The Wallaby skipper twisted his ankle in the build-up to the clash with reigning Super Rugby champions the Chiefs on the Sunshine Coast and could only sit and watched as the Reds, the 2011 winners, were beaten 20-8.
It was the first time in three years the Reds, who face the British & Irish Lions in Brisbane in their third tour outings in June, had lost a trial match. They led 8-0 at half-time, but conceded three second half tries at Stockland Stadium.
As well as missing Horwill, the Reds didn't have Will Genia in their side as he continues his recovery from knee surgery and his half-back partner, Quade Cooper, was making his pro-boxing debut instead.
Ben Lucas stepped into Genia's role and played very well and also got involved in a very unfriendly scuffle with Chief's No 10 Aaron Cruden.
David Pocock was among the try scorers for the Brumbies as they prepared for their Super Rugby opener next week against the Reds with a 38-7 win over an ACT XV in Canberra. Fit again Clyde Rathbone also helped himself to two tries to prove he is ready to return to the big time.
Meanwhile, Michael Cheika has appointed versatile back five forward Dave Dennis as his captain at the NSW Waratahs. He will lead the team into action against the Blues tomorrow in a game which will see back row dynamo Michael Hooper make his debut for the Waratahs.