MAGNETIC RESONANCE IN CHEMISTRY, VOL. 32, 727-731 (1994)

First Discrimination of Enantiorneric Cyclic Herniacetals and Methyl Acetals Derived from Hydroxarnic Acids and Lactarns of Gvamineae by Means of lH NMR Using Various Chiral Solvating Agents

Jens Klein, Holger Hartenstein and Dieter Sicker* Fakultat fur Chemie und Mineralogie der Universitat Leipzig, Talstr. 35, D-04103 Leipzig, Germany

The discrimination of enantiomeric cyclic hemiacetals and methyl acetals derived from hydroxamic acids and lactams with the 2H-1,4-benzoxazin-3(4H)-one and 2H-1,4-benzothiazin-3(4H)sne skeleton was investigated using (S)-( - )-phenylethylamine, (-)-quinine, byclodextrin and, for the first time, (5R,1 1RH + )-2,8dimethyl- 6H,12H-5,1l-methanodibenzo IbJl I 1,5]diazocine, a Troeger's base enantiomer, as chiral solvating agents (CSA). Conditions for the enantiomeric discrimination of six configurationally stable methyl acetals are reported. 2,4- Dihydroxy-2H-1,4-benzoxazin-3(4~-one and its 7-methoxy derivative, bioactive aglucones from Gramineae species, are the first cyclic hemiacetals that could be differentiated into enantiomem by means of 'H NMR, despite their oxo-cyclo tautomerization that prevented enantioseparation by chromatography or capillary electrophoresis. However, 2-hydroxy-2H-1,4-benzothiazin-3(4~-ones (thiohemiacetals) could not be differentiated by the use of these CSA. The influence of the structure of the enantiomers, CSA, temperature and concentration on the size of the chemical shift anisochrony is discussed.

KEY WORDS NMR 'H NMR Enantiomer discrimination acetals Hydroxamic acids

INTRODUCTION

'H NMR is a useful method for determining enantio- meric compositions, supplementing chiral chromatog- raphy, chiral capillary electrophoresis and chiroptical methods. '*' Enantiomers cannot be distinguished by 'H NMR without a chiral influence because enantio- topic nuclei show isochronous resonances. However, these nuclei become diastereotopic in a chiral environ- ment and hence cause anisochronous resonances. Recently, some general methods which facilitate the dis- crimination of enantiomers by 'H NMR have been re~iewed.~ They make use of either chiral derivatizing agents (CDA), chiral lanthanide shift reagents (ClSR) or chiral solvating agents (CSA) for obtaining diastereo- topic protons. The advantages of using the last two methods have been pointed out, but differentiation of acidic enantiomers with CLSR is generally difficult because of the poor stability of the lanthanide complex- es at low pH.3.4 We are interested in the isolation' and synthesis6-' of the bioactive cyclic hemiacetals 1 and 2 with the 2-hydroxy-2H-l,4-benzoxazin-3(4H)-one skele- ton which occur naturally in the form of their (2R)-2-P- D-g luc~s ides~~ '~ These glucosides are active as plant resistance factors" in various species of Gramineae, Acanthaceae and Ranunculaceae, respectively. Recently, we have shown that the aglucones 1 and 2, isolable after

* Author to whom correspondence should be addressed.

Chiral solvating agents Cyclic hemiacetals Cyclic methyl

enzymatic cleavage of their (2R)-glucosides, are racemic and can be separated into enantiomers neither by high- performance liquid chromatography (HPLC) using a P-cyclodextrin-modified stationary phasei2 nor by high- performance capillary electrophoresis (HPCE) using various chiral additive^.'^ This is because the oxo-cyclo tautomerization of both hemiacetals takes place rapidly in comparison with the time-scale of the HPLC or HPCE separation processes (Scheme 1).

Making use of the CLSR method did not seem prom- ising for the discrimination of enantiomers by 'H NMR owing to the high acidity of both 1 and 2 (pK, 6.9) arising from their cyclic hydroxamic acid structure. However, 'H NMR using CSA should be a suitable method for the differentiation of the 2R- and 2S- configurations of the hemiacetals 1 and 2. Suitable CSA are expected to form diastereomeric solvation complex- es with both enantiomers in competition with the achiral solvent. The size of the chemical shift non- equivalence depends not only on the structure of the CSA used, which should effect a clear but distinct com- plexation to both enantiomers, but also on the solvent, temperature and concentration.

