Poultry slaughterhouse wastewater treatment using nanobubble technology Open Access

Poultry slaughterhouses release substantial volumes of wastewater into the environment because of their extensive use of freshwater for ongoing activities such as meat cutting and rinsing. This poultry slaughterhouse wastewater (PSW) is highly contaminated, featuring organic matter measured by biochemical oxygen demand (BOD), chemical oxygen demand (COD), total dissolved solids (TDS), and total suspended solids (TSS). Additionally, it contains high levels of nitrogen and phosphorus components, encompassing substances such as blood, fats, oil and grease (FOG), and proteins (Basitere et al. 2019; Ng et al. 2022; Teo et al. 2023). Improper discharge of inadequately treated PSW poses a substantial risk of contaminating freshwater sources. This poses potential environmental and health hazards, including river deoxygenation, groundwater pollution, eutrophication, and the potential spread of waterborne diseases. Therefore, it is extremely essential to undertake on-site treatment of PSW (Kothari et al. 2024), thereby avoiding or reducing environmental contamination and facilitating the potential reintegration of treated wastewater into plant operations, particularly surface cleaning (Reilly et al. 2019; dos Santos Pereira et al. 2024).

Biological treatment methods, including aerobic and anaerobic processes, have been widely adopted for managing the challenges associated with treating PSW (Philipp et al. 2021). Aeration systems, recognized as one of the oldest and simplest methods, have been extensively used to treat PSW (Gutu et al. 2021; Philipp et al. 2021; dos Santos Pereira et al. 2024). These methods are particularly preferred for handling wastewater concentration of pollutants from industrial effluents, agricultural runoff, or wastewater from urban sources, due to their easy construction and operation, as highlighted by Musa & Idrus (2021) and Ng et al. (2022).

Table 1 shows typical conventional aerobic processes for the treatment of PSW. These processes achieved between 72.2 and 94.7% COD removal. However, conventional aeration methods have their drawbacks, including the build-up of significant sludge, substantial energy consumption for aeration, vulnerability to high organic loads, and extended hydraulic retention times (Ahmadi et al. 2022). Moreover, conventional aeration methods have faced difficulties due to their limited oxygen transfer efficiency, typically ranging from 6 to 10%.

To address these challenges, significant attention has been directed towards technologies aimed at enhancing aerobic processes through the optimisation of aeration methods and gas diffusion, with a specific emphasis on controlling bubble sizes (Sakr et al. 2022). Nanobubble (NB) technology has emerged as a promising advancement in the domain of wastewater treatment. NBs are tiny gas cavities with diameters smaller than 1 μm according to International Organization for Standardisation (2017). Compared to ordinary bubbles, NBs possess distinct characteristics. Their small size and high surface tension enhance the liquid–gas contact area, facilitating physical adsorption, chemical reactions, and mass transport at the gas–liquid interface (An et al. 2019; Shi 2022). Due to their low buoyancy, NBs ascend to the surface more slowly, prolonging their presence in the liquid phase (Kalogerakis et al. 2021). Moreover, they enhance gas mass transfer efficiency by reducing bubble size and increasing internal pressure (Azevedo et al. 2019).

NBs have proven to be highly useful in wastewater treatment. For instance, air-NBs have proven effective in treating various types of wastewater. In their study, Leyva & Flores (2018) demonstrated a reduction of 79.0% in TSS and 85.0% in COD in sugar sector wastewater in under 90 min. Another study by Reyes & Flores (2017) reported a 66.21% removal efficiency for total coliforms. Additionally, Wang & Zhang (2017) investigated the integration of NB into a deep subsurface wastewater infiltration system, achieving removal percentages of 95.1, 98.5, and 99.9% for COD, NH₄⁺-N, and total phosphorus; respectively.

While NB technology has been applied to other high-organic-load wastewater streams such as sugar industry effluent (Leyva & Flores 2018) and synthetic wastewater (Van Leeuwen et al. 2009), its application to PSW remains unexplored. Conventional methods such as aerobic reactors and constructed wetlands have achieved pollutant removal rates of up to 94.7% for COD and 93.3% for TSS (Keerthana & Thivyatharsan 2018), but these methods face limitations, including high sludge production and prolonged hydraulic retention times (Ahmadi et al. 2022). This study demonstrates the potential of NB technology for PSW treatment using air-NBs, ozone-NBs, and air-NBs combined with enzymes. Unlike previous studies, this research provides a comparative analysis of three NB-based systems, elucidating the specific advantages of each approach.

