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EP4362945A1 - Additifs alimentaires pour poisson - Google Patents

Additifs alimentaires pour poisson

Info

Publication number
EP4362945A1
EP4362945A1 EP22831080.1A EP22831080A EP4362945A1 EP 4362945 A1 EP4362945 A1 EP 4362945A1 EP 22831080 A EP22831080 A EP 22831080A EP 4362945 A1 EP4362945 A1 EP 4362945A1
Authority
EP
European Patent Office
Prior art keywords
fish
composition
bead
beads
taste
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Pending
Application number
EP22831080.1A
Other languages
German (de)
English (en)
Other versions
EP4362945A4 (fr
Inventor
Edith Kai Yan TANG
Gavin John PARTRIDGE
Lee Yong Lim
Lindsey Dawn WOOLLEY
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
University of Western Australia
Western Australian Agriculture Authority
Original Assignee
University of Western Australia
Western Australian Agriculture Authority
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Priority claimed from AU2021902007A external-priority patent/AU2021902007A0/en
Application filed by University of Western Australia, Western Australian Agriculture Authority filed Critical University of Western Australia
Publication of EP4362945A1 publication Critical patent/EP4362945A1/fr
Publication of EP4362945A4 publication Critical patent/EP4362945A4/fr
Pending legal-status Critical Current

Links

Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/14—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
    • A61K9/16—Agglomerates; Granulates; Microbeadlets ; Microspheres; Pellets; Solid products obtained by spray drying, spray freeze drying, spray congealing,(multiple) emulsion solvent evaporation or extraction
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P33/00—Antiparasitic agents
    • A61P33/10—Anthelmintics
    • A61P33/12—Schistosomicides
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23K—FODDER
    • A23K10/00—Animal feeding-stuffs
    • A23K10/30—Animal feeding-stuffs from material of plant origin, e.g. roots, seeds or hay; from material of fungal origin, e.g. mushrooms
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23K—FODDER
    • A23K20/00—Accessory food factors for animal feeding-stuffs
    • A23K20/10—Organic substances
    • A23K20/111—Aromatic compounds
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23K—FODDER
    • A23K20/00—Accessory food factors for animal feeding-stuffs
    • A23K20/10—Organic substances
    • A23K20/116—Heterocyclic compounds
    • A23K20/132—Heterocyclic compounds containing only one nitrogen as hetero atom
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23K—FODDER
    • A23K20/00—Accessory food factors for animal feeding-stuffs
    • A23K20/10—Organic substances
    • A23K20/158—Fatty acids; Fats; Products containing oils or fats
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23K—FODDER
    • A23K20/00—Accessory food factors for animal feeding-stuffs
    • A23K20/10—Organic substances
    • A23K20/163—Sugars; Polysaccharides
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23K—FODDER
    • A23K40/00—Shaping or working-up of animal feeding-stuffs
    • A23K40/10—Shaping or working-up of animal feeding-stuffs by agglomeration; by granulation, e.g. making powders
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23K—FODDER
    • A23K40/00—Shaping or working-up of animal feeding-stuffs
    • A23K40/30—Shaping or working-up of animal feeding-stuffs by encapsulating; by coating
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23K—FODDER
    • A23K50/00—Feeding-stuffs specially adapted for particular animals
    • A23K50/80—Feeding-stuffs specially adapted for particular animals for aquatic animals, e.g. fish, crustaceans or molluscs
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/498—Pyrazines or piperazines ortho- and peri-condensed with carbocyclic ring systems, e.g. quinoxaline, phenazine
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/4985—Pyrazines or piperazines ortho- or peri-condensed with heterocyclic ring systems
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/0012—Galenical forms characterised by the site of application
    • A61K9/0053—Mouth and digestive tract, i.e. intraoral and peroral administration
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/14—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
    • A61K9/16—Agglomerates; Granulates; Microbeadlets ; Microspheres; Pellets; Solid products obtained by spray drying, spray freeze drying, spray congealing,(multiple) emulsion solvent evaporation or extraction
    • A61K9/1605—Excipients; Inactive ingredients
    • A61K9/1629—Organic macromolecular compounds
    • A61K9/1652—Polysaccharides, e.g. alginate, cellulose derivatives; Cyclodextrin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/14—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
    • A61K9/16—Agglomerates; Granulates; Microbeadlets ; Microspheres; Pellets; Solid products obtained by spray drying, spray freeze drying, spray congealing,(multiple) emulsion solvent evaporation or extraction
    • A61K9/1605—Excipients; Inactive ingredients
    • A61K9/1664—Compounds of unknown constitution, e.g. material from plants or animals
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
    • Y02A40/00—Adaptation technologies in agriculture, forestry, livestock or agroalimentary production
    • Y02A40/80—Adaptation technologies in agriculture, forestry, livestock or agroalimentary production in fisheries management
    • Y02A40/81—Aquaculture, e.g. of fish

Definitions

  • the present invention relates to the field of oral delivery of therapeutic compounds to fish through fish feed additives.
  • praziquantel 2-(cyclohexylcarbonyl)- 1 ,2, 3, 6, 7,11 b-hexahydro-4H-pyrazino[2,1-a]isoquinolin-4-one
  • praziquantel is used to treat S. lalandi suffering from polyopisthocotylean and monopisthocotylean monogenean flukes.
  • praziquantel due to the poor palatability of praziquantel, as it has a strong bitter taste, fish are bathed in praziquantel, rather than receiving the drug orally.
  • fish are bathed in hydrogen peroxide.
  • Bathing can be labour intensive, time consuming, weather dependant and prolonged exposure can detrimentally impact the fish, causing reduced growth rates and in some cases mortalities (Gaikowski, M. P. et. a/., 1999, Acute toxicity of hydrogen peroxide treatments to selected life stages of cold, cool, and warm water fish Aquaculture, 178, 191-207). Bathing treatments can also be expensive. For example, in Japan, the cost of bathing to treat B. seriolae contributes up to 22% of the production costs of sea caged Seriola species (Ernst, I. et. at., 2002, Monogenean parasites in sea-cage aquaculture Austasia Aquaculture, 16, 46-48).
  • Oral administration of medication to fish on the other hand has advantages over conventional bath treatments. For example, in-feed medications have wider safety margins and do not require crowding or increased handling of fish. Treatment efficiency is also increased, as all cages on the farm can be treated quickly, (Williams, R. et. a!., 2007, Efficacy of orally administered praziquantel against Zeuxapta seriolae and Benedenia seriolae (Monogenea) in yellowtail kingfish Seriola lalandi, Diseases of Aquatic Organisms, 77, 199-205.) reducing the chance of infection from nearby untreated fish. Through feeding medicated diets, the fish can also maintain their natural feeding regime, also reducing stress (Conte, F.
  • W02009/023013 describes multiparticulate in- feed additives that have a number of coatings applied to a core containing the medication (praziquantel and cimetidine).
  • a core containing the medication praziquantel and cimetidine
  • it describes a plurality of granular cores comprising a hydrogel-forming polymer, a second coating disposed on the first coating comprising a barrier material and a third coating disposed on the second coating comprising a taste-masking polymer being a Eudraguard ® polymer.
  • This is however a complex structure from a manufacturing perspective, as the beads comprise several distinct coatings.
  • W01989/12442 describes a pharmaceutical dosage form for administration of medicaments to fish comprising an outer layer of animal or vegetable material, which is substantially impermeable to water, which surrounds an internal chamber which is filled with the active agent.
  • AU2008100441 describes medicated fish feed containing therapeutic quantities of microencapsulated anthelmintic drugs (including praziquantel), as well as attractants such as amino acids, nucleotides and natural and synthetic extracts from various aquatic organisms, which trigger the olfactory and taste receptors of fish, eliciting feeding behaviour in the target fish species. Neither of these methods appear to have been used commercially. [0010] Partridge et. al. (2014) [Partridge, G.J., Michael, R.J.
