Assignment of the 'H and 13cspectra of aspidocarpine and assignment of the structure and stereochemistry of the von Braun reaction product of aspidocarpine by 2D nmr spectroscopy STEWART MCLEAN,WILLIAMF. REYNOLDS, AND XINGPEIZHU' Department of Chemistry, Universiw of Toronto, Toronto, Ont., Canada M5S IAl Received June 17, 1986

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This paper is dedicated to Dr. 0.E. (Ted) Edwards

STEWART MCLEAN, WILLIAMF. REYNOLDS, and XINGPEI ZHU.Can. J. Chem. 65, 200 (1987). The I3cand 'H spectra of aspidocarpine and the spectra, structure, and stereochemistry of the product of its von Braun reaction with cyanogen bromide have been totally assigned by a combination of homonuclear (COSY-45) and direct and indirect heteronuclear shift-correlated 2D nrnr spectra.

STEWART MCLEAN, WILLIAM F. REYNOLDS et XINGPEI ZHU. Can. J. Chem. 65, 200 (1987). En se basant sur une combinaison de techniques impliquant des spectres rmn en 2D homonucleaires (COSY-45) et httCronucleaires comportant une corrtlation directe et indirecte avec les dtplacements, on a pu faire des attributions pour toutes les bandes des spectres rmn 'Het I3cde I 'aspidocarpine et de son produit de reaction avec le bromure de cyanogkne (von Braun); de plus, ces techniques ont permis d'etablir la structure et la sttr6ochimie de ce dernier compost. [Traduit par la revue]

I

1I I

As part of our investigation (1) that led us to the elucidation of the structure of aspidocarpine (I), we subjected the alkaloid to a von Braun degradation by treatment with cyanogen bromide. The bromocyanamide formed without difficulty, but the normal solvolytic completion of the degradation failed and it was observed that merely heating the bromocyanamide in ethanol caused it to revert to aspidocarpine. Aspidospermine had been found to behave in a very similar manner (2) and Conroy et a l . presented evidence that the bromocyanamide included a -CH2Br unit as part of a carbon chain with "few degrees of freedom" (2). Although the unprecedented reversal of the von Braun reaction was of considerable interest, the degradation provided little significant structural information and the quesiion of the structure of the bromocyanamide has lain dormant since then. Once the structure of 1 was established, it was possible to conjecture that the bromocyanamide had structure 2 but alternativestructures could also be suggested.

Aspidocarpine and aspidospermine were among the first alkaloids to which 'H nmr spectroscopy was applied for purposes of structure elucidation (1, 2). However, the techniques available then were of limited power and the 'H spectra were poorly resolved, providing limited and sometimes confusing information (2) regarding molecular structure. The importance of the aspidosperimidine alkaloids subsequently became apparent and a considerable range of nrnr data for them is now available (3). However, only limited H data (1) and no 13cdata are available for 1. Consequently, we decided to apply modem nmr spectroscopic techniques to assign unambiguously the H and 13Cspectra of 1as well as to determine whether the structure

'

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'On leave of absence from the Department of Chemistry, Beijing Normal University, People's Republic of China.

of the von Braun reaction product was indeed 2. We have succeeded not only in confirming 2 and totally.assigning the spectra of 1 and 2, but also have succeeded in deducing the conformations of rings C and D of 1 and 2. These results are reported below.

