**3. Diacetyl and acetoin production**

Diacetyl and acetoin are important compounds of buttery flavor in fermented foods and are used as additives in the food industry. Both compounds are derived from pyruvate, which is converted to -acetolactate by the action of -acetolactate synthase or acetohydroxyacid synthase. Then, acetoin is formed by the activity of -acetolactate decarboxylase on acetolactate and diacetyl results from a non-enzymatic oxidative decarboxilation of acetolactate (Figure 2). Most metabolic engineering approaches to produce diacetyl/acetoin by fermentation have been developed in the model LAB *L. lactis*, in which strains that divert an important part of pyruvate flux towards the production of -acetolactate have been constructed (Hugenholtz *et al.*, 2000; Lopez de Felipe *et al.*, 1998). *ilvBN* genes, encoding acetohydroxyacid synthase from *L. lactis*, have been expressed from the lactose operon in *L. casei*, an organism which shows marginal production of diacetyl/acetoin, resulting in increased diacetyl formation (Gosalbes *et al.*, 2000). In addition, to enhance diacetyl/acetoin production, the amount of pyruvate available for IlvBN was increased by blocking pyruvate alternative pathways in *L. casei*. Thus, the *L. casei* strain that expresses the *ilvBN* genes was mutated in the *ldh* gene and in *pdhC*, encoding the E2 subunit of the pyruvate dehydrogenase enzyme. The introduction of these mutations resulted in an increased capacity to synthesize diacetyl/acetoin from lactose fermentation in whey permeate (1400 mg/l at pH 5.5) (Nadal *et al.*, 2009).
