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Journal of Natural Medicines

, Volume 73, Issue 1, pp 257–261 | Cite as

Two new xanthones and cytotoxicity from the bark of Garcinia schomburgkiana

  • Sutin KaennakamEmail author
  • Kamonwan Mudsing
  • Kitiya Rassamee
  • Pongpun Siripong
  • Santi Tip-pyangEmail author
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Abstract

Two new xanthone derivatives, named schomburgones A (1) and B (2), along with eight known compounds, including xanthones (38) and anthraquinones (910) were isolated from the bark of Garcinia schomburgkiana. Their structures were determined by spectroscopic analysis especially 1D and 2D NMR spectroscopies. All isolated compounds were evaluated for their cytotoxicity against five cancer cell lines (KB, HeLa S-3, HT-29, MCF-7 and HepG-2). Compounds 36 and 8 showed good cytotoxicity against all the five cancer cell lines with IC50 values in the range of 1.45–9.46 µM.

Keywords

Garcinia schomburgkiana Clusiaceae Xanthone Cytotoxicity 

Introduction

Garcinia schomburgkiana Pierre (family Clusiaceae) is a medium-sized tree distributed in Thailand, Laos, Vietnam, and Cambodia. In folk medicine in these countries, its leaves, roots, and fruits are used for the treatment of cough, menstrual disturbances, expectorant, laxative and diabetes [1]. Previous chemical and biological studies on the chemical constituents of G. schomburgkiana showed the presence of xanthones, depsidones, biphenyls, flavonoids, triterpenoids, and phloroglucinols, some of which exhibited antimalarial activity and cytotoxicity [2], [3]. Here, we reported two new xanthone derivatives, named schomburgones A (1) and B (2), along with six known xanthones (38) and two known anthraquinones (910) from the bark of this plant. The structures of all isolated compounds were elucidated using spectroscopic methods especially 1D and 2D NMR spectroscopies and compared with their 1H and 13C NMR spectroscopic data from the literature. The cytotoxicity of all isolated compounds was evaluated using the MTT method against five cancer cell lines.

Results and discussion

Phytochemical investigation of CH2Cl2 crude extract from the bark of G. schomburgkiana led to the isolation of two new xanthone derivatives, named schomburgones A (1) and B (2), along with eight known compounds (Fig. 1), including isocudraniaxanthone B (3) [4], gerontoxanthone I (4) [5], nigrolineaxanthone E (5) [6], isojacareubin (6) [7], dulxanthone A (7) [8], macluraxanthone (8) [9], vismiaquinone A (9) [10], and 3-geranylemodin (10) [11]. The structures of all isolated compounds were elucidated using spectroscopic methods especially NMR spectroscopies and compared with their 1H and 13C NMR spectroscopic data from the literature.
Fig. 1

Chemical structures of 110

Schomburgone A (1) was obtained as yellow oil. Its molecular formula was determined as C24H24O6 by the negative HRESIMS measurement through the ion peak at m/z 407.1527 [M–H]+ (calcd. for C24H23O6, 407.1495). The UV spectrum displayed absorption bands at λmax 395, 315 and 243 nm, which is typical of the xanthone chromophore [12]. The IR spectrum showed phenolic hydroxyl groups and a hydrogen bonded carbonyl group at 3422 and 1632 cm−1. The 1H NMR spectrum showed the presence of a 3,3-dimethylallyl substituent, which was confirmed by two singlets at δH 1.67 (3H, s, H-4) and 1.85 (3H, s, H-5) for the vinyl methyls, a triplet at δH 5.20 (1H, t, J = 7.23 Hz, H-2) for the vinylic proton and a doublet at δH 3.44 (2H, d, J = 7.23 Hz, H-1) for the allylic proton of prenyl group. In addition, the methoxy signal, a hydroxyl signal, two aromatic proton signals and a hydrogen bonded hydroxyl signal appeared as five singlets at δH 3.89 (3H, s, OCH3-3), 6.26 (1H, s, OH-6), 6.33 (1H, s, H-2), 6.85 (1H, s, H-5) and 13.38 (1H, s, OH-1), respectively. The signals at δH 1.50 (6H, s, H-4 and H-5), 5.83 (1H, d, J = 10.23 Hz, H-2) and 8.02 (1H, d, J = 10.23 Hz, H-1) in the spectrum were indicative of a dimethylchromene ring. The angular fusion of the chromene ring at C-8 was deduced from the low field shift of H-1 (δH 8.02), which was located in the deshielding area of the carbonyl group. The 1H and 13C NMR spectroscopic data (Table 1) were shown to be similar to those of the known xanthone, paxanthone B [13], except that the hydroxyl group at C-3 of paxanthone B was replaced by a methoxy group. In the HMBC correlations of 1 (Fig. 2), the methoxy proton at δH 3.89 showed a cross-peak with δC 163.5 (C-3). In addition, a methine proton at δH 8.02 showed cross-peaks with δC 77.1 (C-3), 108.5 (C-8a), and 136.9 (C-7), confirming that a dimethylchromene ring was located at C-8, and a methylene proton at δH 3.44 showed cross-peaks with δC 131.7 (C-3), 153.7 (C-4a), and 163.5 (C-3), indicating that a prenyl group was attached to C-4. Thus, the complete assignment of schomburgone A was determined as 1.
Table 1

