Photochemistry and Photobiology Vol. 34, pp. 107 to 109, 1981 Printed in Great Britain. All rights reserved

003 1-8655/81/070107-03502.00/0 Copyright 0 1981 Pergamon Press Ltd

RESEARCH NOTE

PHOTODIMERIZATION OF 1,3-DIMETHYL-5- CHLOROURACIL”

HARRY MORRISONt, CHRISTINE IZUNOt, STEPHEN B Y R N ~ and ANN MCKENZIES tDepartment of Chemistry, Purdue University, and

$Department of Medicinal Chemistry, Purdue University, W. Lafayette, IN 47907. USA

(Received 20 November 1980; accepted 27 January 1981)

Abstract-1,3-DimethyI-5-chlorouracil dimerizes upon direct photolysis in aqueous solution to give one major photodimer. X-Ray analysis has been used to identify the dimer as the anti head-to-tail isomer, IV.

1NTRODUCTlON

The photochemistry of 5-halouracils has been a sub- ject of continuing interest because of the incorpor- ation of these compounds into DNA and the resultant increase in DNA sensitivity to UV light (reviews: Hutchinson, 1973; Rahn and Patrick, 1976; Wang, 1976; recent examples: Swanson et al., 1980; Rahn and Stafford, 1979; Shim and Lee, 1979). However, by comparison with the fluoro, bromo and iodo deriva- tives, 5-chlorouracil has received relatively little atten- tion (see reviews above, and Southworth and Gehrs, 1976), though its recent identification in sewage efflu- ent has stimulated interest in its chemistry (South- worth and Gehrs, 1976; Pal, 1978).

Our concern with 5-chlorouracil photochemistry was stimulated by the above considerations and by our ongoing inquiry into the role of ground state aggregates in solution phase photodimerization (Mor- rison et al., 1976), particularly in the uracil series (Otten e t al., 1977). The presence of a chlorine atom in several aryl monomers has been shown to direct crys- tal packing into a sandwiched, head-to-head (h-h)t configuration, apparently due to attractive CljCl in- teractions (Cohen and Green, 1973; Desvergne et a/., 1978). Photolysis of such crystals thus primarily gives h-h dimers. Since there is good evidence that 5-substi- tuted-N,N-dimethyluracils photodimerize in solution via aggregates (Otten et a / . , 1977). 5-chloro-N,N- dimethyluracil (DMClU) should provide a test of

*Organic Photochemistry, Part 48. For Part 47, see Morrison, H. and D. Giacherio (1980) J . Chem. Soc. Chem. Commun. 1080-108 1.

?Abbreviations: DMCIU, N,N-dimethyl 5-chlorouracil; DMT, N,N-dimethylthymine; h-h, head-to-head; TMS, tetramethylsilane; TMU, tetramethyluracil.

$Association of DMCIU in water is presumed by ana- logy (Otten et a/., 1977); unfortunately, the solubility of DMCIU is insufficient for accurate osmometry experi- ments.

whether a chlorine atom can direct the photochemis- try of solution phase associates in like fashion, i.e. to preferentially form the DMCIU syn h-h dimer (I).: By comparison, such a dimer constitutes 37% of the pho- todimers formed from N,N-dimethylthymine (DMT) upon photolysis of 0.1 M DMT in water (Kleopfer and Morrison, 1972).

0 PY 0 ri ri 0

DMCIU I (SYn h-h )

MATERIALS AND METHODS

Materials and instruments. N,N-Dimethyl-5-chlorouracil (DMCIU) was synthesized from 5-chlorouracil (Calibio- chem, used as received) in 92’/: yield by a procedure anal- ogous to that used for the preparation of tetramethyluracil (Otten et al., 1977). The DMCIU was recrystallized from water and sublimed at 119-122°C (0.4 mm Hg) to give a sample with mp 150.0-150.2‘C (uncorrected). IR (KBr) 5.84, 6.01, 6.75. 6.92, 7.46, 13.13, 13.22pm; NMR (CDC13) 6

vinyl H); calculated for C6H7NZOZCI, m/e 174.020; found, m/e 174.019.

