REVERSED-PHASE LIQUID-CHROMATOGRAPHIC ISOLATION OF LUBIMIN AND SOLAVETIVONE FROM HYOSCYAMUS-MUTICUS HAIRY ROOT CULTURES

A high-performance liquid chromatographic reversed-phase procedure was developed to estimate the concentration of sesquiterpene phytoalexins secreted into the media by Agrobacterium transformed root cultures of Hyoscyamus muticus. An isolation procedure based on solid-phase extra ction (C-18 Sep-pak) results in very rapid analysis and gives full rec overy, compared to sequential chloroform extraction. A simple isocrati c procedure permits quantification of sesquiterpene production by thes e root cultures in response to exposure to fungal elicitors. Calibrati on curves for lubimin and solavetivone content were obtained by fracti onating sufficient material to provide a measurable weight. The identi ty of these compounds was confirmed by TLC, UV, GC-MS, and NMR. This e xperimental system is being used to examine the regulation of sesquite rpene biosynthesis, and to develop large-scale processing techniques f or the production of secondary metabolites from plant roots grown in b ioreactor systems.