Biomedical subjects

Shang-Tian Yang

Publications and source records attributed to Shang-Tian Yang.

At least 19 recordsLinked to original sources

PubMed2007

Effects of mixing intensity on cell seeding and proliferation in three-dimensional fibrous matrices.

Nonwoven fibrous matrices have been widely used in cell and tissue cultures because their three-dimensional (3-D) structures with large surface areas and pore spaces can support high-density cell growth. Although cell adherence and growth on 2-D surfaces have been thoroughly investigated, very little is known for cells cultured in 3-D matrices. The effects of mixing intensity on cell seeding, adherence, and growth in fibrous matrices were thus investigated. Chinese Hamster Ovary and osteosarcoma cells were inoculated into nonwoven polyethylene terephthalate matrices by dynamic and static seeding methods, of which the former was found to be superior in seeding efficiency and cell distribution in the matrices. Dynamic seeding increased seeding efficiency from approximately 40% to more than 90%. When higher mixing intensities were applied, both cell attachment and detachment rates increased. Cell attachment was transport limited, as indicated by the increased attachment rate with increasing the mass transfer coefficient of the cells. Meanwhile, cell detachment from the 3-D matrix can be described by the Bell model. The effects of matrix pore size on cell adherence and proliferation were also investigated. In general, the smaller pore size is favorable to cell attachment and proliferation. Further analysis revealed that the interaction between mixing intensity and pore size played a vital role in hydrodynamic damage to cells, which was found to be significant when the Kolomogorov eddy size was smaller than the matrix pores. Increasing mixing intensity also increased oxygen transfer, decreased the lactate yield from glucose, and improved cell growth.

Animals↗

PubMed2006

Surface modification for enhancing antibody binding on polymer-based microfluidic device for enzyme-linked immunosorbent assay.

A novel surface treatment method using poly(ethyleneimine) (PEI), an amine-bearing polymer, was developed to enhance antibody binding on the poly(methyl methacrylate) (PMMA) microfluidic immunoassay device. By treating the PMMA surface of the microchannel on the microfluidic device with PEI, 10 times more active antibodies can be bound to the microchannel surface as compared to those without treatment or treated with the small amine-bearing molecule, hexamethylenediamine (HMD). Consequently, PEI surface modification greatly improved the immunoassay performance of the microfluidic device, making it more sensitive and reliable in the detection of IgG. The improvement can be attributed to the spacer effect as well as the functional amine groups provided by the polymeric PEI molecules. Due to the smaller dimensions (140x125 microm) of the microchannel, the time required for antibody diffusion and adsorption onto the microchannel surface was reduced to only several minutes, which was 10 times faster than the similar process carried out in 96-well plates. The microchip also had a wider detection dynamic range, from 5 to 1000 ng/mL, as compared to that of the microtiter plate (from 2 to 100 ng/mL). With the PEI surface modification, PMMA-based microchips can be effectively used for enzyme linked immunosorbent assays (ELISA) with a similar detection limit, but much less reagent consumption and shorter assay time as compared to the conventional 96-well plate.

Animals↗

PubMed2006

Long-term culturing of undifferentiated embryonic stem cells in conditioned media and three-dimensional fibrous matrices without extracellular matrix coating.

ESCs have unlimited proliferation potential and capability to differentiate into all tissue types. They are ideal cell sources for tissue engineering and cell therapy, but their supplies are limited. Current in vitro ESC cultures are carried out in tissue flasks with the surface precoated with extracellular matrix (ECM) proteins. T-flask cultures also require frequent subculturing because their limited surface area cannot support long-term growth of ESCs. In this work, ECM coating and frequent subculturing required in two-dimensional (2D) cultures were circumvented by culturing murine ESCs in three-dimensional (3D) polyethylene terephthalate (PET) fibrous matrices. Also, media conditioned with STO fibroblast cells were used to replace leukemia inhibitory factor and to effectively maintain the pluripotency of murine ESCs in a long-term static culture. However, the lactic acid present in the conditioned medium could inhibit ESC growth and induce spontaneous differentiation when its concentration exceeded 1.5 g/l. In addition, the 3D static culture could be limited by oxygen, which was depleted in the long-term culture when cell density in the matrix was high. However, these problems can be alleviated in dynamic culture with improved oxygen transfer and continuous media perfusion. The matrix pore size also had profound effects on ESCs. The smaller-pore (30-60 mum) matrix gave a higher proliferation rate and Oct-4 and stage specific embryonic antigen-1 expressions. Overall, the 3D culturing method is superior to the 2D culture method and can provide an economical way to mass-produce undifferentiated ESCs in uncoated matrices and conditioned media.

