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INTRODUCTION: This study focuses on metabolic profiling of a robust marine green algal strain Picochlorum sp. MCC39 that exhibits resilient growth under diverse outdoor open pond conditions. Given its potential for producing high-value chemicals through metabolic engineering, understanding its metabolic dynamics is crucial for pathway modification. OBJECTIVES: This study primarily aimed to investigate the metabolic response of Picochlorum sp. to environmental changes. Unlike heterotrophs, algae are subject to diurnal light and temperature, which affect their growth rates and metabolism. Using an environmental photobioreactor (ePBR), we explored how the algal strain adapts to fluctuations in light intensities and temperature within a simulated pond environment. METHODS: We performed a reverse phase ion pairing-LC/MS-MS based metabolome profiling of the MCC39 strain cultivated in simulated pond conditions in ePBR. The experimental setup included diurnal and bi-seasonal variations in light intensities and temperature. RESULTS: The metabolome profile revealed significant differences in 85 metabolites, including amino acids, carboxylic acids, sugar phosphates, purines, pyrimidines, and dipeptides, which exhibited up to 25-fold change in relative concentration with diurnal variations. Seasonal variations also influenced the production of storage molecules, revealing a discernible pattern. The accumulation pattern of metabolites involved in cellular wall formation and energy generation indicated a well-coordinated initiation of photosynthesis and the Calvin cycle with the onset of light. CONCLUSION: The results contribute to a deeper understanding of the adaptability and metabolic response of Picochlorum sp., laying the groundwork for future advancements in algal strain modification.

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Spirulina is capable of using light energy and fixing carbon dioxide to synthesize a spectrum of organic substances, including proteins, polysaccharides, and unsaturated fatty acids, making it one of the most coveted food resources for humanity. Conventionally, Spirulina products are formulated into algal powder tablets or capsules. However, the processing and preparation of these products, involving screw pump feeding, extrusion, high-speed automation, and high-temperature dewatering, often result in the rupture of cell filaments, cell fragmentation, and the unfortunate loss of vital nutrients. In contrast, fresh Spirulina, cultivated within a closed photobioreactor and transformed into an edible delight through harvesting, washing, filtering, and sterilizing, presents a refreshing taste and odor. It is gradually earning acceptance as a novel health food among the general public. This review delves into the manufacturing processes of fresh Spirulina, analyzes its nutritional advantages over conventional algal powder, and ultimately prospects the avenues for fresh Spirulina's application in modern food processing. The aim is to provide valuable references for the research and development of new microalgal products and to propel the food applications of microalgae forward.

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Wastewater resources can be used to produce microbial protein for animal feed or organic fertiliser, conserving food chain resources. This investigation hasemployed thefermented sewage to photoheterotrophically grown purple non-sulfur bacteria (PNSB) in a 2.5 m3 pilot-scaleraceway-pond with infrared light to produce proteinaceous biomass. Fermented sewage with synthetic media consisting of sodium acetate and propionic acids at a surface-to-volume (S/V) ratio of 10 m2/m3 removed 89%, 93%, and 81% of chemical oxygen demand, ammonium nitrogen, and orthophosphate, respectively; whereas respective removal in fermented sewage alone without synthetic media was 73%, 73%, and 72% during batch operation of 120 h. The biomass yield of 0.88-0.95 g CODbiomass /g CODremoved with protein content of 40.3 ± 0.3%-43.9 ± 0.2% w/w was obtained for fermented sewage with synthetic media. The results revealed enhanced possibility of scaling-up the raceway reactor to recover resources from municipal wastewater and enable simultaneous high-rate PNSB single-cell protein production.

