JPWO2019148250A5 - - Google Patents

Description

The present invention relates to a method of producing emulsions or suspensions from biomass. The invention also relates to the production of powders or products produced therefrom. The present invention also relates to emulsions or suspensions produced by the methods as described herein. The present invention also relates to powders produced by the methods as described herein or products produced therefrom.

Bioactive substances such as oxygen-sensitive fatty acids and oils are desirable ingredients for foods, supplements, and/or cosmetics. However, many oils and bioactive molecules are susceptible to oxidation and degradation reactions when exposed to a variety of environments (such as oxygen, heat, pH, or enzymes), making these raw materials unsuitable for ingestion or use. It may degrade before, it cannot be stored in a form suitable for ingestion or use, or it does not reach the desired site in the body after ingestion. Oxygen-sensitive oils include those containing polyunsaturated fatty acids such as omega-3, omega-6, or omega-9 fatty acids. Labile bioactive ingredients include those that are water soluble (such as polyphenols that are labile at high pH) or oil soluble (such as carotenes that are oxygen sensitive) or sparingly soluble in oil or water (such as resveratrol, curcumin) Ingredients included. 58.

Encapsulation has been used for the protection and delivery of lipophilic and hydrophilic bioactive agents, but delivery and selection of the optimal encapsulation system for delivery remains a challenge (Augustin and Sanguansri, 2015; McClements, 2015). Due to the health-promoting properties of omega-3 oils, there is great interest in stabilizing these oils as they are highly susceptible to oxidation (Sanguansri and Augustin, 2006; Drusch and Manino, 2007). ).

Methods of encapsulating oxygen-sensitive oils and bioactive substances are known, but these methods do not involve using purified or substantially purified proteins such as whey protein isolate, soy protein isolate, or casein and carbohydrates. ) is required and is not economical for many products.

Purified proteins and carbohydrates can be used alone or in combination as encapsulation matrices for the delivery of bioactive agents (Augustin and Hemar, 2009; Aditya et al., 2017). For example, starch is commonly used as a wall material for encapsulation (Hoyos-Leyva et al., 2018), and proteins are useful for delivery because they have many desirable functional properties that aid encapsulation. It has been found (Subirade and Chen, 2008; Livney, 2010). MicroMAX® microencapsulation technology uses purified proteins (preferably casein) and purified carbohydrates to produce an encapsulating medium for oils. Heated protein-carbohydrate hybrids (MicroMAX® microencapsulation technology) were found to be superior to corresponding physical protein and carbohydrate hybrids (WO 01/74175; Augustin et al. ., 2006). Purified proteins and carbohydrates used in such processes are selected to be colorless and flavorless and can be costly due to the purification steps involved in their isolation.

Therefore, there is a need for new formulations and processes for producing products containing oxygen-sensitive bioactives such as fatty acids and oils that are susceptible to degradation during storage, processing, and within the gastrointestinal tract.

The inventors have developed a method of producing an emulsion, suspension, or powder containing proteins and carbohydrates obtained from a single source.

In one aspect, the invention provides a method of producing an emulsion or suspension comprising:
i) obtaining an aqueous mixture containing proteins and carbohydrates from the biomass of a single species of organism ;
ii) optionally adding oil to the aqueous mixture;
iii) forming an emulsion or suspension comprising the bioactive agent and/or bioactive precursor.

In one embodiment, the aqueous mixture is an aqueous suspension.

In one aspect, the invention provides a method of producing a powder comprising an entrapped or encapsulated bioactive agent and/or bioactive precursor, comprising:
i) obtaining an aqueous mixture containing proteins and carbohydrates from the biomass of a single species of organism ;
ii) adding oil to the aqueous mixture;
iii) forming an emulsion or suspension comprising the bioactive agent and/or bioactive precursor;
iv) forming a powder comprising an entrapped or encapsulated bioactive agent and/or bioactive precursor from an emulsion or suspension.

In one embodiment, the invention provides a method as described herein, wherein the bioactive agent and/or bioactive precursor comprises
i) components of biomass,
ii) the oil of step ii) or a component thereof,
iii) components added to the oil before the oil is added to the aqueous mixture in step ii);
iv) ingredients injected into the oil before or during step ii);
v) further components of the biomass, and vi) components added in steps i), ii), and iii) of the method.

In one embodiment, the bioactive agents are i) and ii).

In one embodiment, the bioactive precursor is i).

In one embodiment, the bioactive agent is formed in or after step i) or in step ii).

In one aspect, the invention provides a matrix comprising proteins and carbohydrates from the biomass of a single organism .

In one aspect, the present invention provides a bioactive agent and/or bioactive precursor entrapped or encapsulated in a matrix, comprising proteins and carbohydrates of biomass from a single species of organism , wherein the entrapped or encapsulated organism A bioactive agent and/or bioactive precursor wherein the active agent and/or bioactive precursor is resistant to oxygen degradation when compared to the bioactive agent and/or bioactive precursor prior to entrapment or encapsulation. provide the body

In one aspect, the invention provides an emulsion or suspension produced by a method as described herein.

In one aspect, the invention provides an emulsion or suspension produced by a method as described herein.

In one aspect, the present invention provides a powder comprising entrapped or encapsulated bioactive agents and/or bioactive precursors and comprising proteins and carbohydrates from a single species of organism .

In one aspect, the invention provides a powder produced by a method as described herein.

In one aspect, the invention provides an emulsion or suspension produced by a method as described herein, a matrix as described herein, a matrix as described herein, Articles of manufacture are provided that include bioactive agents and/or bioactive precursors entrapped or encapsulated in a matrix, or emulsions or suspensions as described herein.

In one aspect, the invention provides an article of manufacture comprising a powder produced by a method as described herein or comprising a powder as described herein.

The inventors have also surprisingly discovered that lipid-based compositions improve the stability of isothiocyanates (such as sulforaphane) and/or isothiocyanate precursors (such as glucosinolates). Accordingly, in a further aspect, the invention provides pharmaceutical or cosmetic compositions comprising isothiocyanates and/or isothiocyanate precursors, lipids, and pharmaceutical and/or cosmetic excipients.

In one aspect, the invention provides a method of producing an emulsion comprising an isothiocyanate or isothiocyanate precursor, comprising:
A method is provided comprising providing a mixture comprising water, a lipid, and an isothiocyanate or isothiocyanate precursor, thereby forming an emulsion.

In one aspect, the invention provides an emulsion comprising water, lipids, and isothiocyanates and/or isothiocyanate precursors.

In one aspect, the invention provides a method of preparing a powder comprising isothiocyanates and/or isothiocyanate precursors, comprising preparing an emulsion as described herein; subjecting to drying conditions thereby removing water to form a powder.

In one aspect, the invention provides a method of preparing a pharmaceutical or cosmetic composition, comprising preparing an emulsion as described herein or A method is provided comprising preparing a powder and converting the emulsion or dry composition into a pharmaceutical or cosmetic composition.

In one aspect, the invention is a method of treating or preventing a condition, which requires administration of an effective amount of a pharmaceutical composition, emulsion, or powder as described herein. A method is provided that includes performing a subject.

In one aspect, the invention provides a pharmaceutical composition, emulsion or powder as described herein for use in treating or preventing a condition.

In one aspect, the invention provides a method of treating or preventing a condition in a subject, comprising administering to the subject an effective amount of a pharmaceutical composition, emulsion, or powder as described herein. A method is provided, comprising:

In one aspect the invention provides use of an emulsion as described herein or a powder as described herein in the manufacture of a medicament for the treatment of a condition.

In one aspect, the invention provides a method or use as described herein for cancer, diabetes, cardiovascular, autism, osteoporosis, neuroprotective diseases, inflammation, oxidative stress, and cancer. Provided is a method or use selected from gut health conditions.

Any embodiment herein is to be considered as applying mutatis mutandis to any other embodiment unless specifically stated otherwise. For example, as will be appreciated by those skilled in the art, examples of bioactive agents and/or bioactive precursors for the above methods of the invention are the emulsions, suspensions, powders, and products.

This invention is not to be limited in scope by the specific embodiments described herein, which are intended for illustrative purposes only. Functionally equivalent products, compositions and methods, as described herein, are clearly within the scope of the invention.

Throughout this specification, unless otherwise specified or required by context, reference to a single step, composition of matter, group of steps or group of compositions of matter refers to these steps, It shall be construed to include one and more (ie, one or more) of a composition of matter, group of steps, or group of compositions of matter.

The invention will now be described using the following non-limiting examples and with reference to the accompanying figures.

Oil-in-water emulsions using broccoli containing proteins and carbohydrates as encapsulating media compared to oil and water dispersions at A) 0 minutes and B) 120 minutes after emulsion creation. indicates the physical stability of This figure also shows an aqueous phase suspension using lyophilized broccoli powder, adding sufficient amount of water to achieve a fluid mixture (7.46% TS) prior to use as a mounting medium. shows the preparation. A) exhibits 14.29% total solids (TS); B) exhibits 10.64% TS; C) exhibits 8.48% TS; D) exhibits 7.46% TS. Figure 2 shows the preparation of an aqueous phase suspension using fresh broccoli by adding the required amount of water until a fluid mixture is achieved prior to use as an encapsulant. A) 7.66% TS, B) 6.87% TS, C) 6.23% TS, and D) 4.99% TS. Shown are emulsions containing omega-3 oil after A) conditioning, B) after overnight storage, and C) after lyophilization. F1 and F2 using broccoli as encapsulating agent, C1 using heated casein carbohydrate as encapsulating agent, and C2 using Tween as emulsifier. Oxipres showing oxygen induction period (IP)/uptake of sample [emulsion (9.5% TS, 4.8% oil) from FIG. 3A] tested at 80° C. and initial 5 bar oxygen pressure. Test results. The sample tested was 83 g emulsion (4 g matrix solids and 4 g oil in sample). IP(h) is only observed for samples using Tween as emulsifier and heated casein carbohydrate as encapsulating agent when there is a large change in oxygen consumption. Samples using broccoli as the mounting medium have no characteristic IP up to 20 hours when the test is stopped. The slow oxygen uptake in these samples is due in part to oxygen uptake by the broccoli matrix. Oxipres (see Figure 6) for unrefined tuna oil is 9 hours. Oxipres test results showing the IP/uptake of oxygen for a sample [lyophilized powder (50% tuna oil) from FIG. 3C] tested at 80° C. and an initial 5 bar oxygen pressure. The sample tested was 8 g powder (4 g matrix solids and 4 g oil in sample). Samples using broccoli as the mounting medium have no specific IP up to 43 hours when the test is stopped. The slow oxygen uptake in these samples is due in part to oxygen uptake by the broccoli matrix. The IP for unrefined tuna oil in the Oxipres data (see Figure 6) is 9 hours. Oxipres test results showing IP/uptake of oxygen for tuna oil, canola oil, and high DHA canola oil tested at 80° C. and an initial 5 bar oxygen pressure. A distinct IP is observed for each oil. Oxipres test with broccoli matrix (without oil) showing the effect of varying amounts of vegetable matrix on oxygen uptake. Oxipres test results for lyophilized omega-3 broccoli powder (12.5% tuna oil or canola oil) tested at 80° C. and initial 5 bar oxygen pressure. The total solids content of the emulsion before drying was 5.7%. The sample tested was 20 g powder (17.5 g matrix and 2.5 g oil). The slow oxygen uptake in these samples is due in part to oxygen uptake by the broccoli matrix. Oxipres test results for lyophilized omega-3 broccoli powder (25% tuna oil or DHA canola oil) tested at 80° C. and initial 5 bar oxygen pressure. The total solids content of the emulsion before drying was 6.6%. The sample tested was 10 g powder (7.5 g matrix and 2.5 g oil). The slow oxygen uptake in these samples is due in part to oxygen uptake by the broccoli matrix. IP(h) is the point at which there is a large increase in oxygen uptake (a rapid drop in oxygen tension). Oxipres test results for lyophilized omega-3 broccoli powder (50% tuna oil or DHA canola oil) tested at 80° C. and initial 5 bar oxygen pressure. The total solids content of the emulsion before drying was 9.5%. The sample tested was 5 g powder (2.5 g matrix and 2.5 g oil). The slow oxygen uptake in these samples is due in part to oxygen uptake by the broccoli matrix. Oxipres test results showing oxygen uptake of omega-3 broccoli emulsion samples tested at 80° C. and initial 5 bar oxygen pressure. The emulsion was prepared at 4% aqueous solids (3.8% oil and 7.7% total solids) and 6% aqueous solids (5.7% oil and 11.3% total solids). minutes) at 75° C.-2 min and 100° C.-30 min). The sample tested contained 4g of oil and 4g of matrix. Emulsions using broccoli as the encapsulating medium have no specific IP up to 42 hours. The slow oxygen uptake in these samples is due in part to oxygen uptake by the broccoli matrix. Oxipres test results showing IP of lyophilized omega-3 broccoli powder (50% tuna oil) tested at 80° C. and initial 5 bar oxygen pressure. Samples were prepared by two heat treatments (75° C.-2 min and 100° C.-30 min) at 5% and 6% aqueous solids (5.7% oil and 11.3% total solids). bottom. The sample tested contained 4g of oil and 4g of matrix. Oxipres test results showing IP of lyophilized omega-3 broccoli powder (50% tuna oil) tested at 80° C. and initial 5 bar oxygen pressure. Broccoli mounting medium was subjected to two heat treatments (75° C.-2 min and 100° C.-30 min) and either used as is (“fresh broccoli” without drying) or “lyophilized broccoli”. Reconstituted from powder. IP(h) is the point at which there is a large increase in oxygen uptake (a rapid drop in oxygen tension). Oxipres test results showing oxygen uptake of omega-3 broccoli emulsion samples tested at 80° C. and initial 5 bar oxygen pressure. Broccoli encapsulant was used at various processing stages to produce up to 5% aqueous solids. An emulsion was prepared with 9.5% TS and 4.8% oil. IP(h) is the point at which there is a large increase in oxygen uptake (a rapid drop in oxygen tension). The sample tested contained 4g of oil and 4g of matrix. The slow oxygen uptake in these samples is due in part to oxygen uptake by the broccoli matrix. Oxipres test results showing IP of lyophilized omega-3 carrot powder (50% tuna oil) tested at 80° C. and initial 5 bar oxygen pressure. Results showing the two heat treatments used (75° C.-2 min and 100° C.-30 min). The total solids content of the emulsion before drying was 9.5%. The sample tested was 8g powder (4g matrix and 4g oil). No clear IP. Oxygen uptake rates above IP cannot be obtained as a sudden increase in pressure has been shown resulting in the release of volatiles (marked IP). IP(h) is the point at which there is a large increase in oxygen uptake (a rapid drop in oxygen tension). A sharp peak is evidence of an interaction that resulted in a significant increase in pressure. Oxipres test showing IP of omega-3 carrot powder (50% tuna oil) using 'fermented' and 'non-fermented' carrots as encapsulant for omega-3 oil tested at 80°C and initial 5 bar oxygen pressure. result. The total solids content of the emulsion before drying was 9.5%. The sample tested was 8g powder (4g matrix and 4g oil). Oxygen uptake rates above IP cannot be obtained as a sudden increase in pressure has been shown resulting in the release of volatiles (marked IP). A sharp peak is evidence of an interaction that resulted in a significant increase in pressure. There is no clear IP for unfermented carrots as an encapsulant. IP(h) is the point at which there is a large increase in oxygen uptake (a rapid drop in oxygen tension). Oxipres test results showing IP of freeze-dried omega-3 tomato powder (50% tuna oil) tested at 80° C. and initial 5 bar oxygen pressure. No clear IP. Oxygen uptake rates above IP cannot be obtained as a sudden increase in pressure has been shown resulting in the release of volatiles (marked IP). IP(h) is the point at which there is a large increase in oxygen uptake (a rapid drop in oxygen tension). A sharp peak is evidence of an interaction that resulted in a significant increase in pressure. Results showing the two heat treatments used (75° C.-2 min and 100° C.-30 min). The total solids content of the emulsion before drying was 9.5%. The sample tested was 8g powder (4g matrix and 4g oil). Increasing the temperature-time treatment for tomatoes results in longer IP (better protection of omega-3 oil from oxidation). Oxipres test results showing IP of lyophilized omega-3 mushroom powder (25% and 50% oil) tested at 80° C. and initial 5 bar oxygen pressure. There is no clear IP(h) with a large increase in oxygen uptake (rapid drop in oxygen tension). Results showing two heat treatments (75° C.-2 min and 100° C.-30 min) of mushroom as encapsulant for 50% oil powder. The total solids content of the emulsion before drying was 9.5%. The samples tested were 8 g powder for 50% oil powder (4 g matrix and 4 g oil) and 12 g powder for 25% oil powder (9 g matrix and 3 g oil). Low oil loading (25% oil) results in longer IP (better protection of omega-3 oils from oxidation) compared to 50% oil powder. Oxipres test results showing IP of lyophilized omega-3 cauliflower powder (25% and 50% oil) tested at 80° C. and initial 5 bar oxygen pressure. IP was observed for 50% oil powder, but oxygen uptake rate above IP cannot be obtained because there is no clear IP for 25% oil powder. IP(h) is the point at which there is a large increase in oxygen uptake (a rapid drop in oxygen tension). (75° C.-2 min) and results showing heat treatment of cauliflower as encapsulant at two oil loadings (50% and 25% oil). The total solids content of the emulsion before drying was 9.5%. The samples tested were 8 g powder for 50% oil powder (4 g matrix and 4 g oil) and 12 g powder for 25% oil powder (9 g matrix and 3 g oil). Oxipres test results showing the IP of lyophilized omega-3 kale powders (25% and 50% oil) tested at 80° C. and an initial 5 bar oxygen pressure. IP was observed for 25% oil powder, but oxygen uptake above IP cannot be obtained because there is no clear IP for 50% oil powder. IP(h) is the point at which there is a large increase in oxygen uptake (a rapid drop in oxygen tension). The sharp peak for 50% oil powder is evidence of an interaction that resulted in the significant increase in pressure observed for 50% tuna oil powder. (75° C.-2 min) and results showing heat treatment of kale as an encapsulant at two oil loadings (50% and 25% oil). The total solids content of the emulsion before drying was 9.5%. The samples tested were 8 g powder for 50% oil powder (4 g matrix and 4 g oil) and 12 g powder for 25% oil powder (9 g matrix and 3 g oil). Oxipres test results showing the IP of lyophilized omega-3 Brussels sprout powder (25% and 50% oil) tested at 80° C. and an initial 5 bar oxygen pressure. IP was observed for 50% oil powder, but oxygen uptake rate above IP cannot be obtained because there is no clear IP for 25% oil powder. IP(h) is the point at which there is a large increase in oxygen uptake (a rapid drop in oxygen tension). (75° C.-2 min) and results showing heat treatment of Brussels sprouts as an encapsulant at two oil loadings (50% and 25% oil). The total solids content of the emulsion before drying was 9.5%. The samples tested were 8 g powder for 50% oil powder (4 g matrix and 4 g oil) and 12 g powder for 25% oil powder (9 g matrix and 3 g oil). Oxipres test results showing IP of lyophilized omega-3 snow pea powder (25% and 50% oil) tested at 80° C. and initial 5 bar oxygen pressure. Oxygen uptake rates above the IP cannot be obtained because there is no definite IP. IP(h) is the point at which there is a large increase in oxygen uptake (a rapid drop in oxygen tension). The sharp peak for 50% oil powder is evidence of an interaction that resulted in the significant increase in pressure observed for 50% tuna oil powder. (75° C.-2 min) and results showing heat treatment of snow peas as an encapsulant at two oil loadings (50% and 25% oil). The total solids content of the emulsion before drying was 9.5%. The samples tested were 8 g powder for 50% oil powder (4 g matrix and 4 g oil) and 12 g powder for 25% oil powder (9 g matrix and 3 g oil). Oxipres test results showing IP of lyophilized omega-3 garlic powder (25% and 50% oil) tested at 80° C. and initial 5 bar oxygen pressure. IP was observed for 25% oil powder, but oxygen uptake above IP cannot be obtained because there is no clear IP for 50% oil powder. IP(h) is the point at which there is a large increase in oxygen uptake (a rapid drop in oxygen tension). The sharp peak for 50% oil powder is evidence of an interaction that resulted in the significant increase in pressure observed for 50% tuna oil powder. (75° C.-2 min) and results showing heat treatment of garlic as an encapsulant at two oil loadings (50% and 25% oil). The total solids content of the emulsion before drying was 9.5%. The samples tested were 8 g powder for 50% oil powder (4 g matrix and 4 g oil) and 12 g powder for 25% oil powder (9 g matrix and 3 g oil). of lyophilized omega-3 carrot powder (25% tuna oil) supplemented with vegetable protein (pea protein, soy protein (SPI)) or milk protein, sodium caseinate, tested at 80°C and an initial 5 bar oxygen pressure. Oxipres test results. The total solids content of the emulsion before drying was 9.5% (2.4% oil). IP(h) is the point at which there is a large increase in oxygen uptake (a rapid drop in oxygen tension). The sample tested was 10 g powder (2.5 g oil and 7.5 g matrix). IP and oxygen uptake cannot be obtained, except for carrots with pea protein added as encapsulant, as a sudden increase in pressure was noted, resulting in the release of volatiles. A sharp peak is evidence of an interaction that resulted in a significant increase in pressure. IP(h) is the point at which there is a large increase in oxygen uptake (a rapid drop in oxygen tension). Oxipres test showing the IP of spray-dried (25% tuna oil) omega-3 matcha powder with different protein:carbohydrate ratios tested at 80° C. and an initial 5 bar oxygen pressure compared to that of unrefined tuna oil. result. Matcha powder was reconstituted from a commercial sample of matcha (green tea) powder. IP was observed for tuna oil powder only and powders encapsulated at protein to carbohydrate ratios of 1:3 and 1:4, but encapsulated at protein to carbohydrate ratios of 1:2 and 8:9. Since there is no definite IP for powders, it is not possible to obtain an oxygen uptake rate exceeding the IP. The encapsulant containing the 8:9 protein to carbohydrate ratio was of matcha powder only. IP of spray-dried (50% tuna oil) omega-3 broccoli powder tested at 80° C. and initial 5 bar oxygen pressure compared to that of 50% tuna oil powder using heated casein-carbohydrate as encapsulant. Oxipres test results showing. Broccoli mounting medium was reconstituted from "lyophilized broccoli" powder. Oxipres test results showing the IP of lyophilized (50% tuna oil) omega-3 oil powder using broccoli puree and fermented broccoli puree as encapsulating media tested at 80° C. and an initial 5 bar oxygen pressure. Quantitative analysis of secondary lipid oxidation in freeze-dried powders (50% tuna oil) encapsulated in various vegetable matrices after storage at 40°C for 4 weeks. Vegetable encapsulants (left to right: broccoli, carrots, fermented carrots, tomatoes, mushrooms, cauliflower, kale, Brussels sprouts, snow peas, garlic). Induction period (IP ) in Oxipres test results. There is no definite IP(h) for tableted formulations, but a definite IP is given for extruded formulations. The samples tested were 40 g for extrudate (36 g matrix and 4 g oil) and 16 g for tablet format (16 g excipients and 4 g oil). Oxipres test results showing the induction period (IP) of lyophilized omega-3 broccoli powder (50% oil) tested at 80° C. and an initial 5 bar oxygen pressure. There is no clear IP(h) with a large increase in oxygen uptake (rapid drop in oxygen tension). Results showing pre-treatment (using ultrasound or microwave) or post-emulsification treatment (using HPP or microwave) to the aqueous phase. The total solids content of the aqueous phase was 5%, and the total solids content of the emulsion was 9.5%. The sample tested was 8g powder (4g matrix and 4g oil).

