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• Phenolic compounds profiles were differentiated in Rabbiteye and Highbush. • Were identified 44 flavonols and chlorogenic acid in blueberries. • Quercetin HMG glycoside was found for the first time in blueberries. • Glucuronides of syringetin and laricitrin were found for the first time in blueberries. • Rhamnosyl-galactosides of myricetin, quercetin and isorhamnetin were also found for the first time. Although blueberries are widely studied, little information exists on their composition and content of flavonol glycosides. Most studies identify only a few flavonols in blueberries due to separation and identification issues. In the present study, we identified 44 flavonols and chlorogenic acid in 30 samples of Highbush and Rabbiteye blueberry, using HPLC-DAD-ESI-MSn. Highbush group fruits presented mainly quercetin-3-galactoside in their composition, while Rabbiteye group fruits exhibited higher levels of quercetin-3-rhamnoside and quercetin-3-glucuronide. Among the identified flavonols, 8 acylates (acetyl and hydroxymethylglutaroyl) were found, of which quercetin-3- O -[4″-(3-hydroxy-3-methylglutaroyl)]-α-rhamnoside was found for the first time in blueberries. This compound is exclusive to the cultivars Florida and Powderblue, where it is present in high quantities. Glucuronides of syringetin and laricitrin, and rhamnosyl-galactosides of myricetin, quercetin and isorhamnetin were also found for the first time in blueberries. The Principal Component Analysis showed that blueberry groups can be distinguished based on their phenolic compound profile. [ABSTRACT FROM AUTHOR]