Its inaugural customers were a three-member crew from the Italian Air Force and National Research Council of Italy, alongside a Virgin Galactic astronaut instructor occupying the fourth seat.
Named Galactic 01, the 90-minute mission that took off from Spaceport America, New Mexico, conducted several suborbital science experiments, the company confirmed in a statement.
A livestream was made available on Virgin Galactic's website, kicking off at 9:00 am Mountain Time (1500 GMT).
The launch comes nearly two years after Branson's own flight to the final frontier, a test flight designed to mark the beginning of a new era of profitable space tourism.
However, the company experienced a number of setbacks after this, including a temporary grounding by the Federal Aviation Administration (FAA), due to the Branson flight deviating from its assigned airspace and Virgin Galactic's failure to communicate the "mishap" as required.
Later investigations revealed that certain materials used in its vehicles had not met required strength margins, necessitating fleet upgrades.
Virgin Galactic resumed its spaceflight activities with a successful test in May, setting the stage for Thursday's mission.
Virgin Galactic utilizes a "mothership" aircraft with two pilots that takes off from a runway, gains high altitude, and then releases a rocket-powered plane that zooms into space at nearly Mach 3 before gliding back to Earth.
Passengers in the space plane's cabin experienced a few minutes of weightlessness and caught a glimpse of the planet's curvature from more than 50 miles (80 kilometers) above sea level.
The historic flight included Colonel Walter Villadei and Lieutenant Colonel Angelo Landolfi of the Italian Air Force, Pantaleone Carlucci of the National Research Council of Italy, and Colin Bennett of Virgin Galactic.
There were also two pilots on the spaceplane, and two on the mothership.
The crew conducted 13 supervised and autonomous experiments and collected data on their suits and sensors in the cabin.
Experiments included measuring radiation levels in the under-studied mesosphere, and examining how certain liquids and solids mix in microgravity.
Virgin Galactic has already sold around 800 tickets for future commercial flights -- 600 between 2005 and 2014 for $200,000 to $250,000, and 200 more recently for $450,000 each.
Movie stars and celebrities were among the first to purchase seats, but the company's program suffered a setback in 2014 when a spaceplane on a test flight disintegrated midair, resulting in the death of the copilot and serious injury to the pilot.
It competes in the "suborbital" space tourism sector with billionaire Jeff Bezos' company, Blue Origin, which has already sent 32 people into space.
However, since an accident in September 2022 during an unmanned flight, Blue Origin's rocket has been grounded. The company vowed in March to resume spaceflight soon.
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