The palm oil industry is one of the key players in contributing to Malaysia’s economy. Palm oil mill effluent (POME), a significant by-product of the oil extraction process, requires mandatory remediation to ensure proper treatment and disposal. Bioremediation using microalgae is a cost-effective and sustainable approach. This study aims to utilise pure and mixed microalgal species, Chlorella vulgaris and Tetradesmus bernardii, in phycoremediation and biomass production in different concentrations of POME (20%, 40%, 60%, and 80%). Cultivation of microalgae was carried out in 200 mL medium with pH 7–7.8, room temperature of 25±1°C for 21 days and continuous light illumination at 2000 lux. The highest biomass productivity was observed in 20% POME for mixed microalgae (mean = 0.1733 mg.mL^-1 ± 0.0057), followed by C. vulgaris (0.1633 mg.mL^-1 ± 0.0057) and T. bernardii (0.1603 mg.mL^-1 ± 0.0020). Similarly, the highest nutrient removal was observed in 20% POME for mixed microalgae (COD:66.9801%, TN:86.9565%, TP:86.9655%), followed by C. vulgaris and T. bernardii. The results showed positive effects on growth, increased biomass production, and nutrient removal, with 20% POME being the optimal concentration for microalgae. Valuable by-products, such as high-quality pigments and biomass, are also generated by applying microalgae for remediation. Mixed microalgae are superior in the remediation of POME compared to single-culture algae. Treating wastewater through microalgal bioremediation is highly efficient in nutrient removal. This research has contributed towards the use of mixed microalgae to achieve effective nutrient removal and biomass for future industrial applications.

• Aburai, N., Ohkubo, S., Miyashita, H., & Abe, K. (2013). Composition of carotenoids and identification of aerial microalgae isolated from the surface of rocks in mountainous districts of Japan. Algal Research, 2(3), 237-243. https://doi.org/10.1016/j.algal.2013.03.001

• Ahmad, A., Bhat, A. H., & Buang, A. (2017). Immobilized Chlorella vulgaris for efficient palm oil mill effluent treatment and heavy metals removal. Desalination and Water Treatment, 81, 105-117. https://doi.org/10.5004/dwt.2017.20991

• Ahmad, A., Buang, A., & Bhat, A. H. (2016). Renewable and sustainable bioenergy production from microalgal co-cultivation with palm oil mill effluent (POME): A review. Renewable and Sustainable Energy Reviews, 65, 214–234. https://doi.org/10.1016/j.rser.2016.06.084

• Ahmad, M. T., Shariff, M., Yusoff, F. M., Goh, Y. M., & Banerjee, S. (2020). Applications of microalga Chlorella vulgaris in aquaculture. Reviews in Aquaculture, 12(1), 328-346. https://doi.org/10.1111/raq.12320

• Ahmed, S. F., Mofijur, M., Parisa, T. A., Islam, N., Kusumo, F., Inayat, A., Van, G. L., Badruddin, I. A., Khan, T. M. Y., & Ong, H. C. (2022). Progress and challenges of contaminate removal from wastewater using microalgae biomass. Chemosphere, 286(1), Article 131656. https://doi.org/10.1016/j.chemosphere.2021.131656

• Al-Amshawee, S. K., Yunus, M. Y., & Azoddein, A. A. (2020). A review study of biofilm bacteria and microalgae bioremediation for palm oil mill effluent: Possible approach. IOP Conference Series: Materials Science and Engineering 736(2), Article 02203. https://doi.org/10.1088/1757-899X/736/2/022034

• Al-Khiat, S. H., Bukhari, N. A., Ameen, F., & Abdel-Raouf, N. (2023). Comparison of the microalgae Phormidium tenue and Chlorella vulgaris as biosorbents of Cd and Zn from aqueous environments. Environmental Research, 235, Article 116675. https://doi.org/10.1016/j.envres.2023.116675

• Aminot, A., & Rey, F. (2000). Standard procedure for the determination of chlorophyll a by spectroscopic methods. International Council for the Exploration of the Sea, 112, Article 25.

