Topical biopolymers

Abstract

Provided herein are biopolymers comprising repeating polysaccharide units, preparations of the biopolymers, and topical compositions comprising the biopolymers, as well as methods of use.

Description

[Technical field]

[0001] [CROSS REFERENCE TO RELATED APPLICATIONS] This application claims the benefit of priority to U.S. Provisional Application No. 63 / 158,268, filed March 8, 2021, and U.S. Provisional Application No. 63 / 251,205, filed October 1, 2021, each of which is incorporated by reference in its entirety for all purposes.

[0002] Provided herein are biopolymers comprising repeating polysaccharide units, preparations of the biopolymers, and topical compositions comprising the biopolymers, as well as methods of use. [Background technology]

[0003] Polymers that bind and retain water are sought after in personal care and cosmetic products for their anti-aging and anti-wrinkle effects. Polymers with these properties increase epidermal hydration when applied to the surface of the skin. Polymers with sufficiently low molecular weight can pass through the outer layer of the skin epidermis. The ability to bind and retain water is important because increased water retention and moisture content effectively reduces the appearance of wrinkles and results in smoother, more supple skin.

[0004] The most commonly used polymer in anti-aging formulations, especially in premium personal care products, is hyaluronic acid (HA). HA is a high molecular weight (HMW) polymer found naturally in animal skin, connective tissue, and synovial fluid, where it functions by retaining water and providing lubrication. HA is widely used as a dietary supplement, injectable for joint lubrication, surgical aid for adhesion prevention, dermal filler, skin surface moisturizer, and in personal care and cosmetics due to the aforementioned skin penetration and water binding properties of LMW products. HA is typically of animal origin, traditionally derived from rooster combs, and also as a by-product from porcine or bovine processing facilities. HA can also be produced by fermentation of Streptococcus zooepidemicus, or related species, a pathogenic bacterium that naturally incorporates HA into its cell capsule.

[0005] The production costs of both animal- and fermentation-derived HA are high. In the case of animals, the cost is driven by the relatively low yield of HA per amount of tissue and the several downstream processing steps required to produce a purified product. In the case of S. zooepidemicus HA, large-scale fermentation must be carried out under strictly regulated conditions because the production organism is an infectious agent. In addition, downstream processing steps may be required to reduce any potential toxins produced by the bacteria that may make their way into the final product. Finally, both products must be chemically or enzymatically processed to produce HA of a molecular weight that allows skin penetration.

[0006] Despite cost and manufacturing limitations, the HA market continues to grow due to the strong anti-wrinkle and anti-aging performance of HA in cosmetics. Currently, there are few competing technologies that provide the same or better functionality as HA. Alternative products that show improved water binding, can remain on the surface or penetrate the skin, and can be produced at substantially lower cost would allow for significant market growth and expansion. There is a need and shortage of water-binding carbohydrate-based polymers to address a wide range of needs in the personal care and cosmetics industries. Summary of the Invention

[0007] The present disclosure provides naturally produced non-animal derived carbohydrate biopolymers with improved water binding capacity compared to known agents such as HA.As shown in the examples presented herein, the biopolymers of the present disclosure show significantly improved water binding capacity compared to HA.The biopolymers of the present disclosure are particularly suitable for personal care and cosmetic applications and lack cytotoxicity in standard tests.In addition, the biopolymers of the present disclosure show improved skin hydration performance when used as a functional ingredient in lotion bases.

[0008] In some embodiments, the biopolymers of the present disclosure exhibit transdermal permeability. In some embodiments, biopolymers capable of penetrating the skin have low molecular weights. In some embodiments, the molecular weight of the penetrating biopolymer is less than 3 kDa. In some embodiments, the molecular weight of the penetrating biopolymer is in the range of 3 kDa to 0.5 kDa.

[0009] The biopolymers of the present disclosure represent an improvement over agents such as HA, at least because they are not derived from animal sources and can be made by fermentation of non-pathogenic microorganisms using agricultural feedstocks.

[0010] Embodiment 1. A biopolymer preparation comprising a biopolymer composed of repeating disaccharide units comprising glucose and galactose, wherein at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, or at least 90% of the glucose is acetylated and at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% of the galactose is pyruvylated.

[0011] Embodiment 2. The biopolymer preparation of embodiment 1, wherein glucose and galactose are linked by β-1,3 glycosidic and α-1,3 glycosidic bonds.

[0012] Embodiment 3. Structure: [ka] wherein the dotted lines represent bonds between the disaccharide units, wherein no more than 40%, no more than 35%, no more than 30%, no more than 25%, no more than 20%, no more than 15%, no more than 10%, or no more than 5% of the disaccharide units lack acetyl moieties and no more than 60%, no more than 55%, no more than 50%, no more than 45%, no more than 40%, no more than 35%, no more than 30%, no more than 25%, no more than 20%, no more than 15%, no more than 10%, no more than 5%, or no more than 1% of the disaccharide units lack pyruvyl moieties.

[0013] Embodiment 4. A biopolymer preparation of any one of embodiments 1-3, wherein the molar ratio of glucose:galactose:pyruvyl:acetyl in the biopolymer is 1:1:0.4-1:0.6-1.

[0014] Embodiment 5. A biopolymer preparation of any one of embodiments 1 to 4, wherein the average molecular weight of the biopolymers in the biopolymer preparation is less than 3000 kDa, less than 1000 kDa, less than 300 kDa, less than 100 kDa or less than 40 kDa.

[0015] Embodiment 6. A biopolymer preparation of any one of embodiments 1 to 4, wherein the average molecular weight of the biopolymers in the biopolymer preparation is between 0.5 kDa and 40 kDa.

[0016] Embodiment 7. The biopolymer preparation of any one of the above embodiments, wherein at least 75%, at least 80%, at least 85%, at least 90% of the biopolymer preparation is biopolymer, or at least 95% (w / w) is biopolymer.

[0017] Embodiment 8. The biopolymer preparation of any one of the above embodiments, capable of absorbing at least the same amount of water as an equivalent amount of hyaluronic acid, at least 1.5 times, at least 2 times, or at least 3 times.

[0018] Embodiment 9. The biopolymer preparation of any one of the above embodiments, capable of absorbing water in an amount of at least 100%, at least 200%, at least 300%, at least 400% or at least 500% of the initial dry weight of the biopolymer preparation.

[0019] Embodiment 10. The biopolymer preparation of embodiment 9 or 10, wherein the water absorption is measured by placing a dried sample of the biopolymer preparation in a humid chamber at 30° C. for 5 days.

[0020] Embodiment 11. The biopolymer preparation of any one of the above embodiments, wherein less than 25%, less than 20%, less than 15%, less than 10%, less than 5%, less than 3%, or less than 1% of the biopolymer preparation is succinoglycan, or wherein the biopolymer preparation is substantially free of succinoglycan.

[0021] Embodiment 12. The biopolymer preparation of any one of the above embodiments, which is a solid or powder.

[0022] Embodiment 13. The biopolymer preparation of any one of the above embodiments, wherein less than 5%, less than 4%, less than 3%, less than 2%, or less than 1% of the biopolymer preparation is water.

[0023] Embodiment 14. A biopolymer preparation comprising a biopolymer composed of repeating disaccharide units comprising glucose and galactose, wherein at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, or at least 90% of the glucose are acetylated and less than 15%, less than 10%, or less than 5% of the galactose are pyruvylated.

[0024] Embodiment 15. The biopolymer preparation of embodiment 14, wherein glucose and galactose are linked by β-1,3 glycosidic bonds and α-1,3 glycosidic bonds.

[0025] Embodiment 16. A biopolymer preparation, comprising a biopolymer having the structure: [ka] (wherein the dotted lines represent the bonds between the disaccharide units), wherein no more than 40%, no more than 35%, no more than 30%, no more than 25%, no more than 20%, no more than 15%, no more than 10%, or no more than 5% of the disaccharide units lack an acetyl moiety, and no more than 15%, no more than 10%, or no more than 5% of the disaccharide units are pyruvylated.

[0026] Embodiment 17. A biopolymer preparation of any one of embodiments 14 to 16, wherein the molar ratio of glucose:galactose:pyruvyl:acetyl in the biopolymer is 1:1:less than 0.5:0.6-1.

[0027] Embodiment 18. A biopolymer preparation of any one of embodiments 14 to 17, wherein the average molecular weight of the biopolymers in the biopolymer preparation is less than 3000 kDa, less than 1000 kDa, less than 300 kDa, less than 100 kDa or less than 40 kDa.

[0028] Embodiment 19. A biopolymer preparation of any one of embodiments 14 to 17, wherein the average molecular weight of the biopolymers in the biopolymer preparation is between 0.5 kDa and 40 kDa.

[0029] Embodiment 20. The biopolymer preparation of any one of embodiments 14 to 19, wherein at least 75%, at least 80%, at least 85%, at least 90% or at least 95% (w / w) of the biopolymer preparation is biopolymer.

[0030] Embodiment 21. A biopolymer preparation according to any one of embodiments 14 to 20, capable of absorbing at least the same amount of water as an equivalent amount of hyaluronic acid, at least 1.1 times, at least 1.2 times, at least 1.3 times, at least 1.4 times or at least 1.5 times.

[0031] Embodiment 22. A biopolymer preparation according to any one of embodiments 14 to 21, capable of absorbing water in an amount of at least 100%, at least 200%, at least 300% or at least 400% of the initial dry weight of the biopolymer preparation.

[0032] Embodiment 23. The biopolymer preparation of embodiment 21 or 22, wherein the water absorption is measured by placing a dried sample of the biopolymer preparation in a humid chamber at 30° C. for 5 days.

[0033] Embodiment 24. The biopolymer preparation of any one of embodiments 14 to 23, wherein less than 25%, less than 20%, less than 15%, less than 10%, less than 5%, less than 3%, or less than 1% of the biopolymer preparation is succinoglycan.

[0034] Embodiment 25. A biopolymer preparation of any one of embodiments 14 to 24, which is a solid or powder.

[0035] Embodiment 26. The biopolymer preparation of any one of embodiments 14 to 25, wherein less than 5%, less than 4%, less than 3%, less than 2%, or less than 1% of the biopolymer preparation is water.

[0036] Embodiment 27. Structure: [ka] A biopolymer preparation comprising a biopolymer composed of repeating polysaccharide units of the formula: (where the dotted lines represent bonds between the polysaccharide units), wherein no more than 30%, no more than 25%, no more than 20%, no more than 15%, no more than 10%, no more than 5%, or no more than 1% of the polysaccharide units lack acetyl moieties and no more than 10%, no more than 5%, or no more than 1% of the polysaccharide units lack pyruvyl moieties.

