Anti-cancerous/Anti-bacterial Activities of AllicinGenerated In situ from Diastereo Pure Alliins byAlliinase

摘要

Aims: To facilitate allicin generation in-situ from pure diastereomers of alliin by enzymatic reaction of alliinase and assess its anti-cancerous/anti-bacterial activities.Study Design: Chemical synthesis and in-vitro assay of anti-cancerous/anti-bacterial activities.Place and Duration of Study: Protein Research Laboratory, Research Resources Center, University of Illinois at Chicago, between February 2014 and February 2015.Methodology: Cancer cell viability assay MTT assay, bacterial plate-diffusion growth inhibition assay, and flow cytometry cell cycle analysis have been used to demonstrate the anti-cancerous/anti-pathogen activities of the in-situ allicin. Diastereomers of alliin are produced by H2O2 oxidation of deoxyalliin, which is prepared by mixing L-cysteine and allyl bromide. Deoxyalliin and diastereomers of alliin are purified to high purity with repeated fractional crystallization. In addition, fluorenylmethyloxycarbonyl (Fmoc) protected alliin and alliin methyl ester are synthesized and purified with RP-HPLC to test the importance of amino and carboxyl groups of alliin in alliinase enzymatic reaction. Alliinase is produced by a simple and effective method from an aqueous garlic extractResults: Results from spectrophotometric alliinase activity assay indicate that (+)-L-alliin is more reactive toward alliinase than (-)-L-alliin, and both amino and carboxyl groups of the cysteine portion of alliin are critical in alliinase enzymatic reaction. Results from cancer cell viability assay MTT assay, bacterial plate-diffusion growth inhibition, and flow cytometry cell cycle analysis confirm that the in-situ allicin is as active as allicin purified from aqueous garlic extract or allicin synthesized chemically in a dose-dependent manner.Conclusion: We describe here facile pathways to synthesize diastereomerically pure alliins and isolate allinase. The in-situ allicin conversed from alliin by allinase is very active. The data obtained here provide useful information on the design of the in-situ allicin strategy.