IL258550A - Cleaning of water filtration membranes - Google Patents
Cleaning of water filtration membranesInfo
- Publication number
- IL258550A IL258550A IL258550A IL25855018A IL258550A IL 258550 A IL258550 A IL 258550A IL 258550 A IL258550 A IL 258550A IL 25855018 A IL25855018 A IL 25855018A IL 258550 A IL258550 A IL 258550A
- Authority
- IL
- Israel
- Prior art keywords
- dnase
- membrane
- cleaning
- seq
- amino acid
- Prior art date
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Classifications
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01D—SEPARATION
- B01D65/00—Accessories or auxiliary operations, in general, for separation processes or apparatus using semi-permeable membranes
- B01D65/02—Membrane cleaning or sterilisation ; Membrane regeneration
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01D—SEPARATION
- B01D61/00—Processes of separation using semi-permeable membranes, e.g. dialysis, osmosis or ultrafiltration; Apparatus, accessories or auxiliary operations specially adapted therefor
- B01D61/14—Ultrafiltration; Microfiltration
- B01D61/18—Apparatus therefor
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01D—SEPARATION
- B01D61/00—Processes of separation using semi-permeable membranes, e.g. dialysis, osmosis or ultrafiltration; Apparatus, accessories or auxiliary operations specially adapted therefor
- B01D61/14—Ultrafiltration; Microfiltration
- B01D61/22—Controlling or regulating
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01D—SEPARATION
- B01D65/00—Accessories or auxiliary operations, in general, for separation processes or apparatus using semi-permeable membranes
- B01D65/02—Membrane cleaning or sterilisation ; Membrane regeneration
- B01D65/022—Membrane sterilisation
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01D—SEPARATION
- B01D65/00—Accessories or auxiliary operations, in general, for separation processes or apparatus using semi-permeable membranes
- B01D65/08—Prevention of membrane fouling or of concentration polarisation
-
- C—CHEMISTRY; METALLURGY
- C11—ANIMAL OR VEGETABLE OILS, FATS, FATTY SUBSTANCES OR WAXES; FATTY ACIDS THEREFROM; DETERGENTS; CANDLES
- C11D—DETERGENT COMPOSITIONS; USE OF SINGLE SUBSTANCES AS DETERGENTS; SOAP OR SOAP-MAKING; RESIN SOAPS; RECOVERY OF GLYCEROL
- C11D3/00—Other compounding ingredients of detergent compositions covered in group C11D1/00
- C11D3/16—Organic compounds
- C11D3/38—Products with no well-defined composition, e.g. natural products
- C11D3/386—Preparations containing enzymes, e.g. protease or amylase
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/14—Hydrolases (3)
- C12N9/16—Hydrolases (3) acting on ester bonds (3.1)
- C12N9/22—Ribonucleases RNAses, DNAses
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01D—SEPARATION
- B01D2321/00—Details relating to membrane cleaning, regeneration, sterilization or to the prevention of fouling
- B01D2321/16—Use of chemical agents
- B01D2321/166—Use of enzymatic agents
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01D—SEPARATION
- B01D2321/00—Details relating to membrane cleaning, regeneration, sterilization or to the prevention of fouling
- B01D2321/18—Use of gases
- B01D2321/185—Aeration
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- C—CHEMISTRY; METALLURGY
- C02—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F2303/00—Specific treatment goals
- C02F2303/16—Regeneration of sorbents, filters
-
- C—CHEMISTRY; METALLURGY
- C02—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F2303/00—Specific treatment goals
- C02F2303/20—Prevention of biofouling
Landscapes
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Life Sciences & Earth Sciences (AREA)
- Water Supply & Treatment (AREA)
- Health & Medical Sciences (AREA)
- Organic Chemistry (AREA)
- Wood Science & Technology (AREA)
- Separation Using Semi-Permeable Membranes (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Molecular Biology (AREA)
- Genetics & Genomics (AREA)
- Zoology (AREA)
- Biotechnology (AREA)
- Microbiology (AREA)
- Biochemistry (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Biomedical Technology (AREA)
- Oil, Petroleum & Natural Gas (AREA)
- Medicinal Chemistry (AREA)
- External Artificial Organs (AREA)
- Purification Treatments By Anaerobic Or Anaerobic And Aerobic Bacteria Or Animals (AREA)
Description
WO 2017/066510 PCT/US2016/056955
CLEANING OF WATER FILTRATION MEMBRANES
FIELD OF THE INVENTION
The present invention relates to cleaning of water filtration membranes using DNase
containing cleaning compositions.
