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Leaf flavonoids in Cotoneaster wilsonii (Rosaceae) from the island Ulleung-do, Korea
Biochemical Systematics and Ecology 31 (2003) 171–179 www.elsevier.com/locate/biochemsyseco
Leaf flavonoids in Cotoneaster wilsonii (Rosaceae) from the island Ulleung-do, Korea Chin-Sung Chang a,∗, Jeong Ill Jeon b a
Department of Forest Resources and The Arboretum, Seoul National University, Suwon, 441-744, South Korea b Department of Urban Horticulture, Shingu College, Seongnam, 462-743, South Korea Received 12 June 2001; accepted 24 January 2002
Abstract The leaf flavonoids of Cotoneaster wilsonii, an endemic shrub to the island Ulleung-do in East Sea of Korea, were characterized and compared with flavonoids in some eastern Asian Cotoneaster species. The highly specialized taxa belonging to sect. Cotoneaster (sensu Yu¨) including C. wilsonii produced a mixture of flavone O- and C-glycoside and flavonol O-glycosides, including isorhamnetin glycosides. The morphologically similar species, C. multiflorus and C. hebephyllus of central China and C. wilsonii of Korea had similar flavonoid profiles. In addition, C. tenuipes, C. zabelii, and C. dielsianus (sect. Cotoneaster, ser. Integerrimi) had similar flavonoid patterns to taxa in the C. multiflorus complex (sect. Cotoneaster, ser. Multiflori). This indicated that the chemical data cut across Yu¨’s serial treatment within section Cotoneaster. Earlier studies showed that there were few absolute differences between many of the other woody plants growing on this island and on those on the Korean peninsula and mainland China, or the Japanese archipelago. C. wilsonii appears to be another example in which no change in chemistry or morphology has occurred. Many of the woody plants on this island are very recently derived and their progenitors were historically more widely and continuously distributed in eastern Asia. 2002 Elsevier Science Ltd. All rights reserved. Keywords: Flavonoids; Cotoneaster wilsonii; Cotoneaster multiflorus; Island Ulleung-do; Classification
∗
Tel.: +82 31 290 2322; fax: +82 31 295 6660. E-mail address: [email protected] (C.-S. Chang).
0305-1978/03/$ - see front matter 2002 Elsevier Science Ltd. All rights reserved. doi:10.1016/S0305-1978(02)00064-9
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1. Introduction The ornamentally important genus Cotoneaster of the Rosaceae is relatively small with ca. 90 species of shrubs (Yu¨ , 1974; Willis, 1985). The distribution of this genus extends from North Africa to North America, including the Himalayas, Siberia, and China (Rehder, 1940; Kru¨ ssman, 1976). The center of distribution is in China, where almost 60% of species occur, but no species occur in Japan. Cotoneaster wilsonii Nakai, which is characterized by a small number of flowers arranged in a corymb, red fruits and deciduous leaves, is endemic to the small island of Ulleung-do, Korea (Lee, 1980, Fig. 1). C. wilsonii is confined to two areas on the island and comprises fewer than 150 individuals, suggesting it may be endangered. Since this taxon has not previously been investigated, its taxonomic position is not known. The island Ulleung-do is located ca. 150 km off the coast of Korea at 37° 30⬘N and 130° 51⬘E (Lee and Joo, 1958; Pigott, 2000). This island is of late Tertiary origin from three to five million years old (Yang, 1996). Although its origin is controversial, it is known that it was connected to the Korean peninsula and the west coast of Japan until the early Quaternary (Yang, 1996). Ulleung-do has a unique flora of ca. 180 woody species of which ten are endemic to the island (Lee and Joo, 1958).
Fig. 1. Geographic distribution of Cotoneaster wilsonii Nakai in the Island Ulleung-do of Korea and its related two taxa in central China.
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The flora has evolved from isolated species, mainly associated with the Korean peninsula, the Japanese islands, and central China. The two major alternative classifications of Cotoneaster by Rehder (1940) and Yu¨ (1974) conflict unanimity in the circumscription of relationships. Thus, Redher (1940) divided the genus into two sections, Orthopetalum and Chaenopetalum, while Yu¨ (1974) divided the genus into three sections, Densiflos, Cotoneaster, and Uniflos with many series. Rehder recognized only two sections: Uniflos and Cotoneaster combined in sect. Orthopetalum, while he included some species of Yu¨ ’s sects. Densiflos, Cotoneaster, and Uniflos into sect. Chaenopetalum. Rehder’s treatment was based entirely on the position and colour of the petals and the number of styles. On the other hand, Yu¨ indicated that inflorescence type and the number of flowers might be suitable characters for the circumscription of taxa and the determination of natural sectional groupings. According to Yu¨ , sects. Densiflos and Cotoneaster can be separated from sect. Uniflos on the basis of the former having many flowers arranged in a corymb and large leaves. Furthermore, sect. Densiflos differed from sect. Cotoneaster in having many flowers (more than 20) with a compound corymb. Based on the gross morphology, C. wilsonii appears to belong to sect. Cotoneaster sensu Yu¨ . Yu¨ (1974) considered 29 taxa members of sect. Cotoneaster and further divided this section into three series: Multiflori, Acuminati, and Integerrimi, with respect to flower colour, leaf pubescence, fruit colour and the number of seeds. However, this treatment was somewhat artificial because of occurrence of many intermediate taxa among three series. A preliminary study of Cotoneaster taxa in eastern Asia showed that flavonols were the main leaf flavonoid constituents, and flavones and others from C. wilsonii were found unlikely. The purpose of the present study was to 1) identify the leaf flavonoids of C. wilsonii, 2) determine the flavonoid of components in some of the closest postulated mainland relatives, and 3) consider possible evolutionary implications of the flavonoid data.
