Development, characterization and application of 3D printed adsorbents for in situ recovery of taxadiene from microbial cultivations.

Microbial cell factories are an attractive alternative to produce high-value natural products using sustainable processes. However, product recovery is one of the main challenges to reduce production cost and make these technologies economically interesting. In this work, new resins were formulated to 3D print hydrophobic adsorbents for the recovery of biologics from microbial cultivations. Benzyl methacrylate (BEMA) and butyl methacrylate (BUMA) were selected as functional monomers suitable for the adsorption of hydrophobic compounds. Pore morphology was tailored through the inclusion of pore forming agents (porogens) in the resin. Different porogens and porogen concentrations were evaluated resulting in materials with different porous networks. Sudan 1 and the anticancer drug paclitaxel were employed as model compounds to test the adsorption performance of hydrophobic and terpene molecules onto the developed 3D printed materials. The material with greatest adsorption capacity was obtained using BEMA monomer with 40 % (v/v) porogen (BEMA40). The performance of BEMA40 to recover taxadiene from small-scale (5 mL) Saccharomyces cerevisiae cultivations was tested and compared with commercial Diaion HP-20 beads. Taxadiene titres on BEMA40 (46 ± 2 mg/L) and Diaion HP-20 (54 ± 4 mg/L) were comparable, with no taxadiene detected in the cells and cell-free media, suggesting near 100 % taxadiene partition on the adsorbents. Compared to commercial beads, 3D printed adsorbents can be customized with adjustments in the resin formulation, are well adaptable to diverse bioreactor types, do not clog sampling ports and columns and are easier to handle during post processing. The results of this work demonstrate the potential of 3D printing to fabricate hydrophobic interaction adsorbent materials and their application in the recovery of biological products.

In situ recovery of taxadiene using solid adsorption in cultivations with Saccharomyces cerevisiae.

In this study, an engineered strain of Saccharomyces cerevisiae was used to produce taxadiene, a precursor in the biosynthetic pathway of the anticancer drug paclitaxel. Taxadiene was recovered in situ with the polymeric adsorbent Diaion © HP-20. Here we tested two bioreactor configurations and adsorbent concentrations to maximize the production and recovery of taxadiene. An external recovery configuration (ERC) was performed with the integration of an expanded bed adsorption column, whereas the internal recovery configuration (IRC) consisted in dispersed beads inside the bioreactor vessel. Taxadiene titers recovered in IRC were higher to ERC by 3.4 and 3.5 fold by using 3% and 12% (w/v) adsorbent concentration respectively. On the other hand, cell growth kinetics were faster in ERC which represents an advantage in productivity (mg of taxadiene/L*h). High resin bead concentration (12% w/v) improved the partition of taxadiene onto the beads up to 98%. This result represents an advantage over previous studies using a 3% resin concentration where the partition of taxadiene on the beads was around 50%. This work highlights the potential of in situ product recovery to improve product partition, reduce processing steps and promote cell growth. Nevertheless, a careful design of bioreactor configuration and process conditions is critical.

Enhanced production of glutaric acid by biocatalyst-recycled bioconversion process integrated with in situ product recovery by adsorption.

Product inhibition caused by organic acids is a serious issue in establishing economical biochemical production systems. Herein, for enhanced production of glutaric acid by overcoming product inhibition triggered by glutaric acid, a whole-cell bioconversion system equipped with biocatalyst recycling process and in situ product recovery by adsorption was developed successfully. From the whole-cell bioconversion reaction, we found that both dissociated and undissociated forms of glutaric acid acted as an inhibitor in the whole-cell bioconversion reaction, wherein bioconversion was hindered beyond 200 mM glutaric acid regardless of reaction pH. Therefore, as the promising solution for the inhibition issue by glutaric acid, the biocatalyst-recycled bioconversion process integrated with in situ product recovery by adsorption was introduced in the whole-cell bioconversion. As a result, 592 mM glutaric acid was produced from 1000 mM 5-aminovaleric acid with 59.2% conversion. We believe that our system will be a promising candidate for economically producing organic acids with high titer.

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.

Co-culture of Kluyveromyces marxianus and Meyerozyma guilliermondii with In Situ Product Recovery of 2-Phenylethanol.