Results and discussion ( a ) Assignment of 'H and I3cspectra of 1 As a first step in assignment, normal and DEFT-edited (4) 13C spectra of 1 were obtained. Then a two-dimensional (2D) heteronuclear ('H-13C) chemical shift correlation experiment (5) was carried out to determine the chemical shift(s) of the hydrogen(s) directly bonded to each protonated carbon. Next, a 'H COSY-45 spectrum (6) (Fig. 1) was used to establish connectivities between coupled protons. There are four distinct sequences in the saturated carbon region that could be assigned C(6)H2from this experiment: C(2)H-C(3)H2-C(4)H2, C(7)H,C(8)H2, C(lO)H2--C(l 1)H2, and C(a)H2-C(P)H3. The COSY experiment did not allow distinction between C(6)H2 and C(8)H2 in the second sequence and C(10)H2 and C(l 1)H2 in the third sequence (due to the symmetry of each sequence). However, C(8)H2 and C(10)H2 were readily assigned due to their deshielding by the adjacent nitrogen. Assignments are summarized in Table 1. Nonprotonated carbons were assigned by carrying out a heteronuclear shift-correlated experiment with delay times optimized for coupling between indirectly bonded carbons . experiment was and hydrogens (e.g., 2 ~ o or 3 ~ C H )This performed using the recently proposed 2 D pulse sequence XCORFE (7). This sequence detects cross peaks corresponding to 13C-X-'H and 13c-X-C-'H pairs (where X = C, N, 0 , etc.). Two-bond connectivities involving a pair of protonated carbons are distinguished from three-bond connectivities because the former are split by vicinal 'H-'H coupling, while the latter appear as singlets (7). Observed connectivities are summarized in Table 2. The remaining carbons are easily assigned using these connectivities (see Table 1). For example, it is trivial to distinguish C(5) from C(12) on the basis of their numerous cross peaks to adjacent protons. The aromatic carbons are assigned on the basis of cross peaks to OH, 0CH3, and to aromatic and saturated protons. Note that the aromatic protons show much stronger three-bond than two-bond peaks due to the relatively

20 1

McLEAN ET AL.

TABLE1. Assigned I3C and 'H chemical shifts and 'H-'H coupling constants for 1 in CDC13

PPM

OH O=C

0.5

0 8

Q

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1.0-

1.5-

at Q3

Q

2.0 -

8.8

I

CH3

e

o

1

s

2.5 -

1

Carbona

6c

2 3 4

70.25 25.10 22.92

5 6 7 8

35.46 34.02 21.50 53.67

6~ (JHH)~.' 4.07 (dd, 11, 6) 1.86 (m), 1.52 (m) 2.00 (td, 12, 14), 1.15 (dm, 12) 1.11 (td, 12,4), 1.65 (dt, 12,4) 1.72 (tm, 12), 1.53 (dm, 12) 1.98 (td, 12, 4), 3.04 (dm, 12) 2.27 (m), 3.12 (m) 1.57 (m), 2.04 (m) 6.61 (d, 8) 6.69 (d, 8)

0.

4.0-

w I

I

I

I

I

I

4.0

3.5

3.0

2.5

2.0

1.5

I

1.0

1

0.5

PPM

FIG. I . COSY-45 spectrum for 1 . The normal spectrum lies along the diagonal with off-diagonal peaks indicating connectivities between coupled protons.

small magnitudes of 2 ~ C H(12). The fact that the OH proton shows a three-bond cross peak with C(16) but not with C(18) may reflect stereospecific coupling arising from hydrogen bonding to the N-acetyl group (1). The data in Table 2 also support the earlier assignments of C(8)H2 and C(10)H2 (made on chemical shift grounds), which are confirmed by the observation of three-bond connectivities between H(19) and C(8) and between H(8) and C(10). These connectivities, the cross peaks involving OH and 0CH3 protons, as well as several other connectivities in Table 2 illustrate the ability of this experiment to establish connectivities through heteroatoms.

(b)Assignment of the structure and ' H and 13cspectra of 2 Exactly the same combination of experiments as outlined in ( a ) was used to investigate 2 (e.g., see the COSY spectrum in Fig. 2) and spectral assignments were made in a similar manner. Results are summarized in Tables 3 and 4. Several observations clearly indicate that 2 is the correct structure for the reaction product. First, the carbon assigned as C(10) is shifted from 6 52.4 in 1 to 6 27.0 in 2, consistent with replacement of a bond to nitrogen by one to bromine (8). Second, the large nonequivalence of the methylene protons of C(10) in 1 has almost been eliminated in 2, consistent with the fact that a bond has been broken between C(10) and the chiral nitrogen. Finally, the XCORFE experiment revealed two connectivities to the cyano group, one to H(19) and the other to the H(8) proton at 6 3.02, consistent with attachment of the cyano group to the nitrogen, as shown in 2. The fact that no significant cross peak was observed involving the second H(8) proton at 6 3.63 provides useful conformational information. This is discussed below.

