Microcystis aeruginosa Removal and Simultaneous Control of Algal Organic Matter (AOM) Release Using an Electro-Flocculation–Electro-Fenton (EC-EF) System without Chemical Addition

Abstract

Harmful cyanobacterial blooms pose a serious environmental threat to global water ecology and drinking water safety. Microcystis aeruginosa, a dominant cyanobacterial species in cyanobacterial blooms, was removed using the electro-flocculation–electro-Fenton (EC-EF) technology. In the EC-EF system, the iron anode was used as a sacrificial anode to produce iron ions in situ. Combining the aeration device with the graphite felt cathode as one unit realizes a direct and effective air supply to the cathode, and improves the electrical Fenton efficiency for generating oxidizing groups such as hydroxyl radicals. The cyanobacteria removal efficiency was up to 94.6% under optimal process conditions with a current density of 1.08 mA/cm², an electrolysis time of 5 min, and an aeration flow rate of 0.06 L·min⁻¹. At the same time, the microcystins (MCs) and total organic carbon (TOC) content in the water were controlled. The mechanism of cyanobacterial cell removal using this EC-EF system was investigated via characterization of cyanobacterial cells and flocs and cell membrane permeability analysis. The moderate oxidation and iron hydroxide encapsulation of this system are both beneficial to maintaining the integrity of cyanobacterial cells. The results demonstrated that EC-EF is a chemical-free and eco-friendly cyanobacteria removal technology.

1. Introduction

The eutrophication of water has developed into an environmental public health problem. The occurrence of cyanobacterial blooms impacts water quality and threatens drinking water safety. One of the most notorious species is Microcystis aeruginosa (M. aeruginosa), which is a typical cyanobacterium that causes toxins and odor problems [1]. During the blooms, M. aeruginosa seriously influences water treatment processes by plugging the filtration tanks or membranes. Moreover, cyanobacteria can produce algal organic matter (AOM), including toxins, odorants, and precursors of disinfection by-products (DBPs), which are harmful to humans, animals, and aquatic biota [2]. Therefore, many efforts have been taken to effectively remove cyanobacteria.

Various methods have been used to remove M. aeruginosa, such as coagulation [3], peroxidation [4], air flotation [5], approaches using a membrane [6], and ultrasonic methods [7]. Pre-oxidation enhanced coagulation for cyanobacterial removal leads to problems of secondary residual metal ions, the release of intracellular organic matter (IOM) [8], and elevated concentrations of DBPs. The effect of air flotation is poor in removing cyanobacteria because both cyanobacteria and air bubbles are negatively charged, resulting in weak adhesion [9]. Ultrasonic and membrane methods have high energy consumption and cost [10]. With the booming development of power systems and related supporting facilities, the use of electrochemistry to remove pollutants from water has received more attention. The electrochemical method does not require a large amount of flocculant and solid–liquid separation is relatively easy, and this method has thus attracted more and more attention from the researchers in this field [11].

Electrochemical cyanobacteria removal technology is mainly divided into electro-coagulation (EC) and electro-oxidation (EO). Electro-coagulation mainly uses sacrificial anodes (iron or aluminum electrodes) to produce flocculants in situ, which combine with cyanobacterial cells to form flocs and then precipitate them [12]. The main effect of electro-oxidation is that oxidizing substances (e.g., H₂O₂, HClO, ClO₂) and free radical groups (e.g., ·OH, ClO⁻, SO₄⁻·) produced by electrochemical reactions attack cyanobacterial cells to inactivate them [13]. Electro-coagulation and electro-oxidation cyanobacteria removal technologies have been separated in most studies. Studies on electro-oxidation for cyanobacteria removal have mostly focused on anodic oxidation. For example, Liang et al. used Ti-RuO₂ anode and galvanized tube cathode electrochemical cycle treatment for cyanobacteria removal, which took 52 min, and cyanobacterial cells lost their viability and potential for survival and growth [14]. Mascia et al. used a three-dimensional conductive diamond (BDD) anode for cyanobacterial removal, finding that the prevailing mechanism of M. aeruginosa inactivation is the disinfection by bulk oxidants (active chlorine) [15]. Regarding electro-coagulation for cyanobacteria removal, sacrificial anodes are mostly used. For example, the Al-Al electrode system was used to remove cyanobacteria via the electro-coagulation-flotation (ECF) process and achieved complete removal of cyanobacterial cells after 45 min at a current density of 1.0 mA·cm⁻² [16]. Alejandro et al. used lead–aluminum multiple electrodes in parallel for electro-coagulation to remove cyanobacteria and showed that cell removal occurred through charge neutralization with the Al hydroxides generated in the system via bridging and an electrostatic patch aggregation mechanism [17].