• Anto, S., Mukherjee, S. S., Muthappa, R., Mathimani, T., Deviram, G., Kumar, S. S., Verma, T. N., & Pugazhendhi, A. (2020). Algae as green energy reserve: Technological outlook on biofuel production. Chemosphere, 242, Article 125079. https://doi.org/10.1016/j.chemosphere.2019.125079

• Arashiro, L. T., Boto-Ordóñez, M., Van Hulle, S. W., Ferrer, I., Garfí, M., & Rousseau, D. P. (2020). Natural pigments from microalgae grown in industrial wastewater. Bioresource Technology, 303, Article 122894. https://doi.org/10.1016/j.biortech.2020.122894

• Azmi, N. S., & Yunos, K. F. M. (2014). Wastewater treatment of palm oil mill effluent (POME) by ultrafiltration membrane separation technique coupled with adsorption treatment as pre-treatment. Agriculture and Agricultural Science Procedia, 2, 257-264. https://doi.org/10.1016/j.aaspro.2014.11.037

• Bala, J. D., Lalung, J., & Ismail, N. (2015). Studies on the reduction of organic load from palm oil mill effluent (POME) by bacterial strains. International Journal of Recycling of Organic Waste in Agriculture, 4, 1-10. https://doi.org/10.1007/s40093-014-0079-6

• Bauer, D. E., Conforti, V., Ruiz, L., & Gómez, N. (2012). An in-situ test to explore the responses of Scenedesmus acutus and Lepocinclis acus as indicators of the changes in water quality in lowland streams. Ecotoxicology and Environmental Safety, 77, 71-78. https://doi.org/10.1016/j.ecoenv.2011.10.021

• Begum, H., Yusoff, F. M., Banerjee, S., Khatoon, H., & Shariff, M. (2016). Availability and utilization of pigments from microalgae. Critical Reviews in Food Science and Nutrition, 56(13), 2209-2222. https://doi.org/10.1080/10408398.2013.764841

• Bradley, I. M., Pinto, A. J., & Guest, J. S. (2016). Design and evaluation of Illumina MiSeq-compatible, 18S rRNA gene-specific primers for improved characterization of mixed phototrophic communities. Applied and Environmental Microbiology, 82(19), 5878-5891. https://doi.org/10.1128/AEM.01630-16

• Cheah, W. Y., Show, P. L., Juan, J. C., Chang, J. S., & Ling, T. C. (2018). Microalgae cultivation in palm oil mill effluent (POME) for lipid production and pollutants removal. Energy Conversion and Management, 174, 430-438. https://doi.org/10.1016/j.enconman.2018.08.057

• Chew, C. L., Ng, C. Y., Hong, W. O., Wu, T. Y., Lee, Y. Y., Low, L. E., Kong, P. S., & Chan, E. S. (2021). Improving sustainability of palm oil production by increasing oil extraction rate: A review. Food and Bioprocess Technology, 14, 573-586. https://doi.org/10.1007/s11947-020-02555-1

• Ding, G. T., Yaakob, Z., Takriff, M. S., Salihon, J., & Rahaman, M. S. A. (2016). Biomass production and nutrients removal by a newly-isolated microalgal strain Chlamydomonas sp. in palm oil mill effluent (POME). International Journal of Hydrogen Energy, 41(8), 4888-4895. https://doi.org/10.1016/j.ijhydene.2015.12.010

• Dominic, D., & Baidurah, S. (2022). Recent developments in biological processing technology for palm oil mill effluent treatment. Biology, 11(4), Article 525. https://doi.org/10.3390/biology11040525

• El-Din, N. G. S., El-Sheekh, M. M., El-Kassas, H. Y., Essa, D. I., & El-Sherbiny, B. A. (2022). Biological indicators as tools for monitoring water quality of a hot spot area on the Egyptian Mediterranean Coast. Arabian Journal of Geosciences, 15(18), Article 1485. https://doi.org/10.1007/s12517-022-10701-6

