Semi-continuous Culture Research Articles

Semi-continuous cultivation of microalgae to treat coal mine effluent in pilot scale: Nutrient removal, biodesalination and fatty acid composition analysis

Coal excavation is associated with discharge of an enormous amount of mine water rich in salt and trace metal ions, which require efficient treatment before use. Considering the potentiality of microalgae for high salt tolerance, salt removal and metallic pollutant removal from aqueous system, present study focuses semi-continuous cultivation of candidate microalgae Chlorella pyrenoidosa (NCIM 2738) in bubble column reactor (BCR) and open raceway pond (ORP) for nutrient supplemented coal mine effluent (NSCME) treatment. In between different hydraulic retention times (HRTs), HRT 6 d provide maximum average biomass productivity of 950 mg L−1 d−1 in BCR and 728.4 mg L−1 d−1 in ORP. HRT 9 d facilitates a maximum lipid content of 1.8 g L ̶ 1 in BCR and 1.4 g L ̶ 1 in ORP. Further, HRT 9 d had the maximum COD removal efficiency (96.5 % in BCR and 94.2 % in ORP) and maximum salinity removal efficiency (93 % in BCR and 92 % in ORP). Fatty acid methyl esters (FAME) characterization highlighted potential biodiesel applicability with cetane number (CN) of 53.94. Energy dispersive X-ray spectroscopy (EDS) revealed an excess amount of salt and metal ions deposition on the surface of harvested microalgae samples. Outdoor large-scale C. pyrenoidosa cultivation can be integrated with coal mine closure plans to meet sustainable development goals (SDGs) 6, 7 and 13.

A self-isolated acid-tolerant Parachlorella kessleri with high efficiency in treating rare earth mining sewage

Rare earth mining sewage is a significant environmental concern due to its high acidity and ammonia nitrogen levels. Finding a sustainable and cost-effective treatment method is essential. Parachlorella kessleri FM2, a green algae strain isolated in-house, has demonstrated remarkable abilities to grow and remove ammonia nitrogen (NH4+-N) from highly acidic rare earth wastewater without the need for alkaline additives. After optimizing conditions, P. kessleri FM2 achieved an impressive NH4+-N removal rate of 7.94 mg/L/d and a removal efficiency of 98.71% in a 1.5-L photobioreactor. In semi-continuous culture, the average NH4+-N removal rate remained high at 6.67 mg/L/d. When scaled up to continuous culture in a 5-L photobioreactor, P. kessleri FM2 maintained stability with an NH4+-N removal rate of 6.79 mg/L/d. Additionally, we conducted a preliminary analysis of P. kessleri FM2's acid resistance mechanism, further highlighting its potential as a candidate for treating acidic wastewater.

Assessing the potential of a genetically modified Parachlorella kessleri-I with low CO2 inducible proteins for enhanced biomass and biofuel productivity

The biological process is one of the promising approaches for CO2 capture and storage. Recently, biological sequestration using microalgae has gained much interest due to its capability to utilize CO2 as a carbon source to generate valorizable biomass. This algal biomass further can be used as a feedstock for bioenergy and different value-added products. Here, we have heterogeneously overexpressed the low CO2 inducible proteins (LCIA & LCIB) from Chlamydomonas reinhardtii in the marine alga Parachlorella kessleri-I. Incorporation of these two genes boosted the endogenous CCM leading to a hybrid system and improved overall carbon acquisition. The combination of these CCM proteins enhanced the supply of inorganic carbon to RuBisCO with positive effects on growth and biomass productivity. Biochemical analyses revealed that the hybrid CCM in P. kessleri-I was characterised by 2- fold higher carbonic anhydrase activity, increased starch, and 2X more lipid yield in comparison to wild type (WT) strain. Moreover, the semi-continuous cultivation of GM strain with supply of 1 % CO2 led to around 60 % rise in the overall biomass productivity. These traits also persisted in scale-up studies leading to 2X more biomass productivity in the GM strain than WT in 100 L PBR. A comparative life-cycle assessment underlines the sustainability of the process of carbon capture and both biofuel and biochar production from P. kessleri-I with a hybrid CCM.

