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Article

Optimising Galdieria sulphuraria ACUF 427 Biomass for Enhanced Urban Wastewater Treatment: Evaluating Pollutant Removal Efficiency, Algal Growth, and Phycocyanin Production

by
Berhan Retta
1,*,
Manuela Iovinella
2 and
Claudia Ciniglia
2
1
Department of Engineering, University of Campania Luigi Vanvitelli, Via Roma 29, 81031 Aversa, Italy
2
Department of Environmental, Biological and Pharmaceutical Sciences and Technologies, University of Campania Luigi Vanvitelli, Via Vivaldi 43, 81100 Caserta, Italy
*
Author to whom correspondence should be addressed.
Phycology 2025, 5(3), 40; https://doi.org/10.3390/phycology5030040
Submission received: 28 July 2025 / Revised: 18 August 2025 / Accepted: 20 August 2025 / Published: 21 August 2025
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Abstract

Urban wastewater is composed of nutrients such as nitrogen and phosphorus, organic matter, heavy metals, pathogens, and micropollutants. If untreated, these contribute to eutrophication and environmental degradation. Microalgae-based bioremediation offers a sustainable solution, showing promise for pollutant removal and high-value bioproduct generation. This study evaluates the efficacy of Galdieria sulphuraria ACUF 427 in treating urban wastewater, with a focus on nutrient removal and phycocyanin production at different optical densities (OD 2, OD 4, and OD 6). Nutrient removal rates (RRs) were analysed for ammonium nitrogen (N-NH4+), ammonia nitrogen (N-NH3), phosphate phosphorus (P-PO43−), and chemical oxygen demand (COD). The RR for N-NH4+ increased with optical density, reaching 7.49 mg/L/d at an optical density of 6. Similar trends were observed for N-NH3 and P-PO43−, with peak removal at OD 6. COD removal remained high across all ODs, though differences between OD 4 and OD 6 were not statistically significant. Significant variations (p < 0.05) in nutrient removal were noted across the ODs, except for COD between OD 4 and OD 6. Biomass growth and phycocyanin production were significantly higher in the wastewater compared to the control (Allen Medium), with the most effective performance observed at an optical density (OD) of 6. Maximum growth rates were 0.241 g/L/d at OD 6, 0.178 g/L/d at OD 4, and 0.120 g/L/d at OD 2. These results highlight the potential of G. sulphuraria as an agent for wastewater bioremediation and the production of high-value compounds, particularly at elevated cell densities, where we achieved superior nutrient removal and biomass production.