Although enantiomeric purity determination via CSA-induced chemical shift anisochrony has been reviewed for a broad range of cornpo~nds,~* '~~' ' the discrimination of enantiomeric cyclic hemiacetals and methyl acetals has, to the best of our knowledge, not been described. Hence it was the aim of this work to investigate the enantiomeric differentiation of the cyclic

CCC 0749-158 1/94/120727-05 0 1994 by John Wiley & Sons, Ltd.

Received 25 April 1994 Accepted (revised) 11 July 1994

728 J. KLEIN, H. HARTENSTEIN AND D. SICKER

" OH L

(2R)

Scheme 1

hemiacetals 1 and 2 and methyl acetals 3-6, whose N- hydroxy lactam or lactam structure lies within a 1,4- benzoxazin-3(4H)-one skeleton, and also the thioanalo- gous 1,4-benzothiazin-3(4H)-ones 7-10 (Scheme 2). For this purpose, (S)-( -)-phenylethylamine, (-)-quinine, fl- cyclodextrin and, for the first time, (5R,11R)-( +)-2,8- dimethyldfi, 12H-$1 l-methanodibenzo[b& [ 1,5]diazo- cine, a Troeger's base enantiomer, were used as CSA in different solvents at various temperatures.

EXPERIMENTAL

Compounds

Compounds 1-10 were prepared according to the literature procedures cited: 2,4-dihydroxy-2H-1,4- benzoxazin-3(4H)-one, 1 (DIBOA);6 2,4-dihydroxy-7- methoxy-2H-l,4-benzoxazin-3(4H)-one, 2 (DIMBOA);' 4-hydroxy-2-methoxy-2H-1,4-benzoxazin-3(4H)-one, 3 (DIBOA methyl acetal);I6 4-hydroxy-2,7-dimethoxy- 2H-1,4-benzoxazin-3(4H)-one, 4 (DIMBOA methyl acetal);I6 2-methoxy-2H-1,4-benzoxazin-3(4H)-one, 5 (HBOA methyl acetal);" 2,7-dimethoxy-2H-1,4- benzoxazin-3(4H)-one, 6 (HMBOA methyl aceta1);I7

(DIBTA);I8 2-hydroxy-2H-1,4-benzothiazin-3(4H)-one, 8 (HBTA);' 4-hydroxy-2-methoxy-2H- 1,4-benzo- thiazin-3(4H)-one, 9 (DIBTA methyl acetal);" and 2- methoxy-2H-1,4-benzothiazin-3(4H)-one, 10 (HBTA methyl acetal).lg

2,4-dihydroxy-2H- 1,4-benzothiazin-3(4H)-one, 7

Compound

1 2 3 4 5 6 7 8 9

10

X

0 0 0 0 0 0 S S S S

R 2

OH OH OH OH H H OH H OH H

R3

H OCH, H OCH, H OCH, H H H H

Scheme 2

NMR spectra

'H NMR spectra were recorded on a Varian Unity 400 spectrometer at 399.952 MHz. All proton shifts are ref- erenced to internal tetramethylsilane on the 6 scale. Typical measurement parameters were as follows : 90" pulse width, 15 ps at transmission power 55; acquisition time, 2.65 s; spectral width, 6000 Hz; and number of data points, 32K . Compounds 1-10 were studied in 5 mm tubes.

