Aloe megalacantha led to the isolation of four new natural products—1,8-dimethoxynepodinol (1), aloesaponarin III (2), 10-O-methylchrysalodin (3) and methyl-26-O-feruloyl-oxyhexacosanate (4)—along with ten known compounds. All purified metabolites were characterized by NMR, mass spectrometric analyses and comparison with literature data. The isolates were evaluated for their cytotoxic activity against a human cervix carcinoma cell line KB-3-1 and some of them exhibited good activity, with aloesaponarin II (IC50 = 0.98 µM) being the most active compound.Phytochemical investigation of the ethyl acetate extract of the roots of  Aloe is the largest genus in the Asphodelaceae family and comprised of more than 400 species +m/z 283.0916), indicated eight degrees of unsaturation. The UV  and IR  spectra revealed absorptions for conjugated ketone and aromatic moieties. The 13C-NMR spectrum , a methyl  and two methoxy  substituents were also evident. The 1H-NMR spectrum , 7.44  and 7.40 , corresponding to H-5, H-6, and H-7, respectively.Compound spectrum  revealedspectrum  showed tH 3.99, δC 56.3) is present at C-8 (δC 157.0) according to its long range HMBC correlation (C 106.7) and C-8 (δC 157.0). A broad singlet aromatic proton at δH 7.48 (H-4) showed a long-range NOE interaction in the NOESY spectrum with H-5 (δH 6.97) and the methyl group  located as expected according to biosynthetic considerations at C-3 (δC 133.6). The substituents at C-1 (δC 154.8) and C-2 (δC 132.0) were, therefore, established to be the methoxy  and the hydroxymethylketone unit  respectively, based on a 3JC,H HMBC correlation of both H-4 (δH 7.48) and the hydroxymethyl protons (δH 4.55) with C-2 (δC 132.0). These placements were further confirmed by the long range HMBC correlations observed between methoxy protons (δH 3.73) and its nearby C-1 and C-2 carbons. Interestingly, the down-field shifted signal at δC 207.5 for the aryl ketone in the 13C-NMR spectrum is consistent with a group being ortho-disubstituted, resulting in a deviation from planarity as a result of steric repulsion. Therefore, based on the above spectroscopic evidence, the new compound was characterized as 2-hydroxymethylacetyl-1,8-dimethoxy-3-methylnaphthalene, which was given the trivial name 1,8-dimethoxynepodinol (1).One of the methoxy groups  and NMR spectra suggested an anthraquinone skeleton + (m/z = 507.1034) was observed similar to the related dimeric anthraquinones +) and HR-MS (m/z 603.5395 (for [M + H]+) along with the NMR data was compatible with the molecular formula C37H62O6. It has a UV absorption at λmax 213, 278 nm. Its IR spectrum indicated the presence of hydroxyl (3410 cm−1) and conjugated carbonyl (1637 cm−1) functionalities. The 1H-NMR spectrum (in CDCl3) exhibited signals corresponding to a ferulic acid moiety with a pair of doublets at δH 6.29 and 7.61 with a vicinal coupling constant 3JHH = 15.9 Hz. This indicated a trans configuration and the signals were assigned to H-2′ and H-3′, respectively. These protons showed HMBC correlations with the ester carbonyl carbon at δC 167.5, suggesting that these protons were part of an α,β-unsaturated ester moiety. The three aromatic proton signals at δH 7.03 (JHH = 1.9 Hz), 6.92 (JHH = 8.2 Hz) and 7.07  were assigned to H-5′, H-8′, and H-9′, respectively based on their HH-COSY interactions and coupling constants. These assignments were further supported by the HMBC correlations of the downfield shifted proton H-3′ (δH 7.61) with C-5′ (δC 109.4) and C-9′ (δC 123.2) that in turn have HMQC correlations with H-5′ and H-9′, respectively. The 1H-NMR spectrum further showed the presence of a singlet at δH 5.84 corresponding to a phenolic hydroxy group, two singlets integrating for three protons each at δH 3.66 and 3.93 for two methoxy groups, two triplets at δH 4.19 and 2.30 for methylene groups  and multiplets corresponding to long methylene chain. The 13C-NMR spectrum contains signals corresponding to two ester carbonyl carbons , five aromatic/olefinic methine groups , two oxygenated aromatic quaternary carbons , one oxymethylene (δC 64.8), two methoxy  and a number of methylene (34.3–24.9) carbons. The position of one methoxy group  was established at C-6′ (δC 148.0) according to its HMBC correlation to C-6′, while the hydroxy group at δH 5.84 showed a correlation to C-7′ (δC 146.9). The other methoxy group  corresponds to a methyl ester deduced from its upfield shifted (δC 51.6) signal [C 174.5). Comparison of these spectroscopic data with the literature revealed that the compound was very similar to ω-feruloyloxyacid [m/z 625 [M + Na]+, 601 [M−H]−) were in accordance with the presence of a ω-oxygenated C26 fatty acid chain, like ω-feruloyloxyacids with along aliphatic chain (C19–C27), reported from peat soil + . 13C-NMR . ESI-MS Aloesaponarin III (2): Yellow solid. m.p. 257–259 °C. UV (CH3CN): λmax (logε) = 264 (2.80), 298 (2.73), 336 (2.63), 371 (2.44) nm. IR (CH2Cl2) νmax cm−1 3363, 2948, 1732, 1662, 1577, 1235, 712.1H- and 13C-NMR , 319 , 297 . HR-ESI-MS m/z = 297.0753, [M + H]+. 13C-NMR . ESI-MS 10-O-Methylchrysalodin (3): Yellow amorphous solid. m.p. 228–230 °C. UV (CH2Cl2): λmax (logε) = 215 (3.18), 261 (3.14), 384 (2.94), 432 (2.66) nm. 2Cl2). 1H- and 13C-NMR , 537 . HR-ESI-MS m/z = 537.1863, [M − H]− . 13C-NMR . ESI-MS Methyl 26-O-feruloyl-oxyhexacosanoate (4): Colourless solid. m.p. 187–189 °C. UV (CH2Cl2): λmax (logε) = 213 (2.91), 278 (2.87) nm. IR (CH2Cl2) νmax cm−1 3410, 1762, 118, 1637. 1H-NMR (CDCl3) δH 7.61 , 7.07 , 7.03 , 6.92 , 6.29 , 5.84 , 4.19 , 3.93 , 3.66 , 2.30 , 1.18–1.23 . 13C-NMR (CDCl3) δC 174.5 (C-1), 167.5 (C-1′), 148.0 (C-6′), 147.9 (C-7′), 144.8 (C-3′), 127.2 (C-4′), 123.2 (C-9′), 118.8 (C-8′), 115.8 (C-2′), 109.4 (C-5′), 64.8(C-26), 56.1 (6′-OCH3), 51.6 (1-OCH3), 34.3 (C-2), 34.1–24.9 (methylene chain). ESI-MS (rel.int.) m/z = 625 . ESI-MS m/z = 601, [M − H]−. HR-ESI-MS m/z = 603.5395, [M + H]+ .l-glutamine, sodium pyruvate and phenol red, supplemented with 10% fetal bovine serum (FBS) and were maintained at 5.3% CO2 and 37 °C in humidified air. The cells at 70% confluence were detached with trypsin-ethylenediamine tetraacetic acid solution  and placed in sterile 96-well plates in a density of 10,000 cells in 100 μL medium per well. The dilution series of the compounds was prepared from stock solutions in DMSO of concentrations of 100 mM, 50 mM or 25 mM and the stock solutions were diluted with culture medium (10% FBS) down to pM concentrations. The dilution prepared from stock solution was added to the wells and each concentration was tested in at least six replicates. The control contained the same concentration of DMSO as the first dilution. After incubation for 72 h at 37 °C and 5.3% CO2-humidified air, 30 μL of an aqueous resazurin solution (175 μM) was added to each well. The cells were incubated at the same conditions for 5 h. Subsequently, the fluorescence was recorded at a wavelength of 588 nm. The IC50 values were calculated as a sigmoidal dose response curve using Graphpad Prism 4.03 .The human cervix carcinoma cell line KB-3-1 was used in the cytotoxicity assay as previously described . The celA. megalacantha. The two aloesaponarins  showed good cytotoxic activity even greater than one of the reference drug (griseofulvin) against KB-3-1 human cervical cancer cell line, with the highest activity observed for aloesaponarin II.Four new natural products along with ten other known compounds were identified from the root of