1. Introduction

Urbanisation and anthropogenic activities are the primary factors that negatively impact the environment’s health. The expansion of cities results in the generation of wastewater in a very high volume, which can deteriorate water quality and disrupt aquatic ecosystems. Human activities pose a significant threat to the water quality of rivers when pollution exceeds the threshold limit [1], especially in urban areas where industrial activities intensify. The water pollution caused by urban and industrial sources introduces a range of contaminants into natural water bodies, including nutrients [2,3], pathogens [4,5], plastics [6], chemicals [7], and heavy metals [3]. These pollutants cause eutrophication, which gives rise to algal blooms [8] and, hence, biodiversity loss. Moreover, they are hazardous to the environment and human health [9]. Innovative and sustainable wastewater treatment solutions are needed to address these pollutants effectively.
The circular economy concept advocates the efficient use of resources, emphasising the recycling and reuse of materials while reducing waste and minimising environmental impacts. According to Zhang and Liu et al., [10] recovering reusable water, energy and nutrients from municipal wastewater has desirable environmental and economic benefits. Within this framework, microalgae offer a promising alternative to traditional wastewater treatment methods. Microalgae can effectively remove nitrogen, phosphorus, heavy metals, pesticides, organic and inorganic toxins, and pathogens from wastewater [11], making them a suitable candidate for phycoremediation.
According to Usman et al. [9] rapid urbanisation, economic growth and increased water consumption in urban areas have made urban wastewater the largest reservoir of nutrient and energy-rich waste materials. Khan et al. [12] have also highlighted that urban wastewater has an ideal N/P ratio for cultivating microalgae, which converts nitrogen and phosphorus into nutrient-rich biomass. This micro-algal biomass has biostimulants in the form of phytohormones responsible for plant growth stimulation, and it can also be used as a biofertiliser, resulting in increased crop productivity while reducing eutrophication risks.
Galdieria sulphuraria, an extremophilic microalga of the class Cyanidiophyceae [13], thrives in extreme environments, particularly hot, acidic springs and volcanic regions [14], where temperatures can reach up to 56 °C [15] and pH levels range from 0.5 to 4 [16]. It has immense potential in wastewater treatment. This unicellular organism is notable for its ability to perform photosynthesis while growing heterotrophically by consuming organic carbon [17]. Its remarkable adaptability and metabolic flexibility have led to interest in its potential biotechnological applications, such as bioremediation and biofuel production, making it a valuable subject for research on extremophiles. The different cultivation modes, such as autotrophy, heterotrophy and mixotrophy, could result in significant differences in the performance of this alga [18].
A few studies have utilised autotrophic G. sulphuraria in wastewater treatment. Tchinda et al. [19], for instance, reported a continuous fed-batch operation using G. sulphuraria in primary effluent treatment in which consistent removal of BOD5 (5-day biological oxygen demand) and nutrients, as well as a significant reduction in pathogenic bacteria, was demonstrated. Similarly, Selvaratnam et al. [20] studied and recorded nutrient removal from primary wastewater effluent using G. sulphuraria, with seven-day removal efficiencies of 88.3% for ammoniacal-nitrogen, and 95.5% for phosphates, with corresponding removal rates of 4.85 and 1.21 mg/L/d. Moreover, the feasibility of algal systems for sustainable wastewater treatment by growing G. sulphuraria in primary settled urban wastewater indicated that the alga reduced nutrient levels to regulatory discharge levels at reasonable rates [21]. Furthermore, removing dissolved organic carbon and nutrients from primary-settled urban wastewater by G. sulphuraria under batch mode was monitored over various influent and operating conditions at the field level. The result revealed that G. sulphuraria was able to grow well in primary-settled wastewater and reduce organic carbon (measured as BOD5), ammoniacal nitrogen, and phosphate levels to below the respective discharge standards [22].
However, the use and optimisation of G. sulphuraria ACUF 427 for urban wastewater treatment have not yet been explored. This study, therefore, focuses on optimising the use of autotrophic G. sulphuraria for primary settled urban wastewater treatment by examining its performance at various biomass concentrations (OD 2, OD 4, and OD 6). The research evaluates the alga’s ability to reduce key pollutants, namely ammonium nitrogen (N-NH4+), ammonia nitrogen (N-NH3), phosphate phosphorus (P-PO43−), and Chemical Oxygen Demand (COD). In addition to assessing pollutant removal efficiency, the study explores algal growth (measured as Dry Weight-DW) and phycocyanin (PC) production.

2. Materials and Methods

2.1. Allen Medium Preparation

The standard Allen medium (AM) was prepared using the following ingredients per 100 mL of distilled water: KH2PO4 (3 g), K2HPO4 (6 g), CaCl2.2H2O (0.2 g), NaCl (1 g), MgSO4.7H2O (3 g), FeSO4.7H2O (0.0996 g), (NH4)2SO4 (13.2 g), and micronutrients including ZnCl2 (0.14 g), Na2MoO4.2H2O (0.05 g), CuSO4.5H2O (0.1 g), CoCl2.6H2O (0.05 g), and MnCl2.4H2O (0.0208 g). The pH was adjusted to 1.5 using 95% H2SO4 and autoclaved for 3 h at 121 °C before algal cultivation.

2.2. Cultivation of Stock Solution

G. sulphuraria ACUF 427, obtained from the algal collection of the University of Federico II, Naples, Italy (www.acuf.net (accessed on 12 March 2025)), was used in this experiment. A stock solution was prepared by inoculating the alga into a standard growing medium (Allen) in a 1 L Erlenmeyer flask. The culture was maintained at a temperature of 30 °C under continuous fluorescent light, with an intensity of 89 μmol photons/m2/s.