Performance evaluation of Chlorella sp. BRE5 for augmented biomass and lipid production implementing semi-continuous cultivation strategy

Performance evaluation of Chlorella sp. BRE5 for augmented biomass and lipid production implementing semi-continuous cultivation strategy

Outdoor semi-continuous cultivation of Synechococcus sp. for enhanced carotenoid production

The carotenoid productivity of a euryhaline Synechococcus sp. was studied in indoor and outdoor conditions. Indoor experiments revealed that the strain could maintain growth and carotenoid accumulation within a salinity range of 2.5–8.0 % NaCl. In an indoor two phase semi-continuous cultivation in PBRs, every three days, 50 % of the culture was replaced with fresh growth media in phase one, whereas the removed culture was grown for another three days in the second phase to enhance the carotenoid yield. Biomass and carotenoid productivity of the strain in the second phase were similar in five consecutive cultivation cycles. Next, the same two-phase cultivation experiment was conducted in 5 sq. m outdoor raceway tanks. However, for the outdoor experiment, carotenoid-rich biomass was harvested from the second phase by membrane filtration, and the growth media was recycled for cultivating in the first phase. The salinity of the culture continued to increase as the evaporated water was balanced by seawater. The semi-continuous cultivation of the first phase continued for 20 days before the culture salinity reached 7.4 % NaCl. For Synechococcus sp., the growth and carotenoid productivities were not affected within this salinity range.

Establishment of a semi-continuous scale-down clone screening model for intensified perfusion culture.

Perfusion cultures have been extensively used in the biotechnology industry to achieve high yields of recombinant products, especially those with stability issue. The WuXiUP™ platform represents a novel intensified perfusion that can achieve ultra-high productivity. This study describes a representative scale-down 24-deep well plate (24-DWP) cell culture model for intensified perfusion clone screening. Clonal cell lines were expanded and evaluated in 24-DWP semi-continuous culture. Cell were sampled and counted daily with the aid of an automated liquid handler and high-throughput cell counter. To mimic perfusion culture, 24-DWP plates were spun down and resuspended with fresh medium daily. Top clones were ranked based on growth profiles and productivities. The best performing clones were evaluated on bioreactors. The selected clones achieved volumetric productivity (Pv) up to 5g/L/day when expressing a monoclonal antibody, with the accumulative harvest Pv exceeding 60g/L in a 21-day cell culture. Product quality attributes of clones cultured in 24-DWP were comparable with those from bioreactors. A high seeding strategy further shortened the clone screening timeline. In this study, a 24-DWP semi-continuous scale-down model was successfully developed to screen for cell lines suitable for intensified perfusion culture.

Effects of aeration and centrifugation conditions on omega-3 fatty acid production by the mixotrophic dinoflagellate Gymnodinium smaydae in a semi-continuous cultivation system on a pilot scale

High production and efficient harvesting of microalgae containing high omega-3 levels are critical concerns for industrial use. Aeration can elevate production of some microalgae by providing CO2 and O2. However, it may lower the production of others by generating shear stress, causing severe cell damage. The mixotrophic dinoflagellate Gymnodinium smaydae is a new, promising microalga for omega-3 fatty acid production owing to its high docosahexaenoic acid content, and determining optimal conditions and methods for high omega-3 fatty acid production and efficient harvest using G. smaydae is crucial for its commercial utilization. Therefore, to determine whether continuous aeration is required, we measured densities of G. smaydae and the dinoflagellate prey Heterocapsa rotundata in a 100-L semi-continuous cultivation system under no aeration and continuous aeration conditions daily for 9 days. Furthermore, to determine the optimal conditions for harvesting through centrifugation, different rotational speeds of the continuous centrifuge and different flow rates of the pump injecting G. smaydae + H. rotundata cells into the centrifuge were tested. Under continuous aeration, G. smaydaeproduction gradually decreased; however, without aeration, the production remained stable. Harvesting efficiency and the dry weights of omega-3 fatty acids of G. smaydae + H. rotundata cells at a rotational speed of 16,000 rpm were significantly higher than those at 2,000–8,000 rpm. However, these parameters did not significantly differ at injection pump flow rates of 1.0–4.0 L min-1. The results of the present study provide a basis for optimized production and harvest conditions for G. smaydae and other microalgae.