However, electro-coagulation or electro-oxidation has its disadvantages. First, the electrolysis time is too long, mostly around 20 to 60 min, resulting in high energy consumption [16]. Second, electro-oxidation has a very strong destructive effect on cyanobacterial cells, which increases the release of AOM, such as toxins and precursors of DBPs [15]. The composition of AOM is complex, and it is difficult to remove all of it using a conventional water treatment process. There is often residual cyanobacteria AOM in the effluent of water treatment plants, which will reduce the safety of the water ecosystem. Finally, the electrode material is expensive, increasing the cost of the system. Therefore, in this manuscript, electro-coagulation and electro-oxidation (EC-EO) were combined to efficiently remove cyanobacteria without any chemical addition. In this combined system, the electro-coagulation and oxidation possibly cooperate to improve the removal efficiency and water safety.

Among the electro-oxidation processes, the electro-Fenton process (EF) is well known for its simplicity and high efficiency with Fe²⁺ and H₂O₂ being readily available. The ·OH is produced from Fe²⁺ and H₂O₂ and then degrades the organics or other pollutants [18]. H₂O₂ concentration is a critical parameter for EF. The concentration of H₂O₂ determines the potential ·OH that will be generated in the process. In a typical process, air/O₂ is continuously supplied to the cathode, where the generation of H₂O₂ is induced by a two-electron reduction of O₂. With the continuous in situ electrogeneration of H₂O₂, the costs associated with the direct external addition of H₂O₂ are eliminated. Compared to the cost of external addition of H₂O₂ associated with the transportation and storage of H₂O₂, Huang and Chu argued that cost reduction of up to 80% could be obtained with the in situ electrogeneration of H₂O₂ [19]. To enhance the H₂O₂ yield, previous studies have often modified the cathode electrode to increase the active sites of oxygen utilization. However, the oxygen supply mode and utilization rate are often ignored [20]. Previous studies have used aerator heads to inject air or oxygen, and the greater supply of air or oxygen escapes in the form of bubbles and is not well utilized, resulting in extremely low air utilization and increased energy consumption. Therefore, it is necessary to propose a new electric cathode to realize the direct supply of gas in order to improve the air or oxygen utilization efficiency.

In terms of applying the electro-Fenton reaction for cyanobacteria removal, Long et al. (2017) [21], Lian et al. (2020) [22], and Huang et al. (2022) [23] examined a three-dimensional conductive diamond and carbon felt (BDD-CF) electrochemical system, a Pt/Ti and activated carbon fiber/nickel foam (Pt/Ti-ACF/Ni) electrochemical system, and a Pt and graphite felt (Pt-GF) electrochemical system, respectively, for cyanobacteria removal via the addition of Fe²⁺. The results demonstrated that the mechanism of cyanobacteria removal was dominated by homogeneous and non-homogeneous electro-Fenton action, accompanied by a small quantity of electro-coagulation, electro-adsorption, and electro-flotation processes [22]. Although studies of the electro-Fenton reaction for cyanobacteria removal have gradually emerged in recent years, the number of studies is relatively limited at present. Most importantly, these studies improved the removal efficiency with the addition of Fe²⁺. Excessive production of iron sludge is difficult to dispose of and may result in secondary pollution. In this study, iron was used as an anode that could continuously produce Fe²⁺ directly without any chemical addition, which could minimize the potential sludge generation from the precipitation of Fe³⁺. Combining the advantages of EF and EC to act on the removal of cyanobacterial cells in cyanobacteria-containing waters is a research topic that deserves further exploration.

