This process takes place in the presenceof a halogenating agent and under Brønsted acid catalysis andproceeds via a transannular amidohalogenation, followed by elimination.The reaction has been found to be wide in scope, enabling the formationof a variety of tricyclic products in good overall yield, regardlessof the substitution pattern in the initial lactam substrate. The reactionhas also been applied to the total synthesis of a reported topoisomeraseI inhibitor and to the formal synthesis of rosettacin. Further extensionof this methodology allows the preparation of 10-iodopyrroloisoquinolinones by using an excess of halogenating agentand these compounds can be further manipulated through standard Suzukicoupling chemistry into a variety of 10-aryl-substituted pyrroloisoquinolinones.A transannular approach has been developed for the constructionof pyrrolo[1,2- Moreover, isoindoloisoquinolin-5(7H)-one, which also sharesthe same pyrroloisoquinoline motif has shown to exhibit similar topoisomeraseI activity to camptothecin.3 Despite thispromising activity, the use of aromathecin derivatives in clinicaltrials still suffers from poor solubility and dose-limiting toxicity,4 and consequently, there is still a constant needfor the development of effective protocols that enable the constructionof this scaffold with a special interest on those strategies thatprovide a diversity-oriented synthesis approach.The 2,3-dihydropyrroloisoquinolin-5(1H)-one scaffold involve the generation of the B and/or Crings through cyclization or cyclocondensation reactions. In all thesecases, the retrosynthetic design involves disconnections of the C10–C10a,5 C3–N,6 C5–N,7 C5–C5a,8 C9a–C10,9 or C1–C10a10 -one core.12 As an alternative, we propose herein an unconventionaland yet unexplored approach to this scaffold that comprises the generationof the C10a–N bond via formal oxidativetransannular amido functionalization of a benzo-fused medium-sizedunsaturated lactam, as shown in b]isoquinolin-5(1H)-one upon the in situelimination process. Transannular reactions, in which the two reactingsites are located within the cyclic structure of the starting material,have been widely used as a key step in the design of efficient synthesesof rather complex molecular scaffolds,13 including several examples of elegant total syntheses of naturalproducts.14 In most cases, the transannularapproach has demonstrated its performance as a key strategic decisionthat reduces the number of steps involved in the synthetic route and/orenables highly efficient stereocontrol due to the limited degree ofconformational freedom associated with the medium- or large-size cyclicstarting material, the latter effect also being exploited for thedevelopment of several enantioselective variants.15 Specifically, there are several precedents of highly effectivetransannular aminohalogenation or amidohalogenation reactions employedfor the synthesis of bicyclic nitrogen-containing heterocycles suchas pyrrolizidines,16 indolizidines,17 and related derivatives.18 Despite all progress in this field, the transannular approachhas not been already employed for the construction of the isoquinolinecore.In general,the synthetic routes described up to date for the formationof the 2,3-dihydropyrroloisoquinolin-5(1H)-ones 9a–d in excellent yields -ones, which are well-known topoisomerase I inhibitors-configuredlactams 12a–h after N-deprotectionunder standard conditions.20 This approachfor the formation of the medium-sized lactam moiety was especiallysuccessful when arylamide 10-incorporated electron-donatingsubstituents on any of the aryl moieties, providing significantlylower yields in the intramolecular Heck reactions in the cases inwhich fluorine or chlorine atoms were installed. Once the key lactams 12a–h had been prepared, these were submitted to thetransannular amidobromination/elimination process under the standardconditions. However, in an initial attempt, the reaction providedthe amidohalogenated product in which elimination had not taken place.This situation was solved by adding 1 equiv of a Brønsted basesuch as DBU together with NaI; the latter was required to promotea Finkelstein-type process that inverted the relative configurationof the amidohalogenation product to facilitate the elimination reactionthrough the E2 process.21 As it can beseen in b]isoquinolin-5(7H)-one 13a, which had been reported to be a highly active topoisomerase I inhibitorwas formed in excellent yield from lactam 12a under theseconditions. In addition, the other lactams 12b–h prepared were also converted into isoindoloisoquinolin-5(7H)-ones 13b–h through this transannular process in excellent yields regardlessof the substitution patterns in any of the aryl moieties.On the other hand, we also faced the possibilityof using thisapproach to the synthesis of isoindoloisoquinolin-5(1H)-ones using a transannular approach as the key transformationfor the construction of the tricyclic heterocyclic scaffold. Thiscrucial transannular reaction consists of a Brønsted acid-catalyzedamidohalogenationprocess, followed by elimination, and provides the target productsin good yields regardless of the substitution pattern in the cycliclactam. Moreover, when the reaction is carried out in the presenceof an excess of NIS as the electrophilic halogen source, a secondhalogenation reaction took place on the obtained adducts, furnishingthe corresponding iodinated derivatives in good yields, with thesecompounds having a convenient functionality to be further functionalizedthrough Suzuki coupling chemistry. The protocol described herein hasbeen successfully applied to the preparation of the topoisomeraseI inhibitor isoindoloisoquinolin-5(7H)-one and several other related compounds, also includingthe formal total synthesis of rosettacin.In conclusion, we have demonstrated that nine-memberedenelactamscan be used as useful starting materials for the preparation of differentlysubstituted pyrrolo+ calcd for [C13H14ClO2]+, 237.0677; found,237.0689 for compound: 1a.In an oven-dried, two-necked round-bottomflask equipped with a condenser and stir bar with methyl 2-bromobenzoate, PdCl2 balloon with methyl 2-(5-chloropent-1-yn-1-yl)benzoate 1a  in EtOAc (127 mL) and quinoline , Pd on CaCO3  was added. The air flask was evacuated under vacuum and backfilledwith hydrogen three times, and the reaction mixture was allowed tostir at room temperature under a hydrogen atmosphere until full consumption of the starting material, 1 h as judged byTLC. The reaction mixture was filtered through a plug of Celite, andthe filtrate was concentrated under vacuum. The crude was purifiedby silica gel flash chromatography (petroleum ether/EtOAc 19:1) obtaining 2a  as a yellow oil. Rf: 0.62 (petroleum ether/EtOAc). 1H NMR : δ 7.95 , 7.48 , 7.36–7.28 , 6.92 , 5.67 ,3.87 , 3.49 , 2.27 , 1.92–1.78 . 13C {1H} NMR: δ 167.6 (C), 138.9 (C), 131.8 (CH),130.8 (CH), 130.6 (CH), 130.5 (CH), 129.8 (CH), 129.4 (C), 127.0 (CH),52.0 (CH3), 44.5 (CH2), 32.7 (CH2), 25.6 (CH2). IR : 1720(C=O). MS (EI) m/z (%): 238, 161 , 128 , 115 . HRMS(ESI) m/z: [M + H]+ calcdfor [C13H16ClO2]+, 239.0833;found, 239.0838 for compound: 2a.To a two-necked round-bottom flask equippedwith a stirring bar and HZ)-2-(2-(5-chloropent-1-en-1-yl)benzoate) 2a , Cs2CO3 , phthalimide , and KI  in DMF (70 mL) under an Ar atmosphere was heated using a heatingplate at 100 °C for 2 h. Then, the reaction mixture was let tocool down to room temperature and water (100 mL) and EtOAc (50 mL)was added. The aqueous layer was extracted with EtOAc (3 × 25mL), washed with H2O (2 × 75 mL), dried with Na2SO4, filtered and concentrated under vacuum. Thecrude was purified by silica gel flash chromatography (petroleum ether/EtOAc9:1 to petroleum ether/EtOAc 8:2) in order to obtain 3a  as a white solid. Rf: 0.5 (petroleum ether/EtOAc 8:2). mp 69–71°C. 1H NMR : δ 7.89 , 7.80 , 7.68 , 7.40, 7.29–7.19 ,6.88 , 5.72 , 3.85 , 3.66–3.58 , 2.23–2.09, 1.83–1.69 . 