CAPSULES

Abstract

A capsule having an encapsulated material and a capsule wall encapsulating the encapsulated material. The capsule wall includes a N-acetylglucosamine/glucosamine copolymer or a derivative of such a copolymer, wherein the N-acetylglucosamine/glucosamine copolymer or a derivative of such a copolymer is derived from a raw material which has a non-animal origin. The capsules have significant compression resistance while minimizing the amount of polymer incorporated into the capsule wall and are advantageously stable in a range of products including when associated with commercially available protease containing, biological liquid laundry products.

Claims

1-32. (canceled)

33. A capsule comprising an encapsulated material and a capsule wall encapsulating the encapsulated material, wherein the capsule wall includes a N-acetylglucosamine/glucosamine copolymer or a derivative of such a copolymer, wherein the N-acetylglucosamine/glucosamine copolymer or a derivative of such a copolymer is derived from a raw material which has a non-animal origin.

34. The capsule according to claim 33, wherein the N-acetylglucosamine/glucosamine copolymer or a derivative of such a copolymer is derived from mycelium of a fungus; wherein in said capsule, the ratio of the weight of encapsulated material divided by the weight of the capsule wall is at least 8 and the ratio is less than 25; wherein said capsule wall includes: (a) N-acetylglucosamine/glucosamine copolymer or a derivative of such a copolymer; (b) a component (A) or a residue of component (A) after reaction or interaction with material referred to in (a) wherein component (A) is a water-soluble polymer; and (c) a cross-linking moiety which cross-links components in the wall; wherein the ratio of the wt % of components in (a) and (b) is at least 3 and is less than 10.

35. A method of making a capsule according to claim 33, the method comprising an encapsulated material and a capsule wall encapsulating the encapsulated material, the method comprising: (i) selecting a N-acetylglucosamine/glucosamine copolymer or a derivative of such a copolymer, wherein said N-acetylglucosamine/glucosamine copolymer or a derivative of such a copolymer is derived from a micro-organism; (ii) selecting a material to be encapsulated; (iii) subjecting the N-acetylglucosamine/glucosamine copolymer or a derivative of such a copolymer and the material to be encapsulated to conditions thereby causing the N-acetylglucosamine/glucosamine copolymer or a derivative of such a copolymer to be incorporated into a capsule wall which surrounds the material to be encapsulated, thereby defining said capsule.

36. The method according to claim 35, wherein said N-acetylglucosamine/glucosamine copolymer or a derivative of such a copolymer selected in step (i) has an average molecular weight of at least 10 kDa and less than 300 kDa.

37. The method according to claim 35, wherein said N-acetylglucosamine/glucosamine copolymer or a derivative of such a copolymer has a degree of acetylation ranging from 5 to 30 mol %.

38. The method according to claim 35, wherein said N-acetylglucosamine/glucosamine copolymer or a derivative of such a copolymer has a viscosity of 1 to 40 mPa.Math.s in 1 wt % in acetic acid solution.

39. The method according to claim 35, wherein a component (A) and said N-acetylglucosamine/glucosamine copolymer or a derivative of such a copolymer define a coacervate phase arranged to encapsulate the material to be encapsulated in the method, wherein said component (A) is polar and is a polymer.

40. The method according to claim 39, wherein said component (A) is a cellulose or cellulose derivative, or a gum.

41. The method according to claim 39, wherein said component (A) is a hydrophilic polymer; and said material to be encapsulated is hydrophobic.

42. The method according to claim 35, wherein said material to be encapsulated is an oil, a butter, or a wax.

43. The method according to claim 39, wherein the method comprises providing said N-acetylglucosamine/glucosamine copolymer or a derivative of such a copolymer in an aqueous solution (A); providing component (A) in solution in said aqueous solution (A); wherein the ratio of the wt % of said component (A) divided by the wt % of said N-acetylglucosamine/glucosamine copolymer or a derivative of such a copolymer is at least 3; and is less than 10; and wherein the total wt % of dissolved solids in said solution (A) is at least 5 wt %.