2. Materials and methods 2.1. Plant materials Leaf material of C. wilsonii and 29 taxa of Asian Cotoneaster were surveyed using herbarium specimens at The Arboretum of Seoul National University (SNUA) and Institute of Botany, Academia Sinica, Beijing (PE) and fresh samples from the Chollipo Arboretum and the Island Ulleung-do. The collection numbers and places of deposit are given in Appendix A. 2.2. Phytochemical analysis The extraction of flavonoids from leaves followed the methods of Mabry et al. (1970); Chang and Giannasi (1991), and Giannasi (1975). The flavonoids survey
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employed two-dimensional paper chromatography using Whatmann 3 MM paper. TBA (tertiary-butanol: acetic acid : water, 3:1:1) was used as the first solvent and 15% HOAc (acetic acid:water, 15:85) as the second solvent. Flavonoid profiles were viewed under UV light and colours were recorded before and after fuming with ammonia vapour. Compounds were identified using standard UV-visible spectroscopy (Giannasi, 1975; Mabry et al., 1970; Markham, 1982). Aglycones obtained from acid hydrolysis of the glycosides were identified in the same manner to determine the location of sugar(s). Several compounds were compared directly with compounds identified from previous studies (Chang and Giannasi, 1991; Chang, 1998; Chang, 2000). Spectral and Rf data were compared with published data (Mabry et al., 1970). Flavonoid glycoside sugars were identified using trifluoroacetic acid hydrolysis, and co-chromatography with standard sugars in one-dimension, using ethyl acetate: pyridine: water (5:1.6:1) as the solvent. The sugars were visualized by spraying with p-anisidine hydrochloride (Pridham, 1956) followed by heating (100 °C, 10 min). 3. Results The leaf flavonoids identified in the selected taxa are tabulated in Table 1. They included five flavonol glycosides: quercetin-3-O-glucoside, quercetin-3-O-rhamnoside, quercetin-3-O-rhamnosylglucoside, isorhamnetin-3-O-galctodide, and isorhamnetin-3-O-rhamnose-glucose (unknown order); two flavone glycosides: apigenin-7O-glucoside and luteolin-7-O-glucoside, and three flavone C-glycosides: vicenin-A and -B (Markham and Wilson, 1989), and luteolin-8-C-glycoside. Flavonols, primarily as the 3-O-glycosides of quercetin but with occasional isorhamnetin 3-O-glycosides, were detected in all taxa of sections Densiflos and Uniflos and one series (Acuminati) of section Cotoneaster. Flavonoid analysis of C. wilsonii yielded a series of flavone O- and C-glycosides, and a methylated flavonol. Especially noteworthy was the occurrence of flavone O-glycosides, which were characteristic constituents of C. mutiflorus Bge, C. hebephyllus Diels (series Multiflori) and C. tenuipes Rehder et Wilson, C. dielsianus Pritz. and C. zabelii Schneider (ser. Integerrimi of sect. Cotoneaster). The presence of isorhamnetin 3O-rhamnose-glucose in C. wilsonii and some other species e.g., C. submultiflorus Popov, C. mongolicus Pojark., C. mutiflorus, and C. hebephyllus of ser. Multiflori of sect. Cotoneaster and in two taxa of series Integerrimi, C. tenuipes and C. dielsianus, may also have some taxonomic significance. Tabulation of the identified flavonoid (Table 1) failed to distinguish clearly the three sections recognised by Yu¨ (1974), although species in sect. Cotoneaster, ser. Multiflori, except for C. sylvestris, do share the same flavonoid profile. However, four taxa in series Integerrimi and one from section Uniflos (Yu¨ , 1974) also have very similar patterns. Thus, the flavonoid data tend to cut across the serial treatment within section Cotoneaster. In several earlier investigations on flavonoids of European taxa of Cotoneaster (Palme et al., 1994, 1996) the C-glycosylflavones (vitexin, vitexin-2⬙-O-rhamnoside,
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Table 1 Flavonoids distribution of Cotoneaster taxa considered in eastern Asiaa Taxonc
Flavonol O-glycoside 1
sect. Densiflos ser. Salififolii C. salicifolius var. floccosus C. salicifolius cv. Herbstfeuer C. salicifolius cv. Repens ser. Frigidi C. glabratus sect. Cotoneaster ser. Multiflori C. multiflorus var. multiflorus C. multiflorus var. calocarpus C. wilsonii C. mongolicus C. hebephyllus C. soongoricus C. submultiflorus C. sylvestris ser. Acuminati C. acutifolius ser. Integerrimi C. ambiguus C. lucidus (=C. ambiguus?) C. zabelii C. tenuipes C. dielsianus var. dielsianus C. dielsianus var. elegans C. franchetii C. amoenus (=C. franchetii?) C. obscurus C. sikangensis (=C. obscurus?) sect. Uniflos ser. Microphylli C. dammeri cv. Coral Beauty ser. Distichi C. apiculatus C. adppressus C. divaricatus C. horizontalis C. nitens
2
3
Flavone C-glycosylflavone
O-glycoside
6
7
8
9
10
X X X (X) X X X
X
X
X
X
X
X
X X
X X X
X
X X X X
X
X
X
X
O-Meglycoside 4 5
X X X
X X
(X)?b X X
X
X
X
X X X X X X X X
X X X X (X) X X
X X X X X X X
X
X
(X)
(X)
X X X X X
X (X) (X)
X
(X)
X
X
X
X X X X X X X X X
X X X X X X X X X X
X X
X X X X X X X X X X
X
X
X
X X
X
X X X X X
X X X X X
X X X X
X
X X
X X
a 1: quercetin 3-O-glucoside, 2: quercetin 3-O-rhamnoside, 3: quercetin 3-O-rhamnoglucoside, 4:isorhamnetin 3-O-galactoside, 5: isorhamnetin 3-O-rhamnose-glucose (random order), 6:vicenin A, 7: vicenin B, 8: luteolin 6-C-glycoside, 9: apigenin 7-O-glucoside, 10: luteolin 7-O-glucoside. b (x)?=compounds whose presence/absence is not clearly established from examination of the chromatograms and (x)=compounds detected in less than 50% of surveyed individuals. c classification sensu Yu¨ (1974).
