O-geranyltricin (1) and 3′-O-geranylpolloin (2), and a new 2-(2-phenylethyl)-4H-chromen-4-one derivative, 7-hydroxyl-6-methoxy-2-(2-phenylethyl)chromone (3), have been isolated from the stem barks of A. sinensis, together with 21 known compounds 4–24. The structures of new compounds 1–3 were determined through spectroscopic and MS analyses. Compounds 2, 3, 5, 6, and 8–10 exhibited inhibition (IC50 ≤ 12.51 μM) of superoxide anion generation by human neutrophils in response to formyl-l-methionyl-l-leucyl-l-phenylalanine/cytochalasin B (fMLP/CB). Compounds 3, 6, 8, 10, and 19 inhibited fMLP/CB-induced elastase release with IC50 values ≤ 15.25 μM. This investigation reveals bioactive isolates  could be further developed as potential candidates for the treatment or prevention of various inflammatory diseases.In the current study, two new flavones, 4′- Aquilaria sinensis (Lour.) Gilg. (Thymelaeaceae) is an evergreen tree, distributed in southern China +  and this was supported by the 1H-, 13C-, and DEPT-NMR data. The IR spectrum showed the presence of OH (3408 cm−1) and carbonyl (1657 cm−1) groups. Comparison of the 1H-NMR data of 1 with those of tricin (4) +  in the HRESI mass spectrum. IR absorptions for OH (3412 cm−1) and carbonyl (1660 cm−1) functions were observed. The 1H-NMR data of 2 were similar to 5-hydroxy-7,3′,4′-trimethoxyflavone (5)  of 2 replaced the OMe-3′ [δ3.99 ] of 5. This was supported by HMBC correlation observed between H-1′′ (δH 4.72) and C-3′ (δC 148.5), C-2′′ (δC 119.2), and C-3′′ (δC 141.6), and by NOESY correlations observed between H-1′′ (δH 4.72) and H-2′ (δH 7.36), H-2′′ (δH 5.53), and H-10′′ (δH 1.81). On the basis of the above data, the structure of 2 was elucidated as 3′-O-geranylpolloin, which was further confirmed by 1H-1H COSY and NOESY  , except nd NOESY  experimeand HMBC  techniqu3) was obtained as a colorless prisms. The molecular formula C18H16O4 was deduced from a proton adduct ion peak at m/z 297.11209 [M + H]+  in the HRESI mass spectrum. The presence of hydroxy and carbonyl groups was revealed by the bands at 3418 and 1634 cm−1, respectively, in the IR spectrum. The 1H-NMR spectrum indicated the presence of a 2-phenylethyl group , a methoxy group [δ 4.00 ], a hydroxy group [δ 6.24 ], and three singlet aromatic protons . Comparison of the 1H and 13C-NMR data of 3 with those of 6,7-dimethoxy-2-(2-phenylethyl)chromone (10)  of 3 replaced the OMe-7 [δ 3.97 ] of 10. This was supported by HMBC correlations between OH-7 (δ 6.24) and C-6 (δ 145.1), C-7 (δ 151.2), and C-8 (δ 102.7) and NOESY correlations between OH-7 (δ 6.24) and OMe-6 (δ 4.00). Furthermore, the full assignment of 1H- and 13C-NMR resonances was confirmed by the 1H-1H COSY, NOESY (3 was elucidated as 7-hydroxy-6-methoxy-2-(2-phenylethyl)chromone.7-Hydroxy-6-methoxy-2-(2-phenylethyl)chromone (one (10)  suggesteY, NOESY , DEPT, HY, NOESY  experime1H-NMR, [α]D, and MS) with the corresponding authentic samples or literature values, and included six flavones: tricin (4)  and granule proteases  produced by human neutrophils are involved in the pathogenesis of a variety of inflammatory diseases. The effects on neutrophil pro-inflammatory responses of compounds isolated from the stem barks of A. sinensis were evaluated by suppressing fMet-Leu-Phe/cytochalasin B (fMLP/CB)-induced superoxide anion (O2•−) generation and elastase release by human neutrophils. The inhibitory activity data on neutrophil pro-inflammatory responses are summarized in Human neutrophils are known to play an important role in the host defense against microorganisms and in the pathogenesis of various diseases such as rheumatoid arthritis, ischemia-reperfusion injury, asthma, and chronic obstructive pulmonary disease ,47. In re lipids ,48,49. S2•− generation and elastase release, respectively. From the results of our biological tests, the following conclusions can be drawn: (a) 3′-O-Geranylpolloin (2), 7-hydroxy-6-methoxy-2-(2-phenylethyl)chromone (3), 5-hydroxy-7,3′,4′-trimethoxyflavone (5), velutin (6), 3′-hydroxy-genkwanin (8), sakuranetin (9), and 6,7-dimethoxy-2-(2-phenylethyl)chromone (10) exhibited potent inhibition (IC50 ≤ 12.51 μM) of superoxide anion (O2•−) generation by human neutrophils in response to fMLP/CB; (b) 7-hydroxy-6-methoxy-2-(2-phenylethyl)chromone (3), velutin (6), sakuranetin (9), 6,7-dimethoxy-2-(2-phenylethyl)chromone (10), and ergosta-4,6,8(14)-22-tetraen-3-one (19) exhibited potent inhibition (IC50 ≤ 15.25 μM) against fMLP-induced elastase release; (c) 7-hydroxy-6-methoxy-2-(2-phenylethyl)chromone (3) (with a 7-hydroxy group) exhibited more effective inhibition than its analogue, 6,7-dimethoxy-2-(2-phenylethyl)chromone (10) (with a 7-methoxy substituent) against fMLP-induced O2•− generation and elastase release; (d) among the flavone analogues , velutin (6) (with a 4′-hydroxy-3′-methoxyphenyl moiety) and 5-hydroxy-7,3′,4′-trimethoxyflavone (5)  exhibited more effective inhibition than their analogues, 1, 2, 4, 7, and 8, against fMLP-induced O2•− generation; velutin (6) and 3′-hydroxygenkwanin (8)  exhibited more effective inhibition than their analogues, 1, 2, 4, 5, and 7, against fMLP-induced elastase release; (e) among the flavone analogues , only apigenin 7,4′-dimethyl ether (7) (with a 4′-methoxyphenyl moiety) at 10 μg/mL alone elicited superoxide anion generation and elastase release by human neutrophils in the absence of fMLP/CB; (f) sakuranetin (9) and velutin (6) were the most effective among the isolated compounds, with IC50 values of 1.74 ± 0.17 and 1.78 ± 0.35 μM, against fMLP-induced superoxide anion generation; (g) 7-hydroxy-6-methoxy-2-(2-phenylethyl)chromone (3), velutin (6), and 3′-hydroxygenkwanin (8) were the most effective among the isolated compounds, with IC50 value of 3.91 ± 0.87, 4.26 ± 0.12, and 4.56 ± 0.63 μM, against fMLP-induced elastase release.Diphenyleneiodonium and phenylmethylsulfonyl fluoride were used as positive controls for OO-geranyltricin (1) and 3′-O-geranylpolloin (2), and a new 2-(2-phenylethyl)-4H-chromen-4-one derivative, 7-hydroxy-6-methoxy-2-(2-phenylethyl)chromone (3) were isolated from the stem barks of A. sinensis. Known compounds 9, 12–16, and 19–24 were obtained from this plant for the first time. The structures of these compounds were established on the basis of spectroscopic data. Discovery of the two new flavones and a new 2-(2-phenylethyl)-4H-chromen-4-one derivative from the genus Aquilaria may not only provide more structure-activity data of flavones and 2-(2-phenylethyl)-4H-chromen-4-one, but may also contribute to enhancing our understanding of the taxonomy and evolution of the genus Aquilaria.Twenty four compounds, including two new flavones, 4′-2•−), hydrogen peroxide] and produced by human neutrophils contribute to the pathogenesis of inflammatory diseases. Inhibition of the inappropriate activation of neutrophils by drugs has been proposed as a way to ameliorate inflammatory diseases. Based on the results of our biological tests (O-geranylpolloin (2), 7-hydroxy-6-methoxy-2-(2-phenylethyl)chromone (3), 5-hydroxy-7,3′,4′-trimethoxyflavone (5), velutin (6), 3′-hydroxygenkwanin (8), sakuranetin (9), and 6,7-dimethoxy-2-(2-phenylethyl)chromone (10) were the most effective among these compounds, with IC50 values of 12.51 ± 2.75, 4.62 ± 1.48, 4.69 ± 0.94, 1.78 ± 0.35, 7.96 ± 0.76, 1.74 ± 0.17, and 11.54 ± 2.19 μM, respectively, against fMLP-induced superoxide anion generation. 7-Hydroxy-6-methoxy-2-(2-phenylethyl)chromone (3), velutin (6), 3′-hydroxygenkwanin (8), 6,7-di-methoxy-2-(2-phenylethyl)chromone (10), and ergosta-4,6,8(14),22-tetraen-3-one (19) exhibited the most effective among the isolates, with IC50 values of 3.91 ± 0.87, 4.26 ± 0.12, 4.56 ± 0.63, 10.48 ± 1.35, and 15.25 ± 3.75 μM, respectively, against fMLP-induced elastase release. Our study suggests A. sinensis and its isolates  could be further developed as potential candidates for the treatment or prevention of various inflammatory diseases. More experiments should be performed to deduce the action modes of these compounds.Granule proteases  and reactive oxygen species (ROS) e.g., superoxide anion  by venipuncture, using a protocol (No. 102-1595A3) approved by the Institutional Review Board at Chang Gung Memorial Hospital. All donors gave written consent. The Medical Ethics Committee of Chang Gung Memorial Hospital approved this consent procedure.3. Ultraviolet (UV) spectra were obtained on a UV-240 spectrophotometer (Jasco). Infrared (IR) spectra (neat or KBr) were recorded on a 2000 FT-IR spectrometer . The proton nuclear magnetic resonance (NMR) spectra were recorded on a Varian Unity Plus and Mercury 400 , a Varian Inova 500 and a Varian Unity Plus 600 spectrometer operating at 400, 500 and 600 MHz, the carbon NMR spectra, including correlation spectroscopy (COSY), nuclear Overhauser effect spectrometry (NOESY), heteronuclear multiple-bond correlation (HMBC), heteronuclear single-quantum coherence (HSQC) experiments, were recorded on a Varian Unity Plus 600 spectrometer operating at 600 MHz (1H) and 150 MHz (13C), respectively, with chemical shifts given in ppm (δ) using tetramethylsilane (TMS) as an internal standard. Electrospray ionization (ESI), high-resolution electrospray ionization (HRESI) and electron ionization (EI)-mass spectra were recorded on an APEX II mass spectrometer  and Trace GC/Polaris Q MS . Silica gel   was used for column chromatography (CC). Silica gel 60 F-254 (Merck) was used for thin-layer chromatography (TLC) and preparative thin-layer chromatography (PTLC).Melting points were determined on a Yanaco micro-melting point apparatus  and are uncorrected. Optical rotations were measured using a Jasco DIP-370 polarimeter  in