A slow start proved Australia's downfall as they fell behind by double digits in the first quarter although a strong defensive effort in the second term kept them in the contest as they trailed by just six at half-time.
Needing another big effort in the fourth quarter, the Opals trimmed an eight-point advantage to just four inside the final few minutes but they were unable to pull themselves over the line in front of a raucous Spanish crowd.
Steph Talbot led all scorers for Australia, combining 16 points with three steals while Laura Hodges was again in the thick of the action with ten points, three rebounds and two assists.
Rachel Jarry and Katie-Rae Ebzery added eight points apiece but coach Brendan Joyce lamented his team's free throw shooting which saw them hit just 10-26 from the charity stripe.
"Overall our defence was very good but we didn't capitalise on our aggressive offence," said Joyce.
"We got what we expected in front of a very vocal home crowd with good home support.
"[It was] an excellent game for our preparation to play Spain in Spain. The game gave us another scenario of what to expect playing in Brazil."
Completing their first leg of the European tour in Spain, Australia will now head to Italy with a trip to France to come.