The structures were elucidated by spectroscopic analyses including one- and two-dimensional NMR techniques and mass spectrometry. The bioactivities of the compounds were determined by measuring the DPPH radical scavenging activity and by carrying out antioxidant power assays (FRAP), etiolated wheat coleoptile assays and phytotoxicity assays on the standard target species Lycopersicum esculentum W. (tomato). Compounds 1 and 2 exhibited the best free radical scavenging activities, with FRS50 values of 2.0 and 1.9 µM, respectively.A bio-guided study of leaf extracts allowed the isolation of two new macrobicyclic hydrolysable tannins, namely merianin A ( Meriania Swartz (Melastomataceae) is a neotropical genus of shrubs and trees with around 93 species distributed through southern Mexico, Central America, the Greater Antilles, Andean South America, the Guaiana Highlands, and south to southeastern Brazil − and doubly charged ion at m/z 935.0801 [M − 2H]2− in negative mode. The 1H-NMR spectrum revealed the presence of three galloyl groups by three signals that integrate to two protons each at δ 7.23, 7.09 and 6.96, which were assigned as shown in S) configuration of both the HHDP and valoneoyl groups was confirmed by the circular dichroism (CD) spectrum, which exhibited a negative Cotton effect at around 253 nm and a positive effect near to 276 nm, i.e., similar to that of the lignan gomisin D, whose configuration is (S)  and 6.24 . The other glucose proton signals were all assigned based on 1H-1H shift correlation spectroscopy (COSY) and J-resolved spectra, which indicated that the conformation adopted by the two glucoses is 4C1 and that they are fully acylated. A large difference between the chemical shifts of the H-6 proton signals on Glc II (∆H6 = 1.45) suggested the presence of a biphenyl moiety with bridged ester linkages between O-4/O-6 2− in negative mode. The (S) configuration of the HHDP and valoneoyl groups was confirmed by the circular dichroism (CD) spectrum, which exhibited a negative Cotton effect at around 253 nm and positive effect close to 276 nm, as for compound 1,  and glucose II H-1 (6.24) to the ester carbonyl at δ 165.7 provided evidence for the C–O–C oxidative coupling between the galloyl moieties on C-1 and C-2 of telimagrandin II monomeric unit  and 4.34 . The other sugar proton signals were all assigned based on 1H-1H shift correlation spectroscopy (COSY), total correlated spectroscopy (TOCSY) and J-resolved spectroscopy (J-res), which indicated a sequential trans diaxial relationship between H-1’ (4.35) and H-5’ (J = 7–9 Hz) corresponding to β-glucopyranose and a sequential trans diaxial relationship between H-1” (4.34) and H-4” (J = 7–9 Hz) indicated a β-xylopyranose , 5.18  and 5.82  (β-xylopyranosyl-(1”-6’)-β-glucopyranoside (3)   indicatiside (3) . A simil50)  . The res quinone ,27. In aM. hernandoi have high values. This finding correlates with the FRS50 results. The hydrolysable tannins tested had a high antioxidant power—especially the dimeric tannins. In the FRAP assay, the activity values for phenolic compounds seem to depend on the degree of hydroxylation and the extent of conjugation of the phenolic compounds − and m/z 935.0801 [M − 2H]2−; UV maxλ (MeOH) nm log (ε) 273 (8.5), 218 (8.8); CD (MeOH) [θ] (nm): 31.1 × 104 (235), 19.5 × 104 (246), −86.5 × 103 (268), 23.7 × 103 (279), −29.4 × 103 (312). 1H-NMR  δ ppm: see 13C-NMR δ 170.7 (D-7), 170.5 (D’-7), 170.0 (C-3), 169.6 (B’-7), 169.5 (C’-7), 169.1 (E’-7’), 168.0 (A’-7), 165.9 (B-7), 165.7 , 147.6 (D-4), 146.6 , 146.4 , 146.0 (D’-4), 145.9 (D-4), 145.8 , 145.5 (E-6), 145.2 (D’-6), 145.0 (B’-6), 144.9 , 144.8 (D-6), 144.0 (E-5), 141.7 (E-4), 140.8 (E-3), 140.7 (B-4), 140.6 (A-4), 139.9 (A’-4), 137.5 (C’-5), 137.5 , 137.4 (C-5), 137.1 (D’-3), 136.9 (D-5), 136.5 (E-2), 126.2 , 126.1 , 125.9 (C’-2), 125.7 (C-2), 121.2 (A’-2), 119.8 (A-2), 119.6 (B-2), 117.3 (D-1), 116.7 (B’-1), 116.5 (C’-1), 115.3 , 114.6 (D’-1), 114.1 (E-1), 110.5 , 110.4 , 108.5 (B’-3), 108.0 (C’-3), 107.8 (D-3), 107.5 (C-3), 107.3 (D’-3), 103.5 (D-3), for sugar moieties, see O-(trimethylsilyl)-α/β-d-glucopyranoside. tR 9.883/9.590, RI 1772/1756, MS m/z (%): 73 (100), 204 (60.89/69.78), 217 (13.02/11.58). HPLC tR 5.599 min.Brown amorphous powder; −2, MeOH). ESIM: m/z 935.0765 [M − 2H]2−; UV maxλ (MeOH) nm log (ε) 271 (9.6) 218 (9.92); CD (MeOH) [θ] (nm): 41.9 × 103 (235), 25.8 × 103 (246), −12.1 × 103 (268), 4.4 × 103 (279), −4.3 × 103 (312). 13C-NMR δ 169.2 (D-7), 169.1 (D’-7), 168.6 (C-3), 168.2 (B’-7), 168.1 (C’-7), 167.7 (E’-7’), 166.6 (A-7), 164.5 (B-7), 164.3 , 146.2 (C’-4), 145.2 , 145.0 , 145.0 (A’-4), 144.6 (D-4), 144.5 (C-4), 144.4 , 144.1 (C’-6), 143.8 (D’-6), 143.6 (D-6), 143.5 , 143.4 (E’-6), 142.6 (A’-5), 140.3 (A’-4), 139.4 (E-4), 139.3 (A’-3), 139.1 (B-4), 138.5 (A-4), 136.1 , 136.0 (B’-5), 135.7 (C-5), 135.5 (C’-5), 135.1 (A’-2), 124.8 (E’-2), 124.7(C-2), 124.7 , 124.5 (D’-2), 124.3 (C’-2), 119.8 (A-1), 119.0 (A-1), 118.4 (B-2), 118.2 (E-2), 115.9 (C’-1), 115.3 (B’-1), 115.1 (E’-1), 113.9 , 113.2 (D’-1), 112.6 (A’-1), 109.1 (A’-6), 109.0 , 107.1 (D’-3), 106.6 (C-3), 106.4 (B’-3), 106.1 (E’-3), 105.8 (D-3), 102.1 (C’-3), for sugar moieties, see α/β-d-glucopyranoside. Rt 9.883/9.590, RI 1772/1756, MS m/z (%): 73 (100), 204 (60.89/69.78), 217 (13.02/11.58). HPLC tR 6.779 min Brown amorphous powder; −2, MeOH); 1H-NMR  δ ppm: 5.82 , 5.18 , 5.05 , 4.35 , 4.34 , 4.09 , 3.98 , 3.81 , 3.70 , 3.49 , 3.43 , 3.41 , 3.39 , 3.36 , 3.21 , 3.21 , 3.18 , 1.60 , 1.45 , 1.30 , 1.22 , 1.22 , 0.8 . 13C-NMR δ 140.2 (C-2), 116.1 (C-1), 102.5 , 82.1 (C-3), 77.1 (C-3’), 76.6 (C-4’), 75.8 (C-3’’), 74.3 (C-2’’), 73.8 (C-2’), 70.2 (C-4’’), 70.1 (C-5’), 68.7 (C-6’’), 65.9( C-6’), 34.9 (C-4), 32.2 (C-5), 24.9 (C-6), 22.9 (C-7), 14.1 (C-8). 1,2, 3, 4-tetrakis-O-(trimethylsilyl)- α/β-d-xylopyranoside. tR 34343/36.886, RI: 1755/1875, MS m/z (%): 73 (100).Brown syrup; 1, 2 and authentic monosaccharides , including d-(−)-arabinose, d-(−)-lyxose, d-(−)-fructose, l-(−)-fucose, l-rhamnose, d-(+)-xylose, d-(−)-ribose, d-(+)-mannose, d-(+)-galactose and d-(+)-glucose (1 mg), were heated with 2 N methanolic HCl (0.5 mL) for 3 h in a boiling water bath to give 1-O-methyl glycosides. After removal of the solvent by evaporation in vacuo, 1 mL of acetonitrile was added and free hydroxyl groups were trimethylsilylated with 100 μL of N,O-bis(trimethylsilyl)trifluoroacetamide (BSTFA) and trimethylchlorosilane (TMCS) (99:1) (Supelco) for 15 min at 70 °C.Compounds C is the absorbance of the DPPH radicals without the sample or positive control and as is the absorbance of DPPH radicals with sample or positive control. The efficient concentrations of the samples and positive controls that inhibit FRS50 were calculated and are expressed as mg/L.DPPH free radical scavenging was determined using a 96-well microplate format based on the method of Sdiri and co-workers  with sli3+ to Fe2+ by an antioxidant was determined using the method of Benzie and Strain ethanesulfonic acid (MES) and 1 M NaOH and were dissolved in DMSO to obtain working concentrations of 800, 400 and 200 mg L−1 for extracts and 1000, 333, 100, 33 and 10 µM for pure compounds. Each sample contained a constant quantity of 0.5% DMSO. The negative controls were the buffer containing DMSO without the tested compounds and the positive control was Logran®. The measurements  were made using a Fitomed© system, which allowed automatic data acquisition and statistical analysis with the associated software. Results are presented as percentage differences from the control. Zero represents control, positive values represent stimulation, and negative values represent inhibition.The extracts that exhibited activity in the general bioassay were tested in this bioassay. terature . The sam


The structures of these compounds were elucidated through extensive analysis of spectroscopic data including IR, NMR, HRESIMS, and X-ray diffraction experiments. In addition, the above new compounds were detected for their bioactivities against LPS-induced NO production in RAW 264.7 cells. Among them, compound 2 exhibited obvious anti-inflammatory activity with an IC50 value of 8.1 μM.Three new sesquiterpenoids ( Aquilaria sinensis (Lour.) Gilg (Thymelaeaceae), commonly known in different countries as Chenxiang, agarwood, agalloch, jinkoh, or aloeswood, is widely distributed in Southern China in such provinces as Hainan, Fujian, Yunnan and Guangxi + ion peak at m/z 271.1323  in the positive HRESIMS, indicating 6 degrees of unsaturation. The IR spectrum of compound 1 revealed absorptions of carbonyl (1774 cm−1), double bond (1614 cm−1), and ester group (1706 cm−1). Its 1H-NMR data  and one oxymethine  protons. The 13C-NMR data , carbonyl , and acetoxyl  signals could also be observed in the 13C NMR spectrum. In addition, there were no exchanging hydrogen atoms in compound 1, based on the molecular formula by combination with the 13C-NMR and DEPT spectroscopic data. Therefore, the presence of a tricyclic unit was determined by the distinctive signals at δC 180.7 and δC 81.4.Compound NMR data  suggesteNMR data  displaye1H-1H COSY experiment, a long spin-system of CH2CH(CH3) CHCH2CH(CHCH3)CHCH2 [C3/C4(/C14)/C5/C6/C7(/C11/C13)/C8/C9] was