[0037] Embodiment 28. A biopolymer preparation of embodiment 27, wherein the average molecular weight of the biopolymers in the biopolymer preparation is less than 3000 kDa, less than 1000 kDa, less than 300 kDa, less than 100 kDa, or less than 40 kDa.

[0038] Embodiment 29. A biopolymer preparation of embodiment 27, wherein the average molecular weight of the biopolymers in the biopolymer preparation is between 1.6 kDa and 40 kDa.

[0039] Embodiment 30. The biopolymer preparation of any one of embodiments 27 to 29, wherein at least 75%, at least 80%, at least 85%, at least 90% of the biopolymer preparation is biopolymer, or at least 95% (w / w) is biopolymer.

[0040] Embodiment 31. A biopolymer preparation according to any one of embodiments 27 to 30, capable of absorbing at least the same amount of water as an equivalent amount of hyaluronic acid, at least 1.1 times, at least 1.2 times, at least 1.3 times, at least 1.4 times, or at least 1.5 times.

[0041] Embodiment 32. A biopolymer preparation of any one of embodiments 27 to 31, capable of absorbing water in an amount of at least 100%, at least 200%, at least 300%, or at least 400% of the initial dry weight of the biopolymer preparation.

[0042] Embodiment 33. The biopolymer preparation of embodiment 31 or 32, wherein the water absorption is measured by placing a dried sample of the biopolymer preparation in a humid chamber at 30° C. for 5 days.

[0043] Embodiment 34. A biopolymer preparation of any one of embodiments 27 to 33, which is a solid or powder.

[0044] Embodiment 35. The biopolymer preparation of any one of embodiments 27 to 34, wherein less than 5%, less than 4%, less than 3%, less than 2%, or less than 1% of the biopolymer preparation is water.

[0045] Embodiment 36. A biopolymer preparation comprising a biopolymer composed of repeating polysaccharide units, each polysaccharide unit comprising 2-15, or 2-12, or 2-10 monosaccharides, and wherein the ratio of negative charges:monosaccharides in the repeating polysaccharide units of the biopolymer is at least 0.3, or at least 0.35, or at least 0.4, or at least 0.45.

[0046] Embodiment 37. The biopolymer preparation of embodiment 36, wherein the polysaccharide unit comprises at least one galactose linked to at least one glucose.

[0047] Embodiment 38. The biopolymer preparation of embodiment 37, wherein at least one galactose is linked to glucose via a β-1,3 glycosidic bond.

[0048] Embodiment 39. A biopolymer preparation of any one of embodiments 36 to 38, wherein the average molecular weight of the biopolymers in the biopolymer preparation is less than 3000 kDa, less than 1000 kDa, less than 300 kDa, less than 100 kDa or less than 40 kDa.

[0049] Embodiment 40. A biopolymer preparation of any one of embodiments 36 to 38, wherein the average molecular weight of the biopolymers in the biopolymer preparation is between 0.5 kDa and 40 kDa.

[0050] Embodiment 41. The biopolymer preparation of any one of embodiments 36 to 40, wherein at least 75%, at least 80%, at least 85%, at least 90% or at least 95% (w / w) of the biopolymer preparation is biopolymer.

[0051] Embodiment 42. A biopolymer preparation according to any one of embodiments 36 to 41, capable of absorbing at least the same amount of water as an equivalent amount of hyaluronic acid, at least 1.1 times, at least 1.5 times, at least 2 times, or at least 3 times.

[0052] Embodiment 43. A biopolymer preparation of any one of embodiments 36 to 42, capable of absorbing water in an amount of at least 100%, at least 200%, at least 300% or at least 400% of the initial dry weight of the biopolymer preparation.

[0053] Embodiment 44. The biopolymer preparation of embodiment 42 or 43, wherein the water absorption is measured by placing a dried sample of the biopolymer preparation in a humid chamber at 30° C. for 5 days.

[0054] Embodiment 45. A biopolymer preparation of any one of embodiments 36 to 44, which is a solid or powder.

[0055] Embodiment 46. The biopolymer preparation of any one of embodiments 36 to 45, wherein less than 5%, less than 4%, less than 3%, less than 2%, or less than 1% of the biopolymer preparation is water.

[0056] Embodiment 47. A topical composition comprising a biopolymer, the biopolymer being composed of repeating disaccharide units comprising glucose and galactose, wherein at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, or at least 90% of the glucose is acetylated and at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% of the galactose is pyruvylated.

[0057] Embodiment 48. The topical composition of embodiment 47, wherein glucose and galactose are linked by β-1,3 glycosidic bonds and α-1,3 glycosidic bonds.

[0058] Embodiment 49. A topical composition comprising a biopolymer, the biopolymer having the structure: [ka] (wherein the dotted lines represent bonds between the disaccharide units), wherein no more than 40%, no more than 35%, no more than 30%, no more than 25%, no more than 20%, no more than 15%, no more than 10%, or no more than 5% of the disaccharide units lack an acetyl moiety and no more than 60%, no more than 55%, no more than 50%, no more than 45%, no more than 40%, no more than 35%, no more than 30%, no more than 25%, no more than 20%, no more than 15%, no more than 10%, no more than 5%, or no more than 1% of the disaccharide units lack a pyruvyl moiety.

[0059] Embodiment 50. The topical composition of any one of embodiments 47-49, wherein the molar ratio of glucose:galactose:pyruvyl:acetyl in the biopolymer is 1:1:0.4-1:0.6-1.

[0060] Embodiment 51. The topical composition of any one of embodiments 47 to 50, wherein the average molecular weight of the biopolymer in the topical composition is less than 3000 kDa, less than 1000 kDa, less than 300 kDa, less than 100 kDa, or less than 40 kDa.

[0061] Embodiment 52. The topical composition of any one of embodiments 47 to 51, wherein the average molecular weight of the biopolymer in the topical composition is between 0.5 kDa and 40 kDa.

[0062] Embodiment 53. The topical composition of any one of embodiments 47-53, wherein the biopolymer is capable of absorbing at least the same amount of water as an equivalent amount of hyaluronic acid, at least 1.5 times, at least 2 times, or at least 3 times as much water.

[0063] Embodiment 54. The topical composition of any one of embodiments 47-53, wherein the biopolymer is capable of absorbing water in an amount of at least 100%, at least 200%, at least 300%, at least 400%, or at least 500% of the initial dry weight of the biopolymer.

[0064] Embodiment 55. The topical composition of embodiment 53 or 54, wherein the water absorption is measured by placing a dry sample of the biopolymer in a humidity chamber at 30° C. for 5 days.

[0065] Embodiment 56. A topical composition according to any one of embodiments 47 to 55, comprising a biopolymer preparation according to any one of embodiments 1 to 13.

[0066] Embodiment 57. The topical composition of any one of embodiments 47-56, which is substantially free of succinoglycan.

[0067] Embodiment 58. A topical composition comprising a biopolymer, the biopolymer being composed of repeating disaccharide units comprising glucose and galactose, wherein at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, or at least 90% of the glucose is acetylated and less than 15%, less than 10%, or less than 5% of the galactose is pyruvylated.

[0068] Embodiment 59. The topical composition of embodiment 58, wherein the glucose and galactose are linked by β-1,3 and α-1,3 bonds.

[0069] Embodiment 60. A topical composition comprising a biopolymer, the biopolymer having the structure: [ka] (wherein the dotted lines represent bonds between the disaccharide units), wherein no more than 40%, no more than 35%, no more than 30%, no more than 25%, no more than 20%, no more than 15%, no more than 10%, or no more than 5% of the disaccharide units lack an acetyl moiety, and no more than 15%, no more than 10%, or no more than 5% of the disaccharide units are pyruvylated.

[0070] Embodiment 61. The topical composition of any one of embodiments 58-60, wherein the molar ratio of glucose:galactose:pyruvyl:acetyl in the biopolymer is 1:1:less than 0.5:0.6-1.

[0071] Embodiment 62. The topical composition of any one of embodiments 58 to 61, wherein the average molecular weight of the biopolymer in the topical composition is less than 3000 kDa, less than 1000 kDa, less than 300 kDa, less than 100 kDa, or less than 40 kDa.

[0072] Embodiment 63. The topical composition of any one of embodiments 58 to 61, wherein the average molecular weight of the biopolymer in the topical composition is between 0.5 kDa and 40 kDa.

[0073] Embodiment 64. The topical composition of any one of embodiments 58 to 63, wherein the biopolymer preparation is capable of absorbing at least the same amount of water as an equivalent amount of hyaluronic acid, at least 1.1 times, at least 1.2 times, at least 1.3 times, at least 1.4 times, or at least 1.5 times.

[0074] Embodiment 65. The topical composition of any one of embodiments 58 to 64, wherein the biopolymer preparation is capable of absorbing water in an amount of at least 100%, at least 200%, at least 300%, or at least 400% of the initial dry weight of the biopolymer.

[0075] Embodiment 66. The topical composition of embodiment 64 or 65, wherein the water absorption is measured by placing a dry sample of the biopolymer in a humidity chamber at 30° C. for 5 days.

[0076] Embodiment 67. A topical composition according to any one of embodiments 58 to 66, comprising a biopolymer preparation according to any one of embodiments 14 to 26.

[0077] Embodiment 68. The topical composition of any one of embodiments 58-67, which is substantially free of succinoglycan.

[0078] Embodiment 69. A topical composition comprising a biopolymer, the biopolymer having the structure: [ka] (wherein the dotted lines represent bonds between the polysaccharide units), wherein no more than 30%, no more than 25%, no more than 20%, no more than 15%, no more than 10%, no more than 5%, or no more than 1% of the polysaccharide units lack acetyl moieties and no more than 10%, no more than 5%, or no more than 1% of the polysaccharide units lack pyruvyl moieties.

[0079] Embodiment 70. The topical composition of embodiment 69, wherein the average molecular weight of the biopolymer in the topical composition is less than 3000 kDa, less than 1000 kDa, less than 300 kDa, less than 100 kDa, or less than 40 kDa.

[0080] Embodiment 71. The topical composition of embodiment 69, wherein the average molecular weight of the biopolymer in the topical composition is between 1.6 kDa and 40 kDa.

[0081] Embodiment 72. The topical composition of any one of embodiments 69 to 71, wherein the biopolymer preparation is capable of absorbing at least the same amount of water as an equivalent amount of hyaluronic acid, at least 1.1 times, at least 1.2 times, at least 1.3 times, at least 1.4 times, or at least 1.5 times.