BACKGROUND OF THE INVENTION
Membrane fouling is a problem encountered in membrane filtration processes, and it is a
major factor in determining their practical application in water and wastewater treatment and
desalination in terms of technology and economics. Membrane fouling includes inorganic
fouling/scaling, particulate/colloidal fouling, and organic fouling with dead organic matter or live
microorganisms, which would typically form a biofilm. Fouling due to organic and inorganic
components and microorganisms can occur simultaneously, and these components may
interact in terms of mechanism.
A well-known means for cleaning filtration membranes is “cleaning-in-place” (CIP). A
typical CIP cycle will often include the use of chemicals for high pH (e.g., NaOH), low pH (e.g.,
citric acid or nitric acid), water and biocides for disinfection, e.g., bleach. A typical CIP cycle for
cleaning membranes used in the dairy industry consists of many steps which often include:
- Flush/Pre-rinse with clean water flux, which may be heated up to 60-80°C and recirculated for
a period of time.
- Circulation of caustic soda solution (NaOH) at a temperature about 60-80°C for a period of
time.
- Intermediate rinsing with clean water flux.
- Circulation of acidic solution for a period of time.
- Final rinse with clean water flux.
- Final air blow.
Similar processes exist for membrane cleaning in other business segment, and the
individual steps can be applied in different order dependent on the nature of the membrane
fouling. A typical CIP cycle for cleaning membranes used in the enzyme manufacturing industry
includes water flush, a cold and hot caustic rinse, and nitric acid rinse.
The use of harsh chemicals in CIP is undesirable and poses a problem to the
environment. In the past years CIP methods including the use of enzymes have been
developed.
International patent application WO 97/02753 concerns a solution comprising a protease
and a lipase for cleaning-in-place. The solution has been found effective in cleaing process
equipment containing residues of milk or burnt milk.
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Compared to a typical CIP procedure, there are several benefits of improved membrane
cleaning by use of DNase enzymes, such as, saving power to pump liquid through less fouled
membranes, reducing use of harsh and hazardous chemicals, reducing cost for neutralising
acids and bases and discharge after use, extension of membrane lifetime due to milder cleaning
conditions and removal of fouling not removed by chemical or physical means.
SUMMARY
The present invention provides a method for cleaning a water filtration membrane by
contacting the membrane with a liquid cleaning composition comprising a DNase.
The cleaning process may be Cleaning-In-Place (CIP) or Cleaning-Out-of-Place (COP).
In an embodiment, the water flow rate through the membrane (flux) is improved.
In another embodiment, the method is preceded or followed by contacting the
membrane with a biocidal composition.
In yet another embodiment, the water filtration membrane comprises a biofilm, which
preferably includes one or more bacteria selected from the group consisting of Acinetobacter,
Bacillus, Comamonas, Escherichia, Pseudomonas, and Sphingomonas species.
Other aspects and embodiments of the invention will be apparent from the description
and examples.
DETAILED DESCRIPTION OF THE INVENTION
In the context of the present invention, the terms “water filtration” and “water treatment”
include fitration and treatment of aqueous solutions and aqueous suspensions, as well as
substantially pure water.
It has been shown that extracellular DNA plays an important role in initiating attachment
of biotic and abiotic organic matter to membranes. Extracellular DNA (eDNA) has been released
to the environment from dead cells of animal, plant or microbial origin. DNA is sticky and binds
to surfaces and other molecules, thereby inducing membrane fouling either by simply
aggregating smaller molecules into bigger ones or as initiation of biofilm formation. In addition to
the adhesion of microorganisms to surfaces, eDNA also functions as a structural component in
a biofilm. The role of eDNA in biofilms varies from one microbial species to the next, and other
extracellular polymers like proteins and polysaccharides also play important roles, however it is
clear that eDNA plays a major role in microbial attachment to surfaces and early biofilm
formation.
Membranes used for filtration/purification of water or non-aqeous liquids include
ultrafiltration (UF), microfiltration (MF), nanofiltration (NF) and reverse osmosis (RO)
membranes. When the membranes get fouled, in order to maintain a certain flow of liquid
through the membrane, the operator has to increase the pressure over the membrane (trans
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membrane pressure) which results in an increased consumption of power. Above a certain
pressure the integrity of the membrane is irreversible damaged.