Animals↗

PubMed2006

Construction and characterization of ack knock-out mutants of Propionibacterium acidipropionici for enhanced propionic acid fermentation.

Propionibacterium acidipropionici produces propionic acid from glucose with acetic acid, succinic acid, and CO2 as byproducts. In this work, inactivation of ack gene, encoding acetate kinase (AK), by gene disruption and integrational mutagenesis was studied as a method to reduce acetate formation in propionic acid fermentation. The partial ack gene of approximately 750 bp in P. acidipropionici was cloned using a PCR-based method with degenerate primers and sequenced. The deduced amino acid sequence had 88% similarity and 76% identity with the amino acid sequence of AK from Bacillus subtilis. The partial ack gene was used to construct a linear DNA fragment with an inserted tetracycline resistance cassette and a nonreplicative integrational plasmid containing a tetracycline resistance gene cassette. These DNA constructs were then introduced into P. acidipropionici by electroporation, resulting in two mutants, ACK-Tet and TAT-ACK-Tet, respectively. Southern hybridization confirmed that the ack gene in the mutant ACK-Tet was disrupted by the inserted tetracycline resistance gene. As compared to the wild-type, the activities of AK were reduced by 26% and 43% in ACK-Tet and TAT-ACK-Tet mutants, respectively. The specific growth rate of these mutants was reduced by approximately 25% to 0.10/h (0.13/h for the wild-type), probably because of reduced acetate and ATP production. Both mutants produced approximately 14% less acetate from glucose. Although ack disruption alone did not completely eliminate acetate production, the propionate yield was increased by approximately 13%.

Acetate Kinase↗

PubMed2005

Biotransformation of R-2-hydroxy-4-phenylbutyric acid by D-lactate dehydrogenase and Candida boidinii cells containing formate dehydrogenase coimmobilized in a fibrous bed bioreactor.

R-2-hydroxy-4-phenylbutyric acid (R-HPBA) is an important intermediate in the manufacture of angiotensin converting enzyme inhibitors. In this work, a recombinant D-lactate dehydrogenase (LDH) was used to transform 2-oxo-4-phenylbutyric acid (OPBA) to R-HPBA, with concomitant oxidation of beta-nicotinamide adenine dinucleotide (NADH) to NAD(+). The cofactor NADH was regenerated by formate dehydrogenase (FDH) present in whole cells of Candida boidinii, which were pre-treated with toluene to make them permeable. The whole cells used in the process were more stable and easier to prepare as compared with the isolated FDH from the cells. Kinetic study showed that the reaction rate was dependent on the concentration of cofactor, NAD(+), and that both R-HPBA and OPBA inhibited the reaction. A novel method for co-immobilization of whole cells and LDH enzyme on cotton cloth was developed using polyethyleneimine (PEI), which induced the formation of PEI-enzyme-cell aggregates and their adsorption onto cotton cloth, leading to multilayer co-immobilization of cells and enzyme with high loading (0.5 g cell and 8 mg LDH per gram of cotton cloth) and activity yield ( > 95%). A fibrous bed bioreactor with co-immobilized cells and enzyme on the cotton cloth was then evaluated for R-HPBA production in fed-batch and repeated batch modes, which gave relatively stable reactor productivity of 9 g/L . h and product yield of 0.95 mol/mol OPBA when the concentrations of OPBA and R-HPBA were less than 10 g/L.

Bioreactors↗

PubMed2005

Enhanced propionic acid fermentation by Propionibacterium acidipropionici mutant obtained by adaptation in a fibrous-bed bioreactor.

