Antigenotoxic, antiproliferative and antimetastatic properties of a combination of native medicinal plants from Argentina

https://doi.org/10.1016/j.jep.2020.113479Get rights and content

Highlights

  • A ternary herbal mixture, HM2, from “jarillas” was standardized by phenolic profile.

  • Neither single plants nor HM2 induce DNA damage on Ames Test.

  • HM2 shown antigenotoxic effect, while each individual plant did not show it.

  • At low doses, HM2 protects DNA against mutagens on cell lines assays.

  • HM2 demonstrated tumor growth inhibition and anti-metastatic effect in vivo.

Abstract

Ethnopharmacological relevance

Jarilla is the common name of an appreciated group of native plants from the semi-arid region in Argentina (Larrea cuneifolia Cav., Larrea divaricata Cav. and Zuccagnia punctata Cav.) that have been historically consumed to heal respiratory, musculoskeletal and skin ailments, as well as recommended for weakness/tiredness, hypertension, diabetes and cancer treatment. It was previously reported that some biological properties could be improved when these plants are used jointly. Infusions of a defined mixture, composed by three Jarilla species, L. cuneifolia: L. divaricata: Z. punctata (0.5:0.25:0.25) (HM2) showed synergistic and additive effect on antioxidant activity even after passing through the gastro-duodenal tract.

Aim of the study

The main purpose of this work was to evaluate antigenotoxic, antitumor, and anti-metastatic properties of the Jarilla species that grow in the Northwest of Argentina and a herbal combination of them.

Material and methods

Infusions of Jarilla mixture (HM2), and of each single plant species were prepared. Phenolic profiles of infusions were analyzed by HPLC-ESI-MS/MS and two relevant chemical markers were quantified. The antigenotoxic activity was evaluated by using the Ames test and the Cytokinesis-Block Micronucleus (CBMN) assay against direct mutagens. Evaluations of both cytotoxicity and antiproliferative effects were conducted on tumor and non-tumor cell lines. Both in vivo tumoral growth and metastasis inhibition were evaluated by using a carcinoma model on Balb/c mice.

Results

HM2 mix could suppress genetic and chromosome mutations induced by 4-nitro-o-phenylendiamine (4-NPD) and doxorubicin. Herbal mixture and single plant infusions showed cytotoxic effect against mammary, uterus, and brain tumoral cells without a selective action vs normal human cell line. HM2 mix was able to reduce mammary tumor mass on the Balb/c mice model and showed a significant reduction in the number of metastatic nodules in the lungs.

Conclusions

Our results suggest that the combinations of three Jarilla species from northwest Argentina would be a promising alternative to treat or slow down the development of chronic diseases, such as cancer.

Introduction

According to the International Agency for Research on Cancer (IARC) statistics, Argentina is the seventh country in South America with high cancer incidence (IARC, 2018). The only common feature among all types of tumors is the uncontrolled cell proliferation, mainly due to mutations that alter cell regulatory mechanisms (Alexandrov and Zhivagui, 2019; Plankar et al., 2011). Therefore, prevention strategies and efficient treatments that, as far as possible, do not alter the life quality of patients becomes crucial. In this sense, there is a growing interest in developing chemotherapeutic or chemopreventive drugs from medicinal herbs used traditionally, or combination of them (Gras et al., 2018). The fundamental purpose underlying this practice is to obtain synergism, to reduce individual component doses in order to achieve lower toxicity, to minimize or delay resistance and to involve several molecular targets simultaneously. Plant mixtures have been playing a crucial role in ‘omic’ techniques to lead to a new generation of plant-derived drugs useful in multicausal pathologies such as cancer, cardiomyopathies, rheumatism, etc. (Wagner and Ulrich-Merzenich, 2009).

Larrea cuneifolia Cav. (Zygophyllaceae), Larrea divaricata Cav. (Zygophyllaceae) and Zuccagnia punctata Cav. (Fabaceae) belong to an important plant community in the Northwest of Argentina known as the “Jarillal” (Carrizo and Grau, 2014). These plant species are among the most useful and popular in traditional healing by local people in arid ecosystems in Argentina (Ladio and Lozada, 2009; Martínez et al., 2004). It was previously stated that they are commonly consumed as infusions or in taking baths (Carabajal et al., 2020b), and that they can be used to heal mycosis, respiratory, musculoskeletal and skin ailments, as well as weakness/tiredness, hypertension, diabetes and tumor treatment (Carabajal et al., 2020b; Davicino et al., 2011; Isla et al., 2016; Moreno et al., 2018). Regarding the latter, the medicinal use of Jarilla leaves to treate tumor formation was reported in ethnopharmacological surveys by Carabajal et al. (2020b), in an indigenous community in northwestern Argentina. Catalá (1938) also reported the application of these plants on “wound ulcers of a malignant nature” in southern Argentina. Moreover, in vivo antitumor capacity of L. divaricata (“Jarilla hembra”) against mammary carcinoma (Anesini et al., 1998) was demonstrated, and antigenotoxic activity of these species was also reported (Carabajal et al., 2017; Zampini et al., 2008). It is well-known that some phenolic compounds from Jarilla shrubs inhibit pathogenic bacteria and fungi, pro-inflammatory enzymes and show antioxidant, antigenotoxic and cytotoxic potential (Blecha et al., 2007; Chieli et al., 2012; Lü et al., 2010; Youngren et al., 2005).