deduced by the correlations starting from H-3a and H-3b. Coupled with the HMBC correlations of CH3-15 to C-1, C-9, and C-10, correlations of CH3-15 to C-12, correlations of H-5 to C-1 and C-2, and correlations of H-4 to C-2, a guaiane-type sesquiterpenoid skeleton was established as shown in In the 1, the Cu Kα X-ray diffraction experiment was conducted, which confirmed the above conclusion (1 was identified to be 7βH-guaia-1(10)-en-12,8β-olide . The 1H-NMR data of compound 2 , an oxygenated methine proton , and two olefinic protons . The 13C-NMR and DEPT spectra of compound 2 exhibited 15 carbon signals, which was similar to the known compound postiaseco-guaianolide + .1: The crystallographic data were collected with a Bruker APEX2 CCD and graphite monochromated Cu Kα radiation at Bruker SAINT. The structure was solved and refined by SHELXS-97, SHELXL-97, respectively. Compound 1 was crystallized from acetone to yield colorless needles. A single crystal of dimensions 0.18 × 0.11 × 0.06 mm3 was used for the X-ray measurements. Crystal data: C15H20O3, M = 248.31, space group C121, a = 17.6380 (8) Å, b = 6.6237 (4) Å, c = 12.0602 (7) Å, V = 1328.08 (13) Å3, Z = 4, Dcalcd = 1.242 mg/m3, R1 = 0.0478, wR2 =0.1265 [I > 2σ(I)], and the absolute structure parameter 0.1 (3). CCDC 1057220 contains the supplementary crystallographic data. Copy of the data can be obtained free of charge by applying to the Director at CCDC, 12 Union Road Cambridge CB2 1EZ, UK, Fax: +44-1223336033 or E-Mail: data_request@ccdc.cam.ac.uk.X-ray Crystallographic Analysis of compound 1,10-dioxo-4αH-5αH-7βH-11αH-1,10-secoguaia-2(3)-en-12,8β-olide (2): Colorless oil; c 0.1, MeOH); IR νmax 2926, 1771, 1701, 1588, 1457, 1381, 1361, 1173, 1042, 975, 812 cm−1; 1H-NMR  and 13C-NMR  data, see m/z 287.1251 [M + Na]+ .1β-Hydroxy-4βH-5βH-7βH-11αH-8,9-secoguaia-9(10)-en-8,12-olide (3): Colorless oil; c 0.1, MeOH); IR νmax 3447, 2927, 2873, 1757, 1669, 1457, 1380, 1175, 1021, 899 cm−1; 1H-NMR  and 13C-NMR  data, see m/z 291.1594 [M + Na]+ .1α-Hydroxy-4α,10α-dimethyl-5βH-octahydro-azulen-8-one (4): Colorless oil; c 0.1, MeOH); IR νmax 3447, 2954, 2871, 1698, 1457, 1265, 1111, 1013, 978, 896 cm−1; 1H-NMR  and 13C-NMR  data, see m/z 219.1387 [M + Na]+ .2 at 37 °C and plated into 96-well plates at a density of approximately 1 × 105 cells per well. The cells were pretreated with different concentrations of the detected compounds for 2 h and then incubated for 12 h with or without LPS (2 μg/mL). The nitrite content in the culture supernatant was measured by the Griess reaction. The IC50 value was calculated by GraphPad Prism software with the inhibition rates of different concentrations. Moreover, the formula of inhibition rate is (ODModel − ODCompound)/(ODModel − ODControl) × 100%. Cell viability was measured by Cell Counting Kit-8  method.RAW 264.7 cells were maintained in Dulbecco’s Modified Eagles Medium containing 10% fetal bovine serum in a humid atmosphere of 5% CO1 was confirmed by Cu Kα X-ray crystallographic experiment. Besides, the new compound 2 showed significant inhibitory activity against LPS induced NO production in RAW 264.7 cells.A systematic chemical search was performed and resulted in the separations of three new sesquiterpenoids and one sesquiterpenoid derivative, together with three known sesquiterpenoids from Chinese eaglewood. The structures of the new compounds were identified based on detailed spectroscopic analysis and published analogues. In addition, the stereo configuration of compound


All the compounds were evaluated for their cytotoxicity against five tumor cell lines.Two new limonoids, namely 14,15-didehydroruageanin A ( Khaya A. Juss. (Meliaceae) is a genus of eight species mainly distributed in Africa and Madagascar  , signals for a ketone (δC 214.8 C), four tertiary methyls , a furan ring , and an α,β-unsaturated δ-lactone . An epoxy group was also identified by the NMR data . The above signals accounted for seven out of the 13 degrees of unsaturation and suggested the compound to thus be hexacyclic. These observations indicated that compound 1 was possibly a mexicanolide-type limonoid , which suggested that compound 2 was the 3-O-methylbutyryl derivative of seneganolide A. 1H-1H COSY correlations H-4'/H-3'/H-2' and the HMBC correlation H-3/C-1'  20.5D +9.3 ; UV (MeOH) λmax nm (log ε) 204.8 (4.23); IR (KBr) νmax (cm−1): 2972, 2931, 1732, 1634, 1460, 1384, 1271, 1197, 1114, 1028; 1H- and 13C-NMR data, see m/z 555 [M+H]+; HREIMS m/z 554.2501 M .3-O-Methylbutyrylseneganolide A (2). White amorphous powder; [α]15.4D +51.0 ; UV (MeOH) λmax nm (log ε) 203 (4.08), 282.4 (3.89); IR (KBr) νmax (cm−1): 2961, 1630, 1463, 1384, 1295, 1256, 1167, 1119, 1107, 1028, 1001; 1H- and 13C-NMR data, see m/z 553 [M+H]+; HREIMS m/z 552.2715 M .The MTT method  was used1), 3-O-methylbutyrylseneganolide A (2), and seneganolide A (3), along with five known d-seco limonoids, 1,3-dideacetylkhivorin (4), 7-deacetylkhivorin (5), 3-deacetylkhivorin (6), 1-deacetylkhivorin (7) and 3-deacetyl-7-oxokhivorin (8), were isolated from the fruits of Khaya ivorensis. Among these compounds, 1 and 2 were new compounds; and compounds 3, 4, 7, and 8 were obtained from this plant for the first time. The cytotoxicity evaluation showed that only compounds 2, 3, and 4 exhibited cytotoxicity against certain tumor cell lines.In summary, three mexicanolide-type limonoids, 14,15-didehydroruageanin A (


Ganoderma resinaceum has been used as an ethnomedicine for lowering blood sugar. To clarify the bioactive chemical constituents contributing to lower blood sugar, chemical investigation on the fruiting bodies of Ganoderma resinaceum was conducted by chromatographic techniques, and led to the isolation of 14 compounds. Their structures were elucidated as triterpenoid lactones (1–4 and 8) and ganoderma acids (5–7 and 9–14) based on the analysis of extensive spectroscopy (mass spectrometry (MS), nuclear magnetic resonance (NMR), infrared (IR), and ultraviolet (UV)) and comparison with literature data. Compounds 3, 5, 6, and 9–14 were evaluated for α-glucosidase inhibitory activity. Compounds 1–7 are new compounds. Compounds 1–4 and 8 were characteristic of an oxaspirolactone moiety, consisting of a five-membered ether ring, a five-membered lactone ring, and a characteristic C-23 spiro carbon. It is rare for natural products that such an oxaspirolactone moiety occurred in the lanostane-type triterpenoids. Compounds 5–7 and 9–14 may be important intermediates of the biosynthetic pathways of 1–4 and 8. Compounds 1 and 2 showed more potent inhibitory activity against α-glucosidase compared with the positive control drug acarbose with IC50 value of 0.75 ± 0.018 mM and 1.64 ± 0.022 mM, respectively. Ganoderma resinaceum has long been used as an ethnomedicine to treat hyperglycemia, liver diseases, and immunoregulation −  and 1D NMR data (−1), carbonyl (1767 cm−1), and α,β-unsaturated carbonyl (1635 cm−1) groups. The UV spectrum showed an absorption band to 258 nm due to the presence of α,β-unsaturated carbonyl group. The 1H NMR spectrum displayed seven methyl signals  and two oxygenated methine signals (δH 3.52 and 4.72). The 13C NMR and HSQC spectra revealed the presence of 30 carbon signals consisting of seven methyls, eight methylenes, five methines including two oxygenated methines at δC 77.8 and 72.9, six sp3 quaternary carbons , one ketone group (δC 198.6), one carboxylic group (δC 178.7), and two olefinic carbons (δC 165.2 and 139.9). The above spectroscopic data suggested 1 to be lanostane-type triterpenoid. The rings A–D were established by the heteronuclear multiple bond correlation (HMBC) and 1H–1H correlation spectroscopy (COSY) experiment (δH 1.27) and H-29 (δH 1.13) to C-3 (δC 77.8), combined with 1H–1H COSY correlations of H-3 (δH 3.52) with H-2 (δH 1.99 and 2.06), confirmed the presence of 3-OH. The α,β-unsaturated ketone system at C-8/C-9/C-11 was assigned by the HMBC correlations from H-7 (δH 2.96 and 2.76) to C-8 (δC 165.2) and C-9 (δC 139.9), H-19 (δH 1.42) and H-12 (δH 2.46) to C-9 (δC 139.9), and H-12 (δH 3.38 and 2.46) to C-11 (δC 198.6). The 15-OH was established on the basis of the HMBC correlations of H-15 (δH 4.72) with C-8 (δC 165.2), C-14 (δC 54.4), C-16 (δC 47.3), and C-30 (δC 21.8), and H-30 (δH 1.68) with C-15 (δC 72.9), as well as the COSY cross peaks of H-15 (δH 4.72) with H-16 (δH 2.52 and 2.82). Apart from seven degrees of unsaturation ascribing to the rings A–D, two carbonyls, and one double bond, the remaining two degrees of unsaturation indicated that 1 should have two rings in the side chain. According to a carboxylic carbon (δC 178.7), an acetal secondary carbon (δC 113.3), and a downfield oxygenated sp3 tertiary carbon (δC 95.6), it was assumed that the structure of 1 possessed a novel oxaspirolactone moiety consisting of a five-membered ether ring, a five-membered lactone ring, and a characteristic C-23 spiro carbon. The hypothesis of oxaspirolactone moiety was supported by the HMBC correlations from H-21 (δH 0.91) to C-17 (δC 95.6), C-20 (δC 43.7), and C-22 (δC 44.62), H-27 (δH 1.23) to C-24 (δC 44.61), C-25 (δC 35.7), and C-26 (δC 178.7), H-22 (δH 2.64 and 1.76) and H-24 (δH 