The PSW was collected from a local poultry abattoir in Cape Town, South Africa. The wastewater resulted from diverse activities, including slaughtering, feather removal, evisceration, trimming, carcass washing, deboning, chilling, packaging, and the cleaning of facilities and equipment. The collection point was a stream situated between the abattoir and the equalization tank. The raw wastewater was placed in 25 L polyethylene containers, which were then stored in a refrigerator at 4 °C. The composition of the PSW used in this study is presented in Table 2. It was important to note that the composition of poultry PSW can vary significantly depending on several factors. These include the specific practices and methods employed at the slaughterhouse, the variations in the slaughtering process, and the timing of wastewater sample collection. Factors such as the scale of operations, the types of poultry processed, and the cleaning protocols in place also contribute to the variability. The time of sample collection, whether during peak processing periods or after cleaning activities, can lead to differences in the concentration and types of pollutants present in the wastewater.

The PSW composition in this study aligns with the ranges reported in previous studies from internationally published papers. However, except for pH, all measured parameters exceed the general discharge limits established by the South African National Water Act of 1998. This highlights a significant requirement for the treatment of PSW to mitigate the environmental impact associated with PSW discharge, ensuring compliance with regulatory standards, and protecting the environment.

The study focuses on evaluating different NB aeration methods for treating PSW in three phases:

• Phase 1: Treatment with air-nanobubbles (air-NBs)

• Phase 2: Treatment with ozone-nanobubbles(ozone-NBs)

• Phase 3: Treatment with air-NBs combined with enzymes (Ecoflush)

Each phase consisted of a 200 L polyethylene tank filled with PSW. Prior to aeration using the NB generator (MK3), the PSW underwent screening using three Madison test sieves of 100 μm to eliminate suspended solids. The MK3 NB generator, connected to the bottom of the tank for continuous circulation, was equipped with a nozzle for simultaneous air/ozone injection and NB generation. The PSW was aerated for 6 h to allow the breakdown of organics and nutrients, with samples collected from the bottom of the aeration tank at 2 h intervals.

The NB generator used in this study was an MK3 nanobubbler^TM from Fine Bubble Technology (Pty) Ltd, based in Porterville, South Africa. The MK3 NanoBubbler™ has been tested and proven to produce 220,000,000 NBs/mL with an average size of 76 nm (range 10–300 nm) in diameter (Fine Bubble Technologies 2024).

The MK3 NanoBubbler™ uses the venturi method to generate NBs and comprises a porous nanofilm membrane venturi tube directly connected to a water supply source. It is inserted into a pipe-shaped body with a vent, mounted at the centre of a pipeline conduit of a water-supplying pipe, and connected to the vent through a hose for air injection. The generator includes a nano-bubble water generating unit with multiple vortex formation units and pipe couplers positioned on both sides of the nano-bubble generator.

An ozone generator 0Z–3G was used to produce ozone and the ozone was injected into the gas inlet of the venturi of the MK3 NB. The OZ-3G ozone generator uses a fan-cooled corona discharge (CD) tube to produce ozone from various pressurized sources such as compressed air, bottled oxygen, or an oxygen generator. It is equipped with a light-duty air compressor for self-generated compressed air, ensuring high ozone concentration with low energy consumption.

The ozone generator and air compressor feature switches and light-emitting diode (LED) status indicator lights. Ozone discharge concentration is adjustable from 0 to 100% using a dial potentiometer, regulating the current of the CD tube, as indicated on a 0–500 mA ammeter. The unit has electrical protection through an externally accessible fuse.

A volume of 500 mL Ecoflush^TM (Mavu Biotechnologies Pty Ltd SA) was added to the 200 L of raw PSW. The mixture was aerated for 6 h using the MK3 NB generator to allow the activation of the enzymes and biodegradation of FOG. NB aeration ensured a consistent and sufficient provision of dissolved oxygen (DO), promoting optimal proliferation of aerobic bacteria in the Ecoflush^TM.

Ecoflush is a bioremediation agent commercially produced by Ergofito and distributed in South Africa through Mavu Biotechnologies. It is a blend of natural components and bacteria. It remains inactive until exposed to a nutrient-rich organic source, such as PSW, which serves as a substrate. Once activated, it primarily generates enzymes for the hydrolysis of FOG. The natural ingredients in Ecoflush are sourced from glaucids and essential amino acids, forming potent decomposing agents that stimulate specific bacteria to produce enzymes naturally. These enzymes have the capability to break down the hydrocarbon chains present in FOG.

For each phase, PSW samples were collected from the sampling point at the bottom of the aeration tank at 2 h intervals. The following parameters were analysed at a South African National Accreditation System accredited laboratory (Bemlab, Somerset West, South Africa): COD, TSS, ammonia-nitrogen (NH₃-N), total nitrogen (total-N), as well as FOG. Furthermore, pH, temperature, and DO were measured at 1 h intervals using a multi-parameter.