In this manuscript, the iron electrode was selected as the anode and ventilated graphite felt as the cathode. A simultaneous oxidative coagulation and cyanobacteria removal system based on the combination of EF oxidation and EC was constructed. To improve the efficiency of the EF-EC in the removal of cyanobacteria, the aeration device and the graphite felt cathode were combined to form an integrated ventilation cathode to improve its performance. The main objectives were to investigate (i) the efficiency of EF-EC in the removal of M. aeruginosa without any chemical addition and (ii) achieve effective control of algal organic matter (AOM), including MCs. Additionally, the mechanism of the combined system was revealed. This study provides an environmental system to effectively remove cyanobacteria and control its AOM without any chemical addition.

2. Materials and Methods

2.1. Manufacturing Method of Graphite Felt Integrated Ventilation Cathode

The integrated ventilation cathode consists of a base plate, an orifice plate, and graphite felt electrodes stacked together, as shown in Figure S1. The base plate was made by 3D printing technology and has an air inlet for connecting the air inlet hose. Figure S2 shows a schematic diagram of the gas movement in the integrated ventilation cathode. Considering the characteristic of gas floating in water, the air inlet is set at the lowermost part of the bottom plate. To evenly disperse the incoming gas throughout the cathode, an orifice plate is provided between the bottom plate and the electrode to act as a guide for the gas flow. The bottom plate, the orifice plate, and the graphite felt electrode are all of the same size, and the edges of the three are bonded with inorganic silicone to form a closed space called the “gas chamber”, which is the most important operation for improving the utilization of oxygen. The effective electrode area immersed in the electrolytic solution is 5.0 cm × 6.5 cm, which is the same as the counter electrode iron anode.

2.2. Cyanobacterial Strain and Experimental Design

M. aeruginosa (strain FACHB-905) was obtained from the Institute of Hydrobiology, Chinese Academy of Sciences. It was cultured in BG11 medium with a light/dark cycle of 12 h/12 h.

Steady-state M. aeruginosa cells were harvested and centrifuged at 4000 rpm for 10 min. The cyanobacteria precipitates were collected and diluted with deionized water to a concentration of 2.48 × 10⁶ cells·mL⁻¹. Then 400 mL of cyanobacteria solution and 100 mL of sodium sulfate electrolyte solution were used for electrolysis in a 500 mL beaker. The pH value was not specially adjusted in the experiment.

2.3. Analytical Methods

2.3.1. Cell Density and Removal Efficiency

The cell density was determined by measuring the samples’ optical density at 680 nm with a UV–vis spectrophotometer (L6S, Lengguang, Shanghai, China) (OD₆₈₀) [24], which was then positively correlated with cell number. The removal efficiency was calculated based on the variations of cell density using Equation (1).

$$\mathrm{R}=\frac{{\mathrm{C}}_{\mathrm{i}} - {\mathrm{C}}_{\mathrm{f}}}{{\mathrm{C}}_{\mathrm{i}}} \times 100\%$$

(1)

where R is the removal efficiency, and C_i and C_f are the initial and final density of M. aeruginosa (cells·mL⁻¹), respectively.

2.3.2. Determination of TOC and Residual Fe

The supernatant after precipitation was filtered using a 0.45 μm membrane filter, and the TOC content was determined using a total organic carbon meter (TOC-L, Daojin, Kyoto, Japan). The residual iron ion concentration was analyzed using the colorimetric method of o-phenanthroline.

2.3.3. Scanning Electron Microscopy (SEM)

The supernatant was carefully removed with a syringe after sedimentation, leaving just cyanobacterial sediment in the beaker. The beaker containing the cyanobacterial sedimentation was placed in a freeze dryer (LGJ-10, Beijing, China) for three days to dry. Then it was observed via SEM (ZEISS GeminiSEM 300, Carl Zeiss, Oberkochen, Germany).

2.3.4. Flow Cytometry Analysis

The electrochemically treated cyanobacterial solution was stained with the SYTO 9/PI live and dead bacteria double dyeing kit, and then the deactivation of cyanobacterial cells was detected by flow cytometry. The live/dead bacterial double staining kit exploits the working principle that SYTO 9 and PI differ in their spectral characteristics and ability to penetrate healthy bacterial cells. When used alone, SYTO 9 can label all bacteria in a population (i.e., bacteria with intact and damaged membranes); instead, PI can only penetrate the damaged membrane. Therefore, when two dyes are added to the system, the insertion of PI will cause the decrease in SYTO 9 staining fluorescence. Thus, bacteria with intact membrane structures fluoresce green, and bacteria with damaged membrane structures fluoresce red based on a mixture of SYTO 9 and PI in appropriate proportions. The maximum excitation and emission wavelengths of the two dyes are 480/500 nm (SYTO 9) and 490/635 nm (PI).

