Application of acidophilic microorganism in the metal enrichment of electroplating sludge use the membrane bioreactor

Background: Bioleaching is an important technology for treating electroplating sludge. Previous researches have focus on improving the leaching rate of metals in electroplating sludge by bioleaching. However, the concentration of heavy metals in the leachate after single leaching was lower, which is quite unfavorable for subsequent metal recovery. Additionally, membrane bioreactors (MBRs) have been widely used in the field of sewage treatment. Research on the application of bioleaching technology combined with MBRs to enrich metals in electroplating sludge has not been reported. Therefore, in this study, we first combined bioleaching technology and MBRs for metal enrichment in electroplating sludge to obtain the key technology of "acid production - electroplating sludge regeneration -repeated electroplating sludge leaching - achievement of valuable metal enrichment". Results: In this research, through scaling up from the laboratory scale (shake flasks) to a factory-scale application (10 m³ membrane bioreactors), we mastered the key technology of acid production by acidophilic microorganism, and the acid solution can be repeatedly used for metal leaching. The results showed that the MBR maintained high-density cell growth (≈2.1×10 9 /mL) and a stable sulfuric acid production rate (850 L/h) throughout the entire operational period. Under the above conditions, the maximum cycle number (10 times) for enrichment of the target metals in the electroplating sludge was obtained. Additionally, after the end of the cycle enrichment process, the concentrations of the target metals Ni + , Cu 2+ , and Zn 2+ were 13.867 g/L, 18.118 g/L and 21.075 g/L, respectively, which were highly enriched. Conclusions: This study first solved the difficulties in the industrialization of bioleaching electroplating sludge through combining bioleaching technology and MBRs. Furthermore, this research can provide a demonstration project for the industrial application of MBR-bioleaching technology in electroplating sludge, with a view to applying this technology to the disposal of more types of hazardous waste.


Background
Membrane bioreactors (MBRs) are a new water treatment technology that combines membrane a separation unit with a biological treatment unit. The membrane module is used to replace the secondary sedimentation tank in the traditional activated sludge process [1,2]. Membrane bioreactors not only occupy a small area, produce less sludge, and operate flexibly and stably but also achieve high-efficiency solid-liquid separation due to the excellent interception performance of the membrane module [3,4], which makes the effluent water quality better and enables direct recycling of the effluent to achieve wastewater reuse [5,6]. To analyse the current research direction and trends related to MBRs, a Web of Science database was used to search for the keywords "membrane bioreactor" and "bioleaching" from January 2010 to 2020. Using CiteSpace software, the evolution of the keywords in the field of MBR in the past ten years was obtained, and the dynamic evolution of the research hotspots was analysed with respect to time. The results are shown in Fig. 1  The electroplating industry produces a great deal of wastewater, which has a high chemical oxygen demand, a high heavy metal content and a complex composition [7].
Moreover, the types and concentrations of pollutants in wastewater vary with the types of metal plating and the cleaning methods used for plating parts [8]. Therefore, it is necessary to treat electroplating wastewater to reduce the concentration of heavy metals in wastewater and to meet the relevant discharge standards. At present, electroplating wastewater is generally treated by the traditional treatment method of "pollution first, treatment later", and a large amount of electroplating sludge is generated during the treatment process. Statistically, nearly 5×10 4 tons of electroplating sludge is discharged per year in China [9]. Electroplating sludge has the characteristics of a large discharge volume and high harmfulness. If not treated properly, such sludge will cause serious secondary pollution to the environment [10].
Therefore, it is urgent to seek a scientific, green and safe electroplating sludge treatment technology to realize the recovery and utilization of resources. Such technology is highly significant for protecting the environment, improving the level of environmental governance and promoting the sustainable development of the electroplating industry [11].
At present, safe landfilling after stabilization/solidification is the main method used to treat electroplating sludge [12,13]. However, the cost of this method is high (200-500 EUR/ton), which undoubtedly increases the economic burden of electroplating enterprises [14,15]. In addition, safe landfilling not only leads to certain environmental risks but also causes the loss of metal resources. Therefore, the recovery of valuable metals from electroplating sludge is the goal of electroplating enterprises; such recovery can reduce the environmental risk and effectively mitigate the loss of valuable metal resources. Hydrometallurgy and pyrometallurgy are two primary metal recovery processes [16,17]. The principle of hydrometallurgy is based on the feature that metals are easily soluble in acids. After strong acid extraction, metal ions can be separated and purified by different methods. However, this method uses strong acid as the medium, has high requirements in terms of materials and has a high safety risk. The principle of pyrometallurgy is that under high-temperature conditions, metals are separated through oxidation, reduction, decomposition, volatilization, and condensation to achieve the separation of different components.
However, this method requires high energy consumption and a large investment and produces substantial secondary pollution.
Our previous research [18] showed that a mixed bacterial system consisting of This research focuses on shortening the acid production period through a self-designed MBR and improving the pulp density of leaching electroplating sludge through designing the circulation parameters. Additionally, in this study, a combination of MBR and bioleaching technology is first used for the disposal of electroplating sludge to obtain the key technology of "acid productionelectroplating sludge leachingleachate regenerationrepeated electroplating sludge leachingachievement of valuable metal enrichment". Furthermore, this research can provide a demonstration project for the industrial application of MBR-bioleaching technology in electroplating sludge, with a view to applying this technology to the disposal of more types of hazardous waste.

