Development of multifunctional heterocyclic Schiff base as a potential metal chelator: a comprehensive spectroscopic approach towards drug discovery

  • Manojkumar Jadhao
  • Chayan Das
  • Anoop Rawat
  • Himank Kumar
  • Ritika Joshi
  • Sudipta Maiti
  • Sujit Kumar Ghosh
Original Paper


Amyloid-β peptides and their metal-associated aggregated states have been implicated in the pathogenesis of Alzheimer’s disease. The present paper epitomises the design and synthesis of a small, neutral, lipophilic benzothiazole Schiff base (E)-2-((6-chlorobenzo[d]thiazol-2-ylimino)methyl)-5-diethylamino)phenol (CBMDP), and explores its multifunctionalty as a potential metal chelator/fluorophore using UV–visible absorption, steady-state fluorescence, single molecule fluorescence correlation spectroscopic (FCS) techniques which is further corroborated by in silico studies. Some pharmaceutically relevant properties of the synthesized compound have also been calculated theoretically. Steady-state fluorescence and single molecule FCS reveal that the synthesized CBMDP not only recognizes oligomeric Aβ40, but could also be used as an amyloid-specific extrinsic fluorophore as it shows tremendous increase in its emission intensity in the presence of Aβ40. Molecular docking exercise and MD simulation reveal that CBMDP localizes itself in the crucial amyloidogenic and copper-binding region of Aβ40 and undergoes a strong binding interaction via H-bonding and π–π stacking. It stabilizes the solitary α-helical Aβ40 monomer by retaining the initial conformation of the Aβ central helix and mostly interacts with the hydrophilic N-terminus and the α-helical region spanning from Ala-2 to Val-24. CBMDP exhibits strong copper as well as zinc chelation ability and retards the rapid copper-induced aggregation of amyloid peptide. In addition, CBMDP shows radical scavenging activity which enriches its functionality. Overall, the consolidated in vitro and in silico results obtained for the synthesized molecule could provide a rational template for developing new multifunctional agents.


Schiff base Electronic absorption Steady-state fluorescence Amyloid beta Single molecule fluorescence correlation spectroscopy 



SKG gratefully acknowledges the financial support from the Department of Science and Technology, India (Project no. SR/FT/LS/-172/2009) MJ thanks VNIT for his fellowship. The authors thank G. Krishnamoorthy, TIFR Mumbai, for providing steady-state fluorescence facility. The authors appreciate and thank the anonymous reviewers and the respected editor for their kind suggestions to improve the quality of the manuscript.

Supplementary material

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Supplementary material 1 (PDF 1385 kb)


