Autoecological role of steviol glycosides in Stevia rebaudiana Bertoni

Stevia rebaudiana Bertoni is a shrub native to Paraguay now a commercial source of the natural and non-calorific sweeteners of the steviol glycoside class. In stevia leaves, stevioside and rebaudioside A are the major steviol glycosides (SGs), occurring at about 10 and 3-5% of the leaf dw respectively. The large amount stored in the stevia leaves is interesting from an ecological perspective considering that the biosynthesis of SGs results from the up-regulation of a biosynthetic pathway shared with gibberellic acid (GA). The disparity in typical amount of GA (1.2 µg.kg-1 leaf fw) and SGs (10% leaf dw) in a leaf suggests a biochemical investment in SG synthesis which merits an eco-physiological role within the stevia plant. This thesis hopes to initiate this research area by looking at possible physiological roles in carbon storage, osmoregulation, mammalian herbivory and insect deterrence. The role of stevioside and rebaudioside A in carbon storage and osmoregulation was not observed given the relatively stable leaf concentration which was unresponsive to short periods of light and water treatments. Rather change in stevioside and rebaudioside A leaf concentration was gradual and related to the leaf and plant ontogeny with the highest leaf concentration occurring in the top immature leaves. It seems that synthesis occurs mostly in top young leaves with the SGs stored until leaf aging. The build-up of SGs seems to be ecologically beneficial with observed deterrence towards the generalist insect, Valanga irregularis, a grasshopper native to Australia. The leaves also encouraged general mammalian herbivory with the domestic guinea pigs, Cavia porcellus, with possible application in animal feed fortification. Complementary to the ecophysiological investigations, methods for quantification of stevioside and rebaudioside A have also been developed with High Performance Liquid Chromatography (HPLC) as the main reference method. Quick analytical methods with minimal sample preparation were also developed for possible estimation of stevioside and rebaudioside A in field application. Near Infrared Spectroscopy (NIRs) was found useful for estimating the total amount of stevioside and rebaudioside A in dry ground leaves (R = 0.87, RMSECV = 1.63). Potential transfer of the NIRs method to a handheld equipment would find commercial use in quality assessment of leaf harvests managed by stevia manufacturers. Anthrone colorimetry was also useful in measuring the amount of stevioside and rebaudioside A in solution (up to 0.06 µmol with R2 = 0.993 and RMSEC = 0.002 µmol for stevioside and R2 = 0.996, RMSEC = 0.002 µmol for rebaudioside A) and would be useful in quick estimates of liquids containing the sweetener such as beverage application.