Conventional hydrogen-bonding interactions lead to supramolecular tapes in the crystal.The 4-pyridyl residues lie to either side of the central, planar C 14H14N4O2·2C7H6O2, are described. The oxalamide molecule has a (+)-antiperiplanar conformation with the 4-pyridyl residues lying to either side of the central, almost planar C2N2O2 chromophore (r.m.s. deviation = 0.0555 Å). The benzoic acid molecules have equivalent, close to planar conformations [C6/CO2 dihedral angle = 6.33 (14) and 3.43 (10)°]. The formation of hydroxy-O—H⋯N(pyridyl) hydrogen bonds between the benzoic acid molecules and the pyridyl residues of the diamide leads to a three-molecule aggregate. Centrosymmetrically related aggregates assemble into a six-molecule aggregate via amide-N—H⋯O(amide) hydrogen bonds through a 10-membered {⋯HNC2O}2 synthon. These are linked into a supramolecular tape via amide-N—H⋯O(carbonyl) hydrogen bonds and 22-membered {⋯HOCO⋯NC4NH}2 synthons. The contacts between tapes to consolidate the three-dimensional architecture are of the type methylene-C—H⋯O(amide) and pyridyl-C—H⋯O(carbonyl). These interactions are largely electrostatic in nature. Additional non-covalent contacts are identified from an analysis of the calculated Hirshfeld surfaces.The crystal and molecular structures of the title 1:2 co-crystal, C Indeed, in early studies on crystal engineering, the combination of bifunctional nLH2 with dicarboxylic acids such as bis(carboxymethyl)oxalamide -antiperiplanar conformation where the 4-pyridyl residues lie to either side of the central C2N2O2 chromophore. The six atoms comprising the central residue are close to co-planar with their r.m.s. deviation equal to 0.0555 Å, with the maximum deviations to either side of the plane being 0.0719 (5) and 0.0642 (5) Å for the N2 and O2 atoms, respectively; the C6 and C9 atoms lie 0.1908 (14) and 0.0621 (14) Å out of and to one side of the plane (towards the N2 atom), respectively. The N1- and N4-pyridyl rings form dihedral angles of 86.00 (3) and 83.34 (2)°, respectively, with the plane through the C2N2O2 atoms, so are close to perpendicular to the central plane. The dihedral angle between the pyridyl rings is 33.60 (5)°, indicating a splayed disposition as each pyridyl ring is folded away from the rest of the molecule. The carbonyl groups are anti and the molecule features intramolecular amide-N—H⋯O(carbonyl) hydrogen bonds that complete S(5) loops, Table 1The molecular structures of the three constituents comprising the crystallographic asymmetric unit of (I)2 group being 6.33 (14) and 3.43 (10)° for the O3- and O5-benzoic acid molecules, respectively. As expected, the C15—O3(carbonyl) bond length of 1.2162 (13) Å is significantly shorter than the C15—O4(hydroxy) bond of 1.3197 (13) Å; the bonds of the O5-benzoic acid follow the same trend with C22—O5 of 1.2237 (13) Å compared with C22—O6 of 1.3084 (13) Å.There are two independent benzoic acid molecules in (I)via amide-N—H⋯O(amide) hydrogen bonding, which leads to a centrosymmetric ten-membered {⋯HNC2O}2 synthon. The second amide forms an amide-N—H⋯O(carbonyl) bond with the result of that adjacent six-molecule aggregates are connected into a supramolecular tape via 22-membered {⋯HOCO⋯NC4NH}2 synthons, Fig. 2a). The other notable contact within the tape is a pyridyl-C—H⋯O(carbonyl) interaction, which cooperates with a hydroxy-O—H⋯N(pyridyl) hydrogen bond to form a seven-membered {⋯OCOH⋯NCH} pseudo-heterosynthon; no analogous interaction is noted for the O5-benzoic acid. The supramolecular tapes are aligned along the c-axis direction and have a linear topology.As anticipated from the chemical composition, significant conventional hydrogen bonding is noted in the crystal of (I)i.e. methylene-C—H⋯O(amide) and pyridyl-C—H⋯O(carbonyl), the latter involving both pyridyl rings and each carbonyl-O atom, Table 1b).The connections between chains leading to a three-dimensional architecture are of the type C—H⋯O, Crystal Explorer 17  hydrogen bonds was used as the input for calculations. A list of the short interatomic contacts discussed below is given in Table 2dnorm surfaces, Fig. 34LH2 and BA molecules, hereafter BA-I for the O3-containing molecule and BA-II for the O5-molecule, which indicate the presence of close contacts with distances shorter than the sum of the respective van der Waals radii , benzoic acid-O6—H6O⋯N4(pyridyl), amide-N2—H2N⋯O2(amide) and amide-N3—H3N⋯O5(carbonyl) hydrogen-bonding interactions exhibit the most intense red spots on the dnorm surfaces, suggestive of strong interactions.The program a) and (b) [in the order of moderate intensity (m) to weak intensity (w)] were identified for C6—H6B⋯O1 (m), C12—H12⋯O3 (m), C2—H2⋯O5 (m) and C1—H1⋯O3 (w), Table 1m), C28—H28⋯O1 (w), C9—H9B⋯C17 (w), C9—H9B⋯O3 (w), C13—H13⋯C1 (w), C12⋯C21 (w) and C8⋯C26 (w), Table 2PLATON  and (d).Other, relatively less intense red spots in Fig. 34LH2, BA-I and BA-II molecules through DFT-B3LYP/6-31G to further study the nature of the close contacts, Fig. 4-–H⋯N and N—H⋯O hydrogen-bonding contacts that exhibited the most intense red spots on the dnorm map are highly electrostatic in nature, as evidenced from the intense electronegative (red) and electropositive (blue) regions on the Hirshfeld surfaces of the individual molecules. Other regions are relatively pale, indicating the complementary role of the remaining contacts in sustaining the molecular network in the crystal.The electrostatic potential mapping was performed on the individual i.e. the three-molecule aggregate specified above, as well as its individual 4LH2, BA-I and BA-II components, Fig. 54LH2 and benzoic acid in constructing the molecular packing of the system, in contrast to the previously reported benzene monosolvate of 4LH2 i.e. the donor or acceptor atoms internal to the surface) and external  contacts. In contrast, for H⋯C/C⋯H and H⋯O/O⋯H, the distributions are slightly inclined towards -C⋯H- (15.8%) and -O⋯H- (13.4%) as compared to the corresponding counterparts at 11.7 and 11.8%, respectively. A detailed analysis of the di + de distances shows that the closest H⋯O/O⋯H and H⋯C/C⋯H contacts of ∼1.95 Å and ∼2.62 Å, respectively, occur at distances shorter than the sum of the respective van der Waals radii of 2.61 and 2.79 Å, while the H⋯H (∼2.20 Å) and N⋯H/ H⋯N (2.80 Å) contacts are longer than the sum of van der Waals radii of 2.18 and 2.64 Å, respectively.The major surface contacts for (I)4LH2 molecule also displays a shield-like profile with asymmetric spikes which upon further decomposition could be delineated into H⋯H (38.0%), H⋯O/O⋯H (25.6%), H⋯C/C⋯H (21.4%) and H⋯N/N⋯H (9.9%) contacts. The H⋯O/O⋯H contact exhibits a forceps-like profile with the distribution inclined towards internal-H⋯O-external (15.2%) as compared to internal-O⋯H-external (10.4%), and both with tips at di + de ∼1.94 Å which is indicative of significant hydrogen bonding. Similarly, the asymmetric, needle-like profile for the H⋯N/N⋯H contact is inclined towards the internal-N⋯H-external (9.0%) with the tip at di + de = ∼1.6 Å, while the remaining 0.9% is attributed to the internal-H⋯N-external contact with di + de of ∼2.94 Å . The H⋯C/C⋯H contacts are evenly distributed on both sides of the contacts with the di + de of ∼2.64 Å which is slightly shorter than the sum of van der Waals radii. On the other hand, the H⋯H contacts have little direct influence in sustaining the molecular packing as shown from the shortest di + de value of ∼2.2 Å, which is longer than the sum of the van der Waals radii despite the prominent contributions these make to the overall surfaceThe i.e. 31.9% for BA-I cf. 38.7% for BA-II. The discrepancy in the distribution for BA-I is compensated by the increase in O⋯C/C⋯O and C⋯C contacts with the distribution being 4.8 and 2.9%, respectively. The distribution for H⋯C/C⋯H (29.0 vs 29.1%), H⋯O/O⋯H (23.8 vs 24.2%) and H⋯N (6.7 vs 5.7%) contacts is approximately the same in both BA-I and BA-II, except that the H⋯C/C⋯H distribution for BA-II is significantly more inclined towards internal-C⋯H-external (20.5%) than the internal-H⋯C-external (8.6%) in contrast to the relatively balanced distribution for BA-I 15.5% for internal-C⋯H-external vs 13.5% for internal-H⋯C-external. In BA-I, the di + de values for H⋯O/O⋯H, H⋯C/C⋯H and H⋯N/N⋯H at the tips are ∼2.26–2.70, 2.62 and 1.64 Å, respectively, while the equivalent values for the analogous contacts for BA-II have tips at 2.02–2.56, 2.62–2.86 and 1.58 Å, respectively. Among these contact distances, the O⋯H, H⋯C/C⋯H and H⋯N for BA-I as well as H⋯O/O⋯H, H⋯C and H⋯N for BA-II are shorter than the sum of van der Waals radii. As expected, the minimum di + de value for the H⋯H contacts is longer than the sum of van der Waals radii, even if it is the most dominant contact for each molecule. The aforementioned data for BA-I and BA-II clearly distinguishes the independent molecules.As for the pair of BA molecules, both BA-I and BA-II possess similar, claw-like profiles which differ in the diffuse region, with the former being the characteristic of H⋯H contacts while the latter is due to H⋯C/C⋯H interactions. Quantitatively, differences mainly relate to the percentage contribution by H⋯H contacts, CrystalExplorer17  hydrogen bonds complemented by a pair of pyridyl-C13—H13⋯C1(pyridyl) interactions between two oxamide molecules led to the greatest interaction energy (Etot) of −75.2 kJ mol−1. This value is comparable to Etot of −71.7 kJ mol−1 calculated for the classical eight-membered {⋯HOCO}2 interaction  and pyridyl-C1—H1⋯O3(carbonyl) contacts, which combine to generate a seven-membered heterosynthon with Etot of −50.1 kJ mol−1. A diminution in Etot is observed for the other pyridyl terminus, which only comprises a carboxylic-O6—H6O⋯N4(pyridyl) hydrogen bond without a supporting pyridyl-C—H⋯O(carbonyl) interaction, showing an energy of −43.9 kJ mol−1 and ranked third strongest among all interactions in (I)To assess the strength of the specified interactions in the Hirshfeld surface analysis, the molecules in (I)Etot of −36.3 kJ mol−1 involves contributions from the amide-N3—H3N⋯O5(carboxylic acid) and phenyl-C28—H28⋯O1(amide) contacts. Other interactions include the methylene-C6—H6B⋯O1(amide) contacts (–25.1 kJ mol−1), the combination of benzoic acid-C20—H20⋯C8(amide) and benzoic acid-C12⋯C21(pyridyl) (–21.0 kJ mol−1), amide-C8⋯C26(benzoic acid) (–19.4 kJ mol−1), pyridyl-C2—H2⋯O5(carboxylic acid) (–15.6 kJ mol−1), a combination of (pyridine methyl)-C9—H9B⋯O3(carboxylic acid) and methylene-C9—H9B⋯C17(benzoic acid) (−14.7 kJ mol−1), as well as pyridyl-C12—H12⋯O3(carboxylic acid) (–8.3 kJ mol−1). Some inconsistencies are observed between the calculated Etot and Hirshfeld surface analysis, particularly for C2—H2⋯O5 and C12—H12⋯O3. These interactions can be considered weak even though they possess a relatively short contact distance compared to the sum of van der Waals radii, as indicated from the moderately intense red spots on the Hirshfeld surface. The contradiction could arise as a result of the relatively high repulsion terms, which weaken the interaction energy.The next highest interaction energy with 2O}2 synthon as well as the terminal interactions between 4LN2 and BA molecules, through hydroxy-O—H4⋯N(pyridyl) hydrogen bonds, lead to a zigzag electrostatic energy framework, Fig. 6a). The packing system is further stabilized by the dispersion forces contributed by the ten-membered {⋯HNC2O}2 synthon complemented by other peripheral interactions such the pairwise C20—H20⋯C8/C12⋯C21, C8⋯C26 and C6—H6B⋯O1 interactions, which result in a dispersion energy framework resembling a spider web, Fig. 6b). The combination of the electrostatic and dispersion forces leads to an overall energy framework that resembles a ladder, Fig. 6c).Overall, the crystal of (I)Chemical context, 4LH2 molecules have long been known to form co-crystals with carboxylic acids. A