Fed-batch fermentations of glucose by P. acidipropionici ATCC 4875 in free-cell suspension culture and immobilized in a fibrous-bed bioreactor (FBB) were studied. The latter produced a much higher propionic acid concentration (71.8 +/- 0.8 g/L vs. 52.2 +/- 1.1 g/L), indicating enhanced tolerance to propionic acid inhibition by cells adapted in the FBB. Compared to the free-cell fermentation, the FBB culture produced 20-59% more propionate (0.40-0.65 +/- 0.02 g/g vs. 0.41 +/- 0.02 g/g), 17% less acetate (0.10 +/- 0.01 g/g vs. 0.12 +/- 0.02 g/g), and 50% less succinate (0.09 +/- 0.02 g/g vs. 0.18 +/- 0.03 g/g) from glucose. The higher propionate production in the FBB was attributed to mutations in two key enzymes, oxaloacetate transcarboxylase and propionyl CoA: succinyl CoA transferase, leading to the production of propionic acid from pyruvate. Both showed higher specific activity and lower sensitivity to propionic acid inhibition in the mutant than in the wild type. In contrast, the activity of PEP carboxylase, which converts PEP directly to oxaloacetate and leads to the production of succinate from glucose, was generally lower in the mutant than in the wild type. For phosphotransacetylase and acetate kinase in the acetate formation pathway, however, there was no significant difference between the mutant and the wild type. In addition, the mutant had a striking change in its morphology. With a threefold increase in its length and approximately 24% decrease in its diameter, the mutant cell had an approximately 10% higher specific surface area that should have made the mutant more efficient in transporting substrates and metabolites across the cell membrane. A slightly lower membrane-bound ATPase activity found in the mutant also indicated that the mutant might have a more efficient proton pump to allow it to better tolerate propionic acid. In addition, the mutant had more longer-chain saturated fatty acids (C17:0) and less unsaturated fatty acids (C18:1), both of which could decrease membrane fluidity and might have contributed to the increased propionate tolerance. The enhanced propionic acid production from glucose by P. acidipropionici was thus attributed to both a high viable cell density maintained in the reactor and favorable mutations resulted from adaptation by cell immobilization in the FBB.

Adenosine Triphosphatases↗

PubMed2005

Construction and characterization of pta gene-deleted mutant of Clostridium tyrobutyricum for enhanced butyric acid fermentation.

Clostridium tyrobutyricum ATCC 25755 is an acidogenic bacterium, producing butyrate and acetate as its main fermentation products. In order to decrease acetate and increase butyrate production, integrational mutagenesis was used to disrupt the gene associated with the acetate formation pathway in C. tyrobutyricum. A nonreplicative integrational plasmid containing the phosphotransacetylase gene (pta) fragment cloned from C. tyrobutyricum by using degenerate primers and an erythromycin resistance cassette were constructed and introduced into C. tyrobutyricum by electroporation. Integration of the plasmid into the homologous region on the chromosome inactivated the target pta gene and produced the pta-deleted mutant (PTA-Em), which was confirmed by Southern hybridization. SDS-PAGE and two-dimensional protein electrophoresis results indicated that protein expression was changed in the mutant. Enzyme activity assays using the cell lysate showed that the activities of PTA and acetate kinase (AK) in the mutant were reduced by more than 60% for PTA and 80% for AK. The mutant grew more slowly in batch fermentation with glucose as the substrate but produced 15% more butyrate and 14% less acetate as compared to the wild-type strain. Its butyrate productivity was approximately 2-fold higher than the wild-type strain. Moreover, the mutant showed much higher tolerance to butyrate inhibition, and the final butyrate concentration was improved by 68%. However, inactivation of pta gene did not completely eliminate acetate production in the fermentation, suggesting the existence of other enzymes (or pathways) also leading to acetate formation. This is the first-reported genetic engineering study demonstrating the feasibility of using a gene-inactivation technique to manipulate the acetic acid formation pathway in C. tyrobutyricum in order to improve butyric acid production from glucose.

Amino Acid Sequence↗

PubMed2005

Fabrication of well-defined PLGA scaffolds using novel microembossing and carbon dioxide bonding.