2.3. The Urban Wastewater

Primary-settled urban wastewater (UWW), which had not undergone any biological treatment, was obtained from the ‘Foce Regi Lagni’ wastewater treatment plant in Villa Literno (CE), Campania, Italy. Before the experiment, the wastewater composition was analysed (as shown in Table 1). The pH of the wastewater was then adjusted to 4 using 95% H2SO4. The pH of the Allen medium (AM) was also similarly adjusted to 4 and verified using a pH metre.
The initial algal concentration required for the experiment was measured by Optical Density (OD) at 750 nm. Final algal concentrations were adjusted to reach OD values of 2, 4, and 6, in final volumes of 480 mL, 446 mL, and 335 mL, respectively. The required algal biomass was harvested during the exponential growth phase, centrifuged at 3800 rpm for 4 min, and the supernatant was discarded. The algal pellet was then re-suspended in UWW and AM (the control), adjusted to pH 4, and the experiment was performed in triplicate at a temperature of 30 °C under continuous fluorescent light, with an intensity of 89 μmol photons/m2/s.

2.4. Sampling and Data Collection

Algal growth was monitored at regular intervals of 0, 24, 48, 72, and 96 h. Samples were collected at these time points for chemical analysis. Strain growth rate was assessed spectrophotometrically (HP 8453A) by measuring their density as Optical Density (OD) at 750 nm. Additionally, phycocyanin (PC) content was measured at 615/652 nm.

2.5. Chemical Analysis

Ion Chromatography (863 Compact Autosampler) was employed to analyse N-NH3 and N-NH4+ levels in the samples on the collection day. The pH of the media was determined using a pH 50 Violab metre. Additionally, a Thermostat (LT 200) was used to measure PO43− and COD levels.

2.6. Data Analysis

Descriptive statistics and graphical representations were generated using Microsoft Excel [23]. A paired two-sample t-test was conducted in R Studio (Version 2024.09) [24]. The Maximum Growth Rate (MGR) of the alga was calculated using the formula provided by Sirakov et al. [25]:
MGR (1/d) = (Ln(Nt) − Ln(N0))/(t − t0)
where Nt is the dry weight at the final time, N0 is the dry weight at the initial time, t is the final time (days), and t0 is the initial time (days). Additionally, phycocyanin was calculated using the formula described by Bennet and Bogorad [26].
Phycocyanin (mg/mL) = A615 nm − 0.474 (A652)/5.34
A calibration curve with the equation Y = 0.0004x + 0.0003, (R2 = 0.9321), which correlates Optical Density (X) with Dry Weight (Y), was used to determine the dry weight of G. sulphuraria in this experiment. The removal efficiency (RE) of contaminants was determined by employing the formula
RE = (C0 − C)/C0) × 100%
where C0 represents the initial concentration of the chemical (ppm or mg/L), and C denotes the concentration at time t [27]. Moreover, the Removal rate was calculated with the formula depicted by Zhu et al. [28];
Removal rate (mg/L/d) = (Ct − Ci)/(Tt − Ti)
where Ct (mg/L) and Ci (mg/L) were the concentrations of contaminants at Tt (d) and Ti (d), respectively.

