Biomedical subjects

T A Hatton

Publications and source records attributed to T A Hatton.

7 recordsLinked to original sources

PubMed1999

Cell death from bursting bubbles: role of cell attachment to rising bubbles in sparged reactors.

Bursting bubbles are thought to be the dominant cause of cell death in sparged animal or insect cell cultures. Cells that die during the bubble burst can come from three sources: cells suspended near the bubble; cells trapped in the bubble lamella; and cells that attached to the rising bubble. This article examines cell attachment to rising bubbles using a model in which cell attachment depends on cell radius, bubble radius, and cell-bubble attachment time. For bubble columns over 1 m in height and without protective additives, the model predicts significant attachment for 0.5- to 3-mm radius bubbles, but no significant attachment in the presence of protective additives. For bubble columns over 10 cm in height, and without protective additives, the model predicts significant attachment for 50- to 100-micron radius bubbles, but not all protective additives prevent attachment for these bubbles. The model is consistent with three sets of published data and with our experimental results. Using hybridoma cells, serum-free medium with antifoam, and 1.60 +/- 0.05 mm (standard error) radius bubbles, we measured death rates consistent with cell attachment to rising bubbles, as predicted by the model. With 1.40 +/- 0.05 mm (SE) radius bubbles and either 0.1% w/v Pluronic-F68 or 0.1% w/v methylcellulose added to the medium, we measured death rates consistent with no significant cell attachment to rising bubbles, as predicted by the model.

Animals↗

PubMed1998

Protein refolding by reversed micelles utilizing solid-liquid extraction technique.

This article reports that a reversed micellar solution is useful for refolding proteins directly from a solid source. The solubilization of denatured RNase A, which had been prepared by reprecipitation from the denaturant protein solution, into reversed micelles formulated with sodium di-2-ethylhexyl sulfosuccinate (AOT) has been investigated by a solid-liquid extraction system. This method is an alternative to the ordinary protein extraction in reversed micelles based on the liquid-liquid extraction. The solid-liquid extraction method was found to facilitate the solubilization of denatured proteins more efficiently in the reversed micellar media than the ordinary phase transfer method of liquid extraction. The refolding of denatured RNase A entrapped in reversed micelles was attained by adding a redox reagent (reduced and oxidized glutathion). Enzymatic activity of RNase A was gradually recovered with time in the reversed micelles. The denatured RNase A was completely refolded within 30 h. In addition, the efficiency of protein refolding was enhanced when reversed micelles were applied to denatured RNase A containing a higher protein concentration that, in the case of aqueous media, would lead to protein aggregation. The solid-liquid extraction technique using reversed micelles affords better scale-up advantages in the direct refolding process of insoluble protein aggregates.

Acetone↗

PubMed1998

Solvent replacement for green processing.

The implementation of the Montreal Protocol, the Clean Air Act, and the Pollution Prevention Act of 1990 has resulted in increased awareness of organic solvent use in chemical processing. The advances made in the search to find "green" replacements for traditional solvents are reviewed, with reference to solvent alternatives for cleaning, coatings, and chemical reaction and separation processes. The development of solvent databases and computational methods that aid in the selection and/or design of feasible or optimal environmentally benign solvent alternatives for specific applications is also discussed.

Air Pollution↗

PubMed1991

Recovery of biocatalysis products from reversed micellar reaction media: a preliminary evaluation of membrane extractors.

It has been demonstrated that ultrafiltration membranes of sufficiently low molecular weight cutoff can be used to retain reversed micelles and their hosted enzymes, while permitting the recovery of lipophilic products of enzymatically-catalysed, synthesis reactions in a stripping solution on the other side of the membrane. Calculations indicate that hollow fibre membranes having the same rejection characteristics and solvent resistance as the flat sheet membranes, will provide an attractive and efficient means for the recovery of these biosynthesis products; currently, such membrane modules are not available commercially.

Enzymes↗

PubMed1990

On protein partitioning in two-phase aqueous polymer systems.

