Malcom will be out of contention for 10 to 15 days after Barcelona confirmed he suffered an ankle problem in Wednesday's Copa del Rey meeting with Cultural Leonesa.
The Brazilian sustained a sprained right ankle in the 4-1 victory at Camp Nou, which secured a 5-1 aggregate success and a place in the last 16.
Barca had used all three substitutes by that stage and were forced to see out the remaining minutes a man light.
Malcom had netted his side's third goal late in the first half, but is now certain to miss Saturday's derby trip to Espanyol as well as Tuesday's Champions League encounter at home to Tottenham. Barca are already assured progress as winners of Group B.
Malcom left the field in tears after being forced off in midweek.
The Brazilian has struggled to establish himself in Ernesto Valverde’s plans since moving to Spain from Bordeaux in a reported €40million (£36.5m/$48m) deal, having made just nine appearances in all competitions.
Speaking of his decision to withdraw the wide man, Valverde said at the time: "I think it is not a hard injury. I don't know what exactly he has so I prefer to be prudent”.
Malcom’s absence adds to Barcelona’s injury woes, with the La Liga leaders already missing Sergi Roberto, Luis Suarez, Rafinha, Samuel Umtiti, Arthur, and Arturo Vidal.
While Roberto and Suarez are expected to be back in action before the New Year, Rafinha does not look likely to return before the end of the season. It is unclear as to when Umtiti, Arthur, and Vidal will return.
Barcelona’s next match will be a derby day clash with Espanyol on Saturday evening, with the defending champions looking to open up a gap over second-placed Sevilla, who currently lie just one point behind them.
They are then due to face Tottenham in their final outing of the Champions League group stage.