Lilium pumilum (Liliaceae) led to the isolation of four novel steroidal glycosides (1–4) with a 2,3,4-trisubstituted β-d-glucopyranosyl unit. In 1 and 3, the α-l-arabinopyranosyl moiety is linked to C-3 of the inner trisubstituted β-d-glucopyranosyl group and is present as an usual 4C1 conformation. In contrast, in 2 and 4, the α-l-arabinopyranosyl moiety, which is attached to C-4 of the inner trisubstituted β-d-glucopyranosyl group, is present as a 1C4 conformation. The structures of the new steroidal glycosides were determined based on the results of spectroscopic analyses, including two-dimensional (2D) NMR data and hydrolysis.Examination of the bulbs of  Lilium pumilum D.C. (Liliaceae) is described in the Japanese Pharmacopoeia (16th edition) as a plant from which the crude drug Lilium Bulb is derived. Lilium Bulb has long been used as an antitussive and anti-inflammatory agent in traditional Chinese medicine -O-[α-l-rhamnopyranosyl-(1→2)]-β-d-glucopyranosyl was linked to C-3 of the aglycone. A β-d-glucopyranosyl group (Glc (III)) linkage to the C-26 hydroxy group of the aglycone was confirmed by an HMBC correlation between H-1′′′ of Glc (III) at δH 4.82  and C-26 of the aglycone at δC 75.2. The NOE correlations between the signals of the H-20 proton at δH 2.21 and the H2-23 protons at δH 2.03 (2H) confirmed the C-22α configuration -22α-hydroxyfurost-5-en-3β-yl O-α-l-arabinopyranosyl-(1→3)-O-[β-d-glucopyranosyl-(1→4)]-O-[α-l-rhamnopyranosyl-(1→2)]-β-d-glucopyranoside.Compound 2 had the same molecular formula as 1 of C56H92O27, based on the HR-ESI-TOF-MS and 13C-NMR  data. Two singlet signals for tertiary methyl groups at δH 1.04 and 0.89, two doublet signals for secondary methyl groups at δH 1.33  and 1.00 , and five signals for anomeric protons at δH 5.94 (br s), 5.74 , 5.49 , 4.85 , and 4.81  were observed in the 1H-NMR spectrum of 2. The 13C-NMR spectrum contained a signal for an acetal carbon at δC 110.6, and signals for four steroid methyl groups at δC 19.3, 17.4, 16.4, and 16.4. These spectroscopic data for 2 were analogous to those of 1, and suggested that 2 shared the same fundamental furostanol skeleton as 1. Acid hydrolysis of 2 gave 1a, l-arabinose, d-glucose, and l-rhamnose. Although the 1H- and 13C-NMR spectra of 2 indicated that the sugar moieties of 2 also consisted of two terminal β-d-glucopyranosyl units (Glc (II) and Glc (III)), a terminal α-l-arabinopyranosyl unit (Ara), a terminal α-l-rhamnopyranosyl unit (Rha), and a 2,3,4-trisubstituted inner β-d-glucopyranosyl moiety (Glc (I)), it was assumed that the linkage positions of the terminal sugar units to the inner Glc (I) moiety and the conformation of the Ara unit were different from those of 1. The 13C-NMR chemical shifts of C-1 to C-5 for Ara were assigned as δC 102.3, 71.0, 73.3, 65.6, and 62.1, respectively, by combining the 1H-1H COSY, HMQC, and HSQC-TOCSY spectra. Furthermore, the anomeric proton of Ara at δH 5.74  exhibited three-bond coupled strong HMBC correlations with the C-3 and C-5 carbons. These spectral features were consistent with the 1C4 conformation of Ara with an α-orientation of the anomeric center -22α-hydroxyfurost-5-en-3β-yl O-α-l-arabinopyranosyl-(1→4)-O-[β-d-glucopyranosyl-(1→3)]-O-[α-l-rhamnopyranosyl-(1→2)]-β-d-glucopyranoside.Compound 3 was analyzed as C56H90O26 based on the HR-ESI-TOF-MS (m/z 1201.5615 [M + Na]+) and 13C-NMR  data. This molecular formula was smaller than that of 1 by 18.0059 (H2O). The 1H- and 13C-NMR spectral features of 3 were similar to those of 1; however, the Me-21 doublet signal at δH 1.34  in the 1H-NMR spectrum of 1 was replaced by a methyl singlet signal at δH 1.64 in that of 3, and an olefinic functionality (δC 152.3 and 103.6) was observed in addition to the 5(6)-ene group in the 13C-NMR spectrum of 3. These spectroscopic data and the HMBC correlations from H-17 at δH 2.45, H2-23 at δH 2.22, and Me-21 at δH 1.64, to C-22 at δC 152.3 and C-20 at δC 103.6 indicated that 3 was the corresponding Δ20(22)-pseudo-furostanol glycoside of 1. This was supported by all other spectroscopic data and the results of acid hydrolysis. The structure of 3 was assigned as (25R)-26-[(β-d-glucopyranosyl)oxy]-furosta-5,20(22)-dien-3β-yl O-α-l-arabinopyranosyl-(1→3)-O-[β-d-glucopyranosyl-(1→4)]-O-[α-l-rhamnopyranosyl-(1→2)]-β-d-glucopyranoside.Compound 4 was shown to have a molecular formula of C56H90O26 by the HR-ESI-TOF-MS  and 13C-NMR  data, and its aglycone was identified as the same Δ20(22)-pseudo-furostanol derivative as 3, based on its spectroscopic data. Further spectroscopic analysis and acid hydrolysis of 4 indicated that the sugar moieties attached to C-3 and C-26 of the aglycone were in agreement with those of 2, in which the terminal α-l-arabinopyranosyl group was present as an unusual 1C4 conformation. It is notable that the 1C4 α-l-arabinopyranosyl moiety in 4 was converted to the 4C1 moiety on peracetylation (4a). Therefore, 4 was the corresponding Δ20(22)-pseudo-furostanol glycoside of 2, (25R)-26-[(β-d-glucopyranosyl)oxy]-furosta-5,20 (22)-dien-3β-yl O-α-l-arabinopyranosyl-(1→4)-O-[β-d-glucopyranosyl-(1→3)]-O-[α-l-rhamnopyranosyl-(1→2)]-β-d-glucopyranoside.Compound 1–4 were evaluated for their cytotoxic activity against HL-60 cells. They were not cytotoxic to HL-60 cells at sample concentrations up to 15 μM.The isolated compounds 1H-NMR spectra were obtained at 500 MHz by using standard Bruker pulse programs at 300 K . Chemical shifts are given as δ values relative to tetramethylsilane  used as an internal standard. HR-ESI-TOF-MS data were recorded on an LCT mass spectrometer . Diaion HP-20 , BW-300 silica gel , and COSMOSIL 75C18-OPN ODS silica gel  were used for CC. TLC was carried out on precoated silica gel 60 F254  and RP18 F254S plates , and the compounds were visualized by spraying the plates with 10% H2SO4 (aq.) and then heating. HPLC was performed with a system consisting of a CCPM pump , an RI-8021 detector (Tosoh), and a Rheodyne injection port . A TSK gel ODS-100Z column  was used for preparative HPLC. NMR spectra are available in the Optical rotations were measured by using an automatic digital polarimeter . IR spectra were recorded on a spectrophotometer . L. pumilum bulbs were obtained from the Sakata Seed Corporation  in 2008 and were identified by Dr. Yutaka Sashida, professor emeritus of Tokyo University of Pharmacy and Life Sciences. We have retained a voucher specimen in our laboratory (KS-2008-002).The L. pumilum (1.4 kg fr. weight) were extracted with MeOH under reflux for 6 h. After removing the solvent, the MeOH extract (65 g) was passed through a Diaion HP-20 column  and successively eluted with MeOH–H2O , MeOH, EtOH, and EtOAc (6 L of each eluent). CC of the MeOH-eluted fraction (1.8 g) on silica gel , eluted with a stepwise gradient mixture of CHCl3–MeOH–H2O (7:4:1) and finally with MeOH, provided 6 fractions (M-1 to M-6). Fraction M-4 (0.5 g) was chromatographed on ODS silica gel  eluted with MeOH–H2O (2:1) and finally with MeOH, provided 9 fractions (M-4-1–M-4-9). Fraction M-4-5 (148 mg) was separated by HPLC (1.0 cm i.d. × 25 cm) using MeCN–H2O (1:3) to afford 3 (14.3 mg) and 4 (15.0 mg). Fraction M-4-7 (94 mg) was separated by HPLC (1.0 cm i.d. × 25 cm) using MeCN–H2O (1:2) to afford 1 (20.6 mg) and 2 (20.6 mg).The bulbs of 1 (14.5 mg), 2 (14.5 mg), 3 (5.0 mg), or 4 (5.0 mg) in 1 M HCl in dioxane–H2O  were heated at 90 °C for 2 h under an Ar atmosphere, respectively. The reaction mixture was neturalized by passing it through an Amberlite column . The mixture was then eluted through a Diaion HP-20 column  with MeOH–H2O (3:7) and EtOH–Me2CO (1:1). The EtOH–Me2CO (1:1) fraction was purified by CC with CHCl3–MeOH–H2O (7:4:1) to give diosgenin , respectively. The MeOH–H2O (3:7) fraction was analyzed by HPLC under the following conditions: Capcell Pak NH2 UG80 column ; mobile phase of MeCN–H2O (85:15); detection by refractive index and optical rotation; and a flow rate of 1.0 mL/min. l-Arabinose, d-glucose, and l-rhamnose were identified, respectively, by comparing their retention times (tR) and optical rotation with those of authentic samples : l-arabinose , d-glucose , and l-rhamnose .A solution of 4 (8.0 mg) and Ac2O (1.0 mL) in pyridine (7.0 mL) was stirred at room temperature for 24 h. The reaction mixture was purified by CC with CHCl3–EtOAc (1:1) to give the corresponding peracetylation .A mixture of d-Glucopyranosyl)oxy]-22α-hydroxyfurost-5-en-3β-yl O-α-L-arabinopyranosyl-(1→3)-O-[β-d-glucopyranosyl-(1→4)]-O-[α-l-rhamnopyranosyl-(1→2)]-β-d-glucopyranoside(25R)-26--22α-hydroxyfurost-5-en-3β-yl O-α-l-arabinopyranosyl-(1→4)-O-[β-d-glucopyranosyl-(1→3)]-O-[α-l-rhamnopyranosyl-(1→2)]-β-d-glucopyranoside(25R)-26--furosta-5,20(22)-dien-3β-yl O-α-l-arabinopyranosyl-(1→3)-O-[β-d-glucopyranosyl-(1→4)]-O-[α-l-rhamnopyranosyl-(1→2)]-β-d-glucopyranoside(25R)-26--furosta-5,20(22)-dien-3β-yl O-α-l-arabinopyranosyl-(1→4)-O-[β-d-glucopyranosyl-(1→3)]-O-[α-l-rhamnopyranosyl-(1→2)]-β-d-glucopyranoside(25R)-26-[(β- (4); An amorphous solid; c 0.03, MeOH); HR-ESI-TOF-MS ; IR νmax (film) cm−1: 3357 (OH), 2925 (CH); 1H-NMR : for the aglycone moiety, 5.33 , 4.75 , 3.94 , 3.83 , 3.47 , 1.63 , 1.04 , 1.01 , 0.72 . For the sugar moieties, 13C-NMR : Compound4a; An amorphous solid; c 0.16, MeOH); HR-ESI-TOF-MS ; IR νmax (film) cm−1: 2960 and 2924 (CH), 1747 (C=O); 1H-NMR : δH 5.50 , 3.90 , 3.89 , 3.47 , 0.94 , 1.11 , 0.78 , 5.71 , 5.11 , 5.18 ), 4.92 ), 4.89 ), 1.47 , 2.26, 2.24, 2.22, 2.20, 2.16, 2.06, 2.04 × 2, 2.03, 2.02 × 2, 2.00, 1.98 × 2, 1.91 ; 13C-NMR  : δC 152.3 (C-22), 140.4 (C-5), 122.3 (C-6), 84.5 (C-16), 78.5 (C-3), 19.3 (C-19), 16.7 (C-27), 14.1 (C-18), 103.7 (C-20), 11.7 (C-21), 101.4 (C-1′′′ of Glc (III)), 101.2 (C-1 of Ara), 99.9 (C-1′′ of Glc (II)), 99.0 (C-1′ of Glc (I)), 97.8 (C-1 of Rha), 78.6 (C-3′ of Glc (I)), 77.6 (C-2′ of Glc (I)), 76.8 (C-4′ of Glc (I)), 63.1 (C-5 of Ara), 63.1 (C-6′ of Glc (I)), 62.4 (C-6′′′ of Glc (III)), 61.6 (C-6′′of Glc (II)), 17.5 (C-6 of Rha).Cell growth was measured with an MTT reduction assay, as described in a previous paper .1–4) were isolated from the bulbs of L. pumilum, and they were classified into furostanol glycosides (1 and 2) and Δ20(22)-pseudo-furostanol glycosides (3 and 4). Compounds 1–4 are novel steroidal glycosides with a 2,3,4-trisubstituted β-d-glucopyranosyl moiety at C-3 of the aglycone. In general, α-l-arabinopyranosyl groups are stable in a 4C1 conformation in glycosides, except for those glycosylated at C-2 [1 and 3, the α-l-arabinopyranosyl moiety is linked to C-3 of the inner trisubstituted β-d-glucopyranosyl group and is present as an usual 4C1 conformation. In contrast, in 2 and 4, the α-l-arabinopyranosyl moiety, which is attached to C-4 of the inner trisubstituted β-d-glucopyranosyl group, is present as a 1C4 conformation. It is notable that the 1C4 α-l-arabinopyranosyl moiety in 4 was converted to the 4C1 moiety on peracetylation (4a). These interesting steric behaviors of arabinopyranose cannot be explained by steric hindrance only. In a recent study, there are reports of the inner substituted 1C4 conformation of α-l-arabinopyranosyl moiety in triterpene glycoside [In conclusion, four new steroidal glycosides (d at C-2 . In 1 anlycoside ,15. Many


