ROLE OF BIOREACTOR IN BIOPROCESS ENGG.-01

What Is Bioreactor?
A bioreactor is a vessel in which is carried out a chemical process which involves organisms or biochemically active substances derived from such organisms.
Bioreactors are commonly cylindrical, ranging in size from some liter to cube meters,and are often made of stainless steel.
Bioreactor design is quite a complex engineering task. Under optimum conditions the microorganisms or cells will reproduce at an astounding rate. The vessel's environmental conditions like gas (i.e., air, oxygen, nitrogen, carbon dioxide) flowrates, temperature, pH and dissolved oxygen levels, and agitation speed need to be closely monitored and controlled. One bioreactor manufacturer, Broadley-James Corporation, uses vessels, sensors, controllers, and a control system, digitally networked together for their bioreactor system.
Continuous flow stirred tank reactors (chemostat) In the continuous flow, stirred tank reactor (CSTR or chemostat) fresh medium is fed into the bioreactor at a constant rate, and medium mixed with cells leaves the bioreactor at the same rate. A fixed bioreactor volume is maintained and ideally, the effluent stream should have the same composition as the bioreactor contents. The culture is fed with fresh medium containing one and sometimes two growth-limiting nutrients such as glucose. The concentration of the cells in the bioreactor is controlled by the concentration of the growth-limiting nutrient. A steady state cell concentration is reached where the cell density and substrate concentration are constant. The cell growth rate (µ) is controlled by the dilution rate (D) of growth limiting nutrient.
Cell culture bioreactors are categorized into two types: 1. Those that are used for cultivation of anchorage dependent cells (e.g. primary cultures derived from normal tissues and diploid cell lines. 2. Those that are used for the cultivation of suspended mammalian cells (e.g. cell lines derived from cancerous tissues and tumors, transformed diploid cell lines, hybridomas). In some cases the bioreactor may be modified to grow both anchorage dependent and suspended cells. Ideally any cell culture bioreactor must maintain a sterile culture of cells in medium conditions which maximize cell growth and productivity.
Fouling can harm the overall sterility and efficiency of the bioreactor, especially the heat exchangers. To avoid it the bioreactor must be easily cleanable and must be as smooth as possible (therefore the round shape).
Heat exchange is needed to maintain the bioprocess at a constant temperature. Biological fermentation is a major source of heat, therefore in most cases bioreactors need water refrigeration. They can be refrigerated with an external jacket or, for very large vessels, with internal coils.
Optimal oxygen transfer is perhaps the most difficult task to accomplish. Oxygen is poorly soluble in water -and even less in fermentation broths- and is relatively scarce in air (20.8%). Oxygen transfer is usually helped by agitation, that is also needed to mix nutrients and to keep the fermentation homogeneous. There are however limits to the speed of agitation, due both to high power consumption (that's proportional to the cube of the speed) and the damage to organisms due to excessive tip speed.
Bioreactor treatment may be performed using microorganisms growing in suspension in the fluid or attached on a solid growth support medium. In suspended growth systems, such as fluidized beds or sequencing batch reactors, contaminated groundwater is circulated in an aeration basin where a microbial population aerobically degrades organic matter and produces carbon dioxide, water, and biomass. The biomass is settled out in a clarifier, then either recycled back to the aeration basin or disposed of as sludge. In attached growth systems, such as upflow fixed film bioreactors, rotating biological contactors (RBCs), and trickling filters, microorganisms are grown as a biofilm on a solid growth support matrix and water contaminants are degraded as they diffuse into the biofilm. Support media include solids that have a large surface area for bacterial attachment.
A bioreactor landfill operates to rapidly transform and degrade organic waste. The increase in waste degradation and stabilization is accomplished through the addition of liquid and air to enhance microbial processes. This bioreactor concept differs from the traditional “dry tomb” municipal landfill approach.
A bioreactor landfill is not just a single design and will correspond to the operational process invoked. There are three different general types of bioreactor landfill configurations:
Aerobic - In an aerobic bioreactor landfill, leachate is removed from the bottom layer, piped to liquids storage tanks, and re-circulated into the landfill in a controlled manner. Air is injected into the waste mass, using vertical or horizontal wells, to promote aerobic activity and accelerate waste stabilization. Anaerobic - In an anaerobic bioreactor landfill, moisture is added to the waste mass in the form of re-circulated leachate and other sources to obtain optimal moisture levels. Biodegradation occurs in the absence of oxygen (anaerobically) and produces landfill gas. Landfill gas, primarily methane, can be captured to minimize greenhouse gas emissions and for energy projects. Hybrid (Aerobic-Anaerobic) - The hybrid bioreactor landfill accelerates waste degradation by employing a sequential aerobic-anaerobic treatment to rapidly degrade organics in the upper sections of the landfill and collect gas from lower sections. Operation as a hybrid results in the earlier onset of methanogenesis compared to aerobic landfills The Solid Waste Association of North America (SWANA) has defined a bioreactor landfill as "any permitted Subtitle D landfill or landfill cell where liquid or air is injected in a controlled fashion into the waste mass in order to accelerate or enhance biostabilization of the waste." The United States Environmental Protection Agency (EPA) is currently collecting information on the advantages and disadvantages of bioreactor landfills through case studies of existing landfills and additional data so that EPA can identify specific bioreactor standards or recommend operating parameters.
