AHDB

Feed additives in ruminant nutrition

Can feed additives reduce methane and improve performance in

ruminants?

Poppy Frater

Beef and Sheep Scientist

June 2014

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Summary

There is a growing range of feed additives, aimed for use in ruminant diets, that offer

potential to improve rumen fermentation efficiency while reducing methane output. This

review aims to determine how effectively those additives on the market and under

development are meeting these claims.

Methane production is the dominant mechanism for hydrogen disposal in the rumen, a

necessary activity for healthy rumen functioning. Potential opportunities to reduce methane

production involve: inhibiting the methaneogenic archea and/or the accompanying protozoa,

supplying an alternative hydrogen disposal mechanism, encouraging other natural hydrogen

disposal routes and slowing fermentation to improve fermentation efficiency.

A recent review completed by Herefordshire University for the European Food Safety

Authority and a Food and Agriculture Organisation report on technical options for methane

mitigation form the basis of this report. From these, the most promising feed additives were

identified and effectiveness discussed.

Plant extracts, in various forms, hold significant potential for reducing methane, including:

fatty acids, dietary lipids, essential oils and condensed tannins. Polyunsaturated fatty acids

and essential oils are thought to have a similar mode of action to monensin (an ionophore

currently prohibited for use in the EU); they favour propionate producing bacteria in the

rumen, thereby encouraging another hydrogen sink.

Dietary lipids, or their constituent fatty acids alone, reduce methane emissions but at high

intakes they can reduce digestibility and dry matter intake.

Plant saponins are thought to reduce rumen protozoa numbers which can affect the methane

producing bacteria and slow down protein turnover in the rumen, increasing transport of

microbial nitrogen to the duodenum.

Garlic oil is the essential oil with most supporting evidence for methane reduction, but effects

on performance are variable.

A promising group of chemical additives are electron receptors. Nitrate is an electron

receptor that acts as an alternative hydrogen sink to methane, but if used in high protein diets,

excess ammonia production is likely. Nitrate toxicity is a risk to ruminants, but this can be

overcome with gradual introduction of nitrate to the diet.

Overall, most additives require further long term studies in the live ruminant to determine

how effective they are in commercial systems. To be sustainable and taken up by the

industry, the feed additive would need to be effective over long periods of time, non-toxic for

animals, the environmental and consumers and cheap enough for standard use in animal

feeds.

Contents

1. Introduction ..................................................................................................................................... 6

2. Feed additives to reduce methane production in ruminants .......................................................... 10

2.1 EU regulation ........................................................................................................................ 10

2.2 European Food Safety Authority report ................................................................................ 10

2.1.1. Additive development ......................................................................................................... 11

2.3 Food and Agriculture Organization review........................................................................... 14

2.3 The most promising feed additives for reducing methane ......................................................... 16

2.3.1 Fatty acids and plant oils ...................................................................................................... 16

2.3.4 Essential oils (Armoracia rusticana (horseradish) extract, Mentha microphylla, Carvacrol

(oregano)) ...................................................................................................................................... 21

2.3.5 Plant extracts: Saponins ....................................................................................................... 25

2.3.6 Electron receptors ................................................................................................................ 26

3. Persistence ..................................................................................................................................... 32

4. Conclusion .................................................................................................................................... 33

5. References ..................................................................................................................................... 34

Feed additives in ruminant nutrition

1. Introduction

Feed additives are products used in animal nutrition to improve the quality of feed and the

quality of food from animal origin, or to improve the animals’ performance and health. They

are categorised as follows:

Technological additives (e.g. preservatives, antioxidants, emulsifiers, stabilising

agents, acidity regulators, silage additives)

Sensory additives (e.g. flavours, colorants)

Nutritional additives (e.g. vitamins, minerals, amino acids, trace elements)

Zootechnical additives (e.g. digestibility enhancers, gut flora stabilizers)

Coccidiostats and histomonostats (additives used in poultry diets for health

reasons)

In recent years, zootechnical additives1 (the focus of this review from here on) have shown

vast growth in the research field and, consequently, in the feed market. Their suggested

mode of action varies, but in general, they aim to manipulate the rumen fermentation

environment to achieve greater efficiencies.

This range of feed additives has potential to deliver the following improvements in ruminant

nutrition:

1. Increase feed conversion efficiency (FCE) and productivity

2. Stabilise rumen pH to reduce acidosis risk

3. Increase dry matter intake (DMI)

4. Reduce methanogenesis

5. Enhance rumen development

6. Reduce pathogen load and shedding

7. Improve meat quality

8. Enhance rumen stability during dietary transitions

9. Buffer against dietary health risks (e.g. mycotoxins)

1 For the remainder of this review, zootechnical feed additives will be referred to solely as feed additives

This review aims to:

Inform producers/advisors of the most promising feed additives for ruminants in

terms of methane reduction

Provide information on the other effects, risks and practicalities of the feed additives

Where the feed additive is not commercially available, describe those most promising

for future development

1.1 The potential of feed additives

Methane (CH4) is a by-product of carbohydrate breakdown in the reticulo-rumen. Methane

production is energy inefficient, wasting 2-15% of digested energy (McCrabb and Hunter,

1999, Johnson and Ward, 1996, Blaxter and Clapperton, 1965).

Dietary manipulation could reduce methane production while improving performance. A

simplified diagram taken from a Food and Agriculture Organisation report (Alexander N.

Hristov et al., 2013) demonstrates carbohydrate breakdown in Figure 1.

