Chemical constituents of Caragana bungei shoots

Chemical constituents of Caragana bungei shoots

Daniil N. Olennikov^*,I; Larisa M. Tankhaeva^I; Vyacheslav V. Partilkhaev^I; Alexander V. Rokhin^II

^ILaboratory of Medical and Biological Research, Institute of General and Experimental Biology, Siberian Division, Russian Academy of Sciences, Russian Federation

^IIDepartment of Chemistry, Irkutsk State University, Russian Federation

ABSTRACT

The phytochemical study of flowering shoots of Caragana bungei Ledeb., Fabaceae, collected in Tuva Republic (Russian Federation) resulted in the isolation of sixteen compounds identified as β-sitosterol, β-sitosterol-3-O-glucoside, umbelliferone, kaempferol, quercetin, isorhamnetin-3-O-glucoside, isoquercitrin, rutin, narcissin, nicotiflorin, caffeic acid, 3-O-caffeoylquinic acid, 5-O-caffeoylquinic acid, 3,5-di-O-caffeoylquinic acid, gallic acid and sucrose. The structures of the compounds were established by spectral analyses. This is the first phytochemical investigation of C. bungei. The nicotiflorin, phenylpropanoids and gallic acid were isolated from the Caragana genus for the first time.

Keywords:Caragana bungei, Fabaceae, flavonoids, phenylpropanoids, β-sitosterol

Introduction

Caragana Fabr. is a genus of Fabaceae family including about eighty species widely spread in Europe and Asia and used as a medical, decorative and technical raw materials. Despite of wide using the targeted chemical researches of plants of this genus were started relatively recently. To date we have scientific information about chemical compositions of fifteen Caragana species (Meng et al., 2009). In Russian Federation this genus presented by 34 species mainly distributed in the steppe and mountain landscapes.

Caragana bungei Lebeb. is branched spiny shrub growing in the desert steppes and valleys of mountain rivers of Eastern and Western Siberia. The decoction of flowering shoots of this species is used in traditional medicine of Tuva and Buryatia regions as an antiinflamatory and immunostimulant means (Aseeva et al., 2008). There is no any scientific data about chemical composition of C. bungei. In this study, we present the results of phytochemical investigation of C. bungei flowering shoots. As a result sixteen compounds were isolated including β-sitosterol (1), umbelliferone (2), kaempferol (3), quercetin (4), β-sitosterol-3-O-glucoside (5), caffeic acid (6), gallic acid (7), 3,5-di-O-caffeoylquinic acid (8), isorhamnetin-3-O-glucoside (9), isoquercitrin (quercetin-3-O-glucoside, 10), 3-O-caffeoylquinic acid (11), 5-O-caffeoylquinic acid (12), rutin (quercetin-3-O-rutinoside, 13), narcissin (isorhamnetin-3-O-rutinoside, 14), nicotiflorin (kaempferol-3-O-rutinoside, 15) and sucrose (16). This is the first phytochemical investigation of C. bungei.

Material and Methods

Plant material

The shoots of Caragana bungei Ledeb., Fabaceae, were collected in flowering period in Despen locality (Tes-Hemskiĭ region, Tuva Republic, Russian Federation, 50º81'78' N, 93º78'39' E). The plant samples were identified by Prof. Aseeva T.A. (Institute of General and Experimental Biology, Ulan-Ude, Russian Federation). Voucher specimen is deposited at the Herbarium of Department of Biologically Active Substances (Institute of General and Experimental Biology, Ulan-Ude, Russian Federation; voucher specimen number Fb/s-18/02-11/0942).

General experimental procedures

Elemental composition was determined using MAT 8200 spectrometer (Thermo Finnigan). Melting points were determined with a 910 DS calorimeter (DuPont). UV spectra were recorded using a SF-2000 spectrophotometer (OKB Specter). MS spectra were registered on a LCQ mass spectrometer (Thermo Finnigan). ¹³C NMR spectra were recorded on a VXR 500S spectrometer (Varian). Optical rotations were measured on a DIP-1000 digital polarimeter (Jasco). Chromatography column was performed over silica gel 60 (230-400 mesh, Merck), Sephadex LH-20 (25-100 µm, Pharmacia), polyamide (Woelm) and pTLC was performed on Sorbfil-A silica gel TLC plates (layer thickness 1 mm; Imid Ltd.). All chemicals were analytical-grade. The Folin method was used to determine total phenolic content (TPC), as described by (Shetty et al., 1995) using gallic acid as a standard compound. Total flavonoid content (TFC) was determined by spectrophotometric method (Chirikova et al., 2010) using rutin as a standard compound. β-Sitosterol content was determined using HPTLC-densitometric method (Starek et al., 2007).

