These compounds showed approximately equal extinction coefficients of optical absorption over 400–550 nm that corresponds to near-IR two-photon based excitation wavelengths at 780–1,100 nm. Accordingly, they may be utilized as potential precursor candidates to the active-core structures of photosensitizing nanodrugs for 2γ-PDT in the biological optical window of 800–1,050 nm.Broadband nonlinear optical (NLO) organic nanostructures exhibiting both ultrafast photoresponse and a large cross-section of two-photon absorption throughout a wide NIR spectrum may make them suitable for use as nonlinear biophotonic materials. We report here the synthesis and characterization of two C Highly strained curving regions of the cage surface consist of chemically reactive six fulvalenyl bridging olefins that can be utilized for making nucleophilic addition reactions. Chemical modification of C60 on only a limit number of functionalization sites may not lead to much alternation of the cage’s photophysical properties. Conversely, nucleophilic addition of one or two light-harvesting antenna chromophores will largely enhance the cage’s ability to respond and perform various photoinduced electronic and energy-related events by acting as an electron-acceptor fullerene is more readily available commercially in up to kilogram quantities than a number of higher fullerenes. However, its visible absorption extinction coefficient is rather low. This limitation can be overcome by attaching highly fluorescent chromophores as light-harvesting antenna units, such as porphyrin ,21 or dielengths ,13,14. Themistry ,23. In t1), C60(>DPAF-C18) fullerene triads and tetrads was based on both linear and nonlinear optical characteristics of 9,9-dioctadecyl-2-diphenylaminofluorenyl-61-carbonylmethano[60]fullerene (PAF-C18) , and 9,9PAF-C2M) , to cons0–150 fs  that madt manner . Upon tha 1γ-PDT . The obs0–500 nm . The beheviously . By exten and CPAF-Cn in combination as hybrid chromophore addends to a single C60 cage should result in the formation of new methano[60]fullerene triads, C60(>DPAF-C18)(>CPAF-C2M) 3, and tetrads, C60(>DPAF-C18)(>CPAF-C2M)24, as shown in 3 and 4 will then be capable of performing dual-band 2γ-PDT-based photoinduced biocidal effects with enhanced penetration depth at 800–1,100 nm. Synthetically, preparation of 3 and 4 was accomplished by the synthesis of a structurally well-defined monoadduct 1, followed by the attachment of one or two CPAF-C2M antenna in sequence. A key intermediate precursor, 7-α-bromoacetyl-9,9-dioctadecyl-2-diphenylaminofluorene  was prepared by a three-step reaction involving first palladium catalyzed diphenylamination of commercially available 2-bromofluorene at the C2 position of the fluorene ring to afford DPAF 5  in 97% yield. Friedel-Crafts acylation of DPAF-C18 with α-bromoacetyl bromide and AlCl3 in CH2Cl-CH2Cl at 0 °C for a period of 4.0 h afforded the compound 8 in a yield of 96%. Addition reaction of 8 to C60 was carried out in the presence of 1,8-diazabicyclo[5.4.0]undec-7-ene  at ambient temperature for 4.0 h to result in C60(>DPAF-C18) 1 in 65% yield  after column chromatographic purification.Accordingly, selective attachment of these two antenna moiety types DPAF-Cd DPAF 5 . It was 2 by replacing two octadecyl groups with 2-methoxyethyl groups. Thus, 2-methoxyethyl methanesulfonate was used as a leaving group for dialkylation of DPAF 5 followed by Friedel-Crafts acylation with α-bromoacetyl bromide and AlCl3 to yield 7-α-bromoacetyl-9,9-di(2-methoxy)ethyl-2-diphenylaminofluorene , Subsequent conversion of the keto group of 7 to the corresponding 1,1-dicyano-ethylenyl (DCE) group was carried out by the reaction using malononitrile as a reagent, pyridine as a base, and titanium tetrachloride as a deoxygenation agent in dry chloroform at ambient temperature for a short period of 5.0 min. The reaction resulted in the corresponding diphenylaminofluorene BrCPAF-C2M9 in a yield of 89% after chromatographic purification . Attachment of a CPAF-C2M antenna arm to a C60 cage was carried out by identical reaction conditions as those for 1 with DBU (1.0 eq.) at room temperature for 4.0 h to afford 7--9,9-di(2-methoxyethyl)-2-diphenylaminofluorene C60(>CPAF-C2M), 2) as orange red solids in 53% yield (based on recovered C60). The bulkiness of DPAF-C18 and CPAF-C2M in size can prevent these two types of antenna moieties form locating in close vicinity to each other at the cage surface. By considering the regio-location of reactive bicyclopentadienyl olefin bonds on the fullerene surface, when the first antenna is bound at the north-pole location, the second antenna arm is most likely to be pushed away to the equator area of the C60 sphere. Therefore, only a very limited number of multiadduct regioisomers per C60 are likely to form. Indeed, by controlling the reaction kinetic rate with two molar equivalents of CPAF-C2M applied in the reaction with 1 in the presence of DBU (2.0 eq.), only two clear PTLC  bands in the product mixtures were observed in addition to the starting 1 (~15%). The first less polar product band at Rf = 0.5 was found to be the bisadduct C60(>DPAF-C18)(>CPAF-C2M) 3 isolated as orange-brown solids in 28% yield. The second more polar product band at Rf = 0.4 (toluene–ethyl acetate/4:1 as the eluent) was determined to be the trisadduct C60(>DPAF-C18)(>CPAF-C2M)24 isolated as red-brown solids in 40% yield.A similar reaction sequence was applied for the synthesis of the compound 1 and 2 was performed mainly by: (i) the clear detection of a group of molecular mass ion peaks with the maximum peak intensity centered at m/z 1,600 (MH+ of 1) and 1,258 (MH+ of 2)  stretching band centered 2,224 cm−1 for 2, chemical shifts of a keto carbonyl carbon peak at δ 188.33 and three carbons, -C=C(CN)2, –C≡N, and =C(CN)2, in 1,1-dicyanoethylenyl (DCE) moiety of 2 at δ 167.64, 112.99, and 88.07, respectively, in their 13C-NMR spectra fullerene triads 3 and tetrads 4. Upon the attachment of one CPAF-C2M antenna arm to 1, a new cyano stretching band centered at 2,223 cm−1 in addition to the carbonyl stretching band at 1678 cm−1 were detected as expected. Intensity of characteristic half-fullerene cage absorption band at ~526 cm−1 was found to decrease significantly going from that of 1, 3, to 4 (2M addend located at more than 90° away the DPAF-C18 arm (or the other side of the cage surface). Large difference of 1H chemical shifts among alkyl groups of DPAF-C18 (methyl and the most of methylene proton peaks at δ 0.69–1.29) and CPAF-C2M  allowed us to measure a clear proton integration count to verify the structure of 3 and 4 as a bisadduct and trisadduct, respectively, as shown in 4 was also evident by the detection of a higher aromatic proton integration ratio in the region of δ 7.5–7.8 and 8.10–8.15 H+, C60>H+, C60+, respectively, that was consistent well with the molecular structure of triad C60(>DPAF-C18)(>CPAF-C2M) 3. In the case of tetrad C60(>DPAF-C18)(>CPAF-C2M)24, a group of sharp molecular mass ions with the maximum mass at m/z 2,673 (MH+) and similar fragmented mass ions to those of 3 in the low mass region of m/z 720–1,000 were detected 2 by C60> when the number of antenna are more than one. Similarly, three fluorescence bands with λmax at 506, 531, and 615 (broad) nm  and broad bands at 525–650 nm along with the 1(C60>)* emission band centered at 708 nm. The latter broad bands fit in the similar range as those of C60[>1(CPAF)*-C2M]2. As the number of CPAF-C2M antenna being increased by one to the structure of tetrad C60(>DPAF-C18)(>CPAF-C2M)24, the intensity of broad FL bands at 525–650 nm became more pronounced while retaining the same intensity of the 1(C60>)* emission band at 709 nm [60[>1(DPAF)*-C18](>CPAF-C2M)2 state to both 1C60*(>DPAF-C18)(>CPAF-C2M)2 and C60(>DPAF-C18)[>1(CPAF)*-C2M]2 states. The latter energy-transfer is possible since: (i) the energy level of 1(CPAF)*-C2M is lower than that of 1(DPAF)*-C18, (ii) the energy of this FL band at 430–475 nm is slightly higher than that of the CPAF-C2M absorption λmax at 500 nm, and (iii) there is a partial overlap of emission/absorption bands to enhance the energy-transfer efficiency. Conversely, photoexcitation of 3 specifically on the CPAF-C2M antenna moiety at 478 nm, the resulting FL spectrum [1(C60>)* emission band centered at 707 nm. Intensity of the former broad band was significantly increased using 4 [2M antenna moieties. This confirmed the band was contributed from the C60(>DPAF-C18)[>1(CPAF)*-C2M]2 state, which was capable of inducing the 1C60*(>DPAF-C18)(>CPAF-C2M)2 state subsequently.Interestingly, upon photoexcitation of the triad 60(>CPAF-C9), 3, and 4, performed as a function of irradiance intensity using 125-fs laser pulses at either 780 nm (corresponding to the two-photon absorption of DPAF moieties) or 980 nm (corresponding to the two-photon absorption of CPAF moieties) at the concentration of 5 × 10−3 M in toluene, were provided in the 3 and 4 showing dual NIR two-photon absorption capability that led to large nonlinear light-transmittance reduction in intensity in these two wavelength ranges up to the fs-laser fluence of 120 GW/cm2. Observed sufficiently large two-photon absorption cross-section values of 3 and 4 may allow their uses as the nanocarbon core of 2γ-PDT agents after the chemical modification with water-soluble side-chains and cationic targeting segments on the fluorene ring moiety.Data of femtosecond Z-scans and nonlinear light-intensity transmittance reduction measurements of Ct-butoxide, potassium t-butoxide, aluminum chloride, titanium chloride, rac-BINAP, tris(dibenzylideneacetone)dipalladium(0), malononitrile, and 2-methoxy-ethanol, were purchased from Aldrich Chemicals  and used without further purification. The chemical 1-bromooctadecane was purchased from Tokyo Chemical Industry Co., Ltd. . A C60 sample with a purity of 99.0% was purchased from Term USA, Inc. . Both C60(>DPAF-C18)2 and C60(>CPAF-C2M)2 were synthesized by the similar methods described below.The reagents 1,8-diazabicyclo-undec-7-ene (DBU), 1-bromooctadecane, 2-bromofluorene, sodium 1H-NMR and 13C-NMR spectra were recorded on a Bruker Avance Spectrospin-500 spectrometer. UV-vis spectra were recorded on a Perkin Elmer Lambda 750 UV-vis-NIR Spectrometer. Photoluminescence (PL) spectra were measured using PTI Fluorescence Master Systems connected with a photomultiplier (914 Photomultiplier Detection System) with Xenon short arc lamp as the excitation source. Mass spectroscopic measurements were performed by the use of positive ion matrix-assisted laser desorption ionization (MALDI–TOF) technique on a micromass M@LDI-LR mass spectrometer. The sample blended or dissolved in the matrix material was irradiated by nitrogen UV laser at 337 nm with 10 Hz pulses under high vacuum. Mass ion peaks were identified for the spectrum using the MassLynx v4.0 software. In a typical experiment, the samples of C60(>DPAF-C18), C60(>CPAF-C2M), C60(>DPAF-C18)(>CPAF-C2M), or C60(>DPAF-C18)(>CPAF-C2M)2 were dissolved in CHCl3 in a concentration of 1.0 