CN1113089C - Liquid detergents containing proteolytic enzyme and protease inhibitors - Google Patents

Abstract

The present invention describes aqueous liquid detergent compositions comprising a protease wherein the proteolytic activity is reversibly inhibited by a protease inhibitor selected from the group consisting of aldehydes and trifluoromethyl ketones.

Description

Liquid detergents containing proteases and protease inhibitors

technical field

The present invention relates to enzyme-containing liquid detergent compositions. More particularly, the present invention relates to liquid detergent compositions comprising a detersive surfactant, a protease and a protease inhibitor selected from the group consisting of aldehydes and trifluoromethyl ketones.

Background of the invention

Aqueous liquid detergents containing proteases are well known, especially in the field of laundry. A problem commonly encountered in such protease-containing aqueous liquid detergents is the degradation of secondary enzymes such as amylases, lipases and cellulases in the composition by proteases and by proteases on their own proteases . As a result the stability of the second enzyme or protease itself in the detergent composition is affected and thus the detergent composition does not work as well.

In response to this problem, various protease inhibitors or stabilizers have been proposed. For example, various references suggest the use of the following compounds to aid in enzyme stabilization: benzamidine hydrochloride, lower aliphatic alcohols or carboxylic acids, mixtures of polyols and boron compounds, aromatic borates, and calcium, especially formic acid calcium. Recently, certain peptide aldehydes and peptide trifluoromethyl ketones were found to stabilize proteases.

Despite all the success these compounds have had in liquid detergents, they are not without their problems. For example, certain peptide aldehydes can be quite expensive and present complexities to the formulator, especially of liquid detergents. Other inhibitors such as calcium and boric acid are less expensive, but they do not stabilize the enzyme as well as peptide aldehydes. It is therefore an object of the present invention to provide protease inhibitors which are economical, effective and suitable for use in liquid detergent compositions.

Background technique

The use of various protease inhibitors or stabilizers has been proposed. For example US4566985 proposes the use of benzamidine hydrochloride; EP376705 proposes the use of lower aliphatic alcohols or carboxylic acids; EP381262 proposes the use of mixtures of polyhydric alcohols and boron compounds; and EP91870072.5 proposes the use of aromatic borates. See also US Patent 5,030,378 issued Jul. 9,1991. See also US4261868; US4404115; US4318818; and EP130756.

The use of peptide derivatives to inhibit proteases appears to be disclosed in therapeutic applications. EP293881 discloses the use of peptide boronic acids as inhibitors of trypsin-like serine proteases. EP185390 and US4399065 disclose the use of certain peptide aldehyde derivatives to inhibit blood coagulation. J90029670 discloses the inhibition of general enzymes using optically active a-aminoaldehydes. See also "Inhibition of Thrombin and Trypsin by Tripeptide Aldehydes", Int. J. Peptide Protein Res. , Vol. 12 (1978), pp. 217-221; Gaal, Bacsy & Rappay, and "Tripeptide Aldehyde Protease Inhibitors inhibit the release of prolactin and growth hormone in vitro", Endocrinology , Vol. 116, No. 4 (1985), pp. 1426-1432; Rappay, Makara, Bajusz & Nagy. Certain peptide aldehydes are also disclosed in EP-A-473502 for the inhibition of protease mediated skin irritation.

See in particular EP185390, WO94/04651, published March 3, 1994; WO94/04652, published March 3, 1994, EP583536, published February 23, 1994, EP583535, published February 3, 1994, EP583534, Published February 23, 1994, WO93/13125, published July 8, 1993, US4529525, US4537706, US4537707 and US5527487.

Summary of the invention

The present invention is a liquid detergent composition comprising: a) an effective amount of a detersive surfactant; b) an active protease; and c) a protease inhibitor having the formula:

ZA-NH-CH(R)-C(O)-X wherein A is an amino acid moiety; X is hydrogen or _CF3 ; Z is an N-terminated moiety selected from carbamate, urea, sulfonamide, phosphine Amides, thioureas, sulfenamides, sulfonic acids, phosphinamides, thiocarbamates, amidophosphoryl esters, sulfonamide derivatives and phosphonamides; and R is selected from linear or branched C ₁ -C ₆ Unsubstituted alkyl, phenyl and C ₇ -C ₉ alkaryl moieties. Preferred compositions also include a source of calcium ions or boric acid.

Preferably the liquid detergent compositions of the present invention comprise, by weight of the composition: a) from about 1% to about 95%, preferably from about 8% to about 70%, of said detersive surfactants; b) from about 0.0001% to about 5%, preferably about 0.0003% to about 0.1% active protease; c) about 0.00001% to about 5%, preferably about 0.0001% to about 1%, more preferably about 0.0006% to about 0.5% of the protease inhibitor; d ) optionally, about 0.01% to about 1%, preferably about 0.05% to about 0.5% calcium ion; and e) optionally, about 0.25% to about 10%, preferably about 0.5% to about 5% boric acid or can The compound generating boronic acid preferably contains a diol.

The proteases used in the present invention are preferably subtilisin-type proteases, which may be selected from Alcalase ^R , subtilisin BPN', Protease A, Protease B and mixtures thereof.

The source of calcium ions used in the present invention is preferably selected from calcium formate, calcium xylene sulfonate, calcium chloride, calcium acetate, calcium sulfate and mixtures thereof.

The dishwashing compositions of the present invention may contain additional soil release additives including, but not limited to: one or more of the following: suds boosters, chelating agents, polyacrylate polymers, dispersants, dyes, fragrances, processing aids agents and their mixtures. Additionally, for dishwashing compositions, the liquid detergent compositions may also contain an effective amount of an amylase. Additionally, the dishwashing composition may optionally contain an effective amount of a source of boric acid and a glycol. Typically, dishwashing compositions optionally, but preferably comprise from about 0.25% to about 10%, preferably from about 0.5% to about 5%, more preferably from about 0.75% to about 3% by weight of boric acid or a compound capable of generating boric acid and Diols such as 1,2-propanediol.

In a preferred embodiment of the heavy duty detergent composition for laundry, the liquid detergent composition further comprises an effective amount of one or more of the following enzymes: lipase, amylase, cellulase and mixtures thereof. For laundry compositions, it is preferred that the second enzyme is a lipase obtained by cloning a gene from Humicola lanuginosa and expressing the gene in Aspergillus oryzae. Lipase is used in an amount of about 10 to about 18000 lipase units/gram, preferably about 60 to about 6000 units/gram.

In another preferred composition for laundry use, the second enzyme is a cellulase obtained from Humicola Insolens in an amount of from about 0.0001% to about 0.1% by weight of the total composition of said cellulase .

The compositions of the present invention may contain additional soil release additives including, but not limited to: one or more of the following components: suds boosters, builders, soil release polymers, polyacrylate polymers, dispersants, dyes Transfer inhibitors, dyes, fragrances, processing aids, brighteners and mixtures thereof. Additionally, for laundry compositions, detersive surfactants will typically be present at levels from about 10% to about 70% by weight of the total composition. Furthermore, laundry compositions can optionally contain effective amounts of a source of boric acid and a glycol. Laundry compositions generally optionally, but preferably comprise from about 0.25% to about 10%, preferably from about 0.5% to about 5%, more preferably from about 0.75% to about 3% by weight boric acid or a compound capable of generating boric acid and glycol , such as 1,2-propanediol.

Unless specifically stated otherwise, all percentages and ratios herein are by weight and all documents cited are hereby incorporated by reference.

Detailed description of the invention

Definitions - The detergent compositions of the present invention comprise an "effective amount" or "stain removal improving amount" of each ingredient as defined herein. "Effective amount" or "stain removal improving amount" refers to any amount that measurably improves soil cleaning or stain removal from substrates such as soiled fabrics or soiled dishes when laundered by a consumer. Generally, this amount can vary over a wide range.

Aqueous liquid detergent compositions according to the present invention comprise three essential components: (A) a protease inhibitor selected from the group consisting of aldehydes and trifluoromethyl ketones or mixtures thereof as described herein, (B) proteases or mixtures thereof, and (C) detersive surfactants. Compositions according to the invention preferably further comprise (D) a source of calcium ions, (E) a second detergent-compatible enzyme or a mixture thereof, (F) boric acid and a glycol, and may additionally comprise (G) other Optional components.

Protease Inhibitors - The detergent compositions of the present invention comprise as a first essential ingredient a protease inhibitor selected from the group consisting of aldehydes and trifluoromethyl ketones having the formula:

ZA-NH-CH(R)-C(O)-X wherein A is an amino acid moiety (preferably the A moiety is selected from Ala, Gly, Val, Ile, Leu, Phe, Lys); X is hydrogen or _CF3 ; Z is an N-terminated moiety selected from phosphoramidate [(R'O) ₂ (O)P-], sulfenamide [(SR') ₂ -], sulfonamide [(R'(O) ₂ S -], sulfonic acid [SO ₃ H], phosphinamide [(R') ₂ (O)P-], sulfonamide derivatives [R'O(O) ₂ S-], thiourea [(R' ) ₂ N(S)C-], thiocarbamate [R'O(S)C-], phosphonate [R'-P(O)OH], amidophosphoryl ester [R'O( OH)(O)P-], carbamate (R'O(O)C-) and urea (R'NH(O)C-), wherein each R" is independently selected from linear or branched C ₁ -C ₆ unsubstituted alkyl, phenyl, C ₇ -C ₉ alkaryl and cycloalkyl moieties, wherein the cycloalkyl ring can span C ₄ -C ₈ and can contain one or more selected from heteroatoms of O, N, and S; and R are selected from linear or branched C ₁ -C ₆ unsubstituted alkyl, phenyl, and C ₇ -C ₉ alkylaryl moieties.