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The failure of mass production of Spirulina plateaus can be attributed to an imbalance of nutrients (C:N) and an increase in accumulated sodium ions, coupled with the traditional harvesting process. The current study aims at the recovery of stressed and red cultures of Spirulina platensis as well as enhanced phycocyanin accumulation. The stressed Spirulina platensis cultures were obtained from a local Egyptian Spirulina production farms, which were further subjected to water analyses after removing the Spirulina biomass. Optimization was performed within 300-ml water path photobioreactor. Spirulina platensis samples were incubated with Zarrouk medium comparing with those modified using ammonium bicarbonate or ammonium acetate instead of sodium bicarbonate. Continuous batching was performed every 12 days during three sequenced batches. Growth measurements (dry weight and pigments) were performed along the incubation time. It was found that carbon content of the growth medium seems to be more effective in Spirulina growth and biomass characteristics. Under different carbon sources, acetate resulted in the maximum dry weight of 1.48 g·l-1 and recovery percentage of 463.3%. Such effect was extended along the different incubation batches. Various carbon concentrations revealed that moderate concentration of carbon in the form of acetate (0.699 g·l-1) leads to the maximum growth under the same nitrogen content. A similar trend was observed with chlorophyll and phycocyanin accumulation, while carotenoids showed the opposite manner.

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To improve the durability of the photobioreactor antibiofouling surface for microalgal cultivation, a series of photoreactive poly(2,2,2-trifluoroethyl methacrylate) (PTFEMA) were successfully synthesized and used to modify ethylene-vinyl acetate (EVA) films by a surface coating and UV light grafting method. Fourier transform infrared (FT-IR) spectra, X-ray photoelectron spectroscopy analysis (XPS) and fluorescence microscopy results indicated that PTFEMA were fixed successfully onto the EVA film surface through a covalent bond. During the microalgal adhesion assay, the number of EVA-PTFEMA film-adhered microalgae was 41.4% lower than that of the EVA film. Moreover, the number of microalgae attached to the EVA-PTFEMA film decreased by 61.7% after cleaning, while that of EVA film decreased by only 49.1%. It was found that the contact angle of EVA-PTFEMA film surface increased, and remained stable when immersed in acid and alkali solution for up to 90 days. HIGHLIGHTSDurable photobioreactor antibiofouling surfaces for microalgal cultivation were prepared successfully.The contact angle of antibiofouling coating surface remained stable in acid and base environment for 90 days.The attached microalgae on antibiofouling surface decreased 41.4% than those of unmodified surface.The attached microalgae on antibiofouling surface could be cleaned by 61.7% through changing the flow velocity of microalgal suspension.

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Microalgae have emerged as a promising source of biomass to produce renewable biofuels due to their ability to synthesize high-energy density compounds of commercial interest. This study proposes an approach for pilot-scale oil extraction, purification by fractional distillation, hydrocarbon characterization by gas chromatography coupled to mass spectrometry (GC-MS), evaluation of physicochemical parameters of the produced hydrocarbons, preliminary cost analysis, and challenges and future opportunities for green diesel on a commercial scale. Here, the microalgae Tetradesmus obliquus was cultivated in 12 m³ photobioreactors using biodigested swine waste as a culture medium. The resulting biomass was subjected to drying and harvesting, followed by oil extraction using a hot solvent extraction method, followed by distillation to purify the compounds. Three different extraction and distillation experiments were conducted, each using different solvent combinations. The results obtained revealed that extraction with a solvent blend, composed of hexane and ethanol, provided more significant yields compared to extraction with pure hexane. GC-MS analysis showed the presence of alkanes and alkenes in the oil samples, and the proportion of solvent used in the extraction directly influenced the production of alkanes. Additionally, specific hydrocarbons such as 4-methyl-1-decene, 8-heptadecene, 1-pentadecene, 9-heneicosene, and 2-dodecene were identified. The evaluation of the physicochemical parameters demonstrated that the calorific value of the distilled oil samples is within the range of typical values for petroleum diesel. However, it was observed that the distilled oil samples had higher sulfur content compared to conventional diesel. Regarding the cost analysis, it was found that it varies depending on the experimental conditions. In particular, the process using a solvent mixture of 70% hexane and 30% ethanol proved to be more economical than the others, since it extracted a greater quantity of oil with a lower initial biomass requirement. In summary, this microalgae-derived hydrocarbon production process is promising and offers insights for compound purification and future biofuel applications.

