Date of Award

1981

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Abstract

The specific activities of phosphofructokinase, aldolase, and pyruvate kinase was diminished in crude extracts from skeletal muscle of streptozotocin diabetic rats, whereas the activities of glucosephosphate isomerase and phosphoglucomutase were not changed. The decreased phosphofructokinase activity was shown to be independent of allosteric control, and it could be reversed by insulin treatment. A rapid and mild method for the purification of phosphofructokinase from rat skeletal muscle were developed. The partially purified enzyme from muscle of normal and streptozotocin diabetic rats showed identical Michaelis constants for ATP and equal sensitivity to inhibition by excess of ATP. Extracts of quick-frozen muscle from diabetic rats had higher levels of citrate, and lower levels of fructose 1,6-bisphosphate and glucose 1,6-bisphosphate. The levels of fructose 6-phosphate, glucose 6-phosphate, ATP, ADP, and AMP were the same for the two groups. These data suggest that phosphofructokinase activity is decreased in muscle of diabetic animals due to (a) a decrease in extractable activity, when assayed under optimum conditions, and (b) an unfavorable change in the levels of some of its allosteric modulators. A precise and reproducible gas chromatographic procedure for the determination of anomeric composition of glucose in biological fluids was developed. When applied to blood, peaks of the silylated glucose anomers were well resolved and were the only two major peaks found in blood. Blood glucose of normal and alloxan-diabetic rats, fed or fasted, was found statistically to have an anomeric composition slightly favoring the (beta) anomer over the thermodynamic equilibrium composition in water (39.1 (+OR-) 0.6% (alpha)- and 60.9 (+OR-) 0.6% (beta)-). The anomeric composition of glucose in plasma as well as in whole blood remained close to equilibrium after the administration of insulin to normal or diabetic rats. However, there was a statistically noticeable decrease in the percentage of blood (beta)-glucose as a result of insulin treatment of normal animals. A spin echo nuclear magnetic resonance technique for the non-invasive measurement of glycolytic activity in red blood cells from normal and diabetic rats was used. No differences were seen between the two groups. Proton magnetic resonance spectra of serum from normal and diabetic rats were also similar.

Pages

183

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