• Emparan, Q., Jye, Y. S., Danquah, M. K., & Harun, R. (2020). Cultivation of Nannochloropsis sp. microalgae in palm oil mill effluent (POME) media for phycoremediation and biomass production: Effect of microalgae cells with and without beads. Journal of Water Process Engineering, 33, Article 101043. https://doi.org/10.1016/j.jwpe.2019.101043

• Fallahi, A., Hajinajaf, N., Tavakoli, O., & Sarrafzadeh, M. H. (2020). Cultivation of mixed microalgae using municipal wastewater: biomass productivity, nutrient removal, and biochemical content. Iranian Journal of Biotechnology, 18(4), Article e2586. https://doi.org/10.30498/IJB.2020.2586

• Fernando, J. S. R., Premaratne, M., Dinalankara, D. M. S. D., Perera, G. L. N. J., & Ariyadasa, T. U. (2021). Cultivation of microalgae in palm oil mill effluent (POME) for astaxanthin production and simultaneous phycoremediation. Journal of Environmental Chemical Engineering, 9(4), Article 105375. https://doi.org/10.1016/j.jece.2021.105375

• Goh, P. S., Lau, W. J., Ismail, A. F., Samawati, Z., Liang, Y. Y., & Kanakaraju, D. (2022). Microalgae-enabled wastewater treatment: A sustainable strategy for bioremediation of pesticides. Water, 15(1), Article 70. https://doi.org/10.3390/w15010070

• Hadiyanto, & Nur, M. M. A. (2014). Lipid extraction of microalga Chlorella sp. cultivated in palm oil mill effluent (POME) medium. World Applied Sciences Journal, 31(5), 959-967. https://doi.org/10.5829/idosi.wasj.2014.31.05.2006

• Hariz, H. B., Takriff, M. S., Ba-Abbad, M. M., Yasin, N. H. M., & Hakim, N. I. N. M. (2018). CO2 fixation capability of Chlorella sp. and its use in treating agricultural wastewater. Journal of Applied Phycology, 30, 3017-3027. https://doi.org/10.1007/s10811-018-1488-0

• Hariz, H. B., Takriff, M. S., Yasin, N. H. M., Ba-Abbad, M. M., & Hakimi, N. I. N. M. (2019). Potential of the microalgae-based integrated wastewater treatment and CO2 fixation system to treat palm oil mill effluent (POME) by indigenous microalgae; Scenedesmus sp. and Chlorella sp. Journal of Water Process Engineering, 32, Article 100907. https://doi.org/10.1016/j.jwpe.2019.100907

• Haryati, Z., Subramaniam, V., Noor, Z. Z., Hashim, Z., Loh, S. K., & Aziz, A. A. (2022). Social life cycle assessment of crude palm oil production in Malaysia. Sustainable Production and Consumption, 29, 90-99. https://doi.org/10.1016/j.spc.2021.10.002

• Hazman, N. A. S., Yasin, N. H. M., Takriff, M. S., Hasan, H. A., Kamarudin, K. F., & Hakimi, N. I. N. M. (2018). Integrated palm oil mill effluent treatment and CO2 sequestration by microalgae. Sains Malaysiana, 47(7), 1455-1464. https://doi.org/10.17576/jsm-2018-4707-13

• Jasni, J., Arisht, S. N., Yasin, N. H. M., Abdul, P. M., Lin, S. K., Liu, C. M., & Takriff, M. S. (2020). Comparative toxicity effect of organic and inorganic substances in palm oil mill effluent (POME) using native microalgae species. Journal of Water Process Engineering, 34, Article 101165. https://doi.org/10.1016/j.jwpe.2020.101165

• Johan, F., Jafri, M. Z., Lim, H. S., & Maznah, W. W. (2014, December 9-12). Laboratory measurement: Chlorophyll-a concentration measurement with acetone method using spectrophotometer. [Paper presentation]. IEEE International Conference on Industrial Engineering and Engineering Management, Selangor, Malaysia. https://doi.org/10.1109/IEEM.2014.7058737

