Changes of Trigonelline Biosynthesis under Nitrogenous Compounds in Hairy-root Culture of Iranian Fenugreek (Trigonella foenum-graecum L.)

Trigonelline is known as a valuable metabolite of alkaloids group with the therapeutic affects, especially on diabetes. In this study the effect of 3 levels of nitrogen compounds (2x, full, and half) of basic MS media including KNO3 and NH4NO3 were evaluated on trigonelline production in suspension culture for normal and hairy roots. In order to hairy root induction, 2 strains of Agerobacterium rhizogenes (ATCC15834 and K599) via co-cultivation and injection methods were used. After appearing of hairy roots a molecular analysis by PCR and genes primers of rolB and virD was conducted to confirm insertion of T-DNA segments in root genome and that the roots were bacteria-free. Then, the hairy and normal roots for investigations of trigonelline production ability were cultured in suspension MS media and the treatments were arranged in an experimental based on Randomized Complete Block Design with 3 replications. After 28 days, the trigonelline contain of roots were analyzed by HPLC method. The hairy roots were induced in the parts of plant such as crown, stem and leaf by the both strains of bacteria. The highest amount of trigonelline contain was achieved of the hairy roots cultured in MS2x media and the lowest was achieved in MS1/2N media. (13.98 and 8.74 mM/g dry mater respectively). the results indicate a main role of nitrogenous compounds (especially NH4+) and quick growth of hairy roots in enhancing of trigonelline production.