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Article

Utilization of Cheese Whey for Energy Generation in Microbial Fuel Cells: Performance Evaluation and Metagenomic Analysis

by
Rojas-Flores Segundo
1,2,*,
Cabanillas-Chirinos Luis
1,2,
Nélida Milly Otiniano
1,2,
Magaly De La Cruz-Noriega
1,2 and
Moises Gallozzo-Cardenas
3
1
Institutos y Centros de Investigación de la Universidad Cesar Vallejo, Universidad Cesar Vallejo, Trujillo 13001, Peru
2
Renewable Resources Nanotech Group, Universidad Cesar Vallejo, Trujillo 13001, Peru
3
Facultad de Ciencias de la Salud, Universidad César Vallejo, Trujillo 13001, Peru
*
Author to whom correspondence should be addressed.
Fermentation 2025, 11(4), 176; https://doi.org/10.3390/fermentation11040176
Submission received: 20 February 2025 / Revised: 11 March 2025 / Accepted: 21 March 2025 / Published: 26 March 2025
(This article belongs to the Special Issue 10th Anniversary of Fermentation: Feature Papers in Section "Industrial Fermentation")
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Versions Notes

Abstract

:
The dairy industry generates large volumes of whey as a byproduct of cheese production, with a high organic load. Its untreated discharge contaminates water bodies, reduces dissolved oxygen, and damages aquatic ecosystems. In Peru, especially in the rural areas of the Andes, thousands of tons of industrial dairy waste are produced annually, representing an environmental and economic challenge. The lack of sustainable technologies for its management drives the need for innovative solutions, such as microbial fuel cells (MFCs), which combine waste treatment with renewable energy generation. This research uses MFC technology with whey as a substrate to observe its potential to generate electrical energy and treat contaminants. Three liters of whey from a dairy company in Trujillo, Peru, were used and stored at 10 °C. Each MFC contained 800 mL of whey and employed activated carbon as the anode and zinc as the cathode. A maximum voltage of 0.867 ± 0.059 V was reached, with a maximum current of 4.114 ± 0.239 mA recorded on the 11th day. The maximum power density was 1.585 ± 0.061 mW/cm2, with a current density of 4.448 A/cm2, and the internal resistance of the MFCs was 16.847 ± 0.911 Ω. The initial pH of the whey was approximately 3.0, increasing to 4.135 ± 0.264 on the 11th day, and the electrical conductivity increased from 19.101 ± 1.025 mS/cm on the first day to 170.062 ± 9.511 mS/cm on the 11th day. The oxidation-reduction potential (ORP) increased to 104.287 ± 4.058 mV at the peak of electricity generation (day 11). Additionally, a 70% reduction in chemical oxygen demand (COD) was achieved, dropping from 4650.52 ± 10.54 mg/L to 1400.64 ± 23.25 mg/L on the last day. Metagenomic analysis identified two dominant bacterial phyla: Bacteroidota at 48.47% and Proteobacteria at 29.83%. The most abundant families were Bacteroidaceae (38.58%) and Acetobacteraceae (33.39%). The study validates the potential of MFCs to transform whey into an energy resource, aligning with sustainability and circular economy goals, especially in regions with high dairy production, like Peru.