The use of DNase enzymes for cleaning water filtration membranes can reduce the
sticking of organic material and microorganisms to the membrane, whereby the flow of
water/liquid through the membrane is improved. Also, the use of a DNase for membrane
cleaning can be combined with traditional chemical cleaning agents like acids, bases, bleach
and other disinfectants and biocides. The DNase cleaning step can be applied either before or
after cleaning procedures with other cleaning chemicals or formulations.
Typically used biocides and cleaning chemicals include 2,2-dibromo-3-
nitrilopropionamide (DBNPA), rodalon, sodium hypochlorite, H202, HN03, citric acid, oxalic acid,
NaOH, EDTA, surfactants, coagulants, antisealants and dispersants.
The membrane cleaning procedure can be either “Cleaning-In-Place” (CIP) or “Cleaning-
Out-of-Place” (COP). In a CIP cleaning procedure the membranes remain installed in the
treatment facility while subjected to the cleaning formulation either in a soaking procedure
wherein the membranes are submerged or soaked in the cleaning formulation or in a flow
through procedure wherein the cleaning formulation is continuously circulated through the
membrane.
In a COP process the membranes are removed from the facility and cleaned in a
separate soaking or flow through procedure.
Dependent on the nature of the fouling, the CIP or COP cleaning procedures can extend
for a few minutes to several hours, such as over night.
A typical membrane cleaning procedure can have several separate steps separated by
flushing with water. The order of the chemicals and DNase cleaning steps can vary based on
the nature of the membrane foulings.
Dependent on the activity profile of the DNase enzyme, the DNase cleaning procedure
can take place at either low, neutral or high pH, and low or high temperature. Preferably, the
procedure takes place at pH 6-9 and temperature between 30-70°C.
DNase (deoxyribonuclease)
The term “DNase” means a polypeptide with DNase activity that catalyzes the hydrolytic
cleavage of phosphodiester linkages in the DNA backbone, thus degrading DNA. Examples of
enzymes exhibiting DNase activity are those covered by enzyme classes EC 3.1.11 to EC
3.1.31, as defined in the recommendations of the Nomenclature Committee of the International
Union of Biochemistry and Molecular Biology (IUBMB).
The terms “DNases”, “DNase enzymes” and the expression “a polypeptide with DNase
activity” are used interchangeably throughout the application. For purposes of the present
invention, DNase activity is determined according to the procedure described in the Assay I.
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In one aspect, the DNase of the present invention has at least 20%, e.g., at least 40%,
at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, or at least
100% of the DNase activity of the DNase having the amino acid sequence of SEQ ID NO: 1.
The DNase used according to the present invention is a mature polypeptide exhibiting
DNase activity, which comprises or consists of an amino acid sequence having at least 80%
sequence identity to the amino acid sequence shown as SEQ ID NO: 1 or SEQ ID NO: 2.
Preferably, the amino acid sequence of the DNase has at least 85% sequence identity,
more preferably at least 90%, more preferably at least 95%, 96%, 97%, 98%, 99%, and most
preferably 100% sequence identity to the amino acid sequence of SEQ ID NO: 1 or SEQ ID NO:
2. In an embodiment, the amino acid sequence of the DNase is SEQ ID NO: 1 or SEQ ID NO: 2.
In another embodiment, the DNase is a fungal DNase, preferably a filamentous fugal DNase,
more preferably an Aspergillus DNase, and most preferably an Aspergillus oryzae DNase. In
another embodiment, the DNase is an Aspergillus oryzae DNase or a derivative thereof. In yet
another embodiment, the DNase is a DNase as disclosed in International patent application no.
PCT/EP2015/057883 (now WO 2015/155350), which is hereby incorporated by reference.
In an embodiment, the number of amino acid substitutions, deletions and/or insertions
introduced into the amino acid sequence of SEQ ID NO: 1 or SEQ ID NO: 2 is up to 10, e.g., 1,
2, 3, 4, 5, 6, 7, 8, 9, or 10; or up to 5, e.g., 1, 2, 3, 4, or 5; or up to 2. The amino acid changes
may be of a minor nature, that is conservative amino acid substitutions or insertions that do not
significantly affect the folding and/or activity of the protein; small deletions, typically of 1-30
amino acids; small amino- or carboxyl-terminal extensions, such as an amino-terminal
methionine residue; a small linker peptide of up to 20-25 residues; or a small extension that
facilitates purification by changing net charge or another function, such as a poly-histidine tract,
an antigenic epitope or a binding domain.
