**3. Results and discussion**

#### **3.1 Actinobacterial growth**

Of all the six actinobaterial isolates examined, only three display the potential to grow on BPA. All the three actinobacterial isolates (*A. naeslundii, A. bovis*, and *A. israelii*) grew in the presence of BPA indicating their ability to metabolize BPA as a carbon and energy source. Likewise, the actinobacterial growth suggests that the necessary enzymes are required for the degradation of BPA; thus, the production of laccase in basal salt mineral medium confirmed the BPA degrading activity.

#### **3.2 Influence of consortium on enzyme production and BPA degradation**

#### *3.2.1 Effect of consortium on laccase production*

The influence of actinobacterial consortium on the production of laccase was studied over the degradation period of 312 h, as shown in **Figure 1**. The consortium of *A. naeslundii*, *A. bovis*, and *A. israelii* supported a maximum laccase yield of 15.9 U/mL followed by the consortium of *A. naeslundii* and *A. israelii* (9.5 U/mL), *A. naeslundii* and *A. bovis* (8.7 U/mL), while *A. bovis* and *A. israelii* (7.0 U/mL). Initially, at the incubation time of 24 h, the laccase activity observed for the three-member consortium had the lowest enzyme activity; this may be because the three individual strains are adjusting for one another to coexist in the culture media. Interestingly, at this period, the laccase activity increases steadily up to 168 h where maximal laccase

*A Laboratory-Scale Study: Biodegradation of Bisphenol A (BPA) by Different Actinobacterial… DOI: http://dx.doi.org/10.5772/intechopen.105546*

**Figure 1.**

*Laccase activity of the different consortia. Key: An+Ai = Actinomyces naeslundii and Actinomyces israelii; An+Ab = A. naeslundii and A. bovis; Ab+Ai = A. bovis and A. israelii; and An+Ab+Ai = A. naeslundii, A. bovis, and A. israelii.*

activity was recorded. This result suggested that laccase production increases proportionately with the growth of the actinobacterial consortium. This result is similar to the findings of Bogan and Lamar [38], who observed that extracellular enzymes of organisms are produced in response to their growth phases. Tsioulpas et al. reported that maximum laccase activity was measured in the growth medium, while 69–76% of phenolic compounds were removed by Pleurotus spp. [39]. This present work recognized that enzyme secretion also depends on the physical factor, nutritional, physiological, and biochemical nature of the microorganism.

#### *3.2.2 Effect of consortium on BPA degradation*

The influence of the actinobacterial consortium on BPA degradation was studied over the degradation period of 312 h, as shown in **Figure 2**. The consortium of *Actinomyces naeslundii, A. bovis*, and *Actinomyces israelii* supported a maximum BPA degradation of 93.1% followed by that of *A. naeslundii* and *A. israelii* (87.3%), *A. naeslundi* and *A. bovis* (80.4%), and *A. bovis* and *A. israelii* (76.0%). Microbial consortia have the capability of degrading a wide range of hydrocarbons. This research was able to link actinobacterial growth and laccase activity to the rate of BPA degradation because it can be deduced that at 24 h, all the two-member consortium growth gradually increases until 120 h when a steady decrease set in. However, a different growth pattern was observed for the three-member consortium where the growth steadily decreased at 24 h and then sharply increases at 48 h, and the growth increases until 168 h when there was a gradual decline to the incubation period of 312 h. The decrease in growth and laccase activity might be due to the depletion of nutrients or the production of waste or toxic substances into the culture media during this period. BPA degradation rate hardly increases after the 168-h incubation period. However, the enhanced BPA degradation performance displayed by the three-member actinobacterial consortium is due to synergic in the secretion of laccase compared to the two-member consortium. Although all the actinobacterial strains under study exhibited promising potentials to degrade BPA, their interactive or compatibility test was not investigated.

**Figure 2.** *Degradation of BPA by different consortia (a) An+Ai; (b) An+Ab; (c) Ab+Ai; and (d) An+Ab+Ai.*

### **3.3 Degradation by-product of BPA**

The mineral-salt-medium-supplemented BPA as a sole carbon source at pH 7.0 inoculated in each actinobacterial consortium was analyzed after an incubation period of 312 h. Cultures were extracted for GC–MS analysis to determine the biodegradation products of BPA. The by-products of BPA degradation were investigated, and each metabolite produced during the biodegradation process is shown in **Figure 3**. The GC–MS analysis showed that the metabolites could be identified as

**Figure 3.** *Chromatogram generated by GC–MS.*

*A Laboratory-Scale Study: Biodegradation of Bisphenol A (BPA) by Different Actinobacterial… DOI: http://dx.doi.org/10.5772/intechopen.105546*

concerned compounds by comparisons with known authentic compounds using the NIST Chemistry library. The mass peak is found at 38 for 1,2,4-trimethylbenzene, and its relative molecular mass is 120 at the retention time of 4.0 min, while the base peak value was observed at 105. In addition, 2,9-dimethyldecane was identified at mass peak of 21 at the retention time of 4.9 min. The relative molecular mass of the compound 2,9-dimethyldecane was observed as 170 and base peak was noticed at 43. Oxalic acid was also identified as one of the intermediate products, the relative molecular mass of the compound was 216 with a retention time of 7.0 min, and the mass and base peak values were recorded at 12 and 57, respectively. According to Kusvuran and Yildrim [40], oxalic acid was identified as organic intermediate from the degradation of BPA, which is similar to our observation in this study. However, intermediates such as p-hydroxyacetophone, hydroquinone, p-hydroxybenzaldehyde, and p-hydroxbenzoic identified by previous studies [41] were not detected.