Production of 2-phenylethanol (2-PE) via Kluyveromyces marxianus is well-established. However, co-culture with other microbes in combination with in situ product recovery (ISPR) yields improved selectivity and volumetric productivity. Fermentation ofK. marxianus (MUCL 53775) with direct inclusion of absorptive polymer Hytrel3548 achieved ISPR, but accumulation of the byproduct phenylethyl acetate (PEA) was strongly favored. Co-culture of K. marxianus (MUCL 53775) with Meyerozyma guilliermondii (MUCL 28072) with ISPR limited PEA production, thereby improving the 2-PE selectivity from 13 to 90%, compared to a pure culture of K. marxianus (MUCL 53775) under similar conditions. This improved the volumetric productivity by 85% compared to 2-PE ISPR with a pure culture of K. marxianus. This is the first report of co-culture in a two-phase fermentation for 2-PE bioproduction and demonstrates that interactions between co-culture and ISPR techniques can modulate bioproduction between 2-PE and byproduct PEA, and this technique will be explored for other strain combinations and for other high-value molecules of interest.

Development of In Situ Product Recovery (ISPR) System Using Amberlite IRA67 for Enhanced Biosynthesis of Hyaluronic Acid by Streptococcus zooepidemicus.

High broth viscosity due to the accumulation of hyaluronic acid (HA) causes a limited yield of HA. It is a major problem of HA production using Streptococcus zooepidemicus. Extractive fermentation via in situ product recovery (ISPR) was utilized to enhance the HA production. Resins from Amberlite: IRA400 Cl; IRA900 Cl; IRA410 Cl; IRA402 Cl; and IRA67 were tested for the HA adsorption. IRA67 showed high adsorption capacity on HA. The study of the adsorption via a 2 L stirred tank bioreactor of S. zooepidemicus fermentation was investigated to elucidate the adsorption of HA onto IRA67 in dispersed and integrated internal column systems. The application of a dispersed IRA67 improved the HA production compared to the fermentation without resin addition by 1.37-fold. The HA production was further improved by 1.36-fold with an internal column (3.928 g/L) over that obtained with dispersed IRA67. The cultivation with an internal column shows the highest reduction of viscosity value after the addition of IRA67 resin: from 58.8 to 23.7 (mPa·s), suggesting the most effective ISPR of HA. The improved biosynthesis of HA indicated that an extractive fermentation by ISPR adsorption is effective and may streamline the HA purification.

Systematic metabolic engineering of Escherichia coli for the enhanced production of cinnamaldehyde.

Cinnamaldehyde (CAD) derived from cinnamon bark has received much attention for its potential as a nematicide and food additive. Previously, we have succeeded in developing an Escherichia coli strain (YHP05) capable of synthesizing cinnamaldehyde; however, the production titer (75 mg/L) was not sufficient for commercialization. Herein, to develop an economical and sustainable production bioprocess, we further engineered the YHP05 strain for non-auxotrophic, antibiotic-free, inducer-free hyperproduction of CAD using systematic metabolic engineering. First, the conversion of trans-cinnamic acid (t-CA) to CAD was improved by the co-expression of carboxylic acid reductase and phosphopantetheinyl transferase (PPTase) genes. Second, to prevent the spontaneous conversion of CAD to cinnamyl alcohol, 10 endogenous reductase and dehydrogenase genes were deleted. Third, all expression cassettes were integrated into the chromosomal DNA using an auto-inducible system for antibiotic- and inducer-free production. Subsequently, to facilitate CAD production, available pools of cofactors (NADPH, CoA, and ATP) were increased, and acetate pathways were deleted. With the final antibiotic-, plasmid-, and inducer-free strain (H-11MPmR), fed-batch cultivations combined with in situ product recovery (ISPR) were performed, and the production titer of CAD as high as 3.8 g/L could be achieved with 49.1 mg/L/h productivity, which is the highest CAD titer ever reported.

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.

An updated review on advancement in fermentative production strategies for biobutanol using Clostridium spp.