  1. 1.
    Wang Z-M, Xie S-S, Li X-M et al (2015) Multifunctional 3-Schiff base-4-hydroxycoumarin derivatives with monoamine oxidase inhibition, anti-β-amyloid aggregation, metal chelation, antioxidant and neuroprotection properties against Alzheimer’s disease. RSC Adv 5:70395–70409. doi: 10.1039/C5RA13594J CrossRefGoogle Scholar
  2. 2.
    Yanagisawa S (2012) Metal complexes for optical recording material, JP 5036190Google Scholar
  3. 3.
    Choudhury S, Kakoti M, Deb AK, Goswami S (1992) Isomeric complexes of ruthenium(II) with neutral heterocyclic Schiff base ligands. High resolution proton resonance spectra of trans-cis isomeric pairs of RuX2L2 (L = 2-arylpyridinecarboxaldimine, X = Cl, Br) and comparison of their physical properties. Polyhedron 11:3183–3190. doi: 10.1016/S0277-5387(00)83661-X CrossRefGoogle Scholar
  4. 4.
    Sarkar B, Konar S, Gómez-García CJ, Ghosh A (2008) Rare example of mu-nitrito-1kappa2O, O’:2kappaO coordinating mode in copper(II) nitrite complexes with monoanionic tridentate Schiff base ligands: structure, magnetic, and electrochemical properties. Inorg Chem 47:11611–11619. doi: 10.1021/ic8011519 CrossRefPubMedGoogle Scholar
  5. 5.
    Heffern MC, Velasco PT, Matosziuk LM et al (2014) Modulation of amyloid-β aggregation by histidine-coordinating Cobalt(III) Schiff base complexes. ChemBioChem 15:1584–1589. doi: 10.1002/cbic.201402201 CrossRefPubMedPubMedCentralGoogle Scholar
  6. 6.
    Ghosh C, Seal M, Mukherjee S, Ghosh Dey S (2015) Alzheimer’s disease: a heme-Aβ perspective. Acc Chem Res 48:2556–2564. doi: 10.1021/acs.accounts.5b00102 CrossRefPubMedGoogle Scholar
  7. 7.
    Association Alzheimer's (2016) 2016 Alzheimer’s disease facts and figures. Alzheimer’s Dementia 12(4):459–509. doi: 10.1016/j.jalz.2016.03.001 CrossRefGoogle Scholar
  8. 8.
    Sengupta P, Garai K, Sahoo B et al (2003) The amyloid beta peptide (Abeta(1-40)) is thermodynamically soluble at physiological concentrations. Biochemistry 42:10506–10513. doi: 10.1021/bi0341410 CrossRefPubMedGoogle Scholar
  9. 9.
    Rauk A (2009) The chemistry of Alzheimer’s disease. Chem Soc Rev 38:2698–2715. doi: 10.1039/b807980n CrossRefPubMedGoogle Scholar
  10. 10.
    Davies P, Maloney AJF (1976) Selective loss of central cholinergic neurons in Alzheimer’s disease. Lancet 308:1403. doi: 10.1016/S0140-6736(76)91936-X CrossRefGoogle Scholar
  11. 11.
    Hardy JA, Higgins GA (1992) Alzheimer’s disease: the amyloid cascade hypothesis. Science 256:184–185CrossRefPubMedGoogle Scholar
  12. 12.
    Butterfield DA (1997) beta-Amyloid-associated free radical oxidative stress and neurotoxicity: implications for Alzheimer’s disease. Chem Res Toxicol 10:495–506. doi: 10.1021/tx960130e CrossRefPubMedGoogle Scholar
  13. 13.
    Christen Y (2000) Oxidative stress and Alzheimer disease. Am J Clin Nutr 71:621s–629sPubMedGoogle Scholar
  14. 14.
    Sengupta K, Chatterjee S, Pramanik D et al (2014) Self-assembly of stable oligomeric and fibrillar aggregates of Aβ peptides relevant to Alzheimer’s disease: morphology dependent Cu/heme toxicity and inhibition of PROS generation. Dalton Trans 43:13377–13383. doi: 10.1039/c4dt01991a CrossRefPubMedGoogle Scholar
  15. 15.
    Bonda DJ, Lee H, Blair JA et al (2011) Role of metal dyshomeostasis in Alzheimer’s disease. Metallomics 3:267–270. doi: 10.1039/c0mt00074d CrossRefPubMedPubMedCentralGoogle Scholar
  16. 16.
    Dubois B, Feldman HH, Jacova C et al (2010) Revising the definition of Alzheimer’s disease: a new lexicon. Lancet Neurol 9:1118–1127. doi: 10.1016/S1474-4422(10)70223-4 CrossRefPubMedGoogle Scholar
  17. 17.
    Kayed R, Lasagna-Reeves C (2013) Molecular mechanisms of amyloid oligomers toxicity. J Alzheimers Dis 33:S67–S78. doi: 10.3233/JAD-2012-129001 CrossRefPubMedGoogle Scholar
  18. 18.
    Jarosz-Griffiths HH, Noble E, Rushworth JV, Hooper NM (2016) Amyloid-β receptors: the good, the bad, and the prion protein. J Biol Chem 291:3174–3183. doi: 10.1074/jbc.R115.702704 CrossRefPubMedGoogle Scholar