Airlift bioreactor–based strategies for prolonged semi-continuous cultivation of edible Agaricomycetes

Submerged cultivation of edible filamentous fungi (Agaricomycetes) in bioreactors enables maximum mass transfer of nutrients and has the potential to increase the volumetric productivity of fungal biomass compared to solid state cultivation. These aspects are paramount if one wants to increase the range of bioactives (e.g. glucans) in convenient time frames. In this study, Trametes versicolor (M9911) outperformed four other Agaricomycetes tested strains (during batch cultivations in an airlift bioreactor). This strain was therefore further tested in semi-continuous cultivation. Continuous and semi-continuous cultivations (driven by the dilution rate, D) are the preferred bioprocess strategies for biomass production. We examined the semi-continuous cultivation of T. versicolor at dilution rates between 0.02 and 0.1 h−1. A maximum volumetric productivity of 0.87 g/L/h was obtained with a D of 0.1 h−1 but with a lower total biomass production (cell dry weight, CDW 8.7 g/L) than the one obtained at lower dilution rates (12.3 g/L at D of 0.04 and vs 13.4 g/L, at a D of 0.02 h−1). However, growth at a D of 0.1 h−1 resulted in a very short fermentation (18 h) which terminated due to washout (the specific D exceeded the maximum growth rate of the fungal biomass). At a D of 0.04 h−1, a CDW of 12.3 g/L was achieved without compromising the total residence time (184 h) of the fermentation. While the D of 0.04 h−1 and 0.07 h−1 achieved comparable volumetric productivities (0.5 g/L/h), the total duration of the fermentation at D of 0.07 h−1 was only 85 h. The highest glucan content of cells (27.8 as percentage of CDW) was obtained at a D of 0.07 h−1, while the lowest glucan content was observed in T. versicolor cells grown at a D of 0.02 h−1.Key points• The highest reported volumetric productivity for fungal biomass was 0.87 g/L/h.• Semi-continuous fermentation at D of 0.02 h−1 resulted in 13.4 g/L of fungal biomass.• Semi-continuous fermentation at D of 0.07 h−1 resulted in fungal biomass with 28% of total glucans.

Influence of Anaerobically Digested Dairy Waste on Growth and Bio-Active Compounds of Spirulina subsalsa (Cyanobacteria) under Semi-Continuous Culture Conditions

Influence of Anaerobically Digested Dairy Waste on Growth and Bio-Active Compounds of <i>Spirulina subsalsa</i> (Cyanobacteria) under Semi-Continuous Culture Conditions

Low-cost cultivation of Nannochloropsis oceanica in newly designed photobioreactors and its productivity trends in semi-continuous cultivation under inland outdoor conditions

Most marine microalgae are typically cultivated in coastal areas due to challenges in inland cultivation. In this 185 days experiment, Nannochloropsis oceanica was semi-continuously cultivated inland using different photobioreactors (PBRs). The newly designed 700-liter (L) PBR exhibited tolerance to seasonal changes compared to the 150-L PBRs. The innovative in-situ oxygen release rate (ORR) measurement method results indicated that ORR was influenced by light intensity and temperature. The optimal temperature range for N. oceanica growth was 14–25 ℃, demonstrated cold tolerance and lipid accumulation at low temperatures. The maximum lipid content in 700-L and 150-L PBRs was 29 % and 28 %, respectively. Based on the average biomass productivity, the price of N. oceanica was $11.89 kg−1 (or $3.35 kg−1 based on maximum biomass productivity), which is cheaper than the current market price of $20.19 kg−1. From results, smaller PBRs at the same hydro electricity price are more cost-effective.

