GB2221829A

GB2221829A - Improvements in animal feed additives

Info

Publication number: GB2221829A
Application number: GB8916898A
Authority: GB
Inventors: William Leslie Porter
Original assignee: Individual
Current assignee: Individual
Priority date: 1988-08-19
Filing date: 1989-07-24
Publication date: 1990-02-21
Anticipated expiration: 2009-07-24
Also published as: GB2221829B; GB8819718D0; GB8916898D0

Abstract

In a method of manufacture of an animal feed additive, non-pathogenic bacteria are grown in a liquid culture, the bacterial cells are isolated from the bulk of the fluid substrate and are killed before or after isolation, for example by application of heat, and the isolated killed cells are combined with a carrier to provide a solid or liquid product ready for use as an animal feed additive. The carrier may be wheat bran; finely ground cereal, expanded mica.

Description

Improvements in Animal Feed Additives This invention relates to an animal feed additive.

Known probiotic products for oral administration to animals by inclusion in feed consist of fermented cultures of nonpathogenic bacteria, typically lactobacilli and/or streptococci.

Probiotics are used to improve rate of growth and feed conversion efficiency in young animals and to reduce incidence of enteritis and diarrhoea. However, the effects of such products are highly variable.

The essential mode of action of probiotics is not, in fact, understood. It is commonly thought that they enhance the numbers of non-pathogenic bacteria in the gut, which may assist digestion, possibly through enzyme production, and also increase competition with populations of potentially harmful bacteria such as Esoherichia ccli. However, this is not proved. In fact, the normal dosage of a probiotic is usually insufficient to add appreciably to the intestinal population and this casts doubt on the commonly believed mode of action referred to.

Most importantly, since the mode of action is not in fact sufficiently understood, meaningful quality control of probiotic products is virtually impossible.

A principal object of this invention is to provide an animal feed additive and economical method of manufacture thereof based on improved knowledge of the source of the beneficial effects of cultures of non-pathogenic bacteria.

This permits meaningful assay and hence meaningful quality control.

According to one aspect of the invention, there is provided an animal feed additive comprising a carrier supporting killed non-pathogenic microbial cells, the killed cells having been produced by growing the non-pathogenic microbial cells in a liquid culture, isolating the said cells from the bulk of the fluid substrate and killing the cells either before or after isolation in a manner which causes alteration of one or more chemical components of the cells.

According to another aspect of the invention, there is provided a method of manufacturing an animal feed additive which comprises the steps of:- a) growing non-pathogenic microbial cells in a liquid culture; b) isolating the microbial cells from the bulk of the fluid substrate; c) killing the cells in a manner which causes alteration of one or more chemical components of the cells, either before or after step b); and d) combining the killed cells with a carrier to provide a product ready for use.

According to still another aspect of the invention, there is provided an animal feed additive comprising a carrier supporting killed non-pathogenic bacterial cells, the killed cells having been produced by growing the nonpathogenic microbial cells in a liquid culture, isolating the said cells from the bulk of the fluid substrate and heating to a sufficient temperature to kill the cells either before or after isolation.

According to yet another aspect of the invention, there is proivded a method of manufacturing the animal feed additive, which comprises the steps of:- a) growing non-pathogenic bacterial cells in a liquid culture; b) isolating the cells from the bulk of the fluid substrate; c) heating to a sufficient temperature to kill the cells either before or after step b); and d) combining the heat killed cells with a carrier to provide an animal feed supplement ready for use.

Fermentation for at least 24 hours appears necessary to produce a sufficient cell content. The microbial cells are preferably bacterial cells, more especially lactobacilli or streptococci, in particular streptococcus faecium, or Bacillus subtilis or bifidobacterium or micrococci or pediococci, and the cells are preferably killed after isolation from the bulk of the fluid substrate.

It will be understood that, conventionally, probiotic products are products containing live bacterial cells.

The product in accordance with the present invention contains only bacterial cells killed by a method or means, in particular application of heat, which as far as is understood causes alteration and possibly denaturation of one or more chemical components of the cells, but being a product intended for use as an animal feed additive in analogous manner to certain known probiotics.