1. Introduction

The dairy industry faces several environmental challenges stemming from the generation and management of waste during its production processes [1]. The primary waste is whey (a byproduct of cheese production), which has a very high organic load, representing a significant risk to rivers and water bodies if discharged without proper treatment [2,3]. During the transformation of milk to produce cheese, between 8 and 10 L are used to obtain one kilogram of cheese, generating large volumes of byproducts (such as whey) that, if not reused or treated, represent a significant waste [4,5]. It is estimated that global whey production is approximately 180 million tons annually, which may vary depending on the type of milk and the cheese-making process [6]. It has been reported that whey contains approximately 4.5% lactose, 0.8% protein, and 0.5% lipids [7]. Additionally, it contains minerals and vitamins that make it a valuable byproduct [8]. Whey is used in a wide range of food products, including non-alcoholic beverages, fermented drinks, protein concentrates, edible films, and as a source of lactose to sweeten foods [9]. It also produces ethanol, organic acids, and enzymes [10]. Improper management of whey can hurt the environment [11]. In some regions, whey is discharged into rivers and streams, which can cause significant damage to aquatic ecosystems [12]. Implementing clean technologies and sustainable practices is crucial to mitigate these impacts.
Converting whey into energy can be more economical than other waste management methods since it utilizes a resource that would otherwise be discarded [13]. This can result in significant savings for dairy companies [11]. In this regard, microbial fuel cells (MFCs) represent a promising solution that combines renewable energy generation with the purification of organic waste [14]. Their application can transform waste into an energy resource, contributing to reducing pollutants and offering a sustainable alternative to conventional technologies, aligning with the global challenges of climate change and water scarcity [15]. Although the technology is currently emerging, its potential to transform environmental and energy management makes it a crucial area of research and innovation [16]. MFCs consist of several key parts that enable the conversion of organic matter into electrical energy [17]. The anode is where microorganisms oxidize the present substrate, releasing electrons [18]. These electrons are transferred through an external circuit to the cathode, where a reduction, typically of oxygen, occurs [19]. Between the anode and the cathode is the proton exchange membrane (PEM), which allows the passage of protons while keeping the two chambers separated [18,19]. Some studies have reported the use of organic waste as substrates. For example, Aliyu et al. (2024) used fruit waste as substrates in their MFCs, achieving voltage peaks of 0.650 V, operating at an average pH of 7.1 ± 0.23 using carbon electrodes on a metal base to increase conductivity [20]. A power density of 18,228 μW/m2 at a current density of 244 mA/m2 was observed using grape waste with winery wastewater [21]. The agro-industrial wastewater was used as a substrate in their MFCs, achieving maximum power density values of 1350.6 ± 125 mW/m2 and maximum voltage values of 0.76 V [22].
In this context, cheese production in rural areas of Peru generates several wastes, primarily whey, which, if not properly managed, can cause environmental and economic problems [23]. Whey, the main byproduct of cheese making, contains a high organic load (lactose and proteins) [24]. If discharged into rivers or soils without treatment, it causes pollution and decreases oxygen levels in the water, affecting aquatic fauna [25]. The agricultural sector reported that Peru generates much solid waste [23]. In 2014, approximately 1,897,302 tons of solid waste were generated, of which 1,869,618 tons were non-hazardous waste and 22,246 tons were hazardous waste [26]. Similarly, in 2018, the Puno region produced 10,255 tons of milk; from this amount, approximately 15,192.54 tons of liquid effluents were generated during cheese processing, including sweet and salty whey [27]. It is known that for every 100 kg of milk processed to make fresh cheese, approximately 10 kg of cheese and 90 kg of whey are obtained [28]. Dairy waste has a complex composition with high levels of fats, proteins, and lactose, which can negatively affect the performance of MFCs [23,25]. The degradation of these compounds can produce by-products that inhibit microbial activity and reduce the efficiency of organic matter conversion into electricity [19]. One of the main limitations of MFCs is their efficiency [17]. To improve the treatment of dairy waste, it is common to perform pretreatment that reduces the organic load and eliminates inhibitory compounds [25]. This additional process can increase the costs and complexity of the system [20]. Moreover, treating dairy waste can generate by-products, such as whey, which require proper handling to avoid negative environmental impacts [16]. In the literature, the use of waste from the cheese processing industry in rural areas of Peru as a substrate in microbial fuel cells has not been observed. For this reason, utilizing cheese waste should be beneficial for energy production, as it would take advantage of nutrient-rich organic compounds that can be transformed into electricity in a novel approach.
The study aims to analyze the impact on the performance of single-chamber microbial fuel cells (MFCs) using dairy industry waste as a substrate over 18 days. Parameters such as chemical oxygen demand, electric current, redox potential, internal resistance, power density, voltage, electrical conductivity, and current density will be monitored in the MFCs. The conversion of whey into electrical energy drives innovation in waste treatment and energy production technologies. This can lead to the development of new sustainable and efficient solutions; using whey as an energy source reduces the emission of greenhouse gases and other pollutants associated with fossil fuel combustion.

