Elicited Teucrium chamaedrys cell cultures produce high amounts of teucrioside, but not the hepatotoxic neo-clerodane diterpenoids

Teucrium chamaedrys, one of the most common and investigated species of the genus Teucrium, has been used for centuries in traditional medicine for many purposes. Its phytochemical components comprise, among others, phenylethanoid glycosides (PGs) and neo-clerodane diterpenoids. Several reports have demonstrated a wide range of beneficial biological and pharmacological activities of the phenylethanoid components, while the diterpenes were shown to be strongly hepatotoxic. In this work, in vitro cultures were established from leaf explants of T. chamaedrys. Both solid (callus) and liquid (cell suspension) cultures maintained the capacity to produce PGs, with teucrioside (TS) representing the most abundant one. Cell suspensions had a lower TS content than that found in leaf extracts, but higher than that of calli. An NMR-based metabolomics approach was used to compare the product profile of intact plants vs. cell suspension cultures, and results showed that neo-clerodane diterpenes, present in the intact plant, were not detected in cell cultures. Several elicitors were supplied to cell cultures with the aim of increasing TS production, and elicitation was tested at different growth phases and by exposing cells for different periods. Methyl jasmonate and fungal mycelia from Trichoderma viridae and Fusarium moniliforme were able to significantly increase TS production if supplied at the early-exponential growth phase for 24 h. Based on the proposed link between proline and the phenylpropanoid pathways, proline accumulation in cell cultures was followed throughout a 14-day culture period, showing that it strictly reflected that of TS production. Moreover, exogenously supplied proline, and its analogue hydroxyproline, turned out to be very effective in increasing teucrioside production.