3. Results

3.1. G. sulphuraria’s Contaminant Removal Rate and Efficiency

G. sulphuraria was cultured in urban wastewater and AM (control) at pH 4, using different initial algal biomass concentrations (OD 2, OD 4, and OD 6). It was capable of reducing different contaminants while growing in UWW. Table 2 presents the calculated contaminant removal rate (RR) and efficiency (RE) of G. sulphuraria of each biomass group per day. The data show the effectiveness of G. sulphuraria in UWW treatment.
As the algal biomass increases from OD 2 to OD 6, the removal rate and efficiency for N-NH4+ and N-NH3 improved substantially, with the highest values at OD 6, where over 90% of both N-NH4+ and N-NH3 were removed. P-PO43− removal showed an irregular trend, with moderate removal at OD 2, a sharp drop at OD 4, and a recovery at OD 6. COD removal remained consistently high across all biomass concentrations, with the highest efficiency observed at OD 4. These results indicated that increasing biomass concentration generally enhances the removal of N-NH4+, N-NH3, and organic matter, but P-PO43− removal may follow a more complex pattern.
Figure 1 shows a comparative analysis of the efficiency of each biomass group of G. sulphuraria in reducing different contaminants over time from the UWW that was used as the growing medium. Both biomass groups revealed their potential to reduce the contaminants. However, the observed trends in the graphs underscored the variability in removal rates depending on the biomass concentration.
A paired two-sample t-test was conducted in R Studio (Version 2024.09) [24] to evaluate the effects of biomass concentrations (OD 2, OD 4, and OD 6) on the reduction in various parameters, growth and phycocyanin production (Table 3). The means for each parameter are reported with their corresponding standard deviations. The letters (a, b, c) indicate statistically significant differences between the means within each row, where means followed by different letters are significantly different at p < 0.05.

3.2. Growth and Phycocyanin Production:

The impact of initial algal biomass (OD 6, OD 4, OD 2) on the performance of the alga in AM and UWW over the cultivation period (0, 24, 48, 72, and 96 h) was compared. The growth comparison and phycocyanin levels across the three algal biomass groups are illustrated below (Figure 2).
Across both media, higher inoculum densities consistently yielded significantly greater DW and PC values (OD 6 > OD 4 > OD 2, p < 0.05), except at OD 4, where no significant difference was observed (Table 4). This analysis, supported by maximum growth rate (MGR) calculations, focused on how initial biomass influenced growth dynamics across time, rather than on statistical comparisons between t = 0 and t = 96 h within each medium.

3.3. The Maximum Growth Rate

The maximum growth rate (MGR) of G. sulphuraria in UWW and AM varied with biomass concentrations and time. In OD 2, the highest MGR (0.120 g/L/d) was achieved at 48 h in the UWW. For OD 4, the maximum growth rate (0.178 g/L/d) was observed at 96 h in UWW, while the control AM exhibited a maximum growth rate (0.176 g/L/d) at the same time interval. In OD 6, the maximum growth rate (0.241 g/L/d) was reached within just 24 h in UWW, indicating a rapid biomass increase in higher concentrations and shorter time frames. These results highlight the efficiency of higher biomass levels in pollutant removal and accelerated growth under different conditions.

