title: A high-throughput monolithic HPLC method for rapid vitamin C phenotyping of berry fruit.
abstract: A rapid method for the quantification of L-ascorbic acid (1) in berry fruit by HPLC with photodiode array detection is presented. L-Ascorbic acid was resolved on a C18 monolithic column with aqueous buffer, after which the column was washed with acetonitrile to remove lipophilic compounds prior to re-equilibration for analysis of the next sample. Using the monolithic column format with high mobile phase flow rates, the entire separation, wash and re-equilibration were achieved in 3 min. With the exception of gooseberry (Ribes uva-crispa), for which an interfering compound co-eluted, concentrations of 1 could be determined in a wide range of berry fruits after extraction in metaphosphoric acid without further sample preparation. Using this extraction method, recoveries of 1 in excess of 85% were achieved. Fruit or juice extracts were stable in 5% metaphosphoric acid for at least 4 h and stability could be extended to longer than 150 h by the addition of the reducing agent tris(2-carboxethyl)phosphine hydrochloride. Following validation, the method was utilised for the phenotyping of fruit in a Scottish Crop Research Institute (SCRI) Ribes nigrum L. breeding population of 300 individuals. An improved extraction method allowed extraction, quantification of 1 and data analysis to be undertaken in less than one working week.
authors.affiliation: Scottish Crop Research Institute, Invergowrie, Dundee, UK.
authors.forename: Paul G
authors.initials: PG
authors.lastname: Walker
authors.forename: Sandra L
authors.initials: SL
authors.lastname: Gordon
authors.forename: Rex M
authors.initials: RM
authors.lastname: Brennan
authors.forename: Robert D
authors.initials: RD
authors.lastname: Hancock
mesh_terms: D001205:Ascorbic Acid / Q000032:analysis*; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D005638:Fruit / Q000737:chemistry*; D010641:Phenotype; D031965:Ribes / Q000737:chemistry*
chemical_list: D001205:Ascorbic Acid