Sample digestion
In this research, the electroplating sludge came from an electroplating factory in Taizhou, Zhejiang Province. The samples were dried and crushed for analysis. A suitable amount of sample was put into the digestion instrument (SH230N, Haineng Instrument, china), and the electroplating sludge sample was digested by the "hydrofluoric acid and nitric acid and hydrochloric acid method" [19,20]. The concentrations of heavy metals in the samples were analysed by inductively coupled plasma-atomic emission spectrometry (TCAP6300 SERICS, Thermo Scientific, US).

X-ray fluorescence analysis (XRF) of samples
XRF (XRF-1800, Shimadzu, Japan) can analyse the kinds and contents of elements in a sample. X-rays produce fluorescence and excite the tested sample, and different elements in the sample will emit secondary X-rays with different energy characteristics or wavelength characteristics. The secondary X-rays are used in the detection system for energy and quantity analysis, and the analytical information is transformed by the instrumental software to obtain the kinds and contents of different elements in the samples. In this research, the analytical conditions were as follows: X-ray tube pressure: 60 kV max, 150 ma max; X-ray tube target: Rh target; detection range of elements: 4 Be-92 U; and maximum scanning speed: 300/min.

Optimum pulp density for metal leaching of electroplating sludge
Based on our previous research [18], under the optimized process conditions of 33.6 ℃ for the temperature, 3.1 m 3 /min for the aeration rate and 83 rpm for the stirring velocity, a high production efficiency of H2SO4 at pH 1.0 (≈850 L/h) was obtained [21]. The sulfuric acid produced by the membrane bioreactor was used for metal leaching experiments in electroplating sludge. To study the effect of different pulp densities in 10 m³ extraction tanks on the leaching of heavy metals in electroplating sludge, six groups of different pulp densities were designed for comparison. The experimental groups are shown in Table 1.
Step 1: The sludge is crushed and collected for leaching.
Step 2: All circuits, instruments, and equipment are checked for faults.
Step 3: The biological sulfuric acid produced by the membrane bioreactor is pumped into the 10 m³ extraction tanks through an acid-resistant pump.
Step 4: The stirring system is turned on, and the crushed electroplating sludge is slowly added according to the pulp density (w/v) in Table 1. Stirring proceeds for 180 min at 120 rpm.
Step 5: Every 30 minutes, a sample is collected with a 10 ml pipette and centrifuged, and then the centrifuged supernatant is collected and diluted in a test tube. ICP-AES (Inductively Coupled Plasma-Atomic Emission Spectrometry, TCAP6300 SERICS, Thermo Scientific) is used to analyse the heavy metal concentration in the leachate and calculate the leaching rate of each metal.
Step 6: Solid-liquid separation. After stirring, the sludge in the stirring tank is passed into a filter press through a pump, and the supernatant liquid is collected for use.

