AU2006200870B2

AU2006200870B2 - Element deficiency and prevention treatment

Info

Publication number: AU2006200870B2
Application number: AU2006200870A
Authority: AU
Inventors: Wayne Frederick Leech; Donald George Mclaren; Karthigeyan Nanjan
Original assignee: Bomac Research Ltd
Current assignee: Bomac Research Ltd
Priority date: 2005-03-02
Filing date: 2006-03-01
Publication date: 2010-11-11
Anticipated expiration: 2026-03-01
Also published as: GB2423712B; NZ538587A; AU2006200870A1; GB2423712A; IE20060150A1; GB0604022D0; ZA200601797B

Description

AUSTRALIA Patents Act 1990 COMPLETE SPECIFICATION STANDARD PATENT Applicant: Bomac Research Limited Invention Title: ELEMENT DEFICIENCY AND PREVENTION TREATMENT The following statement is a full description of this invention, including the best method of performing it known to us: ELEMENT DEFICIENCY AND PREVENTION TREATMENT TECHNICAL FIELD The invention relates to element deficiency and prevention treatments. 5 More specifically, the invention relates to a medicament for use and methods in the prevention or treatment of deficiencies in the elements iodine and selenium, particularly in relation to farm animals. BACKGROUND ART Element deficiencies in animals, including humans, are known to result in a 10 variety of disease states. As a result many health supplements have been developed for animals and humans to prevent element deficiencies and therefore avoid associated disease states. In the case of farm animals such as sheep and cattle, element deficiencies may occur as a result of nutrient deficiencies in feed which lacks one or more 15 elements required for correct metabolic function. For example, brassica crops often used as an animal feed typically contain low levels of iodine. In such cases, one solution used to prevent or treat the deficiency is to administer to the animal a dose of the element or elements either before onset of the disease state or as a treatment once symptoms of the disease are exhibited. 20 In general, it is desirable that doses be long acting so that the dose need only be administered to an animal on an infrequent basis. Two key elements which are commonly deficient when farming livestock such as sheep and cattle are the elements iodine and selenium. 1A Iodine is an essential element required by the thyroid gland for the manufacture of hormones thyroxine (T4, and tri-iodothyronine (T 3 ). Deficiencies in iodine may lead to a variety of diseases, the symptoms and effects of which are often accentuated in young growing animals. By way of example, iodine deficiency 5 diseases such as goitre and may be associated with prolonged gestation, dystocia and reduced survival of progeny. Also, iodine deficiency of the foetus in a ewe may influence progeny tissue growth and tissue differentiation, resulting in development abnormalities in the central nervous system, gonads, heart, skin, wool and follicles of progeny. 10 One widely used and long acting iodine medicament is Flexidine T M . This is a clear to slightly cloudy oily liquid medicament for administration preferably by injection into the muscle (intramuscularly). The primary constituent of the medicament is iodised peanut oil. The iodine (approximately 26% w/v of the medicament) is bound to ethyl esters of the unsaturated fatty acids in peanut oil 15 (primarily oleic and linoleic). This medicament provides an animal approximately 12 months or more of sustained levels of iodine or at least sufficient amounts to avoid disease states associated with iodine deficiencies. One advantage of this product is that it has few adverse site reactions and does not cause pain. The iodised oil has a delayed mode of action in that, over time, the oil is 20 slowly released from the depot and is taken up by the lymphatic system, making its way to the regional lymph nodes. At the lymph nodes, the oil is metabolised into constituent fatty acids and free iodine over a lengthy duration. A second long acting medicament is DeposeTM. DeposelTM is a yellow coloured free flowing aqueous suspension formulated for administration by 25 injection subcutaneously. The primary constituent of the medicament is barium selenate and the medicament provides approximately 50mg of selenium per ml of medicament. Like Flexidine