list of 4LH2/carboxylic acid co-crystals is given in Table 44LH2, the length of the central C—C bond, recognized as being long diazenyl]benzoic acid, carries a hydroxyl residue and this preferentially forms the hydrogen bonds to the pyridyl-N atoms. This observation is contrary to literature expectation where the carboxylic acid would be expected to form hydrogen bonds preferentially to pyridyl-N atom in instances where there is a competition with putative hydroxyl-O—H⋯N(pyridyl) hydrogen bonds  loop, in accord with Etter’s rules, i.e. ‘six-membered ring intramolecular hydrogen bonds form in preference to intermolecular hydrogen bonds’ oxalamide (4LH2), was prepared according to the literature; M.p.: 486.3–487.6 K; lit. 486–487 K  was mixed with benzoic acid  and the physical mixture was then ground for 15 min in the presence of a few drops of methanol. The procedures were repeated three times. Colourless blocks were obtained through careful layering of toluene (1 ml) on an N,N-dimethylformamide (1 ml) solution of the ground mixture. M.p.: 435.4–436 K. IR (cm−1): 3321 ν(N—H), 3070–2999 ν(C—H), 1702–1662 ν(C=O), 1506 ν(C=C), 1417 ν(C—N).The precursor, 4LH2:benzoic acid in molar ratios of 1:2, 1:3 and 1:4 were also attempted but only the 2:1 co-crystal (I)Similar experiments with Uiso(H) set to 1.2Ueq(C). The oxygen- and nitrogen-bound H atoms were located from a difference Fourier map and refined with O—H = 0.84±0.01 Å and N—H = 0.88±0.01 Å, respectively, and with Uiso(H) set to 1.5Ueq(O) or 1.2Ueq(N).Crystal data, data collection and structure refinement details are summarized in Table 510.1107/S2056989019016840/hb7875sup1.cifCrystal structure: contains datablock(s) I, global. DOI: 10.1107/S2056989019016840/hb7875Isup2.hklStructure factors: contains datablock(s) I. DOI: Click here for additional data file.10.1107/S2056989019016840/hb7875Isup3.cmlSupporting information file. DOI: 1972449CCDC reference:  crystallographic information; 3D view; checkCIF reportAdditional supporting information:


H-chromen-4-yl)oxy]acetic acid dimethylsulfoxide monosolvate is described and the intermolecular contacts in the crystal analysed using Hirshfeld surface analysis and two-dimensional fingerprint plots.The crystal structure of 2-[(2-oxo-2 11H8O5·(CH3)2SO, is a new coumarin derivative. The asymmetric unit contains two coumarin molecules (A and B) and two dimethylsulfoxide solvent molecules (A and B). The dihedral angle between the pyran and benzene rings in the chromene moiety is 3.56 (2)° for molecule A and 1.83 (2)° for molecule B. In molecule A, the dimethyl sulfoxide sulfur atom is disordered over two positions with a refined occupancy ratio of 0.782 (5):0.218 (5). In the crystal, molecules are linked by O—H⋯O hydrogen bonds, forming chains running along the c-axis direction. The chains are linked by C—H⋯O hydrogen bonds, forming layers parallel to the ac plane. In addition, there are also C—H⋯π and π–π interactions present within the layers. The intermolecular contacts in the crystal have been analysed using Hirshfeld surface analysis and two-dimensional fingerprint plots, which indicate that the most important contributions to the packing are from H⋯H (33.9%) and O⋯H/H⋯O (41.2%) contacts.The title compound, C The synthesis, and pharmacological and other properties of coumarin derivatives have been studied intensively and reviewed  and atoms O2A and O2B deviate from this mean plane by 0.007 (3) and 0.039 (3) Å, respectively. The dihedral angle between the pyran and benzene rings in the chromene moiety is 3.56 (16)° for molecule A and 1.83 (16)° for molecule B; this value is in agreement with those found in analogous coumarin derivatives  and 0.218 (5).The 1H-chromen-4-yl)oxy]acetyl}piperazin-1-yl)acetamide -2-[(2-oxo-2H-chromen-4-yl)oxy]acetamide , Cg1⋯Cg4iv = 3.509 (2) and Cg2⋯Cg3iv 3.572 (2) Å where Cg1, Cg2, Cg3 and Cg4 are the centroids of rings O1A/C1A/C6A–C9A, C1A–C6A, O1B/C1B/C6B–C9B, and C1B–C6B, respectively; symmetry code: (iv) x, 1 + y, z].The crystal structure features O—H⋯O and C—H⋯O hydrogen bonds Table 1. In the π Table 1 and π–π et al., 2019et al., 2007CrystalExplorer17  through white to blue . The dnorm surface was mapped over a fixed colour scale of 0.774 (red) to 1.381 (blue) for the title compound, where the red spots indicate the intermolecular contacts involved in the hydrogen bonding.The Hirshfeld surface of the title compound mapped over b) and O⋯H/H⋯O at 41.2% , followed by the C⋯H/H⋯C contacts at 9.6% , C⋯C contacts at 6.3%  and S⋯H/H⋯S contacts at 3.9% .The fingerprint plots are given in Fig. 62% Fig. 6c, follo6% Fig. 6d, C⋯C c3% Fig. 6e and S⋯9% Fig. 6f.et al., 2016et al., 2006H-furo-[1]-benzopyran-7-one monohydrate -4,5-bis(pyrrolidinylcarbonyl)-2,2-dimethyl-1,3-dioxolane cyclopropyl sulfamate  acetate  in THF (10 mL) at 273 K and stirred at 273 K for 1 h. Completion of the reaction was confirmed by TLC (mobile phase ethyl acetate/hexane) and THF was distilled off using a rotavapor. The obtained solution was washed with ethyl acetate (20 mL). The aqueous layer was acidified with 2N HCl (pH 1.0–2.0) and the obtained solid was filtered, washed with hexane and dried under vacuum to give as white solid. The purified compound was recrystallized using dimethyl sulfoxide as solvent.A solution of lithium hydroxide  in water (4 mL) was added to ethyl 2-(2-oxo-2Uiso(H) = 1.5Ueq(C-methyl) or 1.2Ueq(C) for other H atoms. In molecule A, the sulfur atom of the sulfinyldimethane group is disordered over two positions with refined occupancies of 0.782 (5) and 0.218 (5). In the final cycles of refinement, five outliers were omitted.Crystal data, data collection and structure refinement details are summarized in Table 210.1107/S2056989019009447/vm2218sup1.cifCrystal structure: contains datablock(s) global, I. DOI: 10.1107/S2056989019009447/vm2218Isup2.hklStructure factors: contains datablock(s) I. DOI: Click here for additional data file.10.1107/S2056989019009447/vm2218Isup3.cmlSupporting information file. DOI: 1891495CCDC reference:  crystallographic information; 3D view; checkCIF reportAdditional supporting information:


In the crystal, N—H⋯Cl, O–H⋯Cl and N—H⋯O hydrogen bonds connect the anions, cations and water molecules, forming a three-dimensional network.In this hydrated 2,6,13,17-tetraazoniatricyclo[16.4.0.0 20H44N44+·4Cl−·4H2O, has been determined using synchrotron radiation at 220 K. The structure determination reveals that protonation has occurred at all four amine N atoms. The asymmetric unit contains one half-cation , two chloride anions and two water molecules. There are two molecules in the unit cell. The Cl− anions and hydrate molecules are involved in hydrogen bonding. The crystal structure is stabilized by intermolecular hydrogen bonds involving the macrocycle N—H groups and water O—H groups as donors and the O atoms of the water molecules and the Cl− anions as acceptors, giving rise to a three-dimensional network.The crystal structure of the title salt, C These di- or tetraammonium cations may be suitable for the removal of toxic heavy metal ions from water. The macrocycle L contains a cyclam backbone with two cyclohexane subunits. Methyl groups are attached to the 3 and 14 carbon atoms of the propyl chains that bridge opposite pairs of N atoms in the structure. Previously, we have reported the crystal structures of [Cu(L)](NO3)2·3H2O, [Cu(L)](NO3)2, [Cu(L)](ClO4)2 and [Cu(L)(H2O)2](BF4)2·2H2O together with [Zn(L)(OCOCH3)2]. In these structures, the copper(II) or zinc(II) cations have tetragonally distorted octahedral environments with the four N atoms of the macrocyclic ligand in equatorial positions and O atoms of counter-anions, water molecules or acetato ligands in axial positions ·2MeOH The macrocycle 3,14-dimethyl-2,6,13,17-tetraazatricyclo(16.4.0.0− anions and four solvent water molecules and was characterized during studies of the macrocyclic ligand and its copper(II) complexes. An ellipsoid plot of the molecular components in (I)anti with respect to the macrocyclic plane as a result of the molecular inversion symmetry. The six-membered cyclohexane ring is in a stable chair conformation. Within the centrosymmetric tetra-protonated amine unit [C20H44N4]4+, the C—C and N—C bond lengths vary from 1.522 (2) to 1.542 (2) Å and from 1.506 (2) to 1.522 (2) Å, respectively. The ranges of N—C—C and C—N—C angles are 106.85 (10) to 114.32 (11)° and 116.70 (10) to 118.89 (10)°, respectively. The bond lengths and angles within the [C20H44N4]4+ tetra-cation are comparable to those found in the free ligand or the di-cation in C20H40N4·2C11H10O ·2MeOH 3}(CN)3]2·2H2O·2MeOH . The crystal structures of C20H40N4·2C11H10O ·2MeOH 3}(CN)3]2·2H2O·2MeOH  were used as provided. All chemicals were reagent grade and used without further purification. As a starting material, the macrocycle 3,14-dimethyl-2,6,13,17-tetraazatricyclodocosane was prepared according to a published procedure  in water (15 mL). The solution was heated for 1 h at 338 K. After cooling to 298 K, the pH was adjusted to 3.0 with 1.0 M HCl. The solution was filtered and left at room temperature. Colourless crystals suitable for X-ray analysis were obtained unexpectedly from the solution over a period of a few days.Commercially available Uiso(H) values of 1.2Ueq and 1.5Ueq(C-methyl). O-bound H atoms of the water molecules were located in a difference-Fourier map, and the O—H distances and the H—O—H angles were restrained using DFIX and DANG constraints (0.84 and 1.36 Å).Crystal data, data collection and structure refinement details are summarized in Table 210.1107/S2056989018009337/sj5559sup1.cifCrystal structure: contains datablock(s) I. DOI: 10.1107/S2056989018009337/sj5559Isup2.hklStructure factors: contains datablock(s) I. DOI: Click here for additional data file.10.1107/S2056989018009337/sj5559Isup3.cmlSupporting information file. DOI: 1852146CCDC reference:  crystallographic information; 3D view; checkCIF reportAdditional supporting information:


Cav1.2 is the pore‐forming subunit of L‐type voltage‐gated calcium channel (LTCC) that plays an important role in calcium overload and cell death in Alzheimer's disease. LTCC activity can be regulated by estrogen, a sex steroid hormone that is neuroprotective. Here, we investigated the potential mechanisms in estrogen‐mediated regulation of Cav1.2 protein. We found that in cultured primary neurons, 17β‐estradiol (E2) reduced Cav1.2 protein through estrogen receptor α (ERα). This effect was offset by a proteasomal inhibitor MG132, indicating that ubiquitin–proteasome system was involved. Consistently, the ubiquitin (UB) mutant at lysine 29 (K29R) or the K29‐deubiquitinating enzyme TRAF‐binding protein domain (TRABID) attenuated the effect of ERα on Cav1.2. We further identified that the E3 ligase Mdm2 (double minute 2 protein) and the PEST sequence in Cav1.2 protein played a role, as Mdm2 overexpression and the membrane‐permeable PEST peptides prevented ERα‐mediated Cav1.2 reduction, and Mdm2 overexpression led to the reduced Cav1.2 protein and the increased colocalization of Cav1.2 with ubiquitin in cortical neurons in vivo. In ovariectomized (OVX) APP/PS1 mice, administration of ERα agonist PPT reduced cerebral Cav1.2 protein, increased Cav1.2 ubiquitination, and improved cognitive performances. Taken together, ERα‐induced Cav1.2 degradation involved K29‐linked UB chains and the E3 ligase Mdm2, which might play a role in cognitive improvement in OVX APP/PS1 mice. This effect involved lysine 29‐linked ubiquitin chains and the E3 ligase Mdm2 (double minute 2 protein). In OVX APP/PS1 mice, systematic administration of E2 and ERα agonist PPT (propylpyrazoletriol) led to the reduced Cav1.2 protein and the enhanced Cav1.2 ubiquitination in the brain, which were accompanied by the improved cognitive functions.22.1Ca2+density) were measured using whole‐cell patch recordings. As shown in Figure Ca2+density (pA/pF) was significantly decreased by E2 treatment (24 hr), whereas ICI attenuated this effect. These results indicated that E2 reduced Cav1.2 protein in cultured primary cortical neurons, which was mediated by ERs.As shown in Figure Ca2+ density  might mediate E2 effect on Cav1.2, we tested the effect of the selective ERα agonist  or ERβ agonist  on Cav1.2 incubated in cortical neurons for 24 hr. As shown in Figure y Figure . As expe2.2Cav1.2 protein can be regulated at multiple levels  controls Cav1.2 degradation  protein contains seven lysine (K) residues at positions 6, 11, 27, 29, 33, 48, and 63, respectively. In addition, poly‐UB chain assembly can occur at any of these lysine residues   and CHIP  are reportedly associated with ERα signaling   mice were significantly shorter than that in saline‐treated OVX APP/PS1 (AD) mice beginning on the third day Figure a–c. In t3The efficacy of estrogen‐based hormone therapy (HT) in AD is not conclusive. Some studies reveal that HT may reduce the risk of AD in postmenopausal women  mediates proteasomal degradation of Cav1.2 in neuronal cells  were from Abcam  and Alomone, respectively, which produced identical Western blots. Those against β‐actin (ab6276), MAP‐2 (ab32454), ERα (ab3573), ERβ (ab104804), calcineurin (ab3673), Mdm2 (ab16895), and ubiquitin (ab19247) were purchased from Abcam. pSer1928‐Cav1.2 (A010‐70) was from Badrilla (United Kingdom). 