mg/mL. The matrix of 3,5-dimethoxy-4-hydroxycinnamic acid (sinapic acid) was dissolved in THF in a concentration of 10 mg/mL. The solution of matrix (1.0 mL) was taken and mixed with the sample solution (0.1 mL) prior to the deposition on a stainless-steel MALDI target probe. It was subsequently dried at ambient temperature.Infrared spectra were recorded as KBr pellets on a Thermo Nicolet Avatar 370 FT-IR spectrometer. Part A: In a round-bottom flask containing a mixture of triethylamine , 2-methoxyethanol , and anhydrous dichloroethane (150 mL) at 0 °C was added methanesulfonyl chloride  over a period of 20 min. The mixture was warmed to ambient temperature under a nitrogen atmospheric pressure and stirred for 12 h. It was quenched by the addition of water and washed with water (2 × 150 mL), dilute hydrochloric acid (1 × 100 mL), and saturated sodium bicarbonate (1 × 100 mL) in sequence. The organic layer was dried over sodium sulfate and concentrated in vacuo. The crude brownish liquid was vacuum distilled at 120−130 °C to afford 2-methoxyethylmethanesulfonate (17.9 g) in a nearly quantitative yield; 1H-NMR  δ 4.36 , 3.66 , 3.40 , and 3.05 .Part B: In a round-bottom flask containing a mixture of 2-diphenylaminofluorene 5 (DPAF)  and potassium t-butoxide  in dry THF (30 mL) at 0 °C was added 2-methoxyethylmethanesulfonate  over 10 min. The mixture was warmed to ambient temperature under a nitrogen atmosphere and stirred for 4.0 h. The reaction mixture was washed with brine (20 mL) and water (20 mL). Organic layer was dried over sodium sulfate and concentrated in vacuo. The crude product was then purified by column chromatography  via a chromatographic fraction corresponding to Rf = 0.7 on TLC (SiO2) with the same eluent to afford DPAF-C2M6 as white solids in a yield of 94% (0.66 g). Spectroscopic data: MALDI-MS (TOF) m/z 449 calculated for 12C311H3114N116O2; found, m/z 450 (MH+); 1H-NMR  δ 7.61 , 7.55 , 7.38 , 7.31 , 7.28−7.24 , 7.18−7.10 , 7.13−7.00 , 3.04 , 2.79−2.71 , and 2.26−2.20 ; 13C-NMR  δ 150.66, 149.32, 148.32, 147.95, 140.67, 135.72, 129.67, 127.79, 127.08, 124.43, 124.12, 123.18, 120.98, 119.69, 69.08, 58.77, 51.45, and 39.76.2M6  in 1,2-dichloroethane (50 mL). It was added α-bromoacetyl bromide  over a period of 10 min. The mixture was stirred for 4.0 h at 0 °C. The solution was worked up by slow addition of dilute HCl (1.0 N) solution (50 mL) while maintaining the temperature at 0 °C. The resulting organic layer was washed subsequently with dilute brine (2 × 50 mL) and water (2 × 50 mL) at room temperature and dried over magnesium sulfate. It was followed by the solvent removal in vacuo. The crude products were purified by column chromatography  at its chromatographic band corresponding to Rf = 0.2 on TLC (SiO2) with the same eluent to afford BrDPAF-C2M7 in 91% yield (6.3 g). Spectroscopic data: FT-IR (KBr) νmax 3,054 (w), 3037 (w), 2,925 (m), 2,871 (m), 2,804 (w), 1,693(w), 1,673 (m), 1,593 (s), 1,490 (s), 1,467 (m), 1,430 (w), 1,388 (w), 1,320 (w), 1,279 (s), 1,194 (w), 1,113 (m), 1,026 (w), 820 (w), 754 (m), 697 (m), 669 (w), and 627 (w) cm−1; UV-vis (CHCl3) λmax (ε) 299 (1.4 × 104) and 407 (2.1 × 104 L/mol-cm); 1H-NMR  δ 8.03−7.99 , 7.69 , 7.63 , 7.34−7.09 , 7.08−7.06 , 4.53 , 3.06 , 2.84−2.75 , and 2.33−2.21 ; 13C-NMR (CDCl3) δ 190.99, 151.81, 149.32, 149.20, 147.42, 146.14, 132.83, 131.74, 129.38, 129.29, 124.66, 123.46, 122.80, 121.81, 119.06, 117.86, 68.52, 58.29, 51.30, 38.89, and 31.05.To a suspension of aluminum chloride  in 1,2-dichloroethane (200 mL) at 0 °C was added a solution of DPAF-CPart A: In a round-bottom flask containing a mixture of 2-diphenylaminofluorene 5 , potassium t-butoxide  in dry THF (30 mL) at 0 °C was added 1-bromooctadecane  over 10 min. The mixture was warmed to ambient temperature under a nitrogen atmosphere and stirred overnight. The reaction mixture was washed with brine (40 mL) and water (40 mL) in sequence. The organic layer was dried over sodium sulfate and concentrated in vacuo. The crude product was purified by column chromatography  as a chromatographic fraction corresponding to Rf = 0.8 on TLC (SiO2) with the same eluent to give 9,9-dioctadecanyl-2-diphenylaminofluorene DPAF-C18 in 97% yield (2.44 g). Spectroscopic data: FT-IR (KBr) νmax 3,067 (w), 3,036 (w), 2,924 (s), 2,853 (s), 1,599 (m), 1,494 (m), 1,451 (w), 1,331 (w), 1,277 (m), 1,154 (w), 1,075 (w), 1,029 (w), 824 (w), 751 (m), 737 (m), 696 (m), 623 (w), and 513 (w) cm−1; MALDI-MS (TOF) m/z 838 calculated for 12C611H9114N1; found, m/z 839 (MH+); 1H-NMR  δ 7.63 , 7.58 , 7.33−7.25 , 7.15−7.13 , 7.05−7.01 , 1.93−1.81 , 1.33−1.07 , 0.91 , and 0.73−0.62 ; 13C-NMR  δ 152.50, 148.45, 147.44, 141.29, 136.81, 129.52, 124.12, 123.98, 123.09, 122.78, 120.70, 119.93, and 119.49.Part B: To a suspension of aluminum chloride  in 1,2-dichloroethane (50 mL) at 0 °C was added a solution of DPAF-C18  in 1,2-dichloroethane (30 mL). It was then added by α-bromoacetyl bromide  over 10 min. The mixture was stirred for 4.0 h at 0 °C. The solution was diluted by a slow addition of water (100 mL) while maintaining the reaction mixture temperature below 0 °C. The resulting organic layer was washed subsequently with dilute hydrochloric acid  and water (2 × 30 mL), and dried over magnesium sulfate followed by the solvent removal in vacuo. The crude yellow oil was purified by column chromatography  to afford BrDPAF-C188 in 96% yield (1.3 g). The product gave a chromatographic Rf at 0.5 on TLC (SiO2) with the same eluent. Spectroscopic data: FT-IR (KBr) νmax 3,063 (w), 3,034 (w), 2,923 (s), 2,852 (s), 1,677 (m), 1,595 (m), 1,493 (m), 1,466 (w), 1,346 (w), 1,279 (m), 1,182 (w), 1,027 (w), 819 (w), 753 (w), 697 (m), 620 (w), and 508 (w) cm−1; UV-vis (CHCl3) λmax (ε) 292 (1.9 × 104) and 407 (2.5 × 104 L/mol-cm); 1H-NMR  δ 7.95 , 7.93 , 7.64 , 7.59 , 7.27−7.23 , 7.14−7.12 , 7.05−7.02 , 4.49 , 1.97−1.81 , 1.25−1.04 , 0.87 , and 0.72−0.55 ; 13C-NMR (CDCl3) δ 190.99 (-C=O), 153.63 (aminoaryl carbon), 151.06 (aminoaryl carbon), 148.81 (aminoaryl carbon), 147.61, 146.89, 133.96, 131.55, 129.25, 128.80, 124.36, 123.09, 122.78, 121.61, 118.82, 118.20, 55.23, 39.96, 31.90, 31.15, 29.90, 29.67, 29.64, 29.62, 29.57, 29.55, 29.34, 29.29, 23.83, 22.67, and 14.10.60  and 7-α-bromoacetyl-9,9-dioctadecanyl-2-diphenylaminofluorene  in dry toluene (500 mL) was added DBU  under a nitrogen atmosphere. After stirring at room temperature for 5.0 h, suspended solids of the reaction mixture were filtered off and the filtrate was concentrated to a 10% volume. Crude product was precipitated by the addition of methanol and isolated by centrifugation . The isolated solid was found to be a mixture of the monoadduct and its bisadducts. Separation of these two product fractions were made by column chromatography (silica gel) using a solvent mixture of hexane−toluene (3:2) as the eluent. The first chromatographic band corresponding to Rf = 0.7 on TLC  afforded C60(>DPAF-C18) 1 as brown solids . Spectroscopic data: FT-IR (KBr) νmax 3,440 (m), 2,920 (s), 2,849 (s), 1,674 , 1,632 (m), 1,593 (s), 1,491 (m), 1,463 (m), 1,427 (m), 1,346 (w), 1,331 (w), 1,316 (w), 1,273 (m), 1,239 (w), 1,200 (m), 1,186 (w), 1157 (w), 1028 (w), 817 (w), 752 (m), 738 (w), 696 (m), 575 (w), 547 (w), 526 (m), and 490 (m) cm−1; MALDI-MS (TOF) m/z 1598 calculated for 12C1231H9114N116O1; found, m/z 1,601, 1,600 (MH+), 1,599, 866, 839, 762, 734, and 720; UV-vis (CHCl3) λmax (ε) 260 (1.3 × 105), 325 (4.7 × 104), and 411 (3.6 × 104 L/mol-cm) 1H-NMR  δ 8.43 , 8.32 , 7.78 , 7.61 , 7.25−7.22 , 7.11−7.09 , 7.03−7.00 , 5.66 , 2.03−1.84 , 1.29−1.04 , 0.87 , and 0.69 . 13C-NMR (CDCl3) δ 188.33 (-C=O), 153.55 (aminoaryl carbon), 151.20 (aminoaryl carbon), 148.77 (aminoaryl carbon), 147.96 (2C), 147.30 (2C), 147.20 (C), 146.73 (2C), 145.35 (2C), 145.24 (2C), 145.06 (2C), 144.96 (4C), 144.85 (2C), 144.70 (C), 144.52 (2C), 144.43 (4C), 144.13 (2C), 143.74 (2C), 143.49 (2C), 143.14 (2C), 142.96 (C), 142.91 (C), 142.83 (2C), 142.76 (2C), 142.57 (2C), 142.32 (2C), 142.07 (2C), 142.00 (2C), 141.90 (2C), 141.06 (2C), 140.76 (2C), 139.36 (2C), 136.46 (2C), 133.57, 133.22, 129.22, 128.62, 124.40, 123.15, 122.83, 122.42, 121.71, 119.14, 117.78, 72.48 (fullerenyl sp3 carbons), 55.09, 44.58, 40.14 (cyclopropanyl C60> carbon), 32.00, 30.16, 29.81, 29.56, 29.47, 24.11, 22.87, and 14.22. A total of 30 carbon peaks were accounted for 58 fullerenyl sp2 carbons at δ 136–148 indicated a C2-symmetry of the fullerene cage.To a mixture of Cα-bromoacetyl-9,9-di(2-methoxyethyl)-2-diphenylaminofluorene  and malononitrile  in dry chloroform (100 mL) was added pyridine (3.0 mL) while stirring under a nitrogen atmosphere. To this solution, titanium tetrachloride  was added in. After stirring at room temperature for 5.0 min, the reaction mixture was quenched with water (90 mL). The resulting organic layer was washed several times with water (100 mL each), dried over magnesium sulfate, and concentrated in vacuo to afford the crude orange-red oil. It was purified on a preparative chromatographic plate . A product fraction collected at Rf = 0.6 [toluene−ethyl acetate (4:1) as the eluent] gave BrCPAF-C2M9 in 89% yield (2.1 g). Spectroscopic data: FT-IR (KBr) νmax 3,058 (w), 3,035 (w), 2,924 (m), 2,870 (m), 2,855 (m), 2,809 (w), 2,226 (m), 1,593 (s), 1,547 (m), 1,490 (s), 1,468 (m), 1,451 (w), 1,384 (w), 1,346 (m), 1,318 (m), 1,279 (s), 1,191 (w), 1,113 (m), 1,028 (w), 957 (w), 821 (w), 755 (m), 698 (m), and 511 (w) cm−1; UV-vis (CHCl3) λmax (ε) 316 (2.1 × 104) and 489 (1.7 × 104 L/mol-cm); 1H-NMR  δ 7.70−7.66 , 7.59 , 7.30−7.27 , 7.15−7.13 , 7.09−7.06 , 4.61 , 3.03 , 2.87−2.77 , and 2.32−2.18 ; 13C-NMR (CDCl3) δ 170.85 [–C=C(CN)2], 151.77 (aminoaryl carbon), 149.91 (aminoaryl carbon), 149.38 (aminoaryl carbon), 147.28, 145.72, 132.39, 130.13, 129.36, 127.92, 124.74, 123.54, 122.91, 122.55, 121.72, 119.60, 117.55, 112.98 (–C≡N), 112.11 (–C≡N), 84.48 [=C(CN)2], 68.42, 58.31, 51.64, 38.93, and 28.68.To a mixture of 7-60  and 7-[2-bromo-1--1-methyl]-9,9-di(2-methoxyethyl)-2-diphenylaminofluorene  in dry toluene (150 mL) was added 1,8-diazabicyclo-undec-7-ene  under a nitrogen atmosphere. After stirring at room temperature for a period of 5.0 h, the reaction mixture was concentrated to a volume of approximately 10 mL. Crude product was precipitated by the addition of methanol and isolated by centrifugation . The precipitate was further purified by column chromatography  at the corresponding chromatographic Rf = 0.8 on TLC (SiO2) with the same eluent to afford C60(>CPAF-C2M) 2 in 53% yield (0.16 g). Spectroscopic data: FT-IR (KBr) νmax 3,439 (s), 2,980 (w), 2,920 (m), 2,868 (m), 2,824 (w), 2798 (w), 2,224 , 1,629 (m), 1,594 (vs), 1,538 (m), 1,491 (m), 1,466 (m), 1,428 (m), 1,347 (m), 1,319 (m), 1,279 (s), 1,186 (m), 1,115 (m), 1,028 (w), 958 (w), 888 (w), 820 (m), 805 (w), 754 (m), 697 (m), 668 (w), 577 (w), and 527 (m) cm−1; MALDI-MS (TOF) m/z 1,257 calculated for 12C961H3114N316O2; found, m/z 1,260, 1,259, 1,258 (MH+), 1,155, 987, 965, 919, 735, and 720; UV-vis (CHCl3) λmax (ε) 260 (1.2 × 105), 327 (5.1 × 104), and 501 