Includes bibliographical references. / Thesis (Ph.D.)--Massachusetts Institute of Technology, Dept. of Nuclear Engineering, 1980. / by Richard Keith Lester. / Ph.D.

Indiana University-Purdue University Indianapolis (IUPUI) / This thesis describes separations performed by Pressurized Planar Electrochromatography (PPEC), which is a chromatographic method developed at IUPUI. In PPEC the mobile phase is driven by electroosmotic flow, while the system is pressurized to allow temperature control. This results in a highly efficient chromatographic system that has several attractive attributes including the ability to separate multiple samples simultaneously. The first three chapters of the thesis describe the relationship of PPEC to other forms of chromatography, the theoretical background of PPEC, the PPEC apparatus, including the plate holders used, and the different manipulations involved in preparing a plate for a PPEC run. The fourth chapter describes two short studies. The first demonstrates that a very fast separation of steroids on a high efficiency sorbent layer can be effected by PPEC. This is illustrated by the separation of six steroids in three minutes on a Superspher layer, with an efficiency of over 100,000 plates per meter. The second study attempted to improve the efficiency of separation by imposing a temperature gradient. The study was not successful, possibly due to Joule heating within the layer overriding the temperature gradient. The final chapter of the thesis describes two different studies on separating peptides by PPEC. The first study was performed on a bonded C18 sorbent layer that was treated with Brij-35, which is a non-ionic surfactant that prevents irreversible adsorption of the peptides to the sorbent surface while allowing electroosmotic flow. The variables involved in preparing the plates by soaking in a Brij-35 solution were investigated as well as the variables for PPEC (temperature, pressure, electrical potential, and mobile phase composition and pH). It was possible to separate six peptides in eight minutes using this approach. The second study used monolithic sorbent layers prepared by Dr. Frantisek Svec of Lawrence Berkeley National Laboratory. Separations were by conventional PPEC on charged monoliths and by electrophoresis on neutral monoliths. The same variables for PPEC, listed in the above paragraph, were investigated for the monolith study. It was possible to separate six peptides in two minutes on neutral monoliths and in one minute on negatively charged monoliths.

Typically, computational fluid dynamic (CFD) solutions produce large amounts of data that can be used for analysis. The enormous amount of data produces new challenges for effective exploration. The prototype system EVITA, based on ranked access of application-specific regions of interest, provides an effective tool for this purpose. Automated feature detection techniques are needed to identify the features in the dataset. Automated techniques for detecting shocks, expansion regions, vortices, separation lines, and attachment lines have already been developed. A new approach for identifying the regions of flow separation is proposed. This technique assumes that each pair of separation and attachment lines has a vortex core associated with it. It is based on the velocity field in the plane perpendicular to the vortex core. The present work describes these methods along with the results obtained.

The present study addresses a systematic evaluation of the effect of crosslinking via terminal maleimide units on the gas transport properties through a dense polymer film of amorphous Poly(Arylene Ether Ketones) or PEK. The methodology for the preparation of crosslinked maleimide terminated PEK (or MIPEK) films from several blend ratios of different molecular weights of MIPEK is discussed in detail. The primary focus of this work has been to investigate the influence of different degrees of crosslinking on gas permeabilities and selectivities, with an objective of producing a highly selective membrane. It has been demonstrated that crosslinking markedly decreases the permeation of methane, without changing the values for molecules such as helium. / Master of Science

Electrophoresis is a widely used analytical technique in the medical and biotechnology industries. It can provide for the determination of thousands of individual compounds on this small scale. The operating conditions are quite conducive to use with the delicate products of genetic engineering. Due to other complexities associated with scale-up, the process is not widely used on a large scale. A novel recycling electrophoretic instrument was investigated as a preparative protein separation device. The process occurs in a thin film of liquid between two flat plates, in a direction perpendicular to the flow. This device was unique in its use of a relatively high flowrate, and recycling of the process fluid as a method for increasing residence time. The apparatus was operated in three modes, isoelectric focusing (IEF), zone electrophoresis (ZE) and isotachophoresis (ITP). For use in the ITP mode, a computer was used for data acquisition and control functions. Model systems included monoclonal antibodies and lentil lectins.

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An attempt to develop a short and reliable method of caldesmon purification led to the development of three procedures of caldesmon purification. The first method was seldom used because of its low yield and the lack of caldesmon endogenous kinase activity. However, it allowed us to purify MLCK (myosin light chain kinase). The second and third methods gave respectively, a caldesmon sample with and without kinase activity. We were able to localize the endogenous kinase in the 0-30% ammonium sulfate precipitated DEAE pellet but we were unsuccessful at purifying the kinase to homogeneity. We found that caldesmon can also be phosphorylated by rat brain Ca²⁺-calmodulin-dependent kinase II at sites identical to those of caldesmon endogenous kinase but different to those of kinase C. In addition, caldesmon and its endogenous kinase are two different proteins. Furthermore, our study of caldesmon inhibition of actomyosin ATPase activity showed that further research needs to be done to refute F-actin bundling process as a possible cause of caldesmon inhibition of actomyosin ATPase activity. In addition, our studies of caldesmon inhibition of HMM and S-1 ATPase activity suggest that S-2 might be partially involved in the inhibition mechanism. Finally, caldesmon did not affect the 6S-10S transition of myosin conformation and since caldesmon cannot compete against higher affinity calmodulin-binding protein such as MLCK thus, the flip-flop theory is untenable.

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