Gelsemium Alkaloids, Immunosuppressive Agents from Gelsemium Elegans

FITOTE-02441; No of Pages 5 Fitoterapia xxx (2012) xxx–xxx

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1 Gelsemium alkaloids, immunosuppressive agents from Gelsemium elegans

Q12 You-Kai Xu a,⁎, Shang-Gao Liaob b,⁎, Zhi Na c, Hua-Bin Hu a, Yan Li d, Huai-Rong Luo d

3 a CAS Key Laboratory of Tropical Forest Ecology, Xishuangbanna Tropical Botanical Garden, Chinese Academy of Sciences, Mengla, Yunnan Province 666303, PR China 4 b School of Pharmacy, Guiyang Medical College, 9 Beijing Road, Guiyang 550004, PR China 5 c Laboratory of Tropical Plant Resource Science, Xishuangbanna Tropical Botanical Garden, Chinese Academy of Sciences, Mengla, Yunnan 666303, PR China 6 d State Key Laboratory of Phytochemistry and Plant Resources in West China, Kunming Institute of Botany, Chinese Academy of Sciences, Kunming, Yunnan 650204, PR 7 China 8

1110 article info abstract

1612 Article history: Bioassay-guided isolation of the stems of Gelsemium elegans has led to the isolation of two new 1713 Received 14 January 2012 Gelsemium alkaloids, 21-(2-oxopropyl)-koumine (1) and 11-methoxygelselegine (2), and two 1814 Accepted in revised form 22 April 2012 known alkaloids, koumine (3) and gelselegine (4). The structures of 1–2 were determined by 15 19 Available online xxxx spectroscopic (for both) and single-crystal X-ray diffraction (for 1) analysis. All compounds 20 isolated were evaluated for their potential as immunosuppressive, nerve growth factor 2124 Keywords: enhancing, and antitumor agents and the data suggested that Gelsemium alkaloids of different 2225 Gelsemium elegant structural types possibly have potential as immunosuppressive agents. 26 23 21-(2-Oxopropyl)-koumine © 2012 Published by Elsevier B.V. 27 11-Methoxygelselegine 282930 Immunosuppressive activity

32 1. Introduction treatment of malignant tumors, skin diseases (e.g., psoriasis), 47 rheumatism, and rheumatic arthritis. In the previous investi- 48 33 Gelsemium elegans (Loganiaceae), a well-known toxic gation of the leaves and stems of G. elegans, we have reported 49 34 perennial climbing vine native to Southeast Asia, has attracted three Gelsemium alkaloids bearing an iridoid monoterpenoid 50 35 natural products chemists's great attention due to the moiety [6]. Yet, only no to moderate anticancer activity was 51 36 presence of structurally complex and diversified alkaloids. observed for these compounds. Correlations of the traditional 52 37 More than eighty indole alkaloids of six structural skeletons use of G. elegans with biological activities of the alkaloids 53 38 (sarpagine, koumine, humantenine, gelsedine, gelsemine, and produced by the plant remain unclear. Inspired by the 54 39 yohimbane types) have been isolated from the leaves, stems, traditional use of G. elegans in the treatment of rheumatic 55 40 seeds, and roots of the plant [1–11]. Despite the advances in arthritis, an autoimmune disease that causes chronic joint 56 41 the chemical investigations of Gelsemium alkaloids in recent inflammation, we carried out a bioassay-guided investigation 57 42 years, only anticancer activity (and anti-epidermal cell of G. elegans. As a result, two new and two known Gelsemium 58 43 proliferation) was reported for a limited number of com- alkaloids were isolated from the immunosuppressively active 59 44 pounds and only one alkaloid, koumine, was reported to show fractions of G. elegans. The structures of these new alkaloids 60 45 immunosuppressiveUNCORRECTED activity [12,13]. G. elegans, for a very were elucidated by PROOF means of spectroscopic (for 1 and 2) and 61 46 long time, has been used in Chinese folk medicine for the single-crystal X-ray diffraction (for 1) analysis. The alkaloids 62 isolated were evaluated for immunosuppressive, nerve 63 growth factor (NGF)-mediated neurite outgrowth enhancing, 64 and antitumor activity. We report herein the isolation and 65 structural elucidation of these new alkaloids and their 66 ⁎ Corresponding authors. Tel.: +86 691 8715910/851 6908468 8712; biological activity evaluation. 67 fax: +86 691 8715070. – 68 E-mail addresses: [email protected] (Y-K. Xu), [email protected] The structures of 1 2 were determined by spectroscopic (S-G. Liaob). and single-crystal X-ray diffraction analysis (for 1). 69