Clausena anisata led to the isolation and characterization three carbazole alkaloids: girinimbine, murrayamine-A and ekeberginine; two peptide derivatives: aurantiamide acetate and N-benzoyl-l-phenylalaninyl-N-benzoyl-l-phenylalaninate; and a mixture of two phytosterols: sitosterol and stigmasterol. The structures of these compounds were established by nuclear magnetic resonance  spectroscopy and electrospray ionization mass spectrometry (MS).Phytochemical investigations on the stem bark and roots of the tropical shrub  Clausena anisata (Will). Hook.f. ex .benth., is a tropical shrub or tree growing up to ten meters in height in and on the margins of evergreen forests carbazole, confirming the structure of 1 as that of the known alkaloid girinimbine, previously isolated from Murraya koenigii + ion, while all other compounds isolated formed adduct ions with sodium ([M+Na]+). Murrayamine-A (2), ekeberginine (3) and aurantiamide acetate (4) also formed [2M+Na]+ ions. The prominence of the sodium adducts in the mass spectra contrasts with the low abundance of sodium adducts reported in an extensive survey of the ESI(+)MS of natural products − ions obtained upon ESI(-)MS of girinimbine (1), murrayamine-A (2), and ekeberginine (3) most likely are formed by deprotonation at the carbazole N-H. The gas phase acidity of carbazole (ca. 1420 kJ mol−1  a a1–5. Cy+; MS/MS (CID 33%) of m/z 264: m/z 249 (55), 246 (18), 236 (100), 222 (65); ESI(+)TOF-MS m/z 264.1394 [M+H]+ . ESI(-)MS (relative intensity): m/z 262 [M–H]−; MS/MS (CID 38%) of m/z 262: m/z 279 (20), 247 (100), 246 (80); ESI(-)TOF-MS m/z 262.1233 [M–H]− .5–177 °C ). UV λmaFractions 8–11  were combined to give murrayamine-A  as white crystals, C18H17NO2, m.p. 166–167 °C (lit. 162–163 °C +, 302 (100) [M+Na]+; ESI(+)TOF-MS m/z 581.2341 [2M+Na]+ , 302.1120 [M+Na]+ . ESI(-)MS (relative intensity): m/z 278 [M–H]−; MS/MS (CID 36%) of m/z 278: m/z 250 (25), 223 (100), 222 (20); ESI(-)TOF-MS m/z 278.1173 [M–H]− .2–163 °C ). UV λmaFractions 30–33  were combined to give ekeberginine  as a brown powder, C19H19NO2, m.p. 227–228 °C (lit. 230–231 °C +, 316 (100) [M+Na]+; MS/MS (CID 24%) of m/z 316: m/z 298; ESI(+)TOF-MS m/z 609.2694 [2M+Na]+ , 316.1293 [M+Na]+ . ESI(-)MS (relative intensity): m/z 292 (100) [M–H]−, 277 (56); MS/MS (CID 30%) of m/z 292: m/z 277; MS/MS (CID 30%) of m/z 277: m/z 262 (38), 260 (14), 249 (72), 248 (30), 234 (100); ESI(-)TOF-MS m/z 292.1330 [M–H]− .0–231 °C ). UV λmaFractions 58–61 (crude methanol extract roots) (eluent: hexane/ethyl acetate/methanol) were combined to give aurantiamide acetate  as white crystals, C27H28N2O4, m.p. 183–184 °C   +, 467 (100) [M+Na]+; MS/MS (CID 31%) of m/z 467: m/z 449 (10), 407 (100), 385 (62), 274 (8), 224 (4); ESI(+)TOF-MS m/z 467.1936 [M+Na]+ .. 184 °C ). [α]D21= 0.041) ) 1H-NMR Fractions 64–68  were combined to give N-benzoyl-l-phenylalaninyl-N-benzoyl-l-phenylalaninate  as white crystals, C32H30N2O4, m.p. 209–210 °C  δ: 7.72 , 7.69 , 7.53 , 7.46 , 7.42 , 7.34 , 7.23–7.28 , 6.70 , 6.59 , 4.94 , 4.65 , 4.57 , 4.06 , 3.32 , 3.24 , 3.03 , 2.92 . 13C-NMR  δ: 172.9 , 168.4 and 168.2 , 138.2 , 136.8 , 135.2  134.3 , 133.0 , 132.4 , 130.3 , 130.2 , 129.9 , 129.7 , 129.7 , 129.4 , 128.4 , 128.1 , 128.1 , 127.8 , 66.4 , 55.5 , 51.3 , 38.5 , 38.3 . ESI(+)MS (relative intensity): m/z 529 [M+Na]+; MS/MS (CID 28%) of m/z 529: m/z 292 (100), 260 (10); ESI(+)TOF-MS m/z 529.2102 [M+Na]+ . = 0.14) ). 1H-NMR1), murrayamine-A (2) and ekeberginine (3)], two peptide derivatives [aurantiamide acetate (4) and N-benzoyl-l-phenylalaninyl-N-benzoyl-l-phenylalaninate (5)] and a mixture of sitosterol (6) and stigmasterol (7) were isolated from the stem bark and roots of the tropical shrub Clausena anisata. Compounds 1–5 have known biological activities, and 2, 4 and 5 have not been found previously as constituents of Clausena anisata.Three carbazole alkaloids [girinimbine (


R)-norsalvianolic acid L (1) and (S)-norsalvianolic acid L (2), was isolated from the Danshen Injection (lyophilized powder). The structural elucidation and stereochemistry determination were achieved by spectroscopic and chemical methods including 1D, 2D NMR  and circular dichroism experiments. Their antioxidant activities were assessed by the DPPH· and ABTS·+ scavenging methods in vitro with microplate assay. The IC50 values of 1 were 6.9 and 9.7 μM respectively, which was close to the control salvianolic acid B (7.8 and 7.1 μM respectively), while the IC50 values of isomer 2 were 27.1 and 25.3 μM, respectively.A pair of new phenolic acid stereoisomers, ( Salviae miltiorrhizae Radix et Rhizoma, also called Danshen, is an important commonly used Traditional Chinese Medicine. It activates and promotes blood circulation, and is widely used for the treatment of coronary heart diseases, hepatitis and dysmenorrhea , one set of AB coupling system proton signals [δ 6.80  and δ 6.73 ], together with one singlet aromatic proton signal at δ 7.68 . There still existed two sets of AX2 coupling protons . The 13C-NMR spectrum exhibited 26 signals, of which two carbons were methylenes, 11 were methines and 13 were quaternary carbons, as shown in the DEPT spectrum. Further analysis demonstrated two carbonyl signals at δ 171.1 (COOH-9′) and δ 166.8 (COO-17); 11 quaternary aromatic carbons at δ 148.2 (C-7), 145.6 (C-3′), 145.2 (C-13), 144.7 (C-4′), 144.2 (C-14), 143.4 (C-8), 136.1 (C-11), 129.1 (C-1′), 128.5 (C-3), 126.1 (C-10) and 122.7 (C-9); 11 methine carbons at δ 138.4 (C-4), 122.1 (C-5), 121.8 (C-6′), 119.5 (C-16), 117.5 (C-2′), 116.0 (C-5′), 115.6 (C-15), 115.1 (C-12), 113.4 (C-6), 73.8 (C-8′) and 37.2 (C-2); as well as two methylene carbons at δ 37.5 (C-7′) and 30.4 (C-1), as shown in Compound cids A–C ,16. The δ 7.68 (H-4) to 166.8 (COO-17)/136.1 (C-11)/128.5 (C-3)/126.1 (C-10)/122.7 (C-9)/37.2 (C-2), from δ 6.86 (H-2′) to δ 144.7 (C-4′)/121.8 (C-6′), from δ 6.80 (H-5) to δ 148.2 (C-7)/138.4 (C-4)/122.7 (C-9)/δ 143.4 (C-8), from δ 6.76 (H-5′) to δ 145.6 (C-3′)/δ 129.1 (C-1′), from δ 6.73 (H-6) to δ 148.2 (C-7)/143.4 (C-8)/126.1 (C-10), from δ 6.66 (H-6′) to δ 117.5 (C-2′), from δ 6.60 (H-12) to δ 144.2 (C-14)/119.5 (C-16)/115.7 (C-15), from δ 6.56 (H-15) to δ 145.2 (C-13)/136.1 (C-11), from δ 6.44 (H-16) to δ 144.2 (C-14)/115.1 (C-12). Additionally, there were also HMBC correlations from δ 5.11 (H-8′) to δ 171.1 (COOH-9′)/166.8 (COO-17)/129.1 (C-1′)/37.5 (C-7′), from δ 4.01 (H-2) to δ 138.4 (C-4)/136.1 (C-11)/128.5 (C-3)/122.7 (C-9)/119.5 (C-16)/115.1 (C-12), from δ 3.38/2.97(H-1) to δ 143.4 (C-8)/136.1 (C-11)/126.1 (C-10)/122.7 (C-9)/37.2 (C-2), as well as from δ 3.09/3.03 (H-7′) to δ 171.1 (COOH-9′)/129.1 (C-1′)/121.8 (C-6′)/117.5 (C-2′)/73.8 (C-8′) -CH2 (H-7′), CH (H-2)-CH2 (H-1), CH (H-5)=CH (H-6), CH (H-15) =CH (H-16) and CH (H-5′) =CH (H-6′). Based on the combined 1H-, 13C- NMR, HMBC, HSQC and 1H-1H COSY spectral data discussed above, the planar structure of compound 1 was elucidated as 3--2- propanoic acid. The HMBC spectrum presented correlation signals from 8 (C-8′) . MoreoveR) propanoic acid (R)-norsalvianolic acid L.There were two chromophores containing л → л* electron transition systems, which made the application of exciton-coupled circular dichroism (ECCD) feasible to determine the absolute configuration of C2. The CD spectrum  presenteR) . MeanwhiR) 20D −200° . Furthermore, the CD spectrum of 2 (S), while the negative Cotton effect 223 nm (+) was on the contrary to that of compound 1, so the absolute configuration of C8′ was determined as 8′(S). Thus, the structure of compound 2 was assigned as (S)-3--2-propanoic acid (S)-norsalvianolic acid L.In addition, the specific rotation of compound rum of 2  presenteoic acid , and it 50 values of compound 1 were 6.9 μM (DPPH) and 9.7 μM (ABTS), respectively. The result was close to the control salvianolic acid B (7.8 μM and 7.1 μM respectively). Meanwhile, the IC50 values of the isomer 2 were 27.1 and 25.3 μM, respectively  and ABTS  assays are commonly used radical scavenging assays to detect antioxidant capacity. These two methods are simple, rapid, sensitive and reproducible ,14,19. Iectively . The resd6) and with TMS as internal standard. ESIMS were measured on Agilent 6320 Ion Trap mass spectrometer. HRESIMS were measured on an Agilent 6210 TOF mass spectrometer. CD spectra were measured on a JASCO J-815 spectropolarimeter  with a 0.1 cm cell in methanol at room temperature at the following conditions: speed 200 nm/min, time constant 1 s, bandwidth 2.0 nm. Macroporous resin AB-8 , polyamide , Sephadex LH-20  and ODS  were used for column chromatography. Silica gel GF254 plates for TLC were purchased from Qingdao Marine Chemical Company, Qingdao, China. HPLC was carried out on an Elite P230 instrument equipped with a UV 230+ detector , employing a preparative column (Phenomenex Luna Su C18 (2) 21.2 × 250 mm, 5 μm) and an analytical column . In the antioxidant assay, a SpectraMax 190 Absorbance Microplate Reader  and 96 Well Cell Culture Cluster  were used. 1,1-Diphenyl-2-picrylhydrazyl (DPPH) and 2, 2′-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS) were purchased from Sigma . Solvents of analytical grade were purchased from Beijing Chemical Company, Beijing, China, while those of spectroscopy grade were from J. T. Baker . Danshen Injection (lyophilized powder) was provided by Second Chinese Medicine Factory, Harbin Pharm. Group Co. Ltd. . Reference standards of sodium danshensu and salvianolic acid B was from National Institutes for Food and Drug Control . Optical rotations were determined on an AUTOPOL IV Automatic Polarimeter . IR spectra were recorded on a Nicolet 5700 FT-IR spectrometer  by a microscope transmission method. UV spectra were obtained on an Agilent 1200 series UV spectrometer . 1D NMR and 2D NMR experiments were performed on Inova 500 spectrometers . Chemical shifts are given in δ (ppm) with solvent  to afford four fractions (Fr. A–D). Fr. B (26.1 g) was subsequently chromatographed over a polyamide column using a gradient of EtOH-H2O  as eluent to give four fractions (Fr. B1–B4). Fr. B3 (1.70 g) was purified by Sephadex LH-20 and ODS columns. Finally, compounds 1 (33 mg) and 2 (7 mg) were obtained by preparative HPLC with CH3CN-0.05%CF3COOH  as eluting mobile phase.The Danshen Injection (lyophilized powder) (120.7 g) was firstly dissolved in water, and then subjected to AB-8 macroporous resin column chromatography eluted by a gradient of EtOH–H· methanol solution  was added to 200 μL samples of different concentration (3.1‒100 μg/mL) of the test compounds. These solutions were gently mixed and incubated in dark for 30 min at room temperature. The absorbances of the resulting solutions were measured at 517 nm. For preparation of the standard curve, different concentrations of DPPH· methanol solutions (5–50 μg/mL) were used. The DPPH· concentration (μg/mL) in the reaction medium was calculated from the following calibration curve, determined by linear regression (r2: 0.9985):During the experiment, a microplate reader and 96-well plates were used to carry out the determination of the spectrum absorption values ,19. In t517-C is absorbance of a control with no radical scavenger and λ517-S is absorbance of the remaining DPPH· in the presence of scavenger.The scavenging capability of tested compounds was calculated using the following equation:·+ radical were performed by reacting ABTS (1.1 mg/mL) and potassium persulfate , and then storing in the dark for 6 h at room temperature [·+ (50 μL) was added to 200 μL samples of different concentrations (3.1–100 μg/mL). These solutions were gently mixed and incubated in dark for 30 min at room temperature. Then absorbances of the resulting solutions were measured at 734 nm. Different concentrations of ABTS·+ methanol solutions (55–220 μg/mL) were used to prepare the standard curve. The ABTS·+ concentration (μg/mL) in the reaction medium was calculated from the following calibration curve, determined by linear regression (r2: 0.9985):The ABTSperature ,19. Then734-C is absorbance of a control with no radical scavenger and λ734-S is absorbance of the remaining ABTS·+ in the presence of scavenger.The scavenging capability of tested compounds was calculated using the following equation:The bioactive constituents of Danshen Injection (lyophilized powder) were chemically studied and a new pair of isomers was isolated. Most of their spectral data were the same or similar to each other, except for the specific rotation and circular dichroism spectra. In our study, their radical scavenging capacities were measured by DPPH and ABTS assays. According to the results, both stereoisomers possessed potential antioxidant activity and that of compound 1 was closer to that of the control salvianolic acid B. Also it is interesting to find that the stereo structures do have effects on the antioxidant activities.


