Solid-liquid Two-phase Partitioning Bioreactor Research Articles

Improved reactor performance and operability in the biotransformation of carveol to carvone using a solid–liquid two‐phase partitioning bioreactor

In an effort to improve reactor performance and process operability, the microbial biotransformation of (-)-trans-carveol to (R)-(-)-carvone by hydrophobic Rhodococcus erythropolis DCL14 was carried out in a two phase partitioning bioreactor (TPPB) with solid polymer beads acting as the partitioning phase. Previous work had demonstrated that the substrate and product become inhibitory to the organism at elevated aqueous concentrations and the use of an immiscible second phase in the bioreactor was intended to provide a reservoir for substrates to be delivered to the aqueous phase based on the metabolic rate of the cells, while also acting as a sink to uptake the product as it is produced. The biotransformation was previously undertaken in a two liquid phase TPPB with 1-dodecene and with silicone oil as the immiscible second phase and, although improvement in the reactor performance was obtained relative to a single phase system, the hydrophobic nature of the organism caused the formation of severe emulsions leading to significant operational challenges. In the present work, eight types of polymer beads were screened for their suitability for use in a solid-liquid TPPB for this biotransformation. The use of selected solid polymer beads as the second phase completely prevented emulsion formation and therefore improved overall operability of the reactor. Three modes of solid-liquid TPPB operation were considered: the use of a single polymer bead type (styrene/butadiene copolymer) in the reactor, the use of a mixture of polymer beads in the reactor (styrene/butadiene copolymer plus Hytrel(R) 8206), and the use of one type of polymer beads in the reactor (styrene/butadiene copolymer), and another bead type (Hytrel(R) 8206) in an external column through which fermentation medium was recirculated. This last configuration achieved the best reactor performance with 7 times more substrate being added throughout the biotransformation relative to a single aqueous phase benchmark reactor and 2.7 times more substrate being added relative to the best two liquid TPPB case. Carvone was quantitatively recovered from the polymer beads via single stage extraction into methanol, allowing for bead re-use.

Solid-liquid two-phase partitioning bioreactors for the treatment of gas-phase volatile organic carbons (VOCs) by a microbial consortium

A two-phase partitioning bioreactor (TPPB), employing styrene-butadiene co-polymer beads as the sequestering/delivery phase, was used to treat high step change loadings of toluene in a contaminated air stream. The polymers, which are biocompatible and non-bioavailable, allowed the use of a microbial consortium and effectively absorbed and released the toluene vapours for biodegradation, while providing a buffering effect against high toluene transients. Toluene loadings were increased from a base steady state rate of 343-6,000 g/m(3) h for 1 h periods, with the polymer-aqueous system substantially outperforming a single phase system on the basis of improving the toluene removal efficiency and reducing the maximum toluene concentrations emitted during the transients.

Biodegradation of PCBs in two‐phase partitioning bioreactors following solid extraction from soil

This article demonstrates the feasibility of a novel process concept for the remediation of PCB contaminated soil. The proposed process consists of PCB extraction from soil using solid polymer beads, followed by biodegradation of the extracted PCBs in a solid-liquid two-phase partitioning bioreactor (TPPB), where PCBs are delivered from the polymer beads to the degrading organisms. The commercially available thermoplastic polymer Hytrel was used to extract Aroclor 1242 from contaminated artificial soil in bench scale experiments. Initial PCB contamination levels of 100 and 1,000 mg kg(-1) could be reduced to 32% +/- 1 to 41% +/- 7 of the initial value after 48 h mixing in the presence of a mobilizing agent at polymer-to-soil ratios of 1% (w/w) and 10% (w/w). The decrease of detectable PCBs in the soil was consistent with an increase of PCBs in the polymer beads. It was further shown that Aroclor 1242 could be delivered to the PCB degrading organism Burkholderia xenovorans LB400 in a solid-liquid TPPB via Hytrel beads. A total of 70 mg Aroclor 1242 could be degraded in a 1 L solid-liquid TPPB within 80 h of operation.