Preferred R moieties are selected from methyl, isopropyl, sec-butyl, isobutyl, -C ₆ H ₅ , -CH ₂ -C ₆ H ₅ and -CH ₂ CH ₂ -C ₆ H ₅ , which are derived from From the amino acids Ala, Val, Ile, Leu, PGly (phenylglycine), Phe and HPhe (homophenylalanine), by converting the carboxylic acid group into an aldehyde or trifluoromethylketone group. Although these moieties are thus no longer amino acids (and they may or may not be synthesized from amino acid precursors), to simplify the description of inhibitors applicable here, the aldehyde moieties of the inhibitors are denoted as derived from amino acids by Add "H" after [eg, "-AlaH" represents the chemical moiety "-NHCH( _CH3 )C(O)H"]. Trifluoromethyl ketones can be similarly represented by appending " _CF3 " to the analogous amino acid (eg " _-AlaCF3 " represents the chemical moiety "-NHCH( _CH3 )C(O) _CF3 "].

The aldehydes according to the invention can be prepared from the corresponding amino acids, whereby the C-terminus of said amino acids is converted from a carboxyl group to an aldehyde group. Such aldehydes can be prepared by known methods, for example as described in US5015627, EP185930, EP583534 and DE3200812.

The trifluoromethyl ketones according to the invention can be prepared from the corresponding amino acids, whereby the C-terminus of said amino acids is converted from a carboxyl group to a trifluoromethyl ketone group. Such trifluoromethyl ketones can be prepared by known methods, for example as described in EP583535.

While not wishing to be bound by theory, it is believed that the protease inhibitors of the present invention bind proteases in liquid detergent compositions, thereby inhibiting said proteases. When diluted in water, proteolytic activity is restored due to dissociation of the protease/protease inhibitor complex.

The N-terminus of said protease inhibitor of the present invention is protected by a kind of N-blocking partial protecting group, and said N-blocking partial protecting group is selected from carbamate, urea, sulfonamide, phosphonamide , thiourea, sulfenamide, sulfonic acid, phosphinamide, thiocarbamate, amidophosphoramidate, and phosphonamide. However, in a highly preferred embodiment of the invention, the N-terminus of said protease inhibitor is covered with a methyl, ethyl or benzyl carbamate [CH ₃ O-(O)C-; CH ₃ CH ₂ O-( O)C-; or C ₆ H ₅ CH ₂ O-(O)C-]; methyl, ethyl or benzyl urea [CH ₃ NH-(O)C-; CH ₃ CH ₂ NH-(O) C-; or C ₆ H ₅ CH ₂ NH-(O)C-]; methyl, ethyl or benzylsulfonamide [CH ₃ SO ₂ -; CH ₃ CH ₂ SO ₂ -; or C ₆ H ₅ CH ₂ SO ₂ -], and methyl, ethyl or benzyl amidophosphoramido [CH ₃ O(OH)(O)P-; CH ₃ CH ₂ O(OH)(O)P-; or C ₆ H ₅ CH ₂ O(OH)(O)P-] group protection.

The synthesis of N-capping groups can be found in the following references: Protecting Groups in Organic Chemistry , Greene, T., Wuts, P., John Wiley & Sons, New York, 1991, pp. 309-405; March, J, Advanced Organic Chemistry , Wiley Interscience, 1985, pp. 445, 469; Carey, F. Sundberg, R., Advanced Organic Chemistry , Part B, PlenumPress, New York, 1990, pp. 686-89; Atherton, E., Sheppard, R., Solid Phase Peptide Synthesis , Pierce Chemical, 1989, pp. 3-4; Grant, G., Synthetic Peptides , WH Freeman & Co. 1992, pp. 77-103; Stewart, J., Young, J. Phase Peptide Synthesis , 2nd Edition, IRL Press, 1984, pp. 3, 5, 11, 14-18, 28-29. Bodansky, M., Principles of Peptide Synthesis , Springer-Verlag, 1988, pp. 62, 203, 59-69; Bodansky, M., Peptide Chemistry , Springer-Verlag, 1988, pp. 74-81, Bodansky, M., Bodansky, A., Experimental Procedures for Peptide Synthesis , Springer-Verlag, 1984, pp. 9-32.

Examples of protease inhibitors useful in the present invention are: _CH3O- (O)C _- Ala-LeuH _{; CH3CH2O- (O)C-Ala-LeuH; C6H5CH2O- (} O )C-Ala-LeuH; CH ₃ O-(O)C-Ala-LeuCF ₃ ; CH ₃ CH ₂ O-(O)C-Ala-LeuCF ₃ ; C ₆ H ₅ CH ₂ O-(O)C- Ala-LeuCF ₃ ; CH ₃ O-(O)C-Ala-IleH; CH ₃ CH ₂ O-(O)C-Ala-IleH; C ₆ H ₅ CH ₂ O-(O)C-Ala-IleH; _CH3O- (O)C- _Ala - _IleCF3 ; _CH3CH2O- (O) _C -Ala- _IleCF3 ; _C6H5CH2O- (O)C- _Ala - _IleCF3 ; _CH3 O-(O)C-Gly _- LeuH; _CH3CH2O- (O) _C -Gly-LeuH; _C6H5CH2O- (O)C-Gly- _LeuH ; _CH3O- (O) C-Gly-LeuCF ₃ ; CH ₃ CH ₂ O-(O)C-Gly-LeuCF ₃ ; C ₆ H ₅ CH ₂ O-(O)C-Gly-LeuCF ₃ ; CH ₃ O-(O)C- Gly-IleH; _CH3CH2O- (O)C-Gly-IleH; _C6H5CH2O- (O)C- _Gly -IleH; _CH3O- (O) _C -Gly _{- IleCF3} ; _CH3CH2O- (O)C-Gly- _IleCF3 ; _C6H5CH2O- (O)C-Gly- _IleCF3 ; _CH3NH- (O) _{C -} Ala-LeuH _{; CH3CH 2} NH-(O)C-Ala-LeuH; C ₆ H ₅ CH ₂ NH-(O)C-Ala-LeuH; CH ₃ NH-(O)C-Ala-LeuCF ₃ ; CH ₃ CH ₂ NH-( O)C-Ala-LeuCF ₃ ; C ₆ H ₅ CH ₂ NH-(O)C-Ala-LeuCF ₃ ; CH ₃ NH-(O)C-Ala-IleH; CH ₃ CH ₂ NH-(O)C -Ala-IleH; C ₆ H ₅ CH ₂ NH-(O)C-Ala-IleH; CH ₃ NH-(O)C-Ala-IleCF ₃ ; CH ₃ CH ₂ NH-(O)C-Ala-IleCF ₃ ; C ₆ H ₅ CH ₂ NH-(O)C-Ala-IleCF ₃ ; CH ₃ NH-(O)C-Gly-LeuH; CH ₃ CH ₂ NH-(O)C-Gly-LeuH; C ₆ H ₅ CH ₂ NH-(O)C-Gly-LeuH; CH ₃ NH-(O)C-Gly-LeuCF ₃ ; CH ₃ CH ₂ NH-(O)C-Gly-LeuCF ₃ ; C ₆ H ₅ CH ₂ NH-(O)C-Gly-LeuCF ₃ ; CH ₃ NH-(O)C-Gly-IleH; CH ₃ CH ₂ NH-(O)C-Gly-IleH; C ₆ H ₅ CH ₂ NH-( O)C-Gly-IleH; CH ₃ NH-(O)C-Gly-IleCF ₃ ; CH ₃ CH ₂ NH-(O)C-Gly-IleCF ₃ ; C ₆ H ₅ CH ₂ NH-(O)C CH ₃ SO ₂ -Ala-LeuH; CH ₃ CH ₂ SO ₂ -Ala _- LeuH; C ₆ H ₅ CH ₂ SO ₂ -Ala-LeuH; CH ₃ SO ₂ -Ala-LeuCF ₃ ; CH ₃ CH ₂ SO ₂ -Ala-LeuCF ₃ ; C ₆ H ₅ CH ₂ SO ₂ -Ala-LeuCF ₃ ; CH ₃ SO ₂ -Ala-IleH; CH ₃ CH ₂ SO ₂ -Ala-IleH; C ₆ H ₅ CH _{2SO 2} -Ala-IleH; CH ₃ SO ₂ -Ala-IleCF ₃ ; CH ₃ CH ₂ SO ₂ -Ala-IleCF ₃ ; C ₆ H ₅ CH ₂ SO ₂ -Ala-IleCF ₃ ; CH ₃ SO ₂ -Gly _-LeuH ; _{CH3CH2SO2 -} Gly _- LeuH; C6H5CH2SO2- _{Gly - LeuH} ; _{CH3SO2 -} Gly _{- LeuCF3} ; _CH3CH2SO2 - _Gly - _LeuCF3 ; ₆ H ₅ CH ₂ SO ₂ -Gly-LeuCF ₃ ; CH ₃ SO ₂ -Gly-IleH; CH ₃ CH ₂ SO ₂ -Gly-IleH; C ₆ H ₅ CH ₂ SO ₂ -Gly-IleH; CH ₃ SO ₂ -Gly-IleCF ₃ ; CH ₃ CH ₂ SO ₂ -Gly-IleCF ₃ ; C ₆ H ₅ CH ₂ SO ₂ -Gly-IleCF ₃ ; CH ₃ O(OH)(O)P-Ala-LeuH; CH ₃ CH ₂ O(OH)(O)P-Ala-LeuH; C ₆ H ₅ CH ₂ O(OH)(O)P-Ala-LeuH; CH ₃ O(OH)(O)P-Ala-LeuCF ₃ ; CH ₃ CH ₂ O(OH)(O)P-Ala-LeuCF ₃ ; C ₆ H ₅ CH ₂ O(OH)(O)P-Ala-LeuCF ₃ ; CH ₃ O(OH)(O)P-Ala- IleH; _CH3CH2O (OH)(O ₎ P-Ala-IleH; _C6H5CH2O (OH)(O)P- _Ala -IleH; _CH3O (OH) ₍ O)P-Ala -IleCF ₃ ; CH ₃ CH ₂ O(OH)(O)P-Ala-IleCF ₃ ; C ₆ H ₅ CH ₂ O(OH)(O)P-Ala-IleCF ₃ ; CH ₃ O(OH)(O )P-Gly-LeuH; CH ₃ CH ₂ O(OH)(O)P-Gly-LeuH; C ₆ H ₅ CH ₂ O(OH)(O)P-Gly-LeuH; CH ₃ O(OH)( O)P-Gly-LeuCF ₃ ; CH ₃ CH ₂ O(OH)(O)P-Gly-LeuCF ₃ ; C ₆ H ₅ CH ₂ O(OH)(O)P-Gly-LeuCF ₃ ; CH ₃ O (OH)(O)P-Gly-IleH _{; CH3CH2O} (OH _{) (} O)P-Gly-IleH; _C6H5CH2O (OH)(O)P-Gly-IleH; _CH3 O(OH)(O)P-Gly-IleCF ₃ ; CH ₃ CH ₂ O(OH)(O)P-Gly-IleCF ₃ ; and C ₆ H ₅ CH ₂ O(OH)(O)P-Gly- IleCF ₃ .

