RDPBR’ Pd0 and PdII complexes (RDPBR’=(o‐PR2C6H4)2BR’, diphosphinoborane) was analyzed using XRD, 11B NMR spectroscopy and NBO/NLMO calculations. The borane acceptor discriminates between the oxidation state PdII and Pd0, stabilizing the latter. Reaction of lithium amides with [(RDPBR’)PdII(4‐NO2C6H4)I] chemoselectively yields the C−N coupling product. DFT modelling indicates no significant impact of PdII→B coordination on the inner‐sphere reductive elimination rate.The dative Pd→B interaction in a series of  Ligand effects: The borane acceptor in diphosphinoborane ligands discriminates between the oxidation state PdII and Pd0, stabilizing the latter. The impact of the Pd→B interaction on inner‐sphere C−N bond reductive elimination of N,N‐dimethyl‐4‐nitroaniline was investigated (see scheme). We determine how the oxidation state of Pd and co‐ligands affect the strength of the Pd→B interaction in DPB complexes. NBO/NLMO calculations and solid‐state structures are used to assess the strength of Pd→B interactions. The value of the 11B NMR chemical shift as a probe is discussed. The reductive elimination of N,N‐dimethyl‐4‐nitroaniline from [(PhDPBPh)PdII(4‐NO2‐C6H4)NMe2] (5) was studied and modelled with DFT calculations to investigate the assumed influence of the borane acceptor.We speculated that the electron‐withdrawing properties of the borane functionality in diphosphinoborane (DPB) ligands enhances the rate of inner‐sphere reductive elimination from Pd complexes due to 1) overall reduced electron density at the PdPhDPBPh)PdII] complexes was synthesized to examine a possible correlation between the nature of ligands at Pd and the strength of the PdII→B interaction (Scheme A series of [(n Scheme .PhDPBPh)PdIICl2] (7) was produced by reaction of PhDPBPh ligand with [(cod)PdCl2] in DCM and was isolated in 74 % yield  Å is shorter than the sum of the van der Waals radii (3.28 Å),2‐ type coordination to the PdII center. A slightly increased pyramidalization at the boron atom is observed (ΣBα=355.4°) compared to complex [(iPrDPBPh)PdCl2] (ΣBα=359.9°).Complex  and was isolated in 67 % yield. The 31P NMR spectrum displays two broad resonances of equal intensity at δ=45.2 and 38.1 ppm (CD2Cl2), suggesting a similar dynamic process as in 7. Due to the poor solubility of both 7 and 8, no 11B NMR spectra could be obtained.Complex PhDPBPh)PdIICl]SbF6 (9) was produced in 51 % isolated yield by halide abstraction from 7 with AgSbF6 PdII(μ‐Cl)]2(SbF6)2 is observed with an inversion center between the two PdII centers. Within the dimer, the PdII center is coordinated in a square‐pyramidal fashion with the borane located in the apical position. The Pd, B distance in complex 9 is 2.721(5) Å, which is slightly shorter than in [(PhDPBPh)PdIICl2] 7 (2.762(3) Å). However, pyramidalization of the borane is almost identical (ΣBα=355.8°). The absence of a relevant η2→PdII interaction is suggested by the long Pd1,C1 distance of 3.338(4) Å. The Pd,B distance and lack of significant pyramidalization at the borane suggest a weak PdII→B interaction, which is in line with a broad resonance in the 11B NMR spectrum at δ=65 ppm (ω1/2=1900±500 Hz).Cationic complex SbF6 (10) was synthesized by reaction of AgSbF6 with zwitterionic allyl complex [{(o‐PPh2C6H4)2B(OAc)Ph}PdII(C3H5)] (4)  Å, which is in line with a minor pyramidalization at the borane center (ΣBα=354.7°) and a broad 11B NMR resonance at δ=62 ppm (ω1/2=1200±100 Hz). A large Pd,C22 distance of 3.066(6) Å eliminates the possibility of a strong η2→PdII interaction. The η3‐coordinated C3H5‐ligand is disordered. Using the borane as a reference point, a 39:61 mixture of the exo‐ and endo‐isomers is observed. A wider P‐Pd‐P angle of 102.86(5)° is realized by a decrease in the twisting of the ligand backbone . The observed disorder of the C3H5‐ligand is in good agreement with the observed NMR spectra. In the 31P NMR spectrum (CD2Cl2), two singlet resonances are observed in a 40:60 ratio (δ=28.1 and 26.9 ppm) and two sets of C3H5‐units are detected in the 1H NMR spectrum. DFT calculations (BP86/def‐SV(P)) based on the solid‐state structures of endo10‐ and exo10‐ indicate a small Gibbs free energy preference of ΔG=0.74 kcal mol−1 for endo10‐, predicting a 29:71 ratio at 298 K.Cationic allyl complex [(H5)]SbF6 0 was syno‐PPh2C6H2B(OAc)PhII→B interaction on reductive elimination proceeding via an inner‐sphere mechanism, complex [(PhDPBPh)PdII(4‐NO2‐C6H4)I] (5) was reacted with lithium amides. Complex 5 was reacted with LiNMe2 (1.1 equiv) at room temperature in [D8]THF  and 38.3 ppm (14 %). After a total of 4.5 h, all resonances in the 31P NMR spectrum disappeared in favor of the singlet at δ=31.1 ppm. 11B NMR spectroscopy suggested formation of a zero‐valent palladium complex by a broad resonance at δ=19 ppm (ω1/2=400±100 Hz). The concurrent formation of the expected reductive elimination product N,N‐dimethyl‐4‐nitroaniline was confirmed by GC/MS analysis, using an independently prepared sample as a reference. The absence of an intermediate complex cis‐[(PhDPBPh)PdII(4‐NO2‐C6H4)NMe2] suggests that transmetalation is rate‐limiting in this transformation. The intermediate occurrence of the 31P NMR resonance at δ=38.3 ppm is possibly due to a reversible reaction of LiNMe2 with complex 6. In a control experiment complex [(PhDPBPh)Pd0(pyridine)] (1) was reacted with LiNCy2 and LiNMe2 in [D8]THF. In both cases ca. 7 % of a new complex at δ=38.5 (s) and 37.7 ppm (s) were observed.A conversion of 84 % was observed 6 decomposed within hours with simultaneous precipitation of palladium black. Addition of PMe3 as a stabilizing co‐ligand led to the formation of complex [(PhDPBPh)Pd0(PMe3)] 11. The 31P NMR spectrum of 11 showed a doublet at δ=35.3 and a triplet at −40.1 ppm (J=15.1 Hz) in a 2:1 ratio, which is consistent with the expected κ3P‐coordination. The broad resonance in the 11B NMR spectrum at δ=25 ppm (ω1/2=400±100 Hz) suggested a strong Pd0→B interaction. Complex 11 could also be synthesized independently by reaction of PBP pincer 12 with PhLi and PMe3, or reaction of 1 with PMe3, thus confirming unambiguously the identity of 11 (Scheme Complex 5 reacted in a similar fashion with LiNCy2 (26 % 6 after 3 h) and LiNHtBu (14 % 6 after 5.5 h). However, the reaction proceeded slower with these sterically more demanding substrates. The reaction of complex 5 with LiNHtBu was monitored for 96 h by 31P NMR spectroscopy (46 % conversion towards 6) without any side products being observed (cf. Table S1). This is in line with the assumption of a rate‐determining transmetalation followed by a quick reductive elimination.Complex 0/II DPB complexes were analyzed to identify factors which affect the strength of Pd→B interactions. In addition to the new Pd complexes presented in this work (6–10), the structurally characterized DPB complexes cis‐[(PhDPBPh)PdII(4‐NO2‐C6H4)I] (5),PhDPBPh)Pd0(pyridine)] (1),PhDPBMe)Pd0(PMe3)] (13)CyDPBPh)Pd0] (3)0/II center. The shorter Pd,B distances and higher degree of borane pyramidalization =2.194(3) −2.243(2) Å vs. PdII: ΣBα=354–356°, d=2.676(5) −2.762(2) Å). Remarkably, even the generation of cationic PdII complexes (9 and 10) has no significant impact on the strength of PdII→B interactions. The oxidation state at Pd is unambiguously the dominant factor for the strength of the Pd,B bond.The solid‐state structures of PdPBPh)PdII‐NO2‐C6H4s work 6–0, the st1, 3, 5–11 and 13 were geometrically optimized using Turbomole 7.0.1 (BP86/def‐SV(P)). A good agreement was observed between the optimized structures and their corresponding solid‐state structures ‐coordination was found in the NBO calculations. The Wiberg bond index for Pd,Cipso was below 0.02, with the exception of Pd0 complexes 1 (0.0697) and 6 (0.0325). Reactivity studies of [(DPB)Pd]‐complexes presented in this paper thus appear to be unaffected from significant η2‐coordination.The Pd→B interactions were further analyzed using QM calculations. Complexes II→B interactions, a narrow range of NBO stabilizing energies between 8.04 and 11.46 kcal mol−1 was observed. Surprisingly, generation of cationic complexes , exchange of chloro‐ligands by bromide (8) or iodide/aryl (5) had very little effect. In the case of Pd0→B interactions, significantly higher NBO stabilizing energies of 19.53–46.83 kcal mol−1 were found. Regardless of the oxidation state at Pd an approximately linear correlation between the Pd,B distance and the NBO stabilizing energy (E2) associated with the Pd,B interaction was observed  by PCy2‐groups (3) had only a minor effect. The E2 values for the Pd0→B interaction in the 14 VE complexes 3  and 6  significantly deviate from this correlation and are almost twice as much as for 16 VE complexes 1  and 13 . Neither the 11B NMR chemical shift, Pd,B distance or pyramidalization at B indicate a change of the Pd0→B interaction strength in this magnitude between the 14 VE and the 16 VE complexes Pd0(PPh3)] (2) reported by Kameo and Bourissouδ(11B) 27 ppm). In contrast, the 11B NMR resonance shifts linearly towards lower field with an increasing Pd,B distance in case of PdII complexes. 11B NMR spectroscopy therefore can be used as a tool to assess the strength of Pd→B interactions within a given ligand system, provided that the oxidation state at the Pd center is taken into account. However, given the difficulty to determine the precise δ(11B) of [(DPB)PdII] complexes (poor solubility and ω1/2 >1000 Hz ), a certain error for weak PdII→B interactions needs to be factored in.The r Figure . ComplexN,N‐dimethyl‐4‐nitroaniline from complex 14‐B , yielding Pd0 complex 6 and N,N‐dimethyl‐4‐nitroaniline . In order to understand how the PdII→B interaction affects the reductive elimination, the reaction was also modeled for bis[(2‐diphenylphosphino)phenyl]ether (DPEphos) complex 14‐O and diphosphinoamine complex 14‐N. DPEphos is well established as an effective ligand in palladium catalyzed Buchwald–Hartwig‐type coupling reactions,PhDPBPh (Table o‐PPh2C6H4)2NPhN‐Ph bridgehead gives a good model of the B‐Ph group in 14‐B. Elimination of N,N‐dimethyl‐4‐nitroaniline from complexes 14‐O and 14‐N gave very similar Gibbs free reaction energies of ΔG=−38.52 kcal mol−1 and ΔG=−38.63 kcal mol−1, respectively. No Pd0/II→E interactions were observed in complexes featuring DPEphos and the diphosphinoamine ligand  an electronic effect by Pd→B coordination and 2) increased steric bulk of the DPB ligand imposed by the B‐Ph group. In case of diphosphinoamine complex 14‐N the reductive elimination barrier decreased to ΔG≠=5.54 kcal mol−1 , possibly as a result of the increased steric pressure imposed by the N‐Ph group  proceeds via structurally early transition‐state 15‐E  were used to model the inner‐sphere reductive elimination of 15‐B, compared to starting complex 14‐B, as indicated by a slightly elongated Pd,B distance (2.947 Å) in 15‐B compared to 14‐B (2.906 Å). Similarly, the Wiberg bond index for the Pd→B interaction is reduced to 0.162 in 15‐B (14‐B: 0.176), and the NPA charge at the borane remains unchanged (14‐B: +0.737 vs. 15‐B: +0.735). The increase of the Pd→B interaction strength occurs after the reductive elimination, explaining why the inner‐sphere reductive elimination of the C−N bond does not kinetically profit from the substantial increase of the Pd→B strength in the course of the reaction.Unexpectedly, the Pd→B interaction is slightly weakened in transition‐state N,N‐dimethyl‐4‐nitroaniline was also modeled using cis‐[(PMe3)2PdII(4‐NO2C6H4)NMe2]  and its BH3 adduct [(PMe3)2(BH3)PdII(4‐NO2C6H4)NMe2]  as substrates (cf. Scheme S1). Again, a more favorable transition state was found for the acceptor free complex 17 , than for the borane adduct 17‐B .To rule out effects originating from restraints imposed by a chelating ligand frame work, the reductive elimination of 11B NMR spectroscopy has been established as a useful tool to assess the strength of Pd→B interactions in solution. Reaction of lithium amides with [(PhDPBPh)PdII(4‐NO2C6H4)I] (5) chemoselectively yields the C‐N coupling product and [(PhDPBPh)Pd0] (6). Inner‐sphere reductive C−N bond elimination was modelled with DFT methods for the PhDPBPh ligand. In contrast to reports on acceptor promoted outer‐sphere reductive C−N bond elimination,The strength of Pd→B interactions in [(DPB)Pd] complexes depends primarily on the oxidation state of Pd. In contrast, modifications of the DPB ligand or co‐ligands have only a minor effect. n‐hexane were dried over sodium, distilled under argon prior to use and stored over activated molecular sieves (4 Å).All manipulations were performed under an argon atmosphere using standard Schlenk line and glovebox techniques. Glassware was oven dried at 120 °C overnight and dried with a heat gun under vacuum prior to use. Tetrahydrofuran was dried by an MBraun solvent purification system. Benzene and 2Cl2 and C6D6 were degassed employing the freeze‐pump‐thaw technique and stored over activated molecular sieves (4 Å). [D8]THF was dried over activated molecular sieves (3 Å), distilled under an argon atmosphere and degassed employing the freeze‐pump‐thaw technique. PhDPBPh, [(PhDPBPhOAc)Pd(C3H5)] (4), [(PhDPBPh)Pd(4‐NO2C6H4)I] (5) and [{(o‐PPh2C6H4)2BPh}PdI] (12) were synthesized according to published procedures.CD® quick pressure valve NMR tubes. 