Investigation of combwax of honeybees with high-temperature gas chromatography and high-temperature gas chromatography-chemical ionization mass spectrometry. I. High-temperature gas chromatography.
The combwaxes of the honeybee species Apis mellifera, Apis cerana, Apis dorsata, Apis laboriosa, Apis florea and Apis andreniformis have been examined by high-temperature gas chromatography. Combwax consists of a complex mixture of homologous neutral lipids. These compounds containing up to 64 carbons were chromatographed intact on a 10 m x 0.2 mm high-temperature stable SOP-50-PFD (50%-diphenyl/50%-1H,1H,2H,2H-perfluorodecylmethylpolysiloxane)-co ated Duran glass capillary column. The use of this stationary phase results in lower retention values and, at last, in lower thermal stress of the analytes. In order to minimize the discrimination effect due to adsorption and/or degradation, a two-step derivatization was performed resulting in the formation of tert.-butyldimethylsilyl esters of the long chain fatty acids and trimethylsilyl ethers of complex hydroxyesters, respectively. The derivatization procedure was optimized using a modification of the extended Donike test. In addition this test allows the quantification of the thermal stability of the derivatives performed. The derivatization procedure was applied for combwax analysis. More than 80 compounds were separated and their peak areas semiquantitatively exploited.