Examples of conservative substitutions are within the groups of basic amino acids
(arginine, lysine and histidine), acidic amino acids (glutamic acid and aspartic acid), polar amino
acids (glutamine and asparagine), hydrophobic amino acids (leucine, isoleucine and valine),
aromatic amino acids (phenylalanine, tryptophan and tyrosine), and small amino acids (glycine,
alanine, serine, threonine and methionine). Amino acid substitutions that do not generally alter
specific activity are known in the art and are described, for example, by H. Neurath and R.L. Hill,
1979, In, The Proteins, Academic Press, New York. Common substitutions are Ala/Ser, Val/lle,
Asp/Glu, Thr/Ser, Ala/Gly, Ala/Thr, Ser/Asn, Ala/Val, Ser/Gly, Tyr/Phe, Ala/Pro, Lys/Arg,
Asp/Asn, Leu/lle, Leu/Val, Ala/Glu, and Asp/Gly.
Essential amino acids in a polypeptide can be identified according to procedures known
in the art, such as site-directed mutagenesis or alanine-scanning mutagenesis (Cunningham
and Wells, 1989, Science 244: 1081-1085). In the latter technique, single alanine mutations are
introduced at every residue in the molecule, and the resultant mutant molecules are tested for
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DNase activity to identify amino acid residues that are critical to the activity of the molecule. See
also, Hilton et al., 1996, J. Biol. Chem. 271: 4699-4708. The active site of the DNase or other
biological interaction can also be determined by physical analysis of structure, as determined by
such techniques as nuclear magnetic resonance, crystallography, electron diffraction, or
photoaffinity labeling, in conjunction with mutation of putative contact site amino acids. See, for
example, de Vos etal., 1992, Science 255: 306-312; Smith etal., 1992, J. Mol. Biol. 224: 899-
904; Wlodaver etal., 1992, FEBS Lett. 309: 59-64. The identity of essential amino acids can
also be inferred from an alignment with a related polypeptide.
Single or multiple amino acid substitutions, deletions, and/or insertions can be made and
tested using known methods of mutagenesis, recombination, and/or shuffling, followed by a
relevant screening procedure, such as those disclosed by Reidhaar-Olson and Sauer, 1988,
Science 241: 53-57; Bowie and Sauer, 1989, Proc. Natl. Acad. Sci. USA 86: 2152-2156; WO
95/17413; or WO 95/22625. Other methods that can be used include error-prone PCR, phage
display {e.g., Lowman etal., 1991, Biochemistry 30: 10832-10837; U.S. Patent No. 5,223,409;
WO 92/06204), and region-directed mutagenesis (Derbyshire et al., 1986, Gene 46: 145; Ner et
al., 1988, DNA 7: 127).
The relatedness between two amino acid sequences is described by the parameter
“sequence identity”. For purposes of the present invention, the sequence identity between two
amino acid sequences is determined using the Needleman-Wunsch algorithm (Needleman and
Wunsch, 1970, J. Mol. Biol. 48: 443-453) as implemented in the Needle program of the
EMBOSS package (EMBOSS: The European Molecular Biology Open Software Suite, Rice et
al., 2000, Trends Genet. 16: 276-277), preferably version 5.0.0 or later. The parameters used
are gap open penalty of 10, gap extension penalty of 0.5, and the EBLOSUM62 (EMBOSS
version of BLOSUM62) substitution matrix. The output of Needle labeled “longest identity”
(obtained using the -nobrief option) is used as the percent identity and is calculated as follows:
(Identical Residues x 100)/(Length of Alignment - Total Number of Gaps in Alignment).
Biofilm
A biofilm is any group of microorganisms in which cells stick to each other on a surface,
such as a fibrous surface, a metallic surface, or any other hard surface. These adherent cells
are frequently embedded within a self-produced matrix of extracellular polymeric substance
(EPS). Biofilm EPS is a polymeric conglomeration generally composed of extracellular DNA,
proteins, and polysaccharides.
Biofilms may form on living or non-living surfaces. The microbial cells growing in a
biofilm are physiologically distinct from planktonic cells of the same organism, which, by
contrast, are single-cells that may float or swim in a liquid medium. Bacteria living in a biofilm
usually have significantly different properties from free-floating bacteria of the same species, as
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the dense and protected environment of the film allows them to cooperate and interact in
various ways. One benefit of this environment is increased resistance to detergents and
antibiotics, as the dense extracellular matrix and the outer layer of cells protect the interior of the
community.
