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Lack of altitudinal trends in phytochemical constituents of Swertia franchetiana (Gentianaceae)
Biochemical Systematics and Ecology 32 (2004) 861–866 www.elsevier.com/locate/biochemsyseco
Lack of altitudinal trends in phytochemical constituents of Swertia franchetiana (Gentianaceae) Huiling Yang, Yuanwen Duan, Fengzhu Hu, Jianquan Liu Qinghai-Tibet Plateau Biological Evolution and Adaptation Laboratory, Northwest Plateau Institute of Biology, Chinese Academy of Sciences, Xining 810001, Qinghai, China Received 9 June 2003; accepted 20 February 2004
Abstract Diverse biological characters commonly vary with altitude in species that have a wide altitudinal distribution, partly at least as a result of adaptation to differences in aridity, but whether such variation exists for phytochemical constituents remains unknown. Therefore, levels of seven important phytochemical constituents of Swertia franchetiana (swertiamarin, oleanolic acid, swertisin, mangiferin, 1,5,8-trihydroxy-3-methoxyxanthone, 1,8-dihydroxy3,7-dimethoxyxanthone and 1,8-dihydroxy-3,5-dimethoxyxanthone) were studied and statistically compared, using materials collected from sites ranging from 2200 to 3960 m in altitude. Swertiamarin was the most abundant in all samples, then mangiferin, oleanolic acid and the other three xanthones. Throughout the distributional range of this species, no altitudinal trend was detected for other constituents except 1,8-dihydroxy-3,7-dimethoxyxanthone, which showed a negative correlation with altitude. However, the concentration of 1,8-dihydroxy-3,7-dimethoxyxanthone and mangiferin showed a significantly latitudinal and longitudinal correlation. # 2004 Elsevier Ltd. All rights reserved. Keywords: Altitude; Swertia franchetiana; Active constituents; Variation
1. Introduction Swertia franchetiana H. Smith. is a biennial herb of the family Gentianaceae. An extremely important and well-known folk medicine is prepared from this species,
Corresponding author. Tel.: +86-971-6153387; fax: +86-971-6143282. E-mail address: [email protected] (J. Liu).
0305-1978/$ - see front matter # 2004 Elsevier Ltd. All rights reserved. doi:10.1016/j.bse.2004.02.008
862
H. Yang et al. / Biochemical Systematics and Ecology 32 (2004) 861–866
known as ‘‘DiDa’’ in Tibet, that is used to cure a range of diseases, including gall and liver disorders (Yang, 1991). Recent investigations on the action of this drug have revealed that its major active phytochemical constituents consist of swertiamarin, oleanolic acid, mangiferin, swertisin, and other xanthones (Ding et al., 1982, 1988). These active constituents have been found separately or collectively to have hepatoprotective (Liu et al., 1993; Komatsu et al., 1997), hypoglycemic (Song, 1986), anti-inflammatory (Banerjee et al., 2000), anti-oxidative (Ashida et al., 1994; Born et al., 1996), anti-tubercular (Ghosal and Chaudhuri, 1975; Bian et al., 1998), and anti-fungal (Rodriguez et al., 1995) activities, together with other pharmacological properties (Rafatullah et al., 1993; Singha et al., 1993; Ji, 1995). Although endemic to the Tibetan Plateau, this species is widely distributed, from the eastern parts of the Plateau in Qinghai province, to western parts, in Tibet, at altitudes ranging from 2200 to 3800 m (Ho, 1998). Many widely distributed species show physiological trends along altitudinal gradients (Ko¨ner, 1999). For example, contents of carotenoids, flavonoids, sucrose, fructose, glucose and total soluble sugar (Han et al., 1998), the phyotosynthetic rate, and both the light compensation and saturation points tend to rise, while the photorespiration rate tends to decline as altitude increases (Woodward, 1986). However, differences in other characters, such as anatomical structures, are not usually pronounced (Liu and Noshiro, 2003). Geographical variations in levels of phytochemicals can occur in some species (e.g. volatile terpenoids in Juniperus; Adams, 1983, 1986). The purpose of this article is to statistically compare the concentration of seven active constituents in S. franchetiana using materials collected from different altitudes to determine whether the accumulation of these compounds is affected by altitude.
