Abstract
Staphylococcus aureus (S. aureus) is an opportunistic pathogen of the gram-positive variety, known to cause a range of severe infections including cellulitis, pneumonia, osteomyelitis, endocarditis, and sepsis. These infections are associated with significant morbidity and mortality rates in both hospital and community settings. Therefore, it is important to remove S. aureus from the aqueous solution. The present study employed response surface methodology as an effective strategy to optimize the removal of S. aureus through the electro-Fenton (EF), UV/H2O2, and combination EF-UV/H2O2 processes. Under the optimized conditions, the maximum removal efficiency in the EF process of 48.5%, UV/H2O2 process of 36.2%, and combination EF-UV/H2O2 process of 100%. The optimum condition for removal efficiency using combination EF-UV/H2O2 process of 100% was attained at S. aureus concentration of 5 × 106 CFU mL−1, current density of 8.0 mA cm−2, H2O2 dosage of 170 µL L−1, and 2 lamps UV during 7.0 min. Both the production of •OH in the EF and UV/H2O2 process and the additive oxidation effect of UV/H2O2 are the main reasons for the better performance of combination EF-UV/H2O2 process. The results indicate that the EF-UV/H2O2 process is highly promising and environmentally sustainable method for treating wastewater samples contaminated with S. aureus.
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Introduction
The utilization of recycled wastewater is being contemplated as a strategic remedy in numerous regions or countries facing water scarcity (Yong et al. 2023). Nevertheless, the existence of health risks stemming from biological hazards (such as pathogens) and chemical hazards (like micro-pollutants) in the original water sources, reclaimed wastewater, water used for landscaping, and the final drinking water, poses a growing challenge for public health and the ecological/environmental aspects (Li et al. 2023a; Mgidlana et al. 2023). Bacterial contamination represents a distinct category of water pollution. When pathogens contaminate drinking water, they can lead to infections in the gastrointestinal system, resulting in complications such as diarrhea and cholera. Every year, a significant number of children and elderly individuals lose their lives due to water contaminated with Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus), two bacteria that have developed resistance to commonly used antimicrobial agents (Ausbacher et al. 2023). S. aureus, a type of Gram-positive bacterium, is widely recognized as a prominent pathogen associated with severe cases of foodborne and waterborn diseases and has consistently posed a significant challenge in various human infections, especially due to its resistance to antibiotics (Wan Omar et al. 2023; Zhang et al. 2023).
These concerns pose a significant challenge for public health and environmental aspects. Therefore controlling the presence of pathogenic bacteria in water is crucial to avoid potential health risks related to water contamination (Aziz et al. 2013; Iqbal et al. 2019). Disinfection is the final step in wastewater treatment plants aimed at reducing the presence of pathogenic bacteria. Nonetheless, pathogenic bacteria can evade the disinfection processes and consequently be present in treated wastewater, ultimately entering the aquatic environment (Yang et al. 2023; Abbas et al. 2018). This poses a significant concern for human health since treated wastewater, depending on its subsequent utilization, can serve as a potential source of waterborne diseases (Sabatino et al. 2022; Kousar et al. 2022). Traditional water treatment methods have proven to be ineffective in preventing and eliminating pathogenic microorganisms from water (Bokhari et al. 2022). Consequently, there is a pressing need for innovative materials that possess antimicrobial properties in order to enhance the quality of contaminated water (Mphuthi et al. 2023; Liu et al. 2023).
Advanced oxidation processes (AOPs), which operate at or near ambient temperature, include Fenton, photo-Fenton, electro-Fenton, UV-based processes, and photocatalytic redox processes. These methods are specifically employed for water and wastewater treatment. The main objective of AOPs is to generate highly reactive free radicals (Mansour et al. 2023; Babu Ponnusami et al. 2023). Hydroxyl radicals (•OH) are extremely effective in the destruction of organic molecules due to their high standard potential (E = 2.8 vs. SHE, or "Standard Hydrogen electrode "), as well as their react quickly and non-selective (Jiang et al. 2022; Thakur et al. 2023).
The Fenton reaction, being a typical of AOPs, effectively oxidizes organic pollutants by utilizing •OH generated from the reaction between H2O2 and Fe2+. In the typical procedure, the Fenton reaction operates through a cyclic reaction involving H2O2. As represented in reaction (1) and (2), under acidic conditions reaction (1), Fe2+ initially reacts with H2O2 to produce •OH and Fe3+. Following this, the generated Fe3+ reacts with H2O2 to form H2O2 radicals and regenerate Fe2+ reaction. (2) (Qi et al. 2023; Dai et al. 2023).