nm (1.7 × 104 L/mol-cm); 1H-NMR  δ 8.14 , 8.11 , 7.76 , 7.56 , 7.26−7.23 , 7.09−7.08 , 7.06−7.03 , 5.51 , 2.95 , 2.73 , and 2.32−2.18 ; 13C-NMR (CDCl3) δ 167.64 [–C=C(CN)2], 151.83 (aminoaryl carbon), 150.31 (aminoaryl carbon), 149.45 (aminoaryl carbon), 147.23 (2C), 147.06 (2C), 146.07 (C), 145.69 (2C), 145.25 (4C), 145.21 (2C), 145.18 (2C), 145.15 (2C), 145.03 (2C), 144.72 (4C), 144.68 (4C), 144.53 (2C), 144.40 (2C), 144.15 (C), 143.69 (2C), 143.62 (2C), 142.94 (2C), 142.89 (4C), 142.36 (2C), 141.96 (2C), 141.94 (2C), 141.83 (2C), 141.36 (2C), 140.97 (2C), 137.31 (2C), 136.95 (2C), 132.12, 129.36, 128.46, 124.76, 123.63, 122.89, 122.39, 121.89, 119.67, 117.47, 112.99 (–C≡N), 112.92, 88.07 [=C(CN)2], 72.30 (fullerenyl sp3 carbons), 68.33, 58.26, 51.56, 41.22 (cyclopropanyl C60> carbon), and 39.26. A total of 30 carbon peaks representing 58 fullerenyl sp2 carbons at δ 136–148 indicated a C2-symmetry of the fullerene cage. To a mixture of C60(>DPAF-C18) 1  and 7-[2-bromo-1--1-methyl]-9,9-di(2-methoxyethyl)-2-diphenylaminofluorene  in dry toluene (100 mL) was added 1,8-diazabicyclo-undec-7-ene  slowly under a nitrogen atmosphere. After stirring at room temperature for a period of 5.0 h, the reaction mixture was concentrated to a volume of approximately 10 mL. Crude product was precipitated by the addition of methanol and isolated by centrifugation . The isolated solid was found to be a mixture of the fullerene multiadducts. Separation of these mixture was made by column chromatography (silica gel) using a solvent mixture of toluene–ethyl acetate (9:1) as the eluent. The first chromatographic band gave the unreacted starting compound 1 . The second chromatographic band corresponding to Rf = 0.5 on the thin-layer chromatographic plate  afforded the bisadduct product C60(>DPAF-C18)(>CPAF-C2M) 3 as orange-brown solids  in a 28% yield [based on the recovered C60(>DPAF-C18) amount]. The third chromatographic band corresponding to Rf = 0.4 on the thin-layer chromatographic plate  afforded the trisadduct product C60(>DPAF-C18)(>CPAF-C2M)24 as red-brown solids  in a yield of 40% [based on the recovered C60(>DPAF-C18) amount]. Spectroscopic data of C60(>DPAF-C18)(>CPAF-C2M) 3: FT-IR (KBr) νmax 3,424 (w), 3,063 (w), 3,030 (w), 2,921 (m), 2,850 (m), 2,223 (m), 1,678 (m), 1,593 (s), 1,568 (m), 1,537 (w), 1,492 (m), 1,465 (m), 1,426 (m), 1,346 (m), 1,277 (s), 1,202 (m), 1,115 (m), 1,074 (w), 962 (w), 895 (w), 819 (m), 753 (s), 696 (s), 578 (w), 526 (m), and 491 (w) cm−1; MALDI-MS (TOF) m/z 2,135 calculated for 12C1591H12214N416O3; found, m/z 2,136 (MH+), 2,135 (M+), 965, 920, 866, 763 {C60[>(C=O)-H]H+}, 735 (C60>H+), and 720 (C60+); UV-vis (CHCl3) λmax (ε) 255 (1.1 × 105), 304 (7.5 × 104), 413 (3.9 × 104), and 494 nm (2.3 × 104 L/mol-cm); 1H-NMR  δ 8.51−7.52 , 7.28−7.22 , 7.14−7.00 , 5.76−5.18 , 3.01−2.90 , 2.82−2.67 , 2.40−2.14 , 2.06−1.80 , 1.31−1.04 , 0.87 , and 0.67 . Spectroscopic data of C60(>DPAF-C18)(>CPAF-C2M)24: FT-IR (KBr) νmax 3,433 (m), 3,060 (w), 3,027 (w), 2,922 (m), 2,851 (m), 2,224 (m), 1,679 (w), 1,594 (s), 1,539 (w), 1,492 (s), 1,466 (m), 1,426 (m), 1,346 (m), 1,331 (w), 1,279 (s), 1,207 (w), 1,115 (m), 1028 (w), 958 (w), 892 (w), 818 (m), 753 (m), 697 (m), 616 (w), and 525 (w) cm−1; MALDI-MS (TOF) m/z 2,672 calculated for 12C1951H15314N716O5; found, m/z 2,673 (MH+), 2,672 (M+), 965, 920, 866, 763 {C60[>(C=O)-H]H+}, 735 (C60>H+), and 720 (C60+); UV-vis (CHCl3) λmax (ε) 304 (1.1 × 105), 417 (4.6 × 104), and 500 nm (4.6 × 104 L/mol-cm); 1H-NMR  δ 8.44−7.46 , 7.09−7.03 , 5.57−4.72 , 3.08−2.86 , 2.73 , 2.21 , 1.91 , 1.3−0.95 , 0.86 , and 0.64 .To the mixture of 7--9,9-di(octadecyl)-2-diphenylaminofluorene C60-(antenna)x analogous compounds as branched triad C60(>DPAF-C18)(>CPAF-C2M) and tetrad C60(>DPAF-C18)(>CPAF-C2M)2 nanostructures were synthesized and characterized by various spectroscopic methods. The design of 2PA-responsive chromophores was made by covalently attaching multiple light-harvesting donor antenna units on a C60 (acceptor) via a periconjugation linkage within a separation distance of 2.6–3.5 Ǻ. This structural design was intended to facilitate the ultrafast femtosecond intramolecular energy-transfer process from the photoexcited C60[>1(DPAF)*-C18](>CPAF-C2M)1or2 or C60(>DPAF-C18)[>1(CPAF)*-C2M]1or2 to the C60 cage moiety upon two-photon pumping at either 780 or 980 nm, respectively. Interestingly, by adjustment of a higher number of CPAF-C2M antenna, the resulting tetrads showed nearly equal absorption in extinction coefficients over the wavelength range of 400–550 nm that corresponds to near-IR two-photon based excitation wavelengths at 780–1,100 nm for broadband 2γ-PDT applications. We also found that the unique feature of intramolecular Förster energy-transfer phenomena from the photoexcited high-energy DPAF-C18 antenna unit to the low-energy CPAF-C2M moiety at the fullerene cage surface gave the fluorescence emission at slightly longer wavelengths than 600 nm in a cascade fashion. It may be correlated to and provide an interesting mechanism for the enhancement of 2PA cross-section values of these hybrid C60-(antenna)x nanostructures.Two new C


Glycosmis is a genus of evergreen glabrous shrub and distributed all over India. It possesses various medicinal properties and is used in indigenous medicine for cough, rheumatism, anemia, and jaundice. Glycosmis arborea is a rich source of alkaloids, terpenoids, coumarins, as well as flavonoids.G. arborea led to the isolation of one new flavone C-glycoside along with three known flavanoids, named as 5,7-dihydroxy-2-[4-hydroxy-3-(methoxy methyl) phenyl]-6-C-β-d-glucopyranosyl flavone (4), 5,7,4′-trihydroxy-3′-methoxy flavone (1), 5,4′-dihydroxy-3′-methoxy-7-O-β-d-glucupyranosyl flavanone (2), and 5,4′-dihydroxy-3′-methoxy-7-O-(α-l-rhamnosyl-(1‴→6‴)-β-d-glucopyranosyl) flavanone (3), respectively. The structures of all compounds were elucidated with the help of nuclear magnetic resonance spectrometry. Pure compounds and fractions were evaluated for pest antifeedant and antimicrobial activity.The chemical investigation of methanol fraction of the leaves of G. arborea. Among them, compound 4 showed significant antimicrobial activity.Four compounds were isolated from the leaves of  Glycosmis is a genus of evergreen glabrous shrub, distributed in warm and temperate regions of the world, and is a rich source of alkaloids and amide; however, terpenoids, coumarins, and flavonoids were also reported +, 285 [M-CH3]+, 272 [M-CO]+, 257 [M-43]+, 241, 242, 215, 204, 193, 176, 152 [A1]+, 148 [B1]+, 136, 124 [A1-28]+ 107, 105. 1H NMR : δ ppm 6.62  6.05 , 6.38 , 7.63  6.87 , 7.78  3.86 . 13C NMR : δ ppm 161.02 (C-2) 106.32 (C-3), 182.2 (C-4) 160.9 (C-5), 98.7 (C-6), 164.2 (C-7), 93.9 (C-8), 103.6 (C-4a), 157.1 (C-8a), 124.1 (C-1′), 115.6 (C-2′), 148.4 (C-3′), 145.2 (C-4′), 117.0 (C-5′), 122.3 (C-6′), 56.2 (OCH3).M.p. 287°C to 288°C; UV (MeOH) λmax nm: 295, 328. IR (KBr) νmax cm−1: 3423, 1620, 1594, 1351, 1071, 1012, 974, 921. ESIMS (% int.) (C22H24O11) m/z: 464 [M]+ (70), 449 [M-CH3]+ (3), 416 [M-OCH3-OH]+ (17), 381 (88), 353 (93), 302 [M-Glu]+ (4), 301 (4), 203 (6). 1H NMR : δ ppm 12.01 , 9.09 , 5.47  2.8 , 3.29 , 6.13 , 6.10 , 6.92 , 6.89 , 6.92 , 4.67 , 3.42-3.82 , 3.76 . 13C NMR : δ ppm 78.77 (C-2), 42.4 (C-3), 197.44 (C-4), 165.52 (C-5), 96.3 (C-6), 163.43 (C-7), 95.51 (C-8), 103.69 (C-4a), 162.88 (C-8a), 131.27 (C-1′), 112.2 (C-2′), 118.33 (C-3′), 148.36 (C-4′), 146.83 (C-5′), 114 (C-6′), 100.99 (C-1″), 71 to 76 (C-2″-C-5″), 66.41 (C-6″), 56.05 (OCH3).M.p. 264°C; UV (MeOH) λmax nm: 293, 326. IR (KBr) νmax cm−1: 3472 (OH), 1630, 1602 (aromatic), 1521, 1352, 1092, 815. ESIMS (C28H34O15) (% int.) m/z: 610 [M]+, 633 [M + Na]+ (54), 595 (2), 551 (5), 507 (7), 463 (7), 388 (77), 364 (93), 338 (70), 306 (50), 233 (35), 179 (43), 151 (16). 1H NMR : δ ppm 12.24 , 8.84 , 5.45 , 2.85 , 3.22 , 6.49 , 6.60 , 7.11 , 6.96 , 7.51 , 5.69 , 4.26 to 4.66 , 5.43 , 4.14 to 4.60 , 1.57 , 3.71 . 13C NMR : δ ppm 79.50 (C-2), 43.16 (C-3), 197.1 (C-4), 166.48 (C-5), 96.44 (C-6), 164.08 (C-7), 97.32 (C-8), 104.34 (C-4a), 163.49 (C-8a), 132.15 (C-1′), 118.49 (C-2′), 112.31 (C-3′), 148.43 (C-4′), 149.12(C-5′), 115.32 (C-6′), 102.51 (C-1″), 72.79 to 78.45 , 67.37 (C-6″), 101.55 (C-1‴), 69.83 to 77.24 , 18.61 , 55.89 (OCH3).UV (MeOH) λmax nm: 208, 306. IR (KBr) νmax cm−1: 3437, 1660, 1590, 1350, 1074, 901, 836. EIMS (C23H24O11) (% int.) m/z: 493 [M + OH]+ (18), 478 [M + 2H]+ (11), 448 [M-CO]+ (14), 455 (94), 444 (11), 433 (9), 413 (22), 301 (4), 260 (3), 203 (2), 136 (2). 1H NMR  and 13C NMR  δ (see Table UV (MeOH) Spodoptera litura was tested using the leaf dip method -6-C-β-d-glucopyranosyl flavone, a new flavone C-glycoside named as Arboreaside.The structure was supported by the mass spectroscopic studies, which showed molecular ion peak at 478 S. litura L. In a dual-choice leaf disc method, hexane and methanol fractions were tested for pesticidal potential. The hexane fraction showed a percent feeding index (PFI) of 46.71 ± 4.07, while the methanol fraction showed a PFI of 50.21 ± 5.01 as given in Table All the fractions and compounds were tested for antifeedant activity. Among them, the hexane and methanol fractions showed significant antifeedant action against 4 showed antimicrobial activity against the plant bacterial pathogens A. tumifaciens, P. syringae, and P. carotovorum. It was found that moderate inhibitory activities were observed against A. tumifaciens and P. carotovorum at a concentration of 200 μg, whereas P. syringae was found to be fatal at 500 μg. Compound 4 showed moderate inhibition against A. tumifaciens as shown in the Table 3.All the fractions and compounds were also tested for antibacterial activity. The methanol fraction and compound 1H and 13C NMR were scanned on a Bruker AVANCE 400 MHz  at C5D5N-d5, DMSO-d6, CD3OD, and CDCl3 at 400, 300, and 100 MHz with TMS as internal reference. Proton-detected heteronuclear correlations were measured using HMQC (optimized for JHC =14.5 Hz) and HMBC (optimized for JHC =7 Hz). Mass spectra were recorded on a Micromass Quattro II  at 70 eV for ESIMS. Column chromatography was carried out using silica gel /Merck ). Thin layer chromatography was carried out over plates made of silica gel G of Qualigen/Merck.All melting points are uncorrected and were taken in open capillaries. The UV spectra were recorded on a PerkinElmer Lambda 15 UV/VIS spectrophotometer  in methanol as blank. IR spectra were recorded on a PerkinElmer Infrared 15 in KBr pellets and are expressed per centimeter. The 4) along with three known compounds (1 to 3) from methanol fraction of G. arborea leaves. With regard to bioactivity, all fractions and isolated compounds were evaluated for antifeedant and antibacterial activity. Among them, hexane and methanol fractions showed antifeedant activity, whereas methanol fraction and compound 4 showed significant antibacterial activity.In conclusion, the present paper has shown the isolation and structure elucidation of one new flavone C-glycoside (The authors declare that they have no competing interests.


