Aromatic secondary metabolite production from glycerol was enhanced by amino acid addition in Pichia pastoris

ABSTRACT: Aromatic secondary metabolites are widely used in various industries, including the nutraceutical, dietary supplement, and pharmaceutical industries. Their production currently relies on plant extraction. Microbe-based processes have recently attracted attention as sustainable alternatives to plant-based processes. We previously showed that the yeast Pichia pastoris (Komagataella phaffii) is an optimal host for producing aromatic secondary metabolites. Additionally, titers of resveratrol, an aromatic secondary metabolite, increased by 156 % when glycerol was used as a carbon source instead of glucose. However, the mechanisms by which glycerol resulted in higher production has remained unclear. In this study, we aimed to elucidate how P. pastoris produces higher levels of aromatic secondary metabolites from glycerol than from glucose. Titers of p-coumarate, naringenin, and resveratrol increased by 103 %, 118 %, and 157 %, respectively, in natural complex media containing glycerol compared with that in media containing glucose. However, the titers decreased in minimal synthetic medium without amino acids, indicating that P. pastoris cells used the amino acids only when glycerol was the carbon source. Fermentation with the addition of single amino acids showed that resveratrol titers from glycerol varied depending on the amino acid supplemented. In particular, addition of aspartate or tryptophan into the medium improved resveratrol titers by 146 % and 156 %, respectively. These results suggest that P. pastoris could produce high levels of aromatic secondary metabolites from glycerol with enhanced utilization of specific amino acids. This study provides a basis for achieving high-level production of aromatic secondary metabolites by P. pastoris. KEY POINTS: • P. pastoris can produce high levels of aromatic metabolites from glycerol • P. pastoris cells use amino acids only when glycerol is the carbon source • Aromatic metabolite titers from glycerol increase with amino acids utilization SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s00253-023-12798-5.