Please cite this article as: Xu Y-K, et al, Gelsemium alkaloids, immunosuppressive agents from Gelsemium elegans, Fitoterapia (2012), doi:10.1016/j.ﬁtote.2012.04.023 2 Y-K. Xu et al. / Fitoterapia xxx (2012) xxx–xxx

70 2. Experimental extracted with CHCl3 to give a crude alkaloid extract (40 g). 112 Immunosuppressive assay demonstrated that, at 1 μg/mL, the 113 71 2.1. General crude alkaloid showed, respectively, 10% and 37% inhibition of 114 T lymphocyte proliferation stimulated by Con A and B 115 72 Optical rotations were measured on a Perkin-Elmer 241 lymphocyte proliferation stimulated by LPS. The crude alkaloid 116 73 polarimeter. UV spectra were obtained on a UV-210A spec- was therefore subjected to silica gel column chromatography 117

74 trometer. CD spectra were recorded on a JASCO 810 spectrom- (CHCl3/MeOH, 100:0→1:3) to give four major fractions (1–4). 118 75 eter. IR spectra were recorded on a Bio-Rad FTS-135 Fraction 1 (2 g) was subjected to chromatography over silica 119

76 spectrometer with KBr pellets. NMR spectra were recorded in gel (CHCl3/MeOH, 50:1→10:1) to give 4 major fractions 120 77 CD3OD or CDCl3 on a Bruker AM-500 or AM-400 spectrometer (Fr. 1‐1toFr.1‐4). Fraction 1‐3 (300 g) was purified by 121 78 with TMS as the internal standard. HREIMS and ESIMS were semi-preparative HPLC (MeOH/H2O, 50/50→80/20; flow rate: 122 79 carried out on a Finnigan MAT 90 mass spectrometer and VG 4 mL/min) to give 1 (20 mg) and 3 (50 mg). Fraction 2 (3 g) 123

80 Auto-Spec-3000 instrument, respectively. Chromatographic was separated by silica gel (CH3Cl/MeOH, 30:1→5:1) to give 124 81 separations were performed on silica gel (90–150 μm; Qingdao three major fractions (Fr. 2‐1toFr.2‐3). Fr. 2‐2 was purified by 125

82 Haiyang Chemical Plant, Qingdao, China) columns, Sephadex semi-preparative HPLC (MeOH/H2OH, 60:40→90:10) to give 2 126 83 LH-20 (40–70 μm; Amersham Pharmacia Biotech AB, Uppsala, (15 mg). Fraction 3 (5 g) was subjected to silica gel column 127

84 Sweden) columns, and Lichroprep RP-18 gel (40–63 μm; chromatography (CHCl3/MeOH, 10:1→3:1) to give three major 128 85 Merck, Darmstadt, Germany) columns. Semi‐preparative fractions (Fr. 3‐1toFr.3‐2). Fr. 3‐1 was purified by semi- 129

86 HPLC was performed on an XTerra prep RP-18 (10 μm, Waters preparative HPLC (MeOH/H2O, 40:60→80:20) to give 4 130 87 Corp., Ireland) column (10×250 mm) eluted with MeOH/H2O (100 mg). 131 88 from 40:60 to 90:10 for 15 min at a flow rate of 4 mL/min; the 21-(2-Oxopropyl)-koumine (1, Fig. 1): Colorless crystals 132