Extraction and isolation

Air-dried, ground shoots of C. bungei (1462 g) were extracted with 70% EtOH at 80 ºC three times and the extracts were concentrated under reduced pressure to yield 409.4 g of crude extract. The crude extract was re-suspended in water (1:4, v/v) and successively partitioned with CHCl₃, EtOAc, and n-BuOH. The organic layers were brought to dryness in vacuo to yield 87.7, 43.8 and 175.4 g of CHCl3 (CB-C), EtOAc (CB-E) and n-BuOH fraction (CB-B) residue respectively. The CB-C fraction (42 g) was dissolved in 1000 mL of chloroform and chromatographed over Sephadex LH-20 column (7x120 cm), eluting with CHCl₃ containing increasing concentrations of MeOH (100:0→0:100) to obtain fifty fractions (fr.C1-fr.C50). Fr.C3-fr.C6 were combined and chromatographed on a silica column (3x50 cm), eluting with CHCl₃-MeOH mixtures (100:0→70:30) to obtain twelve fractions (fr.C3-6/1-fr.C3-6/12). Fr.C3-6/3 (eluent CHCl₃-MeOH 90:10) after recrystallization (MeOH) was given 263 mg of 1. Fr.C5 was chromatographed on a silica column (2x40 cm), eluting with CHCl₃-MeOH mixtures (90:10→60:40) to obtain fifteen fractions (fr.C5/1-fr.C5/15). Fr.C5/7-fr.C5/8 were combined (eluent CHCl₃-MeOH 74:25) and was chromatographed on a pTLC (solvent toluene-EtOAc-HCOOH 6:3:1) to yield 2 (15 mg), 3 (27 mg) and 4 (37 mg). The CB-E fraction (35 g) was dissolved in 500 mL of MeOH and chromatographed over Sephadex LH-20 column (5x100 cm), eluting with MeOH-H₂O mixtures (100:0→50:50) to obtain 60 fractions (fr.E1-fr.E60). Fr.E3 was chromatographed on a silica column (2x50 cm), eluting with CHCl₃-MeOH mixtures (100:0→60:40) to obtain fifteen fractions (fr.E3/1-fr.E3/15). Fr.E3/8 (eluent-CHCl₃-MeOH 70:30) after recrystallization (MeOH) was given 24 mg of 5. Fr.E5-fr.E8 were combined and chromatographed on a polyamide column (6x100 cm), eluting with H₂O-MeOH mixtures (100:0→0:100) to obtain twenty fractions (fr.E5-8/1-fr.E5-8/20). Fr.E5-8/2-fr.E5-8/3 (eluent H₂O-MeOH 90:10, 85:15) were combined and chromatographed on a pTLC (solvent toluene-EtOAc-HCOOH 5:4:1) to yield 6 (11 mg), 7 (8 mg) and 8 (8 mg). Fr.E5-8/4-fr.E5-8/5 (eluent H₂O-MeOH 80:20, 75:25) were combined and chromatographed on a pTLC (solvent toluene-EtOAc-HCOOH 5:4:1) to yield 8 (18 mg). Fr.E11-fr.E14 were combined and chromatographed on a polyamide column (6x50 cm), eluting with H₂O-MeOH mixtures (100:0→0:100) to obtain twenty fractions (fr.E11-14/1-fr.E11-14/20). Fr.E11-14/11-fr.E11-14/14 (eluent H₂O-MeOH 60:40, 55:45) were combined and chromatographed on a pTLC (solvent EtOAc-toluene-HCOOH 6:4:1.5) to yield 9 (15 mg) and 10 (27 mg). Fr.E19-fr.E23 were combined and chromatographed on a polyamide column (6x100 cm), eluting with H₂O-MeOH mixtures (100:0→0:100) to obtain twenty fractions (fr.E19-23/1-fr.E23/20). Fr.E19-23/3-fr.E19-23/5 (eluent H₂O-MeOH 90:10-80:20) were combined and chromatographed on a pTLC (solvent EtOAc-dioxane-AcOH-H₂O 10:5:1:1) to yield 11 (11 mg) and 12 (9 mg). The CB-B fraction (87 g) was dissolved in 1500 mL of 50% EtOH and chromatographed polyamide column (10x120 cm), eluting with H₂O-EtOH mixtures (100:0→0:100) to obtain fifty fractions (fr.B1-fr.B50). Fr.B20-fr.B23 (eluent-H₂O-EtOH 60:40-54:46) were combined and chromatographed on Sephadex LH-20 column (5x80 cm), eluting with EtOH-H₂O mixtures (96:4→60:40) to obtain twenty fractions (fr.B20-23/1-fr.B20-23/20). Fr.B20-23/5-fr.B20-23/7 (eluent EtOH-H2O 80:20) after recrystallization (80% EtOH) was given 86 mg of 13. Fr.B20-23/7-fr.B20-23/9 were combined and after recrystallization (80% EtOH) was given 43 mg of 14. Fr.B27-fr.B29 (eluent H₂O-EtOH 46:54-42:58) were combined and chromatographed on a pTLC (solvent EtOAc-dioxane-AcOH-H₂O 10:5:1:1) to yield 15 (16 mg). Fr.B1 (eluent H₂O) was concentrated to syrup which crystallized at 2 ºC during 5-7 days to yield colorless prisms of 16 (53 mg).

