Busitema dominated the opening half as they needed to find the goals to potentially progress.
Kampala University became the third side to go through to the semi-finals after edging Busitema University over two legs.
Busitema won Wednesday's return tie 1-0 but it was not enough to offset the advantage Kampala University carried from the first leg. They advanced through the away goals rule.
At Villa Park, Jacob Muga scored the only goal as Busitema struggled to build onto that.
It was the first time they had made it to the quarter-finals of the university tournament and had hoped to go even farther, but came up short.
Their attacks were often kept at bay by a resilient home defense.
Busitema dominated the opening half as - as expected - they needed to find the goals to potentially progress.
Kampala University, on the other hand, defended in numbers but stop Muga from breaking the deadlock early after restart.
The visitors pressed on but a second goal would not come.
Kampala University joins Makerere University and Uganda Martyrs University in the semifinals. The fourth team will be decided when Nkumba University takes on Gulu University.