In Vitro-Cultures of Arctostaphylos-Species. III: Phytochemical Comparison of in Vitro-Cultures and Outdoor Plants

Abstract

Tissue cultures originating from different organs (leaves, leafstalks, seed vessels, anthers, and roots) of Symphyrum officinale L. (Boraginaceae) were initiated under various growth conditions and subcultured several times (first callus generation) (1). From all these calli whole plants could be regenerated. The regenerated plants were used again for the preparation of tissue cultures (second callus generation). The different calli and the regenerated plants were analyzed with respect to the fructan synthesizing capacity. Calli derived from the leaves of the original plant synthesized fructan. In contrast, calli derived from seed vessels, anthers, and roots, which are known to contain large amounts of fructan, were not able to synthesize fructan. The regenerated plant showed an ability for fructan synthesis only if the originating callus synthesized fructan.The calli of the second generation obtained from leaves and roots of regenerated plants containing fructan had the capacity of fructan formation. The calli of the second generation obtained from leaves and roots of regenerated plants without fructan were not able to synthesize fructan.From these data it can be concluded that the calli of the first generation prepared from roots, seed vessels, and anthers have lost the ability for fructan synthesis. Calli initiated from leaves and leaf-stalks preserved the capacity of fructan formation even after many calli generations and regeneration to whole plants.Different phytohormones used in the tissue cultures had only minor effect on the fructan formation.The fructan from leaves and roots of the original plant and from calli was analyzed by TLC (2, 3); by GC (4); by gel filtration with TSK 2000 SW columns; and by GC/MS after methylation analysis (5). The fructan represents the inulin type with a DPof33.Our investigation was concerned with the area-and timedependent variability of the phenolic glycosides arbutin and methylarbutin. We analysed 500 samples of European wild grown plants and of in vitro-cultures raised from explants of two outdoor plants (1).Except the leaves of wild grown plants all samples were freeze dried and fresh (FW) and dry (DW) weights measured. Screening test was an HPTLC-thermoapplication-method (2) with a detection minimum of less than 0,01 % DW phenolic glycoside.Quantitative analysis was performed by UV-scanning of HPTLC-plates and controlled by several statistical examinations as well as by reversed phase HPLC (3).Among the European wild grown plants we found a very high variability of arbutin yields (6,3-14,6% DW) clearly not depending on localities. Moreover we firmly concluded from our results that the frequently mentioned (4), methylarbutin producing chemical race of A. uva-ursi cannot be defined precisely neither in territorial regard nor chemically. During the vegetation period outdoor plants showed changing arbutin yields only in young leaves, the increasing arbutin accumulation being correlated with increasing DW/FW-ratio.Mostly plant in vitro-cultures yield low amounts of secondary...