Somogyi reagent

Fig. 7. —Isolation of Endo-D-galacturonanase on a Column of Cross-linked Pectic Acid.210 [The Column (1.4 cm X 12 cm) was equilibrated with 0.1 M acetate buffer (pH 4.2). The first peak was eluted with the equilibrating buffer. Endo-D-galacturonanase was eluted with 0.1 M acetate buffer (pH 6.0). —, D-galacturonanase activity in 1 ml of eluate per min, determined by Somogyi reagent. The activity is expressed as change in absorbance at 530 nm, —, absorbance at 280 nm.]...

The Somogyi-Nelson assay (see Alternate Protocol 1) requires 30 min for the preparation of the low-alkalinity copper reagent and the same amount of time for the preparation of the arsenomolybdate reagent. An additional 24 hr should be considered for the 37°C incubation when preparing the arsenomolybdate reagent. Therefore, a total of 26 hr is necessary from the beginning of the experiments to obtaining the results. 

Somogyi, M. 1937. A reagent for the copper-io-dometric determination of very small amounts of sugar. J. Biol. Chem. 117 771-776. 

Somogyi s Copper Reagent Dissolve 14.0 g of anhydrous dibasic sodium phosphate and 20.0 g of potassium sodium tartrate tetrahydrate into 250 mL of water. Add 60.0 g of 1 M sodium hydroxide solution. Dissolve 4.0 g of cupric sulfate pentahydrate into 25 mL of water. Add this solution to the tartrate solution. Add 90.0 g of anhydrous sodium sulfate while stirring. Dilute the final solution to 500 mL. 

Procedure Transfer 1.0-mL aliquots of Pullulan Substrate to separate 15- x 150-mm test tubes. Insert a one-hole stopper in each tube, and equilibrate for 15 min in a 40° 0.1° water bath. At time equals zero and at 30-s intervals, add 1 mL of the respective samples, and mix. Exactly 30.0 min after addition of the samples, add 2.0 mL of Somogyi s Copper Reagent to each tube to terminate the reaction. Mix thoroughly, and allow the tubes to come to room temperature. To obtain sample blanks, add the Somogyi s Copper Reagent to the sample before adding the substrate. 

Prepare a standard curve by adding 2.0 mL of each glucose standard and 2.0 mL of Somogyi s Copper Reagent to a test tube, and mix. The blanks and Glucose Standards should not be incubated at 40°. 

Loosely stopper samples, blanks, and Glucose Standards containing Somogyi s Copper Reagent, and place them in a vigorously boiling water bath for exactly 25.0 min. Cool in an ice-water bath for approximately 1 min. 

Colorimetric methods Based on chemical reagents used Anthrone Clegg-anthrone Copper reduction Dinitrosalicylate Phenol/sulfuric acid Nelson-Somogyi Neocuproine Picric... 

Kim, Y. K., and Sakano, Y. (1996). Analyses of reducing sugars on a thin-layer chromatographic plate with modified Somogyi and Nelson reagents, and with copper bicin-choninate. BioscL BiotechnoL, Biochem. 60 594-597. 

The first coulometric titration is credited to Szebel-ledy and Somogyi, who, in 1938, used electrolysis to generate bromine which reacted stoichiometrically with the substance to be determined. The quantity of substance which reacted was computed by Faraday s law from the number of coulombs of electricity rather than from a volume of standard reagent. By keeping the current constant and measuring the time, the number of coulombs was easily calculated. 

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