DEVELOPMENT OF A RAPID MICROPLATE ASSAY FOR DETERMINING PUNGENCY IN ONION BREEDING PROGRAM

Onions with low levels of pungency and sweet flavor are becoming popular with consumers on an international level. The trend of greater demand in the international market will continue as sweet onions are introduced and promoted as a specialty item. Large-scale screening of onion bulbs for pungency requires a fast and cost-efficient method. The original method which was proposed by Schwimmer and Weston (1961), it is slow and tedious when large quantity of samples is analyzed in breeding programs. Moreover, it involves the use of dinitrophenylhydrazine, a chemical difficult to handle and very toxic. In our breeding program for developing low pungency cultivars, the screening is performed for more than 3000 bulbs annually. A 96-well microplate procedure for pyruvic acid analysis was developed in order to screen a large number of onion bulbs in our breeding program. The new method is based on the selective reaction of p-dimethylaminobenzaldehyde with enzymatically produced pyruvic acid. Analysis conditions were optimized with Response Surface Modelling using a Box-Behnken Design. With this method it is possible to determine the pyruvic acid content in 96 onion bulbs in just a few minutes. In a test using our cultivars, this method showed high correlations with spectrophotometric method (R2>0.95).