METHODS FOR SEPARATION OF FREE, SHORT, MEDIUM, AND LONG-CHAIN FATTY-ACIDS AND FOR THEIR DECARBOXYLATION

A system for separation and quantitation of free fatty acids by HPLC i s described. This system provides for separation of short, medium, and long chain, saturated from unsaturated, branched from unbranched free fatty acids, and iso- from anteiso-branched acids of equal carbon num ber, and for their quantitation. An efficient method and inexpensive a pparatus for performing the Schmidt degradation on separated acids is also presented. Standard and plant-derived branched and straight chain fatty acids were investigated using these methods. The latter were ra diolabeled in vivo and released from trichome-exudated sugar esters by saponification. Free fatty acids were separated by reversed-phase C-1 8 HPLC using gradients of acetonitrile/H2O2 containing 0.01 M H3PO4. S eparated acids were converted to their potassium salts and transferred to easily and inexpensively fashioned, triple-well septum vials conta ining a center well for trapping CO2. Sulfuric acid was placed in a ti p well atop the center well and the vial was closed. Tipping the vial allowed the acid to spill to the sample without the need to puncture t he septum or transfer acid via a syringe. The system allowed for the i ntroduction of an additional CO2-trapping reagent, midreaction. This a rrangement improved CO2 recovery to 99-100% and its reproducibility to about +/-3%. (C) 1995 Academic Press, Inc.