A novel biologically benign technique was developed to produce three-dimensional tissue engineering scaffolds with well-defined structure. Photolithography was used to design and pattern a planar scaffold skeletal structure on a photoresist (SU-8), and a variety of microembossing processes including sacrificial layer embossing and bilayer embossing were developed to transfer the skeletal pattern to the poly(DL-lactide-co-glycolide) substrate as scaffold skeletons. Subcritical carbon dioxide was then introduced to assemble these skeletons to a three-dimensional scaffold at a low temperature. Compared with conventional scaffolds, which have a broad pore size distribution and varying pore geometry, these microfabricated scaffolds have a uniform and well-defined geometry and structure. This uniformity of structural parameters allows for the studies of cell attachment, spreading, and proliferation in scaffolds in a controlled and logical manner. The cytocompatibility of these microfabricated scaffolds was tested by seeding three different cell lines with different morphologies and growth patterns into these scaffolds. All three cell lines attached well to the scaffolds and grew to high densities as observed with scanning electron microscopy. This study demonstrates a controllable method to fabricate tissue scaffolds with a well-defined 3D architecture that can be used to better elucidate the effect of structure parameters such as pore geometry and pore size on tissue growth in 3D scaffolds.

Animals↗

PubMed2004

Effect of pH on metabolic pathway shift in fermentation of xylose by Clostridium tyrobutyricum.

The effect of pH (between 5.0 and 6.3) on butyric acid fermentation of xylose by Clostridium tyrobutyricum was studied. At pH 6.3, the fermentation gave a high butyrate production of 57.9 g l(-1) with a yield of 0.38-0.59 g g(-1) xylose and a reactor productivity up to 3.19 g l(-1)h(-1). However, at low pHs (<5.7), the fermentation produced more acetate and lactate as the main products, with only a small amount of butyric acid. The metabolic shift from butyrate formation to lactate and acetate formation in the fermentation was found to be associated with changes in the activities of several key enzymes. The activities of phosphotransbutyrylase (PTB), which is the key enzyme controlling butyrate formation, and NAD-independent lactate dehydrogenase (iLDH), which catalyzes the conversion of lactate to pyruvate, were higher in cells producing mainly butyrate at pH 6.3. In contrast, cells at pH 5.0 had higher activities of phosphotransacetylase (PTA), which is the key enzyme controlling acetate formation, and lactate dehydrogenase (LDH), which catalyzes the conversion of pyruvate to lactate. Also, PTA was very sensitive to the inhibition by butyric acid. Difference in the specific metabolic rate of xylose at different pHs suggests that the balance in NADH is a key in controlling the metabolic pathway used by the cells in the fermentation.

Acetate Kinase↗

PubMed2004

Design of a compact disk-like microfluidic platform for enzyme-linked immunosorbent assay.

This paper presents an integrated microfluidic device on a compact disk (CD) that performs an enzyme-linked immunosorbent assay (ELISA) for rat IgG from a hybridoma cell culture. Centrifugal and capillary forces were used to control the flow sequence of different solutions involved in the ELISA process. The microfluidic device was fabricated on a plastic CD. Each step of the ELISA process was carried out automatically by controlling the rotation speed of the CD. The work on analysis of rat IgG from hybridoma culture showed that the microchip-based ELISA has the same detection range as the conventional method on the 96-well microtiter plate but has advantages such as less reagent consumption and shorter assay time over the conventional method.

Animals↗

PubMed2004

A fibrous-bed bioreactor for continuous production of monoclonal antibody by hybridoma.

