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Five naphthalene glycosides from the roots of Rumex patientia
Phytochemistry 56 (2001) 399±402
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Five naphthalene glycosides from the roots of Rumex patientia OÈmuÈr Demirezer a,*, AysË e KuruuÈzuÈm a, Isabelle Bergere b, H.-J. Schiewe b, Axel Zeeck b a
Department of Pharmacognosy, Faculty of Pharmacy, Hacettepe University, TR-06100 Ankara, Turkey b Institute of Organic Chemistry, Georg-August University, D-37077 GoÈttingen, Germany Received 30 May 2000; received in revised form 14 August 2000
Abstract Three novel and two known naphthalene glycosides were isolated from the roots of Rumex patientia L. (Polygonaceae). The structures of the new compounds were established, respectively as 2-acetyl-3-methyl-6-carboxy-1,8-dihydroxynaphthalene-8-O-b-dglucopyranoside, 4,400 -binaphthalene-8,800 -O,O-di-b-d-glucopyranoside and 2-acetyl-3-methyl-1,8-dihydroxynaphthalene-8-O-b-dglucopyranosyl (1!3) b-d-glucopyranoside on the basis of spectral analysis. The other napthalene glycosides were determined as nepodin-8-O-b-d-glucopyranoside and torachrysone-8-O-b-d-glucopyranoside by comparison of their spectral data with those previously reported. # 2001 Elsevier Science Ltd. All rights reserved. Keywords: Rumex patientia; Polygonaceae; Naphthalene glycosides; Rumexoside; Labadoside; Orientaloside
1. Introduction Rumex species are wide spread plants in the ¯ora of Turkey (Davis, 1967, 1988). The roots of Rumex patientia are used in some parts of Turkey for its purgative, constipate, depurative and tonic properties (Baytop, 1984). Chemical studies on R. patientia have been carried out by some investigators and the presence of the anthraquinone, naphthalene, tannin and napthoquinone derivatives has been reported (Sharma et al., 1977; Suri et al., 1978; Demirezer and KuruuÈzuÈm, 1997). Here, we report the isolation from this plant of ®ve naphthalene glycosides, including the structure elucidation of three new compounds (1±3). 2. Results and discussion Five naphthalene glycosides (1±5) were isolated from the MeOH extract of the roots of R. patientia. Compounds 1±3 were described as 2-acetyl-3-methyl-6-carboxy-1,8-dihydroxynaphthalene-8-O-b-d-glucopyranoside * Corresponding author. Tel.: +90-312-305-10-89; fax: 90-312-31147-77. E-mail address: [email protected] (O. Demirezer).
(1), 4,400 -binaphthalene-8,800 -O,O-di-b-d-glucopyranoside (2) and 2-acetyl-3-methyl-1,8-dihydroxynaphthalene-8O-b-d-glucopyranosyl (1!3) b-d-glucopyranoside (3) on the basis of spectral analysis. Naphthalenes 4 and 5 were identi®ed as nepodin-8-O-b-d-glucopyranoside and torachrysone-8-O-b-d-glucopyranoside by comparison of their spectral data with those previously reported (Tsuboi et al., 1977; Nishina et al., 1991). Compounds 1±3 were obtained as amorphous materials. The UV spectra of 1±3 suggested that their hydroxy naphthalene were observed. The IR spectrum of compound 1 showed the presence of a carboxy group (1732 cmÿ1). Its mass spectrum exhibited the [MÿH]+ peak at m/z 421 and [MÿCOOH]+ peak at m/z 377. In the 13C NMR spectrum, a carbon signal at 127.80 proved the presence of an carboxy group at C-6 in compound 1. Generally 1D (Table 1) and 2D NMR spectral data of 1 were similar to those of nepodin-8-glucoside (4) except for the presence of signals due to the carboxy moiety. Thus compound 1 was assigned as 2-acetyl-3-methyl-6-carboxy1,8-dihydroxynaphthalene-8-O-b-d-glucopyranoside. In the ESI mass spectrum of compound 2, the peak of highest mass number was observed at m/z 777 [M+Na]+ con®rming the molecular weight of 2 corresponding to the molecular formula C38H42O16. Its UV
0031-9422/01/$ - see front matter # 2001 Elsevier Science Ltd. All rights reserved. PII: S0031-9422(00)00368-X
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O. Demirezer et al. / Phytochemistry 56 (2001) 399±402
Table 1 13 C and 1H NMR spectral data for compounds 1 and 3a Compound 1
Compound 3
Position
C atom
C (ppm)
H (ppm), J (Hz)
1 2 3 4 5 6 7 8 9 10 3-CH3 2-COCH3 2-COCH3 6-COOH Glucose 10 20 30 40 50 60