RESULTS AND DISCUSSION

Influence of structures of enantiomers and various CSA on the Occurrence and size of chemical shift anisochrony A6

In non-polar solvents the high acidity of all compounds having hydroxamic acid units allows the formation of diastereoisomeric complexes when a basic CSA is used. This has been demonstrated in the case of chiral car- boxylic acids with (S)-phenylethylamine" and, more recently, with (1 R,2R)- 1,Zdiphenylethane- 1,2-diamine." Figure 1(A) shows that antipodes of DIMBOA methyl acetal 4 can indeed be differentiated by (S)-( -)-pheny- lethylamine in tetrahydrofurane-d8 (THF-d8) with a small chemical shift non-equivalence of both the 2-CH and the 2-OCH3 signal of 0.003 ppm. Using (-)- quinine, surprisingly only once reported as CSA for bin- aphthyls and alkylarylcarbinols,22 a clearly enhanced differentiation of enantiomers was observed [Fig. l(B)]. This may be a result of the two basic centres now avail- able for the formation of diastereomeric complexes which are conformationally more rigid than those formed by the phenylethylamine enantiomer.

Table 1 shows that all racemic methyl acetals, 3, 4 and 9, having hydroxamic acid units, and also 5, 6 and 10 having lactam units, can be discriminated into enan- tiomers using (-)-quinine as CSA in CD,C12 .

Several assessments can be given comparing the chemical shift non-equivalences of structurally related or heteroanalogous compounds. Typically, methyl acetals 3, 4 and 9 belonging to the hydroxamic acid group are more easily discriminated than the lactam methyl acetals 5, 6 and 10. Obviously, this difference originates from the reduced acidity of lactams in com- parison with hydroxamic acids. In general, the signals of the 2-methoxy group of the diastereomeric complexes can serve as better indicators than the methine group for enantiodifferentiation owing to their higher Ad values. Also, the comparison of heteroanalogous pairs

‘H NMR DISCRIMINATION OF ENANTIOMERIC CYCLIC HEMIACETALS AND METHYL ACETALS 729

(sol“.) n

I 5.375 5.365 3.515 3 5 0 5

2-CF 2 - 0 4 e

I I I

5 . 3 5 5 . 3 0 3 . 5 0 3 . 4 5

2 - C H 2-OHe

Figure 1. Partial 399.952 MHz H NMR spectra of racemic 4 at 299 K showing the enantiomeric discrimination for the 2-CH and 2-OCH, signals: (A) in THF-d, (c =0.050 mmol ml-’) in the presence of (S)-(-)-phenylethylamine (0.08 mmol ml-l); (6) in CD,CI, (c=O.O25 mmol ml-’) in the presence of (-)-quinine (c = 0.020 mmol ml-’). Chemical shifts dare given in ppm.

3 9 and 5-10 shows that the 1,4-benzoxazin-3(4H)-one derivatives are better differentiated into enantiomers than their 1,4-benzothiazin-3(4H)-one counterparts, which also needed a higher CSA concentration.

The first enantiodifferentiation of cyclic hemiacetals has been realized for the natural products DIBOA (1) and DIMBOA (2) in THF-d, also using (-)-quinine as CSA (Fig. 2). The 2-OH protons of both hemiacetals could not be detected owing to the 0-D exchange. Therefore, no CHOH coupling was observed in either case. Hence, the singlets at 5.736 and 5.742 ppm for 1 and at 5.730 and 5.737 ppm for 2 have to be assigned to the 2-CH protons of their enantiomers. Hence, at ambient temperature a weak but distinct chemical shift

Table 1. Selected ‘H chemical shifts (6) and chemical shift non-equivalences (A6) for signals from substituents at the chiral centre of racemic methyl acetals (c = 0.025 mmol ml- *) in the presence of the CSA (-)-quinine‘ in CDzCIz at 399.52 MHz and 299 K

‘CSA CHOCH, Compound (mmol ml-’) d (pprn)

3 0.025 3.481 3.51 9

4 0.020 3.449 3.500

5 0.025 3.507 3.518

6 0.025 3.508 3.51 9

9 0.050 3.345 3.372

10 0.075 3.362 3.373

M CHOCH, M (ppm) d h m ) (ppm)

5.390 0.038 5.400 0.010

5.337 0.051 5.359 0.022

5.249 0.011 5.256 0.007

5.226 0.011 5.236 0.010

5.027 0.027 5.034 0.007

4.882 0.011 4.885 0.003

a CSA added in the form of ( - )-quinine hydrochloride dihydrate.

non-equivalence of about 0.006 ppm could be achieved. This differentiation of enantiomeric cyclic hemiacetals is not affected by their oxo-cyclo tautomerization because populations of equal size are always present for the for- mation of diastereomeric associations.