1H, 11B{1H}, 13C{1H}, 19F{1H}, and 31P{1H} NMR spectra were recorded on a Bruker Avance II  or a Bruker Avance  spectrometer. 1H and 13C{1H} NMR spectra were referenced to residual solvent resonances as implemented in MesReNova 10.0.2. Infrared spectra were recorded on an Avatar 360 FT‐IR E.S.P. device by Nicolet. CHN combustion analysis were carried out on an Elementar EL device by Elementar Analysesysteme GmbH.NMR‐experiments were performed in Wilmad7), 1987625 (9) and 1987626 (10) Deposition Number(s) 1987620 ( contain(s) the supplementary crystallographic data for this paper. These data are provided free of charge by the joint Cambridge Crystallographic Data Centre and Fachinformationszentrum Karlsruhe Access Structures service www.ccdc.cam.ac.uk/structures.8]THF (0.25 mL) was added dropwise over a period of 4 min to a stirred solution of nitroarene complex 5  in [D8]THF (0.25 mL). The resulting mixture was stirred for another 5 min and then transferred into an NMR tube. Reductive elimination was monitored by 31P NMR spectroscopy.A solution of the respective lithium amide  in [D2Cl2 (8 mL) was added to a mixture of PhDPBPh  and [(cod)PdCl2] . The mixture was stirred for 30 min at room temperature. Yellow crystals  were formed by overlaying the solution n‐pentane (16 mL). Single crystals suitable for X‐ray diffraction were grown from a solution of [(cod)PdCl2]  and PhDPBPh  in CD2Cl2 (0.7 mL) overlaid with benzene (0.3 mL). 11B and 13C NMR data have not been collected due to poor solubility. 1H NMR : δ 7.81–7.76 , 7.55 , 7.50–7.46 , 7.46–7.38 , 7.35–7.14 , 6.97–6.78 , 5.32 . 31P{1H} NMR : δ 44.5 . IR (KBr): ν˜=3643‐3284 (w), 3049 (w), 1587 (w), 1497 (m), 1433 , 1223 (s), 1158 (vw), 1128 (w), 1093 (vs.), 987 (w), 889 (vw), 864 (vw), 754 (s), 744 (s), 733 (m), 688 (vs.), 667 (w), 611 (m), 600 (s), 542 (m), 523 (vs.), 505 (m) cm−1. Elemental analysis calcd (%) for C42H33BCl2P2Pd⋅CH2Cl2: C 59.18, H 4.04, found: C 59.61, H 4.33.CHPhDPBPh ligand  and [(cod)PdBr2]  were solved in DCM (10 mL) and stirred at r.t. for 30 min. The solution was overlaid with n‐hexane (20 mL) yielding title compound 8 as orange crystals . 11B and 13C NMR data have not been collected due to poor solubility. 1H NMR : δ 7.85–7.76 , 7.59–7.19 . 31P{1H} NMR : δ 45.2 , 38.1 . IR (KBr): ν˜=3424 (s), 3048 (m), 1621 (w), 1587 (w), 1478 (m), 1455 (w), 1432 (s), 1311 (w), 1237 (w), 1220 (s), 1205 (m), 1187 (m), 1153 (w), 1126 (m), 1092 (s), 1027 (w), 1000 (m), 887 (w), 863 (w), 753 (s), 741 (s), 713 (m), 699 (s), 690 (s), 667 (m), 610 (s), 600 (s), 539 (s), 522 (s), 505 (s), 465 (m) cm−1. Elemental analysis calcd (%) for C42H33BBr2P2Pd⋅0.25CH2Cl2: C 56.51; H 3.76, found: C 56.72, H 3.83.The 7  and AgSbF6  were stirred in DCM (15 mL) for 40 minutes. The suspension was filtered through a syringe filter . The clear solution was overlaid with n‐hexane (30 mL) yielding the title compound 9 as long colorless needles . 1H NMR : δ 7.97–7.92 , 7.80 , 7.69 , 7.65 , 7.55 , 7.47–7.34 , 7.27–7.16 , 7.00 , 6.83 . 11B{1H} NMR : δ=65 . 13C{1H} NMR : δ=δ 141.79, 135.43 , 134.88 , 134.25, 133.69 , 133.22 , 132.49 , 129.67 , 129.33–128.82 (m), 128.10, 127.13, 126.74, 126.16. 31P{1H} NMR : δ 49.9 . IR (KBr): ν˜=3441 (s), 3058 (w), 1588 (w), 1482 (w), 1435 (s), 1230 (m), 1200 (w), 1125 (w), 1034 (m), 1001 (w), 867 (vw), 752 (s), 702 (s), 692 (s), 659 (vs.), 614 (m), 538 (s), 517 (s), 697 (w) cm−1. Elemental analysis calcd (%) for C42H33BClF6P2PdSb⋅0.25 C6H14: C 51.75, H 3.64, found: C 51.77, H 3.785.Complex 4  and AgSbF6  were solved in CH2Cl2 (7 mL) and stirred at r.t. for 20 min. The suspension was filtered through a syringe filter . The clear solution was overlaid with n‐hexane (10 mL). The obtained crystals showed insufficient purity and were crystallized again under the same conditions yielding 10 as slightly yellow crystals . 1H NMR : δ 7.72–7.59 , 7.58–7.53 , 7.53–7.44 , 7.43–7.29 , 7.23–7.15 , 7.05–6.87 , 6.78–6.67 , 5.88–5.70 , 3.77–3.61 , 3.59–3.33 , 3.03–2.85 , 2.49–2.29  . 11B{1H} NMR : δ 64 . 13C{1H} NMR : δ 141.1, 140.2, 136.1, 135.5, 135.3, 135.0, 134.4, 134.3, 134.0, 133.2 , 132.3, 132.2, 132.1, 131.6, 131.5, 131.2, 131.0, 129.6 , 129.3, 128.9, 123.1, 80.4, 80.2. 31P{1H} NMR : δ 28.1 , 26.9 . IR (KBr): ν˜=3430 (s), 3000 (m), 1588 (m), 1480 (m), 1458 (w), 1434 (s), 1268 (m), 1227 (s), 1127 (m), 1095 (m), 1031 (w), 999 (w), 950 (vw), 875 (w), 772 (w), 754 (m), 742 (m), 733 (m), 695 (s), 659 (vs.), 609 (s), 537 (m), 521 (s), 478 (w), 430 (w) cm−1. Elemental analysis calcd (%) for C46H40BCl2F6P2PdSb: C 51.22, H 3.74, found: C 51.04, H, 3.86.Allyl complex 2⋅THF  in [D8]THF (0.25 mL) was added over a period of 3 min to a solution of complex 5  in [D8]THF (0.25 mL). The combined solutions were transferred to an NMR tube and NMR spectra were recorded after 1.5 and 4.5 h. 11B{1H} NMR : δ 19 . 31P{1H} NMR : δ 30.93 (s).A solution of LiNMe12  in THF (0.5 mL). After stirring for 10 min at r.t. a solution of PMe3 in toluene  was added. The precipitate was removed by filtration and the solution was concentrated in vacuo. The resulting solid was washed with pentane and dried in vacuo . 1H NMR : δ 8.34 , 7.69–7.58 , 7.44–7.37 , 7.36–7.28 , 7.12 , 7.09–7.05 , 6.85 , 6.68 , 0.64 . 11B{1H} NMR : δ 25 . 13C{1H} NMR : δ 168.7 (bs), 143.2 , 143.0 , 141.5 , 138.9 , 135.8 , 135.7 , 133.5 , 133.0 , 132.3 (s), 132.3 (s), 132.4 , 129.5 (s), 129.0 (s), 128.6 (s), 127.2 (s), 126.1 , 125.2 (s), 18.1 . 31P{1H} NMR : δ 35.44 , −40.13 .A solution of PhLi  in THF (0.5 mL) was slowly added to a solution of complex The 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 title hydrazine carbodithioate chloroform hemi-solvate features approximately planar molecules with splayed phenyl groups. Supramolecular tapes are formed in the crystal through a combination of N—H⋯S, hydroxyl-O—H⋯O(hydroxyl) and hydroxyl-O—H⋯π(phenyl) interactions. 15H14N2O2S2·CHCl3, comprises two independent hydrazine carbodithioate molecules, A and B, and a chloroform molecule; the latter is statistically disordered about its molecular threefold axis. The common features of the organic molecules include an almost planar, central CN2S2 chromophore [r.m.s. deviation = 0.0203 Å (A) and 0.0080 Å (B)], an E configuration about the imine bond and an intramolecular hydroxyl-O—H⋯N(imine) hydrogen bond. The major conformational difference between the molecules is seen in the relative dispositions of the phenyl rings as indicated by the values of the dihedral angles between the central plane and phenyl ring of 71.21 (6)° (A) and 54.73 (7)° (B). Finally, a difference is seen in the disposition of the outer hydroxyl-H atoms, having opposite relative orientations. In the calculated gas-phase structure, the entire molecule is planar with the exception of the perpendicular phenyl ring. In the molecular packing, the A and B molecules assemble into a two-molecule aggregate via N—H⋯S hydrogen bonds and eight-membered {⋯HNCS}2 synthons. The dimeric assemblies are connected into supramolecular chains via hydroxyl-O—H⋯O(hydroxyl) hydrogen bonds and these are linked into a double-chain through hydroxy-O—H⋯π(phenyl) interactions. The double-chains are connected into a three-dimensional architecture through phenyl-C—H⋯O(hydroxyl) and phenyl-C—H⋯π(phenyl) interactions. The overall assembly defines columns along the a-axis direction in which reside the chloroform molecules, which are stabilized by chloroform–methine-C—H⋯S(thione) and phenyl-C—H⋯Cl contacts. The analysis of the calculated Hirshfeld surfaces, non-covalent interaction plots and interaction energies confirm the importance of the above-mentioned interactions, but also of cooperative, non-standard interactions such as π(benzene)⋯π(hydrogen-bond-mediated-ring) contacts.The title hydrazine carbodithioate chloroform hemisolvate, 2C Dithiocarbazato Schiff bases have received considerable attention because of the presence of both soft sulfur and hard nitrogen atoms As part of on-going studies in this area  Å, for the N2 atom, and 0.0319 (16) Å for N1; the C2 and C9 atoms lie, respectively, 0.161 (3) and 0.096 (4) Å out of the plane, in the direction of the N2 atom. The comparable plane for the S3-molecule is significantly more planar with an r.m.s. deviation = 0.0080 Å with maximum deviations of 0.0131 (16) Å for the N3 atom and 0.0104 (12) Å for atom N4; the C17 atom lies 0.018 (3) Å out of the central plane in the direction of the N3 atom, and the C24 lies 0.123 (3) Å out of the plane in the direction of the N4 atom. The small difference in planarity is reflected in the C1—N1—N2—C2 and C16—N3—N4—C17 torsion angles of 171.8 (2) and 179.3 (2)°, respectively. More significant conformational differences are apparent in rest of the molecules: for the S1-molecule, the dihedral angles between the central residue and terminal hydroxybenzene and phenyl rings are 6.18 (13) and 77.21 (6)°, respectively, indicating close to co-planar and perpendicular relationships; the dihedral angle between the terminal rings is 71.22 (8)°. The equivalent dihedral angles for the S3-molecule are 6.07 (13), 54.53 (6) and 54.73 (7)°, respectively. The other notable difference between the molecules relates to the relative orientation of the hydroxy-H atoms in the 4-position, no doubt arising owing to the dictates of the molecular packing.The crystallographic asymmetric unit of (I)E in each case. The comparison of geometric parameters in Table 1The relatively co-planar relationship between the central residue and the appended hydroxybenzene ring allows for the formation of an intramolecular hydroxy-O—H⋯N(imine) hydrogen bond in each molecule, Table 2Gaussian16 An overlay diagram for the experimental and theoretical, gas-phase structures is shown in Fig. 2via O4-hydroxy-O—H⋯π(phenyl) interactions as illustrated in Fig. 3a). The assembly lies parallel to  more than −0.18 a.u., Fig. 6b) and (c). The intramolecular O—H⋯N contacts reveal similar attractive interactions.Among all close contacts present in (I)NCIPLOT results, the strength of interaction for each close contact was quantified by calculation of the interaction energy in Gaussian16 , the combination of π(C3–C8)–quasi-π, quasi-π–quasi-π, C24—H24A⋯S2, C14—H14⋯ π(C25–C30) and C15—H15⋯π(C25–C30) between S1- and S3-molecules exhibits the greatest interaction energy among all close contacts with an E of −65.73 kJ mol−1, Table 52 synthon, being the second strongest interaction with E = −59.79 kJ mol−1. The strength of the N1—H1N⋯S3/N3–H3N⋯S1 interaction is consistent with the energy range of −54.06 to −57.99 kJ mol−1 displayed by the equivalent contacts in the cinnamaldehyde Schiff base of S-(4-methylbenzyl) dithiocarbazates calculated through wB97XD/6-31G C=NN(H)C(=S)SR, five of which have the hydroxyl substituent in the 2-position enabling the formation of an intramolecular hydroxy-O—H⋯N(imine) hydrogen bond. In the most closely related compounds, i.e. with 2-OH substituents, the R group in the ester substituent is methyl  and 2,4-dihydroxybenzaldehyde  were mixed and heated until the initial volume was reduced by half. The yellow precipitate formed after cooling the mixture to room temperature was collected and washed with cold ethanol. It was recrystallized from ethanol solution and dried over silica gel for three days. Light-yellow prisms were obtained from its 1:1 diethyl ether/chloroform solution by slow evaporation.Two solutions, −1): 3310 , 3094 , 1604 , 1100 , 1024 , 948 . 