Semi-continuous cultivation of EPS-producing marine cyanobacteria: A green biotechnology to remove dissolved metals obtaining metal-organic materials

Given the necessity for bioprocesses scaling-up, the present study aims to explore the potential of three marine cyanobacteria and a consortium, cultivated in semi-continuous mode, as a green approach for i) continuous exopolysaccharide-rich biomass production and ii) removal of positively charged metals (Cu, Ni, Zn) from mono and multi-metallic solutions. To ensure the effectiveness of both cellular and released exopolysaccharides, weekly harvested whole cultures were confined in dialysis tubings. The results revealed that all the tested cyanobacteria have a stronger affinity towards Cu in mono and three-metal systems. Despite the amount of metals removed per gram of biomass decreased with higher biosorbent dosage, the more soluble carbohydrates were produced, the greater was the metal uptake, underscoring the pivotal role of released exopolysaccharides in metal biosorption. According to this, Dactylococcopsis salina 16Som2 showed the highest carbohydrate productivity (142 mg L−1 d−1) and metal uptake (84 mg Cu g−1 biomass) representing a promising candidate for further studies. The semi-continuous cultivation of marine cyanobacteria here reported assures a schedulable production of exopolysaccharide-rich biosorbents with high metal removal and recovery potential, even from multi-metallic solutions, as a step forward in the industrial application of cyanobacteria.

A novel multi-stage enrichment workflow and comprehensive characterization for HEK293F-derived extracellular vesicles.

Extracellular vesicles (EVs) are emerging as a promising drug delivery vehicle as they are biocompatible and capable of targeted delivery. However, clinical translation of EVs remains challenging due to the lack of standardized and scalable manufacturing protocols to consistently isolate small EVs (sEVs) with both high yield and high purity. The heterogenous nature of sEVs leading to unknown composition of biocargos causes further pushback due to safety concerns. In order to address these issues, we developed a robust quality-controlled multi-stage process to produce and isolate sEVs from human embryonic kidney HEK293F cells. We then compared different 2-step and 3-step workflows for eliminating protein impurities and cell-free nucleic acids to meet acceptable limits of regulatory authorities. Our results showed that sEV production was maximized when HEK293F cells were grown at high-density stationary phase in semi-continuous culture. The novel 3-step workflow combining tangential flow filtration, sucrose-cushion ultracentrifugation and bind-elute size-exclusion chromatography outperformed other methods in sEV purity while still preserved high yield and particle integrity. The purified HEK293F-derived sEVs were thoroughly characterized for identity including sub-population analysis, content profiling including proteomics and miRNA sequencing, and demonstrated excellent preclinical safety profile in both in-vitro and in-vivo testing. Our rigorous enrichment workflow and comprehensive characterization will help advance the development of EVs, particularly HEK293F-derived sEVs, to be safe and reliable drug carriers for therapeutic applications.

Semi-continuous cultivation for enhanced protein production using indigenous green microalgae and synthetic municipal wastewater

Cultivation of microalgae has gained significant interest as an alternative protein source, potentially becoming a target commodity recovered from microalgae-based wastewater treatment. This study examined a semi-continuous cultivation strategy to optimize protein accumulation of the indigenous freshwater chlorophytes, Lobochlamys segnis and Klebsormidium flaccidum, and simultaneously remove nutrients from wastewater efficiently. A strain-specific regime was made based on a fixed biomass concentration at the start of 24-h cultivation cycle, i.e., a constant initial cell density, which regulated harvesting and fresh medium supply volume according to the dilution rate. Six cultivation cycles were conducted in lab-scale 1L reactors with a synthetic municipal wastewater. Lobochlamys segnis and K. flaccidum grew exponentially in all cycles. The biomass productivity was 573 and 580 mg L–1 day–1, in which the total protein consisted of 62 and 45% of dry cell weight (dw), respectively. When a culture medium deficient in nitrogen and phosphorus was used, protein level was significantly reduced. L. segnis consumed all NH4+ and PO43– supplied by the medium replacement, giving the removal rate of 9.2 and 5.2 mg L–1 day–1. Whereas K. flaccidum removed 13.8 mg L–1 day–1 NH4+ without completing PO43– removal. The amino acid profile of both strains was characterized by glutamic acids content (4–5% dw). We concluded that the designed cultivation regime would support a constant biomass production with stable and high protein content, along with an efficient removal of nutrient from the wastewater.

