Extremophilic microorganisms often produce novel bioactive compounds to survive under harsh environmental conditions. Exopolysaccharides (EPSs), a constitutive part of bacterial biofilm, are functional biopolymers that act as a protecting sheath to the extremophilic bacteria and are of high industrial value. In this study, we elucidate a new EPS produced by thermophilic Bacillus haynesii CamB6 from a slightly acidic (pH 5.82) Campanario hot spring (56.4 °C) located in the Central Andean Mountains of Chile. Physicochemical properties of the EPS were characterized by different techniques: Scanning electron microscopy- energy dispersive X-ray spectroscopy (SEM-EDS), Atomic Force Microscopy (AFM), High-Performance Liquid Chromatography (HPLC), Gel permeation chromatography (GPC), Fourier Transform Infrared Spectroscopy (FTIR), 1D and 2D Nuclear Magnetic Resonance (NMR), and Thermogravimetric analysis (TGA). The EPS demonstrated amorphous surface roughness composed of evenly distributed macromolecular lumps. GPC and HPLC analysis showed that the EPS is a low molecular weight heteropolymer composed of mannose (66%), glucose (20%), and galactose (14%). FTIR analysis demonstrated the polysaccharide nature (–OH groups, Acetyl groups, and pyranosic ring structure) and the presence of different glycosidic linkages among sugar residues, which was further confirmed by NMR spectroscopic analyses. Moreover, D-mannose α-(1→2) and α-(1→4) linkages prevail in the CamB6 EPS structure. TGA revealed the high thermal stability (240 °C) of the polysaccharide. The functional properties of the EPS were evaluated for food industry applications, specifically as an antioxidant and for its emulsification, water-holding (WHC), oil-holding (OHC), and flocculation capacities. The results suggest that the study EPS can be a useful additive for the food-processing industry.

Prodigiosin is a ‘natural red pigment produced by Serratia marcescens which exhibits immunosuppressive and anticancer properties in addition to antimicrobial activities. This work presents an attempt to maximize the production of prodigiosin by two different strategies: one factor at time (OFAT) and statistical optimization. The result of OFAT revealed that sucrose and peptone were the best carbon and nitrogen sources for pigment production with concentration of prodigiosin of about 135 mg/ L. This value was increased to 331.6mg/ L with an optimized ratio of C/N (60:40) and reached 356.8 with pH 6 and 2% inoculum size at end of classical optimization. Statistical experimental design based on Response surface methodology was conducted to optimize the composition of trace element. The design revealed that the predicted prodigiosin production of 406 mg/L can be achieved when concentrations of trace element CaCl2·2H2O, FeSO4·4H2O, MgSO4·7H2O and MnSO4·4H2O were equal to 9.22, 0.32, 0.67 and 2.48 g/L, respectively. The actual production of prodigiosin in the optimized medium was 375 mg/L. Growth kinetics of S. marcescens were evaluated in optimized medium which revealed that prodigiosin was ‘non-associated growth’ secondary metabolite with maximum production of approximately 365.7 mg/L obtained after 54 hours of incubation.

Prodigiosin, a red pigment with diverse biotechnological applications, is produced as a secondary metabolite by Gram-negative bacilli Serratia marcescens. In this study, we implemented an AI-guided hybrid optimization framework combining Response Surface Methodology (RSM) using a Circumscribed Central Composite Design (CCCD) and Artificial Neural Network (ANN) modeling to enhance prodigiosin pigment production. Across 34 experimental runs, we optimized sucrose and peptone concentrations along with inoculum size. The RSM-derived model exhibited a strong correlation (R² = 0.953), while the ANN, trained using a backpropagation algorithm, demonstrated superior predictive power (R² = 0.998; MSE = 0.000414), underscoring the potential of artificial intelligence in modeling complex bioprocesses. Beyond statistical optimization, an induction strategy using 1% of various natural additives (vegetable oils and egg components) identified egg white, rich in albumin, as the most effective enhancer, tripling prodigiosin yield. Further investigation revealed that a 2% egg white concentration maximized production to 1070 mg L⁻¹, a substantial increase compared to the optimized yield of 359.2 ± 12 mg L⁻¹ and predicted value of 391.86 mg L⁻¹. These results highlight the value of integrating machine learning with experimental design and protein-rich inducers to strengthen sustainable microbial pigment production in a cost-effective and scalable manner.

Synthetic azo dyes are the largest class of industrial colorants having widespread application in textile, food, cosmetic, and pharmaceutical industries. Moreover, they are persistent and toxic, threatening aquatic environments as well as human health. Disperse red 1 (DR1), a mono-azo dye belonging to the disperse dye group and widely used in polyester dyeing, cosmetics, and other applications, is of particular concern due to its mutagenic potential and resistance to conventional treatment processes. The present study investigated an integrated DR1 removal strategy using thermotolerant Bacillus licheniformis Tol1 as well as its EPS-stabilized biogenic silver nanoparticles (AgNPs). With a maximum tolerable concentration of 0.5 g L⁻¹, B. licheniformis Tol1 showed a maximum decolorization of 70.86% (0.2 g L⁻¹, 55 °C). However, response surface methodology (RSM) based on the Box–Behnken design showed an actual decolorization efficiency of 73.13%. The artificial neural network (ANN) model predicted an accuracy of R² = 0.9933, confirming the robustness and reliability of the experimental findings. To enhance dye removal efficiency, Tol1 EPS-stabilized AgNPs were synthesized via a green method and characterized using UV-Vis, SEM-EDAX, TEM, AFM, FTIR, DLS and zeta potential. Characterization of AgNP confirmed the formation of spherical stable AgNPs with an average size of 19.99 ± 0.38 nm, indicating polydisperse colloids nature with moderate electrostatic stability. A sunlight/H₂O₂-assisted process (photocatalytic experiments) demonstrated DR1 decolorization (80.72 ± 1.72% within 5 h under sunlight) following pseudo-first-order kinetics (k = 0.271 h⁻¹). Furthermore, FTIR analysis confirmed the degradation of the chemical structure of DR1 through the disappearance of the characteristic azo (–N=N–) bond, indicating cleavage of the dye molecule. Overall, the present study provides a dual biological–nanotechnological approach for DR1 decolorization using single bacteria as well as its polysaccharide-stabilized AgNP, a sustainable eco-friendly future approach. However, further studies on complete mineralization, transformation products, toxicity evaluation, detailed catalyst reusability, and silver (Ag) leaching are needed to facilitate the practical implementation for wastewater treatment.