DESC:
FIELD OF INVENTION
[0001] The present invention relates to microcapsules and method of preparing the same. In particular, the invention relates to dry form of microcapsules comprising algal oil rich in DHA.

BACKGROUND OF THE INVENTION
[0002] The background description includes information that may be useful in understanding the present invention. It is not an admission that any of the information provided herein is prior art or relevant to the presently claimed invention, or that any publication specifically or implicitly referenced is prior art.
[0003] Foods rich in omega 3 fatty acids (PUFAs) are in high demand as their intake is associated with improved cardiovascular health. PUFAs may decrease plasma triglycerides levels, blood pressure, and inflammation. While seafood (fish) is the richest source of PUFAs, access to it may be restricted in places which are not near water bodies. Seafood derived PUFAs are also associated with a strong ‘fishy’ taste, which is hard to remove. Further, people with certain dietary or religious beliefs, such as vegans or vegetarians may not prefer to consume fish derived products.
[0004] While there are numerous food supplements and additives in the market comprising PUFAs, a common limitation to them is their storage requirements (cold dark place) apart from having limited shelf life, strong smell, low bioavailability and low stability. They also suffer from low solubility in water, particularly in ambient to low temperature water, which restrict their application, and increases wastage.
[0005] Indian patent application number 5175/DELNP/2008 describes encapsulated compositions and methods for reducing oxidation of an oxidizable material in a substantially water free environment.
[0006] Indian patent application number 4853/DELNP/2008 describes microcapsules free of animal byproducts.
[0007] Given the limitations in these documents, amongst others, there is a need to develop microencapsulated PUFAs with superior characteristics which is palatable, readily soluble in liquids, particularly water, has an extended shelf life, and has high bioavailability.
[0008] All publications herein are incorporated by reference to the same extent as if each individual publication or patent application were specifically and individually indicated to be incorporated by reference. Where a definition or use of a term in an incorporated reference is inconsistent or contrary to the definition of that term provided herein, the definition of that term provided herein applies and the definition of that term in the reference does not apply.
[0009] Groupings of alternative elements or embodiments of the invention disclosed herein are not to be construed as limitations. Each group member can be referred to and claimed individually or in any combination with other members of the group or other elements found herein. One or more members of a group can be included in, or deleted from, a group for reasons of convenience and/or patentability. When any such inclusion or deletion occurs, the specification is herein deemed to contain the group as modified thus fulfilling the written description of all Markush groups used in the appended claims.

SUMMARY
[00010] An aspect of the present disclosure provides an agglomeration of microcapsules, the agglomeration comprising: the microcapsules comprising at least one loading substance encapsulated by a shell, said shell comprising at least one milk based protein; and alpha cyclodextrin, wherein said loading substance weight percentage in said agglomeration is in the range of about 10% to about 30%. In an embodiment, the loading substance weight percentage in said agglomeration is in the range of about 20% to about 28%.
[00011] In an embodiment, the at least one loading substance is winterized algal oil comprising at least 40% DHA. In an embodiment, the shell comprises: casein having weight percentage in the range of about 10% to about 20% and soy lecithin having weight percentage in the range of about 10% to about 20%. In an embodiment, the soy lecithin acts as an emulsifier.
[00012] In an embodiment, the agglomeration further comprises: at least one stabilizer selected from the group consisting of sodium ascorbate, tri calcium phosphate, mannitol, xylitol and combinations thereof; at least one bulking agent selected from the group consisting of alpha cyclodextrin, beta cyclodextrin, gamma cyclodextrin, waxy maize starch, waxy potato starch, and combinations thereof; at least one humectant selected from the group consisting of liquid glucose, corn syrup, solids, tapioca starch solids and combinations thereof; and at least one anti-oxidant selected from the group consisting of mixed tocopherol oil and natural tocopherol oil.
[00013] In an embodiment, the agglomeration further comprises: at least one sweetener, and wherein the at least one sweetener include one or a plurality of sugars; and at least one odor or taste masking agent, wherein the at least one odor or taste masking agent is trehalose. In an embodiment, the agglomeration exhibits cold water solubility of at least about 10 mg/ml at a temperature of about 25°C. In an embodiment, the agglomeration exhibits cold water solubility of at least about 100 mg/ml at a temperature of about 25°C. In an embodiment, the agglomeration exhibits shelf life of at least about 350 days. In an embodiment, the agglomeration exhibits shelf life of at least about 400 days.
[00014] In an embodiment, the microcapsules comprises 20-30% by weight of winterized algal oil comprising at least 40% DHA, said winterized algal oil encapsulated by a shell comprises casein having weight percentage in the range of 7-17% and soy lecithin having weight percentage in the range of 8-18%, said agglomeration further comprising alpha cyclodextrin having weight percentage in the range of 5-10.5%, ascorbate having weight percentage in the range of 0.2-0.6%, mixed tocopherol oils having weight percentage in the range of 0.1-0.3%, flavoring or masking agent having weight percentage in the range of 5-8%, sugar having weight percentage in the range of 7-11%, calcium phosphate having weight percentage in the range of 0.15-0.35%, mannitol having weight percentage in the range of 2-4%, and liquid glucose having weight percentage in the range of 15-25%. In an embodiment, the agglomeration exhibits cold water solubility of at least about 10 mg/ml at a temperature of about 25°C, and wherein said agglomeration exhibits shelf life of at least about 350 days. In an embodiment, the agglomeration exhibits cold water solubility of at least about 100 mg/ml at a temperature of about 25°C. In an embodiment, the agglomeration exhibits shelf life of at least about 400 days.
[00015] In an embodiment, the microcapsules comprises 20-28 ± 2% by weight of winterized algal oil comprising at least 40% DHA, said algal oil encapsulated by a shell comprising casein having weight percentage in the range of 12-16% ± 5% and soy lecithin having weight percentage in the range of 12-16% ± 5%, said agglomeration further comprising alpha cyclodextrin having weight percentage in the range of 6-9.75% ± 2%, ascorbate having weight percentage in the range of 0.3-0.4% ± 2%, mixed tocopherol oils having weight percentage in the range of 0.15-0.25 ± 2%, flavoring or masking agent having weight percentage in the range of 4.5-6.5 ± 5%, sugar having weight percentage in the range of 8-11 ± 5%, tri calcium phosphate having weight percentage in the range of 0.25-0.35 ± 5%, mannitol having weight percentage in the range of 2.5 -3.5 ± 5%, and liquid glucose having weight percentage in the range of 16-19 ± 2%.