CHClA. sinensis was collected from Pingtung County, Taiwan, in August 2013 and identified by Prof. J.J. Chen. A voucher specimen (AS 201308) was deposited in the Department of Pharmacy, Tajen University, Pingtung, Taiwan.The stem barks of A. sinensis were pulverized and extracted three times with MeOH (20 L each) for 3 days. The MeOH extracts were concentrated under reduced pressure at 35 °C, and the residue (390 g) was partitioned between n-hexane and H2O (1:1). The n-hexane layer was concentrated to give a residue . The water layer was further extracted with EtOAc, and the EtOAc-soluble part  and the water-solubles  were separated. Fraction B (75 g) was chromatographed on silica gel , eluting with n-hexane, gradually increasing the polarity with acetone or MeOH to give 11 fractions: B1 , B2 , B3 , B4 , B5 , B6 , B7 , B8 , B9 , B10 , and B11 . Fraction B1 (5.1 g) was chromatographed further on silica gel  eluting with CH2Cl2/EtOAc (100:1–0:1) to give 10 fractions . Fraction B1-2 (138 mg) was purified further by preparative TLC  to obtain α-tocopherol (21) (5.8 mg) (fR = 0.45). Fraction B1-5 (167 mg) was purified further by preparative TLC  to obtain α-tocospiro A (22) (4.6 mg) (fR = 0.38). Fraction B1-6 (153 mg) was purified further by preparative TLC  to yield β-sitostenone (17) (8.6 mg) (fR = 0.76). Fraction B2 (5.7 g) was chromatographed further on silica gel  eluting with CH2Cl2/EtOAc (80:1–0:1) to give 12 fractions . Fraction B2-3 (405 mg) was purified by MPLC  to afford six subfractions . Fraction B2-3-3 (38 mg) was purified by preparative TLC  to obtain trans-phytol (20) (3.9 mg) (fR = 0.29). Fraction B2-6 (195 mg) was purified further by preparative TLC  to give ergosta-4,6,8(14),22-tetraen-3-one (19) (6.3 mg) (fR = 0.52). Fraction B3 (6.6 g) was chromatographed on silica gel  eluting with CH2Cl2/MeOH (70:1–0:1) to give 10 fractions . Fraction B3-1 (210 mg) was purified further by preparative TLC  to yield tetracosyl trans-ferulate (16) (6.3 mg) (fR = 0.52). Fraction B3-4 (220 mg) was purified further by preparative TLC  to afford β-sitosterol (18) (10.7 mg) (fR = 0.52). Fraction B5 (10.9 g) was chromatographed on silica gel  eluting with CH2Cl2/MeOH (50:1–0:1) to give 12 fractions . Fraction B5-2 (203 mg) was purified further by preparative TLC  to yield 5-hydroxy-7,3′,4′-trimethoxyflavone (5) (8.3 mg) (fR = 0.84). Fraction B5-3 (198 mg) was purified further by preparative TLC  to yield 3′-O-geranylpolloin (2) (4.7 mg) (fR = 0.83). Fraction B5-6 (216 mg) was purified further by preparative TLC  to afford sakuranetin (9) (5.2 mg) (fR = 0.71), 6,7-dimethoxy-2-(2-phenylethyl)chromone (10) (4.6 mg) (fR = 0.62), and taraxacine-A (12) (5.1 mg) (fR = 0.45). Fraction B5-7 (187 mg) was purified further by preparative TLC  to yield tetracosyl caffeate (15) (4.7 mg) (fR = 0.76). Fraction B6 (5.6 g) was chromatographed further on silica gel  eluting with CH2Cl2/acetone (25:1–0:1) to give 8 fractions . Fraction B6-1 (205 mg) was purified further by preparative TLC  to obtain apigenin 7,4′-dimethyl ether (7) (8.7 mg) (fR = 0.67). Fraction B6-3 (169 mg) was purified further by preparative TLC  to obtain 2,6-dimethoxy-1,4-benzoquinone (24) (5.3 mg) (fR = 0.64). Fraction B7 (8.3 g) was chromatographed on silica gel  eluting with CH2Cl2/MeOH (20:1–0:1) to give 11 fractions . Fraction B7-1 (177 mg) was purified further by preparative TLC  to obtain velutin (6) (10.7 mg) (fR = 0.57). Fraction B7-5 (215 mg) was purified further by preparative TLC  to yield 3′-O-geranylpolloin (1) (4.1 mg) (fR = 0.71). Fraction B7-7 (220 mg) was purified further by preparative TLC  to obtain tricin (4) (9.4 mg) (fR = 0.57). Fraction B7-9 (203 mg) was purified further by preparative TLC  to obtain 7-hydroxy-6-methoxy-2-(2-phenylethyl)chromone (3) (5.2 mg) (fR = 0.79). Fraction B8 (6.3 g) was chromatographed on silica gel  eluting with CH2Cl2/MeOH (12:1–0:1) to give 10 fractions . Fraction B8-4 (188 mg) was purified further by preparative TLC  to yield methyl 3,4-dihydroxybenzoate (13) (5.4 mg) (Rf = 0.50). Fraction B8-5 (193 mg) was purified further by preparative TLC  to obtain blumenol (23) (4.9 mg) (fR = 0.69). Fraction B8-6 (213 mg) was purified further by preparative TLC  to yield 3′-hydroxygenkwanin (8) (6.2 mg) (fR = 0.40). Fraction B8-7 (183 mg) was purified further by preparative TLC  to obtain vanillic acid (14) (7.9 mg) (fR = 0.23). Fraction B9 (5.3 g) was chromatographed on silica gel  eluting with CH2Cl2/MeOH (10:1–0:1) to give 9 fractions . Fraction B9-4 (189 mg) was purified further by preparative TLC  to obtain (‒)-syringaresinol (11) (6.8 mg) (fR = 0.48).The dried stem barks (4.1 kg) of 4′-O-Geranyltricin (1). Yellowish needles (CH2Cl2-MeOH); m.p. 256–258 °C; UV (MeOH) λmax (log ε) 207 (4.67), 268 (4.14), 331 (4.21) nm; IR (KBr) νmax 3408 (OH), 1657 (C=O) cm−1; 1H-NMR  δ 1.59 , 1.67 , 1.68 , 2.03 , 2.08 , 3.89 , 3.95 , 4.64 , 5.07 , 5.56 , 6.39 , 6.50 , 6.60 , 7.08 , 12.73 ; 13C-NMR  δ 16.4 (C-10′′), 17.6 (C-9′′), 25.6 (C-8′′), 26.4 (C-5′′), 39.6 (C-4′′), 55.8 (OMe-7), 56.4 (OMe-3′ and OMe-5′), 69.6 (C-1′′), 92.7 (C-8), 98.2 (C-6), 103.8 (C-2′ and C-6′), 105.6 (C-3), 105.6 (C-10), 119.9 (C-2′′), 123.9 (C-6′′), 126.4 (C-1′), 131.7 (C-7′′), 140.4 (C-4′), 142.1 (C-3′′), 154.1 (C-3′ and C-5′), 157.7 (C-9), 162.3 (C-5), 164.0 (C-2), 165.6 (C-7), 182.4 (C-4); ESIMS m/z 481 [M + H]+; HRESIMS m/z 481.22185 [M + H]+ .3′-O-Geranylpolloin (2). Yellowish needles (CH2Cl2-MeOH); m.p. 246–248 °C; UV (MeOH) λmax (log ε) 205 (4.59), 250 (4.20), 268 (4.13), 339 (4.24) nm; IR (KBr) νmax 3412 (OH), 1660 (C=O) cm−1; 1H-NMR  δ 1.58 , 1.64 , 1.81 , 2.10 , 2.12 , 3.89 , 3.95 , 4.72 , 5.07 , 5.53 , 6.37 , 6.48 , 6.56 , 6.97 , 7.36 , 7.51 , 12.80 ; 13C-NMR  δ 16.8 (C-10′′), 17.7 (C-9′′), 25.6 (C-8′′), 26.3 (C-5′′), 39.6 (C-4′′), 55.8 (OMe-7), 56.1 (OMe-4′), 66.2 (C-1′′), 92.7 (C-8), 98.0 (C-6), 104.6 (C-3), 105.6 (C-10), 111.0 (C-2′), 111.4 (C-5′), 119.2 (C-2′′), 120.1 (C-6′), 123.7 (C-1′), 123.7 (C-6′′), 131.9 (C-7′′), 141.6 (C-3′′), 152.9 (C-4′), 148.5 (C-3′), 157.7 (C-9), 164.1 (C-2), 165.5 (C-7), 182.4 (C-4); ESIMS m/z 451 [M + H]+; HRESIMS m/z 451.21133 [M + H]+ .7-Hydroxy-6-methoxy-2-(2-phenylethyl)chromone (3). Colorless prism (CH2Cl2); m.p. 181–183 °C; UV (MeOH) λmax (log ε) 207 (4.69), 226 (4.56), 282 (4.11), 319 (4.15) nm; IR (KBr) νmax 3418 (OH), 1634 (C=O) cm−1; 1H-NMR  δ 2.91 , 3.04 , 4.00 , 6.08 , 6.24 , 6.94 , 7.20 , 7.21 ,7.29 , 7.54  ; 13C-NMR  δ 33.0 (C-7′), 36.0 (C-8′), 56.5 (OMe-6), 102.7 (C-8), 104.4 (C-5), 109.4 (C-3), 117.0 (C-10), 126.5 (C-4′), 128.3 (C-2′ and C-6′), 128.6 (C-3′ and C-5′), 139.8 (C-1′), 145.1 (C-6), 151.2 (C-7), 152.7 (C-9), 167.7 (C-2), 177.6 (C-4); ESIMS m/z 297 [M + H]+; HRESIMS m/z 297.11209 [M + H]+ .The effect of the isolated compounds on neutrophil pro-inflammatory response was evaluated by monitoring the inhibition of superoxide anion generation and elastase release in fMLP/CB-activated human neutrophils in a concentration-dependent manner. The purity of the tested compounds was >98% as identified by NMR and MS.2+)-free HBSS buffer at pH 7.4 and were maintained at 4 °C prior to use.Human neutrophils from venous blood of healthy, adult volunteers (20–28 years old) were isolated using a standard method of dextran sedimentation prior to centrifugation in a Ficoll Hypaque gradient and hypotonic lysis of erythrocytes . Purifiec  and granule proteases  produced by human neutrophils contribute to the pathogenesis of inflammatory diseases. The effects on neutrophil pro-inflammatory responses of isolates were evaluated by suppressing fMLP/CB-induced O2•− generation and elastase release by human neutrophils. The results of anti-inflammatory experiments indicate that compounds 2, 3, 5, 6, 8–10, and 19 can significantly inhibit fMLP-induced O2•− generation and/or elastase release. Sakuranetin (9) and 7-hydroxy-6-methoxy-2-(2-phenylethyl)chromone (3) were the most effective among the isolated compounds, with IC50 values of 1.74 ± 0.17 and 3.91 ± 0.87 μM, respectively, against fMLP-induced O2•− generation and elastase release. Our study suggests A. sinensis and its isolates  are worthy of further biomedical investigation and could be expectantly developed as potential candidates for the treatment or prevention of various inflammatory diseases.Twenty four compounds, including two new flavones, 4′-


Chem. Commun., 2018, DOI: ; 10.1039/c7md00593h.Correction for ‘The critical role of novel benzophenone analogs on tumor growth inhibition targeting angiogenesis and apoptosis’ by Yasser Hussein Eissa Mohammed  The following are the corrected names for compounds 5b: [2-chloro-6-fluoro-4-(4-fluorobenzoyl)phenoxy]acetic acid ethyl ester.5d: [2-chloro-6-fluoro-4-(4-methylbenzoyl)phenoxy]acetic acid ethyl ester.9a: N′-(2-(4-benzoylphenoxy)acetyl)-1-methyl-1H-imidazole-4-carbohydrazide.9b: N′-(2-(2-chloro-6-fluoro-4-(4-fluorobenzoyl)phenoxy)acetyl)-1-methyl-1H-imidazole-4-carbohydrazide.9c: N′-(2-(2-chloro-4-(4-chlorobenzoyl)-6-fluorophenoxy)acetyl)-1-methyl-1H-imidazole-4-carbohydrazide.9d: N′-(2-(2-chloro-6-fluoro-4-(4-methylbenzoyl)phenoxy)acetyl)-1-methyl-1H-imidazole-4-carbohydrazide.10a: N′-(2-(4-benzoylphenoxy)acetyl)-2-oxo-2H-pyran-5-carbohydrazide.10b: N′-(2-(2-chloro-6-fluoro-4-(4-fluorobenzoyl)phenoxy)acetyl)-2-oxo-2H-pyran-5-carbohydrazide.10c: N′-(2-(2-chloro-4-(4-chlorobenzoyl)-6-fluorophenoxy)acetyl)-2-oxo-2H-pyran-5-carbohydrazide.10d: N′-(2-(2-chloro-6-fluoro-4-(4-methylbenzoyl)phenoxy)acetyl)-2-oxo-2H-pyran-5-carbohydrazide.9a was correct in the main manuscript, but wrong in the ESI. The ESI has been corrected.In addition, the structure of The Royal Society of Chemistry apologises for these errors and any consequent inconvenience to authors and readers.