The structures of 1–3 were elucidated on the basis of their spectroscopic data. We also investigated the protective effect of compounds 1–13 on the neurotoxicity of PC12 cells induced by amyloid-beta (Aβ25–25), respectively. As a result, germacrane-type sesquiterpenoids 1–4 and lignans 5–7 were seen to afford protection against Aβ-induced toxicity in PC 12 cells. This study will contribute to revealing the chemical basis for the therapeutic effect of V. amurensis against AD.Three new germacrane-type sesquiterpenoids, heishuixiecaoline A–C (compounds  Valeriana has been taxonomically placed in the family Valerianaceae, comprising ca. 30 species in China , signals for two olefinic protons , and an oxygenated methine proton [δH 4.50 (H-8)], and an aldehydic proton [δH 9.30 (H-14)]. Analysis of its 13C-NMR and DEPT spectra showed 17 carbon resonances, including four methyls, three methylenes, three methines (one oxygenated), two trisubstituted double bond , a carbonyl carbon [δC 172.1 (C-16)], and an aldehydic carbon [δC 196.6 (C-14)]. The acetate group was determined to be located at C-8 by the HMBC correlations from H-8 (δH 4.50) to C-16, C-11 (δC 23.7), C-7 (δC 40.8), and C-9 (δC 47.2). The other correlations in the HMBC and 1H-1H-COSY spectra as shown in 1.The mpound 1  displaye1 were deduced by a ROESY experiment /CH3-15 indicated Δ4,5 and Δ1,10 to be Z- and E-configured, respectively, which was confirmed by a key NOE correlation of H-5 with H-1. Therefore, the structure of compound 1 was established as 10-methyl-6,7-dimethyl-methylene-8β-acetoxy-4-aldehyde--dicyclodecadiene , requiring five degrees of unsaturation. The IR spectrum displayed the presence of hydroxyl (3429 cm−1), α,β-unsaturated aldehyde (1689 cm−1), and double-bond (1624 cm−1) absorptions. The 1H- and 13C-NMR spectra data  and 21.3 (C-17) in compound 2, other differences such as the NMR data in C-7, C-8, and C-9 suggested hydroxy group is linked to C-8 in compound 2. The structure of 2 was further confirmed by the correlations of HMBC and 1H-1H COSY spectra as shown in Compound tra data  of 2 wer2 were deduced by a ROESY experiment /CH3-15, and H-5/H-1. Therefore, compound 2 was established as 10-methyl-6,7-dimethylmethylene-8α-hydroxy-4-aldehyde-)-dicyclodecadiene , indicating five degrees of unsaturation. The IR spectrum showed the presence of hydroxyl (3433 cm−1), α,β-unsaturated aldehyde (1682 cm−1), and double-bond (1631 cm−1) absorptions.Compound 3 could be assigned a similar structure to 2 by comparison of their 1H- and 13C-NMR spectra (3 and the appearance of an exocyclic double bond between C-10 (δC 149.0) and C-15 (δC 113.4) and the oxygenated methine signal at δC 68.5 (C-1) in 3. These were confirmed by the HMBC correlations from H-15  to C-1, C-9 (δC 37.5), and C-10. The other correlations in the HMBC and 1H-1H COSY spectra as shown in 3.Compound  spectra . The maj3 was determined to be the same as that of 2, with H-1 assigned as α-oriented by the correlations of H-7/H-9a (β-H) and H-1/H-9b (α-H). The NOE correlations of H-3a/H-5 confirmed the Z-configuration of the double bond between C-4 and C-5. Thus, the structure of compound 3 (named heishuixiecaoline C) was assigned as 6,7-dimethylmethylene-4-aldehyde-1β-hydroxy-10(15)-ene-(4Z)-dicyclodecylene  + .Heishuixiecaoline B (2): white amorphous powder; c 0.104, MeOH); UV (MeOH) λmax (logε) 261 nm; IR (KBr): 3429, 2938, 2813, 2706, 1689, 1624, 1452, 1370, 1184, 1002 cm−1; 1H-NMR (CD3OD) and 13C-NMR (CD3OD) data are shown in m/z 257.1514 [M+Na]+ .Heishuixiecaoline C (3): white amorphous powder; c 0.112, MeOH); UV (MeOH) λmax (logε) 266.9 nm; IR (KBr): 3433, 1682, 1631, 1443, 1381, 1264, 1190 cm−1; 1H-NMR (CD3OD) and 13C-NMR (CD3OD) data are shown in m/z 257.1510 [M+Na]+ .4 cells per well in growth medium for 24 h in 96-well plates, and then preincubated without or with various concentrations  of compounds 1–13, which was followed 24 h later by exposure to 20 μM aggregated Aβ25–35  prepared as described previously [t-test was used.Cell viability was measured by quantitative colorimetric assay with 3--2,5-diphenyltetrazolium bromide (MTT) method as described previously . Brieflyeviously . Vitamineviously . 25 μL/wV. amurensis based on its activity towards AD for the first time and three new germacrane-type sesquiterpenoids were obtained. Their structures were identified as 10-methyl-6,7-dimethylmethylene-8β-acetoxy-4-aldehyde--dicyclodecadiene (1), 10-methyl-6,7-dimethylmethylene-8α-hydroxy-4-aldehyde-)-dicyclodecadiene (2), and 6,7-dimethyl-methylene-4-aldehyde-1β-hydroxy-10(15)-ene-(4Z)-dicyclodecylene (3), respectively. Seven known cpompounds were also isolated and identified. All these compounds were screened and cell tests showed that germacrane-type sesquiterpenoids 1–4 and lignans 5–7 were the active components of Valeriana amurensis against AD.We investigated the chemical constituents of


Preliminary SARs were obtained, which are in good agreement with the molecular binding model and should prove helpful for future antagonist design. The novel scaffold presented here might also be useful in the development of maraviroc-derived second generation CCR5 antagonists.CCR5, as the major co-receptor for HIV-1 entry, is an attractive novel target for the pharmaceutical industry in the HIV-1 therapeutic area. In this study, based on the structures of maraviroc and 1,4-bis(4-(7-chloroquinolin-4-yl)piperazin-1-yl)butane-1,4- dione ( Almost a decade ago now, the chemokine receptor CCR5 was identified as the major co-receptor for HIV-1 entry, besides the cellular CD4 receptor ,3,4,5. C®, In the last decade, numerous small molecule CCR5 antagonists have been reported. The discovery and development of CCR5 antagonists have been systmatically reviewed by Palani . They in1 (50 = 2.00 μM). Mono-4-(piperazin-1-yl)quinoline derivatives have previously been reported as CCR5 antagonists 28D = -15.6 (c = 1.13 in CH2Cl2) (Lit. 22D =tert-Butyl (S)-2-(methoxycarbonyl)-1-phenylethylcarbamate (6). To a cold (0 °C) mixture of Na2CO3  in THF/H2O  was added 5 in one portion . A solution of (BOC)2O  in THF (4 mL) was added in one portion and the reaction stirred at 0 °C for 1 h, then at 25 °C for 2 h, extracted with EtOAc (3 x 20 mL). The combined organic extracts were dried and concentrated in vacuo. The crude white solid product 6 was used in the next step without further purification.tert-Butyl (S)-2-formyl-1-phenylethylcarbamate (7). To a soln. of 6  in DCM (10 mL) cooled to -78 °C was added DIBAL-H  dropwise at a rate that maintained the temperature below -70 °C. After 2 h the reaction was quenched by the addition of MeOH (1.7 mL) and H2O (1 mL) at -78 °C, then allowed to warm to 25 °C. The mixture was filtered. The filtrate was dried and concentrated in vacuo. The crude colorless oil 7  was used in the next step without further purification. 1H-NMR (CDCl3) δ 1.50 , 2.85-3.05 , 5.10 , 7.25-7.40 , 9.80 .Tert-butyl (S)-3-(4-(7-chloroquinolin-4-yl)piperazin-1-yl)-1-phenylpropylcarbamate (8). To a solution of 7 , 3a  and HOAc (0.14 mL) in DCM (10 mL) was added NaBH(OAc)3  in one portion and the mixture was stirred for 5 h. The reaction was quenched by adding 10% K2CO3 soln. (20 mL) and stirred for an additional 30 min, extracted with DCM (3 x 20 mL). The combined organic layer was dried and concentrated in vacuo. The residue was purified by flash column chromatography on silica gel, eluted with a mixture of MeOH/DCM , to afford 8  as a yellow foam. 1H-NMR (CDCl3) δ 1.35 , 2.15 , 2.61 , 2.91 , 3.35 , 4.69 , 6.80 , 7.22-7.32 , 7.40 , 7.81 , 8.08 , 8.65 .(S)-3-(4-(7-Chloroquinolin-4-yl)piperazin-1-yl)-1-phenylpropan-1-amine (9). A solution of 8  in saturated HCl in MeOH (6 mL) heated at 65 °C for 2 h. The MeOH was evaporated under reduced pressure and the residue cautiously partitioned between DCM (20 mL) and 20% K2CO3 solution (20 mL). The aqueous layer was re-extracted with DCM (3 x 20mL). The combined organic layer was dried and concentrated in vacuo, to afford 9  as a straw yellow oil which was used in the next step without further purification. 1H NMR (CDCl3) δ 1.98-2.12 , 2.58 , 2.81 , 3.25 , 4.18 , 6.81 , 7.20 , 7.32 , 7.35-7.44 , 7.90 , 8.05 , 8.68 .


Ricinodendron heudelotii on selected cancer cell lines. The leaves of the plant were extracted with ethanol and partitioned in sequence with petroleum ether, ethyl acetate, and n-butanol. The ethyl acetate fraction was phytochemically studied using thin layer chromatography (TLC) and column chromatography (CC). Structural elucidation of pure compounds obtained from the ethyl acetate fraction was done using mass spectra, 1H-NMR, and 13C-NMR analysis. The isolated compounds were subsequently screened using five different cancer cell lines: HL-60, SMMC-7721, A-549, MCF-7, SW-480, and normal lung epithelial cell line, BEAS-2B, to assess their cytotoxic effects. Nine compounds were isolated and structurally elucidated as gallic acid, gallic acid ethyl ester, corilagin, quercetin-3-O-rhamnoside, myricetin-3-O-rhamnoside, 1,4,6-tri-O-galloyl glucose, 3,4,6-tri-O-galloyl glucose, 1,2,6-tri-O-galloyl glucose, and 4,6-di-O-galloyl glucose. Corilagin exhibited the most cytotoxic activity with an IC50 value of 33.18 μg/mL against MCF-7 cells, which were comparable to cisplatin with an IC50 value of 27.43 µg/mL. The result suggests that corilagin isolated from R. heudelotii has the potential to be developed as an effective therapeutic agent against the growth of breast cancer cells.This study was designed to explore the in vitro anticancer effects of the bioactive compounds isolated from  Ricinodendron heudelotii (family: Euphorbiceae) plant parts have been used in traditional medicine to treat cough, stomach ailments, and pain related to child birth. Decoctions and infusions are mostly prepared from the leaves and stem bark. Traditional practitioners prescribe it for women suffering from miscarriages −, 1H-NMR : δ 7.10 , 13C-NMR : 121.0 (C-1), 109.0 , 145.9 , 138.3 (C-4), 168.0 (C-7).Gallic acid (2): Off-white amorphous powder, C9H10O5, negative ESI-MS m/z: 198 [M]−, 1H-NMR : δH 0.84 , 3.82  7.05 ; 13C-NMR : δC 120.7 (C-1); 109.4 , 138.0 (C-4), 144.9 , 166.8 (C-7), 60.6 (C-1′), 14.1 (C-2′).Gallic acid ethyl ester (3): Off-white amorphous powder, C27H22O18,negative ESI-MS m/z 633 [M − H]−, 1H-NMR : δH 7.05 ,6.69, 6.65 , 6.36 , 4.95 , 4.80 , 4.51 , 4.46 , 4.16 , 3.98 ; 13C-NMR : HHDP δC 167.1 and 167.6 (C=O), 145.8, 145.1 , 144.9, 144.3 , 136.3, 136.2 , 116.2, 116.1 , 124.1, 123.8 , 107.3, 106.7 ; galloyl δC 165.2 (C=O), 146.2 , 139.7 (C-4′), 119.4 (C-1′), 109.7 ; glucose δC 92.9 (C-1), 78.3 (C-3), 76.6 (C-5), 72.4 (C-2), 64.2 (C-6), 62.8 (C-4).Corilagin (O-rhamnoside (4): Yellow amorphous powder, C21H20O11; negative ESI-MS m/z 447 [M − H]−, 1H-NMR : δ 7.33 , 7.31 , 6.91 , 6.19 , 6.36 , 5.34 , 4.22 , 3.76 , 3.34 , 3.41 , 0.94 ; 13C-NMR : δC 18.7 (C-6″), 71.1 (C-5″), 70.6 (C3″), 71.1 (C-2″), 71.2 (C-4″), 94.3 (C-8), 98.7 (C-6), 102.1 (C-1″), 104.1 (C-10), 115.7 (C-2′), 115.8 (C-5′), 120.8 (C-1′), 122.2 (C-6′), 134.3 (C-3), 145.4 (C-3′), 148.8 (C-4′), 156.7 (C-2), 157.3 (C-9), 161.4 (C-5), 165.6 (C-7), 177.8 (C-4).Quercetin-3-O-rhamnoside (5): Yellow amorphous powder, C21H20O12, negative ESI-MS m/z: 463 [M − H]−, 1H-NMR : δ 6.95 , 6.36 , 6.20 , 5.31 , 3.80 , 3.55 , 3.17 , 3.34 , 0.84 . 