[00016] Another aspect of the present disclosure relates to a method of making an agglomeration, said method comprising the steps of:
a. contacting water at a temperature in the range of 42-48°C with soy lecithin under stirring conditions to dissolve soy lecithin in water to obtain solution A;
b. contacting liquid glucose to solution A under stirring conditions to obtain a slurry B while maintaining a temperature in the range of 42-48°C;
c. contacting ascorbate, casein, flavoring agent, sugar, tri calcium phosphate, sodium CMC, gums, cyclodextrin, trehalose and mannitol to slurry B under stirring conditions while maintaining a temperature in the range of 42-48°C to obtain a slurry C having Brix in the range of 18-20;
d. adjusting the temperature of slurry C to 70-75°C under stirring conditions to obtain a slurry D having Brix in the range of 20-22;
e. contacting algal oil & mixed Tocopherol at a temperature in the range of 70-75°C to slurry D at a temperature in the range of 55-65°C under stirring conditions to obtain an emulsion E having Brix in the range of 18-21;
f. adjusting the temperature of emulsion E to 38-42°C and contacting with masking flavor under stirring conditions to obtain a homogenized emulsion F having Brix in the range of 21-24; and
g. spray drying emulsion F to obtain agglomeration comprising encapsulated algal oil.

DETAILED DESCRIPTION OF THE INVENTION
[00017] The following is a detailed description of embodiments of the disclosure depicted in the accompanying drawings. The embodiments are in such detail as to clearly communicate the disclosure. However, the amount of detail offered is not intended to limit the anticipated variations of embodiments; on the contrary, the intention is to cover all modifications, equivalents, and alternatives falling within the spirit and scope of the present disclosure as defined by the appended claims.
[00018] In the following description, numerous specific details are set forth in order to provide a thorough understanding of embodiments of the present invention. It will be apparent to one skilled in the art that embodiments of the present invention may be practiced without some of these specific details.
[00019] Exemplary embodiments will now be described more fully hereinafter with reference to the accompanying drawings, in which exemplary embodiments are shown. These exemplary embodiments are provided only for illustrative purposes and so that this disclosure will be thorough and complete and will fully convey the scope of the invention to those of ordinary skill in the art. The invention disclosed may, however, be embodied in many different forms and should not be construed as limited to the embodiments set forth herein. Various modifications will be readily apparent to persons skilled in the art. The general principles defined herein may be applied to other embodiments and applications without departing from the spirit and scope of the invention. Moreover, all statements herein reciting embodiments of the invention, as well as specific examples thereof, are intended to encompass both structural and functional equivalents thereof. Additionally, it is intended that such equivalents include both currently known equivalents as well as equivalents developed in the future (i.e., any elements developed that perform the same function, regardless of structure). Also, the terminology and phraseology used is for the purpose of describing exemplary embodiments and should not be considered limiting. Thus, the present invention is to be accorded the widest scope encompassing numerous alternatives, modifications and equivalents consistent with the principles and features disclosed. For purpose of clarity, details relating to technical material that is known in the technical fields related to the invention have not been described in detail so as not to unnecessarily obscure the present invention.
[00020] Various terms as used herein are shown below. To the extent a term used in a claim is not defined below, it should be given the broadest definition persons in the pertinent art have given that term as reflected in printed publications and issued patents at the time of filing.
Definitions:
[00021] For convenience, before further description of the present disclosure, certain terms employed in the specification, and examples are collected here. These definitions should be read in the light of the remainder of the disclosure and understood as by a person of skill in the art. The terms used herein have the meanings recognized and known to those of skill in the art, however, for convenience and completeness, particular terms and their meanings are set forth below.
[00022] Throughout the specification and claims the word "comprise" and other forms of the word, such as "comprising" and "comprises," means including but not limited to, and is not intended to exclude, for example, other additives, components, integers, or steps.
[00023] As used in the description and the appended claims, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise.
[00024] The term "optional" or "optionally" means that the subsequently described event or circumstance can or cannot occur, and that the description includes instances where the event or circumstance occurs and instances where it does not.
[00025] A weight percent (wt.%) of a component, unless specifically stated to the contrary, is based on the total weight of the formulation or composition in which the component is included.
[00026] The term “microcapsule” refers to multicore, single-core, or a mixture of multicore and single-core microcapsules.
[00027] As various changes could be made in the above composition, products and methods without departing from the scope of the invention, it is intended that all matter contained in the above description and in the examples given below, shall be interpreted as illustrative and not in a limiting sense.
[00028] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure belongs. Although any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the disclosure, the preferred methods, and materials are now described. All publications mentioned herein are incorporated herein by reference. The present disclosure is not to be limited in scope by the specific embodiments described herein, which are intended for the purposes of exemplification only. Functionally-equivalent products, compositions, and methods are clearly within the scope of the disclosure, as described herein.
[00029] An aspect of the present disclosure provides an agglomeration of microcapsules, the agglomeration comprising: the microcapsules comprising at least one loading substance encapsulated by a shell, said shell comprising at least one milk based protein; and alpha cyclodextrin, wherein said loading substance weight percentage in said agglomeration is in the range of 10-30%. In an embodiment, the loading substance weight percentage in said agglomeration is in the range of about 20% to about 28%.
[00030] In an embodiment, the at least one loading substance is winterized algal oil comprising at least 40% DHA. In an embodiment, the shell comprises: casein having weight percentage in the range of about 10% to about 20% and soy lecithin having weight percentage in the range of about 10 to about 20%. In an embodiment, the soy lecithin acts as an emulsifier.