2.3.5. Microcystins Concentration

The processed samples were centrifuged at 4000 rpm for 10 min. The supernatant was filtered through a 0.22 μm pore size membrane filter to obtain an extracellular MC sample. The total MC concentrations were evaluated using the enzyme-linked immunosorbent assay (ELISA) method with the kit (Product No. 520011, Abraxis, Warminster, PA, USA).

3. Results and Discussion

3.1. Evaluation of Different Parameters on M. aeruginosa Removal

3.1.1. Influence of Electrolysis Time on Cyanobacteria Removal

The influence of electrolysis time (1, 3, 5, 7, 9, 11, 13, and 15 min) on the cyanobacteria removal was investigated as shown in Figure 1a. At 1 min and 3 min, the cyanobacteria basically had no effect of settling down after coagulation, for which removal efficiencies are quite low. According to the electrochemical Faraday’s law, the amount of Fe²⁺ released from the anode is linearly and positively related to the current size and the electrolysis time [11]. With the electrolysis time increasing, more Fe²⁺ is released to the water. The coagulant surface area and adsorption sites available increase, and Fe(OH)_x precipitates to wrap more cyanobacterial cells [25]. Therefore, the highest cyanobacteria removal rate of 90.58% was achieved at 5 min. After 5 min, the removal rate decreased slightly with increasing electrolysis time. The possible reason for this is that, with the increase in electrolysis time, cyanobacterial cells or formed cyanobacterial flocs are directly damaged by excessive electron attack or oxidation of various types of oxidation groups [26].

The electrolysis time chosen in this study was 5 min, which is the shortest time compared to previous studies. For example, Liang et al. used a 52 min electrochemical cycling treatment to achieve 80% cyanobacterial removal [14]. Graphite felt (GF) was used as an anode at 75 mA for 30 min to achieve 94% removal [23]. Gao et al. utilized a monolithic ceramic electrode (MCE) electrode to achieve 89.2% chlorophyll-a removal over 120 min [27]. A shorter electrolysis time reduces the cost of electricity and energy consumption and also reduces the hydraulic retention time of the treatment process in the development of practical applications.

Figure 1a also shows that the residual iron ion concentration was below the World Health Organization drinking water standard limit of 0.3 mg·L⁻¹ within 11 min of electrolysis, and then rapidly increased to 0.57 mg·L⁻¹ and 0.62 mg·L⁻¹ at 13 and 15 min, respectively, exceeding the concentration limits for drinking water. Figure S3 shows the photos of cyanobacteria removal after sedimentation at different electrolysis times. The color of the solution became brown, indicating more Fe²⁺/Fe³⁺ in the solution and cyanobacterial cells being damaged by oxidation, at 13 and 15 min. The results demonstrated that the EC-EF system can control the residual iron ions in water. Under the optimal electrolysis time of 5 min, the iron ions generated by the electrochemical process were effectively utilized, and their hydrolysis products were coagulated with cyanobacterial cells to form flocs, aiding the removal of cyanobacterial cells.

3.1.2. Influence of Current Density on Cyanobacteria Removal

Current density relates to energy consumption and is an important parameter. The removal efficiencies at different current densities are shown in Figure 1b. The results show that the cyanobacteria removal increased with the increase in current density from 0.15 mA/cm² to 1.08 mA/cm². The cyanobacterial removal rate varied drastically for currents of 0.15 mA/cm² and 0.46 mA/cm², and was less significant at high current variation segments, similar to previous reports [12,27]. The cyanobacteria removal rate reached a high level of 94.3% under 1.08 mA/cm². With the increase in current density, the production of iron ions and iron hydroxyl compounds required for coagulation increased [28]. On the other hand, the electron transfer efficiency and oxidation generation were enhanced with increasing current density, and the study of Zhou et al. also showed that the inactivation efficiency increased with the increase in the applied current [29]. When the current continued to increase to 1.38 and 1.69 mA/cm², there was a decreasing trend of cyanobacteria removal. This EC-EF system achieved the highest cyanobacteria removal efficiency at a smaller current density of 1.08 mA/cm², compared to anodic oxidation systems such as those using a BDD anode (8 mA/cm²) [29] or Ti/RuO₂ anode (optimal current density of 5–10 mA/cm²) [14].