17β‐estradiol (E2758), propylpyrazoletriol , diarylprepionitrile , E2‐BSA (E5630), ICI 182,780 (V900926), nifedipine (N7630), FK506 (F4679), MG132 (C2211), ammonium chloride (A0171), chloroquine (C6628), carfilzomib (791938), and other reagents were purchased from Sigma . Lipid solvents were made in stock solutions in DMSO .4.2pCMV6‐Myc‐Mdm2 and pCMV6‐Myc‐p14ARF were purchased from OriGene (USA). TRABID (DU# 49067) and TRABID mutant C443A (DU# 49089) were purchased from MRC PPU Reagents and Services (University of Dundee). pRK5‐HA‐ubiquitin was obtained from Addgene. All ubiquitin mutants were made from pRK5‐HA‐ubiquitin plasmid by mutagenesis. pRK5‐HA‐Ubiquitin‐K0 (no lysine), pRK5‐HA‐Ubiquitin‐K6 (keep lysine at position 6), pRK5‐HA‐Ubiquitin‐K11, pRK5‐HA‐Ubiquitin‐K27, pRK5‐HA‐Ubiquitin‐K29, pRK5‐HA‐Ubiquitin‐K33, pRK5‐HA‐Ubiquitin‐K48, pRK5‐HA‐Ubiquitin‐K63, pRK5‐HA‐Ubiquitin‐K6R , pRK5‐HA‐Ubiquitin‐K29R, pRK5‐HA‐Ubiquitin‐K63R, CHIP with myc tag, and Derlin‐1 with HA tag were cloned to pcDNA3, respectively.4.36 cells/ml for biochemical experiments and 0.5 × 106 cells/ml for molecular and immunohistochemistry experiments. Cultures were maintained in serum‐free neurobasal (NB) medium  at 37°C with 5% CO2. Fifty percent of the medium was exchanged with fresh medium every other day for 2 weeks. On the day before experiment, culture medium was completely replaced by fresh medium in the presence of accurate dilution of chemicals.Cortical neurons were extracted from prenatal pups on embryonic day 18. Cells were placed on poly‐L‐lysine (0.1 mg/ml)‐coated 6‐well plates, at a density of 2 × 104.42. At ~ 90% confluence, cells were transiently transfected with various plasmids or Mdm2 siRNA using the Lipofectamine 2000 reagent (Invitrogen) according to the manufacturer's instructions. A total of 2.5 μg of DNA in 100 μl Opti‐MEM (Invitrogen) were used in each of six wells. The following human Mdm2 siRNA sequences were used for transfection in SH‐SY5Y cells: sense CGUACGCGGAAUACUUCGATT, antisense AAUCGAAGUAUUCCGCGUACG; NC: sense CGUACGCGGAAUACUUCGATT, antisense AAUCGAAGUAUUCCGCGUACG (100 pM per six wells for 72 hr). The effectiveness of Mdm2 knockdown was verified by Western blotting analysis.HT22  and SH‐SY5Y (human neuroblastoma) cell lines were obtained from the Type Culture Collection of the Chinese Academy of Sciences . Cells were incubated in Dulbecco's modified Eagle's medium  containing 4.5 g glucose/L, supplemented with penicillin/streptomycin (50 units/mL), glutamate (2 mM), and 10% fetal bovine serum , and were maintained at 37°C with 10% CO4.5Western blotting of Cav1.2 was performed as previously described  assays, the cell or tissue extracts (0.5 mg) were pre‐incubated with protein A agarose beads  for 3 hr to reduce nonspecific reaction. The supernatants were then mixed with nonspecific IgG and appropriate antibodies for 1 hr. The mixtures were then incubated with protein A agarose beads  overnight at 4°C. Samples were washed two times with lysis buffer and two times with sterile PBS and denatured with SDS sampling buffer.4.6Brain sections were washed with ice‐cold PBS, then fixed in ice‐cold 4% paraformaldehyde for 10 min, permeabilized in 0.1% Triton X‐100 for 10 min, and blocked with 10% normal goat serum for 1 hr at 37°C with extensive PBS washings between each step. Specimens were incubated with appropriate primary antibodies (1:50 in PBS) at 4°C overnight. The sections were then washed 6 × 2 min with PBS, followed by incubation with the secondary antibody Alexa 549 or Fluro 488 (1:100 in PBS) at 37°C for 1 hr. After sealing with a mixture of glycerol and PBS at a ratio of 1:1, the immunofluorescent‐labeled sections were examined using a laser scanning confocal microscope  with an Olympus IX 70 inverted microscope  equipped with a Fluoview FVX confocal scan head.4.72+) were recorded from cultured cortical neurons from 7 to 11 days in vitro (DIV), at room temperature using a MultiClamp 700B amplifier (Axon Instruments). The recording chamber was perfused with Tyrode's solution containing (in mM): NaCl 100, tetraethylammonium‐Cl 20, 4‐AP 5, KCl 4, MgCl2 1, BaCl210, glycine 0.01, HEPES 25, glucose 30, and TTX 0.001. The glass micropipettes had a resistance of 3–5 MΩ when filled with internal solution containing (in mM): CsCl 135, MgCl2 4, HEPES 10, EGTA 10, MgATP 4, and Na3GTP 0.3. A prepulse protocol consists of 300–700 ms to −30 mV followed by 50 ms to −50 mV, before each test pulse was used to inactivate T‐type Ca2+ channels and Na+ channels. ICa2+ was elicited by 300 ms test pulses of variable amplitude (−40 to +60 mV at a step of 10 mV) from a holding potential of −60 mV. The interval between test pulses was 10 s.Voltage‐dependent calcium‐mediated currents . The primer sequences for Cav1.2 were as follows: 5′‐TACCGTCAGTTCCACACAGC‐3′ and 5′‐CTTCAGAGTCAGGCAGAGCA‐3′. The threshold cycle (Ct) value of each sample was calculated, and the relative mRNA level was normalized to the GAPDH mRNA value.4.9ERα/β shRNAs was produced by Shanghai GeneChem Co., Ltd. . At 10 DIV, ER‐related lentivirus was added to the culture for 72 hr according to the manufacturers’ instruction , while transfection of Mdm2 was performed in HT22 cells.The target shRNAs against rat ERα/β gene (NM_012689/NM_012754) were designed as follows: ERα‐1: GATAAGAACCGGAGGAAGA, ERα‐2: CCAGAATGGCCGAGAGAGA, and ERα‐3: GTAAATGTGTAGAAGGCAT; ERβ‐1: TGAGCAAAGCCAAGAGAAA, ERβ‐2: TCACTAAGCTGGCGGACAA, and ERβ‐3: CCAAATGTGCTATGGCCAA. Negative control was the scrambled sequence with no homology to rat genes. The recombinant lentiviral vector bearing rat 4.10in utero electroporation was performed on pregnant C57/B6L mice on embryonic day 14.5 (E14.5) as previously described  and PEST3 (S840/861) were synthesized and purified by Pepmic Co. Ltd . The sequences were as follows: YGRKKRRQRRR‐DIDPENEDEGMDED‐GFP for TAT‐PEST1, YGRKKRRQRRR‐DIDKENEDEGKDED‐GFP (ΔPEST1) for scrambled, YGRKKRRQRRR‐HSNPNTAGEEDEEEPEMPVGPR‐GFP for TAT‐PEST3, and YGRKKRRQRRR‐HSNPNPAGEEDKEEPAMPVGPR‐GFP (ΔPEST3) for scrabbled control  were obtained from Nanjing University (China). Eight‐month‐old female APP/PS1 mice were bilaterally ovariectomized (OVX) for 14 days. Animals were subcutaneously injected daily with vehicle, 17β‐estradiol , PPT (1 mg/kg), and DPN (1 mg/kg) for two weeks. The dosage of E2 is thought to be within physiological range; and those of PPT and DPN are functional  were recorded by a video and analyzed by image analyzing software . For contextual and cued fear conditioning, mice were tested in a 3‐day paradigm. Behavior was recorded by video camera, and freezing data were measured using FreezeScan software.4.14SEM from at least three independent experiments. The statistical comparisons between two groups were tested using Student's t test. The comparisons among groups were tested using one‐way or two‐way ANOVA, and a post hoc pairwise comparison was used where it applied.Data were presented as means ± None declared.G‐J Chen and Z Yan designed the research; Y‐J Lai performed the research and analyzed the data; B‐L Zhu, F Sun, D Luo, Y‐L Ma, B‐Luo, J Tang, M‐J Xiong, L Liu, X‐T Hu, L He, X‐J Deng, J‐H Zhang, and J Yang provided assistance with the research; G‐J Chen and Y‐J Lai wrote the paper. All authors read and approved the final manuscript. Click here for additional data file. Click here for additional data file. Click here for additional data file. Click here for additional data file. Click here for additional data file.


R)‐[11C]PK11195 was investigated. The synthesis of the reference compound was accomplished in six steps and 9 % overall yield, and the precursor was prepared in eight steps and 8 % overall yield. The chiral separation of the reference and precursor compounds was performed using supercritical fluid chromatography with >95 % ee. The absolute configuration was determined by circular dichroism. Optimisation of reaction conditions for manual radiolabelling revealed acetonitrile as a preferred solvent at 100 °C. Automation of this radiolabelling method provided R and S enantiomers in respective 21.3±16.7 and 25.6±7.1 % decay‐corrected yields and molar activities of 55.8±35.6 and 63.5±39.5 GBq μmol−1 (n=3). Injection of the racemic analogue into a healthy rat confirmed passage through the blood–brain barrier.Translocator protein (TSPO) is a biomarker of neuroinflammation, which is a hallmark of many neurodegenerative diseases and has been exploited as a positron emission tomography (PET) target. Carbon‐11‐labelled PK11195 remains the most applied agent for imaging TSPO, despite its short‐lived isotope and low brain permeability. Second‐generation radiotracers show variance in affinity amongst subjects  caused by the genetic polymorphism (rs6971) of the TSPO gene. To overcome these limitations, a new structural scaffold was explored based on the TSPO pharmacophore, and the analogue with a low‐affinity binder/high‐affinity binder (LAB/HAB) ratio similar (1.2 vs. 1.3) to that of ( R‐1‐(2‐chlorophenyl)‐N‐methyl‐N‐(1‐methylpropyl)‐3‐isoquinoline carboxamide ((R)‐[11C]PK11195, [11C]1, Figure m).R)‐[11C]1 has been used in PET imaging to show neuroinflammation in vivo in cases of stroke, neurodegeneration, traumatic brain injury, and neoplasia.11C]1 in research and clinical settings, such as low brain permeability,t1/2=20 min), which restricts its use to PET centres that have a cyclotron on site.Neuroinflammation, an immune response to neuronal insult, is a core component of many disorders such as stroke, multiple sclerosis (MS), neurodegenerative disorders, and brain tumours.11C]1, multiple new TSPO radiotracers have been developed and studied; these have improved properties for application, including several that are labelled with the longer‐lived radioisotope fluorine‐18 (t1/2=109.9 min).N‐(2‐(methoxy‐11C)benzyl)‐N‐(4‐phenoxypyridin‐3‐yl)acetamide , and good clearance from striatum (region of low TSPO expression) in rats.In 2004, a study by Okubo et al. described a tetracyclic indole‐based pharmacophore .However, while 6 as well as the initial in vivo preclinical PET.Toward this objective informed by structure–activity design features, a novel series of compounds based on the chemical scaffold18F]6, we proposed an SN2 nucleophilic substitution of a good leaving group with [18F]fluoride, for which reason tosylate 25 and mesylate 26 were envisioned as potential precursors. Reference compound 6  to give product 23. A second hydrogenation on crude 23 in the absence