Bioremediation of Phenol by SP-1 Immobilized with Sodium Alginate/Chitosan@Biochar Microspheres

CHEN Lutong; FAN Chenchen; JIANG Chen; HOU Dongmei; ZOU Jianping

doi:10.20078/j.eep.20250701

Abstract：Immobilized microbial technology offers advantages such as strong resistance to toxicity,improved stability,and superior degradation performance.This study developed an innovative embedding-crosslinking co-immobilization strategy using sodium alginate(SA),chitosan(CS),and biochar(BC)to construct SA/CS@BC composite microspheres for encapsulating the deep-sea phenol-degrading bacterial consortium SP-1.Key parameters were optimized through single-factor experiments:SA concentration(1.0％,2.0％,3.0％,4.0％,5.0％),CS concentration(0.25％,0.5％,0.75％,1.0％,1.25％),CaCl2 crosslinker concentration(1.0％,2.0％,3.0％,4.0％,5.0％),and BC dosage(0.25％,0.5％,0.75％,1.0％,1.25％).Following degradation screening at 800 mg/L phenol,the performances of free bacteria,SA/CS microspheres,and SA/CS@BC microspheres were comparatively analyzed at phenol concentrations of 200,400,600,800,1 000,and 1 200 mg/L.The underlying mechanisms were investigated using SEM for microstructural morphology,FTIR for functional group analysis,the BET method for specific surface area measurement,LC-MS for metabolic intermediate identification,TOC analysis for mineralization quantification,and 5-cycle reuse tests for operational stability assessment.The optimized SA/CS@BC microspheres at 3.0％SA,0.75％CS,4.0％CaCl2,and 1.00％BC achieved a 94.6％degradation rate at 1 200 mg/L phenol,which was 3.3-fold higher than that of free bacteria and 76.2％higher than that of SA/CS microspheres.At 1 000 mg/L,SA/CS@BC maintained 95.2％efficiency,significantly exceeding that of SA/CS microspheres and free bacteria.BC incorporation increased the specific surface area from 4.936 m2/g to 32.829 m2/g;SEM confirmed the dense colonization of SP-1 within the BC porous architecture,whereas SA/CS microspheres exhibited a compact structure with sparse microbial loading.FTIR spectra revealed intensified absorption peaks at 3 335 cm-1(-OH),1 590 cm-1(—COOH),and 1 004 cm-1(C—O-C)for SA/CS@BC,with blue shifts compared to SA/CS(3 420 cm 1,1 622 cm 1,1 024 cm 1),indicating enhanced hydrogen-bonding networks.Elevated functional group abundance and strengthened hydrogen bonds jointly promoted phenol adsorption.LC-MS detected key intermediates including catechol,muconic acid,and succinic acid.Phenol was first hydroxylated to catechol,followed by ortho-cleavage of catechol to form muconic acid,which was further oxidized to succinic acid entering the tricarboxylic acid cycle,and ultimately mineralized to CO2 and H2O.TOC analysis demonstrated 93.9％removal after 9 days treatment at 1 200 mg/L phenol,with the IC/TC ratio increasing from 5.8％to 68.5％,verifying complete mineralization to CO2 and H2O.SA/CS@BC microspheres retained 90.5％degradation efficiency after 5 reuse cycles at 1 200 mg/L phenol.This work demonstrates that SA/CS@BC microspheres enhance phenol degradation:the SA/CS hydrogel shields microbes from acute phenol toxicity;BC rapidly concentrates phenol,while SA/CS gel controls the gradual release to maintain sub-inhibitory concentrations.BC's pores increase microbial loading density,and surface oxygen groups facilitate microbe-pollutant interactions.This technology provides an efficient,stable,and engineerable solution for bioaugmentation of high-concentration phenol-laden wastewater.