β-Sitosterol (1): C₂₉H₅₀O, m.p. 141 ºC. ⁺FAB-MC m/z: 415 [M+H]⁺. ¹³C-NMR (CDCl₃, 125 MHz): 11.73 (C-29), 12.14 (C-18), 18.63 (C-26), 19.03 (C-21), 19.39 (C-19), 19.82 (C-27), 21.47 (C-11), 23.08 (C-28), 24.31 (C-15), 26.22 (C-25), 28.18 (C-16), 29.97 (C-23), 31.73 (C-2), 31.95 (C-8), 32.03 (C-7), 34.89 (C-22), 36.11 (C-20), 36.69 (C-10), 37.24 (C-1), 40.27 (C-12), 42.32 (C-4), 42.61 (C-13), 50.33 (C-24), 50.92 (C-9), 56.11 (C-17), 57.02 (C-14), 71.97 (C-3), 121.92 (C-6), 140.87 (C-5).

Umbelliferone (2): C₉H₆O₃, m.p. 232 ºC. UV λ_max nm MeOH: 214 sh, 248 sh, 299 sh, 327. +FAB-MC m/z: 163 [M+H]⁺. ¹³C-NMR (DMSO-d₆, 125 MHz): 103.02 (C-8), 111.60 (C-10), 111.63 (C-3), 113.27 (C-6), 129.97 (C-5), 144.24 (C-4), 156.12 (C-9), 160.71 (C-2), 161.93 (C-7).

Kaempferol (3): C₁₅H₁₀O₆, m.p. 275 ºC. UV λ_max nm MeOH: 258, 325 sh, 370. +FAB-MC m/z: 287 [M+H]⁺. ¹³C-NMR (DMSO-d₆, 125 MHz): 94.38 (C-8), 99.02 (C-6), 103.92 (C-10), 116.18 (C-3',5'), 123.47 (C-1'), 130.38 (C-2',6'), 136.54 (C-3), 147.63 (C-2), 158.03 (C-9), 160.34 (C-4'), 162.14 (C-5), 165.22 (C-7), 176.71 (C-4).

Quercetin (4): C₁₅H₁₀O₇, m.p. >300 ºC. UV λ_max nm MeOH: 259, 308 sh, 376. +FAB-MC m/z: 303 [M+H]⁺. ¹³C-NMR (DMSO-d₆, 125 MHz): 93.74 (C-8), 98.47 (C-6), 104.81 (C-10), 115.21 (C-2'), 115.97 (C-5'), 121.04 (C-6'), 122.49 (C-1'), 134.71 (C-3), 145.33 (C-3'), 146.22 (C-2), 147.27 (C-4'), 158.74 (C-9), 161.52 (C-5), 164.67 (C-7), 178.53 (C-4).

