Use of enzymes for replacing whiteness retention agents in cleaning compositions

Abstract

Use of an enzyme for replacing a whiteness retention agent in a cleaning composition is disclosed, wherein the enzyme is selected from the group consisting of a protease, a mannanase, a cellulase, and combinations thereof.

Description

[0001] References to sequence listings

[0002] This application contains a Sequence Listing in computer readable form, which is incorporated herein by reference. Technical Field

[0003] The present invention relates to the use of enzymes for replacing whiteness / color maintenance components (eg, optical brighteners) in cleaning compositions, wherein the enzymes are selected from the group consisting of proteases, mannanases, cellulases, and combinations thereof. Background Art

[0004] Textiles (e.g., clothes) become dull after being worn and / or washed. One reason is that the textiles lose whiteness / color during the washing process. In order to maintain the whiteness / color of the textiles, whiteness retainers, such as optical brighteners and anti-redeposition polymers, are usually added to laundry detergents. However, since these chemical ingredients come from non-renewable energy sources, they are not sustainable and may cause health or environmental problems. Therefore, it is desirable to provide an alternative detergent with improved sustainability characteristics while maintaining or even improving the key properties of the detergent (e.g., whiteness / color retention effect). Summary of the invention

[0005] The inventors of the present invention have surprisingly found that some enzymes can replace a large amount of whiteness / color maintenance agents (e.g., optical brighteners and / or anti-redeposition polymers) in detergent compositions without compromising the overall whiteness / color maintenance performance. The inventors have found that by such replacement, the whiteness / color of the treated textiles can even be improved.

[0006] In one aspect, the present invention relates to the use of an enzyme for replacing a whiteness maintenance agent in a cleaning composition, wherein the enzyme is selected from the group consisting of: a protease, a mannanase, a cellulase and a combination thereof, preferably, the enzyme is a combination of two or three enzymes selected from protease, mannanase and cellulase.

[0007] In another aspect, the present invention relates to a cleaning composition free of at least one whiteness maintenance agent, wherein the composition comprises an enzyme selected from the group consisting of: protease, mannanase, cellulase and combinations thereof, wherein the whiteness maintenance agent is selected from the group consisting of: optical brightener, anti-redeposition polymer and mixtures thereof. The cleaning composition provided by the present invention has improved sustainability characteristics because at least one whiteness maintenance agent can be replaced by an eco-friendly enzyme while maintaining or even improving the key properties of the composition (e.g., whiteness / color maintenance effect).

[0008] In a further aspect, the present invention relates to a method for maintaining or improving the whiteness and / or color of a textile, the method comprising: (a) exposing the textile to a wash liquor comprising a detergent composition of the present invention; and optionally, (b) rinsing the textile.

[0009] definition

[0010] As used herein, the singular forms “a”, “an” and “the” are intended to include the plural forms as well, unless the context clearly indicates otherwise.

[0011] If not indicated otherwise, all percentages (%) mentioned in the disclosed compositions relate to wt % relative to the total weight of the respective composition.

[0012] Unless defined otherwise or clearly indicated by the context, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.

[0013] Cleaning ingredients: The term "cleaning components" is defined herein to mean the types of chemicals that can be used in cleaning compositions. Examples of cleaning components are alkalis, surfactants, solvents, hydrotropes, builders, co-builders, chelators or chelating agents, bleach systems or bleach components, polymers, foam boosters, suds suppressors, dispersants, dye transfer inhibitors, perfumes, bactericides, fungicides, corrosion inhibitors, soil suspending agents, soil release polymers, whiteness / color maintenance agents (such as optical brighteners and anti-redeposition agents), enzyme inhibitors or stabilizers, enzyme activators, antioxidants, preservatives, solvents and solubilizing agents.

[0014] Whiteness / color maintenance agent: The term "whiteness / color retainer" refers to an additive used in a detergent that can help maintain the original whiteness / color of a fabric, or prevent / delay the loss of whiteness / color, by, for example, preventing dirt (e.g., soluble dirt and insoluble dirt such as clay particles and microfibers) from being redeposited on the fabric after being removed during the washing process, or by masking / shielding the loss of whiteness / color to make the textile appear whiter and brighter. These agents typically include, but are not limited to, optical brighteners and anti-redeposition agents as shown below. In the present invention, the term "whiteness / color retainer" can be used interchangeably with the term "whiteness retainer".

[0015] Optical brightenerAs used herein, the term optical brightener is used interchangeably with the term optical brightener, fluorescent brightener, or fluorescent whitening agent. These additives are typically used to enhance the appearance of color and / or brightness of materials such as textiles and paper, making the material look less yellow / less dull, thereby maintaining the original whiteness and color of the material for a longer period of time.

[0016] Anti-redeposition agents : In the context of the present invention, the term "anti-redeposition agent" refers to an additive used in detergents that helps prevent soil from being redeposited on fabrics after being removed during the washing process. Suitable anti-redeposition agents may include, but are not limited to, anti-redeposition polymers such as polyacrylic acid, modified polyacrylic acid polymers, modified polyacrylic acid copolymers, maleic acid-acrylic acid copolymers, carboxymethyl cellulose (CMC), cellulose gum, methyl cellulose, polyvinyl alcohol (PVA), polyvinyl pyrrolidone (PVP), polyoxyethylene and / or polyethylene glycol (PEG), homopolymers of acrylic acid, copolymers of acrylic acid and maleic acid, ethoxylated polyethyleneimine, and / or combinations thereof. Cellulose-based polymers described below under soil release polymers may also be used as anti-redeposition agents.

[0017] Cleaning composition : The term "cleaning composition (also referred to as "detergent composition")" refers to a composition for removing undesirable compounds from an article to be cleaned. Cleaning compositions can be used, for example, for household cleaning and industrial cleaning. These terms encompass any material / compound and product form (e.g., liquid, gel, powder, granular, paste or spray composition) selected for a particular type of cleaning composition desired. In addition to containing the enzymes of the present invention, the cleaning compositions of the present invention may also contain one or more additional enzymes (e.g., lipases, amylases, hemicellulases, peroxidases, xylanases, phospholipases, esterases, cutinases, keratinases, reductases, oxidases, phenoloxidases, lipoxygenases, ligninases, pullulanases, malanases, β-glucanases, arabinosidases, hyaluronidases, laccases, deoxyribonucleases (DNA enzymes), hexosaminidase and any mixtures thereof) and / or one or more cleaning components as described above.

[0018] Mature peptide : The term "mature polypeptide" means a polypeptide in its final form after translation and any post-translational modifications such as N-terminal processing, C-terminal truncation, glycosylation, phosphorylation and the like.

[0019] It is known in the art that a host cell can produce a mixture of two or more different mature polypeptides (i.e., having different C-terminal and / or N-terminal amino acids) expressed by the same polynucleotide. It is also known in the art that different host cells process polypeptides in different ways, so a host cell expressing a polynucleotide can produce different mature polypeptides (e.g., having different C-terminal and / or N-terminal amino acids) when compared to another host cell expressing the same polynucleotide.

[0020] Sequence identity : The relatedness between two amino acid sequences or between two nucleotide sequences is described by the parameter "sequence identity".

[0021] For purposes of the present invention, the Needleman-Wunsch algorithm (Needleman and Wunsch, 1970, J.Mol.Biol. [Journal of Molecular Biology] 48: 443-453) is used to determine the sequence identity between two amino acid sequences as the output of "longest identity", which is implemented by the Needleman program of the EMBOSS software package (EMBOSS: The European Molecular Biology Open Software Suite [European Molecular Biology Open Software Suite], Rice et al., 2000, Trends Genet. [Genetics Trend] 16: 276-277) (preferred 6.6.0 version or updated version). The parameters used are gap opening penalty 10, gap extension penalty 0.5 and EBLOSUM62 (EMBOSS version of BLOSUM62) substitution matrix. In order to make the Needleman program report the longest identity, it is necessary to specify the non-simplified option in the command line. The output of the "longest identity" marked by Needleman is calculated as follows:

[0022] (number of identical residues x 100) / (length of alignment - total number of gaps in the alignment)

[0023] Variants: The term "variant" means a polypeptide / enzyme having the same type of activity as a parent enzyme and comprising changes, i.e., substitutions, insertions and / or deletions, at one or more positions compared to the amino acid sequence of the parent. Substitution means replacing the amino acid occupying a certain position with a different amino acid; deletion means removing the amino acid occupying a certain position; and insertion means adding an amino acid adjacent to and immediately following the amino acid occupying a certain position.

[0024] In the context of the present invention, a variant may for example be a variant of an identified protease having the activity of the parent enzyme.

[0025] Variant nomenclature: In describing the enzyme variants herein, the nomenclature described below is used for ease of reference. Generally accepted IUPAC single-letter amino acid abbreviations are employed.

[0026] replace :For amino acid substitution, the following nomenclature is used: original amino acid, position, substituted amino acid. Accordingly, the substitution of threonine at position 226 by alanine is represented as "T226A". Multiple mutations can be separated by a plus sign ("+"), such as "G205R+S411F", or separated by a comma, such as "G205R, S411F", representing that glycine (G) and serine (S) at position 205 and position 411 are replaced by arginine (R) and phenylalanine (F), respectively.

[0027] Missing : For amino acid deletions, the following nomenclature is used: original amino acid, position, * Accordingly, the deletion of glycine at position 195 is indicated as "G195*".

[0028] insert : For amino acid insertions, the following nomenclature is used: original amino acid, position, original amino acid, inserted amino acid. Accordingly, the insertion of lysine after the glycine at position 195 is indicated as "G195GK". Indications of insertion at a particular position are understood to be insertions after the original amino acid residue. For example, "insertion at position 195" is understood to be insertions after the original residue at position 195.

[0029] Multiple changes : Variants containing multiple alterations are separated by plus signs ("+") or commas, for example, "R170Y+G195E" or "R170Y,G195E", representing substitution of arginine and glycine at positions 170 and 195 with tyrosine and glutamic acid, respectively.

[0030] Different changes : When different changes can be introduced at one position, the different changes can be separated by commas, for example, "R170Y,E" represents that the arginine at position 170 is replaced by tyrosine or glutamic acid. Therefore, "Y167G,A+R170G,A" represents the following variants: "Y167G+R170G", "Y167G+R170A", "Y167A+R170G", and "Y167A+R170A".

[0031] textile: The term "textile" means any textile material, including yarn, yarn intermediates, fibers, nonwoven materials, natural materials, synthetic materials, and any other textile materials, fabrics made from these materials, and products made from fabrics (e.g., clothing and other articles). The textile or fabric can be in the form of knitwear, woven fabrics, denim, nonwoven fabrics, felt, yarn, and terry cloth. The textile can be cellulose-based, such as natural cellulose, including cotton, flax / linen, jute, ramie, sisal, or coconut shell fiber; or artificial cellulose (e.g., derived from wood pulp), including viscose / rayon, ramie, cellulose acetate fibers (tricell), lyocell fibers, or blends thereof. The textile or fabric can also be based on non-cellulose, such as natural polyamides, including wool, camel hair, cashmere, mohair, rabbit hair and silk; or synthetic polymers, such as nylon, aromatic polyamide, polyester, acrylate, polypropylene and spandex / elastane; or its blends together with blends based on cellulose and based on non-cellulose fibers. Examples of blends are blends of cotton and / or rayon / viscose with one or more accompanying materials, such as wool, synthetic fibers (e.g., polyamide fibers, acrylic fibers, polyester fibers, polyvinyl alcohol fibers, polyvinyl chloride fibers, polyurethane fibers, polyurea fibers, aromatic polyamide fibers) and cellulose-containing fibers (e.g., rayon / viscose fibers, ramie, flax / linen, jute, cellulose acetate fibers, lyocell fibers). Fabrics can be conventional washable clothing, such as household clothing with stains. When the term fabric or clothing is used, it is intended to also include the broad term textiles.