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Cultivation of microalgae using anaerobic digestate is a gain-win strategy for algal biomass production and achieving environmental benefits. However, the low biomass concentration and high harvest cost of the conventional suspended microalgae culture system are troublesome issues. In this study, a novel fluidized bed photobioreactor (FBPBR) based on diatomite powder was constructed for cultivating Scenedesmus quadricauda and treating diluted anaerobic digestate. The optimized diatomite carrier dosage of 750 mg/L increased microalgal biomass concentration to 1.58 g/L compared to suspended microalgae without carrier (0.99 g/L). When the light intensity was increased from 100 to 200 µmol/m2/s, the microalgal biomass in the FBPBR increased to 1.84 g/L and the settling efficiency increased to 93.58 %. This was due to the 1.60-fold enhancement of extracellular polymeric substance (EPS) secretion and changes in EPS properties. The increase in hydrophobic functional groups of EPS under high light intensity, coupled with the reconstitution of protein secondary structure, facilitated the initial attachment of algae to diatomite and the thickening of microalgal biofilm. Moreover, transcriptomic analysis demonstrated that diatomite promoted antioxidant defense and photosynthesis in S. quadricauda cells, alleviating the adverse effect of anaerobic digestate stress. The diatomite addition and elevated light intensity contributed to the highest lipid content (60.37 %), which was owing to the upregulated genes encoding fatty acid and triacylglycerol synthesis under the stress of localized nutrient starvation in the inner layer of microalgae biofilms. Furthermore, the regulation of phosphorus metabolism and NH4+-N assimilation improved nutrient removal (93.24 % and 96.86 % for NH4+-N and TP removal). This work will provide guidance for the development of FBPBR based on diatomite powder.

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This research evaluated a microalgae consortium (MC) in a pilot-scale tubular photobioreactor for municipal wastewater (MWW) treatment, compared with an aeration column photobioreactor. Transitioning from suspended MC to a microalgae-microbial biofilm (MMBF) maintained treatment performance despite increasing influent from 50 L to 150 L in a 260 L system. Carbon and nitrogen removal were effective, but phosphorus removal varied due to biofilm shading and the absence of phosphorus-accumulating organisms. High influent flow caused MMBF detachment due to shear stress. Stabilizing and re-establishing the MMBF showed that a stable phycosphere influenced microbial diversity and interactions, potentially destabilizing the MMBF. Heterotrophic nitrification-aerobic denitrification bacteria were crucial for MC equilibrium. Elevated gene expression related to nitrogen fixation, organic nitrogen metabolism, and nitrate reduction confirmed strong microalgal symbiosis, highlighting MMBF's treatment potential. This study supports the practical application of microalgae in wastewater treatment.

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In this work, we demonstrate the use of capillary electrophoresis and inductively coupled plasma mass spectrometry, as competitive methods primarily for ion chromatography, to determine changes in the concentration of small inorganic ions in the Nostoc sp. culture medium. Although macronutrients were analyzed by capillary electrophoresis with conductivity detection, micronutrients were analyzed by inductively coupled plasma mass spectrometry. The different light settings (light intensity and spectral composition) had a visible effect on the culture growth and depletion of calcium, magnesium, and phosphate ions, and iron and manganese elements when comparing the behavior under red or violet light with that under blue light.

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Cyanobacteria are oxygen-evolving prokaryotes that can be engineered for biofuel production from solar energy, CO2, and water. Isobutanol (IB) has the potential to serve as an alternative fuel and important chemical feedstock. The research involves engineering Synechocystis sp. PCC 6803, for photosynthetic isobutanol production via the 2-keto-acid pathway and their cultivation in lab-scale photobioreactors. This synthetic pathway involves the heterologous expression of two enzymes, α-ketoisovalerate decarboxylase (Kivd) and alcohol dehydrogenase (Yqhd), under a strong light-inducible promotor, psbA2, known to show increased gene expression under high light. The use of psbA2 could be a valuable strategy for isobutanol production as economic scaling up demands the utilization of natural sunlight, which also provides very high light intensity at midday, facilitating increased production. The study reports isobutanol production from engineered strains containing both pathway genes and with only kivd. In shake flask studies, the highest isobutanol titre of 75 mg L-1 (12th day) was achieved from an engineered strain DM12 under optimized light intensity. DM12 was cultivated in a 2 L flat panel photobioreactor, resulting in a maximum isobutanol titre of 371.8 mg L-1 (10th day) with 2 % CO2 and 200 µmol photons m-2 s-1. Cultivation of DM12 in a photobioreactor under mimic diurnal sunlight demonstrated the highest productivity of 39 mg L-1 day-1 with the maximum titre of 308.5 mg L-1 (9th day). This work lays the foundation for sustainable, large-scale biobutanol production using solar energy.

