522 BIOCHEMICAL RESEARCH FOUNDATION. [J. F. I.

found to be inhibited by normal serum, they were inhibited by specific immune serum. This specificity was quite sharp, immune serum to C. histolyticum inhibiting practically com- pletely the proteolytic enzymes of this organism, but not those of C. welchii, C. sporogenes; or even of a Rough strain of C. histolyticum. Since it precipitated with the globulin fraction of the serum, and was so sharply specific, the inhibiting factor in this immune serum was probably antibody.

Synthesis of Organic Compounds Containing Radioactive %&U.-HERBERT K. ALBER. (JOURNAL OF THE FRANKLIN INSTITUTE, 228: 177, 1939.) Recently R. G. Franklin (..%ie?zce, 89: 298 (1939) ; JOURN. FRANKLIN INST., 227: 722, 724 (1939) ) discussed the advantages of using compounds labelled with radioactive sulfur atoms in biochemical investi- gations, especially in tracing the course of sulfur compounds through the animal body and clarifying the role of certain organs in the highly important problems of sulfur metabolism. By means of a strictly biological method glutathione*, i.e. glutathione containing S35, was synthesized. Yeast was grown in a medium containing MgS*04 as the only source of sulfur, the glutathione * formed being isolated through the cuprous salt.

This in viva synthesis has now been supplemented by the in vitro synthesis of organic sulfur compounds* which are intermediates in the preparation of the biologically important sulfanilamide”. This work, because of the small amount of S* available (55 mg.), is an excellent example of a case in which the application of microtechniques is essential to organic synthesis. All operations carried out on a microscale with S* were duplicated by similar syntheses, on the same scale, with ordinary S. This procedure permitted efficient comparisons of physical constants and yields to be made.

S35 was obtained by bombarding CC14 with neutrons accelerated by the cyclotron of the Biochemical Research Foundation; I mg. of purified sulfur containing S35 gave 405 counts per minute, all activity measurements being carried out with the Geiger counter mentioned by Franklin.

I -Chloro-4-nitrobenzene was treated with NazS and S” in a micro container fitted with a reflux condenser, IOO mg. of crude

Oct., 1939.1 BIOCHEMICAL RESEARCH FOUNDATION. 523

material* being obtained (65 per cent. yield). On recrystal- lization from acetone and glacial acetic acid three fractions of 4,4’-dinitrodiphenyl disulfide” were obtained. Each frac- tion was tested for its purity, micro melting point on the Kofler hot stage, elementary composition, etc. The second fraction was the most active and gave micro melting points of 140", 164', and 178" to 179" C. as compared with the melting points of 134O, 164” to 165”, and 178” to 179’ C. given by the inactive compound. It is very probable that three crystal modifications of this compound exist, which may account for the varying melting points reported in the literature.

The radioactive measurements on the 4,4’-dinitrodiphenyl disulfide* were compared with those on the purified S* sample. As is known from studies on other radioactive elements, the ratio between radioactive and ordinary sulfur in the original active sulfur sample remains unchanged in the organic molecule obtained by synthesis. Thus, the ratio of the ob- served activities was I : 1.97 and I : 1.01 respectively, which is in good agreement with the calculated ratio of I : 2 and I : I respectively.

The remaining 4,4’-dinitrodiphenyl disulfide” (20 mg.), was treated in 90 per cent. acetic acid with chlorine gas and the resulting crude p-nitro-benzenesulfonyl chloride* (99 per cent. yield) was purified by repeated recrystallization from benzene-petroleum ether (55 per cent. yield). The micro melting point was 78.5-79” C. as compared with the literature value of 80” C. The elementary analysis confirmed the identity of the compound.

The radioactive measurements on the p-nitrobenzene- sulfonyl chloride* were compared with those on the S* and indicated a ratio of I : 2.4 instead of the calculated I : 2.0. The same results were obtained on comparing the two active sulfur compounds synthesized.

Due to scarcity of material the further steps in the syn- thesis to sulfanilamide” could not be carried out, but the possibility of synthesizing organic compounds containing S* in vitro has been clearly demonstrated even with the small amounts of S* available.