2. Materials and Methods

2.1. Obtaining Whey

The waste was obtained from the dairy company El Rosal, located in La Victoria, Santiago de Chuco, Trujillo, Peru. Three liters of whey were collected and stored in a container until use in the laboratory, where they were kept at 10 °C until used in the microbial fuel cells.

2.2. Design and Assembly of MFCs

The single-chamber microbial fuel cells (MFCs), with a volume of 100 mL, were purchased from Xin Tester in Shanghai, China. For the electrodes, activated carbon (AC) and zinc (Zn) were used as the anode and cathode, respectively, with areas of 20.50 cm2 and 15.50 cm2. Activated carbon was chosen because it has a porous structure that provides a large surface area, which enhances the adsorption of microorganisms and electron transfer, and zinc was selected for its suitable reduction potential, which facilitates electron transfer at the cathode [14,17,18]. The electrodes were connected using an external resistance of 100 Ω in the external circuit. The chambers were separated using Nafion (Nafion is known for its high ionic conductivity, which aids in the transport of protons from the anode to the cathode) as a proton exchange membrane (PEM). Figure 1 presents the schematic of the energy generation process from whey. Each MFC contained 800 mL of whey, and three single-chamber microbial fuel cells were used to obtain the average value and standard deviation for each measured parameter.

2.3. Evaluation of the Performance of MFCs

Voltage and electric current values were obtained using a multimeter (Prasek Premium PR-85), while pH levels were recorded using a pH meter (110 Oakton Series) over 18 days. Power density (PD) and current density (CD) were calculated using external resistances (Rext.) with values of 1.92 ± 0.11, 10.21 ± 1.31, 19.85 ± 3.42, 36.58 ± 4.58, 53.26 ± 6.72, 225.24 ± 11.35, 384.65 ± 32.16, 486.98 ± 41.53, 722.85 ± 69.32, and 1023.63 ± 84.32 Ω, applying the formulas PD = V2cell/(Rext.A) and CD = Vcell/(Rext.A), where Vcell represents the MFC voltage and A is the area [29]. An energy sensor (Vernier-±30 V and ±1000 mA) was employed to evaluate the resistance and power of the MFC. Data on electrical conductivity (EC), chemical oxygen demand (COD), and oxidation-reduction potential were also obtained using a digital multiparameter device. Table 1 shows the physicochemical parameters of the serum used in the research.

2.4. Metagenomic Analysis of Whey

A sterile sample of the substrate (whey) used in the MFCs was collected and transported in a cold chain to Ecobiotech Lab S.A.C. for subsequent analysis. Genomic DNA was extracted using the E.Z.N.A.® Soil DNA Kit (Omega Bio-Tek Inc., Norcross, GA, USA), following the manufacturer’s instructions. The DNA was then quantified and validated using a micro-volume spectrophotometer (EzDrop1000 Blue-Ray Biotech, Trujllo, Peru) and stored at −30 °C. Aliquots of the extracted DNA were analyzed at MR DNA (Molecular Research LP, Shallowater, TX, USA) through bacterial diversity genetic sequencing (16S rRNA sequencing) using the bTEFAP® Illumina Diversity Assay technology (DNA Gene Synthesis, Florida, USA). Diversity and abundance percentages were obtained using Mothur software (version 1.42.3.) and Excel Professional Plus 2019 [30].

