Patent Application
20200268005
Ruminant feed additives may be subject to undesired chemical modifications by rumen organisms. In the context of the modification of polyunsaturated fatty acids by the rumen microorganisms, for example, it can be observed that both the content and the position of double bonds in the carbon chain of the fatty acid may be altered, Usually, the double bonds of the unsaturated fatty acids in the plants are separated by at least two single bonds. Microbial enzymes are capable of generating a conjugated double-bond system in which the double bonds are separated by only one single bond. These include, for example, the conjugated linoleic acid isomers (conjugated linoleic acids, CLA). They are formed mainly by the linoleic acid isomerase enzyme of the rumen bacterium Butyrivibrio fibrisolvens, starting from linoleic acid (C18:2 cis-9, trans-12). The main CLA isomer is cis-9, trans-11-CLA (C18:2 cis-9, trans-11), which amounts to approximately 75-92% of the total CLA content. In contrast, the milk-fat-depressing isomer C18:2 trans-10, cis-12 is present in a small amount of 0.03-1.5% only. Some of these isomers reach the ruminant's organism via absorption. In the case of dairy cows, the rumen-modified fatty acids are also transported into the milk. The CLA content of the milk fat is greatly feed-dependent and amounts to approximately 0.3-1.1% (cf. Kirchgeßner, Tierernährung, 13th edition, DLG Verlag GmbH), in pasture-fed animals even up to over 4% (Troscher et al., 2014; in print).
Owing to their potent physiological or pharmacological activities, CLAs are intensively researched. CLAs show cancer-protective and antiinflammatory activities in cell culture studies and animal experiments. It is also known that conjugated linoleic acid isomers (CLA isomers) inhibit, in ruminants, the expression of a large number of genes which are responsible for the uptake, into the mammary gland, of circulating fatty acids which are responsible for the synthesis of fatty acids and the formation of triglycerides. It is in particular the C18:2 trans-10, cis-12 CLA isomer which has proved to be particularly fat-reducing in this context. Advantages which are discussed in connection with the reduction of the milk fat content are, inter alia, relief of the metabolism and increased milk yields. It has been reported that a reduction of the milk fat content of 0.4-0.5% may result in increased meal yields by up to 3-10% (cf. Kirchgeßner, loc. cit.).
This is why CLAs are currently often added to milk production ration. While these preparations are approved in feeding, they have to be employed in rumen-protected form since unprotected CLAs are further degraded in the rumen into ineffective C18:1 and C18:0 fatty acids.
When these or other molecules which are degraded or chemically modified by rumen micro-organisms so that they lose their biochemical activity are to be administered to ruminants, they will, therefore, have to be protected against such undesired microbial activities in the rumen. The literature describes various methods which are capable of imparting rumen protection to polyunsaturated fatty acids (PUFAs), such as the CLAs. This is also clear from findings, for example by Elgersma et al., in Fresh Herbage for Dairy Cattle, 175-194, 2006, which describe biohydrogenation rates of unsaturated fatty acids in the range of 82-98%. Only approximately 2 to 22% of the polyunsaturated fatty acids (PUFAs) which have been ingested can still be detected after passing through the rumen.
Thus, the literature (cf. Kirchgeßner, loc. cit.) describes various forms of rumen-protected fats which are protected against microbial degradation and microbial modification:
Mentioned are in particular:
a) native protected cell-bound oilseed fats, which are employed in the form of oilseed cake or crushed whole seeds. The principle of the protection here is the slow release of the oils from the plant cells; however, the storage stability of such fats is poor.
b) heat-treated oilseeds, for example by extrusion; here, denatured proteins surround the fat droplet, thus protecting it from microbial degradation.
c) chemically modified fats, for example by suponification of fatty acids with calcium or coating the fat droplets with formaldehyde-treated proteins.
d) Fats which are protected from microbial enzymes by technical methods (for example coating with hardened vegetable fats).
EP-A-1 100 489 discloses a method for reducing the milk fat content in dairy cattle, where an effective amount of CLA is to be administered to the cattle. By way of protection against modification from rumen bacteria, it is proposed to administer the active substance by injection or to provide it in coated form.
FIG. 1 shows the retention profile over time of various in-vitro incubated CLA formulations in pellets and extrudates (T1 and T3 according to the invention).
The object of the invention is to provide a simplified route for converting rumen-unstable feed components into a dosage form with improved rumen stability.
Surprisingly, the object was achieved by providing uncoated, pelleted feeds as per the appended claims.
