Showing papers on "Bioprocess published in 2008"

Bioprocess optimization using design-of-experiments methodology.

Abstract: This review surveys recent applications of design-of-experiments (DoE) methodology in the development of biotechnological processes. Methods such as factorial design, response surface methodology, and (DoE) provide powerful and efficient ways to optimize cultivations and other unit operations and procedures using a reduced number of experiments. The multitude of interdependent parameters involved within a unit operation or between units in a bioprocess sequence may be substantially refined and improved by the use of such methods. Other bioprocess-related applications include strain screening evaluation and cultivation media balancing. In view of the emerging regulatory demands on pharmaceutical manufacturing processes, exemplified by the process analytical technology (PAT) initiative of the United States Food and Drug Administration, the use of experimental design approaches to improve process development for safer and more reproducible production is becoming increasingly important. Here, these options are highlighted and discussed with a few selected examples from antibiotic fermentation, expanded bed optimization, virus vector transfection of insect cell cultivation, feed profile adaptation, embryonic stem cell expansion protocols, and mammalian cell harvesting.

Micro biochemical engineering to accelerate the design of industrial-scale downstream processes for biopharmaceutical proteins

Abstract: The article examines how a small set of easily implemented micro biochemical engineering procedures combined with regime analysis and bioprocess models can be used to predict industrial scale performance of biopharmaceutical protein downstream processing. This approach has been worked on in many of our studies of individual operations over the last 10 years and allows preliminary evaluation to be conducted much earlier in the development pathway because of lower costs. It then permits the later large scale trials to be more highly focused. This means that the risk of delays during bioprocess development and of product launch are reduced. Here we draw the outcomes of this research together and illustrate its use in a set of typical operations; cell rupture, centrifugation, filtration, precipitation, expanded bed adsorption, chromatography and for common sources, E. coli, two yeasts and mammalian cells (GS-NSO). The general approach to establishing this method for other operations is summarized and new developments outlined. The technique is placed against the background of the scale-down methods that preceded it and complementary ones that are being examined in parallel. The article concludes with a discussion of the advantages and limitations of the micro biochemical engineering approach versus other methods.

Switching the mode of sucrose utilization by Saccharomyces cerevisiae.

Abstract: Background Overflow metabolism is an undesirable characteristic of aerobic cultures of Saccharomyces cerevisiae during biomass-directed processes. It results from elevated sugar consumption rates that cause a high substrate conversion to ethanol and other bi-products, severely affecting cell physiology, bioprocess performance, and biomass yields. Fed-batch culture, where sucrose consumption rates are controlled by the external addition of sugar aiming at its low concentrations in the fermentor, is the classical bioprocessing alternative to prevent sugar fermentation by yeasts. However, fed-batch fermentations present drawbacks that could be overcome by simpler batch cultures at relatively high (e.g. 20 g/L) initial sugar concentrations. In this study, a S. cerevisiae strain lacking invertase activity was engineered to transport sucrose into the cells through a low-affinity and low-capacity sucrose-H+ symport activity, and the growth kinetics and biomass yields on sucrose analyzed using simple batch cultures.

Microbioreactors for high-throughput cytotoxicity assays.

Abstract: Cell culture plays a fundamental role in the biotechnology and pharmaceutical industries, impacting both drug discovery and manufacturing as well as regenerative medicine. In drug discovery, cell-based assays are increasingly being used for drug target validation and drug ADMET (absorption, distribution, metabolism, elimination and toxicity) studies because cells can provide more representative responses to drugs than simple molecular assays and are easier to use in a high-throughput format than animals. There are, however, intrinsic drawbacks associated with conventional in vitro cellular tests using two-dimensional cultures, in that they lack a three-dimensional (3D) scaffold to support cell growth and proper tissue function, and cannot mimic in vivo cellular conditions. Tailoring scaffold properties for 3D cell cultures is therefore essential in developing a representative in vitro tissue model for cytotoxicity assays. Recently, microfluidic bioreactors with miniaturized culturing vessels and high controllability for operation and on-line monitoring/sensing have gained popularity in bioprocess development and cell-based assays. The advancement in this field has been enabled by the development of novel cell lines and reporter gene techniques, as well as new microfabrication, microfluidics and optical and electrochemical sensor technologies. Non-invasive detection methods using reporter genes and label-free techniques allow for real-time dynamic monitoring of viable cell number and cellular activities. Microbioreactors with continuous perfusion allow for long-term culturing to study chronic toxicity effects. Systemic toxicity and interactions between different cell types can also be studied on a biochip. High-density microfluidic arrays provide a platform for future high-throughput and high-content screening that will contribute to drug discovery and bioprocess development.