3. Results

Figure 2 shows a typical growth behavior followed by a subsequent decline in the voltage production of microbial fuel cells, where the voltage starts from relatively low values (between 0.2 and 0.4 V) in the first days. This is due to bacteria adapting to the new substrate and beginning to form biofilm on the electrode [31]. On the eleventh day, a maximum voltage increase is observed, reaching 0.867 ± 0.059 V. This peak is associated with the high availability of organic compounds in the whey (lactose, proteins, etc.), which promote chemical reactions, generating a high potential difference between the electrodes [32]. The agricultural waste (rambutan, langsat, and mango) was used as a substrate in their MFCs, showing a maximum voltage of 0.180 V. They mentioned that electrochemical reactions decrease at the end of the monitoring stage due to the depletion of compounds. If the MFC is recharged, the voltages obtained will not be the same as those initially obtained [33]. Similarly, the potato waste was used as a substrate in their MFCs. They achieved a maximum voltage of 1.120 V. These authors point out that the degradation of lactose into organic acids (e.g., lactic, and acetic acid) through microbial hydrolysis rapidly increases voltage values as well as the generation of protons (H+) and their flow to the cathode, maintaining the electrochemical gradient [34].
Figure 3 shows the variation in electricity production in the MFCs over time. On day 1, a value of 0.125 ± 0.001 mA was recorded, indicating a delay in the current generation as the bacteria adapted to the substrate (whey) and begin to form biofilms on the anode [35]. Subsequently, the values progressively increased until the eleventh day, reaching a peak of 4.114 ± 0.239 mA. This increase reflects how the current intensified as the bacteria metabolized the lactose and other nutrients in the whey, releasing electrons that were transferred to the electrode. This peak represents the highest metabolic activity and the maximum efficiency in substrate conversion into electricity [36]. In the final stage, the current slightly decreased until day 18, recording a value of 3.213 ± 0.296 mA. The marine waste was used as a substrate in their MFCs, achieving current peaks of 0.108 µA. Their study mentions that a high organic load can prolong the exponential phase, although an excess could cause inhibition due to the accumulation of acids [37]. Similarly, the dairy industry wastewater was used in MFCs, generating an electric current of 14.3 µA. Their results indicate that the current stabilized when the consumption of whey and the generation of metabolic products (such as organic acids or CO2) reached an equilibrium. Additionally, they noted that limited substrate availability or the accumulation of inhibitors could restrict a further increase in electricity production [38].
In an MFC, microorganisms (primarily electrogenic bacteria) oxidize the organic compounds in cheese whey, releasing electrons and protons [13]. These electrons are transferred to the anode, while the protons migrate to the cathode through a proton exchange membrane [15]. At the cathode, electrons and protons combine with oxygen (or another electron acceptor) to form water [13]. The general oxidation-reduction equations are given by equations 1 and 2 (oxidation at the anode) and 3 (reduction at the cathode) [25,31,32].
Organic compound (e.g., lactose)→CO2 + H+ + e−
where the oxidation of lactose (C12H22O11) can be represented as:
C12H22O11 + 13H2O→12CO2 + 48H+ + 48e−
O2 + 4H+ + 4e−→2H2O
Lactose is hydrolyzed by microbial enzymes (β-galactosidase) into glucose and galactose, which are then metabolized through glycolysis and the Krebs cycle (aerobic pathway) or fermentation (anaerobic pathway) [36]. Several by-products can form during microbial degradation in microbial fuel cells (MFCs) utilizing cheese whey. Organic acids (acetic, lactic, and propionic), which result from the fermentation of carbohydrates in cheese whey, are typical by-products of this type of waste [31,37]. The accumulation of organic acids can decrease the system’s pH, inhibiting microbial activity and reducing the MFC’s efficiency. Additionally, high concentrations of alcohols can be toxic to electrogenic bacteria, affecting their ability to transfer electrons [38]. The accumulation of gases such as methane can displace oxygen in the cathode, reducing the efficiency of oxygen reduction [16]. The long-term stability of MFCs can be evaluated through accelerated and long-term stability studies similar to those used in the pharmaceutical industry [16]. These studies involve subjecting the system to controlled temperature and humidity conditions to observe its behavior and degradation over time [18]. To assess the long-term stability of MFCs utilizing cheese whey, it is essential to consider several factors such as pH parameters, control of by-product accumulation, electrode maintenance, and optimization of operating conditions [20,31].
