NZ554260A

NZ554260A - Selenomethionine andor selenocysteine administration to non-human animals to increase selenium content in protein sources

Info

Publication number: NZ554260A
Application number: NZ554260A
Authority: NZ
Inventors: Scott Oliver Knowles; Neville Donovan Grace; Rex Munday
Original assignee: Agres Ltd
Priority date: 2007-09-30
Filing date: 2007-09-30
Publication date: 2010-05-28
Also published as: WO2008121006A1; WO2008121006A8

Abstract

Disclosed is a method of increasing selenium content in protein sources from an animal, such as milk or meat, by orally administering a tablet containing selenocysteine, selenomethionine, or combinations thereof. Also disclosed is a tablet containing selenomethionine, for oral administration, to increase selenium content of protein sources such as milk and meat from that animal, wherein the tablet releases selenomethionine over at least one month.

Description

<div class="application article clearfix" id="description"> 554260 SELENIUM ADMINISTRATION STATEMENT OF CORRESPONDING APPLICATIONS This application is based on the provisional specification filed in relation to New Zealand Patent Application Number 554260, the entire contents of which are incorporated herein by reference. TECHNICAL FIELD The invention relates to selenium administration. More specifically, the invention relates to a method of orally administering a selenium source to an animal to increase the selenium content of the animal tissue and/or fluids. BACKGROUND ART Nutritional roles and value of dietary selenium Informed consumers increasingly demand foods with benefits beyond simple nourishment, matched to lifestyles, individual preferences and to meet specific dietary requirements. This has spurred interest in foods containing high levels of vitamins and minerals. Nutritional modification to make such foods may be achieved several ways, and in one embodiment utilises changes effected on-farm to directly improve the food without subsequent processing. One mineral of interest is selenium, a trace element essential to growth and health in humans and animals generally. In the body, selenium is incorporated into proteins to make selenoproteins, which are active in cell detoxification, redox cycling, and antioxidant defence against cellular damage from free radicals. Free radicals are natural by-products of oxygen metabolism that contribute to the development of chronic diseases such as cancer and heart disease. Other selenoproteins regulate thyroid function and play a role in the immune system. Except in extreme cases, selenium deficiency does not by itself cause illness. 2 554260 Rather, it can make the body more susceptible to illnesses caused by other nutritional, biochemical or infectious stresses. As for all essential minerals, diet is the source of selenium intake. Criteria in New Zealand and elsewhere define the selenium Recommended Dietary Intake (RDI) or Recommended Daily Allowance (RDA) for healthy consumers of various ages. New Zealand and Australian RDIs were set by the Australian National Health and Medical Research Council, and currently are: Population adult adult sub-group men women boys girls toddlers infants Selenium RDI 70 60-75 60 55 25 10-15 (pg/day) It should be appreciated by those skilled in the art that different countries have differences in RDI/RDA levels and the information above is provided by way of example only. The selenium nutrition status of New Zealanders tends to be below World Health Organisation recommendations. This condition is a consequence of the country's geology, which determines mineral content of soil, which in turn affects mineral uptake by crops and ultimately selenium content of locally grown foods. In other countries such as the UK, recent changes in farming and import practices have also caused a drop in the selenium nutrition status of the general population. People consuming a balanced and ample diet can usually meet their minimum intake requirement. However old habits, dieting or cultural practices that restrict variety of food choice can severely limit access to good selenium sources. Thus many consumers are looking for ways to boost their intake of selenium with nutritional supplements, often via pills but increasingly by eating nutrient-enhanced foods. 3 554260 Meat and milk from New Zealand livestock naturally contain selenium, albeit at low concentrations which reflect composition of the animals' grazing diets. Usual levels are 50 pg Se/kg (range 30-85 pg/kg) in fresh lean beef or lamb (West, 1996), and 2-11 pg Se/litre in whole milk (Grace et al. 2001). The contribution made to a person's daily selenium requirement by eating a 100 g or 100 ml serving of typical meat or milk would be 1-6% and 1-20% for adults and toddlers, respectively. Obviously, higher concentrations of selenium would increase the benefit derived from food servings. In these foods (and in staple grains as well) selenium exists primarily as the organic chemical form selenomethionine, an analogue of the amino acid methionine. Selenomethionine can be incorporated into proteins in place of methionine, and these proteins then serve as selenium storage. Selenomethionine derived from such foods is considered to have high bioavailability for the consumer, in that it is well absorbed and efficiently utilised. Improving food nutritional value by enriching selenium content Enhanced foods can be produced by adding a nutrient such as selenium exoaenouslv i.e. at the factory after harvest, or endoaenouslv by causing extra selenium to be grown into the food prior to harvest or slaughter. The advantage of the latter is that selenium is incorporated into the food in its natural chemical form, is delivered to the consumer in an unadulterated way, and post-harvest processing