2. Materials and methods 2.1. Materials The plant materials were collected from seven populations growing between 2200 and 3960 m (Table 1), covering the entire altitudinal range of this species. All materials were collected at the flowering stage in order to avoid differences associaTable 1 The location and altitude of Swertia franchetiana populations sampled in this study Population Bayi Nanshan Huangzhong Linzhi Jiacha Linzhi Changdu
Location Bayi, Xining, Qinghai Nanshan, Xining, Qinghai Huangzhong, Qinghai Linzhi, Tibet Jiacha, Tibet Linzhi, Tibet Changdu, Tibet
Altitude (m) 2200 2300 2620 2950 3330 3460 3960
Longitude v
Latitude 0
101 52.304 v 101 47.9260 v 101 32.9700 v 94 22.6310 v 92 39.2430 v 94 43.6550 v 97 17.160
v
36 v 36 v 36 v 29 v 29 v 29 v 30
33.6290 35.0010 47.6800 20.3990 06.9950 41.9790 41.380
H. Yang et al. / Biochemical Systematics and Ecology 32 (2004) 861–866
863
ted with different growth stages of the plant. In each population, we randomly chose 30 individuals that were spaced more than 1 m apart. 2.2. Analytical methods All analyses were repeated at least four times, using the methods suggested by Demizu et al. (1986) and Menkovic´ et al. (2000a,b). The air-dried individuals were cut, crushed and mixed completely. Methanol (70%) was then used to extract swertiamarin, mangiferin and swertisin; and 100% ethanol to extract oleanolic acid and the three xanthones. All extracts were analyzed using an HPLC chromatograph (Waters 600E) equipped with an ultraviolet detector (Waters 486) and a Phenomenex Kromasil C18, 5 lm, 250 4:60 mm column. The HPLC protocol developed for analyzing the bitter-tasting swertiamarin, flavonoids (swertisin) and xanthones by Demizu et al. (1986) was adopted, using methanol:water as the mobile phase in the proportions 32:68 for swertiamarin and mangiferin, 45:55 for swertisin, 96:4 for oleanolic acid, and 78:22 for the three xanthones. To calculate the contents of the seven compounds, the previously separated standards were weighed and dissolved in 1 ml of methanol or ethanol. This was diluted to give a series of concentrations (0.1, 0.2, 0.3, 0.4 and 0.5 mg/ml). Three injections were performed for each dilution. Concentrations were determined by injecting 10 ll of the standard solutions or sample extracts at a flow rate of 1 ml/min at room temperature. The peak wavelengths were 227 nm for swertiamarin, 254 nm for mangiferin, swertisin and the three xanthones, and 220 nm for oleanolic acid. A standard calibration curve was generated for each compound using the linear leastsquares regression equation derived from the peak area. 2.3. Statistical analyses All statistical analyses were performed using Microsoft Excel 2000 or the SPSS 8.0 for Windows software package. The mean values obtained for the different groups were compared by one-way ANOVA, post hoc-LSD and t-tests. Differences between means were assumed to be statistically significant at probability levels <0.05. Simple linear correlation analysis was used to obtain a measure of the correlation and the strength of the relationships between the variables.
3. Results and discussion Fig. 1 shows variations in the levels of the seven major active constituents (1–7) of S. franchetiana along the altitudinal gradient in all sampled localities. No distinct trend was revealed for the other constituents except 6, which showed a negative correlation with altitude. This finding is further confirmed by the matrix correlation of all analyzed parameters (Table 2). A significant correlation between the concentration of 6 and altitude was observed, and a significant negative correlation between the levels of 1 and 7.
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H. Yang et al. / Biochemical Systematics and Ecology 32 (2004) 861–866
Fig. 1. Variations in mean content of the seven active constituents: (1) swertiamarin, (2) oleanolic acid, (3) swertisin, (4) mangiferin, (5) 1,5,8-trihydroxy-3-methoxyxanthone, (6) 1,8-dihydroxy-3,7-dimethoxyxanthone and (7) 1,8-dihydroxy-3,5-dimethoxyxanthone along the altitudinal gradient. Values are means. SEM values (not shown for clarity) are 10% of the means, n 4.