[00031] In an embodiment, the agglomeration further comprises: at least one stabilizer selected from the group consisting of sodium ascorbate, tri calcium phosphate, mannitol, xylitol and combinations thereof; at least one bulking agent selected from the group consisting of alpha cyclodextrin, beta cyclodextrin, gamma cyclodextrin, waxy maize starch, waxy potato starch, and combinations thereof; at least one humectant selected from the group consisting of liquid glucose, corn syrup, solids, tapioca starch solids and combinations thereof; and at least one anti-oxidant selected from the group consisting of mixed tocopherol oil and natural tocopherol oil.
[00032] In an embodiment, the agglomeration further comprises: at least one sweetener, and wherein the at least one sweetener include one or a plurality of sugars; and at least one odor or taste masking agent, wherein the at least one odor or taste masking agent is trehalose. In an embodiment, the agglomeration exhibits cold water solubility of at least about 10 mg/ml at a temperature of 25°C. In an embodiment, the agglomeration exhibits shelf life of at least about 350 days. In an embodiment, the agglomeration exhibits cold water solubility of at least about 100 mg/ml at a temperature of 25°C. In an embodiment, the agglomeration exhibits shelf life of at least about 400 days.
[00033] In an embodiment, the microcapsules comprises 20-30% by weight of winterized algal oil comprising at least 40% DHA, said winterized algal oil encapsulated by a shell comprises casein having weight percentage in the range of 7-17% and soy lecithin having weight percentage in the range of 8-18%, said agglomeration further comprising alpha cyclodextrin having weight percentage in the range of 5-10.5%, ascorbate having weight percentage in the range of 0.2-0.6%, mixed tocopherol oils having weight percentage in the range of 0.1-0.3%, flavoring or masking agent having weight percentage in the range of 5-8%, sugar having weight percentage in the range of 7-11%, calcium phosphate having weight percentage in the range of 0.15-0.35%, mannitol having weight percentage in the range of 2-4%, and liquid glucose having weight percentage in the range of 15-25%. In an embodiment, the agglomeration exhibits cold water solubility of at least about 10 mg/ml at a temperature of 25°C, and wherein said agglomeration exhibits shelf life of at least about 350 days. In an embodiment, the agglomeration exhibits cold water solubility of at least about 100 mg/ml at a temperature of 25°C. In an embodiment, the agglomeration exhibits shelf life of at least about 350 days.
[00034] In an embodiment, the microcapsules comprises 20-28 ± 2% by weight of winterized algal oil comprising at least 40% DHA, said algal oil encapsulated by a shell comprising casein having weight percentage in the range of 12-22% ± 5% and soy lecithin having weight percentage in the range of 12-16% ± 5%, said agglomeration further comprising alpha cyclodextrin having weight percentage in the range of 6-9.75% ± 2%, ascorbate having weight percentage in the range of 0.3-0.4% ± 2%, mixed tocopherol oils having weight percentage in the range of 0.15-0.25 ± 2%, flavoring or masking agent having weight percentage in the range of 4.5-6.5 ± 5%, sugar having weight percentage in the range of 8-11 ± 5%, tri calcium phosphate having weight percentage in the range of 0.25-0.35 ± 5%, mannitol having weight percentage in the range of 2.5 -3.5 ± 5%, and liquid glucose having weight percentage in the range of 16-19 ± 2%.
[00035] Another aspect of the present disclosure relates to a method of making an agglomeration, said method comprising the steps of:
a. contacting water at a temperature in the range of 42-48°C with soy lecithin under stirring conditions to dissolve soy lecithin in water to obtain solution A;
b. contacting liquid glucose to solution A under stirring conditions to obtain a slurry B while maintaining a temperature in the range of 42-48°C;
c. contacting ascorbate, casein, flavoring agent, sugar, tri calcium phosphate, sodium CMC, gums, cyclodextrin, trehalose and mannitol to slurry B under stirring conditions while maintaining a temperature in the range of 42-48°C to obtain a slurry C having Brix in the range of 18-20;
d. adjusting the temperature of slurry C to 70-75°C under stirring conditions to obtain a slurry D having Brix in the range of 20-22;
e. contacting algal oil & mixed Tocopherol at a temperature in the range of 70-75°C to slurry D at a temperature in the range of 60-65°C under stirring conditions to obtain an emulsion E having Brix in the range of 18-21;
f. adjusting the temperature of emulsion E to 38-42°C and contacting with masking flavor under stirring conditions to obtain a homogenized emulsion F having Brix in the range of 21-24; and
g. spray drying emulsion F to obtain agglomeration comprising encapsulated algal oil.
[00036] In an embodiment, an agglomeration of microcapsules includes at least one loading substance encapsulated by a shell, said shell including at least one plant based phospholipid and at least one milk based protein, wherein said loading substance weight percentage is in the range of 10-20%, and said at least one plant based phospholipid and at least one milk based protein weight percentage of both in said agglomeration is in the range of 20-40%.
[00037] In an embodiment, the loading substance includes at least one polyunsaturated fatty acid (PUFA). The PUFA can be at least one omega 3 fatty acid. Omega 3 fatty acids can be selected from any or a combination of alpha-linolenic acid (18:3, ALA), stearidonic acid (18:4), eicosatetraenoic acid (20:4), eicosapentaenoic acid (20:5; EPA), docosatetraenoic acid (22:4), n-3 docosapentaenoic acid (22:5; n-3DPA), and docosahexaenoic acid (22:6; DHA), but not limited thereto. In an embodiment, the PUFA can be at least one omega 6 fatty acid, selected from any or a combination of linoleic acid (18:2), gamma-linolenic acid (18:3), eicosadienoic acid (20:2), dihomo-gamma-linolenic acid (20:3), arachidonic acid (20:4), docosadienoic acid (22:2), adrenic acid (22:4), and n-6 docosapentaenoic acid (22:5), but not limited thereto. The fatty acid can be an omega-9 fatty acid, such as oleic acid (18:1), eicosenoic acid (20:1), mead acid (20:3), erucic acid (22:1), and nervonic acid (24:1), but not limited thereto. In a preferred embodiment, said loading substance comprises at least 40% by weight of DHA.