13C {1H}NMR : δ 168.5 (2xC), 167.8 (C), 138.8(C), 134.0 (2xCH), 132.3 (2× C), 131.7 (CH), 130.7 (CH), 130.6(CH), 130.4 (CH), 130.0 (CH), 129.5 (C), 126.9 (CH), 123.3 (2×CH), 52.1 (CH3), 37.8 (CH2), 28.7 (CH2), 25.8 (CH2). IR : 1771(O=C–N–C=O), 1708 (C=O). MS (EI) m/z (%): 317 , 170 . HRMS (ESI) m/z: [M + H]+ calcd for [C21H20NO4]+, 350.1387; found,350.1390 for compound: 3a.An oven-dried two-necked round-bottom flaskprovided with a condenser and a magnetic bar with a suspension ofmethyl (Z)-2-pent-1-en-1-yl)benzoate 3a  and hydrazine  in EtOH (51 mL). The reaction mixture was heated usinga heating plate at 50 °C for 2 h. Then, the reaction mixturewas let to cool down to room temperature and was filtered througha plug of Celite The filtrate was concentrated under reduced pressure,and 1 M HCl (50mL) was added; the aqueous layer was washed with EtOAc(3 × 25 mL), basified with 4 M NaOH to pH = 9, extracted withCH2Cl2 (4 × 25 mL), dried with Na2SO4, filtered, and concentrated under vacuum to obtain 4a  as a yellow oil. Rf: 0.2 (MeOH). 1H NMR : δ 7.96 , 7.49 , 7.37–7.28, 6.90 , 5.74 , 3.90 , 2.68 , 2.19 , 1.57 , 1.36 . 13C {1H} NMR : δ 168.1 (C), 139.2 (C), 132.0 (CH),131.4 (CH), 131.1 (CH), 130.8 (CH), 129.9 (C), 129.7 (CH), 127.2 (CH),52.4 (CH3), 41.3 (CH2), 32.4 (CH2), 26.0 (CH2). IR : 1719(C=O), 1636 (N–H). MS (EI) m/z (%): 219 , 202 , 161 , 115 . HRMS (ESI) m/z: [M + H]+ calcd for [C13H18NO2]+, 220.1332; found, 220.1340 for compound: 4a.A round-bottom flask equipped with a magneticbar was provided with (Z)-2-(5-aminopent-1-en-1-yl)benzoate 4a  in dry THF (115 mL) under an Ar atmosphere,LiHMDS 1 M in THF  was added dropwise at 0 °C.The reaction mixture was stirred at room temperature and monitoredby TLC. Once the starting material (SM) was consumed, MeOH (1 mL)was added, and the solvent was removed under vacuum. The residue wasdissolved in CH2Cl2 and filtrated through Celite.The filtrate was concentrated under vacuum and purified by silicagel flash chromatography (petroleum ether/EtOAc 8:2 to petroleum ether/EtOAc7:3) to obtain 5a  as a whitesolid. Rf: 0.4 (petroleumether/EtOAc 7:3). mp 183–186 °C. 1H NMR : δ 7.42–7.27 , 7.17 , 6.62 , 6.30 , 6.00 , 3.41–2.99 , 2.30–1.78 , 1.51 . 13C {1H}NMR : δ 174.0 (C), 137.1 (C), 136.2(C), 134.3 (CH), 128.7 (CH), 128.7 (CH), 128.3 (CH), 127.3 (CH), 125.2(CH), 45.4 (CH2), 29.3 (CH2), 28.9 (CH2). IR : 3287 (N–H), 1646 (N–C=O).MS (EI) m/z (%): 187 , 158 . HRMS (ESI) m/z: [M + H]+ calcd for [C13H17NO]+, 202.1232;found, 202.1235 for compound: 5a.To an oven-dried two-necked round-bottom flask providedwith a magnetic bar with methyl (Z)-2,3,4,5-tetrahydro-1H-benzo[c]azonin-1-one 5a , a stocksolution of diphenylphosphoric acid  in dry dichloromethane(220 μL) was added at 25 °C, followed by the addition ofNBS . The reaction mixture was subjected to TLC,and when all starting material was consumed , thesolvent was evaporated under vacuum, and Et2O (1 mL) anda standard. aq solution of NaHCO3 (1 mL) were added. Thelayers were separated, and the aqueous phase was extracted with Et2O (3 × 1 mL). All organic layers were washed with water(3 × 1 mL) and brine (1 mL), dried with Na2SO4, filtered, and concentrated under vacuum. The crude was purifiedby silica gel flash chromatography (CH2Cl2 toCH2Cl2/MeOH 99:1) obtaining 6a  as a white solid. Rf: 0.3 (CH2Cl2/MeOH 99:1). mp102–105 °C. 1H NMR : δ 8.31 , 7.51, 7.38 , 7.32 ,6.34 , 4.15–4.06 , 3.02 , 2.20–2.04 . 13C {1H} NMR : δ 161.6 (C), 143.7(C), 138.1 (C), 131.9 (CH), 127.3 (CH), 125.5 (CH), 125.5 (CH), 124.7(C), 100.3 (CH), 47.9 (CH2), 31.3 (CH2), 22.0(CH2). IR : 2985 (C–H),1657 (N–C=O), 1624 (C=C). MS (EI) m/z (%): 184.1 . HRMS (ESI) m/z: [M + H]+ calcd for [C12H12NO]+, 186.0919; found, 186.0919for compound: 6a.To a reaction tube provided with a magnetic bar with (Z)-2,3,4,5-tetrahydro-1H-benzo[c]azonin-1-one 5a , astock solution of diphenylphosphoric acid  indry dichloromethane (290 μL) was added at 25 °C, followedby the addition of NIS . The reaction mixture wassubjected to TLC, and when all starting material was consumed , the solvent was evaporated under vacuum, and Et2O (1 mL) and a standard aq solution of Na2S2O3 (1 mL) were added; the mixture was stirred for 30 minat room temperature. Then, the layers were separated, and the aqueousphase was extracted with Et2O (3 × 1 mL). All organiclayers were washed with water (3 × 1 mL), brine (1 mL), driedwith Na2SO4, filtered, and concentrated undervacuum. The crude was purified by silica gel flash chromatography(CH2Cl2 to CH2Cl2/MeOH99:1) in order to obtain 7a as a pale-yellow solid. Rf: 0.3 (CH2Cl2/MeOH 99:1). mp 141–143°C. 1H NMR : δ 8.37, 7.76 , 7.69 , 7.47 , 4.36, 3.26 , 2.25 . 13C {1H} NMR : δ 160.9 (C),146.7 (C), 138.0 (C), 133.1 (CH), 129.3 (CH), 127.7 (CH), 126.6 (CH),124.9 (C), 68.7 (C), 50.2 (CH2), 37.5 (CH2),20.8 (CH2). IR : 1638 (N–C=O),1609 (C=C). MS (EI) m/z (%):311.0 . HRMS (ESI) m/z: [M + H]+ calcd for [C12H11INO]+, 311.9885; found, 311.9891 for compound: 7a.To a reaction tube provided with a magnetic bar with (b]isoquinolin-5(1H)-one 7a , phenyl boronicacid , Pd(PPh3)4 , and K2CO3 .Under an argon atmosphere, 1,4-dioxane (5 mL) and water (2 mL) wereadded, and the reaction mixture was subjected to vacuum and refilledwith argon three times. The reaction mixture was heated using a heatingplate up at 65 °C for 21 h. After completion ,the mixture was cooled to room temperature and diluted with EtOAc(10 mL). The mixture was filtered through a small pad of Celite. Afterward,the solvent was concentrated in vacuum. The resulting crude was purifiedby flash chromatography (petroleum ether/EtOAc 7:3 to 1:1) in orderto obtain 9a  as a yellow solid. Rf: 0.27 (petroleum ether/EtOAc1:1). mp 169–171 °C. 1H NMR : δ 8.48 ,7.59–7.34 , 7.36–7.23 , 4.28 , 2.93 ,2.15 . 13C {1H} NMR : δ 161.2 (C), 141.4 (C),138.2 (C), 136.5 (C), 132.0 (CH), 130.7 (2× CH), 128.8 (2×CH), 127.7 (CH), 127.5 (CH), 125.7 (CH), 125.1 (C), 124.4 (CH), 113.8(C), 48.6 (CH2), 31.2 (CH2), 22.0 (CH2). IR : 1647 (C=O st), 1622 (CArom–CArom st), 1598 (CArom–CArom st). MS (EI) m/z (%):261.0 , 260.1 (67), 165.0 (36), 163.0 (20), 76.9(26). HRMS (ESI) m/z: [M + Hc+ Calcd for [C18H16NO]+, 262.1226;found, 262.1237 for compound: 9a.An oven-dried 50 mL two-necked flask was charged with10-iodo-2,3-dihydropyrrolo+ calcd for [C16H15INO]+, 364.0193; found, 364.0197.In the second step, to an oven-dried 50 mL two-necked flask, N-(2-iodobenzyl)-2-vinylbenzamide  anddi-tert-butyl dicarbonate  underan Ar atmosphere were dissolved in dry CH2Cl2 (12 mL), and under stirring, DMAP  was added atroom temperature. The reaction mixture was allowed to stir at roomtemperature for 16 h. Then, it was concentrated under vacuum and purifiedby silica gel column chromatography FC (petroleum ether/EtOAc 9:1to 8:2) in order to obtain 10a  as a yellow solid. Rf: 0.47 (petroleum ether/EtOAc8:2). mp 75–76 °C. 1H NMR : δ 7.76 ,7.49 , 7.35–7.16 ,7.10 , 6.91–6.71, 5.65 , 5.28 , 4.97 , 0.99 . 13C {1H} NMR : δ172.1 (C), 152.3 (C), 139.5 (CH), 139.4 (C), 137.1 (C), 135.3 (C),133.8 (CH), 129.7 (CH), 128.7 (CH), 128.4 (CH), 127.3 (CH), 126.3(CH), 126.2 (CH), 125.8 (CH), 117.0 (CH2), 97.7 (C), 83.7(C), 53.3 (CH2), 27.3 (3× CH3). IR : 1733 (C=O), 1671 (N–C=O).MS (EI) m/z (%): 362.0 . HRMS (ESI) m/z: [M + Na]+ calcd for [C21H22INO3Na]+, 486.0537; found, 486.0538 for compound: 10a.In the first step, to an oven-dried 100mL two-necked flask provided with the corresponding methyl 2-vinylbenzoate and (2-iodophenyl)methanamine under an Ar atmosphere, LiHMDS in THF 1 M  wasadded dropwise at room temperature to the stirring mixture, and itwas allowed to stir for 16 h at this temperature. The solvent wasevaporated under vacuum and purified by silica gel column chromatography(petroleum ether/EtOAc 8:2) to obtain tert-butyl(2-iodobenzyl)(2-vinylbenzoyl)carbamate 10a  in dry DMF (12.8 mL) under an Ar atmosphere, sodiumacetate , triphenylphosphine ,and palladium acetate  were added. The reactionmixture was stirred at 100 °C using a heating plate for 72 h.Then, it was cooled down and water (20 mL) and EtOAc (20 mL) wereadded; the mixture was filtered through Celite, the filtrate was extractedwith EtOAc (4 × 20 mL), and the organic phase was washed withwater (3 × 5 mL) and brine (10 mL), dried with Na2SO4 anhydrous, filtered, and concentrated under vacuum.The crude was purified by silica gel column chromatography FC (petroleumether/EtOAc 9:1) in order to obtain 11a  as a yellow solid. Rf: 0.45 (petroleum ether/EtOAc 95:5). mp 74–77 °C. 1H NMR : δ 7.70 , 7.53–7.41 , 7.40–7.25, 6.59 , 6.51 , 5.41 , 4.63 , 1.00 . 