Substitute Balotelli struck the winner for the second time in three games after Jordon Ibe was brought down in the area – but only after despite an apparent disagreement with stand-in skipper Jordan Henderson and Daniel Sturridge over who would take the spot-kick.
A familiar face in Demba Ba had Besiktas’ best chance, going one-on-one with Simon Mignolet in the first half only for the Liverpool goalkeeper to tip his low effort wide, while Adam Lallana blasted over the hosts’ best opportunity from six yards in the second period.
But Balotelli’s late intervention – nine days after he came of the bench to score a crucial winner against Tottenham – means Liverpool will take a slender advantage to Istanbul for the return leg on Thursday, February 26.
Brendan Rodgers brought youngster Ibe into the starting line-up for his first European game, while striker Sturridge also ended his wait for a European appearance for the Reds.
And it was Ibe who fed Sturridge early on at the byline, only for the England striker to force Cenk Gonen into a good save at his near post after a clever turn at the byline.
At the other end Veli Kavlak struck a header from a corner just over at the near post, before Ba came close to bringing hearbreak to the Kop again just 10 months after his famous goal in Chelsea’s 2-0 win at Liverpool from Steven Gerrard’s slip – one that all-but ending the Reds’ Premier League title hopes.
This time, after being put through on goal in the same position, Ba saw his low shot from just outside the area saved by Mignolet down to his left.
Alberto Moreno forced Gonen into another save with a piledriver from 40 yards out, while Henderson’s free-kick from 25 yards dropped a wide yard of the left post.
Lallana had a golden chance to give Liverpool the lead five minutes after half-time, pouncing on a mix-up in the Besiktas defence only to fire the ball over the bar under pressure from close range.
Liverpool struggled to break down Besiktas for the majority of the second half, before fine wing play from Ibe saw him tumbled down from behind by Ramon Motta with five minutes remaining.
Balotelli, who had only been on the pitch for just over 20 minutes, sent Gonen the wrong way having made sure he’d taken the spot kick, with Henderson and Sturridge both also keen take the duty.
The win means Liverpool have lost just once in 17 games in all competitions, and are unbeaten at home in 11.