Figure 1: Carbohydrate metabolism in the rumen, taken from Alexander N. Hristov et al.

(2013)

Reaction stoichiometry is the term to describe the relationship between substances as they

interact in chemical reactions. It is used to estimate the quantity of products formed during a

reaction. If we can understand how carbohydrates break down (the reaction stoichiometry)

and have an understanding of the effect of a specific feed additive on the process, we can

predict how that additive will affect the various product quantities.

In general, the reaction stoichiometry for carbohydrate breakdown has been described by Van

Soest (1994) as follows:

Glucose + ammonia microbes + methane + carbon dioxide + volatile fatty acids*

*butyrate, formate, lactate, propionate, acetate

Note: methanogenic archea are anaerobic, therefore this only occurs under anaerobic conditions

Therefore, the main products of carbohydrate breakdown in the rumen are volatile fatty acids

(VFA), methane and carbon dioxide (CO2). Alcohols and lactate are also formed.

The basic problem in anaerobic metabolism is the storage of oxygen (i.e. as carbon dioxide)

and disposal of hydrogen (i.e. as methane) (Van Soest (1994)). Methane works as a hydrogen

sink to remove hydrogen from the rumen for healthy rumen functioning.

Methane is commonly referred to as “2H” sink because pairs of protons and electrons are

donated and accepted in metabolic reactions. If we can replace this process with an

alternative, there is potential to reduce methane emissions and improve energy-use efficiency,

but in-doing so caution is required not to replace the methane with another pollutant. Ideally,

the hydrogen will be redirected to a useful form of nutrition for the animal.

Although methane can be produced from VFA and alternative sinks for hydrogen do exist in

other environments (acetate, for example), these processes appear to be of little significance

in the rumen (Russell and Wallace, 1997). As shown in Figure 1, acetate and butyrate

produce four and two hydrogen molecules respectively and propionate is a 2H sink.

Therefore propionate is the VFA to encourage if the objective is to reduce methane

production as it decreases the overall amount of 2H available to reduce carbon dioxide to

methane.

There is potential to manipulate carbohydrate breakdown to reduce methane production and

potentially improve performance in the following ways:

Encourage propionate production, thereby providing an alternative H sink (problem

with this approach is that propionate has a lower affinity for hydrogen than methane)

Adding an alternative hydrogen sink into the diet , e.g. nitrate (this has health risks

and may increase ammonia output unless utilised by microbes)

Encourage the production of acetate and butyrate, as these utilise the hydrogen before

it can be directed to produce methane (less potential here as acetate and butyrate still

produce hydrogen)

Inhibit or eliminate the methanogenic microorganisms, thereby forcing the use of

other hydrogen sink pathways (problematic if these organisms have other functions in

the rumen)

Slow down fermentation to improve nitrogen and energy-use efficiency

To be sustainable and taken up by the industry, the feed additive would need to be effective

over long periods of time, non-toxic for animals, the environment and consumers and cheap

enough for standard use in animal feeds.

1.2 The relationship between methane and dry matter intake

To determine how effective an additive is at reducing methane, it is important that the studies

report the effect using the same units, per land area or per unit of product, for example. In

addition, we should also look out for a reduction in methane that is simply due to a reduction

in feed intake and/or a reduction in performance.

Dry matter intake is an important determinant of methane output. A meta-analysis showed

on average, for every kilogram of dry matter intake approximately 30 litres of methane is

produced (Mills et al., 2008). Therefore, when studies report a reduction in methane and a

reduction in dry matter intake, the additive may be reducing digestibility or palatability of the

feed.

2. Feed additives to reduce methane production in ruminants

2.1 EU regulation

According to EU Regulation 1831/2003, only additives that have been authorised by the

European Food Safety Authority (EFSA) can be placed on the market in the EU.

Authorised additives can be put on the market solely for the specific use described for

authorisation. Companies or researchers wishing to get an additive authorised must submit

an application and a dossier to EFSA which must:

Enable assessment of additives based on the current state of knowledge

Demonstrate compliance with the fundamental principles for authorisation

Have a minimum of three long term in vivo studies showing significant effects for the

relevant target species/category. These should be carried out at least two different

locations, one of which should be in the EU. The studies should include the lowest

dose proposed by the applicant to enable recommendations on dose to be made.

Demonstrate the safety of the product ((EFSA), 2012).

Probiotics (fungi, bacteria, enzymes) are the only feed additives categorised under

zootechnical additives for beef and sheep production and therefore, technically, should be the

only feed additives in that category on the market in the UK. However, some additives, with

the claim of offering a dietary nutritional component, may appear on the catalogue of feed

materials European Commission (2013).

2.2 European Food Safety Authority report

A recent review completed by Herefordshire University for the European Food Safety

Authority (EFSA) conducted a meta-analysis of feed additives aimed at mitigation of

environmental impacts of livestock (Lewis et al., 2013).

Many meta-analyses on feed additives report substantial variation in additive effects on

animal performance and other measured variables due to differences in the methodology

employed across the different experiments (Eugène et al., 2008, Sales, 2011, Desnoyers et

al., 2009).

This makes it difficult to determine the efficacy of a substance, as it is hard to disentangle the

effect of the additive from any interaction with other factors. Additionally, the variation in

different diets would likely influence the results (Desnoyers et al., 2009). The review

covered 246 substances in ten major groups, as illustrated in Table 1.

