Sustainable Overproduction of the Bacterial Pigment Indigoidine as a Natural Colorant: Stability Evaluation, Inhibition of Obesity-Diabetes-Related Digestive Enzymes, and Activation of AMPK-AKT.

The increased focus on bacterial pigments can be attributed to their greater safety and eco-friendliness when compared to artificial food colorants, which can lead to severe health problems. This research was designed to optimize IND production at varying growth conditions, to assess its chemical stability, and to study its possible biological activities associated with diabetes, obesity, and insulin resistance. Medium types played an important role in both BL21-DE3 cell growth and IND production. The BL21-DE3 cell growth was maximum in PGB medium (2.76 g/L/24 h), then in LB-O medium (1.29 g/L/24 h), LB medium (1.14 g/L/24 h), and NB medium (0.94 g/L/24 h). The IND production was 75.8, 49.8, and 38.9 mg/L/24 h in LB, NB, and PGB media, respectively, while the experimental maximum IND production was achieved in LB-O medium (177.4 mg/L/24 h), which closely resembled the theoretical IND production of 180 mg/L/24 h. IND exhibited high cellular absorption properties, low cytotoxicity, and dose-dependent inhibition of α-amylase and lipase enzymes at IC50 values of 0.69 and 0.37 µg/mL, respectively. Furthermore, IND enhanced 5'-adenosine monophosphate-activated protein kinase (AMPK) and protein kinase B (AKT) signaling pathways at EC50 values of 1.36 and 4.59 µg/mL, respectively. The binding affinity of IND was also observed towards α-amylase, lipase, AMPK, and AKT enzymes via molecular docking with binding energies (BE) of -7.4, -8.1, -6.9, and -5.5 kcal/mol, respectively. All these findings were confirmed through human spermatozoa culture, where IND stimulated glucose metabolism and AMPK and AKT signaling pathways.