TM

, DeposeTM provides a source of selenium to a 2 - 3 treated animal for over approximately 12 months. One disadvantage of DeposelTM is that the medicament may be associated with adverse site reactions such as irritation and scarring. A further problem with the Deposel"M medicament is that creepage of the depot can occur with the depot eroding away quicker than is desirable. 5 Although both of the above iodine and selenium medicaments are widely used, one disadvantage is that these are separate treatments and therefore, in animals deficient in both elements, separate administration of each medicament is required which is costly in terms of time and labour. It would therefore be an advantage in terms of time and labour as well as ease 10 of use if there could be provided a combination medicament requiring only one administration step. A further advantage of administering both elements at once envisaged by the inventors is that the presence of selenium may increase the absorption of iodine by the animal. 15 Whilst an obvious solution may be to simply mix Flexidine TM and DeposelTM, it is the inventors experience that this is unworkable as: 1. The two medicaments do not mix well as one is oil based and the other is water based and as a result, if injected each constituent would separate and disperse such that the long acting release effect would reduce or not be present at all; 20 and, 2. Each medicament has a preferred route of administration that differs. 3. The two compositions when mixed and stored tend to separate and be difficult to re-mix as caking and clumping of the agent(s) may occur. It would therefore be desirable to have a combination medicament that may 2436143_1 (GHMatters) 12/10110 - 4 be administered for prevention and/or treatment of both selenium and iodine deficiencies and that also minimises pain and site reactions in the animal. All references, including any patents or patent applications cited in this specification are hereby incorporated by reference. No admission is made that any 5 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 10 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 15 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. 20 DISCLOSURE OF INVENTION According to one aspect of the present invention there is provided a medicament formulated for parenteral administration, wherein the medicament includes: (a) idodised unsaturated vegetable oil dissolved in its vegetable oil; (b) micronised barium selenate; and 24361431 (GHMatters) 12110110 - 5 (c) at least one additional solvent selected from mineral oil; almond oil; canola oil; corn oil; cottonseed oil; sesame oil; safflower oil; coconut oil; sunflower oil; palm oil; castor oil; isopropyl myristate; medium chain triglycerides; ethyl oleate; and/or benzoyl benzoate; and combinations thereof. 5 According to a further aspect of the present invention, there is provided use of: (a) iodised unsaturated vegetable oil dissolved in its vegetable oil; (b) micronised barium selenate; at least one solvent selected from mineral oil; almond oil; canola oil; corn oil; cottonseed oil; sesame oil; safflower oil; coconut oil; sunflower oil; palm oil; castor oil; 10 isopropyl myristate; medium chain triglycerides; ethyl oleate; and/or benzoyl benzoate; and combinations thereof; in the manufacture of a medicament for parenteral administration formulated for the prevention or treatment of a deficiency in elements including iodine, selenium, or both. According to a further aspect of the present invention, there is provided use of: 15 (a) iodised unsaturated vegetable oil dissolved in its vegetable oil; (b) micronised barium selenate; at least one solvent selected from mineral oil; almond oil; canola oil; corn oil; cottonseed oil; sesame oil; safflower oil; coconut oil; sunflower oil; palm oil; castor oil; isopropyl myristate; medium chain triglycerides; ethyl oleate; and/or benzoyl benzoate; 20 and combinations thereof; 24361431 (GHMatters) 12/10/10 - 6 in the manufacture or a medicament formulated for parenteral administration in the prevention or treatment of a disease in an animal associated with a deficiency in elements including iodine, selenium or both. According to a further aspect of the present invention there is provided a method of 5 prevention or treatment of a deficiency in iodine or selenium, or both, in an animal comprising parenteral administration, of a medicament including: (a) iodised unsaturated vegetable oil dissolved in its vegetable oil; (b) micronised barium selenate; and (c) at least one additional solvent selected from mineral oil; almond oil; canola oil; 10 corn oil; cottonseed oil; sesame oil; safflower oil; coconut oil; sunflower oil; palm oil; castor oil; isopropyl myristate; medium chain triglycerides; ethyl oleate; and /or benzoyl benzoate; and combinations thereof. 