The vital center of the cell is protected by a membrane that consists mainly of a biomolecular layer of phospholipids with hydrophobic and hydrophilic properties. Protein inclusions within the membrane control the transport of ions in response to changes in the cellular environment. Phospholipid membranes are less susceptible to oxidation, whereas proteins are easily destroyed by the direct action of electric fields. Next, due to the electrical resistance of the cell membrane, the applied current creates a potential difference between the two ends of the cell membrane. This potential difference changes the cell membrane transmembrane potential [30]. Once again, the presence of an electric field restricts the movement of ions (electrons and low mass ions) to a limited area. They are unable to achieve normal membrane transfer, affecting the physiological function of the cell. However, the cell, although unable to transport more ions, can be reactivated in a favorable medium. Its complete destruction requires an oxidizing agent capable of crossing the membrane to reach the center of life [30]. Accordingly, from one side, the electric field synergistic effect is subsequently enhanced by increasing the current, which may increase the dielectric constant of the cell membrane, thus making it easier for oxidants produced by the system to enter the cell, and thus accelerating cell damage [26]. Another mechanism is the electrochemical inactivation of cyanobacterial cells due to direct charge transfer occurring at the electrode surface [31]. Both mechanisms are enhanced with increasing current density [32,33,34], and each cannot be viewed independently and separately.

Figure 1b also shows the residual iron concentrations with different current densities. This clarifies that the residual iron concentration in all groups is lower than the WHO drinking water standard of 0.3 mg·L⁻¹. The residual iron concentration was highest at 0.15 mA/cm², probably because the quantity of iron ions produced at 0.15 mA/cm² was not sufficient to form iron hydroxide compounds to cross-link and coagulate cyanobacterial cells and exist in solution as water-soluble iron ions, resulting in a higher residual iron concentration in solution and low cyanobacterial removal.

3.1.3. Contribution of Aeration on M. aeruginosa Removal

To improve the efficiency of oxygen utilization by the cathode, this study combined the aeration device with the cathode through manual processing to achieve a direct gas supply to the cathode. Different amounts of gas were injected into the integrated ventilated cathode at the optimal electrolysis time (5 min) and optimal current density (1.08 mA/cm²). As shown in Figure 2a, when the amount of gas changed in the range of 0.02–0.1 L·min⁻¹, the corresponding cyanobacterial removal rate was 89.3–94.6%, which was higher than the cyanobacterial removal rate without aeration (85.3%). However, when the aeration flow rate increased to 0.2 L·min⁻¹, the removal efficiency of cyanobacteria decreased to 81.2%. This indicated that excessive aeration was not conducive to the removal of cyanobacteria, which may be related to the influence of aeration on the change in hydraulic conditions, which then affects the aggregation and sedimentation of cyanobacteria.

To further verify the presence and type of active components in the whole reaction process, a study of the EPR test of DMPO-⋅OH for the Fe-GF integrated ventilated cathode electrochemical system was conducted in this study. As shown in Figure 2b, the system showed an obvious “1:2:2:1” ⋅OH signal characteristic peak, which was similar to that in the study of Dai et al. [35]. The signal peaks of the Fe-GF integrated ventilated cathode system at the 5 min test point were significantly higher than those at the 2 min test point, indicating that the integrated cathodic ventilation device can directly provide sufficient oxygen for the electro-Fenton reaction, which is conducive to the occurrence of the electro-Fenton reaction and the production of active substances.

3.2. Effects of EC-EF on the Physiology of M. aeruginosa

3.2.1. Morphology of M. aeruginosa

The SEM images of cyanobacterial cells and flocs after EC-EF treatment are shown in Figure 3a. It can be seen that the flocs containing M. aeruginosa are similar to sponge-like agglomerates. The floc covers and wraps the cyanobacterial cells and is unevenly distributed. Compared with normal spherical shapes and smooth surfaces of M. aeruginosa cells, some cyanobacteria cells were still spherical and intact, but some of the cell walls were wrinkled without clear damage (Figure 3a). This demonstrated that EC-EF only caused a morphology alteration, which is significant for control of intracellular organic matters and MC release.