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vitexin-2⬙-O-arabinoside), flavonols (rutin, quercetin-3-O-rhamnoside, kaempferol-7methyl ether 4⬘-O-glucoside) and 5,7,2⬘,5⬘-tetrahydroxyflavanone and its 7-O-glucoside were found in C. thymaefolia Hort. and flavonols and isoflavones (genistein-5methyl ether 4’-O-glucoside, biochanin and biochanin-7-O-glucoside) were observed in C. simonsii Baker. No C-glycosylflavone, flavanones or isoflavones and only flavonols, however, seemed to occur in Asian taxa. The current flavonoid survey provides some evidence to link ser. Multiflori and ser. Integerrimi. This study showed that the flavonoids’ profiles of C. wilsonii are almost identical to those of C. multiflorus.
4. Discussion 4.1. Systematic implications and biogeography The distribution of flavonoids compounds does not correlate well with the general grouping of sect. Cotoneaster into three series proposed by Yu¨ (1974). Some species of both ser. Multiflori and ser. Integerrimi, which are characterized by having one or two seeds per fruit, appear in each chemical group. With the exception of C. multiflorus and the C. dielsianus complex, no distinct species specific pattern was chemically observed in the studied taxa in the three different series (Multiflori, Acuminati, Integerrimi) of sect. Cotoneaster or in the other two sections. The division of section Cotoneaster into three series proposed on morphology received weak support from the comparative flavonoid data. Although intraspecific variation and the general lack of flavonoid diversity within Cotoneaster as a whole somewhat limit the taxonomic value of flavonoids at the specific level, certain observations are warranted. It appears that C. multiflorus and the C. dielsianus complex, placed in separate series in Yu¨ ’s classification, are actually more closely related than this classification implies. Our results suggest that these taxa may best be included within the same group. However, pending a complete survey of all Chinese taxa, no formal nomenclatural changes are proposed here. The taxonomic significance of the flavonoid data in C. wilsonii lies in their correlation with morphological evidence. C. wilsonii is placed in ser. Multiflori of sect. Cotoneaster with C. multiflorus, C. submultiflorus, C. hebephyllus, C. mongolicus, and C. soongoricus). Among these, only C. multiflorus and C. hebephyllus have been found to have the identical flavone O-glycosides of C. wilsonii. The finding of identical flavonoid patterns in the morphologically similar species C. multiflorus of central and northeastern China and C. wilsonii is difficult to explain as these two taxa are geographically disjunct. No change in morphology and/or flavonoids was the trend during the evolution of several woody plants on Uleung-island. There are few absolute differences between island endemics and mainland counterparts in Korea, Japan, or central China according to previous studies [Acer takesimense Nakai vs A. pseudosieboldianum (Pax) Komarov, Chang and Giannasi, 1991, Chang, 1992; Acer okmotoanum Nakai vs A. pictum Thunb. ex Murray var. mono (maxim.) maxim. ex Franch., Chang, 2001; Tilia insularis Nakai vs T. amurensis Rupr., Pigott, 2000;
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Lonicera insularis Nakai vs L. morrowii A. Gray, Hara, 1983; and Fagus multinervis Nakai vs F. engleriana Seemen ex Diels, Shen, 1992; Abelia insularis Nakai vs A. biflora Turcz., Hara, 1983; P. takesimense Nakai vs Prunus sargentii Rehder, Chang, unpublished]. Cotoneaster wilsonii appears to be another example in which no change has occurred in chemistry or morphology, except for the degree of leaf pubescence. It seems that many of the woody plants on the island are recently derived narrow endemic taxa. The extent and pattern of flavonoid variation in two closely related taxa of Cotoneaster (C. wilsonii and C. multiflorus) indicated that C. multiflorus was historically more widely and continuously distributed in eastern Asia, and that the lack of differentiation between plants on the island and the inland was a result of recent divergence. 4.2. Evolution of the flavonoids According to Harborne (1977) and Gornall and Bohm (1978), the presence of flavonols is thought to be a primitive character state, while the presence of flavonols, together with flavone O-glycosides, and O-methylated compounds is considered to represent a more advanced character state. Thus, species in section Densiflos with their many flowers arranged in a corymb and which produce only flavonols may represent the basal ancestral stock, which has given rise to the more advanced taxa in section Cotoneaster, series Multiflori, including C. wilsonii and to some members of series Integerrimi. The two series within section Cotoneaster except for C. tenuipes, C. dielsianus, and C. zabelii of series Intergerrimi do not reflect phylogenetic relationships. In summary, due to the similarity between C. wilsonii and C. multiflorus with respect of chemical and morphological data, we do not believe that further taxonomic splitting of C. multiflorus is warranted like many other endemic woody species on this island. Acknowledgements Our thanks to Dr. Qin, H. N. at PE and Mr. M. H. Chung at Chollipo Arboretum for their help and consideration. Appreciation is also expressed to W. K. Min and E. Y. Ji for their laboratory help. At lastly, I would like to dedicate this paper to my dear friend, the late Dr. Michael O. Moore at the University of Georgia, USA. This research was supported by a grant (PF001302-00) from the Plant Diversity Research Center of 21st Frontier Research Program funded by Ministry of Science and Technology of Korean Government. Appendix A. Origin and accession number for specimens utilized for flavonoids surveys. All voucher specimens are deposited at Seoul National University, The Arboretum (SNUA) or as otherwise indicated. C. acutifolius Turcz. K. Clausen et al. 79–30 (Arnold Arboretum) C. adppressus Bois K. Clausen et al. 79–100 (Arnold Arboretum) C. ambiguus Rehder. et Wilson S.