13C-NMR : δC 177.6 (C-4), 163.8 (C-7), 161.4 (C-5), 157.1 (C-9), 156.6 (C-2), 145.9 , 135.4 (C-4′), 135.2 (C-3), 119.7 (C-1′), 107.9 (C-2′), 107.9 (C-6′), 104.3 (C-10), 102.4 (C-1″), 98.8 (C-8), 93.7 (C-6), 71.9 (C-4″), 71.3 (C-5″), 70.8 (C-3″), 70.3 (C-2″), 17.8 (C-6″).Myricetin 3-O-galloyl-β-d-glucose (6): Off-white amorphous powder, C27H24O18, negative ESI-MS m/z 635 [M − H]−; 1H-NMR : δH 7.15, 7.10, 7.06 , 5.78 , 5.22 , 4.45 , 4.22 , 4.06 , 3.84 , 3.65 .1,4,6-Tri-O-galloyl-d-glucose (7): Off-white amorphous powder, C27H24O18, negative ESI-MS m/z 635 [M − H]−, 1H-NMR : δH 7.07, 6.99, 6.94 , 4.42 , 4.26 , β-glucose form: δH 4.76 , 3.56 , 5.45 , 5.35 , 4.07 , α-glucose form: δH 5.25 , 3.81 , 5.68 , 5.36 , 4.47 .3,4,6-Tri-O-galloyl-β-d-glucose (8): Off-white amorphous powder, C27H24O18, negative ESI-MS: m/z 635 [M − H]−, 1H-NMR : δH 7.10, 7.04, 7.00 , 5.92 , 5.23 , 4.56 , 4.47 , 3.83 , 3.81 , 3.66 ; 13C-NMR : δC 63.1 (C-6), 69.6 (C-4), 72.7 (C-5), 74.4 (C-3), 74.2 (C-2), 93.2 (C-1), 117.0, 119.1, 119.2 , 109.2, 108.8, 108.8 , 145.3, 145.4, 145.8 , 138.6, 138.7, 139.1 , 164.1, 164.9, 165.7 (C=O).1,2,6-Tri-O-galloyl-β-d-glucose (9): Off-white amorphous powder, C20H20O14, negative ESI-MS: m/z 483 [M − H]−, 1H-NMR : δH 7.09, 7.07, 7.08, 7.06 , 5.13 , 4.39 , 4.18 , α-glucose form: 5.17 , 3.98 , 4.53 , β-glucose form: δH 4.60 , 3.89 , 3.70 , 3.31 .4,6-Di-1–5 were applied to HL-60, SMMC-7721, A-549, MCF-7, and SW-480 cancer cells to assess their cytotoxic effects. Compound 3 (Corilagin) exhibited the most cytotoxic activity against MCF-7 cells with IC50 values of 33.18 and 25.81 µg/mL for HL-60 cells. The IC50 value of compound 3 in MCF-7 cells is comparable with the positive control drug (cisplatin), having an IC50 value of 27.43  wasless than 50% at the concentration of 40 µM .R. heudelotii and its potential to be developed for use for cancer treatment.After many years of research on cancer, cancer still remains a destructive disease responsible for over one quarter of the deaths recorded globally. It is one of the major health obstacles, representing the second largest cause of mortality . This is3) was isolated and structurally identified from the ethyl acetate fraction of the ethanolic extract of R. heudelotii leaves in our study. Only compound 3 showed significant and comparable cytotoxic property relative to cisplatin. Corilagin has been previously identified in several other plants, especially those of the Euphorbiaceae family. It was firstly isolated from Caesalpinia coriaria in 1951, hence the name of the molecule [Terminalia arjuna bark extract induced apoptosis in A549 cells and in human breast adenocarcinoma MCF-7 cells via blockage of cell-cycle progression in the G0/G1 phase. The cytotoxic properties of tannins are characterised by the release of reactive oxygen species (ROS), which is followed by a reduction in the thioredoxin, superoxide dismutase, and in the level of redox active proteins [3. The chemopreventive activity of tannins, especially the ellagitannins and hydrolysable tannins, has been linked to activation of apoptosis [cdc 2 after corilagin treatment on cancer cells was discovered [cdc 2, and pAKT. Selectivity index (SI) signifies the differential activity of compounds; the greater the SI value, the more selective the compound [Corilagin  operating at 600 MHz for 1H-NMR, and 150 MHz for 13C-NMR, respectively. Coupling constants are expressed in hertz (Hz), and chemical shifts are given on a δ  scale with tetramethylsilane (TMS) as an internal standard. ESI mass spectra were recorded on a VG Auto Spec-300 spectrometer . Column chromatography (CC) was performed on Sephadex LH-20 , MCI-gel CHP20P , and Toyopearl HW-40F . Thin-layer chromatography (TLC) was performed on silica gel H-precoated plates, 0.2–0.25 mm thick , with benzene–ethylformate–formic acid , and compounds were detected by spraying with 2% ethanolic FeCl3 and 10% sulfuric ethanol solution followed by heating.Ricinodendron heudelotii leaves were collected within Covenant University premises Ota, Ogun, Nigeria. The plant was identified by Dr. J.O. Popoola  and a voucher specimen was prepared and submitted in the Forest research Institute of Nigeria, Ibadan with voucher number, FHI 110573. Leaves were dried at room temperature (25 °C) and blended using an electric blender into a coarse powder.n-butanol (8 L). The solvent fractions were concentrated to dryness to afford four (4) fractions as: Pet, EtOAc, n-butanol, and aqueous fractions. The EtOAc fraction was selected for subsequent isolation procedures.The powdered leaf samples (10 kg) were extracted using 95% ethanol by macerationat room temperature, the filtrate was further condensed under reduced temperature and pressure, and the yield of the extract obtained was 11.75%. The concentrated crude extract (1 kg) was suspended in 1 L distilled water and partitioned in sequence with petroleum ether , ethyl acetate , and 1 (8.53 mg), 2 (2.1 mg), and 3 (12 mg). Fraction 12 (3.8 g) was subjected to repeated column chromatography also eluting with H2O and methanol (60:1) to afford compound 4 (98 mg) and 5 (13.2 mg). Compounds 6 (9.9 mg), 7 (2.6 mg), 8 (2.7 mg), and 9 (4.4 mg) were obtained from fraction 13 (14.5 g) after repeated purification by chromatography on a Sephadex LH-20, MCI-gel CHP20P and Toyopearl HW-40F columns eluting with H2O/methanol.The ethylacetate fraction (246 g) was loaded into a column containing Sephadex LH-20 (800 g). The eluent was a mixture of water and methanol in different ratios starting with 100:0 to 0:100. Fractions with similar TLC patterns were combined to afford 13 fractions. Fraction 11 (36 g) was chromatographed on a Sephadex LH-20 column and MCI-gel CHP20P eluting with water:methanol (4:1) to afford compounds 2 incubator for 48 h and the cell density was adjusted to 5 × 104 cells/well.Cancer cell lines, HL-60, SMMC-7721, A-549, MCF-7, and SW-480, were obtained from the American Type Culture Collection Center (ATCC) . They were cultured in RMPI-1640 or DMEM medium supplemented with 10% bovine serum albumin . Prior to exposure to drugs, the cancer cell lines were cultured in a CO2O depending on their solubility. Samples (2 µL) were suspended in centrifuge tubes containing 48 µL of complete medium to make a final volume of 100 µL.Compounds (1 mg/mL) were dissolved in either DMSO or HH-tetrazolium, inner salt (MTS) method by measuring the absorbance of formazan precipitate formed in the cells based on MTS reduction [50, which was calculated using Reed and Muench’s method [50 was calculated by the following equation:50 = 10 Log10(IC50); IH: I% above 50%; IL: I% below 50%; CH: High drug concentration; and CL: Low drug concentration.The cytotoxicity assay was evaluated using the 3--5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2eduction . In thiss method ,47. The 50 value of the compound on a normal cell line to the IC50 value of the compound on cancer cell lines [The selectivity index (SI) for the cytotoxicity of corilagin was calculated using the ratio of the ICll lines .Samples are expressed as mean ± standard deviation or standard error of mean.R. heudelotii on a breast cancer cell line, and its potential for development as an anticancer agent to reduce the burden of cancer diseases globally.In conclusion, we report significant cytotoxic activities of corilagin isolated from leaf extract of


The performance of these devices has been analyzed, showing higher efficiencies of those sensitized with ZnP compounds. These results have been explained, on one hand, taking into account the electronic character of the metal ion, which acts as mediator in the injection step, and, on the other, considering the number of anchoring groups, which determines both the stereoelectronic character of the dye and the way it binds to TiO2 surface.A series of porphyrin compounds, free base (H UV-vis (THF) λmax/nm (log Ԑ): 421 (5.58), 515 (4.29), 549 (3.88), 591 (3.68), 649 (3.18). HRMS (MALDI-TOF): m/z calcd, for C54H46N4O10 ([M+]): 910.32, found 910.45. 2P-(CO2Me)3 9).5,10,15-tris(4-methoxycarbonylphenyl)-20-porphyrin (H1H NMR (CDCl3: 300 MHz), δ ppm: −2.80 , 3.97 , 4.12  4.18 , 7.46 , 8.31 , 8.43 , 8.81 , 8.98 . UV-vis (THF) λmax/nm (log Ԑ): 419 (5.44), 514 (4.13), 550 (3.73), 590 (3.48), 649 (2.30). HRMS (MALDI-TOF): m/z calcd, for C53H42N4O9 ([M+]): 878.30, found 878.64.4 and evaporated. The residue was recrystallized in hexane to get the pure powder. 50 mg of free-base porphyrin was dissolved in a mixture of THF/MeOH (20/14 mL) and 6 mL of NaOH 20% aqueous solution were added. The crude was heated for 2 h. After cooling, the reaction crude was then diluted with dichloromethane and washed, first with HCl (1M) and then with water. The organic layer dried over MgSO2P-(CO2H)22, to isolate cis and trans isomers .Column chromatography was conducted for H2P-CO2H 1).5-(4-carboxyphenyl)-10,15,20-trisporphyrin (H Yield: 95%. 1H NMR (DMSO: 300 MHz), δ ppm: −2.93 , 3.90 , 4.00 , 7.53 , 8.33 , 8.39 , 8.81 , 8.96 . UV-vis (THF) λmax/nm (log Ԑ): 420(5.38), 515(4.13), 550(3.70), 592(3.70), 650 (3.30). HRMS (MALDI-TOF) calcd for C54H48N4O11 ([M+]): 928.33, found 928.38.2P-(CO2H)2 2-t).5,10-bis(4-carboxyphenyl)-15,20-bisporphyrin, trans isomer (trans-H Yield: 23%. 1H NMR (DMSO: 300 MHz), δ ppm: −2.93 , 3.89 , 3.99 , 7.54 , 8.33 , 8.38 , 8.82 , 8.98 . UV-vis (THF) λmax/nm (log Ԑ): 420 (5.59), 515 (4.27), 550 (3.93), 592 (3.78), 651(3.65). HRMS (MALDI-TOF): m/z calcd, for C52H42N4O10 ([M+]): 882.29, found 882.518.2P-(CO2H)2 2-c).5,10-bis(4-carboxyphenyl)-15,20-bisporphyrin, cis isomer (cis-H Yield: 71%. 1H NMR (DMSO: 300 MHz), δ ppm: −2.93 , 3.90 , 3.99 , 7.53 , 8.32 , 8.38 , 8.83 , 8.98 . UV-vis (THF) λmax/nm (log Ԑ): 420 (5.59), 515 (4.27), 550 (3.93), 591 (3.78), 649 (3.65). HRMS (MALDI-TOF): m/z calcd, for C52H42N4O10 ([M+]): 882.29, found 882.52.2P-(CO2H)3 3).5,10,15-tris(4-carboxyphenyl)-20-porphyrin (H Yield: 94%. 1H NMR (DMSO: 300 MHz), δ ppm: −2.93 , 3.90 , 3.99 , 7.55 , 8.35 , 8.38 , 8.85 , 8.98 . UV-vis (THF) λmax/nm (log Ԑ): 419(5.52), 515(4.18), 549(3.78), 590(3.54), 649 (3.40). HRMS (MALDI-TOF): m/z calcd, for C50H36N4O9 ([M+]): 836.25, found 836.50.7, 8 or 9, 50 mg) and zinc acetate (1:5 mol ratio) were refluxed in dichloromethane and methanol  until free-base porphyrin was completely metalated, checked by TLC and UV-vis absorption spectroscopy. The reaction mixture was then diluted with dichloromethane and washed, first with HCl (1 M) and then with water. The organic layer was collected and, after evaporation of solvent, the crude compound was purified by silica gel column using hexane/ethyl acetate eluent mixtures. Quantitative yields were obtained in all cases and isolated compounds were hydrolized following the same procedure used for free-base porphyrins hydrolisis (see Free-base porphyrin (isis see .2H)25 to isolate cis and trans isomers .Column chromatography was conducted for ZnP-(CO2H 4Zinc(II) 5-(4-carboxyphenyl)-10,15,20-trisporphyrinate (ZnP-CO). Yield: 98%. 1H NMR (DMSO: 300 MHz), δ ppm: 3.90 , 3.99 , 7.45 , 8.29 , 8.37 , 8.75 , 8.90 . UV-vis (THF) λmax/nm (log Ԑ): 426 (5.63), 557 (4.20), 597 (3.65). HRMS (MALDI-TOF) calcd for C54H46N4O11 ([M+]): 990.25, found 989.22.2H)2 5-t).Zinc(II) 5,10-bis(4-carboxyphenyl)-15,20-bisporphyrinate, trans isomer (trans-ZnP-(CO Yield: 20%. 1H NMR (DMSO: 300 MHz), δ ppm: 3.89 , 3.98 , 7.46 , 8.23 , 8.35 , 8.76 , 8.91 . HRMS (MALDI-TOF): m/z calcd, for C52H40N4O10Zn ([M+]): 944.20, found 943.45.2H)2 5-c).Zinc (II) 5,10-bis(4-methoxycarbonylphenyl)-15,20-bisporphyrinate, cis isomer (cis-ZnP-(CO Yield: 69%. 1H NMR (DMSO: 300 MHz), δ ppm: 3.89 , 3.98 , 7.45 , 8.23 , 8.34 , 8.76 , 8.91 . UV-vis (THF) λmax/nm (log Ԑ): 426 (5.56), 557 (4.29), 597 (3.80). HRMS (MALDI-TOF): m/z calcd, for C52H40N4O10Zn ([M+]): 944.20, found 943.48.2H)3 6).Zinc(II) 5,10,15-tris(4-carboxyphenyl)-20-porphyrinate (ZnP-(CO Yield: 98%. 