To better understand the formation of flocs, EDX analysis was performed on two sites, as shown in Figure 3b,c. One site was located on the cyanobacterial cell (Figure 3b) and the other on the floc surface (Figure 3c). In the elemental composition analysis in Figure 3b, C, N, and O are the main elements of the cell, accounting for the largest proportion of the elemental composition, i.e., 66.1%, 10.14%, and 20.69% of the total weight, respectively. In Figure 3c, the composition ratios of C, N, and O elements in flocs were not significantly different from those of cyanobacterial cells (Figure 3b), which is similar to the results of Yue Dong et al. [36]. The percentage of iron detected in cyanobacterial cells was 0.09% (Figure 3b), and in cyanobacterial flocs it was 1.02% (Figure 3c). The percentage of elemental oxygen also increased slightly. These results indicate that the flocculants with coagulation effects are iron and their hydrolysates, confirming that flocculants are the main acting factors for coagulation and sedimentation of cyanobacterial cells. The iron hydroxyl compounds produced by the electro-coagulation-Fenton reaction are partially adsorbed on the surface of cyanobacterial cells and act as flocculants, bridging between cyanobacterial cells, forming flocs, netting, sweeping, and settling. There is also some hydroxylated iron that binds to AOM released from cyanobacterial cells, binds to MCs from stressed cyanobacterial cells, and then precipitates.

3.2.2. Variations of Cyanobacterial Cell Integrity

Figure 4 shows the flow cytometry results of M. aeruginosa via EF-EC treatment with different electrolysis times. In Figure 4, B3 represents live cells, quadrant B4 represents early apoptotic cells, and quadrant B2 represents late apoptotic cells. From 5 to 9 min, there was an increasing trend in cell inactivation due to the cumulative effect of EF. This indicates that the oxidation inactivation becomes stronger at 7 min and 9 min, resulting in a decrease in the activity of cyanobacteria.

The inactivated cyanobacterial cells mainly involve the following causes: Firstly, the electrode exhibits adsorption and direct oxidation of cyanobacterial cells. The surface of cyanobacterial cells is negatively charged, and some of them are adsorbed on the surface of the anode. Direct electron exchange occurs between them and the electrode, resulting in direct inactivation. Secondly, electromagnetic fields are harmful to cells [37]. For cells, transmembrane potentials above 1 V and long pulse times lead to irreversible osmosis and cell death [38]. This transmembrane potential depends on experimental conditions such as the pulse or duration of the electric field [39]. The larger inactivation observed at longer electrolysis times is due to the avoidance of membrane recovery, which greatly increases ion transfer and triggers irreversible cellular damage. In addition, these pores may allow short-lived oxidants such as ·OH, ·OOH, ${\mathrm{SO}}_4^{·-}, {\mathrm{and} \mathrm{HSO}}_4^{·}$, and long-lived oxidants such as H₂O₂, to freely enter the interior of the cell and aid in the inactivation process [13].

3.3. The Effects of AOM Release and Control

3.3.1. TOC Changes and Control

The AOM release with different electrolysis times, as indicated by changes in soluble TOC in the supernatant after EC-EF treatment, are shown in Figure 5. The TOC value of cyanobacteria-laden water before treatment was 5.6 mg·L⁻¹. When the electrolysis time increased from 1 to 15 min, the TOC values after EC-EF varied from 0.34 to 2.15 mg·L⁻¹, which are lower than the regulatory limits of “Standards for drinking water quality” (GB5749-2022, China) (5 mg·L⁻¹). The TOC values were higher at 1 min and 3 min with 1.69 mg·L⁻¹ and 2.15 mg·L⁻¹, respectively, and ranged from 0.34–0.65 mg·L⁻¹ during the 5–15 min interval of electrolysis. When the electrolysis time was too short, the number of flocs formed by hydroxide iron was less. Therefore, the cyanobacteria cells could not settle. With the increasing electrolysis time, the flocs played a key role in coagulation to settle with AOM. The residual TOC was lowest at 9 min but increased at 11 min. This may be due to the destruction of the cyanobacterial cells caused by ·OH and other oxidants, resulting in the intracellular organic matter released from the cyanobacterial cells. The results demonstrated that the EC-EF system only caused limited damage to the structure of M. aeruginosa cells, which was collaborated by the results of Figure 3a. AOM could be released from the interior or the surface of the cell into the solution, especially with longer electrolysis time, and then be decomposed by ·OH and other oxidant compounds, which corresponds to the 13–15 min decline in the figure. During the EC-EF process, both coagulation and oxidation processes could lead to a decrease in TOC, which would contribute significantly to the control of DBPs.