2.25 (s) 0.93 (m), 1.44 (m) 0.63 (t, 7.5) 3.88 (s) 2.33 (s)

'Numbering shown below. bSplittingpattern in parentheses (multiplicity, coupling constant (in Hz) where resolved). 'In cases of protons in rings C and D, the first listed proton of a methylene pair is the axial proton, while the second is equatorial. For C(10), C(11), and C(u), the protons are listed in chemical shift order.

(c)Deduction of the conformations of 1 and 2 from three-bond 'H-'~c connectivities The solution conformation of 1 is unknown. The N-methiodide of the closely related alkaloid aspidospermine has been shown by X-ray crystallography to have a chair conformation for ring C but a boat conformation for ring D (see 3) (9). However, on the basis of infrared spectroscopy, it was concluded that ring D most probably is in a chair conformation in the free base (see 4) (10). The nmr data strongly suggest that the conformation of 1 is very close to that shown in 4. For example, the 'H spectrum of 1 shows multiplets for the methylene protons of C(4), C(6), and C(8), all of which show the expected couplings for axial and equatorial proton pairs

202

CAN. I. CHEM. VOL. 65, 1987

TABLE2. Observed two-bond and three-bond I3c-'H connectivities from XCORFE experiments on 1

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Carbon

'H connectivitiesa

n

"The number indicates the proton while the subscript indicates whether it is axial (a) orequatorial (e) in ring C or D, or high field (h) or low field (I) for other methylene pairs. Weak connectivities are indicated in parentheses. d = a doublet splitting, while b = a broadened line with unresolved splitting. Observation of coupling indicates a two-bond connectivity between a protin and an adjacent protonated carbon (see ref. 7). Couplings of less than 7 Hz are not detected due to limited data point resolution.

TABLE3. Assigned I3C and 'H chemical shifts and 'H-'H coupling constants for 2 in CDC13 Carbona

6c

6~ (JHH)

2 3 4 5 6 7 8 10 11 12 13 14 15 16 17 18 19

65.67 24.71 21.93 38.47 34.38 20.12 53.51 26.97 41.03 50.07 129.38 114.01 108.92 150.42 138.44 126.13 64.68 31.16 6.93 56.45 22.80 169.74 119.94

4.56 (dd, 11, 6) 2.15 (m), 1.56 (m) 2.24 (td, 12, 4), 1.33 (dm, 12)

CL

P

0CH3 COCH3 COCH3 CN -

-

1.11 (td, 12,4), 1.41 (dt, 12,4) 2.02 (tm, 12), 1.65 (dm, 12) 3.02 (td, 12, 4), 3.63 (dm, 12) (3.27) (mlb 1.93 (m), 2.82 (m) 6.56 (d, 8) 6.77 (d, 8)

3.29 (s) 0.72 (rn), 1.17 (m) 0.58 (t, 7.5) 3.91 (s) 2.38 (s) OH 10.90

"Numbering is identical to that in Table 1. bThe C(10) protons appear as a complex multiplet centered at 8 3.27. The nonequivalence of these protons is approximately 0.03 ppm.

- -.-.8

"