The interactions of 3-formyl-4-chlorocoumarin (3) with nitrogen-containg nucleophiles leading to the corresponding substituted chromen-pyrazol-4-ones 5 are described. The structures of the obtained compounds were established on the basis of 1D NMR, 2D NMR and IR and further the compounds were evaluated for possible antioxidant activities. The coumarinic chalcone 4a has been found to be the most active (IC50 = 2.07 μM) in this study. A series of new coumarin derivatives  Coumarin derivatives constitute an important class of compounds with a wide range of biological activities ,2,3. In 1) and 4-chloro-3-formyl coumarin (3) systems , the method of Dholakia et al. pyrazol-4-ones 5 in short times and in high yields (67 to 89%). The proposed structures of the new substituted chromenpyrazol-4(1H)-ones 5 were validated by their spectral data, which were consistent with available literature data for similar substitution patterns in coumarin derivatives pyrazol-4(1H)-ones 5a–c displayed IR stretching at 1,711–1,742 cm−1 associated with lactone carbonyl groups, also confirmed by the signals appearing between δ 155 and 162 ppm in the 13C-NMR spectra. The derivatives 5a can exist in α, β and γ tautomeric forms, as depicted in The H)-ones nucleus gave poor attention to this tautomerism problem pyrazol-4-ones 5a–c, recorded in chloroform solution, showed lactone bands at 1,704–1,712 cm−1, confirmed by typical carbonyl patterns in the 13C-NMR spectra. 1H- and 13C-NMR analyses show that compounds 5 exist as the tautomeric form containing a proton on the N atom. In our case the β tautomer can be ruled out because all the IR spectra of the chromenpyrazol-4 which is currently used in the interpretation of experimental data from the method.One such method that is currently popular is based upon the use of the stable free radical diphenylpicrylhydrazyl (DPPH). The purpose of our study was to examine the use of the parameter “EC• and the donor molecule by AH, the primary reaction is: • is free radical produced in this first step. This latter radical will then undergo further reactions which control the overall stoichiometry, that is, the number of molecules of DPPH reduced (decolorized) by one molecule of the reductant.When a solution of DPPH is mixed with that of a substance that can donate a hydrogen atom, this gives rise to the reduced form with the loss of this violet colour . Representing the DPPH radical by Z• is thus intended to represent the free radicals formed in the system whose activity is to be suppressed by the substance AH• Representing the DPPH radical by Z• and the coumarinic chalcones by ROH, the initial reaction is then Z• + ROH= ZH+ RO• -2(H)-chromen-2-one (4b). Solid (yield 85%), mp = 182 °C, IR: ν 3168 (-OH), 1622 (>C=O), 1577 (C=C), 1028(s) (sym) (C-O-C); 1H-NMR: δ (ppm) 6.26–7.84 , 19.58 ; 13C-NMR (ppm): 191.2 (CO); 172.8 (C4); 172.1 (C2); 102.2–168.8 (Carom+Cethyl). C18H11O4Br calc. C 58.22, H 2.96, O 17.25, found C 58.30, H 2.90, O 17.30.3-((2E)-3-prop-2-enoyl)-2(H)-chromen-2-one (4c). Solid (yield 83%), mp = 193 °C, IR: ν 3368 (-OH), 1716(s) (>C=O), 1577 (C=C), 1018(s) (sym) (C-O-C); 1H-NMR: δ (ppm) 7.30 , 7.51  6.90–7.96 , 19.50 ; 13C-NMR (ppm): 191.6 (CO); 59.7 (O-CH3); 59.8 (O-CH3); 60.0 (O-CH3) 103.47–153.3 (Carom); 164.8 (C4); 163.2 (C2); 102.8 (C3). C21H18O7 calc. C 65.96, H 4.71, O 29.31; found C 65.90, H 4.80, O 29.30.3-[(2E)-3-(Naphtyl)prop-2-enoyl]-2(H)-chromen-2-one (4d). Solid (yield 80%), mp = 185 °C, IR: ν 3365 (-OH), 1718(s) (>C=O), 1578 (C=C), 1019(s) (sym) (C-O-C); 1H-NMR: δ (ppm) 7.30 , 7.51 , 6.75–7.92 , 19.40 ; 13C-NMR (ppm): 192.6(CO); 102.47–154.2 (Carom); 164.6(C4); 164.2 (C2); 101.8 (C3). C22H14O4 calc. C 77.20, H 4.10, O 18.71; found C 77.10, H 4.10, O 18.70.3-((2E)-3-(4-Dimethylamino-phenyl)prop-2-enoyl)-2(H)-chromen-2-one (4e). Solid (yield 85%), mp = 192 °C, IR: ν 3125 (-OH), 1712(s) (>C=O), 1550 (C=C), 1028 (sym) (C-O-C); 1H-NMR: δ (ppm) 2.9 , 6.54–8.1 , 18.62 ; 13C-NMR (ppm): 191.0 (CO); 40.1 (CH3-N); 102.8–142.9 (Carom); 179.2 (C4); 162.2 (C2); 102.8 (C3). C20H17O4N calc. C 71.64, H 5.1, O 19.10, N 4.17; found C 71.60, H 5.10, O 19.30, N 4.20.3  dissolved on boiling in ethanol (10 mL) and was then cooled to 15–20 °C. A solution prepared from aryl or alkylhydrazine hydrochloride  and triethylamine (4 mmol) in 60%) ethanol (10 mL) was slowly added dropwise so that the temperature of the reaction mixture did not exceed 25 °C. After addition of the reagent was complete a yellowish precipitate formed rapidly. The precipitate was filtered off and recrystallized from ethanol and then twice more from DMF to give fine colourless needlesThe aldehyde 1H-Chromenopyrazol-4-one (5a). Solid (Yiled 67%), mp = 183 °C; IR: ν 3368 (-NH), 1716(s) (lactone >C=O), 1577 (C=C), 1018(s) (sym) (C-O-C); 1H-NMR: δ (ppm) 7.40 , 7.13–7.46 , 13.2 ; 13C-NMR (ppm): 158.2 (C2); 102.1 (C3); 149.2 (C4); 146.7 (C=N); 122.1–151.2 (Carom). C10H5O2N2 calc. C 64.86, H 2.70, O 17.30, N 15.13; found C 64.80, H 2.70, O 17.20, N 15.2.1-Methyl-1H-chromenopyrazol-4-one (5b). Solid(Yiled 75%), mp = 184 °C; IR: ν 1720(s) (lactone >C=O), 1578 (C=C), 1020(s) (sym) (C-O-C); 1H-NMR: δ (ppm). 3.5 , 6.4 ; 7.13–7.45 , 13C-NMR (ppm): 157.2 (C2); 112.2 (C3); 144.6 (C4); 140.6 (Céthyl); 122.1–151.2 (Carom). C11H7O2N2 calc. C 60.53, H 6.27, O 33.20; found C 60.50, H 6.30, O 33.20.1-Phenyl-1H-chromenopyrazol-4-one (5c). Solid (Yiled 89%), mp = 185 °C; IR: ν 1718(s) (lactone >C=O), 1579 (C=C), 1022(s) (sym) (C-O-C); 1H-NMR: δ (ppm): 7.13–7.45 , 7.2 (Héthy) 13C-NMR (ppm): 158.2 (C2); 112.4 (C3); 144.2 (C4); 138.6 (Céthyl); 122.1–151.2 (Carom). C16H10O2N2 calc. C 58.89, H 3.06, O 9.81, N 8.58; found, C 58.90, H 3.10, O 9.80, N 8.60.et al. [32] was used. Half a millilitre of each sample concentration was mixed with an equal volume of DPPH ethanolic solution. After incubation for 30 minutes in darkness at 25 °C, absorbance was read at 520 nm wavelength. A mixture of DPPH solution (0.5 mL) and ethanol (0.5 mL) was taken as a blank . Inhibitory concentration (IC50) values denoting the concentration  required to scavenge 50% of DPPH radicals were calculated. All measurements were performed in triplicate. Results were expressed in inhibition percentage versus sample concentrations (mg/mL) at 30 minutes.The DPPH radical scavenging capacity of the obtained coumarinic derivatives was measured from the bleaching of purple colored ethanol solution of DPPH. The method described by Hatano 5 in good yields. The flexibility of this methodology should provide access to many variously substituted coumarinyl pyrazoles. Our synthesis of coumarinic derivatives allowed us to obtain and identify 12 compounds. The presence of the coumarin nucleus endows this species with important pharmacological and therapeutic interest. The antioxidant activities of these compounds against the stable free radical DPPH show that these species are good source of compounds that could help to increase the overall antioxidant capacity of an organism.In summary, we developed a new and simple method for the synthesis of new coumarinyl chalcone derivatives through a condensation reaction. Our improved synthesis of coumarinyl chalcones was effective in terms of time and yields. The reaction conditions are simple, since they are not sensitive to oxygen and water, which make it easy to operate at room temperature. We have shown a new synthetic method by which to synthesize substituted chromenpyrazol-4-ones


Acetyl groups were introduced using acetic anhydride and microwave irradiation. The introduction of one or two acetoglycoside moieties in the framework of 3,7-dihydroxyflavone (4) was performed using two synthetic methods: the Michael reaction and the Koenigs-Knorr reaction. The in vitro cell growth inhibitory activity of compounds 5, 6, 7, 9, and 10 was investigated in six human tumor cell lines: A375-C5 , MCF-7 (breast adenocarcinoma), NCI-H460 , U251 (glioblastoma astrocytoma), U373 (glioblastoma astrocytoma), and U87MG (glioblastoma astrocytoma). The new flavonoid 3-hydroxy-7- flavone (10) was the most potent compound in all tumor cell lines tested, with GI50 values < 8 μM and a notable degree of selectivity for cancer cells.Natural flavonoids and xanthone glycosides display several biological activities, with the glycoside moiety playing an important role in the mechanism of action of these metabolites. Herein, to give further insights into the inhibitory activity on cell growth of these classes of compounds, the synthesis of four flavonoids ( Pr. Pr5) waon times . The syn2) and mangiferin (3). Diosmin peracetate (6) was successfully obtained in only 35 min (62% yield) in the presence of NaF, under microwave (MW) irradiation (3) was achieved in only 15 min (78% yield) under MW irradiation, but only when iodine was used as a catalyst  ancatalyst .6 and 7 used highly toxic pyridine as solvent and catalyst, which was applied room temperature, leading to long reaction times -4H-chromen-4-one (5) ; mp 119–120 °C (petroleum ether 60–80 °C).Synthesis of diosmin peracetate (6). Diosmin  and NaF  were mixed in acetic anhydride (4 mL) and the mixture was kept under MW irradiation (400 W) at 130 °C, for 35 min. After cooling, the solution obtained was poured into ice and then extracted with CH2Cl2. The organic layer was extracted with NaHCO3 followed by crystallization from MeOH/H2O provided 5,3-O-acetyl-2-(3-O-acetyl-4-methoxyphenyl)-7- oxychromen-4-one (6) as a yellow solid ; mp 135–138 °C.Synthesis of mangiferin peracetate (7). Mangiferin  and iodine  were mixed in acetic anhydride (7 mL) and the reaction was kept under MW irradiation (400 W), at 130 °C, for 15 min. After cooling, a saturated solution of sodium thiosulfate was added to convert iodine (dark yellow) into iodide (yellow). The crude product was extracted with CH2Cl2 and the organic layer was extracted with a saturated solution of NaHCO3 twice, dried with anhydrous Na2SO4, and filtered. The solvent was evaporated under reduced pressure and the oil obtained was dissolved in ethyl acetate. A yellow solid was obtained with petroleum ether 60–80 °C corresponding to 1,3,6,7-tetra-O-acetyl-2-C--9H-xanthen-9-one (7) ; mp 143–147 °C (petroleum ether 60–80 °C).Synthesis of 3,7- flavone (9) and 3-hydroxy-7- flavone (10). 3,7-Dihydroxyflavone  and K2CO3  were mixed in dry acetone (10 mL) and the mixture was kept under stirring for 15 min. 2,3,4,6-Tetra-O-acetyl-α-d-glucopyranosyl bromide  was added and the mixture was kept under stirring at room temperature, for 48 h. The suspension was filtered to eliminate the K2CO3 and the filtrate was evaporated. The oil obtained was further purified through flash column chromatography (100% CHCl3). The obtained fractions were gathered and crystallized from MeOH to achieve 3,7- flavone  and 3-hydroxy-7- flavone .3,7- flavone (9). Mp 105–108 °C; IR (KBr) υmax: 2962, 2917, 2843, 1750, 1623, 1449, 1381, 1231, 1066, 1040 cm−1; 1H NMR  δ: 8.17 , 8.02–7.98 , 7.50–7.47 , 7.10 , 7.05 , 5.71 , 5.34–5.04 , 4.31–4.19 , 3.98–3.91 , 3.65–3.60 , 2.09 , 2.08 , 2.06 , 2.05 , 2.04 , 2.01 , 2.00 , 1.90  ppm; 13C NMR  δ: 173.4 (C-4), 170.6 (COCH3), 170.5 (COCH3), 170.3 (COCH3), 170.2 (COCH3), 170.0 (COCH3), 169.7 (COCH3), 169.5 (COCH3), 169.4 (COCH3), 160.9 (C-7), 157.4 (C-2), 156.6 (C-9), 136.3 (C-3), 131.0 (C-1′), 130.6 (C-4′), 129.1 , 128.3 , 127.6 (C-5), 119.9 (C-10), 115.5 (C-6), 104.4 (C-8), 98.9 (C-glucose), 98.5 (C-glucose), 72.9 (C-glucose), 72.6 (C-glucose), 72.6 (C-glucose), 71.8 (C-glucose), 71.8 (C-glucose), 68.4 (C-glucose), 68.2 (C-glucose), 62.1 (C-glucose), 61.5 (C-glucose), 21.0 (COCH3), 20.8 (COCH3), 20.8 (COCH3), 20.7 (COCH3), 20.7 (COCH3) ppm; HRMS (ESI+) m/z calcd for C29H28O13 [M + H] 915.25535, found 915.25410.3-Hydroxy-7- flavone (10). Mp 222–223 °C; IR (KBr) υmax: 3442, 2961, 2917, 2847, 1758, 1621, 1572, 1501, 1457, 1235, 1211, 1071 cm−1; 1H NMR  δ: 9.62 (3-OH), 8.24-8.18 , 7.57–7.48 , 7.13 , 7.06 , 5.36–5.16 , 4.33–4.20 , 4.03–3.97 , 2.09 , 2.08 , 2.07 , 2.06  ppm; 13C NMR  δ: 172.9 (C-4), 170.6 (COCH3), 170.3 (COCH3), 169.6 (COCH3), 169.4 (COCH3), 160.9 (C-7), 156.8 (C-9), 138.4 (C-3), 131.1 (C-1′), 130.4 (C-4′), 128.8 , 127.7 , 127.4 (C-5), 116.6 (C-10), 115.4 (C-6), 104.3 (C-8), 98.4 (C-1″), 72.6 (C-3″), 72.6 (C-5″), 71.1 (C-2″), 68.3 (C-4″), 62.1 (C-6″), 20.8 (COCH3), 20.8 (COCH3), 20.7 (COCH3), 20.7 (COCH3) ppm; HRMS (ESI+) m/z calcd for C29H28O13 [M + H] 585.16027, found 585.15875.2 humidified atmosphere . Cell viability was routinely determined with Trypan Blue (Sigma-Aldrich) exclusion assay and all experiments were performed with exponentially growing cells, showing more than 95% viability.The human tumor cell lines A375-C5 (melanoma), MCF-7 (breast adenocarcinoma), and NCI-H460  were grown in culture medium RPMI-1640 , U251 (glioblastoma astrocytoma), U373 (glioblastoma astrocytoma), and U87MG (glioblastoma astrocytoma) were grown in culture medium DMEM (Biochrom), both supplemented with 5% heat-inactivated fetal bovine serum . All cell lines were maintained at 37 °C in a 5% COm/v) trichloroacetic acid , washed with distilled water and stained with Sulforhodamine B  for 30 min at room temperature. The SRB-stained cells were washed 5 times with 1% (v/v) acetic acid (Merck Millipore) and left to dry at room temperature. SRB complexes were solubilized, by adding 10 mM Tris buffer (Sigma-Aldrich) for 30 min. Absorbance was measured at 515 nm in a microplate reader . A dose–response curve was obtained for each cell line with each test compound, and the concentration that caused cell growth inhibition of 50% (GI50) was determined.The effect of compounds on the cell growth was evaluated according to the procedure adopted by the National Cancer Institute (NCI) in the ‘In vitro Anticancer Drug Discovery Screen’, which uses the protein-binding dye sulforhodamine B (SRB) to assess cell growth. Cells were plated in 96-well plates (0.05 × 106 cells/well) in complete culture medium and incubated at 37 °C. Twenty-four hours later, cells were treated with two-fold serial dilutions of the test compounds, ranging from 0 to 150 µM, for 48 h. Control groups received the same amount of sterile Dimethyl Sulfoxide , used as compounds solvent, up to 0.25% concentration. Then, cells were fixed in situ with 50%  over GI50 in cancer cells. An SI value less than 2.0 indicates the general toxicity of the compound [The degree of selectivity of the compounds was assessed by determining the selectivity index (SI) as the ratio of GIcompound .5, 6, 9, and 10), and one xanthonoside (7) were synthesized. As far as we know, compounds 9 and 10 are described for the first time. Compound 5 was synthesized without pyridine in a shorter reaction time. Applying MW irradiation to the synthesis of compounds 6 and 7 allowed to shorten the reaction times.In this work, four acetylated flavonosides  flavone (10) was the most active compound in all the human tumor cell lines tested with notable selectivity for cancer cells. The mechanism of action of this promising compound deserves to be further explored. The position of the sugar moiety seems to influence the activity of these compounds.The in vitro growth inhibitory activity of compounds These results are a starting point from which to investigate more stable chemical modifications in order to optimize such molecules and develop detailed structure-activity relationship studies in future.