Optimizing Mixed-Culture Bioprocessing To Convert Wastes into Bioenergy

Abstract: The most efficient systems for biodegradation of polymeric organic compounds are mixed cultures that have evolved in some insect and mammalian guts. The efficiency and economic viability of converting organic wastes to biofuels depends on the characteristics of the waste material, especially the chemical composition and the concentrations of the components that can be converted into products that can be used as fuels. As mixed-culture fermentation involves large microbial communities, only certain compounds can be produced. Some products cannot be generated because they are converted to other compounds by the mixed culture more quickly than they are formed. When glucose-containing waste streams, such as those that are high in starch or cellulose, are used to produce bioenergy, butyrate may be one of the most important organic acid products. The hydrogen yield in mixed-culture bioprocessing can be increased by physically separating the anaerobic oxidation of sugars from hydrogen production by conducting the reactions in the anode and cathode, respectively, of a microbial fuel cell (MFC). Diverse microbial communities with metabolic flexibility should be more resistant to bacteriophage attack because different species or strains with similar metabolic functions can take over. Bioaugmentation can be used when modeling or systems biology analysis shows that a metabolic pathway that is needed to produce a useful energy carrier or its precursor is missing from the community metabolome.

Bioprocess Parameters and Oxygen Transfer Characteristics in β‐Lactamase Production by Bacillus Species

Abstract: After screening potential β-lactamase producers in a medium containing penicillin G, an inducible ( Bacillus subtilis NRS 1125) and a constitutive ( Bacillus licheniformis 749/C ATCC 25972) β-lactamase producer were selected. As the highest enzyme activity was obtained with B. licheniformis 749/C, the effects of the concentration of carbon sources, i.e., glucose, fructose, sucrose, citric acid, and glycerol, and nitrogen sources, i.e., (NH4)2HPO4, NH4Cl, yeast extract, casamino acids and peptone, pH, and temperature on β-lactamase production were investigated with B. licheniformis 749/C in laboratory scale bioreactors. Among the investigated media, the highest volumetric activity was obtained as 270 U cm−3 in the medium containing 10.0 kg m−3 glucose, 1.18 kg m−3 (NH4)2HPO4, 8.0 kg m−3 yeast extract, and the salt solution at 32 °C and pH0 = 6.0. By using the designed medium, fermentation and oxygen transfer characteristics of the bioprocess were investigated at V = 3.0 dm3 bioreactor systems with a VR = 1.65 dm3 working volume at QO/ VR = 0.5 vvm and N = 500 min-1. At the beginning of the process the Damkohler number was <1, indicating that the process was at biochemical reaction limited condition; at t = 2–5 h both mass-transfer and biochemical reaction resistances were effective; and at t = 6–10 h ( Da ≫1) the bioprocess was at mass transfer limited condition. Overall oxygen transfer coefficients ( KLa) varied between 0.01 and 0.03 s−1, enhancement factor ( KLa/ KLaO) varied between 1.2 and 2.3, and volumetric oxygen uptake rate varied between 0.001 and 0.003 mol m−3 s−1 throughout the bioprocess. The specific oxygen uptake and the specific substrate consumption rates were the highest at t = 2 h and then decreased with the cultivation. The maximum yield of cells on substrate and the maximum yield of cells on oxygen values were obtained, respectively, as YX/S = 0.34 and YX/O = 1.40, at t = 5 h, whereas the highest yield of substrate on oxygen was obtained as YS/O = 6.94 at t = 3.5 h. The rate of oxygen consumption for maintenance and the rate of substrate consumption for maintenance values were found, respectively, as mO = 0.13 kg kg−1 h−1 and mS = 3.02 kg kg−1 h−1.