A significant concern of our fuel-dependent era is the unceasing exhaustion of petroleum fuel supplies. In parallel to this, environmental issues such as the greenhouse effect, change in global climate, and increasing global temperature must be addressed on a priority basis. Biobutanol, which has fuel characteristics comparable to gasoline, has attracted global attention as a viable green fuel alternative among the many biofuel alternatives. Renewable biomass could be used for the sustainable production of biobutanol by the acetone-butanol-ethanol (ABE) pathway. Non-extinguishable resources, such as algal and lignocellulosic biomass, and starch are some of the most commonly used feedstock for fermentative production of biobutanol, and each has its particular set of advantages. Clostridium, a gram-positive endospore-forming bacterium that can produce a range of compounds, along with n-butanol is traditionally known for its biobutanol production capabilities. Clostridium fermentation produces biobased n-butanol through ABE fermentation. However, low butanol titer, a lack of suitable feedstock, and product inhibition are the primary difficulties in biobutanol synthesis. Critical issues that are essential for sustainable production of biobutanol include (i) developing high butanol titer producing strains utilizing genetic and metabolic engineering approaches, (ii) renewable biomass that could be used for biobutanol production at a larger scale, and (iii) addressing the limits of traditional batch fermentation by integrated bioprocessing technologies with effective product recovery procedures that have increased the efficiency of biobutanol synthesis. Our paper reviews the current progress in all three aspects of butanol production and presents recent data on current practices in fermentative biobutanol production technology.

Production of indole by Corynebacterium glutamicum microbial cell factories for flavor and fragrance applications.

BACKGROUND: The nitrogen containing aromatic compound indole is known for its floral odor typical of jasmine blossoms. Due to its characteristic scent, it is frequently used in dairy products, tea drinks and fine fragrances. The demand for natural indole by the flavor and fragrance industry is high, yet, its abundance in essential oils isolated from plants such as jasmine and narcissus is low. Thus, there is a strong demand for a sustainable method to produce food-grade indole. RESULTS: Here, we established the biotechnological production of indole upon L-tryptophan supplementation in the bacterial host Corynebacterium glutamicum. Heterologous expression of the tryptophanase gene from E. coli enabled the conversion of supplemented L-tryptophan to indole. Engineering of the substrate import by co-expression of the native aromatic amino acid permease gene aroP increased whole-cell biotransformation of L-tryptophan to indole by two-fold. Indole production to 0.2 g L-1 was achieved upon feeding of 1 g L-1 L-tryptophan in a bioreactor cultivation, while neither accumulation of side-products nor loss of indole were observed. To establish an efficient and robust production process, new tryptophanases were recruited by mining of bacterial sequence databases. This search retrieved more than 400 candidates and, upon screening of tryptophanase activity, nine new enzymes were identified as most promising. The highest production of indole in vivo in C. glutamicum was achieved based on the tryptophanase from Providencia rettgeri. Evaluation of several biological aspects identified the product toxicity as major bottleneck of this conversion. In situ product recovery was applied to sequester indole in a food-grade organic phase during the fermentation to avoid inhibition due to product accumulation. This process enabled complete conversion of L-tryptophan and an indole product titer of 5.7 g L-1 was reached. Indole partitioned to the organic phase which contained 28 g L-1 indole while no other products were observed indicating high indole purity. CONCLUSIONS: The bioconversion production process established in this study provides an attractive route for sustainable indole production from tryptophan in C. glutamicum. Industrially relevant indole titers were achieved within 24 h and indole was concentrated in the organic layer as a pure product after the fermentation.

Bioproduction of 2-Phenylethanol through Yeast Fermentation on Synthetic Media and on Agro-Industrial Waste and By-Products: A Review.

Due to its pleasant rosy scent, the aromatic alcohol 2-phenylethanol (2-PE) has a huge market demand. Since this valuable compound is used in food, cosmetics and pharmaceuticals, consumers and safety regulations tend to prefer natural methods for its production rather than the synthetic ones. Natural 2-PE can be either produced through the extraction of essential oils from various flowers, including roses, hyacinths and jasmine, or through biotechnological routes. In fact, the rarity of natural 2-PE in flowers has led to the inability to satisfy the large market demand and to a high selling price. Hence, there is a need to develop a more efficient, economic, and environmentally friendly biotechnological approach as an alternative to the conventional industrial one. The most promising method is through microbial fermentation, particularly using yeasts. Numerous yeasts have the ability to produce 2-PE using l-Phe as precursor. Some agro-industrial waste and by-products have the particularity of a high nutritional value, making them suitable media for microbial growth, including the production of 2-PE through yeast fermentation. This review summarizes the biotechnological production of 2-PE through the fermentation of different yeasts on synthetic media and on various agro-industrial waste and by-products.