Polymer Selection for Biphenyl Degradation in a Solid-Liquid Two-Phase Partitioning Bioreactor

The commercially available thermoplastic polymer Hytrel was selected as the delivery phase for the hydrophobic model compound biphenyl in a solid-liquid two-phase partitioning bioreactor (TPPB), and 2.9 g biphenyl could successfully be degraded in 1-L TPPBs by a pure culture of the biphenyl-degrading bacterium Burkholderia xenovorans LB400 in 50 h and by a mixed microbial consortium isolated from contaminated soil in 45 h. TPPBs consist of an aqueous cell-containing phase and an immiscible second phase that partitions toxic and/or poorly soluble substrates (in this case biphenyl) on the basis of maintaining a thermodynamic equilibrium. This paper illustrates a rational strategy for selecting a suitable solid polymeric substance for the delivery of the poorly water-soluble model compound biphenyl. The partitioning of biphenyl between the selected polymers and water was analogous to partitioning of solutes between two immiscible liquid phases. The partitioning coefficients varied between 180 for Nylon 6.6 and 11,000 for Desmopan, where the later numerical value is comparable to biphenyl partitioning coefficients between water and organic solvents. Employing a solid delivery phase enabled the utilization of a surfactant-producing microbial mixed culture, which could not be cultivated in liquid-liquid TPPBs and thereby extended the range of biocatalysts that can be employed in TPPBs.

A novel solid–liquid two‐phase partitioning bioreactor for the enhanced bioproduction of 3‐methylcatechol

The bioproduction of 3-methylcatechol from toluene via Pseudomonas putida MC2 was performed in a solid-liquid two-phase partitioning bioreactor with the intent of increasing yield and productivity over a single-phase system. The solid phase consisted of HYTREL, a thermoplastic polymer that was shown to possess superior affinity for the inhibitory 3-methylcatechol compared to other candidate polymers as well as a number of immiscible organic solvents. Operation of a solid-liquid biotransformation utilizing a 10% (w/w) solid (polymer beads) to liquid phase ratio resulted in the bioproduction of 3-methylcatechol at a rate of 350 mg/L-h, which compares favorably to the single phase productivity of 128 mg/L-h. . HYTREL polymer beads were also reconstituted into polymer sheets, which were placed around the interior circumference of the bioreactor and successfully removed 3-methylcatechol from solution resulting in a rate of 3-methylcatechol production of 343 mg/L-h. Finally, a continuous biotransformation was performed in which culture medium was circulated upwards through an external extraction column containing HYTREL beads. The design maintained sub lethal concentrations of 3-methylcatechol within the bioreactor by absorbing produced 3-methylcatechol into the polymer beads. As 3-methylcatechol concentrations in the aqueous phase approached 500 mg/L the extraction column was replaced (twice) with a fresh column and the process was continued representing a simple and effective approach for the continuous bioproduction of 3-methylcatechol. Recovery of 3-methylcatechol from HYTREL was also achieved by bead desorption into methanol.

Biodegradation of biphenyl in a solid–liquid two-phase partitioning bioreactor

Biphenyl was successfully degraded by Burkholderia xenovorans LB400, initially described as Pseudomonas sp. LB400, in a solid–liquid two-phase partitioning bioreactor (TPPB). Solid–liquid TPPBs are comprised of an aqueous, cell containing phase, and a solid polymeric phase that partitions toxic and/or poorly soluble substrates (in this case biphenyl) based on maintaining a thermodynamic equilibrium. The employed polymer was Hytrel™, a thermoplastic polyester elastomer. The surface area available for mass transfer of biphenyl was limiting and resulted in mass transfer limited growth, as demonstrated experimentally by employing two different geometric shapes (cylinders with different aspect ratios) of the polymer phase with different specific surface area, while keeping all other parameters constant. The linear microbial growth rates were substantially higher when more polymer surface area was provided. The mass transfer coefficient of biphenyl from Hytrel™ to water was measured under experimental conditions, which allowed predicting the release rate based on the biphenyl concentration gradient. The partitioning behaviour of biphenyl between Hytrel™ and culture medium was measured as well, which allowed the development of a simple mechanistic model describing microbial growth based on known microbial properties in combination with substrate delivery from the solid polymer phase. The model was capable of describing the experimental data well and can be used to predict degradation rates for other geometric shapes of a solid delivery phase such as sheets or rods, which might be of operational advantage in various applications of TPPBs to the controlled uptake and release of other recalcitrant molecules.