of triethylamine was performed to remove the benzyl protecting group to give crude 24 in quantitative yield. Using crude alcohol 24, we prepared both crude tosylate 25 and crude mesylate precursor 26 . However, the mesyl precursor 26 required purification by column chromatography, yielding a viscous oil (37 %), thus making tosyl precursor 25 practically easier to purify and handle than the mesyl precursor 26.Analogously, 2‐chloro‐5‐methoxyaniline hydrochloride 18F]6 by establishing the chiral separation of both reference compound 6 as well as its radiolabelling precursor (25). A clear choice for chiral separation was found in the preparation of diastereomeric mixture by application of a chiral derivatisation group. We identified an acid intermediate 21 for further functionalisation with several potential enantiomerically pure alcohols. The alcohols selected were l‐menthol, (R)‐(−)‐2‐butanol, and (R)‐1‐phenyl‐2‐propyn‐1‐ol. The ester 28 with (R)‐(−)‐2‐butanol was formed in 53 % yield using DBU as a base and a mesylate analogue 27 of the alcohol as an intermediate. However, a higher‐yielding and a more direct method of EDCI/DMAP coupling was used to form the esters with l‐menthol and (R)‐1‐phenyl‐2‐propyn‐1‐ol, which afforded 29 and 30 in 54 and 31 % yields, respectively ‐1‐phenyl‐2‐propynyl analogues. Flash chromatography using diethyl ether and cyclohexane yielded two diasteriomerically enriched samples with both esters 29 (in 78 and 40 % diastereomeric excess) and 30 (in 82 and 35 % diastereomeric excess) ‐29, ‐29 and ‐30, ‐30). To demonstrate the presence of a single enantiomer in a mixture, the chiral l‐menthol functionality was first removed under a variety of basic conditions (Table 2O in EtOH (entry 1), which were the hydrolytic conditions used in the synthesis of the reference compound 6 and precursors 25 and 26. Reaction with LiOH in a mixture of THF/H2O (entry 2) gave a minor byproduct.While NMR analysis demonstrated the presence of two diastereomers in all three product mixtures, TLC analysis of the ester mixtures using a variety of solvent systems showed the two diastereomers could be separated readily by flash column chromatography in two cases, that of 2CO3 in MeOH (entry 3) gave an equal amount of the desired product and an unidentified byproduct,l‐menthol using EDCI/DMAP. To our disappointment, 1H NMR analysis indicated complete racemisation from all investigated hydrolytic reaction conditions. This was further confirmed by chiral HPLC analysis of the acid samples obtained from hydrolysis of diastereomerically enriched ester samples ‐21, Scheme 21 in a solution presumably due to the presence of a highly acidic proton is a likely explanation for such rapid racemisation of the stereocenter. In the case of the ester analogue 30, while we were able to obtain diastereomerically enriched samples through multiple rounds of flash column chromatography in diastereomeric excess ‐30, ‐30), a previously described CuCl hydrolysis protocol failed to give the enantiomerically enriched product 21 (Scheme 6 and tosylate 25 (see the Supporting Information for details).Using K6 was next determined by electronic circular dichroism (CD) spectroscopy whereby CD spectra of 6 were compared with that of 5, for which stereochemistry had already been established ‐5 was composed of a maximum positive Cotton effect at 200 nm and a corresponding maximum negative Cotton effect at 226 nm. The enantiomer of 6, which similarly exhibited a maximum positive Cotton effect at 204 nm and a corresponding maximum negative Cotton effect at 228 nm, was in analogy assigned as the S isomer. The other isomer showed the mirror image bisignate band in the CD spectrum and was therefore identified as R enantiomer.The absolute stereochemistry of enantiomers of d Figure . The speS enantiomer (Supporting Information).To correlate enantiomers obtained via SFC separation to CD data, the chiral HPLC separation was performed on a Whelk‐O1 column to unambiguously identify first eluting peak as the 5 was shown to bind to wild‐type TSPO with a higher affinity than compound 1, but lost affinity at A147T TSPO (Table S)‐6 bound to wild‐type TSPO with a similar affinity to compound 1, and both compounds 1 and (S)‐6 only showed minor loss in affinity at A147T TSPO ‐6: 36.3 nm, while 1.04 nm for rat TSPO.Using an assay based on human embryonic kidney cell lines stably overexpressing human TSPO wild‐type and TSPO A147TPO Table . Compoun18F]5, manual radiochemical reactions were performed first. Both precursors 25 and 26 were assessed and the effect of temperature, solvent, and time was investigated (Table 25 (entries 1–6), a single radiolabelled product formed which was identified as [18F]6 via co‐injection of reference compound 6. However, when using 26 (entries 7–12), a second unidentified radiolabelled product was formed during the reaction. The amount of this radiolabelled side product which formed immediately, did not change during the course of the reaction.To establish suitable reaction conditions for radiolabelling of [ed Table . For all18F]fluoride to [18F]6 was observed: the highest being 18 % when MeCN was used as a solvent and mesylate 25 as a precursor. When the temperature was increased to 100 °C or 120 °C reactions performed in DMF similarly showed poor conversion (entries 6 and 12) after 20 min. In DMSO, [18F]6 formed with 3 % conversion after 20 min (entry 10) when 26 was used as precursor, whereas tosylate 25 afforded 47 % conversion into [18F]6 under analogous conditions (entry 4). The highest conversion in the case of both precursors 25 and 26 was observed after 20 min in MeCN: 81 and 39 %, respectively (entries 2 and 8).At 80 °C, independent of solvent and precursor, low conversion of <20 % of [25 was used for automated radiolabelling on a GE Healthcare FXFN TRACERlab instrument. [18F]Fluorination was achieved with cyclotron‐produced [18F]fluoride in MeCN at 100 °C for 20 min. Product [18F]6 was purified by semipreparative reversed‐phase HPLC.With this result in hand , next tosylate 18F]6 was 60 min from end of bombardment (EOB). Radiochemical purity was over 99 %, and radiolabelling yield was 20.8±4.5 % decay‐corrected to EOB or 13.6±2.8 % non‐decay‐corrected, and molar radioactivity 93.2±50.6 GBq μmol−1 (n=9) at the end of synthesis (EOS). Furthermore, the same automated protocol was used for the radiosyntheses of (R)‐[18F]6 and (S)‐[18F]6 as follows: 21.3±16.7 and 25.6±7.1 % decay‐corrected yields, radiochemical purity >98 % and molar radioactivities 55.8±35.6 and 63.5±39.5 GBq μmol−1 (n=3) at the EOS, respectively.Total radiosynthesis time for the preparation for [18F]6 enters the brain  was distilled over P2O5 and stored over KOH. Reported density values are for ambient temperature. Purity of compounds was ≥95 % as determined by analytical HPLC on a Thermo Dionex 3000 HPLC system. Preparative chromatographic separations were performed on Material Harvest silica gel 60 (35–75 μm) and reactions followed by TLC analysis using Sigma–Aldrich silica gel 60 plates (2–25 μm) with fluorescent indicator (254 nm) and visualised with UV or potassium permanganate.All reactions requiring anhydrous conditions were conducted in oven‐dried glass apparatus under an atmosphere of inert gas. Reagents were purchased from Sigma–Aldrich or Alfa Aesar. Reagents were used without further purification unless otherwise noted. Triethylamine =7.26 ppm, δC=77.2 ppm. Multiplicities in the 1H NMR spectra are described as: s=singlet, d=doublet, t=triplet, q=quartet, quint.=quintet, s=sextet, h=heptet, m=multiplet, b=broad; coupling constants are reported in Hz.18 1.7 μm, 2.1×50 mm; flow rate: 0.6 mL min−1, 4 min method, 5–95 % MeCN/H2O with 0.1 % formic acid. HRMS data were obtained from the EPSRC Mass Spectrometry Service at the University of Swansea. Ion mass/charge (m/z) ratios are reported as values in atomic mass units.LC–MS data were obtained using a Waters Acquity‐H/Xevo TQD LC–MS instrument. Column: Acquity UPLC® BEH CFN TRACERlab. Semipreparative purification of radiolabelled material was performed on a Merck‐Hitachi L6200A system equipped with Knauer variable wavelength detector and an Eberline radiation detector using a reversed‐phase column  and eluting with 48 % aq. MeCN at a flow rate of 5 mL min−1. Analytical HPLC samples were analysed by an Agilent HPLC 1100 system equipped with UV multi‐wavelength detector and Raytest Gabi star radiation detector using reversed‐phase column (Phenomenex Luna 5 μm C18 (2) 100 Å, 250×4.6 mm, 675295‐11) and eluting with 0–3 min isocratic 10 % aq. MeCN, 3–5 min gradient 10–80 % aq. MeCN, 6–12 min isocratic 80 % aq. MeCN at a flow rate of 1.5 mL min−1.The automated radiosynthesis was performed on a GE Healthcare FX2‐Fluoroethyl 4‐methylbenzenesulfonate (8). Commercially available 2‐fluoroethanol 7  was dissolved in pyridine (2.5 mL) under nitrogen. The solution was stirred at 0 °C and tosyl chloride  added portion‐wise to the solution over a period of 30 min, keeping the temperature below 5 °C. The reaction was stirred at RT for 18 h. The reaction was then quenched by careful addition of ice followed by water (5 mL). The reaction mixture was extracted into EtOAc (10 mL) and washed with water (3×10 mL), 1 m HCl solution (10 mL), 1 m aqueous sodium carbonate (10 mL), and copper sulfate (2×10 mL). The organic layer was washed with brine (10 mL), dried (MgSO4) and concentrated in vacuo to give 8 as an oil . LC–MS: Rf 1.86 (−ESI) m/z 311.5 ([M+94]+); 1H NMR : δH=7.81 , 7.36 , 4.57 , 4.26 , and 2.45 . Characterization data for this compound are in complete agreement with previously published data.N‐(2‐fluoroethyl)‐5‐methoxyaniline (10)2‐Chloro‐. Commercially available 2‐chloro‐5‐methoxyaniline hydrochloride 9  was dissolved in DMF (60 mL) and sodium hydride  was added. The reaction was stirred for 30 min at RT under nitrogen. The tosylate 8  in DMF was then added dropwise and the reaction was stirred at RT for 2 h. The reaction was then heated at 100 °C for 18 h. The reaction was allowed to cool and the solvent was removed under reduced pressure. The residue was dissolved in EtOAc (100 mL) and washed with water (5×100 mL). The organic layers were combined, dried (MgSO4) and concentrated in vacuo to give a brown oil. The crude mixture was then purified by silica flash chromatography eluting with a gradient of 5–30 % EtOAc/petroleum ether (40–60 °C) to yield 10 as a yellow oil . LC–MS: Rf 2.01 (+ESI) m/z 204.3 ([M+H]+); 1H NMR : δH=7.16 CHCH), 6.25–6.22 CHCH), 4.64 , 4.65–4.55 , 3.77 , and 3.48 . Characterization data for this compound are in complete agreement with an internal GE publication.Ethyl 3‐bromo‐2‐hydroxycyclohex‐1‐ene‐1‐carboxylate (11). Ethyl 2‐oxocyclohexanecarboxylate S1 was dissolved in diethyl ether and cooled to 0 °C under nitrogen. Bromine was added dropwise over 15 min and the reaction mixture was allowed to warm to RT over 90 min. The mixture was slowly poured into ice‐cold saturated aqueous potassium carbonate and extracted with ethyl acetate. The combined organic layers were dried over magnesium sulfate, filtered, concentrated in vacuo and dried on the vacuum line for 18 h to afford 11 as a yellow oil. LC–MS: Rf 2.17; 1H NMR : δH=11.92 , 4.93 ), 4.20 OCH2CH3), 2.53–2.46 , 2.12–2.03 , 1.79–1.67 , 1.24 OCH2CH3). Characterization data for this compound are in complete agreement with an internal GE publication.Ethyl‐3‐((2‐chloro‐5‐methoxyphenyl)(2‐fluoroethyl)amino)‐2‐hydroxycyclohex‐1‐ene‐1‐carboxylate (12). A solution of the aniline 10  in THF (60 mL) was cooled to −40 °C. Potassium bis(trimethylsilyl)amide  was added dropwise and the reaction stirred for 30 min at −40 °C. The carboxylate 11  in THF (10 mL) was added dropwise at −40 °C. The cooling bath was removed and the reaction was stirred at RT for 4 h. The reaction was quenched with brine (100 mL) and extracted into EtOAc (2×200 mL), dried (MgSO4) and