Rapid, sustainable RP-HPLC-DAD protocols for simultaneous determination of flavonols, anthocyanins, and tryptophan in onion bulbs: A multi-analytical approach

Various onion ( Allium cepa ) varieties – common onion (var. cepa ), top onion (var. proliferum ), and potato and shallot (var. aggregatum ) – share the same secondary metabolites. Here, we developed and validated single-step, sustainable ultrasound-assisted extraction (5 min) and rapid HPLC separation (15 min) protocols, designed to reduce analytical workload, minimize toxic solvent consumption, and shorten analysis time while enabling the simultaneous determination of 10 flavonols, 5 anthocyanins, and tryptophan in onion bulbs. At a 1:100 ( w / v ) material-to-solvent ratio, neither solvent type (methanol vs. ethanol) nor concentration (40–95%, v / v ) significantly affected extraction efficiency, and re-extraction was found to be unnecessary, except for quercetin 3,7,4′-O-triglucoside. The effects of genetic diversity across 40 genotypes of potato onion (11 red, 29 yellow), bulb layer, and 1 yellow common onion on secondary metabolites accumulation were investigated. Aqueous ethanol (55%, v / v ) with 1% formic acid was selected as a balanced, sustainable extraction medium providing high-efficiency simultaneous recovery of the target phytochemicals for high-throughput genotype screening in long-term onion breeding programs.