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Davis et al. 80–256 (Arnold Arboretum) C. amoenus Wilson Chang 3610 (Chollipo Arboretum) C. apiculatus Rehder et Wilson K. Clausen et al. 79–101 (Arnold Arboretum) C. dammeri Schneider cv. Coral Beauty Chang 3618 (Chollipo Arboretum) C. dielsianus Pritz. var. dielsianus K. Clausen et al. 79–89 (Arnold Arboretum) C. dielsianus Pritz. var. elegans Rehd. et Wilson [as C. sabrinus Hadden ex Kru¨ ssman] L. Segel and A. Thompson 78–49 (Arnold Arboretum) C. divaricatus Rehd. et Wils. K. Clausen et al. 79–177 (Arnold Arboretum) C. franchetii Bois A. Logan and N. Gilsdorf 404 (Arnold Arboretum) C. glabratus Rehder et Wilson Chang 3614 (Chollipo Arboretum) C. hebephyllus Diels China; Gansu, PE 0603155, Sichuan, PE 541627, PE 0549097, Yunnan, PE 05812661, PE 0570597 C. horizontalis Decne K. Clausen et al. 79-82 (Arnold Arboretum) C. lucidus Schlecht Chang 3609 (Chollipo Arboretum) C. multiflorus Bge. var. multiflorus China: Shaanxi, PE 0943369, PE 354332, PE 354327, Sichuan, PE 672388, K. Clausen et al. 79–103 (Arnold Arboretum) C. multiflorus var. calocarpus Rehder et Wilson A. Logan and N. Gilsdorf 406 (Arnold Arboretum) C. mongolicus Pojark. China: Neimonggu, PE 817345 C. nitens Rehder et Wilson Chang 3612 (Chollipo Arboretum) C. obscurus Rehder et Wilson Chang 3611 (Chollipo Arboretum), L. Segel and A. Thompson 78–67 (Arnold Arboretum) C. salicifolius Franch. var. floccosus Rehder et Wilson Chang 3616 (Chollipo Arboretum) C. salicifolius Franch. cv. Herbstfeuer Chang 3610 (Chollipo Arboretum) C. salicifolius Franch. cv. Repens Chang 3617 (Chollipo Arboretum) C. sikangensis Flinck et Hylmo Chang 3608 (Chollipo Arboretum) C. soongoricus (Rehder et Herd.) Popov China: Neimonggu, PE 728788, PE 500612, Gansu, PE 29608, PE s.n. (Huang-He-Dui (8781) C. submultiflorus Popov China: Xinjiang, PE 0594032, PE s.n. (T.N. Liou (3003), Tibet, PE 1161440, PE 0881746, PE s.n. (P. Q. Zhong (5639) C. sylvestris Pamp. [as C. racemiflorus (Desf.) K. Koch var. veitchii Rehder et Wilson] A. Logan and N. Gilsdorf 407 (Arnold Arboretum) C. tenuipes Rehder et Wilson K. Clausen et al. 79–32 (Arnold Arboretum) C. wilsonii Nakai Korea: Ulleung-do, T. Lee s.n. (July 27, 1961; July 25, 1966; Nov.1, 1970), DY Choi295, Chang 3615 (Chollipo Arboretum) C. zabelii C. K. Schneid. K. Clausen et al. 79–31 (Arnold Arboretum) References Chang, C.S., Giannasi, D.E., 1991. Foliar flavonoids of Acer sect. Palmata series Palmata. Syst. Bot. 16, 225–241. Chang, C.S., 1992. A morphometric analyses of genus Acer L., section Palmata Pax, series Palmata. Kor. J. Plant Taxonomy 21, 165–186. Chang, C.S., 2001. Reconsideration of Acer pictum complex in Korea. Kor. J. Plant Taxonomy 31, 283– 309 (in Korean). Chang, C.S., 1998. Flavonoid chemistry of Weigela (Caprifoliaceae) in Korea. J. Plant Res. 110, 275–281. Chang, C.S., 2000. Foliar flavonoids of eastern Asian birch (Betula)—with respect to Korean plants. Kor. J. Plant Taxonomy 30, 75–91 (in Korean). Giannasi, D.E., 1975. The flavonoids systematics of the genus Dahlia (Compositae). Mem. N.Y. Bot. Gard 26, 1–128. Gornall, R.J., Bohm, B.A., 1978. Angiosperm flavonoid evolution: A reappraisal. Syst. Bot 3, 353–368. Hara, H., 1983. A Revision of Caprifoliaceae of Japan with Reference to Allied Plants in Other Districts and the Adoxaceae. Gingkoana 5. Academic Scientific Book Inc., Tokyo.
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Harborne, J.B., 1977. Flavonoids and the evolution of the Angiosperms. Biochem. Syst. and Ecology 5, 7–22. Kru¨ ssman, G., 1976. Manual of Cultivated Broad-Leaved Trees and Shrubs. Timber Press, Beaverton, OR. Lee, D.Y., Joo, S.U., 1958. Reinvestigation of the Flora of the Dagelet Island. Korea Univ. Nat. Sci. Bull. 3, 223–296 (in Korean). Lee, T.B., 1980. Illustrated Flora of Korea. Hyang-mun Pub. Co., Seoul (in Korean). Markham, K.R., 1982. Techniques of Flavonoid Identification. Academic Press, New York. Markham, K.R., Wilson, R.D., 1989. Paper chromatographic mobilities of a range of flavonoid C-glycosides. Phytochemical Bulletin 21, 2–4. Mabry, T.J., Markham, B.L., Thomas, M.B., 1970. The Systematic Identification of Flavonoids. SpringerVerlag, New York, NY. Palme, E.A., Rita, R., De Feo, V., Morelli, I., 1994. Flavonoid glycosides from Cotoneaster thymaefolia. Phytochemistry 35, 1381–1382. Palme, E.A., Rita, R., Morelli, I., 1996. Flavonols and isoflavones from Cotoneaster simonsii. Phytochemistry 42, 903–905. Pigott, D., 2000. The taxonomic status of Tilia insularis. New Plantsman 7, 178–183. Pridham, J.B., 1956. Determination of sugars on paper chromatograph with p-anisidine hydrochloride. Anal. Chem. 28, 1967–1968. Rehder, A., 1940. Manual of Cultivated Trees and Shrubs, Second ed. The Macmillan Company, New York, NY. Shen, C.D., 1992. A monograph of the genus Fagus Tourn. ex L. (Fagaceae). Ph. D. Diss. City University of New York. Willis, J.C., 1985. A Dictionary of the Flowering Plants and Ferns. Cambridge University Press, Cambridge. Yang, S.Y., 1996. A Pioneer Study on Geology of Korean Peninsula. Kyungbuk National University Press, Taegu (Korean translated version). Yu¨ , T.T., 1974. Rosaceae. Flora Republicae Popularis Sinicae, 36. Science Press, Beijing (in Chinese).
