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Citation of this paper

The effects of plant essential oils on in vitro camel rumen fermentation

I Medila, I Toumi, A Adamou1 and S Maaroufi2

Department of Cell and Molecular Biology, Faculty of Natural Science and Life, University Echahid Hamma Lakhder, El-oued, Algeria
1 Laboratoire protection des écosystèmes dans les zones arides et semi-aride, Université Kasdi Merbah, Ouargla
2 Department of Animal Production: National Agronomy Institute- Tunis


The objective of this study was to determine the effects of different doses of three natural essential oils (EOs) from semi-arid medicinal plants (Cotula cinerea, Juniperus phoenicea, and Artemisia campestris) on in vitro camel rumen fermentation of Atriplex halimus, a plant that is eaten by the dromedary. The levels added were 0.0, 0.10, 0.25 and 1.0 g/liter of the fermentation medium.

All the oils decreased the production of gas and methane per unit of substrate DM incubated. N-NH3 concentration was reduced by the oil from Juniperus phoenicea.

Key words: ammonia, dromedary, essential oils, methane


In recent studies, a variety of compounds and substances have been evaluated for their ability to reduce methane emissions from rumen fermentations in vitro and in vivo (Beauchemin et al 2008; Bunglavan et al 2010; Buddle et al 2011; Patra et al 2012; Adelusi et al 2015; Vongkhamchanh et al 2015; Sengsouly and Preston 2016). The inclusion of essential oils of medicinal plants, as natural alternatives, has been investigated for inclusion in ruminant diets for improving feed efficiency, as well as decreaet alsing the adverse envirental effects of ruminants (Wallace 2004).

The aim of the present study was to investigate the effect of different levels of essential oils on methane pproduction fromCotula cinerea, Juniperus phoenicea, Artemisia campestris in anin vitro camel rumen fermentation.

Materials and methods

Forage Samples

Sampling of plant material was done in a salty area named Benguecha (Taleb El Oued Province-Algeria). The plant Atriplex halimus (Amaranthaceae) was selected as the feed substrate for incubation. It was chosen as being the main component of the herbaceous layer along the route taken by camels. A preference by camels for this plant is well known; also that it is effectively degraded by the dromedary (Medila 2015).

Plant essential oil

Semi-arid native medicinal plants (Cotula cinerea, Juniperus phoenicea, Artemisia campestris) were collected from the El Oued region of Algeria. The essential oils from these plants were obtained through steam distillation using the “Celevenger” procedure (Clevenger, 1928). The distillation was carried out for four hours in accordance with the recommendations of the European Pharmacopoeia (AFNOR 2000). The extractions were repeated five times to confirm the return earned by the mode used. The essential oil was stored and maintained optimally at 4 °C protected from light.

Treatments and design

Three different doses were taken in the case of each essential oil: 0.10, 0.25 and 1.0 g/liter of in vitro fermentation medium. Each treatment was incubated in triplicate.

In vitro gas production

The in vitro fermentation was conducted according to the technique of calibrated glass syringes (100ml capacity) described by Menke and Steingass (1988). 200 mg dry weight of the samples were placed in triplicate in 100 ml calibrated glass syringes. Rumen liquid was collected from 3 dromedaries slaughtered in the morning. These animals were randomly selected, of different age and sex. The rumen liquor was bubbled with C0 2 for about 2 minutes and filtered through 4 layers of muslin cloth. The strained rumen liquor was mixed with a phosphate-bicarbonate buffer (Menke et al 1979) in a 2:1 ratio and 40ml of this mixture were introduced in each syringe for incubation. The level of the piston was recorded and the syringes were placed in an incubator (39 ± 0.5°C). Gas volumes were recorded after 24 h of incubation.

At the end of the incubation, the total gas produced in each fermenter was collected in gas bags and quantified by displacement of water. Samples were drawn from the total gas produced and fractioned for methane and carbon dioxide by the method of Fievez et al (2005). A 2ml sample was taken from the gas bag with the help of syringe and needle. It was then injected through the nozzle of another syringe containing 2 ml of 10M NaOH. The displacement of gas caused by the 10M NaOH indicated the volume of methane.

N-NH3 was determined by the colorimetric technique of Chaney and Marbach (1962).

Statistical analysis

The data were analyzed using SAS software (SAS 2013). The effects of treatments were compared with those of the control using the Dunnett test. Significant differences were declared at P < 0.05.


Relative to the control, all three sources of essential oils reduced production of methane and total gas (Table 1; Figures 1-3). Rumen ammonia levels were reduced by oil from Juniperus phoenicea but were not affected by essential oils from Cotula cinereal and Artemisia campestris.

Table 1. Mean values for gas and methane production, and N-NH3 concentration,
in an in vitro rumen microbial fermentation of three plant essential oils


Gas production







Cotula cinerea





















Juniper us phoenicea





















Artemisia campestris





















Figure 1. Effect of Cotula cinérea essential oil on methane
production in an in vitro fermentation of Atriplex halimus
Figure 2. Effect of Juniperus phoenicea essential oil on methane
production in an in vitro fermentation of Atriplex halimus
Figure 3. Effect of Artemisia campestris essential oil on methane
production in an in vitro fermentation of Atriplex halimus


In the present study, essential oil from Juniperus phoenicea was more potent than oil from Cotula cinérea andArtemisia campestris in lowering methane and N-NH3 production. The two latter sources of essential oil appeared to reduce methane production but did not affect N-NH3 concentration. The decrease in N-NH3 concentration with the addition of Juniperus phoenicea essential oil was probably consistent with direct inhibition of proteolysis, and peptidolitic and deamination processes. Patra (2011) demonstrated that essential oils can reduce NH 3 concentration and protein deamination by inhibiting hyper-ammonia producing bacteria. McIntosh et al (2003) demonstrated that a commercial blend of essential oil compounds reduced the rate of amino acid deamination and inhibited the growth of a specific group of ammonia hyper-producing bacteria. Addition of thymol (an essential oil) to a medium containing rumen liquid resulted in an accumulation of amino acid nitrogen and a decrease in the ammonia nitrogen according to Castillejos et al (2007).

Many studies have documented reduction in methane production by essential oils (Agarwal et al 2009, Patra et al 2012, Jahani-Azizabadi 2014, Joch et al 2017). Oils from eucalyptus, garlic, origanum and peppermint supplementations all educed methane production in the range of 18 to 87.0% according to Parra and Yu (2012). Knapp et al (2014) indicated that the effect of essential oils in reducing in vitro CH4 production was through a direct inhibition of methanogenic archaea and/or an indirect depression of some microbial metabolic processes involved in methanogenesis. However, Beauchemin and McGinn (2006) did not reveal any effect on methanogenesis in a study done in vivo.

The practical application of the findings reported in this paper have yet to be demonstrated.



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Received 14 October 2017; Accepted 29 October 2017; Published 2 November 2017

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