Structures and absolute configurations of 1 and 2 were determined by extensive spectroscopic methods, especially 2D NMR and electronic circular dichroism (ECD) analysis. Possible biosynthetic pathways for 1–3 were also proposed and discussed. Compounds 1 and 2 significantly inhibited human cancer K562, HL-60, HeLa, BGC-823 and MCF-7 cells, and compound 3 also inhibited the K562 and HL-60 cells. Both bioassay and chemical analysis  demonstrated that the parent strain G59 did not produce 1–3, and that DES-induced mutation(s) in the mutant BD-1-6 activated some silent biosynthetic pathways in the parent strain G59, including one set for 1–3 production.Two new drimenyl cyclohexenone derivatives, named purpurogemutantin ( In the LC-ESIMS analysis, 1–3 were also detected only in BD-1-6 but entirely not in G59  in the mutant BD-1-6 activated some silent biosynthetic pathways in the parent strain G59, including one set of those for the oduction , althoug1H and 100 MHz 13C NMR) NMR spectrometer using the solvent signals  as references, respectively.The melting point was measured on a Beijing Tiandiyu X-4 exact micro melting point apparatus (uncorrected). Optical rotations were measured on a Rudolph Research Autopol II spectropolarimeter. ESIMS was recorded on an Applied Biosystems API 3000 LC-MS spectrometer. HRESIMS was measured on an Agilent 6520 Q-TOF LC-MS spectrometer. UV data were recorded on a GBC Cintra 20 spectrophotometer, and IR spectra were taken on a Bruker Tensor-27 infrared spectrophotometer. CD data were recorded on a Biologic Science MOS 450 CD or a JASCO J-815 spectropolarimeter. 1D and 2D NMR spectra were obtained on a JEOL JNM-GX 400  were used in TLC and spots were detected under UV light (254 and 365 nm) or by using Vaughan’s reagent +, 441 [M + Na]+, 837 [2M + H]+, 859 [2M + Na]+; negative ESI-MS m/z: 531 [M + CF3CO2]−. Positive HRESIMS m/z: measured 419.2431 [M + H]+, calculated for C24H35O6 [M + H] 419.2434; measured 436.2692 [M + NH4]+, calculated for C24H38NO6 [M + NH4] 436.2699; measured 441.2234 [M + Na]+, calculated for C24H34O6Na [M + Na] 441.2253.Purpurogemutantin (2): Colorless oil (MeOH),  −13.7 ,  −9.3 ;  in literature +; negative ESIMS m/z: 341 [M − H2O − H]−, 359 [M − H]−, 395 [M + Cl]−. Positive HRESIMS m/z: measured 743.4486 [2M + Na]+, calculated for C44H64O8Na [2M + Na] 743.4499; measured 399.1939 [M + K]+, calculated for C22H32O4K [M + K] 399.1938; measured 383.2193 [M + Na]+, calculated for C22H32O4Na [M + Na] 383.2198; measured 361.2377 [M + H]+, calculated for C22H33O4 [M + H] 361.2379; measured 343.2269 [M − H2O + H]+, calculated for C22H33O4 [M − H2O + H] 343.2273. Negative HRESIMS m/z: measured 395.1995 [M + Cl]−, calculated for C22H32O4Cl [M + Cl] 395.1989; measured 359.2223 [M − H]−, calculated for C22H31O4 [M − H] 359.2222; measured 341.2118 [M − H2O − H]−, calculated for C22H29O3 [M − H2O − H] 341.2117.Purpurogemutantidin (terature ). UV λma3): a pale-colored solid (MeOH),  +25.7 , ([α]D +29 (MeOH) in literature [m/z: 361 [M + H]+, 378 [M + NH4]+, 383 [M + Na]+; negative ESIMS m/z: 359 [M − H]−, 395 [M + Cl]−, 405 [M + HCOO]−. IR νmax cm−1: 3400, 2938.6, 2867.8, 2844.7, 1677.4, 1458.9, 1439.9, 1387.8, 1365.7, 1278.9, 1204.3, 1103.6, 1030.4, 882.1, 669.1. CD  Δε (nm): 0 (389), +2.76 (334), 0 (280), −4.41 (243), 0 (218), +0.31 (215), 0 (212), −1.92 (204). CD λmax nm (mdeg) in MeOH at 1.0 mg/mL: 389 (0), 334 (+25.33), 280 (0), 243 (−40.40), 218 (0), 215 (+2.80), 212 (0), 204 (−17.56). 1H NMR  δ: 5.91 , 4.77 , 4.52 , 4.29 , 4.21  , 3.67 , 2.33 , 2.31 , 1.92 , 1.81 , 1.76 , 1.75 , 1.72 , 1.60 , 1.49 , 1.38 , 1.30 , 1.21 , 1.18 , 1.12 , 0.70 , 0.79 , 0.85 . 13C NMR δ: 195.4 (C-1′), 161.2 (C-3′), 150.5 (C-8), 120.2 (C-2′), 107.4 (C-12), 66.2 (C-4′), 62.3 (C-5′), 62.2 (C-7′), 61.2 (C-6′), 56.9 (C-5), 52.9 (C-9), 43.3 (C-3), 40.7 (C-10), 40.0 (C-1), 39.3 (C-7), 34.5 (C-4), 34.1 (C-13), 25.6 (C-6), 22.2 (C-14), 22.0 (C-11), 20.4 (C-2), 15.0 (C-15). The above 1H and 13C NMR data were assigned on the basis of DEPT, 1H–1H COSY, HMQC, HMBC and NOESY experiments.Macrophorin A (terature ). Positi1–3 were dissolved in MeOH to prepare a 10 mg/mL sample solution for HPLC analysis. HPLC analysis of the samples was carried out on a Venusil MP C18 column  using the same Waters HPLC equipment mentioned. Each 5 μL of sample solutions was injected into the column after filtration by a 0.22 μm pore membrane filter. Then, the elution was performed using MeOH–H2O in linear gradient . The acquired photodiode array (PDA) data were processed by Empower PDA software to obtain targeted HPLC data.The EtOAc extracts of G59 and BD-1-6 cultures and the compounds 1–3 samples were used as control. HPLC was carried out on a Venusil MP C18 column  using MeOH–H2O in linear gradient  as mobile phase. The mass detector was set to scan a range from m/z 150 to 1500 in positive (for 1–3) or negative (for 1) mode. The acquired data were processed by Analyst 1.4 software to obtain targeted LC-ESIMS data.The LC-ESIMS analysis was performed on a LC-MS equipment equipped with Agilent 1100 HPLC system, AB Sciex API 3000 LC-MS/MS system, and AB Sciex Analyst 1.4 software. The above mentioned EtOAc extracts of G59 and BD-1-6, at 10 mg/mL in MeOH, were employed also for the LC-ESIMS analysis and crude 1) and purpurogemutantidin (2), and the known macrophorin A (3) were isolated from a mutant BD-1-6 obtained by random DES mutagenesis of a marine-derived fungal strain Penicillium purpurogenum G59. The structures and absolute configurations of 1 and 2 were determined by extensive spectroscopic methods, especially 2D NMR and ECD analysis. Possible biosynthetic pathways for 1–3 were also proposed and discussed. Compounds 1 and 2 significantly inhibited human cancer K562, HL-60, HeLa, BGC-823 and MCF-7 cells, and 3 also inhibited the K562 and HL-60 cells. Both bioassay and chemical analysis (HPLC and LC-ESIMS) demonstrated that the parent strain G59 did not produce 1–3, and that the DES-induced mutation(s) in BD-1-6 activated some silent biosynthetic pathways in the parent strain G59, including one set for the 1–3 production.Two new drimenyl cyclohexenone derivatives, purpurogemutantin (


S)-mandelate and irradiation of this complex led to the formation of the dibenzosemibullvalene with moderate stereoselectivity (68:32 er). In contrast, the thioureido derivative showed no such effect upon complexation of chiral anions.Bisureido- and a bisthioureido-substituted dibenzobarrelene derivative were synthesized and the photoreactivity of two representative examples were studied. Direct irradiation of the ureido-substituted derivative induces a di-π-methane rearrangement to the corresponding dibenzosemibullvalene derivative, whereas the thioureido-substituted derivative is almost photoinert. Complexes of the latter derivative with chloride, carboxylates, or sulfonate anions, however, are efficiently transformed to the dibenzosemibullvalene product upon irradiation, presumably by suppressing the self-quenching of the thiourea units in the complex. The association of the ureido-substituted dibenzobarrelene derivative with ( The control of the selectivity of a photoreaction by supramolecular interactions has recently received much attention –3. For e1a) and its derivatives has been shown to be an appropriate model reaction for the assessment of substituent effects on the selectivity of organic photoreactions  rearrangement –28 of dieactions  29–30].–30.1a) areaction –30. In t1b crystallizes in the chiral space group P212121, and irradiation of the chiral crystals gives dibenzosemibullvalene 2b with a high enantiomeric excess, >95% ee . The corresponding Job plot confirms the 1:1 complex between 1h and SMD -mandelate (SMD). Thus, a complex of the dibenzobarrelene 1h with SMD was irradiated in acetone solution at different concentrations and with varied host–guest ratios  led to a decrease of the stereoselectivity. In addition, changes in the concentration of the dibenzobarrelene 1h did not have a significant influence on the stereoselectivity of the reaction. Based on these results, the following experiments were carried out with a concentration of 0.25 mM for the dibenzobarrelene derivative 1h and 1.1 molar equiv of the chiral additive (R)-enantiomer of mandelate induced the same extent of stereoselectivity with the reverse ratio of products. For comparison, the photoreaction of dibenzobarrelene 1h was performed in the presence of other chiral anions, namely (R)-thiazolidine-4-carboxylate (RTZ), (S)-camphor-10-sulfonate (SCS), (R)-carnitine (RCN), and (2S)-1-[(benzyloxy)carbonyl]-2-pyrrolidinecarboxylate (SCP) (S)-mandelate.Since it was demonstrated that the ureido- and thioureido-substituted dibenzobarrelene derivatives rs of 2h . The abste (SCP) . In each1h was also investigated in the presence of (S)-mandelate , whereas in acetonitrile (40:60 er), THF (45:55 er) or MeOH, EtOH or 2-PrOH (50:50 er) the DPM rearrangement of compound 1h proceeds with very low or no selectivity.The influence of the solvent on the DPM rearrangement of compound 1i were studied with chiral mandelate, camphorsulfonate and binaphthyl phosphonate in a variety of solvents including acetone, acetonitrile, ethyl acetate, dichloromethane, and benzene. Although the DPM rearrangement of the dibenzobarrelene 1i took place readily upon irradiation and the semibullvalene photoproducts were isolated by column chromatography in very good yields, none of these products proved to be enantiomerically enriched, as determined by 1H NMR experiments with SMD as the chiral shift reagent.In additional experiments, the photoreactions of the thioureido-substituted dibenzobarrelene derivative 1h and 1i provide useful initial information about the potential of anion-controlled photoreactions.It is well established that the regio- and stereoselectivity of a photoreaction may be induced by the selective preorganization of the substrates by hydrogen bonding between neutral organic functionalities with an appropriate substitution pattern or by complexation of crown-ether units to cationic guest molecules –3. In co1h takes place even without external sensitizers suggests that an efficient intersystem crossing (ISC) process exists for the excited chromophore 1h that directs the photoreaction predominately to the triplet pathway. The 3,5-bis(trifluoromethyl)phenyl substituent may be responsible for the ISC, because m-bis(trifluoromethyl)benzene has an ISC quantum yield of ΦISC = 0.12 (λex = 254 nm) in the gas phase, and the latter compound is able to sensitize a triplet-state E/Z-isomerization of alkenes . Absorption spectra were recorded on a Varian 100 Bio spectrometer at 25 °C. Melting points were determined on a Büchi 510K and are uncorrected. Mass spectra were recorded on a Hewlett-Packard HP 5968 (EI) and a Finnigan LCQ Deca instrument (ESI). Elemental analyses were performed on a KEKA-tech EuroEA combustion analyzer by Mr. H. Bodenstedt, Organic Chemistry I, University of Siegen. TLC analyses were performed on silica-gel sheets (Macherey-Nagel Polygram Sil G/UV254). Unless otherwise mentioned, commercially available chemicals were reagent grade and were used without further purification. Tetrabutylammonium hydroxide in MeOH (1.0 M) and (S)-camphor-10-sulfonic acid were obtained from Aldrich. (R)-Mandelic acid and (S)-mandelic acid were obtained from Fluka. (R)-Thiazolidine-4-carboxylic acid and (2S)-1-[(benzyloxy)carbonyl]-2-pyrrolidinecarboxylic acid were obtained from Acros. (R)-Carnitine was obtained from Alfa-Aesar. Preparative column chromatography was performed on MN Silica Gel 60 M .Irradiations were performed with a TQ150 middle-pressure mercury lamp , which was placed inside a quartz cooling tube. The reaction mixture was placed ca. 10–15 cm in front of the lamp.General procedure for the preparation of bisurea- and bisthiourea derivatives of dibenzobarrelene (GP1): The isocyanate or isothiocyanate derivative (1.1 molar equiv) was added to a stirred solution of 11,12-bis(aminomethyl)-9,10-dihydro-9,10-ethenoanthracene  : δ 0.88 , 1.23–1.34 , 2.95–2.99 , 3.83 , 5.08 , 5.83–5.85 , 6.91, 7.22 . 13C NMR : δ 14.1 (CH3), 19.9 (CH2), 32.6 (CH2), 38.0 (CH2), 53.1 (CH), 122.9 (CHar), 124.5 (CHar), 143.3 (Cq), 146.6 (Cq), 158.6 (C=O). Anal. Calcd for C34H28N4O2 (460.6): C, 73.01; H, 7.88; N, 12.16. Found: C, 73.25; H, 7.95; N, 12.04.11,12-Bis(N’-phenylureidomethyl)-9,10-dihydro-9,10-ethenoanthracene (1f): Prepared from dibenzobarrelene 1d  according to GP1 and collected by filtration and obtained as a white powder , mp 309–312 °C (dec.). 1H NMR : δ 4.07 , 5.15 , 6.37 , 6.90–6.93 , 7.20–7.41 , 8.29 . 13C NMR : δ 37.9 (CH2), 53.3 (CH), 118.3 (CHar), 121.5 (CHar), 123.0 (CHar), 124.6 (CHar), 129.0 (CHar), 140.8 (Cq), 143.5 (Cq), 146.5 (Cq), 155.8 (C=O). MS (EI): m/z = 500 [M+]. Anal. Calcd for C34H28N4O2 (500.6): C, 76.78; H, 5.64; N, 11.19. Found: C, 76.68; H, 5.67; N, 11.12.11,12-Bis[N’-(4-n-butylphenyl)ureidomethyl]-9,10-dihydro-9,10-ethenoanthracene (1g): Prepared from dibenzobarrelene 1d  according to GP1, collected by filtration and dried in vacuo. White amorphous solid, yield 0.21 g , mp > 320 °C. 1H NMR : δ 0.89 , 1.26–1.55 , 2.47–2.50 , 3.97 , 5.14 , 6.06 , 6.90–6.92 , 7.03–7.05 , 7.25–7.29 , 8.42 . 13C NMR : δ 14.2 (CH3), 22.1 (CH2), 33.7 (CH2), 34.5 (CH2), 37.9 (CH2), 53.3 (CH), 118.4 (CHar), 123.0 (CHar), 124.6 (CHar), 128.8 (CHar), 135.3 (CHar), 138.4 (Cq), 143.5 (Cq), 146.5 (Cq), 155.9 (C=O). MS (EI): m/z (%) = 613 [M+]. Anal. Calcd for C40H44N4O2 (612.8): C, 78.40; H, 7.24; N, 9.14. Found: C, 78.12; H, 7.25; N, 9.09.11,12-Bis{N’-ureidomethyl}-9,10-dihydro-9,10-ethenoanthracene (1h): Prepared from dibenzobarrelene 1d  according to GP1. After filtration of the precipitate, the product was purified by recrystallization from CH2Cl2/hexane and obtained as a white solid , mp > 300 °C. 1H NMR : δ 4.18 , 5.25 , 6.43 , 6.86 , 7.27 , 7.51 , 8.03 , 8.64 . 13C NMR : δ 38.3 (CH2), 53.3 (CH), 113.9 (CHar), 117.7 (CHar), 122.4 (CHar), 123.0 (CHar), 124.6 (CHar), 125.1 (CHar), 130.7 (CHar), 131.1 (CHar), 142.7 (Cq), 143.4 (Cq), 146.4 (Cq), 155.4 (C=O). UV (MeCN): λmax (log ε) = 229 (4.49), 246 (4.86), 272 (4.01), 280 (4.13). MS (ESI–): m/z (%) = 771 (100) [M − H]−. Anal. Calcd for C36H24F12N4O2 (772.6): C, 55.97; H, 3.13; N, 7.25. Found: C, 55.58; H, 2.85; N, 7.04.11,12-Bis{N’-thioureidomethyl}-9,10-dihydro-9,10-ethenoanthracene (1i): Prepared from dibenzobarrelene 1d  according to GP1 and obtained by recrystallization from CH2Cl2/hexane as a white solid , mp > 300 °C. 1H NMR : δ 4.66 , 5.36 , 6.92 , 7.30–7.32 , 7.73 , 8.25–8.26 , 9.57 . 