Natural bioink of interpenetrating network hydrogels mimicking extracellular polymeric substances for microbial...

Artificial biomanufacturing has been developed as a promising biotechnology for water pollution control. Effective bioimmobilization techniques are limited in application because of low productivity and the difficulty in achieving both mechanical strength and biocompatibility. Bioprinting technology, using biomaterials as bioink to enable the rapid on-demand production of bioactive structures, opens a new path for bioimmobilization. In this study, mimicking extracellular polysaccharide and protein of aerobic granular sludge (AGS), sodium alginate (SA) and silk fibroin methacryloyl (SilMA) were developed as the dual-component bioink with a suitable viscosity for bioprinting hydrogel. Interpenetrating network (IPN) hydrogel beads were manufactured using 1.5% (w/v) SA combined with 20% (w/v) SilMA through physical and covalent crosslinking, which exhibited excellent structural stability and bioactivity. The addition of SilMA provided a solution to the poor mechanical stability of SA-Ca hydrogels limited by Ca²⁺-Na⁺ ionic exchange. The unique structure of SilMA contributed to the reduction of hydrogel swelling as well as the prevention of SA loss. IPN hydrogels showed a swelling rate of less than 20% compared to the high swelling rate of more than 60% for SA hydrogels. On the other hand, SA controlled the hardening induced by excessive self-assembly of SilMA and improved mass transport in SilMA hydrogels. Compared to IPN hydrogels, SilMA hydrogels experienced a 15% volumetric shrinkage and exhibited a low water content of 92%. Sonication pretreatment of the dual-component bioink not only increased the intermolecular chain entanglement to form IPN, but also led to β-sheet content in SiMA reaching 46%-48%, which resulted in the formation of stable IPN hydrogels dominated entirely by physical crosslinking. Satisfactory proliferation and viability were achieved for the encapsulated bacteria in IPN hydrogels (μ_max 1.49-2.18 d^-1). Further, the IPN biohydrogels could maintain structural stability as well as achieve pollutant removal for treating synthetic wastewater with high Na⁺ concentration of 300 mg/L. The novel SA/SilMA hydrogel bioprinting strategy established in this study offers a new direction for bioimmobilization in water pollution control and other environmental applications.