  • the invention provides an aquatic animal feed additive composition capable of delivering a therapeutically effective amount of a therapeutic to an aquatic animal, said additive comprising a plurality of therapeutic beads, wherein each bead comprises:
  • the carrier matrix is digestible by the aquatic animal.
  • the therapeutically effective compound is substantially stable in the carrier matrix when the amount of leaching of the therapeutically effective compound from the carrier matrix is less than 20% w/w over a 60 minute period in an aqueous environment, at room temperature, wherein the aqueous environment is an environment in which the aquatic animal feed additive is used (such as water or more preferably such as sea water).
  • the additive is generally durable in the aqueous environment in which the aquatic animal lives, that is to be treated.
  • the carrier matrix is generally durable if it breaks down at room temperature by less than 1 , 1.1 , 1.2, 1.3, 1.4, 1 .5, 1 .6, 1 .7, 1 .8, 1 .9, 2, 2.5, 3, 3.5, 4, 4.5, 5, 5.5, 6, 6.5, 7, 7.5, 8, 8.5, 9, 9.5 or 10% w/w.
  • the additive preferably disintegrates, at least in part, in the digestive tract of the aquatic animal that is treated by the feed additive.
  • the additive breaks down by at least 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95% or 99% w/w in the digestive tract of the aquatic animal.
  • the additive may also include a wetting agent or surfactants polymer or pharmaceutical disintegrant to assist in disintegration of the additive.
  • Wetting agents or surfactants may include, for example and without limitation, anionic or cationic detergents or a non-ionic wetting agent.
  • the wetting agent is Cremophor ® RH40 or Tween.
  • Disintegrants can also be starches and modified starches, cellulose etc.
  • the additive or carrier matrix can include a wetting agent or surfactants polymer or pharmaceutical disintegrant, that may or may not breakdown in an animal’s digestive tract, which assists in the manufacture of the additive and or enhances the wetting of the additive in an aqueous environment.
  • Wetting agents or surfactants suitable for this purpose include, for example and without limitation, anionic or cationic detergents or a non-ionic wetting agent. Disintegrants can also be starches and modified starches, cellulose etc.
  • the wetting agent is Cremophor ® RH40 or Tween.
  • the therapeutic is substantially stable in the additive for at least 6 months, more preferably 9, 12, 15, 18, 24 months.
  • the carrier matrix is at least partially prepared from a blend of alginate and an acid soluble polymer (such as chitosan and/or Eudraguard ® ) or alginate; or agar.
  • an acid soluble polymer such as chitosan and/or Eudraguard ®
  • alginate or agar.
  • taste-masking agent is a garlic powder or a garlic derivative.
  • the therapeutic is an anthelmintic drug, such as praziquantel, mebendazole and fenbendazole.
  • Praziquantel is widely used to treat trematode, cestode and monogenean infestations in both freshwater and marine fish and elasmobranchs and therefore provides a model therapeutic.
  • the aquatic animal feed additive will include an appropriate amount of a therapeutic agent to achieve therapeutic efficacy. That amount of therapeutic agent will depend on, inter alia, the nature of the pathogen, the size of the aquatic animal and temperature of the water (amongst other matters). Where praziquantel is administered for blood feeding gill flukes the dose rate is approximately 50mg/kg. Where praziquantel is administered for mucus feeding gill flukes the dose rate is approximately 150mg/kg. The dietary inclusion level required to achieve these doses is something that a person skilled in the art can easily ascertain depending on water temperature (fish eat less in cold water) and the size of the fish.
  • the effective dietary inclusion level having regard to the average of all fish sizes and all water temperatures, could be achieved with a dietary inclusion level of between 10 g PZQ/kg to 20 g PZQ/kg of food. In many respects, the higher the dietary inclusion level the better.
  • the invention provides that the therapeutically effective compound is present in an amount of at least about 30% to 70% w/w of the beads.
  • the beads of the invention are about 0.1 to 5 millimetres in diameter.
  • the composition comprises: (a) the therapeutically effective compound comprises praziquantel which is present in an amount of about 70-90% w/w of each bead; (b) the taste masking agent comprises a garlic derivative which is present in an amount of about 5% w/w of each bead; and (c) the carrier matrix comprises a combination of an acid soluble polymer and alginate, wherein the acid soluble polymer is present in an amount of 8-10% w/w and the alginate is present in an amount of 8-11% w/w of each bead.
  • the composition comprises: (a) the therapeutically effective compound comprises praziquantel which is present in an amount of about 70-90% w/w of each bead; (b) the taste masking agent comprises a garlic derivative which is present in an amount of about 5% w/w of each bead; (c) the carrier matrix comprises alginate, wherein the alginate is present in an amount of 5-11 % w/w of each bead; and (d) the wetting agent is Cremophor ® RH40, wherein the Cremophor ® RH40 is present in an amount of 5-10% w/w of each bead.
  • the composition comprises: (a) the therapeutically effective compound comprises praziquantel which is present in an amount of about 70-90% w/w of each bead; (b) the taste masking agent comprises a garlic derivative which is present in an amount of about 2-5 % w/w of each bead; and (c) the carrier matrix comprises a combination of agar, which is present in an amount of 18-20% w/w of each bead.
  • the composition comprises: (a) the therapeutically effective compound comprises praziquantel which is present in an amount of about 70-90% w/w of each bead; (b) the taste masking agent comprises a garlic derivative which is present in an amount of about 5% w/w of each bead; (c) the carrier matrix comprises a combination of an acid soluble polymer and alginate, wherein the acid soluble polymer is present in an amount of 8-10% w/w and the alginate is present in an amount of 8-11% w/w of each bead; and (d) the wetting agent is Cremophor ® RH40 or Tween, wherein the RH40 or Tween is present in an amount of 5-10% w/w of each bead.
  • the beads are further covered with a taste-masking agent.
  • the invention provides a fish feed composition comprising the fish feed additive of the invention.
  • the invention provides a method of treating or preventing a disease in a fish comprising administering to the fish the composition, fish feed or fish feed additive of the invention.
  • the disease is a parasitic infection, such as an infection caused by cestodes or trematodes, such as polyopisthocotylean and monopisthocotylean monogenean flukes.
  • the fish feed or fish additive is administered in a sufficient quantity to deliver to the fish a dose of the therapeutically effective compound in an amount of 50 mg/kg to 150 mg/kg.
  • the invention provides a method for preparing the aquatic animal feed additive composition of the invention comprising the steps of: (a) preparing a dry mixture of the therapeutically effective compound and the taste-masking agent;
  • the dry mixture is dry because no water is added.
  • the dry mixture is substantially or completely free of water.
  • the invention provides a method for preparing the fish feed additive composition of the invention comprising the steps of: (a) mixing the aquatic animal feed additive with crushed fish feed to form a mixture; and (b) forming pellets from the mixture in (a).
  • Figure 1 A depicts the level of disintegration of beads comprising formulations A to E in seawater after 5 hours.
  • Figure 1 B depicts the level of disintegration of beads comprising formulations A to E in 0.1 M HCI after 5 hours.
  • Figure 2A depicts the cumulative percent dissolution of praziquantel (PZQ) from pure drug powder, Formulation B, and Formulation C after 180 min incubation in seawater.
  • Figure 3A depicts DSC thermograms of Formulation B in comparison with the DSC thermograms of PZQ powder and the corresponding blank beads.
  • Figure 3B depicts DSC thermograms of Formulation C in comparison with the DSC thermograms of PZQ powder and the corresponding blank beads.
  • Figure 4A depicts the percentage of the total ration offered of fish feed comprising formulation A or E which was consumed by 175 gram (small) and 2000 gram (large) kingfish.
  • Figure 4B depicts the time taken to consume the total ration offered of fish feed comprising formulation A or E which was consumed by 175 gram (small) and 2000 gram (large) kingfish.
  • Figure 5A depicts the percentage of the total ration offered of fish feed comprising formulations B, C or D which was consumed by 260 gram kingfish.
  • Figure 5B depicts the time taken to consume the total ration offered of fish feed comprising formulations B, C or D which was consumed by 260 gram kingfish.