[0082] Embodiment 73. The topical composition of any one of embodiments 69-72, wherein the biopolymer preparation is capable of absorbing water in an amount of at least 100%, at least 200%, at least 300%, or at least 400% of the initial dry weight of the biopolymer.

[0083] Embodiment 74. The topical composition of embodiment 72 or 73, wherein the water absorption is measured by placing a dry sample of the biopolymer in a humidity chamber at 30° C. for 5 days.

[0084] Embodiment 75. The topical composition of any one of embodiments 69 to 74, comprising a biopolymer preparation of any one of embodiments 27 to 35.

[0085] Embodiment 76. A topical composition comprising a biopolymer, wherein the biopolymer is composed of repeating polysaccharide units, each polysaccharide unit comprising 2-15, or 2-12, or 2-10 monosaccharides, and wherein the ratio of negative charges:monosaccharides in the repeating polysaccharide units of the biopolymer is at least 0.3, or at least 0.35, or at least 0.4, or at least 0.45.

[0086] Embodiment 77. The topical composition of embodiment 76, wherein the polysaccharide unit comprises at least one galactose linked to at least one glucose.

[0087] Embodiment 78. The topical composition of embodiment 77, wherein at least one galactose is linked to glucose via a β-1,3 glycosidic bond.

[0088] Embodiment 79. The topical composition of any one of embodiments 76 to 78, wherein the average molecular weight of the biopolymer in the topical composition is less than 3000 kDa, less than 1000 kDa, less than 300 kDa, less than 100 kDa, or less than 40 kDa.

[0089] Embodiment 80. The topical composition of any one of embodiments 76 to 78, wherein the average molecular weight of the biopolymer in the topical composition is between 0.5 kDa and 40 kDa.

[0090] Embodiment 81. The topical composition of any one of embodiments 76 to 80, wherein the biopolymer preparation is capable of absorbing at least the same amount of water as an equivalent amount of hyaluronic acid, at least 1.1 times, at least 1.5 times, at least 2 times, or at least 3 times.

[0091] Embodiment 82. The topical composition of any one of embodiments 76-81, wherein the biopolymer preparation is capable of absorbing at least the same, at least 1.5 times, at least 2 times, at least 2.5 times, at least 3 times, at least 4 times, or at least 5 times the amount of water compared to the initial dry weight of the biopolymer.

[0092] Embodiment 83. The topical composition of embodiment 81 or 82, wherein the water absorption is measured by placing a dry sample of the biopolymer in a humidity chamber at 30° C. for 5 days.

[0093] Embodiment 84. The topical composition of any one of embodiments 76 to 83, comprising a biopolymer preparation of any one of embodiments 36 to 46.

[0094] Embodiment 85. The topical composition of any one of embodiments 47 to 84, comprising 0.01% to 10% (w / w) or 0.05% to 5% (w / w) of a biopolymer.

[0095] Embodiment 86. The topical composition of any one of embodiments 47-85, comprising one or more thickeners, stabilizers, emulsifiers, emollients, moisturizers, rheology modifiers, film formers, antioxidants, additives, actives, butters, essential oils, exfoliants, clays, muds, extracts, hydrosols, exfoliants, supplements, waxes, thickeners, salts, minerals, acids, bases, carrier and fixed oils, surfactants, preservatives, pearlizing agents, quality improvers, structuring agents, brighteners, humectants, osmolytes, occlusive agents, cleansers, colorants, pigments, fragrances, UV-A and UV-B blockers, and / or nutrients.

[0096] Embodiment 87. The topical composition of any one of embodiments 47-86, comprising hyaluronic acid.

[0097] Embodiment 88. The topical composition of any one of embodiments 47-86, which does not contain hyaluronic acid.

[0098] Embodiment 89. The topical composition of any one of embodiments 47-88, which is a cream, lotion, ointment, balm, tincture, liniment, shampoo, soap, conditioner, sunscreen, rinse, deodorant, or cosmetic.

[0099] Embodiment 90. A method of treating or preventing a skin condition in a subject comprising applying a topical composition of any one of embodiments 47-89.

[0100] Embodiment 91. The method of embodiment 90, wherein the skin condition is one or more of dry skin, wrinkled skin, sagging skin, aged skin, blemished skin, injured skin, blemished skin, acne, and / or sun-damaged skin.

[0101] Embodiment 92. The method of embodiment 90, wherein the skin condition is an inflammatory skin condition.

[0102] Embodiment 93. The method of embodiment 92, wherein the inflammatory skin condition is psoriasis, eczema, or atopic dermatitis.

[0103] Embodiment 94. The method of any one of embodiments 90-93, wherein the topical composition is applied to the subject's face, ears, forehead, neck, arms, upper chest, and / or hands.

[0104] Embodiment 95. The method of any one of embodiments 90-94, wherein the topical composition reduces transepidermal water loss, reverses transepidermal water loss, improves epidermal water retention, reduces the appearance of wrinkles, reduces sagging skin, increases skin smoothness, increases skin swelling, improves skin flexibility, improves skin texture, reduces skin blemishes, and / or reduces skin dryness. [Brief description of the drawings]

[0105] [Figure 1] FIG. 1 shows a representative repeating unit of a polysaccharide biopolymer. Individual sugar residues are linked by glycosidic bonds. Polysaccharide-based biopolymers consist of a backbone of sugar residues connected by glycosidic bonds. The backbone may also have side chains made of sugar residues, each of which may have different chemical modifications. The biopolymer backbone and side chains, together with the chemical modifications, make up the repeating unit. The repeating units are connected by glycosidic bonds to produce longer polymers. [Diagram 2] 1 shows biopolymer structures: Galactoglucan is a repeating dimer of galactose and glucose with pyruvyl and acetyl modifications; Succinoglycan is a repeating octamer of one galactose and seven glucose residues with pyruvyl, acetyl and succinyl modifications; Glucuronoglycan is a repeating nonamer of two galactose, two glucuronic acid and five glucose residues with pyruvyl and acetyl modifications. [Diagram 3] Figure 1 shows water absorption of isolated and blended biopolymers compared to HA. The ability to bind water, as measured by mass gain, was determined for individual and blended biopolymers. The mass gain of each biopolymer was divided by the mass gain of hyaluronic acid (HA) to derive the relative fold change in water retention. [Figure 4] Figure 1 shows the correlation between negative charge and water absorption. The charge of each biopolymer was calculated at physiological pH based on the pKa of the acidic modifying group or sugar acid. This value was then divided by the number of sugar residues in the repeating unit to obtain a ratio for each biopolymer. The charge density was plotted against the percent mass gain of each biopolymer. [Diagram 5] Figure 1 shows the water absorption of non-pyruvylated galactoglucan compared to HA. The water binding capacity was measured as described in Example 4 and Figure 3. Mass gain was normalized to the value for HA and fold change was calculated. [Figure 6] Figure 1 shows the cytotoxicity of biopolymers. Cytotoxicity tests were performed according to the ISO 10993-5 standard. Biopolymers were dissolved in PBS at a concentration of 1% (w / v), heat sterilized, and used in the cytotoxicity assay. Values ​​for each biopolymer were compared with both positive and negative controls. [Figure 7] Figure 1 shows skin water retention measurements. Measurements were performed using a Corneometer 825 (Courage + Khazaka). Two healthy volunteers applied either a lotion base containing 0.2% (w / v) galactoglucan or a control base (containing an equal amount of water) to two different sites on the body, namely the hands and elbows, for 48 hours. The mean change in water retention (arbitrary units measured by the Corneometer) from the baseline measurement for each site on the body for each participant is shown. [Figure 8] Figure 10: Reduction in the appearance of wrinkles with galactoglucan treatment. Image of the back of the hand of participant P1 from Example 8. The circles highlight the areas where a reduction in the appearance of wrinkles can be observed. [Figure 9]Figure 1 shows the results of transdermal permeability and skin retention assays. Unidirectional permeability through ex vivo human skin tissue was determined using Franz cells. Results are shown for the amount of carbohydrate in a representative receptor chamber (permeate) after 4 hours of treatment with a donor solution of 5% (w / v) non-pyruvyl galactoglucan. After completion of the Franz cell assay, the skin samples were rinsed and then immersed in PBS to elute carbohydrate absorbed or associated with the skin surface. Total carbohydrate in the receptor solution or eluted skin samples was determined using the Anthrone assay. Background levels from control receptor chambers and skin samples were subtracted to derive reported values. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0106] Natural product biopolymers with improved water-binding properties that are low molecular weight (LMW) and produced by fermentation of non-pathogenic microorganisms are attractive for a number of different reasons. First, such biopolymers are more cost-effective to produce than traditional hyaluronic acid and can be added at higher concentrations to personal care and cosmetic products to achieve improved anti-wrinkle and anti-aging performance. Second, these biopolymers can be added to a substantially wider range of personal care products, such as lip balms, shaving creams, sunscreens, shampoos, conditioners, and soaps, where skin penetration and water retention improve performance. Third, biopolymers produced by microbial fermentation are an attractive alternative to animal-derived HA, reducing the social and / or ethical issues associated with animal use. Biopolymers derived from harmless microorganisms, as opposed to pathogenic microorganisms, require fewer downstream processing steps to produce a safe product. Finally, biopolymers produced naturally by LMW have significant advantages over current LMW HA manufacturing processes, which require several additional downstream processing steps to achieve performance specifications.

[0107] The present invention provides compositions of biopolymers based on exopolysaccharides produced by non-pathogenic species of soil bacteria, and their cosmetic and / or dermatological or pharmaceutical uses. High water binding capacity and skin retention / penetration are desirable properties of personal care ingredients with anti-aging and anti-wrinkle effects. The most widely used polymer with these properties is hyaluronic acid, which is derived from either animals or pathogenic bacteria. Provided herein are isolated biopolymers with substantially improved water binding properties compared to hyaluronic acid. In some embodiments, the isolated biopolymers are derived from Rhizobiaceae bacteria. These bacteria naturally produce low molecular weight biopolymers with the ability to penetrate the skin. The production of these biopolymers by fermentation is highly advantageous compared to the production process of hyaluronic acid.