β-Sitosterol-3-O-glucoside (5): C₃₅H₆₁O₆, m.p. 281 ºC. +FAB-MC m/z: 577 [M+H]⁺. ¹³C-NMR (CDCl₃, 125 MHz): 11.62 (C-29), 12.46 (C-18), 18.35 (C-26), 19.37 (C-19), 19.83 (C-21), 20.51 (C-27), 21.82 (C-11), 23.70 (C-28), 25.72 (C-15), 25.94 (C-25), 29.48 (C-23), 29.74 (C-16), 29.93 (C-2), 33.07 (C-8), 33.19 (C-7), 37.15 (C-10), 37.38 (C-20), 38.82 (C-1), 39.73 (C-22), 40.33 (C-12), 42.97 (C-13), 44.01 (C-4), 50.21 (C-9), 50.41 (C-24), 56.26 (C-17), 57.31 (C-14), 61.58 (Glc, C-6'), 70.93 (Glc, C-4'), 74.77 (Glc, C-2'), 76.63 (Glc, C-5'), 77.02 (Glc, C-3'), 81.12 (C-3), 103.57 (Glc, C-1'),122.15 (C-6), 140.52 (C-5).

Caffeic acid (6): C₉H₈O₄, m.p. 224 ºC. UV λ_max nm MeOH: 238, 301 sh, 325. +FAB-MC m/z: 181 [M+H]⁺. ¹³C-NMR (CD₃OD, 125 MHz): 114.92 (C-8), 115.09 (C-2), 116.11 (C-5), 121.24 (C-6), 125.71 (C-1), 144.37 (C-7), 145.37 (C-3), 148.38 (C-4), 168.98 (C-9).

Gallic acid (7): C₇H₆O₅, m.p. 250 ºC. UV λ_max nm MeOH: 275. +FAB-MC m/z: 171 [M+H]⁺. ¹³C-NMR (DMSO-d₆, 125 MHz): 109.08 (C-2, C-6), 120.47 (C-1), 137.84 (C-4), 145.11 (C-3, C-5), 167.74 (C-7).

3,5-Di-O-caffeoylquinic acid (8): C₂₅H₂₄O₁₂, m.p. 170 ºC. UV λ_max nm MeOH: 220, 246 sh, 297 sh, 333. +FAB-MC m/z: 517 [M+H]⁺. ¹³C-NMR (CD₃OD, 125 MHz): 74.62 (C-1), 36.08 (C-2), 72.21 (C-3), 70.54 (C-4), 72.04 (C-5), 37.42 (C-6), 114.98 (C-8''), 115.20 (C-2''), 115.27 (C-2'), 115.51 (C-8'), 116.41 (C-5''), 116.41 (C-5'), 123.01 (C-6'), 123.42 (C-6''), 127.71 (C-1''), 127.81 (C-1'), 146.29 (C-3''), 146.74 (C-3'), 147.08 (C-7'), 147.37 (C-7''), 149.20 (C-4'), 149.37 (C-4''), 168.64 (C-9''), 168.91 (C-9'), 176.87 (C-7);

Isorhamnetin-3-O-glucoside (9): C₁₅H₁₀O₆, m.p. 267 ºC. UV λ_max nm MeOH: 258, 305, 361. +FAB-MC m/z: 479 [M+H]⁺. ¹³C-NMR (CD₃OD, 125 MHz): 55.24 (3'-OCH₃), 61.37 (Glc, C-6''), 71.48 (Glc, C-4''), 75.63 (Glc, C-2''), 76.40 (Glc, C-5''), 77.11 (Glc, C-3''), 94.37 (C-8), 99.12 (C-6), 104.52 (Glc, C-1''), 105.74 (C-10), 112.87 (C-2'), 114.12 (C-5'), 123.24 (C-6'), 123.97 (C-1'), 135.22 (C-3), 148.61 (C-4'), 150.24 (C-3'), 158.14 (C-2), 158.61 (C-9), 164.27 (C-5), 166.12 (C-7), 179.21 (C-4).

Isoquercitrin (10): C₂₁H₂₀O₁₂, m.p. 226 ºC. UV λ_max nm MeOH: 257, 359. +FAB-MC m/z: 465 [M+H]⁺. ¹³C-NMR (CD₃OD, 125 MHz): 61.42 (Glc, C-6''), 70.95 (Glc, C-4''), 75.31 (Glc, C-2''), 76.37 (Glc, C-5''), 77.25 (Glc, C-3''), 94.52 (C-8), 99.03 (C-6), 102.67 (Glc, C-1''), 103.04 (C-10), 114.93 (C-2'), 115.98 (C-5'), 122.54 (C-6'), 123.24 (C-1'), 135.74 (C-3), 144.35 (C-3'), 149.60 (C-4'), 159.05 (C-9), 160.71 (C-2), 163.58 (C-5), 165.81 (C-7), 179.55 (C-4).

