Über die Bestimmung von d-Glutaminsäure in Tumorhydrolysaten mit Deuterium als Indicator. 9. Mitteilung über die Chemie der Tumoren

Abstract

Deuterium was detd. by the interferometer and falling drop methods. For the former method, which requires 400 mg. H2O, the portable Zeiss liquid interferometer, calibrated with standard mixtures of H2O. D2O, was used. For the latter method, 1 drop of the purified combustion H2O was allowed to fall through a tube containing o-fluorotoluene, which was in a thermostat at a temp. such that 1 drop H2O fell about 15 cm. in 180 sec. The D2O conc. in the sample was calculated by comparison with the rate of fall of standard mixtures of H2O . D2O. To prepare deuterium glutamic acid by the exchange reaction, 1 g. PtO2. H2O was reduced in 8 cc. D2O with deuterium gas, 6 g. 1- or d-glutamic acid and 6 cc. conc. HC1 added, the mixture melted and shaken at 95[degree] for 9 days. After purification of the product obtained, the deuterium content of the l-form was 7.4 atom %, and of the d-form, 7.5 atom %. To prepare deuterium glutamic acid by the Knoop reaction, 10.2 g. [alpha]-ketoglutaric acid were shaken with 2.45 g. palladium oxide, 122.5 cc. H2O and 32.9 cc. 25% NH3 for 12 hrs. under D2. After purification, the deuterium content of the glutamic acid obtained was 10.47 atom %. The tumor material was ground and 5 times its wt. of 0.9% NaCl and toluene added. The mixture was shaken, allowed to stand 24 hrs. in the ice box, centrifugalized, and, after washing with alcohol and ether, the insoluble material was dried over P2O5 in vacuo. The filtrate was treated with 5 vols. alcohol and the precipitate obtained washed and dried as above. The dried material plus 3 times its wt. of 37% HC1 was heated to gentle boiling for about 20 hrs. in an oil bath. The hydrolysate was cooled, an equal vol. H2O added and filtered. The filtrate was concd. in vacuo, and the brown syrup obtained dissolved in 10 vols. H2O. Humus impurities were removed with Cu2O, the soln. concd. and the syrup saturated at 0[degree] with HC1, the glutamic acid HC1 filtered off after 7 days and recrystallized. When the deuterium glutamic acid was added to the dried tumor material before hydrolysis, it was found that the degree of racemism of the acid by benign tumors was 3-4%; primary Flexner-Jobling carcinoma, 17% chemical tumors (methyl-cholanthrene, benzpyrene), 27%; human tumors with metastasis, 42-44%; Brown-Pearce with metastasis, 40-48%. Under the same conditions, preps. of normal tissues gave only negligible amts. of d-glutamic acid.