Analytical and Bioanalytical Chemistry

, Volume 411, Issue 10, pp 2189–2200 | Cite as

Enhanced His@AuNCs oxidase-like activity by reduced graphene oxide and its application for colorimetric and electrochemical detection of nitrite

  • Lu Liu
  • Jie DuEmail author
  • Wen-e Liu
  • Yongliang Guo
  • Guofan Wu
  • Weinan Qi
  • Xiaoquan LuEmail author
Research Paper


Enzyme-mimicking (nanozyme)-based biosensors are attractive owing to their unique catalytic efficiency, multifunctionality, and tunable activity, but examples of oxidase-like nanozymes are quite rare. Herein, we demonstrated that histidine-capped gold nanoclusters (His@AuNCs) possessed intrinsic oxidase-like activity, which could directly oxidize 3,3′,5,5′-tetramethylbenzidine (TMB) to blue colored ox-TMB without H2O2. The assembly of reduced graphene oxide (RGO) with His@AuNCs could further improve its oxidase-like activity and the His@AuNCs/RGO nanocomposites had a lower Michaelis constant (Km) and higher catalytic constant (Kcat) for TMB oxidation. Furthermore, compared to other nanomaterials, the as-prepared His@AuNCs/RGO also exhibited enhanced electrocatalytic activity toward TMB. Interestingly, nitrite inhibited the catalytic (chromogenic) and electrocatalytic processes of His@AuNCs/RGO in the oxidation of TMB. The oxidase-like and electrocatalytic activity of His@AuNCs/RGO was evaluated with nitrite and TMB as substrates, and the results indicated that TMB and nitrite might share the same catalytic active sites. On the basis of these findings, a colorimetric and electrochemical sensor was developed with the His@AuNCs/RGO composite as an oxidase mimic for determination of nitrite with linear ranges of 10–500 μM and 2.5–5700 μM, respectively. The developed method was successfully applied to the detection of nitrites in real samples. The present work suggests that the oxidase-like nanozyme is not only suitable for colorimetric assay but also for development of electrochemical sensors in bioanalysis.

Graphical abstract

The colorimetric and electrochemical detection of nitrite using His@AuNCs/RGO.


Nanozyme, oxidase-like Colorimetric Electrocatalytic Nitrite 



This work was supported by the National Natural Science Foundation of China (21565020).

Compliance with ethical standards

Conflict of interests

The manuscript was written through contributions of all authors. The authors declare that they have no competing interest.

Supplementary material

216_2019_1655_MOESM1_ESM.pdf (716 kb)
ESM 1 (PDF 715 kb)