In the Synthetic Examples below, methods for the synthesis of certain of these peptide protease inhibitors are disclosed.

Synthesis Example 1:

Synthesis of Aldehyde Protease Inhibitors

Moc-Leu-OH - L-Leucine (5.0 g, 38.2 mmol) was dissolved in 38 ml 1 N NaOH and cooled to 0°C. Methyl chloroformate (3.1 ml, 40.0 mmol) was added dropwise while adding 1 N NaOH in a separate addition funnel, maintaining the pH at 9.0-9.5. After the addition was complete and the pH stabilized at 9.0-9.5, the solution was washed with 200 ml EtOAc and the aqueous phase was then acidified to pH=2. The mixture was extracted with EtOAc (2 x 100ml), dried ( _MgSO4 ), filtered and the solvent removed to give 7.15g of pure product.

Moc-Leu-Leucinol - To a solution of 3.5 g (18.52 mmol) Moc-Leu-OH in 100 ml THF cooled to -15°C was added 2.04 ml (18.52 mmol) N-methylmorpholine, Then 2.4 ml (18.52 mmol) of isobutyl chloroformate were added immediately. After stirring for 10 min, 2.37 ml (18.52 mmol) leucinol in 25 ml THF was added and the reaction was stirred at -15°C for 0.5 h and at room temperature for 1 h. The mixture was then diluted with 100ml _H2O and THF was evaporated. The remaining aqueous phase was partitioned between EtOAc and 1N HCl, the organic phase was washed with _NaHCO3 , dried ( _MgSO4 ) and evaporated to give 5.33g of pure product.

Moc-Leu-LeuH - A solution containing _4.4 g (10.41 mmol) of Dess-Martin periodinane suspended in 100 ml _CH2Cl2 was prepared and stirred for 10 minutes. To this solution was added 1.0 g (3.47 mmol) Moc-Leu-leucinol, and the solution was stirred at room temperature for 2 h, then poured into 100 ml saturated NaHCO containing 18 g (72.87 _mmol ) Na ₂ S ₂ O ₃ . The solution was stirred for 10 minutes, then extracted with EtOAc (2X, 125ml), dried ( _MgSO4 ) and the solvent evaporated. Chromatography on silica gel afforded 0.550 g of pure product.

Synthesis Example 2

Synthesis of Trifluoromethyl Ketoprotease Inhibitor

N-Trityl leucine methyl ester: To a solution of 2.50 g (13.8 mmol) Leu-OMe.HCl in 100 ml _CH2Cl2 was added dropwise 3.86 ml TEA (27.5 _mmol ). After the dropwise addition was complete, 3.76 g (13.5 mmol) of triphenylmethane chloride in 15 ml _CH2Cl2 were added _dropwise . The mixture was stirred for 4 hours. The solution was diluted with 5% EtOAc/petroleum ether and washed with water. The organic phase was dried ( _MgSO4 ), filtered and the solvent was removed. The residue was chromatographed on silica gel to give 4.8 g of pure product (90% yield).

N-trityl leucine aldehyde: To a cold (0° C.) solution of 4.70 g (12.2 mmol) of N-trityl leucine methyl ester in 100 ml THF was added dropwise 28.1 ml of 1.5 M Diisobutylaluminum hydride (42.2 mol) solution. The solution was stirred at this temperature for 6 hours, the reaction was quenched with saturated Na-K tartrate, extracted with EtOAc, dried ( _MgSO4 ), filtered, and the solvent was removed. The 4.13 g of desired material recovered was used without purification. To a solution of 1.29 g (14.9 mmol) oxalyl chloride in 20 ml _CH2Cl2 was added dropwise 2.26 ml DMSO (29.8 _{mmol )} in 5 ml _CH2Cl2 at -78°C. After the addition was complete, 4.13 _g (11.5 mmol) of crude N-tritylleucinol in 10 ml _CH2Cl2 were added. The solution was warmed to 0°C and poured into a mixture of water and ether. The phases were separated, the ether phase was dried ( _MgSO4 ) and evaporated to give 1.37 g of the desired compound.

5-Methyl-3-tritylamino-1,1,1-trifluoro-2-hexanol: To 1.37g (3.83mmol) N-trityl leucinaldehyde and 0.653ml in THF (4.59 mmol) of CF ₃ TMS solution was added in one portion of 0.121 g (0.383 mmol) of tetrabutylammonium fluoride trihydrate. The solution was stirred at room temperature for 3 hours and the solvent was removed. The residue was dissolved in EtOAc, washed with water, dried ( _MgSO4 ) and the solvent removed to give 1.20 g of product which was chromatographed on silica gel (0.760 g of pure product recovered).

Moc-Ala-OH-Alanine (5.0 g, 56.2 mmol)) was dissolved in 56 ml 1 N NaOH and cooled to 0°C. Methyl chloroformate (5.57ml, 58.9mmol) was added dropwise while adding 1N NaOH in a separate addition funnel, maintaining the pH at 9.0-9.5. After the addition was complete and the pH stabilized at 9.0-9.5, the solution was washed with 200 ml EtOAc and the aqueous phase was then acidified to pH=2. The mixture was extracted with EtOAc (2 x 100ml), dried ( _MgSO4 ), filtered and the solvent removed to give 7.15g of pure product.

3-(N-(Moc-Ala))-5-methyl-1,1,1-trifluoro-2-hexanol: To 1.21g (2.83mmol) 5-methyl- To the solution of 3-tritylamino-1,1,1-trifluoro-2-hexanol was added 5 ml of 4.0M HCl in dioxane. The solution was stirred at room temperature for 2 hours, and the solvent was removed. The residue was triturated with ether and the solid material was filtered. The resulting HCl salt (0.627 g, 2.83 _mmol ) was suspended in 10 ml _CH2Cl2 and Moc-Ala-OH (0.416 g, 2.83 mmol) was added. To this mixture was added 0.870ml (6.23mmol) TEA followed immediately by 0.473ml (3.12mmol) DEPC. The mixture was stirred overnight and the solvent was removed. The residue was dissolved in EtOAc, washed with 1N HCl, saturated NaHCO ₃ and brine. The product solution was dried (MgSO ₄ ), filtered, and the solvent was removed to obtain 0.650 g of product.

Moc-Ala-LeuCF ₃ : To a slurry of _{2.63 g (6.21 mmol) of Dess-Martin periiodane in 15 ml CH 2 Cl 2 was added} 0.650 g (2.07 mmol) of 3-(Moc -Ala)-5-methyl-1,1,1-trifluoro-2-hexanol, the slurry was stirred for 3 hours. To this mixture was added 10.88 g (43.47 mmol) _Na2S2O3 in 50 ml saturated _NaHCO3 and the resulting solution was _stirred for 10 _minutes . The solution was extracted with EtOAc, the organic phase was dried ( _MgSO4 ), filtered and the solvent was removed. The residue was chromatographed on silica gel to give 0.425 g of pure product. Z = benzyl ester group Gly = glycine Ala = alanine Leu = leucine Phe = phenylalanine OMe = methyl ester TEA = triethylamine DECP = diethyl cyanophosphonate TLC = thin layer chromatography MeOH = Methanol Pd/C = palladium on activated carbon EtOH = ethanol THF = tetrahydrofuran Mac = methylaminocarbonyl Moc = methoxycarbonyl

Proteases - Another essential ingredient in the liquid detergent compositions of the present invention are active proteases. Also included are mixtures of proteases. Proteases may be of animal, vegetable or microbial (preferred) origin. Proteases useful in the detergent compositions of the present invention include, but are not limited to, trypsin, subtilisin, chymotrypsin and elastase-type proteases. Preferred for use in the present invention are subtilisin-type proteases. Particularly preferred are bacterial serine proteases derived from Bacillus subtilis and/or Bacillus licheniformis. Proteases are typically present in such liquid detergent compositions in an amount sufficient to provide 0.005-0.1 Anson Units (AU) of activity per gram of composition.

Suitable proteases include Novo Industri A/S Alcalase ^_ (preferred), Esperase ^_ , Savinase ^_ (Copenhagen, Denmark), Gist-brocades' Maxatase ^_ , Maxacal ^_ and Maxapem 15 ^_ (Maxacal ^_ of Protease Engineering) (Delft, Netherlands) and the subtilisins BPN and BPN' (preferred), which are commercially available. Preferred proteases are also modified bacterial serine proteases, such as those prepared by Genencor International, Inc. (San Francisco, California), which are described in European Patent 251446, filed April 28, 1987 (particularly Nos. 17, 24 and 98 pages), referred to herein as "Protease B," and U.S. Patent 5,030,378 to Venegas, issued July 9, 1991, which relates to modified bacterial serine proteases (Genencor International), referred to herein as is "Protease A" (same as BPN'). See especially US Patent No. 5,030,378, columns 2 and 3, for a complete description, including the amino acid sequences of Protease A and variants thereof. Preferred proteases are then selected from ^Alcalase® (Novo Industri A/S), BPN', Protease A and Protease B (Genencor) and mixtures thereof. Protease B is most preferred.

