Gold Bulletin

, Volume 52, Issue 2, pp 87–97 | Cite as

Horseradish peroxidase-AuNP/LDH heterostructures: influence on nanogold release and enzyme activity

  • Elena-Florentina Grosu
  • Renato FroidevauxEmail author
  • Gabriela CarjaEmail author
Original Paper


Gold nanostructures (AuNP) are important as strong platforms for targeted therapeutic and diagnostic applications. Tireless effort has been devoted nowadays to explore the multifunctionality of AuNP in multicomponent biostructures. Herein, we report the fabrication of horseradish peroxidase enzyme (HRP)-AuNP/ZnAlLDH heterostructure by the facile synthesis of AuNP on the biocompatible matrices of layered double hydroxides (LDH) followed by the immobilization of the enzyme on AuNP/LDH assemblies. During this process, ZnAlLDH have a dual function of exploring its structural “memory effect” for the synthesis of nanogold and acting as a support for the enzyme immobilization. X-ray diffraction (XRD), UV-Vis spectrometry, transmission electronic microscopy (TEM), energy-dispersive X-ray spectroscopy (EDX), and infrared (FTIR) spectroscopy have been used to characterize the structural, chemical composition, optical, and morphology of the novel materials. We present here the release of AuNP from HRP-AuNP/ZnAlLDH by using as controlled variables HRP:LDHs ratio and the pH of the solution. Results show that AuNP established close interactions with HRP and formed an HRP-AuNP bioconjugate. Results reveal that HRP suffers a significant loss of the activity in the presence of nanoparticles of gold, such that, AuNP act to inhibit the activity of the enzyme. AuNP behavior in enzyme-bio-heterostructures should be inspiring for future applications of AuNP in nanomedicine.


Layered double hydroxides Horseradish peroxidase enzyme Nanoparticles of gold Controlled release 



The authors thank Joelle Thuriot (REALCAT platform - “Future Investments” program (PIA), with the contractual reference ANR-11-EQPX-00370) and Eric Gautron (Institut des Materiaux Jean Rouxel - NANTES -France), for their help in obtaining the XRD, TEM, and EDX data. E-F Grosu thanks the Governments of Romania and France for the financial support during the doctoral studies. FEDER is also acknowledged for supporting and funding partially this work.

Supplementary material

13404_2019_256_MOESM1_ESM.docx (1.6 mb)
ESM 1 (DOCX 1629 kb)