Despite the general absence of altitude-associated trends in phytochemical constituents of S. franchetiana, there was considerable variation in the content of certain compounds among some populations, which was statistically significant (p < 0:001). For example, the contents of 1 showed significant differences between all of the populations (p < 0:001) except for Bayi (2200 m), Nanshan (2300 m) and Jiacha (3330 m) (p > 0:05). The content of 2 did not differ significantly between Bayi (2200 m) and Nanshan (2300 m) (p > 0:05), but it varied significantly among the other populations (p < 0:001). The contents of both 4 and 6 were not significantly different in the Jiacha (3330 m) and Changdu (3960 m) populations (p > 0:05), but differed significantly among the other populations (p < 0:001). Matrix correlation analyses confirmed significant correlations between concentration of these two compounds and latitude and longitude (Table 2). It seems Table 2 Matrix correlation of altitude, latitude, longitude and analyzed constituents: (1) swertiamarin, (2) oleanolic acid, (3) swertisin, (4) mangiferin, (5) 1,5,8-trihydroxy-3-methoxyxanthone, (6) 1,8-dihydroxy-3,7dimethoxyxanthone and (7) 1,8-dihydroxy-3,5-dimethoxyxanthone 1 Altitude Latitude Longitude 1 2 3 4 5 6
0.021 0.381 0.357
2 0.620 0.224 0.425 0.369
3 0.436 0.560 0.437 0.647 0.439
4 0.047 0.896 0.901 0.236 0.366 0.374
5 0.382 0.565 0.669 0.278 0.522 0.168 0.546
6
7
0.849 0.911 0.873 0.264 0.154 0.631 0.709 0.345
significant at the 0.05 probability level, significant at the 0.001 probability level.
0.012 0.227 0.387 0.763 0.005 0.699 0.289 0.039 0.497
H. Yang et al. / Biochemical Systematics and Ecology 32 (2004) 861–866
865
likely that the differences in phytochemical constituents of S. franchetiana are more geographical than altitudinal. Two possible factors might account for these variations. First, ecological factors other than altitude, e.g. soil type, temperature, and precipitation, might affect the synthesis and turnover of secondary compounds (Nobel, 1991). Second, their ability to synthesize the seven selected compounds in these populations might be subject to genetic controls. Genetic differentiation generally has stronger effects on the contents of secondary compounds than ecological factors (Hashimoto and Yamada, 1994) and the mutation of a single gene might affect the production of certain compounds (Vogel et al., 1996). More studies are needed to clarify the relationships between synthesis of these compounds, genetic controls and ecological factors.
Acknowledgements We thank Miss Song Yali for her help with the HPLC experiments. We are also indebted to Dr John Blackwell and Prof. Robert Adams for polishing and improving the manuscript. Support for this research was provided by the Chinese Academy of Sciences (Key Innovation Plan KSCX-SW-106, Special Fund of Outstanding PhD Dissertation and Training Qualified People Plan ‘‘Hope of the West China’’) and the Chinese Research Project for Students Returned from Overseas fund.
References Adams, R.P., 1983. Infraspecific terpenoid variation in Juniperus scopulorum: evidence for Pleistocene refugia and recolonization in western North America. Taxon 32, 30–46. Adams, R.P., 1986. Geographic variation in Juniperus silicola and J. virginiana of the southeastern United States: multivariate analyses of morphology and terpenoids. Taxon 35, 61–76. Ashida, S., Noguchi, S.F., Suzuki, T., 1994. Antioxidative components, xanthone derivatives, in Swertia japonica Makino. J. Am. Oil Chem. Soc. 71 (10), 1095–1099. Banerjee, S., Sur, T., Kumar, M.S., Chandra, D.P., Sikdar, S., 2000. Assessment of the anti-inflammatory effects of Swertia chirata in acute and chronic experimental models in male albino rats. Indian J. Pharm. 32, 21–24. Bian, Q.Y., Luo, N.C., Xiao, P.G., 1998. Four xanthone glycosides from Swertia calycina Franch. Pharm. Pharm. Com. 4 (12), 597–598. Born, M., Carrupt, P.A., Zini, R., Bree, F., Tillement, J.P., Hostettmann, K., Testa, B., 1996. Electrochemical behavior and antioxidant activity of some natural polyphenols. Helv. Chim. Acta 79 (4), 1147–1158. Demizu, S., Ohshima, Y., Hiraga, Y., Takahashi, K., 1986. High-performance liquid chromatographic analysis of Swertia herb. J. Chromatogr. 360, 307–311. Ding, J.Y., Fan, S.F., Hu, B.L., Sun, H.F., 1988. On the xanthone glycosides and flavonoid glucoside from Swertia franchetiana H. Smith. Acta Bot. Sin. 30, 414–419, (in Chinese). Ding, J.Y., Fan, S.F., Hu, B.L., Sun, H.F., 1982. Separation and identification of xanthone glycosides in Swertia franchetiana H. Smith. Acta Biol. Plateau Sin. 1, 267–269, (in Chinese). Ghosal, S., Chaudhuri, R.K., 1975. Chemical constituents of Gentianaceae. XVI. Antitubercular activity of xanthones of Canscora decussata. J. Pharm. Sci. 64, 888–889.
