Date of Award

2001

Degree Type

Thesis

Degree Name

Master of Science (MS)

Department

Pharmaceutical Sciences

First Advisor

Yuzuru Shimizu

Abstract

Protein phosphatases play significant roles in signal transduction pathways that regulate cellular processes in response to external/internal stimuli. They are crucial for the growth, division, and differentiation of all organisms. Examples of cell functions involving reversible phosphorylation include ion transport, metabolism, cell cycle progression, developmental control, and stress responses. Serine/threoninespecific protein phosphatases are of particular interest in this study. There are two types of serine/threonine-specific protein phosphatases: type 1 & type 2. Both protein phosphatase 1 (PPl) and protein phosphatase 2A (PP2A) are inhibited by a structurally diverse group of natural toxins produced by marine organisms that cause diarrhetic shellfish poisoning. Recent research into diarrhetic shellfish poisoning has contributed to the understanding of some of the mechanism of actions of cathartics. It was learned that hydroxy acid moieties are essential for receptor binding of diarrhetic shellfish toxins, such as okadaic acid, to the protein phosphatase enzymes in the intestines which regulate ion channels. The loss of ion channel regulation in the intestines leads to an efflux of electrolytes and water, causing diarrhea. This study was undertaken in order to learn if four different crude cathartic plant drugs whose active components contain hydroxy-acid moieties, will act similarly to these diarrhetic shellfish toxins by inhibiting PP2A. First, two resin glycosides, jalapin and convolvulin, were isolated from Ipomoea purga and Pharbitis nil, of the Convolvulaceae family, respectively. After verifying their molecular structures by proton nuclear magnetic resonance (1H NMR) spectroscopy, they were tested against PP2A. Pure ricinoleic acid from Ricinis communis and podophyllotoxin from Podophyllum peltatum were purchased and also tested against PP2A. A fluorometric assay, recently developed in Dr. Shimizu's lab, was used to determine their activities toward protein phosphatase enzyme PP2A. The assay results indicated weak but significant inhibition of PP2A by these compounds. The doses of these crude drugs used to produce catharsis are six orders larger than the dose of okadaic acid which causes diarrhea in humans. Thus the weak activities of these compounds may be sufficient to account for the cathartic action of these drugs. The bioavailability of these compounds in the human body is unknown. Since the traditional standard dosages of these cathartics are too large to prepare as samples for a micro-titer assay due to their solubilities, they were not able to be assayed at higher concentrations at which more or total inhibition might take place. The possibility that these crude cathartic drugs inhibit PP2A can not be discounted based on the results of this study. It is also possible that these compounds may affect other enzymatic activities which were not tested. This matter remains open to question.

Share

COinS