Features Unique to Bioreactor Landfills: The bioreactor accelerates the decomposition and stabilization of waste. At a minimum, leachate is injected into the bioreactor to stimulate the natural biodegradation process. Bioreactors often need other liquids such as stormwater, wastewater, and wastewater treatment plant sludges to supplement leachate to enhance the microbiological process by purposeful control of the moisture content and differs from a landfill that simple recirculates leachate for liquids management. Landfills that simply recirculate leachate may not necessarily operate as optimized bioreactors. J Biomech, 2005 Mar, 38(3), 543 - 93-D computational modeling of media flow through scaffolds in a perfusion bioreactor; Porter B et al.; Media perfusion bioreactor systems have been developed to improve mass transport throughout three-dimensional (3-D) tissue-engineered constructs cultured in vitro . In addition to enhancing the exchange of nutrients and wastes, these systems simultaneously deliver flow-mediated shear stresses to cells seeded within the constructs . Local shear stresses are a function of media flow rate and dynamic viscosity, bioreactor configuration, and porous scaffold microarchitecture . We have used the Lattice-Boltzmann method to simulate the flow conditions within perfused cell-seeded cylindrical scaffolds . Microcomputed tomography imaging was used to define the scaffold microarchitecture for the simulations, which produce a 3-D fluid velocity field throughout the scaffold porosity . Shear stresses were estimated at various media flow rates by multiplying the symmetric part of the gradient of the velocity field by the dynamic viscosity of the cell culture media . The shear stress algorithm was validated by modeling flow between infinite parallel plates and comparing the calculated shear stress distribution to the analytical solution . Relating the simulation results to perfusion experiments, an average surface shear stress of 5x10(-5)Pa was found to correspond to increased cell proliferation, while higher shear stresses were associated with upregulation of bone marker genes . This modeling approach can be used to compare results obtained for different perfusion bioreactor systems or different scaffold microarchitectures and may allow specific shear stresses to be determined that optimize the amount, type, or distribution of in vitro tissue growth.J Biotechnol, 2005 Mar 2, 116(1), 61 - 77 Epub 2004 Nov 30.Pellet morphology, culture rheology and lovastatin production in cultures of Aspergillus terreus; Casas Lopez JL et al.; Pellet growth of Aspergillus terreus ATCC 20542 in submerged batch fermentations in stirred bioreactors was used to examine the effects of agitation (impeller tip speed u(t) of 1.01-2.71 ms(-1)) and aeration regimens (air or an oxygen-enriched mixture containing 80% oxygen and 20% nitrogen by volume) on the fungal pellet morphology, broth rheology and lovastatin production . The agitation speed and aeration methods used did not affect the biomass production profiles, but significantly influenced pellet morphology, broth rheology and the lovastatin titers . Pellets of approximately 1200mum initial diameter were reduced to a final stable size of approximately 900mum when the agitation intensity was >/=600rpm (u(t)>/=2.03ms(-1)) . A stable pellet diameter of approximately 2500mum could be attained in less intensely agitated cultures . These large fluffy pellets produced high lovastatin titers when aerated with oxygen-enriched gas but not with air . Much smaller pellets obtained under highly agitated conditions did not attain high lovastatin productivity even in an oxygen-enriched atmosphere . This suggests that both an upper limit on agitation intensity and a high level of dissolved oxygen are essential for attaining high titers of lovastatin . Pellet size in the bioreactor correlated equally well with the specific energy dissipation rate and the energy dissipation circulation function . The latter took into account the frequency of passage of the pellets through the high shear regions of the impellers . Pellets that gave high lovastatin titers produced highly shear thinning cultivation broths.Appl Microbiol Biotechnol . 2005 Jan 13; {Epub ahead of print}Oligomeric compounds formed from 2,5-xylidine (2,5-dimethylaniline) are potent enhancers of laccase production in Trametes versicolor ATCC 32745; Kollmann A et al.; Numerous chemicals, including the xenobiotic 2,5-xylidine, are known to induce laccase production in fungi . The present study was conducted to determine whether the metabolites formed from 2,5-xylidine by fungi could enhance laccase activity . We used purified laccases to transform the chemical and then we separated the metabolites, identified their chemical structure and assayed their effect on enzyme activity in liquid cultures of Trametes . versicolor . We identified 13 oligomers formed from 2,5-xylidine . (4E)-4-(2,5-dimethylphenylimino)-2,5-dimethylcyclohexa-2,5-dienone at 1.25x10(-5) M was an efficient inducer, resulting in a nine-fold increase of laccase activity after 3 days of culture . Easily synthesized in one step (67% yield), this compound could be used in fungal bioreactors to obtain a great amount of laccases for biochemical or biotechnological purposes, with a low amount of inducer.Ann N Y Acad Sci, 2004 Nov, 1027, 85 - 98Modeling of phosphate ion transfer to the surface of osteoblasts under normal gravity and simulated microgravity conditions; Mukundakrishnan