Systematic significance of xanthones, benzophenones and biflavonoids in Garcinia

g/ochem/ce/~t/cs andS,

Print~ in GreatBritain.

Vol. 11, No. 1, pp. 21-28, 1883.

0Q05-19"/8/83/010021-08lI03.00/0

PergamonPrMsLtd.

Systematic Significance of Xanthones, Benzophenones and Biflavonoids in Garcinia PETER G. WATERMAN and RAOUF A. HUSSAIN Phytochemistry ResearchLaboratory, Departmentof Pharmaceutical Chemistry, Universityof Strsthclyde, GlasgowG1 1XW, UK Key Word Index - Garcinia;Guttiferee; xanthones;benzophenones;bifievonoids; chemowstamatica. Abstract - Variousparts of ten African speciesof Garciniaproduce a rangeof xanthones,benzophenones,biflavonoidsand lactones. Condensedtannins basedon procyanidin are also a majorfeature of some of the speciesexaminedand thesetend to be relativelydeficient in biflavonoids. Distributionof compoundsbetween plant partscan varymarkedlybut, in general, the distribution of xanthones, biflavonoids and benzophenonesamong these and other African speciesrelateswell to classical taxonomic divisions. However,when Asian speciesare included in the analysisonly the biflavonoids appearto show general promiseastaxonomicmarkerswithin the genus.All threeclassesof compoundsseemto possessvalueas taxonomic markers at the supra-genericlevelwithin the subfamilyClusioideae.

the investigation of a number of Garcinia species of west African origin [3-5, 7-10] and from the stem barks of these we have isolated and characterized a total of 19 compounds, including known and novel biflavonoids, xanthones, polyisoprenyl benzophenones and lactones. In this paper we report on the distribution of these compounds among the seeds, wood and leaves of some of the species already examined and, for the first time, on the constituents of the barks of G. afzelii, G. punctata and the east African taxon G. huillensis. We examine the potential importance of the isolated compounds to an understanding of the systematics of African species of Garcinia and then proceed to an analysis of the value of xanthones and related compounds and of bifiavonoids as chemotaxonomic markers in the genus as a whole.

Inmxluction Garcinia L., containing about 400 species, is the largest genus of the tropical family Guttiferae (excluding the Hypericaceae) [1]. Species, which are most commonly rain forest trees of the canopy and sub-canopy or more rarely shrubs, are restricted in distribution to the Old World, with a major concentration in south-east Asia and with secondary centres in the Indian subcontinent and in tropical Africa [1]. The fruit of several species are edible, that of G. mangostana L., the mangosteen, being quite widely cultivated throughout the tropics. Garcinia species are also characterized by the production of a yellow or occasionally white latex in the endocarp of the fruit, in the bark and perhaps also in the wood. There have, to date, been phytochemical investigations reported on about 30 species but many of these have been extremely fragmentary. They have, however, cleady demonstrated that the genus is a significant source of xanthones, which are characteristic of the Guttiferae as a whole [2], and is the major or sole known producer of partially reduced polyisoprenyl xanthones, similar polyisoprenyl benzophenonea and 3,8"-linked biflavonoids. Over the past 5 years we have been involved in

Results and discussion

Distn'bution of Compounds in Investigated Species Table 1 presents the known distribution of compounds, and where isolated the concentrations found, within and between the ten African species that we have investigated. All compounds have been isolated and characterized from at least one source, usually more, and these have then

(Received18June 1982) 21

2

BK(DE) 181 BK(K) 141 WD(DE)" LF(DE)* SD(DE) °

BK(DE) 191

G. mannii

G ovNffoh'a

BK(DE)I101 SO(DE)"

BK(DE) [ 10l

G. quadrifana

G. staudtii

0.32 0.15 + 4-

0.13

+

~+

0.12

0.15 0.06 +

0.19

0.22 44.

4-

+

0.03

4.

4-

0.23

4-

5

0.012

+

4

Biflavonoids 3

+

+

6

0.41

0.42 + +

10.4 +

0.8

4-

+

7

0.012 0.006

8

0.004

9

0.00~

10

Xantho~es

0.02

11

13

0.08

4-

+

0.56 0.02 + 4-

44.

0.008

12

B a n z ~

0.9

14

+

0.4

1B

0.35

+

0.8

1§

0.001

+

17

Lactones

19

CT

12.93

2.69 1.89

0.60

15.94 16.83 2.14 14.44 17.25

0.67 0.35 0.25 0.67 0.30

0.30 0.009 0.009 10.00 + + 33.55 1.00 + + 11.50 + + 5.29 0.45 0.43 0,20 6.25 1.15

18

*Reported here for first time. BK = stem bark, WD = wood, LF = leaf, SD = seed, FR = fruit, TW = twigs, DE = from Oouala-Edea Forest Reserve, Cameroun; K = from Korup National Park, Cameroun; G = from Gombe National Park, Tanzania. CT - condensed tannin content (procyanidin tannin). Vouchers: G. afzel6 Engl. - Thomes-6~ at Kew; G. contauana Engl. - Waterman and McKey-807 at Kew (for DE) and ~ 1 at Kew (for K); G. ~ Engl. - Waterman and McKey-8~ at Kew (for DE) and Thomas- 1175 at Kew (for K); G. huillensis Welw. ex Oliv. - Verdcourt (no number) at East African Hed=~rium, Nairobi; G. ko/a Heckel - no voucher; G. mantra" Oily. - Waterman and McKey-877 at Kew (for DE) and Thomas-729 at Kew (for K ); G. ovalifolia Oliv. - Waterman and McKey-79B and 810 (for DE ); G. punctata Oliv. - McKey-293 at Kew; G. quadrifariaBaiH.exP~rre WatermanandMeKey-802atKew;G. staudtiiEngl..-McKey-270atKew. The biflavonoids 1, 3, § and 7 from the previously uninvestigated G. huillensis and G. punctata were isolated as previously reported for compounds of this type 15l and were characterized as the malhyl ethers (UV, IR. ELMS, 1H NMR, t3C NMR) 15, 8, 151. TLC analysis of biflavonoids was performed on silica gel using a two-dirnertsior~ system consisting of (i) CHCI3-MeOH (4:1). and (li) toluene-ethyl formate-formic acid (5:4:1).

TW(DE)*

G.punctata

WD(DE)* LF(DE)" SD(DE)*

BK(K)*

BKIG)"

SD( I 16l FR( - ) 171

0.7 ÷ 4-

WD~DE)* LF(DE)" SDIDE)"

G. kola

÷

4

0.53 0.10

BKIK)"

0.15 0.Q2

1

BK(K)' BKIDE) 131 BKIK) 14l WD(DE)* LF(DE)" SO(DE)* BK(DE)I5I

G. hu#/an.~

G densivenia

G. afzelh" G. conrauana

Species, plant part and source

TABLE 1. DISTRIBUTION OF BIFLAVONOIDS, XANTHONES, BENZOPHENONES, LACTONES, CONDENSED TANNINS AND MISCELLANEOUS COMPOUNDS WITHIN AND BE'nNEEN TEN AFRICAN SPECIES OF GARCINIA (percentage content or ( 4- ) = TLC only)

7

_>

C

P

0

Z

XANTHONES,BENZOPHENONESANDBIFLAVONOIDSIN GARCINIA

been used as standards for TLC comparisons with extracts of plant parts that were available in small amounts only. Many samples also yielded procyanidin-based condensed tannins and these were quantified using the assay of Bate-Smith [11]. The results of some of the condensed tannin analyses have been reported previously [12], others are reported here for the first time. By far the most widely distributed group of metabolites are the biflavonoids, which have been found in all species examined except G. staud~7 and appear to be the major constituents in seven of the remaining nine, the exceptions being G. conrauana and G. ovalifolia. The biflavonoids isolated can be divided into two subgroups, those made up of one flavone and one flavanone sub-unit (1-2) and those made up of two flavanone units (3-7). Of these two types the biflavanones appear to be the more dominant being the sole or major components in G. afzelii, G. huillends, G. kola, G. mann# and G. punctata, whereas the mixed dimers predominate only in G. densivenia and G. quadrifaria. Co-occurrence of

NO ~

/0~. ~ J K , .~~ OH

R

!

R =OH

2

R =

R

OMe

~

OH

H0..,.~'-,,.,/0

R,

HO

0

:3 R=OH, R=,Rz=H 41 R=Rt=R2"H R" R==OH,Rz=H a R•OH, R=•OMe, R2•H 7 R" R='R2=OH SO Rp=OH,R'R2=H

23

the two types in a plant part is rare, being recorded only in the wood of G. densivenia and twigs of G. punctata among the 18 plant parts in which biflavonoids were found. However, the types change between plant parts, as is demonstreted by the occurrence of mixed dimers in the woods of G. conrauana and G. ovalifolia but biflavanones in the seeds of the former and leaves of the latter. The procyanidin condensed tannins have been found to occur in very high concentrations in G. conrauana, G. ovMifolia and G. s~ud~7. These are the three taxa that are devoid of or relatively deficient in biflavonoids and it is tempting to consider the formation of large amounts of biflavonoids or of flavonoid-based polymers (condensed tannins) as deriving from alternate biosynthetic routes. Xanthones (8-11) have been found in very low yields in the barks of four species, G. densivenia, G ovalifolia, G. quadrifaHa and G. staud~7, but not in either the eight other parts of three of these species or in any of the other six species examined. The woods of two of the xanthoneproducing species, G. densivenia and G. ovalifolia, were available in quite large quantities but an intensive examination failed to reveal the occurrence of xanthones: this is somewhat surprising in that most previous records of xanthones from Garcinia spp. have involved extraction from the wood. It should be noted that in all cases the xanthones isolated were present only in very small amounts and it appears probable that they form a less significant proportion of the secondary metabolic output among west African species than they do in some Asiatic species. The chemically closely allied benzophenones were found in all of the xanthoneproducing species except G. quadrifaria and in two species not found to contain xanthones, G. mannii and G. kola. In two species, G. ovalifolia and G. staudfii, xanthochymol (12) is the metabolite found in greatest quantity while in G. kola fruit pulp kolanone (14) is similarly important. In all other cases they appear to be minor components and benzophenones were not found in the woods of any species studied. The unusual lactones conrauanalactone (15) and its prenylated derivative (16) have only been found in G. conrauana where they are the major metabolites. Their distribution is rather interesting,

24

PETERG. WATERMANANDRAOUFA. HUSSAIN 0

OH

I

OH OH

15 R=H 16 R = CHzCH:CIMe)2 OH

0

Ho,,,,,, I~1

O~ 0.-'~,CHz)7CH:CH,CHz)7Me IO

17 OH H

O

~

OH

II 18

H 19 12

O

~

oo

H

with 15 having been found alone in stem bark collected in the Douala-Edea Forest Reserve whereas 16 replaces it in stem bark material from the Korup National Park and in wood from individuals at Douala-Edea that yield 15 from the bark. Both lactones are found together in leaves collected at Douala-Edea. The related furanolactone 17 occurs in the bark of G. mann# from either site but the quantities present are very small, in contrast to the high yields of 15 and 16 in G. conrauana. Similarly the flavanone eriodictyol

XANTHONES, BENZOPHENONES AND BIFLAVONOIDS IN GARCINIA

(18) and 5,7-dihydroxychromone (18), which may be derived from 18, are so far recorded only from G. conrauane, in which they have been found in all parts except the wood.

Chemical Systematics of the African Garcinia Species In the classification of Garcinia adopted in Engler and Prentl [13] the genus is divided into a large number of sections; the nine west African species examined falling into five of these (Table 2). In general the distribution of compounds shows good agreement with the sectional separation TABLE 2. CLASSIFICATION OF CHEMICALLY INVESTIGATED SPECIES OF GARCINIA OF AFRICAN ORIGIN [13] Section II III IV V Xlll

Species Rheediopsis Pierre Xanthochyrnop~s Engl. Te~ngium Engl. Xanthochymus 1". Anders Togmanthera Pierre

G. ovalifolia, G. staudtii G. volkensii G. conrauana G. densivenia, G. quadrifaria G. afzeli~ G. mannii, G. punctata, G. huillensi$, G. buchananii G kola

~5

distant from the other species in the section Teoaphelangium and its isolation is confirmed by its concentration on the synthesis of simple lactones. It has been suggested [14] that G. huillensis is impossible to separate from G. buchanenii Baker. The two texa are certainly very similar in their ability to produce large quantities of biflavanones; the heartwood of the latter yielding 3-6 and 20 [15], while the bark of G. huillensis gave 3, § and 7. However, the wood of G. buchanan# has also yielded xanthones [16] but these could not be detected in the bark of G. huBensis. The only other information on an African species concerns the biflavonoids of the heartwood of G. volkensii Engl. from Kenya [17], which has yielded two blflavanones and two mixed dimers. The relative importance of the mixed dimers may reflect an affinity with G. densivenia and G. quadrifaria, which is suggested by its placement in the allied section Xanthochymopsis.

Chemical Systema~'csat the Genericand Higher Levels The available information on the occurrence of xanthones, benzophenones and biflavonoids in among these species. Thus the two species 31 species of Garcinia is given in Table 3. For the included in section Rheediopsis, G. ovalifolia and xanthones substitution patterns are given and a G. staudti~ both have stem barks devoid of points score based on the system proposed by biflavonoids and that have yielded both 1,3,5,6- Razende and Gottlleb [2]. oxygenated xanthones and the benzophenone Just over half of the species investigated to xanthochyrnol. G. quadrifaria and G. densivenia date have yielded xanthones. These have exhibiare placed together in section Xanthochymus and ted a large number of substitution patterns that are the only other species to produce xanthones suggest reduction from both 1,3,5,6-oxygenated and the only biflavonoid-producing species to and 1,3,6,7-oxygenated types. Methyletion and concentrate on mixed dimers rather than biflava- C-prenylation, but not O-prenylation, appear nones. The three species that concentrate mainly quite common with the formation of additional on the production of blflavanones, G. rnannii, G. 2,2-dirnethylpyran rings a feature of many afzelii and G. punctate, are all placed in section species. Application of the Razende and Gottlieb Tagrnanthera although in the case of G. mannii system [2] shows lowest scores among souththere appears to be some confusion as it was also east Asian species producing 1,3,6,7-substituted placed in section Co#anThera. Clearly the compounds and African species producing chemistry of this species favours its relationship 1,3,5,6-substituted derivatives. Highest scores are with others placed in Tegrnanthera. seen among the Indian and Sri Lankan species Of the remaining two species, G. kola, which is and in G. eugeniifolie from Sarawak. At present it placed in section Paragarcinia, bears obvious does not appear possible to demonstrate any affinities to the Tagmantheragroup in its produc- relationship between the occurrence of tion of large amounts of biflavanones. On the xanthones and the sectional classification of the other hand, the absence of A-ring oxygenation in genus [13]. kolanone (14) is unique among the benzopheBy contrast, less than a third of investigated nones of the genus. G. conrauana is also placed species have yielded benzophenones and, so far, XXXlV

Paragarcinia VeSclue

28

PETER G. WATERMAN AND RAOUF A. HUSSAIN

TABLE 3. DISTRIBUTION OF XANTHONES, BENZOPHENONES AND BIFLAVONOIDS IN GARCINIA SPP,

Section and specias I

Xanthone= [substitution patterns and points score)

Benzophenones

Bifiavonoids A O

Refs.

Teracentrum Pierre

It

G, liw'ngstonii T. Anders

India

Rheed/ops/s G. ova~o//a G. staudU7

W. Africa W. Africa

III

Xanthochymopsis

IV

Tetraphalangium G. conmuana Xanthochyrnus G spicate Hook, f. G. densivenia

G. v~/kendi

V

Source

G. du/c/s Kurz G. qudldrffm'ta

G. xanthochymus Hook, f. VI Plinthostigrna Pierre G. multiflora Champ. VII C~mogin Pierre G. indica Choisy G. Cambogia Desr. VIII Anisostigma Pierre G. pedunculata Roxb. Xltl Tagmanthera G. mann# G. punctata G. buchananii G. huil/en$is G. afzelii XV Mangostana Choisy G. hombroniana Pierre G. mangostana XIX Discostigma Pierre G. terpnophy/la Thw. G. eugeniifoliaWaH. XXIV Echinocarpa Pierre G. achinocarpa Thw. XXIX Oxycarpus Lour. G cowa Roxb. G. rubra Merr. XXXI Hebradendron PI. et Tr. G. more/la Desr G. handburyi Hook. f. XXXIV Paragarcinia G. kale

+

118]

+

+

19] [10]

E. Africa

+

+

117]

W. Africa

+

+

13, 4)

+ +

+ + + + +

119, 20} [5] [21 ] 110] 120, 22, 231

+

+

[20, 24}

1356 1356

1.0 1.0

+ +

SE Asia W. Africa India W. Africa India

1356

1.0

+

135 15/17/125/135

1.0 3.8

+

Taiwan

1367

2.0

India Thailand

17 136"

4.0 3.0

+

+

India

13571136"7

2.5

+

+

+

+

+ +

+

W. Africa W, Africa E. Africa E. Africa W. Africa

15/156

3.3

SE Asia SE Asia

135/137/1367/1358

1.0

Sri Lanka SE Asia

15/17/1367 17/137/156/167 d347

2.7 3.4

Sri Lanka

15/1367

3.0

SE Asia SE Asia

1367 1367

0.2 0.0

India SE Asia

136° 136°

2.0 2.0

W. Africa

+

1251 122, 27] 125} 14, 8}

+ + +

[15, 16]

+

125]

+

[30-33]

+ +

[34] 135, 36] +

{34]

137, 38}

[39] +

+

+

125, 40-42] [39] [6, 7]

Not Assigned

G. linii Chang G. talbotii Roxb.

SE Asia India

+

+

120]

A = biflavanones; B = mixed flavanone-flevone dimers and biflavone dimers; ° = xanthones with reduced B-ring.

they are restricted to west African and Indian species. The majority are polyisoprenylated compounds which have undergone reduction and cyclization reactions analogous to those seen in the xanthones of G. morella. As with the xanthones it does not, at present, appear possible

to relate benzophenone distribution to sectional classification. Two points that may be of systematic value are: (i) the occurrence of a number of species that can be termed "benzophenone specialists", in that this type of compound dominates secondary metabolism

27

XAN'FHONES, BENZOPHENONESAND BIFLAVONOIDS IN GARCINIA TABLE4. DISTRIBUTIONOF XANTHONES,BENZOPHENONESAND BIFLAVONOIDSWITHIN THE GENERAOF THE CLUSlOIDEAE Tribe and genus Ctuskme Ous~ L. TovorniraAubl. Chrysochlarnys Poepp. Garciniae G~dn~ Alenb/ack~ Oily. R/~esYaL. Pentaphdang/umWarb.

Xanthones (substitution patterns)

Benzophenones

Biflavonoids

146,46] 147,48]

1,6,7/1,2,3,5/1,3,4,5/1,3,6,7

Sss Table3 1, 5/1, 7/1, 3, 5/1, 3, 7/1, 3, 6, 7 1, 5/1, 7/I, 5, 6/1, 3, 5, 6 1.5/1, 3, 5/1, 3, 7/1, 3, 6, 7

Ref.

+ +

+ + +

[50, 51) [52]

(e.g.G. ovalifolia, G. staudtii, G. xanthochymus, lady many of the xanthones found in both AllanG. cambogia, G. homobroniana); (ii) all benzo- b/ac/ob and Pentaphalangium have also been phenone-producing species that have also yielded found in Garcinia and all of the oxygenation patterns of the xanthones are shared between xanthones exhibit 5-oxygenation of the latter. Of the 31 species in Table 3 at least 25 yield these genera. By contrast Tovomita, the only biflavonoids, 17 biflavanones and 17 mixed genus of the tribe Clusieae that has yielded dimers with nine producing both. There is still xanthones to date, exhibits oxygenation patterns insufficient data to permit a realistic appraisal of (1,2,3,5 and 1,3,4,5) yet to be found in any of the the value of these compounds as taxonomic genera of the tribe Garciniae. The dose relationship between Garcinia, Allanmarkers but at present they do appear to hold out some promise. For example, production of mixed b/ac/o~ and Pentaphalangium is further demonclimars dominates in all five species of section strated by the co-occurrence of 3,8"-linked Xanthochymus whereas biflavanones are just as biflavonoids. At present these remain the only dearly dominant among the five species of sources of biflavonoids linked in this way section Tagrnanthera and two species of Disco- although 3',8"-linked dimers are known from s~gma. It would seem likely that the biflavanones both G. dulcis [21] and G. livingstonii [18], and are the precursors of the flavanone-flavone 8,8"-linked from Mesua ferrea L. [53]. On the dimars, the latter being derived via dehydration of other hand, the link between the two vibes of the Clusioideae is confirmed by the co-occurrence of a 3"-hydroxybiflavanone (e.g. §-* 1). If this is so then the mixed dimers can probably be assigned a the complex polyisoprenylated benzophenones, derived position relative to the biflavanones from with and without A-ring oxygenation, in Garcinia and in Clusia, a genus that has yet to yield an evolutionary view. At present the systematic value of these xanthones but which has been little studied. compounds is demonstrated more clearly above Simpler benzophenonas some with prenylation but the generic level. Garcinia forms part of the sub- lacking the complex cyclizations of 12-14, have family Clusioideae [44], which is in turn divided been found in A/lanblackia and in Moronobea into the tribes Clusleae and Garciniae. The distri- pulchra Drake [54]. They also occur in Vismia bution of xanthones, benzophenones and biflavo- decipiens Cham. of the closely allied Hyperinoids among the six investigated genera of the aceae [55]. subfamily is shown in Table 4, and clearly shows close agreement with the proposed classification. Among the xanthones, those found in Rheedia Acknowledgements - The following are gratefully acknowappear perticulady close to those of Garcinia, ledged for assistance in the collection of plant materiel: Dr. J. Gartisn, Wisconsin Regional Primate Research Center; most notably the co-occurrence of macluraxan- S. Dr. D. B. McKey, Zoologisches Institut, Universit~t Basel; thone (10) and rheediaxanthone-A (11) in Dr. R. W. Wrengham, Department of Anthropology, Rheedia benthamiana PI. et Tr. and in the species University of Michigan. Dr. N. K. D. Ross, British Museum, of Garcinia, section Rheediopsis (Table 3). Simi- gave valuable assistance in identifying the species studied.

28

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