No abstract

As flaxseed mainly accumulates lignans (secoisolariciresinol diglucoside and matairesinol), these compounds were barely or not detected in plant cell suspensions initiated from Linum usitatissimum. In contrast, these cell suspensions were shown to accumulate substantial amounts of a neolignan identified as dehydrodiconiferyl alcohol-4-beta-D: -glucoside (DCG) (up to 47.7 mg g(-1) DW). The formation of this pharmacologically active compound was evaluated as a function of cell growth and in relation to phytohormone balance of the culture media. After establishment of efficient culture conditions, production of DCG was investigated in immobilized plant cell suspensions initiated from plantlet roots of L. usitatissimum. The results indicate that immobilization enhances the DCG production up to 60.0 mg g(-1) DW but depresses the cell growth resulting in no improvement of the total DCG yield. Nevertheless, with immobilized cell suspensions, a release of DCG into the medium is observed allowing an easier recovery.

Phenylpropanoids represent a broad range of secondary metabolites in plants, in which they are involved in defense mechanisms. This study deals with the NMR identification of a phenylpropanoid, which belongs to the class of neolignans, dehydrodiconiferyl alcohol-4-b-d-glucoside (DCG), in in vitro cultures of Linum usitatissimum. The combination of 1-and 2-D NMR experiments such as COSY, HMBC and HMQC allowed the identification of this compound's structure unambiguously. In order to evaluate its implication in defense mechanism, the L. usitatissimum suspension cells were placed together with fungal extracts. Consequently, the DCG concentration decreased dramatically after 96 h of treatment. In correlation, the phenylcoumaran benzylic ether reductase (PCBER) expression increased rapidly and constantly immediately after elicitation until 96 h post elicitation, as shown by semi-quantitative reverse transcriptase polymerase chain reaction (RT-PCR). These two results are in agreement, since the aglycone form of DCG is one of the two substrates of PCBER, thus suggesting PCBER activation in plant defense mechanisms.

De-differentiation of transformed root cultures of Datura stramonium has previously been shown to cause a loss of tropane alkaloid synthetic capacity. This indicates a marked shift in physiological status, notably in the flux of primary metabolites into tropane alkaloids. Nitrogen metabolism in transformed root cultures of D. stramonium (an alkaloid-producing system) and de-differentiated suspension cultures derived therefrom (a non-producing system) has been compared using Nuclear Magnetic Resonance (NMR) spectroscopy. (15)N-Labelled precursors [((15)NH(4))(2)SO(4) and K(15)NO(3)] were fed and their incorporation into nitrogenous metabolites studied using Heteronuclear Multiple Bond Coherence (HMBC) NMR spectroscopy. In both cultures, the same amino acids were resolved in the HMBC spectra. However, marked differences were found in the intensity of labelling of a range of nitrogenous compounds. In differentiated root cultures, cross-peaks corresponding to secondary metabolites, such as tropine, were observed, whereas these were absent in the de-differentiated cultures. By contrast, N- acetylputrescine and gamma-aminobutyric acid (GABA) accumulated in the de-differentiated cultures to a much larger extent than in the root cultures. It can therefore be suggested that the loss of alkaloid biosynthesis was compensated by the diversion of putrescine metabolism away from the tropane pathway and toward the synthesis of GABA via N-acetylputrescine.