O-(threo-β-guaiacylglyceryl) ether 7''-O-β-d-glucopyranose (4) and tricin-4'-O-(erythro-β-guaiacylglyceryl) ether 7''-O-β-d-glucopyranose (5) were isolated from the roots of Zizania latifolia, together with tricin-7-O-β-d-glucopyranose (1), tricin-4'-O-(threo-β-guaiacylglyceryl) ether 7-O-β-d-glucopyranose (2), and tricin-4'-O-(erythro-β-guaiacylglyceryl) ether 7-O-β-d-glucopyranose (3). Their structures were identified on the basis of spectroscopic techniques, including HR-ESI/MS, 1D-NMR , 2D-NMR , and IR spectroscopy.Two new flavonolignan glycosides, tricin-4'- Zizania latifolia Turcz (wild rice) is cultivated in various regions of Southeastern Asia, including Korea, Japan, and China. Particularly, in China, wild rice grain is used as an important traditional medicine to treat anemia and fever in addition to heart, kidney, and liver disorders. It has been reported that wild rice species possess rich nutrient content , confirming that compound 4 was a flavonoid with a phenylpropanoid moiety and hexose moiety. In the low-magnetic field region, a conjugated ketone carbon δ 183.9 (C-4), seven oxygenated olefine quaternary carbons , two olefine quaternary carbons , and five olefin methine carbons  signals were observed, indicating the presence of a flavone moiety. In addition, two oxygenated olefine quaternary carbons , one olefine quaternary carbon δ 133.1 (C-1''), and three olefine methine carbons , two oxygenated methine carbons  and one oxygenated methylene carbon δ 62.7 (C-9'') signals were observed, indicating the presence of a phenylpropanoid moiety. In the HMBC spectrum, the oxygenated methine proton δH 5.19  showed a correlated cross peak with an oxygenated olefine quaternary carbon δC 141.3 (C-4′), indicating that the flavone and phenylpropanoid moieties were linked between δC C-8'' and δC C-4' via an ether linkage, as well as a cross-peak between an anomer proton signal at δH 5.54  and an oxygenated methine carbon signal at δC 82.6 (C-7''), indicating that glucose was attached to the C-7'' of the phenylpropanoid through a glycosidic likage (4) was determined to be tricin-4'-O-(threo-β-guaiacylglyceryl) ether 7''-O-β-d-gluco-pyranose, which is salcolin A 7''-O-β-d-glucopyranose. The molecular weight was determined to be 688.2003 from the pseudomolecular ion peak at m/z 687.1962 [M−H]− in the negative high resolution ESI-MS .Compound ectively . The 13Cc likage . Finally1H-NMR  and 13C-NMR  spectra of compound 5 were almost the same as those of compound 4, with the exception of the relative stereostructure of C-7'' and C-8''. Compound 5 was confirmed to be erythro type from the coupling constant between the oxygenated methine signals of H-7'' and H-8'' (J = 4.8 Hz). The ether linkage (C-8''/C-4') and glycosidic likage (C-1'''/C-7'') were confirmed from the cross peak between the oxygenated methine proton δH 5.16  and oxygenated olefine quaternary carbon δC 142.0 (C-4'), as well as the cross-peak between the anomer proton signal at δH 5.35  and a oxygenated methine carbon signal at δC 81.9 (C-7'') in the HMBC spectrum. Consequently, the structure of compound 5 was determined to be tricin-4'-O-(erythro-β-guaiacylglyceryl) ether 7''-O-β-d-glucopyranose, which is salcolin B 7''-O-β-d-glucopyranose. The molecular weight was determined to be 688.2003 from the pseudomolecular ion peak at m/z 687.1952 [M−H]− in the negative high resolution ESI-MS .Signals in the 254 plate (Merck) and a RP-18 F254s plates (Merck), and detection was performed by a UV lamp at 254 and 365 nm and 10% H2SO4 solution by spraying and heating. 1H and 13C-NMR along with 2D-NMR data were obtained on a Bruker AVANCE II 400  spectrometer  in pyridine-d5 with TMS as internal standard. ESI/MS was obtained on a AB SCIEX triple TOF 5600 mass spectrometer . All the reagent grade chemicals were purchased from Sigma Chemical Co. .Preparative-liquid chromatography (prep-LC)  was carried out using cartridge YMC-DispoPack . Silica gel  and Sephadex LH-20 (Merck) were used for column chromatography. Thin–layer Chromatography (TLC) analysis was performed using on a Kiesel gel 60FPlant sample. Dried aerial parts of Z. latifolia was purchased from store located at Yeongcheon City, Gyeongbuk, Korea. A voucher specimen (KHU-0130301) was deposited at the Laboratory of Natural Products Chemistry, Kyung Hee University, Yongin, Korea.2O extract . The EtOAc extract was applied to a silica gel c.c. (12 × 14 cm), and eluted with CHCl3 : MeOH  monitoring by thin layer chromatography (TLC) to produce 13 fractions (ZLE-1 to ZLE-13). Fraction ZLE-12 (5.0 g) was combined and subjected to MPLC column chromatography: YMC-DispoPack AT . The fractions were analyzed by a UV detector at 254 and 356 nm, and the mobile phase consisted of A: 0.1% formic acid in water, B: 0.1% formic acid in acetonitrile with a flow rate of 10 mL/min. The gradient consisted of 10% B held for 20 min to 30% B at a rate of 0.25%/min, from 30% B to 50% B at a rate of 0.20%/min, from 50% B to 70% B at a rate of 0.25%/min, and from 70% B to 90% B at a rate of 0.30%/min. A total 11 fractions (ZLE-12-1 to ZLE-12-15) were obtained. ZLE-12-10 (642 mg) was subjected to MPLC column chromatography: YMC-DispoPack AT , with A: 0.1% formic acid in water, B: 0.1% formic acid in acetonitrile as the mobile phase at a flow rate of 5 mL/min. The detector was maintained at 254 and 356 nm. The gradient consisted of 20% B to 50% B at a rate of 0.5%/min, and 50% B to 70% B at a rate of 0.6%/min. Compound 1 , compound 2 , compound 3 , compound 4  and compound 5  were obtained.Dried powder (5 kg) was extracted at room temperature with 80% MeOH (45 L). The concentrated extract was partitioned with water (3 L), EtOAc (3 L) and n-BuOH (3 L) successively to produce. EtOAc extract , n-BuOH extract  and Hd-glucopyranoseTricin-7-O-β- (1). Pale yellow powder (CH3OH); negative ESI/MS m/z 491 [M−H]−; IR (KBr) 3380, 2921, 1730, 1652, 1613, 1491,1358, 1084 cm−1; 1H-NMR (pyridine-5d) δ 7.55 , 7.17 , 7.02 , 6.86 , 5.78 , 4.55 , 4.38-4.10 , 3.87 . 13C-NMR (pyridine-5d) δ 182.8 (C-4), 164.9 (C-2), 164.0 (C-7), 162.5 (C-5), 157.8 (C-9), 149.5 , 142.2 (C-4'), 121.0 (C-1'), 106.5 (C-10), 105.1 , 104.5 (C-3), 101.7 (C-1''), 100.6 (C-6), 95.5 (C-8), 79.2 (C-3''), 78.4 (C-5''), 74.7 (C-2''), 71.0 (C-4''), 62.2 (C-6''), 56.5 .Tricin-4'-O-(threo-β-guaiacylglyceryl) etherd-glucopyranose7-O-β- (2). Yellow amorphous powder (CH3OH); Negative ESI/MS m/z 687 [M−H]−; IR (KBr) 3369, 2938, 1652, 1611, 1588, 1512, 1495, 1356, 1263, 1159, 1124, 841 cm−1; 1H-NMR (pyridine-5d) δ 7.53 , 7.37 , 7.31 , 7.26 , 7.14 , 7.01 , 6.85 , 5.76 , 5.70 , 5.08 , 4.65 , 4.54 , 4.28 , 4.36-4.27 , 3.84 , 3.74 . 13C-NMR (pyridine-5d) δ 184.1 (C-4), 165.5 (C-7), 165.4 (C-2), 163.7 (C-5), 159.1 (C-9), 155.3 , 149.6 (C-3''), 148.6 (C-4''), 141.9 (C-4'), 135.6 (C-1''), 127.7 (C-1'), 121.8 (C-6''), 117.3 (C-5''), 112.9 (C-2''), 107.8 (C-10), 107.1 (C-3), 106.1 , 103.0 (C-1'''), 102.1 (C-6), 96.8 (C-8), 89.4 (C-8''), 80.5 (C-3''′), 79.7 (C-5'''), 76.0 (C-4'''), 75.1 (C-7''), 72.4 (C-2'''), 63.5 (C-6'''), 62.9 (C-9''), 57.7 , 57.1 (C-3''-OCH3).Tricin-4'-O-(erythro-β-guaiacylglyceryl) etherd-glucopyranose7-O-β- (3). Yellow amorphous powder (CH3OH); Negative ESI/MS m/z 687 [M−H]−; IR (KBr) 3364, 2933, 1649, 1607, 1590, 1495, 1457, 1358, 1121, 832 cm−1; 1H-NMR (pyridine-5d) δ 7.57 , 7.46 , 7.29 , 7.26 , 7.13 , 6.99 , 6.86 , 5.77 , 5.76 , 4.95 , 4.54 , 4.43 , 4.37–4.10 , 4.06 , 3.82 , 3.77 . 13C-NMR (pyridine-5d) δ 183.5 (C-4), 165.0 (C-7), 164.9 (C-2), 163.3 (C-5), 158.6 (C-9), 154.6 , 149.1 (C-3''), 148.2 (C-4''), 141.9 (C-4'), 134.6 (C-1''), 127.2 (C-1'), 121.4 (C-6''), 116.8 (C-5''), 112.6 (C-2''), 107.4 (C-3), 106.6 (C-10), 105.9 , 105.7 (C-1'''), 101.3 (C-6), 96.3 (C-8), 87.8 (C-8''), 80.0 (C-3'''), 79.2 (C-5'''), 75.5 (C-4'''), 74.4 (C-7''), 71.9 (C-2'''), 63.9 (C-6'''), 62.5 (C-9''), 57.6 , 57.1 (C-3''-OCH3).Tricin-4'-O-(threo-β-guaiacylglyceryl) etherd-glucopyranose7''-O-β- (4). Yellow amorphous powder (CH3OH); HRESI/MS m/z 687.1962 [M−H]−; IR (KBr) 3406, 1639, 1614, 1495, 1358, 1121 cm−1; 1H-NMR (pyridine-5d) and 13C-NMR (pyridine-5d), see Tricin-4'-O-(erythro-β-guaiacylglyceryl) etherd-glucopyranose7''-O-β- (5). Yellow amorphous powder (CH3OH); HRESI/MS m/z 687.1952 [M−H]−; IR (KBr) 3432, 2502, 1685, 1510, 1420, 1399, 1112 cm−1; 1H-NMR (pyridine-5d) and 13C-NMR (pyridine-5d), see Zizania latifolia using a prep-LC instrument equipped with cartridges of various sizes and were identified based on spectroscopic data analysis, including NMR, UV, and ESI-MS. Four flavonolignan glycosides, including two new flavonolignan glycosides, showed structural similarities in being derived from tricin-7-O-β-d-glucopyranose (1), and were shown to be diastereomers of each other. Tricin-4'-O-(threo-β-guaiacylglyceryl) ether 7-O-β-d-glucopyranose (2) and tricin-4'-O-(erythro-β-guaiacylglyceryl) ether 7-O-β-d-glucopyranose (3) were shown to be diastereomers distinguished by threo and erythro configurations, whereas tricin-4'-O-(threo-β-guaiacylglyceryl) ether 7''-O-β-d-glucopyranose (4) and tricin-4'-O-(erythro-β-guaiacylglyceryl) ether 7''-O-β-d-glucopyranose (5) were shown to be threo and erythro, respectively.Five compounds were isolated from the aerial parts of


Satisfactory results were obtained, with excellent yields and a simple experimental procedure. The catalyst could be recycled and reused up to three times without any noticeable decrease in the catalytic activity.The Brønsted acid ionic liquid [PyN(CH ILs have gone far beyond the role of solvent, showing their significant importance as new catalysts in controlling the reactions in which they participate ,19. Rececatalyst .2)4SO3H][p-CH3PhSO3] under different reaction conditions, the reaction of indole and chalcone was selected as a model reaction. The results are presented in 2)4SO3H][p-CH3PhSO3] required (2)4SO3H][p-CH3PhSO3], the reaction did not yield the desired product (2)4SO3H][p-CH3PhSO3], indole and chalcone at 80 °C for 4 h using acetonitrile as solvent.To evaluate the effect of the catalyst [PyN4SO3H][p-CH3PhSO3] could be recovered easily and directly reused in subsequent runs. As shown in b). This indicated that the ionic liquid [PyN(CH2)4SO3H][p-CH3PhSO3] was an efficient and recyclable catalyst for the conjugated addtion of indoles to α, β-unsaturated ketones.When optimizing the reaction conditions, the recycling of the catalyst in the reaction of indole and chalcone was investigated. After the separation of products, ethyl acetate was added to the filtrate, which split into two layers. The upper ethyl acetate layer contained unreacted raw material. The lower aqueous layer included the ionic liquid. The lower layer was concentrated on a rotary evaporator at 80 °C for 1 h under vacuum to remove excess solvent. [PyN4SO3H][p-CH3PhSO3] catalyst under the optimized experimental conditions to afford the corresponding β-indolylketones in excellent yields. 4SO3H][p-CH3PhSO3] coordinates with oxygen atom of α,β-unsaturated ketones (2) to give intermediate 4. The electron rich β-position of the indole ring then attacks the electron deficient conjugated carbon-carbon double bond of 4 to afford 5, followed by a hydrogen transfer to yield 6. Finally 6 rearranged to give target compound 3 and [PyN(CH2)4SO3H][p-CH3PhSO3] catalyzes the next cycle.A plausible mechanism for the conjugate addition of indoles to chalcones is proposed in 2)4SO3H][p-CH3PhSO3] as an efficient catalyst for the synthesis of β-indolylketones. The procedure reported here has the advantages of mild reaction conditions, high yields of products, operational simplicity and catalyst recyclability.In conclusion, we have described for the first use of the Brønsted acid ionic liquid [p-CH3PhSO3] was synthesized according to previous literature [p-CH3PhSO3] (0.2 mmol) was refluxed at 80 °C in acetonitrile (10 mL) for 4 h with stirring. The completion of the reaction was monitored by TLC. After cooling, the reaction mixture was poured onto crushed ice (30 g). The resulting precipitate was filtered under suction, and then recrystallized from ethanol to afford the pure product. The results are summarized in Tables 2. All products  are known compounds, which were characterized by mp, IR, 1H-NMR spectra, elemental analyses.A mixture of indole or substituted indole (2 mmol), 3-(4-Chlorophenyl)-3-(1H-indol-3-yl)-1-(4-methoxylphenyl)propan-1-one (3e): light yellow powder. Mp 202-205 °C; 1H-NMR δ: 3.74 , 3.83 , 3.86 , 4.85 , 6.88-8.00 , 10.83 ; IR : 3383, 2971, 1657, 1599, 1489, 1256, 1180, 742; Anal. calcd. (%) for C24H20NO2Cl: C 73.94, H 5.17, N 3.59. Found: C 73.79, H 5.13, N 3.67.1-(4-Chlorophenyl)-3-(1H-indol-3-yl)-3-(4-methylphenyl)propan-1-one (3g): light yellow powder. Mp 146-149 °C; 1H-NMR δ: 2.19 , 3.77 , 3.88 , 4.80 , 6.85-8.03 , 10.83 ; IR : 3418, 2914, 1674, 1587, 1455, 1244, 1179, 741; Anal. Calcd. (%) for C24H20NOCl: C 77.10, H 5.39, N 3.75. Found C 77.27, H 5.34, N 3.68.3-(4-Chlorophenyl)-1-(4-methoxylphenyl)-3-(2-methyl-1H-indol-3-yl)propan-1-one(3i): brown powder. Mp 169-172 °C; 1H-NMR δ: 2.38 ; 3.81 ; 3.87 , 3.90 , 4.88 , 6.82-8.19 , 10.74 ; IR : 3312, 1664, 1596, 1464, 1258, 1169, 748; Anal. Calcd. (%) for C25H22NO2Cl: C 74.34, H 5.49, N 3.47. Found C 74.45, H 5.43, N 3.52.1-(4-Methoxylphenyl)-3-(2-methyl-1H-indol-3-yl)-3-(4-methylphenyl)propan-1-one (3j): light yellow powder. Mp 172-175 °C; 1H-NMR δ: 2.2 ; 2.36 ; 3.80 ; 3.84 , 3.86 , 4.85 , 6.81-7.94 , 10.68 ; IR : 3338, 1663, 1595, 1463, 1260, 1168, 747; Anal. Calcd. (%) for C26H25NO2: C 81.43, H 6.57, N 3.65. Found C 81.29, H 6.51, N 3.73.1,3-Diphenyl-3-(5-methoxyl-1H-indol-3-yl)propan-1-one (3m): white powder. Mp 150-153 °C; 1H-NMR δ 3.67 , 3.80 , 3.90, 4.82 , 6.66-8.02 , 10.68 ; IR : 3365, 1678, 1597, 1485, 1214, 1169, 723; Anal. Calcd. (%) for C24H21NO2: C 81.10, H 5.96, N 3.94. Found C. 80.93, H 6.01, N 3.90.


