MXPA00005831A - Matrix granule - Google Patents

Abstract

Granules that include a protein core are described. The protein core includes a protein matrix which includes a protein mixed together with a combination of a sugar or sugar alcohol and a structuring agent such as a polysaccharide or a polypeptide. The protein matrix can be layered over a seed particle or the protein granule can be homogeneous. The protein can be an enzyme or a therapeutic protein such as a hormone. Also described are methods for making the granules.

Description

MATRIX GRANUL Background of the Invention Proteins such as pharmaceutically important proteins such as industrially important protein hormones such as enzymes become more widely used. Enzymes are used in various industries that include, for example, the starch industry, the dairy industry, and the detergent industry. It is well known in the detergent industry that the use of enzymes, particularly proteolytic enzymes, has created industrial hygiene concerns for workers in detergent factories, particularly because of the health risks associated with the dustiness of available enzymes.

Since the introduction of enzymes in the detergent business, many developments in the granulation and coating of enzymes have been offered by the industry.

The patent of E.U. 4,106,991 describes an improved formulation of enzymatic granules included within the REF .: 120475 composition undergoing granulation, to finely divided cellulose fibers in an amount of 2-40% w / w based on the dry weight of the complete composition. In addition, this patent discloses that waxy substances can be used to coat the particles of the granulated substance.

US Patent 4,689,297 describes particles containing enzymes comprising a core of particles and dispersible in water having 150-2,000 microns in its longest dimension, a uniform layer of enzyme around the core particle giving a total 10% -35% by weight of the weight of the core particle, a layer of a macro-molecular, film-forming, water-soluble or dispersible coating agent uniformly surrounding the enzyme layer wherein the combination of the enzyme and the covering agent is from 25-55% of the weight of the core particle. The core material described in this patent includes clay, a sugar crystal encased in layers of corn starch which is coated with a layer of dextrin, agglomerated potato starch, particulate salt, agglomerated trisodium citrate, NaCl flakes, bread -crystallized, granules or bentonite beads converted to a solid form converted to spherical pellets, granules containing bentonite, Kaolin and earth, diatomaceous with sodium citrate crystals. The film-forming material can be an ester of fatty acids, an alkoxylated alcohol, a polyvinyl alcohol or an ethoxylated alkylphenol.

U.S. Patent 4,740,469 describes a granular enzymatic composition consisting essentially of 1-35% by weight of an enzyme and from 0.5-30% by weight of a synthetic fibrous material having an average length of 100-500 microns and a fineness in the range from 0.05- 0 7denier, with the balance being a filler or diluent. The granular composition may further comprise a fused waxy material, such as polyethylene glycol, and optionally a dye such as titanium dioxide.

The U.S. Patent 5,324,649 describes granules containing enzymes having a core, an enzymatic layer and an outer coating layer. The enzyme layer and, optionally, the core and the outer coating layer contain a vinyl polymer.

WO 91/09941 describes a preparation containing enzymes wherein at least 50% of the enzymatic activity is present in the preparation as enzymatic crystals. The preparation can be either a granulated material or a thick paste.

WO 97/12958 discloses a microgranular enzymatic composition. The granules are made by agglomeration in a fluid bed that results in granules with various carrier particles or seeds coated with an enzyme and joined together by a binder.

Two of the methods known to prepare the granulated enzymes in fluid bed coating apparatus include fluid bed agglomeration and fluid bed aerosol coating. In the fluid bed agglomeration, one or more enzymes and a binder are atomized into fine powder-bearing solids, which are constructed in size by agglomerating the carrier particles together. In these agglomerated substances, the agglutinator and the enzyme serve to transform the multiple carrier particles into granules of irregular size and shape. In the fluid bed aerosol coating, the enzyme can be arranged in layers of uniform core particles together with an optional binder.

It is desirable to produce enzymatic granules with improved stability, particularly in detergents containing bleach at high temperature and humidity. Current enzyme granules coated by fluid bed aerosol contain the enzyme in a relatively thin layer near the surface of the granule. This geometry makes the enzyme more vulnerable to depost the granule in a concentrated layer during handling and transport operations, increasing the probability and levels of airborne enzymatic aerosols in the work environment. This geometry also makes the enzyme more vulnerable to attack by penetrating moisture and inactivating substances.

However, even in light of these developments offered by the industry (as described above) there is a continuing need for enzymatic granules that produce little powder and have additional beneficial characteristics. The additional beneficial features needed in the enzyme granulation industry are formulations of low-residue granules (where few residues are defined as a reduced tendency to leave significant undissolved residues in clothing or other material), and stability improved during storage in, for example, detergent formulations containing bleach, for example, those containing peroxygen bleach such as sodium perborate or sodium carbonate. To achieve all these desired characteristics simultaneously is a particularly challenging task since, for example, many agents of prolonged release or that produce little dust such as fibrous cellulose or kaolin leave behind insoluble residues.

As such, there is a need for, for example, an enzymatic detergent granule that simultaneously does not produce dust, is stable when stored in detergents, and that is easy to manufacture in controlled distribution size. The granules of a controlled size distribution are desirable in order to confer a good circulation property to handle and mix in the detergents, and to resist segregation and settling once they are formulated in the ingredients. A controlled distribution of the particle size and a uniform shape of the particles are also important contributions to achieve a particle that produces little dust.

Therefore, it is an object of the present invention to provide enzymatic granules that produce little dust, produce few residues, highly soluble and have an increased stability particularly in detergents containing bleach. It is another object of the present invention to provide processes that allow the formation of these improved granules.

Brief description of the invention The present invention provides a granule that includes a protein core that includes a protein matrix. The protein matrix includes a protein that is mixed together with a combination of sugar or sugar alcohol and a structuring agent. Optionally, a protective layer may be layered on the protein core or the protective material may be included in the protein core. Also, optionally, a coating may be applied on the seed particle, the protein matrix and / or the protective layer. Preferably, the structuring agent is a polysaccharide or a polypeptide.