A gamma sterilizable rfid system that prevents unauthorized operation of associated disposable bioprocess components

Abstract: This invention provides a system and apparatus that is able to authenticate and prevent illegal manufacturing and unauthorized operation of disposable bioprocess components. This invention utilizes a ferro-electric random access memory (FRAM) chip to store error-correctable information on a RFID tag attached to the disposable bioprocess components, where the error-correctable information is written into the memory chip, so that the information can remain in the chip when the RFID tag and disposable bioprocess component is gamma-sterilized. Also, this invention includes a method for authenticating the disposable bioprocess component that reduces liability in that a counterfeit poor quality disposable component is not used on the hardware so the user will not file an unjustified complaint.

Applications of PAT‐Process Analytical Technology in Recombinant Protein Processes with Escherichia coli

Abstract: Monitoring of bioprocesses and thus observation and identification of such processes is one of the main aims of bioprocess engineering. It is of vital importance in bioprocess development to improve the overall productivity by avoiding unintentional limitations to ensure not only optimal process conditions but also the observation of established production processes. Furthermore, reproducibility needs to be improved and final product quality and quantity be guaranteed. Therefore, an advanced monitoring and control system has been developed, which is based on different in-line, on-line and at-line measurements for substrates and products. Observation of cell viability applying in-line radio frequency impedance measurement and on-line determination of intracellular recombinant target protein using the reporter protein T-Sapphire GFP based on in-line fluorescence measurement show the ability for the detection of critical process states. In this way, the possibility for the on-line recognition of optimal harvest times arises and disturbances in the scheduled process route can be perceived.

Expression of nucleic acid sequences for production of biofuels and other products in algae and cyanobacteria

Abstract: Various embodiments provide, for example, vectors, expression cassettes, and cells useful for transgenic expression of nucleic acid sequences. In various embodiments, vectors can contain plastid-based sequences of unicellular photosynthetic bioprocess organisms for the production of food- and feed-stuffs, oils, biofuels, pharmaceuticals or fine chemicals.

Development of a New Bioprocess for Production of 1,3-propanediol I.: Modeling of Glycerol Bioconversion to 1,3-propanediol with Klebsiella pneumoniae Enzymes

Abstract: Glycerol is a renewable resource for it is formed as a byproduct during biodiesel production. Because of its large volume production, it seems to be a good idea to develop a technology that converts this waste into products of high value, for example, to 1,3-propanediol (1,3-PD). We suggested an enzymatic bioconversion in a membrane reactor in which the NAD coenzyme can be regenerated, and three key enzymes are retained by a 10-kDa ultrafilter membrane. Unfortunately, some byproducts also formed during successful glycerol to 1,3-PD bioconversion runs, as we used crude enzyme solution of Klebsiella pneumoniae. To study the possibilities to avoid this byproduct formation, we built a mathematical description of this system. The model was also used for simulation bioconversions of high glycerol concentration with and without elimination of byproduct formation and of continuous operation.

Method for preventing an unauthorized use of disposable bioprocess components

Abstract: This invention provides a system and apparatus that is able to authenticate and prevent illegal manufacturing and unauthorized operation of disposable bioprocess components This invention utilizes a ferro-electric random access memory chip (FRAM) chip to store error-correctable information on a RFID tag attached to the disposable bioprocess components, where the error-correctable information is written in sequence into the memory chip, so that the redundant information can remain in the chip when the RFID tag and disposable bioprocess component is gamma-sterilized Also, this invention includes a method for authenticating the disposable bioprocess component that reduces liability in that a counterfeit poor quality disposable component is not used on the hardware so the user will not file an unjustified complaint

Sugar beet leaves as new source of hydroperoxide lyase in a bioprocess producing green-note aldehydes

Abstract: Hydroperoxide lyase activity was found in sugar beet leaves. Its optimum pH and temperature were, respectively, 6.7 and 22°C. Under these conditions, conversion of linolenic acid 13-hydroperoxide to cis-3-hexenal with a maximum yield of 80% was reached after only 2 min. The stability of cis-3-hexenal was improved by acidifying the reaction medium. Based on these studies, a bioprocess producing green-note aldehydes in a laboratory-scale was achieved.

At-line monitoring of bioreactor protein production by Surface Plasmon Resonance.

Abstract: An innovative and automated method for the at-line monitoring of secreted protein was developed by harnessing a Surface Plasmon Resonance-based biosensor to a bioreactor. The proof of concept was performed by following at-line the relative concentration of a secreted protein produced by transient transfection of mammalian cells in a bioreactor. Our results suggest that our approach can be readily applied to the at-line determination of both protein concentration and bioactivity. Our experimental setup and strategy can thus satisfy the needs related to the development of novel bioprocess control protocols in the context of the new process analytical technology that arises in the biopharmaceutical industry.