Figure 4a shows the pH values recorded in the MFCs over 18 days. The pH is relatively low in the early days, around 3, suggesting an initially acidic environment, likely due to organic acids in the whey [39]. Over time, the pH gradually increases, reaching an optimal operating value of 4.135 ± 0.264. This increase could be due to the metabolic activity of the bacteria, which consume the organic acids and generate less acidic or more alkaline byproducts. The agro-industrials waste were used in MFCs, achieving voltage and current peaks of 0.608 ± 0.002 V and 0.40 ± 0.011 mA, respectively. In their study, the MFCs operated at an optimal pH of 6, and the researchers mentioned that this value is reached when metabolic activity stabilizes, and equilibrium is maintained in the internal environment of the fuel cells [40]. Similarly, researchers generated a maximum electric current of 10.126 ± 0.093 mA and a voltage of 0.816 ± 0.017 V using lemon waste as a substrate. Their study highlighted that operating at a pH of 3 ± 0.12 requires neutralizing the acids generated during fermentation to avoid bacterial inhibition [41]. Figure 4b shows the evolution of electrical conductivity in the MFCs using whey over 18 days. A progressive increase is observed from day 1 (19.101 ± 1.025 mS/cm) to reach a maximum value of approximately 170.062 ± 9.511 mS/cm on day 11. Subsequently, conductivity decreases until day 18 (145.585 ± 11.251 mS/cm). Scientific reports indicate that, in the initial phase, the degradation of lactose by bacteria releases ions (e.g., H+, HCO3−), which temporarily increases conductivity [42]. However, in the later days, the accumulation of acidic metabolites (e.g., lactic acid) can lower the pH and cause ion precipitation, decreasing conductivity [19]. Additionally, high electrical conductivity favors proton transfer to the cathode, reducing internal resistance and improving current generation. In contrast, low conductivity limits ionic flow, increasing ohmic losses and reducing energy efficiency [43]. The evolution of electrical conductivity in the MFCs suggests that ion concentration in the solution influences microbial activity and electricity production [43]. Optimizing MFC performance would be beneficial in controlling pH, replenishing nutrients, and preventing the accumulation of inhibitory products [44].
Figure 4c shows the chemical oxygen demand (COD) values obtained from monitoring the MFCs, showing a significant reduction in COD over time, indicating efficient biodegradation of whey by the microbial community [45]. The initial COD is 4650.52 ± 10.54 mg/L, decreasing to a value close to 1400.64 ± 23.25 mg/L by day 18. The COD reduction is more pronounced in the first days (0–6), suggesting high initial metabolic activity [46]. Subsequently, the reduction rate seems to stabilize, possibly due to the reduction of available substrate or the accumulation of inhibitory metabolic products [46]. The decrease in COD suggests that the microbial fuel cell system effectively treats the whey while generating electricity from substrate degradation [45,46]. Dairy industry waste has been used as a substrate in MFCs, achieving voltage peaks of 0.576 V while simultaneously reducing COD values by 63 ± 5%, indicating that a well-developed anodic biofilm improves electron transfer and substrate utilization, promoting complete organic matter oxidation and COD reduction [47]. Similarly, the wastewater was used as a substrate in their MFCs, observing a 57% reduction in COD, mentioning that proper pretreatment or dilution of the substrate mitigates inhibitors (e.g., high salinity, lipids), allowing uninterrupted microbial activity and complete substrate oxidation [48]. Figure 4d shows the oxidation-reduction potential (ORP) variation over time in microbial fuel cells using whey as a substrate. The ORP rapidly increased from 11.163 ± 0.513 mV to 104.287 ± 4.058 mV on day 11. Then, the ORP gradually decreased to 73.557 ± 7.158 mV by day 18. The increase in ORP indicates higher metabolic activity of electroactive microorganisms in the MFCs, favoring electron transfer to the anode [49]. As microorganisms consume the whey, intermediate metabolites are generated that affect the redox balance [47]. Initially, the oxidation of organic compounds generates electrons, increasing the ORP. However, with the depletion of the substrate, the process slows down, and the ORP begins to decrease [50]. The ORP trend is inverse to the decrease in COD observed in the figure, suggesting that as organic compounds are degraded, the cell reaches its maximum electrochemical performance before substrate depletion reduces microbial activity [48,49,50].
The power density values initially increase with the current density (Figure 5a), reaching a PDmax of 1.585 ± 0.061 mW/cm2 at a CD of 4.448 A/cm2 with a peak voltage of 801.235 ± 25.155 mV. Subsequently, the power density decreases as the current density continues to increase. As the power density increases in the optimal region, it indicates that the microorganisms are generating more electrons from the degradation of whey [46]. However, when the current density is too high, the MFCs become less efficient, which could indicate electron transfer limitations or the available substrate’s consumption [51]. A PD of 1.78541 mW/cm2 was observed using marine waste in MFCs, mentioning that losses can occur due to ohmic polarization (due to the resistance of the electrolyte and conductive materials) [37]. Similarly, dairy wastewater was used as a substrate, generating a PD of 62 mW/m2 and a maximum voltage of 0.66 V, mentioning that excess current can cause a voltage drop, reducing energy conversion efficiency [50]. It has also been observed that biofilm formation on the electrodes can affect electron transfer and internal resistance, impacting the relationship between power density and current [51]. Figure 5b shows the voltage-current (V-I) curve with a linear fit, characteristic of an internal resistance analysis in a microbial fuel cell. According to the fit equation y = a + bx and following Ohm’s Law, where the slope determines the internal resistance, an internal resistance of 16.8471 ± 0.911 Ω was calculated. The value of 16.8471 ± 0.911 Ω is relatively low, suggesting good conductivity and efficiency in electron transfer within the system [50]. If the internal resistance were higher, it would indicate possible energy losses due to limitations in the electrolyte conductivity, the quality of electrode materials, or restrictions in the diffusion of electroactive species [52]. The lower the internal resistance, the higher the energy conversion efficiency, and this value is key to optimizing cell design, as high internal resistance reduces the generated power and system efficiency [53]. Researchers calculated an internal resistance of 117 Ω and a power density of 0.1047 mW/m2 using municipal organic waste as substrates in their MFCs, mentioning that the composition and design of the electrodes affect internal resistance and that materials with higher electrical conductivity, such as graphite or activated carbon, can reduce internal resistance [54]. Researchers showed an internal resistance of 4.9 Ω and a power density of 226 mW/m2 using bamboo waste as substrates in their MFCs, mentioning that a high density of active microorganisms can improve electron transfer, reducing internal resistance. However, high density can also cause problems, such as accumulating waste products that increase resistance [48].