A fibrous-bed bioreactor (FBB) has been developed to culture hybridoma cells for long-term continuous production of monoclonal antibody (MAb). A non-woven polyester fibrous matrix was used to immobilize the cells to reach a high viable cell density of 3 x 10(8) cells cm(-3) packed bed, which gave a high volumetric MAb productivity of 1 g L(-1) day(-1) under continuous feed conditions with the medium containing 10% serum. Reducing the medium serum content to 1% increased MAb production to 6.5 g L(-1) day(-1) in a repeated batch FBB culture. MAb production was higher at higher dissolved oxygen (DO) levels in the range between 10% and 70% of air saturation, although DO did not significantly affect glucose metabolism and lactate production. The medium LDH (lactate dehydrogenase) level increased dramatically when the DO level was decreased from 30% to 10%, suggesting that a critical DO level of approximately 30% is necessary for maintaining the FBB culture for long-term operation. Compared with suspension cultures in T-flasks and spinner flasks, the FBB culture had a lower lactate yield from glucose (0.80 vs. 0.91 g g(-1), produced MAb at a higher concentration (up to 442 mg L(-1) vs. 83.5 mg L(-1), and was stable for continuous long-term operation (more than 1 month). The superior FBB performance was attributed to the highly porous fibrous matrix that enabled the efficient mass transfer, cell immobilization, and continued growth and regeneration that are critical to maintaining a high density of viable and productive cell populations. The cells immobilized in the fibrous matrix had high viability (>85%) even though many of them were in growth arrest (G1/G0 phase) as indicated by their smaller cell size (<10 microm). Scanning electron microscopic studies of the cell-matrix showed that the high density of cells formed large clumps within the interstitial spaces of the fibrous matrix. Their close contact and interaction with each other might have contributed to their ability to survive well under adverse conditions such as low DO and low serum content in the medium. It was also found that the cells present inside the fibrous matrix had a higher viability and lower apoptosis than those present in the liquid suspension, indicating that the fibrous matrix had selectively retained healthy, nonapoptotic cells and dislodged apoptotic and dead cells; this also might have contributed to the stability of the long-term culture. This work demonstrated that the FBB originally developed for microbial fermentation also gave excellent results in achieving high cell density, productivity, and product concentrations, and should have a good potential for industrial animal cell culture applications.

Antibodies, Monoclonal↗

PubMed2004

A hollow-fiber membrane extraction process for recovery and separation of lactic acid from aqueous solution.

An energy-efficient hollow-fiber membrane extraction process was successfully developed to separate and recover lactic acid produced in fermentation. Although many fermentation processes have been developed for lactic acid production, an economical method for lactic acid recovery from the fermentation broth is still needed. Continuous extraction of lactic acid from a simulated aqueous stream was achieved by using Alamine 336 in 2-octanol contained in a hollow-fiber membrane extractor. In this process, the extractant was simultaneously regenerated by stripping with NaOH in a second membrane extractor, and the final product is a concentrated lactate salt solution. The extraction rate increased linearly with an increase in the Alamine 336 content in the solvent (from 5 to 40%). Increasing the concentration of the undissociated lactic acid in the feed solution by either increasing the lactate concentration (from 5 to 40 g/L) or decreasing the solution pH (from 5.0 to 4.0) also increased the extraction rate. Based on these observations, a reactive extraction model with a first-order reaction mechanism for both lactic acid and amine concentrations was proposed. The extraction rate also increased with an increase in the feed flow rate, but not the flow rates of solvent and the stripping solution, suggesting that the process was not limited by diffusion in the liquid films or membrane pores. A mathematical model considering both diffusion and chemical reaction in the extractor and back extractor was developed to simulate the process. The model fits the experimental data well and can be used in scale up design of the process.

Biotechnology↗

PubMed2004

Continuous production of butanol by Clostridium acetobutylicum immobilized in a fibrous bed bioreactor.

We explored the influence of dilution rate and pH in continuous cultures of Clostridium acetobutylicum. A 200-mL fibrous bed bioreactor was used to produce high cell density and butyrate concentrations at pH 5.4 and 35degreesC. By feeding glucose and butyrate as a cosubstrate, the fermentation was maintained in the solventogenesis phase, and the optimal butanol productivity of 4.6 g/(L h) and a yield of 0.42 g/g were obtained at a dilution rate of 0.9 h-1 and pH 4.3. Compared to the conventional acetone-butanol-ethanol fermentation, the new fermentation process greatly improved butanol yield, making butanol production from corn an attractive alternative to ethanol fermentation.

Acetone↗

PubMed2003

Extractive fermentation for butyric acid production from glucose by Clostridium tyrobutyricum.