Another preferred protease known as "Protease D" is a carbonyl hydrolase variant having an amino acid sequence not found in nature, which is derived from a carbonyl hydrolase precursor according to the Numbering of the bacillin, substituting a different amino acid for the amino acid residue corresponding to position +76 in said carbonyl hydrolase, also preferably combining substitutions corresponding to the group selected from +99, +101, +103, +104 , +107, +123, +27, +105, +109, +126, +128, +135, +156, +166, +195, +197, +204, +206, +210, +216, + One or more amino acid residues at positions 217, +218, +222, +260, +265, and/or +274 as described in WO 95/10615 published April 20, 1995 by Genencor International.

Useful proteases are also described in PCT publications: WO95/30010, published November 9, 1995, The Procter & Gamble Company; WO95/30011, published November 9, 1995, The Procter & Gamble Company; 1995, The Procter & Gamble Company WO95/29979 published November 9.

Detersive Surfactants - An effective amount, typically about 1-95%, preferably about 8-70% by weight, of detersive surfactants is another essential component of the present invention. The detersive surfactant may be selected from anionic, nonionic, cationic, amphoteric, zwitterionic surfactants and mixtures thereof. By selection of the type and amount of detersive surfactant and other additional ingredients disclosed herein, the detergent compositions of the present invention can be formulated for use in laundry cleaning or other various cleaning applications, including especially dishwashing. The particular surfactant employed can thus vary widely depending upon the particular end use envisioned.

The benefits of the present invention are especially judged in compositions containing enzyme harsh components such as certain detergency builders and surfactants. These include, but are not limited to, anionic surfactants such as alkyl ether sulfates, linear alkylbenzene sulfonates, alkyl sulfates, and the like. Suitable surfactants are described below.

Anionic Surfactants - One class of anionic surfactants that may be used includes alkyl ester sulfonates. They are desirable because they can be prepared from renewable, non-petroleum sources. The preparation of the alkyl ester sulfonate surfactant component can be carried out according to known methods disclosed in the technical literature. For example, linear esters of C ₈ -C ₂₀ carboxylic acids can be obtained in the gaseous state according to the method of "The Journal of the American Oil Chemists Society", 52 (1975), pp. 323-329. SO ₃ sulfonation. Suitable materials include natural fatty materials such as those derived from tallow, palm oil and coconut oil.

Preferred alkyl ester sulfonate surfactants particularly suitable for laundry applications include those having the formula:

R ³ -CH(SO ₃ M)-C(O)-OR ⁴ wherein R ³ is a C ₈ -C ₂₀ hydrocarbon group, preferably an alkyl group, or a combination thereof, R ⁴ is a C ₁ -C ₆ hydrocarbon group, preferably an alkyl group, or a combination thereof, M is a cation that forms a soluble salt. Suitable salts include metal salts such as sodium, potassium and lithium salts, and substituted or unsubstituted ammonium salts such as methyl-, dimethyl-, trimethyl- and quaternary ammonium cations such as tetramethylammonium and dimethylammonium Alkyridiniums and cations derived from alkanolamines such as monoethanolamine, diethanolamine and triethanolamine. Preferably R ³ is C ₁₀ -C ₁₆ alkyl and R ⁴ is methyl, ethyl or isopropyl. Particularly preferred are methyl ester sulfonates, wherein R ³ is C ₁₄ -C ₁₆ alkyl.

Alkyl sulfate surfactants are another important class of anionic surfactants for use in the present invention. When combined with polyhydroxy fatty acid amides (see below), in addition to excellent all-around cleaning power, including good grease/oil cleaning power over a wide range of temperatures, wash concentrations and wash times, alkyl sulfates are also available Salt solubilization and improved formulatability in liquid detergent formulations, which are water-soluble salts or acids of the formula ROSO ₃ M, wherein R is preferably a C ₁₀ -C ₂₄ hydrocarbyl group, preferably with a C ₁₀ -C ₂₀ alkyl group Part of an alkyl or hydroxyalkyl group, more preferably a _C12 - _C18 alkyl or hydroxyalkyl group, M is H or a cation, such as an alkali metal cation (e.g. sodium, potassium, lithium), a substituted or unsubstituted ammonium cation Examples include methyl, dimethyl, and trimethylammonium and quaternary ammonium cations such as tetramethylammonium and dimethylpyridinium, and those derived from alkanolamines such as ethanolamine, diethanolamine, triethanolamine, and mixtures thereof, etc. . In general for lower wash temperatures (e.g. below about 50°C) alkyl chains of C ₁₂ -C ₁₆ are preferred and for higher wash temperatures (e.g. above about 50°C) C ₁₆ -C ₁₈ alkanes are preferred base chain.

Alkyl alkoxylated sulfate surfactants are another class of suitable anionic surfactants. These surfactants are water soluble salts or acids generally having the formula RO(A) _m SO ₃ M where R is an unsubstituted C ₁₀ -C ₂ 4 alkyl or hydroxyalkane having a C ₁₀ -C ₂₄ alkyl moiety Group, preferably C ₁₂ -C ₂₀ alkyl or hydroxyalkyl, more preferably C ₁₂ -C ₁₈ alkyl or hydroxyalkyl, A is an ethoxy or propoxy unit, m value is greater than 0, generally in the range of about 0.5 to Between about 6, more preferably between about 0.5 and about 3, M is H or a cation, which can be, for example, a metal cation (eg, sodium, potassium, lithium, calcium, magnesium, etc.), ammonium or substituted ammonium cations. Alkyl ethoxylated sulfates as well as alkyl propoxylated sulfates are contemplated herein. Specific examples of substituted ammonium cations include methyl, dimethyl, trimethylammonium cations and quaternary ammonium cations such as tetramethylammonium and dimethylpyridinium, and those formed from alkanolamines such as monoethanolamine, diethanolamine, triethanolamine, Cations derived from ethanolamine and mixtures thereof. Exemplary surfactants are C ₁₂ -C ₁₈ alkyl polyethoxylate (1.0) sulfate, C ₁₂ -C ₁₈ alkyl polyethoxylate (2.25) sulfate, C ₁₂ -C ₁₈ alkyl Polyethoxylate (3.0) sulfate and C ₁₂ -C ₁₈ alkyl polyethoxylate (4.0) sulfate, wherein M is conveniently selected from sodium and potassium.

Other Anionic Surfactants - Other anionic surfactants useful for detersive purposes may also be included in the compositions of the present invention. These may include salts of soaps (including, for example, sodium, potassium, ammonium and substituted ammonium salts such as mono-, di- and triethanolamine salts), C ₉ -C ₂₀ linear alkylbenzene sulfonates, C ₈ -C ₂₂ primary or Secondary alkane sulfonates, C ₈ -C ₂₄ olefin sulfonates, sulfonated polycarboxylic acids prepared by sulfonating pyrolysis products of alkaline earth metal citrates, such as described in British Patent Specification No. 1,082,179 ; Alkyl glycerol sulfonates, fatty acyl glycerol sulfonates, fatty oleoyl glycerol sulfates, alkylphenol oxirane ether sulfates, paraffin sulfonates, alkyl phosphates, isethionates such as Acyl isethionates, N-acyl taurates, fatty acid amides of methyl taurine, alkyl succinamidates and sulfosuccinates, sulfosuccinic acid monoesters (especially saturated and unsaturated saturated C ₁₂ -C ₁₈ monoesters) and sulfosuccinic acid diesters (especially saturated and unsaturated C ₆ -C ₁₄ diesters), N-acyl sarcosinates, sulfates of alkyl polysaccharides such as alkyl Polyglucoside sulfates (nonionic, nonsulfated compounds, described below), branched primary alkyl sulfates, and alkyl polyethoxy carboxylates such as those having the formula RO( _{CH2CH 2} O) salts of _k CH ₂ COO ^- M ⁺ , wherein R is an alkyl group of C ₈ -C ₂₂ , k is an integer from 0 to 10, M is a cation that forms a soluble salt and is esterified with isethionate and replaced with hydrogen Sodium oxide neutralized fatty acids. Resin acids and hydrogenated resin acids are also suitable, such as rosin, hydrogenated rosin, and resin acids and hydrogenated resin acids present in or derived from tall oil. Further examples are given in "Surface Active Agents and Detergents" (Volumes I and II, edited by Schwartz, Perry and Berch). Many such surfactants are also generally disclosed in US Patent 3,929,678, issued December 30, 1975 to Laughlin et al., column 23, line 58 through column 29, line 23 (incorporated herein by reference).

Nonionic Detergent Surfactants - Suitable nonionic detergent surfactants are generally disclosed in U.S. Patent 3,929,678, Laughlin et al., issued December 30, 1975, column 13, line 14 to column 16, line 6, at Text cited by reference. Illustrative, non-limiting types of nonionic surfactants that can be used are listed below.

Polyethylene oxide, polypropylene oxide and polybutylene oxide condensation products of alkylphenols. In general, condensation products of polyethylene oxide are preferred. These compounds include the condensation products of alkylphenols with alkylene oxides having an alkyl group having from about 6 to about 12 carbon atoms in a linear or branched configuration. In a preferred embodiment, the ethylene oxide is present in an amount equal to about 5 to about 25 moles per mole of alkylphenol. Commercially available nonionic surfactants of this type include ^Igepal_CO -630, sold by GAF Corporation; and ^Triton_X -45, X-114, X-100 and X-102, all of which are Sold by Rohm & Haas. These compounds are commonly referred to as alkylphenol alkoxylates (eg, alkylphenol ethoxylates).

