Team Europcar finished 19th in Tuesday's 25km team time trial in Nice after crossing the line one minute and 13 seconds down on winners, Orica-GreenEdge.
Rolland, who wear wearing the polka dot jersey for the second day running, fell to 63rd on GC, 1:13 behind the new race leader Simon Gerrans.
"We knew that time trials were not our strength," said a disappointed Rolland. "We have made a big effort to improve but the other teams have come on strides too and so it's not that easy."
Team Europcar set the fourth best time of the five French teams in the Tour, with only Cofidis finishing below, a further seven seconds in arrears.
Rolland is now 1:10 behind Chris Froome (Sky) and 1:04 down on Alberto Contador (Saxo-Tinkoff) on GC. He also trails Ryder Hesjedal (Garmin-Sharp) by 56 seconds, Movistar pair Alejandro Valverde and Nairo Quintana by 53 seconds, BMC's Cadel Evans and Tejay van Garderen by 47 seconds, and Joaquim Rodriguez (Katusha) by 45 seconds.
With two individual time trials still to ride in the 100th edition of the Tour, Rolland will have to ride with attacking flair in the mountains if he wants to build on last year's eighth place in Paris. But having a deficit could well act in Rolland's favour ahead of the Pyrenees.
"I'm going to try and stay calm and recover as quickly as possible," said Rolland. "The three stages in Corsica were difficult and the team time trial took away a lot of energy. I will take each day as it comes."
The next few stages ahead of Saturday's stage in the Pyrenees look to be suited to the sprinters - although there are four lower category climbs on the menu on Wednesday.
"I will rely heavily on my team-mates to help support me in this kind of situation. Yohann Gene, Kevin Reza and Jerome Cousin will be by my side. And once the terrain goes uphill, I know that I can rely on the support of Davide Malacarne, David Vielleux and Cyril Gautier."
Having been extremely active in the opening road stages in Corsica, Team Europcar will be eager to get over this little blip and put in some strong performances as the race crosses the south of France and heads towards the Pyrenees.