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A molecularly imprinted polymer (MIP) was prepared using (E)-piceatannol, a natural potential anti-epidermal growth factor receptor (EGFR) inhibitor, as the template and 4-vinylpyridine as the functional monomer. The template was isolated from a Chinese traditional Tibetan medicinal herb, Caragana jubata, by a solid-phase extraction procedure. The crude extract of this herb was loaded on the MIP column for the binding test, and two different compounds besides the template itself were specifically recognized by the polymer, which were identified to be butein and quercetin possessing potent anti-EGFR tyrosine kinase activities with I[C.sub.50] values of 10 and 15 [micro]M, respectively. Affinity and selectivity for these inhibitors and another three compounds coexisting with the template in this herb were evaluated in the chromatographic mode. For the first time, the affinity of a molecularly imprinted polymer was investigated to be correlative to the bioactivities of the analytes. The chromatographic behavior of the analytes was consistent with their activity values: the more active inhibitor was retained longer on the MIP. This research work afforded us a new approach for the effective recognition of novel anti-EGFR inhibitors from herbs by using the MIP as the receptor mimic to assay the bioactivities of reserved components, which will be very helpful in the direct separation of lead candidates for anticancer drugs.

Frontal affinity chromatography (FAC) is a simple but powerful method to analyze molecular interactions between an analyte and an immobilized ligand by calculating the extent of retardation of the elution front. By combination of FAC with a PE-Mariner electrospray ionization mass spectrometry, a very efficient and straightforward procedure was developed herein for analyzing the binding properties of different inhibitors of the epidermal growth factor receptor (EGFR). In this study, a polyclonal antibody prepared with a known anti-EGFR inhibitor coupled with bovine serum albumin was adopted as the stationary phase in the FAC system. Using the antibody to mimic the receptor, other different anti-EGFR inhibitors as well as the small-molecule half-antigen itself were recognized directly from the crude extract of herb, which afforded us a novel promising approach for the efficient screening of lead compounds or drug candidates from natural resources.

Frontal affinity chromatography (FAC) using immobilized polyclone antibodies of compound A coupled with mass spectrometry was used for the screening of affinity compounds from an extract of Phyllanthus urinaria L. Mass spectrometry was used as an analyzer of FAC. It can analyze the frontal affinity chromatogram of each compound of the extract in one program. The extract was dissolved in 2 mM N[H.sub.4]OAc at a concentration of 10 [micro]g/mL, then loaded on the immobilized antibody column, and data were collected from mass spectrometry to get a frontal affinity chromatogram. The screening of extract resulted in brevifolin, brevifolin carboxylic acid, corilagin, ellagic acid, and phyllanthusiin U. Activity analyses give high inhibitory activities to these compounds. This research work afforded us a new approach to find new leading compounds from nature or a man-made combinatorial library that have different structure styles or to find substitutes for the synthetic active compound that has high toxicity.