  1. 1.
    Cheng HJ, Zhang L, He J, Guo WJ, Zhou ZY, Zhang XJ, et al. Integrated nanozymes with nanoscale proximity for in vivo neurochemical monitoring in living brains. Anal Chem. 2016;88(10):5489–97.CrossRefGoogle Scholar
  2. 2.
    Gao LZ, Zhuang J, Nie L, Zhang JB, Zhang Y, Gu N, et al. Intrinsic peroxidase-like activity of ferromagnetic nanoparticles. Nat Nanotechnol. 2007;2(9):577–83.CrossRefGoogle Scholar
  3. 3.
    Liu BW, Liu JW. Surface modification of nanozymes. Nano Res. 2017;10(4):1125–48.CrossRefGoogle Scholar
  4. 4.
    Asati A, Santra S, Kaittanis C, Nath S, Perez JM. Oxidase-like activity of polymer-coated cerium oxide nanoparticles. Angew Chem Int Ed. 2009;48(13):2308–12.CrossRefGoogle Scholar
  5. 5.
    Lin YH, Ren JS, Qu XG. Catalytically active nanomaterials: a promising candidate for artificial enzymes. Acc Chem Res. 2014;47(4):1097–105.CrossRefGoogle Scholar
  6. 6.
    Mu JS, Wang Y, Zhao M, Zhang L. Intrinsic peroxidase-like activity and catalase-like activity of Co3O4 nanoparticles. Chem Commun. 2012;48(19):2540–2.CrossRefGoogle Scholar
  7. 7.
    Tseng CW, Chang HY, Chang JY, Huang CC. Detection of mercury ions based on mercury-induced switching of enzyme-like activity of platinum/gold nanoparticles. Nanoscale. 2012;4(21):6823–30.CrossRefGoogle Scholar
  8. 8.
    Nasir M, Nawaz MH, Latif U, Yaqub M, Hayat A, Rahim A. An overview on enzyme-mimicking nanomaterials for use in electrochemical and optical assays. Microchim Acta. 2017;184(2):323–42.CrossRefGoogle Scholar
  9. 9.
    Pan N, Wang LY, Wu LL, Peng CF, Xie ZJ. Colorimetric determination of cysteine by exploiting its inhibitory action on the peroxidase-like activity of Au@Pt core-shell nanohybrids. Microchim Acta. 2017;184(1):65–72.CrossRefGoogle Scholar
  10. 10.
    Shao K, Wang BR, Ye SY, Zuo YP, Wu L, Li Q, et al. Signal-amplified near-infrared ratiometric electrochemiluminescence aptasensor based on multiple quenching and enhancement effect of graphene/gold nanorods/G-quadruplex. Anal Chem. 2016;88(16):8179–87.CrossRefGoogle Scholar
  11. 11.
    Masud MK, Yadav S, Isam MN, Nguyen NT, Salomon C, Kline R, et al. Gold-loaded nanoporous ferric oxide nanocubes with peroxidase-mimicking activity for electrocatalytic and colorimetric detection of autoantibody. Anal Chem. 2017;89(20):11005–13.CrossRefGoogle Scholar
  12. 12.
    Xu Q, Yuan H, Dong X, Zhang Y, Asif M, Dong Z, et al. Dual nanoenzyme modified microelectrode based on carbon fiber coated with AuPd alloy nanoparticles decorated graphene quantum dots assembly for electrochemical detection in clinic cancer samples. Biosens Bioelectron. 2018;107:153–62.CrossRefGoogle Scholar
  13. 13.
    Cheng HJ, Lin SC, Muhammad F, Lin YW, Wei H. Rationally modulate the oxidase-like activity of Nanoceria for self regulated bioassays. Acs Sensors. 2016;1(11):1336–43.CrossRefGoogle Scholar
  14. 14.
    Guo LL, Mao L, Huang KX, Liu HM. Pt-Se nanostructures with oxidase-like activity and their application in a selective colorimetric assay for mercury(II). J Mater Sci. 2017;52(18):10738–50.CrossRefGoogle Scholar
  15. 15.
    Wang G-L, Jin L-Y, Dong Y-M, Wu X-M, Li Z-J. Intrinsic enzyme mimicking activity of gold nanoclusters upon visible light triggering and its application for colorimetric trypsin detection. Biosens Bioelectron. 2015;64:523–9.CrossRefGoogle Scholar
  16. 16.
    Yu CJ, Chen TH, Jiang JY, Tseng WL. Lysozyme-directed synthesis of platinum nanoclusters as a mimic oxidase. Nanoscale. 2014;6(16):9618–24.CrossRefGoogle Scholar
  17. 17.
    Deng HH, Lin XL, Liu YH, Li KL, Zhuang QQ, Peng HP, et al. Chitosan-stabilized platinum nanoparticles as effective oxidase mimics for colorimetric detection of acid phosphatase. Nanoscale. 2017;9(29):10292–300.CrossRefGoogle Scholar
  18. 18.
    Borghei YS, Hosseini M, Ganjali MR. Oxidase-like catalytic activity of Cys-AuNCs upon visible light irradiation and its application for visual miRNA detection. Sens Actuators B Chem. 2018;273:1618–26.CrossRefGoogle Scholar
  19. 19.
    Wang GL, Jin LY, Wu XM, Dong YM, Li ZJ. Label-free colorimetric sensor for mercury(II) and DNA on the basis of mercury(II) switched-on the oxidase-mimicking activity of silver nanoclusters. Anal Chim Acta. 2015;871:1–8.CrossRefGoogle Scholar