The microbial metabolites of 1 were formed by epoxidation or hydroxylation of the prenyl group, and HPLC, NMR, and CD analyses revealed that all of the products were diastereomeric pairs composed of (2S)- and (2R)- isomers. The structures of these metabolic compounds were elucidated to be - and -4′-hydroxy-5-methoxy-7,8--flavanones (2 and 3), (2S)- and (2R)-7,4′-dihydroxy-5-methoxy-8--flavanones (4 and 5) which were new oxygenated derivatives, along with (2R)- and (2S)-4′-hydroxy-5-methoxy-2″-(1-hydroxy-1-methylethyl)dihydrofuroflavanones (6 and 7) on the basis of spectroscopic data. These results could contribute to understanding the metabolic fates of the major beer prenylflavanone isoxanthohumol that occur in mammalian system.A metabolic conversion study on microbes is known as one of the most useful tools to predict the xenobiotic metabolism of organic compounds in mammalian systems. The microbial biotransformation of isoxanthohumol ( The UV spectrum of both compounds displayed typical absorptions for a flavanone, with a maximum absorption peak at ~287 nm and an inflection at 318 nm, which were similar to those observed for 1. However, their 1H-NMR spectra showed major differences in the isoprenyl group, which showed two methyl signals shifted upfield (H-4″ and H-5″) at δH 1.31, 1.28  and 1.33, 1.34  as well as a new oxymethine signal (H-2″) at δH 3.76  and 3.73  in 2 and 3, respectively. These observations suggested that the two sp2 carbons of the isoprenyl group in 1 had been metabolized to oxygenated sp3 carbons. The 13C-NMR data of both compounds showed resonances for an sp3 oxymethine at δC 69.6 and 69.9 (C-2″) and an oxygenated sp3 quaternary carbon at δC 79.9 (C-3″), and the loss of the olefin signals at δC 124.0 and 131.9 that were observed in 1. An oxygenated aromatic carbon at δC 161.9 (C-7) in metabolites 2 and 3 was shifted upfield relative to the corresponding carbon in 1 (δC 164.0). These results indicated that a dihydropyran ring had been formed by oxidation and subsequent cyclization of the isoprenyl moiety, possibly going through an epoxide intermediate. Two-dimensional NMR data, including HSQC and HMBC experiments, supported the presence of a dihydropyran ring. The gem-dimethyl protons H3-4″ and H3-5″ correlated with C-2″ and C-3″, and the H2-1″ methylene protons correlated with C-8 and C-3″. Based on these spectroscopic analyses, the planar structure of diastereomeric metabolites 2 and 3 was assigned as 4′-hydroxy-5-methoxy-7,8--flavanone. The absolute configurations of the two asymmetric carbons, C-2 and C-2″, were established by circular dichroism + peaks at m/z 389.1612 and m/z 389.1600  respectively, which suggested a molecular formula of C21H24O7 corresponding to dihydroxylated derivatives of 1. The UV spectra of 4 and 5 displayed characteristic absorptions of a flavanone moiety at 225, 285 and ~310 nm, which were similar to their substrate 1. However, the 1H-NMR spectra of 4 and 5 exhibited major differences in the isoprenyl group resonances. They showed two pairs of methyl signals shifted upfield (H-4″ and H-5″) at δH 1.12, 1.15  and 1.14, 1.16 , as well as oxymethine signals (C-2″) at δH 3.52  and 3.55 , suggesting formation of two hydroxylated sp3 carbons from the olefinic group of 1. The 13C-NMR spectra of both compounds showed the presence of a hydroxymethine signal (C-2″) at δC 80.2 in 4 and δC 80.1 in 5, and an oxygenated quaternary carbon at δC 74.1 (C-3″) in both. In addition, 13C-NMR signals of a gem-dimethyl group at δC 25.0 and 26.0, were consistent with their substitution on an oxygenated carbon. HMBC correlations from H3-4″ and H3-5″ to C-2″ and C-3″, as well as from H2-1″ to C-8 and C-2″, and H-2″ to C-8 helped to establish the structure of diastereomers 4 and 5 as 7,4′-dihydroxy-5-methoxy-8--flavanone. The absolute configuration of the asymmetric center at C-2 was assigned by CD studies + peaks at m/z 393.1307 and m/z 393.1316 , respectively, which established their molecular formula as C21H22O6 which was isomeric with the mono-oxygenated metabolites 2 and 3. However, several significant differences were observed in the 1H- and 13C-NMR spectra of 6 and 7. Highly downfield shifted oxymethine proton  and carbon (δC 92.5) resonances of C-2″ indicated that a dihydrofuran ring was formed from the isoprenyl group substituted on the A-ring. The NMR data of 6 and 7 were in good agreement with those of (2S)-4′-hydroxy-5-methoxy-7,8-flavanone, a metabolite of xanthohumol by Pichia membranifaciens (ATCC 2254) flavanone, a metabolite of isoxanthohumol by F. equiseti (AM15) flavanones, and the absolute configurations of the asymmetric center at C-2 were identified by CD D +37.0° ; UV λmax (MeOH) (log ε) 224 (4.35) 285 (4.18), 319 (3.65) nm; CD (MeOH) λext (Δε): 288 (−13.5), 311 (0.0), 331 (+4.7); IR (KBr) νmax: 3421, 1658, 1608, 1579, 1519, 1485, 1338, 1206, 1133, 1106, 835 cm−1; 1H-NMR  δ 7.33 , 6.82 , 6.09 , 5.37 , 3.80 , 3.76 , 3.02 , 2.82 , 2.70 , 2.55 , 1.33 , 1.31 ; 13C-NMR  δ 192.7 (C-4), 164.1 (C-8a), 162.0 (C-5), 161.9 (C-7), 159.1 (C-4′), 131.4 (C-1′), 129.0 , 116.5 , 106.4 (C-4a), 101.9 (C-8), 94.9 (C-6), 80.4 (C-2), 79.9 (C-3″), 69.6 (C-2″), 56.3 (5-OCH3), 46.2 (C-3), 26.7 (C-1″), 25.7 (C-5″), 22.0 (C-4″); ESIMS m/z 371 [M + H]+; HRESIMS m/z 371.1504 [M + H]+ .-4′-Hydroxy-5-methoxy-7,8--flavanone (3): pale yellow amorphous powder, [α]D +66.4° ; UV λmax (MeOH) (log ε) 224 (4.47), 288 (4.30), 318 (3.77) nm; CD (MeOH) λext (Δε): 288 (+16.4), 313 (0.0), 334 (−5.4); IR (KBr) νmax: 3421, 1658, 1608, 1578, 1519, 1485, 1337, 1206, 1133, 1106, 835 cm−1; 1H-NMR  δ 7.32 , 6.82 , 6.08 , 5.36 , 3.79 , 3.73 , 3.00 , 2.83 , 2.69 , 2.50 , 1.34 , 1.28 ; 13C-NMR  δ 192.7 (C-4), 163.9 (C-8a), 162.0 (C-5), 161.9 (C-7), 159.1 (C-4′), 131.4 (C-1′), 129.0 , 116.5 , 106.4 (C-4a), 102.2 (C-8), 94.9 (C-6), 80.3 (C-2), 79.9 (C-3″), 69.9 (C-2″), 56.3 (5-OCH3), 46.2 (C-3), 26.7 (C-1″), 25.9 (C-5″), 21.3 (C-4″); ESIMS m/z 371 [M + H]+; HRESIMS m/z 371.1482 [M + H]+ .(2S)-7,4′-Dihydroxy-5-methoxy-8--flavanone (4): pale yellow amorphous powder, [α]D −31.7° ; UV λmax (MeOH) (log ε): 225 (4.42), 285 (4.17), 310 (3.73) nm; CD (MeOH) λext (Δε): 289 (−9.1), 313 (0.0), 331 (+4.0); IR (KBr) νmax: 3424, 1600, 1515, 1463, 1351, 1280, 1150, 1099, 828 cm−1; 1H-NMR  δ 7.34 , 6.81 , 6.16 , 5.33 , 3.81 , 3.52 , 3.00 , 2.93 , 2.69 , 2.62 , 1.14 , 1.12 ; 13C-NMR  δ 192.9 (C-4), 165.5 (C-7), 164.2 (C-8a), 162.5 (C-5), 159.0 (C-4′), 131.5 (C-1′), 129.0 , 116.4 , 108.5 (C-8), 105.9 (C-4a), 94.6 (C-6), 80.4 (C-2), 80.2 (C-2″), 74.1 (C-3″), 56.1 (5-OCH3), 46.3 (C-3), 26.6 (C-1″), 26.0 (C-4″), 25.0 (C-5″); ESIMS m/z 389 [M + H]+; HRESIMS m/z 389.1612 [M + H]+ .(2R)-7,4′-Dihydroxy-5-methoxy-8--flavanones (5): pale yellow amorphous powder, [α]D −53.0° ; UV λmax (MeOH) (log ε): 225 (4.30), 285 (4.08), 318 (3.65) nm; CD (MeOH) λext (Δε): 288 (+10.8), 311 (0.0), 331 (−3.8); IR (KBr) νmax: 3422, 2975, 1599, 1514, 1463, 1349, 1281, 1210, 1150, 1101, 829 cm−1; 1H-NMR  δ 7.34 , 6.81 , 6.17 , 5.34 , 3.82 , 3.55 , 2.98 , 2.95 , 2.70 , 2.61, 1.16 , 1.15 ; 13C-NMR  δ 192.9 (C-4), 165.5 (C-7), 164.0 (C-8a), 162.6 (C-5), 159.0 (C-4′), 131.6 (C-1′), 128.9 , 116.4 , 108.6 (C-8), 105.8 (C-4a), 94.7 (C-6), 80.5 (C-2), 80.1 (C-2″), 74.1 (C-3″), 56.1 (5-OCH3), 46.4 (C-3), 26.7 (C-1″), 26.0 (C-4″), 25.0 (C-5″); ESIMS m/z 389 [M + H]+; HRESIMS m/z 389.1600 [M + H]+ .1) for 5 d. Production of a metabolite was monitored by reversed-phase C18 TLC (MeOH 70%). The red-colored cultures were extracted with EtOAc two times and the organic layers were combined and concentrated in vacuo. The EtOAc extract (280 mg) was subjected to silica gel  column chromatography with n-hexane-EtOAc (2:1) mixture to give three fractions. Fraction 2 (89 mg) containing a metabolite was chromatographed with MPLC  using MeOH 45% isocratic solvent system to afford isoxanthohumol as a mixture of metabolites 6 and 7 . An aliquot of compound mixture (3.4 mg) was further chromatographed by HPLC with a gradient solvent system of 20% A to 35% A for 25 min to afford two isomers 6  and 7 .Scale-up fermentations were carried out under the same condition with two 1 L Erlenmeyer flasks each containing 250 mL medium and 25 mg isoxanthohumol (3 (1.0 mg in 0.2 mL pyridine) was treated with 15 μL (20.3 mg) of (R)-(−)-α-methoxy-α-(trifluoromethyl)phenylacetyl chloride (MTPA-chloride), and stirred overnight at room temperature under nitrogen gas. The reaction mixture was evaporated in vacuo, and chromatographed by HPLC using a SunFire™ Prep C18  with a gradient solvent MeCN–H2O (70:30 → 90:10) at 3 mL/min for 20 min, afforded the (S)-Mosher ester derivative 3a . The derivative of (R)-Mosher ester 3b  was prepared by reaction of 3 (1.0 mg) with 15 μL of (S)-(−)-MTPA-chloride reagent under the same condition as described above.Compound 3a: 1H-NMR  δ 6.03 , 5.44 , 5.16 , 3.86 , 2.99 , 2.93 , 2.83 , 2.65 , 1.36 , 1.32 . 3b: 1H-NMR  δ 6.06 , 5.46 , 5.16 , 3.87 , 3.00 , 2.95 , 2.87 , 2.80 , 1.31 , 1.27 .1) resulted in the production of three oxygenated pairs of metabolites (2–7). These include two novel metabolite pairs possessing a dihydropyran  and a dihydroxymethylbutane  substituted flavanone core structure. It was widely reported that cyclization at the prenyl side chain resulted in forming a five-membered furan or a six-membered pyran heterocycle attached to the A-ring in prenylated flavanones [Brosimum acutifolium, a rich source of 8-prenylated flavonoids [Microbial transformation studies of isoxanthohumol (avanones ,30. The avanones . These tavanones , and theavonoids .2–7, the regioselectively mono-hydroxylated prenyl side chain metabolites of isoxanthohumol, namely cis- and trans-prenyl alcohols, were previously identified as the most abundant oxidation metabolites of isoxanthohumol during in vitro metabolism studies using human liver microsomes, which was catalyzed by hepatic cytochrome P450 enzymes [Unlike the microbial metabolites  enzymes ,33. Howe enzymes ,35. It w