However, it was impossible to investigate the thiohe- miacetals 7 and 8 using the CSA (-)-quinine in THF-d8 because of the signal of interest for the methine proton is covered by resonances of the CSA. Overcoming this difficulty required another CSA having properties similar to those of quinine but an open spectral window at about 5.50 ppm. It was tentatively expected that a Troeger’s base enantiomer, hitherto undescribed as a

5 . 7 6 5 . 7 4 5 . 7 5 5 . 7 3

2-CH 2-CH

Figure 2. Partial 399.952 MHz ’H NMR spectra for the hemi- acetals (A) DIBOA (1) and (8) DIMBOA (2). c,,, = 0.050 mmol ml-’ for both 1 and 2 in THF-d,, CSA (-)-quinine (c = 0.10 mmol ml-’) at 299 K. Chemical shifts 6 are given in ppm for 2-CH protons.

730 J. KLEIN, H. HARTENSTEIN AND D. SICKER

Table 2. Selected 'H chemical shifts (6) and chemical shift non-equivalences (A6) for signals from the chiral centre of racemic 4 and 9 (c = 0.050 mmol ml-') caused by &CD (c = 0.060 mmol ml-') in D,O at 399.952 MHz and 298 K

CHOCH, M CHOCH, M Compound d (ppm) (ppm) d (ppm) (ppm)

5.508 0.01 2

3.427 0.004 5.280 0.004

b 5.496 4"

9 3.423 5.276

'To increase its solubility, 4 was measured in 0.05 M NaOD. Obscured by B-CD resonances.

CSA, might be suitable. Indeed, by addition of (5RJ 1R)- ( +)-2,8-dimethyl-6H, 12H-5,1l-methanodibenzo[bfl [1,5]diazocine, it was possible to discriminate the 2- OCH3 singlets of the racemic methyl acetals 4 and 9 with a chemical shift anisochrony of 0,003 (craC = 0.025 mmol ml-' for both 4 and 9, cCSA = 0.025 mmol ml-' in CD,CL2 at 299 K). However, this weak discrimi- nating effect prevents a discernible differentiation of the 2-CH protons in both compounds. Similarly, the methine protons of the thiohemiacetals DIBTA (7) and HBTA (8), now observable owing to the spectral window, could not be differentiated, because in this case the Troeger's base enantiomer did not act as a CSA.

The application of cyclodextrins (CDs) as CSA for the determination of enantiomeric excesses by 'H NMR has been reviewed several and has recently been shown to be useful even for the differentiation of non-polar cl-pinene enantiomers.26 Usually, a CD that can effect chromatographic separation of enantiomers is also expected to act as a CSA because of the similar mechanism for the interaction of a chiral guest with the CD cavity. Recently, we have reported enantio- separations of racemic DIBOA methyl acetal (3) and DIMBOA methyl acetal (4) by means of HPLCI2 and HPCE', using p-CD as chiral selector.

Table 2 shows the differentiation of enantiomers of 4 and 9 on addition of /I-CD. In each case the methine protons show a chemical shift non-equivalence. Only the 2-OCH3 group of 9 could be discriminated, because the 2-OCH3 of 4 is covered by 8-CD resonances. Fur- thermore, clear evidence for the complexation of both methyl acetals by /I-CD is also provided by the observa- tion of a guest-induced shift of the inner protons (H-3, H-5, H-6) to higher field.