2.36 and 1.96) to C-23 (δC 113.3), together with the COSY correlations of H-21 (δH 0.91) and H-22 (δH 2.64) with H-20 (δH 2.14), and H-24 (δH 2.36 and 1.96) and H-27 (δH 1.23) with H-25 (δH 3.02). The planar structure of 1 was highly similar to that of ganotropic acid (8), except for the disappearance of hydroxy group attached to C-7 in 1.Compound NMR data . Its IR periment . The key13C NMR chemical shifts of C-20, C-22, and C-24 for -isomer were almost equal, however, those of , , and -isomers showed dramatical differences larger than 5 ppm − , implying ten degrees of unsaturation. The IR and UV spectra showed the presence of hydroxy (3442 cm−1), carbonyl (1767 cm−1), and α,β-unsaturated carbonyl (1665 cm−1 and 272 nm) groups. The 1H NMR, 13C NMR , 1.01 , 1.03 , 1.05 , 1.24 , 1.25 , and 1.29 ), seven methylenes, five methines including two oxygenated methine at δC 77.3 and 72.7, six sp3 quaternary carbons including two oxygenated quaternary carbons at δC 94.7 and 112.7, one tetrasubstituted olefinic group (δC 150.3 and 154.5), and three carbonyl groups . The abovementioned data suggested 2 to be similar to 1 except for the presence of an additional ketone in 2. The HMBC correlations from H-5 (δH 1.55) and H-6 (δH 2.56 and 2.57) to C-7 (δC 205.0) confirmed that a ketone was located at C-7. The relative configuration of 2 was established based on the rotating-frame nuclear overhauser effect correlation spectroscopy (ROESY) experiment, coupling constant, and comparison of its NMR spectroscopic data with that of 1. The 3-OH was determined to be β-oriented by the ROESY correlations . The ROESY correlation of H-18 (δH 1.05) with H-15 (δH 4.45) assigned 15-OH as α-orientation. the Me-21 and Me-27 was established as α-orientation and β-orientation, respectively, based on the ROESY correlations of H-20 (δH 2.26) with H-18 (δH 1.05), H-21 (δH 1.01) with H-24α (δH 2.47), and H-24β (δH 2.03) with H-27 (δH 1.25). Therefore, compound 2 was determined to be -17,23-epoxy-3β,15α-dihydroxy-7,11-dioxo-5α-lanosta-8-en-26,23-olide.Compound  13C NMR , and HSQelations  and the 3 was isolated as a white powder. Its molecular formula was defined as C30H42O7 by the HRESIMS ion peak at m/z 513.2855 [M−H]− . The presence of hydroxy (3474 cm−1), carbonyl (1774 and 1750 cm−1), and α,β-unsaturated carbonyl (1656 cm−1 and 255 nm) groups was confirmed by the IR and UV spectra. The 1H NMR, 13C NMR , 1.04 , 1.12 , 1.18 , 1.21 , 1.29 , and 1.41 ], seven methylenes, five methines including two oxygenated methine groups δC 66.7 and 72.7, six sp3 quaternary carbons including two oxygenated quaternary carbons at δC 91.8 and 113.0, one tetrasubstituted olefinic group (δC 142.4 and 157.5), and three carbonyl groups . The 1D NMR spectroscopic data indicated the gross structure of 3 was same as that of 2. The differences were that an oxygenated methine carbon signal (δC 66.7) resonated at higher field and a ketone carbon signal (δC 216.1) at lower field in 3 compared with those of 2. The HMBC correlations from H-7 (δH 4.81) to C-8 (δC 157.5) and C-9 (δC 142.4), combined with the 1H–1H COSY correlations of H-7 (δH 4.81) with H-6 (δH 2.18 and 1.59), confirmed the presence of 7-OH. The ketone group at C-15 was supported by the HMBC correlations from H-30 (δH 1.41) and H-16 (δH 3.35 and 2.47) to C-15 (δC 216.1). The ROESY correlation of H-7 (δH 4.81) with H-30 (δH 1.41) assigned 7-OH as having β-orientation. The relative configurations of Me-21 and Me-27 were all determined to be α-orientation on the basis of the ROESY correlations of H-18 (δH 1.18) with H-20 (δH 2.38), H-22α (δH 1.93) with H-21 (δH 1.12) and H-24α (δH 1.93), and H-24α (δH 1.93) with H-27 (δH 1.29). Therefore, compound 3 was assigned as -17,23-epoxy-3β,7β-dihydroxy-11,15-dioxo-5α-lanosta-8-en-26,23-olide.Compound  13C NMR , and HSQ4 was obtained as a white powder. Its molecular formula was established as C30H40O7 by the HRESIMS ion peak at m/z 511.2700 [M−H]− . The IR and UV spectra displayed the presence of hydroxy (3490 cm−1), carbonyl (1774 and 1750 cm−1), and α,β-unsaturated carbonyl (1672 cm−1 and 265 nm) groups. A comparison of its NMR spectroscopic data with those of 3 showed their structural similarities. The only difference was the presence of ketone at C-7 in 4 instead of the hydroxy group. The ketone group was located at C-7 due to the key HMBC correlations of H-6 (δH 2.62 and 2.58) and H-5 (δH 1.63) with C-7 (δC 198.5). The relative configuration for 4 was determined based on the ROESY experiment and comparison of NMR spectroscopic data with those of 3. The ROESY correlations of H-3 (δH 3.30) with H-28 (δH 1.03), H-20 (δH 2.38) with H-18 (δH 1.03), H-22α (δH 1.91) with H-21 (δH 1.10) and H-24α (δH 2.09), and H-24α (δH 2.09) with H-27 (δH 1.28) assigned 3-OH, Me-21, and Me-27, were β-, α-, and α-orientation, respectively. Therefore, compound 4 was established as -17,23-epoxy-3β-hydroxy-7,11,15-trioxo-5α-lanosta-8-en-26,23-olide.Compound 5 was obtained as a white powder. Its molecular formula was determined to be C30H40O7 based on the HRESIMS peak ion at m/z 511.2684 [M−H]− . The 1H NMR, 13C NMR (5 was lanostane ganoderic acid closely similar to ganoderesin C − . The 1H NMR, 13C NMR (6 was lanostane-type triterpenoid closely similar to 5. The only difference was an oxygenated methine group at C-15 in 6 instead of carbonyl group in 5. The key HMBC correlations from H-30 (δH 1.17) to C-15 (δC 78.4), and H-15 (δH 4.95) to C-16 (δC 124.5) and C-30 (δC 23.2), combined with a 1H–1H COSY cross peak of H-15 (δH 4.95) with H-16 (δH 5.26), confirmed the presence of 15-OH. The 15-OH was assigned as α-orientation on the basis of the ROESY correlation of H-15 (δH 4.95) with H-30 (δH 1.17). Based on detail examination of HMBC, 1H-1H COSY, and ROESY experiments, compound 6 was identified as 3β,15α-dihydroxy-7,11,23-trioxo-5α-lanosta-8,16-dien-26-oic acid.Compound  13C NMR , and HSQ7 was obtained as a white powder. Its molecular formula was determined to be C30H44O6 by the HRESIMS ion peak at m/z 499.3048 [M−H]− . A comparison of NMR spectroscopic data  with C-6 (δC 18.7), C-8(δH 166.5), and C-9 (δH 140.9), as well as COSY correlations of H-6 (δH 1.81 and 1.51) with H-7 (δH 2.55). Therefore, compound 7 was identified as 3β,15α-dihydroxy-11,23-dioxo-5α-lanosta-8,16-dien-26-oic acid.Compound pic data  with tho8–14 were also isolated from G. resinaceum. By comparison of their NMR spectroscopic data and MS data with those reported in the literature, their structures were identified as ganotropic acid (8) − .White powder; [α]c 0.13, MeOH); UV (MeOH) λmax (log ε) 202 (3.55), 272 (3.85) nm; IR (KBr) vmax 3442, 2967, 2932, 2866, 1767, 1665, 1459, 1384, 1344, 1263, 1182, 1158, 1119, 1095, 1057, 1027, 961, 922, 882 cm−1; For 1H NMR and 13C NMR spectroscopic data, see m/z 513.2852 [M−H]− .White powder; [α]c 0.12, MeOH); UV (MeOH) λmax (log ε) 201 (3.79), 255 (4.17) nm; IR (KBr) vmax 3474, 2974, 2933, 2870, 1769, 1729, 1656, 1455, 1381, 1329, 1301, 1265, 1185, 1139, 1114, 1073, 1037, 959, 916, 882 cm−1; For 1H NMR and 13C NMR spectroscopic data, see m/z 513.2855 [M−H]− .White powder; [α]c 0.11, MeOH); UV (MeOH) λmax (log ε) 265 (3.79) nm; IR (KBr) vmax 3490, 3040, 2976, 2932, 2867, 1774, 1750, 1672, 1455, 1382, 1328, 1258, 1188, 1141, 1075, 1031, 960, 919, 883 cm−1; For 1H NMR and 13C NMR spectroscopic data, see m/z 511.2700 [M−H]−1 .White powder; [α]c 0.23, MeOH); UV (MeOH) λmax (log ε) 234 (4.02), 271 (3.80) nm; IR (KBr) vmax 3432, 2968, 2933, 2872, 1718, 1677, 1601, 1459, 1381, 1189 cm−1; For 1H and 13C NMR data, see m/z 511.2684 [M−H]− , 1023.5451 [2M−H]− .White powder; [α]c 0.23, MeOH); UV (MeOH) λmax (log ε) 234 (4.05), 272 (3.82) nm; IR (KBr) vmax 3424, 2970, 2938, 2875, 1709, 1686, 1664, 1461, 1406 cm−1; For 1H and 13C NMR data, see m/z 513.2843 [M−H]− , 1027.5767 [2M−H]− .White powder; [α]c 0.18, MeOH); UV (MeOH) λmax (log ε) 203 (3.77), 259 (3.77) nm; IR (KBr) vmax 3430, 2963, 2932, 2878, 1715, 1653, 1577, 1460, 1375 cm−1; For 1H and 13C NMR data, see m/z 499.3048 [M−H]− , 999.6182 [2M−H]− .White powder; [α]α-Glucosidase inhibitory activity was examined by the method described by Li et al. [α-glucosidase inhibitor, was used as the positive control drug.i et al. . AcarbosG. resinaceum led to the isolation of 14 triterpenoids, including seven new triterpenoids (1–7) and seven known analogues (8–14) from the fruiting bodies of G. resinaceum. Compounds 1–4 and 8 were determined to be triterpenoid lactones which possessed an oxaspirolactone moiety in the side chain, consisting of a five-membered ether ring, a five-membered lactone ring, and a characteristic C-23 spiro carbon. It is rare that such an oxaspirolactone moiety ring occurred in the lanostane-type triterpenoids. Based on the analysis of plausible biogenetic pathway, compounds 5–7 and 9–14 may be important intermediates in the biosynthesis of 1–4 and 8. Compounds 1 and 2 showed stronger α-glucosidase inhibitory activity than the positive control drug acarbose, which are strong α-glucosidase inhibitors with IC50 value of 0.75 ± 0.018 mM and 1.64 ± 0.022 mM, respectively.Phytochemical investigation of