Table 1: The substance groups covered by the EFSA report (Lewis et al., 2013)

Substance group Number of substances Number of studies

Organic acids & their salts 24 47

Fatty acids 12 15

Amino acids 8 11

Plant & animal oils, fats 16 33

Plant extracts

General 6 6

Saponins 7 34

Tannins 27 28

Essential oils & spices 42 48

Antibiotic drugs

General 11 13

Ionophores 9 47

Bacteria, enzymes & yeasts 29 62

Mineral salts and complexes 11 9

Miscellaneous substances

Chemical substances 21 31

Natural substances 23 29

TOTAL 246 -

2.1.1. Additive development

Unfortunately, many of the additives, particularly those that had the greatest effect (Table 2)

were only tested in vitro, and not in vivo. In vitro research gives insight into the potential of

the additive with different substrates, but it does not account for the capacity for rumen

microbes to adapt to the additives and degrade them (Benchaar and Greathead, 2011).

Additionally, dose rate in vitro can be significantly higher than when fed to the ruminant,

therefore a response can be induced that may not be practically feasible in a live animal

(Benchaar and Greathead, 2011).

Table 2: Summary of the top ten feed additives for methane reduction taken from the EFSA

report (Lewis et al., 2013)

n.b. these were all in vitro studies

Looking only at the substances with five or more studies and some in vivo experimentation,

we can ascertain which substances have the greatest evidence to reduce methane production

(Table 3). Tables 2 and 3 were used to compile a list of the most promising feed additives on

which to base the rest of the review (Table 4).

Substance group Substance Methane reduction (%)

and range in brackets

No. of

studies

Species

Essential oils Mentha

microphylla

-96 (-92 to -100) 1 Sheep

Plant extracts:

tannins

Rheum

officinale

(root)

-75 (No range) 1 Sheep

Fatty acids Linolenic

acid

-71 (-46 to -99) 3 Sheep (1) and

cattle (2)

Miscellaneous

chemical substances

9, 10-

anthraquinon

e

-70 (-58 to -91) 2 Cattle (1) and

goats (1)

Miscellaneous

chemical substances

2-

iodopropane

-61 (-25 to -97) 1 Cattle

Fatty acids Lauric acid -59 (-11 to -99) 4 Cattle

Plant and animal oil,

fats

Palm kernel

oil

-58 (-32 to -85) 2 Cattle

Miscellaneous

chemical substances

Pyromellitic

diimide

-57 (-7 to -100) 3 Cattle

Essential oils Carvacrol -56 (-13 to -98) 1 Sheep

Plant extracts:

general

Cashew nut

shell liquid

-55 (-36 to -70) 2 Cattle

Table 3: Summary of substances with five or more studies including some in vivo

experiments from the EFSA report (Lewis et al., 2013)

Substance

group

Substance Average

methane change

in vivo

Number of

in vivo

studies

Species Total

studies

Fatty Acids Myristic acid -36.3 2 Sheep 5

Plant & animal

oils, fats

Coconut oil -23.8 5 Cattle (4)

and Sheep

(1)

10

Plant & animal

oils, fats

Canola oil -16.0 3 Cattle 6

Essential oils &

Spices

Allium

arenarium oil

-12.0 1 Sheep 9

Organic acids

and their salts

Fumaric acid -11.5 2 Cattle and

Sheep

5

Antibacterial

drugs:

Ionophores

Monensin

sodium

-11.3 10 Cattle (9)

and Sheep

(1)

21

Plant extracts:

Saponins

Tea saponins -10.1 7 Cattle (3)

and Sheep

(4)

7

Plant extracts:

Saponins

Quillaja

saponaria

extract

-4.3 4 Cattle (3)

and Sheep

(1)

9

Plant extracts:

Saponins

Yucca

Schidigera

extract

-2.4 5 Cattle (4)

and Sheep

(1)

10

Organic acids

and their salts

Fumaric acid,

sodium salt

0.0 1 Cattle 12

2.3 Food and Agriculture Organization review

A recent report, Mitigation of Greenhouse gas emissions in livestock production; a review of

technical options for non-CO2 emissions, of the Food and Agriculture Organization of the

United Nations (Alexander N. Hristov et al., 2013) discussed the major feed additive groups

with potential to reduce methane as follows:

Inhibitors (i.e. targeted chemical compounds that inhibit rumen archea, e.g

bromochloromethane)

Verdict: Bromochloromethane is promising but cannot be used because it is an ozone

depleting substance, there was no sufficient data for other substances in this category

Electron receptors, e.g. nitrate

Verdict: Nitrate may be promising in low protein diets that may benefit from non-

protein nitrogen supplementation. An adaptation period is crucial and must beware of

pollution swapping (i.e. ammonia). Long term effects unknown.

Ionophores

Verdict: Likely to provide a moderate CH4 mitigating effect, prohibited in Europe.

Plant bioactive compounds (includes tannins, saponins, essential oils)

Verdict: hydrolysable and condensed tannins show promise but intake and animal

production may be compromised. Tea saponins require more long term studies.

Most essential oils or their active ingredients do not reduce methane. Studies that

demonstrated a reduction were over short timescales, therefore longer studies are

required.

Dietary lipids (also termed plant and animal oils)

Verdict: Effective at reducing methane but uptake will depend on cost-effectiveness

and potential negative effects on feed intake and productivity. Distillers grains were

also categorised here, but results have been variable, some increases in methane

observed due to increased fibre intake.

Exogenous enzymes

Verdict: May increase feed efficiency but inconsistent results.

Direct fed microbials

Verdict: Insufficient evidence, although yeast appears to stabilise pH and promote

rumen function in high concentrate diets.

