Field of the Invention

The present invention is concerned with compositions comprising enzymes. More particularly, the present invention is concerned with oral care compositions containing enzymes.

Background of the Invention

Oral care compositions containing enzymes are described in EP3192496, EP3192495 and EP3192565 and EP3192564. However there remains the need to formulate compositions with enhanced enzyme activity.

The present application relates to a composition and method, for enhancing the activity of enzymes in oral care products.

Description of the Invention

The present application relates to a composition comprising a surfactant, an enzyme and silica, in which at least 50wt % of at least one enzyme is adsorbed to a silica.

The application further relates to a process for preparing a composition according to any preceding claim in which the enzyme is added to a pre-mix of silica.

Adsorption is a surface phenomenon in which particles or molecules bind to the top layer of material.

For the avoidance of doubt, any feature of one aspect of the present invention may be utilised in any other aspect of the invention. Any feature described as‘preferred’ should be understood to be particularly preferred in combination with a further preferred feature or features. Herein, any feature of a particular embodiment may be utilized in any other embodiment of the invention. All percentages are weight/weight percentages unless otherwise indicated.

Detailed Description of the Invention

The application relates to compositions comprising enzymes, preferably the compositions are personal care compositions and more preferably the compositions are oral care compositions. Preferred product forms for compositions of the invention are those which are suitable for brushing and/or rinsing the surfaces of the oral cavity.

The composition of the invention is most preferably in the form of a dentifrice. The term "dentifrice" denotes an oral composition which is used to clean the surfaces of the oral cavity. Such a composition is not intentionally swallowed for purposes of systemic administration of therapeutic agents, but is applied to the oral cavity, used to treat the oral cavity and then expectorated. Typically such a composition is used in conjunction with a cleaning implement such as a toothbrush, usually by applying it to the bristles of the toothbrush and then brushing the accessible surfaces of the oral cavity.

Preferably the dentifrice/toothpaste is in the form of an extrudable semi-solid such as a cream, paste or gel (or mixture thereof).

Compositions according to the invention comprise enzymes. Preferably the enzymes are selected from the following groups:

i) a glycoside hydrolase, more preferably an amylase, most preferably amylglucosidase;

ii) an oxoreductase acting on OH groups of donors, more preferably an oxidase with oxygen as acceptor, most preferably glucose oxidase;

iii) a hemeperoxidase more preferably a haloperoxidase most preferably a lactoperoxidase.

It is preferred if the enzyme is a mixture of 2 of the above, more preferably 3 of the above, most preferably a mixture of amylglucosidase, glucose oxidase and lactoperoxidase.

In the context of the present invention glucose oxidase either alone or part of the enzyme mix is particularly preferred.

Preferably the total level of enzyme is from 0.01 wt% to 2wt% of the total composition. More preferably from 0.05 wt% to 1 wt% of the total composition.

Preferably the total level of glycoside hydrolase in the composition is from 0.05 to 1 wt% of the total composition.

Preferably the total level of oxoreductase acting on OH groups of donors is from 0.05 to 1 wt% of the total composition. Preferably the total level of hemeperoxidase is from 0.0005 to 0.01 wt% of the total composition.

Preferably the weight ratio of glycoside hydrolase to other enzymes in the composition is greater than 1 :1 , more preferably greater than 3:2.

Compositions of the invention may also comprise other proteins such as lysozymes, lactoferrins and colostrum. Preferably these other proteins are present at total levels from 0.001 to 0.5 wt% of the total composition, preferably from 0.005 to 0.1 wt% of the total composition.

Composition according to the invention comprise a silica. The preferred silica used in the present invention is a silica with a low refractive index. It may be used as the sole abrasive silica, or in conjunction with a low level of other abrasive silicas, e.g. those according to EP 236070. The low refractive index silicas, used as abrasives in the present invention are preferably silicas with an apparent refractive index (R.l.) in the range ffornl .41 to1.47, more preferably from 1.43 to1.45, preferably having a weight mean particle size of from 5 to 15 pm, a BET (nitrogen) surface area of from 10 to 150 m ² /g, more preferably and an oil absorption of about 70 - 150 cm ³ /100 g, but abrasive silicas with a lower apparent refractive index may also be used. Typical examples of suitable low refractive index abrasive silicas (e.g. having an R.l. of between 1.435 and 1.445) are Tixosil 63 and 73 ex Rhone Poulenc; Sident 10 ex Degussa; Zeodent 113 ex Zeofinn; Zeodent 124 ex Huber, Sorbosil AC 77 ex Crosfield Chemicals (having an R.l. of approximately 1.440). The amount of these silicas in the composition generally ranges from 5 to 60% by weight of the total composition more preferably from 5 to 20% by weight.

It is preferred if the enzyme is adsorbed onto a thickening silica. Preferably the silica is has an oil absorbing level of less than 150 cm ³ /100g, more preferably less than 1 cm ³ /100g. Preferably the silica is a milled silica. Preferably the silica has a average particle size from 8 to 14 microns .more preferably from 10 to 12 microns. Particularly preferred silicas are Zeodent 165 and Tixosil 43 as silicas of this type are preferred as they enhance enzyme activity.

Preferably the ratio of total silica to total enzyme is from 150: 1 to 5: 1 more preferably from 100: 1 to 10:1 , most preferably from 60: 1 to 20: 1.

The enzyme is preferably adsorbed onto the silica. Preferably the adsorption can occur either by a main batch method in which enzyme(s) are added immediately after silica dispersion and before polymer thickener / structurants and / or flavour or by using a side pot in which silica is added to an enzyme premix solution, the premix is then added to the main batch before polymer thickener/ structurants and / or flavour.

The oral care composition will preferably contain a surfactant in an amount from 0.2 to 7% by weight of the composition. Preferred are nonionic surfactants such as polyethoxylated fatty acid sorbitan esters, ethoxylated fatty acids, esters of polyethylene glycol, ethoxylates of fatty acid

monoglycerides and diglycerides, and ethylene oxide/propylene oxide block polymers. Particularly preferred are polyethylene glycol ethers of fatty alcohols in particular polyethylene glycol ethers having from 20 to 40 ethylene oxide groups per unit. Examples of such suitable surfactant include the group of surfactants known as Steareth surfactants such as Steareth 30. Preferably the level of non-ionic surfactant is from 0.5 to 7 wt% of the total composition, more preferably from 1 to 5 wt% of the total composition.

Although other surfactants may be present it is preferred if they are limited to levels below 0.5 wt% of the total composition preferably less than 0.1 wt% of the total composition.

The oral care composition, especially in the cases of dentifrices, may also contain structurants and agents such as binders and thickening agents. These structurants will generally be present in an amount of from 0.1 to 1 % by weight based on the total weight of the composition. Suitable binders or thickening agents include carboxyvinyl polymers (such as polyacrylic acids cross-linked with polyallyl sucrose or polyallyl pentaerythritol), hydroxyethyl cellulose, hydroxypropyl cellulose, natural gums (such as carrageenan, gum karaya, guar gum, xanthan gum, gum arabic, and gum tragacanth). Natural gum based thickeners, in particular carrageenan are particularly preferred.

In preferred product forms, the amount of water and/or polyhydric alcohol will generally be at least 10 percent, preferably at least 30 percent, more preferably at least 50 percent by total weight water and/or polyhydric alcohol based on the total weight of the composition. Preferably the level of water and/or polyhydric alcohol will be less than 90 wt% of the total composition, more preferably less than 85 wt%.

A dentifrice composition according to the invention will usually contain, as the aqueous continuous phase, a mixture of water and polyhydric alcohol in various relative amounts, with the amount of water generally ranging from 10 to 50% by weight (based on the total weight of the dentifrice) and the amount of polyhydric alcohol generally ranging from 20 to 60% by weight (based on the total weight of the dentifrice). Typical polyhydric alcohols for use in the oral care composition, particularly a dentifrice composition according to the invention include humectants such as glycerol, sorbitol, polyethylene glycol, polypropylene glycol, propylene glycol, xylitol (and other edible polyhydric alcohols), hydrogenated partially hydrolysed polysaccharides and mixtures thereof. Glycerol and sorbitol are preferred, sorbitol being particularly preferred.

The oral care composition, in particular dentifrice compositions comprises abrasive materials.

Abrasive materials will generally be present in an amount of from 0.5 to 75%, preferably 3 to 60% by weight based on the total weight of the dentifrice. Suitable abrasive cleaning agents include silica xerogels, hydrogels and aerogels and precipitated particulate silicas; calcium carbonate, dicalcium phosphate, tricalcium phosphate, calcined alumina, sodium and potassium

metaphosphate, sodium and potassium pyrophosphates, sodium trimetaphosphate, sodium hexametaphosphate, particulate hydroxyapatite and mixtures thereof. Silicas are particularly preferred as they can perform a dual function an abrasive and a means of enhancing the activity of the enzyme. Preferably the composition, particularly a toothpaste, comprises a silica based abrasive as described above.

The composition, particularly if a toothpaste preferably comprises an inorganic or a natural or synthetic thickener or gelling agent in proportions of about 0.10 to about 15% by weight of the total composition, depending on the material chosen. These proportions of thickeners in the dentifrice compositions of the present invention form an extrudable, shape-retaining product which can be squeezed from a tube onto a toothbrush and will not fall between the bristles of the brush but rather, will substantially maintain its shape thereon. Suitable thickeners or gelling agents useful in the practice of the present invention include finely divided silicas, hectorites, calcium carbonate, colloidal magnesium aluminium silicates and mixtures thereof. Preferred are inorganic thickening silicas such as amorphous silicas available from Huber Corporation under the trade designation Zeodent 165, and/or gums such as Irish moss, iota-carrageenan, gum tragacanth, and polyvinylpyrrolidone.

Compositions of the invention may comprise a metal ion such as tin or zinc. Preferably the sources of zinc ions are zinc chloride, zinc acetate, zinc gluconate, zinc sulphate, zinc fluoride, zinc citrate, zinc lactate, zinc oxide, zinc monoglycerolate, zinc tartrate, zinc pyrophosphate zinc maleate and mixtures thereof.

Compositions of the invention may comprise fluoride sources such as sodium fluoride, stannous fluoride, sodium monofluorophosphate, zinc ammonium fluoride, tin ammonium fluoride, calcium fluoride, cobalt ammonium fluoride and mixtures thereof. Compositions of the present invention may also contain further optional ingredients customary in the art such as antimicrobial agents, e.g. chlorhexidine, sanguinarine extract, metronidazole, quaternary ammonium compounds, such as cetylpyridinium chloride; bis-guanides, such as chlorhexidine digluconate, hexetidine, octenidine, alexidine; and halogenated bisphenolic compounds, such as 2,2' methylenebis-(4-chloro-6-bromophenol);

anti-inflammatory agents such as ibuprofen, flurbiprofen, aspirin, indomethacin etc.;

anti-caries agents such as sodium- and stannous fluoride, aminefluorides, sodium

monofluorophosphate, sodium trimeta phosphate and casein;

plaque buffers such as urea, calcium lactate, calcium glycerophosphate and strontium

polyacrylates;

vitamins such as Vitamins A, C and E;

plant extracts;

plant-derivable antioxidants such as flavonoid, catechin, polyphenol, and tannin compounds and mixtures thereof;

desensitising agents, e.g. potassium citrate, potassium chloride, potassium tartrate, potassium bicarbonate, potassium oxalate, potassium nitrate and strontium salts;

anti-calculus agents, e.g. alkali-metal pyrophosphates, hypophosphite-containing polymers, organic phosphonates and phosphocitrates etc.;

biomolecules, e.g. bacteriocins, antibodies, enzymes, etc.;

flavours, e.g. peppermint and spearmint oils;

proteinaceous materials such as collagen;

preservatives;

opacifying agents;

amino acids such as arginine

colouring agents;

pH-adjusting agents;

sweetening agents;

pharmaceutically acceptable carriers, e.g. starch, sucrose, water or water/alcohol systems etc.; surfactants, such as anionic, nonionic, cationic and zwitterionic or amphoteric surfactants;

humectants such as glycerol, sorbitol, propyleneglycol, xylitol, lactitol etc.;

binders and thickeners such (Natrosol®), xanthan gum, gum arabic etc. as well as synthetic polymers such as polyacrylates and carboxyvinyl polymers such as Carbopol®;

polymeric compounds which can enhance the delivery of active ingredients such as antimicrobial agents can also be included;

buffers and salts to buffer the pH and ionic strength of the oral care composition; and other optional ingredients that may be included are e.g. bleaching agents such as peroxy compounds e.g. potassium peroxydiphosphate, effervescing systems such as sodium bicarbonate/citric acid systems, colour change systems, and so on.

The invention will now be illustrated by the following non-limiting Examples:

Examples

The composition of the glucose oxidase premix, is as in Table 1 :

Table 1

The samples were prepared following the amounts in Table 2. Powders were dispersed using a magnetic stirrer for 30 minutes at room temperature. Zeta potential was measured using dynamic light scattering using a Malvern ZetaSizer NanoSeries with a universal dip cell and a backscatter angle of 173 degrees. After diluting the samples using water to minimise turbidity. All measurements were repeated three times. The zeta potential results demonstrate a significant adsorption of enzyme, Glucose oxidase (GOx) onto silicas indicated by the large drop in surface charge of the particles.

Table 2

Glucose Oxidase was incubated at 37°C with individual formula ingredients in 0.05 mM citrate- phosphate buffer, pH 7. After 10 mins Glucose oxidase activity was measured using Standard Kits and Reagents

• Amplex® Red Glucose/Glucose Oxidase Assay Kit - ex Invitrogen - Catalog no. A22189 · Amplex® UltraRed Reagent - ex Invitrogen - Catalog no. A36006]

Table 3

j j j

I

j !

Table 3 demonstrates when enzyme is adsorbed onto silica the activity increases. Zeodent 113 gives enhanced enzyme activity compared with Tixosil 43

An Example according to the invention is as follows:

Table 4

(composition contains 1450ppm F as NaF) * Precipitated silica

** Thickening silica

In this Example the glucose oxidase is added to the thickening silica prior to addition to the toothpaste mix.