However, traditional Fenton technology suffers from certain drawbacks such as the need for chemical dosing, generation of substantial sludge, lengthy treatment duration, and difficulties in control, which restrict its widespread implementation. Additionally, the pH tolerance range of the conventional Fenton process is limited, as it can only operate efficiently under acidic conditions.
The electro-Fenton (EF) process has gained significant attention due to its numerous advantages, including high efficiency, minimal land usage, user-friendly operation, and environmental friendliness. This process addresses several drawbacks of the conventional Fenton process, such as mitigating the risks associated with the storage and transportation of Ferric sulfate, as well as reducing the formation of excessive iron sludge (Li et al. 2023b). UV-based AOPs represent a promising category of advanced treatment technologies that have been developed with the aim of eliminating pollutants and enhancing the quality of treated water and wastewater. UV-AOPs employ a collaborative approach involving UV light, chemical oxidants (such as H2O2, persulfate, and chlorine), or photocatalysts to efficiently oxidize pollutants (Wang et al. 2018). UV irradiation combined with H2O2 has garnered significant attention from environmentalists among various AOPs due to its remarkable ability to effectively eliminate pollutants present in treated effluents (Salim et al. 2022). Enhanced Fenton-based and UV processes, including the innovative photoelectro-Fenton (PEF) process, have emerged to minimize or even eliminate sludge production. These advanced methods are specifically engineered to operate efficiently at elevated pH levels, even under near-neutral conditions (Brillas 2022; Arshad et al. 2020).
This research focuses on the removal of S. aureus investigation and optimization of various parameters in the EF process, with the aim of studying. Initially, key factors such as the initial concentration of S. aureus, current density, and H2O2 dosage are investigated using response surface methodology (RSM). Subsequently, the removal of S. aureus is analyzed by employing the UV/H2O2 process. Finally, the removal process is thoroughly examined and optimized through the application of the PEF process.
Experimental and methods
Chemicals
Analytical grade sodium hydroxide (NaOH, purity: ≥ 98.0%), Hydrochloric acid (HCl, purity: 37.0%), sodium sulfate (Na2SO4, purity: ≥ 99.0%), acetone (C3H6O, purity: ≥ 99.0%), potassium chloride (KCl, purity: ≥ 99.0%), hydrogen peroxide (H2O2, purity: 30.0%), chloroform (CHCl3, purity: ≥ 99.0%), isoamyl alcohol ((CH3)2CHCH2CH2OH, purity: ≥ 95.0%), ethanol (C2H6O, purity: ≥ 70%.), isopropyl alcohol (C3H8O, purity: ≥ 99.5%), lactose, glucose, and sucrose were prepared from Merck® company. Moreover, safranin, triple sugar iron agar, MacConkey agar, trypticase soy agar with 5% sheep blood, plate count agar was prepared from Merck company.
Electrochemical reactor
The EF experiments were carried out under batch conditions using an electrochemical reactor. An iron plate with 99.5% purity and dimensions of 60 × 6 × 1 mm was utilized as both the anode and cathode electrodes, which were positioned vertically and parallel to each other. The electrodes were of the same size, with a constant inter-distance of 3.0 cm within the electrochemical reactor. The reaction time and pH were kept constant at 15 min and 6.5, respectively, throughout the EF process. The removal efficiency of S. aureus and the electrical energy consumption (EEC) of the process were determined using Eq. (3) and (4), respectively.
where CS.aureus-0 and CS.aureus-t (CFU mL−1) denote the S. aureus concentration before EF process and time t, respectively.
where U, I, t, and V are applied voltage (U), applied electrical current (A), reaction time (h) and volume of sample (L), respectively.
The UV/H2O2 process was conducted by introducing synthetic wastewater and H2O2 into a 250 ml UV/H2O2 reactor in UV cabinet. To ensure the temperature and radiation output of the lamps were balanced, a period of 30 min was allocated before each experimental run for the lamps (consisting of 1 to 3 lamps) to be switched on and reach equilibrium.
Previous experiments have demonstrated that a period of continuous exposure to radiation lasting 10 min is adequate to achieve a state of equilibrium. Therefore, for the purpose of this experiment, a fixed reaction time of 10 min was established. After each sampling in EF, UV/H2O2, and PEF, the samples were promptly quenched by adding Na2S2O3. This addition serves to eliminate any excess H2O2 and halt the reaction. To minimize errors, performed three repetitions of all experiments and presented the average value for each run.
