WO2011076155A1

WO2011076155A1 - Feed supplement based on milk thistle, method of its production and its use

Info

Publication number: WO2011076155A1
Application number: PCT/CZ2009/000165
Authority: WO
Inventors: Zdenek Svehllk; Martin Buchta
Original assignee: Irel, Spol. S R.O.
Priority date: 2009-12-22
Filing date: 2009-12-22
Publication date: 2011-06-30

Abstract

This is a feed supplement suitable, in particular, for feeding animals, wherein it contains the whole and/or treated fruits of Milk Thistle (Silybum marianum (L.) GAERTNER) and 10 % to 35 % by weight of crushed and/or milled and/or pressed parts of at least one additional plant species, optionally chosen from the group comprising the genus Dragonhead, Leuzea, Amaranth and Clintonia, relatively to weight of the fruits of Milk Thistle. The method of producing this feed supplement and its use are also disclosed.

Description

FEED SUPPLEMENT BASED ON MILK THISTLE, METHOD OF ITS

PRODUCTION AND ITS USE

Technical Field

The invention relates to the feed supplement containing whole fruits and/or treated fruits of Milk Thistle (Silybum marianum (L.) GAERTNER) in a mixture with other plant materials, to which chemically treated oil from the fruits of Milk Thistle can be preferably added, for use in feeding animals.

Background Art

The fruits of Milk Thistle contain several substances of which the pharmacologically most important are substances of the flavonol-lignan type, in aggregate named flavonolignans or also flavolignans. For their hepatoprotective, antioxidative, galactogenic, and vascular- beneficial effects, these substances are, according to the state of the art, utilized as efficient phytopharmacons in classical (see e.g. disclosed international application WO 2000/078325, European patent EP 1 774 963 Bl, and Russian patents RU 2 291 706 and RU 2 274 400), and in alternative medicine (see e.g. Czech patents CZ 293 046 and CZ 296 773, and published Czech application PV 2004-905), and in cosmetic preparations (see e.g. published Japan application JP 2006282568 and published application DE 102005063063). According to the state of the art, however, use of flavonolignans is not limited to human area. Extracts containing flavonolignans are also fed in the form of feeding mixes or batchs, for instance, to farm animals, in particular to cattle and poultry, with an objective to protect health and improve utility value of these animals and/or increase quality of products obtained from them - as it can be found e.g. in the publication Tedesco D. et al. J. Dairy Sci. 87(7), 2239-2247, 2004; Tedesco D. et al. J. Vet. Med. A Physiol. Pathol. Clin. Med. 51(2), 85-89, 2004; Shiavone A. et al. Journal of Animal Physiology and Animal Nutrition 9_I, 256-262, 2007, and also in French patent FR 2 719 451.

If animals were fed with the fruits of Milk Thistle these were, in particular, parts of the fruits after extraction of active substances (flavonolignans), as mentioned e.g. in Russian patent RU 2 099 076 where cows were fed with extracted cake after its previous extraction with aqueous ethanol. These experiments with feeding the fruits of Milk Thistle after previous extraction of flavonolignans or feeding the separated inner parts of the fruits (endosperm) containing flavonolignans just in very low concentration were also carried out elsewhere in the world with logical stress on the nutritional energy utilization only (Frankiewicz A. et al. Ann. Anim. Sci., 3 (1), 81-87, 2003) in feeding pigs or horses (Stopyra A. et al. Annales Universitas Mariae Curie-Sklodowska Lublin - Polonia, 61(11), 95-102, 2006) or calves (Potkanski A. et al. Rozcniki Akademii Rolniczej w Poznaniu, 209, 85-93, 1991). The fruits of Milk Thistle fed in this way, however, represent predominantly just a source of energy for animals the basis of which is fat (fatty oil), protein component, and starch contained in the fruits. Absolute amount and mutual ratio of these components vary depending on the said methods of processing the Milk Thistle fruits; however, the amount of bioactive flavonolignans received by the organism of an animal in this way into its digestive tract is generally low. Moreover, due to an intact matrix of plant cells and very low solubility of flavonolignans in stomach and intestinal fluids, bioavailability of these substances is quite limited; hence, for reaching their effective concentration in blood, considerable feeding batchs are required of which just a tiny part of flavonolignans is transferred into the internal environment of the organism.

