United required a victory in the final game of the Premier League campaign and defeat for Tottenham against Aston Villa to steal sixth spot, but results transpired against Ryan Giggs' men.
Tottenham's win over Villa coupled with a share of the spoils at St Mary's Stadium means United miss out on Europe having played in continental competition for the past 23 seasons.
Juan Mata levelled for the visitors early in the second half with a sumptuous free-kick that gave Artur Boruc absolutely no chance in the Southampton goal after Rickie Lambert had broken the deadlock after 28 minutes.
Both sides pushed for a winner and were guilty of missing chances to seal all three points. The draw, combined with Tottenham's win over Villa, sees United finish the campaign in seventh, while Southampton are one position behind them.
The hosts, who named three potential FIFA World Cup hopefuls for the watching England coach Roy Hodgson to monitor from the stands, started the game with more attacking intent, but failed to test David De Gea.
Victor Wanyama was first to threaten the United goal with a low strike in the 10th minute and even Adam Lallana's flick in front of the Spanish goalkeeper was not able to find the back of the net.
Luke Shaw and Lallana both saw good chances wasted before Southampton took the lead just before the half-hour mark through Lambert, although there was controversy surrounding the goal.
An elbow from Lambert in the build-up left Nemanja Vidic on the floor, but play was allowed to continue and the burly forward emphatically fired past De Gea from close range after being played in by Steven Davis' neat pass.
United's players surrounded Mike Dean to bemoan the decision, but the goal stood, while Vidic – making his final appearance before joining Serie A giants Inter - received treatment on a bleeding nose.
The visitors, who left Tuesday's hero James Wilson out of their squad, only created their first sight of goal in the 34th minute as Chris Smalling met a loose ball on the edge of the penalty area, but he skewed his low shot wide without testing Boruc.
Giggs' decision to introduce Michael Carrick at half-time had an immediate impact for the visitors as it released Mata into a more creative role.
And, in the 54th minute, the Spaniard curled a glorious free-kick over the Southampton wall and into the top corner to bring United level.
United's second-half revival continued and just after the hour they should have been ahead through Darren Fletcher.
After neat build-up on the right, Smalling delivered a perfect cross to the back post where the midfielder was able to escape the attentions of Davis, but the Scot placed his free header wide from seven yards.
Danny Welbeck came close to putting United ahead in the closing minutes, but was denied by Boruc, while Southampton struggled to create any chances of note as their season ended with a comfortable display.
With nine minutes remaining, Hodgson left his seat in the stands having seen his potential England players perform well and all come through the game without injury, with Lallana and Shaw both substituted to standing ovations.
