3976
COMMUNICATIONS TO THE EDITOR
the conclusion has been drawn that this isotope effect will always have a value near unity, and this value has been used to deduce the relative acid strengths of the species HzO and D20.2bWe wish here to report a deuterium isotope effect on ratedetermining proton transfer from the solvated proton which is considerably greater than unity. Acid-catalyzed aromatic hydrogen exchange in 1,3,5-trimethoxybenzene is known to proceed by the two-step reaction sequence4
NHS-
,
acetylenedicnrboh) dirnrtliyl ldte
pha ph P?
,
Vol. 84
k1
PI1
H’Ar 3- H+(HnO),
/
)r H’ArH+ + nHnO kz
R’
Ph R’
\CO&H3
IIIa, R = Ph, R ’ = C02CH3 b, R’=Ph, R=CO,CHs
deep blue with antimony pentachloride. Trifluoroacetic acid produces a deep blue color with I and a fluorescent red color with 11. Addition of hydroxylic solvents immediately discharges these intense colors. Heating I with excess dimethyl acetylenedicarboxylate a t 160’ affords in 63% yield the dicarbomethoxyhexaphenylazulene, 111 (a or b) [dark greenish-blue crystals, m.p. 247-248’; calcd. for C6oHo6Oa:C, 85.69; H, 5.18; mol. wt., 700.8. Found: C, 85.54; H , 5.10; mol. wt., 664. Ultravioletvisible spectrum (chloroform) Xma, mp (log E ) : 265 (4.64), 339 (4.96), 634 (2.80). Infrared (KBr) : 3.27, 3.39, 5.75, 6.24, 6.67, 6.91, 7.84, 8.17, 9.25, 9.71, 13.09, 14.321.” Thus, the pentalene structure of I is consistent with: (1) the mode of formation (two routes), ( 2 ) the analytical results, (3) the unusual spectral properties, and (4) the unique two carbon ring-expansion to form an azulene. The author wishes to thank Mr. R. B. La Count for assistance in obtaining the spectral data. MELLONINSTITUTE EUGENELE GOFF PITTSBURGH 13, PENNSYLVANIA RECEIVED AUGUST13, 1962 THE ISOTOPE EFFECT ON ACID-CATALYZED SLOW PROTON TRANSFER IN DILUTE AQUEOUS SOLUTION’
Sir:
HAr slow
kz
ki
+ H’+(HzO)
(1)
The first forward step in this reaction is a slow proton transfer from catalyzing acid to substrate. The isotope effect on this step (klH~O/klDZ0) as well as that on the second step (kzH/kzD) can be obtained by performing the experiments (I), (11) and (111).
+ ~ H z J_ O + T’(H2O)n (2) D.4r + H + ( H Z O )J_ ~ DArH+ + nHiO HAr + D+(HzO), (3) TAr + D+(D20), TArD+ + nDz0 2
I . TAr 11. 111.
+ H+(HzO),
TArH+ HAr
DAr
+ T+(DzO),
(4)
These reactions are practically non-reversible, and their observed second order rate constants are related to the rate constants for the individual steps in the following way (5)
With the aid of a relationship between deuterium and tritium isotope effect^,^ equations of the form of ( 5 ) for reactions I, I1 and 111 can be solved for
(7)
the rate ratios klHz0/klD20 and k 2 H / k 2 D . (This treatment neglects secondary isotope effects in the aromatic molecule, but there are both empiricale and t h e ~ r e t i c areasons l~~~~ to~expect these to be absent.) For 1,3,5-trimethoxybenzene in 0.05 M HCIOP, the data (Table I) give the values: k l H ~ O / k l D ~ o= 2.93 u = 0.07, and kzH/k2* = 6.68, u = 0.18 (u is the standard deviation of the mean value). The isotope effect kP/kZD is near the maximum value for C-H bond-breaking in a methylene group. The predicted rate ratio based on a simple consid-
Few examples of slow proton transfer from catalyzing acid t o substrate are known, and not many measurements of the isotope effect on this reaction have been made. In the several cases reported so far, the deuterium isotope effect ( k H / k D ) has not been larger than two whenever the catalyzing acid has been the solvated proton in dilute aque(3) F. A. Long and J. Bigeleisen, Trans. Faraday Soc., 6 6 , 2077 ous solution.2 Since this small isotope effect is in accord with a t least one theoretical p r e d i ~ t i o n , ~(1959). (4) A. J. Kresge and Y. Chiang, J . A m . Chcm. Soc., 81, 5509 (1959); (1) Work supported by the Atomic Energy Commission, in part under USAEC Oontract AT(l1-1)-1025 and in part by Brookhaven National Laboratory. (2) (a) S. H. Maron and V. K. LaMer, J . A m . Chcm. SOC.,61, 692 (1939); A. V. Willi, Z . Naturfoorschg., 18% 997 (1958); 2. physik. Chem. N . F . , 27, 221 (1961); H . 0. Kuivila and K. V. Nahabedian, J . A m . Chem. Soc., 83, 2164 (1961); ( b ) F. A. Long and D. Watson, J . Chem. Soc., 2019 (1958); T. Riley and P. A. Long, J . A m . Chcm. Sac., 84, 522 (1962).
