Stevia: Steviol

Steviol is ent-13-hydroxykaur-16-en-19-oic acid (C20H30O3; molecular weight 318). The compound is well absorbed by mouth.

Pharmacological actions

Energy metabolism

Steviol inhibits oxidative phosphorylation, with 40 µM producing 50% inhibition. It is, therefore, more potent than the related aglycones atractyligenin and dihydrosteviol, which produce 50% inhibition at 210 µM and 100 µM, respectively. Some of the mitochondrial actions of steviol are summarized on Table. The effects on oxidative phosphorylation are complex, involving at least three components: (i) inhibition of adenine nucleotide exchange; (ii) inhibition of NADH oxidase; and (iii) inhibition of L-glutamate dehydrogenase. Adenine nucleotide exchange between intra- and extramitochondrial spaces is involved in the shuttling of high energy phosphate groups generated in the mitochondrion to their sites of consumption in the cytoplasm. Inhibition of this exchange, therefore, implies a profound disturbance of energy flow in the cell. Unlike atractyligenin, the inhibition produced by steviol is non-competitive. Inhibition of nucleotide exchange is dependent on the presence of a free CO2H group, with inhibitory action not being seen with the methylated analog, 13-hydroxystevane, or the glucosylated analog, stevioside. The exocyclic methylene group is also involved, as reduction to dihydrosteviol decreases activity.

The complexity the action of steviol on mitochondrial function is indicated by the finding that 0.5 mM steviol inhibits DNP-stimulated ATPase by 92%, NADH oxidase by 45%, succinate oxidase by 42%, succinate dehydrogenase by 46% and glutamate dehydrogenase by 46%. In the absence of an uncoupling agent, steviol at low concentrations (0.03 mM) stimulated mitochondrial ATPase activity.

In hamster intestine, steviol (1 mM) inhibited glucose uptake and altered the morphology of intestinal absorptive cells. At 2mM, steviol also lowered mucosal mitochondrial NADH cytochrome C reductase and mucosal ATP concentration without affecting ATPase activity. The time course of fall in ATP correlates with the decrease in glucose transport. This effect on glucose transport, therefore, appears to be a consequence of inhibition of mitochondrial adenine nucleotide exchange. In fasted rats given a fructose load, steviol (0.2 mM) increased glycogen deposition in the liver. In the absence of a sugar load, steviol was without effect on glycogen levels.

Effects on blood pressure and renal function

Steviol interferes with energy metabolism in rat renal tubules, blocking oxygen uptake and glucose production. Gluconeogenesis is inhibited 50% at 0.3 mM steviol, and complete inhibition is produced at 1 mM. Oxygen uptake is inhibited 50% at 0.4 mM steviol in the presence of pyruvate (10 mM) as substrate. Steviol inhibited p-aminohippuric acid uptake in rat renal cortical slices.

Other metabolic effects

Steviol inhibits phorbol ester-stimulated ornithine decarboxylase activity in mouse skin. Steviol (200 nM) applied to the skin one hour before phorbol ester administration lowers the ornithine decarboxylase response by 63%. A stimulation in ornithine decarboxylase activity is an early event in trophic responses.

Chromosomal and mutagenic effects

There is general agreement that steviol is mutagenic in the forward mutation assay (an assay on a ‘normal’ or unmutated gene), although not in the reverse (Ames) assay (an assay on a gene containing a designed mutation). Steviol is mutagenic as a result of bioactivation. In the presence of a metabolic activating system, it is mutagenic in both Salmonella typhimurium and in human and rat liver microsomes and rat liver S-9 fraction. In some strains of S. typhimurium and E. coli, however, steviol is not mutagenic, even in the presence of a metabolic activating system. ‘Activated’ steviol binds covalently to calf thymus DNA. In S. typhimurium TM677 strain, it induces mutations of the guanine phosphoribosyltransferase (gpt) gene. Sequence studies showed that steviol induced mutations near a putative pausing site for DNA synthesis, leading to DNA duplication, deletion and untargeted mutagenesis by stimulation of misalignment and realignment of developing DNA strands.

The nature of the mutagenic metabolite is unknown. Further metabolism of steviol in rat liver is complex, with at least nine metabolites being detected. The presence of a C-13 hydroxyl and a C-16/C-17 double bond are required for activation. The major metabolite is 15α-hydroxysteviol, which is non-mutagenic both in the presence and absence of a metabolic activating system. Other metabolites include 7β-hydroxysteviol, 17-hydroxyisosteviol and ent-16-oxo-17-hydroxybey eran-19-oic acid, the latter probably being formed from the 16,17-oxide. It had been shown to be a product of acid rearrangement of steviol-16,17-oxide. The mutagenic substance has been proposed to be 15-oxosteviol, although this compound has not been detected as a metabolite of steviol. 15-Oxosteviol has been reported as bactericidal and weakly mutagenic. It forms an adduct with cysteine via an allylic Michael addition. Others, however, have strongly disputed these findings, reanalyzing the original data to show that 15-oxosteviol is without mutagenic action. On repetition of the earlier studies, no mutagenicity was found for 15-oxosteviol in Salmonella or Escherichia. These latter authors suggest that the apparent mutagenicity of steviol may be due to an impurity .

The nature of the mutagenic metabolite thus remains in doubt. It appears to be formed via a P450-mediated oxidation, as pretreatment of rats with Aroclor 1254 or phenobarbital induces metabolism. Activation is NADPH-dependent. The compound appears to be a nucleophile, as mutagenicity is decreased by GSH, cysteamine and cysteine, although not by methionine. Neither isosteviol nor the 16a,17-epoxide of steviol are mutagenic. The non-involvement of an epoxide is indicated by the finding that epoxide hydrolase does not inhibit mutagenicity. The 13-hydr o xy g roup is a requirement, as ent-kaurenoic acid is not mutagenic. Dihydrosteviol A and B, grandifloric acid and steviol acetate are also non-mutagenic, indicating that unsaturation at C16-C17 is also important for expression of mutagenicity.

On metabolic activation, steviol is mutagenic in Chinese hamster lung fibroblasts in vivo, and in a gene mutation assay in the same cells. It was without chromosomal effects on cultured human lymphocytes.

Reproductive effects

At doses of 0.75 g/kg body weight/day, steviol is toxic to pregnant hamsters and their fetuses when given on days six through ten of gestation. Steviol produces decreased maternal weight gain and high maternal mortality. The number of live births per litter is decreased and the mean fetal weight is lower. The no effect dose is 0.25 g/kg body weight/day. If 100% conversion of stevioside to steviol is assumed, this intake is equivalent to 625 mg/kg/day of stevioside. This is about 80 times the acceptable daily intake calculated by Xili et al. ().


Although much more information is needed, it does appear that steviol has the potential to be mutagenic. However, two conditions have yet to be established for humans: (i) that Stevia glycosides are converted to steviol in meaningful amounts; and (ii) that steviol is further metabolized to the mutagenic substance. In the words of a recent paper, ‘further work is necessary to predict the genotoxic risk of steviol to human beings’.


Selections from the book: “Stevia. The genus Stevia”. Edited by A.Douglas Kinghorn. Series: “Medicinal and Aromatic Plants — Industrial Profiles”. 2002