CN101370934B - Detergent compositions - Google Patents

Abstract

The present invention relates to detergent compositions comprising a detergent ingredient and a lipase variant with reduced potential for odor generation obtained by introducing mutations in one or more regions identified in a parent lipase.

Description

Detergent composition

Invention field

The present invention relates to detergent composition, especially comprise the laundry detergent of lipolytic enzyme.

Background of invention

To detergent manufacturers, the especially manufacturers of clothes washing industry, improving the oily dirt removal effect is its constant target.Although used the combination of many effective tensio-active agents and tensio-active agent, surfactants based product still can't be realized the fully removal of fats/oils dirt, especially when product uses under low water temperature.Lipase just is used in the washing composition from the later stage 1980s, in order to remove fatty dirt by fatty dirt is resolved into triglyceride level.

Until not long ago, main commercially available lipase such as Lipolase (trade(brand)name, Novozymes) just can the baking stage of washing process than producing especially effectively effect under the low moisture content.These enzymes only are tending towards producing significant cleaning effect at the second washing step, because the dirt that only stays at the washed clothing of baking stage just significant steatolysis can occur, then remove the fat of decomposition in next washing step.Yet developed recently more efficient lipase, its washing stage at cleaning course also can work effectively.Therefore the cleaning action in second washing step, owing to lipase can be present in first cycles of washing, so cleaning effect is significantly improved.US 6,939, described the example of these enzymes among 702 B1, WO00/60063 and the Research DisclosureIP6553D.Hereinafter this kind of enzyme is called the first washing lipase.

In addition, the human consumer wishes that goods (such as clothes) are clean as much as possible.The smell of the goods that these human consumers usually will clean or process is associated with the clean-up performance of these goods.Therefore, from human consumer's viewpoint, cleaning and/or the validity for the treatment of compositions directly are associated with the smell that said composition is given the goods that clean or processed with said composition usually.The applicant has recognized that Cucumber for example esterase and lipase can produce unpleasant lipid acid smell, especially short chain fatty acid smell, for example smell of butyric acid.Yet these materials can be especially effective cleaning agent.Regrettably, the human consumer usually will be owing to the smell that uses these reagent to produce is associated with inadequate cleaning.Have the variant example terminal extension sequence of C-, that reduce smell referring to WO02/062973, but these lipase Variants do not show the potent scourability of the first washing lipase, the lipase such as among those WO00/60063 comprises with trade(brand)name

The variant of selling.

Therefore, need a kind of detergent composition that comprise lipolytic enzyme obtaining excellent fats/oils dirt removal effect, and do not produce any tedious lipid acid smell.

Summary of the invention

The present invention relates to comprise the detergent composition of detergent ingredients and lipase Variant, the smell that described lipase Variant has minimizing produces trend, and without the terminal extension sequence of C-.Lipase Variant is introduced sudden change by one or more zones of determining and is obtained in parent lipase.

Description of drawings

Fig. 1 represents the lipase sequence alignment.

Sequence list

Sequence identification number: the dna sequence dna of 1 presentation code thermophilic fungus lipase.

Sequence identification number: the aminoacid sequence of 2 expression thermophilic fungus lipase.

Sequence identification number: the aminoacid sequence of the mould lipase of 3 expression colters.

Sequence identification number: the aminoacid sequence of 4 expression absidia corymbifera lipase.

Sequence identification number: the aminoacid sequence of 5 expression rhizomucor miehei lipases.

Sequence identification number: the aminoacid sequence of 6 expression Rhizopus oryzae lipase.

Sequence identification number: the aminoacid sequence of 7 expression lipase from Aspergillus Nigers.

Sequence identification number: the aminoacid sequence of 8 expression Tabin aspergillus lipase.

Sequence identification number: the aminoacid sequence of 9 expression Fusarium oxysporum lipase.

Sequence identification number: the aminoacid sequence of 10 expression fusarium heterosporium lipase.

Sequence identification number: the aminoacid sequence of 11 expression aspergillus oryzae lipase.

Sequence identification number: the aminoacid sequence of 12 expression penicillium camembertii lipase.

Sequence identification number: the aminoacid sequence of the smelly aspergillus niger lipase enzyme of 13 expressions.

Sequence identification number: the aminoacid sequence of 14 expression lipase from Aspergillus Nigers.

Sequence identification number: the aminoacid sequence of 15 expression aspergillus oryzae lipase.

Sequence identification number: the aminoacid sequence of 16 expression Landerina penisapora lipase.

Detailed Description Of The Invention

Lipase Variant

Parent lipase

Parent lipase can be fungal lipase, and described in " homology and sequence alignment " part, its aminoacid sequence has and sequence identification number: the homology of the thermophilic fungus lipase sequence at least 50% that shows in 2.

Parent lipase can be yeast polypeptides, for example candiyeast, kluyveromyces, pichia spp, yeast, fission yeast or Ye Shi yeast polypeptides; Or filamentous fungus polypeptide more preferably, for example top spore mould, aspergillus tubigensis, short stalk mould, cryptococcus, line black powder yeast, sickle mycete, humic mould, Pyricularia oryzae, mucormycosis, ruin a mould, Xin Kaoma fat mould, neurospora, paecilomycerol, Penicillium notatum, Rumen Fungi, Split-gill, yellow silk aspergillus tubigensis, thermophilic ascomycete, grass roots mould, Trichoderma or Trichoderma polypeptide.

One preferred aspect, parent lipase is Ka Ersibai yeast, cereuisiae fermentum, saccharomyces diastaticus, Douglas yeast, kluyveromyces, Nuo Shi yeast or the ellipsoideus yeast polypeptide with lipase activity.

Another preferred aspect, parent lipase is microorganism Aspergillus aculeatus, Aspergillus awamori, Aspergillus fumigatus, smelly aspergillus, aspergillus japonicus, Aspergillus nidulans, aspergillus niger, aspergillus oryzae, Tabin aspergillus, sickle spore bacterioide, the wheat wilt, Fusariumsp, fusarium culmorum, Fusarium graminearum, the red sickle spore of standing grain, fusarium heterosporium, the five-leaved chaste tree sickle-like bacteria, Fusarium oxysporum, netted sickle-like bacteria, the moniliform sickle-like bacteria, fusarium sambucinum, colour of skin sickle spore, Fusarium sporotrichioides, fusarium sulphureum, bunch capsule sickle-like bacteria, fusarium trichothecioides, fusarium, Humicola insolens, thermophilic fungus (synonym: the high temperature chactomium globosum), the rice black wool is mould, thermophilic fungus destroyed wire, Neurospora, penicillium purpurogenum, trichoderma harziarum, healthy and free from worry wood is mould, long shoot wood is mould, Trichodermareesei, or viride polypeptide.

Another preferred aspect, parent lipase is that thermophilic fungus belongs to lipase.

One preferred aspect, parent lipase is thermophilic fungus lipase.In a preferred embodiment, parent lipase is sequence identification number: 2 lipase.

Zone identification and replacement

The regional I that hereinafter mentions is sequence identification number to the site of regional IV: the amino-acid residue site in 2.Use the described method of " homology and sequence alignment " part to find in the different lipase corresponding (or homology) site.

Replacement among the I of zone

Zone I is comprised of the amino-acid residue around N-terminal residue E1.Preferably in this zone replace amino acid in the parent lipase with the amino acid with more positive electricity.Zone I comprises the amino-acid residue in corresponding following site: 2 to 11 and 223 to 239.Following site has special significance: 4,8,11,223,227,229,231,233,234,236.Specifically, following replacement: X4V, X227G, X231R and X233R have been confirmed.

In a preferred embodiment, parent lipase and sequence identification number: 2 have at least 80%, such as 85% or 90%, and the identity such as at least 95% or 96% or 97% or 98% or 99%.In a preferred embodiment, parent lipase and sequence identification number: 2 is identical.

Replacement among the II of zone

Zone II is comprised of the amino-acid residue that contacts substrate on the one side of acyl chain with one side at alcohol moiety.The preferred amino acid with replacing with the amino acid of more positive electricity or low hydrophobic amino acid in the parent lipase in this zone.Zone II comprises the amino-acid residue in corresponding following site: 202 to 211 and 249 to 269.Following site has special significance: 202,210,211,253,254,255,256.Specifically, confirmed following replacement: X202G, X210K, X255Y/V and X256K/R.

In a preferred embodiment, parent lipase and sequence identification number: 2 have at least 80%, such as 85% or 90%, and the identity such as at least 95% or 96% or 97% or 98% or 99%.In a preferred embodiment, parent lipase and sequence identification number: 2 is identical.

Replacement among the III of zone

Zone III forms thereby the permission substrate enters the amino-acid residue of avtive spot by forming flexible structure.The preferred amino acid with replacing with the amino acid of more positive electricity or low hydrophobic amino acid in the parent lipase in this zone.Zone III comprises the amino-acid residue in corresponding following site: 82 to 102.Following site has special significance: 83,86,87,90,91,95,96,99.Specifically, following replacement: X83T, X86V and X90A/R have been confirmed.

In a preferred embodiment, parent lipase and sequence identification number: 2 have at least 80%, such as 85% or 90%, and the identity such as at least 95% or 96% or 97% or 98% or 99%.In a preferred embodiment, parent lipase and sequence identification number: 2 is identical.

Replacement among the IV of zone

Zone IV is comprised of the amino-acid residue of electrostatical binding to the surface.Preferably in this zone replace amino acid in the parent lipase with the amino acid with more positive electricity.Zone IV comprises the amino-acid residue in corresponding following site: 27 and 54 to 62.Following site has special significance: 27,56,57,58,60.Specifically, following replacement: X27R, X58N/AG/T/P and X60V/S/G/N/R/K/A/L have been confirmed.

In a preferred embodiment, parent lipase and sequence identification number: 2 have at least 80%, such as 85% or 90%, and the identity such as at least 95% or 96% or 97% or 98% or 99%.In a preferred embodiment, parent lipase and sequence identification number: 2 is identical.

The amino acid in other site

Parent lipase optionally comprises other amino acid whose replacement, especially is less than 10 or be less than 5 such replacements.Example is the replacement in the one or more sites in the corresponding parent lipase 24,46,74,81,83,127,131,137,147,150,203,206,211,263,264,265,267 and 269.In a specific embodiment, replace one of them site that occurs in corresponding 81,147,150 and 249 sites.In a preferred embodiment, at least one replacement is selected from the group that is comprised of following replacement: X81Q/E, X147M/Y, X150G and X249R/I/L.

For example, can more replace the replacement described in WO 92/05249, WO 94/25577, WO 95/22615, WO 97/04079 and WO 97/07202 according to rule known in the art.

Parent's fat variant

In one aspect, when comparing with described parent, described variant comprises at least three replacements of total, and described replacement is selected from following one or more groups replacement:

A) at least two kinds of replacements in regional I,

B) at least a replacement in regional II,

C) at least a replacement in regional III, and/or

D) at least a replacement in regional IV.

Variant is compared with the parent of variant can comprise replacement, those replacements of listing in the corresponding following table 1 of described replacement.

Zone I	Zone II	Zone III	Zone IV	Outside the zone
					X4V+X227G+X231R+ X233R	X210K+ X256K	X83T+ X86V	X58A+X60S	X150G
X227G+X231R+X233R	X256K	X86V	X58N+X60S	X150G
					X231R+X233R	X255Y
X231R+X233R	X202G
					X227G+X231R+X233R	X256K	X86V
X4V+X231R+X233R			X58N+X60S
					X231R+X233R		X90R	X58N+X60S
X231R+X233R	X255V	X90A
					X227G+X231R+X233R	X256K	X86V	X58N+X60S	X150G
X231R+X233R	X211L		X58N+X60S	X147M

Table 1: the variant that some are concrete.

Parent lipase and sequence identification number: 2 is identical more specifically among the embodiment at one, thereby the variant of table 1 will be:

Zone I	Zone II	Zone III	Zone IV	Outside the zone
					Q4V+L227G+T231R+ N233R	E210K+ P256K	S83T+ I86V	S58A+V60S	A150G
L227G+T231R+N233R	P256K	I86V	S58N+V60S	A150G
					T231R+N233R	1255Y
T231R+N233R	I202G
					L227G+T231R+N233R	P256K	I86V
Q4V+T231R+N233R			S58N+V60S
					T231R+N233R		I90R	S58N+V60S
T231R+N233R	1255V	I90A
					L227G+T231R+N233R	P256K	I86V	S58N+V60S	A150G
T231R+N233R	F211L		S58N+V60S	L147M

Table 2: sequence identification number: the concrete variant of some of 2

Amino acid modified name

When describing as described in the present invention lipase Variant, use following nomenclature to be beneficial to reference: initial amino acid: site: substituted amino acid.