In case of disturbed fruits of Milk Thistle, stability of all nutritional and bioactive components (e.g. flavonolignans) is also considerably limited. The reason lies in propensity of some components to fast oxidation by air oxygen as these components themselves are considered very strong antioxidants (fat containing polyunsaturated fatty acids, flavonolignans, etc.). Action of oxygen on large specific surface of the disturbed fruits, in particular in synergism with sun radiation and/or elevated temperature, results in fast degradation of biologically active substances and, on contrary, in formation of dangerous products of oxidation, such as oxidized fatty acids (peroxides, oxiranes, etc.), i.e. substances causing digestive problems and, possibly also other deeper health damages as a result of their reactivity against living cells.

Disclosure of the Invention

The said shortcomings are solved by the feed supplement based on Milk Thistle according to this invention that provides stable plant mixtures suitable for feeding animals and leading to protection of their organism against oxidative stress, protection of their liver tissue (liver parenchyma), increased utility value of farm animals, and improved physical condition and performance of sports animals. The subject of this invention is the feed supplement particularly suitable for feeding animals, wherein it contains the whole and/or treated Milk Thistle fruits (Silybum marianum (L.) GAERTNER) and 10 % to 35 % by weight of crushed and/or milled and/or pressed parts of at least one additional plant species, relatively to weight of the fruits of Milk Thistle.

Solution according to this invention comprises also the method of producing this feed supplement, wherein this method includes the following steps:

a) crushing and/or milling and/or pre-pressing of the added part of at least one additional plant species;

b) mixing of thus treated plant parts with the whole and/or with treated fruits of Milk Thistle;

which is followed by

c) pressing of the obtained mixture under pressure of 30 to 50 MPa, wherein 5 % to 10 % of fatty oil is separated which consists of more than 85 % by weight of esters of higher fatty acids of chain length C₁₄ - C₂₂ with tribasic alcohol, and a tough pressed mass - the cake is formed; and

d) crushing and sieving of the cake to specified distribution of particle size.

The intact fruits of Milk Thistle are a form of living mass that is able, under certain conditions, to germinate and produce new plant. In this form, all components of the fruit, including present secondary metabolites, are thus naturally protected in maximum way against adverse conditions of the external environment. If, however, the fruits are disturbed by an appropriate method the cell walls can be destroyed, which enables subsequent easier releasing of substances into the external environment, e.g. in the digestive tract of the organism. If this method of mechanical treatment, i.e. pressing, is performed under high pressure (30 to 50 MPa) at low temperature from 25 to 40°C, this can result in disintegration of cell walls accompanied by compaction of the material so that a material of high specific weight is obtained and a pressed cake of low specific surface is formed. In this cake, all its components are enclosed and protected against influence of the surrounding environment similarly to the case of the intact fruits. In the disclosed method of processing, a part of the fat (oil) is separated from the pressed material so that its total content in the final cake is reduced to about one half to one third of the original content (Table 1). In this process, the whole material is uniformly saturated with oil from the disrupted endosperm cells where oil is mainly contained. Oil on the material surface dries out quickly to form a polymeric product that protects the cake from influence of the external environment. In this way, it is possible to store the final materials under prescribed conditions (temperature to 30°C, original closed containers without access of light) without apparent decrease in concentration of determinable biologically active substances for the period of up to three years. Materials with potential risk of decomposition of biologically active substances by oxidation can be easily stabilized by presence of an inert gas (for instance, nitrogen or argon) that expels air oxygen from a hermetically closed container unit.

