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Colony integrity and survival in honeybees is regulated by chemical signals that are actively produced by glands and synergically transmitted between the members. A number of these signals regulate the typical age-related division of labor among the worker bees performing different tasks at different ages. In this study, the analysis of the cuticular profiles in Apis mellifera ligustica Spinola, 1806 workers at various tasks (newly emerged, nurse and forager bees) was performed using in vivo solid-phase microextraction gas chromatography-mass spectrometry (SPMEGC- MS). The use of in vivo SPME shed new light on the complexity of the molecular pattern on the epicuticle of honeybees. The identified molecules are mainly hydrocarbons, saturated and unsaturated carboxylic acids, and to a less extent, esters, sterols, aldehydes, and alcohols. Their relative abundance between the three task groups was evaluated using descriptive statistics and multivariate pattern recognition analysis (i.e., principal component analysis, PCA, and linear discriminant analysis, LDA). Eleven molecules namely nonacosane, pentacosane, (Z)-12-pentacosene, 11-tricosene, 11-methylnonacosane, squalene, 13-methylheptacosane, heptacosane, heneicosane, docosane and tricosane, occur with high frequency in newly emerged, nurse, and forager bees. The compounds that contributed the most for the separation of the three task groups in the PCA were pentacosane, (Z)-12-pentacosene, 13-methylheptacosane and squalene; while for LDA, nonacosane, 11-methylnonacosane and pentacosane were the molecules that contributed most to the discrimination.