Adaptive control strategies for a class of anaerobic depollution bioprocesses

Abstract: This paper presents the design and the analysis of some nonlinear adaptive control strategies for a class of anaerobic depollution processes that are carried out in continuous stirred tank bioreactors. The controller design is based on the input-output linearization technique. The adaptive control structure is based on the nonlinear model of the process and is combined with a state observer and a parameter estimator which play the role of the software sensors for the on-line estimation of biological states and parameter variables of interest of the bioprocess. The resulted control methods are applied in depollution control problem in the case of the anaerobic digestion bioprocess for which dynamical kinetics are strongly nonlinear and not exactly known, and not all the state variables are measurable. The effectiveness and performance of both estimation and control algorithms are illustrated by simulation results.

REVIEW: BIOCATALYSTS AND BIOREACTOR DESIGN Bioprocess Optimization Using Design-of-Experiments Methodology

Abstract: This review surveys recent applications of design-of-experiments (DoE) methodology in the development of biotechnological processes. Methods such as factorial design, response surface methodology, and (DoE) provide powerful and efficient ways to optimize cultivations and other unit operations and procedures using a reduced number of experiments. The multitude of interdependent parameters involved within a unit operation or between units in a bioprocess sequence may be substantially refined and improved by the use of such methods. Other bioprocess-related applications include strain screening evaluation and cultivation media balancing. In view of the emerging regulatory demands on pharmaceutical manufacturing processes, exemplified by the process analytical technology (PAT) initiative of the United States Food and Drug Administration, the use of experimental design approaches to improve process development for safer and more reproducible production is becoming increasingly important. Here, these options are highlighted and discussed with a few selected examples from antibiotic fermentation, expanded bed optimization, virus vector transfection of insect cell cultivation, feed profile adaptation, embryonic stem cell expansion protocols, and mammalian cell harvesting.

Transgenic Plants Producing Polyhydroxyalkanoates

Abstract: Currently, the polyhydroxybutyrate (PHB) copolymer, polyhydroxy-co-valerate (PHBV) is being produced commer- cially using a two-stage glucose / propionate fed batch fermentation process using Ralstonia eutropha. The economics of the manufacturing process are still a major barrier to the widespread use of polyhydroxyalkanoates (PHAs) and intensive efforts are being made to reduce the cost of production by means of bioprocess design and metabolic engineering of production strains. However, despite these improvements, the production costs are still high compared to petroleum-derived commodity plastics. An alternative strategy for lowering production costs that has been proposed is to develop transgenic plants that produce PHAs. This strategy is considerably cheaper because the PHAs production from plants does not require expensive fermentation equip- ment and processing facilities.

Potential of on-line CIMS for bioprocess monitoring.

Abstract: Chemical-ionization mass spectrometry (CIMS) using flow reactors is an emerging method for on-line monitoring of trace concentrations of organic compounds in the gas phase. In this study, a flow-reactor CIMS instrument, employing the H 3 O + cation as the ionizing reagent, was used to simultaneously monitor several volatile metabolic products as they are released into the headspace during bacterial growth in a bioreactor. Production of acetaldehyde, ethanol, acetone, butanol, acetoin, diacetyl, and isoprene by Bacillus subtilis is reported. Ion signal intensities were related to solution-phase concentrations using empirical calibrations and, in the case of isoprene, were compared with simultaneous gas chromatography measurements. Identification of volatile and semivolatile metabolites is discussed. Flow-reactor CIMS techniques should be useful for bioprocess monitoring applications because of their ability to sensitively and simultaneously monitor many volatile metabolites on-line.

Extractables and Leachables : Study Approach for Disposable Materials Used in Bioprocessing

Abstract: To help maintain product safety, quality, and efficacy, a general approach to designing extractables and leachables studies should include evaluating and qualifying bioprocess materials used in a disposable manufacturing environment. This paper presents a systematic approach to the evaluation, identification, quantitation, and specification setting of extractables and leachables that builds product quality into the manufacturing process. It also discusses analytical thresholds of extractables for the evaluation of disposable materials.