Condensation products of fatty alcohols with from about 1 to about 25 moles of ethylene oxide. The alkyl chains of the fatty alcohols can be straight or branched, primary or secondary, and generally contain from about 8 to about 22 carbon atoms. Particularly preferred are condensation products of alcohols having alkyl groups having from about 10 to about 20 carbon atoms with from about 2 to about 18 moles of ethylene oxide per mole of alcohol. Examples of commercially available nonionic surfactants of this type include Tergitol ^_ 15-S-9 (condensation product of C ₁₁ -C ₁₅ linear secondary alcohols with 9 moles of ethylene oxide), Tergitol ^_ 24 -L-6 NMW (condensation product of a _C12 - _C14 primary alcohol with 6 moles of ethylene oxide and has a narrow molecular weight distribution), both sold by Union Carbide Corporation; ^Neodol_45 sold by Shell Chemical Company -9 (condensation product of C ₁₄ -C ₁₅ straight chain alcohol and 9 moles of ethylene oxide), ^{Neodol_23-6.5} (condensation product of C ₁₂ -C ₁₃ straight chain alcohol and 6.5 moles of ethylene oxide), Neodol ^_ 45-7 (condensation product of C ₁₄ -C ₁₅ linear alcohol with 7 moles of ethylene oxide), Neodol ^_ 45-4 (condensation product of C ₁₄ -C ₁₅ linear alcohol with 4 moles of ethylene oxide) ; Kyro ^® EOB (condensation product of a C ₁₃ -C ₁₅ alcohol with 9 moles of ethylene oxide) sold by the Procter & Gamble Company. Such nonionic surfactants are generally referred to as "alkyl ethoxylates".

The condensation product of ethylene oxide with a hydrophobic base formed by the condensation of propylene oxide with propylene glycol. The hydrophobic portion of these compounds preferably has a molecular weight of from about 1500 to about 1800 and exhibits water insolubility. Addition of a polyoxyethylene moiety to the hydrophobic moiety tends to increase the water solubility of the overall molecule, and the liquid character of the product is maintained to the point where the polyoxyethylene content is about 50% by weight of the total condensation product, which is equivalent to Up to about 40 moles of ethylene oxide condense. Examples of compounds of this type include certain commercially available ^Pluronic® surfactants, which are sold by BASF.

The condensation product of ethylene oxide with the product resulting from the reaction of propylene oxide and ethylenediamine. The hydrophobic portion of these products consists of the reaction product of ethylenediamine with excess propylene oxide and generally has a molecular weight of from about 2500 to about 3000. The hydrophobic moiety is condensed with ethylene oxide to such an extent that the condensation product contains from about 40% to about 80% by weight polyoxyethylene and has a molecular weight of from about 5,000 to about 11,000. Examples of this type of nonionic surfactant include some of the commercially available ^Tetronic® compounds, sold by BASF.

Semi-polar nonionic surfactants are a special class of nonionic surfactants that include water-soluble amine oxides comprising an alkyl moiety of about 10 to about 18 carbon atoms and two alkyl moieties selected from the group consisting of about 1 to Alkyl and hydroxyalkyl moieties of about 3 carbon atoms; water-soluble phosphine oxides comprising an alkyl moiety of about 10 to about 18 carbon atoms and 2 and hydroxyalkyl moieties; and water-soluble sulfoxides comprising an alkyl moiety of about 10 to about 18 carbon atoms and an alkyl and hydroxyalkyl moiety selected from about 1 to about 3 carbon atoms part.

Semi-polar nonionic detergent surfactants include amine oxide surfactants having the formula:

R ³ (OR ⁴ ) _x N(O)(R ⁵ ) ₂ wherein R ³ is alkyl, hydroxyalkyl or alkylphenyl containing about 8 to about 22 carbon atoms, or a mixture thereof; R ⁴ is Alkylene or hydroxyalkylene containing about 2 to about 3 carbon atoms or mixtures thereof; x is 0 to about 3; each R is an alkyl or ^{hydroxyalkane} containing about 1 to about 3 carbon atoms groups or polyethylene oxide groups containing from about 1 to about 3 ethylene oxide groups. The ^R groups may be linked to each other, for example via an oxygen or nitrogen atom to form a ring structure.

These amine oxide surfactants include in particular C ₁₀ -C ₁₈ alkyl dimethyl amine oxides and C ₈ -C ₁₂ alkoxyethyl dihydroxyethyl amine oxides.

Alkyl polysaccharides disclosed in U.S. Patent 4,565,647 issued to Llenado on January 21, 1986, having a hydrophobic group of about 6 to about 30 carbon atoms, preferably about 10 to about 16 carbon atoms, and a polysaccharide For example, the hydrophilic group of a polyglycoside contains from about 1.3 to about 10, preferably from about 1.3 to about 3, most preferably from about 1.3 to about 2.7 sugar units. Any reducing sugar containing 5 or 6 carbon atoms may be used, eg glucose, galactose, and glycosyl moieties may be substituted for galactosyl moieties. (The hydrophobic group is optionally attached at the 2-, 3-, 4-, etc. positions, resulting in glucose or galactose as opposed to glucoside or galactoside). An intersugar linkage may be, for example, between the 2-, 3-, 4- and/or 6-position of a previous saccharide unit and a position of another saccharide unit.

Optionally but less preferably polyoxyalkylene chains may link the hydrophobic moiety and the polysaccharide moiety. The preferred alkylene oxide is ethylene oxide. Typical hydrophobic groups include saturated or unsaturated branched or unbranched alkyl groups containing from about 8 to about 18, preferably from about 10 to about 16, carbon atoms. Preferably, the alkyl group is a straight chain saturated alkyl group. The alkyl group may contain up to about 3 hydroxyl groups and/or the polyoxyalkylene chain may contain up to about 10, preferably less than 5, oxyalkylene moieties. Suitable alkyl polysaccharides are octyl, nonyl, decyl, undecyl, dodecyl, tridecyl, tetradecyl, pentadecyl, hexadecyl, heptadecyl and Octadecyl, di-, tri-, tetra-, penta- and hexa-glucosides, galactoside, lactoside, glucose, fructoside, fructose and/or galactose. Suitable mixtures include coconut alkyl, di, tri, tetra and pentaglucosides and tallow alkyl tetra, penta and hexaglucosides.

Preferred alkyl polyglycosides have the formula:

R ² O(C _n H _2n O) _t (glycosyl) _x wherein R ² is selected from the group consisting of alkyl, alkylphenyl, hydroxyalkyl, hydroxyalkylphenyl and mixtures thereof, wherein the alkyl group contains from about 10 to about 18 carbon atoms, preferably about 12 to about 14 carbon atoms; n is 2 or 3, preferably 2; t is 0 to about 10, preferably 0; x is about 1.3 to about 10, preferably about 1.3 to about 3, most Preferably from about 1.3 to about 2.7. The glycosyl is preferably derived from glucose. To prepare these compounds, the alcohol or alkyl polyethoxy alcohol is first prepared and then reacted with glucose or a source of glucose to form the glucoside (attachment at the 1 position). The further glycosyl unit may then be linked between its 1-position and the 2-, 3-, 4- and/or 6-position of the previous glycosyl unit, preferably mainly between the 2-position.

Fatty acid amides having the formula:

R ⁶ -C(O)-N(R ⁷ ) ₂

wherein ^R is an alkyl group containing about 7 to about 21 (preferably about 9 to about 17) carbon atoms, and ^each R is selected from hydrogen, C ₁ -C ₄ alkyl, C ₁ -C ₄ hydroxyalkyl and -( _C2H4O ) _XH , wherein x is from about 1 _to about 3.

Preferred amides are C ₈ -C ₂₀ aminoamides, monoethanolamides, diethanolamides and isopropanolamides.

Cationic Surfactants - Cationic detersive surfactants can also be included in the detergent compositions of the present invention. Cationic surfactants include ammonium salt surfactants such as alkyl dimethyl ammonium halides, and those of the formula:

[R ² (OR ³ ) _y ][R ⁴ (OR ³ ) _y ] ₂ R ⁵ N ⁺ X ^- wherein R ² is an alkyl or alkyl benzyl having about 8 to about 18 carbon atoms in the alkyl chain Each R ³ is selected from the group consisting of -CH ₂ CH ₂ -, -CH ₂ CH(CH ₃ )-, -CH ₂ CH(CH ₂ OH)-, -CH ₂ CH ₂ CH ₂ - and mixtures thereof; each R ⁴ is selected from C ₁ -C ₄ alkyl, C ₁ -C ₄ hydroxyalkyl, benzyl, a ring structure formed by linking two R ⁴ groups, -CH ₂ CHOHCHOHCOR ⁶ CHOHCH ₂ OH, wherein R ⁶ is Any hexose or hexose polymer having a molecular weight of less than about 1000, and when y is other than 0 ^R6 is hydrogen; ^R5 is the same group as ^R4 , or an alkyl chain, wherein ^R2 is added to ^R5 The total number of carbon atoms is not more than about 18; each y is from 0 to about 10 and the sum of the y values is from 0 to about 15; x is any compatible anion.

Other cationic surfactants suitable for use in the present invention are also described in US Patent 4,228,044, Cambre, issued October 14, 1980, which is incorporated herein by reference.

Other Surfactants - Amphoteric surfactants can be incorporated into the detergent compositions of the present invention. These surfactants can be broadly described as aliphatic derivatives of secondary or tertiary amines, or aliphatic derivatives of heterocyclic secondary and tertiary amines, where the aliphatic group may be straight or branched. One of the aliphatic substituents contains at least about 8 carbon atoms, typically from about 8 to about 18 carbon atoms, and at least one of the aliphatic substituents contains a water-solubilizing anionic group, such as carboxyl, sulfonate, sulfate . See US Patent 3,929,678, issued December 30, 1975 to Laughlin et al., column 19, lines 18-35, for examples of amphoteric surfactants.

