Biflavonoids from Brazilian pine Araucaria angustifolia as potentials protective agents against DNA damage and lipoperoxidation

doi:10.1016/j.phytochem.2004.11.014

Phytochemistry

Volume 66, Issue 18, September 2005, Pages 2238-2247

https://doi.org/10.1016/j.phytochem.2004.11.014 Get rights and content

Abstract

A biflavonoid fraction (BFF) obtained from Araucaria angustifolia needles was effective to quench singlet oxygen (¹O₂), to protect plasmid DNA against single strand break (ssb) caused by ¹O₂ or Fenton reaction and to inhibit Fenton or UV radiation-induced lipoperoxidation in phosphatidylcholine liposomes. The activity of the biflavonoid fraction (BFF) was compared with quercetin, rutin (flavonoids), ginkgetin, amentoflavone (biflavonoids), α-tocopherol and Trolox^®. The BFF displayed a higher quenching rate constant compared to flavonoids and biflavonoids and protected against ssb induced by ¹O₂. Although the BFF was not as efficient as either flavonoids, α-tocopherol or Trolox^® in protection against ssb induced by Fenton-reaction or lipoperoxidation, these scavenging properties suggest that BFF is still an excellent candidate for successful employment as an antioxidant and photoprotector.

Graphical abstract

Biflavonoids from Araucaria angustifolia were able to protect against DNA damage and lipoperoxidation promoved by reactive species (RNOS). The ¹O₂ quenching rate constant using the time-resolved near infrared luminescence technique was also determined.

Introduction

The generation of reactive nitrogen and oxygen species (RNOS) has been shown to occur in multiple physiological processes, resulting in oxidation of several biological targets, including proteins, enzymes, lipids and DNA (Sies, 1991, Martinez et al., 2003). Singlet molecular oxygen (¹O₂) is an electronically excited species involved in mammalian cells under both normal and pathological conditions (Halliwell and Gutteridge, 1998, Pierlot et al., 2000). Due to its relatively long half-life in aqueous systems, ¹O₂ is able to move through appreciable distances in the cellular environment affecting different targets (Piette, 1991, Ravanat et al., 2000, Miyamoto et al., 2003, Martinez et al., 2002).

There is a much evidence indicating that RNOS are involved in in situ DNA damage, contributing to the aetiology of several human diseases (Helbock et al., 1999). Thus the search for new compounds able to prevent oxidative damage is an important strategy aimed at preventing injuries (Sies, 1993). Several studies have revealed antioxidant activities of compounds, such as lycopene (Di Mascio et al., 1989a, Di Mascio et al., 1992), curcumin (Subraminian et al., 1994), flavonoids (Williams et al., 2004), and related polyphenols (Arteel and Sies, 1999, Klotz and Sies, 2003) as inhibitors of RNOS-induced damage to biomolecules.

Flavonoids are widely distributed secondary metabolites in the plant kingdom and have several important roles as pigments, bactericides, fungicides, etc. Besides, they have other characteristic, properties, such as anti-inflammatory, antibacterial, anticarcinogenic and oestrogenic agents (Middleton and Kandswami, 1986). Additionally, flavonoids can inhibit some enzymes, such as cyclooxygenase, lipoxygenase and phospholipase A₂ (Lee et al., 1996, Kim et al., 1998, Saponara and Bosisio, 1998), and are also able to chelate metals, which play an important role in metal-induced free radical reactions (Deng et al., 1997). Biflavonoids are dimers of flavonoids, linked by a C–O–C or C–C bond, their biological properties include anti-inflammatory and antiarthritic activites in animals (Harborne, 1967). Such capacities were associated with an ability to suppressing reactive species and to inhibit cyclooxygenases as well (Kim et al., 1999).

The needles of Araucaria angustifolia, an endemic conifer in southern Brazil, contain several amentoflavone-type (C8″–C3′) biflavones (Fonseca et al., 2000), which differ from each other in the number and position of methoxyl groups (Table 1).