13C {1H} NMR : δ177.8 (C), 151.4 (C), 140.0 (C) 137.8 (C), 137.7 (C), 136.3 (CH),133.0 (C), 130.1 (CH), 129.5 (CH), 128.3 (CH), 128.1 (CH), 128.0 (CH),127.9 (CH), 127.5 (CH), 127.3 (CH), 125.1 (CH), 82.0 (C), 52.0 (CH2), 27.5 (3× CH3). IR : 1722 (C=O), 1680 (N–C=O). MS (EI) m/z (%): 232.9 (M+ –Boc). HRMS (ESI) m/z: [M + Na]+ calcd for [C21H21NNaO3]+, 358.1414; found, 358.1420 for compound: 11a.To an oven-dried 250 mL flask provided with tert-Butyl(E)-5-oxo-5,7-dihydro-6H-dibenzoazonine-6-carboxylate 11a  was dissolved in CH2Cl2 (5 mL),and silica gel (230–400 mesh) (2.00 g) was added. The solventwas vacuumed, and the powdered solid obtained was irradiated in themicrowave oven in an open Erlenmeyer flask at 540 W. The reactionwas checked by TLC every 6 min until it was completed . The reaction mixture was deadsorbed by thoroughly washing thesilica gel with petroleum ether/EtOAC 1:1 with pressure in a column.The crude was purified by silica gel column chromatography (petroleumether/EtOAc 8:2 to 7:3) in order to obtain 12a  as a white solid. Rf: 0.39 (petroleum ether/EtOAc 9:1). mp 205–208°C. 1H NMR : δ 7.66–7.60, 7.44–7.28 , 7.28–7.17 , 7.17–7.11, 6.65 , 6.41 , 5.38 , 4.82 , 4.18 . 13C {1H} NMR : δ 176.0 (C), 139.1 (C), 138.4 (C), 138.3(C), 136.0 (CH), 132.6 (C), 130.4 (CH), 129.3 (CH), 129.0 (CH), 128.7(CH), 128.1 (CH), 128.0 (CH), 127.9 (CH), 127.3 (CH), 125.7 (CH),48.9 (CH2). IR : 3280 (N–H),1637 (N–C=O), 1521 (C=C). MS (EI) m/z (%): 235.1 . HRMS (ESI) m/z: [M + H]+ calcd for [C16H14NO]+, 236.1075; found, 236.1074for compound: 12a.E)-6,7-dihydro-5H-dibenzoazonin-5-one 12a , astock solution of diphenylphosphoric acid  indry dichloromethane (200 μL) was added at 25 °C, followedby the addition of NBS . The reaction mixture wassubjected to TLC (CH2Cl2/MeOH 98:2). Once thestarting material was consumed (5h), DBU and NaI  were added to the reaction mixture at25 °C. The new reaction mixture was allowed to stir and was subjectedto TLC until the intermediate was consumed (16 h). Then, a standardaq. solution of NH4Cl (1 mL) was added; the layers wereseparated, and the aqueous phase was extracted with CH2Cl2 (3 × 1 mL). All organic layers were washed withwater (3 × 1 mL) and brine (1 mL), dried with Na2SO4, filtered, and concentrated under vacuum. The crude was purifiedby silica gel flash chromatography (CH2Cl2 toCH2Cl2/MeOH 99:1) to obtain 13a  as a pale-yellow solid. Rf: 0.49 (petroleum ether/EtOAc 1:1).mp 186–188 °C. 1H NMR : δ 8.52–8.44 , 7.82–7.75 , 7.70–7.60, 7.58–7.52 , 7.53–7.41 , 7.01, 5.18 . 13C {1H} NMR : δ 161.2 (C), 142.2 (C), 138.0 (C), 137.7(C), 134.1 (C), 132.2 (CH), 129.9 (CH), 128.4 (CH), 127.5 (CH), 126.4(CH), 126.2 (CH), 124.8 (C), 123.5 (CH), 121.1 (CH), 98.1 (CH), 52.1(CH2). IR : 1660 (N–C=O),1627 (C=C). MS (EI) m/z (%):233.0 . HRMS (ESI) m/z: [M + H]+ calcd for [C16H12NO]+, 234.0919; found, 234.0922 for compound: 13a.To a reaction tube provided with a magnetic bar with (


Various strategies to overcome the MDR phenomenon have been developed, and one of the most attractive research directions is focused on the inhibition of MDR transporters, membrane proteins that extrude cytotoxic drugs from living cells. Here, we report the results of our studies on a series newly synthesized of 5-arylidenerhodanines and their ability to inhibit the ABCB1 efflux pump in mouse T-lymphoma cancer cells. In the series, compounds possessing a triphenylamine moiety and the carboxyl group in their structure were of particular interest. These amphiphilic compounds showed over 17-fold stronger efflux pump inhibitory effects than verapamil. The cytotoxic and antiproliferative effects of target rhodanines on T-lymphoma cells were also investigated. A putative binding mode for  Multidrug resistance (MDR) is a severe problem in the treatment of various diseases such as cancer, bacterial, fungal, and parasitic infections. One of the mechanisms of cancer drug resistance is associated with increased drug efflux from cells, mediated by the ATP-binding cassette (ABC) transporters . More thThe MDR-ABC transporter most studied in cancer is P-glycoprotein . A wide range of compounds with beneficial inhibitory activity toward P-gp have been reported, however, most clinical trials targeting P-gp inhibition have failed, mainly due to the toxicity of the compounds . Multipl3, 7, and 11 were determined by the X-ray diffraction method to compare molecular geometries and to search for differences that could be responsible for the observed biological activity/inactivity. Additionally, for the representative of the most active amphiphilic derivative, 11, the molecular docking studies into the P-gp structure have been performed. The selected physicochemical properties for the whole series of compounds have been estimated as well.Rhodanine-derived compounds have been reported to possess a wide spectrum of biological activities, e.g., antidiabetic, antibacterial, antifungal, antimalarial, antitubercular, antiviral and anticancer ,14,15,1610–12  sho sho43] sm/z, positive ionization mode. CID fragmentation was in the ion trap analyzer with the aid of helium gas. The collision energy was set to ca. 1 eV. The samples were introduced into the mass spectrometer in a CH3OH:CHCl3 1:1 solution with 0.1% HCOOH acidification. The 1H-NMR spectra and 13C-NMR spectra were recorded on a JEOL JNM-ECZR500 RS1 (ECZR version) at 500 and 126 MHz, respectively, and were reported in parts per million (ppm) using deuterated solvent for calibration (DMSO-d6). The coupling constants values (J) were reported in hertz (Hz).All reagents for syntheses were purchased from Merck KGaA  and used without further purification. Melting point (uncorrected) has been determined on the Boetius apparatus (Carl Zeiss Jena). The IR spectrum has been recorded with Jasco FT IR-670 Plus spectrophotometer  in the KBr disk (1 mg sample/400 mg KBr). The MS analyses were obtained on the AmaZon ETD mass spectrometer . Scan parameters: scan range 100–1000 Two mmol of rhodanine or the appropriate rhodanine-3-carboxyalkyl acid, 3 g of 4-Å molecular sieves, 20 mL of isopropyl alcohol, 2 mmol of the appropriate aldehyde, and 1.4 mL (5 eq) of triethylamine were placed in the round bottom flask. The reaction mixture was heated under reflux for 5 h. After heating, the hot solution was filtered. The filtrate was cooled, and 2M hydrochloric acid was added thereto until the solution was acidic. The obtained precipitate was filtered off and crystallized from glacial acetic acid.Five mmol of rhodanine, 7 g of 4Å molecular sieves, 30 mL of isopropyl alcohol, 5 mmol of the appropriate aldehyde, and 3.5 mL (5 eq) of triethylamine were placed in the round bottom flask. The reaction mixture was heated under reflux for 6 h. After heating, the hot solution was filtered. The filtrate was cooled, and 2 M hydrochloric acid was added thereto until the solution was acidic. The resulting mixture was extracted with two 25 mL portions of chloroform. The organic fractions were combined and washed with three 25 mL portions of water. The solution was dried over anhydrous magnesium sulfate, filtered, and chloroform was evaporated from the filtrate. The obtained precipitate was crystallized from isopropanol.1)5--2-thioxothiazolidin-4-one  cm−1: 1719.23 N-CO-C, 1705.73 C=O conj., 1642.09 C=C exo., 1173.47 C=S; 1H-NMR  δ 13.82 , 7.80 , 7.70 , 7.63 , 7.45 , 7.37 ; 13C-NMR  δ 196.0 (C=S), 169.9 (N-C=O), 142.6 (C4′-Ar), 139.3 (Ar-C1″), 132.5 C=CH-Ar, 131.7 (C1′), 129.6 , 128.9 (C4″), 128.0 , 127.4 , 125.8 (C=CH-Ar).Yield 85.2%, m.p. 239–241 °C, MS [M+1]2)2--4-oxo-2-thioxothiazolidin-3-yl)acetic acid  cm−1: 1735.62 N-CO-C, 1711.51 C=O conj., 1641.13 C=C exo., 1199.51 C=S; 1H-NMR  δ 13.42 , 7.89 , 7.83 , 7.73–7.70 , 7.46 , 7.39 , 4.71 ; 13C-NMR  δ 193.6 (C=S), 167.9 (HOOC-), 166.9 (N-C=O), 143.0 (C4′), 139.2 (Ar-C1″), 134.1 , 132.4 (C1′), 132.1 (C=CH-Ar), 129.7 , 129.0 (C4″), 128.1 , 127.4 , 122.0 (C=CH-Ar), 45.6 (HOOC-CH2-).Yield 64.0%, m.p. 257–258 °C, MS [M+1]3)2--4-oxo-2-thioxothiazolidin-3-yl)propionic acid  cm−1: 1737.55 N-CO-C, 1707.66 C=O conj., 1641.13 C=C exo., 1171.54 C=S; 1H-NMR  δ 12.51 , 7.81–7.79 , 7.70 , 7.66 , 7.45 , 7.38 , 4.19 , 2.60 ; 13C-NMR  δ 193.7 (C=S), 172.3 (HOOC-), 167.2 (N-C=O), 142.8 (C4′-Ar), 139.2 (Ar-C1″), 133.0 , 132.5 (C=CH-Ar), 131.9 (C1′), 129.7 , 128.9 (C4″), 128.1 , 