3-O-Caffeoylquinic acid (11): C₁₆H₁₈O₉, m.p. 206 ºC. UV λ_max nm MeOH: 240, 302 sh, 324. +FAB-MC m/z: 354 [M+H]⁺. ¹³C-NMR (CD₃OD, 125 MHz): 40.84 (C-6), 38.72 (C-2), 71.54 (C-5), 72.67 (C-4), 73.27 (C-3), 77.41 (C-1), 115.14 (C-2'), 115.61 (C-8'), 116.51 (C-5'), 122.71 (C-6'), 127.84 (C-1'), 146.50 (C-3'), 146.90 (C-7'), 149.33 (C-4'), 169.51 (C-9'), 182.38 (C-7).

5-O-Caffeoylquinic acid (12): C₁₆H₁₈O₉, m.p. 210 ºC. UV λ_max nm MeOH: 241, 305 sh, 326. +FAB-MC m/z: 354 [M+H]⁺. ¹³C-NMR (CD₃OD, 125 MHz): 39.18 (C-2), 40.73 (C-6), 71.87 (C-3), 72.41 (C-4), 73.68 (C-5), 77.75 (C-1), 115.18 (C-2'), 115.52 (C-8'), 116.58 (C-5'), 122.93 (C-6'), 127.78 (C-1'), 146.80 (C-3'), 146.94 (C-7'), 149.75 (C-4'), 169.37 (C-9'), 181.10 (C-7).

Rutin (13): C₂₇H₃₀O₁₆, m.p. 243 ºC. UV λ_max nm MeOH: 263, 364. +FAB-MC m/z: 611 [M+H]⁺. ¹³C-NMR (CD₃OD, 125 MHz): 17.81 (Rha, C-6'''), 68.31 (Glc, C-6''), 69.84 (Rha, C-5'''), 71.34 (Rha, C-2'''), 72.23 (Rha, C-3'''), 73.83 (Glc, C-4''), 74.34 (Rha, C-4'''), 75.68 (Glc, C-2''), 77.25 (Glc, C-5''), 78.14 (Glc, C-3''), 94.63 (C-8), 99.82 (C-6), 102.08 (Rha, C-1'''), 104.51 (Glc, C-1''), 105.67 (C-10), 115.47 (C-2'), 116.67 (C-5'), 123.27 (C-6'), 123.75 (C-1'), 135.77 (C-3), 145.61 (C-3'), 149.92 (C-4'), 159.05 (C-9), 159.21 (C-2), 163.41 (C-5), 165.97 (C-7), 179.32 (C-4).

Narcissin (14): C₂₈H₃₂O₁₆, m.p. 187 ºC. UV λ_max nm MeOH: 254, 356. +FAB-MC m/z: 625 [M+H]⁺. ¹³C-NMR (CD₃OD, 125 MHz): 17.73 (Rha, C-6'''), 55.01 (3'-OCH₃), 68.37 (Glc, C-6''), 69.52 (Rha, C-5'''), 71.18 (Rha, C-2'''), 71.48 (Glc, C-4''), 71.82 (Rha, C-3'''), 74.59 (Rha, C-4'''), 75.63 (Glc, C-2''), 76.40 (Glc, C-5''), 77.11 (Glc, C-3''), 94.32 (C-8), 99.30 (C-6), 101.92 (Rha, C-1'''), 104.52 (Glc, C-1''), 105.11 (C-10), 112.96 (C-2'), 114.82 (C-5'), 123.02 (C-6'), 123.74 (C-1'), 135.18 (C-3), 148.71 (C-4'), 150.02 (C-3'), 158.30 (C-9), 158.37 (C-2), 163.37 (C-5), 165.42 (C-7), 178.84 (C-4).

