**5. The potential of microalgae in absorbing heavy metals**

Microalgae are considered microscopic photosynthetic organisms that are found in all aquatic environments (freshwater and saltwater) and their cultivation is possible in closed and open environments. The size of microalgae can be from several micrometers to several hundred micrometers. Criteria and various methods are used to classify microalgae, including their pigment, life cycle, or their primary cell structure. Considering their abundance, the most important types of microalgae are diatoms (Bacillariophyceae), green microalgae (Chlorophyceae), and golden microalgae (Chrysophyceae). The difference between these types of seaweed is mainly in the structure of the cell walls, where the absorption of heavy metal ions occurs. The cell wall of microalgae generally contains significant amounts of starch and glycogen, as well as cellulose, hemicellulose, and polysaccharides. These compounds contain numerous reactive active groups (e.g., amino, hydroxyl, carboxyl, sulfate, etc.) that can It is involved in chemical bonding with metal ions and are known as the main factor of very good biological absorption potential of microalgae [17–19].

Investigations have shown that microalgae are at the forefront of wastewater treatment because, in addition to the biological absorption of heavy elements, they also do nitrogen removal, phosphorus removal, and COD reduction well [20–22].

Dirbaz and Rosta conducted a study on the kinetics and thermodynamics of cadmium biosorption by Parachlorella microalgae. They observed that the absorption capacity of this microalgae at a temperature of 30 degrees Celsius and a pH of 7 is 90.72 mg/g, which is between 3 and 5.5 times that of other studied absorbents [23].

PhongVo et al. presented a review of different designs and applications of microalgae-based photobioreactors for pollutant treatment. In their review article, in addition to summarizing the progress made in the field of removing pollutants with the help of microalgae, they provided a vision of the future of using photobioreactors in this field [24].

Moreira et al. investigated the biological removal of copper metal using the microalgae *Chlorella pyrenoidosa* with experiments designed by the factorial Box–Behnken method. They reported the removal of 83.14% under optimal conditions of pH 3.6, metal ion concentration of 5 mg/L, and adsorbent dose of 1.28 g/L [25].

Saavedra et al. conducted a comparative study on the removal of toxic elements arsenic, boron, copper, manganese, and zinc from solutions containing single metal ions and mixed metal ions by four different species of green microalgae [26].

Their results, in addition to confirming microalgae as efficient and economic adsorbents for removing heavy metals, showed that the presence of other metal ions strongly affects the removal rate of metal ions by microalgae.

Areco et al. studied the effect of zinc metal ions on the growth and photosynthetic metabolism of microalgae *Botryococcus braunii* and the ability of this microalgae to remove metal ions from aqueous solutions. They studied concentrations of 0 to 80 mg/L of metal ions and observed that increasing the concentration of zinc metal ions in the solution significantly reduces the growth of microalgae. Also, the metal absorption capacity in the period of 200 days was 3.4 grams per gram of absorbent [27].

In another study, Pradhan et al. studied the removal of hexavalent chromium using the Scenedesmus microalgae. They investigated the factors affecting this process, including initial pH, contact time, initial metal ion concentration, adsorbent dose, particle size, and temperature, and reported the effective removal of hexavalent chromium with a maximum of 92.89%. They also found the presence of aldehyde, amide, carboxylic acid, phosphate, and halide functional groups to be effective in this process by examining the FTIR spectra taken from the microalgae used. Regarding the adsorption mechanism, they concluded that the removal is done by anionic surface adsorption [28].

### **5.1 The mechanism of removing heavy metals by microalgae**

Adsorption processes are usually very complex, and the mechanism of metal adsorption includes a combination of various elementary mechanisms such as electrostatic collisions, ion exchange, complex formation, adsorption with chelate formation, micro-deposition, etc., which occur simultaneously or sequentially. The basic mechanism of the biological absorption process can be divided into two categories: chemical biological absorption and physical biological absorption. As their names suggest, the first category includes chemical reactions and the second category involves the absorption of metal ions by van der Waals forces or electrostatic attraction forces. Ion exchange, complex formation, and microscopic sedimentation are the main mechanisms of heavy metal absorption by microalgae [29].

