PAyrockmufry. Vol 26. No I. pi 171-174. 1987 0031 -w2 87 $3.00 + 0 00 Pnnlcd on Great Bnutn Pcrgmwn Jounuls Ltd

SYSTEMATIC IMPLICATIONS OF CHROMENES AND BENZOFURANS FROM ENCELZA (ASTERACEAE)

PETER PROKSCH and Cuttrts CLARK*

Ins~itut f&r Pharmazeutischc Biologic der Tachairhen Univmibt Brounschweig Mendchsohnsrr I. D3MO Braunschwclg. War Germany; *Biological scimca. Cahfomia Stale Polytechnic University, Pomona. CA 91768, U.S.A.

Key Wad Index-Encelio; As~eraceac; chronxna; bcnzofunns: chcmosplcmalics.

- .-_. _. .__ ._- _ Abetract-Phytochemil analysis of Encelia species yielded numerous chromenc and benzofuran derivatives. Systematic comparison of the chemical pro&s revealed three distinct groups of species. The spcctcs affinittcs, as clucidatcd by the chromcnc and bcnxofuran patterns. largely paralkkd those based on recent morphological and anatomical findings and are thought IO reflect the phylogeny of the genus.

- -

IKlRODlJCTlON

Enceliu Adans. comprises around 18 species of shrubby perennials inhabiting the arid and semi-arid parts of southwatem North America [I J. The distribution of the genus largely overlaps the Sonoran and Mojave deserts. whcrc several spccics. notably Efarinosu, are widespread and dominant ckmcnts of the desert floras. Two specks are disjunct in South America; one, E. cunescens, grows along the coasts of Peru and Chik and in northcm interior ArgentiM; the other, E. hispido, is endemic to the Galapagos Islands.

Encelia is a mcmbcr of the tribe Helianthcae and has born assigned either to the subtribc Helianthinae 123 or Ecliptinae [3]. Robinson considers Em&a to belong to a group of related genera including Enceliopsis. Gaaea, Flouretuia and Phoebonhs; they share morphological features such as pale anther thccae and sterile rays 133. Historically, this group of genera has been a difficult subject for systematic study. The generic ~undarics have been repeatedly rcdcfmcd; many species originally as- signal to Encelia have been transferred lo Enceliopsis. Geraeo and even gcncra apparently outside this group, such as Simslo and Helianrhus [I J. Reproductive barriers arc lacking within the gcncra. while intergmeric hybrids have sharply reduced fertility (14,s); C. Clark and D. W. Kyhos, unpublished results).

As part of a hrgcr investigation of this group of genera we have studied Encelio with emphasis on its chrommc and bcnxofuran chemistry. Chromencs and bcnxofurans are prominent natural products of many genera of the Astcraccac and are especially common in the tribes Astcreae. Eupatorieae. Hcliantheac, lnukac and Scnecioneae [6,7]. Whereas the presence of these com- pounds in species of Encelio has occasionally been rcportcd before [&IO]. this study provides a comprehen- sive analysis on the distribution and systematic impli- cations of chromcncs and bcnzofurans in this genus.

Chromcncs and bcnzofurans are among the major natural prducts of Encelia and may in some spec~a

account for up IO So/, of the dry weight of kavcs and stems. The compounds usually occur in all organs anal- yscd, including roots, stems. kavcs, inflorcsccnses and achencs [ 1 I]. Within the plant tissues the compounds are stored in resin ducts, as was demonstrated by chemical analysis of the rain of E. /uritwsu and by fluorescent microscopic analysis of the plant tissues [ 121.

To obtain rcproducibk and reliabk information on the chromencand bcnxofuran content and patterns within the genus, equal amounts of air-dried leaves and slcms of each taxon were extracted separately. The extracts were rc- duccd m volume and used for comparative TLC or HPLC. All major chromcnes and bcnzofurans detected (compounds I-14) were consecutively isolated from bulk samples and identified spectroscopically. Companson of the chemical data obtained revealed thra distinct groups that could be clearly distinguishcd by their chromene and/or bcnxofuran constituents (Table 1).

The largest group. Group 1. consists of 1 I taxa that are characterized by an abundant accumulatton ofchromencs and bcnzofurans. up to a total concentration of 2-S “/, of the dry weight. The chromenc and bcnzofuran patterns of all taxa of Group I proved to be remarkably homoge- neous, with simpk compounds like encccalin (3) or cuparin (4)prcdominating. Howcver,all taxa of this group also elaborated unusual dimers that regularly consisted of a euparin and encccalin motcty. Such dimcrs have only been lsolatcd from Encelia so far [ 131 and ckarly demonstrate the close rclatiomhip of the taxa in this group. The dimen reached their highest concentrations in the stems; in the leaves they were somctima prcscnt only as trace amounts.

