摘要:

AbstractIn this study, different mole fractions of pureThermomonospora fuscaE5and E3, plusTrichoderma reeseiCBHI were studied for reducing sugar production at 2 h, degree of synergism, and cellulose binding. In addition, the effects of introducing theCaldocellum saccharolyticumβ‐glucosidase into this cellulase system were investigated. The cellulases used were purified to homogeneity. Avicel PH 102 (4% w/w solution in 0.05 sodium acetate pH 5.5 buffer) was the substrate. Reactions were run at 50°C for 2 h using total cellulase concentrations of 8.3 or 12.2 μM. A bimixture ofT. fuscaE3andT. reeseiCBHI was very effective in hydrolyzing microcrystalline cellulose (9.1% conversion). The addition of endoglucanase E5to the mixture only increased conversion to 9.8%. However, when both E5and β‐glucosidase were added, conversion increased to 14%. It was also observed that increasing total cellulase concentration beyond 8.3 μMdid little to increase percent conversion of cellulose into glucose. The results of the binding studies indicate no competition for binding sites between the endo‐ and exocellulases. © 1993 John Wile

ISSN:0006-3592    

DOI:10.1002/bit.260420902

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1993

数据来源: WILEY

摘要:

AbstractThe use of a temperature switch to control the growth and productivity of temperature‐sensitive (ts) mutants was investigated to extend the productive life span of recombinant Chinese hamster ovary (CHO) cells in batch culture. Bromodeoxyuridine was used at 39°C to select mutagenized CHO‐K1 cells, which resulted in the isolation of 31 temperature‐sensitive mutants that were growth inhibited at 39°C. Two of these mutants were successfully transfected with the gene for tissue inhibitor of metalloproteinases (TIMP) using glutamine synthetase amplification, and a permanent recombinant cell line established (5G1‐B1) that maintains thetsphenotype.Continuous exposure to the nonpermissive temperature (npt) of 39°C led to a rapid decline in cell viability. However, a temperature regime using alternating incubations at 34°C and 39°C arrested the 5G1‐B1 cells while retaining a high cell viability for up to 170 h in culture. The specific production rate of the growth‐arrested cells was 3–4 times that of control cultures maintained at a constant 34°C over the crucial 72–130‐h period of culture, which resulted in a 35% increase in the maximum product yield. Glucose uptake and lactate production both decreased in arrested cells. Flow cytometric analysis indicated that 5G1‐B1 cells arrested in the G1or G0phase of the cell cycle, and no major structural damage was caused to these cells by the alternating temperature regime.These results demonstrate that growth‐arrestedtsCHO cells have increased productivity compared to growing cultures and maintain viability for longer periods. The system offers the prospect of enhancing the productivity of recombinant mammalian cells grown in simple batch fermentors.

ISSN:0006-3592    

DOI:10.1002/bit.260420903

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1993

数据来源: WILEY

摘要:

AbstractUsing monomers that polymerize to form electrically conducting polymers, one can control the thickness of the polymer film and the amount of enzyme that can be immobilized in the films. First, an investigation of the major variables that influence the immobilization of glucose oxidase by entrapment in polypyrrole films, prepared by electropolymerization from aqueous solutions containing the enzyme and monomer, was carried out. Then the optimized conditions were used to assess the effects of film thickness on the activity and stability of immobilized enzyme. For the films ranged in thickness from 0.1 μm to 1.6 μm, the resulting apparent activity and stability of the immobilized enzyme were found to be a strong function of the polymer film thickness. Above a thickness of 1.0 μm, the apparent activity of the immobilized enzyme increases linearly with increasing film thickness. The nonlinearity observed for films of thickness less than 1.0 μm can be attributed to the changes observed in the morphology of the resulting polypyrrole films. Furthermore, it was noted that when the glucose oxidase/polypyrrole films are stored in phosphate buffer, at 4°C, the observed rate of loss in apparent activity of the immobilized enzyme is highest for the first few days, also being higher for the thinner films. However, after the loosely entrapped enzyme is leached from the polymer film, the rate of loss in activity is very low indicating that the well‐entrapped enzyme, as well as the polypyrrole films, exhibit good stability. Finally, the reproducibility of the immobilization technique is excellent. © 1993 John Wiley&So