127.4 , 122.7 (C=CH-Ar), 40.5 (HOOC-CH2-CH2-), 31.3 (HOOC-CH2-CH2-). Yield 97.2%, m.p. 253–255 °C, MS [M+1]4)2--4-oxo-2-thioxothiazolidin-3-yl)butyric acid  cm−1: 1736.58 N-CO-C, 1712.48 C=O conj., 1641.13 C=C exo., 1167.69 C=S; 1H-NMR  δ 12.11 , 7.82 , 7.78 , 7.71 , 7.67 , 7.46 , 7.38 , 4.04 , 2.27 , 1.89–1.83 ; 13C-NMR  δ 194.0 (C=S), 174.3 (HOOC-), 167.7 (N-C=O), 142.7 (C4′-Ar), 139.2 (Ar-C1″), 132.8 (C=CH-Ar), 132.6 (C1′), 131.9 , 129.7 , 128.9 (C4″), 128.1 , 127.4 , 122.8 (C=CH-Ar), 44.3 (HOOC-CH2-CH2-CH2-), 31.5 (HOOC-CH2-CH2-CH2-), 22.5 (HOOC-CH2-CH2-CH2-).Yield 57.3%, m.p. 197–198 °C, MS [M+1]5)5-(4-(benzyloxy)benzylidene)-2-thioxothiazolidin-4-one  δ 13.72 , 7.56 , 7.52 , 7.42 , 7.36 , 7.30 , 7.14 , 5.17 ; 13C-NMR  δ 196.1 (S=C-S), 170.1 (N-C=O), 160.9 (C4 Ar), 137.0 (C1′ Ar), 133.2 , 132.3 (=CH-Ar), 129.0 , 128.6 , 128.4 (C4′ Ar), 126.2 (C=CH-Ar), 123.0 (C1 Ar), 116.4 , 70.1 (O-CH2-Ar).Yield 74.8%, m.p. 232–234 °C, MS [M-1]ˉ 326, IR (1 mg/400 mg KBr) cm6) 7)2-(5-(4-(benzyloxy)benzylidene)-4-oxo-2-thioxothiazolidin-3-yl)propionic acid  cm−1: 1738.51 N-CO-C, 1711.51 C=O conj., 1642.09 C=C exo., 1260.25 O-CH2-, 1171.54 C=S; 1H-NMR  δ 12.47 , 7.72 , 7.56 , 7.44–7.42 , 7.38–7.35 , 7.32–7.29 , 7.16–7.14 , 5.16 , 4.18 , 2.61–2.57 ; 13C-NMR  δ 193.6 (S=C-S), 172.3 (HOOC-), 167.3 (N-C=O), 161.2 (C4 Ar), 136.9 (C1′ Ar), 133.7 , 133.5 (=CH-Ar), 129.0 , 128.6 (C4′ Ar), 128.4 , 126.2 (C=CH-Ar), 119.7 (C1 Ar), 116.5 , 70.1 (O-CH2-Ar), 40.5 (HOOC-CH2CH2), 31.3 (HOOC-CH2CH2).Yield 85.9%, m.p. 190–192 °C, MS [M+1]8)2-(5-(4-(benzyloxy)benzylidene)-4-oxo-2-thioxothiazolidin-3-yl)butyric acid  cm−1: 1738.50 N-CO-C, 1710.01 C=O conj., 1643.31 C=C exo., 1262.18 O-CH2-, 1173.44 C=S; 1H-NMR  δ 12.10 , 7.71 , 7.56 , 7.43 , 7.38–7.35 , 7.32–7.29 , 7.15 , 5.16 , 4.03 , 2.25 , 1.87–1.82 ; 13C-NMR  δ 194.0, 174.2 (HOOC-), 167.7 (N-C=O), 161.1 (C4 Ar), 136.9 (C1′ Ar), 133.5 , 133.4 (=CH-Ar), 129.0 , 128.6 (C4′ Ar), 128.4 , 126.3 (C=CH-Ar), 119.8 (C1 Ar), 116.4 , 70.1 (O-CH2-Ar), 44.2 (HOOC-CH2CH2CH2), 31.5 (HOOC-CH2CH2CH2), 22.5 (HOOC-CH2CH2CH2).Yield 42.3%, m.p. 165–167 °C, MS [M+1]9)5-(4-(diphenylamino)benzylidene)-2-thioxothiazolidin-4-one  cm−1: 1720.19 N-CO-C, 1682.59 C=O conj., 1643.05 C=C exo., 1174.44 C=S; 1H-NMR  δ 13.65 , 7.46 , 7.36 , 7.32 , 7.13 , 7.07 , 6.85 ; 13C-NMR  δ 195.8, 170.0, 150.2, 146.2, 132.9, 132.3, 130.4, 126.4, 125.6, 125.4, 121.8, 120.3.Yield 25.0%, m.p. 237–239 °C, MS [M+1]L5178Y mouse T-cell lymphoma cells (PAR)  were transfected with pHa MDR1/A retrovirus, as previously described by Cornwell et al. . The ABC® flow cytometer , measuring 10,000 individual cells in the given sample. The inhibition of the MDR efflux pump P-gp (ABCB1) was detected by the retention of rhodamine 123 by ABCB1 (P-glycoprotein) in MDR mouse T-lymphoma cells using flow cytometry according to the protocol published by the former study . The flu4 cells in 100 μL of medium and in the case of the antiproliferative assay, 6 × 103 cells in 100 μL medium were pipetted to each well, with the exception of the medium control wells. The plates were incubated at 37 °C for 24 h or 72 h, respectively; at the end of the incubation period, 20 μL of MTT  solution (from a 5 mg/mL stock) were added to each well as described previously , R1 = 0.0650, wR2 = 0.1074 .7: C20H17NO4S2, Mr = 399.46, triclinic, space group P-1, a = 11.1930(3) Å, b = 13.2410(4) Å, c = 13.9940(4) Å, α = 113.250(2)°, β = 93.208(2)°, γ = 101.598(2)°, V = 1845.5(1) Å3, Z = 4, T = 293(2)K, 14691 reflections collected, 8425 unique reflections (Rint = 0.0254), R1 = 0.0481, wR2 = 0.1149 [I > 2σ(I)], R1 = 0.0626, wR2 = 0.1223 .11: C22H20N2O3S2, Mr = 460.55, triclinic, space group P-1, a = 7.9420(2) Å, b = 9.5630(3) Å, c = 16.0670(4) Å, α = 77.790(2)°, β = 76.769(2)°, γ = 69.832(1)°, V = 1103.25(5) Å3, Z = 2, T = 100(2)K, 8969 reflections collected, 5028 unique reflections (Rint = 0.0279), R1 = 0.0358, wR2 = 0.0816 [I > 2σ(I)], R1 = 0.0508, wR2 = 0.0889 .www.ccdc.cam.ac.uk/data_request/cif (accessed on 22 May 2022).CCDC 2174182-2174184 contains the supplementary crystallographic data. These data can be obtained free of charge from The Cambridge Crystallographic Data Centre via The cryo-EM structure of the human P-gp bound to encequidar was retrieved from the PDB database (PDB code 7O9W). The homology model of the human P-gp was built by multiple comparative modeling using X-ray structures 4Q9H, 4XWK, and 4M1M of the murine P-gp as templates) . The seq11 and verapamil was performed in Schrödinger Suite [11, obtained in this work under crystallographic studies, was used as the initial conformation of the ligand. The protonation states of the 11 and verapamil were generated in the LigPrep module to give neutral or ionized structures that were further optimized by minimization of energy, using options of the MacroModel module  Flexible receptor docking of compound er Suite . Both pr11, the receptor grid box was defined as a large binding cavity limited by the transmembrane region of the hP-gp, reported for the P08183 sequence. For verapamil, the putative binding site was built around a centroid of P-gp residues found by Loo in the mutagenesis experiments to interact with this drug [11 and verapamil to individual conformations of hP-gp.An induced-fit docking study was performed according to the protocol implemented in Schrödinger Suite. In the case of compound his drug ,41. DuriThe graphic presentation of selected protein-ligand complexes with the highest docking scores was prepared in the PyMOL software, v. 2.5.2 .10–12 (substituted with the carboxyl group) exhibit greater potential than 9 (no carboxyl substituent) to inhibit the P-gp. The obtained results indicate derivatives of rhodanine as a new group of compounds for medicinal chemistry and drug design in the search for new inhibitors of efflux pump P-gp.The present work demonstrated the previously unknown biological activity of rhodanine derivatives and the influence of the carboxyl group in their structure on this activity. Importantly, 8, showing additive interaction. An emerging research strategy could be the application of efflux pump inhibitors (EPIs) and their use as adjuvant compounds or chemosensitizers to improve the outcome of antitumor therapy by co-administering them with chemotherapeutic agents. For this reason, the ability of the compounds to inhibit ABCB1 should be investigated; furthermore, their interaction with standard chemotherapeutic drugs should be evaluated. Based on our results, potent ABCB1 modulators have been found; however, their combination with doxorubicin was not synergistic. They cannot be applied in combination with doxorubicin except for compound


Eryngium cymosum F. Delaroche, a plant that is used in traditional Mexican medicine to treat type 2 diabetes. The chemical structures of the isolated compounds were elucidated using a variety of spectroscopic techniques, including 1D and 2D nuclear magnetic resonance (NMR) and mass spectrometry (MS). Chlorogenic acid (1), rosmarinic acid (2), caffeic acid (3), protocatechuic acid (4), kaempferol-3-O--β-d-glucopyranoside (5), and the new acylated flavonol glucoside quercetin-3-O--β-d-glucopyranoside (6) were isolated. This is the first report on the natural occurrence of quercetin-3-O--β-D-glucopyranoside (6). In addition, according to the HPLC profile obtained for the water extract (WE), chlorogenic acid (1) and rosmarinic acid (2) were identified as the main compounds, while kaempferol-3-O--β-d-glucopyranoside (5) were the main compound in the butanolic extract. We demonstrate the important role of compound 5 over the inhibition of G6Pase and FBPase. The isolated compounds may play an important role in the hypoglycemic effect of the extract and may act in a synergic way, but more experiments are needed to corroborate these findings.One undescribed acylated flavonol glucoside and five known compounds were isolated from the aerial parts of  Eryngium genus comprises approximately 250 species that are widely distributed worldwide  and is the largest and most complex genus in the Apiaceae family −, which would correspond to the expected mass of compound m/z 756 minus one molecule of water (6 showed the presence of OH (3254 cm−1), C-H (2924 and 2854 cm−1), C=O (1625 cm−1), and the phenyl