C C C CH CH C CH C C C CH3 CH3 C C
153.02 125.18 133.55 118.17 127.34 127.80 122.59 154.59 111.09 136.30 19.63 31.65 205.16 170.50
± ± ± 7.14 7.23 ± 7.41 ± ± ± 2.22 2.50 ± ±
CH CH CH CH CH CH2
103.02 73.45 74.43 69.97 76.03 63.26
5.03 d (7.3) 3.24±3.68b 3.24±3.68b 3.24±3.68b 3.24±3.68b 4.12 dd (12/6.8) 4.31 dd
CH CH CH CH CH CH2
± ± ± ± ± ±
± ± ± ± ± ±
Glucose 100 200 300 400 500 600 a b c d
HMBC
s d (4)
C-2, CH3 C-7
d (4)
C-8, C-9
s s
C-2, C-3, C-4 COCH3
C-8
C atom
C (ppm)
H (ppm), J (Hz)
C C C CH CH CH CH C C C CH3 CH3 C ±
152.65 126.57 138.94 118.56 123.99 128.62 112.92 157.14 114.30 135.14 20.47 31.13 208.26 ±
± ± ± 7.14 7.40 7.42 7.36 ± ± ± 2.29 2.65 ± ±
CH CH CH CH CH CH2
105.30 73.65c 75.04 71.69 78.09 62.55 3.74 m
CH CH CH CH CH CH2
99.99 73.71c 75.24 71.42 77.25 67.51
HMBC
s m m m
C-2, C-5, CH3 C-4, C-8 C-8, C-10 C-5
s s
C-2, C-3, C-4
5.14 3.61 3.57 3.31 3.49 3.62
d (7.8) md m m m m
C-8
4.90 3.63 3.71 3.48 3.76 3.84 3.96
d (7.5) mc,d m m m m m
C-30
Assigments are based on APT, COSY, HMQC and HMBC. Signal pattern unclear due to overlapping. Values for C-20 and C-200 can be interchanged. Values for H-20 and H-200 can be interchanged.
and IR spectra indicated the presence of a hydroxy naphthalene skeleton. The 1H NMR spectrum (Table 2) of 2 was similar to that of 4 whereas its 1H NMR spectrum contains resonances for two anomeric protons of two sugar units at 5.10 (d, H-10 , J=7.0 Hz) and 5.11 (d, H-100 , J=7.0 Hz). The anomeric proton and other proton signals of sugars are characteristic for glucose. The anomeric con®guration of glucose was proposed to be b on the basis of the coupling constant (J=7 Hz) and the chemical shifts. The 1H NMR spectrum of 2 was signi®cant in that it indicated duplication of resonances attributable to the glucosidic naphthalene function. 13C NMR assigments were supported by the APT experiment are shown in Table 2. 1H, 1H-Homonuclear COSY spectrum of 2 showed the symetrically same spin system for double naphthalene ring and double sugar units. The assigments for all protonated carbons and protons of 2 were determined with short range and long range heteronuclear correlations spectra (HMQC and
HMBC). Thus the dimeric naphthalene glucosides were clearly de®ned in the COSY, HMQC and HMBC spectra of compound 2. However, the long range 1H, 13C correlations of H-10 with C-8 and H-1000 with C-800 supports the position of the attachment of the sugars. The lack of the typical H-4/H-400 signals in the 1H NMR spectrum of 2 suggested that C-4 and C-400 are the positions of dimerization. In addition, the down®eld shift (=5.16 ppm) in the 13C NMR spectra of C-4/C-400 in comparison to that of nepodin-8-glucoside (4) is supporting this point. Evidence for the dimer glucosidic naphthalene structure was con®rmed by a combination of the mass, 1D NMR (1H, 13C) and 2D NMR (COSY, HMQC, HMBC) spectrometry. Therefore, compound 2 was determined as 4,400 -binaphthalene-8,800 -O,O-di-b-dglucopyranoside. The UV and IR spectra of compound 3 were similar to those of 1. Its 1H and 13C NMR spectra (Table 1) displayed many similarities with those of 4, especially
O. Demirezer et al. / Phytochemistry 56 (2001) 399±402
for the resonances assigned to the naphthalene moiety and the glucose unit. However, the set of additional protons, apart from the b-anomeric proton at 4.90 (d, J=7.5 Hz) and the corresponding carbon signals, were in agreement with the presence of another hexose unit, which was identi®ed as b-glucose by COSY and HMQC experiments. The heteronuclear multiple bond correlation (HMBC) experiment observed between H-10 of inner glucose and C-8 of the naphthalene aglycone and H-100 of terminal glucose and C-30 of the inner glucose. Consequently, compound 3 was established as 2-acetyl3-methyl-1,8-dihydroxynaphthalene-8-O-b-d-glucopyranosyl (1!3) b-d-glucopyranoside. Compound 1±3 were isolated from nature for the ®rst time, for which rumexoside, labadoside, and orientaloside are proposed as the trivial names, respectively. To our knowledge this is the ®rst report of the occurrence of the dimeric type naphthalene in the genus Rumex until now.