Temperature and concentration dependences of chemical shift anisochrony

Assuming an optimum enantiomer to CSA ratio, the appropriate choice of concentration and temperature is most important to maximize the size of chemical shift anis~chrony.~ This is illustrated by the results of temperature-dependent measurements listed in Tables 3 and 4. Lowering the temperature of a dilute solution (c = 0.005 mmol m1-I) of the hemiacetal DIBOA (1) and the CSA (-)-quinine causes a distinct increase of the chemical shift non-equivalence for the methine

Table 3. Temperature-dependent 'H chemical shifts (6) and chemical shift non-equivalences (A6) for the 2-CH sig- nals' of DIBOA (l)b in the presence of ( -)quinineb in THF-d, at 399.952 MHz

T (lo

298 283 273 263 253 233

d (ppm)

5,73215,743 5.741 15.755 5,75115,770

5.77015.820

5.71 815.725

5.759p.785

M (ppm)

0.007 0.01 1 0.01 4 0.01 9 0.026 0.050

a Signals occur as singlets owing to a complete exchange of the 2-OH proton. b~ = 0.005 mmol ml-' for each.

protons of its enantiomers (Table 3). These data confirm the linearity of a In A6 us. [ T (K)]-' plot. This arises from the fact that a preferred conformer in the dia- stereoisomeric complex is increasingly populated as the temperature is lowered.

However, lowering the temperature of a solution of higher concentration gives rise to the opposite effect, as shown by measuring DIMBOA methyl acetal (4) (c = 0.025 mmol m1-l) under comparable conditions (Table 4). The decrease in chemical shift non- equivalence observed in this case may result from com- peting intermolecular aggregation of hemiacetals which becomes possible at higher concentration and is increas- ingly favoured at lower temperature. However, dia- stereoisomeric solvation complexes with the CSA still exist. Interestingly, at - 30 "C and lower two differen- tiated signals for the 7-OCH3 group remote from the chiral centre of enantiomeric 4 are to be observed. This discrimination may result from the reduced mobility of the solvation complexes at low temperatures. Finally, also a DIBOA (1) solution of higher concentration (0.050 mmol ml- ') shows the phenomenon of decreas- ing chemical shift anisochrony at lower temperatures. However, an exact observation is hindered by the occurrence of doublets due to the CHOH coupling at

Table 4. Temperature-dependent 'H chemical shifts (6) and chemical shift non-equivalences (A6) for the CHOCH, and CHOCH, protons of racemic 4' in the presence of (-)quinine' in THF-d8 at 399.952 MHz

T CHOCH, (K) 6 bpm)

298 3.449 3.500

273 3.457 3.497

263 3.462 3.499

243b 3.471 3.503

223" 3.477 3.503

CHOCH, M (ppm) 6 (wm) M (ppm)

5.337 0.051 5.359 0.022

5.354 0.040 5.370 0.01 6

5.365 0.037 5.379 0.01 4

5.389 0.032 5.397 0.008

5.41 0 0.026 5.41 6 0.006

'C = 0.025 mmol ml-' for each. b3.741 (s. 3H, 7-OCH3); 3.747 (s. 3H. 7-OCH3). '3.731 (S, 3H. 7-OCH3); 3.742 (s, 3H, 7-OCH3).

'H N M R DISCRIMINATION OF ENANTIOMERIC CYCLIC HEMIACETALS AND METHYL ACETALS 73 I

reduced temperatures. Thus, CH singlets for enantio- DIMBOA (2) are the first cyclic hemiacetals to be differ- meric 1 at ambient temperature at 5.713 and 5.716 ppm entiated into enantiomers despite the occurrence of an become doublets (5.770 and 5.780 ppm; J = 6.0 Hz for oxo-cyclo tautomerization which hitherto prevented both) at - 50 "C. enantioseparations by HPLC or HPCE. However, dis-

crimination of enantiomeric thiohemiacetals 7 and 8 could not be accomplished because of interferences by CSA resonances. Generally, maximized chemical shift non-equivalence can be obtained from dilute solutions (0.005 mmol ml - I ) at reduced temperatures.

CONCLUSIONS

The first discrimination of enantiomeric cvclic methvl acetals M, 9 and 10, derived from hydro-xamic acids and lactams with the 2H- 1,4,-benzoxazin-3(4H)-one and

2H-1'4-benzothiazin-3(4H)-one was best The financial support of this investigation by the Studienstiftung des accomP1ished using ( - )-quinine as CSA in a non-polar Deutschen Volkes (for J.K), the Deutsche Forschungsgemeinschaft solvent. Analogously, natural products DIBOA (1) and and the Fonds der Chemischen Industrie i s gratefully acknowledged.

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