Zwitterionic surfactants can also be incorporated into the detergent compositions herein. These surfactants can be broadly described as derivatives of secondary and tertiary amines, derivatives of heterocyclic secondary and tertiary amines, or derivatives of quaternary ammonium salts, quaternary phosphonium or tertiary sulfonium compounds. See US Patent 3,929,678, issued December 30, 1975 to Laughlin et al., column 19, line 38 through column 22, line 48, for examples of zwitterionic surfactants. Amphoteric and zwitterionic surfactants are generally used in combination with one or more anionic and/or nonionic surfactants.

Polyhydroxy Fatty Acid Amide Surfactant - The liquid detergent compositions of the present invention may also contain an enzyme enhancing amount of a polyhydroxy fatty acid amide surfactant. By "enhancing enzyme amount" is meant that the formulator of the composition can select the amount of polyhydroxy fatty acid amide incorporated into the composition such that the cleaning performance of the enzyme in the detergent composition is improved. Typically, about 1% by weight of polyhydroxy fatty acid amide is incorporated to enhance enzyme performance for conventionally used amounts of enzyme.

The detergent compositions herein will generally comprise about 1% by weight polyhydroxy fatty acid amide surfactant, preferably from about 3% to about 30% polyhydroxy fatty acid amide. The polyhydroxy fatty acid amide surfactant component includes compounds of the following structural formula:

R ² -C(O)-N(R ¹ )-Z wherein: R ¹ is hydrogen, C ₁ -C ₄ hydrocarbyl, 2-hydroxyethyl, 2-hydroxypropyl, or a mixture thereof, preferably C ₁ -C ₄ alkyl, more preferably C ₁ or C ₂ alkyl, most preferably C ₁ alkyl (ie methyl); R ² is C ₅ -C ₃₁ hydrocarbon group, preferably straight chain C ₇ -C ₁₉ alkyl or alkenyl , more preferably straight chain C ₉ -C ₁₇ alkyl or alkenyl, most preferably straight chain C ₁₁ -C ₁₅ alkyl or alkenyl or mixtures thereof; and Z is a straight chain having at least 3 hydroxyl groups directly attached Polyhydroxyhydrocarbyl groups of hydrocarbyl chains, or alkoxylated derivatives thereof (preferably ethoxylated or propoxylated). Z is preferably derived from a reducing sugar in a reductive amination reaction; more preferably Z is a glycityl. Suitable reducing sugars include glucose, fructose, maltose, lactose, galactose, mannose and xylose. As raw materials, high dextrose corn syrup, high fructose corn syrup, and high maltose corn syrup may be used in addition to the individual sugars listed above. The mixed sugar components for Z are derived from these corn syrups. It should be recognized that it is by no means intended to exclude other suitable starting materials. Z is preferably selected from _-CH2 (CHOH) _n - _CH2OH , -CH( _CH2OH )(CHOH _{) n-1- CH2OH} , -CH2-(CHOH) ₂ (CHOR')(CHOH)- _CH2OH , and alkoxylated derivatives thereof, wherein n is an integer from 3 to 5 inclusive, and R' is H or a cyclic or aliphatic monosaccharide. Most preferred are glycityls where n is 4, specifically _-CH2- (CHOH) ₄ - _CH2OH .

R ¹ can be, for example, N-methyl, N-ethyl, N-propyl, N-isopropyl, N-butyl, N-2-hydroxyethyl or N-2-hydroxypropyl.

R ² -CO-N< can be, for example, cocoamide, stearamide, oleamide, laurylamide, myristamide, capricamide, palmitamide, tallowamide, and the like.

Z can be 1-deoxyglucosyl, 2-deoxyfructosyl, 1-deoxymaltosyl, 1-deoxylactosyl, 1-deoxygalactosyl, 1-deoxymannosyl, 1-deoxymaltotriosyl, etc. .

Methods of preparing polyhydroxy fatty acid amides are well known in the art. In general, they are prepared by reacting an alkylamine with a reducing sugar in a reductive amination reaction to form the corresponding N-alkyl polyhydroxylamine, followed by the condensation/amidation step of the N-alkyl Polyhydroxylamines react with fatty esters or triglycerides to form N-alkyl, N-polyhydroxy fatty acid amide products. Methods for preparing compositions containing polyhydroxy fatty acid amides are disclosed, for example, in British Patent Specification 809060, Thomas Hedley & Co., Ltd., issued February 18, 1959; , and U.S. Patent 2,703,798 to Anthony M. Schwartz, issued March 8, 1955, and U.S. Patent 1,985,424 to Piggott, issued December 25, 1934, which are all incorporated herein by reference.

Second Enzyme - Preferred compositions of the present invention also comprise a performance enhancing amount of a second detergent compatible enzyme. "Detergent compatible" means compatible with other ingredients in liquid detergent compositions such as detersive surfactants and detergency builders. These second enzymes are preferably selected from lipases, amylases, cellulases and mixtures thereof. The term "second enzyme" excludes the proteases discussed above, so each composition with a second enzyme contains at least two enzymes, including at least one protease. The amount of the second enzyme used in the composition varies according to the type of enzyme. Generally, it is preferred to use about 0.0001-0.3% by weight, more preferably 0.001-0.1% by weight of these second enzymes. Mixtures of enzymes of the same type (eg lipase) or mixtures of two or more types (eg cellulase and lipase) may also be used. Enzymes can be used in refined or unrefined form.

Any lipolytic enzyme suitable for use in liquid detergent compositions can be used in these compositions. Lipases suitable for use in the present invention include those obtained from bacterial and fungal sources.

Suitable bacterial lipases include those produced by microorganisms of the Pseudomonas group, such as Pseudomonas stutzeri ATCC 19.154, as disclosed in British Patent 1372034, incorporated herein by reference as refer to. Suitable lipases include those that exhibit a positive immunological cross-reaction with antibodies to lipase produced by the microorganism Pseudomonas fluorescens IAM1057. This lipase and its purification method are described in Japanese Patent Application No. 53-20487 published on February 24, 1978 . This lipase is commercially available from Amano Pharmaceutical Co. Ltd., Nagoya, Japan under the tradename Lipase P "Amano", hereinafter referred to as "Amano-P". This lipase should show a positive immunological cross-reaction with the Amano-P antibody using the standard and known immunodiffusion method according to Quchterlony (Acta, Med. Scan., 133, pp. 76-79 (1950)). These lipases and their method of immunological cross-reactivity with Amano-P are also described in US Patent 4,707,291, Thom et al., issued November 17, 1987, which is incorporated herein by reference. Typical examples thereof are Amano-P lipase, lipase from Pseudomonas fragi FERM P1339 (commercially available under the tradename Amano-B), lipolyticum FERM P1338 from Pseudomonas azoreducens variant (available under the tradename Amano-CES), lipases from Chromobacter viscosum such as Chromobacter viscosum var. lipolyticum NRRLB 3673, which are commercially available from Toyo Jozo Co., Tagata, Japan, and additionally from the U.S. Biochemical Corporation, USA and Disoynth Co., Netherlands purchased Chromobacter viscosum lipase and lipase from Pseudomonas gladioli.

Suitable fungal lipases include those produced by Humicola lanuginosa and Thermomyces lanuginosus . Most preferred is the lipase obtained by cloning the gene from Humicola lanuginosa and expressing the gene in Aspergillus oryzae, as described in European Patent Application 0258068 (Novo Industri A/S), commercially available under the trade name ^Lipolase_ was purchased from Novo Nordisk A/S.

From about 10 to 18,000, preferably from about 60 to 6,000 lipase units per gram of lipase (LU/g) may be used in these compositions. Lipase units are at a fixed pH, i.e. pH 9.0, temperature 30°C, matrix is an emulsion of 3.3% by weight olive oil and 3.3% gum arabic, 13mmol in 5mmol/l Tris buffer The amount of lipase that produces 1 mmol titratable fatty acid per minute at Ca ⁺⁺ /l Ca++ and 20 mmol/l NaCl.

Any cellulase enzyme suitable for use in liquid detergent compositions can be used in these compositions. Suitable cellulases for use in the present invention include those derived from bacterial and fungal sources. Preferably their pH optimum is between 5-9.5. About 0.0001-0.1% by weight cellulase can be used.

Suitable cellulases are disclosed in US Patent No. 4,435,307, Barbesgoard et al., issued March 6, 1984, incorporated herein by reference, which discloses fungal cellulases produced by Humicola insolens. Suitable cellulases are also disclosed in GB-A-2075028; GB-A-2095275 and DE-OS-2247832.

Examples of such cellulases are cellulases produced by strains of Humicola insolens (Humicola griseus var. thermoidea ), in particular Humicola strain DSM1800, and cellulases produced by Bacillus N molds or produced by bacteria belonging to the Cellulase 212 produced by molds of the genus Aeromonas, and cellulase extracted from the hepatopancreas of marine molluscs (Dolabella Auricula Solander).

Any amylase suitable for use in liquid detergent compositions can be used in these compositions of the present invention. Amylases include, for example, amylases obtained from a special strain of Bacillus licheniformis, described in more detail in British Patent Specification 1296839 (Novo). Amylolytic proteins include, for example, Rapidase ^R , International Bio-Synthetics, Inc., and Termamyl ^R Novo Industries.

About 0.0001%-0.55%, preferably 0.0005-0.1% by weight of amylase may be used.

Calcium - The compositions of the present invention may optionally contain a source of calcium ions. Any water-soluble calcium salt can be used as the source of calcium ions, including calcium acetate, calcium formate, calcium xylene sulfonate, and calcium propionate. Divalent ions such as zinc and magnesium ions can replace calcium ions completely or partially. Thus, in the liquid detergent compositions of the present invention, the source of calcium ions may be partially replaced by another source of divalent ions.