The animal feed supplement of the present invention can take several forms. Thus, for example, the killed and altered cells, in the form of a wet slurry, can have a bacteriostat added thereto for storage and administration in the wet state. Alternatively, the killed and altered cells can be dried and mixed with wheat bran or finely ground cereal to form the additive. Yet again, the killed cells can be absorbed in a drying agent, such as expanded mica, to produce a concentrated animal feed additive.

Preferred methods of killing the cells in the required manner which is believed to cause alteration of one or more chemical components of the cells comprise:1) Application of heat to a temperature not less than 65eC preferably at least 700C and even more preferably at least 800C or 900C and up to and including 1000C at normal pressure, or still higher temperatures at elevated pressures, as by autoclaving. Lower temperatures require more prolonged periods of heating.

2) Application of heat by radiation, such as microwaving.

Possible but less preferred methods are:3) Chemically by treatment with substances which effect alteration or denaturation, which substances may include: (A) a halogen such as iodine, bromine or chlorine, (B) an acid such ås performic acid, (C) an alkali, (D) alcohols and/or phenolics, (E) salts such as sodium chloride, zinc sulphate and/or ammonium sulphate, (F) specific denaturing chemicals such as guanidine hydrochloride, urea, betamercapto ethanol, and/or enzymes. Performic acid previously referred to is also an example of such a specific chemical.

The animal feed additive in accordance with the invention has been tested on chicks in trials lasting 14 days or more. Different groups of chicks were fed a basic diet as a control diet and other diets having allegedly beneficial additives, the additive for one group being the probiotictype product in accordance with the invention. Trials were initially carried out using a commercial broiler feed as a control diet, to assess the effects of differing allegedly beneficial additives. Additives used were live bacterial probiotics known by the Trade Marks Pronifer and Maxipro.The growth and feed conversion efficiency of groups of chicks receiving these products were compared with similar groups of chicks receiving the same originally live bacterial products, but where the products had been subject to autoclaving, i.e. a treatment to kill and alter one or more chemical components of the bacterial cells present in the products. It was found that substantially improved results were obtained when the live bacterial probiotics were so treated as to kill the cells in the specified manner. Further trials were then carried out in which bacterial cultures were used as additives, both as whole cultures and after centrifugal separation into the bulk liquid substrate and the cellular concentrate or residue of killed cells. The results were compared against the same control diet as before and also the control diet with added killed yeast cells.

The statistical results of these trials have shown that, especially after a full trial of at least 14 days, the best liveweight gain and best feed conversion ratio is to be expected from the cellular concentrate or residue containing killed and altered cells.

The cellular concentrate thus has, to an unexpected extent, nutritive benefit to the growth of the animals.

Quality control is enabled firstly because the cellular content of the bacterial culture may be determined in conventional manner before autoclaving, application of heat or other treatment, for example colony counting techniques using plate cultures, and secondly because an index of the content is readily obtained either by dry matter estimation of the culture or the concentrated slurry or by using acid or gas production as parameters of bacterial growth.

More generally, various advantages arising from use of the probiotic-type product in accordance with the invention are as follows:- a) the total cell mass can be estimated and related to the effectiveness of the product; b) stability is improved over conventional probiotics as no live bacteria are present; c) the low volume of the cellular concentrate enables production of a solid, powdered or granular material, when this is required for inclusion in dry animal feeds; d) liquid products can be manufactured using the killed and altered bacterial cells, since the matter is inert (it is difficult to keep cells alive for long periods in a liquid); e) the liquid substrate is a by-product of the method of manufacture which, when used as a bulk liquid feed, is also found to possess useful nutritional properties.

A practical animal feed additive and method of production thereof in accordance with the invention will now be described by way of example.

First, one exemplary formulation of a base culture will be given: Milk powder 60 g/l Yeast extract 4 g/l Sucrose 10 g/l Beef and vegetable extracts 1 g/l Vitamin B12 5 mg/l Magnesium sulphate 0.2 g/l Manganous sulphate 0.05 g/l -'Tween 80' (Trade Mark) 1 m/l To the above formulation may be added: D. Potassium H. Phosphate 2.0 g/l Sodium acetate 5.0 g/l Tri-ammonium citrate 2.0 g/l, more especially to act as buffers if pH is not otherwise controlled during the subsequent fermentation process.

The medium is made up to one litre with de-ionised water.

Production of the animal feed additive is then carried out generally in accordance with the following'steps:- 1. The base culture is mixed using a shearing-type mixer.

2. The culture is sterilised by autoclaving at a pressure not less than 5 p.s.i., conveniently for 15 minutes at 121 degrees C. The culture could be sterilised in other known ways, if desired, which will ensure that any contaminants are killed.

3. The culture is cooled or allowed to cool to about 41 degrees C.

4. The base culture is inoculated with a small volume of an active starter culture, more especially but not exclusively a commercially available strain of a lactobacillus culture (such as Lactobacillus fermentum) or a streptococcus faecium culture or bacillus subtilis culture or a bifidobacterium culture.

5. The culture is allowed to ferment, at a temperature of about 41 degrees C, for a period of about 72 hours, whilst being subjected to gentle agitation.

6. During the fermentation step, a pH value of between 5 and 6, conveniently about 5.5, is maintained by means of NaOH and/or by means of the buffers previously referred to.

If, as is preferred, pH is controlled by use of sodium hydroxide, this mav be added initially, and then automatically, responsively to the output of a pH sensor.

7. During the fermentation process, gas production is monitored with a gas flowmeter, acid production is monitored by titration, and optionally cell production may be monitored by cell counting.

8. At the end of the fermentation period, the fermented culture is cooled or allowed to cool to ambient temperature.

Settling occurs during this period of cooling.

9. The supernatent liquid is then syphoned off. This leaves a residue in the form of a slurry containing the cells produced by fermentation.

10. The slurry residue is mixed by a shearing-type mixer.

11. The slurry is then subject to application of heat, which may be by direct application of heat, by microwaving or by autoclaving. A minimum temperature of at least 70 degrees C for a prolonged period is necessary to alter the cells in the required manner. The heating period can be reduced at higher temperatures, for example to the extent that autoclaving at 5 p.s.i. at about 120 degrees C for about 20 minutes is sufficient to achieve the desired result.

12. The killed cell slurry is again mixed by a shearingtype mixer.

13. The killed cell slurry is then dried, as by freeze drying or addition of a bacteriostat and mixing with a drying agent. Again, spray drying, centrifugation, vacuum evaporation or other drying by use of heat may be employed instead.

The The resulting killed cell residue in dried condition is combined with a carrier and constitutes an animal feed additive ready for use.

Alternatively te steps 13 and 14, a flocculating agent may be employed to effect separation of the cells, reducing the original volume by at least 80 per cent. A predetermined quantity of the resulting killed cell concentrate may then be mixed whilst simultaneously drying into a given amount of wheat bran, crushed barley or other cereal which is then ground to produce an animal feed additive.

It is alternatively possible to produce a liquid product, again suitable for use as an animal feed supplement, by mixing the wet residue of step 12 or the dried residue of step 13 with a liquid carrier together with a bacteriostat, although a solid product is preferred for inclusion in dry feed.

A cellular concentrate containing only autoclaved cells of Lactobacillus fermentum and added to a basic control diet has been tested on broiler chicks over a period of O to 21 days of age with the following results. In the table, the column headings designate the number of parts by weight of cellular concentrate added to one million parts of weight of control feed, and the successive rows in the table indicate weight gain (WG) in grams, weight gain as a percentage of control (WG%), the feed conversion ratio (FCR) and the feed conversion ratio as a percentage (FCR%), taking the FCR for control as 100.

Table 0 50 100 200 400 WG 335.25 381.08 401.81 390.52 371.36 WG% 100 113.67 119.85 116.49 110.77 FCR 2.963 2.216 2.096 2.139 2.541 FCR% 100 74.79 70.74 72.19 85.75 A prototype commercial food additive produced substantially by the method hitherto described has been tested on broiler chicks over a period of O to 18 days. The control diet used for this test was a commercially available mix containing wheat (63.4), soya extract (25.0), full fat soya (3.6), dicalcium sulphate (2.-0), DL-Methionine (0.22), salt (0.2), Minvite 204 (0.5) and soya bean oil (4.8), with added choline chloride (Q.5), the figures being percentages.

In the following table, the column headings indicate grams/ ton of additive employed, first when the fermented slurry residue was autoclaved and second when this residue was boiled.

Table Control Autoclaved Boiled Approx.30 g/tonne Approx. 30 /tone WG 360.17 381.75 376 WG% 100 105.99 104.04 This result indicates that boiling is nearly as effective as autoclaving as a means of processing the bacterial cells.

It is also important to note that trials with broiler chicks have been conducted in which WG and FCR were compared when a) the cellular concentrate was used as a food additive, b) the bulk liquid substrate emerging as a by-product of the process was used as an additive, c) the bulk fermented fluid autoclaved but without separation of the bulk fluid substrate was used as an additive, d) the bulk dried culture without separation and autoclaving was used as an additive, and e) a concentrate of cells killed and ruptured by a combination of mechanical shearing and sonication, but without boiling or autoclaving. These trials have shown that a substantially improved WG and FCR are obtained with the cellular concentrate of cells killed by application of heat, as compared with any of the other additives.It is therefore to be understood that process step 9) of the above-described method of production, in which the bulk liquid substrate is syphoned off or otherwise removed, is an essential step in production of the animal feed additive in accordance with the invention. Clearly, step 11 of the process, which involves killing the cells by application of heat, is also essential.

The identity of the substance obtained from bacterial cells killed by application of heat and having considerable beneficial effect on animal growth and feed conversion efficiency is not yet known. However, test for antibiotic activity have proved negative.

It will be understood that various modifications of the above-described method of production are possible within the scope of this invention. For example, the bacterial cells may be killed, instead of by autoclaving or boiling, by microwaving.

It should also be made clear that the feed additive of the invention is useful for administration to animals other than chicks, such as pigs and other monogastrics, as well as ruminants.

Claims

1. An animal feed additive comprising a carrier supporting killed non-pathogenic microbial cells, the killed cells having been produced by growing the nonpathogenic microbial cells in a liquid culture, isolating the said cells from the bulk of the fluid substrate and killing the cells either before or after isolation in a manner which causes alteration of one or more chemical components of the cells.

2. A method of manufacturing an animal feed additive which comprises the steps of:- a) growing non-pathogenic microbial cells in a liquid culture; b) isolating the microbial cells from the bulk of the fluid substrate; c) killing the cells in a manner which causes alteration of one or more chemical components of the cells, either before or after step b); and d) combining the killed cells with a carrier to provide a product ready for use.

3. An animal feed additive comprising a carrier supporting killed non-pathogenic bacterial cells, the killed cells having been produced by growing the nonpathogenic microbial cells in a liquid culture, isolating the said cells from the bulk of the fluid substrate and heating to a sufficient temperature to kill the cells either before or after isolation.

4. A feed additive according to claim 1 or claim 3, wherein the bacterial cells include at least one of lactobacilli, streptococci, bacillus faecium, bifidobacillus, micrococci and pediococci.

5. A feed additive according to claim 1 or claim 3, comprising a wet slurry containing the heat killed cells and with a bacteriostat added thereto.

6. A feed additive according to claim 1 or claim 3, incorporating the heat killed cells in a dried state.

7. A method of manufacturing the animal feed additive of claim 1, which comprises the steps of:- a) growing nonpathogenic bacterial cells in a liquid culture; b) isolating the cells from the bulk of the fluid substrate; c) heating to a sufficient temperature to kill the cells either before or after step b); and d) combining the heat killed cells with a carrier to provide an animal feed supplement ready for use.

8. A method according to claim 2 or claim 7, wherein the cells are killed by one of autoclaving, boiling or micro waving.

9. A method according te claim 2 or claim 7 or claim 8, wherein the cells are killed by application of heat after separation of the bulk fluid substrate.

10. A method according to claim 2 or claim 7 or claim 8 or claim 9, wherein the heat killed cells are dried prior to combination with the carrier.

11. A method according to claim 2 or claim 7 or claim 8 or claim 9, wherein the heat killed cells are combined with a bacteriostat to form a liquid animal feed additive.

12. A method according to claim 2 or claim 7, including the step of estimating the cellular content of the heat killed cell concentrate to enable quality control