According to this nomenclature, for example the L-glutamic acid with 195 places, site is substituted by glycine, is expressed as G195E.Words at identical site disappearance glycine are expressed as G195 ^*, be expressed as G195GK and insert additional amino-acid residue such as Methionin.Compare with other lipase for one, comprise one in the site " disappearance " amino acid of 36, the concrete lipase that inserts simultaneously an aspartic acid in this site is expressed as ^*36D.A plurality of sudden changes are separated with plus sige, that is: R170Y+G195E is illustrated in site 170 and 195 places and respectively tyrosine and L-glutamic acid is substituted by arginine and glycine.

When using described sequence alignment method, X231 is illustrated in the amino acid in corresponding site 231 in parent's polypeptide.X231R represents that this amino acid replaces with R.Concerning sequence identification number: 2 X are T, and therefore be illustrated in 231 places, site replaces T with R to X231R.When the amino acid of a site (for example 231) can be selected from one group of amino acid whose aminoacid replacement with another, for example, described group was comprised of R and P and Y, represents with X231R/P/Y.

In all cases, use IUPAC single-letter or the trigram amino acid abbreviations of generally acknowledging.

The amino acid grouping

In this manual, amino acid is divided into electronegative, positively charged or electroneutral amino acid according to its electric charge under the pH10 condition.Therefore, electronegative amino acid is E, D, C (halfcystine) and Y, especially E and D.The amino acid of positively charged is R, K and H, especially R and K.When forming disulfide linkage, neutral amino acids is G, A, V, L, I, P, F, W, S, T, M, N, Q and C.Be called as conservative the replacement by the another kind of aminoacid replacement in same group (electronegative, positively charged or electric neutrality).

That neutral amino acids can be divided into is hydrophobic or nonpolar (G, A, V, L, I, P, F, W and as the C of the part of disulfide linkage) and (S, T, M, N, the Q) of hydrophilic or polarity.

Amino acid identity

With parameter " identity " dependency between two aminoacid sequences or two nucleotide sequences is described.

For purposes of the invention, by using the comparison from two aminoacid sequences of Needle program determination of EMBOSS package (http://emboss.org) 2.8.0 version.The Needle program is used the overall comparison algorithm, and this arthmetic statement is in Needleman, S.B. and Wunsch, C.D. (1970) J.Mol.Biol.48,443-453.The substitution matrix that uses is BLOSUM62, and the open point penalty in room is 10, and it is 0.5 that point penalty is expanded in the room.

Aminoacid sequence of the present invention (" invention sequence "; Sequence identification number for example: 2 amino acid/11 to 269) and the identity degree between the difference aminoacid sequence (" external sequence ") calculate by the following method: the exact matching number of two sequence alignments is divided by the length of " invention sequence " or the length of " external sequence ", and the length of whichever sequence is lacked most.The result is expressed as identity per-cent.

When " invention sequence " and " external sequence " when the same loci of lap has identical amino-acid residue, exact matching can occur.Sequence length is the number (for example sequence identification number: 2 length is 269) of the amino-acid residue in the sequence.

Parent lipase and thermophilic fungus lipase (sequence identification number: 2) have at least 50%, especially at least 55%, at least 60%, at least 75%, at least 85%, at least 90%, surpass 95% or surpass 98% amino acid identity.In a specific embodiment, parent lipase and thermophilic fungus lipase (sequence identification number: 2) identical.

Can use aforesaid method calculating identity and homology and be used for sequence alignment.Homology and sequence alignment are calculated as follows in situation of the present invention.

Homology and sequence alignment

For purposes of the invention, by computer program known in the art, such as GAP (Program Manual for the Wisconsin Package, the 8th edition, in the August, 1994 that provides in the GCG routine package, Genetics Computer Group, 575 Science Drive, Madison, Wisconsin, USA 53711) (Needleman, S.B. and Wunsch, C.D., (1970), " Journal of Molecular Biology ", 48,443-45), use to have the GAP that following peptide sequence contrast arranges, measuring aptly homology degree: GAP generation point penalty is 3.0, and GAP extension point penalty is 0.1.

In the present invention, (synonym: corresponding the high temperature chactomium globosum) and in the Landerina penisapora lipase sequence (or homology) site is determined by the sequence alignment that shows among Fig. 1 for mould, the absidia corymbifera of colter, rhizomucor miehei, Dai Shi head mold, aspergillus niger, Tabin aspergillus, Fusarium oxysporum, fusarium heterosporium, aspergillus oryzae, penicillium camembertii, smelly aspergillus, aspergillus niger, thermophilic fungus.

In order to find the homologous site that is not presented in the lipase sequence in the sequence alignment, the sequence that shows among the sequence paid close attention to and Fig. 1 is compared.Carry out the GAP sequence alignment by the highest sequence of homology that the GAP program is found, the existing sequence among new sequence and Fig. 1 is compared.(August 1994 for Program Manual for the Wisconsin Package, Version8 for the GCG routine package, Genetics Computer Group, 575 Science Drive, Madison, Wisconsin, USA 53711) (Needleman, S.B. and Wunsch, C.D., (1970), Journal of Molecular Biology, 48,443-45) provide GAP.It is 3.0 that peptide sequence relatively uses following setting: GAP to produce point penalty, and GAP extension point penalty is 0.1.

Parent lipase and thermophilic fungus lipase (sequence identification number: 2) have at least 50%, especially at least 55%, at least 60%, at least 75%, at least 85%, at least 90%, surpass 95% or surpass 98% amino acid identity.In a specific embodiment, parent lipase and thermophilic fungus lipase (sequence identification number: 2) identical.

Hybridization

The present invention also relates to by polynucleotide encoding, isolated polypeptide with lipase activity, described polynucleotide are under low-down preciseness condition, preferably under low preciseness condition, more preferably under middle preciseness condition, more preferably in-Gao preciseness condition under, even more preferably under high preciseness condition, most preferably under very high preciseness condition, hybridize with following nucleotide sequences: (i) Nucleotide 178 to 660, sequence identification number: 1, (ii) be included in the cDNA sequence of Nucleotide 178 to 660, sequence identification number: 1, (iii) (i) or sequence (ii), or (iv) (i), (ii) or complementary strand (J.Sambrook (iii), E.F.Fritsch, and T.Maniatus, 1989, MolecularCloning, A Laboratory Manual, second edition, Cold Spring Harbor, New York).Sequence identification number: 1 subsequence comprises at least 100 in abutting connection with Nucleotide, or preferably at least 200 in abutting connection with Nucleotide.In addition, the subsequence codified has the polypeptide fragment of lipase activity.

Concerning length is the long probe of at least 100 Nucleotide, prehybridization and the hybridization of very high preciseness conditional definition for carrying out under the following conditions will be low to moderate very much: 42 ℃, 5X SSPE, 0.3%SDS, that 200 μ g/mL have sheared and the milt DNA of sex change, perhaps 25% formyl ammonia is used for very low and low preciseness condition, 35% formyl ammonia is used for neutralization-Gao preciseness condition, perhaps 50% formyl ammonia is used for high and very high preciseness condition, standard Southern hybridization program hybridization most preferably 12 to 24 hours.

Concerning length is the long probe of at least 100 Nucleotide, use 2X SSC, 0.2%SDS, preferably at least 45 ℃ (low-down preciseness), more preferably at least 50 ℃ (low preciseness), more preferably at least 55 ℃ (middle preciseness), more preferably at least 60 ℃ (in-Gao preciseness).Even more preferably at least 65 ℃ (high preciseness), most preferably at least 70 ℃ (very high preciseness), last wash vehicle material three times, each 15 minutes.

Dna sequence dna, expression vector, host cell, lipase production

The invention provides code book invention lipase dna sequence dna, comprise the expression vector of described dna sequence dna and comprise the transformed host cell of described dna sequence dna or described expression vector.They can obtain by methods known in the art.

The present invention also provides the method for producing lipase, and this method is by cultivating transformant and regain lipase yielding lipase in next life from the gained broth culture under being of value to the condition of producing lipase.Described method is carried out according to principle known in the art.

Lipase activity

-under neutral pH (LU) condition to the lipase activity of tributyrin

Use Sudan Gum-arabic as emulsifying agent, emulsification tributyrin (tributyrin) prepares the lipase substrate.Then in the constant burette test of pH-, at 30 ℃, pH 7 or pH9 Water Under solution tributyrin.One unit lipase activity (1LU) is equivalent to can be under the pH7 condition, and per minute discharges the enzyme amount of 1 micromole's butyric acid.

-effect danger

With describe performance and smell reduce risks than effect danger Coefficient Definition be: BR=RP _Avg/ R.Lipase Variant as herein described can have greater than 1, greater than 1.1, or even greater than the dangerous coefficient of 1 to about 1000 effect.

-average relative performance

The program of calculating average relative performance (RPavg) is present in the example 5 of this specification sheets.Lipase Variant as herein described can have at least 0.8, at least 1.1, at least 1.5, or even at least 2 to about 1000 average relative performance (RPavg).

Detergent ingredients

Detergent composition used herein comprises goods and cleaning and treatment compositions.Except as otherwise noted, term " cleaning and/or treatment compositions " multi-functional or " heavy duty type " washing composition, especially laundry detergent of tablet, granule type or powder-type that comprise as used herein; Liquid, gel or paste type multifunctional detergent, especially so-called heavy-filth liquid type; Liquid high-count fabric washing composition; The washing composition of detergent for washing dishware with hand or light-duty dishwashing agent, especially those high alveolitoids; Dishwashing detergent for machine washing comprises the washing composition of multiple tablet form, granule type, liquid-type and rinse aid type, is used for household and enterprise.Described composition also can comprise those unit dose packagings known in the art in unit dose packaging, and those water miscible, water-insoluble and/or permeable unit dose packagings of water.

Detergent composition of the present invention can comprise one or more lipase Variants of the present invention.Except lipase Variant, detergent composition also comprises detergent ingredients.The non-limiting tabulation of detergent ingredients described below is applicable to cleaning compositions of the present invention, and be fit to it is joined among some embodiment of the present invention, for example to promote or to improve clean-up performance, process substrate to be cleaned or usually to improve the aesthetic property of cleaning compositions with tinting material, dyestuff etc.The physical form that the clear and definite character of these annexing ingredients and incorporation thereof will depend on composition with and the character of the clean operation used.Suitable detergent ingredients includes but not limited to tensio-active agent, washing assistant, sequestrant, dye transfer inhibitor, dispersion agent, enzyme and enzyme stabilizers, bleach-activating agent, hydrogen peroxide, hydrogen peroxide cource, preformed peracid, polymeric dispersant, whitening agent suds suppressor, dyestuff, corrosion inhibitor, tarnish inhibitor, spices, fabric softener, carrier, hydrotropic agent, processing aid, solvent and/or pigment.

Typical washing composition will comprise any combination weight 5% to 30% tensio-active agent by following composition, and the preferred anionic tensio-active agent is such as linear alkylbenzene sulfonate and hydroxyl epoxythio hydrochlorate; 0.005% to 0.1% protease activity albumen, wherein said proteolytic enzyme is preferably selected from Coronase ^TM, FN4FNA or Savinase ^TM, 0.001-0.1% amylase activity albumen, wherein said amylase is preferably selected from Termamyl ^TMNatalase ^TM, Stainzyme ^TMAnd Purastar ^TM, and 0.1% to 3% sequestrant, preferred DTPA.For granule type and tablet product, these typical washing composition will comprise by weight 5% to 20% SYNTHETIC OPTICAL WHITNER in addition, preferred SPC-D; 1% to 4% bleach-activating agent, preferred TAED and/or 0% to 30% washing assistant, preferred 5% to 30%, the washing assistant more preferably less than 10% is such as aluminosilicate zeolite A and/or tri-polyphosphate.

SYNTHETIC OPTICAL WHITNER-detergent composition of the present invention can comprise one or more SYNTHETIC OPTICAL WHITNER.

