling firmly into retirement.
He ran an unsuccessful campaign for the United States House of Representatives in 1810, and in 1816 was elected sheriff of Philadelphia County, serving a four-year term. He also published several books, well-known at the time, covering navigation and naval tactics. He is buried at Christ Church Burial Ground.
Several Navy ships have since been named USS Truxtun in his honor.