or concentrating steps are avoided. There are many known methods for endogenously enriching the selenium content of animal-derived foods. Feed additives One method is by adding selenium in its 'organic' form of selenomethionine or selenocysteine to the animal feed, as described for example in the Patent abstracts TW0565432B, CN1094899A and CN1217153A. Generally, these methods involve 4 554260 daily feeding of a selenium-rich feedstuff derived from selenised yeast, silages, and seleniferous forage plants. Alternatively, the feed additive may contain sodium selenite and sodium selenate, two inorganic forms of selenium salts. Examples of sodium selenite feed supplements are described in at least the abstracts of Chinese Patent No. CN1452884A, CN1439284A, CN1452883A and CN1341374. All these methods can be costly in just producing the feed, without taking into consideration labour costs for their daily distribution. Also, the amount of specialised feed consumed by the animal may vary on a day to day basis, as this depends on the appetite of the animal. Many farming systems do not allow for easily monitoring the amount of feed consumed by the animals, particularly if the animals graze in large herds or flocks. Additionally, the feed additive products may by poorly defined, and contain mixed composition and variation in the profile of selenium chemical forms present in the feed. This in turn varies the amount of selenium taken up by the animal. Oral dosages Oral doses and liquid 'drenches' have been used for increasing the nutrient content of the meat and milk of an animal. Knowles S.O. et al. (1999) describes drenches containing either un-purified raw yeast enriched with selenoamino acids or inorganic sodium selenate to treat selenium-deficient cows. The study showed that by administering drenches three times a week, the concentration of selenium in the animals' blood, liver, milk and casein could be substantially increased. However, this method has a number of practical disadvantages, as the administration can be messy and time consuming, especially if a large number of animals have to receive the formulation. Further, the use of a short-acting drench means that the drench must be administered at frequent intervals to obtain the desired result, therefore adding to an increase in both time and money. 5 554260 Other methods for endogenously enriching the nutrient content of animal-derived products are summarized in Knowles, S.O. et al. (2004 and 2006). The methods described in these publications include specific diets, parenteral supplements and modification of the ruminal microflora. These articles describe increased concentrations of selenium in meat and milk of animals through using organic and inorganic feed additives, drenches and subcutaneous injections. However, that article only summarises the various techniques available, which have been reviewed above, without providing any experimental description of the techniques. The use of purified selenomethionine as an active compound is not disclosed. Injections Injectable selenium supplements are available in New Zealand that contain inorganic sodium selenite or sodium selenate. Their purpose is therapeutic, with none claiming to affect meat or milk composition. Parental administration via injection of a selenomethionine formulation, as described in the applicants' patent, New Zealand Patent No. 540958, is another example of a method for administering a nutrient to an animal. Tablets and boluses Rumen boluses are a common method for delivering nutrients to ruminant animals. There are a number of rumen boluses available on the market, to deliver various elements, including inorganic and elemental selenium. However, the large number of documents that are publicly available referring to bolus technology describe the use of a bolus only for the maintenance of animal health or as a therapeutic use -i.e. to act as a nutritional supplement for the animal's dietary intake. None of these documents describe the use of this technology for a non-therapeutic use - i.e. to increase the selenium concentration in the animal's meat or milk leading to enriched foods for consumers. 6 554260 Grace et al. (1997) describes administering to a number of cows two to three 30 g boluses (containing elemental selenium of 10% Se and 90% Fe), to assess the effects on the concentration of selenium in the cows' blood and milk. The results showed that there was an increase in selenium concentration due to bolus administration. However, the study demonstrates that such a treatment scheme is impractical for farmers, with a number of disadvantages, such as: • As many as three boluses were required to change milk selenium concentration; • The total selenium dose per cow was large; • The milk selenium response was statistically significant (at about 2-fold increase) but not very great, such that the additional nutritional benefit for consumers would not be markedly advantageous nor would the economic added-value of collecting and processing that milk; and • Elemental selenium in the bolus is an inorganic chemical form that is distributed into and stored in the animal's tissues with poor efficiency, such that a significant amount of the selenium dose is lost through animal excretion into the environment. To overcome the numerous disadvantages of the various techniques described in the prior art, the inventors of the present invention considered developing and using a bolus containing purified selenomethionine that would be long-acting to deliver a constant supply of bioavailable selenomethionine for increasing the selenium content of animal tissues and products. A controlled release bolus would provide a more consistent means of selenium supplementation than do feedstuffs or salt-mineral