Leaf flavonoids in Cotoneaster wilsonii (Rosaceae) from the island Ulleung-do, Korea Chin-Sung Chang a,∗, Jeong Ill Jeon b a
Department of Forest Resources and The Arboretum, Seoul National University, Suwon, 441-744, South Korea b Department of Urban Horticulture, Shingu College, Seongnam, 462-743, South Korea Received 12 June 2001; accepted 24 January 2002
Abstract The leaf flavonoids of Cotoneaster wilsonii, an endemic shrub to the island Ulleung-do in East Sea of Korea, were characterized and compared with flavonoids in some eastern Asian Cotoneaster species. The highly specialized taxa belonging to sect. Cotoneaster (sensu Yu¨) including C. wilsonii produced a mixture of flavone O- and C-glycoside and flavonol O-glycosides, including isorhamnetin glycosides. The morphologically similar species, C. multiflorus and C. hebephyllus of central China and C. wilsonii of Korea had similar flavonoid profiles. In addition, C. tenuipes, C. zabelii, and C. dielsianus (sect. Cotoneaster, ser. Integerrimi) had similar flavonoid patterns to taxa in the C. multiflorus complex (sect. Cotoneaster, ser. Multiflori). This indicated that the chemical data cut across Yu¨’s serial treatment within section Cotoneaster. Earlier studies showed that there were few absolute differences between many of the other woody plants growing on this island and on those on the Korean peninsula and mainland China, or the Japanese archipelago. C. wilsonii appears to be another example in which no change in chemistry or morphology has occurred. Many of the woody plants on this island are very recently derived and their progenitors were historically more widely and continuously distributed in eastern Asia. 2002 Elsevier Science Ltd. All rights reserved. Keywords: Flavonoids; Cotoneaster wilsonii; Cotoneaster multiflorus; Island Ulleung-do; Classification
∗
Tel.: +82 31 290 2322; fax: +82 31 295 6660. E-mail address: [email protected] (C.-S. Chang).
0305-1978/03/$ - see front matter 2002 Elsevier Science Ltd. All rights reserved. doi:10.1016/S0305-1978(02)00064-9
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1. Introduction The ornamentally important genus Cotoneaster of the Rosaceae is relatively small with ca. 90 species of shrubs (Yu¨ , 1974; Willis, 1985). The distribution of this genus extends from North Africa to North America, including the Himalayas, Siberia, and China (Rehder, 1940; Kru¨ ssman, 1976). The center of distribution is in China, where almost 60% of species occur, but no species occur in Japan. Cotoneaster wilsonii Nakai, which is characterized by a small number of flowers arranged in a corymb, red fruits and deciduous leaves, is endemic to the small island of Ulleung-do, Korea (Lee, 1980, Fig. 1). C. wilsonii is confined to two areas on the island and comprises fewer than 150 individuals, suggesting it may be endangered. Since this taxon has not previously been investigated, its taxonomic position is not known. The island Ulleung-do is located ca. 150 km off the coast of Korea at 37° 30⬘N and 130° 51⬘E (Lee and Joo, 1958; Pigott, 2000). This island is of late Tertiary origin from three to five million years old (Yang, 1996). Although its origin is controversial, it is known that it was connected to the Korean peninsula and the west coast of Japan until the early Quaternary (Yang, 1996). Ulleung-do has a unique flora of ca. 180 woody species of which ten are endemic to the island (Lee and Joo, 1958).
Fig. 1. Geographic distribution of Cotoneaster wilsonii Nakai in the Island Ulleung-do of Korea and its related two taxa in central China.
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The flora has evolved from isolated species, mainly associated with the Korean peninsula, the Japanese islands, and central China. The two major alternative classifications of Cotoneaster by Rehder (1940) and Yu¨ (1974) conflict unanimity in the circumscription of relationships. Thus, Redher (1940) divided the genus into two sections, Orthopetalum and Chaenopetalum, while Yu¨ (1974) divided the genus into three sections, Densiflos, Cotoneaster, and Uniflos with many series. Rehder recognized only two sections: Uniflos and Cotoneaster combined in sect. Orthopetalum, while he included some species of Yu¨ ’s sects. Densiflos, Cotoneaster, and Uniflos into sect. Chaenopetalum. Rehder’s treatment was based entirely on the position and colour of the petals and the number of styles. On the other hand, Yu¨ indicated that inflorescence type and the number of flowers might be suitable characters for the circumscription of taxa and the determination of natural sectional groupings. According to Yu¨ , sects. Densiflos and Cotoneaster can be separated from sect. Uniflos on the basis of the former having many flowers arranged in a corymb and large leaves. Furthermore, sect. Densiflos differed from sect. Cotoneaster in having many flowers (more than 20) with a compound corymb. Based on the gross morphology, C. wilsonii appears to belong to sect. Cotoneaster sensu Yu¨ . Yu¨ (1974) considered 29 taxa members of sect. Cotoneaster and further divided this section into three series: Multiflori, Acuminati, and Integerrimi, with respect to flower colour, leaf pubescence, fruit colour and the number of seeds. However, this treatment was somewhat artificial because of occurrence of many intermediate taxa among three series. A preliminary study of Cotoneaster taxa in eastern Asia showed that flavonols were the main leaf flavonoid constituents, and flavones and others from C. wilsonii were found unlikely. The purpose of the present study was to 1) identify the leaf flavonoids of C. wilsonii, 2) determine the flavonoid of components in some of the closest postulated mainland relatives, and 3) consider possible evolutionary implications of the flavonoid data.
