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Three Metschnikowia strains marketed as bioprotection yeasts were studied to compare their antimicrobial effect on a mixture of two Hanseniaspora yeast strains in synthetic must at 12 °C, mimicking pre-fermentative maceration by combining different approaches. The growth of the different strains was monitored, their nitrogen and oxygen requirements were characterised, and their metabolomic footprint in single and co-cultures studied. Only the M. fructicola strain and one M. pulcherrima strains colonised the must and induced the rapid decline of Hanseniaspora. The efficiency of these two strains followed different inhibition kinetics. Furthermore, the initial ratio between Metschnikowia and Hanseniaspora was an important factor to ensure optimal bioprotection. Nutrient consumption kinetics showed that apiculate yeasts competed with Metschnikowia strains for nutrient accessibility. However, this competition did not explain the observed bioprotective effect, because of the considerable nitrogen content remaining on the single and co-cultures. The antagonistic effect of Metschnikowia on Hanseniaspora probably implied another form of amensalism. For the first time, metabolomic analyses of the interaction in a bioprotection context were performed after the pre-fermentative maceration step. A specific footprint of the interaction was observed, showing the strong impact of the interaction on the metabolic modulation of the yeasts, especially on the nitrogen and vitamin pathways.