1H NMR (DMSO: 300 MHz), δ ppm: 3.90 , 3.99 , 7.47 , 8.29 , 8.37 , 8.78 , 8.94 . UV-vis (THF) λmax/nm (log Ԑ): 426 (5.70), 557 (4.33), 598 (3.89). HRMS (MALDI-TOF): m/z calcd, for C50H34N4O9Zn ([M+]): 898.16, found 897.40.2 photoanodes were prepared coating pastes of anatase TiO2 nanoparticles having two different diameters, 20 nm (Dyesol’s 90 T) and 400 nm (Dyesol’s WER2-O), onto TiCl4 treated FTO glass plates , by repetitive screen printing to obtain the required thickness. These electrodes were gradually heated under a programmed flow: at 370 °C for 10 min and 450 °C for 10 min. Their apparent surface area was 0.16 cm2 (0.4 cm × 0.4 cm), and revealed a total thickness of 8–10 μm, containing a 3–4 μm scattering layer. The TiO2 electrodes were treated again with TiCl4 under 70 °C for 30 min and sintered at 500 °C for 30 min, before they were dipped into dye solution. The nanocrystalline TiO2 films were immersed into 5 mM dye solutions, without any other additives, i.e. co-adsorbents, and kept at RT for 20 h. Finally, dye adsorbed TiO2 photoanodes and thermally platinized and drilled FTO counter electrodes , were assembled into sandwich type cells, separated by a 30 μm thick hot-melt gasket , and sealed by heating. An electrolyte solution  was introduced in the assembled devices. A series of three devices with each dye were prepared and their photovoltaic performance measured. The values described are, in all cases, the best obtained, with no significant differences between devices sensitized with the same dye, which ensures the reproducibility and consistency of the results.Double-layered nanoporous TiO2 , prior to every measurement, using a calibrated photovoltaic reference cell . The applied potential and cell current were registered with a Keithley 2401 low voltage digital sourcemeter. The incident photon-to-current conversion efficiency (IPCE) was measured as a function of wavelength from 400 to 800 nm by using an IPCE-DC system (Lasing SA).An ABET 150W xenon light source equipped with an AM 1.5 G correcting filter was employed. The light intensity was adjusted to 100 mW/cm2P-CO2H 1, H2P-(CO2H)22 (cis and trans) and H2P-(CO2H)33 were prepared through a two-step synthetic sequence, as it is described in 7–9) were obtained as pure compounds, following the traditional Adler-Longo method  and Zn metallated [ZnP-(CO2H)n4–6], with one, two or three carboxyphenyl anchoring groups, were synthesized, characterized and used as sensitizers in TiO2-based DSSC devices. The comparison of their performances shows the utility of these compounds for this use, reflecting that ZnP-(CO2H)n4–6 sensitized solar cells offer better efficiencies, compared to those sensitized with H2P-(CO2H)n1–3. This is due to the presence of a zinc ion in the porphyrin inner cavity, acting as electronic mediator in the injection step. The observed order of efficiency for the zinc complexes, i.e. ZnP-CO2H 4 > ZnP-(CO2H)36 > cis-ZnP-(CO2H)2, 5-c is in agreement with a rapid injection of the photoexcited electrons to the TiO2 conduction band, where electronic characteristics of the dye prevail over its structural features. On the other hand, the observed order of efficiency for free-base dyes, i.e., cis-H2P-(CO2H)22-c > (H2P-(CO2H)33 > H2P-CO2H 1, agrees with the absence of a metallic mediator, slowing down the injection step and making structural features of the dye to gain prominence over its electronic characteristics.A series of unsymmetric porphyrin compounds, free-base [H


Of these; costunolide (2), dehydrocostus lactone (3) and scopoletin (4) showed significant inhibition on seedling growth in a concentration-dependent manner. Detection and observation demonstrated that the antioxidase system was found to be affected by these chemicals, resulting in the accumulation of ROS and membrane damage. To investigate their release ways, the compounds were traced back and volumes quantified in rhizosphere soil and plant tissues. This work made clear the chemical bases and their physiological effects on the plants. These chemicals were found to be the secondary metabolites of the plants and included in the rhizosphere soil. The findings identified a potential pathway of plant-plant interactions, which provided theoretical basis to overcoming replanting problems. This research was also useful for exploring ecological effects of allelochemicals in green agriculture.Allelochemicals are the media of allelopathy and form the chemical bases of plant-environment interactions. To determine true allelochemicals and their autotoxic effects, seven compounds were isolated and identified from  Saussurea lappa (Decne.) Sch. Bip. is an alpine medicine plant frequently used as an anti-inflammatory, digestion aid and liver protection agent in Traditional Chinese Medicine +, C17H34NaO2, measured m/z 293.2447, calculated m/z 293.2451, err 1.5 ppm. 1H-NMR (CDCl3) δ 0.88 , 1.261–0.28 , 1.62 , 2.30 , 3.66 . 13C-NMR (CDCl3) δ 14.09 (C-16), 22.6934–0.12 (C: 215–), 51.39 (C-17), 174.29 (C-1).Costunolide (2): HR-ESI-MS: [M + Na]+, C15H20NaO2, measured m/z 255.1365, calculated m/z 255.1356, err −3.9 ppm. 1H-NMR (CD3COCD3) δ 1.47 , 1.72 , 1.741−.81 , 2.182–0.37 , 2.66 , 4.69 , 4.82 , 4.90 , 5.61 , 6.10 . 13C-NMR (CD3COCD3) δ 15.39 (6-CH3), 16.47 (10-CH3), 25.89 (C-8), 27.68 (C-4), 39.07 (C-5), 40.80 (C-9), 50.13 (C-3), 81.34 (C-12), 118.10 (2-CH2), 126.59(C-7), 127.88 (C-11), 137.11 (C-6), 140.61 (C-10), 141.08 (C-2), 169.61 (C-1).Dehydrocostus lactone (3): HR-ESI-MS: [M + Na]+, C15H18NaO2, measured m/z 253.1190, calculated m/z 253.1199, err 3.5 ppm. 1H-NMR (CDCl3) δ 1.371–0.97 , 2.122−.58 , 2.842−.95 , 4.82 , 4.90 , 5.27 , 5.07 , 5.49 , 6.22 . 13C-NMR (CDCl3) δ 30.32 (C-8), 30.95 (C-4), 32.61 (C-9), 36.27(C-5), 45.15 (C-3), 47.64 (C-7), 52.05(C-11), 85.25 (C-12), 109.64 (10=C), 112.64 (6=C), 120.19 (2=C), 139.78 (C-2), 149.25 (C-6), 151.25 (C-10), 170.27 (C-1).Scopoletin (4): HR-ESI-MS: [M + Na]+, C10H8NaO4, measured m/z 215.0316, calculated m/z 215.0315, err 0.8 ppm. 1H-NMR (CDCl3) δ 3.96 , 6.27 , 6.85 , 6.92 , 7.60 . 13C-NMR (CDCl3) δ 56.42 (6-OCH3), 103.21 (C-8), 107.50 (C-5), 111. 50 (C-4), 113.42 (C-2), 143.32 (C-3), 144.02 (C-7), 149.72 (C-6), 150.27 (C-9), 161.47 (C-1).Syringaldehyde (5): HR-ESI-MS: [M + Na]+, C9H10NaO4, measured m/z 205.0467, calculated m/z 205.0471, err 2.0 ppm. 1H-NMR (CDCl3) δ 3.98 , 6.12 , 7.16 , 9.82 . 13C-NMR (CDCl3) δ 56.49 , 106.70 , 128.41 (C-1), 140.83 (C-4), 147.37 , 190.78 (CHO).5-Hydroxymethylfurfural (6): HR-ESI-MS: [M + Na]+, C6H6NaO3, measured m/z 149.0208 calculated m/z 149.0209, err 0.7 ppm. 1H-NMR (CDCl3) δ 2.77 , 4.72 , 6.52 , 7.23 , 9.56 .13C-NMR (CDCl3) δ 57.59 (5-C-OH), 110.02 (C-4), 122.95 (C-3), 152.35 (C-2), 160.71 (C-5), 177.74 (CHO).Chlorogenic acid (7): HR-ESI-MS: [M + Na]+, C16H18NaO9, measured m/z 377.0844, calculated m/z 377.0843, err 0.2 ppm. 1H-NMR (CD3OD) δ 2.012–0.09 , 2.142–0.24 , 3.73 , 4.16 , 5.32 , 6.25 , 6.77 , 6.94 , 7.04 , 7.55 . 13C-NMR (CD3OD) δ 36.81 (C-6), 37.38 (C-2), 69.90 (C-5), 70.58 (C-3), 72.07 (C-4), 74.77 (C-1), 113.80 (C-2′), 113.86(C-5′), 115.09 (-C=), 121.59 (C-6′), 126.40 (C-1′), 145.40 (=C-Ar), 145.70 (C-4′), 148.18 (C-3′), 167.28 (-CO-), 175.66 (COOH).S. lappa and lettuce were immersed in warm water at 37 °C for 3 h to accelerate germination, then sterilized with 5% sodium hypochlorite solution for 2 min and washed with distilled water to remove the residue of solution. The seeds were germinated on wet filter paper and kept humid in dark at 25 °C for 3 days and 24 h for S. lappa and lettuce, respectively. Uniformly germinated seeds were cultured in 6-well plates for further tests when their coats were just broken. Crude extract was dissolved in DMSO, diluted with water and administered on the seedlings at concentrations of 400, 200, 100 and 50 μg/mL, respectively, with three replicates for each treatment. The concentrations of DMSO were all less than 1% (v/v) in the samples, and the solution containing only the same concentration of DMSO was used in the controls. Purified compounds were used with the same method at the concentrations of 200, 100, 50 and 25 μg/mL. After being incubated in a constant-temperature humidity chamber in dark at 25 °C , the root length, stem length and fresh weight of S. lappa and lettuce seedlings were measured. The inhibition ratios were calculated according to the following formula:Inhibition to the seedlings was used for evaluating the bioactivity of the purified compounds. Seeds of S. lappa and lettuce were ground in cold PBS  and centrifuged  at 4 °C. The suspensions were used for the spectrophotometric determination methods [Peroxidase (POD), superoxide dismutase (SOD) and malondialdehyde (MDA) are all important physiological indexes in plant and usually used for the investigation of antioxidant activity and plasmolemma structure status. Seedling roots of  methods ,46,47. P methods . The resH-tetrazolium monosodium salt solution , xanthine oxidase ammonium sulfate suspension , catalase solution  and hypoxanthine solution  were added into the reaction system. For the control groups, all the conditions were the same except the sample suspension was replaced by distilled water. The reaction system was incubated at 25 °C for 30 min and measured at 560 nm. SOD activity was calculated by the formula:SOD activities were measured by a modified hydroxylamine method . SuspensMDA concentration was tested by the thiobarbituric acid (TBA) method . TBA sol2 showing high allelopathic activity was used to treat the seedlings, comparing with the controls only treated with DMSO. Freehand sections of the roots were washed with distilled water four times and labeled for 8 min in dark with DCFH-DA solution. Then the sections were washed with distilled water six times to remove the dye residue and viewed under a fluorescence microscope .ROS in the cells of treated seedlings were observed by the method in  with slip value, 0.05.Data was expressed as the mean ± standard error (SE). Variance analysis to the values was conducted by SPSS Statistics . Results of inhibition to the seedling growth were subject to one-way analysis of variance (ANOVA) with a Dunnett’s test at probability levels of 0.05 and 0.01. Values of stress effects were analyzed by a two-way ANOVA test with the S. lappa was resolved and filtered (0.22 μm) before analysis. Analysis of the samples was carried out on an HPLC system at 35 °C. The detection wavelength was 230 nm and the injection volume was 20 µL. Ultrapure water (A) and acetonitrile (B) were used to prepare the mobile phase as follows: linearly varying (A) from 85% to 10% in 70 min at the flow rate of 1.0 mL/min and then held for 10 mins.To investigate the enrichment location of the allelochemicals in the environment and support the measures for dealing with the replanting obstacles, the allelochemicals were traced back and their possible release pathways were studied. Extract of plant tissue and soil of S. lappa. They caused intensive inhibition on the tested seedlings and are considered to be the main allelochemicals. Their contents in rhizosphere soil and release pathways were also determined. Activities of SOD and POD in the treated seedlings evidently respond to the stress from the allelochemicals, and ROS exhibited an accumulation trend. The increase of MDA indicated that the plasma membrane was damaged by the accumulated ROS. The results suggested that the allelochemicals stressed the receptor plant, and this caused a disequilibrium in the self-protection mechanism of the cells and damaged the cell structure. The allelochemicals affected the growth and the antioxidant enzymes of the seedlings, which would explain the obstacles to many physiological activities in the growth of the plant, and it might be an initial inducement to replanting obstacles. It also might provide a clue worth exploring to help mitigate replanting problems of S. lappa.In this study, costunolide, dehydrocostus lactone and scopoletin were obtained from the rhizosphere soil of