Typical biofilms comprise one or more bacteria selected from the group consisting of
Acinetobacter, Bacillus, Comamonas, Escherichia, Pseudomonas, and Sphingomonas species;
such as one or more bacterial species selected from the group consisting of Acinetobacter
calcoaceticus, Bacillus amyloliquefaciens SC100, Bacillus amyloliquefaciens SC168,
Comamonas denitrificans, Escherichia coli K-12, Pseudomonas aeruginosa, and
Sphingomonas mucosissima.
Non-DNase enzyme
The non-DNase enzyme to be optionally combined with the DNase, according to the
invention, may be one or more non-DNase enzymes selected from the group consisting of
amylase, arabinase, carbohydrase, cellulase, cutinase, galactanase, lipase, mannanase,
oxidoreductase (haloperoxidase, laccase, oxidase, peroxidase), pectate lyase, pectinase,
perhydrolase, protease, and xylanase.
In general, the properties of the selected enzyme(s) should be compatible with the
cleaning conditions (i.e., pH-optimum, compatibility with other enzymatic and non-enzymatic
ingredients, etc.), and the enzyme(s) should be present in effective amounts.
Protease
Suitable proteases include those of bacterial, fungal, plant, viral or animal origin, e.g.,
vegetable or microbial origin. Microbial origin is preferred. Chemically modified or protein
engineered mutants are included. It may be an alkaline protease, such as a serine protease or a
metalloprotease. A serine protease may for example be of the S1 family, such as trypsin, or the
S8 family such as subtilisin. A metalloproteases protease may for example be a thermolysin
from, e.g., family M4 or other metalloprotease such as those from M5, M7 or M8 families.
The term "subtilases" refers to a sub-group of serine protease according to Siezen et al.,
1991, Protein Engng. 4: 719-737, and Siezen etal., 1997, Protein Science 6: 501-523. Serine
proteases are a subgroup of proteases characterized by having a serine in the active site, which
forms a covalent adduct with the substrate. The subtilases may be divided into 6 sub-divisions,
i.e., the Subtilisin family, the Thermitase family, the Proteinase K family, the Lantibiotic
peptidase family, the Kexin family and the Pyrolysin family.
Examples of subtilases are those derived from Bacillus such as Bacillus alkalophilus, B.
amyloliquefaciens, Bacillus gibsonii, Bacillus lentus, Bacillus pumilus, and Bacillus subtilis
described in; US 7,262,042 and WO 2009/021867, and subtilisin lentus, subtilisin Novo,
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subtilisin Carlsberg, Bacillus licheniformis, subtilisin BPN’, subtilisin 309, subtilisin 147 and
subtilisin 168 described in WO 89/06279 and protease PD138 described in (WO 93/18140).
Other useful proteases may be those described in WO 92/17577, WO 01/16285, WO 02/26024
and WO 02/16547. Examples of trypsin-like proteases are trypsin (e.g., of porcine or bovine
origin) and the Fusarium protease described in WO 89/06270, WO 94/25583 and WO
2005/040372, and the chymotrypsin proteases derived from Cellumonas described in WO
2005/052161 and WO 2005/052146.
A further preferred protease is the alkaline protease from Bacillus lentus DSM 5483, as
described for example in WO 95/23221, and variants thereof which are described in WO
92/21760, WO 95/23221, EP 1921147 and EP 1921148.
Examples of metalloproteases are the neutral metalloprotease as described in WO
2007/044993 (Genencor Int.) such as those derived from Bacillus amyloliquefaciens.
Examples of useful proteases are the variants described in: WO 92/19729, WO
96/34946, WO 98/20115, WO 98/20116, WO 99/11768, WO 01/44452, WO 03/006602, WO
2004/03186, WO 2004/041979, WO 2007/006305, WO 2011/036263, WO 2011/036264,
especially the variants with substitutions in one or more of the following positions: 3, 4, 9, 15,
27, 36, 57, 68, 76, 87, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 106, 118, 120, 123, 128,
129, 130, 160, 167, 170, 194, 195, 199, 205, 206, 217, 218, 222, 224, 232, 235, 236, 245, 248,
252 and 274 using the BPN’ numbering. More preferred the subtilase variants may comprise the
mutations: S3T, V4I, S9R, A15T, K27R, *36D, V68A, N76D, N87S,R, *97E, A98S, S99G,D,A,
S99AD, S101G,M,R S103A, V104I,Y,N, S106A, G118V,R, H120D.N, N123S, S128L, P129Q,
S130A, G160D, Y167A, R170S, A194P, G195E, V199M, V205I, L217D, N218D, M222S,
A232V, K235L, Q236H, Q245R, N252K, T274A (using BPN’ numbering).
