The terminal peptides of insulin

Abstract

While this work was in progress an outline of a fluorimetric method for the determination of Nmethyl-2-pyridone-5-carboxylamide in urine was reported by Rosen, Perlzweig & Handler (1948). The authors considered that their values for normal urines (5.0-13.2 mg./day) may have been somewhat low, but it is obvious that they are of the same order as those obtained by the present colorimetric method (see Table 1, and also Dean & Holman, 1949). SUMMARY 1. Methods are described for the determination of N-methyl-2-pyridone-5-carboxylamide and Nmethyl-2-pyridone-3-carboxylamideinhuman urine. The methods are based on the nitration of the compounds, after isolation from urine by adsorption on Lloyd's reagent followed by extraction with chloroform, and on their colorimetric determination by means of the yellow colours given by the nitro derivatives in alkaline solution.2. The accuracy and specificity of the methods is discussed.3. Results are shown which indicate that the 2:3amide is of no significance in nicotinic acid metabolism, but that the 2:5-amide is an important metabolite. The 2:5-amide was excreted by two normal adults in amounts of the order of 7 mg./24 hr. After the ingestion of 500 mg. ofnicotinamide, the urinary excretion of the 2:5-amide increased greatly, 47-57 % of the dose being excreted in 72 hr. as the latter compound.4. It was possible to account for 73-80 % of an orally administered dose of 500 mg. of nicotinamide in the urine of two adult subjects as N-methyl-2pyridone-5-carboxylamide, N-methylnicotinamide and the total acid-hydrolysable derivatives of nicotinic acid.

I. Denaturation of enzymes by the direct action of ionization radiation. II. Enzymically active components of the crystalline chymotrypsins

“…Sulfhydryl and disulfide analyses were performed by a modification of the metho~s of Boyer (16) These data are presented in Figure 1. The log percent r_emaining specific activity is plotted versus dose in eV.…”

Limited Hydrolysis of Ricin D with Alkaline Protease fromBacillus subtilis

“…2 ) Digestion of ricin D with nagarse (purchased from Nagase Inc., Osaka) was carried out at pH 8.0 and 30°C and the N-terminal amino acid of the digest was , determined by FDNB-method. 3 ) Polyacrylamide gel electrophoresis was performed in Tris-glycine buffer (pH 8.3) with a gel concentration of 7.5 % by the method of Orstein. 4 ) Cytoagglutinating activity was measured with sarcoma 180 ascites tumor cells and the toxicity was determined by intraperitomeal injection into purebred mice weighing 20 ~ 25 g as previously described.…”

Purification and Properties of an Extracellular Proteinase of Psychrophilic Escherichia freundii

“…The analysis was made by the method of Sanger [20]. I n order to prevent autolysis, the formic acidinactivated enzyme preparation was used for the determination of the NH,-terminal amino acid of the enzyme and was prepared in the following manner.…”