  1. 1.
    Ovais M, Raza A, Naz S, Islam NU, Khalil AT, Ali S, Khan MA, Shinwari ZK (2017) Current state and prospects of the phytosynthesized colloidal gold nanoparticles and their applications in cancer theranostics. Appl Microbiol Biotechnol 101:3551–3565. CrossRefGoogle Scholar
  2. 2.
    Boisselier E, Astruc D (2009) Gold nanoparticles in nanomedicine: preparations, imaging, diagnostics, therapies and toxicity. Chem Soc Rev 38:1759–1782. CrossRefGoogle Scholar
  3. 3.
    Huang X, El-Sayed MA (2010) Gold nanoparticles: optical properties and implementations in cancer diagnosis and photothermal therapy. J Adv Res 1:13–28. CrossRefGoogle Scholar
  4. 4.
    Rajeshkumar S (2016) Anticancer activity of eco-friendly gold nanoparticles against lung and liver cancer cells. J Genet Eng Biotechnol 14:195–202. CrossRefGoogle Scholar
  5. 5.
    Ahirwal GK, Mitra CK (2009) Direct electrochemistry of horseradish peroxidase-gold nanoparticles conjugate. Sensors 9:881–894. CrossRefGoogle Scholar
  6. 6.
    Chen PC, Mwakwari SC, Oyelere A (2008) Gold nanoparticles: from nanomedicine to nanosensing. Nanotechnol Sci Appl Volume 1:45–66. CrossRefGoogle Scholar
  7. 7.
    Ginzburg AL, Truong L, Tanguay RL, Hutchison JE (2018) Synergistic toxicity produced by mixtures of biocompatible gold nanoparticles and widely used surfactants. ACS Nano 12:5312–5322. CrossRefGoogle Scholar
  8. 8.
    Mukha I, Vityuk N, Severynovska O, Eremenko A, Smirnova N (2016) The pH-dependent structure and properties of Au and Ag nanoparticles produced by tryptophan reduction. Nanoscale Res Lett 11:1–7. CrossRefGoogle Scholar
  9. 9.
    Mishra A, Sardar M (2015) Cellulase assisted synthesis of nano-silver and gold: application as immobilization matrix for biocatalysis. Int J Biol Macromol 77:105–113. CrossRefGoogle Scholar
  10. 10.
    Keshavarzi M, Davoodi D, Pourseyedi S, Taghizadeh S (2018) The effects of three types of alfalfa plants (Medicago sativa) on the biosynthesis of gold nanoparticles: an insight into phytomining. Gold Bull 51:99–110CrossRefGoogle Scholar
  11. 11.
    Roshmi T, Soumya KR, Jyothis M, Radhakrishnan EK (2015) Effect of biofabricated gold nanoparticle-based antibiotic conjugates on minimum inhibitory concentration of bacterial isolates of clinical origin. Gold Bull 48:63–71. CrossRefGoogle Scholar
  12. 12.
    Durán N, Marcato PD, Durán M, Yadav A, Gade A, Rai M (2011) Mechanistic aspects in the biogenic synthesis of extracellular metal nanoparticles by peptides, bacteria, fungi, and plants. Appl Microbiol Biotechnol 90:1609–1624. CrossRefGoogle Scholar
  13. 13.
    Kalishwaralal K, Deepak V, Ram Kumar Pandian SB, Kottaisamy M, BarathManiKanth S, Kartikeyan B, Gurunathan S (2010) Biosynthesis of silver and gold nanoparticles using Brevibacterium casei. Colloids Surfaces B Biointerfaces 77:257–262. CrossRefGoogle Scholar
  14. 14.
    Mishra A, Singh P, Sardar M (2015) Peroxidase assisted biosynthesis of silver and gold nanoparticles: characterization and computational study. Adv Mater Lett 6:194–200. CrossRefGoogle Scholar
  15. 15.
    Mukherjee P, Senapati S, Mandal D, Ahmad A, Khan MI, Kumar R, Sastry M (2002) Extracellular synthesis of gold nanoparticles by the fungus Fusarium oxysporum. Chembiochem 3:461–463.<461::AID-CBIC461>3.0.CO;2-X CrossRefGoogle Scholar
  16. 16.
    Li L, Weng J (2010) Enzymatic synthesis of gold nanoflowers with trypsin. Nanotechnology 21:305603. CrossRefGoogle Scholar
  17. 17.
    Govindaraju K, Kiruthiga V, Manikandan R, Ashokkumar T, Singaravelu G (2011) β-Glucosidase assisted biosynthesis of gold nanoparticles: a green chemistry approach. Mater Lett 65:256–259. CrossRefGoogle Scholar
  18. 18.