44. The method according to claim 43, wherein said solution (A) has a pH of greater than 1 or is adjusted to a pH of greater than 1; wherein the method comprises producing a formulation (B) which comprises aqueous solution (A) and said material to be encapsulated, wherein formulation (B) is subjected to conditions whereby complex coacervation of the N-acetylglucosamine/glucosamine copolymer or a derivative of such a copolymer and component (A) occurs, wherein said conditions comprise increasing the pH of formulation (B).

45. The method according to claim 44, wherein a base is contacted with formulation (B) to increase the pH, wherein the pH is greater than 2 and less than 4 after contact with said base.

46. The method according to any of claim 44, wherein when the coacervate phase is formed, the temperature of formulation (B) does not increase to greater than 30 C.; and/or formulation (B) is not actively heated.

47. The method according to claim 35, wherein after formation of a coacervate phase around said material to be encapsulated, the combination is treated with a component (B) which is arranged to effect cross-linking between components in the wall.

48. The method according to claim 47, wherein component (B) is arranged to react with hydroxy moieties in said capsule wall to effect cross-linking.

49. A formulation comprising: (i) capsules according to claim 33; and (ii) a protease enzyme or an alcohol.

50. The method according to claim 35, wherein said N-acetylglucosamine/glucosamine copolymer or a derivative of such a copolymer selected in step (i) includes a moiety of structure ##STR00003##

51. The method according to claim 35, wherein: said N-acetylglucosamine/glucosamine copolymer or a derivative of such a copolymer selected in step (i) has an average molecular weight of at least 10 kDa and less than 300 kDa; said N-acetylglucosamine/glucosamine copolymer or a derivative of such a copolymer has a degree of acetylation in the range 5 to 30 mol %; a component (A) and said N-acetylglucosamine/glucosamine copolymer or a derivative of such a copolymer define a coacervate phase which is arranged to encapsulate the material to be encapsulated in the method, wherein said component (A) is a cellulose or cellulose derivative, or a gum; said material to be encapsulated is an oil, a butter or a wax; said N-acetylglucosamine/glucosamine copolymer or a derivative of such a copolymer selected in step (i) includes a moiety of structure ##STR00004##

52. The method according to claim 35, wherein: said N-acetylglucosamine/glucosamine copolymer or a derivative of such a copolymer selected in step (i) has an average molecular weight of at least 60 kDa and less than 150 kDa; said N-acetylglucosamine/glucosamine copolymer or a derivative of such a copolymer has a degree of acetylation in the range 5 to 30 mol %; said component (A) is a gum; said material to be encapsulated is an oil, a butter or a wax; the method comprises providing said N-acetylglucosamine/glucosamine copolymer or a derivative of such a copolymer in an aqueous solution (A); providing component (A) in solution in said aqueous solution (A); wherein the ratio of the wt % of said component (A) divided by the wt % of said N-acetylglucosamine/glucosamine copolymer or a derivative of such a copolymer is at least 3 and is less than 10; and wherein the total wt % of dissolved solids in said solution (A) is at least 5 wt %; said solution (A) has a pH of greater than 1 or is adjusted to a pH of greater than 1; wherein the method comprises producing a formulation (B) which comprises aqueous solution (A) and said material to be encapsulated, wherein formulation (B) is subjected to conditions whereby complex coacervation of the N-acetylglucosamine/glucosamine copolymer or a derivative of such a copolymer and component (A) occurs, wherein said conditions comprise increasing the pH of formulation (B); wherein a base is contacted with formulation (B) to increase the pH, wherein the pH is greater than 2 and less than 4 after contact with said base.

Description

[0080] Specific embodiments of the invention will now be described by way of example, with reference to the accompanying drawings, in which:

[0081] FIG. 1 is a graph comparing average break strengths of capsules made using different sources of chitosan;

[0082] FIG. 2 is a graph comparing compression before breakage of capsules made using different sources of chitosan;

[0083] FIG. 3 is a graph comparing probe distance travelled before failure of capsules made using different sources of chitosan; and

[0084] FIG. 4 is a graph comparing compressibility of capsules made using different sources of chitosan;

[0085] The following materials are referred to hereinafter:

[0086] Copolymer (I)biomass derived N-acetylglucosamine/glucosamine copolymer (commonly referred to as chitosan) having a molecular weight of 80 kDa obtained from Kitozyme.

