An Escherichia coli acid phosphatase gene with phytase activity was cloned into pBBRT plasmid and transformed into Pseudomonas putida. A complex medium was developed for phytase production from this recombinant strain. After cultivation at 30 degreesC for 48 h crude phytase was isolated and characterized. The optimum temperature and pH for activity were 55 degreesC and pH 4.0, respectively. Enzyme activity was enhanced by storage for 2 h at temperatures ranging from 30 to 50 degreesC. Upon storage at various pH, there was one high-activity-peak at pH 4.0 and another at pH 8.0 with remaining activities of 100 and 110%, respectively. Since this recombinant phytase was sensitive to trypsin and the bile acids. taurochloric and deoxycholic acid it would not be suitable for direct addition to animal feed but might be useful for pretreatment of raw materials prior to feed formulation. Digestion of a pre-mixed chicken feed increased the available phosphate and protein for animal consumption. (C) 2004 Elsevier Ltd. All rights reserved.