concentrated in vacuo to give 12 as a brown oil  which was used crude in the next step. LC–MS: Rf 2.29 (+ESI) m/z 372.4 ([M+H]+). Characterization data for this compound are in complete agreement with an internal GE publication.H‐carbazole‐4‐carboxylate (13)Ethyl 8‐chloro‐9‐(2‐fluoroethyl)‐5‐methoxy‐2,3,4,9‐tetrahydro‐1. The intermediate 12  was dissolved in Et2O (100 mL) and zinc chloride  was added. The reaction was heated at reflux for 16 h. EtOAc (300 mL) was added to dissolve everything and was washed with 2 m HCl (200 mL), water (200 mL), 10 % aqueous potassium carbonate (200 mL), dried (MgSO4) and concentrated in vacuo. The crude material was purified by silica gel chromatography eluting with a gradient of 5–20 % of EtOAc/petroleum ether (40–60 °C) to afford 13 as a yellow oil . Material could not be obtained in a pure form, but regardless, the reaction was progressed to the next step. LC–MS: Rf 2.38 (+ESI) m/z 354.4 ([M+H]+); 1H NMR : δH=6.95 , 6.35 , 4.90–4.40 , 4.20–4.10 , 2.80–2.65 , 3.79 , 2.10–1.80 , and 1.30–1.20 . Characterization data for this compound are in complete agreement with an internal GE publication.H‐carbazole‐4‐carboxylate (14)Ethyl 9‐(2‐fluoroethyl)‐5‐methoxy‐2,3,4,9‐tetrahydro‐1. The chloro intermediate 13  was dissolved in methanol (50 mL) and triethylamine  and 10 % Pd/C (0.414 g) were added. The mixture was stirred for 18 h after purging twice (over 2 h) with hydrogen gas under atmospheric pressure. The reaction was filtered through a pad of Celite under nitrogen atmosphere, washed with methanol (50 mL), and the solvent was removed in vacuo. The residue was dissolved in EtOAc (100 mL) and washed with 10 % aqueous potassium carbonate (100 mL), dried (MgSO4) and concentrated in vacuo to give 14 as an oil . LC–MS: Rf 2.18 (+ESI) m/z 320.5 , Rf 2.20 (−ESI) m/z 318.4 ([M−H]−); 1H NMR : δH=7.04 , 6.84 , 6.46 , 4.64 , 4.40–4.00 , 3.82 , 2.90–2.60 , 2.20–1.80 , and 1.30–1.20 . Characterization data for this compound are in complete agreement with an internal GE publication.boxylate . The chlH‐carbazole‐4‐carboxylic acid (15)9‐(2‐Fluoroethyl)‐5‐methoxy‐2,3,4,9‐tetrahydro‐1. The ester 14  was dissolved in ethanol (10 mL). A solution of sodium hydroxide  dissolved in 10 mL of water, was added. The reaction mixture was heated at reflux for 18 h. The solvent was removed in vacuo and the crude mixture diluted with water (30 mL), acidified with 2 m HCl dropwise until acidic, and washed with CH2Cl2 (50 mL). The organic layers were combined and dried (MgSO4) and concentrated in vacuo to give 15 , which was used crude into the next step. LC–MS: Rf 1.81 (+ESI) m/z 292.4 ([M+H]+).H‐carbazole‐4‐carbonyl chloride (16)9‐(2‐Fluoroethyl)‐5‐methoxy‐2,3,4,9‐tetrahydro‐1. A solution of 15  in anhydrous CH2Cl2 (5 mL) was stirred under nitrogen. Oxalyl chloride  was added followed by a drop of DMF. The reaction mixture was stirred at RT under nitrogen for 2 h then evaporated in vacuo to give 16  as an oil, which was used into the next step without purification.NN‐ethyl‐9‐(2‐fluoroethyl)‐5‐methoxy‐2,3,4,9‐tetrahydro‐1H‐carbazole‐4‐carboxamide (GE387) (6)‐Benzyl‐. The acid chloride 16  was dissolved in anhydrous CH2Cl2 (8 mL) and cooled to 0 °C. N‐Ethylbenzylamine  was then added and the reaction was stirred for 18 h at RT. The reaction was quenched with 10 % aqueous potassium carbonate (6 mL). The CH2Cl2 layer was collected via a separatory funnel, dried (MgSO4), and then concentrated in vacuo. The crude material was purified by silica gel chromatography eluting with a gradient of 50–100 % EtOAc/petroleum ether (40–60 °C) to afford the crude product as a yellow oil. The oil was then recrystallised from EtOAc to afford 6 as a pale‐yellow solid . LC–MS: Rf 2.30 (+ESI) m/z 409.6 , Rf 2.35 (+ESI) m/z 409.6 ([M+H]+); 1H NMR : δH=7.42–7.21 , 7.02 , 6.84 , 6.43 , 4.80–4.40 , 4.31 , 3.62 , 3.83–3.41 2), 2.88–2.61 , 2.20–1.80 , and 1.32 2).N‐(2‐(Benzyloxy)ethyl)‐2‐chloro‐5‐methoxyaniline (18). Commercially available 9  was converted into the free base with 1 m aqueous sodium carbonate (300 mL). The mixture was extracted with CH2Cl2 (2×200 mL), the organic layer dried (MgSO4) and evaporated to give an oil. Compound 9 was then dissolved in CH2Cl2 (50 mL), in a dry flask under nitrogen. Benzyloxyacetaldehyde 17  and acetic acid  were added. After 15 min sodium triacetoxyborohydride  was added. The mixture was stirred for 18 h at RT and then poured into saturated aqueous ammonium chloride solution (100 mL) and extracted with CH2Cl2 (2×100 mL). The combined organic layers were dried (MgSO4) and evaporated to give an oil. The crude product was purified via silica flash chromatography eluting with CH2Cl2 to yield 18 as an oil . Material could not be obtained in a pure form, but regardless, the reaction was progressed to the next step. LC–MS: Rf 2.42 (+ESI) m/z 292.3 ([M+H]+); 1H NMR : δH=7.37–7.30 , 7.14 CHCH), 6.23–6.18 CHCH), 4.71 , 3.74 , 3.73 , and 3.35 . Characterization data for this compound are in complete agreement with an internal GE publication.Ethyl 3‐((2‐(benzyloxy)ethyl)(2‐chloro‐5‐methoxyphenyl)amino)‐2‐hydroxycyclohex‐1‐ene‐1‐carboxylate (19). The aniline 18  was stirred in dry THF (150 mL) at −40 °C under nitrogen and potassium bis(trimethylsilyl) amide  was added over 30 min. The carboxylate 11  in dry THF (50 mL) was then added and allowed to warm to RT over a period of 1.5 h. Acetic acid was added (15 mL) and concentrated in vacuo to remove the THF. EtOAc (200 mL) and 10 % aqueous potassium carbonate (200 mL) was added and the mixture vigorously shaken. The EtOAc solution was separated, dried over (MgSO4) and concentrated in vacuo to afford 19 as an oil , which was used crude in the next step. LC–MS: Rf 2.39 (−ESI) m/z 458.5 ([M−H]−), Rf 2.53 (+ESI) m/z 460.5 ([M+H]+), Rf 2.39 (+ESI) m/z 460.5 ([M+H]+). Characterization data for this compound are in complete agreement with an internal GE publication.H‐carbazole‐4‐carboxylate (20)Ethyl 9‐(2‐(benzyloxy)ethyl)‐8‐chloro‐5‐methoxy‐2,3,4,9‐tetrahydro‐1. Zinc chloride  was added to carboxylate 19  in dry Et2O (400 mL) under nitrogen and heated at reflux for 5.5 h. As the reaction was held at reflux, a thick brown dense oil formed in the reaction. The reaction was then cooled and the supernatant Et2O decanted off, EtOAc (300 mL) was added, washed with 2 m HCl (150 mL) and with 10 % aqueous potassium carbonate (150 mL). The EtOAc layer was separated, dried (MgSO4) and concentrated in vacuo to afford an oil. The crude material was purified by silica gel chromatography eluting with a gradient of 10–40 % EtOAc/petroleum ether (40–60 °C) to afford 20 as an oil . The thick dense brown layer was treated with EtOAc (300 mL) and 2 m HCl (150 mL). The EtOAc layer was separated, washed with 10 % aqueous potassium carbonate (150 mL), dried (MgSO4) and concentrated in vacuo to give an oil. Et2O (400 mL) and anhydrous zinc chloride  were added. The mixture was heated at reflux for a further 5 days. The Et2O layer and the dark gum were both diluted with EtOAc (200 mL) and then washed with 2 m HCl (150 mL), dried (MgSO4) and concentrated in vacuo to give a gum. This gum was purified by silica gel chromatography eluting with a gradient of 5–35 % EtOAc/petroleum ether (40–60 °C) to afford 20 as an oil . LC–MS: Rf 2.66 (+ESI) m/z 442.5 ([M+H]+); 1H NMR : δH=7.31–7.16 , 6.93 , 6.34 , 4.58 , 4.40 , 4.20–4.09 , 3.86–3.80 , 3.79 , 2.82–2.63 , 2.10–1.80 , and 1.23 . Characterization data for this compound are in complete agreement with an internal GE publication.H‐carbazole‐4‐carboxylic acid (21)9‐(2‐(Benzyloxy)ethyl)‐8‐chloro‐5‐methoxy‐2,3,4,9‐tetrahydro‐1. To the ester 20  in ethanol (12 mL) was added sodium hydroxide  in water (1 mL) and heated at 80 °C for 18 h. The ethanol was then removed by evaporation in vacuo and the residue partitioned between Et2O (30 mL) and water (30 mL). The Et2O layer was separated, dried (MgSO4) and concentrated in vacuo to give a gum. The aqueous layer was acidified to pH 1 with 2 m HCl dropwise and extracted with CH2Cl2 (3×20 mL). The organic layers were dried (MgSO4) and concentrated in vacuo to afford 21 as a solid . LC–MS: Rf 2.34 (+ESI) m/z 414.4 ([M+H]+); 1H NMR : δH=7.31–7.15 , 6.97 , 6.41 , 4.68–4.52 , 4.40 , 4.18 , 3.88 , 3.82 , 2.86–2.63 , and 2.27–1.86 . Characterization data for this compound are in complete agreement with an internal GE publication.NN‐ethyl‐5‐methoxy‐2,3,4,9‐tetrahydro‐1H‐carbazole‐4‐carboxamide (22)‐Benzyl‐9‐(2‐(benzyloxy)ethyl)‐8‐chloro‐. The acid 21  was dissolved in THF (135 mL) under N2 at RT. The solution was allowed to cool to 0 °C and N‐ethylbenzylamine  was added followed by 1‐hydroxybenzotriazole hydrate  and N‐(3‐dimethylaminopropyl)‐N′‐ethylcarbodiimide hydrochloride . Finally, triethylamine  was added via syringe and the mixture stirred under N2 and warmed to RT over 48 h. The reaction was then diluted with EtOAc (100 mL) and filtered through Celite. The Celite was washed with more EtOAc (2×100 mL). The combined filtrates were washed with 1 m aqueous HCl (2×150 mL), H2O (2×150 mL), brine (150 mL), dried (MgSO4) and concentrated in vacuo to give the product 22 as an oil , which was used crude in the next step. LC–MS: Rf 2.74 (+ESI) m/z 531.6 ([M+H]+).NN‐ethyl‐5‐methoxy‐2,3,4,9‐tetrahydro‐1H‐carbazole‐4‐carboxamide (23)‐Benzyl‐9‐(2‐(benzyloxy)ethyl)‐. The carboxamide 22  in methanol (120 mL) was shaken with 10 % palladium on charcoal (1.5 g), triethylamine  and stirred for 18 h at RT after purging twice (over 2 h) with hydrogen gas under atmospheric pressure. The reaction was then filtered through a pad of Celite under nitrogen atmosphere and the filtrate concentrated in vacuo. The concentrate was then taken up in CH2Cl2 (200 mL) and washed with 5 % aqueous potassium carbonate solution (200 mL). The CH2Cl2 solution was then separated, dried (MgSO4) and concentrated in vacuo to afford 23 as an oil , which was used crude in the next step. LC–MS: Rf 2.62 (+ESI) m/z 497.5 ([M+H]+).NN‐ethyl‐9‐(2‐hydroxyethyl)‐5‐methoxy‐2,3,4,9‐tetrahydro‐1H‐carbazole‐4‐carboxamide (24)‐Benzyl‐. The benzyl‐protected intermediate 23  in methanol (50 mL) was shaken with 10 % palladium on charcoal (0.333 g) and stirred for 18 h at RT after purging twice (over 2 h) with hydrogen gas under atmospheric pressure. The reaction was then filtered through a pad of Celite under nitrogen atmosphere and the filtrate concentrated in vacuo to give 24 as an oil , which was used crude in the next step. LC–MS: Rf 2.08 (+ESI) m/z 407.5 ([M+H]+).H‐carbazol‐9‐yl)ethyl4‐methylbenzenesulfonate (25)2‐(4‐(Benzyl(ethyl)carbamoyl)‐5‐methoxy‐1,2,3,4‐tetrahydro‐9. The alcohol 24  was dissolved in anhydrous pyridine (5 mL) under N2. The solution was cooled to 0 °C and 4‐toluenesulfonyl chloride  added potion‐wise to the solution over 30 min. 4‐Dimethylaminopyridine  was then added, and the solution was stirred and warmed to RT for 18 h. The reaction was then quenched by careful addition of ice followed by water (5 mL). The mixture was then extracted into EtOAc and washed with water (3×10 mL). Excess pyridine was removed by washing with 1 m HCl (2×10 mL) and aqueous copper sulfate (2×10 mL). Excess 4‐toluenesulfonyl chloride was removed by washing with 1 m aqueous sodium carbonate (2×10 mL). The organic layer was washed with brine (10 mL), dried (MgSO4) and concentrated in vacuo. The crude product was recrystallised from EtOAc to afford 25 as an off‐white solid . LC–MS: Rf 2.46 (+ESI) m/z 561.6 ([M+H]+); 1H NMR : δH=7.51 (CH2)CH3), 7.35–7.15 , 7.09 (CH2)CH3), 6.91 , 6.62 , 6.37 , 4.70–4.50 , 4.30–4.15 , 3.61 , 3.80–3.40 2), 2.80–2.50 , 2.33 , 2.1–1.60 , and 1.30 .H‐carbazol‐9‐yl)ethyl methanesulfonate (26)2‐(4‐(Benzyl(ethyl)carbamoyl)‐5‐methoxy‐1,2,3,4‐tetrahydro‐9. The phenol 24  in CH2Cl2 (20 mL) was cooled to 0 °C and methanesulfonyl chloride  and triethylamine  were added and allowed to warm to RT for 18 h. The reaction was diluted with CH2Cl2 (20 mL) washed with 5 % aqueous potassium carbonate solution (40 mL). The layers were separated. The combined organic layers were dried (MgSO4) and concentrated in vacuo to give a gum. The crude material was purified by silica gel flash chromatography eluting with a gradient of 50–100 % EtOAc/petroleum ether (40–60 °C) to afford 26 as an oil . LC–MS: Rf 2.12 (+ESI) m/z 485.5 ([M+H]+). 