• Justine, I., Chin, G. J. W. L., Yong, W. T. L., & Misson, M. (2023). Characterization and optimization of Rhodotorula toruloides and Ankistrodesmus falcatus co-culture in palm oil mill effluent for efficient COD removal and lipid production. Biocatalysis and Agricultural Biotechnology, 51, Article 102782. https://doi.org/10.1016/j.bcab.2023.102782

• Kamarudin, K. F., Yaakob, Z., Rajkumar, R., & Tasirin, S. M. (2013). Bioremediation of palm oil mill effluents (POME) using Scenedesmus dimorphus and Chlorella vulgaris. Advanced Science Letters, 19(10), 2914-2918. https://doi.org/10.1166/asl.2013.5044

• Kamyab, H., Din, M. F. M., Keyvanfar, A., Majid, M. Z. A., Talaiekhozani, A., Shafaghat, A., & Ismail, H. H. (2015). Efficiency of microalgae Chlamydomonas on the removal of pollutants from palm oil mill effluent (POME). Energy Procedia, 75, 2400-2408. https://doi.org/10.1016/j.egypro.2015.07.190

• Kamyab, H., Fadhil, M., Lee, C., Ponraj, M., Soltani, M., & Eva, S. (2014). Micro-macro algal mixture as a promising agent for treating POME discharge and its potential use as animal feed stock enhancer. Jurnal Teknologi, 68(5), 1-4.

• Kamyab, H., Yuzir, M. A., Riyadi, F. A., Ostadrahimi, A., Khademi, T., Ghfar, A. A., & Kirpichnikova, I. (2022). Electrochemical oxidation of palm oil mill effluent using platinum as anode: Optimization using response surface methodology. Environmental Research, 214(3), Article 113993. https://doi.org/10.1016/j.envres.2022.113993

• Karim, A., Islam, M. A., Khalid, Z. B., Yousuf, A., Khan, M. M. R., & Faizal, C. K. M. (2021). Microbial lipid accumulation through bioremediation of palm oil mill effluent using a yeast-bacteria co-culture. Renewable Energy, 176, 106-114. https://doi.org/10.1016/j.renene.2021.05.055

• Khalid, A. A. H., Yaakob, Z., Abdullah, S. R. S., & Takriff, M. S. (2016). Enhanced growth and nutrients removal efficiency of Characium sp. cultured in agricultural wastewater via acclimatized inoculum and effluent recycling. Journal of Environmental Chemical Engineering, 4(3), 3426-3432. https://doi.org/10.1016/j.jece.2016.07.020

• Khatun, R., Reza, M. I. H., Moniruzzaman, M., & Yaakob, Z. (2017). Sustainable oil palm industry: The possibilities. Renewable and Sustainable Energy Reviews, 76, 608-619. https://doi.org/10.1016/j.rser.2017.03.077

• Kumaran, M., Palanisamy, K. M., Bhuyar, P., Maniam, G. P., Rahim, M. H. A., & Govindan, N. (2023). Agriculture of microalgae Chlorella vulgaris for polyunsaturated fatty acids (PUFAs) production employing palm oil mill effluents (POME) for future food, wastewater, and energy nexus. Energy Nexus, 9, Article 100169. https://doi.org/10.1016/j.nexus.2022.100169

• Lau, P. S., Tam, N. F. Y., & Wong, Y. S. (1995). Effect of algal density on nutrient removal from primary settled wastewater. Environmental Pollution, 89(1), 59-66. https://doi.org/10.1016/0269-7491(94)00044-E

• Medvedeva, L. N., Zorkina, O. V., & Moskovets, M. V. (2022). Use of Chlorella vulgaris as a dietary supplement for quails bred at private farms. Journal of Agronomy and Animal Industries, 17(4), 499-513. https://doi.org/10.22363/2312-797X-2022-17-4-499-513

• Michalak, I., Baśladyńska, S., Mokrzycki, J., & Rutkowski, P. (2019). Biochar from a freshwater macroalga as a potential biosorbent for wastewater treatment. Water, 11(7), Article 1390. https://doi.org/10.3390/w11071390

• Mohammed, R. R. (2013). Decolourisation of biologically treated palm oil mill effluent (POME) using adsorption technique. International Refereed Journal of Engineering and Science, 2(10), 1-11.