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H. Yang et al. / Biochemical Systematics and Ecology 32 (2004) 861–866
Han, F., Ben, G.Y., Shi, S.B., 1998. A comparative study on the resistance of Kobresia humilis grown at different altitudes in Qinghai–Tibet Plateau. Acta Ecol. Sin. 18, 654–659, (in Chinese). Hashimoto, T., Yamada, Y., 1994. Alkaloid biogenesis: molecular aspects. Annu. Rev. Plant Physiol. Plant Mol. Biol. 45, 257–285. Ho, T.N., 1988. Flora Reipublicae Popularis Sinicae. Tomus 62. Science Press, Beijing, (in Chinese). Ji, Y.B., 1995. Pharmacology and Application of the Active Constituents in Chinese Traditional Medicines. Heilongjiang Science and Technology Press, Heilongjiang, pp. 453, (in Chinese). Komatsu, K., Purusotam, B., Yamaji, S., Kadota, S., Namba, T., 1997. A comparative study on Swertia herba from Japan, Nepal, and China and their hypoglycemic activities in streptozotocin (STZ)induced diabetic rats. Nat. Med. 51, 265–268. Ko¨ner, C., 1999. Alpine Plant Life—Functional Plant Ecology of High Mountain Ecosystems. Springer, Berlin, pp. 259–290. Liu, J.Q., Noshiro, S., 2003. Lack of latitudinal trends in wood anatomy of Dodoneaea viscosa (Sapindaceae), a species with a worldwide distribution. Am. J. Bot. 90, 532–539. Liu, J.Q., Lu, Y.P., Madhu, C., Klaassen, C., 1993. Protective effects of oleanolic acid on acetaminophen-induced hepatotoxicity in mice. J. Pharm. Exp. Ther. 266, 1607–1613. Menkovic´, N., Sˇavikin-Fodulovic´, K., Savin, K., 2000a. Chemical composition and seasonal variations in the amount of secondary compounds in Gentian lutea leaves and flowers. Plant. Med. 66, 178–180. Menkovic´, N., Sˇavikin-Fodulovic´, K., Momcilovic´, I., Grubisˇic´, D., 2000b. Quantitative determination of secoiridoid and c-pyrone compounds in Gentiana lutea cultured in vitro. Plant. Med. 66, 96–98. Nobel, P.S., 1991. Physicochemical and Environmental Plant Physiology. Academic Press, San Diego, CA. Rafatullah, S., Tariq, M., Mossa, J.S., Al-Yahya, M.A., Al-Said, M.S., Ageel, A.M., 1993. Protective effect of Swertia chirata against indomethacin and other ulcerogenic agent-induced gastric ulcers. Drugs Exp. Clin. Res. 19 (2), 69–73. Rodriguez, S., Wolfender, J.-L., Hakizamungu, E., Hostettmann, K., 1995. An antifungal naphthoquinone, Xanthones and secoiridoids from Swertia calycina. Planta Med. 61 (4), 362–364. Singha, U.K., Guru, P.Y., Sen, A.B., Tandon, J.S., 1993. Antileishmanial activity of traditional plants against Leishmania donovani in golden hamsters. Int. J. Pharm. 30, 289–295. Song, W.Z., 1986. A general survey on medicinal plants of Gentianaceae in China. Bull. Chin. Materia Med. 11, 643–647, (in Chinese). Woodward, F.I., 1986. Ecophysiological studies on the shrub Baccinium myrtillus L. taken from a wide altitudinal range. Oecologia 70, 580–586. Vogel, B.S., Wildung, M.R., Vogel, G., Croteau, R., 1996. Abietadiene Synthase from grand fir (Abies grandis). J. Biol. Chem. 38, 23262–23268. Yang, Y.C., 1991. Tibetan Medicines. Qinghai People Press, Qinghai, (in Chinese).