K et al.; We have modeled the transport and accumulation of phosphate ions at the remodeling site of a trabecular bone consisting of osteoclasts and osteoblasts situated adjacent to each other in straining flows . Two such flows are considered; one corresponds to shear levels representative of trabecular bone conditions at normal gravity, the other corresponds to shear level that is representative of microgravity conditions . The latter is evaluated indirectly using a simulated microgravity environment prevailing in a rotating wall vessel bioreactor (RWV) designed by NASA . By solving the hydrodynamic equations governing the particle motion in a RWV using a direct numerical simulation (DNS) technique, the shear stress values on the surface of the microcarriers are found . In our present species transfer model, osteoclasts release phosphate ions (Pi) among other ions at bone resorption sites . Some of the ions so released are absorbed by the osteoblast, some accumulate at the osteoblast surface, and the remainder are advected away . The consumption of Pi by osteoblasts is assumed to follow Michaelis-Menten (MM) kinetics aided by a NaPi cotransporter system . MM kinetics views the NaPi cotransporter as a system for transporting extracellular Pi into the osteoblast . Our results show, for the conditions investigated here, the net accumulation of phosphate ions at the osteoblast surface under simulated microgravity conditions is higher by as much as a factor of three . Such increased accumulation may lead to enhanced apoptosis and may help explain the increased bone loss observed under microgravity conditions.Artif Organs, 2005 Jan, 29(1), 58 - 66A novel bioartificial liver with culture of porcine hepatocyte aggregates under simulated microgravity; Hochleitner B et al.; An extracorporeal bioartificial liver device could provide vital support to patients suffering from acute liver failure . We designed a novel, customized bioreactor for use as a bioartificial liver (patent pending) . The Innsbruck Bioartificial Liver (IBAL) contains aggregates of porcine hepatocytes grown under simulated microgravity . The culture vessel rotates around its longitudinal axis and is perfused by two independent circuits . The circuit responsible for exchange of plasma components with the patient consists of a dialysis tube winding spirally around the internal wall of the culture vessel . IBAL was evaluated in vitro . Viability tests showed sufficient viability of hepatocytes for up to 10 days . Cytologic examination of samples from the bioreactor showed liver cell aggregates . These were also examined by electron microscopy . A number of biochemical parameters were analyzed . In conclusion, cell culture is possible for at least 10 days in the IBAL system, organoid hepatocyte aggregates are formed and synthetic activity of the hepatocytes was demonstrated.Biotechnol Bioeng . 2005 Jan 10; {Epub ahead of print}Novel application of oxygen-transferring membranes to improve anaerobic wastewater treatment; Kappell AS et al.; Anaerobic biological wastewater treatment has numerous advantages over conventional aerobic processes; anaerobic biotechnologies, however, still have a reputation for low-quality effluents and operational instabilities . In this study, anaerobic bioreactors were augmented with an oxygen-transferring membrane to improve treatment performance . Two anaerobic bioreactors were fed a synthetic high-strength wastewater (chemical oxygen demand, or COD, of 11,000 mg l(-1)) and concurrently operated until biomass concentrations and effluent quality stabilized . Membrane aeration was then initiated in one of these bioreactors, leading to substantially improved COD removal efficiency (>95%) compared to the unaerated control bioreactor ( approximately 65%) . The membrane-augmented anaerobic bioreactor required substantially less base addition to maintain circumneutral pH and exhibited 75% lower volatile fatty acid concentrations compared to the unaerated control bioreactor . The membrane-aerated bioreactor, however, failed to improve nitrogenous removal efficiency and produced 80% less biogas than the control bioreactor . A third membrane-augmented anaerobic bioreactor was operated to investigate the impact of start-up procedure on nitrogenous pollutant removal . In this bioreactor, excellent COD (>90%) and nitrogenous (>95%) pollutant removal efficiencies were observed at an intermediate COD concentration (5,500 mg l(-1)) . Once the organic content of the influent wastewater was increased to full strength (COD = 11,000 mg l(-1)), however, nitrogenous pollutant removal stopped . This research demonstrates that partial aeration of anaerobic bioreactors using oxygen-transferring membranes is a novel approach to improve treatment performance . Additional research, however, is needed to optimize membrane surface area versus the organic loading rate to achieve the desired effluent quality . (c) 2005 Wiley Periodicals, Inc.Yi Chuan, 2004 Nov, 26(6), 977 - 83{The application of chloroplast genome transformation in higher plant.