The observed chemical patterns proved IO bc consistent when scvcral populations of one species were compared (Tabk I), even though there wcrc great differences m the amounts of the compounds accumulated. Ten sampks of Ejarinosa colkctcd within 2 days in August 1984. along a wesl*t transect from Palm Springs. Californta to Tcmpc. Arizona, rcvcakd a gradual decline in the concen- tratioo of cncecalin. resulting in a ten-fold difference bctwan the eastern and wcstem extremes (Figs I and 2).

Group II consists of four taxa that show a markedly reduced capacity IO accumulate chromcncs and bcnxo-

172 P. PROKSCH and c. CURK

1 R=H

2. R - OH

3 R=OMe

L R=OH

5 R=OMe

6 RI=

9

0 7 R,= 4 ,Rz=OH ? ’

R

+=Y Me0 :I I

0

0

0 8. R : ? ’ 4 0

9. R =

furans. Compounds like encecalin (3) of tuparin (4)

produced so abundantly by the taxa of Group I are. with the exception of E. renui/o/m, only trace compounds barely vistbk on TLC plates (Table 1). Dimcrs were missing totally. All the taxa of Group II. however, elaborated new cstcritied 2,Sdtacetylbenzofuran derivat- ives (8.9) [ 141. These novel compounds provided cvi- dencc of the affinities of all taxa in this group.

Me0

Group 111 consists of only two taxa. As in Group II, there was a marked trend towards reduction of the capacity to accumulate chromcna and bcnzofurans. Again only trace amounts of cncaalin were detected (Tabk I). But in contrast to Group II, no diacetyl- bcruofurans were present, allowing a clear distinction between these last two groups. The chemical data proved agam to bc stable when different populations of the taxa were compared (Table 1).

X remaining two taxa, 15. srenophyllu and E. roposa. both elaborated compounds that were not observed in any of the previously described taxa. Estcritied derivatives of tremctonc and hydroxytremetonc (6.7) were isolated from E. srenophyllo (Tabk I). E. scoposu yielded several novel bcnzofuran derivatrvcs (1 l-14). The spectroscopic elucidation of the structures will be published elsewhere (M. Breuer. P. Proksch and H. Budzikicwk in prcpar- ation). One compound (11) was a 2-acetylbctuofuran dertvative recently isolated from Eupurorium srerber- gionum [IS]. Compounds 12-14 were novel estertied bctuofuran derivatives. The acid substitucnts (angelic, xnccic and tiglic acids) were cstcrihed via the hydro- xyethyl group at C-4. These compounds were further charactertxed by an acetyl substttucnt at C-2 of the furan ring. The unusual nature of the bcnxofurans (no acetyl group present at the aromatic ring of II, hydroxyethyl group present at C-4 of compounds 12-14) clearly differentiated E. xaposu from the rest of the taxa analyscd in this study.

These chemical data agra very well with morphological features to substantiate general ideas of phylogcnctic relationship. The presence of the unique dimcr in Group I suggests that it LS a monophylctrc group, and this is supported by the prcscncc in all the members of ray corollas that reflect UV light (vrsiblc to insect pollinators) [ 163 and of a unique moniliform uniseriatc trichomc [17. 181.

The reduced capacny of Groups II and 111 to ac- cumulate chromcncs and benzofurans is a unique feature, at least among Enceliu and its relatives. suggesting that these groups taken together are also monophyktic. This, too, is supported by morphology. al) members having a

cluomena and bcnzofurans from Enceliu

Tabk 1. Dwribution of chromena and bcnzofurans in Encelia*

spacia Cdkclionst Compound8

- .._- __-_ _.. .- - --. - _.--- --.. -- ._- GROUP I

6. coli~omico NutI. 6, I IS. 123. 166 1, t 3. 4. 5. 10

E. CanesccN lam. 267. 268. 273, 281 1. t 3. 4. 5, 10

E. Benth. cimspersa 189 1. 2 3, 4. 5. 10

E. ‘denstjolio” (undescnbcd sp.) 184 I. r 3. 4. 5. 10

E. /uwtosu A. Gray In Ton I. 2 3. 4. 5. 6. 7. 8. 9 I. 2. 3, 4, 5. 10

var. fwitwsa 10.13.18.37,247.248

E./winosa var. phenicodoara (S. F. Blake) I. M. Johnston 82-163.8. 183. 186. 190. 202 1. 2 3.4.5. 10

