The authors wish to express their indebtedness to Mrs. Barbara Bingham for fine technical assistance, and to Drs. Robert B. Loftfield and Mahlon B. Hoagland for helpful advice and criticism. * This is publication No. 1,000 of the Cancer Commission of Harvard University. This work was supported by grants from the American Cancer Society, Inc., the U. S. Atomic Energy Commission, and the U. S. Public Health Service. 1 Hecht, L. I., M. L. Stephenson, and P. C. Zamecnik, Biochim. et Biophys. Acta, 29,460 (1958). 2 Zachau, H. G., G. Acs, and F. Lipmann, these PROCEEDINGS, 44, 885 (1958). 3Preiss, J., P. Berg, E. J. Ofengand, F. H. Bergmann, and M. Dieckmann, these PROCEEDINGS, 45, 319 (1959). 4Hecht, L. I., M. L. Stephenson, and P. C. Zamecnik, these PROCEEDINGS, 45, 505 (1959). 5 Hoagland, M. B., M. L. Stephenson, J. F. Scott, L. I. Hecht, and P. C. Zamecnik, J. Biol. Chem., 231, 241 (1958). 6 Lipmann, F., W. C. Hulsmann, G. Hartmann, H. G. Boman, and G. Acs, J. Cell. Comp. Physiol., 54, 75 (1959). 7Smith, K. C., E. Cordes, and R. S. Schweet, Biochim. et Biophys. Acta, 33, 286 (1959). 8 Holley, R. W., and S. H. Merrill, J. Am. Chem. Soc., 81, 753 (1959). 9 Brown, G. L., A. V. W. Brown, and J. Gordon, in Brookhaven Symposia in Biology, No. 12, (1959). 10 Zamecnik, P. C., and M. L. Stephenson, Federation Proc., 19, 346 (1960). 11 Monier, R., M. L. Stephenson, and P. C. Zamecnik, Biochim. et Biophys. Acta (in press). 12 Warner, R. C, and P. Vaimberg, Federation Proc., 17, 331 (1958). 13 Ashbel, R., and A. M. Seligman, Endocrinology, 44, 565 (1959). 14 Kirby, K. S., Biochem. J., 64, 405 (1956). 11 Gierer, A., and G. Schramm, Nature, 177, 702 (1956). 16 Whitfeld, P. R.. Biochem. J., 58, 390 (1954). 17 Dixon, J. S., and D. Lipkin, Anal. Chem., 26, 1092 (1954). 16 Hecht, L. I., P. C. Zamecnik, M. L. Stephenson, and J. F. Scott, J. Biol. Chem., 233, 954 (1958). 19 Canellakis, E. S., and E. Herbert, these PROCEEDINGS, 46, 170 (1960). 20 Lehrer, G. M., and L. Ornstein, J. Biophys. Biochem. Cytel., 6, 399 (1959). 21 Barka, T., and T. Ornstein, J. Histochem. and Cytochem., 7, 385 (1959). 22 Webster, G., Federation Proc., 19, 345 (1960). 23 Holley, R. W., and B. P. Doctor, Federation Proc., 19, 348 (1960).

A NEW REACTION OF CYCLIC ALIPHATIC ANHYDRIDES* BY D. RITTENBERGt AND LAURA PONrICORVO DEPARTMENT OF , COLLEGE OF PHYSICIANS AND SURGEONS, COLUMBIA UNIVERSITY Communicated April 11, 1960 During the course of another investigation, it was found that the addition of catalytic amounts of to maleic anhydride induces a of some violence which yields and a black brittle residue. Further studies show that this reaction is a general one, occurring with most aliphatic cyclic anhydrides and with all tertiary tested except . Primary and secondary amines (e.g., a-naphthylamine, diethylamine, ) do not catalyze the reaction. The anhydrides and active amines tested are listed in Table 1. The reaction varies in rate with the various anhydrides; it is rapid Downloaded by guest on September 25, 2021 VOL. 46, 1960 CHEMISTRY: RITTENBERG AND PONTICORVO 823