}; Wang YF et al.; The chloroplast genome transformation of higher plant has become the research hotspot of plant genetic engineering with several advantages over nuclear genetic engineering, and it has become into a powerful approach for both basic and applied research . This paper presents a brief summary for the principle and methods of chloroplast genome transformation in higher plant . The particular emphasis was placed on its application in the basic and applied research . These applications include the studies of rearrangement of Rubisco and the structure, transcription, translation and RNA editing of genes in chloroplast; the producing of antibodies, vaccines, poly-beta-hydroxybutyrate and bio-elastic protein using chloroplast as bioreactor; the generating of transgenic plants with resistance to insect, disease, herbicide and drought; and the decreasing the gene flow of transgenic plants.Yi Chuan, 2004 Sep, 26(5), 567 - 73{Mammary Gland Dual-Expression Construct Containing Prolactin and Foreign DNAs Can Promote the Expression of Foreign Proteins in Mammary Cells.}; Liu FT et al.; To investigate the feasibility in the promotion of the expression of foreign proteins, such as human lactinferrin (hLF) and thrombopoietin (TPO), in the transgenic animals-mammary gland bioreactor by bovine prolactin (bPRL), two types of mammary gland dual-expression vectors containing bPRL and foreign DNAs were constructed . In one type of vector, both foreign and bPRL DNAs were linked by internal ribosome entry site (IRES) which shared one goat 6.7 kb of beta-casein promoter; while in the other type of vector, the transcriptions of the foreign gene aswell as bPRL were respectively directed by 6.7 kb goat beta-casein promoter and 2.0 kb of goat beta-casein promoter . After transfection of the vectors with lipofectin method, the expression of foreign proteins were measured by RT-PCR as well as ELISA assay . The results showed that bPRL could obviously promote the expression of foreign proteins (hLF and TPO) in cultured cells . The hLF expression level was increased from 12.6 mug/L to 18.4 mug/L and 37.2 mug/L in the COS-7 cells (African green monkey kidney cell) (P<0.05), and from 13.7 mug/L to 20.7 mug/L and 19.9 mug/L in the HC-11 cells (epithelial cell of mouse mammary gland) (P<0.05) by two vectors, respectively . TPO expression level was increased from 572 ng/L to 1340 ng/L in the COS-7 cell (P<0.05), and from 783 ng/L to 1040 ng/L in the HC-11 cell (P<0.05) by the vector which have two promoters . This work indicate that bovine prolactin can effectively increase the expression of foreign genes in mammal cells.Yi Chuan, 2004 Jul, 26(4), 537 - 43{Achievements and Applications in Making Chicken Chimeras Using BCs.}; Yan HF et al.; The technology of producing chicken chimeras using blastodermal cells is very important not only in the field of transgenic chicken bioreactor but also in searching for efficient ways to conserve avian genetic resource . The basic processes for producing chicken chimeras consist of: (1) Setting up the color model; (2) Separating and dissociating of donor embryos; (3) Compromising of the recipient embryos; (4) Windowing and recovering the recipient eggs; (5) Cells injecting; (6) Method of hatching . The progress, obstacles and prospects of producing chicken chimeras via BCs were discussed in this paper.Yi Chuan, 2003 Jan, 25(1), 107 - 12{Plants as bioreactor for the production of pharmaceutical proteins.}; Xiao NZ et al.; Transgenic plant as bioreactor has been used to produce recombinant proteins for medicinal purposes,including mammalian antibodies,blood substitutes and vaccines.As the demand for biopharmaceuticals is expected to increase,transgenic plants have the potential to provide virtually unlimited quantities of proteins for use as tools in both human health care and the bioscience.This paper reviews the recent developments in this field and the prospect of commercial applications.J Biotechnol, 2005 Feb 23, 115(4), 379 - 386 Epub 2004 Nov 21.Selection of new over-producing derivatives for the improvement of extracellular lipase production by the non-conventional yeast Yarrowia lipolytica; Fickers P et al.; The non-conventional yeast Yarrowia lipolytica produces an extracellular lipase encoded by the LIP2 gene . Mutant strains with enhanced productivity were previously obtained either by chemical mutagenesis or genetic engineering . In this work, we used one of these mutants, named LgX64.81 to select new overproducing strains following by amplification of the LIP2 gene . We also developed a process for lipase production in bioreactors and compared lipase production levels in batch and fed-batch cultures . Batch culture led to a lipase production of 26450Uml(-1) in a media containing olive oil and tryptone as carbon and nitrogen sources . Feeding of a combination of tryptone and olive oil at the end of the exponential growth phase yielded to lipase activity of 158246Uml(-1) after 80h of cultivation . In addition this production system developed for the extracellular lipase could also be applied for other heterologous protein production since we have demonstrated that LgX64.81 is an interesting alternative host strain.J Biotechnol, 2005 Feb 23, 115(4), 333 - 43 Epub 2004 Nov 24.Comparison of different fungal enzymes for bleaching high-quality paper pulps; Sigoillot C et al.; Wild and recombinant hydrolases and oxidoreductases with a potential interest for environmentally sound bleaching of high-quality paper pulp (from flax) were incorporated into a totally chlorine free (TCF) sequence that also included a peroxide stage . The ability of feruloyl esterase (from Aspergillus niger) and Mn(2+)-oxidizing peroxidases (from Phanerochaete chrysosporium and Pleurotus eryngii) to decrease the final lignin content of flax pulp was shown . Laccase from Pycnoporus cinnabarinus (without mediator) also caused a slight improvement of pulp