24361431 (GHMatters) 12/10/10 The inventors have found that by including a solvent, preservative and wetting agent in the medicament as well as using fine particle size barium selenate, it is possible to overcome mixing and dispersion problems associated with simply mixing iodised unsaturated vegetable oil and barium selenate. The result of combining the 5 above factors is that the barium selenate more readily disperses and re-disperses into the iodised oil and, on injection, forms a depot, gradually releasing medicament over time. In addition, the formulation ensures that the barium selenate particles do not cake or clump together. It is understood by the inventors that this improved mixing, dispersion and avoidance of caking may be due to the factors mentioned above 10 decreasing the viscosity of the iodised oil sufficient to allow dispersion of barium selenate particles within the oil but not so much that the barium selenate cakes together. The formulation therefore has the advantage of being unexpectedly storage stable as well. Preferably, the unsaturated vegetable oil is selected from: peanut oil; poppy 15 seed oil; rape seed oil; olive oil; soya oil; walnut oil; and combinations thereof. Most preferably, the iodised oil is peanut oil. Preferably, the iodised oil used in the above medicament may include approximately 26% w/v iodine bound to ethyl esters of the unsaturated fatty acids (primarily oleic and linoleic) in the oil. This is comparable to the existing Flexidine M 20 product which is known to be successful and provide long acting and adequate amounts of iodine to animal to avoid iodine deficiencies. In preferred embodiments it is envisaged that the iodised oil may constitute approximately 90% w/v of the medicament. Preferably, barium selenate may be in the form of a micronised powder 25 dispersed within the iodised oil. More preferably, the barium selenate may be a micronised powder with a particles distribution whereby 50% of the particles are 24361431 (GHMatters) 12110/10 - 8 less than 5 microns, 95% of the particles are less than 11 microns and 100% of the particles are less than 22 microns in size. It is envisaged by the inventors that micronised barium selenate may constitute approximately 8.75% w/v of the medicament. 5 Preferably, the solvent or solvents may be selected from either fixed oils or parenteral solvents or combinations thereof. For the purposes of this specification, the term 'fixed oils' refer to animal, vegetable or mineral oils that are non-volatile (stable) on heating. The term 'parenteral solvents' refers to solvents not administered orally. 10 Preferably, fixed oils may be selected from: almond oil; canola oil; corn oil; cottonseed oil; peanut oil; sesame oil; safflower oil; coconut oil; sunflower oil; palm oil; castor oil; and combinations thereof. Preferably, parenteral solvents may be selected from: isopropyl myristate; medium chain triglycerides; ethyl oleate; benzoyl benzoate; and combinations thereof. is Most preferably, the solvent is ethyl oleate although, this should not been seen as limiting as other solvents such as those described above may also have similar dispersion and/or viscosity reduction properties. In preferred embodiments envisaged by the inventors, ethyl oleate is added to make up the final volume to q.s. Preferably, the medicament may also include one or more excipients such as 20 preservatives and wetting agents. In a preferred embodiment, the inventors have found that the preservative may be benzyl alcohol and the wetting agent may be sorbitan monooleate in order to produce optimum re-suspension characteristics and to prevent caking or clumping of micronised barium selenate particles. However, other preservatives and wetting agents with similar re-suspension and anti-caking 24361431 (GHMatters) 12/10/10 - 9 properties are also envisaged as being used. In one preferred embodiment, the medicament may include: iodised peanut oil, ethyl oleate, micronised barium selenate, benzyl alcohol and sorbitan monooleate. Preferably, the medicament may be administered to an animal, for example a 5 sheep or cow. This should not be seen as limiting as the invention may also be used in many other animals. Preferably, the formulation is shaken before administration to fully suspend and distribute micronised barium selenate particles in the formulation. Preferably, the medicament is administered subcutaneously but may be 10 administered intramuscularly. In one embodiment, the medicament may be administered into the anterior half of the neck of the animal. Preferably, where the medicament is used in treatment of an element deficiency or associated disease, the medicament may be administered when the deficiency is diagnosed. 15 Preferably, where the medicament is used to prevent a deficiency, the medicament may be administered: > To a ewe before mating or at least two months prior to lambing; > To lambs as necessary, most preferably at weaning; > To rams approximately one month before mating; 20 > To feed stock at least two months before feeding goitre drops. It is envisaged by the inventors that the medicament from the preferred embodiment may provide approximately 25 mg of selenium per ml of medicament 2436143_1 (GHMatters) 12/10/10 administered. It is further envisaged that the medicament from the preferred embodiment may provide approximately 195 mg of iodine per ml of medicament administered. According to a further aspect of the present invention there is provided a 5 method of manufacture of a medicament containing iodine and selenium formulated for parenteral administration including the steps of: 1. benzyl alcohol is mixed with sorbitan monooleate; 2. the mixture of step 1 is mixed with iodised unsaturated vegetable oil; 3. the mixture of step 2 is further mixed with micronised barium selenate 10 powder; and, 4. the mixture of step 3 is further mixed with ethyl oleate. Preferably, nitrogen gas is bubbled through the mixture of step 4. Preferably, steps in the method are undertaken in aseptic conditions. It should be appreciated from the above description that there is provided a 15 medicament capable of providing a long acting source of both iodine and selenium. As the selenium source is dispersed in oil, the medicament has the advantage of being able to be administered intramuscular or subcutaneous and avoid problems associated with parenteral administration such as site reactions, irritation and egress of medicament. The medicament may be used as a prevention method to 20 avoid a deficiency or, as a treatment method once a deficiency is diagnosed. BRIEF DESCRIPTION OF DRAWINGS 10 - 11 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 of sheep serum selenium levels for Trial 2; 5 Figure 2 Shows a graph of sheep plasma iodine levels for Trial 2; and, Figure 3 Shows a graph comparing sheep urinary iodine levels for Trial 2. BEST MODES FOR CARRYING OUT THE INVENTION The invention is now described with respect to two sets of experiments. Trial 1 describes the preparation method for the composition of the present invention, and 10 administration details. Trial 2 below describes an animal trial to verify the efficacy of the formulation. TRIAL 1 The invention is now described showing both the preferred embodiment as well as mixtures that were found to not be ideal. It should be appreciated that these 15 examples are provided by way of illustration only and should not be seen as limiting. [1A] Experimental Objective The aim of the experiments was to prepare a long acting medicament for injection to deliver an adequate dose of both iodine and selenium to an animal to prevent deficiencies in these elements. 20 [1B] Initial Experiments Conducted Various trials were conducted to incorporate miconised barium selenate in iodised peanut oil. 24361431 (GHMatters) 12/10/10 - 12 Due to the inherent high viscosity of iodised peanut oil and solid nature of micronised barium selenate powder, resulting mixtures of iodised oil and micronised barium selenate exhibited a lack of uniformity in dispersion. In worst cases, micronised barium selenate powder separated out of the iodised oil. 5 One variation tested was to reduce the amount of micronised barium selenate powder from 175 mg/ml (Deposel TM equivalent) to 87.5 mg/ml of micronised barium selenate, equivalent to providing 25mg/mi selenium. [1C] Use of a Solvent During testing, one option tested was the addition of a solvent. This 10 unexpectedly improved the dispersion and uniformity. It is thought that this may be attributable to the viscosity of the iodised peanut oil being reduced by the solvent. Solvents tested included fixed oils including: almond oil, canola oil, corn oil, castor oil, cottonseed oil, peanut oil, sesame oil, safflower oil, coconut oil, sunflower oil and palm oil. Other parenteral solvents were also tested including, isopropyl myristate, 15 medium chain triglycerides, ethyl oleate and benzoyl benzoate. Solvents were found to reduce the viscosity sufficiently to allow uniform dispersion of micronised barium selenate into the iodised oil. [1D] Preferred Medicament Formulation The best medicament formulation found by the inventors that was long acting 20 included micronised barium selenate (providing 25mg/ml of selenium) with iodised oil (providing 195 mg/ml of iodine). Using the above medicament, a total dose was found to be a 2 ml dose to provide 50mg of selenium and 390mg of iodine. 24361431 (GHMatters) 12/10110 - 13 The ideal solvent used to achieve the best dispersion was found to be ethyl oleate. It is thought by the inventors that this is because of the low relative density of ethyl oleate (approximately 0.87 g/ml) compared to other solvents and compared to iodised oil (approximately 1.20 g/ml). 5 Ethyl oleate is a desirable solvent to use as it is a well accepted oleaginous vehicle in many parenteral formulations. It was also found that the addition of excipients, sorbitan monooleate (0.6 % w/v) as wetting agent, and benzyl alcohol (1% w/v) as a preservative are also advantageous. It should be appreciated that a variety of other excipients may also be 10 added without departing from the invention. In summary, a preferred medicament formulation arrived at in experiments by the inventor was as follows: Table 1: Preferred Formulation Ingredients % w/v Barium selenate (equivalent to selenium) 8.75 Benzyl alcohol 1.00 Sorbitan monooleate 0.60 lodised peanut oil 90.0 Ethyl Oleate q.s. to 100 ml (87.5mg/ml of micronised barium selenate is equivalent to approximately 25 mg/ml 15 selenium in the medicament and 900 mg/ml of iodised oil is equivalent to approximately 390 mg/mI of iodine). [1 E] Preparation Method An envisaged manufacturing procedure is described below. This should not be seen as limiting as it should be appreciated by those skilled in the art that 24361431 (GHMatters) 12110110 - 14 variations may be made without departing from the scope of the invention as described. Further, amounts and times given below are described by way of illustration only and should not be seen as limiting. The method is as follows: 5 1. Mix 3-7g of benzyl alcohol with 1-4g of sorbitan monooleate and stir well for approximately less than 15 minutes; 2. Add 440-490g of iodised peanut oil to the mixture of step 1 and stir well for approximately less than 20 minutes; 3. Add 40-50g of micronised barium selenate powder to the mixture of step 2 and stir for 10 approximately less than 50 minutes; 4. Add ethyl oleate up to the final volume (500ml) to be produced to the mixture of step 3 and stir well for approximately 25 minutes; 5. Homogenise the mixture from step 4 for approximately 15 minutes; 6. Package the formulation of step 5. 15 7. Bubble nitrogen gas through the mixture of step 5 for approximately 10 minutes while stirring. [1F] Treatment Regime A preferred animal for treatment is a sheep. It is envisaged that the medicament may be used as a treatment method when goitre 20 (or any iodine deficiency) is diagnosed. The medicament may also be used as a prevention method administered for example: either to a ewe before mating or at least two months prior to lambing; or administered to lambs as necessary, most likely at weaning. Rams may also be treated for example, by 2436143_1 (GHMatters) 12/10110 - 15 following administration approximately one month before mating and all stock treated at least two months before feeding goitre drops. [1G] Route of Administration The primary method of administration is designed to be by subcutaneous 5 injection into the anterior half of the neck. TRIAL 2 This trial was a comparative animal study to determine blood levels of selenium and iodine, and urinary iodine concentrations, following the administration of FlexidineTM, DeposelTM and the composition of the present invention in sheep. 10 Flexidine TM is a product already on the market comprising an iodised oil administered to an animal in order to provide a long acting dose of iodine. DeposelTM is a product already on the market containing micronised barium selenate particles which provide a long acting dose of selenium. The invention composition in this trial included iodised peanut oil, micronised 15 barium selenate particles and ethyl oleate as the solvent in amounts approximately the same as that of Trial 1. [2A] Trial 2 Objective This study was carried out to evaluate the properties and use characteristics of the composition of the present invention which, provides both selenium and iodine in 20 the one formulation. It was anticipated that the composition of the present invention would provide similar levels of selenium and iodine as existing products currently adminstered separately. Consequently, it was expected that the response 24361431 (GHMatters) 12/10/10 following administration (measured as a change in the serum or plasma levels of the two minerals) would be comparable to the established products, Flexidine TM (iodine) and DeposeTM (selenium). The trial aims to quantify the level and duration of the response to the 5 invention product under field conditions. [2B] Trial 2 Methodology Duration of Study The study commenced in early winter with screening tests to determine serum selenium and plasma iodine status in available sheep. 10 The trial was divided into two parts with 3 sheep tested for adverse reactions in a tolerance study (Part 1) and 47 sheep tested in the main study (Part 2) measuring selenium and iodine levels over time. The sheep ranged in age from 2-8 months at day 0 of the trial when the formulations were administered. Careful selection was made of the test site. The animals had been reared 15 on the same farm. On this farm, a marginal selenium deficiency is normally expected and in previous years selenium had been applied in fertilizer form to the pasture. Selected animals were not grazed on pastures supplemented in the current season, and were not given any selenium supplementation in drenches. Iodine deficiency was not anticipated to occur as the pasture used in the trial is not 20 in a region known to have low iodine levels. [2B(a)] Part I - Tolerance Study Initially, a small-scale evaluation of the compatibility of the new formulation was performed, using three sheep. This was a preliminary tolerance study involving three sheep that were treated as outlined below in Table 1. Flexidine T M 16 and the invention composition were injected subcutaneously at two dose rates and observations for injection site reactions were made after treatment. Table 2: Treatment group: for subcutaneous injection site study Treat Sheep Treatment Dose Rate Administration Administration No. ID. Route Site The side of the 1 1 Invention 2 mnLf neck into cranio Composition 50kg dorsal quadrant Subcutaneous in the region Invention 1 mU/ bounded 2 2 Composition 50kg centrally by the cervical vertebrae 3 3 Flexidine T M 1.5mL/ caudally by the 50kg shoulder blade Subcutaneous and dorsally by M 075mL/ the nuchal 4 2 Flexidine T M Q50kg ligament For Part 1, treatments were given to the 3 selected sheep at day -34 5 (relative to Part 2 treatments described below). Injection site and general tolerance observations were then made and recorded at 1, 4, 24, 48, and 72 hours after treatment, then 7, 10 and 14 days after treatments on Day -34. [2B(b)] Part 2: Serum and Urine Level Study Following on from Part 1 of the study, Part 2 of Trial 2 was commenced. 10 Selenium and iodine serum levels before treatment was measured in each sheep with the aim of selecting trial groups with approximately the same average levels of these minerals. Five treatment groups were selected based on pre-treatment selenium and iodine levels as summarised in Table 2 below. 15 17 Table 3: Trial Groups and Treatments Group Treatment Dose rate Dose route Selenium Iodine Dose Dose Control Group 1 - Nil Nil N/A Nil Nil Treatment 2 FlexidineTM 1.5mL/ Intramuscular Nil 9.4mg/kg 50kg 3 FlexidineTM .5m Subcutaneous Nil 9.4mg/kg Flexidine

TM

50kg _____ 4 Invention 2mU Subcutaneous 49.9mg/kg 9.4mg/kg Composition 50kg 5 Deposel T M lmL/ Subcutaneous 49.3mg/mL Nil __ __ _ __ __ __ I_ 50kg I _ _ _ __ _ _ I _ _ _ _ _ _ _ _ _ At the time of each injection, wool over the injection site was marked with a felt pen. This was completed to identify the site of injection for later observations. The trial animals were weighed using calibrated, electronic scales. Dose 5 rates for each treatment were based on the options presented in Table 3. Table 4: Dose Rates FlexidineTM Dose DeposelTM Dose Invention Composition Dose Live mL mg iodine mL mg mL mg mg weight /kg selenium/ iodine selenium/ kg /kg kg up to 10 kg 0.3 9.4 0.2 1 0.4* 9.4 1 11-20 kg 0.6 9.4 0.4 1 0.8+ 9.4 1 21-30 kg 0.9 9.4 0.6 1 1.2* 9.4 1 31-40 kg 1.2 9.4 0.8 1 1.6+ 9A 1 41-50 kg 1.5 9.4 1.0 1 2.0+ 9.4 1 51-60 kg 1,8 94 1.2 1 2.4* 9.4 1 + dose for a formulation delivering the required doses of iodine and selenium to a 50 kg ewe in 2mL. [2B(c)] Samples collected 10 Blood and urine samples were collected initially at day -34 and day -1, before administration and then at days 14, 28, 42 and 56 post administration. 18 These days are each relative to day 0, when the treatments described for Part 2 of the study were administered. On each sampling day, clotted blood samples, heparin treated blood samples and urine samples were collected. These were chilled and transported to 5 a laboratory within 24 hours for analysis. [2B(d)] Part 2 Tolerance Observations For Part 2, injection site and general tolerance were observed closely at 1, 4, 24, 48, and 72 hours after treatments. Then after each sample collection (14 28, 42, & 56 days after treatment). 10 [2C] Results [2C(a)] Tolerance Observations [2C(b)] Part 1 For the three sheep given separate treatments, no reaction was apparent, either at injection sites or on clinical observation. 15 [2C(c)] Part 2 At 1 hour post-injection, swellings of 1-2 cm were apparent in selected sheep at the injection sites in some cases. These swellings were observed in none of group 1, two of group 2, five of group 3, four of group 4, and one of the group 5 sheep. 20 At 4 hours post-injection, swellings were smaller or had disappeared in many cases. Often these swellings were dark coloured, indicating that a small, subcutaneous haematoma had formed. All of these swellings resolved without any sequelae, and most were not observable at later inspections of injection sites. It is 19 considered that they should be ascribed to vascular trauma rather than any specific property of the treatments. It is notable that the highest incidence was observed 4 hours after intramuscular injections. From observations 72 hours after treatments and later, no abnormalities 5 related to injection sites or general health were seen in any sheep. Generally the observations did not disclose any change that suggested a specific reaction to any of the injected treatments. [2C(d)] Serum selenium levels Pre-treatment means (from samples taken at days -34 and -1) were not 10 significantly different between groups on each day. Subsequent to treatments, differences emerged in mean serum selenium between treatments, and these were consistently ranked DeposeTM > Invention Composition > Control. The significance of differences varies with time of sample, and also with the 15 method of evaluation. In graphic form (see Figure 1), the bars show 95% confidence intervals and thus contain the true mean 95% of the time. If there is no overlap of a given pair of 95% confidence intervals, they can for practical purposes be considered significantly different. The confidence intervals for the controls and the invention composition do 20 not overlap at day 56, but are very close. By either method it is apparent that a significant difference in serum selenium is obtained by treatment with the invention composition (Group 4) compared to controls (Group 1), which persists until day 56. Differences between DeposelTM and the invention composition are not significant, but approach 25 significance at day 56. This suggests that the invention composition provides a 20 similar effect in terms of supplementing selenium as an existing selenium-only product on the market (Deposel

T

m. [2C(e)] Plasma iodine levels Again pre-treatment means (from samples taken at days -34 and -1) were 5 not significantly different between groups on each day. Subsequent to treatments, differences of control from other groups emerged, but generally differences in mean plasma inorganic iodine between treatments were not as large and their 95 % confidence intervals overlapped substantially. 10 The plasma profiles produced by subcutaneous administration of Flexidine TM (iodine only) and subcutaneous administration of the invention composition were very similar. The significance of differences varies with time of sample, and also with the method of evaluation. In graphic form (see Figure 2), the bars show 95% 15 confidence intervals. The confidence intervals for the controls and the invention composition do not overlap at days 28, 42 or 56. Confidence intervals for subcutaneous treatments are particularly large (indicating wide variability) at 28 days. It is apparent that a significant difference in plasma iodine is obtained 20 between Group 1 (controls) and: 1. Group 2 (Flexidine TM intramuscular) at each sample time post-treatment; 2. Group 3 (Flexidine TM subcutaneous) at 42 and 56 days post treatment; 3. Group 4 (Invention composition) also at 42 and 56 days post treatment. 21 FlexidineM intramuscular has a lower peak value than with subcutaneous administration, but appears to have a similar AUC (bioavailability) and has tighter confidence intervals. The above results show that the invention composition provides a 5 comparable level of iodine to the animal compared to existing iodine-only products as measured in blood plasma. The results also show that administration via the intramuscular route versus subcutaneous is also likely to provide comparable results at least in terms of bioavailability. [2C(f)] Urinary Iodine Levels 10 Utilising the visual information shown in Figure 3, it is apparent that the treated and untreated controls are significantly different at days 28 and 42. It is also apparent that there is likely to be no significant difference between the treated groups, as the confidence intervals of each treatment, indicated by bars, overlap substantially. 15 Thus each treatment has a significant difference in urinary iodine result from group 1 (controls), but no treatment differs significantly from another. The results therefore also show that the invention composition provides comparable results in terms of iodine urinary levels as existing iodine-only compositions. [2D] Conclusions 20 The invention composition when administered as a subcutaneous injection to sheep, improved both selenium and iodine status to a similar extent compared with the reference treatments that supplement each mineral separately. The observed improvement in selenium status was not significantly different from the result with the reference product. 22 The improvement in iodine status resulting from test product treatment appeared to be equivalent to that provided by the reference product. It should be appreciated from the above experiments that the present invention provides a medicament including both iodine and selenium for use in the 5 treatment and prevention of element deficiencies and associated disease states. The combination medicament also maintains its long acting ability, remaining stable for a duration of months within the animal. 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 10 thereto without departing from the scope thereof as defined in the appended claims. 23

Claims

1. A medicament formulated for parenteral administration, wherein the medicament includes: (a) iodised unsaturated vegetable oil dissolved in its vegetable oil; 5 (b) micronised barium selenate; and (c) at least one additional solvent selected from mineral oil; almond oil; canola oil; corn oil; cottonseed oil; sesame oil; safflower oil; coconut oil; sunflower oil; palm oil; castor oil; isopropyl myristate; medium chain triglycerides; ethyl oleate; and/or benzoyl benzoate; and combinations thereof. 10

2. The medicament as claimed in claim 1 wherein the unsaturated vegetable oil of component (a) is selected from : peanut oil; poppy seed oil; rape seed oil; olive oil; soya oil; walnut oil; and combinations thereof.

3. The medicament as claimed in claim 1 or claim 2 wherein the iodised unsaturated vegetable oil is peanut oil including approximately 26% w/v iodine bound 15 to ethyl esters of the unsaturated fatty acids in the peanut oil.

4. The medicament as claimed in any one of the above claims wherein the iodised unsaturated vegetable oil constitutes approximately 90% w/v of the medicament.

5. The medicament as claimed in any one of the above claims wherein micronised barium selenate is in the form of a micronised powder suspended within the iodised oil 20 immediately prior to administration.

6. The medicament as claimed in any one of the above claims wherein the micronised barium selenate constitutes approximately 8.75% w/v of the medicament. 24361431 (GHMatters) 12110/10 - 25

7. The medicament as claimed in any one of the above claims, which includes a preservative, benzyl alcohol.

8. The medicament as claimed in any one of the above claims, which includes a wetting agent, sorbitan monooleate. 5

9. Use of: (a) iodised unsaturated vegetable oil dissolved in its vegetable oil; (b) micronised barium selenate; at least one solvent selected from mineral oil; almond oil; canola oil; corn oil; cottonseed oil; sesame oil; safflower oil; coconut oil; sunflower oil; palm oil; castor oil; 10 isopropyl myristate; medium chain triglycerides; ethyl oleate; and/or benzoyl benzoate; and combinations thereof; in the manufacture of a medicament for parenteral administration formulated for the prevention or treatment of a deficiency in elements including iodine, selenium, or both.

10. Use of: 15 (a) iodised unsaturated vegetable oil dissolved in its vegetable oil; (b) micronised barium selenate; at least one solvent selected from mineral oil; almond oil; canola oil; corn oil; cottonseed oil; sesame oil; safflower oil; coconut oil; sunflower oil; palm oil; castor oil; isopropyl myristate; medium chain triglycerides; ethyl oleate; and/or benzoyl benzoate; 20 and combinations thereof; in the manufacture or a medicament formulated for parenteral administration in the 24361431 (GHMatters) 12/10110 - 26 prevention or treatment of a disease in an animal associated with a deficiency in elements including iodine, selenium or both.

11. A method of prevention or treatment of a deficiency in iodine or selenium, or both, in an animal comprising parenteral administration of a medicament including: 5 (a) iodised unsaturated vegetable oil dissolved in its vegetable oil; (b) micronised barium selenate; and (c) at least one additional solvent selected from mineral oil; almond oil; canola oil; corn oil; cottonseed oil; sesame oil; safflower oil; coconut oil; sunflower oil; palm oil; castor oil; isopropyl myristate; medium chain triglycerides; ethyl oleate; and/or 10 benzoyl benzoate; and combinations thereof.

12. A method for the prevention or treatment of a disease in an animal associated with a deficiency in elements including iodine, selenium or both, comprising parenteral administration of a medicament including: (a) iodised unsaturated vegetable oil dissolved in its vegetable oil; 15 (b) micronised barium selenate; and (c) at least one additional solvent selected from mineral oil; almond oil; canola oil; corn oil; cottonseed oil; sesame oil; safflower oil; coconut oil; sunflower oil; palm oil; castor oil; isopropyl myristate; medium chain triglycerides; ethyl oleate; and/or benzoyl benzoate; and combinations thereof. 20

13. A method as claimed in claim 11 or claim 12 wherein the unsaturated vegetable oil of component (a) is selected from : peanut oil; poppy seed oil; rape seed oil; olive oil; soya oil; walnut oil; and combinations thereof. 2436143_1 (GHMatters) 12/10/10 - 27

14. A method as claimed in any one of claims 11 to 13 wherein the iodised unsaturated vegetable oil is peanut oil including approximately 26% w/v iodine bound to ethyl esters of the unsaturated fatty acids in the peanut oil.

15. A method as claimed in any one of claims 11 to 14 wherein the iodised 5 unsaturated vegetable oil constitutes approximately 90% w/v of the medicament.

16. A method as claimed in any one of claims 11 to 15 wherein micronised barium selenate is in the form of a micronised powder suspended within the iodised oil immediately prior to administration.

17. A method as claimed in any one of claims 11 to 16 wherein the micronised 10 barium selenate constitutes approximately 8.75% w/v of the medicament.

18. A method as claimed in any one of claims 11 to 17, which includes a preservative, benzyl alcohol.

19. A method as claimed in any one of claims 11 to 18, which includes a wetting agent, sorbitan monooleate. 24361431 (GHMatters) 12/10110