NMR spectroscopic data (400 MHz, CDCl3) for 1 and 2

Position

1

2

δH (J in Hz)

δ C

δH (J in Hz)

δ C

1

 

162.0

 

163.0

2

6.33 (s)

94.1

6.43 (s)

96.2

3

 

163.5

 

166.1

4

 

107.2

 

113.9

4a

 

153.7

 

154.0

5

6.85 (s)

102.6

 

145.9

6

 

151.1

7.26 (d, 7.62)

120.3

7

 

136.9

7.23 (t, 7.62)

124.6

8

 

119.9

7.71 (d, 7.62)

116.6

8a

 

108.5

 

121.0

9

 

183.0

 

182.0

9a

 

103.9

 

104.1

10a

 

153.5

 

144.8

1

3.44 (d, 7.23)

21.7

 

42.0

2

5.20 (t, 7.23)

122.4

6.70 (dd, 10.63,17.67)

156.3

3

 

131.7

5.22 (d, 17.67), 5.07 (d, 10.63)

104.5

4

1.67 (s)

25.9

1.61 (s)

28.4

5

1.85 (s)

18.0

1.61 (s)

28.4

1

8.02 (d, 10.23)

121.2

  

2

5.83 (d, 10.23)

132.5

  

3

 

77.1

  

4

1.50 (s)

27.5

  

5

1.50 (s)

27.5

  

1-OH

13.38 (s)

 

13.25 (s)

 

5-OH

  

6.42 (s)

 

6-OH

6.26 (s)

   

3-OCH3

3.89 (s)

56.1

3.91 (s)

56.2

Fig. 2

Key HMBC (arrow curves) and COSY (bold lines) correlations of 1 and 2

Schomburgone B (2) was obtained as yellow oil. Its molecular formula was determined as C19H18O5 by the negative HRESIMS measurement through the ion peak at m/z 325.1098 [M–H]+ (calcd. for C19H17O5, 325.1076). The UV spectrum displayed absorption bands at λmax 394, 315 and 244 nm. The IR spectrum showed phenolic hydroxyl groups and a hydrogen bonded carbonyl group at 3432 and 1642 cm−1. The 1H NMR spectrum showed the presence of a 1,1-dimethylallyl group, which was confirmed by a singlets at δH 1.61 (6H, s, H-4 and H-5) for two methyls, a doublet of doublet at δH 6.70 (1H, dd, J = 10.63 Hz, 17.67, H-2) for the methine proton and two doublets at δH 5.07 (1H, d, J = 10.63 Hz, H-3) and 5.22 (1H, d, J = 17.67 Hz, H-3) for the methylene protons. Moreover, the methoxy signal, hydroxyl signal, aromatic proton signal and hydrogen bonded hydroxyl signal appeared as four singlets at δH 3.91 (3H, s, OCH3-3), 6.42 (1H, s, OH-5), 6.43 (1H, s, H-2) and 13.25 (1H, s, OH-1), respectively. The ABC-type aromatic protons were assigned at δH 7.23 (1H, t, J = 7.62 Hz, H-7), 7.26 (1H, d, J = 7.62 Hz, H-6) and 7.71 (1H, d, J = 7.62 Hz, H-8). The 1H and 13C NMR spectroscopic data (Table 1) were shown to be similar to those of the known xanthone, pancixanthone A [14], except that the hydroxyl group at C-3 of pancixanthone-A was substituted by a methoxy group. In the HMBC correlations of 2 (Fig. 2), the methoxy proton at δH 3.91 showed a cross-peak with δC 166.1 (C-3). Moreover, two methyl protons at δH 1.61 showed cross-peaks with δC 113.9 (C-4) and 156.3 (C-2), confirming that a 1,1-dimethylallyl group was connected at C-4. Thus, the completed assignment of schomburgone B was determined as 2.

In previous research many xanthones showed cytotoxicity [15]. Therefore, all isolated compounds were evaluated in vitro for their cytotoxicity against five cancer cell lines (KB, HeLa S-3, HT-29, MCF-7 and HepG-2) (Table 2). Compounds 36 and 8 showed good cytotoxicity against all five cancer cell lines with IC50 values in the range of 1.45–9.46 µM. Compounds 1 and 7 showed weak cytotoxicity against all five cancer cell lines with IC50 values in the range of 34.69–73.10 µM. Compounds 2, 9 and 10 showed inactive cytotoxicity against all five cancer cell lines with IC50 values >100 µM. The SAR studied data (Fig. 1; Table 2) of xanthones suggest that the ortho hydroxy group at C-5 and C-6 and the 1,1-dimethylallyl group at C-4 might improve the cytotoxicity as inferred from the comparison of their cytotoxicity of compounds 110.
Table 2

In vitro cytotoxicity of compounds 110 against five cancer cell lines

Compounds

IC50 (μM) ± SD

KB

HeLa S-3

HT-29

MCF-7

HepG-2

1

45.05 ± 2.08

69.22 ± 4.02

61.92 ± 2.40

52.21 ± 1.71

73.19 ± 1.14

2

> 100

> 100

> 100

> 100

> 100

3

5.23 ± 0.19

7.95 ± 0.25

7.87 ± 0.30

6.70 ± 0.81

5.93 ± 0.94

4

4.69 ± 0.21

7.57 ± 0.26

9.18 ± 0.38

5.26 ± 0.55

4.89 ± 0.83

5

5.08 ± 0.36

5.82 ± 0.15

4.17 ± 0.07

7.19 ± 0.36

9.46 ± 0.45

6

4.30 ± 0.12

6.60 ± 0.24

5.92 ± 0.40

3.21 ± 0.71

3.19 ± 0.14

7

38.17 ± 6.83

65.26 ± 3.89

34.69 ± 2.29

46.03 ± 1.29

54.80 ± 1.18

8

1.45 ± 0.09

1.62 ± 0.20

1.87 ± 0.30

1.70 ± 0.81

1.93 ± 0.94

9

> 100

> 100

> 100

> 100

> 100

10

> 100

> 100

> 100

> 100

> 100

Doxorubicin

0.13 ± 0.006

0.03 ± 0.001

0.31 ± 0.07

0.42 ± 0.14

1.23 ± 0.02

IC50 ≤ 10 = good activity, 10 < IC50 ≤ 30 = moderate activity, IC50 > 100 = inactive

Experimental

General experimental procedures

NMR spectra were recorded on Bruker 400 AVANCE spectrometer. HRESIMS spectra were obtained using a Bruker MICROTOF model mass spectrometer. The UV–visible absorption spectra were recorded on a UV-2550 UV–Vis spectrometer (Shimadzu, Kyoto, Japan). The IR spectra were measured on a Nicolet 6700 FT-IR spectrometer using KBr discs.

Plant material

The bark of G. schomburgkiana was collected from Bang Ramat Road, Khwaeng Bang Ramat, Khet Taling Chan, Bangkok Thailand (13°4542N, 100°2456E), in June 2017. The plant material was identified by Dr. Suttira Sedlak, a botanist at the Walai Rukhavej Botanical Research Institute, Mahasarakham University, and a specimen retained as a reference (Khumkratok no. 92-08).

Extraction and isolation

The air-dried bark of G. schomburgkiana (2.0 kg) was extracted with CH2Cl2 at room temperature for 7 days (2 × 25 L). The CH2Cl2 crude extract (91.0 g) was further separated by column chromatography (CC) over silica gel CC and eluted with a gradient of Hexane–EtOAc (90, 70, 50 and 30% Hexane–EtOAc each 5 L) to give six fractions (A-F). Fraction A (4.0 g) was purified by Sephadex LH-20 column eluted with 80% CH2Cl2–MeOH (2 L) and further applied to a radial chromatography (chromatotron) with 95% hexane–EtOAc (200 mL) to afford compound 9 (3.2 mg). Fraction B (10.5 g) was purified by Sephadex LH-20 column eluted with 80% CH2Cl2–MeOH (2 L) and further applied to a chromatotron with 50% hexane–CH2Cl2 (200 mL) to obtain compounds 2 (4.2 mg), 4 (2.5 mg) and 7 (2.3 mg). Compound 1 (7.2 mg) was separated by Sephadex LH-20 column eluted with 50% CH2Cl2–MeOH (2 L) from fraction C (2.0 g). Fraction D (6.5 g) was purified by Sephadex LH-20 column eluted with 50% CH2Cl2–MeOH (2 L) to give compounds 5 (8.5 mg) and 8 (4.6 mg). Fraction E (8.5 g) was purified by Sephadex LH-20 column eluted with 50% CH2Cl2–MeOH (2 L) and further applied to a chromatotron with 70% hexane–EtOAc (200 mL) to obtain compounds 3 (5.2 mg) and 10 (5.5 mg). Finally, fraction F (1.2 g) was subjected to silica gel CC eluted with 100% CH2Cl2 and further purified by Sephadex LH-20 column eluted with 80% CH2Cl2–MeOH (2 L) to yield compound 6 (6.5 mg).

Schomburgone A (1): yellow oil; UV (CHCl3) λmax (log ε): 395 (3.6), 315 (4.2) and 243 (4.4) nm,. IR νmax (KBr): 3422 and 1632 cm−1; 1H (400 MHz, CDCl3) and 13C NMR (100 MHz, CDCl3) spectroscopic data, see Table 1; HRESIMS m/z 407.1527 [M–H]+ (calcd. for C24H23O6, 407.1495).

Schomburgone B (2): yellow oil; UV (CHCl3) λmax (log ε): 394 (3.5), 315 (4.0) and 244 (4.2) nm,. IR νmax (KBr): 3432 and 1642 cm−1; 1H (400 MHz, CDCl3) and 13C NMR (100 MHz, CDCl3) spectroscopic data, see Table 1; HRESIMS m/z 325.1098 [M–H]+ (calcd. for C19H17O5, 325.1076).

Cytotoxicity assay

All isolated compounds (110) were subjected to cytotoxic evaluation against KB (human epidermoid carcinoma), HeLa S-3 (human cervical carcinoma), HT-29 (human colon adenocarcinoma), MCF-7 (human breast adenocarcinoma) and HepG-2 (human liver carcinoma) cell lines employing the colorimetric method [16]. Doxorubicin was used as the reference substance which exhibits activity against five cancer cell lines. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (Sigma Chemical Co., USA) was dissolved in saline to make a 5 mg/mL stock solution. Cancer cells (3 × 103 cells) suspended in 100 μg/wells of MEM medium containing 10% fetal calf serum (Gibco BRL, Life Technologies, NY, USA) were seeded onto a 96-well culture plate (Costar, Corning Incorporated, NY, USA). After 24 h pre-incubation at 37 °C in a humidified atmosphere of 5% CO2/95% air to allow cellular attachment, various concentrations of test solution (0.1, 0.3, 1.0, 3.0, 10.0, 30.0, and 100.0 μM, each 10 μL/well) were added and these were then incubated for 48 h under the above conditions. At the end of the incubation, 10 μL of tetrazolium reagent was added into each well followed by further incubation at 37 °C for 4 h. The supernatant was decanted, and DMSO (100 μL/well) was added to allow formosan solubilization. The optical density of each well was detected using a Microplate reader at 550 nm and for correction at 595 nm. Each determination represented the average mean of six replicates. The 50% inhibition concentration (IC50 value) was determined by curve fitting.

Notes

Acknowledgments

The authors are grateful to the Graduate School of Chulalongkorn University for a Postdoctoral Fellowship (Ratchadaphiseksomphot Endowment Fund) to SK. We also thank Dr. Suttira Sedlak, Walai Rukhavej Botanical Research Institute, Mahasarakham University, Mahasarakham 44000, Thailand for identification and deposition of the plant material.

Supplementary material

11418_2018_1240_MOESM1_ESM.pdf (972 kb)
Supplementary material 1 (PDF 972 kb)

References

  1. 1.
    Mungmee C, Sitthigool S, Suttisri R, Buakeaw A (2012) Xanthones and biphenyls from Garcinia schomburgkiana wood and their cytotoxicity. Thai J Pharm Sci 36:6–9Google Scholar
  2. 2.
    Le DH, Nishimura K, Takenaka Y, Mizushina Y, Tanahashi T (2016) Polyprenylated Benzoylphloroglucinols with DNA polymerase inhibitory activity from the fruits of Garcinia schomburgkiana. J Nat Prod 79:1798–1807CrossRefPubMedGoogle Scholar
  3. 3.
    Sukandar ER, Siripong P, Khumkratok S, Tip-Pyang S (2016) New depsidones and xanthone from the roots of Garcinia schomburgkiana. Fitoterapia 111:73–77CrossRefPubMedGoogle Scholar
  4. 4.
    Kobayashi M, Mahmud T, Yoshioka N, Shibuya H, Kitagawa I (1997) Indonesian medicinal plants. XXI. Inhibitors of Na+/H+ exchanger from the bark of Erythrina variegata and the roots of Maclura cochinchinensis. Chem Pharm Bull 45:1615–1619CrossRefPubMedGoogle Scholar
  5. 5.
    Chang CH, Lin CC, Kawata Y, Hattori M, Namba T (1989) Prenylated xanthones from Cudrania cochinchinensis. Phytochemistry 28:2823–2826CrossRefGoogle Scholar
  6. 6.
    Rukachaisirikul V, Ritthiwigrom T, Pinsa A, Sawangchote P, Taylor WC (2003) Xanthones from the stem bark of Garcinia nigrolineata. Phytochemistry 64:1149–1156CrossRefPubMedGoogle Scholar
  7. 7.
    Helboe P, Arends P (1973) Xanthone studies. VI. Synthesis of jacareubin, isojacareubin, and some hydroxyxanthones with allylic substituents. Arch Pharm Chemi Sci Ed 1:549–555Google Scholar
  8. 8.
    Ito C, Miyamoto Y, Nakayama M, Kawai Y, Rao KS, Furukawa H (1997) A novel depsidone and some new xanthones from Garcinia species. Chem Pharm Bull 45:1403–1413CrossRefGoogle Scholar
  9. 9.
    Wolfrom ML, Komitsky FJ, Fraenkel G, Looker JH, Dickey EE, McWain P (1963) Macluraxanthone and two accompanying pigments from the root bark of the osage orange. Tetrahedron Lett 4:749–755CrossRefGoogle Scholar
  10. 10.
    Delle Monache F, Ferrari F, Marini-Bettolo GB, Maxfield P, Cerrini S, Fedeli W (1979) Vismiones from Vismia baccifera var. dealdata (H. B. K.): chemistry and X-ray structure determination. Gazz Chim Ital 109:301–310Google Scholar
  11. 11.
    Botta B, Delle Monache F, Delle Monache G, Marini Bettolo GB, Oguakwa JU (1983) 3-Geranyloxy-6-methyl-1,8-dihydroxyanthraquinone and vismiones C, D, and E, from Psorospermum febrifugum. Phytochemistry 22:539–542CrossRefGoogle Scholar
  12. 12.
    Kaennakam S, Siripong P, Tip-Pyang S (2015) Kaennacowanols A-C, three new xanthones and their cytotoxicity from the roots of Garcinia cowa. Fitoterapia 102:171–176CrossRefPubMedGoogle Scholar
  13. 13.
    Ishiguro K, Nakajima M, Fukumoto H, Isoi K (1995) Co-occurrence of prenylated xanthones and their cyclization products in cell suspension cultures of Hypericum patulum. Phytochemistry 38:867–869CrossRefGoogle Scholar
  14. 14.
    Ito C, Miyamoto Y, Rao KS, Furukawa H (1996) A novel dibenzofuran and two new xanthones from Calophyllum panciflorum. Chem Pharm Bull 44:441–443CrossRefGoogle Scholar
  15. 15.
    Vo HT, Nguyen NTT, Nguyen HT, Do KQ, Connolly JD, Maas G (2012) Cytotoxic tetraoxygenated xanthones from the bark of Garcinia schomburgkiana. Phytochem Lett 5:553–557CrossRefGoogle Scholar
  16. 16.
    Kongkathip N, Kongkathip B, Siripong P, Sangma C, Luangkamin S, Niyomdecha M (2003) Potent antitumor activity of synthetic 1,2-naphthoquinones and 1,4-naphthoquinones. Bioorganic Med Chem 11:3179–3191CrossRefGoogle Scholar

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Authors and Affiliations

  1. 1.Center of Excellence in Natural Products Chemistry, Department of Chemistry, Faculty of ScienceChulalongkorn UniversityBangkokThailand
  2. 2.Natural Products Research Section, Research DivisionNational Cancer InstituteBangkokThailand

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