1H NMR : δH=7.40–7.10 , 7.03 , 6.86 , 6.44 , 4.65–4.50 , 4.50–4.45 , 3.64 , 3.90–3.20 2), 2.90–2.60 , 2.51 , 2.30–1.70 , and 1.31 .R,2S,5R)‐2‐Isopropyl‐5‐methylcyclohexyl 9‐(2‐(benzyloxy)ethyl)‐8‐chloro‐5‐methoxy‐2,3,4,9‐tetrahydro‐1H‐carbazole‐4‐carboxylate (29) and l‐menthol in CH2Cl2 (40 mL) at 0 °C was added N‐(3‐Dimethylaminopropyl)‐N′‐ethylcarbodiimide hydrochloride  and 4‐Dimethylaminopyridine . Stirring was continued at RT for 16 h. After this period, H2O (100 mL) and CH2Cl2 (100 mL) were added. The organic layer was washed with 1 m HCl, saturated NaHCO3 solution, and brine. The solution was dried (MgSO4), filtered, and concentrated. The crude racemic mixture 29  was separated into two diasteriomerically enriched samples via silica flash chromatography eluting with 30 % Et2O/cyclohexane. LC–MS: Rf 3.12 (+ESI) m/z 368.3 ; 1H NMR : δH=7.36–7.12 , 6.94 , 6.31 , 4.90 , 4.38 , 4.09 , 3.86–3.77 , 3.75 , 2.7–2.44 , 2.15–2.01 , 2.00–1.82 , 1.81–1.56 , 1.46–1.40 2, and 1.02–0.83 .R)‐1‐Phenylprop‐2‐yn‐1‐yl 9‐(2‐(benzyloxy)ethyl)‐8‐chloro‐5‐methoxy‐2,3,4,9‐tetrahydro‐1H‐carbazole‐4‐carboxylate (30), N‐(3‐Dimethylaminopropyl)‐N′‐ethylcarbodiimide hydrochloride  and 4‐Dimethylaminopyridine  in CH2Cl2 (5 mL) was added (R)‐1‐phenyl‐2‐propyn‐1‐ol  at RT. The reaction was stirred for 4 h at RT. The reaction mixture was diluted with CH2Cl2 (20 mL) and washed with H2O (5 mL). The organic layer was dried (MgSO4) and concentrated. The crude mixture was purified via silica flash column chromatography eluting with a gradient of 20–40 % Et2O/petroleum ether (40–60 °C) to yield 30 as a racemate . Separation of the two diastereomer was done via silica flash column chromatography eluting with a gradient of 10–20 % Et2O/cyclohexane. LC–MS: Rf 2.72 (+ESI) m/z 528.5 , 2.73 (+ESI) m/z 528.5 ; 1H NMR : δH=7.60–7.14 , 6.90 , 6.25 , 4.71–4.53 , 4.39 , 4.15 , 3.81 , 3.40 , 2.85–2.51 , 2.60 , and 2.20–1.79 .NN‐ethyl‐9‐(2‐fluoroethyl)‐5‐methoxy‐2,3,4,9‐tetrahydro‐1H‐carbazole‐4‐carboxamide (6)‐Benzyl‐. 49.8 mg of a racemic mixture of 6 was sent to Reach Separations to be separated into the two enantiomers. They provided the following methods for separation and purification. The racemate 6 was dissolved to 12 mg mL−1 in MeOH/CH2Cl2 (1:1) and was then purified by SFC using a Lux C1 column  at 40 °C and eluting with isocratic 50:50 MeOH/CO2 with 0.2 % v/v NH3 at a flow rate of 50 mL min−1. The injection volume was 1500 μL and peaks were detected at 223 nm. Combined fractions of each of the two enantiomers were then evaporated to near dryness using a rotary evaporator, transferred into final vessels with CH2Cl2, which was removed under a stream of compressed air at 40 °C before being stored in a vacuum oven at 40 °C and 5 mbar for 4 h to afford the two enantiomers as yellow gums. Chiral purity of the enantiomers was analysed using a Lux C1 column  at 40 °C and eluting with isocratic 50:50 MeOH/CO2 with 0.2 % v/v NH3 at a flow rate of 4 mL min−1. The injection volume was 1.0 μL and the peaks were detected at 210–400 nm. Chemical purity of the enantiomers was analysed using an Acquity BEH C18 column  at 40 °C and eluting with 0–4 min gradient of 5–95 % MeCN/H2O with 0.1 % TFA, 4‐4.02 min gradient of 95–100 % MeCN/H2O with 0.1 % TFA, 4.03 min–4.5 min isocratic 100 % MeCN with 0.1 % TFA, 4.5–4.52 min gradient of 100–5 % MeCN/H2O with 0.1 % TFA, and 4.53–6 min isocratic of 5 % MeCN/H2O with 0.1 % TFA at flow rate of 0.6 mL min−1. The injection volume was 1.0 μL and the peak was detected at 220 nm. The final yield is 23.9 mg of enantiomer (S)‐6 with 98.02 % chemical purity and in 100 % ee, and 22.1 mg of enantiomer (R)‐6 with 100 % chemical purity and in 98.8 % ee.H‐carbazol‐9‐yl)ethyl 4‐methylbenzenesulfonate (25)2‐(4‐(Benzyl(ethyl)carbamoyl)‐5‐methoxy‐1,2,3,4‐tetrahydro‐9. 162.4 mg of a racemic mixture of 25 was sent to Reach Separations to be separated into the two enantiomers. They provided the following methods for separation and purification. The racemate 25 was dissolved to 23 mg mL−1 in MeOH/CH2Cl2 (1:1) and was then purified by SFC using a Lux C1 column  at 40 °C and eluting with isocratic 50:50 MeOH/CO2 with 0.2 % v/v NH3 at a flow rate of 50 mL min−1. The injection volume was 1000 μL and peaks were detected at 223 nm. Combined fractions of each of the two enantiomers were then evaporated to near dryness using a rotary evaporator, transferred into final vessels with CH2Cl2, which was removed under a stream of compressed air at 40 °C before being stored in a vacuum oven at 40 °C and 5 mbar for 4 h to afford the two enantiomers as white solids. Chiral purity of the enantiomers was analysed using a Lux C1 column  at 40 °C and eluting with isocratic 50:50 MeOH/CO2 with 0.2 % v/v NH3 at a flow rate of 4 mL min−1. The injection volume was 1.0 μL and the peaks were detected at 210–400 nm. Chemical purity of the enantiomers was analysed using an Acquity BEH C18 column  at 40 °C and eluting with 0–4 min gradient of 5–95 % MeCN/H2O with 0.1 % TFA, 4‐4.02 min gradient of 95–100 % MeCN/H2O with 0.1 % TFA, 4.03 min–4.5 min isocratic 100 % MeCN with 0.1 % TFA, 4.5–4.52 min gradient of 100‐5 % MeCN/H2O with 0.1 % TFA, and 4.53–6 min isocratic of 5 % MeCN/H2O with 0.1 % TFA at flow rate 0.6 mL min−1. The injection volume was 1.0 μL and the peak was detected at 220 nm. The final yield is 82.7 mg of enantiomer (S)‐25 with 97.42 % chemical purity and in 98.2 % ee, and 77.1 mg of enantiomer (R)‐25 with 98.75 % chemical purity and in 99.6 % ee.m Tris⋅HCl, pH 7.4) were incubated with ≈Kd concentration of [3H]1  and test compounds (0.3 nm–10 μm) at 4 °C for 90 min. Reactions were terminated by filtration through a 96‐well glass‐fibre filter plate (Millipore), and washed eight times with ice‐cold 50 mm Tris⋅HCl. Microscint 0 was added to the dry filters, and radioactivity read in a Microbeta2 2450 Microplate Counter (PerkinElmer). Data were analysed using GraphPad Prism 6.0, and a four‐parameter nonlinear regression curve fit was used to calculate Ki values. Data are expressed as the mean±SEM from at least three independent experiments.Binding affinities were measured as per our published protocol.Circular dichroism spectroscopy. Solutions of (S)‐6, (R)‐6, and (S)‐5 were prepared in concentrations of 0.1–0.2 mg mL−1 in acetonitrile and CD spectra were collected on an Aviv 410 instrument.Radioisotope production. No‐carrier‐added aqueous [18F]fluoride ion was produced on a GE PETtrace cyclotron by irradiation of a 2.3‐mL silver‐bodied water target with a 25‐mA current and a 16.5‐MeV proton beam on 95 % enriched 18O‐H2O via the nuclear 18O18F reaction.18F]KF‐Kryptofix‐222 complexPreparation of the [. [18F]Fluoride in [18O]H2O was transferred and immediately trapped on an anion‐exchange resin (Waters Sep‐Pak Accell Light QMA cartridge in the carbonate form) under vacuum. Trapped 18F‐fluoride was eluted from the Sep‐Pak cartridge and transferred to the reaction vessel with an eluent solution containing 0.25 % wild‐type Kryptofix‐222® solution (1 mL) in basic (0.05 % wild‐type K2CO3) aq. MeCN (75 % v/v). The solvents were evaporated in vacuo (130 mbar) with a stream of N2 gas at 95 °C over 5 min. Anhydrous MeCN (3×0.7 mL) was then added and the mixture was azeotropically dried in vacuo (130 mbar) with a stream of N2 at 95 °C.18F]6Manual preparation of fluoride solution (125 μL) of equivalent solvent. The vials were heated in a heating block to the desired temperature  and a single vial was removed at each time point  and quenched with 1:1 MeCN/H2O (300 μL) before analyzing directly by HPLC using a gradient method: (pre‐injection) 1 min equilibration at 10:90 MeCN/H2O. (post‐injection) 1 min isocratic at 10:90 MeCN/H2O, 2 min gradient to 70:30 MeCN/H2O, 3 min isocratic at 70:30 MeCN/H2O. Flow rate: 1.5 mL min−1. Run time: 6 min (7 min including equilibration time). Column: ACE® UltraCore, Super C18, 2.5 μm, 50×4.6 mm; Serial No. A133239. A portion of the sample from 20 min was taken and used for co‐injection with reference compound 6 to confirm synthesis of desired product.18F]6Automated preparation and formulation of KF‐Kryptofix‐222 complex and heated at 100 °C for 20 min. The reaction mixture was then cooled with compressed air and diluted with anhydrous MeCN (1 mL). The crude mixture was injected into an ACE 160433 C18 5 μm (100×10 mm) semipreparative reversed‐phase HPLC column. With a mobile phase of 48 % aq. MeCN at a flow rate of 5 mL min−1. [18F]6 was collected (tR: 19.6 min) and immediately diluted with H2O (10 mL). The aqueous solution was passed through a C18 cartridge . The cartridge was washed with H2O (15 mL) and the radiolabelled product was eluted with EtOH (0.5 mL). The radiolabelled product was then formulated with 0.9 % w/v aq. NaCl in a vial to afford the radiolabelled title compound.18F]6Quality control for 6 at a tR of 9.8 min. The area of the UV absorbance peak at 220 nm corresponding to the carrier product was measured (integrated) on the HPLC chromatogram and compared with a standard curve relating mass to UV absorbance. The identity of the radiolabelled product was confirmed by HPLC co‐injection of the reference compound 6.n=2; weight: 230 and 302 g) were obtained from Charles River Laboratories, UK. The rats were housed in Techniplast 2000P IVC cages on a layer of Aspen bedding in a room with constant temperature (21±2 °C) and fixed 12 h light–dark regime (lights on at 7:00 am). Food and water were available ad libitum. After arrival, the rats were allowed to acclimatise for at least seven days. This research was regulated under the Animals (Scientific Procedures) Act 1986 Amendment Regulations 2012 following ethical review by the University of Cambridge Animal Welfare and Ethical Review Body (AWERB).Male Wistar rats .68Ge point source was made for attenuation and scatter correction of 511 keV photons. The radiotracer, formulated in saline, was injected to the rats and the emission scan was started with tracer injection. A list‐mode protocol was used with an acquisition time of 60 min.Prior to scanning, the animals were anaesthetised using isofluorane at a concentration of 5 % in OImage reconstructions were performed using microPET Manager 2.4.1.1, ASIPro 6.7.1.2 (Siemens). Acquisition data were then Fourier re‐binned in 22 time frames  and the data were reconstructed per timeframe employing an iterative reconstruction algorithm . The final datasets consisted of 95 slices with a slice thickness of 0.8 mm, and an in‐plane image matrix of 128×128 pixels. Voxel size was 0.8×0.8×0.8 mm. Data sets were corrected for decay, random coincidences, scatter and attenuation.−1 for brain tissue.Three‐dimensional regions of interest (ROIs) were drawn over the whole brain on an MRI template using PMOD software . PET images were then co‐registered with this MRI template and the regions of interest transferred from MRI to PET. Whole‐brain TACs were then obtained for each of the animals. The results were expressed as dimensionless standardised uptake values (SUV=[(tissue activity)×(body weight)]/injected dose). SUVs were calculated assuming a specific gravity of 1 g mLThe authors declare no conflict of interest.As a service to our authors and readers, this journal provides supporting information supplied by the authors. Such materials are peer reviewed and may be re‐organized for online delivery, but are not copy‐edited or typeset. Technical support issues arising from supporting information (other than missing files) should be addressed to the authors.SupplementaryClick here for additional data file.


