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`US. PATENT DOCUMENTS
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`4,129,578
`4,138,498
`4,159,322
`4,405,609
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`1211978 Celmeretal.
`. 260845.? R
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`2.!1979 Das ...........
`426m?
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`424MB}
`611979 Cloyd
`9.31983 Potter
`424! 1??
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`4,695,466
`4.761.426
`5,104,662
`5,244,669
`5,244,681
`
`9f198? Morishita at a].
`8.11988 Marlin el al.
`#1992 Kalsta et a1.
`911993 Saluh etal.
`911993 Vlnci el al.
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`424f456
`51411460
`4241451
`4244438
`42672
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`US. Patent
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`Mar. 26, 1996
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`FIGURE 1
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`US. Patent
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`Mar. 26, 1996
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`Sheet 2 of 7
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`5,501,857
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`FIGURE2d
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`FIGURE2b
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`CPU/Bolus(logarithmicscale)
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`VIABLE BACTERIAL COUNTS PER BOLUB
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`Bolus #
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`FIGURE 3
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`1
`ORAL NUTRITIONAL AND DIETARY
`COMPOSITION
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`5,501,857
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`2
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`This application is a continuation-in-part of U.S. patent
`application Ser. No. 07(920,104 filed Jul. 24, 1992, now US.
`Pat. No. 5,310,555.
`
`TECHNICAL FIELD
`
`This invention relates generally to veterinary nutritional
`and dietary compositions for livestock, and specifically to
`compositions which combine incompatible substances such
`as nutrient supplements and viable direct-fed microbials.
`The invention also relates to a method of preparing a
`shelf-stable
`composition of
`incompatible agents. The
`present invention is particularly well suited to deliver in
`vivo, nearly simultaneously, incompatible supplements such
`as vitamins and minerals and microorganism to cattle,
`sheep, goats, ostriches, and coins in the form of a capsule-
`in-a—capsule.
`
`BACKGROUND OF THE INVENTION
`
`In the past, the development of effective treatments for
`feeding disorders in cattle, sheep and goats has been spurred
`by a desire to maximize yields of meat and dairy products.
`Existing drug-based treatments (see, e.g., US. Pat. No.
`4.?61,426 issued to Martin, et al., and U.S. Pat. No. 4,6105,
`609 issued to Potter), however, have the serious drawback of
`rendering products from treated animals unsalahle for long
`periods under laws designed to protect consumers from
`harmful drug residues. Farmers, unhappy with the need to
`choose between low yields or unsalable products, have long
`sought the development of alternative, drug-free dietary
`treatments. As farmers expand into other livestock markets,
`such as the raising of ostriches and cmus for yields of meat
`and eggs, similar problems occur. The goals of drug-free
`dietary treatments are generally, improved growth and per—
`forrnanee, and especially, appetite stimulation and reestab-
`lishment of the rumen or gastrointestinal microbial popula-
`tions necessary for proper digestion.
`Much attention has been given in recent years to the use
`of certain microorganisms as dietary adjuncts in efi‘orts to
`improve the growth and performance of livestock in general.
`and rcestablishment of rumen or gastrointestinal microbial
`populations in ruminant animals. Such dietary cultures are
`known as probiotics or direct-fed microbials. (Gilliland, S.
`13., 8th lnr’l. Biatech. Syn. Proc, Vol. 2, pp. 923—933
`(1988)). Generally, the microorganisms of such probiotics
`are those that are expected to grow andlor function in the
`intestinal tract or in the rumen of the particular animal and
`can exert certain metabolic actions that influence that ani-
`mal. Various gastrointestinal tract microorganisms which
`have been considered for this type of usage include lacto-
`bacillus oct'dophilus, [acrobacillus plantanon, [actobecil-
`t'us femrenrnm, Lactobocillur caret, Lactobacillns lactis,
`Pediococcos cerevisioe and Streptococcus fizecr'um. These
`bacteria perform one or more of the following functions:
`They compete for villi attachment sites with pathogenic
`microorganisms, control Ph{acidi1y) within the gastrointes-
`tinal tract, produce enzymes and other metabolites which
`benefit digestion, and produce substances capable of inhib—
`iting die growth of other harmful microorganisms. Addi-
`tional microorganisms that could be used for this purpose
`include the live cell yeast, Saccharomyces cerevirioe, the
`fungus, Aspergillus otyzae, and bacteria, Bhidobacrerium
`longum and Pmpionibacten'wn fieudenreichii or the taxo-
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`Page 10 of 28
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`admin equivalents thereof. Live cell yeast cultures and
`fiingi, such as Sacchammyces cerevisiae and Aspergt'llos
`oryzae, have been shown to produce beneficial enzymes and
`raise the pH in the rumen by enhancing the growth of
`bacteria that either utilize lactic acid or ferment fecdstuffs to
`absorbable fatty acids. Bifidobacteria species have been
`shown to be a major colonizer of the undeveloped rumen of
`newborn cattle and the lower intestine of all newborn
`animals. Brfidobacten‘wn longwn may also aid in intestinal!
