The Netherlands international, who joined the club from Ajax in a £34million transfer this summer, was substituted and helped off the pitch by club medics.
Tests to diagnose the full extent of the injury and a subsequent recovery timeframe are ongoing.
The news will come as a huge blow to Mikel Arteta’s side after he had made an excellent start to life in English football.
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Arteta was asked about the extent of Timber’s injury after the victory, saying: “The doctors looked at him and they were happy for him to continue, he was as well. Straight away in the first action of the second half he made a movement and it was a bit funny.
“We took him out straight after and now we have to assess him and see what he has.”
Timber has impressed in his short time at the club, playing in several positions during pre-season.
He played at right-back on the U.S tour as well as deputising for the injured Oleksandr Zinchenko at left-back against Manchester City in the FA Community Shield and against Forest in the opening game of the season.
Arsenal are next in action against Crystal Palace at Selhurst Park next Monday.
How damaging is Timber’s injury for Arsenal?
Analysis by Jordan Campbell
It’s a huge blow for Arsenal as the Dutch defender had settled in seamlessly since joining from Ajax, providing Mikel Arteta with a variety of options across the back line.
He started both the Community Shield against Manchester City and the opening league game against Nottingham Forest at left-back, showing that he offers a lot both in defence and attack in place of Oleksandr Zinchenko.
In the pre-season games, he played all across the back four, inverting from right-back and also slotting in at centre-back for a brief spell. It underlined how important his signing was as it has made Arsenal impossible to predict.
The injury deprives them of someone who has hit the ground running with his aggressive 1v1 defending and driving runs impressing.
How will Arsenal cope with the news?
The good thing for Arsenal is that they have several versatile options.
Ben White continues to be reliable at right-back and Zinchenko is nearing a return from the calf injury that kept him out at the end of last season.
Timber would have been challenging on both flanks but Takehiro Tomiyasu is similar to him in that he can play both sides comfortably.
Kieran Tierney should not be forgotten about despite being left out the squad on Saturday, and the news could potentially change how his future is viewed with several interested suitors.
Jakub Kiwior can also cover centre-back and left-back so Arsenal don’t need to panic.
(Photo: Getty Images)