mixes because the appetite and 7 554260 food consumption of the animal does not play a role in daily intake, nor does diversity in feedings systems within and between dairying regions. Because selenomethionine is efficiently absorbed by the animal and transferred to edible tissues (milk and meat), only insignificant amounts of selenium are excreted into the environment. The boluses formulated by the applicant (as referred to above) dissolve completely in the rumen and leave no residue or waste that might affect slaughter and sale of the animal. Examples of the bolus technology can be seen in the applicants' own patents and/or patent applications. For example, New Zealand Patent No. 504631 describes a device that enables the delivery of a biological agent, such as a fungus to reduce nematode parasites in animal, over a period of time. Also, New Zealand Patent No. 250544 refers to the therapeutic use of a bolus containing various elements, including selenium. However this patent refers only to the therapeutic use of the element and not to the non-therapeutic use the element may have. Another example of the use of a bolus for delivering elements for a therapeutic effect only is described in W005082270A1. The mechanisms and compositions of controlled release rumen boluses range from simple to complex. Examples include a blend of elemental iron plus elemental selenium, a homogeneous solid matrix of water-soluble glass containing sodium selenite plus other elements, and Push-Melt™ technology utilising a semipermeable membrane, osmotic tablet, wax plug piston and capscreen to force a sodium selenite formulation out through an exit port. None of these boluses use selenomethionine. Further, none of these boluses describe application of their product for anything other than animal therapeutic use. It should be appreciated that it would be desirable to have an orally delivered selenium source that is precisely chemically defined, easily and accurately dosed 8 554260 to animals, provides a known and reproducible delivery of active ingredient, and efficiently increases the selenium content of animal tissue and products for use in nutritious foods. It is an object of the present invention to address the foregoing problems or at least to provide the public with a useful choice. All references, including any patents or patent applications cited in this specification are hereby incorporated by reference. No admission is made that any reference constitutes prior art. The discussion of the references states what their authors assert, and the applicants' reserve the right to challenge the accuracy and pertinency of the cited documents. It will be clearly understood that, although a number of prior art publications are referred to herein, this reference does not constitute an admission that any of these documents form part of the common general knowledge in the art, in New Zealand or in any other country. It is acknowledged that the term 'comprise' may, under varying jurisdictions, be attributed with either an exclusive or an inclusive meaning. For the purpose of this specification, and unless otherwise noted, the term 'comprise' shall have an inclusive meaning - i.e. that it will be taken to mean an inclusion of not only the listed components it directly references, but also other non-specified components or elements. This rationale will also be used when the term 'comprised' or 'comprising' is used in relation to one or more steps in a method or process. Further aspects and advantages of the present invention will become apparent from the ensuing description which is given by way of example only. DISCLOSURE OF INVENTION The inventors of the present invention have found that by increasing the selenium content of an animal's proteinaceous tissues, a non- or extra-therapeutic benefit or 9 554260 effect is achieved. For the purposes of the specification, the term 'oral administration' refers to administration via the mouth. The term 'non-therapeutic' or 'extra-therapeutic' refers to when the element (for example selenium) being deposited into animal protein sources results in effects that are not exclusively therapeutic. The term 'protein sources' refers to various tissues and/or fluid secretions of an animal which have a composition that may be substantially high in protein. Preferably, the protein sources may be milk from an animal. In other preferred embodiments, the protein sources may be meat from an animal. It will be appreciated that the meat from an animal may refer to various edible parts of an animal, including the muscle meat, and offal such as the liver or the heart. Preferably, the animal may be a ruminant animal. More preferably, the animal may be cattle, sheep, goats or deer. However this should not be seen as limiting as various types of other ruminant animals may be food sources in other cultures and countries. According to first aspect of the present invention there is provided a method of increasing selenium content of protein sources of an animal by oral administration of a tablet containing at least one selenium source. According to another aspect of the present invention there is provided a method of inducing a non-therapeutic effect in an animal by oral administration of a tablet containing selenium source including selenomethionine. According to a further aspect of the present invention there is provided a composition containing selenomethionine formulated as a tablet for oral administration. 10 554260 According to a further aspect of the present invention there is provided use of a composition, containing a selenium source including selenomethionine, formulated as a tablet, to increase the selenium content of a protein source in an animal. According to a further aspect of the present invention there is the use of a tablet for oral administration, containing at least one selenium source, to provide an increase in selenium content of animal protein sources. According to further aspect of the present invention, there is provided the use of a tablet containing selenium source including selenomethionine, for oral administration, to induce a non-therapeutic effect in an animal. It is envisaged that a person skilled in the art will know of the various ways to manufacture a controlled release bolus. For example, the applicants' own patents, New Zealand Patent No. 504631, New Zealand Patent No. 250544 and New Zealand Patent No. 278977 which are all examples as to how a bolus may be manufactured and are incorporated herein, by way of example only. Preferably, the selenium source may be an 'organic' form of selenium. For example, the selenium source may be selenocysteine or selenomethionine, or their enantiomeric or racemic mixtures, or a combination thereof. In preferred embodiments, the selenium source may be selenomethionine. In an alternative embodiment, both an inorganic and organic selenium source may be used, in order to address a selenium deficiency and to enhance the selenium content of animal protein sources. In some preferred embodiments, selenomethionine may be in the chemical structure of either; the L-enantiomer form or D-enantiomer form. In other preferred embodiments, the chemical structure of selenomethionine may be a mixture of both L- and D-enantiomer forms. 11 554260 In one embodiment the selenomethionine used may have a chemical structure that can be represented by formula (I): o nh2 FORMULA (i) In one embodiment, the tablet may contain purified selenomethionine as a small percentage of the total bolus mass. Preferably, the selenium source may be in a finely divided powder form. More preferably, the selenium source may be suited to homogeneous incorporation into a solid matrix of inert carrier compounds. Preferably, the content of selenomethionine may be 100-5000 milligram per bolus. As selenomethionine contains about 40% elemental selenium, the concentration of selenomethionine is equivalent to 40-2000 milligram of selenium. Preferably, the selenium may be released slowly by the bolus into the animal's rumen so as to deliver 0.3-20 milligram selenium per day over a period of at least one month. Preferably, the tablet may be a long-acting, controlled release device. Preferably, the tablet releases the selenium source at a controlled and steady rate. More preferably, the tablet releases the selenium source over a time period of at least one month. Preferably, the tablet may be a bolus. The term 'bolus' refers to a large pill or tablet type composition that is administered orally and which tends to stay and erode or dissolve in the rumen of the animal over a prolonged time period. 12 554260 The inventors of the present invention consider that by oral administration of a tablet containing selenomethionine, large increases in the concentration of selenium in the animal protein sources may be seen, in comparison to no treatment. By providing this endogenous enrichment, fresh milk, milk fractions or dry powders made from the animal milk would not need to have selenium added exogenously during powder processing such as in the production of infant formula or other nutritious foods, which is current manufacturing practice. Other food examples include selenium-enriched meats. The composition may include selenomethionine and a carrier. Preferably, the carrier may be suitable binding agents and/or solubilising agents that serve to control the size, shape and dissolution rate of the bolus. It is will be appreciated that a person skilled in the art will know of suitable binding agents and/or solubilising agents, for example, a binding agent may be a fatty acid ester. While a suitable solubilising agent may be polyethylene glycol stearate. It will be appreciated that the composition may be used to supply selenomethionine to provide an increase in the selenium content of a protein source in an animal. Preferably, the use of the composition may be for a non-therapeutic effect. In an alternative embodiment the composition has both a therapeutic and non-therapeutic effect. According to a further aspect of the present invention there is provided a method of elevating the concentration of selenium in an animal by oral administration of a composition substantially as described above. According to a further aspect of the present invention there is provided a method of increasing the concentration of selenium in protein sources of an animal by oral administration of a composition substantially as described above. 13 554260 It is envisaged that the preferred embcxiiments of the present invention will have a number of advantages over the prior art, including: • decreased time and labour costs for administration, in comparison compared to daily feed additives; • precise chemical composition, in comparison to selenium-rich additives derived from selenised yeast, silages or seleniferous forage plants; • more accurate dose rates and reproducible delivery of active ingredient to the animal, without variation due to differences in animal dietary intake; • decreased animal handling and stress, in comparison to short acting oral drenches that require frequent dosing; • greater selenium response in animal protein sources, in comparison to inorganic forms of selenium; • lesser selenium wastage and excretion into the environment. BRIEF DESCRIPTION OF DRAWINGS Further aspects of the present invention will become apparent from the ensuing description which is given by way of example only and with reference to the accompanying drawings in which: Figure 1 shows a graph correlating bolus in situ lifespan and the amount of Lipomulse™ in the core formula, as discussed in Example 1; Figure 2 shows the in situ erosion rate over time of six instances of one bolus formulation containing 1.79% Lipomulse™, as discussed in Example 3; and 14 554260 Figure 3 compares the mean blood selenium concentrations of the test cows from Example 3 to the correlation of milk and blood selenium concentrations from cows treated with organic selenium by Knowles et al 1999. BEST MODES FOR CARRYING OUT THE INVENTION Examples describing the invention are now provided below. It should be appreciated that the following best modes are given by way of example only to perform the invention. Example 1 • Investigations into the best formula for rumen bolus prototypes To create a slow-release bolus with lifespan of 1-4 months, the effect of altering the percentage of releasing agent (Lipomulse™) in the core formula was tested. As known in the art, Lipomulse™ is a blend of glyceryl stearate and PEG-100 stearate. It is a non-ionic emulsifier and viscosity builder that affects density and in situ erosion rate of the bolus, which determines its longevity or lifespan and the daily release rate of any active ingredient mixed with the core formula. The purpose of this example was to assess the carrier dissolvability prior to adding the active ingredient, therefore these bolus prototypes did not contain any selenomethionine as the active ingredient. The basic composition of all the boluses was the same. The boluses were made as follows: 60 g of glycerol monostearate (Mono-Di) plus the set amount or percentage of Lipomulse™ was melted in a glass beaker at ~ 140°C. 200 g sieved barium sulphate and 200 g iron powder were added gradually, with stirring. The cooled mixture was ground to a fine powder and boluses made individually by compression, as described in the lab-scale manufacturing method discussed in Example 2 below. 15 554260 As stated above, the percentage of Lipomulse™ in the mixture was varied between 1.0 and 4.0%. Three boluses of each formulation were manufactured. The lifespan of the resulting boluses was tested in situ in a field trial using fistulated cattle. Fistulated animals have been surgically fitted with a permanent resealable port into their rumen that allows materials to be placed and withdrawn without trouble or discomfort. The boluses were placed individually in the rumen on Day 1 of the trial and were removed approximately weekly for examination and accurate weighing. Because the bolus composition eventually dissolves leaving no residue, the lifespan of a bolus extends until it is completely gone from the rumen. The lifespan (days) of the boluses in comparison with the percentage of Lipomulse™ are outlined in Table 1 below and in Figure 1. Percentage of Lipomulse™ Mean measured lifespan (days) 1.00% 130 1.25% 100 1.50% 70 1.75% 48 2.00% 40 2.00% (repeat) 50 2.50% 38 3.00% 25 3.50% 24 4.00% 20 TABLE 1: Comparison of bolus lifespan (days) and percentage of Lipomulse™ As shown in Table 1 and Figure 1, at low concentrations of Lipomulse™, the lifespan was dependent on concentration. At the higher concentrations, adding more Lipomulse™ had little effect on lifespan, suggesting that some other factor is controlling erosion rate, for example the wax coating. 16 554260 At 1% Lipomulse™, the boluses lasted 130 days (4.3 months), indicating that it is possible to make quite long-lasting boluses with this type of composition. Example 2 - Preferred formula and method for lab-scale manufacture of rumen boluses for slow-release delivery of selenomethionine to livestock The following lab-scale manufacturing procedure was used to create a bolus weighing 140 g and containing 0.695 g of selenomethionine: 1. 60 g of glycerol monostearate (Mono-Di) and 8.43 g Lipomulse™ (releasing agent) were melted in a glass beaker and brought to - 140°C. 2. 200 g sieved barium sulphate and 200 g iron powder (densifiers) were added gradually with vigorous stirring. 3. The paste was scraped from the beaker onto a sheet of aluminium foil and allowed to cool. During cooling, the paste was moved around with a spatula to avoid setting in a lump. 4. When cool, the material was ground in a coffee grinder and the resulting powder was passed through a flour sieve. 5. To 140 g of the powder mixture, 0.695 g selenomethionine powder was added and mixed thoroughly. 6. A compression die, 25 mm diameter by approximately 150 mm length (about 75 ml volume), was prepared. 7. 70 g of the powder mixture was loaded into the die, tamping firmly with a spatula. This was slowly pressed to 1,000 pounds per square inch (psi). A further 70 g of mixture was then added, and slowly pressed to 5,000 psi. 17 554260 8. The bolus was carefully removed from the die, and the moulding ridge removed from the rounded end with fine sandpaper. 9. An ethanolic solution of shellac was prepared by heating 20 g shellac with 150 ml absolute ethanol to boiling. After leaving overnight at room temperature, the mixture was filtered. 10. The bolus was painted with the shellac solution. 11. A small hole was made in the flat end of the bolus by drilling, and a cup hook was screwed into the hole. 12. A molten mixture of 245 g beeswax, 43.5 g carnauba wax and 210 g sieved barium sulphate was prepared at ~ 110°C. The mixture was constantly stirred in order to maintain the barium sulphate in suspension. 13. The bolus was dipped 3 times into the molten mixture. After cooling, the cup hook was removed and the flat end scraped with a scalpel. The finished bolus contained 0.695 g selenomethionine. Other quantities are easily possible, in order to create boluses with different delivery rates and durations. Example 3 - Investigations into the erosion rate and biological effect of rumen bolus prototypes tested in situ in fistulated cattle The following field trial was carried out to assess the bolus erosion rate and biological effect. Two experimental groups of fistulated non-lactating dairy cows were assessed. These were: 1. Three control cows. No bolus was administered. 18 554260 2. Six treated cows. Each cow was administered 1 selenomethionine bolus each. The boluses were prepared as described in Example 2 with 1.79% Lipomulse™ and 0.695 g selenomethionine, and were designed to last up to 2 months. The boluses were placed in the rumen on Day 1 and were removed weekly for examination and accurate weighing. Blood samples were collected from the cows on day 38 and day 56 of the experiment. Evaluation of bolus erosion rate As each bolus dissolves it liberates its active ingredient, so measuring bolus total weight loss is an accurate index of the release rate. Slow and steady weight loss indicates controlled sustained release of the selenomethionine payload. Good bolus design will prevent early spiking of release rate (as the bolus settles in to the rumen environment) as well as end-stage spiking (as the last remnants of the bolus crumble and dissolve). Figure 2 shows that these boluses eroded and dissolved appropriately, releasing their payload in a predictable manner. The erosion rate of boluses with core formula containing 1.79% Lipomulse™ was relatively constant at about 22 g per week, which equates to a selenomethionine release of 0.11 g per week or 0.015 g per day. As selenomethionine contains about 40% elemental selenium, this is 0.0062 g selenium per day (6.2 mg Se/day). Effect of selenomethionine release on cow blood selenium concentrations Selenomethionine released from the bolus into the rumen is readily absorbed into the cow and distributed to tissues including blood. In the above experiment absorption by the cows of 6.2 mg Se/day for 5-7 weeks substantially increased 19 554260 their selenium status. This is indicated in Table 2 below by way of the mean (± std error) blood selenium concentrations measured on two days. Mean blood selenium concentration Day 38 Day 56 Control cows ug Se/L 55 ±7 78 ±5 Test cows ug Se/L 145 ± 15 188 ±20 TABLE 2: Comparison of blood selenium concentrations in control and test cows at day 38 and day 56 of the experiment. Theoretical milk selenium concentration due to selenomethionine treatment These were non-lactating cows so direct assessment of milk selenium concentration was not possible. However we can estimate the milk response based on the inventors experience with lactating cows treated with oral selenomethionine. An experiment performed and described by Knowles et al 1999 mimicked the effect of sustained release of selenomethionine by manually giving cows a daily oral dose of selenised yeast equivalent to 2 or 4 mg Se/day for at least 11 weeks. Selenium concentrations were measured in both blood and milk, and correlations were determined. We can use that insight to estimate the theoretical milk effect of the selenomethionine bolus treatment performed in Example 3. Figure 3 shows data from the 1999 experiment: blood selenium concentrations of each cow are plotted against milk selenium concentrations measured at the same time during 11 weeks of the trial. The blood-milk correlation and dose-response characteristics of selenomethionine treatment are apparent. Overlaid on the figure are data from the bolus experiment performed in Example 3. Mean blood selenium 20 554260 concentrations of the control and treated groups on Day 38 and Day 56 are extrapolated to show theoretical milk selenium concentrations. Given that the current blood concentrations are where we would expect them for a selenomethionine bolus release rate of 6.2 mg Se/day, it is reasonable to expect that milk selenium concentrations would also respond as expected if boluses were administered to lactating dairy cows. Aspects of the present invention have been described by way of example only and it should be appreciated that modifications and additions may be made thereto without departing from the scope thereof as defined in the appended claims. 21 554260 REFERENCES Givens Dl, Allison A, Cottrill B, Blake JS. Enhancing the selenium content of bovine milk through alteration of the form and concentration of selenium in the diet of the dairy cow. Journal of the Science of Food and Agriculture 84:811-817, 2004. Grace ND, Ankenbauer K, Alexander AM, and Marchant RM. Relationship between blood selenium concentration or glutathione peroxidase activity, and milk selenium concentrations in New Zealand dairy cows. NZ Veterinary Journal 49:24-28, 2001. Grace ND, Lee J, Mills AR and Death AF. The influence of Se status on milk Se concentrations in dairy cows. NZ Journal of Agricultural Research 40:75-78, 1997. Heard JW, Walker GP, Royle PJ, Mcintosh GH, Doyle PT. Effects of short-term supplementation with selenised yeast on milk production and composition of lactating cows. Australian Journal of Dairy Technology 59,199-203, 2004. Knowles SO, Grace ND, Wurms K, and Lee J. Significance of amount and form of dietary selenium on blood, milk, and casein selenium concentrations in grazing cows. Journal of Dairy Science 82:429-437, 1999. Knowles SO, Grace ND, Knight TW, McNabb WC, and Lee J. Adding nutritional value to meat and milk from pasture-fed livestock. NZ Veterinary Journal 52:342-351,2004. Knowles SO, Grace ND, Knight TW, McNabb WC, Lee J. Reasons and means for manipulating the micronutrient composition of milk from grazing dairy cattle. Animal Feed Science and Technology 131:154-167, 2006. West J. Compilation of nutritional data for New Zealand beef and lamb in domestic and export trade. Project 96MZ 64/4.3. New Zealand Meat Research and Development Council, Wellington, NZ, 1996. Unpublished report. 22 554260 </div>