rings (1571 and 1485 cm−1) , 6.85 , and 7.56  assignable to H-6′, H-5′, and H-2′, respectively, which, according to the HSQC experiment  and H-8  protons that are in the A ring of flavonol. Furthermore, in the 1H-NMR spectrum , which corresponds, according to the HSQC experiment  of the anomeric proton H-1″ was indicative of the β-form glucose.Compound of water . The inf85 cm−1) . The 1H- , 6.44 (J = 15.9 Hz), 7.42 (J = 15.9 Hz), and 6.10 (J = 15.9 Hz) with a characteristic coupling constant of trans-olefinic protons and the COSY correlation are of the protons that resonate at δH 7.73–6.44 and 7.42–6.10  and 6.83  for the first system, and 7.33  with 6.83  for the second system  of the sugar. The other cinnamic molecule present in compound 6 is bound, according to the HMBC experiment, to the C-6″ (δC 64.2) of the sugar, observing a correlation of the carbonyl group that resonates at δC 168.7 with the H-6a″  and H-6b″  protons -β-d-glucopyranoside , rosmarinic acid (2), and caffeic acid (4) were identified in the extracts by coelution with the standards in HPLC and by comparing retention time and UV spectrum data of the peaks that were identified in the chromatographic profiles; in comparison, Compound 5 was identified by being coeluted with the extracts and by comparing UV spectrum data and retention time of the peak in the chromatographic profiles. Because Compounds 3 and 6 were not abundant in the extracts, there was a lack of standards and the samples rapidly decomposed; these compounds were not identified in the extracts. The two chromatograms were qualitatively different; however, the main components of the WE  were also observed in BuS , but in different proportions, and 5 was the major compound in this extract  showed a modest inhibition over G6Pase and no inhibition over FBpase, while kaempferol-3-O--β-d-glucopyranoside (5) showed an important inhibition over the two enzymes, suggesting that this compound could be involved in the hypoglycemic effect of the plant.The results of the inhibition assays of glucose-6-phosphatase (G6Pase) and fructose-1,6-bisphosphatase (FBPase) are shown in Eryngium cymosum, a plant commonly used in Tlanchinol, Hidalgo, Province of Mexico, for the management of T2D mellitus.In this study, we described the phytochemical composition of the aerial parts of Eryngium. Compounds 1, 2, and 4 have been identified in species such as E. alpinum, E. planum, and E. bourgatii −.Amorphous powder; 1H NMR  δ 7.23 , 6.82 , 6.75 , 6.74 , 6.68 , 6.66 , 6.59 , 5.97 , 5.06 , 2.96 , 2.91 ; 13C NMR  δ 173.9 (C-9′), 168.2 (C-9), 147.0 (C-3), 146.8 (C-7), 143.9 (C-4), 143.7 (C-3′), 142.8 (C-4′), 128.4 (C-1′), 126.5 (C-1), 122.7 (C-6), 121.9 (C-6′), 117.1 (C-2′), 116.0 (C-5′), 115.8 (C-5), 115.1 (C-2), 113.0 (C-8), 73.5 (C-8′), 36.0 (C-7′); ESIMS (negative-ion mode) m/z 358.5 [M − H]−.Amorphous powder; 1H NMR  δ 7.53 , 7.47 , 6.89 . 13C NMR  δ 166.55 (C-7), 149.77 (C-4), 144.63 (C-3), 122.71(C-6), 122.23 (C-1), 116.55 (C-2), 114.79 (C-5).Amorphous powder; 1H NMR  δ 7.54 , 7.16 , 7.04 , 6.87 , 6.26 . 13C NMR  δ 166.55 (C-9), 147.74 (C-4), 145.39 (C-3), 144.96 (C-7), 126.80 (C-1), 121.54 (C-6), 115.46 (C-5), 114.91 (C-8), 114.26 (C-2).Amorphous powder; −1; 1H NMR  δ 12.74 , 8.11 , 7.73 , 7.59 , 7.46 , 7.44 , 6.99 , 6.91 , 6.90 , 6.45 , 6.44 , 6.21 , 6.19 , 5.92 , 5.16 , 4.42 , 4.22 , 3.84 , 3.75–3.64 , 3.56. 13C NMR  δ 177.79 (C4), 166.3 (C9⁗), 165.93 (C9‴), 164.0 (C7), 162.2 (C5), 159.9 (C4′), 159.7 (C4‴), 159.6 (C4⁗), 157.0 (C2), 156.8 (C9), 145.1 (C7‴), 144.6 (C7⁗), 133.2 (C3), 131.1 , 130.1 , 126.2 (C1‴), 126.0 (C1⁗), 121.9 (C1′), 115.82 , 115.8 , 115.10 , 114.8 (C8‴), 114.23 (C8⁗), 104.7 (C10), 98.9 (C1″), 98.7 (C6), 93.60 (C8), 75.0 (C3″), 74.6 (C5″), 74.11 (C2″), 70.81 (C4″), 62.79 (C6″); ESIMS (positive-ion mode) m/z 740.7 [M]+.Amorphous yellow powder;−1; 1H NMR  δ 7.73 , 7.56 , 7.55 , 7.50 , 7.42 , 7.33 , 6.85 , 6.83 , 6.44 , 6.27 , 6.10 , 6.09 , 5.63 , 5.13 , 4.38 , 4.26 , 3.68 , 3.56 , 3.46 ;13C NMR  δ 179.2 (C4), 168.8 (C9⁗), 168.6 (C9‴), 165.7 (C7), 163.0 (C5), 161.3 (C4‴), 161.2 (C4⁗), 158.8 (C2), 158.3 (C9), 149.7 (C4′), 147.1 (C7‴), 146.6 (C7⁗), 146.0 (C3′), 134.6 (C3), 131.3 , 131.2 , 127.3 (C1⁗), 127.1 (C1‴), 123.5 (C6′), 123.1 (C1′), 117.1 (C2′), 116.8 , 116.8 , 116.0 (C5′), 115.2 (C8‴), 114.7 (C8⁗), 105.7 (C10), 100.4 (C1″), 99.8 (C6), 94.6 (C8), 76.2 (C3″), 76.0 (C5″), 75.7 (C2″), 72.0 (C4″), 64.2 (C6″); ESIMS (negative-ion mode) m/z 738.6 [M − H2O + H]−.Amorphous yellow powder; ® Kinetex C18 100 Å  reversed-phase column. The column temperature was kept at 35 °C. Working solutions of samples were prepared by dissolving 10, 1 and 1 mg of extracts of E-cymosum (WE and BuS), isolated compounds, and standards, respectively, with the appropriate solvent for each . All samples were filtered on membrane filters  and injected (2 μL). Elution was carried out with water containing 0.1% (v/v) formic acid as solvent A and acetonitrile (MeCN) as solvent B, starting with a mixture of 99% of A and 1% of B, increasing the amount of solvent B as follows: 25% at 14 min, 30% at 14–18 min, 35% at 18–22 min, 95% at 22–27, holding this mixture for a minute and returning to the initial conditions at 30 min. The flow rate was 0.35 mL/min. For UV detection, the wavelength program was set at an acquisition of λ 240, 254, 280, 320 and 365 nm; the UV spectra were recorded from 230 at 400.The HPLC profiles were developed using an Agilent 1260 HPLC instrument with the specification mentioned in 2SO4, 10% SDS, and 10% ascorbic acid. Finally, they were incubated at 45 °C for 20 min and measured at 830 nm.Rat liver microsomal fractions isolated through differential centrifugation were diluted 1:1 in buffer  and added to reaction mixtures, containing buffer and different concentrations of inhibitor samples (chlorogenic acid or compounds). Enzymatic activity started by adding 20 mM of glucose-6-phosphate (G6P). Next, the mixtures were incubated at 22 °C for 20 min and ended with the addition of 900 μL of a solution of 0.42% ammonium molybdate in 1 N H2, 50 mM Tris-HCl, pH 7.2) enriched with 0.1 mM of fructose-1,6-bisphosphate was added and incubated for 5 min with different concentrations of inhibitor or samples (adenosine 5′-monophosphate or compounds) in reaction mixtures of 1 mL. Enzymatic activity started by adding rat liver cytosolic supernatant diluted 9:1 and, 15 min later, ended with the addition of a color reagent previously prepared by mixing 1 volume of 7.5% ammonium molybdate and 0.17% TWEEN 20 with 4 volumes of stock solution . Finally, the reaction mixtures were incubated for 10 min at room temperature and measured at 630 nm.Buffer , rosmarinic acid (2), caffeic acid (3), protocatechuic acid (4), kaempferol-3-O--β-d-glucopyranoside (5), and quercetin-3-O--β-d-glucopyranoside (6) from the aerial parts of Eryngium cymosum. Moreover, this is the first report of the isolation and elucidation of quercetin-3-O--β-d-glucopyranoside (6). Recently, the acute hypoglycemic effect of the aqueous extract of Eryngium cymosum has been proved [5 over G6Pase and FBPase could play an important role in the hypoglycemic effect of the plant in the fasting state, but it is possible that the six isolated compounds play a major role in the overall hypoglycemic effect of the extract since they could act in a synergic way. More experiments are required to corroborate it.This work is the first report on isolation of chlorogenic acid (


