Title: Dihydroxyacetone Phosphate, Fructose-l,6-diphosphate and d-Glyceraldehyde-3-phosphate
Abstract: This chapter elaborates the method for the enzymatic determination of dihydroxyacetone phosphate, fructose-1, 6-diphosphate, and D-glyceraldehyde-3-phosphate using glycerol-l-phosphate dehydrogenase, aldolase, and triosephosphate isomerase. The same indicator reaction can be used for the spectrophotometric determination of dihydroxyacetone phosphate, fructose diphosphate and glyceraldehyde phosphate. The method for dihydroxyacetone phosphate is based on the principle that glycerol-l-phosphate dehydrogenase (GDH) catalyzes the reduction of dihydroxyacetone phosphate (DAP) with reduced diphosphopyridine nucleotide (DPNH). The equilibrium constant of this reaction Kc is 1.78 × 1011 (moles/1)−1 at pH 0 and 25°C; even at pH 7 (Kc’ = 1.78 × 104 (moles/1)−1) the equilibrium lies far to the right so that dihydroxyacetone phosphate reacts quantitatively. With a measurable excess of DPNH and sufficient enzyme the reaction proceeds rapidly. The decrease in optical density at 340 or 334 (also 366) mμ because of the oxidation of DPNH is a measure of the reaction. In case of fructose-1,6-diphosphate, aldolase cleaves fructose-1,6-diphosphate (FDP) to give glyceraldehyde-3-phosphate (GAP) and dihydroxyacetone phosphate (DAP). GAP is isomerized to DAP by triosephosphate isomerase (TIM) and the DAP is reduced by DPNH and GDH.
Publication Year: 1965
Publication Date: 1965-01-01
Language: en
Type: book-chapter
Indexed In: ['crossref']
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Cited By Count: 40
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