Inhibition of Rabbit Muscle Enolase by Ascorbate and Fatty Acid Derivatives

We have investigated the role of vitamin C (ascorbate) in glycogen synthesis and glycolysis in rabbit muscle. Previous research has also shown that ascorbate (vitamin C) inhibits muscle phosphofructokinase-1 (PFK-1), and muscle lactate dehydrogenase (LDH), but does not inhibit the PFK-1 from liver or the LDH. We show that ascorbic acid also inhibits enolase, an enzyme needed for glycolysis. Ascorbate inhibits muscle enolase as effectively as it inhibits muscle LDH. Previous studies showed that fatty acid derivatives of ascorbate, L-ascorbyl 2,6-dipalmitate (AADP) and L-ascorbyl 6-stearate (AAS) were 30-times and 7-times more inhibitory to PFK-1, respectively, and were irreversible. Our studies with enolase show that AADP and AAS are also irreversible inhibitors.  The AADP is twice and AAS is half as inhibitory as ascorbate. Other fatty acid derivatives of ascorbate are currently being tested for enolase inhibition L-ascorbyl 2,6-dibutyrate, L-ascorbyl 6-palmitate. Previous studies showed that PFK-1 was protected from loss of activity due to ascorbate, dilution, fatty acid derivatives of ascorbate, and dilution was prevented by the aldolase, the enzyme following PFK-1 in glycolysis; we are testing whether of not enolase can protect similarly. We are also testing whether or not aldolase can prevent enolase losses of activity. Since most cancer cells depend solely on glycolysis as a source of energy, we believe that these studies have some relevance to chemotherapeutic pursuits.