Immobilisation of bacteria onto magnetic nanoparticles for the decolorisation and degradation of azo dyes

Abstract

Azo dyes are widely used in industries and their release in the environment contributes to the pollution of effluents. The authors aim to develop a new eco-friendly water treatment method for the degradation of azo dyes based on in situ magnetic separation and immobilisation of bacterial cells. The immobilisation was achieved using superparamagnetic Fe₃O₄ nanoparticles and offers the possibility of reusing bacteria by magnetic separation for several degradation cycles. The iron-oxide nanoparticles were synthesised by reverse co-precipitation. The Gram-positive bacteria Bacillus subtilis were immobilised using iron-oxide nanoparticles by adsorption and then separated with an external magnetic field. Transmission electron microscopy observation showed that the particles' diameter was ∼20 nm with a narrow size distribution. Moreover, the iron-oxide nanoparticles were adsorbed onto the surface in order to coat the cells. B. subtilis has proved its ability to decolorise and degrade several azo dyes at different values of pH, with the highest decolorisation rate for Congo red. Furthermore, immobilised cells have a degradation activity similar to that of free cells. The system provided a degradation rate up to 80% and could be reused for seven batch cycles.

Fig. 1

Azo dyes degradation process using B. subtilis immobilised by iron–oxide nanoparticles

Fig. 2

XRD patterns of magnetic nanoparticles obtained by reverse co‐precipitation

Fig. 3

TEM images of iron–oxide nanoparticles synthesised by reverse co‐precipitation

Fig. 4

Infrared spectrum of magnetic nanoparticles obtained by reverse co‐precipitation

Fig. 5

TEM Images of iron–oxide nanoparticles coating B. subtilis (a) 4 g/L, (b) 40 g/L, (c) Free cell

Fig. 6

SEM images of immobilised bacteria incubated with different concentrations of magnetic iron–oxide nanoparticles (a) Free cells, (b) 4 g/L, (c) 10 g/L, (d) 40 g/L

Fig. 7

Enumeration results of immobilised bacteria by iron–oxide nanoparticles Data points indicate the mean of three independent replicates; ±standard errors of mean (SEM) is indicated by error bars

Fig. 8

Decolorisation test of the azo dyes using free B. subtilis ATCC6633 (a) T ₀, (b) After 24 h of incubation at 37°C, (c) Results of decolorisation rate measured by UV‐Vis spectroscopy Data points indicate the mean of three independent replicates; ±SEM is indicated by error bars

Fig. 9

Amino acid sequences of B. subtilis ATCC6633 azoreductase 1

Fig. 10

Optical density of decolorisation solutions at 630 nm after 24 h of incubation Data points indicate the mean of three independent replicates; ±SEM is indicated by error bars

Fig. 11

Biosorption test and Extracellular decolorisation test on CR (a) Before, (b) After decolorisation and centrifugation, (c) Decolorisation of CR using extracellular supernatant of B. subtilis Data points indicate the mean of three independent replicates; ±SEM is indicated by error bars

Fig. 12

GC/MS Chromatogram of CR metabolite after decolorisation by B. subtilis

Fig. 13

Proposed biodecolorisation pathway of CR dye by B. subtilis

Fig. 14

Reusability test of immobilised B. subtilis in decolorisation process of CR