Repression of Carotenoid Accumulation by Nitrogen and NH4+ Supply in Carrot Callus Cells In Vitro

Abstract

The effect of mineral nutrition on the accumulation of the main health beneficial compounds in carrots, the carotenoid pigments, remains ambiguous; here, a model-based approach was applied to reveal which compounds are responsible for the variation in carotenoid content in carrot cells in vitro. For this purpose, carotenoid-rich callus was cultured on either BI (modified Gamborg B5) or R (modified Murashige and Skoog MS) mineral media or on modified media obtained by exchanging compounds between BI and R. Callus growing on the BI medium had abundant carotene crystals in the cells and a dark orange color in contrast to pale orange callus with sparse crystals on the R medium. The carotenoid content, determined by HPLC and spectrophotometrically after two months of culture, was 5.3 higher on the BI medium. The replacement of media components revealed that only the N concentration and the NO3:NH4 ratio affected carotenoid accumulation. Either the increase of N amount above 27 mM or decrease of NO3:NH4 ratio below 12 resulted in the repression of carotenoid accumulation. An adverse effect of the increased NH4+ level on callus growth was additionally found. Somatic embryos were formed regardless of the level of N supplied. Changes to other media components, i.e., macroelements other than N, microelements, vitamins, growth regulators, and sucrose had no effect on callus growth and carotenoid accumulation. The results obtained from this model system expand the range of factors, such as N availability, composition of N salts, and ratio of nitrate to ammonium N form, that may affect the regulation of carotenoid metabolism.