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Biomedical subjects

A J Daugulis

Publications and source records attributed to A J Daugulis.

30 records · Page 2Linked to original sources
PubMed1994

Integrated fermentation and recovery processes.

The application of integrated fermentation and product recovery continues to be improved for products such as ethanol, acetone and butanol. An extractive fermentation process for ethanol has now been scaled up and is approaching commercialization. Integrated fermentation process design is expanding into higher value materials, such as amino acids, plant cell products and antibodies. The general concept of selective partitioning of molecules between an aqueous and second (organic) phase has also been successfully applied to controlled substrate addition.

1-Butanol↗
PubMed1993

Importance of enzyme and solvent physical properties for the biocompatibility relationship of alpha-amino acid ester hydrolase.

The enzyme alpha-amino acid ester hydrolase was used with a variety of organic solvents to further explore the relationships between biocompatibility and solvent physical properties. Biocompatibility was shown to be system-specific; that is, the infection point of the characteristic log P trend (approximately 1.5) was specific to the enzyme employed, while the biocompatibility of solvents that were exceptions to the trend was dependent on the extent of agitation. In addition, phase toxic solvents all possessed a high interfacial tension with water. Finally, while the source of the enzyme may be significant, the extent of purification does not appear to be a factor in biocompatibility.

Biotransformation↗
PubMed1993

Polysaccharide microcapsules and macroporous beads for enhanced chromatographic separation.

One recent solution to the diffusion problem found in conventional chromatographic separation is the use of dual porosity beads in which large pores allow for convective flow and smaller pores allow for molecular diffusion. In our studies, dual porosity beads were prepared from carrageenan using an emulsion method. Effects of polymer type, polymer concentration, toluene content, gelling temperature, and stirring speed on the structure of porous beads were investigated. In an alternative approach, one of the interacting molecules in affinity chromatography, can be entrapped within semipermeable microcapsules. This has the advantage of increased adsorption capacities. Using blue dextran and albumin as a model system, alginate-chitosan capsules, containing blue dextran, were employed in the recovery of albumin from a saline solution.

Alginates↗
PubMed1992

Recombinant protein production in insect cell cultures infected with a temperature-sensitive baculovirus.

Spodoptera frugiperda (IPLB-SF-21) insect cells were grown in shake-flasks and infected with a temperature-sensitive baculovirus to express the gene of chloramphenicol acetyl transferase (CAT) in serum-free medium (SF-900) and two serum-supplemented media (IPL-41 and Grace's). In temperature-shift experiments (cell growth at 33 degrees C followed by virus replication at 27 degrees C 3-4 days later), virus and CAT production were much poorer in the serum-free medium than in serum-supplemented media, though cell growth was virtually the same in the different media tested. In all the three media, highest virus and CAT titers were obtained at the lowest MOI (multiplicity of infection 0.02). This result is contrary to that obtained in constant-temperature culture (27 degrees C for both cell growth and virus replication). Virus and CAT production was greatly improved when the entire culture was run at constant temperature. It appeared that infected cells were severely damaged at 33 degrees C (6 degrees C above the optimal 27 degrees C), resulting in little or no virus and protein production. As a result of these temperature-shift experiments, a larger-scale (14 1 air-lift bioreactor) serum-free culture of Sf-9 insect cells was conducted at constant temperature (27 degrees C) to produce recombinant protein (beta-galactosidase). A cell density as high as 1 x 10(7) cells.ml-1, and a beta-gal concentration of up to 104,000 unit.ml-1 were achieved.

Animals↗
PubMed1991

Integrated product formation and recovery.

As continues to be demonstrated, the in situ recovery of selected products froma bioreactor can have a significant positive impact on production. The strategies that are focused on here are: aqueous two-phase biocatalysis; non-aqueous biocatalysis; and membrane-enhanced biocatalysis. Additional fundamental understanding of molecular partitioning and biocatalytic activity in these environments will facilitate the rational selection of the components involved in these processing strategies.

Biotechnology↗
PubMedDate not supplied

Solvent selection strategies for extractive biocatalysis.

This report follows the development of systematic solvent screening strategies for the identification of superior pure solvents and introduces techniques for the identification of effective coextractants. Specifically, methods to predict the biocompatibility and extractant capability of solvents are discussed. Biocompatibility is predicted by using heuristic data or the correlations between bioactivity and the logarithm of the partition coefficient of the solvent or the concentration of solvent in the cell membrane. A computer program, known as the extractant screening program or ESP, has been developed to effectively predict the behavior of virtually any product in any solvent/aqueous system. It is demonstrated that a biocompatible yet poor solvent can be mixed with a toxic solvent that has better extractant properties to yield a mixture with improved solvent characteristics that is still biocompatible. The fact that solvents do not mix in an ideal manner is exploited by using ESP to identify solvent mixtures that are still biocompatible at relatively high concentrations of toxic solvent.

Animals↗
PubMedDate not supplied

Recombinant beta-galactosidase production in serum-free medium by insect cells in a 14-L airlift bioreactor.

Spodoptera frugiperda (Sf9) insect cells were successfully cultured in serum-free medium in a 14-L airlift bioreactor. Cell densities as high as 1 x 10(7) cells/mL were achieved with specific growth rates of approximately 0.0286 h-1 (doubling time of 24 h). This system was also used to demonstrate the expression of a reported gene, beta-galactosidase (beta-gal), when cells were infected with a recombinant baculovirus. Approximately 0.33 mg of beta-gal/mL (i.e., 104,000 units/mL) of medium were obtained at the 14-L scale, while about 0.95 mg of beta-gal/mL (i.e., 285,000 units/mL) of medium were obtained in small-scale shaker flasks. The difference was attributed to a suboptimal infection in the large scale. Specific oxygen consumption rates decreased from 5.58 x 10(-17) mol O2/cell.s in early exponential growth to 3.13 x 10(-17) mol O2/cell.s at 3 days post-infection.