    Kawasaki H, Hamaguchi K, Osaka I, Arakawa R (2011) Ph-dependent synthesis of pepsin-mediated gold nanoclusters with blue green and red fluorescent emission. Adv Funct Mater 21:3508–3515. CrossRefGoogle Scholar
  19. 19.
    Manivasagan P, Venkatesan J, Kang KH, Sivakumar K, Park SJ, Kim SK (2015) Production of α-amylase for the biosynthesis of gold nanoparticles using Streptomyces sp. MBRC-82. Int J Biol Macromol 72:71–78. CrossRefGoogle Scholar
  20. 20.
    Faramarzi MA, Forootanfar H (2011) Biosynthesis and characterization of gold nanoparticles produced by laccase from Paraconiothyrium variabile. Colloids Surf B Biointerfaces 87:23–27. CrossRefGoogle Scholar
  21. 21.
    Chaudiere J, Tappel AL (1984) Interaction of gold(I) with the active site of selenium-glutathione peroxidase. J Inorg Biochem 20:313–325. CrossRefGoogle Scholar
  22. 22.
    Wang P, Wang X, Wang L, Hou X, Liu W, Chen C (2015) Interaction of gold nanoparticles with proteins and cells. Sci Technol Adv Mater 16:34610. CrossRefGoogle Scholar
  23. 23.
    Dahl JA, Maddux BLS, Hutchison JE (2007) Toward greener nanosynthesis. Chem Rev 107:2228–2269. CrossRefGoogle Scholar
  24. 24.
    Thomas M, Klibanov AM (2003) Conjugation to gold nanoparticles enhances polyethylenimine’s transfer of plasmid DNA into mammalian cells. Proc Natl Acad Sci 100:9138–9143. CrossRefGoogle Scholar
  25. 25.
    Niidome T, Akiyama Y, Yamagata M, Kawano T (2009) Poly (ethylene glycol)-modified gold nanorods as a photothermal nanodevice for hyperthermia. J Biomater Sci Polym Ed 20:1203–1215.
  26. 26.
    Vanderkooy A, Chen Y, Gonzaga F, Brook MA (2011) Silica shell/gold core nanoparticles: correlating shell thickness with the plasmonic red shift upon aggregation. ACS Appl Mater Interfaces 3:3942–3947. CrossRefGoogle Scholar
  27. 27.
    Yue Y, Liu TY, Li HW, Liu Z, Wu Y (2012) Microwave-assisted synthesis of BSA-protected small gold nanoclusters and their fluorescence-enhanced sensing of silver(i) ions. Nanoscale 4:2251–2254. CrossRefGoogle Scholar
  28. 28.
    Mieszawska AJ, Mulder WJM, Fayad ZA, Cormode DP (2013) Multifunctional gold nanoparticles for diagnosis and therapy of disease. Mol Pharm 10:831–847. CrossRefGoogle Scholar
  29. 29.
    Zhang XD, Wu HY, Wu D et al (2010) Toxicologic effects of gold nanoparticles in vivo by different administration routes. Int J Nanomedicine 5:771–781. CrossRefGoogle Scholar
  30. 30.
    Parapat RY, Saputra OHI, Ang AP, Schwarze M, Schomäcker R (2014) Support effect in the preparation of supported metal catalysts via microemulsion. RSC Adv 4:50955–50963. CrossRefGoogle Scholar
  31. 31.
    Parapat RY, Wijaya M, Schwarze M, Selve S, Willinger M, Schomäcker R (2013) Particle shape optimization by changing from an isotropic to an anisotropic nanostructure: preparation of highly active and stable supported Pt catalysts in microemulsions. Nanoscale 5:796–805. CrossRefGoogle Scholar
  32. 32.
    Carja G, Kameshima Y, Nakajima A, Dranca C, Okada K (2009) Nanosized silver-anionic clay matrix as nanostructured ensembles with antimicrobial activity. Int J Antimicrob Agents 34:534–539. CrossRefGoogle Scholar
  33. 33.
    Carja G, Grosu EF, Petrarean C, Nichita N (2015) Self-assemblies of plasmonic gold/layered double hydroxides with highly efficient antiviral effect against the hepatitis B virus. Nano Res 8:8–3523. CrossRefGoogle Scholar
  34. 34.
    Mishra G, Dash B, Pandey S, Mohanty PP (2013) Antibacterial actions of silver nanoparticles incorporated Zn-Al layered double hydroxide and its spinel. J Environ Chem Eng 1:1124–1130. CrossRefGoogle Scholar
  35. 35.
    Krainer FW, Glieder A (2015) An updated view on horseradish peroxidases: recombinant production and biotechnological applications. Appl Microbiol Biotechnol 99:1611–1625. CrossRefGoogle Scholar
  36. 36.
