Characterization of 4 Cupressus arizonica extracts using different solvents

Abstract

Rationale Cypress spp. pollen induced pollinosis is increasing worldwide. The objective of this study was to characterize 4 Cupressus arizonica extracts using different solvents and in vivo and in vitro techniques. Methods Commercially available Cupressus arizonica pollen was used to prepare 4 types of extracts extracts. Extract A was extracted in PBS (0.01M); extract B in ammonium bicarbonate (40 mM); extract C, in ammonium bicarbonate (40 mM), followed by precipitation with ammonium sulfate; and extract D consisted of the supernatant obtained after the centrifugation of extract C. All extracts were characterized by SDS-PAGE and immunoblot. Cup a 1 content was measured by scanning densitometry. Thirty individuals were skin tested with the 4 extracts at 2 mg/ml of freeze-dried material. Specific IgE was evaluated by direct ELISA and immunoblots. Polysaccharide content was evaluated by the Anthrone method. Results The diagnosis capacity of extract C was better than of extracts A, B and D. All skin prick tests were negative with extract D. Extract C had the highest protein and major allergen content and induced greater wheal sizes than the other extracts (p<0.05). Cup a 1 content was 7.2, 14.1 and 23.1 μg/mg of freeze-dried material in extracts A, B and C, respectively. No differences were found in the polysaccharide content in these extracts. Conclusions The increase in the protein and major allergen content and the removal of irrelevant material produces a better extract for the in vivo and in vitro diagnosis of C. arizonica induced respiratory allergies.