Dimeric sesquiterpenoid esters from Chloranthus serratus

0031 9422j92 %5.00+0.00 f> 1992 Pergamon Press plc

Vol. 31, No. 4, pp. 1293 1296, 1992 Phytochemistry, Printed in Great Bntam.

DIMERIC SESQUITERPENOID

ESTERS FROM CHLORANTHUS

JUN KAWABATA

and

JUNYA

SERRA TUS*

MIZUTANI

Department of Agricultural Chemistry, Faculty of Agriculture, Hokkaido University, Kita-Ku, Sapporo 060 Japan (Receiwd Key Word Index-Chloranthus lit lactone.

10

June

1991)

serratus; Chloranthaceae; roots; sesquiterpene dimer; lmdenane skeleton; macrocyc-

Abstract-Three novel dimeric sesquiterpenes, shizukaols, B, C and D, which consist of two lindenane (modified eudesmane) units were isolated from the roots of C’hloranthus serratus. Their structures were elucidated by ID- and 2DNMR analyses and by chemical methods. Among them, shizukaol B has a unique pendent macrocyclic lactone ring. The biogenetic relationship of the lindenane dimers is briefly discussed.

INTRODUCTION

In the course of our continuing search for sesquiterpenes in plants of the Chloranthaceae, a series of unusual sesquiterpene lactones having a lindenane skeleton have been isolated from Chloranthusjaponicus Sieb. [2--41 and C. serratus Roem. et Schult. [4,5]. Recently, we reported that C. juponicus contains a dimeric lindenane named shizukaol A (1) [l]. We now report the isolation and structural elucidation of the related dimer, shizukaol B (Z), which contams a unique macrocyclic lactone rmg as a partial structure. The biogenetically related dimers, shizukaols C (3) and D (4), and 1 were also isolated from C. serratus. RESULTS AND DISCUSSION

Shizukaol B (2) showed a molecular ion at m/z 732 (FD-MS) and the molecular formula was determined to be C4,,H44013 with the aid of the ‘H and 13C NMR data. The presence of a prominent fragment peak at m/z 274 in the FD mass spectrum and of two sets of spin system for 1,Zdisubstituted cyclopropane rings (60.33-1.02-1X-2.06 and 0.74-1.34-1.40-1.61) in the ‘H NMR spectrum revealed 2 to be a lindenane dimer like shizukaol A. The ‘H NMR spectrum of 2 (Table 1) showed a rather complex pattern in the region of 62.5-3.2 compared with the corresponding region of 1. The difference in the molecular formulae of the two compounds (CgHioO,), and the presence of the five ester carbonyl absorptions (6167.0, 170.2, 171.5, 171.7 and 172.0) in the 13C NMR spectrum (Table 2), indicated a highly oxygenated and acylated structure for 2. It should be noted that another low field carbon (6 174.5) in the ester carbonyl region was assigned to be olefinic from the results of the heteronuclear multiple bond correlation (HMBC) spectrum. The HH and CH COSY spectra made it possible to analyse the complex region of the ‘H NMR spectrum. The spin systems of -CH,-CH,and -CH=C(Me)-CH,-, in *Part 8 in the series, ‘Studies on the Chemical Constituents of Chloranthaceae Plants’. For Part 7, see ref. [l].