    Chiodo F, Marradi M, Calvo J, Yuste E, Penadés S (2014) Glycosystems in nanotechnology: gold glyconanoparticles as carrier for anti-HIV prodrugs. Beilstein J Org Chem 10:1339–1346. CrossRefGoogle Scholar
  37. 37.
    Rautio J, Kumpulainen H, Heimbach T, Oliyai R, Oh D, Järvinen T, Savolainen J (2008) Prodrugs: design and clinical applications. Nat Rev Drug Discov 7:255–270. CrossRefGoogle Scholar
  38. 38.
    Walther R, Rautio J, Zelikin AN (2017) Prodrugs in medicinal chemistry and enzyme prodrug therapies. Adv Drug Deliv Rev 118:65–77. CrossRefGoogle Scholar
  39. 39.
    Mikami G, Grosu F, Kawamura S, Yoshida Y, Carja G, Izumi Y (2016) Harnessing self-supported Au nanoparticles on layered double hydroxides comprising Zn and Al for enhanced phenol decomposition under solar light. Appl Catal B Environ 199:260–271. CrossRefGoogle Scholar
  40. 40.
    Carja G, Grosu EF, Mureseanu M, Lutic D (2017) A family of solar light responsive photocatalysts obtained using Zn 2+ Me 3+ (Me = Al/Ga) LDHs doped with Ga 2 O 3 and in 2 O 3 and their derived mixed oxides: a case study of phenol/4-nitr. Catal Sci Technol 7:5402–5412. CrossRefGoogle Scholar
  41. 41.
    Conterosito E, Gianotti V, Palin L, Boccaleri E, Viterbo D, Milanesio M (2018) Facile preparation methods of hydrotalcite layered materials and their structural characterization by combined techniques. Inorganica Chim Acta 470:36–50. CrossRefGoogle Scholar
  42. 42.
    Xiao G, Zeng H, Huang Q et al (2018) Facile preparation of modifying layered double hydroxide nanoparticles for drug delivery. J Nanosci Nanotechnol 18:5256–5265. CrossRefGoogle Scholar
  43. 43.
    Carja G, Dartu L, Okada K, Fortunato E (2013) Nanoparticles of copper oxide on layered double hydroxides and the derived solid solutions as wide spectrum active nano-photocatalysts. Chem Eng J 222:60–66. CrossRefGoogle Scholar
  44. 44.
    Saifullah B, Hussein M (2015) Inorganic nanolayers: structure, preparation and biomedical applications. Int J Nanomedicine 10:5609-33.
  45. 45.
    Cavani F, Trifiro F, Vaccari A (1991) Hydrotalcite-type anionic clays: preparation, properties and applications. Catal Today 11:173–301CrossRefGoogle Scholar
  46. 46.
    Karunakaran C, Anilkumar P, Gomathisankar P (2011) Photoproduction of iodine with nanoparticulate semiconductors and insulators. Chem Cent J 5:31. CrossRefGoogle Scholar
  47. 47.
    Zhao X, Wang L, Xu X, Lei X, Xu S, Zhang F (2012) Fabrication and photocatalytic properties of novel ZnO/ZnAl2O4 nanocomposite with ZnAl2O4 dispersed inside ZnO network. AICHE J 58(2):573–582. CrossRefGoogle Scholar
  48. 48.
    Wang Z, Zhang Q, Kuehner D, Niu L (2008) Green synthesis of 1-2 nm gold nanoparticles stabilized by amine-terminated ionic liquid and their electrocatalytic activity in oxygen reduction †. Society 1:907–909.
  49. 49.
    Grosu E-F, Cârjă G, Froidevaux R (2018) Development of horseradish peroxidase/layered double hydroxide hybrid catalysis for phenol degradation. Res Chem Intermed 44:7731–7752. CrossRefGoogle Scholar
  50. 50.
    Jobbágy M, Regazzoni AE (2011) Dissolution of nano-size Mg-Al-Cl hydrotalcite in aqueous media. Appl Clay Sci 51:366–369. CrossRefGoogle Scholar
  51. 51.
    Manohara GV, Vishnu Kamath P, Milius W (2012) Reversible hydration and aqueous exfoliation of the acetate-intercalated layered double hydroxide of Ni and Al: observation of an ordered interstratified phase. J Solid State Chem 196:356–361. CrossRefGoogle Scholar
  52. 52.
    Liu X, Luo L, Ding Y, Xu Y (2011) Amperometric biosensors based on alumina nanoparticles-chitosan-horseradish peroxidase nanobiocomposites for the determination of phenolic compounds. Analyst 136:696–701. CrossRefGoogle Scholar
  53. 53.
    Bhattacharyya PRC, KG (2015) Dalton transactions. Dalt Trans 44:6809–6824. CrossRefGoogle Scholar
  54. 54.