Wastewater treatment for PSW is challenging due to high levels of organics and nutrients. NB aeration technologies offer innovative solutions by improving mass transfer, oxidation, and biological activity. This section presents and discusses the performance of three NB-based treatments: air-NB treatment (Phase 1), ozone-NB treatment (Phase 2), and air-NB treatment combined with enzymes (Ecoflush) (Phase 3) in treating PSW. First, the performance of each phase is analysed individually in removing pollutants.

The analysis is presented in two parts. First, the individual performance of each treatment phase is evaluated, highlighting their strengths and limitations in addressing specific pollutants. Second, a comparative assessment of the three phases is conducted, providing insights into their relative efficiency and suitability for treating PSW. This comprehensive evaluation aims to identify the most effective method for optimizing wastewater treatment in this context.

Ahmed et al. (2023) generated NBs by injecting gases such as oxygen and air through a ceramic membrane to enhance secondary effluent (municipal wastewater) treatment. They reported that air-NB injection resulted in a 28% reduction in TSS, a 26% decrease in COD, a 43% decrease in BOD₅, an 11% decrease in organic nitrogen, and a 96% decrease in NH₃-N after 2 h of aeration. Similarly, Oktafani et al. (2019) investigated the effect of aeration on chicken slaughterhouses to assess organics removal using the granular activated sludge–sequencing batch reactor (GAS–SBR) system. The findings showed that after 2 h of aeration, the removal of COD, and BOD was 72.8%. Extending the aeration period to 4 h resulted in a total NH₃-N removal of 65.8%.

The results in this study after 6 h of air-NB of PSW achieved higher COD reduction (24.2% higher) compared to the conventional aeration GAS–SBR chicken wastewater treatment by Ahmed et al. (2023). This is attributed to the high surface area of NBs, facilitating enhanced mass transfer and subsequent oxidation of pollutants combined with their ability to release hydroxyl radicals, which can interact directly and non-selectively with organic pollutants. Lastly, the increased contact between bubble surfaces and contaminants contributes to this effect (Fan et al. 2019).

In Table 3, the results of air-NB treatment after 6 h compared to the standard discharge limits outlined in the National Water Act 36 of 1998 show that only total-N met the discharge standards. NH₃-N, COD, TSS, and FOG are still above the required discharge standards and may require extended aeration times of more than 6 h to further reduce their levels and meet the discharge standards. Further exploration into extending the aeration duration is necessary.

Ozone has been applied to remediate organics, ammonia, and disinfection in water and wastewater treatment since the 1970s (Shangguan et al. 2018; Sakr et al. 2022). Despite its capabilities in decomposing organics and inactivating microorganisms, the broader utilization of ozone is constrained by challenges such as low mass transfer efficiency, limited saturation solubility, and a short half-life. These constraints often result in reduced reaction efficiency and underutilization of ozone in water treatment (Andinet et al. 2016). To address these limitations, the application of NB technology is explored to enhance the ozonation process in water and wastewater treatment (Shangguan et al. 2018; Xia & Hu 2018). In Phase 2 (Figure 1(b)), ozone-NBs were used to treat PSW, and the removal rates for COD, TSS, and FOG were 86.7, 93.5, and 99.5%, respectively, after 6 h. For total-N and NH₃-N, ozone-NB treatment achieved 40.0 and 41.2%, respectively. The low removal rates of total-N and NH₃-N can be attributed to several factors. The process of nitrogen produces intermediate compounds such as nitrites and nitrates, which contribute to total nitrogen (Luo et al. 2022). Additionally, high pH and alkalinity can reduce ozone effectiveness, while competing reactions with other contaminants lower ozone availability for nitrogen removal (Ma et al. 2023; Chen et al. 2024). The pH of ozone-NB treatment increased from 7.2 to 7.5. Further optimisation of the pH is required for the removal of ammonia and total-N by ozone-NB.

These results exhibit similarities to previous studies. For instance, Van Leeuwen et al. (2009) treated synthetic wastewater containing methylene blue using ozonation during the process. They reported an average COD removal of about 80.5% in the ozonated and 79.6% in the un-ozonated control. Similarly, Wu et al. (2022) utilized an integrated approach combining NB and ozone oxidation to enhance ammonia (NH₃-N) removal from wastewater. They found that ammonia concentration decreased slightly to 910 mg/L with a removal efficiency of 44.2% in the control group. In contrast, the NB treatment group saw a faster decrease, reaching 277 mg/L with a removal efficiency of 82.5% in 30 min due to the NB's ability to slowly release gas into the water.