`rumen pH control and enzyme production. Certain strains of
`Propionibacreriomfi-eudenreichii utilize nm'ien nitrates as a
`food substrate and are,
`therefore, beneficial
`in reducing
`nitrate toxicity in animals exposed to excessive amounts of
`that chemical. This might occur where the cattle are exposed
`to excess nitrates in feeds, to fresh pastures grown during
`drought conditions and to nitrates in water.
`As known in the art, the Food and Drug Administration,
`Center for Veterinary Medicine has published a list of
`microorganisms which they have reviewed and have found
`to present no safety concerns when used in direct-fed
`microbial products in the Official Publication of the Asso-
`ciation of American Food Control Dfiicials, Inc. (AAFCO)
`(1993} at pp. 148—149. the disclosure of which is hereby
`incorporated by reference. Many commercial direct~fed
`microbials are especially important in their non-spore forms,
`i.e. vegetative forms. These vegetative forms may be in the
`dormant state. These microbials are approved for animal
`feeding and an: available to the public from several suppli~
`ers
`
`Common feed additives (AAFCO p. 248—253, 21 CFR
`§573 and 21 CFR §534) are known in the art as carriers for
`animal or livestock feeding purposes. Those which have
`been found to be safe when used in feed, are on the FDA’s
`Generally Recognized as Safe (GRAS) lists. 21 CFR 584.
`The ingredient additives not defined by AAFCO appear on
`the “Least Common Federal Ingredients” list, (AAFCO p.
`248-253).
`'Ib derive maximum benefit from use of probiotics, the
`microorganisms must survive and grow in the rumen andl'or
`intestine. It is thus imperative that the probiotic contain
`viable and active microorganisms at the time of consump-
`tion. The microorganisms used as probiotics, therefore, must
`be stable during preparation and during storage prior to
`consumption.
`The simplest approach to delivery of probiotics is to add
`cultures to animal feed. However, it appears that few direct—
`fed microbials are stable in feed for more than 3—5 days.
`(Aimutis, W. R., Feeds Management, Vol. 42, pp. 26—32
`(1991)). Moreover, some feed contains antibiotics which are
`contrary to microbials stability. Yet other feed is pelleted,
`and most Lactobacillus species, which are predominant and
`beneficial
`intestinal species. are susceptible to the high
`temperatures, compression, aeration and mixing abrasion to
`which they are exposed during the pellcting process.
`Another approach is to provide the bacteria themselves as
`a pellet or bolus. Many such bolus products are commer-
`cially available.
`More recently, bolus or pellet formulations have been
`developed which include a combination of the microorgan-
`isms and dry vitamin and trace mineral supplements. as
`nearly simultaneous administration in vivo of these compo-
`nents has been suggested as being highly beneficial to
`achieving the goals of appetite stimulation and microbial
`population reestablishrnent. Many of these bolus formula-
`tions are available commercially. It has been found, how-
`ever, that the supplements and microorganisms are incom-
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`5,501,85'lr
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`3
`patiblc as the vitamin and mineral levels commonly used
`and elficacious for livestock are toxic to the microorganisms
`in many formulations. The toxicity is dependent upon con-
`centration of the vitamins andlor minerals and the microor-
`ganisms used. By “toxic“ is meant that the vitamins andror
`minerals inhibit, or prevent growth or diminish viability of
`the microorganisms. By viability is meant the capability of
`life. The toxicity is demonstrated in a reduction of shelf life
`of a microorganism. As the time period in which the
`microbials are in contact with the toxic substance increases,
`the concentration of the mierobials (often expressed as
`colony forming units per gram i.e., CFUlg or colony forming
`units per bolus i.e., CFUfBolus) as measured by standard
`testing procedures, decreases. This is also expressed as
`Percent Survival Rate of the microorganism observed over
`a period of tune. As the time period increases, the Percent
`Survival Rate decreases.