The partitioning of proteins between the coexisting phases of two-phase aqueous polymer systems reflects an intricate and delicate balance of interactions between proteins, polymers, salts and water. Experimental investigations have suggested that a large number of factors influence protein partitioning, including the types of polymers, their molecular weight and concentration; the protein sizes, conformation and composition; salt type and concentration, and solution pH; and the presence of ligands attached to the polymer which may interact with surface sites of the protein. Complementary modelling attempts have been successful in illuminating several molecular-level mechanisms influencing protein partitioning using lattice-model techniques, viral expansions and a scaling-thermodynamic approach. In spite of these experimental and modelling approaches, many of the physical phenomena associated with these complex systems are not well understood. Notably, the precise nature of the protein-polymer interactions and the potent effect of inorganic salts on the partitioning of proteins in these systems remains poorly understood.

Methods↗

PubMedDate not supplied

Extraction behavior of hemoglobin using reversed micelles by dioleyl phosphoric acid.

A new surfactant, dioleyl phosphoric acid (DOLPA), has been applied to the extraction of hemoglobin using reversed micelles. The reversed micelles formed by DOLPA can easily extract hemoglobin from aqueous to reversed micellar solutions. DOLPA is the first surfactant to extract hemoglobin completely without using any cosurfactants. On the basis of the difference between DOLPA and AOT reversed micelles in the forward extraction behavior of hemoglobin, the nature of the interfacial complex that would be formed between surfactants and hemoglobin at the oil--water interface was found to be the dominant factor in determining the extraction efficiency of hemoglobin be reversed micelles. In addition, back-transfer studies of hemoglobin from the DOLPA reversed micelles were also carried out by the phase transfer method. It was found that hemoglobin, once dissolved into the DOLPA reversed micelles, is not transferred to a fresh aqueous solution even when the conditions are adjusted to not allow the forward transfer of hemoglobin. However, the addition of several kinds of alcohol drastically improved the yield in the back-transfer of hemoglobin. The efficiency in the back-transfer of hemoglobin strongly depends on the aqueous conditions that are in contact with the reversed micelles, such as pH, ionic strength, and alcohol concentration. A pH higher than the pI of hemoglobin, a salt concentration lower than that of the water pool, and the proper concentration of alcohol are required for the recovery aqueous phase to ensure the back-transfer of hemoglobin from the DOLPA reversed micelles.

Alcohols↗

PubMedDate not supplied

Extraction and activity of chymotrypsin using AOT-DOLPA mixed reversed micellar systems.

Novel reversed micellar solutions formulated with a mixture of AOT (dioctyl sulfosuccinate) and DOLPA (dioleyl phosphoric acid) show good potential for use in reversed micellar protein extraction operations. Chymotrypsin is easily extracted from an aqueous phase into organic isooctane containing 10 mM AOT and DOLPA in a 4:1 ratio. The extraction ability of the mixed reversed micelles of 10 mM was higher than that of 200 mM AOT alone. The results of extraction indicated that the AOT-DOLPA mixed reversed micelles are very useful for separating and enriching chymotrypsin. Back-extraction of chymotrypsin from the organic phase to a fresh aqueous phase is also accomplished by adding an alcohol to the organic phase. Although the back-transfer of chymotrypsin from the reversed micelles formed by AOT alone is very slow and difficult, in the AOT-DOLPA mixed reversed micelles, the back-extraction can be achieved completely by addition of 10% (v/v) isobutyl alcohol to the reversed micellar phase. The time to attain to the equilibrium of back-extraction was reduced from more than 24 to 2 h by adding the alcohol. On the basis of the activity data, the best composition of AOT and DOLPA was a 4:1 ratio and the total surfactant concentration was 10 mM. The activity of chymotrypsin recovered from the mixed reversed micelles was higher than that of the initial protein before the forward-transfer. This result means that the novel mixed reversed micellar solutions are useful not only in separation but also in purification of proteins.

Animals↗