Cyrtopodium paniculatum. Eight new compounds, including one phenanthrene 1, one 9,10-dihydro-phenanthrene 2, one hydroxybenzylphenanthrene 3, two biphenanthrenes 4–5, and three 9,10 dihydrophenanthrofurans 6–8, together with 28 known phenolic compounds, mostly stilbenoids, were isolated from the CH2Cl2 extract of its leaves and pseudobulbs. The structures of the new compounds were established on the basis of extensive spectroscopic methods.We report the first phytochemical study of the neotropical orchid  Arundina, Bletilla, Dendrobium, Gastrodia and Pleione at the expense of the New World orchids In fact, the first report on a phytochemical study of South American orchids was in the late 1990s and was mainly focused on species in the genus Scaphyglottis + peak at m/z 331.1188  in the HRESIMS, indicating a molecular formula of C18H18O6 and suggesting the existence of ten degrees of unsaturation. The UV maximal absorptions at 216, 261, 281, 312 and 344 nm indicated the presence of a phenanthrene derivative +; , suggesting the presence of nine degrees of unsaturation. The UV spectrum showed absorption maxima at 220, 262 and 282 nm suggesting a 9,10 dihydrophenanthrene moiety + signal at m/z 407.1508  in the HRESIMS, indicating a molecular formula of C24H22O6 and fourteen degrees of unsaturation. Its UV spectrum showed maximal absorptions at 212, 268, 289, 297, 301 and 314 nm suggesting a phenanthrene moiety. The 1D-NMR data indicated that the structure of 3 comprised a confusarin (11) + molecular ion peak at m/z 511.1742 indicating a molecular formula of C31H26O7  which is consistent with nineteen degrees of unsaturation. Its UV spectrum showed maximal absorptions at 213, 263, 296, 310, 349 and 367 nm. The IR spectrum showed typical absorption bands of a hydroxyl group at 3245 cm−1 and of aromatic rings at 1608, 1585, 948, 865 and 830 cm−1. In 1D-NMR spectrum  and δH 8.28  suggested that 4 was a dimeric molecule formed from the fusion of a phenanthrene unit -2,2,7,7′-tetraol and named lusidol A.This was also supported by the downfield shift in the 5 was obtained as a pale yellow amorphous powder. Its molecular formula was determined as C31H26O7 by the HRESIMS [M + H]+ peak at m/z 511.1740  which indicated that it has the same molecular formula as compound 4. The UV and IR spectra were identical to those of 4. Comparison of the NMR spectral data of these two compounds revealed their structural similarities, suggesting that the structure of 5 comprises the same phenanthrene (nudol) and 9,10-dihydrophenanthrene (lusianthridin) moieties  in 5. The NOESY correlations from H-8 to H-9, H-10 to H-4′, as well as subsequent HMBC correlations from H-10 and H-4′ to C-1; H-8 to C-4b, C-6 and C-9 confirmed the position of this linkage. The optical rotation experiment result was zero, thus suggesting that 5 is a racemic compound. Therefore, compound 5 was identified as a 3,4,7′-trimethoxy-9′,10′-dihydro--2,2′,5′,7-tetraol and named lusidol B.Compound moieties . The onlage in 4 . This wa6 was obtained as a white amorphous powder. Its molecular formula was determined to be C33H32NO7 based on its HRESIMS [M + H]+ molecular ion peak at m/z 554.2169 . Its UV spectrum showed maximal absorption bands at 208, 281, 305 and 318 nm, indicative of a dihydrophenanthrene derivative. Its IR spectrum exhibited absorption bands at 3241 cm−1 (hydroxyl), 1704 and 1519 cm−1 (amide), 1199, 1118, 1015, 950, 814 and 773 cm−1 (aromatic rings). Analysis of the 13C-NMR and DEPT-135 spectra of 6 disclosed the presence of signals belonging to 24 aromatic carbons, comprising of eleven methine, seven quaternary, and six oxygenated tertiary carbons. The 1H-NMR spectrum , 6.68 , 6.68  and one aromatic proton singlet at δH 6.54 . Two methylene proton signals at δH 2.45–2.55  and 2.55–2.60  and the corresponding carbon signals at δC 30.7 and 27.5, respectively, were characteristic of the methylene groups of a 9,10-dihydrophenanthrene skeleton. A 4′-hydroxy-3′-methoxyphenyl moiety comprising of three aromatic protons at δH 6.82 , 6.82  and 6.99  that formed the second ABX system. The presence of an A2B2 system characteristic of a symmetrical hydroxybenzyl ring was inferred by the resonance at δH 7.01 , 6.73 , as well as signals attributed to an ethylamide moiety at δH 3.46 , 2.72 , 7.28  and a amide carbonyl at δC 172.3, thus achieving a hydroxybenzylethylamide moiety. The connection between the three partial structures of 9,10-dihydrophenanthrene, 4′-hydroxy-3′-methoxyphenyl and hydroxybenzylethylamide moiety was achieved through a central furan ring as observed by the presence of two coupled methines at δH 4.11  and the second being oxygenated at δH 5.67 . This was confirmed by extensive analysis of HMBC and NOESY spectra (H 5.67) showed HMBC correlations to C-2, C-4′, C-5′, C-9′ and the amide carbonyl at C-1′; H-2′ to C-1, C-2, C-1′, C-3′, and C-4′. Additional HMBC correlations were observed from the two aromatic protons at H-5′ to C-3′ and H-9′ to C-3′.Compound spectrum  showed r spectra . The oxyH 3.88  and 3.82  as well as the three hydroxyl groups on position C-7, C7′ and C-7′′ were confirmed by HMBC and NOESY analysis and supported by the characteristic chemical shift of the carbon bearing the groups. The hydroxyl bearing carbon position was settled in C-7 based on the HMBC correlation between H-5 and C-7 and by the characteristic chemical shift at δC 156.2. The positioning of the methoxyl on C-4 was affirmed from the NOESY cross peaks between H-3, H-5 and 4-OCH3.This was further supported by NOESY correlations between H-2′ to H-10, H-5′, H-9′ and 1′′-NH; H-3′ to H-5′, H-9′, and 1′′-NH. The assignments and locations of the two methoxyl groups at δtrans- based on NOESY correlations between H-2′ and H-5′′/9′′, suggesting these protons are on the same side of the furan ring. This was also supported by the coupling constant of 6.6 Hz, typical of trans-dihydrophenanthrenofuran derivatives furan-3-carboxamide, and it was named moupilonin.The relative configuration of the methine protons H-2′ and H-3′ on the furan ring was defined as ivatives ,56, whicure of 6  obtained7 was obtained as a white amorphous powder. Its molecular formula was determined as C26H26O7 as deduced from its HRESIMS [M + H]+ ion peak at m/z 451.1753 , suggesting the presence of fourteen degrees of unsaturation. The UV spectrum showed absorption maxima at 210, 274 and 284 nm, suggesting that compound 7 has a dihydrophenanthrene skeleton. The IR absorption bands at νmax at 3303, 1601, 1452 and 773 cm−1 were characteristic to a hydroxyl and aromatic moieties.Compound 1H-NMR spectrum , 6.73 , 6.73 , and 6.56 , as well as two methylenes at δH 2.69  and 2.67 , which corresponds to resonances attributed to a 9,10-dihydrophenanthrene ring. A singlet aromatic proton integrating for two protons at δH 6.75  suggested the presence of a symmetrical 1′,3′,4′,5-tetrasubstituted aromatic ring. Three methoxy groups at δH 3.80 , 3.60  and two hydroxyl groups at δH 8.29  and 7.24 , were also noticed. The linkage between the 9,10-dihydrophenanthrene unit and 1′,3′,4′,5-tetrasubstituted aromatic moiety of compound 7 was achieved through a furan ring as supported by signals assigned to an oxymethine proton at δH 5.66 , a methine at δH 3.73  and an oxygenated methylene at δH 3.83  and δH 4.08  which coupled to an hydroxyl group at δH 4.17 . Analysis of the COSY spectrum (2(13)-CH(12)-CH(11) proton spin systems. Furthermore, HMBC correlations from H-11 to C-2, C-2′/6′; H-2′/6′ to C-11 as well as NOESY cross peaks between H-11 and H-2′/6′; H-2′/6′ and H-12 supported the structure as shown, connecting the phenyl to the furan ring at C-11.The spectrum  showed rspectrum  validate3 to C4 and 3′/5′-OCH3 to C-3′/5′ suggested that the methoxy groups were attached to C-4 and C-3′/5′, respectively. NOESY correlations between 4-OCH3 and H-5, and between H-2′/H-6′ to 3′/5′-OCH3 also validated this assignment. The hydroxyl groups were assigned to positions C-7 and C-4′ based on HMBC long range correlations between 7-OH to C-7 , and 4′-OH to C-4′ . The relative stereoconfiguration of 7 was established on the basis of NOESY experiments, which displayed cross peaks between H-12 on the furan ring and H-2′/6′ indicated that both protons reside on the same side. In addition, the coupling constant of 5.4 Hz between H-11 and H-12 was in agreement with the reported literature values for the relative trans-configuration furan-3-ol and named cyrtonesin A .The assignment of the substitution pattern was achieved based on HMBC correlations from 4-OCHguration . The abs reports ,59. The 8 was obtained as a white amorphous powder and its molecular formula was established as C26H26O7 by the HREIMS peak at m/z 451.1748 [M + H]+ . The mass, UV, IR and NMR spectra  as well as HMBC correlations --2--3-(hydroxymethyl)-10-methoxy-2,3,4,5-tetra-hydrophenanthrofuran-7-ol. Compound 8 was named cyrtonesin B.The methylene at position C-13 also had NOESY correlation with the proton H-10 on the dihydrophenanthrene subunit. The relative configuration between H-11 and H-12 was assumed to be ivatives . The CD 13C/1H Cryoprobe . Acetone-6d and methanol-4d  were used as deuterated solvents and their protonated residual signals were used as internal standard at 2.05 ppm and 3.31 ppm respectively, relative to TMS. The HRESIMS analysis were performed on a HPLC-DAD/UV-MS Agilent 1200 series coupled to a 6520 Q-ToF mass spectrometer . The acquisition of mass spectra was conducted in ESI positive ion mode. Column chromatography and vacuum liquid chromatography were carried out using 40–60 µm silica gel . The obtained fractions were monitored by TLC and the spots were visualized under UV light (254 nm) and by 2% sulfuric vanillin reagent. Sephadex LH-20 , was used for gel chromatography. RP-HPLC experiments were conducted on a Gilson LC system  equipped with a semi preparative Kinetex Axia C-18 column . Experiments were conducted at a wavelength of 280 nm. Preparative TLC was performed on a glass supported silica gel 60F254 . Analytical grade solvents of HPLC quality were purchased from Sigma Aldrich.Optical rotations were measured with a P-2000 polarimeter  and circular dichroïsm spectra were recorded on a Jasco J-510 spectropolarimeter apparatus (Jasco). UV spectra were recorded on a UV-2401 PC spectrometer . IR spectra were obtained on a 380 FT-IR spectrophotometer .The 1D and 2D NMR spectra were recorded on a Bruker 500 MHz Avance III spectrometer  equipped with a DCH C. paniculatum (Ruiz & Pav.) Garay were purchased from the orchid farm Orquidea del Valle, Ginebra, Colombia in October 2013 and imported to France, according to the Convention of Natural Trades in Endangered Species (CITES). The voucher specimens (No 58054 and 58056) were deposited at the Herbarium of CUVC, Universidad del Valle, Cali, Colombia.Fifteen fresh specimens of 2Cl2 and n-BuOH to yield after solvent removal a cyclohexane extract (0.60 g), a CH2Cl2 extract (1.67 g) and an n-BuOH extract (7.88 g). The CH2Cl2 extract was subjected to silica column chromatography  to afford 24 fractions (A–X). Fraction D (24.4 mg) was subjected to preparative thin layer chromatography  to afford compound 9 (6.3 mg). Fraction F (51.6 mg) was fractionated on Sephadex LH-20 (CH3OH) to obtain sub-fractions F1-F8. Sub-fractions F4 (7.4 mg) and F5 (25.7 mg) were further purified using PTLC (CHCl3/CH3OH 94:6) to afford 10 (4 mg), 11 (5 mg), 12 (5 mg) and 13 (0.9 mg). Fraction G (165.4 mg) was purified using Sephadex LH-20 (CH3OH) to obtain eight sub-fractions (G1–G8). Sub-fraction G8 was obtained as a pure compound 14 (2.8 mg). Fraction H (71.6 mg) was also subjected to Sephadex LH-20 (CH3OH) to obtain four sub-fractions (H1–H4). H1 (12.2 mg) led to the isolation of 15 (1.4 mg) and 16 (1.3 mg) on PTLC eluting with CHCl3/CH3OH (94:6). Sub-fraction H2 (4.7 mg) led to the isolation of 17 (1 mg) and 18 (0.9 mg) using PTLC (CHCl3/CH3OH). Fraction I (87 mg) was purified on Sephadex LH-20 (CH3OH) to obtain seven sub-fractions (I1-I7). Sub-fraction I1 (25.6 mg) led to the isolation of 19 (13.1 mg), while sub-fraction I2 (13.3 mg) afforded compounds 20 (5.3 mg) and 21 (4.1 