The Timbers were beaten 3-1 by Olimpia at the Estadio Tiburcio Carias Andino as the Hondurans narrowly advanced to the championship stage.
Both teams finished level on nine points and head-to-head goal difference from four games in Group Five.
But Olimpia claimed top spot due to the number of goals scored away from home head-to-head, which was two in the 4-2 loss in Portland last month.
Olimpia made a dream start to proceedings, scoring two goals in the opening four minutes.
Romell Quioto made the breakthrough in the second minute - firing a shot past Timbers goalkeeper Jake Gleeson that hit the underside of the crossbar - before Anthony Lozano tapped in from close range two minutes later.
Portland struggled to create anything clear-cut in the first half but the visitors started strongly after the break.
Ben Zemanski found the back of the net seven minutes into the second half, albeit courtesy of a deflection.
Olimpia hit back immediately, however, with Lozano bagging his second of the match just two minutes later.
A draw against Cruz Azul was enough for Alajuelense to progress from Group Six.
Armando Alonso's second-minute opener was cancelled out by Gerardo Torrado six minutes later as Alajuelense were held to a 1-1 draw by their Mexican visitors.
But a point still done the job for the Costa Ricans, who finished one point above Cruz Azul.
At the Estadio Azteca, Liga MX pacesetters America won through to the next round.
Needing a win to guarantee their passage into knockout rounds, Mexico international Oscar Peralta scored twice as America accounted for Comunicaciones 2-0.
America topped Group Eight ahead of the Guatemalans, who were three points adrift.
Meanwhile, DC United went through the group stages with a 100 per cent record after edging Tauro in Panama.
Conor Shanosky's 24th-minute goal was all that separated the two teams as United made it four wins from four games.