Lack of altitudinal trends in phytochemical constituents of Swertia franchetiana (Gentianaceae) Huiling Yang, Yuanwen Duan, Fengzhu Hu, Jianquan Liu Qinghai-Tibet Plateau Biological Evolution and Adaptation Laboratory, Northwest Plateau Institute of Biology, Chinese Academy of Sciences, Xining 810001, Qinghai, China Received 9 June 2003; accepted 20 February 2004
Abstract Diverse biological characters commonly vary with altitude in species that have a wide altitudinal distribution, partly at least as a result of adaptation to differences in aridity, but whether such variation exists for phytochemical constituents remains unknown. Therefore, levels of seven important phytochemical constituents of Swertia franchetiana (swertiamarin, oleanolic acid, swertisin, mangiferin, 1,5,8-trihydroxy-3-methoxyxanthone, 1,8-dihydroxy3,7-dimethoxyxanthone and 1,8-dihydroxy-3,5-dimethoxyxanthone) were studied and statistically compared, using materials collected from sites ranging from 2200 to 3960 m in altitude. Swertiamarin was the most abundant in all samples, then mangiferin, oleanolic acid and the other three xanthones. Throughout the distributional range of this species, no altitudinal trend was detected for other constituents except 1,8-dihydroxy-3,7-dimethoxyxanthone, which showed a negative correlation with altitude. However, the concentration of 1,8-dihydroxy-3,7-dimethoxyxanthone and mangiferin showed a significantly latitudinal and longitudinal correlation. # 2004 Elsevier Ltd. All rights reserved. Keywords: Altitude; Swertia franchetiana; Active constituents; Variation
1. Introduction Swertia franchetiana H. Smith. is a biennial herb of the family Gentianaceae. An extremely important and well-known folk medicine is prepared from this species,
Corresponding author. Tel.: +86-971-6153387; fax: +86-971-6143282. E-mail address: [email protected] (J. Liu).
0305-1978/$ - see front matter # 2004 Elsevier Ltd. All rights reserved. doi:10.1016/j.bse.2004.02.008
862
H. Yang et al. / Biochemical Systematics and Ecology 32 (2004) 861–866
known as ‘‘DiDa’’ in Tibet, that is used to cure a range of diseases, including gall and liver disorders (Yang, 1991). Recent investigations on the action of this drug have revealed that its major active phytochemical constituents consist of swertiamarin, oleanolic acid, mangiferin, swertisin, and other xanthones (Ding et al., 1982, 1988). These active constituents have been found separately or collectively to have hepatoprotective (Liu et al., 1993; Komatsu et al., 1997), hypoglycemic (Song, 1986), anti-inflammatory (Banerjee et al., 2000), anti-oxidative (Ashida et al., 1994; Born et al., 1996), anti-tubercular (Ghosal and Chaudhuri, 1975; Bian et al., 1998), and anti-fungal (Rodriguez et al., 1995) activities, together with other pharmacological properties (Rafatullah et al., 1993; Singha et al., 1993; Ji, 1995). Although endemic to the Tibetan Plateau, this species is widely distributed, from the eastern parts of the Plateau in Qinghai province, to western parts, in Tibet, at altitudes ranging from 2200 to 3800 m (Ho, 1998). Many widely distributed species show physiological trends along altitudinal gradients (Ko¨ner, 1999). For example, contents of carotenoids, flavonoids, sucrose, fructose, glucose and total soluble sugar (Han et al., 1998), the phyotosynthetic rate, and both the light compensation and saturation points tend to rise, while the photorespiration rate tends to decline as altitude increases (Woodward, 1986). However, differences in other characters, such as anatomical structures, are not usually pronounced (Liu and Noshiro, 2003). Geographical variations in levels of phytochemicals can occur in some species (e.g. volatile terpenoids in Juniperus; Adams, 1983, 1986). The purpose of this article is to statistically compare the concentration of seven active constituents in S. franchetiana using materials collected from different altitudes to determine whether the accumulation of these compounds is affected by altitude.
