The 2-0 scoreline condemned Dean Saunders' crestfallen side to the drop, while Barnsley drew at Huddersfield and rivals Peterborough lost 3-2 at Crystal Palace.
Wolves' second successive relegation means they have an unwanted slice of history as the first English club to suffer the drop from the first to the third tier in successive seasons twice.
And Brighton were again the team to deliver the final blow, just as they did when Wolves last dropped into the old Third Division exactly 28 years ago on May 4, 1985.
Relegation had been on the cards for two months with Saunders unable to stop the slide after the sacking of Stale Solbakken in January following just seven months in charge.
Brighton 2 Wolves 0 - Match report
Wolves slipped into the bottom three on February 19 with the 2-1 defeat at Barnsley and have flirted with danger ever since.
Owner Steve Morgan said relegation was a 'devastating blow for everyone who loves Wolves'.
He acknowledged he had made poor decisions which had contributed to the club's crisis but also blamed the players for letting fans down with sub-standard performances.
And he vowed Wolves would 'fight our way back' next season, restoring 'pride and passion' with a core of younger players. Mr Morgan pledged to bankroll new signings and said a huge clearout of the current squad would take place this summer.
He made no comment on the future of Saunders, whose own comments following today's game suggest he expects to remain in his job.
Full quotes from Steve Morgan after relegation
Dean Saunders said: "It's a massively disappointing day for everyone and I am devastated."
"The game was symptomatic of our season – we missed chances and just couldn't put the ball in the net and Brighton did.
"Sometimes it takes hitting rock bottom before you realise it has got to change, and I have got to change everything and rebuild the club.
"I knew what I was coming into when I walked into the club – I have done this before with clubs who have been struggling and had lots of problems.
"I have no hesitation in wanting to try and take this club forward."
Full quotes from Dean Saunders here
Saunders has blamed the slide on injuries to key players Sylvan Ebanks-Blake, Bakary Sako, Dave Edwards, David Davis and Carl Ikeme.
But the squad should have performed far better over the season and avoided being in the desperate situation the club now finds itself in.
The future of Saunders is now one of many issues to be decided by under-fire owner and chairman Steve Morgan as they prepare for the grim transition to League One football, while huge changes are expected in the squad and possibly behind the scenes with jobs on the line.
Wolves relegation season - in pictures