In ancient cultures, mixing medicinal plants led to improve the individual properties of chemical components by increasing their solubility or bioavailability, once they were consumed (Grosso et al., 2015). Diaguita-Calchaqui people indicated that they generally consumed these plants one at a time but in some cases, they mixed them along with other shrubs (Carabajal et al., 2020b). Although no reports describe traditional uses of mixtures of these three Jarilla species, it was previously demonstrated that some such mixtures from Z. punctata/L. divaricata, Z. punctata/L. cuneifolia, and Z. punctata/L. nitida showed a synergistic effect in bi-combinations on antifungal activity against Saccharomyces cerevisiae, Candida albicans, and non-Albicans strains (Butassi et al., 2015; Moreno et al., 2020b). It was also stated that a ternary herbal combination, namely, 50% L. cuneifolia, 25% L. divaricata and 25% Z. punctata, has a synergistic and additive effect on antioxidant activity through several mechanisms of actions, even after passing through the gastro-duodenal tract (Carabajal et al., 2020a). Nevertheless, little is known about the chemotherapeutic potential of Jarilla mixtures against cancer.

Considering the traditional use of Jarilla leaves to treat tumor formation, this work aimed to provide new knowledge on the antigenotoxic potential, as well as the antitumor and anti-metastatic properties of Jarilla species and a ternary combination from them.

Section snippets

Chemicals

HPLC grade acetic acid and methanol were purchased from Merck (Darmstadt, Germany), β-nicotinamide adenine dinucleotide (NADH), nordihydroguaiaretic acid (NDGA), dimethyl sulfoxide (DMSO), L-Histidine, D-Biotin, 4-nitrophenylene-1,2-diamine (4-NPD), 2- aminofluorene (2-AF), penicillin-streptomycin solution and trypsin-EDTA solution, methyl methanesulfonate (MMS), cytochalasin B, oxaliplatin, Cole's hematoxylin/eosin, Aroclor-1254, Hepes solution, Cell Proliferation Kit (XTT-based colorimetric

Phytochemical profiling of ternary mixture infusion

Thirty-three chemical compounds belonging to different phenolic families (tannins, flavonoids, esters and lignans) were identified in HM2 infusion, according to literature and standard fragmentation patterns (Table 1). Most of the compounds belong to Z. punctata components, such as flavonols (compounds 46, 13, 24, 33), flavanones (8, 9, 12, 16, 17, 21), hydroxylated and methoxylated chalcones (7, 22, 31, 32), hydrolyzable tannins (1, 3), esters (19, 25, 26) and condensed tannins (catechin

Discussion

Numerous potential chemotherapeutic drugs have been derived from natural sources. Secondary metabolites synthesized by plants did provide several drugs for the treatment of various human diseases and 75% of the modern synthetic drugs have been derived mainly from plants (Newman and Cragg, 2016).

Plant species belonging to “Jarillal” inhabit arid ecosystems in Argentina and are recognized by local people as important plants for its medical, tinctorial, and spiritual uses (Carabajal et al., 2020b

Conclusion

A ternary combination of three Argentine native Jarilla species (HM2), composed by L. cuneifolia (50%), L. divaricata (25%), and Z. punctata (25%) showed anti-cancer effect and the antigenotoxic potential as mechanisms for cancer prevention were revealed. HM2 showed cytotoxic activity on different cell lines derived from human tumors, as well as inhibited tumor growth, and presents an anti-metastatic effect in vivo.

From these results, it could be inferred that the proposed herbal mixture is an

Authors’ contributions

M.P.A.C, H. D-N, I.S–S, A.P.P-G, T.R-E performed the micronucleus assay and antiproliferative in vitro experiments. M.P.A.C and I.C.Z, performed chemical profile analyses and the Ames test. J.P.F designed and performed in vivo experiments. I.C.Z., M.I.I. and D.C.T. designed the experiments and supervised the research. All authors analyzed the data. I.C.Z and M.P.A.C drafted the manuscript. I.C.Z and M.I.I. conceived and initiated the project.

Declaration of competing interest

The authors declare that they have no competing financial interests or personal relationships that could have influenced the work reported in this paper.

Acknowledgements

The authors are very grateful to the following Argentinian institutions: Universidad Nacional de Tucumán (PIUNT 2018-G637 Project), Agencia Nacional de Promoción Científica y Técnica (ANPCyT PICT2014, 3136 Project, PICT 2017, 4416 Project), and Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET- PUE 2018-0011 Project); Brazilian institutions: São Paulo Research Foundation (FAPESP; grant # 2016/24269–7), National Council for Scientific and Technological Development (CNPq) and

References (77)

  • E. Chieli et al.

    Effects of Zuccagnia punctata extracts and their flavonoids on the function and expression of ABCB1/P-glycoprotein multidrug transporter

    J. Ethnopharmacol.

    (2012)
  • R.v. Edenharder et al.