Issues in Plant Cell Culture Engineering for Enhancement of Productivity

Abstract: Plant cell culture is seen as an alternative source to whole plants for the production of useful compounds, such as dyes, pharmaceuticals, perfumes and insecticides. Despite intensive research for the last 30 years, only a few products have reached commercial production in bioreactors, and the number is far less than those commercialized for bacterial and animal cells. Several biological challenges including slow cell growth, low productivity, compartmentation, metabolite channelling and poor metabolite secretion have hampered the realization of commercialization of plant cell products. Other engineering and technological problems include cell aggregation, plant cell shear sensitivity, foaming and lack of automation. Several strategies are being explored to improve productivity including medium optimization and cultural conditions, and the understanding of biochemical and signal transduction pathways. In plant cell culture process development, there is a great need to explore a more rational approach through metabolic and genetic engineering by making use of advanced technologies in genomic, transcriptomic, proteomic and metabolomic analyses. These should be integrated with developments in bioprocess and systems engineering.

Recombinant Protein Production Yields from Mammalian Cells : Past, Present, and Future

Abstract: in the last two decades, recombinant protein yields from mammalian cells in batch and extruded-batch bioprocesses have increased, dramatically. In general, this has been a result of the utilization of cell lines with high specific productivities, the formulation of media to allow the suspension cultivation of cells to high densities, a better understanding of bioprocess conditions, and the enhancement of cell viability in these high-density suspension cultures. This article discusses the contributions of these factors to yield improvement. It also describes a theoretical model to show how volumetric productivities may increase in the future. The major consequence of higher volumetric productivities is expected to he the decline in the time or volume of production runs. This outcome is expected to lead to a substantial improvement in the efficiency of protein manufacturing.

Process analysis of macrotetrolide biosynthesis during fermentation by means of direct infusion LC‐MS

Abstract: The optimization of the biosynthetic pathways is highly attractive for the large-scale preparation of macrotetrolides, because overall yields in the chemical synthesis of compounds like nonactin have been very low. A key success factor determining the outcome of such optimizations is the adequate process analysis for the envisioned product. The analytical methods for process control involved in the past spectrophotometric and chromatographic measurements. LC-MS offers a modern approach to obtain more detailed data than the spectrophotometric and chromatographic measurements used in the past. In this work, a fast and versatile analytical LC-MS method has been set up, which has proven of much value for the in-process analysis of macrotetrolides during fermentation and which has allowed rapid large-scale bioprocess development.

Ultrasonic monitor for a bioreactor

Abstract: A method and apparatus for monitoring the state of a microbiological or animal cell culture, or other bioprocess wherein the attenuation and in some embodiments, the speed of an ultrasonic wave passed through the culture is used to determine viscous and viscoelastic properties of the culture medium.

The Potential of Mid- and Near-infrared Spectroscopy for Reliable Monitoring of Bioprocesses

Abstract: The processes involved in bioreactors are complex and a detailed monitoring of the chemical composition of gaseous, liquid and solid matter involved in the bioprocess is of crucial importance for efficient operation. However, only a few parameters can be reliably monitored in real time with commercially available instrumentation, but there is a growing demand and need for effective and efficient process monitoring tools. Infrared (IR) spectroscopy has proved to be a powerful method for the study of various biomedical and bioanalytical applications. In this paper, applications of this analytical method for bioprocess monitoring have been discussed by reviewing the relevant literature from the last ten years. Furthermore, recent advances in our laboratory in the field of bio-reactor monitoring by use of mid-IR attenuated total reflection (ATR) spectroscopy and the development of mid-IR sensor systems for continuous biofluid monitoring based on transmission spectroscopy have also been addressed.

Fuzzy-Neural control of a distributed parameter bioprocess plant

Abstract: In the paper it is proposed a new recurrent fuzzy-neural multi-model (FNMM) identifier applied for decentralized identification of a distributed parameter anaerobic wastewater treatment digestion bioprocess, carried out in a fixed bed and a recirculation tank. The distributed parameter analytical model of the digestion bioprocess is reduced to a lumped system using the orthogonal collocation method, applied in three collocation points (plus the recirculation tank), which are used as centers of the membership functions of the fuzzyfied space variable of the plant. The states of the proposed FNMM identifier are implemented by a direct feedback-feedforward hierarchical fuzzy-neural controller. The comparative graphical simulation results of the digestion wastewater treatment system identification and control, obtained via learning, exhibited a good convergence, and precise reference tracking outperforming the optimal control.