Intensification of p-coumaric acid heterologous production using extractive biphasic fermentation.

p-coumaric acid (p-CA) can be produced from D-glucose by an engineered S. cerevisiae strain. p-CA has antimicrobial properties and retro-inhibition activity. Moreover, p-CA is a hydrophobic compound, limiting its accumulation in fermentation broth. To overcome these issues all at once, a liquid-liquid extraction in-situ product recovery process using oleyl alcohol as extractant has been implemented in order to continuously extract p-CA from the broth. Media and pH impacts on strain metabolism were assessed, highlighting p-CA decarboxylase endogenous activity. Biphasic fermentations allowed an increase in p-CA respiratory production rates at both pH assessed (13.65 and 9.45 mg L-1.h-1 at pH 6 and 4.5, respectively) compared to control ones (10.5 and 7.5 mg L-1.h-1 at pH 6 and 4.5, respectively). Biphasic fermentation effects on p-CA decarboxylation were studied showing that continuous removal of p-CA decreased its decarboxylation into 4-vinylphenol at pH 4.5 (57 mg L-1 in biphasic fermentation vs 173 mg L-1 in control one).

In situ product recovery techniques aiming to obtain biotechnological products: A glance to current knowledge.

Biotechnology and bioengineering techniques have been widely used in the production of biofuels, chemicals, pharmaceuticals, and food additives, being considered a "green" form of production because they use renewable and nonpolluting energy sources. On the other hand, in the traditional processes of production, the target product obtained by biotechnological routes must undergo several stages of purification, which makes these processes more expensive. In the past few years, some works have focused on processes that integrate fermentation to the recovery and purification steps necessary to obtain the final product required. This type of process is called in situ product recovery or extractive fermentation. However, there are some differences in the concepts of the techniques used in these bioprocesses. In this way, this review sought to compile relevant content on considerations and procedures that are being used in this field, such as evaporation, liquid-liquid extraction, permeation, and adsorption techniques. Also, the objective of this review was to approach the different configurations in the recent literature of the processes employed and the main bioproducts obtained, which can be used in the food, pharmaceutical, chemical, and/or fuel additives industry. We intended to elucidate concepts of these techniques, considered very recent, but which emerge as a promising alternative for the integration of bioprocesses.

Using poly(vinyldodecylimidazolium bromide) for the in-situ product recovery of n-butanol.

The mitigation of end-product inhibition during the biosynthesis of n-butanol is demonstrated for an in-situ product recovery (ISPR) system employing a poly(ionic liquid) (PIL) absorbent. The thermodynamic affinity of poly(vinyldodecylimidazolium bromide) [P(VC12 ImBr)] for n-butanol, acetone and ethanol versus water was measured at conditions experienced in a typical acetone-ethanol-butanol (ABE) fermentation. In addition to providing a high n-butanol partition coefficient (PC = 6.5) and selectivity (αBuOH/water = 46), P(VC12 ImBr) is shown to be biocompatible with Saccharomyces cerevisiae and Clostridium acetobutylicum. Furthermore, the diffusivity of n-butanol in a hydrated PIL provides absorption rates that support ISPR applications. Using a 5 wt% PIL phase fraction relative to the aqueous phase mass, P(VC12 ImBr) improved the volumetric productivity of a batch ABE ISPR process by 31% relative to a control fermentation. The concentration of n-butanol in the P(VC12 ImBr) phase was sufficient to increase the alcohol concentration from 1.5 wt% in the fermentation medium to 25 wt% in the saturated PIL, thereby facilitating downstream n-butanol recovery.

Improved Production and In Situ Recovery of Sesquiterpene (+)-Zizaene from Metabolically-Engineered E. coli.