Nicotiflorin (15): C₂₇H₃₀O₁₅, m.p. 185 ºC. UV λ_max nm MeOH: 268, 351. +FAB-MC m/z: 595 [M+H]⁺. ¹³C-NMR (CD₃OD, 125 MHz): 17.68 (Rha, C-6'''), 68.45 (Glc, C-6''), 69.63 (Rha, C-5'''), 71.52 (Glc, C-4''), 71.95 (Rha, C-2'''), 72.27 (Rha, C-3'''), 74.45 (Rha, C-4'''), 75.71 (Glc, C-2''), 76.94 (Glc, C-5''), 78.39 (Glc, C-3''), 94.75 (C-8), 99.97 (C-6), 102.18 (Rha, C-1'''), 104.49 (Glc, C-1''), 105.71 (C-10), 116.40 (C-3',5'), 123.11 (C-1'), 132.37 (C-2',6'), 135.65 (C-3), 158.57 (C-9), 158.63 (C-2), 159.71 (C-4'), 161.82 (C-5), 166.07 (C-7), 179.29 (C-4).

Sucrose (16): C₁₂H₂₂O₁₁, m.p. 185 ºC. [α]_D +65.9º (H₂O; c 5.0). ¹³C-NMR (D₂O, 125 MHz): 62.82 (Glc, C-6), 64.02 (Frc, C-1'), 65.05 (Frc, C-6'), 72.01 (Glc, C-4), 73.79 (Glc, C-2), 75.11 (Glc, C-5), 75.37 (Glc, C-3), 76.89 (Frc, C-4'), 79.18 (Frc, C-3'), 84.01 (Frc, C-5'), 95.15 (Glc, C-1), 106.21 (Frc, C-2').

HPTLC-analysis conditions

Powdered plant material (500 mg) was extracted for 30 min with 25 mL of MeOH at 80 ºC in a water bath two times. The samples were then filtered through a cellulose acetate filter (0.50 µm; Reakhim) and concentrated. The residue was re-dissolved in 5 mL of MeOH. The sample solutions were stored at 4 ºC until used. Chromatography was performed on 15 cm x 10 cm Sorbfil-A silica gel HPTLC plates (layer thickness 0.1 mm; Imid Ltd), which used without pretreatment. Standard (1 µL) and sample solutions (2 µL) were applied as 8 mm bands, 5 mm apart, 15 mm from the left of the plate and 15 mm from the bottom edge, by means of a APA-1 automatic sample applicator (Imid Ltd). Plates were developed at room temperature (20 ºC) with toluene-EtOAc-HCOOH 5:4:1 (v/v, for flavonol aglycones, caffeic and gallic acids) or EtOAc-1,2-dichloroethane-AcOH-HCOOH-H₂O 10:2.5:1:1:0.8 (v/v, for flavonol glycosides and caffeoylquinic acids) as mobile phases, in a flat-bottomed vertical chamber previously saturated with mobile phase vapor (40 min); the development distance was 6 cm. After development, plates were dried for 10 min in cool air and derivatized with Neu reagent (freshly prepared 0.5% 2-aminoethyl diphenylborinate solution in EtOAc/5% PEG 400 in 1,2-dichloroethane). After derivatization the plates were heated at 100 ºC for 5 min with a TLC Plate Heater USP-1M (Imid Ltd) and then scanned 387 nm by use of a Sorbfil Videodensitometer 2.0 TLC Scanner (Imid Ltd) controlled by Sorbfil software; the slit dimensions were 3 mm x 0.40 mm.

Results and Discussion

The 70% EtOH-extract of Caragana bungei Ledeb., Fabaceae, flowering shoots was partitioned with CHCl₃, EtOAc, and n-BuOH to yield three fractions which were separated by chromatographic column (gel permeation, silica gel and polyamide chromatography) and pTLC, given sixteen compounds. The isolated compounds including seven flavonoids [kaempferol (3), quercetin (4) (Olennikov et al., 2011b), isorhamnetin-3-O-glucoside (9) (Manguro et al., 2004), isoquercitrin (quercetin-3-O-glucoside, 10) (Liu et al., 2010), rutin (quercetin-3-O-rutinoside, 13) (Hou et al., 2005), narcissin (isorhamnetin-3-O-rutinoside, 14) (Lu et al., 2000), nicotiflorin (kaempferol-3-O-rutinoside, 15) (Hou et al., 2005)], four phenylpropanoids [caffeic acid (6) (Olennikov et al., 2011a), 3,5-di-O-caffeoylquinic acid (8) (Satake et al., 2007), 3-O-caffeoylquinic acid (11), 5-O-caffeoylquinic acid (12) (Olennikov et al., 2011a)], two triterpenes [β-sitosterol (1), β-sitosterol-3-O-glucoside (5) (Tomimatsu & Hashimoto, 1968)], umbelliferone (2) (Olennikov & Tankhaeva, 2005a), gallic acid (7) (Olennikov et al., 2005b) and sucrose (16) (Jung et al., 2000) were identified by comparison of their m.p., [α]D, UV, MS and ¹³C-NMR data with those reported in the literature.