Table 1: Composition of important substances contained in Milk Thistle fruits (% by weight)

The said method of preparation makes it possible to integrate additional plant or other materials into the pressed material, either in the phase of the pressing itself (then this is common pressing, co-extrusion) or in the phase when the already pressed cake is mixed with this other material. Advantage of the first method is excellent homogenization of all materials, their integration and maximum preserving of contents of all active substances. The second method can be preferably utilized, in particular, in cases when the pressed cake serves for applying additional liquid material, e.g. chemically treated oil. Thus, the cake, that can, of its own, be a mixture of commonly pressed materials, serves as a carrier of the liquid phase. Even in this case, the materials are excellently mixed if an appropriate method of homogenization (e.g. in a conical rotary screw homogenizer) is used, wherein the liquid phase is uniformly spread on the surface of the carrier into which it can also be partially sucked.

The resulting complex material need not be stabilized with antioxidants or other stabilizers. Taste properties are acceptable for wide range of higher organisms; hence, these mixtures can be directly and without treatment fed to individual animal species. On the other hand, the resulting mixtures according to the invention can also be incorporated into complex feeding mixes by mixing, pressing, or other methods ordinary in preparing complex feeding mixes.

Main active substances are, above all, flavonolignans and then also biologically active substances from added other plant species. Concentration of these substances also determines recommended dosage of respective mixtures to certain animal species in certain indication. The base of dosing remains reaching of sufficient effective concentration of biologically active components in the internal environment of the organism so that intended biological effect could take place.

Influence of the prepared feeding mixes on conditions of animals has been verified on the bellow given specific examples by carried out clinical experiments. These experiments were thematically focused on safety of feeding and monitoring, in particular, of biochemical parameters of tested animals in comparison with reference groups, their utility value in case of farm animals, and overall physical condition in case of other animals.

For the purpose of this invention, the farm animals are, in particular, milk cattle - cows (e.g. breeds Holstein-Friesland, Jersey, Ayrshire, Brown-Swiss), meet cattle (e.g. breeds Hereford, Aberdeen Argus, Limousine), combined cattle (e.g. breed Bohemian parti-coloured cattle), meet poultry - hens (e.g. Plymouth White, Kornys White, hybrids: ISA 220, Ross 208, Ross 308), meet poultry - ducks (e.g. Tree-type, Aylesburg, Peking White), layers (e.g. Lenghorn White, Rodailand Red, hybrids: Hisex Brown, Dominant Brown, ISA Brown, Moravia BSL), police dogs (e.g. German shepherd), sports animals, in particular race horses (e.g. English thoroughbred), show-jumping horses (e.g. Bohemian hot-blood), draught horses (e.g. Kladruby Whitehorse), camels (e.g. one-humped camel), hunting dogs (e.g. Bohemian wire-haired dog, breeds of dachshunds).

Best Mode for Carrying Out of the Invention

Example 1 Preparation of the feed supplement from the fruits of Milk Thistle and the fruits of Moldavian Dragonhead.

Provided batch (15,000 kg) of the fruits of Milk Thistle (loss on drying 7.21 % by weight; content of flavonolignans 3.17 % by weight) was abraded on a peeling machine (type: „Ekonos" 3.22, manufacturer TMS Group, a.s., Pardubice) at weight flow-rate 800 kg.h^*1. At its outlet, the machine was fitted with a sieve of mesh size 1.2 x 20 mm. 2,200 kg of abraded outer layers of the fruits and 12,600 kg of the peeled fruits (Product 1) were obtained. Provided batch (450 kg) of the fruits of Moldavian Dragonhead was pressed on a press with multiple-run screw (type: "Shark" PL 200 LCPV7, manufacturer: Farmet, spol. s r. o, Ceska Skalice) under pressure 50 MPa; 372 kg of the pressed cake (Product 2) and 60 kg of fatty oil was obtained. Product 2 was crushed and sieved on the crushing machine (type: PS- 12, manufacturer Haiva, spol. s r. o., Tasovice) fitted with a sieve of mesh size 8 mm at the outlet. Flow rate of the material through the crusher was 6,000 kg.h^"1. In total, 367 kg of the crushed and sieved cake (Product 3) was obtained.

A part of the Product 1 (979 kg) and all Product 3 (367 kg) - weight ratio 8 : 3 - were also processed by mixing on a conical rotary screw homogenizer (type: HV.3000.1, manufacturer ERGON, n.p., Kromefiz) with the screw speed 1.0 s^"1 and the arm speed 0.033 s^"1 for 30 min; subsequently, the homogenized material was pressed on a press with multiple- run screw (type: "Shark" PL 200 LCPV7, manufacturer: Farmet, spol. s r. o, Ceska Skalice) under pressure 50 MPa; 1 140 kg of the pressed Product 4 and 200 kg of the fatty oil 2 was obtained. Product 4 was crushed and sieved on a crushing machine (type: PS- 12, manufacturer Haiva, spol. s r. o., Tasovice) fitted with a sieve of mesh size 12 mm at its outlet. Flow rate of the material through the crusher was 6,000 kg.h^"1. In total, 1134 kg of the crushed and sieved cake (Product 5) was obtained.

The Products 3 and 5 were sampled for pertinent analyses. Results of the analyses are shown in Tables 2 and 3.

Table 2: Results of analyses of individual products of processing the fruits of Milk Thistle and the fruits of Moldavian Dragonhead

Table 3: Results of sieve analysis of feed supplement from fruits of Milk Thistle and fruits of Moldavian Dragonhead

Example 2 Preparation of feed supplement from fruits of Milk Thistle and dried haulm of Leuzea Carthamoides

Provided batch (15,000 kg) of the fruits of Milk Thistle (loss on drying 7.21% by weight; content of flavonolignans 3.17 % by weight) underwent abrading on a peeling machine (type:„Ekonos" 3.22, manufacturer TMS Group, a.s., Pardubice) at weight flow-rate 800 kg.h^"1. At its outlet, the machine was fitted with a sieve of mesh size 1.2 x 20 mm. 2200 kg of abraded outer layers of the fruits and 12,600 kg of the peeled fruits (Product 6) was obtained.

Provided batch (500 kg) of the dried haulm (over-ground part) of Leuzea Carthamoides was crushed and sieved on a cylindrical grinder (type: S-300, manufacturer Romill, spol. s r. o., Brno) fitted at its outlet with a sieve of mesh size 8 mm. Flow rate of the material through the crusher was 1500 kg.h^"1. In total, 495 kg of the crushed and sieved haulm (Product 7) was obtained.

A part of product 6 (1000 kg) and a part of product 7 (100 kg) - weight ratio 10 : 1 - were also processed by mixing in a conical rotary screw homogenizer (type: HV.3000.1, manufacturer ERGON, n.p., Kromefiz) at the screw speed 1.0 s^"1 and the arm speed 0.033 s^"1 for 30 min; subsequently, the homogenized material was pressed on a press with double-run screw (type: L02X100.1 1, manufacturer: Tovarny mlynskych stroju, a.s., Pardubice) at speed 0,55 s^"1 under pressure 35 MPa; 825 kg of the pressed Product 8 and 223 kg of the fatty oil were obtained. Product 8 was crushed and sieved on a crushing machine (type: PS- 12, manufacturer Haiva, spol. s r. o., Tasovice) fitted at its outlet with a sieve of mesh size 12 mm. Flow rate of the material through the crusher was 6,000 kg.h^"1. In total, 820 kg of the crushed and sieved cake (Product 9) was obtained.

Product 9 was sampled for pertinent analyses. Results of the analyses are given in Tables 4 and 5.

Table 4: Results of analyses of product of processing Milk Thistle fruits and dried Leuzea

Carthamoides haulm

Product Loss on drying, Oil content, Content of flavonolignans, number % by weight % by weight % by weight, dry basis

9 7.17 8.66 4.26 Table 5: Results of sieve analysis of feed supplement from Milk Thistle fruits and dried Leuzea Carthamoides haulm

Example 3 Preparation of the feed supplement from the fruits of Milk Thistle and dried haulm of Leuzea Carthamoides in high concentration

Provided batch (500 kg) of the dried haulm (over-ground part) of Leuzea Carthamoides was crushed and sieved on a hammer grinder (type: DM-3/8, manufacturer Skiold A/S, Denmark) fitted at its outlet with a sieve of mesh size 8 mm. Flow rate of the material through the crusher was 1300 kg.h^"1. In total, 493 kg of the crushed and sieved haulm (Product 10) was obtained.