Statistical analysis and modeling using RSM
Response Surface Methodology (RSM) was utilized to develop a statistical model that evaluates and optimizes the impact of operating parameters as independent variables on the removal efficiency of the EF process, which serves as the response variable. For the current study, three independent variables were chosen as A: S. aureus concentration (X1), B: current density (X2), C: H2O2 dosage (X3). The effect of variables was investigated at five different coded levels of − α, − 1, 0, + 1, and + α, in which the number of α is equal to 1.68. The S. aureus removal efficiency was selected as response. Table 1 summarizes the actual and coded values of the variables.
The obtained data from the experiments were analyzed using Design Expert 11.0 software. The analysis of variance (ANOVA) was investigated. The results obtained indicated that the experimental data showed the highest level of agreement with the second-order polynomial model presented below (Eq. (5)) (Wang et al. 2022):
where Y, Xi, and Xj denote the predicted response and the input variables of i and j, respectively. Moreover, the parameters β0, βi, βii, and βij denote the regression constants for intercept, linear, quadratic, and interaction coefficients, respectively. Also, ε and n denote the error, and the number of variables studied. The experiments were conducted in a random manner, and the designed experiments along with the corresponding values of the model responses are presented in Table 2.
Results and discussion
Statistical analysis and model fitting
To evaluate the appropriateness and significance of the constructed model, we conducted an ANOVA, utilizing the statistical indicators of F value and p value. The findings from this analysis are presented in Table 3.
Table 3 shows that the developed model achieved an F value of 209.5 for the removal efficiency of S. aureus. Additionally, the p values associated with the developed model was found to be less than 0.0001. Therefore, it can be concluded that the fitted model was statistically significant (p value < 0.05) at a 95% confidence level (CI).
The removal efficiency of S. aureus, as influenced by the independent variables, is represented by the coded form in Eq. (6), which illustrates the relationship between the independent variables and response.
X1, X2, and X3 are the code values of S. aureus concentration, current density, and the H2O2 dosage, respectively. Moreover, X32 is the quadratics effects of H2O2 dosage. The presence of negative or positive signs associated with the terms in the equation signifies the respective synergistic and antagonistic effects of each factor on the removal efficiency of S. aureus in the EF process, which serves as the dependent variable. The quadratic model proposed for the removal efficiency of S. aureus demonstrated that the developed regression model was the most suitable fit for the examined parameters.
The coefficient of determination (R2) for the removal efficiencies, as predicted by the developed model (S. aureus removal efficiency), was 0.9824. This value indicates the extent to which the expected and observed values of removal efficiency align with each other. The proposed model achieved an adjusted R2 (Adj.R2) value of 0.9777 for the removal efficiency. Additionally, the predicted R2 (Pred.R2) value was determined to be 0.9663.
Based on the discrepancy between Adj.R2 and Pred.R2 being below 0.20, it can be concluded that the developed model accurately predicts the removal efficiency of the proposed EF process. Additionally, this prediction demonstrates a significant level of agreement with the actual values, indicating a high level of satisfaction. The adequate precision (AP) value, expressed as the signal-to-noise ratio, achieved a value of 51.7 for the S. aureus removal efficiency. As the obtained value exceed 4, it is confirmed that the proposed model effectively explores the design space with great precision and dependability. The calculated p-value of 0.0881, indicating the lack of fit in the proposed regression model for removal efficiency, exceeds the significance level of 0.05. This suggests that the relationship between the input variables and the model response is deemed appropriate, affirming the significance of the developed model.
Diagnostic diagrams were plotted in order to assess the normal distribution and adequacy of the data, as demonstrated in Fig. 1.
Based on Fig. 1a, it is clear that there is a satisfactory agreement between the predicted values and the experimentally obtained values for the S. aureus removal efficiency of the proposed model. This agreement confirms the competence of the model in accurately predicting the response values. Furthermore, the normal distribution of the model, as depicted in Fig. 1b, indicates that the externally observed residual points align closely along a nearly straight line.
Pareto analysis
The Pareto analysis was performed in order to examine the influential factors' contribution to the response variable for the predicted model. To evaluate the significance of operating parameters on the S. aureus efficiency removal, Pareto analysis was conducted. The result, presented in Fig. 2, clearly demonstrate the operating parameter with the most substantial influence on the response. As can be seen, the impact of H2O2 dosage and the initial concentration of S. aureus on the removal efficiency are evident. Furthermore, the quadratic effect of H2O2 dosage (X32) influenced the removal efficiency.