According to the results presented in Figure 3a, the cell structure was kept intact after EC-EF processes. It can be speculated that intracellular matters did not leak out or, if they did, only to a minimal degree. The AOM removed by the oxidation process was mainly extracellular organic matter.

3.3.2. Release and Control of MCs

The release of MCs at different electrolysis times is shown in Figure 6. The quantity of MCs in the supernatant after EC-EF treatment showed an overall increasing trend with the increase in electrolysis time. The result indicates that the cyanobacterial cells are stressed to release more MCs due to the attack of electrons and the influence of oxidation groups generated by electrochemical processes. In addition, MCs could be degraded or settled during the EC-EF process. Therefore, the residue MCs in all treatment groups were below the national drinking water limit of 1.0 μg·L⁻¹ at different electrolysis times from 1 to 15 min. The EC-EF did not cause cell tearing and effectively controlled the release of MCs, which is a promising cyanobacteria removal technique for application.

Alejandro et al. [17] investigated the release of MCs during electro-coagulation of aluminum electrodes with cyanobacterial densities of 10⁵ and 10⁶, at currents of 2.5–12.5 A and a hydraulic retention time of 2.2 min. The MC concentrations exceeded the standard limit of 1.0 μg·L⁻¹, and both the current and the MC release were several orders of magnitude higher than those in this study, but corresponded to an inactivation efficiency of lower than 40% for M. aeruginosa. The trend of MC release was the same in both studies.

The chemical structure of MCs is shown in Figure S5. MCs are small monocyclic peptides consisting of seven amino acids linked by peptide bonds. Their general structure is cyclic (D-alanine-X-erythro-β-methyl-D-isoaspartate-Y-Adda-D-isoglutamic acid-N-methyl dehydroalanine). X and Y indicate the positions occupied by variable L-type amino acids [40]. This substitution also allows the existence of numerous MC isomers in natural waters, and MC-LR is generally chosen as a model for study. As shown by its molecular structure, MC-LR contains two carboxyl ionizable groups and one amino ionizable group with pK_a values of 2.09, 2.19, and 12.48, respectively. Because of the possibility of (COOH)₂(NH₂⁺) and (COO⁻)(COOH)(NH₂⁺) dominant species formation in extremely acidic environments, and in general pH and alkaline environments, MC-LR carries a net negative charge (in the form of (COO⁻)₂(NH₂⁺) or (COO⁻)₂(NH)) [41]. Therefore, influenced by the electric field in the system, MC-LR will move toward the anode and be degraded by the catalytic oxidation of the anode. In addition, Gao Yu et al. used Ti/Pt electrode electrochemical treatment, and the transcript levels of MCs synthesizing genes mcyB and mcyD decreased to be undetectable on days 2 and 6. The study showed that the electrochemical system played an important role in down-regulating the transcript levels of MC synthesizing gene clusters, MC-LR concentration, and cell density [42].

3.4. The Processes and Mechanism of Cyanobacteria Removal and Simultaneous AOM Control by EC-EF

Cyanobacterial-laden water is a heterogeneous system. The process and mechanism in this EC-EF system to remove cyanobacteria are different from those of a homogeneous system. The schematic diagram in Figure 7 shows the processes of EC-EF for removal of cyanobacteria. In the system, the iron acts as a sacrificial anode, releasing Fe²⁺ into solution (Equation (2)), which is the initial source of a range of iron species in the system.