2. Materials and methods 2.1. Materials The plant materials were collected from seven populations growing between 2200 and 3960 m (Table 1), covering the entire altitudinal range of this species. All materials were collected at the flowering stage in order to avoid differences associaTable 1 The location and altitude of Swertia franchetiana populations sampled in this study Population Bayi Nanshan Huangzhong Linzhi Jiacha Linzhi Changdu
Location Bayi, Xining, Qinghai Nanshan, Xining, Qinghai Huangzhong, Qinghai Linzhi, Tibet Jiacha, Tibet Linzhi, Tibet Changdu, Tibet
Altitude (m) 2200 2300 2620 2950 3330 3460 3960
Longitude v
Latitude 0
101 52.304 v 101 47.9260 v 101 32.9700 v 94 22.6310 v 92 39.2430 v 94 43.6550 v 97 17.160
v
36 v 36 v 36 v 29 v 29 v 29 v 30
33.6290 35.0010 47.6800 20.3990 06.9950 41.9790 41.380
H. Yang et al. / Biochemical Systematics and Ecology 32 (2004) 861–866
863
ted with different growth stages of the plant. In each population, we randomly chose 30 individuals that were spaced more than 1 m apart. 2.2. Analytical methods All analyses were repeated at least four times, using the methods suggested by Demizu et al. (1986) and Menkovic´ et al. (2000a,b). The air-dried individuals were cut, crushed and mixed completely. Methanol (70%) was then used to extract swertiamarin, mangiferin and swertisin; and 100% ethanol to extract oleanolic acid and the three xanthones. All extracts were analyzed using an HPLC chromatograph (Waters 600E) equipped with an ultraviolet detector (Waters 486) and a Phenomenex Kromasil C18, 5 lm, 250 4:60 mm column. The HPLC protocol developed for analyzing the bitter-tasting swertiamarin, flavonoids (swertisin) and xanthones by Demizu et al. (1986) was adopted, using methanol:water as the mobile phase in the proportions 32:68 for swertiamarin and mangiferin, 45:55 for swertisin, 96:4 for oleanolic acid, and 78:22 for the three xanthones. To calculate the contents of the seven compounds, the previously separated standards were weighed and dissolved in 1 ml of methanol or ethanol. This was diluted to give a series of concentrations (0.1, 0.2, 0.3, 0.4 and 0.5 mg/ml). Three injections were performed for each dilution. Concentrations were determined by injecting 10 ll of the standard solutions or sample extracts at a flow rate of 1 ml/min at room temperature. The peak wavelengths were 227 nm for swertiamarin, 254 nm for mangiferin, swertisin and the three xanthones, and 220 nm for oleanolic acid. A standard calibration curve was generated for each compound using the linear leastsquares regression equation derived from the peak area. 2.3. Statistical analyses All statistical analyses were performed using Microsoft Excel 2000 or the SPSS 8.0 for Windows software package. The mean values obtained for the different groups were compared by one-way ANOVA, post hoc-LSD and t-tests. Differences between means were assumed to be statistically significant at probability levels <0.05. Simple linear correlation analysis was used to obtain a measure of the correlation and the strength of the relationships between the variables.
3. Results and discussion Fig. 1 shows variations in the levels of the seven major active constituents (1–7) of S. franchetiana along the altitudinal gradient in all sampled localities. No distinct trend was revealed for the other constituents except 6, which showed a negative correlation with altitude. This finding is further confirmed by the matrix correlation of all analyzed parameters (Table 2). A significant correlation between the concentration of 6 and altitude was observed, and a significant negative correlation between the levels of 1 and 7.
864
H. Yang et al. / Biochemical Systematics and Ecology 32 (2004) 861–866
Fig. 1. Variations in mean content of the seven active constituents: (1) swertiamarin, (2) oleanolic acid, (3) swertisin, (4) mangiferin, (5) 1,5,8-trihydroxy-3-methoxyxanthone, (6) 1,8-dihydroxy-3,7-dimethoxyxanthone and (7) 1,8-dihydroxy-3,5-dimethoxyxanthone along the altitudinal gradient. Values are means. SEM values (not shown for clarity) are 10% of the means, n 4.