N-methyl-D-aspartate (NMDA) receptor in the central nervous system (CNS). We have reported that low doses of ketamine administrated to patients before incision significantly reduced post-operative inflammation as reflected by reduced interleukin-6 (IL-6) sera-levels. Our data demonstrated in a rat model of Gram-negative bacterial-sepsis that if we inject a low dose of ketamine following bacterial inoculation we reduce mortality from approximately 75% to 25%. Similar to what we have observed in operated patients, the levels of TNF-α and IL-6 in ketamine-treated rats were significantly lower than in septic animals not treated with ketamine. On the base of these results, we have designed and synthesized series of new analogues of ketamine applying a thermal rearrangement of alicyclic α-hydroxyimines to α-aminoketones in parallel arrays. One of the analogues (compound 6e) displayed high activity in down-regulating the levels of IL-6 and TNF-α in vivo as compared to ketamine.Ketamine is an analgesic/anesthetic drug, which, in combination with other drugs, has been used as anesthetic for over 40 years. Ketamine induces its analgesic activities by blocking the  Uncontrolled inflammation and immune response lie at the heart of bacterial sepsis. Sepsis is a major complication among post-operative and trauma patients which leads to multi-organ dysfunction, multisystem failure and eventually to death in up to 60% of septic patients in chirurgical intensive care units . The sepN-methyl-D-aspartate (NMDA) receptor in the central nervous system (CNS) , 7[a–g], 6'[a–g] and 7'[a–g] in a one pot reaction using both classical or microwave heating of mixtures of the corresponding primary amines R2-NH2 and the different hydroxyalcohols 3a,b. In prior works, this type of process needed high temperatures (200 °C) and long reaction times (10–24 h) that were also employed here.Our approach is described in 4[a–g], 5[a–g] for the thermal rearrangement were hypothesized (but not isolated) by Stevens  were conducted using microwave assisted reactions. A reduced model was chosen using Rnditions .6 and 6' after short reactions. The products were easily separated by preparative HPLC. Interestingly, yields varied for the different amines, while using classical heating method an overall yield of 48% was observed for 6a/6'a mixtures, the use of microwave improved the yield to 90% with a different distribution of isomers. On the other hand, hexyl-  and propyl-  amine that gave relatively high yields using classical heating, led to reduced yields with microwave energy. Finally, while reacting aniline (6f and 6'f), yields were similarly low for both methods, but in contrast to classical heating that resulted in 1:1 isomer mixtures, the microwave reaction led exclusively to isomer 6'f. vs. microwave heating methods.To our surprise we could obtain separable mixtures of compounds 8 of synthetic interest has been isolated from the microwave assisted reaction using benzylamine in 8% yield (see 6a-6'a]). This compound appears to be the result of aromatization of the cyclohexane ring followed by the thermal migration (rearrangement) of the N-benzyl group to the para position in the generated aromatic system in a similar way to the previously reported photo- , 3–20 min [50/50], 20–30 min [0/100], 30–40 min [0/100], 41 min [100/0]; flow, 1 mL/min. column Chromolith Performance RP-18e 100–4.6 mm, gradient H2O/MeCN, Method B: 1 min [100/0], 1–8 min [0/100], 8–11 min [0/100], 11.1 min [100/0]; flow, 6 mL/min. Preparative: column C18 Vydac-218TP-101510 or column C18 Phenomenex 58 × 21.2 × 10 mm, gradient H2O/MeCN. Method D: 3 min [100/0], 3–25 min [50/50], 25–35 min [0/100], 35–50 min [0/100], 51 min [0/100]; flow, 15 mL/min. Method E: 18 min [100/0], 18–40 min [50/50], 40–50 min [0/100], 50–65 min [0/100], 66 min [0/100]; flow, 15 mL/min. Method F: 3 min [100/0], 3–20 min [50/50], 20–30 min [0/100], 30–60 min [0/100], 61 min [0/100]; flow, 15 mL/min.Reagents, unless otherwise mentioned, were purchased from Aldrich and used without further purification. 1,2-Cyclohexanedione was purchased from Acros, Tetrahydrofuran (THF) and diethyl ether  hexane and (B) EtOAc. Separation conditions are as follows: Preparative: column GL Sciences Inc-2GI95001 Inertsil PREP-SIL µm 50 × 250 mm, manual gradient hexane/EtOAc, Method G: 100% hexane until 2:1 hexane/EtOAc ; flow, 70 mL/min.Microwave irradiations were carried out with a professional “Initiator” microwave from Biotage at pre-fixed temperature. The intensity differed from 0 to 300W at frequency of 2.456 GHz. 1H and 13C-NMR spectra were recorded on Bruker DPX-300 and Avance DMX-600 spectrometers. 1H-NMR data was obtained by 2D, COSY and HOHAHA methods (t = 40 ms). 13C-NMR data was obtained by 2D techniques, NOESY, HMQC and HMBC methods with a delay of 3.45 60 ms respectively in the reverse mode. Chemical shifts are in ppm relative to TMS internal standard or relative to the residual solvent resonance.4) and by ESI (electron spray ionization) mass spectrometry on a Q-TOF (quadropole time of flight) low-resolution micromass spectrometer . Microwave reactions were performed using a mono-mode Initiator station from Biotage. Purity of compounds was over 95% as assessed by two different HPLC conditions, QTOF-MS-MS was used for assessing purity of the molecular peaks. Mass spectra analyses were recorded on an AUTOSPEC-FISSONS VG (Micromass) high-resolution mass spectrometer under DCI  conditions : A solution of 1,2-cyclohexanedione  in toluene (300 mL), an equimolar amount of ethylene glycol  and p-toluenesulfonic acid (100 mg) were heated at reflux for 8 h in a Dean-Stark apparatus (water was continuously separated). The resulting solution was washed twice with 1 N NaOH solution, dried over MgSO4 and concentrated under reduced pressure to afford 9.61 g  of the crude yellow oily product. The crude product containing a certain amount of the diacetal 1′ , was used without further purification in the next step. 1H-NMR  δ 3.93 , 2.47 , 1.44–1.91 ; 13C-NMR  δ 206.19 (C-1), 106.62 (C-2), 65.05 , 39.51 (C-6), 36.77 (C-3), 26.07 (C-5), 22.55 (C-4); MS(CI+): 157 (MH+); 128 (M-CO). HRMS: m/z calc. for C8H12O3 (MH+) 157.0865; found 157.0835.Preparation of 6-phenyl-1,4-dioxaspiro[4.5]decan-6-ol (2): To a solution of 1 containing 1 mmol of monoketone in dry THF (25 mL) at −78 °C under N2 was added dropwise a solution of phenylMgBr (2 mmol in 50 mL ether) The mixture was stirred at −78 °C and then was warmed to room temperature with stirring overnight. The reaction mixture was quenched by the addition of saturated aqueous NH4Cl. The organic layer was removed, and the aqueous layer was extracted twice with ether. The combined organic layers were washed with brine, dried over MgSO4, and concentrated under reduced pressure. The residue was purified by preparative normal phase-HPLC  to give 4.27 g of the desired product as a white solid (60% yield). HPLC analysis Rt = 26.0 min ; 1H-NMR  δ 7.57 , 7.29 , 7.24 , 3.69 , 3.64 , 3.34 , 2.83 , 2.65 , 2.30 , 2.12 , 1.79 , 1.78 , 1.74 , 1.64 , 1.59 , 1.55 ; 13C-NMR  δ 143.83 (C-1'), 127.22 (C-3'), 127.05 (C-2'), 126.68 (C-4'), 110.74 (C-1), 76.70 (C-2), 65.52 (C-7"), 65.09 (C-7'), 35.72 (C-3), 32.72 (C-6), 23.26 (C-5), 20.61 (C-4); MS(CI+): 234 (M+); 217 (M-OH+). HRMS: m/z calc. for C14H18O3 (M+) 234.1256; found 234.1240.Preparation of 2-hydroxy-2-phenylcyclohexanone (3): Compound 2 (0.15 mmol) was treated with aqueous 80% TFA (3 mL) at 0 °C. The reaction mixture was stirred for 3 h at room temperature and then treated with aqueous saturated NaHCO3 solution at 0 °C until it became alkaline. The resulting mixture was extracted with CH2Cl2 (×2). The organic layer was washed with brine, dried over MgSO4, and concentrated under reduced pressure to give the desired product as a yellow oil (86–98% yield); HPLC analysis Rt = 22.8 min ; 1H-NMR  δ 7.27 , 2.91 , 2.40 , 1.57–2.04 ; 13C-NMR  δ 211.59 (C-1), 138.92 (C-1'), 128.12 (C-3'), 127.30 (C-4'), 125.37 (C-2'), 79.04 (C-2), 37.88 (C-6), 37.82 (C-3), 27.31 (C-5), 22.04 (C-4); MS(CI+): 190 (M+); 173 (M+-OH); 162 (M+-CO). HRMS: m/z calc. for C12H14O2 (M+) 190.0994; found 190.0984.Preparation of 6-methyl-1,4-dioxaspiro[4.5]decan-6-ol (2′): Prepared similarly to 2, but using MeMgBr as Grignard reagent. The residue was purified by silica gel column chromatography  to give 1.88 g of the desired product as a yellowish oil (60.5% yield). 