1

S O B I

o

6

a'@ •

@ o *

0

0

I

I

I

I

I

I

I

1

4.5

4.0

3.5

3.0

2.5

2.0

1.5

1.0

I

0.5

PPM

FIG.2. COSY-45 spectrum for 2. in a chair form (11). i.e., the axial protons show two large (ca. 12 Hz) splittings corresponding to 2 ~ H ( a ) H ( e and ) 3~H(a)H(a) plus a smaller splitting (ca. 4 HZ) corresponding to 3~H(a)Hse while the equatorial protons show one large 2~H(a)H(e, splitting and two smaller splittings assignable as 3 ~ H ( e ) H ( a )and JH(eIH(e) (see Table 1). Similar splitting patterns are observed for the corresponding protons in 2 (see Table 2), suggesting that rings C and D in both 1and 2 are in chair conformations. Further evidence concerning the specific chair conformations in each case is provided by the indirect shift-correlated experiments. 3 ~ C Hcouplings are conformationally dependent with typical values for anti, syn, and gauche couplings being, respectively, ca. 8, 5, and 2 Hz (12). Polarization transfer between ' H and 13C is maximum when the fixed delay, T, equals (2JcH)-' (7). Most spectra were run using T = 0.06 s (see Experimental), corresponding to an optimum JCH = 8 HZ. Consequently, gauche 3 ~ C H coupling vectors are still far removed from the antiphase orientation that leads to maximum polarization transfer (5). After polarization transfer, a second delay allows the I3C-'H coupling vectors to return to an in-phase orientation prior t o 'H decoupling and I3C detection (5). Once again the delay is too short for gauche 3JCH coupling vectors to reach this optimum orientation. Consequently threebond gauche C-H connectivities are effectively suppressed, while those corresponding to anti pathways are optimized.2 This allows one to probe conformations of rings C and D in terms of observed or missing three-bond peaks. Specifically, equatorial hydrogen should show three-bond 'H-13C cross peaks, while axial hydrogens do not. For example, the isolated 'A somewhat similar approach has recently been used to deduce conformational information for ryanodine (13).

203

McLEAN ET AL.

TABLE4. Observed two-bond and three-bond I 3 c - l ~ connectivities from XCORFE experiments on 2

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Carbon

H connectivities"

suggests an explanation for the reversal of the reaction: the equatorially disposed lone-pair electrons o n nitrogen are well placed to effect a n intramolecular displacement of the bromide. I t is reasonable to suppose that this process corresponds to the microscopic reversal of the bond cleavage in the forward direction and leads to the conclusion that the configuration at the nitrogen of the cyanoammonium intermediate is opposite to that in the methiodide (see 3). It follows then that t h e cyanogen bromide in the von Braun reaction undergoes a facile reaction with the free base in its dominant conformation (4), while methyl iodide fails to react with the nitrogen in this conformation and selectively quaternizes the nitrogen o f the minor conformational isomer.

Experimental

"See footnote a of Table 2

proton, H(19), is in an axial orientation with respect to ring D and a n equatorial orientation with respect to ring C . As expected, it shows strong three-bond cross peaks with C(2) and C(4) of ring C but n o significant cross peaks with C(6) or C(8) in ring D. By contrast, in both 1 and 2 C(19) shows strong cross peaks with hydrogens H(4), ( 6 1.33), H(6), ( 6 1.82), and H(8), but not from H(2), which is in a pseudo-axial orientation with respect to ring C . Similarly, there is a cross peak between C(2) and H(4), but not between C(4) and H(2), while strong cross peaks are also observed between C(6) and H(8), and C(8) and H(6),. All of these observations are entirely consistent with chair forms for rings C and D of 1 and 2, similar to 4. T h e connectivity data also strongly imply an axial orientation for the cyano group bonded to nitrogen in 2 (see 5). Three-bond connectivities are observed between the cyano group and both H(8), and H(19) but not with H(8),. Since this indicates an anti orientation of the cyano group with respect to the two axial hydrogens, it follows that the former group is axial. This was further supported by selectively ' H decoupled I3C spectra for the cyano group, which indicated couplings of 9 , 2 , and 9 H z to H(8), , H(8), , and H(19), respectively. While the mechanism of the von Braun cyanogen bromide reaction received little earlier attention, it is now established that the reaction proceeds through the N-cyanoammonium salt (14). T h e reversal of the reaction, eliminating CNBr from the bromocyanamide, is rarely observed. The conformation deduced for the bromocyanamide (5) in the present case

