Quantifying Hypochlorous Acid Concentration in Environmental Water Using Smartphone Colorimetry


In this study, we developed an effective method to detect hypochlorite acid (HClO) by using methylene blue (MB) derivative (BPY1). BPY1 was selectively oxidized through HClO, and the solution color changed from colorless to blue. In the presence of HClO, the ultraviolet–visible (UV–vis) spectra and concentration of HClO had a linear relationship with a detection limit of 0.5 μM. Furthermore, a test paper for HClO monitoring was successfully prepared using the BPY1 probe, and the observed detection limit by the naked eye was estimated at 5 μM. Additionally, using the BPY1 probe, HClO could also be detected through smartphone colorimetry, and the method showed a good recovery ranging from 98.7 to 104.0% for HClO detection in an actual water sample. Especially for developing countries, such a low-cost and highly sensitive detection method provides a simple and practical method for monitoring HClO in water.

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  1. 1.

    Aoki T, Munemori M. Continucus flow determination of free chlorine in water. Anal Chem. 1983;55(2):209–12.

    CAS  Article  Google Scholar 

  2. 2.

    Yang YM, Zhao Q, Feng W, Li FY. Luminescent chemodosimeters for bioimaging. Chem Rev. 2013;113(1):192–270.

    CAS  Article  Google Scholar 

  3. 3.

    Guo J, Zhang ZX, Kuai ZY, Wang R, Yang QB, Shan YM, Li YX. A new turn-on fluorescent probe towards hypochlorite in living cells. Anal Methods. 2017;9(5):864–70.

    CAS  Article  Google Scholar 

  4. 4.

    Chen GW, Song FL, Wang JY, Yang ZG, Sun SG, Fan JL, Qiang XX, Wang X, Dou BR, Peng XJ. FRET spectral unmixing: FRET spectral unmixing:a ratiometric fluorescent nanoprobe for hypochlorite. Chem Commun. 2012;48(24):2949–51.

    CAS  Article  Google Scholar 

  5. 5.

    Yap YW, Whiteman M, Cheung NS. Chlorinative stress: an under appreciated mediator of neurodegeneration? Cell Signal. 2007;19(2):219–28.

    CAS  Article  Google Scholar 

  6. 6.

    Li DW, Sun JJ, Gan ZF, Chen HY, Guo D. Reaction-based SERS nanosensor for monitoring and imaging the endogenous hypochlorous acid in living cells. Anal Chim Acta. 2018;1018(14):104–10.

    CAS  Article  Google Scholar 

  7. 7.

    Klebanoff SJ. Myeloperoxidase: friend and foe. J Leukocyte Biol. 2005;77(5):598–625.

    CAS  Article  Google Scholar 

  8. 8.

    Pattison DI, Davies MJ. Absolute rate constants for the reaction of hypochlorous acid with protein side chains and peptide bonds. Chem Res Toxicol. 2001;14(10):1453–64.

    CAS  Article  Google Scholar 

  9. 9.

    Kettle AJ, Winterbourn CC. Myeloperoxidase: a key regulator of neutrophil oxidant production. Redox Rep. 1997;3(1):3–15.

    CAS  Article  Google Scholar 

  10. 10.

    Iwase H. Routine high-performance liquid chromatographic determination of ascorbic acid in foods using l-methionine for the pre-analysis sample stabilization. Talanta. 2003;60(5):1011–21.

    CAS  Article  Google Scholar 

  11. 11.

    Jiang Y, Zheng G, Cai N, Zhang H, Tan Y, Huang M, He Y, He J, Sun H. A fast-respone fluorescent probe for hypochlorous acid detection and its application in exogenous and endogenous HOCl imaging of living cells. Chem Commun. 2017;53(91):12349–52.

    CAS  Article  Google Scholar 

  12. 12.

    Li HY, Ma HM. New progress in spectroscopic probes for reactive oxygen species. J Anal Test. 2018;2(1):2–19.

    Article  Google Scholar 

  13. 13.

    Gallina A, Pastore P, Magno F. The use of nitrite ion in the chromatographic determination of large amounts of hypochlorite ion and of traces of chlorite and chlorate ions. Analyst. 1999;124(10):1439–42.

    CAS  Article  Google Scholar 

  14. 14.

    Xue MG, Wang H, Chen J, Ren JY, Shu C, Yang HP, Zeng RJ, Long YF, Zhang PS. Ratiometric fluorescent sensing of endogenous hypochlorous acid in lysosomes using AIE-based polymeric nanoprobe. Sensor Actuat B-Chem. 2019;282(1):1–8.