General Techniques and Definitions Unless specifically defined otherwise, all technical and scientific terms used herein shall be deemed to have the same meaning as commonly understood by one of ordinary skill in the art. .

The term "and/or", e.g., "X and/or Y", is understood to mean either "X and Y" or "X or Y", both meanings or either meaning being expressly should be construed to include

Throughout this specification, the word "comprise", or variations such as "comprises" or "comprising" refer to a given element, integer or step or group of elements, integers or steps. is not meant to exclude any other element, integer or step, or group of elements, integers or steps.

As used herein, the term "about," unless stated to the contrary, is +/- 10%, more preferably +/- 5%, even more preferably +/- 1% of the specified value. point to

As used herein, "component" refers to a part or element of a larger whole.

As used herein, "protein" or "polypeptide" refers to macromolecules containing carbon, hydrogen, oxygen, nitrogen, and usually sulfur, comprising polymers of amino acids linked together by peptide bonds. Point.

As used herein, "carbohydrate" refers to a class of molecules of general formula Cx(H2O)y.

As used herein, the term "resistant to oxygen degradation," or similar phrases, refers to reducing the susceptibility of biologically active substances, such as fatty acids, to oxidation. In one embodiment, the susceptibility of a substance to oxidation is reduced by entrapping or encapsulating the substance to reduce exposure to oxygen. In one embodiment, this involves entrapping or encapsulating the substance with molecules that have oxygen sequestering capabilities. Evaluation of oxidation resistance can be performed by any method known to those skilled in the art. For example, the oxidation resistance of fats and oils can be based on oxidation of the oil by oxygen under pressure. In such tests, the consumption of oxygen causes a pressure drop during the test, which is due to the uptake of oxygen by the sample during oxidation. Performing at high pressure and temperature accelerates the oxidation rate. In one embodiment, oxidative resistance is assessed using Oxipres (eg, Mikrolab Aarhus A/S apparatus Hojbjerg, Denmark). In one embodiment, emulsions, suspensions, and/or powders containing fats or oils (eg, polyunsaturated oils) are exposed to elevated temperatures and oxygen pressures. In one embodiment, oxidation resistance is evaluated at 80° C. and an initial oxygen pressure of 5 bar. In one embodiment, the induction period (IP,h) associated with the oxidative stability of the sample is determined. A longer IP(h) indicates that the sample is more resistant to oxidation during storage (more stable in the presence of oxygen). Other methods for measuring lipid oxidation include, for example, peroxide value, para-anisidine value, headspace analysis of volatiles (e.g., propanal and EE, which are secondary oxidation products from oxidation of omega-3 fatty acids). -Aldehydes such as 2,4-heptadienal), and changes in the % of individual unsaturated fatty acids (eg, EPA and DHA) in the stored samples. In one embodiment, oxidation is not necessarily related to solvent extractable free fat (ie, free fat level is not an indicator of oil IP or susceptibility to oxidation in powder).

As used herein, "thermolysis," or similar phrases, refers to the degradation of bioactive substances (eg, fatty acids) or bioactive precursors due to exposure to low or high temperatures. In one embodiment, the susceptibility to degradation by temperature is measured during the method of producing an emulsion, suspension, or powder as described herein, by adding a bioactive agent or bioactive agent to a protein or carbohydrate. Decreased by precursor binding.

As used herein, "moisture degradation," or similar phrases, refers to the degradation of bioactive substances (eg, fatty acids) or bioactive precursors due to exposure to low or high humidity. In one embodiment, susceptibility to degradation by moisture reduces the bioactive agent or bioactivity of the protein or carbohydrate during the method of producing an emulsion, suspension, or powder as described herein. Decreased by precursor binding.

As used herein, the term "pH degradation," or similar phrases, refers to the degradation of bioactive substances (e.g., fatty acids) or bioactive precursors due to exposure to low or high pH. . In one embodiment, low pH is pH<7. In one embodiment, high pH is pH>7. In one embodiment, susceptibility to degradation by pH is measured during the process of producing emulsions, suspensions, or powders as described herein bioactive substances or bioactive substances to proteins or carbohydrates. Reduced by binding of precursors (eg, phytonutrients).

As used herein, the term "photodegradation", or like phrases, refers to the degradation of bioactive substances (eg, carotenoids) or bioactive precursors due to exposure to light. In one embodiment, susceptibility to photodegradation is determined by bioactive substances or bioactive precursors to proteins or carbohydrates during methods of producing emulsions, suspensions, or powders as described herein. Lowered by body binding.

As used herein, "capturing" or "captured" or "capturing" refers to bioactive substances, such as phytonutrients, or bioactive precursors, as described herein. Refers to binding or partitioning into one or more components of an emulsion, suspension, or encapsulant matrix. In one embodiment, the component is a carbohydrate or protein. In one embodiment, scavenging increases the resistance of the bioactive agent or bioactive precursor to degradation by one or more of oxygen, temperature, pH, moisture, and light.

As used herein, "encapsulated" or "encapsulated" refers to the lipid and lipid in the emulsion, suspension, or encapsulant matrix of the present invention or produced by the methods of the present invention. Refers to the formation of a functional barrier around a bioactive agent or bioactive precursor such as a lipid soluble component. In one embodiment, encapsulation increases the resistance of the bioactive agent or bioactive precursor to degradation by one or more of oxygen, temperature, pH, moisture, and light.

As used herein, " species " refers to a subdivision of a genus. In one embodiment, " species " refers to a group of organisms consisting of individuals that can breed among themselves.

As used herein, "polyphenol" refers to compounds containing more than one phenolic hydroxyl group. In one embodiment, the polyphenols are anthocyanins, dihydrochalcone, flavan-3-ols, flavanones, flavones, flavonols and isoflavones, curcumin, resveratrol, benzoic acid, phenylacetic acid, hydroxycinnamic acid, coumarins, naphthoquinones, xanthones, stilbenes , chalcones, tannins, phenolic acids, and catechins such as epigallocatechin gallate (EGCg), epigallocatechin (EGC), epicatechin gallate (ECg), epicatechin (EC), and their geometric isomers gallocatechin gallate (GCg), gallocatechin (GC), gallocatechin gallate (Cg), and catechin).

As used herein, "bioactive organic sulfur-containing compound(s)" includes glucosinolates, isothiocyanates, and allium compounds (e.g., alliin, allicin, ajoene, allylpropyldisulfide, diallyltrisulfide). , salylcysteine, vinyldithyin, S-allylmercaptocysteine).

Biomass The present invention produces emulsions, suspensions, powders, or products produced therefrom, at least in part from biomass comprising proteins and carbohydrates from a single (first) species of organism . about how to Proteins and carbohydrates are therefore not separated from each other prior to use in the method of the invention.

In some embodiments, whole biomass can be used, while in other embodiments the biomass has been treated to remove or reduce the concentration of one or more components of the biomass. there is In one embodiment, less than about 50% of the biomass is removed prior to use in the methods of the invention. In one embodiment, less than about 40% of biomass is removed prior to use in the methods of the invention. In one embodiment, less than about 30% of biomass is removed prior to use in the methods of the invention. In one embodiment, less than about 20% of biomass is removed prior to use in the methods of the invention. In one embodiment, less than about 10% of biomass is removed prior to use in the methods of the invention. In one embodiment, less than about 5% of biomass is removed prior to use in the methods of the invention. In one embodiment, less than about 1% of biomass is removed prior to use in the methods of the invention. In one embodiment, biomass is not removed prior to use in the methods of the invention.

In one embodiment, the biomass is dried or concentrated to remove water. In one embodiment, drying removes from about 60% to about 90% by weight of biomass. In one embodiment, drying removes about 70% to about 90% of the weight of the biomass. In one embodiment, drying removes about 80% to about 90% of the weight of the biomass.

In one embodiment, the biomass contains proteins and carbohydrates from only a single species of organism (no proteins or carbohydrates from additional species ). In one embodiment, the biomass further comprises proteins and carbohydrates from one or more additional species (eg, second, third, fourth, fifth, etc. species of organisms ). Proteins and carbohydrates from additional species, as well as biomass from the first species, have not been separated from one another prior to use in the method of the present invention. In one embodiment, the biomass further comprises proteins and carbohydrates from a second species . In one embodiment, the biomass further comprises proteins and carbohydrates from second and third species . In one embodiment, the biomass further comprises proteins and carbohydrates from second, third and fourth species .

In one embodiment, the biomass and/or further biomass comprises fibers that have not been separated from the proteins and carbohydrates of the biomass and/or further biomass.

In one embodiment, the biomass and/or further biomass comprises catechins that have not been separated from the proteins and carbohydrates of the biomass and/or further biomass.

In one embodiment, the biomass and/or further biomass has a protein to carbohydrate ratio of 1:1 to 1:10.5. In one embodiment, the biomass and/or further biomass has a protein to carbohydrate ratio of about 1:4.5 to 4:1. In one embodiment, the biomass and/or further biomass has a protein to carbohydrate ratio of about 1:2.5 to 2:1. In one embodiment, the biomass and/or further biomass has a protein to carbohydrate ratio of about 1:2.4. In one embodiment, the biomass and/or further biomass additionally comprises fibers. In one embodiment, the biomass or further biomass has a protein to carbohydrate ratio as shown in Table 1.

In one embodiment, the biomass or further biomass is green tea leaf powder (maccha). In one embodiment, matcha (about 2% moisture) is about 35.5% protein, about 39.6% carbohydrate, about 5.9% fat, and about 6.0% fat on a dry basis. including. In one embodiment, matcha contains about 13.1% catechins.

In one embodiment, protein is added to the biomass and/or additional biomass to form a protein to carbohydrate ratio of about 1:1 to 1:10.5. In one embodiment, protein is added to the biomass and/or additional biomass to form a protein to carbohydrate ratio of about 1:4.5 to 4:1. In one embodiment, protein is added to the biomass and/or additional biomass to form a protein to carbohydrate ratio of about 1:2.5 to 2:1.

In one embodiment, carbohydrate is added to the biomass and/or additional biomass to form a protein to carbohydrate ratio of about 1:1 to 1:10.5. In one embodiment, carbohydrate is added to the biomass and/or additional biomass to form a protein to carbohydrate ratio of about 1:4.5 to 4:1. In one embodiment, carbohydrate is added to the biomass and/or additional biomass to form a protein to carbohydrate ratio of about 1:2.5 to 2:1.

In one embodiment, biomass can be a whole organism or one or more parts thereof.

In one embodiment, the biomass and/or further biomass comprises whole biomass (or pieces thereof) in fresh/raw or dried form. In one embodiment, the biomass and/or further biomass is fresh/raw. In one embodiment, the biomass and/or additional biomass is pretreated as described herein.

In one embodiment, the biomass and/or further biomass is subjected to extraction or separation processes as described herein suitable for removing one or more components from the biomass and/or from the further biomass. It is a thing.

In one embodiment, the biomass and/or further biomass comprises bioactive substances. In one embodiment, the biomass and/or further biomass comprises bioactive precursors.

In one embodiment, bioactive agents and/or bioactive precursors are added to the biomass or to additional biomass.

In one embodiment, the biomass and/or further biomass is eukaryotic. In one embodiment, the biomass and/or further biomass is prokaryotic (eg, algae). In one embodiment, the biomass and/or further biomass is from Plantae or the fungi kingdom.

The material can be any part of a Plantae or fungus, including, but not limited to, one or more of leaves, stems, flowers, florets, seeds, and roots, where relevant.