The present invention also provides a granule that includes an enzymatic core that includes an enzymatic matrix. The enzyme matrix includes an enzyme mixed together with a combination of sugar or sugar alcohol and a structuring agent. Optionally, a protective layer can be layered on the enzyme core or a protective material can be included in the enzyme core. Also, optionally, a coating may be applied on the seed particle, the enzyme matrix and / or the protective layer. Preferably, the structuring agent is a polysaccharide or a polypeptide.

The present invention further provides a granule that includes an enzyme core that includes an enzymatic matrix arranged in layers on a seed particle. The enzyme matrix includes an enzyme mixed together with a combination of sugar or sugar alcohol and a structuring agent. Optionally, a protective layer can be layered on the enzyme core or a protective material can be included in the enzyme core. Also, optionally, a coating may be applied on the seed particle, the enzyme matrix and / or the protective layer. Preferably, the structuring agent is a polysaccharide or a polypeptide.

A method for producing the above granules is also provided which includes providing a seed particle and coating the seed particle with a protein matrix comprising a protein mixed together with sugar or sugar alcohol and a structuring agent. Optionally, a protective layer can be layered on the protein core. Also, optionally, a coating may be applied on the seed particle, the protein matrix and / or the protective layer. In addition, a method for producing the above granules is also provided which includes providing a homogeneous protein matrix core comprising a protein mixed together with a sugar or a sugar alcohol and a structuring agent. Optionally, a protective layer may be layered on the protein core or a protective material may be included in the protein core. Also, optionally, a coating may be applied on the seed particle, on the protein matrix and / or the protective layer.

Detailed description of the invention One embodiment of the invention is a granule that includes a protein core that includes a protein matrix. The protein matrix includes a protein mixed together with a combination of sugar or sugar alcohol and a structuring agent. Optionally, a protective layer can be layered on an enzyme core or a protective material can be included in the enzyme core. Also, optionally, a coating may be applied on the seed particle, the enzyme matrix and / or the protective layer. Preferably, the structuring agent is a polysaccharide or a polypeptide.

Another embodiment of the invention is a granule that includes a protein core that includes a protein matrix and arranged in layers on the seed particle. The protein matrix includes a protein mixed together with a combination of sugar or sugar alcohol and a structuring agent. Optionally, a protective layer can be layered on an enzyme core or a protective material can be included in the enzyme core. Also, optionally, a coating may be applied on the seed particle, the enzyme matrix and / or the protective layer. Preferably, the structuring agent is a polysaccharide or a polypeptide.

Another embodiment of the invention is a grain that includes an enzymatic core that includes an enzymatic matrix. The enzyme matrix includes an enzyme mixed together with a combination of sugar or sugar alcohol and a structuring agent. Optionally, a protective layer can be layered on the enzyme core or a protective material can be included in the enzyme core. Also, optionally, a coating may be applied on the seed particle, the enzyme matrix and / or the protective layer. Preferably, the structuring agent is a polysaccharide or a polypeptide.

Another embodiment of the invention is a granule that includes an enzymatic core that includes an enzymatic matrix arranged in layers on the seed particle. The enzyme matrix includes an enzyme mixed together with a combination of sugar or sugar alcohol and a structuring agent. Optionally, a protective layer can be layered on the enzyme core or a protective material can be included in the enzyme core. Also, optionally, a coating may be applied on the seed particle, the enzyme matrix and / or the protective layer. Preferably, the structuring agent is a polysaccharide or a polypeptide.

A "protein core", an "enzyme core" or a "core" includes a protein matrix, for example, an enzymatic matrix in the case of an enzymatic core, the matrix can be homogeneous through the nucleus or can be layered on The seed particle There may be one or more layers between the seed particle and the matrix or the matrix and the protective layer, for example, a coating such as polyvinyl alcohol (PVA).

The seed particles are inert particles in which the enzyme matrix can be arranged in layers and can be composed of inorganic salts, sugars, sugar alcohols, small organic molecules such as acids or organic salts, minerals such as clays or silicates or a combination of two or more of these. Suitable soluble ingredients for incorporation into the seed particles include: sodium chloride, potassium chloride, ammonium sulfate, sodium sulfate, sodium sesquicarbonate, urea, citric acid, citrate, sorbitol, mannitol, oleate, sucrose, lactose and what it looks like Soluble ingredients can be combined with dispersible ingredients such as talc, kaolin or bentonite. The seed particles can be manufactured by a variety of granulation techniques including: crystallization, precipitation, pan-coating, fluid bed coating, fluid bed agglomeration, rotary atomization, extrusion, by the combination of a solid converted to spherical pellets the fluid Free, spheronization, granulation in high shear agglomeration drum. In the granules of the present invention, if a seed particle is used then the ratio of seed particles to granules is 1: 1.

The "protein matrix", "enzyme matrix" or "matrix" is a mixture of one or more proteins such as an enzyme, sugar or sugar alcohol and a structuring agent. The protein, sugar or sugar alcohol, and the structuring agent can be mixed, for example, in a solution or as a slurry. The protein can be applied from a solution or applied as a slurry of crystals or precipitated protein. The matrix of the present invention comprises between about 20-80% of the weight of the granule.

By burying the protein within the matrix, the protein can better protect itself from the twin dangers of loss of activity and wear. However, it has not previously been possible to granulate the enzymes of the sugar or sugar alcohol matrices, since the sugars and sugar alcohols exhibit "binding" characteristics, that is, they are sticky and tend to plasticize the particles together (as occurs intentionally in the case of granulation by agglomeration).

Also, to achieve a granular protein product that is produced under dust, it is necessary to control the distribution of the size and shape of the granules. Uniform and reproducible size and shape also contribute to granule stability, since particle breakage and re-agglomeration attract some protein near the granular surface.

Surprisingly, it has been found that by adding a structuring agent to the formula of the sugar matrix, the protein can be applied uniformly to individual seed particles at fast speeds without agglomeration or attrition. The particle size distribution or wear can be controlled in a precise way, based on the knowledge of the size distribution of the raw seed and the amount of solids to be added. The resulting particles are approximately spherical in shape, have a high cohesive force, are resistant to wear and penetration of moisture and inactivating substances.

