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Saccharomyces cerevisiae cells incubated with D-glucose (D-Glc), D-galactose or D-mannose (D-Man) synthesised D-erythroascorbic acid (D-EAA) but not L-ascorbic acid (L-AA). Accumulation of D-EAA was observed in cells incubated with D-arabinose (D-Ara) whilst accumulation of L-AA occurred in cells incubated with L-galactose (L-Gal), L-galactono-1,4-lactone and L-gulono-1,4-lactone. When S. cerevisiae cells were incubated with D-[U-¹⁴C]Glc, D-[U-¹⁴C]Man or L-[1-¹⁴C]Gal, incorporation of radioactivity into L-AA was observed only with L-[1-¹⁴C]Gal. Pre-incubation of yeast cells with D-Ara substantially reduced the incorporation of L-[1-¹⁴C]Gal into L-AA. Our results indicate that, under appropriate conditions, yeast cells can synthesise L-AA via the pathway naturally used for D-EAA biosynthesis.