In our previous paper (Biochim. Biophys. Acta 1379 (1998) 257-263), we demonstrated that bicarbonate promotes a cleavage of lactone ring of dehydroascorbate (DHA) on the basis of in vitro experiments. In the present study, we examined the degradation of DHA in blood circulation in vivo by using a high-performance liquid chromatographic method for the determination of ascorbate (AsA), DHA and 2,3-diketogulonate (2,3-DKG), which required no pretreatment of biological fluids. When DHA was intravenously administered to rats, a rapid disappearance of DHA (t1/2 < 1 min) and a concomitant appearance of 2,3-DKG in blood circulation were observed. Approximately 90% of the administered DHA were excreted into urine as resulting 2,3-DKG (55%) and AsA (31%), respectively. Furthermore, we elucidated that rat plasma lacks an enzyme having an aldonolactonase-like activity. The result of the present study suggests that this DHA disappearance is a function of both a chemical degradation to 2,3-DKG and a reduction to AsA.