  • Figure 6 depicts the dissected digestive tract showing undigested beads in the a) midgut and b) hindgut of a large kingfish fed fish feed comprising formulation E.
  • Figure 7 depicts the dissected digestive tract of fish 3 hours post-feeding, for fish offered fish feed comprising formulations B, C or D.
  • Figure 8A depicts the percentage of the total ration of 4 month old fish feed containing formulations B and C compared with fresh fish feed which was consumed by the fish.
  • Figure 8B depicts the average time taken to consume the total ration of 4 month old fish feed containing pure praziquantel, formulation B, or formulation C compared with fresh fish feed.
  • Figure 9A depicts the percentage of the total ration offered of fish feed comprising pure praziquantel, formulation B, or formulation C consumed by 1600 gram kingfish.
  • Figure 9B depicts the percentage of the total ration offered of fish feed comprising praziquantel, formulation B, or formulation C consumed by 1600 gram kingfish over the 6 day experimental period.
  • Figure 9C depicts the average daily dose of praziquantel received by the fish that were offered fish feed comprising pure praziquantel, or formulation B, or formulation C.
  • Figure 10 depicts the percentage reduction of Zeuxapta fluke in fish that were offered fish feed comprising pure praziquantel, formulation B, or formulation C.
  • the present invention is based on the discovery that encapsulating a taste-masking agent together with a therapeutically effective compound in a carrier matrix that is ingestible for fish and which does not substantially dissolve or degrade in freshwater or seawater, can improve the palatability of therapeutically effective compounds that are otherwise difficult to administer to fish orally.
  • the invention described herein may include one or more range of values (e.g. size, percentage, concentration, etc.).
  • a range of values will be understood to include all values within the range, including the values defining the range, and values adjacent to the range which lead to the same or substantially the same outcome as the values immediately adjacent to that value which defines the boundary to the range. Accordingly, unless indicated to the contrary, the numerical parameters set forth in the specification and claims are approximations that may vary depending upon the desired properties sought to be obtained by the present invention. Hence “about 80%” means “about 80%” and also “80%”. At the very least, each numerical parameter should be construed in light of the number of significant digits and ordinary rounding approaches.
  • fish feed means any material such as plant, animal or other organic material that is intended for consumption by fish.
  • Fish feed is used to provide nutrients for captive or farmed fish, and usually contains macronutrients, trace elements as well as vitamins.
  • Sources of nutrients can be fishmeal and other marine sources, vegetable proteins as well as binding agents such as wheat.
  • Fish feed is commonly provided in solid form, such as granules, pellets or flakes.
  • aquatic animal includes fish.
  • aquatic animal feed additive means a composition that can be added to aquatic animal feed, preferably fish feed.
  • Aquatic animal feed additives can contain medications, or other biologically effective agents, taste-masking agents, including flavouring agents, carriers and other components that are desired to be ingested by an aquatic animal population.
  • Aquatic animal feed additives can be incorporated within aquatic animal feed granules or pellets during the manufacturing of the aquatic animal feed. Alternatively, they can be mixed with manufactured aquatic animal feed granules or pellets prior to feeding, or otherwise coated on the surface of aquatic animal feed granules or pellets.
  • Beads includes particles, granules and pellets. Beads can be any shape, or size, including spherical or flattened.
  • therapeutically effective amount refers to the amount or dose of the therapeutically effective compound determined to produce a therapeutic response in an aquatic animal. Such therapeutically effective amounts or doses are readily ascertained by one of ordinary skill in the art.
  • treat and “treatment” includes therapeutic treatments, prophylactic treatments, and applications in which one reduces the risk that a subject will develop a disorder or other risk factor. Treatment does not require the complete curing of a disorder and encompasses embodiments in which one reduces symptoms or underlying risk factors.
  • palatable means that the taste and/or odour of a food or additive is sufficiently acceptable to the subject such that it is consumed by the subject.
  • a food or additive does not need to be completely consumed in a given period (i.e. consumed 100%) in order to be considered palatable to the subject.
  • a feed or additive containing a therapeutically effective compound is considered “palatable” if the subject finds it acceptable to consume it in a sufficient quantity to have a therapeutic effect.
  • a feed or additive containing a therapeutically effective compound is considered “palatable” if the presence of the therapeutically effective compound does not significantly affect the amount of feed consumed by an aquatic animal population. This can be measured by comparing the amount of medicated aquatic animal feed or aquatic animal feed additive consumed by the aquatic animal population to the amount of non-medicated aquatic animal feed or aquatic animal feed additive consumed by the aquatic animal population.
  • the invention provides an aquatic animal feed additive capable of delivering a therapeutically effective amount of a therapeutic to an aquatic animal, said additive comprising a plurality of therapeutic beads, wherein each bead comprises:
  • (b) is substantially stable for at least 60 minutes at room temperature in an aqueous environment in which the aquatic animal feed additive is used; and wherein, the taste-masking agent alone or in combination with the carrier matrix masks the taste of the therapeutically effective compound.
  • the carrier matrix is digestible by the aquatic animal.
  • the present invention provides a particulate aquatic animal feed additive.
  • the beads of the invention can be of any shape or size that can be ingested by the target aquatic animal population, and which can be incorporated into aquatic animal feed.
  • the beads are about 0.1 millimetres to 5 millimetres in diameter.
  • the beads are 0.1 , 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1 , 1.2, 1 .3, 1 .4, 1 .5, 1 .6, 1 .7, 1 .8, 1 .9, 2.0, 2.1 , 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1 , 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, 4.0, 4.1 , 4.2, 4.3, 4.4, 4.5, 4.6, 4.7, 4.8, 4.9 or 5.0 millimetres in diameter.
  • the beads have a diameter selected from the group consisting of: 5000 pm, ⁇ 5000 pm, 1000 pm, ⁇ 1000 pm; 500 pm; ⁇ 500 pm; 355-500pm; 212-355 pm; 150-355 pm; 150 pm; ⁇ 150 pm; 100 pm and ⁇ 100 pm.
  • Each bead comprises a therapeutically effective compound, a taste-masking agent and a carrier matrix.
  • the relative amounts of each of these elements in a bead depends on the nature of the therapeutically effective compound, the taste-masking agent and the carrier matrix.
  • the amount of therapeutically effective compound present in each bead will be at least 45% w/w, although a skilled reader will understand that the final amount will depend on the therapeutic being delivered by each bead and the number of beads in the additive.
  • the amount of therapeutically effective compound present in each bead will be at least
  • the taste-masking agent is between 1% and 10% w/w such as 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10 percent w/w.
  • the taste-masking agent is between 1% and 10% w/w such as 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10 percent w/w.
  • each taste-masking agents may be present in an amount of between 1% and 10% w/w such as 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10 percent w/w.
  • the carrier matrix is between 2% and 35% w/w such as 1 , 2, 3, 4, 5, 6, 7, 8,
  • each of the therapeutically effective compound, taste-masking agent and carrier matrix are preferably distributed throughout the beads of the invention, rather than coated in separate layers.
  • the percent w/w of each of the therapeutically effective compound, taste- masking agent and carrier matrix will amount to between 80 and 100 percent w/w such as 80, 81 , 82, 83, 84, 85, 86, 87, 88, 89, 90, 91 , 92, 93, 94, 95, 96, 97, 98, 99, 100 percent w/w.
  • the therapeutically effective compound is substantially stable in the carrier matrix when the amount of leaching of the therapeutically effective compound from the carrier matrix is less than 20% w/w over a 60 minute period in an aqueous environment, at room temperature, wherein the aqueous environment is an environment in which the aquatic animal feed additive is used (such as water or more preferably such as sea water).
  • the amount of leaching is less than 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2% or 1% w/w over a 60 minute period in the aqueous environment at room temperature. More preferably the amount of leaching is less than 5% w/w over a 60 minute period in an aqueous environment at room temperature.
  • Chemical stability of an additive preparation depends upon the amount of therapeutic leaching of the active ingredient in that preparation. Commonly, stability analysis of a preparation may be performed under accelerated temperature conditions, such as in an oven at temperatures higher than room temperature. The kinetic methods used in the accelerated stability analysis need not involve detailed studies of mechanism of degradation to be able to predict stability, but they are preferably based upon sound scientific principles and compliance with regulatory requirements.
  • the additive is generally durable in the aqueous environment in which the aquatic animal lives, that is to be treated.
  • the carrier matrix is generally durable if it breaks down at room temperature by less than 0.01 , 0.02, 0.03, 0.04, 0.05, 0.06, 0.07, 0.08, 0.09, 0.1 , 0.11 , 0.12, 0.13, 0.14, 0.15, 0.16, 0.17, 0.18, 0.19, 0.2, 0.21 , 0.22, 0.23, 0.24, 0.25, 0.26, 0.27, 0.28, 0.29, 0.3, 0.31 , 0.32, 0.33, 0.34, 0.35, 0.36, 0.37, 0.38, 0.39, 0.4, 0.41 , 0.42, 0.43, 0.44, 0.45, 0.46, 0.47, 0.48, 0.49, 0.5, 0.51 , 0.52, 0.53, 0.54, 0.55, 0.56, 0.57, 0.58, 0.59, 0.6, 0.61 , 0.62, 0.63, 0.64
  • the additive preferably disintegrates, at least in part, in the digestive tract of the aquatic animal that is treated by the feed additive.
  • the additive breaks down by at least 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95% or 99% w/w in the digestive tract of the aquatic animal.
  • the additive may also include a wetting agent or surfactants polymer or pharmaceutical disintegrant to assist in disintegration of the additive.
  • Wetting agents or surfactants may include, for example and without limitation, anionic or cationic detergents or a non-ionic wetting agent.
  • the wetting agent is Cremophor ® RH40 or Tween.
  • Disintegrants can also be starches and modified starches, cellulose etc.
  • the additive or carrier matrix can include a wetting agent or surfactants polymer or pharmaceutical disintegrant, that may or may not breakdown in an animal’s digestive tract, which assists in the manufacture of the additive and or enhances the wetting of the additive in an aqueous environment.
  • a wetting agent can be added to help with the wetting of the drug powder so that it can be uniformly dispersed in the matrix.
  • the wetting agent also assists in displacing entrapped air between the drug and the carrier matrix to give a stronger bind between the carrier matrix and the therapeutic- entrapped air gives rise to friable beads that are then easily crushed when incorporated into the fish feed, resulting in the fish being able to taste the exposed therapeutic.
  • wetting agents or surfactants suitable for this purpose include, for example and without limitation, anionic or cationic detergents or a non-ionic wetting agent. Disintegrants can also be starches and modified starches, cellulose etc. In a particular form of the invention, the wetting agent is Cremophor ® RH40 or Tween.
  • the therapeutic is substantially stable in the additive for at least 6 months, more preferably 9, 12, 15, 18, 24 months.
  • the beads may include other components or additional materials such as additional taste-masking agents may be deposited on (including as a coating or covering) or included in the beads.
  • the beads may be dried. Any suitable method can be used for drying the beads.
  • the bead may be air-dried using a fume-hood over a period of several days, as described below. The dried beads may then be incorporated into an aquatic animal feed.
  • any therapeutically effective compound that can be administered orally to aquatic animals, preferably fish, may be used in the present invention.
  • the therapeutically effective compound can be an anthelmintic drug, a probiotic, a synbiotic or an antibiotic or a combination thereof.
  • the therapeutically effective compound is an anthelmintic drug.
  • Anthelmintic drugs that are used in the treatment and prevention of parasitic diseases in aquatic animals such as fish include trichlorfon, mebendazole, fenbendazole, praziquantel and 40% phoxim.
  • the anthelmintic drugs is selected from florfenicol, oxytetracycline dehydrate, sulfadimethoxine/ormetoprim, bioinsecticides based on B. thuringiensis and spores of the Gram-positive bacteria of the genus Bacillus.
  • the antibiotic is selected from a tetracycline, oxolinic acid and chloramphenicol.
  • the probiotic main purpose will be to establish or to maintain a relationship between beneficial and harmful bacteria, which is usually present in the intestine or gut of fish.
  • Effective probiotics should possess certain qualities as specified below: a. The probiotics should have a beneficial effect on the growth, development and protection of fish against various pathogenic bacteria. b. The probiotic bacteria should not have any harmful effect on the host. c. The probiotics should not have the ability of drug resistance, and should have the ability to keep up hereditary traits. d. For the utilization of probiotics in an efficient feed, they should exhibit the following properties: i. acid and bile tolerance ii. resistance to gastric juices iii.
  • probiotic microorganisms belong to Bacillus, Lactobacillus and Bifidobacterium genus.
  • Various species of Lactobacillus, Bifidobacterium and Streptococcus reported for use in aquaculture as probiotics include L. acidophilus, L. casei, L fermentum, L. gasseri, L. plantarum, L. salivarius, L. rhamnosus, L. johnsonii, L. paracasei, L. reuteri, L helveticus, L.
  • Lactococcus lactis RQ516 probiotic shows inhibitory action against Aeromonas hydrophila when given to Tilapia (Oreochromis niloticus).
  • L. lactis probiotic has anti-bacterial activity against two pathogens - Yersinia rukeri and Aeromonas salmonicida that can affect fish growth.
  • Leuconostoc mesenteroides has the potential to inhibit the fish pathogens found in Nile tilapia ( O . niloticus).
  • Lactic acid bacteria such as Lactobacillus acidophilus, Lactobacillus buchneri, Lactobacillus fermentum, Lactococcus lactis, and Sterptococcus salivarius were isolated from Spanish mackerel ⁇ Scomberomorus commerson) intestine and were capable of inhibiting the Listeria innocua growth.
  • the inhibition of viruses can also occur by secretion of extracellular enzymes produced by the bacteria.
  • Aeromonas species, Corynebacterium, Pseudomonas and Vibrio species show the antiviral activity against the IHNV (Infectious hematopoietic necrosis virus).
  • Feeding of probiotic strain Bacillus megaterium has increased the resistance against WSSV (white-spot syndrome virus) in the shrimp, Litopenaeus vannamei.
  • the probiotics strains Bacillus and Vibrio species are effective against WSSV and efficiently protect Litopenaeus vannamei.
  • Application of Lactobacillus as a probiotic, either as a single strain or as a mixture with Sporolac resulted in better resistance against lymphocystis viral disease, which is found in Paralichthys olivaceus der).
  • Probiotics can also exhibit antifungal activity.
  • the Aeromonas strain A199 from Anguilla australis (eel) culture water can inhibit the Saprolegnia species.
  • Pseudomonas species M162, Pseudomonas species M174 and Janthinobacterium species M169 can increase the animal’s immunity against saprolegniasis, as has been demonstrated with Oncorhynchus mykiss (rainbow trout).
  • Lactobacillus plantarum FNCC 226 has also shown inhibitory potential in catfish (Pangasius hypophthalamus) against Saprolegnia parasitica.
  • a major advantage of the present invention is that the aquatic animal feed additive provides a means for improving the palatability of therapeutically effective compounds. Therefore, in some embodiments of the invention, the therapeutically effective compound is a poorly palatable or pungent compound that is difficult to directly administer orally to aquatic animals.
  • the therapeutically effective compound is selected from the group consisting of: a probiotic; a symbiotic; an anti-invertebrate compound; an anti-helminitic compound; an anti-viral compound; an antinematode compound; an antibiotic compound; an algaecide; an insecticide compound; an antifungal compound; an antiprotozoal compound; and a larvicide compound.
  • the therapeutically effective compound is selected from the group consisting of: a tetracycline; a sulfa-antibiotic; a diaminopyrimidine; a fluoroquinolone; a quinolone; a sulphonamide; a avermectin; a macrolide; a chlorinated bisphenol; a benzoylurea; a monochlorobenzene; an insect growth regulator; a sulfur compound; a salicylamide; a phosphorus compound; a benzimidazole; a pyrethrin; a triazine; a tetramisole; and an anticoccidal; a irreversible organophosphate acetylcholinesterase inhibitor; a organophosphate insecticide; a chloramphenicol; and a tetracycline;
  • the therapeutically effective compound is selected from the group consisting of: praziquantel; sulfadiazine; trimethoprim; flumequine; oxytetracycline; oxolinic acid (also known as terramycin); emamectin benzoate; trichlorfon; mebendazole; fenbendazole; 40% phoxim; florfenicol; oxytetracycline dehydrate; sulfadimethoxine/ormetoprim; sulfadimethoxine; ormetoprim; bioinsecticides based on B. thuringiensis and spores of the Gram positive bacteria of the genus Bacillus; tetracycline; and chloramphenicol.
  • the therapeutically effective compound is selected from the group consisting of: macrolide anthelmintic; bithionol; diflubenzuron; triflumuron; pyriproxyfen; sulfur powder; salicylanilide; organic phosphorus; benzimidazole; pyrethroid; triazine; tetramisole anthelmintic; levamisole anthelmintic; sulfonamide; and an anticoccidial drug.
  • the therapeutically effective compound is praziquantel.
  • Praziquantel is an anthelmintic drug which is known to have a particularly bitter taste that is rejected by fish when administered directly orally to fish, simply mixed in with fish feed, or coated on fish feed.
  • Praziquantel is used frequently in the treatment of flukes among certain commercially cultured fish species, such as the yellowtail kingfish, but is generally administered through bathing due to its bitter taste.
  • the yellowtail kingfish is known to have a particularly sensitive palate.
  • the amount of the therapeutically effective compound in an aquatic animal feed additive bead of the invention is dependent on the nature of the therapeutically effective compound, and the amount required for achieving a therapeutic effect in the aquatic animal population.
  • the amount of aquatic animal feed additive (or aquatic animal feed comprising the aquatic animal feed additive) consumed by the aquatic animal population influences the amount of therapeutically effective compound required in the beads.
  • the size of the aquatic animal, and the temperature of the water can also influence the amount of aquatic animal feed consumed by the aquatic animal population. For example, larger fish tend to eat a smaller percentage of their bodyweight in fish feed compared with smaller fish. Fish populations also tend to consume a smaller amount of feed in colder water conditions. This means that both for larger fish and in colder water conditions, a greater amount of the therapeutically effective compound must be present in the fish feed in order for it to be consumed in a therapeutically effective dose by the fish population.
  • a particular advantage of some embodiments of this invention is that very high drug loading can be achieved in the beads.
  • beads contain at least about 70% w/w of the therapeutically effective compound. This can be useful in delivering some therapeutically effective compounds in certain conditions and to some aquatic animal populations, as high drug loading is necessary in order to deliver the therapeutically effective compound in a sufficient quantity to have a therapeutic effect in the aquatic animal.
  • the therapeutically effective dose of praziquantel is between 50 mg/kg and 150 mg/kg.
  • drug loading of about 70% to 90% w/w praziquantel is present in the beads, which ensures that the aquatic animal population ingests enough aquatic animal feed such that a therapeutically effective dosage of praziquantel is delivered to the aquatic animal population.
  • the present invention provides for a taste-masking agent that is incorporated within the beads of the invention.
  • the encapsulation of the taste-masking agent within the beads of the invention provides the advantage of ensuring that the taste-masking agent remains proximal to the therapeutically effective compound and is not dispersed independently of the therapeutically effective compound. It is therefore better able to mask the taste of the therapeutically effective compound.
  • the taste-masking agent can be any agent that has the capacity to sufficiently conceal the flavour and/or smell of the therapeutically effective compound that is also comprised within the beads.
  • Taste-masking agents can exert this effect in numerous ways. Some taste-masking agents do not have a taste or flavour of their own. Instead, these agents can influence the perception of the taste of the therapeutically effective compound, by for example, exerting an effect on the gustatory system of the aquatic animal, binding to the therapeutically effective compound and preventing its release in saliva, or reducing the dissolution of the therapeutically effective compound in the saliva. Other taste-masking agents are flavouring agents.
  • Flavouring agents themselves have an odour and/or flavour that is more palatable to aquatic animals, which then have the capacity to mask the unpalatable flavour and/or odour of the therapeutically effective compound.
  • the taste-masking agent is a flavouring agent.
  • a large number of taste-masking agents for aquatic animals, particularly fish, are known in the art, including sucrose, free amino acids, nucleotides and nucleosides, organic acids, fish hydrolysates, and other compounds. The appropriate taste-masking agent can depend on the specific species of the aquatic animal population.
  • the taste-masking agent is a derivative or extract of garlic (e.g. garlic powder, diluted or pure, synthetic garlic component (allicin) or garlic oil), including odorous compounds found in garlic.
  • the taste- masking agent is an amino acid, such as betaine, L-alanine, L-glutamic acid, L-arginine, glycine and inosine.
  • the taste-masking agent is natural fish oil (cod liver oil or tuna oil), anise oil, or castor oil.
  • the amount of the taste-masking agent required in the aquatic animal feed depends on the nature of the taste-masking agent, and is preferably in the range of 1 - 10% w/w.
  • the taste-masking agent comprises about 2-8% w/w of the bead.
  • the flavouring or taste-masking agent comprises about 4-5% w/w of the bead.
  • the carrier matrix provides a means of encapsulating the therapeutically effective compound and the taste masking agent together.
  • the carrier matrices used in the compositions of the invention are capable of forming beads incorporating the taste masking agent, and the therapeutically effective compound such that the beads are of a suitable size, shape and consistency for incorporation into aquatic animal feed and consumption by the target aquatic animal population.
  • the carrier matrices also assist in maintaining the integrity of the beads when compressed into the aquatic animal feed.
  • the beads are digestible by aquatic animals such that the therapeutically effective compound is released into the digestive tract of the animal once it is ingested, rather than regurgitated or excreted intact.
  • the carrier matrices used in the present invention are preferably capable of at least partially breaking down in the aquatic animal digestive tract so that the therapeutically effective compound is able to be absorbed by the animal. This allows for targeted drug release in the gut.
  • Aquatic animal populations can find therapeutically effective compounds more palatable when provided in some carrier matrices rather than others.
  • the carrier matrix acts together with the taste-masking agent to improve the palatability of the therapeutically effective compound.
  • the therapeutically effective compound when incorporated into a suitable carrier matrix, exhibits reduced leaching into the surrounding environment when it is delivered for consumption by a population of aquatic animals. The reduced leaching of the therapeutically effective compound reduces the detection of malodour/taste of the therapeutically effective compound by the aquatic animals, improving its palatability. This has the additional effect of reducing the amount of therapeutically active compound in the aquatic animal feed additive itself.
  • the carrier matrix preferably does not substantially dissolve or disintegrate in freshwater or seawater.
  • the carrier matrix is substantially stable in sea water for at least 60 minutes, in that the beads will not disintegrate in sea water during this period.
  • the rate of leaching of the therapeutic is less than 10% over a 60 minute period in sea water. More preferably, the rate of leaching is less than 5% over a 60 minute period in sea water.
  • suitable carrier matrices are at least partially prepared from hydrogel based compounds derived from marine environments, which do not substantially dissolve or disintegrate in seawater.
  • Suitable marine based hydrogels include agar, agarose, carrageenan, fucoidan, chitosan, alginate or agar.
  • suitable carrier matrices include a combination of alginate and an acid soluble polymer (such as chitosan and/or Eudraguard ® ), alginate alone, or agar.
  • the suitable carrier matrix is a marine based collagen.
  • the carrier matrix is a combination of alginate and an acid soluble polymer (such as chitosan and/or Eudraguard ® ).
  • the carrier matrix is comprised of about 5 - 15% w/w acid-soluble polymer and 5 - 15% w/w alginate.
  • the carrier matrix is comprised of about 8-10 w/w% acid-soluble polymer and about 8-11% w/w alginate.
  • the beads are prepared by cross-linking a suspension containing the alginate/ acid-soluble polymer carrier matrix, therapeutically effective compound and taste masking agent in a solution of calcium chloride or zinc chloride.
  • the carrier matrix is comprised of alginate alone.
  • the carrier matrix is comprises 5 - 15% w/w alginate.
  • the carrier matrix is comprised of about 5-11% alginate.
  • the carrier matrix is prepared from agar. Any type of agar can be used as the carrier matrix, including commercial food-grade agar, agarose and other pharmaceutical or microbiological grade agar.
  • the carrier matrix is comprised of about 15 - 25% agar.
  • the carrier matrix comprises food-grade agar and comprises about 18-20% w/w agar.
  • these beads are prepared by setting the suspension containing the agar carrier matrix, therapeutically effective compound and taste-masking agent into an oil, emulsifier (e.g. Tween 80), or buffer.
  • an oil emulsifier or buffer used to set the agar carrier matrix has taste-masking properties as a flavouring agent, in that it has a flavour which is palatable to the aquatic animal.
  • an oil is used to set the suspension containing the agar carrier matrix.
  • the oil used may be garlic oil (that is, oil containing a garlic derivative), and/or cod liver oil.
  • the beads additionally comprise one or more wetting agents, surfactants or disintegrants.
  • the additive or carrier matrix is poorly soluble in an animal’s digestive tract (such as the gastric or intestinal tracts)
  • the additive may include a wetting agent or surfactant polymer or pharmaceutical disintegrant to assist in disintegration of the additive.
  • the additive or carrier matrix can include a wetting agent or surfactants polymer or pharmaceutical disintegrant, that may or may not breakdown in an animal’s digestive tract, which assists in the manufacture of the additive and or enhances the wetting of the additive in an aqueous environment.
  • a wetting agent can be added to help with the wetting of the drug powder so that it can be uniformly dispersed in the matrix.
  • the wetting agent also assists in displacing entrapped air between the drug and the carrier matrix to give a stronger bind between the carrier matrix and the therapeutic- entrapped air gives rise to friable beads that are then easily crushed when incorporated into the fish feed, resulting in the fish being able to taste the exposed therapeutic.
  • the wetting agent has the effect of reducing the amount of air trapped in the suspension from which the beads are formed.
  • the amount of air trapped in the suspension has the capacity to affect the brittleness of the beads.
  • a reduction of the air trapped in the suspension increases the compressibility of the bead, and therefore reduces the risk of crushing and consequent leaching of the therapeutic agent when the bead is incorporated into fish feed.
  • the wetting agent can be added to help with the wetting of the drug powder so that it can be uniformly dispersed in the alginate solution.
  • the wetting agent also helps to displace the entrapped air between the drug and the sodium alginate solution (a result of poor wetting) to give strong beads - entrapped air in the final beads gave rise to friable beads that were then easily crushed when incorporated into the fish feed, resulting in the fish being able to taste the exposed praziquantel.
  • wetting agents or surfactants include, for example, and without limitation, anionic or cationic detergents or a non-ionic wetting agent.
  • wetting agents or surfactants include, for example and without limitation, anionic detergents such as sodium lauryl sulfate, dioctyl sodium sulfosuccinate and dioctyl sodium sulfonate.
  • anionic detergents such as sodium lauryl sulfate, dioctyl sodium sulfosuccinate and dioctyl sodium sulfonate.
  • Cationic detergents might be optionally used and could include, without limitation, benzalkonium chloride or benzethomium chloride.
  • nonionic wetting agents that could be included in the formulation include Cremophor ® RH40, lauromacrogol 400, polyoxyl 40 stearate, polyoxyethylene hydrogenated castor oil 10, 50 and 60, glycerol monostearate, polysorbate 40, 60, 65 and 80 (Tween), sucrose fatty acid ester, methyl cellulose and carboxymethylcellulose.
  • Cremophor ® RH40 lauromacrogol 400
  • polyoxyl 40 stearate polyoxyethylene hydrogenated castor oil 10
  • 50 and 60 glycerol monostearate
  • polysorbate 40, 60, 65 and 80 Tween
  • sucrose fatty acid ester methyl cellulose and carboxymethylcellulose.
  • the wetting agent is Cremophor ® RH40 or polysorbate (Tween).
  • the wetting agent is 21 , 21 , 40, 60, 61 , 65, 80, 81 and/or 85 Tween.
  • the bead is comprised of about 5 - 15% Cremophor ® RH40.
  • Disintegrants can be, for example, starches and modified starches, cellulose, etc.
  • suitable carrier matrices include a combination of alginate and an acid soluble polymer (such as chitosan and/or Eudraguard ® ) and a wetting agent (such as Cremophor ® RH40 or Tween).
  • an acid soluble polymer such as chitosan and/or Eudraguard ®
  • a wetting agent such as Cremophor ® RH40 or Tween
  • the carrier matrix is comprised of about 5 - 15% w/w acid- soluble polymer and 5 - 15% w/w alginate and a wetting agent.
  • the wetting agent is selected from Cremophor ® RH40 and/or Tween. Most preferably, the wetting agent is present at about 5 - 10% w/w. Most preferably, the wetting agent is Cremophor ® RH40 and is present at about 5 - 10% w/w.
  • the carrier matrix is comprised of alginate and a wetting agent.
  • the carrier matrix is comprised 5 - 15% w/w alginate and a wetting agent.
  • the carrier matrix is comprised of about 5-11% w/w alginate and a wetting agent.
  • the bead additionally comprises a wetting agent selected from Cremophor ® RH40 and/or Tween. Most preferably, the wetting agent is Cremophor ® RH40 and is present at about 5 - 10% w/w.
  • Aquatic Animal Feed comprising the Aquatic Animal Feed Additive
  • the invention provides an aquatic animal feed comprising the aquatic animal feed additive of the invention in a therapeutically effective amount.
  • the aquatic animal feed additive beads of the invention can be incorporated into aquatic animal feed, which can then be delivered to the target aquatic animal population.
  • the aquatic animal is a fish. Any aquatic animal feed that can safely be offered to the target aquatic animal population can be used to prepare the aquatic animal feed of the invention.
  • the beads of the invention are mixed with crushed aquatic animal feed during the manufacturing process, compressed and then cut into pellets or granules of an appropriate size for the target aquatic animal population.
  • the beads of the invention are dispersed throughout the aquatic animal feed, and not merely coated on the surface of the aquatic animal feed.
  • a person skilled in the art can readily determine the amount of aquatic animal feed additive that must be added to the aquatic animal feed in a given situation in order to constitute a therapeutically effective amount. This will depend on a range of factors including the nature of the therapeutically effective compound, the aquatic animal population, and the percentage drug loading in the beads.
  • this invention provides a method of treating a disease in an aquatic animal comprising administering an aquatic animal feed additive or aquatic animal feed of the invention to the aquatic animal population.
  • the aquatic animal is a fish.
  • compositions of the invention can be used to treat a disease that an aquatic animal population is suffering.
  • the compositions of the invention can be used prophylactically to prevent an aquatic animal population from developing a disease.
  • the therapeutically effective compound is an anthelmintic drug, such as praziquantel.
  • the aquatic animal feed additive or aquatic animal feed of the invention comprising praziquantel is administered to a fish population such that the fish population receives a dose of between 50mg/kg and 150mg/kg of praziquantel.
  • a range of different diseases can be treated or prevented using the compositions of the invention including parasitic diseases such as those caused by cestodes or trematodes including monogenean flukes, and blood flukes.
  • parasitic diseases such as those caused by cestodes or trematodes including monogenean flukes, and blood flukes.
  • Families of monogenean parasites that causes disease in aquatic animals include Capsalidae, Dactylogyrus vastator, Centrocestus formosanus and Digenea Heterophydae.
  • the disease is a parasitic infection caused by monogenean flukes. In a further preferred embodiment, the disease is a parasitic infection caused by Zeuxapta flukes.