Figure 6 presents the metagenomic analysis of microbial fuel cells (MFCs) using whey as a substrate, which reveals the composition of the bacterial community involved in substrate degradation and bioelectricity generation. The pattern identified three phyla, four classes, three orders, four families, and two bacterial species. Among the most abundant bacterial phyla are Bacteroidota (48.47%), followed by Proteobacteria (29.83%). Bacteroidia (38.98%) is the most abundant bacterial class, predominant in the Bacteroidota phylum. In terms of bacterial order, the most abundant are Bacteroidales (38.63%) and Acetobacterales (33.39%). Regarding bacterial families, Bacteroidaceae shows a high presence at 38.58%, followed by Acetobacteraceae at 33.39%. Additionally, it is observed that within the dominant phyla, there are well-defined classes, with a notable presence of Caproiciproducens sp. and Stenotrophomonas maltophilia. Similar research has demonstrated that bacteria such as Proteobacteria are abundant in whey and are associated with carbohydrate degradation and electron production. At the same time, Bacteroidota thrive in whey rich in lactose and proteins, favoring fermentative bacteria like Bacteroidaceae, which specialize in metabolizing complex compounds [55,56]. Studies with MFCs fed with whey have found a predominance of Proteobacteria and Bacteroidota, which aligns with these results. Bacteroidaceae (class Bacteroidia) is linked to polysaccharide fermentation, consistent with lactose degradation in whey [57].
Bacteroidota are known for their ability to degrade complex compounds such as polysaccharides and proteins present in whey [55]. This is due to their extensive repertoire of hydrolytic enzymes that break down these compounds into simpler molecules, which can be utilized by other microorganisms or directly for electricity generation [57]. During the degradation of organic compounds, Bacteroidota produces volatile fatty acids (VFAs) such as acetate and butyrate. These VFAs are crucial for electron transfer in MFCs, as they can be oxidized by other electroactive microorganisms, generating electrons that are transferred to the anode [55,56]. Similarly, Proteobacteria have a highly versatile metabolism that allows them to utilize a wide range of organic compounds in whey [57]. This includes oxidizing both simple and complex compounds, contributing to electron generation [58]. Some species of Proteobacteria, such as those in the genus Geobacter, are known for their ability to transfer electrons directly to electrodes [57]. This is achieved through bacterial nanowires or outer membrane proteins that facilitate electron transfer from the bacterial cell to the anode [59]. Additionally, Caproiciproducens sp. is associated with caproate production, a compound of interest in biofuel synthesis [60]. Its presence suggests potential for the valorization of dairy waste [61]. Stenotrophomonas maltophilia, a versatile organism, could contribute to protein degradation in whey and electron transfer, as observed in MECs treating nitrogenous effluents [62]. The abundance of Bacteroidaceae and Acetobacteraceae suggests an efficient pathway for converting whey into electroactive intermediates (e.g., acetate), which could optimize bioelectricity production compared to systems using fewer complex substrates [63].
Figure 7 shows the energy generation process scheme, which begins with milk collection and cheese production, during which whey is generated as a byproduct. This whey is then utilized as a substrate in MFCs, where microorganisms degrade it and generate electricity. Each MFC containing 800 mL of cheese whey was connected in series, achieving a recorded voltage of 2.14 V, demonstrating the capability of cheese whey to generate renewable energy. This process reduces the environmental impact of whey and provides a sustainable source of electricity.
The economic feasibility and potential application of microbial fuel cells (MFCs) with cheese whey can be analyzed from the material cost perspective [64]. MFCs can be constructed using low-cost materials such as carbon electrodes and accessible proton exchange membranes, significantly reducing initial implementation costs [17]. Cheese whey is a by-product of the dairy industry, making it an economical and abundant substrate [65]. Utilizing cheese whey not only reduces operational costs but also helps manage industrial waste [14]. The maintenance and operational costs of MFCs are relatively low compared to other energy generation technologies [16]. However, it is essential to consider the costs associated with electrode cleaning and replacement and the control of by-product accumulation [17]. Integrating MFCs into a circular economy model allows for the reuse of industrial waste, such as cheese whey, for energy generation, and the production of valuable by-products, such as eco-friendly fertilizers [66]. MFCs are scalable and can adapt to different environments and applications, from small domestic units to large industrial installations. This makes them versatile and suitable for various applications [67].

