Studies on synthesis of hypericins by undifferentiated cultures of Hypericum perforatum l.

Abstract

The high demand for the therapeutically valuable natural hypericins has necessitated to look into alternative sources such as biotechnological methods for their production. Hypocotyl portions of Hypericum perforatum L. seedlings were cultured on Murashighe and Skoog (MS), and Gamborg’s B5 (B5) media supplemented with different concentrations of auxins [2,4-dichlorophenoxy acetic acid (2,4-D) or α-naphthalene acetic acid (NAA)] and cytokinins [kinetin (KN) or benzyl adenine (BA)] for the establishment of callus cultures. The frequency of callus formation was high in MS media but only two callus lines cultured under a photoperiod of 16 h in the presence of phytohormone combination NAA (1 mg/l) + KN (0.25 mg/l) and NAA (0.25 mg/l) + KN (0.025 mg/l) produced pseudohypericin and hypericin in trace amounts. The objective was to study the accumulation of hypericins by in vitro cultures and to improve or induce production by means of elicitation and precursor feeding. However, initiation of cell suspensions from such calli led to a loss of biosynthetic capacity, which could not be restored by the addition of either elicitors (copper sulphate and chitosan) or precursors (sodium acetate and emodin). The inability of cell suspensions to produce hypericins may reflect the absence of the desired biosynthetic pathways due to lack of cellular differentiation suggesting that organ cultures are appropriate culture systems for their production. Elicitation has been found to improve accumulation of hypericins in cell cultures but no reports exist regarding precursor feeding. We tested unreported elicitors and precurors but these treatments did not induce synthesis. One of the callus lines which developed 3-5 shoots accumulated hypericns confirming the results reported previously.

Keywords: hypericins, Hypericum perforatum, callus cultures, elicitors, precursors