    Alshammari A, Köckritz A, Kalevaru VN, Bagabas A, Martin A (2012) Direct oxidation of cyclohexane to adipic acid using nano-gold catalysts. Appl Petrochemical Res 2:61–67. CrossRefGoogle Scholar
  55. 55.
    Benaissi K, Helaine V, Prevot V, Claude Forano LH (2011) Efficient immobilization of yeast transketolase on layered double hydroxides and application for ketose synthesis. Adv Synth Catal 353:1497–1509. CrossRefGoogle Scholar
  56. 56.
    Thapa R, Bhagat C, Shrestha P, Awal S, Dudhagara P (2017) Enzyme-mediated formulation of stable elliptical silver nanoparticles tested against clinical pathogens and MDR bacteria and development of antimicrobial surgical thread. Ann Clin Microbiol Antimicrob 16:1–10. CrossRefGoogle Scholar
  57. 57.
    Lv X, Weng J (2013) Ternary composite of hemin, gold nanoparticles and graphene for highly efficient decomposition of hydrogen peroxide. Sci Rep 3:1–10. Google Scholar
  58. 58.
    Zuber A, Purdey M, Schartner E, Forbes C, van der Hoek B, Giles D, Abell A, Monro T, Ebendorff-Heidepriem H (2016) Detection of gold nanoparticles with different sizes using absorption and fluorescence based method. Sensors Actuators B Chem 227:117–127. CrossRefGoogle Scholar
  59. 59.
    Yakavets I, Yankovsky I, Bezdetnaya L, Zorin V (2017) Soret band shape indicates mTHPC distribution between β-cyclodextrins and serum proteins. Dyes Pigments 137:299–306. CrossRefGoogle Scholar
  60. 60.
    Keighron JD, Keating CD (2013) Enzyme–gold nanoparticle bioconjugates: quantification of particle stoichiometry and enzyme specific activity. In: NanoBiotechnology Protocols, Methods in Molecular Biology, pp 163–174CrossRefGoogle Scholar
  61. 61.
    Seida Y, Nakano Y (2001) Removal of phosphate in dissolution-coagulation process of layered double hydroxide. J Chem Eng Japan 34:906–911. CrossRefGoogle Scholar
  62. 62.
    Osuji AC, Eze SOO, Osayi EE, Chilaka FC (2014) Biobleaching of industrial important dyes with peroxidase partially purified from garlic. Sci World J 2014:1–8. CrossRefGoogle Scholar
  63. 63.
    Chen X, Fu C, Wang Y, Yang W, Evans DG (2008) Direct electrochemistry and electrocatalysis based on a film of horseradish peroxidase intercalated into Ni-Al layered double hydroxide nanosheets. Biosens Bioelectron 24:356–361. CrossRefGoogle Scholar
  64. 64.
    Ajdary M, Negahdary M, Chelongar R, Zadeh S (2015) The antioxidant effects of silver, gold, and zinc oxide nanoparticles on male mice in in vivo condition. Adv Biomed Res 4:69. CrossRefGoogle Scholar
  65. 65.
    Ortego L, Cardoso F, Martins S, Fillat MF, Laguna A, Meireles M, Villacampa MD, Gimeno MC (2014) Strong inhibition of thioredoxin reductase by highly cytotoxic gold(I) complexes. DNA binding studies. J Inorg Biochem 130:32–37. CrossRefGoogle Scholar
  66. 66.
    Gabbiani C, Mastrobuoni G, Sorrentino F, Dani B, Rigobello MP, Bindoli A, Cinellu MA, Pieraccini G, Messori L, Casini A (2011) Thioredoxin reductase, an emerging target for anticancer metallodrugs. Enzyme inhibition by cytotoxic gold(iii) compounds studied with combined mass spectrometry and biochemical assays. Medchemcomm 2:50–54. CrossRefGoogle Scholar
  67. 67.
    Farhad Behzad BM (2014) The effects of gold and silver nanoparticles on an enzymatic reaction between horseradish peroxidase and 3,3′,5,5’-tetramethylbenzidine. Biochem Pharmacol Open Access 03:3–5. CrossRefGoogle Scholar

Copyright information

© Springer Nature Switzerland AG 2019

Authors and Affiliations

  1. 1.Department of Chemical Engineering, Faculty of Chemical Engineering and Environmental ProtectionTechnical University “Gheorghe Asachi” of IasiIasiRomania
  2. 2.Univ. Lille, INRA, ISA, Univ. ArtoisUniv. Littoral Côte d’Opale, EA 7394, ICV - Institut Charles ViolletteLilleFrance

Personalised recommendations