addition to CHCH,and -CHCH,C=C-CHwhich were also found in 1, were disclosed. The presence of two pairs of AB doublets assignable to isolated methylenes at 64.54 and 5.07 (5=11.9 Hz) and 63.63 and 4.60 (J = 11.4 Hz) in place of an allylic methyl and an exomethylene in 1 suggested that acyloxy groups were substituted at C-13’ and C-15’. Acetylation of 2 gave a monoacetate (5, [Ml+ at m/z 774 by FD-MS) and an acetylation shift observed in H-9 (63.88+5.25) showed that OH-9 in 2 is free. The carbocyclic skeleton of the sesquiterpene dimer portion was unambiguously determined from the results of COLOC experiments. For the pendent acyl groups, two acyl units, succinyl and 4hydroxy-2-methyl-2-butenoyl, were identified from the COLOC data. In addition, alkaline hydrolysis of 2 yielded succinic and y-hydroxytiglic acids which were identified as their methyl esters by GC-MS, whereas efforts to isolate a deacyl-lindenane dimer product failed. The position of the acyl groups was also determined by long range CH correlation experiments (COLOC and HMBC) as follows. One of the nonequivalent methylenes at 64.54 and 5.07, which were assignable to H,-13’ from the correlation with lactonic C-12’ (6 171.7), gave a cross peak with one of the carbonyl carbons (i, 6172.0) of the succinyl unit. Furthermore, another succinyl carbonyl (f, 6 171.5) showed three bond connectivity with y-methylene protons (d, 64.64 and 5.06) of the hydroxytiglyl unit. A cross peak between H,-15’ (63.63 and 4.60) and a carbonyl carbon (a, 6 167.0) of the hydroxytiglyl unit completed the assignments of the acylated part of 2. Finally, lack of H-4’ by hydroxyl substitution accounted for the assignment of the remaining oxygen, and the plane structure was thus determined. The stereochemistry was determined by NOESY experiments. With regard to the dimeric lindenane skeleton, similar NOE interactions were found to those observed for 1 such as H-l/H-9, H-6/H-14, H-6/H-13, H-9/H-14 and H-9/H-5’. In the 4-hydroxy-2-methyl-2-butenoyl substructure, an NOE interaction was observed between the allylic methyl (e, 6 1.92) and the methylene (d, 64.64 and 5.06). Hence, the double bond has the E-configuration. Moreover, an additional NOE between H-3’ and

1293 PHYTO

31:4-A

J. KAWABATA and J.

1294

MIZLJTANI

4

3

2

R,=

R, = OH

OH

d

R, =

R3=

Ri

R2

C

H

R3 =H

Table 1. ‘H NMR spectral data of compounds 2-4 (500 MHz. CDCI,) H

2 (pyridine-d,)

3 (pyridine-d,)

4

1

2.06 (2.33) 1.02 (0.99) 0.33 (0.40) 1.88 (1.93) 3.96 (4 23) 3.88 (4.31) 1.96 (2.14) 1.03 (1.37) 2.81 (2.84) 2.59 (2.66) 1.61 (1.69) 0.74 (0.74) 1 34 (1.62) 1.40(1.22)

ddd (7.6, 7.6, 3.9) ddd (7.6, 7.3, 3.8) ddd (3.9, 3.8. 3.2) m d (3.1) s s s br d (16.2) ddd (16.2, 4.6, 3.1) ddd (8.6, 7.6, 4.0) ddd (8.6, 8.6, 5 3) ddd (5.3. 4.0, 3.4) ddd (8.6, 7.6, 3.4)

2.09 (2.35) 1.02 (0.99) 0.34 (0.40) 1.84 (1.97) 3.93 (4.24) 3 95 (4.44) 1.90(2.17) 1.01 (1.35) 2.83 (2.83) 2.57 (2.69) 1 62 (1.72) 0 71 (0.77) 1.27 (1.55)

1.86 2.50 2.70 1.85

dd (13.3, 6.2) dd (18.5, 6.2) dd (18.5. 13.3)

1.90 2.27 2.70 1.97 4.33

2.06 m 1.00 ddd (7.8, 7.8, 4.2) 0.30 ddd (4.2, 4.2, 3.1) 1.86 m 3.91 br d (3.5) 4.06 s 1.90 s 1.02 5 2.77 dd (16.4, 1.5) 2.61 ddd (16.4, 5.9. 3.5) 1.45 ddd (8.3, 8.1, 3.8) 0.77 ddd (8.3, 8.3, 5.8) 0.83 ddd (5.8, 3.8, 3.6) l.lOdddd(8.3.8.1, 3.6, 3.5) 1.58 dddd (10.9, 8.3, 6.5. 3.5) 1.83 dt (10.9. 10.1 (t)) 2.46 d (10.1 I 2.46 d (10.1) 1.92 dd (5.9.1.5) 4.33 d (13.6) 4.39 d (13.6) 0.66 s 3.78 dd (11.2, 8.3) 3.98 dd (11.2, 6.5) 2.08 s

2a 2P 3 6 9 13 14 15a 15s 1’ 2’r 2’B 3’ 4 5’ 6’~ 6’P 9’ 13’ 13’ 14 15’ 15’ b c d d e g E h

OMe

i2 15) (3.02) (3.21) (1.90)

dd (4.6,0.9)

