Sustainable 3D-printed immobilization system with soybean meal coating for high-yield Lactococcus lactis bioprocessing

This study utilized 3D-printed carriers coated with soybean meal, an agricultural by-product, to optimize the immobilization of Lactococcus lactis subsp. lactis , enhancing bacteriocin production and efficiency. Among the six experimental groups, the soybean-only (S) group achieved the highest yield of 1275.36 ± 17.49 IU/mL within two hours, which was 246 % higher than the non-coated group and significantly greater than the control (C) group (no 3D-printed carriers) (non-detect). After eight hours, the S group’s production rate was 2191.10 ± 21.24 IU·mL⁻¹ ·h⁻¹ , representing a 42.68 ± 2.20 % increase compared to the C group and 25.41 ± 1.80 % compared to the NC group. Under repeated batch fermentation, the S-only group achieved the highest total yield of 170,606.61 ± 3896.23 IU/mL over 10 cycles of shake flask fermentation, which was 20.42 % higher than static cultivation. Furthermore, after nine cycles, 72.2 % of activity was retained. The immobilized carrier also demonstrated improved hydrophilicity, with the water contact angle decreasing from 92.37° to 25.03°. SEM and FTIR analyses further confirmed the successful immobilization. Moreover, real-time RT-PCR analysis indicated that the increase in bacteriocin production was associated with the upregulation of 11 nisin-related genes, with nisA and nisB increasing by 4.74- and 4.86-fold, respectively. This study presents a novel approach for producing nisin by immobilizing Lactococcus lactis on 3D-printed structures coated with agricultural waste, demonstrating that immobilization can influence gene regulation. This technique offers a novel approach to valorizing agricultural waste while promoting environmentally responsible and economically viable industrial practices.