4. Conclusions

Microbial fuel cells (MFCs) fed with whey demonstrated outstanding electrochemical performance, reaching a maximum voltage of 0.867 ± 0.059 V and a maximum current of 4.114 ± 0.239 mA on the eleventh day. The pH increased from 3 to 4.135 ± 0.264, indicating partial neutralization of organic acids by bacterial activity, showing an optimal pH of 4.135 ± 0.264 on day 11. The electrical conductivity reached 170.062 ± 9.511 mS/cm, favoring proton transfer and reducing internal resistance (16.8471 ± 0.911 Ω), which optimized energy efficiency. The redox potential (ORP) showed compensation with microbial activity, reaching 104.287 ± 4.058 mV at the peak of electricity generation (eleventh day). A 70% reduction in chemical oxygen demand (COD) was achieved, decreasing from 4650.52 ± 10.54 mg/L to 1400.64 ± 23.25 mg/L in 18 days. This demonstrates the MFCs’ ability to efficiently degrade the organic compounds in whey, mitigating its environmental impact and aligning with circular economy strategies. Furthermore, the low internal resistance and high-power density (1.585 ± 0.061 mW/cm2) highlight their scalability for applications in rural areas like Peru, where whey is an abundant and underutilized waste. The metagenomic analysis revealed the predominance of Bacteroidota (48.47%) and Proteobacteria (29.83%), with key families such as Bacteroidaceae (38.58%) and Acetobacteraceae (33.39%). The results validate using MFCs as a dual technology for both whey treatment and renewable energy production. This reduces waste management costs in the dairy industry and prevents pollutant emissions from untreated discharges.
Although the system showed stability over 18 days, the decrease in current and conductivity in the final stages suggests substrate depletion or accumulation of inhibitors. Future studies could explore continuous whey recharge strategies and electrode optimization to improve electron transfer. In summary, this provides solid evidence regarding the potential of MFCs to transform whey from an environmental liability into an energy resource, promoting sustainability in the dairy industry and contributing to the Sustainable Development Goals (SDGs).