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Accordingly to literature data some of isolated compounds were previously detected in other Caragana species: kaempferol in C. spinosa (Shpekina, 1990); quercetin in C. arborescens (Boinik et al., 1986), C. alaica (Butayarov et al., 1999), C. jubata (Umarov et al., 1971), C. pygmaea (Polovinko & Yakovlev, 1985), C. sinica (Tai et al., 2010) and C. spinosa (Shpekina, 1990); isorhamnetin-3-O-glucoside in C. alaica (Butayarov, 1999); isoquercitrin in C. arborescens (Boinik, 1986), C. alaica (Butayarov, 1999), C. sinica (Tai et al, 2010) and C. spinosa (Shpekina, 1990); rutin in C. arborescens (Boinik, 1986), C. aurantiaca (Polovinko & Shostakovskaya, 1987), C. pygmaea (Polovinko & Yakovlev, 1985) and C. spinosa (Shpekina, 1990); narcissin in C. aurantiaca (Polovinko & Shostakovskaya, 1987), C. pygmaea (Polovinko & Yakovlev, 1985), C. sinica (Tai et al, 2010) and C. spinosa (Shpekina, 1990); β-sitosterol in C. conferta (Khan et al., 2009) and C. microphylla (Huo et al., 2007); β-sitosterol-3-O-glucoside in C. conferta (Khan et al., 2009); umbelliferone in C. frutex (Boinik et al., 1983); sucrose in C. microphylla (Huo et al., 2007). The presence of nicotiflorin, caffeic, 3-O-caffeoylquinic, 5-O-caffeoylquinic, 3,5-di-O-caffeoylquinic and gallic acids was firstly shown for Caragana species in this work.

Using HPTLC analysis we were investigated the distribution of phenolic compounds in different C. bungei organs (leaves, flowers, stems). It was shown that leaves were concentrated the flavonoid glucosides and phenylpropanoids and traces of flavonoid aglycones and gallic acid (Table 1). The composition of flowers phenols was similar to those of leaves excluding of kaempferol, nicotiflorin, caffeic, 3,5-di-O-caffeoylquinic and gallic acids. Characteristic feature of stems was ability of outer bark (epidermis) to concentration of quercetin, kaempferol and gallic acid with absence or traces amounts of flavonoids and phenylpropanoids. Only traces of quercetin were detected in inner bark of stems. The values of total phenolic content in leaves, flowers, outer and inner barks of stems were 40.09, 36.24, 12.60 and 1.94 mg/g from dry plant weight respectively, and values of total flavonoid content in leaves, flowers and outer bark of stems were 29.32, 18.67 and 2.13 mg/g from dry plant weight respectively.

The known data about biological activity of plant sterols as a hypocholesterolemic and antiatherosclerotic substances (Lees et al., 1977) was initiated the quantitative analysis of β-sitosterol in C. bungei. It was established that β-sitosterol contents in leaves, flowers, outer and inner barks of stems were 1.39, 4.52, 1.43 and 0.97 mg/g respectively (Table 1). High content of β-sitosterol was allowed to consider C. bungei flowers as a potential raw material for production of hypocholesterolemic preparations.

Conclusions

The phytochemical study of Caragana bungei Ledeb. flowering shoots using in traditional medicine of Tuva and Buryatia Republics (Russian Federation) indicated that this plant species contain flavonols (kaempferol, quercetin, isorhamnetin-3-O-glucoside, isoquercitrin, rutin, narcissin, nicotiflorin), phenylpropanoids (caffeic acid, caffeoylquinic acids), triterpenes (β-sitosterol, β-sitosterol-3-O-glucoside), coumarins (umbelliferone), simple phenols (gallic acid) and carbohydrates (sucrose) and can be regarded as perspective for further pharmacological investigation.

Acknowledgment

The authors acknowledge the financial support provided by the Presidium of Siberian Division of Russian Academy of Science under Project VI.52.1.3.

Received 21 Jun 2011

Accepted 27 Sep 2011

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Publication Dates

• Publication in this collection
  17 Jan 2012
• Date of issue
  June 2012

History

• Received
  21 June 2011
• Accepted
  27 Sept 2011