Suitable sugars include sugars such as sucrose, fructose, glucose, raffinose, trehalose, lactose and maltose. Suitable sugar alcohols include sorbitol, mannitol and inositol. The amount of sugar or sugar alcohol is preferably 0.1-90% by weight of the protein matrix. The sugar or sugar alcohol of the matrix can be sugar or sugar alcohol added to the protein and can be from the fermentation culture in which the protein is present.

The structuring agent may be a polysaccharide or a polypeptide. These classes of compounds have simultaneous desirable high molecular weight properties of high water solubility. Without wishing to be bound by theory, it is believed that the high molecular weight of the structured agent contributes to two important properties which a single sugar or sugar alcohol matrix is lacking: (1) provide cohesion and strength to the particle, reducing by much the tendency of the particle to become dust; and (2) functioning as a diffusion barrier to water and small molecules by virtue of forming a network or polymer "cage" along the structure of the matrix. This greatly improves the stability of the granule.

Particular structuring agents that are chosen for polysaccharides and polypeptides typically have an anti-binding feature that is useful for reducing the binding characteristic of sugar or sugar alcohol, and allowing matrix layers to pile up, for example, in bedding. fluid - in fast speeds without agglomeration occurring.

Sugars and sugar alcohols and structuring agents have high solubility in water or dispersibility. A matrix formula can easily be prepared that includes sugars or sugar alcohols, structuring agents, and enzymes in solution or slurry with a high concentration of total solids. Concentrations of total solutions or solids in slurry of 20-50% w / w) or more can be formulated. These concentrated mixtures are highly desirable because they can be formed into granules with minimal need to evaporate water, an advantage in any granulation and drying process.

Preferred structuring agents include starch, modified starch, Irish moss, cellulose, modified cellulose, gum arabic, guar gum, acacia gum, xanthene gum, locust bean gum, cytosan, gelatin, collagen, casein, polyaspartic acid and acid. polyglutamic Preferably, the structuring agent has a low allergenicity. A combination of two or more structuring agents can be used in the granules of the present invention.

Proteins that are within the scope of the present invention include pharmaceutically important proteins such as hormones or other therapeutic proteins and industrially important proteins such as enzymes.

Any enzyme or combination of enzymes can be used in the present invention. Preferred enzymes include those enzymes capable of hydrolyzing substrates, for example, dyes. These enzymes are known as hydrolases which include, but are not limited to, proteases (bacterial, fungal, acidic, neutral or alkaline), amylases (alpha or beta), lipases, cellulases and mixtures thereof. Particularly preferred enzymes are subtilisins and cellulases. More preferred are subtilisins such as those described in US Patent 4,760,025, EP 130 756 Bl and EP Patent Application WO 91/06637, which are incorporated herein for reference, and cellulases such as Multifect L250 TM Puradax ™, commercially available from Genecor International. Other enzymes can be used in the present invention and include oxidases, transferases, dehydratases, reductases, hemicellulases and isomerases.

The matrix of the granules of the present invention may further comprise one or more synthetic polymers or other excipients as are known to those skilled in the art. Suitable synthetic polymers include polyethylene oxide, polyvinyl alcohol, polyvinyl pyrrolidone, polyethylene glycol, and polyethylene oxide-polypropylene oxide.

The matrix may further comprise plasticizers and anti-caking agents. Suitable plasticizers useful in the present invention include polyols such as glycerol, propylene glycol, polyethylene glycol (PEG), urea or other known plasticizers such as triethyl citrate, dibutyl or dimethyl phthalate or water. Suitable anti-caking agents include insoluble or barely soluble fines such as talc, TiO2, clays, amorphous silica, magnesium stearate, stearic acid and calcium carbonate.

The granules of the present invention may further comprise a protective layer, a protective layer being used to retard or prevent the diffusion of substances that may adversely affect the protein or enzyme of the matrix. The protective layer is made of a protective material and can be covered over the protein core or the protective material can be included in the protein core. Suitable protective materials include, for example, inorganic salts or acids and organic salts. The matrix without the protein can also be used as a protective layer.

The granules of the present invention may also comprise one or more coating layers. For example, these coating layers can be one or more intermediate coating layers or these coating layers can be one or more outer coating layers or a combination thereof. The coating layers can serve a variety of functions in the granular composition, depending on the final use of the enzyme granule. For example, coatings can make the enzyme resistant to oxidation by the bleach, remove the rates of solution salts at the introduction of the granule in an aqueous medium, or provide a barrier against environmental humidity in order to intensify the storage stability of the enzyme and reduce the possibility of microbial growth within the granule.

Suitable coatings include water-dispersible film-forming polymers such as polyvinyl alcohol (PVA), polyvinyl pyrrolidone (PVO), cellulose derivatives such as methyl cellulose, hydroxypropyl methyl cellulose, hydroxy cellulose, ethyl cellulose, carboxymethyl cellulose, hydroxypropyl cellulose, polyethylene glycol, polyethylene oxide, gum arabic, xanthene, Irish moss, cytosan, latex polymers, and enteric coatings. In addition, the coating agents may be used in conjunction with other active agents of the same or different categories.

Suitable PVAs for incorporation into the granule coating layer (s) include PVA partially hydrolyzed, fully hydrolyzed and moderately hydrolyzed and having high or low viscosity grades. Preferably, the outer coating layer comprises a partially hydrolyzed PVA having low viscosity. Other vinyl polymers that may be useful include polyvinyl acetate and polyvinyl pyrrolidone. Useful copolymers include, for example, PVA-methyl methacrylate copolymer and PVP-PVA copolymer.

