Title page for ETD etd-1454132679612381


Type of Document Master's Thesis
Author Gardner, Tara Conti
URN etd-1454132679612381
Title Delipidation Treatments for Large-Scale Protein Purification Processing
Degree PhD
Department Chemical Engineering
Advisory Committee
Advisor Name Title
Conger, William L.
Davis, Richey M.
Velander, William H. Committee Chair
Keywords
  • delipidation
  • lipid removal
  • protein purification
  • transgenic milk
  • sephadex
  • TNBP
Date of Defense 1998-08-12
Availability unrestricted
Abstract
Triglycerides are the majority lipid component

of most biochemical mixtures and are virtually

water insoluble. Lipid removal is desired prior

to protein purification processing to decrease

nonspecific fouling of downstream

chromatographic matrices. Transgenic pig milk

was used as a model system to study

delipidation from therapeutic protein sources.

The majority of triglycerides was extracted

from stable lipid micelles and removed with a

method that can be incorporated in

downstream protein purification processing

without denaturing the target protein. An

efficient delipidation treatment used TNBP, a

non-polar solvent, to extract lipid micelles and

then phase transfer milk lipids into a

TNBP-swelled dextran particulate. A batch

incubation of a whey/TNBP mixture with

pre-swollen Sephadex LH-20 or

hydroxyalkoxypropyl dextran (HAPD) beads

at 4 C for 24 hours removed 67 + 2 %

(0.645 mg triglycerides/ml Sephadex LH-20)

and 71 o + 1 % (0.628 mg triglycerides/ml

HAPD) of the triglycerides present in the

skimmed transgenic whey, respectively. Fully

swollen beads removed 20% more

triglycerides than beads which were wetted

but not swollen in TNBP, indicating that a

larger phase volume and internal adsorption of

the lipids onto the Sephadex matrix dominates

over surface adsorption. Polyclonal ELISAs

indicated that 89 + 6% of the recombinant

human Protein C was still present in the

transgenic whey after this delipidation

treatment, indicating this treatment did not

denature or harm the target protein.

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