Production of rosmarinic acid in ocimum sanctum (l.) cell suspension cultures by the influence of growth regulators

Abstract

The rosmarinic acid (RA) accumulation in cell suspension cultures of Ocimum sanctum (L.) was investigated. Callus was initiated from leaf explant on Murashige and Skoog’s (MS) medium supplemented with 2,4 dichlorophenoxyacetic acid (2,4-D) 1 mg/L and kinetin (KIN) 0.1 mg/L. Suspensions were established by transferring friable callus to MS liquid medium supplemented with growth regulators such as 2,4-D (1 mg/L) + KIN (0.1-0.5 mg/L), 2,4-D (0.5–2.5 mg/L), NAA (0.5-2.5 mg/L), and IAA (0.5-2.5 mg/L) individually. After stabilization of cell suspension with five subcultures, continuous culture duration of six weeks on MS liquid medium of the same growth regulator combinations resulted in progressive callus growth and RA accumulation. The highest RA (104 mg/L) content and biomass growth (17.8 g/L) was observed in the cultures supplemented with 2,4-D (1 mg/L) and KIN (0.1 mg/L). It was two-fold higher than that found in leaf callus induced on MS solid medium. RA formation was paralleled with cell growth. RA was quantified using reverse phase high performance liquid chromatography with reference standard. RA was isolated from suspension harvested biomass and characterized by spectral analysis.