Councillors voted down the proposals on the grounds of visual impact on the Crichton site in Dumfries.
The Dumfries and Galloway College plan had been recommended for approval.
A spokesman for the college said it was "disappointed" by the decision but would now have to look at a smaller scheme or alternative location.
The further education body had earmarked a shared funding package for the project with Sportscotland and Dumfries and Galloway Council.
It will now explore the possibility of a smaller-scale development in the same area or a full-size pitch at another site.
A Â£410,000 plan to develop a first full-size, third-generation all-weather football pitch in Dumfries and Galloway has been rejected.