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A Novel Method to Extract Juglone from Juglans mandshurica Waste Branches Using a Water-in-Oil Microemulsion


Pruning of Juglans mandshurica produces many branches, which are potentially a rich source of juglone although they are usually discarded as waste. Herein, we describe a water-in-oil microemulsion-based microwave-assisted extraction (MBMAE) method to isolate the juglone found in waste branches from J. mandshurica. In our experiment, J. mandshurica waste branch powder was added to the MBMAE system. Using the optimized microemulsion system comprising Tween 80:n-propanol:n-hexane:water at 27:13.5:4.5:55 (w/w/w/w), and a pH of 5.6, at an operating temperature of 40 °C, operating time of 63 s, and operating power of 400 W, the juglone yield was 4.58 mg/g from a ratio of microemulsion to branch power of 20:1 (mL/g). The juglone extraction yield using the MBMAE method was 1.86- and 6.65-fold better than that obtained from microwave-assisted extraction by ethanol (Ethanol-MAE) and heat reflux extraction by ethanol (Ethanol-HRE), respectively. The MBMAE method may be useful as an alternative to traditional extraction methods to isolate juglone.

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  1. 1.

    Wang, R.J., Wang, S., Xia, Y.J., Tu, M.W., Zhang, L.J., Wang, Y.M.: Antitumor effects and immune regulation activities of a purified polysaccharide extracted from Juglan regia. Int. J. Biol. Macromol. 72, 771–775 (2015)

    Article  Google Scholar 

  2. 2.

    Zhang, Y.W., Lin, H., Li, S.S., Chen, J.B., Sun, Y.S., Li, Y.X.: High-speed counter-current chromatography assisted preparative isolation of bioactive compounds from stem bark of Juglans mandshurica. J. Sep. Sci. 40(3), 767–778 (2017)

    Article  Google Scholar 

  3. 3.

    Lou, L.L., Cheng, Z.Y., Guo, R., Yao, G.D., Song, S.J.: Alkaloids from Juglans mandshurica maxim induce distinctive cell death in hepatocellular carcinoma cells. Nat. Prod. Res. 33(6), 911–914 (2019)

    Article  Google Scholar 

  4. 4.

    Yao, D.L., Zhang, C.H., Luo, J., Jin, M., Zheng, M.S., Cui, J.M., Son, J.K., Li, G.: Chemical constituents from the leaves of Juglans mandshurica. Arch. Pharm. Res. 38(4), 480–484 (2015)

    Article  Google Scholar 

  5. 5.

    Zhou, Y.Y., Gao, H.R., Song, H.J., Ma, D.D., Zhang, X.J., Sun, Y.P., Liu, Y., Wang, X.L., Yang, B.Y., Kuang, H.X.: Two new dammarane-type triterpenoids from the green walnut husks of Juglans mandshurica. J. Nat. Prod. Res. 15, 1–8 (2020)

    Google Scholar 

  6. 6.

    Zhu, X.D., Liu, Y., Wang, W.C., Dong, N.W., Zheng, S.Y.: Effect of clear painting on surface color change of thermochromic wood veneers. J. Northwest For. Univ. 34(4), 223–229 (2019)

    Google Scholar 

  7. 7.

    Shigaeva, J., Darr, D.: On the socio-economic importance of natural and planted walnut (Juglans regia L.) forests in the Silk Road countries: a systematic review. For. Policy Econ. 118, 102233–102249 (2020)

    Article  Google Scholar 

  8. 8.

    Zhang, Y.W., Lin, H., Bao, Y.L., Wu, Y., Yu, C.L., Huang, Y.X., Li, Y.X.: A new triterpenoid and other constituents from the stem bark of Juglans mandshurica. J. Biochem. Syst. Ecol. 44, 136–140 (2012)

    Article  Google Scholar 

  9. 9.

    Park, S.J., Kim, N.Y., Yoo, G.J., Kim, S.N., Kwon, H.J., Jung, K.W., Oh, D.C., Lee, Y.H., Kim, S.H.: Phenolics and neolignans isolated from the fruits of Juglans mandshurica and their effects on lipolysis in adipocytes. Phytochemistry 137, 87–93 (2017)

    Article  Google Scholar 

  10. 10.

    Daneiva, C., David, R., Yanet, O., Klaus, M., Luis, A.F.: A promising naphthoquinone [8-hydroxy-2-(2-thienylcarbonyl)naphtho[2,3-b]thiophene-4,9-dione] exerts anti-colorectal cancer activity through ferroptosis and inhibition of MAPK signaling pathway based on RNA sequencing. Open Chem. 18(1), 1242–1255 (2020)

    Article  Google Scholar 

  11. 11.

