EP4133055A1 - Compositions comprenant des algues et leurs méthodes d'utilisation pour augmenter la production de produit animal - Google Patents

Compositions comprenant des algues et leurs méthodes d'utilisation pour augmenter la production de produit animal

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Publication number
EP4133055A1
EP4133055A1 EP21785593.1A EP21785593A EP4133055A1 EP 4133055 A1 EP4133055 A1 EP 4133055A1 EP 21785593 A EP21785593 A EP 21785593A EP 4133055 A1 EP4133055 A1 EP 4133055A1
Authority
EP
European Patent Office
Prior art keywords
biomass
bromoform
iodine
present technology
asparagopsis
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
EP21785593.1A
Other languages
German (de)
English (en)
Other versions
EP4133055A4 (fr
Inventor
Vivienne HAY
Matthew ROTHE
Joan SALWEN
Michael BRACCO
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.)
Blue Ocean Barns
Original Assignee
Blue Ocean Barns
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Filing date
Publication date
Application filed by Blue Ocean Barns filed Critical Blue Ocean Barns
Publication of EP4133055A1 publication Critical patent/EP4133055A1/fr
Publication of EP4133055A4 publication Critical patent/EP4133055A4/fr
Pending legal-status Critical Current

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Classifications

    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23K—FODDER
    • A23K10/00—Animal feeding-stuffs
    • A23K10/10—Animal feeding-stuffs obtained by microbiological or biochemical processes
    • A23K10/12—Animal feeding-stuffs obtained by microbiological or biochemical processes by fermentation of natural products, e.g. of vegetable material, animal waste material or biomass
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
    • A01G33/00—Cultivation of seaweed or algae
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
    • A01H13/00—Algae
    • 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
    • A23K20/105—Aliphatic or alicyclic compounds
    • 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
    • A23K40/35—Making capsules specially adapted for ruminants
    • 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
    • A23K50/10—Feeding-stuffs specially adapted for particular animals for ruminants
    • 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

Definitions

  • the present technology generally relates to algae-derived biomass and to compositions comprising same.
  • the present technology also generally relates to use of such algae- derived biomass and compositions comprising same in methods for feeding animals, in methods for increasing animal product production; and in methods of decreasing methane emission from animals.
  • Methanogenesis i.e., the production of methane by ruminant animals
  • CH4 methane
  • the CH4 analogues include bromochloromethane (BCM), bromoform and chloroform and have been proven to be the most effective feed additives for reducing CH4 production.
  • BCM bromochloromethane
  • Naturally synthesized bromoform administered via supplementation of Asparagopsis spp. was found more effective than synthetic methane analogs (Machado et. al. 2018). Supplementation with gametophytes of Asparagopsis taxiformis comprising 7.8 mg/g of bromoform by dry weight has been found to mitigate an 80% reduction of methane gas production over a 147-day period in steers fed a 0.5% seaweed supplemented low forage total mixed ration (LF-TMR) (Roque, 2020).
  • LF-TMR low forage total mixed ration
  • Asparagopsis taxiformis Asparagopsis taxiformis (AT) as a feed additive to inhibit methanogenesis in ruminant animals is constrained by several factors. These include its unpleasant odor, high iodine content, epiphytic nature, and the lack of capacity, especially in male AT specimens, to synthesize material concentrations of the halogenated compounds.
  • US Patent Application 2019/0174793 describes a formulation such that the animal is provided with of red marine macroalgae per day for animals maintained at pasture and states that amounts to about 1-5% of algae on a dry matter basis or 1-3% on an organic matter basis per day.
  • US Patent Application 2019/0174793 also discloses providing an animal with about 200-600 g/day of algae for animals on a finishing diet in feedlots.
  • this application discloses the filamentous tetrasporophyte lifestage of Asparagopsis as a potential feedstock for a feed pre mix.
  • FIG. 1 depicts the target metabolites comprising halogenated metabolites and iodine.
  • FIG. 2 shows the growth of tetrasporophyte material in the nursery as the
  • filamentous form uniform red material in the Erlenmeyer flasks, and the “puffball” form as the darker red floating spheres.
  • FIG. 3 shows a closeup of the non-filamentous tetrasporophyte form.
  • FIG. 4 shows another closeup of the non-filamentous tetrasporophyte form.
  • FIG. 5 shows a flow chart of a method for culture enhancement cycles that enable step-wise improvement in the synthesis of halogenated metabolites that forces iodine concentrations lower within target biomass in the tetrasporophyte life stage according to one embodiment of the present technology.
  • FIG. 6 shows naturally occurring gametophytes (the parent plant) and some anatomical differences between tetrasporophytes and gametophytes, for example in terms of the size and shape of the plants.
  • FIG. 7 shows a photomicrograph of tetrasporophyte material growing in the laboratory. Note the unusually large gland cells (the orange/black dots). The gland cells are what contains the active ingredient (bromoform). Large gland cells indicate an unusually high concentration of bromoform.
  • FIG. 8 is a photomicrograph of tetrasporophytes creating undesired spores. Spores detract energy from growth and bromoform synthesis.
  • FIG. 9 is a photo of wild tetrasporophytes used as starting material for the breeding and cultivation program.
  • FIG. 10 depicts a graph illustrating a system that calculates precise inclusion rates of red algae in livestock feed and supplements according to one embodiment of the present technology.
  • FIG. 11 depicts a graph illustrating a system in which biological methods are used to synthesize and encapsulate bromoform according to one embodiment of the present technology.
  • FIG. 12 depicts a graph showing projected intermittent feeding impact for dairy.
  • FIG. 13 depicts a graph showing projected intermittent feeding impact on beef.
  • the present technology relates to a biomass derived from
  • the biomass comprising a ratio of halogenated metabolite to iodine that is equal to or less than about 700:1.
  • the present technology relates to a biomass derived from Asparagopsis taxiformis, the biomass comprising a ratio of halogenated metabolite to iodine that is between about 5: 1 and about 700: 1.
  • the present technology relates to a biomass derived from Asparagopsis taxiformis, the biomass comprising a ratio of halogenated metabolite to iodine that is equal or greater than about 700:1.
  • the Asparagopsis taxiformis is Asparagopsis taxiformis tetrasporopgyte.
  • the halogenated metabolite is selected from any one of structures 1 to 78 of FIG. 1 or any combination thereof.
  • the present technology relates to a method for cultivating the biomass of as defined herein, the method comprising: i) collecting a parent Asparagopsis taxiformis plant; ii) manipulating the plant of i) to obtain filaments of Asparagopsis that are substantially free of contaminants; and selecting for Asparagopsis taxiformis aragopsis that exhibit enlarged gland cells.
  • the present technology relates to a method for reducing methane production in a ruminant, the method comprising administering between about 10 g and about 60 g per day of the biomass as described herein to the ruminant.
  • the present technology relates to a method for reducing methane production in a ruminant, the method comprising administering between about 10 g and about 30 g per day of the biomass as defined herein to the ruminant.
  • the present technology relates to a method for reducing methane production in a ruminant, the method comprising administering between about 5 g and about 25 g per day of the biomass as defined herein to the ruminant.
  • the present technology relates to a kit comprising an algal feed supplement comprising: at least about 20% of neutral dietary fiber (NDF) by dry weight of the algal feed supplement; at least about 16% protein by dry weight of the algal feed supplement; less than about 3000 ppm iodine by dry weight of the algal feed supplement; and at least about 2.5% of a halogenated metabolite by dry weight of the algal feed supplement.
  • NDF neutral dietary fiber
  • the term “about” is used herein explicitly or not, every quantity given herein is meant to refer to the actual given value, and it is also meant to refer to the approximation to such given value that would reasonably be inferred based on the ordinary skill in the art, including equivalents and approximations due to the experimental and/or measurement conditions for such given value.
  • the term “about” in the context of a given value or range refers to a value or range that is within 20%, preferably within 15%, more preferably within 10%, more preferably within 9%, more preferably within 8%, more preferably within 7%, more preferably within 6%, and more preferably within 5% of the given value or range.
  • a quantifier such as “less than”, “more than”, or “greater than” modifies a comma delimited list of numbers or quantities, or ratios, such as “1, 2.5, 3, ...” or “one, two, three...” the intention is that the quantifier applies to every member of the list.
  • “alpha is greater than about 1, 2, or 3, or 4” means “alpha is greater than about 1, or, alpha is greater than about 2 or alpha is greater than about 3.”
  • the Asparagopsis taxiformis variety of the present technology is anatomically distinguished from the parent plant in a number of ways and was accomplished by a vegetative breeding program to increase bromoform concentration under novel cultivation environments. Tests were initially carried out to test typical wild Asparagopsis taxiformis gametophytes for bromoform and it was found a typical sample comprised 6.63 mg/g of bromoform for freeze dried material, matching closely the value reported by Machado. Next, wild Asparagopsis taxiformis tetrasporophytes was harvested, as shown in FIG. 9 and it was found to comprised only 1.1 mg/g of bromoform. Even with the increased ease of cultivation, this level of bromoform would make commercial supplementation of cattle costly.
  • tetrasporophytes quickly mature to a filamentous form that has been previously studied as a feed supplement, under our conditions, the tetrasporophytes maintain a non-filamentous appearance, grow rapidly, exhibit high bromoform concentrations, have low iodine concentration / bromoform concentration ratios and can be kept in these stages in bioreactors for indefinite times.
  • These morphotypes can be described as, “puff ball” forms. Both the filamentous and non-filamentous puffball forms are shown in FIGS. 2-4.
  • the supplement comprising the tetrasporophyte variety grown under the herein described conditions can be supplemented without the necessity of adding molasses to the biomass that is typically needed to increase palatability and prevent feed refusal.
  • the feed supplement compositions of the present technology do not comprise sweeteners such as, but not limited to, molasses, high fructose corn syrup, sucrose, fructose, xylitol, sorbitol, or other alcohol sugar appetants.
  • the feed supplement compositions of the present technology may be used to supplement animals at a rate of less than 10 g/day, or 7.5 g/day, or 5 g/day of the algal biomass described here for animals grazing on a pasture.
  • the feed supplement compositions of the present technology may be used to supplement feedlot animals on finishing diets at a daily supplementation rate of less than 200 g/day, less about 150 g/day, less than 100 g/day, or 50 g/day of the algal biomass described here for animals on finishing diets.
  • animals will have different supplementation rates depending on whether they are being raised for dairy or meat, are grazed solely on pasture, solely on grain, or on transition diets.
  • the algorithms and supplementation rates and methods described herein will take into account the amount of neutral dietary fiber (NDF, or sometimes referred to as aNDF).
  • the present investigators discovered how to induce the growth and indefinite maintenance of “puffball” forms of the tetrasporophyte that appear to be microscopic oligocellular forms which are very morphologically and chemotypically distinct from the gametophyte macroalgal stage shown in FIG 6.
  • the present investigators discovered that certain of these tetrasporophyte varieties in particular morphotypes are very well suited for use as feed supplements as a source of halogenated compounds to inhibit methanogenesis, promote growth of animals (e.g., ruminant animals) as well as increase the quality of products derived therefrom.
  • the present technology thus relates to a Asparagopsis taxiformis that is created through the aforementioned collection, manipulation, dissection and selection process.
  • the resulting plant is a small red alga comprising microscopic branched chains of cells shown in FIG 7.
  • the cells in the tetrasporophyte are not highly differentiated. Instead, each cluster of four cells is roughly equivalent and these clusters string together into long chains. The color ranges from pale pink to red to dark cherry.
  • Each branch contains gland cells where the bromoform is stored. These gland cells are a dark red to brown in color, with deeper color indicating higher bromoform concentration.
  • the plant is not rooted, but rather free-floating in water. It obtains all its organic and inorganic nutrients from the water and can live in this state indefinitely.
  • this variety is anatomically distinguished from others by stasis in the third phase, the tetrasporophyte phase.
  • Wild AT typically follows a progression through three life stages (gametophyte, carposporophyte and tetrasporophyte).
  • the present variety is static in the tetrasporophyte phase. This is particularly beneficial because one hundred (100%) percent of tetrasporophytes create high levels of bromoform, in contrast to just fifty (50%) percent of gametophytes synthesize meaningful amounts of bromoform.
  • the present tetrasporophytes are static in phase they are not producing spores, unlike the tetrasporophyte form shown in FIG 8. This means they can devote all of their energy to growth, which is correlated with even higher bromoform concentrations.
  • the present variety is specific even within the tetrasporophyte class. While tetrasporophytes, left to their own devices, devolve from “puffballs” into a filamentous form, the present tetrasporophytes can be maintained in the “puffball” phase. This is advantageous because the “puffball” form grows faster than the filamentous form. Again, this may be correlated with higher bromoform concentrations.
  • composition of the present variety is materially different to the mother variety.
  • it has a much higher bromoform to iodine ratio.
  • the lower iodine levels may, in part, be due to high rates of bromoform synthesis and storage displacing iodine in gland cells (where bromoform is stored), as outlined in Table 1 below:
  • Table 1 Comparison of Bromoform and Iodine content in Gametophyte and Tetrasporophyte in the types of algal biomass studied here, we assert that the to iodine content of the wild tetrasporophyte assayed here for bromoform is significantly higher than in our AT brominate variant.
  • AT tends to grow as epiphytes.
  • the present variety is distinct because it grows as an isolated algae species. This has a number of advantages for algal culture, including the fact that all nutrients go towards the growth of AT rather than competitive species and increased product purity. Nevertheless, AT is a fragile species, highly vulnerable to pests, diseases and competitive algae.
  • the introduction of pests or contaminants may be prevented through a variety of mechanisms such as, but not limited to: purification cycles, maintaining positive air pressure in the flasks, using stoppers on flasks to prevent ingress of materials, wearing lab coats and using shoe dips to prevent pests or contaminants entering the lab.
  • AT has low resistance to shipping or environmental changes. It can be killed or bleached by changes in temperature or light intensity.
  • the plant is grown under controlled environmental conditions.
  • Light is provided by incandescent, halogen, LED, fluorescent, high intensity discharge, metal halide, high pressure sodium or other suitable lights and maintained at lO-lOOmE in the seed bank and nursery using 60-80% Blue Pearl shade cloth.
  • filtered and controlled natural light if available, properly filtered, and of sufficient duration may also be used as the main, light source, or as a supplement or complement to the artificial light sources named herein, but tightly controlled artificially supplied light is preferable.
  • the photoperiod is maintained at 12 hours per day to prevent spore formation.
  • flasks are aerated to ensure algae have sufficient supply of CO2 for photosynthesis and O2 for respiration.
  • Aeration also serves to promote movement of biomass. This ensures all algae have access to light, reduces the formation of biofilm, and prevents clumping of algae, which can create an anoxic environment where bacteria or contaminants grow. Nutrients are provided through F/2 medium in approximately the concentrations depicted in Table 2. Temperature is maintained at 65-85°F (between about 18 and 30° Celsius) throughout the day.
  • the present Asparagopsis taxiformis variety is a stable and uniform culture that is distinct from the parent plant. Wild type Asparagopsis has unpleasant odor, high iodine content, epiphytic nature, and lack of capacity, especially in male specimens, to synthesize material concentrations of the halogenated compounds.