ISSN:0006-3592    

DOI:10.1002/bit.260420904

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1993

数据来源: WILEY

摘要:

AbstractAspergillus nigerwas grown in a 7‐L chemostat at biomass levels of 7 to 9 gL−1; dilution rates of 0.03, 0.05, 0.075, and 0.009 h−1; and dissolved oxygen tensions of 7%, 12%, and 40% of air saturation. Broth rheological measurements were made on‐line, while off‐line image analysis was used to measure mycelial morphology, including characterization of mycelial aggregates (clumps). Under all conditions, more than 87% of the hyphase were in clumps, the shape of which determined the rheological characteristics of the broth. In particular, the power law consistency index could be correlated with the biomass concentration and the roughness factor of the clumps, which describes their hairiness. A decrease in specific growth rate decreased roughness, possibly due to changes in the amount of clump breakup. However, decreases of roughness with increasing dissolved oxygen tension might rather imply some effect on hyphal–hyphal interactions within the clumps. © 1993 John Wil

ISSN:0006-3592    

DOI:10.1002/bit.260420905

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1993

数据来源: WILEY

摘要:

AbstractThe ability to genetically alter the product‐formation capabilities ofClostridium acetobutylicumis necessary for continued progress toward industrial production of the solvents butanol and acetone by fermentation. Batch fermentations at pH 4.5, 5.5, or 6.5 were conducted usingC. acetobutylicumATCC 824 (pFNK6). Plasmid pFNK6 contains a synthetic operon (the “ace operon”) in which the three homologous acetone‐formation genas (adc,ctfA, andctfB) are transcribed from theadcpromoter. The corresponding enzymes (acetoacetate decarboxylase and CoA‐transferase) were best expressed in pH 4.5 fermentations. However, the highest levels of solvents were attained at pH 5.5. Relative to the plasmid‐free control strain at pH 5.5, ATCC 824 (pFNK6) produced 95%, 37%, and 90% higher final concentrations of acetone, butanol, and ethanol, respectively; a 50% higher yield (g/g) of solvents on glucose; and a 22‐fold lower mass of residual carboxylic acids. At all pH values, the acetone‐formation enzymes were expressed earlier with ATCC 824 (pFNK6) than in control fermentations, leading to earlier induction of acetone formation. Furthermore, strain ATCC 824 (pFNK6) produced butanol significantly earlier in the fermentation and produced significant levels of solvents at pH 6.5. Only trace levels of solvents were produced by strain ATCC 824 at pH 6.5. Compared with ATCC 824, a plasmid‐control strain containing a vector without theaceoperon also produced higher levels of solvents [although lower than those of strain ATCC 824 (pFNK6)] and lower levels of acids. Strains containing plasmid‐borne derivatives of theaceoperon, in which either the acetoacetate decarboxylase or CoA‐transferase alone were expressed at elevated levels, produced acids and solvents at levels similar to those of the plasmid‐control strain. © 1

ISSN:0006-3592    

DOI:10.1002/bit.260420906

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1993

数据来源: WILEY

摘要:

AbstractAn inactivation model previously developed to characterize the rate of enzyme activity loss in unstirred solutions was extended to take into account orthokinetic interactions resulting from convective mixing. A synergistic relationship between shear rate and temperature was observed; the rate of inactivation of the enzyme dextransucrase was unaffected by the action of shear below 25°C, but was increased by the shear rate at 30°C. Shear rate does not appear to influence the equilibrium between native and denatured dextransucrase either directly in solution or indirectly by augmenting the turnover of the gas–liquid interface. However, a second‐order plot of the inverse of relative activity (AO/A) versusGt(shear rate × time) of dextransucrase at a constant temperature was linear because of the influence of shear on the coagulation of the denatured enzyme. The addition of 0.01 g L−1of polyethylene glycol (MW 20,000) blocked this coagulation reaction, thereby completely inhibiting the shear‐induced inactivation of dextransucrase at 30°C. © 1993 John Wil

ISSN:0006-3592    

DOI:10.1002/bit.260420907

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1993

数据来源: WILEY

摘要:

AbstractThis article is concerned with the development of a model to maximize the overall yield of isolation of antibiotics recovered from fermented broths by the so‐called direct precipitation method. In this process, as in most antibiotics isolation processes, a second filtration of the semiexhaust mycellium from the first filtration, after slurrying it in water at the suitable pH, is required. The maximization of the overall yield of isolation implies the use of an optimal amount of water, measured as a volume to fermented broth mass ratio, which can be calculated by using the model derived here. The model also allows for the calculation of the partial and overall yields of the isolation process, and its validity is demonstrated by its ability to describe reasonably well the isolation data of two different tetracycline‐fermented broths produced according to two different technologies.The application of the model requires only the knowledge of easily obtainable fermented broth parameters and is illustrated for two different types of tetracycline‐fermented broths. Although the model had been derived for the optimization of the overall yield of isolation of antibiotics recovered by direct precipitation, it can easily be adapted to be used for the optimization of the overall yield of isolation of antibiotics recovered by other isolation processes. © 1993 John Wiley&Son

ISSN:0006-3592    

DOI:10.1002/bit.260420908

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1993

数据来源: WILEY

摘要:

AbstractThe physiology of a recombinant Chinese hamster ovary cell line in glucose‐limited chemostat culture was studied over a range of dilution rates (D= 0.008 to 0.20 h−1). The specific growth rate (μ) deviated fromDat low dilution rates due to an increased specific death rate. Extrapolation of these data suggested a minimum specific growth rate of 0.011 h−1(μmax= 0.025 h−1) The metabolism at each steady state was characterized by determining the metabolic quotients for glucose, lactate, ammonia, amino acids, and interferon‐γ (IFN‐γ). The specific rate of glucose uptake increased linearly with μ, and the saturation constant for glucose (Ks) was calculated to be 59.6 μM. There was a linear increase in the rate of lactate production with a higher yield of lactate from glucose at high growth rates. The decline in the rate of production of lactate, alanine, and serine at low growth rate was consistent with the limitation of the glycolytic pathway by glucose. The specific rate of IFN‐γ production increased with μ in a manner indicative of a growth‐related product. Despite changes in the IFN‐γ production rate and cell physiology, the pattern of IFN‐γ glycosylation was similar at all except the lowest growth rates where there was increased production of nonglycosylated IFN

ISSN:0006-3592    

DOI:10.1002/bit.260420909

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1993

数据来源: WILEY

摘要:

AbstractIn this report, we show that it is possible to separate valuable proteins from egg‐white using a ProductivTMCM ion‐exchanger column operated at flow rates significantly higher than those than can be achieved using traditional particulate adsorbents. In the approach taken, sample pretreatment is restricted to a simple dilution of the egg‐white, which can then be applied to the column at superficial velocities (Vs) of up to 13.8 m/h. Under a loading of 220 mg total protein per milliliter of ion‐exchanger, the resolution (Rs) between the eluted conalbumin and lysozyme fractions was found to be almost constant during nine consecutive adsorption/desorption cycles. For all nine consecutive batches, the column average adsorption capacity was greater than 30 mg/mL, with 90% recovery of adsorbed protein being achieved in each run. The overall productivity achieved was 12.6 kg/m3h for lysozyme and 31.2 kg/m3h for conalbumin. © 1993 John Wiley&S

ISSN:0006-3592    

DOI:10.1002/bit.260420910

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1993

数据来源: WILEY

摘要:

AbstractThis article deals with the production by fermentation of a mycostatic and aromatic food additive based on propionic acid. Membrane bioreactors have been used from laboratory scale up to pilot and industrial production plants. Due to the high cell densities achieved by the sequential recycling mode of operation, a mixed acids solution was rapidly produced from whey permeate. The sterile fermented broth obtained was subsequently concentrated at different levels by evaporation and spray drying according to the projected use. ConcentratedPropionibacteriumcells (200 g · L−1DW) were obtained from the process by periodic bleeds and could be used to good effect as cheese starters, silage preservatives, or probiotics. Propionic acid concentrations from 30 to 40 g · L−1were easily achieved with no residual lactose. The highest volumetric productivity was 1.6 g · L−1· h−1for total acid and 1.2 g · L−1· h−1for propionic acid with a specific productivity of 0.035 h−1. © 1993

ISSN:0006-3592    

DOI:10.1002/bit.260420911

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1993

数据来源: WILEY