Provided batch (1000 kg) of the fruits of Milk Thistle (loss on drying 7.21 % by weight; content of flavonolignans 3.17 % by weight) and a part of Product 10 (250 kg) - weight ratio 4 : 1 - were also processed by mixing on a conical rotary screw homogenizer (type: HV.3000.1, manufacturer ERGON, n.p., Kromefiz) at the screw speed 1.0 s^"1 and the arm speed 0.033 s^'1 for 30 min; subsequently, the homogenized material was pressed on a press with a double-run screw (type: L02X100.i l, manufacturer: Tovarny mlynskych stroju, a.s., Pardubice) at speed 0.55 s^"1 under pressure 30 MPa; 1050 kg of the pressed Product 1 1 and 155 kg of the fatty oil were obtained. Product 11 was crushed and sieved on a crushing machine (type: PS- 12, manufacturer Haiva, spol. s r. o., Tasovice) fitted at its outlet with a sieve of mesh size 12 mm. Flow rate of the material through the crusher was 6,000 kg.h^'1. In total, 1044 kg of the crushed and sieved cake (Product 12) was obtained.

Product 12 was sampled for pertinent analyses. Results of the analyses are shown in Tables 6 and 7.

Table 6: Results of analyses of product of processing fruits of Milk Thistle and dried haulm of Leuzea Carthamoides in high concentration Product Loss on drying, Oil content, Content of flavonolignans, number % by weight % by weight % by weight, dry basis

12 8.31 6.9 2.97

Results of sieve analysis of feed supplement from fruits of Milk Thistle and dried haulm of Leuzea Carthamoides in high concentration

Example 4 Preparation of feed supplement from the fruits of Milk Thistle, dried haulm of

Leuzea Carthamoides in high concentration and conjugated fatty acid ethyl esters

Provided batch (500 kg) of Product 12 and 26.3 kg of Milk Thistle oil treated by conjugation of double bonds of fatty acids and transferred to the form of ethyl esters (content of conjugated linoleic acid ethyl esters 48 % by weight) were processed by mixing in a conical rotary screw homogenizer (type: HV.3000.1, manufacturer ERGON, n.p., Kromefiz) at the screw speed 1.0 s^"1 and the arm speed 0.033 s^"1 for 60 min. In total, 526 kg of the product (Product 13) was obtained.

Product 13 was sampled for pertinent analyses. Results of the analyses are shown in Tables 8 and 9.

Results of analyses of the product of processing the fruits of Milk Thistle and dried haulm of Leuzea Carthamoides in high concentration and conjugated fatty acid ethyl esters

13 7.89 7.4 2.81 Results of sieve analysis of feed supplement from fruits of Milk Thistle and dried haulm of Leuzea Carthamoides in high concentration and conjugated fatty acid ethyl esters

Summary of results of clinic experiments

Experiment 1 Flock of dairy cows of the breed Holstein-Friesland in stable breeding

The experiment was carried out with the flock of dairy cows in stable breeding fed with a feeding batch composed of haylage, dried lucerne fodder, fodder concentrate, and maize silage. The flock was divided into two halves - experimental and reference under otherwise the same conditions of feeding and stabling. The experimental group was fed with the feeding mix enriched with an additive of the processed fruits of Milk Thistle Product 5 in the amount of 150 g per one dairy cow and day for two subsequent 22-day periods. Monitoring in the subsequent third 22-day period was carried out in order to follow the parameters after stopping feeding the processed fruits of Milk Thistle. Evaluation indices: utility values of milk production of whole groups and utility values and biochemical clinic parameters of 12 individuals selected from both groups paired with respect to physical condition of an animal and the order of lactation. The results showed that average utility values of milk production in the experimental group were higher in respective periods, as shown in Table 10. The influence on increased utility value of milk production with respect to the order of lactation is shown in Table 11 - here, the highest positive effect of feeding the processed fruits of Milk Thistle was proved in the second or third lactation cycles. The monitored biochemical parameters (liver enzymes - ALT, AST, GMT, ionic balance, creatinine, and specific proteins of inflammation) did not show statistically significant influence.

Table 10: Overall utility value of milk production in dm per dairy cow and day Parameter 1st period of 22 2nd period of 22 3rd period of 22 days days days

Reference group 28.3 27.0 26.8

Experimental group 30.9 30.0 28.4

Relative increase in 9.2 1 1.1 6.0

% of experimental

group compared with

reference group

Table 11: Utility value of milk production in dm³ for 12 selected paired individuals with respect to order of lactation

Experiment 2 Group of dairy cows of the breed Holstein-Friesland with introduced ruminal cannula

The experiment was carried out with three experimental dairy cows fed with a feeding batch composed of dried lucerne fodder, fodder concentrate, and maize silage, with an introduced ruminal cannula. The first one-day period was reference; it was followed by the 11 -day experimental period when the standard feeding batch was enriched with 150 g of the material of Product 5 per dairy cow and day.

The aim of the experiment was to monitor dynamics of transition of the active substances (flavonolignans) from the administered processed fruits of Milk Thistle into the internal environment of experimental individuals by the method in sacco (nylon bags filled with the experimental material introduced into the rumen and subsequently taken out at specified intervals). The experiment proved that the administered batch of flavonolignans from the processed fruits of Milk Thistle was degraded in the rumen by about 30 %. Dynamics of degradation is shown in Table 12. The data in the table, in columns "Effective degradability at outflow rate k = 0.06 and/or k = 0.08" mean what amount, in percentage, of substances originally present in the nylon bag were transferred into solution and thus also to other parts of the digestive tract of the animal during the tested period. Here, the outflow rate is defined as a ratio of volume of the rumen juice transferred into other parts of the digestive tract per unit of time (one hour) to total volume of the rumen fluid. The most degraded substances were: polar flavonolignans (silydianin and silychristin); silybinin A and silybinin B, same as isosilybinin A and isosilybinin B, were degraded to lesser extent. By far the most degraded substance was taxifolin (more than 50 %) which, however, is not ranked among flavonolignans, even though it is their biosynthetic precursor. Total plasmatic concentrations of flavonolignans in blood of experimental individuals were determined on the 9th or 10th day from the start of feeding the tested material. The values found correspond to the range of 0.190 to 0.485 mg.dm^" of the sum of flavonolignans present predominantly in the form of their conjugates of the II. phase of metabolism of xenobiotics (glucuronides and sulphates).

Biochemical parameters in blood of the experimental individuals, profile of lower fatty acids and pH value in the rumen fluid were also monitored. In these parameters, no significant differences were found between the values of the reference and experimental periods.

Table 12: Degradability of dry matter and flavonolignans from processed fruits of Milk

Thistle. The degradability data are in percents.

Parameter Effective degradability at Effective degradability at outflow rate k - 0.06 outflow rate k = 0.08

Dry matter 47.2 45.9

Taxifolin 59.1 55.1

Silychristin 34.4 31.5

Silydianin 35.2 32.2

Silybinin A 25.1 22.8

Silybinin B 27.8 25.7

Isosilybinin A 23.3 21.1

Isosilybinin B 23.6 21.8

Sum of flavonolignans 28.0 25.5

without taxifolin

Sum of flavonolignans 30.2 27.7

including taxifolin Experiment 3 Group of race sports horses of the breed English thoroughbred

The experiment was carried out in two groups, reference and experimental, each of 5 horses of the breed English thoroughbred aged 3 - 8 years for the period of 60 days. Both groups were fed with the same feeding batch consisting of meadow hay, oats, load granules of firm Mikrop Cebin, plant oil, and mineral additive of firm Mikrop Cebin; in addition, the experimental group obtained a batch of 150 g of the processed fruits of Milk Thistle of Product 9 per individual and day. Biochemical parameters of blood were tested with stress on metabolism of parenchymatous organs (liver, kidney) with respect to protein, energy, mineral, and muscular metabolism. Moreover, clinic symptoms of the tested animals were evaluated: intake, fodders, activity were subjectively evaluated - influence on performance and vitality, allergic reaction, welfare - well behaviour. Results of the tests proved improvement of clinical symptom in the experimental group as shown in Table 13.

From the monitored biochemical parameters, haematological profile was not adversely influenced and the values of all monitored haematological parameters and mineral metabolism ranged within physiological values. On the other hand, parameters of the liver metabolism (ALT, AST, GMT and bilirubin) in the experimental group were dramatically improved in comparison with the reference group. Key results of relevant monitored parameters are shown in Table 14. These values suggest that the administered feed supplement according to this invention has a positive effect on all values of liver enzymes (transaminases - ALT, AST, GMT), which documents fast regeneration of liver cells in case of feeding horses with the feed supplement according to this invention in comparison with the reference group that was not fed with the given additive. In case of the group fed with the feed supplement according to this invention, lower loading of the organism is also documented by significant decrease of metabolites represented by reduced concentration of bilirubin. Similarly, the parameters of muscular metabolism (lactic acid, LDH, and creatinekinase) were improved in the experimental group compared with the reference group under the same muscular load.

Table 13: Clinical symptoms of feeding the processed fruits of Milk Thistle according to the invention in comparison with common fodder of race horses: scale 0 to 10 points (0 points = worst, 10 points = best) - average values of both groups

Parameter Reference group Experimental Difference

group Feeding batch intake 8.30 9.32 1.02

Activity 8.06 9.52 1.46

Allergic reaction 0.00 0.00 0.00

Welfare 9.20 9.50 0.30

Relevant biochemical parameters of liver enzymes in feeding the processed fruits of Milk Thistle according to the invention in comparison with common fodder of race horses - average values of both groups

0. day ... day before start of feeding the processed fruits of Milk Thistle

Experiment 4 Group of show-jumping sports horses of the breed Bohemian hot-blood

The experiment was carried out with the experimental group of 5 horses of the breed of Bohemian hot-blood aged 3 - 8 years for 60 days, in the so-called shortened form of monitoring. The group was fed with a feeding batch consisting of meadow hay, oats, load granules of firm Mikrop Cebin, plant oil, mineral additive of firm Mikrop Cebin, and 150 g of the processed fruits of Milk Thistle - Product 9 per horse and day. The tests included biochemical parameters of blood with stress on metabolism of parenchymatous organs (liver, kidney) with respect to protein, energy, mineral, and muscular metabolism.

Although the type of load of the show-jumping horses differs significantly from the load of the race horses, thee parameters of liver metabolism (ALT, AST, GMT, and bilirubin) were improved on feeding the processed fruits of Milk Thistle according to this invention, similarly to the case of Experiment 3. Similarly, two parameters of the muscular metabolism (lactic acid and LDH) showed improvement compared with the initial values of these biochemical parameters before the start of feeding the feed supplement according to this invention (0th day of the experiment). The level of creatinekinase increased, which can be explained as a result of permanent load of the locomotive organs. Key results of relevant monitored parameters are shown in Table 15. The results prove that the administered feed supplement according to this invention has a positive effect on all values of liver enzymes (transaminases - ALT, AST, GMT), which confirms fast regeneration of liver cells in case of feeding the feed supplement according to this invention compared with the initial state of the organism immediately before giving the feed supplement according to this invention. The tested individuals showed lower load of the organism after being fed the feed supplement according to this invention, as documented by the decrease of metabolites represented by reduced concentration of bilirubin. In the parameters of muscular metabolism, improved conditions were demonstrated of the tested individuals after they were fed with the feed supplement according to this invention, in particular, by dramatic decrease of level of lactic acid. This documents dramatic improvement of the muscular metabolism of the animal under after-load conditions and its ability to adapt more quickly to additional physical performance.

Table 15: Relevant biochemical parameters of liver enzymes in feeding show-jumping horses with the processed fruits of Milk Thistle according to the invention - average values

Industrial Applicability

The feed supplements based on the processed fruits of Milk Thistle according to this invention are an important form of feed supplements for a wide group of animals. Due to safety of feeding and showed positive effects on health of animals, they can be used for preventative and curative feeding of animals with a positive effect on their productivity, in case of farm animals, and overall health conditions and performance, in case of sports and domestic animals. Long-term shelf life of the said feed supplements, together with a possibility to feed them to animals either directly or integrated into various types of feeding mixes and products for animal nutrition, ensure their universal applicability and enable active substances from the feed supplements to be administered to animals in an optimum batch and form.

The said method of producing the feed supplements based on the processed fruits of Milk Thistle transfers active substances from this fruits (flavonolignans) into forms showing their high availability from the digestive tract and transfer into the internal environment of the animal where their biological properties beneficial to the animal health can be subsequently asserted.

Claims

1. Feed supplement, suitable, in particular, for feeding animals, characterized in that it contains whole and/or treated fruits of Milk Thistle {Silybum marianum (L.) GAERTNE ) and 10 % to 35 % by weight of crushed and/or milled and/or pressed parts of at least one additional plant species, relatively to weight of the fruits of Milk Thistle.

2. Feed supplement according to Claim 1, characterized in that the fruits of Milk Thistle are treated in such a way that its outer parts are removed.

3. Feed supplement according to Claim 2, characterized in that the outer parts of the fruits are removed in the range of 10 % to 40 % of original weight of the fruits of Milk Thistle.

4. Feed supplement according to Claims 1 to 3, characterized in that the outer parts of the fruits of Milk Thistle are removed using peeling and/or abrading.

5. Feed supplement according to any of Claims 1 to 4, characterized in that at least one additional plant species is chosen from the group comprising the genus Dragonhead and/or genus Leuzea and/or genus Amaranth and/or genus Clintonia.

6. Feed supplement according to Claim 5, characterized in that the genus Dragonhead contains at least the species - Moldavian Dragonhead {Dracocephalum moldavica L.), genus Leuzea contains at least the species - Leuzea Carthamoides {Leuzea Carthamoides L., syn. Rhaponticum Carthamoides Willd.), genus Amaranth contains at least the species - love-lies-bleeding {Amaranthus caudatus L.) or Amaranthus quitensis Kunth, and genus Clintonia contains at least the species - Yellow Clintonia {Prunella vulgaris L.).

7. Feed supplement according to any of Claims 1 to 6, characterized in that it further contains 5 to 15 % by weight of oil from the seeds of Milk Thistle treated by conjugations of double bonds.

8. Feed supplement according to Claim 7, characterized in that the added oil from the seeds of Milk Thistle treated by conjugation of double bonds consists of a mixture of higher fatty acid ethyl esters of chain length from C₁₄ to C₂₂.

9. Feed supplement according to Claim 8, characterized in that concentration of linoleic acids (C₁₈) ethyl esters with conjugated double bonds in the added treated oil falls within the range of 20 % to 60 % by weight.

10. Feed supplement according to Claims 1 to 9, characterized in that it is a form of tough, powdered, non-sintering particles of loose consistence of size up to 12 mm.

11. Method of producing the feed supplement according to Claims 1 to 6 comprising the steps:

a) crushing and/or milling and/or pre-pressing of added parts of at least one additional plant species;

b) mixing of thus treated plant parts with the whole and/or treated fruits of Milk

Thistle according to Claims 1 to 4, characterized in that it is followed by c) pressing of the obtained mixture at pressure 30 to 50 MPa, wherein 5 % to 10 % of fatty oil is separated which is composed of more than 85 % by weight of esters of higher fatty acids of chain length C₁₄-C₂₂ with tribasic alcohol, and a tough pressed mass - the cake is formed; and

d) crushing and sieving of the cake to specified distribution of particle size.

12. Method of production according to Claim 11, characterized in that the tribasic alcohol is propane- 1,2,3-triol.

13. Method production according to Claims 11 or 12, characterized in that the particle size distribution is purpose-controlled depending on the contemplated way of feeding and on the animal species.

14. Use of the feed supplement according to Claims 1 to 10 to direct feeding of animals or to integration into the feeding batch.

15. Use of the feed supplement according to Claims 1 to 10 for increasing utility value of farm animals.

16. Use of the feed supplement according to Claims 1 to 10 for increasing performance and improving liver and muscular metabolism of sports, domestic, and farm animals.