In one of Chile's final friendlies before they open the South American showpiece against Ecuador in June, Jorge Sampaoli saw his side fall to goals in either half from captain Javad Nekounam and Vahid Amiri.
Carlos Queiroz's Iran started brightly and were rewarded after 21 minutes when Nekounam converted Andranik Teymourian's corner.
Sampaoli, who named Arturo Vidal and Alexis Sanchez on the bench, introduced the latter at the break, although Amiri added to Chile's frustrations when he doubled Iran's advantage early in the second half.
Brazil are next up for Chile - as the sides meet for the first time since their last 16 World Cup clash - while Iran face Sweden following victory in their first outing since the Asian Cup.
Eduardo Vargas fired over following a smart turn 12 minutes in, after Chile had survived some early Iran pressure.
Masoud Shojaei then went down under a clumsy challenge from Gonzalo Jara, although the referee waved away the half-hearted penalty protests.
Nekounam eventually made Iran's pressure pay when the skipper bundled home the opener amid slack marking from Rodrigo Millar and questionable goalkeeping from Barcelona's Claudio Bravo.
The goal sparked Chile into life, with Juan Cornejo flashing a shot wide before the half-hour mark, before Fabian Orellana hit the side-netting from close range.
Mark Gonzalez saw a header beaten away by Alireza Haghighi as Chile piled on the pressure prior to the interval to no avail.
Sampaoli opted to introduce star man Sanchez at the interval, but it was Iran who found the target next as Reza Ghoochannejhad latched on to a throughball and picked out Amiri.
The Naft Tehran forward calmly finished from inside the area to put Queiroz's side firmly in control.
Matias Fernandez almost reduced the deficit with a fierce drive prior to the hour with Jose Fuenzalida angling a strike wide of the far post as Sampaoli's men continued to knock on the door.
Veteran David Pizarro was brought on to try to find a way back into the game for the South Americans, but Chile found Haghighi in fine form as the goalkeeper smartly kept out Fernandez's free-kick going into the final 20 minutes.
Despite the late pressure, Chile failed to avert a second defeat in eight post-World Cup outings, leaving Sampaoli questions to answer with the Copa clash against Ecuador in Santiago less than three months away.
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