Optimum stirring velocity for metal leaching of electroplating sludge
To study the effect of different stirring velocities in the 10 m³ extraction tank on the leaching of heavy metals in electroplating sludge, five groups of comparative experiments with different stirring velocities were designed, as shown in Table 2.
Step 1: The sludge is crushed and collected for leaching.
Step 2: All circuits, instruments, and equipment are checked for faults.
Step 3: The biological sulfuric acid produced by the membrane bioreactor is pumped into the 10 m³ extraction tanks through an acid-resistant pump.
Step 4: The stirring system is turned on, and the crushed electroplating sludge is slowly added at a pulp density (w/v) of 8%. Stirring is performed for 3 min at the experimental stirring velocity specified in Table 2. Step 5: Every 30 minutes, a sample is collected with a 10 ml pipette and centrifuged, and then the centrifuged supernatant is collected and diluted in a test tube. ICP-AES is used to analyse the heavy metal concentration in the leachate and calculate the leaching rate of each metal.
Step 6: Solid-liquid separation. After stirring, the sludge in the stirring tank is passed into a filter press through a pump, and the supernatant liquid is collected for use. Solid-liquid separation. After stirring, the sludge in the stirring tank is passed into a filter press through a pump, and the supernatant liquid is collected for use.

Metal enrichment in electroplating sludge
According to the concentration analysis of heavy metal ions in the leaching supernatant of electroplating sludge by ICP-AES, at 8% pulp density, the target metal concentration in the leaching solution was low after one round of leaching (the Ni + /Cu 2+ /Zn 2+ concentrations were 1401 ppm/1959 ppm/2020 ppm, respectively). It was quite difficult to recover the target metals. To increase the concentrations of the target metals in the leaching solution, a cycle experiment was designed as follows: Step 1: The sludge is crushed and collected for leaching. Step 2: All circuits, instruments, and equipment are checked for faults. Step 3: The biological sulfuric acid produced by the membrane bioreactor is pumped into the 10 m³ extraction tanks through an acid-resistant pump.
Step 4: The stirring system is turned on, and the crushed electroplating sludge is slowly added at a pulp density (w/v) of 8%, stirring velocity of 100 rpm, and leaching time of 150 min.
Step 5: A sample is collected with a 10 ml pipette and centrifuged, and then the centrifuged supernatant is collected and diluted in a test tube. ICP-AES is used to analyse the heavy metal concentration in the leachate and calculate the leaching rate of each metal.
After stirring, the sludge in the stirring tank is passed into a filter press through a pump, and the supernatant liquid is collected for use. Step 7: The separated liquid is returned to the membrane bioreactor through a pump for microbial acid production.
Steps 1-7 were repeated. The sample after each cycle of solid-liquid separation was marked as a raw material for later experiments.
The flow chart of the 10 m³ membrane bioreactor target metal cycle enrichment experiment is shown in Fig. 3: Criteria for the end of the cycle enrichment experiments: One criteria: When the cell number of microorganisms in the MBR drops sharply (i.e., when the cell number is less than 10 9 cells/ml) or when the pH of the MBR fluctuates greatly (i.e., when the pH in the MBR is higher than 1.0), the acid production and cycle enrichment experiments should be stopped. The other criteria: When the leaching rate of the target metal in the leaching experiment in the stirred tank is less than 100%, the acid production and cycle enrichment experiments should be stopped.

Morphological structure and target metal content of electroplating sludge
Digestion analysis of target metal content in electroplating sludge.
The sample was digested, and it was found that the electroplating sludge mainly contained Ni, Cu and Zn and that the contents of the target metals were 1.75% (Ni), 2.45% (Cu), and 2.53% (Zn), respectively.  [22,23], areas with a small electron scattering angle are bright, indicating that the atom density is low or there are pores in this area. Areas with a large electron scattering angle are dark, indicating that the atom density is higher in this region; the darkest part indicates that the atom density is the highest in this area. Combined with the SEM-EDS data for the electroplated sludge sample in Fig. 6, it can be seen that the contents of the target metals Ni and Cu in the sample were 1.67% and 2.23%, respectively, which is consistent with the XRF results. Zn was not detected in the EDS test, which may be due to the poor uniformity of the target metal distribution in the electroplating sludge and the lack of Zn in the area of the limited energy spectrum sweep.