401
3. Experimental UV spectra were determined in spectroscopic grade MeOH on a Shimadzu UV-160 A spectrophotometer. IR spectra were measured on a Perkin-Elmer FTIR 1720 X spectrometer as pressed KBr disks. NMR spectra were recorded on Bruker AMX 300 and AMX 500 NMR, operating at 300 and 500 MHz for proton and 75.5 MHz for carbon. EIMS (70 eV) and DCIMS were recorded on Finnigan MAT 311 A and Varian MAT 731, respectively. TLC was carried out on precoated silica gel 60 F 254 aluminium sheets (Merck). For column chromatography, normal phase silica gel 60 (0.063±0.200 mm, Merck), reversed phase silica gel (Li Chroprep RP-18, Merck), and polyamide (PolyamidMN-Polyamid SC 6, Macherey-Nagel, DuÈren) were used. Compounds were detected by UV ¯ourescence and/or spraying with vanillin±H2SO4 reagent followed by heating at 100 C for 5±10 min. 3.1. Plant material The roots of R. patientia L. were collected from BorNigÆde (1050 m), Turkey in September 1996. A voucher specimen has been deposited at the herbarium of the Faculty of Pharmacy, Hacettepe University, Ankara, Turkey (HUEF 96003). Table 2 13 C and 1H NMR spectral data for compound 2a Position 00
1, 1 2, 200 3, 300 4, 400 5 500 6 600 7 700 8, 800 9, 900 10, 1000 3-CH3 300 -CH3 2, 200 -COCH3 2, 200 -COCH3 Glucose 10 1000 20 , 2000 30 , 3000 40 , 4000 50 , 5000 60 , 6000 a b
C atom
C (ppm)
H (ppm), J (Hz)
C C C C CH CH CH CH CH CH C C C CH3 CH3 CH3 C
158.47 126.32 132.10 126.11 120.49 120.49 128.01 128.01 111.04 111.04 154.95 113.83 135.18 16.70 16.64 31.36 205.88
± ± ± ± 6.57 6.53 7.22 7.20 7.33 7.32 ± ± ± 1.75 1.74 2.59 ±
CH CH CH CH CH CH CH2
102.83 102.83 73.55 76.32 69.92 77.85 60.81
5.11 d (7) 5.10 d (7) 3.17±3.80b 3.17±3.80b 3.17±3.80b 3.17±3.80b 3.17±3.80b
d (8) d (8) dd (8/8) dd (8/8) d (8) d (8)
s s s
HMBC
C-4, C-7, C-9 C-8, C-10 C-5, C-8, C-9
C-2, C-3, C-4 COCH3 C-8 C-800
Assignments are based on APT, COSY, HMQC and HMBC. Signal pattern unclear due to overlapping.