    CAS  Article  Google Scholar 

  15. 15.

    Lou XD, Zhang Y, Qin JG, Li Z. Colormetric hypochlorite detection using an azobenzene acid in pure aqueous solutions and real application in tap water. Sensor Actuat B-Chem. 2012;161(1):229–34.

    CAS  Article  Google Scholar 

  16. 16.

    Zhang J, Wang XL, Yang XR. Colorimetric determination of hypochlorite with unmodified gold nanoparticles through the oxidation of a stabilizer thiol compound. Analyst. 2012;137(12):2806–12.

    CAS  Article  Google Scholar 

  17. 17.

    Zhang J, Yang XR. A simple yet effective chromogenic reagent for the rapid estimation of bromate and hypochlorite in drinking water. Analyst. 2013;138(2):434–7.

    CAS  Article  Google Scholar 

  18. 18.

    Zhu JG, Liu SP, Liu ZF, Li YF, Qiao M, Hu XL. Enhanced spectrofluorimetric determination of hypochlorite based on the catalytic oxidation of thiamine to thiochrome in the presence of trace ferrocyanide. RSC Adv. 2014;4(12):5990–4.

    CAS  Article  Google Scholar 

  19. 19.

    Hao YQ, Chen WS, Wang LQ, Zhou BB, Zang QG, Chen S, Lou YN. A naphthalimide-based azo colorimetric and ratiometric probe: synthesis and its application in rapid detection of cyanide anions. Anal Methods. 2014;6(8):2478–83.

    CAS  Article  Google Scholar 

  20. 20.

    Sumriddetchkajorn S, Chaitavon K, Intaravanne Y. Mobile device-based self-referencing colorimeter for monitoring chlorine concentration in water. Sensor Actuat B-Chem. 2013;182:592–7.

    CAS  Article  Google Scholar 

  21. 21.

    Zeng XL, Hu J, Zhang M, Wang FL, Wu L, Hou XD. Visual detection of fluoride anions using mixed lanthanide metal-organic frameworks with a smartphone. Anal Chem. 2019;92(2):2097–102.

    Article  Google Scholar 

  22. 22.

    Kumar RS, Kumar SKA, Vijayakrishna K, Sivaramakrishna A, Brahmmananda Rao CVS, Sivaraman N, Sivaraman N, Sahoo SK. Development of the smartphone-assisted colorimetric detection of thorium by using new schiff’s base and its applications to real time samples. Inorg Chem. 2018;57(24):15270–9.

    Article  Google Scholar 

  23. 23.

    Manna A, Goswami S. Ratiometric detection of hypochlorite applying the restriction to 2-way ESIPT: simple design for “naked-eye” tap water analysis. New J Chem. 2015;39(6):4424–9.

    CAS  Article  Google Scholar 

  24. 24.

    Wei P, Liu LY, Wen Y, Zhao GL, Xue FF, Yuan W, Li RH, Zhong YP, Zhang MF, Yi T. Release of amino- or carboxy-containing compounds triggered by HOCl: application for imaging and drug design. Angew Chem. 2019;58(14):4547–51.

    CAS  Article  Google Scholar 

  25. 25.

    Wei P, Yuan W, Xue FF, Zhou W, Li RH, Zhang DT, Yi T. Deformylation reaction-based probe for in vivo imaging of HOCl. Chem Sci. 2018;9(2):495–501.

    CAS  Article  Google Scholar 

  26. 26.

    Sun ZN, Liu FQ, Chen Y, Tam PKH, Yang D. A Highly specific BODIPY-based fluorescent probe for the detection of hypochlorous acid. Org Lett. 2008;10(11):2171–4.

    CAS  Article  Google Scholar 

  27. 27.

    Hu JJ, Wong N-K, Gu Q, Bai X, Ye S, Yang D. HKOCl-2 series of green BODIPY-based fluorescent probes for hypochlorous acid detection and imaging in live cells. Org Lett. 2014;16(13):3544–7.

    CAS  Article  Google Scholar 

  28. 28.

    Pan H, Liu YJ, Liu SZ, Ou ZP, Chen HB, Li HM. A dual-function colorimetric probe based on carbazole-cyanine dyad for highly sensitive recognition of cyanide and hypochlorous acid in aqueous media. Talanta. 2019;202(1):329–35.

    CAS  Article  Google Scholar 

  29. 29.