A “pellet” or a “pelleted form” comprises solid feed formulations as are obtainable in a manner known per se with the aid of conventional pelleting devices, but also conventional extrusion or expanding devices. Usually, such pellets have a length of approximately 0.5 to 2 or 0.8 to 1.5 cm, such as, for example, approximately 1 cm, and a diameter in the range of from 0.1 to 0.8 or 0.4 to 0.5 cm. Also encompassed by these terms are what are known as “crumbled” pellets, i.e. pellets which are converted into smaller particles by the action of mechanical force. The purpose of crumbling is frequently to make it easier for the animal to take up the pellets, or else better distribution in the final feed. Crumbling allows the particle size to be reduced in a targeted manner and the number of particles to be increased, giving, for example, a particle mixture with a content of >50%, for example 55 to 95 or 60 to 90 or 70 to 80% of particles with a diameter in the range of from approximately 3 to 6 or 4 to 5 mm.
A “rumen-unstable constituent” (pure substances, but also mixtures of natural or synthetic substances) comprises a chemical compound which, without sufficient protection, may be subjected to chemical modifications (modification of the chemical empirical formula, such as, for example, by biohydrogenation, and/or changes in configuration and/or stereochemistry) upon passing through the ruminant's rumen. As a rule, this will be an organochemical substance, in particular one which is of importance as a food/feed component or a food/feed additive. Those which must be mentioned in particular are saturated or mono- or polyunsaturated carboxylic acids, such as, for example, those with at least 6 carbon atoms, such as e.g. the CS, PUFA, MUFA or CLA stated below. Further feed additives, or feed components or straight feeds which must/may be administered in rumen-protected form are amino acids (in particular methionin, lysine and the like), enzymes, cholin and other active substances from the class of the vitamins.
A “rumen-unstable mixture” or “rumen-unstable feed mixture” comprises at least one “rumen-unstable” constituent and is not or insufficiently rumen-protected, i.e. the constituent, if fed and passing through the ruminant's rumen, would be exposed to chemical changes as described above.
In the context of the present invention, “rumen stability” or “rumen protection” means that a “rumen-unstable” constituent is converted by the above-described “pellet” form into a state of reduced “rumen instability” up to an essentially complete rumen protection. This improved rumen stability or reduced rumen instability can be determined in a simple manner by comparing the stability of the constituent in a pelleted or nonpelleted mixture using experimental approaches (in vitro or in vivo) which are described in the experimental part.
“Carboxylic acids” (CS) are, in particular, straight-chain or branched, in particular straight-chain, saturated or mono- or polyunsaturated, optionally substituted C6-C30-monocarboxylic acids. Examples of saturated unbranched fatty acids are caproic acid, oenanthic acid, caprylic acid, pelargonic acid, capric acid, undecanoic acid, lauric acid, tridecanoic acid, myristic acid, pentadecanoic acid, palmitic acid, margaric acid, stearic acid, nonadecanoic acid, arachidic acid, behenic acid, lignoceric acid, cerotic acid and melissic acid. Examples of monounsaturated fatty acids are palmitoleic acid, oleic acid and erucic acid. Examples of diunsaturated fatty acids are sorbic acid and linoleic acid. Examples of triunsaturated fatty acids are linolenic acid and elaeosteric acid. Examples of tetra- and polyunsaturated fatty acids are arachidonic acid, clupanodonic acid, eicosapentaenoic acid and docosahexaenoic acid. Examples of substituted fatty acids are ricinoleic acid ((R)-12-hydroxy-(Z)-9-octadecenoic acid). Further suitable fatty acids are naturally occurring fatty acids such as gondo acid and neronic acid. If double bonds are present in the fatty acids, they may be present both in cis and in trans form. The substituents are preferably selected among hydroxyl and lower alkyl groups, such as, for example, methyl and ethyl groups. Keto groups or epoxy groups may furthermore be present in the hydrocarbon radical, as is the case for example in vernolic acid. Further functional groups are cyclopropane, cyclopropene and cyclopentene rings, which may be formed by bridging of two adjacent carbon atoms in the hydrocarbon radical of the fatty acid (cf. malvalic acid and chaulmoogric acid).
“PUFAs” are polyunsaturated fatty acids with at least two conjugated or nonconjugated C═C double bonds in the fatty acid molecule. Examples which may be mentioned are: linolenic acid, eicosapentaenoic acid (EPA) ((5Z,8Z,11Z,14Z,17Z)-eicosa-5,8,11,14,17-pentaenoic acid; or C20:5 (ω-3)) and docosahexaenoic acid (DHA) ((4Z,7Z,10Z,13Z,16Z,19Z)-docosa-4,7,10,13,16,19-hexaenoic acid) or C22:6 (ω-3)).
“MUFAs” are monounsaturated fatty acids which may be present in cis or trans configuration, such as, for example, oleic acid or vaccenic acid.
Conjugated linoleic acids (CLA) are a group of isomers of the C18-monocarboxylic acid “linoleic acid”, which is diunsaturated and whose two double bonds are in positions 9 and 12 and are, therefore, not conjugated. Linoleic acid is also designated by the abbreviation “018:2 cis-9, cis-12”.
“CLA” comprises, in principle, all conjugated, diunsaturated isomers of linoleic acid (C18:2 cis-9,cis-12), where the position of the two double bonds in the carbon chain may be shifted towards the chain end or towards the carboxyl group and where the stereochemistry of the conjugated double bonds may furthermore comprise any variation (cis/cis, trans/trans, cis/trans), where “cis/trans” comprises the two sequences “trans-cis” or “cis-trans”, with the first-mentioned configuration of the two sequences in each case relating to the double bond which is closest to the carboxyl group:
By way of example, reference may be made to the conjugated linoleic acid “C18:2 cis-9, trans-11”, which is shown hereinbelow:
Therefore, the names cis/trans-9,11-linoleic acid, cis/trans-8,10-linoleic acid, cis/trans-11,13-linoleic acid, and cis/trans-10,12-linoleic acid, used herein comprise both the cis-trans and the trans-cis isomers, in other words:
cis/trans-9,11-linoleic acid comprises: C18:2 cis-9, trans-11 and C18:2 trans-9,cis 11
cis/trans-8,10-linoleic acid comprises: C18:2 cis-8, trans-10 and C18:2 trans-8,cis 10
cis/trans-11,13-linoleic acid comprises: C18:2 cis-11, trans-13 and C18:2 trans-11,cis 13
cis/trans-10,12-linoleic acid comprises: C18:2 cis-10, trans-12 and C18:2 trans-10,cis 12
The same applies analogously to linolenic acid, which is triunsaturated (C18:3, cis-9, cis-12, cis-15) and its cis/trans isomers.
The above information on the substances and the uses described herein of these substances primarily relate to the respective pure substance, but also to natural or synthetic substance mixtures which comprise at least one of these substances, for example at least one PUFA or at least one CLA.
Natural substance mixtures are, for example, fish oils or microbial oils, which may be high in PUFAs, and linseed oil, soya oil, sunflower oil, castor oil and the like.
Synthetic substance mixtures are, for example, commercial products which are high in CLA, such as Lutalin® by BASF SE.
Others which must be mentioned are rumen-unstable derivatives of the abovementioned carboxylic acids (CS, PUFA, MUFA, CLA), such as, in particular, substituted derivatives. Examples which must be mentioned are compounds which are mono- or polysubstituted on the hydrocarbon radical of the carboxylic acid, such as, for example, by hydroxyl groups. An example of such a compound which must be mentioned is: 10-hydroxy-cis-12-octadecadienoic acid.
The present invention relates in particular to the following:
14. Feed according to one of the embodiments 1 to 9, prepared by a method according to embodiment 13.
To prepare the pelleted feed compositions, the rumen-unstable constituent(s) to be formulated in accordance with the invention are mixed, optionally together with further added customary animal feed components (such as, for example, milk production ration). The amount of constituents is chosen such that it is for example in the range of from 0.1 to 20, 1 to 15, 2 to 10 or 3 to 5% by weight. Thereafter, the feed is pelleted with the aid of a suitable pelleting press. To this end, the feed mixture is usually conditioned by passing in steam (with hot steam of a temperature of up to 95° C., for example saturated steam), for example for 20 to 1240 seconds. The increase in temperature during the pelleting process may be controlled via the added amount of saturated steam. This steam conditioning may however also be omitted.
Pelleting is usually carried out in commercially available pelleting presses (for example from Bühler GmbH), which may be designed as an annular die press. Here, rollers press the pre-conditioned substance mixture through an annular die (gap width between roller and die for example 1 mm). Depending on the die, pellets of approximately 2 to 12 mm in diameter may thus be prepared. For dairy cattle feed, one will use, for example, dies with a bore diameter of 4 to 5 mm and a slot length of 40 to 50 mm. The highest process temperature prevails when the mixture is pressed through the die. Temperatures in the range of approximately 60 to 100° C. may be reached at this stage.
The hot material which leaves the die outlet is cut off continuously by means of blades, and the pellets thus formed are immediately cooled in a cooler and optionally subjected to a final drying process. The residual moisture content may, after cooling and optionally drying, be in the range of approximately 1 to 15% by weight, based on the total pellet weight.
Alternatively, the pelleting may be replaced by extrusion or expansion (for example by means of feed expanders from Kahl, Germany) (cf., for example, “Feed Manufacturing Technology IV” Ed: McEllhiney, Kansas State Univ.; Pub. by American Feed Industry Assoc, 1994; Arlington, Va. in which may be found: Extrusion Cooking Systems, Bob Hauck et al., p. 131-139; Wilson et al. 1998, Journal of Poultry Science, 77 (Supp1.1): 41). The products of these processes may then be crumbled.
Typical feed components which are useful in the production of pellets according to the invention comprise straight feeds of vegetable or animal origin as per Futtermittelverordnung (FMV; German Feed Regulation), such as, for example, cereal by-products, wheat bran shorts, wheat bran; extracted meals, pomace, dried molassed beet pulp, fish meal, meat-and-bone meals; and/or straight mineral feeds as per FMV, such as, for example, carbonates, phosphates, sulfates, propionates. Others which are suitable are cereals such as wheat, rye, barley, oats, corn, millet or triticale; cereal by-products (by-products of milling) such as brans, feeds, wheat feeds, bran shorts or middlings; by-products of oil production (extracted meals, expellers, cake); by-products of sugar production (molasses, dry beet chips, feeding sugar, pulp, potato starch, maize gluten, wheat gluten); by-products of the fermentation industry, brewer's grains, yeast, malt culms, stillage; and feedstuffs of animal and other origin, such as blood meal, fish meal, pressed must, potato protein.
Those which must be mentioned in particular are wheat, grain corn, barley, oats; soybeans; cereal meals such as wheat or corn meal, soybean meal, molassed beet chips, wheat feed, wheat middlings, corn gluten feed, extracted soybean meal, extracted rapeseed meal, brewer's grains, cereal stillage, beet molasses, oat bran; sugars such as glucose and sugar alcohols, furthermore proteinaceous components such as soybean concentrate, fish meal, glutens such as corn or wheat gluten, oils and fats, and nutraceuticals such as, for example, free amino acids, their salts, vitamins (such as, for example, A, D, E) and trace elements (such as, for example, Cu as CuSO4), mineral constituents such as calcium carbonate, sodium chloride; phosphates and optionally processing aids, for example glidants, inert fillers and the like, and optionally preservatives.
Typical milk production ration compositions comprise, for example, corn, wheat, barley, oats, rye, citrus pomace, extracted soybean meal, extracted rapeseed meal, soybean husks, palm kernel expellers, DDGS, corn gluten feed, sugarbeet chips, wheat feeds, wheat bran, linseed, molasses, lime, salt, vitamin and trace element premix.
The invention will now be illustrated in greater detail with reference to the following use examples.
Pelleting press: Manufacturer Simon Heessen, Type V3-30C.
Capacity 600-650 kg/h
(1) Lutalin®: (CLA-containing commercial product)
1 GC area %
Lutrell is composed of 34% Lutalin and the additives 15% silica, 5% gypsum and 46% hydrogenated soya oil, with the rumen-stabilizing function being effected mainly by the latter.
The composition (fatty acid profile) of Lutrell is shown in the following table
(3) Lutalin plus Silafett
Fatty acid composition of Silafett=hydrogenated soya oil
0-1%
0-3%
Four experimental variants (see table hereinbelow) are tested.
Variant 1 (T1) is the control (C) and does not comprise any CLA,
Variant 2 (T2) comprises the CLA formulation “Lutrell”,
Variant 3 (T3) comprises the CLA formulation “Lutalin” and
Variant 4 (T4) comprises the CLA formulation “Lutalin plus Silafett”.
1)Data in grams
The CLA preparations are mixed into a milk production ration (25% by weight extracted soybean meal; 75% by weight corn meal) and the mixture is pelleted.
The experimental design corresponds to a 4×4 Latin square (see the following table). In the test groups, the pure CLA is metered in identical amounts of in each case 16.7 g per cow per day.
Each period is 21 days; i.e. an adaptation time of 14 days is followed by a measuring period of 7 days. All four test rations are based on the same basic components.
Each treatment is tested according to the above experimental design on 20 cows. A further 4 cows are assigned to the test groups by way of reserve animals. At the beginning of the experiment, the test cows are in the middle lactation stage. The mean milk yield at the beginning of the experiment is approx. 32 kg FCM per day. The test cows (German Holstein) are selected from the experimental station's herd (herd performance approx. 11 000 kg milk, 3.9% fat, 3.4% protein) and assigned to the four groups by the criteria
All cows are provided with the same total mixed ration (TMR), which is fed once per day ad libitum. The TMR is composed of corn silage, grass silage, hay and approx. 45% concentrate. The TMR target values are shown in the table which follows. This TMR ensures that the cows are provided with sufficient nutrients for a milk yield of 33 kg if they take in approx. 21 kg DM of this feed.
The three CLA formulations are first used for preparing pelleted premixes with a milk production feed. Suitable amounts of these premixes (corresponding to 1 kg per animal per day) are then admixed by hand to the TMR in the feed troughs with the aim of obtaining an amount of in each case 16.7 g of the pure substances in 20 kg DM of the TMR. The control group receives the same concentrate in the same amount, without CLA.
For determining the dry matter, one daily TMR sample is taken, and the samples are combined to give a cumulative sample per measuring period (7 days). This cumulative sample is used for determining the crude nutrient contents as per Weender analysis and the energy content as per the Hohenheimer feed value test.
The TMR uptake, the milk yield and the live weight are recorded daily (individually) for each cow. During the 7-day measuring period, milk samples are taken three times in the form of aliquots from the evening and the morning milk and tested for the fat, total protein (Nx6.38), lactose and urea contents and the somatic cell count by the Baden-Württemberg milk testing organization. An additional milk test is performed in the middle of the first and the second week of each experimental period (preparation phase). A cumulative milk sample which comprises the 7-day measuring period is additionally formed for each cow and each treatment and then frozen so as to be able to optionally perform a fatty acid analysis of the milk fat therein.
Surprisingly, treatments T3 and T4 resulted in the same milk fat depression as treatment T2, although the active substance CLA in T3 and T4 had been admixed in a non-rumen-protected form into the milk production ration. Accordingly, pelleting protected the active substance CLA of T3 and T4 in the rumen to the same extent as this is known from Lutrell, and has also been demonstrated by T2.
Stability in the rumen is the essential prerequisite for the milk fat depression by CLA-containing feeds. To this end, the feeds tested in example 1 (75% corn plus 25% extracted soybean meal) were mixed with 1.66% Lutalin or 5% Lutrell, and the mixture was pelleted or extruded. The pellets and extrudates produced were then incubated in vitro for 4, 12 and 24 hours. After the predetermined times, the incubation was stopped, and all of the contents were transferred into a vessel and freeze-dried. The CLA content was subsequently determined on this freeze-dried sample material.
Preparation of the meal: Corn and extracted soybean meal were weighed in together, ground and mixed in a horizontal mixer. Lutalin or Lutrell were added and mixed in. The mixing time amounted to 15 minutes. These two mixtures were divided in each case. 80 kg were pelleted and 30 kg were extruded.
Pelleting press: Manufacturer Simon Heesen, horizontal die, 3.3×35 mm, nominal capacity 300 kg/hr.
Extruder: Werner & Pfleiderer 37, die 2×0 3.9 mm, nominal capacity up to 50 kg/hr.
Pelleting: Pelleting temperatures of 67-68.5° C. were reached.
Extrusion: An extrusion temperature of 85° C. was reached, approximately 14% of water having been added. A residual moisture of 10 to 12% has been found in the end product.
4 test variants (see table hereinbelow) were tested.
Variant 1 (T1) One pellet and contains the CLA formulation “Lutalin”
Variant 2 (T2) One pellet and contains the CLA formulation “Lutrell”
Variant 3 (T3) One extrudate and contains the CLA formulation “Lutalin”
Variant 4 (T4) One extrudate and contains the CLA formulation “Lutrell”
An analysis performed immediately after the processing step revealed that the active substance CLA survived the pelleting process very well, but that extrusion resulted in an activity loss of approx. 40%.
The Hohenheimer feed value test (HFT) is an in-vitro method for estimating the energetic feed value of ruminant feed. An adapted variant makes it possible also to test active substances in the HFT in respect of their rumen stability, when incubation is stopped after defined periods of time and the remainder of the active substance in the incubation solution is analyzed.
The Hohenheimer feed value test (HFT) is carried out as per the prescribed VDLUFA method (Methodenbuch [Methods book] vol. III, chapter 25.1).
The incubation period depends on the purpose of the test in question; for the degradation of CLA formulations, it is expediently between 4 and 24 hours. The in-vitro system is stable for up to 48 hours; beyond this point in time, however, deviations from a physiological course of the fermentation must be expected. In each case 500 mg of the test variants T1 to T4 (corresponding to approx. 5 mg CLA, or approx. 3 mg for the extruded variants) were weighed in. The incubation period amounted to 4, 12 and 24 h. A triple determination (3 incubation vessels per test variant T1 to T4) was carried out for each incubation period.
After the relevant times, the incubation was stopped by cooling in ice-water. Each of the rumen fluid/buffer mixtures was quantitatively transferred into glass vessels, which are suitable for the subsequent freeze-drying step. The drying vessels are weighed empty, so that, after drying, it was possible to determine the residual weight and use the data for a subsequent quantitative analysis. The freeze-drying system used was a Gamma 1-20 from Christ, 37507 Osterode.
Owing to the specific matrix and the low CLA content, the processing as described in the analytical method (see Chapter 7.4.2) had to be modified as described hereinbelow. The modifications performed are shown in italics.
The sample is weighed or introduced into a 250-ml Erlenmeyer flask as described in 7.4.2; thereafter, the stated amounts of BHT, sodium ascorbate and water are added.
Then, a spatula-tip full of the enzyme Pronase is added, and the Erlenmeyer flask is placed for 15 minutes into an ultrasonic bath at a temperature of 60° C.
After adding the amounts of ethanol and acetic acid which are described in 7.4.2, the sample is returned for 15 minutes into the ultrasonic bath at a temperature of 60° C.
After cooling to room temperature, extraction solution cyclohexane/ethyl acetate 80:20 (v/v) is added. However, only 50 ml of the extraction solution are used, instead of normally 100 ml.
Thereafter, the sample is stirred for 30 minutes on the magnetic stirrer as described in 7.4.2, 70 ml saturated sodium chloride solution are added, and stirring is continued for 10 minutes. Thereafter, the stirrer is switched off, and the mixtures are left to stand until the phases separate.
3 ml of the organic phase are withdrawn, the solvents cyclohexane and ethyl acetate are removed in the stream of nitrogen, and the residue is taken up in 2 ml of the extraction solution cyclohexane/ethyl acetate 80:20 (v/v) (concentration).
An aliquot of the resulting sample solution is directly filtered, by a disposable 0.45-μm-filter, into an HPLC vial.
The injection volume is 20 μl, instead of normally 10 μl.
In this in-vitro experiment, it was possible to reproduce the identical milk fat depression of pelleted Lutalin versus pelleted Lutrell, which had already been found, surprisingly, in example 1, as the CLA retentions measured in the two pelleted variants T1 and T2 were identical. Therefore, using this test, it is possible to assess with certainty at least those variants which are based on the same feed and which merely differ in a physical treatment. Surprisingly, the extruded variants T3 and T4 showed in each case substantially higher retention rates—after 4 and 12 h, and after 12 and 24 h, respectively—than the pelleted variants (cf. FIG. 1).
It can therefore be concluded that Lutalin (non-rumen-stabilized CLA) in extruded form, too, has an at least equivalent rumen stability as a comparable Lutrell extrudate (CLA which has been rumen-stabilized by stabilizing additives).
Reference is expressly made to the disclosure of the publications mentioned herein.
| Number | Date | Country | Kind |
|---|---|---|---|
| 14159496.0 | Mar 2014 | EP | regional |
This application is a divisional application of U.S. application Ser. No. 15/124,403 filed Sep. 8, 2016, which is incorporated by reference. U.S. application Ser. No. 15/124,403 is a national stage application (under 35 U.S.C. § 371) of PCT/EP2015/055006, filed Mar. 11, 2015, which claims benefit of European Application No. 14159496.0, filed Mar. 13, 2014, which is incorporated herein by reference in its entirety. The invention relates to pelleted ruminant feed, comprising, in pelleted form, a mixture of at least one solid particulate feed component with at least one rumen-labile constituent which has been added to the mixture; to processes for their preparation and to their use in methods of altering the milk fat concentration.
| Number | Date | Country | |
|---|---|---|---|
| Parent | 15124403 | Sep 2016 | US |
| Child | 16804086 | US |