Cytotoxicity assay shows that RuRe-1 and RuRe-2 exhibit higher anticancer activity than cisplatin, and exist certain selectivity toward human cancer cells over normal cells. The anticancer mechanistic studies reveal that RuRe-1 and RuRe-2 can induce apoptosis through the regulation of cell cycle, depolarization of mitochondrial membrane potential (MMP), elevation of intracellular reactive oxygen species (ROS), and caspase cascade. Moreover, RuRe-1 and RuRe-2 can effectively inhibit cell migration and colony formation. Taken together, heteronuclear Ru(Ⅱ)-Re(Ⅰ) metal complexes possess the prospect of developing new anticancer agents with high efficacy.The development of heteronuclear metal complexes as potent anticancer agents has received increasing attention in recent years. In this study, two new heteronuclear Ru(Ⅱ)-Re(Ⅰ) metal complexes, [Ru(bpy) Compareoperties . Brenda operties . Pt(II)-operties . The resoperties . There aoperties  and pH loperties  effects 2LRe(CO)3(DIP)](PF6)3 and [Ru(phen)2LRe(CO)3(DIP)](PF6)3 imidazoliophenanthroline, bpy = 2,2′-bipyridine, DIP = 4,7-diphenyl-1,10-phenanthroline, phen = 1,10-phenanthroline), were designed and synthesized , reactive oxygen species (ROS), cell migration and colony formation. These findings will contribute to the development of heteronuclear metal complexes in anticancer field.In this context, two heterobimetallic complexes based on Ru(II) and Re(I) units, [Ru(bpy)thesized . We firsRuRe-1 and RuRe-2 are shown in Ru-1 2Cl2]·2H2O (cis-[Ru(bpy)2Cl2]·2H2O  and heteronuclear RuRe-1 and RuRe-2 were recorded in phosphate buffered saline (PBS), dichloromethane and acetonitrile at 298 K  of mononuclear complexes  and heteronuclear RuRe-1 and RuRe-2 have also been determined. The Фem of RuRe-1 and RuRe-2 were similar to those of Ru-1 and Ru-2. The photophysical data were summarized in The electronic absorption and emission spectra of mononuclear complexes  were tested through UV–Vis spectroscopy. As shown in RuRe-1 and RuRe-2 collected at 0, 24, and 48 h, indicating that these complexes are stable under physiological conditions.The stabilities of Po/w)  . Herein,RuRe-1 and RuRe-2 was first qualitatively investigated by confocal microscopy. As shown in RuRe-1 and RuRe-2 for 6 h, the ratio of intracellular ruthenium and rhenium content was approximately 1:1 , HepG2 (human hepatocellular carcinoma), A549 (human lung cancer cells), A549R (cisplatin-resistant A549) and one normal cell line LO2  were determined by 3--2,5-diphenyltetraazolium bromide (MTT) assay, while the mononuclear complexes Ru-1, Ru-2, Re-1 and cisplatin were selected as controls. As shown in RuRe-1 and RuRe-2 showed the best anticancer effects on HeLa cells, with IC50 values of 3.1 μM and 2.5 μM, respectively. And the in vitro antiproliferative efficacies of the compounds against HeLa cells were in the following order: RuRe-2 > RuRe-1 > Re-1 > cisplatin > Ru-1 > Ru-2. The mononuclear ruthenium complexes Ru-1 and Ru-2 showed negligible antitumor activity against all cancer cell lines screened. The anticancer activity of the mononuclear rhenium complex Re-1 was between 5.6 μM and 8.0 μM, which contributed the most to the antitumor activities of the heteronuclear Ru(II)-Re(Ⅰ) metal complexes. In addition, RuRe-1 and RuRe-2 displayed approximately 8.7-fold and 5.4-fold greater ability to kill A549R cells than cisplatin, indicating that they can conquer the resistance of cisplatin. Furthermore, the cytotoxicity of RuRe-1 and RuRe-2 against LO2 cells was lower than that of HepG2 cells, revealing their selectivity to cancer cells.The anticancer activities of Apoptosis is an evolutionarily conserved form of programmed cell death that is essential for animal development and tissue homeostasis . ApoptosRuRe-1 and RuRe-2 were examined with 2'-(4-ethoxyphenyl)-5-(4-methyl-1-piperazinyl)-2,5′-bi-1H-benzimidazole trihydrochloride (Hoechst 33342) staining. Hoechst 33342 is a blue fluorescent dye that can penetrate cell membranes and is commonly used to stain cell nuclei  double-staining. In the early stage of apoptosis, the surface of the cell membrane is broken, at which point the phosphatidylserine on the surface of the apoptotic cell flips from the inner cell membrane to the outer cell membrane, where it can be labelled by Annexin V  increased in a concentration-dependent manner. Specifically, treatment with RuRe-1 (12.4 μM) or RuRe-2 (10.0 μM) significantly increased the percentage of apoptotic cells from 1.81% (control) to 22.38% (RuRe-1) and 22.26% (RuRe-2), respectively. Overall, the results indicate their ability to induce apoptosis in HeLa cells.The ability of nnexin V . The memnnexin V . As showRuRe-1 or RuRe-2 for 24 h, the expression of caspase-3 and the caspase substrate PARP apoptotic protein were detected by western blot. As shown in RuRe-1 and RuRe-2 induced the cleavage of caspase-3 and PARP in a dose-dependent manner. It was further shown that RuRe-1 and RuRe-2 are activators of caspase-3, which can trigger apoptosis in HeLa cells.Cysteine aspartate proteases play an important role in apoptosis, especially caspase-3 protein, which is one of the most important execution factors in the apoptotic pathway . The polRuRe-1 and RuRe-2, we analyzed the cell cycle of HeLa cells treated with RuRe-1 and RuRe-2 by flow cytometry with PI staining. The results (RuRe-1 (12.4 μM) and RuRe-2 (10.0 μM) treated groups increased from 14.4 to 33.8% and 40.0%, respectively. A corresponding decrease in cell content in G1 and G2 phases was observed. The result suggests that RuRe-1 and RuRe-2 may induce cell death by regulating the cell cycle.Cell cycle arrest studies provide a good understanding of cell apoptosis induced by metal drugs . To furt results  showed tRuRe-1 and RuRe-2 on MMP. A decrease in fluorescence intensity of rhodamine 123 (Rh123) can indicate the loss of MMP . H2DCFDA is non-fluorescent and can be oxidized by intracellular ROS to highly fluorescent 2′,7′-dichlorofluorescein (DCF)  . The intRuRe-1 and RuRe-2 on inhibiting cell migration were studied through wound healing. As compared to control group, HeLa cells treated with RuRe-1 (3.1 μM) and RuRe-2 (2.5 μM) exhibited a significant time-dependent inhibition of wound healing integrity  and RuRe-2 (7.5 μM). The quantitative graphs showed that the cell populations at this time were only 4.8% and 3.9% compared to the control group , bpy (J&K), phen (J&K), DIP (J&K), Re(CO)5Cl (Sigma Aldrich), silver trifluoromethanesulfonate (Alfa Aesar), 4-pyridinecarboxaldehyde (Alfa Aesar), 5,6-diamino-1,10-phenanthroline (Alfa Aesar), NH4PF6 (Alfa Aesar), 4% paraformaldehyde (Beyotime), crystal violet (Beyotime), Dulbecco’s Modified Eagle Medium , Roswell Park Memorial Institute 1640 , Fetal bovine serum , penicillin-streptomycin (Gibco), MTT (J&K), Rh123 (J&K), H2DCFDA (J&K), Hoechst 33342 (J&K), Annexin V-FITC Apoptosis Detection Kit (Beyotime). Primary antibodies against caspase-3 and PARP were purchased from Cell Signaling Technology. RuRe-1, RuRe-2 were dissolved in DMSO just before the experiments, and the concentration of DMSO in biological experiments was 1% (v/v). Cisplatin was dissolved in 0.9% sodium chloride solution just before use.RuCl1H NMR spectra. A SpetraMax M2 plate reader was used for determining cell viability. A Nikon A1R/A1 laser-scanning confocal microscope was used for obtaining cell imaging images. A CyFlow Space flow cytometer was used for performing the flow cytometry analysis.A LCQ DECA XP spectrometer was used for obtaining ESI-MS spectra. A Bruker Avance 600 spectrometer was used for obtaining Ru-1 2Cl2]·2H2O (cis-[Ru(bpy)2Cl2]·2H2O  δ 9.08 , 8.87 , 8.78 , 8.40 , 8.23 , 8.14 , 8.06 , 7.83 , 7.77 . ESI-MS (CH3CN): m/z 758.1345 [M-PF6]+, 379.5706 [M-2PF6]2+.RuRe-1: The synthetic route of RuRe-1 is shown in Ru-1  and Re-1  were dissolved in 60 ml acetone. After stirred at 329 K for 24 h under nitrogen, the reaction solution was concentrated to 5 ml. A red precipitate was obtained by dropwise addition of saturated NH4PF6 aqueous solution. Then, the solid was purified by silica gel column chromatography . The PF6 salt of RuRe-1 was again formed by adding saturated NH4PF6 aqueous solution, and then dried under vacuum. Yield: 79% (red solid). 1H NMR  δ 9.88 , 8.97–8.71 , 8.31–8.22 , 8.24–8.12 , 8.08 , 7.99 , 7.83 , 7.77–7.64 , 7.58 , 7.32 . FT-IR (KBr) νCO/cm−1: 2032.45, 1914.89. ESI-MS (CH3CN): m/z 656.6029 [M-3PF6-H]2+, 603.0723 [M-Ru(bpy)2L-2PF6]+, 355.5717 [M-Re(DIP)(CO)3-3PF6]2+. Elemental analysis: calcd (%) for C65H43F18N11O3P3ReRu: C, 44.66; H, 2.48; N, 8.81; found: C, 44.56; H, 2.63; N, 8.92.RuRe-2: Complex RuRe-2 was prepared following a similar procedure to that of RuRe-1, except that Ru-2 was used instead of Ru-1. Yield: 82% (red solid). 1H NMR  δ 9.89 , 8.89 , 8.81–8.68 , 8.39 , 8.31–8.24 , 8.20–8.12 , 8.08 , 7.95 , 7.79–7.64 . FT-IR (KBr) νCO/cm−1: 2031.69, 1918.85. ESI-MS (CH3CN): m/z 680.6032 [M-3PF6-H]2+, 603.0723 [M-Ru(phen)2L-2PF6]+, 379.5722 [M-Re(DIP)(CO)3-3PF6]2+. Elemental analysis: calcd (%) for C69H43F18N11O3P3ReRu: C, 46.14; H, 2.41; N, 8.58; found: C, 46.46; H, 2.66; N, 8.42.


