The PGFC Academy team were edged out 2-1 by Brisbane Roar in Saturday's Foxtel Youth League clash at Lions Stadium.
Chris Harold fired Richard Garcia's men into an early lead from the penalty spot, but the hosts hit back later in the first half through Mirza Muratovic and Shannon Brady and held on to secure a victory which took them to the top of the Conference.
Glory pressed hard for an equaliser, especially in the final quarter, but were unable to avoid a third consecutive defeat which Garcia put down to his young charges switching off at key times.
"It was very similar to last week against Adelaide," he said. "We played some very good stuff in patches, but lapses of concentration have cost us again.
"We had good spells in the game, but when the lads get tired, they tend to forget the lessons that we are trying to get them to learn.
"We're asking them to make more independent choices on the field and think about the game in terms of how we want to play and what we want to. We want them to be more self-sufficient on the park and it's a steep learning curve.
"Progress is being made and now we need to push through that barrier and make sure that they can keep concentrating and focused when they're tired and under pressure.
"The boys are disappointed themselves, but it's an experience that will stand them in good stead."
A handball in the box gifted Glory the best possible start, with Harold rattling home the resulting spot-kick.
But Daniel Leck then went close as Brisbane roared back before Brady restored parity, converting from close-range after Jackson Lee had parried Muratovic's initial effort.
Oliver Duncan duly struck the woodwork as the hosts continued to threaten and on 25 minutes they bagged what proved to be the winner when Muratovic beat Lee with a fiercely-struck shot.
Daniel Stynes went close after the break and the visitors came closing to grabbing a share of the spoils in the final minute of normal time, a goalmouth scramble culminating in the ball bouncing just over the bar as a cluster of Glory and Roar players converged.
Glory return to action next Sunday when Melbourne Victory will be the visitors to Dorrien Gardens.
Brisbane Roar 2 (Brady 19', Muratovic 25') Perth Glory 1 (Harold (pen) 8')
Brisbane Roar: 1.Macklin FREKE (gk), 2.Emlyn WELLSMORE, 3.Luke DE VERE, 4.Aaron REARDON, 6.Zachary DUNCAN (17.Daniel MASKIN 72'), 7.Shannon BRADY (22.Brodie BOYCE 87'), 8.Oliver DUNCAN (19.Jay BARNETT 56'), 9.Mirza MURATOVIC, 11.Daniel LECK, 15.Stefan NIGRO, 18.Joe CALETTI
Substitutes not used: 20.Lachlan HUNTER (gk)
Yellow cards: Duncan, Barnett
Red cards: Nil
Perth Glory: 1.Jackson LEE, 2.Daniel WALSH,3.Walter SCOTT, 4.Zayd FARAH, 5.Kristian POPOVIC, 6.Henry HORE, 7.Etahn BROOKS, 8.Cameron MURRAY, 9.Chris HAROLD, 10.Daniel STYNES, 11.Gabriel POPOVIC
Substitutes not used: 12.Nicholas SPECA
Yellow cards: K.Popovic
Red cards: Nil