Simple and efficient isolation of cordycepin from culture broth of a Cordyceps militaris mutant

•The culture broth of the Cordycepin militaris mutant G81-3 had high cordycepin concentration.•Cordycepin was efficiently isolated from the culture broth by crystallization.•The combination of temperature and pH shifts was the most effective crystallizing way.•The purification by crystallization provided a highly purified cordycepin with a high yield.

Isolation of cordycepin from the culture broth of Cordyceps militaris mutant was investigated. Based on the solubility curve, three crystallizing processes, temperature shift (process I), pH shift (process II), and pH shift followed by temperature shift (process III) were carried out. Process III was the most promising method regarding both purity and yield.