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Fossil fuel limitations and their influence on climate change through atmospheric greenhouse gas emissions have made the excessive use of fossil fuels widely recognized as unsustainable. The high lipid content, carbon-neutral nature and potential as a biofuel source have made microalgae a subject of global study. Microalgae are a promising supply of biomass for third-generation biofuels production since they are renewable. They have the potential to produce significant amounts of biofuel and are considered a sustainable alternative to non-renewable energy sources. Microalgae are currently incapable to synthesize algal biofuel on an extensive basis in a sustainable manner, despite their significance in the global production of biofuels. Wastewater contains nutrients (both organic and inorganic) which is essential for the development of microalgae. Microalgae and wastewater can be combined to remediate waste effectively. Wastewater of various kinds such as industrial, agricultural, domestic, and municipal can be used as a substrate for microalgal growth. This process helps reduce carbon dioxide emissions and makes the production of biofuels more cost-effective. This critical review provides a detailed analysis of the utilization of wastewater as a growth medium for microalgal - biofuel production. The review also highlights potential future strategies to improve the commercial production of biofuels from microalgae.

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Microalgae have the potential to exceed current nutrient recovery limits from wastewater, enabling water resource recovery facilities (WRRFs) to achieve increasingly stringent effluent permits. The use of photobioreactors (PBRs) and the separation of hydraulic retention and solids residence time (HRT/SRT) further enables increased biomass in a reduced physical footprint while allowing operational parameters (e.g., SRT) to select for desired functional communities. However, as algal technology transitions to full-scale, there is a need to understand the effect of operational and environmental parameters on complex microbial dynamics among mixotrophic microalgae, bacterial groups, and pests (i.e., grazers and pathogens) and to implement robust process controls for stable long-term performance. Here, we examine a full-scale, intensive WRRF utilizing mixed microalgae for tertiary treatment in the US (EcoRecover, Clearas Water Recovery Inc.) during a nine-month monitoring campaign. We investigated the temporal variations in microbial community structure (18S and 16S rRNA genes), which revealed that stable system performance of the EcoRecover system was marked by a low-diversity microalgal community (DINVSIMPSON = 2.01) dominated by Scenedesmus sp. (MRA = 55 %-80 %) that achieved strict nutrient removal (effluent TP < 0.04 mg·L-1) and steady biomass concentration (TSSmonthly avg. = 400-700 mg·L-1). Operational variables including pH, alkalinity, and influent ammonium (NH4+), correlated positively (p < 0.05, method = Spearman) with algal community during stable performance. Further, the use of these parameters as operational controls along with N/P loading and SRT allowed for system recovery following upset events. Importantly, the presence or absence of bacterial nitrification did not directly impact algal system performance and overall nutrient recovery, but partial nitrification (potentially resulting from NO2- accumulation) inhibited algal growth and should be considered during long-term operation. The microalgal communities were also adversely affected by zooplankton grazers (ciliates, rotifers) and fungal parasites (Aphelidium), particularly during periods of upset when algal cultures were experiencing culture turnover or stress conditions (e.g., nitrogen limitation, elevated temperature). Overall, the active management of system operation in order to maintain healthy algal cultures and high biomass productivity can result in significant periods (>4 months) of stable system performance that achieve robust nutrient recovery, even in winter months in northern latitudes (WI, USA).