• Morais, F. P., Simões, R. M. S., & Curto, J. M. R. (2020). Biopolymeric delivery systems for cosmetic applications using Chlorella vulgaris algae and tea tree essential oil. Polymers, 12(11), Article 2689. https://doi.org/10.3390/polym12112689

• Nakarmi, K. J., Daneshvar, E., Mänttäri, M., & Bhatnagar, A. (2023). Removal and recovery of nutrients from septic tank wastewater using microalgae: Key factors and practical implications. Journal of Environmental Management, 345, Article 118922. https://doi.org/10.1016/j.jenvman.2023.118922

• Nobre, B. P., Villalobos, F., Barragan, B. E., Oliveira, A. C., Batista, A. P., Marques, P. A. S. S., Mendes, R. L., Savova, H., Palavra, A. F., & Gouveia, L. (2013). A biorefinery from Nannochloropsis sp. microalga–extraction of oils and pigments. Production of biohydrogen from the leftover biomass. Bioresource Technology, 135, 128-136. https://doi.org/10.1016/j.biortech.2012.11.084

• Orusmurzaeva, Z., Maslova, A., Tambieva, Z., Sadykova, E., Askhadova, P., Umarova, K., Merzhoeva, A., Albogachieva, K., Ulikhanyan, K., & Povetkin, S. (2022). Investigation of the chemical composition and physicochemical properties of Chlorella vulgaris biomass treated with pulsed discharges technology for potential use in the food industry. Slovak Journal of Food Sciences, 16, 777-789.

• Otondo, A., Kokabian, B., Stuart-Dahl, S., & Gude, V. G. (2018). Energetic evaluation of wastewater treatment using microalgae, Chlorella vulgaris. Journal of Environmental Chemical Engineering, 6(2), 3213-3222. https://doi.org/10.1016/j.jece.2018.04.064

• Qin, L., Wang, Z., Sun, Y., Shu, Q., Feng, P., Zhu, L., & Yuan, Z. (2016). Microalgae consortia cultivation in dairy wastewater to improve the potential of nutrient removal and biodiesel feedstock production. Environmental Science and Pollution Research, 23, 8379-8387. https://doi.org/10.1007/s11356-015-6004-3

• Ratnasari, A., Zaidi, N. S., Syafiuddin, A., Boopathy, R., Kueh, A. B. H., Amalia, R., & Prasetyo, D. D. (2021). Prospective biodegradation of organic and nitrogenous pollutants from palm oil mill effluent by acidophilic bacteria and archaea. Bioresource Technology Reports, 15, Article 100809. https://doi.org/10.1016/j.biteb.2021.100809

• Resdi, R., Lim, J. S., & Idris, A. (2021). Batch kinetics of nutrients removal for palm oil mill effluent and recovery of lipid by Nannochloropsis sp. Journal of Water Process Engineering, 40, Article 101767. https://doi.org/10.1016/j.jwpe.2020.101767

• Rowland, D., Zanello, G., Waliyo, E., & Ickowitz, A. (2022). Oil palm and gendered time use: A mixed-methods case study from West Kalimantan, Indonesia. Forest Policy and Economics, 137, Article 102682. https://doi.org/10.1016/j.forpol.2021.102682

• Saidu, H., Jamaluddin, H., & Mohamad, S. E. (2017). Nutrient removal and biokinetic study of freshwater microalgae in palm oil mill effluent (POME). Indian Journal of Science and Technology, 10(24), 1-10. https://doi.org/10.17485/ijst/2017/v10i24/114584