  20. 20.
    Tanaka S, Kaneti YV, Bhattacharjee R, Islam MN, Nakahata R, Abdullah N, et al. Mesoporous Iron oxide synthesized using poly(styrene-b-acrylic acid-b-ethylene glycol) block copolymer micelles as templates for colorimetric and electrochemical detection of glucose. ACS Appl Mater Interfaces. 2018;10(1):1039.CrossRefGoogle Scholar
  21. 21.
    Bhattacharjee R, Tanaka S, Moriam S, Masud MK, Lin J, Alshehri SM, et al. Porous nanozymes: peroxidase-mimetic activity of mesoporous iron oxide for colorimetric and electrochemical detection of global DNA methylation. J Mater Chem B. 2018.
  22. 22.
    Chong Y, Dai X, Fang G, Wu R, Zhao L, Ma X, et al. Palladium concave nanocrystals with high-index facets accelerate ascorbate oxidation in cancer treatment. Nat Commun. 2018;9(1):4861.CrossRefGoogle Scholar
  23. 23.
    Hu YH, Cheng HJ, Zhao XZ, Wu JJ, Muhammad F, Lin SC, et al. Surface-enhanced Raman scattering active gold nanoparticles with enzyme-mimicking activities for measuring glucose and lactate in living tissues. ACS Nano. 2017;11(6):5558–66.CrossRefGoogle Scholar
  24. 24.
    Canfield DE, Glazer AN, Falkowski PG. The evolution and future of Earth’s nitrogen cycle. Science. 2010;330(6001):192–6.CrossRefGoogle Scholar
  25. 25.
    Maia LB, Pereira V, Mira L, Moura JJG. Nitrite reductase activity of rat and human xanthine oxidase, xanthine dehydrogenase, and aldehyde oxidase: evaluation of their contribution to NO formation in vivo. Biochemistry. 2015;54(3):685–710.CrossRefGoogle Scholar
  26. 26.
    Halsted TP, Yamashita K, Hirata K, Ago H, Ueno G, Tosha T, et al. An unprecedented dioxygen species revealed by serial femtosecond rotation crystallography in copper nitrite reductase. IUCrJ. 2018;5:22–31.CrossRefGoogle Scholar
  27. 27.
    Hematian S, Siegler MA, Karlin KD. Heme/copper assembly mediated nitrite and nitric oxide interconversion. J Am Chem Soc. 2012;134(46):18912–5.CrossRefGoogle Scholar
  28. 28.
    Maia LB, Moura JJG. How biology handles nitrite. Chem Rev. 2014;114(10):5273–357.CrossRefGoogle Scholar
  29. 29.
    D’Ischia M, Napolitano A, Manini P, Panzella L. Secondary targets of nitrite-derived reactive nitrogen species: nitrosation/nitration pathways, antioxidant defense mechanisms and toxicological implications. Chem Res Toxicol. 2011;24(12):2071.CrossRefGoogle Scholar
  30. 30.
    Cross AJ, Ferrucci LM, Risch A, Graubard BI, Ward MH, Park YY, et al. A large prospective study of meat consumption and colorectal cancer risk: an investigation of potential mechanisms underlying this association. Cancer Res. 2010;70(6):2406.CrossRefGoogle Scholar
  31. 31.
    Shen Y, Rao DJ, Bai WS, Sheng QL, Zheng JB. Preparation of high-quality palladium nanocubes heavily deposited on nitrogen-doped graphene nanocomposites and their application for enhanced electrochemical sensing. Talanta. 2017;165:304–12.CrossRefGoogle Scholar
  32. 32.
    Uzer A, Saglam S, Can Z, Ercag E, Apak R. Electrochemical determination of food preservative nitrite with gold nanoparticles/p-aminothiophenol-modified gold electrode. Int J Mol Sci. 2016;17(8):17.CrossRefGoogle Scholar
  33. 33.
    Zou CE, Yang BB, Bin D, Wang J, Li SM, Yang P, et al. Electrochemical synthesis of gold nanoparticles decorated flower-like graphene for high sensitivity detection of nitrite. J Colloid Interface Sci. 2017;488:135–41.CrossRefGoogle Scholar
  34. 34.
    Zhang X-X, Song Y-Z, Fang F, Wu Z-Y. Sensitive paper-based analytical device for fast colorimetric detection of nitrite with smartphone. Anal Bioanal Chem. 2018;410(11):2665–9.CrossRefGoogle Scholar
  35. 35.
    Bos ES, van der Doelen AA, van Rooy N, Schuurs AH. 3,3′,5,5′-Tetramethylbenzidine as an Ames test negative chromogen for horseradish peroxidase in enzyme-immunoassay. J Immunoass. 1981;2(3-4):187–204.CrossRefGoogle Scholar
  36. 36.
    Hummers WS Jr, Offeman RE. Preparation of graphitic oxide. J Am Chem Soc. 1958;80(6):1339.CrossRefGoogle Scholar
  37. 37.
    Titelman GI, Gelman V, Bron S, Khalfin RL, Cohen Y, Bianco-Peled H. Characteristics and microstructure of aqueous colloidal dispersions of graphite oxide. Carbon. 2005;43(3):641–9.CrossRefGoogle Scholar