Usually, when using SYNTHETIC OPTICAL WHITNER, composition of the present invention can comprise by the weighing scale of this theme cleaning compositions about 0.1% to about 50%, perhaps even about 0.1% to about 25% SYNTHETIC OPTICAL WHITNER.The example of suitable SYNTHETIC OPTICAL WHITNER comprises:

(1) hydrogen peroxide cource, for example inorganic over hydrogenation adduct salt comprises following an alkali metal salt such as sodium salt: perborate (being generally monohydrate or tetrahydrate), percarbonate, persulphate, superphosphate, persilicate and their mixture.In one aspect of the invention, inorganic over hydrogenation adduct salt is selected from the group that is comprised of following material: peroxyboric acid sodium salt, percarbonic acid sodium salt and their mixture; Soap; With

(2) have the bleach-activating agent of R-(C=O)-L, wherein R is alkyl, optional branched-chain alkyl.When bleach-activating agent when being hydrophobic, it has 6 to 14 carbon atoms or 8 to 12 carbon atoms.When bleach-activating agent when being hydrophilic, its have less than 6 carbon atoms or even less than 4 carbon atoms; And L is leavings group.The example of suitable leavings group is phenylformic acid and derivative thereof, especially benzene sulfonate.Suitable bleach-activating agent comprises dodecane acyl-oxygen base benzene sulfonate, last of the ten Heavenly stems acyloxy benzene sulfonate, caprinoyl aminobenzoic acid and salt, 3 thereof; 5,5-trimethyl acetyl oxygen base benzene sulfonate, tetra acetyl ethylene diamine (TAED) and nonanoly acyloxy benzene sulfonate (NOBS).Suitable bleach-activating agent also is disclosed among the WO 98/17767.Although can adopt any suitable bleach-activating agent, in one aspect of the invention, the cleaning compositions of this theme can comprise NOBS, TAED or their mixture.

(3) preformed peracid.

If exist, peracid and/or bleach-activating agent usually with based on described composition about 0.1 to about 10% weight, about 0.5 to about 60% weight or even about 0.6 content to about 40% weight be present in the described composition.Its one or more hydrophilic precursor can be united use with one or more hydrophilic peracid or its precursor.

Can select the amount of hydrogen peroxide cource and peracid or bleach-activating agent, so that the mol ratio of available oxygen (from peroxide source) and peracid is 1: 1 to 35: 1, perhaps even 2: 1 to 10: 1.

Tensio-active agent-detergent composition can comprise tensio-active agent or surfactant system as described in the present invention, and wherein said tensio-active agent can be selected from nonionogenic tenside, anion surfactant, cats product, amphoterics, zwitterionics, semi-polar nonionic surfactants and their mixture.If exist, the content of tensio-active agent is generally about 0.1% to about 60%, about 0.1% to about 40%, about 0.1% to about 12%, about 1% to about 50% by the weighing scale of tested composition, or even about 5% to about 40%.

If exist, described washing composition will comprise about 1% to about 40% anion surfactant usually, such as linear alkylbenzene sulfonate, alhpa olefin sulfonate, alkyl-sulphate (aliphatic alcohol sulfate), hydroxyl epoxythio hydrochlorate, secondary alkyl sulfonate, α sulfo methyl ester, alkyl or alkenyl Succinic Acid or soap.

Described washing composition optionally comprises about 0.2% to about 40% nonionogenic tenside, such as alcohol ethoxylate, nonyl phenol ethoxylate, alkyl polyglycoside, alkyl dimethyl amine oxide, ethoxylated fat enzyme one glycollic amide, lipid acid one glycollic amide, polyhydroxy alkyl fatty acid amide or glycosamine N-acyl group N-alkyl derivative (" glucamide ").

Washing assistant-detergent composition of the present invention can comprise one or more detergent builder or builder system.When using washing assistant, this theme composition will comprise weighing scale by this theme composition usually at least about 1%, and about 5% to about 60%, perhaps even about 10% to about 40% washing assistant.Washing assistant includes but not limited to various an alkali metal salts, ammonium salt and the substituted ammonium salt of basic metal, ammonium and alkanol ammonium poly-phosphate, alkalimetal silicate or layered silicate, alkaline earth and alkaline carbonate, silico-aluminate washing assistant, polynary acetic acid (such as ethylenediamine tetraacetic acid (EDTA) and nitrilotriacetic acid(NTA)), and polycarboxylate is (such as mellitic acid, succsinic acid, citric acid, oxygen di-succsinic acid, polynary toxilic acid, benzene-1,3,5-tricarboxylic acid, carboxymethyl oxosuccinic acid) and their soluble salt.

The detergent composition of sequestrant-this paper can comprise sequestrant.Suitable sequestrant comprises copper, iron and/or manganese sequestrant and their mixture.When using sequestrant, this theme composition can comprise by the weighing scale of this theme composition about 0.005% to about 15%, or even about 3.0% to about 10% sequestrant.

Whitening agent-detergent composition of the present invention also can comprise the annexing ingredient that can change products appearance to be cleaned, such as white dyes.These whitening agent absorbing ultraviolet light also send visible light.Suitable fluorescent brightener levels comprises from about 0.01% weight, about 0.05% weight, about 0.1% weight, or even lower aq to 0.5% weight of about 0.2% weight, or even the high level of 0.75% weight.

Dispersion agent-composition of the present invention also can comprise dispersion agent.Suitable water soluble organic substance comprises homopolymerization or co-polymeric acids or its salt, and wherein polycarboxylic acid comprises at least two and is separated by and is no more than the carboxyl of two carbon atoms.

Enzyme-except lipase Variant of the present invention, detergent composition also can comprise one or more enzymes again so that clean-up performance and/or fabric care benefit effect to be provided, such as proteolytic enzyme, another kind of lipase, at, amylase, carbohydrase, cellulase, polygalacturonase, mannonase arabinase, Galactanase, zytase, oxydase, for example laccase and/or peroxidase.

The characteristic of the enzyme of usually selecting should be compatible with the washing composition of selecting, and (that is, best pH and other enzyme are compatible with non-enzyme component etc.), and enzyme should be significant quantity.

Suitable proteolytic enzyme comprises those animals, plant or microbe-derived proteolytic enzyme.The preferred microorganism source.Comprise mutant chemical modification or protein engineering.Proteolytic enzyme can be serine protease or metalloprotease, preferred microorganism Sumizyme MP or trypsin-like proteolytic enzyme.The example of Sumizyme MP is subtilisin, especially those are derived from the proteolytic enzyme of bacillus, for example subtilisin Novo, subtilysin, subtilisin 309, subtilisin 147 and subtilisin 168 (being described in WO 89/06279), sequence identification number 4 and sequence identification number 7 are described among the WO 05/103244.Other suitable serine protease comprises that those are obtained from some kind of micrococci suborder, especially is disclosed in serine protease and the variant thereof of Cellulonas kind, is disclosed among the WO2005052146.The example of trypsin-like proteolytic enzyme is for example trypsinase of pig or Niu Laiyuan of trypsin) and the reaping hook fungi protease, be described among WO 89/06270 and the WO 94/25583.

The example of available proteolytic enzyme is WO 92/19729, WO 98/20115, the variant of describing among WO 98/20116 and the WO 98/34946, especially the variant that has replacement in one or more following sites: 27,36,57,68,76,87,97,101,104,106,120,123,167,170,194,206,218,222,224,235,245,252 and 274, and other variant with following sudden change: (K27R, V104Y, N123S, T124A), (N76D, S103A, V104I), or (S101G, S103A, V104I, G159D, A232V, Q236H, Q245R, N248D, N252K).The example of other available proteolytic enzyme is the variant of describing among the WO 05/052146, especially especially has the variant of replacement in one or more following sites: 14,16,35,65,75,76,79,123,127,159 and 179

The proteolytic enzyme of preferred commercially available acquisition comprises Alcalase ^TM, Savinase ^TM, Primase ^TM, Duralase ^TM, Esperase ^TM, Coronase ^TM, Polarzyme ^TMAnd Kannase ^TM(NovozymesA/S), Maxatase ^TM, Maxacal ^TM, Maxapem ^TM, Properase ^TM, Purafect ^TM, PurafectPrime ^TM, Purafect OxP ^TM, FN2, FN3 and FN4 (Genencor International Inc.).

Lipase comprises the lipase of those bacteriums or originated from fungus.Comprise mutant chemical modification or protein engineering.The example of available lipase comprises and is obtained from humic mould (synonym: lipase thermophilic fungus), for example be obtained from high temperature chactomium globosum (synonym: lipase thermophilic fungus), be described among EP 258 068 and the EP 305 216, or be obtained from the lipase of Humicola insolens, be described among the WO 96/13580, Rhodopseudomonas lipase, for example be obtained from Pseudomonas alcaligenes or pseudomonas pseudoalcaligenes (EP 218 272), pseudomonas cepacia (EP 331376), (GB 1 for the Si Shi pseudomonas, 372,034), pseudomonas fluorescens, pseudomonas strain SD705 (WO 95/06720 and WO 96/27002), the lipase of prestige state fluted disc pseudomonas (WO 96/12012), bacillus lipase, such as being obtained from the bacterial strain subtilis (people such as Dartois, (1993), Biochemica et Biophysica Acta, 1131,253-360), the lipase of bacillus stearothermophilus (JP 64/744992) or bacillus pumilus (WO 91/16422).

Other example is the lipase Variant of describing in WO 92/05249, WO 94/01541, EP 407 225, EP 260 105, WO 95/35381, WO 96/00292, WO 95/30744, WO 94/25578, WO 95/14783, WO 95/22615, WO 97/04079 and WO 97/07202 such as those.

The lipase of other commercially available acquisition comprises Lipolase ^TM, Lipolase Ultra ^TMAnd Lipex ^TM(Novozymes A/S).

Suitable amylase (α and/or β) comprises the amylase of those bacteriums or originated from fungus.Comprise mutant chemical modification or protein engineering.Amylase for example comprises α-the derive from amylase of bacillus, and a kind of special lichem bacillus strain for example, this bacterial strain be at GB 1,296, more detailed description is arranged in 839.

The example of available starches enzyme is the variant of describing among WO 94/02597, WO 94/18314, WO 96/23873 and the WO 97/43424, especially has the variant of replacement in one or more following sites: 15,23,105,106,124,128,133,154,156,181,188,190,197,202,208,209,243,264,304,305,391,408 and 444.

The amylase of commercially available acquisition is Duramyl ^TM, Termamyl ^TM, Stainzyme ^TM, Stainzyme Ultra ^TM, Fungamyl ^TMAnd BAN ^TM(Novozymes A/S), Rapidase ^TMAnd Purastar ^TM(from Genencor International Inc.).

Suitable cellulase comprises the cellulase of those bacteriums or originated from fungus.Comprise mutant chemical modification or protein engineering.Suitable cellulase comprises the cellulase that is obtained from Bacillaceae, Rhodopseudomonas, the mould genus of humic, fusarium, careless Rhizopus, Acremonium, for example originate from Humicola insolens, thermophilic Huang and ruin the fungal cellulase of the mould and Fusarium oxysporum of silk, they are disclosed in US 4,435, and 307, US 5,648,263, US 5,691, and 178, US 5,776,757 and WO89/09259 in.

Especially suitable cellulase is alkalescence or the neutral cellulase with color care benefit effect.The example of these cellulases is cellulases of describing among EP 0 495 257, EP 0 531 372, WO96/11262, WO 96/29397, the WO 98/08940.Other example is the cellulase variants of describing 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 PCT/DK98/00299 such as those.

The cellulase of commercially available acquisition comprises Renozyme ^TM, Celluclean ^TM, Endolase, ^TMCelluzyme ^TM, and Carezyme ^TM(Novozymes A/S), Clazinase ^TM, and PuradaxHA ^TM(Genencor International Inc.) and KAC-500 (B) ^TM(Kao Corporation).

Peroxidase/oxydase:

Suitable peroxidase/oxydase comprises the enzyme of those plants, bacterium or originated from fungus.Comprise mutant chemical modification or protein engineering.Available peroxidase example comprises the peroxidase that is obtained from terrible agaric, for example is obtained from the peroxidase of Coprinus cinereus, and its variant is described among WO 93/24618, WO 95/10602 and the WO 98/15257.

The peroxidase of commercially available acquisition comprises Guardzyme ^TM(Novozymes A/S).

In the time of in being present in cleaning compositions, above-mentioned enzyme can be by weight of the composition about 0.00001% to about 2%, about 0.0001% to about 1%, perhaps in addition about 0.001% content to about 0.5% zymoprotein exist.

Enzyme stabilizers-can stablize the enzyme that is used for washing composition with multiple technologies.The enzyme that the present invention uses can be stablized by the calcium that exists in the final composition and/or magnesium ion water-soluble sources, and final product offers enzyme with this ion.Also can use other conventional stablizer, polyvalent alcohol for example, such as propylene glycol or glycerine, sugar or sugar alcohol, lactic acid, boric acid or boric acid derivatives aromatic borate for example, or the phenyl-boron dihydroxide derivative is such as 4-benzoyl boric acid, the method preparation that described composition can be described according to for example WO92/19709 and WO 92/19708.