Baccharis genus have been shown to possess important biomedical properties, including cytotoxic activity. In this study, we examined the cytotoxic effect of methanol extract from Baccharis obtusifolia (Asteraceae) in cancer cell lines of prostate (PC-3), colon (RKO), astrocytoma (D-384), and breast (MCF-7). The methanolic extract displayed the largest substantial cytotoxic effect in lines of colon cancer (RKO) and cerebral astrocytoma (D-384). Chromatographic purification of the B. obtusifolia methanolic extract led to the isolation and identification of 5,4′-dihydroxy-7-methoxyflavone (1) and 5-hydroxy-7,4′-dimethoxyflavone (2) compounds of the flavonoid type.Some species of the Baccharis is the largest genus in the family Asteraceae, with over 500 species distributed throughout North and South America -3--2-(4-sulfo-phenyl)-2H-tetrazolium inner salt) cell viability assay was used to assess the inhibitory effects of the extracts on the survival of human cancer cell lines. A total of 3–5 × 10 standard error of the mean (SEM) of three independent experiments. The nonlinear regression was determined by GraphPad Prism 5 .All of the data were reported as means ±1), and the fraction F 43-45 eluted in Hex:EtOAc 90:10, from which we isolated 11.2 mg of a yellow crystalline solid identified as 5-hydroxy-7,4′-dimethoxyflavone (2)  (vone (2) . All of 1): m.p. 240–243°C, EIMS m/z (%) 286 , 269 (36), 241 (16), 167 (13), 166 (14), 138 (14), 128 (16), 121 (7), 118 (10). 1H NMR (DMSO-d6) δ ppm: 2.8 , 3.1 , 3.89 , 6.40 , 6.79 , 6.85 , 6.96 , 7.98 , 12.95  13C NMR (DMSO-d6) δ ppm: 79 (C-2) 43.1 (C-3) 196.0 (C-4) 164.1 (C-5) 95.1 (C-6) 168.0 (C-7) 94.2 (C-8) 162.8 (C-9) 103.5 (C-10) 130.5 (C-1ʼ) 128.0 (C-2ʼ) 114.2 (C-3ʼ) 156.1 (C-4ʼ) 115.6 (C-5ʼ) 128.0 (C-6ʼ) 55.6 (OCH3-7).Physical and spectroscopic constants from 5,4′-dihydroxy-7-methoxyflavone (2): m.p. 160–163°C. EIMS m/z (%) 298 , 269 (28), 255 (14), 166 (11), 138 (16), 135 (10), 132 (16). 1H NMR (CDCI3) δ ppm: 3.89, 3.90 , 6.37 , 6.49 , 6.58 , 7.02 , 7.85 , 12.80 . 13C NMR (CDCl3) δ ppm: 164.0 (C-2) 104.3 (C-3) 182.4 (C-4) 165.4 (C-5) 98.0 (C-6) 165.4 (C-7) 92.6 (C-8) 162.1 (C-9) 104.3 (C-10) 123.5 (C-1ʼ) 128.0 (C- 2ʼ) 114.4 (C-3ʼ) 157.6 (C-4ʼ) 114.5 (C-5ʼ) 128.3 (C-6ʼ) 55.6 (OCH3 7ʼ) 55.7 (OCH3-4ʼ).Physical and spectroscopic constants from 5-hydroxy-7,4′-dimethoxyflavone ( B. obtusifolia (50 μg/ml). The extract exhibited a strong inhibitory effect on RKO 89.2%, as in D-384 cells with 60.6%. However, the MCF-7 and PC3 cell lines exhibited moderate inhibitory activity with 20.2% and 18.3%, respectively  and 34.30 μM (10.22 μg/ml), for compounds 1 and 2, respectively. Compounds 1 and 2 were specific for the RKO line, and compound 2 was more effective than compound 1 (Compoundsmpound 1 .1) and 5-hydroxy-7,4′-dimethoxyflavone (2). These compounds have been identified for the first time in Baccharis obtusifolia, but they have been reported in other species: compound 1 in Chromolaena subscandens [ Trixis vauthieri [ Trigona spinipes [ 2 in Baccharis polycephala [ Thymus vulgaris [ Combretum erythrophyllum [ Kaempferia parviflora [ Boesenbergia pandurata [The species belonging to the Asteraceae family are characterized by their abundant biological activity , which rscandens , Trixis authieri , and Trispinipes  and compvulgaris , Combretophyllum , Kaempferviflora , and Boeandurata .Our evaluation of the biological activity of methanolic extract revealed a high percentage of inhibition in RKO cell lines (89.2%), followed by D-384 cells (60.6%). This activity can be attributed to the presence of flavonoids in the extract. Some flavonoids act on these stages in very different cell types, which may lead to the use of these compounds as cytostatic agents in the later stages of carcinogenesis rather than as early-stage preventive elements , 21.50 of flavonoids 1 and 2 shows similar values to the other compounds of the same type [ICame type –24. Numeame type . The act6-C3-C6, which may form a third ring. Natural flavonoids are often present in at least three phenolic hydroxyls [1H data from compounds 1 and 2 show a substitution pattern “para” in the ring B [ 1, the substituent corresponds to a hydroxyl group (OH), and for compound 2, the substituent corresponds to a methoxy group (OCH3).Chemically, flavones are substances of phenolic nature and are characterized by having two benzene aromatic rings united by a bridge with three carbon atoms. They have the general structure Cydroxyls . The NMRe ring B ; substit13C from compound 1 in the aliphatic zone δ (55.6 ppm), there appears a signal that belongs to a methoxy group. The signal in δ (43.1 ppm) is assigned to C-3 [1H NMR spectrum. The spectral data of compound 2 show two signals in δ (55.6–55.7 ppm) belonging to two methoxy groups present in this structure, and these two groups are also observed in the 1H NMR spectrum.In the NMR data of d to C-3  and the  in vitro via various mechanisms in the oncogenic process, which renders them possible useful agents in the early stages of cancer or in the inhibition of later stages of progression or invasion. This activity may be explained by studying the Structure-Activity Relationship (SAR) between several flavonoids and cancer [ breast cancer resistance protein (BCRP) [2) possesses important structural characteristics like the double bond between C-2 and C-3 and the presence of an OH group at the 5-position and the carbonyl group at the 4-position. Previous studies have suggested that the position, number, and substitution of hydroxyl groups in rings A and B, as well as the saturation in the C2-C3 bond, may be important factors that increase the cytotoxic or antiproliferative activities of flavonoids [1) does not exhibit the double bond at C-2 and C-3, which in mono- or dihydroxylated flavones results in the loss of cellular growth inhibitory activity [Flavonoids have characteristics that make them attractive for anticancer research. They actd cancer . An examn (BCRP) . The stravonoids , which wactivity . It woul


Malus, Pyrus, Sorbus, and Chaenomeles. Here, cell suspension cultures of Sorbus pohuashanensis were established to study biphenyl phytoalexins formation after yeast extract (YE) treatment. An ultra-performance liquid chromatography (UPLC) method coupled with quadrupole time of flight mass spectrometry (Q-TOF-MS) LC−MS/MS was applied to determine the time course of these biphenyl biomarkers accumulation in YE-treated S. pohuashanensis suspension cells. The results of quantitative analyses show the content of Noraucuparin, 2′-Hydroxyaucuparin, and their glycosides initially increased, then decreased over time. The Noraucuparin content reached its highest (225.76 μg·g−1) at 18 h after treatment, 6 hours earlier than that of Noraucuparin 5-O-β-d-glucopyranoside. The content of 2′-Hydroxyaucuparin reached its highest (422.75 μg·g−1) at 30 h after treatment, also earlier than that of its glycoside. The understanding of phytoalexin metabolism in this study may provide a basis for improving Maloideae resistance to pathogens.Biphenyls are unique phytoalexins de novo synthesized in plants in response to pathogen attack. These compounds are found in Maloideae, a subfamily of the Rosaceae. The anti-microbial activities of biphenyls have been reported in a number of studies and they appear to represent an important defense strategy against pathogens common in the Maloideae, such as species in  Malus domestica) and pear (Pyrus communis), could de novo synthesize phytoalexins after infection by bacteria and fungi. Even so, there is significant economic loss because yield can be greatly affected by these types of pathogenic infections  > 0.8—a total of 18 marker compounds accountable for the different metabolite profiles of control and treatment groups were obtained  was subsequently used. The Robtained .m/z 215.0706 [M − H]−, m/z 245.0814 [M − H]−, m/z 377.1229 [M − H]− and m/z 407.1331 [M − H]− with predicted molecular formulae of C13H11O3−, C14H13O4−, C19H11O8− and C20H23O9−, respectively. These metabolites could be used as potential markers in different groups of time course of metabolites differentially accumulated in YE-treated S. pohuashanensis suspension cells. The makers may be attributed to differences in the activity of key enzymes, and differences in the regulation of metabolic pathways. Furthermore, we separated these compounds from the methanol extract of YE-treated cell cultures of S. pohuashanensis by guided LC-MS, and identification structures of these target compounds based on NMR : Yellow amorphous powder (MeOH); UV (MeOH) λmax (log ε) 279 (5.5), 226 (4.6) nm; ESI-MS (negative) m/z 407 [M − H]−, 245[M − H − 162]−; HR-ESI-MS (negative) m/z 407.1331 [M − H]− ; 1H-NMR : 7.18–7.21 , 6.97–7.01 , 6.67 , 4.89 , 3.20–3.75 , 3.87 ; 13C-NMR : δ 131.1 (C-1), 106.7 , 148.8 , 135.9 (C-4), 131.2 (C-1′), 153.8 (C-2′), 116.5 (C-3′), 128.1 (C-4′), 120.7 (C-5′), 130.3 (C-6′), 56.2 , 101.7 (glc-C-1), 73.7 (glc-C-2), 77.8 (glc-C-3), 70.2 (glc-C-4), 76.3 (glc-C-5), 61.2 (glc-C-6). The spectroscopic properties were in accordance with published data −, 215[M − H − 162]−; HR-ESI-MS (negative) m/z 377.1229 [M − H]− ; 1H-NMR : δ 7.52 , 7.45 , 7.26 , 6.87 , 6.65 , 4.44 , 3.93 , 3.85 , 3.69 , 3.50–3.53 , 3.18–3.39 ; 13C-NMR : δ 131.7 (C-1), 102.1 (C-2), 