Using the results of the two reviews, the most promising additives are summarised in Table 4.

Table 4: Most promising feed additives of the EFSA and FAO reports, as categorised by the

reports

Taking into account the findings of the EFSA and FAO reports, the next section will provide

further details of the mode of action of these additives, describe the practicality of feeding

them, health or environmental risks and effects on other pollutants.

Substance group Substances

Fatty Acids Myristic acid Linolenic acid, Lauric acid

(table 2)

Plant & animal oils, fats/dietary lipids Coconut oil, Canola oil, Palm Kernel oil

Essential oils and spices Allium arenarium oil, Mentha microphylla,

Carvacrol

Plant extracts: Saponins Tea saponins, Quillaja saponaria extract,

Yucca Schidigera extract

Electron receptors Nitrate, sulphate

2.3 The most promising feed additives for reducing methane

2.3.1 Fatty acids and plant oils

Table 5: Characteristics of the most promising fatty acids

Fatty acid Characteristics Main sources

Myristic acid Saturated,

Medium chain

Coconut oil,

Palm kernel oil

Lauric acid Saturated,

Medium chain

Coconut oil

Linolenic acid Polyunsaturated,

Long chain

Linseed

Linoleic acid Unsaturated, long

chain, omega 6

Soybean,

sunflower

Oleic acid Monounsaturated,

long chain

Rapeseed meal,

Canola oil

Medium-chain saturated fatty acids have strong antimicrobial properties (Hristov et al.,

2004), particularly affecting methanogenic archea (Machmuller et al., 2003). This inhibition

should reduce methane production. Conversely, it has been suggested that long chain fatty

acids do not have this mode of action (Mosoni et al., 2008).

Polyunsaturated fatty acids have toxic effects on cellulolytic bacteria and protozoa (Doreau

and Ferlay, 1995). They act against lactate producers (Kubo et al., 1993, Nagaraja et al.,

1997) thereby favouring propionate producers.

Coconut oil contains lauric and myristic acid. Both fatty acids have substantial potential to

reduce methane production (Tables 2 and 3) but there is a large range in effects observed and

lauric acid is only explored in vitro in the EFSA report. There is some indication that the

effect of the two fatty acids on methane production is additive (Machmüller and Kreuzer,

1999, Soliva et al., 2004).

Linolenic acid and linoleic acid also reduce methane by an average of 71% and 51%,

respectively, in vitro (Lewis et al., 2013) but further work is required in vivo.

Diet effect

Diet composition will affect the success of the additive, but results are not definitive.

Machmuller et al. (2003) investigated the interaction of myristic acid supplementation with

diet type (concentrate: forage ratios: 1 : 1·5 and 1 : 0·5) and dietary calcium (Ca) content (4.2

and 9g/kg DM) in sheep. Myristic acid suppressed methane in both diets, although more in

the concentrate diet than the forage diet and the highest dose of Ca reduced this effect.

When averaged across both diets, myristic acid increased DMI, as a result, methane

emissions reported in kg/megajoule (MJ) of gross energy intake and g/kg organic matter

digested were reduced for both diet types.

Lovett et al. (2003) showed no interaction between diet (forage to concentrate ratios: 65: 35,

40: 60 and 10: 90) and coconut oil supplementation in finishing beef heifers. Although

supplementing with coconut oil reduced dry matter intake, this was outweighed with greater

reductions in methane, therefore methane produced per kg DMI, liveweight gain, carcase

gain and as percentage of gross energy intake was reduced.

Martin et al. (2009) explored linseed supplementation in dairy cows on a hay- and maize

silage-based diet. In the four treatments (0, 5, 10 and 15% linseed in ration), the cows on the

maize silage diet produced less methane than those on the hay-based diet. The methane

reduction effect from linseed supplementation was greater on the hay-based diet than the

maize diet, perhaps this is due to a greater potential to improve rumen fermentation efficiency

in the hay based diet. 15% extruded linseed with a maize diet had the lowest total methane

emissions (l/d). The authors did not present results on dry matter intake, however, they found

that supplementation did not alter milk yield, suggesting extruded linseed (providing an

expected oil supplementation of 6% of the diet) is a viable methane mitigation option.

Feeding

The best way to feed dietary lipids of interest is unclear. Wholeseeds tend to be cheaper than

the refined oils (Martin et al., 2010), but sunflower seeds have been reported to reduce NDF

digestibility in heifers (Beauchemin et al., 2007) and average daily gain may be enhanced

with the oil form compared to whole seed/bean form (refined soy oil, crossbred beef bulls

(Jordan et al., 2006a)).

In terms of methane reduction, the oil form was better at reducing methane when feeding soy

(young bulls, Jordan et al. (2006a)) and linseed (lactating dairy cows, Martin et al. (2008))

than the whole form, but sunflower seed has been reported to reduce methane more than

sunflower oil (Aberdeen angus heifers, Beauchemin et al. (2007)). For many seeds, the hard

seed coating is difficult to digest by enzymes, therefore crushing, bruising, extrusion or

expansion is required to release the oil (Raes et al., 2004). These papers suggest the oil form

would be the most attractive option in terms of performance and methane reduction as long as

the cost of the ingredient is outweighed by improved performance.

Unintended consequences

Only five studies additionally looked at ammonia emissions in the EFSA report (three in vivo

and two in vitro), illustrating that lauric acid and myristic acid have potential reduce

ammonia emissions too (Lewis et al., 2013).