  • the present variety has higher bromoform content, lower odor, lower iodine, an absence of epiphytes and is static in the tetrasporophyte phase.
  • the current technology provides for a Asparagopsis taxiformis derived biomass with a bromoform to iodine ratio of equal to or greater than about 5:1, 10:1, 15: 1, 20: 1, 25: 1, 50:1, 75:1, 100: 1, 150:1, 200: 1, 300: 1, 400:1. 500:1, 600: 1, or 700:1.
  • the current technology provides for a non-filamentous
  • Asparagopsis taxiformis derived biomass comprising greater than about 5, about 6, about 7, about 8, about 9, 10, about 11 , about 12, about 15, or about 20 mg/g of bromoform w/w of dried material.
  • the current technology provides for non-filamentous
  • Asparagopsis taxiformis tetrasporophyte derived biomass obtained by unattached cultivation in a continuous aeration induced circulatory flow bioreactor.
  • the current technology provides for non-filamentous Asparagopsis taxiformis derived biomass continuously grown for 1, 2, 3, 4, 5, 6, 7, 9, or 10 weeks.
  • the present technology also provides non-filamentous Asparagopsis taxiformis algal biomass that is produced through a cultivation cycle comprising a seedbank, nursery, and outdoor phase that spans about 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, or 24 months.
  • the present technology also provides non-filamentous Asparagopsis taxiformis algal biomass that is produced through a cultivation cycle comprising a seedbank, nursery, and outdoor phase that spans more than about 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, or 24 months.
  • the present technology also provides non-filamentous Asparagopsis taxiformis algal biomass that is produced through a cultivation cycle comprising a seedbank, nursery, and outdoor phase that spans more than about 16, 17, 18, 19, 20, 21, 22, 23, or 24 months.
  • the present technology also provides non-filamentous Asparagopsis taxiformis algal biomass that is produced through a cultivation cycle comprising a seedbank, nursery, and outdoor phase that spans more than about 20, 21, 22, 23, or 24 months.
  • the present technology also provides non-filamentous Asparagopsis taxiformis algal biomass that is produced through a continuous cultivation cycle where free-floating tetrasporophytes comprising an acceptable concentration of bromoform may be harvested while leaving the free floating tetrasporophytes that have not achieved acceptable levels of bromoform in the bioreactor for further maturation.
  • the levels of bromoform may include levels from about 3000 to about 20000 pg/g of freeze-dried biomass. In another variation, the levels of bromoform may include levels from about 5000 to about 18000 pg/g of freeze-dried biomass. In another variation, the levels of bromoform may include levels from about 7000 to about 12000 pg/g of freeze-dried biomass.
  • the levels of bromoform may include levels from about 9000 to about 11000 pg/mg of freeze-dried biomass. In still another variation, the levels of bromoform may include levels from about 20000 to about 30000 pg/g of freeze-dried biomass. [0058]
  • the present technology also provides non-filamentous Asparagopsis taxiformis algal biomass that is produced through a cultivation cycle where the biomass density is less than about 10 g/L in the growth vessels.
  • the present technology provides for non-filamentous
  • Asparagopsis taxiformis algal biomass that is produced through a continuous cultivation cycle at a density of less than about 1 Og/1 of fresh biomass in a bioreactor for a period of between about 8 months to about 24 months yielding a bromoform content of about between about 9000 pg/g and about 20000 pg/g as measured on the freeze-dried biomass.
  • the present technology provides for non-filamentous
  • Asparagopsis taxiformis algal biomass that is produced through a continuous cultivation cycle at a density of less than about 1 Og/1 of fresh biomass in a bioreactor for a period of between about 8 months to about 24 months yielding a bromoform content of about between about 2000 pg/g and about 30000 pg/g as measured on the freeze-dried biomass.
  • the present technology relates to a method for cultivating a variety of Asparagopsis taxiformis with a higher bromoform concentration, lower odor, lower iodine and higher purity than the parent plants.
  • the features of the variety are suitable for culture in large-scale algaculture and for use as a cattle feed additive.
  • This method comprises 3 phases as shown in FIG. 5: i) Collection of parent plant; ii) Manipulation, dissection and growth in a “seed bank” room; and iii) Selection of appropriate material from seed stock.
  • wild type Asparagopsis taxiformis is collected from algal turfs or as free-floating algae in the wild.
  • dissection and growth in a “seed bank” room samples are observed and manipulated under a dissecting microscope to isolate, to the extent possible, clean filaments of AT and separate out contaminants (e.g., epiphytes, other algae, marine animals, contaminated or unhealthy AT).
  • Tiny branches are cut from the mother plant and placed in sterile well plates with seawater, each well containing 360 pL of water. These samples are maintained in a “seed bank” (a room with controlled temperature conditions, contamination protection and carefully-calibrated light with 12- hour photoperiod daily). Cultures are regularly examined. When more than doubled in size, they are stepped-up to larger sterile well plates and then again to sterilized test tubes with 30 ml seawater.
  • step of selecting appropriate material from seed stock after seven days in test tubes, material that is growing rapidly and, under magnification, appears completely free of epiphytes and fouling organisms is promoted to 250 ml flasks and moved to the nursery.
  • An additional selection step may include selecting for promotion organisms exhibiting larger than usual gland cells. From material that has not achieved those standards, tips representing new growth are cut from the material, which is returned to the smallest sterile well plates with seawater, beginning the process again.
  • the light levels and temperature within the seed bank are controlled to 10-100 mE and 65-85°F to ensure sustained growth.
  • the growing medium is supplemented with micronutrients in the form of F/2 medium.
  • material in the nursery is grown in an environment that is carefully maintained, including control of light (intensity, spectrum, photoperiod), temperature, micronutrients and aeration. Furthermore, flasks are aerated to ensure algae have sufficient supply of CO2 for photosynthesis and O2 for respiration. Aeration also promotes movement of biomass (beneficial to ensure access to light and prevent formation of biofilm).
  • the present technology provides for methods of calculating inclusion rates, methane reduction levels, and methods of intermittent feeding of Asparagopsis spp. generally, and more specifically, those disclosed in US provisional patent application 63/117,390 which is incorporated herein in its entirety.
  • Asparagopsis tends to be fed to cows regularly as part of their TMR (total mixed ration). However, recently it was found that the methane-preventing effect of Asparagopsis persisted for several days after the cows stopped eating the seaweed. This discovery opens up the possibility of maintaining the same, or similar, impact of Asparagopsis supplementation, but with intermittent rather than regular feeding. There are several possible time regimes for this new approach.
  • Red marine algae contain levels of minerals, energy and macronutrients commensurate with commonly fed grass forage type feedstuffs, such as alfalfa hay and grass hay, except its level of iodine, which is higher than common forages.
  • Milk iodine levels are directly correlated to iodine intake levels of the cow, with about 2% of consumed iodine being directly passed into the milk. If cows exceed their daily iodine intake levels significantly, their milk will exceed the recommended iodine intake levels for humans.
  • Meat iodine levels from cattle fed a diet including red marine algae at 0.5% of organic matter daily are elevated relative to cattle not fed red marine algae.
  • the concentration of iodine is inversely correlated with the concentration of target components that reduce gas emissions in red marine algae. In one aspect, this target component is bromoform.
  • a target component may be a compound selected from the set of structures illustrated in FIG 1. The concentration of iodine is generally inversely correlated with the concentration of target components that promote growth performance improvements in red marine algae.
  • Cows not fed algae typically have milk iodine levels of about 0.4 mg/kg.
  • the present investigators found that with an intermittent feeding schedule, a similar level of methane reduction can be achieved while lowering the iodine content of the milk to about 0.8 mg/kg, or less, particularly when the cows are fed the “puffball” morphotype of the Asparagopsis taxiformis or Asparagopsis armata tetrasporophyte.
  • milk from unsupplemented cows contains about 0.3 mg of iodine per kg.
  • milk from cows continuously supplemented with the gametophyte form of Asparagopsis spp. to achieve an 80% methane reduction contains about 6.5 mg of iodine per kg. This is an increase of 6.2 mg/kg.
  • intermittent feeding feeding every other day, we predict an increase of only 3.1 mg/kg, for a total of about 3.4 mg/kg. Therefore, intermittent feeding methods may also be used alone, or in combination with using algal biomass derived supplements with low iodine content to lower total iodine exposure in supplemented animals.
  • Red marine algae have complex triphasic life histories with distinct life stages with unique traits and characteristics. Additionally, we have discovered that within some of these life stages, different morphotypes can exist that are particularly effective as feed supplements, especially for promoting efficient growth and methane gas emission reduction in mammals, including ruminants.
  • red marine seaweeds with a triphasic life history generations alternate between diploid and haploid stages.
  • the mature haploid, or gametophyte, stage is characterized by a holdfast which resembles a root structure that lodges to a reef or other substrate. In the diploid stage, a tetrasporophyte forms and spreads out around the surrounding turf.
  • This morphotype that is particularly useful in the implementation of the current technology is a small red alga comprising branched chains of cells. Unlike the gametophyte form, where cells have differentiated functions (holdfast, stipe, fronds etc.), the cells in the tetrasporophyte are not highly differentiated. Instead, each cluster of four cells is roughly equivalent and these clusters string together into long chains. The color ranges from pale pink to red to dark cherry. Depending on the conditions it is cultivated in, it can take on a “puffball” form, which is distinct from the naturally occurring filamentous form mentioned in the literature.
  • Male red marine macroalgal gametophytes contain lower concentrations of the target components that promote growth performance improvement in ruminants than do female red marine macroalgae gametophytes.
  • Male red marine macroalgal gametophytes contain higher concentrations of nitrogen than do female red marine macroalgae gametophytes.
  • the cystocarps wall of the female have the highest concentration of the target components that reduce gas emissions in ruminants and the lowest concentration of iodine.
  • the cystocarps wall of the female have the highest concentration of the target components that promote growth performance improvement in ruminants and the lowest concentration of iodine.
  • the carpospores contained within the cystocarps contain virtually no bromoform.
  • the cystocarps of the female gametophytes are club-shaped structures at the tips of the gametophyte fronds. They protrude slightly from the fronds on stems. Effective inclusion of gametophyte-based biomass into animal feeds can be impacted by inadequate concentration of target component to achieve the desired outcomes at cost-effective inclusion rates.
  • the mature gametophyte life stage of red marine macroalgae both male and female, contains materially higher levels of odor triggering compounds than the tetrasporophyte life stage of the plant. It is thought that these odor triggering compounds comprise iodine or iodide comprising chemical species.
  • the present investigators elucidated how to induce the growth and indefinite maintenance of “puffball” form of the tetrasporophyte that appear to be microscopic oligocellular forms which are very morphologically and chemotypically distinct from the gametophyte macroalgal stage.
  • the present investigators have discovered that certain of these tetrasporophyte varieties in particular, certain morphotypes, are very well suited for use as feed supplements as a source of halogenated compounds to inhibit methanogenesis, promote growth of animals (e.g., ruminant animals) as well as increase the quality of products derived therefrom.
  • cultivation methods capable of achieving targeted metabolite concentrations are needed to enable effective use of a composition of algae biomass with a ratio of halogenated metabolites pg/g to iodine ppm of greater than about 150:1 at differentially lower levels than 10-30 g/day while minimizing odor and the over-supplementation of iodine.
  • the expected supplementation rates of non-filamentous Asparagopsis taxiformis tetrasporophytes is about 40 g/day, or about 20-60 g/day for dairy cows and less than about 40 g/day for beef cattle.
  • the low iodine content allows for this level of supplementation and results in methane reductions greater than can be achieved by dosing gametophytes or filamentous tetrasporophytes between 10-30 g per day.
  • the additional methane reduction is greater than 10%, 20%,
  • the current technology also includes methods of calculating intermittent dosing schedules that provide for decreased of algal to feed ratios for given methane reduction levels, increased propionate to acetate ratios, and decreased iodine concentrations in milk, meat or fat products derived from the supplemented animals.
  • Inclusion calculation methods capable of achieving targeted component concentrations are needed to enable effective use of red marine algae biomass with a ratio of target components pg/g to iodine ppm of greater than 20:1 so that the biomass can be supplemented at lower levels than 10-30 g/day thus optimizing the concentration of iodine, while enhancing the beneficial effects and minimizing capital and marginal costs.
  • Algae may contain malodorous components, here called “odor triggering components”. These odor triggering components reduce the palatability of the feed that has been supplemented with compositions derived from algal biomass or alga constituents. Therefore, it is desirable to minimize the levels of these components in the final feed either by reducing the concentration of these components in the algal derived composition, or, by enhancing the concentration of the desired bioactive components in relationship to the undesired odor triggering components, thus reducing the amount of the algal derived composition that needs to be added to the animal feed. Described herein are methods and systems of cultivating red marine algae biomass which exhibits the desired characteristics of lower odor, higher halogenated metabolite levels and lower iodine volume than whole plants harvested at the gametophyte stage.
  • Some embodiments of the present technology include tetrasporophyte life stage algaculture systems. In some implementations of these embodiments, cultivation is achieved in managed systems on land; in others, cultivation is achieved in rack systems on barges, on rafts or in shallow ocean waters. In some embodiments, processes are enabled by an apparatus and in other processes are performed by hand.
  • embodiments of the methods and systems of the subject disclosure may include establishment of high purity growth media; culture scale selection and isolation of samples for propagation; culture enhancement cycles that enable selection for robustness within target biomass; growth stage control; sustainable harvest; and preservation of halogenated metabolites.
  • generations alternate between diploid and haploid stages.
  • the mature haploid stage is characterized by a holdfast which resembles a root structure that lodges to a reef or other substrate.
  • a tetrasporophyte forms and spreads out around the surrounding turf. As it grows it takes on a form that can be dislodged from the reef, becoming a filamentous free-living organism so different from the gametophyte that it once was considered a different species.
  • tetrasporophytes are harvested from the ocean or obtained from an in-production biomass generating system.
  • tetrasporophytes grow throughout the water column and may be visible on rocks or on man-made structures including boats and docks.
  • Available samples may be given a cursory inspection for color, purity or texture, as plants that appear healthy and reasonably free of fouling organisms are selected for propagation. Samples may first be placed in bags and bags may be placed on ice during post-collection transport.
  • FIG. 5 illustrates a flow chart of a process method for culture enhancement cycles that enable step-wise improvement in the synthesis of halogenated metabolites that forces iodine concentrations lower within target biomass in the tetrasporophyte life stage.
  • the objective of the culture enhancement cycles is to effect stepwise improvement in biomass stock by selecting for robustness within target biomass to promote growth rate and halogenated metabolite accumulation, naturally lowering iodine, while removing non-target species.
  • FIG. 1 illustrates the structures of halogenated metabolites and iodine that may be found in algae. High purity starter medium is prepared for settling the tetrasporophytes into a controlled environment.