  • compositions of the inventions can be used to treat or prevent diseases in an aquatic animal.
  • the aquatic animal is any species of fish, including yellowtail kingfish or Pacific Bluefin Tuna.
  • the fish is of a species that is known to be particularly taste and/or odour sensitive.
  • the fish is of the species Seriola lalandi.
  • the skilled person will know the amount of the aquatic animal feed or aquatic animal feed additive to deliver to the aquatic animal population, and the dosage schedule required in order to achieve a therapeutic effect in the aquatic animal population.
  • the aquatic animal feed additives of the invention are prepared using a method comprising the following steps:
  • the dry mixture in step (a) is prepared by mixing an effective amount of a therapeutically effective compound such as praziquantel with a taste-masking agent such as garlic powder.
  • a therapeutically effective compound such as praziquantel
  • a taste-masking agent such as garlic powder.
  • This step can also optionally include mixing a portion of the carrier matrix (such as chitosan) and/or wetting agent.
  • a solution containing a carrier matrix is added to the dry mixture to form a homogenous suspension.
  • the carrier matrix is an acid soluble polymer (such as chitosan or Eudraguard ® ), and the solution is made by mixing acid-soluble polymer powder with acetic acid.
  • chitosan powder is dissolved in 0.2M acetic acid to prepare a 1% chitosan solution.
  • the carrier matrix is alginate, preferably in a 1% w/v solution.
  • the carrier matrix is agar, and is added in a 2% w/v solution.
  • the carrier matrix is alginate.
  • the carrier matrix is chitosan dispersed in alginate, where chitosan is included as part of the dry mixture in step (a) and alginate solution is then added to the dry mixture, stirred and sonicated in a sonicating bath to form a homogenous suspension (stirring time may range from 30 minutes to overnight).
  • a wetting agent Preferably, the wetting agent is Cremophor ® RH40 and/or Tween.
  • Cremophor ® RH40 is mixed together with the therapeutically effective compound and taste-masking agent (and optionally a portion of the carrier matrix) in step (a), and a solution of alginate preferably 1% w/v is added to the dry mixture.
  • the homogenous suspension is dropped in a liquid medium to form beads.
  • the appropriate liquid medium is selected based on carrier matrix used, and will be known to persons in the art.
  • the beads may be required to sit in the liquid medium for some time in order to form properly.
  • the carrier matrix comprises alginate, and the liquid medium is a calcium chloride solution.
  • the calcium chloride solution is at a concentration of 1% w/v.
  • the carrier matrix comprises agar, and the liquid medium is a cold oil or mixture of oils, emulsifier or buffer.
  • the oil is selected from one or more of garlic oil, cod liver oil and soybean oil.
  • the oil contains garlic oil, which can have an additional taste-masking effect.
  • step (d) the beads are taken out of the liquid medium and washed with water, preferably deionised water.
  • the beads are dried.
  • the beads may be dried through any of the methods known in the art.
  • the beads may be dried through air drying, spray drying, convective drying, vacuum drying, microwave drying, lyophilisation and combinations thereof.
  • the beads are air dried, for example, in a fume hood.
  • the beads are dried for at least 7 days.
  • the dried beads are additionally mixed with a taste-masking agent in order to coat the beads with this agent.
  • this taste-masking agent is garlic powder.
  • the aquatic animal feed additive can be incorporated into aquatic animal feed.
  • the fish feed is prepared by mixing the aquatic animal feed additive prepared by the above process with a commercial fish feed, and then pelleting the mixture.
  • the mixture may be pelleting using any machine or device suitable for that process.
  • the commercial fish feed used in the process, and the size and shape of the pellets can be chosen based on the requirements of the target aquatic animal population.
  • the aquatic animal feed additive composition is prepared using an extruder.
  • the extruder is subjected to the following heat and pressure parameters: >90 °C pre-conditioner for approximately 30 seconds, followed by extruder temperature of 90 °C to 120 °C at >20 bar cylinder pressure in the zone of high pressure, followed by drying at 90 °C to 110 °C for 44 minutes.
  • the clearance between the screw and barrel within an extruder is less than 1 mm.
  • the aquatic animal feed additive composition is not substantially disrupted or degraded by these parameters.
  • the bead size is kept to a diameter of 500 pm (0.5 mm) and preferably under 1 mm.
  • the extruder is subjected to the following heat and pressure parameters: 70 °C and pressure at 20 - 30 bar, with a extruder retention time (mixing through the extruder) of 25 - 30 seconds.
  • the aquatic animal feed additive composition is not substantially disrupted or degraded by these parameters.
  • Example 1 Preparation of beads.
  • PZQ praziquantel
  • Formulations A and E (the chitosan beads) were prepared by dissolving the chitosan powder in 0.2M acetic acid to prepare a 1% chitosan solution. The praziquantel and garlic derivative was then mixed with the chitosan solution. The resulting suspension was then dropped in a bath of 2% w/v sodium triphosphate solution in order to form beads. Formulation A was allowed to complex for 3 hours, whereas formulation E was allowed to complex overnight. The beads were then air dried in a fume hood for at least 7 days. Once dried, garlic powder equating to 1% of the total bead weight was mixed with the dry beads.
  • Formulation B was prepared by preparing a dry mixture of the praziquantel, chitosan and garlic derivative. The 1% (w/v) alginate solution was then added to the dry mixture, and stirred to form a homogenous suspension. The suspension was then dropped into a 1% (w/v) calcium chloride solution in order to form the beads. The beads were allowed to sit in the calcium chloride solution for another 10-15 minutes and washed with deionised water. The beads were then air dried in a fume hood for at least 7 days. Once dried, garlic powder equating to 1% of the total bead weight was mixed with the dry beads.
  • Formulation B * was prepared by preparing a dry mixture of the praziquantel, chitosan, garlic derivative and Cremophor ® RH40. The 1% (w/v) alginate solution was then added to the dry mixture, and stirred to form a homogenous suspension. The suspension was then dropped into a 1% (w/v) calcium chloride solution in order to form the beads. The beads was then allowed to sit in the calcium chloride solution for another 10-15 minutes and washed with deionised water. The beads was then air dried in a fume hood for at least 7 days. Once dried, garlic powder equating to 1% of the total bead weight was mixed with the dry beads.
  • Formulation C was prepared by preparing a dry mixture of the praziquantel and garlic derivative.
  • the 2% (w/v) agar solution (when hot and runny) was then added to the dry mixture and stirred to form a homogenous suspension.
  • the suspension was then dropped into cold garlic oil (which was comprised of garlic oil: soybean oil: cod liver oil in a ratio of 5:3:1 ) to form the beads.
  • the beads were then washed with deionised water and then air dried in a fume hood for at least 7 days. Once dried, garlic powder equating to 1% of the total bead weight was mixed with the dry beads.
  • Formulation D was prepared by preparing a dry mixture of the praziquantel, Cremophor® RFI40 and garlic derivative. The 1% (w/v) alginate solution was then added to the dry mixture and stirred to form a homogenous suspension. The suspension was then dropped into a 1 % (w/v) calcium chloride solution in order to form the beads. The beads were allowed to sit in the calcium chloride solution for another 10-15 minutes and washed with deionised water. The beads were then air dried in a fume hood for at least 7 days. Once dried, garlic powder equating to 1% of the total bead weight was mixed with the dry beads.
  • Beads were placed in vials containing 10ml_ 0.1 M HCI or seawater and were observed over a period of 5 hours they were stirred using a magnetic stirrer at 10Orpm.
  • simulated fish gastric fluid SGF, seawater adjusted to pH 2.0 ⁇ 0.05 with HCI, with 0.1% w/v Polysorbate 80 and 0.8 mg/ml_ pepsin
  • SIF simulated fish intestinal fluid
  • the respective dissolution medium was sampled (1 ml.) at 0 and 5 min (seawater), 35 and 65 min (SGF), and 125 and 185 min (SIF). Withdrawn samples were filtered (0.45 _m) and analysed for praziquantel content using the HPLC assay.
  • Control experiments were performed using pure PZQ powder (7-13 mg per basket). However, as the basket could not be transferred from one dissolution medium to the next without loss of powder, the experiments for the PZQ powder were performed separately in the three dissolution media as follows: 5 min in 500 ml. of seawater, 60 min in 100 ml. of SGF, and 120 min in 100 ml. of SIF. Additionally, the dissolution profiles of Formulation B and Formulation C beads were determined over 3 h in seawater using the same equipment set-up.
  • Formulation B and Formulation C beads did not disintegrate even after 3 h incubation in 500 mL of seawater, and only 6.51 ⁇ 0.59% and 1.36 ⁇ 0.71% of the drug loads from the respective beads were leached into the seawater at 3 h ( Figure 2a). Simulation of the bead passage from seawater into the fish GIT showed undetectable drug release after 5 min in seawater from both formulations.
  • the Formulation B beads remained intact after a further 60 min incubation in SGF, releasing only 2.3 ⁇ 0.4% of the drug load; however, bead disintegration was noted in the SIF accompanied by the release of 84.7 ⁇ 2.9% of the drug load at 185 min ( Figure 2b).
  • Example 4 Analysis by Differential Scanning Calorimetry (DSC).
  • DSC25 System TA Instruments, Newcastle, DE, USA
  • Bead samples were analysed within 14 days of manufacture. Samples ( ⁇ 3 mg) were analysed in standard aluminium pans (DSC Consumables Incorporated, Austin, MN, USA) over 0 to 250°C at a heating rate of 10°C/min with empty aluminium pans as reference. DSC thermograms were analysed using the TRIOS Software (TA Instruments, New Castle, DE, USA.
  • PZQ peak in Formulation C (3.4 mg) had onset at 138.03°C and peak temperature at 141.93°C. Its enthalpy of 71 .645 J/g was 71 .74% that of pure PZQ, which again corresponded closely to the drug loading of Formulation C. Thus, it may be concluded from the respective DSC thermograms that PZQ retained its crystalline characteristics, and did not interact with the matrix materials in Formulations B and C.
  • Example 5 Fish palatability trials.
  • Each of the dried beads of formulations A to E were mixed with crushed commercial yellowtail kingfish fish feed, and pelleted.
  • the feed contained the equivalent of 10 grams of pure praziquantel per kilogram of feed.
  • Healthy yellowtail kingfish were offered a fixed ration based on their weight and water temperature of the fish feed pellets containing the formulations. The fish were fed for a maximum duration of 3 minutes in a single morning feed over a 5 day period.
  • Healthy yellowtail kingfish fish were offered a fixed ration per tank per day in a single morning feed over a day period, calculated on the basis of their size, and the water temperature.
  • Figure 3 presents the results for fish feed containing formulations B, C and D.
  • Example 8 Drug Loading Stability.
  • the drug loading capacity (or more specifically residual intact drug content) of the beads of formulations B, C and D was determined 18 months after the beads were initially prepared according to the method described in Example 7. The results are set out in Table 3 below: Table 3
  • Example 9 Palatability of fresh beads compared with stored beads.
  • a second palatability trial was conducted to compare formulation B and C, with fish feed incorporating pure praziquantel (which had not been incorporated within bead, or with garlic extract).
  • Beads of formulation B or C were mixed with ground 3mm commercial yellowtail kingfish feed, and were reconstituted into pellets using a Dolly pasta maker.
  • Pellets were also made with pure praziquantel powder using the same process. All pellets contained the equivalent of 10 grams/kg of pure praziquantel.
  • the control diet was also made using the same process, which contained no praziquantel. Yellowtail kingfish fish with an average weight of 1600 grams, infected with Zeuxapta flukes were offered a fixed ration of 155 grams per tank per day in a single morning feed over a 6 day period.
  • Figure 9C presents the average daily dose of praziquantel ingested by the fish receiving the different treatments in mg/kg.
  • the actual praziquantel dose ingested by fish fed the pure praziquantel fish feed composition was 21 mg/kg.
  • the results therefore demonstrate that formulations B and C clearly allow the oral delivery of greatly increased quantities of praziquantel to the fish. It is generally considered that fish must receive approximately 50 mg/kg for three consecutive days to eliminate Zeuxapta. This dose was easily achieved in the fish feed containing formulations B and C but not in the pure praziquantel treatment.
  • Example 11 Treatment of fish populations suffering from fluke.
  • Example 12- Methods to prepare beads of different sizes Beads of different sizes (diameters) in the range 0.4 to 2.5 mm were prepared using methods described in the following paragraph. The bead sizes were regulated by choosing the appropriate chitosan particle size, extrusion needle and stirring speed. The chitosan size ranges were: (1) 355-500pm; (2) 212-355 pm; (3) 150-355 pm; and (4) ⁇ 150 pm. [00197] Firstly, a mixture of the praziquantel, chitosan and garlic derivative (with and without
  • Cremophor ® RH40 Cremophor ® RH40 was prepared.
  • the 1% (w/v) alginate solution was then added to the mixture, stirred and sonicated to form a homogenous suspension. Stirring of the suspension ranged from 30 minutes to overnight.
  • the suspension was then dropped into a 1% (w/v) calcium chloride solution in order to form the beads. Syringe needles of different sizes were used to drop the suspension, e.g. 18G or 21 G needles were used to form smaller beads with the smaller chitosan particles.
  • the beads were allowed to sit in the calcium chloride solution for another 10-15 minutes and washed with deionised water. The beads were then air dried in a fume hood for at least 7 days. Once dried, garlic powder equating to 1% of the total bead weight was mixed with the dry beads.

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Abstract

La présente invention concerne le domaine de l'administration orale de composés thérapeutiques à des poissons par l'intermédiaire d'additifs alimentaires pour poissons, y compris des compositions d'additifs alimentaires pour animaux aquatiques, ainsi que des méthodes de traitement ou de prévention de maladies chez des animaux aquatiques.
EP22831080.1A 2021-07-01 2022-07-01 Additifs alimentaires pour poisson Pending EP4362945A4 (fr)

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AU2021902007A AU2021902007A0 (en) 2021-07-01 Fish Feed Additives
PCT/AU2022/050690 WO2023272362A1 (fr) 2021-07-01 2022-07-01 Additifs alimentaires pour poisson

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AU2022416992A1 (en) * 2021-12-13 2024-06-27 Clean Seas Seafood Limited Solid dosage forms for aquaculture
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NO882653D0 (no) 1988-06-15 1988-06-15 Apothekernes Lab Doseringsform.
FR2745979B1 (fr) * 1996-03-15 1998-05-29 Additif alimentaire pour animaux
GB2403407B (en) 2003-07-03 2006-12-13 Michael Hilary Burke Palatable oral anthelmintic composition
AU2008100441A4 (en) 2007-07-25 2008-06-12 Diggles, Ben Dr Medicated fish feed with feeding attractant
WO2009023013A1 (fr) 2007-08-13 2009-02-19 Alpharma, Inc. Compositions de praziquantel et de cimétidine et procédés
HU231017B1 (hu) * 2012-05-08 2019-11-28 LAVET Gyógyszeripari Kft. Praziquantel tartalmú ízfedett formulációk
WO2013187897A1 (fr) 2012-06-14 2013-12-19 Empire Technology Development Llc Compositions à ingérer de densité neutre
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CN105661005A (zh) * 2014-11-21 2016-06-15 广西大学 一种驱虫并提高免疫力的鱼用中草药饲料添加剂
AU2016281617A1 (en) * 2015-06-26 2018-01-18 Advanced Bionutrition Corporation Composition for delivery of active agents to an animal
AU2015101787A4 (en) * 2015-12-11 2016-01-28 Virbac (Australia) Pty Limited Antiparasitic formulations for oral administration
CN113134008B (zh) * 2020-01-17 2022-08-30 中国农业大学 宠物用伊维菌素吡喹酮咀嚼片及其制备方法

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