[0032] Whiteness / color retention performance: The term is defined herein as a broad term with different meanings in different areas and for different customers. The loss of fabric whiteness / color can be attributed to graying, yellowing, or removal of optical brighteners / toners, for example. Graying and yellowing can be attributed to soil redeposition, body soil, coloring or dye transfer. In the present invention, the term "whiteness / color retention performance" can be used interchangeably with the term "whiteness retention effect" or "whiteness performance". For the purposes of the present invention, whiteness / color retention performance can be measured according to the ΔR460 value as described in the Examples section. BRIEF DESCRIPTION OF THE DRAWINGS

[0033] A more complete understanding of the invention may be obtained by referring to the detailed description and claims when considered in conjunction with the accompanying drawings.

[0034] Figure 1 Shown are pictures of CN-42 cotton textiles washed with detergents (with or without whiteness maintenance agents) in the presence or absence of an enzyme of the present invention, wherein:

[0035] Row 1, from left to right:

[0036] D1: Textiles washed with standard detergent 1;

[0037] D2: Textiles washed with standard detergent 1 + optical brightener (OB);

[0038] D3: Textiles washed with standard detergent 1 + optical brightener (OB) + anti-redeposition polymer;

[0039] D4: textiles washed with standard detergent 1 + cellulase B;

[0040] Row 2, from left to right:

[0041] D5: textiles washed with standard detergent 1 + cellulase A;

[0042] D6: textiles washed with standard detergent 1 + protease;

[0043] D7: textiles washed with standard detergent 1 + mannanase;

[0044] D8: textiles washed with standard detergent 1 + cellulase B + protease;

[0045] Row 3, from left to right:

[0046] D9: textiles washed with standard detergent 1 + cellulase B + mannanase;

[0047] D10: textiles washed with standard detergent 1 + cellulase B + protease + mannanase;

[0048] D11: textiles washed with standard detergent 1 + cellulase A + protease + mannanase;

[0049] D12: textiles washed with standard detergent 1 + cellulase A + protease;

[0050] D13: Textiles washed with Standard Detergent 1 + Cellulase A + Mannanase. DETAILED DESCRIPTION

[0051] The inventors have found that the reduction in R460 values ​​obtained for textiles treated with detergent compositions containing no whiteness / color maintenance agents (eg optical brighteners / anti-redeposition polymers) can be fully compensated by the addition of one or more enzymes of the present invention.

[0052] Therefore, one aspect of the present invention relates to the use of enzymes for replacing whiteness / color maintenance agents in cleaning compositions, wherein the enzyme is selected from the group consisting of proteases, mannanases, cellulases, and combinations thereof. By adding said one or more enzymes, the amount of whiteness / color maintenance agents can be reduced by at least 20%, e.g., at least 30%, at least 40%, at least 50%, or even 100%.

[0053] Accordingly, in one embodiment of the present invention, the amount of optical brightener is reduced by at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 75%, at least 80% or at least 90%.

[0054] In one embodiment of the invention, the amount of anti-redeposition polymer is reduced by at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 75%, at least 80% or at least 90%.

[0055] In one embodiment, both the optical brightener and the anti-redeposition polymer in the detergent composition are completely replaced by the one or more enzymes.

[0056] Another aspect of the present invention relates to a method for maintaining and / or improving the whiteness / color of a textile, wherein the textile is exposed to a wash liquor containing a detergent comprising an enzyme selected from the group consisting of: a protease, a mannanase, a cellulase, and a combination thereof. In one embodiment, the enzyme is a combination of a protease and a mannanase. In another embodiment, the enzyme is a combination of a protease and a cellulase. In yet another embodiment, the enzyme is a combination of a protease, a mannanase, and a cellulase.

[0057] Suitable amounts of one or more enzymes of the present invention may be present in an amount ranging from 0.0001 wt% to 3 wt% based on the total weight of the cleaning composition, for example, from 0.0003 wt% to 2 wt%, from 0.005 wt% to 1 wt%, from 0.05 wt% to 1.5 wt% or from 0.1 wt% to 0.5 wt% based on the total weight of the cleaning composition.

[0058] In another embodiment, the amount of one or more enzymes of the present invention ranges from 0.0001 mg to 100 mg enzyme protein per liter of wash liquor, for example 0.01 mg to 50 mg enzyme protein, 0.1 mg to 30 mg enzyme protein, 0.2 mg to 10 mg enzyme protein or 1 mg to 5 mg enzyme protein per liter of wash liquor.

[0059] In one embodiment of the present invention, the optical brightener is selected from the group comprising: benzenesulfonic acid, 2,2'-(1,2-ethylenediyl)bis[5-[4-(4-morpholinyl)-6-(phenylamino)-1,3,5-triazin-2-yl]amino]-, disodium salt; 2,2'-([1,1'-biphenyl]-4,4'-diyldi-2,1-ethylenediyl)bis-, disodium salt; diaminostilbene-sulfonic acid derivatives, including the following sodium salts: 4,4'-bis-(2-diethanolamino-4-anilino-s-triazin-6-ylamino)stilbene-2,2'-disulfonate; 4,4'-bis-(2,4-diphenylamino-s-triazin-6-ylamino)stilbene-2.2'-disulfonate; 4,4'-bis-(2-anilino-4(N-methyl-N-2-hydroxy-ethylamino)stilbene-2.2'-disulfonate. 4,4'-bis-(2-anilino-4(1-methyl-2-hydroxy-ethylamino)-s-triazin-6-ylamino)stilbene-2,2'-disulfonate, 4,4'-bis-(4-phenyl-2,1,3-triazol-2-yl)stilbene-2,2'-disulfonate; 4,4'-bis-(2-anilino-4(1-methyl-2-hydroxy-ethylamino)-s-triazin-6-ylamino)stilbene-2,2'-disulfonate, and / or 2-(stilbene-4"-naphthalene-1.,2':4,5)-1,2,3-triazole-2"-sulfonate; or 4,4'-bis-(2-morpholino-4-anilino-s-triazin-6-ylamino)stilbene disulfonate or 2,2'-bis-(phenyl-stilbene) disulfonate, or 4,4'-bis-(sulfostyryl)-biphenyl disodium salt, bistriazinylstilbene derivatives, and mixtures thereof.

[0060] In one embodiment, the optical brightener is 4,4'-bis-(sulfostyryl)-biphenyl disodium salt or a bistriazinylstilbene derivative.

[0061] In other embodiments of the present invention, the preferred optical brightener is Tinopal DMS or Tinopal CBS or Tinopal CBS-X available from BASF. Tinopal DMS is the disodium salt of 4,4'-bis-((2-morpholino-4-anilino-s-triazine-6-yl)amino)stilbene disulfonate. Tinopal CBS is the disodium salt of 2,2'-bis-(phenyl-styryl) disulfonate. Tinopal CBS-X is 4.4'-bis-(sulfostyryl)-biphenyl disodium salt, also known as distyryl biphenyl disulfonic acid disodium salt. Also preferred is the commercially available Parawhite KX, supplied by Paramount Minerals and Chemicals of Mumbai, India. Other fluorescent agents suitable for use in the present invention include 1-3-diarylpyrazoline and 7-alkylaminocoumarin.

[0062] In one embodiment, the anti-redeposition polymer is selected from the group comprising: (carboxymethyl)cellulose (CMC), poly(vinyl alcohol) (PVA), poly(vinyl pyrrolidone) (PVP), poly(ethylene glycol) or poly(ethylene oxide) (PEG), ethoxylated poly(ethyleneimine), carboxymethyl inulin (CMI), and polycarboxylic acids (such as PAA, PAA / PMA, polyaspartic acid, and lauryl methacrylate / acrylic acid copolymer), hydrophobically modified CMC (HM-CMC) and silicones, copolymers of terephthalic acid and oligoethylene glycols, copolymers of poly(ethylene terephthalate) and poly(oxyethylene terephthalate) (PET-POET), PVP, poly(vinylimidazole) (PVI), poly(vinylpyridine-N-oxide) (PVPO or PVPNO) and polyvinylpyrrolidone-vinylimidazole (PVPVI). Suitable examples include PVP-K15, PVP-K30, ChromaBond S-400, ChromaBond S-403E and ChromaBond S-100 from Ashland Aqualon, and ChromaBond S-100 from BASF. HP 165, HP 50 (dispersant), HP 53 (dispersant), HP 59 (dispersant), HP 56 (dye transfer inhibitor), HP 66K (dye transfer inhibitor). Additional exemplary polymers include sulfonated polycarboxylates, polyethylene oxide and polypropylene oxide (PEO-PPO), and diquaternary ammonium ethoxy sulfates, ACUSOL from DOW TM 445N detergent polymer and ethoxylated homopolymer from BASF HP 20.

[0063] In one embodiment, the detergent composition comprising the one or more enzymes improves the whiteness of textiles, i.e., a ΔR460 value of at least 1.0, e.g., at least 2.0, 5.0, 8.0, 10.0, or at least 15.0, compared to a detergent containing an optical brightener and / or an anti-redeposition polymer but not containing one or more enzymes of the present invention.

[0064] In one embodiment, in addition to one or more enzymes of the present invention, the detergent composition further comprises an enzyme selected from the group consisting of amylases, lipases, hemicellulases, peroxidases, xylanases, phospholipases, esterases, cutinases, keratinases, reductases, oxidases, phenoloxidases, lipoxygenases, ligninases, pullulanases, β-glucanases, arabinosidases, hyaluronidases, laccases, deoxyribonucleases (DNases), hexosaminidase and / or any mixture thereof.

[0065] In one embodiment, the enzyme suitable for replacing the whiteness maintenance agent is an enzyme with cellulase activity. Suitable cellulase can be selected from glycoside hydrolase family 5 (GH5), glycoside hydrolase family 7 (GH7), glycoside hydrolase family 12 (GH12), glycoside hydrolase family 44 (GH44) and glycoside hydrolase family 45 (GH45), preferably family GH45 cellulase. Suitable family GH44 cellulase can be a family 44 xyloglucanase having a sequence with at least 60% identity to position 40-559 of SEQ ID NO:2 of WO 2001 / 062903.

[0066] In one embodiment, the enzyme having cellulase activity is a polypeptide having at least 60% sequence identity to the polypeptide of SEQ ID NO: 13, 14, 15, 16, 17, 18, 19, 20, 21, 22 or 23. Preferably, the enzyme having cellulase activity is a polypeptide having at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% sequence identity to the polypeptide of SEQ ID NO: 13, 14, 15, 16, 17, 18, 19, 20, 21, 22 or 23, or a fragment thereof having cellulase activity.

[0067] In one embodiment, the enzyme having cellulase activity is a polypeptide having at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99% or 100% sequence identity to the mature polypeptide of SEQ ID NO: 13, or a fragment thereof having cellulase activity. In a specific embodiment, the cellulase is from Thermothielavioides, preferably from Thermothielavioides terrestris.

[0068] In another embodiment, the enzyme having cellulase activity is a polypeptide having at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99% or 100% sequence identity to the mature polypeptide of SEQ ID NO: 14, or a fragment thereof having cellulase activity. In one embodiment, the mature polypeptide is amino acids 22 to 294 of SEQ ID NO: 14. In a specific embodiment, the enzyme is from Paenibacillus, preferably from Paenibacillus polymyxa.

[0069] In another embodiment, the enzyme having cellulase activity is a polypeptide having at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99% or 100% sequence identity to the mature polypeptide of SEQ ID NO: 15, or a fragment thereof having cellulase activity. In a specific embodiment, the enzyme is from the genus Thermoclostridium, preferably from Thermoclostridium terrestris.

[0070] In another embodiment, the enzyme having cellulase activity is a polypeptide having at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99% or 100% sequence identity to the mature polypeptide of SEQ ID NO: 16, or a fragment thereof having cellulase activity. In a specific embodiment, the enzyme is from the genus Thermoclostridium, preferably from Thermoclostridium terrestris.