13C NMR : δ = 43.4 (CH2), 54.2 (CH), 117.9 (CHar), 121.5 (CHar), 123.6 (CHar), 125.3 (CHar), 127.0 (CHar), 132.0 (CHar), 142.4 (Cq), 144.1 (Cq), 146.8 (Cq), 182.4 (C=O). UV (MeCN): λmax (log ε) = 230 (4.49), 246 (4.59), 272 (4.46), 280 (4.47). MS (ESI): m/z (%) = 803 (100) [M − H]−. Anal. Calcd for C36H24F12N4S2 (804.7): C, 53.73; H, 3.01; N, 6.96; S, 7.97. Found: C, 53.58; H, 2.79; N, 6.84; S, 7.97.General procedure for the synthetic photolysis in solution (GP2): Solutions of the substrate (10−3–10−2 mol/l) were placed in a Duran flask (acetone) or quartz test tube (other solvents), and argon gas was bubbled carefully through the solution for at least 20 min. The solution was irradiated for 4–15 h with stirring until the starting material was fully converted as determined by TLC or 1H NMR spectroscopic analysis. After evaporation of the solvent in vacuo, the photolysate was analyzed by 1H NMR spectroscopy. In preparative experiments, the photoproduct was isolated by recrystallization or column chromatography.4b,8b-Dihydro-8c,8e-bis{N’-ureidomethyl}dibenzocyclopropapentalene (2h): Prepared by irradiation of 1h  in acetone solution according to GP2 and obtained as white crystals , mp 246–247 °C. 1H NMR : δ 3.22 , 3.78 , 3.82 , 3.94 , 4.44 , 4.63 , 6.42 , 6.53 , 6.99–7.05 , 7.21–7.25 , 7.36–7.38 , 7.49 , 8.09 , 8.13 , 8.75 , 8.81 . 13C NMR : δ 41.3 (CH2), 41.4 (CH2), 46.7 (CH), 53.3 (Cq), 58.8 (Cq), 67.4 (CH), 115.6 (CHar), 115.6 (CHar), 115.6 (CHar), 115.7 (CHar), 119.3 (CHar), 119.3 (CHar), 123.4 (CHar), 123.4 (CHar), 124.5 (CHar), 124.6 (Cq), 125.8 (CHar), 126.3 (Cq), 126.4 (CHar), 126.6 (CHar), 128.1 (CHar), 128.2 (CHar), 128.3 (CHar), 128.5 (CHar), 133.2 (Cq), 133.2 (Cq), 133.4 (Cq), 133.4 (Cq), 139.9 (CHar), 140.1 (CHar), 144.4 (Cq), 144.6 (Cq), 151.8 (Cq), 152.8 (Cq), 156.7 (C=O), 157.2 (C=O). MS (ESI): m/z (%) = 771 (100) [M − H]−. An analytical sample was obtained by recrystallization from ethyl acetate/hexane and contained one equiv of ethyl acetate as the lattice solvent. Anal. Calcd for C36H24F12N4O2·EtOAc (860.7): C, 55.82; H, 3.75; N, 6.51. Found: C, 55.93; H, 3.51; N, 6.45.4b,8b-Dihydro-8c,8e-bis{N’-thioureidomethyl}dibenzocyclopropapentalene (2i): Prepared by photoreaction of 1i  in acetonitrile solution in the presence of 2 molar equiv of ammonium chloride according to GP2. After the irradiation the inorganic components were removed by column filtration , and the residue was recrystallized from ethyl acetate/hexane to give light yellow crystals , mp 181–182 °C. 1H NMR : δ 1.20 , 3.43 , 4.05 , 4.22 , 4.29 , 4.48 , 4.74 , 4.76 , 7.00–7.09 , 7.24–7.28 , 7.36–7.38 , 7.67 , 7.81 , 7.89 , 8.23 , 8.32 , 9.45 , 9.53 . 13C NMR : δ 14.5 (CH3), 20.8 (CH2), 44.2 (CH2), 45.2 (CH2), 50.3 (Cq), 57.2 (CH), 59.7 (CH), 65.1 (Cq), 116.5 (CHar), 116.7 (c), 121.5 (CHar), 121.6 (CHar), 122.1 (CHar), 122.5 (CHar), 123.8 (CHar), 124.3 (CHar), 124.3 (CHar), 124.9 (CHar), 126.4 (CHar), 126.4 (CHar), 126.6 (CHar), 126.9 (CHar), 130.6 (CHar), 130.6 (CHar), 130.8 (CHar), 130.9 (CHar), 131.1 (Cq), 131.1 (Cq), 137.6 (Cq), 137.7 (Cq), 141.8 (Cq), 142.0 (Cq), 149.9 (Cq), 150.8 (Cq), 181.4 (C=S), 181.8 (C=S), two Cq signals are overlapped in the aromatic region. MS (ESI): m/z (%) = 803 (100) [M − H]−. Anal. Calcd for C36H24F12N4S2·EtOAc (892.8): C, 53.81; H, 3.61; N, 6.28; S, 7.18. Found: C, 54.03; H, 3.28; N, 6.32; S, 7.09.Preparation of the tetrabutylammonium salts of chiral acids: The tetrabutylammonium salts of the chiral carboxylates SMD, RMD, RTZ, and SCP were prepared by the neutralization of the corresponding chiral acids with tetrabutylammonium hydroxide (1.0 M in MeOH) [in MeOH) . The resSCS was prepared according to the literature procedure [The tetrabutylammonium salt of (S)-camphor-10-sulfonate rocedure , and usePhotoreaction of dibenzobarrelene derivatives 1h and 1i in the presence of chiral anions: The dibenzobarrelene derivatives 1h or 1i (50 μmol) were dissolved in a stock solution  of the chiral tetrabutylammonium salt, and the solvent was removed in vacuo. The resulting residue was re-dissolved in the solvent of choice (200 mL). Argon gas was bubbled through the solution for 20 min to remove residual oxygen from the solvent. The reaction container  was placed ca. 15 cm in front of the light source, and the solution was irradiated for 3–4 h (TLC control). The solvent was removed in vacuo and the major photoproduct purified by column chromatography . The photoproduct was analyzed by 1H NMR spectroscopy in the presence of 5 equiv of tetrabutylammonium SMD as chiral shift reagent. The enantiomeric ratio of the semibullvalene mixture was determined by the integration of the aromatic proton signals from each isomer (δ in the range of 8.4–8.7 ppm), and by the integration of the NH and cyclopropane CH signals. Each spectroscopic measurement was repeated twice to ensure the reproducibility.1H NMR and 13C NMR spectra of compounds 1e–i and 2h–i; 1H NMR spectra of 2h with (S)-mandelate (SMD) at different molar ratios and corresponding Job plot; 1H NMR spectra of 2h with (S)-mandelate (SMD) as chiral shift reagent.File 1Supporting Information for: Effects of anion complexation on the photoreactivity of bisureido- and bisthioureido-substituted dibenzobarrelene derivatives.


Background: α,ω-Difunctional substrates are useful intermediates for polymer synthesis. An attractive, sustainable and selective (but as yet unused) method in the chemical industry is the oleochemical cross-metathesis with preferably symmetric functionalised substrates. The current study explores the cross-metathesis of methyl oleate (1) with cis-2-butene-1,4-diyl diacetate (2) starting from renewable resources and quite inexpensive base chemicals.Results: This cross-metathesis reaction was carried out with several phosphine and N-heterocyclic carbene ruthenium catalysts. The reaction conditions were optimised for high conversions in combination with high cross-metathesis selectivity. The influence of protecting groups present in the substrates on the necessary catalyst loading was also investigated.Conclusions: The value-added methyl 11-acetoxyundec-9-enoate (3) and undec-2-enyl acetate (4) are accessed with nearly quantitative oleochemical conversions and high cross-metathesis selectivity under mild reaction conditions. These two cross-metathesis products can be potentially used as functional monomers for diverse sustainable polymers. Quantitative conversions of methyl 10-undecenoate were obtained with a catalyst loading of 5 mol % [Ru]-1 -1 .1) with cis-2-butene-1,4-diyl diacetate (2). The self-metathesis of methyl oleate (1), which yields octadec-9-ene (5) and dimethyl octadec-9-enedioate (6), is the only concurrent metathesis reaction which had to be suppressed  and undec-2-enyl acetate (4). Both are thus derived from a renewable precursor and are interesting substrates for the synthesis of different types of polymers. Substrate 2 can be considered as an important cross-metathesis partner for the metathetical conversion of methyl oleate 1 in terms of its short chain length. Thus, the chain length of the cross-metathesis products is appropriate for certain polymer applications. Therefore, in contrast to polymers prepared from short-chain monomers, these polymers have a higher flexibility and higher stability against hydrolysis -1 to [Ru]-8 (1) with cis-2-butene-1,4-diyl diacetate (2). The reactions were performed with a fivefold excess of 2 in toluene at 50 °C for 5 h to shift the reaction equilibrium towards the cross-metathesis products 3 and 4.The catalytic activity of the ruthenium complexes o [Ru]-8  using a 1 and yields of the cross-metathesis products 3 and 4 were determined by gas chromatography with internal standard.The differences in metathesis activity of these investigated metathesis initiators are presented in 1 (about 15%) and yields of each of the desired cross-metathesis products 3 and 4 (about 5%) were achieved using the ruthenium phosphine complexes [Ru]-1 and [Ru]-3 ([Ru]-1 and its indenylidene counterpart [Ru]-3. The self-metathesis of methyl oleate (1) mentioned above could not be suppressed. Other side-reactions were not observed. No double-bound isomerisation took place. Promising results were obtained with catalysts bearing N-heterocyclic carbene ligands (1) was converted and the yields of the cross-metathesis products 3 and 4 of ca. 28% were achievable with ruthenium catalysts [Ru]-2 and [Ru]-4  were obtained with ruthenium complexes [Ru]-5–[Ru]-8. Due to their bidentate Schiff base ligands, they must be chemically activated by the addition of 100 equiv of phenyltrichlorosilane [[Ru]-7 and [Ru]-8 were the most active catalysts due to their space-filling isopropyl substituted Schiff base ligands -7 and [Ru]-8 is decisively higher compared to the other catalysts used.Comparable or higher conversions of rosilane –31. With1 (from 15 to 94%) and of yield of each of the cross-metathesis products 3 and 4 (from 2 to 66%) were obtained with a catalyst loading of [Ru]-7 in a range of 0.1 and 1.5 mol %. Cross-metathesis selectivity was increased by 60% to 67%. With a catalyst loading of 1.5 mol % of [Ru]-7, a nearly quantitative conversion of unsaturated fatty ester 1 was achievable (1 were obtained in lower amounts (ca. 14% of each self-metathesis product 5 and 6). From the results, it can be concluded that a catalyst loading of above 1.0 mol % of the ruthenium catalyst [Ru]-7 was necessary for efficient conversion of cis-2-butene-1,4-diyl diacetate (2). Further increasing the quantity of [Ru]-7 led neither to an essentially higher oleochemical conversion nor to higher cross-metathesis yields  was quite high (<96%). The cross-metathesis yields reached its maximum at 77% using an eightfold excess of 2 .Independent of the excess of ess of 2 , entry 4d yields , entry 51) was already converted  appears to be temperature-independent (1) underwent a rapid self-metathesis at low reaction temperatures  is required. On increasing the reaction temperature from 30 to 50 °C an increase in the yields of the cross-metathesis products (up to 77%) was observed. At the same time the self-metathesis reaction of 1 was hindered and only 10% of each of the self-metathesis products 5 and 6 was produced.Moreover, the conversion of methyl oleate  with cis-2-butene-1,4-diyl diacetate (2) were applied to the cross-metathesis reaction of the corresponding fatty acid 7 with the diol 8. The oleic acid (7) was reacted with both cis-2-butene-1,4-diyl diacetate (2) and the diol 8  with the protected diol 2 was nearly quantitative using 1.5 mol % of the ruthenium complex [Ru]-7 at 50 °C within 5 h (7) with 2 were only achieved with the use of 3.0 mol % of the same ruthenium catalyst. Under otherwise similar reactions conditions, 75% of oleic acid was converted  with cis-2-butene-1,4-diyl diacetate (2) was feasible with the relatively low catalyst loading of the Schiff base ruthenium catalyst [Ru]-7 to yield two value-added and sustainable intermediates in one step. Methyl 11-acetoxyundec-9-enoate (3) and undec-2-enyl acetate (4) are both very interesting substrates for polymer synthesis. They could be prepared under mild reaction conditions within 5 h. Moreover, this is an advantageous contribution towards the synthesis of sustainable monomer units because a new α,ω-difunctional substrate class starting from a renewable compound and an inexpensive base chemical was prepared.In conclusion, the cross-metathesis of methyl oleate  (97%), solvents and reagents were purchased from Sigma-Aldrich. Benzylidene ruthenium catalysts [Ru]-1 and [Ru]-2 were obtained from Sigma-Aldrich and the remaining indenylidene ruthenium catalysts [Ru]-3–[Ru]-8 were provided by Umicore AG & Co. KG and were used as received.Sunflower oil with a high oleic content (91.4% oleic acid) was obtained from Emery Oleochemicals. 1) was prepared by transesterifcation of high oleic sunflower oil with methanol using hypostoichiometric amounts of sodium methoxide (30% in methanol). cis-2-Butene-1,4-diyl diacetate (2) was prepared by the pyridine catalysed acylation of the diol 8 with acetic anhydride according to -1–[Ru]-8 were added in the range of 0.1–2.0 mol % to the reaction mixture. In the case of Schiff base ruthenium catalysts [Ru]-5–[Ru]-8, 100 equiv of phenyltrichlorosilane  were also added. The reaction mixture was stirred magnetically at the appropriate temperature (20–50 °C) for the appropriate time (1–9 h). After completion of the reaction, the mixture was cooled to ambient temperature. Conversion and yield analyses were performed by gas chromatography. The metathesis products were isolated after removing toluene in vacuo by flash chromatography on silica gel with cyclohexane/ethyl acetate (from 10/1 to 1/2) as eluent, and subsequently characterised by NMR spectroscopy.A flame-dried Schlenk tube was charged with 0.050 g (0.17 mmol) methyl oleate : δ (ppm) = 0.88 , 1.28 , 1.61 -CH2-CH2-), 2.00 , 2.30 -CH2-), 3.66 , 5.34 . 13C NMR : δ (ppm) = 14.0, 22.6, 24.9, 27.0, 27.1, 28.9, 29.0, 29.1, 29.2, 29.4, 29.6, 29.7, 31.8, 34.0, 51.3, 129.6, 129.9, 174.2. MS : m/z (%) = 296 (4) [M+], 264 (28), 246 (2), 235 (3), 222 (17), 207 (3), 194 (3), 180 (16), 166 (8), 152 (10), 137 (14), 123 (24), 110 (29), 97 (54), 83 (55), 74 (65), 69 (68), 55 (100), 41 (76), 29 (29).1H NMR : δ (ppm) = 1.96 -CH3), 4.57 , 5.64 . 13C-NMR : δ (ppm) = 20.6, 59.7, 127.8, 170.3. MS : m/z (%) = 172 (14) [M+], 113 (7), 99 (1), 82 (2), 70 (46), 61 (4), 53 (2), 43 (100), 39 (6), 27 (6).1H NMR : δ (ppm) = 1.61 , 1.66 -CH2-CH2-), 2.05 -CH3), 2.07 , 2.30 -CH2-), 3.66 , 4.67 , 5.87 . 13C NMR : δ (ppm) = 21.0, 24.9, 28.7, 29.0, 29.3, 29.7, 32.2, 51.4, 65.3, 129.7, 130,9, 170.9, 174.2. MS : m/z (%) = 256 (2) [M+], 213 (3), 196 (3), 182 (23), 164 (31), 154 (8), 135 (14), 122 (14), 107 (8), 98 (15), 81 (29), 67 (24), 55 (34), 43 (100), 29 (9).1H NMR : δ (ppm) = 0.88 , 1.29 , 2.56 , 4.50 , 5.66 . 13C NMR : δ (ppm) = 11.4, 21.0, 22.7, 29.2, 29.2, 29.3, 29.4, 31.9, 32.2, 65.3, 129.8, 136.8, 179.3. MS : m/z (%) = 212 (4) [M+], 170 (5), 152 (3), 141 (4), 124 (12), 110 (8), 96 (19), 82 (26), 67 (25), 54 (31), 43 (100), 39 (11), 29 (14).1H NMR : δ (ppm) = 0.89 , 1.28 , 2.00 , 5.37 . 13C NMR : δ (ppm) = 13.1, 25.9, 26.2, 28.2, 28.5, 28.8, 31.0, 33.4, 129.4. MS : m/z (%) = 252 (5) [M+], 154 (1), 139 (2), 125 (10), 111 (29), 97 (59), 91 (1), 83 (68), 79 (7), 69 (79), 65 (4).1H NMR : δ (ppm) = 1.27 , 1.59 -CH2-CH2-), 1.93 , 2.26 -CH2-), 3.63 , 5.34 . 13C NMR : δ (ppm) = 24.9, 28.9, 29.4, 29.5, 29.6, 32.5, 34.0, 51.3, 130.2, 174.2. MS : m/z (%) = 340 (1) [M+], 308 (7), 290 (3), 276 (16), 265 (1), 207 (1), 165 (7), 151 (11), 133 (12), 121 (13), 109 (18), 95 (38), 81 (59), 74 (44), 67 (58), 55 (100).


