Production of shikonin derivatives in transgenic roots of Lithospermum canescens (Michx.) Lehm. cultivated in mist bioreactor

Lithospermum canescens (Boraginaceae) is a common prairie plant native to North America. The natural roots of L. canescens yielded the isolation of acetylshikonin (ACS) 24.35mg/g D.W and isobutyrylshikonin (IBS) 14.48mg/g D.W. The biological studies of ACS and IBS demonstrate their immunomodulatory effect on the cellular and humoral immunity, ACS also inhibited cutaneous angiogenesis induced by L-1 sarcoma cells in Balb/c mice [1, 2].

The influence of two media LS [3] and M9 [4] on shikonin derivatives production in two L.canescens hairy root lines Lc-1A and Lc-1D was investigated in 3l mist bioreactor. After 4 weeks of culture the growth and shikonin derivatives content in roots and medium was determined. Better growth of both cultivated hairy root lines was observed in M9 than in LS medium. Biomass increase achieved by hairy roots of Lc-1A line was 10-fold and 27-fold, and for Lc-1D roots 2-fold and 8-fold in LS and M9 media, respectively.

The M9 medium showed beneficial effect on red pigments accumulation in comparison to LS medium. The highest total ACS and IBS content 6.17mg/l was found in the culture of Lc-1A hairy root line. The roots of Lc-1D line produced 4.73mg/l of ACS and IBS. The ACS and IBS total content obtained in root culture performed in LS medium was 1.68mg/l for Lc-1A while for Lc-1D 1.11mg/l. The 4-fold rise in shikonin derivatives yield was obtained during culturing hairy roots in M9 medium.

References: 1. Pietrosiuk, A. et al. (2004) Die Pharm. 59: 199–204.

2. Pietrosiuk, A. et al. (2004) Acta Polon Pharm Drug Res 61: 379–382.

3. Linsmaier, E.M., Skoog, F. (1965) Planta 72: 146–154.

4. Fujita, Y. (1988) Biotechnology in Agriculture and Forestry (ed by Y.P.S. Bajaj). Medicinal and Aromatic Plants, II.3, 4: 225–236