89 detector used was PDA (200–400 nm) at 33 °C. Precoated silica (MeOH). M.p 256–258 °C; [α]D −56 (c 0.1, MeOH); CD 133 90 gel GF254 and HF254 plates (Qingda Haiyang Chemical Plant, (MeOH): λ (Δε)=265 (−3.70), 224 (2.83), 214 (−0.11); 134

91 Qingdao, China) were used for TLC. UV(MeOH), λmax (log ε): 258 (1.79), 221 (1.21), 216 (1.23), 135 196 (1.43); IR (KBr) νmax 3462, 2920, 2877, 1630, 1587, 1446, 136 − 1 1 13 92 2.2. Plant material 1080, 775 cm ; H and C NMR (in CD3OD), see Table 1; 137 EIMS m/z: 362 (34), 322 (18), 308 (70), 307 (85), 306 (100), 138 93 The perennial stems (2–3cmindiameter)ofG. elegans were 305 (95), 291 (65), 279 (72), 277 (75), 263 (82), 251 (76), 139 94 collected in Xishuangbanna Tropical Botanical Garden, Chinese 235 (72), 223 (82), 218 (82), 206 (84), 194 (84), 180 (82), 140 95 Academy of Sciences, Yunnan Province, China, in October. 2008. 167 (76); HREIMS m/z : 362.1995 [M]+ (calcd for 141

96 A voucher specimen (accession number: 2008081203) was C23H26N2O2, 362.1994). 142 97 deposited in the Herbarium of Xishuangbanna Tropical Botanical 11-Methoxygelselegine (2, Fig. 1): white amorphous 143

98 Garden, Chinese Academy of Sciences. powder; [α]D −114 (c 0.20, EtOH); CD (MeOH): λmax (Δε): 144 270 (−9.08), 236 (43.70), 215 (−51.79); UV (MeOH), λmax 145 99 2.3. Extraction and bioassay-guided isolation (log ε): 218 (0.46), 251 (1.13), 257 (1.12), 262 (1.13), 283 146

(1.17) nm; IR (KBr) νmax 3482, 3408, 3186, 3049, 2951, 2836, 147 100 The air-dried perennial stems of G. elegans (5.0 kg) were 1686, 1626, 1502, 1433, 1222, 985, 753 cm− 1;1H and 13C 148 + 101 percolated three times with 10 L of 95% EtOH at room NMR (in CDCl3), see Table 1; ESIMS m/z: 389 [M+H] ; 149 + 102 temperature to give an ethanol extract (310 g). Immunosup- HRESIMS m/z: 389.2072 [M+H] (calcd for C21H29N2O5, 150 103 pressive activity and cytotoxicity assay indicated that the 389.2076). 151 104 extract, at 10 μg/mL, showed 30% and 40%, respectively, X-ray single-crystal structure determination of 21-(2- 152

105 inhibition of T lymphocyte proliferation stimulated by Con A oxopropyl)-koumine (1). C23H26O2N2•2H2O, M=398.49; 153 106 and B lymphocyte proliferation stimulated by LPS. The Monoclinic, space group, P21; a=8.4184(1) Å, b=15.2473 154 107 ethanolic extract (300 g) was then dissolved in water (1.0 L) (3) Å, c=8.9574(2) Å, α=90°, β = 117.8410(10) °, γ=90°, 155 3 3 108 to form a suspension, which was acidified with 10% H2SO4 to V=1016.66(3)A ,Z=2,d=1.302g/cm , crystal dimensions 156 109 around pH 3. The acidic suspension was first partitioned with 0.10 × 0.05 × 0.05 mm were used for measurement on a Bruker 157 110 EtOAc to remove the neutral compounds, and the aqueous Smart Apex-II CCD diffractometer equipped with a graphite- 158