The results of ozone-NB treatment presented in Table 3 are compared to the standard discharge limits outlined in the National Water Act 36 of 1998 of Republic of South Africa (1998), only FOG met the discharge standards. NH₃-N, COD, TSS, and total-N may require extended aeration time (more than 6 h) to further reduce their levels and meet the discharge standards. Further exploration into extending the aeration duration is necessary.

Enzymes are used in the hydrolysis of fats and greases in wastewater such as PSW (Affes et al. 2017). The enzymatic approach enhances the performance of microorganisms in subsequent biological treatment processes, as indicated by Jamie et al. (2016). Ecoflush, a blend of natural components and bacteria, remains inactive until exposed to a nutrient-rich organic source, such as PSW, which serves as a substrate. Once activated, it primarily generates enzymes for the hydrolysis of FOGs (Mdladla et al. 2021; Meyo et al. 2021). Figure 2(c), demonstrates that the removal rates of COD, TSS, FOG, NH₃-N, and total-N were 86.8, 65.7, 88.8, 99.2, and 88.8%, respectively. It can be noted that NH₃-N, which could not be removed efficiently by air-NB (40.2% removal) and ozone-NB (41.2% removal) after 6 h of aeration, achieved a much higher removal rate (99.2%) when air-NB was combined with an enzyme such as Ecoflush. Enzymes combined with NBs achieve higher ammonia removal rates compared to air-NB aeration alone due to several factors. Enzymes specifically target and break down ammonia more efficiently than microbial processes alone (Liu & Smith 2021). NBs enhance oxygen transfer, supporting both enzyme activity and aerobic microbial processes involved in nitrification (An et al. 2019; Azevedo et al. 2019; Shi 2022). The combination of enzymes and NBs creates a synergistic effect, optimizing conditions for enzyme action and microbial degradation.

Dlamini et al. (2021) used enzymes (Ecoflush) to treat PSW using conventional aeration (macro bubbles), the treatment resulted in an average removal rate of 80 ± 6.3% for FOG, 38 ± 8.4% for COD, and 56 ± 7.2% for TSS after 24 h of aeration, which pale in comparison to air-NB with the same concentration of enzymes (Ecoflush). Similarly, Ngobeni et al. (2022) reported removal rates of FOG by 85–99%, and COD by 20–50% using Ecoflush with macrobubbles after 24 h of aeration treating PSW. These results highlight the superior performance of air-NB combined with Ecoflush compared to Ecoflush with macro bubbles aeration.

The findings presented in Table 3 are compared to the standard discharge limits outlined in the National Water Act 36 of 1998 of Republic of South Africa (1998). After 6 h of aeration, NH₃-N and total-N levels complied with the specified limits. However, COD, TSS, and FOG still exceeded the discharge limits. Further exploration into extending the aeration duration is necessary to comply with the discharge standards for COD, TSS, and FOG.

COD serves as a standard parameter to gauge water pollution levels, with its analysis revealing changes in water-soluble oxygen content and indicating the ease or difficulty of decomposition. The heightened COD levels observed in PSW stem from the significant presence of organic waste, including residual organs, blood, and unused chicken parts (Basitere et al. 2019).

Air-NB and ozone-NB exhibited the most significant treatment effect on PSW, leading to 88.6 and 86.7% reduction in COD after 4 h of treatment, compared to air-NB combined with Ecoflush (19.4%). This is attributed to the higher concentration of DO in NB, facilitating enhanced mass transfer and subsequent oxidation of pollutants. Furthermore, the collapse of NBs can release hydroxyl radicals, which can interact directly and non-selectively with organic pollutants. Lastly, the increased contact between bubble surfaces and contaminants contributes to this effect (Fan et al. 2019).

It was observed that the COD removal by air-NB and ozone-NB reached a plateau after 2 h of aeration due to several factors. The plateau can be attributed to the system reaching a saturation point where available oxygen is fully utilized. The depletion of easily degradable substrates and the formation of biofilms, which limit oxygen and nutrient diffusion, can also contribute to the observed plateau in COD reduction (Yaparatne et al. 2022).

The COD removal by Ecoflush combined with NB was delayed and slow during the first 4 h due to several factors. First, enzymes require time to activate and catalyse the breakdown of organic matter, and there may be initial diffusion limitations of nanobubbles and enzymes throughout the wastewater (Liu & Smith 2021). Additionally, inhibitory substances such as FOG in PSW can temporarily reduce enzyme activity, and the system needs time to reach an equilibrium state for optimal COD removal (Povis & Pérez 2023).