The fatty acids above are described as slight to strong skin and eye irritants and lauric acid is

toxic to aquatic life (Lewis et al., 2013).

Performance effects

Many studies report a reduction in methane with various types of fat supplementation to

ruminant diets but this is often accompanied by a reduction in dry matter intake and

digestibility (Table 6). When this is reported on a fat-corrected milk basis, increased feed

efficiency is observed in dairy production (Eugene et al., 2008, Rabiee et al., 2012).

Table 6 is adapted from a recent review of the benefits of supplementary fats in ruminant

rations (Rasmussed and Harrison, 2011). From this table, we can see that, apart from one

study, all the plant oils in the studies had potential to decrease methane. However,

digestibility and intake were reduced in some studies, particularly with high dose rates

therefore the negative effect on production is likely to increase methane output per unit of

liveweight gain.

Table 6: In vivo studies analysing the effects of plant oils on methane and other parameters (adapted from (Rasmussed and Harrison, 2011))

Animal n Durati

on

Ration (forage/concentrate dry

matter basis)

Fat source (% of

DM)

Methane

(%)

NH3 DMI Digestibility Reference

Charolais/Limousin

heifers

4

1

93d 50/50 Coconut oil

(10%)

-18 NS NS (Jordan et al.,

2006c)

250g/day

Copra meal*

(10%)

-14.9 NS decrease

250 g/day

coconut oil

Charolais/Limousin

heifers

1

6

35d 50/50 Coconut oil

(14%)

-20.5 NS NS (Jordan et al.,

2006b)

250g/day

Coconut oil

(42%)

-39 decreas

e

decrease

375g/day

Charolais/Limousin

bulls

3

6

103 d 10/90 Soya oil (10%) -40 NS (Jordan et al.,

2006a)

Soybean (12%) -25.3 decreas

e

Lactating cows 5 12 wk 70/30 Cotton seed

(48%)

-23 NS decreas

e

(Grainger et al.,

2010)

Lactating cows 1

6

4 X

28d

45/55 TMR Sunflower seed

(3.3%)

-10 increa

se

decreas

e

decrease (Beauchemin et

al., 2009)

Linseed oil

(3.3%)

-18 NS decreas

e

decrease

Rape seed (3.3%) -16 NS NS NS

Huzhou sheep 3

2

60 d 60/40 Soya oil (3%) -14 decre

ase

Decrease (Mao et al., 2010)

Swiss White Hill

sheep

1

2

3 x

21d

60/40 Coconut oil (6%) -26 NS (Machmüller et al.,

2000)

Sunflower seed

(6%)

-27 Decrease

Linseed oil (6%)

-10 Decrease

Table 6 continued: In vivo studies analysing the effects of plant oils on methane and other parameters (adapted from (Rasmussed and Harrison, 2011))

Animal n Durati

on

Ration (forage/concentrate) Fat source (% of

DM)

Methane

(%)

NH3 DMI Digestibil

ity

Reference

Holstein bulls 1

6

21 d 75/25 Sunflower oil

(5%)

-22 decrease (McGinn et al., 2004)

400g/d

Lactating Holstein

cows

3

6

319d 50/50 TMR Cotton seed (4%) NS increa

se

(Johnson et al., 2002)

Cotton seed

(5.6%)

NS increa

se

Rape seed (4%) NS increa

se

Rape seed (5.6%) NS increa

se

Lactating Holstein

cows

1

2

18 d 60/40 TMR Myristic acid

(5%)

-36 NS decrease (Odongo et al., 2007b)

Lactating Holstein

cows

8 4 wk 65/35 Linseed oil

(5.7%)

-64 decre

ase

decrease (Martin et al., 2008)

Extruded linseed

(5.7%)

-38 decre

ase

decrease

Crude linseed

(5.7%)

-12 NS decrease

Huzhou Sheep 8 21 d 60/40 Coconut oil (7%) -38 (Yuan et al., 2007)

*copra meal based concentrate with 250g of Coconut/day from copra meal

DMI = dry matter

intake

TMR = total mixed

ration

NS = no statistically significant effect

2.3.4 Essential oils

Essential oils cover a wide variety of products extracted from plants using steam distillation

with water or aqueous alcohol.

Mode of action

Similar to ionophores and polyunsaturated fatty acids, essential oils are thought to select

against gram positive bacteria, favouring propionate producing bacteria, thereby encouraging

the alternative hydrogen sink to methane – propionate.

Response

Within the category 'Essential oils', 42 substances were covered by the EFSA review and 31

were found to offer potential for reducing ruminal methane and ammonia, in many cases

reduction in these pollutants occurred simultaneously (Lewis et al., 2013). The most

promising essential oil with over 5 studies, that included in vivo work, was Allium arenarium

oil (garlic oil) which reduced methane by 12 % in vivo and an average of 36% reduction was

reported in vitro (Lewis et al., 2013).

Garlic oil is unique in that some active compounds present in the oil are not present in the

whole plant; they are formed during the steam treatment process (Pentz and Siegers, 1996).

It is active against a wide range of gram positive and gram negative bacteria, fungi, parasites

and viruses – this is likely due to interaction with the sulfhydryl groups (-SH) of other active

compounds (Reuter et al., 1996). The activity of the oil is more powerful than the individual

compounds alone – suggesting synergism between constituent compounds (Busquet et al.,

2005b).