brightness that was increased in the presence of aryl-alcohol oxidase . However, the best results were obtained when the laccase treatment was performed in the presence of a mediator, 1-hydroxybenzotriazol (HBT), enabling strong delignification of pulps . The enzymatic removal of lignin resulted in high-final brightness values that are difficult to attain by chemical bleaching of this type of pulp . A partial inactivation of laccase by HBT was observed but this negative effect was strongly reduced in the presence of pulp . The good results obtained with the same laccase expressed in A . niger at bioreactor scale, revealed the feasibility of using recombinant laccase for bleaching high-quality non-wood pulps in the presence of a mediator.J Biotechnol, 2005 Feb 9, 115(3), 307 - 15Efficient production of a soluble fusion protein containing human beta-defensin-2 in E . coli cell-free system; Chen H et al.; Human beta-defensin-2 (hBD2), a small cationic peptide, exhibits a broad range of antimicrobial activity and does not cause microbial resistance . In order to produce hBD2 efficiently, an Escherichia coli cell-free biosynthesis system has been developed as an alternative method . A specific plasmid pIVEX2.4c-trxA-shBD2 was constructed for the cell-free expression of fusion protein (hBD2 linked with His-Tag and Trx-Tag) . This allowed enhancement of protein stability and facilitated downstream purification process . Significant amount of target fusion protein was synthesized in the batch-mode bioreactor by optimizing the reaction conditions . About five-fold improvement of productivity (ca . 2.0mg/ml soluble fusion protein) could be achieved by using a continuous exchange cell-free (CECF) system compared to batch system . One-step affinity chromatographic process was developed to recover high purity fusion protein (95.2%) with overall recovery ratio of about 50% . The fusion protein was cleaved by cyanogens bromide (CNBr), and the mature hBD2 had demonstrated strong inhibition on the growth of E . coli D31 at low concentration.J Biotechnol, 2005 Feb 9, 115(3), 291 - 301 Epub 2004 Nov 05.The influence of the chemical composition of cell culture material on the growth and antibody production of hybridoma cells; Heilmann K et al.; The multiplication and antibody production of murine hybridoma cells cultured on five different polymer membranes were tested and compared with conventional tissue culture polystyrene (TCPS) . Membranes were prepared from polyacrylonitrile (PAN) and acrylonitrile copolymerized with N-vinylpyrrolidone (NVP20, NVP30), Na-methallylsulfonate (NaMAS) and N-(3-amino-propyl-methacrylamide-hydrochloride) (APMA) . Cell number and antibody concentration were quantified as criteria for viability and productivity . Adhesion of hybridoma cells was characterized by vital and scanning electron microscopy . The results suggest that a strong adhesion of cells, observed on APMA and TCPS, increased cell growth but reduced monoclonal antibody production . In contrast membranes with lowered adhesivity such as NVP20 provided favourable conditions for monoclonal antibody production . In addition it was shown that this membrane also possessed a minor fouling as indicated by the low decrease of water flux across the membrane after protein adsorption . It was concluded that NVP20 could be a suitable material for the development of hollow fibre membranes for bioreactors.Protein Pept Lett, 2005 Jan, 12(1), 85 - 8Production of minichaperone (sht GroEL191-345) and its function in the refolding of recombinant human interferon gamma; Guan YX et al.; The recombinant minichaperone sht GroEL191-345 was cultivated in a 3.7 L stirred bioreactor with the high yield of 216.2 mg/L broth . In the refolding of recombinant human interferon gamma (rhuIFN-gamma) inclusion bodies, more than 2-3 fold enhancement in protein mass recovery and total activity were observed in the presence of free or immobilized minichaperone to the refolding buffer.Res Microbiol, 2005 Jan-Feb, 156(1), 82 - 7Optimisation of growth conditions for continuous culture of the hyperthermophilic archaeon Thermococcus hydrothermalis and development of sulphur-free defined and minimal media; Postec A et al.; The hyperthermophilic archaeon Thermococcus hydrothermalis was cultivated in continuous culture in a gas-lift bioreactor in the absence of elemental sulphur on both proteinaceous and maltose-containing media . Optimal conditions (pH, temperature and gas flow rate), determined on complex media that yielded maximal growth rate and maximal steady state cell density, were obtained at 80 degrees C, pH 6 and gas sparging at 0.2 v v(-1) min(-1) . Higher steady state cell densities were obtained on a medium containing maltose and yeast extract . In order to design a defined and minimal media, the nutritional requirements of T . hydrothermalis were then investigated using continuous culture in the absence of elemental sulphur in the gas-lift bioreactor . First, the complex nutriments were replaced and a defined medium containing maltose, 19 amino acids and the two nitrogenous bases adenine and thymine, was determined . Secondly, selective feedings and withdrawal of amino acids showed requirements for 14 amino acids.Biotechnol Appl Biochem . 