1H NMR : δ 13.19 , 10.21 , 10.07 , 8.35 , 7.45 , 7.36 , 7.21 , 6.29 , 6.26 , 4.44 . 13C{1H}-NMR : δ ppm. 194.4 (C=S), 162.1, 159.6 (C—OH), 146.5 (N=C), 129.7–127.8 (Ph & benzene), 38.0 (CH2); GCMS (DI): m/z calculated for C15H14N2O2S2+ [M+]: 318, found 318.Yield: 4.98 g, 62%; m.p. 463–465 K. FT–IR UATR (solid), λ (cmUiso(H) set to 1.2Ueq(C). The O- and N-bound H atoms were located in a difference-Fourier map but, were refined with O—H (0.84±0.01 Å) and N—H (0.88±0.01 Å) distance restraints, and with Uiso(H) set to 1.5Ueq(O) and to 1.2Ueq(N), respectively. The CHCl3 solvent molecule is statistically disordered about the molecular threefold axis. The C31 atom is common to both conformations and the individual Cl atoms were refined anisotropically. A loose distance restraint for C—Cl was applied, i.e. C—Cl = 1.76±0.02 Å. The maximum and minimum residual electron density peaks of 1.04 and 1.22 e Å−3, respectively, are located 1.03 and 0.90 Å from the Cl3′ atom.Crystal data, data collection and structure refinement details are summarized in Table 610.1107/S2056989020007070/hb7919sup1.cifCrystal structure: contains datablock(s) . DOI: 10.1107/S2056989020007070/hb7919Isup2.hklStructure factors: contains datablock(s) I. DOI: Click here for additional data file.10.1107/S2056989020007070/hb7919Isup3.cmlSupporting information file. DOI: 2005815CCDC reference:  crystallographic information; 3D view; checkCIF reportAdditional supporting information:


The cation and anion are linked together by supramolecular interactions [graph-set notation of hydrogen bonding a-axis direction]. Additionally, there is π–π stacking involving the salicylate anion and the piperazinium cation in adjacent asymmetric units as well as a C—H⋯π interaction between a hydrogen atom on the piperazine ring and the phenyl ring within the salicyclate anion. In bis[4-(4-nitrophenyl)piperazin-1-ium] bis(4-fluorobenzoate) trihydrate (2C10H14N3O2+·2C7H4FO2−·3H2O), there are two cations, two anions, and three water molecules of solvation in the asymmetric unit, all linked by hydrogen bonds [graph-set notation of hydrogen bonding R22(20) between adjacent cations and R33(9) between a cation and its adjacent anion]. In the anion, the 4-nitrophenyl ring occupies an axial substitution position in the piperazinium ring, which is relatively rare. Within the asymmetric unit, the phenyl groups in the cations show an offset π–π interaction. Additionally, there is a C—H⋯π interaction between a hydrogen atom on the phenyl ring within a cation and the phenyl ring within an anion. In 4-(4-nitrophenyl)piperazin-1-ium 3,5-dinitrobenzoate (C10H14N3O2+·C7H4N2O6−), there is a strong N—H⋯O hydrogen bond linking the cation and anion and the 4-nitrophenyl ring occupies an axial substitution position in the piperazinium ring, as seen in the previous structure. In the crystal, the cation and the anion form a complex three-dimensional hydrogen-bonded array involving R22(8), R44(12) and R44(20) rings propogating in the a-axis direction. The nitrophenyl group is disordered with occupancies of 0.806 (10) and 0.194 (10).The structures and Hirshfeld surface analysis of three salts of 1-(4-nitrophenyl)piperazine are discussed. In 4-(4-nitrophenyl)piperazin-1-ium salicylate (C Additionally, ring B′s piperazine substituent forms a weak C—H⋯O interaction with an oxygen atom in ring C′s terminal nitro group . Ring C′s piperazine substituent forms a similar interaction with ring B′s terminal nitro group . In the conformation of rings A–D, the dihedral angles between the 4-nitrophenyl rings in rings B and C, the 4-nitrophenyl ring and nitro group in ring B, the 4-nitrophenyl ring and nitro group in ring C, the piperazine ring and the 4-nitrophenyl ring in ring B, the piperazine ring and 4-nitrophenyl ring in ring C, the fluorobenzene ring in ring D and the phenyl ring in ring C, and the fluorobenzene ring in A and the phenyl ring in C are 11.4 (4), 1.1 (2), 0.2 (2), 141.72 (16), 145.17 (17), 101.47 (17) and 103.32 (17)°, respectively. The third and fourth angles listed indicate that the 4-nitrophenyl ring occupies an axial position in both cations, which is relatively rare. In a previous paper containing eleven analogous structures, only one had this substitution pattern , crystallizes in the monoclinic space group C2/c with eight formula units in the unit cell. The structure consists of a 4-nitrophenylpiperazinium cation and a 3,5-dinitrobenzoate anion linked by a strong N—H⋯O hydrogen bond . Additionally, there is a C—H⋯π interaction between a hydrogen atom in the piperazine ring and the phenyl ring within the salicyclate anion  x, y, z).In discussing the supramolecular features of the three structures, the direct hydrogen bonding involving the linking of the 4-nitrophenylpiperazinium cations and organic acid anions is omitted since it has already been discussed in the previous section. For 2, there are two anions and two cations as well as three water molecules of solvation in the asymmetric unit. This leads to a complex three-dimensional array of hydrogen bonding involving both B and C, and R33(9) motifs between rings C and D as well as one water molecule, as seen in Fig. 6x, y, −z; see Table 2B forms an offset π–π interaction with the phenyl ring in C . Additionally, there is a C–H⋯π interaction between a hydrogen atom on the phenyl ring in C and the phenyl ring in D .For 3, the cation and the anion form a complex three-dimensional array of hydrogen bonding involving a-axis direction between the 4-nitrophenyl group of one cation with the piperazinium ring of an adjacent cation , two cations and two anions in adjacent asymmetric units , two cations and two anions in adjacent asymmetric units , respectively, as seen in Fig. 9Cg2⋯Cg2i distance, 4.4132 (9) Å; perpendicular distance: 3.5596 (9) Å; slippage of 2.609 Å, symmetry code: (i) x, y, 1 − z; Cg2 is the centroid of the C1–C6D ring].For 4.et al., 1997et al., 2014et al., 2019Related structures containing 1-phenylpiperazine or the 1-phenylpiperazinium cation include racemic perhydrotriphenylene (PHTP), which has been shown to form a polar inclusion compound with 1-(4-nitrophenyl)piperazine (NPP) as a guest molecule -4-(4-nitrophenyl)piperazine (BIQYIM), 1-(4-bromobenzoyl)-4-(4-nitrophenyl)piperazine (BIRHES), 1-(3-bromobenzoyl)-4-(4-nitrophenyl)piperazine (BIRHIW) and bis[(4-fluorophenyl)methanone] (BIRGOB) have been reported  4-nitrophenylpiperazine  in methanol (10 ml) was mixed with equimolar solutions of the appropriate acids in methanol (10 ml) and ethyl acetate (10 ml) viz., salicylic acid (67 mg) for 1, 4-fluorobenzoic acid (68 mg) for 2 and 3,5-dinitrobenzoic acid (102 mg) for 3  for a week. For 3, DMF (3 ml) was used for crystallization. The corresponding melting points were 453–458 K (1), 373–378 K (2) and 445–447 K (3).For the synthesis of salts 6.Uiso(H) = 1.2Ueq(C) while the N–H and water O–H hydrogen atoms were refined isotropically. In 3 the nitrophenyl group is disordered with occupancies of 0.806 (10)/0.194 (10) and constrained to have similar metrical parameters.Crystal data, data collection and structure refinement details for the three structures are summarized in Table 410.1107/S2056989023002517/vm2276sup1.cifCrystal structure: contains datablock(s) 1, 2, 3. DOI: 10.1107/S2056989023002517/vm22761sup2.hklStructure factors: contains datablock(s) 1. DOI: 10.1107/S2056989023002517/vm22762sup3.hklStructure factors: contains datablock(s) 2. DOI: 10.1107/S2056989023002517/vm22763sup4.hklStructure factors: contains datablock(s) 3. DOI: Click here for additional data file.10.1107/S2056989023002517/vm22761sup5.cmlSupporting information file. DOI: Click here for additional data file.10.1107/S2056989023002517/vm22762sup6.cmlSupporting information file. DOI: Click here for additional data file.10.1107/S2056989023002517/vm22763sup7.cmlSupporting information file. DOI: 2248697, 2248696, 2248695CCDC references:  crystallographic information; 3D view; checkCIF reportAdditional supporting information:


Blood Cancer Journal (2023) 13:111 10.1038/s41408-023-00883-x, published online 20 July 2023Correction to: N = 3111)” and “Non-ASCT (N = 2437)” were assigned in incorrect lines. Correct Table In Table ASCT N = 437” wereThe original article has been corrected.


In the published article, there was an error in the name and abbreviation of the measure “mean corpuscular hemoglobin concentration” abbreviated to “MCHC”. It should be “mean corpuscular hemoglobin” and abbreviated to “MCH”. Throughout the text of the article, “mean corpuscular hemoglobin concentration” has been corrected to “mean corpuscular hemoglobin” and the abbreviation “MCHC” has been corrected to “MCH”.In the published article, there was an error in the header, title and legend for The corrected In the published article, there was an error in the legend for In the published article, there was an error. A correction has been made to the section Results, subsection “Analysis of anemia prevalence and hematological phenotype indexes in pregnant women with various thalassemia genotypes”, paragraph 1. This sentence previously stated:SEA, –α3.7/–α3.7, –α3.7/–α4.2, and –α4.2/–α4.2 ranged from 35.96 to 37.82% (P < 0.05).”“The anemia rates of αα/–The corrected sentence appears below:SEA, –α3.7/–α3.7, –α3.7/–α4.2, and –α4.2/–α4.2 ranged from 35.96 to 37.82% (P > 0.05).”“The anemia rates of αα/–A correction has been made to the section Discussion, subsection “Analysis of anemia prevalence and hematological phenotype indexes of pregnant women with various thalassemia genotypes”, paragraph 1. This sentence previously stated:SEA, –α3.7/–α3.7, –α4.2/–α4.2, and –α3.7/–α4.2 ranged from 35.96 to 37.82% (P < 0.05), and no significant differences were observed in Hb levels among those genotypes.”“The anemia rate among carriers of αα/–The corrected sentence appears below:SEA, –α3.7/–α3.7, –α4.2/–α4.2, and –α3.7/–α4.2 ranged from 35.96 to 37.82% (P > 0.05), and no significant differences were observed in Hb levels among those genotypes.”“The anemia rate among carriers of αα/–A correction has been made to the section Discussion, subsection “Prevalence of iron deficiency in anemic pregnant women with various genotypes of thalassemia”, paragraph 1. This sentence previously stated:“Although there is no large sample, multicenter randomized controlled study that has yet confirmed the need for routine iron supplementation in PW with thalassemia minor genotypes, we suggest that the iron load of APW with β- thalassemia minor genotypes should be monitored dynamically during iron supplementation”The corrected sentence appears below:“Although there is no large sample, multicenter randomized controlled study that has yet confirmed the need for routine iron supplementation in PW with thalassemia minor genotypes, we suggest that the ferritin levels of APW with β- thalassemia minor genotypes should be monitored dynamically during iron supplementation”The authors apologize for these errors and state that this does not change the scientific conclusions of the article in any way. The original article has been updated.