4.54(4.90)d (11.9) 5.07(5.34)d (11.9) 0.82 (1.05) s 3.63 (3.87) d (11.4) 4.60 (4.89)d (11.4) 6 62 (6.91)br dd (6.9, 4.7)

4 64 5.06 1.92 2.48 2.89 2 67 2.79 3.71

(4.67) (5.05) (1.79) (2.61) (3.11) (2.84) (2.95) (3 67)

dd (14.9. 6.9) dd (14.9, 4.7) s ddd (17.3, 7.3, 2.7) ddd (17.3, 10.2. 2.9) ddd (17.2, 7.3, 2.9) ddd (17 2, 10.2. 2.7) s

complex 1.53 (1.55) ddd (8.8. 7.6, 3.5)

._

(2.15) (2.98) (3.47) (1.98) (4.76)

dd (13.7, 6.1) dd (18.3, 6.1) dd (18.3, 13.7) dd (6.1. 1.5) d (13.7) 441(4.80) d (13.7) 087(1.20) s 3.88 (4.29) d (11.6) 4.23 (4.56) d (11.6) 6.88 (6.92) qq (7.1, 1.0) 1.84 (1.58) d (7.1)

._ 1.85(1.X1)br s

3 79(3.69)s

indicated the orientation of OH-4 was p, which was also supported by the large pyridine-induced solvent shift [6] ofH-2B (A= -0.28)and H-14’(A= -0.23)in the ‘H NMR spectra. Shizukaol C (3) showed a molecular ion at m/z 634 by FD-MS. Characteristic fragments of m/z 360 and 274 strongly suggested this compound to be an acylated H,-15’

ddd (8.0, 7.7, 4.3) ddd (7.7, 7.6, 4.3) ddd (4.3. 4.3, 3.1) r?* d (3.5) s s s dd (16.3, 1.5) ddd (16.3, 6.1, 3.5) ddd (8.5, 7.6, 4.1) ddd (8.8, 8.5, 5.8)

3.79 s

hndenane dimer and the molecular formula was assigned as C,,H,,O,,. The difference of the molecular formula from that of 2 was C,H,O,. In the ‘H NMR spectrum, the complex succinyl proton region at 62.5 -3.0 in 2 was simplified in 3. Furthermore, non-equivalent methylene protons coupled with an olefinic proton in the y-hydroxytiglyl residue in 2 were replaced by an allylic methyl

Sesquiterpenoid

Table

2. 13C NMR spectral

C

2

3

4

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15

26.0 16.0 24.8 142.5 132.2 41.1 131.3 200.7 79.9 51.0 147.5 170.2 20.1 15.3 25.4

26.1 16.0 24.9 142.7 132.3 41.2 130.8 200.0 80.0 51.1 147.5 171.0 20.4 15.6 25.3

25.7 15.9 24.8 142.5 131.5 40.7 131.4 200.5 80.0 51.1 147.3 171.0 20.6 15.3 25.6

C 1’ 2’ 3’ 4 5’ 6 7 8 9’ 10 11’ 12 13’ 14 15’

esters from Chloranthus

data of compounds

2-4 (125 MHz, CDCI,)

