Effects of bee artificial diets on the composition of royal jelly: Chemical characterization and marker identification by untargeted and targeted metabolomic approaches

Non-migratory apiaries supply artificial diets to bees to sustain royal jelly (RJ) production during nectar shortages. To investigate the effects of artificial diets on RJ composition and identify characteristic markers for artificially-fed RJ, samples from various dietary combinations, including natural foods such as pollen and honey, along with alternative nutrients like soybean meal (SBM) and sucrose, were analyzed for physicochemical properties, major royal jelly proteins (MRJPs), and metabolic profiles. Our results demonstrate that sucrose feeding significantly increased sucrose, erlose, and δ¹ ³C values in RJ (p < 0.01), while SBM supplementation had a limited effect but notably reduced MRJP 4 and 5 levels (p < 0.05). Distinct metabolite profiles of amino acids, nucleotides, and isoflavones, were observed between RJ from natural and artificial diets, with daidzein (11.76–187.58 μg/kg) and genistein (16.06–292.25 μg/kg) identified as novel characteristic markers for SBM-fed RJ. The forms and concentrations of these isoflavone markers varied among SBM, SBM-fed RJ, and the midgut and hypopharyngeal glands of nurse bees, suggesting their transformation through biological processes of nurse bees. Our findings provide comprehensive insights into the authentication and quality assessment of RJ, addressing limitations inherent in conventional methodologies based on fundamental physicochemical parameters.