  19. 19.
    Masters CL, Simms G, Weinman NA et al (1985) Amyloid plaque core protein in Alzheimer disease and Down syndrome 82:4245–4249Google Scholar
  20. 20.
    Hane FT, Lee BY, Petoyan A et al (2014) Testing synthetic amyloid-β aggregation inhibitor using single molecule atomic force spectroscopy. Biosens Bioelectron 54:492–498. doi: 10.1016/j.bios.2013.10.060 CrossRefPubMedGoogle Scholar
  21. 21.
    Hane FT, Leonenko Z (2014) Effect of Metals on Kinetic Pathways of Amyloid-β Aggregation. Biomolecules 4:101–116. doi: 10.3390/biom4010101 CrossRefPubMedPubMedCentralGoogle Scholar
  22. 22.
    Balland V, Hureau C, Saveant J-M (2010) Electrochemical and homogeneous electron transfers to the Alzheimer amyloid–copper complex follow a preorganization mechanism. Proc Natl Acad Sci 107:17113–17118. doi: 10.1073/pnas.1011315107 CrossRefPubMedPubMedCentralGoogle Scholar
  23. 23.
    Sharma AK, Pavlova ST, Kim J et al (2012) Bifunctional compounds for controlling metal-mediated aggregation of the aβ42 peptide. J Am Chem Soc 134:6625–6636. doi: 10.1021/ja210588m CrossRefPubMedPubMedCentralGoogle Scholar
  24. 24.
    Hindo SS, Mancino AM, Braymer JJ et al (2009) Small molecule modulators of copper-induced Aβ aggregation. J Am Chem Soc 131:16663–16665. doi: 10.1021/ja907045h CrossRefPubMedPubMedCentralGoogle Scholar
  25. 25.
    Rodríguez-Rodríguez C, Sánchez de Groot N, Rimola A et al (2009) Design, selection, and characterization of thioflavin-based intercalation compounds with metal chelating properties for application in Alzheimer’s disease. J Am Chem Soc 131:1436–1451CrossRefPubMedGoogle Scholar
  26. 26.
    León R, Garcia AG, Marco-Contelles J (2013) Recent advances in the multitarget-directed ligands approach for the treatment of Alzheimer’s disease. Med Res Rev 33:139–189. doi: 10.1002/med.20248 CrossRefPubMedGoogle Scholar
  27. 27.
    Cavalli A, Bolognesi ML, Minarini A et al (2008) Multi-target-directed ligands to combat neurodegenerative diseases. J Med Chem 51:347–372. doi: 10.1021/jm7009364 CrossRefPubMedGoogle Scholar
  28. 28.
    Braymer JJ, Choi JS, Detoma AS et al (2011) Development of bifunctional stilbene derivatives for targeting and modulating metal-amyloid-β species. Inorg Chem 50:10724–10734. doi: 10.1021/ic2012205 CrossRefPubMedPubMedCentralGoogle Scholar
  29. 29.
    Choi J-S, Braymer JJ, Nanga RPR et al (2010) Design of small molecules that target metal-A{beta} species and regulate metal-induced A{beta} aggregation and neurotoxicity. Proc Natl Acad Sci USA 107:21990–21995. doi: 10.1073/pnas.1006091107 CrossRefPubMedPubMedCentralGoogle Scholar
  30. 30.
    Lee S, Zheng X, Krishnamoorthy J et al (2014) Rational design of a structural framework with potential use to develop chemical reagents that target and modulate multiple facets of Alzheimer’s disease. J Am Chem Soc 136:299–310. doi: 10.1021/ja409801p CrossRefPubMedGoogle Scholar
  31. 31.
    Sengupta P, Balaji J, Maiti S (2002) Measuring diffusion in cell membranes by fluorescence correlation spectroscopy. Methods 27:374–387. doi: 10.1016/S1046-2023(02)00096-8 CrossRefPubMedGoogle Scholar
  32. 32.
    Halgren TA, Murphy RB, Friesner RA et al (2004) Glide: a new approach for rapid, accurate docking and scoring. 2. Enrichment factors in database screening. J Med Chem 47:1750–1759. doi: 10.1021/jm030644s CrossRefPubMedGoogle Scholar
  33. 33.
    Sahoo B, Balaji J, Nag S et al (2008) Protein aggregation probed by two-photon fluorescence correlation spectroscopy of native tryptophan. J Chem Phys 129:075103. doi: 10.1063/1.2969110 CrossRefPubMedGoogle Scholar
  34. 34.
    Garai K, Sureka R, Maiti S (2007) Detecting amyloid-beta aggregation with fiber-based fluorescence correlation spectroscopy. Biophys J 92:L55–L57. doi: 10.1529/biophysj.106.101485 CrossRefPubMedPubMedCentralGoogle Scholar
  35. 35.