Polymer Selection for Biphenyl Degradation in a Solid-Liquid Two-Phase Partitioning Bioreactor

Polymer Selection for Biphenyl Degradation in a Solid-Liquid Two-Phase Partitioning Bioreactor

Oxygen transfer in a gas–liquid system containing solids of varying oxygen affinity

An air sparged, mechanically agitated bioreactor containing spherical solids was studied in order to determine the effect of the solid phase on oxygen mass transfer. It was found that both nylon 6,6 and glass beads cause an enhancement of the volumetric mass transfer coefficient of up to 268%, whereas particles of silicone rubber and styrene–butadiene copolymer reduce the volumetric mass transfer coefficient by up to 63%, relative to a system without a solid phase. A simple transport in series model has been proposed to account for the observed phenomena, which includes both the physical enhancement effects of particles on gas–liquid mass transfer as well as absorption of oxygen into the polymer. Even though volumetric mass transfer coefficient reductions were observed in the system containing silicone rubber, it was demonstrated that an increased oxygen transfer rate into the working volume of a two-phase system occurs relative to a system without a solid phase. This study provides an explanation for previous results regarding the enhanced effect of the presence of a styrene–butadiene copolymer phase in reducing oxygen limitations in a solid–liquid two-phase partitioning bioreactor. Results from this study can be applied to two-phase aerobic fermentation systems that will benefit from reducing oxygen limiting conditions.

Biodegradation of a phenolic mixture in a solid–liquid two-phase partitioning bioreactor

A solid-liquid two-phase partitioning bioreactor (TPPB) in which the non-aqueous phase consisted of polymer (HYTREL) beads was used to degrade a model mixture of phenols [phenol, o-cresol, and 4-chlorophenol (4CP)] by a microbial consortium. In one set of experiments, high concentrations (850 mg l(-1) of each of the three substrates) were reduced to sub-inhibitory levels within 45 min by the addition of the polymer beads, followed by inoculation and rapid (8 h) consumption of the total phenolics loading. In a second set of experiments, the beneficial effect of using polymer beads to launch a fermentation inhibited by high substrate concentrations was demonstrated by adding 1,300 and 2,000 mg l(-1) total substrates (equal concentrations of each phenolic) to a pre-inoculated bioreactor. At these levels, no cell growth and no degradation were observed; however, after adding polymer beads to the systems, the ensuing reduced substrate concentrations permitted complete destruction of the target molecules, demonstrating the essential role played by the polymer sequestering phase when applied to systems facing inhibitory substrate concentrations. In addition to establishing alternative modes of TPPB operation, the present work has demonstrated the differential partitioning of phenols in a mixture between the aqueous and polymeric phases. The polymeric phase was also observed to absorb a degradation intermediate (arising from the incomplete biodegradation of 4CP), which opens the possibility of using solid-liquid TPPBs during biosynthetic transformation to sequester metabolic byproducts.

Polymer Development for Enhanced Delivery of Phenol in a Solid-Liquid Two-Phase Partitioning Bioreactor

Two-Phase Partitioning Bioreactors (TPPBs) have traditionally been used to partition toxic concentrations of xenobiotics from a cell-containing aqueous phase by means of an immiscible organic solvent and to deliver these substrates back to the cells on the basis of metabolic demand and the maintenance of thermodynamic equilibrium between the phases. A limitation of TPPBs, which use organic liquid solvents, is the possibility that the solvent can be bioavailable, and this has therefore limited organic liquid TPPBs to the use of pure strains of microbes. Solid polymer beads have recently been introduced as a replacement for liquid organic solvents, offering similar absorption properties but with the capability to be used with mixed microbial populations. The present work was aimed at identifying a polymer with a greater capacity for and more rapid uptake and release of phenol for use as the second phase in a mixed culture TPPB. Polarity and hydrogen bonding capabilities between polymer and phenol were considered in the screening and selection process of candidate polymers. Hytrel (a copolymer of poly(butylene terephthalate) and butylene ether glycol terephthalate) polymer beads, offered improved capacity (19 mg phenol/g polymer at a fixed initial phenol concentration of 2000 mg/L) and a greater diffusivity (1.54 x 10(-7) cm2/s) when compared to the capacity and diffusivity of previously used EVA (ethylene vinyl acetate) beads (12.4 mg phenol/g polymer and 3.73 x 10(-9) cm2/s, respectively). Hytrel polymer beads were then used in a TPPB for the investigation of various substrate feeding strategies (fed-batch, bead replacement, and concentrated spikes of phenol), with rapid and complete phenol degradation shown in all cases.