To compare the quality of life with patients suffering from JIA, attending school and taught at home.st group , 2nd group - , 2-A , 2B- .Basing on data of 99 patients 14-17 old with JIA boys/girls - 27%/73%, 1- group of children attending school 62 (63%), 2 – 37 patients taught at home; among whom - 20 (group 2A) on medical indications; 17 (group 2B) at will of the patient and parents. 1Questionnaire: SF-36, EQ-5D, HAQ, CHAQ; Kettel, author's method of social activity, Spielberger-Khanin scale; Visual analogue scale (VAS) – assessment of pain, VAS – global evaluation of health, Well-Being Index (WHO-5)VAS of pain .PF 1st group-77, 2nd group-54, 2A-55, 2B-56; RP- 1gr -62, 2 - 48, 2A - 50, 2B - 50); BP: 70, 47, 57, 37; p=0.07), GH: .HAQ, CHAQ: 0.3* ; 0.9; 1.12*: 0.6.p=0.02WHO-5 (Norm >13): .st group -82%; 2nd group - 43%; 2A group - 40%; 2B group 45%Regular communication with equals in age: 1st group - 50%, in the 2nd group, 30%, 2A group - 20%, 2B group -36%.Attendance of cultural events at least once a month in the 1st group - 68%; 2nd group - 80%, 2A-77%, 2B-72%; fears: 26%, 43%, 46%, 54%. Family stress: 71%, 40%, 38%, 36%.Communication disorders: 1The quality of life was higher with children attending school and considerably lower with children groundlessly taught at home.


AbstractProgradungula otwayensis Milledge, 1997 is endemic to the Great Otway National Park  and known from only one male and a few juvenile specimens. In a recent survey we recorded 47 specimens at several localities across the western part of the Great Otway National park. Our field data suggest that this species is dependant on the microclimate in the hollows of old myrtle beech trees since other hollow trees were very much less inhabited. Furthermore, we describe the female for the first time and study the male palpal organ by using X-ray microtomography. The female genitalia are characterized by eight spermathecae which are grouped in two quartets. The spermophor resembles the general organization of gradungulids, but is similar to Kaiya Gray, 1987 by a convoluted appearance within the embolus. The muscle 30 is located in the cymbium and resembles the organization of other non-entelegyne Araneomorphae.The recently described cribellate gradungulid  Gradungulidae consists of seven genera with 16 described species from eastern Australia and New Zealand  and Progradungula  can be distinguished from the remaining gradungulids by being cribellate. These spiders use a very conspicuous catching ladder for prey capture . Nevertheless, only a single adult male and several immatures were reported at that time. In the present paper, we describe the female for the first time and reconstruct the male palpal organ by using X-ray microtomography (micro-CT). Moreover, we provide new data on the natural history of this species.The family  Zealand . The gen capture . The enith Wales  and thusWe observed a total of 47 specimens  in the localities depicted in https://www.morphdbase.de/?P_Michalik_20130802-M-5.1).For the micro-CT analyses of the male palp, the sample was dehydrated in graded ethanol and stained with a 1% iodine solution for 12 hours. After washing in pure ethanol, the sample was scanned in ethanol with an Xradia MicroXCT-200 X-ray imaging system  at 20 KV and 4 W . The female genitalia were digested with enzymatic cleaner for contact lenses Ultrazyme, and dried after the iodine treatment using hexamethyldisilazane (HMDS). The scan was performed using the same system at 30 kV and 6 W . The obtained data were processed using the 3D analysis software AMIRA v. 5.4.2 . The spermophor was reconstructed by delineation of the contours in each section and a smooth surface was computed using the surface editor. The image stack is stored in MorphDBase under creative commons attribution MACN Museum of Victoria MV Zoologisches Institut und Museum Greifswald (Germany)ZIMGMilledge, 1997http://species-id.net/wiki/Progradungula_otwayensis38°42’S, 143°29’E, 31 Jan 1995, G. Milledge .Male holotype: AUSTRALIA: Victoria. Otway Ranges, Aire Crossing Track, 0.5 km N of Aire River crossing, 38.67032°S, 143.49810°E , elev. 330m, 11 Feb 2013, Nothofagus, Eucalyptus, tree ferns wet forest, hand collecting, P.Michalik & M.J.Ramírez (MJR-Loc-125), two females (MACN-Ar 30666); Triplet Falls Trail, E of Lavers Hill, 38.67188°S, 143.49673°E , elev. 300m (GPS), 10 Feb 2013, Nothofagus, Eucalyptus, tree ferns wet forest, hand collecting, P.Michalik & M.J.Ramírez (MJR-Loc-124), three females ; Maits Rest Trail, W Apollo Bay, 38.75492°S, 143.55495°E , elev. 240m, 13–14 Feb 2013, Nothofagus, tree ferns wet forest, hand collecting, P.Michalik & M.J.Ramírez (MJR-Loc-126), one female (ZIMG II/28130); Melba Gully Trail, 38°41.726’S, 143°22.312’E , elev. 328m, 15 Feb 2013, Nothofagus, tree ferns wet forest, hand collecting, P.Michalik & M.J.Ramírez (MJR-Loc-128), one male (ZIMG II/28127).AUSTRALIA: VICTORIA: Great Otway National Park: Little Aire Cascade Trail, E of Lavers Hill, Progradungula carraiensis by the single process on the embolus of the male palpal organ and the presence of eight spermathecae in the female genitalia.This species can be distinguished from –4.PageBreakPageBreak: I 11.97 + 14.10 + 11.57 + 2.66 = 40.30, II 9.31 + 10.77 + 8.25 + 2.53 = 30.86, III 7.98 + 9.18 + 7.85 + 2.13 = 27.14, IV 9.98 + 10.91 + 9.44 + 2.13 = 32.46, palp 2.93 + 2.66 + - + 2.79 = 8.38.. Carapace length 6.92; carapace width 4.66; caput width 2.66; abdomen length 8.51; abdomen width 7.18. Legs: length of segments Carapace yellowish brown, darker on cephalic area, particularly in eye region. Chelicerae, maxillae and labium reddish brown. Sternum reddish brown marginally, paler centrally. Legs yellowish brown. Abdomen fawn-coloured with dark brown dorsal pigmentation consisting of 3 chevron markings. Carapace longer than wide, widest between coxae II and III. Cephalic area elevated behind eyes . ClypeusFrontal view : anterioStrong, vertical, slightly divergent. Three strong prolateral teeth, evenly spaced. Five very small retromarginal teeth (or denticles) in row on basal half of groove, with a spine on the apico-dorsal side. Stridulatory ridges absent.Subparallel, external margin strongly curved and ending anteriorly in a bluntly pointed apex . SerrulaFree. Length 0.83, width 0.87. Apical margin indented, lateral margin subparallel below, sloping in toward apex above .Length 3.33, width 2.20. Elongate, shield-shaped with pointed apex which extends back midway between coxae IV .1423. Trochanters shallowly notched. Superior claws of 1st and 2nd legs dissimilar, with raptorial proclaws long and strongly developed , retrocl Femur I d (r2p1)-r2-r1-p1-1-2-p1-2-p1-2-2-3ap v 0-p1-0-0-0; patella r 1; tibia d 2-0-2-p1-0-p1-3 v 2-0-0-2-p1-0-2-0; metatarsus d r1-p1-p1-0-0-0-0 v p1-r1-p1-p1-r1-p1-r1-2ap; II; femur d (r2p1)-r2-2-p1-2-p1-2- 2-p1-3 v 0-p1-1-0-0; patella p 1; tibia d 2-p1-1-2-2-p1-p1-1-3 v 2-0-p1-r1-p1-2-0-2ap; metatarsus d 2-p1-2-r1-p1-r1-p1-p1-2ap v p1-r1-2-2-r1-p1-3ap; III; femur d (r3p1)-1-r1-2-p1-2-2-2-r1-3 v r1-p1-2-0-0; patella p 1 d 1 r 1; tibia d 2-1-p1-3-2-2 v 2-0-p1-r1-p1-2ap; metatarsus d 2-p1-p1-r1-p1-2- 2ap v 2-p1-p1-r1-p1-p1-r1-3ap; tarsus v 0-r1-0; IV femur d (r4p2)-r2-1-r2-p1-2-2-2-3 v 0-p1-p1-0-0-0; patella p 1 d 1; tibia d 2-2-p1-1-3-r1 v 2-0-0-p1-p1-2ap; metatarsus d 2-p1-2-r1-2ap v p1-2-p1-p1-p1-2-r1-3ap; tarsus v 0-1-0.Single claw. Spines: femur d 0-0-1-1-3ap patella p 1 d 1-1, tibia d 1-1, p 0-2-0; tarsus d 2-2-1-2-3ap v 2-2-2ap.PageBreakPageBreakCribellum undivided, with narrow spinning field, as wide as basal span of anterior spinnerets . SpiraclGenital area distinct and dome-shaped , 4A. Int trees, the bases of large tree ferns . In addition to the locations reported by PageBreakPageBreakProgradungula species  and their tendons are in accordance with the description of Gradungula sorenseni Forster, 1955 by Araneomorphae (Gradungulidae and different to the organization found in austrochilines (As revealed by micro-CT analysis of the male palpal organ of omorphae . Thus, tchilines .


