An intermittent feeding strategy in fed-batch cultivation has been widely applied to control the accumulation of overflow toxic metabolites and oxygen limitation. However, successive depletions of carbon source by carbon-limited feeding might induce considerable stress to host cells, such as nutrient starvation, which may evoke the enhanced degradation of secreted recombinant proteins by proteolysis. To prevent this degradation, we evaluated novel nutrient feeding strategies for the production of MOX promoter-driven recombinant human serum albumin in the fed-batch cultivation of Hansenula polymorpha. Compared to conventional feeding strategies such as exponential or intermittent feeding strategy, feeding strategies developed in the present study significantly reduced the degradation of recombinant products and yielded high levels of intact recombinant human serum albumin in fed-batch Cultivation. (C) 2008 Elsevier Ltd. All rights reserved.