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Medical use and toxic properties of rue (Ruta graveolens) and peppermint


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Medical use and toxic properties of rue (Ruta graveolens) and peppermint (Mentha piperita)
Nóra Kováts, András Ács, Maha Refaey, Zsuzsanna Simonné Baán
University of Pannonia, Institute of Environmental Engineering,

8200 Veszprém P.O.Box 158, E-mail: toxlab@almos.vein.hu




Abstract
Several chemicals have been identified which are produced by different herbs, also, their mode of action is also well-known. The purpose of the study was to find out if herbs are producing chemicals ‘on purpose’, for self-defence. Specificity of chemicals produced by two selected medical plants, rue (Ruta graveolens) and peppermint (Mentha piperita) was represented by different ecotoxicity tests: insecticide property by standard Daphnia test and bactericide property by the Vibrio fischeri bioluminescence inhibition test.

Keywords: rue, peppermint, ecotoxicology, bioassay, Vibrio fischeri, Daphnia magna



1. Introduction
Herbs are widely used for their medical properties but it is less frequently discussed what purpose chemicals produced by these plants might serve for the plant itself. The purpose of the study was to find out if herbs are producing chemicals ‘on purpose’, for self-defence. Our null hypothesis was that plant productions can be (1) antibacterial/antifungal agents; (2) of insecticide properties; (3) against grazing herbivores or (4) allelochemicals against competitors. In order to find out their specificity, the first two target groups were represented by different (standardised) ecotoxicity tests:

  • Bacteria were represented by Vibrio fischeri

  • Arthropods by Daphnia magna.

For our study two herbs were selected, differing significantly in their taxonomy, chemicals produced and medical use. Rue or rude (Ruta graveolens) is a rather neglicted and even controversial medical plant. The Greeks regarded it as an antimagical herb, because it served to remedy the nervous indigestion they suffered when eating before strangers which they attributed to witchcraft. Also, in the Middle Ages it was considered in many parts of Europe a powerful defence against witches. “Rub the twigs with the sprig of fresh rue and then place the sprig of rue in the bowl of water with the twine. Hold one twig in each hand and repeat the following words aloud: right to left and left to right bind these twigs and tie them tight return each wicked to evil spell to its sender and teach them well! Remove the twine from the bowl of water and tie the twigs together to form an x shape with them. Allow the whole to dry in the light of the sun and then keep it in the room where you sleep.” This spell can be found at http://www.spells-magic.com/protectfromevilspells.html which gives a collection of rare magic spells and rituals.

Ruta graveolens L. belongs to the family Rutaceae. Its main active principles are: glycosides (such as rutine, a flavonoid); alkaloids (quinolones: coquisagenine, skimmianine and graveoline); furocoumarins (psoralens: bergaptene and xantotoxine); essential oils: methyl-nonyl-ketone, methyl-n-octyl-ketone and methyl-heptyl-ketone; alcohols: methyl-ethyl-carbinol, pinene, limenenes.

Rue has a long history of use as a domestic remedy, being especially valued for its strengthening action on the eyes. Rue oil and infusions of rue were formerly used as antispasmodics and emmenagogues. It is recommended in herbal treatment of insomnia, headaches, nervousness, abdominal cramps, and renal troubles. Although it is beyond our scope of investigation, it is interesting to mention that rue was used by indigenous communities for fertility regulation and does have abortifacient effect (Ciganda and Laborde, 2003; Conway and Slocumb, 1979). Gutiérrez-Pajares et al. (2003) demonstrated that Ruta graveolens aqueous extract can retard mouse preimplantation embryo development. Clinical toxications are reported in countries where voluntary abortion is illegal and relative large doses are administered on purpose. Several websites advise to use it internally with some caution, there is even one stating: Highly toxic, may be fatal if eaten! (http://www.ces.ncsu.edu/depts/hort/consumer/poison/Ruta_gr.htm).