The primary interactions in the ion exchange mechanism can be from electrostatic or van der Waals forces to chemical bonds (ionic or covalent). In general, microalgae have mobile metal ions in their structure such as K+, Na+, Ca2+, Mg2+, etc. attached to the functional groups of microalgae. In the biosorption process, these mobile metal ions are exchanged with heavy metal ions according to the following reaction:

$$R^- \text{--} X^\* \text{+} M^\* \leftrightarrow R^- \text{--} M^\* \text{+} X^\* \tag{2}$$

where R- is the functional group of the microalgae surface, X+ is the mobile metal ion and M+ is the heavy metal ion in the aqueous solution. The mechanism of complex formation includes the formation of a complex on the cell surface, between heavy metal ions in the solution and a functional group of microalgae. For example, it has been shown in research that the absorption of Cu(II) ions on *Chlorella vulgaris* is done by a complex formation mechanism in which dative bonds are formed between metal ions and the amino and carboxyl groups of the microalgae cell wall polysaccharide. Microscopic sedimentation occurs when the pH of the biosorption solution increases sharply and/or the concentration of metal ions in the aqueous solution increases to the saturation level. In this case, heavy metals in an aqueous solution can be precipitated and the resulting microscopic sediments settle on the surface of the bioabsorbent [30].

### **5.2 Effective factors on the biological absorption of heavy metals by microalgae**

In the case of microalgae, the most important factors affecting their biological performance in the process of removing heavy metals can be divided into two categories:


Growing conditions can affect the biological performance of microalgae. Although the data reported in the articles show that microalgae grown in saline environments contain higher amounts of polysaccharides than freshwater microalgae, their efficiency in the biological absorption process varies widely.

It was also shown that microalgae that have a large number of functional groups available on their surface show better biosorption characteristics. Of course, this depends on the nature of the microalgae and the pretreatment of the biomass cells before being used as a biosorbent. Normally, to obtain raw bioabsorbent, microalgae biomass is separated by centrifugation at different speeds and at different time intervals. This biomass is then pretreated. In most cases, pretreatment involves drying the biomass so that it can be stored more easily and for a longer period of time. The main process factors that affect the biological performance of microalgae and should be optimized for the discontinuous system are solution pH, adsorbent dose, contact time, and temperature. These factors for the continuous process are the pH of the solution, height of the bioabsorbent bed, flow rate of wastewater containing heavy metals, and initial concentration of heavy metal ions. A summary of the optimal process conditions for the removal of heavy metals from wastewater with microalgae is presented in **Table 2**.

The pH of the solution is one of the most important experimental parameters that not only affects the characteristics and solubility of heavy metal ions but also the degree of separation and dissociation of functional groups that are considered adsorption sites (such as hydroxyl, carboxyl, carbonyl, amino, etc.) from the bio absorbent surfaces [31].

According to research and review articles, the highest absorption rate of algae occurs at pH between 3 and 5 (because the acidity of the environment affects the surface bands of ions and biomass and the chemical structure of ions), and the dried biomass of algae has a much higher capacity and in the first 120 minutes, most of the absorption process will take place.

Biosorbent dosage is another parameter that should be optimized in order to ensure the economic and environmental efficiency of the bioremediation process. Using large amounts of bioabsorbent not only increases the cost of the bioabsorption process but also leaves a large amount of waste contaminated with heavy metals, which has a negative impact on the environment. On the other hand, the use of very small amounts of microalgae will significantly affect the efficiency of biological absorption, and the biological treatment process with low efficiency will not be used for industrial applications.