Despite the general absence of altitude-associated trends in phytochemical constituents of S. franchetiana, there was considerable variation in the content of certain compounds among some populations, which was statistically significant (p < 0:001). For example, the contents of 1 showed significant differences between all of the populations (p < 0:001) except for Bayi (2200 m), Nanshan (2300 m) and Jiacha (3330 m) (p > 0:05). The content of 2 did not differ significantly between Bayi (2200 m) and Nanshan (2300 m) (p > 0:05), but it varied significantly among the other populations (p < 0:001). The contents of both 4 and 6 were not significantly different in the Jiacha (3330 m) and Changdu (3960 m) populations (p > 0:05), but differed significantly among the other populations (p < 0:001). Matrix correlation analyses confirmed significant correlations between concentration of these two compounds and latitude and longitude (Table 2). It seems Table 2 Matrix correlation of altitude, latitude, longitude and analyzed constituents: (1) swertiamarin, (2) oleanolic acid, (3) swertisin, (4) mangiferin, (5) 1,5,8-trihydroxy-3-methoxyxanthone, (6) 1,8-dihydroxy-3,7dimethoxyxanthone and (7) 1,8-dihydroxy-3,5-dimethoxyxanthone 1 Altitude Latitude Longitude 1 2 3 4 5 6
0.021 0.381 0.357
2 0.620 0.224 0.425 0.369
3 0.436 0.560 0.437 0.647 0.439
4 0.047 0.896 0.901 0.236 0.366 0.374
5 0.382 0.565 0.669 0.278 0.522 0.168 0.546
6
7
0.849 0.911 0.873 0.264 0.154 0.631 0.709 0.345
significant at the 0.05 probability level, significant at the 0.001 probability level.
0.012 0.227 0.387 0.763 0.005 0.699 0.289 0.039 0.497
H. Yang et al. / Biochemical Systematics and Ecology 32 (2004) 861–866
865
likely that the differences in phytochemical constituents of S. franchetiana are more geographical than altitudinal. Two possible factors might account for these variations. First, ecological factors other than altitude, e.g. soil type, temperature, and precipitation, might affect the synthesis and turnover of secondary compounds (Nobel, 1991). Second, their ability to synthesize the seven selected compounds in these populations might be subject to genetic controls. Genetic differentiation generally has stronger effects on the contents of secondary compounds than ecological factors (Hashimoto and Yamada, 1994) and the mutation of a single gene might affect the production of certain compounds (Vogel et al., 1996). More studies are needed to clarify the relationships between synthesis of these compounds, genetic controls and ecological factors.
Acknowledgements We thank Miss Song Yali for her help with the HPLC experiments. We are also indebted to Dr John Blackwell and Prof. Robert Adams for polishing and improving the manuscript. Support for this research was provided by the Chinese Academy of Sciences (Key Innovation Plan KSCX-SW-106, Special Fund of Outstanding PhD Dissertation and Training Qualified People Plan ‘‘Hope of the West China’’) and the Chinese Research Project for Students Returned from Overseas fund.
References Adams, R.P., 1983. Infraspecific terpenoid variation in Juniperus scopulorum: evidence for Pleistocene refugia and recolonization in western North America. Taxon 32, 30–46. Adams, R.P., 1986. Geographic variation in Juniperus silicola and J. virginiana of the southeastern United States: multivariate analyses of morphology and terpenoids. Taxon 35, 61–76. Ashida, S., Noguchi, S.F., Suzuki, T., 1994. Antioxidative components, xanthone derivatives, in Swertia japonica Makino. J. Am. Oil Chem. Soc. 71 (10), 1095–1099. Banerjee, S., Sur, T., Kumar, M.S., Chandra, D.P., Sikdar, S., 2000. Assessment of the anti-inflammatory effects of Swertia chirata in acute and chronic experimental models in male albino rats. Indian J. Pharm. 32, 21–24. Bian, Q.Y., Luo, N.C., Xiao, P.G., 1998. Four xanthone glycosides from Swertia calycina Franch. Pharm. Pharm. Com. 4 (12), 597–598. Born, M., Carrupt, P.A., Zini, R., Bree, F., Tillement, J.P., Hostettmann, K., Testa, B., 1996. Electrochemical behavior and antioxidant activity of some natural polyphenols. Helv. Chim. Acta 79 (4), 1147–1158. Demizu, S., Ohshima, Y., Hiraga, Y., Takahashi, K., 1986. High-performance liquid chromatographic analysis of Swertia herb. J. Chromatogr. 360, 307–311. Ding, J.Y., Fan, S.F., Hu, B.L., Sun, H.F., 1988. On the xanthone glycosides and flavonoid glucoside from Swertia franchetiana H. Smith. Acta Bot. Sin. 30, 414–419, (in Chinese). Ding, J.Y., Fan, S.F., Hu, B.L., Sun, H.F., 1982. Separation and identification of xanthone glycosides in Swertia franchetiana H. Smith. Acta Biol. Plateau Sin. 1, 267–269, (in Chinese). Ghosal, S., Chaudhuri, R.K., 1975. Chemical constituents of Gentianaceae. XVI. Antitubercular activity of xanthones of Canscora decussata. J. Pharm. Sci. 64, 888–889.