N-heterocycles , as a result of the spontaneous intramolecular heterocyclization of the intermediate reaction product of an SN2 substitution of a hydrogen atom in the oxime group of the amidoxime fragment by a nitrobenzenesulfonyl group, produce spiropyrazolinium ortho- or para-nitrobenzenesulfonates. An exception is ortho-nitrobenzenesulfochlorination of β-(thiomorpholin-1-yl)propioamidoxime, which is regioselective at room temperature, producing two spiropyrazolinium salts (ortho-nitrobezenesulfonate and chloride), and regiospecific at the boiling point of the solvent, when only chloride is formed. The para-Nitrobezenesulfochlorination of β-(benzimidazol-1-yl)propioamidoxime, due to the reduced nucleophilicity of the aromatic β-amine nitrogen atom, is regiospecific at both temperatures, and produces the O-para-nitrobenzenesulfochlorination product. The antidiabetic screening of the new nitrobezenesulfochlorination amidoximes found promising samples with in vitro α-glucosidase activity higher than the reference drug acarbose. 1H-NMR spectroscopy and X-ray analysis revealed the slow inversion of six-membered heterocycles, and experimentally confirmed the presence of an unfavorable stereoisomer with an axial N–N bond in the pyrazolinium heterocycle.Nitrobenzenesulfochlorination of β-aminopropioamidoximes leads to a set of products depending on the structure of the initial interacting substances and reaction conditions. Amidoximes, functionalized at the terminal C atom with six-membered  Heterocycles with potential bioactive properties are of great interest, first of all, for medical chemists working in the field of heterocyclic compounds synthesis. Among the new drugs approved by the FDA in 2021, almost 50% are substances with nitrogen-containing heterocycles . . 23]. S,6S)-3-benzhydryl 4′-methoxycarbonyl- and -3-benzhydryl 4′-benzoyl-5′,5′-diphenyl-4′,5′-dihydrospiro-3-carboxylates up to 75%. The change of the alkylidene substituent to CO2t-Bu leads to a stereoselective, but regioisomeric, mixture of 1,2- and 2,3-spiropyrazolines: -3-benzhydryl 4′-tert-butoxycarbonyl-5′,5′-diphenyl-4′,5′-dihydrospiro-3-carboxylate and (6R)-3-benzhydryl 3′-acetyl-1′,5′-dihydrospiro-3-carboxylate in a ratio of 46 and 10% -3N2 reaction mechanism  of β-(morpholin-1-yl)aminopropioamidoxime -dec-1-ene-5-ammonium nitrobenzenesulfonates and chloride and the product of para-nitrobenzenesulfochlorination, on the oxygen atom of β-(benzimidazol-1-yl)propioamidoxime were obtained. Among them, samples with high in vitro antidiabetic activity were found. The significance of the present work is the establishment of the ambiguity of amidoximes sulfochlorination and the possibility of obtaining a wide range of potentially biologically active products.Previously, we obtained new spiropyrazolium compounds via the hydrolysis of 3-(β-heteroamino)ethyl-5-aryl-1,2,4-oxadiazoles , by the midoxime  and by tmidoxime . In the 1–5) with ortho-, para-nitrobenzenesulfochlorides in CHCl3 was carried out at room temperature and by heating the reaction mixture to the solvent boiling point. A change in the electronic properties of the sulfochlorinating agent, the transition from tosyl chloride to ortho-, para-nitrobenzenesulfochlores lead to an increase in the reaction time at room temperature from 15–20 h in the case of tosylation dec-1-en-5-ammonium chloride monohydrate (13b) is preferred by −8.1, −6.18, −3.08 kJ/mol, respectively dec-1-en-5-ammonium 4-nitrobenzenesulfonate (6). The reaction mixture consisting of 0.5 g (0.0029 mol) of compound 1 in 20 mL of CHCl3 and 0.38 g (0.0029 mol) of DIPEA was cooled to -1℃. Then, 0.64 g (0.0029 mol) of 4-nitrobenzenesulfochloride was added dropwise. When the reaction mixture was kept at r.t. for 38 h, 0.80 g (77%) of white solid 6 was obtained  of white solid 6 was obtained); m.p. 151 °C, Rf 0.12. IR : 1665 (C=N), 1607 (C=C), 1232 (SO2 as) and 1190 (SO2 sy), 1529 (NO2 as) and 1352 (NO2 sy), 3302 [N(-H)2], 2938, 2864, 2815 (Csp3−H), 3032, 3257 (Csp2−H). 1H-NMR : 3.10 , 3.81 , 1.55 m, 1.75 m, 1.87 m, , 3.35  and 3.44 , 7.23 , 7.83−8.21 H). 13C-NMR : 21.0, 21.9, 31.5, 60.7, 64.3 (2C), 126.0 (2C), 128.5 (2C), 138.0 (1C), 145.3 (1C), 168.5. Anal. Calcd for C14H20N4O5S (356.40): C, 47.18; H, 5.66. Found: C, 47.36; H, 5.37.2-Amino-8-oxa-1,5-diazaspiro[4.5]dec-1-ene-5-ammonium 4-nitrobenzenesulfonate (7).The reaction mixture consisting of 0.5 g (0.0029 mol) of compound 2 in 20 mL of CHCl3 and 0.38 g (0.0029 mol) of DIPEA was cooled to −1 °C. Then, 0.64 g (0.0029 mol) of 4-nitrobenzenesulfochloride was added dropwise. When the reaction mixture was kept at r.t. for 120 h, 0.73 g (70%) of white solid 7 was obtained  of white solid 7 was obtained); m.p. 187–188 °C, Rf 0.10. IR : 1650 (C=N), 1600 (C=C), 1231 (SO2 as) and 1194 (SO2 sy), 1520 (NO2 as) and 1362 (NO2 sy), 3466 [N(-H)2], 2967, 2854 (Csp3−H), 3236, 3328, 3388 (Csp2−H). 1H-NMR : 3.13 , 3.92 , 3.32 , 3.41  and 3.65 , 7.28 , 7.80−8.19 H). 13C-NMR : 31.4, 62.1, 62.4, 63.2, 123.8 (2C), 127.4 (2C), 147.7 (1C), 154.8 (1C), 170.1. Anal. Calcd for C13H18N4O6S (358.37): C, 43.57; H, 5.06. Found: C, 43.48; H, 5.13.2-Amino-8-thia-1,5-diazaspiro[4.5]dec-1-en-5-ammonium 4-nitrobenzenesulfonate (8). The reaction mixture consisting of 0.55 g (0.0029 mol) of compound 3 in 20 mL of CHCl3 and 0.38 g (0.0029 mol) of DIPEA was cooled to −1 °C. Then, 0.64 g (0.0029 mol) of 4-nitrobenzenesulfochloride was added dropwise. When the reaction mixture was kept at r.t. for 84 h, 0.74 g (68%) of white solid 8 was obtained  of white solid 8 was obtained); m.p. 230 °C, Rf 0.10. IR : 1642 (C=N), 1588 (C=C), 1229 (SO2 as) and 1197 (SO2 sy), 1519 (NO2 as) and 1349 (NO2 sy), 3308, 3396 [N(-H)2], 2944 (Csp3−H), 3105, 3199, 3245 (Csp2−H). 1H-NMR : 3.11 , 3.86 , 2.87  and 3.15 , 3.60  and 3.72 , 7.37 , 7.83−8.20 H). 13C-NMR : 23.2, 31.4, 62.5, 64.7, 123.8 (2C), 127.3 (2C), 147.7 (1C), 154.8 (1C), 169.0. Anal. Calcd for C13H18N4O5S2 (374.44): C, 41.70; H, 4.85. Found: C, 41.59; H, 4.33.2-Amino-8-phenyl-1,5,8-triazaspiro[4.5]dec-1-en-5-ammonium 4-nitrobenzenesulfonate (9). The reaction mixture consisting of 0.72 g (0.0029 mol) of compound 4 in 20 mL of CHCl3 and 0.38 g (0.0029 mol) of DIPEA was cooled to −1 °C. Then, 0.64 g (0.0029 mol) of 4-nitrobenzenesulfochloride was added dropwise. When the reaction mixture was kept at r.t. for 60 h, 0.88 g (70%) of white solid 9 was obtained  of white solid 9 was obtained); m.p. 203 °C, Rf 0.15. IR : 1649 (C=N), 1599 (C=C), 1240 (SO2 as) and 1189 (SO2 sy), 1515 (NO2 as) and 1350 (NO2 sy), 3421 [N(-H)2], 2790, 2880, 2915 (Csp3−H), 3118, 3290 (Csp2−H). 1H-NMR : 3.17 , 3.95 , 3.56 , 3.49  and 3.98 , 7.25 , 7.81−8.53 H). 13C-NMR : 31.5, 44.5, 61.5, 62.9, 115.3 (2C), 120.4 (1C), 123.8 (2C), 127.4 (2C), 129.5 (2C), 147.7 (1C), 149.9 (1C), 156.7 (1C), 169.0. Anal. Calcd for C19H23N5O5S (433.48): C, 52.64; H, 5.35. Found: C, 52.35; H, 5.21.3-(1H-benzo[d]imidazol-1-yl)-N′-{[(4-nitrophenyl)sulfonyl]oxy}propanimidamide (10). The reaction mixture consisting of 0.59 g (0.0029 mol) of compound 5 in 20 mL of CHCl3 and 0.38 g (0.0029 mol) of DIPEA was cooled to −1 °C. Then, 0.64 g (0.0029 mol) of 4-nitrobenzenesulfochloride was added dropwise. When the reaction mixture was kept at r.t. for 80 h, 0.93 g (82%) of white solid 10 was obtained  of white solid 10 was obtained); m.p. 158 °C, Rf 0.75. IR : 1648 (C=N), 1617 (C=C), 1240 (SO2 as) and 1187 (SO2 sy), 1520 (NO2 as) and 1365 (NO2 sy), 3417 [N(-H)2], 2791, 2920 (Csp3−H), 3110, 3237 (Csp2−H). 1H-NMR : 2.50 , 4.33 , 6.50 , 7.78−8.44 H), 8.05 H). 