  • the water may be nutrient-rich deep ocean water, near-surface seawater, or other saltwater suitable for growing marine species. Collected water is autoclaved, then purified through ultraviolet light. Collected water is further purified through canister filtration. High purity starter medium filtered to 0.2-0.35 microns is brought to an appropriate growth temperature and nutrients are added. Collected tetrasporophytes are removed from bags and added at target densities to small vessels containing the high purity starter medium where they can acclimate to new environmental condition such as light, ambient temperature and biodiversity changes. Purified growing medium is prepared for the cultivation system for the tetrasporophytes. The water may be nutrient-rich deep ocean water, near surface seawater, or other saltwater suitable for growing marine species.
  • tetrasporophytes may be transferred to photo bioreactors on land with managed nutrients, lighting, temperature and aeration.
  • a valve-activated system may be used to release an amount of growing medium from the photobioreactor.
  • Biomass that exits the photobioreactor may be captured in a filter apparatus. Captured biomass is transferred from the Ffilter within minutes to preserve bioactive compounds.
  • Biomass may be blast frozen, then freeze dried under vacuum and temperature control for 30 hours.
  • Biomass may be dried using a vacuum tray dryer. Biomass may be dried in the open sun. Biomass may be dried in a solar conduction dryer.
  • the halogenated metabolites may be extracted from the biomass using oil extraction. The halogenated metabolites may be extracted from the biomass through fractionation.
  • biomass that has been combined into a bag or vessel at a stocking density that promotes growth while inhibiting fouling is transferred to racks on rafts or barges or upright in shallow ocean water where nutrients, lighting, and temperature are managed and aeration is enabled by a solar pump.
  • the biomass may be blast frozen, then freeze dried under vacuum and temperature control for 30 hours.
  • the biomass may be dried using a vacuum tray dryer.
  • the biomass may be dried in the open sun.
  • the biomass may be dried in a solar conduction dryer.
  • the halogenated metabolites may be extracted from the biomass using oil extraction.
  • the oil extract may be used to prepare encapsulated halogenated metabolite containing oil using the methods described herein.
  • the halogenated metabolites may be extracted from the biomass through fractionation.
  • Inclusion calculation methods capable of achieving targeted metabolite concentrations are needed to enable effective use of a composition of algae biomass with a ratio of non-iodinated halogenated metabolites mg/g to iodine ppm of greater than about 150:1 at differentially lower levels than 10-30 g/day or 20-60 g/day, thus minimizing odor and optimizing palatability.
  • Animals whose feed is supplemented with the algae biomass, or target components derived therefrom, obtained from these systems and methods provide improvements in the quantity or quality of meat, milk, manure, leather, meal, and fats in addition to reducing harmful methane emissions or example, the milk, meat and manure may have optimal nutritional iodine content and fatty acid composition. Animals fed this feed grow more quickly even on lower quality diets and produce more milk and leather.
  • the algae to be feed on an intermittent schedule is a
  • the algae is Asparagopsis armata, Asparagopsis taxiformis, Dictyota spp (e.g. Dictyota bartayresii), Oedogonium spp, Ulva spp, or C. patentiramea.
  • the algae is an Asparagopsis species.
  • the algae is Asparagopsis taxiformis.
  • the algae is Asparagopsis armata.
  • the animals supplemented with the compositions of the present technology gain about 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10% or 11% more weight than animals fed the same unsupplemented diet.
  • This weight gain difference may be average weight at slaughter or other time in the growth cycle.
  • the animals supplemented with the compositions of the present technology gain about 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20% or 21% more weight than animals fed the same unsupplemented diet.
  • This weight gain difference may be average weight at slaughter or other time in the growth cycle.
  • the animals supplemented with the compositions of the present technology grow about 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10% or 11% faster than animals fed the same unsupplemented diet. This weight gain difference between supplemented and supplemented animals may defined as average daily weight gain. In one embodiment, the animals supplemented with the compositions of the present technology grow about 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20% or 21% faster than animals fed the same unsupplemented diet. This weight gain difference between supplemented and supplemented animals may defined as average daily weight gain.
  • the animals supplemented with the compositions of the present technology provide meat or milk that is about 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10% or 11% lower in trans-fat than animals fed the same unsupplemented diet. In one embodiment, the animals supplemented with the compositions of the present technology provide meat or milk about 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20% or 21% lower in trans-fat than animals fed the same unsupplemented diet.
  • the supplemented animals require about 1%, 2%, 3%, 4%,
  • the animals supplemented with the compositions of the present technology provide meat or milk that is about 10% to 20%, 21% to 30%, 31% to 40%, 41% to 50%, 51% to 60%, 61% to 70%, 71% to 80%, 81% to 90%, 91% to 99% or 100% lower in trans fat than animals fed the same unsupplemented diet.
  • the animals supplemented with the compositions of the present technology have a propionate to acetate ratio in their rumen about 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10% or 11% greater than animals fed the same unsupplemented diet. In one embodiment, the animals supplemented with the compositions of the present technology have a propionate to acetate ratio in their rumen about 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20% or 21%greater than animals fed the same unsupplemented diet.
  • the animals supplemented with the compositions of the present technology have a propionate to acetate ratio in their rumen about 10% to 20%, 21% to 30%, 31% 40%, 41% to 50%, 51% to 60%, 61% to70%, 71% to 80%, 81% to 90%, 91% to 99% or 100% greater than animals fed the same unsupplemented diet.
  • the animals supplemented with the compositions of the present technology provide about 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10% or 11% more milk than animals fed the same unsupplemented diet. In one embodiment, the animals supplemented with the compositions of the present technology provide about 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20% or 21% more milk than animals fed the same unsupplemented diet. [0097] In one embodiment, the animals supplemented with the compositions of the present technology provide about 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30% or 31% more milk than animals fed the same unsupplemented diet.
  • the ratio of methane-reducing quality-quantity enhancing feed supplements added to animals’ normal diet is calculated by the methods, systems, and devices of the present technology.
  • animals fed the supplemented diet exhale more hydrogen than animals fed the unsupplemented diet.
  • the animals supplemented with the compositions of the present technology exhale about 10% to 20%, 21% to 30%, 31% to 40%, 41% to 50%, 51% to 60%, 61% to 70%, 71% to 80%, 81% to 90%, 91% to 99% or 100% more hydrogen than animals fed the same unsupplemented diet. In one embodiment, the animals supplemented with the compositions of the present technology exhale about 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10% or 11% more hydrogen than animals fed the same unsupplemented diet.
  • the animals supplemented with the compositions of the present technology exhale no less than about 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10% or 11% of hydrogen than animals fed the same unsupplemented diet. In one embodiment, the animals supplemented with the compositions of the present technology exhale no less than about 10% to 20%, 21% to 30%, 31% to 40%, 41% to 50%, 51% to 60%, 61% to 70%, 71% to 80%, 81% to 90%, 91% to 99% or 100% of hydrogen than animals fed the same unsupplemented diet.
  • animals fed the supplemented diet exhale less methane than animals fed the unsupplemented diet.
  • the animals supplemented with the compositions of the present technology exhale about 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10% or 11% less methane than animals fed the same unsupplemented diet. In one embodiment, the animals supplemented with the compositions of the present technology exhale about 10% to 20%, 21% to 30%, 31% to 40%, 41% to 50%, 51% to 60%, 61% to 70%, 71% to 80%, 81% to 90%, 91% to 99% or 100% less methane than animals fed the same unsupplemented diet.
  • the animals supplemented with the compositions of the present technology exhale no less than about 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10% or 11% of carbon dioxide than animals fed the same unsupplemented diet. In one embodiment, the animals supplemented with the compositions of the present technology exhale no less than about 10% to 20%, 21% to 30%, 31% to 40%, 41% to 50%, 51% to 60%, 61% to 70%, 71% to 80%, 81% to 90%, 91% to 99% or 100% of carbon dioxide than animals fed the same unsupplemented diet.
  • the present technology provides for cultivation methods capable of achieving targeted metabolite concentrations needed to enable effective use of a composition of algae biomass with a ratio of the concentration of halogenated metabolites (mg/g) to iodine (ppm) of greater than 150:1, thus allowing the inclusion of the algae biomass in animal feed at lower levels than between about 10 g/day and about 60 g/day therefore minimizing odor and the over-supplementation of iodine while maintaining the beneficial effects of reduced methane generation, faster growth, higher final body mass, fatty acid content quality, manure quality, leather quality, meat quality, and milk quality.
  • mg/g concentration of halogenated metabolites
  • ppm iodine
  • the inclusion rate may be 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, or 60 g/day on a particular day. In other embodiments, the inclusion rate may be 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, or 60 g/day averaged over 2, 3, 4, 5, 6, or 7-day period. In one embodiment the inclusion rate is about 40 g/day about every 48 h.
  • the algal supplement is administered every 1.5 days, every 2 days, every 3 days or every 4.5 days. In still another embodiment, the algal supplement is administered every 7 days.
  • the inclusion rate is determined by an algorithm in which one input is the amount of neutral detergent fiber (NDF) in the animal’s diet. In another embodiment, the amount of neutral detergent fiber in an animal’s diet is determined by multiplying the animal’s dry matter intake by the percentage of neutral detergent fiber in the dry matter that is fed to the animal.
  • NDF neutral detergent fiber
  • a regression constant is determined.
  • the regression constant relates the amount of bromoform required to achieve a set percentage of methane reduction per amount of neutral detergent fiber present in the animal’s diet.
  • the algorithm comprises the steps of: i) Determining the target methane reduction percentage and subsequently the target absolute reduction (g CH4/ kg milk); ii) Determining cow's NDF intake, based on dry matter intake (DMI) and NDF proportion; iii) Determining the normalized bromoform concentration required (equivalent to the desired absolute reduction divided by the regression constant, where the regression constant is the Reduction in methane intensity (g CH4 / kg milk) per unit normalized bromoform intake (mg / kg of NDF); iv) Multiplying by the NDF intake to determine the required bromoform concentration (mg) and v) Dividing by the bromoform concentration in that seaweed to determine amount of seaweed required.
  • the present technology provides for cultivation methods capable of achieving targeted metabolite concentrations are needed to enable effective use of a composition of algae biomass with a ratio of the concentration of non-iodinated halogenated metabolites (mg/g) to iodine (ppm) of greater than 150:1, thus allowing the inclusion of the algae biomass in animal feed at lower levels than between about 10 g/day and about 30 g/day therefore minimizing odor and the over-supplementation of iodine while maintaining the beneficial effects of reduced methane generation, faster growth, higher final body mass, fatty acid content quality, manure quality, leather quality, meat quality, and milk quality.
  • mg/g non-iodinated halogenated metabolites
  • ppm iodine
  • the cultivation method of the present technology provides a composition comprising algae biomass with a ratio of the concentration of halogenated metabolites (pg/g) to iodine (ppm) of greater than 1:1. 2:1, 3:1, 4:1, 5:1, 6:1, 7:1, 8:1, 9:1, 10:1, 11:1, 12:1, 13:1, 14:1, 15:1, 16:1, 17:1, 18:1, or 19:1.
  • the cultivation method of the present technology provides a composition comprising algae biomass with a ratio of the concentration of non-iodinated halogenated metabolites (pg/g) to iodine (ppm) of greater than 1:1.
  • the cultivation method of the present technology provides a composition comprising algae biomass with a ratio of the concentration of halogenated metabolites (pg/g) to iodine (ppm) of greater than 21:1. 22:1, 23:1, 24:1, 25:1, 26:1, 27:1, 28:1, 29:1, 30:1, 31:1, 32:1, 33:1, 34:1, 35:1, 36:1, 37:1, 38:1, or 39:1.
  • pg/g concentration of halogenated metabolites
  • ppm iodine
  • the cultivation method of the present technology provides a composition comprising algae biomass with a ratio of the concentration of non-iodinated halogenated metabolites (pg/g) to iodine (ppm) of greater than 21:1. 22:1, 23:1, 24:1, 25:1, 26:1, 27:1, 28:1, 29:1, 30:1, 31:1, 32:1, 33:1, 34:1, 35:1, 36:1, 37:1, 38:1, or 39:1. [00110] In one embodiment, the cultivation method of the present technology provides a composition comprising algae biomass with a ratio of the concentration of bromoform (pg/g) to iodine (ppm) of greater than 20:1.
  • the cultivation method of the present technology provides a composition comprising algae biomass with a ratio of the concentration of bromoform (pg/g) to iodine (ppm) of greater than 1:1. 2:1, 3:1, 4:1, 5:1, 6:1, 7:1, 8:1, 9:1, 10:1, 11:1, 12:1, 13:1, 14:1, 15:1, 16:1, 17:1, 18:1, or 19:1.
  • the cultivation method of the present technology provides a composition comprising algae biomass with a ratio of the concentration of bromoform (pg/g) to iodine (ppm) of greater than 21:1.
  • the cultivation method of the present technology provides a composition comprising algae biomass with a ratio of the concentration of bromoform (pg/g) to iodine (ppm) of equal to or greater than 700:1, 600:1, 500:1, 400:1, 300:1 200:1, 150:1, 140:1, 130:1, 120:1, 100:1, 75:1, 50:1, 25:1, or 20:1.
  • the algae biomass obtained by the methods of the present technology has a ratio of concentration of bromoform (pg/g) to iodine (ppm) of greater than 140.
  • the present disclosure provides methods of estimating average concentration of target components, the range of component concentrations within the sample, or trending of component concentration levels over time.
  • the methods include measurement or manipulation of the intensity or wavelength distribution spectrum of the light source, measurement or manipulation of the sample turbidity in an image-based target component measurement system, or photobioreactor, or other algae growth vessel.
  • the methods include computer vision-based feature recognition and neural network based optical image analysis.
  • the present disclosure provides for methods for triggering alerts or actions based on positive or negative developments of the measurement of target components within the microalgae or algae sample.
  • the methods include triggering a cultivation harvesting cycle, adjustment of environmental conditions, or request for personnel intervention.
  • target component includes halogenated metabolites, iodine, bromoform or other components of pre-harvest, post-harvest, or post processed algae biomass that are determined to affect the function of the derived composition as an animal feed supplement.
  • the halogenated metabolites include iodine containing compounds, but not elemental iodine.
  • the non-iodinated halogenated metabolites exclude iodine containing compounds and elemental iodine.
  • the iodine content comprises any target component that contains iodine, including elemental iodine, organic iodine and inorganic iodine.
  • the expression “inorganic iodine” means iodide anions, salts, hypoiodites and the like. In one embodiment, the expression “organic iodine” refers to any compound comprising at least one iodine atom bound to at least one carbon atom.
  • the target components may be metabolite components contained within algae, microalgae, or algae.
  • Target component includes, but is not be limited to: primary metabolites, secondary metabolites, substances absorbed or concentrated from the environment, substances present due to the actions of parasites or symbiotes, substances formed due to environmental factors, substances formed through the actions of electromagnetic radiation; acoustic energy; fermentation by bacteria, yeasts or other organisms; oxidation, dehydration, elimination, hydration, decarboxylation, isomerization, racemization, chelation, inclusion, fragmentation; or substances that are substrates for the actions of electromagnetic radiation; acoustic energy; fermentation by bacteria, yeasts or other organisms; oxidation, dehydration, elimination, hydration, decarboxylation, isomerization, racemization, chelation, inclusion, fragmentation.
  • a target component is bromoform. In one embodiment, a target component is a halogenated metabolite. In one embodiment, a target component is a non-iodinated metabolite. In another embodiment, a target component is elemental iodine (I2). In another embodiment, a target component is organic iodine. In another embodiment, a target component is inorganic iodine. In another embodiment, a target component is a substance that liberates elemental iodine post-harvest of the algae biomass. A representative set of target components is shown in FIG. 1.
  • bromoform (compound 1) is a major contributor to the methane reduction and animal product improvement results of seaweed supplementation.
  • level may mean an absolute level, an amount over time, or a concentration, depending on context.
  • level of consumption the term “levels” should be construed as an amount eaten by an animal over a set period of time, such as a day, a week, month, etc.
  • level means the concentration of that component in a particular weight or volume of biomass. In the last case, the level may be determined on living biomass in a photoreactor, wet harvested biomass, dried biomass, or biomass at each stage of processing from start to the final commercial product.
  • the inclusion calculations can take into account the addition of other, non-algae derived components that provide additive or synergistic beneficial effects when combined with the compositions described herein.
  • the non-algae derived components are selected from the group consisting of; 3-nitrooxypropanol, Mootral (a product of Mootral, a Swiss Agritech company), garlic extract, Yucca extract, Yucca powder, saponin, furostanol aglycone, spirostanol aglycone, chloroform, sarsapogenin, markogenin, smilagenin, samogenin, gitogenin, neogitogenin, monodesmosidic saponins, YS-I, YS-II, YS-III, YS-IV, YS- V, YS-VI, YS-VII, YS-VIII, YS-IX, YS-
  • compositions of the present technology may be used in combination with other methane-reducing, quality and quantity enhancing components as disclosed in A. Cieslak, M. Szumacher-Strabel, A. Stochmal and W. Oleszek, Animal (2013), 7:s2, pp 253-265 & The Animal Consortium 2013, doi: 10.1017/S1751731113000852, which is incorporated herein in its entirety.
  • Red algae in contrast to green and brown algae, produce a broad set of halogenated metabolites including peptides, polyketides, indoles, terpenes, phenols and volatile halogenated hydrocarbons.
  • halogenated hydrocarbon Bromoform CHBr3.
  • concentrations of Bromoform and other halogenated compounds present in Asparagopsis taxiformis and other red algae have been shown to vary widely based on the growth environment, seasonality, species, strain, lifestage, cultivation method, and other known and unknown factors.
  • In vivo testing has identified a strong positive correlation between the level of methane reduction in ruminants and the ratio of the bromoform component delivered from a red algae feed supplement, relative to specific components of the animal’s diet.
  • In vivo testing has identified a predictive relationship among the concentration of bromoform in algae, neutral detergent fiber (NDF) in livestock diets, and the percentage of enteric methane reduced.
  • NDF neutral detergent fiber
  • bromoform is safely and usefully degraded in anaerobic environments like livestock rumens where the enzyme methyl-coenzyme M reductase is present.
  • the degradation of the bromoform component delivered from red algae beneficially increases the propionate:acetate ratios in livestock rumens, which enables conservation of feed energy.
  • Described herein is a system and method that calculates and accounts for the enteric methane emissions that result from livestock which consume red macroalgae.
  • the components of this system and method include a knowledge base of the interrelationships among variables related to red macroalgae, inclusion rates of red macroalgae, and livestock diets, as well as the combinatorial impact of these variables on livestock enteric methane emissions; an algorithm that calculates enteric methane emissions based on the interrelationships among variables related to red macroalgae, inclusion rates of red macroalgae, and livestock diets; and a system that accounts for enteric methane emissions reductions that result from livestock which consume red macroalgae.
  • the system calculates the enteric methane reductions that result from a certain inclusion rate of red macroalgae in livestock feed and supplements.
  • the system documents, accounts, tracks and verifies enteric methane emissions from livestock which consume red macroalgae.
  • Further embodiments include integration with digital or analogue processes that document, account, track and verify methane emissions reductions from livestock which consume red macroalgae.
  • processes that document, account, track, and verify enteric methane emissions reductions from livestock which consume red macroalgae operate dependently on other, existing systems. In other embodiments they operate independently of other, existing systems.
  • the system is accessed by a graphical user interface.
  • the system is accessed by an application programming interface. Further embodiments enable and allow users to enter and manipulate input data and objective functions, and to observe the results of an algorithmic calculation based on those inputs.
  • FIG. 10 Shown in FIG. 10 is an embodiment of a system (1) of the present technology that calculates and accounts for the enteric methane emissions reductions that result from livestock which consume red macroalgae.
  • the system includes a knowledge base and algorithm (11) that calculates the enteric methane reductions that result from a certain inclusion rate of red macroalgae in livestock feed and supplements.
  • This knowledge base and algorithm (11) is programmed into computer software (12) which resides and is accessed on a mainframe computer server (13).
  • a database (16) which resides on the mainframe computer (13) documents, accounts, tracks, and verifies enteric methane emissions from livestock which consume red macroalgae.
  • system users may input data (14) into the database (16) through a unique graphical interface (15) or through an existing software program (18) that is capable of electronic data interchange (17).
  • users of the system receive a unique identification number associated with their data.
  • this unique identification number is used for verifying and tracking livestock enteric methane emissions reductions.
  • Described herein is a system and method that calculates precise inclusion rates of red algae in livestock feed and supplements. This system and method accounts for variables related to red algae and livestock diets in ways that deliver specific intended biological impacts and outcomes to livestock and their byproducts.
  • the system also comprises supplementation schedules which may be intermittent or continuous.
  • intermittent feeding is where the variation in feeding is done on a daily or weekly timescale. For example, one might imagine feeding Asparagopsis in the morning TMR but not the evening, or on weekdays but not weekends. This could yield a number of the benefits above, for example reducing labor. Note that, though the actual feeding of the Asparagopsis taxiformis (AT) is done on sub- week timescales, this will likely be part of a feeding regimen lasting a couple weeks or more.
  • Asparagopsis taxiformis AT
  • intermittent feeding is where the variation in feeding is done on a period of longer than a week. For example, one might imagine feeding Asparagopsis during lactation but not during pregnancy for dairy cows or removing beef steers from the feed additive two weeks before harvest.
  • feeding ruminants a higher dose of Asparagopsis at the beginning of the dosing period tapering to a lower dose towards the end of the dosing period. This could be useful, for example, to “kickstart” the benefits of AT.
  • feeding ruminants a lower dose of Asparagopsis at the beginning of the dosing period increasing to a higher dose at the end of the dosing period. This could be useful, for example, to maintain or elongate the benefits of AT.
  • the current technology is used to alter the dose of AT based on the concentration of the active ingredient (bromoform) within the seaweed.
  • the current technology provides for a more efficacious use of AT as calculated by the methane reduction per gram of AT composition fed per animal.
  • the increase of efficacy of the amount of methane emissions reduced per gram of the AT composition is at least about 5%, 10%, 20%, 30%, 40%, 50%, 75%, 100%, 150%, 200% or greater than 200%.
  • the method of feeding ruminant animals a time- varying dose of AT composition is a time- varying dose of AT composition.
  • a time varying dose of AT composition is a supplementation schedule describing the regularly occurring intervals and amounts of feeding the AT composition.
  • the AT composition is administered once every 48 h
  • the AT composition is administered every 72 h.
  • a time varying dose of AT composition is a supplementation schedule describing the supplementation time window and dose of the AT composition based on discrete events such as reproductive status or time to market, or other such events that would limit the amount of iodine or halogenated organic materials allowable in the animal product or animal.
  • time varying dose supplementation schedule time window is adjusted to comport with animal feed regulations or consumer perception.
  • the current technology allows for the more flexible crew scheduling since fewer crew members would be required to be familiar with the AT composition administration.
  • the current technology provides for healthier animals and their offspring as compared to unsupplemented or continuously supplemented animals.
  • the current technology provides for higher quality animal products compared to unsupplemented or continuously supplemented animals.
  • the feed supplement compositions of the present technology do not comprise sweeteners, such as, but not limited to, molasses, high fructose corn syrup, sucrose, fructose, xylitol, sorbitol, or other sugars or alcohol sugar appetants.
  • the feed supplement compositions of the present technology may comprise sweeteners, such as, but not limited to, molasses, high fructose corn syrup, sucrose, fructose, xylitol, sorbitol, or other sugars or alcohol sugar appetants.
  • sweeteners such as, but not limited to, molasses, high fructose corn syrup, sucrose, fructose, xylitol, sorbitol, or other sugars or alcohol sugar appetants.
  • the feed supplement compositions of the present technology may be used to supplement feedlot animals on finishing diets at a daily supplementation rate of less than 200 g/day, less than about 150 g/day, less than about 100 g/day, or about or less than about 50 g/day of the algal biomass described here for animals on finishing diets.
  • the components of this system and method include a knowledge base of the interrelationships among variables related to red algae, variables related to inclusion rates of red algae, variables related to livestock diets, as well as the combinatorial impact of these variables on specific biological outcomes to livestock and their byproducts; an algorithm that calculates and predicts specific biological impact and outcomes to livestock and their byproducts based on the interrelationships among variables related to red algae, inclusion rates of red algae, and livestock diets; and guidance for persons skilled in the art of formulating livestock feed and supplements that achieves specific and intended biological impact and outcomes to livestock and their byproducts.
  • the system calculates and predicts the enteric methane reductions that result from a certain inclusion rate of red algae in livestock feed and supplements. [00159] In one embodiment, the system calculates and predicts specific improvements in livestock feed conversion efficiency ratios, or improvements in livestock productivity gains and outputs that result from a certain inclusion rate of red algae in livestock feed and supplements. [00160] In one embodiment, the system calculates and predicts propionate:to acetate ratios in livestock rumens that result from a certain inclusion rate of red algae in livestock feed and supplements.
  • the system calculates and predicts valuable attributes in the byproducts of livestock that result from a certain inclusion rate of red algae in livestock feed and supplements. [00162] In a further embodiment, the system calculates and predicts valuable attributes in the meat and milk produced by livestock which have consumed a certain inclusion rate of red algae in their feed and supplements.
  • the system calculates and predicts economically optimal inclusion rates of red algae in livestock feed and supplements.
  • the system calculates and formulates livestock feed and supplement rations in which red algae is added. Further embodiments include integration with digital or analogue processes that calculate and formulate livestock feed and supplement rations. In some embodiments, calculations and formulations operate dependently on other existing systems. In other embodiments they operate independently of other, existing systems.
  • the system is accessed by a graphical user interface.
  • the system is accessed by an application programming interface. Further embodiments enable and allow users skilled in the art of formulating livestock feed and supplements to enter and manipulate input data and objective functions, and to observe the results of an algorithmic calculation based on those inputs.
  • the system is used by commercial feed mills.
  • the system is used in on-farm feeding operations.
  • FIG. 11 Shown in FIG. 11 is an embodiment of a system (1) that calculates precise inclusion rates of red algae in livestock feed and supplements.
  • the system includes a knowledge base and algorithm (11) that calculates and predicts specific biological impact and outcomes to livestock and/or their byproducts based on the interrelationships among variables related to red algae, inclusion rates of red algae, and livestock diets.
  • This knowledge base and algorithm (11) is programmed into computer software (12) which resides and is accessed on a mainframe computer server (13).
  • the knowledge base and algorithm provides instructions (14) to persons skilled in the art of formulating livestock feed and supplements for incorporating red algae into livestock feed and supplements.
  • instructions are based on common characteristics related to species, breed, gender, age, and stage of reproductive cycle, as well as predetermined objective functions related to specific intended biological impact and outcomes to livestock which possess these common characteristics and/or the byproducts produced by livestock which possess these common characteristics.
  • a unique graphical user interface allows users skilled in the art of formulating livestock feed to enter and manipulate input data and objective functions, and to observe the results (16) rendered by the knowledge based and algorithm (11).
  • the knowledge base and algorithm are accessed by an application programming interface (17) and the observed results are rendered and observed within an existing software program (18).
  • FIG. 1 Shown in FIG. 1, are exemplary structures of the halogenated metabolites and other halogen containing compounds that are a subset of the target compounds whose levels are an input into the methods of the present technology that calculate their levels of inclusion in the feed supplements and supplemented feeds of the present technology.
  • the current technology also includes novel varieties of AT that are particularly suitable for intermittent feeding.
  • compositions of the current technology comprise non- gametophyte derived algal biomass exhibiting a bromoform content of more than 2.5, 2.4, 2.3, 2.2, 2.1, 2.0, 1.9, 1.8, 1.7, 1.6, 1.5, 1.4, 1.3, 1.2, 1.1, 1.0, 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, 0.3, 0.2, or 0.1 w/w% dry weight.
  • compositions of the current technology comprise non- gametophyte derived algal biomass exhibiting a bromoform content of more than 1.8 w/w% dry weight.
  • compositions of the current technology comprise non- gametophyte derived algal biomass exhibiting an iodine content of less than 0.1, 0.09, 0.08, 0.07, 0.06, 0.05, 0.04, 0.03, 0.02 or 0.01 w/w% dry weight.
  • compositions of the current technology comprise non- gametophyte derived algal biomass exhibiting an iodine content of less than 0.145 w/w% dry weight.
  • compositions of the current technology comprise algal biomass exhibiting an iodine to bromoform ratio of less than 0.1, 0.09, 0.08, 0.07, 0.06, 0.05, 0.04, 0.03, 0.02, 0.012 or 0.01.
  • compositions of the current technology comprise algal biomass exhibiting an iodine to bromoform ratio of less than 0.04, 0.03, 0.02. 0.012 or 0.01. [00178] Even more preferably, the compositions of the current technology comprise algal biomass exhibiting an iodine to bromoform ratio of less than 0.012.
  • the iodine content of the algal biomass may comprise elemental iodine, organic iodine compounds, inorganic iodine compounds, iodide, iodate or periodate, or a combination thereof.
  • the iodine content of the algal biomass may liberate elemental iodine upon harvesting, processing, or storage.
  • the current technology provides for a method of reducing methane reductions from ruminants by at least 80% by supplementing the food rations of such ruminants with non-gametophyte derived algal biomass exhibiting a bromoform content of more than 2.5, 2.4, 2.3, 2.2, .2.0, 1.9, 1.8, 1.7, 1.6, 1.5, 1.4, 1.3, 1.2, 1.1, 1.0, 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, 0.3, 0.2, or 0.1 w/w% dry weight.
  • the current technology provides for a method of reducing methane reductions from ruminants by at least 80% by supplementing the food rations of such ruminants with non-gametophyte derived algal biomass exhibiting an iodine content of less than 0.1, 0.09, 0.08, 0.07, 0.06, 0.05, 0.04, 0.03, 0.02 or 0.01 w/w%
  • the current technology provides for a method of reducing methane reductions from ruminants by at least 80% by supplementing the food rations of such ruminants with algal biomass exhibiting an iodine to bromoform ratio of less than 0.1, 0.09, 0.08, 0.07, 0.06, 0.05, 0.04, 0.03, 0.02, 0.012 or 0.01.