    Klein SM, Cohen G, Cederbaum AI (1981) Production of formaldehyde during metabolism of dimethyl sulfoxide by hydroxyl radical-generating systems. Biochemistry 20:6006–6012. doi: 10.1021/bi00524a013 CrossRefPubMedGoogle Scholar
  36. 36.
    Darghal N, Garnier-Suillerot A, Salerno M (2006) Mechanism of thioflavin T accumulation inside cells overexpressing P-glycoprotein or multidrug resistance-associated protein: role of lipophilicity and positive charge. Biochem Biophys Res Commun 343:623–629. doi: 10.1016/j.bbrc.2006.03.024 CrossRefPubMedGoogle Scholar
  37. 37.
    Begley D (2004) ABC transporters and the blood–brain barrier. Curr Pharm Des 10:1295–1312. doi: 10.2174/1381612043384844 CrossRefPubMedGoogle Scholar
  38. 38.
    Mathis CA, Bacskai BJ, Kajdasz ST et al (2002) A lipophilic thioflavin-T derivative for positron emission tomography (PET) imaging of amyloid in brain. Bioorg Med Chem Lett. doi: 10.1016/S0960-894X(01)00734-X PubMedGoogle Scholar
  39. 39.
    Nesterov EE, Skoch J, Hyman BT et al (2005) In vivo optical imaging of amyloid aggregates in brain: design of fluorescent markers. Angew Chemie Int Ed 44:5452–5456. doi: 10.1002/anie.200500845 CrossRefGoogle Scholar
  40. 40.
    Ioakimidis L, Thoukydidis L, Mirza A et al (2008) Benchmarking the reliability of QikProp. Correlation between experimental and predicted values. QSAR Comb Sci 27:445–456. doi: 10.1002/qsar.200730051 CrossRefGoogle Scholar
  41. 41.
    Carrico D, Ohkanda J, Kendrick H et al (2004) In vitro and in vivo antimalarial activity of peptidomimetic protein farnesyltransferase inhibitors with improved membrane permeability. Bioorg Med Chem 12:6517–6526. doi: 10.1016/j.bmc.2004.09.020 CrossRefPubMedGoogle Scholar
  42. 42.
    Löscher W, Potschka H (2005) Drug resistance in brain diseases and the role of drug efflux transporters. Nat Rev Neurosci 6:591–602. doi: 10.1038/nrn1728 CrossRefPubMedGoogle Scholar
  43. 43.
    Löscher W, Potschka H (2005) Blood-brain barrier active efflux transporters: ATP-binding cassette gene family. NeuroRx 2:86–98. doi: 10.1602/neurorx.2.1.86 CrossRefPubMedPubMedCentralGoogle Scholar
  44. 44.
    Telpoukhovskaia MA, Rodríguez-Rodríguez C, Cawthray JF et al (2014) 3-hydroxy-4-pyridinone derivatives as metal ion and amyloid binding agents. Metallomics 6:249–262. doi: 10.1039/c3mt00135k CrossRefPubMedGoogle Scholar
  45. 45.
    LeVine H (1999) Amyloid, prions, and other protein aggregates. Methods Enzymol 309:274–284. doi: 10.1016/S0076-6879(99)09020-5 CrossRefPubMedGoogle Scholar
  46. 46.
    Kirschner DA, Inouye H, Duffy LK et al (1987) Synthetic peptide homologous to beta protein from Alzheimer disease forms amyloid-like fibrils in vitro. Proc Natl Acad Sci USA 84:6953–6957. doi: 10.1073/pnas.84.19.6953 CrossRefPubMedPubMedCentralGoogle Scholar
  47. 47.
    Wood SJ, Wetzel R, Martin JD, Hurle MR (1995) Prolines and amyloidogenicity in fragments of the Alzheimer’s peptide beta/A4. Biochemistry 34:724–730CrossRefPubMedGoogle Scholar
  48. 48.
    Garbuzynskiy SO, Lobanov MY, Galzitskaya OV (2010) FoldAmyloid: a method of prediction of amyloidogenic regions from protein sequence. Bioinformatics 26:326–332. doi: 10.1093/bioinformatics/btp691 CrossRefPubMedGoogle Scholar
  49. 49.
    Sticht H, Bayer P, Willbold D et al (1995) Structure of amyloid A4-(1-40)-peptide of Alzheimer’s disease. Eur J Biochem 233:293–298. doi: 10.1111/j.1432-1033.1995.293_1.x CrossRefPubMedGoogle Scholar
  50. 50.
    Kepp KP (2012) Bioinorganic chemistry of Alzheimer’s disease. Chem Rev 112:5193–5239. doi: 10.1021/cr300009x CrossRefPubMedGoogle Scholar
  51. 51.
    Miller Y, Ma B, Nussinov R (2012) Metal binding sites in amyloid oligomers: complexes and mechanisms. Coord Chem Rev 256:2245–2252. doi: 10.1016/j.ccr.2011.12.022 CrossRefGoogle Scholar
  52. 52.
    Xu L, Wang X, Wang X (2013) Effects of Zn 2+ binding on the structural and dynamic properties of amyloid Β peptide associated with Alzheimer’s disease: Asp 1 or Glu 11 ? ACS Chem Neurosci 4:1458–1468. doi: 10.1021/cn4001445 CrossRefPubMedPubMedCentralGoogle Scholar