Claims

<div class="application article clearfix printTableText" id="claims"> CLAIMS

1. A method of increasing selenium content of protein sources of a non-human animal by oral administration of a tablet containing at least one selenium source to the non-human animal selected from; selenocysteine; selenomethionine; and combinations thereof.

2. A method of inducing a non-therapeutic increase in the selenium content of a non-human animal's protein sources by oral administration of a tablet containing a selenium source including seienomethionine to the non-human animal.

3. The method as claimed in any one of the above claims, wherein the protein source is the animal's milk.

4. The method as claimed in any one of the above claims, wherein the protein source is the animal's meat.

5. The method as claimed in any one of the above claims, wherein the selenium source is selenomethionine which has the chemical structure of either; the L-enantiomer form, D-enantiomer form, or a combination thereof.

6. The method as claimed in any one of the above claims, wherein the selenium source is selenomethionine which has a chemical structure that can be represented by formula (I): FORMULA (I)

7. The method as claimed in any one of the above claims, wherein the selenium source is selenomethionine which is in a finely divided powder form.

8. The method as claimed in claim 7, wherein the powdered selenium source is suited to homogeneous incorporation into a solid matrix of inert carrier compounds.

9. The method as claimed in any one of the above claims, wherein the selenomethionine is at a concentration of 100-5000 milligram per tablet.

10. The method as claimed in anyone of the above claims, wherein the selenomethionine is at a concentration equivalent to 40-2000 milligram of elemental selenium. o H nh2 15 554260

11. The method as claimed in any one of the above claims, wherein the selenium is released to deliver 0.3-20 milligram selenium per day over a period of at least one month.

12. The method as claimed in any one of the above claims, wherein the animal is a ruminant animal.

13. The method as claimed in any one of the above claims, wherein the tablet releases the selenium source at a controlled and steady rate.

14. The method as claimed in any one of the above claims, wherein the tablet releases the selenium source over a time period of at least one month.

15. The method as claimed any one of the above claims, wherein the tablet is a bolus.

16. A tablet formulated for oral administration and which on administration increases the selenium content of a non-human animal's protein sources containing selenomethionine which has a chemical structure represented by the formula (I): o NH2 FORMULA (I), and wherein on administration, selenomethionine is released from the tablet over a time period at Seast one month.

17. The tablet as claimed in claim 16, wherein the selenomethionine is in a finely divided powder form.

18. The tablet as claimed in claim 17, wherein the powdered selenomethionine is incorporated into a solid matrix of inert carrier compounds.

19. The tablet as claimed in any one of claims 16 to 18, wherein the selenomethionine is at a concentration of 100-5000 milligram per tablet.

20. The tablet as claimed in any one of claims 16 to 18, wherein the selenomethionine Is at a concentration equivalent to 40-2000 milligram of elemental selenium.

