Liquid / solid hybrid materials as stabilized bromoform and other Anti-methanogenic formulations for administration to ruminants

Abstract

The present disclosure provides anti-methanogenic compositions comprising: (i) a haloform; (ii) a glycol, alcohol, wax, oil, triglyceride, or combination thereof; and (iii) a hydrophobic polymer. The disclosure also concerns animal feed additives and nutritional preparations for ruminant animals comprising such anti-methanogenic compositions as well as methods for making such compositions. The disclosure further concerns methods for reducing methane emissions from a ruminant.

Description

TECHNICAL FIELD The present disclosure relates to the use of bromoform (and other anti-methanogens) mixed with viscous organic liquids / solids dispersed in cross-linked porous polymeric materials. These haloform containing viscous liquids are homogeneously deposited throughout the polymeric material allowing for controllable release in the rumen resulting in a reduction in enteric methane emissions. BACKGROUND In the realm of livestock management, one of the significant challenges is the control of methane emissions. Ruminants, such as cows, sheep, and goats, are known to produce methane as a byproduct of their digestive process. This methane is released into the atmosphere, contributing to greenhouse gas emissions and global warming. Furthermore, the production of methane also represents a loss of energy for the animal, as the energy contained in the methane could have been used for growth or milk production. A reduction in methane from rumen fermentation has been hypothesized to cause an improvement in productivity through re-direction of energy that would be otherwise be expended in the production of methane (see Environ. Monit. Assess., 184 (2012), pp. 1929-1952). Bromoform is being commercialized as a chemical agent to reduce methane emissions from ruminant livestock. Unfortunately, aqueous solutions of bromoform are unstable due to its unfavorable air / water partition coefficient leading to significant volatility which makes effective dosing in a commercial livestock setting very difficult. Various materials are being explored for their potential to stabilize bromoform and other haloforms in a manner suitable for administration to ruminants. Ideally, the material would bind or encapsulate the haloform sufficiently to stabilize it long enough to be fed to the animal and then allow for controlled release in the rumen. Asparagopsis seaweed is a natural source of bromoform and various formulations have been developed where the bromoform is stabilized through extraction into a vegetable oil. In vegetable oils, the intermolecular interaction between bromoform and the organic liquid is much stronger (relative to bromoform / water intermolecular interactions) resulting in a substantial reduction in volatility. Waxes and high melting point fats (e.g., hydrogenated coconut oil) can also be used to stabilize bromoform. However, in both cases, the volatility will be significantly impacted by the specific surface area of either the liquid or solid. Comparing a solid formulation to a liquid formulation, it might be expected that the volatility would be reduced due to greater viscosity which would restrict diffusion to the surface (as predicted by Einstein-Stokes relationship) where it can evaporate into the air. Similarly, the release rate in the rumen will be impacted by the surface area. As a result, it is desirable to develop a formulation where the surface area and resulting release characteristic and evaporation rate can be controlled, to provide both uniform carrier characteristics (i.e., the properties will not be dependent on the surface area to volume ration) and a more homogeneous release in the rumen. Furthermore, for ruminants, liquids have limited utility as either a feed additive or as a supplement. Vegetable oils (e.g., canola oil) often form a component of feed or supplements given to ruminants. The addition of these oils can serve several purposes including increasing fat content of the diet, reducing dust during feed / supplement manufacture and lubricating manufacturing equipment. However, when dispersed in a solid matrix the surface area will increase significantly and hence the volatility will increase along with an increase in releasability in the rumen. With this approach, it is challenging to achieve a regime for the effective dosing in the rumen. The present disclosure addresses a need for stabilized forms of bromoform for use in reducing methane production in ruminants without negatively affecting their health or productivity. SUMMARY In some aspects, the present disclosure relates to cross-linked polymers which contain pores of variable size ranging from micropores to mesopores that are utilized effectively as sponges for the disclosed bromoform (or other haloform) containing liquids / solids. Viscous organic liquids have been incorporated into porous cross-linked polymers for carbon dioxide capture applications (see Ind. Eng. Chern. Res. 2021, 60, 41, 14758-14767, AU2021408141A1, and Journal of Polymers and the Environment (2022) 30:5228-5238). These materials, whether they have a high surface area or not, are generally characterized as materials with a high sorption capacity for organic liquids (i.e., greater than 0.2 mL / g sorbent). In essence, they are either rigid polymer structures that contain a pore structure for hosting the bromoform-containing liquid / wax, or they are flexible cross-linked polymer structures that can swell in the presence of the haloformcontaining liquid / wax. In these instances, the liquid presents itself as microdroplets on the pore surfaces of the polymer material thereby drastically improving the liquid / water or liquid / gas interfacial surface area. In this context, the surface area for interaction between the microdroplets supported on the solid matrix and the surrounding medium are greatly enhanced. With the use of WO 2025 / 120610                                   PCT / IB2024 / 062351 highly crosslinked porous polymer beads, a uniform solid formulation is achieved where the liquid / solid containing the haloform is dispersed throughout the pore space. In other aspects, the cross-linked polymer material can be easily dispersed as a solid in either feed or supplements and the bromoform (or other anti-methanogen) containing liquid / solid dispersed throughout the polymer matrix can thereby interact effectively with water comprising the rumen. This allows for the effective dosing of bromoform resulting in methane emissions reduction from ruminants. In certain aspects, the disclosure concerns anti-methanogenic composition comprising: i) a haloform; ii) a glycol, alcohol, wax, oil, triglyceride, or combination thereof; and iii) a hydrophobic porous polymer. Preferably the haloform is bromoform. In other aspects, the disclosure concerns animal feed additives and nutritional preparations for an animal comprising the anti-methanogenic compositions described herein. Preferably the anti-methanogenic agent is a haloform, more preferably bromoform. In still other aspects, the disclosure concerns methods of reducing methane emission by administering a feed additive or nutritional preparation described herein to ruminants, such as cows, steers, bulls, sheep, goats, and deer. Yet other aspects of the disclosure concern processes of making an anti-methanogenic composition, wherein the process comprises: i) combining an aqueous solution comprising a haloform with a wax, oil, or combination thereof to extract the haloform from the aqueous solution into an organic phase; ii) mixing a hydrocarbon solvent with the organic phases iii) separating the organic phase from the aqueous phase; iv) evaporating the hydrocarbon solvent from the organic phase; v) combining the bromoform-containing organic phase with a miscible glycol or alcohol to improve extraction into the aqueous phase of the rumen; and iv) combining this mixture with a