Comparison of plant biostimulating properties of Chlorella sorokiniana biomass produced in batch and semi-continuous systems supplemented with pig manure or acetate

Microalgae-derived biostimulants provide an eco-friendly biotechnology for improving crop productivity. The strategy of circular economy includes reducing biomass production costs of new and robust microalgae strains grown in nutrient-rich wastewater and mixotrophic culture where media is enriched with organic carbon. In this study, Chlorella sorokiniana was grown in 100 l bioreactors under sub-optimal conditions in a greenhouse. A combination of batch and semi-continuous cultivation was used to investigate the growth, plant hormone and biostimulating effect of biomass grown in diluted pig manure and in nutrient medium supplemented with Na-acetate. C. sorokiniana tolerated the low light (sum of PAR 0.99 ± 0.18 mol/photons/(m2/day)) and temperature (3.7–23.7° C) conditions to maintain a positive growth rate and daily biomass productivity (up to 149 mg/l/day and 69 mg/l/day dry matter production in pig manure and Na-acetate supplemented cultures respectively). The protein and lipid content was significantly higher in the biomass generated in batch culture and dilute pig manure (1.4x higher protein and 2x higher lipid) compared to the Na-acetate enriched culture. Auxins indole-3-acetic acid (IAA) and 2-oxindole-3-acetic acid (oxIAA) and salicylic acid (SA) were present in the biomass with significantly higher auxin content in the biomass generated using pig manure (> 350 pmol/g DW IAA and > 84 pmol/g DW oxIAA) compared to cultures enriched with Na-acetate and batch cultures (< 200 pmol/g DW IAA and < 27 pmol/g DW oxIAA). No abscisic acid and jasmonates were detected. All samples had plant biostimulating activity measured in the mungbean rooting bioassay with the Na-acetate supplemented biomass eliciting higher rooting activity (equivalent to 1–2 mg/l IBA) compared to the pig manure (equivalent to 0.5–1 mg/l IBA) and batch culture (equivalent to water control) generated biomass. Thus C. sorokiniana MACC-728 is a robust new strain for biotechnology, tolerating low light and temperature conditions. The strain can adapt to alternative nutrient (pig manure) and carbon (acetate) sources with the generated biomass having a high auxin concentration and plant biostimulating activity detected with the mungbean rooting bioassay.

An integrated process for enhanced production and purification of fucoxanthin and sulfated polysaccharides in diatom Hyalosynedra toxoneides cultures

BackgroundThis study explores the efficient production and purification of health-promoting compounds, fucoxanthin and sulfated polysaccharides, from the diatom Hyalosynedra toxoneides, contributing to the growing demand for sustainable bioactive compounds. MethodsSemi-continuous cultivation of H. toxoneides (AQ9) in three medium concentrations (2.5f, 5f, 10f) was conducted in a custom-made photobioreactor. Fucoxanthin was extracted using 95 % ethanol, purified through silica gel chromatography, and characterized via high-performance liquid chromatography. Crude polysaccharides were obtained, purified using DEAE-Sephacel resins, and characterized for uronic acid and sulfate content. Significant findingsHigher culture medium concentrations led to increased biomass, fucoxanthin, and polysaccharide yields, providing insights for large-scale cultivation. With a 10f medium in semi-continuous cultures, we achieved the highest yields of approximately 1.18 g/L for cell mass, 4.45 mg/L for fucoxanthin, and 17.20 mg/L for crude polysaccharides, with corresponding productivities of 0.20 g/L/day, 0.74 mg/L/day, and 2.46 mg/L/day. Fucoxanthin was purified from crude pigments with 91 % purity and 70 % recovery. Meanwhile, two uncharacterized sulfated polysaccharides were isolated from crude polysaccharides using anion-exchange chromatography. Notably, the culture media influenced sulfated polysaccharide distribution. This integrated approach contributes to efficient production and purification of bioactive compounds from diatom cultures, with potential applications in diverse industries, including nutraceuticals and cosmetics.