The sesquiterpene (+)-zizaene is the direct precursor of khusimol, the main fragrant compound of the vetiver essential oil from Chrysopogon zizanioides and used in nearly 20% of men's fine perfumery. The biotechnological production of such fragrant sesquiterpenes is a promising alternative towards sustainability; nevertheless, product recovery from fermentation is one of the main constraints. In an effort to improve the (+)-zizaene recovery from a metabolically-engineered Escherichia coli, we developed an integrated bioprocess by coupling fermentation and (+)-zizaene recovery using adsorber extractants. Initially, (+)-zizaene volatilization was confirmed from cultivations with no extractants but application of liquid-liquid phase partitioning cultivation (LLPPC) improved (+)-zizaene recovery nearly 4-fold. Furthermore, solid-liquid phase partitioning cultivation (SLPPC) was evaluated by screening polymeric adsorbers, where Diaion HP20 reached the highest recovery. Bioprocess was scaled up to 2 L bioreactors and in situ recovery configurations integrated to fermentation were evaluated. External recovery configuration was performed with an expanded bed adsorption column and improved (+)-zizaene titers 2.5-fold higher than LLPPC. Moreover, internal recovery configuration (IRC) further enhanced the (+)-zizaene titers 2.2-fold, whereas adsorption velocity was determined as critical parameter for recovery efficiency. Consequently, IRC improved the (+)-zizaene titer 8.4-fold and productivity 3-fold from our last report, achieving a (+)-zizaene titer of 211.13 mg L-1 and productivity of 3.2 mg L-1 h-1. This study provides further knowledge for integration of terpene bioprocesses by in situ product recovery, which could be applied for many terpene studies towards the industrialization of fragrant molecules.

Stabilization and scale-up of photosynthesis-driven ω-hydroxylation of nonanoic acid methyl ester by two-liquid phase whole-cell biocatalysis.

Photoautotrophic organisms are promising hosts for biocatalytic oxyfunctionalizations because they supply reduction equivalents as well as O2 via photosynthetic water oxidation. Thus far, research on photosynthesis-driven bioprocesses mainly focuses on strain development and the proof of principle in small-scale biocatalytic reaction setups. This study investigates the long-term applicability of the previously developed cyanobacterial strain Synechocystis sp. PCC 6803_BGT harboring the alkane monooxygenase system AlkBGT catalyzing terminal alkyl group oxyfunctionalization. For the regiospecific ω-hydroxylation of nonanoic acid methyl ester (NAME), this biocatalyst showed light intensity-independent hydroxylation activity and substantial hydrolysis of NAME to nonanoic acid. Substrate mass transfer limitation, substrate hydrolysis, as well as reactant toxicity were overcome via in situ substrate supply by means of a two-liquid phase system. The application of diisononyl phthalate as organic carrier solvent enabled 1.7-fold increased initial specific activities (5.6 ± 0.1 U/gCDW ) and 7.6-fold increased specific yields on biomass (3.8 ± 0.1 mmolH-NAME /gCDW ) as compared with single aqueous phase biotransformations. Finally, the whole-cell biotransformation system was successfully scaled from glass tubes to a stirred-tank photobioreactor. This is the first study reporting the application of the two-liquid phase concept for efficient phototrophic whole-cell biocatalysis.

Enhanced production of styrene by engineered Escherichia coli and in situ product recovery (ISPR) with an organic solvent.

BACKGROUND: Styrene is a large-volume commodity petrochemical, which has been used in a wide range of polymer industry as the main building block for the construction of various functional polymers. Despite many efforts to produce styrene in microbial hosts, the production titers are still low and are not enough to meet the commercial production of styrene. RESULTS: Previously, we developed a high L-phenylalanine producer (E. coli YHP05), and it was used as a main host for de novo synthesis of styrene. First, we introduced the co-expression system of phenylalanine-ammonia lyase (PAL) and ferulic acid decarboxylase (FDC) genes for the synthesis of styrene from L-phenylalanine. Then, to minimize cell toxicity and enhance the recovery of styrene, in situ product recovery (ISPR) with n-dodecane was employed, and culture medium with supplementation of complex sources was also optimized. As a result, 1.7 ± 0.1 g/L of styrene was produced in the flask cultures. Finally, fed-batch cultivations were performed in lab-scale bioreactor, and to minimize the loss of volatile styrene during the cultivation, three consecutive bottles containing n-dodecane were connected to the air outlet of bioreactor for gas-stripping. To conclude, the total titer of styrene was as high as 5.3 ± 0.2 g/L, which could be obtained at 60 h. CONCLUSION: We successfully engineered E. coli strain for the de novo production of styrene in both flask and fed-batch cultivation, and could achieve the highest titer for styrene in bacterial hosts reported till date. We believe that our efforts in strain engineering and ISPR strategy with organic solvent will provide a new insight for economic and industrial production of styrene in a biological platform.