Performance and practicality

Garlic and other essential oils could affect the palatability of the feed. Garlic oil and diallyl

disulphide (garlic oil constituent) additions to mature ewe diets did increase feed refusal

initially (Klevenhusen et al., 2011). With time, the sheep adjusted to diallyl disulphide

additions and feed intake resumed to the level of the control (no additive). This adjustment

did not occur with the garlic oil treatment.

Robertson et al. (2006) looked at improving the palatability of straw by adding different food

flavourings and found that applying a low rate of garlic flavouring (0.05 g/kg (0.005%)) to

straw improved acceptability to sheep. However, higher rates of garlic inclusion (5, 10, 15,

20, and 25% DM) reduced diet palatability (Nolte and Provenza, 1992a, Nolte and Provenza,

1992b).

Performance effects of added garlic to ruminant diets are variable, as illustrated in Table 7.

Diet effect

There are few studies investigating the interaction between diet and garlic supplementation.

One in vitro study suggests garlic oil additives are more effective with moderate concentrate

diets (50 : 50 alfalfa hay : concentrate ) than high concentrates (15 : 85 barley straw :

concentrate) at reducing the acetate:propionate ratios (Mateos et al., 2013). In both diets the

garlic oil reduced methane production significantly.

Risks

High rates of any plant extracts may be toxic, therefore dose will need careful consideration.

No adverse effects on the target species or human health (as typical usage doses) have been

identified for these substances.

The main barrier to the use of garlic in dairy diets it the risk of milk taint (Babcock, 1938),

but there is no evidence to suggest it will impact meat quality. In fact, Strickland et al.

(2011) found that lamb reared on diets containing garlic additives was accepted by taste

panellists more positively than those reared on garlic-free diets.

Table 6: Studies reporting the effect of garlic supplementation in ruminants

Form Dose Diet Animals Response Author

ADG Dry

matter

intake

FCE (DM

intake:kg

growth)

Methane Other

Garlic

oil

200mg/kg

dietary

DM

Barley based 10 ewe lambs/treatment NS NS NS NA Meat quality

measurements: NS,

Carcase

characteristics: NS,

except liver weight

(+20%)

VFAs, pH and

ammonia: NS

(Chaves et al.,

2008)

Garlic

extract*

250 mg/kg

BW/day/c

alf

Milk, forage and

concentrate (~1:6

conc:forage ratio)

12 pre-ruminant Holstein

cross calves, weaned just

after birth, 5 days old

+18 +8% NS NA Faecal score

decreased an average

of one score (four

point scoring scale),

NS change in feed

cost/kg gain

(Ghosh et al.,

2011)

Garlic

extract*

250 mg/kg

BW/day/c

alf

Milk, forage and

concentrate (~1:6

conc:forage ratio)

38 pre-ruminant Holstein

cross calves, weaned just

after birth, 5 days old

+44% +12% -44% NA Faecal score

decreased an average

of 0.8 score (four

point scoring scale),

30% reduction in feed

cost/kg gain

(Ghosh et al.,

2010)

Table 6 continued: Studies reporting the effect of garlic supplementation in ruminants

Form Dose Diet Animals Response Author

ADG Dry

matter

intake

FCE

(DM

intake:kg

growth)

Methane Other

Raw

Garlic

**

53 g/kg

dietary DM

(1.5% DMI)

Concentrate based Brown swiss x Limousin

crossbred bulls

NS NS NS NS (Staerfl et al.,

2012)

Raw

Garlic

Diet

inclusion:

0.9%

Pelleted ration

40 Merino wether lambs

aged 6 months

-18% NS NS NA Worm count: NS

(Strickland et al.,

2009)

1.8% -22% NS NS NA

3.6% -41% NS +62% NA

Garlic

oil***

4g/day Lucerne hay and

concentrate at

45:55ratio

Four ruminally fistulated

Baloochi lambs

NA NA NA NA No effect on organic

matter digestibility

or fibre digestibility

(Vakili et al.,

2011)

*prepared using electric mixer and filtered through muslin cloth, **acacia tannin, maca and lupines also compared, ***Tumeric powder and monensin also compared ADG = average daily gain, BW = body weight, DM = dry matter, DMI = dry matter intake, FCE = feed conversion efficiency, NA = not applicable to study, NS = no statistically significant

effect, VFA = volatile fatty acids.

2.3.5 Plant extracts: Saponins

Although there has been a lot of research interest in tannins and other plant extracts, saponins

were the only plant extract- based additive that had sufficient evidence described in the EFSA

review to reduce methane emissions.

Saponins are sugar and non-carbohydrate complexes that characteristically foam when

shaken in water. They are steroid or triterpene glycoside compounds found in a variety of

plants.

Commercial additives are derived from Yucca shidigera or Quillaya saponaria and Sapindus

sp (temperate and tropical plant species).

As Table 3 shows, in vivo effects of saponin addition in the studies for the EFSA report show

mean methane reductions of 2.4%, 4.3% and 10.1% for Yucca Schidigera extract, Quillaja

saponaria and Tea saponins, respectively.

Mode of action

In vitro and in vivo experiments show how saponins can be used to remove or reduce

protozoa numbers in the rumen (described in a review by Wina et al. (2005)). Protozoa cause

protein turnover by predating on bacteria. Theoretically their removal should lead to an

increase in the number of bacteria in the rumen, this slows the protein turnover leading to an

increase in bacterial N flow to the duodenum. Therefore, the removal of protozoa may

increase nitrogen utilisation leading to improvements in performance output and reductions in

ammonia produced. This theory has been supported by in vitro work of Makkar and Becker

(1996), whereby quillaja saponins (0.4-1.2 mg/mL) on a hay substrate increased efficiency of

microbial protein synthesis.