1H-NMR  δ 4.00 , 2.19 , 1.82 , 1.36–1.67 , 1.21 ; 13C-NMR  δ 111.13 (C-1), 73.91 (C-2), 65.60 (C-7"), 65.39 (C-7'), 37.85 (C-6), 31.57 (C-3), 23.32 (C-5), 22.49 (Me), 21.74 (C-4); MS(CI+): 172 (M+); 155 (M+-OH). HRMS: m/z calc. for C9H16O3 (M+) 172.1099; found 172.1084.Preparation of 2-hydroxy-2-methylcyclohexanone (3′): Prepared similarly to 3 but starting from 2’. Yellow oil (82–91% yield); 1H-NMR  δ 3.83 , 2.54 , 2.11 , 1.61–1.85 , 1.41 ; 13C-NMR  δ 214.37 (C-1), 76.46 (C-2), 42.05 (C-6), 37.78 (C-3), 27.88 (C-5), 25.07 (Me), 23.01 (C-4); MS (CI+): 129 (MH+); 111 (M-OH). HRMS: m/z calc. for C7H12O2 (MH+) 129.0916; found 129.0928.3, 190 mg, 1 mmol) was added the appropriate primary amine . The tubes were flushed with argon, sealed and heated at 200 °C overnight in a Radley’s combinatorial station with stirring. The residues were diluted with ether and extracted with 0.1 M HCl. The acidic phase washed with ether and concentrated and the combined ether phase was dried over MgSO4 and concentrated. Preparative HPLC conditions were selected among methods D-F, according to the elution times obtained by analytical HPLC using method A.To seven pressure-tubes each containing 2-hydroxy-2-phenylcyclohexanone -2-phenylcyclohexanone (6a): White solid (36% yield); HPLC analysis Rt = 22.3 min ; 1H-NMR , 7.54–7.67 , 7.54–7.67 , 7.46–7.56 , 7.46–7.56 , 7.31–7.42 , 3.78 , 3.74 , 3.33 , 2.54 , 2.08–2.22 , 2.08–2.22 , 1.96–2.00 , 1.73–1.88 , 1.73–1.88 ; 13C-NMR  δ 206.96 (C-1), 132.43* (C-1′′), 131.98* (C-4′), 131.51* (C-1′), 131.29* (C-3′), 131.17* (C-3′′), 130.59* (C-4′′), 130.14* (C-2′′), 129.90* (C-2′), 74.28 (C-2), 47.90 (CH2-Bn), 40.47 (C-6), 34.38 (C-3), 28.74 (C-5), 22.80 (C-4); MS(CI+): 280 (MH+); 91 (C7H7+). HRMS: m/z calc. for C19H21NO (MH+) 280.1701; found 280.1734.(1-(Benzylamino)cyclopentyl)(phenyl)methanone (6′a): White solid (12% yield); HPLC analysis Rt =23.7 min ; 1H-NMR  δ 8.23 , 7.43 , 7.52 , 7.21 , 7.19 , 7.05 , 3.49 , 2.40 , 1.90 , 1.80 , 1.69 ; 13C-NMR  δ 204.30 (COPh), 140.24 (C-1′′), 136.28 (C-1′), 132.17 (C-4′), 129.33 (C-2′), 128.30 (C-3′′), 128.22 (C-2′′), 128.10 (C-3′), 126.99 (C-4′′), 75.17 (C-1), 49.32 (CH2-Bn), 36.68 (C-2), 24.66 (C-3); MS(CI+): 280 (MH+); 174 (M−COPh)+. HRMS: m/z calc. for C19H21NO (MH+) 280.1701; found 280.1726.6b-6′b: Hexylamine was used as amine component in the reaction. The mixture was purified by preparative HPLC using method D to give the desired products.Synthesis of 2-(Hexylamino)-2-phenylcyclohexanone (6b): Colorless oil (38% yield); HPLC analysis Rt = 25.3 min ; 1H-NMR  δ 7.49–7.61 , 7.49–7.61 , 7.49–7.61 , 3.29 , 2.72 , 2.47 , 2.47 , 1.96–2.16 , 1.96–2.16 , 1.96–2.16 , 1.82 , 1.82 , 1.59 , 1.28 , 1.28 , 1.28 , 0.88 ; 13C-NMR  δ 207.00 (C-1), 131.77 (C-4′), 131.49 (C-1′), 131.14 (C-3′), 129.59 (C-2′), 73.48 (C-2), 43.47 (C-1′′), 40.24 (C-6), 33.61 (C-3), 32.27 (C-2′′), 28.60 (C-5), 27.24 (C-3′′), 27.20 (C-4′′), 23.32 (C-5′′), 22.74 (C-4), 14.20 (C-6′′); MS(CI+): 274 (MH+); 245 (M-Et)+; 174 (M−NHHex)+. HRMS: m/z calc. for C18H27NO (MH+) 274.2170; found 274.2175.(1-(Hexylamino)cyclopentyl)(phenyl)methanone (6'b): Yellowish oil (13% yield); HPLC analysis Rt = 26.3 min ; 1H-NMR  δ 7.86 , 7.66 , 7.55 , 2.96 , 2.61 , 2.15 , 2.15 , 1.77 , 1.41 , 1.41 , 1.41 , 0.94 ; 13C-NMR  δ 199.81 (COPh), 135.06 (C-1′), 134.59 (C-4′), 130.02 (C-2′), 129.87 (C-3′), 79.34 (C-1), 45.63 (C-1′′), 36.31 (C-2), 32.48 (C-2′′), 27.99 (C-3′′), 27.31 (C-4′′), 26.70 (C-3), 23.50 (C-5′′), 14.30 (C-6′′); MS(CI+): 274 (MH+); 168 (M-COPh)+. HRMS: m/z calc. for C18H27NO (MH+) 274.2170; found 274.2184.6c-6′c]: n-Propylamine was used as amine component in the reaction. The mixture was purified by preparative HPLC using method D to give the desired products.Synthesis of : i-Propylamine was used as amine component in the reaction. The mixture was purified by preparative HPLC using method D to give the desired products.Synthesis of : n-Butylamine was used as amine component in the reaction. The mixture was purified by preparative HPLC using method D to give the desired products.Synthesis of : Aniline was used as amine component in the reaction. The mixture was purified by preparative HPLC using method F to give the desired products.Synthesis of : 4-Chloroaniline was used as amine component in the reaction. The mixture was purified by preparative HPLC using method F to give the desired products.Synthesis of : n-Hexylamine was used as amine component in the reaction. The residue was purified by silica gel column chromatography  to give the desired product. Synthesis of : n-propylamine was used as amine component in the reaction. The residue was purified by silica gel column chromatography  to give the desired products. Synthesis of : iso-propylamine was used as amine component in the reaction. The residue was purified by silica gel column chromatography  to give product [7d]. Synthesis of : n-butylamine was used as amine component in the reaction. The residue was purified by silica gel column chromatography  to give the desired products. Synthesis of : aniline was used as amine component in the reaction. The organic phase was purified by preparative reverse-HPLC (method D) to give the product [7'f].Synthesis of : 4-Cl-aniline was used as amine component in the reaction. The organic phase was purified by preparative reverse-HPLC (method D) to give the product [7'g].Synthesis of + compound 8: To a microwave flask containing 95 mg (0.5 mmol) of 2-hydroxy-2-phenylcyclohexanone 3, was added 0.575 mmol of benzylamine and NMP (1.5 mL). The flask was flushed with argon and sealed. The reaction mixture was heated at 230 °C for 10 min . The residue was diluted with ten folds of water and TFA was added until clearness. The obtained solution was filtered and purified by preparative reverse-HPLC (method E).Synthesis of -2-amine (8). (8% yield). HPLC analysis Rt = 4.8 min ; 1H-NMR  δ 7.41–7.43 , 7.32 , 7.27 , 7.21 , 7.17 , 6.98 , 6.97 , 6.72 , 3.91 , 3.60 ; 13C-NMR  δ 144.75 (C-1"), 144.28 (C-1), 139.46 (C-1'), 131.55 (C-4), 130.90 (C-3), 129.09 (C-3'), 128.93 (C-5), 128.81* (C-2"), 128.75* (C-2'), 128.39 (C-3'), 127.91 (C-2), 127.13 (C-4'), 125.88 (C-4"), 116.02 (C-6), 41.02 (CH2-Bn); MS(CI+): 260 (MH+); 259 (M+). HRMS: m/z calc. for C19H17N (M+) 259.1361; found 259.1368.6b-6′b]: To a microwave flask containing 95 mg (0.5 mmol) 2-hydroxy-2-phenylcyclohexanone 3, was added 0.575 mmol of hexylamine and NMP (1.5 mL). The flask was flushed with argon and sealed. The reaction mixture was heated at 230 °C for 20 min . The residue was diluted with ten folds of water and TFA was added to clearness. The resulting solution was filtered and purified by preparative reverse-HPLC (method E).Synthesis of : To a microwave flask containing 95 mg (0.5 mmol) 2-hydroxy-2-phenylcyclohexanone 3, was added 0.575 mmol of propylamine and NMP (1.5 mL). The flask was flushed with argon and sealed. The reaction mixture was heated at 230 °C for 30 min . The residue was diluted with ten folds of water and TFA was added to clearness. The resulting solution was filtered and purified by preparative reverse-HPLC (method E).Synthesis of : To a microwave flask containing 95 mg (0.5 mmol) 2-hydroxy-2-phenylcyclohexanone 3, was added 0.575 mmol of aniline and NMP (1.5 mL). The flask was flushed with argon and sealed. The reaction mixture was heated at 230 °C for 30 min . The residue was diluted with twenty fold of saturated NaHCO3 solution and extracted with EtOAc. The organic phase was concentrated, diluted with ether and extracted with 0.1 M HCl. The acidic phase was washed with ether and concentrated. The residue was purified by preparative reverse-HPLC (method E).Synthesis of [phenyl(1-(phenylamino)cyclopentyl)methanone (6'f). Yellow-brown solid (14% yield); HPLC analysis Rt = 5.4 min ; 1H-NMR  δ 8.07 , 7.46 , 7.35 , 7.11 , 6.80 , 6.80 , 6.66 , 2.54 , 2.09 , 1.84 ; 13C-NMR  δ 203.72 (COPh), 143.07 (C-1′′), 135.57 (C-1′), 132.68 (C-4′), 129.45 (C-2′), 129.15 (C-3′′), 128.33 (C-3′), 120.60 (C-4′′), 116.41 (C-2′′), 74.92 (C-1), 37.47 (C-2), 25.08 (C-3); MS(ES+): 266 (MH+). HRMS: m/z calc. for C18H19NO (M+•) 265.1467; found 265.1483.Physical properties of Escherichia coli (E. coli). Inoculation of animals with live bacteria has been a common tool for studying sepsis mechanisms and represents a model relevant to clinical practice [E. coli-induced sepsis. This effect was attributed to the anti-inflammatory activity of ketamine, as evidenced by decreased IL-6 and TNF-α levels in serum and peritoneal lavage of treated animals [To examine the protective effect of the synthesized ketamine derivatives, we used a sepsis model and examined the inflammatory response of treated animals. Sepsis was induced in mice by intra-peritoneal (i.p.) inoculation of a lethal dose