[00038] In an embodiment, the loading substance can be obtained from land based animals, water based animals, land based plants, or water based plants. The water based animals or plants may be fresh water or sea water based. In a preferred embodiment, the loading substance is obtained from algae. Commercially available algae-derived oils include those from Crypthecodinium cohnii and Schizochytrium sp. Other suitable species of algae, from which oil can be extracted, include, but not limited to, Aphanizomenonflos-aquae, Bacilliarophy sp., Botryococcus braunii, Chlorophyceae sp., Dunaliella tertiolecta, Euglena gracilis, Isochrysis galbana, Nannochloropsis salina, Nannochloris sp., Neochloris oleoabundans, Phaeodactylum tricornutwn, Pleurochrysis carterae, Prymnesiwn parvum, Scenedesmus dimorphus, Spirulina sp., and Tetraselmis chui.
[00039] In a preferred embodiment, the loading substance weight percentage is in the range of 20-30%, more preferably 20-28 ± 2%, and said loading substance is winterized algal oil including at least 40% DHA.
[00040] In an embodiment, the at least one plant based phospholipid may be a single purified phospholipid, such as distearoylphosphatidylcholine. In another embodiment, the phospholipid may be mixture of purified phospholipids, such as a mix of phosphatidylcholines. In still another embodiment, the phospholipid may be a mixture of different types of purified phospholipids, such as a mix of phosphatidylcholines and phosphatidylinositols or a mixture of phosphatidylcholines and phosphatidylethanolamines. In a preferred embodiment, the phospholipid may be a complex mix of phospholipids, such as a lecithin. Commercial sources of lecithin include soybeans, rice, sunflower seeds, chicken egg yolks, milk fat, bovine brain, bovine heart, and algae, but not limited thereto. Soy lecithin is rich in phosphatidylcholine, phosphatidylethanolamine, phosphatidylinositol, and phosphatidic acid. Lecithin may be de-oiled and treated such that it is an essentially pure mixture of phospholipids. Lecithin may be modified to make the phospholipids more water-soluble. In an exemplary embodiment, the lecithin is soy lecithin. Soy lecithin is also a very good emulsifier apart from having encapsulating properties. In an embodiment, soy lecithin can also act an emulsifier.
[00041] In an exemplary embodiment, the milk based protein is casein. Casein naturally forms micelles ranging in size from about 50 nm to about 500 nm, which are quite stable during processing. Caseinates are solubilized forms of casein produced by reaction with an alkaline substance. Common caseinate include: sodium caseinate, calcium caseinate, potassium caseinate, and ammonium caseinate, but not limited thereto. "Caseinates" is used herein to generally refer to these and other caseinates. Sodium caseinate is highly soluble and is used as an emulsifier.
[00042] In an embodiment, xanthan gum is another suitable encapsulating material that is not an animal by-product. Xanthan gum is a natural gum polysaccharide. It is produced by fermentation of glucose or sucrose by Xanthomonas campestris. Further examples of suitable encapsulating materials are polyphosphate, alginate, chitosan, carrageenan, starch, modified starch, oligofructans, konnyaku, alpha-lactalbumin, beta-lactoglobumin, ovalbumin, polysorbiton, maltodextrin (DE18, DE 21, DE40 etc.), cyclodextrins (alpha-, beta- and gamma-cyclodextrins), cellulose, cellulose ether, methyl cellulose, ethyl cellulose, hydropropylmethylcellulose, carboxymethylcellulose, hydroxypropyl cellulose, milk protein, canola protein, albumin, chitin, polylactides, poly-lactide-co-glycolides, derivatized chitin, poly-lysine, dilutan gum, locus bean gum, welan gum, and xanthan gum including combinations and mixtures thereof, but not limited thereto. In an embodiment, xanthan gum can act as a viscosity enhancer.
[00043] In an embodiment, the at least one plant based phospholipid is preferably soy lecithin and at least one milk based protein is preferably casein. In a preferred embodiment, casein weight percentage in the range of 10-20% and soy lecithin weight percentage in the range of 10-20%. In a more preferred embodiment, casein weight percentage in the range of 7-17% and soy lecithin weight percentage in the range of 8-18%. In a still more preferred embodiment, casein weight percentage in the range of 12-16% ± 5% and soy lecithin weight percentage in the range of 12-16% ± 5%.
[00044] In an embodiment, the agglomeration can further include at least one anti-oxidant. The anti-oxidant can stabilize the loading substance, which is susceptible to oxidation. The anti-oxidant may be synthetically or naturally derived. The anti-oxidant may be a mix of both. Suitable antioxidants include, but are not limited to, ascorbic acid and its salts, ascorbyl palmitate, ascorbyl stearate, anoxomer, N-acetylcysteine, benzyl isothiocyanate, o-, m- or p-amino benzoic acid (o is anthranilic acid, p is PABA), butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), caffeic acid, canthaxantin, alpha-carotene, beta-carotene, beta-caraotene, beta-apo-carotenoic acid, carnosol, carvacrol, cetyl gallate, chlorogenic acid, citric acid and its salts, clove extract, coffee bean extract, p-coumaric acid, 3,4-dihydroxybenzoic acid, N,N'-diphenyl-p-phenylenediamine (DPPD), dilauryl thiodipropionate, distearyl thiodipropionate, 2,6-di-tert-butylphenol, dodecyl gallate, edetic acid, ellagic acid, erythorbic acid, sodium erythorbate, esculetin, esculin, 6-ethoxy-l,2-dihydro-2,2,4-trimethylquinoline, ethyl gallate, ethyl maltol, ethylenediaminetetraacetic acid (EDTA), eucalyptus extract, eugenol, ferulic acid, flavonoids (e.g., catechin, epicatechin, epicatechin gallate, epigallocatechin (EGC), epigallocatechin gallate (EGCG), polyphenol epigallocatechin-3-gallate), flavones (e.g., apigenin, chrysin, luteolin), flavonols (e.g., datiscetin, myricetin, daemfero), flavanones, fraxetin, fumaric acid, gallic acid, gentian extract, gluconic acid, glycine, gum guaiacum, hesperetin, alpha-hydroxybenzyl phosphinic acid, hydroxycinammic acid, hydroxyglutaric acid, hydroquinone, Nhydroxysuccinic acid, hydroxytryrosol, hydroxyurea, lactic acid and its salts, lecithin, lecithin citrate; R-alpha-lipoic acid, lutein, lycopene, malic acid, maltol, 5-methoxy tryptamine, methyl gallate, monoglyceride citrate; monoisopropyl citrate; morin, beta-naphthoflavone, nordihydroguaiaretic acid (NDGA), octyl gallate, oxalic acid, palmityl citrate, phenothiazine, phosphatidylcholine, phosphoric acid, phosphates, phytic acid, phytylubichromel, pimento extract, propyl gallate, polyphosphates, quercetin, trans-resveratrol, rice bran extract, rosemary extract, rosmarinic acid, sage extract, sesamol, silymarin, sinapic acid, succinic acid, stearyl citrate, syringic acid, tartaric acid, thymol, tocopherols (i.e., alpha-, beta-, gamma- and deltatocopherol), tocotrienols (i.e., alpha-, beta-, gamma- and delta-tocotrienols), tyrosol, vanilic acid, 2,6-di-tert-butyl-4-hydroxymethylphenol (i.e., lonox 100), 2,4-(tris-3',5'-bi-tert-butyl-4'-hydroxybenzyl)-mesitylene (i.e., lonox 330), 2,4,5-trihydroxybutyrophenone, ubiquinone, tertiary butyl hydroquinone (TBHQ), thiodipropionic acid, trihydroxy butyrophenone, tryptamine, tyramine, uric acid, vitamin K and derivates, vitamin Q10, wheat germ oil, zeaxanthin, or combinations thereof. In a preferred embodiment, the anti-oxidant is mixed tocopherols. Mixed tocopherols oil includes naturally occurring d-alpha, d-beta, d-gamma and d-delta tocopherols, which can extend the shelf life of the algal oil comprising DHA. In an embodiment, the anti-oxidant weight percentage is in the range of 0.1-0.3%, more preferably in the range of 0.15-0.25 ± 2%, and most preferably in the range of 0.18-0.23% ± 2%.
[00045] In an embodiment, the agglomeration can further include at least one stabilizer selected from any or a combination of sodium ascorbate, calcium ascorbate, potassium ascorbate, magnesium ascorbate, Ester-C, mannitol and xylitol,but not limited thereto. In a preferred embodiment, stabilizer(s) include(s) a combination of sodium ascorbate and mannitol. Mannitol is a sugar alcohol having low hygoscopicity, which makes it useful as a coating agent and stabilizer. Mannitol can be produced by industrial synthesis, biosynthesis, or natural extraction. Industrially mannitol is typically produced by hydrogenation of fructose. Mannitol can also be extracted from seaweed. Sodium ascorbate is a mineral salt of ascorbic acid (vitamin C), which can be used for stabilization of dry form of algal oil comprising DHA. Tri calcium phosphate can act as an anti-caking agent. Stabilizer weight percentage is in the range of 0.15-0.35, and preferably in the range of 0.25-0.35 ± 5%. In a preferred embodiment, stabilizer(s) include a combination of ascorbates. In an embodiment, ascorbate is present as sodium ascorbate having weight percentage in the range of 0.2-0.6%, preferably in the range of 0.3-0.45%, and more preferably in the range of 0.3-0.4% ± 2%. In an embodiment, mannitol weight percentage is in the range of 2-4%, preferably in the range of 2-4%, and more preferably in the range of 2.5-3.5% ± 5%.
[00046] In an embodiment, the agglomeration can further include at least one bulking agent selected from any or a combination of alpha cyclodextrin, beta cyclodextrin, gamma cyclodextrin, waxy maize starch, waxy potato starch, and the likes. In a preferred embodiment, bulking agent is alpha cyclodextrin. Alpha cyclodextrin is a polysaccharide of six glucose units that are covalently attached end to end via a-1, 4 linkages alpha cyclodextrin is multifunctional, which is also useful in encapsulation of oils, and can act as a masking agent for masking the taste and odor. In a preferred embodiment, the bulking agent is alpha cyclodextrin. In an embodiment, alpha cyclodextrin weight percentage is in the range of 5-15%, preferably 5-10.5%, and more preferably in the range of 6-9.75% ± 2%.
[00047] In an embodiment, the agglomeration can further include at least one at least one anti-caking agent including any or a combination of tri calcium phosphate, magnesium stearate, and the likes. In a preferred embodiment, the anti-caking agent is tri calcium phosphate. In an embodiment, tri calcium phosphate weight percentage is in the range of 0.15-0.4%, preferably in the range of 0.15-0.35%, and more preferably in the range of 0.25-0.35% ± 5%.
[00048] In an embodiment, the agglomeration can further include at least one humectant including any or a combination of liquid glucose, corn syrup, solids, tapioca starch solids, and combinations thereof. In a preferred embodiment, the humectant is liquid glucose. Liquid glucose can also be used as a thickening agent. Industrially, liquid glucose is manufactured by enzymatic degradation of vegetable raw materials, such as corn or potatoes. Starch from the raw materials is liquefied either by heat treatment or using a-amylase. Then cyclodextrin glycosyltransferase (CGTase) is added for enzymatic conversion. CGTases can synthesize all forms of cyclodextrins, thus for the production of the a-cyclodextrin, one specific enzyme is used. In an embodiment, alpha cyclodextrin weight percentage is in the range of 5-15%, preferably 5-10.5%, and more preferably in the range of 6-9.75% ± 2%.
[00049] In an embodiment, the agglomeration can further include at least one viscosity enhancer including any or a combinations of xanthan gum, carrageenan, sodium caboxy methyl cellulose, and the likes. In a preferred embodiment, the viscosity enhancer is a combination of sodium CMC and xanthan gum. In an embodiment, viscosity enhancer weight percentage is in the range of 0.5-1.5%, preferably having weight percentage in the range of 0.6-1.2%, and more preferably in the range of 0.7-0.9% ± 5%. In a preferred embodiment, sodium CMC weight percentage is in the range of 0.75-0.85% ± 5% and xanthan gum weight percentage is in the range of 0.25-0.35% ± 5%.
[00050] In an embodiment, the agglomeration can further include at least one sweetener including any or a combination of sugars such as dextrose, corn syrup, fructo-oligosaccharide, maltitol, natural sugar, fructose, fruit sugar, high maltose sugar powder and the likes. In an embodiment, sugar weight percentage is in the range of 7-11, preferably in the range of 8-10%, and more preferably in the range of 8-9.5% ± 5%.
[00051] In an embodiment, the agglomeration can further include at least one odor or taste masking agent including any or a combination of trehalose, vanilla, and the likes. In a preferred embodiment, trehalose acts as an odor masking agent. In a preferred embodiment, vanilla acts as a taste masking agent. In an embodiment, odor or taste masking agent weight percentage is in the range of 1-3%. In a preferred embodiment, trehalose weight percentage is in the range of 1-3%, preferably in the range of 1.5-2.5%, more preferably 2% ± 5%. In a preferred embodiment, vanilla weight percentage is in the range of 2-4%, preferably in the range of 2.5-3.5%, and more preferably in the range of 2.7-3% ± 5%.
[00052] In an embodiment, the agglomeration of microcapsules can include 20-30% by weight of winterized algal oil including at least 40% DHA, said algal oil encapsulated by a shell including casein having weight percentage in the range of 7-20% and soy lecithin having weight percentage in the range of 8-20%, said agglomeration further including alpha cyclodextrin having weight percentage in the range of 5-10.5%, ascorbate having weight percentage in the range of 0.2-0.6%, mixed tocopherol oils having weight percentage in the range of 0.1-0.3%, flavoring or masking agent having weight percentage in the range of 5-8%, sugar having weight percentage in the range of 7-11%, tri calcium phosphate having weight percentage in the range of 0.15-0.35%, mannitol having weight percentage in the range of 2-4%, viscosity enhancer having weight percentage in the range of 0.5-1.5%, and liquid glucose having weight percentage in the range of 15-25%. However, it should be appreciated that the agglomeration can include any of the abovementioned and/or other ingredients and/or excipients in any other proportion (wt.%) as known to or appreciated by a person skilled in the art without departing from the scope and spirit of the present invention.
[00053] In an embodiment, the agglomeration of microcapsulesincludes 20-28% ± 2% by weight of winterized algal oil including at least 40% DHA, said algal oil encapsulated by a shell including casein having weight percentage in the range of 12-16% ± 5% and soy lecithin having weight percentage in the range of 12-16% ± 5%, said agglomeration further including alpha cyclodextrin having weight percentage in the range of 6-9.75% ± 2%, ascorbate having weight percentage in the range of 0.3-0.4% ± 0.2%, mixed tocopherol oils having weight percentage in the range of 0.15-0.25% ± 0.2%, flavoring or masking agent having weight percentage in the range of 4.5-8% ± 4%, sugar having weight percentage in the range of 8-11% ± 5%, tri calcium phosphate having weight percentage in the range of 0.25-0.35% ± 0.15%, mannitol having weight percentage in the range of 2.5-3.5% ± 0.5%,, and liquid glucose having weight percentage in the range of 16-19% ± 2%,
[00054] Another aspect of the present disclosure relates to a method of making an agglomeration, said method including the steps of: contacting water at a temperature in the range of 42-48°C with soy lecithin under stirring conditions to dissolve soy lecithin in water to obtain solution A; contacting liquid glucose to solution A under stirring conditions to obtain a slurry B while maintaining a temperature in the range of 42-48°C; contacting ascorbate, casein, flavoring agent, sugar, tri calcium phosphate, sodium CMC, gums, cyclodextrin, trehalose and mannitol to slurry B under stirring conditions while maintaining a temperature in the range of 42-48°C to obtain a slurry C having Brix in the range of 18-20; adjusting the temperature of slurry C to 70-75°C under stirring conditions to obtain a slurry D having Brix in the range of 20-22; contacting algal oil & mixed Tocopherol at a temperature in the range of 70-75°C to slurry D at a temperature in the range of 70-75°C under stirring conditions to obtain an emulsion E having Brix in the range of 18-20; adjusting the temperature of emulsion E to 38-42°C and contacting with masking flavor under stirring conditions to obtain a homogenized emulsion F having Brix in the range of 21-24; and spray drying emulsion F to obtain agglomeration comprising encapsulated algal oil.
[00055] The agglomeration of microcapsules, as realized in accordance with embodiments of the present disclosure, surprisingly exhibits exceptional cold water solubility of at least 10 mg/ml at a temperature of about 25°C, preferably at least about 50 mg/ml at a temperature of about 25°C, and most preferably at least about 100 mg/ml at a temperature of about 25°C. Further, the agglomeration of microcapsules, as realized in accordance with embodiments of the present disclosure, exhibits superior shelf life as compared to conventional compositions, wherein the agglomeration exhibits shelf life of at least 350 days, preferably at least 400 days.
[00056] The agglomeration of microcapsules, as realized in accordance with embodiments of the present disclosure, can be used as additives in beverages, confectionery, dry blend powders, gummies, bellies, malted beverages, medical nutrition, weaning formulas, milk and milk products, bakery products, etc.
[00057] While the invention has been described with reference to an exemplary embodiment(s), it will be understood by those skilled in the art that various changes may be made and equivalents may be substituted for elements thereof without departing from the scope of the invention. In addition, many modifications may be made to adapt a particular situation or material to the teachings of the invention without departing from the essential scope thereof. Therefore, it is intended that the invention not be limited to the particular embodiment(s) disclosed, but that the invention will include all embodiments falling within the scope of the appended claims.
[00058] Although the subject matter has been described in considerable detail with reference to certain preferred embodiments thereof, other embodiments are possible.

EXAMPLES
[00059] The disclosure will now be illustrated with working examples, which is intended to illustrate the working of disclosure and not intended to take restrictively to imply any limitations on the scope of the present disclosure. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood to one of ordinary skill in the art to which this disclosure belongs. Although methods and materials similar or equivalent to those described herein can be used in the practice of the disclosed methods and compositions, the exemplary methods, devices and materials are described herein. It is to be understood that this disclosure is not limited to particular methods, and experimental conditions described, as such methods and conditions may vary.
[00060] Free flowing, freely cold water soluble agglomeration of microcapsules comprising algal oil
[00061] Table 1 below provide an exemplary formula for a free flowing and freely cold water soluble agglomeration of microcapsules in powder form encapsulating algal oil comprising at least 40% DHA .

Table 1
SL. No. Raw Material Name Function (at least in part) measurement Quantity
1 DHA Oil (from Algae) (40% DHA) Active Ingredient Gram 23.4
2 Alpha Cyclodextrin Encapsulation effect, thickening agent & product stability Gram 9.22
3 Sodium caseinate (90%) Protein source & dispersion support of final product in Cold water Gram 14.74
4 Sodium ascorbate Buffering agent and stabilizer Gram 0.38
5 Soy lecithin liquid Emulsifier & encapsulating agent Gram 14.95
6 Mixed Tocopherols oil Anti-oxidant effect on fatty oil Ml 0.21
7 Vanilla flavor – powder Flavoring agent Gram 2.95
8 Sugar – Pharma grade Sweetness Gram 8.48
9 Tri Calcium Phosphate anticaking/ anti-lumping effect Gram 0.26
10 Mannitol Low calorie substitute of sugar Gram 2.76
11 Xanthum Gum Thickness Gram 0.3
12 Liquid Glucose Sweetner & bricks providing agent Gram 17.94
13 Trehalose Masking off notes Gram 2.021
14 Sodium CMC viscosity enhancer Gram 0.82
15 Masking Flavor Masking Gram 1.6

[00062] Method of preparing agglomeration of microcapsules comprising algal oil
[00063] The formulation provided in Table 1 of Example 1 above was prepared by the following methodology:
[00064] Firstly, water at a temperature of about 45°C was mixed with soy lecithin under stirring conditions in order to dissolve the soy lecithin in water to obtain a solution A.
[00065] Secondly, liquid glucose was contacted with solution A under stirring conditions to obtain a slurry B while maintaining the temperature in a range of 42-48°C.
[00066] Thirdly, ascorbate, casein, flavoring agent, sugar, tri calcium phosphate, Sodium CMC, Gums, cyclodextrin, Trehalose and mannitol were contacted with slurry B under stirring conditions while maintaining a temperature in the range of 42-48°C to obtain a slurry C having Brix in the range of 18-20. Subsequently, the temperature of slurry C was adjusted to 70-75°C under stirring conditions to obtain a slurry D having Brix in the range of 20-22.
[00067] Separately, algal oil comprising at least 40% DHA and mixed tocopherol were heated to a temperature in the range of 70-75°C and contacted with slurry D under stirring conditions to obtain an emulsion E having Brix in the range of 18-21.
[00068] Next, the temperature of emulsion E was adjusted to 38-42°C and contacted with masking flavor under stirring conditions to obtain a homogenized emulsion F having Brix in the range of 21-24.
[00069] Finally, the emulsion F was spray dried to obtain an agglomeration of microcapsules comprising encapsulated algal oil.
[00070] Accelerated Stability Studies for agglomeration of microcapsules comprising algal oil
[00071] Freely cold water soluble agglomeration of microcapsules comprising algal oil as prepared in Example 1 was subjected to accelerated stability studies. 50 gm of the agglomeration of microcapsules was filled in a 150 CC HDPE bottle and subjected to accelerated stability testing at a temperature of 40°C ± 2°C and at a relative humidity of 75% ± 5% (as per ICH guidelines for Zone 4B). Various test parameters were observed vis-à-vis standard for 6 months and results of the accelerated stability testing is provided in Table 2 below.
Table 2
Sr. No. Test Parameter Initial Results Standards Observations after completion of number of Months
1 2 3 5 6
1 Appearance Pass Light yellowish powder Pass Pass Pass Pass Pass
2 Odor Pass Nil Pass Pass Pass Pass Pass
3 Taste Pass Mild characteristic taste Pass Pass Pass Pass Pass
4 Texture Pass Free flowing Pass Pass Pass Pass Pass
5 Moisture 4.53% Not more than 5% 4.65% 4.02% 3.73% 3.70% 3.71%
6 DHA content 10.02% 10.0% ± 5% 10.02% 10.02% 10.00% 9.98% 9.95%
7 Emulsification test (10% solution) Pass Evenly dispersed and no surface oil droplets Pass Pass Pass Pass Pass
8 pH (10% solution) 6.20 5.0 to 6.5 6.18 6.16 6.18 6.15 6.01
9 Ash content 2.49% Not more than 5% 2.52% 2.50% 2.49% 2.50% 2.50%
10 Total Plate count Pass Not more than 1000 cfu/gm Pass Pass Pass Pass Pass

[00072] Based on the accelerated stability studies it could be observed that the composition exhibits shelf life of at least about 350 days (or 12 months).
[00073] Table 3 below provides the working range of parameters of spray dryer used to prepare a free flowing, cold water soluble formulation of Table 1.
Table 3
SL.No. Parameter(s) Set Value (& UOM) Values during Operation (±2%)
1 Inlet Temperature 190°C 191°C
2 Outlet Temperature 80°C to 90°C 87°C
3 Inlet Temperature High 220°C 219°C
4 Outlet Temperature High 100°C 101°C
5 Hot Plate Temperature 45°C 45°C
6 Feed Pump Flow rate 2ml/Minute 1.90 ml/Minute
7 Atomization Pressure 1.25 Kg/cm2 1.24 Kg/cm2
8 Aspirator 100 95
9 Stirrer/Plate Speed 30 rpm 30 rpm
10 D1 Block On 01 minute 01 minute
11 D2 Block Off 30 minutes 30 minutes
12 Cycle time 225 Minutes 225 Minutes
13 Vacuum 85-95 91
14 Cooling temperature 65°C 64°C

[00074] Variables and effect on final product
[00075] Table 4 depicts the critical process parameters which may affect the characteristic features of the formulation of Table 1.

Table 4
SL. No. Variable(s) In-Process Parameter Function Effect on the product in case of variable is out of operative limit(s)
1. pH (5% Soln. W/V) Play important role in application and stability Low pH leads to oxidation of DHA oil in the encapsulation and product will be unfit for consumption.
2. Moisture Content Important for organoleptic properties and shelf life Product(s) with higher microbial load is not safe for consumption so moisture needs to be in limit. Also Shelf life will shorten to a great extent in case higher moisture content.
3. Flavor Essential to mask fishy taste and odor to certain extent DHA oils has fishy odor and flavor fortification during Microencapsulation helps in masking it to a good extent. Deviation will lead to a product with strong fishy notes and that will limit the application of the product
4. Inlet temperature of Spray Dryer To get desired encapsulated DHA powder The process of Microencapsulation and DHA load depends on this on this step majorly. Slight deviation lead to undesired product
5. Cooling Temperature Important for the productivity of whole process Deviation from the range can lead to the formation of Coarse and clumpy material with very poor dissolution in water
6. Matrix section & Emulsion consistency Important to get DHA powder as per specification Shelf life, DHA load in the encapsulated powder and overall physical property depends on quality of emulsion and its matrix

ADVANTAGES
[00076] The present disclosure provides an agglomeration of microcapsules that exhibits minimal “fishy” odor.
[00077] The present disclosure provides an agglomeration of microcapsules that is freely soluble in cold water.
[00078] The present disclosure provides an agglomeration of microcapsules that exhibits extended shelf life without any change in its physical and chemical properties.
[00079] The present disclosure provides an agglomeration of microcapsules that exhibits better bioavailability.
[00080] The present disclosure provides an agglomeration of microcapsules that require no special storage or transport conditions.
,CLAIMS:
1. An agglomeration of microcapsules, the agglomeration comprising:
the microcapsules comprising at least one loading substance encapsulated by a shell, said shell comprising at least one milk based protein; and
alpha cyclodextrin,
wherein said loading substance weight percentage in said agglomeration is in range of 10-30%.
2. The agglomeration as claimed in claim 1, wherein said at least one loading substance is winterized algal oil comprising at least 40% DHA.
3. The agglomeration as claimed in claim 1, wherein said shell comprises: casein having weight percentage in the range of 10-20%; and soy lecithin having weight percentage in the range of 10-20%.
4. The agglomeration as claimed in any of claims 1 through 3, the agglomeration further comprising:
a. at least one stabilizer selected from the group consisting of sodium ascorbate, tri calcium phosphate, mannitol, xylitol, and combinations thereof;
b. at least one bulking agent selected from the group consisting of alpha cyclodextrin, beta cyclodextrin, gamma cyclodextrin, waxy maize starch, waxy potato starch, and combinations thereof;
c. at least one humectant selected from the group consisting of liquid glucose, corn syrup, solids, tapioca starch solids, and combinations thereof; and
d. at least one anti-oxidant selected from the group consisting of mixed tocopherol oil and natural Tocopherol oil.
5. The agglomeration as claimed in claim 4, further comprising:
a. at least one sweetener; and
b. at least one odor or taste masking agent.
6. The agglomeration as claimed in any of claims 1 through 5, wherein said agglomeration exhibits cold water solubility of at least about 10 mg/ml at a temperature of about 25°C.
7. The agglomeration as claimed in any of claims 1 through 5, wherein said agglomeration exhibits shelf life of at least about 350 days.
8. An agglomeration of microcapsules, said microcapsules comprises winterized algal oil comprising at least 40% DHA in an amount of about 20-30% by weight, said algal oil encapsulated by a shell comprises casein having weight percentage in the range of 7-17%; and soy lecithin having weight percentage in the range of 8-18%, said agglomeration further comprising alpha cyclodextrin having weight percentage in the range of 5-10.5%, ascorbate having weight percentage in the range of 0.2-0.6%, mixed tocopherol oils having weight percentage in the range of 0.1-0.3%, flavoring or masking agent having weight percentage in the range of 5-8%, sugar having weight percentage in the range of 7-11%, calcium phosphate having weight percentage in the range of 0.15-0.35%, mannitol having weight percentage in the range of 2-4%, and liquid glucose having weight percentage in the range of 15-25%.
9. The agglomeration as claimed claim 8, wherein said agglomeration exhibits cold water solubility of at least about 10 mg/ml at a temperature of about 25°C, and wherein said agglomeration exhibits shelf life of at least about 350 days.
10. A method of making the agglomeration as claimed in any of the claims 1 through 9, said method comprising:
a. contacting water at a temperature in the range of 42-48°C with soy lecithin under stirring conditions to dissolve soy lecithin in water to obtain solution A;
b. contacting liquid glucose to solution A under stirring conditions to obtain a slurry B while maintaining a temperature in the range of 42-48°C;
c. contacting ascorbate, casein, flavoring agent, sugar, tri calcium phosphate, Sodium CMC, Gums, cyclodextrin, Trehalose and mannitol to slurry B under stirring conditions while maintaining a temperature in the range of 42-48°C to obtain a slurry C having Brix in the range of 18-21;
d. adjusting the temperature of slurry C to 70-75°C under stirring conditions to obtain a slurry D having Brix in the range of 20-22;
e. contacting algal oil & mixed Tocopherol at a temperature in the range of 70-75°C to slurry D at a temperature in the range of 55-65°C under stirring conditions to obtain an emulsion E having Brix in the range of 18-21;
f. adjusting the temperature of emulsion E to 38-42°C and contacting with masking flavor under stirring conditions to obtain a homogenized emulsion F having Brix in the range of 21-24; and
g. spray drying emulsion F to obtain agglomeration comprising encapsulated algal oil.