hydrophobic polymer to form the anti-methanogenic composition. In some aspects, the present disclosure relates to an approach where hydrophobic crosslinked polymer materials with a combination of micropores (<2 nm), macropores (2-50 nm) and mesopores (>50 nm) are used to host an organic liquid / solid that has bromoform dissolved in it. The viscous liquid or solid is held in the matrix of this polymeric material under capillary forces with a fraction adsorbed onto the surface of the porous polymer. If the viscous liquid / solid is water-soluble, when the bromoform loaded polymer material is introduced into the rumen, then it is expected that the release will be very quick. If the viscous liquid / solid is oil-soluble, when the bromoform loaded polymer material is introduced into the rumen, the release will be slower as the aqueous fraction of the rumen contents will extract the bromoform over time through a partitioning or diffusion process. In other aspects, the disclosed anti-methanogenic composition, animal feed additive, or nutritional preparation is the form of beads or granules. In some aspects the diameter of the beads or granules is between 0.01 mm and 20 mm, between 0.01 mm and 15 mm, between 0.01 mm and 10 mm, between 0.01 mm and 5 mm, between 0.01 mm and 4 mm, between 0.01 mm and 3 mm, between 0.01 mm and 2 mm, between 0.01 mm and 1 mm, between 0.1 mm and 20 mm, between 0.1 mm and 15 mm, between 0.1 mm and 10 mm, between 0.1 mm and 5 mm, between 0.1 mm and 4 mm, between 0.1 mm and 3 mm, between 0.1 mm and 2 mm, between 0.1 mm and 1 mm. In other aspects, the lower limit of the diameter of the beads or granules is 0.2 mm, or 0.3 mm, or 0.4 mm, or 0.5 mm. The upper limit of the diameter may be, for example, 20 mm, or 15 mm, or 10 mm, or 5 mm, or 4 mm, or 3 mm, or 2 mm, or 1 mm. In one preferable aspect, the beads or granules have a particle size between 0.3 and 2.0 mm. The following description is intended only by way of example, and simply illustrates certain selected embodiments of devices, systems, and processes that are consistent with the disclosed subject matter as claimed herein. BRIEF DESCRIPTION OF THE DRAWINGS FIG. 1 depicts bromoform retention in water (curve on the left) and in PEG6000 / tributyrin on PAD600 (curve on the right) as measured by GCMS over time. FIG. 2A and 2B depict bromoform retention in various waxes, oils, and glycols / alcohols as measured by GCMS over time. DETAILED DESCRIPTION Preferred features, embodiments and variations of the invention may be discerned from the following detailed description which provides sufficient information for those skilled in the art to perform the invention. The detailed description is not to be regarded as limiting the scope of the preceding summary of the invention in any way. In compliance with the statute, the invention has been described in a language more or less specific to structural or methodical features. The term “comprises” and its variations, such as “comprising” and “comprised of’ is used throughout in an inclusive sense and not to the exclusion of any additional features. It is to be understood that the invention is not limited to specific features shown or described since the means herein described comprises preferred forms of putting the invention into effect. The invention is, therefore, claimed in any of its forms or modifications within the proper scope of the appended claims appropriately interpreted by those skilled in the art. WO 2025 / 120610                                   PCT / IB2024 / 062351 Throughout the specification and claims (if present), unless the context requires otherwise, the term “substantially” or “about” will be understood to not be limited to the value for the range qualified by the terms. It is to be understood that unless specifically stated otherwise, references to “a,” “an,” and / or “the” may include one or more than one and that reference to an item in the singular may also include the item in the plural. Reference to an element by the indefinite article “a,” “an” and / or “the” does not exclude the possibility that more than one of the elements are present, unless the context clearly requires that there is one and only one of the elements. As used herein, the term “comprise,” and conjugations or any other variation thereof, are used in its non-limiting sense to mean that items following the word are included, but items not specifically mentioned are not excluded. A “ruminant” is a mammal of the order Artiodactyla that digests plant-based food by initially softening and fermenting it within the animal's first stomach chambers, then regurgitating the semi-digested mass, now known as cud, and chewing it again. The process of rechewing the cud to further break down plant matter and stimulate digestion is called “ruminating”. Ruminants have a digestive tract with four chambers, namely the rumen, reticulum, omasum and abomasum. In the first two chambers, the rumen and the reticulum, the food is mixed with saliva and separates into layers of solid and liquid material. Solids clump together to form the cud, mat, or bolus. The cud is then regurgitated, chewed slowly to completely mix it with saliva, which further breaks down soluble carbohydrates . Fiber, especially cellulose, is broken down into glucose in the rumenreticulum by symbiotic anaerobic bacteria, protozoa and fungi. The degraded fiber, which is now in the liquid part of the contents, then passes through the rumen into the next stomach chamber, the omasum. The food in the omasum is digested much like it would be in the monogastric stomach and water is absorbed in the abomasum. Digested gut contents finally move to the small intestine, where the absorption of the nutrients , including microbial protein, occurs. Almost all the glucose produced by the degradation of cellulose is used by the symbiotic bacteria. Ruminants get their energy from the volatile short chain fatty acids (VFAs) produced by the bacteria, namely acetate, propionate, butyrate, valerate, and isovalerate. Ruminants include cattle, goats, sheep, giraffes, yaks, deer, antelope, and other grazing herbivores. As used herein (if present), the term “bovid” includes any member of the family Bovidae, which include hoofed mammals such as antelope, goats, and cattle, among others. As used herein, the term “reducing” includes the reduction of amount of substance in comparison with a reference. For example, the reduction in the amount of total gas and / or methane produced by a ruminant animal or animals administered a composition according to the present invention, relative to an animal or animals that were not administered a composition of the present invention. The reduction can be measured in vitro with an artificial rumen system that simulates anaerobic fermentation, or in vivo with animals confined in respiration chambers. It is within the knowledge and skill of those trained in the art to assess enteric methanogenesis by a ruminant animal. As used herein, the term “reducing methane production” refers to the reduction and / or mitigation of methane produced in the gastro-intestinal tract. The term includes the specific volume of methane generated as a result of anaerobic fermentation, for example, in the systems described herein. Fermentation in the rumen and the gut of a ruminant gives rise to production of methane. The present invention aims to reduce this process, such as to reduce the total amount of methane produced in the gastro-intestinal tract. It is within the knowledge and skill of those trained in the art to assess methane production by a ruminant animal. The term “methanogenic” refers to the process of producing methane by an anaerobic microbially generated pathway. An anti-methanogenic composition is one that interferes with the methanogenic pathway so as to reduce the amount of methane produced. As used herein, the phrase “molecular weight” refers to a weight average molecular weight (Mw) unless otherwise noted. When a polymer type followed by a number