146.5 (C-3), 132.3 (C-4), 149.7 (C-5), 107.5 (C-6), 141.3 (C-1′), 126.9 , 128.5 , 126.4 (C-4′), 56.1 (3-OCH3), 101.7 (glc-C-1), 74.7 (glc-C-2), 78.2 (glc-C-3), 71.5 (glc-C-4), 78.1 (glc-C-5), 62.7 (glc-C-6). The spectroscopic properties were in accordance with published data − ; 1H-NMR : δ 7.22–7.27 , 6.96–6.99 , 6.66 , 3.91 ; 13C-NMR : δ 128.2 (C-1), 105.7 , 147.6 , 134.5 (C-4), 129.0 (C-1′), 152.5 (C-2′), 115.6 (C-3′), 127.9 (C-4′), 120.7 (C-5′), 130.0 (C-6′), 56.4 . The spectroscopic properties were in accordance with published data − ; 1H-NMR : δ 7.53 , 7.40 , 7.30 , 6.86 , 6.70 , 3.93 ; 13C-NMR : δ 132.0 (C-1), 102.3 (C-2), 147.1 (C-3), 133.6 (C-4), 144.1 (C-5), 107.8 (C-6), 141.1 (C-1′), 126.8 , 128.7 , 126.9 (C-4′), 56.2 (3-OCH3). The spectroscopic properties were in accordance with published data .Raw profile data (raw) from UPLC-Q-TOF-MS were imported into the Progenesis QI software for removal of isotope peak, alignment and peak-picking. This software approach employs peak alignment, matching, and comparison. After peak extraction and grouping, nonlinear retention time correction of peaks was accomplished in two iterative cycles with descending bandwidth (bw). The resulting data matrix consisting of retention time, S. pohuashanensis guided by LC-MS. YE-treated cell cultures of S. pohuashanensis (400.0 g) were extracted three times using the same process described in section 4.6. The filtered extract was combined and condensed at below 40 °C to remove most of methanol. The residue was suspended in H2O (about 400 mL) and then successively partitioned with petroleum ether (PE), EtOAc, and BuOH. The EtOAc extract was freeze-dried and yielded 10.5 g of yellow powder.Based on biomarkers screening from 4.8, the compounds of biphenyl and biphenyl glycosides were isolated from YE-treated cell cultures of 2Cl2-MeOH  to afford fractions I-V. The Fraction I was further chromatographed on silica gel 60 with a stepwise gradient of CH2Cl2–acetone  to obtain 6 fractions (Fractions 1.1→1.6). Fr.1.3 was subjected to be repeated CC over silica gel 60  to afford 3 (6 mg) and 4 (8 mg). Preparative HPLC was performed on Shimadzu LC-10A instrument equipped with a SPD-M20A detector  using YMC-Pack ODS-A columns . Fraction IV was chromatographed over silica gel 60 eluting with CHCl3–MeOH–EtOAc–H2O  to afford Fraction IVa, Fraction V with CHCl3–MeOH–EtOAc–H2O  to afford Fraction Va. They were further purified by preparative HPLC using acetonitrile and water as the mobile phase (flow rate 8.0 mL·min−1) and the eluate was monitored at 276 nm. Compound 2 (3 mg) was obtained from Fraction IVa with gradient elution (23%–25% acetonitrile from 25–35 min), and Compound 1 (10 mg) was obtained from Fraction Va by using 21.5% acetonitrile.The yellow powder (10.5 g) was subjected to column chromatography over silica gel and eluted with CH18 column  thermostated at 40 °C. The binary gradient consisted of solvent system A (formic acid/water 0.1:99.9 v/v) and solvent system B (formic acid/acetonitrile 0.1:99.9 v/v). The conditions were as follows: 0 min, 10% B; 4 min, 40% B; 5 min, 52% B. The injection volume for all samples was 1 μL and the flow rate was 0.6 mL·min−1.Chromatographic analyses were performed on an Acquity UPLC I-Class system (Waters). UPLC separation was carried out on Acquity UPLC BEH CTandem mass spectrometry was performed on the QTRAP 6500 system  for characterization equipped with electrospray ionization (ESI) source (AB SCIEX). MS analysis was carried out in negative ionization mode (ESI) by monitoring the protonated molecular ions at the following operating conditions: ion source voltage of −4500 V; entrance potential (EP) of −10 V; nebulizer gas (Gas 1) of 55 psi; heater gas (Gas 2) of 55 psi; curtain gas of 30 psi and turbo spray temperature (TEM) of 550 °C. MS parameters were manually optimized as shown in O-β-d-glucopyranoside were detected at 6 h after YE treatment. These showed that de novo biosynthesis of biphenyl phytoalexins was rapidly induced by YE in S. pohuashanensis cell cultures. Through analyzing the changes in the four main phytoalexins in terms of different contents with treatment time, we found that Noraucuparin could transfer to Noraucuparin 5-O-β-d-glucopyranoside by glycosylation, while 2′-Hydroxyaucuparin could also transfer to a corresponding glucoside, indicating that Noraucuparin and 2′-Hydroxyaucuparin could transfer not only to their downstream mediators but also to their glucosides via glycosyltransferase.Herein, Noraucuparin and Noraucuparin 5-


V. luteola, and among these, three new (1–3) were characterized by spectroscopic and mass spectrometric analyses. The isolated constituents were assessed for anti-inflammatory potential evaluation, and several purified principles exhibited significant superoxide anion and elastase inhibitory effects.Seventy-three compounds were identified from the methanol extract of  Plenty of phytochemicals isolated from dietary and medicinal plants, such as epigallocatechin gallate , soy isoInflammation is the first response of the immune system to infection or irritation due to bacteria, virus, wound, or other various environmental factors resulting in injuring. Recently, the overexpression of neutrophils has been demonstrated to be related with various human diseases and causing serious threat to human health ,13,14,15V. luteola (Jacq.) Benth. is a trailing or twining herb belonging to the Vigna genus usually distributed in the tropical regions. In Taiwan, V. luteola is grown in the open seaside at elevation below 100 m and spread over the island -β-d-galactopyranoside (56), chrysoeriol (57), astragalin (58), kaempferol-3-O-sophoroside (59), kaempferol 3-O-sophoroside-7-O-rhamnoside (60), robinin (61), isorhamnetin 3-O-[α-l-rhamnopyranosyl(1→6)]-β-d-galactopyranoside 7-O-α-l-rhamnopyranoside (62); eight isoflavonoids, daidzein (63), 8-O-methylretusin (64), 5,7,4′-trihydroxyisoflavone (65), 7,2′,4′-trihydroxyisoflavone (66), tectorigenin (67), afromosin (68), 3′-methoxydaidzein (69), 7,4′-dihydroxy-8-methoxyisoflavone (70); three others, 2-furanoic acid (71), maltol glucoside (72), sodium 5-hydroxymethylfuran-2-carboxylate (73), respectively, were verified by inspection of their reported spectroscopic data  analysis. The infrared (IR) absorption bands at 3416 and 1647 cm−1 were in agreement with the presences of hydroxy and carbon–carbon double bond functionalities, respectively. In the 1H-NMR spectrum, there were proton signals for two methyl singlets at δ 0.96 , 1.06 , one methyl doublet of doublets 1.73 , two oxymethylene protons at δ 3.61  and 3.81 , two oxymethine at δ 3.80  and 4.18 , three methines at δ 1.36 , 1.51 , 2.44 , and two trans olefinic protons at δ 5.67  and 5.78 . The 13C-NMR and distortionless enhancement by polarization transfer (DEPT) spectra exhibited three methyl carbons at δ 18.2 (C-10), 24.0 (C-12) and 25.9 (C-11); three methylene carbons at δ 31.6 (C-4), 46.8 (C-2), and 69.7 (C-13); five methines at δ 43.7 (C-5), 68.5 (C-3), 84.4 (C-7), 129.6 (C-8), and 132.1 (C-9); and two quaternary carbons at δ 37.7 (C-1) and 81.2 (C-6). The correlation spectroscopy (COSY) indicated the linkage of H-2/H-3/H-4/H-5/H-13 and H-7/H-8/H-9/H-10  heteronuclear multiple bond correlation (HMBC) spectrum  established its relative stereochemistry configuration . The UV absorption maxima at 274 and 221 nm were indication of the heteroaromatic chromophore  was equal to 0.8 ppm in the 10 ppm sample. After acidification of 4 with 0.1 M HCl, the spectral data was changed to be the same as that of (−)-phaseic acid Vigna luteola (Jacq.) Benth. was gathered in the side of Ching-Shui River, Nantou, Taiwan (March 2006), and it was authenticated by Prof. C. S. Kuoh . A voucher specimen (PCKuo_2006002) was deposited in the herbarium of School of Pharmacy, National Cheng Kung University, Tainan, Taiwan.The plant material V. luteola (dried weight: 3.5 kg) were grounded and extracted with methanol (20 L) exhaustively under reflux (85 °C) for 8 hours, and the resulting liquid was concentrated in vacuo to give a dark brown syrup (640 g). The methanol extract was partitioned between chloroform and water to produce chloroform soluble layer (190 g) and water soluble layer (450 g), respectively.The herbs of n-hexane and acetone (100:1 to 1:1) to afford seven fractions (CF 1–7), as monitored by TLC. CF 3 was further column chromatographed on silica gel with a mixture of n-hexane and ethyl acetate (step gradient from 50:1 to 1:1) to afford fourteen subfractions (CF 3-1–3-14). CF 3-2 was purified by silica gel column chromatography (SiO2 CC) and the minor fraction was recrystallized from chloroform to give 18 (0.3 g). CF 3-3 was isolated by SiO2 CC to yield 16 (0.8 mg), 17 (1.1 mg), 40 (1.8 mg), a mixture of 19 and 20 (2.5 mg), and a mixture of 23 and 24 (2.2 mg), respectively. CF 3-5 was separated by SiO2 CC and further preparative thin-layer chromatography (pTLC) purification afforded compounds 6 (1.6 mg), 7 (4.3 mg), 37 (4.2 mg), 69 (0.6 mg), 8 (5.4 mg), 14 (1.2 mg), 55 (1.8 mg), and 63 (2.0 mg), respectively. CF 6 was further subjected on a silica gel column and eluted with a chloroform and methanol mixture (step gradient