2

3

4

25.6 11.7 27.8 77.1 61.2 23.4 174.5 93.2 55.5 44.9 123.4 171.7 54.3 26.0 72.0

25.5 11.8 28.4 77.6 59.9 22.2 168.3 93.6 54.8 44.9 127.4 172.3 54.9 26.1 71.0

24.4 16.6 21.8 43.0 59.1 25.1 168.5 93.4 54.5 44.1 126.7 172.4 55.0 24.0 66.1

doublet (61.84, 5=7.1 Hz) in 3. These results indicated that 3 has a structure which lacks a succinyl residue and a y-hydroxyl of the hydroxytiglyl unit found in 2. Extensive analysis of the 2D-NMR data (HH and HC COSY and HMBC) supported this structure. In addition, methanolysis of 3 gave methyl tiglate which was identified by GC-MS. A strong cross peak of C-a (6 167.0)with H,-15’ (63.63 and 4.60) in the HMBC spectrum unambiguously determined the position of the tiglyl residue on C-15’. NOESY experiments provided evidence that 3 has the same relative stereochemistry as 2. The third compound, shizukaol D (4), showed a molecular ion at m/z 578 (C,,H,,O,). In the ‘H and 13C NMR spectra, there were a set of signals assignable to an acetyl group C62.08 (s), and 620.8 (Me) and 171.1 (CO)]. However, no other signals attributable to acyl moieties were detected. It was also shown that the coupling pattern of one of the cyclopropyl methine protons (61.10) became more complex than seen in the spectrum of 3. In addition, a complex methine proton assignable to H-4’ appeared at 6 1.58 (dddd, J = 10.8, 8.3, 6.5 and 3.5 Hz) and the resulting loss of OH-4’ found in 3 accounted for an upfield shift of H-2’fi (A= -0.44) and H-14’ (A= -0.21) for 4 compared with 3. The stereochemistry was again deduced from the NOESY results to be the same as in 3 except for the C-4’ position. Regarding the C-4’ position, the coupling constants of H-4’ to H-5’ (10.9 Hz) and to H-3’ (3.5 Hz) clearly indicated that the proton at C-4’ is P-orientated [4]. Compound 2 contains a unique macrocyclic la&one ring which is unprecedented in plant constituents. As far as we know, verrucarins [7] isolated as microbial metabolites are the only sesquiterpene compounds containing this type of macrocyclic lactone ring. From a biogenetic point of view, 1 should be first synthesized by a Diels-Alder-type reaction of 6 and 7 ( = shizukanolide B [2]), followed by oxidation at C-13’ and hydration of the 4’/15’ double bond to give 8 as a key intermediate. Acetylation of the 15’-hydroxyl of 8 affords 4, whereas further oxidation at C-4’ and tiglylation of the 15’hydroxyl gives 3. Finally, completion of the macrocyclization by introducing a succinyl unit to C-13’ and to the oxidized y-position of the tiglyl residue of compound 3 yields 2 (Scheme 1).

1295

serratus

C

it i

f” :: i OMe

2

3

4

167.0 129.2 135.5 61.6 13.0 171.5 28.6 29.2 172.0 52.4

168.2 128.0 138.9 14.2 12.1 52.7

171.1 20.8 -

52.7

EXPERIMENTAL Mps: uncorr. ‘H and “C NMR spectra were measured in CDCI, or pyridine-d, using TMS as mt. standard. The air-dried roots (390 g) of C. serratus were extracted with Et20 at room temp. The extracts were washed with 5% NaHCO, and chromatographed over silica gel using a pentane-Et,O-MeOH gradient. The Et,0 eluates were rechromatographed on a silica gel column (pentane-EtzO, 1:l) followed by silica gel prep. TLC (CHCI,-MeOH, 160: 1) to give 1 (6 mg) as needles. The physiocochemical properties of 1 agreed with those of an authentic specimen [l]. The Et,O-MeOH (9:l) eluates of the first column were rechromatographed on silica gel (CHCl,-MeOH, 50: 1) followed by silica gel prep. TLC (CHCl,-MeOH, 30: 1) to give 2 (53 mg), 3 (47 mg) and 4 (17 mg). Shizukaol B (2). Amorphous powder, mp 116-118”; [a]:: - 110” (CHCl,; c 0.28); FDMS m/z (rel. int.): 732 [M]’ (37), 714 (12),459 (17), 274 (lOO);IR v!$!icrn-‘: 3450,1732,1255,1153, 1076,985. Shizukaol B monoacetate (5). Oil, FD-MS m/z (rel. mt.): 774 [M] + (lOO), 316 (56); ‘H NMR 6 (500 MHz, CDCl,): 0.29 (lH, ddd,5=4.5,4.3,3.0Hz,H-2B),0.74(1H,ddd,J=8.8,8.6,5.8Hz, H-2’a), 0.83 (3H, s, H-14’), 1.00 (lH, ddd, J=8.2, 7.8,4.5 Hz., H2a), 1.16 (3H, s, H-14), 1.35 (lH, m, H-2’@, 1.5-1.6 (2H, complex, H-l’ and H-3’), 1.68 (lH, ddd, J=8.2. 5.7,4.3 Hz., H-l), 1.72 (IH, dd,J=13.3,6.5 Hz,H-5’), 1.76(1H,dd,J=6.0,1.9 Hz,H-9’), 1.87 (lH, m, H-3), 1.87 (3H, d, J = 1.0 Hz, H-e), 1.95 (3H, s, H-13), 2.21 (3H, s, OAc), 2.4-2.7 (5H, m), 2.75 (lH, br d, J= 16.9 Hz, H-15a), 2.8-2.9 (2H, m), 3.73 (3H, s, OMe), 3.79 (lH, d, J= 12.5 HZ, H15’), 3.95 (lH, br d, 5=3.5 Hz, H-6), 4.57 (lH, d, 5=12.0 Hz, H13’),4.61 (lH,brd,J=16.0,4.3 Hz,H-d),4.85(1H,d,J=12.5 Hz, H-15’), 5.00 (lH, d, 5=12.0 Hz, H-133, 5.05 (lH, dd, 5=16.0, 5.3 Hz, H-d), 5.25 (lH, s, H-9), 6.68 [lH, ddq, 5=5.3,4.3, 1.0 (q) Hz, H-c]. Alkaline-hydrolysis of compound 2. To a soln of 2 (10 mg) in MeOH (1 ml) was added a soln of K,CO, (130mg) in Hz0 (0.5 ml). After 3 hr at room temp., the mixt. was acidified and extracted with EtOAc. The extract was coned and methylated with ethereal CH,N,. The methylated products were directly subjected to the GLC analysis (5% PEG-ZOM, 1 m) to give two peaks. GC-MS m/z (rel. int.), peak 1: 115 [M -OMe]+ (lOO), 87 [M-CO,Me]+ (31), 59(26), 55 (45); peak 2: 101 [M-CHO]+ (lOO), 98 [M-OMe]+ (54), 71 (41), 69 (53). These data were identical with those of authentic dimethyl succinate (peak 1) and