H-pyrazol-5-yl)(methyl)carbamoyl)benzene-diazonium hydrogen sulfate (10) was reacted with copper sulfate and sodium chloride, in the presence of ascorbic acid as reducing agent, to afford a mixture of the chlorinated epimers 4′-chloro-2,2′,5′-trimethyl-2′,4′-dihydrospiro-3-one (18) and (19), the epimers 4′-hydroxy-2,2′,5′-trimethyl-2′,4′-dihydrospiro-3-one (20) and (21), and N-benzamide (22). Under the foregoing conditions, diazonium salt 10 affords neither the 2-chloro-N--N-methylbenzamide (23) nor the tricyclic derivative 24, the classical products of the Sandmeyer and Pschorr reactions, respectively. Finally, by heating 20 at 210 °C the compound 1,3-dimethylisochromenopyrazol-5(1H)-one (24) was obtained. The transformation under the above conditions of 2-((4-chloro-3-methyl-1-phenyl-1H-pyrazol-5-yl)(methyl)carbamoyl)benzendiazonium hydrogen sulphate (11) afforded 4′,4′-dichloro-2,5′-dimethyl-2′-phenyl-2′,4′-dihydrospiro-3-one (29) as the sole reaction product.The compound 2-((1,3-dimethyl-1 This reaction was carried out with CuSO4 and NaCl, in the presence of ascorbic acid as a reducing agent isoquinoline-5(4H)-one (24), the expected product of a possible competing Pschorr ring closure.Condensation of 11 started with the reaction of the 2-nitrobenzamide derivative 25 with a seven-fold excess of nitrous acid in acetic acid, resulting in the direct introduction of a diazo group at the 4-position of the pyrazole nucleus -3-one (18) and 19, respectively in 8% overall yield; GLC-MS: two peaks, 228 (m/z) for each, (M+-Cl); IR : 1,710 (CO); 1H-NMR : 2.05 , 2.07 , 2.37 , 2.44 , 2.79 , 2.81 , 5.54 ; 5.83 ; 7.59–7.77 ; Anal. Calcd. for C13H14ClN3O (263.72) : C, 59.21%; H, 5.35%; N, 15.93%. Found: C, 59.28%; H, 5.33%; N, 15.93%.To a cold (0–5 °C) solution. (400 mL) of CuSO2SO4) and evaporated under reduced pressure. The oily residue (850 mg) was chromatographed following the flash procedure -3(2H)-one (20) as a colourless solid, mp 160–163 °C (ethyl acetate); MS (m/z): 245 (M+), 214 (M+-CH3NH2); IR: 3,263 (OH), 1,671 (CO); 1H-NMR : 1.94 , 2.34 , 2.78 , 5.11 , 6.12 , 7.52-7.70 ; Anal. Calcd. for C13H15N3O2 (245.28): C, 63.66%; H, 6.16%; N, 17.13%. Found: C, 63.42%; H, 6.55%; N, 17.02%.The mother liquors were saturated with NaCl and extracted with diethyl ether (3 × 100 mL). The combined extracts were dried (Narocedure : externa1H-NMR) with an authentic specimen of N-benzamide (22) -3-one (21), as a colorless solid, mp 177–180 °C; MS (m/z): 245 (M+), 214 (M+-CH3NH2); 1H-NMR : 1.97 , 2.22 , 2.85 , 4.92 , 5.85 , 7.43-7.70 ; 13C-NMR : 12.75 (CH3), 24.74 (CH3), 34.02 (CH3), 77.76 (CH), 91.86 (C), 111.21 (C), 122.10 (CH), 124.65 (CH), 128.83 (CH), 130.45 (CH), 131.52 (C), 139.47 (C), 152.38 (C), 166.02 (CO); Anal. Calcd. for C13H15N3O2 (245.28) : C, 63.66%; H, 6.16%; N, 17.13%. Found: C, 63.45%; H, 6.17%; N, 17.20%.The combined fractions 11–14 when evaporated gave a residue 170 mg) which was chromatographed by preparative TLC on silica gel . The plate showed three bands (U.V. light at 254 nm) among which the intermediate one underwent extraction with ethyl acetate. The extract was evaporated under vacuum and the residue obtained was crystallized to give a compound which was identical in all respects  was melted at 210 °C for 10 min. The obtained material was crystallized from ethanol to give 39, as colorless solid, in 69% yield, mp 237–239 °C; MS (m/z): 214 (M+); IR : 1715 (CO); 1H-NMR : 2.37 , 4.23 , 7.55–8.46 ; 13C-NMR : 9.96 (CH3), 40.07 (CH3), 119.86 (C), 120.47 (CH), 122.41 (C), 128.10 (CH), 128.58 (C), 132.47 (CH), 132.91 (C), 134.97 (CH), 136.95 (C), 162.08 (CO); Anal. Calcd. for C12H10N2O2 (214.22) : C, 67.28%; H, 4.71%; N, 13.08%. Found: C, 67.56%; H, 4.87%; N, 13.09%.Compound 2 by the CuSO4/NaCl/ascorbic acid reagent combination: the diazonium hydrogen sulfate 2 derived from compound 1 [10. The suspension obtained was filtered, and the solid was crystallized from ethanol (95% V/V) to give epimers 7, 8. The mother liquors when evaporated leave a residue (350 mg) which was processed by preparative TLC . Work up allowed us to obtain a product (140 mg) which was identical in all respects  to an authentic specimen of compound 4 [4 (20 mg) was obtained following the above procedure, overall yield 10%.From the transformation of the diazonium salt 25 [2 in 1.8 mL of H2O) was added dropwise and stirring was continued for a further 24 h. The obtained suspension was filtered and the filtrate was treated first with an aqueous solution (300 mL) of hydrochloric acid (0.24 M), copper sulphate pentahydrate (0.3 M) and sodium chloride (0.75 M) and then with ascorbic acid . The mixture was stirred for 1 h at r.t. and then filtered. The solid product obtained was crystallized from ethanol (95% V/V) to give 27 (yield 60%) identical in all respects  to an authentic specimen of compound 27 [This compound was prepared by modifying the procedure reported in reference . Compoun25   was added in four portions, with intervals of 20 min. between additions. After the last addition, stirring was continued at 100 °C for 1 h, then the mixture was cooled to r.t. The pH of the suspension was adjusted to 7–8 with a saturated NaHCO3 solution. The solid was separated by filtration, air dried overnight, and then extracted with boiling CHCl3 (3 × 30 mL). The combined extracts were evaporated under vacuum, leaving an residue which was crystallized from ethyl acetate to give 28 in 80% yield, mp 165–166 °C; MS (m/z): 342 (M+); IR (cm−1): 3485, 3358 (NH2), 1640 (CO); 1H-NMR (ppm): 2.26 , 3.40 ; 4.33 , 6.40–7.35 ; Anal. Calcd. for C18H17ClN4O (340.81) : C, 63.44%; H, 5.03%; N, 16.44%. Found: C, 63.35%; H, 5.21%; N, 16.56%.A suspension of Fe filings (12.41 g) in 5% (V/V) aqueous AcOH (16 mL) was heated at about 100 °C under stirring on the water bath of a rotavapor until H28  was dissolved in cooled (0–5 °C) aqueous sulfuric acid (10 M) (17.6 mL) and aqueous sodium nitrite (2.5 M) (1.82 mL) was added drop-wise to the stirred solution. The solution was stirred for a further 15 min in the ice bath and was then checked for excess nitrous acid with potassium iodide starch paper. The eventual excess can be destroyed by addition of urea.The pulverized amine 4·5H2O (0.3 M) and NaCl (0.75M), first the soln. of 11 obtained from the previous procedure, and then ascorbic acid  were added under stirring. The mixture was stirred for 1 h at r.t. and then filtered. The solid product obtained was dried in desiccator (anhydrous CaCl2) for 24 h and then crystallized from ethyl acetate to give compound 29 in 80% yield, mp 202–203 °C; MS (m/z): 359 (M+); IR (cm−1):1713 (CO); 1H-NMR : 2.38 , 2.66 , 6.70–7.96 ; 13C-NMR : 11.13 (CH3), 26.64 (CH3), 96.61 (C), 93.23 (C), 115.59 (2xCH), 122.45 (CH), 124.30 (CH), 127.21 (CH), 129.05 (2xCH), 130.98 (CH), 131.62 (CH), 132,66 (C), 139.08 (C), 142.21 (C), 147 (C), 167 (CO); Anal. Calcd. for C18H15Cl2N3O (359.06) : C, 60.01%; H, 4.20%; N, 11.66%. Found: C, 60.20%; H, 4.58%; N, 11.69%.To a cold (0–5 °C) soln. (220 mL) of CuSO29 (10 mg) in acetonitrile (1 mL) was added water (1 mL) and the mixture was refluxed for 30 min. The solution was evaporated under vacuum to give a pure residue which was identical in all respects to an authentic specimen of compound 31 [To a solution of 29 0 mg in a2 and 10 show as unique structural diversity the different substituent at the 1-position of the pyrazole nucleus, that is phenyl or methyl. The above diversity does not lead to a different chemical reactivity of these diazonium salts when they are reacted with the reagents copper sulphate/ascorbic acid/sodium chloride. In fact, the observed difference in the composition of the reaction mixtures of 10 and 2 is not due to different electronic and/or steric effects of the pyrazole substituents at 1-position, but rather to a remarkable differential solubility between the pairs of epimers 18, 19 and 7, 8. Epimers 18, 19 partially dissolve in the reaction mixture and undergo nucleophilic replacement of the chloro atom to give the hydroxy spiro derivatives 20, 21, whereas epimers 7, 8 do not follow this pathway. As regards diazonium salt 11, the presence of a substituent at the 4-position of the pyrazole moiety does not hinder this position, and a dichloro spiro derivative 29 is obtained.The diazonium salts


Atractylodes macrocephala; 1, 2--6-methyl-2, 5-cyclohexadiene-1, 4-dione; 2, 1-acetoxy-tetradeca-6E,12E-diene-8, 10-diyne-3-ol; 3, 1,3-diacetoxy-tetradeca-6E, 12E-diene-8, 10-diyne. Compounds 1–3 showed concentration-dependent inhibitory effects on production of nitric oxide (NO) and prostaglandin E2 (PGE2) in lipopolysaccharide (LPS)-activated RAW 264.7 macrophages. Western blotting and RT-PCR analyses demonstrated that compounds 1–3 suppressed the protein and mRNA levels of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). Furthermore, compounds 1–3 inhibited transcriptional activity of nuclear factor-κB (NF-κB) and nuclear translocation of NF-κB in LPS-activated RAW 264.7 cells. The most active compound among them, compound 1, could reduce the mRNA levels of pro-inflammatory cytokines  and suppress the phosphorylation of MAPK including p38, JNK, and ERK1/2. Taken together, these results suggest that compounds 1–3 from A. macrocephala can be therapeutic candidates to treat inflammatory diseases.In relation to anti-inflammatory agents from medicinal plants, we have isolated three compounds from  Atractylodes macrocephala, is a perennial herb that has been used as a traditional medicine in Korea, China, and Japan for thousands of years to treat gastrointestinal dysfunction. Examples may include loss of appetite, abdominal distention, and diarrhea -6-methyl-2,5-cyclohexadiene-1,4-dione (1), 1-acetoxy-tetradeca-6E,12E-diene-8,10-diyne-3-ol (2), and 1,3-diacetoxy-tetradeca-6E,12E-diene-8, 10-diyne (3)  A by spec. lancea  and A. mocephala , and com koreana  and A. chinensis .E)-3,7-Dimethy-2,6-octadienyl]-6-methyl-2,5-cyclohexadiene-1,4-dione (1) yellow- brownish oil, HREIMS m/z 258.1646 , 1H-NMR : δ 6.55 , 6.47 , 5.15 , 5.08 , 3.13, 2.08 , 2.06 , 1.70 ,1.62 , 1.60 ; 13C-NMR : δ 188.0 (C-1 and C-4), 148.5 (C-3), 145.9 (C-5), 140.0 (C-3′), 133.1 (C-6), 132.2 (C-2), 131.8 (C-7′), 123.9 (C-6′), 118.0 (C-2′), 39.6 (C-4′), 27.5 (C-1′), 26.4 (C-5′), 25.7 (C-9′), 17.7 (C-8′), 16.0 (C-10′), 16.0 (C-7).2-+ 303.1468 , 1H-NMR : δ 6.33 , 6.25 , 5.57 , 5.00 , 4.10 , 2.09 , 2.06 , 1.89 , 1.84  1.70 ; 13C-NMR : δ 170.9 , 170.5 , 146.5 (C-6), 143.5 (C-13), 109.8 (C-12), 109.5 (C-7), 80.0 (C-11), 79.2 (C-8), 73.1 (C-10), 72.3 (C-9), 70.3 (C-3), 60.6 (C-1), 33.0 (C-2), 33.0 (C-4), 29.1 (C-5), 21.0 , 20.9 , 18.9 (C-14).1,3-Diacetoxy-tetradeca-62 in a humidified atmosphere.RAW 264.7 murine macrophage cells  were cultured in Dulbecco’s Modified Eagle Medium (DMEM) containing 10% FBS, 100 U/mL penicillin, and 100 μg/mL streptomycin  at 37 °C in 5% CORAW 264.7 cells were stimulated with LPS (1 μg/mL) in the absence or presence of compounds for 20 h. NO released from cells was assessed by detecting nitrite in culture supernatant. Aliquots (100 μL) of culture media were incubated with 150 μL of Griess reagent  at room temperature for 10 min. Absorbance at 540 nm was read by using a microplate reader . The concentration of NO was determined by the sodium nitrite standard curve.2 levels in culture media were determined using enzyme immunoassay kit  according to the manufacturer’s instruction. In brief, 50 μL of supernatant of the culture medium and 50 μL PGE2 tracer were put into the PGE2 EIA plate and incubated for 18 h at room temperature. The wells were washed with 10 mM phosphate buffer (pH 7.4) containing 0.05% Tween 20. Then 200 μL of Ellman’s reagent was added to the well and incubated in the dark. Following the developing step, the absorbance was read at 405 nm by a microplate reader. A standard curve was prepared simultaneously with PGE2 standard ranging from 0.06 to 6 pg/mL.To observe the effects of compounds on COX-2, cells were seeded with aspirin (500 µM) to inactivate the COX-1. After 2 h, cells were washed with fresh media three times and incubated with LPS (1 µg/mL) in presence of compounds for 20 h. The PGE5 cells/60 mm dish) were treated with or without test compounds in presence of LPS (1 μg/mL) for 20 h. Cells were lysed gently with cell lysis buffer . Cell lysates were centrifuged at 4 °C and supernatant were subjected to the quantitation of protein concentrations by the Bradford method. For preparation of cytosol and nuclear extracts, cells were treated with test compounds for 30 min prior to the stimulation with 1 μg/mL LPS. Following 15 min treatment of LPS, cells were collected by using NE-PER nuclear and cytoplasmic extraction reagents according to the manufacturer’s instructions . The protein lysates were then subjected to SDS-PAGE and transferred onto PVDF membranes. After blocking with 5% non-fat milk for 1 h, membranes were incubated with the primary antibody overnight at 4 °C. Antibodies against iNOS , COX-2 , I-κB-α, p65 , ERK1/2, phospho-ERK1/2, p38, phospho-p38, JNK, phospho-JNK, and β-actin  were used for immunoblot analysis. After incubation with the secondary antibody for 1 h at room temperature, proteins were detected by VersaDoc 3000  with ECL reagents .RAW 264.7 cells (5 × 106 cells/60mm dish) were stimulated for 6 h with LPS (1 μg/mL) in the presence or absence of test compounds. Total RNA was extracted by TRIzol  according to the manufacturer’s instructions. RNA was reverse-transcribed into cDNA using reverse-transcriptase  and random-hexamer . The cDNA amplification was performed by using a recombinant Taq polymerase  and primers for iNOS, COX-2, IL-6, IL-1β, TNF-α, and β-actin. The amplified PCR products were separated on 2% agarose gels and stained with ethidium bromide.Cells  as described previously with some modifications ,37. T-RAp-value of <0.05 was considered as significantly different.All values were expressed as mean ± S.D. of three experiments. Statistical analysis was carried out with Student’s t-test. A 1–3 from Atractylodes macrocephala. Compounds 1–3 reduce NO and PGE2 production, and also suppress the iNOS and COX-2 expression in LPS-stimulated RAW 264.7 cells. The underlying mechanism proved that compounds 1–3 suppressed NF-κB through inhibiting I-κB degradation and the NF-κB nuclear accumulation. Moreover, compound 1, the most potent among three components, decreased levels of pro-inflammatory cytokines including IL-6, IL-1β, and TNF-α, and also suppressed MAPKs phosphorylation in LPS-activated RAW 264.7 cells. Taken together, compounds 1–3 from A. macrocephala can be therapeutic candidates to treat inflammatory diseases.In the present study, we explored the anti-inflammatory potential of compounds


These functional dendrimers suggest that the apparent molecular sizes of the  Dendrimers are unique macromolecules with a regular, three-dimensional, spherical structure. They have attracted much attention as functional materials because they can integrate functional groups under similar conditions on their molecular surface. In our ongoing synthetic studies of carbosilane dendrimers uniformly covered with functional molecules, we have previously reported the preparation and characterization of carbosilane dendrimers carrying mesogenic groups  and sugatrans→cis) and thermal (cis→trans) isomerization processes -hexanoates 1, 3 and 4 were synthesized in a manner similar to that described above. In contrast, the hydrosilation of 7 with HSiMeCl2, subsequent allylation and introduction of an azobenzene derivative gave 5, which had a decreased number of functional groups, compared to that of 4. terature . These deactions . Then 104 in THF solution both before and after irradiation (360 nm) were measured at room temperature  because the total length of the rans-one . To obseTrans-isomers of 3 and 4 in THF solution were injected into the GPC apparatus, and the corresponding retention times were measured. The retention times of trans3 and 4 were 12.87 and 11.97 min, respectively  were analyzed in a manner similar to that described above. The retention times of the cis isomers of 3 and 4 were 13.00 and 12.09 min, respectively, and the corresponding weight-average molecular weights were 5,800 and 12,300. However, it is difficult to conclude directly that these data show the difference in molecular sizes of the trans-isomer and the cis-isomer because there are complicated interactions arising from polarities between the gel and the azobenzene unit in the GPC column. It should be noted that the differences in retention times were reproducible. The 1~5 was then investigated using 1H-NMR experiments. The heat-isomerization process of the cis-isomers was followed by placing them in a fixed temperature NMR probe , and the cis→trans isomerization rates were calculated from the 1H-NMR signals in the aromatic region . The activation energies and the frequency factors were then calculated with Arrhenius plots, and the results are summarized in The thermal isomerization behavior of 2 and 3 are almost equal to that of 1, while the activation energy of 4 is significantly higher than that of 2 and 3. It is thought that the cis→trans thermal-isomerization is slightly prevented in the higher generation dendrimer because the azobenzene units in 4 would be in more crowded conditions and are piled up one upon another. In order to provide support for this notion, the activation energy of 5, which has a smaller number of azobenzene units than 4, was further measured. The cis→trans activation energy of 5 obtained was 119.7 KJ/mol, which is similar to that of 1, 2 and 3. The activation energies of T1 value of 1H on the azobenzene units was carried out , using a differential refractive index detector (RI-50). The analytical GPC was a Shimadzu LC-VP with a polystyrene standard . UV-spectrum measurement was performed with a JASCO V550 U/V-spectrometer. Matrix-assisted laser desorption/ionization time-of-flight mass spectra (MALDI-TOF-MS) were obtained using a Perseptive Biosystems Voyager Elite spectrometer. Melting point measurement was performed with a Büchi B-540 Melting Point apparatus.THF was freshly distilled from sodium just before use. For UV irradiation, a 400 W High-Pressure Mercury Light with a glass filter (Toshiba UV-36C) was used as the light source. Ethyl 6-[4-(4-cyanophenylazo)phenoxy]hexanoate (1). A solution of ethyl 6-bromohexanoate  in 2-butanone (EMK) (30 mL) was added to a mixture of 4-(4-hydroxyphenylazo)-benzonitrile  and K2CO3  in EMK (30 mL). The mixture was stirred at 90 °C for 24 h. After the precipitates had been filtered off, THF (50 mL) was added to the filtrate and the solution was washed with water (50 mL × 3). The extract was dried over anhydrous Na2SO4. The solvent was evaporated, and the resulting red crystals were recrystallized from EMK to give 2.48 g (85.4%) of pure 1: mp 133.0-133.6 °C; 1H-NMR (CDCl3) δ 1.2-1.3 , 1.5-1.8 , 2.3-2.4 CH2), 4.0-4.1 , 4.1-4.2 , 7.0-8.0 ; 13C-NMR (CDCl3) δ 14.2 (CH3), 24.6 (OCOCH2CH2), 25.6 (CH2CH2CH2-O), 28.7 (CH2CH2CH2-O), 34.2 (OCOCH2), 60.3 (CH3CH2), 68.0 (CH2-O-PAB), 114-133 (Ar).Tetrakis[3-{6-[4-(4-cyanophenylazo)phenoxy]hexanoyloxy}propyl]silane (2). A solution of tetrakis[3-(6-iodohexanoyloxy)propyl]silane 18 (141 mg 0.115 mmol) in EMK (30 mL) was added to a mixture of 4-(4-hydroxyphenylazo)benzonitrile (154 mg 0.690 mmol) and K2CO3 (105 mg 0.761 mmol). The mixture was stirred at 90 °C for 48 h. THF (50 mL) was added to the mixture, and the suspension was washed with water (50 mL × 3). The organic solution was dried over anhydrous Na2SO4. The solvent was evaporated and purified by preparative GPC with THF as the eluent to give 135 mg (73.4%) of 2: 1H-NMR (CDCl3) δ 0.45-0.65 , 1.20-1.80 ), 2.30-2.45 , 4.00-4.15 , 6.95-8.05; 13C-NMR (CDCl3) δ 173-114 (Ar), 68.0 (CH2), 66.7 (CH2), 34.1 (CH2), 28.8 (CH2), 25.6 (CH2), 24.6 (CH2), 23.0 (CH2), 7.86 (SiCH2).Tetrakis{3-[tris(3-{6-[4-(4-cyanophenylazo)phenoxy]hexanoyloxy}propyl)silyl]propyl}silane (3). Compound 3 was prepared in a manner similar to that described for 2: 1H-NMR (CDCl3) δ 0.50-0.70 ), 1.20-1.40 ), 1.40-1.90 ), 2.30-2.40 , 3.90-4.10 , 6.90-7.80 ; 13C-NMR (CDCl3) δ 173-114 (Ar), 118 (C=O), 113 (CN), 68.0 , 66.7 , 34.0 , 28.9 , 25.6 , 24.7 , 23.2 , 18.4 , 7.99 .Tetrakis[3-(tris{3-[tris(3-{6-[4-(4-cyanophynylazo)phenoxy]hexanoyloxy}propyl)silyl]propyl}silyl)-propyl]silane (4). Compound 4 was prepared in a manner similar to that described for 2: 1H-NMR (CDCl3) δ 0.45-0.78 ), 1.20-1.80 , 2.20-2.40 , 3.85-4.10 , 6.80-7.90 ; 13C-NMR (CDCl3) δ 173-114 (Ar), 128 (C=O), 113 (CN), 68.0 , 66.7 , 34.0 , 28.9 , 25.6 , 24.7 , 23.2 , 18.8 , 7.99 ; MALDI-TOF mass Found: m/z 14797.0. Calcd. for C840H960N108O108Si17Na: [M+Na]+, 14797.7.Tetrakis[3-(tris{3-[bis(3-{6-[4-(4-cyanophynylazo)phenoxy]hexanoyloxy}propyl)methylsilyl]propyl}-silyl)propyl]silane (5). Compound 5 was prepared in a manner similar to that described for 2. 1H-NMR (CDCl3) δ -0.10-0.10 , 0.40-0.80 ), 1.20-2.00 , 2.20-2.40 , 3.85-4.10 , 6.80-8.00 ; 13C-NMR (CDCl3) δ 173-114 (Ar), 128 (C=O), 113 (CN), 68.0 , 66.7 , 34.0 , 28.9 , 25.6 , 24.7 , 23.2 , 18.8 , 9.50 , -5.30 (SiCH3); MALDI-TOF mass Found: m/z 10436.6. Calcd. for C588H708N72O72Si17Na: [M+Na]+, 10436.8.Although various azobenzene-substituted dendrimers have been prepared in terms of their photo-responsiveness ,17,18,19


Moutabea guianensis roots led to the isolation of five polyoxygenated xanthones, including two new ones named moutabeone B  and moutabeone C , along with the three known xanthones, 1,8-dihydroxy-4,6-dimethoxyxanthone, 1,8-dihydroxy-4,5,6-trimethoxyxanthone and augustin A . Structural characterization of all compounds was established on the basis of spectroscopic methods, mainly 1D and 2D nuclear magnetic resonance (NMR) and comparison with literature data. The antioxidant activity of compounds was tested through a thin layer chromatography (TLC) bioautography assay using 1,1-diphenyl-2-picryl-hydrazyl radical (DPPH·) as detection reagent. All tested compounds were more active (DL < 0.13–0.03 µg) than Trolox (DL < 0.15 µg), used as reference standard.The phytochemical investigation of  Monnina, Bredemeyera, Securidaca and Polygala are the main sources of xanthones in the Polygalaceae + peak at m/z 363.1077  in the HRESIMS. The 1H- and 13C-NMR data of compound 5 were very similar to those reported for compound 2. The main difference is the presence of five methoxyl groups instead of four and the presence of only one hydroxyl group instead of two, indicating that compound 5 could have a methoxyl group instead a hydroxyl group at C-8. Moreover, the B ring of 5 is similar to the totally substituted ring of 1-hydroxy-3,5,6,7,8-pentamethoxyxanthone [13C-NMR data of C-4b, C-5, C-6, C-7, C-8 and C-8a were very close. Compound 5 was thus identified as 1-hydroxy-4,5,6,7,8-pentamethoxyxanthone, a new natural product, named moutabeone C. HMBC correlations observed for compound 5 are shown in Compound 1 (1.3 µg) < 5 (0.6 µg) < 3 and 4 (0.3 µg) < 2 (<0.15 µg). The detection limit of 2 was equivalent to Trolox (<1.5 µg).Thin-layer chromatography, combined with both biological and chemical detection methods, is an effective and inexpensive technique for the study of bioactive compounds . In the 1 has only two substituents in the B ring and showed less activity. However, the highest activity was observed for xanthone 2, which has four substituents in the B ring (one hydroxyl and three methoxyl groups). The increase in antioxidant activity may be attributed to the number of hydroxyl and methoxyl groups, which are strong protons and electrons donors, respectively [Xanthones have been reported to inhibit lipid peroxidation, antioxidant activity, neuroprotective properties . The xanectively ,16.1H- and 13C-, respectively) using TMS as internal standard. HRESIMS was carried out on a Waters Xevo G2-S QTof/Tof spectrometer . IR was carried out on a Shimadzu Prestige 21 . Column chromatography was performed on silica gel 60 . Precoated sheets of silica gel with UV254 indicator (thickness 200 µm) were used for TLC . Spots were visualized either with a UV lamp (254 nm) or by spraying with aqueous H2SO4 (15%) saturated with CeSO4 solution, followed by heating. Semipreparative HPLC was carried using a Varian liquid chromatograph with UV detector model ProStar 335; using a Phenomenex Gemini  C18 column .NMR spectra were recorded on a Mercury-300 NMR spectrometer  was kept in the Herbarium MG of the Museu Paraense Emílio Goeldi (MPEG). Roots were dried under forced air circulation at 40 °C for five days and powdered in a knife mill.The roots of M. guianensis (928 g) were submitted to successive extractions with hexane (3 L), ethyl acetate (3 L) and methanol (3 L) at room temperature for five days each solvent. After removal of the solvent in vacuo, the hexane, ethyl acetate and methanol extracts were obtained, respectively. The ethyl acetate extract (2.5 g) was subjected to column chromatography (CC) on silica gel, using hexane and increasing proportions of EtOAc as eluents, collecting 25 fractions of 125 mL each. The fractions 1–5 eluted with 95:5 (hexane/EtOAc) were combined according to TLC to afford fraction A (185 mg). Fraction A was purified by semi-preparative HPLC  yielding compounds 1 (8 mg), 2 (25 mg), 3 (7 mg), 4 (13 mg) and 5 (7 mg) which showed chromatographic peaks with retention times 21.57, 22.8, 28.59, 31.93 and 37.01 min, respectively.Dried and powered roots of 13C-NMR  chemical shifts of compounds 1, 3 and 4 obtained in this work: 1,8-Dihydroxy-4,6-dimethoxyxanthone (1), δC 162.8 (C1), 97.9 (C2), 167.4 (C3), 93.1 (C4), 157.7 (C4a), 154.1 (C4b), 139.9 (C5), 120.4 (C6), 109.3 (C7), 145.4 (C8), 108.1 (C8a), 184.6 (C9), 102.8 (C8b), 57.4 (4-OCH3), 55.9 (5-OCH3).1,8-Dihydroxy-4,5,6-trimethoxyxanthone (3), δC 154.3 (C1), 109.4 (C2), 121.6 (C3), 140.3 (C4), 145.7 (C4a), 149.1 (C4b), 129.2 (C5), 160.5 (C6), 95.1 (C7), 158.1 (C8), 107.8 (C8a), 184.9 (C9), 107.2 (C8b), 57.3 or 56.5 (4-OCH3), 60.9 (5-OCH3), 57.3 or 56.5 (6-OCH3).1,8-Dihydroxy-4,6,7-trimethoxyxanthone (augustin A) (4), δC 154.0 (C1), 109.2 (C2), 120.0 (C3), 139.9 (C4), 145.4 (C4a), 153.3 (C4b), 91.2 (C5), 160.7 (C6), 132.2 (C7), 153.6 (C8), 103.2 (C8a), 184.9 (C9), 107.9 (C8b), 57.3 (4-OCH3), 56.5 (6-OCH3), 60.9 (7-OCH3). · scavenging activity were observed as white or yellow bands on a purple background. The antioxidant Trolox  was used as reference standard.The antioxidant activity of xanthones was performed by TLC bioautography analysis . The comM. guianensis resulted in the isolation of five xanthones by chromatographic methods. The structures of the isolated compounds were proposed based on NMR methods. Compounds 1,8-dihydroxy-4,5,6,7-tetramethoxyxanthone (2), 1-hydroxy-4,5,6,7,8-pentamethoxyxanthone (5) were identified as new natural products. Compound 1,8-dihydroxy-4,6,7-trimethoxyxanthone (4) was isolated for the first time in the family Polygalaceae. This study shows that Moutabea genus is an important source of natural xanthones. All isolated compounds showed significant antioxidant activity in the thin layer chromatography (TLC) bioautography assay, equivalent to Trolox used as reference standard.The chemical study of the roots of


The very short sequences involve the simultaneous introduction of the side-chain and seco-B,A-ring fragments via Suzuki and Sonogashira coupling reactions. The analogues are devoid of relevant biological activity.Three analogues of 1α,25-dihydroxyvitamin D A turnisymmetry  6]..2a, 2b aS2-symmetric trans-decalin core functionalized as in 2 was dictated both by structural and synthetic considerations. Indeed symmetrical non-steroidal derivatives have been described that were found to possess vitamin D-like activity  nonane (9-BBN) in THF  is added. After stirring for 5 hours at 60 °C the reaction is brought to room temperature. To the red-orange solution is added via double-tipped needle a degassed solution of bisenol triflate 8 , 1,1’-bis(diphenylphosphino)ferrocene palladium(II) chloride (PdCl2(dppf), 9.8 mg, 0.012 mmol) and triphenylarsine  in THF, followed by a degassed 2 m sodium carbonate solution . After further stirring for 60 hours in the dark at room temperature the mixture is poured into water and extracted with methyl tert-butyl ether. After drying (magnesium sulfate) and concentration in vacuo the brown residue is purified by column chromatography  to yield 57.1 mg of 11a (57%) as an oil which is further subjected to deprotection.11a  in THF (2 mL) is added a 1 m solution of tetrabutylammonium fluoride in THF . After stirring overnight at room temperature the mixture is concentrated in vacuo. The brown residual oil is purified by repeated (2 times) column chromatography on silica gel  to yield 70 mg of analogue 2a as a white crystalline solid (70%). Rf = 0.34 ; m.p. 195 °C (decomposition); optical rotation: [α]D = −86° ; IR (KBr): 3386 (s), 3330 (s), 2932 (s), 1620 (m), 1040 (s), 972 (s), 964 (s) cm-1; 1H-NMR : δ = 6.24 , 5.83 , 5.73−5.72 , 4.43−4.38 , 4.16−4.11 , 2.56 , 2.31−2.17 , 2.03 , 1.88 , 1.82−1.76 , 1.22−1.12  ppm; 13C-NMR : δ = 141.03 (C), 137.15 (C), 131.06 (CH), 129.87 (CH), 129.81 (CH), 129.45 (CH), 126.75 (CH), 126.66 (CH), 123.41 (CH), 66.03 (CH), 64.96 (CH), 42.95 (CH), 42.89 (CH), 40.70 (CH2), 34.59 (CH2), 28.10 (CH2), 27.82 (CH2) ppm; MS: m/z (%) = 374 (4), 356 (23), 338 (75), 179 (28), 167 (57), 154 (31), 141 (82), 128 (68), 115 (61), 91 (100), 86 (31), 49 (48), 45 (46).To a solution of -1,2,4a,5,6,8a-hexahydro-4,8-di((E)-2--3,5-dihydroxycyclohex-1-enyl)ethynyl)napht-halene (2b). To a solution of bisenol triflate 8  and enyne 9  in diethylamine/dimethyl formamide  are added copper(I) iodide  and bistriphenylphosphine palladium(II) acetate . The reaction mixture is further stirred at room temperature in the dark for 3 hours. The reaction mixture is poured into water and further extracted with diethyl ether. The combined organic phases are washed with brine and dried (sodium sulfate). After concentration in vacuo the residue is purified by column chromatography on silica gel with pentane/dichloromethane (7:3) as the eluent, followed by HPLC , to yield 1.0 g of white crystalline 11b which is further subjected to deprotection.11b  in THF (3 mL) is added a 1 m solution of TBAF in THF . After stirring for 17 hours at room temperature the reaction mixture is directly eluted  on a short column of silica gel. The residue obtained after concentration in vacuo is purified by crystallization (2 times) from methanol to yield 24 mg of white crystalline 2b (51% yield). Rf = 0.59 ; m.p. 225 °C (decomposition); optical rotation: [α]D = −52° ; IR (KBr): 3360 (s), 2929 (s), 1261 (s), 1086 (s), 1048 (s), 809 (s) cm -1; 1H-NMR : δ = 6.07 , 5.94 , 4.74 , 4.64 , 4.24−4.18 , 3.95−3.90 , 2.30−2.22 , 1.97−1.90 , 1.67−1.59 , 1.28−1.23  ppm; 13C-NMR : δ = 136.09 (CH), 135.45 (CH), 123.75 (C), 119.20 (C), 90.51 (C), 87.24 (C), 63.06 (CH), 62.05 (CH), 39.47 (CH), 38.90 (CH2), 38.21 (CH2), 26.29 (CH2), 25.96 (CH2); MS (APCI): m/z = 389 .To a solution of -1,2,4a,5,6,8a-hexahydro-4,8-di((Z)-2--3,5-dihydroxycyclohex-1-enyl)vinyl)naphtha-lene (2c). To a solution of enyne 11b  in ethyl acetate is added quinoline  and Lindlar catalyst . The mixture is stirred under an atmosphere of hydrogen at room temperature and the reaction followed by TLC. After disappearance of starting material diethyl ether is added and the mixture filtered over Celite®. After concentration in vacuo the residue is purified by column chromatography on silica gel with isooctane/ethyl acetate (98:2) as the eluent and by HPLC with 0.2% methyl tert-butyl ether in isooctane as the eluent to yield 157 mg of 11c as a colorless oil (78%) which is further subjected to deprotection.11c  in THF (7 mL) is added TBAF  dropwise. After stirring for 24 hours at room temperature the reaction mixture is directly eluted  on a column of silica gel to yield 55 mg of white crystalline 2c (81%). Rf = 0.27 ; m.p. 135 °C (decomposition); optical rotation: [a]D = −41° ; IR (KBr): 3362 (s), 2916 (s), 1654 (w), 1637 (w) cm -1. 1H-NMR : δ = 5.91−5.80 , 5.65 , 5.63 , 5.45 , 5.41 , 4.60 , 4.56 , 4.53 , 4.51 , 4.22−4.19 , 3.87−3.79 , 2.38 , 2.25 , 2.18−2.10 , 1.99−1.91 , 1.86 , 1.70−1.64 , 1.58−1.51 , 1.23−1.15  ppm; 13C-NMR : δ = 138.55 (C), 138.49 (C), 134.73 (C), 134.55 (C), 131.63 (CH), 131.47 (CH), 131.05 (CH), 129.81 (CH), 129.72 (CH), 126.65 (CH), 126.28 (CH), 63.70 (CH), 62.68 (CH), 40.80 (CH), 40.72 (CH), 38.04 (CH2), 37.56 (CH2), 26.47 (CH2), 26.10 (CH2) ppm; MS: m/z (%) = 392 , 141 (28), 129 (34), 128 (31), 117 (37), 114 (37), 105 (36), 91 (100), 78 (41), 43 (75), 40 (45). To a solution of


Cellular uptakes of 99mTc-1 and 99mTc-2 were tested using a S180 cells line. Under anoxic conditions, the cellular uptakes of 99mTc-1 and 99mTc-2 were 33.7 ± 0.2% and 35.0 ± 0.7% at 4 h, whereas the normoxic uptakes of the two complexes were 6.0 ± 1.6% and 4.6 ± 0.9%, respectively. Both 99mTc-1 and 99mTc-2 displayed significant anoxic/normoxic differentials. The cellular uptakes were highly dependent on oxygen and temperature. Biodistribution studies revealed that both 99mTc-1 and 99mTc-2 showed a selective localization in tumor and slow clearance from it. At 4 h, the tumor-to-muscle ratios (T/M) were 3.79 for 99mTc-1 and 4.58 for 99mTc-2. These results suggested that 99mTc-labeled PnAO complexes 99mTc-1 and 99mTc-2 might serve as novel hypoxia markers. By introducing a second nitrotriazole redox center, the hypoxic accumulation of the marker was slightly enhanced.Two propylene amine oxime (PnAO) complexes,  However, its hypoxic cellular uptake was lower than that of [18F]FMISO, and the biodistribution studies and the PET imaging showed that the tumor to muscle ratios of [18F]3-NTR were also lower than those of [18F]FMISO FMISO .99mTc is an ideal diagnostic radionuclide with versatile chemistry -technetium (BMS181321)  (3-)-N,N',N'',N''']-technetium (BRU59-21) + 205.06960, found 205.06923.3-Chloro-3-methyl-1-butan-2-one oxime (4). Compound 3  and isoamyl nitrite  was cooled to −30 °C in an acetone-dry ice bath and vigorously stirred. Concentrated hydrochloric acid (1.4 mL) was added dropwise over 30 min. The mixture was stirred at −30 °C to −20 °C for another 2 h. The precipitated was isolated by filtration and washed with cold methanol-ether = 1:5 (10 mL) to give 4 as a white solid . 1H-NMR  δ 11.97, 8.88 , 5.33 , 1.87 2-); HRMS (ESI): calculated for C7H11ClN5O3 [M+H]+ 248.05449, found 248.05444.3-(3-(3-(Hydroxyimino)-2-methylbutan-2-ylamino)propylamino)-3-methyl-1-butan-2-one oxime (1). Compound 4 , compound 5  and (i-Pr)2NEt (100 μL) were added in dry acetonitrile (2 mL), and the mixture was stirred at room temperature for 24 h. Acetonitrile was removed on a rotary evaporator. The crude product was purified by column chromatography  and recrystallized from CH2Cl2to provide a yellow solid , m. p. 63.5–64.3 °C; 1H-NMR δ 11.32 , 10.35 , 8.79 , 5.16 , 2.23 , 1.69 , 1.40 , 1.23 2), 1.11 2); 13C-NMR  δ 161.4, 160.1, 155.7, 147.9, 56.7, 56.6, 43.1, 41.1, 31.3, 25.6, 25.4, 8.9; HRMS (ESI): calculated for C15H29N8O4 [M+H]+ 385.23063, found 385.23008.2). Compound 4 , 1,3-diaminopropane  and (i-Pr)2NEt (50 μL) were added in dry acetonitrile (2 mL), and the mixture was stirred at 40 °C for 0.5 h. Acetonitrile was removed. The crude product was purified by column chromatography  and recrystallized from CH2Cl2 to provide a white solid , m.p. 158.2–159.1 °C; 1H-NMR  δ 11.32 , 8.79 , 5.16 , 2.25 , 1.40 , 1.23 2), 0.98 . 13C-NMR  δ 161.4, 155.7, 147.8, 56.7, 43.1, 40.9, 31.3, 25.4; HRMS (ESI): calculated for C17H29N12O6 [M+H]+ 497.23275, found 497.23254. 3-(3-(3-(Hydroxyimino)-2-methyl-4-butan-2-ylamino)propylamino)-3-methyl-1-butan-2-one oxime , the ligand solution , phosphate buffer solution , saline (400 µL), diethylenetriaminepentaacetic acid (DTPA)  were mixed in a 2 mL centrifugal tube. The mixture was purged with argon and fresh SnCl2 solution  was added. The labeling mixture was stirred in ice water bath for 15 min and filtrated by 0.22 µm filter. The radio high-performance liquid chromatography (radio-HPLC) method had been described in previous work  × 100%. The trypan blue exclusion assay revealed that S180 cells maintained more than 90% viability for 4 h.The cellular accumulation experiments were performed according to the literature methods ,26. MuriTo imitate the physiological circumstances, the temperature in the accumulation experiments was 37 °C. At 1, 2 and 4 h, aliquots of supernatant were filtrated by 0.22 µm filter. Filtered supernatant was analyzed with radio-HPLC . To inve99mTc-1 and 99mTc-2 was evaluated in Kunming male mice (20–25 g) bearing the S180 tumor in the left front leg (diameter of 10–15 mm). The 99mTc-complex was (1 MBq in 0.1 mL) administered by tail vein injection. The mice were sacrificed by cervical dislocation at different times after injection. Various organs and tissues were excised, weighed and counted. The percent injected dose per gram (%ID/g) of each organ/tissue or tumor was calculated by the above data. The final results were expressed as means ± SD. of five parallel experiments.All the animal experiments were performed in accordance with the national laws related to the conduct of animal experimentation. The biodistribution of 1 and 2 containing 3-nitro-1,2,4-triazole moieties were synthesized and labeled with 99mTc at high labeling efficiencies (95%). Both 99mTc-1 and 99mTc-2 were stable at 8 h after labeling. In vitro cellular uptake studies, 99mTc-1 and 99mTc-2 displayed significant anoxic/normoxic differentials. The anoxic cellular uptakes of 99mTc-1 and 99mTc-2 reached to 33.7 ± 0.8% and 35.0 ± 0.7% at 4 h, whereas the normoxic cellular uptakes were 6.0 ± 1.6% and 4.6 ± 0.9%, respectively. The entry of oxygen restrained the cellular uptakes of 99mTc-1 and 99mTc-2, and the anoxic cellular uptakes were highly dependent on the temperature. In biodistribution studies, the tumor-to-muscle ratios (T/M) of 99mTc-1 and 99mTc-2 increased with time. At 4 h, the tumor-to-muscle ratios (T/M) were 3.79 for 99mTc-1 and 4.58 for 99mTc-2. These results suggested that nitrotriazole derivatives 99mTc-1 and 99mTc-2 might serve as novel hypoxia markers. Moreover, by introducing a second nitrotriazole redox center, 99mTc-2 showed a higher selective localization in tumor hypoxia compared with 99mTc-1.Two propylene amine oxime derivatives