Optimum pulp density for metal leaching of electroplating sludge
The variation in the leaching rate of the target metal ions over time in the bioleaching of electroplating sludge with different pulp densities is shown in Fig. 7. It can be seen from Fig. 7 that the leaching rates of the target metal ions over time in electroplating sludge with different pulp densities show similar trends. In the first 30 minutes of the electroplating sludge leaching reaction, the leaching rate of the target metal ions in the sludge increased rapidly, and then the leaching rate slowed. When the leaching reaction time was 150 min, the leaching rate showed a maximum value.
In addition, the target metals could be completely leached when the pulp density was 4%, 6%, and 8%; when the pulp density was 10% or 12%, the target metals could not be fully leached. These results show that the pulp density of bioleaching electroplating sludge has a significant effect on the leaching rate of heavy metal ions in the sludge. The smaller the pulp density was, the higher the leaching rate of heavy metal ions was at a given time. The above results indicate that under the studied conditions, the optimal pulp density of bioleaching electroplating sludge is 8% under the premise of increasing the treatment amount of electroplating sludge as much as possible.

Optimum leaching time for metal leaching of electroplating sludge
The variation in the leaching rate of the target metal ions over time in bioleaching electroplating sludge with different leaching times is shown in Fig. 8. It can be seen from Fig. 8 that the leaching rates of the target metal ions over time in electroplating sludge with different leaching times show similar trends. In the first 30 minutes of the electroplating sludge leaching reaction, the leaching rates of the target metal ions in the sludge increased rapidly, and then the leaching rates of the target metal ions slowed. When the leaching reaction time was 150 min, the leaching rates showed a maximum value. The above results indicate that under these conditions, the optimal extraction time for bioleaching electroplating sludge is 150 min.

Optimum stirring velocity for metal leaching of electroplating sludge
The variation in the leaching rates of the target metal ions over time in bioleaching electroplating sludge with different stirring velocities is shown in Fig. 9.
It can be seen from Fig. 9  Therefore, the pH slightly fluctuated, but the fluctuation was small. The fluctuation in the cell concentration of the leaching strain was mainly caused by the slow changes in the environment, including the pH of the leaching solution (1.3-1.5) being higher than the pH (1.0) in the membrane bioreactor and the temperature of the leaching solution (room temperature) being lower than that of the membrane reactor (33 ℃). During the cycle process, we found that when the cycle experiment was carried out for the 8 th , 9 th , and 10 th cycles, the pH in the membrane bioreactor showed a gradual increase, which was inconsistent with the pH variation during cycles 1 th -7 th . Accordingly, the cell concentration of the leaching strain in the membrane bioreactor showed an irreversible decreasing trend at the 8 th , 9 th , and 10 th cycles. These results heralded the end of the cycle enrichment experiment.
Based on the analysis of the above data, we performed leaching toxicity analysis of the electroplating sludge residues after 10 heavy metal cycle enrichment experiments. The results are shown in Fig. 12. It can be seen from Fig. 12 that as the number of heavy metal enrichment cycles increased, the toxic leaching concentrations of the target metals (Ni + , Cu 2+ and Zn 2+ ) gradually approached the national standard limits. Moreover, for the electroplating sludge residues after the 10 th cycle of enrichment, the toxic leaching value was higher than the national standard limit. Therefore, based on the above analysis, the maximum number of cycles for the enrichment of heavy metals in electroplating sludge is 10. After 10 cycles, the enrichment concentrations of Ni + , Cu 2+ and Zn 2+ were 13.867 g/L, 18.118 g/L and 21.075 g/L, respectively.

The mechanism analysis of heavy metals release in electroplating sludge
Consistent with our previous bioleaching experiments at the same experimental conditions [16], the acidophilic microorganism did not directly affect the metal removal efficiencies, but contributed to accelerate the pH decrease and Fe(II/III) ion conversion. The schematic diagram of acidophilic microorganism sulfur metabolic pathway ( Fig. 13) depicting the mechanism of H + release (Equations (1)-(3) ). The mechanism of heavy metals release in electroplating sludge can be describe through the Equations (4)

Conclusions
In this research, we concluded that the MBR maintained a high cell