[0108] The present inventors have identified certain biopolymers that have superior water retention properties to HA, making them ideal for use as anti-aging products in a wide range of personal care and cosmetic formulations. The first molecule, galactoglucan, is a repeating disaccharide of galactose and glucose with pyruvyl and acetyl modifications. Galactoglucan is derived from the bacterium Sinorhizobium meliloti (also known as Ensifer meliloti) and is naturally produced at low molecular weight (LMW) during fermentation. Galactoglucan is one of two biopolymers naturally produced by this organism, and water-binding properties can only be observed in galactoglucan, independent of the other biopolymer, succinoglycan. The present inventors have demonstrated that galactoglucan has a water retention capacity greater than 3.5 times that of HA. A mutant of galactoglucan lacking the pyruvyl modification exhibits a water retention capacity 1.7 times that of HA. The second molecule, glucuronoglycan, is a repeating nonasaccharide containing galactose, glucuronic acid, and glucose with pyruvyl and acetyl modifications. Glucuronoglycan is derived from the bacterium Sinorhizobium fredi (also known as Ensifer fredii) and is produced naturally as a mixture of high and low molecular weight molecules. We have demonstrated that glucuronoglycan has a water-holding capacity 1.7 times higher than that of HA.

[0109] In addition to increasing water retention performance, galactoglucans and glucuronoglycans are non-toxic in standard preclinical models, which is a prerequisite for the suitability of the compounds in cosmetic and dermatological applications. In some embodiments, these LMW biopolymer fractions penetrate skin samples in an in vitro Franz cell assay. In personal care formulations, in some embodiments, the addition of biopolymers from the Rhizobium family to a typical lotion base leads to improved water retention in the skin.

[0110] The present invention provides microbial polymers with water retention, skin surface retention and / or penetration properties that are beneficial performance characteristics for use as anti-aging and anti-wrinkle ingredients in personal care and cosmetic formulations. These microbial biopolymers exhibit substantially improved water retention compared to hyaluronic acid, an industry standard active ingredient used in many skin care products for its anti-aging properties. The Rhizobiaceae-derived biopolymers described herein are suitable for use in a wide range of aqueous skin care formulations and can provide increased skin water retention efficacy in many consumer products. Furthermore, the production of LMW biopolymers by fermentation requires little downstream processing and provides an economically advantageous production method compared to existing technologies for the production of animal- or microbial-derived HA of specific molecular weights.

[0111] Biopolymers and Biopolymer Preparations Rhizobiaceae, a family of symbiotic bacteria living in soil, has been studied for decades for its ability to provide fixed nitrogen to legume hosts, but until now has been underutilized as a fermenting microorganism for the production of bioproducts, pharmaceuticals or cosmetics. These bacteria naturally produce water-soluble exopolysaccharides, or biopolymers, that are involved in both symbiosis with the host plant and in biofilm formation. The diversity of exopolysaccharides produced by Rhizobiaceae suggests a range of novel biopolymers with novel functionalities that could add substantial value to several markets.

[0112] Succinoglycan and Galactoglucan Sinorhizobium (ensifer) meliloti naturally produces two acidic exopolysaccharides: succinoglycan (EPS I) and galactoglucan (EPS II) (Barnett 2018). Succinoglycan is the major exopolysaccharide produced by S. meliloti. The repeating unit of succinoglycan (Figure 2) consists of glucose and galactose in a 7:1 ratio, with acetyl, pyruvyl, and succinyl modifications (Reuber 1993). Succinoglycan is naturally produced by S. meliloti in both high and low molecular weights. The general mechanism of succinoglycan biosynthesis is relatively well understood and is likely common to related organisms producing similar biopolymers.

[0113] Galactoglucan production is restricted to species phylogenetically close to S. meliloti. The repeating unit of galactoglucan (Figure 2) consists of glucose and galactose in a 1:1 ratio, with acetyl and pyruvyl modifications (Glazebrook 1989). The specific linkage is β-D-Glcp-(1-3)-α-D-Galp-(1-3), with 6-O-acetyl on most D-glucose residues and 4,6-O-pyruvyl on all D-galactose residues (Her 1990). Galactoglucan is naturally produced by S. meliloti at low molecular weight. Compared to succinoglycan, relatively little is known about the biosynthetic pathway of galactoglucan.

[0114] Glucuronoglycan Sinorhizobium (ensifer) fredii naturally produces an acidic exopolysaccharide consisting of glucose, galactose, and glucuronic acid in a 5:2:2 ratio, with acetyl and pyruvyl modifications (Djordjevic 1986) (Figure 2). The biopolymer shown in Figure 2 is produced by several S. fredii type strains, including NGR234, HH103 (ATCC51809), and possibly USDA257 (Gray 1991, Pueppke 1999, Rodriguez-Navarro 2014). Structural analysis indicates that the terminal galactose on the side chain of the glucuronoglycan is 4,6-pyruvylated and may be acetylated at either the second, third, or both carbons of the same sugar residue. A third acetyl group was also detected in glucuronoglycan from strain HH103, and thus likely in other strains as well, but its location remains to be elucidated (Staehelin 2006, Rodriguez-Navarro 2014). The exoregion of these S. fredii species spans 28 kb and has a high degree of synteny with the S. meliloti cluster involved in succinoglycan biosynthesis. Many of its genes share homology with exogenes in S. meliloti (Zhan 1990). Glucuronoglycan has a similar structure to succinoglycan, but contains glucuronic acid and is not succinylated. The main chain of glucuronoglycan consists of six sugar residues, whereas the main chain of succinoglycan contains four sugar residues. Glucuronoglycan is produced by S. fredii strains in both high and low molecular weights (Staehelin 2006).

[0115] In some embodiments, a biopolymer is provided that is composed of repeating disaccharide units comprising glucose and galactose, wherein at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, or at least 90% of the glucose are acetylated and at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% of the galactose are pyruvylated. In some embodiments, the glucose and galactose are linked by β-1,3 glycosidic bonds and α-1,3 glycosidic bonds. In some embodiments, the structure: [ka] (where the dotted line represents the bond between the disaccharide units), wherein no more than 40%, no more than 35%, no more than 30%, no more than 25%, no more than 20%, no more than 15%, no more than 10%, or no more than 5% of the disaccharide units lack an acetyl moiety and no more than 60%, no more than 55%, no more than 50%, no more than 45%, no more than 40%, no more than 35%, no more than 30%, no more than 25%, no more than 20%, no more than 15%, no more than 10%, no more than 5%, or no more than 1% of the disaccharide units lack a pyruvyl moiety. In some embodiments, the biopolymer is comprised in a biopolymer preparation. In some such embodiments, the molar ratio of glucose:galactose:pyruvyl:acetyl in the biopolymer preparation is 1:1:0.4-1:0.6-1. In some embodiments, biopolymer preparations are provided that contain less than 25%, less than 20%, less than 15%, less than 10%, less than 5%, less than 3%, or less than 1% succinoglycan, hi some embodiments, the biopolymer preparations are substantially free of succinoglycan.

[0116] In some embodiments, a biopolymer is provided that is composed of repeating disaccharide units comprising glucose and galactose, where at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, or at least 90% of the glucose are acetylated and less than 15%, less than 10%, or less than 5% of the galactose are pyruvylated. In some embodiments, the glucose and galactose are linked by β-1,3 glycosidic and α-1,3 glycosidic bonds. In some embodiments, the biopolymer has the structure: [ka] Provided are biopolymers composed of repeating disaccharide units of the formula: (where the dotted lines represent bonds between the disaccharide units), in which no more than 40%, no more than 35%, no more than 30%, no more than 25%, no more than 20%, no more than 15%, no more than 10%, or no more than 5% of the disaccharide units lack an acetyl moiety and no more than 15%, no more than 10%, or no more than 5% of the disaccharide units are pyruvylated. In some embodiments, the biopolymer is included in a biopolymer preparation. In some such embodiments, the molar ratio of glucose:galactose:pyruvyl:acetyl in the biopolymer preparation is 1:1:less than 0.5:0.6-1. In some embodiments, biopolymer preparations are provided that contain less than 25%, less than 20%, less than 15%, less than 10%, less than 5%, less than 3%, or less than 1% succinoglycan. In some embodiments, the biopolymer preparation is substantially free of succinoglycan.

[0117] In some embodiments, the biopolymer has the structure: [ka] (where the dotted lines represent bonds between polysaccharide units), wherein no more than 30%, no more than 25%, no more than 20%, no more than 15%, no more than 10%, no more than 5%, or no more than 1% of the polysaccharide units lack an acetyl moiety and no more than 10%, no more than 5%, or no more than 1% of the disaccharide units lack a pyruvyl moiety. In some embodiments, the polysaccharide units contain one, two, or three acetyl groups. In some embodiments, the average number of acetyl groups per polysaccharide unit is 1-3.

[0118] In some embodiments, a biopolymer is provided, wherein the biopolymer is composed of repeating polysaccharide units, each polysaccharide unit comprising 2-15, or 2-12, or 2-10 monosaccharides, and wherein the negative charge:monosaccharide ratio in the repeating polysaccharide units of the biopolymer is at least 0.3, or at least 0.35, or at least 0.4, or at least 0.45. In some embodiments, the polysaccharide units comprise at least one galactose linked to at least one glucose. In some such embodiments, the at least one galactose is linked to the glucose via a β-1,3 glycosidic bond.

[0119] In various embodiments, biopolymer preparations are provided, wherein at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% (w / w) of the biopolymer preparation is a biopolymer provided herein. In some embodiments, the biopolymer preparation is a solid or powder. In some embodiments, less than 5%, less than 4%, less than 3%, less than 2%, or less than 1% of the biopolymer preparation is water.

[0120] In various embodiments, the average molecular weight of the biopolymers in the biopolymer preparation is less than 3000 kDa, less than 1000 kDa, less than 300 kDa, less than 100 kDa, or less than 40 kDa. In some embodiments, the average molecular weight of the biopolymers in the biopolymer preparation is between 0.5 kDa and 40 kDa, or between 1.6 kDa and 40 kDa.

[0121] In some embodiments, the biopolymer can absorb at least the same amount of water as an equivalent amount of hyaluronic acid, at least 1.1 times, at least 1.2 times, at least 1.3 times, at least 1.4 times, 1.5 times, at least 2 times, or at least 3 times. In some embodiments, the biopolymer can absorb at least 100%, at least 200%, at least 300%, at least 400%, or at least 500% of the initial dry weight of the biopolymer preparation. Water absorption can be measured, for example, by placing a dry sample of the biopolymer or biopolymer preparation in a humidified chamber at a substantially constant temperature for a period of time. In some embodiments, the temperature is about 20° C., about 25° C., about 30° C., about 35° C., or about 37° C. In some embodiments, the period of time is 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, or 1 week.

[0122] Strain construction Both S. meliloti and S. fredii are amenable to genetic modification, and a common method of strain engineering is to delete specific regions in the genome using homologous recombination, antibiotic resistance, and sucrose counterselection (Quandt 1993). Plasmids containing modified genomic regions can then be constructed and used to replace the native regions with the targeted changes. These non-replicating plasmids are introduced by conjugal transfer, allowing strains with single integrants to be selected by antibiotic resistance and confirmed by PCR. Secondarily, the integrated plasmid can be counterselected by the presence of the sacB gene, which encodes a levansucrase that is lethal to Gram-negative bacteria in the presence of sucrose. The antibiotic-sensitive, sucrose-resistant strains then recombine into the wild type or incorporate the deletion, insertion, or other modification that was present in the constructed plasmid. The modified strains can be confirmed by PCR and sequencing.

[0123] Unmodified, unadapted strains of S. meliloti produce both succinoglycan and galactoglucan and are suitable for the simultaneous production of both biopolymers. In certain type strains, such as Rm1021 (ATCC51124), the ability to produce galactoglucan has been lost due to strain adaptation in the laboratory (Charoenpanich 2015). In the case of adapted strains, there are several ways in which galactoglucan-producing strains can be constructed. Examples include restoration of an intact expR gene, knockout of mucR, overexpression of WggR (Bahlawane 2008) or growth in phosphate-limiting medium (Mendrygal 2000).

[0124] For the production of succinoglycan in the absence of galactoglucan, the reference strain Rm1021 can be used. Several regulatory genes can be modified to obtain strains that overproduce succinoglycan. These genes include exoR, exoS, chvI, syrM, and nodD3 (Barnett 2015). Others include syrA, mucR (Keller 1995), and exoX (Zhan 1990). When using unadapted strains of S. meliloti, it is necessary to knock out the galactoglucan biosynthesis genes to generate strains that produce only succinoglycan. The genes required for galactoglucan biosynthesis are contained in a 32 kb region of pSymB and include six predicted glycosyltransferases and four genes predicted to code for proteins required for the synthesis of dTDP-glucose and dTDP-rhamnose (Becker 1997). To eliminate production of galactoglucan, any of several glycosyltransferases, such as wgaB or wgeB, can be ablated.

[0125] To produce galactoglucan in the absence of succinoglycan, wild-type strains of S. meliloti with mutations in succinoglycan biosynthetic genes can be generated using pJQ200SK. Additionally, domesticated strains that have been restored the ability to produce galactoglucan by any of the methods described above may be used. A succinoglycan-specific biosynthetic cluster is located in a 22 kb region on pSymB. Structural and regulatory roles have been assigned to some of the genes in this cluster (Reuber 1993). To eliminate succinoglycan biosynthesis, any of several genes, such as exoA, exoF, exoL, exoM, exoP, exoQ, exoT, or exoY, can be genetically excised.

[0126] Glucuronoglycan is the major product of wild-type S. fredii and no modification of the strain is required for the production of this biopolymer. Any of the above strains can be used for production of the material and further analysis.

[0127] Methods for making biopolymers Several different liquid growth media can be used for the production of biopolymers. S. meliloti strains grow well in LB or TY medium, which may be supplemented with additional carbon sources such as glucose, sucrose or succinate to enhance product production. S. fredii can be grown in TY medium, and the addition of additional carbon sources is beneficial for production. Both S. meliloti and S. fredii can be grown in defined minimal media such as M9 or MOPS-mannitol, resulting in higher yields. Minimal media allows precise control of fermentation variables such as phosphate concentration, pH, micronutrients, sulfate concentration, and carbon source.

[0128] Alcohol precipitation may be used to purify the biopolymer after fermentation. Typically, cells are removed from the fermentation broth by centrifugation or filtration. The high viscosity of the fermentation broth may require the addition of 1-2 volumes of water to aid in the cell separation procedure. The cell-free supernatant may be incubated with a protease to further remove residual cells or cell debris. To precipitate the biopolymer, isopropanol or ethanol in the concentration range of around 1 mM, as well as monovalent or divalent cations such as KCl or CaCl2, typically at 1-2 times the culture volume, may be added to the cell-free supernatant. The biopolymer is precipitated by mixing and can be isolated by centrifugation or filtration. At this point, further purification steps may be performed to reduce the salt concentration or any cell debris that may have precipitated with the polymer. These steps may include additional alcohol washes, protease treatment, rehydration, centrifugation, dialysis, solvent washes, lyophilization, etc., appropriate to the desired end use. The purified product may be dried in an oven until mass is stable (all unbound water has evaporated). The dried product can be ground, milled or otherwise processed to produce the final purified biopolymer.

[0129] Topical Compositions Hyaluronic acid (HA) plays an important role in skin aging and HA addition can be found in many personal care lotions and creams. In certain common formulations, HMW HA is used because it stays on the surface of the skin and retains moisture by absorbing water from the air. In other formulations, LMW HA is used because it can penetrate the skin and provide anti-aging or anti-wrinkle effects. It has been shown that skin permeability is directly related to the length of the polymer (Essendoubi 2016, Witting 2015). Penetration into the lower layers of the epidermis, binding of water and swelling of the skin contribute to reducing the appearance of wrinkles. Molecules with these capabilities are therefore highly sought after for skin care formulations that reduce the appearance of aging.

[0130] Functional ingredients are typically included in personal care products at concentrations of 2% (w / v) or less. Higher concentrations may be more effective in improving skin appearance, but raw material costs likely do not allow ingredients to be added at levels above 2%. High costs may also prevent more widespread use in commonly used personal care products. Despite these limitations, there are few alternative products that can compete with the performance and efficacy of HA in dermal fillers and topical anti-aging personal care formulations. Dermal filler alternatives include products such as biomineral hydroxylapatite or petroleum-derived products such as polymethylmethacrylate. Biological alternatives include agarose, which is often used in combination with HA, non-carbohydrate-based polyglutamic acid (PGA), or polymers derived from tamarind seed extract.

[0131] Examples of topically applied products include any aqueous, alcoholic, or oil-in-water emulsion, such as a cream, lotion, serum, ointment, balm (such as lip balm), tincture, liniment, shampoo, soap, conditioner, sunscreen, rinse, cosmetic, deodorant, or any other treatment used directly on the skin. The topical compositions provided herein may be generally fluid and may be in the form of a salve, emulsion, cream, milk, ointment, impregnated pad, detergent, solution, serum, gel, spray or aerosol, foam, suspension, lotion, or stick.

[0132] In various embodiments, a topical composition is provided that includes at least one biopolymer provided herein. In some embodiments, the topical composition includes one, two, or three of the biopolymers provided herein. In some embodiments, the composition includes 0.01%-30%, 1%-30%, 0.01%-20%, 1%-20%, 0.01%-10%, or 0.05%-5% (w / v) of the biopolymer provided herein. In some embodiments, the topical composition includes at least one biopolymer provided herein and hyaluronic acid. In some such embodiments, the hyaluronic acid is present in a concentration of 0.01%-5%, 0.01%-3%, 0.01%-2%, 0.01%-1%, 0.1%-2%, or 0.1%-1% (w / v).

[0133] In various embodiments, the topical compositions provided herein include one or more pharma- ceutically and / or cosmetically acceptable thickeners, stabilizers, emulsifiers, emollients, moisturizers, rheology modifiers, film formers, antioxidants, additives, actives, butters, essential oils, exfoliants, clays, muds, extracts, hydrosols, exfoliants, supplements, waxes, thickeners, salts, minerals, acids, bases, carrier and fixed oils, surfactants, preservatives, pearlizing agents, texture improvers, structurants, brighteners, humectants, osmolytes, occlusive agents, detergents, colorants, pigments, fragrances, UV-A and UV-B blockers, and / or nutrients. Those skilled in the art can select suitable ingredients for topical compositions based on the desired application. Numerous lists and descriptions of suitable ingredients are available in the art, including, for example, cir-safety.org and / or www.fda.gov / cosmetics / cosmetic-products-ingredients. Non-limiting examples of viscosity and / or rheology adjusting agents include polyacrylates, their derivatives and copolymers, polysorbates and derivatives, myristates, polyquaterniums, cellulose and derivatives, cetearyl alcohol, carbomer, xanthan gum, diutan gum, capric acid glyceride, processed sugar, other polysaccharide polymers, paraffin, polyethylene glycol, glycerol, propanediol, oleic acid derivatives, and hyaluronic acid.Non-limiting examples of moisturizing agents include hyaluronic acid, methyl glucose ether, ethylated methyl glucose, glycerol, polyethylene glycol, glycol derivatives, collagen, urea, sorbitol, allantoin, and alpha hydroxy acid.Non-limiting examples of emollients include mineral oil, lanolin, shea butter, cocoa butter, coconut oil, beeswax, sunflower oil, other vegetable oils, vegetable and animal fats, petrolatum, and squalene. Additional ingredients of topical compositions include, but are not limited to, silicone derivatives (lubricants), tocopherol (vitamin E), parabens (preservatives), stearic and oleic acids, sodium lauryl sulfate (surfactants), keratin, elastin (proteins), amino acids and peptides, and bentonite clay.

[0134] Use of the topical composition Skin is the human body's first line of defense against external challenges such as fever, infection, exposure to toxic substances, and water loss. The outermost layer of the epidermis, called the stratum corneum, is composed of stacked non-viable cells that do not retain water. The primary function of the stratum corneum is to maintain skin homeostasis by preventing excessive transepidermal water loss. Beneath the stratum corneum lies the rest of the epidermis, which is composed of various types of metabolically active cells such as keratinocytes and melanocytes. Beneath the epidermis lies the dermis and subcutaneous tissue, which contain blood vessels, nerve cells, and apocrine glands (Abd 2016).

[0135] Although the stratum corneum is the primary barrier to water loss, the presence of HA in metabolically active layers of the epidermis has been shown to be the most important factor in the skin's overall water retention level. Aging or aged skin is characterized by both a decrease in polymer length and an overall loss of polymer in the epidermis (Papakonstantinou 2012). A water-retaining polymer that can pass the stratum corneum barrier and penetrate the epidermis could potentially counteract the effects of skin aging mentioned above. Application of a cream, lotion, serum or other topical composition to the skin may allow the polymer to penetrate deeper layers of the epidermis, increasing the overall amount of water retained by the skin. This may result in swelling of the skin and an overall reduction in the appearance of wrinkles. Water-retaining polymers may also be used in a number of other personal care products where additional water retention is useful to counter the drying effects of other active ingredients or to create formulations that provide additional moisturization and / or value to the consumer.

[0136] In some embodiments, a method for treating or preventing a skin condition is provided, comprising applying a topical composition comprising a biopolymer provided herein. In some embodiments, the skin condition is one or more of dry skin, wrinkled skin, sagging skin, aged skin, scarred skin, wounded skin, blemished skin, acne, and / or sun-damaged skin. In some embodiments, the skin condition is an inflammatory skin condition, such as psoriasis, eczema, or atopic dermatitis. In some embodiments, the skin condition is wounded skin. In some embodiments, the topical composition is applied to the face, ears, forehead, neck, arms, upper chest, legs, feet, and / or hands of a subject.

[0137] In various embodiments, application of the topical composition reduces transepidermal water loss, reverses transepidermal water loss, improves epidermal water retention, reduces the appearance of wrinkles, reduces sagging skin, increases skin smoothness, increases skin swelling, improves skin suppleness, improves skin texture, reduces skin blemishes, and / or reduces skin dryness. In some embodiments, the improvement is determined at a second time point compared to a first time point immediately prior to the first application of the topical composition. In some embodiments, the topical composition is applied three times a day, twice a day, once a day, once every two days, once every three days, or once a week. In some embodiments, the topical composition is applied for one week, two weeks, one month, two months, three months, six months, one year, or more. In some embodiments, the improvement is evident after using the topical composition for two weeks, one month, two months, three months, six months, or one year, or more. EXAMPLES

[0138] Example 1. Natural biopolymers produced by S. meliloti and S. fredii The naturally occurring biopolymers produced by selected Rhizobiaceae strains are shown in Figure 2. Succinoglycan, also called EPSI, is produced by Sinorhizobium meliloti. The repeating unit consists of a linear backbone of one galactose and three glucose monosaccharides with four glucose molecules on the side. The backbone sugars are linked by β-1,3 and β-1,4 glycosidic bonds. The side chain sugars are linked by β-1,3 and β-1,6 glycosidic bonds. Succinoglycan is acetylated at glucose 3 of the backbone, succinylated at glucose 3 of the side chain, and pyruvylated at the terminal glucose of the side chain (Reuber 1993). Galactoglucan, also called EPSII, is produced by S. meliloti. Galactoglucan is a linear repeating dimer of galactose and glucose linked by β-1,3 and α-1,3 glycosidic bonds (Glazebrook 1989). The galactose residues are fully pyruvylated and approximately 70% of the glucose units are acetylated (Her 1990). Glucuronoglycan is produced by Sinorhizobium fredii. Glucuronoglycan consists of a main chain composed of one galactose and five glucose monosaccharides linked by β-1,3, β-1,4 and β-1,6 glycosidic bonds, and two glucuronic acid and terminal galactose side chains linked by α-1,3, α-1,4 and β-1,4 glycosidic bonds. The terminal galactose on the side chain is acetylated (at carbon 2 and / or carbon 3) and 4,6-pyruvylated (Djordjevic 1986). Glucuronoglycans may be acetylated at a third, unknown position (Rodriquez-Navarro 2014).

[0139] Example 2. Strain construction For targeted deletion of selected ORFs, excision of insertion elements or correction of SNPs, non-replicative plasmid vectors with positive and negative selection markers were used. Both S. meliloti and S. fredii are amenable to genetic modification using standard molecular biology and strain engineering techniques, making it possible to rapidly and precisely alter their genomes and generate the desired genotypes. First, derivatives of the pJQ200SK plasmid (Quandt 1993) carrying deletion cassettes were generated. For deletion cassettes, regions (usually 500 bp) upstream and downstream of the target ORF, including the start and stop codons, were amplified by PCR. For introduction of wild-type DNA, regions upstream and downstream of the insertion element were amplified by PCR. Plasmids were then assembled using the CPEC method (Quan 2009) and sequences were verified before introduction into S. meliloti. Plasmids were introduced into S. meliloti by triparental mating, and strains containing a single integration in the homologous genomic region were selected for antibiotic resistance and verified by PCR with primers outside the amplified region. Strains positive for integration of the plasmid were then streaked, purified, and selected for the ability to grow on sucrose. The presence of the sacB gene on the integrated pJQ200 plasmid causes lethality when the strain is grown on sucrose. Thus, strains that grow on sucrose will either have a mutation in sacB itself, or have recombined to "loop out" the integrated plasmid and revert to wild type, or will have deleted or altered sequences that were originally present in the plasmid.

[0140] Domesticated strains of S. meliloti, such as strain Rm1021, have lost the ability to produce galactoglucan (Pellock 2002), and there are several genetic modifications that can be introduced to restore this function. These modifications include deletion of the mucR gene, restoration of the wild-type allele expR101 to the expR locus (Gonzalez 1996), or introduction of the wild-type expR ORF and promoter (Charoenpanich 2015). All these changes result in Rm1021-derived strains that produce galactoglucan in addition to succinoglycan. In this study, we introduced wild-type expR into strain Rm1021 using the methods described above. This resulted in strain EXO3, which produces both biopolymers of S. meliloti simultaneously.

[0141] Using the EXO3 strain, derivatives were generated that produce either succinoglycan or galactoglucan alone by deleting the ORFs known to be involved in the biosynthesis of either exopolysaccharide. For example, a strain that produces succinoglycan can be generated by deleting any of several glycosyltransferases, including wgaB or wgeB, which are involved in the synthesis of galactoglucan (Becker 1997). A strain that produces galactoglucan can be generated by deleting any of several glycosyltransferases, such as exoF, exoA, or exoY, which are involved in the early steps of succinoglycan biosynthesis (Gonzalez 1996, Glazebrook 1989). Using the above methodology, the wgeB ORF was deleted to generate an EXO3 derivative capable of producing only succinoglycan. To generate a galactoglucan-producing strain, the exoY ORF was deleted in EXO3. These strains, EXO1 and EXO2 (Table 1), were subsequently used for the production of succinoglycan or galactoglucan, respectively.

[0142] Using the targeted deletion methods described above, strains producing mutant biopolymers lacking chemical modifications can be generated. For example, exoH and exoZ, genes involved in succinylation and acetylation of succinoglycan, can be deleted from the S. meliloti genome. To produce modified galactoglucans, wgaE, a gene involved in pyruvylation, was excised from S. meliloti.

[0143] [Table 1]

[0144] Example 3. Production and purification of biopolymers For bench-scale growth and biopolymer production, batch cultures in shake flasks were used. The production strains were inoculated from culture plates into TY medium and grown overnight in a shaking incubator at 30°C. The next day, the overnight cultures were diluted into production medium, typically at a ratio of 1:100 or 1:200. The production medium consisted of a defined minimal medium such as M9, containing a carbon source of either glucose or sucrose at a concentration of 2%-4% (w / v), a nitrogen source such as ammonium sulfate, a buffer to maintain neutral pH, divalent cations such as MgSO4 and CaCl2, trace elements, and vitamins (US Pat. No. 7,371,558). Strains were grown in production medium for up to 3 days and then harvested for purification.

[0145] Recovery and purification of the biopolymers was performed by alcohol precipitation after initial cell separation. For high molecular weight polymers, the culture was diluted with either 2 or 3 volumes of water and the supernatant was separated from the cells by centrifugation. For low molecular weight biopolymers, it was not necessary to dilute the culture broth before cell separation. Approximately 1 mM CaCl2 was then added to the supernatant and the biopolymer was precipitated at room temperature by the addition of 2 volumes of isopropyl alcohol. The precipitate was isolated by centrifugation and then washed with either 70% or 90% ethyl alcohol. After the washing step, the precipitate was again isolated by low speed centrifugation and dried overnight in a 60°C oven until the weight loss stabilized, indicating no residual water. The final product was then ground using a mortar and pestle or using a bench scale mill.

[0146] Example 4. Characterization of biopolymers Biopolymers are characterized by analytical methods. In some cases, NMR spectroscopy can be used to determine structural information about composition, sequence distribution, substitution pattern, and molecular weight. Biopolymers may be assayed by solution NMR or solid-state NMR. For solution NMR of polysaccharides, samples may be subjected to enzymatic digestion or pretreatment due to the high viscosity of the material (Her et al. 1990). 13 Samples can be assayed without pretreatment using solid-state methods such as C cross-polarization magic angle spinning (CPMAS) NMR (Schaefer 1976). More detailed structural analysis and / or quantification can be assessed by 2D NMR, for example as described in Yao 2021. Those skilled in the art will appreciate that each analytical method has distinct advantages and disadvantages and can select the appropriate analytical method to generate the desired information regarding the structure, degree of modification, and / or purity level of the biopolymer. These methods can be used to quantify the degree of modification of sugars in polysaccharide chains. For example, the level of acetylation, pyruvylation, succinylation or other modified chemical groups can be determined for a sample of biopolymer.

[0147] Example 5. Water absorption rate To determine the water binding capacity of the biopolymers, samples were placed in a closed humid chamber for 5 days and the mass gain was measured in comparison to hyaluronic acid. The biopolymers were purified according to the procedure in Example 3. Before performing the water absorption experiments, the biopolymer samples were dried at 60°C for 30 minutes to ensure that all residual water had evaporated. Small amounts (usually 25mg to 50mg) were then weighed (value m0) and placed in individual tared plastic or aluminum trays. All samples were placed on a platform in a closed plastic chamber containing 250ml of warm water (approximately 35°C). The entire chamber with all samples was then placed in an incubator at 30°C. After 5 days, the humid chamber was opened, the individual samples were weighed (value m) and the mass gain was calculated. The water binding capacity (WBC) of each sample was calculated according to the following formula: (m-m0) / m0. This raw value represents the degree of swelling and can be expressed as a percentage mass gain by multiplying by 100.

[0148] A pure 5 kDa preparation of hyaluronic acid (HAworks) was used as a control for water binding experiments. WBC for hyaluronic acid measured as above was typically 200%-300%. Higher molecular weight HA (100 kDa, HAworks and >1000 kDa, Acros Organics) was tested and showed similar water uptake capacity as the lower molecular weight samples. To calculate the fold change for experimental samples, WBC values ​​for biopolymer mass gain were normalized to the WBC value for HA within the experiment.

[0149] Figure 3 shows that the WBC of isolated galactoglucan was increased by 3.5-fold compared to HA. This WBC value was reproducible across multiple experiments. The raw percent mass increase was 650% for galactoglucan, considerably higher than the 185% increase measured for HA. After further incubation in a humidified chamber, the mass increase of galactoglucan reached 720% of its initial mass. Results were similar when galactoglucan was derived from multiple carbon sources including glucose, sucrose, and corn syrup. Glucuronoglycan isolated from S. fredii showed a 1.7-fold increase in water binding compared to HA. The raw value of the percent mass increase for glucuronoglycan was 311%. Isolated succinoglycan, in contrast to galactoglucan and glucuronoglycan, showed reduced water binding compared to HA (0.7-fold reduction or 129% increase in raw mass), and the mixture of succinoglycan and galactoglucan was further reduced (0.3-fold reduction or 60% increase in raw mass). In the case of galactoglucan, improvements in WBC were only observed when galactoglucan was purified independently, and not as a naturally occurring mixture of both galactoglucan and succinoglycan derived from S. meliloti.

[0150] Although the type and content of monosaccharides, chemical modifications, glycosidic linkages, and molecular weight can all affect the behavior of biopolymers, the degree of negative charge appears to be the dominant factor in WBCs for biopolymers from these Rhizobiaceae species. Figure 4 shows that there is a correlation between negative charge and water binding ability for S. meliloti and S. fredii biopolymers. For the x-axis values, the negative charge of each repeating unit was calculated based on the pKa of the chemical group or sugar acid at physiological pH. For example, the two glucuronic acid and pyruvate modifications of glucuronoglycan each result in one negative charge at neutral pH, resulting in a ratio of negative charge to total sugar in the repeating unit of 1:3, or 0.33. For galactoglucan, the ratio is 1:2, and for succinoglycan, the ratio is 1:4. These ratios were plotted against the percent mass gain values ​​for each biopolymer. As shown in Figure 4, galactoglucan, the molecule with the highest ratio of negative charges to monosaccharides in the repeating unit, had the highest water binding capacity. Other rhizobia biopolymers did not show a high R 2 The fit was exact with values ​​of 0.001 and 0.001. Hyaluronic acid (grey circle), which also has a 1:2 ratio of charge to monosaccharides, did not fit the fitted curve, indicating that something other than or in addition to the charge ratio affects water retention for this molecule. Xanthan gum (Modernist Pantry) showed a lower water binding capacity compared to HA and similarly did not fit the fitted curve.

[0151] Figure 5 shows the structure of a nonpyruvylated galactoglucan molecule (NP-galactoglucan) derived from a S. meliloti strain in which the wgaE gene was excised. The NP-galactoglucan molecule also retained more water than the HA control, although not as much as the fully pyruvylated galactoglucan. Compared to HA, the NP-galactoglucan molecule showed a 1.7-fold increase in its ability to bind water.

[0152] Example 6. Cytotoxicity To test for cytotoxicity, biopolymers were purified according to Example 3 and resuspended in Ca, Mg-free PBS solution at a concentration of 1% (w / v). These solutions were then heat sterilized in a water bath at 60° C. for 30 minutes. The following cytotoxicity assays were performed at Pacific Biolabs in Hercules, CA.

[0153] Test Procedure: Total surface area 1.0 cm 2 A sterile filter paper with a flat surface of 10 cm is saturated with approximately 0.1 mL of the test solution and 2 The test articles were placed directly on the cell culture monolayer in the center of the wells. Triplicate preparations were prepared. Triplicate positive and negative controls were tested as well as the test articles. All wells were incubated for at least 24 hours at 37±1°C in a humidified incubator with 5±1% CO2. After incubation, the test articles and controls were gently removed from the wells. The cell cultures were tested for cytotoxic response under an inverted microscope at 100x magnification. The response was graded on a scale of 0-4. Achieving a grade number greater than 2 is considered a cytotoxic effect.

[0154] The study was performed according to ISO 10993-5:2009. A value of 0 is considered no reactivity and a value of 1 is considered slightly reactive. Figure 6 shows that isolated succinoglycan, galactoglucan, glucuronoglycan, and a mixture of succinoglycan and galactoglucan are not cytotoxic by this assay.

[0155] Example 7. Topical formulations for skin care Biopolymers and / or their derivatives are produced and purified according to Example 3. The purified biopolymers are added to the following commercially available lotion bases: Dermabase Cream™ (Paddock Laboratories), Crafter's Choice™ Basic Lotion Base (Crafter's Choice Brands LLC) and Stephenson Easy Lotion Base (Stephenson), or other similar bases. These lotion bases are a variety of room temperature stable oil-in-water emulsions for skin care. The biopolymers are added directly to the lotion bases or resuspended in water or PBS (Ca, Mg free) prior to addition to achieve the appropriate final concentration. Typical formulations include a final concentration of 0.1%-2% (w / v) of biopolymer or derivative.

[0156] Lotion Base Ingredients: Dermabase Cream™ - Purified water, mineral oil, petrolatum, cetostearyl alcohol, propylene glycol, sodium lauryl sulfate, isopropyl palmitate, imidazolidinyl urea, methylparaben and propylparaben. Crafter's Choice™ Basic Lotion Base - Water, Cetearyl Alcohol, Glyceryl Stearate, Polysorbate-60, Stearyl Alcohol, Sunflower Oil, Allantoin, Propylene Glycol, Isopropyl Palmitate, Petrolatum, Diazolidinyl Urea, Methylparaben, Propylparaben, BHT, Carbomer, Benzophenone-4, Stearic Acid. Stephenson Easy Lotion Base - Water, Sunflower Seed Oil, Polysorbate 20, Cetearyl Alcohol, Glyceryl Stearate, Phenoxyethanol, Carbomer, Potassium Sorbate, Sodium Hydroxide.

[0157] For skin hydration testing of biopolymers, the following oil-in-water base lotion was used: distilled water, Papaver Somniferum (Poppy Seed) Seed Oil, glycerin, pentylene glycol (vegetable derived), olive squalane, polyglyceryl-2 stearate, glyceryl stearate, stearyl alcohol, tocopherol, Lonicera Japonica (Honeysuckle) Flower Extract, Lonicera Caprifolium (Honeysuckle) Flower Extract, 0.2% (w / v) Galactoglucan, Gum Arabic, Xanthan Gum, Sodium Phytate, Alcohol, and Lactic Acid.

[0158] Example 8. In vivo skin water retention study The biopolymer-containing lotion-based formulations described in Example 7 are used in a comparative in vivo study of skin moisture along with a placebo composition. Each lotion-based placebo composition containing the biopolymer is generated by adding an equal volume of PBS or sterile water.

[0159] In the skin moisture retention study, subjects are instructed to apply a standard amount of lotion base containing biopolymers to the right side of the face and a placebo composition to the left side of the face. Subjects are instructed to always apply the same composition to the same side of the face. Subjects are instructed to apply these compositions twice a day for the specified times and to refrain from using any other topical compositions unrelated to the study composition. Skin moisture retention measurements are performed using a Corneometer 825 (Courage and Khazaka).

[0160] Galactoglucan was purified according to the procedure described above and used in the skin water retention study. Skin water retention was measured over a 48-hour period using poppy seed oil formulations as described in Example 7, containing either 0.2% (w / v) galactoglucan or an equal amount of water as a control. Baseline skin water retention measurements (arbitrary units with Corneometer 825) for two healthy volunteers, a 47-year-old man (P1) and a 49-year-old woman (P2), were taken before the start of the study. In this study, Corneometer measurements in arbitrary units typically range from 20 to 50, with the maximum value of the instrument being 120. Participants were instructed to apply approximately 0.3 ml to two different sites on the body, namely the back of the hand and the elbow. Test and control samples were applied four times per day, and changes in skin water retention were monitored before each application. Figure 7 shows the mean change in water retention from baseline for each site on the body for measurements taken after 24 hours of treatment. Treatment with the lotion base containing galactoglucan resulted in a consistent increase in water retention values ​​at both body sites for both P1 and P2 compared to the lotion base control. Of note, the water retention trend was well established 24 hours after treatment, suggesting that the water retention effect was cumulative and that longer treatments would result in even greater differences in water retention between the lotion containing galactoglucan and the control. Figure 8 shows an image of the back of participant P1's hand 36 hours after treatment, where a clear reduction in the appearance of wrinkles can be observed on the hand treated with galactoglucan compared to the hand treated with the control lotion base. The circles highlight the areas where the reduction in the appearance of wrinkles is particularly evident. Participants did not report any redness, irritation, swelling, sensitivity or discomfort with either the experimental or control treatments.

[0161] Example 9. In vitro percutaneous absorption of biopolymers Ex vivo skin studies are used to assess skin penetration, rank biopolymers in terms of permeability or accumulation, and optimize formulations for efficacy. Studies are performed in Franz diffusion cells (Franz 1975) using human or porcine skin to assess percutaneous absorption. Biopolymer formulations are applied to the upper (external) surface of the Franz cell, and samples are removed and measured at predefined time points from a buffer-containing reservoir in contact with the lower (serosal) surface. Control polymers or other compounds are either co-administered (if the test biopolymer is in solution) or run in parallel (if the test biopolymer is in some other type of formulation) for quality control. Skin may be excised at the end of the study to quantify accumulation of the test compound.

[0162] Franz cell assays were performed with non-pyruvylated galactoglucan samples. Specifically, human cadaveric skin excised from a single donor was measured for thickness and then mounted in a Franz vertical diffusion cell (FDC-6, Logan instruments, Somerset, NJ) thermostatically controlled at 37°C. Receptor solution (PBS, pH 7.4) was added and equilibrated at 37°C for 30 min (to reach a skin surface temperature of 32°C). After the equilibration period, the entire volume of receptor solution (approximately 11 ml) was removed and discarded, and replaced with fresh pre-warmed receptor solution to remove endogenous background released into the buffer before the start of the experiment.

[0163] Dosing solutions were prepared by adding 5% (w / v) biopolymer to PBS (pH 7.4) and incubating at 50°C for 30 min. 1 ml of dosing solution was added to each donor chamber. Thus, the total amount of biopolymer in each chamber was 50 mg. Donor solutions were sampled at 0, 8, and 24 h. Receptor solutions were sampled at 0.5, 1, 2, 4, 8, and 24 h. After 24 h, tissues were removed, briefly rinsed in blank PBS after the permeabilization period, weighed, and stored separately at -20°C. Biopolymer quantification used the Anthrone method (Morris 1948) with Carrez reagent (EMD Millipore) to reduce protein background in receptor samples. The Anthrone method is commonly used to detect the presence of carbohydrates in solutions. For receptor samples, 4 μL of Caretz Reagent I was added to 400 μL of sample in a 1.5 mL tube, and the tube was mixed by vortexing. An additional 4 μL of Caretz Reagent II was then added, and the tube was mixed again. The reagent was neutralized with 2.5 μL of 1 N NaOH, and the tube was centrifuged at 16000×g for 5 min. 625 μL of 0.2% anthrone in concentrated sulfuric acid was mixed with 325 μL of the supernatant on ice. The sample was heated at 99°C for 15 min, then cooled on ice.

[0164] To detect biopolymers retained in the skin, the removed samples were immersed in 1 mL of PBS in a 2 mL tube at 37°C for 4 h and then at 4°C for 3 days. The PBS was collected and centrifuged twice at 16,000 x g for 5 min to remove debris. Samples were clarified as above with Caretz's reagent and then diluted 10-fold in PBS.

[0165] The absorbance was measured at 620 nm for all samples and carbohydrate concentrations were determined by comparison with a standard curve developed using Karez's reagent. The background absorbance of the control Franz cell (no biopolymer) was calculated and subtracted from the values ​​to derive the carbohydrate concentration. Figure 9 shows representative results from the receptor chamber (permeate) after 4 hours of incubation. The carbohydrate detected in this sample indicates that the biopolymer had penetrated the epidermal layer. The highest concentration of biopolymer in the receptor chamber was usually observed at these time points. Figure 9 also shows that after immersing the removed sample in PBS as described above, a certain amount of biopolymer is contained or associated with the skin itself (skin). These findings indicate that a portion of the material in the donor chamber of the Franz cell has completely penetrated the skin sample and another portion is retained or associated with the skin sample. Without intending to be bound by any particular theory, it is possible that the skin-penetrating fraction of non-pyruvylated galactoglucan is the lowest molecular weight fraction in the sample, and that the higher molecular weight fractions are absorbed and / or retained at the epidermal surface.

Claims

1. 1. A biopolymer preparation comprising a biopolymer composed of repeating disaccharide units comprising glucose and galactose, wherein at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, or at least 90% of the glucose is acetylated and at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% of the galactose is pyruvylated.

2. 2. The biopolymer preparation of claim 1, wherein the glucose and galactose are linked by β-1,3 glycosidic and α-1,3 glycosidic bonds.

3. structure: 【Chemistry 1】 wherein the dotted line represents the bond between the disaccharide units, wherein no more than 40%, no more than 35%, no more than 30%, no more than 25%, no more than 20%, no more than 15%, no more than 10%, or no more than 5% of the disaccharide units lack an acetyl moiety and no more than 60%, no more than 55%, no more than 50%, no more than 45%, no more than 40%, no more than 35%, no more than 30%, no more than 25%, no more than 20%, no more than 15%, no more than 10%, no more than 5%, or no more than 1% of the disaccharide units lack a pyruvyl moiety.

4. The biopolymer preparation according to any one of claims 1 to 3, wherein the molar ratio of glucose:galactose:pyruvyl:acetyl in the biopolymer is 1:1:0.4-1:0.6-1.

5. 5. The biopolymer preparation of claim 1, wherein the average molecular weight of the biopolymers in the biopolymer preparation is less than 3000 kDa, less than 1000 kDa, less than 300 kDa, less than 100 kDa or less than 40 kDa, optionally the average molecular weight of the biopolymers in the biopolymer preparation is between 0.5 kDa and 40 kDa.

6. 6. The biopolymer preparation of any one of claims 1 to 5, wherein the biopolymer preparation is at least 75%, at least 80%, at least 85%, at least 90% biopolymer, or at least 95% (w / w) biopolymer.

7. 7. The biopolymer preparation of any one of claims 1 to 6, which is capable of absorbing at least the same amount of water, at least 1.5 times, at least 2 times or at least 3 times as much water as an equivalent amount of hyaluronic acid, and optionally is capable of absorbing an amount of water that is at least 100%, at least 200%, at least 300%, at least 400% or at least 500% of the initial dry weight of the biopolymer preparation, and optionally is capable of absorbing an amount of water that is at least 100%, at least 200%, at least 300%, at least 400% or at least 500% of the initial dry weight of the biopolymer preparation.

8. 8. The biopolymer preparation of any one of claims 1 to 7, wherein less than 25%, less than 20%, less than 15%, less than 10%, less than 5%, less than 3% or less than 1% of the biopolymer preparation is succinoglycan, or the biopolymer preparation is substantially free of succinoglycan.

9. 9. The biopolymer preparation of any one of claims 1 to 8, which is a solid or powder, and optionally has less than 5%, less than 4%, less than 3%, less than 2% or less than 1% of the biopolymer preparation being water.

10. 1. A biopolymer preparation comprising a biopolymer composed of repeating disaccharide units comprising glucose and galactose, wherein at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85% or at least 90% of the glucose is acetylated and less than 15%, less than 10% or less than 5% of the galactose is pyruvylated.

11. 11. The biopolymer preparation of claim 10, wherein the glucose and galactose are linked by β-1,3 glycosidic and α-1,3 glycosidic bonds.

12. 1. A biopolymer preparation, comprising a biopolymer having the structure: 【Chemistry 2】 (wherein the dotted line represents the bond between the disaccharide units), wherein no more than 40%, no more than 35%, no more than 30%, no more than 25%, no more than 20%, no more than 15%, no more than 10%, or no more than 5% of the disaccharide units are lacking an acetyl moiety, and no more than 15%, no more than 10%, or no more than 5% of the disaccharide units are pyruvylated.

13. 13. The biopolymer preparation of any one of claims 10 to 12, wherein the molar ratio of glucose:galactose:pyruvyl:acetyl in the biopolymer is 1:1:<0.5:0.6-1.

14. 14. The biopolymer preparation of any one of claims 10 to 13, wherein the average molecular weight of the biopolymers in the biopolymer preparation is less than 3000 kDa, less than 1000 kDa, less than 300 kDa, less than 100 kDa or less than 40 kDa, optionally wherein the average molecular weight of the biopolymers in the biopolymer preparation is between 0.5 kDa and 40 kDa.

15. A biopolymer preparation according to any one of claims 10 to 14, wherein at least 75%, at least 80%, at least 85%, at least 90% or at least 95% (w / w) of the biopolymer preparation is biopolymer.

16. 16. The biopolymer preparation of any one of claims 10 to 15, capable of absorbing at least the same, at least 1.1 times, at least 1.2 times, at least 1.3 times, at least 1.4 times or at least 1.5 times as much water as an equivalent amount of hyaluronic acid, and optionally capable of absorbing an amount of water that is at least 100%, at least 200%, at least 300% or at least 400% of the initial dry weight of the biopolymer preparation, optionally wherein the water absorption is measured by placing a dried sample of the biopolymer preparation in a humid chamber at 30°C for 5 days.

17. 17. The biopolymer preparation of any one of claims 10 to 16, wherein less than 25%, less than 20%, less than 15%, less than 10%, less than 5%, less than 3%, or less than 1% of the biopolymer preparation is succinoglycan.

18. 18. A biopolymer preparation according to any one of claims 10 to 17, which is a solid or powder, and optionally has less than 5%, less than 4%, less than 3%, less than 2% or less than 1% of the biopolymer preparation being water.

19. 1. A biopolymer preparation comprising a biopolymer composed of repeating polysaccharide units, each polysaccharide unit comprising 2 to 15, or 2 to 12, or 2 to 10 monosaccharides, and wherein the negative charge:monosaccharide ratio in said repeating polysaccharide units of said biopolymer is at least 0.3, or at least 0.35, or at least 0.4, or at least 0.

45.

20. 20. The biopolymer preparation of claim 19, wherein the polysaccharide units comprise at least one galactose linked to at least one glucose, and optionally, at least one galactose is linked to the glucose via a β-1,3 glycosidic bond.

21. 21. The biopolymer preparation of claim 19 or 20, wherein the average molecular weight of the biopolymers in the biopolymer preparation is less than 3000 kDa, less than 1000 kDa, less than 300 kDa, less than 100 kDa or less than 40 kDa, optionally wherein the average molecular weight of the biopolymers in the biopolymer preparation is between 0.5 kDa and 40 kDa.

22. 22. A biopolymer preparation according to any one of claims 19 to 21, wherein at least 75%, at least 80%, at least 85%, at least 90% or at least 95% (w / w) of the biopolymer preparation is biopolymer.

23. 23. The biopolymer preparation of any one of claims 19 to 22, capable of absorbing at least the same, at least 1.1 times, at least 1.5 times, at least 2 times, or at least 3 times as much water as an equivalent amount of hyaluronic acid, and optionally capable of absorbing an amount of water that is at least 100%, at least 200%, at least 300% or at least 400% of the initial dry weight of the biopolymer preparation, optionally wherein the water absorption is measured by placing a dried sample of the biopolymer preparation in a humid chamber at 30°C for 5 days.

24. 24. The biopolymer preparation of any one of claims 19 to 23, which is a solid or powder, and optionally has less than 5%, less than 4%, less than 3%, less than 2% or less than 1% of the biopolymer preparation being water.

25. A topical composition comprising a biopolymer, comprising the biopolymer preparation of any one of claims 1 to 24.

26. 26. The topical composition of claim 25, comprising 0.01% to 10% (w / w) or 0.05% to 5% (w / w) of said biopolymer.

27. 27. The topical composition of claim 25 or 26, comprising one or more thickeners, stabilizers, emulsifiers, emollients, moisturizers, rheology modifiers, film formers, antioxidants, additives, actives, butters, essential oils, exfoliated oils, clays, muds, extracts, hydrosols, exfoliants, supplements, waxes, thickeners, salts, minerals, acids, bases, carrier oils, fixed oils, surfactants, preservatives, pearlizing agents, conditioning agents, structuring agents, brighteners, humectants, osmolytes, occlusive agents, cleansing agents, colorants, pigments, fragrances, UV-A blockers, UV-B blockers, and / or nutrients.

28. A topical composition according to any one of claims 25 to 27, comprising hyaluronic acid.

29. The topical composition according to any one of claims 25 to 27, which is free of hyaluronic acid.

30. 30. The topical composition of any one of claims 25 to 29, which is a cream, lotion, serum, ointment, balm, tincture, liniment, shampoo, soap, conditioner, sunscreen, rinse, deodorant, or cosmetic.

31. A topical composition according to any one of claims 25 to 30 for treating or preventing a skin condition in a subject, optionally wherein the skin condition is one or more of dry skin, wrinkled skin, sagging skin, aged skin, flawed skin, injured skin, blemished skin, acne and / or sun damaged skin, optionally wherein the topical composition is applied to the face, ears, forehead, neck, arms, upper chest, legs, feet and / or hands of the subject, and optionally wherein the topical composition reduces transepidermal water loss, reverses transepidermal water loss, improves epidermal water retention, reduces the appearance of wrinkles, reduces skin sagging, increases skin smoothness, increases skin swelling, improves skin suppleness, improves skin texture, reduces skin blemishes and / or reduces skin dryness.

32. 32. The topical composition of claim 31, wherein the skin condition is an inflammatory skin condition, optionally wherein the inflammatory skin condition is psoriasis, eczema, or atopic dermatitis.

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