mg) using PTLC eluting with CHCl3/CH3OH (94:6). Sub-fraction I7 (4.2 mg) was purified on a PTLC  to obtain compound 1 (1.3 mg). Fraction J (32.2 mg) was fractioned on Sephadex LH-20 (CH3OH) and PTLC  to obtain 22 (0.9 mg) and 23 (1 mg). Fraction K (29.5 mg) was purified on Sephadex LH-20 (CH3OH) affording 7 sub-fractions (K1–K7). Sub-fraction K6 (4.7 mg) was subjected to a PTLC eluting with CHCl3/CH3OH (94:6) affording compounds 3 (0.6 mg), 24 (2 mg) and 25 (0.6 mg). Fraction L (26.6 mg) was also subjected to Sephadex LH-20 (CH3OH) to yield 8 sub-fractions (L1–L8). Sub-fraction L5 (2.2 mg) was purified using PTLC with CHCl3/CH3OH (94:6) as the eluting solvent to yield 26 (0.6 mg). Fraction M (90.0 mg) was subjected to Sephadex LH-20 and eluted with CH3OH to afford 10 sub-fractions (M1–M10). Compounds 27 (2.4 mg) and 28 (6.7 mg) were obtained from sub-fraction M2 (19.9 mg) by PTLC using CHCl3/CH3OH as the mobile phase. Sub- fraction M7 (13.2) was further purified by PTLC using CHCl3/CH3OH (94:6) to obtain 29 (2.1 g) and 30 (2.6 mg). Sub-fraction M8 (2.9 mg) was passage over PTLC that was eluted with CHCl3/CH3OH (94:6) to yield compound 31 (1.1 mg). Sub-fraction M9 (11.6 mg) was purified by PTLC with CHCl3/CH3OH (94:6) to yield compounds 4 (0.7 mg), 5 (0.6 mg) and 32 (0.9 mg). Fraction O (71.7 mg) was subjected to Sephadex LH-20 eluting with CH3OH, to obtain 5 sub-fractions (O1–O5). Sub-fraction O3 (9.8 mg) was purified by PTLC with CHCl3/CH3OH (94:6) to obtain compound 2 (2.8 mg). Sub-fraction O5 (2.4 mg) was purified by PTLC with CHCl3/CH3OH (94:6) to give compound 33 (0.7 mg). Fraction P (63 mg) was fractionated on Sephadex LH-20 (CH3OH) to obtain six sub-fractions (P1–P6). Sub-fraction P4 (3.7 mg) was further subjected to PTLC eluting with CHCl3/CH3OH (94:6) to obtain compounds 6 (0.6 mg), 7 (0.9 mg) and 8 (0.6 mg). Fraction Q (19.5 mg) was purified on Sephadex LH-20 (CH3OH) affording six sub-fractions. Compounds 34 (3 mg) and 35 (0.8 mg) were obtained from sub-fraction Q4 (7 mg) through a PTLC with CHCl3/CH3OH (94:6) as the eluting solvent.The pseudobulbs (7.5 kg) were cut into small pieces, ground and soaked in demineralized water for 30 min to get rid of the mucilage content. The remaining part was then macerated with ethanol and the filtrates were evaporated under reduced pressure to obtain a crude ethanolic extract (60 g). The ethanolic extract was suspended in water and successively partitioned with cyclohexane, CH2Cl2 and CH3OH (1 g/15 mL × 3) to afford a cyclohexane extract (6.99 g), a CH2Cl2 extract (1.23 g) and a CH3OH extract (4.98 g). The CH2Cl2 extract was subjected to vacuum liquid chromatography (VLC) using cyclohexane, EtOAc and CH3OH to afford six main fractions (A–F). Fraction E (98 mg) was subjected to semi-preparative HPLC  to afford compounds 13 (3.5 mg), 18 (0.7 mg), 19 (1 mg), and 21 (1.7 mg). Fractions F (158.4 mg) was also subjected to semi- preparative RP-HPLC (35% B for 35 min) to afford compound 36 (9.6 mg).Fresh leaves were lyophilized and grinded to afford 70 g of dried powder and were successively extracted with cyclohexane, CHCyrtopodin (1). Brown amorphous powder (1.3 mg); UV (CH3OH) λmax (log ε): 216 (3.96), 261 (4.38), 281 (4.02), 312 (3.48), 344 (2.52); IR (FT-IR) νmax: 3370, 2935, 2831, 1616, 1573, 1465, 1268, 1118, 1061, 1021, 953, 860, 770 cm−1; 1H-NMR and 13C-NMR see m/z 331.1188 [M + H]+ .(+)-9S-Hydroxyerianthridin (2). White amorphous powder (2.8 mg); 3OH); CD (CH3OH): nm (Δε): 236.2 (−20.69), 280.9 (+5.39); (UV (CH3OH) λmax (log ε): 220 (4.01), 262 (3.91), 282 (4.00), 348 (2.28), 364 (2.26); IR (FT-IR) νmax: 3227, 2936, 2834, 1611, 1584, 1446, 1412, 1348, 1213, 1112, 1066, 999, 946, 870, 828 cm−1; 1H-NMR and 13C-NMR see m/z 311.0884 [M + Na]+; .Gastrodiconfusarin (3). Yellow amorphous powder (0.6 mg); UV (CH3OH) λmax (log ε): 212 (4.11), 268 (4.09), 289 (3.63), 297 (4.00), 301 (3.59), 314 (3.45); IR (FT-IR) νmax: 3336, 2931, 2936, 1606, 1512, 1454, 1352, 1221, 1106, 1017, 822, 773 cm−1; 1H-NMR and 13C-NMR see m/z 407.1508 [M + H]+ .Lusidol A (4). Pale yellow amorphous powder (0.7 mg); 3OH); UV (CH3OH) λmax (log ε): 213 (4.80), 263 (4.82), 296 (4.47), 310 (4.40), 349 (3.58), 367 (3.55); IR (FT-IR) νmax: 3245, 2931, 2835, 1608, 1585, 1451, 1393, 1351, 1210, 1162, 1118, 1076, 999, 948, 865, 830 cm−1; 1H-NMR and 13C-NMR see m/z 511.1742 [M + H]+ .Lusidol B (5). Pale yellow amorphous powder (0.6 mg); 3OH); UV (CH3OH) λmax (log ε): 212 (4.72), 264 (4.66), 297 (4.37), 309 (4.29), 350 (3.28), 367 (3.27). IR (FT-IR) νmax: 3271, 2931, 2835, 1608, 1585, 1451, 1393, 1351, 1210, 1162, 1118, 1076, 999, 948, 865, 830; 527 cm−1; 1H-NMR and 13C-NMR see m/z 511.1740 [M + H]+ .(±)-Moupilonin (6). White amorphous powder (0.9 mg); 3OH); UV (CH3OH) λmax (log ε): 208 (4.37), 281 (3.97), 305 (3.73), 318 (3.51); IR (FT-IR) νmax : 3241, 2935, 2829, 1704, 1519, 1447, 1349, 1237, 1199, 1118, 1015, 950, 814, 773 cm−1; 1H-NMR and 13C-NMR see m/z 554.2169 [M + H]+ .(+)-Cyrtonesin A (7). White amorphous powder (0.6 mg); 3OH); CD (CH3OH): nm (Δε): 233.1 (+0.84), 284.2 (−0.95); UV (CH3OH) λmax (log ε): 210 (4.30), 274 (3.73), 284 (3.76); IR (FT-IR) νmax: 3303, 2935, 2831, 1601, 1452, 1359, 1239, 1217, 1114, 1068, 1033, 1008, 977, 951, 824, 773 cm−1; 1H-NMR and 13C-NMR see m/z 451.1753 [M + H]+ .(+)-Cyrtonesin B (8). White amorphous powder (0.6 mg); 3OH); CD (CH3OH): nm (Δε): 235.0 (+1.11), 272.8 (−7.26); UV (CH3OH) λmax (log ε): 207 (4.29), 292 (3.73), 301 (3.77); IR (FT-IR) νmax: 3320, 2936, 2825, 1599, 1460, 1354, 1239, 1221, 1060, 1035, 1008, 977, 955, 825, 773 cm−1; 1H-NMR and 13C-NMR see m/z 451.1748 [M + H]+ .2Cl2 extract from the aerial parts of C. paniculatum led us to the isolation of an long list of polyphenolic derivatives, mainly stilbenoids. Monomeric stilbenoid derivatives, comprising bibenzyls, 9,10-dihydrophenanthrenes, phenanthrenes and phenanthrenequinones, are well represented in this orchid. The second set of isolated compounds consisted of phenolic adducts (4-hydroxybenzyl) coupled to a 9,10-dihydrophenanthrene or phenanthrene derivative. This type of 4-hydroxybenzyl adduct (gastrodigenin) is very common in orchids and mostly found in the genus Arundina [Bletilla [Pleione [trans-feruloyltyramine for 6, synapyl alcohol for 7 and 8). The connection between the two units allows a new cyclization leading to a furan ring. It is noteworthy that this kind of dihydrophenanthrene derivative is considered as a chemotaxonomic marker for the species in the genus Pleione [Bulbophyllum [Cremastra [Bletilla [Bulbophyllum [Cremastra [Cyrtopodium paniculatum.Due to extreme environmental and territorial conditions as well as predation, plant species have evolved to produce complex secondary metabolites as a means of survival. In orchids, stilbenoids and phenanthrene derivatives are produced as major phytoalexins produced in response to various biotic and abiotic stressors, mostly fungal, bacterial and worm attacks ,64,65,66Arundina ,68, BletBletilla ,70,71,72[Pleione ,73,74. T Pleione ,76,77, bophyllum  and Cremremastra . The lasBletilla ,79,80, Bophyllum ,81,82,83remastra ,86,87,88remastra  or an enremastra ,90,91. T


H-1,2,3-triazol-4-yl)-1,3,4-thiadiazoles 2-4 have been synthesized by the reaction of 2-aryl-2H-1,2,3-triazole-4-carboxylic acids 1 with thiosemicarbazide. Their reaction with phenacyl (p-substituted phenacyl) bromides led to formation of the respective 6-aryl-2-imidazo-1,3,4-thiadiazoles 5. Reactivity of the latter fused ring towards reaction with different electrophilic reagents afforded the corresponding 5-substituted derivatives 6-8. The structure of the above compounds was confirmed from their spectral characteristics. Some of these compounds were found to possess slight to moderate activity against the microorganisms Staphylococcus aureus, Candida albicans, Pseudomonas aeruginosa, and Escherichia coli. 2-Amino-5-(2-aryl-2 Somele rings , antituble rings , antibacle rings , antifunle rings , anticonle rings , antiserle rings  and antile rings  propertiammatory , cardiotammatory , diuretiammatory  and herbammatory  agents aammatory ,29,30. WH-1,2,3-triazole-4-carboxylic acids 1 -1,3,4-thiadiazoles 5a-i. The reaction proceeds via the formation of the iminothiadiazole intermediate, followed by dehydrative cyclization to form the desired fused heterocycle -1,3,4-thiadiazole (3). Yield (77%); m.p. 259–260 °C ; IR (cm−1): 3485, 3300 (NH2), 1639 (thiadiazole ring C=N), 1597 (triazole ring C=N) and 660 (thiadiazole ring C-S-C); 1H-NMR : 7.55 , 7.61 , 7.98  and 8.52 ; MS, m/z (%): 324 , 322 , 266 , 239 , 244 , 188 , 184 , 169 ,161 , 145 , 133 , 105  and 91 ; Anal. Calc. for C10H7BrN6S (323.17): C, 37.17; H, 2.18; N, 26.00%, found. C, 37.00; H, 2.60; N, 25.70%.2-Amino-5--1,3,4-thiadiazole (4). Yield (65%); m.p. 269–270 °C ; IR (cm−1): 3465, 3290 (NH2), 1626 (thiadiazole C=N), 1532 (triazole C=N) and 694 (thiadiazole C-S-C); Anal. Calc. for C10H7ClN6S (278.72): C, 43.09; H, 2.53; N, 30.15%, found. C, 43.12; H, 2.51; N, 30.17%. 2-4 (0.01 mol) and phenacyl (p-substituted phenacyl) bromides (0.01 mol) was refluxed in dry ethanol (50 mL) for 24 hours. The excess of solvent was distilled off and the solid hydrobromide salt that separated out was collected by filtration, suspended in water and neutralized by sodium carbonate to get free base 5a-i. The product was filtered, washed with water, dried and crystallized from carbon tetrachloride. Their IR, 1H-NMR spectra and analytical data are listed in A mixture of equimolar quantities of thiadiazoles 5a-i (0.01 mol) in acetic acid (10 mL) with stirring for one to eight days at room temperature. The reaction mixture was poured onto ice cold water. The separated solid was collected, washed with water, dried and crystallized from either chloroform or benzene in methanol mixture. Their IR, 1H-NMR spectra and analytical data are listed in A solution of bromine (iodine monochloride) (0.01 mol) in acetic acid (10 mL) was gradually added to a suspension of 5a  was added to the reagent and stirred at 0 °C for 30 min. The mixture was further stirred at room temperature for 2 hours and then at 60 °C for an additional 2 hours. The reaction mixture was then poured onto sodium carbonate solution and stirred at 90 °C for 2 hours. After cooling, the mixture was diluted with water and extracted with chloroform. The chloroform layer was washed with water, dried over anhydrous sodium sulfate and the residue obtained after removal of the solvent was crystallized from chloroform/ methanol. Yield (80%); m.p. 184–185 °C; IR (cm−1): 2921, 2885 (formyl hydrogen), 1676 (CO), 1629 (thiadiazole C=N), 1589 (imidazole C=N), 1492 (triazole C=N), 1444 (imidazole C=C) and 664 (thiadiazole C-S-C); 1H-NMR : 7.44 , 7.54 , 7.89 , 8.15 , 8.53  and 10.1 ; Anal. Calc. for C19H12N6OS (372.40): C, 61.28; H, 3.25; N, 22.57%, found. C, 61.30; H, 3.30; N, 22.57%.The Vilsmeir-Haack reagent was prepared by adding phosphorus oxychloride (3 mL) to dimethyl-formamide (20 mL) at 0 °C with stirring. Then, compound The antibacterial activity tests were performed according to agar diffusion method  using amb]-1,3,4-thiadiazole fused heterocyles. The antibacterial activities of the synthesized compounds were also examined.In conclusion, formation of diverse 2-amino-5-substituted-1,3,4-thiadiazoles using carbohydrates as precursors was successfully achieved, followed by their conversion into the novel target imidazo[2,1-


