Metodologia direta para derivatização de ácidos graxos em abelhas Apis mellifera Linnaeus, 1758 (Hymenoptera: Apidae)

Abstract

Lipids in living organisms are involved in important metabolic functions. For years, the study of fatty acids in bees has been growing, with the aim of understanding physiology, behavioral activities, communication, and nutrition, among other things. The honeybee Apis mellifera L. is one of the most studied species, due to its economic and ecological importance. Traditional methodologies are used for the preparation and analysis of fatty acids in animal samples. The Bligh and Dyer method and the Soxhlet technique are the most widely used, but they have disadvantages, such as the need for a large sample size and the thermal decomposition of temperature-sensitive compounds in the case of Soxhlet. Therefore, this study aims to apply a direct methodology to obtain methyl esters from small samples of the bee species A. mellifera, in less time and with fewer samples. To this end, Complete Factorial Design 22 was used to evaluate the best combination of basic (NaOH) and acid (H2SO4) reaction time factors in samples of worker bees. The data obtained were analyzed by Analysis of Variance (ANOVA) and visualized using response surface and contour graphs to determine the parameters that had a statistically significant influence on fatty acids. In the most promising condition of the study, samples of worker bees, drones, and queens were prepared using both the direct fatty acid derivatization method and the traditional Bligh & Dyer method, followed by a derivatization step. Subsequently, the methyl esters obtained were analyzed by gas chromatography coupled with a flame ionization detector (GC-FID). The results revealed that the OP1 treatment, with a reaction time of 5 minutes for the basic stage and 5 minutes for the acid stage, proved to be particularly efficient for the analysis of oleic acid in bees. Compared to the Bligh & Dyer method, the direct method showed a higher relative percentage of oleic acid in queens (81.81%), workers (83.45%), and drones (85.27%). In addition, it proved to be efficient for myristic acid in queens (1.34%), workers (1.93%), and drones (1.99%), and for lauric acid in workers (0.58%) and drones (0.31%). Thus, the direct method, with a total time of 10 minutes, proved its effectiveness for determining the abundance of oleic, myristic, and lauric acids. In addition to its efficiency in this lipid profile, it is a simplified and rapid methodology, ideal for adoption in laboratory routines for the analysis of the fatty acid profile in bees.