Suitable commercially available protease enzymes include those sold under the trade
names Alcalase™, Duralase™, Durazym™, Relase™, Relase™ Ultra, Savinase™, Savinase™
Ultra, Primase™, Polarzyme™, Kannase™, Liquanase™, Liquanase™ Ultra, Ovozyme™,
Coronase™, Coronase™ Ultra, Blaze™, Neutrase™, Everlase™ and Esperase™ (Novozymes
A/S), those sold under the tradename Maxatase™, Maxacal™, Maxapem™, Purafect™,
Purafect Prime™, Purafect MA™, Purafect Ox™, Purafect OxP™, Puramax™, Properase™,
FN2™, FN3™, FN4™, Excellase™, Eraser™, Opticlean™ and Optimase™ (Danisco/DuPont),
Axapem™ (Gist-Brocases N.V.), BLAP (sequence shown in Figure 29 of US 5,352,604) and
variants hereof (Henkel AG) and KAP (Bacillus alkalophilus subtilisin) from Kao.
Proteases, but also other types of enzymes, may be used together with protease
inhibitors, which are reversible inhibitors of protease activity, e.g., serine protease activity.
Preferably, the protease inhibitor is a (reversible) subtilisin protease inhibitor. In particular, the
protease inhibitor may be a peptide aldehyde, boric acid, or a boronic acid; or a derivative of
any of these.
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Suitable boronic acid derivatives are described in US 4,963,655, US 5,159,060, WO
95/12655, WO 95/29223, WO 92/19707, WO 94/04653, WO 94/04654, US 5,442,100, US
,488,157 and US 5,472,628.
Suitable peptide aldehydes are described in WO 94/04651, WO 95/25791, WO
98/13458, WO 98/13459, WO 98/13460, WO 98/13461, WO 98/13461, WO 98/13462, WO
2007/141736, WO 2007/145963, WO 2009/118375, WO 2010/055052 and WO 2011/036153.
Alternatively, the peptide aldehyde may have the formula as described in WO 2011/036153.
The peptide aldehyde may be converted into a water-soluble hydrosulfite adduct by
reaction with sodium bisulfite, as described in textbooks, e.g., March, J. Advanced Organic
Chemistry, fourth edition, Wiley-lnterscience, US 1992, p 895; and as described in WO
2013/004636.
Lipase/cutinase
Suitable lipases and cutinases include those of bacterial or fungal origin. Chemically
modified or protein engineered mutants are included. Examples include lipase from
Thermomyces, e.g., from T. lanuginosus (previously named Humicola lanuginosa) as described
in EP 258068 and EP 305216, cutinase from Humicola, e.g., H. insolens as described in WO
96/13580, a Pseudomonas lipase, e.g., from P. alcaligenes or P. pseudoalcaligenes (EP
218272), P. cepacia (EP 331376), P. stutzeri (GB 1372034), P. fluorescens, Pseudomonas sp.
strain SD 705 (WO 95/06720 and WO 96/27002), P. wisconsinensis (WO 96/12012), a Bacillus
lipase, e.g., from B. subtilis (Dartois etai, 1993, Biochemica et Biophysica Acta 1131: 253-360),
B. stearothermophilus (JP 64/744992) or B. pumilus (WO 91/16422), GDSL-type Streptomyces
lipases (WO 2010/065455), cutinase from Magnaporthe grisea (WO 2010/107560), cutinase
from Pseudomonas mendocina (US 5,389,536), lipase from Thermobifida fusca (WO
2011/084412), Geobacillus stearothermophilus lipase (WO 2011/084417), lipase from Bacillus
subtilis (WO 2011/084599), and lipase from Streptomyces griseus (WO 2011/150157) and S.
pristinaespiralis (WO 2012/137147).
Other examples are lipase variants such as those described in WO 92/05249, WO
94/01541, EP 407225, EP 260105, WO 95/35381, WO 96/00292, WO 95/30744, WO 94/25578,
WO 95/14783, WO 95/22615, WO 97/04079, WO 97/07202, WO 00/60063, WO 2007/087508
and WO 2009/109500.
Preferred commercially available lipase enzymes include Lipolase™, Lipolase Ultra™,
and Lipex™; Lecitase™, Lipolex™; Lipoclean™, Lipoprime™ (Novozymes A/S). Other
commercially available lipases include Lumafast (Genencor Int Inc); Lipomax (Gist-
Brocades/Genencor Int Inc) and Bacillus sp. lipase from Solvay.
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Carbohydrase
A carbohydrase is a general term for enzymes that cleave carbohydrates. In general
carbohydrases are named after the substrates they act on, for example amylases act on
amylase and cellulases act on cellulose. Many carbohydrases have found use in cleaning and
laundry applications, such as amylase, cellulase, pectinase, pectate lyase, mannanase,
arabinase, galactanase and xylanase, and all these can be applied in the liquid composition.
Amylase
Suitable amylases include those of bacterial or fungal origin. Chemically modified or
protein engineered mutants are included. Amylases include, for example, a-amylases obtained
from Bacillus, e.g., a special strain of Bacillus licheniformis, described in more detail in GB
1,296,839.
Examples of suitable amylases include amylases having SEQ ID NO: 2 in WO 95/10603
or variants having 90% sequence identity to SEQ ID NO: 3 thereof. Preferred variants are
described in WO 94/02597, WO 94/18314, WO 97/43424 and SEQ ID NO: 4 of WO 99/19467,
such as variants with substitutions in one or more of the following positions: 15, 23, 105, 106,
124, 128, 133, 154, 156, 178, 179, 181, 188, 190, 197, 201, 202, 207, 208, 209, 211, 243, 264,
304, 305, 391, 408, and 444.
Different suitable amylases include amylases having SEQ ID NO: 6 in WO 02/10355 or
variants thereof having 90% sequence identity to SEQ ID NO: 6. Preferred variants of SEQ ID
NO: 6 are those having a deletion in positions 181 and 182 and a substitution in position 193.
Other amylases which are suitable are hybrid alpha-amylase comprising residues 1-33 of the
alpha-amylase derived from B. amyloliquefaciens shown in SEQ ID NO: 6 of WO 2006/066594
and residues 36-483 of the B. licheniformis alpha-amylase shown in SEQ ID NO: 4 of WO
2006/066594 or variants having 90% sequence identity thereof. Preferred variants of this hybrid
alpha-amylase are those having a substitution, a deletion or an insertion in one of more of the
following positions: G48, T49, G107, H156, A181, N190, M197, 1201, A209 and Q264. Most
preferred variants of the hybrid alpha-amylase comprising residues 1-33 of the alpha-amylase
derived from B. amyloliquefaciens shown in SEQ ID NO: 6 of WO 2006/066594 and residues
36-483 of SEQ ID NO: 4 are those having the substitutions:
M197T;
H156Y+A181T+ N190F+A209V+Q264S; or
G48A+T49I+G107A+H156Y+A181T+ N190F+I201F+A209V+Q264S.
Further amylases which are suitable are amylases having SEQ ID NO: 6 in WO
99/19467 or variants thereof having 90% sequence identity to SEQ ID NO: 6. Preferred variants
of SEQ ID NO: 6 are those having a substitution, a deletion or an insertion in one or more of the
following positions: R181, G182, H183, G184, N195, I206, E212, E216 and K269. Particularly
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preferred amylases are those having deletion in positions R181 and G182, or positions H183
and G184.
Additional amylases which can be used are those having SEQ ID NO: 1, SEQ ID NO: 3,
SEQ ID NO: 2 or SEQ ID NO: 7 of WO 96/23873 or variants thereof having 90% sequence
identity to SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3 or SEQ ID NO: 7. Preferred variants of
SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3 or SEQ ID NO: 7 are those having a substitution,
a deletion or an insertion in one or more of the following positions: 140, 181, 182, 183, 184, 195,
206, 212, 243, 260, 269, 304 and 476. More preferred variants are those having a deletion in
positions 181 and 182 or positions 183 and 184. Most preferred amylase variants of SEQ ID
NO: 1, SEQ ID NO: 2 or SEQ ID NO: 7 are those having a deletion in positions 183 and 184
and a substitution in one or more of positions 140, 195, 206, 243, 260, 304 and 476.
Other amylases which can be used are amylases having SEQ ID NO: 2 of WO
2008/153815, SEQ ID NO: 10 in WO 01/66712 or variants thereof having 90% sequence
identity to SEQ ID NO: 2 of WO 2008/153815 or 90% sequence identity to SEQ ID NO: 10 in
WO 01/66712. Preferred variants of SEQ ID NO: 10 in WO 01/66712 are those having a
substitution, a deletion or an insertion in one of more of the following positions: 176, 177, 178,
179, 190, 201, 207, 211 and 264.