4. Discussion

This study investigates the contaminant removal potential and growth performance measured as dry weight and phycocyanin synthesis of autotrophic G. sulphuraria in UWW and AM (control) at pH 4 with different initial algal biomass concentrations measured as optical density (OD) at 750 nm (OD 2, OD 4, and OD 6). The difference in initial algal biomass resulted in different pollutant removal rates at the end of the experiment, whereby a significant (p < 0.05) decrease in pollutants was observed as biomass increased. This result is in line with a literature report [12] for microalgae in which the removal rates for nitrogen and phosphorus increase with increasing microalgal biomass density, even though growth also depends on the concentration and types of nutrients, among other abiotic factors. For N-NH4+, the removal rate and efficiency improve significantly with increasing OD, from 5.17 mg/L/d and 62.73% at OD 2 to 7.49 mg/L/d and 90.74% at OD 6. A similar trend is observed with N-NH3, where the removal rate increases from 4.64 mg/L/d at OD 2 to 6.43 mg/L/d at OD 6, and the removal efficiency rises from 66.01% to 91.55%. These outcomes suggest that G. sulphuraria is highly efficient at assimilating nitrogen because of its capacity to utilise N-NH4+ and N-NH3 as nitrogen sources for growth. The increasing trend in nitrogen removal as biomass density increases indicates that higher algal concentrations enhance the uptake of nitrogen compounds from wastewater, making G. sulphuraria a promising candidate for large-scale nitrogen removal. Literature data report comparable findings. For example, Selvaratnam et al. [20] observed a removal rate of 4.85 mg/L/d using autotrophic G. sulphuraria 5587.1 cultured in modified cyanidium medium with primary effluent. However, their study used an initial N-NH4+ concentration of 40 mg/L measured over seven days, and the primary effluent was autoclaved.
In contrast, this study used non-autoclaved primary settled urban wastewater with an initial N-NH4+ concentration of 31.93 mg/L, measured over three days. Despite these differences, the results of this study are superior. Similarly, Tchinda [19] reported an N-NH4+ removal rate of 6.1 mg/L/d by G. sulphuraria from primary effluent with an initial NH4+ concentration of 42 mg/L, which is comparable to our result in the OD 4 algal group. Moreover, Selvaratnam et al. [21] reported removal rates of 4.70 mg/L/day and 4.97 mg/L/day using filter-sterilised primary effluent and non-autoclaved wastewater effluent, respectively, measured over seven days. The N-NH4+ removal rate obtained in this study’s OD 6 algal group was the highest compared to all available results in the literature using autotrophic G. sulphuraria. Abeysiriwardana-Arachchige and Nirmalakhandan et al. [29] reported an N-NH3 removal rate of 4.14 mg/L/day from primary-settled urban wastewater using a mixotrophic strain of G. sulphuraria, which was still lower than the result of this study.
P-PO43− removal shows more variability, with a lower removal rate and efficiency at OD 4 (0.315 mg/L/d and 18.28%) compared to OD 2 and OD 6, where the removal efficiencies are 45.749% and 49.966%, respectively. Several factors might have caused the inconsistency, such as nutrient competition, saturation of phosphate uptake mechanisms, or variations in the biochemical pathways activated under different algal densities. Although the removal efficiency at OD 6 improves, the fluctuation highlights the need for further investigation into optimising conditions for phosphate removal. Galdieria’s phosphate removal is less consistent than nitrogen removal, suggesting that supplementary strategies may be necessary to enhance phosphate extraction. A literature value reported a P-PO43− removal rate of 1.21 mg/L/day using G. sulphuraria CCMEE 5587.1 in modified Cyanidium medium prepared with autoclaved primary effluent, starting with an initial P-PO43− concentration of 10 mg/L and measured over seven days [20]. In contrast, this study began with a lower initial P-PO43− concentration of 7.2 mg/L, measured over a shorter treatment duration (three days), and achieved higher removal rates of 0.789 mg/L/d (OD 2), 0.315 mg/L/d (OD 4), and 0.862 mg/L/d (OD 6). Moreover, Selvaratnam et al. [21] observed P-PO43− removal rates of 1.68 mg/L/day from filter-sterilised primary effluent and 1.47 mg/L/day from non-autoclaved wastewater effluent, both measured over seven days. Furthermore, a PO43- removal rate of 1.4 mg/L/d over 120 days from primary effluent with an initial value of 5.6 mg/L was reported [19]. Our study demonstrated a more efficient removal rate in a shorter time frame.
In terms of COD, the removal rate increases from 47.92 mg/L/d at OD 2 to 51.83 mg/L/d at OD 4 but slightly decreases to 50.58 mg/L/d at OD 6, with efficiencies remaining high around 72–74%. The decrease at OD 6 possibly indicated that the alga was reaching a saturation point in its capacity to degrade organic matter. Despite this, COD removal remains effective, suggesting that G. sulphuraria can contribute substantially to reducing organic pollutants in wastewater, even at higher biomass concentrations. This COD removal is particularly relevant for wastewater treatment, as organic matter reduction is critical for reducing the overall pollutant load.