[0071] In another embodiment, the enzyme having cellulase activity is a polypeptide having at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99% or 100% sequence identity to the mature polypeptide of SEQ ID NO: 17, or a fragment thereof having cellulase activity. In a specific embodiment, the enzyme is from Acremonium, preferably from Acremonium thermophilum.

[0072] In another embodiment, the enzyme having cellulase activity is a polypeptide having at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99% or 100% sequence identity with the mature polypeptide of SEQ ID NO: 18, or a fragment thereof having cellulase activity. In a specific embodiment, the enzyme is from the genus Paenibacillus, preferably from Paenibacillus polymyxa.

[0073] In another embodiment, the enzyme having cellulase activity is a polypeptide having at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99% or 100% sequence identity with the mature polypeptide of SEQ ID NO: 19, or a fragment thereof having cellulase activity. In a specific embodiment, the enzyme is from Neurospora, preferably from Neurospora tetrasperma.

[0074] In another embodiment, the enzyme having cellulase activity is a polypeptide having at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99% or 100% sequence identity with the mature polypeptide of SEQ ID NO: 20, or a fragment thereof having cellulase activity. In a specific embodiment, the enzyme is from Staphylotrichum, preferably from Staphylotrichum coccosporum.

[0075] In another embodiment, the enzyme having cellulase activity is a polypeptide having at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99% or 100% sequence identity with the mature polypeptide of SEQ ID NO: 21, or a fragment thereof having cellulase activity. In a specific embodiment, the enzyme is from Humicola, preferably from Humicola insolens.

[0076] In another embodiment, the enzyme having cellulase activity is a polypeptide having at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99% or 100% sequence identity with the mature polypeptide of SEQ ID NO: 22, or a fragment thereof having cellulase activity. In a specific embodiment, the enzyme is from the genus Paenibacillus, preferably from Paenibacillus polymyxa.

[0077] In another embodiment, the enzyme having cellulase activity is a polypeptide having at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99% or 100% sequence identity with the mature polypeptide of SEQ ID NO: 23, or a fragment thereof having cellulase activity. In a specific embodiment, the enzyme is from Chaetomium, preferably from Chaetomium thermophilum.

[0078] Suitable cellulases in addition are alkaline or neutral cellulases with color care benefits. Examples of such cellulases are the cellulases described in EP 0 495 257, EP 0 531 372, WO 96 / 11262, WO 96 / 29397, WO 98 / 08940. Other examples are those cellulase variants such as described in WO 94 / 07998, EP 0 531 315, US 5,457,046, US 5,686,593, US 5,763,254, WO 95 / 24471, WO 98 / 12307 and WO99 / 001544.

[0079] Commercially available cellulases include Celluzyme, Carezyme, Carezyme Elite, Carezyme Premium, Luminous, Celluclean, Celluclean Classic, Cellusoft, Whitezyme, Celluclean 4500T, and Celluclean 5000L (all registered trademarks of Novozymes A / S), Clazinase and Puradax HA (registered trademarks of Genencor International Inc.), KAC-500(B) (registered trademark of Kao Corporation), and Revitalenz 2000 (registered trademark of Danisco / Dupont), and Biotouch FLX1, Biotouch FCL75, Biotouch DCL, and Biotouch FCC45 (registered trademarks of AB Enzymes), and Lavergy C Bright from BASF.

[0080] The preparation of an enzyme having cellulase activity as described herein can be carried out, for example, as described in WO 2015 / 036579 (incorporated herein by reference). The enzyme of the present invention can also be prepared by any method known in the art.

[0081] In the present invention, polypeptides or mannanases with mannanase activity suitable for use in place of whiteness / color retainers are enzymes that hydrolyze compounds known as mannans. Mannanases are enzymes that catalyze the hydrolysis of 1,4-β-D-mannosidic bonds in mannans, galactomannans, glucomannans, and galactoglucomannans. Mannans are polysaccharides with a main chain of D-pyranomannosyl residues connected in b-1,4-, which may contain galactose or acetyl substitutions and may have glucose residues in the main chain. The main enzyme type involved in the degradation of mannans is endo-1,4-p-mannanase (EC 3.2.1.78), which hydrolyzes internal glycosidic bonds in the main chain of mannans. For the purposes of the present invention, mannanase activity is determined according to the procedure described in Assay II. According to CAZy (www.cazy.org), endo-1,4-p-mannanase can be found in glycoside hydrolase families 5, 26 and 113. Suitable mannanases include those of bacterial or fungal origin. Chemical or genetically modified mannanases are included. It can be a wild type from Bacillus or Humicola, particularly B. agaradhaerens, B. licheniformis, B. halodurans, B. clausii or Humicola insolens. Suitable mannanases are described in WO 1999 / 064619. Commercially available mannanases are Mannaway (Novozymes), EFFECTENZ from DuPont TM M1000 and Preferenz M 100, (Danisco / DuPont), and Biotouch M9 from AB Enzymes.

[0082] Other preferred mannanases include GH26 mannanase, mannanase from Preussiae emulans (mature sequence of SEQ ID NO: 2 of WO2017 / 021515), mannanase from Yunnania penicillata (mature sequence of SEQ ID NO: 2 of WO2017 / 021516), mannanase from Myrothecium roridum (mature sequence of SEQ ID NO: 2 of WO2017 / 021517), mannanase from Chaetomium brasiliense (mature sequence of SEQ ID NO: 2 of WO2017 / 021518), mannanase from Ascobolus stictoideus, or mannanase from Chaetomium chlororaphis. virescens) (SEQ ID NOs: 3 and 6 of WO2015 / 040159).

[0083] In one embodiment, the enzyme having mannanase activity or the mannanase is a polypeptide having at least 60% sequence identity to the polypeptide of SEQ ID NO: 6, 7, 8, 9, 10, 11 or 12. Preferably, the enzyme having mannanase activity is a polypeptide having at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% sequence identity to the polypeptide of SEQ ID NO: 6, 7, 8, 9, 10, 11 or 12, or a fragment thereof having mannanase activity.

[0084] In one embodiment, the enzyme having mannanase activity is a polypeptide having at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99% or 100% sequence identity to the mature polypeptide of SEQ ID NO: 6, or a fragment thereof having mannanase activity. In a specific embodiment, the mannanase is from Alkalihalobacillus, preferably from Alkalihalobacillus bogoriensis.

[0085] In one embodiment, the enzyme having mannanase activity is a polypeptide having at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99% or 100% sequence identity to the mature polypeptide of SEQ ID NO: 7, or a fragment thereof having mannanase activity. In a specific embodiment, the mannanase is from the genus Halobacillus, preferably from Halobacillus bogoria.

[0086] In one embodiment, the enzyme having mannanase activity is a polypeptide having at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99% or 100% sequence identity to the mature polypeptide of SEQ ID NO: 8, or a fragment thereof having mannanase activity.

[0087] In one embodiment, the enzyme having mannanase activity is a polypeptide having at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99% or 100% sequence identity to the mature polypeptide of SEQ ID NO: 9, or a fragment thereof having mannanase activity.

[0088] In one embodiment, the enzyme having mannanase activity is a polypeptide having at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99% or 100% sequence identity to the mature polypeptide of SEQ ID NO: 10, or a fragment thereof having mannanase activity. In a specific embodiment, the mannanase is from Bacillus, preferably from Bacillus clausii.

[0089] In one embodiment, the enzyme having mannanase activity is a polypeptide having at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99% or 100% sequence identity to the mature polypeptide of SEQ ID NO: 11, or a fragment thereof having mannanase activity. In a specific embodiment, the mannanase is from Bacillus species.

[0090] In one embodiment, the enzyme having mannanase activity is a polypeptide having at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99% or 100% sequence identity to the mature polypeptide of SEQ ID NO: 12, or a fragment thereof having mannanase activity. In a specific embodiment, the mannanase is from a Paenibacillus species.

[0091] Proteases suitable for replacing whiteness / color retainers can be of any origin, but are preferably of bacterial or fungal origin, optionally in the form of protein engineered or chemically modified mutants. Proteases can be alkaline proteases, such as serine proteases or metalloproteases. Serine proteases can be, for example, S1 family (such as trypsin) or S8 family (such as subtilisin). Metalloproteases can be, for example, thermolysin, such as thermolysin from M4 family, or another metalloprotease, such as those from M5, M7 or M8 family.

[0092] The term "subtilase" refers to a subgroup of serine proteases according to Siezen et al., Protein Eng. [Protein Engineering] 4 (1991) 719-737 and Siezen et al., Protein Sci. [Protein Science] 6 (1997) 501-523. Serine proteases are a subgroup of proteases characterized by having a serine in the active site that forms a covalent adduct with the substrate. Subtilases can be divided into six subclasses: the subtilisin family, the thermophilic protease family, the proteinase K family, the lanthionine antibiotic peptidase family, the Kexin family, and the Pyrolysin family.

[0093] Although proteases suitable for detergent use can be obtained from a variety of organisms, including fungi such as Aspergillus, detergent proteases are typically obtained from bacteria, particularly from Bacillus and related genera (see Patel and Gupta, supra). Examples of Bacillus species from which subtilases are derived include Bacillus lentus, Bacillus alcalophilus, Bacillus subtilis, Bacillus amyloliquefaciens, Bacillus licheniformis, Bacillus pumilus, and Bacillus gibsonii. Particular subtilisins include subtilisin lentus, subtilisin Novo, subtilisin Carlsberg, subtilisin BPN', subtilisin 309, subtilisin 147 and subtilisin 168, and, for example, protease PD138 (described in WO 93 / 18140). Other useful proteases are, for example, those described in WO 01 / 16285 and WO 02 / 16547.

[0094] Examples of trypsin-like proteases include the Fusarium protease (described in WO 94 / 25583 and WO 2005 / 040372), and the chymotrypsin derived from Cellumonas (described in WO 2005 / 052161 and WO 2005 / 052146).

[0095] Examples of metalloproteases include neutral metalloproteases described in WO 2007 / 044993 (such as those derived from Bacillus amyloliquefaciens), and metalloproteases described, for example, in WO 2015 / 158723 and WO 2016 / 075078.

[0096] Examples of useful proteases are the protease variants described in WO 89 / 06279, WO 92 / 19729, WO 96 / 34946, WO 98 / 20115, WO 98 / 20116, WO 99 / 11768, WO 01 / 44452, WO 03 / 006602, WO 2004 / 003186, WO 2004 / 041979, WO 2007 / 006305, WO 2011 / 036263, WO 2014 / 207227, WO 2016 / 087617 and WO 2016 / 174234. Preferred protease variants may, for example, comprise one or more mutations selected from the group consisting of S3T, V4I, S9R, S9E, A15T, S24G, S24R, K27R, N42R, S55P, G59E, G59D, N60D, N60E, V66A, N74D, S85R, A96S, S97G, S97D, S97A, S97SD, S99E, S99D, S99G, S99M, S99N, S99R, S99H, S101A, V102I, V102Y, V102N, S104A, G116V, G116R, H118D, H118N, A120S, S126L, P126L 27Q, S128A, S154D, A156E, G157D, G157P, S158E, Y161A, R164S, Q176E, N179E, S182E, Q185N, A188P, G189E, V193M, N198D, V199I, Q200L, Y203W, S206G, L211Q, L211D, N212D, N212S, M216S, A226V, K229L, Q230H, Q239R, N246K, S253D, N255W, N255D, N255E, L256E, L256DT268A, and R269H, where the position numbers correspond to WO The position of the Bacillus lentus protease shown in SEQ ID NO: 1 of WO 2016 / 001449. The protease variant having one or more of these mutations is preferably the Bacillus lentus protease shown in SEQ ID NO: 1 of WO 2016 / 001449 ( Also referred to as a variant of subtilisin 309) or a variant of Bacillus amyloliquefaciens protease (BPN') shown in SEQ ID NO: 2 of WO2016 / 001449. Such a protease variant preferably has at least 80% sequence identity with SEQ ID NO: 1 or SEQ ID NO: 2 of WO 2016 / 001449.

[0097] Another protease of interest is the alkaline protease from Bacillus lentus DSM 5483 (as described, for example, in WO 91 / 02792) and variants thereof (these variants are described, for example, in WO 92 / 21760, WO 95 / 23221, EP 1921147, EP1921148 and WO 2016 / 096711).

[0098] Alternatively, the protease may be a variant of the TY145 protease having SEQ ID NO: 1 of WO 2004 / 067737, for example a variant comprising a substitution at one or more positions corresponding to positions 27, 109, 111, 171, 173, 174, 175, 180, 182, 184, 198, 199 and 297 of SEQ ID NO: 1 of WO 2004 / 067737, wherein the protease variant has at least 75% but less than 100% sequence identity with SEQ ID NO: 1 of WO 2004 / 067737.

[0099] In some embodiments of the present invention, the preferred protease according to the present invention can be a protease having at least 60%, such as at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% sequence identity with the polypeptide of SEQ ID NO: 1, 2, 3, 4 or 5.

[0100] In another embodiment, the preferred protease according to the present invention may be a variant of the polypeptide of SEQ ID NO: 2, comprising an alteration at one or more positions corresponding to positions 3, 4, 9, 15, 43, 68, 76, 99, 101, 103, 104, 160, 167, 170, 194, 199, 205, 206, 209, 217, 218, 222, 245, 261 and 262, wherein the position numbers correspond to the positions of SEQ ID NO: 1, wherein each alteration is independently a substitution, deletion or insertion, and wherein the variant has protease activity and has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99%, but less than 100% sequence identity with the polypeptide of SEQ ID NO: 2.

[0101] In one embodiment, the protease of the invention is a variant of the polypeptide of SEQ ID NO: 2 comprising one or more substitutions selected from the group consisting of S3T, V4I, S9E, S9R, A15T, V68A, N76D, S99D, S99G, S99A, S99SE, S101E, S101N, S101R, S103A, V104I, G160S, Y167A, R170S, A194P, V199M, V205I, Q206L, Y209W, L217D, L217Q, N218D, M222S, Q245R, N261W, and L262E, wherein the position numbers correspond to the positions of SEQ ID NO: 1, and wherein the variant has protease activity and is identical to the polypeptide of SEQ ID NO: 2. NO:2 has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, but less than 100% sequence identity.

[0102] In one embodiment, the protease of the invention is a variant of the polypeptide of SEQ ID NO: 2 comprising one or more substitutions selected from the group consisting of S3T, V4I, S9E, S9R, A15T, V68A, N76D, S99D, S99G, S99A, S99SE, S101E, S101N, S101R, S103A, V104I, G160S, Y167A, R170S, A194P, V199M, V205I, Q206L, Y209W, L217D, L217Q, N218D, M222S, Q245R, N261W, and L262E, wherein the position numbers correspond to the positions of SEQ ID NO: 1, and wherein the variant has protease activity and is identical to the polypeptide of SEQ ID NO: 2. NO:2 has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, but less than 100% sequence identity.

[0103] In one embodiment, the protease of the present invention is a variant of the polypeptide of SEQ ID NO:2, comprising the substitution S87N, wherein the variant has protease activity and wherein the position corresponds to the position of SEQ ID NO:1, and wherein the variant has protease activity and has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99%, but less than 100% sequence identity to SEQ ID NO:2.

[0104] In one embodiment, the protease of the present invention is a variant of the polypeptide of SEQ ID NO:2, comprising the substitutions Y167A+R170S+A194P, wherein the position numbers correspond to the positions of SEQ ID NO:1, and wherein the variant has protease activity and has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99%, but less than 100% sequence identity to SEQ ID NO:2.

[0105] In one embodiment, the protease of the present invention is a variant of the polypeptide of SEQ ID NO:2, comprising the substitutions S9E+N43R+N76D+V205I+Q206L+Y209W+S259D+N261W+L262E, wherein the position numbers correspond to the positions of SEQ ID NO:1, and wherein the variant has protease activity and has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99%, but less than 100% sequence identity to SEQ ID NO:2.

[0106] In one embodiment, the protease of the present invention is a variant of the polypeptide of SEQ ID NO:2, which comprises the substitutions S3T+N43R+N76D+S87N+G118M+S128Q+N184E+V205I+Q206L+Y209W+S259D+N261W+L262E, wherein the position numbers correspond to the positions of SEQ ID NO:1, and wherein the variant has protease activity and has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99%, but less than 100% sequence identity to SEQ ID NO:2.

[0107] In one embodiment, the protease of the present invention is a variant of the polypeptide of SEQ ID NO:4, comprising the substitutions A68S+T77N+T78I+G127S+A128P+G165Q+N184Q+A202V+N217S+S258P, wherein the position numbers correspond to the positions of SEQ ID NO:4, and wherein the variant has protease activity and has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99%, but less than 100% sequence identity to SEQ ID NO:4.

[0108] In one embodiment, the protease of the present invention is a variant comprising a substitution at one or more positions corresponding to position 171, 173, 175, 179 or 180 of SEQ ID NO: 1 of WO2004 / 067737, wherein the variant has protease activity and has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99%, but less than 100% sequence identity with SEQ ID NO: 1 of WO2004 / 067737.

[0109] In one embodiment, the protease of the present invention is a variant comprising one or more substitutions compared to a parent protease, wherein the one or more substitutions are selected from the group consisting of: X3V, X9[E,R], X22[R,A], X43R, X61[E,D], X62[E,D], X76[D], X87N, X101[E,G,D,N,M], X103A, X104I, X118[V,R], X120V, X128[A,L,S], X12 9Q, X130A, X160D, X185[E,D], 188[E,D], X191N, X194P, X205I, X206L, X209W, X216V, X217[Q,D,E], X218[D,E,S], X232V, X245R, X248D, X256[E,D], X259[E,D], X261[E,D,W], and X262[E,D], where the position numbers correspond to BPN'(SEQ ID NO: 1). ID NO:1), wherein "X" represents any amino acid residue present in the specified position in the parent protease, and wherein the variant has protease activity and has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99%, but less than 100% sequence identity to SEQ ID NO:1 or SEQ ID NO:2.

[0110] In one embodiment, the protease of the present invention is a variant comprising any of the following substitution groups compared to a parent protease, wherein the parent protease has an amino acid sequence of SEQ ID NO: 1 or SEQ ID NO: 2 or has at least 80% sequence identity with SEQ ID NO: 1 or SEQ ID NO: 2, wherein the position number corresponds to the position of BPN' (SEQ ID NO: 1), wherein "X" represents any amino acid residue present in the specified position in the parent protease, and wherein the substitution group is selected from the group consisting of:

[0111] i.X9R+X15T+X68A+X218D+X245R,

[0112] ii.X9R+X15T+X68A+X245R,

[0113] iii.X61E+X194P+X205I+X261D,

[0114] iv.X61D+X205I+X245R,

[0115] v.X61E+X194P+X205I+X261D,

[0116] vi. X87N + X118V + X128L + X129Q + X130A,

[0117] vii. X87N + X101M + X118V + X128L + X129Q + X130A,

[0118] viii. X76D + X87R + X118R + X128L + X129Q + X130A,

[0119] ix. X22A + X62D + X101G + X188D + X232V + X245R,

[0120] x. X103A + X104I,

[0121] xi. X22R + X101G + X232V + X245R,

[0122] xii. X103A + X104I + X156D,

[0123] xiii. X103A + X104I + X261E,

[0124] xiv. X62D + X245R,

[0125] xv. X101N + X128A + X217Q,

[0126] xvi. X101E + X217Q,

[0127] xvii. X101E + X217D,

[0128] xviii. X9E + X43R + X262E,

[0129] xix. X76D + X43R + X209W,

[0130] xx. X205I + X206L + X209W,

[0131] xxi. X185E + X188E + X205I,

[0132] xxii. X256D + X261W + X262E,

[0133] xxiii. X191N + X209W,

[0134] xxiv. X261E + X262E,

[0135] xxv. X261E + X262D, and

[0136] xxvi.X167A+X170S+X194P,

[0137] wherein the protease variant has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99%, but less than 100% sequence identity to SEQ ID NO: 1 or 2.

[0138] In one embodiment, a protease variant of the present invention comprises the amino acid sequence of SEQ ID NO: 2 having substitutions Y167A+R170S+A194P, wherein the variant has protease activity and wherein the positions correspond to the positions of SEQ ID NO: 1.

[0139] In one embodiment, a protease variant of the present invention comprises the amino acid sequence of SEQ ID NO: 2 having substitutions S9E+N43R+N76D+V205I+Q206L+Y209W+S259D+N261W+L262E, wherein the variant has protease activity and wherein the position numbers correspond to the positions of SEQ ID NO: 1.

[0140] In one embodiment, the protease of the present invention is an enzyme having the amino acid sequence of SEQ ID NO: 2, 3, 4 or 5 or consisting of the amino acid sequence.

[0141] Suitable commercially available proteases may include those sold under the following trade names: Duralase TM 、Durazym TM , Ultra, Ultra、Primase TM , Ultra, Ultra, Pro, Blaze 100T, Blaze 125T、Blaze 150T, Blaze 200T, Uno, In and Excel (Novozymes), those sold under the following trade names: Maxatase TM Maxacal TM , Ox, OxP, FN2 TM 、FN3 TM 、FN4 exTM , Excellenz TM P1000、Excellenz TM P1250、Eraser TM , P100, P300, Purafect Prime, Preferenz P110 TM 、EffectenzP1000 TM , Effectenz P1050 TM , Ox, Effectenz TM P2000, Purafast TM , Opticlean TM and (Dansonic / DuPont), BLAP (sequence shown in Figure 29 of US 5352604) and its variants (Henkel AG), Puzhi 1.0L from Bestzyme, and KAP (alkaliphilic Bacillus subtilisin) from Kao Corporation.

[0142] In one aspect, variants of enzymes (proteases, mannanases and cellulases) according to the invention comprise substitutions, deletions and / or insertions at one or more positions. In one embodiment, the number of amino acid substitutions, deletions and / or insertions introduced into, for example, the mature polypeptides of SEQ ID NO: 2, 3, 5, 6, 10 or 11 is no more than 20, for example 1-15, such as 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10. Amino acid changes may be of a minor nature, i.e., conservative amino acid substitutions or insertions that do not significantly affect the folding and / or activity of the protein; typically small deletions of 1-30 amino acids; small amino-terminal or carboxyl-terminal extensions, such as amino-terminal methionine residues; small linker peptides of up to 20-25 residues; or small extensions that promote purification by changing the net charge or another function (such as a polyhistidine segment, an antigenic epitope or a binding domain).

[0143] Examples of conservative substitutions are within the group consisting of basic amino acids (arginine, lysine, and histidine), acidic amino acids (glutamic acid and aspartic acid), polar amino acids (glutamine and asparagine), hydrophobic amino acids (leucine, isoleucine, and valine), aromatic amino acids (phenylalanine, tryptophan, and tyrosine), and small amino acids (glycine, alanine, serine, threonine, and methionine). Amino acid substitutions that generally do not alter specific activity are known in the art and are described, for example, by H. Neurath and RL Hill, 1979, The Proteins, Academic Press, New York. Common substitutions are Ala / Ser, Val / Ile, Asp / Glu, Thr / Ser, Ala / Gly, Ala / Thr, Ser / Asn, Ala / Val, Ser / Gly, Tyr / Phe, Ala / Pro, Lys / Arg, Asp / Asn, Leu / Ile, Leu / Val, Ala / Glu, and Asp / Gly.

[0144] Alternatively, these amino acid changes have such a property that the physicochemical properties of the polypeptide are changed. For example, amino acid changes can improve the thermal stability of the polypeptide, change the substrate specificity, change the optimal pH, and the like.

[0145] Essential amino acids in polypeptides can be identified according to procedures known in the art, such as site-directed mutagenesis or alanine scanning mutagenesis (Cunningham and Wells, 1989, Science 244: 1081-1085). In the latter technique, single alanine mutations are introduced at every residue in the molecule, and the resulting mutant molecules are tested for protease activity, cellulase activity, or mannanase activity to identify amino acid residues that are critical for the activity of the molecule. See also, Hilton et al., 1996, J. Biol. Chem. 271: 4699-4708. Active sites of enzymes or other biological interactions can also be determined by physical analysis of the structure, such as by techniques such as nuclear magnetic resonance, crystallography, electron diffraction, or photoaffinity labeling, together with mutations to putative contact site amino acids. See, e.g., de Vos et al., 1992, Science 255:306-312; Smith et al., 1992, J. Mol. Biol. 224:899-904; Wlodaver et al., 1992, FEBS Lett. 309:59-64. The identities of essential amino acids can also be inferred from alignments with related polypeptides.

[0146] In some other embodiments, the enzymes suitable for replacing the whiteness / color maintenance agents of the detergent composition are enzyme combinations of proteases, mannanases and / or cellulases as described above.

[0147] In some other embodiments, the enzyme suitable for replacing the whiteness / color retaining agent is an enzyme combination of a protease and a mannanase, wherein the protease is a polypeptide comprising or consisting of SEQ ID NO: 2, wherein the mannanase is a polypeptide having or consisting of SEQ ID NO: 6, 7, 8, 9, 10 or 11. In a specific embodiment, the protease is a polypeptide of SEQ ID NO: 2 having the substitutions S9E+N43R+N76D+V205I+Q206L+Y209W+S259D+N261W+L262E, wherein the position numbering is based on the numbering of SEQ ID NO: 1, and wherein the mannanase is a polypeptide having or consisting of SEQ ID NO: 6 or 7.

[0148] In some embodiments, the enzyme suitable for replacing the whiteness / color maintenance agent is an enzyme combination of a protease and a cellulase, wherein the protease comprises or consists of a polypeptide of SEQ ID NO: 2, wherein the cellulase is a polypeptide having or consisting of SEQ ID NO: 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22 or 23. In a specific embodiment, the protease is a polypeptide of SEQ ID NO: 2 having the substitutions S9E+N43R+N76D+V205I+Q206L+Y209W+S259D+N261W+L262E, wherein the position numbering is based on the numbering of SEQ ID NO: 1, and wherein the cellulase is a polypeptide having or consisting of SEQ ID NO: 18, 21 or 22.

[0149] In some embodiments, the enzyme suitable for replacing the whiteness / color retaining agent is an enzyme combination of a mannanase and a cellulase, wherein the mannanase is a polypeptide comprising or consisting of SEQ ID NO: 6, 7, 8, 9, 10 or 11, wherein the cellulase is a polypeptide comprising or consisting of SEQ ID NO: 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22 or 23. In a specific embodiment, the mannanase is a polypeptide comprising or consisting of SEQ ID NO: 6 or 7, and wherein the cellulase is a polypeptide having or consisting of SEQ ID NO: 18, 21 or 22.

[0150] In some embodiments, the enzyme suitable for replacing the whiteness / color retaining agent is an enzyme combination of a protease, a mannanase, and a cellulase, wherein the protease is a polypeptide comprising or consisting of SEQ ID NO: 2, wherein the mannanase is a polypeptide having or consisting of SEQ ID NO: 6, 7, 8, 9, 10 or 11, and wherein the cellulase is a polypeptide having or consisting of SEQ ID NO: 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22 or 23. In a specific embodiment, the protease is a polypeptide of SEQ ID NO: 2 having the substitutions S9E+N43R+N76D+V205I+Q206L+Y209W+S259D+N261W+L262E, wherein the position numbering is based on the numbering of SEQ ID NO: 1, and the mannanase is a polypeptide having or consisting of SEQ ID NO: 6 or 7, and the cellulase is a polypeptide having or consisting of SEQ ID NO: 18, 21 or 22. In yet another embodiment, the protease is a polypeptide of SEQ ID NO: 2, and the mannanase is a polypeptide having or consisting of SEQ ID NO: 7, and the cellulase is a polypeptide having or consisting of SEQ ID NO: 18 or 21.

[0151] The one or more enzymes of the detergent composition of the present invention may be stabilized using conventional stabilizers, for example polyols, such as propylene glycol or glycerol, sugars or sugar alcohols, lactic acid, boric acid or boric acid derivatives, such as aromatic borate esters, or phenylboronic acid derivatives, such as 4-formylphenylboronic acid, and the composition may be formulated as described in, for example, WO92 / 19709 and WO92 / 19708.

[0152] In one embodiment of the invention, the textile is selected from the group consisting of: cellulose-based textiles and textiles partially made of cellulose-based textiles. In one embodiment, the textile is cotton, or a mixture of cotton and other types of fabrics, such as a mixture of at least 50% polyester and at least 20% cotton, or a mixture of at least 30% polyester and at least 40% cotton.

[0153] The optical brightener may be present at up to 0.6% by weight of the composition, e.g., up to 0.4% by weight of the composition, up to 0.3% by weight of the composition, up to 0.25% by weight of the composition, up to 0.2% by weight of the composition, up to 0.15% by weight of the composition, up to 0.1% by weight of the composition, up to 0.08% by weight of the composition, up to 0.05% by weight of the composition, up to 0.04% by weight of the composition, up to 0.02% by weight of the composition, or up to 0.01% by weight of the composition.

[0154] The anti-redeposition polymer may be present at up to 1% by weight of the composition, for example up to 0.8% by weight of the composition, up to 0.6% by weight of the composition, up to 0.4% by weight of the composition, up to 0.2% by weight of the composition, up to 0.15% by weight of the composition, up to 0.1% by weight of the composition, up to 0.08% by weight of the composition, up to 0.05% by weight of the composition, up to 0.04% by weight of the composition.

[0155] Surfactants

[0156] The detergent composition can comprise one or more surfactants, which can be anionic and / or cationic and / or nonionic and / or semi-polar and / or zwitterionic or its mixture. In a specific embodiment, the detergent composition comprises a mixture of one or more nonionic surfactants and one or more anionic surfactants. One or more surfactants typically exist by weight from about 0.1% to 60%, for example, about 1% to about 40% or about 3% to about 20% or about 3% to about 10%. Select the one or more surfactants based on the desired cleaning application, and the surfactant comprises any one or more conventional surfactants as known in the art. Any surfactant for use in detergents as known in the art can be utilized.

[0157] When included therein, the detergent will typically contain from about 1% to about 40% (e.g., from about 5% to about 30%, including from about 5% to about 15%, or from about 20% to about 25%) by weight of anionic surfactant. Non-limiting examples of anionic surfactants include sulfates and sulfonates, particularly linear alkylbenzene sulfonates (LAS), isomers of LAS, branched alkylbenzene sulfonates (BABS), phenylalkanesulfonates, alpha-olefin sulfonates (AOS), olefin sulfonates, alkene sulfonates, alkane-2,3-diylbis(sulfates), hydroxyalkanesulfonates and disulfonates, alkyl sulfates (AS) such as sodium dodecyl sulfate (SDS), fatty alcohol sulfates (FAS), primary alcohol sulfates (PAS ), alcohol ether sulfates (AES or AEOS or FES, also known as alcohol ethoxy sulfates or fatty alcohol ether sulfates), secondary alkane sulfonates (SAS), paraffin sulfonates (PS), ester sulfonates, sulfonated fatty acid glycerides, α-sulfonic fatty acid methyl esters (α-SFMe or SES) (including methyl sulfonate (MES)), alkyl succinic acid or alkenyl succinic acid, dodecenyl / tetradecenyl succinic acid (DTSA), fatty acid derivatives of amino acids, diesters and monoesters of sulfosuccinic acid or soap, and combinations thereof.

[0158] When included therein, detergents will typically contain from about 1% to about 40% by weight of a cationic surfactant. Non-limiting examples of cationic surfactants include alkyl dimethylethanol quaternary ammonium (ADMEAQ), cetyl trimethyl ammonium bromide (CTAB), dimethyl dioctadecyl ammonium chloride (DSDMAC), and alkyl benzyl dimethyl ammonium and combinations thereof, alkyl quaternary ammonium compounds, alkoxylated quaternary ammonium (AQA).

[0159] When included therein, the detergent will typically contain from about 0.2% to about 60% (e.g., from about 0.3% to about 40%, such as from about 0.5% to about 30%, especially from about 1% to about 20%, from about 3% to about 10%, such as from about 3% to about 5%, or from about 8% to about 12%) by weight of nonionic surfactant. Non-limiting examples of nonionic surfactants include alcohol ethoxylates (AE or AEO), alcohol propoxylates, propoxylated fatty alcohols (PFA), alkoxylated fatty acid alkyl esters (e.g., ethoxylated and / or propoxylated fatty acid alkyl esters), alkylphenol ethoxylates (APE), nonylphenol ethoxylates (NPE), alkyl polyglycosides (APG), alkoxylated amines, fatty acid monoethanolamide (FAM), fatty acid diethanolamide (FADA), ethoxylated fatty acid monoethanolamide (EFAM), propoxylated fatty acid monoethanolamide (PFAM), polyhydroxyalkyl fatty acid amides, or N-acyl N-alkyl derivatives of glucosamine (glucamide (GA), or fatty acid glucamide (FAGA)), as well as products available under the trade names SPAN and TWEEN, and combinations thereof.

[0160] When included therein, the detergent will typically contain from about 1% to about 40% by weight of a semi-polar surfactant. Non-limiting examples of semi-polar surfactants include amine oxides (AO) such as alkyl dimethyl amine oxides, N-(coconut oil alkyl)-N,N-dimethyl amine oxides, and N-(tallow-alkyl)-N,N-bis(2-hydroxyethyl) amine oxides, fatty acid alkanolamides and ethoxylated fatty acid alkanolamides, and combinations thereof.

[0161] When included therein, detergents will typically contain from about 1% to about 40% by weight of a zwitterionic surfactant.Non-limiting examples of zwitterionic surfactants include betaines, alkyl dimethyl betaines, and sulfobetaines, and combinations thereof.

[0162] Builders and co-builders

[0163] The detergent composition may contain about 0-65% by weight of a detergent builder or co-builder, or a mixture thereof. In dishwashing detergents, the builder level is typically 40%-65%, especially 50%-65%. The builder and / or co-builder may be particularly useful with Ca 2+ and Mg 2+ The chelating agent that forms a water-soluble complex. Any builder and / or co-builder known in the art for use in laundry detergents can be utilized. Non-limiting examples of builders include zeolites, diphosphates (pyrophosphates), triphosphates (such as sodium triphosphate (STP or STPP)), carbonates (such as sodium carbonate), soluble silicates (such as sodium metasilicate), layered silicates (e.g., SKS-6 from Hoechst), disilicates, ethanolamines (such as 2-aminoethane-1-ol (MEA), iminodiethanol (DEA) and 2,2',2"-nitrotriethanol (TEA)), and carboxymethyl inulin (CMI), and combinations thereof.

[0164] The detergent composition can also contain 0-65% by weight of detergent co-builder, or its mixture. The detergent composition can include a co-builder including a single co-builder or a combination of a builder (e.g., a zeolite builder). The limiting examples of co-builders include homopolymers or copolymers of polyacrylates, such as poly(acrylic acid) (PAA) or copolymers (acrylic acid / maleic acid) (PAA / PMA). Other limiting examples include citrates, chelating agents (such as aminocarboxylates, aminopolycarboxylates, and phosphonates), and alkyl succinic acids or alkenyl succinic acids. Further specific examples include 2,2′,2″-nitrilotriacetic acid (NTA), ethylenediaminetetraacetic acid (EDTA), diethylenetriaminepentaacetic acid (DTPA), iminodisuccinic acid (IDS), ethylenediamine-N,N′-disuccinic acid (EDDS), methylglycinediacetic acid (MGDA), glutamic acid-N,N-diacetic acid (GLDA), 1-hydroxyethane-1,1-diylbis(phosphonic acid) (HEDP), ethylenediaminetetra(methylene)tetra(phosphonic acid) (EDTMPA), diethylenetriaminepenta(methylene)penta(phosphonic acid) (DTPMPA), N-(2-hydroxyethyl)iminodiacetic acid (EDG), aspartic acid-N-monoacetic acid (ASMA), aspartic acid-N,N-diacetic acid (ASDA), aspartic acid-N-monopropionic acid (ASMP), iminodisuccinic acid (IDA), N-(2-sulfomethyl)aspartic acid (SMAS), N-(2-sulfoethyl)aspartic acid (SE N-(2-sulfomethyl)glutamic acid (SMGL), N-(2-sulfoethyl)glutamic acid (SEGL), N-methyliminodiacetic acid (MIDA), α-alanine-N,N-diacetic acid (α-ALDA), serine-N,N-diacetic acid (SEDA), isoserine-N,N-diacetic acid (ISDA), phenylalanine-N,N-diacetic acid (PHDA), anthranilic acid-N,N-diacetic acid (ANDA), para-aminobenzenesulfonic acid-N,N-diacetic acid (SLDA), taurine-N,N-diacetic acid (TUDA) and sulfomethyl-N,N-diacetic acid (SMDA), N-(hydroxyethyl)-ethylenediaminetriacetic acid (HEDTA), diethanolglycine (DEG), diethylenetriaminepenta(methylenephosphonic acid) (DTPMP), aminotri(methylenephosphonic acid) (ATMP), and combinations and salts thereof. Additional exemplary builders and / or co-builders are described, for example, in WO 09 / 102854, US 5977053.

[0165] Bleaching system

[0166] The detergent may contain 0-40%, for example about 5% to about 25%, by weight of a bleaching system. Any bleaching system known in the art for use in laundry detergents may be utilized. Suitable bleaching system components include bleach catalysts, photobleaches, bleach activators, hydrogen peroxide sources such as sodium percarbonate and sodium perborate, preformed peracids, and mixtures thereof.

[0167] polymer

[0168] In addition to the anti-redeposition polymers as mentioned above, the detergent may further contain 0-10% by weight (e.g. 0.5%-5%, 2%-5%, 0.5%-2% or 0.2%-1%) of another type of polymer. The polymer may act as a co-builder as mentioned above, or may provide fiber protection, soil release, dye transfer inhibition, greasy cleaning and / or defoaming properties. Some polymers may have more than one of the above-mentioned properties and / or more than one of the motifs mentioned below. Any polymer known in the art for use in detergents may be utilized.

[0169] Other enzymes

[0170] The detergent additives and detergent compositions may comprise one or more additional enzymes, such as lipases, amylases, hemicellulases, peroxidases, xylanases, phospholipases, esterases, cutinases, keratinases, reductases, oxidases, phenoloxidases, lipoxygenases, ligninases, pullulanases, maranases, β-glucanases, arabinosidases, hyaluronidases, laccases, deoxyribonucleases (DNases), hexosaminidase, and any mixtures thereof.

[0171] Typically, the properties of the one or more selected enzymes should be compatible with the selected detergent (i.e., optimal pH, compatibility with other enzymes and non-enzyme ingredients, etc.), and the one or more enzymes should be present in an effective amount. Typically, when present in the cleaning composition, the one or more enzymes can be present at a level of at least 0.001 mg of enzyme protein, at least 0.006 mg of enzyme protein, at least 0.008 mg of enzyme protein, at least 0.01 mg of enzyme protein, at least 0.1 mg of enzyme protein, at least 0.5 mg of enzyme protein, at least 1 mg of enzyme protein, or at least 2 mg of enzyme protein.

[0172] Formulation of enzymes in co-granules

[0173] Enzyme of the present invention can be formulated as granules, for example, as co-granules combining one or more enzymes. Then, each enzyme will be present in a variety of granules, which ensure that the enzyme is more evenly distributed in the detergent. This also reduces the physical isolation of different enzymes caused by different particle sizes. The method for producing multi-enzyme co-granules for the detergent industry is disclosed in IP.com disclosure IPCOM000200739D.

[0174] Another example of a formulation of enzymes by using co-granules is disclosed in WO 2013 / 188331, which relates to a cleaning composition comprising: (a) a multi-enzyme co-granule; (b) less than 10 wt% zeolite (on anhydrous basis); and (c) less than 10 wt% phosphate (on anhydrous basis), wherein the enzyme co-granule comprises from 10 wt% to 98 wt% of a moisture sink component, and the composition additionally comprises from 20 wt% to 80 wt% of a detergent moisture sink component. WO 2013 / 188331 also relates to a method of treating and / or cleaning a surface, the method comprising the steps of: (i) contacting the surface in an aqueous washing liquor with a cleaning composition as claimed and described herein, (ii) rinsing and / or drying the surface.

[0175] The multi-enzyme co-granule can comprise an enzyme of the invention and one or more enzymes selected from the group consisting of lipase, amylase, hemicellulase, peroxidase, xylanase, phospholipase, esterase, cutinase, keratinase, reductase, oxidase, phenoloxidase, lipoxygenase, ligninase, pullulanase, maranaase, β-glucanase, arabinosidase, hyaluronidase, laccase, deoxyribonuclease (DNase), hexosaminidase, and any mixture thereof.

[0176] The present invention is further summarized in the following paragraphs:

[0177] 1. Use of an enzyme for replacing a whiteness maintenance agent in a cleaning composition, wherein the enzyme is selected from the group consisting of proteases, mannanases, cellulases and combinations thereof.

[0178] 2. The use as described in paragraph 1, wherein the whiteness maintenance agent is selected from the group consisting of optical brighteners, anti-redeposition polymers and mixtures thereof.

[0179] 3. The use according to paragraph 1 or 2, wherein the replacement is partial or complete replacement.

[0180] 4. The use according to any one of the preceding paragraphs, wherein the enzyme is a combination of a protease and a cellulase.

[0181] 5. The use as described in any of the preceding paragraphs, wherein the enzyme is a combination of a protease and a mannanase.

[0182] 6. The use as described in any of the preceding paragraphs, wherein the enzyme is a combination of cellulase and mannanase.

[0183] 7. The use as described in any of the preceding paragraphs, wherein the enzyme is a combination of protease, mannanase and cellulase.

[0184] 8. The use of any of the preceding paragraphs, wherein the protease has at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% sequence identity to the polypeptide of SEQ ID NO: 1, 2, 3, 4 or 5.

[0185] 9. The use of any of the preceding paragraphs, wherein the protease is a variant of the polypeptide of SEQ ID NO: 2 comprising an alteration at one or more positions corresponding to positions 3, 4, 9, 15, 43, 68, 76, 99, 101, 103, 104, 160, 167, 170, 194, 199, 205, 206, 209, 217, 218, 222, 245, 261 and 262, wherein the position numbers correspond to positions of SEQ ID NO: 1, wherein each alteration is independently a substitution, deletion or insertion, and wherein the variant has protease activity and has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99%, but less than 100% sequence identity with the polypeptide of SEQ ID NO: 2.

[0186] 10. The use according to any one of the preceding paragraphs, wherein the protease is:

[0187] (a) a variant of the polypeptide of SEQ ID NO: 2, comprising one or more substitutions selected from the group consisting of S3T, V4I, S9E, S9R, A15T, V68A, N76D, S99D, S99G, S99A, S99SE, S101E, S101N, S101R, S103A, V104I, G160S, Y167A, R170S, A194P, V199M, V205I, Q206L, Y209W, L217D, L217Q, N218D, M222S, Q245R, N261W and L262E, wherein the position numbers correspond to the positions of SEQ ID NO: 1, and wherein the variant has protease activity and is identical to the polypeptide of SEQ ID NO: 2. NO:2 has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, but less than 100% sequence identity;

[0188] (b) a variant of the polypeptide of SEQ ID NO:2 comprising the substitutions N76D+Y209W, wherein the positions are numbered corresponding to the positions of SEQ ID NO:1, and wherein the variant has protease activity and has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100% sequence identity to SEQ ID NO:2;

[0189] (c) a variant of the polypeptide of SEQ ID NO:2 comprising the substitution S87N, wherein the variant has protease activity and wherein the position corresponds to the position of SEQ ID NO:1, and wherein the variant has protease activity and has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99%, but less than 100% sequence identity to SEQ ID NO:2;

[0190] (d) a variant of the polypeptide of SEQ ID NO:2, comprising the substitutions Y167A+R170S+A194P, wherein the position numbers correspond to the positions of SEQ ID NO:1, and wherein the variant has protease activity and has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99%, but less than 100% sequence identity to SEQ ID NO:2;

[0191] (e) a variant of the polypeptide of SEQ ID NO:2, comprising the substitutions S9E+N43R+N76D+V205I+Q206L+Y209W+S259D+N261W+L262E, wherein the positions are numbered corresponding to the positions of SEQ ID NO:1, and wherein the variant has protease activity and has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100% sequence identity to SEQ ID NO:2;

[0192] (f) a variant of the polypeptide of SEQ ID NO:4, comprising the substitutions A68S+T77N+T78I+G127S+A128P+G165Q+N184Q+A202V+N217S+S258P, wherein the position numbers correspond to the positions of SEQ ID NO:4, and wherein the variant has protease activity and has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100% sequence identity to SEQ ID NO:4;

[0193] (g) a variant comprising a substitution at one or more positions corresponding to position 171, 173, 175, 179 or 180 of SEQ ID NO: 1 of WO2004 / 067737, wherein the variant has protease activity and has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99%, but less than 100% sequence identity to SEQ ID NO: 1 of WO2004 / 067737;

[0194] (h) a protease variant comprising one or more substitutions compared to a parent protease, the one or more substitutions being selected from the group consisting of: X3V, X9[E,R], X22[R,A], X43R, X61[E,D], X62[E,D], X76[D], X87N, X101[E,G,D,N,M], X103A, X104I, X118[V,R], X120V, X128[A,L,S], X129Q, X130 30A, X160D, X185[E,D], 188[E,D], X191N, X194P, X205I, X206L, X209W, X216V, X217[Q,D,E], X218[D,E,S], X232V, X245R, X248D, X256[E,D], X259[E,D], X261[E,D,W], and X262[E,D], where the position numbers correspond to BPN'(SEQ ID NO: ... ID NO:1), wherein "X" represents any amino acid residue present in the specified position in the parent protease, and wherein the variant has protease activity and has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99%, but less than 100% sequence identity to SEQ ID NO:1 or SEQ ID NO:2; and

[0195] (i) a protease variant comprising any of the following substitution groups compared to a parent protease, wherein the parent protease has an amino acid sequence of SEQ ID NO: 1 or SEQ ID NO: 2 or has at least 80% sequence identity to SEQ ID NO: 1 or SEQ ID NO: 2, wherein the position number corresponds to the position of BPN' (SEQ ID NO: 1), wherein "X" represents any amino acid residue present in the specified position in the parent protease, and wherein the substitution group is selected from the group consisting of:

[0196] i.X9R+X15T+X68A+X218D+X245R,

[0197] ii.X9R+X15T+X68A+X245R,

[0198] iii.X61E+X194P+X205I+X261D,

[0199] iv.X61D+X205I+X245R,

[0200] v.X61E+X194P+X205I+X261D,

[0201] vi. X87N + X118V + X128L + X129Q + X130A,

[0202] vii. X87N + X101M + X118V + X128L + X129Q + X130A,

[0203] viii. X76D + X87R + X118R + X128L + X129Q + X130A,

[0204] ix. X22A + X62D + X101G + X188D + X232V + X245R,

[0205] x. X103A + X104I,

[0206] xi. X22R + X101G + X232V + X245R,

[0207] xii. X103A + X104I + X156D,

[0208] xiii. X103A + X104I + X261E,

[0209] xiv. X62D + X245R,

[0210] xv. X101N + X128A + X217Q,

[0211] xvi. X101E + X217Q,

[0212] xvii. X101E + X217D,

[0213] xviii. X9E + X43R + X262E,

[0214] xix. X76D + X43R + X209W,

[0215] xx. X205I + X206L + X209W,

[0216] xxi. X185E + X188E + X205I,

[0217] xxii. X256D + X261W + X262E,

[0218] xxiii. X191N + X209W,

[0219] xxiv. X261E + X262E,

[0220] xxv. X261E + X262D, and

[0221] xxvi.X167A+X170S+X194P,

[0222] wherein the protease variant has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99%, but less than 100% sequence identity to SEQ ID NO: 1 or 2.

[0223] 11. The use as described in any of the preceding paragraphs, wherein the protease variant comprises the amino acid sequence of SEQ ID NO: 2 having substitutions Y167A+R170S+A194P, wherein the variant has protease activity and wherein the position numbers correspond to the positions of SEQ ID NO: 1.

[0224] 12. The use of any of the preceding paragraphs, wherein the protease variant comprises the amino acid sequence of SEQ ID NO: 2 having the substitutions S9E+N43R+N76D+V205I+Q206L+Y209W+S259D+N261W+L262E, wherein the variant has protease activity and wherein the positions are numbered corresponding to the positions of SEQ ID NO: 1.

[0225] 13. The use of any of the preceding paragraphs, wherein the protease variant comprises the amino acid sequence of SEQ ID NO: 2 having the substitutions S3T+N43R+N76D+S87N+G118M+S128Q+N184E+V205I+Q206L+Y209W+S259D+N261W+L262E, wherein the variant has protease activity and wherein the positions are numbered corresponding to the positions of SEQ ID NO: 1.

[0226] 14. The use as described in any one of the preceding paragraphs, wherein the protease is an enzyme having or consisting of the amino acid sequence of SEQ ID NO: 2, 3, 4 or 5.

[0227] 15. The use of any of the preceding paragraphs, wherein the cellulase is an enzyme having at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% sequence identity to SEQ ID NO: 13, 14, 15, 16, 17, 18, 19, 20, 21, 22 or 23.

[0228] 16. The use of any of the preceding paragraphs, wherein the mannanase is an enzyme having at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% sequence identity to SEQ ID NO: 6, 7, 8, 9, 10, 11 or 12.

[0229] 17. A cleaning composition free of at least one whiteness maintenance agent, wherein the composition comprises at least two enzymes selected from the group consisting of proteases, mannanases and cellulases.

[0230] 18. The composition of paragraph 17, wherein the whiteness maintenance agent is selected from the group consisting of optical brighteners, anti-redeposition polymers and mixtures thereof, and preferably, the cleaning composition is a laundry detergent.

[0231] 19. The composition of paragraph 17 or 18, wherein the enzyme is as defined in any of the preceding paragraphs 4 to 16.

[0232] 20. The composition of any of the preceding paragraphs 17-19, wherein the whiteness performance of the composition is improved compared to a comparable or reference cleaning composition, wherein the reference cleaning composition comprises the at least one whiteness maintenance agent but does not comprise the enzyme.

[0233] 21. A method for maintaining or improving the whiteness and / or color of a textile, the method comprising: (a) exposing the textile to a wash liquor comprising a detergent composition as described in any of the preceding paragraphs 17-19; and optionally, (b) rinsing the textile.

[0234] The present invention is further illustrated in the following non-limiting examples.

[0235] Examples

[0236] Assay I: Test for protease activity

[0237] Proteolytic activity can be determined by a method using a Suc-AAPF-pNA substrate. Suc-AAPF-pNA is an abbreviation for N-succinyl-alanine-alanine-proline-phenylalanine-p-nitroaniline and is a closed peptide that can be cleaved by an endoprotease. After proteolytic cleavage, free pNA molecules with a yellow color are released and can be measured by visible light spectrophotometry at a wavelength of 405 nm. The Suc-AAPF-PNA substrate is manufactured by Bachem (Cat. No. L1400, dissolved in DMSO).

[0238] The protease samples to be analyzed were diluted in residual activity buffer (100 mM Tris pH 8.6). The assay was performed by transferring 30 μl of the diluted enzyme sample to a 96-well microtiter plate and adding 70 μl of substrate working solution (0.72 mg / ml in 100 mM Tris, pH 9). The solution was mixed at room temperature and the absorption was measured at OD 405 nm every 20 seconds for 5 minutes.

[0239] Under a given set of conditions, the slope of the time-dependent absorbance curve (absorbance / minute) is directly proportional to the activity of the protease in question. The protease sample should be diluted to a level where the slope is linear.

[0240] Assay II: Test for mannanase activity

[0241] Mannanase activity was determined by using an insoluble azo-carob-galactomannan substrate from Megazyme. The substrate and enzyme were incubated at 25°C for 20 min with shaking at 800 rpm. The reaction mixture was kept static for 10 min to allow the insoluble substrate to settle. Enzyme activity was measured by reading the optical density of the supernatant at 590 nm.

[0242] Assay III: Test for cellulase activity

[0243] Cellulase activity is determined as the ability of the enzyme to catalyze the hydrolysis of 1,4-β-D-glycosidic bonds in β-1,4-glucan (cellulose). For the purposes of the present invention, AZCL-HE-cellulose (from Magzyme) is used as a reaction substrate to determine cellulase activity. The substrate and enzyme are incubated at 30°C for 20min with shaking at 800rpm. The reaction mixture is kept static for 10min to allow the insoluble substrate to settle. Enzyme activity is measured by reading the optical density of the supernatant at 590nm.

[0244] Detergent:

[0245] Use standard detergent 1, ingredients:

[0246] 5.3% LAS, 10.7% AEOS, 1% soap, 5.3% nonionic surfactant AEO, 2% sodium citrate, 0.4% TEA, 0.73 NaOH, 0.02% CaCl2, and water to 100% (all percentages are w / w)

[0247] Use standard detergent 2, ingredients:

[0248] 21.0% LAS, 25.0% nonionic surfactant AEO, 15.0% monopropylene glycol, 9.0% soap, 8.0% glycerol, 6.8% monoethanolamine, 0.5% DTMPA, and water to 100% (all percentages are w / w).

[0249] Enzymes used in the examples

[0250] Protease: The polypeptide of SEQ ID NO:2, having the following mutations: S9E+N43R+N76D+V205I+Q206L+Y209W+S259D+N261W+L26E, wherein the position numbering is based on the numbering of SEQ ID NO:1.

[0251] Mannanase: A mannanase corresponding to the polypeptide of SEQ ID NO:7.

[0252] Cellulase A: A cellulase corresponding to the polypeptide of SEQ ID NO:18.

[0253] Cellulase B: A cellulase corresponding to the polypeptide of SEQ ID NO:21.

[0254] Cellulase C: A cellulase corresponding to the polypeptide of SEQ ID NO:14.

[0255] Terg-O-tometer (TOM) washing assay

[0256] Tergo-To-Meter (TOM) is a medium-scale model washing system, which can be applied to test 16 different washing conditions simultaneously. TOM is basically a large temperature-controlled water bath, in which up to 16 open metal beakers are immersed. Each beaker constitutes a small top-loading washing machine and during the experiment, each of them will contain a solution of a specific detergent / enzyme / polymer system and test its performance on soiled and unsoiled fabrics. Mechanical stress is obtained by rotating agitator arms, which stir the liquid in each beaker.

[0257] In a TOM experiment, factors such as the ballast to soil ratio and the fabric to wash liquor ratio can be varied. Thus, TOM provides a link between small-scale experiments and more time-consuming full-scale experiments.

[0258] Equipment: The water bath has 16 steel beakers and each beaker has a rotating arm, each beaker has a capacity of 1 L of detergent solution. The temperature range is from 5°C to 80°C. The water bath must be filled with deionized water. The rotation speed can be set from 70 to 120 rpm / min.

[0259] Set the temperature in the Terg-0-Tometer and start the rotation in the water bath. Wait for the temperature to adjust (tolerance is + / - 0.5°C). Prepare a washing solution or washing liquor with the desired amount of detergent, temperature and water hardness in a bucket. Allow the detergent to dissolve during magnetic stirring for 10 min. The washing solution should be used within 30 to 60 min after preparation.

[0260] 1L of washing solution is added to a TOM beaker. The washing solution or washing liquid is stirred at 120 rpm and one or more enzymes, optical brighteners or polymers are added to the beaker. Whiteness tracer is sprinkled into the beaker, followed by ballast loading. Time measurement begins when the swatch and ballast are added to the beaker. The swatch is washed for 20 or 30 minutes, after which stirring is stopped.

[0261] Subsequently, the wash load is transferred from the TOM beaker to the sieve and rinsed with cold tap water. The soiled swatches are separated from the ballast load. Under running water, the soiled swatches are transferred to a 5 L beaker containing cold tap water for 5 minutes. For the upcoming inactivation, the ballast load is stored separately. The water in the swatches is gently pressed out by hand and placed on a tray covered with paper. The swatches are allowed to dry overnight and then analyzed, such as measuring ΔREM.

[0262] Washing conditions and materials are listed in Tables 1-2 below.

[0263] Table 1: TOM washing conditions

[0264]

[0265] Table 2: List of white tracers

[0266]

[0267]

[0268] Light reflectance measurement

[0269] After washing and rinsing, the small pieces of cloth are spread out and allowed to air dry overnight at room temperature. Whiteness can be expressed as reflectance (R), which is the measure of light reflected or scattered from the test material when irradiated with white light. The reflectance of textiles is measured at 460nm using a Macbeth Color Eye 7000 reflection spectrophotometer with a very small aperture. The measurement is carried out without UV in the incident light, and the reflectance at 460nm (R460) is extracted. Measure according to the manufacturer's protocol. Whiteness performance is evaluated by R460 or ΔR460. ΔR460 can be calculated as follows: the R460 value of the tracer washed with detergent + other additives (OB, ARD polymers or enzymes) minus the R460 value of the tracer washed with a comparable detergent (e.g., a detergent without at least one of the additives).

[0270] Example 1: Whiteness performance evaluation of white tracer

[0271] The whiteness performance was evaluated by the TOM wash test as shown above. The wash results are summarized in Table 3 below. In the table below, the sum of R460 (or ΔR460) represents the sum of the R460 (or ΔR460) values ​​of the 9 washed white tracers (see Table 2). The higher the ΔR460, the better the whiteness performance / effect of the corresponding detergent composition.

[0272] Additionally, in the following Table 3, the whiteness performance brought about by the addition of one or more enzymes, or in other words, the whiteness effect (WE) of the enzyme replacing the whiteness maintaining agent is calculated as follows:

[0273] WE1 = sum of ΔR460 (detergent + enzyme(s)) minus sum of ΔR460 (detergent + OB + ARD polymer). Thus, "WE1" represents the whiteness improvement relative to a detergent comprising OB and ARD polymer but without the enzyme(s) of the present invention.

[0274] WE2 = Sum of ΔR460 (detergent + enzyme(s)) minus sum of ΔR460 (detergent + OB). Thus, "WE2" represents the whiteness improvement relative to a detergent containing an optical brightener but without the enzyme(s) of the present invention.

[0275] Table 3: Sum of reflectance and delta reflectance of white tracers washed under different conditions

[0276]

[0277]

[0278]

[0279] It is clear from Table 3 above that the addition of whiteness maintenance agents (such as OB or ARD polymers) can improve the whiteness performance of the detergent composition, in other words, can prevent / reduce the whiteness loss of textiles. Replacing the agents with a single enzyme (protease, mannanase or cellulase) of the present invention can further improve the whiteness performance of the detergent composition.

[0280] The results also show that the enzyme combination of protease, mannanase and / or cellulase provides even better whiteness performance than the individual enzymes. This clearly shows that there is a synergistic effect between two or three enzymes in improving the whiteness performance of detergent compositions.

[0281] Example 2: Whiteness performance evaluation of white tracer

[0282] Table 4: TOM washing conditions under European washing conditions

[0283]

[0284]

[0285] Table 5: List of white tracers that meet EU conditions

[0286]

[0287] The whiteness performance was evaluated by the TOM wash test as shown above. The wash results are summarized in Table 6 below. In the table below, the sum of R460 represents the sum of the R460 values ​​of the 7 washed white tracers (see Table 5). The higher the R460, the better the whiteness performance / effect of the corresponding detergent composition.

[0288] Additionally, in the following Table 6, the whiteness performance brought about by the addition of one or more enzymes, or in other words, the whiteness effect (WE) of the enzyme replacing the whiteness maintaining agent is calculated as follows:

[0289] WE3 = sum of R460 (detergent + enzyme(s)) minus sum of R460 (detergent + OB + cleaning polymer). Thus, "WE3" represents the whiteness improvement relative to a detergent comprising OB and cleaning polymer but without the enzyme(s) of the present invention.

[0290] WE4 = sum of R460 (detergent + enzyme(s)) minus sum of R460 (detergent + OB). Thus, "WE4" represents the whiteness improvement relative to a detergent containing an optical brightener but without the enzyme(s) of the present invention.

[0291] WE5 = sum of R460 (detergent + enzyme(s)) minus sum of R460 (detergent + cleaning polymer). Thus, "WE5" represents the whiteness improvement relative to a detergent comprising a cleaning polymer but without the enzyme(s) of the present invention.

[0292] Table 6: Sum of reflectance and delta reflectance of white tracers washed under different conditions

[0293]

[0294]

[0295] It is clear from Table 6 above that replacing whiteness maintenance agents (such as OB or cleaning polymer) with single enzymes (protease, mannanase or cellulase) of the present invention can maintain the same level of whiteness or further improve the whiteness performance of detergent compositions.

[0296] The results also show that the enzyme combination of protease, mannanase and / or cellulase provides even better whiteness performance than the individual enzymes. This clearly shows that there is a synergistic effect between two or three enzymes in improving the whiteness performance of detergent compositions.

[0297] Example 2: Whiteness performance evaluation with commercial detergents

[0298] Table 7: TOM washing conditions under European washing conditions using commercial detergents

[0299]

[0300]

[0301] The whiteness performance was evaluated by the TOM wash test as shown above. The wash results are summarized in Table 8 below. In the table below, the sum of R460 (or ΔR460) represents the sum of the R460 (or ΔR460) values ​​of the 7 washed white tracers (see Table 5). The higher the ΔR460, the better the whiteness performance / effect of the corresponding detergent composition.

[0302] Additionally, in the following Table 6, the whiteness performance brought about by the addition of one or more enzymes, or in other words, the whiteness effect (WE) of the enzyme replacing the whiteness maintaining agent is calculated as follows:

[0303] WE2 = Sum of ΔR460 (detergent + enzyme(s)) minus sum of ΔR460 (detergent + OB). Thus, "WE2" represents the whiteness improvement relative to a detergent containing an optical brightener but without the enzyme(s) of the present invention.

[0304] Table 8: Sum of reflectance and delta reflectance of white tracers washed under different conditions

[0305]

[0306] It is clear from the above table that adding a whiteness maintenance agent (such as OB) does not always improve the whiteness performance of the detergent composition, in other words, it may not prevent / reduce the whiteness loss of textiles. Replacing the agent with a combination of enzymes of the present invention (protease, mannanase and cellulase) can further improve the whiteness performance of the detergent composition.

Claims

1. Use of an enzyme for replacing a whiteness maintenance agent in a cleaning composition, wherein the enzyme is selected from the group consisting of proteases, mannanases, cellulases and combinations thereof.

2. The use as claimed in claim 1, wherein the whiteness maintenance agent is selected from the group consisting of optical brighteners, anti-redeposition polymers and mixtures thereof.

3. The use according to claim 1 or 2, wherein the replacement is partial or complete replacement.

4. Use as claimed in any one of the preceding claims, wherein the enzyme is a combination of a protease and a cellulase.

5. Use according to any one of the preceding claims, wherein the enzyme is a combination of a protease and a mannanase.

6. Use as claimed in any one of the preceding claims, wherein the enzyme is a combination of protease, mannanase and cellulase.

7. Use as described in any of the preceding claims, wherein the protease has at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% sequence identity with the polypeptide of SEQ ID NO: 1, 2, 3, 4 or 5.

8. The use of any of the preceding claims, wherein the protease is a variant of the polypeptide of SEQ ID NO: 2 comprising an alteration at one or more positions corresponding to positions 3, 4, 9, 15, 43, 68, 76, 99, 101, 103, 104, 160, 167, 170, 194, 199, 205, 206, 209, 217, 218, 222, 245, 261 and 262, wherein the position numbers correspond to positions of SEQ ID NO: 1, wherein each alteration is independently a substitution, deletion or insertion, and wherein the variant has protease activity and has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99%, but less than 100% sequence identity with the polypeptide of SEQ ID NO:

2.

9. Use as claimed in any one of the preceding claims, wherein the protease is: (a) a variant of the polypeptide of SEQ ID NO: 2, comprising one or more substitutions selected from the group consisting of S3T, V4I, S9E, S9R, A15T, V68A, N76D, S99D, S99G, S99A, S99SE, S101E, S101N, S101R, S103A, V104I, G160S, Y167A, R170S, A194P, V199M, V205I, Q206L, Y209W, L217D, L217Q, N218D, M222S, Q245R, N261W and L262E, wherein the position numbers correspond to the positions of SEQ ID NO: 1, and wherein the variant has protease activity and is identical to the polypeptide of SEQ ID NO:

2. NO:2 has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, but less than 100% sequence identity; (b) a variant of the polypeptide of SEQ ID NO:2, comprising the substitutions N76D+Y209W, wherein the position numbers correspond to the positions of SEQ ID NO:1, and wherein the variant has protease activity and has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99%, but less than 100% sequence identity to SEQ ID NO:2; (c) a variant of the polypeptide of SEQ ID NO:2 comprising the substitution S87N, wherein the variant has protease activity and wherein the position corresponds to the position of SEQ ID NO:1, and wherein the variant has protease activity and has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100% sequence identity to SEQ ID NO:2; (d) a variant of the polypeptide of SEQ ID NO:2, comprising the substitutions Y167A+R170S+A194P, wherein the position numbers correspond to the positions of SEQ ID NO:1, and wherein the variant has protease activity and has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99%, but less than 100% sequence identity to SEQ ID NO:2; (e) a variant of the polypeptide of SEQ ID NO:2, comprising the substitutions S9E+N43R+N76D+V205I+Q206L+Y209W+S259D+N261W+L262E, wherein the positions are numbered corresponding to the positions of SEQ ID NO:1, and wherein the variant has protease activity and has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100% sequence identity to SEQ ID NO:2; (f) a variant of the polypeptide of SEQ ID NO:4, comprising the substitutions A68S+T77N+T78I+G127S+A128P+G165Q+N184Q+A202V+N217S+S258P, wherein the position numbers correspond to the positions of SEQ ID NO:4, and wherein the variant has protease activity and has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100% sequence identity to SEQ ID NO:4; (g) a variant comprising a substitution at one or more positions corresponding to position 171, 173, 175, 179 or 180 of SEQ ID NO: 1 of WO2004 / 067737, wherein the variant has protease activity and has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99%, but less than 100% sequence identity to SEQ ID NO: 1 of WO2004 / 067737; (h) a protease variant comprising one or more substitutions compared to a parent protease, the one or more substitutions being selected from the group consisting of: X3V, X9[E,R], X22[R,A], X43R, X61[E,D], X62[E,D], X76[D], X87N, X101[E,G,D,N,M], X103A, X104I, X118[V,R], X120V, X128[A,L,S], X129Q, X130 30A, X160D, X185[E,D], 188[E,D], X191N, X194P, X205I, X206L, X209W, X216V, X217[Q,D,E], X218[D,E,S], X232V, X245R, X248D, X256[E,D], X259[E,D], X261[E,D,W], and X262[E,D], where the position numbers correspond to BPN'(SEQ ID NO: ... ID NO:1), wherein "X" represents any amino acid residue present in the specified position in the parent protease, and wherein the variant has protease activity and has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99%, but less than 100% sequence identity to SEQ ID NO:1 or SEQ ID NO:2; and (i) a protease variant comprising any of the following substitution groups compared to a parent protease, wherein the parent protease has an amino acid sequence of SEQ ID NO: 1 or SEQ ID NO: 2 or has at least 80% sequence identity to SEQ ID NO: 1 or SEQ ID NO: 2, wherein the position number corresponds to the position of BPN' (SEQ ID NO: 1), wherein "X" represents any amino acid residue present in the specified position in the parent protease, and wherein the substitution group is selected from the group consisting of: i.X9R+X15T+X68A+X218D+X245R, ii.X9R+X15T+X68A+X245R, iii.X61E+X194P+X205I+X261D, iv.X61D+X205I+X245R, v.X61E+X194P+X205I+X261D, vi.X87N+X118V+X128L+X129Q+X130A, vii.X87N+X101M+X118V+X128L+X129Q+X130A, viii.X76D+X87R+X118R+X128L+X129Q+X130A, ix.X22A+X62D+X101G+X188D+X232V+X245R, x.X103A+X104I, xi.X22R+X101G+X232V+X245R, xii.X103A+X104I+X156D, xiii.X103A+X104I+X261E, xiv.X62D+X245R, xv.X101N+X128A+X217Q, xvi.X101E+X217Q, xvii.X101E+X217D, xviii.X9E+X43R+X262E, xix.X76D+X43R+X209W, xx.X205I+X206L+X209W, xxi.X185E+X188E+X205I, xxii.X256D+X261W+X262E, xxiii.X191N+X209W, xxiv.X261E+X262E, xxv.X261E+X262D, and xxvi.X167A+X170S+X194P, wherein the protease variant has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99%, but less than 100% sequence identity to SEQ ID NO: 1 or 2.

10. Use as claimed in any one of the preceding claims, wherein the protease variant comprises the amino acid sequence of SEQ ID NO: 2 with the substitutions Y167A+R170S+A194P, wherein the variant has protease activity and wherein the position numbers correspond to the positions of SEQ ID NO:

1.

11. The use of any one of the preceding claims, wherein the protease variant comprises the amino acid sequence of SEQ ID NO: 2 having the substitutions S9E+N43R+N76D+V205I+Q206L+Y209W+S259D+N261W+L262E, wherein the variant has protease activity and wherein the positions are numbered corresponding to the positions of SEQ ID NO:

1.

12. Use as claimed in any one of the preceding claims, wherein the protease is an enzyme having or consisting of the amino acid sequence of SEQ ID NO: 2, 3, 4 or 5.

13. Use as described in any of the preceding claims, wherein the cellulase is an enzyme having at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% sequence identity to SEQ ID NO: 13, 14, 15, 16, 17, 18, 19, 20, 21, 22 or 23.

14. Use as described in any of the preceding claims, wherein the mannanase is an enzyme having at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% sequence identity to SEQ ID NO: 6, 7, 8, 9, 10, 11 or 12.

15. A cleaning composition free of at least one whiteness maintenance agent, wherein the composition comprises an enzyme selected from the group consisting of proteases, mannanases, cellulases, and combinations thereof.

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