    Jiménez-Brenes, F.M., López-Granados, F., Castro, A.I., Torres-Sánchez, J., Serrano, N., Peña, J.M.: Quantifying pruning impacts on olive tree architecture and annual canopy growth by using UAV-based 3D modelling. J. Plant Methods 13(1), 55–70 (2017)

    Article  Google Scholar 

  12. 12.

    Ersin, A.: A new insight into pruning strategy in the biennial cycle of fruiting: vegetative growth at shoot and whole-tree level, yield and fruit quality of apple. Not. Bot. Horti Agrobot. Cluj-Napoca 45, 232–237 (2017)

    Article  Google Scholar 

  13. 13.

    Mohammadi, A., Mahmoudi, M.J., Rezaee, R.: Vegetative and reproductive responses of some apple cultivars (Malus domestica Borkh.) to heading back pruning. Int. J. AgriSci. 3, 628–635 (2013)

    Google Scholar 

  14. 14.

    Cosmulescu, S., Trandafir, I., Achim, G., Bout, M., Bacit, A., Gruit, M.: Phenolics of green husk in mature Juglans fruits. Not. Bot. Horti Agrobot. Cluj-Napoca 38(1), 53–56 (2010)

    Google Scholar 

  15. 15.

    Stampar, F., Solar, A., Hudina, M.: Traditional Juglans liqueur—cocktail of phenolics. Food Chem. 95, 627–631 (2006)

    Article  Google Scholar 

  16. 16.

    Gökalp, F.: The chemical activity of Juglone in different phases as a protective agent. J. Fluor. Chem. 242, 109701–109704 (2021)

    Article  Google Scholar 

  17. 17.

    Kao, C.C., Kung, P.H., Tai, C.J., Tsai, M.C., Cheng, Y.B., Wu, C.C.: Juglone prevents human platelet aggregation through inhibiting Akt and protein disulfide isomerase. J. Phytomed. 82, 153449–153449 (2020)

    Article  Google Scholar 

  18. 18.

    Wu, C.Y., Wang, H., Fan, X.H., Yue, W., Wu, Q.N.: Waste Euryale ferox Salisb. leaves as a potential source of anthocyanins: extraction optimization, identification and antioxidant activities evaluation. Waste Biomass Valoriz. 11, 4327–4340 (2020)

    Article  Google Scholar 

  19. 19.

    Orioli, M.L., Regazzoni, F., Saligari, C., Marinello, G., Rossoni, G., Aldini, M.: CariniCoffee silver skins as a source of polyphenols: high resolution mass spectrometric profiling of components and antioxidant activity. J. Funct. Foods 20, 472–485 (2016)

    Article  Google Scholar 

  20. 20.

    Gîrzu, M., Fraisse, D., Carnat, A.P., Carnat, A., Lamaison, J.L.: High-performance liquid chromatographic method for the determination of juglone in fresh walnut leaves. J. Chromatogr. A 805(1–2), 315–318 (1998)

    Google Scholar 

  21. 21.

    Laroze, L., Soto, C., Zúñiga, M.E.: Phenolic antioxidants extraction from raspberry waste assisted by-enzymes. Electron. J. Biotechnol. 13(6), 11–12 (2010)

    Article  Google Scholar 

  22. 22.

    Nima, R., Foad, R., Mashallah, R., Mohammad, Y.: Juglone extraction from walnut (Juglans regia L.) green husk by supercritical CO2: Process optimization using Taguchi method. J. Environ. Chem. Eng. 8(3), 103776 (2020)

    Article  Google Scholar 

  23. 23.

    Yong, N., Bi, Z.H., Su, H., Yan, L.F.: Deep eutectic solvent (DES) as both solvent and catalyst for oxidation of furfural to maleic acid and fumaric acid. Green Chem. 21, 1075–1079 (2019)

    Article  Google Scholar 

  24. 24.

    Cui, Q., Liu, J.Z., Xu, W.J., Kang, Y.F., Wang, X.Y., Li, Y.Y., Fu, Y.J.: Enhanced extraction and preconcentration of main target saponins from Panax notoginseng root using green and efficient formulated surfactant aqueous systems. J. Clean. Prod. 186, 297–308 (2019)

    Google Scholar 

  25. 25.

    Joanna, N., Kaveh, S., Marliya, I., Mohammed, M.F.: Approach for polygodial extraction from Pseudowintera colorata (Horopito) leaves using deep eutectic solvents. ACS Sustain. Chem. Eng. 6, 862–871 (2018)

    Article  Google Scholar 

  26. 26.