Ficus tikoua Bur. was investigated. One new antifungal pyranoisoflavone, 5,3′,4′-trihydroxy-2″,2″-dimethylpyrano  isoflavone (1), together with two known isoflavones, wighteone (2) and lupiwighteone (3) , were isolated from ethyl acetate extract by bioassay-guided fractionation. Their structures were determined by spectroscopic analysis, such as NMR , IR, UV and HRMS, as well as ESI-MSn analyses. The antifungal activities of 1–3 against Phytophthora infestans were evaluated by direct spore germination assay, and the IC50 values were 262.442, 198.153 and 90.365 μg·mL−1, respectively.Considering the undesirable attributes of synthetic fungicides and the availability of Ficus species in China, the stem of  Ficus tikoua Bur., a wood plant of the Ficus genus, is widely distributed in southern China, India, Vietnam and Laos. It has long been used in traditional folk medicine to treat diseases, such as chronic bronchitis, diarrhea, dysentery, rheumatism, edema, impetigo, and so on D +19.0° , IR : 3410, 1686, 1580; 1H-NMR  δ 8.01 , 7.03 , 6.68 , 6.39 , 6.36 , 6.37 , 5.71 , 1.45 ; 13C-HMR  δ 156.85 (CH-2), 122.66 (C-3), 182.84 (CO-4), 157.58 (C-5), 95.86 (CH-6), 160.90 (C-7), 106.56 (C-8), 158.87 (C-9), 106.93 (C-10), 110.63 (C-1′), 133.23 (CH-2′), 160.25 (C-3′), 157.78 (C-4′), 104.23 (CH-5′), 108.13 (CH-6′), 79.25 (C-2″), 129.67 (CH-3″), 116.09 (CH-4″) and 28.55 . ESI-MS/MS: m/z (%) 351 (30) [M-H]−, 333 (20) [M-H-H2O]−, 323 (35) [M-H-CO]−, 307 (100) [M-H-CO2]−, 265 (40) [M-H-C4H6O2]−, 217 (100) [M-H-C8H6O2]−, 201 (30) [M-H-C8H6O3]−. High-resolution ESI-MS: m/z [M + H]+ calcd for C20H17O6 353.1020; found 353.1010.5,3′,4′-trihydroxy-2″,2″-dimethylpyrano isoflavone (1), was isolated using bioactivity-guided bioassay-guided fractionation. The in vitro antifungal activity of 1 against P. infestans was preliminarily evaluated using the spore germination assay. The IC50 value of 1 was 262.442 μg·mL−1, which showed the equivalent antifungal activity as a reference fungicide, pyrimethanil with an IC50 value 210.050 μg·mL−1.In summary, a new isoflavone derivative, 5,3′,4′-trihydroxy-2″,2″-dimethylpyrano  isoflavone (


Reactions of N-(2-substituted-aryl)-3-hydroxyisoindolinones with N-vinylpyrrolidone under identical conditions resulted in the formation of 2-(2-substitued-aryl)-3-(2-(2-oxopyrrolidin-1-yl)vinyl)isoindolin-1-one analogues indicating steric hinderance as the cause of deviation. The probable mechanism of the reaction based on the results from X-ray crystallography and molecular modelling is discussed.The synthesis of 5-substituted 6,6a-dihydroisoindolo[2,1- Our literature search also infuses confidence in the proposed scheme because the synthesis of N-aryl-3-hydroxyisoindolinones is well studied and standardized quinolin-11(5H)-one derivatives were synthesized from N-aryl-3-hydroxyisoindolinones and N-vinyl lactams under Lewis acid-catalysed anhydrous conditions in fair to good yields. As expected, the reaction appeared to proceed via the formation of an N-acyliminium ion which underwent an inverse-electron demand imino-Diels–Alder reaction. While reactions of this type are known to occur, this is the first report of the use of N-vinyl lactams as a dienophile for the inverse-electron demand imino-Diels–Alder reaction with N-acyliminium cations as dienes. The most important aspect of this investigation is the dissimilar outcome of the reactions of N-(2-substituted-aryl)-3-hydroxyisoindolinones with N-vinylpyrrolidone under identical conditions which resulted in the electrophilic addition of an N-acyliminium ion on the vinyl group followed by β-elimination leading to 2-(2-substitued-aryl)-3-(2-(2-oxopyrrolidin-1-yl)vinyl)isoindolin-1-one analogues. This indicated that the ortho substituent on the N-acyliminium ion became detrimental for cyclization to occur.6,6a-Dihydroisoindoloquinolin-11(5H)-ones (1a–h); 5-(2-oxoazepan-1-yl)-6,6a-dihydroisoindoloquinolin-11(5H)-ones (2a–h) and (E)-2-(2-substitued-phenyl)-3-(2-(2-oxopyrrolidin-1-yl)vinyl)isoindolin-1-ones (3a–i)N-aryl-3-hydroxyisoindolinones (500 mg) from the previous step were dissolved in dichloromethane (10 mL) in a 50 mL round bottom flask which was then sealed with a rubber septum. BF3:Et2O (1.5 equiv) was added to the mixture slowly through the septum by a syringe. This resulted in the formation of a transparent solution. Appropriate tert-enamide (1.5 equiv) dissolved in dichloromethane (5 mL) was slowly added to the flask through a syringe over a period of 5 minutes. The reaction was then allowed to stir at room temperature until precipitates formed. The reaction progress was monitored by TLC. Upon completion of the reaction, the residue was suction filtered and the solid obtained was extracted with ethyl acetate (60 mL) and washed with water (30 mL × 3). The organic layer was dried over anhydrous Na2SO4 and evaporated in vacuum to dryness to obtain pure products. In the cases where precipitates did not form, the solvents were evaporated to dryness under reduced pressure. The residue was extracted by ethyl acetate (60 mL) and washed with water (30 mL × 3). The organic layer was dried over anhydrous Na2SO4 and evaporated in vacuum to dryness. The purification of the crude product was achieved by column chromatography . Characterization data for one representative compound from each series is presented here.Appropriate a]quinolin-11(5H)-one (1a): Yield: 56%. White solid. mp: 225–227 °C; 1H NMR (CD3OD) δ 1.68–1.84 , 1.98–2.14 , 2.57–2.65 , 2.68–2.78 , 3.03–3.18 , 3.22–3.32 , 5.04 , 5.73–5.75 , 7.10–7.25 , 7.32 , 7.50–7.70 , 7.83 , 8.45 ; 13C NMR (CD3OD) δ 18.9, 32.0, 32.2, 44.3, 50.0, 59.7, 121.5, 123.5, 124.7, 125.2, 125.7, 127.9, 129.3, 130.0, 133.0, 133.8, 137.8, 145.6, 167.8, 178.9; FTIR νmax (NaCl): 3051, 2953, 2921, 1689, 1490, 1456, 1380, 1282, 1213, 1094, 906, 759, 730 cm−1; HRMS m/z: [M + Na]+ calcd for C20H18N2O2Na, 341.1260; found, 341.1252.5-(2-Oxopyrrolidin-1-yl)-6,6a-dihydroisoindoloquinolin-11(5H)-one (2a): Yield: 47%. White solid. mp: 228–230 °C; 1H NMR (CD3OD) δ 1.30–1.40 , 1.48–1.90 , 2.50–2.67 , 2.68–2.85 , 2.90–3.34 , 5.03 , 6.15–6.23 , 7.15 , 7.28–7.37 , 7.52–7.60 , 7.68 , 7.85 , 8.43 ; 13C NMR (CD3OD) δ 24.5, 30.3, 30.7, 33.2, 38.1, 46.3, 53.0, 60.0, 121.7, 123.4, 124.8, 125.6, 126.5, 128.5, 129.1, 130.0, 133.2, 133.8, 138.4, 146.0, 167.8, 179.4; FTIR νmax (NaCl): 3047, 2929, 2851, 1695, 1683, 1597, 1489, 1454, 1380, 1198, 1070, 759 cm−1; HRMS m/z: [M + Na]+ calcd for C22H22N2O2Na, 369.1573; found, 369.1560.5-(2-Oxoazepan-1-yl)-6,6a-dihydroisoindolo+ calcd for C20H17FN2O2Na, 359.1166; found, 359.1149.(File 11a, 1h, 2b and 3a.Synthetic procedure and characterization data for all compounds and crystallographic data for


ZJ9-6B, isolated from the mangrove Aegiceras corniculatum collected in the South China Sea. Their structures were elucidated by one- and two-dimensional NMR spectroscopy, MS data analysis and circular dichroism measurements. Compounds 1, 2 and 3 were first isolated alterporriols with a C-2–C-2′ linkage. The crystallographic data of tetrahydroaltersolanol B (7) was reported for the first time. In the primary bioassays, alterporriol K and L exhibited moderate cytotoxic activity towards MDA-MB-435 and MCF-7 cells with IC50 values ranging from 13.1 to 29.1 μM.Three new bianthraquinone derivatives, alterporriol K ( AnthrAlternaria porri by Suemitsu et al. in 1984 . ,13. 1 waelations . In the 1H-1H COSY spectrum combined with the HMBC correlations from H-5 to C-6, C-7 and C-8a, from H-6 to C-5, C-8, and C-10a, and from H-7 to C-5, C-8, C-8a, C-9 and C-11, established the substructure of the cyclohexene ring  and 4′-OH (δH = 13.63 ppm) to C-4, C-3, C-4a and to C-4′, C-3′, C-4′a, respectively, indicated their location at C-4 and C-4′ of the bianthraquinone scaffold.HMBC correlations from each of 4-OH  and H-3′ . Moreover, HMBC correlations of H-3 with C-1, C-2, C-2′, C-4, C-4a and C-10, and of H-3′ with C-1′, C-2, C-2′, C-4′, C-4′a and C-10′, respectively  with C-1 and C-3. Likewise, correlations were observed from methoxyl proton  to C-1′. These allowed us to assign two methoxyl groups to C-1 and C-1′.The presence of a bond connecting C-2 and C-2′ was suggested by the absence of two sets of ectively  provided1H-1H NOESY experiments  and H-5 suggested that they were on the same side of the cyclohexene ring. The axial position of H-5 was confirmed by coupling constant JH-5,H-6a = 12.5 Hz, indicating CH3-11 to also have an axial orientation. Therefore, compound 1 was determined as -4,4′,5,7′,8-pentahydroxy-1,1′-dimethoxy-6′,8-dimethyl-5,6,7,8-tetrahydro--9,9′,10,10′-tetraone. We propose the trivial name alterporriol K.The relative configuration of the chiral centers of C-5 and C-8 were deduced by 2D eriments  and the onstants . A NOESY2 and 3 were obtained as a mixture after separating with Sephadex LH-20 chromatography. Upon HPLC analysis, the mixture exhibited two partially overlapped peaks (area ratio ca. 4:1). Then compounds 2 and 3 were isolated with re-separation by preparative HPLC, respectively. Compound 2 was a red amorphous powder, c = 1.0, MeOH). The HR-ESI-TOF-MS exhibited a peak at m/z = 601.1340 [M – H]− indicating a molecular formula of C32H26O12 . Comparison of the 1H and 13C NMR spectral data of 2  and an additional hydroxyl group signal  and the absence of the methylene signals corresponding to H-7 in the 1H NMR spectrum of 2. The substructure of the cyclohexene ring was established by HMBC correlations from H-8 to C-6, C-7, C-8a, C-9, C-10a and C-11 and 1H-1H COSY correlations between H-5 and H-6  was irradiated, no enhancement of signals H-6 and H-8 was observed. Meanwhile, irradiation of H-8 caused an enhancement of H-6. These data suggested that CH3-11 with H-6 and H-8 was trans-configuration in the cyclohexene ring  indicated an axial location for H-6, likewise suggesting an axial location for H-8. Thus, the relative configuration at C-6, C-7, and C-8 was 6S*, 7R* and 8R*. Compound 2 was finally defined as -4,4′,6,7,7′,8-hexahydroxy-1,1′-dimethoxy-6′,7-dimethyl-5,6,7,8-tetrahydro--9,9′,10,10′-tetraone, named as alterporriol L.Compounds ata of 2  with tha and H-6 . NOE difonstants  enabled ene ring . A large3, obtained as a red amorphous power, c = 1.0, MeOH), had the molecular formula C32H26O12, as determined by HR-ESI-TOF-MS at m/z = 601.1340 [M – H]− . The MS analysis revealed 3 to be an isomer of 2. The UV and IR absorptions, 1H and 13C NMR data, HMBC and 1H-1H COSY correlations  resulted in obvious enhancements of the signals for both H-6 and H-8. The coupling constant for H-5a/H-6 was measured to be JH-5a,H-6 = 9.9 Hz suggesting axial locations for H-6 and, by extension, H-8. These data suggested that 2 and 3 are epimers at C-7. Therefore compound 3 was elucidated as -4,4′,6,7,7′,8-hexahydroxy-1,1′-dimethoxy-6′,7-dimethyl-5,6,7,8-tetrahydro--9,9′,10,10′-tetraone, named alterporriol M.Compound elations  of 3 werAb initio calculations of CD spectra for several phenylanthraquinones with chiral axes have recently been reported +.Alterporriol L: Red powder. c = 1.0, MeOH). UV (MeOH): λmax (log ε) = 280.20 (1.24), 225.40 (1.61) nm. IR (KBr): νmax = 3440, 2926, 2857, 1592, 1462, 1430, 1389, 1288, 1208, 1109, 1070, 974, 927, 846, 793, 605 cm−1. For 1H, 13C and 2D NMR spectroscopic data, see 32H25O12, 601.1346; found m/z = 601.1340 [M – H]−.Alterporriol M: Red powder. c = 1.0, MeOH). UV (MeOH): λmax (log ε) = 279.00 (1.33), 225.60 (1.77) nm. IR (KBr): νmax = 3450, 2928, 2027, 1638, 1463, 1389, 1280, 1207, 1111, 1046, 977, 929, 858, 618 cm−1. For 1H, 13C and 2D NMR spectroscopic data, see m/z = 601.1 [M – H]−. HR-ESIMS calcd. for C32H25O12, 601.1346; found m/z = 601.1340 [M – H]−.Crystallographic data of 7: The X-ray diffraction data for 7  Å, b = 5.8251(10) Å, c = 7.9087(13) Å, α = 90.00°, β = 102.210(3)°, γ = 90.00°; V = 1679.8(5) Å3; Density: 1.397 g/cm3; Z = 2, F (000) = 756.0; Goodness-of-fit on F2: 1.087; R Indices : R1 = 0.0399, wR2 = 0.1208. CCDC-751689 contains the supplementary crystallographic data for this paper. These data can be obtained free of charge via www.ccdc.cam.ac.uk/conts/retrieving.html .ta for 7  was meas3.4.1 and 2 were evaluated using human breast cancer cell lines MDA-MB-435 and MCF-7 by MTT assay. Cell viability was measured using the CellTiter 96 aqueous nonradioactive cell proliferation assay. Results were expressed as the mean value of triplicate data points.The cytotoxicity of compounds 4.Alternaria sp. ZJ9-6B is a prolific producer of bioactive metabolites. Nine compounds have been isolated from this fungal strain, including three new alterporriols. These three alterporriols all possess dimeric structures with a C-2–C-2′ linkage. In the primary bioassay, compounds 1 and 2 showed moderate cytotoxic activity against human breast cancer cell lines.


