Phytochemical analysis of Juglans regia oil and kernel exploring their antinociceptive and anti-inflammatory potentials utilizing combined bio-guided GC-FID, GC-MS and HPLC analyses


Juglans regia L., Juglandaceae, is broadly used due to its immunomodulatory effects, potentials in protecting against many sever-disorders, and high safety-profile. The aim of this work is to make a phytochemical analysis of J. regia oil and kernel exploring their antinociceptive and anti-inflammatory potentials utilizing combined bio-guided gas chromatography with mass spectrometer (GC-MS), gas chromatography with flame ionization detection (GC-FID) and reversed-phase high-performance liquid chromatography (RP-HPLC) analyses. Combined bio-guided GC-MS, GC-FID and RP-HPLC analyses is an innovative-combined-technique aiming at efficiently analyzing various-extracts phytochemical and biological characters. The J. regia oil and kernel ethyl-acetate extract were monitored during exploring their possible acute-antinflammatory, antidiabetic and antidiabetic-neuropathy. Glycated-hemoglobin, serum-insulin, serum-catalase and lipid-peroxidation levels have been also monitored. Combined bio-guided GC-FID, GC-MS and HPLC analyses have shown to be an efficient analyzing-method through identifying the most active compound, linoleic acid. Linoleic acid has shown the highest improvement of the acute inflammatory-pain, chronic blood-glucose level reduction, serum-insulin elevation, and normalization of glycated-hemoglobin levels. J. regia oil has shown more lipid-peroxidation reduction, while kernel ethyl-acetate extract has shown more acute-blood-glucose level reduction and serum-catalase levels elevation. Compared to tramadol, the highest-doses of J. regia oil, kernel ethyl-acetate extract, and linoleic acid have shown higher antinociceptive-potentials in amelioration of thermal-hyperalgesic and anti-allodynic neuropathic-pain. Thus, the antinflammatory, the reduction of oxidative-stress, and the insulin-secretagogue potentials might be among the possible mechanisms of improvement of neuropathic-pain. In correlation to conventional-techniques, the combined bio-guided analyses have shown to be an efficient innovative-combined technique. After further clinical studies, J. regia might be utilized as a possible-remedy for various painful-syndromes.


  1. AOAC, 1998. Fatty acids in oils and fats. In: Preparation of Methyl Esters, 16th ed. AOAC International, Gaithersburg.

    Google Scholar 

  2. Bakirel, T., Bakirel, U., Keles, O.U., Ulgen, S.G., Yardibi, H., 2008. In vivo assessment of antidiabetic and antioxidant activities of rosemary (Rosmarinus officinalis) in alloxan-diabetic rabbits. J. Ethnopharmacol. 116, 64–73.

    Article  PubMed Central  Google Scholar 

  3. Boulton, A.J., Malik, R.A., Arezzo, J.C., Sosenko, J.M., 2004. Diabetic somatic neuropathies. Diabetes Care 27, 1458–1486.

    Article  Google Scholar 

  4. Galer, B.S., Gianas, A., Jensen, M.P., 2000. Painful diabetic polyneuropathy: epidemiology, pain description, and quality of life. Diabetes Res. Clin. Pr. 47, 123–128.

    CAS  Article  Google Scholar 

  5. Gardmark, M., Hoglund, A.U., Hammarlund-Udenaes, M., 1998. Aspects on tail-flick, hot-plate and electrical stimulation tests for morphine antinociception. Pharmacol. Toxicol. 83, 252–258.

    CAS  Article  PubMed Central  Google Scholar 

  6. Hayes, D., Angove, M.J., Tucci, J., Dennis, C., 2016. Walnuts (Juglans regia) chemical composition and research in human health. Crit. Rev. Food Sci. Nutr. 56, 1231–1241.

    CAS  Article  PubMed Central  Google Scholar 

  7. Hosseini, S., Huseini, H.F., Larijani, B., Mohammad, K., Najmizadeh, A., Nourijelyani, K., Jamshidi, L., 2014. The hypoglycemic effect of Juglans regia leaves aqueous extract in diabetic patients: a first human trial. Daru 22,

  8. Koleilat, M., Raafat, K., El-Lakany, A., Aboul-Ela, M., 2017. Designing monographs for Rosmarinus officinalis L. and Lavandula angustifolia L: two Lebanese species with significant medicinal potentials. Pharmacogn. J. 9, 452–474.

    CAS  Article  Google Scholar 

  9. Mathias, E.V., Halkar, U.P., 2004. Separation and characterization of lignin compounds from the walnut (Juglans regia) shell oil using preparative TLC, GC-MS and 1H NMR. J. Anal. Appl. Pyrol. 71, 515–524.

    CAS  Article  Google Scholar 

  10. Micov, A., Tomic, M., Pecikoza, U., Ugresic, N., Stepanovic-Petrovic, R., 2015. Levetiracetam synergises with common analgesics in producing antinociception in a mouse model of painful diabetic neuropathy. Pharmacol. Res. 97, 131–142.