Quantitative analysis for DMCIU was via HPLC using a composite system consisting of a Waters Model 6000A sol- vent delivery system, a Rheodyne 70-10 sample injection valve with a 20$ loop, a Laboratory Data Control UV monitor with 254 nm detector and a Hewlett-Packard 3380 A electronic integrator. With a Whatman Partisil-10 (11 pn) 25 cm x 4.2 mm column, an eluent of CH2C12:n-hexane:CH3CN (3:3:1), a flow rate of 1.4 m//min and 2-bromonaphthalene as an internal stan- dard, retention times were: standard, 1.9 min; DMCIU, 4.2 min.

IR spectra were recorded in KBr using a Perkin-Elmer Infracord, UV spectra were measured on a Cary IS or Beckman-Gilford Spectrophotometer. proton NMR spec-

3.41 (s, 3H. N-CHA), 3.44 (s, 3H, N-CH,), 7.43 (s, 1H.

107

108 HARRY MORRISON et al.

tra were measured on a Perkin-Elmer R-32 90 MHz spec- trometer with TMS as reference, carbon NMR spectra were recorded on a Varian CFT-20 spectrometer with TMS as reference and the satellite experiment was carried out on a Nicollet NT-360 spectrometer.

Phorodimerization of' DMCIC'. DMCIU (800 mg. 0.046M) in aqueous solution was degassed with Ar for 30 min and irradiated for 1W150 h with a Hanovia 450 W medium pressure Hg arc and Corex or Pyrex filters. White crystals of the dimer form during the photolysis and could be isolated in 2&40";, conversions. Several washings of the filtered dimer with water provided pure (by NMR) samples (mp 213-214-C. sealed tube), for elemental and spectral analysis (see Results), (Attempts to recrystallize the dimer gave samples with broad and varied melting ranges in the region of 205-222 C . )

Quantum qfficirncj of disappearance of DMCIU. Two 5 m/ aqueous solutions of 0.046 M DMCIU were degassed for 15 min with Ar and irradiated in vycor tubes using the 313 nm line of a Hg arc (filtered by 0.5cm of 62mM K2Cr04 + 6.4mM KOH, and by 1 cm of 1.07M CoSO, .7H 20). (The DMCIU solutions completely absorb at this wavelength.) Uranyl oxalate actinometry was used to monitor the light intensity and the DMCIU was assayed in one tube by HPLC, and in the other, by UV spec- troscopy at 313 nm. The HPLC analysis was conducted on aliquots taken to dryness and redissolved in the organic eluent (see above); UV measurements were conducted on diluted aliquots of the photolysis mixture. The HPLC and U V analyses indicated 6.66 and 6.45oi, loss of DMCIU. with photon counts of 6.31 x lozo and 6.16 x lozo, re- spectively. Both results correspond to 4di% = 0.015.

bony1 peaks in the IR, and the disapperance of vinyl hydrogens in the NMR. The I3C NMR (360MHz, CDC13) provides further confirmation: 629.1 and 35.4

and 165.4 (s, C 4 ) . The question then reduces to which of the four

possible cis-fused, cyclobutane isomers are in hand. These are the syn h-h structure (I) noted above, and the anti h-h (II), syn head-to-tail (syn h-t) (111), and anti h-t (IV) dimers. The fact that the DMClU dimer precipitates from the photolysis solution suggests the anti h-t structure (IV), for the equivalent isomer like- wise precipitates from both the DMT and tetremethy- luracil (TMU) photolyses in water (Kleopfer and Morrison, 1972; Otten et al., 1977). The h-t relation- ship receives further support from I3C-H 'satellite' spectra. Working at high concentrations, one can ob- serve the proton signals for hydrogens attached to the naturally abundant 13C atoms in a molecule. These 'satellite' peaks are, at the minimum, doublets with a spacing equal J,,c-,,; however, the vicinal 13CH--"CH relationship in the h-h dimers (1,II) leads to proton-proton coupling which converts each of the 'satellite' peaks into a doublet with a spacing equal to JH-,, (Anet, 1965). This vicinal relationship, and the consequent splitting, is lacking in h-t dimers (111, IV), and we in fact observe the satellite peaks for