Sida rhombifolia L.  aims to strengthen the chemosystematics and pharmacology of Sida genera and support the ethnopharmacological use of this species as hypotensive herb. The present work reports phytoconstituents isolated and identified from aerial parts of S. rhombifolia by using chromatographic and spectroscopic methods. The study led to the isolation of scopoletin (1), scoporone (2), ethoxy-ferulate (3), kaempferol (4), kaempferol-3-O-β-d-glycosyl-6′′-α-d-rhamnose (5), quindolinone (6), 11-methoxy-quindoline (7), quindoline (8), and the cryptolepine salt (9). The alkaloids quindolinone (6) and cryptolepine salt (9) showed vasorelaxant activity in rodent isolated mesenteric arteries.The follow-up of phytochemical and pharmacological studies of  Sida genus is considered native to Brazil and presents 95 species oxy]-5,7-dihydroxy-2-(4-hydroxyphenyl)-. Yellow powder: 1H-NMR (500 MHz) (CD3OD) δ: 6.16 , 6.34 , 8.04 , 6.87 , 5.08 , 3.38-3.82 , 4.50 , 3.38-3.82  and 1.12 . 13C-NMR (125 MHz) (CD3OD) δ: 159.4 (C-2), 137.4 (C-3), 179.3 (C-4), 162.8 (C-5), 100.0 (C-6), 166.6 (C-7), 94.9 (C-8), 158.5 (C-9), 105.5 (C-10), 122.6 (C-1′), 132.3 (C-2′/6′), 116.1 (C-3′/5′), 161.4 (C-6′), 104.5 (C-1′′), 71.3 (C-2′′), 78.0 (C-3′′), 75.6 (C-4′′), 69.7 (C-5′′), 68.5 (C-6′′), 102.3 (C-1′′′), 72.0 (C-2′′′), 72.2 (C-3′′′), 77.1 (C-4′′′), 73.8 (C-5′′′) and 17.9 (C-6′′′). In agreement with literature [11H-Quindolin-11-one-5,10-dihydro . Yellow powder: 1H-NMR (500 MHz) (DMSO-d6) δ: 8.35 , 7.28 , 7.68 , 7.73 , 12.43 , 8.19 , 7.20 , 7.47 , 7.52  and 11.67 . 13C-NMR (125 MHz) (DMSO-d6) δ: 125.1 (C-1), 120.4 (C-2), 130.6 (C-3), 117.7 (C-4), 139.0 (C-4a), 128.8 (C-5a), 115.8 (C-6a), 120.7 (C-6), 118.8 (C-7), 127.3 (C-8), 112.5 (C-9), 138.5 (C-9a), 123.0 (C-10a), 167.3 (C-11) and 122.8 (C-11a). In agreement with literature [10H-Quindoline, 11-methoxy . Yellow powder: 10H-Quindoline . Orange crystal. High Resolution Mass Spectrometry by Electronspray (HRESI) (+)-MS m/z 219.09 . 1H-NMR (500 MHz) (DMSO-d6) δ: 7.63 , 7.39 , 7.67 , 8.29 , 8.44 , 7.24 , 7.57 , 11.72  and 8.52 . In agreement with literature [terature .10H-Quindolinium, 5-methyl : NMR data published [6 (quindolinone) and 9  were evaluated by increasing cumulative addition of the compound (10−12–10−3 M) using rodent mesenteric arteries pre-contracted with phenylephrine (PHE) (1 μM). The experiments were carried out using arteries with functional endothelium and without functional endothelium .The vasorelaxant activity of compounds othelium . Data we1H-NMR spectra of compound 7 showed signals with a pattern of substitution indicating an indoquinoline skeleton [H 8.37 J = 7.85 Hz (H-6); δH 8.30 J = 8.45 Hz (H-1); δH 8.18 J = 8.5 Hz (H-4); δH 7.64 J = 7.1 Hz (H-9), besides four broad triplets: δH 7.70 J = 7.45 Hz (H-3); δH 7.63 J = 7.6 Hz (H-8); δH 7.58 J = 7.55 Hz (H-2); δH 7.30 J = 7.0 Hz (H-7)  . H 11.50 (1H) was detected indicating the N-H of indol (NH-10) found in this group of alkaloids [H 4.39  were attributed to a methoxy group bonded to C-11, since the chemical shift at δH 9.02 characteristic of H-11 was not found [A singlet at δot found . 13C-NMR data supported the proposal of indoquinoline nucleous for compound 7 by showing signals for 15 carbons. The chemical shift at δC 60.7 indicated a methoxy group as proposed by 1H-NMR, being confirmed by the HMQC ; H-9 with C-6a (J3); H-8 with C-9a (J3) and H-1 with C-9a (J2) (The HMBC spectrum of -9a (J2) .7 showed correlations between neighbour hydrogens, as well as confirmed that the methoxy is bonded to C-11, by showing correlation between δH 4.39(O-CH3) with δH 11.50 (NH-10) and δH 8.30 .The NOESY spectra of compound 6) and the salt of cryptolepine (9) promoted vasorelaxation in mesenteric artery rings with functional endothelium and after endothelium removal. The PHE potency in promoting arterial contraction was significantly reduced by compounds 6 and 9  (The tested substances quindolinone ( < 0.05) . 2) into inositol 1,4,5-trisphosphate (IP3) and diacylglycerol (DAG) by the action of phospholipase C-β (PLCβ). Then, the IP3 receptors at the sarcoplasmic reticulum are activated, releasing Ca2+ into the cytosol. DAG and the elevation of intracellular Ca2+ activate kinase proteins, including protein kinase C (PKC) in smooth muscle. It results in phosphorylation and inactivation of enzymes related to the contractile process, such as myosin light chain phosphatase (MLCP) [PHE is an agonist of α1- adrenergic receptors ,14, whose (MLCP) ,16. -derived relaxing factor (EDRF), such as nitric oxide (NO), prostacyclin, bradykinin, endothelium-derived hyperpolarizing factor (EDHF), and metabolites of monooxygenases, among others [-derived constrictor factors (EDCF) promote contraction, i.e., angiotensin II, endothelin, vasoconstrictors prostanoids, and reactive oxygen species, such as superoxide anion [After endothelium removal, the vasorelaxant effects of both tested compounds were significantly altered. The vascular endothelium secretes several biologically active substances, and many of them are active in vessels’ muscle cells, modulating the vascular tone. The vasodilator mechanism of endothelium is mediated by endotheliumg others ,18,19,20de anion ,22.S. rhombifolia has been previously reported [Thus, the vasorelaxant activity of tested compounds seems to involve endothelium-derived substances, since the effect was decreased after the endothelium removal. However, the compounds also seem to act in smooth muscle cells, since in endothelium-free preparations it was possible to observe some vasorelaxant effect. The vasorelaxant effect of cryptolepinone isolated from reported .1H- and 13C-NMR spectra were recorded on a Varian Oxford 200 NMR spectrometer  and on a Varian 500 NMR spectrometer . Mass spectra were obtained on a Xevo® G2-XS QT of mass spectrometer . Silica gel 60  7734 , neutral alumina, Sephadex LH-20, and XAD-2 were used for the chromatographic procedures. Sida rhombifolia was collected in Santa Rita city  in September 2008 and identified by Dr. Maria de Fátima Agra. A voucher specimen (Agra 7045) is kept at the herbarium Professor Lauro Pires Xavier (CCEN/UFPB). The dried and powdered aerial parts of the plant (5.5 kg) were extracted with 95% EtOH  for 72 h. The EtOH extract was concentrated under reduced pressure at 40 °C, providing 570 g of crude ethanol extract (CEE).S. rhombifolia showed positive results when tested with Dragendorff, Bouchadart, and Mayer reagents for detection of alkaloids. Thus, the classic alkaloid extraction was performed using 100 g of the CEE.The CEE of 3 fraction (1.2 g) was subjected to silica gel column chromatography (CC) using hexane (Hex), dichloromethane (CH2Cl2), and methanol (MeOH), alone or in binary mixtures of increasing polarity, resulting in the isolation of compounds 1 (21 mg), 2 (9 mg), and 3 (8 mg). The total alkaloids fraction (800 mg) was submitted to CC using neutral alumina as stationary phase. The column fraction 14/43 (28 mg) was chromatographed on preparative, Analytical Thin Layer Chromatography (TLC) using alumina and eluted with Hex:ethyl acetate (8:2), yielding the pure compounds 7 (12 mg) and 8 (5 mg).The acid CHClPart of the CEE (200 g) was subjected to filtration under reduced pressure (VLC) using silica gel 60 as stationary phase and eluted with hexane (Hex.), ethyl acetate (EtOAc), and methanol (MeOH) alone or in binary mixtures increasing polarity.2O, H2O:MeOH (7:3), H2O:MeOH (1:1), MeOH, Hex, acetone, and ethyl acetate (AcOEt). Fractions of MeOH (162 mg) and H2O:MeOH (7:3) (475 mg) were fractioned on a Sephadex LH-20 (MeOH) leading to isolation of compounds 4 (5 mg) and 5 (10 mg).An sample of the MeOH phase (10 g) was submitted to CC on XAD-2 using the eluents: H2O (9:1) and submitted to liquid-liquid chromatography using Hex, CH2Cl2, EtOAc and n-butanol, obtaining the respective phases. The CH2Cl2 phase (26 g) was chromatographed on VLC (silica gel 60) eluted with CH2Cl2 and MeOH alone and in mixtures increasing polarity. The fraction CH2Cl2:MeOH (9:1) (7 g) was chromatographed on silica gel CC using the same solvents. The fraction 66/74 (85 mg) was purified on a Sephadex LH-20 (MeOH) yielding compound 6 (23 mg).Another part of the CEE (200 g) was dissolved in EtOH–H2 and 5% CO2), and kept at pH 7.4. All preparations were stabilised under a resting tension of 0.75 g for 1 h. The solution was replaced every 15 min in order to prevent the accumulation of metabolites. The force of contraction was isometrically recorded by a force transducer  coupled to an amplifier-recorder (Miobath-4) and to a computer equipped with an analog-to-digital converter board as described earlier. The presence of functional endothelium was assessed by the ability of acetylcholine (10 µM) to induce more than 80% relaxation of vessels pre-contracted with phenylephrine (10 µM). Less than 10% of relaxation to acetylcholine was taken as evidence that the vessel segments were functionally denuded of endothelium [−12–10−3 M) added to the preparations until a maximum response was observed as indicated by a plateau response (approximately 3–5 min).The experiments were approved by the Federal University of Paraíba Ethical Committee for Animal Use under the protocol CEPA#305/09. After animal euthanasia by decapitation using a guillotine, the cranial mesenteric artery was isolated, placed in Tyrode’s solution and dissected in order to make it free of adhering tissue. In endothelium-denuded experiments, endothelium was removed by rubbing the intimal surface of the vessels. Rings with 1–2 mm were obtained and placed in physiological Tyrode’s solution, maintained to 37 °C, gassed with carbogenic mixture .The results were expressed as mean ± standard error of the mean (SEM). Differences between means were considered significant when S. rhombifolia aerial parts, using chromatographic and spectroscopic techniques, led to the identification of eight substances, among them two coumarins: scopoletin (1) and escoporone (2); a ferulic acid derivative, ethoxy-ferulate (3); two flavonoids: kaempferol (4) and kaempferol-3-O-β-d-glycosyl-6′′-α-d-rhamnose (5); and three indoquinoline alkaloids: quindolinone (6), 11-methoxy-quindoline (7) and quindoline (8). Compound 3 is being first reported in the Sida genus, and others have been previously reported strengthening Sida chemosystematics [The study of tematics ,24,25.6) and the salt of cryptolepine (9) induced vasorelaxation dependent on the vascular endothelium, justifying the use of the species in popular medicine in India. The vasorelaxation effect may be related to indoquinoline core, but more protocols are required to elucidate their complete mechanism of action.The quindolinone (