Both the algal groups reduced the pH of the media as they grew, but the reduction was more pronounced in the less dense OD 2 group (2.837 ± 0.153) than the others, perhaps due to a reduced algal buffering capacity. This reduction was in line with what Oesterhelt et al. [30] reported, whereby G. sulphuraria cultures showed a decreasing pH in response to growth. The OD 4 and OD 6 groups showed a pH reduction of 3.258 ± 0.168 and 3.175 ± 0.171, respectively, which was not significantly different (p > 0.05). The obtained result might suggest that G. sulphuraria with higher OD values might possess fundamental buffering mechanisms that help maintain a favourable pH environment for growth and metabolic activity.
G. sulphuraria showed significantly (p < 0.05) higher growth and phycocyanin levels in UWW than in AM in OD 2 and OD 6 biomass groups. However, the dry weight and phycocyanin levels observed between the two media at OD 4 were not statistically significant. These absences of significant variations at OD 4 might have resulted from an equilibrium state where the initial biomass density allows for similar growth and phycocyanin synthesis efficiency in the nutrient-rich wastewater and the controlled standard AM. At this intermediate level, the alga’s growth demands are met sufficiently by both environments, unlike at the lower and higher biomass levels (OD 2 and OD 6), where the advantages of wastewater with high amounts of nutrients and energy-rich materials become more pronounced [9]. The higher growth and phycocyanin synthesis observed in the UWW than the control AM in this study were in line with other studies that reported higher or comparable growth in different wastewater than the controls [20,21,22,31].
The initial algal biomass concentrations significantly (p < 0.05) affected both growth and phycocyanin levels, giving higher values to the highest biomass groups (OD 6 > OD 4 > OD 2). The enhanced nutrient availability in the UWW is the primary factor for robust algal growth and biomass accumulation. Furthermore, high phycocyanin production at higher biomass levels might be a coping strategy for G. sulphuraria in optimising pigment composition and light-harvesting capabilities that maximise its photosynthetic efficiency and support continued growth. The recorded autotrophic growth (dry weight) of G. sulphuraria ACUF 427 was higher compared to the literature values for autotrophic G. sulphuraria, despite differences in algal strain and density, wastewater type, and culture conditions used, which would lead to differences in the growth of the alga. Our experiment yielded dry weight values of 1.524 ± 0.094 g/L, 2.421 ± 0.407 g/L, and 3.419 ± 0.292 g/L of G. sulphuraria ACUF 427 at OD 2, OD 4 and OD 6, respectively, using non-autoclaved primary effluent over four days. According to Selvaratnam [20], G. sulphuraria CCMEE 5587.1 reached a density of 1.2 g/L using a modified cyanidium medium prepared with autoclaved primary effluent at the end of the exponential phase (4 days). It was also reported that a biomass of less than 0.5 g/L in four days was obtained in an experiment that utilised G. sulphuraria CCMEE 5587.1 in filter-sterilised and non-autoclaved primary effluent [21]. Additionally, it was reported that G. sulphuraria biomass of approximately 1 g/L in seven days was achieved in a single-step treatment of primary effluent [19]. All these reported values were lower than the results of this experiment using G. sulphuraria at OD 2, OD 4 and OD 6. The study also found that the levels of phycocyanin were 0.356 ± 0.040 mg/mL, 0.643 ± 0.121 mg/mL, and 0.905 ± 0.085 mg/mL for OD 2, OD 4, and OD 6, respectively. These values were significantly different from each other (p < 0.05). Comparing these values with the literature data was challenging due to the limited information on phycocyanin synthesis using waste material. For example, Lim et al. [32] reported a phycocyanin yield of 159 ± 14.55 mg/L (0.159 mg/mL) by cultivating G. sulphuraria in an optimised tangerine peel medium. Similarly, di Cicco et al. [33] indicated an average yield of 94 ± 6 mg/L (0.094 mg/mL) for G. phlegrea cultured in raw municipal wastewater. Both of these values were lower than the ones obtained in this study.
The maximum growth rates (MGRs) at different biomass concentrations (OD 2, 4, and 6) illustrate how algal biomass density impacts nutrient uptake and growth in urban wastewater. At OD 2, a lower MGR of 0.120 g/L/d at 48 h suggests slower nutrient utilisation due to reduced biomass. At OD 4, MGR increased to 0.178 g/L/d, but peak growth was delayed to 96 h, indicating that nutrient uptake takes longer despite the rich wastewater. In contrast, OD 6 showed a much higher MGR of 0.241 g/L/d within 24 h, suggesting that higher biomass allows for faster nutrient uptake and metabolism. This result indicated the efficiency of higher biomass in optimising both pollutant removal and growth in wastewater treatment. Comparing the results of OD 4 and OD 6 with literature reports that utilised G. sulphuraria reveals apparent differences in growth rate. At OD 4, the growth rate exceeds that reported by Russo et al. [34] (0.147 g/L/d in 2.5% concentration of cheese whey), Pan et al. [31] (0.159 g/L/d in 20% concentration of raw landfill leachate), and Selvaratnam et al. [20] (0.133 g/L/d in autoclaved primary urban wastewater effluent). OD 6 achieves a much higher growth rate, exceeding all these values quickly. The result demonstrates that OD 6 offers the highest productivity and fastest growth, making it the most efficient condition for maximising algal growth.