In one embodiment, the Plantae is Brassicaceae. As used herein, "Brassicaceae" refers to members of the Brassicaceae family, commonly referred to as mustard, cruciferous plants, or the cabbage family.

In one embodiment, the Brassicaceae is selected from the genus Brassica or Cardamine. In one embodiment, the Brassica is Brassica balearica, Brassica carinata, Brassica elongate, Brassica fruticulosa, Brassica hilarionis, Brassica juncea, Brassica napus (rapeseed or canola), Brassica sica narinosa, Brassica nigra, Brassica oleracea, Brassica perviridis, Brassica rapa, Brassica rupestris, Brassica septiceps, and Brassica tournefortii.

In one embodiment, the Brassica is Brassica oleracea.

In one embodiment, the Brassica is Brassica napus (rapeseed or canola).

In one embodiment, the Brassica is Brassica oleracea var. oleracea (wild cabbage), Brassica oleracea var. capitate (cabbage), Brassica rapa subsp. pobrassica (rutabaga), Brassica rapa var. rapa (turnip), Brassica oleracea var. alboglabra (Chinese kale), Brassica oleracea var. viridis (collared greens), Brassica oleracea var. longata (Jersey cabbage), Brassica oleracea var. acephal a (habotan), Brassica oleracea var sabellica (kale), Brassica oleracea var. palmifolia (Carvolonero), Brassica oleracea var. ramose (perpetual kale), Brassica oleracea var. medullosa (marrow cabbage), Brassica oler acea var. costata (tronchuda kale), Brassica oleracea Brassica oleracea var. italica (broccoli), Brassica oleracea var. botrytis (cauliflower, Romanesco broccoli, broccoli ditor bole)), Brassica oleracea var. botrytis × italica ( brocco flour), and one or more of Brassica oleracea var. italica x alboglabra (stick senor). In one embodiment, the Brassica oleracea is kale.

In one embodiment, the Brassica is Brassica oleracea var. italica (broccoli).

In one embodiment, the Brassica is Brassica oleracea var. botrytis (cauliflower).

In one embodiment, the Brassica is Brassica oleracea var. gemmifera (Brussels sprouts).

In one embodiment, Brassicaceae is Cardamine hirsuta, Iberis sempervirens, Sinapis arvensis, Armoracia rusticana, Pringlea antiscorbutica cabbage), Thlaspi arvense, Raphanus raphanistrum Species sativus (radish), Eruca sativa (radish), Anastatica hierochuntica (rose of Jericho), Crambe maritima (hamana), Cakile maritima (radish), Capsella bursa-pastoris (shepherd's purse), sweet alyssum, Arabidopsis thaliana , Nasturtium officinale (water mustard), Sinapis alba (white mustard), Erophila verna (whitlow grass), Raphanus raphanistrum (wild radish), Isatis tinctoria (hosoba rotunda), and Nasturtium microphyllum selected from one or more of be done.

In one embodiment, the Plantae is Cannabis. In one embodiment, the Cannabis is Cannabis sativa (hemp).

In one embodiment the Plantae is a fruit or vegetable. In one embodiment, the fruit is selected from one or more of simple, clustered, and multiple fruits. In one embodiment, the fruit or vegetable is from the family Umbelliferae, Asparagaceae, Arecaceae, Myrtaceae, Rosaceae, Musaceae, Ericaceae, Saxifragaceae, Cucurbitaceae, Nightshade, Family Capparaceae, Family Adoxaceae, Family Vitaceae, Rutaceae Family, Actinidiaceae, Sapindaceae, Anacardiaceae, Moraceae, Oleaceae, Cactaceae, Passifloraceae, Bromeliaceae, Cactaceae, Lythraceae, Polygonac They are from the families eae, Cucurbitaceae, Oxalidaceae, and Caesalpinioideae.

In one embodiment, Umbelliferae is a carrot.

In one embodiment, the Asparagaceae is asparagus.

In one embodiment, Polygonaceae is selected from one or more of buckwheat, sorrel, and rhubarb.

In one embodiment, Cucurbitaceae is selected from one or more of cucumber, pumpkin, squash, and zucchini.

In one embodiment, the fruit is apple, apricot, avocado, banana, bilberry, blackberry, blackcurrant, blueberry, coconut, gooseberry, cherry, cherimoya, clementine, cloudberry, damson, durian, elderberry, fig, feijoa, Gooseberry, Grape, Grapefruit, Guava, Huckleberry, Jackfruit, Jumble, Jujube, Kiwifruit, Kumquat, Lemon, Lime, Loquat, Lychee, Mandarin, Mango, Melon, Cantaloupe, Honeydew, Watermelon, Nectarine, Orange, Passionfruit , pawpaw, peach, pear, plum, plumcot, pineapple, pomegranate, pomelo, purple mangosteen, raspberry, rambutan, red currant, satsuma, starfruit, strawberry, tangerine, tomato, and agrifruit be done.

In one embodiment, a Plantae is a Compositae. In one embodiment, the Compositae are selected from one or more of artichoke, chamomile, chicory, dandelion, endive, Jerusalem artichoke, lettuce, romaine lettuce, safflower balamondin, and sunflower.

In one embodiment, the Plantae is Amarantaceae/Chenopodiacae. In one embodiment, the Amaranthaceae/Chenopodiacae are selected from one or more of amaranth, beet, chard, chloza, quinoa, spinach, and sugar beet.

In one embodiment, the Plantae is Malvaceae. In one embodiment, Malvaceae is selected from one or more of cocoa, cotton, and okra.

In one embodiment, the Plantae is from the family Amarylidaceae. In one embodiment, the Amarylidaceae is from the subfamily Allioideae. In one embodiment, the Allioideae is from the genus Allium. In one embodiment, the Allium is selected from one or more of Allium sativum (garlic), Allium cepa (onion), Allium ampeloprasum (leeks), Allium schoenoprasum (chives), and Allium oschaninii (shallots).

In one embodiment, Allium is Allium sativum (garlic).

In one embodiment, the Plantae is from the family Fabaceae. In one embodiment, the Fabaceae are soy alfalfa, beans, carob, chickpeas, kidney beans, jicama, lentils, peas, snow peas, and peanuts.

In one embodiment, the Fabaceae is snow peas.

In one embodiment, Plantae is a grain. In one embodiment, the grain is an ancient grain. In one embodiment, the grain is rice, corn, wheat, triticale, barley, millet, sorghum, spelt, oats, freeka, bulgur, sorghum, emmer wheat, einkorn, teff, emmer, and/or buckwheat. selected from one or more.

In one embodiment, the Plantae is from the Arecaceae family. In one embodiment, Arecaceae is coconut palm. In one embodiment, the biomass and/or further biomass is coconut drupe.

In one embodiment, Plantae is a grass. In one embodiment, the grass is from the Poaceae family. In one embodiment, the grass is selected from one or more of bamboo, lemongrass, sugarcane, corn, and wheatgrass.

In one embodiment, the Plantae is from the Camellia sinensis family. In one embodiment, Camellia sinensis is green tea leaf (Matcha).

In one embodiment, the fungi are mushrooms. In one embodiment, the fungi are of the families Boletaceae, Cantharellaceae, Tricholomataceae, Cortinariaceae, Cantharellaceae, Meripilaceae, Discinaceae, Pleurotaceae, Tricholomataceae, and It is from the Tuberaceae family.

In one embodiment, the fungus is Boletus edulis, Cantharellus cibarius, Cantharellus tubeaeformis, Clitocybe nuda, Cortinarius caperatus, Craterellus cornucopioides, Grifola frondosa, Hericium erin aceus, Hydnum repandum, Lactarius deliciosus, Morchella conica var. deliciosa, Morchella esculenta var. selected from one or more of rotunda, Pleurotus ostreatus, Tricholoma matsutake, Tuber brumale, Tuber indicum, Tuber macrosporum, Tuber mesentericum, and Tuber aestivum.

In one embodiment, the biomass and/or further biomass is not animal biomass or animal-generated products. In one embodiment, the biomass and/or further biomass is non-avian. In one embodiment, the biomass and/or further biomass is not bone or bone marrow. In one embodiment the biomass and/or further biomass is not animal milk.

In one embodiment, the biomass and/or further biomass is not milk, skim milk, or refined milk proteins and carbohydrates.

In one embodiment, the biomass and/or further biomass is Plantae or fungal material that does not meet cosmetic retail standards or is no longer suitable for fresh sale but is still edible.

Bioactive Agent As used herein, “bioactive agent” refers to a substance that has a biological effect. In one embodiment, the bioactive agent is susceptible to degradation by one or more of oxygen (oxidation), temperature, pH, moisture, and light. In one embodiment, the bioactive agent is an oil, or an oil-soluble agent.

In one embodiment, the bioactive agents are fatty acids, isothiocyanates, quercetin, allicin, ajoene, vitamin A, vitamin D, vitamin E, tocopherols, tocotrienols, vitamin K, beta carotene, lycopene, lutein, zeaxanthin, stigmasterol, selected from one or more of beta-sitosterol, campesterol, antioxidants, coenzyme Q10, astaxanthin, cannabinoids, cannabidiol, and polyphenols.

In one embodiment, the bioactive agent is selected from one or more of quercetin, allicin, phenolic acid. In one embodiment, the bioactive agent is allicin. In one embodiment, the bioactive agent is ajoene.

In one embodiment, the bioactive agent is a polyphenol. In one embodiment, the polyphenols are selected from one or more of catechins, flavonols, flavanols, anthocyanadins, resveratrol, and/or curcumin. Additional polyphenols are described herein.

In one embodiment, the bioactive agent is an isothiocyanate. As used herein, "isothiocyanate" refers to sulfur-containing phytochemicals having the general structure RN=C=S, which are the products of myrosinase activity on glucosinolates and their biological It is an active derivative. In one embodiment, the isothiocyanate is sulforaphane (1-isothiocyanato-4-methylsulfinylbutane). In one embodiment, the isothiocyanate is allyl isothiocyanate (3-isothiocyanato-1-propene). In one embodiment the isothiocyanate is benzyl isothiocyanate. In one embodiment the isothiocyanate is phenethyl isothiocyanate. In one embodiment the isothiocyanate is 3-butenyl isothiocyanate. In one embodiment the isothiocyanate is 5-vinyl-1,3-oxazolidine-2-thione. In one embodiment the isothiocyanate is 3-(methylthio)propylisothiocyanate. In one embodiment the isothiocyanate is 3-(methylsulfinyl)-propylisothiocyanate. In one embodiment the isothiocyanate is 4-(methylthio)-butyl isothiocyanate. In one embodiment the isothiocyanate is 1-methoxyindole-3-carbinol isothiocyanate. In one embodiment, the isothiocyanate is 2-phenylethyl isothiocyanate (also known as phenylethyl isothiocyanate or PEITC). In one embodiment the isothiocyanate is iverine.

In one embodiment, when the bioactive agent is an isothiocyanate, the biomass and/or further biomass further comprises one or more isothiocyanate bioactive derivative(s) or oligomers thereof. In one embodiment, the isothiocyanate bioactive derivative is any derivative of an isothiocyanate as described herein. In one embodiment, the isothiocyanate bioactive derivative is a derivative of sulforaphane. In one embodiment, the isothiocyanate bioactive derivative is indole-3-carbinol. In one embodiment, the isothiocyanate bioactive derivative is methoxy-indole-3-carbinol. In one embodiment, the isothiocyanate bioactive derivative is ascorbigen. In one embodiment, the isothiocyanate bioactive derivative is neoascorbigen.

In one embodiment, the bioactive agent is a component of biomass. In one embodiment, the bioactive agent is a component of additional biomass. In one embodiment, the bioactive agent is not present in the biomass or additional biomass and is added before, during, or after preparation of the aqueous mixture as described herein. In one embodiment, the bioactive agent is added before, during or after step ii) of the method as described herein. In one embodiment, the bioactive agent is added before or during step iii) of the method as described herein. In one embodiment, the bioactive agent is the oil of step ii) or a component thereof. In one embodiment, the bioactive agent is an ingredient added to the oil before it is added to the aqueous mixture suspension in step ii). In one embodiment, the bioactive agent is an ingredient that is injected into the oil before or during step ii). In one embodiment the bioactive agent is formed in step i) or after step i) or in step ii).

In one embodiment, the bioactive agent is a synthetically produced bioactive agent. In one embodiment, the bioactive agent is a synthetically produced isothiocyanate. In one embodiment, the bioactive agent is synthetically produced sulforaphane.

In one embodiment, if the biomass and/or further biomass comprises: i) Brassicaceae, the bioactive agent is an isothiocyanate; ii) if Brassicaceae is included, the bioactive precursor is a glucosinolate; ) if containing onion, the bioactive agent is one or more of quercetin, allicin, and phenolic acid; and iv) if containing garlic, the bioactive agent is one or more of allicin and ajoene. or v) fruits and/or vegetables containing polyphenols. In one embodiment, the Brassicaceae is broccoli and the isothiocyanate is sulforaphane. In one embodiment, the biomass and/or bioactives from further biomass are infused into the oil in step ii) or step iii) of the method as described herein.

In one embodiment, the bioactive agent is a phytonutrient. As used herein, "phytonutrients" refer to plant-derived substances associated with positive health effects. In one embodiment, the biomass and/or further biomass as described herein comprises one or more phytonutrient(s). In one embodiment, the phytonutrients are betalains, indoles, organic sulfides, phenols, terpenes, triterpenes, carotenoids, curcuminoids, flavonoids, glucosinolates, isothiocyanates, hydroxycinnamic acids, lignans, lipids, stilbenes, sulfides, tocopherols, of lutein, zeantin, isoflavones, flavonoids, coumestan, lycopene, ellagic acid, caffeoylquinic acid, hydroxybenzoic acid, hesperetin, flavonols, terpenoids, phthalides, flavonols, allicine quercetin, sulfides, anthocyanins, resveratrol, and antoxanthins selected from one or more of

In one embodiment, the phytonutrient is a colored phytonutrient. In one embodiment, the colored phytonutrients are selected from one or more of anthocyanins, lutein, zeaxanthin, lycopene, carotenoids, and/or antoxanthins.

In one embodiment, the bioactive agent is a fatty acid. As used herein, the term "fatty acid" refers to a carboxylic acid (or organic acid) with a long aliphatic tail, often either saturated or unsaturated. Typically, fatty acids have carbon-carbon bond chains of at least 4 carbon atoms (C4) or at least 8 carbon atoms (C8) in length, more preferably at least 12 carbons in length. Preferred fatty acids of the present invention have 18-22 carbon atoms (C18, C20, C22 fatty acids), more preferably 20-22 carbon atoms (C20, C22), most preferably 22 carbon atoms (C22). of carbon chains. Most naturally occurring fatty acids have an even number of carbon atoms because their biosynthesis involves acetate with two carbon atoms. Fatty acids can be free (non-esterified) or esterified forms such as triglycerides, diacylglycerides, monoacylglycerides, acyl-CoA (thioester) linked forms, or some of the other linked forms. Fatty acids can be esterified as phospholipids, such as in the form of phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, phosphatidylglycerol, phosphatidylinositol, or diphosphatidylglycerol. In one embodiment, fatty acids are esterified to methyl or ethyl groups, such as, for example, methyl or ethyl esters of C20 or C22 polyunsaturated fatty acids. Preferred fatty acids are methyl or ethyl esters of eicosatrienoic acid, docosapentaenoic acid or docosahexaenoic acid, or mixtures of eicosapentaenoic acid and docosahexaenoic acid, or mixtures of eicosapentaenoic acid, docosapentaenoic acid and docosahexaenoic acid, or It is a mixture of eicosapentaenoic acid and docosapentaenoic acid.

In one embodiment the fatty acid is a polyunsaturated fatty acid. As used herein, "polyunsaturated fatty acid" refers to a fatty acid containing more than one double bond in its backbone. In one embodiment, the polyunsaturated fatty acids are selected from one or more of omega-3, omega-6, or omega-9. In one embodiment, omega-3 is hexadecatrienoic acid, alpha-linolenic acid, stearidonic acid, eicosatrienoic acid, eicosatetraenoic acid, eicosapentaenoic acid, heneicosapentaenoic acid, docosapentaenoic acid, docosahexaenoic acid , tetracosapentaenoic acid, and tetracosahexaenoic acid. In one embodiment, the bioactive agent(s) is one or more or all of eicosapentaenoic acid, docosapentaenoic acid, and docosahexaenoic acid. In one embodiment, the omega-6 is linoleic acid, gamma-linolenic acid, eicosadienoic acid, dihomo-gamma-linolenic acid, arachidonic acid, docosadienoic acid, adrenic acid, docosapentaenoic acid, tetracosatetraenoic acid, and tetracosatetraenoic acid. selected from one or more of cosapentaenoic acids. In one embodiment, the omega-9 oil is selected from one or more of oleic acid, eicosenoic acid, mead acid, erucic acid, and nervonic acid. In one embodiment, the bioactive agent is a triglyceride.

In one embodiment the bioactive agent is an oil.

Bioactive Precursors In one embodiment, the biomass and/or further biomass as described herein comprises bioactive precursors. In one embodiment, bioactive precursors are added to oils as described herein.

In one embodiment, the bioactive precursor is a glucosinolate. As used herein, "glucosinolate" refers to secondary metabolites found in at least the Brassicaceae family, which are metabolized via a sulfur atom as described in Halkier and Gershenzon (2006). They share a chemical structure consisting of a β-D-glucopyranose residue linked to a (Z)-N-hydroxyiminosulfate ester and various R groups derived from amino acids. Examples of glucosinolates are provided in Halkier and Gershenzon (2006) and Agerbirk and Olsen (2012). Hydrolysis of glucosinolates can produce isothiocyanates, nitriles, epithionitrile, thiocyanates, and oxazolidine-2-thiones. Many glucosinolates play a role in plant defense mechanisms against pests and diseases.