    Antimutagenic effects of flavoniods, chalcones and structurally related compounds on the activity of 2-amino-3-methylinidazo [4, 5-ƒ] quinoline (IQ) and other heterocyclic amine mutagens from cooked food

    Mut. Res./Fund. Mol. M

    (1993)
  • M. Fenech

    The in vitro micronucleus technique

    Mut. Res./Fund. Mol. M

    (2000)
  • M. Ferrer et al.

    Antimutagenic mechanisms of Phyllanthus orbicularis when hydrogen peroxide is tested using Salmonella assay. Mut. Res./Gen. Tox

    Eng.

    (2002)
  • J. Gnabre et al.

    Creosote bush lignans for human disease treatment and prevention: perspectives on combination therapy

    J. Tradit. Complement. Med.

    (2015)
  • A. Gras et al.

    Folk medicinal plant mixtures: establishing a protocol for further studies

    J. Ethnopharmacol.

    (2018)
  • C. Grosso et al.

    HPLC–DAD analysis and in vitro enzyme inhibition: an integrated approach to predict herbal binary mixture behaviour employing median effect equation

    Microchem. J.

    (2015)
  • X. Guo et al.

    Biphasic regulation of spindle assembly checkpoint by low and high concentrations of resveratrol leads to the opposite effect on chromosomal instability. Mut. Res./Gen. Tox

    Eng.

    (2018)
  • L.V. Jacociunas et al.

    Protective activity of Cynara scolymus L. leaf extract against chemically induced complex genomic alterations in CHO cells

    Phytomedicine

    (2013)
  • A.H. Ladio et al.

    Human ecology, ethnobotany and traditional practices in rural populations inhabiting the Monte region: resilience and ecological knowledge

    J. Arid Eng.

    (2009)
  • J.D. Lambert et al.

    Cytotoxic lignans from Larrea tridentata

    Phytochemistry

    (2005)
  • D.K. Mahapatra et al.

    Anti-cancer chalcones: structural and molecular target perspectives

    Eur. J. Med. Chem.

    (2015)
  • D.M. Maron et al.

    Revised methods for the Salmonella mutagenicity test. Mut. Res./Envir

    Mutagenesis

    (1983)
  • M.R. Martínez et al.

    Environment and illness in the Calchaquí Valley (Salta, Argentina): phytotherapy for osteo-articular and cardio-circulatory diseases

    J. Ethnopharmacol.

    (2004)
  • M.A. Moreno et al.

    Active properties of edible marine polysaccharide-based coatings containing Larrea nitida polyphenols enriched extract

    Food Hydrocolloids

    (2020)
  • M.A. Moreno et al.

    Crosslinked electrospun zein-based food packaging coatings containing bioactive chilto fruit extracts

    Food Hydrocolloids

    (2019)
  • M.A. Moreno et al.

    Antifungal, anti-inflammatory and antioxidant activity of bi-herbal mixtures with medicinal plants from Argentinean highlands

    J. Ethnopharmacol.

    (2020)
  • K. Mortelmans et al.

    The Ames Salmonella/microsome mutagenicity assay

    Mutat. Res./Fund. Mol. M

    (2000)
  • V. Nandakumar et al.

    Multi-targeted prevention and therapy of cancer by proanthocyanidins

    Canc. Lett.

    (2008)
  • M. Novak et al.

    Cytotoxicity and genotoxicity of anticancer drug residues and their mixtures in experimental model with zebrafish liver cells

    Sci. Total Environ.

    (2017)
  • J. Piloto-Ferrer et al.

    Xanthium strumarium's xanthatins induces mitotic arrest and apoptosis in CT26WT colon carcinoma cells

    Phytomedicine

    (2019)
  • M. Plankar et al.

    On the origin of cancer: can we ignore coherence?

    Prog. Biophys. Mol. Biol.

    (2011)
  • J.L. Quiles et al.

    Antioxidant nutrients and adriamycin toxicity

    Toxicology

    (2002)
  • D. Raffa et al.

    Recent discoveries of anticancer flavonoids

    Eur. J. Med. Chem.

    (2017)
  • É.d.M. Reis et al.

    Assessment of the genotoxic potential of two zinc oxide sources (amorphous and nanoparticles) using the in vitro micronucleus test and the in vivo wing somatic mutation and recombination test

    Food Chem. Toxicol.

    (2015)
  • A. Rostami-Vartooni et al.

    Photocatalytic degradation of azo dyes by titanium dioxide supported silver nanoparticles prepared by a green method using Carpobrotus acinaciformis extract

    J. Alloys Compd.

    (2016)
  • L.C. Rufatto et al.

    Mikania laevigata: chemical characterization and selective cytotoxic activity of extracts on tumor cell lines

    Phytomedicine

    (2013)
  • J.S. Santos et al.

    Optimized Camellia sinensis var. sinensis, Ilex paraguariensis, and Aspalathus linearis blend presents high antioxidant and antiproliferative activities in a beverage model

    Food Chem.

    (2018)
  • View full text