High-Level Production of scFv-Fc Antibody Using an Artificial Promoter System with Transcriptional Positive Feedback Loop of Transactivator in CHO Cells.

With the increasing demand for therapeutic antibodies, CHO cells have become the de facto standard as producer host cells for biopharmaceutical production. High production yields are required for antibody production, and developing a high-titer production system is increasingly crucial. This study was established to develop a high-production system using a synthetic biology approach by designing a gene expression system based on an artificial transcription factor that can strongly induce the high expression of target genes in CHO cells. To demonstrate the functionality of this artificial gene expression system and its ability to induce the high expression of target genes in CHO cells, a model antibody (scFv-Fc) was produced using this system. Excellent results were obtained with the plate scale, and when attempting continuous production in semi-continuous cultures using bioreactor tubes with high-cell-density suspension culture using a serum-free medium, high-titer antibody production at the gram-per-liter level was achieved. Shifting the culture temperature to a low temperature of 33 °C achieved scFv-Fc concentrations of up to 5.5 g/L with a specific production rate of 262 pg/(cell∙day). This artificial gene expression system should be a powerful tool for CHO cell engineering aimed at constructing high-yield production systems.

Comparison of microalgae cultivation modes for advanced wastewater purification and pollutant-resource conversion: A pilot study

Microalgae absorb CO2 and nutrients (e.g., nitrogen, phosphorus) for biomass growth, which has high potential for wastewater purification and pollutant-resource conversion. However, current studies mainly focused on short-term effect on stimulated water in lab. In this study, the feasibility of pilot-scale microalgae cultivation in real wastewater with low pollutant content and no sterilization was evaluated, and the optimal culture mode was proposed. It was found that microalgae could remove >95 % of nitrogen and phosphorus during the batch cultivation, proving the strong purifying ability of wastewater with low level of nitrogen and phosphorus. However, the high ratio of nitrogen to phosphorus led to long-term phosphorus shortage in the semi-continuous cultivation mode. The fluctuation of wastewater quality/quantity broke the balance of suspended microalgae growth and wash-out in the continuous cultivation mode. Besides, the aggregation of microalgae at the bottom inhibited the sustainable wastewater purification in both semi-continuous and continuous cultivation modes. Finally, continuous cultivation mode of attached microalgae (CCMA) was proposed with stable and efficient performance of wastewater treatment, and the process of microalgae-water separation could be omitted. Under this mode, the average removal of COD, TN, and TP reached 39.33 %, 21.27 % and 88.10 %, respectively, which could be further enhanced by optimizing the carrier density. Therefore, CCMA was a promising approach for scale-up low-pollutant wastewater treatment.

Increased paclitaxel recovery from Taxus baccata vascular stem cells using novel in situ product recovery approaches