2005 Jan 6; {Epub ahead of print}Application of radial-flow bioreactor on production of beta1, 3-N-acetylglucosaminyltransferase-2 fused with GFP uv using stably transformed insect cell lines; Kwon MS et al.; A radial-flow bioreactor (RFB) with a reactor volume of 5 ml was applied to produce human beta1, 3-N-acetylglucosaminyltransferase (beta3GnT) using two stably transformed insect cell lines . When air was supplied to the RFB, cell growth was stopped at 4-d culture and beta3GnT was not detected . However, with a supply of pure oxygen, the cell concentration, assumed from glucose consumption, increased 1.3 x 10 7 cells/ml . Insect cells attached to polyvinyl alcohol (PVA) matrixes packed in the RFB and grew confluently . The 5.6 mU/ml of beta3GnT was produced under the conditions of a pure oxygen supply and the addition of glucose and glutamine . This RFB was first applied on the beta3GnT production using stably transformed insect cells . The amount of beta3GnT production in only a 5 ml scale RFB was comparable to that of a 100 ml shaking flask culture.Int J Artif Organs, 2004 Nov, 27(11), 962 - 70Hydrodynamic stimulation and long term cultivation of nucleus pulposus cells: a new bioreactor system to induce extracellular matrix synthesis by nucleus pulposus cells dependent on intermittent hydrostatic pressure; Gokorsch S et al.; A novel bioreactor system was constructed to induce extracellular matrix (ECM) synthesis by intervertebral disc (ID) cells due to intermittent hydrostatic pressure . The developed system is completely sterilizable and reusable . It is viable for cultivation, immobilization, and stimulation of various other cell types and tissues especially for cartilage . The custom made lid allows long-run cultivation through semi-continuous operation . Manual interferences and therefore the risk of contamination are reduced . Sampling, medium changing and addition of supplements are easily performed from the connected conditioning vessel, which could be placed in an incubator . For the present investigations nucleus pulposus cells from pigs were taken and immobilized in agarose to obtain three-dimensional cell matrix constructs which were subjected to intermittent hydrostatic pressure . Afterwards the construct was biochemically examined . The proven constituents of ECM were found to be released in dependence of the magnitude and profile of the applied pressure.Biotechnol Bioeng . 2005 Jan 5; {Epub ahead of print}Relationships between hydrodynamics and rheology of flocculating yeast suspensions in a high-cell-density airlift bioreactor; Klein J et al.; In this article a hydrodynamic and rheological analysis of a continuous airlift bioreactor with high-cell-density system is presented . A highly flocculating recombinant strain of Sacharomyces cerevisiae containing genes for lactose transport (lactose permease) and hydrolysis (beta-galactosidase) was exploited to ferment lactose from cheese whey to ethanol . The magnetic particle-tracer method was used to assess the effect of operational conditions (air-flow rate, biomass concentration) on hydrodynamic behavior of an airlift bioreactor during the fermentation process . Measurements of liquid circulation velocity showed the existence of a critical value of biomass concentration at which a dramatic deceleration of net liquid flow appeared with increasing biomass quantity . Rheological analysis revealed exponential increase of viscosity of the yeast floc suspension at the same biomass concentration of about 73 g/dm(3) corresponding to 42.8% v/v of solid fraction . These facts have a particular importance for the successful processing of a high-cell-density airlift bioreactor as only a circulated flow regime will be favorable to keep the solid particles in suspension state and evenly distributed throughout the bioreactor . (c) 2004 Wiley Periodicals, Inc.Zhonghua Yi Xue Za Zhi, 2004 Nov 2, 84(21), 1832 - 5{In vitro immunocompatibility of a novel bioartificial liver reactor material: propylene-acidamide grafted polypropylene membrane.}; Peng CH et al.; OBJECTIVE: To evaluate the immunocompatibility of a novel bioartificial liver bioreactor material: propylene-acidamide grafted polypropylene membrane (PP-g-AAm) in vitro on peripheral blood mononuclear cells (PBMCs) . METHODS: Fifty milliliters of peripheral blood were collected from 30 healthy young people . PBMCs were extracted and inoculated on the 24-well culture plate preset with sterilized PP-g-AAm membrane and ungrafted polypropylene (PP) membrane . Automatic biochemical analyzer was used to detect the lactic dehydrogenase (LDH) value of the PMBCs after 2 and 6 hours . The PMBCs on these 2 kinds of membrane were cultured for 6 hours and then added with lipopolysaccharide and cultured continually . Six, twelve, and thirty-six hours later the supernatant was collected . ELISA was used to detect the values of tumor necrosis factor (TNF)alpha, interleukin (IL)-1beta, and IL-6 . Flow cytometry was used to detect the expression of CD69 antigen . Scanning electron microscopy was used to observe the adhesion of PMBCs on the materials . RESULTS: After 2 hours' epimembranous inoculation the LDH value was 43.50 U/L +/- 12.71 U/L in the PP group, significantly higher than that in the PP-g-AAm group (29.13 U/L +/- 8.74 U/L, P = 0.008) and the newly extracted PMBC group (0 h group, 19.89 U/L +/- 4.67 U/L, P = 0.000) . However, the difference between the 0 h group and PP-g-AAm group (P = 0.080) was insignificant . After 6 hours' epimembranous inoculation the LDH value was 50.25 U/L +/- 13.38 U/L in the PP group, and 32.50 U/L +/- 9.21 U/L in the PP-g-AAm group (P = 0.001), both significantly higher than that in the 0 h group (P = 0.000, and P = 0.019) . There was no significant