Solvent-suitable solvent comprises water and other solvent such as lipophilic fluid.The example of suitable lipophilic fluid comprises siloxanes, other siloxanes, hydrocarbon, glycol ether, glycerol derivative such as glyceryl ether, perfluoroamine, perfluorination and hydrogen fluorine ether solvents, the floride-free organic solvent of low volatility, diol solvent, other environment amenable solvent and their mixture.

Washing methods

The present invention includes a kind of a certain position of cleaning and/or process, the method of surface or fabric particularly, aforesaid method may further comprise the steps: with applicant's cleaning compositions embodiment (with the pure substance form or be diluted in the washing liq) and at least part of surface or clothing in contact, and the then randomly above-mentioned surface of rinsing or fabric.Can before above-mentioned rinse step, carry out washing step to described surface or fabric.For the present invention, washing includes but not limited to clean and mechanical stirring.Understand as those skilled in the art, cleaning compositions of the present invention can be used for the laundry purposes ideally.Therefore, the present invention includes a kind of method for laundering of textile fabrics, said method comprising the steps of: fabric that will be to be washed contacts with described cleaning washing soln, and described washing soln comprises at least one embodiment of applicant's cleaning compositions, cleaning additive or their mixture.Fabric can comprise any most of any fabric that can wash under the normal consumer working conditions.Described solution preferably has about 8 to about 10 pH.The strength of solution that described composition adopts be about 100ppm, and preferably 500ppm is extremely about 15,000ppm.Water temperature is generally about 5 ℃ to about 90 ℃.The present invention in low water temperature as being especially useful below 30 ℃ or below 25 ℃ or 20 ℃.The ratio of water and fabric is generally about 1: 1 to about 30: 1.

The lipase Variant example

The commerical prod of SILVER REAGENT at least as the chemical substance of buffer reagent and substrate.

-substratum and solution: LAS (Surfac PS ^TM) and Wessalith CS (Wessalith P ^TM).Other used composition is standard laboratory reagent.

-material: EMPA221, from EMPA St.Gallen, Lerchfeldstrasse 5, CH-9014 St.Gallen, Switzerland

Example 1: enzyme production

Use the standard method of this area, make up the plasmid of a gene that comprises the lipase of encoding and its transfection is entered the appropriate host cell.

Use 34 ℃ of constant temperature, the substratum that initial volume is 1.2 liters carries out fed-batch fermentation.The initial pH of substratum is made as 6.5.In case pH rises to 7.0, keep this value constant by adding 10%H3PO4.Dissolved oxygen levels in the substratum is by changing stir speed (S.S.) and using the fixedly rate of venting of every liter of substratum per minute 1.0 litres of air to control.In the whole fed-batch fermentation stage, feeding rate maintains a constant level.The batch culture base comprises malt syrup as carbon source, and urea and yeast extract are as nitrogenous source, and the mixture of trace-metal and salt.The charging that adds continuously in the fed-batch fermentation stage comprises malt syrup as carbon source, yet adding urea and yeast extract are the nitrogenous source supplies in order to ensure abundance.

Use standard method known in the art to carry out the purifying of lipase, for example by filtering fermented supernatant fluid and follow-up hydrophobic chromatography and anionresin liquid, for example EP 0 851913 described methods in the example 3.

Example 2:AMSA-automation stress detection analytical method-for calculating relative performance (RP)

Use automation stress to detect the enzyme variants of analytical method (AMSA) test present patent application.By the AMSA test, can detect the scourability of a large amount of small volume enzyme-detergent solutions.The AMSA plate has a plurality of slits for test soln, and one firmly choke fabric sample, so that its capping that is washed at all slotted opening places.During washing, acutely shake described plate, test soln, fabric and capping so that the test soln contact fabric, and apply mechanical stress.More detailed description can be referring to WO 02/42740, especially the 23rd to 24 page paragraph " Special method embodiments ".The container that comprises the washing composition test soln is comprised of the cylindrical cavity in a metal sheet (diameter 6mm, dark 10mm).Dirty fabric (test substances) is positioned at the top of metal sheet, as capping and the sealing member on container.Another metal sheet is positioned at the top of dirty fabric to avoid overflowing any liquid from any one container.Two metal sheets together with dirty fabric with the frequency of 30Hz and the amplitude up-down vibration of 2mm.

Described detection analytical method is carried out under the test conditions of appointment hereinafter:

Test soln	0.5g/L LAS 0.52g/L Na 2CO3 1.07g/L Wessalith CS 0.52g/L trisodium citrate
		The test soln volume	160 microlitres
PH value	In statu quo (≈ 9.9.)
		Washing time	20 minutes
Temperature	30℃
		Water hardness
	15°dH Ca 2+/Mg 2+/NaHCO 3Ratio be 4: 1: 7.5
		Enzyme concn in the test soln	0.125,0.25,0.50,1.0mg zymoprotein/liter (mg ep/L)
Dry	Performance: pieces of fabric is in immediately rinsing in tap water after the washing, and at 85 ℃ of lower air-dry 5 minutes smells: pieces of fabric is in immediately rinsing in tap water after the washing, and in room temperature (20 ℃) lower dry 2 hours
		Test substances	White cream turmeric sample as described below (as the EMPA221 of cotton-spinning fabric)

Table 3

By under 50 ℃, mixing 5g (Santa Maria, Denmark) turmeric and 100g (38% fat, Arla, Denmark) white cream prepares white cream turmeric sample, and mixture kept under this temperature about 20 minutes and filtered (50 ℃) and removes any undissolved particle.Mixture is cooled to 20 ℃, and with the cotton spinning sample, EMPA221 immerses this white cream turmeric mixture, and then at room temperature dried overnight is also freezing until use.The preparation method of white cream turmeric sample is disclosed in the patent PA 200500775 that submitted on May 27th, 2005.

The performance of enzyme variants can be measured by the colour brightness with the fabric sample of concrete enzyme variants washing.Brightness also represents from the light intensity of fabric sample reflection during available white-light illuminating.When fabric was dirty, the intensity of reflected light of its catoptrical strength ratio clean textile was low.Therefore catoptrical intensity can be used for measuring the scourability of enzyme variants.

Use a kind of plane scanner (PFU DL2400pro) of specialty to carry out color measuring, this scanning device is used for obtaining the image of laundering of textile fabrics sample.Scanning resolution is 200dpi, 24 of output color depths.In order to obtain accurate result, with Kodak reflection IT8 colour atla frequent calibration scanner.

Use a specially designed application software (Novozymes Color Vector Analyzer) to come from scanning image, to extract light intensity value.Described program is retrieved 24 pixel values and is converted it into red, green and blue (RGB) value from image.By with the rgb value addition as carrier, then extract the length of gained carrier and come computed strength value (Int):

$\mathrm{Int}=\sqrt{{\mathrm{<figure-callout\; id="2"\; label="r"\; filenames="S2007800029559E00021.png,S2007800029559E00031.png"\; state="\{\{state\}\}">r</figure-callout>}}^2+{\mathrm{<figure-callout\; id="2"\; label="g"\; filenames="S2007800029559E00021.png,S2007800029559E00031.png"\; state="\{\{state\}\}">g</figure-callout>}}^2+{\mathrm{<figure-callout\; id="2"\; label="b"\; filenames="S2007800029559E00021.png,S2007800029559E00031.png"\; state="\{\{state\}\}">b</figure-callout>}}^2}$

The scourability of variant (P) is calculated according to following formula: P=Int (v)-Int (r) wherein

Int (v) is the light intensity value with the fabric face of tested enzyme washing, and Int (r) is the light intensity value without the fabric face of tested enzyme washing.

According to definition, regulation relative performance score is the result of AMSA washing: relative performance score (RP) is performance (P) sum and the ratio of reference enzyme performance: RP=P (the tested enzyme)/P (reference enzyme) of tested enzyme variant.

RPavg refers to the average relative performance that (0.125,0.25,0.5,1.0mg ep/l) compares with reference enzyme under all four kinds of enzyme concns

RPavg＝avg(RP(0.125)，RP(0.25)RP(0.5)，RP(1.0))

If the variant performance is better than reference enzyme, think that namely it shows the scourability of improvement.In situation of the present invention, reference enzyme is sequence identification number: 2 lipase, it has the replacement of T231R+N233R.

Example 3: the GC-gas-chromatography that is used for calculating the effect danger

By solid-phase microextraction vapor-phase chromatography (SPME-GC), the butyric acid that uses following methods to measure in the lipase washing sample discharges.To specify four pieces of fabric (diameter 5mm) of washing in the solution to be transferred to gas-chromatography (GC) bottle at the table 3 that comprises 1mg/L lipase.Analytic sample on a Varian3800 GC who is furnished with a Stabilwax-DA w/Integra-Guard post (30m, 0.32mm internal diameter and 0.25micro-m df) and a Carboxen PDMS SPME fiber (75micro-m).Each sample is 40 ℃ of precultures 10 minutes, then with sampling in the headspace of SPME fiber on pieces of fabric 20 minutes.Subsequently sample is injected chromatographic column (injector temperature=250 ℃).Post flow=2mL helium/minute.The column oven thermograde: 0 minute=40 ℃, 2 minutes=40 ℃, 22 minutes=240 ℃, 32 minutes=240 ℃.Detect butyric acid with fid detector, and calculate the amount of butyric acid based on the butyric acid typical curve.

The risk performance smell (R) of lipase Variant be the butyric acid that discharges from the lipase Variant washing sample with from sequence identification number: the ratio between the butyric acid amount of 2 lipase washing sample release, sequence identification number: 2 lipase has the replacement of T231R+N233R (reference enzyme), and two values all use the butyric acid amount that discharges from the washing sample of fat-free enzyme to proofread and correct.Calculate the risk (R) of variant according to following formula:

The microgram butyric acid that smell=the 1mg zymoprotein records/l blank correction

α _{Tested enzyme}=smell _{Tested enzyme}-blank

α _{Reference enzyme}=smell _{Reference enzyme}-blank

R=α _{Tested enzyme}/ α _{Reference enzyme}

If the R coefficient of a variant, thinks namely that it shows than the smell with reference to minimizing less than 1.

Example 4: with respect to the activity (LU) in the absorbancy of 280nm

Measure lipase with respect to the activity in the absorbancy of 280nm by following detection analytical method LU/A280:

The method that use is partly described at lipase activity is above measured lipase activity.Measure lipase in the absorbancy (A280) of 280nm and calculate the LU/A280 ratio.The LU/A280 of variant calculates relative LU/A280 divided by the LU/A280 of reference enzyme.In situation of the present invention, reference enzyme is sequence identification number: 2 lipase, it has the replacement of T231R+N233R.

Example 5:BR-is imitated the danger

With describe performance and smell reduce risks than effect danger coefficient therefore be defined as: BR=RP _Avg/ R

If the BR coefficient of a variant is higher than 1, think that namely it shows the scourability of improvement and the smell of minimizing.

The result who uses above method acquisition is as follows:

Variant	In sequence identification number: 2	Average RP (RP avg)	BR	LU/A280
					1	I202G+T231R+N233R	0.84	1.41	Undetermined
2	I86V+L227G+T231R+N233R+P256K	1.08	1.52	1700
					3	Q4V+S58N+V60S+T231R+N233R	0.87	1.73	1950
4	S58N+V60S+I90R+T231R+N233R	1.06	1.27	2250
					5	I255Y+T231R+N233R	1.19	1.17	3600
6	I90A+T231R+N233R+I255V	1.13	1.14	2700
					Reference	T231R+N233R	1.00	1.00	3650
7	G91A+E99K+T231R+N233R+Q249R +270H+271T+272P+273S+274S+ 275G+276R+277G+278G+279H+ 280R	0.43	Undetermined	850
					8	G91A+E99K+T231R，N233R+Q249R +270H+271T+272P+273S+274S+ 275G+276R+277G+278G	0.13	Undetermined	500

Table 4

WO 2000/060063 has described reference lipase and variant 7 and 8 in the table 4.

The washing composition example

The affirmation of abbreviation component that is used for example is as follows:

The LAS straight chain C _11-13Sodium alkyl benzene sulfonate.

CxyAS C _1x-C _1ySodium alkyl sulfate

The C of CxyEzS and average z moles of ethylene oxide condensation _1x-C _1ySodium alkyl sulfate

CxyEy C _1x-C _1yAlcohol has average degree of ethoxylation z

QAS R ₂.N+ (CH ₃) ₂(C ₂H ₄OH), R wherein ₂=C ₁₀-C ₁₂

Silicate amorphous sodium silicate (SiO ₂: Na ₂The ratio of O is 1.6: 1 to 3.2: 1).

The Wessalith CS molecular formula is Na ₁₂(AlO ₂SiO ₂) ₁₂.27H ₂The hydrated sodium aluminosilicate of O has the primary particle sizes (weight is based on anhydrous statement) in 0.1 to 10 micrometer range.

(Na-) SKS-6 crystalline layered silicate, molecular formula are δ-Na ₂Si ₂O ₅.

Citrate trianion two hydration trisodium citrates

The anhydrous citric acid citric acid.

The carbonate anhydrous sodium carbonate.

The vitriol anhydrous sodium sulphate.

The random copolymers of 4: 1 acrylate/maleic acid esters of MA/AA, molecular-weight average are about 70,000 to 80,000.

AA polymer poly sodium acrylate polymkeric substance, molecular-weight average is about 4,500.

PB1/PB4 anhydrous sodium perborate monohydrate/tetrahydrate.

Anhydrous SPC-D [the 2.74Na of PC3 ₂CO ₃.3H ₂O ₂]

The TAED tetra acetyl ethylene diamine.

NOBS nonanoyl Sodium dobesilate.

DTPA ethylidene triamine five acetic acid.

HEDP hydroxyl ethane diphosphonates

EDDS 1，2-diaminoethane-N, the sodium salt of N '-disuccinic acid, (S, S) isomer

The STPP tripoly phosphate sodium STPP

The protease protein lytic enzyme is with trade(brand)name

Be sold by Novozymes A/S, with trade(brand)name

Purafect

And Purafect

Be sold by Genencor, and proteolytic enzyme such as FNA, the FN3 and/or the FN4 that in patent WO 91/06637 and/or WO 95/10591 and/or EP 0 251 446, describe.

Amylase is with trade(brand)name

Purafect

Be sold by Genencor;

With

Be sold by the amylolytic enzyme of Novozymes A/S.

Lipase has been described any in the lipase Variant 1 to 5 and their combination in the example 5 of table 4.

Mannase is sold by Novozymes's

CMC or carboxymethyl or hydroxyethyl or ester modified Mierocrystalline cellulose

HEC or

EMC

SS agglomerate suds suppressor agglomerate: 12% siloxanes/silicon-dioxide, 18% stearyl alcohol, 70% starch is particle form.

TEPAE tetren ethoxylate

In 1% distilled water solution, measure during 20 ℃ of pH.

Example A

1. the bleach detergent compositions that has the granular laundry detergent form by following formula examples explanation.

	A	B	C	D	E	F
							LAS	20	22	20	15	20	20
QAS	0.7	1	1	0.6	0.0	0.7
							C25E3S	0.9	0.0	0.9	0.0	0.0	0.9
C25E7	0.0	0.5	0.0	1	3	1
							STPP	23	30	23	17	12	23
Wessalith CS	0.0	0.0	0.0	0.0	10	0.0
							Silicate	7	7	7	7	7	7
Carbonate	15	14	15	18	15	15
							The AA polymkeric substance	1	0.0	1	1	1.5	1
CMC	1	1	1	1	1	1
							Protease 3 2.89mg/g	0.1	0.07	0.1	0.1	0.1	0.1
Amylase 8 .65mg/g	0.1	0.1	0.1	0.0	0.1	0.1
							Lipase 18mg/g	0.03	0.07	0.3	0.1	0.07	0.1
Whitening agent-Tinopal AMS (Ciba)	0.06	0.0	0.06	0.18	0.06	0.06
							Whitening agent-Tinopal CBS-X (Ciba)	0.1	0.06	0.1	0.0	0.1	0.1
DTPA	0.6	0.3	0.6	0.25	0.6	0.6
							MgSO 4	1	1	1	0.5	1	1
PC3	0.0	5.2	0.1	0.0	0.0	0.0

PB1	4.4	0.0	3.85	2.09	0.78	3.63
							NOBS	1.9	0.0	1.66	1.77	0.33	0.75
TAED	0.58	1.2	0.51	0.0	0.015	0.28
							Vitriol/moisture	Surplus to 100%	Surplus to 100%	Surplus to 100%	Surplus to 100%	Surplus to 100%	Surplus to 100%

Under 25 ℃, any composition that is used for laundering of textile fabrics among the example A is 600ppm to 10000ppm in the concentration of water, and typical intermediate conditions is 2500ppm, and the ratio of water and fabric is 25: 1.Typical pH is about 10, but can regulate pH with the ratio of the sodium-salt form of alkyl benzene sulphonate (ABS) by changing acid.

Example B

The bleach detergent compositions that has the granular laundry detergent form by the explanation of following formula examples.

	A	B	C	D
					LAS
	8	7.1	7	6.5
					C25E3S	0	4.8	0	5.2
C68S	1	0	1	0
					C25E7	2.2	0	3.2	0
QAS	0.75	0.94	0.98	0.98
					(Na-)SKS-6	4.1	0	4.8	0
Wessalith CS	20	0	17	0
					Citric acid	3	5	3	4
Carbonate	15	20	14	20
					Silicate	0.08	0	0.11	0
Stain remover	0.75	0.72	0.71	0.72
					MA/AA	1.1	3.7	1.0	3.7
CMC	0.15	1.4	0.2	1.4
					Proteolytic enzyme (56.00mg active substance/gram)	0.37	0.4	0.4	0.4
Termamyl (the 21.55mg active substance/g)	0.3	0.3	0.3	0.3
					Lipase (18.00mg active substance/gram)	0.05	0.15	0.1	0.5

Amylase (8.65mg active substance/gram)	0.1	0.14	0.14	0.3
					TAED	3.6	4.0	3.6	4.0
PC3	13	13.2	13	13.2
					EDDS	0.2	0.2	0.2	0.2
HEDP	0.2	0.2	0.2	0.2
					MgSO 4	0.42	0.42	0.42	0.42
Spices	0.5	0.6	0.5	0.6
					The SS agglomerate	0.05	0.1	0.05	0.1
Soap	0.45	0.45	0.45	0.45
					Vitriol	22	33	24	30
Water and other composition	Surplus to 100%	Surplus to 100%	Surplus to 100%	Surplus to 100%

20 ℃ under 90 ℃, any above-mentioned composition that is used for laundering of textile fabrics among the example B is 10,000ppm in the concentration of water, and the ratio of water and fabric is 5: 1.Typical pH is about 10, but can regulate pH with the ratio of the sodium-salt form of alkyl benzene sulphonate (ABS) by changing acid.

Example C

	A (% by weight)	B (% by weight)	C (% by weight)	D (% by weight)	E (% by weight)	F (% by weight)
							C25E1.8S	11	10	4	6.32	6.0	8.2
LAS	4	5.1	8	3.3	4.0	3.0
							Sodium formiate	1.6	0.09	1.2	0.04	1.6	1.2
Sodium hydroxide	2.3	3.8	1.7	1.9	2.3	1.7
							Monoethanolamine	1.4	1.490	1.0	0.7	1.35	1.0
Glycol ether	5.5	0.0	4.1	0.0	5.500	4.1
							C23E9	0.4	0.6	0.3	0.3	2	0.3
DTPA	0.15	0.15	0.11	0.07	0.15	0.11
							Citric acid	2.5	3.96	1.88	1.98	2.5	1.88
C 12-14Dimethyl oxidation amine	0.3	0.73	0.23	0.37	0.3	0.225
							C 12-18Lipid acid	0.8	1.9	0.6	0.99	0.8	0.6

Borax	1.43	1.5	1.1	0.75	1.43	1.07
							Ethanol	1.54	1.77	1.15	0.89	1.54	1.15
TEPAE 1	0.3	0.33	0.23	0.17	0.0	0.0
							The hexamethylene-diamine of ethoxylation 2	0.8	0.81	0.6	0.4	0.0	0.0
1,2-PD	0.0	6.6	0.0	3.3	0.0	0.0
							Proteolytic enzyme *	36.4	36.4	27.3	18.2	36.4	27.3
Mannase *	1.1	1.1	0.8	0.6	1.1	0.8
							Amylase *	7.3	7.3	5.5	3.7	7.3	5.5
Lipase *	10	3.2	0.5	3.2	2.4	3.2
							Water, spices, dyestuff and other component	Surplus	Surplus	Surplus	Surplus	Surplus	Surplus

^*Numerical value in mg enzyme/100g

¹Such as US 4,597, described in 898.

²With trade(brand)name

Derive from BASF, and described in WO 01/05874 those

The All Files of quoting in detailed Description Of The Invention is all introduced for your guidance in relevant portion; Should not be interpreted as admitting that for quoting of any file it is relevant prior art of the present invention.When any implication of term in any implication of term among the present invention or definition and the file of introducing for your guidance or when defining contradiction, should obey implication or the definition of giving in the present invention this term.

Although illustrated and described specific embodiments of the present invention, it will be apparent to one skilled in the art that and in the situation that does not deviate from essence of the present invention and scope, can make a plurality of other changes and modification.Therefore, claims all such changes and modification of being intended to be included in the scope of the present invention.

Sequence table

<110>The Procter & Gamble Company

＜120〉detergent composition

<130>CM3051ML

<160>16

<170>PatentIn version 3.3

<210>1

<211>807

<212>DNA

＜213〉thermophilic fungus (Thermomyces lanuginosus)

<220>

<221>CDS

<222>(1)..(807)

<220>

<221>mat_peptide

<222>(1)..()

<400>1

gag gtc tcg cag gat ctg ttt aac cag ttc aat ctc ttt gca cag tat 48

Glu Val Ser Gln Asp Leu Phe Asn Gln Phe Asn Leu Phe Ala Gln Tyr

1 5 10 15

tct gca gcc gca tac tgc gga aaa aac aat gat gcc cca gct ggt aca 96

Ser Ala Ala Ala Tyr Cys Gly Lys Asn Asn Asp Ala Pro Ala Gly Thr

20 25 30

aac att acg tgc acg gga aat gcc tgc ccc gag gta gag aag gcg gat 144

Asn Ile Thr Cys Thr Gly Asn Ala Cys Pro Glu Val Glu Lys Ala Asp

35 40 45

gca acg ttt ctc tac tcg ttt gaa gac tct gga gtg ggc gat gtc acc 192

Ala Thr Phe Leu Tyr Ser Phe Glu Asp Ser Gly Val Gly Asp Val Thr

50 55 60

ggc ttc ctt gct ctc gac aac acg aac aaa ttg atc gtc ctc tct ttc 240

Gly Phe Leu Ala Leu Asp Asn Thr Asn Lys Leu Ile Val Leu Ser Phe

65 70 75 80

cgt ggc tct cgt tcc ata gag aac tgg atc ggg aat ctt aac ttc gac 288

Arg Gly Ser Arg Ser Ile Glu Asn Trp Ile Gly Asn Leu Asn Phe Asp

85 90 95

ttg aaa gaa ata aat gac att tgc tcc ggc tgc agg gga cat gac ggc 336

Leu Lys Glu Ile Asn Asp Ile Cys Ser Gly Cys Arg Gly His Asp Gly

100 105 110

ttc act tcg tcc tgg agg tct gta gcc gat acg tta agg cag aag gtg 384

Phe Thr Ser Ser Trp Arg Ser Val Ala Asp Thr Leu Arg Gln Lys Val

115 120 125

gag gat gct gtg agg gag cat ccc gac tat cgc gtg gtg ttt acc gga 432

Glu Asp Ala Val Arg Glu His Pro Asp Tyr Arg Val Val Phe Thr Gly

130 135 140

cat agc ttg ggt ggt gca ttg gca act gtt gcc gga gca gac ctg cgt 480

His Ser Leu Gly Gly Ala Leu Ala Thr Val Ala Gly Ala Asp Leu Arg

145 150 155 160

gga aat ggg tat gat atc gac gtg ttt tca tat ggc gcc ccc cga gtc 528

Gly Asn Gly Tyr Asp Ile Asp Val Phe Ser Tyr Gly Ala Pro Arg Val

165 170 175

gga aac agg gct ttt gca gaa ttc ctg acc gta cag acc ggc gga aca 576

Gly Asn Arg Ala Phe Ala Glu Phe Leu Thr Val Gln Thr Gly Gly Thr

180 185 190

ctc tac cgc att acc cac acc aat gat att gtc cct aga ctc ccg ccg 624

Leu Tyr Arg Ile Thr His Thr Asn Asp Ile Val Pro Arg Leu Pro Pro

195 200 205

cgc gaa ttc ggt tac agc cat tct agc cca gag tac tgg atc aaa tct 672

Arg Glu Phe Gly Tyr Ser His Ser Ser Pro Glu Tyr Trp Ile Lys Ser

210 215 220

gga acc ctt gtc ccc gtc acc cga aac gat atc gtg aag ata gaa ggc 720

Gly Thr Leu Val Pro Val Thr Arg Asn Asp Ile Val Lys Ile Glu Gly

225 230 235 240

atc gat gcc acc ggc ggc aat aac cag cct aac att ccg gat atc cct 768

Ile Asp Ala Thr Gly Gly Asn Asn Gln Pro Asn Ile Pro Asp Ile Pro

245 250 255

gcg cac cta tgg tac ttc ggg tta att ggg aca tgt ctt 807

Ala His Leu Trp Tyr Phe Gly Leu Ile Gly Thr Cys Leu

260 265

<210>2

<211>269

<212>PRT

＜213〉thermophilic fungus (Thermomyces lanuginosus)

<400>2

Glu Val Ser Gln Asp Leu Phe Asn Gln Phe Asn Leu Phe Ala Gln Tyr

1 5 10 15

Ser Ala Ala Ala Tyr Cys Gly Lys Asn Asn Asp Ala Pro Ala Gly Thr

20 25 30

Asn Ile Thr Cys Thr Gly Asn Ala Cys Pro Glu Val Glu Lys Ala Asp

35 40 45

Ala Thr Phe Leu Tyr Ser Phe Glu Asp Ser Gly Val Gly Asp Val Thr

50 55 60

Gly Phe Leu Ala Leu Asp Asn Thr Asn Lys Leu Ile Val Leu Ser Phe

65 70 75 80

Arg Gly Ser Arg Ser Ile Glu Asn Trp Ile Gly Asn Leu Asn Phe Asp

85 90 95

Leu Lys Glu Ile Asn Asp Ile Cys Ser Gly Cys Arg Gly His Asp Gly

100 105 110

Phe Thr Ser Ser Trp Arg Ser Val Ala Asp Thr Leu Arg Gln Lys Val

115 120 125

Glu Asp Ala Val Arg Glu His Pro Asp Tyr Arg Val Val Phe Thr Gly

130 135 140

His Ser Leu Gly Gly Ala Leu Ala Thr Val Ala Gly Ala Asp Leu Arg

145 150 155 160

Gly Asn Gly Tyr Asp Ile Asp Val Phe Ser Tyr Gly Ala Pro Arg Val

165 170 175

Gly Asn Arg Ala Phe Ala Glu Phe Leu Thr Val Gln Thr Gly Gly Thr

180 185 190

Leu Tyr Arg Ile Thr His Thr Asn Asp Ile Val Pro Arg Leu Pro Pro

195 200 205

Arg Glu Phe Gly Tyr Ser His Ser Ser Pro Glu Tyr Trp Ile Lys Ser

210 215 220

Gly Thr Leu Val Pro Val Thr Arg Asn Asp Ile Val Lys Ile Glu Gly

225 230 235 240

Ile Asp Ala Thr Gly Gly Asn Asn Gln Pro Asn Ile Pro Asp Ile Pro

245 250 255

Ala His Leu Trp Tyr Phe Gly Leu Ile Gly Thr Cys Leu

260 265

<210>3

<211>265

<212>PRT

＜213〉colter mould (Absidia reflexa)

<400>3

Ser Ser Ser Ser Thr Gln Asp Tyr Arg Ile Ala Ser Glu Ala Glu Ile

1 5 10 15

Lys Ala His Thr Phe Tyr Thr Ala Leu Ser Ala Asn Ala Tyr Cys Arg

20 25 30

Thr Val Ile Pro Gly Gly Arg Trp Ser Cys Pro His Cys Gly Val Ala

35 40 45

Ser Asn Leu Gln Ile Thr Lys Thr Phe Ser Thr Leu Ile Thr Asp Thr

50 55 60

Asn Val Leu Val Ala Val Gly Glu Lys Glu Lys Thr Ile Tyr Val Val

65 70 75 80

Phe Arg Gly Thr Ser Ser Ile Arg Asn Ala Ile Ala Asp Ile Val Phe

85 90 95

Val Pro Val Asn Tyr Pro Pro Val Asn Gly Ala Lys Val His Lys Gly

100 105 110

Phe Leu Asp Ser Tyr Asn Glu Val Gln Asp Lys Leu Val Ala Glu Val

115 120 125

Lys Ala Gln Leu Asp Arg His Pro Gly Tyr Lys Ile Val Val Thr Gly

130 135 140

His Ser Leu Gly Gly Ala Thr Ala Val Leu Ser Ala Leu Asp Leu Tyr

145 150 155 160

His His Gly His Ala Asn Ile Glu Ile Tyr Thr Gln Gly Gln Pro Arg

165 170 175

Ile Gly Thr Pro Ala Phe Ala Asn Tyr Val Ile Gly Thr Lys Ile Pro

180 185 190

Tyr Gln Arg Leu Val His Glu Arg Asp Ile Val Pro His Leu Pro Pro

195 200 205

Gly Ala Phe Gly Phe Leu His Ala Gly Glu Glu Phe Trp Ile Met Lys

210 215 220

Asp Ser Ser Leu Arg Val Cys Pro Asn Gly Ile Glu Thr Asp Asn Cys

225 230 235 240

Ser Asn Ser Ile Val Pro Phe Thr Ser Val Ile Asp His Leu Ser Tyr

245 250 255

Leu Asp Met Asn Thr Gly Leu Cys Leu

260 265

<210>4

<211>264

<212>PRT

＜213〉absidia corymbifera (Absidia corymbifera)

<400>4

Ser Ser Ser Thr Gln Asp Tyr Arg Ile Ala Ser Glu Ala Glu Ile Lys

1 5 10 15

Ala His Thr Phe Tyr Thr Ala Leu Ser Ala Asn Ala Tyr Cys Arg Thr

20 25 30

Val Ile Pro Gly Gly Gln Trp Ser Cys Pro His Cys Asp Val Ala Pro

35 40 45

Asn Leu Asn Ile Thr Lys Thr Phe Thr Thr Leu Ile Thr Asp Thr Asn

50 55 60

Val Leu Val AlaVal Gly Glu Asn Glu Lys Thr Ile Tyr Val Val Phe

65 70 75 80

Arg Gly Thr Ser Ser Ile Arg Asn Ala Ile Ala Asp Ile Val Phe Val

85 90 95

Pro Val Asn Tyr Pro Pro Val Asn Gly Ala Lys Val His Lys Gly Phe

100 105 110

Leu Asp Ser Tyr Asn Glu Val Gln Asp Lys Leu Val Ala Glu Val Lys

115 120 125

Ala Gln Leu Asp Arg His Pro Gly Tyr Lys Ile Val Val Thr Gly His

130 135 140

Ser Leu Gly Gly Ala Thr Ala Val Leu Ser Ala Leu Asp Leu Tyr His

145 150 155 160

His Gly His Asp Asn Ile Glu Ile Tyr Thr Gln Gly Gln Pro Arg Ile

165 170 175

Gly Thr Pro Glu Phe Ala Asn Tyr Val Ile Gly Thr Lys Ile Pro Tyr

180 185 190

Gln Arg Leu Val Asn Glu Arg Asp Ile Val Pro His Leu Pro Pro Gly

195 200 205

Ala Phe Gly Phe Leu His Ala Gly Glu Glu Phe Trp Ile Met Lys Asp

210 215 220

Ser Ser Leu Arg Val Cys Pro Asn Gly Ile Glu Thr Asp Asn Cys Ser

225 230 235 240

Asn Ser Ile Val Pro Phe Thr Ser Val Ile Asp His Leu Ser Tyr Leu

245 250 255

Asp Met Asn Thr Gly Leu Cys Leu

260

<210>5

<211>269

<212>PRT

＜213〉rhizomucor miehei (Rhizomucor miehei)

<400>5

Ser Ile Asp Gly Gly Ile Arg Ala Ala Thr Ser Gln Glu Ile Asn Glu

1 5 10 15

Leu Thr Tyr Tyr Thr Thr Leu Ser Ala Asn Ser Tyr Cys Arg Thr Val

20 25 30

Ile Pro Gly Ala Thr Trp Asp Cys Ile His Cys Asp Ala Thr Glu Asp

35 40 45

Leu Lys Ile Ile Lys Thr Trp Ser Thr Leu Ile Tyr Asp Thr Asn Ala

50 55 60

Met Val Ala Arg Gly Asp Ser Glu Lys Thr Ile Tyr Ile Val Phe Arg

65 70 75 80

Gly Ser Ser Ser Ile Arg Asn Trp Ile Ala Asp Leu Thr Phe Val Pro

85 90 95

Val Ser Tyr Pro Pro Val Ser Gly Thr Lys Val His Lys Gly Phe Leu

100 105 110

Asp Ser Tyr Gly Glu Val Gln Asn Glu Leu Val Ala Thr Val Leu Asp

115 120 125

Gln Phe Lys Gln Tyr Pro Ser Tyr Lys Val Ala Val Thr Gly His Ser

130 135 140

Leu Gly Gly Ala Thr Ala Leu Leu Cys Ala Leu Asp Leu Tyr Gln Arg

145 150 155 160

Glu Glu Gly Leu Ser Ser Ser Asn Leu Phe Leu Tyr Thr Gln Gly Gln

165 170 175

Pro Arg Val Gly Asp Pro Ala Phe Ala Asn Tyr Val Val Ser Thr Gly

180 185 190

Ile Pro Tyr Arg Arg Thr Val Asn Glu Arg Asp Ile Val Pro His Leu

195 200 205

Pro Pro Ala Ala Phe Gly Phe Leu His Ala Gly Glu Glu Tyr Trp Ile

210 215 220

Thr Asp Asn Ser Pro Glu Thr Val Gln Val Cys Thr Ser Asp Leu Glu

225 230 235 240

Thr Ser Asp Cys Ser Asn Ser Ile Val Pro Phe Thr Ser Val Leu Asp

245 250 255

His Leu Ser Tyr Phe Gly Ile Asn Thr Gly Leu Cys Thr

260 265

<210>6

<211>271

<212>PRT

＜213〉Rhizopus oryzae (Rhizopus oryzae)

<400>6

Ser Ala Ser Asp Gly Gly Lys Val Val Ala Ala Thr Thr Ala Gln Ile

1 5 10 15

Gln Glu Phe Thr Lys Tyr Ala Gly Ile Ala Ala Thr Ala Tyr Cys Arg

20 25 30

Ser Val Val Pro Gly Asn Lys Trp Asp Cys Val Gln Cys Gln Lys Trp

35 40 45

Val Pro Asp Gly Lys Ile Ile Thr Thr Phe Thr Ser Leu Leu Ser Asp

50 55 60

Thr Asn Gly Tyr Val Leu Arg Ser Asp Lys Gln Lys Thr Ile Tyr Leu

65 70 75 80

Val Phe Arg Gly Thr Asn Ser Phe Arg Ser Ala Ile Thr Asp Ile Val

85 90 95

Phe Asn Phe Ser Asp Tyr Lys Pro Val Lys Gly Ala Lys Val His Ala

100 105 110

Gly Phe Leu Ser Ser Tyr Glu Gln Val Val Asn Asp Tyr Phe Pro Val

115 120 125

Val Gln Glu Gln Leu Thr Ala His Pro Thr Tyr Lys Val Ile Val Thr

130 135 140

Gly His Ser Leu Gly Gly Ala Gln Ala Leu Leu Ala Gly Met Asp Leu

145 150 155 160

Tyr Gln Arg Glu Pro Arg Leu Ser Pro Lys Asn Leu Ser Ile Phe Thr

165 170 175

Val Gly Gly Pro Arg Val Gly Asn Pro Thr Phe Ala Tyr Tyr Val Glu

180 185 190

Ser Thr Gly Ile Pro Phe Gln Arg Thr Val His Lys Arg Asp Ile Val

195 200 205

Pro His Val Pro Pro Gln Ser Phe Gly Phe Leu His Pro Gly Val Glu

210 215 220

Ser Trp Ile Lys Ser Gly Thr Ser Asn Val Gln Ile Cys Thr Ser Glu

225 230 235 240

Ile Glu Thr Lys Asp Cys Ser Asn Ser Ile Val Pro Phe Thr Ser Ile

245 250 255

Leu Asp His Leu Ser Tyr Phe Asp Ile Asn Glu Gly Ser Cys Leu

260 265 270

<210>7

<211>267

<212>PRT

＜213〉aspergillus niger (Aspergillus niger)

<400>7

Thr Ala Gly His Ala Leu Ala Ala Ser Thr Gln Gly Ile Ser Glu Asp

1 5 10 15

Leu Tyr Ser Arg Leu Val Glu Met Ala Thr Ile Ser Gln Ala Ala Tyr

20 25 30

Ala Asp Leu Cys Asn Ile Pro Ser Thr Ile Ile Lys Gly Glu Lys Ile

35 40 45

Tyr Asn Ser Gln Thr Asp Ile Asn Gly Trp Ile Leu Arg Asp Asp Ser

50 55 60

Ser Lys Glu Ile Ile Thr Val Phe Arg Gly Thr Gly Ser Asp Thr Asn

65 70 75 80

Leu Gln Leu Asp Thr Asn Tyr Thr Leu Thr Pro Phe Asp Thr Leu Pro

85 90 95

Gln Cys Asn Gly Cys Glu Val His Gly Gly Tyr Tyr Ile Gly Trp Val

100 105 110

Ser Val Gln Asp Gln Val Glu Ser Leu Val Lys Gln Gln Val Ser Gln

115 120 125

Tyr Pro Asp Tyr Ala Leu Thr Val Thr Gly His Ser Leu Gly Ala Ser

130 135 140

Leu Ala Ala Leu Thr Ala Ala Gln Leu Ser Ala Thr Tyr Asp Asn Ile

145 150 155 160

Arg Leu Tyr Thr Phe Gly Glu Pro Arg Ser Gly Asn Gln Ala Phe Ala

165 170 175

Ser Tyr Met Asn Asp Ala Phe Gln Ala Ser Ser Pro Asp Thr Thr Gln

180 185 190

Tyr Phe Arg Val Thr His Ala Asn Asp Gly Ile Pro Asn Leu Pro Pro

195 200 205

Val Glu Gln Gly Tyr Ala His Gly Gly Val Glu Tyr Trp Ser Val Asp

210 215 220

Pro Tyr Ser Ala Gln Asn Thr Phe Val Cys Thr Gly Asp Glu Val Gln

225 230 235 240

Cys Cys Glu Ala Gln Gly Gly Gln Gly Val Asn Asn Ala His Thr Thr

245 250 255

Tyr Phe Gly Met Thr Ser Gly Ala Cys Thr Trp

260 265

<210>8

<211>266

<212>PRT

＜213〉Tabin aspergillus (Aspergillus tubingensis)

<400>8

Thr Ala Gly His Ala Leu Ala Ala Ser Thr Gln Gly Ile Ser Glu Asp

1 5 10 15

Leu Tyr Ser Arg Leu Val Glu Met Ala Thr Ile Ser Gln Ala Ala Tyr

20 25 30

Ala Asp Leu Cys Asn Ile Pro Ser Thr Ile Ile Lys Gly Glu Lys Ile

35 40 45

Tyr Asn Ser Gln Thr Asp Ile Asn Gly Trp Ile Leu Arg Asp Asp Ser

50 55 60

Ser Lys Glu Ile Ile Thr Val Phe Arg Gly Thr Gly Ser Asp Thr Asn

65 70 75 80

Leu Gln Leu Asp Thr Asn Tyr Thr Leu Thr Pro Phe Asp Thr Leu Pro

85 90 95

Gln Cys Asn Ser Cys Glu Val His Gly Gly Tyr Tyr Ile Gly Trp Ile

100 105 110

Ser Val Gln Asp Gln Val Glu Ser Leu Val Gln Gln Gln Val Ser Gln

115 120 125

Phe Pro Asp Tyr Ala Leu Thr Val Thr Gly His Ser Leu Gly Ala Ser

130 135 140

Leu Ala Ala Leu Thr Ala Ala Gln Leu Ser Ala Thr Tyr Asp Asn Ile

145 150 155 160

Arg Leu Tyr Thr Phe Gly Glu Pro Arg Ser Asn Gln Ala Phe Ala Ser

165 170 175

Tyr Met Asn Asp Ala Phe Gln Ala Ser Ser Pro Asp Thr Thr Gln Tyr

180 185 190

Phe Arg Val Thr His Ala Asn Asp Gly Ile Pro Asn Leu Pro Pro Ala

195 200 205

Asp Glu Gly Tyr Ala His Gly Val Val Glu Tyr Trp Ser Val Asp Pro

210 215 220

Tyr Ser Ala Gln Asn Thr Phe Val Cys Thr Gly Asp Glu Val Gln Cys

225 230 235 240

Cys Glu Ala Gln Gly Gly Gln Gly Val Asn Asn Ala His Thr Thr Tyr

245 250 255

Phe Gly Met Thr Ser Gly His Cys Thr Trp

260 265

<210>9

<211>276

<212>PRT

＜213〉Fusarium oxysporum (Fusarium oxysporum)

<400>9

Ala Val Gly Val Thr Thr Thr Asp Phe Ser Asn Phe Lys Phe Tyr Ile

1 5 10 15

Gln His Gly Ala Ala Ala Tyr Cys Asn Ser Glu Ala Ala Ala Gly Ser

20 25 30

Lys Ile Thr Cys Ser Asn Asn Gly Cys Pro Thr Val Gln Gly Asn Gly

35 40 45

Ala Thr Ile Val Thr Ser Phe Val Gly Ser Lys Thr Gly Ile Gly Gly

50 55 60

Tyr Val Ala Thr Asp Ser Ala Arg Lys Glu Ile Val Val Ser Phe Arg

65 70 75 80

Gly Ser Ile Asn Ile Arg Asn Trp Leu Thr Asn Leu Asp Phe Gly Gln

85 90 95

Glu Asp Cys Ser Leu Val Ser Gly Cys Gly Val His Ser Gly Phe Gln

100 105 110

Arg Ala Trp Asn Glu Ile Ser Ser Gln Ala Thr Ala Ala Val Ala Ser

115 120 125

Ala Arg Lys Ala Asn Pro Ser Phe Asn Val Ile Ser Thr Gly His Ser

130 135 140

Leu Gly Gly Ala Val Ala Val Leu Ala Ala Ala Asn Leu Arg Val Gly

145 150 155 160

Gly Thr Pro Val Asp Ile Tyr Thr Tyr Gly Ser Pro Arg Val Gly Asn

165 170 175

Ala Gln Leu Ser Ala Phe Val Ser Asn Gln Ala Gly Gly Glu Tyr Arg

180 185 190

Val Thr His Ala Asp Asp Pro Val Pro Arg Leu Pro Pro Leu Ile Phe

195 200 205

Gly Tyr Arg His Thr Thr Pro Glu Phe Trp Leu Ser Gly Gly Gly Gly

210 215 220

Asp Lys Val Asp Tyr Thr Ile Ser Asp Val Lys Val Cys Glu Gly Ala

225 230 235 240

Ala Asn Leu Gly Cys Asn Gly Gly Thr Leu Gly Leu Asp Ile Ala Ala

245 250 255

His Leu His Tyr Phe Gln Ala Thr Asp Ala Cys Asn Ala Gly Gly Phe

260 265 270

Ser Trp Arg Arg

275

<210>10

<211>273

<212>PRT

＜213〉fusarium heterosporium (Fusarium heterosporum)

<400>10

Thr Val Thr Thr Gln Asp Leu Ser Asn Phe Arg Phe Tyr Leu Gln His

1 5 10 15

Ala Asp Ala Ala Tyr Cys Asn Phe Asn Thr Ala Val Gly Lys Pro Val

20 25 30

His Cys Ser Ala Gly Asn Cys Pro Asp Ile Glu Lys Asp Ala Ala Ile

35 40 45

Val Val Gly Ser Val Val Gly Thr Lys Thr Gly Ile Gly Ala Tyr Val

50 55 60

Ala Thr Asp Asn Ala Arg Lys Glu Ile Val Val Ser Val Arg Gly Ser

65 70 75 80

Ile Asn Val Arg Asn Trp Ile Thr Asn Phe Asn Phe Gly Gln Lys Thr

85 90 95

Cys Asp Leu Val Ala Gly Cys Gly Val His Thr Gly Phe Leu Asp Ala

100 105 110

Trp Glu Glu Val Ala Ala Asn Val Lys Ala Ala Val Ser Ala Ala Lys

115 120 125

Thr Ala Asn Pro Thr Phe Lys Phe Val Val Thr Gly His Ser Leu Gly

130 135 140

Gly Ala Val Ala Thr Ile Ala Ala Ala Tyr Leu Arg Lys Asp Gly Phe

145 150 155 160

Pro Phe Asp Leu Tyr Thr Tyr Gly Ser Pro Arg Val Gly Asn Asp Phe

165 170 175

Phe Ala Asn Phe Val Thr Gln Gln Thr Gly Ala Glu Tyr Arg Val Thr

180 185 190

His Gly Asp Asp Pro Val Pro Arg Leu Pro Pro Ile Val Phe Gly Tyr

195 200 205

Arg His Thr Ser Pro Glu Tyr Trp Leu Asn Gly Gly Pro Leu Asp Lys

210 215 220

Asp Tyr Thr Val Thr Glu Ile Lys Val Cys Glu Gly Ile Ala Asn Val

225 230 235 240

Met Cys Asn Gly Gly Thr Ile Gly Leu Asp Ile Leu Ala His Ile Thr

245 250 255

Tyr Phe Gln Ser Met Ala Thr Cys Ala Pro Ile Ala Ile Pro Trp Lys

260 265 270

Arg

<210>11

<211>278

<212>PRT

＜213〉aspergillus oryzae (Aspergillus oryzae)

<400>11

Asp Ile Pro Thr Thr Gln Leu Glu Asp Phe Lys Phe Trp Val Gln Tyr

1 5 10 15

Ala Ala Ala Thr Tyr Cys Pro Asn Asn Tyr Val Ala Lys Asp Gly Glu

20 25 30

Lys Leu Asn Cys Ser Val Gly Asn Cys Pro Asp Val Glu Ala Ala Gly

35 40 45

Ser Thr Val Lys Leu Ser Phe Ser Asp Asp Thr Ile Thr Asp Thr Ala

50 55 60

Gly Phe Val Ala Val Asp Asn Thr Asn Lys Ala Ile Val Val Ala Phe

65 70 75 80

Arg Gly Ser Tyr Ser Ile Arg Asn Trp Val Thr Asp Ala Thr Phe Pro

85 90 95

Gln Thr Asp Pro Gly Leu Cys Asp Gly Cys Lys Ala Glu Leu Gly Phe

100 105 110

Trp Thr Ala Trp Lys Val Val Arg Asp Arg Ile Ile Lys Thr Leu Asp

115 120 125

Glu Leu Lys Pro Glu His Ser Asp Tyr Lys Ile Val Val Val Gly His

130 135 140

Ser Leu Gly Ala Ala Ile Ala Ser Leu Ala Ala Ala Asp Leu Arg Thr

145 150 155 160

Lys Asn Tyr Asp Ala Ile Leu Tyr Ala Tyr Ala Ala Pro Arg Val Ala

165 170 175

Asn Lys Pro Leu Ala Glu Phe Ile Thr Asn Gln Gly Asn Asn Tyr Arg

180 185 190

Phe Thr His Asn Asp Asp Pro Val Pro Lys Leu Pro Leu Leu Thr Met

195 200 205

Gly Tyr Val His Ile Ser Pro Glu Tyr Tyr Ile Thr Ala Pro Asp Asn

210 215 220

Thr Thr Val Thr Asp Asn Gln Val Thr Val Leu Asp Gly Tyr Val Asn

225 230 235 240

Phe Lys Gly Asn Thr Gly Thr Ser Gly Gly Leu Pro Asp Leu Leu Ala

245 250 255

Phe His Ser His Val Trp Tyr Phe Ile His Ala Asp Ala Cys Lys Gly

260 265 270

Pro Gly Leu Pro Leu Arg

275

<210>12

<211>278

<212>PRT

＜213〉penicillium camembertii (Penicillium camemberti)

<400>12

Asp Val Ser Thr Ser Glu Leu Asp Gln Phe Glu Phe Trp Val Gln Tyr

1 5 10 15

Ala Ala Ala Ser Tyr Tyr Glu Ala Asp Tyr Thr Ala Gln Val Gly Asp

20 25 30

Lys Leu Ser Cys Ser Lys Gly Asn Cys Pro Glu Val Glu Ala Thr Gly

35 40 45

Ala Thr Val Ser Tyr Asp Phe Ser Asp Ser Thr Ile Thr Asp Thr Ala

50 55 60

Gly Tyr Ile Ala Val Asp His Thr Asn Ser Ala Val Val Leu Ala Phe

65 70 75 80

Arg Gly Ser Tyr Ser Val Arg Asn Trp Val Ala Asp Ala Thr Phe Val

85 90 95

His Thr Asn Pro Gly Leu Cys Asp Gly Cys Leu Ala Glu Leu Gly Phe

100 105 110

Trp Ser Ser Trp Lys Leu Val Arg Asp Asp Ile Ile Lys Glu Leu Lys

115 120 125

Glu Val Val Ala Gln Asn Pro Asn Tyr Glu Leu Val Val Val Gly His

130 135 140

Ser Leu Gly Ala Ala Val Ala Thr Leu Ala Ala Thr Asp Leu Arg Gly

145 150 155 160

Lys Gly Tyr Pro Ser Ala Lys Leu Tyr Ala Tyr Ala Ser Pro Arg Val

165 170 175

Gly Asn Ala Ala Leu Ala Lys Tyr Ile Thr Ala Gln Gly Asn Asn Phe

180 185 190

Arg Phe Thr His Thr Asn Asp Pro Val Pro Lys Leu Pro Leu Leu Ser

195 200 205

Met Gly Tyr Val His Val Ser Pro Glu Tyr Trp Ile Thr Ser Pro Asn

210 215 220

Asn Ala Thr Val Ser Thr Ser Asp Ile Lys Val Ile Asp Gly Asp Val

225 230 235 240

Ser Phe Asp Gly Asn Thr Gly Thr Gly Leu Pro Leu Leu Thr Asp Phe

245 250 255

Glu Ala His Ile Trp Tyr Phe Val Gln Val Asp Ala Gly Lys Gly Pro

260 265 270

Gly Leu Pro Phe Lys Arg

275

<210>13

<211>270

<212>PRT

＜213〉smelly aspergillus (Aspergillus foetidus)

<400>13

Ser Val Ser Thr Ser Thr Leu Asp Glu Leu Gln Leu Phe Ala Gln Trp

1 5 10 15

Ser Ala Ala Ala Tyr Cys Ser Asn Asn Ile Asp Ser Lys Asp Ser Asn

20 25 30

Leu Thr Cys Thr Ala Asn Ala Cys Pro Ser Val Glu Glu Ala Ser Thr

35 40 45

Thr Met Leu Leu Glu Phe Asp Leu Thr Asn Asp Phe Gly Gly Thr Ala

50 55 60

Gly Phe Leu Ala Ala Asp Asn Thr Asn Lys Arg Leu Val Val Ala Phe

65 70 75 80

Arg Gly Ser Ser Thr Ile Glu Asn Trp Ile Ala Asn Leu Asp Phe Ile

85 90 95

Leu Glu Asp Asn Asp Asp Leu Cys Thr Gly Cys Lys Val His Thr Gly

100 105 110

Phe Trp Lys Ala Trp Glu Ser Ala Ala Asp Glu Leu Thr Ser Lys Ile

115 120 125

Lys Ser Ala Met Ser Thr Tyr Ser Gly Tyr Thr Leu Tyr Phe Thr Gly

130 135 140

His Ser Leu Gly Gly Ala Leu Ala Thr Leu Gly Ala Thr Val Leu Arg

145 150 155 160

Asn Asp Gly Tyr Ser Val Glu Leu Tyr Thr Tyr Gly Cys Pro Arg Ile

165 170 175

Gly Asn Tyr Ala Leu Ala Glu His Ile Thr Ser Gln Gly Ser Gly Ala

180 185 190

Asn Phe Arg Val Thr His Leu Asn Asp Ile Val Pro Arg Val Pro Pro

195 200 205

Met Asp Phe Gly Phe Ser Gln Pro Ser Pro Glu Tyr Trp Ile Thr Ser

210 215 220

Gly Asn Gly Ala Ser Val Thr Ala Ser Asp Ile Glu Val Ile Glu Gly

225 230 235 240

Ile Asn Ser Thr Ala Gly Asn Ala Gly Glu Ala Thr Val Ser Val Leu

245 250 255

Ala His Leu Trp Tyr Phe Phe Ala Ile Ser Glu Cys Leu Leu

260 265 270

<210>14

<211>270

<212>PRT

＜213〉aspergillus niger (Aspergillus niger)

<400>14

Ser Val Ser Thr Ser Thr Leu Asp Glu Leu Gln Leu Phe Ser Gln Trp

1 5 10 15

Ser Ala Ala Ala Tyr Cys Ser Asn Asn Ile Asp Ser Asp Asp Ser Asn

20 25 30

Val Thr Cys Thr Ala Asp Ala Cys Pro Ser Val Glu Glu Ala Ser Thr

35 40 45

Lys Met Leu Leu Glu Phe Asp Leu Thr Asn Asn Phe Gly Gly Thr Ala

50 55 60

Gly Phe Leu Ala Ala Asp Asn Thr Asn Lys Arg Leu Val Val Ala Phe

65 70 75 80

Arg Gly Ser Ser Thr Ile Lys Asn Trp Ile Ala Asp Leu Asp Phe Ile

85 90 95

Leu Gln Asp Asn Asp Asp Leu Cys Thr Gly Cys Lys Val His Thr Gly

100 105 110

Phe Trp Lys Ala Trp Glu Ala Ala Ala Asp Asn Leu Thr Ser Lys Ile

115 120 125

Lys Ser Ala Met Ser Thr Tyr Ser Gly Tyr Thr Leu Tyr Phe Thr Gly

130 135 140

His Ser Leu Gly Gly Ala Leu Ala Thr Leu Gly Ala Thr Val Leu Arg

145 150 155 160

Asn Asp Gly Tyr Ser Val Glu Leu Tyr Thr Tyr Gly Cys Pro Arg Val

165 170 175

Gly Asn Tyr Ala Leu Ala Glu His Ile Thr Ser Gln Gly Ser Gly Ala

180 185 190

Asn Phe Pro Val Thr His Leu Asn Asp Ile Val Pro Arg Val Pro Pro

195 200 205

Met Asp Phe Gly Phe Ser Gln Pro Ser Pro Glu Tyr Trp Ile Thr Ser

210 215 220

Gly Thr Gly Ala Ser Val Thr Ala Ser Asp Ile Glu Leu Ile Glu Gly

225 230 235 240

Ile Asn Ser Thr Ala Gly Asn Ala Gly Glu Ala Thr Val Asp Val Leu

245 250 255

Ala His Leu Trp Tyr Phe Phe Ala Ile Ser Glu Cys Leu Leu

260 265 270

<210>15

<211>269

<212>PRT

＜213〉aspergillus oryzae (Aspergillus oryzae)

<400>15

Asp Val Ser Ser Ser Leu Leu Asn Asn Leu Asp Leu Phe Ala Gln Tyr

1 5 10 15

Ser Ala Ala Ala Tyr Cys Asp Glu Asn Leu Asn Ser Thr Gly Thr Lys

20 25 30

Leu Thr Cys Ser Val Gly Asn Cys Pro Leu Val Glu Ala Ala Ser Thr

35 40 45

Gln Ser Leu Asp Glu Phe Asn Glu Ser Ser Ser Tyr Gly Asn Pro Ala

50 55 60

Gly Tyr Leu Ala Ala Asp Glu Thr Asn Lys Leu Leu Val Leu Ser Phe

65 70 75 80

Arg Gly Ser Ala Asp Leu Ala Asn Trp Val Ala Asn Leu Asn Phe Gly

85 90 95

Leu Glu Asp Ala Ser Asp Leu Cys Ser Gly Cys Glu Val His Ser Gly

100 105 110

Phe Trp Lys Ala Trp Ser Glu Ile Ala Asp Thr Ile Thr Ser Lys Val

115 120 125

Glu Ser Ala Leu Ser Asp His Ser Asp Tyr Ser Leu Val Leu Thr Gly

130 135 140

His Ser Tyr Gly Ala Ala Leu Ala Ala Leu Ala Ala Thr Ala Leu Arg

145 150 155 160

Asn Ser Gly His Ser Val Glu Leu Tyr Asn Tyr Gly Gln Pro Arg Leu

165 170 175

Gly Asn Glu Ala Leu Ala Thr Tyr Ile Thr Asp Gln Asn Lys Gly Gly

180 185 190

Asn Tyr Arg Val Thr His Thr Asn Asp Ile Val Pro Lys Leu Pro Pro

195 200 205

Thr Leu Leu Gly Tyr His His Phe Ser Pro Glu Tyr Tyr Ile Ser Ser

210 215 220

Ala Asp Glu Ala Thr Val Thr Thr Thr Asp Val Thr Glu Val Thr Gly

225 230 235 240

Ile Asp Ala Thr Gly Gly Asn Asp Gly Thr Asp Gly Thr Ser Ile Asp

245 250 255

Ala His Arg Trp Tyr Phe Ile Tyr Ile Ser Glu Cys Ser

260 265

<210>16

<211>251

<212>PRT

<213>Landerina penisapora

<400>16

Pro Gln Asp Ala Tyr Thr Ala Ser His Ala Asp Leu Val Lys Tyr Ala

1 5 10 15

Thr Tyr Ala Gly Leu Ala Tyr Gln Thr Thr Asp Ala Trp Pro Ala Ser

20 25 30

Arg Thr Val Pro Lys Asp Thr Thr Leu Ile Ser Ser Phe Asp His Thr

35 40 45

Leu Lys Gly Ser Ser Gly Tyr Ile Ala Phe Asn Glu Pro Cys Lys Glu

50 55 60

Ile Ile Val Ala Tyr Arg Gly Thr Asp Ser Leu Ile Asp Trp Leu Thr

65 70 75 80

Asn Leu Asn Phe Asp Lys Thr Ala Trp Pro Ala Asn Ile Ser Asn Ser

85 90 95

Leu Val His Glu Gly Phe Leu Asn Ala Tyr Leu Val Ser Met Gln Gln

100 105 110

Val Gln Glu Ala Val Asp Ser Leu Leu Ala Lys Cys Pro Asp Ala Thr

115 120 125

Ile Ser Phe Thr Gly His Ser Leu Gly Gly Ala Leu Ala Cys Ile Ser

130 135 140

Met Val Asp Thr Ala Gln Arg His Arg Gly Ile Lys Met Gln Met Phe

145 150 155 160

Thr Tyr Gly Gln Pro Arg Thr Gly Asn Gln Ala Phe Ala Glu Tyr Val

165 170 175

Glu Asn Leu Gly His Pro Val Phe Arg Val Val Tyr Arg His Asp Ile

180 185 190

Val Pro Arg Met Pro Pro Met Asp Leu Gly Phe Gln His His Gly Gln

195 200 205

Glu Val Trp Tyr Glu Gly Asp Glu Asn Ile Lys Phe Cys Lys Gly Glu

210 215 220

Gly Glu Asn Leu Thr Cys Glu Leu Gly Val Pro Phe Ser Glu Leu Asn

225 230 235 240

Ala Lys Asp His Ser Glu Tyr Pro Gly Met His

245 250

Claims (14)

1. detergent composition, described composition comprises the variant of detergent ingredients and parent lipase, when comparing with described parent, described variant comprises at least three kinds of replacements of total, and described parent lipase variant is the sequence identification number with T231R+N233R+I255Y or I90A+T231R+N233R+I255V replacement: 2.

2. detergent composition as claimed in claim 1, described detergent composition also comprises 0.1% to 40% anion surfactant.

3. detergent composition as claimed in claim 2, the content of wherein said anion surfactant is 0.1% to 12%.

4. detergent composition as claimed in claim 2, wherein said anion surfactant is oxyalkylated alkyl-sulphate.

5. detergent composition as claimed in claim 1, described detergent composition also comprises 5% to 30% silico-aluminate and/or phosphate builders.

6. detergent composition as claimed in claim 1, described detergent composition also comprises peroxide source and bleach-activating agent.

7. detergent composition as claimed in claim 1, wherein said detergent composition is liquid detergent composition or solid detergent composition.

8. detergent composition as claimed in claim 7, wherein said detergent composition is detergent granules.

9. detergent composition as claimed in claim 1, wherein said detergent composition are the units dosage compositions of solid tablet or the liquid sealed with dissolvable film.

10. washing methods, described method is included in the aqueous wash medium that comprises detergent composition as claimed in claim 1 and washs textiles.

11. washing methods as claimed in claim 10, wherein said method are suitable for dirt and spot are removed from the surface, said method comprising the steps of:

A) detergent composition as claimed in claim 1 with significant quantity is added to the water, and comprises the wash water solution of the described composition of 500ppm to 10000ppm with formation;

B) make described wash water solution and described Surface Contact.

12. washing methods as claimed in claim 10, the temperature of the wherein said aqueous solution are lower than 30 ℃.

13. washing methods as claimed in claim 11 said method comprising the steps of:

A) with the described dirt of detergent composition pre-treatment as claimed in claim 1 and spot to form the surface of pretreated mistake;

B) detergent composition as claimed in claim 1 with significant quantity is added to the water, and comprises the wash water solution of the described composition of 500ppm to 10000ppm with formation;

C) make the described Surface Contact of described wash water solution and pretreated mistake.

14. washing methods as claimed in claim 11, described method also comprises steps d): the surface to described wash water solution and pretreated mistake provides stirring.

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