83, 2877 (1961); Proc. Chcm. SOC.,81 (1961). (5) (a) C. G. Swain, E. C. Stivers, J. F. Reuwer, Jr., and L. 1. Schaad, J . A m . Chem. Soc., 80, 5885 (1958); L. Melander, “Isotope Effects on Reaction Rates,” The Ronald Press Company, h’ew York. N. Y.,1960, p. 23; (b) S. Olsson, Arkiu K e m i , 1 6 , 4 8 9 (1960). (6) (a) A. Streitwieser, Jr., R. H. Jagow, R. C. Fabey and S. Suzuki, J . A m . Chcm. Soc., 80, 2326 (1058); (b) L. Melander, “Isotope Effect, on Reaction Rates,” The Ronald Press Company, h’ew York, N. Y. 1960, 107 ff.
Oct. 20, 1962
COMMUNICATIONS TO THE EDITOR
3977
eration of zero-point energy differences for a C-H the two forces holding the proton in the transition stretching vibration of 2900 cm.-' is 7.8, and the state should be more nearly equal. Thus, the isolargest deuterium isotope effect reported in elec- tope effect with trimethoxybenzene should be trophilic aromatic substitution is 6.67.7 Thus, the stronger than those reported before. isotope effect weakening produced by the symmetriThis isotope effect on proton transfer to trical stretching vibration in the transition state of a methoxybenzene is twice the value which was used three-center reaction must be near its minimum to estimate that HzO is five times as strong an acid value in this case. as Dz0.2b Similar reasoning with the data for The 0-H stretching vibration in the solvated trimethoxybenzene gives a ten-fold difference in proton occurs a t 2900 cm.-1,8 and the value pre- acid strength between H20 and DzO. This undicted for the other isotope effect, klH20/klDa0, is 7.8 reasonably high isotope effect emphasizes the as well. This is considerably greater than the ob- danger inherent in basing conclusions on the asserved value of 2.93. But the transition states of sumption that isotope effects will have essentially the two steps in the exchange reaction are the same, constant values. and, if the observed valueof k2H/k2D is near its DEPARTMENT OF CHEMISTRY maximum value, the observed value of k~H20/klD~0ILLINOIS INSTITUTE OF TECHNOLOGY 16, ILLINOIS A . J . KRESGE must be near its maximum value also. This dis- CHICAGO UNIONCARBIDE RESEARCH INSTITUTE crepancy can be understood in terms of a predicted TARRYTOWN, NEW YORK Y. CHIANG secondary effect of the water molecules solvating RECEIVED JUNE 18, 1962 When a proton is transferred from its the p r o t ~ n . ~ solvent shell, the solvating water molecules revert to ordinary water. Since the 0-H stretching vibraCELLULOSE COLUMNS CONTAINING POLYRIBONUCLEOTIDES AND RIBONUCLEIC tion in liquid water is 3400 cm.-', this process is ACIDS accompanied by considerable bond-tightening. It has been estimated that the deuterium isotope Sir : Recent studies2, have shown that polydeoxyrieffect on this change is 0.7 per 0-H bond for an equilibrium process, 9a and the prediction has been bonucleotides can be covalently linked to cellulose made that this will reduce the kinetic isotope effect and then be employed as chromatographic adsoron proton transfer to a maximum value of about bents which selectively bind polynucleotides com3.6.9b The observed effect of 2.93 is in good agree- plementary to them in base sequence. The methods of synthesis utilized the glucosidic hydroxyl groups ment with this prediction. of T4 DNA2 and the terminal phosphate groups of TABLE I thymidine