5. Conclusions

The findings suggest that G. sulphuraria can be highly effective in removing nitrogen and organic matter from wastewater, especially at higher biomass concentrations, with some variability in phosphate removal. The high removal efficiencies for N-NH4+, N-NH3, and COD highlight the potential of Galdieria for wastewater bioremediation. However, the inconsistent phosphate removal suggests that further research is needed to fully understand nutrient uptake dynamics at varying algal concentrations. Additionally, the slight decrease in COD removal at the highest biomass level indicates a possible limit to the system’s capacity, suggesting that there may be an optimal biomass concentration for maximising removal efficiency without oversaturation. Furthermore, enhanced growth and phycocyanin production were recorded in urban wastewater compared to the control (Allen Medium), with significant differences (p < 0.05) across all optical densities. The maximum growth rates were achieved at OD 6, underscoring the advantage of higher optical densities for biomass production and pollutant removal. These findings support the use of G. sulphuraria in sustainable wastewater management strategies, particularly in applications requiring effective nutrient remediation and biomass generation. Further research may explore this approach’s long-term viability and scalability in diverse wastewater treatment scenarios.

Author Contributions

Conceptualisation, B.R., M.I. and C.C.; Methodology, B.R., M.I. and C.C.; Data curation, B.R.; Writing—original draft preparation, B.R.; Writing—review and editing, B.R., M.I. and C.C. All authors have read and agreed to the published version of the manuscript.

Funding

This research received no funding.

Data Availability Statement

The raw data are available from the corresponding author upon a reasonable request.

Acknowledgments

The authors would like to thank the “algal collection of the University of Federico II, Naples, Italy” for the provision of G. sulphuraria ACUF 427.

Conflicts of Interest

The authors declare that they have no conflicts of interest.