Wu et al. show that the mitotic kinesin CENP-E promotes full expansion of the outer layer of the kinetochore known as the fibrous corona. This new function of CENP-E is independent of its motor activity. The findings expand the roles of CENP-E in spindle assembly and chromosome congression in mitosis. Correct chromosome segregation during cell division depends on proper connections between spindle microtubules and kinetochores. During prometaphase, kinetochores are temporarily covered with a dense protein meshwork known as the fibrous corona. Formed by oligomerization of ROD/ZW10/ZWILCH-SPINDLY (RZZ-S) complexes, the fibrous corona promotes spindle assembly, chromosome orientation, and spindle checkpoint signaling. The molecular requirements for formation of the fibrous corona are not fully understood. Here, we show that the fibrous corona depends on the mitotic kinesin CENP-E and that poorly expanded fibrous coronas after CENP-E depletion are functionally compromised. This previously unrecognized role for CENP-E does not require its motor activity but instead is driven by farnesyl modification of its C-terminal kinetochore- and microtubule-binding domain. We show that in cells, CENP-E binds Spindly and recruits RZZ-S complexes to ectopic locations in a farnesyl-dependent manner. CENP-E is recruited to kinetochores following RZZ-S, and—while not required for RZZ-S oligomerization per se—promotes subsequent fibrous corona expansion. Our comparative genomics analyses suggest that the farnesylation motif in CENP-E orthologs emerged alongside the full RZZ-S module in an ancestral lineage close to the fungi–animal split (Obazoa), revealing potential conservation of the mechanisms for fibrous corona formation. Our results show that proper spindle assembly has a potentially conserved non-motor contribution from the kinesin CENP-E through stabilization of the fibrous corona meshwork during its formation. The cell division cycle culminates in the segregation of duplicated sister chromatids to nascent daughter cells. Segregation is driven by the microtubule-based spindle apparatus . MicrotuThe fibrous corona houses over two dozen proteins to perform its various roles in spindle assembly and chromosome segregation . It is fcomet, the Cyclin B1–CDK1 kinase complex, the CLASP1/2 microtubule dynamics regulators, and the microtubule-binding protein centromere protein F  was fully sufficient to restore fibrous corona formation after CENP-E loss, we next attempted to further narrow down the essential region. Expression in HeLa cells of a series of MT-binding domain mutants of CENP-E 2111-C  revealedC2698A delayed chromosome congression  have sm ZWINT-1 . The kin ZWINT-1 . In cont ZWINT-1 . Further ZWINT-1 , showing ZWINT-1  or expre ZWINT-1  abolisheOur data thus far showed that farnesylated CENP-E is important for fibrous corona formation and that CENP-E can quite robustly recruit RZZ-S to ectopic locations in a manner dependent on its farnesylation. To understand more about how CENP-E promotes fibrous corona formation, we asked when and how CENP-E localizes to kinetochores, relative to RZZ-S. Again we made use of the monopolar spindle assay, in which fibrous corona proteins are depleted from kinetochores and accumulate at the spindle pole upon chemical inhibition of CENP-E. Subsequent addition of nocodazole to these cells to deplete microtubules caused detectable relocalization of ZW10 back to the kinetochores within ∼1 min, followed by CENP-E after ∼8 min . ImportaΔN) bypasses the need for regulatory mechanisms of fibrous corona formation and does not bind dynein/dynactin, resulting in persistent fibrous coronas  should be able to bypass this step in the absence of CENP-E. This is not so, showing CENP-E contributes to a parallel or downstream event. An alternative hypothesis is that CENP-E is an integral component of the RZZ-S meshwork. However, RZZ-S can oligomerize in vitro without CENP-E . Our current analysis pinpoints emergence of SPINDLY one branch earlier than the cysteine in CENP-E, but we note that the CENP-E ortholog sequences in Breviata are incomplete. Better sampling of genomes from Breviata species might therefore reveal FL CENP-E ORF sequences with a cysteine at −4 from the carboxy terminus, which would indicate near-simultaneous emergence of SPINDLY and the potential for CENP-E farnesylation. Beyond SPINDLY, the RZZ-S complex and the potential for CENP-E farnesylation show strongly cohesive presence/absence in Obazoa, with notable coherent losses close in the species tree. Interestingly, kinetochores in colchicine-treated Drosophila S2 cells do not appear to form clear fibrous coronas  supplemented with 9% tetracycline-free fetal bovine serum and 100 μg/ml penicillin-streptomycin .ΔN, and LAP-SPINDLYΔN/C602A in pCDNA5 vector were previously cloned by PCR-based strategy; SPINDLYΔN: SPINDLY 65-605; SPINDLYΔN/C602A: SPINDLY 65-605 with C602A  to the C-terminal CENP-E 2111-C (comprising KT-binding domain and MT-binding domain). LAP-tagged CENP-E 2,111–2,697^CAAXLAP-ROD in pCDNA5 vector was cloned based on the ROD cDNA, which was a kind gift from Reto Gassmann , TagRFP-CENP-E 2111-C and H2B-mScarlet were cloned into the pLVX-IRES-Puro vector (Clontech) by PCR-based Gibson assembly method.CENP-E-1aa-XhoI-GA-F: 5′-CGGAGGTGGATCCACTAGTCTCGAGATGGCGGAGGAAGGAGCCGT-3′CENP-E-2,701aa-NotI-GA-R: 5′-GCGGGTTTAAACGGGCCCGCGGCCGCCTACTGAGTTTTGCACTCAGGCACATC-3′CENP-E-1,301-BamHI-F: 5′-GCACATGGACAGCGGAGGTGGATCCACTCAGGAAACAATGAATGAACTGGAGT-3′CENP-E-2,111-BamHI-F: 5′-GCACATGGACAGCGGAGGTGGATCCGATGATCATTATGAGTGCTTGAATAGATTGTCTCT-3′CENP-E 2,599-NotI-R: 5′-GCGGGTTTAAACGGGCCCGCGGCCGCCTATTGTTTGTGAGCCTCTCTTTTAAGGGTT-3′CENP-E 2,643-NotI-R: 5′-GCGGGTTTAAACGGGCCCGCGGCCGCCTAACAAGATTTTGGTGATTCCTTTGGCACA-3′CENP-E-2,600-mutation10AA-GA-R: 5′-TGCAGCCGCTGCAGCTGCCGCTGCGGCAGCTTGTTTGTGAGCCTCTCTTTTAAGGGTT-3′CENP-E-2,609-mutation10AA-GA-F: 5′-GCGGCAGCTGCAGCGGCTGCAAAAGTGACTGGAACAGCTTCTAAAAAGAAAC-3′CENP-E-2,610-mutation10AA-GA-R: 5′-TGCAGCCGCTGCAGCTGCCGCTGCGGCAGCAGGAGACTTTGGAGAATTCTCACAAG-3′CENP-E-2,619-mutation10AA-GA-F: 5′-GCGGCAGCTGCAGCGGCTGCACAAATTACACCCTCTCAATGCAAGGAAC-3′CENP-E-2,620-mutation10AA-GA-R: 5′-TGCAGCCGCTGCAGCTGCCGCTGCGGCAGCTTTCTTTTTAGAAGCTGTTCCAGTCAC-3′CENP-E-2,629-mutation10AA-GA-F: 5′-GCGGCAGCTGCAGCGGCTGCAAATTTACAAGATCCTGTGCCAAAGGAATC-3′CENP-E-2,630-mutation10AA-GA-R: 5′-TGCAGCCGCTGCAGCTGCCGCTGCGGCAGCCCGTTCCTTGCATTGAGAGGGTG-3′CENP-E-2,639-mutation10AA-GA-F: 5′-GCGGCAGCTGCAGCGGCTGCACCAAAATCTTGTTTTTTTGATAGCCGATC-3′CENP-E-2,640-mutation10AA-GA-R: 5′-TGCAGCCGCTGCAGCTGCCGCTGCGGCAGCTGATTCCTTTGGCACAGGATCTTG-3′CENP-E-2,649-mutation10AA-GA-F: 5′-GCGGCAGCTGCAGCGGCTGCAAAGTCTTTACCATCACCTCATCCAG-3′CENP-E-2,650-mutation10AA-GA-R: 5′-TGCAGCCGCTGCAGCTGCCGCTGCGGCAGCTGATCGGCTATCAAAAAAACAAGATTTTG-3′CENP-E-2,659-mutation10AA-GA-F: 5′-GCGGCAGCTGCAGCGGCTGCATATTTTGATAACTCAAGTTTAGGCCTTTGTC-3′CENP-E-2,660-mutation10AA-GA-R: 5′-TGCAGCCGCTGCAGCTGCCGCTGCGGCAGCGCGAACTGGATGAGGTGATGGTAA-3′CENP-E-2,669-mutation10AA-GA-F: 5′-GCGGCAGCTGCAGCGGCTGCACCAGAGGTGCAAAATGCAGGAG-3′CENP-E-2,670-mutation10AA-GA-R: 5′-TGCAGCCGCTGCAGCTGCCGCTGCGGCAGCACAAAGGCCTAAACTTGAGTTATCAAAAT-3′CENP-E-2,679-mutation10AA-GA-F: 5′-GCGGCAGCTGCAGCGGCTGCAGTGGATTCTCAGCCAGGTCCTTG-3′CENP-E-2,680-mutation10AA-GA-R: 5′-TGCAGCCGCTGCAGCTGCCGCTGCGGCAGCACTCTCTGCTCCTGCATTTTGC-3′CENP-E-2,689-mutation10AA-GA-F: 5′-GCGGCAGCTGCAGCGGCTGCATCCTCAGGCAAGGATGTGCCTG-3′CENP-E-2,689-NotI-R: 5′-GCGGGTTTAAACGGGCCCGCGGCCGCCTAGGCGTGCCAAGGACCTGGCTGAG-3′CENP-E-2,701-NotI-S2690A-R2: 5′-GCGGGTTTAAACGGGCCCGCGGCCGCCTACTGAGTTTTGCACTCAGGCACATCCTTGCCTGAAGCGGCGTGCCAAGGACCTG-3′CENP-E-2,701-NotI-S2691A-R2: 5′-GCGGGTTTAAACGGGCCCGCGGCCGCCTACTGAGTTTTGCACTCAGGCACATCCTTGCCAGCGGAGGCGTGCCAAGGAC-3′CENP-E-2,701-NotI-K2693A-R2: 5′-GCGGGTTTAAACGGGCCCGCGGCCGCCTACTGAGTTTTGCACTCAGGCACATCAGCGCCTGAGGAGGCGTGCCAAGGAC-3′CENP-E-2,701-NotI-D2694A-R2: 5′-GCGGGTTTAAACGGGCCCGCGGCCGCCTACTGAGTTTTGCACTCAGGCACAGCCTTGCCTGAGGAGGCGTGCCAAGGAC-3′CENP-E-2,701-NotI-V2695A-R: 5′-GCGGGTTTAAACGGGCCCGCGGCCGCCTACTGAGTTTTGCACTCAGGAGCATCCTTGCCTGAGGAGGCGTG-3′CENP-E-2,701-NotI-P2696A-R: 5′-GCGGGTTTAAACGGGCCCGCGGCCGCCTACTGAGTTTTGCACTCAGCCACATCCTTGCCTGAGGAGGC-3′CENP-E-2,701-NotI-E2697A-R: 5′-GCGGGTTTAAACGGGCCCGCGGCCGCCTACTGAGTTTTGCAAGCAGGCACATCCTTGCCTGAGGAG-3′CENP-E-C-C2698A-R2: 5′-GCGGGTTTAAACGGGCCCGCGGCCGCCTACTGAGTTTTAGCCTCAGGCACATCCTTGCCTGAGGAGGC-3′CENP-E-2,701-NotI-K2699A-R: 5′-GCGGGTTTAAACGGGCCCGCGGCCGCCTACTGAGTAGCGCACTCAGGCACATCCTTGCC-3′CENP-E-2,701-NotI-T2700A-R: 5′-GCGGGTTTAAACGGGCCCGCGGCCGCCTACTGAGCTTTGCACTCAGGCACATCCTTGC-3′CENP-E-2,701-NotI-Q2701A-R: 5′-GCGGGTTTAAACGGGCCCGCGGCCGCCTAAGCAGTTTTGCACTCAGGCACATCCT-3′CENP-E-2,701-NotI-C2698S-R: 5′-GCGGGTTTAAACGGGCCCGCGGCCGCCTACTGAGTTTTTGACTCAGGCACATCCTTGCCTGAGGAGGCGTG-3′CENP-E^KRAS_caax_R: 5′-GCGGGTTTAAACGGGCCCGCGGCCGCCTACATTATAATGCACTCAGGCACATCCTTGCCTGAGGAGGCGTG-3′TagRFP-kozak-EcoR1-F: 5′-CTACTAGAGGATCTATTTCCGGTGAATTCGCCACCATGGTGTCTAAGGGCGAAGAGCTGATTAAG-3′TagRFP-C-toCENP-E2111-R: 5′-TCTATTCAAGCACTCATAATGATCATCGGATCCACCTCCGCTATTAAGTTTGTGCCCCAGTTTGCT-3′ROD-1aa-XhoI-GA-F: 5′-CGGAGGTGGATCCACTAGTCTCGAGATGTGGAATGATATTGAGCTGCTAACAAATG-3′ROD-1,100aa-overlap-GA-R: 5′-AATAGCAATTTCCCAGTGTCAGCATT-3′ROD-1,100aa-overlap-GA-F: 5′-AATGCTGACACTGGGAAATTGCTATT-3′ROD-2,209aa-NotI-GA-R: 5′-GCGGGTTTAAACGGGCCCGCGGCCGCTTACGATAATCCACTAAGAAACATCTTCAGAAT-3′TurboID-N-BamHI-F: 5′-GCACATGGACAGCGGAGGTGGATCCAAAGACAATACTGTGCCTCTGAAGCT-3′TurboID-C-BamHI-R: 5′-TCAAGCACTCATAATGATCATCGGATCCGCTGAATTCCTTTTCGGCAGACCGCAGACT-3′CENP-E-1aa-BamHI-cut-GA-F: 5′-GCACATGGACAGCGGAGGTGGATCCATGGCGGAGGAAGGAGCCGTG-3′CENP-E-426aa-BamHI-cut-GA-R: 5′-ATTCAAGCACTCATAATGATCATCGGATCCGCCAAGGCACCAAGTAACTCTTC-3′Lentiviruses were produced by cotransfection of HEK 293T cells with the lentiviral vector containing TagRFP-CENP-E 2111-C or H2B-mScarlet and separate plasmids that express Tat, vesicular stomatitis virus G glycoprotein, Rev, and Gag-Pol, with the transfection reagent of Fugene HD . 72 h after transfection, cell culture supernatant was harvested and filtered.RPE1 FlpIn cell line stably expressing EB3-TagRFPT-ires-GFP-ZWILCH was made previously by transduction of viruses that were produced by cotransfection of HEK 293T cells with the lentiviral vector containing gene of EB3-TagRFPT-ires-GFP-Zwilch, which was cloned into the pLVX-IRES-Puro vector .