oligonucleotides.s This suggested to us that the free hydroxyl groups on ribose in polyRATESOF AROIIATIC HYDROGEN EXCHANGE BETWEEN 1,3,5ribonucleotides might be available for similar TRIMETHOXYBENZENE AND 0.050 M HCIOl AT 25' reactions. We have found that phosphocellulose Substrate Solvent lo* kz ( M - 1 m h - 1 ) No. of runs can indeed be linked to synthetic polyribonucleoTMB-t Hz0 3.722 A 0.030" 9 tides and made into columns capable of binding and TMB-d HzO 7.98 A . i o 0 7 desorbing polynucleotides. The specificity of adTMB-t Dz0 6.286 A .012" 5 sorption with respect to the nature of the bases, Error estimates are standard deviations of the mean salt concentration and temperature, is very similar values. to that for the formation of helical complexes in The difference between this approximately solution. Columns also have been prepared from maximum isotope effect on proton transfer to 1,3,5- natural ribonucleic acids. Use of these columns trimethoxybenzene and the other smaller isotope may constitute a chromatographic method for effects on slow proton transfer2 is understandable in isolating cellular components complementary in terms of the relative basicities of the various pro- base sequence to the RNAs. ton donors and acceptors.gb Isotope effects in We followed a procedure similar to Bautz and three-center reactions will be less than their Hall's2 adaptation of Khorana's carbodiimide reacmaximum value whenever the transition state is tion for forming phosphate-ester bonds between not truly symmetrical, that is, whenever the two acetylated phosphocellulose (Serva, 0.78 meq. P/g.) force constants governing the symmetrical stretch- and each of the listed polyribonucleotides: pylying vibration in the transition state are not equal.lO A, poly-C, poly-I, poly-U, bacteriophage virus I n the slow proton transfer reactions on which iso- RNA, E. coli transfer and ribosomal RNAs. tope effects have heretofore been reported, the pro- Nucleotide polymers and cellulose were dissolved in ton acceptor has usually been a strongly basic pyridine and reacted with dicyclohexyl carbodianion. In these transition states, therefore, the imide a t 115' for one hour. After the reaction proton is not bound with equal strength to the ac- product was isolated, it was chopped in a Waring ceptor molecule and the solvating water which it is blender a t 4 O , ground in a mortar, and washed exleaving. Trimethoxybenzene, on the other hand, tensively with neutral buffer a t 80' to liberate is a weaker base than these anions, and in this case pyridine and starting materials. After removal of (7) H.Zollinger, Rclu. Chim. Acta, 88, 1957,1617,1623 (1955). (8) M. Falk and P. A. Giguere, Can. J . Chem., 8 5 , 1185 (1959); C. G.Swain and R. F. Bader, Tetyahcdron, 10, 182 (1960). (9) (a) C. G. Swain and E. R. Thornton, J. A m . Chem. Soc., 88, 3884 (1961); (h) C. A. Bunton and V. J. Shiner, Jr., {bid., 8 8 , 42, 3207, 3214 (lY6l). (10) F. H Westheimer, Chcm. Reus.. 61, 265 (1961).
(1) Abbreviations used: poly-A or simply A, polyriboadenylic acid; poly-C or C, polycytidylic acid; poly-I or I, polyinosinic acid; poly-U or U, polyuridylic acid; tris, tris-(hydroxymethy1)-aminomethane. (2) E. K.F. Bautz and B. D. Hall, Proc. Nail. Acad. Sci. U.S., 48, 400 (19621. (3) P.T.Gilham, J . A m . Chcm. Soc., 84, 1311 (1962).