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Phycology 05 00040 g001aPhycology 05 00040 g001b
Figure 1. Comparative analyses of different concentrations of G. sulphuraria in contaminant removal. (Error bars show std. deviation and the number of independent replicates, n, is equal to 3).
Figure 1. Comparative analyses of different concentrations of G. sulphuraria in contaminant removal. (Error bars show std. deviation and the number of independent replicates, n, is equal to 3).
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Phycology 05 00040 g002aPhycology 05 00040 g002b
Figure 2. Dry weight and phycocyanin of G. sulphuraria in AM and UWW at different initial biomass concentrations. (Error bars show std. deviation and the number of independent replicates, n, is equal to 3).
Figure 2. Dry weight and phycocyanin of G. sulphuraria in AM and UWW at different initial biomass concentrations. (Error bars show std. deviation and the number of independent replicates, n, is equal to 3).
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Table 1. Characteristics of the Urban Wastewater.
Table 1. Characteristics of the Urban Wastewater.
UWW CompositionValues (ppm)
Ammonium nitrogen (N-NH4+)31.93
Nitrogen dioxide (N-NO2)0.1
Nitrous oxide (N-N2O)0.1
Ammonia nitrogen (N-NH3)24.8
Phosphate phosphorus P-PO43−7.2
COD (mg/L)279
pH7.5
Table 2. The contaminant removal rate and efficiency of G. sulphuraria in UWW.
Table 2. The contaminant removal rate and efficiency of G. sulphuraria in UWW.
ParametersOD 2OD 4OD 6
RR (mg/L/d)RE (%)RR (mg/L/d)RE (%)RR (mg/L/d)RE (%)
N-NH4+5.17 ± 0.3662.73 ± 4.355.8 ± 0.6970.61 ± 8.417.49 ± 0.1990.74 ± 2.26
N-NH34.64 ± 0.2866.01 ± 3.965.14 ± 0.5473.20 ± 7.676.43 ± 0.1491.55 ± 2.05
P-PO43−0.79 ± 0.0445.75 ± 0.590.32 ± 0.0618.28 ± 3.590.86 ± 0.0649.97 ± 3.34
COD47.92 ± 1.668.70 ± 2.3351.83 ± 1.6674.31 ± 2.3850.58 ± 0.7272.52 ± 1.03
Note: The number of independent replicates, n, is equal to 3.
Table 3. The Effect of initial G. sulphuraria biomass concentrations on its growth and ability to reduce various pollutants from UWW.
Table 3. The Effect of initial G. sulphuraria biomass concentrations on its growth and ability to reduce various pollutants from UWW.
Parameters
(ppm)		Biomass
(OD 2)		Biomass
(OD 4)		Biomass
(OD 6)
N-NH421.949 ± 7.418 a18.582 ± 6.986 b12.433 ± 7.903 c
N-NH317.045 ± 5.760 a14.679 ± 5.562 b9.654 ± 6.137 c
P-PO43−4.539 ± 0.599 a5.726 ± 0.573 b3.775 ± 0.610 c
pH2.837 ± 0.153 a3.258 ± 0.168 b3.175 ± 0.171 b
COD (mg/L)104.750 ± 15.882 a82 ± 10.954 b75.333 ± 9.948 b
DW (g/L)1.524 ± 0.094 a2.421 ± 0.407 b3.419 ± 0.292 c
PC (mg/mL)0.356 ± 0.040 a0.643 ± 0.121 b0.905 ± 0.085 c
Note: Means in the same row followed by different letters are significantly different (p < 0.05). (The number of independent replicates, n, is equal to 3).
Table 4. Comparison of growth parameters of G. sulphuraria of different biomass in AM and UWW.
Table 4. Comparison of growth parameters of G. sulphuraria of different biomass in AM and UWW.
VariablesValuesOD 2OD 4OD 6
AMUWWp-ValuesAMUWWp-ValuesAMUWWp-Values
DW (g/L)Mean1.3081.5245.34 × 10−62.2422.4210.2163.0933.4190.005
SD0.1140.094 0.3680.407 0.3060.292
PC (g/L)Mean0.2940.3560.00010.6030.6430.3530.8190.9050.019
SD0.0350.040 0.1140.121 0.1030.085
The number of independent replicates, n, is equal to 3.
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Retta, B.; Iovinella, M.; Ciniglia, C. Optimising Galdieria sulphuraria ACUF 427 Biomass for Enhanced Urban Wastewater Treatment: Evaluating Pollutant Removal Efficiency, Algal Growth, and Phycocyanin Production. Phycology 2025, 5, 40. https://doi.org/10.3390/phycology5030040

AMA Style

Retta B, Iovinella M, Ciniglia C. Optimising Galdieria sulphuraria ACUF 427 Biomass for Enhanced Urban Wastewater Treatment: Evaluating Pollutant Removal Efficiency, Algal Growth, and Phycocyanin Production. Phycology. 2025; 5(3):40. https://doi.org/10.3390/phycology5030040

Chicago/Turabian Style

Retta, Berhan, Manuela Iovinella, and Claudia Ciniglia. 2025. "Optimising Galdieria sulphuraria ACUF 427 Biomass for Enhanced Urban Wastewater Treatment: Evaluating Pollutant Removal Efficiency, Algal Growth, and Phycocyanin Production" Phycology 5, no. 3: 40. https://doi.org/10.3390/phycology5030040

APA Style

Retta, B., Iovinella, M., & Ciniglia, C. (2025). Optimising Galdieria sulphuraria ACUF 427 Biomass for Enhanced Urban Wastewater Treatment: Evaluating Pollutant Removal Efficiency, Algal Growth, and Phycocyanin Production. Phycology, 5(3), 40. https://doi.org/10.3390/phycology5030040

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