Effect of parameters
Effect of S. aurous initial concentration
The effect of S. aureus initial concentration on the removal efficiency using the EF process was investigated at various concentrations ranging from 1 × 104–10 × 106 CFU mL−1. The simultaneous effect of current density and initial S. aureus concentrations on the removal efficiency using EF process were investigated and displayed in the contour plot of Fig. 3.
The observation revealed a decrease in removal efficiency as the initial concentration of S. aureus increased under constant conditions, including a current density of 7.5 mA cm−2, H2O2 dosage of 110 µL L−1, reaction time of 15 min, and pH of 6.5. Specifically, the removal efficiency of S. aureus decreased from 46.2% to 31.5% when the initial concentration increased from 1 × 104 to 10 × 106 CFU mL−1.
It is important to note that under constant conditions, the generation of •OH remains consistent. However, as the S. aureus concentration increases to higher levels, the proportion of •OH compared to the S. aureus count decreases. The decrease in the proportion of hydroxyl radicals (•OH) subsequently leads to a reduction in the efficiency of removing S. aureus.
Effect of current density
Effect of the current density on S. aureus removal efficiency was investigated in the range of 3–12 mA m−2 during reaction time of 15 min, S. aureus concentration of 5 × 106 CFU mL−1, and pH of 6.5. The obtained experimental results were illustrated by contour plot in Fig. 3.
It was observed that the S. aureus removal efficiency is increased with increasing the current density. The removal efficiency of S. aureus was increased from 33.8% to 44.5% when the current density increased from 3 to 12 mA cm−2. It is evident that the use of a higher current density to the electrochemical reactor leads to an increase in the rate of Fe+2 ion generations, as indicated by reaction (7). The acquired response ascribed to the accelerating of Fe2+ ion generation, which resulted in the progression of Fenton reaction and generation of more •OH (Amarzadeh et al. 2023; Javed et al. 2021).
Effect of H2O2 dosage
According to literature surveys, the removal efficiency is significantly influenced by the dosage of H2O2 used as an oxidizing agent in EF process. Therefore, the present study conducted a thorough investigation to identify the optimal dosage of H2O2 within the range of 20–200 µL L−1. Effect of H2O2 dosage for S. aureus removal efficiency was investigated under center point conditions, including a current density of 7.5 mA m−2, S. aureus concentration of 5 × 106 CFU mL−1, pH of 6.5, and a reaction time of 15 min during the EF process. The results obtained are depicted in Fig. 4.
The removal efficiency of S. aureus was increased from 22.7% to 42.6% when the H2O2 dosage increased from 20 to 170 µL L−1. The increase in removal efficiency observed when the dosage of H2O2 was increasing from 20 to 170 µL L−1 can be attributed to the acceleration of the Fenton reaction and the generation of a greater amount of •OH (reactions 5 and 6). However, when the dosage of H2O2 exceeds 170 µL L−1, the removal efficiency of S. aureus was decreases.
Similar studies have attributed the reason for this phenomenon to the following reactions, as demonstrated in side reactions (reactions 10–2), the observed phenomenon may be attributed to an excessive quantity of H2O2 ranging from 170 to 200 µL L−1, leading to the consuming of free •OH. HO2•, acting as a weak oxidizing agent, deactivates the hydroxyl radicals, as described. Furthermore, in high dosage of H2O2, the interaction between •OH intensifies, leading to a greater depletion of oxidizing agents used for the removal efficiency (Iqbal et al. 2023; Rahman et al. 2023).
Optimal operating condition for EF process
The optimization of analytical procedures has garnered significant attention with the use of RSM in comparison to the classical approach known as One factor at a time (OFAT). This is primarily because RSM has the ability to yield valuable information with a limited number of designed experiments. In order to attain the highest efficiency in removing S. aureus (response), the optimization of operating conditions was carried out to identify the optimal values for these parameters. The process of optimization involved the utilization of a numerical technique integrated within the Design-Expert software. The variables were set with the objective of achieving values within a specified range, while the primary focus was placed on maximizing S. aureus removal efficiency. Based on the outcome findings it was found that the best S. aureus removal efficiency of 48.5% was achieved at S. aureus concentration of 5 × 106 CFU mL−1, H2O2 dosage amount of 170 µL L−1, pH 6.5, reaction time 15 min and current density 8.0 mA.cm−2.