Glucosinolates are conserved in storage sites in Brassicaceae. As used herein, a "storage site" is a site within Brassicaceae where glucosinolates are present and myrosinase is absent.

As used herein, "myrosinase", also referred to as "thioglucosidases", "siniglases" or "siniglinases", is a family of enzymes involved in plant defense mechanisms that can cleave thio-linked glucose (EC 3. 2.1.147). Myrosinase catalyzes the hydrolysis of glucosinolates, resulting in the production of isothiocyanates. Myrosinase is sometimes stored as myrosin granules in vacuoles of certain atypical cells called myrosinocytes, but has also been reported in protein granules or vacuoles and as a cytosolic enzyme that tends to be membrane-bound. there is

In one embodiment, pretreatment as described herein improves myrosinase access to glucosinolates that produce isothiocyanate bioactives. As used herein, "improve access" or "improved access" refers to increasing the availability of glucosinolates to myrosinase enzymes that allow the production of isothiocyanates. . In one embodiment, access is improved by release of glucosinolates from glucosinolate storage sites. In one embodiment, the glucosinolate storage sites are mechanically disrupted (ie, by maceration). In one embodiment, access is improved by allowing entry of myrosinase into glucosinolate conserved sites. In one embodiment, access is improved by release of myrosinase from myrosin cells. In one embodiment, about 10% to about 90% of the glucosinolates are released from the glucosinolate storage sites. In one embodiment, about 20% to about 80% of the glucosinolates are released from the glucosinolate storage sites. In one embodiment, about 30% to about 70% of the glucosinolates are released from the glucosinolate storage sites. In one embodiment, about 40% to about 60% of the glucosinolates are released from the glucosinolate storage sites. In one embodiment, about 45% to about 55% of the glucosinolates are released from the glucosinolate storage sites. In one embodiment, about 10% of the glucosinolates are released from glucosinolate storage sites. In one embodiment, about 20% of the glucosinolates are released from glucosinolate storage sites. In one embodiment, about 30% of the glucosinolates are released from glucosinolate storage sites. In one embodiment, about 40% of the glucosinolates are released from glucosinolate storage sites. In one embodiment, about 50% of the glucosinolates are released from glucosinolate storage sites. In one embodiment, about 60% of the glucosinolates are released from glucosinolate storage sites. In one embodiment, about 70% of the glucosinolates are released from glucosinolate storage sites. In one embodiment, about 80% of the glucosinolates are released from glucosinolate storage sites. In one embodiment, about 90% of the glucosinolates are released from glucosinolate storage sites.

In one embodiment, the glucosinolate(s) is selected from one or more of aliphatic, indole, or aromatic glucosinolates.

In one embodiment, the aliphatic glucosinolates are glucoraphanin (4-methylsulfinylbutyl or glucoraphanin), sinigrin (2-propenyl), gluconapine (3-butenyl), glucobrasicanapine (4-pentenyl) , progoitrin (2(R)-2-hydroxy-3-butenyl), epiprogoitrin (2(S)-2-hydroxy-3-butenyl), gluconapoleiferin (2-hydroxy-4-pentenyl), glucoibervirine (3-methylthiopropyl), glucoercin (4-methylthiobutyl), dehydroerucine (4-methylthio-3-butenyl), glucoiberin (3-methylsulfinylpropyl), glucoraphenin (4-methylsulfinyl-3- butenyl), glucoalysin (5-methylsulfinylpentenyl), and glucoerytholin (3-methylsulfonylbutyl, 4-mercaptobutyl).

In one embodiment, the indole glucosinolates are glucobrasicine (3-indolylmethyl), 4-hydroxyglucobrasicine (4-hydroxy-3-indolylmethyl), 4-methoxyglucobrasicine (4-methoxy -3-indolylmethyl), and neoglucobrasin (1-methoxy-3-indolylmethyl).

In one embodiment, the indole glucosinolate is selected from one or more of glucotropeolin (benzyl) and gluconasturtiin (2-phenylethyl).

In one embodiment, the glucosinolate is selected from one or more of benzyl glucosinolate, allyl glucosinolate, and 4-methylsulfinylbutyl. In one embodiment, the glucosinolate is glucoraphanin (4-methylsulfinylbutyl). In one embodiment, the glucosinolate is glucobrasin (3-indolylmethyl).

In one embodiment, the glucosinolate is converted to an isothiocyanate before or during steps i), ii), and/or iii) of the method of producing a powder as described herein.

In one embodiment, the bioactive precursor is a component of biomass. In one embodiment, the bioactive precursor is a component of additional biomass. In one embodiment, the bioactive precursor is not present in the biomass or further biomass and is added before, during, or after preparation of the aqueous mixture as described herein. In one embodiment, the bioactive precursor is added before, during or after step ii) of the method as described herein. In one embodiment, the bioactive precursor is added before or during step iii) of the method as described herein. In one embodiment, the bioactive precursor is the oil or component thereof in step ii). In one embodiment, the bioactive precursor is a component added to the oil before it is added to the aqueous mixture in step ii). In one embodiment, the bioactive precursor is a component that is injected into the oil before or during step ii).

Pretreatment In one embodiment, biomass and/or additional biomass as described herein is pretreated. As used herein, "pretreatment" or "pretreating" or "pretreated" decomposes material into smaller components or removes biomass and/or further components of biomass (e.g. removing certain components that are not suitable for ingestion, or extracting certain components, e.g. oils, for different uses) or biomass and/or to modify the composition of the biomass and/or further biomass. Or refers to further processing of biomass. In one embodiment, modifying a component includes, for example, producing a bioactive agent or producing an oligo- or polysaccharide. In one embodiment, the pretreatment does not alter the carbohydrate to protein ratio in the biomass or further biomass.

In one embodiment, the pretreatment comprises: i) heating, ii) maceration, iii) microwave treatment, iv) exposure to low frequency sound waves (ultrasound), v) pulsed electric field treatment, vi) static high pressure. , vii) extrusion, viii) enzymatic treatment, ix) fermentation, x) extraction or separation processes, and xi) drying.

In one embodiment, the biomass and/or further biomass is processed in fuel-based heating systems, electric-based heating systems (i.e., oven or ohmic heating), radio frequency heating, high pressure heat treatment, ultra-high temperature (UHT) processing plants during retorting. , or heated with a steam-based heating system (indirect or direct application of steam). In one embodiment, the biomass and/or additional biomass is heated in an oven, water bath, bioreactor, stove, water blancher, or steam blancher. In one embodiment, the biomass and/or additional biomass is heated via high pressure thermal heating. In one embodiment, the biomass and/or additional biomass is heated via ohmic heating. In one embodiment, the biomass and/or additional biomass is heated via high frequency heating. In one embodiment, the biomass and/or additional biomass is heated via high pressure heat treatment. In one embodiment, the biomass and/or additional biomass is placed in a sealed pack or container for high pressure heat treatment.

In one embodiment, pretreatment includes heating the biomass and/or additional biomass to about 50°C to about 140°C. In one embodiment, heating comprises heating the biomass and/or additional biomass to about 55°C to about 70°C. In one embodiment, heating comprises heating the biomass and/or additional biomass to about 60°C to about 70°C. In one embodiment, heating comprises heating the biomass and/or additional biomass to about 65°C to about 70°C. In one embodiment, heating comprises heating the biomass and/or additional biomass to about 70°C to about 140°C. In one embodiment, heating comprises heating the biomass and/or additional biomass to about 80°C to about 130°C. In one embodiment, heating comprises heating the biomass and/or additional biomass to about 90°C to about 120°C. In one embodiment, heating comprises heating the biomass and/or additional biomass to about 100°C to about 110°C. In one embodiment, heating comprises heating the biomass and/or additional biomass to about 75° C. for about 2 minutes. In one embodiment, heating comprises heating the biomass and/or additional biomass to about 100° C. for about 30 minutes. In one embodiment, the pretreatment comprises heating at the lower end of the above temperature range for an extended period of time or treatment at the upper end of the above temperature range for a short period of time.

In one embodiment, heating includes steaming the biomass and/or additional biomass. In one embodiment, the biomass and/or further biomass is steamed to a temperature of about 100°C. In one embodiment, the biomass and/or additional biomass is steamed for at least about 30 seconds. In one embodiment, the biomass and/or further biomass is steamed for at least 1 minute. In one embodiment, the biomass and/or additional biomass is steamed for at least 2 minutes. In one embodiment, the biomass and/or additional biomass is steamed for at least 3 minutes. In one embodiment, the biomass and/or additional biomass is steamed for at least 4 minutes. In one embodiment, the biomass and/or additional biomass is steamed for at least 5 minutes. In one embodiment, the biomass and/or additional biomass is steamed to a temperature of about 100° C. for 30 minutes.

In one embodiment, heating includes ultra-high temperature (UHT) treatment of the biomass and/or additional biomass. In one embodiment, the biomass and/or further biomass is UHT treated at a temperature of about 140°C.

In one embodiment, heating includes retorting the biomass and/or further biomass. In one embodiment, the biomass and/or additional biomass is retorted at a temperature of about 116°C to about 130°C.

In one embodiment, pretreatment includes maceration of the biomass and/or additional biomass. In one embodiment, the biomass and/or additional biomass is macerated in a shredder, blender, colloid mill, grinder, or pulverizer. In one embodiment, the biomass and/or further biomass is macerated such that at least 80% of the biomass and/or further biomass is 2 mm or less in size. In one embodiment, the biomass and/or further biomass is macerated such that at least 80% of the biomass and/or further biomass is 1 mm or less in size. In one embodiment, the biomass and/or further biomass is macerated such that at least 80% of the biomass and/or further biomass is 0.5 mm or less in size. In one embodiment, the biomass and/or further biomass is macerated such that at least 80% of the biomass and/or further biomass is 0.25 mm or less in size. In one embodiment, the biomass and/or further biomass is macerated such that at least 80% of the biomass and/or further biomass is 0.1 mm or less in size. In one embodiment, the biomass and/or further biomass is macerated such that at least 80% of the biomass and/or further biomass is 0.05 mm or less in size. In one embodiment, the biomass and/or further biomass is macerated such that at least 80% of the biomass and/or further biomass is 0.025 mm or less in size. In one embodiment, the biomass and/or further biomass is heated during maceration. In some embodiments, heating facilitates conversion of bioactive precursors to bioactive substances such as sulforaphane and ajoene. In one embodiment, the biomass and/or additional biomass is heated to a temperature of about 25°C to about 80°C during maceration. In one embodiment, the biomass and/or additional biomass is heated to a temperature of about 40°C to about 70°C during maceration. In one embodiment, the biomass and/or additional biomass is heated to a temperature of about 50°C to about 70°C during maceration. In one embodiment, the biomass and/or additional biomass is heated to a temperature of about 60°C to about 70°C during maceration. In one embodiment, the biomass and/or additional biomass is heated to a temperature of about 70° C. for about 2-5 minutes during maceration. In one embodiment, the biomass and/or additional biomass is heated to a temperature of about 30° C. to about 80° C. for about 1 to about 5 hours during maceration.

In one embodiment, pretreatment includes heating and maceration of the biomass and/or additional biomass.

Those skilled in the art will understand that "microwaves" or "microwave treatment" heats a material, such as biomass and/or additional biomass, by passing microwave radiation through the material. In one embodiment, pretreating comprises microwave treating the biomass and/or additional biomass. In one embodiment, the biomass and/or further biomass is pretreated with domestic or industrial microwaves. In one embodiment, the industrial microwave is a continuous microwave system, such as, but not limited to, MIP 11 Industrial Microwave Continuous Cooking Over (Ferrite Microwave Technologies). In one embodiment, pretreating comprises microwave treating the biomass and/or additional biomass. In one embodiment, the biomass and/or additional biomass is microwaved at about 0.9 to about 2.45 GHz. In one embodiment, the biomass and/or further biomass is microwaved for at least 30 seconds, or at least 1 minute, or at least 2 minutes, or at least 3 minutes. In one embodiment, microwave treatment raises the temperature of the biomass and/or additional biomass to about 70 to about 80°C, preferably about 76°C.

In one embodiment, the pretreatment comprises exposing the biomass and/or additional biomass to low to medium frequency ultrasound. In one embodiment, pretreatment comprises exposing the biomass and/or additional biomass to thermal sonication (low to medium frequency ultrasound with heat from about 50° C. to about 140° C.). In one embodiment, ultrasound is generated using an industrial scale sonicator. In one embodiment, the sonicator is a continuous or batch sonicator. In one embodiment, the sonicator is for example, but not limited to, a UIP500hd or UIP4000 (Hielscher, Ultrasound Technology). In one embodiment, the ultrasound is of a frequency between about 20 kHz and about 600 kHz. In one embodiment, the biomass and/or further biomass is removed for at least 30 seconds, or at least 1 minute, or at least 2 minutes, or at least 3 minutes, or about 5 minutes, or about 6 minutes, or about 7 minutes, or about 7 minutes. .5 minutes, or approximately 8 minutes of exposure to sound waves.

In one embodiment, pretreatment includes exposing the biomass and/or additional biomass to a pulsed electric field treatment. Pulsed electric field treatment is a non-thermal treatment technique that involves the application of short pulses of high voltage. The pulses induce cell electroporation of biomass and/or additional biomass. In one embodiment, the pulsed electric field treatment heats the biomass and/or additional biomass to a temperature of about 50°C to about 140°C. In one embodiment, the pulsed electric field treatment heats the biomass and/or additional biomass to a temperature of about 70°C to about 110°C. In one embodiment, the pulsed electric field treatment heats the biomass and/or additional biomass to a temperature of about 80°C to about 100°C. In one embodiment, pulsed field treatment includes treating the biomass and/or additional biomass with a voltage pulse of about 20 to about 80 kV.

In one embodiment, the pretreatment includes hydrostatic pressure. In one embodiment, hydrostatic pressure includes treating the biomass and/or additional biomass at about 100 to about 600 MPa.

In one embodiment, pretreatment includes extrusion. In one embodiment, extrusion typically involves applying force at elevated temperature and/or pressure to the biomass or product through a barrel prior to discharge of the material through an orifice. In one embodiment, the high temperature, high pressure, and mechanical force applied during extrusion change the functional properties of the material. In one embodiment, the extrusion process comprises a co-rotating twin-screw extruder (MPF 18:25, APV Baker Ltd., Peterborough, UK) or a lab-scale co-rotating, intermeshing twin-screw laboratory extruder ( KDT30-II, Jinan Kredit Machinery Co. Ltd., China). In one embodiment, the extrusion process produces a Maillard reaction product.

In one embodiment, pretreatment includes enzymatic treatment to convert one or more components in the biomass and/or further biomass to new components. For example, enzymes convert monosaccharides into oligosaccharides or polysaccharides. In one embodiment, the enzyme is a glycosyltransferase, ii) a glycosidase, iii) a pectinase, iv) an esterase, v) an oxidoreductase, vi) a protease, vii) a pectinase, viii) a polygalacturonase, ix) an amylase, and x ) pullulanases. In one embodiment, the glycosyltransferases are selected from one or more or all of i) dextrasucrase, ii) alternansucrase, and iii) fructosyltransferase. In one embodiment, the fructosyltransferase is, for example, levansucrase and/or inulosucrase. In one embodiment, the oxidoreductase is mannitol dehydrogenase.

In one embodiment, pretreatment includes fermenting the biomass and/or additional biomass. As used herein, "fermentation" refers to the biochemical degradation of biomass and/or further biomass by bacteria such as, for example, lactic acid and/or acetic acid bacteria. As used herein, a "lactic bacterium" or "lactic acid bacterium" is a bacterium that produces lactic acid as the primary product of carbohydrate fermentation. As used herein, an "acetic bacterium" or "acetic acid bacterium" is a bacterium that produces acetic acid as the end product of carbohydrate fermentation.

In one embodiment, the lactic and/or acetic acid bacteria produce enzymes that catalyze the production of mannitol, oligosaccharides and/or polysaccharides.

In one embodiment, the lactic acid bacterium is Genera Lactobacillus, Leuconostoc, Pediococcus, Lactococcus, Streptococcus, Aerococcus, Carnobacterium, Enterococcus, Oenococcus, Sporolactobacillus, Tetragenoco ccus, Vagococcus, and/or Weissella. In one embodiment, the lactic acid bacteria are selected from one or more of Leuconostoc mesenteroides, Lactobacillus reuteri, and/or Lactobacillus gasseri.

Leuconostoc mesenteroides is a Gram-positive epiphytic bacterium (McCleskey et al., 1947). Leuconostoc mesenteroides also produce an antibacterial protein bacteriocin that is used as a natural preservative in the meat industry. In one embodiment, the lactic acid bacterium is Leuconostoc mesenteroides. In one embodiment, Leuconostoc mesenteroides is as described in Olvera et al. (2007), ATCC 8293 (equivalent to NRRL B-1118) and/or NRRL B-512F.

In one embodiment, the lactic acid bacteria are from Acetobacteraceae. In one embodiment, the Acetobacteraceae is Gluconacetobacter.

In one embodiment, the biomass and/or additional biomass comprises fermentation for about 8 hours to about 30 hours. In one embodiment, fermentation is for at least 8 hours. In one embodiment, fermentation is for at least 10 hours. In one embodiment, the fermentation is for at least 15 hours. In one embodiment, fermentation is for at least 20 hours. In one embodiment, fermentation is for at least 24 hours. In one embodiment, the fermentation is for at least 30 hours. In one embodiment, fermentation is at a pH of about 5 to about 7. In one embodiment, fermentation is at a pH of about 5.3. In one embodiment, the material from step i) has a pH of about 4 at the end of fermentation. In one embodiment, the temperature of fermentation is from about 24°C to about 36°C. In one embodiment, the temperature of fermentation is from about 28°C to about 32°C. In one embodiment, the temperature of fermentation is about 30°C.

In one embodiment, the pretreatment releases or aids in the release of glucosinolates from the glucosinolate storage sites and/or allows myrosinase to enter the glucosinolate storage sites in the biomass and/or further biomass. enable In one embodiment, pretreatment increases the exposure of glucosinolates to myrosinase, allowing myrosinase to convert glucosinolates to isothiocyanates.

In one embodiment, the pretreatment converts about 10% to about 90% of the glucosinolates to isothiocyanates. In one embodiment, the pretreatment converts about 20% to about 80% of the glucosinolates to isothiocyanates. In one embodiment, the pretreatment converts about 30% to about 70% of the glucosinolates to isothiocyanates. In one embodiment, the pretreatment converts about 40% to about 60% of the glucosinolates to isothiocyanates. In one embodiment, the pretreatment converts about 10% of the glucosinolates to isothiocyanates. In one embodiment, the pretreatment converts about 20% of the glucosinolates to isothiocyanates. In one embodiment, the pretreatment converts about 30% of the glucosinolates to isothiocyanates. In one embodiment, the pretreatment converts about 40% of the glucosinolates to isothiocyanates. In one embodiment, the pretreatment converts about 50% of the glucosinolates to isothiocyanates. In one embodiment, the pretreatment converts about 60% of the glucosinolates to isothiocyanates. In one embodiment, the pretreatment converts about 70% of the glucosinolates to isothiocyanates. In one embodiment, the pretreatment converts about 80% of the glucosinolates to isothiocyanates. In one embodiment, the pretreatment converts about 90% of the glucosinolates to isothiocyanates.

In one embodiment, the pretreatment includes treating the biomass and/or further biomass with an extraction or separation process such that the biomass and/or further biomass (e.g., the biomass is depleted or partially depleted of canola oil). reducing the amount of one or more ingredients in the canola meal). In one embodiment, the other component is suitable for producing other products or is a non-edible or palatable component of the biomass and/or further biomass.

In one embodiment, the extraction or separation process is for removing components selected from oils, bioactive substances or bioactive precursors, polyphenols, carotenoids, or juices from the biomass. In one embodiment, the extraction or separation process can be used as biomass in the methods as described herein canola meal, nut meal, soybean meal, coconut meal, palm kernel meal, hemp oil presscake, chia oil. Produce seed cake, or rice bran. In one embodiment, the extraction or separation process produces pomace (eg, olive or apple pomace) that can be used as biomass in methods as described herein. In one embodiment, the extraction or separation process may include removing non-edible components (eg, seeds or stems) from the biomass. In one embodiment, the extraction or separation process may include grinding, cutting, milling, centrifuging, and/or filtering.

As used herein, "reduced" means lower levels of constituents in the biomass or further biomass after treatment by the extraction process than in the biomass or further biomass before treatment by the extraction process. means that

In one embodiment, the level of ingredients is reduced by about 5% to about 90%. In one embodiment, the level of ingredients is reduced by about 5%. In one embodiment, the level of ingredients is reduced by about 10%. In one embodiment, the level of ingredients is reduced by about 15%. In one embodiment, the level of ingredients is reduced by about 20%. In one embodiment, the level of ingredients is reduced by about 30%. In one embodiment, the level of ingredients is reduced by about 40%. In one embodiment, the level of ingredients is reduced by about 50%. In one embodiment, the level of ingredients is reduced by about 60%. In one embodiment, the level of ingredients is reduced by about 70%. In one embodiment, the level of ingredients is reduced by about 80%. In one embodiment, the level of ingredients is reduced by about 90%. In one embodiment, the level of ingredients is reduced by about 100%.

In one embodiment, pretreatment includes drying or partially drying the biomass. In one embodiment, drying includes tray drying, drum drying, roller drying, fluid bed drying, impingement drying, spray drying, freeze drying (lyophilization or low temperature drying), thin film belt dryer, vacuum microwave drying, ultrasonic Assisted drying, extrusion porosification technology, or any other method known to those skilled in the art. In one embodiment, pretreatment comprises freeze-drying the biomass. In one embodiment, pretreatment includes heating the biomass and then freeze-drying. In one embodiment, pretreatment includes drum drying. In one embodiment, pretreatment comprises spray drying.

Those skilled in the art will appreciate that pretreatment does not include separate purification of proteins and purification of carbohydrates from biomass.

In one embodiment, pretreatment does not alter the protein to carbohydrate ratio in the biomass.

Preparation of Aqueous Mixtures As described herein, the methods of the present invention involve obtaining an aqueous mixture from biomass from a first species comprising proteins and carbohydrates. As used herein, "aqueous mixture" refers to a mixture containing biomass containing water. In one embodiment, the aqueous mixture further comprises proteins and carbohydrates from at least one additional biomass (eg, second, third, fourth, fifth, etc.) from the species . In one embodiment, the aqueous mixture is homogeneous. In one embodiment, the aqueous mixture is a suspension.

In one embodiment, the method further comprises forming an aqueous mixture. In one embodiment, the aqueous mixture is formed by combining water at a temperature of about 40° C. to about 100° C. with biomass and optionally further biomass. In one embodiment, the aqueous mixture is formed by combining water at a temperature of about 40°C to about 80°C with biomass and optionally further biomass. In one embodiment, the aqueous mixture is formed by combining water at a temperature of about 45°C to about 70°C with biomass and optionally further biomass. In one embodiment, the aqueous mixture is formed by combining water at a temperature of about 55°C to about 65°C with biomass and optionally further biomass. In one embodiment, the aqueous mixture is formed by combining water at a temperature of about 60° C. with biomass and optionally further biomass.

In one embodiment, the aqueous mixture includes bioactive agents and/or bioactive precursors. In one embodiment, bioactive substances and/or bioactive precursors present in the biomass and/or further biomass are used for injection into the oil in step ii) or iii) of the method as described herein. It is present in the aqueous mixture in a suitable form.

In one embodiment, bioactive substances and/or bioactive precursors suitable for injection into the oil in step ii) or iii) of the method are added to the aqueous mixture.

In one embodiment, minerals are added to the biomass prior to preparation of the aqueous mixture or in step i) or ii) of the method as described herein. In one embodiment, the minerals are selected from one or more of zinc, calcium, magnesium, selenium, and chromium.

Lipids In one embodiment, the methods as described herein further comprise the addition of lipids. As used herein, "lipid" refers to esters of long linear carboxylic acids that are insoluble in water but soluble in organic solvents. In one embodiment, the lipid is saponifiable. In one embodiment the lipid is an oil as described herein. In one embodiment the lipid is a wax as described herein.

Oil In one embodiment, a method as described herein further comprises adding oil to the aqueous mixture. As used herein, "oil" refers to a viscous liquid that is both hydrophobic and lipophilic and immiscible with water. In one embodiment, the oil is susceptible to degradation by one or more of oxidation, temperature, pH, moisture, and light. In one embodiment, the oil is the bioactive substance.

In one embodiment, the oil comprises fatty acids as described herein. In one embodiment, the oil comprises polyunsaturated fatty acids as described herein. In one embodiment, the polyunsaturated fatty acids are selected from one or more of omega-3, omega-6, or omega-9 fatty acids. In one embodiment, omega-3 is hexadecatrienoic acid, alpha-linolenic acid, stearidonic acid, eicosatrienoic acid, eicosatetraenoic acid, eicosapentaenoic acid, heneicosapentaenoic acid, docosapentaenoic acid, docosahexaenoic acid , tetracosapentaenoic acid, and tetracosahexaenoic acid. In one embodiment, the omega-6 is linoleic acid, gamma-linolenic acid, eicosadienoic acid, dihomo-gamma-linolenic acid, arachidonic acid, docosadienoic acid, adrenic acid, docosapentaenoic acid, tetracosatetraenoic acid, and tetracosatetraenoic acid. selected from one or more of cosapentaenoic acids. In one embodiment, the omega-9 is selected from one or more of oleic acid, eicosenoic acid, mead acid, erucic acid, and nervonic acid.

In one embodiment, the oil is Plantae oil. In one embodiment, the oil is vegetable oil. In one embodiment the oil is an animal oil. In one embodiment the animal oil is marine or fish oil.

In one embodiment, the oil is fish oil, krill oil, marine oil, canola oil, sunflower oil, avocado oil, soybean oil, borage oil, evening primrose oil, safflower oil, linseed oil, olive oil, pumpkin seed oil, hemp seed oil, wheat selected from one or more of germ oil, palm oil, palm olein, palm kernel oil, coconut oil, medium chain triglycerides (MCT), and grape seed oil. In one embodiment, canola oil can be obtained from transgenic Brassica that encodes the necessary elongases and desaturases such as eicosapentaenoic acid (EPA), docosapentaenoic acid (DPA), and docosahexaenoic acid (DHA). and long chain polyunsaturated fatty acids (see, for example, WO2015/089587).

In one embodiment, the fish oil is selected from one or more of tuna oil, herring oil, mackerel oil, anchovy oil, sardine oil, cod liver oil, and shark oil.

In one embodiment, the essential oil is one of oregano oil, mint oil, basil oil, rosemary oil, tea tree oil, thyme oil, camphor oil, cardamom oil, citrus oil, clove oil, and/or saffron oil. selected from the above.

In one embodiment, the oil comprises milk fat.

In one embodiment, the oil is olive oil.

In one embodiment, the oil is sunflower oil.

In one embodiment, the oil is canola oil.

In one embodiment, the oil comprises one or more bioactive substance(s) and/or bioactive precursor(s). Thus, in some embodiments, oil functions as a bioactive carrier. In one embodiment, the bioactive agent and/or bioactive precursor are added to the oil before the oil is added to the aqueous mixture. In one embodiment, the bioactive agent and/or bioactive precursor is infused into the oil in step ii) of the method as described herein. In one embodiment, the bioactive agent and/or bioactive precursor is infused into the oil in step iii) of the method as described herein. In one embodiment, the bioactive agent and/or bioactive precursor is from biomass and/or further biomass as described herein. In one embodiment, the bioactive agent and/or bioactive precursor is not from biomass and/or further biomass.

Wax In one embodiment, the method as described herein further comprises the addition of wax. As used herein, "waxes" or "waxes" are esters of long-chain saturated and unsaturated fatty acids and long-chain alcohols. In one embodiment, the long chain saturated fatty acids are C14-C26. In one embodiment, unsaturated fatty acids with long chain alcohols are C16-C30. In one embodiment, the wax is selected from one or more of candelilla wax, carnauba wax, beeswax, rice bran wax, sugarcane wax, and sunflower wax.

Preparation of Emulsions or Suspensions As used herein, an “emulsion” is a dispersion of droplets/particles of one liquid into another liquid that is insoluble or immiscible. Point. In one embodiment, the droplets are oil dispersed in an aqueous mixture. In one embodiment the emulsion is a wet emulsion. In one embodiment, the emulsion is dried into a powder. In one embodiment, the emulsion is extruded. In one embodiment, the emulsion is extruded with a powder matrix.

In one embodiment, the oil droplets produced by the methods described herein are between about 0.2 μm and about 10 μm. In one embodiment, the oil droplets produced by the methods described herein are between about 1 μm and about 10 μm. In one embodiment, the oil droplets produced by the methods described herein are between about 2 μm and about 8 μm. In one embodiment, the oil droplets produced by the methods described herein are between about 2 μm and about 4 μm.

In one embodiment, the average oil droplet size is from about 0.2 μm to about 10 μm. In one embodiment, the average oil droplet size is from about 1 μm to about 10 μm. In one embodiment, the average oil droplet size is from about 2 μm to about 8 μm. In one embodiment, the average oil droplet size is about 2 μm to about 4 μm.

As used herein, "suspension" refers to a dispersion of droplets/particles of one material through a bulk of another material. In one embodiment, the droplets are oil dispersed in an aqueous mixture.

As used herein, the production or formation of an emulsion or suspension refers to entrapment or encapsulation of a substance in an aqueous mixture that reduces the exposure of the substance to degradation. In one embodiment, the substance is oil. In one embodiment, the substance is a bioactive substance. In one embodiment, the bioactive agent is a fatty acid.
In one embodiment, the oil is heated when added to the aqueous mixture in step ii) as described herein. In one embodiment, the oil is heated to about 30°C to about 80°C. In one embodiment, the oil is heated to about 40°C to about 70°C. In one embodiment, the oil is heated to about 45°C to about 65°C. In one embodiment, the oil is heated to about 50°C to about 60°C.

In one embodiment, the bioactive agent and/or bioactive precursor are added to the oil prior to addition to the aqueous mixture. In one embodiment, the bioactive agent and/or bioactive precursor is added to the aqueous mixture before, during, or after the oil is added to the aqueous mixture.

In one embodiment, forming the emulsion or suspension as described in step iii) comprises mixing the oil and the aqueous mixture. In one embodiment, mixing comprises stirring under high shear. In one embodiment, mixing includes homogenization to obtain a small oil droplet size. In one embodiment, the oil droplets produced by homogenization are from about 0.2 μm to about 10 μm in diameter. In one embodiment, the oil droplets produced by homogenization are about 1 μm to about 10 μm in diameter. In one embodiment, the oil droplets produced by homogenization are about 2 μm to about 8 μm in diameter. In one embodiment, the oil droplets produced by homogenization are about 2 μm to about 4 μm in diameter. In one embodiment, homogenization forms a homogeneous emulsion.

In one embodiment, one or more bioactive agent(s) and/or bioactive precursor(s) are added prior to step ii) or step iii) of the method as described herein or It is present in the aqueous solution that is injected into the oil in between.

In one embodiment, bioactive agents and/or bioactive precursors entrapped or encapsulated in emulsions or suspensions by the methods described herein are obtained using the MicroMAX® encapsulation method (WO 01/74175). less susceptible to oxygen degradation than the same bioactive agent and/or bioactive precursor entrapped or encapsulated by .

In one embodiment, the oil content of the emulsion or suspension is from about 1% weight/volume to about 10% weight/volume. In one embodiment, the oil content of the emulsion or suspension is from about 1.2% weight/volume to about 9% weight/volume. In one embodiment, the oil content of the emulsion or suspension is from about 1.3% weight/volume to about 8% weight/volume. In one embodiment, the oil content of the emulsion or suspension is from about 1.4% weight/volume to about 7% weight/volume. In one embodiment, the oil content of the emulsion or suspension is from about 1.5% weight/volume to about 6% weight/volume.

In one embodiment, about 5% to about 50% by weight of the oil is entrapped or encapsulated in the biomass after the emulsion or suspension dries. In one embodiment, about 10% to about 50% by weight of the oil is entrapped or encapsulated in the biomass after the emulsion or suspension dries. In one embodiment, about 20% to about 40% by weight of the oil is entrapped or encapsulated in the biomass after the emulsion or suspension dries. In one embodiment, the emulsion contains dispersed probiotics.

Post-Treatment In one embodiment, a method as described herein comprises post-treating the emulsion or suspension to reduce microbial activity.

As used herein, "post-treatment," "post-treated," or "post-treating" refer to an emulsion or suspension as described herein for reducing microorganisms. Refers to the treatment of turbid liquids.

Those skilled in the art will appreciate that post-processing includes, for example, one or more of heat treatment (including pasteurization), microwave treatment, ultrasonication, UV treatment, high pressure treatment, ultra high temperature treatment (UHT), and retort treatment. It will be understood that any method of inactivating microorganisms or altering product properties (eg, stability as well as physical structure) includes.

In one embodiment, the emulsion or suspension is post-treated with heat treatment. In one embodiment, the emulsion or suspension is post-treated with high pressure treatment. In one embodiment, the emulsion or suspension is in a sealed package during post-processing. In one embodiment, the emulsion or suspension is in a sealed package during high pressure processing. In one embodiment, the emulsion or suspension is in a sealed package during heat treatment. In one embodiment, high pressure treatment comprises treating the emulsion or suspension at a hydrostatic pressure of about 100 to about 600 MPa. In one embodiment, high pressure processing comprises processing the emulsion or suspension at a hydrostatic pressure of about 350 to about 550 MPa. In one embodiment, high pressure treatment comprises treating the emulsion or suspension at a hydrostatic pressure of about 300 to about 400 MPa. In one embodiment, the high pressure process is about 25° C. for about 1 minute, or about 2 minutes, or about 3 minutes, or about 4 minutes. In one embodiment, the heat treatment comprises heating the microparticles to a temperature of about 60°C to about 80°C. In one embodiment, the heat treatment comprises heating the emulsion or suspension to a temperature of about 65°C to about 75°C. In one embodiment, the heat treatment comprises heating the emulsion or suspension under a retort (120°C). In one embodiment, the heat treatment comprises heating the emulsion or suspension under UHT conditions (>120-140°C).

In one embodiment, post-processing includes microwave treatment. In one embodiment, microwave treatment comprises treatment at about 750 W for about 1 minute, or about 2 minutes, or about 2.5 minutes, or about 3 minutes. In one embodiment, microwave treatment raises the temperature of the biomass and/or additional biomass to about 70 to about 80°C, preferably about 76°C.

Powder Preparation In one embodiment, emulsions or suspensions as described herein are partially dried or dried to reduce water content. In one embodiment, the method as described herein comprises drying the emulsion or suspension to reduce the water content to about 1 to about 14%. In one embodiment, the method as described herein comprises drying the emulsion or suspension to reduce the water content to about 1 to about 13%. In one embodiment, the method includes drying the emulsion or suspension to reduce the water content to about 1 to about 12%. In one embodiment, the method includes drying the emulsion or suspension to reduce the water content to about 1 to about 10%. In one embodiment, the method includes drying the emulsion or suspension to reduce the water content to about 2 to about 8%. In one embodiment, the method includes drying the emulsion or suspension to reduce the water content to about 2 to about 6%. In one embodiment, the method includes drying the emulsion or suspension to reduce the water content to about 2 to about 4%. In one embodiment, the method includes drying the emulsion or suspension to reduce the water content to about 2 to about 3%.

In one embodiment, the method as described herein comprises drying the emulsion or suspension to reduce the water activity to a low water activity of about 0.1 to about 0.7. include. In one embodiment, the method includes drying the emulsion or suspension to reduce the water activity to a low water activity of about 0.2 to about 0.6. In one embodiment, the method includes drying the emulsion or suspension to reduce the water activity to a low water activity of about 0.2 to about 0.5. In one embodiment, the method includes drying the emulsion or suspension to reduce the water activity to a low water activity of about 0.3 to about 0.4. In one embodiment, the method includes drying the emulsion or suspension to reduce the water activity to a low water activity of about 0.4.

In one embodiment, the method as described herein comprises drying the emulsion or suspension to form a powder. Drying includes, for example, spray drying, freeze drying (ryophilization or low temperature drying), tray drying, drum drying, roller drying, fluidized bed drying, impingement drying, Refractance Window drying, thin film belt drying, vacuum Microwave drying, ultrasonically assisted drying, extrusion porosification techniques, or any other method known to those skilled in the art may be included.

In one embodiment, the emulsion or suspension is dried to produce an average dry particle size of about 10 μm to about 4000 μm. In one embodiment, the emulsion or suspension is dried to produce an average dry particle size of about 10 μm to about 3000 μm. In one embodiment, the emulsion or suspension is dried to produce an average dry particle size of about 20 μm to about 2000 μm. In one embodiment, the emulsion or suspension is dried to produce an average dry particle size of about 10 μm to about 1000 μm. In one embodiment, the emulsion or suspension is dried to produce an average dry particle size of about 10 μm to about 500 μm.

In one embodiment, the emulsion or suspension is dried by spray drying (eg, a Drytec laboratory spray dryer) to form a powder. For example, emulsions or suspensions can be prepared by heating the feed to 60° C. prior to atomization using Drytec laboratory spray dryers with rotary atomizers, ultrasonic nozzles, or twin-fluid nozzles. at a spray pressure of 2.0-4.0 bar, the inlet and outlet air was 180° C. and 80° C., respectively. In one embodiment, the spray dryer has a granulation function. In one embodiment, the spray dryer is fitted with a granulation dryer.

In one embodiment, spray drying produces individual particles or aggregates of particles.

In one embodiment, spray drying produces an average dry particle size of about 10 μm to about 3000 μm. In one embodiment, spray drying produces an average dry particle size of about 20 μm to about 2000 μm. In one embodiment, spray drying produces an average dry particle size of about 10 μm to about 1000 μm. In one embodiment, spray drying produces an average dry particle size of about 10 μm to about 500 μm.

In one embodiment, the emulsion or suspension is dried by lyophilization to form a powder. In one embodiment, a cryoprotectant is added to the emulsion or suspension prior to lyophilization. In one embodiment, the cryoprotectant is a mono-, di- or polysaccharide, polyalcohol, or derivative thereof. In one embodiment, the cryoprotectant is selected from one or more of trehalose, sucrose, and mannitol.

In one embodiment, the emulsion or suspension is dried by drum drying to form a powder.

In one embodiment, the powder comprises from about 5% to about 50% oil by weight. In one embodiment, the powder comprises from about 10% to about 50% oil by weight. In one embodiment, the powder comprises from about 20% to about 50% oil by weight. In one embodiment, the powder comprises from about 20% to about 50% oil by weight. In one embodiment, the powder comprises from about 20% weight/volume to about 40% weight/volume oil. In one embodiment, the powder comprises from about 20% to about 30% oil by weight.

In one embodiment, the powder comprises particles from about 20 μm to about 1200 μm. In one embodiment, the powder comprises particles from about 100 μm to about 900 μm. In one embodiment, the powder comprises particles from about 400 μm to about 700 μm. In one embodiment, the powder comprises particles between about 500 μm and about 600 μm. In one embodiment, the powder comprises particles of approximately 1000 μm. In one embodiment, the powder is milled to further reduce particle size. In one embodiment, milling can reduce particle size to less than about 10 μm, or less than about 8 μm, or less than about 6 μm, or less than about 4 μm, or less than about 2 μm.

In one embodiment, the bioactive agents and/or bioactive precursors captured or encapsulated in the powder by the methods described herein are captured or encapsulated by the MicroMAX® encapsulation method (WO 01/74175). Less susceptible to oxygen degradation than the same bioactive agent and/or bioactive precursor (eg, oil).

In one embodiment, the bioactive agents and/or bioactive precursors entrapped or encapsulated in the powder by the methods as described herein were compared in time to IP, About 500% to about 4000% more resistant to oxygen degradation than bioactive agents and/or bioactive precursors (see Table 7). In one embodiment, the bioactive agents and/or bioactive precursors entrapped or encapsulated in the powder are higher than the bioactive agents and/or bioactive precursors that are not entrapped or encapsulated in the compared time to IP. are also about 500% to about 3000% more resistant to oxygen decomposition. In one embodiment, the bioactive agents and/or bioactive precursors entrapped or encapsulated in the powder are higher than the bioactive agents and/or bioactive precursors that are not entrapped or encapsulated in the compared time to IP. are also about 500% to about 2000% more resistant to oxygen decomposition. In one embodiment, the bioactive agents and/or bioactive precursors entrapped or encapsulated in the powder are higher than the bioactive agents and/or bioactive precursors that are not entrapped or encapsulated in the compared time to IP. are also about 800% to about 2000% more resistant to oxygen decomposition. In one embodiment, the bioactive agents and/or bioactive precursors entrapped or encapsulated in the powder are higher than the bioactive agents and/or bioactive precursors that are not entrapped or encapsulated in the compared time to IP. are also about 800% to about 1500% more resistant to oxygen decomposition. In one embodiment, the bioactive agents and/or bioactive precursors entrapped or encapsulated in the powder are higher than the bioactive agents and/or bioactive precursors that are not entrapped or encapsulated in the compared time to IP. are also about 900% to about 1300% more resistant to oxygen decomposition.

In one embodiment, the bioactive agent and/or bioactive precursor entrapped or encapsulated in the powder lasts at least 3 months compared to the bioactive agent and/or bioactive precursor that is not entrapped or encapsulated. Resistant to oxygen decomposition. In one embodiment, the bioactive agent and/or bioactive precursor entrapped or encapsulated in the powder lasts at least 6 months compared to the bioactive agent and/or bioactive precursor that is not entrapped or encapsulated. Resistant to oxygen decomposition. In one embodiment, the bioactive agent and/or bioactive precursor entrapped or encapsulated in the powder lasts at least 12 months compared to the bioactive agent and/or bioactive precursor that is not entrapped or encapsulated. Resistant to oxygen decomposition. In one embodiment, the bioactive agent and/or bioactive precursor entrapped or encapsulated in the powder lasts at least 18 months compared to the bioactive agent and/or bioactive precursor that is not entrapped or encapsulated. Resistant to oxygen decomposition. In one embodiment, the bioactive agent and/or bioactive precursor entrapped or encapsulated in the powder lasts at least 24 months compared to the bioactive agent and/or bioactive precursor that is not entrapped or encapsulated. Resistant to oxygen decomposition.

In one embodiment, the bioactive agent and/or bioactive precursor entrapped or encapsulated in the powder lasts at least 3 months compared to the bioactive agent and/or bioactive precursor that is not entrapped or encapsulated. Resistant to moisture decomposition. In one embodiment, the bioactive agent and/or bioactive precursor entrapped or encapsulated in the powder lasts at least 6 months compared to the bioactive agent and/or bioactive precursor that is not entrapped or encapsulated. Resistant to moisture decomposition. In one embodiment, the bioactive agent and/or bioactive precursor entrapped or encapsulated in the powder lasts at least 12 months compared to the bioactive agent and/or bioactive precursor that is not entrapped or encapsulated. Resistant to moisture decomposition. In one embodiment, the bioactive agent and/or bioactive precursor entrapped or encapsulated in the powder lasts at least 18 months compared to the bioactive agent and/or bioactive precursor that is not entrapped or encapsulated. Resistant to moisture decomposition. In one embodiment, the bioactive agent and/or bioactive precursor entrapped or encapsulated in the powder lasts at least 24 months compared to the bioactive agent and/or bioactive precursor that is not entrapped or encapsulated. Resistant to moisture decomposition.

In one embodiment, bioactive agents and/or bioactive precursors that are entrapped or encapsulated in the powder are pH degraded during the process compared to bioactive agents and/or bioactive precursors that are not entrapped or encapsulated. more tolerant. In one embodiment, the bioactive agent and/or bioactive precursor entrapped or encapsulated in the powder has a higher pH during gastrointestinal transit compared to a bioactive agent and/or bioactive precursor that is not entrapped or encapsulated. Resistant to decomposition.

In one embodiment, the powder-encapsulated oil is more resistant to oxygen degradation for at least three months compared to non-encapsulated oil. In one embodiment, the powder-encapsulated oil is more resistant to oxygen degradation for at least 6 months compared to non-encapsulated oil. In one embodiment, the powder-encapsulated oil is more resistant to oxygen degradation for at least 12 months compared to non-encapsulated oil. In one embodiment, the powder-encapsulated oil is more resistant to oxygen degradation for at least 18 months compared to non-encapsulated oil. In one embodiment, the powder-encapsulated oil is more resistant to oxygen degradation for at least 24 months compared to non-encapsulated oil.

In one embodiment, the powder-encapsulated oil is more resistant to thermal degradation for at least three months compared to non-encapsulated oil. In one embodiment, the powder-encapsulated oil is more resistant to thermal decomposition for at least six months compared to non-encapsulated oil. In one embodiment, the powder-encapsulated oil is more resistant to thermal decomposition for at least 12 months compared to non-encapsulated oil. In one embodiment, the powder-encapsulated oil is more resistant to thermal decomposition for at least 18 months compared to unencapsulated oil. In one embodiment, the powder-encapsulated oil is more resistant to thermal degradation for at least 24 months compared to non-encapsulated oil.

In one embodiment, the powder-encapsulated oil is more resistant to moisture degradation for at least 3 months compared to non-encapsulated oil. In one embodiment, the powder-encapsulated oil is more resistant to moisture degradation for at least 6 months compared to non-encapsulated oil. In one embodiment, the powder-encapsulated oil is more resistant to moisture degradation for at least 12 months compared to non-encapsulated oil. In one embodiment, the powder-encapsulated oil is more resistant to moisture degradation for at least 18 months compared to non-encapsulated oil. In one embodiment, the powder-encapsulated oil is more resistant to moisture degradation for at least 24 months compared to non-encapsulated oil.

In one embodiment, the powder-encapsulated oil is more resistant to pH degradation for at least 3 months compared to non-encapsulated oil. In one embodiment, the powder-encapsulated oil is more resistant to pH degradation for at least 6 months compared to non-encapsulated oil. In one embodiment, powder-encapsulated oil is more resistant to pH degradation during gastrointestinal transit than non-encapsulated oil.

Products In one aspect, the invention provides a matrix comprising proteins and carbohydrates from biomass from a first species. In one embodiment, the matrix comprises one or more bioactive agent(s) or bioactive precursor(s) as described herein. In one embodiment, the matrix comprises sulforaphane. In one embodiment, the matrix comprises glucosinolates. In one embodiment, the matrix comprises glucoraphanin.

In one aspect, the present invention provides a matrix comprising oil droplets or bioactive agents and/or bioactive precursors, wherein proteins and carbohydrates are present in additional species of organisms (e.g., second, third, fourth organisms ). A matrix is provided that is from one or more additional biomass from (species such as , 5th, etc.).

In one aspect, the invention provides a bioactive agent and/or bioactive precursor entrapped or encapsulated in a matrix, comprising proteins and carbohydrates from biomass from a first species, wherein the entrapped or encapsulated The bioactive agent and/or bioactive precursor is more resistant to oxygen degradation when compared to the bioactive agent and/or bioactive precursor prior to entrapment or encapsulation. Provide an active precursor.

In one embodiment, the bioactive agent and/or bioactive precursor is not from the first species .

In one aspect, the present invention provides a bioactive agent and/or bioactive precursor entrapped or encapsulated in a matrix, comprising proteins and carbohydrates from broccoli, wherein the entrapped or encapsulated bioactive agent and/or The bioactive agent and/or bioactive precursor provides a bioactive agent and/or bioactive precursor that is more resistant to oxygen degradation when compared to the bioactive agent and/or bioactive precursor prior to entrapment or encapsulation.

In one embodiment, the entrapped or encapsulated bioactive agents and/or bioactive precursors are fatty acids. In one embodiment the bioactive agent is an oil.

In one embodiment, the matrix comprises proteins and carbohydrates from at least one additional biomass from the first species .

In one embodiment, the biomass and/or further biomass comprises i) a protein to carbohydrate ratio of about 1:1 to about 1:10.5, ii) a protein to carbohydrate ratio of about 1:4.5 to about 4:1. and ii) a protein to carbohydrate ratio of from about 1:2.5 to about 2:1.

In one embodiment, the biomass and/or further biomass comprises bioactive substances and/or bioactive precursors.

In one embodiment, the biomass is broccoli.

In one aspect, the invention provides an emulsion or suspension produced by a method as described herein. In one embodiment, the emulsion or suspension has an induction period of from about 10 hours to about 300 hours at 80°C, measured using Oxipres at 80°C and an initial 5 bar oxygen pressure. In one embodiment, the emulsion or suspension has an induction period of about 100 hours to about 300 hours at 80°C, measured using Oxipres at 80°C and an initial 5 bar oxygen pressure.

In one aspect, the invention provides a powder comprising entrapped or encapsulated bioactive agents and/or bioactive precursors produced by a method as described herein. In one embodiment, the powder has an induction period of about 10 hours to about 300 hours at 80° C. measured using Oxipres at 80° C. and an initial 5 bar oxygen pressure. In one embodiment, the powder has an induction period of about 50 hours to about 300 hours at 80° C. measured using Oxipres at 80° C. and an initial 5 bar oxygen pressure. In one embodiment, the powder has an induction period of about 80 hours to about 300 hours at 80° C. measured using Oxipres at 80° C. and an initial 5 bar oxygen pressure. In one embodiment, the powder has an induction period of about 100 hours to about 300 hours at 80° C. measured using Oxipres at 80° C. and an initial 5 bar oxygen pressure. In one embodiment, the powder has an induction period of at least 10 hours at 80°C, measured using Oxipres at 80°C and an initial oxygen pressure of 5 bar. In one embodiment, the powder has an induction period of at least 50 hours at 80°C, measured using Oxipres at 80°C and an initial oxygen pressure of 5 bar. In one embodiment, the powder has an induction period of at least 100 hours at 80°C, measured using Oxipres at 80°C and an initial oxygen pressure of 5 bar.

In one embodiment, the moisture content of the powder is from about 1 to about 14%. In one embodiment, the moisture content of the powder is from about 1 to about 10%. In one embodiment, the moisture content of the powder is about 10% or less. In one embodiment, the moisture content of the powder is about 8% or less. In one embodiment, the moisture content of the powder is about 7% or less. In one embodiment, the moisture content of the powder is about 6% or less. In one embodiment, the moisture content of the powder is about 5% or less. In one embodiment, the moisture content of the powder is about 4% or less. In one embodiment, the moisture content of the powder is about 3% or less.

In one embodiment, the powder includes oil. In one embodiment, the powder comprises omega-3 polyunsaturated fatty acids. In one embodiment, the powder comprises an isothiocyanate bioactive.

In one embodiment, a powder is a material that can be used as is or added to or combined with other materials to form a product (eg, food or cosmetic).

In one embodiment, powders can be used to form powders (eg, combined with one or more other powdered ingredients), tablets, liquids, pills, capsules, or extruded products. In one embodiment, the powder is extruded. In one embodiment, the powder is compressed to form, for example, tablets.

In one embodiment, the powder is a food, food ingredient, beverage ingredient, or cosmetic ingredient.

In one embodiment, an emulsion, suspension, or powder can be combined with one or more other ingredients to form a product.

In one embodiment, the product is a cream, gel, tablet, liquid, pill, capsule, or extruded product.

In one embodiment, the product is a food product, food ingredient, beverage ingredient supplement, cosmetic or cosmetic ingredient. In one embodiment, the cosmetic product is a skin moisturizing product such as a moisturizer or face mask.

In one embodiment, the product comprises omega-3 polyunsaturated fatty acids.

In one embodiment, the food product is animal feed. In one embodiment, the animal feed comprises omega-3 polyunsaturated fatty acids. In one embodiment, the omega-3 polyunsaturated fatty acid is one of alpha-linolenic acid (ALA), eicosapentaenoic acid (EPA), and docosahexaenoic acid (DHA), and docosapentaenoic acid (DPA) selected from the above. In one embodiment, the animal feed comprises astaxanthin and/or alpha-lipoic acid.

In one embodiment the animal feed is an aquaculture feed.

In one embodiment, the product is, for example, infant formula, children's formula, adult formula, yoghurt, beverages, geriatric supplements, ultra high temperature processed (UHT) beverages (e.g. milk), soups, dips , pasta products, breads, snacks, other bakery products; processed cheese; and/or food ingredients for animal feed (including aquaculture feed). In one embodiment, the entrapped or entrapped bioactive agent and/or bioactive precursor in the food ingredient is less than the unencapsulated or unencapsulated bioactive agent and/or bioactive precursor when added to the product. is also stable.

In one embodiment the product is suitable for use as a cosmetic or cosmetic ingredient, for example as a lipstick, cream, lotion or ointment.

In one embodiment, the product is a powdered supplement. In one embodiment, the powdered supplement is dissolved in water or added to food, beverages, or meals.

In one embodiment the product is an emulsion, suspension or powder.

Preparation of Pharmaceutical Compositions In one aspect, the present invention provides a method of producing an emulsion comprising isothiocyanates and/or isothiocyanate precursors, comprising: providing a mixture, thereby forming an emulsion. Such emulsions are suitable for use in pharmaceutical compositions.

In one embodiment, a mixture comprising water and isothiocyanates and/or isothiocyanate precursors is mixed with lipids. In one embodiment, an aqueous suspension comprising isothiocyanates and/or isothiocyanate precursors and comprising proteins and/or carbohydrates is mixed with lipids. In one embodiment, isothiocyanates and/or isothiocyanate precursors are mixed with lipids and the resulting composition is mixed with an aqueous medium. In one embodiment, the aqueous medium comprises proteins and/or carbohydrates. In one embodiment, isothiocyanates and/or isothiocyanate precursors are mixed with a mixture comprising water and lipids. In one embodiment, isothiocyanates and/or isothiocyanate precursors are mixed with a mixture comprising water and lipids. In one embodiment, the proteins or carbohydrates are from the same single species organism . In one embodiment, the proteins, carbohydrates, or isothiocyanates and/or isothiocyanate precursors are from the same single species of microorganism.

In one embodiment, the concentration of isothiocyanate and/or isothiocyanate precursors in the emulsion is reduced after about 1 month of storage at about 4 to about 10°C or about -18°C to a corresponding composition devoid of lipids. at least twice the concentration of isothiocyanate and/or isothiocyanate precursor in the product. In one embodiment, the concentration of isothiocyanate and/or isothiocyanate precursors in the emulsion is reduced after about 1 month of storage at about 4 to about 10°C or about -18°C to a corresponding composition devoid of lipids. at least 3 times the concentration of isothiocyanate and/or isothiocyanate precursor in the product. In one embodiment, the concentration of isothiocyanate and/or isothiocyanate precursors in the emulsion is reduced after about 2 months of storage at about 4 to about 10°C or about -18°C to a corresponding composition devoid of lipids. at least twice the concentration of isothiocyanate and/or isothiocyanate precursor in the product.

In one aspect, the invention provides a method of preparing a powder comprising isothiocyanates and/or isothiocyanate precursors, comprising preparing an emulsion as described herein; and subjecting to drying conditions thereby removing water. In one embodiment, the emulsion is subjected to freeze-drying or spray-drying conditions to form a powder.

In one embodiment, the concentration of isothiocyanates and/or isothiocyanate precursors in the powder is reduced to that of the corresponding powder lacking lipids after about 1 month of storage at -18°C. at least one-fold the body concentration.

In one embodiment, the concentration of isothiocyanates and/or isothiocyanate precursors in the powder is reduced to that of the corresponding powder lacking lipids after about 1 month of storage at -18°C. at least twice the body concentration.

In one embodiment, the concentration of isothiocyanates and/or isothiocyanate precursors in the powder is reduced to that of the corresponding powder lacking lipids after about 2 months of storage at -18°C. at least twice the body concentration.

In one aspect, the invention provides a method of preparing a pharmaceutical or cosmetic composition, comprising preparing an emulsion as described herein or A method is provided comprising preparing a powder and converting the emulsion or dry composition into a pharmaceutical or cosmetic composition.

Pharmaceutical Compositions In one embodiment, the present invention is a pharmaceutical or cosmetic composition produced by the method or from the emulsions or powders described herein, comprising an isothiocyanate and/or an isothiocyanate A pharmaceutical or cosmetic composition is provided comprising a precursor, a lipid, and a pharmaceutical and/or cosmetic excipient.

In one aspect, the present invention provides pharmaceutical or cosmetic compositions comprising isothiocyanates and/or isothiocyanate precursors, lipids, and pharmaceutical and/or cosmetic excipients. In one embodiment, the pharmaceutical or cosmetic composition further comprises protein and/or carbohydrate.

In one aspect, the invention provides an emulsion comprising water, lipids, and isothiocyanates and/or isothiocyanate precursors. Such emulsions are suitable for use in pharmaceutical and cosmetic compositions.

In one embodiment, the composition is for topical, enteral/gastrointestinal, or parenteral administration. In one embodiment, application to topical areas of the skin is included, including transdermal administration (administration via absorption through the skin). In one embodiment, includes enteral/gastrointestinal, eg, oral, rectal, gastric, gastrointestinal, sublabial, buccal, sublingual. In one embodiment, parenteral, eg, transdermal, intramuscular, and intravenous are included. In one embodiment, the composition is in the form of a cream, ointment, gel, tablet, liquid, pill, capsule, powder, or extruded product.

In one embodiment, about 10% to about 90% of the isothiocyanate and/or isothiocyanate precursor remains in the composition after storage for a period of about one month. In one embodiment, at least 10% of the isothiocyanate and/or isothiocyanate precursor remains in the composition after storage for a period of about one month. In one embodiment, at least 20% of the isothiocyanate and/or isothiocyanate precursor remains in the composition after storage for a period of about one month. In one embodiment, at least 30% of the isothiocyanate and/or isothiocyanate precursor remains in the composition after storage for a period of about one month. In one embodiment, at least 40% of the isothiocyanate and/or isothiocyanate precursor remains in the composition after storage for a period of about one month. In one embodiment, at least 50% of the isothiocyanate and/or isothiocyanate precursor remains in the composition after storage for a period of about one month.

In one embodiment, the isothiocyanate is selected from one or more of sulforaphane, allyl isothiocyanate, benzyl isothiocyanate, and phenethyl isothiocyanate.

In one embodiment, the isothiocyanate precursor is selected from one or more of glucosinolates, glucoraphanin, sinigrin, glucotropeolin, and gluconasturtiin.

In one embodiment the lipid is an oil as described herein. In preferred embodiments, the oil is selected from canola oil, olive oil, sunflower oil, fish oil, or algae oil. In one embodiment, the composition comprises from about 10% to about 90% oil. In one embodiment, the composition comprises from about 20% to about 80% oil. In one embodiment, the composition comprises about 30% to about 70% oil. In one embodiment the lipid is a wax as described herein.

In one embodiment, the emulsion or powder contains one or more excipients that should be pharmaceutically or cosmetically acceptable in the sense of being compatible with the other ingredients of the formulation and not being unduly harmful to the recipient thereof. combined with excipients, carriers, or excipients, including, for example, polyvinylpyrrolidone, derivatized celluloses such as hydroxymethylcellulose, hydroxyethylcellulose, and hydroxypropylmethylcellulose; ficoll (a polymeric sugar); hydroxyethyl starch (HES); Dextrates (eg, cyclodextrins such as 2-hydroxypropyl-β-cyclodextrin and sulfobutylether-β-cyclodextrin), polyethylene glycols, and pectin may be included. The composition may contain diluents, buffers, binders, disintegrants, thickeners, lubricants, preservatives (including antioxidants), flavoring agents, taste-masking agents, inorganic salts (e.g. sodium chloride), antimicrobial agents. (e.g. benzalkonium chloride), sweeteners, antistatic agents, sorbitan esters, lipids (e.g. phospholipids such as lecithin and other phosphatidylcholines, phosphatidylethanolamines, fatty acids and fatty acid esters, steroids (e.g. cholesterol)), and chelating agents such as EDTA, zinc, and other suitable cations. Other pharmaceutical excipients, carriers, and/or additives suitable for use in the compositions are described in "Remington: The Science & Practice of Pharmacy", 19. sup. th ed. , Williams & Williams, (1995), and "Physician's Desk Reference", 52. sup. nd ed. , Medical Economics, Montvale, N.J. J. (1998), and "Handbook of Pharmaceutical Excipients", Third Ed. , Ed. A. H. Kibbe, Pharmaceutical Press, 2000.

In one aspect, the invention is a method of treating or preventing a condition, which requires administration of an effective amount of a pharmaceutical composition, emulsion, or powder as described herein. A method is provided that includes performing a subject. In one aspect, the invention provides a pharmaceutical composition, emulsion or powder as described herein for use in treating or preventing a condition. In one aspect, the invention provides a method of treating or preventing a condition in a subject, comprising administering to the subject an effective amount of a pharmaceutical composition, emulsion, or powder as described herein. A method is provided, comprising: Use of a pharmaceutical composition as described herein in the manufacture of a medicament for treating or preventing a condition. Use of an emulsion as described herein or a powder as described herein for the manufacture of a medicament for the treatment or prevention of a condition.

In one aspect, the invention provides a method of treatment or prevention, use, or method of treatment or prevention as described herein, wherein the condition is cancer, diabetes, cardiovascular disease, autism. , osteoporosis, neuroprotective diseases, metabolic syndrome, inflammation, oxidative stress, and gut health. In one embodiment, the intestinal health condition is selected from ulcerative colitis, irritable bowel syndrome, Crohn's disease, small bowel overgrowth, leaky gut, and lactose intolerance. In one embodiment, the condition is cancer.

Example 1 - Demonstration of Emulsification and Physical Function of Broccoli as an Encapsulating Agent to Prepare a Physically Stable Oil-in-Water Emulsion Fresh broccoli was mixed with additional water (1:1 ratio). To this was added oil such that the broccoli:water:oil ratio was 1:1:1 and the entire mixture was blended using a benchtop blender. The oil-in-water emulsion was physically stable for more than 2 hours. When the oil-water mixture without broccoli was mixed, it immediately separated into two phases, as expected (FIGS. 1A-B).

Example 2 - Preparation of an Aqueous Phase Suspension Using Lyophilized Broccoli Powder as Encapsulating Agent Lyophilized broccoli powder is placed in a beaker and mixed using an overhead mixer until a fluid mixture is obtained. , water (60° C.) was added (7.46% total solids (TS)) (FIGS. 1C-F). The pH of the mixture was adjusted to 6.01-7.50 using 2N NaOH. The mixture was then heat treated at 75°C for 2 minutes or 100°C for 30 minutes and then cooled to 60°C.

Example 3 - Preparation of aqueous phase suspension using fresh broccoli as encapsulant Fresh broccoli (10% TS) was cut into small pieces, boiling water was added first and the mixture was blended to give different total solids content. Broccoli suspensions were obtained (Figure 2A: 7.66% (TS), (Figure 2B: 6.87% TS), (Figure 2C: 6.23% TS), (Figure 2D: 4.99% TS ) From this initial experiment, a 4.99% TS mixture was selected for the preparation of the encapsulant The pH of this mixture was adjusted to 6.23-7.50 using 2N NaOH. After that, the mixture was heat treated at 75°C for 2 minutes or 100°C for 30 minutes and then cooled to 60°C.

Example 4 Demonstration of Freeze-Dried Broccoli Powder for Encapsulating Omega-3 Oil in Emulsion and Powder Formats An aqueous phase suspension (5% TS) was used as the mounting medium. Tuna oil (1:1 broccoli solids:oil ratio) was added to the aqueous phase suspension (60° C.) described in Example 2 and filtered using an Ultra Turrax at 15,000 rpm for 3 minutes. F1 emulsions were homogenized and heated at 75° C. for 2 min, and F2 was prepared at 100° C. for 30 min (FIG. 3A).

An aqueous phase formulation using 1 sodium caseinate (NaCas)-1 glucose (Glu)-1 dry glucose syrup (DGS) solution was prepared in water at 60° C. for 40 minutes (25% TS) to pH 7.50. heat treated at 100° C. for 30 minutes, cooled to 60° C. (C1 as control), Tween 80 (0.5 g) was dissolved in 87 g of water at 60° C. as another control using a low molecular weight emulsifier. for 5 minutes (C2).

Omega-3 oil-in-water emulsions were made by combining oil with protein (sodium caseinate) and carbohydrate (glucose and dried glucose syrup) C1, or low molecular weight emulsifier C2 (FIG. 3A).

All F1, F2, C1, C2 emulsions were physically stable after overnight storage at 20° C. (FIG. 3B). F1, F2, C1 were also lyophilized to obtain omega-3 oil powder containing 50% omega-3 oil (Fig. 3C).

Example 5 - Oxidative Stability of Omega-3 Oil Emulsions and Powders Tested Under Accelerated Oxidation , F2) were tested under accelerated oxidation at 80° C. under an initial oxygen pressure of 5 bar using an Oxipres apparatus (Mikrolab Aarhus A/S, Hojbjerg, Denmark). Fish oil emulsions and powders made with broccoli matrices (F1 and F2) were encapsulated using CSIRO's MicroMAX® technology and the corresponding fish oil emulsion made with Tween. (F1 and F2) and the emulsion (C1) absorbed oxygen more slowly (Figures 4 and 5 and Table 2).

Example 6 - Demonstration of Use of Broccoli Powder to Stabilize Omega-3 Oil in Powder Oxipres data for unrefined oil is shown in FIG. The effect of the amount of encapsulant matrix on oxygen uptake was evaluated with the Oxipres test against broccoli matrix (no oil). The results are shown in FIG.

Broccoli heads were cut into quarters, macerated by adding water (5% TS), heated to 79°C for 4 minutes and freeze dried. Lyophilized broccoli powder was reconstituted with water (5% TS) and used for encapsulation. Oil (tuna oil, DHA canola oil or canola oil) was added to the suspension to obtain 12.5%, 25% and 50% oil powder and homogenized using an Ultraturrax at 15,000 rpm for 3 minutes. to prepare an emulsion. The emulsions (5.7%, 6.6%, and 9.5% total solids, respectively) were lyophilized and subjected to accelerated oxidation conditions using an Oxipres unit at room temperature at 80° C. with an initial oxygen pressure of 5 bar. tested below.

The results are shown in Figures 8-10. The slow oxygen uptake in these samples is due in part to oxygen uptake by the broccoli matrix. No clear IP was observed for 12.5% and 25% oil powder up to 300 hours.

Example 7 - Demonstration of Using Fresh Broccoli for the Preparation and Stabilization of Omega-3 Oil-in-Water Emulsions % TS) was used as the mounting medium. Omega-3 oil (1:1 broccoli solids:oil ratio) was added to the aqueous phase suspension (60° C.) described in Example 3 and spun at 15,000 rpm for 3 minutes using an Ultraturrax. E1 and E2 emulsions were prepared by homogenizing at 75° C. for 2 minutes and E3 and E4 by heating at 100° C. for 30 minutes. The final emulsions on total solids were 7.7% and 11.3%, respectively.

The emulsion was tested under accelerated oxidation conditions using an Oxipres unit at 80° C. and room temperature with an initial oxygen pressure of 5 bar. The results are shown in FIG. 11 and Table 3. Both cooked and uncooked broccoli performed the same way with respect to oxygen uptake.

Example 8 - Oxipres results for fresh broccoli tuna oil lyophilized powder Fresh broccoli (5% and 6% TS) was mixed with boiling water at high speed for 3 minutes (temperature measured 68.3°C). The pH was adjusted to 7.5. One half was pasteurized at 75°C for 2 minutes and the other half was heat treated at 100°C for 30 minutes. Both of them were cooled to 60° C., tuna oil was added and ultra-turraxed at 15,000 rpm for 2 minutes. The resulting emulsions containing 9.5% and 11.3% TS respectively were then lyophilized. Using lyophilized broccoli, an emulsion formulation (9.5% total solids) using sodium caseinate and carbohydrates as encapsulating agents was also prepared, lyophilized and tested for comparison.

The freeze-dried emulsions were tested at room temperature at 80° C. and an initial oxygen pressure of 5 bar using an Oxipres unit. The results are shown in Figures 12, 13 and Table 4.

Example 9 - Oxygen uptake of omega-3 broccoli emulsion samples made using pretreated broccoli as mounting medium The oxygen uptake of broccoli emulsions using steamed shredded broccoli, steamed pureed drum-dried broccoli) followed by drum drying was evaluated by the Oxipres test at 80°C and an initial oxygen pressure of 5 bar. .

The results are shown in FIG. Broccoli encapsulant was used at various processing stages to produce up to 5% aqueous solids. An emulsion was prepared with 9.5% TS and 4.8% oil. IP(h) is the point at which there is a large increase in oxygen uptake (a rapid drop in oxygen tension). The sample tested contained 4g of oil and 4g of matrix. The slow oxygen uptake in these samples is due in part to oxygen uptake by the broccoli matrix.

Example 10 - Biomass as encapsulant Raw biomass was cut into small pieces, boiling water was first added and the mixture was mixed to obtain an aqueous suspension containing biomass (5% TS). The pH of this mixture was adjusted to 7.50 using 2N NaOH. The mixture was then heat treated at 75°C for 2 minutes or 100°C for 30 minutes and then cooled to 60°C. Omega-3 oil (1:1 biomass solids:oil ratio) was added to the aqueous phase suspension (60° C.) and homogenized using an Ultraturrax at 15,000 rpm for 3 minutes to form an emulsion. prepared and lyophilized to obtain a powder (50% oil).

The results are shown in Table 5 and Figures 15-23. These results indicate the relative oxidative stability of the oil, with the length of the induction period related to the amount of protection the encapsulant affords the oil under accelerated conditions.

Example 11 - Effect of protein added to biomass as encapsulant Raw carrots were cut into small pieces, added to boiling water and mixed to obtain an aqueous suspension containing biomass (5% TS). The pH of this mixture was adjusted to 7.50 using 2N NaOH. The mixture was then heat treated at 90°C for 5 minutes and then cooled to 60°C. Different protein dispersions (10% TS) (sodium caseinate, soy protein isolate, pea protein) were added to the carrot suspension to obtain a 1:2 protein:CHO ratio. Add tuna oil to the carrot-protein mixture (60° C.) and homogenize using an Ultraturrax at 15,000 rpm for 3 minutes to prepare an emulsion, which is lyophilized to powder (25% oil). got

The results are shown in FIG. Only the carrot-pea protein omega-3 powder had a clear IP (38 hours). All other samples experienced a sudden increase in pressure during the Oxipres test.

Example 12 - Matcha as an encapsulating agent with and without added carbohydrate Matcha powder (with or without added maltodextrin) was reconstituted in water (45°C, 1 hour). The protein-to-carbohydrate (CHO) ratio (weight ratio) in the different formulations was 8:9 (matcha only), 1:2, 1:3, and 1:4 for the formulation with added maltodextrin. Tuna oil was added and dispersed for 3 minutes using a Silverson mixer (Silverson L4R, Silverson Machines Ltd., Chesham, Buckinghamshire, UK) and the emulsion was homogenized at 250/100 bar (Avestin Emulsiflex C5, Avestin Inc., Ottawa, Ontario, Canada). The emulsion (15% total solids) was spray dried to obtain a powder containing 25% fish oil (dry basis).

The Oxipres test at 80° C. and an initial 5 bar oxygen pressure is shown in FIG. The slow oxygen uptake in these samples with protein-carbohydrate ratios of 1:2 and 8:9 is due in part to oxygen uptake by the matrix. A clear IP is shown for tuna oil only and samples with protein-carbohydrate ratios of 1:4 and 1:3.

Example 13 - Oxipres Results Showing Stability of Spray Dried Omega-3 Oil Powder Comparing "Lyophilized Broccoli" Powder Versus "Sodium Caseinate + Carbohydrate" as Encapsulating Agent of water to 5% TS and allowed to hydrate for 1 hour. Tuna oil heated to 60° C. was added and homogenized for 5 minutes at maximum speed using a Silverson emulsifier. The emulsion was then spray dried. For comparison, a MicroMAX® formulation using a heated sodium caseinate-glucose dry glucose sugar solution heated at 100° C. for 30 minutes was used as the encapsulating medium. The same oil was added and homogenized at 60° C. and 180/80 bar pressure. Both formulations were spray dried in a laboratory scale Drytec spray dryer using a twin fluid nozzle (4 bar pressure). The spray-dried powder was tested at room temperature at 80° C. and an initial oxygen pressure of 5 bar using an Oxipres unit.

Oxipres test results (Figure 26) were tested at 80°C and an initial 5 bar oxygen pressure compared to the induction period for 50% tuna oil powder using heated casein-carbohydrate as the encapsulant, spray drying (50 % tuna oil) shows the induction period (IP) of omega-3 broccoli powder. There is a clear IP for both samples. The results show that encapsulated oil via the method as described herein is more resistant to oxygen degradation than encapsulated oil using MicroMAX® technology.

Example 14 - Comparison between the use of fermented and non-fermented broccoli as an encapsulant to produce omega-3 oil broccoli powder.
Broccoli puree (heated at 75° C. for 2 minutes or 100° C. for 30 minutes) or fermented broccoli puree (± heat treatment before fermentation) was prepared (5% TS). Hi-DHA tuna oil was added and homogenized using a Silverson emulsifier mixer (60° C. for 5 minutes) to form an emulsion. All formulations were lyophilized. The powder was tested on the Oxipres at 80° C. and an initial oxygen pressure of 5 bar and the results are shown in FIG. There is a clear IP for the sample with unfermented broccoli powder, but no clear IP for the sample with fermented broccoli puree.

Example 15 - Comparison of stability of EPA and DHA in tuna oil powder versus unencapsulated tuna oil Powders selected from Examples 5 and 8 and unrefined tuna oil (unencapsulated) were lightly capped. Stored in an oven at 40°C in sealed bottles. Fatty acid analysis of oils in oil and powder samples measured using gas chromatography. Table 6 shows the content of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) in the original tuna oil and in the stored samples. The higher EPA and DHA content in the preserved powder compared to the EPA and DHA in the unrefined oil after 1 month at 40°C clearly demonstrates the protection of the EPA and DHA encapsulants against oxidation. Proving.

Example 16 - Headspace Analysis of Secondary Oxidation Products from Omega-3 Oil Powders Using Selected Biomass as Encapsulating Media Lyophilized omega-3 oil powders from Examples 8 and 10 were C. for 14 months, selected powders from these examples were sampled and stored in lightly capped bottles in a 40.degree. C. oven for 4 weeks for analysis. Analysis of headspace secondary oxidation products was performed using GC-MS.

The results are shown in FIG. A lower amount of propanal and EE-2,4-heptadienal means a higher stability to oxidation of the sample. From these results, the best protection against omega-3 oxidation in these examples was obtained in broccoli, mushrooms and Brussels sprouts.

Example 17 Oxygen Uptake of Tableted and Extruded Formulations Using 50% Omega-3 Oil Broccoli Powder Lyophilized omega-3 oil broccoli powder (50% Hi-DHA tuna oil) was used. For extruded samples, freeze-dried omega-3 oil broccoli powder (20 wt%) and corn flour (80 wt%) were dry blended and subjected to low shear snack extrusion screw profiles at 60°C, 100°C, 140°C, Feed through the extruder (DSE32-II Twinscrew Lab Extruder) using a barrel temperature of 140° C. (powder feed throat to mold end) and a mold pressure of about 5 bar. For the tableting formulation, freeze-dried omega-3 oil broccoli powder (50% by weight) and skim milk powder (50% by weight) as an excipient were dry blended to prepare tablets (φ1.3 cm x 0.5 cm). . The oxygen uptake of the samples was evaluated by the Oxipres test at 80° C. and an initial oxygen pressure of 5 bar.

The results are shown in FIG. The samples tested were 40 g in extrudate (36 g matrix and 4 g oil) and 16 g in tablet form (16 g excipients and 4 g oil).

Example 18 - Oxygen uptake of omega-3 oil broccoli powder made by pre-treating broccoli biomass and/or post-treating emulsion Make up to 5% aqueous solids (50°C, 60 min hydration), add oil and homogenize to obtain a stable emulsion or suspension (9.5% TS and 4.8% oil) was formed. The emulsion was lyophilized to produce a 50% oil powder. The sample tested contained 4g of oil and 4g of matrix. In this example, the biomass (aqueous phase) is either pretreated in a water bath using ultrasound (40 KHz/180 W) for 7.5 minutes or microwave (750 W) up to 76.3° C. for 2.5 minutes. , or high pressure (6000 bar) at 25° C. for 3 minutes, or microwave (750 W) up to 76.3° C. for 2.5 minutes followed by post-treatment. Oxygen uptake of freeze-dried broccoli powder prepared by biomass pretreatment (5% TS broccoli suspension) or emulsion post-treatment (4.8% oil and 9.5% TS emulsion) was evaluated by the Oxipres test at 80° C. and an initial oxygen pressure of 5 bar.

The results are shown in FIG. The slow oxygen uptake in these samples is due in part to oxygen uptake by the broccoli matrix.

Example 19 - Sulforaphane Content of Broccoli Aqueous Suspensions and Emulsions Fresh broccoli florets were steamed (core temperature 60°C/5 min) and cooled to room temperature. An aqueous suspension (5 wt% broccoli solids) and an emulsion with medium chain triglycerides (5 wt% broccoli solids and 5 wt% medium chain triglycerides) were prepared and held at 25°C for 4 hours. . Aqueous suspensions and emulsions were extracted with ethyl acetate and sulforaphane content was measured using UPLC.
The sulforaphane content was 13% higher in the emulsion than in the aqueous suspension (expressed as mg/g dry weight of broccoli solids). This demonstrates that sulforaphane is more stable in emulsion.

Example 20 - Sulforaphane Content of Aqueous Suspensions and Emulsions of Preserved Freeze-Dried Broccoli was prepared as The sulforaphane content of the powder was determined after cryopreservation (-18°C for several months). After extracting sulforaphane from the samples, sulforaphane content was measured (using ethyl acetate/hexane mixtures). The sulforaphane content (expressed as mg/g dry weight of broccoli solids) in freeze-dried omega-3 oil broccoli powder (50% oil) was about two times higher than freeze-dried broccoli powder (no oil). This demonstrates that the presence of oil stabilized sulforaphane during long-term storage.

Example 21 - Discussion Instead of using purified proteins and carbohydrates as the encapsulant, these experiments demonstrate that whole biomass can be used as the encapsulant, obviating the need to purify and separate proteins and carbohydrates from the biomass source. demonstrated. Emulsions and suspensions, encapsulating media, powders, and entrapped and encapsulated bioactive agent(s) and/or bioactive precursor(s) can be used by using proteins and carbohydrates from the same biomass source. ) can be reduced. Methods as described herein may contribute to stabilization and may serve as a delivery vehicle for bioactive agent(s) and/or bioactive precursor(s). offers the added advantage of utilizing all the components inherent in biomass. Many biomass sources (e.g. plants, fruits and vegetables, algae, fungi) contain many nutrients (proteins, carbohydrates, fiber) and phytonutrients (e.g. carotenoids, polyphenols, sulfur-containing compounds, tocopherols, glucosinolates, etc.). ), which also have good nutritional and health-promoting properties (Hounsome et al., 2008). Due to the ubiquitous presence of proteins, carbohydrates, phytonutrients, and other minor components (e.g., vitamin C) present in biomass, whole biomass (or fractions thereof) can be used instead of refined individual components. The phytonutrients in the biomass can be utilized in place of components purified from the biomass for encapsulation and delivery of sensitive bioactive agent(s) and/or bioactive precursor(s). presents an advantage when using In addition, methods such as those described herein provide encapsulated oils with high protection/resistance to oxidative degradation. A comparison of IP for unrefined oil and oil encapsulated using the method as described herein is shown in Table 7.

It will be appreciated by those skilled in the art that many changes and/or modifications may be made to the present invention, as shown in the specific embodiments thereof, without departing from the spirit or scope of the invention as broadly described. will be understood. As such, embodiments of the present invention are to be considered in all respects as illustrative and not restrictive.

This application claims priority from Australian Provisional Application No. 2018900326, entitled "Method of producing an emulsion or suspension and products produced therefrom", filed on February 2, 2018, the entire contents of which incorporated herein by reference.

All publications discussed and/or referenced herein are incorporated herein in their entirety.

Any description of documents, acts, materials, devices, articles, etc. contained herein is solely for the purpose of providing a context for the invention. It is acknowledged that some or all of these matters were part of the prior art base or common general knowledge in the fields relevant to the present invention as they existed prior to the priority date of each claim in this application. shall not be deemed to have been made.

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Claims (13)

1. A method of producing a powder comprising entrapped or encapsulated bioactive agents and/or bioactive precursors comprising:
i) obtaining an aqueous mixture containing proteins and carbohydrates from the biomass of a single species of organism ;
ii) adding oil to said aqueous mixture;
iii) forming an emulsion or suspension comprising said bioactive agent and/or bioactive precursor;
iv) forming a powder comprising entrapped or encapsulated bioactive agents and/or bioactive precursors from said emulsion or suspension ;
Said method , wherein said biomass is selected from vegetables, tomatoes and avocados . 2. The method of claim 1, wherein the vegetables are selected from broccoli, cauliflower, kale, carrots, onions, garlic, Brussels sprouts, spinach, snow peas, and asparagus. The powder has the following characteristics:
a) having an induction period of 10 to 300 hours , measured at 80° C. and an initial oxygen pressure of 5 bar;
b ) 5 % to 50 % oil by weight;
c ) 10 % to 40 % by weight of oil; and d) the oil content of the emulsion or suspension before forming the powder is 1 % to 10% by weight.
2. The method of claim 1, comprising one or more of: said aqueous mixture further comprising proteins and carbohydrates from at least one further biomass from a single species of organism , and/or said biomass and/or said further biomass is
a ) a protein to carbohydrate ratio of 1 :1 to 1:10.5;
b ) a protein to carbohydrate ratio of 1 :4.5 to 4 :1;
c ) a protein to carbohydrate ratio of 1 :2.5 to 2 :1;
d) the Brassicaceae plant and, as said bioactive substance, an isothiocyanate;
e) Brassicaceae plants and, as said bioactive precursors, glucosinolates and/or glucoraphanin;
f) onion and, as said bioactive substance, one or more of quercetin, allicin and phenolic acid;
g) garlic and, as said bioactive substances, one or more of allicin and ajoene; and h) fruits and/or vegetables containing polyphenols,
including one or more of
The method according to any one of claims 1-3. wherein the bioactive agent and/or bioactive precursor is
a ) components of said biomass;
b ) said oil of step ii) or a component thereof;
c ) ingredients added to said oil before said oil is added to said aqueous mixture in step ii);
d ) ingredients injected into said oil before or during step ii);
e ) a component of said further biomass;
f ) ingredients added in steps i), ii) and iii) of the method;
g ) said bioactive agent is a ) and b );
h ) said bioactive agent is formed at or after step i), ii) or iii);
i ) sensitive to degradation by one or more of oxygen, temperature, pH, moisture, and light;
j ) is a phytonutrient;
k ) fatty acids, isothiocyanates, quercetin, allicin, ajoene, vitamin A, vitamin D, vitamin E, tocopherols, tocotrienols, vitamin K, beta-carotene, lycopene, lutein, zeaxanthin, stigmasterol, beta-sitosterol, campesterol, antioxidants coenzyme Q10, astaxanthin, cannabinoids, cannabidiol, and polyphenols;
l ) when said bioactive agent is a fatty acid, said fatty acid is an omega-3, omega-6, or omega-9 fatty acid;
m ) when said bioactive agent is an omega-3 fatty acid, said omega-3 fatty acid is alpha-linolenic acid (ALA), eicosapentaenoic acid (EPA), docosapentaenoic acid (DPA), and docosahexaenoic acid (DHA) ), the powder according to any one of claims 1 to 4 . a) pretreating said biomass;
where the pretreatment is
i) heating,
ii) maceration;
iii) microwave treatment,
iv) exposure to low frequency sound waves (ultrasound);
v) pulsed electric field treatment;
vi) static high pressure,
vii) extrusion,
viii) enzymatic treatment,
ix) fermentation,
x) an extraction or separation process, and xi) drying; and/or b) post-treating said emulsion or suspension to reduce microbial activity.
wherein the post-treatment is
i) heating,
ii) microwave treatment;
iii) UV treatment, and iv) high pressure treatment,
The method of any one of claims 1-5 , further comprising: Said oil has the following characteristics:
a) containing one or more fatty acids;
b) fish oil, krill oil, marine oil, canola oil, sunflower oil, avocado oil, soybean oil, borage oil, evening primrose oil, safflower oil, linseed oil, olive oil, pumpkin seed oil, hemp seed oil, wheat germ oil, palm oil, selected from one or more of palm olein, palm kernel oil, coconut oil, medium chain triglycerides, and grape seed oil; and c) fish or marine oil, wherein said fish or marine oil is tuna oil, herring oil. oil, mackerel oil, sardine oil, cod liver oil, menhaden oil, shark oil, algal oil, squid oil, and squid liver oil;
The method of any one of claims 1-6, comprising one or more of forming said powder comprises spray drying, freeze drying, reflectance window drying, or drum drying; , resistant to degradation. A method according to any preceding claim, wherein the plant is broccoli (Brassica oleracea variety italica). A method according to any one of the preceding claims, wherein the plant is cauliflower (Brassica oleracea variety botrytis). A powder produced by the method according to any one of claims 1-10 . A product comprising the powder of claim 11. The powder has the following characteristics:
a) said entrapped or encapsulated bioactive agents and/or bioactive precursors are resistant to oxygen degradation compared to bioactive agents and/or bioactive precursors that are not entrapped or encapsulated;
b) said bioactive substances and/or bioactive precursors are fatty acids and/or oils;
c) said powder has an induction period of 10 hours to 300 hours , measured at 80° C. and an initial oxygen pressure of 5 bar;
d) said powder comprises 5 % to 50% by weight of oil;
e) the powder contains 10 % to 40% by weight of oil;
and/or the product has the following characteristics:
a) the entrapped or encapsulated bioactive agents and/or bioactive precursors in the product are reduced by degradation compared to the same product comprising unencapsulated or unencapsulated bioactive agents and/or bioactive precursors; resistant;
b) said product is a cream, gel tablet, liquid, pill, capsule, powder, or extruded product;
c) the product is a food, food ingredient, supplement, cosmetic, or cosmetic ingredient;
d) said product comprises omega-3 polyunsaturated fatty acids;
e) when said product is food, said food is animal feed; and f) when said product is animal feed, said animal feed is aquaculture feed;
having one or more of
A powder according to claim 11 or a product according to claim 12 .