The propynyl substituent is nearly perpendicular to the plane formed by the rails of the boat. In the crystal, inversion dimers are formed by weak C—H⋯F hydrogen bonds with the dimers forming oblique stacks along the  18H12FNOS, is built up from a 4-fluorobenzylidene moiety and a dihydrobenzothiazine unit with a propynyl substituent, with the heterocyclic portion of the dihydrobenzothiazine unit adopting a shallow boat conformation with the propynyl substituent nearly perpendicular to it. The two benzene rings are oriented at a dihedral angle of 43.02 (6)°. In the crystal, C—HFlurphen⋯FFlurphen (Flurphen = fluorophenyl) hydrogen bonds link the molecules into inversion dimers, enclosing R22(8) ring motifs, with the dimers forming oblique stacks along the a-axis direction. Hirshfeld surface analysis of the crystal structure indicates that the most important contributions to the crystal packing are from H⋯H (33.9%), H⋯C/C⋯H (26.7%), H⋯F/F⋯H (10.9%) and C⋯C (10.6%) interactions. Hydrogen bonding and van der Waals interactions are the dominant interactions in the crystal packing. Density functional theory (DFT) optimized structures at the B3LYP/6–311 G level are compared with the experimentally determined molecular structure in the solid state. The HOMO–LUMO behaviour was elucidated to determine the energy gap.The title compound, C The propynyl substituent is nearly perpendicular to the plane defined by C1, C6, C7 and C8, as shown by the C6—N1—C9—C10 torsion angle of 81.3 (2)°. A puckering analysis of the heterocyclic ring  of the dihydrobenzothiazine unit shows that it adopts a shallow boat conformation with puckering parameters QT = 0.3759 (14) Å, q2 = 0.3639 (15) Å, q3 = −0.0938 (17) Å, φ = 173.6 (3)° and θ = 104.5 (3)°. In the heterocyclic ring B, the C1—S1—C8 [101.73 (8)°], S1—C8—C7 [119.93 (12)°], C8—C7—N1 [119.23 (14)°], C7—N1—C6 [125.59 (14)°] and C6—C1—S1 [122.07 (13)°] bond angles are enlarged, while the N1—C6—C1 [120.91 (15)°] bond angle is narrowed when compared with the corresponding values in the closely related compounds 4-methyl-3,4-dihydro-2H-1,4-benzothiazin-3-one, (II) -one, (III) -2-(2-chlorobenzylidene)-4-(prop-2-ynyl)-2H-1,4-benzothiazin-3(4H)-one, (IV), Flurphen⋯FFlurphen (Flurphen = fluorophenyl) hydrogen bonds  analysis  have a nearly symmetrical distribution of points, Fig. 7c, with the thick edges at de + di ∼2.70 Å. The pair of characteristic wings in the fingerprint plot delineated into H⋯F/F⋯H contacts  arises from the C—H⋯F hydrogen bonds  have an arrow-shaped distribution of points with the tip at de = di ∼1.68 Å. The pair of characteristic wings in the fingerprint plot delineated into H⋯O/O⋯H contacts  have a pair of spikes with the tips at de + di = 2.54 Å. Finally, the H⋯S/S⋯H contacts  are viewed as A pair of wide spikes with the tips at de + di = 3.02 Å. The Hirshfeld surface representations with the function dnorm plotted onto the surface are shown for the H⋯H, H⋯C/C⋯H, H⋯F/F⋯H, C⋯C, H⋯O/O⋯H and H⋯S/S⋯H interactions in Fig. 8a–f, respectively.The overall two-dimensional fingerprint plot, Fig. 7s Table 1 as well s Table 1 and is set al., 2015The Hirshfeld surface analysis confirms the importance of H-atom contacts in establishing the packing. The large number of H⋯H, H⋯C/C⋯H and H⋯O/O⋯H interactions suggest that van der Waals interactions and hydrogen bonding play the major roles in the crystal packing  using standard B3LYP functional and 6–311G basis-set calculations -2-(4-fluorobenzylidene)-4-(prop-2-ynyl)-2H-1,4-benzothiazin-3(4H)-one ring. The energy band gap [ΔE = ELUMO - EHOMO] of the molecule was about 3.92 eV, and the frontier molecular orbital energies, EHOMO and ELUMO were −5.85 and −1.93 eV, respectively.The optimized structure of the title compound, (I)II  in the Cambridge Crystallographic Database  to 36° (IId). The other three  have the benzothiazine unit nearly planar with the corresponding dihedral angle of ca 3–4°. In the case of IIa, the displacement ellipsoid for the sulfur atom shows a considerable elongation perpendicular to the mean plane of the heterocyclic ring, suggesting disorder, and a greater degree of non-planarity but for the other two, there is no obvious source for the near planarity.Using the search fragment Z)-2-(4-fluorobenzylidene)-2H-1,4-benzothiazin-3(4H)-one (1.6 mmol), potassium carbonate (4 mmol) and tetra-n-butyl ammonium bromide (0.15 mmol) in DMF (20 ml). Stirring was continued at room temperature for 24 h. The salts were removed by filtration and the filtrate was concentrated under reduced pressure. The residue was separated by chromatography on a column of silica gel with ethyl acetate–hexane (2/8) as eluent. The solid product obtained was recrystallized from ethanol to afford colourless crystals (yield: 89%).Propargyl bromide (4 mmol) was added to a mixture of  I, global. DOI: 10.1107/S2056989019002354/lh5893Isup2.hklStructure factors: contains datablock(s) I. DOI: Click here for additional data file.10.1107/S2056989019002354/lh5893Isup3.cdxSupporting information file. DOI: Click here for additional data file.10.1107/S2056989019002354/lh5893Isup6.cmlSupporting information file. DOI: 1897371CCDC reference:  crystallographic information; 3D view; checkCIF reportAdditional supporting information:


In the title hydrated azo dye, the benzene and thiazole make a dihedral angle of 4.69 (17)°. In the crystal, hydrogen bonds, C—H⋯π and π–π interactions resulting in the formation of a three-dimensional framework. 10H10N4OS·H2O, the benzene and thiazole, are nearly coplanar, with a dihedral angle between their mean planes of 4.69 (17)°. The aromatic rings on the –N=N– moiety exhibit a trans configuration. The crystal structure features many types of intermolecular interactions involving all the functional groups – strong hydrogen bonds (N⋯H and O⋯H), weak hydrogen bonds (C—H⋯O and C—H⋯N), C—H⋯π and π–π interactions – resulting in the formation of a three-dimensional framework.In the title hydrated azo dye, C Azo dyes have wide range applications in the cosmetic, food, textile industry, chemical sensing, and pharmaceutical . The methoxy and amino groups on the benzene ring are co-planar with the ring with atoms O1 and N4 deviating by −0.010 (2) and −0.019 (4) Å, respectively. The dihedral angle between the thiazole and benzene rings is 4.69 (17)°, nearly coplanar.The molecular structure of (I)a, Table 12), azo (–N=N–) and thiazole groups and the water molecule of crystallization  are the primary interactions responsible for the formation of the three dimensional structure. In addition, the crystal structure is supported by other intermolecular interactions as a secondary weak interactions, C—H⋯X (X = O and N), C—H⋯π and offset π—π interactions. The weak hydrogen bonds are formed between the C—H moieties in the benzene and thiazole rings with amine, azo, methoxy groups of adjacent molecules and water molecules . On the shape index surface , convex blue regions represent hydrogen-donor groups and concave red regions represent hydrogen-acceptor groups. In addition, concave red regions represent C—H⋯π and offset π–π interactions. The amino group behaves as both a donor and an acceptor. The methyl part of the methoxy group acts as a donor while the oxygen atom is an acceptor.Hirshfeld surfaces and fingerprint plots were generated using ce Fig. 3b, conveet al., 2007The two-dimensional fingerprint plots Fig. 4 quantifet al., 1992trans configuration and the phenolic oxygen atom is linked to an azo nitrogen atom by intramolecular hydrogen bonding. The crystal structure features only van der Waals interactions. To form complexes with metal ions, both thiazole and naphthol rings are rotated by 180° to coordinate to the metal through the phenolic oxygen atom, the azo nitrogen atom adjacent to the naphthol ring and the thiazole nitrogen atom, resulting the formation of five-membered chelate rings. Complexes of TAR and TAC are formed in a similar way due to the presence of a hydroxyl group in the structure M HCl (16 ml), and 8.236 mmol of sodium nitrate solution was added slowly under stirring at low temperature 268–273 K until the diazonium salt was obtained. m-Anisidine (1.12 ml in 40 ml of 4 M HCl) was slowly dropped into the mixture and stirred at a temperature between 268 and 273 K for 1 h. After the reaction was complete, conc. NH3 was dropped into the mixture (pH 6) until the red–orange crude produce appeared. The products were filtered, washed with cold water, purified by column chromatography and recrystallized from an acetonitrile–water (1:1) mixture by vapour diffusion.2-Aminothiazole (9.986 mmol) was dissolved in 6 1H NMR : δ 3.806 , 6.364  , 6.374 , 7.546  ,7.629 , 7.697 , 7.883 . Mass spectroscopy: m/z 235.0654 [C10H11N4OS+], 205.0548 [C9H9N4S.+], 150.0662 [C7H8N3O.+], 122.0601 [C7H8NO.+]. IR (KBr cm−1): 3,413 cm−1 ; 821 cm−1 ; 1,617 ; 1,222 cm−1 ; 1,103 cm−1 ; 1,152 cm−1 ; 1,541cm−1 ; 1,021 cm−1 . Elemental analysis calculated for C10H10N4OS·H2O: C, 51.27; H, 4.30; N, 23.92. Found: C, 51.34; H, 4.20; N, 23.98.Uiso = 1.2 Ueq(C-aromatic) and 1.5Ueq (C-methyl).Crystal data, data collection and structure refinement details are summarized in Table 210.1107/S205698901900207X/dx2014sup1.cifCrystal structure: contains datablock(s) global, I. DOI: 10.1107/S205698901900207X/dx2014Isup2.hklStructure factors: contains datablock(s) I. DOI: 1895710CCDC reference:  crystallographic information; 3D view; checkCIF reportAdditional supporting information:


The structure has triclinic (P2(O2CCPh3-μ-κO:κO′)]2 is located about an inversion centre. The triphenylacetate ligand displays a μ-κO:κO′-bridging coordination mode, leading to the formation of an octagonal Al2O4C2 core. The complex displays HPh⋯CPh intermolecular interactions.Single crystals of the title compound, [Al(iBu) The triphenylacetate ligand exhibits a μ2-κO:κO′-bridging coordination mode. The CPh—CPh [1.3788 (17) Å for C5—C6 to 1.4016 (14) Å for C15—C16], CiBu—CiBu [1.523 (2) Å for C26—C28 to 1.5381 (16) Å for C21—C22], C1—C2 [1.5473 (13) Å] and C1—Cipso [1.5425 (14) Å for C2—C9 to 1.5455 (14) Å for C2—C3] bond lengths inside the ligands are within the expected ranges. The complex has a nearly flat eight-membered Al2O4C2 core, with the greatest deviations from the plane being 0.0548 (6) Å for the O2 and O2i atoms. The bond angles inside the core are 106.84 (4) (O1—Al—O2i), 151.00 (7) (Al—O1—C1), 123.64 (9) (O1—C1—O2) and 156.79 (8)° (C1—O2—Ali), summing to a value of 1076.54° for the entire core, which deviates from a flat octagon by 3.46°. The Al—X bond lengths are given in Table 1R2(μ-O2CR′)]2 compounds (see §4 below) having the Al2O4C2 core (23 independent core fragments) have Al—X bond lengths varying from ca 1.77 to 1.86 Å (average 1.82 Å) for Al—O, 1.92–2.00 Å (average 1.96 Å) for Al—C and 1.23–1.29 Å (average 1.26 Å) for C—O bonds. The bond lengths in the title complex (Table 12-bridging RO− ligands), but shorter than the Al—O distances in complexes with either Al–O=CR2  or Al—Oether fragments  due to different types of Al—O interactions, changing from the ion–ion type in the case of Al—Oalkyl/aryl bonds to the ion–dipole one in the case of Al—O=CR2 or Al—Oether fragments.The title compound crystallizes in the triclinic space group ]2 Fig. 2 located x Table 1 are closA⋯H20 and 2.30 Å for H25A⋯H12. Two intermolecular interactions involving aromatic H atoms with the π-system of a arene group have been found, i.e. 2.89 Å for H6⋯C12 and 2.98 Å for H7⋯C11 (see Table S1 for details). Interacting arene rings are located nearly perpendicular to one another, with the corresponding angle between the C3–C8 and C9–C14 planes being 82.83 (3)° .The crystal lattice exhibits weak intermolecular van der Waals contacts between methyl or methylene and aromatic H atoms, with the distances being 2.49 Å for H23)° Fig. 3. The laset al., 20162O4C2 core and having the [AlR2(μ-O2CR′)]2 motif, where R is alkyl or C6F5. Records for crystal structures with other R groups connected to Al via the C atom have not been found in the CSD. 14 complexes have bridging carboxylate ligands, the others have a heteroatom in the α-position .According to the Cambridge Structural Database ]2 type are represented by structures with R = R′ = Me and aryl = Ph 3 and aryl = 4-MeC6H4  and R = isobutyl (iBu)  ]2 complexes with a substituted acetate anion possess R = Et and CX3 = CPh3 ]2, with X = –C4H(CH3)2Zr(η5-C5Me5)2 2(μ-O2CC6F5)]2 ]2 ]2, with R = iBu  depends greatly on various interactions within the complex, including nonvalence ones. See also related ab initio calculations in the literature 3(THF)3] was prepared according to a previously published method  was added dropwise to a suspension of [Nd(Ph3CCOO)3(THF)3]  in 15 ml of hexane at room temperature. The suspension dissolved within a few minutes upon addition. The resulting solution was stirred overnight at room temperature. Crystals of [Al(iBu)2(Ph3CCOO)]2 were isolated from the reaction mixture by crystallization at 243 K for 2 d. The mother liquor was decanted and crystals were dried under dynamic vacuum. The yield was 56 mg . Calculated for C56H66Al2O4 (%): C 78.48, H 7.76; found: C 78.17, H 8.01.A solution of Al(iBu)Uiso(H) = 1.5Ueq(C) for methyl H atoms and 1.2Ueq(C) otherwise. A rotating group model was applied for methyl groups.Crystal data, data collection and structure refinement details are summarized in Table 210.1107/S2056989019003396/tx2010sup1.cifCrystal structure: contains datablock(s) global. DOI: 10.1107/S2056989019003396/tx2010sup4.pdfC...H interactions and packing plots. DOI: 1902149CCDC reference:  crystallographic information; 3D view; checkCIF reportAdditional supporting information:


In the crystal. hydroxyl-O—H⋯O(hydroxyl) and hydroxyl-O—H⋯S(thione) hydrogen bonds give rise to a supramolecular layer in the ab plane.The title tri-substituted thiourea derivative is twisted with a dihedral angle of 72.12 (9)° between the planes through the CN 13H18N2O3S, the thione-S and carbonyl-O atoms lie, to a first approximation, to the same side of the molecule [the S—C—N—C torsion angle is −49.3 (2)°]. The CN2S plane is almost planar (r.m.s. deviation = 0.018 Å) with the hydroxyethyl groups lying to either side of this plane. One hydroxyethyl group is orientated towards the thioamide functionality enabling the formation of an intramolecular N—H⋯O hydrogen bond leading to an S(7) loop. The dihedral angle [72.12 (9)°] between the planes through the CN2S atoms and the 4-tolyl ring indicates the molecule is twisted. The experimental molecular structure is close to the gas-phase, geometry-optimized structure calculated by DFT methods. In the molecular packing, hydroxyl-O—H⋯O(hydroxyl) and hydroxyl-O—H⋯S(thione) hydrogen bonds lead to the formation of a supramolecular layer in the ab plane; no directional interactions are found between layers. The influence of the specified supramolecular interactions is apparent in the calculated Hirshfeld surfaces and these are shown to be attractive in non-covalent interaction plots; the interaction energies point to the important stabilization provided by directional O—H⋯O hydrogen bonds.In the title tri-substituted thiourea derivative, C R1(R2)NC(=S)N(R3)R4 for R1–4 = alkyl/aryl. The present study concerns a tri-substituted example, i.e. an N,N′-di-N′-benzoylthiourea derivative, notable for having a carbonyl group connected to the thiourea framework. Thiourea molecules are of interest in themselves and as ligands for metal ions 2, (I)The amine-H atoms in thiourea, Hsyn as the S1—C1—N2—C6 torsion angle is −49.3 (2)°; the O3—C6—N2—C1 torsion angle is −6.8 (3)°. The hydroxyethyl groups lie to either side of the CN2S plane (r.m.s. deviation = 0.018 Å). The O1-hydroxyethyl group is folded toward the thioamide part of the molecule, an orientation that allows for the formation of an intramolecular N2—H⋯O1 hydrogen bond that closes an S(7) loop, Table 22S atoms and the terminal aryl ring. The C1—N1 bond length is considerably shorter than the C1—N2 bond, which suggests some delocalization of π-electron density over the S1—C1—N1 atoms that does not extend over the C1—N1—C6 atoms, consistent with the large twist about the C1—N2 bond (see above). The bond angles subtended at the C1 and C6 atoms follow the expected trends in that those involving the formally doubly bonded atoms are wider, by approximately 10°, compared with the other angles, Table 1The title compound, (I)d,p) basis set  N1—C2—C3—O1 and N1—C4—C5—O2 torsion angles, which are disparate, by about 12°, in the experimental structure but are symmetric, i.e. ±69°, in the optimized structure.Compound (I)1 symmetry to generate helical chains along the b-axis direction. The O—H⋯S hydrogen bonding serves to connect translationally related chains along the a-axis direction and these contacts are reinforced by phenyl-C—H⋯O(carbonyl) interactions. In this way, a supramolecular layer in the ab plane is formed, Fig. 3a). Layers stack along the c-axis direction without directional interactions between them, Fig. 3b).In the crystal of (I)Crystal Explorer 17  for (I)dnorm in Fig. 5O atoms, Table 3The Hirshfeld surface mapped over electrostatic potential in Fig. 4ER) descriptor, which is derived from the analysis of the Hirshfeld surface , for a pair of elements  is defined as the ratio between proportion of actual contacts in the crystal to the theoretical proportion of random contacts. This ratio is greater than unity for a pair of elements having a high likelihood to form contacts in a crystal, while it is less than one for a pair which tends to avoid contacts with each other. A listing of ER values for (I)The intermolecular contacts in the crystal of (I)a)–(e), respectively. A summary of the percentage contributions from the various contacts in the crystal are given in Table 5b) and has an ER value of 0.92, i.e. close to unity. The contribution from O⋯H/H⋯O contacts is viewed as long spikes at de + di ∼1.8 Å, with points scattered around different regions in the delineated fingerprint plot, Fig. 6c). In the fingerprint delineated into C⋯H/H⋯C contacts in Fig. 6d), a pair of small tips at de + di < 2.8 Å is the result of short interatomic contacts, Table 3A⋯O1). The percentage contribution from S⋯H/H⋯S contacts (13.1%) reflect the presence of O—H⋯S hydrogen bonds and are apparent through the appearance of asymmetric spikes at de + di ∼2.1 Å in Fig. 6e).The overall fingerprint plots for (I)EBSSEint) listed in Table 6−1 more stable than the O—H⋯S interaction despite phenyl-C—H being a weak hydrogen-bond donor and thione-S a weak acceptor, and that such interactions are known to be dispersive in nature  of all pairwise interactions sums to −36.11 kcal mol−1, while the total dispersion energy term (Edispersion) computes to −43.83 kcal mol−1.As the molecular packing is governed directionally by hydrogen bonding between molecules, the energy frameworks were simulated (Turner 2CH2OH)2, (II), has been reported twice i.e. of the type phenyl-C—H⋯π(phenyl), to sustain a three-dimensional architecture. The other closely related structure is that of 4-MePhC(=O)N(H)C(=S)N(Me)CH2CH2OH) N(H)C(=S)N. The optical absorption spectra were measured on 10 and 100 µM ethanol:acetonitrile (1:1) solutions in the range 190–1100 nm on a double-beam Shimadzu UV 3600 Plus UV–vis spectrophotometer. The thermogravimetric analysis (TGA) was performed on a Perkin Elmer STA 6000 Simultaneous Thermogravimetric Analyzer in the range of 35–900°C under a nitrogen atmosphere at a flow rate of 10°C min−1. The experimental powder X-ray diffraction pattern was measured on a Rigaku MiniFlex diffractometer with Cu Kα1 radiation (λ = 1.54056 Å) in the 2θ range of 5–70° and a step size of 0.02°. The experimental PXRD patterns were compared to the simulated PXRD patterns calculated from the CIF using the Rigaku PDXL structure analysis software package. The patterns matched indicating that the reported crystal structure is representative of the bulk material.All chemicals and solvents were used as purchased without purification. The reactions were carried out under ambient conditions. The melting point was measured using a Hanon MP-450 melting point apparatus. The CHN elemental analysis was performed on a LECO TruSpec Micro analyser under helium atmosphere with glycine being used as the standard. The IR spectrum was measured on a Bruker Vertex 70v FT–IR spectrophotometer from 4000 to 400 cm13H18N2O3S, found : C 55.59 (55.30), H 6.57 (6.43), N 9.79 (9.92). IR : 3312 , 3158 , 3061 , 2955–2881 , 1686 , 1539 , 1250 , 1054 , 747 . UV : λmax nm  354.4 , 294.0 , 246.4 , 202.6 . 1H NMR , 7.76 , 7.31 , 5.66 , 4.87 , 3.98 , 3.76 , 3.70 , 2.37 . 13C{1H} NMR : δ 180.63 (C1), 163.78 (C6), 141.88 (C7), 130.13 (C10), 128.47 , 127.28 , 58.58 (C5), 56.95 (C3), 54.42 (C4), 54.29 (C2), 20.42 (C13).Synthesis of (I)2O between 135 and 165°C, which corresponds to approximate 6% of the weight for (I)The pyrolytic processes for (I)Uiso(H) set to 1.2–1.5Ueq(C). The O- and N-bound H atoms were located from a difference map and refined with O—H and N—H = 0.84±0.01 and 0.88±0.01 Å, respectively, and with Uiso(H) = 1.5Ueq(O) and 1.2Ueq(N).Crystal data, data collection and structure refinement details are summarized in Table 710.1107/S2056989019012581/hb7854sup1.cifCrystal structure: contains datablock(s) I, global. DOI: 10.1107/S2056989019012581/hb7854Isup2.hklStructure factors: contains datablock(s) I. DOI: 1919878CCDC reference:  crystallographic information; 3D view; checkCIF reportAdditional supporting information:


The second, C8H11N9O2S, contains two azide groups with an average Nα—Nβ distance of 1.249 (2) Å and an average Nβ—Nγ distance of 1.132 (2) Å. Each compound contains a bulky protecting group.The structures of two azide containing imidazole derivatives are reported. The first, C 8H12N6O2S, contains one azide group with an Nα—Nβ distance of 1.229 (2) Å and an Nβ—Nγ distance of 1.128 (2) Å. The second, C8H11N9O2S, contains two azide groups with an average Nα—Nβ distance of 1.249 (2) Å and an average Nβ—Nγ distance of 1.132 (2) Å. Each compound contains a bulky protecting group (dimethylaminosulfonyl) which can be easily removed under mildly acidic conditions.The structures of two azide containing imidazole derivatives are reported. Allylic azides are fairly reactive making them attractive starting compounds to convert into amides. The first, C These compounds were synthesized in the previous study but the structures were not reported. Figs. 11 and 2, respectively.The efficient synthesis of nagelamide alkaloids  has garnered interest  Å and an N4—N5 distance of 1.128 (2) Å. The N3—N4—N5 angle is 172.32 (13)°. The azide groups in 2 show an N3—N4 distance of 1.253 (2) Å, N4—N5 distance of 1.129 (2) Å, N6—N7 distance of 1.239 (2) Å, and N7—N8 distance of 1.134 (2) Å. The N3—N4—N5 angle is 171.58 (15)° and the N6—N7—N8 angle is 173.95 (15)°. All three azide moieties in both compounds show the same general trend of a longer Nα—Nβ distance and shorter Nβ—Nγ distance with a quasilinear geometry. This is typical for covalent azides with terminal Nβ—Nγ demonstrating more triple-bond character. A previously reported covalent azide occurring in the compound ethyl-2-[(azidocarbonyl)amino]benzoate demonstrated bond lengths Nα—Nβ of 1.264 (2) Å and Nβ—Nγ of 1.131 (2) Å and an Nα—Nβ—Nα angle of 174.7 (2)°  in 1 was −115.21 (13)° while the related torsion angle (C5—C6—N6—N7) in 2 was 50.25 (18)°. 2 contains one azide group bound to the imidazole at C2 and shows a torsion angle N1—C2—N3—N4 of −174.82 (11)°. The allylic azides in both compounds exhibit a similar dihedral angle between the azide and the imidazole ring, 70.3 (11)° for 1 and 77.3 (17)° for 2. While the imidazole-bound azide in 2 shows a dihedral angle of 5.0 (10)°. Indeed, the torsion angle and dihedral angle for this particular azide demonstrate the near planarity between the imidazole and its covalently bound azide. Figs. 31 and 2, respectively.The torsion angles for the azides and dihedral angles between the azides and imidazole rings for both compounds have been measured. The allylic azide torsion angles between sp3-hybridized, as validated by the C—N—C bond angles C6—N6—C8 = 113.86 (10)° for 1 and C7—N9—C8 = 113.93 (12)° for 2. Both compounds also contain a double bond between C4 and C5. The measured bond distance is 1.333 (2) Å for 1 and 1.340 (2) Å for 2.Both title compounds contain a DMAS protecting group. The amine component of this protecting group is 1 is substituted at the N1 and C3 position with no substitution at C2. The N1—C2 distance is 1.378 (2) Å while the N2—C2 distance is 1.301 (2) Å. However, in 2, the imidazole ring is substituted with an azide group at C2 but this seemingly has no effect on the ring bond distances. The measured bond distances for N1—C2 and N2—C2 in 2 are 1.385 (2) and 1.310 (2) Å, respectively.The imidazole ring in 1 is 1.686 (1) Å and 1.718 (1) Å for 2. The disparity may be attributed to the presence of azide, which is substituted at the C2 position for 2.There is, however, a significant difference in the measured N1—S1 distance for the two compounds. The imidazole ring is substituted at the N1 position for both compounds with DMAS. The N1—S1 distance for 1 shows C1—H1⋯O1i and C2—H2⋯O2ii interactions of 2.53 and 2.39 Å, respectively. There is also a C4—H4⋯N5iii interaction of 2.70 Å acrylate, was reported  Å which is similar to the bond distances in 1 and 2 [1.333 (2) and 1.340 (2) Å respectively].A search of related compounds was conducted in the Cambridge Structural Database  = 1.5Ueq(C) for methyl H and 1.2Ueq(C) for other H atoms.Crystal data, data collection and structure refinement details for 10.1107/S205698901900519X/zl2753sup1.cifCrystal structure: contains datablock(s) compound_1, compound_2. DOI: 10.1107/S205698901900519X/zl2753compound_1sup2.hklStructure factors: contains datablock(s) compound_1. DOI: 10.1107/S205698901900519X/zl2753compound_2sup3.hklStructure factors: contains datablock(s) compound_2. DOI: Click here for additional data file.10.1107/S205698901900519X/zl2753compound_1sup4.cmlSupporting information file. DOI: Click here for additional data file.10.1107/S205698901900519X/zl2753compound_2sup5.cmlSupporting information file. DOI: 1910313, 1910312CCDC references:  crystallographic information; 3D view; checkCIF reportAdditional supporting information: