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ORGANIC ACIDS AND ESSENTIAL OILS

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ORGANIC ACIDS AND ESSENTIAL OILS Powered By Docstoc
					           ORGANIC ACIDS AND ESSENTIAL
        OILS, A REALISTIC ALTERNATIVE TO
       ANTIBIOTIC GROWTH PROMOTERS IN
                                POULTRY
                                                                              ROBERT GAUTHIER D.V.M., DIPL. ACPV
                                                                                   JEFO NUTRITION INC., CANADA
                                                                                                              rgauthier@jefo.ca

        INTRODUCTION
        Since 1967, the year of the Swann report in England, the use of antibiotics in farm animals as a growth promoter has been
      questioned. More recently, in the late 80’s and early 90’s, strong regulatory actions have removed most of the antibiotic
      growth promoters from the European Union market, the last ones will be removed in January 2006.
        The adjustments following the withdrawal of these products in animal production have been difficult at times and many
      replacement solutions have been proposed, more or less successfully, by the feed additive industry. It is not easy to replace
      products that have proven to be generally efficacious for the last 50 years.
        A consensus seems to develop among the scientific community concerned by this subject [1] and one approach is definitely
      standing out, for its efficacy, technological and economical feasibility. We are talking about organic acids. Another option is,
      under the generic name of “botanicals”, essential oils (plant extracts or related compounds).

        ORGANIC ACIDS (OA)
        Organic acids have been used successfully in pig production for more than 25 years and continue to be the alternative of
      choice. Even if much less work has been done in poultry [2], we can confirm today that the OA are very efficacious provided
      their use is adapted to the physiology and anatomy of poultry.




        Organic acids (C1-C-7) are widely distributed in nature as normal constituents of plants or animal tissues. They are also
      formed through microbial fermentation of carbohydrates mainly in the large intestine [5]. They are also found in their sodium,
      potassium or calcium form.




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  THE MODE OF ACTION OF ORGANIC ACIDS
  Over the years, it was thought that a pH reduction of the GIT content was the mode of action. Research has proven di-
fferently and it is what we will review more precisely. Research in the food preservation field has brought clear explanations
on the mode of action of organic acids on bacteria and numerous trials have shown that the concept works both in pigs and
poultry.
  The mode of action of organic acids on bacteria is related to: [7]
  •       Undissociated organic acids entering the bacteria cell
  •       Bacteria membrane disruption (leakage, transport mechanisms)
  •       Inhibition of essential metabolic reactions (ex. of glycolysis)
  •       Stress on intracellular pH homeostasis (normal bacteria pH is ± neutral)
  •       Accumulation of toxic anions
  •       Energy stress response to restore homeostasis
  •       Chelation as permeabilizing agent of outer membrane and zinc binding
  The key basic principle on the mode of action of organic acids on bacteria is that non-dissociated (non-ionized) organic
acids can penetrate the bacteria cell wall and disrupt the normal physiology of certain types of bacteria that we call “pH
sensitive” meaning that they cannot tolerate a wide internal and external pH gradient. Among those bacteria we have E. coli,
Salmonella spp., C. perfringens, Listeria monocytogenes, Campylobacter spp.

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        Upon passive diffusion of organic acids into the bacteria, where the pH is near of above neutrality, the acids will dissociate
      and lower the bacteria internal pH, leading to situations that will impair or stop the growth of bacteria.
        On the other hand, the anionic part of the organic acids that cannot escape the bacteria in its dissociated form, will accu-
      mulate within the bacteria and disrupt many metabolic functions and lead to osmotic pressure increase, incompatible with
      the survival of the bacteria.




        It has been well demonstrated that the state of the organic acids (undissociated or dissociated) is extremely important to
      define their capacity to inhibit the growth of bacteria. As a general rule, we need more than ten to twenty times the level
      of dissociated acids to reach the same inhibition of bacteria, compared to undissociated acids [8].




        Too often, “in vitro” assays showing the antibacterial capacity of organic acids are done at a low pH, making sure that the
      acids are not dissociated. At a pH below 3.0-3.5, almost all organic acids are very efficacious in controlling bacteria growth.
      This does not reflect at all what is happening in the GIT of poultry and pigs.




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  Many authors have studied the effects of organic acids on animals, trying to find an explanation on their mode of action
as a growth promoter.
  Their findings are more related to experiments in pigs but could be partially extrapolated to poultry. Among the explanations,
some are still believing that the gastrointestinal tract (GIT) content pH change is important even if recent and not so recent
publications [9, 10] are showing differently, even with very high acid levels in the feed or water. Many are underestimating
the capacity of the animal to maintain its GIT environment homeostasis in order to warrant the normal functioning of all
digestive functions. Also the strong buffering capacity of the feed prevents any significant GIT pH modification.
  Logically, organic acids added to feeds should be protected to avoid their dissociation in the crop and in the intestine (high
pH segments) and reach far into the GIT, where the bulk of the bacteria population is located.




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        Other modes of action have been investigated mainly in pigs and cannot apply to poultry; increased digestibility of protein
      and energy, slower emptying of the stomach (crop – gizzard ), modification of the fermentation pattern in the intestine (mostly
      the large intestine), stimulation of enzyme secretion, modification of the enterocyte development. All these observations were
      made in trials using extremely high levels of organic acids incompatible with poultry nutrition [5]. Even the modification of
      the fermentation pattern in the intestine, which is important in pigs, may not apply to poultry because the relative impor-
      tance of fermentation bacteria in domesticated poultry is much less than in pigs or other mammals. High intake of organic
      acids could be harmful to animals. It has been shown that organic acids may lead to a reduction in bone mineral deposition
      in piglets [11] and many veterinary clinicians are reporting cases of bone decalcification, both in pigs and poultry.
        More likely, the organic acids in poultry might play a direct role on the GIT bacteria population, reducing the level of some
      pathogenic bacteria (ex. C. perfringens) and mainly controlling the population of certain types of bacteria that compete with
      the birds for nutrients. [12]. After 50 years of usage of antibiotic growth promoters in poultry, there is still a lot of speculation
      on their mode of action.
        From the use of organic acids in poultry we expect an improvement in performance similar or better than the antibiotic
      growth promoters, without the public health concern, a preventive effect on intestinal problems like necrotic enteritis and a
      reduction of the carrier state for Salmonella spp. & Campylobacter spp.

        ESSENTIAL OILS (EO)
        Contrary to organic acids, a wealth of research has been done on the use of essential oils, herbs, botanicals in poultry
      production, both as a growth promoter or a disease prevention product.
        The scientific and popular press use a lot of different names (plant extracts, phytogenic additives etc.) and to better un-
      derstand what we are working with, let’s have some definitions.
        •        Essential oils: Are any of a class of volatile oils obtained from plants, possessing the odour and other characteristic
      properties of the plant, used chiefly in the manufacture of perfumes, flavours and pharmaceuticals (extracts after hydro-
      distillation) [13].
        •        Herbs: Are a flowering plant whose stem above the ground does not become woody and persistent. A plant when
      valued for its medical properties, flavour, scent or the like [13].
        •        Botanicals: Are drugs made of a plant, as from roots, leaves, barks etc. [13].
        Essential oils or plant extracts can be used as appetite stimulant, aroma, stimulant of saliva production, gastric and pan-
      creatic juices production enhancer and antioxidant. However there is no clear demonstration of the importance of these




152
factors on the chicken performance.
   However the antibacterial property of essential oils is the most widely studied area, both in human nutrition, food preserva-
tion and animal production. Because the control (modulation) of the GIT microflora is the most important aspect in replacing
antibiotic growth promoters, we will concentrate on this aspect.
   Plants contain hundreds of substances having different properties but essential oils composed mainly of nine groups (and
many sub-groups) of molecules are of interest to us. There are many chemical constituents but no two oils are alike in their
structure and effect.
   One must make a difference between non purified plant extracts containing numerous different molecules interacting
and pure active compounds, either extracted from plants or synthesized (nature identical). According to the plant chosen,
one or more active compounds are dominant and the quantity found will differ according to factors like; plant variety, soil,
moisture, climate, time of harvest etc..
   It is counter productive to test every plants that can have interesting properties, concentrating on the active compounds
and selecting the right plants or the right synthetic molecules is easier and will be more acceptable on a regulatory point
of view.
   Nutritionally, metabolically and toxicologically, we have a clear interest in using as low as possible levels of essential oils
in animal nutrition. Essential oils are extremely potent substances, they can lead to feed intake reduction, GIT microflora
disturbance, accumulation in animal tissues and products.
   Most essential oils are GRAS (generally recognized as safe) but they must be used cautiously because they can be toxic
(allergens) and potent sensitizers and their odour/taste may contribute to feed refusal [14, 15]. They are also very volatile
and will evaporate rapidly, leading to large variation in concentration in the finished products. Encapsulation of essential
oils could solve the problem [14].
   The “in vitro” level of most of the essential oils needed to reach a MIC on various bacteria is high and not applicable in
animal nutrition. The number of EO really interesting and efficacious as alternative to antibiotic growth promoters is very
limited.




  THE MODE OF ACTION OF ESSENTIAL OILS
  It is extremely difficult to generalize on the mode of action of essential oils on bacteria and yeasts because each EO has
different properties and each type of microorganism has a different sensitivity. Generally, Gram+ bacteria are considered
more sensitive to EO than Gram- bacteria [14] because of their less complex membrane structure.
  The consensus on the mode of action of EO on bacteria is now that these compounds influence the biological membranes
of bacteria. The cytoplasmic membrane of bacteria has two principal functions [17]:
  A: barrier function and energy transduction, which allow the membrane to form ion gradients that can be used to drive
various processes.

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        B: formation of a matrix for membrane-embedded proteins influencing the ATP-synthase complex.
        In a study on the mode of action of carvacrol on Bacillus cereus, Ultee et al. [17] have shown many aspects of the mode
      of action of EO.
        • Carvacrol inhibits the growth of the bacteria effectively, it is a function of concentration, temperature and time.
        • A sharp reduction of the intracellular ATP pool through a reduction of ATP synthesis and increased hydrolysis, not obviously
      related to an increase in membrane permeability.
        • Reduction of the membrane potential (transmembrane electrical potential) which the driving force for ATP synthesis. The
      membrane becoming more permeable to protons.
        • Reduction of the bacteria internal pH with high level of carvacrol (1mM, pH from 7.1 to 5.8) related to ion gradient
      across cell membrane.
        • Potassium efflux; 1 mM of carvacrol reduced internal bacteria K level from 12 µmol/mg of cell protein to 0.99 µmol in
      5 minutes. K plays a role in the activation of cytoplasmic enzymes, in maintaining osmotic pressure and the regulation of
      cytoplasmic pH. K efflux is the first indication of membrane damage.
        Lambert et al.[15] report that the mode of action of EO is related to an impairment of a variety of enzyme systems, mainly
      involved in energy production and structural components synthesis. They also explain the mode of action through leakage
      of ions, ATP, nucleic acids, amino acids. They have shown that potassium and phosphate ion concentrations are affected at
      levels below the MIC concentration.
        It is interesting to note that most of the levels used “in vitro” to determine MICs are higher than the levels considered
      acceptable in animal nutrition.

        A REALISTIC ALTERNATIVE TO ANTIBIOTIC GROWTH PROMOTERS (AGP)
        Both OA and EO (mostly non purified plant extracts) have been tested as AGP alternative, alone or in combination. Results
      are lacking consistency due to the varied composition of products, varied growing conditions during the trials (environment,
      feeds, health etc.).
        In our own experiments with OA, we have experienced very consistent results, both under research station and field conditions,
      our rate of positive response exceeded 90% for weight gain and feed conversion, using a blend of protected organic acids.




154
  Not only protected organic acids can act as growth promoter but also play a role in the prevention of necrotic enteritis
and in the reduction of intestinal Salmonella spp. It appears that the amplitude of the response is often related to the level
of contamination or intestinal disease challenge in the flock.
  Plants, plant extracts (with their essential oil components) or pure EO have been tested quite extensively in many countries
with very high result variability. This could be explained by the lack of consistency of the products themselves or simply by the
lack of efficacy of certain molecules. As previously outlined, the level of EO needed to achieve results is often not compatible
with the level allowing the best feed intake. This leads to using levels that are not efficacious.
  Single EO or blend of EO, at low level, are difficult to justify economically and zootechnically.
  More and more, the concept of combining EO and OA is proving to be efficacious [18, internal data] because there appears
to be a synergy between the two concepts [19, 20, internal data].
  Our own experiments in field trials or when using a chicken necrotic enteritis challenge model have shown a strong synergy
between EO and OA.
  Some authors suggest [20] that the EO are damaging the bacteria cell membrane facilitating the penetration of organic
acids into the bacteria cytoplasm. This hypothesis has not been demonstrated clearly yet. Usually the bacteria cell membrane
damage leads to an efflux of material from the bacteria, not an influx into the bacteria. However it has been shown that a
damaged bacteria cell membrane allowed the uptake of a nuclear fluorescent stain, binding to cellular DNA and RNA [15].
The question raised is; are the organic acids behaving like ethidium bromide fluorescent nuclear stain?




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       OVERCOMING THE TECHNICAL PROBLEMS OF USING ORGANIC ACIDS AND ESSENTIAL
      OILS
        The use of organic acids and essential oils in the feed industry are often a source of problems; corrosion, worker’s safety,
      handling, vitamin stability in premixes, environmental concern, stability of products.
        It has been demonstrated that when both OA and EO are protected in a special matrix, the quantity required to achieve
      maximum performance in poultry can be reduced drastically. The active ingredients can be delivered into the intestine, directly
      where the bulk of gastrointestinal bacteria are located [21].
        Without protection, organic acids are readily dissociated in the first part of the chicken GIT and are rendered useless [2].
      Essential oils are very rapidly absorbed in the duodenum and cannot interact with the microflora.

        CONCLUSION
        There is a general consensus on the efficacy of organic acids as the best alternative to antibiotic growth promoters.
        Essential oils have a limited effect as a replacement of antibiotic growth promoters but they can act in synergy with organic
      acids both for their growth promoting effect and prevention of specific intestinal diseases.
        Now we have an encapsulation technology that enhances the efficacy of organic acids and essential oils, at low level of
      inclusion.

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