  • the current technology provides for a method of reducing methane reductions from ruminants by at least 80% by supplementing the food rations of such ruminants with algal biomass exhibiting an iodine to bromoform ratio of less than 0.04, 0.03, 0.02, 0.012 or 001
  • the current technology provides for a method of reducing methane production from ruminants by at least 80% by supplementing the food rations of such ruminants with algal biomass such that the ruminant consumes less than 50 mg of iodine per 1 kg of dry matter intake.
  • the current technology provides for a method of reducing methane production from ruminants by at least 80% by supplementing the food rations of such ruminants with algal biomass such that the ruminant consumes less than 40 mg of iodine per 1 kg of dry matter intake.
  • the current technology provides for a method of reducing methane production from ruminants by at least 80% by supplementing the food rations of such ruminants with algal biomass such that the ruminant consumes less than 30 mg of iodine per 1 kg of dry matter intake.
  • the current technology provides for a method of reducing methane production from ruminants by at least 80% by supplementing the food rations of such ruminants with algal biomass such that the ruminant consumes less than 20 mg of iodine per 1 kg of dry matter intake.
  • the current technology provides for a method of reducing methane production from ruminants by at least 80% by supplementing the food rations of such ruminants with algal biomass such that the ruminant consumes less than 10 mg of iodine per 1 kg of dry matter intake.
  • the current technology provides for a method of reducing methane production from ruminants by at least 80% by supplementing the food rations of such ruminants with algal biomass such that the ruminant consumes less than 5 mg of iodine per 1 kg of dry matter intake.
  • the current technology provides for a method of reducing methane production from ruminants by at least 80% by supplementing the food rations of such ruminants with algal biomass such that the ruminant consumes less than 4 mg of iodine per 1 kg of dry matter intake.
  • the current technology provides for a method of reducing methane reductions from ruminants by at least 80% by supplementing the food rations of such ruminants with algal biomass such that the ruminant consumes less than 3 mg of iodine per 1 kg of dry matter intake.
  • the current technology provides for a method of reducing methane production from ruminants by at least 80% by supplementing the food rations of such ruminants with algal biomass such that the ruminant consumes less than 2 mg of iodine per 1 kg of dry matter intake.
  • the current technology provides for a method of reducing methane production from ruminants by at least 80% by supplementing the food rations of such ruminants with algal biomass such that the ruminant consumes less than 1 mg of iodine per 1 kg of dry matter intake.
  • the current technology provides for a method of reducing methane production from ruminants by at least 70% by supplementing the food rations of such ruminants with non-gametophyte derived algal biomass exhibiting a bromoform content of more than 2.5, 2.4, 2.3, 2.2, .2.0, 1.9, 1.8, 1.7, 1.6, 1.5, 1.4, 1.3, 1.2, 1.1, 1.0, 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, 0.3, 0.2, or 0.1 w/w% dry weight.
  • the current technology provides for a method of reducing methane production from ruminants by at least 70% by supplementing the food rations of such ruminants with non-gametophyte derived algal biomass exhibiting an iodine content of less than 0.1, 0.09, 0.08, 0.07, 0.06, 0.05, 0.04, 0.03, 0.02 or 0.01 w/w%
  • the current technology provides for a method of reducing methane production from ruminants by at least 70% by supplementing the food rations of such ruminants with algal biomass exhibiting an iodine to bromoform ratio of less than 0.1, 0.09, 0.08, 0.07, 0.06, 0.05, 0.04, 0.03, 0.02, 0.012 or 0.01.
  • the current technology provides for a method of reducing methane production from ruminants by at least 70% by supplementing the food rations of such ruminants with algal biomass exhibiting an iodine to bromoform ratio of less than 0.04, 0.03, 0.02, 0.012 or 001
  • the current technology provides for a method of reducing methane production from ruminants by at least 70% by supplementing the food rations of such ruminants with algal biomass such that the ruminant consumes less than 50 mg of iodine per 1 kg of dry matter intake.
  • the current technology provides for a method of reducing methane production from ruminants by at least 70% by supplementing the food rations of such ruminants with algal biomass such that the ruminant consumes less than 40 mg of iodine per 1 kg of dry matter intake.
  • the current technology provides for a method of reducing methane production from ruminants by at least 70% by supplementing the food rations of such ruminants with algal biomass such that the ruminant consumes less than 30 mg of iodine per 1 kg of dry matter intake.
  • the current technology provides for a method of reducing methane production from ruminants by at least 70% by supplementing the food rations of such ruminants with algal biomass such that the ruminant consumes less than 20 mg of iodine per 1 kg of dry matter intake.
  • the current technology provides for a method of reducing methane production from ruminants by at least 70% by supplementing the food rations of such ruminants with algal biomass such that the ruminant consumes less than 10 mg of iodine per 1 kg of dry matter intake.
  • the current technology provides for a method of reducing methane production from ruminants by at least 70% by supplementing the food rations of such ruminants with algal biomass such that the ruminant consumes less than 5 mg of iodine per 1 kg of dry matter intake.
  • the current technology provides for a method of reducing methane production from ruminants by at least 70% by supplementing the food rations of such ruminants with algal biomass such that the ruminant consumes less than 4 mg of iodine per 1 kg of dry matter intake.
  • the current technology provides for a method of reducing methane production from ruminants by at least 70% by supplementing the food rations of such ruminants with algal biomass such that the ruminant consumes less than 3 mg of iodine per 1 kg of dry matter intake.
  • the current technology provides for a method of reducing methane production from ruminants by at least 70% by supplementing the food rations of such ruminants with algal biomass such that the ruminant consumes less than 2 mg of iodine per 1 kg of dry matter intake.
  • the current technology provides for a method of reducing methane production from ruminants by at least 70% by supplementing the food rations of such ruminants with algal biomass such that the ruminant consumes less than 1 mg of iodine per 1 kg of dry matter intake.
  • the current technology provides for a method of reducing methane production from ruminants by at least 85% by supplementing the food rations of such ruminants with non-gametophyte derived algal biomass exhibiting a bromoform content of more than 2.5, 2.4, 2.3, 2.2, .2.0, 1.9, 1.8, 1.7, 1.6, 1.5, 1.4, 1.3, 1.2, 1.1, 1.0, 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, 0.3, 0.2, or 0.1 w/w% dry weight.
  • the current technology provides for a method of reducing methane production from ruminants by at least 85% by supplementing the food rations of such ruminants with non-gametophyte derived algal biomass exhibiting an iodine content of less than 0.1, 0.09, 0.08, 0.07, 0.06, 0.05, 0.04, 0.03, 0.02 or 0.01 w/w%
  • the current technology provides for a method of reducing methane production from ruminants by at least 85% by supplementing the food rations of such ruminants with algal biomass exhibiting an iodine to bromoform ratio of less than 0.1, 0.09, 0.08, 0.07, 0.06, 0.05, 0.04, 0.03, 0.02, 0.012 or 0.01.
  • the current technology provides for a method of reducing methane production from ruminants by at least 85% by supplementing the food rations of such ruminants with algal biomass exhibiting an iodine to bromoform ratio of less than 0.04, 0.03, 0.02, 0.012 or 001
  • the current technology provides for a method of reducing methane production from ruminants by at least 85% by supplementing the food rations of such ruminants with algal biomass such that the ruminant consumes less than 50 mg of iodine per 1 kg of dry matter intake.
  • the current technology provides for a method of reducing methane production from ruminants by at least 85% by supplementing the food rations of such ruminants with algal biomass such that the ruminant consumes less than 40 mg of iodine per 1 kg of dry matter intake.
  • the current technology provides for a method of reducing methane production from ruminants by at least 85% by supplementing the food rations of such ruminants with algal biomass such that the ruminant consumes less than 30 mg of iodine per 1 kg of dry matter intake.
  • the current technology provides for a method of reducing methane production from ruminants by at least 85% by supplementing the food rations of such ruminants with algal biomass such that the ruminant consumes less than 20 mg of iodine per 1 kg of dry matter intake.
  • the current technology provides for a method of reducing methane production from ruminants by at least 85% by supplementing the food rations of such ruminants with algal biomass such that the ruminant consumes less than 10 mg of iodine per 1 kg of dry matter intake.
  • the current technology provides for a method of reducing methane production from ruminants by at least 85% by supplementing the food rations of such ruminants with algal biomass such that the ruminant consumes less than 5 mg of iodine per 1 kg of dry matter intake.
  • the current technology provides for a method of reducing methane production from ruminants by at least 85% by supplementing the food rations of such ruminants with algal biomass such that the ruminant consumes less than 4 mg of iodine per 1 kg of dry matter intake.
  • the current technology provides for a method of reducing methane production from ruminants by at least 85% by supplementing the food rations of such ruminants with algal biomass such that the ruminant consumes less than 3 mg of iodine per 1 kg of dry matter intake.
  • the current technology provides for a method of reducing methane production from ruminants by at least 85% by supplementing the food rations of such ruminants with algal biomass such that the ruminant consumes less than 2 mg of iodine per 1 kg of dry matter intake.
  • the current technology provides for a method of reducing methane production from ruminants by at least 85% by supplementing the food rations of such ruminants with algal biomass such that the ruminant consumes less than 1 mg of iodine per 1 kg of dry matter intake.
  • the present technology provides for an algal feed supplement for beef cattle, dairy cattle and other ruminants comprising a maximum of 3000 ppm of iodine by dry weight.
  • the present technology provides for an algal feed supplement for beef cattle, dairy cattle and other ruminants comprising a maximum of 2000 ppm of iodine by dry weight.
  • the present technology provides for an algal feed supplement for beef cattle, dairy cattle and other ruminants comprising a maximum of 1000 ppm of iodine by dry weight.
  • the present technology provides for an algal feed supplement for beef cattle, dairy cattle and other ruminants comprising a maximum of 500 ppm of iodine by dry weight.
  • the present technology provides for an algal feed supplement for beef cattle, dairy cattle and other ruminants comprising a minimum of 2.5% of bromoform by dry weight.
  • the present technology provides for an algal feed supplement for beef cattle, dairy cattle and other ruminants comprising a minimum of 3.5% of bromoform by dry weight.
  • the present technology provides for an algal feed supplement for beef cattle, dairy cattle and other ruminants comprising a minimum of 16% of protein by dry weight.
  • the present technology provides for an algal feed supplement for beef cattle, dairy cattle and other ruminants comprising a minimum of 20% of aNDF by dry weight.
  • the present technology provides for an algal feed supplement for beef cattle, dairy cattle and other ruminants comprising a minimum of 20% of aNDF by dry weight, a minimum of 16% protein by dry weight, a maximum of 3000 ppm iodine by dry weight, and a minimum of 2.5% bromoform by dry weight.
  • the present technology provides for a method of reducing methane belching by ruminants comprising supplementing the feed of such ruminants with an algal feed supplement comprising a minimum of 20% of aNDF by dry weight, a minimum of 16% protein by dry weight, a maximum of 3000 ppm iodine by dry weight, and a minimum of 2.5% bromoform by dry weight.
  • the present technology provides for a method of reducing methane belching by ruminants by at least 50% comprising supplementing the feed of such ruminants with an algal feed supplement comprising a minimum of 20% of aNDF by dry weight, a minimum of 16% protein by dry weight, a maximum of 3000 ppm iodine by dry weight, and a minimum of 2.5% bromoform by dry weight.
  • the present technology provides for a method of reducing methane belching by ruminants by at least 80% comprising supplementing the feed of such ruminants with an algal feed supplement comprising a minimum of 20% of aNDF by dry weight, a minimum of 16% protein by dry weight, a maximum of 3000 ppm iodine by dry weight, and a minimum of 2.5% bromoform by dry weight at an average rate of 40 g .per day.
  • the present technology provides for a kit comprising an algal feed supplement comprising a minimum of 20% of aNDF by dry weight, a minimum of 16% protein by dry weight, a maximum of 3000 ppm iodine by dry weight, and a minimum of 2.5% bromoform by dry weight accompanied by instructions for substituting the algal feed supplement for total mixed ration (TMR) up to about 0.5% as fed.
  • TMR total mixed ration
  • the present technology provides for a kit comprising an algal feed supplement comprising a minimum of 20% of aNDF by dry weight, a minimum of 16% protein by dry weight, a maximum of 3000 ppm iodine by dry weight, and a minimum of 2.5% bromoform by dry weight accompanied by instructions for supplementing the total mixed ration with about 6.5 g of product per kg of neutral dietary fiber (aNDF).
  • an algal feed supplement comprising a minimum of 20% of aNDF by dry weight, a minimum of 16% protein by dry weight, a maximum of 3000 ppm iodine by dry weight, and a minimum of 2.5% bromoform by dry weight accompanied by instructions for supplementing the total mixed ration with about 6.5 g of product per kg of neutral dietary fiber (aNDF).
  • the present technology provides for a kit comprising an algal feed supplement comprising a minimum of 20% of aNDF by dry weight, a minimum of 16% protein by dry weight, a maximum of 3000 ppm iodine by dry weight, and a minimum of 2.5% bromoform by dry weight accompanied by instructions for supplementing the total mixed ration from about 4.5 g to 8.5 g of product per kg of neutral dietary fiber (aNDF).
  • aNDF neutral dietary fiber
  • compositions and methods of the present technology provide for the use of manure from Asparagopsis spp.., Asparagopsis taxiformis, or a composition comprising Asparagopsis taxiformis brominata treated ruminants to produce methane using a manure digester to obtain improved yields of methane than those obtained from the manure of untreated cows
  • compositions and methods of the present technology provide for the use of manure from Asparagopsis spp., Asparagopsis taxiformis, or a composition comprising Asparagopsis taxiformis brominata treated ruminants to produce methane using covered manure lagoon to obtain improved yields of methane than those obtained from the manure of untreated cows.
  • the present technology also offers for either treating the manure itself with the algal biomass compositions described herein, other antimethanogenic compounds described herein, or combinations thereof.
  • This method of treating the manure with the compositions and methods of the present technology may be performed on the manure from animals supplemented with the compositions and methods of the present technology or on the manure from untreated animals.
  • compositions and methods of the present technology provide for the treatment of ruminant manure with a composition comprising about 1 g/ton, lOg/ton, 50g/ton, lOOg/ton, 500g/ton, 1 kg/ton, or 10 kg/ton of Asparagopsis spp., Asparagopsis taxiformis, or a composition comprising Asparagopsis taxiformis brominata.
  • compositions and methods of the present technology provide for the treatment of ruminant manure with a composition comprising about 1 g,/ton, lOg/ton, 50g/ton, lOOg/ton, 500g/ton, 1 kg/ton, or 10 kg/ton of bromoform, encapsulated bromoform, or a compound or element selected from the compounds or elements depicted in Figure 2.
  • compositions and methods of the present technology provide for the use of manure from Asparagopsis spp.., Asparagopsis taxiformis, or a composition comprising Asparagopsis taxiformis brominate treated ruminants to produce methane using a manure digester to obtain improved yields of methane than those obtained from the manure of untreated cows
  • compositions and methods of the present technology provide for the use of manure from Asparagopsis spp.., Asparagopsis taxiformis, or a composition comprising Asparagopsis taxiformis brominata treated ruminants to produce methane using covered manure lagoon to obtain improved yields of methane than those obtained from the manure of untreated cows
  • the composition used to treat the manure may comprise one or more compounds selected from the group consisting of: 3-nitrooxypropanol, Mootral (a product of Mootral, a Swiss Agritech company), garlic extract, Yucca extract, Yucca powder, saponin, furostanol aglycone, spirostanol aglycone, chloroform, sarsapogenin, markogenin, smilagenin, samogenin, gitogenin, neogitogenin, monodesmosidic saponins, YS-I, YS-II, YS-III, YS-IV, YS- V, YS-VI, YS-VII, YS-VIII, YS-IX, YS-X, YS-XI, YS-XII, YS-XIII, bidesmoside saponin components, monodesmoside saponin components, monodesmoside saponin components
  • compositions and methods of the present technology provide for the treatment of ruminant manure with a composition comprising of one or more compounds selected from the group consisting of: 3-nitrooxypropanol, Mootral (a product of Mootral, a Swiss Agritech company), garlic extract, Yucca extract, Yucca powder, saponin, furostanol aglycone, spirostanol aglycone, chloroform, sarsapogenin, markogenin, smilagenin, samogenin, gitogenin, neogitogenin, monodesmosidic saponins, YS-I, YS-II, YS-III, YS-IV, YS- V, YS-VI, YS-VII, YS-VIII, YS-IX, YS-X, YS-XI, YS-XII, YS-XIII, bid
  • composition used to treat the manure may comprise other methane-reducing, quality and quantity enhancing components as disclosed in A. Cieslak, M. Szumacher-Strabel, A. Stochmal and W.
  • compositions of the present technology further comprise 3-nitrooxylpropanol (3-NOP).
  • compositions of the present technology further comprise Agolin Ruminant (Agolin S.A. Of Biere, Switzerland).
  • compositions of the present technology further comprise Agolin Ruminant (Agolin S.A. Of Biere, Switzerland) and 3-nitrooxylpropanol (3-NOP).
  • Agolin Ruminant Agolin S.A. Of Biere, Switzerland
  • 3-nitrooxylpropanol 3-nitrooxylpropanol
  • the methods of the present technology provide for the administration of Agolin Ruminant (Agolin S.A. Of Biere, Switzerland) in combination with a tetrasporophyte derived algal biomass to an animal.
  • Agolin Ruminant Agolin S.A. Of Biere, Switzerland
  • the methods of the present technology provide for the administration of 3-nitrooxylpropanol (3-NOP). in combination with a tetrasporophyte derived algal biomass to an animal.
  • the methods of the present technology provide for the administration of 3-nitrooxylpropanol (3-NOP) and Agolin Ruminant (Agolin S.A. Of Biere, Switzerland) in combination with a tetrasporophyte derived algal biomass to an animal.
  • the methods of the present technology provide for the treatment of manure with Agolin Ruminant (Agolin S.A. Of Biere, Switzerland) in combination with a tetrasporophyte or gametophyte derived algal biomass to an animal.
  • the methods of the present technology provide for the treatment of manure with 3-nitrooxylpropanol (3-NOP) in combination with a tetrasporophyte or gametophyte derived algal biomass to an animal.
  • the methods of the present technology provide for the treatment of manure with Agolin Ruminant (Agolin S.A. Of Biere, Switzerland) and 3- nitrooxylpropanol (3-NOP) in combination with a tetrasporophyte or gametophyte derived algal biomass to an animal.
  • Agolin Ruminant Agolin S.A. Of Biere, Switzerland
  • 3- nitrooxylpropanol 3- nitrooxylpropanol
  • the methods of the present technology provide for the treatment of manure with 3-nitrooxylpropanol (3-NOP).
  • the present technology provides for the treatment of manure from animals not supplemented with the compositions and methods described herein.
  • the algal biomass provided to the ruminant animals is formulated into a feed supplement comprising further nutrient sources and excipients described below.
  • a beef cow requires energy, protein, minerals, and vitamins in its diet. What determines how much of these nutrients is required? What determines if they need to be supplemented in the diet?
  • a female performs many functions — body maintenance, activity, weight gain, reproduction, and milk production — that all require nutrients.
  • the amount of nutrients required depends on body size, environmental conditions, how far an animal travels, desired rate of gain, stage of gestation, and level of milk production.
  • the nutritional value and quantity of available forage determine if nutrients need to be supplemented in the diet. During most of the year, warm-season forages are likely to be deficient in some minerals, especially phosphorus and certain trace elements like copper and zinc. In most situations, supplementation should include at least year -round provision of salt and a mineral with 8 per cent to 12 percent phosphors and a similar level of calcium. Vitamin A, which usually is low in dry or weathered forages, should be injected or fed in mineral or other supplements if it is suspected to be deficient. Mineral and vitamin supplementation should be a high priority because deficiencies can be corrected for relatively little cost. [00264] After addressing mineral and vitamin needs, protein and energy deficiencies must be considered. Forage protein and energy vary seasonally. Warm- season forage typically becomes deficient in protein in mid-summer and again in winter. Forage lacks adequate energy content primarily in winter, but energy available to the animal is restricted more often by a limited supply of forage rather than by deficiencies in plant composition.
  • Forage Quantity The amount of available forage obviously affects the need for supplemental feed. If grazing or hay will be limited, take immediate action. Reduce the number of animals in order to lessen the need for supplemental feeding of the remaining cows. As forage supply declines, the opportunity for animals to selectively graze decreases, and so does diet quality. Then, supplementation may become necessary even if animal numbers are reduced.
  • the amount a cow can eat in a day ranges from as little as 1.5 percent of body weight for very low quality forage to near 3.0 percent for very high quality forage.
  • the typical amount is 2.0 percent to 2.5 per cent.
  • Body Condition The level of body condition (amount of fat) affects supplemental requirements. Low body condition markedly increases the need for supplemental nutrients, and meeting such needs often is cost prohibitive. Moderate body condition significantly reduces or eliminates the need for supplements. Fleshy cows generally need little if any supplement and the daily amount of forage required often can be reduced. If forage consumption is not reduced, higher production is possible or reserves of stored body energy can be maintained. [00269] Body Size. The potential for forage consumption is related to body size, so larger animals may not require more supplement than smaller ones. Adjustments in stocking rate, to allow adequate amounts of forage per cow, may offset differences in size but will increase the cost per cow.
  • compositions and methods of the current technology can be used in conjunction with the below described supplement forms:
  • Oilseed Meals Cottonseed, soybean, and peanut meals often are manufactured as large pellets or cubes for feeding convenience. These are high protein (38 percent to 45 percent CP), medium to high energy sources, commonly fed at 1 pound to 3 pounds a day. Although relatively costly per ton, they often are the cheapest source of protein. These feeds are most useful when supplemental protein, and little or no energy, is needed. Oilseed meals are especially suitable for dry cows in moderate to good flesh when they have access to adequate amounts of low protein, medium energy forages.
  • Grain Corn and grain sorghum (milo) are the most comm on low protein, high energy sources. Other grains include oats, wheat, and barley. Grains often are the cheapest sources of supplemental energy. Similar feeds include processed by products such as wheat mids, soybean hulls, and rice bran. These by products are slightly higher in protein and a little lower in energy than grains and are relatively low in starch. Starch can interfere with forage digestibility, so these are excellent supplements to forage. Feeds in this category commonly are found in breeder /range cubes.
  • breeding/Range Cubes are most commonly 20 percent CP but also are found as 30 percent to 32 percent products. These feeds are designed to provide a combination of protein and energy, fed in larger amounts (3 to 6 pounds a day) than high protein feeds.
  • the equivalent of a 20 per cent cube can be prepared with a mix of about one- third oilseed meal and two-thirds grain. A mix of about three- fourths meal and one-fourth grain is the equivalent of a 32 percent cube.
  • Some cubes use nonprotein nitrogen (NPN), usually urea, to supply nitrogen for potential synthesis of rumen microbial protein. Cubes with low crude fiber (below 10 per cent) generally are highest in energy. Whole cottonseed, brewers grains, and some corn gluten meals are similar in protein and energy content to these cubes.
  • Protein Blocks and Liquids These feeds usually contain 30 per cent to 40 percent CP and typically are low to medium in energy. Their formulation or physical structure limits consumption to around 1 pound to 3 pound s daily. The protein portion often consists of 50 per cent to 90 per cent from NPN, but can be considerably lower. Their primary use is to provide supplemental protein on low protein, medium energy forages (below 7 percent CP, 50 per cent to 52 per cent TDN) where convenience of self-feeding is a priority. These feeds generally will not fill large voids of nutrient deficiency, nor support higher levels of animal performance.
  • Syrup Blocks and Tubs These generally range from 12 percent to 24 per cent CP (often about half from NPN) and are medium in energy. Consumption of these blocks usually is very low (typically 1/2 pound to 1 1/2 pounds a day), so higher protein versions probably are most useful. These products are not intended to directly supply much supplemental protein or energy. Rather, their theoretical function is to stimulate rumen microbes to digest more forage and produce microbial protein, which can be utilized in the small intestine. For this to occur, sufficient amounts of at least moderately digestible forage must be available. These feeds work best when supplied year round, allowing accumulation of body fat reserves that animals can utilize during typical fall and winter decline in forage quality and quantity. They generally will not support high performance.
  • Hays High quality hays, such as alfalfa, peanut, and soybean, can be used as supplements. These medium protein (usually 15 percent to 20 percent CP), medium energy sources can be limit-fed in place of one of the feeds discussed previously. Such hays also can be fed free choice, although protein is wasted, if their cost is competitive.
  • Supplements must be chosen to meet particular nutrient deficiencies. Body condition is a key factor in the choice of supplements. Thin cows are relatively more deficient in dietary energy than in protein. In contrast, fleshier cows may need extra protein, if they need anything. To minimize supplementation, use forage supplies logically. In general, hay (excluding supplemental alfalfa, etc.) should not be limit-fed with standing forage. Limit-feeding of hay encourages cows to reduce grazing and fails to use pastures while quality is reasonably good. For example, assume available forage for grazing or feeding includes some tame pasture (such as coastal bermudagrass), some native range, and some hay.
  • compositions and methods of the present technology provide for animal supplements and animal supplementation methods that inhibit methane production in ruminants and do not require any changes in the typical supplementation regimes described below.
  • compositions and methods of the present technology provide for animal supplements and animal supplementation methods that inhibit methane production in ruminants and include the appropriate mineral supplements as part of the methane inhibiting formulation.
  • Self-fed, controlled consumption can be accomplished with some feeds, especially oilseed meals and meal-grain mixes, by including an intake limiter such as salt. Cattle then will consume salt in maximum amounts of approximately 0.1 percent of body weight, or about 1 pound of salt consumption daily by a 1,000-pound cow. So, to obtain supplement consumption of 3 pounds daily in a 1,000- pound cow, a mix of 1 pound salt to 3 pounds supplement should be provided. When using salt to limit consumption, plenty of high quality water must be available. Also, cows consume more of a salt-limited supplement when it is located close to a water supply. [00285] Perhaps the most common supplement is a high quality 20 percent CP breeder/range cube (high or all-natural protein and low crude fiber), or the equivalent.
  • Such a supplement often is a compromise for the common situation of low quality forage and low to medium body condition. But this must be fed in adequate amounts, typically 3 to 6 pounds a day, to be effective. In fact, with the exception of managing weight loss in fleshy cows, there are few situations where feeding smaller amounts of such cubes is applicable. If a producer is unwilling or unable to assume the cost of required amounts of these cubes (or the equivalent), then a lower amount of a higher protein feed should be fed. But realize, however, that body condition, reproduction, productivity, and profit are likely to decline if nutrient requirements are not met. [00286] Minerals and vitamins account for a very small proportion of daily dry matter intake in beef cattle diets and can sometimes be overlooked in a herd nutritional program.
  • minerals and vitamins are needed as a very small percentage of dietary nutrients, they are very important in beef cattle nutritional programs for proper animal function, such as bone development, immune function, muscle contractions, and nervous system function. Cattle growth and reproductive performance can be compromised if a good mineral program is not in place.
  • a good mineral and vitamin supplementation program costs approximately $15 to $25 per head per year. With annual cost of production per cow generally being several hundred dollars, the cost of a high-quality mineral and vitamin supplement program is a relatively small investment.
  • Many free-choice mineral and vitamin mixes are formulated for 2- or 4-ounce daily consumption rates. For illustration purposes, if a beef cow consumes 4 ounces (1/4 pound) of a supplement per day for 365 days, then she consumes 91.25 pounds of the supplement in a year.
  • Macrominerals required by beef cattle include calcium, magnesium, phosphorus, potassium, sodium, chlorine, and sulfur.
  • Required microminerals include chromium, cobalt, copper, iodine, iron, manganese, molybdenum, nickel, selenium, and zinc. Nutrient requirements of specific mineral elements vary, depending on animal age, weight, stage of production, lactation status, breed, stress, and mineral bioavailability (the degree to which a mineral becomes available to the target tissue after administration) from the diet.
  • Dietary mineral sources include forages, concentrate feedstuffs, mineral supplements, and water.
  • Cattle can tolerate high concentrations of dietary calcium if other mineral levels are adequate in the diet.
  • Calcium recommendations are expressed in terms of a calcium to phosphorus ratio (Ca:P), where approximately 1.6:1 is ideal, with a range of 1:1 to 4:1 being acceptable.
  • Supplemental calcium sources include calcium carbonate, feed-grade limestone, dicalcium phosphate, defluorinated phosphate, monocalcium phosphate, and calcium sulfate.
  • Feed-grade limestone is approximately 34 percent calcium and is commonly added to beef cattle diets to increase the calcium levels of the diet.
  • Dicalcium phosphate is approximately 22 percent calcium and 19.3 percent phosphorus and is added to beef cattle diets to help balance the calcium to phosphorus ratio. It adds both calcium and phosphorus to the diet.
  • Recommended phosphorus levels in a mineral supplement are generally from 4 to 8 percent, largely depending on forage conditions and other levels of dietary sources of phosphorus.
  • bromoform delivered from a red macroalgae feed supplement is degraded in livestock rumens in such a way that bromoform is not absorbed into the rumen wall, or other organs, and it is not found in metabolic byproducts produced by livestock, such as their milk, meat, or manure
  • Bromoform may be artificially synthesized by several methods, including a haloform reaction using acetone and sodium hypobromite, electrolysis of potassium bromide in ethanol, or by treating chloroform with aluminum bromide.
  • the present technology solves the problem of having to build and maintain large-scale algaculture installations and the issues of excess iodine ingestion by animals and people who consume their products by providing compositions, and methods for making those compositions of target component infused feed products or target component feed supplements.
  • compositions and methods of the present technology provide for encapsulated bromoform composition comprising a bromoform bearing core and an edible polymeric material that forms an encapsulation barrier.
  • the encapsulation material forming the encapsulation barrier is an edible polymeric material and may be selected from, for example, polymers; resins; carbohydrates; modified carbohydrates; mono-, di-, oligo- or poly-saccharides; starches; modified starches; proteins; fatty acids; polyglycerol fatty acid esters; acrylics; vegetable gums; polyvinyl acetate; polyvinylpyrrolidone; poly(l-vinylpyrrolidone-co-vinyl acetate); povidone; crospovidone; Kollidon® polymers; Kollidon®-CL; Kollidon®-25; Kollidon®-30; Kollidon®- 90; Kollidon®-12 PF; Kollidon®-17 PF; Kollidon®-VA 64; Aquacoat® aqueous dispersions; halocarbons; Aquateric® enteric coatings; hydrocarbon resins; polyvinyl alcohol; cellulose acetate; hydroxyl propyl
  • the encapsulation barrier may comprise other additives such as, but not limited to: dextrose, dextrin, gum arabic, guar gum, maltose, sucrose, pectin, hydroxyl propyl cellulose (HPC), hydroxypropyl methyl cellulose (HPMC), methylcellulose, Eudragit® polymers (polyacrylates and methyacrylic acid-ethyl acrylate copolymers), CarbowaxTM SentryTM polyethylene glycol (e.g., PEG-8000), SentryTM PolyoxTM WSR N12K-NF Grade, SentryTM PolyoxTM WSR 301-NF Grade, water-soluble shellacs (preferably refined food-grade confectioners glaze), starch, modified starches, sodium chloride, alanine, arginine, asparagines, aspartic acid, cysteine, glutamine, glutamic acid, glycine, histidine, isoleucine, leucine,
  • the bromoform bearing core may further comprise: dextrose, dextrin, gum arabic, guar gum, maltose, sucrose, pectin, hydroxyl propyl cellulose (HPC), hydroxypropyl methyl cellulose (HPMC), methylcellulose, Eudragit® polymers (polyacrylates and methyacrylic acid-ethyl acrylate copolymers), CarbowaxTM SentryTM polyethylene glycol (e.g., PEG-8000), SentryTM PolyoxTM WSR N12K-NF Grade, SentryTM PolyoxTM WSR 301-NF Grade, water-soluble shellacs (preferably refined food-grade confectioners glaze), starch, modified starches, sodium chloride, alanine, arginine, asparagines, aspartic acid, cysteine, glutamine, glutamic acid, glycine, histidine, isoleucine, leucine, lysine, methionine
  • the encapsulation barrier may further comprise, or be coated with sweetners such as, but not limited to: sucrose, L-aspartyl-L-phenylalanine methyl ester, sorbitol, xylitol, and mannitol, fructose, molasses, beet sugar, brown sugar, cane sugar, confectioner's sugar, powdered sugar, raw sugar, turbinado, maple syrup, carob powder, corn syrup, sugar cane syrup, honey, sweetened condensed milk, and chocolate, saccharin, aspartame, acesulfame potassium, sucralose, and stevia.
  • sweetners such as, but not limited to: sucrose, L-aspartyl-L-phenylalanine methyl ester, sorbitol, xylitol, and mannitol, fructose, molasses, beet sugar, brown sugar, cane sugar, confectioner's sugar, powdered sugar, raw sugar,
  • the encapsulation barrier may comprise other additives such as, but not limited to: Advanta-GelTM P75, Batter Bind® S, Crisp Coat UC, Crisp Film®, Crystal Gum, Crystal TeXTM 627, Crystal TexTM 644, Crystal TexTM 648, ElastigelTM 1000J, Encapsul 855, Flojel® 60, Flojel® 65, Flojel® G, Hi-Set® 322, Hi-Set® 377, Hi-Set® C, Hi-Set® CHG, Hylon® V, Hylon® VII, ImpressionTM, K4484, Melojel®, NadexTM 772, National 0280, National 814, N-TACK®, Purity® 21D, Purity® TF, Superset® FV, Ultra-Set® FT, Dry-Tack® 250, Versa-SheenTM, Baka-PlusTM, Baka-Snak®, Capsul®, Capsul® TA, Gel N Melt
  • the bromoform bearing core may further comprise: Advanta- GelTM P75, Batter Bind® S, Crisp Coat UC, Crisp Film®, Crystal Gum, Crystal TeXTM 627, Crystal TexTM 644, Crystal TexTM 648, ElastigelTM 1000J, Encapsul 855, Flojel® 60, Flojel® 65, Flojel® G, Hi- Set® 322, Hi-Set® 377, Hi-Set® C, Hi-Set® CHG, Hylon® V, Hylon® VII, ImpressionTM, K4484, Melojel®, NadexTM 772, National 0280, National 814, N-TACK®, Purity® 21D, Purity® TF, Superset® LV, Ultra-Set® LT, Dry-Tack® 250, Versa-SheenTM, Baka-PlusTM, Baka-Snak®, Capsul®, Capsul® TA, Gel N Melt®, H-50, Hi-CapTM 100, Hi-Cap
  • the microcapsules of the invention are prepared by (i) dissolving the edible encapsulation material (e.g., polymeric or resin) in a suitable organic solvent; (ii) mixing the solubilized encapsulation material with a core material comprising bromoform adsorbed or absorbed on a carrrier material slowly adding to the mixture, with stirring, a nonsolvent for the encapsulation material.
  • a core material comprising an acid, a base, effervescent couples, and/or combinations of these components
  • coated with a permeable encapsulation barrier comprising a water-insoluble edible organic polymeric material that is optionally water-swellable.
  • slowly adding” and “slow addition” refer herein to the speed of addition which results in the even distribution of encapsulation material onto the core material. Such speed of addition can be determined without undue experimentation by those skilled in the art.
  • solvents and non-solvents include, but are not limited to: acetic acid, acetone, acetonitrile, acetyl acetone, acrolein, acrylonitrile, allyl alcohol, 1,3- butanediol, 1,4-butanediol, 1 -butanol, 2-butanol, tert-butanol, 2-butoxy ethanol, n-butyl amine, butyl dioxitol acetate, butyraldehyde, butyric acid, 2-chloroethanol, decane, diacetone alcohol, diacetyl, diethylamine, diethylene glycol diethyl ether, diethylene glycol dimethyl ether, diethylene glycol monobutyl ether, diethylene glycol monobutyl ether acetate, diethylene glycol monoethyl ether, diethylene glycol monoethyl ether acetate, diethylene glycol monomethyl methyl ether, diethylene
  • Suitable carriers include, but are not limited to, kaolin, silica, polyethylene glycol, clay nanoparticles, magnesium stearate, silica gel, surface derivatized silica, fumed silica, hectorite, colloidal magnesium-aluminum silicate, magnesium trisilicate, aluminum hydroxide, activated charcoal, talc, Neusilin, calcium silicate, magnesium oxide, zinc oxide, microcrystalline cellulose, croscarmellose sodium, and polymethacrylate.
  • Described herein are systems and methods that artificially synthesize and encapsulate bromoform in a way that is safe for livestock to consume, safe for the environment, and effective in reducing livestock enteric methane emissions as well as increasing the quantity and quality of the products derived from such livestock.
  • the components of these systems include methods that artificially synthesize bromoform and methods that encapsulate artificially synthesized bromoform.
  • a chemical reaction is used to synthesize bromoform.
  • this chemical reaction occurs in specific and controlled environmental conditions.
  • bromoform is encapsulated by a coating material.
  • bromoform is embedded in a homogeneous or heterogeneous matrix.
  • the method of encapsulation of the present technology prevents bromoform from volatizing, or chemically reacting to environmental conditions prior to digestion in livestock rumens.
  • the method of encapsulation of the present technology allows bromoform to react with common enzymes in livestock rumens thereby causing the bromoform to degrade safely and usefully.
  • a genetic screen is used to identify genes related to the synthesis and glandular encapsulation of bromoform in red macroalgae.
  • genetic screens may include methods of forward genetic screens or methods of reverse genetic screens.
  • genes related to the synthesis and glandular encapsulation of bromoform in red macroalgae are isolated and copied using recombinant DNA methods.
  • genes related to the synthesis and glandular encapsulation of bromoform in red macroalgae are copied by methods that artificially synthesize DNA.
  • organisms which are capable of hosting genes related to the synthesis and glandular encapsulation of bromoform are identified and selected.
  • host organisms are identified and selected according to their ability to produce encapsulated bromoform in a production or manufacturing system.
  • genes related to the synthesis and glandular encapsulation of bromoform are inserted into the DNA of a host organism using a vector.
  • insertion results in a transgenic host organism.
  • insertion results in a genetically edited host organism.
  • inserted genetic material is replicated by the host organism.
  • inserted genetic material is expressed by the host organism.
  • organisms which contain the genes in red macroalgae that synthesize and encapsulate bromoform are propagated or cultivated in a unique growing medium. In further embodiments, these organisms produce encapsulated bromoform which are harvested. In further embodiments, this method of synthesis and encapsulation prevents bromoform from volatizing, or chemically reacting to environmental conditions prior to digestion in livestock rumens. In further embodiments, this method of synthesis and encapsulation allows bromoform to react with common enzymes in livestock rumens thereby causing the bromoform to safely and usefully degrade.
  • FIG. 11 Shown in Figure 11 is an embodiment of a system (1) in which a chemical reaction involving one or more chemicals or reagents (11) is introduced to one or more other other chemicals or reagents (12) in the context of specific and controlled environmental conditions (13) to result in the chemical formation of bromoform CHBr3 (14).
  • a chemical reaction involving one or more chemicals or reagents (11) is introduced to one or more other other chemicals or reagents (12) in the context of specific and controlled environmental conditions (13) to result in the chemical formation of bromoform CHBr3 (14).
  • the bromoform is encapsulated by either a coating material (15) or is embedded in or encapsulated by a homogeneous or heterogeneous matrix (16).
  • FIG. 11 Shown in FIG. 11 is an embodiment of a system (2) in which biological methods are used to synthesize and encapsulate bromoform.
  • the system includes a method to identify (21) and isolate (22) genes related to the synthesis and glandular encapsulation of bromoform in red macroalgae. After these genes are identified and isolated, a method is used to copy them (23). Once copied, a vector (24) is used to insert the genes into the DNA of a host organism (25). The host organism is then propagated and cultivated in a unique medium (26). The result of the system (2) is bromoform which is encapsulated in a safe and useful way (27).
  • Oral administration constitutes the preferred route of administration for a majority of target components, such as bromoform.
  • target components that have an undesirable or bitter taste leads to lack of patient compliance in the case of orally administered dosage forms.
  • taste masking is an essential tool to improve patient compliance.
  • target components e.g., bromoform
  • the presently disclosed compositions also comprise one or more target component taste masking agents.
  • target components taste-masking agents include dry milk as described above, as well as menthol, sweeteners, sodium bicarbonate, ion-exchange resins, cyclodextrin inclusion compounds, adsorbates, and the like.
  • the formulation agent is an edible oil or fat, a protective colloid, or both a protective colloid and an edible oil or fat.
  • the bioavailability enhancing agent is also a lipophilic active agent taste masking agent.
  • protective colloids include, but are not limited to, polypeptides (such as gelatin, casein, and caseinate), polysaccharides (such as starch, dextrin, dextran, pectin, and gum arabic), as well as whole milk, skimmed milk, milk powder or mixtures of these.
  • polypeptides such as gelatin, casein, and caseinate
  • polysaccharides such as starch, dextrin, dextran, pectin, and gum arabic
  • whole milk skimmed milk, milk powder or mixtures of these.
  • polyvinyl alcohol vinyl polymers, for example polyvinylpyrrolidone, (meth)acrylic acid polymers and copolymers, methylcellulose, carboxymethylcellulose, hydroxypropylcellulose, cyclodextrins and alginates.
  • the bioavailability of the target component in a subject is at least about 1.5 times, about 2 times, about 2.5 times, about 3 times, about 3.5 times, about 4 times, about 4.5 times, about 5 times, about 5.5 times, about 6 times, about 6.5 times, about 7 times, about 7.5 times, about 8 times, about 8.5 times, about 9 times, about 9.5 times, or about 10 times greater than the bioavailability of the target component agent in the methanogenic organisms in the absence of the bioavailability enhancing agent.
  • bioavailability refers to the bioavailability of the target component to the methanogenic and other symbiotic organisms that colonize the rumen of ruminant animals, and not the bioavailability of the target components to the subject animal itself. This distinction is thought to be undesirable to expose the inner tissues of the animal to the target components, as this diminishes the amount of target component available to affect the methanogenic organisms as well as subjecting the subject animal to undesirable effects of the target components.
  • compositions and methods of the present technology comprise dosages of target components ranging from about 0.01 mg to about 1,000 mg, from about 0.5 mg to about 500 mg, from about 1 mg to about 100 mg, from about 5 mg to about 50 mg, and about from 10 mg to about 25 mg.
  • compositions and methods of the present technology comprise dosages of target components of about 0.01 mg, about 0.05 mg, about 0.1 mg, about 0.25 mg, about 0.5 mg, about 0.75 mg, about 1 mg, 5 mg, about 10 mg, about 15 mg, about 20 mg, about 25 mg, about 30 mg, about 35 mg, about 40 mg, about 45 mg, about 50 mg, about 55 mg, about 60 mg, about 65 mg, about 70 mg, about 75 mg, about 80 mg, about 85 mg, about 90 mg, about 95 mg, about 100 mg, about 150 mg, about 200 mg, about 250 mg, about 300 mg, about 350 mg, about 400 mg, about 450 mg, about 500 mg, about 550 mg, about 600 mg, about 650 mg, about 700 mg, about 750 mg, about 800 mg, about 850 mg, about 900 mg, about 950 mg, or about 1,000 mg.
  • compositions and methods of the present technology comprise dosages of target components of about 1500 mg, about 2000 mg, about 2500 mg, about 3000 mg, about 3500 mg, about 4000 mg, about 4500 mg, about 5000mg, about 5500 mg, about 6000 mg, about 6500 mg, about 7000 mg, about 7500 mg, about 8000 mg, about 8500 mg, about 9000 mg, about 9500 mg, or about 10,000 mg.
  • compositions and methods of the present technology comprise dosages of target components of about 15,000 mg, about 20,000 mg, about 25,000 mg, about 30,000 mg, about 35,000 mg, about 40,000 mg, about 45,000 mg, about 50,000 mg, about 55,000 mg, about 60,000 mg, about 6,5000 mg, about 70,000 mg, about 7,5000 mg, about 80,000 mg, about 85,000 mg, about 90,000 mg, about 95,000 mg, or about 100,000 mg.
  • the present technology is directed to dry solid lipid compositions useful for the oral delivery of lipophilic substances, and to methods for preparing and using such compositions.
  • the present technology provides for dry solid lipid mixtures that include a first component of a target component in an amount sufficient to provide a therapeutic effect when administered to an animal; a second component of a lipid comprising at least one solid fat; and a third component of at least one phospholipid, wherein the second and third components are present in an amount sufficient to increase the oral availability of the lipophilic substance when administered to the animal.
  • the dry solid lipid mixtures may include one or more of an antioxidant, a cryoprotectant or a free-flow imparting agent.
  • the present technology further relates to methods for producing such dry solid lipid mixture compositions by dissolving the lipophilic substance together with lipid components comprising at least one solid fat and at least one phospholipid in a suitable organic solvent; evaporating the solvent to dryness; hydrating the dry solid lipid mixture with an aqueous phase, with mechanical shaking, to obtain a lipid dispersion in water; homogenizing the resultant lipid dispersion, such as by high-pressure homogenization, to reduce the particle size to the submicron range; and drying the submicron dispersion.
  • the dry solid lipid mixtures according to the present technology may be prepared by directly drying the lipid mixture that is dissolved in the organic solvent.
  • the solid lipid mixture formulations can be spray dried or freeze- dried to obtain dry compositions suitable for the preparation of solid-dosage forms, such as hard gelatin capsules or tablets.
  • These solid dosage forms may further comprise cryoprotectants, antioxidants, free flowing imparting agents, surface active materials and/or emulsifiers.
  • lipid compositions are suitable for the oral delivery of target components of methane reducing and animal product quality enhancing additives.
  • the present technology is directed to dry solid lipid compositions for the oral delivery of lipophilic substances, and to methods for preparing and using such compositions.
  • the dry solid lipid mixtures of the present technology are composed of: i) a lipophilic substance, ii) a lipid or lipid mixture comprising at least one solid fat, and iii) one or more phospholipids.
  • the dry lipid mixtures of the present technology may further comprise an antioxidant, a cryoprotectant and/or a free-flow imparting agent.
  • any of a wide variety of target components in addition to bromoform can be utilized in these mixtures.
  • examples include, but are not limited to: lipophilic drugs, vitamins, and hormones. These lipophilic substances include steroids, steroid antagonists, non-steroidal anti inflammatory agents, antifungal agents, antibacterial agents, antiviral agents, anticancer agents, anti-hypertensives, anti-oxidants, anti-epileptic agents and antidepressants among many others.
  • lipophilic drugs with very poor water solubility and low oral bioavailability which could benefit from oral dosage forms are the neurohormone melatonin, the antifungal agent amphotericin B, the anticancer drug etoposide, as well as tamoxifen and its analogs. More specific compounds include cannabinoids, as exemplified by dexanabinol, and vitamins, enzymes or coenzymes, as exemplified by CoQlO. Some lipophilic substances are those which have a water solubility of ⁇ 200 pg/ml in water at room temperature (25°C), and others ⁇ 50 pg/ml.
  • the content of the lipophilic substance in the final dry solid lipid mixture may range from between about 0.01% and about 50% of the total solid weight of the mixture, or between about 5% and about 40% of the total solid weight of the mixture, or between about 7% and about 30% of the total solid weight of the mixture.
  • solid fat denotes any lipid or mixture of lipids, provided that the melting characteristics of the lipid or mixture are such that they exhibit a solid or liquid crystal phase at about 25° C.
  • Triglycerides which are solid at room temperature may be used in the preparation of the lipid mixture.
  • the solid triglycerides may be composed of a single pure triglyceride, usually available as a synthetic triglyceride, or may be a mixture of several triglycerides. Fats isolated from natural sources usually are available only as mixtures of triglycerides. Such natural mixtures are suitable for the preparation of dry lipid mixtures, provided that the melting characteristics of the mixture are such that they exhibit a solid or liquid crystal phase at about 25°C.
  • solid fats suitable for the preparation of dry lipid mixtures of the present technology are triglycerides composed of natural, even-numbered and unbranched fatty acids with chain lengths in the C10-C18 range, or microcrystalline glycerol tri esters of saturated, even-numbered and unbranched fatty acids of natural origin such as tricaprin, trilaurin, trimyristin, tripalmitin, and tristearin.
  • the content of solid triglycerides in the final dry lipid mixture is in the range of between about 20% and about 75% of the total solid weight of the mixture, or between about 25% and about 50% of the total solid weight of the mixture, or between about 30% and about 45% of the total solid weight of the mixture.
  • Phospholipids which may enter into the composition of the dry lipid mixture of the present technology include, but are not limited, natural phospholipids, such as: soybean lecithin, egg lecithin, phosphatidylglycerol, phosphatidylinositol, phosphatidylethanolamine, phosphatidic acid, sphingomyelin, diphosphatidylglycerol, phosphatidylserine, phosphatidylcholine, cardiolipin; synthetic phospholipids, such as dimyristoylphosphatidylcholine, dimyristoyl- phosphatidylglycerol, distearoylphosphatidylglycerol, dipalmitoylphosphatidylcholine; and hydrogenated or partially hydrogenated lecithins and phospholipids.
  • natural phospholipids such as: soybean lecithin, egg lecithin, phosphatidylglycerol, phosphatidyl
  • the phospholipid component may be either saturated or unsaturated and may have a gel to fluid phase transition temperature either above or below about 25°C.
  • Egg or soy phosphatidylcholines egg or soy PC
  • DMPC Dimyristoyl phosphatidylcholine
  • DPPC and DSPC Dipalmitoyl and distearoyl phosphatidylcholines
  • Acceptable dry lipid mixtures may be made with these and many other phospholipids.
  • Dry lipid mixtures may be prepared with molar ratios of phospholipid to total lipid in the range of between about 0.1 and about 0.75 (about 10 to 75 mol %), or between about 0.1 and about 0.5 (about 10 to 50 mol %).
  • the molar ratio of phospholipid to total lipid typically may be in the range of between about 0.1:1 and about 2:1, or between about 0.1:1 and about 1:1, or between about 0.2:1 and about 0.9:1, or between about 0.2:1 and 0.8:1, or between about 0.25:1 and about 0.6:1.
  • the ratio of phospholipid to total lipid is between about 0.1:1 and about 2:1, or between about 0.2:1 and about 1:1, or between about 0.4:1 and about 1.5:1, or between about 0.5:1 and about 1.25:1.
  • the content of phospholipids in the final dry solid lipid mixture is in the range of between about 2% and about 40% of the total solid weight of the mixture, or between about 5% and about 35% of total solid weight of the mixture, or between about 10% and about 30% of total solid weight of the mixture.
  • the dry solid lipid mixture of the present technology may comprise one or more additional antioxidants.
  • Antioxidants lessen the formation of oxidative degradation products, such as peroxides, from the unsaturated lipids, or other components.
  • a non-limiting example of such a preferred antioxidant is a-tocopherol, or its derivatives (such as tocopherol succinate), which are members of the Vitamin E family.
  • Many other antioxidants which are known in the art as safe for human consumption may be used, such as butylated hydroxytoluene (BHT).
  • BHT butylated hydroxytoluene
  • the content of the antioxidant in the final dry solid lipid mixture is commonly in the range of between about 0.01% and about 5% of the total solid weight of the mixture, or between about 0.1 and about 1% of the total solid weight of the mixture.
  • Dry solid lipid mixtures may further comprise a cryoprotectant material as known in the art, such as a sugar or an amino compound, to preserve the formulation during freeze-drying or
  • cryoprotectants examples include, but are not limited to: glucose, sucrose, lactose, maltose, and trehalose; polysaccharides, such as dextrose, dextrins, and cyclodextrins; and non-natural polymers, such as polyvinylpyrrolidone (PVP).
  • Other types of cryoprotectants may also be used, including amino acids, as disclosed in U.S. Pat. No. 5,472,706, incorporated herein by reference. Examples of range of cryoprotectant to total solids in the dry solid lipid mixtures include between about 0.1% and about 50% (w/w), or between about 20% and about 40% may be used.
  • the dry solid lipid mixtures of the present technology may further comprise any suitable nontoxic carrier or diluent powder, known in the art, to serve as a free-flow imparting agent.
  • suitable nontoxic carrier or diluent powder known in the art, to serve as a free-flow imparting agent.
  • suitable nontoxic carrier or diluent powder known in the art, to serve as a free-flow imparting agent.
  • additives are silicon dioxide, starch, lactose, sucrose, sorbitol, talc, stearic acid, magnesium stearate, and dicalcium phosphate.
  • the tablet or pill can be coated or otherwise compounded with pharmaceutically acceptable materials known in the art to provide a dosage form affording prolonged action or sustained release.
  • the dry solid lipid mixtures may also be prepared in gelatin capsules. But, theses mixtures may simply be mixed with any sort of animal feed product by co-milling or other mixing process.
  • the dry solid lipid mixtures are further mixed with fumed silica such as CAB-O-SIL® (Cabot Corp., Ill., US), which is fumed silicon dioxide.
  • fumed silica such as CAB-O-SIL® (Cabot Corp., Ill., US), which is fumed silicon dioxide.
  • This compound is a powdery material with extremely small particle size and enormous surface area. Fumed silica can act as a dry lubricant, promoting the free flow of the powdery mixture and preventing the mixture from caking or lumping.
  • the free-flow, anti-caking and anti-clogging characteristics of this compound are the result of several actions.
  • the submicroscopic size of the silica aggregates permits them to move easily between the larger particles of the other dry agents, and, in most cases, fumed silica probably forms a coating on the powder particles.
  • the fumed silica layer also decreases bulk tensile strength and shear strength, while neutralizing the electrostatic charge on the particles.
  • fumed silica After blending with the other powders, fumed silica adsorbs some or all the moisture which may be present in or on the product particles.
  • the fumed silica aggregates therefore, prevent other particles from contacting each other and, in turn, from forming the nuclei that would otherwise lead to the formation of larger lumps and cakes.
  • This spacing and lubricating action helps to keep materials moving through apertures, such as process equipment valves, spray heads, storage bin openings, bag and drum spouts and aerosol nozzle orifices.
  • Most powdered materials can be kept free flowing by adding a concentration of fumed silica in the final product range between about 0.5% and about 50% (total solid weight). The optimum concentration can be determined by working up or down in small steps. The weight percent of fumed silica in the final product will be in the range of between about 1% and about 40% (total solid weight). Even powders which have already become caked can usually be rendered free flowing by blending them in fumed silica (about 2% of the total solid weight, or less). The tremendous surface area of fumed silica is the reason very small amounts can provide effective action.
  • Products which cannot be processed beyond a sticky or tacky powder can be made free flowing by adding the proper level of fumed silica as a final finishing step.
  • Fumed silica can also be used to promote free flow in spray-dried or freeze-dried products. In some cases it can be introduced into the original emulsion, suspension or solution, or blended in later. Fumed silica has also been used to coat powdered and pelletized products to prevent them from caking later.
  • the content of silicon dioxide in the final dry solid lipid mixture is in the range of between about 5% and about 40% of the total solid weight of the mixture.
  • the dry target component-lipid mixtures of the present technology may be prepared by different methods as described in the following non- limiting examples appearing below.
  • Table 1 shows the Asparagopsis quantity calculation for a given reduction in methane production in dairy cattle based on normal dietary requirements and measured bromoform level in the algal biomass supplement.
  • Table 2a shows the iodine exposure based on a 120 g daily dose of supplement comprising Asparagopsis taxiformis brominata comprising a total iodine content of lOOOppm (0.1%) and a total DMI of 20kg.
  • the methods of supplementation rate determination of the present technology can be used in a variety of circumstances, taking into account the bromoform content of the supplement and the type of diet an animal is receiving.
  • table 2b illustrative supplementation rates of an algal biomass derived supplement comprising 35,000 pg/g (35 mg/g or 3.5% by dry weight of bromoform are given for dairy cows on feed, dairy cows on pasture, beef steers on pasture, beef steers on a transitioning diet, and beef steers on a finishing diet.
  • DMI dry matter intake
  • NDF Neutra Detergent Fiber
  • EXAMPLE 3 ENCAPSULATION: PREPARATION OF DRY BROMOFORM-LIPID MIXTURE BY FREEZE-DRYING FROM AN AQUEOUS DISPERSION
  • Bromoform 11.7 % w/w Tricaprin: 33.7 % w/w Lecithin: 16.8 % w/w Tocopherol succinate: 0.4 % w/w Sucrose: 23.9 % w/w Silicon dioxide: 13.5 % w/w.
  • Bromoform is obtained from Sigma- Aldrich, Inc.
  • D-a tocopherol succinate is purchased from Merck (Germany).
  • Lecithin is obtained from Lipoid KG (Germany).
  • Tricaprin is obtained from Hulls (Germany).
  • CAB-O-SIL is obtained from Cabot Corp.
  • bromoform is dissolved together with the lipid agents (phospholipids, tocopherol succinate and solid triglycerides) in dichloromethane. The solvent is evaporated until complete dryness, and the dry solid lipid mixture is then hydrated with the aqueous phase by mechanical shaking.
  • the resultant lipid dispersion is consequently homogenized by high-pressure homogenization (800 bar) using an EMULSIFLEXTM C-30 high pressure homogenizer (Avestin Inc., Canada) to reduce the particle size to the submicron range.
  • the cryoprotectant sucrose (from a 40% w/w water solution)
  • the free-flowing imparting agent CAB-O-SIL fumed silicon dioxide (from a 5% w/w suspension in water)
  • the formulation is then freeze- dried using a Christ lyophilizer (Germany).
  • the weight ratio of phospholipids to total lipids is 0.33:1.
  • EXAMPLE 4 Bromoform-Iodine Ratio Variation from different AT compositions Table 3 shows some possible bromoform-iodine ration variation from different AT compositions from seedstock harvested from various locations and grown at different light intensities and photoperiods. All but Nos. 9 and 10 are tetrasporophytes.
  • Table 4 shows the typical gametophyte Heavy Metal and Iodine Analysis and natural variation.
  • Table 8 Example of two commercially blended mineral supplements with calculated exposures at recommended supplementation rates.

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Abstract

La présente technologie se rapporte globalement à une méthode de détermination de taux d'inclusion d'algues rouges dans des aliments pour bétail et dans des compléments pour bétail.
EP21785593.1A 2020-04-10 2021-04-12 Compositions comprenant des algues et leurs méthodes d'utilisation pour augmenter la production de produit animal Pending EP4133055A4 (fr)

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WO2023108208A1 (fr) * 2021-12-14 2023-06-22 John Mickle Plantes transgéniques produisant du bromoforme
WO2023212773A1 (fr) * 2022-05-06 2023-11-09 Sea Forest Limited Procédé de préparation d'une composition anti-méthanogène
WO2023215945A1 (fr) * 2022-05-12 2023-11-16 Sea Forest Limited Améliorations du cycle de vie de l'asparagopsis
WO2023215946A1 (fr) * 2022-05-12 2023-11-16 Sea Forest Limited Procédés de culture d'asparagopsis
WO2024073579A1 (fr) * 2022-09-29 2024-04-04 Ch4 Global, Inc. Produits alimentaire à base d'algues rouges et procédés de traitement d'algues rouges
US12115248B2 (en) 2022-11-02 2024-10-15 Ruminant Biotech Corp Limited Devices and methods for delivering methane inhibiting compounds to animals
WO2024133548A1 (fr) 2022-12-21 2024-06-27 Dansk Landbrugs Grovvareselskab A.M.B.A. Procédé de réduction de la production de méthane d'un ruminant et/ou d'amélioration des performances d'un ruminant
US20240260611A1 (en) * 2023-02-08 2024-08-08 Rumin8 Pty Ltd Compositions and methods for reducing greenhouse gas

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WO2015109362A2 (fr) * 2014-01-21 2015-07-30 Commonwealth Scientific And Industrial Research Organisation Procédé de réduction de production de gaz et/ou de production de méthane totale(s) chez un animal ruminant
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