OXFORD United suffered their second away defeat in five days as a strong Burton Albion side took the points at the Pirelli Stadium.
Adam McGurk put the promotion-chasing Brewers in front with the game's first serious attack, on 14 minutes.
It was a rare opening in a tight first half, but United, who lost at Accrington on Tuesday night, scrapped to stay in the game and improved with the introduction of Roofe on the hour mark.
He created one good chance which was shot wide by Danny Hylton, before winning a free-kick five minutes from time.
Josh Ashby came within inches of an equaliser with the resulting set piece, which hit the crossbar.
Having survived the scare, Burton made the game safe late on through Lucas Akins, who neatly finished to make it 2-0.
It left United, who have won just twice in their last dozen outings, just four points above the bottom two.
Roofe was named on the bench, but fellow new signing George Baldock came straight in for his debut.
The right back was one of three changes from the side which started the 1-0 defeat at Accrington Stanley in midweek.
Andy Whing came in to shield the back four in a 4-1-4-1 system, where Ashby was handed his second senior start in midfield.
Burton came into the game with a formidable recent record, but they spent much of the opening exchanges chasing United.
The visitors made an assured start - and then conceded the first time Burton came forward with purpose.
Damien McCrory got goal side of Hylton and fired a fierce shot from the edge of the box, which Ryan Clarke parried well, but it looped up perfectly for McGurk to nod in from close range.
Those two efforts were the only on target at either end in a first half low on incident.
Burton were the side asking most of the questions, but for the most part the visitors kept them at arm's length.
Making any headway in possession was another matter, though.
The Brewers had not conceded more than one goal in a game since October and they proved a difficult unit to break down, particularly with a lead to protect.
They were really stretched just once in the first half, when a long free-kick fell to Johnny Mullins on 43 minutes.
The defender took it down well, but a tackle came in just before he was able to pull the trigger.
It took until the stroke of half-time for the visitors to finally muster an effort, although Callum O'Dowda's shot was sliced so wide it went for a throw-in.
The shot count doubled early in the second half, when Ashby prodded wide after a burst out wide from O'Dowda.
But it was not the start of a brighter spell from the visitors, who were quickly on the back foot.
Many Burton fans were cheering a second goal on 49 minutes, when Shane Cansdell-Sherriff met a corner, only for his shot to be cleared off the line.
McGurk quickly had a chance to double his tally when he outmuscled a dithering Jake Wright, before shooting over wastefully.
Akins also headed onto the roof of the net at a corner in a good spell for Burton at the start of the half.
United needed to change the game and on the hour Roofe came on for his debut, replacing Whing.
It saw them switch to 4-4-2, opening the game up.
The West Brom loanee took 15 minutes to make his mark, before creating their first clear chance of the match.
Roofe danced past a couple of defenders by the corner flag and crossed low for Hylton, who could only find the side netting.
He worried Burton again five minutes later with a run which was only halted by a John Mousinho foul a yard outside the box.
Ashby hit it sweetly, but could not quite get enough dip on the free-kick, which rattled the crossbar.
Burton were ruffled for the first time all afternoon as United sensed an equaliser.
A goal was just around the corner, but it came at the other end.
With the U's pushing on, Akins got on to a through ball and slotted past an exposed Clarke to end the contest.
Burton Alb: McLaughlin, Edwards, Mousinho, Cansdell-Sherriff, McCrory, Lenihan, Weir, Akins, McGurk (Naylor 71), Calero (El Khayati 77), Beavon (Taft 90).
Unused subs: Phillips, Shearer, Harness, Slade.
Oxford Utd: Clarke, Baldock, Mullins, Wright, Skarz, Whing (Roofe 60), Hylton (Roberts 86), Collins, Ashby, O'Dowda, Hoban.
Unused subs: Ashdown, Rose, Dunkley, Ruffels, Long.
Referee: Peter Bankes (Merseyside).
Attendance: 2,954.
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