*6.3.2. Solid phase extraction method*

Despite the efficiency of liquid‐liquid extraction method of removal of chemicals from waste‐ water, the technique comparatively consumes a lot of time and is expensive with possible associated injuries from the large quantity of organic solvents (some toxic) used in the pro‐ cess. Solid phase extraction technique, which requires minimal time and organic solvents, highly selective and environmentally friendly, is therefore regarded as an appropriate alter‐ native for liquid‐liquid extraction method [5].

Solid phase extraction system consists of a syringe containing a merged silica fibre, which is coated with an immobilised phase. The aqueous solution containing the analyte is exposed to the fibre with the subsequent accumulation of the analyte on the stationary phase. The fibre is then removed from the aqueous solution followed by desorption of the extracted analyte in a column injector or gas chromatography. Polydimethylsiloxane is normally used as the stationary phase for removal of halogenated and polycyclic aromatic hydrocarbons and poly‐ chlorinated biphenyls [60].

Möder et al. [61] used a polyacrylate‐coated fibre as a solid phase extractant for phenolic com‐ pound elimination from wastewater. Effects of humic acid and surfactant concentrations on the extraction efficiency were analysed. They attributed the successful extraction of naphthols, alkylated phenols and Tetra‐ols from the wastewater to the fact that the polyacrylate coating demonstrated high specificity for polar hydroxylated aromatic compounds. Non‐polar mole‐ cules hardly interacted with the extractant within the optimum 45 min extraction time. Another experiment on solid‐phase extraction of phenols from wastewater was carried out by Tavallali and Shiri [62]. Their study involved the use of iron oxide nanoparticles modified with activated carbon as the solid adsorbent. They demonstrated that development of solid phase extraction method based on magnetised activated carbon prior to their spectrophotometric determination is an appropriate technique. Their result showed 98% removal of phenol from water, indicat‐ ing the effectiveness of the iron oxide nanoparticle modified activated carbon solid adsorbent.

### **6.4. Biological method**

Biological method of phenolic compound removal from wastewater is subdivided into micro‐ bial and enzymatic methods. The microbial method involves the deployment of bacteria, yeast and fungi in breaking down the phenolics into harmless products such as carbon dioxide and water. This method of phenolics removal is feasible as a result of the fact that some microorgan‐ isms are known to depend on aromatic compounds, including phenolics, as their source of car‐ bon or nutrient [63, 64]. It has the advantage of a comparatively low operational cost. Microbial removal of phenolic compounds occurs through either aerobic or anaerobic processes and begins with hydroxylation (introduction of hydroxyl groups) of the aromatic ring [65]. Hydroxylation through aerobic degradation involves two steps with catechol being the end product [66]:


Cleavage of the catechol aromatic rings then passes through various stages with specific enzymes, based on the type of microorganisms, resulting in the conversion of the phenolic compounds to compounds such as carbon dioxide and water [67]. Anaerobic degradation occurs whenever oxidising agents such as sulphates, nitrates and CO2 , or light are present. This process is believed to be initiated by carboxylation of phenol to 4‐hydroxybenzoate [68].

In general, the aerobic process is known to be better suited for the degradation of phenolics with minimal substituents consisting of halogens. On the other hand, the anaerobic process is mostly appropriate for reduction of chlorinated phenolic compounds [69]. The anaerobic system produces methane in addition to carbon dioxide and water. A major advantage of the anaerobic system of degradation is the absence of aeration cost, recovery of methane and minimum excess biomass generation [70].

Kukadiya et al. [71] studied the effectiveness of using a moving bed biofilm reactor for phe‐ nolic compounds removal from wastewater. The laboratory scale model moving bed biofilm reactor was observed to be effective against the removal of phenol with about 98% efficiency. In their experiment, Sinha et al. [72] studied the p‐chlorophenol and phenol microbial deg‐ radation as a single and mixed substrates by using *Rhodococcus* sp. RSP8 bacteria strain. The experiment was performed with a liquid mineral salt medium in a shake flask experiment at a neutral pH and a temperature of 37 °C. The two compounds (p‐chlorophenol and phenol) served as the main source of carbon and energy for the cells and were consumed completely as individual solutions by the cells. The two pollutants, however, repressed each other's deg‐ radation by the cells in the mixed substrate experiment.

The enzymatic method of degradation, however, employs enzymes (biological catalysts). Enzymes can be used effectively to selectively eliminate pollutants in water since they catalyse specific reactions under modest temperature, pH and ionic strengths [73]. In addi‐ tion, the enzymatic reaction is known to occur at much faster rates compared to other types of reactions [74]. As an advantage over the microbial system of pollutant degradation, the enzymatic system of pollutant removal can occur under conditions, which are unfavourable or toxic to bacteria. This system can operate under different pollutant concentration (high or low), eliminates the time requirement for biomass acclimatisation, involves no shock loading effect and with no generated biomass [74]. This method receives a high level of consideration due to its high pollutant removal efficiency, operation in wide temperature and pressure ranges and formation of harmless end products [75, 76]. The enzyme with a high promise for dephenolisation of phenolics in water is tyrosinase (KF1.14.18.1). This enzyme oxidises the phenolics to quinones, which are further broken down into the non‐toxic intermediate prod‐ uct. The intermediate products are then removed through the addition of binding agents [77].

There has been a series of reported research works where enzymes have been used for the removal of phenolic compounds from wastewater [78, 79]. Among these reports is the work done by Shesterenko and co‐workers [79]. They used tyrosinase isolated from *Agaricus bisporus* and immobilised it on polymer carriers, and inorganic coagulants to remove phenols from water. Peroxidase extracted from horseradish, hydrogen peroxide and polyethylene glycol (PEG) was also used to catalyse phenol removal from simulated wastewater [80]. Optimum degradation of 1 mM phenol (80%) was attained at 0.3 U/ml horseradish peroxidase and 3.0 mM hydrogen peroxide concentrations at pH 7 and 273 mg/l of PEG.
