Messi’s 61st-minute strike in Eindhoven was his 106th goal in Europe’s top competition, one more than Ronaldo managed for Real Madrid before his switch to Juventus.
Messi’s milestone goal was typically brilliant, the 31-year-old exhibiting close control in a crowded box before rifling past Jeroen Zoet.
He has scored five times in the Champions League this term and looks set for more following Barca’s progression to the knockout rounds as Group B winners.
Madrid great Raul is third on the list with 66 goals, while Karim Benzema sits fourth thanks to the 47 he has netted since arriving at the Santiago Bernabeu in 2009.
Thomas Muller – still regularly involved for Bayern Munich – and Juventus legend Alessandro Del Piero are joint-fifth, having struck 42 apiece.