    Douglas, G.H., Ran, Y., Rachel, N.D., Matthew, J.C., Sai, V.P., Hugh, M.O., Volker, S.U.: Protein extraction into the bicontinuous microemulsion phase of a water/SDS/pentanol/dodecane winsor-III system: effect on nanostructure and protein conformation. J. Colloids Surf. B 160, 144–153 (2017)

    Article  Google Scholar 

  27. 27.

    Mehdi, J.J., Soleiman, A., Martin, G.S.: Microemulsion as nanoreactor for lutein extraction: optimization for ultrasound pretreatment. J. Food Biochem. 43(8), 12929–12941 (2019)

    Google Scholar 

  28. 28.

    Atefeh, A.R., Soleiman, A.: Extraction of lycopene using a lecithin-based olive oil microemulsion. Food Chem. 272, 568–573 (2019)

    Article  Google Scholar 

  29. 29.

    Liu, F., Yang, Y., Lu, Y., Shang, K., Lu, W., Zhao, X.: Extraction of germanium by the AOT microemulsion with N235 System. Ind. Eng. Chem. Res. 49, 10005–10008 (2010)

    Article  Google Scholar 

  30. 30.

    Guo, Y., Li, H.Y., Yuan, Y.H., Huang, J., Diao, J., Xie, B.: ME: an efficient way for simultaneous detoxification and resource recovery of hazardous wastewater containing V(V) and Cr(VI). J. Hazard. Mater. 386, 121948–121959 (2020)

    Article  Google Scholar 

  31. 31.

    Jalali, J.M., Abbasi, S.: Novel approach for lutein extraction: food grade microemulsion containing soy lecithin & sunflower oil. J. Innov. Food Sci. Emerg. Technol. 66, 102505–102512 (2020)

    Article  Google Scholar 

  32. 32.

    Yang, X.Y., Jie, F.P., Wang, B.J., Bai, Z.S.: High-efficient synergistic extraction of Co(II) and Mn(II) from wastewater via novel microemulsion and annular centrifugal extractor. J. Sep. Purif. Technol. 209, 997–1006 (2018)

    Article  Google Scholar 

  33. 33.

    Aramburu, A., Bonifazi, E.L., Gerschenson, L.N., Rojas, A.M., Basanta, M.F.: Sweet cherry (Prunus avium) fibers extracted from microwave- and steam-blanched recovered fruits: photo-antioxidant activity in milk proteins. Bioact. Carbohydr. Diet. Fibre 24, 100247–100258 (2020)

    Article  Google Scholar 

  34. 34.

    Guo, Y., Li, H.Y., Shen, S., Wang, C.J., Diao, J., Xie, B.: Recovery of vanadium from vanadium slag with high phosphorus content via recyclable ME. Hydrometallurgy 198, 105509–105519 (2020)

    Article  Google Scholar 

  35. 35.

    Imen, B.A., Georgia, A., Ioanna, N., Maria, A., Moncef, N., Tahar, M., Gerasimos, L.: On the evaluation of different saccharification schemes for enhanced bioethanol production from potato peels waste via a newly isolated yeast strain of Wickerhamomyces anomalus. Bioresour. Technol. 289, 121614–121623 (2019)

    Article  Google Scholar 

  36. 36.

    Kadouche, S., Farhat, M., Lounici, H., Fiallo, M., Sharrock, P., Mecherri, M., Hadioui, M.: Low cost chitosan biopolymer for environmental use made from abundant shrimp wastes. Waste Biomass Valoriz. 8(2), 401–406 (2017)

    Article  Google Scholar 

  37. 37.

    Hajimohammadi, A., van Deventer, J.S.J.: Solid reactant-based geopolymers from rice hull ash and sodium aluminate. Waste Biomass Valoriz. 8(6), 2131–2140 (2017)

    Article  Google Scholar 

  38. 38.

    Ma, L., Zhang, Y.L., Han, J.Q., Zhou, X.Y., Xia, S.S.: Separation and purification of juglone from walnut green peel by macroporous resin. Food Ferment. Ind. 42(01), 108–113 (2016)

    Google Scholar 

  39. 39.

    Lawrence, J.: Surfactant systems: microemulsions and vesicles as vehicles for drug delivery. Eur. J. Drug Metab. Pharmacokinet. 19(3), 257–269 (1994)

    Article  Google Scholar 

  40. 40.

    Du, Y.H., Jia, R.Y., Yin, Z.Q.: Acaricidal activity of extracts of neem (Azadirachta indica) oil against the larvae of the rabbit mite Sarcoptes scabiei var. cuniculi in vitro. Vet. Parasitol. 157(1–2), 144–148 (2008)

    Article  Google Scholar 

  41. 41.

    Yuan, Y., Li, S.M., Mo, F.K.: Investigation of microemulsion system for transdermal delivery of meloxicam. Int. J. Pharm. 32(1), 117–123 (2006)

    Article  Google Scholar 

  42. 42.