is used, the number generally refers to the approximate molecular weight of the polymer. For example, “polyethylene glycol 3000” refers to a polyethylene polymer having an average molecular weight, or a molecular weight range, of about 3000 Daltons. However, for polysorbates, such as polysorbate 20, the number in the name of the polysorbate instead indicates the average number of moles of ethylene oxide that has been reacted per mole of sorbitol in producing the polysorbate. Animal Feed Also disclosed herein is combination of an animal feed and the composition(s) described herein. The animal feed itself may be solid (e.g., powder, granules, pellets), semi-solid (e.g. gel, ointment, cream, paste), liquid (e.g. solutions, suspensions, emulsions), or any combination thereof. The composition being added to the animal feed may independently be solid (e.g., powder, granules, pellets), semi-solid (e.g., gel, ointment, cream, paste) or liquid (e.g. solutions, suspensions, emulsions). For example, the animal feed and the composition to be added to the feed may both be liquid or both be semi-solid or both be solid. Alternatively, the animal feed and composition may each be in a different physical state. For example, the animal feed may be solid or semi-solid and the composition may be liquid. The composition may, for example, be used to WO 2025 / 120610                                   PCT / IB2024 / 062351 "top-dress" (added on top) a ruminant feedlot ration or may be used to blend into a total mixed ration. The three main types of animal feed include roughages, concentrates and mixed feeds. In general, roughages contain a higher percentage of crude fiber and a lower percentage of digestible nutrients than concentrates. For example, roughages may be defined as containing equal to or greater than 20 wt.% crude fiber and equal to or less than 60 wt.% total digestible nutrients. Roughages may include, for example, dry roughages (e.g., hay, straw, artificially dehydrated forages containing at least 90 wt.% dry matter), silages (formed from green forages such as grass, alfalfa, sorghum and corn and preserved in a silo at dry matter contents of 20 to 50 %), and pastures (e.g. green growing pastures providing forage that has a high water content and generally less than 30 % dry matter). The two basic types of roughages include grasses and legumes. Grasses are generally higher in fiber and dry matter than legumes. Legumes are generally higher in protein, metabolizable energy, vitamins and minerals. Concentrates contain a relatively lower percentage of crude fiber and a higher percentage of digestible nutrients than roughages. For example, concentrates may be defined as containing less than 20 wt.% crude fiber and greater than 60 wt.% total digestible nutrients. Concentrates may include, for example, energy-rich grains and molasses. Corn, wheat, oats, barley and milo (sorghum grain) are energy-rich grains, containing about 70 to 80 wt.% total digestible nutrients. Mixed feeds are generally a mixture of roughages and concentrates to provide "complete" balanced rations and may be either high or low in energy, protein or fiber. The disclosed compositions, for example, can be combined with animal feed in various amounts depending on the total amount of the composition intended to be administered to the animal. The animal feed may, for example, comprise from about 0.0001 wt.% to about 10 wt.% of the disclosed compositions based on the total dry weight of the animal feed. The animal feed may, for example, comprise from about 0.01 wt.% to about 10 wt.% of disclosed composition, based on the total dry weight of the animal feed. For example, the animal feed may comprise from about 0.001 wt.% to about 9.5 wt.%, or from about 0.005 wt.% to about 9 wt.%, or from about 0.01 wt.% to about 8.5 wt.%, or from about 0.05 wt.% to about 8 wt.%, or from about 0.1 wt.% to about 7.5 wt.%, or from about 0.9 wt.% to about 7 wt.%, or from about 1 wt.% to about 6 wt.%, or from about 1.5 wt.% to about 5.5 wt.%, or from about 2 wt.% to about 5 wt.%, or from about 2.5 wt.% to about 4.5 wt.%, or from about 3 wt.% to about 4 wt.% disclosed composition based on the total dry weight of the animal feed. For example, the animal feed may comprise from about 0.4 wt.% to about 9.5 wt.%, or from about 0.5 wt.% to about 9 wt.%, or from about 0.6 wt.% to about 8.5 wt.%, or from about 0.7 wt.% to about 8 wt.%, or from about 0.8 wt.% to about 7.5 wt.%, or from WO 2025 / 120610                                   PCT / IB2024 / 062351 about 0.9 wt.% to about 7 wt.%, or from about 1 wt.% to about 6 wt.%, or from about 1.5 wt.% to about 5.5 wt.%, or from about 2 wt.% to about 5 wt.%, or from about 2.5 wt.% to about 4.5 wt.%, or from about 3 wt.% to about 4 wt.% disclosed composition based on the total dry weight of the animal feed. In one embodiment, the disclosed composition is administered at a dose of preferably at least 16.67, 10, 5, 3,2, 1, 0.5, 0.25 0.125 or 0.067% of the dry matter administered to the ruminant animal. For example, if a 450 kg ruminant animal (e.g., steer) consumes 2.5% to 3% of its body weight per day of feed, then the disclosed composition is administered at a dose proportional to the amount of dry matter administered to the ruminant. In the case of a 450 kg ruminant animal, and where 80% of the feed is dry matter, if the animal consumes about 2.5% of its body weight per day, then the disclosed composition is administered at a dose of about 0.27, 0.18, 0.09, 0.045, 0.0225, 0.01125 or 0.00603 kg per day to result in a dose at least 3,2, 1,0.5, 0.25 0.125 or 0.067% of the dry matter administered to the ruminant animal. In some embodiments, the anti-methanogenic compositions disclosed herein can be manufactured by a process comprising: i) combining an aqueous solution comprising a haloform with wax, oil, triglyceride, or combination thereof to extract the haloform from the aqueous solution into an organic phase; ii) mixing a hydrocarbon solvent with the organic phase to facilitate separation of the organic phase from the aqueous solution; iii) separating the organic phase from the aqueous phase; iv)    evaporating the hydrocarbon solvent from the organic phase; and v)     combining the organic phase with a hydrophobic polymer to form the anti- methanogenic composition. In some embodiments, the hydrocarbon solvent is hexane, heptane, pentane, petroleum ether, diethyl ether, toluene, xylene, or any mixture thereof. Certain compositions are obtained by directly dissolving the haloform into the glycol, alcohol, wax, oil, triglyceride, or combinations thereof to form a solution. The solution can then be combined with the hydrophobic polymer. In some compositions, the haloform is selected from the group consisting of bromoform, chloroform, iodoform, fluoroform, and combinations thereof. Although all compositions are effective, certain compositions use bromoform only. In some embodiments, the glycol is a polyethylene glycol having a molecular weight of about 200 to about 7500. In other embodiments, the glycol molecular weight is about 5000 to about 7500. In some compositions, the glycol is selected from the group consisting of polyethylene WO 2025 / 120610                                   PCT / IB2024 / 062351 glycol 200, polyethylene glycol 300, polyethylene glycol 400, polyethylene glycol 600, polyethylene glycol 1000, polyethylene glycol 1500, polyethylene glycol 3000; polyethylene glycol 3350, polyethylene glycol 4000, polyethylene glycol 6000, polysorbate 20, polysorbate 40, polysorbate 60, polysorbate 65, polysorbate 80, polysorbate 85, and combinations thereof. In certain embodiments, the wax is selected from the group comprising paraffin wax, carnauba wax, beeswax, and candelilla wax, and combinations thereof. In one embodiment, the wax is carnauba wax. Some compositions use an oil selected from canola oil, soybean oil, sunflower oil, safflower oil, com oil, coconut oil, hydrogenated coconut oil (one version sold under the brand name