The coating layers of the present invention may also contain one or more of the following: plasticizers, expanders, lubricants, pigments and optionally optional enzymes. Suitable plasticizers useful in the coating layers of the present invention are plasticizers including, for example, polyols such as sugars, sugar alcohol, or polyethylene glycols (PEG), urea, glycol, propylene glycol and other known plasticizers such such as triethyl citrate, dibutyl or dimethyl phthalate or water. Suitable pigments useful in the coating layers of the present invention include, but are not limited to, finely divided whiteners such as titanium dioxide or calcium carbonate or colored dyes and pigments or the combination thereof. Preferably these pigments are pigments of low residues upon dissolution. Suitable expanders include sugars such as sucrose or starch hydrolysates such as maltodextrin and solids in corn syrup, clays such as kaolin and bentonite and talc. Suitable lubricants include nonionic surfactants such as Neodol, bait alcohols, fatty acids, salts of fatty acids such as magnesium stearate and fatty acid esters.

The adjunct ingredients can be added to the enzyme granules of the present invention. The attached ingredients may include: metal salts, solubilizers; activators; antioxidants; dyes; inhibitors; agglutinators; fragrances; enzyme protecting agents / scavengers such as ammonium sulfate, ammonium citrate, urea, guanidine hydrochloride, guanidine carbonate, guanidine sultanate, thiourea dioxide, monoethanolamine, and diethanolamine, triethanolamine, amino acids such as glycine, Sodium glutamate and what it looks like, proteins such as bovine serum albumin, casein and whatever that looks like etc .; surfactants including anionic surfactants, ampholytic surfactants, non-ionic surfactants, cationic surfactants and salts of long-chain fatty acids; builders; inorganic or alkaline electrolytes; bleaching agents; indigo agents and fluorescent dyes and bleaches; and blockage inhibitors.

The granules described herein can be made by methods known to those skilled in the art of enzymatic granulation, including pan-coating, fluid bed coating, converting the solid material to spherical pellets, disc granulation, aerosol drying, extrusion. , centrifugal extrusion, spheronization, drum granulation, high shear agglomeration, or combinations of these techniques.

The following examples are representative and are not intended to be limiting. One skilled in the art can choose other enzymes, matrices, seed particles, methods and coating agents based on the teachings herein.

Examples Example 1 Atomization Coating in Fluid Bed of Alkaline Protease Laboratory 1119 grams of non-pareil particles are loaded (prepared by atomizing a colloidal mixture of sucrose and corn starch into sucrose crystals and followed by an atomization of a final coating of PVA and corn starch and then passed through a mesh screen. between 20 and 50) in a Vector FL1 fluid bed coater and then fluidized. 159 grams of an aqueous solution containing 15% w / w of Elvanol 51-05 (PVA marketed by Dow Chemical) are added to 1128 grams of an aqueous protease solution with 19.7% dry total solids and 8.4% w / w of active protease. The protease / PVA solution is atomized on the non-pareil particles under the following conditions: Fluid feed rate 18 g / min Atomization pressure 54 psi Temperature set point of 100 ° C inlet air Output air temperature range 55 to 58 ° C Input air speed 81 cyfm The coated particles are then coated with an aqueous solution containing 444 grams (40% w / w) of magnesium sulfate heptahydrate. This coating is applied under the following conditions: Fluid feed speed 23 g / min Atomization pressure 54 psi Temperature set point of 100 ° C inlet air Output air temperature range 55 to 58 ° C Inlet air speed 88 cfm Magnesium sulphate coated particles are cosmetically coated with 2356 grams of an aqueous solution that It contains 146 grams (6.2% o / p) of dioxide «Titanium, 118 grams (5% .p / p) of methyl cellulose (Methocel A -LV, Dow Chemical), 24 grams (1% w / w) of Neodol 23 / 6.5 (Shell Chemical Co.) and 39 grams (1.67% -, / p) of polyethylene glycol at a molecular weight (MW) of 600. The cosmetic coating is applied under the following conditions: Fluid feed speed 24 g / min Atomization pressure 54 psi Temperature set point of 100 ° C inlet air Output air temperature range 51 to 58 ° C Inlet air speed 88 cfm A total of 1912 grams of enzymatic granules is harvested as batch A. The overall mass balance for this experiment is 78%.

Example 2: Fluidized Bed Atomization Coating Laboratory of an Alkaline Protease Matrix / Sucrose Starch 404 grams of sieved anhydrous sodium sulfate crystals are charged between 50 and 70 mesh size in a Vector FL1 fluid bed coater and fluidized. 781 grams of an aqueous protease solution with 19.7% dry total solids and 8.4% w / w of active protease are added to 1605 grams of an aqueous solution containing 670 grams of sucrose, 186 grams of common yellow-tooth starch and 74 grams of Ethylex 2015 (AE Staley, Decatur, Illinois) that has been fully hydrated by "external cooking" at 87.78 ° C for 15 minutes. The proportion of enzymatic solids for other solids in the combined solution remains identical to that of Example 1, but the amounts are reduced to take into account an additional step in this example. The combined solution is sprayed on sodium sulphate under the following conditions: Fluid feed speed 27 g / min Atomization pressure 54 psi Setpoint of temperature 100 ° C of intake air Range of outlet air temperature 56 to 61 ° C Input air speed 80 sfm The coated particles are then coated with an aqueous solution containing 444 grams (40% w / w) of magnesium sulfate heptahydrate. This coating is applied under the following conditions: Speed of "fluid feed 27 g / min Atomizing pressure 50 psi Temperature set point 100 ° C inlet air Temperature range of outlet air 54 to 57 ° C Input air speed 79 cfm The particles coated with magnesium sulfate are cosmetically coated with 2356 grams of an aqueous solution containing 146 grams (6.2% w / w) of titanium dioxide, 118 grams (5% w / w) of methyl cellulose, 24 grams (1% w / w) of Neodol 23 / 6.5 and 39 grams (1.67% w / w) of polyethylene glycol at a MW of 600. The cosmetic coating is applied under the following conditions: Fluid feed speed 23 g / min Atomization pressure 56 psi Temperature set point 100 ° C inlet air Output temperature range 53 to 58 ° C Input air speed 83 cfm A total of 2050 grams of enzyme granules are harvested as batch. The overall mass balance for this experiment is 88.6%.

Example 3 Analysis of the Lots The granules of Examples 1 and 2 are analyzed to determine the amount of powder they generate and their stability in a stability stress analysis for three days. The methods for these procedures are as follows and the results are shown in Table 1.

Accelerated Stability Analysis The stability of various enzymatic granules that are formulated in detergents containing bleach is generally excellent, they do not really show more than about 10-20% loss in activity during six storage periods from 30 to 37 ° C and from 70% to 80 % relative humidity. However, to facilitate the development and exploration of granular formulations, it is desirable to have an accelerated medium to determine the relative stability of the granule. The conditions of accelerated stability analysis (AST) is much more severe than enzymatic granules or detergents would never find in realistic storage or transport. The AST is a "stress analysis" designed to discriminate differences between formulations that are otherwise not evident for weeks or months.

In this analysis, a detergent base to be analyzed was made from the following ingredients: 72% WFK-1 base detergent (WF, K Forschunginsti ut fuer Reinigungsteahnologie e V., Krefeld, Germany) % perborate from (Degussa Corp., Allendale Sodium Monohydrate Park, New Jersey) 3% TAED activator (Warwick International, from Mostyn bleach, United Kingdom) (= tetraacetylethylenediamine) For each sample of the enzyme to be analyzed, three identical tubes are prepared by adding 1 gram of the analysis base and 30 mg of enzymatic granules to a 15 ml conical tube and mixing by inverting the capped tube 5-8 times with the hand. Drill a hole in the tube cap with a 1/16 inch drill bit. One of the three tubes is immediately evaluated the other two are stored in a humidity chamber set at 50 ° C and at 70% relative humidity. One of the two stored tubes is evaluated after a day of storage; the second, after three days of storage. Storage stability is reported for Day 1 and Day 3 by dividing the remaining activity by the original activity on Day 0, expressed as a percentage.

Enzyme activity is determined by adding to each tube 30 ml of 0.25M MES buffer at a pH of 5.5 containing 20 μl of Catalase HP L5000 (Genencor International, Rochester, NY) and incubated for 40 minutes to inactivate perborate. After this, the enzyme is titrated by adding 10 μl of the same mixture from the test tube and 10 μl of the protease substrate sAAPF to 980 μl of 0.1M Tris pH 8.6, then incubated at 25 ° C for 3 minutes, and quantifying the Optical absorbance at 410 nm. The slope of the absorbance against time is multiplied by the dilution factor and the known extinction coefficient for the specific protease to obtain an enzymatic activity as concentration in mg / ml.

Heubach analysis of wear and dust elutriation Two methods commonly used to quantify the powder of the enzyme granule are the Heubach wear analysis and the elutriation analysis. These analyzes attempt to quantify the tendency of enzymatic granules to generate airborne protein thicknesses which can potentiate allergic reactions among workers in detergent plants. These analyzes are designed to reproduce certain typical mechanical actions of, handling, transport and combination operations used to mix the enzymatic granules in detergents on a commercial scale.

In the elutriation analysis, 60 grams of enzymatic granules are placed in a glass frit inside a glass tube that is 175 cm high and 3.54 cm in diameter, and is fluidized with a constant stream of dry air at 0.8 m / sec for 40 minutes.

In the Heubach wear analysis, 13.5 g of granules are placed in a small chamber, and cylindrical provided with a rotating blade and 4 steel balls; the granules are pushed around by the paddle and the balls, while percolating dry air through the chamber at 20 lpm for 20 minutes.

In both analyzes, the powder that leaves the particles through the air is captured in a 15 cm glass fiber filter for the subsequent quantification of the weight and determination of the activity by the sAAPF method described above. Enzyme powder for Heubach is reported as ng enzymes per gram of granule. Enzyme powder for elutriation is converted from activity to GU by 60 grams of granules, using specific enzyme conversion factors.

Table 1 Example 4 Atomization Coating in Fluid Bed at Pilot Scale of Protease Alkaline Matrix / Sucrose Starch 73.4 kg of screened sucrose crystals are loaded between 35 and 50 mesh size in a Glatt WSG-120 modified fluid bed coater and fluidized. 174.67 kg of an aqueous protease solution with 19.98% dry total solids and 6.365% w / w of active protease are added to 117 kg of an aqueous solution containing 36.25 kg of sucrose, 29 kg of common yellow-tooth starch and 7.25 kg of Ethilex 2015 that has been totally hydrated by "external cooking" at 87.78 ° C for 15 minutes. The combined solution is atomized on sucrose under the following conditions: Fluid feed rate 1.0 LPM Atomization pressure 75 psi Set point of the inlet air NA temperature Output air temperature range 70C Inlet air velocity 70 cubic meters / min The lμego coated particles are coated with an aqueous solution containing 75 kg (400.3% w / w) of magnesium sulfate heptahydrate. This coating is applied under the following conditions: Fluid feed rate 2.3 LPM Atomization pressure 50 psi Set point of the inlet air temperature NA Output air temperature range 70 ° C Inlet air velocity 70 cubic meters / min The particles coated with magnesium sulfate are cosmetically coated with 208.93 kg of an aqueous solution containing 12.97 kg (6.2% w / w) of titanium dioxide, 10.59 kg (5% w / w) of methyl cellulose, 2.12 kg (1 % w / w) of Neodol 23 / 6.5 and 3.57 kg (1.67% w / w) of polyethylene glycol at a MW of 600. The cosmetic coating is applied under the following conditions: Fluid feed rate 1.1 LPM Atomization pressure 75 psi Setpoint temperature NA inlet air Range of outlet air temperature 75 ° C Inlet air velocity 70 cubic meters / min A total of 199.35 kg is harvested of enzymatic granules as lot D. The overall mass balance for this experiment is 83.84%.

Example 5 Atomization in Fluid Bed at Pilot Scale of an Alkaline Protease Matrix / Sucrose Starch A. 65.75 kg of sieved crystals of sieve are loaded between a mesh size of 35 and 50 with a modified fluid bed coating device Glatt WSG -120 and fluidize. Add 180.42 kg of an aqueous protease solution with 20.74% dry total solids and 6.71% w / w of active protease to 145.13 kg of an aqueous solution containing 37.57 kg of sucrose, 29.94 kg of common yellow tooth starch and 7.62 kg of Ethylex 2015 that has been fully hydrated by "external cooking" at 87.78 ° C for 15 minutes. The combined solution is atomized on sucrose under the following conditions: B. The coated particles are then coated with one. aqueous solution containing 86.95 kg (40.3% w / w) of magnesium sulfate heptahydrate. This coating is applied under the following conditions: The particles coated with magnesium sulfate are cosmetically coated with 240.79 kg of an aqueous solution containing 16.97 kg (6.2% w / w) of magnesium dioxide, 6.84 kg (2.5% w / w) of methyl cellulose, 6.84 kg (2.5 % w / w) of maltodextrin M150 (DE = 15 from Grain Processing Corp., Muscatine, Iowa), 2.74 kg (1% w / w) of Neodol 23 / 6.5 and 4.57 kg (1.67% w / w) of glycol of polyethylene at a MW of 600. The cosmetic coating is applied under the following conditions: Fluid feed rate 1.2 LPM Atomizing pressure 75 psi Set point of the NA temperature inlet air Point determined for the temperature 60 ° C of the product Inlet air velocity 58 meters subie / min A total of 199.35 kg of enzymatic granules are harvested as batch E. The overall mass balance for this experiment is 97.13%.

Example 6 Atomization Coating in Fluid Bed at Pilot Scale of the Protease Alkaline Matrix / Sucrose Starch Enzymatic nuclei are made according to section A of Example 5.

In the next three granules, magnesium sulfate heptahydrate is applied as a 50% solution to make up 15% by weight of the final weight of the granule. The conditions are as follows: Atomizing pressure 50 psi Set point of the inlet air temperature NA Point determined for the temperature 47-54 ° C of the product Inlet air velocity 58 cubic meters / min The coating polymers are applied as 15% w / w of soluble solid solutions, and are batch processed in order to provide the following coating ratios, given as weight percentages of the final granules in Table 2. The conditions are as follows: follow: Atomizing pressure 50 psi Set point of the NA temperature inlet air Point determined for the product temperature 46-55 ° C Inlet air speed 58 cubic meters / min Table 2 The granules are analyzed as described in Example 3 and the results are shown in Table 3.

Table 3 Example 7 Three large-scale matrix pellets are produced in a Glatt WSG 120 fluidized bed spray apparatus. In batch J, 50.5 kg of sucrose seeds with a mesh size of -35 / + 50 are loaded in the coating apparatus and fluidizes. A carrier matrix solution is prepared by externally sewing 0.4 kg of Ethylex 2015 starch, as in the previous examples, and 46.7 kg of sucrose and 23.4 kg of yellow tooth starch are added, together with water to give a final weight of the solution of 337.4 kg. The carrier matrix solution is combined with 243.2 kg of an aqueous protease solution containing 51.89 g / L subtilisin GG36 and 19% total solids, to form the enzyme solution of the matrix. The enzymatic solution of the matrix is atomized on the sucrose seeds under the following conditions: Bed temperature 60 ° C Fluidisation air 48 scfm Speed gradient 0.3 to 1.0 lpm during 240 atomization minutes Atomization air 50-75 psig for 240 minutes A solution of ammonium sulfate is prepared by dissolving 58.3 kg of ammonium sulfate in 135.9 kg of water and this is atomized on the seeds coated with the matrix under the following conditions: Bed temperature 70 ° C Fluidizing air 48-57 scfm Speed gradient 1.5 lpm atomization Air atomization 75 psig Finally, a coating solution is prepared by dissolving or suspending 17.9 kg of polyvinyl alcohol Elvanol 51-05, 22.4 kg of titanium dioxide, and 4.5 kg of nonionic surfactant Neodol 23.5-6T in water for a net weight of 224.1 kg. This coating solution is applied under the following conditions: Bed temperature 72 ° C Fluidization air 56 scfm Speed gradient 0.5-1.2 lpm for 300 atomization minutes Atomizing air 75 psig After the coating has been completed, 255.5 kg of granules are harvested from the coating material and sieved to retain the mesh size cut -16 / + 50. The granule is titrated at 4.54% w / w of active subtilisin, and the powder and stability quantifications, reported in the following table, are carried out.

Two additional batches of matrix granules, Batches K and L, are produced in the Glatt WSG 120 modified coating apparatus under essentially the same processing conditions, but with the formulation changes noted in the following table. A pellet is produced in layers as described in Example 1.

Next, Table 4 summarizes the four formulations and reports both stability and dust for each sample.

Example 8 Atomization Coating in Fluid Bed at Pilot Scale of Amylase / Starch Matrix 26 kg of sieve crystals sieved between a mesh size of 35 and 50 are loaded in a Deseret 60 fluid bed covering apparatus. 15.3 kg of an aqueous amylase solution are added with 31% total dry solids and 12.5% w / Active amylase w to 43.5 kg of an aqueous solution containing 23.5 kg of corn starch. The combined solution is atomized on sucrose under the following conditions: The coated particles are then coated with. an aqueous solution containing 66.7 kg (40% w / w) of magnesium sulfate heptahydrate. This coating is applied under the following conditions: The particles coated with magnesium sulphate are then cosmetically coated with 92.6 kg of an aqueous solution containing 7.1 kg (6.2% w / w) of titanium dioxide, 2.9 kg (2.5% w / w) of methylcellulose, 2.9 kg (2.5 %) of Purecote B790, 1.2 kg (1.5% w / w) Neodol 23 / 6.5, and 2.0 kg (1.67% w / w) of polyethylene glycol at a MW of 600. The cosmetic coating is applied under the following conditions: Example 9 Atomization Coating in Fluid Bed at Pilot Scale of Amylase Matrix / Sucrose Starch 26 kg of sieved crystals of sucrose between a mesh size of 35 and 50 are loaded in a Deseret 60 fluid bed coater and in a fluidizer. 15.3 kg of an aqueous amylase solution with 31% dry total solids and 12.5% w / w of active amylase are added to 59.3 kg of an aqueous solution containing 7.8 kg of sucrose and 23.5 kg of corn starch. The combined solution is atomized on sucrose under the following conditions: The MgSO4 and the cosmetic coating are run exactly as described above in Example 8 Example 10 In a Glatt WSG 120 fluidized bed spray apparatus, 47.37 kg of sucrose crystals are added, sized to a mesh size of 30-50, and fluidized at 40-60 m3 / min and at 45 ° C. A suspension of amylase enzyme is prepared by making slurry 67.72 kg of corn starch common starch in 105 kg of amylose UF concentrate (LAT) with an activity of 30,000 TAU / g or 85.7 mg / g of amylase and containing 24.2 mg / ml of sugars that will be carried forward from the fermentation and recovery processes.

The enzyme suspension is coated on the sucrose seeds under the following conditions (where a range is shown, the values increase linearly over a period in slope): Once the enzyme suspension is coated on the sucrose crystals, 80 kg of a 50% solution of MgSO4 heptahydrate is atomized in the fluidized granules under the following conditions: Finally, a coating solution is prepared by adding 5.29 kg of Methocel A-15 methylcellulose (Dow Chemical), 12.71 kg of titanium dioxide (DuPont), 5.29 kg of modified starch Pure Cote B-790 (Grain Processing Corp. ), 2.12 kg of Neodol 23-6.5T (Shell) and 3.54 kg of polyethylene glycol, with molecular weight 600 (Union Carbide) to 174.91 kg of hot water and cooled to approximately 20 ° C to completely dissolve the polymers. The coating solution is applied under the following conditions: The 180 kg resulting from the coated amylase matrix granules are harvested from the coating apparatus, with an enzymatic production of 85%.

Various examples and modifications of the foregoing description of the Examples are apparent to one skilled in the art upon reading the descriptions without departing from the spirit and scope of the invention, and it is intended that all these examples or modifications be included within the scope of the drawings. appended claims. All the publications and patents referred to are incorporated in their entirety here for your reference.

It is noted that in relation to this date, the best known method for the applicant to carry out the aforementioned invention, is the conventional one for the manufacture of the objects or products to which it refers.

Having described the invention as above, property is claimed as contained in the following:

Claims (56)

Claims

1. A granule comprising a protein core comprising a protein matrix, characterized in that here, the protein matrix comprises a protein mixed together with a combination of a sugar and a structuring agent.

2. The granule according to claim 1, characterized in that the structuring agent is selected from a group consisting of a polysaccharide and a polypeptide.

3. The granule according to claim 2, characterized in that the structuring agent is selected from a group consisting of starch, modified starch, cellulose, modified cellulose, Irish moss, gum arabic, acacia gum, xanthene gum, gum of the lobster bean, and guar gum.

4. The granule according to claim 2, characterized in that the structuring agent is selected from a group consisting of chitosan, gelatin, casein, collagen, polyaspartic acid, polyglutamic acid.

5. The granule according to claim 1, characterized in that the sugar is selected from a group consisting of glucose, fructose, raffinose, maltose, lactose, trehalose and sucrose.

6. The granule according to claim 1, characterized in that it also comprises a synthetic polymer, wherein the synthetic polymer is selected from a group consisting of polyethylene oxide, polyvinyl alcohol, polyvinyl pyrrodilone, polyethylene glycol and polyethylene oxide. polypropylene oxide.

7. The granule according to claim 1, characterized in that the protein core comprises the protein matrix arranged in layers on a seed particle.

8. The granule according to claim 1, characterized in that it also comprises a coating layer.

9. The granule according to claim 8, characterized in that the coating layer is on the seed particle.

10. The granule according to claim 8, characterized in that the coating layer is on the protein matrix.

11. The granule according to claim 8, characterized in that the coating is selected from a group consisting of polyvinyl alcohol, polyvinyl pyrrodilone, cellulose derivatives such as methylcellulose, hydroxypropyl methylcellulose, hydroxycellulose, ethylcellulose, carboxymethyl cellulose, hydroxypropyl cellulose , polyethylene glycol, polyethylene oxide, cytosan, gum arabic, xanthene and moss Ireland.

12. A granule comprising a protein core comprising a protein matrix, characterized in that the protein matrix comprises a protein mixed together with a combination of sugar alcohol and a structuring agent.

13. The granule according to claim 12, characterized in that the structuring agent is selected from a group consisting of a polysaccharide and a polypeptide.

14. The granule according to claim 13, characterized in that the structuring agent is selected from a group consisting of starch, modified starch, cellulose, modified cellulose, Irish moss, gum arabic, acacia gum, xanthene gum, gum of the lobster bean, and guar gum.

15. The granule in accordance with the claim 13, characterized in that the structuring agent is selected from a group consisting of chitosan, gelatin, casein, collagen, polyaspartic acid and polyglutamic acid.

16. The granule according to claim 12, characterized in that the sugar alcohol is selected from a group consisting of mannitol, sorbitol and inositol.

17. The granule according to claim 12, characterized in that it also comprises a synthetic polymer, wherein the synthetic polymer is selected from a group consisting of polyethylene oxide, polyvinyl alcohol, polyvinyl pyrrodilone, polyethylene glycol and polyethylene oxide. polypropylene oxide.

18. The granule according to claim 12, characterized in that the protein core comprises the protein matrix arranged in layers on a seed particle.

19. The granule according to claim 12, characterized in that it also comprises a coating layer.

20. The granule according to claim 19, characterized in that the coating layer is on the seed particle.

21. The granule according to claim 19, characterized in that the coating layer is on the protein matrix.

22. The granule according to claim 19, characterized in that the coating is selected from a group consisting of polyvinyl alcohol, polyvinyl pyrrodilone, cellulose derivatives such as methylcellulose, hydroxypropyl methylcellulose, hydroxycellulose, ethylcellulose, carboxymethyl cellulose, hydroxypropyl cellulose , polyethylene glycol, polyethylene oxide, chitosan, gum arabic, xanthene and Irish moss.

23. A granule comprising an enzymatic core comprising an enzymatic matrix, characterized in that the enzyme matrix here comprises an enzyme mixed together with a combination of a sugar and a structuring agent.

24. The granule in accordance with the claim 23, wherein the structuring agent is selected from a group consisting of a polysaccharide and a polypeptide.

25. The granule in accordance with the claim 24, characterized in that the structuring agent is selected from a group consisting of starch, modified starch, cellulose, modified cellulose, Irish moss, gum arabic, acacia gum, xanthene gum, locust bean gum, and guar gum

26. The granule according to claim 24, characterized in that the structuring agent is selected from a group consisting of chitosan, gelatin, casein, collagen, polyaspartic acid and polyglutamic acid.

27. The granule according to claim 23, characterized in that the sugar is selected from a group consisting of glucose, fructose, raffinose, maltose, lactose, trehalose and sucrose.

28. The granule according to claim 23, characterized in that it also comprises a synthetic polymer, wherein the synthetic polymer is selected from a group consisting of polyethylene oxide, polyvinyl alcohol, polyvinyl pyrrodilone, polyethylene glycol and polyethylene oxide. polypropylene oxide.

29. The granule according to claim 23, characterized in that the protein core comprises an enzymatic matrix arranged in layers on a seed particle.

30. The granule according to claim 23, characterized in that it also comprises a coating layer.

31. The granule according to claim 30, characterized in that the coating layer is on the seed particle.

32. The granule according to claim 30, characterized in that the coating layer is on the enzymatic matrix.

33. The granule according to claim 30, characterized in that the coating is selected from a group consisting of polyvinyl alcohol, polyvinyl pyrrodilone, cellulose derivatives such as methylcellulose, hydroxypropyl methylcellulose, hydroxycellulose, ethylcellulose, carboxymethyl cellulose, hydroxypropyl cellulose , polyethylene glycol, polyethylene oxide, cytosan, gum arabic, xanthene and Irish moss.

34. A granule comprising an enzymatic core comprising an enzymatic matrix, characterized in that the enzyme matrix here comprises an enzyme mixed together with a combination of sugar alcohol and a structuring agent.

35. The granule in accordance with the claim 34, where the structuring agent is selected from a group consisting of a polysaccharide and a polypeptide.

36. The granule in accordance with the claim 35, characterized in that the structuring agent is selected from a group consisting of starch, modified starch, cellulose, modified cellulose, Irish moss, gum arabic, acacia gum, xanthene gum, locust bean gum, and guar gum

37. The granule according to claim 35, characterized in that the structuring agent is selected from a group consisting of chitosan, gelatin, casein, collagen, polyaspartic acid and polyglutamic acid.

38. The granule according to claim 34, characterized in that the sugar alcohol is selected from a group consisting of mannitol, sorbitol and inositol.

39. The granule according to claim 33, characterized in that it also comprises a synthetic polymer, wherein the synthetic polymer is selected from a group consisting of polyethylene oxide, polyvinyl alcohol, polyvinyl pyrrodilone, polyethylene glycol and polyethylene oxide. polypropylene oxide.

40. The granule according to claim 33, characterized in that the protein core comprises the enzymatic matrix arranged in layers on a seed particle.

41. The granule according to claim 33, characterized in that it also comprises a coating layer.

42. The granule according to claim 41, characterized in that the coating layer is on the seed particle.

43. The granule according to claim 41, characterized in that the coating layer is on the enzyme matrix.

44. The granule according to claim 41, characterized in that the coating is selected from a group consisting of polyvinyl alcohol, polyvinyl pyrrodilone, cellulose derivatives such as methylcellulose, hydroxypropyl methylcellulose, hydroxycellulose, ethylcellulose, carboxymethyl cellulose, hydroxypropyl cellulose , polyethylene glycol, polyethylene oxide, cytosan, gum arabic, xanthene and Irish moss.

45. A method for making a granule characterized in that the method comprises: a) providing a seed particle; Y . b) coating the seed particle of step a) with a protein matrix comprising a protein mixed together with a sugar or sugar alcohol and a structuring agent.

46. The method according to claim 45, characterized in that it also comprises applying a protective material.

47. The method according to claim 45, characterized in that it also comprises applying a coating layer.

48. The method according to claim 47, characterized in that the coating layer is applied on the seed particle.

49. The method according to claim 47, characterized in that the coating layer is applied on the protein matrix.

50. The method according to claim 47, characterized in that the coating is selected from a group consisting of polyvinyl alcohol, polyvinyl pyrrodilone, cellulose derivatives such as methylcellulose, hydroxypropyl methylcellulose, hydroxycellulose, ethylcellulose, carboxymethyl cellulose, hydroxypropyl cellulose , polyethylene glycol, polyethylene oxide, cytosan, gum arabic, xanthene and Irish moss.

51. A method for making a granule characterized in that the method comprises: a) providing a homogeneous protein matrix core comprising a protein mixed together with a sugar or sugar alcohol and a structuring agent.

52. The method according to claim 51, characterized in that it also comprises applying a protective material.

53. The method according to claim 51, characterized in that it also comprises applying a coating layer.

54. The method according to claim 53, characterized in that the coating layer is applied on protective material.

55. The method according to claim 53, characterized in that the coating layer is applied on the protein matrix.

56. The method according to claim 53, characterized in that the coating is selected from a group consisting of polyvinyl alcohol, polyvinyl pyrrodilone, cellulose derivatives such as methylcellulose, hydroxypropyl methylcellulose, hydroxycellulose, ethylcellulose, carboxymethyl cellulose, hydroxypropyl cellulose , polyethylene glycol, polyethylene oxide, chitosan gum arabic, xanthene and Irish moss.