    Silva Camilo, J.V., Martelloti, R.R., Costa, J.A., Mansur, C.R.E.: Development of oil-in-water microemulsions and evaluation of its presence in the treatment of produced water. J. Nanosci. Nanotechnol. 19(8), 8143–8150 (2019)

    Article  Google Scholar 

  43. 43.

    Liu, W.L., Liu, C.M., Liu, W., Chen, H.L., Liu, J.H.: Study on preparation of O/W MCFA micro-emulsion and its characteristics. Food Sci. 30(6), 102–105 (2009)

    Google Scholar 

  44. 44.

    Brown, D.G., Jaffé, P.R.: Effects of nonionic surfactants on the cell surface hydrophobicity and apparent hamaker constant of a Sphingomonas sp. Environ. Sci. Technol. 40(1), 195–201 (2006)

    Article  Google Scholar 

  45. 45.

    Zhu, F.S., Yang, X.Q., Bai, L.: Effect of cosurfactant on phase behavior in acetochlor microemulsion. Fine Chem. 27(7), 646-649+677 (2010)

    Google Scholar 

  46. 46.

    Wang, L.Z., Yan, W.R., Tian, W.R., Xue, H.H., Tang, J.K., Zhang, L.W.: Self-microemulsifying drug delivery system of phillygenin: formulation development, characterization and pharmacokinetic evaluation. Pharmaceutics 12(2), 130 (2020)

    Article  Google Scholar 

  47. 47.

    Candioti, L.V., De Zan, M.M., Cámara, M.S., Goicoechea, H.C.: Experimental design and multiple response optimization. Using the desirability function in analytical methods development. Talanta 15(124), 123–138 (2014)

    Article  Google Scholar 

  48. 48.

    Sun, H.Y., Li, C.Y., Ni, Y.J., Yao, L.P., Jiang, H.W., Ren, X.T., Fu, Y.J., Zhao, C.J.: Ultrasonic/microwave-assisted extraction of polysaccharides from Camptotheca acuminata fruits and its antitumor activity. Carbohydr. Polym. 15(206), 557–564 (2019)

    Article  Google Scholar 

  49. 49.

    Zhang, S., Feng, L.L., Peng, X.X., Mao, M.M., Chi, Y., Wang, F.: Effect of sludge pellets addition on combustion characteristics and ash behaviour of municipal solid waste. Waste Biomass Valoriz. 11(10), 5351–5361 (2020)

    Article  Google Scholar 

  50. 50.

    Ebrahimi, I., Gashti, M.P.: Extraction of juglone from Pterocarya fraxinifolia leaves for dyeing, anti-fungal finishing, and solar UV protection of wool. Color. Technol. 131(6), 451–457 (2016)

    Article  Google Scholar 

  51. 51.

    Elavarasan, S., Gopalakrishnan, M.: Synthesis, structural analysis, theoretical studies of some lawsone derivatives. Spectrochim. Acta A 133(10), 1–6 (2014)

    Article  Google Scholar 

  52. 52.

    Singh, D.K., Luqman, S., Mathur, A.K.: Lawsonia inermis L.—a commercially important primaeval dying and medicinal plant with diverse pharmacological activity: a review. Ind. Crops Prod. 65, 269–286 (2015)

    Article  Google Scholar 

  53. 53.

    Luo, X.: Infrared and THz Spectroscopy Study of Naphthoquinone and Its Derivatives. Capital Normal University, Beijing (2008)

    Google Scholar 

  54. 54.

    Izadyar, E., Mazeyar, P.G.: Extraction of polyphenolic dyes from henna, pomegranate rind, and Pterocarya fraxinifolia for nylon 6 dyeing. Color Technol. 132(2), 162–176 (2016)

    Article  Google Scholar 

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We thank Renee Mosi, PhD, from Liwen Bianji, Edanz Editing China (, for editing the English text of a draft of this manuscript.


This work was financially supported by the Fundamental Research Fund for Central Universities (2572019CZ01), the 111 Project of China (B20088) and the Heilongjiang Touyan Innovation Team Program (Tree Genetics and Breeding Innovation Team).

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CZ and CL conceived the underlying ideas of the study, XM and MT contributed to the practical aspects of the research work, SL, YT and TW analyzed the data, and CL, XM and MT wrote the manuscript.

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Correspondence to Chunjian Zhao.

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Li, C., Meng, X., Tian, M. et al. A Novel Method to Extract Juglone from Juglans mandshurica Waste Branches Using a Water-in-Oil Microemulsion. Waste Biomass Valor (2021).

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  • Juglans mandshurica
  • Juglone
  • Waste branches
  • Microemulsion