[0087] Copolymer (II)biomass derived N-acetylglucosamine/glucosamine copolymer having a molecular weight of 10-20 kDa obtained by from Kitozyme.

[0088] Marine Chitosan (HCMF)obtained from Chitinor, Norway and having a molecular weight of 50-100 kDa.

[0089] Gum Arabic (Instant Gum) obtained from Nexrra, France.

[0090] Other reagents obtained from Sigma Aldrich.

[0091] Chitosan, or deacetylated chitin, is a linear copolymer comprised of randomly repeating glucosamine and N-acetylglucosamine units connected by .fwdarw.(1,4) type linkages. The chemical structure is as shown below:

##STR00002##

[0092] The N-acetylglucosamine/glucosamine copolymer is a positively charged polyelectrolyte and will undergo complex coacervation with negatively charged polyelectrolytes such as Gum Arabic. However, N-acetylglucosamine/glucosamine copolymer is not soluble above pH 5 (depending on degree of deacetylation) and therefore the controlled deposition of a coacervate cannot be achieved by adjustment of pH from neutral to acidic to allow precipitation to occur. Instead, a different approach can be utilised whereby coacervate is formed by controlling the charge on Gum Acabic Below pH 2.0, the ionisation of the carboxyl groups is minimal and an interaction with N-acetylglucosamine/glucosamine copolymer is not observed. By increasing the pH gradually, the anionic moieties on Gum Arabic become ionised which gives rise to interaction between N-acetylglucosamine/glucosamine copolymer and Gum Arabic and the coacervate can be seen as a concentrated liquid droplets, which can then be used to coat oil droplets. More particularly, a matrix encapsulation technique can be employed to create beads of N-acetylglucosamine/glucosamine copolymer encasing an active material. Material to be encapsulated is mixed into a solution of N-acetylglucosamine/glucosamine copolymer. Then, via a dropping mechanism, an alkali solution is dripped into the mixture which causes N-acetylglucosamine/glucosamine copolymer precipitation. The resultant beads which form entrap the active material in a spherical matrix, which can be recovered and maturated.

[0093] Microcapsules formed via complex coacervation will remain mechanically fragile or will maintain the potential to be re-solubilised by warm water or pH changes unless cross-linked. This crosslinking is achieved by application of a cross linking agent, which is typically a solution of glutaraldehyde or formaldehyde, although genepin, oleuropein, epichlorhydrin, sodium tripolyphosphate and transglutaminase have also been used.

[0094] In the following, Examples 1 and 2 describe the preparation of microcapsules using biomass derived N-acetylglucosamine/glucosamine (i.e. chitosan) and, subsequently, results of mechanical tests are provided.

EXAMPLE 1PREPARATION OF MICROCAPSULES MADE USING HIGH MOLECULAR WEIGHT COPOLYMER (I)

[0095] The internal oil phase component was:

[0096] High oleic sunflower oil (80 wt %)

[0097] -carotene suspension (30 wt %) in vegetable oil (20 wt %).

[0098] The external water phase components comprised the following:

[0099] 4 wt % solution (in deionised water) of Copolymer (I) (28.85 wt %).

[0100] 16 wt % solution (in deionised water) of gum arabic solution (48.08 wt %).

[0101] Deionised water (23.07 wt %).

[0102] The external water phase components were combined (130 g) and mixed with an overhead stirrer (at 320 rpm), using a 4 blade star propeller in a beaker (400 ml), and adjusted to pH 1.7 with hydrochloric acid (25%, 2.3 g).

[0103] The internal oil phase components (100 ml) were mixed on a stirrer plate until homogenous. The internal oil phase was then added to the external water phase and the stirrer speed was increased to 1400 rpm for 120 seconds, yielding oil droplets with a modal average particle size of 50 microns. The stirrer speed was then reduced to 450 rpm.

[0104] In a separate beaker, 1 litre deionised water (480 ml) was adjusted to pH 1.7 with hydrochloric acid (25%). The emulsion comprising oil droplets was added to this beaker and stirring continued at 500 rpm.

[0105] Coacervation formation was as follows: A dropping funnel (50 ml) was charged with 50 mL of triethanolamine solution (5% wt in Deionised Water). The pH was monitored as the funnel was set to release approximately 0.5 ml/min. The emulsion was regularly checked under the microscope for wall formation and quality.

[0106] The flow of triethanolamine solution was stopped when the pH reached 2.78.

[0107] Glutaraldehyde solution (4 g, 50%) was then added to effect cross-linking. The solution was left to stir overnight.

[0108] The capsule suspension was transferred to another 2 litres beaker and diluted with deionised water. When the capsules had settled to the top, the suspension was filtered over 60 micron mesh fabric and washed with 4 litres of deionised water. The dry capsule slurry was then transferred to a beaker (400 ml) and weighed, yielding 125.68 g of approximately 85% solids. Preservative solution (87.97 g) was added, comprised of deionised water (98.5%), carboxy methylcellulose (1%) and potassium sorbate (0.5%), adjusted to pH 4.8 with citric acid solution (10%).

EXAMPLE 2PREPARATION OF MICROCAPSULES USING LOWER MOLECULAR WEIGHT COPOLYMER (II)

[0109] The internal oil phase component used was:

[0110] High oleic sunflower oil (80 wt %)

[0111] -carotene suspension (30 wt %) in vegetable oil (20 wt %).

[0112] The external water phase component comprised the following:

[0113] 4 wt % solution (in deionised water) of Copolymer II (28.85 wt %)

[0114] 16 wt % solution (in deionised water) of gum arabic solution (48.08 wt %)

[0115] Deionised water (23.07 wt %)

[0116] The external water phase components were combined (130 g) and mixed with an overhead stirrer (at 320 rpm), using a 4 blade star propeller in a beaker (400 ml), and adjusted to pH 2.21 with hydrochloric acid (25%, 1.4 g).

[0117] The oil phase (100 ml) was then added to the external water phase and the stirrer speed was increased to 1200 rpm for 180 seconds, yielding an emulsion comprising oil droplets with a modal average particle size of 60 microns. The stirrer speed was then reduced to 400 rpm.

[0118] In a separate 1 litre beaker, deionised water (428 ml) was adjusted to pH 1.7 with hydrochloric acid (25%). The emulsion comprising oil droplets was added to this beaker and stirring continued at 500 rpm.

[0119] Coacervation formation was as follows:

[0120] A dropping funnel (50 ml) was charged with 50 mL of triethanolamine solution (5% wt in deionised water). The pH was monitored as the funnel was set to release approximately 0.33 ml/min. The emulsion was regularly checked under the microscope for wall formation and quality.

[0121] The flow of triethanolamine solution was stopped when the pH reached 3.17.

[0122] Glutaraldehyde solution (3 g, 50%) was then added to effect cross-linking. The solution was left to stir overnight.

[0123] The following test was used to assess the microcapsules.

Test IMicrocapsule Mechanical Strength Determination

[0124] One capsule was isolated and centred underneath a probe (set at 5 mm height from plate) of a Stable Microsystem Texture Analyzer. The analyser was used to assess the degree of force required to break a single microcapsule.

[0125] After initiation of a test, the first drop in resistance to applied force was recorded. The test parameters (on Stable Micro Systems TA.XT plus Texture Analyser) were as follows:

[0126] Start height: 5 mm

[0127] Pre-test speed: 0.5 mm sec.sup.1

[0128] Test speed: 0.2 mm sec.sup.1

[0129] Trigger Force: 0.05 g

[0130] Maximum force: 100 g

[0131] Post-test speed: 10 mm sec.sup.1

[0132] Test sequence: return to start

[0133] Probe: Perspex cylinder

[0134] After a test, the probe and surrounding area were wiped down with a small amount of ethanol and a test repeated.

Results

[0135] In tests, capsules of Example 1 were found to have significantly higher break strength compared to capsules of Example 2.

EXAMPLES 3 AND 4COMPARISON OF CAPSULES PREPARED USING BIOMASS DERIVED AND MARINE DERIVED CHITOSAN

[0136] Two batches of capsules based on a combination of Gum Arabic with different sources of chitosan were prepared and assessed.

[0137] The internal phase of the capsules consisted of the following:

TABLE-US-00001 Constituent Amount (wt %) High oleic sunflower oil 97.78 Marula oil 2 Lutein suspension 0.2 Green 6 0.02

[0138] The Lutein and Green 6 colourants were included to make particle size determination easier.

[0139] The external phase of the capsules consisted of the following:

TABLE-US-00002 Example Chitosan Instagum Deionised No. Chitosan Type amount (g) AA water (g) 3 HCMF 1.5 10 410 4 Copolymer (I) 1.5 10 410

[0140] For both Examples 3 and 4, the deionised water was lowered to pH 1.95 (21.5 C.) and the chitosan was added. When it had fully dissolved, the pH and viscosity were measured. The Gum Arabic was then added and stirred until dissolved, and hydrochloric acid (25% wt.) was used to lower the pH of the systems to 1.9, and the viscosity was again measured. Results are provided in the table below.

TABLE-US-00003 Example 3 Example 4 Time taken for full chitosan solvation 9 minutes 18 minutes pH after chitosan solvation 2.16 (22.2 C.) 2.20 (21.9 C.) Viscosity* (cP) after chitosan solvation 32.4 (21.0 C.) 22.6 (21.0 C.) Time taken for full gum arabic solvation 11 minutes 7 minutes pH after gum arabic solvation 2.47 (22.5 C.) 2.52 (22.1 C.) Viscosity* (cP) after gum arabic 30.4 (21.0 C.) 22.6 (21.0 C.) solvation HCl (25% wt.) (g) to lower system to 1.67 1.76 starting pH Starting pH 1.88 (22.5 C.) 1.87 (22.3 C.) *All measurement taken on Brookfield Viscometer, spindle 02, speed 100 rpm

[0141] The Example 3 formulation was clear, whereas the Example 4 formulation was a golden colour. The HCMF was faster to dissolve and had a higher viscosity than the Copolymer (I), though the Gum Arabic then took longer to dissolve in HCMF. For Example 4, approximately 5% more HCl (25% wt.) was required to achieve the appropriate pH.

[0142] The Internal Phases were then added, by being carefully dropped into the aqueous phase's stirring vortex, to prevent oil slick formation. The batches were stirred at 295 rpm.

[0143] Two dropping funnels were charged with triethanolamine (TEA) solution (5%), and set to drip around 1 ml/min. The TEA was then delivered. It was noted that the pH of the batches responded in lockstep with each other, proportionate to how much TEA solution had been added. The response of the encapsulations to TEA addition was noted and photomicrographs taken to record the progress.

[0144] After 20 ml of TEA addition, the two batches were around pH 2. No coacervate had yet formed in either batch. Local coacervate formation was macroscopically visible when TEA was added, at pH 2.18 for HCMF and Copolymer (I) at pH 2.28. However this did not translate to microscopically visible coacervate.

[0145] The first visible, lasting coacervate was for HCMF at pH 2.55, and Copolymer (I) at pH 2.56, though neither batch had yet formed walls. The Copolymer (I) batch had a finer, less visible coacervate at this point; the HCMF batch had more visible, larger coacervate.

[0146] At 78 ml of TEA addition, the pH was 2.63 for HCMF and 2.65 for Copolymer (I). Both batches had started forming walls around the smaller capsules. The Copolymer (I) coacervate was more discreet and the walls formed were clearer and more uniform.

[0147] Wall formation on all capsules began at pH 2.68 for HCMF, with thinner walls round the larger oil droplets. The coacervate appeared to drop in quality at this point, becoming less discrete droplets and more amorphous material from pH 2.68 up to the finishing pH of 2.85.

[0148] Walls formed more consistently around the various capsule sizes present in the Copolymer (I) batch. The coacervate remained recognisable as discrete droplets until a higher pH than HCMF, around pH 2.8.

[0149] At the final pH of 2.85, there were several differences between the batches. The Copolymer (I) batch was still producing coacervate locally upon addition of TEA solution. The walls were smoother and more consistent and optically clearer, and did not incorporate smaller oil droplets into the walls as the HCMF did.

[0150] HCMF stopped producing locally visible coacervate at pH 2.75. The walls appeared thicker than the Copolymer (I) capsules and less optically clear. The coacervate was more amorphous.

[0151] Both batches were then cross-linked with 6 g glutaraldehyde and stirred overnight. Both batches of capsules appeared stable and of good quality. The copolymer (I) batch had clearer walls and a lack of oil droplets incorporated into the walls; compared to the HCMF-based batch.

[0152] Yields were roughly the same: the filtered weight of the capsules was 121 g for HCMF and 119 g for Copolymer (I).

[0153] Each batch was split, with some half preserved in 0.5% xanthan, 0.5% rokonsal solution, and a quarter each in 0.5% rokonsal and either 2.5% CMC or 0.4% Gellan. The capsules were stable in Xanthan and Gellan.

[0154] The capsules were assessed and results obtained as described further below.

Test (A)Particle Size Comparison

[0155] Samples of xanthan preserved batches were taken and micrographs acquired. For each batch, five photographs and a total of 50 capsules were sized and the following results obtained.

TABLE-US-00004 Capsules formed Average Particle from components of particle size Example No. size (m) range (m) 3 555 206 to 866 4 440 139 to 787

EXAMPLES 5 TO 9

[0156] The procedure described for Examples 3 and 4 was used to produce a range of particles for comparison as detailed in the table below.

TABLE-US-00005 Example No. Summary 5 Capsules formed from Copolymer I with modal average particle size of 1000 m 6 Capsules made from Marine Chitosan HCMF with modal average particle sizes of 500 m 7 Capsules formed from Copolymer I with modal average particle size of 500 m 8 Capsules made from Marine Chitosan HCMF with modal average particle sizes of 250 m 9 Capsules formed from Copolymer I with modal average particle size of 250 m

[0157] The capsules described in Examples 5 to 9 were further assessed for break strength, compression before breakage, total distance travelled by the probe used to apply the force to the capsules and compressibility (%). Results are presented in FIGS. 1 to 4. It is found that, in general, capsules based on Copolymer I have higher break strength and are generally more flexible or compressible compared to comparable capsules derived from marine chitosan.

[0158] As an alternative to the gum arabic, the following materials may be used:

[0159] Sodium carboxymethylcellulose or other cellulose derivatives, sodium polyacrylate, polyacrylic or methacrylic acid, sodium tripolyphosphate, albumen, alginates such as sodium alginate, alginic acid, polyphosphates, polyvinyl acetate, polyvinyl alcohol, carrageenan, casein, calcium caesinate, agar-agar, starch, pectins, Irish moss and xanthan gum.

[0160] The capsules prepared as described have been proven to offer reasonable stability (ie capsule walls remain intact and the capsules contents remain within the capsule) in commonly used personal care products, such as moisturising creams, shower gel, hand wash, shampoo and hydro-alcoholic formulations; in homecare applications, such as hand dish liquid, biological laundry detergent; in beverage formulations including dairy based beverages, like milkshakes and soft drinks, both still and carbonated.

[0161] Additionally, the capsules retain a relevant encapsulated material when associated with commercially available protease containing, biological liquid laundry products. Other prior art capsules were found to prematurely degrade, releasing the capsule content. Thus, the capsules can be advantageously used in protease enzyme-containing environments.

[0162] The invention is not restricted to the details of the foregoing embodiment(s). The invention extends to any novel one, or any novel combination, of the features disclosed in this specification (including any accompanying claims, abstract and drawings), or to any novel one, or any novel combination, of the steps of any method or process so disclosed.