Br

The isolation of aspidocarpine has been described elsewhere (1). The von Braun reaction was carried out using previously described conditions (1). All nmr data were obtained on a Varian XL-400 nmr spectrometer (13C 90" pulse width = 13.5 ps, 'H 90" pulse width 4 13.6 ps, 'H 90" pulse width through decoupler coil of multinuclear probe = 27.0 ps, probe temperature = 18°C). In each case, 70 mg of 1 or 2 was dissolved in ca. 0.5 rnL of CDC13 in a 5-rnm tube. Typical parameters for the COSY spectrum were f l = f 2 = 2200 Hz with 512 time increments (zero filled to 1024), 1024 data points, 16 transients per time interval, and 0.5 s relaxation delay. Pseudo-echo processing was used in both domains, followed by symmetrization by triangular folding. Typical parameters for direct heteronuclear shift-correlated spectra were f , = 2200 Hz, f 2 = 7500 Hz, 256 time increments (zero filled to 1024), 2048 data points, 64 transients per time interval, and 0.5 s relaxation delay. Separate spectra were obtained for the aromatic region. Two sets of XCORFE spectra were obtained in each case with fixed times T = 0.062 s (272 time increments for 2200 Hz, f, spectral width) and T = 0.08 s (352 time increments for 2200 Hz, f l spectral width) and 256 transients per time interval. These are, respectively, optimized for JCH= 8 HZ and JCH= 6 HZ. Note that in XCORFE (7) and other similar 2D experiments (15, 16) one not only must choose T ( 2 ~ C H ) -but L also avoid the condition T (2JHH)-Isince polarization transfer is eliminated if 'H-'H coupling vectors are antiphase at the end of T (16). T = 0.062 is a good choice in most cases since JHH is generally less than 5 Hz or greater than 10 Hz. However, in some cases (H(2), H(a), H(P), and the aromatic protons) JHH 8 HZ. The alternative time generally gave better cross peaks for these protons.

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Acknowledgements Financial support from the Natural Sciences and Engineering Research Council o f Canada (to S . M c L . and W.F.R.) and the Government of the People's Republic of China (to X.Z.) is gratefully acknowledged. 1. S. MCLEAN,K. PALMER,and L. MARION.Can. J. Chem. 38, 1547 (1960). 2. H. CONROY,P. R. BROOK,M. K. ROUT,and N. SILVERMAN. J. Am. Chem. Soc. 80, 5178 (1958). 3. G. A. CORDELL.In The alkaloids. Vol. 17. Edited by R. H. F. Manske and R. Rodrigo. Academic Press, New York. 1979. 4. D. M. DODDRELL, D. T. PEGG,and M. R. BENDALL.J. Magn. Reson. 48, 323 (1982). 5. A. BAXand G . A. MORRIS.J. Magn. Reson. 42, 501 (1981). 6. A. BAXand R. FREEMAN. J. Magn. Reson. 44,542 (1981). and 7. W. F. REYNOLDS,D. W. HUGHES,M. PERPICK-DUMONT, R. G . ENRIQUEZ.J. Magn. Reson. 63,413 (1985). 8. G. E. MACIEL.I n Topics in carbon-13 nmr spectroscopy. Vol. 1. Edited by G . C. Levy. Wiley-Interscience, New York. 1974. 9. J. F. D. MILLSand S. C. NYBURG.J. Chem. Soc. 1458 (1960). 10. G. F. SMITHand J. T. WROBEL.J. Chem. Soc. 1463 (1960). and S. STERNHELL. Applications of nuclear mag11. L. JACKMAN

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netic resonance spectroscopy in organic chemistry. Pergamon Press, Oxford. 1969. 12. P. E. HANSEN.Prog. Nucl. Magn. Reson. Spectrosc. 14, 175 (1981). I. HOLDEN,and J. E. CASIDA. J. Chem. Soc. 13. A. L. WATERHOUSE, Perkin Trans. 2, 1011 (1985).

1987

14. G. FODOR,S. ABIDI,and T. C. CARPENTER. J. Org. Chem. 39, 1507 (1974). 15. H. KESSLER,C. GRIESENGER, J. ZARBOCK,and H. R. L O O ~ I . J. Magn. Reson. 57, 331 (1984). 16. C. BAUER,R. FREEMAN, and S. WIMPERIS.J. Magn. Reson. 58, 526 (1984).