In this study, various ratios of combined red, blue, and amber light-emitting diodes (rbaLEDs) were investigated for their effect on the expression of carotenoid biosynthetic genes and carotenoid accumulation in eight Brassica microgreens. Total and individual (β-carotene, lutein, α-carotene, neoxanthin, and violaxanthin) carotenoids were increased 20-44 and 10-55%, respectively, under dose-dependent increasing amber-blue light and decreasing red in most microgreens. Lipophilic 2,2-diphenyl-1-picrylhydrazyl and ferric reducing antioxidant power antioxidant activities were significantly increased under higher amber and blue light fractions, while oxygen radical absorbance capacity was generally decreased. Under rbaLED in mizuna () microgreens, the lycopene epsilon cyclase (LYCε) expression was 10-15-fold higher, which resulted in downstream accumulation of α-carotene and lutein. Lycopene beta cyclase (LYCβ) was not significantly changed, suggesting that β-carotene, violaxanthin and neoxanthin were mainly controlled by upstream phytoene synthase and branch-point LYCε. Increased beta-ring carotenoid hydroxylase (CHXβ) expression was also consistent with lutein accumulation. This study demonstrated for the first time that amber LED was involved in the regulatory mechanism of carotenoid biosynthesis, thus a potential novel approach to production of antioxidant-rich microgreens.