General method for extraction of blueberry anthocyanins and identification using high performance liquid chromatography–electrospray ionization-ion trap-time of flight-mass spectrometry

A systematic approach for optimizing the extraction and identification of anthocyanins from blueberries was explored using HPLC-UV and HPLC–ESI-IT-TOF-MS. Sample homogenization effects, extraction solvent selection, type of acid, and amount used in extraction solvent were investigated. A mixture of methanol:water:trifluoroacetic acid (70:30:1, v/v/v) was found to be the best solvent system for blueberry anthocyanin extraction. Differences in total anthocyanin content due to commercial blueberry processing were explored as an application using the optimized extraction technique and HPLC-UV analysis. A methodical system for anthocyanin identification by HPLC–ESI-IT-TOF-MS without the use of standards was also reviewed and applied. Consideration was given to elution order by chromatographic separation with selective detection at 520 nm, high mass accuracy m/z values, tandem MS fragmentation, and previously published literature. Overall, 25 anthocyanins from a wild type highbush blueberry were identified and reported.