Production of pyranoanthocyanins using Escherichia coli co-cultures.

Hydroxyphenyl-pyranoanthocyanins are one of the pyranoanthocyanins found in red wines and some fruit juices. Since they have a fourth ring (pyran or ring D) which provides higher color intensity and exceptional stability toward pH variations in comparison to their anthocyanin precursors, these molecules are one of the most important candidates as natural colorants especially for low- and medium-acidic food and beverages. However, their isolation and characterization are difficult due to their very low concentration. In this study, we co-cultured recombinant E. coli strains to synthesize pyranoanthocyanins with improved titers and yields. To accomplish this task, firstly we engineered 4-vinylphenol and 4-vinylcatechol producer modules then we co-cultured each one of these strains with cyanidin-3-O-glucoside producer recombinant cells to obtain pyranocyanidin-3-O-glucoside-phenol (cyanidin-3-O-glucoside with vinylphenol adduct) and pyranocyanidin-3-O-glucoside-catechol (cyanidin-3-O-glucoside with vinylcatechol adduct). By optimizing the co-culture conditions, we were able to significantly increase final titers and yields, allowing our co-culture approach to easily outperform production of pyranoanthocyanins from red wine. Finally, we demonstrate that the produced pyranoanthocyanins are far more stable than the starting plant-produced cyanidin 3-O-glucoside.