(q, N-CH,), 60.8 (s, C--Cl), 66.9 (d, C-H), 151.6

RESULTS AND DISCUSSION

Photolysis of Ar degassed, 0.04M aqueous sol- utions of DMCIU, using a Pyrex or Corex filtered medium pressure Hg arc (i > 260nm), gave an - 25" conversion to white crystals which precipitated during the course of the reaction. [A similar crystal- lization of photodimer is observed with DMT and tetramethyluracil (TMU).] Elemental analysis (calcu- lated for Cl2HI4N4O4Cl2: C, 41.31; H, 4.04; N, 16.06",; found: C. 41.36; H. 3.90: N, 15.86",;) and mol wt determination by vapor pressure osmometry (cal- culated for 348.9; found 347.9) confirm that the new product is a dimer. That the dimer is of the typical 2 + 7 cyclobutane type is demonstrated by UV, IR and NMR absorption spectra; UV (H,O). shoulder 265 nm ( E = 1.7 x lo3): shoulder 245 nm (E = 5.8 x lo3): IR (KBr) 5.84 and 5 . 9 7 ~ ; NMR (CDCI3) 6:3.18

Particularly noteworthy is the disappearance of the characteristic UV chromophore of the monomer [Emax 287 nm (c = 8.33 x lo3)], the presence of the two car-

(3H. N-CH,). 3.37 (3H. N--CH,), 4.37 ( lH, C-H).

the 4.37 6 methine protons to be an unsplit doublet with J , ,< -# , = 159 Hz.

The ultimate confirmation that the DMCIU dimer is indeed IV has been provided by X-ray crystallogra- phy. The dimer crystals belong to space group P2,/c with a = 6.435 (I), b = 16.914 (5), c = 13.584,

= 81.42 (2)", Z = 4. Crystallographic data were col- lected on a Syntex P3 diffractometer out to a 28 of 135'. The structure was solved using the MULTAN

program and refined to a final R factor of 0.064. This refinement was performed using anisotropic thermal parameters for all non-hydrogen atoms. The hydro- gen atoms were placed in calculated positions except for the two cyclobutane hydrogen atoms. All hydro- gen atoms were fixed during refinement. A final differ- ence map revealed no recognizable molecular frag- ments and the highest peak on this map was 0.54 e/(A')3. An ORTEP stereoscopic view of the molecule is given in Fig. 1.

The fact that DMCIU dimerizes virtually exclus- ively in the anti h-t fashion is quite striking; using NMR analysis of the photolysis mixture filtrate, we

Research Note 109

Figure 1. An ORTEP steroscopic view of the DMCIU photodimer.

estimate that IV constitutes -86% of the dimers formed, whereas the corresponding figure for D M T is only 187; (Kleopfer and Morrison, 1972). Clearly, the chlorines are not directing dimerization in a fashion akin to that observed for certain crystals (see Intro- duction), and s t e m and dipole effects are dominating.

The quantum efficiency for loss of DMClU at 0.046 M is 4d,s = 0.015; at the same concentration, $dlq for D M T is 0.016 (Lisewski and Wierzchowski, 1969). However. DMCIU photolysis involves some C-CI cleavage in the monomer as well as dimeriza-

tion, since HCI formation occurs. I t is worth noting that 5-fluorouracil is reported not t o photodimerize upon direct photolysis (Shim and Lee, 1979).

The extent of carbon-halogen cleavage and the mechanistic details of DMCIU photochemistry are under study.

Acknowledgements-This investigation was supported, in part, by NIH Research Grant RR01077 from the Division of Research Resources, and NIH Grant ES00929 to S. B. and A. M. We are grateful to John Saddler for his assist- ance in obtaining the I3C satellite spectra.

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