111 phase was then basified with Na2CO3 to around pH 10 and monochromated CuKa radiation (40 kV, 30 mA), Wavelength 159 UNCORRECTED PROOF

Fig. 1. Structure of compounds 1–4.

a b c d t1:4 δC δH δC δH t1:5 2 187.4 175.7 3.62 (br d, J=4.4 Hz) t1:6 3 72.0 4.83 (br s) 75.4 4.67 (br d, J=9.7 Hz) t1:7 5 59.1 3.15 (s) 60.1 2.25 (overlapped) t1:8 6 27.4 2.66 (d, J=18.5 Hz) 32.0 2.37 (overlapped) t1:9 2.36 (dd, J=18.5,3.3 Hz) t1:10 7 61.1 57.2 t1:11 8 143.6 122.0 7.30(d, J=8.2 Hz) t1:12 9 125.2 7.90 (bd, J=7.8,2.8 Hz) 126.1 6.66(dd, J=8.2,2.2 Hz) t1:13 10 128.0 7.42 (overlapped) 109.6 t1:14 11 130.0 7.44 (overlapped) 161.3 6.58 (d, J=2.2 Hz) t1:15 12 121.8 7.52 (bd, J=8.2,2.8 Hz) 96.0 t1:16 13 155.2 139.0 2.22 (overlapped) t1:17 14 25.6 1.97 (overlapped) 23.3 2.29 (overlapped) t1:18 2.74 (overlapped) 2.40 (overlapped) t1:19 15 25.0 2.85 (overlapped) 37.5 3.13 (m) t1:20 16 40.6 2.85 (overlapped) 38.4 4.21 (br s) t1:21 17 62.0 3.63 (d, J=15.2 Hz) 62.9 4.21 (br s) t1:22 4.27 (d, J=15.2 Hz) 0.96 (t, J=7.4 Hz) t1:23 18 120.9 4.88 (d, J=15.7 Hz) 9.2 t1:24 4.91 (overlapped) 2.12 (dd, J=7.4,7.4 Hz) t1:25 19 135.7 4.44 (dd, J=22.5,14.3 Hz) 22.1 2.37 (overlapped) t1:26 20 52.1 75.1 3.77 (d, J=13.0 Hz) t1:27 21 61.1 3.91 (s) 63.5 3.93 (d, J=13.0 Hz) t1:28 22 45.1 Not detected t1:29 23 208.6 t1:30 24 30.5 2.01 (s) t1:31 N-Me 40.7 2.74 (s) 4.00 (s) t1:32 N-OMe 64.0 3.81 (s) t1:33 OMe 56.0

a t1:34 Measured at 125 MHz in CD3OD. b t1:35 Measured at 500 MHz in CD3OD. c t1:36 Measured at 100 MHz in CDCl3. d t1:37 Measured at 400 MHz in CDCl3.

160161162163164165166167168169170171172173 1.54178 Å. The total number of reflections measured was 4505, method SHELXS-97 and expanded using difference Fourier 176

174 of which 3191 were observed. Final indices: R1 =0.0406, technique, refined by the program SHELXL-97 (Sheldrick 1997) 177 175 wR2 =0.1042. The crystal structure of 1 was solved by direct and the full-matrix least-squares calculations. Crystallographic 178

UNCORRECTED PROOF

Fig. 2. Key 1H–1H COSY, HMBU and ROESY correlations of compound 1.