of practice ,35. Intr animals ,13. To d animals . The ELICD1 female mice aged 10 to 12 weeks  were maintained in the animal laboratory of the Soroka Medical Center. Experiments were done with the permission of the Ben-Gurion University of the Negev Committee for Ethical Care and Use of Animals in Experiments . E. coli were grown in Luria-Bertani broth  and harvested during the log phase. Bacteria aliquots were stored frozen in LB broth containing 30% glycerol. Ketamine was purchased from Parke Davis .E. coli (3.6 × 109 colony forming unit). Prior to bacteria inoculation mice were injected subcutaneously with 200 µL of ketamine (10 mg/kg), its analogues (10 mg/kg) or saline.Ketamine analogues were reconstituted in saline solution to a concentration of 1.25 mg per mL. Sepsis was induced in mice by intra-peritoneal (i.p.) inoculation of a lethal dose of E. coli inoculation, animals were anesthetized with sodium pentobarbital . A 1 mL syringe flushed with heparin was used to draw intra-cardial blood sample. The samples were stored on ice before centrifugation at 1,000 g at 40 °C for 10 minutes. The cell-free supernatants were collected and frozen at −200 °C for future analysis. Peritoneal lavage was performed with 5 mL phosphate buffer saline  containing 2% bovine serum albumin  and 5 mM ethylenediaminetetraacetic acid . After centrifugation at 400 g for 10 minutes, the cell-free supernatants were removed and frozen at −20 °C until analysis. TNFα and IL-6 levels were determined by commercial ELISA kits .At 16 hours after Results are expressed as mean ± S.E.M. To compare levels between treated groups and saline group, one-way analysis of variance was used. P values of <0.05 were considered significant.in vivo. Analog 6e was shown to downregulate the levels of IL-6 and TNF-α  in vivo, which correlates with a normal behavior of mice treated with ketamine in contrast to a hunched posture, ruffled fur and reduced activity in mice treated with inactive compounds or saline. Compound 6e with a statistically significant improved effect on reduction of IL-6 and TNF-α secretion upon E. coli inoculation as compared to ketamine is a good candidate for the treatment of sepsis. Toxicology and other drug-safety in vivo studies are on-going.We have designed and synthesized libraries of new analogues of ketamine as potential anti-sepsis agents. The synthetic strategy included a thermal rearrangement of alicyclic α-hydroxyimines to five- and six-membered α-aminoketone rings. The use of microwave energy is advantageous in terms of time reactions, but gives ambiguous results in terms of yields. We have confirmed that a α-hydroxy-imine hypothesized by Stevens 40 years ago, is the intermediate of the thermal rearrangement between the five- and six-membered ring ketones. We speculate that the five-membered ring compounds are the kinetic products while the six-membered rings are the thermodynamic products. The compounds were assayed for IL-6 and TNF-α regulation


Galinsoga parviflora herb (GP) in some aspects of the endothelial cell function necessary for anti-inflammatory activity and wound healing and relate these to the GP phytochemical profile. This study demonstrated that the GP extract caused a dose-dependent reduction of IL-6 secretion on IL-1β-stimulated endothelial cells. The IL-6 release was decreased to 33% ± 9% while this did not influence the IL-6 secretion without stimulation. Additionally, the GP extract exhibited an anti-hyaluronidase activity (IC50 = 0.47 mg/mL), which was evidently stronger than the positive control kaempferol (IC50 = 0.78 mg/mL) as well as a moderate and concentration-dependent, antioxidant activity. The results of the scratch assay showed that exposure of the endothelial cells to GP induced complete healing of the damage after 12 h of the study. The phytochemical profile of the extract was studied by using spectrophotometric  and UPLC (phenolic acids) methods. The main compound in the GP extract was a chlorogenic acid (2.00 ± 0.01 mg/g by UPLC). The total content of polyphenols was 98.30 ± 0.14 mg of chlorogenic acid equivalent/g of the dry herb and content of flavonoids amounted to 6.15 ± 0.41 mg quercetin equivalent/g of the dry herb. Moreover, the presence of flavonoids in G. parviflora was provided after their isolation and identification by spectroscopic methods. In conclusion, it demonstrated that application of GP in the treatment of skin lesions gives possibility of wound healing based on antioxidant, anti-inflammatory, and hyaluronidase-inhibiting activities of G. parviflora herb extract.The objective of this study was to evaluate the usefulness of a hydroalcoholic extract from  Inflammation is a key protective response to tissue injury. If uncontrolled, however, it can produce some collateral damage and contribute to further tissue destruction. The inflammatory reaction is regulated at many levels by cytokines, eicosanoids, adhesion molecules, reactive oxygen, and nitrogen species. Pharmacological attempts to curtail inflammation include cytokine inhibitors, soluble receptors or antagonists, antioxidants, and pro-resolving mediators.Many medicinal plants used successfully in folk medicine display anti-inflammatory properties, albeit the exact molecular mechanisms by which they produce these beneficial effects are often poorly defined. It was proven that they cause transcriptional or post-transcriptional control of gene expression, inhibition of pro-inflammatory cytokines and eicosanoids, and/or antioxidant activity .Galinsoga parviflora Cav.) (GP) is a cosmopolitan annual herb from the Asteraceae family. Fresh leaves and juice of GP have been used in folk medicine throughout the world to treat dermatological disorders including eczema, lichen, and non-healing and/or bleeding wounds −, m/z 317 [A − H]−; ESI-MS positive: m/z 481 [M + H]+, m/z 319 [A + H]+.Compound 2 (quercetin 7-O-β-glucoside = quercimeritrin): ESI-MS negative: m/z 463 [M − H]−, m/z 301 [A − H]−; ESI-MS positive: m/z 465 [M + H]+, m/z 303 [A + H]+.1H-NMR (DMSO-d6) δ (ppm): 6.41 , 6.76 , 7.72 , 6.89 , 7.55 , 5.07 , 13C-NMR (DMSO-d6) δ (ppm): 147.90 (C-2), 136.09 (C-3), 176.01 (C-4), 160.34 (C-5), 98.73 (C-6), 162.66 (C-7), 94.23 (C-8), 155.71 (C-9), 104.64 (C-10), 121.80 (C-1′), 115.56 (C-2′), 145.05 (C-3′), 147.56 (C-4′), 115.34 (C-5′), 120.02 (C-6′), 99.85 (C-1′′), 73.10 (C-2′′), 77.13 (C-3′′), 69.53 (C-4′′), 76.38 (C-5′′), 60.59 (C-6′′).Compound 3 (patuletin-7-O-β-glucopyranoside = patulitrin): ESI-MS negative: m/z 493 [M − H]−, m/z 331 [A − H]−, ESI-MS positive: m/z 495 [M + H]+, m/z 333 [A + H]+.1H-NMR (CD3-OD) δ (ppm): 6.88 , 7.76 , 6.88 , 7.67 , 3.92 , 5.10 , 13C-NMR (DMSO-d6) δ (ppm): 147.69 (C-2), 135.80 (C-3), 176.13 (C-4), 151.04 (C-5), 131.80 (C-6), 156.36 (C-7), 93.84 (C-8), 151.39 (C-9), 104.96 (C-10), 121.84 (C-1′), 115.41 (C-2′), 145.06 (C-3′), 147.91 (C-4′), 115.58 (C-5′), 120.03 (C-6′), 60.32 (OCH3), 100.13 (C-1′′), 73.20 (C-2′′), 76.68 (C-3′′), 69.56 (C-4′′), 77.22 (C-5′′), 60.60 (C-6′′).Compound 4 (quercetin-3-O-β-glucopyranoside = isoquercitrin): ESI-MS negative: m/z 463 [M − H]−, m/z 301 [A − H]−, ESI-MS positive: m/z 465 [M + H]+, m/z 303 [A + H]+.1H-NMR (DMSO-d6) δ (ppm): 12.64 , 6.19 , 6.40 , 7.57 , 6.84 , 7.57 , 5.46 , 13C-NMR (DMSO-d6) δ (ppm): 156.12 (C-2), 133.29 (C-3), 177.40 (C-4), 161.21 (C-5), 98.68 (C-6), 164.26 (C-7), 93.51 (C-8), 156.31 (C-9), 103.90 (C-10), 121.12 (C-1′), 115.20 (C-2′), 144.81 (C-3′), 148.47 (C-4′), 116.16 (C-5′), 121.58 (C-6′), 100.85 (C-1′′), 74.08 (C-2′′), 76.49(C-3′′), 69.91 (C-4′′), 77.55 (C-5′′), 60.94 (C-6′′).Compound 5 (quercetin-3-O-β-glucuronide = miquelianin): ESI-MS negative: m/z 477 [M − H]−, m/z 301 [A − H]−; ESI-MS positive: m/z 479 [M + H]+, m/z 303 [A + H]+.1H-NMR (DMSO-d6) δ (ppm): 12.26 , 6.17 , 6.36 , 8.28 , 6.82 , 7.34 , 5.20 , 3.38 , 13C-NMR (DMSO-d6) δ (ppm): 157.61 (C-2), 134.05 (C-3), 177.57 (C-4), 160.92 (C-5), 98.95 (C-6), 164.92 (C-7), 93.78 (C-8), 156.50 (C-9), 103.55 (C-10), 120.61 (C-1′), 118.02 (C-2′), 144.79 (C-3′), 148.42 (C-4′), 115.40 (C-5′), 120.45 (C-6′), 103.08 (C-1′′), 73.95 (C-2′′), 76.63 (C-3′′), 71.78 (C-4′′), 74.16 (C-5′′), 172.20 (C-6′′).Compound 6 : ESI-MS negative: m/z 447 [M − H]−, m/z 285 [A − H]−, ESI-MS positive: m/z 449 [M + H]+, m/z 287 [A + H]+.1H-NMR (DMSO-d6) δ (ppm): 12.61 , 6.21 , 6.43 , 8.04 , 6.88 , 5.46 ; 13C-NMR (DMSO-d6) δ (ppm): 156.23 (C-2), 133.16 (C-3), 177.46 (C-4), 161.21 (C-5), 98.67 (C-6), 164.12 (C-7), 93.63 (C-8), 156.36 (C-9), 103.99 (C-10), 120.88 (C-1′), 130.88 (C-2′), 115,09 (C-3′), 159.94 (C-4′), 115.09 (C-5′), 130.88 (C-6′), 100.82 (C-1′′), 74.20 (C-2′′), 77.49 (C-3′′), 69.87 (C-4′′), 76.40 (C-5′′), 60.81 (C-6′′).Compound 7 (quercetin-3-O-α-rhamnopyranosylo (1→6)-β-glucopyranoside = rutin): ESI-MS negative: m/z 609 [M − H]−, m/z 463 [M-rhamnosyl-H]¯, m/z 301 [A − H]−; ESI-MS positive: m/z 611 [M + H]+, m/z 465 [M-rhamnosyl + H]+, m/z 303 [A + H]+. 