  53. 53.
    Ito M, Johansson J, Strömberg R, Nilsson L (2012) Effects of ligands on unfolding of the amyloid β-peptide central helix: mechanistic insights from molecular dynamics simulations. PLoS ONE 7:e30510. doi: 10.1371/journal.pone.0030510 CrossRefPubMedPubMedCentralGoogle Scholar
  54. 54.
    Xu Y, Shen J, Luo X et al (2005) Conformational transition of amyloid beta-peptide. Proc Natl Acad Sci USA 102:5403–5407. doi: 10.1073/pnas.0501218102 CrossRefPubMedPubMedCentralGoogle Scholar
  55. 55.
    Raffa DF, Rauk A (2007) Molecular dynamics study of the beta amyloid peptide of Alzheimer’s disease and its divalent copper complexes. J Phys Chem B 111:3789–3799. doi: 10.1021/jp0689621 CrossRefPubMedGoogle Scholar
  56. 56.
    Geng J, Li M, Wu L et al (2012) Liberation of copper from amyloid plaques: making a risk factor useful for Alzheimer’s disease treatment 55:9146–9155Google Scholar
  57. 57.
    Hane FT, Hayes R, Lee BY, Leonenko Z (2016) Effect of copper and zinc on the single molecule self-affinity of Alzheimer’s amyloid-β peptides. PLoS ONE 11:e0147488. doi: 10.1371/journal.pone.0147488 CrossRefPubMedPubMedCentralGoogle Scholar
  58. 58.
    Lakowicz JR (2006) Principles of fluorescence spectroscopy, 3rd edn. Springer, New YorkCrossRefGoogle Scholar
  59. 59.
    Jadhao M, Ahirkar P, Kumar H et al (2015) Surfactant induced aggregation–disaggregation of photodynamic active chlorin e6 and its relevant interaction with DNA alkylating quinone in a biomimic micellar microenvironment. RSC Adv 5:81449–81460. doi: 10.1039/C5RA16181A CrossRefGoogle Scholar
  60. 60.
    Sharma AK, Kim J, Prior JT et al (2014) Small bifunctional chelators that do not disaggregate amyloid β fibrils exhibit reduced cellular toxicity. Inorg Chem 53:11367–11376CrossRefPubMedPubMedCentralGoogle Scholar
  61. 61.
    Lovell M, Robertson J, Teesdale W et al (1998) Copper, iron and zinc in Alzheimer’s disease senile plaques. J Neurol Sci 158:47–52. doi: 10.1016/S0022-510X(98)00092-6 CrossRefPubMedGoogle Scholar
  62. 62.
    Huang X, Atwood CS, Hartshorn MA et al (1999) The Aβ peptide of Alzheimer’s disease directly produces hydrogen peroxide through metal ion reduction. Biochemistry 38:7609–7616. doi: 10.1021/bi990438f CrossRefPubMedGoogle Scholar
  63. 63.
    Huang X, Cuajungco MP, Atwood CS et al (1999) Cu(II) potentiation of Alzheimer a neurotoxicity: correlation with cell-free hydrogen peroxide production and metal reduction. J Biol Chem 274:37111–37116. doi: 10.1074/jbc.274.52.37111 CrossRefPubMedGoogle Scholar
  64. 64.
    Mayes J, Tinker-Mill C, Kolosov O et al (2014) Amyloid fibrils in alzheimer disease are not inert when bound to copper ions but can degrade hydrogen peroxide and generate reactive oxygen species. J Biol Chem 289:12052–12062. doi: 10.1074/jbc.M113.525212 CrossRefPubMedPubMedCentralGoogle Scholar
  65. 65.
    Gutteridge JMC (1984) Lipid peroxidation initiated by superoxide-dependent hydroxyl radicals using complexed iron and hydrogen peroxide. FEBS Lett 172:245–249. doi: 10.1016/0014-5793(84)81134-5 CrossRefPubMedGoogle Scholar
  66. 66.
    Cooke MS, Evans MD, Dizdaroglu M, Lunec J (2003) Oxidative DNA damage: mechanisms, mutation, and disease. FASEB J 17:1195–1214. doi: 10.1096/fj.02-0752rev CrossRefPubMedGoogle Scholar
  67. 67.
    Xu G, Chance MR (2007) Hydroxyl radical-mediated modification of proteins as probes for structural proteomics. Chem Rev 107:3514–3543. doi: 10.1021/cr0682047 CrossRefPubMedGoogle Scholar

Copyright information

© SBIC 2016

Authors and Affiliations

  • Manojkumar Jadhao
    • 1
  • Chayan Das
    • 1
  • Anoop Rawat
    • 2
  • Himank Kumar
    • 1
  • Ritika Joshi
    • 1
  • Sudipta Maiti
    • 2
  • Sujit Kumar Ghosh
    • 1
  1. 1.Department of ChemistryVisvesvaraya National Institute of TechnologyNagpurIndia
  2. 2.Tata Institute of Fundamental Research (TIFR)ColabaIndia

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