Crystallization Assisted Dynamic Kinetic Resolution for the Synthesis of (R)-ß-Methylphenethylamine.

This study explores a combination of the concept of enantioselective enzymatic synthesis of ß-chiral amines through transamination with in situ product crystallization (ISPC) to overcome product inhibition. Using 2-phenylpropanal as a readily available and easily racemizing substrate of choice, (R)-ß-methylphenethylamine ((R)-2-phenylpropan-1-amine) concentrations of up to 250 mM and enantiomeric excesses of up to 99 % are achieved when using a commercially available transaminase from Ruegeria pomeroyi in a fed-batch based dynamic kinetic resolution reaction on preparative scale. The source of substrate decomposition during the reaction is also investigated and the resulting unwanted byproduct formation is successfully reduced to insignificant levels.

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.

Honoring two stalwarts of photosynthesis research: Eva-Mari Aro and Govindjee.

On behalf of the entire photosynthesis community, it is an honor, for us, to write about two very eminent scientists who were recently recognised with a Lifetime Achievement Award from the International Society of Photosynthesis Research (ISPR) on August 5, 2022; this prestigious Award was given during the closing ceremony of the 18th International Congress on Photosynthesis Research in Dunedin, New Zealand. The awardees were: Professor Eva-Mari Aro (Finland) and Professor Emeritus Govindjee Govindjee (USA). One of the authors, Anjana Jajoo, is especially delighted to be a part of this tribute to professors Aro and Govindjee as she was lucky enough to have worked with both of them.

Foam fractionation methods in aerobic fermentation processes.

Inherently occurring foam formation during aerobic fermentation of surface-active compounds can be exploited by fractionating the foam. This also serves as the first downstream processing step for product concentration and is used for in situ product recovery. Compared to other foam prevention methods, it does not interfere with fermentation parameters or alter broth composition. Nevertheless, parameters affecting the foaming behavior are complex. Therefore, the specific foam fractionation designs need to be engineered for each fermentation individually. This still hinders a widespread industrial application. However, few available commercial approaches demonstrate the applicability of foam columns on an industrial scale. This systematic literature review highlights relevant design aspects and process demands that need to be considered for an application to fermentations and proposes a classification of foam fractionation designs and methods. It further analyses substance-specific characteristics associated with foam fractionation. Finally, solutions for current challenges are presented, and future perspectives are discussed.

Design and evaluation of a continuous semipartition bioreactor for in situ liquid-liquid extractive fermentation.

Extractive fermentation (or in situ product removal (ISPR)) is an operational method used to combat product inhibition in fermentations. To achieve ISPR, different separation techniques, modes of operation and physical reactor configurations have been proposed. However, the relative paucity of industrial application necessitates continued investigation into reactor systems. This article outlines a bioreactor designed to facilitate in situ product extraction and recovery, through adapting the reaction volume to include a settler and solvent extraction and recycle section. This semipartition bioreactor is proposed as a new mode of operation for continuous liquid-liquid extractive fermentation. The design is demonstrated as a modified bench-top fermentation vessel, initially analysed in terms of fluid dynamic studies, in a model two-liquid phase system. A continuous abiotic simulation of lactic acid (LA) fermentation is then demonstrated. The results show that mixing in the main reaction vessel is unaffected by the inserted settling zone, and that the size of the settling tube effects the maximum volumetric removal rate. In these tests the largest settling tube gave a potential continuous volumetric removal rate of 7.63 ml/min; sufficiently large to allow for continuous product extraction even in a highly productive fermentation. To demonstrate the applicability of the developed reactor, an abiotic simulation of a LA fermentation was performed. LA was added to reactor continuously at a rate of 33ml/h, while continuous in situ extraction removed the LA using 15% trioctylamine in oleyl alcohol. The reactor showed stable LA concentration of 1 g/L, with the balance of the LA successfully extracted and recovered using back extraction. This study demonstrates a potentially useful physical configuration for continuous in situ extraction.

In-situ muconic acid extraction reveals sugar consumption bottleneck in a xylose-utilizing Saccharomyces cerevisiae strain.

BACKGROUND: The current shift from a fossil-resource based economy to a more sustainable, bio-based economy requires development of alternative production routes based on utilization of biomass for the many chemicals that are currently produced from petroleum. Muconic acid is an attractive platform chemical for the bio-based economy because it can be converted in chemicals with wide industrial applicability, such as adipic and terephthalic acid, and because its two double bonds offer great versatility for chemical modification. RESULTS: We have constructed a yeast cell factory converting glucose and xylose into muconic acid without formation of ethanol. We consecutively eliminated feedback inhibition in the shikimate pathway, inserted the heterologous pathway for muconic acid biosynthesis from 3-dehydroshikimate (DHS) by co-expression of DHS dehydratase from P. anserina, protocatechuic acid (PCA) decarboxylase (PCAD) from K. pneumoniae and oxygen-consuming catechol 1,2-dioxygenase (CDO) from C. albicans, eliminated ethanol production by deletion of the three PDC genes and minimized PCA production by enhancing PCAD overexpression and production of its co-factor. The yeast pitching rate was increased to lower high biomass formation caused by the compulsory aerobic conditions. Maximal titers of 4 g/L, 4.5 g/L and 3.8 g/L muconic acid were reached with glucose, xylose, and a mixture, respectively. The use of an elevated initial sugar level, resulting in muconic acid titers above 2.5 g/L, caused stuck fermentations with incomplete utilization of the sugar. Application of polypropylene glycol 4000 (PPG) as solvent for in situ product removal during the fermentation shows that this is not due to toxicity by the muconic acid produced. CONCLUSIONS: This work has developed an industrial yeast strain able to produce muconic acid from glucose and also with great efficiency from xylose, without any ethanol production, minimal production of PCA and reaching the highest titers in batch fermentation reported up to now. Utilization of higher sugar levels remained conspicuously incomplete. Since this was not due to product inhibition by muconic acid or to loss of viability, an unknown, possibly metabolic bottleneck apparently arises during muconic acid fermentation with high sugar levels and blocks further sugar utilization.

Current status and perspectives of 2-phenylethanol production through biological processes.

2-Phenylethanol (2-PE), an important flavor and fragrance compound with a rose-like smell has been widely used in the cosmetics, perfume, and food industries. Traditional production of 2-PE was mainly through the extraction from plant materials or by chemical synthesis. However, the increasing demand of consumers for natural flavors cannot be met by these methods. Biological production of 2-PE has emerged to be an appealing solution due to an environmental friendly process and the definition of a "natural" product. In this review, we have comprehensively summarized the current status and perspectives for biological 2-PE production in terms of its advantages over classical chemical synthesis and extraction from natural plants. A comprehensive description of 2-PE synthetic pathways and global regulation mechanisms, strategies to increase 2-PE production, and the utilization of agro-industrial wastes as feedstocks has been systematically discussed. Furthermore, the application of in situ product removal techniques have also been highlighted.

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.

Development of in situ product removal strategies in biocatalysis applying scaled-down unit operations.

An experimental platform based on scaled-down unit operations combined in a plug-and-play manner enables easy and highly flexible testing of advanced biocatalytic process options such as in situ product removal (ISPR) process strategies. In such a platform, it is possible to compartmentalize different process steps while operating it as a combined system, giving the possibility to test and characterize the performance of novel process concepts and biocatalysts with minimal influence of inhibitory products. Here the capabilities of performing process development by applying scaled-down unit operations are highlighted through a case study investigating the asymmetric synthesis of 1-methyl-3-phenylpropylamine (MPPA) using ω-transaminase, an enzyme in the sub-family of amino transferases (ATAs). An on-line HPLC system was applied to avoid manual sample handling and to semi-automatically characterize ω-transaminases in a scaled-down packed-bed reactor (PBR) module, showing MPPA as a strong inhibitor. To overcome the inhibition, a two-step liquid-liquid extraction (LLE) ISPR concept was tested using scaled-down unit operations combined in a plug-and-play manner. Through the tested ISPR concept, it was possible to continuously feed the main substrate benzylacetone (BA) and extract the main product MPPA throughout the reaction, thereby overcoming the challenges of low substrate solubility and product inhibition. The tested ISPR concept achieved a product concentration of 26.5 gMPPA · L-1 , a purity up to 70% gMPPA · gtot-1 and a recovery in the range of 80% mol · mol-1 of MPPA in 20 h, with the possibility to increase the concentration, purity, and recovery further. Biotechnol. Bioeng. 2017;114: 600-609. © 2016 Wiley Periodicals, Inc.

Demonstration of in situ product recovery of butyric acid via CO2 -facilitated pH swings and medium development in two-phase partitioning bioreactors.

Production of organic acids in solid-liquid two-phase partitioning bioreactors (TPPBs) is challenging, and highly pH-dependent, as cell growth occurs near neutral pH, while acid sorption occurs only at low pH conditions. CO2 sparging was used to achieve acidic pH swings, facilitating undissociated organic acid uptake without generating osmotic stress inherent in traditional acid/base pH control. A modified cultivation medium was formulated to permit greater pH reduction by CO2 sparging (pH 4.8) compared to typical media (pH 5.3), while still possessing adequate nutrients for extensive cell growth. In situ product recovery (ISPR) of butyric acid (pKa = 4.8) produced by Clostridium tyrobutyricum was achieved through intermittent CO2 sparging while recycling reactor contents through a column packed with absorptive polymer Hytrel® 3078. This polymer was selected on the basis of its composition as a polyether copolymer, and the use of solubility parameters for predicting solute polymer affinity, and was found to have a partition coefficient for butyric acid of 3. Total polymeric extraction of 3.2 g butyric acid with no CO2 mediated pH swings was increased to 4.5 g via CO2 -facilitated pH shifting, despite the buffering capacity of butyric acid, which resists pH shifting. This work shows that CO2 -mediated pH swings have an observable positive effect on organic acid extraction, with improvements well over 150% under optimal conditions in early stage fermentation compared to CO2 -free controls, and this technique can be applied other organic acid fermentations to achieve or improve ISPR.

The use of high pressure CO2 -facilitated pH swings to enhance in situ product recovery of butyric acid in a two-phase partitioning bioreactor.

Through the use of high partial pressures of CO2 (pCO2 ) to facilitate temporary pH reductions in two-phase partitioning bioreactors (TPPBs), improved pH dependent partitioning of butyric acid was observed which achieved in situ product recovery (ISPR), alleviating end-product inhibition (EPI) during the production of butyric acid by Clostridium tyrobutyricum (ATCC 25755). Through high pressure pCO2 studies, media buffering effects were shown to be substantially overcome at 60 bar pCO2 , resulting in effective extraction of the organic acid by the absorptive polymer Pebax® 2533, yielding a distribution coefficient (D) of 2.4 ± 0.1 after 1 h of contact at this pressure. Importantly, it was also found that C. tyrobutyricum cultures were able to withstand 60 bar pCO2 for 1 h with no decrease in growth ability when returned to atmospheric pressure in batch reactors after several extraction cycles. A fed-batch reactor with cyclic high pCO2 polymer extraction recovered 92 g of butyric acid to produce a total of 213 g compared to 121 g generated in a control reactor. This recovery reduced EPI in the TPPB, resulting in both higher productivity (0.65 vs. 0.33 g L(-1) h(-1) ) and yield (0.54 vs. 0.40). Fortuitously, it was also found that repeated high pCO2 -facilitated polymer extractions of butyric acid during batch growth of C. tyrobutyricum lessened the need for pH control, and reduced base requirements by approximately 50%. Thus, high pCO2 -mediated absorptive polymer extraction presents a novel method for improving process performance in butyric acid fermentation, and this technique could be applied to the bioproduction of other organic acids as well.

Production of (hydroxy)benzoate-derived polyketides by engineered Pseudomonas with in situ extraction.

Polyketides from (hydroxy)benzoates are an interesting group of plant polyphenolic compounds, whose biotechnological production is so far underrepresented due to their challenging heterologous biosynthesis. Efficient heterologous production of 2,4,6-tri- and 2,3',4,6-tetrahydroxybenzophenone, 3,5-dihydroxybiphenyl, and 4-hydroxycoumarin by whole-cell biocatalysis in combination with in situ product extraction with an organic solvent was demonstrated. Production was highly dependent on the used CoA ligase and polyketide synthase type III. Therefore, different combinations of polyketide synthases and benzoate-CoA ligases were evaluated for their biosynthesis performance in the solvent-tolerant Pseudomonas taiwanensis VLB120. A solvent screening yielded 2-undecanone as biocompatible, extraction-efficient solvent with good phase separation. In aqueous-organic two-phase cultivations, this solvent extraction circumvents product instability in the aqueous cultivation medium, and it increases yields by reducing inhibitory effects. Complete de novo synthesis from glucose of all (hydroxy)benzoate-derived polyketides was achieved in two-phase cultivations with metabolically engineered strains. Additionally, mutasynthesis was applied to obtain fluorinated benzophenone derivatives.

The liquid-liquid extractive fermentation of L-lactic acid in a novel semi-partition bioreactor (SPB).

Fermentation technology is commonly used as a mature process to produce numerous products with the help of micro-organisms. However, these organisms are sometimes inhibited by the accumulation of these products or their by-products. One route to circumvent this is via extractive fermentation, which combines the fermentation process with extraction. To facilitate this, novel bioreactor designs are required, such as the semi-partition bioreactor (SPB) which has been recently proposed for in-situ extractive fermentation. The latter combines a fermentation and an extraction unit into a single vessel using a mixer-settler principle. Where the bioproduct is produced in the mixer and removed continuous in the settler. As the SPB functionality is a subject of interest, this study builds on demonstrating different process conditions in the production of a sample bioprocess (lactic acid (LA)) which is susceptible to product inhibition. The results showed a 34.5 g/L LA concentration was obtained in the pH-controlled condition. While LA production can suffer from product inhibition, neutralizing agents can be easily used to curb inhibitory problems, however, the LA fermentation is a simple (and well-studied) example, which can demonstrate an alternative route to avoiding product inhibition (for systems which cannot be rescued using pH control). Hence, to replicate a scenario of product inhibition, two different process conditions were investigated, no pH control with no extraction (non-integrated), and no pH control with integrated extractive fermentation. Key findings showed higher LA concentration in integrated (25.10 g/L) as compared to the non-integrated (14.94 g/L) case with improved yield (0.75 gg-1 (integrated) versus 0.60 gg-1 (non-integrated)) and overall productivity (0.35 gL-1h-1(integrated) versus 0.20 gL-1h-1(non-integrated)) likewise. This is the first demonstration of an SP bioreactor, and shows how the reactor can be applied to improve productivity. Based on these results, the SPB design can be applied to produce any product liable to product inhibition.

Ionic liquid-based in situ product removal design exemplified for an acetone-butanol-ethanol fermentation.

