Aerobic culture of Propionibacterium freudenreichii ET-3 can increase production ratio of 1,4-dihydroxy-2-naphthoic acid to menaquinone

This is the first report on the production of both 1,4-dihydroxy-2-naphthoic acid (DHNA) and menaquinone by Propionibacterium freudenreichii ET-3. DHNA can be a stimulator of bifidogenic growth, and menaquinone has important roles in blood coagulation and bone metabolism. During anaerobic culture, DHNA and menaquinone concentrations reached 0.18 mM and 0.12 mM, respectively. The molar ratio between these products was approximately 3:2, which was not affected by culture pH and temperature over the ranges of 6.0–7.0 and 31–35°C, respectively. As for organic acid, propionate and acetate accumulated at concentrations of 0.3 M and 0.15 M, respectively, and the propionate accumulation particularly inhibited further production of DHNA. To improve DHNA production, we switched from anaerobic condition to aerobic condition during the culture when lactose was depleted. DHNA concentration continued to increase even after lactose exhaustion, reaching 0.24 mM. In contrast to DHNA production, menaquinone production stopped after the switch to aerobic condition. The total molar production of DHNA and menaquinone was 0.3 mM irrespective of aerobic culture and anaerobic-aerobic switching culture. Therefore, the anaerobic-aerobic switching culture could increase the production ratio of DHNA to menaquinone. The DHNA concentration obtained from the anaerobic-aerobic switching culture was 1.3-fold higher than that in the anaerobic culture, because P. freudenreichii ET-3 utilized propionate accumulated in the medium via the reversed methylmalonyl CoA pathway under aerobic condition. The culture method proposed in this study could be applicable to industrial-scale fermentation using 1000 l of media, by which 0.23 mM DHNA was produced.