The major aim of this study were: to determine the kinetic parameters of ¹O₂ quenching of A. angustifolia biflavonoids, using a time-resolved near infrared luminescence technique; to examine the biflavonoid effect on DNA damage induced by a clean source of ¹O₂ generated chemically by a naphthalene endoperoxide derivative and by Fenton reaction in plasmid pBluescript and PUC 19. The protector capacity of these biflavonoids against UV radiation-induced lipoperoxidation was also determined.

Section snippets

Identification of biflavonoids

The BFF, precipitated from MeOH–H₂O (3:7) solution was analyzed by HPLC/MS and contained six major biflavonoids (Table 1). The determination of a common 3′-8″-interflavonoid linkage (amentoflavone type) between the flavonoid units was based on the analysis of ¹H and ¹³C NMR spectroscopic on data of the per-methylated product of BFF. Spiking with authentic samples of biflavonoids from Ginkgo biloba allowed the identification and characterization of amentoflavone (1, 2.4 % of relative abundance

Conclusions

Araucaria angustifolia needles contain six major biflavones amentoflavone, mono-O-methylamentoflavone, di-O-methylamentoflavone, ginkgetin, tri-O-methylamentoflavone and tetra-O-methylamentoflavone (BFF). Since the per-methylation of BFF produced hexamethylated amentoflavone, determined by analysis of MS, ¹H, and ¹³C NMR data, all components in the BFF were identified as derivatives of amentoflavone-type biflavonoids.

A biflavonoids fraction obtained from A. angustifolia needles showed a

General

NMR analyses were performed in a Bruker DRX 500 MHz using deuterated DMSO as solvent and TMS as internal standard.

The 1-H-phenalen-1-one was purchased from Acros (NJ, USA), whereas the hydrogen peroxide was supplied by Peróxidos do Brasil (Curitiba, Brazil). The biflavonoids, ginkgetin and amentoflavone from Ginkgo biloba, were kindly provided by Dr. Nicola Fuzzati from Indena Chemicals (Milano, Italy). Fresh needles of A. angustifolia (Bert) O. Kuntz were collected from a tree growing in the

Acknowledgements

This work was supported by the “Fundação de Amparo à Pesquisa do Estado de São Paulo”, FAPESP (Brazil), by “Conselho Nacional para o Desenvolvimento Científico e Tecnológico”, CNPq (Brazil), by “Programa de Apoio aos Núcleos de Excelência”, PRONEX/FINEP (Brazil) “Pró-Reitoria de Pesquisa da Universidade de São Paulo (Brazil)” and the John Simon Guggenheim Memorial Foundation (fellowship to P.D.M.). We gratefully thank Clécio F. Klitzke for the contribution to mass spectrometry analyses. L.F.Y

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Biflavonoids from Brazilian pine Araucaria angustifolia as potentials protective agents against DNA damage and lipoperoxidation

Abstract

Graphical abstract

Introduction

Section snippets

Identification of biflavonoids

Conclusions

General

Acknowledgements

FEBS Lett.

J. Photochem. Photobiol. B

Arch. Biochem. Biophys.

Biochim. Biophys. Acta

Phytochemistry

Free Radic. Biol. Med.

Meth. Enzymol.

Prostaglan. Leucotr. Essent. Fatty Acids

Toxicol. Lett.

Mutat. Res.

J. Photochem. Photobiol. B

J. Biol. Chem.

Free Radic. Biol. Med.

Mut. Res.

J. Photochem. Photobiol. B

Cancer Lett.

Scavenger and antioxidant properties of ten synthetic flavones

Free Radic. Biol. Med.

Flavonoids function as antioxidants: by scavenging reactive oxygen species or by chelating iron

Rad. Phys. Chem.

Quantification of singlet oxygen generated by thermolysis of 3,3′-(l,4–naphthylidene) dipropionate. Monomol and dimol photoemission and the effect of 1,4-diazabicyclo[2.2.2.]octane

J. Am. Chem. Soc.

UV-C irradiation-induced peroxidative degradation of microsomal fatty acids and proteins: protection by an extract of Ginkgo biloba (EGb 761)

Free Radic. Biol. Med.