Elsevier

Food Chemistry

Volume 106, Issue 3, 1 February 2008, Pages 1014-1020
Food Chemistry

Walnut (Juglans regia) leaf extracts are strong scavengers of pro-oxidant reactive species

https://doi.org/10.1016/j.foodchem.2007.07.017Get rights and content

Abstract

The growing interest in the substitution of synthetic food antioxidants by natural ones has fostered research on vegetable sources and on the screening of raw materials, for identifying new antioxidants. In the present study, an ethanol:water (4:6) extract from Juglans regia leaves was evaluated for its putative in vitro scavenging effects on reactive oxygen species (ROS) [hydroxyl radical (HOradical dot), superoxide radical (O2-), peroxyl radical (ROOradical dot) and hydrogen peroxide (H2O2)] and reactive nitrogen species (RNS) [nitric oxide (radical dotNO) and peroxynitrite anion (ONOO)]. The extract presented a potent scavenging activity against all the reactive species tested, all the IC50s being found at the μg/mL level. IC50s (mean ± SE) for the ROS O2- and H2O2 were 47.6 ± 4.6, 383 ± 17 μg/mL, respectively. The oxygen radical absorbance capacity (ORAC) value obtained for ROOradical dot was 2.17 ± 0.22 μmol Trolox equivalents/mg extract. The IC50s for radical dotNO and ONOO were 1.95 ± 0.29 and 1.66 ± 0.10 μg/mL, respectively. The content of total phenolics was 270 ± 3 mg of gallic acid equivalents (GAE)/g of lyophilised extract.

These results showed that J. regia leaf extracts can be used as an easily accessible source of natural antioxidants.

Introduction

The prevention of the oxidative reactions in foods, pharmaceuticals and cosmetics and the management of oxidative stress-related diseases are some of the potential applications of antioxidants. The most widely used antioxidants, to prevent the oxidation of lipids in foods, are butylated hydroxyanisole (BHA), propyl gallate and 2-tert-butylhydroquinone (TBHQ) (Moure et al., 2001). However, there has been growing concern over the safety of some of the commercial antioxidants because several studies documented the mutagenesis and carcinogenesis potential associated with some synthetic antioxidants (Ito et al., 1986, Witschi, 1986), although other authors reported anti-carcinogenic effects (Williams, Iatropoulos, & Whysner, 1999). Therefore, the extraction and characterisation of natural antioxidants are of considerable interest. Natural antioxidants, such as tocopherols, ascorbic acid and plant polyphenols have gained increasing importance.

Walnut (Juglans regia L.) is a deciduous tree native in southeastern Europe of the Juglandaceae family. The walnut fruits are a highly nutritious food, which are rich in oil composed of unsaturated fatty acids. Antioxidant effects of isolated polyphenols obtained from walnuts have been previously reported (Fukuda, Ito, & Yoshida, 2003). Scavenging of hydroxyl radicals and superoxide radicals is documented for water and methanol extracts of the kernel of J. regia (Ohsugi et al., 1999). Walnut liqueur, obtained with green walnuts, also presents antioxidant activity which was correlated with its polyphenolic composition (Alamprese, Pompei, & Scaramuzzi, 2005). Walnut leaf has been widely used in folk medicine for the treatment of skin inflammations, hyperhidrosis and ulcers and for its antidiarriec, anti-helmintic, antiseptic and astringent properties (Bruneton, 1999, Proença da Cunha et al., 2003).

No studies concerning the scavenging effect of reactive oxygen species (ROS) or reactive nitrogen species (RNS) have been reported for J. regia leaves.

The objective of the present study is thus to assess the in vitro scavenging activity of pro-oxidant reactive species exerted by the J. regia leaf ethanol:water (4:6) extract, namely on the ROS [hydroxyl radical (HOradical dot), superoxide radical (O2-), peroxyl radical (ROOradical dot) and hydrogen peroxide (H2O2)] and the RNS [nitric oxide (radical dotNO) and peroxynitrite anion (ONOO)].

Section snippets

Plant material

J. regia leaves were collected during Summer, July 2003, in Mirandela, Northern Portugal, in a highland with high thermal amplitude. The leaves were dried at room temperature. Voucher specimens were preserved in our laboratory for further reference.

Chemicals

Dihydrorhodamine 123 (DHR), 4,5-diaminofluorescein (DAF-2), 3-(aminopropyl)-1-hydroxy-3-isopropyl-2-oxo-1-triazene (NOC-5), nitroblue tetrazolium chloride (NBT), lucigenin, diethylenetriaminepentaacetic acid (DTPA), β-nicotinamide adenine nucleotide

Total phenols

The total phenolic content of the J. regia ethanol:water (4:6) extract was 270 ± 3 mg GAE per g of lyophilised extract (mean ± SE). The extract yield was 0.804 g, which corresponds to a recovery of 20.1%. Several polyphenols have been identified on J. regia leaves, namely cinnamic acids (3-caffeoylquinic acid, 3-p-coumaroylquinic and 4-p-coumaroilquinic acids) and flavonoids (quercetin 3-galactoside, quercetin 3-arabinoside, quercetin 3-xyloside, quercetin 3-rhamnoside and a kaempferol derivative).

Conclusions

The studied J. regia (ethanol:water) extract was shown to be very effective against the evaluated pro-oxidant species. The scavenging effects of the studied reactive species have been described for some of the polyphenols that have been identified in J. regia leaves. These data imply that at least part of the observed antioxidant activity may be a result of the extract’s phenolic compounds.

The results of the present study showed that the J. regia leaf can be used as an easily accessible source

Acknowledgement

The authors greatly acknowledge José Madureira for the collection of Juglans regia leaves.

References (40)

  • G.R.M.M. Haenen et al.

    Peroxynitrite scavenging by flavonoids

    Biochemical and Biophysical Research Communications

    (1997)
  • B. Halliwell et al.

    Role of free radicals and catalytic metal ions in human disease: An overview

    Methods in Enzimology

    (1990)
  • B. Halliwell et al.

    The deoxyribose method: a simple “test-tube” assay for the determination of rate constants for reactions of hydroxyl radicals

    Analytical Biochemistry

    (1987)
  • N. Ito et al.

    Studies on antioxidants: Their carcinogenic and modifying effects on chemical carcinogenesis

    Food and Chemical Toxicology

    (1986)
  • N.W. Kooy et al.

    Peroxynitrite-mediated oxidation of dihydrorodamine 123

    Free Radical Biology and Medicine

    (1994)
  • C.S. Lau et al.

    Identification and quantification of glycoside flavonoids in the energy crop Albizia julibrissin

    Bioresource Technology

    (2007)
  • A. Moure et al.

    Food Chemistry

    (2001)
  • M. Ohsugi et al.

    Active-oxygen scavenging activity of traditional nourishing-tonic herbal medicines and active constituents of Rhodiola sacra

    Journal of Ethnopharmacology

    (1999)
  • G.L. Squadrito et al.

    Oxidative chemistry of nitric oxide: the roles of superoxide, peroxynitrite, and carbon dioxide

    Free Radical Biology and Medicine

    (1998)
  • M. Valko et al.

    Free radicals and antioxidants in normal physiological functions and human disease

    The International Journal of Biochemistry and Cell Biology

    (2007)
  • Cited by (0)

    View full text