from 50:1 to 1:1) to produce thirteen subfractions (CF 6-1–6-13). A mixture of 21 and 22 (4.3 mg) was obtained from CF 6-7 by recrystallization of ethyl acetate. CF 6-8 was purified by SiO2 CC and the minor fraction was further isolated by pTLC with a solvent mixture of chloroform and acetone (10:1) to obtain 41 (1.0 mg). CF 7 was isolated on a SiO2 CC eluted with chloroform and methanol (step gradient from 50:1 to 1:1) to give twelve subfractions (CF 7-1–7-12). CF 7-3, 7-4, 7-5, and 7-8 were purified by SiO2 CC and the resulting minor fractions were subjected to pTLC to yield 68 (2.3 mg), 25 (1.9 mg) and 64 (1.5 mg), 10 (1.6 mg) and 43 (5.1 mg), and 36 (2.2 mg), respectively.The chloroform layer was subjected to a silica gel column eluted with a step gradient of 26 (3.1 mg) and 38 (1.5 mg). WF 1-4 was separated by SiO2 CC and further purified by pTLC to afford 65 (2.2 mg), 67 (6.0 mg), 35 (24.0 mg), and 54 (2.7 mg). WF 3 was purified by Sephadex LH-20 CC eluted with a step gradient mixture of water and methanol  to produce thirteen subfractions (WF 3-1–3-13). WF 3-4 and 3-9 were separated by pTLC to yield 70 (1.3 mg), 48 (2.4 mg), and 56 (7.3 mg). WF 3-10 was purified by SiO2 CC eluted with a step gradient mixture of chloroform and methanol (50:1 to 1:1) to produce eight minor fractions (WF 3-10-1–3-10-8). WF 3-10-2 was isolated by reversed-phase HPLC with a Gemini 5u C18 column  eluted with a MeOH-H2O mixture  to yield 61 (2.7 mg) and 62 (3.5 mg). WF 5 was isolated by Diaion HP-20 CC to give six subfractions (WF 5-1 ~ 5-6). WF 5-4 was subjected on a Sephadex LH-20 column eluted with a mixture of water and methanol , and then recrystallization of the resulting minor fractions afforded 66 (2.2 mg), 71 (1.1 mg), 4 (3.7 mg), 39 (1.6 mg), 44 (1.8 mg), and 57 (5.0 mg), respectively. WF 5-5 was purified by reversed-phase HPLC with a Gemini 5u C18 column  eluted with a MeOH-H2O mixture  to afford 52 (4.2 mg). WF 6 was subjected to Diaion HP-20 CC eluted with water and a step gradient of methanol (10:0 to 0:10) to afford six subfractions (WF 6-1–6-6). WF 6-2 and 6-6 were further purified by SiO2 CC and pTLC to obtain 5 (4.0 mg), 12 (2.6 mg), 49 (3.3 mg), 15 (3.5 mg), and 58 (0.4 g). WF 7 was purified by Diaion HP-20 CC eluted with water and a step gradient of methanol (10:0 to 0:10) to obtain seven subfractions (WF 7-1–7-7). WF 7-6 was separated by repeated Sephadex LH-20 CC eluted with a step gradient mixture of water and methanol (10:0 to 0:10) to yield 1 (2.8 mg), 9 (17.0 mg), 11 (5.2 mg), 47 (2.6 g), 59 (1.5 g), and 60 (0.5 g). WF 8 was purified by SiO2 CC eluted by chloroform and a step gradient with methanol and water (100:1:0.1 to 1:1:0.1) to obtain seven subfractions (WF 8-1–8-7). WF 8-4 was isolated by SiO2 CC and pTLC to give 45 (2.7 mg), 13 (1.1 mg), and 72 (1.9 mg). WF 10 was subjected to Sephadex LH-20 CC eluted with water and a step gradient of methanol (10:0 to 0:10) to obtain ten subfractions (WF 10-1–10-10). WF 10-3, 10-6, and 10-9 were further purified by pTLC to yield 42 (0.8 mg); 31 (4.2 mg), 46 (12.3 mg), and 50 (11.0 mg), and 51 (1.3 mg) and 53 (2.2 mg), respectively. WF 12 was subjected to Diaion HP-20 CC eluted with a mixture of water and methanol (step gradient from 10:0 to 0:10) to afford seven subfractions (WF 12-1–12-7). WF 12-3 was purified by Sephadex LH-20 CC and further resolved on pTLC to produce 30 (1.9 mg), 34 (2.1 mg), and 73 (1.4 mg). WF 12-6 was isolated by repeated SiO2 CC yield 2 (4.2 mg). WF 13 was resolved on a Sephadex LH-20 column eluted with water and methanol (step gradient from 10:0 to 0:10) to produce eight subfractions (WF 13-1–13-8). WF 13-3 was separated by SiO2 CC and pTLC to obtain 32 (1.6 mg) and 33 (40.0 mg). WF 13-7 was resolved on Sephadex LH-20 CC and the minor fraction was isolated by repeated SiO2 CC to result in 3 (6.3 mg), 27 (33.0 mg), and 28 (43.3 mg). WF 14 was subjected to SiO2 CC eluted with ethyl acetate and methanol (step gradient from 300:1 to 1:1) to give nine subfractions (WF 14-1–14-9). Compound 29 (7.5 mg) was obtained from the WF 14-7 by repeated SiO2 CC followed by recrystallization.The water soluble layer was resolved on a Diaion HP-20 column and eluted with a step gradient mixture of water and methanol  to result in sixteen fractions (WF 1–16). WF 1 was subjected to Diaion HP-20 CC eluted with the same program as mentioned above to obtain nine subfractions (WF 1-1–1-9). WF 1-3 was purified by pTLC with a solvent mixture of chloroform and methanol (50:1) to give D25 + 36.9 ; IR (neat) νmax: 3416, 2928, 1647, 1580, 1383, 1045 cm−1; ESI-MS (rel. int. %) m/z 227 , 209 (43); HR-ESI-MS m/z 249.1460 ([M + Na]+) ; 1H-NMR  δ 5.78 , 5.67 , 4.18 , 3.81 , 3.80 , 3.61 , 2.44 , 1.73 , 1.73 , 1.51 , 1.36 , 1.31 , 1.06 , 0.96 ; 13C-NMR  δ 132.1 (C-9), 129.6 (C-8), 84.4 (C-7), 81.2 (C-6), 69.7 (C-13), 68.5 (C-3), 46.8 (C-2), 43.7 (C-5), 37.7 (C-1), 31.6 (C-4), 25.9 (C-11), 24.0 (C-12), 18.2 (C-10).Colorless powder; [α]D25 − 23.6 ; UV (MeOH) λmax (log ε): 221 (3.78), 274 (3.38) nm; IR (neat) νmax: 3430, 2924, 1637 cm−1; ESI-MS (rel. int. %) m/z 272 ; HR-ESI-MS m/z 294.0952 ([M + Na]+) ; 1H-NMR  δ 8.03 , 7.32 , 7.21 , 4.78 , 4.65 , 4.51 , 3.85 , 3.67 , 3.60 , 3.38 , 1.25 ; 13C-NMR  δ 155.9 (C-3), 148.6 (C-6), 138.1 (C-2), 125.0 (C-4), 124.8 (C-5), 101.4 (C-1’), 74.0 (C-4’), 72.4 (C-3’), 72.3 (C-2’), 70.3 (C-7), 70.1 (C-5’), 18.0 (C-6’).Yellowish amorphous powder; [α]max (log ε): 299 (3.44) nm; IR (neat) νmax: 3493, 2953, 2133, 1703, 1631, 1387, 1054 cm−1; ESI-MS (rel. int. %) m/z 197 ; HR-ESI-MS m/z 197.0449 ([M − H]−) ; 1H-NMR  δ 6.97 , 6.25 , 3.60 , 2.60 , 1.84 ; 13C-NMR  δ 165.8 (C-10), 164.8 (C-2), 162.3 (C-4), 156.4 (C-6), 114.8 (C-3), 110.4 (C-5), 61.7 (C-9), 32.6 (C-7), 32.1 (C-8).Colorless syrup; UV (MeOH) λD25 − 9.1 ; UV (MeOH) λmax (log ε): 249 (3.55) nm; IR (neat) νmax: 3424, 2937, 1713, 1644, 1636, 1550, 1401, 1334, 1254 cm−1; HR-ESI-MS m/z 279.1234 ([M − Na]−) ; 1H-NMR  δ 7.87 , 6.20 , 5.86 , 3.93 , 3.65 , 2.81 , 2.71 , 2.45 , 2.37 , 1.95 , 1.22 , 1.00 ; 13C-NMR  δ 211.3 (C-3), 175.0 (C-11), 141.6 (C-9), 134.1 (C-8), 129.2 (C-7), 127.9 (C-10), 87.8 (C-5), 82.8 (C-6), 78.5 (C-12), 54.0 (C-4), 53.2 (C-2), 49.0 , 20.5 (C-15), 19.4 (C-14), 15.8 (C-13).Colorless powder; [α]max (log ε): 282 (3.83) nm; IR (neat) νmax: 3403, 2941, 1550, 1414, 1389, 1250, 1085 cm−1; ESI-MS (rel. int. %) m/z 163 ; HR-ESI-MS m/z 163.0396 ([M − Na]−) ; 1H-NMR  δ 7.36 , 7.33 , 6.75 , 6.33 ; 13C-NMR  δ 176.4 (C-9), 160.0 (C-4), 141.0 (C-7), 130.1 (C-2 and -6), 128.6 (C-1), 123.2 (C-8), 116.6 (C-3 and -5).Colorless powder; UV (MeOH) λThe assays of the generation of superoxide anion and elastase release inhibition examinations were determined as described previously . The exp


The structure and mechanistic pathways for all products were discussed and proved based on spectral results, in addition to conformational studies. Our aim after the synthesis is to investigate their antimicrobial activity against various types of bacteria and fungi species. Preceeding such an investigation, a molecular docking study was carried out with selected conformers, as representative examples, against three pathogen-proteins. This preliminary stage could support the biological application. The potency of these compounds as antimicrobial agents has been evaluated. The results showed that derivatives which have di- and trithiazole rings displayed high activity that exceeds the used standard antibiotic.Based on the extensive biological activities of thiazole derivatives against different types of diseases, we are interested in the effective part of many natural compounds, so we synthesized a new series of compounds containing di-, tri- and tetrathiazole moieties. The formation of such derivatives proceeded via reaction of 2-bromo-1-(4-methyl-2-(methylamino)thiazol-5-yl)ethan-1-one with heterocyclic amines,  It is not coincidental that the first effective antibiotics used to treat microbes , which was prepared through bromination of 5-acetyl-4-methyl-2-(methylamino)thiazole (1) in acidic medium, as previously reported [2 was reacted with a mole equivalent of 2-aminothiazole (3) in ethanol under reflux to give one of two possible isomers 6A or 6B as indicated by the presence of only one spot in TLC. The spectral data  and elemental analysis cannot distinguish the actual product among the suggested possibilities. 