1H-NMR (DMSO-d6) δ (ppm): 12.59 , 6.18 , 6.37 , 7.52 , 6.83 , 7.54 , 5.34 , 4.38 , 0.99 , 13C-NMR (DMSO-d6) δ (ppm):156.42 (C-2), 133.27 (C-3), 177.32 (C-4), 161.20 (C-5), 98.70 (C-6), 164.23 (C-7), 93.59 (C-8), 156.56 (C-9), 103.88 (C-10), 121.13 (C-1′), 115.20 (C-2′), 144.75 (C-3′), 148.43 (C-4′), 116.22 (C-5′), 121.57 (C-6′), 101.18 (C-1′′), 74.05 (C-2′′), 75.88 (C-3′′), 69.98 (C-4′′), 76.42 (C-5′′), 66.97 (C-6′′), 100.73 (C-1′′′), 70.35 (C-2′′′), 70.53 (C-3′′′), 71.82 (C-4′′′), 68.23 (C-5′′′), 17.72 (C-6′′′).Compound The stock solutions (1 mg/mL) of phenolic acids: chlorogenic, caffeic, gallic, isovanilic, 4-hydroxybenzoic, and protocatechuic were freshly prepared in methanol. The working solutions were prepared by diluting the stock solution using mobile phase A. 2.0 μL volume at each concentration level of each phenolic acids working solutions (5–40 µg/mL) was injected eight times into the UPLC system for the construction of the calibration curves.v/v) to 2.0 mL, and then purified through a syringe filter .The GP extract was added to the SPE process. The cartridge  was conditioned with 5 mL of methanol followed by 5 mL of grade water. 1.0 mL of the GP extract was eluted with 50 mL of 70% methanol. The organic eluate was collected in a flask and then concentrated under the vacuum at 40°C to dryness, diluted with mixture methanol-water  at the column temperature of 24 °C. Elution was conducted using mobile phase A: HPLC grade water and mobile phase B: HPLC grade acetonitrile. Each solvent (1000 mL) was acidulated by using formic acid (50 µL). The linear gradient was as follows: 3% to 13% B over 0.0 to 4.0 min, 13% to 17.5% B over 4.0 to 5.0 min, 17.5% B over 5.0 to 9.0 min, 17.5% to 3.0% B over 9.0 to 13.0 min with a flow rate of 0.275 mL/min. The spectral data of signals from the PDA detector were collected during the whole run in the range of 234 to 330 nm and processed by using Empower 2 Software Build 2154 . The presence of phenolic acids in the GP extract was confirmed by comparison of retention time and spectra of particular substances.Identification and quantification of phenolic acids of GP extract were carried out by using an ultra-high performance liquid chromatography . The method was performed on an Acquity UPLC HSS T3 column  with a guard pre-column of Acquity UPLC BEH CQuantitative analyses of phenolic acids by the UPLC-PDA were validated with respect to linearity, sensitivity, and precision. Linearity was evaluated by the correlation coefficient of the curves of studied compounds. A limit of detection (LOD) and quantification (LOQ) were determined by the injection of a series of dilutions of phenolic acids with LOD determined as the concentration that resulted in a peak area three times greater than the noise level and LOQ is a concentration that resulted in a peak area 10 times greater than the noise level. The precision of the method was estimated by running eight injections of six different concentrations, which were given on the same day, and the values of relative standard deviation were calculated to determine the intra-day precision.2 = 0.9954) in a concentration range 0.002 mg/mL to 0.012 mg/mL.TPC was determined using the Folin–Ciocalteu method  with min3 in methanol the GP extract (3 mg/mL) were mixed and left for 10 min. The absorbance was measured in pentaplicate at 415 nm using a blank sample of water and methanol without AlCl3. TFC was expressed as mg of quercetin (6.25–100 mg/mL) per 100 g of dry weight .TFC was determined as described by Meda et al. . Equal v3·6H2O solution. The working FRAP solution was freshly prepared by mixing 25 mL acetate buffer, 2.5 mL TPTZ solution, and 2.5 mL FeCl3·6H2O solution and then warmed at 37 °C before usage. The examination of the GP extract (50 μL) was allowed to react with the FRAP solution (1500 μL) at 37 °C for 30 min. in the dark condition. Then the absorbance was read at 593 nm in pentaplicate. The standard curve was linear in the range of 10 to 100 μg Vitamin C/mL with R2 = 0.9998. The results were expressed as the IC0.5 (µg/mL), which corresponded to the extract concentration required to produce 0.5 O.D. value.The FRAP assay was performed according to Tiveron el al.  with somInhibition of hyaluronidase (HA) by the GP extract was determined by a turbidimetric method described by Grabowska et al.  with minST—absorbance of the enzyme + substrate (HA) + substance sample. CT—absorbance of the enzyme + substrate (HA). HT—absorbance of the HA + substance sample.v/v fetal calf serum .The experiments were performed using human umbilical vein endothelial cells (HUVECs) of the EA.hy926 line  . Cells wTo ensure that the GP extract produced was not contaminated with endotoxin, GP extract was tested with the Limulus amebocyte lysate (LAL) assay.v/v) with 0.4% Trypan blue solution. After 10 min, the number of blue-stained non-viable cells was counted in a hemocytometer.The viability of cells treated with GP (0.125–1 mg/mL) was determined using the Trypan blue exclusion test. Cells were suspended in Hanks′ Balanced Salt Solution and mixed 1:1  in the presence or absence of IL-1β (1 ng/mL). After 24 h, the medium was collected and frozen at −80°C until it was assayed. The cells were lysed with 0.1 N NaOH and measured for protein concentration with the Bradford method using the Protein Assay Dye Reagent. IL-6 concentrations were measured with the DuoSet Immunoassay Development Kits  as per manufacturer′s instructions. The sensitivity of the assay was estimated to be 2.6 pg/mL.The assay was performed as described previously ,43. The The wound scratch assay is a well-established method to study wound healing in vitro and is thought to be particularly suited for measuring cell migration .2DCFDA) that is trapped inside the cells and activated by intracellular ROS. Following the exposure to test solutions, 2 × 104 cells were loaded with 10 µM H2DCFDA for 30 min and then treated with the lysis buffer. Fluorescence emitted by cell lysates was measured in a Wallac Victor spectrofluorometer  using wavelengths of 485 nm and 535 nm for excitation and emission, respectively. During the experiments of ROS detection in vitro, the internal positive control (100 µM H2O2) was also detected. In addition, the HUVEC line and EA.hy926 line were incubated ± GP extract (0.5–1.0 mg/mL) for 24 h in 6 well dishes. Trypsinized cells (2.5–3 × 105) were washed twice in cold PBS and dihydrorhodamine 123 (10 μg/mL) was added. After 30 min of incubation at 37 °C, the samples were measured by a BDFACS Aria III cytometer and results were analyzed by FACS Diva Software. Dihydrorhodamine 123 (DHR) infiltrates the cell membrane and, after oxidation by ROS, is converted to rhodamine 123, which emits a green fluorescent signal after excitation by a blue laser (λ = 488 nm). ROS levels were measured as the difference in median fluorescence intensity (MdFI). As a control, we used cells with correct concentrations of GP extract without DHR.The generation of ROS by endothelial cells treated with the GP extract (0.5 to 1.0 mg/mL) was assessed by labeling with 2′–7′-dichlorodihydrofluorescein diacetate . Results were expressed as means ± SD. A The anti-inflammatory activity of the rich polyphenol extract from GP herb is, at least in part, related to free radical scavenging activity, inhibition of IL-6 release, and an inhibitory effect on hyaluronidase activity. The results of this study demonstrate that such a plant extract could have a potential as a drug in the treatment of skin disorders.


Rumex japonicus by high-speed counter-current chromatography (HSCCC). The extract was separated with n-hexane–ethanol–water  as the two-phase solvent system and yielded 3.4 mg of emodin, 24.1 mg of chrysophanol, and 2.0 mg of physcion from 500 mg of sample with purities of 99.2 %, 98.8% and 98.2%, respectively. The HSCCC fractions were analyzed by high-performance liquid chromatography (HPLC) and the chemical structures of the three anthraquinones were confirmed by 1H-NMR and 13C-NMR analysis. This is the first time these anthraquinones have been obtained from R. japonicus by HSCCC.Three anthraquinones—emodin, chrysophanol, and physcion—were successfully purified from the dichloromethane extract of the Chinese medicinal herb  Rumex japonicus Houtt., a rich source of anthraquinones, has been traditionally used for the treatment of heat phlegm, jaundice, constipation, scabies, and uterine hemorrhage in East Asian countries such as China, Korea, and Japan +; 1H-NMR  δ: 11.96 , 11.86 , 7.80 , 7.71 , 7.55 , 7.38 , 7.22 , 2.44 ; 13C-NMR  δ: 191.4 (C-9), 181.3 (C-10), 161.4 (C-8), 161.1 (C-1), 149.0 (C-3), 137.2 (C-6), 133.2 (C-10a)，132.8 (C-4a), 124.2 (C-2), 123.9 (C-7), 120.4 (C-4), 119.2 (C-5), 115.7 (C-8a), 113.6 (C-9a)，21.6 (-CH3). Compared with the data given in reference +; 1H-NMR  δ: 12.31 , 12.11 , 7.63 , 7.37 , 7.08 , 6.69 , 3.94 , 2.46 ; 13C-NMR  δ: 190.7 (C-9), 181.9 (C-10), 166.5 (C-8), 165.1 (C-1), 162.4 (C-6), 148.4 (C-3), 135.2 (C-10a), 133.2 (C-4a), 124.4 (C-4), 121.2 (C-2), 113.7 (C-9a), 110.2 (C-8a), 108.2 (C-5), 106.8 (C-7), 56.1 (-OCH3), 22.2(-CH3). Compared with the data given in reference [Fraction III: EI-MS eference ,28, fracRumex japonicus by HSCCC for the first time. These compounds can be isolated on a sufficiently large scale with high purities which may then be used as reference substances for chromatography or for bioactivity studies. The method is a feasible, economical, and efficient technique for rapid preparative isolation of complicated natural products.Three anthraquinones –emodin, chrysophanol, and physcion – were separated from the crude extract of


Structural elucidation of new isolates was based on nuclear magnetic resonance  and other spectroscopic analyses. Bioactivity evaluation indicated that 1-6 possessed anti-inflammatory activities with dose-dependent relationships for lipopolysaccharide (LPS)-induced nitric oxide production. Furthermore, 1-4 also showed moderate antiproliferative effects against human laryngeal carcinoma (HEp-2) (IC50 = 14.81~20.06 μg/mL) and human colon adenocarcinoma (WiDr) (IC50 = 12.89~21.14 μg/mL).Six azaphilonoid derivatives, including two new blue fluorescent monapurfluores A ( A growing body of evidence suggests a direct link between inflammation and cancer. It has been found that various steps in tumorigenesis, such as cellular transformation, promotion, proliferation, and metastasis, can be influenced by chronic inflammation . Nitric Monascus species, has been used as a food additive to enhance color and flavor and as a remedy for digestive and vascular diseases in Chinese traditional medicine +, suggesting a molecular formula of C23H32O4, which contains eight required degrees of unsaturation. Compound -1) indicated the existence of carbonyl groups and olefinic groups in 1, and the 1H-NMR spectrum of 1 exhibited five olefinic , one methoxyl (δH 3.09), and three methyl  protons. The 13C-NMR spectrum of 1  and a conjugated carbonyl carbon (δC 195.7). Three singlet olefinic protons  in the 1H-NMR spectrum, along with HMBC long-range correlations   and the COSY cross peaks of H-9/H-10 and H-10/H-11. Based on the HMBC correlations between H-9/C-3 and between CH3-12/C-6, C-7 and C-8, the trans-propenyl and CH3-12 groups were assigned at C-3 and C-7, respectively. The above corroboration, together with the number of degrees of unsaturation, allowed us to assign 1 to have a pendant ring. The nature of the ring was determined by the COSY correlation between H-8/H-13, and the HMBC correlation between H-13/C-7 and C-14, indicating that 1 possessed a five-membered furan ring fused to C-7 and C-8 of the azaphilonoid structure. Moreover, cross peaks from H-15 to H-21 were observed in the COSY spectrum of 1, proving the presence of a heptyl side chain. In the HMBC spectrum of 1, correlations between H-15/C-14 and OCH3-22/C-14 were found; thus, both the alkyl side chain and OCH3-22 were located at C-14 of the five-membered furan ring.The IR absorbance bands  , suggest3) experiment. After the irradiation of H-8, the protons of CH3-12 were enhanced, whereas no enhancement was observed for OCH3-22, suggesting that the relative configurations between H-8/OCH3-22 and between H-8/CH3-12 were trans- and cis-, respectively. Based on this evidence, monapurfluore A (1) was determined to be 8-heptyl-9,9a-dihydro-8β-methoxy-6a-methyl-3-[(E)-prop-1-enyl]-6aH-furoisochromen-6(8H)-one, a new natural azaphilone derivative. The relative configuration of the tetrahydrofuran moiety was established based on a 1D NOE . The fluorescence, IR, UV, 1H-, and 13C-NMR spectra showed that 2 and 1 were azaphilone stereoisomers. The chemical shifts of C-8, C-13 and C-14 in 2 were shifted upfield by ∆δc 1.2, 1.5 and 0.5 ppm, respectively, compared with 1. After the irradiation of H-8 in the 1D NOE experiment, enhanced signals for CH3-12 and OCH3-22 were seen, suggesting that these protons were of the α-configuration. Thus, the structure of 2 (monapurfluore B) was established as 8-heptyl-9,9a-dihydro-8α-methoxy-6a-methyl-3-[(E)-prop-1-enyl]-6aH-furo iso-chromen-6(8H)-one. The HREI-MS of 3-6 were identified as the known compounds monascopyridine C, monascopyridine D, monasfluore A and monasfluore B, respectively, by comparison with authentic samples and literature data 25D: +18.84 ; UV (MeOH) λmax: (log ε) 368 (4.1), 284 (3.9); IR: vmax (KBr) 2,954, 2,926, 2,855, 1,706, 1,623, 1,572, 1,548, 1,453, 1,370, 1,299, 1,255, 1,176, 1,097, 9,18 cm-1; EIMS m/z 372 [M]+; HREI-MS m/z 372.2292 [M]+ ; 1H- and 13C-NMR data, see Monapurfluore B (2). Slightly yellowish oil; [α]25D: -71.43 ; UV (MeOH) λmax: (log ε) 367 (4.2), 284 (3.9); IR: vmax (KBr) 2,918, 2,851, 1,714, 1,627, 1,572, 1,548, 1,441, 1,374, 1,310, 1,239, 1,176, 1,093, 914 cm-1; EI-MS m/z 372 [M]+; HREI-MS m/z 372.2308 [M]+ ; 1H- and 13C-NMR data, see 2.HEp-2 , WiDr (human colon adenocarcinoma), and RAW 264.7 (murine macrophage) were obtained from Food Industry Research and Development Institute . All cell lines were maintained in MEM containing 5% foetal bovine serum and were kept in a 37 ºC incubator with 5% CO3 per well). After 4 h, test agents (20 μL), dissolved in PBS solution, were added to reach final concentrations of 6.25, 12.5, 25, 50 and 100 μg/mL . Twenty μL of MTT solution (2 mg/mL) was added to each well and incubated for 4 h in a 37 ºC incubator with 5% CO2. After three days of incubation, the cellular conversion of a tetrazolium salt into a formazan product was achieved. The supernatant was removed and DMSO (200 μL) was added to dissolve the formazan, which was finally detected by spectrophotometry at a wavelength of 570 nm, and the relative estimate of cell proliferation was calculated.HEp-2 and WiDr were seeded in MEM (180 μL) in 96-well plates (3 × 104 per well) were seeded and maintained with DMEM (90 μL) in 96-well plates. After incubating for 12 h, wells were treated with LPS (1 μg/mL) and test agents (10 μg/mL) dissolved in DMEM. The nitrite concentrations of the supernatants were determined using a Griess reagent kit  after 24 h. The cell proliferation was evaluated by a cell growth inhibitory assay.RAW 264.7 cells (5 × 10t-test. Differences were considered significant at p < 0.05.The data on cancer cell growth inhibition and nitrite production were presented as mean ± standard deviation for three independently performed experiments (n = 3). Significant difference was analyzed by Student‘s M. purureus NTU 568 fermented red mold rice. The structures of new azaphilone compounds 1 and 2 were elucidated by spectral methods. Bioassays revealed that the isolates 1-4 showed moderate anti-proliferation effects against human cancer cell lines HEp-2 and WiDr and anti-inflammatory activity by the inhibition of LPS-induced NO production. The current results, together with the exhibition of anti-NO activity by the crude M. purpureus NTU 568 fermented RMR extracts [In this study, two new and four known azaphilone derivatives were isolated from extracts , suggest


Their structures were elucidated on the basis of one- and two-dimensional spectroscopic analysis. Isolation of compounds 4, 6–13 from this plant is reported here for the first time.Two new limonoids, toonins A ( Toona sinensis  is a woody plant native to eastern and southeastern Asia that has more than 2,000 years of cultivation history in China + , 567.23561 [M+K]+  in positive mode. Its NMR spectra generally resembled compound 1 and showed strong similarity to the evodulane type nomilins carapolide H and carapolide I . In addition to the characteristic α-orientation furan signals, two oxymethine resonances were clearly discernable at δ: 4.94  and 4.70 . The HMBC cross-signals can deduce the OAc groups were attached at C-1 and C-7. The correlations of the carbonyl C-3 resonance at δ 170.00 to the proton signals of H-1 and CH3-28 suggested the existence of a lactone moiety at A-ring of 2. Compared with carapolide H , however, it is a CH2 signal of C-16  instead of a ketone carbon. Through further literature research, the NMR data of the D-ring was found to be very similar to the D-ring of surenin . And H-17/C-20 (δC-20: 123.03), H-17/C-21(δC-21: 138.84) revealed the furan ring  was attached at C-17. Therefore, structure 2 was attributed to 14,15-epoxycarapolide I and named toonin B.Compound I 2a,2b,  21]. Th. Th2 wasde H 2a, , the D-renin 2c,  22] who who2 was3 was obtained as an amorphous white powder (CH3OH). The molecular formula C19H18O7 was established by HR-ESIMS+: m/z 359.1125 [M+H]+ , 381.0945 [M+Na]+, 739.1997 [2M+Na]+ in positive mode. Its IR spectrum showed characteristic absorptions for hydroxyl group (3445 cm−1), carbonyl group (1712 cm−1), and phenyl group . The NMR data was very similar to that of the known compound cedralin A  of 3, suggested methyl esterification of the carboxylic acid group at C-1′ of cedralin A  benzofuran-1′-carboxylic acid methyl ester) and named toonin C -pyran-2-one, had previously been reported : 0, 0; 9.5, 100; 12.2 100; 12.3, 0; 15.0, 0. Detection was carried out at 200 nm, with peak scanning between 195-600.2 nm. All of the analyses were carried out using a Turbo Ionspray source in the negative mode with the following settings: capillary exit, 100 V; nebulizer, curtain, collision, and drying gas (N2). Full scan acquisition was performed by scanning from m/z 100 to 1,750 U in profile at a cycle time of 2 s with a step size of 0.1 U and a pause between each scan of 2 ms. MS/MS product ions were produced by collision-activated dissociation (CAD) of the selected precursor ions in the collision cell of the triple quadrupole mass spectrometer and analyzed using the second analyzer of the instrument. In all of the experiments, quadrupoles (Q1 and Q3) were operated at unit resolution. Different MS/MS experiments, neutral loss scan, product ion scan, and precursor ion scan of selected molecules were carried out to confirm the structure of the compounds previously identified by full scan mode. Analytical HPLC was performed on a Purospher STAR RP-18 column, 3 μm, 55 mm length × 4 mm ID, Gradient elution was performed with water/0.01% TFA (solvent A) and acetonitrile/0.01% TFA (solvent B) at a constant flow rate of 1.5 mL/min. An increasing linear gradient (v/v) of solvent B was used : 0, 10; 7.5, 100; 10, 100; 10.5, 10; 13.0, 10. Detection was carried out at 200 and 220 nm, with peak scanning between 195-600.2 nm. Millennium software was used for data acquisition and processing. An Agilent ChemicalStation was used off-line for library building and RT-UV library search. Preparative HPLC (Varian) for fractions 18 and 6 was performed on a Purospher RP-18 column, 12 μm, 250 mm length × 50 mm ID, Gradient elution was performed with water (solvent A) and acetonitrile (solvent B) at a constant flow rate of 40 mL/min. An increasing linear gradient (v/v) of solvent B was used : 0, 20; 65, 85; 80, 100; 95, 100. Detection was carried out at 210 nm. Preparative HPLC (Varian) for the 16th fraction: Purospher RP-18 column, 12 μm, 250 mm length × 25 mm ID, Gradient elution was performed with water (solvent A) and acetonitrile (solvent B) at a constant flow rate of 10 mL/min. An increasing linear gradient (v/v) of solvent B was used : 0, 15; 60, 45; 90, 100; 111, 100. Detection was carried out at 270 nm. Preparative HPLC (Varian) for the 30–39th fractions: Purospher RP-18 column, 12 μm, 250 mm length × 25 mm ID, Gradient elution was performed with water (solvent A) and acetonitrile (solvent B) at a constant flow rate of 10 mL/min. An increasing linear gradient (v/v) of solvent B was used : 0, 30; 60, 62; 90, 100; 111, 100. Detection was carried out at 250 nm.All solvents used in the extraction and isolation processes were redistilled prior to use. IR spectra (KBr) were obtained on a Perkin Elmer Spectrum 2000 instrument. All NMR spectra were recorded on a Bruker DRX-500 instrument in CDClPlant Material. The roots of T. sinensis were initially collected in Tonglin County of Anhui Province in October, 2011, and identified by one of the authors (G.-H.B.). A voucher specimen (No. AHAU2011102-1) was deposited in the herbarium of Anhui Agricultural University.3 (4 × 4 L), EtOAc (3 × 4 L), and n-BuOH (4 × 4 L). The CHCl3 extract (355 g) was fractionated on a silica gel (100–200 mesh) column (11 × 100 cm) and gradually eluted by hexane-dichloromethane-methanol to yield 18 fractions. All of the 18 fractions were examined by TLC and LC-PAD-MS and some interesting constituents were found in fractions 6 (2.0 g), 14 (7.9 g), 15 (7.1 g), 16 (8.3 g) and fraction 18 (5 g). These five fractions were further fractionated by repeated preparative HPLC. Further preparative HPLC of Fraction 15 gave three limonoids: the new compounds toonin A , toonin B , and the known one proceranone . The new compound toonin C  and the known compound lyoniresinol  were obtained from Fraction 6. Further HPLC purification of fraction 18 yielded 4-methoxy-6--pyran-2-one  and syringic acid . Fraction 14 was subjected to repeated preparative HPLC yielding bourjotinolone A  and matairesinol . 4-Hydroxy-3-methoxybenzeneethanol , isoscopoletin , aloeemodin (12), and β-sitosterol (13) were obtained by repeated HPLC purification of Fraction 16. The air-dried and powdered roots (15 kg) were extracted with 95% EtOH (3 × 30 L) at room temperature. After removal of the solvent by evaporation under vacuum, the residue was diluted with 70% aqueous methanol (4 L) and then extracted successively with CHClToonin A (1). White amorphous powder, [α]D25 −25.3 . IR (KBr) νmax (cm−1): 2987, 1741, 1720, 1508, 1444, 1370, 1221, 1123, 1024, 876, 774. ESIMS−: m/z 615.3 [M−H]− (6), 677.2 [M+HCO3]− (38), 156.9 (100) in negative mode, ESIMS+: m/z 639.2 [M+Na]+ (100) in positive mode; HRESIMS+: m/z 639.2411 [M+Na]+ , 634.2860 [M+NH4]+ in positive mode. 1H and 13C-NMR data (CDCl3), see Toonin B (2). White powder, [α]D25 −43.7 . IR (KBr) νmax (cm−1): 2984, 2948, 1802, 1731, 1600, 1504, 1459, 1435, 1374, 1235, 1124, 1027, 874, 789 cm−1; EIMS: m/z 528 [M]+ (16), 468 (5), 410 (7), 350 (8), 208 (24), 107 (100), 79 (13), 43 (51); ESIMS−: m/z 527.2 [M−H]− (94) in negative mode; ESIMS+: m/z 529.2 [M+H]+ (30), 546.2 [M+NH4]+ (100), 1074.1 [2M+NH4]+ (15) in positive mode; HRESIMS+: m/z 551.26166 [M+Na]+ , 567.23561 [M+K]+ in positive mode. 1H-NMR (CDCl3), see 13C-NMR (CDCl3) δ: 70.48 (C-1), 34.08 (C-2), 170.00 (C-3), 85.24 (C-4), 44.08 (C-5), 26.47 (C-6), 73.22 (C-7), 41.73 (C-8), 37.05 (C-9), 44.25 (C-10), 15.80 (C-11), 29.42 (C-12), 41.46 (C-13), 72.48 (C-14), 56.73 (C-15), 31.54 (C-16), 39.16 (C-17), 21.02 (C-18), 15.82 (C-19), 123.03 (C-20), 138.84(C-21), 110.92 (C-22), 142.71 (C-23), 34.03 (C-28), 23.01 (C-29), 18.85 (C-30), 169.96 (OCOCH3-1), 21.07 (OCOCH3-1), 169.54 (OCOCH3-7), 20.60 (OCOCH3-7).Toonin C (3). white powder (CH3OH); [α]D25 −22.6 ; IR (KBr) νmax (cm−1): 3445, 2948, 1712, 1613, 1599, 1492, 1435, 1331, 1250, 1108, 1038, 936, 769 cm−1; ESIMS+: m/z 359.2 [M+H]+ (52), 381.1 [M+Na]+ (100), 739.0 [2M+Na]+ (26); in positive mode; HRESIMS+: m/z 359.1125 [M+H]+ , 381.0945 [M+Na]+, 739.1997 [2M+Na]+ in positive mode. 1H-NMR (DMSO-d6) δ: 6.88 , 6.91 , 6.84 , 5.60 , 3.49 , 3.69 , 5.11 , 7.44 , 7.55 , 6.01 , 3.81 , 3.83 . 13C-NMR (DMSO-d6) δ: 134.94 (C-1), 106.17 (C-2), 146.99 (C-3), 148.36 (C-4), 108.36 (C-5), 119.30 (C-6), 86.67 (C-7), 52.45 (C-8), 62.58 (C-9), 121.94 (C-1′), 113.35 (C-2′), 142.66 (C-3′), 151.72 (C-4′), 129.44 (C-5′), 118.93 (C-6′), 100.98 (OCH2O), 166.30 (OCOCH3-1′), 51.92 (OC3OCH-1′), 56.08 (OCH3-3′). 4-Methoxy-6--pyran-2-one (4). White powder (CH3OH); IR (KBr) νmax (cm−1): 3335, 2942, 1680, 1610 1558, 1514, 1454, 1406, 1336, 1269, 1222, 1161, 840. ESIMS−: m/z 247.1 [M−H]− (100), 495.1 1 [2M−H]− (5) in negative mode; ESIMS+: m/z 249.1 [M+H]+ (100), 271.1 [M+Na]+ (12), 497.1 [2M+H]+ (3) in positive mode; HRESIMS−: m/z 247.06118 [M−H]−  in negative mode. 1H-NMR (DMSO-d6) δ: 5.60 , 6.12 , 6.22 , 6.15 , 3.83 , 9.55, 9.66 , 2.08 . 13C-NMR (DMSO-d6) δ: 164.90 (C-2), 88.62 (C-3), 171.74 (C-4), 105.34 (C-5), 159.30 (C-6), 111.57 (C-1′), 157.68 (C-2′), 99.96 (C-3′), 159.91 (C-4′), 109.24 (C-5′), 139.12 (C-6′), 56.28 (OCH3-4), 19.75 (CH3-6′). 1H-1H COSY, HSQC, HMBC, and ROESY spectra.The assignments were confirmed by a combination of T. sinensis yielded three new and ten known compounds, among which eight known phenols and four Meliaceae limonoids were found. The considerable number of phenols in the plants may account for its strong free radical scavenging activity. Isolation of compounds 4, 6–13 from this plant is reported here for the first time.The chemical investigation on the free radical scavenging active fraction of the roots of


The new compounds were determined on the basis of spectroscopic and chemical methods to be 3β-acetoxy-4β-hydroxy-1α,7α, 10β,11αH-pseudoguaia-12,8β-olide (1), 1β,7β,9β,10β,13αH-guaia-4(5)-en-12,6β-olide 9-O-β-d-glucoside (2) and 4β-hydroxy-1α,5α,7α,9αH-guaia-10(14),11(13)-dien-12-acid 9-O-β-d-glucoside (3). The isolated compounds were evaluated for cytotoxicity against human promyelocytic leukemia HL-60 cell lines in vitro, but were all inactive.A new pseudoguaianolide  Ambrosia artemisiifolia L. (Asteraceae), an invasive alien plant species in China, is nowadays widespread in an area ranging from Guangdong Province in the south to Heilongjiang Province in the north  , a quaternary carbon, and two quaternary carbonyl groups  to C-16; H3-14 (δH 0.90) to C-1, C-9 and C-10; H3-13 (δH 1.02) to C-7, C-11 and C-12; H3-15 (δH 1.18) to C-1, C-4, C-5 and C-6)  and three fragments as 14CH3-10CH(1CH)-9CH-, 13CH3-11CH (7CH)-12COOH, and 15CH3-5C(4CH/1CH)-6CH2-  and C-8 (δH 4.58 and δC 79.7)  with C-16 (δC 169.8) observed in the HMBC spectrum , and corroborated by an NOE between H-10 and H3-15. Finally, compound 1 was established as 3β-acetoxy-4β-hydroxy-1α,7α,10β,11αH-pseudoguaia-12,8β-olide.Compound , H-13)] . The 13Cl groups . The strand C-6) ] establiH)-6CH2- . Those felations . Conside3H, s, H-, 2.04 ; δC 102.5, 75.2, 77.9, 71.8, 78.2, 62.8], which was confirmed by acid hydrolysis and then co-chromatography with an authentic sample. Besides, the remaining 15 carbon signals, which belong to the aglycone, were attributable to three methyls, three methylenes, six methines (including two oxy-methines), two olefinic quaternary carbons, and a quaternary carbonyl group -9CH(OR)-8CH2-7CH(6CHOR)-11CH-13CH3] which was corroborated by HMBC correlations (C 82.6 (C-9) showed a long-range correlation with the anomeric proton of glucose at δH 4.38 , indicating that the hydroxyl group at C-9 is glucosylated. The low-field chemical shift of δC 181.2 (C-12) and δC 83.3 (C-6), as well as the molecular formula C21H32O8 revealed that a lactone ring was formed between C-12 and C-6 .,18.2 has and C-6 . Therefoone ring . Moreovel at C-7 , the ste3 was obtained as a colorless gum. The molecular formula was shown to be C21H32O9 by its HRESIMS and 13C-NMR data. The structure of 3 contained a β-d-glucopyranosyl, which was confirmed by methods previously described in compound 2. Moreover, two pair of exomethylene singlets at δH 5.55 /6.42  and 5.27 /6.18 , along with a methyl singlets δH 1.32  was clearly observed in 1H-NMR spectrum. The 13C-NMR and DEPT spectra . The HMBC correlations from the H-atoms of the exomethylenes and methyl groups to corresponding C-atoms revealed three C4 units as 13CH2=11C (7CH)-12COOH, 14CH2=10C(1CH)-9CH-, and 15CH3-4C(3CH2)-5CH-, respectively. Those C4 units in combination with a C8 unit -3CH2-2CH2-1CH-5CH-6CH2-7CH-8CH2-9CHOR deduced from 1H-1H COSY correlations  and a methine carbon signal at δC 79.5 (C-9) in the HMBC spectrum (3 was established as 4-hydroxyguaia-10(14),11(13)-dien-12-acid 9-O-β-d-glucoside. The key NOESY correlations of H-1 with H-5 and H-9, and H-5 with H-7, H-9 and H3-15 showed that all of those protons  were cofacial. Meanwhile, the coupling constants of H-9  revealed its α-oriented position. Thus compound 3 was determined to be 4β-hydroxy-1α,5α,7α,9αH-guaia-10(14),11(13)-dien-12-acid 9-O-β-d-glucoside.Compound  spectra  showed aelations , establispectrum , implyin1–5 were tested against human promyelocytic leukemia HL-60 cell by the MTT method in vitro. The results revealed that all compounds were inactive (LC50 > 100 μM).The cytotoxicity of compounds Optical rotations were measured on WZZ2B automatic polarimeter ; melting points were obtained on a SGW X-4 micromelting point apparatus . The 1D- and 2D-NMR spectra were measured with a Bruker DRX-400 instrument  with TMS as internal standard. ESIMS data were recorded on an API QSTAR mass spectrometer . Column chromatography was performed on silica gel 60 , Sephadex LH-20  and Develosil ODS . Preparative HPLC was performed on a Waters 1525 Binary HPLC pump and a Waters 2414 refractive index detector  using a YMC-Pack ODS-A column .A. artemisiifolia were collected from Miluo, Hunan Province, P. R. China, in August 2012, and identified by Prof. Zhongshi Zhou, Institute of Plant Protection, Chinese Academy of Agricultural Sciences. The voucher specimen (No. 20120821) was deposited in Hunan Agricultural University.The aerial parts of A. artemisiifolia (10.0 kg) were powdered and extracted three times with MeOH (95% v/v) at room temperature, then concentrated under reduced pressure to obtain a crude residue (0.5 kg). The residue was further suspended in H2O (2 L) and extracted with petroleum ether (PE) and EtOAc successively, to yield a PE-soluble fraction (90.0 g), an EtOAc-soluble fraction (90.0 g). The EtOAc-soluble fraction was subjected to silica gel column chromatography (CC) (100–200 mesh) with elution of CHCl3–MeOH  to give six fractions (Fr. C1–C6). The Fr. C2 was further separated by ODS-C18 CC  to produce seven sub-fractions (C2-1–C2-7), then sub-fraction C2-3 was further purified by Sephadex LH-20 column (MeOH) and normal silica gel CC  to yield colourless crystal 4 (100.0 mg) and 5 (30.0 mg). Similarly, Fractions C3 (6.0 g) and C5 (8.3 g) were fractionated by an ODS-C18 column with elution of MeOH–H2O , respectively. Compound 1 (60.0 mg) was obtained from sub-fraction C3-5 by Sephadex LH-20 column (MeOH) and semi-preparative HPLC chromatography . Compounds 2 (8.0 mg) and 3 (6.0 mg) were obtained from sub-fraction C5-2 which purified successively on Sephadex LH-20 column (MeOH) and semi-preparative HPLC chromatography . Dried aerial parts of 1: white powder, mp 290–293 °C, c 0.9, MeOH); IR (KBr) υmax 3342, 2951, 1769, 1740, 1727, 1704, 1660, 1176, 1035 cm−1; 1H-NMR  and 13C-NMR  spectroscopic data, see m/z: 311 [M+H]+, 333 [M+Na]+; negative ESIMS m/z: 655 [2M+Cl]−; HRESIMS m/z: 311.1852 [M+H]+ .Compound 2: white powder, mp 305–307 °C, c 0.5, MeOH); IR (KBr) υmax 3392, 2921, 2850, 1754, 1688, 1458, 1384, 1229, 1176 cm−1; 1H-NMR  and 13C-NMR  spectroscopic data, see m/z: 413 [M+H]+, 435 [M+Na]+; negative ESIMS m/z: 411 [M−H]−, 859 [2M+Cl]−; HRESIMS m/z: 435.1971 [M+Na]+ .Compound 3: yellowish syrup, c 1.1, MeOH); IR (KBr) υmax 3400, 2923, 2870, 1702, 1458, 1380, 1221 cm−1; 1H-NMR  and 13C-NMR  spectroscopic data, see m/z: 451 [M+Na]+, 879 [2M+Na]+; negative ESIMS m/z: 427 [M−H]−, 463 [M+Cl]−; HRESIMS m/z: 451.1935 [M+Na] + .Compound 2 (2.0 mg) in 1 N HCl  was heated under reflux for 8 h. After removal of the solvent, the residue was partitioned between Et2O and H2O. The water layer was neutralized with 5% NaOH and desalted  to afford the sugar residue (1.0 mg). The sugar residue was derivatized with Sigma Sil-A for 35 min at 70 °C and analyzed by GC-MS . In the acid hydrolysate of 2, d-Glucose was confirmed by comparison of the retention times of the aforementioned derivative with the authentic d-glucose derivative prepared in a similar way, both of which showed identical retention time of 12.17 min. By the same method, the sugar moiety of 3 was identified as d-glucose.Compounds in vitro at the Second Xiangya Hospital of Central South University, and was determined by the MTT assay [The cytotoxic activity of each compound against human promyelocytic leukemia HL-60 cell was examined TT assay ,21.A. artemisiifolia afforded a new pseudoguaianolide (1), two new guaiane-type sesquiterpene glucosides (2 and 3), as well as two known sesquiterpene dilactones psilostachyin B (4) and psilostachyin (5). Although all the isolated compounds were inactive against human promyelocytic leukemia HL-60 cell, the discovery of these new compounds further expands our knowledge of the structural diversity of the sesquiterpenoids produced by the plant A. artemisiifolia.The phytochemical investigation of the aerial parts of