SYNTHESIS AND PURIFICATION OF 36 AROMATIC CHOLINE ESTERS AS STANDARDS

Natural aromatic choline esters occurring in seeds of Brassicaceae are determined, after extraction with a mixture of methanol and acetic acid 0.01N (70/30), by high performance liquid chromatography. So, from a qualitative and a quantitative point of view, it is very useful to have in the laboratory pure standards for confirm retention times, response factors and for the constitution of u.v., infra-red or mass spectra libraries. As choline esters are not available by a commercial way, we decided to synthesise different aromatic choline esters (36). The structure of the acidic part of all the choline esters prepared are derived from benzoic or cinnamic acids with hydroxy or methoxy substituants in ortho, para or meta position. Pure aromatic choline esters were synthesised according to a fast method using bromocholine bromide. An aromatic acid (in excess) in methanol is first neutralised by NaOH (0.1 N). Then bromocholine bromide in methanol is added in the evaporation flask. Solvent is evaporated to dryness using a rotative evaporator Büchi (40°C). The flask containing the dried mixture is placed in an oven (107°C) during 5 hours, avoiding carbonisation. The choline ester is taken up with 2 x 3 ml of distilled water and purified on a SP Sephadex C25-120 resin. After washing with distilled water, the ester is eluted HCl (1N). The elution is followed by measuring u.v. absorbance at 280 nm. The eluate is evaporated to dryness with a rotative evaporator (Büchi) and the crystals obtained are washed with acetone. The purity obtained for all the choline esters produced was close to 98% and total amount between 5 to 100 mg. An u.v. spectra library of each aromatic choline esters has been constituted for rapid identification when a diode array detector is coupled with the HPLC.