Oligostilbenes from Gnetum Species and Anticarcinogenic and

Chapter 4

Oligostilbenes from Gnetum Species and Anticarcinogenic and Antiinflammatory Activities of Oligostilbenes 1

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Downloaded by UNIV OF ARIZONA on June 29, 2014 | http://pubs.acs.org Publication Date: September 1, 2008 | doi: 10.1021/bk-2008-0987.ch004

Ka-Wing Cheng , Mingfu Wang , Feng Chen , and Chi-Tang Ho

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1

Department of Botany, The University of Hong Kong, Pokfulam Road, Hong Kong, People's Republic of China Department of Food Science, Rutgers, The State University of New Jersey, 65 Dudley Road, New Brunswick, NJ 08901

2

Interest in the physiological roles of phytochemicals has increased dramatically over the last twenty years. Oligostilbenes, which are widely distributed in several plant families and liverworts, have attracted much attention due to emerging evidence suggesting them to be potential disease preventive and curing agents against several chronic diseases, particularly through their anti-inflammatory, antioxidant and anti-carcinogenic activities. The genus Gnetum is of particular interest in search of this group of polyphenols owing to its large genus size and wide distribution of its species in both hemispheres. In this paper we summarize the oligostilbenes that have been isolated from Gnetum species and we also discuss the anti-carcinogenic and anti-inflammatory activities of oligostilbenes in general.

36

© 2008 American Chemical Society

In Dietary Supplements; Ho, C., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 2008.


37 Stilbenes are a group of naturally occurring phenolic compounds characterized by the presence of diphenylethylene backbone and have been isolated mainly from angiosperms and gymnosperms (7). In plants, stilbenes are synthesized through specific stilbene synthase with p-coumaroyl-CoA and malonyl-CoA as major precursor molecules (2). During the course of identifying the paramount active compounds in prevention and treatment of the age-related diseases, stilbenes have emerged to hold important positions stemming from various in vitro and in vivo studies which strongly support their potential benefits in a wide range of health disorders through their antioxidant, anti-inflammatory and anticarcinogenic activities, among others (5-7). The most remarkable monomeric stilbene compound discovered so far is resveratrol (3,5,4trihydroxystilbene), which exhibits a wide range of pharmacological effects, including cardiovascular protection, antioxidation and cancer chemoprevention (8-16). Stilbenes, which exist in oligomeric forms, are known as oligostilbenes. They are believed to arise from oxidative condensation of stilbene nucleus, and are known to be present mainly in nine plant families, Dipterocarpaceae, Vitaceae, Cyperaceae, Gnetaceae, Leguminosae, Celastraceae, Paeoniaceae, Moraceae and Iridaceae, in woody tissues, as constitutive components, though various other tissues have also been reported to contain such diphenyl ethenebridged compounds (77-27). Oligostilbenes have also been shown to possess strong health-promoting effects (22-27) and a number of them have demonstrated more potent anticarcinogenic and anti-inflammatory activities than the corresponding monomers. Most oligostilbenes are not regular dietary components in human populations (28), while plants from Gnetum are an exception. Gnetum is a perennial plant belonging to the Gymnosperms (29). It embodies over 40 species that can be monoecious or dioecious (30). Most of them are trees or shrubs and some are woody climbers (29). They are widely distributed in both hemispheres, particularly inhabiting the tropics of Asia, Africa, South America and certain islands between Asia and Australia (29,31). Some species of this perennial plant have a long history in the diets of some populations in under-developed countries in central Africa, where they are a significant source of dietary protein and mineral elements (29,32), though other biological roles have not been evaluated in these populations. In addition, various species of the genus have long been used in medical prescriptions as a blood invigorating agent, for treatment of chronic diseases like arthritis, bronchitis and asthma and have been shown to contain oligostilbenes (33-37). In recent years, the genus Gnetum has attracted considerable attention due to emerging evidence suggesting them to be potential rich sources of oligostilbenes. Myriad oligostilbenes have been isolated and structurally characterized from Gnetum species, ranging from homo-, hetero-dimers, to trimers and tetramers, along with their derivatives, but more extensive in vitro and in vivo studies are demanded to fill the current gap in our understanding of their pharmacological


38 potentials. This review will focus on the chemistry of oligostilbenes from Gnetum and the anti-inflammatory and anticarcinogenic activities of oligostilbenes in general.

Chemistry of Gnetum Oligostilbenes


Structural Features and Classification Since the isolation of gnetin A from G. leyboldii in 1982 (36), more than 80 oligostilbenes have been obtained from this genus, mainly from their liana and root, but also from wood, bark and fruit. Among the Gnetum species, G. hainanense and G. gnemon have been most extensively evaluated for their oligostilbene content. In Table 1, selected Gnetum species, their corresponding oligostilbenes together with order of polymerization, molecular formula and constitutional monomers were presented in order of their isolation from the species. These species exhibit wide variation in their oligostilbene constituents on the basis of the types of oligostilbenes that have been isolated and characterized. This thus confirms the need to systematically screen and identify oligostilbenes from this genus. It appears that geographical variation also significantly affects the stilbene contents. A s an example, several studies have reported the isolation of stilbene dimmers from G. montanum, Xiang et al. obtained no dimeric stilbenes from the same species of a different geographical origin (38). These geographical factors complicate species-based profding of oligostilbene contents. Similar to oligostilbenes isolated from other plant genus, most of the oligostilbenes of Gnetum origin contain resveratrol as the fundamental unit. The simplest are those derived from direct oxidative condensation of resveratrol, such as gnetins and (-)-e-viniferin. More complicated structures result from homo- or heteromeric condensation of resveratrol and its derivatives like oxyresveratrol, piceatannol, isorhapontigenin, gnetol and resveratrolosides (Figure 1). These mainly include dimmers, trimers and tetramers, which are substituted by varying number and pattern of hydroxyl groups. The number of hydroxy! group can be confirmed by the results of acetylation and methylation experiments (39). Acetylation is generally performed in a mixture of pyridine and acetic anhydride for 24 hours under room temperature, while methylation usually carried out with M e l and K C 0 in dry acetone under reflux for 3 hours (40). Different connectivity between monomers also gives rise to different oligostilbenes. Based on the structural characteristics of their connection, these oligomers can be divided into five groups (A-E). Group A contains at least one oxygen heterocyclic ring, which may be five-membered or six-membered. The former, usually in the form of /rara-2-aryl-2,3-dihydrobenzofuran moiety, 2

3


39


Table I. Oligostilbenes from Selected Gnetum Species Part Compound Oligomer type Constitutional examined isolated (Mot formula) monomers Gnetum wood gnetin A dimer Res leyboldii (C H 0 ) wood gnetin B dimer Res (C H 0 ) wood gnetin C dimer Res (C H 0 ) wood gnetin D dimer Res & oxyRes (C H 0 ) wood gnetin E trimer Res (C4 H 09) ? Gnetum fruits gnetin C dimer schwa(C H 0 ) ckeanum fruits ? gnetin E trimer (C H 09) ? Gnetum lianas gnetifolin C dimer parvi(C H O ) ? folium lianas gnetifolin D dimer (C H O ) lianas e-viniferin dimer Res 28

22

6

28

24

6

28

22

6

28

22

7

2

bark bark bark bark

Gnetum fruit venosum kernels fruit kernels fruit kernels fruit kernels Gnetum ula

parvifolol A parvifolol B parvifolol C parvifolol D 2b-hydroxyampelopsin F gnetin C gnetin E gnetin J gnetin K gnetulin

22

36 36 36

32

30

26

8

30

28

8

dimer (C H dimer (C H dimer (C H dimer (C H dimer (C H 28

22

0)

28

22

0)

28

22

0)

30

26

28

22

Res & oxyRes

40

Res & oxyRes

40

oxyRes

40

Isorhapontigenin

40

Res & oxyRes

40

Res

37

Res

37

Res & piceatannol

37

Res & isorhapon tigenin Isorhapontigenin

37

8

0) 7

42

32

9

26

48

O)

6

34

48

8

22

43

48

7

28

32

36

7

dimer (C H 0 ) trimer (C H 0 ) trimer (C H Oio) trimer (C H Oio) dimer (C H O ) 42

36

6

42

30

36

32

28

bark

36

94

8

Continued on next page.


40 Table I. Continued. Source Gnetum montanum

Part Compound Oligomer type Constitutional Ref examined isolated (Molformula) monomers lianas gnetifolin M dimer isorhapontigenin 46 (C30H28O9)

lianas

gnetifolin N

dimer

isorhapontigenin

46

isorhapontigenin

47

isorhapontigenin

47

Res & oxyRes

95

Res & oxyRes

95

Res & oxyRes

95

Res & oxyRes

95

Res & oxyRes

95

isorhapontigenin

35

isorhapontigenin

35

isorhapontigenin

35

isorhapontigenin

35

isorhapontigenin & oxyRes Isorhapontigenin & gnetol

35

Isorhapontigen-in & oxyRes Res & oxyRes

96 34

Isorhapontigenin

34

Isorhaponti-genin

34

Isorhapontigenin

34

(C30H28O9)

lianas

gnetifolin L

lianas

gnetifolin 0

dimer (C3 H 8O ) dimer 0

2

8


(C30H28O9)

Gnetum Hainanense

lianas lianas lianas lianas lianas lianas lianas lianas lianas lianas lianas

lianas lianas lianas lianas lianas

gnetuhainin A gnetuhainin B gnetuhainin C gnetuhainin D gnetuhainin E gnetuhainin F gnetuhainin G gnetuhainin H gnetuhainin I gnetuhainin J gnetuhainin K gnetuhainin L gnetuhainin M gnetuhainin N gnetuhainin 0 gnetuhainin P

dimer (C28H 0 ) dimer 22

7

(C28H20O7)

dimer (C 8H 20 ) dimer (C H240 ) dimer (C H 408) dimer 2

2

7

28

8

28

2

(C30H24O8)

dimer (C30H22O9)

dimer (C30H24O9

dimer (C3oH 0 ) dimer 28

9

(C29H24O8)

dimer (C H240 ) 29

8

dimer (C29H24O8)

trimer (C4 H 20 ) trimer 2

3

96

n

(Q5H38O12)

trimer (C45H38O12)

dimer (C3oH2 0 ) 8

9


41 Table I. Continued. Part Compound examined isolated Gnetum lianas gnetuhainin hainanQ ense lianas gnetuhainin (continued) R lianas gnetuhainin S Gnetum stem Gnemonoside gnemonA oides stem Gnemonoside B stem Gnemonoside C stem Gnemonoside D stem Gnemonoside lianas E stem Gnemonoside lianas F stem Gnemonoside lianas G stem Gnemonoside lianas H stem Gnemonoside lianas I stem Gnemonoside lianas J stem gnemonol C lianas stem gnetin E lianas stem 2b-hydroxylianas ampelopsin F Gnetum lianas gnetupendin pendulum C lianas gnetupendin D lianas gnetin D


Source

Oligomer type Constitutional Ref (Mol formula) monomers dimer Res & isorha 96 pontigenin (C29H24O7) tetramer Isorhapontigenin 51 (Q0H51O16)

dimer

Res & oxyRes

51

Resveratroloside

97

Resveratroloside

97

Res & resvera troloside Res & resvera troloside Resveratroloside

97

Resveratroloside

44

Res & resvera troloside Res & Resdiglucopyranoside Resveratroloside & oxyRes NA

44

44

Res & oxyRes

39

Res

39, 36 39, 40

(C28H23O7)

dimer (C40H42O16)

dimer (C40H42O16)

dimer (C 4H 20„) dimer 3

3

(C34H32O1,)

dimer

97 39

(C40H42O16)

trimer (C60H62O24)

trimer (C54H52O19)

trimer (C46H52O21)

dimer (C40H42O17)

dimer

44 44

(C40H42O17)

Tetramer (C H4 0i ) trimer 56

2

3

(C42H32O9)

dimer

Res & oxyRes

(C28H22O7)

dimer

Res & oxyRes

50

isorhapontigenin

92

Res

92

(C28H22O7)

dimer C H 0i3) dimer 36

36

(C28H22O7



42 Table I. Continued Part Compound Oligomer type Constitutional examined isolated (Molformula) monomers shegansu B dimer lianas isorhapontigenin (C H 0 ) Gnetum stem oxyRes & isorha gneafricanins dimer africanum lianas pontigenin A (C29H24O8) stem gneafricanins dimer isorhapontigenin lianas B & piceatannol (C29H24O8)


36

Gnetum gnemon

stem lianas stem lianas stem lianas stem lianas root

gneafricanins C gneafricanins D gneafricanins E gneafricanins F gnemonol A

26

dimer

gnemonol B

92, 98 99 99

piceatannol

100

Res

100

Res & oxyRes

100

isorhapontigenin

100

Res & oxyRes

39

Res

39

Res

30

Res

30

Res & oxyRes

30

Res & oxyRes

30

Res

30

Res

30

Res

30

Res

30

Res

30, 101

Res & oxyRes

102

(C28H22O8)

dimer (C H 0 ) dimer 28

24

6

(C28H24O7)

dimer (C H26O ) trimer (C H 0i6) tetramer 30

8

42

root

8

Ref

MXCJ

32

(C56H42O12)

root

gnemonol G

dimer (C28H20O7)

root

gnemonol H

trimer (C 2H Oio) trimer (C 2H 0i6) trimer (C 2H 0i6) dimer 4

root

gnemonol I

root

gnemonol J

root

gnetin C

32

4

32

4

32

(C28H22O6)

root

gnetin D

dimer (C28H22O7)

root

gnetin E

trimer (C 2H 09) trimer (C4 H 0 ) trimer (C4 H 09) 4

root root

root

ampelopsin E cisampelopsin p gnemonol D

32

2

2

32

9

32

trimer (C H Oio) 42

32


43 Table I. Continued. Sourrp

U l / M l L/C-

Part Compound Oligomer type Constitutional Rpf examined isolated (Molformula) monomers nej root gnemonol E trimer 102 Res & oxyRes

Gnetum gnemon (continued) root

(C42H32O10)

gnemonol F

trimer

Res

102

Res

49

Res

49

isorhapontigenin

49

Res

49

resveratroloside

Res

49, 97 49, 97 97, 49 49, 43 49

Res

43

Res

isorhapontigenin

43, 36 43, 36 43, 36 43, 18 42

isorhapontigenin

42

isorhapontigenin

96

(C42H32O10)

root

gnemonol K

trimer (C42H32O9)

root

gnemonol L

trimer (C42H32O9)


root

gnemonol M

dimer (C30H26O&O

root root root root root

Gnemonoside K gnemonoside A gnemonoside B gnemonoside F latifolol

trimer (C oH 024) dimer 6

62

(C40H42O16)

dimer

resveratroloside

(C40H42O16)

trimer

resveratroloside

(C60H62O24)

trimer

Res & oxyRes

(C42H32O10)

Root

(-)-e-viniferin

dimer (C 8H 20 ) trimer 2

Gnetum stem latifolium stem

latifolol

2

(C42H32O10)

gnetin C

dimer (C 8H 20 ) dimer 2

stem

6

gnetin D

2

6

Res

(C28H22O7)

stem

gnetin E

trimer

Res

(C42H32O9)

stem

(-)-e-viniferin

dimer

Res

(C28H22O6)

Gnetum lianas cleistostachyum lianas lianas

bisisorhapontigenin A c/s-shegansu B gnetuhainin P

dimer (C30H26O8)

dimer (C3 H 6O ) dimer 0

2

8

(C30H28O9)



44 Table I. Continued. Source

Gnetum klossii

Part Compound Oligomer type Constitutional Ref monomers examined isolated (Mol formula) 94 isorhapontigenin lianas gnetulin dimer (C oH260 ) 103, stem Res (-)-E-viniferin dimer 18 (C28H22O6) isorhapontigenin 103, stem dimer gnetulin 94 (C oH260 ) 103 Res stem gnetin C dimer (C H 20 ) 103 stem gnetin E trimer Res (C H 0 ) 103, Res & oxyRes stem trimer latifolol 43 (C 2H Oio) 3

8

3

8


28

42

4

2

32

6

9

32

Res = resveratrol

predominates among oligostilbenes isolated from Gnetum (41). Group B contains both oxygen heterocyclic ring and homocyclic ring. Group C contains only homocyclic structure in its connective part. Group D has a carbonyl group in its cyclic connective part (bicyclo-octanoid) and its presence can be easily revealed in the IR spectrum with characteristic absorption band at around 1700 cm' (36). Group E does not possess any cyclic structure in connecting the monomeric units. Moreover, some oligostilbenes exist as isomers. Whilst most Gnetum oligostilbenes have /raw-configuration, a few cw-oligostilbenes have also been identified (1,17,30,42). These structural differences may affect their bioavailability, affinity for receptor, and thus their biological activity. There has not been a clear pattern in the distribution of these oligostilbenes among Gnetum species except for group B which has mainly been isolated from G. parvifolium and G. gnemon; and group D which has so far been identified only in G. leyboldii. 1

Extraction, isolation and identification Oligostilbenes are usually soluble in moderately polar to polar solvents such as ethyl acetate, ethanol and methanol. A variety o f solvents have been used in extraction of oligostilbenes from Gnetum species. Ethanol (60-95%) is commonly employed for obtaining the crude extract. The typical extraction method has been refluxing (35,42) or percolation with aqueous ethanol (37). Alternatively, successive extraction with acetone, MeOH and M e O H - H 0 mixture has also been adopted to obtain crude extract for further separation (30, 43). The first separation is usually carried out on silica gel column and eluted 2



45

OH Isorhapontigenin / 3methoxyresveratrol

Gnetol

Resveratroloside

Figure 1. Monomeric units constituting oligostilbenes from Gnetum spp.

with solvent system of increased polarity, in particular, by gradually increasing proportion of M e O H in C H C l - M e O H mixture. A preliminary partition step may also be conducted prior to the above column chromatography. Separation of individual oligostilbenes usually requires further chromatographic procedures, including repeating the column chromatographic process on silica gel, with similar or different elution solvent, such as cyclohexane-acetone mixture (55); column chromatography over Sephadex LH-20 often eluted with M e O H ; and over reversed phase silica gel. The final purification process is usually by preparative T L C and H P L C (40,43,44)- In some cases, especially for the more highly hydroxylated oligostilbenes, acetylation may be carried out prior to isolation to avoid oxidative modification or polymerization (45-47). The oligostilbenes obtained have a wide range of colors: brown (39), yellowish (48), 3


46 greenish (44), white (55) and colorless (49). Many of them show a positive reaction to Gibbs reagent. Structural elucidation is usually accomplished with a combination of spectroscopic techniques. U V spectra usually provide evidence for presence of conjugation or fiiran ring in the molecule, while IR spectra reveal presence of characteristic functional groups such as hydroxyl, aromatic, olefinic and carbonyl groups. Full elucidation of these oligostilbenes can be best accomplished with M S and various N M R techniques including H - H C O S Y , H B Q C , H M B C and N O E S Y . Fast atom bombardment ( F A B ) and electron ionization (EI) are currently the most common modes in molecular mass determination for oligostilbenes (35,50). Very few oligostilbenes obtainable from Gnetum have been determined for its absolute configuration. A previous circular dichroism study conducted by Kurihara and coworkers proposed a 7R, 8R absolute configuration for (-)-e-viniferin which was obtained from Carex pumila (18). There have also been rare studies, which used X-ray crystallography for analysis of the absolute configuration of some oligostilbenes (51).


!

!

Anticarcinogenic Activities of Oligostilbenes Studies, on the anticarcinogenic activity of oligostilbenes are limited, especially those from Gnetum species, yet among those tested, appreciable number of them have shown potent activities in different cancer cell lines. These studies mainly focused on their abilities to prohibit tumor progression rather than prevent its initiation. The plausible anticarcinogenic mechanisms have been ascribed as induction of apoptosis (52,72,106), cytotoxic and antiproliferative, antimutagenic activity (55-55), and inhibitory effects on the key enzymes that have been implicated in carcinogenesis.

Cytotoxicity Several studies have demonstrated the cytotoxic activities of oligostilbenes against different cancer cell lines (54,56,57,104,105). With the use of three human colon cancer cell lines (SW480, DLD-1 and COLO201), Ito and coworkers evaluated the in vitro cytotoxicity of two resveratrol oligomers, vaticanols B and C with resveratrol as the reference (56). A l l these three stilbenoids exhibited cytotoxicity, with vaticanol C being the most potent and the growth inhibition induced by vaticanol C was concentration-dependent. It was suggested that presence of a dibenzobicyclo[3.2.1]octadien framework may contribute to its higher cytotoxic activity (56). Seo et al. showed that vatdiospyroidol, a resveratrol tetramer, was cytotoxic against human oral


47 epidermoid carcinoma (KB), colon cancer, and breast cancer cell lines with E C values of 1.0 jig/mL, 1.9 ng/mL, and 3.8 jig/mL, respectively (57). Ohyama and coworkers elaborately tested the cytotoxic effect of 11 oligostilbenes against seven cancer cell lines including those of nasopharynx, lung, ileocecal, melanoma, renal, breast and ovary (54). Among these oligostilbenes, hopeaphenol, a resveratrol tetramer, showed most potent cytotoxicity against K B cells with an E D value of 1.2 ng/mL and moderate cytotoxicity against several other cancer cell lines. In contrast, the remaining oligomers (dimmers and trimers) did not exhibit significant cytotoxicity. These findings are in accordance with the hypothesis that oligostilbenes with more than four basic units are endowed with higher cytotoxicity (52). Although it was observed that resveratrol oligostilbenes constituting of more than four monomeric units are generally more potent cytotoxic agents, with very high molecular weight (from many monomeric units), it is probable that other factors like steric hindrance may also affect their bioactivities. Oligostilbenes also exhibited high selectivity against different cancer cells. Using five different cancer cell lines, K i m and coworkers found that fraws-E-viniferin, cw-e-viniferin and gnetin H possessed marked cytotoxicity against Hela (cervicse) and HCF-7 (breast) human cancer cell lines with I C values of 20.4, 21.5 and 12.9 fig/mL, respectively, but not effective against HepG2 (liver hepatoma) and HT-29 (colon) (53). Pronounced cytotoxicity of viniferin and gnetin H was also observed with H L 60 cells and in a dose dependent manner (58). The potential anticarcinogenic activities of oligostibenes are further exemplified by the fact that some oligostilbenes demonstrate good tumor specificity (relative sensitivity between the tumor cell lines and normal cells). In a comparative study engaging both stilbenes and flavonoids, the stilbene trimers demonstrated higher tumor-specific cytotoxicity, especially sophorastilbene A and (+)-a-viniferin, which yielded tumor-specific indices of 3.6 and 4.7, respectively (59). However, tumor specificity did not correlate with their ability to induce apoptosis as it was observed that (+)-a-viniferin exhibited the highest tumor specificity, yet induced internucleosomal D N A fragmentation and activation of caspases - 3 , -8 and - 9 in HL-60 cells at concentration much higher than other stilbenes tested (>200 \M) (59). Structural modification can significantly affect cytotoxic activity of oligostilbenes. As mentioned above, hopeaphenol was a strong cytotoxic agent against K B cells. However, its oxidative derivative stenophyllol A was ineffective in the same study and it was speculated that the 4-hydroxyphenyl moiety on the seven-membered rings of hopeaphenol may account for its enhanced cytotoxicity (54). Vatdiospyroidol (I) was potently cytotoxic against a panel of cancer cell lines, but conversion of the hydroxyl groups to methoxyl groups abrogated its activity, suggesting the importance of the hydroxyl groups in mediating cytotoxicity (57). Acetylation could be another strategy to alter cytotoxic activity. By adding an acetate group, Ohyama and coworkers significantly increased cytotoxicity of the dimmers, which displayed negligible activity in their native state (54). They suggested that 5 0


5 0

50


48 the presence of a lipophilic acetate group could aid in cellular penetration and thus access to the targeted point. However this enhancement seems to be restricted to dimmers as the trimers and tetramers remained inactive after acetylation. Information concerning the effect of structural modification is scarce, especially for oligomers of higher orders. More systematic structureactivity relationship (SAR) studies are demanded to characterize this group of phenolic compounds.


Anti-proliferation Horoshi et al demonstrated that some oligostilbenes possessed potent inhibitory activity against the isomerase at concentration of

Oligostilbenes from Gnetum Species and Anticarcinogenic and

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