The calcium used in the present invention is enzyme accessible. Accordingly, preferred compositions are substantially free of chelating agents, such as polyacids capable of forming calcium complexes which are soluble in the composition. However, small amounts of chelating agents such as polyacids or mixtures of polyacids may be used. Enzyme accessible calcium is defined as the amount of calcium ions available to the enzyme components. Thus, from a practical point of view, in the absence of any depot chelating agent, the calcium accessible to the enzyme is soluble calcium in the composition, eg, the equilibrium constant of the calcium complex is equal to or greater than 1.5 at 20°C.

Boronic Acid - The compositions of the present invention also optionally contain from about 0.25% to about 10%, preferably from about 0.5% to about 5%, more preferably from about 0.75% to about 3% by weight of boric acid or a compound capable of forming boric acid in the composition ( Calculated on the basis of boric acid). Boric acid is preferred, although other compounds such as boron oxide, borax, and other alkali metal borates (eg, sodium ortho-, meta-, pyroborate, sodium pentaborate) are also suitable. Substituted boronic acids (eg, phenylboronic acid, butaneboronic acid, and p-bromophenylboronic acid) can also be used in place of boric acid.

The compositions of the invention may also contain polyols, especially diols, containing only carbon, hydrogen and oxygen atoms. They preferably contain from about 2 to about 6 hydroxyl groups. Examples include propylene glycol (especially 1,2-propanediol, which is preferred), ethylene glycol, glycerin, sorbitol, mannitol, glucose, and mixtures thereof. Polyols generally comprise from about 1% to about 15%, preferably from about 1.5% to about 10%, more preferably from about 2% to about 7%, by weight of the composition.

Optional Components - Detergent builders can optionally be included in the compositions herein, especially for laundry compositions. Inorganic as well as organic builders can be used. When present, the composition will generally contain at least about 1% builder, and this may be inorganic or organic. Liquid laundry formulations preferably comprise from about 3% to 30%, more preferably from about 5% to 20%, by weight, of detergency builder.

Inorganic detergency builders include, but are not limited to, the alkali metal, ammonium, and alkanolammonium salts of the following: polyphosphates (such as tripolyphosphates, pyrophosphates, and glassy polymeric metaphosphates), phosphonates, creatine Alcohol hexaphosphoric acid, silicates, carbonates (including bicarbonates and sesquicarbonates), sulfates and aluminosilicates. Borate builders and builders containing borate-forming materials which generate borate under detergent storage or wash conditions (hereinafter collectively referred to as "borate builders") can also be used. Non-borate builders are preferred for use in compositions herein intended for use at wash conditions below about 50°C, especially below about 40°C.

Examples of silicate builders are alkali metal silicates, especially those having a _SiO2 : _Na2O ratio in the range of 1.6:1 to 3.2:1 and layered silicates such as Layered sodium silicates are described in US Patent No. 4,664,839, HP Rieck, issued May 12, 1987, which is incorporated herein by reference. But other silicates are also useful, such as magnesium silicate, which can be used as a crisping agent in granular formulations, as a stabilizer for oxygen bleaches and as a component of foam control systems.

Examples of carbonate builders are the alkaline earth and alkali metal carbonates in German Patent Application 2321001 published on November 15, 1973 (the disclosure of which is incorporated herein by reference), including sodium carbonate and sesqui Sodium carbonates and their mixtures with superfine calcium carbonate.

Aluminosilicate builders are useful herein. Aluminosilicate builders are of paramount importance in the most popular commercially available heavy duty granular detergent compositions and can also be an important builder ingredient in liquid detergent formulations. Aluminosilicate builders include builders having the following empirical formula:

M _Z (zAlO ₂ .ySiO ₂ ) where M is sodium, potassium, ammonium, or substituted ammonium, z is about 0.5 to about 2; y is 1; the magnesium ion exchange capacity of the material is per gram of anhydrous aluminosilicate Hardness of at least about 50 meq CaCO ₃ . Preferred aluminosilicates are zeolite builders having the formula:

Na _z [(AlO ₂ ) _Z (SiO ₂ ) _y ].xH ₂ O wherein z and y are integers of at least 6, the molar ratio of z to y is in the range of 1.0 to about 0.5, and x is from about 15 to about Integer of 264.

Useful aluminosilicate ion exchange materials are commercially available. These aluminosilicates may be of crystalline or amorphous structure and may be naturally occurring aluminosilicates or synthetically derived. Methods for preparing aluminosilicate ion exchange materials are disclosed in US Patent 3,985,669, Krummel et al., issued October 12, 1976, incorporated herein by reference. Preferred synthetic crystalline aluminosilicate ion exchange materials for use herein are commercially available under the registered trademarks Zeolite A, Zeolite P(B) and Zeolite X. In a particularly preferred embodiment, the crystalline aluminosilicate ion exchange material has the formula:

Na ₁₂ [(AlO ₂ ) ₁₂ (SiO ₂ ) ₁₂ ].xH ₂ O wherein x is about 20 to about 30, especially about 27. This material is called Zeolite A. The aluminosilicate preferably has a particle size of about 0.1-10 microns in diameter.

Specific examples of polyphosphate salts are alkali metal tripolyphosphate, sodium, potassium and ammonium pyrophosphate, sodium, potassium and ammonium pyrophosphate, sodium and potassium orthophosphate, sodium polymeta/phosphate, wherein the degree of polymerization is about 6 - about 21, and salts of phytic acid.

Examples of phosphonate builder salts are the water-soluble salts of ethane 1-hydroxy-1,1-diphosphonic acid, especially the sodium and potassium salts, the water-soluble salts of methylene diphosphonic acid such as the trisodium salt and Tripotassium salt, and water-soluble salts of substituted methylene diphosphonic acids, such as trisodium and tripotassium ethylene, isopropylidene, benzylmethylene, and halomethylene phosphonates. Phosphonate builder salts of the type described above are disclosed in U.S. Patents 3,159,581 and 3,213,030, issued December 1, 1964, and October 19, 1965 to Diehl; US Patents 3,400,148 and 3,422,137, issued September 3, 1969 and January 14, 1969 to Quimby, the disclosures of which are incorporated herein by reference.

Preferred organic detergent builders for the purposes of the present invention include various polycarboxylate compounds. As used herein, "polycarboxylate" means a compound having a plurality of carboxylate groups, preferably at least 3 carboxylate groups.

Polycarboxylate builders can generally be incorporated into the compositions in acid form, but can also be included in neutralized salt form. When used in salt form, alkali metals such as sodium, potassium and lithium or alkanolammonium salts are preferred.

Included in polycarboxylate builders are various types of useful materials. One important class of polycarboxylate builders comprises the ether polycarboxylates. A number of ether polycarboxylates have been disclosed for use as detergent builders. Examples of useful ether polycarboxylates include oxydisuccinates as disclosed in U.S. Patent 3,128,287, Berg, issued April 7, 1964, and U.S. Patent 3,635,830, Lamberti et al., issued January 18, 1972, Both documents are incorporated herein by reference.

Particular types of ether polycarboxylates useful as builders herein include those having the general formula:

CH(A)(COOX)-CH(COOX)-O-CH(COOX)-CH(COOX)(B) where A is H or OH; B is H or -O-CH(COOX)-CH ₂ (COOX ); X is H or a salt-forming cation. For example, in the general formula above, if A and B are both H, then the compound is oxydisuccinic acid and its water-soluble salts. If A is OH and B is H, then the compound is tartrate monosuccinic acid (TMS) and its water-soluble salts. If A is H and B is -O-CH(COOX) _-CH2 (COOX), then the compound is tartrate disuccinic acid (TDS) and its water-soluble salts. Mixtures of these builders are especially preferred for use herein. Particularly preferred are mixtures of TMS and TDS in a weight ratio TMS:TDS of from about 97:3 to about 20:80. These builders are disclosed in US Patent 4,663,071, Bush et al., issued May 5,1987.

Suitable ether polycarboxylates also include cyclic compounds, especially cycloaliphatic compounds, such as those described in US Patent Nos. 3,923,679; 3,835,163;

Other useful builders include ether hydroxy polycarboxylates represented by the formula:

HO-[C(R)(COOM)-C(R)(COOM)-O] _n -H wherein M is hydrogen or a cation such that the resulting salt is water-soluble, preferably an alkali metal, ammonium or substituted ammonium cation, n is about 2 to about 15 (preferably n is about 2 to about 10, more preferably n averages about 2 to about 4), each R is the same or different, and is selected from hydrogen, C _1-4 alkyl or C _1-4 substituted alkyl (preferably R is hydrogen).

Other ether polycarboxylates include copolymers of maleic anhydride and ethylene or vinyl methyl ether, 1,3,5-trihydroxybenzene-2,4,6-trisulfonic acid, and carboxymethoxysuccinic acid .

Organic polycarboxylate builders also include the various alkali metal, ammonium and substituted ammonium polyacetic acids. Examples include ethylenediaminetetraacetic acid and sodium, potassium, lithium, ammonium and substituted ammonium salts of nitrilotriacetate.

Also included are polycarboxylates such as mellitic acid, succinic acid, oxydisuccinic acid, polymaleic acid, benzene-1,3,5-tricarboxylic acid, carboxymethoxysuccinic acid and soluble salts thereof.

Citrate builders, such as citric acid and its soluble salts (especially the sodium salt), are polycarboxylate builders of particular importance in heavy duty liquid detergent formulations, but can also be included in granular compositions.

Other carboxylate builders include the carboxylated carbohydrates disclosed in Diehl, US Patent 3,723,322, issued March 28, 1973, incorporated herein by reference.

Also suitable for use in the detergent compositions of the present invention are the 3,3-dicarboxy-4-oxa-1,6-hexanedioates disclosed in U.S. Patent No. 4,566,984, Bush, issued January 28, 1986 and For the compounds concerned, this document is incorporated herein by reference. Useful succinic acid builders include _C5 - _C20 alkyl succinic acids and their salts. A particularly preferred compound of this type is dodecenylsuccinic acid. Alkyl succinic acid generally has the general formula: R—CH(COOH)CH ₂ (COOH), that is, a derivative of succinic acid, wherein R is a hydrocarbon group, such as C ₁₀ -C ₂₀ alkyl or alkenyl, preferably C ₁₂ - C ₁₆ alkyl or alkenyl, or where R may be substituted by hydroxy, sulfo, sulfoxy, sulfone substituents, as described in the aforementioned patents.

Succinate builders are preferably used in the form of their water-soluble salts, including sodium, potassium, ammonium and alkanolammonium salts.

Specific examples of succinate builders include: lauryl succinate, myristyl succinate, palmityl succinate, 2-dodecenyl succinate (preferred), 2-pentadecanyl succinate alkenyl succinate, etc. Lauryl succinate is a preferred builder of this group and is described in European Patent Application 86200690.5/0200263, published November 5,1986.

Examples of useful builders also include carboxymethyloxymalonic acid, carboxymethyloxysuccinic acid, cis-cyclohexanehexacarboxylic acid, sodium and potassium cis-cyclopentane tetracarboxylate; water-soluble polyacrylates (These polyacrylates having a molecular weight above about 2000 are also effective as dispersants), and copolymers of maleic anhydride and vinyl methyl ether or ethylene.

Other suitable polycarboxylates are the polyacetal carboxylates disclosed in US Patent 4,144,226, Crutchfield et al., issued March 13, 1979, which is incorporated herein by reference. These polyacetal carboxylates can be prepared by bringing together under polymerization conditions a glyoxylic acid ester and a polymerization initiator. The resulting polyacetal carboxylate is then attached with chemically stable end groups to stabilize the polyacetal carboxylate against rapid depolymerization in alkaline solution, converted into the corresponding salt, and added to the surfactant.

Polycarboxylate builders are also disclosed in US Patent 3,308,067, Diehl, issued March 7, 1967, which is incorporated herein by reference. Such materials include the water-soluble salts of homo- and copolymers of aliphatic carboxylic acids such as maleic acid, itaconic acid and methylenemalonic acid.

Other organic builders known in the art can also be used. For example, monocarboxylic acids having long chain hydrocarbyl groups and soluble salts thereof can be used. These include substances commonly known as "soaps". Typically chain lengths C ₁₀ -C ₂₀ are used. The hydrocarbyl group may be saturated or unsaturated.

Other optional ingredients include soil release agents, sequestrants, clay soil removal/antiredeposition agents, polymeric dispersants, bleaches, brighteners, suds suppressors, solvents and aesthetic agents.

The detergent compositions of the present invention can be formulated in a variety of compositions, eg as laundry detergents and hard surface cleaners or dishwashing compositions.

The compositions according to the invention are further illustrated by the following examples.

Example I

Prepare the following compositions by mixing the listed ingredients in the listed proportions: combination A B C D. E. f linear alkylbenzene sulfonic acid 8.5 15 6.5 10 12.5 4 Sodium C _12-15 Alkyl Sulfate 1 2 1 2 - - C _14-15 Alkyl 2.5x Ethoxylated Sulfate 10 5 10.5 - 11 9 C ₁₂ Glucamide - - 9 - - 5 C _12-15 Alcohol 7x Ethoxylates 3 10 4 7 2.5 - fatty acid 2 5 5 4 2 2 citric acid 6 7 4 6 4 5 C _12-14 alkenyl substituted succinic acid - 6 - 5 - 6 sodium hydroxide 2 6 2 4 1 1.5 ethanol 2 1.5 2 4 2 1.5 Monoethanolamine 6 5 4 - - - 1,2-propanediol 12 10 5 5 4 6 Amylase (143KNU/g) - - 0.1 - - 0.2 ^Lipolase_ (100KLU/g commercial solution) 0.5 0.2 0.5 0.5 0.4 - Protease B (34 g/L commercial solution) 0.9 - 0.5 - 1.2 - ^Savinase_ (commercially available solution) - 0.3 - 0.4 0.2 0.3 ^Carezyme_ 0.5 1 0.8 - 0.2 0.8 Aldehyde Inhibitor ¹ 0.009 0.005 0.001 0.0005 0.0003 0.01 calcium ion 0.01 0.5 0.1 0.05 0.9 0.25 water and minor components Balance to 100% 1) The aldehyde protease inhibitor according to Synthesis Example 1.

Example II

Prepare the following recipe: components weight(%) Alkyl 1.4 Ethoxylated Sulfate 30 Amine oxide 6 polyhydroxy fatty acid amides 4 Nonionic Surfactant (C11E9) 5 Magnesium ions from _MgCl2 1 Calcium ions from _CaCl2 0.2 Aldehyde Inhibitor ^* 0.0025 Sodium xylene sulfonate 4 solvent 6 water to 100% pH to 8 *The aldehyde of Example 1 was synthesized.

Example III

Prepare the following compositions by mixing the listed ingredients in the listed proportions: components A (weight%) B (weight%) C (weight%) D (weight%) LAS 0 0 0 12 AE _X S ¹ 22.1 24.7 33.5 3 polyhydroxy fatty acid amides 4.6 1.2 4.2 0 Amine oxide 4.6 1.2 4.8 0 Betaine 0 1.2 0 0 nonionic surfactant 6.7 4.1 0 0 Mg(OH) ₂ 0.5 0.5 0.7 0 Calcium ions from _CaCl2 0.1 0.3 0.4 0.1 Calcium xylene sulfonate 4.5 0 4 0 polyethylene glycol 3 0 0 0 Polypropylene Glycol 2000 1.5 0 0 0 balance, water to 100% to 100% to 100% to 100% Protease A or Protease B 0.001-0.01 0.001-0.01 0.005-0.01 0.0003-0.01 Aldehyde Inhibitor ² 0.00025-0.0025 0.00025-0.0025 0.00025-0.0025 0.00125-0.0025 ¹ x = degree of ethoxylation. For each composition, the average degree of ethoxylation was A=2.2, B=0.6, C=1.4, D=2.2. ² The aldehyde of Synthesis Example 1 is used here.

Claims (17)

1. A liquid detergent composition comprising: a) by weight of the composition, 1%-95% detersive surfactant; b) active protease; and c) protease inhibitors having the formula: Z-A-NH-CH(R)-C(O)-X wherein A is an amino acid moiety; X is hydrogen; Z is an N-terminated moiety selected from the group consisting of sulfonamides, sulfenamides, sulfonic acids, phosphinamides, amidophosphoramidates, sulfonamide derivatives, and phosphonamides; and R is selected from linear or branched C ₁ -C ₆ unsubstituted alkyl, phenyl and C ₇ -C ₉ alkaryl moieties. 2. A liquid detergent composition according to claim 1, wherein R is selected from the group consisting _of methyl, isopropyl, sec-butyl, _isobutyl , _-C6H5 , _-CH2 - _C6H5 and _{-CH2 CH2} - _{C6H5 .} 3. A liquid detergent composition according to claim 1 comprising: a) 8%-70% of said detersive surfactant; b) 0.0001%-5% active protease; c) 0.00001%-5% protease inhibitors. 4. A liquid detergent composition according to claim 3, further comprising a source of calcium ions. 5. A liquid detergent composition according to claim 4, wherein the N-terminal moiety Z is selected from the group consisting of (R'O) ₂ (O)P-, (SR') _2- , R'(O) _2S- , SO ₃ H, (R') ₂ (O)P-, R'O(O) ₂ S-, R'-P(O)OH and R'O(OH)(O)P-, where each R' independently selected from linear or branched C ₁ -C ₆ unsubstituted alkyl, phenyl, C ₇ -C ₉ alkaryl and cycloalkyl moieties, wherein the cycloalkyl ring may span C ₄ -C ₈ , and One or more heteroatoms selected from O, N and S may be contained. 6. The liquid detergent composition according to claim 1, further comprising a source of calcium ions selected from the group consisting of calcium formate, calcium chloride, calcium acetate, calcium xylene sulfonate, calcium sulfate and mixtures thereof. 7. A liquid detergent composition according to claim 6, wherein said protease is selected from the group consisting of trypsin, subtilisin, chymotrypsin, elastase and mixtures thereof. 8. A liquid detergent composition according to claim 7, wherein said protease is selected from the group consisting of subtilisin BPN, subtilisin BPN', protease A, protease B, protease D and mixtures thereof. 9. A liquid detergent composition according to claim 1, further comprising suds boosters, chelating agents, polyacrylate polymers, dispersants, dyes, perfumes, processing aids or mixtures thereof. 10. A liquid detergent composition according to claim 9, further comprising an amylase. 11. A liquid detergent composition according to claim 10, further comprising 0.25% to 10% by weight of boric acid or a compound capable of forming boric acid and a polyol. 12. A liquid detergent composition according to claim 1, further comprising an effective amount of a second enzyme selected from the group consisting of lipases, amylases, cellulases and mixtures thereof. 13. A liquid detergent composition according to claim 12, further comprising suds boosters, builders, soil release polymers, polyacrylate polymers, dispersants, dye transfer inhibitors, dyes, perfumes, processing aids , whitening agents or their mixtures. 14. A liquid detergent composition according to claim 12, wherein said second enzyme is lipase. 15. A liquid detergent composition according to claim 14, wherein the lipase is obtained by cloning a gene from Humicola Lanuginosa and expressing the gene in Aspergillus oryzae. 16. The composition according to claim 12, wherein said second enzyme is a cellulase produced by Humicola insolens , and wherein said composition comprises 0.0001% to 0.1% of said fiber by total weight of the composition prime enzyme. 17. A liquid detergent composition according to claim 1, further comprising 0.25% to 10% by weight of boric acid or a compound capable of forming boric acid and a polyol.