• Salazar, J., Santana-Sánchez, A., Näkkilä, J., Sirin, S., & Allahverdiyeva, Y. (2023). Complete N and P removal from hydroponic greenhouse wastewater by Tetradesmus obliquus: A strategy for algal bioremediation and cultivation in Nordic countries. Algal Research, 70, Article 102988. https://doi.org/10.1016/j.algal.2023.102988

• Simon, D., & Helliwell, S. (1998). Extraction and quantification of chlorophyll a from freshwater green algae. Water Research, 32(7), 2220-2223. https://doi.org/10.1016/S0043-1354(97)00452-1

• Singh, R. P., Ibrahim, M. H., Esa, N., & Iliyana, M. S. (2010). Composting of waste from palm oil mill: A sustainable waste management practice. Reviews in Environmental Science and Biotechnology, 9(4), 331–344. https://doi.org/10.1007/s11157-010-9199-2

• Song, X., Bo, Y., Feng, Y., Tan, Y., Zhou, C., Yan, X., Ruan, R., Xu, Q., & Cheng, P. (2022). Potential applications for multifunctional microalgae in soil improvement. In Frontiers in Environmental Science, 10, Article 1035332. https://doi.org/10.3389/fenvs.2022.1035332

• Takaichi, S. (2011). Carotenoids in algae: Distributions, biosynthesis and functions. Marine Drugs, 9(6), 1101-1118. https://doi.org/10.3390/md9061101

• Talib, S. L. A., Yasin, N. H. M., Takriff, M. S., & Japar, A. S. (2023). Comparative studies on phycoremediation efficiency of different water samples by microalgae. Journal of Water Process Engineering, 52, Article 103584. https://doi.org/10.1016/j.jwpe.2023.103584

• Tan, K. A., Lalung, J., Wijaya, D., Ismail, N., Omar, W. M. W., Wabaidur, S. M., Siddiqui, M. R., Alam, M., & Rafatullah, M. (2022). Removal of nutrients by using green microalgae from lab-scale treated palm oil mill effluent. Fermentation, 8(11), Article 658. https://doi.org/10.3390/fermentation8110658

• Udaiyappan, A. F. M., Hasan, H. A., Takriff, M. S., Abdullah, S. R. S., Yasin, N. H. M., & Ji, B. (2021). Cultivation and application of Scenedesmus sp. strain UKM9 in palm oil mill effluent treatment for enhanced nutrient removal. Journal of Cleaner Production, 294, Article 126295. https://doi.org/10.1016/j.jclepro.2021.126295

• Udaiyappan, A. F., Hasan, H. A., Takriff, M. S., Abdullah, S. R. S., Maeda, T., Mustapha, N. A., Mohd Yasin, N. H., & Hakimi, N. I. N. M. (2020). Microalgae-bacteria interaction in palm oil mill effluent treatment. Journal of Water Process Engineering, 35, Article 101203. https://doi.org/10.1016/j.jwpe.2020.101203

• Velmurugan, A., & Muthukaliannan, G. K. (2022). Genetic manipulation for carotenoid production in microalgae an overview. Current Research in Biotechnology, 4, 221-228. https://doi.org/10.1016/j.crbiot.2022.03.005

• Wynne, M. J., & Hallan, J. K. (2015). Reinstatement of Tetradesmus GM Smith (Sphaeropleales, Chlorophyta). Feddes Repertorium, 126(3‐4), 83-86. https://doi.org/10.1002/fedr.201500021

• Zhao, D., Cheah, W. Y., Lai, S. H., Ng, E. P., Khoo, K. S., Show, P. L., & Ling, T. C. (2023). Symbiosis of microalgae and bacteria consortium for heavy metal remediation in wastewater. Journal of Environmental Chemical Engineering, 11(3), Article 109943. https://doi.org/10.1016/j.jece.2023.109943