13C-NMR : 31.2, 42.9, 110.8 (2C), 119.8 (2C), 121.9 (1C), 122.7 (1C), 128.5 (2C), 130.0 (2C), 133.5 (2C), 134.0 (1C), 143.7 (1C), 144.3 (1C), 144.7 (1C), 160.0. Anal. Calcd for C16H15N5O5S (389.39): C, 49.35; H, 3.88. Found: C, 49.28; H, 3.45.The synthesis of β-aminopropioamidoximes2-nitrobenzenesulfochlorinationproducts6–10. To a solution of 0.0029 mol of β-aminopropioamidoximes 1–5 in 20 mL of CHCl3, 0.0029 mol of DIPEA was added. The reaction mixture was cooled to −1 °C, and a solution of 0.0029 mol of 2-nitrobenzenesulfochloride in 2 mL of CHCl3 was added dropwise with stirring. The reaction mixture was then allowed to warm to r.t. (or up to the b.p. of CHCl3) and stirred until the completion of the reaction. The progress of the reaction was monitored by TLC. The formed white precipitates of the products were filtered off and recrystallized from 2-PrOH.2-Amino-1,5-diazaspiro[4.5]dec-1-en-5-ammonium 2-nitrobenzenesulfonate (11). The reaction mixture consisting of 0.5 g (0.0029 mol) of compound 1 in 20 mL of CHCl3 and 0.38 g (0.0029 mol) of DIPEA was cooled to −1 °C. Then, 0.64 g (0.0029 mol) of 2-nitrobenzenesulfochloride was added dropwise. When the reaction mixture was kept at r.t. for 35 h, 0.80 g (77%) of white solid 11 was obtained  of white solid 11 was obtained); m.p. 153 °C, Rf 0.05. IR : 1657 (C=N), 1593 (C=C), 1221, 1232 (SO2 as) and 1024 (SO2 sy), 1541 (NO2 as) and 1377 (NO2 sy), 3399 [N(-H)2], 2949 (Csp3−H), 3204, 3398 (Csp2−H). 1H-NMR : 3.05 , 3.77 , 1.54m, 1.71m, 1.84m, , 3.35 , 7.20d, 7.46−7.81 H). 13C-NMR : 21.0, 21.9(2C), 31.6, 60.7, 64.3, 122.0, 129.5, 130.5, 131.3, 140.0, 148.3, 168.6. Anal. Calcd for C14H20N4O5S (356.40): C, 47.18; H, 5.66. Found: C, 47.57; H, 5.33.2-Amino-8-oxa-1,5-diazaspiro[4.5]dec-1-ene-5-ammonium 2-nitrobenzenesulfonate (12). The reaction mixture consisting of 0.5 g (0.0029 mol) of compound 2 in 20 mL of CHCl3 and 0.38 g (0.0029 mol) of DIPEA was cooled to −1 °C. Then, 0.64 g (0.0029 mol) of 2-nitrobenzenesulfochloride was added dropwise. When the reaction mixture was kept at r.t. for 25 h, 0.97 g (93%) of white solid 12 was obtained  of white solid 12 was obtained); m.p. 148 °C, Rf 0.01. IR : 1637 (C=N), 1593 (C=C), 1207, 1228 (SO2 as) and 1022 (SO2 sy), 1549 (NO2 as) and 1377 (NO2 sy), 3387 [N(-H)2], 2907 (Csp3−H), 3250, 3306, 3328 (Csp2−H). 1H-NMR : 3.09 , 3.88 , 3.35  and 3.60 , 3.88 , 7.20 , 7.45−7.81 H). 13C-NMR : 31.5, 62.2 (2C), 63.2, 122.0, 129.5, 130.6, 131.3, 140.0, 148.3, 169.2. Anal. Calcd for C13H18N4O6S (358.37): C, 43.57; H, 5.06. Found: C, 43.89; H, 5.47.2-Amino-8-thia-1,5-diazaspiro[4.5]dec-1-en-5-ammonium 2-nitrobenzenesulfonate (13a) and 2-Amino-8-thia-1,5-diazaspiro[4.5]dec-1-en-5-ium chloride hydrate (13b). The reaction mixture consisting of 0.55 g (0.0029 mol) of compound 3 in 20 mL of CHCl3 and 0.38 g (0.0029 mol) of DIPEA was cooled to −1 °C. Then, 0.64 g (0.0029 mol) of 2-nitrobenzenesulfochloride was added dropwise. When the reaction mixture was kept at r.t. for 104 h, the white precipitate of the mixture of products 13a and 13b was obtained. After recrystallization, 0.16 g (25%) of 13b was obtained. After the evaporation of the filtrate from the recrystallization to ½ volume. 0.27 g (25%) of 2-nitrobenzenesulfonate 13a was formed. When the reaction mixture was kept at CHCl3 b.p. for 24 h, only 0.37 g (56%) of white solid 13b was obtained.13a: m.p. 138−140 °C, Rf 0.08. IR : 1647 (C=N), 1604 (C=C), 1207, 1215 (SO2 as) and 1024 (SO2 sy), 1545 (NO2 as) and 1373 (NO2 sy), 3497 [N(-H)2], 2936, 2964 (Csp3−H), 3152, 3258, 3302 (Csp2−H). 1H-NMR : 3.10 , 3.83 , 3.10  and 3.55 , 3.10  and 3.68 , 7.26 , 7.47−7.81 H). 13C-NMR : 23.2, 31.5, 62.6, 64.7, 122.9, 129.5, 130.6, 131.3, 139.7, 148.3, 169.1. Anal. Calcd for C13H18N4O5S2 (374.44): C, 41.70; H, 4.85. Found: C, 41.67; H, 4.46.13b: m.p. > 280 °C, Rf 0.08. IR : 1659 (C=N); 1612 ; 670 [S−C]; 3135, 3230, 3380, 3384 . 1H-NMR : 3.10 , 3.85 , 2.85  and 3.55 , 3.10  and 3.68 , 7.26 . 13C-NMR : 23.2, 31.5, 62.6, 64.7, 169.1. Anal. Calcd for C7H16ClN3OS (225.74): C, 37.24; H, 7.14. Found: C, 37.52; H, 7.48.2-Amino-8-phenyl-1,5,8-triazaspiro[4.5]dec-1-en-5-ammonium 2-nitrobenzenesulfonate (14). The reaction mixture consisting of 0.72 g (0.0029 mol) of compound 4 in 20 mL of CHCl3 and 0.38 g (0.0029 mol) of DIPEA was cooled to −1 °C. Then, 0.64 g (0.0029 mol) of 2-nitrobenzenesulfochloride was added dropwise. When the reaction mixture was kept at r.t. for 34 h, 0.99 g (79%) of white solid 14 was obtained (when reaction mixture was kept at CHCl3 b.p. for 24 h 1.02 g (81%) white solid 14 was obtained); m.p. 185−187 °C, Rf 0.08. IR : 1647 (C=N), 1593 (C=C), 1217, 1224 (SO2 as) and 1024 (SO2 sy), 1535 (NO2 as) and 1366 (NO2 sy), 3373 [N(-H)2], 2837 (Csp3−H), 3161, 3300 (Csp2−H). 1H-NMR : 3.12 , 3.91 , 3.40 , 3.71 , 3.54 , 6.82−7.81 H). 13C-NMR : 31.6, 44.6, 61.5, 62.9, 116.4, 120.5, 122.9, 129.5, 129.7, 130.6, 131.3, 139.8, 148.3, 150.0, 169.2. Anal. Calcd for C19H23N5O5S (433.48): C, 52.64; H, 5.35. Found: C, 52.92; H, 5.63.14 was obtained [when reaction mixture was kept at CHCl3 b.p. for 24 h 1.02 g (81%) white solid 14 was obtained].When reaction mixture was kept at r.t. for 34 h, 0.99 g (79%) white solid 6–14 was assessed by the degree of inhibition of α-amylase and α-glucosidase activity. Pure DMSO was used as a solvent. The final concentration of the sample substances was 10 mg/mL. A total of 100 µL of α-amylase or α-glucosidase (1 U/mL) and 200 µL of the test sample solution (10 mg/mL) were added to 500 µL of phosphate buffer . The resulting mixture was incubated for 15 min at +37 °C, and 200 µL of P-NPG solution (5 mM) was added.The in vitro antidiabetic activity of samples Then, the resulting mixture was incubated again at +37 °C for 20 min. The reaction was stopped by the addition of 500 μL of sodium carbonate (0.1 M). Since the samples showed too much absorbance at 405 nm, they were diluted 5 times with 5 mL of water and 1 mL of sodium carbonate solution (0.1 M). A solution of α-amylase or α-glucosidase (1 U/mL) was used as a blank. As a negative control, 200 μL of pure DMSO was used in triplicate. As a reference drug, acarbose was obtained at a concentration of 10.0 mg/mL (positive control).Simultaneously, a negative control was placed without the addition of the test compounds. All the samples were examined in triplets. The inhibitory activity was expressed as a percentage (%) of the degree of inhibition of α-glucosidase, in comparison to the negative control.8, 9, and 11–14 were collected on a Bruker Apex II diffractometer  equipped with an Oxford Cryostream cooling unit and a graphite monochromated Mo anode (λ = 0.71073 Å). The intensities of the reflections for 10 were collected at 100 K at the “Belok” beamline of the Kurchatov Synchrotron Radiation Source , at the wavelength of 0.745 Å using a MAR CCD 165 detector. The image integration was performed using the iMosflm software -dec-1-ene-5-ammonium ortho-nitrobezenesulfonate and chloride hydrate. Heating leads to a regiospecific course of the reaction with the formation of only chloride hydrate.The set of the reaction products of β-aminopropioamidoximes nitrobenzenesulfochlorination depends on the structure of the initial substrates and temperature. β-Aminopropioamidoximes with six-membered heterocycles in the β-aminogroup produce good yields of 2-amino-1,5-diazaspiro[4.5]-dec-1-ene-5-ammonium nitrobenzenesulfonates at r.t. and CHClpara-Nitrobenzenesulfochlorination of β-(benzimidazol-1-yl)propioamidoxime produces the O-para-nitrobenzenesulfochlorination product. The reaction time when the reaction mixture is heated is reduced by 2–3 times. The in vitro screening of the library of nitrobenzenelsulfochlorination products for antidiabetic activity reveals two samples with high α-glucosidase activity exceeding the activity of the acarbose standard: products of para-nitrobenzeneulfochlorination of β-(thiomorpholin-1-yl)- and β-(benzimidazol-1-yl)propioamidoximes. The arsenal of the physicochemical and spectral methods made it possible to establish the structural features of the studied spiropyrazolinium organic salts. Thus, in DMSO-d6 solutions in the 1H NMR spectra, the slow inversion of six-membered nitrogen-containing β-heterocycles can be observed. These spiropyrazoline derivatives can be interesting objects in dynamic NMR spectroscopy, which would allow for the rotational barriers of six-membered heterocycles to be measured. It is assumed that the bulky substituted arylsulfonate anion anchors the spiroheterocycles in the most thermodynamically favorable chair-like position. According to X-ray diffraction data, the axial location of the N–N bond in the spiropyrazoline heterocycles is unambiguously determined. The NMR and XRD data demonstrate that two various conformations of spirocation are present both in solution and in solids. The cation can take part in different types of intermolecular interactions, depending on the conformation and the nature of the six-membered cycle.The