E. Minujalio Cav. 143 1. L 3. 4 s, 10

E. Y ltxvviato Vasey & Rose 82-164 I, 2, 3. 4. 5. 10

E. polnvri Vasey & Row 58. II8 1. 2 3. 4. 5. 10

E. rdhnr Bran&gee 203 1. L 3. 4. 5. 10

E. LWIIO~KW Brandqce I31 I. z 3, 4. 5. 10

GROUP II

E ocront Elmer II. IO& 1% 3 @r). a 9

E. /NICYCN A. Gray var. reshow Jones ex Blake 195. 198 3 w a 9 E. rownii Wiggms I65 3 w a 9

E. ‘renu~olti” (undescnbcd sp.) 200 3 w a 9

GROUP Ill E. asperlfolia (S. F. Blake) Clark & Kyhos I -

106. I29 3 w

132 3 w). 4 &I E. jrurermr A. Gray var. ~rurrscens ls4.210 3 (tr)

174. I76 -

OTHER SPECIES

E. woposa A. Gray I87 11-14

E. srenophyllo Gromc I33 1.4ls.6,7

*The reportal compounds were analyscd in Leaves ami stems. tColkctmn datn of thrcedlgit numbers can be obtaiocd from C. Clark. thoac of onedigit and twodiglt numbers

from P. Proksch. Voucher spenmms referring to the coIlaction numbers reported have ban deposited at CSPU.

I73

Fig. I. Popula~ional analysu of major chrorrkmcs and btmzofuran from Encr/tu /cyinosa.

174

J

Fip. 2. Southwestern Umted States and adjacent Mexico. TM hat&d area resembles the natural datnbution of &rrrc/iu farmosa (mod&d after ref. [23] ). E. j&row, ts gradually replaced by E. farinou var. pknbdonta and E. rOdi4N (pre- vmusly E. jurinoul var. rnd~cvu) from north to south in the

penmsula of BaJa Cahfornia.

specialized development of the pakae in the fruiting head ([ 193; C. Clark, unpublished results).

It would further appear that Group II itself ts mono- phyktic. diagnosed by the unusual diacetylbenzofurans. However, at present morphology contradicts this assess- ment. Two of the taxa of Group II. E. tenuijo/iolio and E. /rurescens var. resino.w, share with both taxa of Group 111 a unique broad-based uniseriatc trichome [ 17. IHI; the group possessing this character overlaps Group II, so at most one group can be monophyktic. It seems perhaps more reasonable to postulate that Group III secondarily lost the ability to synthesize diacetylbenzofurans, than to assume that a complicated trichome evolved twrce independently.

The clear chemical differences between E. jrufescens

and its variety resinosu call into question the conspecificity of these taxa. Preliminary studies of their Aavonoids also show significant dtfferences (P. Proksch. in preparation). The taxa are morphologically quite distinctive as well, and specific status is being proposed for the variety resin0.w (C. Clark, in preparation).

Encelia asperijolia is especially interestmg in light of its apparent origin from the stabilization of diploid hybrids between E. co@rnico (Group I) and E.frurescenr (Group III) [2-O]. It shares with E.jrurescens the inability to accumulate chromenes and benzofurans. and with E. colijornico the presence of UV-reflecting ray corollas.

The chemical differences between E. sfenophyllo and E. scaposu and the rest of the genus are paralkkd by morphological differences (C. Clark, unpublished results), suggesting that these species, like many other species that have been removed from Encelio in the past, have their a!Tinities eLsewhere.

Whereas chromenes and benxofurans are already a- tabhshed as chemical markers which serve in delineating tribal affinities within the Astcrruzeae [6.7,21]. this study for the first time demonstrates their importancc for chemosystematic analyses at the s&generic level. Furthermore, recent findings indicate this class of natural products to be a useful tool for gaining insight into the generic affinities between the r&ted genera Encelio. Enceliopsis, Floutensia and Geraeu (P. Proksch and C. Clark, in preparation),

LXPERIMCNTAL

The compoundr reported ID th1.5 study were ong~nally isolated from bulk sampks of one colkctioo of each (axon analysed and identified by tberr ‘HNMR and mass qmtra The spectral cbaracteristtcs as well as details on the cxtractmn and isotxuon prccedura employed can be obtained from refs [Il. 13. 14.221. Comparottve analyses on the chromene and benzofuran patterns of the various populations studmd of each taxon were conducted by TLC and HPLC of crude extracts [22].

Acknowledgrnwnrs We thank Jutta Bodden for expert tschnwml assistance. This project was supported by grants of the DFG (Pr 229. I-l and 229,‘l 2) awarded to P. P.

I. 2.

3. 4.

5. 6

7. 8

9. IO.

Il.

12.

I3

14.

I 5.

16. Il.

18.

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