The Frenchman picked up a hamstring injury in Saturday's 1-1 draw at Sporting Gijon that will keep him out for the visit of Dinamo Zagreb.
Nasri has starred since joining on-loan from Manchester City as Sampaoli's excellent start in Seville has seen the Spanish side pick up seven points from their first three Group H games without even conceding a goal.
However, the Frenchman picked up a hamstring injury in Saturday's 1-1 draw at Sporting Gijon that will keep him out for the visit of the Croatian champions and makes him a doubt for Barcelona's trip to the Sanchez Pizjuan on Sunday.
"It is a difficult situation because Nasri's performances recently have been excellent," said Sampaoli on Tuesday.
Sevilla won three consecutive Europa League titles under former boss Unai Emery, but haven't reached the last 16 of the Champions League since the 2009/10 season.
Victory over Zagreb will guarantee the Andalusians' place in the knockout stages as long as Lyon don't win at Juventus in the other game in the group.
"Hopefully, tomorrow in his (Nasri's) absence the team takes flight to achieve something exciting in the best competition in the world," added Sampaoli.
"We need to learn to play without him. It is difficult, but it is our obligation."
Sampaoli admitted there will also be other changes from the side that started against Sporting.
However, the Argentine rejected suggestions he is resting players for the visit of Barca.
"I try to put out the best team for every game," he said.
"Right now I am thinking about tomorrow and after Zagreb we will think about Barcelona. Honestly, I am not thinking about the game on Sunday."
Dinamo have yet to take a point in the group and have lost all 10 of their last away games in the Champions League group stage.
However, they have improved both domestically and in their one European outing under new coach Ivaylo Petev.
A 1-0 defeat to Sevilla two weeks ago is Dinamo's only loss in six games under Petev and he expects more attacking threat from his side in Spain.
"We have to be brave," he told UEFA.com.
"I don't think I will change much from the first match in Zagreb. We didn't look dangerous in the first half in Zagreb, but after the break we had some chances to score.
"We had a team full of youngsters, so I can't say anything against them but I am sure we will be more dangerous in Seville."