In addition, some methanogens are closely associated with protozoa, therefore the reduction

of protozoa may also reduce methanogen activity.

Diet effect

A review of studies by Wina et al. (2005) indicated that improvements in liveweight gain are

most likely to be seen with saponin addition to high roughage diets, although this was not

clear cut, some moderate to high roughage diets still showed no growth enhancing effect

(Hussain and Cheeke, 1995). Also, sex can interplay with the response: a higher growth

response was reported in male lambs compared to females lambs with 40 mg quillaja

saponins/kg in low roughage diets (Hussain and Cheeke, 1995).

Performance and practicality

The main issue with feeding saponins to ruminants is evidence that the rumen microbe adapt

and protozoa return to pre-supplementation levels (Newbold et al., 1997), intermittent

additions may mitigate this effect (Thalib et al., 1996). There is also evidence that they can

be degraded in the rumen (Wang et al., 2000). Further long term experimentation will need

to fully explore this effect and develop ways to overcome it.

Saponins have been described to reduce fibre digestibility but not total tract digestibility,

however acid detergent fibre (ADF) digestibility of a legume-grass diet was reduced and

neutral detergent fibre digestibility of a grass diet were reduced with addition of S.Saponaria

fruit (Abreu et al., 2004), further emphasising the important diet-additive interactions.

Risks

Some saponin containing plants are toxic which can lead to photosensitisation and subsequent

liver and kidney degeneration, and gut problems. However it is unlikely these would be used

in commercial additives.

2.3.6 Electron receptors

Electron receptors such as nitrate and sulphate were not considered in the EFSA review, yet

are prominent in other recent reviews on methane mitigation (Alexander N. Hristov et al.,

2013, Hristov et al., 2013). They offer an alternative a hydrogen sink to carbon dioxide,

thereby reducing methane emissions and potentially improving digestion efficiency

(Ungerfeld and Kohn, 2006). Nitrate (NO3−) likely holds the greatest potential for this

because it has a greater affinity for hydrogen (H2) than carbon dioxide therefore – instead of

methane – nitrite and ammonia are produced. Ammonia generated would supply fermentable

nitrogen for animals receiving protein deficient diets where ammonia is limiting protein

synthesis for microbial growth (Dijkstra et al., 1998). Additionally, the reaction to produce

nitrite is more energetically efficient than methanogenesis (Ungerfeld and Kohn, 2006).

Risks

Sudden introduction of nitrate into the diet may induce methaemoglobinemia, a state of

general anoxia; in mild cases this will depress animal behaviour but in severe cases can be

fatal (Ozmen et al., 2005). Methaemoglobinemia is caused by nitrite in the blood. The

accumulation of nitrite in the rumen is readily absorbed across the rumen wall and converts

blood haemoglobin (Hb) from the ferrous (Fe2+

) to the ferric (Fe3+

) form- methaemoglobin

(MetHb)- which is incapable of transporting oxygen (Morris et al., 1958). Gradual

introduction of nitrate into the diet can allow the rumen microbes to adapt and increase their

ability to reduce nitrite (Alaboudi and Jones, 1985). In animals unadapted to nitrate in their

diet, the capacity of the microbes to reduce nitrate to nitrite, exceeds the capacity for nitrite

reduction (Lewis, 1951). MetHb formation coincides with reduced oxygen intake, carbon

dioxide production and metabolic rate (Takahashi et al., 1998).

A comprehensive review suggests that sheep can tolerate up to 4% nitrate in high quality

concentrate or forage based diets without showing clinical signs of methamoglobinemia

(Leng, 2008).

Table 7: Studies reporting nitrate supplementation in ruminant diets

Paper Additive(s) Form Administration Dose Animals Diet Adaptation

period

(days)

Methanoglobin

>2% of

haemoglobin?

Methane

change (%

difference

in output

(L/day)

(Li et

al.,

2012)

Nitrate Calcium

nitrate

(1.9%

nitrate)

Dietary pellet 3% of DM 10 weaned

ewe lambs

(Poll dorset

sire x

Dohne

ewe)

Limited info. 7 No -34

(P=0.06)

(van

Zijderv

eld et

al.,

2011)

Nitrate Calcinit:

5Ca(NO3)

2NO3·10H

2O; 75%

NO3 in

DM

Added to

concentrates

8.8% of DM 20 lactating

holstein-

fresian

cows 33.2

±6.0kg

milk/d

104±58 d

in milk at

start

TMR-Corn silage

based ration

(66:34,

forage:concentrate)

Isoenergetic and

isonitrogenous

28 Yes, max.

observed:4.2%

of Hb

-16

(van

Zijderv

eld et

al.,

2010)

Nitrate and

Sulphate

5Ca(NO3)

2·NH4NO3

·10H2O;75

% NO3 in

DM

(highly

soluble

and

available

nitrate)

MgSO4

Added to

concentrates

2.6% of DM 20 male

cross bred

lambs

lambs

Maize silage basal

ration (90%) plus

concentrates based

on soybean mean

and additives

(10%).

Urea substitutes

nitrate in sulphate

experiment

28 Yes-max.

observed: 7% of

Hb for high

nitrate inclusion

diet

Nitrate -

32%

Sulphate -

16%,

Both -47%

Table 7 continued: Studies reporting nitrate supplementation in ruminant diets

Paper Additive(s) Form Administr

ation

Dose Animals Diet Adaptation

period

(days)

Methanoglobin

>2% of

haemoglobin?