The herb was a very common spice in ancient Rome, nowadays it is only scarcely used in Europe. Its popularity is greatest in Ethiopia, where fresh leaves are sometimes used as a coffee flavourant. Ethiopian cuisine is unique in using not only rue leaves, but also the dried fruits (rue berries).

Ruta sp. cause contact dermatitis being similar to burn injury (Furniss and Adams, 2007). There are scarce data available on rue toxicity to mammals. Gonzalez-Trujano et al. (2006) showed that Ruta chalepensis ethanol extract until 5000 mg/kg, p.o., did not produce mortality, nor macroscopic tissue injury or weight loss during the observation period of 14 days.

Rue has been rather used for its insect repellent property. In the Medieval Ages ‘it was the custom for judges sitting at assizes to have sprigs of rue placed on the bench of the dock against the pestilental infection brought into courts from gaol by the prisoner’ says Mrs. M. Grieve in her Modern Herbal (1931). It was believed that ‘rue-water sprinkled in the house kills all the fleas’.

It is known to disrupt feeding behavior in butterfly larvae (Hypsipylagrandella) (Mancebo et al., 2001). Abramson et al. (2007) investigated if rue can be used as a natural insecticide against Rhodnius prolixus which is the main vector of Chagas disease in Venezuela. They found that in comparison to the most effective agent, cinnamon, rue was not able to stop molting of larvae but did influence the life-cycle, delaying it.

On the contrary, peppermint (Mentha piperita) is on the FDA’s GRAS list (generally recognized as safe). It is one of the world’s oldest medicinal herbs, nowadays it is cultivated in huge quantities and peppermint oil is the most extensively used of all the volatile oils. The plant which belongs to the family Lamiaceae is most likely a natural hybrid between spearmint (Mentha spicata) and water mint (Mentha aquatica). Its Latin name, Mentha, comes from the Greek Mintha, the name of a mythical nymph who to have metamorphosed into a plant.

Its potentially active chemicals include volatile oils such as menthol (which is the main ingredient), menthone, menthyl acetate, neomenthol, isomenthone, menthofuran, limonene, pulegone, alpha and beta pinene, and trans-sabinene hydrate7; monoterpenes; caffeic acids; flavonoids and tannins. Peppermint is considered mainly to have antiseptic and antimicrobial activity but is also considered by herbalists as an astringent, antipruritic, antispasmodic, antiemetic, carminative, diaphoretic, mild bitter, analgesic, anticatarrhal, rubefacient, stimulant, and emmenagogue. Peppermint oil vapor is used as an inhalant for respiratory congestion. Peppermint tea is used to treat coughs, bronchitis, and inflammation of the oral mucosa and throat. It has traditionally been used to treat a variety of digestive complaints such as colic in infants, diarrhea, indigestion, nausea and even anorexia. Peppermint oil is used to relieve menstrual cramps and (externally) to treat neuralgia, headaches and migraines.

In addition to traditional medical applications, recent studies have revealed its antimutagenic properties as well (Yu et al., 2004; Samarth et al., 2006)



2. Materials and methods
Aqueous extracts were prepared following the method given by Economou (2007). Leaves and stems were oven-dried at 75°C for 48 h and then ground. 30 g of ground tissue was diluted in 500 ml of deionized water and then treated by shaking for 24 h at room temperature.


2.1 Vibrio fischeri bioluminescence inhibition test
For the Vibrio fischeri bioluminescence inhibition test the ToxAlert®100 luminometer developed by Merck was used, following the protocol described by BS EN ISO 11348.3, Part 3 – Method using freeze-dried bacteria. Bioluminescence is a rapid indicator of the metabolic status and of the viability of the cell. The enzyme involved in the process is bacterial luciferase. A toxic substance will cause changes in some cellular structures or functions such as the electron transport system, cytoplasmic constituents or the cell membrane, which are directly reflected in a decrease in bioluminescence. The luminometer automatically calculates bioluminescence inhibition in %. Main benefit of the test is its rapidity, with the exposure of only 30 minutes.