ΔN were made previously by cotransfection of pCDNA5-constructs with gene of interest and construct of pOG44 recombinase in a 1:2 ratio with electroporation machine (Amaxa Nucleofector II) using protocol U-017. 3 d after transfection, RPE1 cells were selected with 100 μM hygromycin  for about 3 wk (RPE1 FlpIn cell line inducible expressing LAP-SPINDLY FL and LAP- SPINDLYout 3 wk .RPE1 FlpIn cell lines were cotransfected with pCDNA5-constructs with gene of interest and construct of pOG44 recombinase in a 1:2 ratio with an electroporation machine (Amaxa Nucleofector II) using protocol U-017. 3 d after transfection, RPE1 cells were selected with 100 μM hygromycin  for about 3 wk.HeLa FlpIn were cotransfected with pCDNA5 constructs with the gene of interest and construct of pOG44 recombinase in a 1:9 ratio with Fugene HD  under the manufacturer’s instructions. 1 d after transfection, HeLa cells were selected with 200 μM hygromycin  for about 3 wk.To generate HeLa FlpIn cells stably expressing TagRFP-CENP-E 2111-C and inducible expressing LAP-ROD, HeLa FlpIn cells inducible expressing LAP-ROD were infected with lentiviruses and selected with 1 μg/ml puromycin for 1 wk.To generate HeLa FlpIn cell stably expressing H2B-mScarlet, HeLa FlpIn were infected with lentiviruses and selected with 1 μg/ml puromycin for 1 wk.To generate HeLa FlpIn cells inducible expressing CENP-E FL WT or CENP-E FL C2698A and stably expressing H2B-mScarlet, HeLa FlpIn cells inducible expressing CENP-E FL WT or CENP-E FL C2698A were infected with lentiviruses and selected with 1 μg/ml puromycin for 1 wk.The siRNAs targeting GAPDH , CENP-E  for 15 h (overnight) for fibrous corona formation before the fixation. HeLa FlpIn cells were treated with 6.6 μM nocodazole for 6∼8 h before the fixation. For cells transfected with siRNA, nocodazole was added 48 h after siRNA transfection. For the rescue experiments, 1 μg/ml doxycycline  was added to cells 40 h after siRNA transfection to induce the expression of protein mutants. For nocodazole washout assay, 6.6 μM nocodazole was added to the culture medium for 15 h before washout. For CENP-E inhibition, cells were treated with 250 nM GSK923295  for 15 h (overnight). For farnesyl transferase inhibition, cells were treated with 5 µm lonafarnib  for 15 h (overnight). For the fast disassembly of microtubules, cells were treated with 13.2 μM nocodazole for 4 h. Cells were treated with 10 mM STLC  for 15 h (overnight). For the experiments in which translocated fibrous corona proteins were released from spindle poles, cells were cotreated with STLC and GSK923295 for 15 h (overnight) and were further treated with 13.2 μM nocodazole for indicated time before fixation. In the TurboID pulldown experiment, HeLa cells were treated with 1 μg/ml doxycycline for 24 and 1 h of 250 μM biotin  before harvest. For spindle end-on attachment experiments, HeLa cells were cotreated with STLC and ZM447439  for 6 h before fixation.Glued . We used the following secondary antibodies: Goat anti-guinea pig Alexa Fluor 647 , Goat anti-rabbit Alexa Fluor 488 , Goat anti-rabbit Alexa Fluor 568 , Goat anti-rabbit Alexa Fluor 405 , Goat anti-mouse Alexa Fluor 568 , Goat anti-mouse Alexa Fluor 647 , and GFP-Booster Atto 488 .We used guinea pig polyclonal antibody against CENPC , and rabbit polyclonal antibodies against CENP-E , ZW10 , and were blocked with 2% BSA diluted in PBST for 30 min. After blocking, cells were incubated with primary antibodies diluted in PBST containing 2% BSA for 1 h at room temperature in humid conditions. Subsequently, cells were washed three times with PBST and incubated with secondary antibodies diluted in PBST together with or without DAPI for 1 h at room temperature. Then, cells were washed three times with PBST. In the end, coverslips were sequentially rinsed in 70% and 100% ethanol, air-dried, and mounted on glass slides with Prolong Gold antifade.n = 1.5160  using SoftWorx 6.0 software . Images were acquired as z-stacks at 0.2-μm intervals for 32 stacks using Photometrics Coolsnap HQ2 cameral (Roper Scientific) and deconvolved using SoftWoRx. The images are maximum-intensity projections of deconvoluted stacks.All images were acquired on a deconvolution system  with a 100×/1.40 NA UPlanSApo objective (Olympus) and imaging medium with https://fiji.sc). For quantification of protein levels, all images of immunostaining experiments were acquired with identical illumination settings. Protein levels near the kinetochores were determined on maximum projections of z-stacks images using an ImageJ macro . 488 and 561 nm lasers were used for sample excitation and images were acquired using an Andor iXon-888 EMCCD camera. Nocodazole washout was done during the imaging with a warm culture medium. Before and after nocodazole washout, cells were kept at 37°C and 5% CO2 using a cage incubator and Boldline temperature/CO2 controller (OKO-Lab).For imaging of RPE1 cells coexpressing EB3-TagRFPT and GFP-Zwilch in nocodazole washout assay, RPE1 cells were cultured in a 24-well glass bottom plate . Live-cell imaging was performed on a Nikon TiE microscope, controlled by NIS Elements software , by acquiring images every 20 or 30 s with z-stacks at 1-μm intervals for 15 stacks at 1  ×  1 binning on an Andor CSU-W1 spinning disk (50 μm disk) with Nikon 100× 1.45 NA oil objective and immersion oil with n = 1.406 . 488 and 561 nm lasers were used for sample excitation and images were acquired using an Andor iXon-888 EMCCD camera. Cells were kept at 37°C and 5% CO2 using a cage incubator and Boldline temperature/CO2 controller (OKO-Lab).Imaging of HeLa FlpIn cells expressing H2B-mScarlet and HeLa FlpIn cells coexpressing LAP-CENP-E FL WT or LAP-CENP-E FL C2698A with H2B-mScarlet that were cultured in 8-well plates  was performed on a Nikon TiE microscope, controlled by NIS Elements software , by acquiring images every 3 min with z-stacks at 2-μm intervals for eight stacks at 1  ×  1 binning on an Andor CSU-W1 spinning disk (50-μm disk) with Nikon 30× silicon objective and silicon immersion oil with In the TurboID-based proteins pulldown experiments, HeLa cells inducible expressing LAP-TurboID-CENP-E 2111-C mutants were treated with 1 μg/ml doxycycline for 24 h to induce the expression of LAP-TurboID-CENP-E 2111-C mutants and 1 h of 250 μM biotin before harvest. The cells were harvested by trypsinization and centrifuge. The cells were washed once with ice-cold PBS and lysated with lysis buffer  on ice for 10 min. After lysis, the samples were centrifuged at 14,000 rpm for 20 min at 4°C. The supernatants of the centrifuged samples were incubated with the streptavidin magnetic beads  for 1 h in a 4°C cold room. The beads were washed with wash buffer  three times. The wash buffer was removed after the final wash and the beads were stored in a −80°C freezer.2 Fold Change and −log10 P value higher than 2. The program used for the analyses was R (version 4.0.4) through RStudio (version 1.5.64).Precipitated proteins were denatured and alkylated in 50 µl 8 M urea, 1 M ammonium bicarbonate (ABC) containing 10 mM tris (2-carboxyethyl) phosphine hydrochloride and 40 mM 2-chloro-acetamide for 30 min. After fourfold dilution with 1 M ABC and digestion with trypsin (20 µg/200 µl), peptides were separated from the beads and desalted with homemade C-18 stage tips (3 M), eluted with 80% acetonitrile (ACN) and, after evaporation of the solvent in the speedvac, redissolved in buffer A (0.1% formic acid [FA]). After separation on a 30-cm pico-tip column  in-house packed with C-18 material  using a 140 min gradient , delivered by an easy-nLC 1,000 (Thermo Fisher Scientific), peptides were electrosprayed directly into a Orbitrap Fusion Tribrid Mass Spectrometer (Thermo Fisher Scientific). The latter was set in data-dependent top speed mode with a cycle time of 1 s, in which the full scan over the 400–1,500 mass range was performed at a resolution of 240,000. Most intense ions  were isolated by the quadrupole, where after they were fragmented with a higher-energy collisional dissociation collision energy of 30%. The maximum injection time of the ion trap was set to 50 ms with injection of ions for all available parallelizable time. Raw data were analyzed with MaxQuant (version 1.6.3.4) using the Homo Sapiens (taxonomy ID: 9606) fasta file, extracted from UniprotKB. To determine proteins of interest, the protein group output file was used to perform a differential enrichment analysis. Proteins with less than one unique peptide and proteins that have not been identified in at least two out of three of the replicates of one condition were filtered out. Then, a background correction and normalization of the data were performed by variance stabilizing transformation, shifting and scaling the protein intensities by sample group. A left-shifted Gaussian distribution randomization was used to impute since the data presented a pattern of missingness not at random. Finally, a differential enrichment analysis was performed to identify those proteins that were differentially enriched and selected those falling inside the threshold for logThe mass spectrometry proteomics data have been deposited to the ProteomeXchange Consortium via the PRIDE  partner Preprint), with some minor adjustments detailed in The eukaryotic sequence database used for the evolutionary analysis consisted of 177 predicted proteomes derived from genomes or transcriptomes and spans a representative set of species across the eukaryotic tree of life. This dataset is derived from earlier work  and Prism 9 . Data distribution was assumed to be normal but this was not formally tested. The results of The data generated are available in the published article and its online supplemental material. The mass spectrometry proteomics data have been deposited to the ProteomeXchange Consortium via the PRIDE partner repository with the dataset identifier PXD040338.