Solvent-free membrane extraction of volatile fatty acids from acidogenic fermentation.

Diversification of anaerobic digestion into higher value products, namely volatile fatty acids (VFAs), is receiving interest. One of the biggest challenges with this is recovery of the VFAs. Membrane extraction can be used, and a novel process configuration using a non-porous silicone membrane and water for an extractant is proposed here. This process would enable the reduction in the number of downstream unit operations compared to other membrane extraction processes. Selective recovery in favour of longer chain VFAs was demonstrated. Testing with a synthetic solution resulted in an overall mass transfer coefficient of 0.088 µm s-1 for butyric acid, and 0.157 µm s-1 when fermentation broth was used. This indicates this process is not hindered by fouling, but improved somehow. Although the preliminary economic analysis showed this process to require a larger membrane area compared to porous membrane alternatives, it also has a significantly reduced cost associated with the extractant.

Resolving mismatches in the flexible production of ethanol and butanol from eucalyptus wood with vacuum fermentation.

In flexible ethanol-butanol plants, low tolerance to butanol by solventogenic clostridia (and resulting dilute fermentation) results in considerable number of empty fermentors whenever production focuses on ethanol. This research identified scenarios in which vacuum fermentation (in-situ vacuum recovery) may be applied to solve this problem. We conducted ethanol (Saccharomyces cerevisiae) and ABE (Clostridium beijerinckii NCIMB 8052) batch vacuum fermentations of eucalyptus hydrolysates according to the distribution of sugars in a flexible plant. Based on the experiments and performance targets set for the ABE fermentation, we simulated a flexible plant that processes 1000 dry t eucalyptus/day using pretreatment and enzymatic hydrolysis steps with moderate solids loading (15% w/w). The simulation showed that the number of fermentation tanks can decrease by 62% (eliminating 10 idle tanks, 3748 m3 each) by applying vacuum recovery only to the fermentation of mixed (cellulose + hemicellulose) hydrolysates to ABE. We concluded that this configuration can result in savings of up to 2 MMUS$/year in comparison with flexible plants having only conventional batch fermentors, and additional cost savings are expected from reduced wastewater footprint.

A novel close-circulating vapor stripping-vapor permeation technique for boosting biobutanol production and recovery.

BACKGROUND: Butanol derived from renewable resources by microbial fermentation is considered as one of not only valuable platform chemicals but alternative advanced biofuels. However, due to low butanol concentration in fermentation broth, butanol production is restricted by high energy consumption for product recovery. For in situ butanol recovery techniques, such as gas stripping and pervaporation, the common problem is their low efficiency in harvesting and concentrating butanol. Therefore, there is a necessity to develop an advanced butanol recovery technique for cost-effective biobutanol production. RESULTS: A close-circulating vapor stripping-vapor permeation (VSVP) process was developed with temperature-difference control for single-stage butanol recovery. In the best scenario, the highest butanol separation factor of 142.7 reported to date could be achieved with commonly used polydimethylsiloxane membrane, when temperatures of feed solution and membrane surroundings were 70 and 0 °C, respectively. Additionally, more ABE (31.2 vs. 17.7 g/L) were produced in the integrated VSVP process, with a higher butanol yield (0.21 vs. 0.17 g/g) due to the mitigation of butanol inhibition. The integrated VSVP process generated a highly concentrated permeate containing 212.7 g/L butanol (339.3 g/L ABE), with the reduced energy consumption of 19.6 kJ/g-butanol. CONCLUSIONS: Therefore, the present study demonstrated a well-designed energy-efficient technique named by vapor stripping-vapor permeation for single-stage butanol removal. The butanol separation factor was multiplied by the temperature-difference control strategy which could double butanol recovery performance. This advanced VSVP process can completely eliminate membrane fouling risk for fermentative butanol separation, which is superior to other techniques.