Author Contributions

Conceptualization, R.-F.S.; methodology, R.-F.S. and M.D.L.C.-N.; software, C.-C.L.; validation, M.G.-C.; formal analysis, M.D.L.C.-N. and M.G.-C.; investigation, N.M.O.; resources, N.M.O.; data curation, M.G.-C., C.-C.L. and M.G.-C.; writing—original draft preparation, M.D.L.C.-N.; writing—review and editing, N.M.O. and R.-F.S.; visualization, M.G.-C.; supervision, R.-F.S.; project administration, R.-F.S.; funding acquisition, R.-F.S. All authors have read and agreed to the published version of the manuscript.

Funding

This research was funded by Universidad de Cesar Vallejo via approved project No. P-2024-172.

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Data Availability Statement

The original contributions presented in this study are included in the article. Further inquiries can be directed to the corresponding author.

Acknowledgments

The authors thank the Universidad de Cesar Vallejo for technical support.

Conflicts of Interest

The authors declare no conflicts of interest.

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Fermentation 11 00176 g001
Figure 1. Schematic of the experiment.
Figure 1. Schematic of the experiment.
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Figure 2. Voltage evolution in microbial fuel cells over time.
Figure 2. Voltage evolution in microbial fuel cells over time.
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Figure 3. Evolution of current in microbial fuel cells over time.
Figure 3. Evolution of current in microbial fuel cells over time.
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Fermentation 11 00176 g004aFermentation 11 00176 g004b
Figure 4. Evolution of the values of (a) pH, (b) electrical conductivity, (c) COD, and (d) ORP.
Figure 4. Evolution of the values of (a) pH, (b) electrical conductivity, (c) COD, and (d) ORP.
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Figure 5. Performance of the MFCs in their (a) PD vs. CD and (b) internal resistance.
Figure 5. Performance of the MFCs in their (a) PD vs. CD and (b) internal resistance.
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Figure 6. Composition of bacterial communities in MFC samples: relative abundance of dominant groups at all taxonomic levels, including (a) phylum, (b) class, (c) order, (d) family, and (e) species.
Figure 6. Composition of bacterial communities in MFC samples: relative abundance of dominant groups at all taxonomic levels, including (a) phylum, (b) class, (c) order, (d) family, and (e) species.
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Figure 7. Bioelectricity generation process from cheese whey.
Figure 7. Bioelectricity generation process from cheese whey.
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Table 1. Physicochemical parameters of whey.
Table 1. Physicochemical parameters of whey.
ParameterValue
pH3.07 ± 0.01
Oxidation-Reduction Potential (mV)11.163 ± 0.513
Electrical Conductivity (mS/cm)19.101 ± 1.025
Fat (%)0.4
Chemical Oxygen Demand (mg/L)4650.52 ± 10.54
Density (g/cm3)1.026
Temperature (°C)23
Lactose (%)4.71
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MDPI and ACS Style

Segundo, R.-F.; Luis, C.-C.; Otiniano, N.M.; De La Cruz-Noriega, M.; Gallozzo-Cardenas, M. Utilization of Cheese Whey for Energy Generation in Microbial Fuel Cells: Performance Evaluation and Metagenomic Analysis. Fermentation 2025, 11, 176. https://doi.org/10.3390/fermentation11040176

AMA Style

Segundo R-F, Luis C-C, Otiniano NM, De La Cruz-Noriega M, Gallozzo-Cardenas M. Utilization of Cheese Whey for Energy Generation in Microbial Fuel Cells: Performance Evaluation and Metagenomic Analysis. Fermentation. 2025; 11(4):176. https://doi.org/10.3390/fermentation11040176

Chicago/Turabian Style

Segundo, Rojas-Flores, Cabanillas-Chirinos Luis, Nélida Milly Otiniano, Magaly De La Cruz-Noriega, and Moises Gallozzo-Cardenas. 2025. "Utilization of Cheese Whey for Energy Generation in Microbial Fuel Cells: Performance Evaluation and Metagenomic Analysis" Fermentation 11, no. 4: 176. https://doi.org/10.3390/fermentation11040176

APA Style

Segundo, R.-F., Luis, C.-C., Otiniano, N. M., De La Cruz-Noriega, M., & Gallozzo-Cardenas, M. (2025). Utilization of Cheese Whey for Energy Generation in Microbial Fuel Cells: Performance Evaluation and Metagenomic Analysis. Fermentation, 11(4), 176. https://doi.org/10.3390/fermentation11040176

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