UV/H2O2 process
The optimization of light intensity emitted by 1 to 3 UV lamps was investigated during the UV/H2O2 process under the following conditions: S. aureus concentration of 5 × 106 CFU mL−1, H2O2 dosage of 170 µL L−1, and pH 6.5. These conditions were selected based on the optimized values in the EF process. Samples were manually collected at reaction time 0 and 10 min of each experiment. The results obtained are depicted in the Fig. 5.
The results showed that the removal efficiencies increased from 19.1%, 36.2%, and 41.4% by increasing the number of lamps: (Electrical power) from 1 lamp (6 W), 2 lamps (12 W), and 3 lamps (18 W). On the other hand, increasing the number lamps from 1 to 3 lamps increased the electrical energy consumption during UV/H2O2 process. Increased levels of S. aureus removal efficiency was observed as the intensity of radiation escalated, primarily attributed to the generation of •OH via the photolysis of the oxidant (Vitale et al. 2023; Bhatti et al. 2020).
de Souza et al. and Srithep et al. reported in her study that the UV radiation in wavelength of 254 nm breaks the chemical bond of –O–O– in H2O2 and creates •OH, according to reaction (5) (de Souza et al. 2023; Srithep et al. 2017; Bokhari et al. 2020). The generation of •OH enhances the effectiveness of the removal process (Zhu et al. 2023). Based on the obtained results, the removal efficiency of using 2 lamps (36.2%) is very close to that of using 3 lamps (41.5%). Therefore, 2 lamps (12 W), is selected as the optimal conditions.
Combination EF- UV/H2O2 process
The combined effect of the EF and UV/H2O2 processes was investigated under the specified conditions: a concentration of S. aureus at 5 × 106 CFU mL−1, dosage of 170 µL L−1 of H2O2, pH of 6.5, current density of 8.0 mA cm−2, and 2 lamps UV. These conditions were chosen based on optimized values in the EF and UV/H2O2 process. The samples were conducted at reaction times ranging from 1 to 10 min for each experiment. Figure 6 illustrates the results obtained.
The results clearly indicate that the removal efficiencies of S. aureus increased with the extension of reaction time. Specifically, the removal efficiencies improved from 35.2%, 58.1%, 83.3%, 97.4%, 99.8%, to 100% when the reaction time was increased to 1, 2, 3, 5, 7, and 10 min, respectively. According to obtained result, the combination of EF and UV/H2O2 process was more efficient for S. aureus removal efficiency than EF and UV/H2O2 alone. The increase in removal efficiency may be attributed to the enhancement of the Fenton reaction rate through simultaneous UV irradiation. This phenomenon assists in the regeneration of the required Fe2+ species (reaction 13).
The obtained results show that the coupling of EF process with UV/H2O2 process can be a promising approach for the removal of S. aureus.
Conclusions
In this study, substantial efforts were undertaken to remove S. aureus from contaminated water. The present research aimed to optimize the operational parameters of the EF, UV/H2O2, and combination processes using response surface methodology (RSM) based on central composite design (CCD). These measures were implemented to improve the efficacy of S. aureus removal. According to our findings, the combination of the EF and UV/H2O2 process was more efficient for removal efficiency than EF process (48.5%) and UV/H2O2 process (36.2%) alone. Furthermore, the analysis of variance showed the adequacy of the correspondence of the experimental data to the theoretically determined ones, i.e., the obtained mathematical model. The significant improvement of removal efficiency in combination of the EF and UV/H2O2 process highlighted the synergistic effect between EF and UV/H2O2 process. The results show that •OH was the main oxidant species contributed in removal efficiency of S. aureus under the EF and UV/H2O2, and combination process. The findings indicate that the combination EF and UV/H2O2 has significant potential as a highly efficient and environmentally friendly method for the removal of S. aureus.
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Acknowledgements
The authors would like to express their appreciation to Kerman University of Medical Sciences [Grant Number 402000584] for supporting the current work.
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This work received a grant from the Kerman University of Medical Sciences [Grant Number 402000584].
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Dokhani, A., kheirkhah, B., Kalantar-Neyestanaki, D. et al. Removal of Staphylococcus aureus using electro-fenton, UV/H2O2, and combination of electro-fenton and UV/H2O2 processes; optimization of operational parameters. Appl Water Sci 14, 100 (2024). https://doi.org/10.1007/s13201-024-02151-0
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DOI: https://doi.org/10.1007/s13201-024-02151-0