Contact time also plays an important role in ensuring the efficiency of the biological absorption process. The inappropriate value of this parameter can significantly limit the practical and industrial use of a biological adsorption process, even if its efficiency in removing heavy metal ions is high. The absorption rate of heavy metal ions on microalgae increases with increasing contact time, and the absorption process usually reaches equilibrium in about 180 min [32].


### **Table 2.**

*Optimal conditions for removing heavy metals using microalgae.*

The important of the effect of temperature in the case of microalgae bioabsorbents is more important for the thermodynamic description of the absorption process than increasing the efficiency of heavy metal absorption. Many studies have shown that increasing or decreasing the temperature (even up to 40°C) has a small effect on the absorption of microalgae.

Discontinuous systems are usually only suitable for the biological treatment of small volumes of wastewater, and for larger scales, it is necessary to use continuous systems in which biological absorbents are used in several cycles of absorption and desorption (recovery). However, it should be noted that the use of microalgae in continuous systems has an important drawback, which is column clogging due to the small size of the bioabsorbent particles. Therefore, in order to ensure the sufficient intensity of wastewater flow through the column, in many types of research, the immobilization of microalgae in different matrices has been proposed, which increases the mechanical strength, particle size, and resistance to chemicals in wastewater [30].

According to the results reported in the articles in continuous systems, the metal absorption capacity and breakthrough time increase with the increase in bed height, which means an increase in the total surface area of the surface absorber. The metal adsorption capacity during adsorption decreases with an increasing initial metal concentration in the solution because the biosorbent becomes saturated faster at high concentrations. For this reason, optimal values for each of these parameters should be determined for practical applications.

Generally, the process of biological absorption in non-living microalgae follows a chemical mechanism, and the main factors that determine the nature of the primary processes are the type of functional groups on the microalgae surface, the nature of heavy metals in the aqueous solution, and the characteristics of the aqueous solution (pH, ionic strength, presence of competing ions, etc.). The absorption of various heavy metals such as Pb(II), Cd(II), Cu(II), Zn(II), etc. using different types of microalgae is mainly done by ion exchange. In confirmation of this point, laboratory

### *Biological Treatment of Heavy Metals with Algae DOI: http://dx.doi.org/10.5772/intechopen.110301*

studies showed that the concentration of light metal ions increases at the end of the biological absorption process [33].

In the complex formation mechanism, both electrostatic interactions and covalent and/or dative bonds are involved and compared to the ion exchange mechanism, the formed surface complexes are more stable. For this reason, the recovery of such biological attractants requires the use of strong agents. Nevertheless, the complex formation mechanism has been proven as the primary interaction in many adsorption processes on different types of microalgae, especially in high initial concentrations of heavy metal ions [34].

Microscopic sedimentation can occur depending on the nature of the microalgae or independent of it and can distort the results of biological absorption and prevent the determination of the absorption rate of metal ions. Although processes such as liming and turning into activated carbon are also used to increase the absorption capacity of microalgae. If the pretreatment of microalgae is done only by drying at 50–60°C (usually for 12–24 h), biomass is not decomposed and the functional groups of its surface are not changed [35–37].

Many studies have shown that pH values in the range of 2–8 lead to an increase in the absorption capacity of most heavy metals by microalgae. In this pH range, heavy metals have high solubility and are in solution as simple ions with the most toxic effect and the highest biological absorption. At lower pH values, the adsorption capacity of microalgae is lower due to the competition between protons and heavy metal ions to bind to biosorbent sites. At higher pH, heavy metal ions are precipitated as hydroxides and only a small amount of heavy metals remain in solutions to be absorbed by interacting with surface groups of microalgae. Due to the insignificant effect of temperature on the absorption capacity of heavy metals by microalgae, it is recommended that, for large-scale applications, the absorption of heavy metals from aqueous solutions in microalgae is carried out at an ambient temperature, because operating costs will be lower in this case. The use of microalgae to absorb heavy metals in continuous systems facilitates the treatment of a large volume of aqueous wastewater, however, research in the fields related to biological absorption in continuous systems is still ongoing [38, 39].