`
`Thus, the CFU’s decrease as time increases. The popula-
`tion of viable microorganisms can be greatly reduced within
`a week or within approximately a month. As indicated
`previously, microorganisms are also sensitive to mixing
`abrasion, aeration, compression and high temperatures, all
`of which occur during conventional hard bolus production.
`Moreover, the bolus formulations also require binding, wet-
`ting and disintegrating agents, any or all of which may
`adversely afiect the viability of the microorganisms. Such
`bolus products, therefore, have limited shelf stability or shelf
`life, in that, the population of viable microorganisms can be
`greatly reduced within about a week or about a month.
`Thus, a persistent and vexatious problem, largely unat-
`tended by the prior art, is the lack of a method for simul—
`taneously delivering incompatible substances in vivo to
`animals, when one of the substances is a viable microor-
`ganism culture.
`Various prior art methods of physical separation, e.g.,
`encoating, encapsulation and microencapsulation, of nutri-
`tional supplements are known. however, none adequame
`address the preparation and storage requirements of sensi-
`tive direct-fed microbial agents. For example, conventional
`microencapsulation subjects microorganisms to a number of
`potentially fatal packaging procedures arid requires expen-
`sive materials, complex equipment, and carefully controlled
`environmental conditions. Polymeric microeapsules also
`require specific pH ranges or enzyme activities to elfeet
`release of their contents in vivo. These requirements often
`frustrate conventional laboratory assessment techniques and
`prevent efiective nutrient release in animals whose rumen
`andt'or gastrointestinal pH of enzyme balances have been
`disrupted by microbial depopuiation.
`U.S. Pat. No. 4,695,466 to Mori shita discloses a multiple-
`encapsulation method. The Morishita process includes suc-
`cessively encapsulating oil solutions or suspensions in soft
`capsules. Although the method of Morishita has potential for
`delivery of two components in a single vehicle, the use of oil
`carriers presents insurmountable obstacles to the delivery of
`microorganisms and vitamin supplement components. It is
`unlikely that Morishita‘s soft outer capsules will be able to
`withstand common shipping, storage and administration
`conditions and also is unlikely applicable to commonly
`available microbial forms.
`
`Despite recognition of the known drawbacks of prior art
`products, the art has not adequately responded to date with
`a method for delivery in vivo of the incompatible compo—
`nents, namely, direct-fed microbials and nutrient supple~
`ments nearly simultaneously to cattle, sheep, goats, and
`ratites.
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`4
`SUMMARY or THE MENTION
`
`The present invention responds specifically to the long-
`felt need heretofore unmet by the prior art, and especially
`with a view to overcoming the inherent inadequacies of
`combination supplements and direct—fed microbials for oral
`delivery to animals. The composition is a dietary adjunct or
`feedstull, providing the convenience and reliability of oral
`administration, while providing near simultaneous delivery
`in vivo of direct—fed microbials and incompatible sub
`stances. such as vitamins and minerals in a partitioned
`unitary structure, a double capsule. The composition is shelf
`stable, i.e., allows substantially greater viability of micro-
`bials. and does not require binding. wetting and disintegrat-
`ing agents necessary for pellet or bolus formulations. The
`composition provides accurate unit dosage, virtually simul—
`taneous in vivo delivery of the components of the double
`capsule, and rapid in vivo dissolution.
`This invention is directed to the probiotie classes of
`publicly available direct-fed microbials which are fed to
`both ruminant animals and to non-ruminant animals. Where
`the direct~fed-microbial is ruminant specific, it is understood
`that the application is to feeding ruminant animals. However
`the scope of this invention also includes non-mminant
`animals such as other livestock. Other livestock includes,
`but is not limited to, ratites, such as ostriches and emus.
`The foregoing, and other advantages of the present inven-
`tion, are realized in one aspect thereof in an oral nutritional
`composition, i.e., a dietary adjunct, useful for treating feed-
`ing disorders and improving feed efficiency in livestock,
`e.g., cattle, sheep and goats, especially ruminants, but not
`limited to ruminants. Non-ruminants include, but are not
`limited to ratites [such as, but not limited to ostriches and
`emus). For ruminant animals,
`the gastrointestinal
`tract
`includes the rumen; for non-ruminant animals,
`the gas-
`trointestinal tract is absent a ruruen. Gastrointestinal micro-
`orgainsms inhabit the gastrointestinal tract. Gastrointestinal
`microorganisms include bacteria, live cell yeasts or fungi,
`among others.
`The dietary adjunct composition comprises a double
`capsule which includes live cultures of rumen andlor gas-
`trointestinal microorganisms in a first capsule which is
`enclosed with vitamin andl'or mineral supplements in a
`second capsule. The capsules are preferably made of a
`dissolvahle material, preferably gelatin. The microorgan-
`isms and supplements may be combined with acceptable
`feed grade
`carriers. Gastrointestinal microorganisms,
`include bacteria, live cell yeasts, fungi or a combination
`thereof. By live cell yeast is meant, a yeast culture contain-
`ing live yeast cells but not containing yeast metabolites
`andlor the yeast growth media.
`In another embodiment of this invention, this invention
`provides a dietary adjunct composition comprising a double
`capsule which includes a vitamin andfor mineral supplement
`in a first capsule which is enclosed with live gastrointestinal
`microorganisms in a second capsule. The first capSule wall
`separates the live microorganisms from the vitamins auditor
`minerals. The capsules are preferably made of gelatin. The
`microorganisms and supplements may be combined with
`acceptable feed grade carriers. Gastrointestinal nficroorgan-
`isms, include bacteria, live cell yeasts, fungi or a combina—
`tion thereof. By live cell yeast is meant, a yeast culture
`containing live yeast cells but not containing yeast metabo-
`lites andr'or the yeast growth media.
`In still another aspect, this invention comprises a double
`capsule which includes a gastrointestinal nucroorganism
`separated by a capsule wall from a feed ingredient which
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`5,501,857
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`5
`when stored in contact with the microorganism for a period
`of time will cause the microorganism to diminish its full
`growth potential.
`In still another embodiment, this invention provides a
`dietary adjunct composition comprising a double capsule
`which includes an inner capsule and an outer capsule. The
`outer capsule is spaced apart from and encloses the inner
`capsule. The inner capsule includes a dissolvable shell and
`a first substance therein. The outer capsule includes a
`dissolvable shell and a second substance therein. The shells
`are preferably made of gelatin. One of the substances is
`viable gastrointestinal microorganisms and the other sub-
`stance is a nutritional supplement. The nutritional supple-
`ment has the property of diminishing the shelf life of the
`microorganisms. That is, direct contact between the micro-
`organisms and the nutritional supplement for an extended
`period of time will cause the number of colony forming units
`expressed per unit quantity, i.e., CFUlg or CFUt'bolus, of the
`gastrointestinal microorganisms to decrease.
`In another aspect, the invention is a method of simulta-
`neously delivering incompatible compounds to animals in
`vivo. Such delivery is achieved by feeding an animal a
`double capsule containing a first substance in a first capsule,
`which is enclosed with a second substance, in a second
`larger capsule. One of the two substances is a viable sub-
`stance. The other substance is such that the viable substance
`is rendered nonviable when both substances are stored and
`administered simultaneously in a bolus or in a single capsule
`formulation. By “incompatible" is meant that one of the nvo
`substances prevents or inhibits the growth audlor effects the
`viability of the other substance. By viable is meant capabil-
`ity of life. The substance can come out of a dormant
`condition, such as a freeze dried state to a active vegetative
`state. The viable substance is not completely in a spore state.
`The viable substance may be in a form that has both spore
`forming and non spore forming components. As is known in
`the art, only certain of the direct-fed niicrobials have a spore
`component. In this instance, CFU include typically spore
`and nouspore forms.
`In another embodiment, this invention provides a method
`for preparing shelf-stable compositions of incompatible sub-
`stances, which includes the use of multiple capsules of
`variable composition. Such method is accomplished manu-
`ally or by machine.
`Other advantages and a fuller appreciation of the specific
`adaptations, compositional variations, and physical and
`chemical attributes of the present invention will be gained
`upon an examination of the following detailed description of
`the invention, taken in conjunction with the accompanying
`drawings and appended claims.
`
`BRIEF DESCRIPTION OF THE DRAWINGS
`
`The present invention will hereinafter he described in
`conjunction with the appended drawings, wherein like des—
`ignations refer to like elements tluonghout and in which:
`FIG. 1 shows an enlarged sectional view of the capsule-
`in-a-capsule structure in accordance with the present inven-
`tion with the microorganisms within the inner capsule;
`FIGS. 2a—2f illustrate a method by which each capsule-
`in-a-capsule structure of FIG. 1 is assembled;
`FIG. 3 compares the efficacy of double capsules in
`accordance with the present invention and hard boluses in
`simultaneously delivering live microorganisms and incom-
`patible nutrient supplements;
`
`6
`
`FIG. 4 shows an enlarged sectional view of the capsule-
`in-a—capsule structure in accordance with the present inven-
`tion, with the microorganisms outside of the inner capsule
`and with the vitamins andfor minerals admixture in the inner
`capsule; and
`FIGS. 50-53“ illustrates a method by which each capsule-
`in«a~capsule structure of FIG. 4 is assembled.
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`DETAILED DESCRIPTION
`
`invention relates broadly to nutritional
`The present
`supplements and dietary adjuncts for animals, such as cattle,
`sheep, goats, and ratites and specifically to compositions and
`nutrient delivery systems which permit delivery of direct-fed
`microbials with incompatible substances. However,
`the
`composition of the present invention is most particularly
`adapted for use in oral supplementation formulations which
`combine nutrients, such as vitamins andr'or minerals, and
`viable gastrointestinal microorganisms,
`such as
`fungi,
`yeasts, or bacteria. Gastrointestinal microorganisms include
`intestinal microorganisms, andfor rumen microorganisms
`depending upon the animal. The gastrointestinal microor-
`ganisms include but are not limited to those dwelling in the
`stomach, large and small intestines. Accordingly, the present
`invention will now be described in detail with respect to
`such fields of endeavor; however, those skilled in the art will
`appreciate that such description of the invention is meant to
`be exemplary only and should not be viewed as limitative of
`the full scope thereof.
`The present invention provides a nutritional composition
`useful for ameliorating drug-induced, stress-related, and
`other feeding disorders in food—producing animals. The
`composition is partiCularly useful as arumlnant feedstufi for
`improving feed efficiency and promoting growth for cattle,
`sheep and goals, but may also be used in non-ruminant food
`producing animals such as ratitcs. Ratitae is a classification
`of birds and includes birds, such as ostriches, rheas, cas~
`sowaries. emus, elipbantbirds, moas, kiwis. Raising cents
`and ostricbes is popular for production of meat, hides,
`feathers and eggs. For ruminants, the composition avoids the
`milk and slaughter withdrawal periods required after drug
`treatments and enhances the general nutritional status of the
`animal. Additionally, the composition is shelf-stable and
`provides a general packaging system for incompatible mate-
`rials, and is particularly useful for direct-fed nucrohial
`agents or probiotics. These attributes are achieved through a
`particular composition meeting a special combination of
`physical parameters.
`As used herein, the term “incompatible" is meant to refer
`to substances which deleteriously react with one another
`when combined in desired levels or concentrations. The
`microbial is a substance which has a capacity of viability or
`is viable. The incompatible substance renders the microbial
`non viable when the probiotic and the incompatible sub-
`stance are stored and administered simultaneously in abolus
`or in a single capsule formulation.
`In one embodiment, the invention provides a nutritional
`composition whose components are incompatible, and
`which incompatible components are physically separated
`from each other until they reach their in vivo situs. The
`composition includes two components. One of the compo-
`nents is cultures of viable microorganisms, e.g., bacteria or
`fungi or live cell yeasts, or combinations thereof. By live cell
`yeasts is meant. a yeast culture containing live yeast cells but
`not containing yeast metabolites andlor the yeast growth
`media. The other of the components is nutritional supple-
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`5,501,857
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`meats, e.g.. vitamins andror minerals. Each component
`necessary to make and use the present invention is commer—
`cially available or can be synthesized using known meth-
`odologies. The two components are separated from one
`another. One of the components is enclosed in the inner
`capsule. The outer capsule contains the inner capsule and the
`other component. In a preferred embodiment, the microbial
`cultures are enclosed in a first capsule which is then enclosed
`with the vitamin andlor mineral supplements in a second
`such capsule, i.e., a “capsule-in—a—capsule” structure. How-
`ever the invention also comprehends the reversal of the
`components, i.e., the vitamins andfor mineral supplements
`are enclosed in a first capsule which is then enclosed with
`the microorganisms in a second capsule. The microorgan-
`isms of the one component serve the valuable function of
`repopulating the rumen andlor intestines,
`thus enabling
`digestion to resume, producing digestive enzymes, and
`correcting acid imbalances which result from rumen or
`intestinal microbial microorganism depopulation.
`The vitamins andfor minerals of the other component
`increase the nutritional status of animals laboring under
`conditions of malnutrition caused by feeding disorders.
`Further, once dispersed throughout the rumen andfor intes-
`tines,
`these vitamins andlor minerals support
`the rapid
`growth of the microorganisms of the first component. Oral
`administration of these vitamins andlor minerals contempo-
`raneously with the administration of microorganisms is
`preferable to separate administration. Separate administra-
`tion increases the risk that microorganisms will not encoun-
`ter dispersed vitamins and minerals in the rumen andlor in
`the intestine and thus fail
`to exhibit
`their full growth
`potential. Also separate adminisuation increases the end
`user cost for materials and labor.
`
`In one embodiment. it has been found that the bacterial
`survival rate with the capsule-in—a—capsule structure of the
`present invention after up to six months storage, i.e., a
`shelf—life of six months after preparation, is nearly 500 times
`that of an admixture of the bacteria and nutrient supple-
`ments. Art admixture of bacterial and nutrient supplements
`is typically a single capsule or a bolus formulation.
`FIG.
`1 illustrates a capsule-in-a-capsnle structure in
`accordance with the present invention and is generally
`designated as 20. Capsule—in—a—capsule 20 includes an inner
`capsule 22 and an outer capsule 24. The inner capsule 22 is
`within the outer capsule 24. Inner capsule 22 contains viable
`microorganisms 26 and outer capsule 24 contains vitamins
`andl'or minerals generally designated as 28. Inner capsule 22
`includes a top member 30 and a bottom member 32 which
`is bigger than top member 30. Top member 30 and bottom
`member 32 are locked together after filling by a locking
`mechanism 33 which includes a groove 34 proximate the top
`35 of bottom portion 32 and a complementary ridge 36
`substantially about the mid-portion 3'? of top portion 3|),
`forming a circumferentially nested ridge and groove. Simi—
`larly, outer capsule 24 has a top member 38 and a bottom
`member 40 in which top member 38 is locked to bottom
`member 40 with a groove 42 and a ridge 44. Alternatively
`the content of the inner and outer capsules may be reversed
`as is shown in FIG. 4 and wherein the capsule—in—a~capsule
`vehicle 20' is depicted with microorganisms 26' contained
`Within the outer capsule and with the vitamin andlor mineral
`admixture 28' contained in the inner capsule. FIG. 4 illus—
`trates a capsule-in-a-capsule structure in accordance with the
`present invention and is generally designated as 20'. Cap-
`sule-in-a—capsule 20' includes an inner capsule 22' and an
`outer capsule 24'. The inner capsule 22' is within the outer
`capsule 24'. Inner capsule 22' contains vitamins andfor
`
`10
`
`15
`
`30
`
`35
`
`40
`
`45
`
`50
`
`55
`
`65
`
`minerals generally designated as 28' and outer capsule 24‘
`contains viable microorganisms 26'.
`Inner capsule 22'
`includes a top member 30' and a bottom member 32' which
`is bigger than top member 30'. Top member 30' and bottom
`member 32‘ are locked together after filling by a locking
`mechanism 33' which includes a groove 34' proximate the
`top 35' of bottom portion 32' and a complementary ridge 36‘
`substantially about the mid~portion 37' of top portion 30'.
`forming a circumferentially nested ridge and groove. Simi-
`larly, outer capsule 24‘ has a top member 38‘ and a bottom
`member 40' in which top member 38' is locked to bottom
`member 40' with a groove 42' and a ridge 44'.
`The capsules, in either FIG. 1 or FIG. 4. are preferably
`made of gelatin, however other materials which dissolve in
`the animal‘s gastrointestinal system, as are known in the art,
`may also be used and are encompassed by the invention.
`Capsule shells are, however. easily reformulated to meet a
`myriad of size, transportation, storage, and administration
`requirements, e.g., excessive heat or cold, vibration, humid-
`ity, compression or impact, aeration, or ultraviolet light. The
`shell of the inner capsule forms a wall, separating the
`contents contained within the inner capsule from the sub-
`stance extemal to the inner capsule, but contained within the
`outer capsule.
`In snucntre 20, the microorganisms 26 of the first com-
`ponent, i.e., of inner capsule 22, include one or more of the
`indigenous gastrointestinal bacteria selected from [acroba-
`cilltts acidophilas, Lacrobact'llus lacrir, Lacrobnct’llns casei,
`Streptococcus fascittm, and Pediococcus cerevirt'ae andt'or
`microorganisms including the live cell yeast Saccharomyces
`cerevisiae. fungus Aspergillus oryzae, andr'or bacteria Bifi-
`dobacterium and Propionibacterium. All of these microbes
`are publicly available from commercial sources such as
`Far-Mot Biochem, Milwaukee, Wis. Chr. Hansen's Labora-
`tory, Inc, Milwaukee, Wis. Red Star® Yeast Products.
`Milwaukee, Wis. and Agtech Products, Inc., Waukesha, Wis.
`By live cell yeast culture is meant, a yeast culture containing
`live yeast cells but not containing yeast metabolites andl‘or
`the yeast growth media. The microorganisms can be pro-
`cessed in accordance with conventional methods of bacte—
`nology to produce direct~fed microbial agents suitable for
`encapsulation in gelatin-shelled capsules and administration
`to cattle, sheep and goats, and ratites. The in vitro viability
`of the microorganisms of the inner capsule is determined by
`counting the colony-forming units per gram (CFUI'g) of the
`culnire administered, according to standard feed industry
`protocols such as those developed by the American Feed
`Ingredients Association (AFIA) or the National Feed Ingre-
`dients Association (NFIA). At time of preparation,
`the
`microbial count in the inner capsule in accordance with the
`present invention depends upon the microorganism used and
`is suitably about 20X109 CFUlg for Iactobnct'llas acidophi—
`Eur, [acrobacillus tom's, [acrobact'lltts carei, Streptococcus
`fascium, and Pediccoccus cerevisiae; about 10x109 CFUl'g
`for live cell yeast Saccharomyces cerevisiae; about 1x107
`CFUIg for Aspen'gillus oryzae; about 20x109 CFUl’g for
`Bifidobacterium; and about 2x1011 CFUIg for Propionibac-
`terium. As is known in the art, the invention comprehends
`the use of the standard formulation overages for determining
`the amount of microbial culture present. The standard over-
`age depends on the culture used and ranges typically from 0
`to 50%. For example for the lactic acid producing bacteria
`an average of 50% is typical, for yeasts 50%, for Bifido-
`bacterium 50% for Propionibacterium 50%, and for Aspen-
`gt'llus oryzae about 0%.
`The microorganisms can be employed in admixtures with
`conventional excipients, e.g., acceptable feed grade carriers
`
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`5,501,857
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`9
`suitable for enteral [e.g. oral) administration which do not
`deleteriously react with the microorganisms. By the term
`“deleteriously react“ it is meant that the feed grade carrier
`does not inhibit, or prevent growth or diminish the viability
`of the microorganisms. Suitable feed grade carriers include,
`but are not limited to, calcium carbonate, nonhygroscopic
`whey. rice hulls, and sucrose.
`The microbial preparations can also be mixed with aux-
`iliary agents, e.g., whole dried milk, dextrose, enzymes,
`plasma proteins or amino acids to promote the growth and
`nutritional status of the animal and the microbials in vivo.
`
`The vitamins and minerals of the second component, i.e.,
`outer capsule. are selected from one or more vitamins,
`namely, A, Big, C, D, E, and K, niacin, thiamine. choline,
`biotin, folic acid, riboflavin, pantothenic acid, andl‘or one or
`more minerals, namely, cobalt, copper, iron, manganese,
`selenium and zinc.
`
`The vitamins andfor minerals of the outer capsule 28 can
`be processed in accordance with conventional methods of
`pharmacy to produce agents suitable for encapsulation in
`gelatin-shelled capsules and administration to cattle, sheep,
`goats, and ratites. For example, the vitamins and minerals
`can be administered in alternative sulfate, oxide, chelated, or
`other chemical forms to promote efficient dissolution and
`absorption in vivo.
`The vitamins andt'or minerals can also be employed in
`admixtures with conventional excipients, e.g., acceptable
`feed grade c