TABLE 1 ANHYDRIDES WHICH YIELD CO2 ON TREATMENT WITH PYRIDINE Maleic fl-methylglutaconic Succinic Glutaric Adipic Citraconic Chloromaleic Sebacic BASES WHICH CATALYZE THE REACTION Triethylamine Tri-n-butylamine 5,6-Benzoquinoline Quinaldine Pyridine Collidine 2,6-Lutidine Quinaldine with maleic anhydride and slow with succinic. The has only the role of a catalyst because the product formed is free of . With most anhydrides the reaction occurs in two stages. In the first a red to red-brown color appears. This red color has been observed by Pfeiffer and Bottlerl on the addition of maleic anhydride to dimethylaniline. In contrast, the addition of a primary or secondary to these anhydrides, at room temperature or at the of the anhydride, yields either no color or a yellow one. The addition of triethylamine to maleic anhydride at room temperature pro- duces within 30 sec a sequence of colors which pass from yellow to red to purple to purple-black. The color sequence with triethylamine and has been observed by Barb,2 who offered it as a specific test for detection of tertiary aliphatic amines.3 The reddish-colored pigment usually formed initially is soluble in acetone, aceto- phenone, and halogenated hydrocarbons and remains unchanged in these for some hours. If water is added, the color fades. The reddish pigment can also be produced when the reaction between maleic anhydride and pyridine is carried out in acetone or chloroform solution. The second stage of the reaction is an exothermic . The rate of the reaction is determined by the amount of catalyst added and the temperature. Experimental.-In a typical experiment 4 ml of pyridine was added to 100 gm. of maleic anhydride. The mixture spontaneously warmed up (turning brownish-red in the process) and began to decarboxylate rapidly when the temperature reached 90°C. The temperature then rose rapidly to 160°C. On cooling, a spongy, friable black solid remained. The yield of carbon dioxide per mole of anhydride varies from compound to compound (see Table 2). TABLE 2 YIELD OF CO2 Temperature Pyridine of reaction C02 Anhydride mM ml 0C Evolved mM Maleic 2.07 0.04 245 1.81 Chloromaleic 1.97 0.05 200 1.92 Citraconic 2.23 0.05 200 1.03 0-Methylglutaconic 2.03 0.05 150-190 1.88 Succinic 4.0 0.20 210 0.69 2.51 (a) 200 1.06 2.21 (b) 245 0.91 Succinic 500 2.0 190-200 350 Adipic 3.0 0.05 200 1.58 (a) 0.05 ml of lutidine. (b) 0.10 ml of triethylamine. Downloaded by guest on September 25, 2021 824 ENGINEERING: BOELTER AND BRANCH PROC. N. A. S.

Anhydrous acetate will serve as the catalyst with maleic, chloro-maleic, and methylglutaconic anhydrides, giving yields of 0.54, 0.22, and 1.02 moles CO2 per mole of anhydride. The products of all these reactions are black and brown-black, are soluble in acetone, acetophenone, dioxane, and aqueous alkali and are insoluble in , toluene, , chloroform and carbon tetrachloride, and aqueous . The reac- tion can be carried on in solution in , diphenylmethane, acetone, and acetophenone. The product is heterogeneous since it can be fractionated by solution in acetone and precipitation by ether. The most soluble fraction is brown; the least soluble one tends to be black. Cyclopentylmaleic,4 dodecenylsuccinic,5 1,2-cyclohexanedicarboxylic,8 and cy- clohexene-1,2-dicarboxylic6 anhydrides do not produce CO2 on treatment with ter- tiary amines. Summary.-Many cyclic aliphatic anhydrides on treatment with catalytic amounts of a tertiary amine undergo a reaction with the evolution of CO2. The evolution of CO2 is preceded by the appearance of a red to purple intermediate. The products are brittle and amorphous, soluble in polar solvents and in alkali, in- soluble in nonpolar solvents and acid. * This research was supported by a grant from the American Cancer Society on recommendation of the Committee on Growth of the National Academy of Sciences-National Research Council (MET-33B) and by contracts to Columbia University from the Office of Naval Research, Depart- ment of the Navy (ONR 26602); from the Atomic Energy Commission (AT(30-1)1803), and from the National Institutes of Health (E-2839). Reproduction of this article in whole or in part is per- mitted for any purpose of the United States Government. t Part of the work described here was carried out while one of us (D. R.) was a guest at the Weizmann Institute of Science at Rehovoth, Israel. We are indebted to Dr. E. Katchalsky for many helpful discussions. I Pfeiffer, P., and T. Bottler, Ber., 51, 1819 (1918). 2 Barb, W. G., J. Chem. Soc., 1647 (1955). 3 Sass, S., J. J. Kaufman, A. D. Cardenas, and J. J. Martin, Anal. Chem., 30, 529 (1958). 4We are indebted to Dr. W. R. Vaughn of the University of Michigan for a sample of this anhydride. 6 Obtained from Allied Chemical and Dye Corp., Buffalo, N. Y. Used without further purifica- tion. 6Obtained from Brothers Chemical Co., Orange, N. J. Used without further purification.


COLLEGE OF ENGINEERING, UNIVERSITY OF CALIFORNIA AT LOS ANGELES, AND THOMPSON RAMO WOOLDRIDGE INC., LOS ANGELES Communicated by Augustus B. Kinzel, April 29, 1960 As a consequence of established trends in our contemporary society, more plan- ning is required and practiced in the United States with each passing year. Plan- ning is applied in many forms and areas of activity. It is therefore of growing importance that the American people have the opportunity to choose among the Downloaded by guest on September 25, 2021