2. Materials and methods 2.1. Plant materials Leaf material of C. wilsonii and 29 taxa of Asian Cotoneaster were surveyed using herbarium specimens at The Arboretum of Seoul National University (SNUA) and Institute of Botany, Academia Sinica, Beijing (PE) and fresh samples from the Chollipo Arboretum and the Island Ulleung-do. The collection numbers and places of deposit are given in Appendix A. 2.2. Phytochemical analysis The extraction of flavonoids from leaves followed the methods of Mabry et al. (1970); Chang and Giannasi (1991), and Giannasi (1975). The flavonoids survey
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employed two-dimensional paper chromatography using Whatmann 3 MM paper. TBA (tertiary-butanol: acetic acid : water, 3:1:1) was used as the first solvent and 15% HOAc (acetic acid:water, 15:85) as the second solvent. Flavonoid profiles were viewed under UV light and colours were recorded before and after fuming with ammonia vapour. Compounds were identified using standard UV-visible spectroscopy (Giannasi, 1975; Mabry et al., 1970; Markham, 1982). Aglycones obtained from acid hydrolysis of the glycosides were identified in the same manner to determine the location of sugar(s). Several compounds were compared directly with compounds identified from previous studies (Chang and Giannasi, 1991; Chang, 1998; Chang, 2000). Spectral and Rf data were compared with published data (Mabry et al., 1970). Flavonoid glycoside sugars were identified using trifluoroacetic acid hydrolysis, and co-chromatography with standard sugars in one-dimension, using ethyl acetate: pyridine: water (5:1.6:1) as the solvent. The sugars were visualized by spraying with p-anisidine hydrochloride (Pridham, 1956) followed by heating (100 °C, 10 min). 3. Results The leaf flavonoids identified in the selected taxa are tabulated in Table 1. They included five flavonol glycosides: quercetin-3-O-glucoside, quercetin-3-O-rhamnoside, quercetin-3-O-rhamnosylglucoside, isorhamnetin-3-O-galctodide, and isorhamnetin-3-O-rhamnose-glucose (unknown order); two flavone glycosides: apigenin-7O-glucoside and luteolin-7-O-glucoside, and three flavone C-glycosides: vicenin-A and -B (Markham and Wilson, 1989), and luteolin-8-C-glycoside. Flavonols, primarily as the 3-O-glycosides of quercetin but with occasional isorhamnetin 3-O-glycosides, were detected in all taxa of sections Densiflos and Uniflos and one series (Acuminati) of section Cotoneaster. Flavonoid analysis of C. wilsonii yielded a series of flavone O- and C-glycosides, and a methylated flavonol. Especially noteworthy was the occurrence of flavone O-glycosides, which were characteristic constituents of C. mutiflorus Bge, C. hebephyllus Diels (series Multiflori) and C. tenuipes Rehder et Wilson, C. dielsianus Pritz. and C. zabelii Schneider (ser. Integerrimi of sect. Cotoneaster). The presence of isorhamnetin 3O-rhamnose-glucose in C. wilsonii and some other species e.g., C. submultiflorus Popov, C. mongolicus Pojark., C. mutiflorus, and C. hebephyllus of ser. Multiflori of sect. Cotoneaster and in two taxa of series Integerrimi, C. tenuipes and C. dielsianus, may also have some taxonomic significance. Tabulation of the identified flavonoid (Table 1) failed to distinguish clearly the three sections recognised by Yu¨ (1974), although species in sect. Cotoneaster, ser. Multiflori, except for C. sylvestris, do share the same flavonoid profile. However, four taxa in series Integerrimi and one from section Uniflos (Yu¨ , 1974) also have very similar patterns. Thus, the flavonoid data tend to cut across the serial treatment within section Cotoneaster. In several earlier investigations on flavonoids of European taxa of Cotoneaster (Palme et al., 1994, 1996) the C-glycosylflavones (vitexin, vitexin-2⬙-O-rhamnoside,
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Table 1 Flavonoids distribution of Cotoneaster taxa considered in eastern Asiaa Taxonc
Flavonol O-glycoside 1
sect. Densiflos ser. Salififolii C. salicifolius var. floccosus C. salicifolius cv. Herbstfeuer C. salicifolius cv. Repens ser. Frigidi C. glabratus sect. Cotoneaster ser. Multiflori C. multiflorus var. multiflorus C. multiflorus var. calocarpus C. wilsonii C. mongolicus C. hebephyllus C. soongoricus C. submultiflorus C. sylvestris ser. Acuminati C. acutifolius ser. Integerrimi C. ambiguus C. lucidus (=C. ambiguus?) C. zabelii C. tenuipes C. dielsianus var. dielsianus C. dielsianus var. elegans C. franchetii C. amoenus (=C. franchetii?) C. obscurus C. sikangensis (=C. obscurus?) sect. Uniflos ser. Microphylli C. dammeri cv. Coral Beauty ser. Distichi C. apiculatus C. adppressus C. divaricatus C. horizontalis C. nitens
2
3
Flavone C-glycosylflavone
O-glycoside
6
7
8
9
10
X X X (X) X X X
X
X
X
X
X
X
X X
X X X
X
X X X X
X
X
X
X
O-Meglycoside 4 5
X X X
X X
(X)?b X X
X
X
X
X X X X X X X X
X X X X (X) X X
X X X X X X X
X
X
(X)
(X)
X X X X X
X (X) (X)
X
(X)
X
X
X
X X X X X X X X X
X X X X X X X X X X
X X
X X X X X X X X X X
X
X
X
X X
X
X X X X X
X X X X X
X X X X
X
X X
X X
a 1: quercetin 3-O-glucoside, 2: quercetin 3-O-rhamnoside, 3: quercetin 3-O-rhamnoglucoside, 4:isorhamnetin 3-O-galactoside, 5: isorhamnetin 3-O-rhamnose-glucose (random order), 6:vicenin A, 7: vicenin B, 8: luteolin 6-C-glycoside, 9: apigenin 7-O-glucoside, 10: luteolin 7-O-glucoside. b (x)?=compounds whose presence/absence is not clearly established from examination of the chromatograms and (x)=compounds detected in less than 50% of surveyed individuals. c classification sensu Yu¨ (1974).
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vitexin-2⬙-O-arabinoside), flavonols (rutin, quercetin-3-O-rhamnoside, kaempferol-7methyl ether 4⬘-O-glucoside) and 5,7,2⬘,5⬘-tetrahydroxyflavanone and its 7-O-glucoside were found in C. thymaefolia Hort. and flavonols and isoflavones (genistein-5methyl ether 4’-O-glucoside, biochanin and biochanin-7-O-glucoside) were observed in C. simonsii Baker. No C-glycosylflavone, flavanones or isoflavones and only flavonols, however, seemed to occur in Asian taxa. The current flavonoid survey provides some evidence to link ser. Multiflori and ser. Integerrimi. This study showed that the flavonoids’ profiles of C. wilsonii are almost identical to those of C. multiflorus.