1H-NMR of the isolated product showed a remarkable singlet at δ = 8.20 ppm for the imidazole proton. The high value of such a chemical shift indicated that this =CH is adjacent to a sp3 hybridized nitrogen atom [6A rather than isomer 6B. As illustrated in I, followed by in-situ dehydrative cyclization to form the desired fused heterocycle 6A [2O [4) reacted in a similar way with compound 2 to afford 5--N,4-dimethylthiazol-2-amine (7A). The other possible isomer 7B -N,4-dimethylthiazol-2-amine (8A) was formed via reaction of 2-aminothiophenol with 2-bromo-1-(4-methyl-2-(methylamino)thiazol-5-yl)ethan-1-one (2) in ethanol under reflux . Furthermore, the IR spectra for all derivatives 6–8 were free from any absorption bands characteristic for C=O, but, revealed the presence of two characteristic bands for NH and C=N at 3417-3238 and 1634-1612 cm−1, respectively. The starting material required in this study was 2-bromo-1-(4-methyl-2-(methylamino)thiazol-5-yl)ethan-1-one , displayed significant bands at, ύ = 3430 (NH) and 1653 (C=O), which suggested the presence of 17A tautomer in the solid state, while, the 1H-NMR spectrum suggested the existence of 17C tautomer, through the appearance of a singlet signal for the CH2 group at δ = 4.44 ppm and lack of from any signals for a NH group (17b (R = CH3) exists in only the tautomeric form 17A as proved from its spectral data . The formation of products 17a,b started with substitution reactions through the formation of intermediate 16, which was followed by concurrent elimination of a water molecule , reflect the comparative stability that is consistent with the suggested conformers (A compounds). With respect to tautomers 3A and 3B, we considered the reaction-determining step of 3A tautomer. Also, regarding the two tautomers, the charge on the N(6) atom was −0.51307 or −0.338355, respectively. The two values indicate the highly nucleophilic nature of the nitrogen atom in compound 3A. This agrees excellently with the previously suggested reaction pathway. Another additional proof was obtained from their electrostatic maps (3A (−6.935 × 10−2) but not the 3B tautomer (−5.868 × 10−2). Such an observation suggests the superiority of 3A tautomer over the 3B one. Regarding intermediate 16, the one in the reaction pathway in −2) over the whole molecule. Also, the charge on N(18) (which is adjacent to the carbonyl group) was −0.782807, while the charge on N (14) was −0.580283, which points to the high nucleophilicity of the N(18) atom, which was already proposed in the reaction mechanism. All synthesized conformers as well as tautomers, were structurally investigated , to confenergies  point totic maps , which eA conformers, while, the LUMO mainly appeared condensed in a defined part in the compound. This is somewhat logical, that the two orbitals consider the counterparts in the compounds. Essential physical parameters were calculated for all optimized conformers, to gain some insight into their general characteristics. Electronegativity (χ), chemical potential (μ), global hardness (η), global softness (S), global electrophilicity index (ω) and absolute softness (σ), were the calculated parameters (ω), reflect significant toxicity for most of the tested compounds, especially of 19, 21, 24 and 25. (ii) The global hardness (η) and absolute softness (σ) values, clarify the high degree of softness for compounds 13, 19, 21 and 24 [HOMO and LUMO frontier images were extracted and displayed . The HOMrameters . The dif1 and 24 ,30,31,3214, 17a, 19f and 24 conformers and 3k4p, 1ydo and 1ecl PDB co-crystals of Aspergillus niger, Bacillis subtilis and Escherichia coli, respectively [This study aimed to provide a good simulation of what happens inside infected cells after treatment with the suggested inhibitors. Implementing the MOE module, the docking process was executed between ectively . Fundame17a, 24 and 19f against the 1ydo protein of Bacillis subtilis bacteria [The underlying mechanism of action was suggested by all docking processes, which varied from the best interaction to others with moderate binding, based on the following remarks;. (i) The energy value scores (S) appear in the −5.8156 to −7.0064 and −4.3294 to −5.3916 ranges, which represent the best to moderate interactions, respectively. (ii) The ligation mode was mainly focused on, S, N, 5-ring and 6-ring sites. (iii) The main binding receptor-backbones were lysine, asparagine, glutamate, alanine, serine and tryptophan. (iv) The interactions appeared to covering all possible types, such as π-H, π-π, H-donor and H-acceptor. (v) From all the previous remarks the best inhibition may be expected for bacteria ,35. The 2 and the seventeen synthetic derivatives 6–8, 12–14, 17a,b, 19a, 19c–f, 21a,b, 24, and 25 against two fungal species, and four Gram-positive as well as four Gram-negative bacteria, as illustrated in The antimicrobial activity was assayed for the starting α-bromoketone derivative 2 showed less activity than the the used antifungal and antibacterial standards against most of the tested fungi and bacteria. The activity of the thiazole ring was affected by the conversion of the α-bromoketone group to heterocyclic rings at position 5. The presence of an imidazotetrazole ring at position-5 of the thiazole ring, enhanced the activity of compound 12 (IZ = 23.3 µg/mL) to a level similar to that of amphotercin B against Aspergillus niger, whereas, upon insertion of imidazotriazole, thiazolopyrimidine, thiazole rings in the same position-5, a favorable activity was recorded for compounds 14, 17a, 19a, 19c–f, 21a and 24 that exhibited higher activity than the standard drug toward Aspergillus niger (Staphylococcus aureus), three derivatives 14, 19a and 21a are more potent than the reference drug (ampicillin), but only derivative 14 exceed the activity of the standard against Staphylococcus epidermidis. Observing Gram negative bacteria, six derivatives 17a,b, 19a,e,f, 24 are more potent than the reference drug, as depicted in Streptococcus pyogenes and Gram-negative Pseudomonas aeruginosa. It is easy to note that the starting α-bromoketone derivative us niger . Investi7, 13, 17a,b, 19c, and 21b that showed significant inhibition zones. The results revealed that most of these compounds displayed promising activity against Aspergillus niger, whereas, only compound 17a showed MIC similar to Amphotericin B against Geotricum candidum. Focusing on the results of G (+) bacteria and G (−) bacteria, the most active derivative is 17a. Moreover, compounds 7 and 13 showed high reactivity toward the bacterium Salmonella typhimurium (MIC = 0.49 μg/mL). Finally, from the inhibition zone and MIC investigations of the tested compounds, we found that the insertion of a heterocyclic system at position-5 of the thiazole ring, improved the antimicrobial activity. Also, with the presence of more than one thiazole ring in the system, promising activity concluded to be more than that of the reference antibiotics used was observed. −1, JASCO, Easton, MD 21601, United States). 1H and 13C-NMR spectral analysis were obtained on a Bruker 400 MHz instrument . Mass spectral analysis was obtained at 70 eV on Shimadzu GCMS-QP 1000 EX mass spectrometer  at a heating rate 40 °C/min. The elemental analysis was carried out using a 2400 CHN Elemental Analyzer  instrument. The biological evaluation of the products was carried out at the Regional Center of Mycology and Biotechnology, Al-Azhar University, Cairo, Egypt. Theoretical calculations were performed using Gaussian 09 and AutoDock Tools 4.2. 2-Bromo-1-(4-methyl-2-(methylamino)thiazol-5-yl)ethan-1-one (2) was synthesized as previously reported [All melting points were measured on a Electrothermal IA 9000 series digital melting point apparatus . IR spectral analyses were recorded on a FT-IR-4100 spectrophotometer (400–4000 cmreported .2-bromo-1-(4-methyl-2-(methylamino)thiazol-5-yl)ethan-1-one  and heterocyclic amines 3, 4, 9–11 or 2-aminothioplenol 5 (5 mmol) in ethanol (30 mL) was refluxed for 10 h (the progress of the reactions was monitored by TLC). After the reaction was complete, the reaction mixture was filtered, the filtrate washed with methanol, and crystallized from a suitable solvent to give derivatives 6, 7, 12–14 and 8, respectively. A mixture of equimolar quantities of max 3417 (NH), 3038 (sp2 C-H), 2997 (sp3 C-H), 1634 (C=N) cm−1; 1H-NMR (DMSO-d6) δ 2.45 , 3.56 , 7.37 , 7.94 , 8.20 , 9.60 ; MS m/z (%) 250 , 239 (53), 234 (47), 224 (89), 220 (16), 206 (62), 191 (45), 188 (57), 183 (100), 150 (30), 119 (34), 91 (33), 65 (42), 48 (54). Anal. Calcd. for C10H10N4S2 (250.34): C, 47.98; H, 4.03; N, 22.38. Found: C, 48.05; H, 3.89; N, 22.24%.Yellow solid, (78% yield), mp 243–244 °C; (EtOH); IR (KBr) νmax 3238 (NH), 3047 (sp2 C-H), 2987 (sp3 C-H), 1623 (C=N) cm−1; 1H-NMR (DMSO-d6) δ 2.59 , 3.56 , 7.40 , 7.54 , 8.01 , 8.09 , 8.76 , 9.62 ; MS m/z (%) 300 , 285 (22), 270 (29), 254 (40), 175 (26), 153 (27), 147 (23), 150 (28), 138 (71), 129 (29), 77 (17). Anal. Calcd. for C14H12N4S2 (300.40): C, 55.98; H, 4.03; N, 18.65. Found: C, 55.87; H, 3.87; N, 18.58%.Yellow solid, (83% yield), mp 265–266 °C; (EtOH); IR (KBr) νmax 3374, 3293 (2NH), 3059 (sp2 C-H), 2926 (sp3 C-H), 1612 (C=N) cm−1; 1H-NMR (DMSO-d6) δ 2.51 , 2.89 , 5.45 , 6.42 , 6.73 , 7.01 , 7.07 , 7.4 , 9.07 ; MS m/z (%) 275 , 272 (100), 256 (7), 166 (10), 154 (3), 152 (17), 147 (4), 139 (86), 127 (7), 113 (37), 102 (23), 91 (31), 83 (72), 76 (17). Anal. Calcd. for C13H13N3S2 (275.39): C, 56.70; H, 4.76; N, 15.26. Found: C, 56.85; H, 4.58; N, 15.17%.Yellow solid, (88% yield), mp 110–112 °C; (EtOH/Dioxane); IR (KBr) νmax 3428, 3203 (2NH), 3106, 2924 (sp3 C-H), 1646, 1599 (C=N) cm−1; 1H-NMR (DMSO-d6) δ 2.51 , 2.89 , 6.92 , 7.03 , 8.85 ; MS m/z (%) 235 , 220 (16), 210 (55), 192 (24), 191 (33), 189 (38), 170 (23), 123 (32), 106 (24), 103 (26), 96 (34), 82 (20), 64 (100). Anal. Calcd. for C8H9N7S (235.27): C, 40.84; H, 3.86; N, 41.68. Found: C, 41.10; H, 3.65; N, 41.59%.Yellow solid, (76% yield), mp 155–157 °C; (EtOH); IR (KBr) νmax 3438, 3165 (2NH), 