A novel extractive fermentation for butyric acid production from glucose, using immobilized cells of Clostridium tyrobutyricum in a fibrous bed bioreactor, was developed by using 10% (v/v) Alamine 336 in oleyl alcohol as the extractant contained in a hollow-fiber membrane extractor for selective removal of butyric acid from the fermentation broth. The extractant was simultaneously regenerated by stripping with NaOH in a second membrane extractor. The fermentation pH was self-regulated by a balance between acid production and removal by extraction, and was kept at approximately pH 5.5 throughout the study. Compared with conventional fermentation, extractive fermentation resulted in a much higher product concentration (>300 g/L) and product purity (91%). It also resulted in higher reactor productivity (7.37 g/L. h) and butyric acid yield (0.45 g/g). Without on-line extraction to remove the acid products, at the optimal pH of 6.0, the final butyric acid concentration was only approximately 43.4 g/L, butyric acid yield was 0.423 g/g, and reactor productivity was 6.77 g/L. h. These values were much lower at pH 5.5: 20.4 g/L, 0.38 g/g, and 5.11 g/L. h, respectively. The improved performance for extractive fermentation can be attributed to the reduced product inhibition by selective removal of butyric acid from the fermentation broth. The solvent was found to be toxic to free cells in suspension, but not harmful to cells immobilized in the fibrous bed. The process was stable and provided consistent long-term performance for the entire 2-week period of study.

Bioreactors↗

PubMed2002

Production of L(+)-lactic acid from glucose and starch by immobilized cells of Rhizopus oryzae in a rotating fibrous bed bioreactor.

A rotating fibrous-bed bioreactor (RFB) was developed for fermentation to produce L(+)-lactic acid from glucose and cornstarch by Rhizopus oryzae. Fungal mycelia were immobilized on cotton cloth in the RFB for a prolonged period to study the fermentation kinetics and process stability. The pH and dissolved oxygen concentration (DO) were found to have significant effects on lactic acid productivity and yield, with pH 6 and 90% DO being the optimal conditions. A high lactic acid yield of 90% (w/w) and productivity of 2.5 g/L.h (467 g/h.m(2)) was obtained from glucose in fed-batch fermentation. When cornstarch was used as the substrate, the lactic acid yield was close to 100% (w/w) and the productivity was 1.65 g/L.h (300 g/h.m(2)). The highest concentration of lactic acid achieved in these fed-batch fermentations was 127 g/L. The immobilized-cells fermentation in the RFB gave a virtually cell-free fermentation broth and provided many advantages over conventional fermentation processes, especially those with freely suspended fungal cells. Without immobilization with the cotton cloth, mycelia grew everywhere in the fermentor and caused serious problems in reactor control and operation and consequently the fermentation was poor in lactic acid production. Oxygen transfer in the RFB was also studied and the volumetric oxygen transfer coefficients under various aeration and agitation conditions were determined and then used to estimate the oxygen transfer rate and uptake rate during the fermentation. The results showed that the oxygen uptake rate increased with increasing DO, indicating that oxygen transfer was limited by the diffusion inside the mycelial layer.

Bioreactors↗

PubMed2002

A fibrous-bed bioreactor for continuous production of developmental endothelial locus-1 by osteosarcoma cells.

Genetically engineered human osteosarcoma cells containing developmental endothelial locus-1 (del-1) gene were studied for production of Del-1, a protein that has the properties of an extracellular matrix protein and can regulate vascular morphogenesis and remodeling. Del-1 has been studied as a potential anti-angiogenesis drug targeting solid tumors. In this study, osteosarcoma cells were cultured in a fibrous-bed bioreactor (FBB) to continuously produce Del-1. The FBB was constructed by packing a polyester fibrous matrix into a 1.5-l spinner flask. The effects of media composition, including the serum content in the medium, and dilution rate on cell growth, metabolism, and Del-1 production were studied. A gradual reduction of serum content from 10% (v/v) to 0.5% (v/v) caused no loss in Del-1 production. However, the production of Del-1 decreased significantly in a serum-free medium, suggesting some nutrients present in the serum were important to culture viability and Del-1 production. The continuous FBB culture was stable for long-term production of Del-1, with a higher Del-1 titer than that normally obtained in T-flask cultures and overall productivity similar to the total production from 300 25-cm(2) T-flasks. Reducing geneticin in the medium from 250 microg ml(-1) to zero at later culturing stages had no significant effect on Del-1 production. The FBB was operated for a period of more than 4 months without any notable degeneration, and reached a final cell density of 3 x 10(8) cells ml(-1) of packing volume with >90% cell viability. The good reactor performance can be attributed to the three-dimensional environment provided by the fibrous matrix that allows for efficient mass transfer and cell immobilization and growth. Scanning electron microscopic and confocal scanning laser microscopic studies of the cell-matrix showed that cells formed large aggregates in the fibrous matrix and cell density was relatively uniform in the matrix.