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3.2. Extraction and isolation
3.4. Labadoside (2)
Powdered dried material (500 g) was extracted with MeOH (32.5 l) at 40 C. The MeOH extracts were combined and evaporated to dryness in vacuo (100 g). The crude extract (40 g) was fractionated by column chromatography on polyamide by gradient elution with H2O±MeOH mixtures. The fractions eluted with 50% MeOH in water were puri®ed by repeated silica gel column chromatographies eluted with CHCl3±MeOH (95:5, 90:10) to give compound 1 (8 mg). The fractions eluted with 60% MeOH were subjected to a polyamide column, eluting with H2O±MeOH (60:40!20:80) to give 13 fractions. The fractions 5±7 were rechromatographed over silica gel columns using EtOAc±MeOH±H2O (100:5:1!100:17:13) solvent system and a sephadex LH-20 column using MeOH to yield compound 2 (8.6 mg). The fractions eluted with 80% MeOH were rechromatographed an a polyamide column using MeOH gradient as eluent and a silica gel column using EtOAc± MeOH±H2O (100:5:2!100:17:13) solvent system to give compound 3 (16 mg). The fractions eluted with 70% MeOH were applied to a polyamide column using increasing amounts of MeOH in H2O (40±100% MeOH). The fractions eluted with 40±60% MeOH were subjected to MPLC using RP-18 material as stationary phase and H2O±MeOH gradient as solvent system (30±100%). Fractions 33±36 were further puri®ed by silica gel column using EtOAc± MeOH±H2O (100:17:13) to yield compounds 4 (12 mg) and 5 (14 mg).
Amorphous; ESIMS m/z: 777 [M+Na]+, 615 [(MÿC6H10O5)+Na]+; UV lmax (MeOH) nm: 230, 314, 337; IR max (KBr) cmÿ1: 3399 (OH), 1693 (CO), 1621 (CÿC), 1078 (CÿO); 1H (300 MHz; DMSO±D2O) and 13 C NMR (75.5 MHz; DMSO±D2O): Table 2.
3.3. Rumexoside (1) Amorphous; ESIMS m/z: 868 [2M+Na]+, 445 [M+Na]+, 421 [MÿH]ÿ, 377 [MÿCOOH]ÿ, 215 [(MÿC6H10O5)±COOH]ÿ; UV lmax (MeOH) nm: 224, 260, 301, 334; IR max (KBr) cmÿ1: 3399 (OH), 1732 (COOH), 1690 (CO), 1605 (CC, arom.), 1079 (CÿO); 1H (300 MHz; DMSO±D2O) and 13C NMR (75.5 MHz; DMSO-d6): Table 1.
3.5. Orientaloside (3) Amorphous; DCIMS m/z: 234 [aglycon+NH4]+, 217 [aglycon+H]+, 197 [C6H10O5+NH4+NH3]+, 180 [C6H10O5+NH4]+; UV lmax (MeOH) nm: 225, 260, 301, 333, 345; IR max (KBr) cmÿ1: 3400 (OH), 1632 (CÿC), 1028 (CÿO); 1H (300 MHz; CD3OD) and 13C NMR (75.5 MHz; CD3OD): Table 1. Acknowledgements This . study was supported partly by grants from TUÈBITAK-BAYG (Scienti®c and Technical Research Council of Turkey), Turkey and Researching Project Programs of Hacettepe University (98T01102011), Turkey. References Baytop, T., 1984. Therapy with Medicinal Plants (Past and Present). Nobel Tip Kitabevleri, Istanbul. Davis, P.H., 1967. Flora of Turkey and the East Aegean Islands. University Press, Edinburgh. Davis, P.H., 1988. Flora of Turkey and the East Aegean Islands. University Press, Edinburgh. Demirezer, LOÈ, KuruuÈzuÈm, A., 1997. A comparative chemotaxonomic study on eleven Rumex species growing in Turkey. FABAD Journal of Pharmaceutical Sciences 22, 153±158. Nishina, A., Kubota, K., Kameoka, H., Osawa, T., 1991. Antioxidizing component, musizin, in Rumex japonicus Houtt. Journal of American Oil Chemical Society 68, 735±739. Sharma, M., Sharma, P., Rangaswami, S., 1977. Orientalone, a new 1,4-naphthoquinone from Rumex orientalis. Indian Journal of Chemistry 15B, 544±545. Suri, J.L., Dhar, K.L., Atal, C.K., 1978. Chemical constituents of Rumex orientalis Bernh. Journal of Indian Chemical Society 55, 292±293. Tsuboi, M., Minami, M., Nonaka, G.I., Nishioka, I., 1977. Studies on Rhubarb (Rhei Rhizoma) IV. Naphthalene glycosides. Chemical and Pharmaceutical Bulletin 25, 2708±2712.