  38. 38.
    Phiri J, Johansson LS, Gane P, Maloney T. A comparative study of mechanical, thermal and electrical properties of graphene-, graphene oxide- and reduced graphene oxide-doped microfibrillated cellulose nanocomposites. Composites Part B Eng. 2018;147:104–113.Google Scholar
  39. 39.
    Guo Y, Zhao X, Long T, Lin M, Liu Z, Huang C. Histidine-mediated synthesis of chiral fluorescence gold nanoclusters: insight into the origin of nanoscale chirality. RSC Adv. 2015;5(75):61449–54.CrossRefGoogle Scholar
  40. 40.
    Zhang N, Qiu H, Liu Y, Wang W, Li Y, Wang X, et al. Fabrication of gold nanoparticle/graphene oxide nanocomposites and their excellent catalytic performance. J Mater Chem. 2011;21(30):11080–3.CrossRefGoogle Scholar
  41. 41.
    Mao Z, Hu HH, Su R, Liu PZ, Li YX, Zhang WT, et al. Confining gold nanoclusters in highly defective graphitic layers to enhance the methanol electrooxidation reaction. ChemCatChem. 2018;10(1):141–7.CrossRefGoogle Scholar
  42. 42.
    Zhan L, Li CM, Wu WB, Huang CZ. A colorimetric immunoassay for respiratory syncytial virus detection based on gold nanoparticles-graphene oxide hybrids with mercury-enhanced peroxidase-like activity. Chem Commun (Camb). 2014;50(78):11526–8.CrossRefGoogle Scholar
  43. 43.
    He Y, Wang X, Zhu J, Zhong S, Song G. Ni2+-modified gold nanoclusters for fluorescence turn-on detection of histidine in biological fluids. Analyst. 2012;137(17):4005–9.CrossRefGoogle Scholar
  44. 44.
    Zhang X, Wu F-G, Liu P, Gu N, Chen Z. Enhanced fluorescence of gold nanoclusters composed of HAuCl4 and histidine by glutathione: glutathione detection and selective cancer cell imaging. Small. 2014;10(24):5170–7.CrossRefGoogle Scholar
  45. 45.
    Yang HK, Xiao JY, Su L, Feng T, Lv QY, Zhang XJ. Oxidase-mimicking activity of the nitrogen-doped Fe3C@C composites. Chem Commun. 2017;53(27):3882–5.CrossRefGoogle Scholar
  46. 46.
    Gao M, Lu XF, Nie GD, Chi MQ, Wang C. Hierarchical CNFs/MnCo2O4.5 nanofibers as a highly active oxidase mimetic and its application in biosensing. Nanotechnology. 2017;28(48).Google Scholar
  47. 47.
    Liu Y, Ding D, Zhen YL, Guo R. Amino acid-mediated ‘turn-off/turn-on’ nanozyme activity of gold nanoclusters for sensitive and selective detection of copper ions and histidine. Biosens Bioelectron. 2017;92:140–6.CrossRefGoogle Scholar
  48. 48.
    Chen Y, Liu XM, Wu X, Liu XC, Dong WH, Han BK, et al. An array of poly-l-histidine functionalized multi-walled carbon nanotubes on 4-aminothiophenol self-assembled monolayer and the application for sensitively glucose sensing. Electrochim Acta. 2017;258:988–97.CrossRefGoogle Scholar
  49. 49.
    Gevaerd A, Blaskievicz SF, Zarbin AJG, Orth ES, Bergamini MF, Marcolino LH. Nonenzymatic electrochemical sensor based on imidazole-functionalized graphene oxide for progesterone detection. Biosens Bioelectron. 2018;112:108–13.CrossRefGoogle Scholar
  50. 50.
    Liu Y, She P, Gong J, Wu WP, Xu SM, Li JG, et al. A novel sensor based on electrodeposited Au-Pt bimetallic nano-clusters decorated on graphene oxide (GO)-electrochemically reduced GO for sensitive detection of dopamine and uric acid. Sens Actuators B Chem. 2015;221:1542–53.CrossRefGoogle Scholar
  51. 51.
    Wang JM, Wang XY, Wu S, Song J, Zhao YQ, Ge YQ, et al. Fabrication of highly catalytic silver nanoclusters/graphene oxide nanocomposite as nanotag for sensitive electrochemical immunoassay. Anal Chim Acta. 2016;906:80–8.CrossRefGoogle Scholar
  52. 52.
    Palanisamy S, Thirumalraj B, Chen SM. A novel amperometric nitrite sensor based on screen printed carbon electrode modified with graphite/beta-cyclodextrin composite. J Electroanal Chem. 2016;760:97–104.CrossRefGoogle Scholar
  53. 53.
    Yang B, Wang J, Duan B, Zhu M, Yang P, Du Y. A three dimensional Pt nanodendrite/graphene/MnO2 nanoflower modified electrode for the sensitive and selective detection of dopamine. J Mater Chem B. 2015;3(37):7440–8.CrossRefGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  1. 1.College of Life ScienceNorthwest Normal UniversityLanzhouChina
  2. 2.Key Laboratory of Bioelectrochemistry & Environmental of Gansu ProvinceLanzhouChina

Personalised recommendations