In addition, peptide fragments from solution phase synthesis can be purified, and the pure intermediates are elaborated to the desired target peptide. Also, the smaller fragments used to make larger peptides give way for easier purifications (by size exclusion chromatography) of the target peptides by taking advantage of their size differences. Here, we report the synthetic routes to hemopressin and other related peptides using this approach. The protecting groups  used her-OH (15) , the N-tut)-(28) , and the4) was prepared in two steps by condensation of 1 with 2 followed by hydrogenolysis of the resulting intermediate 3 -Pro-Val-OH (11), was accomplished by hydrogenolysis. For the synthesis of the third key N-terminal pentapeptide, Fmoc-Arg(Pbf)-Val-Asp(OBut)-Pro-Val-OH (15), intermediate 10 was converted to 12 by exposure to piperidine followed by coupling with Fmoc-Arg(Pbf)-OH (13) to give 14, which on hydrogenolysis afforded 15.The dipeptide fragment Boc-Pro-Val-OH (ediate 3 . Preparasy steps . Brieflyt)-)-His(Trt)-OBut)-(28), which is common to all target peptides was assembled as outlined in 18) and H-Lys(Boc)-AA-OBn (21), where AA is either Phe or Leu, were prepared from condensations of 16 with 17 and 19 with 20, respectively. Further, condensation of dipeptide 18 with dipeptide 21 provided tetrapeptide 22 after hydrogenolysis. Elaboration of 22 to pentapeptide 24 was accomplished by addition of C-terminal residue 23 followed by hydrogenolysis (t)-)-His(Trt)-OBut)-(27), was assembled by coupling Cbz-Ser(But)-)-OH (25) with HCl·His(Trt)-OBut)-(26) followed by removal of the Cbz group. The final steps in the assembly of C-terminal heptapeptide fragment 28 involve the (5 + 2) fragment condensation of pentapeptide 24 with dipeptide 27 followed by exposure to piperidine.Next, the final C-terminal heptapeptide fragment, H-Asn(Trt)-Phe-Lys(Boc)-Phe/Leu-Leu-Ser(Buenolysis . The dip4 was condensed with heptapeptide fragment 28 under HATU/HOAt/DIEA coupling conditions . The crude peptide was purified to homogeneity by preparative reversed phase HPLC. The HPLC conditions were as follows: Vydac C18 column (218TP1022); flow rate: 15 mL/min; using a gradient (10% B → 65% B over 30 min) where A = 0.1% TFA/H2O and B = 0.1% TFA/CH3CN; UV detection 220 nm.] All target peptides: TFA·Pro-Val-Asn-Phe-Lys-Phe-Leu-Ser-His-OH (rHP) (30a), TFA·Pro-Val-Asn-Phe-Lys-Phe-Leu-Ser-His-OH (hHP) (30b), TFA·Val-Asp-Pro-Val-Asn-Phe-Lys-Phe-Leu-Ser-His-OH (VD-hHP) (32), and TFA·Arg-Val-Asp-Pro-Val-Asn-Phe-Lys-Phe-Leu-Ser-His-OH (RVD-hHP) (34), were purified to homogeneity by preparative reversed phase HPLC and characterized by TLC, HPLC, MS (ESI), and amino acid analysis. D22 −26° ; amino acid analysis: found  Pro, 1.10 (1.00); Val 0.96 (1.00); Asn, 0.72 (1.00); Phe, 2.00 (2.00), Lys, 0.61 (1.00), Leu, 1.30 (1.00), Ser, 1.13 (1.00), His, 0.26 (1.00); (b) hHP: MS (ESI) m/z 1055 (M+H); [α]D22 −5.3° ; amino acid analysis: found  Pro, 1.03 (1.00); Val 0.96 (1.00); Asn, 1.00 (1.00); Phe, 0.84 (1.00), Lys, 1.03 (1.00), Leu, 2.03 (2.00), Ser, 0.94 (1.00), His, 0.97 (1.00); (c) VD-hHP: MS (ESI) m/z 1269.1 (M+H), m/z 635.5 (M+H)+2; [α]D22 −21° ; amino acid analysis: found  Pro, 1.00 (1.00); Val 1.90 (2.00); Asx, 2.13 (2.00); Phe, 1.00 (1.00), Lys, 1.06 (1.00), Leu, 1.96 (2.00), Ser, 0.89 (1.00), His, 0.91 (1.00); (d) RVD-hHP: MS (ESI) m/z 1424.81 (M+H), m/z 712.90 (M+H)+2, m/z 475.60 (M+H)+3; [α]D22−35° ; amino acid analysis: Found  Pro, 0.90 (1.00); Val 1.90 (2.00); Asx, 2.2 (2.00); Arg, 0.80 (1.00); Phe, 1.10 (1.00), Lys, 1.20 (1.00), Leu, 1.90 (2.00), Ser, 0.80 (1.00), His, 1.00 (1.00).]  Similarly, VD-hemopressin (ectively  involvin


Musa acuminata, Cavendish cultivar). In banana peels, chlorophyll catabolites were found in an unprecedented structural richness: a variety of new fluorescent chlorophyll catabolites (FCCs) and nonfluorescent chlorophyll catabolites (NCCs) were detected. As a rule, FCCs exist only "fleetingly" and are hard to observe. However, in bananas several of the FCCs (named Mc-FCCs) were persistent and carried an ester function at the propionate side-chain. NCCs were less abundant, and exhibited a free propionic acid group, but functional modifications elsewhere. The modifications of NCCs in banana peels were similar to those found in NCCs from senescent leaves. They are presumed to be introduced by enzymatic transformations at the stage of the mostly unobserved, direct FCC-precursors. The observed divergent functional group characteristics of the Mc-FCCs versus those of the Mc-NCCs indicated two major "late" processing lines of chlorophyll breakdown in ripening bananas. The "last common precursor" at the branching point to either the persistent FCCs, or towards the NCCs, was identified as a temporarily abundant "secondary" FCC. The existence of two "downstream" branches of chlorophyll breakdown in banana peels, and the striking accumulation of persistent Mc-FCCs call for attention as to the still-elusive biological roles of the resulting colorless linear tetrapyrroles.The disappearance of chlorophyll is a visual sign of fruit ripening. Yet, chlorophyll breakdown in fruit has hardly been explored; its non-green degradation products are largely unknown. Here we report the analysis and structure elucidation of colorless tetrapyrrolic chlorophyll breakdown products in commercially available, ripening bananas ( Breakdown of chlorophyll is a major contributor to the diagnostic color changes in fall leaves. De-greenHordeum vulgare), named Hv-NCC-1.-Mc-FCC-71, in which a trideutero-methyl ester was present. In addition, 1H atoms at C1 and C20 were also partially replaced by 2 H. Mc-FCC-71 was thus indicated to represent an isolation artifact and to be formed in the extract by a trans-esterification of hmFCCs (such as Mc-FCC-56) by MeOH.The least polar FCC named Mc-NCCs) in extracts of the peels of ripe bananasNonfluorescent catabolites  and classified tentatively, based on their UV spectra. Seven of the Mc-NCCs were isolated by preparative HPLC, and further analyzed by mass spectrometry and NMR spectroscopy .Mc-NCC-26 was the most polar NCC identified in extracts from yellow banana peels. From mass spectrometric analysis, the molecular formula of Mc-NCC-26 was deduced as C34H40N4O10, providing the basis for a tentative structure assignment. Mc-NCC-26 was indicated to have no ester function at C(132), but a 31,32-dihydroxyethyl group (at C3) and a 82-hydroxyethyl group  from 65 bananas; see Mc-NCC-42 to have the molecular formula of C35H42N4O10. The molecular constitution of Mc-NCC-42 was elucidated with the help of homo- and hetero-nuclear NMR-correlations -phytoporphyrin, and was revealed to carry 31,32-dihydroxyethyl and 82-hydroxyethyl groups, respectively, as side chains at C-3 and C-8 of the NCCs tetrapyrrole core .+; found: 645.314. See Mc-FCC-62.Mc-FCC-62: Mc-FCC-71: UV/Vis: λmax =262 , 317 (1.00), 359 nm (0.65). For 1H- and 13C NMR and ESI-MS, see the Supporting Information.Mc-NCC-61: tR=61 min; 1H NMR : δ=1.93 ), 1.96 ), 2.13 ), 2.25 ), 2.33 ), 2.48 ), 2.62  superimposed by H2C(81)), 2.71 ), 2,85 ), 3.48 ), 3.75 ), 3.94 ) superimposed by 3.95 ), 5.35 ), 6.09 ), 6.44 ), 9.34 ppm ); 13C NMR : δ=111.1 (12), 114.1 (18), 119.3 (17), 119.4 (8), 123.4 (19), 124.8 (13), 127.6 (3), 128.2 (6), 132.8 (11), 155.7 (2), 170.4 ppm (133); UV/Vis: λmax =245 (1.00), 313 (0.89); MS : m/z (rel. abundance)=699.20 , 683.19 , 667.21 (27 [M+Na]+), 646.19 (40), 645.21 , 613.19 , 522.10 . HR-FAB (matrix NOBA): m/z calcd for C39H41N4O8: 645.292 [M+H]+; found: 645.291.Mc-NCC-59: (tR=59 min); UV/Vis ): λmax (εrel)=243 (1.00), 314 (0.87). MS (nano-HPLC-ESI): m/z (rel. abundance)=829.33 , 809.35 (14), 808.35 (48), 807.35 , 775.32 , 732.22 (6), 684.22 , 661.21 (6), 645.22 , 613.17 ; HR-FAB matrix: glycerol: m/z calcd for C41H51N4O13+: 807.345 [M+H]+; found: 807.342.Mc-NCC-58: (tR=58 min); UV/Vis: λmax =242 (1.00), 314 (0.82). CD : λmax/nm(Δε)=222 (21.2), 245 (−5.4), 283 (−12.0), 316 (6.3); ESI-MS: m/z (%): 945.23 , 929.30 , 907.25 , 891.32 , 871.31 (29), 870.28 (57), 869.31 , 854.32 (23), 853.37 , 833.28 (14), 832.30 (42), 831.37 .Mc-NCC-55: (tR=55 min); UV/Vis: λmax=242 (1.00), 314 (0.84). CD : λmax/nm (Δε)=230 (−16.1), 283 (6.9), 320 (−5.0); MS (ESI): m/z (%)=891.32 , 869.31 , 853.31 (30 [M+Na]+), 833.39 (19), 832.35 (55), 831.31 , 705.25 (13), 683.25 (13), 667.21 .Mc-NCC-49: (tR=49 min): 1H NMR : δ=1.92 ), 1.94 ), 2.10 ), 2.24 ), 2.37 ), 2.45 ), 2.63 ), superimposed by 2.66 ), 2.78 ), 2,83 ), 3.46 ), 3.92 ), 3.98 ), 5.34 ), 6.08 ), 6.42 ), 9.34 ppm ); UV/Vis: λmax =242 , 313 nm (0.90); MS (nano-HPLC-ESI): m/z (rel. abundance)=677.23 (2), 659.18 83), 633.18 (6), 632.18 (17), 631.17 , 613.15 , 609.16 (6), 588.17 (20), 587.16 , 508.06 , 464.11 (3), 424.96 (2); HR-FAB matrix: glycerol: m/z calcd for C34H39N4O8+: 631.276 [M+H]+; found: 631.279.Mc-NCC-42: (tR=42 min): UV/Vis ): 1H NMR : δ=1.93 ), 2.02 ), 2.08 ), 2.26 ), 2.32 ), 2.54 ), 2.62 ) superimposed by 2.63 ), 2.72 ), 2,87 ), 3.47 ), 3.62 ), 3.69 ), 3.75 , 3.97 ), 4.01 ), 4.56 ), 9.36 ppm ). 13C NMR : δ=8.93 (71), 9.16 (121), 9.31 (181), 12.4 (21), 21.9 (171), 23.8 (10), 28.1 (81), 29.7 (20), 37.2 (15), 39.1 (171), 52.8 (135), 62.4 (1), 62.5 (82), 65.9 (32), 68.6 (31), 113 (12), 115 (18), 120 (17), 121 (8), 124 (16), 124 (19), 126 (13), 129 (4), 132 (3), 135 (7), 135 (11), 139 (9), 159 (2) ppm (see λmax (εrel)=248 (0.77), 319 nm (1.00); Nano-HPLC-ESI-MS: m/z (rel. abundance)=718.19 (8), 717.23 , 702.21 (15), 701.22 (39 [M+Na]+), 681.26 (11), 680.25 (40), 679.24 , 648.21 (18), 647.18 , 522.12 ; HR-FAB matrix: glycerol: m/z calcd for C35H43N4O10+: 679.297 [M+H]+; found: 679.300. ppm see ; λmax (εMc-NCC-26: (tR=26 min): UV/Vis ): λmax (εrel)=246 , 316 nm (1.00); FAB-MS: m/z (rel. abundance)=666.14 (40), 665.10 .Citric acid induced isomerization ofMc-FCC-56: Aqueous citric acid  was added to a solution of Mc-FCC-56 (3 mL of a 30.9 μm solution) and Mc-FCC-53 (3:1 mixture of FCC-isomers) in MeOH. The mixture was stored for 159 h under an Ar atmosphere in the dark at 15 °C. Water (1 mL) then was added and the resulting solution was subjected to analytical HPLC. Complete disappearance of FCCs and appearance of two major NCCs were indicated . The two most abundant NCCs (Mc-NCC-55 and Mc-NCC-58) were isolated by semi-preparative HPLC and their mass spectra, CD spectra .ctra see  and UV/VCo-elution experimentsMc-NCC-42 withHv-NCC-1 andSo-NCC-2: The most abundant NCC fraction found in extracts of yellow-banana peels, Mc-NCC-42, was isolated and characterized (as described above). The purified Mc-NCC-42 sample was applied to analytical HPLC, as were 1:1 mixtures of Mc-NCC-42 with Hv-NCC-1[So-NCC-2,[Mc-NCC-42 (tR=42) with So-NCC-2 (tR=42), whereas it showed Mc-NCC-42 to differ from Hv-NCC-1 (tR=40) (tR=retention time (in min); see the Supporting Information, Figure S6). Hv-NCC-1 and So-NSo-NCC-2, respectiAnalysis of FCCs in ripening bananas during storage: Ethylene-ripened green–yellow bananas (at rs=5) were stored at ambient temperature and a normal daylight night rhythm. Banana peels were extracted at day zero, one, two, three and four. The extracts were produced from 6–8 cm2 of the peel, ground with liquid nitrogen in a mortar. MeOH (500 μL) and potassium phosphate buffer  were added in the mortar. The mixture was then ultra-filtrated ; 150 μL of the filtrate were applied to the analytical HPLC system. Relative amounts of FCCs were determined by integration of the peaks of the luminescence trace resulting from an FCC; the area was normalized according to the area of peel used for extraction (and was averaged over three to five experiments, see