Analysing the distribution of TSS serves as a common approach to evaluating water quality. Elevated TSS levels indicate increased pollution, obstructing light penetration into the water and disrupting photosynthesis. Additionally, TSS can absorb solar thermal energy, potentially elevating water temperature and subsequently reducing DO levels.

The performance of ozone-NBs significantly reduced the TSS content of the PSW by 93.5%, marking the most substantial decrease compared to air-NBs, which achieved a reduction of only 78.9 and 65.7% when combined with Ecoflush. This highlights the practical effectiveness of NBs in TSS reduction. This is attributed to the NBs' capability to broaden the range of flotation particle sizes, enhance particle surface hydrophobicity, and improve froth flotation efficiency (Wu et al. 2021). This technique has been widely recognized for its efficacy in TSS removal due to the similar size and opposing charge of bubbles and suspended particles, facilitating enhanced collisions and adherence. Additionally, the organic nature of these solids enables further oxidation with ozone-NBs, contributing to their effectiveness in reducing TSS content (Kyzas et al. 2021). The findings of this study align closely with previous research. For instance, Rameshkumar et al. (2019) examined the impact of ionization-induced NBs on domestic wastewater treatment, noting a complete reduction of TSS by nearly 100%.

Ammonia compounds pose significant risks to aquatic life, often stemming from sources like urine, faeces, and the microbial decomposition of organic matter in both natural and industrial water bodies. Even at concentrations as low as 1 mg/L, ammonia can diminish oxygen levels in water, posing a severe threat to aquatic organisms and potentially resulting in fatalities.

The outcomes of this study exhibit similarities to prior research accomplishments. For instance, Atkinson et al. (2019) introduced NBs into a wastewater treatment facility in Missouri, witnessing a rapid reduction in organics and turbidity, although with a significant decline in ammonia removal. Furthermore, Wang et al. (2020) explored the oxygenation and concurrent regulation of nitrogen and phosphorus release at the sediment-water interface using oxygen-NB modified material, resulting in notable reductions in NH3-N, and total nitrogen (TN) by 96.4, 51.1, and 24.9%, respectively.

This study demonstrates the potential of NB technology in the treatment PSW, particularly in achieving high removal efficiencies of ammonia, COD, TSS, and FOG. To advance the practical application and scalability of this technology, several key areas merit further investigation. Future research should focus on optimising aeration duration to ensure complete pollutant removal. While significant reductions were achieved within 6 hours, certain parameters – especially COD and TSS – remained above discharge limits. Investigating extended aeration times, along with the effects on additional contaminants such as nitrates and nitrites, could provide deeper insights into treatment efficiency. Integrating NB technology with other treatment processes offers a promising avenue for performance enhancement. Hybrid systems incorporating biological reactors, advanced oxidation processes, or membrane technologies may overcome current limitations, such as incomplete degradation or excessive sludge generation, by leveraging complementary mechanisms. Assessing the long-term performance and sustainability of NB systems under real-world conditions is crucial. Pilot- and full-scale studies are needed to evaluate operational reliability, energy consumption, and cost-effectiveness, thereby supporting industrial-scale implementation. Finally, mechanistic studies on the interaction between NB and enzymatic agents – such as Ecoflush – could clarify the synergistic effects observed in this study, particularly for ammonia and FOG removal. Elucidating the activation pathways and enhancement of enzymatic activity in the presence of nanobubbles may inform the development of more effective enzyme-assisted NB treatments.

This study demonstrates the efficacy of NB technology in the treatment of PSW, particularly in the removal of pollutants such as COD, TSS, ammonia, and FOG. The results indicate that air-NB aeration and ozone-treated NBs achieved impressive COD removal rates of over 80% within just 2 h, highlighting their rapid pollutant removal capabilities. Furthermore, the study shows that NBs combined with Ecoflush enzymes, although initially slower in pollutant removal, can achieve high removal rates after 4–6 h of aeration, particularly for ammonia and FOG. The steady TSS removal efficiency across all methods after 4 h underscores the reliability of NB technology in TSS reduction. Additionally, the study highlights the importance of enzyme treatments like Ecoflush in enhancing FOG removal efficiency. Overall, the findings suggest that NB technology, especially when coupled with enzyme treatments, holds promise for efficient and sustainable PSW treatment.

This research was made possible through financial support from the Thuthuka and Black Academic Advancement Research Funding programmes of the National Research Foundation (NRF). We also gratefully acknowledge the equipment support provided by Fine Bubble Technologies (Pty) Ltd We extend our sincere gratitude to both institutions for their commitment to advancing research in the field of NB technology.

All relevant data are included in the paper or its Supplementary Information.

The authors declare there is no conflict.