Abstract
Microcystin-LR (MC-LR) treatment benefits from rapid parent removal and control of transformation products. We co-immobilized cell-free MlrA and MlrC preparations on chitosan- and glutaraldehyde-functionalized magnetic biochar to form a dual-enzyme magnetic biochar composite (DMBC). At 1 mg·L−1 MC-LR, measured removal reached 82.85 ± 0.26% within 30 min and 95.28 ± 0.34% within 1 h. No difference in 30 min removal was detected between DMBC and the MlrA-only composite (80.00 ± 5.66%). MlrC-only and induced empty-vector BL21 lysate composites gave similar lower responses, indicating a substantial carrier-associated contribution. Linear MC-LR and 3-amino-9-methoxy-2,6,8-trimethyl-10-phenyldeca-4(E),6(E)-dienoic acid (Adda) were identified in recovered DMBC extracts. Linear MC-LR was also detected qualitatively in recovered MlrA-only material. Together, these observations are consistent with the established MlrA/MlrC cleavage sequence. A separate Adda experiment showed a 97.02 ± 4.22% decrease in normalized aqueous chromatographic response at 30 min. Six weekly reuse cycles maintained mean MC-LR removal between 74.60% and 81.50%. Treatment of centrifuged urban lake water showed concentration-dependent removal at 60 min. The results support a reusable cell-free treatment material that combines MlrA-associated transformation and carrier-associated removal, with product evidence consistent with downstream cleavage.