Methane

change (%

difference

in output

(L/day)

(Nolan

et al.,

2010)

Nitrate Potassium

nitrate (KNO3)

Sprinkled

on hay

4% of DM 8 merino

wethers

rumen-

cannulated

sheep

Chaffed oaten hay

1kg/day

18 No -23

(Guo et

al.,

2009)

i) Nitrate-N

ii) urea N

iii) tryptone-N (a

pancreatic digest

of casein and a

source of amino

acids and

peptides)

Sodium nitrate

(NaNO3)

in vitro 12.6% DM 3

cannulated

Simmental

x Luxi

steers

60% roughage

(corn stover cubes

and alfalfa hay

pellets) and 40%

mixed concentrate

(ground corn,

soybean mean and

soy hulls)

NA Not measured -74

Table 7 continued: Studies reporting nitrate supplementation in ruminant diets

Paper Additive(s) Form Administr

ation

Dose Animals Diet Adaptation

period

(days)

Methanoglobin

>2% of

haemoglobin?

Methane

change (%

difference

in output

(L/day)

(Sar et

al.,

2004)

Nitrate with or

without:

Beta 1-4 galacto-

oligosaccharides

or Nisin

30%(w/v)

aqueous solution

of NaNO3/kg0.75

GOS-oligomate

55

Nisin-Sigma

Via fistula

30 mins

after

morning

meal

1.3g

NaNO3/kg0.

75 of body

wgt

GOS

20g/day

Nisin

3mg/kg0.75

of BW

4

ruminally-

fistulated

wethers

Chopped timothy

hay, alfalfa hay

cube, and

concentrate

(40:40:20 on DM

basis)

Fed at maintenance

energy level

7 Yes-peaked at

18.4% in

nitrate-only

treatment

-50

No diff.

with GIS

or nisin

(Alabo

udi and

Jones,

1985)

Nitrate Initially low

dose of nitrate

(NO3-) alone

(0.05% of BW)

then potassium

nitrate (KNO3-)

Added to

ration

2.5g

KNO3/kg

body

wgt/day

reached in

0.5g

increments

4 Dorset-

Columbia

crossbred

ewes

44% cereal grain,

50% hay

63 No NA

Unintended consequences

Nitrate may divert hydrogen away from other uses such as propionogenesis- therefore the

VFA profile often shifts away from propionate and butyrate in favour of acetate (Farra and

Satter, 1971, Allison and Reddy, 1984, Alaboudi and Jones, 1985). This could potentially

reduce the efficiency of the rumen as acetate production is less energy efficient than

propionate and butyrate production.

Additionally, it can lead to an increase in ammonia production, which can be a good source

of nitrogen for microbes but in high protein diets it is likely to be excreted, thereby producing

another pollutant.

Effect longevity

Although overall, potassium nitrate addition did reduce methane emissions in the study by

Nolan et al. (2010) (further details in table 10) after the immediate reduction observed post

feeding, there was a gradual rise in methane over the two hours between feeding periods.

The authors suggest a ‘slow release’ form of nitrate might enhance methane mitigation and,

in addition, reduce nitrate and nitrite absorption from the rumen and thereby reduce the

potential for nitrite toxicity.

More in vivo studies are needed to fully understand the impact of nitrate supplementation on

whole-farm GHG emissions (animal, manure storage and manure-amended soil), animal

production and animal health. The long-term effects of these compounds have

not been established.

3. Persistence

Of the studies considered in the EFSA review, very few studied the long term effects of

feeding additives. Where the mode of action depends on shifting the microbial population of

the rumen there is always potential for adaption of the microbes to take place over time and

the response to diminish, this issue was highlighted in a review by McAllister and Newbold

(2008). Where the mode of action is related to providing exogenous alternative hydrogen

sinks then there may be less potential for adaptation over time to occur.

Sauer et al. (1998) reported that ruminal microflora adapted over time to the use of

monensin, a view supported by Guan et al. (2006) who found that monensin reduced methane

production by around a third but this benefit was short-lived and methane production

gradually returned to its pre-supplementation level over a 16 week period. However, Odongo

et al. (2007a) studied the effects of monensin on methane production in steers and identified a

modest yet long term (6-months) reduction in methane of around 7%.

In a study with dairy cows on pasture, Woodward (2006) examined the effect of vegetable

and fish oils on milk production and CH4 emission in short- (14 days) and long-term (12

weeks) experiments. Lipids significantly decreased CH4 production in the short-term study,

but this effect was not observed after 11 weeks of feeding lipids in the long-term study.

Promising results in vitro demonstrating inhibition of methane production is not always seen

in vivo. Several processes in the live animal, such as: adaptation of the microbes to the

additive or degradation of the additive, toxicity for the host animal, negative effects on

overall digestion and productive performance can temper the results achieved in vivo

(VanNevel and Demeyer, 1996).

4. Conclusion

Several strategies have been explored to mitigate methane production using feed additives in

ruminants. Recent reviews suggest that plant extracts (dietary lipids, fatty acids, essential oils

and saponins) and some chemical substances (electron receptors) hold the most potential.

Condensed tannins and electron receptors (e.g., nitrate) have known toxicity risks to animals,

therefore dose and adaptation periods are necessary. All substances require more in vivo

studies over long time periods to determine whether the effect will be sustained.

Their successful implementation will depend on the cost-effectiveness and practicality in

commercial farm systems. Solutions must be evaluated in combination with other aspects of

livestock production.

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