1296

J. KAWABATA

and J. MIZUTANI

/

Scheme

1. Plausible

biogenetic

pathway

methyl y-hydroxytlglate (peak 2) which was synthesized by the method of ref. r81. Shizukaol C (3,. Powder, mp 104-106”; [a]~z-6S” (CHCI,; c 1.9); FD-MS m/z (rel. int.): 634 [M]’ (71), 360(70), 274(100); IR vfzi cm -‘: 3420, 2920, 1735, 1260, 1078. Methanolysis ofcompound 3. To a soln of 3 (5.6 mg) in 3 drops of MeOH was added a drop of 28% NaOMe in MeOH. After 2 hr at room temp., the mixt. was acidified and extracted with Et,O. The extract gave a single peak by the GLC analysis. GCMS m/: (rel. int.): 114 CM]’ (48), 99 [M-Me]+ (16), 83 [M -OMe] ’ (59), 55 (100). These data were identical with those of authentic methyl @ate. Shizukaol D (4). 011, [a]y - 174” (CHCI,; c 1.l); FD-MS m/z (rel. mt.): 578 [M]’ (100). 304 (23), 274 (44); IR vk\y cm-‘: 3430, 2930, 1730, 1600, 1225.

Acknowledgements- We are gratiful to Mr Kenji Watanabe and MS Eri Fukushi (GC-MS & NMR Laboratory of our faculty), for measuring mass spectra. Part of this work was supported by a

of lindenane

Grant-in-Aid the Ministry

dimers in C. serratus.

for Scientific Research (No. 02454064 to J. K.) from of Education, Science and Culture of Japan.

REFERENCES J., Fukushi, Y., Tahara, S. and Mizutani, J. (1990) 1. Kawabata, Phytochemistry 29, 2332. 2. Kawabata, J., Tahara, S. and Mizutani, J. (1981) Agric. Biol. Chem. 45, 1447. 3. Tahara, S., Fukushi, Y., Kawabata, J. and Mizutani, J. (1981) Agric. Biol. Chem. 45, 1511. 4. Kawabata, J. and Mizutani, J. (1989) Agric. Biol. Chem. 53, 203. 5. Kawabata, J., Fukushi, Y., Tahara, S. and Mizutani, J. (1985) Agric. Biol. Chem. 49, 1479. 6. Demarco, P. V.. Farcus, E.. Doddrell, D., Mylari, B. L. and Wenkert, E. (1968) J. Am. Chem. Sot. 90, 5480. 7. Gutzwiller, J. and Tamm, Ch. (1965) Helo. Chim. Acta 48,157; Breitenstein, W. and Tamm, Ch. (1977) Helo. Chim. Acta 60, 1522. 8. Ishida, A. and Mukaiyama, T. (1978) Bull. Chem. Sot. Jpn 51, 2077.