The re-enactment took place 70 years to the minute after the first test flight took off from Southampton Airport.
Five spitfires flew in formation along Southampton Water, providing a spectacle which may never be repeated.
Among those who took part in the event was Alex Henshaw, a former chief test pilot of the Spitfire who is now 93.
Alex was a passenger in one of the few two-seater versions of the aircraft during the memorial flight.
To mark the anniversary, a special exhibition celebrating the fighter aircraft which made its first flight from Eastleigh Airport in 1936 is being staged at Eastleigh Museum.
The exhibition runs from 19th April - 27th May, Tuesdays to Fridays, 10am - 5pm and Saturday 10am - 4pm, admission is free.