Selecting an appropriate separation technique is essential for the application of in situ product removal (ISPR) technology in biological processes. In this work, a three-stage systematic design method is proposed as a guide to integrate ionic liquid (IL)-based separation techniques into ISPR. This design method combines the selection of a suitable ISPR processing scheme, the optimal design of an IL-based liquid-liquid extraction (LLE) system followed by process simulation and evaluation. As a proof of concept, results for a conventional acetone-butanol-ethanol fermentation are presented (40,000 ton/year butanol production). In this application, ILs tetradecyl(trihexyl)phosphonium tetracyanoborate ([TDPh][TCB]) and tetraoctylammonium 2-methyl-1-naphthoate ([TOA] [MNaph]) are identified as the optimal solvents from computer-aided IL design (CAILD) method and reported experimental data, respectively. The dynamic simulation results for the fermentation process show that, the productivity of IL-based in situ (fed-batch) process and in situ (batch) process is around 2.7 and 1.8fold that of base case. Additionally, the IL-based in situ (fed-batch) process and in situ (batch) process also have significant energy savings (79.6% and 77.6%) when compared to the base case.

Comprehensive investigations of 2-phenylethanol production by high 2-phenylethanol tolerating Meyerozyma sp. strain YLG18.

2-Phenylethanol (2-PE) production through bio-synthesis method has become an appealing option owning to the mild conditions and high product selectivity. However, 2-PE is toxic to cells, which is an important limiting factor for the biosynthesis of 2-PE. In this study, a novel 2-PE generating Meyerozyma sp. strain YLG18 was first isolated, which could produce 2-PE through both Ehrlich and Shikimate pathways. Moreover, the indigenous high 2-PE tolerance makes it a promising candidate for high 2-PE production. Response surface methodology and in situ product recovery technology could improve the final 2-PE production to 3.20 g/L, representing the highest 2-PE production by using Meyerozyma sp. Furthermore, genes involved in 2-PE synthesis were identified and their expression levels between Shikimate pathway and Ehrlich pathway were compared. Based on the genomic and transcriptional analysis, a penta-functional enzyme AroM and an aspartate aminotransferase (AAT) with the potential to convert phenylalanine into phenylpyruvate were identified. These findings would help broaden our knowledge and add the pool of known 2-PE generating microbes and genes.

Gold nanoparticle-enhanced multiplexed imaging surface plasmon resonance (iSPR) detection of Fusarium mycotoxins in wheat.

A rapid, sensitive and multiplexed imaging surface plasmon resonance (iSPR) biosensor assay was developed and validated for three Fusarium toxins, deoxynivalenol (DON), zearalenone (ZEA) and T-2 toxin. The iSPR assay was based on a competitive inhibition format with secondary antibodies (Ab2) conjugated to gold nanoparticles (AuNPs) used as a signal amplification tag. Signal was amplified nearly 25-fold for DON, 90-fold for ZEA and 12-fold for T-2 toxin assay using Ab2-AuNPs. Analyses, including steps to regenerate the sensor, took 17.5min. The antigen coated sensor chip was used for more than 46 cycles without affecting signal intensity (< 12%). Matrix matched calibration curves were used to determine Fusarium toxins in wheat. The mean recoveries ranged from 87% to 103% with relative standard deviations of repeatability of less than 5%. The limits of detection were 15µg/kg for DON, 24µg/kg for ZEA and 12µg/kg for T-2 toxin. This provided sufficient sensitivity to monitor contamination of these mycotoxins in wheat in accordance with European Commission (EC) limits. Cut off levels for all three Fusarium toxins were validated using blank wheat and wheat spiked either at the EC regulated levels (100µg/kg for ZEA and T-2 toxin) or at one third of the EC level (for DON: 400µg/kg). The assay was successfully applied and further validated with naturally contaminated wheat samples. This is the first reported AuNP enhanced iSPR assay to detect and classify three agriculturally important Fusarium toxins in wheat.

An Imaging Surface Plasmon Resonance Biosensor Assay for the Detection of T-2 Toxin and Masked T-2 Toxin-3-Glucoside in Wheat.

