Production of Coenzyme Q10 from Tobacco Cell Culture

Abstract

A callus from explant of tobacco was induced with MS medium (supplemented with 2.0 ppm of 2,4-D, 0.2 ppm of kinetin, 3 w/v% of sucrose). The leaf of Nicotiana tabacum L. cv. Wisconsin 38 were used for the callus development. Coenzyme Q10 was produced by suspension culture of induced callus. After suspension culture system was established, fractionated the cells into different sizes by using stainless steel sieves (mesh no. 10, 18, 45) for synchronizing the cells. The growth curve was obtained and the suitable subculture period was determined. Q10 was determined by HPLC with Nacalai Cosmosil 5C18-AR column by UV detector under 275 nm. Cells entered stationary phase on the eighth day. Dry cell weight, total Q10 and Q10 content reached 10.8 g/L, 668 μg/L, 61.9 μg/g dry wt, respectively.
The optimum concentrations of carbon source (sucrose, glucose ,fructose and mannitol), nitrogen source (peptone, yeast extract, casein, casamino acid, (NH4)2SO4 and KNO3) and hormones were found. The results showed that the optimum concentration of sucrose was 3 w/v %. Mannitol (7%) favored the accumulation of Q10 content, but suppressed the cell growth. The NH4+/NO3－ ratio of 20/40 maximized the total Q10 productivity and the ratio of 30/30 maximized the Q10 content.
Under pH 5.7 and temperature 26℃, both cell growth and Q10 formation were high. 1000 ppm of L-tyrosine dosed as a precursor enhanced Q10 productivity. The other precursor, 4-hydroxybenzoic acid, p-coumaric acid and mevalonic acid did not enhance the productivity of Q10. It was found that the Mg2+ repressed Q10 formation and cell growth.