4. Discussion 4.1. Systematic implications and biogeography The distribution of flavonoids compounds does not correlate well with the general grouping of sect. Cotoneaster into three series proposed by Yu¨ (1974). Some species of both ser. Multiflori and ser. Integerrimi, which are characterized by having one or two seeds per fruit, appear in each chemical group. With the exception of C. multiflorus and the C. dielsianus complex, no distinct species specific pattern was chemically observed in the studied taxa in the three different series (Multiflori, Acuminati, Integerrimi) of sect. Cotoneaster or in the other two sections. The division of section Cotoneaster into three series proposed on morphology received weak support from the comparative flavonoid data. Although intraspecific variation and the general lack of flavonoid diversity within Cotoneaster as a whole somewhat limit the taxonomic value of flavonoids at the specific level, certain observations are warranted. It appears that C. multiflorus and the C. dielsianus complex, placed in separate series in Yu¨ ’s classification, are actually more closely related than this classification implies. Our results suggest that these taxa may best be included within the same group. However, pending a complete survey of all Chinese taxa, no formal nomenclatural changes are proposed here. The taxonomic significance of the flavonoid data in C. wilsonii lies in their correlation with morphological evidence. C. wilsonii is placed in ser. Multiflori of sect. Cotoneaster with C. multiflorus, C. submultiflorus, C. hebephyllus, C. mongolicus, and C. soongoricus). Among these, only C. multiflorus and C. hebephyllus have been found to have the identical flavone O-glycosides of C. wilsonii. The finding of identical flavonoid patterns in the morphologically similar species C. multiflorus of central and northeastern China and C. wilsonii is difficult to explain as these two taxa are geographically disjunct. No change in morphology and/or flavonoids was the trend during the evolution of several woody plants on Uleung-island. There are few absolute differences between island endemics and mainland counterparts in Korea, Japan, or central China according to previous studies [Acer takesimense Nakai vs A. pseudosieboldianum (Pax) Komarov, Chang and Giannasi, 1991, Chang, 1992; Acer okmotoanum Nakai vs A. pictum Thunb. ex Murray var. mono (maxim.) maxim. ex Franch., Chang, 2001; Tilia insularis Nakai vs T. amurensis Rupr., Pigott, 2000;
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Lonicera insularis Nakai vs L. morrowii A. Gray, Hara, 1983; and Fagus multinervis Nakai vs F. engleriana Seemen ex Diels, Shen, 1992; Abelia insularis Nakai vs A. biflora Turcz., Hara, 1983; P. takesimense Nakai vs Prunus sargentii Rehder, Chang, unpublished]. Cotoneaster wilsonii appears to be another example in which no change has occurred in chemistry or morphology, except for the degree of leaf pubescence. It seems that many of the woody plants on the island are recently derived narrow endemic taxa. The extent and pattern of flavonoid variation in two closely related taxa of Cotoneaster (C. wilsonii and C. multiflorus) indicated that C. multiflorus was historically more widely and continuously distributed in eastern Asia, and that the lack of differentiation between plants on the island and the inland was a result of recent divergence. 4.2. Evolution of the flavonoids According to Harborne (1977) and Gornall and Bohm (1978), the presence of flavonols is thought to be a primitive character state, while the presence of flavonols, together with flavone O-glycosides, and O-methylated compounds is considered to represent a more advanced character state. Thus, species in section Densiflos with their many flowers arranged in a corymb and which produce only flavonols may represent the basal ancestral stock, which has given rise to the more advanced taxa in section Cotoneaster, series Multiflori, including C. wilsonii and to some members of series Integerrimi. The two series within section Cotoneaster except for C. tenuipes, C. dielsianus, and C. zabelii of series Intergerrimi do not reflect phylogenetic relationships. In summary, due to the similarity between C. wilsonii and C. multiflorus with respect of chemical and morphological data, we do not believe that further taxonomic splitting of C. multiflorus is warranted like many other endemic woody species on this island. Acknowledgements Our thanks to Dr. Qin, H. N. at PE and Mr. M. H. Chung at Chollipo Arboretum for their help and consideration. Appreciation is also expressed to W. K. Min and E. Y. Ji for their laboratory help. At lastly, I would like to dedicate this paper to my dear friend, the late Dr. Michael O. Moore at the University of Georgia, USA. This research was supported by a grant (PF001302-00) from the Plant Diversity Research Center of 21st Frontier Research Program funded by Ministry of Science and Technology of Korean Government. Appendix A. Origin and accession number for specimens utilized for flavonoids surveys. All voucher specimens are deposited at Seoul National University, The Arboretum (SNUA) or as otherwise indicated. C. acutifolius Turcz. K. Clausen et al. 79–30 (Arnold Arboretum) C. adppressus Bois K. Clausen et al. 79–100 (Arnold Arboretum) C. ambiguus Rehder. et Wilson S.