In the cathode, with air aeration, H₂O₂ was continuously generated through two-electron oxygen reduction (Equation (3)). Meanwhile, in situ Fe²⁺, from anode oxidation, reacted with H₂O₂ to produce ·OH (Equation (4)) and then regenerated via a direct cathodic reaction (Equation (5)). In this heterogeneous system, cyanobacteria (M. aeruginosa) were oxidized to inactive cells by O₂/H₂O₂/·OH (Equation (6)). During this process, M. aeruginosa were possibly destroyed by the oxidants, as many studies have reported [13,26]. However, in this study, the results of SEM demonstrated that most cells remained intact and maintained a round shape with less activation without obvious cell lysis (Figure 3). The values of TOC and MC also clarify that intracellular organics were not largely released (Figure 5 and Figure 6).

$$\mathrm{Fe} -2{\mathrm{e}}^{-}\to {\mathrm{Fe}}^{2+}$$

(2)

$${\mathrm{O}}_2+2{\mathrm{H}}^{+}+2{\mathrm{e}}^{-}\to {\mathrm{H}}_2{\mathrm{O}}_2$$

(3)

$${\mathrm{Fe}}^{2+}+{ \mathrm{H}}_2{\mathrm{O}}_{2 }+{ \mathrm{H}}^{+}\to { \mathrm{Fe}}^{3+}+{\mathrm{H}}_2\mathrm{O}+·\mathrm{OH}$$

(4)

$${\mathrm{Fe}}^{3+}+{\mathrm{e}}^{-}\to {\mathrm{Fe}}^{2+}$$

(5)

$${\mathrm{O}}_{2 }/{\mathrm{H}}_2{\mathrm{O}}_{2 }/·\mathrm{OH}+\mathrm{cyanobacteria}\to \mathrm{cyanobacteria}* +\mathrm{AOM}+{\mathrm{H}}_2\mathrm{O}$$

(6)

$${\mathrm{Fe}}^{2+}+2\mathrm{O}{\mathrm{H}}^{-}\to \mathrm{Fe}(\mathrm{OH}{)}_{2 }$$

(7)

$${\mathrm{Fe}}^{3+}+3\mathrm{O}{\mathrm{H}}^{-}\to \mathrm{Fe}(\mathrm{OH}{)}_{3 }$$

(8)

$$\mathrm{Fe}(\mathrm{OH}{)}_{2 }/\mathrm{Fe}(\mathrm{OH}{)}_{3 }+\mathrm{cyanobacteria}*+\mathrm{AOM}+\mathrm{MCs} \to \mathrm{Flocs}$$

(9)

With the production and cycling of Fe²⁺ and Fe³⁺, the in situ coagulants, Fe(OH)₂ and Fe(OH)₃, were continuously generated in the cyanobacteria-laden solution (Equations (7) and (8)). These coagulants, Fe(OH)₂ and Fe(OH)₃, could sweep and flocculate the cyanobacterial cells to form flocs, promote the continued growth of flocs, and play a role in the final bridging settlement. Moderate oxidation causes shrinkage of cyanobacterial cells, which affects their surface properties and cellular activity, making them more prone to coagulation and sedimentation [43]. Consequently, the combined action of the flocs’ adsorption sweeping and hydroxyl radical oxidation leads to the maximum removal of M. aeruginosa, coincidently causing the AOM and MCs to settle (Equation (9)).

The EC-EF system has more prominent advantages. First, because of the in situ coagulants and oxidants, extra chemical reagents do not need to be added to the whole system. Second, the relatively short reaction time (5 min) leads to low energy consumption. Finally, AOM and MC release was controlled simultaneously without secondary pollution. This is a promising technology that can ensure water safety while efficiently removing cyanobacteria.

4. Conclusions

Coupled simultaneous oxidation and coagulation for cyanobacteria removal using an electro-Fenton/electro-flocculation system was proposed. Iron electrodes were chosen as the anode and a bespoke integrated ventilated cathode as the cathode. A cyanobacterial removal rate of 94.6% was achieved via electrolysis at 1.08 mA/cm² for 5 min without chemical addition. The water quality parameters, such as TOC, residual iron, and MCs, were effectively controlled to ensure the safe water quality. From the results of EPR, SEM, etc., it can be seen that the ventilated cathode helps the system to generate hydroxyl radicals and other oxidizing groups to produce moderate oxidation of cyanobacterial cells so that the activity of the cyanobacterial cells is reduced to be more conducive to the removal of coagulation and sedimentation. Overall, this is a low-carbon, green, and non-polluting cyanobacteria removal technology.