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Han, F., Ben, G.Y., Shi, S.B., 1998. A comparative study on the resistance of Kobresia humilis grown at different altitudes in Qinghai–Tibet Plateau. Acta Ecol. Sin. 18, 654–659, (in Chinese). Hashimoto, T., Yamada, Y., 1994. Alkaloid biogenesis: molecular aspects. Annu. Rev. Plant Physiol. Plant Mol. Biol. 45, 257–285. Ho, T.N., 1988. Flora Reipublicae Popularis Sinicae. Tomus 62. Science Press, Beijing, (in Chinese). Ji, Y.B., 1995. Pharmacology and Application of the Active Constituents in Chinese Traditional Medicines. Heilongjiang Science and Technology Press, Heilongjiang, pp. 453, (in Chinese). Komatsu, K., Purusotam, B., Yamaji, S., Kadota, S., Namba, T., 1997. A comparative study on Swertia herba from Japan, Nepal, and China and their hypoglycemic activities in streptozotocin (STZ)induced diabetic rats. Nat. Med. 51, 265–268. Ko¨ner, C., 1999. Alpine Plant Life—Functional Plant Ecology of High Mountain Ecosystems. Springer, Berlin, pp. 259–290. Liu, J.Q., Noshiro, S., 2003. Lack of latitudinal trends in wood anatomy of Dodoneaea viscosa (Sapindaceae), a species with a worldwide distribution. Am. J. Bot. 90, 532–539. Liu, J.Q., Lu, Y.P., Madhu, C., Klaassen, C., 1993. Protective effects of oleanolic acid on acetaminophen-induced hepatotoxicity in mice. J. Pharm. Exp. Ther. 266, 1607–1613. Menkovic´, N., Sˇavikin-Fodulovic´, K., Savin, K., 2000a. Chemical composition and seasonal variations in the amount of secondary compounds in Gentian lutea leaves and flowers. Plant. Med. 66, 178–180. Menkovic´, N., Sˇavikin-Fodulovic´, K., Momcilovic´, I., Grubisˇic´, D., 2000b. Quantitative determination of secoiridoid and c-pyrone compounds in Gentiana lutea cultured in vitro. Plant. Med. 66, 96–98. Nobel, P.S., 1991. Physicochemical and Environmental Plant Physiology. Academic Press, San Diego, CA. Rafatullah, S., Tariq, M., Mossa, J.S., Al-Yahya, M.A., Al-Said, M.S., Ageel, A.M., 1993. Protective effect of Swertia chirata against indomethacin and other ulcerogenic agent-induced gastric ulcers. Drugs Exp. Clin. Res. 19 (2), 69–73. Rodriguez, S., Wolfender, J.-L., Hakizamungu, E., Hostettmann, K., 1995. An antifungal naphthoquinone, Xanthones and secoiridoids from Swertia calycina. Planta Med. 61 (4), 362–364. Singha, U.K., Guru, P.Y., Sen, A.B., Tandon, J.S., 1993. Antileishmanial activity of traditional plants against Leishmania donovani in golden hamsters. Int. J. Pharm. 30, 289–295. Song, W.Z., 1986. A general survey on medicinal plants of Gentianaceae in China. Bull. Chin. Materia Med. 11, 643–647, (in Chinese). Woodward, F.I., 1986. Ecophysiological studies on the shrub Baccinium myrtillus L. taken from a wide altitudinal range. Oecologia 70, 580–586. Vogel, B.S., Wildung, M.R., Vogel, G., Croteau, R., 1996. Abietadiene Synthase from grand fir (Abies grandis). J. Biol. Chem. 38, 23262–23268. Yang, Y.C., 1991. Tibetan Medicines. Qinghai People Press, Qinghai, (in Chinese).