Anti-Bacterial Agents↗

PubMed2002

Production of galacto-oligosaccharides from lactose by Aspergillus oryzae beta-galactosidase immobilized on cotton cloth.

The production of galacto-oligosaccharides (GOS) from lactose by A. oryzae beta-galactosidase immobilized on cotton cloth was studied. The total amounts and types of GOS produced were mainly affected by the initial lactose concentration in the reaction media. In general, more and larger GOS can be produced with higher initial lactose concentrations. A maximum GOS production of 27% (w/w) of initial lactose was achieved at 50% lactose conversion with 500 g/L of initial lactose concentration. Tri-saccharides were the major types of GOS formed, accounting for more than 70% of the total GOS produced in the reactions. Temperature and pH affected the reaction rate, but did not result in any changes in GOS formation. The presence of galactose and glucose at the concentrations encountered near maximum GOS greatly inhibited the reactions and reduced GOS yield by as much as 15%. The cotton cloth as the support matrix for enzyme immobilization did not affect the GOS formation characteristics of the enzyme, suggesting no diffusion limitation in the enzyme carrier. The thermal stability of the enzyme increased approximately 25-fold upon immobilization on cotton cloth. The half-life for the immobilized enzyme on cotton cloth was more than 1 year at 40 degrees C and 48 days at 50 degrees C. Stable, continuous operation in a plugflow reactor was demonstrated for 2 weeks without any apparent problem. A maximum GOS production of 21 and 26% (w/w) of total sugars was attained with a feed solution containing 200 and 400 g/L of lactose, respectively, at pH 4.5 and 40 degrees C. The corresponding reactor productivities were 80 and 106 g/L/h, respectively, which are at least several-fold higher than those previously reported.

Aspergillus oryzae↗

PubMed2002

Production of carboxylic acids from hydrolyzed corn meal by immobilized cell fermentation in a fibrous-bed bioreactor.

Corn meal hydrolyzed with amylases was used as the carbon source for producing acetic, propionic, and butyric acids via anaerobic fermentations. In this study, corn meal, containing 75% (w/w) starch, 20% (w/w) fibers, and 1.5% (w/w) protein, was first hydrolyzed using amylases at 60 degrees C. The hydrolysis yielded approximately 100% recovery of starch converted to glucose and 17.9% recovery of protein. The resulting corn meal hydrolyzate was then used, after sterilization, for fermentation studies. A co-culture of Lactococcus lactis and Clostridium formicoaceticum was used to produce acetic acid from glucose. Propionibacterium acidipropionici was used for propionic acid fermentation, and Clostridium tyrobutylicum was used for butyric acid production. These cells were immobilized on a spirally wound fibrous matrix packed in a fibrous-bed bioreactor (FBB) developed for multi-phase biological reactions or fermentation. The bioreactor was connected to a stirred-tank fermentor that provided pH and temperature controls via medium circulation. The fermentation system was operated at the recycle batch mode. Temperature and pH were controlled at 37 degrees C and 7.6, respectively, for acetic acid fermentation, 32 degrees C and 6.0, respectively, for propionic acid fermentation, and 37 degrees C and 6.0, respectively, for butyric acid production. The fermentation demonstrated a yield of approximately 100% and a volumetric productivity of approximately 1 g/(1 h) for acetic acid production. The propionic acid fermentation achieved an approximately 60% yield and a productivity of 2.12 g/(1 h), whereas the butyric acid fermentation obtained an approximately 50% yield and a productivity of 6.78 g/(1 h). These results were comparable to, or better than those fermentations using chemically defined media containing glucose as the substrate, suggesting that these carboxylic acids can be efficiently produced from direct fermentation of corn meal hydrolyzate. The corn fiber present as suspended solids in the corn meal hydrolyzate did not cause operating problem to the immobilized cell bioreactor as is usually encountered by conventional immobilized cell bioreactor systems. It is concluded that the FBB technology is suitable for producing value-added biochemicals directly from agricultural residues or commodities such as corn meal.

Acetic Acid↗