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The authenticity of honey is generally a worldwide concern, and there is a pressing need to establish a suitable entomological method to identify the authenticity of Apis cerana cerana (A. cerana) and Apis mellifera ligustica (A. mellifera) honey. Hydroxy fatty acids as bee-derived components are known to widely exist in honey and other biosamples. Herein, we present an identification strategy for hydroxy fatty acids based on the relative quantification with reference to royal jelly and targeted quantification combined with multivariate statistical analysis to identify the honey entomological origin. Multivariate statistical analysis was used to further determine differential hydroxy fatty acids between A. cerana honey and A. mellifera honey. Results showed that 8-hydroxyoctanoic acid (96.20-253.34 versus 0-32.46 mg kg-1) and 3,10-dihydroxydecanoic acid (1.96-6.56 versus 0-0.35 mg kg-1) could be used as markers for accurate identification of the honey entomological origin, while the three fraud honey samples were recognized using this method. This study provides the novel marker hydroxy fatty acids to identify A. cerana honey and A. mellifera honey from the perspective of bee-derived component differences.