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Membrane bioreactors (MBRs) hold significant promise for wastewater treatment, yet the persistent challenge of membrane fouling impedes their practical application. One promising solution lies in the synergy between microalgae and bacteria, offering efficient nutrient removal, reduced energy consumption, and potential mitigation of extracellular polymeric substances (EPS) concentrations. Inoculating microalgae presents a promising avenue to address membrane fouling in MBRs. This review marks the first exploration of utilizing microalgae for membrane fouling control in MBR systems. The review begins with a comprehensive overview of the evolution and distinctive traits of microalgae-MBRs. It goes further insight into the performance and underlying mechanisms facilitating the reduction of membrane fouling through microalgae intervention. Moreover, the review not only identifies the challenges inherent in employing microalgae for membrane fouling control in MBRs but also illuminates prospective pathways for future advancement in this burgeoning field.

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Produced water (PW) from oil and gas exploration adversely affects aquatic life and living organisms, necessitating treatment before discharge to meet effluent permissible limits. This study first used activated sludge to pretreat PW in a sequential batch reactor (SBR). The pretreated PW then entered a 13 L photobioreactor (PBR) containing Scenedesmus obliquus microalgae culture. Initially, 10% of the PW mixed with 90% microalgae culture in the PBR. After the exponential growth of the microalgae, an additional 25% of PW was added to the PBR without extra nutrients. This study reported the growth performance of microalgae in the PBR as well as the reduction in effluent's total organic carbon (TOC), total dissolved solids (TDS), electrical conductivity (EC), and heavy metals content. The results demonstrated removal efficiencies of 64% for TOC, 49.8% for TDS, and 49.1% for EC. The results also showed reductions in barium, iron, and manganese in the effluent by 95, 76, and 52%, respectively.

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Microalgae are a promising renewable feedstock that can be produced on non-arable land using seawater. Their biomass contains proteins, lipids, carbohydrates, and pigments, and can be used for various biobased products, such as food, feed, biochemicals, and biofuels. For such applications, the production costs need to be reduced, for example, by improving biomass productivity in photobioreactors. In this study, Picochlorum sp. (BPE23) was cultivated in a prototype of a novel outdoor V-shaped photobioreactor on Bonaire (12°N, 68°W). The novel photobioreactor design was previously proposed for the capture and dilution of sunlight at low-latitude locations. During several months, the biomass productivity of the local thermotolerant microalgae was determined at different dilution rates in continuous dilution and batch dilution experiments, without any form of temperature control. Reactor temperatures increased to 35°C-45°C at midday. In the continuous dilution experiments, high average biomass productivities of 28-31 g m-2 d-1 and photosynthetic efficiencies of 3.5%-4.3% were achieved. In the batch dilution experiments, biomass productivities were lower (17-23 g m-2 d-1), as microalgal cells likely experienced sudden light and temperature stress after daily reactor dilution. Nonetheless, dense cultures were characterized by high maximum photosynthetic rates, illustrating the potential of Picochlorum sp. for fast growth under outdoor conditions.

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Microalgae-mediated industrial flue gas biofixation has been widely discussed as a clean alternative for greenhouse gas mitigation. Through photosynthetic processes, microalgae can fix carbon dioxide (CO2) and other compounds and can also be exploited to obtain high value-added products in a circular economy. One of the major limitations of this bioprocess is the high concentrations of CO2, sulfur oxides (SOx), and nitrogen oxides (NOx) in flue gases, according to the origin of the fuel, that can inhibit photosynthesis and reduce the process efficiency. To overcome these limitations, researchers have recently developed new technologies and enhanced process configurations, thereby increased productivity and CO2 removal rates. Overall, CO2 biofixation rates from flue gases by microalgae ranged from 72 mg L-1 d -1 to over 435 mg L-1 d-1, which were directly influenced by different factors, mainly the microalgae species and photobioreactor. Additionally, mixotrophic culture have shown potential in improving microalgae productivity. Progress in developing new reactor configurations, with pilot-scale implementations was observed, resulting in an increase in patents related to the subject and in the implementation of companies using combustion gases in microalgae culture. Advancements in microalgae-based green technologies for environmental impact mitigation have led to more efficient biotechnological processes and opened large-scale possibilities.