Animals↗
PubMedDate not supplied

Protective effects of polymer additives on animal cells exposed to rapidly falling liquid films.

The protective effects of polymer additives on insect cells against fluid mechanical damage was investigated in a falling film-flow device. The falling liquid film creates rapidly moving air-liquid interfaces and high fluid shear stress, mimicking the characteristics of a bursting bubble in aerated cell culture. The additives tested included a group of surface-active polymers, (i.e., Pluronic F68, poly(ethylene glycol)s, and Tween 80) and a group of viscosity-enhancing polymers (i.e., dextrans, methyl-cellulose, and (carboxymethyl)cellulose). We found that methylcellulose, which was previously considered a viscosity-enhancing polymer, actually had significant surface-active properties. All of the surface-active polymers exhibited significant protective effects, with Pluronic F68 and the higher molecular weight poly(ethylene glycol), PEG 20M, providing the best protection. In contrast, the viscosity-enhancing polymers, with the exception of methylcellulose, showed little or no protection for insect cells in the film flow. All of the protective polymers had surface-active properties, even though some of them did not change the surface tension in the actual insect cell medium. There was no correlation between the protective effect and the changes in liquid viscosity and surface tension due to the polymer additives. The level of protection was shown to be dependent upon the type of polymer, its concentration in the culture medium, and the polymer molecular weight. We concluded that the mechanism of protection of these surface-active polymers was through interaction of the polymer molecules with the cell plasma membranes: a fast-acting biological mechanism.

Animals↗
PubMedDate not supplied

Expanded application of a two-phase partitioning bioreactor through strain development and new feeding strategies.

This research demonstrated the microbial treatment of concentrated phenol wastes using a two-phase partitioning bioreactor (TPPB). TPPBs are characterized by a cell-containing aqueous phase and an immiscible and biocompatible organic phase that partitions toxic substrates to the cells on the basis of their metabolic demand and the thermodynamic equilibrium of the system. Process limitations imposed by the capability of wild-type Pseudomonas putida ATCC 11172 to utilize long chain alcohols were addressed by strain modification (transposon mutagenesis) to eliminate this undesirable biochemical characteristic, enabling use of a range of previously bioavailable organics as delivery solvents. Degradation of phenol in a system with the modified strain as catalyst and industrial grade Adol 85 NF (primarily oleyl alcohol) as the solvent was demonstrated, with the system ultimately degrading 36 g of phenol within 38 h. Volumetric phenol consumption rates by wild type P. putida ATCC 11172 and the genetically modified derivative revealed equivalent phenol degrading capabilities (0.49 g/L x h vs 0.47 g/L x h respectively, in paired fermentations), with the latter presenting a more efficient remediation option due to decreased solvent losses arising from the modified strain's forced inability to consume the delivery solvent as a substrate. Two feeding strategies and system configurations were evaluated to expand practical applications of TPPB technology. The ability to operate with a lower solvent ratio over extended periods revealed potential for long-term application of TPPB to the treatment of large masses of phenol while minimizing solvent costs. Repeated recovery of 99% of phenol from concentrated phenol solutions and subsequent treatment within a TPPB scheme demonstrated applicability of the approach to the remediation of highly contaminated "effluents" as well as large masses of bulk phenol. Operation of the TPPB system in a dispersed manner, rather than as two distinct phases, resulted in volumetric consumption rates similar to those previously achieved only in systems operated with enriched air.

Bioreactors↗
PubMedDate not supplied

Simultaneous biodegradation of benzene, toluene, and p-xylene in a two-phase partitioning bioreactor: concept demonstration and practical application.

In this work, a mixture of benzene, toluene, and p-xylene was simultaneously biodegraded by Pseudomonas sp. ATCC 55595 in a two-phase partitioning bioreactor. This bioreactor consisted of a 1-L cell-containing aqueous medium phase and a 500-mL immiscible organic phase. The organic solvent systematically selected for use in the bioreactor was Adol 85 NF, an industrial-grade, biocompatible solvent. In the first of three experiments, the organic phase was loaded with 2.0 g of benzene, 10.15 g of toluene, and 2.1 g of p-xylene, which partitioned into the aqueous phase at concentrations of 25, 50, and 8 mg/L, respectively. The system ultimately biodegraded all of the substrates within 144 h. During the rapid growth phase of this fermentation, the cells were oxygen-limited. This fermentation was therefore repeated using an enriched air supply to remove the oxygen limitation. The use of enriched air ultimately reduced the length of the fermentation to 108 h, thereby improving the overall volumetric consumption rates. Finally, 500 mL of Adol were used to recover 2.0 g of benzene, 10.15 g of toluene, and 2.1 g of p-xylene from silica sand that was contaminated as part of a simulated soil "spill". The solvent washing procedure was able to recover greater than 99% of each compound from the contaminated soil. The Adol was then transferred to the two-phase bioreactor to permit biological treatment of the BTX contaminants. This process was repeated when the initial BTX load had been consumed almost to exhaustion, and the solvent was able to recover the contaminants at greater than 99% efficiency once again. The system was ultimately able to degrade 4.0 g of benzene, 20.2 g of toluene, and 4.2 g of p-xylene within 144 h. These results represent an unprecedented level of BTX degradation and illustrate a potential practical application for this novel biotechnology.

Benzene↗