In this study, several approaches were tested to optimise the production and recovery of the widely used anticancer drug Taxol® (paclitaxel) from culturable vascular stem cells (VSCs) of Taxus baccata, which is currently used as a successful cell line for paclitaxel production. An in situ product recovery (ISPR) technique was employed, which involved combining three commercial macro-porous resin beads (HP-20, XAD7HP and HP-2MG) with batch and semi-continuous cultivations of the T. baccata VSCs after adding methyl jasmonate (Me-JA) as an elicitor. The optimal resin combination resulted in 234 ± 23 mg of paclitaxel per kg of fresh-weight cells, indicating a 13-fold improved yield compared to the control (with no resins) in batch cultivation. This resin treatment was further studied to evaluate the resins’ removal capacity of reactive oxygen species (ROS), which can cause poor cell growth or reduce product synthesis. It was observed that the ISPR cultivations had fourfold less intracellular ROS concentration than that of the control; thus, a reduced ROS concentration established by the resin contributed to increased paclitaxel yield, contrary to previous studies. These paclitaxel yields are the highest reported to date using VSCs, and this scalable production method could be applied for a diverse range of similar compounds utilising plant cell culture.Graphical

The synergetic effect from the combination of different adsorption resins in batch and semi-continuous cultivations of S. Cerevisiae cell factories to produce acetylated Taxanes precursors of the anticancer drug Taxol.

In situ product recovery is an efficient way to intensify bioprocesses as it can perform adsorption of the desired natural products in the cultivation. However, it is common to use only one adsorbent (liquid or solid) to perform the product recovery. For this study, the use of an in situ product recovery method with three combined commercial resins (HP-20, XAD7HP, and HP-2MG) with different chemical properties was performed. A new yeast strain of Saccharomyces cerevisiae was engineered using CRISPR Cas9 (strain EJ2) to deliver heterologous expression of oxygenated acetylated taxanes that are precursors of the anticancer drug Taxol ® (paclitaxel). Microscale cultivations using a definitive screening design (DSD) were set to get the best resin combinations and concentrations to retrieve high taxane titers. Once the best resin treatment was selected by the DSD, semi-continuous cultivation in high throughput microscale was performed to increase the total taxanes yield up to 783 ± 33 mg/L. The best T5α-yl Acetate yield obtained was up to 95 ± 4 mg/L, the highest titer of this compound ever reported by a heterologous expression. It was also observed that by using a combination of the resins in the cultivation, 8 additional uncharacterized taxanes were found in the gas chromatograms compared to the dodecane overlay method. Lastly, the cell-waste reactive oxygen species concentrations from the yeast were 1.5-fold lower in the resin's treatment compared to the control with no adsorbent aid. The possible future implications of this method could be critical for bioprocess intensification, allowing the transition to a semi-continuous flow bioprocess. Further, this new methodology broadens the use of different organisms for natural product synthesis/discovery benefiting from clear bioprocess intensification advantages.

Application of Immersed Membrane Bioreactor for Semi-Continuous Production of Polyhydroxyalkanoates from Organic Waste-Based Volatile Fatty Acids.

Volatile fatty acids (VFAs) appear to be an economical carbon feedstock for the cost-effective production of polyhydroxyalkanoates (PHAs). The use of VFAs, however, could impose a drawback of substrate inhibition at high concentrations, resulting in low microbial PHA productivity in batch cultivations. In this regard, retaining high cell density using immersed membrane bioreactor (iMBR) in a (semi-) continuous process could enhance production yields. In this study, an iMBR with a flat-sheet membrane was applied for semi-continuous cultivation and recovery of Cupriavidus necator in a bench-scale bioreactor using VFAs as the sole carbon source. The cultivation was prolonged up to 128 h under an interval feed of 5 g/L VFAs at a dilution rate of 0.15 (d-1), yielding a maximum biomass and PHA production of 6.6 and 2.8 g/L, respectively. Potato liquor and apple pomace-based VFAs with a total concentration of 8.8 g/L were also successfully used in the iMBR, rendering the highest PHA content of 1.3 g/L after 128 h of cultivation. The PHAs obtained from both synthetic and real VFA effluents were affirmed to be poly(3-hydroxybutyrate-co-3-hydroxyvalerate) with a crystallinity degree of 23.8 and 9.6%, respectively. The application of iMBR could open an opportunity for semi-continuous production of PHA, increasing the feasibility of upscaling PHA production using waste-based VFAs.