The left-back completed the full game for his country but could not prevent his side from falling to a 2-0 defeat to Denmark.
Having claimed just one point from four games going into this clash, the Оrlovi knew their chances of progression were slim - and they were dealt a significant blow at the Parken Stadium.
Despite dominating possession for much of the game, Serbia were wasteful in front of goal and found former Blue goalkeeper Kasper Schmeichel in inspired form, while the hosts were simply more clinical.
Yussuf Poulsen opened the scoring with the home side’s first attempt on target, collecting Nicklas Bendtner’s lay-off before drilling into the bottom corner.
Just after the half-hour mark, the Danes should have stretched their lead, when the referee awarded a penalty for a challenge by Matija Nastasic on Pierre-Emile Hojbjerg. Daniel Agger however, saw his resulting spot-kick saved.
Just after the break, Kolarov would have been left wondering how he hadn’t managed to claim an assist, when he delivered a cross to Aleksandar Mitrovic, whose header was somehow superbly pushed onto the crossbar by Schmeichel.
Serbia spurned more chances as they pressed for an equaliser - and they were dealt a suckerpunch minutes before the end when Jakob Poulsen sealed the win for the hosts.
On the counter-attack, he drove forward and slotted into the bottom corner to keep his side second in Group I, leaving Kolarov’s men joint bottom with a single point.
Elsewhere, France – who will host the 2016 tournament and therefore do not need to qualify – fell to a friendly defeat to Albania in Elbasan.
Following last week’s 4-3 loss to Belgium, Les Bleus were eager to return to winning ways – but they came up against an inspired home side, who restricted their visitors to half-chances and in fact should have increased their margin of victory.
Bacary Sagna completed the final 20 minutes of the encounter, replacing Christophe Jallet with the away side already trailing, courtesy of Ergys Kace’s first-half free-kick.
Despite a late rally, the defender could not help his team back into the game and Albania hung on for the win.
Eliaquim Mangala was an unused substitute for Didier Deschamps’ men.