In the title compound, which is related to the herbicide flufenacet, the amide group and fluorobenzene ring are almost perpendicular. A short O⋯π contact is observed in the crystal. 12H16FNO3S, which is related to the herbicide flufenacet, are presented. The dihedral angle between the amide group and the fluorinated benzene ring is 87.30 (5)° and the N—C—C—S torsion angle defining the orientation of the methylsulfonyl substituent relative to the amide group is 106.91 (11)°. In the crystal, inversion-related molecules form dimers as a result of pairwise C—H⋯O hydrogen bonds, which appear to be reinforced by short O⋯π contacts [O⋯Cg = 3.0643 (11) Å]. A Hirshfeld surface analysis was used to quantify the various types of intermolecular contacts, which are dominated by H atoms.The synthesis and crystal structure of the title compound, C For example, substituted phenylacetamides and their use as protease inhibitors was reported by Kreutter et al. -yl]acetamides was given by Ahmad et al. (2013N4-substituted sulfonamide–acetamide derivatives as dihydrofolate reductase (DHFR) inhibitors was reported by Hussein et al. (2019N-(substituted phenyl)-N-(substituted)acetamide derivatives as potent analgesic agents was described by Verma et al. -2-phenylacetamide derivatives as potential antimycobacterial agents was reported by Güzel-Akdemir et al. , systematic name N-(4-fluorophenyl)-N-propan-2-yl-2-{oxy}acetamide, is an herbicide, xenobiotic and environmental contaminant   Fig. 1.2.I, the nitrogen atom of the amide group is close to planar, the sum of bond angles about N1 being 358.92 (19)°, which places N1 0.0862 (14) Å from the plane passing through C1, C4, and C7. The amide group is also almost planar, having an r.m.s. deviation from the mean plane of N1, C1, O1, C2 of 0.0095 Å [maximum = 0.0165 (11) Å for C1], and is almost perpendicular to the fluorobenzene ring (C7–C12), subtending a dihedral angle of 87.30 (5)°. The overall conformation of the molecule is defined by the torsion angles C7—N1—C1—C2 [14.68 (17)°], N1—C1—C2—S1 [106.91 (11)°], C1—C2—S1—C3 [74.53 (10)°] and by the orientation of the ipropyl group, e.g., C1—N1—C4—C6 [139.85 (13)°]. Otherwise, all bond lengths and angles lie within the expected ranges.In the crystal structure of 3.I  1 – x, 1 – y, 1 – z]. These dimers also feature close contacts between the sulfone O3 atom and the inversion-related benzene ring to give an O3⋯Cg(C7–C12)i distance of 3.0643 (11) Å . The other weak C—H⋯O interactions involve inversion, translation, and c-glide related molecules (Table 1a). A Hirshfeld surface analysis using CrystalExplorer .There are no strong hydrogen bonds in the crystal structure of I Fig. 2, but thes Table 1a. A Hir4.et al., 2016N-phenylacetamide with ‘any non-H group’ attached at the nitrogen atom, the 4-position of the benzene ring, and replacing one hydrogen of the methyl group, yielded 259 hits. A similar fragment, but with ‘any halogen’ at the 4-position on the ring, gave 92 hits. With the halogen restricted to fluorine, twelve hits were returned, and with an isopropyl group attached to the nitrogen atom, only one match was found: CSD refcode QEMHOG  was presented by Gung et al. (2008A search of the CSD for non-bonded close contacts (up to 3.1Å) between S=O oxygen atoms and a benzene-ring centroid (with ‘any substituent’) returned 154 hits, none of which have much else in common with  al. 2008.d-threo-2,2-dichloro-N-{2-hydroxy-1-(hydroxymethyl)-2-[4-(methylsulfonyl)phenyl]ethyl}acetamide -2-hydroxy-2-[4- (methylsulfonyl)phenyl]ethyl}acetamide -2-(2-{3-[4-(methylsulfonyl)phenyl]-1,2,4-oxadiazol-5-yl}phenoxy)acetamide -N-(methylsulfonyl)acetamide -N-(methylsulfonyl)acetamide phenyl)acetamide -4-nitrophenyl]-N-methylacetamide acetamide acetyl chloride were added. The mixture was stirred at room temperature for 5 h. After this, 100 ml of water were added and the mixture was extracted three times, each with 100 ml of methyl tert-butyl ether (MTBE). The combined organic phases were dried over MgSO4 and the solvent was evaporated under reduced pressure. The crude product, N-(4-fluorophenyl)-N-isopropyl-2-(methylthio)acetamide, was used for the next stage with purification (7.5 g).In a 250 ml flask (with a nitrogen inlet and a septum) was placed 5 g of 4-fluoro-N-(4-fluorophenyl)-N-isopropyl-2-(methylthio)acetamide dissolved in 150 ml of dichloromethane. After cooling to 263–273 K, 13.37 g of meta-chloroperbenzoic acid in 100 ml dichloromethane was added slowly at the same temperature. The mixture was stirred at room temperature for 5 h. After this, 200 ml of water were added and the organic layer was separated, and washed with 100 ml of 10% sodium bicarbonate twice. The organic phases were dried over MgSO4 and the solvent was evaporated under reduced pressure. The crude product was purified by chromatography over SiO2 (hexane:ethyl acetate 9:1 v/v). The title compound was recrystallized from diethyl ether solution in the form of colorless plates. The overall reaction scheme is shown in Fig. 4To a 250 ml round-bottomed flask (with a nitrogen inlet and a septum) was added 7.5 g of 1H NMR: CDCl3 : 1.097–1.08 ; 3.198 ; 3.664 ; 5.006–4.938 ; 7.273–7.132 . MS m/z: 273.45 (M)+.6.sp2H), 0.98 Å (RCH3), 0.99 Å (R2CH2) and 1.00 Å (R3CH). Uiso(H) parameters were set to values of either 1.2Ueq or 1.5Ueq (RCH3 only) of the attached atom.Crystal data, data collection and structure refinement details are summarized in Table 210.1107/S2056989023003675/hb8062sup1.cifCrystal structure: contains datablock(s) I, global. DOI: 10.1107/S2056989023003675/hb8062Isup2.hklStructure factors: contains datablock(s) I. DOI: Click here for additional data file.10.1107/S2056989023003675/hb8062Isup3.cmlSupporting information file. DOI: 2258160CCDC reference:  crystallographic information; 3D view; checkCIF reportAdditional supporting information:


The capsid p24 (CA-p24) antigen is a component of the viral capsid of human immunodeficiency virus (HIV) that has been commonly used for clinical diagnosis and monitoring of HIV infections in Enzyme-linked Immunosorbent Assays (ELISAs). Commercial CA-p24 ELISAs are widely used in research settings, but these kits are costly and have limited breadth for detecting diverse HIV isolates.Commercial CA-p24 antibodies were used as capture and detection antibodies. Specific CA-p24 ELISAs were established with these antibodies and tested for the detection of HIV-1 isolates with the aim of developing in-house protocols to recognize HIV-1 infections in vitro for research purposes.Here we present four protocols for in-house ELISAs to detect HIV CA-p24 using commercial antibodies. The assays were able to detect the CA-p24 antigen of different HIV-1 isolates tested. Comparison between the protocols showed that these in-house ELISAs exhibit high specificity, sensitivity, and reproducibility for CA-p24 quantitation but their reactivity varied per HIV-1 isolate and subtype.These optimized ELISA protocols represent valuable tools to investigate HIV-1 infections in research facilities at a lower price than commercial CA-p24 kits.The online version contains supplementary material available at 10.1186/s12985-023-02242-5. HIV infections are prevalent worldwide. If left untreated, HIV can progress to AIDS (Acquired Immunodeficiency Syndrome) and cause more than 600,000 deaths every year . The curCurrent research approaches include viral genome editing through CRISPR-Cas systems, stem cell transplants, and mRNA vaccines –7. SucceIn this study, we aimed to optimize and compare four in-house CA-p24 ELISA systems to detect and quantitate distinct HIV-1 isolates. We determined the reproducibility and sensitivity of the ELISA systems and described their characteristics for testing CA-p24 antigen.q = 0.0065, Kruskal–Wallis’ test), and 45 × lower than very high-cost commercial kits (q = 0.0048) , Anogen (ANG), Sino Biological (SB), and R&D Systems (RND). The costs per each in-house assay range between €25 and €30 per one 96-well plate, which is 15 × lower than medium-cost commercial kits, 25 × lower than high-cost commercial kits , possibly due to the use of the same detection antibody. Similarly, SB-CA-p24 ELISA showed a significantly high correlation with ABR-CA-p24 ELISA . RND-CA-p24 ELISA also exhibited a significant high correlation with ABR-CA-p24 ELISA  , suggesting a similar sensitivity between both assays, possibly due to their similar reactivity for HIV-1 B subtypes. The lowest correlations were observed between RND- and SB-CA-p24 ELISAs  and between ANG- and SB-CA-p24 ELISAs  , suggesting a high between-assay reproducibility for the ELISA systems.We then assessed the reproducibility of the ELISA systems, examining both their consistency within individual assays and their reliability between different assays. In order to assess within-assay reproducibility, we employed the NL4-3 HIV-1 isolate as a test model. We quantified its concentration in each assay by utilizing technical replicates and applying different inactivation treatments, including heat and/or detergent. We observed that the concentration of the NL4-3 isolate remained consistent between the replicates within the tested conditions  guidelines classify HIV-1 as a Risk Group 3 agent. Research involving HIV-1 was approved by the institutional biosafety office and carried out according to laboratory biosafety 3 guidelines.Twenty-two different HIV-1 isolates were produced according to standard protocols . The HIVVirus stocks were generated on CD8 + -depleted human peripheral blood mononuclear cells (PBMCs) for each isolate, including the lab-adapted strains, and virus production was measured by CA-p24 ELISA (in-house assays). The HIV stocks were inactivated with 0.1% Empigen  followed by incubation at 56 °C for 30 min.Aalto Bio Reagents (ABR) system2O. Adjust pH to 7.4 with HCl and add ddH2O up to a volume of 1,000 mL. Store at room temperature (20–25 °C). Dilute 1:10 with ddH2O for 1 × TBS buffer.10 × Tris buffered saline (TBS) buffer: 1.37 M NaCl (80 g) + 27 mM KCl (2 g) + 2.4 g of Tris in a volume of 950 ml of ddHFirst wash buffer solution and dilution buffer:1 × TBS + 0.05% Empigen: Add 83 ml of 30% Empigen into 500 ml of 1 × TBS buffer (Table 2O. Adjust pH to 7.4 with HCl up to 1000 mL and purify with a 0.2 µm filter and store at room temperature (20–25 °C). Dilute 1:10 with ddH2O for 1 × PBS buffer.10 × phosphate-buffered saline (PBS): 20 PBS tablets and up to 950 mL of ddHSecond wash buffer solution:2O for 1 × PBS buffer + 0.1% Tween.10 × PBS buffer + 1% Tween. Add 10 ml of Tween 20 into 990 ml of 10 × PBS buffer. Store at room temperature (20–25 °C). Dilute 1:10 with ddH3: 4.2 g NaHCO3 and up to 450 mL of ddH2O. Adjust pH to 8.5 and add ddH2O up to a volume of 500 mL. Store at room temperature (20–25 °C).0.05 M NaHCOCoating buffer and antibody:3 (10 µg/mL) and dispense 25 µl of capture antibody solution into each well.Anti-HIV Gag CA-p24: reconstitute according to manufacturer’s instructions. Make 100 µl aliquots and store at − 20 °C until use. To coat one 96-well plate, add 25 µl of aliquoted capture antibody into 2,475 µl of 0.05 M NaHCOCA-p24 protein: reconstitute with 1 × TBS, 20% Sheep serum, and 1% Empigen to a concentration of 10 µg/mL. Dilute 1:10 with 1 × TBS + 1% Empigen to a concentration of 1 µg/mL. Make 30 µl aliquots and store at -80 °C until use.CA-p24 standard curve: add 25 µl of aliquoted CA-p24 standard into 225 µl of dilution buffer (1 × TBS + 0.05% Empigen) (working stock). Prepare standard curve dilutions as specified in Additional File CA-p24 standard protein and curve:ABR, ANG):HIV-CA-p24, Alkaline Phosphatase Conjugate monoclonal antibody (AP-mAb): reconstitute and dilute 1:5 by adding 20 µl of antibody into 80 µl triethanolamine. Store at 4 °C until use.Prepare the conjugate for one 96-well plate as followed: 2 mL of 1 × TBS + 0.5 mL of sheep serum + 2.5 µl of Tween 20 + 0.05 g of skim milk (2%) + 0.31 µl of the reconstituted conjugated . Dispense 25 µl of conjugate solution into each well.Detection antibody, conjugate, and substrate  into each well Table .2.Sino Biological (SB) and Anogen (ANG) systemsABR protocol). Store at room temperature (20–25 °C). Dilute 1:10 with ddH2O for 1 × PBS buffer + 0.05% Tween : to prepare the blocking buffer needed for one 96-well plate, add 0.2 g of BSA into 10 ml of 1 × PBS buffer + 0.05% Tween.1 × PBS + 0.05% Tween + 0.1% BSA: to prepare the dilution buffer needed for one 96-well plate, dilute 1:20 the blocking buffer into new 1 × PBS buffer + 0.05% Tween as needed.SB): reconstitute according to manufacturer’s instructions. To coat one 96-well plate, add 5.6 µl of coating antibody into 2.8 mL of 1 × PBS (2 µg/mL) and dispense 25 µl of coating antibody solution into each well (Table Mouse Anti-HIV-CA-p24 (Coating antibodies:ANG): reconstitute according to manufacturer’s instructions. To coat one 96-well plate, add 25 µl of aliquoted coating antibody into 2,475 µl of 1 × PBS (10 µg/mL) and dispense 25 µl of coating antibody solution into each well  into 225 µl of dilution buffer (1 × PBS + 0.05% Tween + 0.1% BSA) (working stock). Prepare standard curve dilutions as specified in Additional File CA-p24 standard curve:SB): reconstitute according to manufacturer’s instructions. Dilute 1:1,000 in HRP stabilizer (Table Mouse Anti-HIV-CA-p24 horseradish peroxidase (HRP) antibody (SB): to prepare the conjugate for one 96-well plate, dilute 1:1,000 aliquoted detection antibody by adding 3 µl of antibody into 3,000 µl of dilution buffer (100 ng/mL) and dispense 25 µl of conjugate solution into each well.Conjugate (SB): dilute 1:10 Lumiphos A and B solutions by adding 150 µl of solution A and 150 µl of solution B into 2.7 ml of ddH2O. Dispense 25 µl of LumiPhos A + B solution into each well SB/ANG protocols) with ddH2O.1 × PBS buffer + 0.05% Tween: dilute 1:10 10 × PBS buffer + 0.5% Tween  into 45 µl of dilution buffer (1 × PBS + 0.2% TritonX-100 + 1% BSA) (working stock). Prepare standard curve dilutions as specified in Additional File CA-p24 standard curve: add 5 µl of aliquoted CA-p24 standard (CA-p24 standard curve:Biotinylated Mouse Anti-HIV-1 Gag CA-p24 Detection Antibody: reconstitute according to manufacturer’s instructions with 1.0 ml of dilution buffer (1 × PBS + 0.2% TritonX-100 + 1% BSA). Make 45 µl aliquots and store at − 20 °C until use. To prepare the antibody solution for one 96-well plate, add 41.8 µl of aliquoted detection antibody into 5 mL of 1 × PBS + 0.2% TritonX-100 + 1% BSA buffer (125 ng/mL) and dispense 50 µl of detection antibody solution into each well.Conjugate: to conjugate the detection antibody in one 96-well plate, dilute 1:80 Streptavidin-HRP solution by adding 62.5 µl of antibody into 5 ml of 1 × PBS + 0.2% TritonX-100 + 1% BSA buffer and dispense 50 µl of solution into each well.2O  was used to analyze datasets and determine the sensitivity of the assays and Pearson’s Additional file 1: Supplementary Figs. 1–4. (.pdf). Fig. S1. Comparison of costs between in-house and commercial ELISA kits. Fig. S2. Standard curves generated by ABR-, ANG-, SB-, and RND-CA-p24 ELISAs. Fig. S3. Effect of viral inactivation on assay performance and within-assay reproducibility of ELISA systems. Fig. S4. Between-assay reproducibility of the ELISA systems.Additional file 2: Supplementary Tables 1–4 (.pdf). Table S1. List of commercial ELISA kits and their costs. Tables S2–S4. Four tables with the volumes and details needed to prepare the standard curves for the ABR-, ANG-, SB-, and RND-CA-p24 ELISAs.