Please cite this article as: Xu Y-K, et al, Gelsemium alkaloids, immunosuppressive agents from Gelsemium elegans, Fitoterapia (2012), doi:10.1016/j.ﬁtote.2012.04.023 4 Y-K. Xu et al. / Fitoterapia xxx (2012) xxx–xxx

(2-oxopropyl)-koumine. Compound 1 was therefore a new 200 Gelsemium alkaloid with a three-carbon extension of the 201 alkaloid backbone. 202 Compound 2 was obtained as white amorphous powder and 203

was assigned the molecular formula C21H28N2O5 on the basis of 204 the HRESIMS peak at m/z 389.2072 [M+H]+, showing 30 mass 205 units more than that of gelselegine (4) [3], a known alkaloid 206 also isolated in this investigation. The 1HNMRdataof2 were 207 very close to those of gelselegine (4) [3], the only difference 208 being the presence of an additional aromatic methoxy (δ 3.81) 209 in 2 and the replacement of a 1,2-disubstituted benzene ring 210 (δ=7.41, d, J=9.5Hz; δ=7.34, t, J=9.5Hz; δ=7.17, t, 211 J=9.5 Hz; δ=7.00, d, J=9.5Hz)in 4 by a 1,2,4-trisubstituted 212 counterpart (δ=7.30, d, J=8.2Hz;δ=6.66, dd, J=8.2,2.2Hz; 213 δ=6.58, d, J =2.2 Hz) in 2. These observations suggested that 214 2 was a 9- or 11-methoxyl derivative of 4. The coupling patterns 215 and chemical shifts of the two upfield shifted protons (δ=6.66, 216 Fig. 3. Single-crystal X-ray structures of 1. dd, J=8.2, 2.2 Hz; δ=6.58, d, J =2.2 Hz) indicated that the 217 methoxy group was located at C-11. Distinct chemical shifts for 218 C-8 (Δδ=−8.6), C-10 (Δδ=−14.1), C-11 (Δδ=−23.3), C-12 219 179 data for the structure of 1 have been deposited in the (Δδ=−11.2) also supported this assignment. Detailed analysis 220 180 Cambridge Crystallographic Data Centre (deposition number: of the HMBC and ROESY spectra (Fig. 4) further verified this 221 181 CCDC 821172). Copies of these data can be obtained free of deduction. In particular, the HMBC correlation from the methoxy 222 182 charge via www.ccdc.cam.ac.uk/conts/retrieving.html (or from to C-11 confirmed the presence of a 11-methoxy group. The 223 183 the Cambridge Crystallographic Data Centre, 12 Union Road, structure of 2 was thus established as 11-methoxygelsegine. 224 184 Cambridge CB2 IEZ. UK; fax:(+44)1223-336-033; or email: The absolute stereochemistries of alkaloids 1 were estab- 225 185 [email protected]). lished as mentioned above by X-ray crystallography using 226 anomalous dispersion effects. The results deduced through 227 186 3. Results and discussion comparison of their CD spectra with those of koumine [15] 228 were in agreement with the conclusions. Similar CD curves 229 187 Compound 1 was obtained as a colorless crystal (m.p.256– observed for compounds 2 [(MeOH): λ (Δε)270(−9.08), 236 230

188 258 °C). Its molecular formula C23H26N2O2 was established by (43.70), 215 (−51.79)] and 4 [(MeOH): λ (Δε) 263 (−18.79), 231 189 the HREIMS peak at m/z 362.1995 [M]+, showing 56 mass units 224 (15.25), 200 (−12.11)] indicated that both compounds 232 190 more than that of a known alkaloid, koumine (3) [14],isolated shared the same stereochemistry as depicted. 233 191 from this plant. The 1Hand13CNMRdataof1 were very similar Identification of the known compounds koumine (3) and 234 192 to those reported for koumine (3), the only difference being the gelselegine (4) was carried out by comparing its NMR and 235

193 presence of an additional 2-oxopropyl group (δC 45.1, CH2; EIMS data with those reported in the literature. 236 194 208.6, C; 30.5, CH3)in1 and the replacement of a methylene at UPLC-DAD-MS and UPLC-Q-TOF analyses indicated that 1 237 195 δC 57.2 in 3 by a methine at δC 61.1 in 1 for C-21. HMBC did not exist in the ethanolic extract of the plant material and 238 196 correlations (Fig. 2) from H-24 to C-23 and C-22 verified the was an artifact possibly generated from 3 and acetone. 239 197 about deduction. Single-crystal X-ray diffraction study of 1 Although acetone was not used in the current investigation, 240 198 (Fig. 3) using anomalous dispersion effects further confirmed the formation of 1 might be ascribed to the presence of trace 241 199 these assignments and allowed the establishment of 1 as 21- acetone in the partially recovered EtOH solvent. 242

UNCORRECTED PROOF

Fig. 4. Key 1H–1H COSY, HMBC and ROESY correlations of compound 2.

Please cite this article as: Xu Y-K, et al, Gelsemium alkaloids, immunosuppressive agents from Gelsemium elegans, Fitoterapia (2012), doi:10.1016/j.ﬁtote.2012.04.023 Y-K. Xu et al. / Fitoterapia xxx (2012) xxx–xxx 5 t2:1 Table 2 Cytotoxicity and immunosuppressive activity of compounds 1–4. t2:2 t2:3 Group Concentration MTT OD570 nm T cell B cell (μM) value t2:4 CPM Inhibitory rate (%) CPM Inhibitory rate (%) t2:5 Normal control 0.124 ±0.001 322±35 277±59 t2:6 Con A or LPS 24911±822 11984±1287 t2:7 1 10 −0.015 ±0.006** 10297±1246 59 2569±745 79 t2:8 1 0.050 ±0.004** 27074±576 −9 6785±800 43 t2:9 0.1 0.100 ±0.004** 24139±2242 3 9770±928 18 t2:10 2 10 0.048 ±0.013** 17743±279 29 3495±767 71 t2:11 1 0.089 ±0.019 25483±879 −2 7112±1061 41 t2:12 0.1 0.088 ±0.026 19749±3074 21 6586±1007 45 t2:13 3 10 0.116 ±0.010 24662±4932 1 11145±5127 7 t2:14 1 0.106 ±0.012 22918±15355 8 9228±2583 23 t2:15 0.1 0.098 ±0.09** 22420±2638 10 11742±2583 2 t2:16 4 10 0.025 +0.007** 24996±663 0 4515±555 62 t2:17 1 0.047 +0.005** 28199±919 −13 8316±325 31 t2:18 0.1 0.052 +0.008** 28995±641 −16 8090±485 32 t2:19 CsA CC50 1.32 μMIC50 1.37 nM IC50 0.175 μM Cytotoxicity was tested on murine spleen cells and immunosuppressive activity on Con A-induced T cell proliferation and LPS-induced B cell proliferation. Data are the mean±S.D. of 3 separate wells. **pb0.01, *pb0.05 vs. normal control. The degree of growth inhibition was calculated by the following equation: Inhibitory rate (%)=[CMP of Con A (or LPS) group − (CMP of experiment group)/CMP of Con A (or LPS) group]×100. Con A and LPS were at 5 and 10 μg/ml, t2:20 respectively. CPM=counts per minute.

244245246247248249250251252253254255256257258259260261 All the compounds isolated were evaluated for their in vitro (Grant No. KSCX2-YW-R-172) of the Chinese Academy of 292 262 immunosuppressive activity by testing their cytotoxicity on Sciences is gratefully acknowledged. Special thanks were also 293 263 murine spleen cells and inhibitory activity on concanavalin A given to the National Center for Drug Screening for the 294 264 (Con A)-induced T cell proliferation and lipopolysaccharide immunosuppressive activity evaluation. 295 265 (LPS)-induced B cell proliferation (Table 2). Our data demon- 266 strated that the Gelsemium alkaloids isolated in this investigation 267 inhibited, to varying degrees, the proliferation of T lymphocyte References 296 268 stimulated by Con A and the proliferation of B lymphocyte 269 stimulated by LPS on mice. At the concentration of 10 μM, [1] Liu ZJ, Lu RR. Chemistry and Pharmacology. In: Arnold B, editor. The 297 – 298 270 Alkaloids, vol. 33. New York: Academic Press; 1988. p. 83 140. compound 1 showed 59% inhibition of the proliferation of T [2] Ponglux D, Wongseripipatana S, Subhadhirasakul S, Takayama H, Yokota 299 271 lymphocyte stimulated by Con A and 79% the proliferation of B M, Ogata K, Phisalaphong C, Aimi N, Sakai S. Tetrahedron 1988;44: 300 272 lymphocyte stimulated by LPS. The immunosuppressive effects 5075–94. 301 – 302 273 [3] Lin LZ, Cordell GA, Ni CZ, Clardy J. Phytochemistry 1990;29:3013 7. of these alkaloids did not show a significant correlation to their [4] Lin LZ, Hu SF, Cordell GA. Phytochemistry 1996;43:723–6. 303 274 cytotoxicity. At some non-cytotoxin concentrations, some of the [5] Takayama H, Sakai SI. Chemistry and Pharmacology. In: Geoffrey AC, 304 275 alkaloids (e.g., 3) still showed obvious suppression of the editor. The Alkaloids, vol. 49. New York: Academic Press; 1997. p. 1–78. 305 306 276 mitogen-induced proliferation of lymphocyte but not in a dose- [6] Xu YK, Yang SP, Liao SG, Zhang H, Lin LP, Ding J, Yue JM. J Nat Prod 2006;69:1347–50. 307 277 dependent manner or may show an immunostimulating effect at [7] Kogure N, Ishii N, Kitajima M, Wongseripipatana S, Takayama H. Org 308 278 certain concentrations (see Table 2), which limited our further Lett 2006;8:3085–8. 309 310 279 effort to characterize their immunoregulatory effects by IC .The [8] Yamada Y, Kitajima M, Kogure N, Takayama H. Tetrahedron 2008;64: 50 7690–4. 311 280 peculiar immunoregulatory effects may suggest a complex [9] Yamada Y, Kitajima M, Kogure N, Wongseripipatana S, Takayama H. 312 281 mechanism of action for these alkaloids. These observations Tetrahedron Lett 2009;50:3341–4. 313 314 282 indicate that Gelsemium alkaloids of different structural types [10] Zhang Z, Di YT, Wang YH, Zhang Z, Mu SZ, Fang X, Zhang Y, Tan CJ, Zhang Q, Yan XH, Guo J, Li CS, Hao XJ. Tetrahedron 2009;65:4551–6. 315 283 possibly have potential as immunosuppressive agents. The [11] Kitajima M, Kobayashi H, Kogure N, Takayama H. Tetrahedron 2010;66: 316 284 results suggested that the traditional use of G. elegans for the 5987–92. 317 285 treatment of rheumatic arthritis was attributed to the presence [12] Sun LS, Lei LS, Fang FZ, Yang SQ, Wang J. Pharmacol Clin Chin Mat Med 318 – 319 286 1999;15:11 3. of immunosuppressively active agents, and the plant extracts, [13] Z.R. Wang, C.Q. Huang, Z.Y. Zhang, L.L. Zhang, J.M. Lin. J First Mil Med 320 287 particularly the alkaloid fractions obtained from this plant, might Univ 25; 562–64. 321 Q2 288 be a potential source for immunosuppressive drug development. [14] Lin LZ, Cordell GA, Ni CZ, Clardy J. Phytochemistry 1990;29:965–8. 322 UNCORRECTED[15] Kitajima M, Takayama PROOF H, Sakai SI. J Chem Soc Perkin 1 1991;8:1773–9. 323 289 Acknowledgment

290 Financial support from the key project of the Tropical 291 Characteristic Medical Resources Branch Research Centre 324

Please cite this article as: Xu Y-K, et al, Gelsemium alkaloids, immunosuppressive agents from Gelsemium elegans, Fitoterapia (2012), doi:10.1016/j.ﬁtote.2012.04.023