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Five naphthalene glycosides from the roots of Rumex patientia OÈmuÈr Demirezer a,*, AysË e KuruuÈzuÈm a, Isabelle Bergere b, H.-J. Schiewe b, Axel Zeeck b a
Department of Pharmacognosy, Faculty of Pharmacy, Hacettepe University, TR-06100 Ankara, Turkey b Institute of Organic Chemistry, Georg-August University, D-37077 GoÈttingen, Germany Received 30 May 2000; received in revised form 14 August 2000
Abstract Three novel and two known naphthalene glycosides were isolated from the roots of Rumex patientia L. (Polygonaceae). The structures of the new compounds were established, respectively as 2-acetyl-3-methyl-6-carboxy-1,8-dihydroxynaphthalene-8-O-b-dglucopyranoside, 4,400 -binaphthalene-8,800 -O,O-di-b-d-glucopyranoside and 2-acetyl-3-methyl-1,8-dihydroxynaphthalene-8-O-b-dglucopyranosyl (1!3) b-d-glucopyranoside on the basis of spectral analysis. The other napthalene glycosides were determined as nepodin-8-O-b-d-glucopyranoside and torachrysone-8-O-b-d-glucopyranoside by comparison of their spectral data with those previously reported. # 2001 Elsevier Science Ltd. All rights reserved. Keywords: Rumex patientia; Polygonaceae; Naphthalene glycosides; Rumexoside; Labadoside; Orientaloside
1. Introduction Rumex species are wide spread plants in the ¯ora of Turkey (Davis, 1967, 1988). The roots of Rumex patientia are used in some parts of Turkey for its purgative, constipate, depurative and tonic properties (Baytop, 1984). Chemical studies on R. patientia have been carried out by some investigators and the presence of the anthraquinone, naphthalene, tannin and napthoquinone derivatives has been reported (Sharma et al., 1977; Suri et al., 1978; Demirezer and KuruuÈzuÈm, 1997). Here, we report the isolation from this plant of ®ve naphthalene glycosides, including the structure elucidation of three new compounds (1±3). 2. Results and discussion Five naphthalene glycosides (1±5) were isolated from the MeOH extract of the roots of R. patientia. Compounds 1±3 were described as 2-acetyl-3-methyl-6-carboxy-1,8-dihydroxynaphthalene-8-O-b-d-glucopyranoside * Corresponding author. Tel.: +90-312-305-10-89; fax: 90-312-31147-77. E-mail address: [email protected] (O. Demirezer).
(1), 4,400 -binaphthalene-8,800 -O,O-di-b-d-glucopyranoside (2) and 2-acetyl-3-methyl-1,8-dihydroxynaphthalene-8O-b-d-glucopyranosyl (1!3) b-d-glucopyranoside (3) on the basis of spectral analysis. Naphthalenes 4 and 5 were identi®ed as nepodin-8-O-b-d-glucopyranoside and torachrysone-8-O-b-d-glucopyranoside by comparison of their spectral data with those previously reported (Tsuboi et al., 1977; Nishina et al., 1991). Compounds 1±3 were obtained as amorphous materials. The UV spectra of 1±3 suggested that their hydroxy naphthalene were observed. The IR spectrum of compound 1 showed the presence of a carboxy group (1732 cmÿ1). Its mass spectrum exhibited the [MÿH]+ peak at m/z 421 and [MÿCOOH]+ peak at m/z 377. In the 13C NMR spectrum, a carbon signal at 127.80 proved the presence of an carboxy group at C-6 in compound 1. Generally 1D (Table 1) and 2D NMR spectral data of 1 were similar to those of nepodin-8-glucoside (4) except for the presence of signals due to the carboxy moiety. Thus compound 1 was assigned as 2-acetyl-3-methyl-6-carboxy1,8-dihydroxynaphthalene-8-O-b-d-glucopyranoside. In the ESI mass spectrum of compound 2, the peak of highest mass number was observed at m/z 777 [M+Na]+ con®rming the molecular weight of 2 corresponding to the molecular formula C38H42O16. Its UV
0031-9422/01/$ - see front matter # 2001 Elsevier Science Ltd. All rights reserved. PII: S0031-9422(00)00368-X
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O. Demirezer et al. / Phytochemistry 56 (2001) 399±402
Table 1 13 C and 1H NMR spectral data for compounds 1 and 3a Compound 1
Compound 3
Position
C atom
C (ppm)
H (ppm), J (Hz)
1 2 3 4 5 6 7 8 9 10 3-CH3 2-COCH3 2-COCH3 6-COOH Glucose 10 20 30 40 50 60
C C C CH CH C CH C C C CH3 CH3 C C
153.02 125.18 133.55 118.17 127.34 127.80 122.59 154.59 111.09 136.30 19.63 31.65 205.16 170.50
± ± ± 7.14 7.23 ± 7.41 ± ± ± 2.22 2.50 ± ±
CH CH CH CH CH CH2
103.02 73.45 74.43 69.97 76.03 63.26
5.03 d (7.3) 3.24±3.68b 3.24±3.68b 3.24±3.68b 3.24±3.68b 4.12 dd (12/6.8) 4.31 dd
CH CH CH CH CH CH2
± ± ± ± ± ±
± ± ± ± ± ±
Glucose 100 200 300 400 500 600 a b c d
HMBC
s d (4)
C-2, CH3 C-7
d (4)
C-8, C-9
s s
C-2, C-3, C-4 COCH3
C-8
C atom
C (ppm)
H (ppm), J (Hz)
C C C CH CH CH CH C C C CH3 CH3 C ±
152.65 126.57 138.94 118.56 123.99 128.62 112.92 157.14 114.30 135.14 20.47 31.13 208.26 ±
± ± ± 7.14 7.40 7.42 7.36 ± ± ± 2.29 2.65 ± ±
CH CH CH CH CH CH2
105.30 73.65c 75.04 71.69 78.09 62.55 3.74 m
CH CH CH CH CH CH2
99.99 73.71c 75.24 71.42 77.25 67.51
HMBC
s m m m
C-2, C-5, CH3 C-4, C-8 C-8, C-10 C-5
s s
C-2, C-3, C-4
5.14 3.61 3.57 3.31 3.49 3.62
d (7.8) md m m m m
C-8
4.90 3.63 3.71 3.48 3.76 3.84 3.96
d (7.5) mc,d m m m m m
C-30
Assigments are based on APT, COSY, HMQC and HMBC. Signal pattern unclear due to overlapping. Values for C-20 and C-200 can be interchanged. Values for H-20 and H-200 can be interchanged.
and IR spectra indicated the presence of a hydroxy naphthalene skeleton. The 1H NMR spectrum (Table 2) of 2 was similar to that of 4 whereas its 1H NMR spectrum contains resonances for two anomeric protons of two sugar units at 5.10 (d, H-10 , J=7.0 Hz) and 5.11 (d, H-100 , J=7.0 Hz). The anomeric proton and other proton signals of sugars are characteristic for glucose. The anomeric con®guration of glucose was proposed to be b on the basis of the coupling constant (J=7 Hz) and the chemical shifts. The 1H NMR spectrum of 2 was signi®cant in that it indicated duplication of resonances attributable to the glucosidic naphthalene function. 13C NMR assigments were supported by the APT experiment are shown in Table 2. 1H, 1H-Homonuclear COSY spectrum of 2 showed the symetrically same spin system for double naphthalene ring and double sugar units. The assigments for all protonated carbons and protons of 2 were determined with short range and long range heteronuclear correlations spectra (HMQC and
HMBC). Thus the dimeric naphthalene glucosides were clearly de®ned in the COSY, HMQC and HMBC spectra of compound 2. However, the long range 1H, 13C correlations of H-10 with C-8 and H-1000 with C-800 supports the position of the attachment of the sugars. The lack of the typical H-4/H-400 signals in the 1H NMR spectrum of 2 suggested that C-4 and C-400 are the positions of dimerization. In addition, the down®eld shift (=5.16 ppm) in the 13C NMR spectra of C-4/C-400 in comparison to that of nepodin-8-glucoside (4) is supporting this point. Evidence for the dimer glucosidic naphthalene structure was con®rmed by a combination of the mass, 1D NMR (1H, 13C) and 2D NMR (COSY, HMQC, HMBC) spectrometry. Therefore, compound 2 was determined as 4,400 -binaphthalene-8,800 -O,O-di-b-dglucopyranoside. The UV and IR spectra of compound 3 were similar to those of 1. Its 1H and 13C NMR spectra (Table 1) displayed many similarities with those of 4, especially
O. Demirezer et al. / Phytochemistry 56 (2001) 399±402
for the resonances assigned to the naphthalene moiety and the glucose unit. However, the set of additional protons, apart from the b-anomeric proton at 4.90 (d, J=7.5 Hz) and the corresponding carbon signals, were in agreement with the presence of another hexose unit, which was identi®ed as b-glucose by COSY and HMQC experiments. The heteronuclear multiple bond correlation (HMBC) experiment observed between H-10 of inner glucose and C-8 of the naphthalene aglycone and H-100 of terminal glucose and C-30 of the inner glucose. Consequently, compound 3 was established as 2-acetyl3-methyl-1,8-dihydroxynaphthalene-8-O-b-d-glucopyranosyl (1!3) b-d-glucopyranoside. Compound 1±3 were isolated from nature for the ®rst time, for which rumexoside, labadoside, and orientaloside are proposed as the trivial names, respectively. To our knowledge this is the ®rst report of the occurrence of the dimeric type naphthalene in the genus Rumex until now.
401
3. Experimental UV spectra were determined in spectroscopic grade MeOH on a Shimadzu UV-160 A spectrophotometer. IR spectra were measured on a Perkin-Elmer FTIR 1720 X spectrometer as pressed KBr disks. NMR spectra were recorded on Bruker AMX 300 and AMX 500 NMR, operating at 300 and 500 MHz for proton and 75.5 MHz for carbon. EIMS (70 eV) and DCIMS were recorded on Finnigan MAT 311 A and Varian MAT 731, respectively. TLC was carried out on precoated silica gel 60 F 254 aluminium sheets (Merck). For column chromatography, normal phase silica gel 60 (0.063±0.200 mm, Merck), reversed phase silica gel (Li Chroprep RP-18, Merck), and polyamide (PolyamidMN-Polyamid SC 6, Macherey-Nagel, DuÈren) were used. Compounds were detected by UV ¯ourescence and/or spraying with vanillin±H2SO4 reagent followed by heating at 100 C for 5±10 min. 3.1. Plant material The roots of R. patientia L. were collected from BorNigÆde (1050 m), Turkey in September 1996. A voucher specimen has been deposited at the herbarium of the Faculty of Pharmacy, Hacettepe University, Ankara, Turkey (HUEF 96003). Table 2 13 C and 1H NMR spectral data for compound 2a Position 00
1, 1 2, 200 3, 300 4, 400 5 500 6 600 7 700 8, 800 9, 900 10, 1000 3-CH3 300 -CH3 2, 200 -COCH3 2, 200 -COCH3 Glucose 10 1000 20 , 2000 30 , 3000 40 , 4000 50 , 5000 60 , 6000 a b
C atom
C (ppm)
H (ppm), J (Hz)
C C C C CH CH CH CH CH CH C C C CH3 CH3 CH3 C
158.47 126.32 132.10 126.11 120.49 120.49 128.01 128.01 111.04 111.04 154.95 113.83 135.18 16.70 16.64 31.36 205.88
± ± ± ± 6.57 6.53 7.22 7.20 7.33 7.32 ± ± ± 1.75 1.74 2.59 ±
CH CH CH CH CH CH CH2
102.83 102.83 73.55 76.32 69.92 77.85 60.81
5.11 d (7) 5.10 d (7) 3.17±3.80b 3.17±3.80b 3.17±3.80b 3.17±3.80b 3.17±3.80b
d (8) d (8) dd (8/8) dd (8/8) d (8) d (8)
s s s
HMBC
C-4, C-7, C-9 C-8, C-10 C-5, C-8, C-9
C-2, C-3, C-4 COCH3 C-8 C-800
Assignments are based on APT, COSY, HMQC and HMBC. Signal pattern unclear due to overlapping.
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O. Demirezer et al. / Phytochemistry 56 (2001) 399±402
3.2. Extraction and isolation
3.4. Labadoside (2)
Powdered dried material (500 g) was extracted with MeOH (32.5 l) at 40 C. The MeOH extracts were combined and evaporated to dryness in vacuo (100 g). The crude extract (40 g) was fractionated by column chromatography on polyamide by gradient elution with H2O±MeOH mixtures. The fractions eluted with 50% MeOH in water were puri®ed by repeated silica gel column chromatographies eluted with CHCl3±MeOH (95:5, 90:10) to give compound 1 (8 mg). The fractions eluted with 60% MeOH were subjected to a polyamide column, eluting with H2O±MeOH (60:40!20:80) to give 13 fractions. The fractions 5±7 were rechromatographed over silica gel columns using EtOAc±MeOH±H2O (100:5:1!100:17:13) solvent system and a sephadex LH-20 column using MeOH to yield compound 2 (8.6 mg). The fractions eluted with 80% MeOH were rechromatographed an a polyamide column using MeOH gradient as eluent and a silica gel column using EtOAc± MeOH±H2O (100:5:2!100:17:13) solvent system to give compound 3 (16 mg). The fractions eluted with 70% MeOH were applied to a polyamide column using increasing amounts of MeOH in H2O (40±100% MeOH). The fractions eluted with 40±60% MeOH were subjected to MPLC using RP-18 material as stationary phase and H2O±MeOH gradient as solvent system (30±100%). Fractions 33±36 were further puri®ed by silica gel column using EtOAc± MeOH±H2O (100:17:13) to yield compounds 4 (12 mg) and 5 (14 mg).
Amorphous; ESIMS m/z: 777 [M+Na]+, 615 [(MÿC6H10O5)+Na]+; UV lmax (MeOH) nm: 230, 314, 337; IR max (KBr) cmÿ1: 3399 (OH), 1693 (CO), 1621 (CÿC), 1078 (CÿO); 1H (300 MHz; DMSO±D2O) and 13 C NMR (75.5 MHz; DMSO±D2O): Table 2.
3.3. Rumexoside (1) Amorphous; ESIMS m/z: 868 [2M+Na]+, 445 [M+Na]+, 421 [MÿH]ÿ, 377 [MÿCOOH]ÿ, 215 [(MÿC6H10O5)±COOH]ÿ; UV lmax (MeOH) nm: 224, 260, 301, 334; IR max (KBr) cmÿ1: 3399 (OH), 1732 (COOH), 1690 (CO), 1605 (CC, arom.), 1079 (CÿO); 1H (300 MHz; DMSO±D2O) and 13C NMR (75.5 MHz; DMSO-d6): Table 1.
3.5. Orientaloside (3) Amorphous; DCIMS m/z: 234 [aglycon+NH4]+, 217 [aglycon+H]+, 197 [C6H10O5+NH4+NH3]+, 180 [C6H10O5+NH4]+; UV lmax (MeOH) nm: 225, 260, 301, 333, 345; IR max (KBr) cmÿ1: 3400 (OH), 1632 (CÿC), 1028 (CÿO); 1H (300 MHz; CD3OD) and 13C NMR (75.5 MHz; CD3OD): Table 1. Acknowledgements This . study was supported partly by grants from TUÈBITAK-BAYG (Scienti®c and Technical Research Council of Turkey), Turkey and Researching Project Programs of Hacettepe University (98T01102011), Turkey. References Baytop, T., 1984. Therapy with Medicinal Plants (Past and Present). Nobel Tip Kitabevleri, Istanbul. Davis, P.H., 1967. Flora of Turkey and the East Aegean Islands. University Press, Edinburgh. Davis, P.H., 1988. Flora of Turkey and the East Aegean Islands. University Press, Edinburgh. Demirezer, LOÈ, KuruuÈzuÈm, A., 1997. A comparative chemotaxonomic study on eleven Rumex species growing in Turkey. FABAD Journal of Pharmaceutical Sciences 22, 153±158. Nishina, A., Kubota, K., Kameoka, H., Osawa, T., 1991. Antioxidizing component, musizin, in Rumex japonicus Houtt. Journal of American Oil Chemical Society 68, 735±739. Sharma, M., Sharma, P., Rangaswami, S., 1977. Orientalone, a new 1,4-naphthoquinone from Rumex orientalis. Indian Journal of Chemistry 15B, 544±545. Suri, J.L., Dhar, K.L., Atal, C.K., 1978. Chemical constituents of Rumex orientalis Bernh. Journal of Indian Chemical Society 55, 292±293. Tsuboi, M., Minami, M., Nonaka, G.I., Nishioka, I., 1977. Studies on Rhubarb (Rhei Rhizoma) IV. Naphthalene glycosides. Chemical and Pharmaceutical Bulletin 25, 2708±2712.