A short and directional tetrel bond between tin and oxygen is identified in the title compound. 6H5)3]·C18H21OP, is reported. The 1:1 cocrystal features a short and directional tetrel bond between tin and oxygen. The tin–oxygen distance is 2.346 (4) Å, representing 62% of the sum of the van der Waals radii of Sn and O. The Cl—Sn⋯O angle is 174.0 (1)° and this nearly linear arrangement is consistent with a tetrel bond formed via a σ-hole opposite the tin–chlorine covalent bond. Some weak C—H⋯Cl interactions are noted between adjacent molecules.The single-crystal X-ray diffraction structure of the title compound, [SnCl(C The asymmetric unit consists of one complete molecule of each type. The tin–oxygen distance is 2.346 (4) Å and the Cl—Sn⋯O TB angle is 174.0 (1)°  Å and the Cl—Sn⋯O TB angle is 177.57 (7)° (p-tolyl)tin]-μ-1,2-bis(diphenylphosphoryl)ethane-κ2O:O′-[bromidobis(p-chlorophenyl)(p-tolyl)tin] 2·(Me2N)2PO tin–hexamethylphosphoramide , a moderately strong and directional noncovalent interaction, has received renewed interest in recent years as a useful structure-directing element and crystal engineering tool ° Fig. 1. This divia slow evaporation of the solvent in a fume hood over a period of 5 d. Evidently, during the synthesis, the phosphane was oxidized to give the phosphane oxide, as the process was not carried out under an inert atmosphere.In a typical procedure, triphenyltin(IV) chloride (0.0614 g) and cyclohexyldiphenylphosphane (0.0894 g) were added to hexane (60 ml) in a beaker. The mixture was heated and stirred until the solids were completely dissolved. Cocrystals grew The crystal data, data collection and structure refinement details are summarized in Table 210.1107/S2414314623006375/bv4048sup1.cifCrystal structure: contains datablock(s) I. DOI: 10.1107/S2414314623006375/bv4048Isup2.hklStructure factors: contains datablock(s) I. DOI: Click here for additional data file.10.1107/S2414314623006375/bv4048Isup3.cmlSupporting information file. DOI: 2267964CCDC reference:  crystallographic information; 3D view; checkCIF reportAdditional supporting information:


Hirshfeld surface analysis indicates that H⋯H and H⋯C/C⋯H contacts are the most important interactions.A new copper(II) complex with 3-methylbenzoate and 2,2′-bipyridine synthesized displays chains of hydrogen-bonded complex units along the  2N,N′)bis(3-methylbenzoato)-κ2O,O′;κO-copper(II) 0.68-hydrate, [Cu(C8H7O2)2(C10H8N2)(H2O)]·0.68H2O or [Cu(3-mb)2(bipy)(H2O)]·0.68H2O. The coordination environment of CuII is a distorted octahedron. The metal atom is attached to two 3-mb moieties, which bind in monodentate and bidentate fashions. One of the 3-mb units is disordered. The coordination environment is completed by one bipy ligand and a water molecule. A second water molecule is outside the coordination sphere of the CuII atom and its occupancy refined to 0.68. The structure consists of chains along the b-axis direction formed by complex units joined via hydrogen bonds between the coordinated water molecule and an O atom of a coordinated 3-mb unit. Hirshfeld surface analysis indicates that the most abundant contacts are H⋯H (56.8%), H⋯C/C⋯H (21.7%) and H⋯O/O⋯H (13.7%).3-Methylbenzoic acid (3-mbH) and 2,2′-bipyridine (bipy) reacted with a copper(II) salt forming a new mixed ligand complex, aqua(2,2′-bipyridine-κ The complex was octahedral with 3-mb acting as monodentate  complexes continues to be of interest. Copper is an important part of various metalloenzymes. It takes part in many metabolic processes such as iron metabolism, mitochondrial oxidative phosphorylation and catecholamine production  to 2.0131 (18) Å and the N1—Cu1—N2 angle is 80.58 (7)° (Table 1carboxylate distances are 1.842 (17)–2.2988 (18) Å. The Cu—O and Cu—N values are very close to those reported for copper(II) complexes involving benzoate (BZA) as a ligand, for example [Cu(BZA)2(bipy)(H2O)] [Cu—O = 1.9951 (12)–1.9633 (12) Å and Cu—N = 2.0064 (14)–2.0111 (13) Å; Devereux et al., 2007Complex 1 Fig. 1 crystall° Table 1. The Cu13.B⋯O4) leads to the formation of a linear chain in the b-axis direction  refined to 0.68, which seems to be due to water escaping the crystal through the channels that run along the b-axis direction.In the crystal, hydrogen bonding between H atoms of the coordinated water molecule and the O atoms of the coordinated 3-mb , H⋯C/C⋯H  and H⋯O/O⋯H  contacts are 56.8, 21.7 and 13.7%, respectively.⋯H Fig. 4c and H⋯⋯H Fig. 4d contac6.3)2·3H2O  in water (20 ml) under stirring. A solution of 2,2′-bipyridine  in EtOH (25 ml) was added and the color changed from greenish blue to blue. The precipitate was filtered off, washed with water and dried. Blue single crystals of the title complex suitable for X-ray diffraction studies were obtained after evaporation of an ethanol solution after several days.3-Methylbenzoic acid  and sodium hydroxide  in water (20 ml) were added to a solution of Cu(NO7.A–O6–H6B refined to 0.680 (10). The coordinates of the ordered water atom were refined with Uiso(H) = 1.5Ueq(O). All other H atoms were positioned geometrically and refined as riding with Uiso(H) = 1.2–1.5Ueq(parent atom).Crystal data, data collection and structure refinement details are summarized in Table 210.1107/S2056989023006904/dj2063sup1.cifCrystal structure: contains datablock(s) I. DOI: 10.1107/S2056989023006904/dj2063Isup2.hklStructure factors: contains datablock(s) I. DOI: 2117143CCDC reference:  crystallographic information; 3D view; checkCIF reportAdditional supporting information:


The formation of a co-crystal sustained by π-type halogen bonds involving 1,4-diiodoperchlorobenzene and naphthalene is reported. 6I2Cl4) as the halogen-bond donor along with naphthalene (nap) as the acceptor is reported. The co-crystal  generates a chevron-like structure that is held together primarily by π-type halogen bonds (i.e. C—I⋯π contacts) between the components. In addition, C6I2Cl4 also interacts with the acceptor via C—Cl⋯π contacts that help stabilize the co-crystal. Within the solid, both aromatic components are found to engage in offset and homogeneous face-to-face π–π stacking interactions. Lastly, the halogen-bond donor C6I2Cl4 is found to engage with neighboring donors by both Type I chlorine–chlorine and Type II iodine–chlorine contacts, which generates an extended structure.The formation and crystal structure of a co-crystal based upon 1,4-diiodoperchlorobenzene (C Recently, we reported the formation of photoreactive co-crystals based upon C6I2Cl4 along with trans-1,2-bis(pyridine-4-yl)ethylene  resulting in a chevron-like structure. In addition to the π-type halogen bond, the co-crystal (C6I2Cl4)·(nap) is also held together by the combination of C—Cl⋯π contacts, homogeneous face-to-face π–π stacking interactions, Type I chlorine–chlorine contacts, and Type II iodine–chlorine contacts.A continued goal within our research groups has been in the design and formation of halogen-bonded co-crystals based upon 1,4-diiodoperchlorobenzene (C2.6I2Cl4)·(nap) crystallizes in the centrosymmetric triclinic space group Pī. The asymmetric unit contains half a molecule of both C6I2Cl4 and nap where inversion symmetry generates the remainder of each molecule ·(nap), the donor C6I2Cl4 is found to engage in Type I chlorine–chlorine contacts  1 − x, -y, 1 − z], with a distance of 3.499 (1) Å and a C—Cl⋯Cl bond angle of θ1 = θ2 = 132.16 (6)°  1 − x, -y, 1 − z], with a distance of 3.808 (1) Å and a C—I⋯Cl bond angle of 111.83 (4)°. Both the aromatic halogen-bond donor and acceptor are found to engage in an offset and homogeneous face-to-face π–π stacking arrangement that stabilizes the co-crystal  Å measured for Cl1⋯C5.In addition to π-type halogen bond within ·(nap). This Hirshfeld surface analysis along with the observed bond lengths confirms the ability of C6I2Cl4 to engage in π-type halogen bonds to a polycyclic aromatic hydrocarbon, namely nap.These various non-covalent interactions were also investigated and visualized by utilizing a Hirshfeld surface analysis  in which the I atom is within the van der Waals radius of an aromatic surface revealed only one structure, refcode HONBIY ·(nap).A search of the Cambridge Crystallographic Database  was synthesized utilizing a previously published method  were both purchased from Sigma-Aldrich Chemical  and used without any additional purification. The halogen-bond donor 1,4-diiodoperchlorobenzene ·(nap) was achieved by dissolving 50.0 mg of C6I2Cl4 in 2.0 mL of toluene and then combined with a 2.0 mL toluene solution containing 13.7 mg of nap . Within two days, single crystals suitable for X-ray diffraction were formed upon loss of some of the solvent by slow evaporation.The formation of  I. DOI: 10.1107/S2056989023008356/jy2037Isup2.hklStructure factors: contains datablock(s) I. DOI: Click here for additional data file.10.1107/S2056989023008356/jy2037Isup3.cmlSupporting information file. DOI: 2291675CCDC reference:  crystallographic information; 3D view; checkCIF reportAdditional supporting information:


In the title complex, bis{μ-1,1′-(o-xylylene)bis[4-(pyridin-2-yl)triazole]-κN3:N3′}dipalladium(II) tetrakis(tetrafluoridoborate)–dimethylformamide–diethyl ether (1/2/1), as the BF4 salt, and including dimethylformamide and diethyl ether solvent molecules, with stoichiometry [Pd2(C22H18N8)2](BF4)4·2C3H7NO·C4H10O, the Pd complex and the disordered diethyl ether molecule lie on inversion centers. The ligand coordinates to the PdII centers with square-planar geometry, forming a dimeric macrocycle. The Pd⋯Pd separation in the complex [Pd2(o-xpt)2]4+ cation is 3.6184 (4) Å. In the crystal, the complex molecules are stacked along the b axis, with π–π interactions between the pyridyltriazole ligands of two molecules.A tetradentate ligand, namely,  Herein, we report the synthesis and crystal structure of the title compound.The self-assembly of polydentate ligands with transition-metal ions to create functional metal–organic supramolecules has been of great interest in recent years. In particular, the complexation of 2-pyridyl-1,2,3-triazole chelating pockets with transition-metal ions has intensified due to the ease of synthesis of the ligands through the copper-catalyzed azide–alkyne cycloaddition (CuAAC) reaction 2]4+ lying on an inversion center, two BF4− anions in general positions, a dimethylformamide solvent molecule in a general position and a diethyl ether solvent molecule disordered about an inversion center  atoms of two pyridyltriazole moieties. The N—Pd—N chelating angles are N1—Pd1—N2 = 79.46 (8)° and N7—Pd1—N8 = 79.75 (8)°. The N(py)—Pd bonds  are slightly longer than N(trz)—Pd bonds , suggesting the triazolium N atom coordinates more strongly to the Pd center than the pyridyl N atom of the ligand. These values are typical for the reported mononuclear PdII complex of pyridyltriazole ligands  with an average interplanar distance of 3.364 Å, indicating π–π interaction between the molecules; this is shown along the 3CN)4]BF4  in acetonitrile (5 ml), o-xpt  in acetonitrile (5 ml) was added dropwise. The solution was stirred for 1 h at room temperature. The volatiles were removed in vacuo. The residue was washed with dichloromethane (2 ml), followed by methanol (2 ml), and dried under vacuum to give [Pd2(o-xpt)2](BF4)4  as a pale-yellow solid. Crystals suitable for X-ray analysis were obtained by slow vapor diffusion of diethyl ether into a dimethylformamide (DMF) solution of the complex at room temperature. Our attempts to obtain a clean 1H NMR spectrum in DMSO-d6 were not successful, possibly due to the labile nature of the complex in solution. High resolution ESI–MS analysis showed a monocationic signal at m/z 1255.1478 {calculated 1255.1540 for [Pd2(o-xpt)2(BF4)4]+}.To a stirred solution of [Pd values were assigned as 1.2Ueq for the attached atom (1.5 for methyl). A torsional parameter was refined for each methyl group, except for those of the diethyl ether molecule, which were staggered with respect to CH2. The diethyl ether solvent molecule is disordered about an inversion center with two half-populated sites. A number of distance and displacement parameter restraints were necessary to model the disorder.Crystal data, data collection and structure refinement details are summarized in Table 110.1107/S2414314623003620/xu4050sup1.cifCrystal structure: contains datablock(s) I, glonal. DOI: 10.1107/S2414314623003620/xu4050Isup2.hklStructure factors: contains datablock(s) I. DOI: 2257865CCDC reference:  crystallographic information; 3D view; checkCIF reportAdditional supporting information:


The dihedral angles between the A and B rings and their attached phenyl rings are 49.15 (8) and 80.78 (8)°, respectively. In the extended structure, the molecules are linked by weak C—H⋯O and C—H⋯π bonds and π–π stacking interactions, which variously generate C(11) chains and Edis at −428.6 kJ mol−1 being the major contributor.In the title compound, C 38H28O6, the dihedral angles between the naphthalene ring system and its pendant benzyloxy rings A and B are 88.05 (7) and 80.84 (7)°, respectively. The dihedral angles between the A and B rings and their attached phenyl rings are 49.15 (8) and 80.78 (8)°, respectively. In the extended structure, the molecules are linked by weak C—H⋯O and C—H⋯π hydrogen bonds, and π–π stacking interactions, which variously generate C(11) chains and R22(12) loops as part of a three-dimensional network. The Hirshfeld surface  and intermolecular interaction energies are reported, with dispersion (Edis = −428.6 kJ mol−1) being the major contributor.In the title compound, C A variety of banana-shaped, bow-shaped or bent-core ferroelectric liquid crystals were developed by incorporating a benzene ring as a rigid core  and its pendant C26–C31 (A) and C12–C17 (B) benzyloxy rings are 88.05 (7) and 80.84 (7)°, respectively. The dihedral angles between the A and B rings and their attached C33–C38 and C19–C24 phenyl rings are 49.15 (8) and 80.78 (8)°, respectively. Key torsion angles include C1—O4—C25—C26 [−160.98 (13)°], C28—O2—C32—C33 [−172.04 (14)°], C10—O1—C11—C12 [−168.94 (14)°] and C14—O3—C18—C19 [172.84 (14)°]. Otherwise, the geometrical data for the title compound may be regarded as normal.The title compound crystallizes with one molecule in the asymmetric unit Fig. 1 in the s3.C(11) chain (arising from the C21—H21⋯O2ii hydrogen bond), which runs along [010], and centrosymmetric i hydrogen bond) between the molecules as shown in Fig. 2In the crystal, the molecules are linked by numerous C—H⋯O and C—H⋯π interactions Table 1. Promine4.et al., 2007CrystalExplorer17  are electrostatic (Eele), polarization (Epol), dispersion (Edisp) and exchange–repulsion (Erep), and the cylinder-shaped energy frameworks represent the relative strengths of the interaction energies in individual directions, as well as the topologies of pairwise intermolecular interaction energies within the crystal  in DCM (20 ml). Filtration was used to remove the precipitated N,N-dicyclohexylurea and the solvent was evaporated. To obtain the pure product, the solid residue was purified using column chromatography on silica gel with DCM as an eluent, followed by recrystallization from ethyl alcohol solution.Under an inert atmosphere, 1,2-dihydroxynaphthalene (1.00 mmol), a catalytic amount of 4-dimethylaminopyridine and 3-benzyloxybenzoic acid (2.00 mmol) were dissolved in 50 ml of dry dichloromethane (DCM). The above mixture was stirred for 2 h at room temperature with a solution of 7.Uiso(H) = 1.2Ueq(C).Crystal data, data collection and structure refinement details are summarized in Table 210.1107/S2056989023005571/hb8069sup1.cifCrystal structure: contains datablock(s) I, global. DOI: 10.1107/S2056989023005571/hb8069Isup4.hklStructure factors: contains datablock(s) I. DOI: Click here for additional data file.10.1107/S2056989023005571/hb8069sup3.docxSupplementary Figures showing C--H...pi interactions. DOI: 2271880CCDC reference:  crystallographic information; 3D view; checkCIF reportAdditional supporting information:


Compounds (I)–(III) are constructed from vertex-sharing ZnO3N tetrahedra (the organic molecule acting as a ligand) and HPO3 pseudo pyramids in a 1:1 ratio to generate the same motif of infinite 4-ring ‘ladder’ chains propagating in the [010], [101] and [100] directions, respectively, whereas (IV) consists of (010) layers of vertex-sharing ZnO4 and HPO3 units in a 3:4 ratio with the protonated organic molecule acting as a template. When an excess of HCl is used in the synthesis, the simple hydrated molecular salt, bis(2-amino-3-methylpyridinium) tetrachlorozincate monohydrate, (C6H8N2)2[ZnCl4]·H2O, (V), arises. Compounds (I)–(V) feature extensive networks of hydrogen bonds, both classical  and non-classical  in nature, which help to consolidate the extended structures.The syntheses and crystal structures of four hydrothermally prepared organo–zinc phosphites,  Each O atom is bonded to one Zn and one P atom [mean Zn—O—P = 130.2°], thus there are no ‘dangling’ The asymmetric unit of (I)C2/c, consists of two Zn2+ ions, two [HPO3]2– hydrogen phosphite anions, and two C6H7N2 2-amino-4-methylpyridine molecules acting as ligands, i.e., Z′ = 2. Unlike (I)3N tetrahedra and [HPO3]2– dianions – and the major structural features of (II)4′ = 0.97; comparable data for Zn2 = 1.940 Å, 100.10 (5)–121.28 (5)° and 0.91, respectively; mean P1—O = 1.525 Å; spread of O—P1—O bond angles = 110.64 (6)–113.78 (6)°; P1 displacement from its attached O atoms = 0.4278 (7) Å; comparable data for P2 = 1.520 Å, 111.44 (6)–113.14 (7)° and −0.4269 (8) Å, respectively. All six O atoms are bridging between Zn and P atoms with a mean bond angle of 131.3° [range = 123.26 (6)–144.09 (8)°]. The extended structure of (II)The asymmetric unit of (II)P212121 with a well-defined absolute structure and its asymmetric unit  and 115.09 (8)°, respectively, τ4′ = 0.96] and [HPO3]2– units . The three O atoms bridge adjacent zinc and phosphorus atoms with a mean Zn—O—P bond angle of 126.7°. For the extended structure of (III)Compound (III)it Fig. 3 consistsP4 tetrahedra and four [HPO32–] pseudo pyramids as well as two protonated 2-amino-4-methylpyridinium cations, which therefore act as templates rather than ligands; a water molecule of crystallization (O13) completes the structure. Geometrical data for the zinc polyhedra are as follows: mean Zn1—O = 1.941 Å, spread of bond angles = 100.42 (8)–122.18 (9)°, τ4’ = 0.90; equivalent data for Zn2: 1.936 Å, 98.33 (8)–115.30 (9)° and 0.98, respectively; equivalent data for Zn3: 1.945 Å, 99.70 (8)–117.10 (8)° and 0.96, respectively. The four [HPO3]2– anions adopt their normal geometries: mean P1—O = 1.519 Å, minimum and maximum O—P1—O = 111.00 (11) and 112.70 (11)°, respectively, deviation of P1 from its attached O atoms = 0.4498 (13) Å; equivalent data for P2: 1.522 Å, 110.08 (10)°, 115.33 (11)° and −0.4122 (12) Å, respectively; equivalent data for P3: 1.516 Å, 110.19 (11)°, 114.49 (12)° and −0.4123 (13) Å, respectively; equivalent data for P4: 1.516 Å, 112.68 (12)°, 114.13 (12)° and 0.3903 (13) Å, respectively. The twelve unique O atoms all bridge Zn and P atoms , maximum = 146.90 (13), spread = 21.5°). For the extended structure of (IV)In (IV)it Fig. 4 reveals P6H8N2+ cations protonated at their pyridine N atoms, a [ZnCl4]2– anion and a water molecule of crystallisation. The tetrachlorozincate ion has a mean Zn—Cl separation of 2.2704 Å [range = 2.2536 (13)–2.2867 (13) Å] and smallest and largest Cl—Zn—Cl bond angles of 104.48 (5) and 113.75 (5)°, respectively. The synthetic intent here was to lower the pH with HCl and establish if a dihydrogen phosphite (H2PO3−) anion containing a terminal P—OH moiety could be incorporated into the structure 3.3N and HPO3 polyhedra are linked by Zn—O—P bonds into [010] polyhedral 4-ring (two Zn and two P nodes) ‘ladder’ chains in which the zinc and phosphorus nodes strictly alternate  to adjacent P atoms (and a fourth bond to the organic species) and that the P atom forms three links to zinc atoms (and a fourth P—H vertex), the 1:1 Zn:P stoichiometry is to be expected and hence no charge compensating, protonated template is needed. In (II)Z′ = 2, every other 4-ring is generated by inversion symmetry and translation in the [101] direction leads to the extended array. In (III)motif is again apparent te Fig. 6: the chant Fig. 6, althougns Fig. 6.4 and HPO3 polyhedra sharing corners. One way to visualize this rather complex arrangement  is in terms of contorted chains of 4-rings featuring atoms Zn1, Zn2, Zn3, P2, P3 and P4 as the nodes propagating in the [001] direction. One out of every three 4-rings in a chain is generated by inversion symmetry. These chains are cross-linked in the a-axis direction by the P1-centred hydrogen phosphite groups to form the (010) layers, which encapsulate 8-ring voids built up from four Zn and four P nodes although there is no suggestion of ‘zeolitic’ porosity. So far as stoichiometry is concerned, in this case the zinc nodes forming four bonds  to nearby phosphorus atoms and the P nodes forming three bonds to Zn atoms leads to the 3:4 ratio of zinc and phosphorus, which is the proportion most commonly seen in this family of phases 4] unit, hence the two protonated template molecules. The template cations and water molecules of crystallisation occupy the inter-layer regions.The extended structure of (IV)Various classical  and non-classical (C—H⋯O and C—H⋯Cl) hydrogen bonds occur in these structures. As is normal, the hydrogen phosphite P—H unit does not participate in hydrogen bonding  and inter-chain (via H1N and H3N) links, with the latter serving to cross-link the [101] chains into a three-dimensional network (Table 2B⋯O2 bond (Table 3A) of the amine grouping does not participate in a hydrogen bond, the closest acceptor O atom being some 2.77 Å distant. There are no significant π–π stacking interactions in (III)In (II)k Table 2. The arok Table 2, the sind Table 3 cross-livia two O—H⋯O hydrogen bonds. The N—H⋯O hydrogen bonds originating from the protonated pyridine N atoms and the –NH2 groups of the organic species all link to the same sheet for each template cation, i.e., there are no inter-sheet hydrogen bonds associated with the templates. Significant aromatic π–π stacking interactions occur between centrosymmetric pairs of each template cation, as indicated by the centroid–centroid separation of 3.6167 (15) Å (slippage = 1.196 Å) for the C1 species and 3.4695 (17) Å (0.146 Å) for the C7 cation.In (IV)w (w = water) and O—H⋯Cl interactions to generate [001] chains; within these chains, centrosymmetric assemblages of two C6H8N2+ cations, two [ZnCl4]2– anions and two water molecules are apparent t Fig. 12.4.et al., 20165H6N2·Zn(HPO3)]n, catena-[(μ3-hydrogenphosphito)(2-aminopyridine)zinc] (CSD refcode LUZYOU)  layers of 4- and 8-rings.A survey of the Cambridge Structural Database (Groom 3 units in (I)motif in the wider ZnPO phase space: two other examples with very different ligating molecules to those in (I)–(III) are [C4H8N2O3·Zn(HPO3)]n (C4H8N2O3 = l-asparagine) ]n  –(III).The fact that the N-bonded zinc ions and HPO6H8N2 organic molecule acts as a ligand (a Zn—N bond and a 1:1 Zn:P ratio) and (IV)6H9N2+ template (N—H⋯O hydrogen bonds and a 3:4 Zn:P ratio) arose from similar syntheses, with the only difference being the source of zinc ions . Assuming that hydrothermal synthesis is not just an impenetrable ‘black box’ 5.3PO3 and 1.14 g of 2-amino-3-methylpyridine (Zn:P:template ratio ≃ 1:1:1), which were placed in a 50 ml polypropylene bottle with 20 ml of water and shaken well to result in a white slurry. The bottle was placed in an 353 K oven for 24 h and then removed and allowed to cool to room temperature. The solids were recovered by vacuum filtration to result in a mass of needle-like transparent crystals. IR: 2383 cm−1 (P—H stretch). Increasing the heating time to one week led to the same product, with a slight improvement in crystallinity, as indicated by sharper peaks in its IR spectrum and X-ray powder diffraction pattern.Compound (I)3PO3 and 1.10 g of 2-amino-4-methylpyridine (Zn:P:template ratio ≃ 1:1:1); otherwise following the same procedure as for (I)−1 (P—H stretch). Two peaks may arise because of the two different P—H groups in the asymmetric unit 2, 0.86 g of H3PO3 and 1.09 g of 2-amino-5-methylpyridine (Zn:P:template ratio ≃ 1:1:1) and 20 ml of water were placed in a 50 ml polypropylene bottle and heated to 353 K for three days. Upon cooling, the product consisted of a mass of colourless blocks. IR: 2406 cm−1 (P—H stretch).To prepare compound (III)2, 0.77 g of H3PO3 and 1.03 g of 2-amino-4-methylpyridine (Zn:P:template ratio ≃ 1:1:1) and 20 ml of water. These components were placed in a 50-ml polypropylene bottle and heated to 353 K for 24 h. Upon cooling, the product consisted of a mass of colourless blocks. IR: 3000–3600 (broad) (O—H stretch), 2391, 2381 cm−1 (P—H stretch). The same product arises if the mixture is heated for one week.Compound (IV)M HCl.Compound (V)6.Uiso(H) = 1.2Ueq(N or O). The phosphite H atoms were geometrically placed (P—H = 1.32 Å) and refined as riding atoms with Uiso(H) = 1.2Ueq(P). All the C-bound H atoms were located geometrically (C—H = 0.95–0.98 Å) and refined as riding atoms with Uiso(H) = 1.2Ueq(C) or 1.5Ueq(methyl C). The methyl groups were allowed to rotate, but not to tip, to best fit the electron density. Two peaks greater than 1 e Å−3 were found in the final difference map for (IV)Crystal data, data collection and structure refinement details are summarized in Table 610.1107/S2056989023002062/pk2681sup1.cifCrystal structure: contains datablock(s) I, II, III, IV, V, global. DOI: 10.1107/S2056989023002062/pk2681Isup2.hklStructure factors: contains datablock(s) I. DOI: 10.1107/S2056989023002062/pk2681IIsup3.hklStructure factors: contains datablock(s) II. DOI: 10.1107/S2056989023002062/pk2681IIIsup4.hklStructure factors: contains datablock(s) III. DOI: 10.1107/S2056989023002062/pk2681IVsup5.hklStructure factors: contains datablock(s) IV. DOI: 10.1107/S2056989023002062/pk2681Vsup6.hklStructure factors: contains datablock(s) V. DOI: 2246111, 2246110, 2246109, 2246108, 2246107CCDC references:  crystallographic information; 3D view; checkCIF reportAdditional supporting information: