Yeast strain Y68, producing a large amount of pullulan, was isolated from the leaves collected in the south of China. This strain was identified to be Rhodotorula bacarum by BIOLOG analysis and routine yeast identification. This is the first time to report that pullulan was produced by R. bacarum. The optimal medium for pullulan production by this strain was 8.0% (w/v) glucose, 2.0% (w/v) soybean cake hydrolysate, 0.5% (w/v) K2HPO4, 0.1% (w/v) NaCl, 0.02% (w/v) MgSO4.7H(2)O, 0.06% (w/v) (NH4)(2)SO4, pH 7.0. The optimal cultivation conditions for pullulan production by this strain in 300-ml shake flask containing 50 ml of medium were observed at 28 C and with 180 rpm. Under these conditions, 5.9% (w/v) pullulan was produced within 60 h. This was the highest pullulan yield produced by yeasts obtained so far. No pigment in the medium was observed during the fermentation, suggesting that strain Y68 was a non-pigmented yeast strain. (C) 2003 Elsevier Science Inc. All rights reserved.