Aldolases are lyases enabling C-C bond formation and offering the benefit to work in eco-friendly conditions (water as solvent, neutral conditions, room temperature) without any need for protection and deprotection steps . Another advantage of these biocatalysts mainly relies on their capabilities to control the configuration of the newly created chiral centre(s). Using an ketoaldehyde as electrophilic substrate (hydroxypyruvaldehyde phosphate, HPP, as a proof of principle) with different aldolases, we were able to prepare various 2,5-dicarbonyl adducts. In particular, due to the high homology of HPP with glyceraldehyde-3-phosphate, deoxyribose-5-phosphate aldolase (DERA) natural substrate, the reactions involving this aldolase proceeded with good efficiency. Amazingly, a very interesting result was observed since exceptional multiple additions of the nucleophilic substrate acetaldehyde till fourfold was demonstrated, leading to a C11 aldoketose monosaccharide. Interestingly, the stereochemistries of the four asymmetric centers were controlled, starting from achiral material. HPP was generated using a rhamnulose-1-phosphate aldolase, allowing the implementation of a one pot two-steps cascade reactions (figure 1). The results obtained with the other aldolases will also be presented herein. Figure 1 : DERA catalyzed reaction using hydroxypyruvaldehyde phosphate as electrophile References: 1 P. Clapés, X. Garrabou (2011), Current Trends in Asymmetric Synthesis with Aldolases, Adv. Synth. Catal. : 13, 2263-2283. 2 V. Laurent, E. Darii, A. Aujon, M. Debacker, J.-L. Petit, V. Hélaine, T. Liptaj, M. Breza, A. Mariage, L. Nauton, M. Traïkia, M. Salanoubat, M. Lemaire, C. Guérard-Hélaine, V. de Berardinis (2018), Synthesis of Branched-Chain Sugars with a DHAP-Dependent Aldolase: Ketones are Electrophile Substrates of Rhamnulose-1-phosphate Aldolases, Angew. Chem. Int. Ed. : 19, 5467-5471.