21. The tablet as claimed in any one of claims 16 to 18, wherein the selenium is released to deliver 0.3-20 milligram selenium per day over a period of at least one month.

22. The tablet as claimed in any one of claims 16 to 21, wherein on administration, 16 554260 selenomethionine is released from the tablet at a controlled and steady rate.

23. The tablet as claimed in any one of claims 16 to 22, wherein the tablet is a bolus.

24. The tablet as claimed in any one of claims 16 to 23, wherein the tablet is administered to a ruminant animal.

25. The tablet as claimed in any one of claims 16 to 24, wherein the protein source is the animal's milk.

26. The tabiet as claimed in any one of claims 16 to 25, wherein the protein source is the animal's meat.

27. Use of a selenium source selected from selenocysteine; selenomethionine; and combinations thereof, in the manufacture of a tablet formulated for oral administration to provide an increase in the selenium content of animal protein sources.

28. Use of a selenium source selected from selenocysteine; selenomethionine; and combinations thereof in the manufacture of a tablet formulated for oral administration, to induce a non- or extra-therapeutic increase in the selenium content of an animal's protein sources.

29. The use as claimed in either one of claim 27 or claim 28, wherein the protein source is the animai's milk.

30. The use as claimed in any one of claims 27 to 29, wherein the protein source is the animal's meat.

31. The use as claimed in any one of claims 28, wherein the selenomethionine has the chemical structure of either; the L-enantiomer form, D-enantiomer form, and combinations thereof,

32. The use as claimed in any one of claims 28 to 32, wherein the selenomethionine has a chemical structure represented by formula (I): o nh2 FORMULA (I).

33. The use as claimed in any one of claims 28 to 33, wherein the seienium source is 17 554260 selenomethionine which is in a finely divided powder form.

34. The use as claimed in claim 34, wherein the powder form of selenomethionine is mixed homogeneously into a solid matrix of inert carrier compounds.

35. The use as claimed in any one of claims 28 to 35, wherein the selenomethionine is at a concentration of 100-5000 milligram per tablet.

36. The use as claimed in any one of claims 28 to 35, wherein the selenomethionine is at a concentration equivalent to 40-2000 milligram of elemental selenium.

37. The use as claimed in any one of claims 28 to 35, wherein the tablet releases selenomethionine sufficient to deliver 0.3-20 milligram selenium per day over a period of at least one month,

38. The use as claimed in any one of claims 28 to 29, wherein the animal is a ruminant animal.

39. The use as claimed in any one of claims 28 to 30, wherein, on administration, the selenium source is released at a controlled and steady rate.

40. The use as claimed in any one of claims 28 to 31, wherein, on administration, the selenium source is released over a time period of at least one month,

41. The use as claimed in any one of claims 28 to 32, wherein the tablet is a bolus.

42. A food product containing a protein source derived from an animal subjected to the method as claimed in any one of claims 1 to 15.

43. Meat product containing elevated levels of selenium derived from an animal subjected to the method as claimed in any one of claims 1 to 15.

44. Milk or milk derived products containing elevated levels of selenium derived from an animal subjected to the method as claimed in anyone of claims 1 to 15.

45. Use as claimed in any one of claims 27to 41 to produce a food endogenously enriched with selenium.

46. Use as claimed in any one of claims 27to 41 to produce meat endogenously enriched with selenium.

47. Use as claimed in any one of claims 27to 41 to produce milk or a milk derived product endogenously enriched with selenium. 18 554260

48. A method of increasing the selenium content of protein sources of a non-human animal by oral administration of a tablet containing at least one selenium source to the non-human animal, substantially as described herein with reference to Examples 1 to 3.

49. A method of inducing a non-therpeutic increase in the selenium content of a non-human animal's protein sources by oral administration of a tablet to the non-human animal substantially as described herein with reference to Examples 1 to 3.

50. A tablet formulated for oral administration and which, on administration, increases the selenium content of a non-human animal's protein sources, substantially as described herein with reference to Examples 1 to 3.

51. Use of a selenium source in the manufacture of a tablet formulated for oral administration, to increase the concentration of selenium in animal protein sources, substantially as described herein with reference to Examples 1 to 3.

52. Use of a a selenium source selected from selenocysteine; selenomethionine; and combinations thereof in the manufacture of a tablet formulated for oral administration, to induce a non- or extra-therapeutic increase in the selenium content of an animal's protein sources, substantially as described herein with reference to Examples 1 to 3. AgResearch Limited by their Attorneys JAMES & WELLS 19 </div>