2.2 Daphnia magna mortality test
Daphnids are perhaps the most commonly used test organisms: they occur world-wide and also, they have a key role in aquatic ecosystems, providing important food for fish. During toxicity testing the ISO 6341:1996 standard was followed. End-point of the test is the inhibition of motility (practically mortality). This is an acute toxicity test, where the exposure is 24 – 48 hours.

2.3 Statistical evaluation
Both bioluminescence inhibition and mortality were expressed in the form of EC50 which is the concentration causing 50% of effect. EC50 values were calculated using the probit software developed by USEPA.

3. Results and conclusions
Table 1 gives the calculated EC50 values for Vibrio fischeri and Daphnia magna tests, comparing the toxicity of Ruta graveolens and Mentha piperita.
Table 1

Calculated EC50 values for Vibrio fischeri and Daphnia magna tests, comparing the toxicity of Ruta graveolens and Mentha piperita. EC50 values are given in g/l.







Ruta graveolens

Mentha piperita

EC50 Vibrio fischeri (g/l)

1.483

0.667

EC50 Daphnia magna (g/l)

0.839

1.43

These results confirmed our null hypothesis, that is, rue was suspected to have higher toxicity for arthropods and mint which is known for its antibacterial property was suspected to have higher toxicity in the bacterial bioassay.

As both herbs seem to have an inherent capacity for self-protection, the question immediately arises if this capacity can be used in biological/ecological pest control. Meepagala et al. (2005) extracted rutacridone epoxide which showed significantly higher activity than commercial fungicides captan and benomyl in micro-bioassay against the agriculturally important pathogenic fungi Colletotrichum fragariae, C. gloeosporioides, C. acutatum, and Botrytis cineara and Fusarium oxysporium. Unfortunately, rutacridone epoxide is reported as a direct-acting mutagen, and none of the synthetic analogs tested by the authors showed comparable activities to it. Paulini et al. (1987) also detected mutagenic compounds in rue. However promising pesticide activity rue might have, these detected mutagenic components preclude its use as an agrochemical.

Mentha piperita, again, seems to be much unambiguous to use, without predictable side-effects. Its pesticide properties on pathogen microorganisms has been well demonstrated, here only agrological aspects are discussed. Essential oil of a close relative, Mentha rotundifolia (Moroccan mint) inhibits growth of Aspergillus niger, Escherichia coli, Bacillus subtilus (El-Arch et al., 2003). Its fungicidal activity of potato late blight (Phytophthora infestans) was reported by Quintanilla et al. (2002); in fact, of selected essential oils peppermint was the second most effective, following thyme.

In addition to its bactericidal and fungicidal effectiveness, insecticide properties has been widely investigated. Abivardi & Benz (1984) studied plant extracts as feeding repellents (antifeedants) against Pieris brassicae (L.), the large white butterfly. They found that extracts of peppermint significantly reduced larval ceding in glasshouse experiments. Lamiri et al. (2001) tested the effectiveness of a close relative, Mentha pulegium against Hessian fly (Mayetiola destructor), which is the major pest for wheat in Morocco. They found that essential oil produced by M. pulegium was one of the most effective amongst nineteen essential oils tested, causing 100% mortality in adults. Repellent activity of peppermint against mosquitoes has been perhaps the most widely studied. Essential oil extracted from Mentha piperita provided almost 100% protection against Culex quinquefasciatus and Anopheles culicifacies (Ansari et al., 1999) and is recommended for personal protection, avoiding or minimising the use of sysnthetic chemicals. Larvicidal effects of the essential oil was also demonstrated against Aedes aegypti, Anopheles stephensi, and Culex quinquefasciatus (Pathak et al., 2000).

Our study confirms that standard ecotoxicological tests can be applied to assess the bactericide/insecticide potential of medical herbs. As these bioassays use standard test organisms and are carried out following a standard protocol, such data are produced which are expressed in a unified form and a large range of herbs can be screened and compared.
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