A sensitive, rapid, and reproducible imaging surface plasmon resonance (iSPR) biosensor assay was developed to detect T-2 toxin and T-2 toxin-3-glucoside (T2-G) in wheat. In this competitive assay, an amplification strategy was used after conjugating a secondary antibody (Ab2) with gold nanoparticles. Wheat samples were extracted with a methanol/water mixture (80:20 v/v), then diluted with an equal volume of primary antibody (Ab1) for analysis. Matrix-matched calibration curves were prepared to determine T-2 toxin and T2-G. Recovery studies were conducted at three spiking levels in blank wheat. Mean recoveries ranged from 86 to 90%, with relative standard deviations for repeatability (RSDr) of less than 6%. Limits of detection were 1.2 ng/mL of T-2 toxin and 0.9 ng/mL of T2-G, equivalent to their levels in wheat, of 48 and 36 µg/kg, respectively. The developed iSPR assay was rapid and provided enough sensitivity for the monitoring of T-2 toxin/T2-G in wheat. This is the first iSPR assay useful for detecting the "masked" T2-G in wheat.

Integrated foam fractionation for heterologous rhamnolipid production with recombinant Pseudomonas putida in a bioreactor.

Heterologeous production of rhamnolipids in Pseudomonas putida is characterized by advantages of a non-pathogenic host and avoidance of the native quorum sensing regulation in Pseudomonas aeruginosa. Yet, downstream processing is a major problem in rhamnolipid production and increases in complexity at low rhamnolipid titers and when using chemical foam control. This leaves the necessity of a simple concentrating and purification method. Foam fractionation is an elegant method for in situ product removal when producing microbial surfactants. However, up to now in situ foam fractionation is nearly exclusively reported for the production of surfactin with Bacillus subtilis. So far no cultivation integrated foam fractionation process for rhamnolipid production has been reported. This is probably due to excessive bacterial foam enrichment in that system. In this article a simple integrated foam fractionation process is reported for heterologous rhamnolipid production in a bioreactor with easily manageable bacterial foam enrichments. Rhamnolipids were highly concentrated in the foam during the cultivation process with enrichment factors up to 200. The described process was evaluated at different pH, media compositions and temperatures. Foam fractionation processes were characterized by calculating procedural parameter including rhamnolipid and bacterial enrichment, rhamnolipid recovery, YX/S, YP/X, and specific as well as volumetric productivities. Comparing foam fractionation parameters of the rhamnolipid process with the surfactin process a high effectiveness of the integrated foam fractionation for rhamnolipid production was demonstrated.

Debottlenecking product inhibition in 1,3-propanediol fermentation by In-Situ Product Recovery.

The present work describes the application of liquid-liquid extraction as an In-Situ product recovery (ISPR) technique to overcome the problem of product inhibition in 1,3-PD fermentation. As a part of initial screening experiments, six solvents were subjected to phase separation and biocompatibility tests to find the best extractant for in-situ removal of 1,3-PD from the bioreactor. These included tributylphosphate, ethyl acetate, butyl acetate, oleyl alcohol, oleic acid and hexanol. Of these, ethyl acetate was found to be the most suitable solvent for 1,3-PD extraction. Use of the selected extractant in continuous integrated fermentation-extraction was established by batch and fed-batch extractive fermentations which demonstrated a significantly improved 1,3-PD production of 35g/L and 74.5g/L, respectively. A steady state 1,3-PD concentration of 58g/L was obtained in continuous extractive system. Continuous cultivation with in-situ cell retention and in-situ 1,3-PD removal demonstrated a 5-fold enhancement in 1,3-PD productivity over non-extractive batch.

Advances in in-situ product recovery (ISPR) in whole cell biotechnology during the last decade.

The review presents the state-of-the-art in the applications of in-situ product recovery (ISPR) in whole-cell biotechnology over the last 10years. It summarizes various ISPR-integrated fermentation processes for the production of a wide spectrum of bio-based products. A critical assessment of the performance of various ISPR concepts with respect to the degree of product enrichment, improved productivity, reduced process flows and increased yields is provided. Requirements to allow a successful industrial implementation of ISPR are also discussed. Finally, supporting technologies such as online monitoring, mathematical modeling and use of recombinant microorganisms with ISPR are presented.