difference between the values 2 hours and 6 hours after inoculation for the two groups . No expression of TNFalpha, IL-1beta, and IL-6 was found in the supernatant of the 2 groups without LPS stimulation . Expressions of TNFalpha, IL-1beta, and IL-6 could be found at a low level 12 hours after LPS stimulation for both groups and peaked 24 hours after LPS stimulation . The expressions of TNFalpha, IL-1beta, and IL-6 were lower in the PP-g-AAm group than in the PP group (P = 0.004, P = 0.003, and P = 0.022) . The TNFalpha, IL-1beta, and IL-6 levels all decreased after 36 hours . The CD69 antigen expression rates were 17.20% +/- 3.45%and 12.02% +/- 2.44% respectively in the PP group and PP-g-AAm group, both significantly higher than that of the blank control group (3.38% +/- 1.30%, both P = 0.000) . SEM showed that the PMBCs adhered on the PP-Aam membrane were significantly less then those adhered on the PP membrane . And the PMBCs adhered on the PP-g-AAm membrane were smaller and with less microvilli . CONCLUSION: PP-g-AAm membrane has weaker activation capability to PMBCs and has better immunocompatibility in comparison with the PP membrane.IEEE Trans Nanobioscience, 2004 Dec, 3(4), 243 - 50Complex permittivity measurement as a new noninvasive tool for monitoring in vitro tissue engineering and cell signature through the detection of cell proliferation, differentiation, and pretissue formation; Bagnaninchi PO et al.; In in vitro tissue engineering, microporous scaffolds are commonly used to promote cell proliferation and differentiation in three-dimensional structures . Classic measurement methods are particularly time consuming, difficult to handle, and destructive . In this study, a new nondestructive method based on complex permittivity measurement (CPM) is proposed to monitor and track the osteoblast and macrophage differentiation through their morphological variation upon cell attachment and proliferation inside the microporous scaffolds . CPM is performed using a vector network analyzer and a dielectric probe under sterile conditions in a laminar-flow hood . A suitable effective medium approximation (EMA) is applied to fit the data in order to extract the parameters of the different constituents . Our data show that the EMA depolarization factor can be monitored to assess the variation of cell morphology characterizing cell attachment . Discrimination between two batches of scaffolds seeded, respectively, with 2 million and 1 million osteoblast cells is possible; the ratio of their CPM-derived cell volume fractions is in agreement with the ratio of their cell seeding numbers . In addition, cell proliferation inside scaffolds seeded with osteoblasts cultured in alpha minimum essential medium and inside scaffolds seeded with osteoblasts cultured in alpha minimum essential medium supplemented to induce the formation of extracellular matrix is monitored via CPM over several days . CPM-determined cell volume fraction is compared to DNA assay cell counts . Extracellular matrix formation and cell presence was confirmed by scanning electron microscopy . A set of three signature parameters (epsilon'mem, epsilon'cyt, kappa'cyt) characteristic of cell line is extracted from CPM . Distinct signatures are recorded for osteoblasts and macrophages, thus confirming the ability of CPM to discriminate between different cell types . This study demonstrates the potential of CPM as a diagnostic tool to monitor quickly and noninvasively cell growth and differentiation inside microporous scaffolds . Our findings suggest that the use of CPM could be extended to many biomedical applications, such as drug detection and automation of tissue and bacterial cultures in bioreactors.Appl Microbiol Biotechnol . 2005 Jan 4; {Epub ahead of print}Enhancement of ginsenoside biosynthesis in high-density cultivation of Panax notoginseng cells by various strategies of methyl jasmonate elicitation; Wang W et al.; A single addition of 200 muM methyl jasmonate (MJA) to high-density cell cultures of Panax notoginseng enhanced ginsenoside production in both shake-flask (250 ml) and airlift bioreactor (ALR; 1 l working volume) . Repeated elicitation with two additions of 200 muM MJA during cultivation further induced the ginsenoside biosynthesis in both cultivation vessels . The content of ginsenosides Rg(1), Re, Rb(1) and Rd in the ALR was increased from, respectively, 0.18+/-0.01, 0.21+/-0.01, 0.21+/-0.02 and 0 mg per100 mg dry cell weight (DW) in untreated cell cultures (control) to 0.32+/-0.02, 0.36+/-0.02, 0.72+/-0.06 and 0.08+/-0.01 mg per100 mg DW with a single addition of MJA and further increased to 0.43+/-0.02, 0.46+/-0.03, 1.09+/-0.07 and 0.14+/-0.02 mg per100 mg DW with two additions of MJA . Interestingly, the activity of the Rb(1) biosynthetic enzyme (UDPG-ginsenoside Rd glucosyltransferase), was also increased with a single elicitation by MJA and increased again by a repeated elicitation, which coincided well with the trend in the increase in Rb(1) content . In order to further improve the cell density and ginsenoside production, a strategy of MJA repeated elicitation combined with sucrose feeding was adopted . The final cell density and total ginsenoside content in the ALR reached 27.3+/-1.5 g/l and 2.02+/-0.06 mg per100 mg DW; and the maximum production of ginsenoside Rg(1), Re, Rb(1) and Rd was 111.8+/-4.7, 117.2+/-4.6, 290.2+/-5.1 and 32.7+/-8.1 mg/l, respectively . The strategies demonstrated and the information obtained in this work are useful for the efficient large-scale production of bioactive ginsenosides by plant cell cultures.Yi Chuan, 2004 Jan, 26(1), 75 - 83{Analysis of Codon Usage in Potato and Its Application in the Modification of t-PA Gene.