Copha®), palm oil, rice bran oil, grapeseed oil or a combination thereof. In some compositions, the triglyceride is tributyrin, tricaproin, tricaprylin, triacetin, trivalerin, tripropionin, or a combination thereof. Some hyper-crosslinked polymers comprise a poly(divinylbenzene) or styrenedivinylbenzene group with a macropore structure. In certain compositions, the hyper-crosslinked polymer is selected from the group consisting of PUROSORB® PAD400, PAD500, PAD600, PAD900, PAD1200, PAD350, PAD610, PAD910, PAD950, PAD950C, Amberlite® FPX66, FPX68, Amberlite® XAD2, XAD4, XAD16, XAD1 180, XAD200, XAD2010, XAD16N, XAD1600N, XAD18, XAD1 180N, XAD7HP, DIADION® Sepabe, XAD761 ® HP20, HP20SS, HP21 , SP70, SP700, SP825L, SP850, CHP20, CHP50, SP207, HP2MGL, LEWATIT® AF 5, SEPLITE® CT10, LX20, LX 207, LXA8, LXA10, LXA17, LXA680, LXA1600, LXA1 180, LXA81, LXA816, LXA817, LXA8302, LXA88, LXS868, and AB-8. The hydrophobic polymer may be cellulose or a derivative thereof in some compositions. Derivatives of cellulose include methylcellulose, cellulose acetate, hydroxyethyl cellulose, hydroxypropyl cellulose, carboxymethyl cellulose, ethyl cellulose, cellulose gum, microcrystalline cellulose, or a combination thereof. In some embodiments, the hydrophobic polymer is a hyper-crosslinked polymer having a surface area of about 300 to about 4000 m2 / g. In certain embodiments, the surface area about 500 to about 1500 m2 / g or 1000 m2 / g. In other embodiments, the hydrophobic polymer is a hyper-crosslinked polymer having a swelling capacity of at least 0.01 mL / g, at least 0.05 mL / g, at least 0.10 mL / g, at least 0.15 mL / g, at least 0.20 mL / g, at least 0.25 mL / g, at least 0.30 mL / g, at least 0.35 mL / g, at least 0.40 mL / g, at least 0.45 mL / g, or at least 0.50 mL / g. In one embodiment, the hydrophobic polymer is a hypercrosslinked polymer having a swelling capacity of between 0.01 mL / g and 1.0 mL / g, between 0.01 WO 2025 / 120610                                   PCT / IB2024 / 062351 mL / g and 0.9 mL / g, between 0.01 rnL / g and 0.8 mL / g, between 0.01 mL / g and 0.7 mL / g, between 0.01 mL / g and 0.6 mL / g, or between 0.01 mL / g and 0.5 mL / g. Some anti-methanogenic compositions disclosed herein are used as an animal feed additive. Certain anti-methanogenic compositions disclosed herein are used as a nutritional preparation for an animal. Use of the anti-methanogenic compositions in an effective amount in the animal feed additives or the nutritional preparations disclosed herein reduce methane emission of a ruminant. In some embodiments, the disclosed anti-methanogenic composition, animal feed additive, or nutritional preparation further comprises biochar. Nonlimiting examples of biochar include the following: • Wood-based Biochar: Produced from woody biomass such as hardwood or softwood. • Crop Residue Biochar: Made from agricultural residues such as corn stalks, rice husks, wheat straw, or other crop residues. • Manure-based Biochar: Produced from animal manure, often mixed with other organic materials. • Green Waste Biochar: Created from green waste, which includes yard trimmings, leaves, and other plant materials. • Nutshell Biochar: Produced from the shells of nuts, such as coconut shells or walnut shells. • Bamboo Biochar: Made from bamboo, which is a fast-growing and renewable resource. • Municipal Solid Waste (MSW) Biochar: Derived from the organic fraction of municipal solid waste after proper treatment and processing. • Algae-based Biochar: Created from micro or macro algae biomass. • Paper-based Biochar: Made from paper waste or other cellulose-rich materials. • Switchgrass Biochar: Derived from the perennial grass species switchgrass. Any embodiment of the invention is meant to be illustrative only and is not meant to be limiting to the invention. Therefore, it should be appreciated that various other changes and modifications can be made to any embodiment described without departing from the spirit and scope of the invention. All headings are for the convenience of the reader and should not be used to limit the meaning of the text that follows the heading, unless so specified. WO 2025 / 120610                                   PCT / IB2024 / 062351 The present invention is further illustrated by the following examples that should not be construed as limiting. The contents of all references, patents, and published patent applications cited throughout this application, as well as the Figures, are incorporated herein by reference in their entirety for all purposes. EXAMPLES Example 1. Measurement of Bromoform with Gas Chromatography-Mass Spectroscopy Gas chromatography-mass spectroscopy (GCMS) was used to analyse the bromoform content in water solutions and in the stabilized solid samples that are the subject of this patent. A headspace technique is used where the sample placed inside of a 20 mL sample vial with a silicone septum is exposed to an elevated temperature. When the sample reaches equilibrium after a period, the vial is pressurized with helium gas and then a portion of this volume is analyzed by the GCMS. To quantify the amount of material, the temperature and volume of liquid / solid are chosen such that the concentration in the headspace at elevated temperature is proportional to the amount in the original unheated sample. For bromoform in water, 10 mL of solution is used, and the temperature is 70 °C; a temperature of 180 °C is used to analyse the bromoform content with the solid / liquid infused porous polymers (e.g., canola oil infused into poly(divinylbenzene) beads) where 600 mg of sample is assayed. At these elevated temperatures, the bromoform in its respective matrices has demonstrated a linear response using a standard addition method and the generation of numerous calibration curves. A Shimadzu GC-2030 gas chromatograph is used in conjunction with a HS-20NX headspace sampler module and a GCMS-QP2020NX single quadrupole mass spectrometer. A Shimadzu SH-I-5Sil column (with a silylene phase comprising a 5% diphenyl and 95% dimethyl polysiloxane). This column was chosen for its ability to separate various halomethanes (e.g., chloroform, bromochloromethane). For the headspace analyzer, the oven temperature used is dependent on the matrix (see above). Sample line and transfer line temperatures of 150 °C and 150 °C, respectively, where used. A shaking level of 4 was used for a 20-minute equilibrating time. The sample was pressurized for 1 minute to 55.2 kPA with helium and allowed to equilibrate for 0.50 minutes. Load and load equilibration times of 0.50 and 0.10 minutes, respectively, were used. An injection time of 1.50 minutes was used followed by a 4-minute needle flush time. The GC oven temperature is set to 50 °C initially and ramped to 150 °C at a 35 °C / minute rate, then to 200 °C at a 50 °C / minute rate and finally to 250 °C at a 70 °C / minute rate. A 50-ratio split injection is used with a gas pressure of 147.0 kPa and total column flow of 54.0 mL / minute (comprising 1.00 mL / minute column flow and 3.0 mL / minute purge flow). For the mass WO 2025 / 120610                                   PCT / IB2024 / 062351 spectrometer, ion source and interface temperatures of230 °C and 220 °C, respectively, were used. The spectrometer is tuned using perfluorotributylamine to maximize the m / z 502 peak. A detector voltage of -0.1 kV relative to this tuning is used for this analysis. Example 2. Stability of 100 pM Bromoform in Water Background Bromoform at a concentration of 10 mM has been tested for its stability in water previously where it was found to very quickly evaporate into the atmosphere (i.e., within hours of putting it into the water). Here, the Inventors attempted with a much smaller concentration of bromoform of 100 pM in water to analyze if bromoform at this lower concentration is effective in remaining in the water solution for longer. This study was conducted as a basis for comparison with the bromoform stabilized on solid carriers. Materials and Methods Sample Preparation A 100 pM solution of bromoform was made in 500 ml of water. Approximately 12.6 mg of synthetic bromoform (equivalent volume) was pipetted into the water and thoroughly mixed for about a minute to ensure homogeneity. Experimental Parameters / Conditions The prepared solution was kept in an open bowl with a magnetic stirrer continuously stirring at approximately 50 rpm. The bowl was kept inside a fume cupboard to minimize bromoform stratification along the surface of the water since bromoform is of higher density than air. The airflow from the cupboard should have been able to reduce bromoform stratification from the water surface. Sample Collection The first sample was taken just after preparing the 100 pM solution to mark the starting concentration. At every 1-hour interval around 10 ml of sample was taken into a headspace vial and sealed using crimp caps. Samples collected for the first day were kept more frequently than for the other two days. This sample collection frequency was followed as it was seen before with a 10 mM concentration that bromoform in water is not stable and was seen to be lost in air in less than 24 hours. Results The headspace vials of the collected samples were run in GCMS. Due to the low concertation of bromoform in the water, no dilution was required for these samples. Following the parameters for water analysis relating to the gas chromatogram method, an appropriate method file was loaded, and the results obtained were documented (see FIG. 1). The GCMS analysis was stopped after sample 9 because after sample 6 there were trace levels or no bromoform content registered in the GCMS read. Bromoform levels from the 100 pM aqueous solution had dissipated to trace amounts within the first 5 hours of measurement (see the curve on the left in FIG. 1). Example 3. Formulation of Bromoform in Glycols, Waxes, or Oils with PAD600 Background One way to administer bromoform to a ruminant is in the form of a glycol, wax, or oil. For example, a vegetable oil which is immiscible with water can be added to a water solution containing bromoform and mixed where the bromoform transfers into the oil phase. Alternatively, bromoform can be added directly into the vegetable oil at the desired concentration. This bromoform infused oil can then be added to a porous polymer solid. The Inventor’s initial testing utilized hyper-crosslinked poly( divinylbenzene), which is sold as PUROSORB™ PAD600 (polydivinylbenzene macroporous, adsorbent resin, non-ionic form) (“PAD600”) by Purolite. This material has applications in the water and food processing industries. It is anticipated that crosslinked hydrophobic polymers (similar to hydrophilic hydrogel materials) as well as hydrophobic cellulose materials could be utilized for this purpose. Another consideration is the availability of bromoform in the rumen; it is expected that greater availability will be critical for attaining anti-methanogenic efficacy. On the one hand, it is advantageous for the bromoform to be stabilized in the oil phase. An undesired side effect is that the octanol-water partition coefficient predicts that the concentration in the water phase (where it is likely needed to affect a reduction in methane emissions) may be limited. There is work in the pharmaceutical industry (see Journal of Drug Delivery Science Technology, 22 (2012), 181-187) where glycols (e.g., PEG, PPG, propylene glycol, triacetin, fatty acids) are added to help with solubility between hydrophobic / hydrophilic components. Adding these components to the vegetable oil even with limited miscibility will help with increasing bromoform concentration in the aqueous phase within the rumen. Another approach would be to directly add pure bromoform to a low molecular weight glycol (e.g., poly(ethylene glycol) - molecular weight 300 or 600 or PEG300 / PEG600) where both will stabilize bromoform (ethanol is a common stabilizer used by the chemical industry so PEG is WO 2025 / 120610                                   PCT / IB2024 / 062351 likely to elicit both chemical stability and volatility reduction). However, PEG is soluble in water and would not be useful for extraction of bromoform from water; vegetable oils are useful in part because they are immiscible with water. This analysis is associated with the bioproduction of bromoform using a VHPO-catalyzed reaction in aqueous media to form hypobromous acid from hydrogen peroxide and potassium bromide which then reacts with a carbon substrate (e.g., acetyl acetone, dimedone, etc.) to form bromoform via the haloform reaction. Vegetable oils due to their hydrophobicity and immiscibility with water are useful for the liquid-liquid extraction of bromoform. However, this may limit the bioavailability in the aqueous phase of the rumen with the bromoform remaining in the oil phase. In this case, solvents such as tributyrin, which are largely hydrophobic and useful for liquid-liquid extraction with limited water solubility, are useful in place of vegetable oils (e.g., canola oil). Furthermore, it is possible to add water soluble solvents (e.g., PEG300) to these limited water solubility solvents (e.g., tributyrin) following the liquid-liquid extraction. In doing so, the dissolution of the solvent infused into porous polymer in the aqueous phase of the rumen will be further facilitated. The Stokes-Einstein relation predicts an inverse relation between the solute diffusion coefficient and liquid viscosity. The diffusion coefficient is strongly correlated with the volatilization rate as predicted by liquid-gas mass transfer theory assuming that the film thicknesses is relatively unchanged at low solute concentrations. This suggests that increasing the viscosity of the oil will reduce volatilization. As such, the Inventors decided to evaluate copha (i.e. hydrogenated coconut oil). In addition, various molecular weight PEG liquids (i.e., 300, 400 and 600) were also selected for testing and comparison. Materials and Methods With PAD-600, mass ratios of 0.7-1 to 1 (liquid / solid to polymer) resulted in a mixture that had a dry appearance and was free-flowing as a powder. Bromoform was directly added to either the liquid or solid at the desired concentration. As a melt or viscous liquid, this was combined with PAD600. A 1:1 mass ratio of bromoform loaded liquid / solid and PAD600 performed well in maximizing loading while achieving a freely flowing bead material. The following materials were prepared for testing: • Bromoform / canola oil on PAD600 (9.4 wt.%) • Bromoform / PEG300 on PAD600 (9.9 wt.%) • Bromoform / copha on PAD600 (9.0 wt.%) • Bromoform / carnauba wax on PAD600 (8.4 wt.%) • Bromoform / Tween 20 on PAD600 ((8.0 wt. %) • Bromoform / PEG600 on PAD600 (9.7 wt.%) • Bromoform / PEG6000 onPAD600 (9.1 wt.%) • Bromoform / Tributyrin with PEG6000 on PAD600 Preparation of Diluted Solid Samples for Retention Test For GCMS analysis, these solid samples were diluted so that they could be directly analyzed using a split ratio of 50 using the GCMS at a 180°C headspace oven temperature. A weight-by-weight dilution of 40-fold using PAD600 resin beads (i.e., 0.5 grams of sample is combined with 19.5 grams of dried PAD600 resin beads in a glass container) was carried out. Each container was gently rotated to ensure thorough mixing of the sample and the polymer to produce a homogeneous diluted sample. From the diluted samples prepared, roughly half of it was distributed to eight other separate glass jars for the purpose of keeping a control to be kept in the freezer for each of the samples that is to be kept inside the oven. GCMS Calibration Curve and Analysis of Samples 125 uM, 250 pM, 375 pM, 500 pM, 625 pM, 750 pM, 875 pM and 1000 pM solutions of bromoform (made up from 100 mM bromoform in ethanol solution) in 10 mL of canola was used to establish a calibration curve in a 20 mL headspace vial at a GCMS oven temperature of 130°C. For these tests, the diluted solid samples are placed in oven at 40°C temperature and GCMS tested on an approximately weekly basis to measure retention. The samples were placed in 120 mL clear glass jars as a thin layer (approximately 1 cm thick) which were then placed in a fan forced oven with the jar lids removed. Results The retention analysis performed with the various glycols, waxes, and oils demonstrated similar stabilization of the bromoform by each carrier extending the period in which bromoform could be detected from a matter of hours (for an aqueous solution of bromoform, see the left curve in FIG. 1) to a matter of weeks (see FIGs. 2 A and 2B). Among the glycols / alcohols in FIG. 2A, there is not a significant difference in the behavior based on viscosity. The Stokes-Einstein relationship would suggest that PEG6000 would more strongly retain bromoform relative to PEG300 as an example. This is not observed suggesting that the diffusion distance within the microdroplets dispersed through the polymer matrix is very small and that the rate limiting step for loss is evaporation of the liquid surface to the environment. Comparing FIGs. 2A and 2B, it appears that there is a slightly stronger retention for the waxes and oils (FIG. 2A) (which are hydrophobic) relative to glycols / alcohols (FIG. 2B) (which are hydrophilic). Similarly, comparing carnauba wax and canola oil, the retention does not seem to be affected by viscosity. Example 4. Methanogen Assay Results Background Unprotected formulations of methane inhibitors, such as bromoform will be rapidly lost from the rumen due to the volatile nature of these compounds and their rate of passage through the rumen. Formulations that “entrap” bromoform for longer in the rumen are likely to provide higher levels of efficacy. However, these stabilised formulations will still need to be able to release the active compound to elicit a response. A simple in vitro bioassay using Methanobrevibacter smithii was used to assess the effectiveness of these stabilised formulations in relation to the bioavailability of the bromoform inhibitor and its effect on methane production. Materials and Methods Culturing and media Pure cultures of Methanobrevibacter smithii were inoculated from glycerol stocks into modified anaerobic BRN media (Maczulak et al., 1989, Miller et al., 1982) (30% clarified rumen fluid and Cysteine-HC1 as a reductant) in 10 mL Balch Tubes (Balch and Wolfe, 1976). The same media was used for all methane inhibition assays. Inoculated cultures were pressurised with UHP hydrogen gas to 150 kPa over pressure and incubated at 39°C with gentle shaking at 70 rpm on an orbital shaker. Cultures were regularly transferred into new media (1 / 100 dilution) every 48h (ODeoo approx. 0.3) and used as inoculum for assays with stabilised formulations. Culture headspace gases were analysed for CH4 production and quantification after 24 and 48 hours of incubation. Stabilised formulation preparation For each formulation a matrix control (containing no bromoform) and the bromoform charged matrix were assessed. As these formulations cannot be dissolved in a diluent, a weighted amount was added to each individual tube / bottle. A minimum amount of compound was chosen that would provide a final concentration of 5 pM of bromoform in a 100 ml bottle for the bromoform formulations (see Table 1). This was necessary as a sub-milligram amount of material which is difficult to accurately weight would be necessary to do these experiments in a 10 mL tube. The matrix only formulation assays were performed in 10 ml tubes with the same weighed amount as for the 100 ml bottles, equating to a 10-fold increase in the matrix for these tubes compared with the bottles. Methanogen assay All assays were set up in an anaerobic chamber (approx. 95% CO2 + 5% H2). Pre-warmed bulk modified BRN media (~ 2 L) was inoculated with 1:50 vol / vol of a 36-40 hour growing (ODeoo approx. 0.3) M. smithii culture and mixed thoroughly to ensure uniform distribution of methanogens and dispensed into replicate tubes or bottles with pre weighted formulations. A M. smithii control culture (no additive) and a bromoform control culture (0.5 pM) were also used. All formulations were performed in duplicate cultures for both matrix and bromoform assays. All tubes and bottles were capped in the anaerobic chamber and pressurised with UHP Hydrogen (BOC, Australia) using a gassing-manifold and pressure gauge to 150 KPa overpressure. Tubes and bottles were incubated, at 39°C in an orbital shaker incubator at 70 rpm. Headspace pressures were recorded using microswitch pressure transducers (Cat# 185PC15DT, Honeywell, IL) connected to a datalogger (Wellman. Belgium), after 24h; if unchanged, they were allowed to be incubated until 48h. Methane and H2 analysis was done by drawing approximately 2ml headspace gas in a gastight syringe with a 25G x 25mm needle (Terumo) and injected into a Shimadzu GC-2014 with a TCD detector as described by Gagen et al (Gagen et al., 2014) with N2 as carrier gas; CH4 and H2 calibration was done using a 5-level standard mixture of CH4, H2 & CO2 in N2 (Air Liquide, Australia) with linear R2 = > 0.99 for each gas standard. Gas percentages (CH4) were converted using headspace volumes and gas pressures and reported as micromoles of CH4 for 10ml or 100ml using the Ideal Gas Law (PV = nRT). Results In vitro methane production from M. smithii was as predicted for the control cultures with a production of about 325 pmol / 10 ml culture in 24 hrs (see Table 1). The inclusion of bromoform at a concentration of 0.5 pM completely inhibited all growth and production of methane from M. smithii. There was no inhibition of growth or methane production for all control matrix tubes. Control matrix tubes were at 10X the concentration of the bromoform matrix tubes. Overall, bromoform formulations did provide an inhibitory effect on methane production from M. smithii. We also tested alpha-cyclodextrin (which is a solid that has been previously used for host-guest chemistry to stabilize bromoform, see M. Honan et al., Animal Production Science 62(14) 13031317) at a 18.9 wt.% bromoform loading for comparison. With copha (as an oil), carnauba wax (as a solid wax) and Tween 20 (as an alcohol / glycol) loaded with bromoform and incorporated 5 into PAD600 (which is a porous polymer), a strong reduction in bromoform production relative to matrix was seen (see Table 1). Table 1. Amounts of material used for assay and methane production results Formulation Quantity (mg) CH4 Produced with Matrix Only CH4 Produced with Matrix and Bromoform Control (pure bromoform) -0.5 pM — 328 0 Bromoform on alphacyclodextrin (18.9 wt.%) -12.5 pM 1.68 356 0 Bromoform / carnauba wax on PAD600 (8.4 wt.%) - 5 pM 1.5 292 0 Bromoform / copha on PAD600 (9.0 wt.%) - 5 pM 1.4 212 0 Bromoform / Tween20 on PAD600 (8.0 wt.%) - 5 pM 1.58 376 0 Example 5. Protocol for Liquid-Liquid Extraction and Stabilization onto PAD600 10         VHPO enzyme can be used to catalyze the production of bromoform through the haloform reaction. We describe here a procedure for the isolation and purification of bromoform derived from a complex supernatant mixture containing VHPO enzyme and biomass. This experimental procedure encompasses an initial extraction step using tributyrin (boiling point of 305°C) as a solvent followed by a liquid-liquid extraction process employing hexane (boiling point 69°C) as 15 the solvent of choice. The resulting solution mixture was subsequently combined with a highly crosslinked poly(divinyl benzene) (i.e., PAD 600), yielding a distinct solid formulation that holds the compound of interest in a freely flowing bead material. With canola oil and water, we have demonstrated molar basis partition coefficients of approximately 700 to 800; with liquid-liquid extraction, hydrophobic solvents will readily extract bromoform from an aqueous solution. The WO 2025 / 120610                                   PCT / IB2024 / 062351 isolation and purification of specific compounds from complex mixtures are fundamental processes to develop a stabilized formulation that can be implemented in an agricultural setting to reduce enteric methane. Preparation of sample The experimental process began with the introduction of 100 ml of tributyrin into a homogenized supernatant solution, 10 L in volume, containing bromoform at approximately 9.2 mM concentration (as measured by GCMS) yielding 23.2 grams of bromoform. The resulting mixture was vigorously shaken to achieve uniform homogeneity of the tributyrin within the solution. The tributyrin (which is a low molecular weight triglyceride fat) extracts a substantial amount of the bromoform from the aqueous solution. The presence of a substantial amount of biomass makes separation of the two phases difficult. Subsequently, the homogenized supernatant was divided into two separate 5000 ml Erlenmeyer flasks, which served as containers for the treatment with hexane. Hexane was chosen for its excellent solvating properties for oil-based substances and was introduced into the two flasks. Its low boiling point and high evaporation rate make hexane ideal for the liquid-liquid extraction process. During this phase, 400 ml of hexane was added to facilitate the dissolution of the bromoform-tributyrin mixture from the supernatant into the hexane phase. This selectively transfers the desired components, tributyrin-bromoform, into the hexane phase while leaving unwanted constituents behind due to hexane's compatibility with oil-based compounds. The resulting mixture was subsequently dried using magnesium sulphate. The supernatant solution from the Erlenmeyer flasks were then carefully transferred into 500 mL separating funnels and allowed to settle. Hexane and the supernatant undergo phase separation, forming two distinct layers, with hexane settling at the top. The lower supernatant layer was carefully filtered out, leaving the hexane undisturbed at the upper layer. This process was repeated for the entire 10 L supernatant, with hexane-tributyrin mixture being collected at the top of the separating funnel. The collected mixture was afterwards filtered into a separate glass container and left to evaporate inside a fume cupboard. The final solution that remains is enriched with tributyrin containing bromoform, a significant achievement in compound isolation. More tributyrin could have been used as an alternative to hexanes; however, it is not possible to concentrate the solution (i.e., through evaporation of a solvent more volatile than bromoform). Alternatively, the biomass could have WO 2025 / 120610                                   PCT / IB2024 / 062351 been removed prior to the liquid-liquid extraction making phase separation easier with a small amount of tributyrin. Approximately 60 g of purified tributyrin was recovered from the supernatant representing a significant loss of solution during the separation funnel and filtration processes. To this, 60 g of PEG 6000 was measured, melted, and added to increase the viscosity. Subsequently, 120 g of PAD600 polymer was introduced. A thorough mixing and stirring was maintained to ensure uniform distribution of the polymer within the tributyrin and PEG 6000 mixture, resulting in a distinct solid mixture formulation. This solid formulation is a concentrated mixture with a bromoform loading as measured by GCMS of 3 wt.%. This represents a total bromoform quantity of approximately 7 grams corresponding to an overall yield of approximately 30%. This mixture was kept in an oven with a regulated temperature of 25°C and 60% relative humidity (RH) for stability testing. Stability testing The solid mixture was separated into six different 20 ml glass jars containing 20 g of the solid formulation in each of the glass jars. Of the 6 jars, 3 of them were kept inside the oven with lids open, and the other 3 had the lids closed inside oven. These samples were tested under GCMS parameters at a temperature of 180 degrees every 7 days to check the retention capability and how the samples performed at the conditions of 25°C and RH 60%. GCMS sample preparation The six jars of sample were all concentrated. Therefore, a solid-to-solid dilution was required for all six jars using the PAD600 polymers before putting them in the headspace vials for GCMS analysis. A 40-fold dilution was performed for all 6 samples. Around 0.25g of the oven samples was measured using an analytical balance in a separate jar and was followed by an addition of 9.75 g of PAD600 polymer to dilute. The resulting mixture was shaken and stirred thoroughly ensuring homogeneity after dilution. The dilution procedure was performed for the other samples: closed lid jars, open lid ones and control (kept in the freezer) as well and labelled accordingly. Finally, into a 20 ml headspace vial approximately 0.6 g / 600 mg of the diluted solid sample was measured and sealed using crimp caps and crimper. Stability testing results During the testing phase for the samples, it was seen from the GCMS data that the three glass jars that were kept open had approximately 55% loss after 1 week and 87% loss after 3 weeks (see the curve on the right in FIG. 1). While the loss is significant, this stabilized formulation is significantly more stable than an aqueous bromoform solution (see the curve on the left in FIG. 1). The comparatively large loss is attributed to the high surface area that is imparted by the liquid mixture being dispersed throughout the high surface area porous polymer (i.e., PAD600 has a surface area of approximately 850 m2 / g). The liquid on its own has significantly lower volatility as the surface area is much smaller, and hence liquid-gas interaction will be much less leading to a lower evaporation rate. While the invention has been described in connection with specific embodiments thereof, it will be understood that it is capable of further modifications and this application is intended to cover any variations, uses, or adaptations of the invention following, in general, the principles of the invention and including such departures from the present disclosure as come within known or customary practice within the art to which the invention pertains and as may be applied to the essential features hereinbefore set forth. All references, articles, publications, patents, patent publications, and patent applications cited herein within the above text and / or cited below are incorporated by reference in their entireties for all purposes. However, mention of any reference, article, publication, patent, patent publication, and patent application cited herein is not, and should not be taken as acknowledgment or any form of suggestion that they constitute valid prior art or form part of the common general knowledge in any country in the world. REFERENCES Balch, W. E. & Wolfe, R. S. 1976. New approach to the cultivation of methanogenic bacteria: 2-mercaptoethanesulfonic acid (HS-CoM)-dependent growth of Methanobacterium ruminantium in a pressureized atmosphere. Appl Environ Microbiol, 32, 781-91. Gagen, E. J., Wang, J., Padmanabha, J., Liu, J., De Carvalho, I., Liu, J., Webb, R. 1., Al Jassim, R., Morrison, M., Denman, S. E. & Mcsweeney, C. S. 2014. Investigation of a new acetogen isolated from an enrichment of the tammar wallaby forestomach. BMC Microbiol, 14, 314. Honan, M., Feng, X., Tricarico, J.M. 2020. Feed additives as a strategic approach to reduce enteric methane production in cattle: modes of action, effectiveness and safety. Animal Production Science 62(14) 1303-1317 Maczulak, A. E., Wolin, M. J. & Miller, T. L. 1989. Increase in colonic methanogens and total anaerobes in aging rats. Appl Environ Microbiol, 55, 2468-73. Miller, T. L., Wolin, M. J., De Macario, E. C. & Macario, A. J. 1982. Isolation of Methanobrevibacter smithii from human feces. Applied and Environmental Microbiology, 43, 227-232. Magnusson, M,. Vucko, M,. Neoh, T,. & de Nys, R. 2020. Using oil immersion to deliver a naturally-derived, stable bromoform product from the red seaweed Asparagopsis taxiformis. Algal Research 51, 102065.

Claims

1. An anti-methanogenic composition comprising:i)      a haloform;ii) a glycol, alcohol, wax, oil, triglyceride, or combination thereof; and iii)    a hydrophobic polymer.

2. The anti-methanogenic composition of Claim 1, wherein the haloform is selected from the group consisting of bromoform, chloroform, iodoform, fluoroform, and combinations thereof.

3. The anti-methanogenic composition of Claim 2, wherein the haloform is bromoform.

4. The anti-methanogenic composition of any one of Claims 1-3, wherein the glycol is a polyethylene glycol having an average molecular weight of about 200 to about 7500.

5. The anti-methanogenic composition of Claim 4, wherein the polyethylene glycol is selected from the group consisting of polyethylene glycol 300, polyethylene glycol 400, polyethylene glycol 600, polyethylene glycol 1000, polyethylene glycol 1500, polyethylene glycol 3000; polyethylene glycol 3350, polyethylene glycol 4000, polyethylene glycol 6000, polysorbate 20, polysorbate 40, polysorbate 60, polysorbate 65, polysorbate 80, polysorbate 85, and combinations thereof.

6. The anti-methanogenic composition of any one of Claims 1-5, wherein the wax is selected from the group comprising paraffin wax, carnauba wax, beeswax, and candelilla wax, and combinations thereof.

7. The anti-methanogenic composition of Claim 6, wherein the wax is carnauba wax.

8. The anti-methanogenic composition of any one of Claims 1-7, wherein the oil isselected from the group consisting of canola oil, soybean oil, sunflower oil, safflower oil, corn oil,WO 2025 / 120610                                   PCT / IB2024 / 062351coconut oil, hydrogenated coconut oil (also known as copha), palm oil, rice bran oil, grapeseed oil and combinations thereof.

9. The anti-methanogenic composition of any one of Claims 1-8, wherein the triglyceride is tributyrin, tricaproin, tricaprylin, triacetin, trivalerin, tripropionin, or a combination thereof.

10. The anti-methanogenic composition of any one of Claims 1-9, wherein the haloform is dissolved in the glycol, alcohol, wax, oil, triglyceride, or combination thereof to form a solution, and the solution is combined with the hydrophobic polymer.

11. The anti-methanogenic composition of any one of Claims 1-10, wherein the hydrophobic polymer is a hyper-crosslinked polymer having a surface area of about 300 to about 4000 m2 / g and / or a swelling capacity of at least 0.2 mL / g.

12. The anti-methanogenic composition of Claim 11, wherein the hyper-crosslinked polymer comprises a styrene-divinylbenzene polymer or poly(divinylbenzene) polymer with a macropore structure.

13. The anti-methanogenic composition of Claim 11, wherein the hyper-crosslinked polymer is selected from the group consisting of PUROSORB® PAD400, PAD500, PAD600, PAD900, PAD1200, PAD350, PAD610, PAD910, PAD950, PAD950C, Amberlite® FPX66, FPX68, Amberlite® XAD2, XAD4, XAD16, XAD1 180, XAD200, XAD2010, XAD16N, XAD1600N, XAD18, XAD1 180N, XAD7HP, DIADION® Sepabe, XAD761 ® HP20, HP20SS, HP21 , SP70, SP700, SP825L, SP850, CHP20, CHP50, SP207, HP2MGL, LEWATIT® AF 5, SEPLITE® CT10, LX20, LX 207, LXA8, LXA10, LXA17, LXA680, LXA1600, LXA1 180, LXA81, LXA816, LXA817, LXA8302, LXA88, LXS868, AB-8, and combinations thereof.

14. The anti-methanogenic composition of Claim 13, wherein the hyper-crosslinked polymer is a non-ionic polydivinylbenzene macroporous, adsorbent resin, preferably PUROSORB® PAD600.

15. The anti-methanogenic composition of any one of Claims 1-10, wherein the hydrophobic polymer is cellulose or a derivative thereof.WO 2025 / 120610                                   PCT / IB2024 / 06235116. The anti-methanogenic composition of Claim 15, wherein the derivative of cellulose is selected from the group consisting of methylcellulose, cellulose acetate, hydroxyethyl cellulose, hydroxypropyl cellulose, carboxymethyl cellulose, ethyl cellulose, cellulose gum, microcrystalline cellulose, and combinations thereof.

17. An animal feed additive comprising the anti-methanogenic composition of any one of Claims 1-16.

18. A nutritional preparation for an animal comprising the anti-methanogenic composition of any one of Claims 1-16.

19. A method for reducing methane emission, the method comprising administering the anti-methanogenic composition of any one of Claims 1-16, the animal feed additive of Claim 16, or the nutritional preparation of Claim 17 to a ruminant in an effective amount to reduce methane emission from the ruminant.

20. The method of Claim 19, wherein the ruminant is a cow, steer, bull, sheep, goat, or deer.

21. A process for preparing an anti-methanogenic composition, wherein the process comprises:i) combining an aqueous solution comprising a haloform with a glycol, alcohol, wax, oil, triglyceride, or combination thereof to extract the haloform from the aqueous solution into an organic phase;ii) mixing a hydrocarbon solvent with the organic phase;iii) separating the organic phase from the aqueous phase;iv.)    evaporating the hydrocarbon solvent from the organic phase; andv)     combining the organic phase with a hydrophobic polymer to form the anti-methanogenic composition.

22. The process of Claim 21, wherein the hydrocarbon solvent is hexane, heptane, pentane, petroleum ether, diethyl ether, toluene, or xylene.WO 2025 / 120610                                   PCT / IB2024 / 06235123. The process of Claim 21 or 22, wherein the hyper-crosslinked polymer comprises a styrene-divinylbenzene or poly(divinylbenzene) group with a macropore structure.

24. The process of any one of Claims 21-23, wherein the hydrophobic polymer is a hyper-5 crosslinked polymer having a surface area of about 300 to about 4000 m2 / g and / or a swelling capacity of at least 0.2 mL / g.

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