    Li DX, Feng Y, Lin JZ, Chen M, Wang SX, Wang X, Sheng HT, Shao SL, Zhu MZ, Meng XM. A mitochondria-targeted two-photon fluorescent probe for highly selective and rapid detection of hypochlorite and its bio-imaging in living cells. Sensor Actuat B-Chem. 2016;222:483–91.

    CAS  Article  Google Scholar 

  30. 30.

    Dong H, Zhou YL, Zhao L, Hao YQ, Zhang YT, Ye BX, Xu MT. Dual-response ratiometric electrochemical microsensor for effective simultaneous monitoring of hypochlorous acid and ascorbic acid in human body fluids. Anal Chem. 2020;92(22):15079–86.

    CAS  Article  Google Scholar 

  31. 31.

    Gong YJ, Lv MK, Zhang ML, Kong ZZ, Mao GJ. A novel two-photon fluorescent probe with long-wavelength emission for monitoring HClO in living cells and tissues. Talanta. 2019;192(15):128–34.

    CAS  Article  Google Scholar 

  32. 32.

    Yuan L, Lin WY, Xie YN, Chen B, Song JZ. Fluorescent detection of hypochlorous acid from turn-on to FRET-based ratiometry by a HOCl-mediated cyclization reaction. Chem Eur J. 2012;18(9):2700–6.

    CAS  Article  Google Scholar 

  33. 33.

    Chen SM, Lu JX, Sun CD, Ma HM. A highly specific ferrocene-based fluorescent probe for hypochlorous acid and its application to cell imaging. Analyst. 2010;135(3):577–82.

    CAS  Article  Google Scholar 

  34. 34.

    Zou XM, Liu YJ, Zhu XJ, Chen M, Yao LM, Feng W, Li FY. An Nd3+-sensitized upconversion nanophosphor modified with a cyanine dye for the ratiometric upconversion luminescence bioimaging of hypochlorite. Nanoscale. 2015;7(9):4105–13.

    CAS  Article  Google Scholar 

  35. 35.

    Yue YK, Huo FJ, Yin CX, Chao JB, Zhang YB, Wei X. An ICT based ultraselective and sensitive fluorescent probe for detection of HClO in living cells. RSC Adv. 2015;5(95):77670–2.

    CAS  Article  Google Scholar 

  36. 36.

    Goswami S, Das AK, Manna A, Maity AK, Saha P, Quah CK, Fun HK, Abdel-Aziz HA. Nanomolar detection of hypochlorite by a rhodamine-based chiral hydrazide in absolute aqueous media: application in tap water analysis with live-cell imaging. Anal Chem. 2014;86(13):6315–22.

    CAS  Article  Google Scholar 

  37. 37.

    Fan J, Mu H, Zhu H, Zhu H, Du J, Jiang N, Wang J, Peng X. Recognition of HClO in live cells with separate signals using a ratiometric fluorescent sensor with fast response. Ind Eng Chem Res. 2015;54(36):8842–6.

    CAS  Article  Google Scholar 

  38. 38.

    Xiao HD, Li JH, Zhao J, Yin G, Quan YW, Wang J, Wang RY. A colorimetric and ratiometric fluorescent probe for ClO- targeting in mitochondria and its application in vivo. J Mater Chem B. 2015;3(8):1633–8.

    CAS  Article  Google Scholar 

  39. 39.

    Zhang J, Wang X, Yang X. Colorimetric determination of hypochlorite with unmodified gold nanoparticles through the oxidmion of a stabilizer thiol compound. Analyst. 2012;137(12):2806–12.

    CAS  Article  Google Scholar 

  40. 40.

    GB 5749–2006, Set by the standardization a dministration of the people’s republic of China

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We are grateful to the National Natural Science Foundation of China (Nos. U1404215, 22074089, 21804085, 21675109), Innovation Scientists and Technicians Troop Construction Projects of Henan Province (No: 41) and Key Scientific Research Projects of Colleges and Universities in Henan Province (No: 21A150043) for support.

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Correspondence to Hui Dong or Guoqing Xiao or Maotian Xu.

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He, S., Dong, H., Hao, Y. et al. Quantifying Hypochlorous Acid Concentration in Environmental Water Using Smartphone Colorimetry. J. Anal. Test. (2021). https://doi.org/10.1007/s41664-021-00156-1

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  • Smartphone colorimetry
  • RGB
  • Test strips
  • Hypochlorite acid
  • Methylene blue