}; Bai X et al.; Bioperl-1.0 was used under Hongqi LINUX system to programm the codon analysis software.According to the analysis of 98 codon DNA sequences with this software,the codon usage in potato was calculated and 4 codons have been inferred to the optimal codons.The codons of tissue plasminogen activator (t-PA) gene sequence have been reconstructed according to the results.The t-PA gene sequence containing the optimal codons of potato will be used for t-PA production by potato bioreactor.Biotechnol Bioeng . 2004 Dec 29; {Epub ahead of print}Effect of culture pH on erythropoietin production by Chinese hamster ovary cells grown in suspension at 32.5 and 37.0 degrees C; Yoon SK et al.; To investigate the effect of culture pH in the range of 6.85-7.80 on cell growth and erythropoietin (EPO) production at 32.5 and 37.0 degrees C, serum-free suspension cultures of recombinant CHO cells (rCHO) were performed in a bioreactor with pH control . Lowering culture temperature from 37.0 to 32.5 degrees C suppressed cell growth, but cell viability remained high for a longer culture period . Regardless of culture temperature, the highest specific growth rate (mu) and maximum viable cell concentration were obtained at pH values of 7.00 and 7.20, respectively . Like mu, the specific consumption rates of glucose and glutamine decreased at 32.5 degrees C compared to 37.0 degrees C . In addition, they increased with increasing culture pH . Culture pH at 32.5 degrees C affected specific EPO productivity (q(EPO)) in a different fashion from that at 37 degrees C . At 37 degrees C, the q(EPO) was fairly constant in the pH range of 6.85-7.80, while at 32.5 degrees C, the q(EPO) was significantly influenced by culture pH . The highest q(EPO) was obtained at pH 7.00 and 32.5 degrees C, and its value was approximately 1.5-fold higher than that at pH 7.00 and 37.0 degrees C . The proportion of acidic EPO isoforms, which is a critical factor for high in vivo biological activity of EPO, was highest in the stationary phase of growth, regardless of culture temperature and pH . Although cell viability rapidly decreased in death phase at both 32.5 and 37.0 degrees C, the significant degradation of produced EPO, probably by the action of proteases released from lysed cells, was observed only at 37.0 degrees C . Taken together, through the optimization of culture temperature and pH, a 3-fold increase in maximum EPO concentration and a 1.4-fold increase in volumetric productivity were obtained at pH 7.00 and 32.5 degrees C when compared with those at 37.0 degrees C . These results demonstrate the importance of optimization of culture temperature and pH for enhancing EPO production in serum-free, suspension culture of rCHO cells . (c) 2004 Wiley Periodicals, Inc.Biotechnol Bioeng . 2004 Dec 29; {Epub ahead of print}Low-cost noninvasive optical CO(2) sensing system for fermentation and cell culture; Ge X et al.; High-throughput bioprocessing is a very promising technique for bioprocess development and optimization because of its high efficiency . The key to its development has been the availability of simple and inexpensive sensors to monitor the bioprocesses conducted in its small-scale bioreactors . Here we report on a low-cost noninvasive CO(2) sensing system suitable for any transparent vessel . The system was composed of a CO(2) sensing patch, a coaster, an interface, and a computer . The sensing film was prepared using the ion-pair technique . The coaster was a small self-made device with necessary optics and electronics for ratiometric measurements with a component cost of less than $100 . Results show that the system was stable and reliable despite its simplicity and low cost . The sensitivity of the CO(2) sensing system was not affected by pH, media type, or temperature . It was shown to be stable for at least 10 days, long enough for most bioprocesses . (c) 2004 Wiley Periodicals, Inc.Huan Jing Ke Xue, 2004 Sep, 25(5), 102 - 5{Printing and dyeing wastewater treatment using combined process of anaerobic bioreactor and MBR}; Zheng X et al.; This paper describes a labor-scale experiment for printing and dyeing wastewater treatment of woolen mill using a combined process of an anaerobic reactor and a membrane bioreactor (MBR) . The experimental results showed that when the concentration of COD, BOD5 and color in the influent were 128-321 mg/L, 36-95 mg/L and 40-70 dilution times (DT), the average concentrations of COD, BOD5, color and turbidity in the effluent were 36.9 mg/L, 3.7 mg/L, 21 DT and 0.24 NTU, respectively, and the corresponding removal rates were 80.3%, 95%, 59% and 99.3%, respectively . A new integrated membrane bioreactor by gravity drain of liquid level in the bioreactor was developed in this study . It not only lessens suction pump for effluent and controlling unit comparing to the traditional integrated membrane bioreactor, but also has the characters of high and continuous flux, concise configuration and simple operation and maintenance . According to the experimental results, the air flow rate through the membrane module is a significant factor to affect the flux rate and cake layer deposited on the membrane . With application of optimal air flow rate, it is effective to reduce membrane fouling and maintain high flux rate.