## 4 Discussion  
Lignin peroxididases (EC 1.11.1.14) are heme-containing enzymes that catalyze hydrogen peroxide-dependent oxidation in lignin. Dyes are structurally complex like lignin. Thus, there is a high demand for lignin peroxidase in the textile, dye manufacturing, pulp and paper, and cosmetic industries that routinely use dyes in their processes [9]. The need for lignin peroxidase by these industries cannot be over-emphasized. The two lignin peroxidase preparations, separately obtained from _E. coli_ and _P. aeruginosa_ used in this study, exhibited appreciable dye decolorization potential of the synthetic dyes, Azure B (AZ) and Malachite Green (MG) (Table 1). Given that these preparations are crude enzymes, it is encouraging to note that they showed about 52-63% efficiency (the purified enzyme was 80% efficient). The inability of the lignin peroxidase or ligninolytic bacteria to effectively decolorize CR and RBBR (as observed in this study) has been reported by other researchers [3,9]. These bacterial isolates could not degrade these dyes because of the structural stability of CR and RBBR, which makes them highly resistant to biodegradation as well as their toxicity [9]. The characterization of an enzyme is crucial since factors such as substrate and enzyme concentrations, pH, and temperature play a vital role in influencing the enzymatic processes, and thus the industrial application of enzymes produced a 65 kDa lignin peroxidase from _Bacillus megaterium_[26,31]. In this study, the crude enzyme concentration at 50 mg/L effectively degraded the dyes (Figure 1). This was contrary to the work of [PERSON] and [PERSON] [32], in which the dyes were degraded at 200 mg/L. This implied that at 50 mg/L, there was an increase in the nucleic acids content ratio (RNA/DNA), which promote protein synthesis [33]. These results supported the concentration efficacy of the crude enzymes to degrade the dyes. A temperature either increases or badly inhibits enzyme activity. If temperature gets too high, enzyme activity will diminish, and enzymes are denatured [34]. Likewise, a change in pH can also alter the enzyme functional group, thus influencing the enzyme capability [35]. The lignin peroxididases produced in this study seem to prefer a neutral pH environment. These observations indicated that the enzymes could also efficiently neutralize weakly acidic dyes' waste [33]. In previous studies, _Citrobacter_ sp. CK3 achieved the best decolorization of reactive red at a neutral pH of 6-7. In contrast, _Aspergillus niger_ decolorized red azo dye at pH 9 [36]. The broad pH profiles (Figure 2) underscore the ability of the studied preparations to perform effectively in various industries. The mesophilic nature of the organisms reflects the optimum performance of the enzymes at 30-40 \({}^{\circ}\)C (Figure 3). Similar pH and temperature profiles have been reported for other lignin peroxididases [37]. It is noted that the enzyme preparations were inhibited by oxidoreductase inhibitors (Table 3). Lignin peroxidase, being a metalo-(heme)-enzyme, was inhibited by EDTA, a known metal-chelating agent [30,37,38]. Various lignin peroxididases have shown similar results [39,40,41]. The observed high survival rate of bacteria of dye treated with crude enzymes (Figures 6 and 7) is an indication of the efficacy of the crude enzyme to degrade the dye without toxic effects. This suggested the potential of the crude enzymes to simultaneously degrade the dyes and detoxify the dye's metabolite in producing no toxic products [42]. The calculated enzyme kinetics (Table 4) indicate that the enzyme produced by the _E. coli_ had a higher affinity (Km) and catalytic efficiency (\(k_{\text{cat}}/\text{K}_{\text{m}}\)) for the dyes than the enzyme of _P. aeruginosa_. It is apparent that the _E. coli_ enzyme was more effective on MG than on AZ. The measure of this (catalytic efficiency) is confirmed in the toxicity testing.