A molecularly imprinted polymer (MIP) has been prepared using diniconazole, a triazole-based fungicide with a broad antifungal spectrum, as the template and the mixture of methacrylic acid and acrylamide as the functional monomers, and ethylene glycol dimethacrylate as the crosslinker. HPLC was used to study the molecular recognition mechanism that regulates the binding behavior, and to evaluate the binding performance of these MIPs for the template and for paclobutrazol. Under optimal condition, baseline separation could be obtained. The results showed that the MIP had high affinity and selectivity for diniconazole in water-containing system, and the retention of diniconazole became stronger on increasing the water content. This behaviour gives the MIP the potential for use in the enrichment, separation and detection of diniconazole in biological fluids. In addition, a solid-phase extraction column packed with diniconazole-imprinted polymers was used to enrich the diniconazole extraction recovery range from 63.8% to 80.5% (C) 2005 Elsevier B.V. All rights reserved.