3094 (sp2 C-H), 2991 (sp3 C-H), 1649 (C=N) cm−1; 1H-NMR (DMSO-d6) δ 2.25 , 2.85 , 7.12 , 7.24 , 8.0–8.21 , 8.29 , 10.44 , 10.58 ; MS m/z (%) 283 , 264 (41), 245 (56), 238 (17), 230 (49), 207 (100), 181 (66), 167 (41), 137 (83), 124 (22), 115 (42), 98 (62), 67 (45). Anal. Calcd. for C14H13N5S (283.35): C, 59.34; H, 4.62; N, 24.72. Found: C, 59.17; H, 4.55; N, 24.60%.Yellow solid, (65% yield), mp 140–142 °C; (EtOH/Dioxane); IR (KBr) νmax 3436, 3124(2NH), 2978 (sp3 C-H), 1629 (C=N) cm−1; 1H-NMR (DMSO-d6) δ 2.41 , 3.03 , 6.75 , 6.85 , 9.55 ; MS m/z (%) 234 , 224 (7), 221 (15), 187 (5), 166 (12), 153 (5), 145 (25), 127 (14), 118 (28), 107 (4), 84 (11), 73 (28), 67 (6), 65 (54), 63 (100). Anal. Calcd. for C9H10N6S (234.28): C, 46.14; H, 4.30; N, 35.87. Found: C, 46.25; H, 4.29; N, 35.75%.Yellow crystals, (91% yield), mp 245–146 °C; (EtOH); IR (KBr) ν15a or 15b or thiosemicarbazone derivatives 18a–f, 20a,b 22, or 23 (5 mmole) in dioxane (30 mL) was added 2-bromo-1-(4-methyl-2-(methylamino)thiazol-5-yl)ethan-1-one  and triethylamine (0.7 mL) then the reaction mixture was refluxed for 6 h. After cooling, the formed solid product was collected by filtration and washed with aqueous ethanol to dissolve any triethylamine hydrochloride crystals, dried and crystallized from ethanol/dioxane to give thiazolopyrimidines 17a,b or hydrazine-thiazole derivatives 19a–f, 21a,b, 24 and 25.To a stirred hot solution of pyrimidinethione derivatives max 3430 (NH), 2926 (sp3 C-H), 1653 (C=O), 1564 (C=N) cm−1; 1H-NMR (DMSO-d6) δ 2.51 , 3.30 , 4.44 , 6.69 , 7.36–7.95 ; 13C-NMR (DMSO-d6) δ 18.9 (CH3), 30.1 (CH2), 34.0 (N-CH3), 110.9, 112.0, 114.3, 119.8, 125.5, 127.32, 129.6, 131.9, 132.6, 145.7, 147.6, 166.7 (C=O) ppm; MS m/z (%) 354 , 338 (28), 331 (40), 327 (30), 311 (41), 269 (44), 258 (69), 201 (30), 173 (43), 155 (56), 111 (83), 86 (47), 60 (100). Anal. Calcd. for C17H14N4OS2 (354.45): C, 57.61; H, 3.98; N, 15.81. Found: C, 57.73; H, 3.76; N, 15.72%.Yellow solid, (81% yield), mp 200–201 °C; IR (KBr) νmax 3430 (NH), 2928 (sp3 C-H), 1636 (C=O), 1560 (C=N) cm−1; 1H-NMR (DMSO-d6) δ 2.07 , 2.51 , 3.37 , 5.68 , 12.24 . MS m/z (%) 292 , 284 (59), 277 (23), 265 (21), 256 (63), 246 (32), 231 (25), 202 (44), 163 (25), 139 (16), 127 (40), 108 (53), 87 (35), 82 (25). Anal. Calcd. for C12H12N4OS2 (292.38): C, 49.30; H, 4.14; N, 19.16. Found: C, 49.45; H, 4.05; N, 19.02%.Dark orange solid, (92% yield), mp 259–260 °C; IR (KBr) νmax br. 3419 (2NH), 2929 (sp3 C-H), 1554 (C=N) cm−1; 1H-NMR (DMSO-d6) δ 2.31 , 2.51 , 2.89 , 7.39–8.27 , 10.22 ; MS m/z (%) 343 , 334 (14), 303 (17), 282 (30), 218 (20), 163 (10), 127 (100), 93 (61), 91 (49), 85 (12). Anal. Calcd. for C16H17N5S2 (343.47): C, 55.95; H, 4.99; N, 20.39. Found: C, 56.14; H, 4.80; N, 20.27%.Green solid, (77% yield), mp 235–237 °C; IR (KBr) νmax br. 3423 (2NH), 2925 (sp3 C-H),1552 (C=N) cm−1; 1H-NMR (DMSO-d6) δ 2.29 , 2.33 , 3.37 , 7.42–7.98 , 10.26 ; MS m/z (%) 379 , 378 , 377 , 364 (28), 346 (25), 337 (34), 328 (100), 265 (50), 251 (49), 239 (43), 225 (22), 216 (30), 188 (49), 128 (52), 83 (53). Anal. Calcd. for C16H16ClN5S2 (377.91): C, 50.85; H, 4.27; N, 18.53. Found: C, 51.02; H, 4.13; N, 18.44%.Green solid, (68% yield), mp 226–228 °C; IR (KBr) νmax br. 3416 (2NH), 2919 (sp3 C-H), 1552 cm-1; 1H-NMR (DMSO-d6) δ 2.28 , 2.33 , 3.36 , 7.56–7.92 , 8.31 , 10.26 ; MS m/z (%) 422 , 406 (52), 390 (100), 371 (56), 347 (32), 325 (37), 302 (38), 270 (67), 238 (36), 111 (24), 72 (20). Anal. Calcd. for C16H16BrN5S2 (422.36): C, 45.50; H, 3.82; N, 16.58. Found: C, 45.75; H, 3.68; N, 16.46%.Green solid, (64% yield), mp 220–222 °C; IR (KBr) νmax 3423, 3249 (2NH), 3161, 2936 (sp3 C-H), 1602, 1560 (C=N) cm−1; 1H-NMR (DMSO-d6) δ 2.28 , 2.51 , 3.51 , 7.17–8.00 , 8.24 , 10.19 ; MS m/z (%) 361 , 338 (22), 299 (66), 273 (45), 271 (100), 252 (16), 202 (16), 189 (15), 136 (39). Anal. Calcd. for C16H16FN5S2 (361.46): C, 53.17; H, 4.46; N, 19.38. Found: C, 53.24; H, 4.31; N, 19.29%.Green solid, (72% yield), mp 249–250 °C; IR (KBr) νmax br. 3411 (2NH), 2919 (sp3 C-H), 1552 cm−1; 1H-NMR (DMSO-d6) δ 2.25 , 2.35 , 2.50 , 3.26 , 6.94 , 7.07 , 7.76 , 8.50 , 10.21 ; MS m/z (%) 357 , 346 (53), 324 (100), 298 (60), 270 (45), 229 (34), 174 (68), 145 (44), 135 (81), 115 (50), 65 (56). Anal. Calcd. for C17H19N5S2 (357.49): C, 57.12; H, 5.36; N, 19.59. Found: C, 57.30; H, 5.33; N, 19.46%.Green solid, (68% yield), mp 229–230 °C; IR (KBr) νmax 3386, 3265 (2NH), 2926 (sp3 C-H), 1597 (C=N) cm−1; 1H-NMR (DMSO-d6) δ 2.00 , 2.51 , 2.80 , 3.86 , 7.30 , 7.44 , 7.57 , 8.29 , 10.39 ; MS m/z (%) 373 , 369 (52), 346 (100), 332 (55), 284 (24), 270 (63), 250 (34), 177 (36), 123 (37), 111 (63), 76 (38). Anal. Calcd. for C17H19N5OS2 (373.49): C, 54.67; H, 5.13; N, 18.75. Found: C, 54.76; H, 5.24; N, 18.65%.Green solid, (75% yield), mp 195–197 °C; IR (KBr) νmax br. 3423 (3NH), 2927 (sp3 C-H), 1590 (C=N) cm−1; 1H-NMR (DMSO-d6) δ 2.34 , 2.39 , 2.51 , 3.35 , 6.69–8.08 , 9.39 , 10.91 ; MS m/z (%) 456 , 420 (24), 367 (24), 267 (26), 249 (52), 200 (44), 139 (33), 104 (21), 85 (60), 76 (29). Anal. Calcd. for C19H20N8S3 (456.61): C, 49.98; H, 4.42; N, 24.54. Found: C, 50.11; H, 4.27; N, 24.35%.Green solid, (68% yield), mp 270–271 °C; IR (KBr) νmax br. 3425 (3NH), 2991 (sp3 C-H), 1621 (C=N) cm−1; 1H-NMR (DMSO-d6) δ 1.92 , 2.41 , 2.68 , 7.50–8.11 , 8.66 , 10.92 ; MS m/z (%) 479 , 477 , 461 (41), 382 (33), 313 (15), 222 (26), 202 (36), 111 (15), 108 (32), 96 (100). Anal. Calcd. for C18H17ClN8S3 (477.02): C, 45.32; H, 3.59; N, 23.49. Found: C, 45.43; H, 3.41; N, 23.28%.Green solid, (65% yield), mp 278–280 °C; IR (KBr) νmax 3419, 3262 (3NH), 1650 (C=O), 1600 (C=N) cm−1; 1H-NMR (DMSO-d6) δ 2.51 , 3.39 , 6.92 , 7.07 , 7.33 , 7.65 , 8.68 , 9.03 , 11.21 , 12.48 ; MS m/z (%) 370 , 356 (50), 340 (32), 244 (45), 337 (50), 305 (40), 297 (63), 282 (25), 264 (58), 224 (32), 211 (44), 190 (100), 87 (44), 82 (19), 77 (32). Anal. Calcd. for C16H14N6OS2 (370.45): C, 51.88; H, 3.81; N, 22.69. Found: C, 52.02; H, 3.65; N, 22.53%.Brown solid, (79% yield), mp 210–212 °C; IR (KBr) νmax 3415, 3240 (2NH), 2937 (sp3 C-H), 1575 (C=N) cm−1; 1H-NMR (DMSO-d6) δ 2.51 , 3.38 , 3.83 , 7.51–8.03 , 8.13, 8.40 , 10.02 ; MS m/z (%) 593 , 567 (14), 548 (23), 531 (20), 510 (14), 439 (20), 404 (20), 360 (100), 352 (15), 276 (31), 226 (11), 211 (13), 200 (58). Anal. Calcd. for C25H24N10S4 (592.78): C, 50.66; H, 4.08; N, 23.63. Found: C, 50.76; H, 4.14; N, 23.58%.Brown solid, (72% yield), mp 216–217 °C; IR (KBr) ν2 and the new thiazole compounds 6–8, 12–14, 17a,b, 19a, 19c–f, 21a,b, 24 and 25 were assayed in vitro for their antimicrobial activity against two fungal species, namely Aspergillus niger, Geotricum candidum, four Gram-positive bacteria: Staphylococcus aureus, Staphylococcus epidermidis, Bacillis subtilis, Streptococcus pyogenes, and four Gram-negative bacteria: Pseudomonas aeruginosa, Escherichia coli, Klebsiella pneumoniae, Salmonella typhimurium using the known the agar diffusion procedure [The starting α-bromoketone derivative rocedure . FirstlyThe synthesis pathways were also confirmed based on structural configuration studies which were performed using the Gaussian 09 software . To deteAspergillus niger, Bacillis subtilis and Escherichia coli, which cover fungi and bacteria, were selected, which will simulate the intended screening in vitro study. 3k4p, 1ydo and 1ecl, were the co-crystalline structures as PDB files, which are attributed to the reported microorganisms, respectively. This introductory study may strengthen the experimental screening, which will be displayed later. Firstly the docked synthesis must be configured by the addition of hydrogen atoms, after removing water molecules around the skeletons. Then the potential energy must be fixed after displaying the atomic charges and other important parameters adapted by the MMFF94x force field. After that, the oriented compounds must be saved as MDB format, to be ready for the docking process. The PDB files of co-crystallized proteins are configured by addition of hydrogen, choosing the receptor and generating the α-site spheres in the site finder [Through the MOE module (v. 2015), the molecular docking process was implemented forselected compounds against three microorganisms. Regarding the tested inhibitors, the selection aimd to cover all the various synthesis pathways, to give a broad view about their interactions with pathogen proteins. Regarding the selected pathogens, e finder . The docA main and growing health problem around the world is microbial resistance to existing treatments. Several studies have been conducted aiming to synthesize thiazole derivatives with excellent antimicrobial activity. In this context we focused on the synthesis for a new series of thiazole derivatives with varying substituents at position-5. The atomic assembly and the mechanistic pathway for all products have been investigated and proved based on our spectral and conformational studies. Antimicrobial results for the starting material (α-bromoketone) and the products indicated that the antimicrobial activity of some new synthetic products is more efficient than that of the starting material itself. This activity of the derivatives was somewhat improved to exceed the activity of standard drugs.