    CAS  Article  PubMed Central  Google Scholar 

  11. Nasiry, D., Khalatbary, A.R., Ahmadvand, H., Talebpour Amiri, F., Akbari, E., 2017. Protective effects of methanolic extract of Juglans regia L. leaf on streptozotocin-induced diabetic peripheral neuropathy in rats. BMC Complement. Altern. Med. 17,

  12. Ohkawa, H., Ohishi, N., Yagi, K., 1979. Assay for lipid peroxides in animal tissues by thiobarbituric acid reaction. Anal. Biochem. 95, 351–358.

    CAS  Article  PubMed Central  Google Scholar 

  13. Ohsawa, M., Aasato, M., Hayashi, S.S., Kamei, J., 2011. RhoA/Rho kinase pathway contributes to the pathogenesis of thermal hyperalgesia in diabetic mice. Pain 152, 114–122.

    CAS  Article  PubMed Central  Google Scholar 

  14. Pan, A., Sun, Q., Manson, J.E., Willett, W.C., Hu, F.B., 2013. Walnut consumption is associated with lower risk of type 2 diabetes in women. J. Nutr. 143, 512–518.

    CAS  Article  PubMed Central  Google Scholar 

  15. Paudel, P., Satyal, P., Dosoky, N.S., Maharjan, S., Setzer, W.N., 2013. Juglans regia and J. nigra, two trees important in traditional medicine: A comparison of leaf essential oil compositions and biological activities. Nat. Prod. Commun. 8, 1481–1486.

    CAS  PubMed  PubMed Central  Google Scholar 

  16. Raafat, K., El-Darra, N., Saleh, F.A., Rajha, H.N., Maroun, R.G., Louka, N., 2018. Infrared-assisted extraction and HPLC-analysis of Prunus armeniaca L. pomace and detoxified-kernel and their antidiabetic effects. Phytochem. Anal. 29, 156–167.

    CAS  Article  PubMed Central  Google Scholar 

  17. Raafat, K., El-Haj, R., Shoumar, D., Alaaeddine, R., Fakhro, Y., Tawil, N., Shaer, F., Daher, A., Awada, N., Sabra, A., Atwi, K., Khaled, M., Messi, R., Abouzaher, N., Houri, M., AlJallad, S., 2017. Neuropathic pain: literature review and recommendations of potential phytotherapies. Pharmacogn. J. 9, 425–434.

    CAS  Article  Google Scholar 

  18. Salama, R.A.M., El Gayar, N.H., Georgy, S.S., Hamza, M., 2016. Equivalent intraperitoneal doses of ibuprofen supplemented in drinking water or in diet: a behavioral and biochemical assay using antinociceptive and thromboxane inhibitory dose-response curves in mice. Peer J 4, e2239.

    Article  PubMed Central  Google Scholar 

  19. Sullivan, K.A., Hayes, J.M., Wiggin, T.D., Backus, C., Su Oh, S., Lentz, S.I., Brosius 3rd, F., Feldman, E.L., 2007. Mouse models of diabetic neuropathy. Neurobiol. Dis. 28, 276–285.

    CAS  Article  PubMed Central  Google Scholar 

  20. Tesfaye, S., 2013. Impact of Painful Diabetic Polyneuropathy on Patients. Springer, New York.

    Google Scholar 

  21. Wallace, M.S., Marcotte, T.D., Umlauf, A., Gouaux, B., Atkinson, J.H., 2015. Efficacy of inhaled cannabis on painful diabetic neuropathy. J. Pain 16, 616–627.

    CAS  Article  PubMed Central  Google Scholar 

  22. Willoughby, D.A., DiRosa, M., 1972. Studies on the mode of action of non-steroid antiinflammatory drugs. Ann. Rheum. Dis. 31, 540.

    CAS  Article  PubMed Central  Google Scholar 

  23. Wu, Q.L., Liang, X.C., 2007. Survey of current experimental studies of effects of traditional Chinese compound recipe on diabetic peripheral neuropathy. Zhongguo Zhong Yao Za Zhi 32, 775–778.

    PubMed  PubMed Central  Google Scholar 

  24. Xu, H.B., Jiang, R.H., Chen, X.Z., Li, L., 2012. Chinese herbal medicine in treatment of diabetic peripheral neuropathy: a systematic review and meta-analysis. J. Ethnopharmacol. 143, 701–708.

    Article  PubMed Central  Google Scholar 

  25. Yasmineh, W.G., Kaur, T.P., Blazar, B.R., Theologides, A., 1995. Serum catalase as marker of graft-vs-host disease in allogeneic bone marrow transplant recipients: pilot study. Clin. Chem. 41, 1574–1580.

    CAS  Article  PubMed Central  Google Scholar 

Download references

Author information



Corresponding author

Correspondence to Karim Raafat.

Rights and permissions

This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Raafat, K. Phytochemical analysis of Juglans regia oil and kernel exploring their antinociceptive and anti-inflammatory potentials utilizing combined bio-guided GC-FID, GC-MS and HPLC analyses. Rev. Bras. Farmacogn. 28, 358–368 (2018).

Download citation


  • Bio-guided GC-FID
  • GC-MS
  • HPLC analyses
  • Oil analysis
  • Diabetes mellitus
  • Acute inflammation
  • Neuropathic pain