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Prior research has emphasized the potential of microalgae in biodiesel production, driven by their ability to replace fossil fuels. However, the significant costs associated with microalgae cultivation present a major obstacle to scaling up production. This study aims to develop an eco-friendly microalgae cultivation system by integrating carbon dioxide from flue gas emissions with an affordable photobioreactor, providing a sustainable biomass production. The research evaluates the growth performance of Chlorella sorokiniana and Chlorella vulgaris across this integrated system for biomass and lipid production. Results indicate substantial biomass yields of 1.97 and 1.84 g/L, with lipid contents of 35 % and 41 % for C. sorokiniana and C. vulgaris, respectively. The macrobubble photobioreactor demonstrates high potential for microalgae biomass and lipid production, yielding quality fatty acid methyl esters such as palmitic, linoleic and stearic. This study presents an environmentally friendly system for efficient microalgae cultivation, generating lipid-rich biomass suitable for biodiesel production.

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Diatoms are a diverse group of phytoplankton usually dominating areas characterized by rapidly shifting light conditions. Because of their high growth rates and interesting biochemical profile, their biomass is considered for various commercial applications. This study aimed at identifying strains with superior growth in a photobioreactor (PBR) by screening the natural intraspecific diversity of ecotypes isolated from different habitats. We investigated the effect of PBR light fluctuating on a millisecond scale (FL, simulating the light in a PBR) on 19 ecotypes of the diatom Skeletonema marinoi isolated from the North Sea-Baltic Sea area. We compare growth, pigment ratios, phylogeny, photo-physiological variables and photoacclimation strategies between all strains and perform qPCR and absorption spectra analysis on a subset of strains. Our results show that the ecotypes responded differently to FL, and have contrasting photo-physiological and photoprotective strategies. The strains from Kattegat performed better in FL, and shared common photoacclimation and photoprotection strategies that are the results of adaptation to the specific light climate of the Kattegat area. The strains that performed better with FL conditions had a high light (HL)-acclimated phenotype coupled with unique nonphotochemical quenching features. Based on their characteristics, three strains were identified as good candidates for growth in PBRs.

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Laboratory scale conventional single-use bioreactor was used to investigate the effect of different stirrer speeds on the Arthrospira platensis (Spirulina platensis) culture. Experiments were handled in two steps. First step was the selection of the stirring speeds, which was simulated via using CFD, and the second was the long term cultivation with the selected speed. During 10 days of batches as the first step, under identical culture conditions, stirrer speed of 230 rpm gave higher results, compared to 130 and 70 rpm, with respect to dry biomass weight, absorbance value (AB) and chlorophyll-a concentration. Volumetric productivity during the growth phase of the cultures were calculated as 0.39 ± 0.03, 0.28 ± 0.01, and 0.19 ± 0.02 g L-1 d-1, from the fast to the slower speeds. According to the results a 17 day batch was handled with 230 rpm in order to monitor the effects on the culture. The culture reached a volumetric productivity of 0.33 ± 0.04 g L-1 d-1. Statistical analysis showed the significance of the parameters related with the stirring speed.

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The development of innovative production processes and the optimization of photobioreactors play an important role in generating industrial competitive production technologies for phototrophic biofilms. With emerse photobioreactors a technology was introduced that allowed efficient surface attached cultivation of terrestrial cyanobacteria. However, the productivity of emerse photobioreactors depends on the available cultivation surface. By the implementation of biocarriers to the bioreactor volume, the cultivation surface can be increased which potentially improves productivity and thus the production of valuable compounds. To investigate the surface attached cultivation on biocarriers new photobioreactors need to be developed. Additive manufacturing (AM) offers new opportunities for the design of photobioreactors but producing the needed transparent parts can be challenging using AM techniques. In this study an emerse fixed bed photobioreactor was designed for the use of biocarriers and manufactured using different AM processes. To validate the suitability of the photobioreactor for phototrophic cultivation, the optical properties of three-dimensional (3D)-printed transparent parts and postprocessing techniques to improve luminous transmittance of the components were investigated. We found that stereolithography 3D printing can produce parts with a high luminous transmittance of over 85% and that optimal postprocessing by sanding and clear coating improved the clarity and transmittance to more than 90%. Using the design freedom of AM resulted in a bioreactor with reduced part count and improved handling. In summary, we found that modern 3D-printing technologies and materials are suitable for the manufacturing of functional photobioreactor prototypes.