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Davis et al. 80–256 (Arnold Arboretum) C. amoenus Wilson Chang 3610 (Chollipo Arboretum) C. apiculatus Rehder et Wilson K. Clausen et al. 79–101 (Arnold Arboretum) C. dammeri Schneider cv. Coral Beauty Chang 3618 (Chollipo Arboretum) C. dielsianus Pritz. var. dielsianus K. Clausen et al. 79–89 (Arnold Arboretum) C. dielsianus Pritz. var. elegans Rehd. et Wilson [as C. sabrinus Hadden ex Kru¨ ssman] L. Segel and A. Thompson 78–49 (Arnold Arboretum) C. divaricatus Rehd. et Wils. K. Clausen et al. 79–177 (Arnold Arboretum) C. franchetii Bois A. Logan and N. Gilsdorf 404 (Arnold Arboretum) C. glabratus Rehder et Wilson Chang 3614 (Chollipo Arboretum) C. hebephyllus Diels China; Gansu, PE 0603155, Sichuan, PE 541627, PE 0549097, Yunnan, PE 05812661, PE 0570597 C. horizontalis Decne K. Clausen et al. 79-82 (Arnold Arboretum) C. lucidus Schlecht Chang 3609 (Chollipo Arboretum) C. multiflorus Bge. var. multiflorus China: Shaanxi, PE 0943369, PE 354332, PE 354327, Sichuan, PE 672388, K. Clausen et al. 79–103 (Arnold Arboretum) C. multiflorus var. calocarpus Rehder et Wilson A. Logan and N. Gilsdorf 406 (Arnold Arboretum) C. mongolicus Pojark. China: Neimonggu, PE 817345 C. nitens Rehder et Wilson Chang 3612 (Chollipo Arboretum) C. obscurus Rehder et Wilson Chang 3611 (Chollipo Arboretum), L. Segel and A. Thompson 78–67 (Arnold Arboretum) C. salicifolius Franch. var. floccosus Rehder et Wilson Chang 3616 (Chollipo Arboretum) C. salicifolius Franch. cv. Herbstfeuer Chang 3610 (Chollipo Arboretum) C. salicifolius Franch. cv. Repens Chang 3617 (Chollipo Arboretum) C. sikangensis Flinck et Hylmo Chang 3608 (Chollipo Arboretum) C. soongoricus (Rehder et Herd.) Popov China: Neimonggu, PE 728788, PE 500612, Gansu, PE 29608, PE s.n. (Huang-He-Dui (8781) C. submultiflorus Popov China: Xinjiang, PE 0594032, PE s.n. (T.N. Liou (3003), Tibet, PE 1161440, PE 0881746, PE s.n. (P. Q. Zhong (5639) C. sylvestris Pamp. [as C. racemiflorus (Desf.) K. Koch var. veitchii Rehder et Wilson] A. Logan and N. Gilsdorf 407 (Arnold Arboretum) C. tenuipes Rehder et Wilson K. Clausen et al. 79–32 (Arnold Arboretum) C. wilsonii Nakai Korea: Ulleung-do, T. Lee s.n. (July 27, 1961; July 25, 1966; Nov.1, 1970), DY Choi295, Chang 3615 (Chollipo Arboretum) C. zabelii C. K. Schneid. K. Clausen et al. 79–31 (Arnold Arboretum) References Chang, C.S., Giannasi, D.E., 1991. Foliar flavonoids of Acer sect. Palmata series Palmata. Syst. Bot. 16, 225–241. Chang, C.S., 1992. A morphometric analyses of genus Acer L., section Palmata Pax, series Palmata. Kor. J. Plant Taxonomy 21, 165–186. Chang, C.S., 2001. Reconsideration of Acer pictum complex in Korea. Kor. J. Plant Taxonomy 31, 283– 309 (in Korean). Chang, C.S., 1998. Flavonoid chemistry of Weigela (Caprifoliaceae) in Korea. J. Plant Res. 110, 275–281. Chang, C.S., 2000. Foliar flavonoids of eastern Asian birch (Betula)—with respect to Korean plants. Kor. J. Plant Taxonomy 30, 75–91 (in Korean). Giannasi, D.E., 1975. The flavonoids systematics of the genus Dahlia (Compositae). Mem. N.Y. Bot. Gard 26, 1–128. Gornall, R.J., Bohm, B.A., 1978. Angiosperm flavonoid evolution: A reappraisal. Syst. Bot 3, 353–368. Hara, H., 1983. A Revision of Caprifoliaceae of Japan with Reference to Allied Plants in Other Districts and the Adoxaceae. Gingkoana 5. Academic Scientific Book Inc., Tokyo.
C.-S. Chang, J.I. Jeon / Biochemical Systematics and Ecology 31 (2003) 171–179
179
Harborne, J.B., 1977. Flavonoids and the evolution of the Angiosperms. Biochem. Syst. and Ecology 5, 7–22. Kru¨ ssman, G., 1976. Manual of Cultivated Broad-Leaved Trees and Shrubs. Timber Press, Beaverton, OR. Lee, D.Y., Joo, S.U., 1958. Reinvestigation of the Flora of the Dagelet Island. Korea Univ. Nat. Sci. Bull. 3, 223–296 (in Korean). Lee, T.B., 1980. Illustrated Flora of Korea. Hyang-mun Pub. Co., Seoul (in Korean). Markham, K.R., 1982. Techniques of Flavonoid Identification. Academic Press, New York. Markham, K.R., Wilson, R.D., 1989. Paper chromatographic mobilities of a range of flavonoid C-glycosides. Phytochemical Bulletin 21, 2–4. Mabry, T.J., Markham, B.L., Thomas, M.B., 1970. The Systematic Identification of Flavonoids. SpringerVerlag, New York, NY. Palme, E.A., Rita, R., De Feo, V., Morelli, I., 1994. Flavonoid glycosides from Cotoneaster thymaefolia. Phytochemistry 35, 1381–1382. Palme, E.A., Rita, R., Morelli, I., 1996. Flavonols and isoflavones from Cotoneaster simonsii. Phytochemistry 42, 903–905. Pigott, D., 2000. The taxonomic status of Tilia insularis. New Plantsman 7, 178–183. Pridham, J.B., 1956. Determination of sugars on paper chromatograph with p-anisidine hydrochloride. Anal. Chem. 28, 1967–1968. Rehder, A., 1940. Manual of Cultivated Trees and Shrubs, Second ed. The Macmillan Company, New York, NY. Shen, C.D., 1992. A monograph of the genus Fagus Tourn. ex L. (Fagaceae). Ph. D. Diss. City University of New York. Willis, J.C., 1985. A Dictionary of the Flowering Plants and Ferns. Cambridge University Press, Cambridge. Yang, S.Y., 1996. A Pioneer Study on Geology of Korean Peninsula. Kyungbuk National University Press, Taegu (Korean translated version). Yu¨ , T.T., 1974. Rosaceae. Flora Republicae Popularis Sinicae, 36. Science Press, Beijing (in Chinese).