Further suitable amylases are amylases having SEQ ID NO: 2 of WO 2009/061380 or
variants having 90% sequence identity to SEQ ID NO: 2 thereof. Preferred variants of SEQ ID
NO: 2 are those having a truncation of the C-terminus and/or a substitution, a deletion or an
insertion in one of more of the following positions: Q87, Q98, S125, N128, T131, T165, K178,
R180, S181, T182, G183, M201, F202, N225, S243, N272, N282, Y305, R309, D319, Q320,
Q359, K444 and G475. More preferred variants of SEQ ID NO: 2 are those having the
substitution in one of more of the following positions: Q87E,R, Q98R, S125A, N128C, T131I,
T165I, K178L, T182G, M201L, F202Y, N225E.R, N272E.R, S243Q,A,E,D, Y305R, R309A,
Q320R, Q359E, K444E and G475K and/or deletion in position R180 and/or S181 or of T182
and/or G183. Most preferred amylase variants of SEQ ID NO: 2 are those having the
substitutions:
N128C+K178L+T182G+Y305R+G475K;
N128C+K178L+T182G+F202Y+Y305R+D319T+G475K;
S125A+N128C+K178L+T182G+Y305R+G475K; or
S125A+N128C+T131I+T165I+K178L+T182G+Y305R+G475K wherein the variants are C-
terminally truncated and optionally further comprises a substitution at position 243 and/or a
deletion at position 180 and/or position 181.
Other suitable amylases are the alpha-amylase having SEQ ID NO: 12 in WO 01/66712
or a variant having at least 90% sequence identity to SEQ ID NO: 12. Preferred amylase
variants are those having a substitution, a deletion or an insertion in one of more of the following
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positions of SEQ ID NO: 12 in WO 01/66712: R28, R118, N174; R181, G182, D183, G184,
G186, W189, N195, M202, Y298, N299, K302, S303, N306, R310, N314; R320, H324, E345,
Y396, R400, W439, R444, N445, K446, Q449, R458, N471, N484. Particularly preferred
amylases include variants having a deletion of D183 and G184 and having the substitutions
R118K, N195F, R320K and R458K, and a variant additionally having substitutions in one or
more position selected from the group: M9, G149, G182, G186, M202, T257, Y295, N299,
M323, E345 and A339, most preferred a variant that additionally has substitutions in all these
positions.
Other examples are amylase variants such as those described in WO 2011/098531, WO
2013/001078 and WO 2013/001087.
Commercially available amylases are Stainzyme™, Stainzyme Plus™, Amplify™,
Resilience™, Everest™, Duramyl™, Termamyl™, Termamyl Ultra™; Natalase™, Fungamyl™
and BAN™ (Novozymes A/S), Rapidase™ and Purastar™/Effectenz™, Powerase™ and
Preferenz S100 (from Genencor International Inc./DuPont).
Lyases
The lyase may be a pectate lyase derived from Bacillus, particularly B. agaradhaerens or
B. licherniformis, or a variant derived of any of these, e.g., as described in US 6,124,127, WO
99/27083, WO 99/07084, WO 02/06442, WO 02/092741, WO 03/095638, Commercially
available pectate lyases are XPect™, Pectawash™, and Pectaway™ (Novozymes A/S).
Mannanase
The mannanase may be an alkaline mannanase of Family 5 or 26. It belongs It may be a
wild-type from Bacillus or Humicola, particularly B. agaradhaerens, B. clausii, B. halodurans, B.
licheniformis, or H. insolens. Suitable mannanases are described in WO 99/64619. A
commercially available mannanase is Mannaway™ (Novozymes A/S).
Cellulase
Suitable cellulases may be of bacterial or fungal origin. Chemically or genetically
modified mutants are included. Suitable cellulases include cellulases from the genera
Acremonium, Bacillus, Fusarium, Humicola, Pseudomonas, Thielavia, e.g., the fungal cellulases
produced from Fusarium oxysporum, Humicola insolens, and Myceliophthora thermophila
disclosed in US 4,435,307, US 5,648,263, US 5,691,178, US 5,776,757 and WO 89/09259.
Especially suitable cellulases are the alkaline or neutral cellulases having color care
benefits. Examples of such cellulases are cellulases described in EP 0 495 257, EP 0 531 372,
WO 96/11262, WO 96/29397, WO 98/08940. Other examples are cellulase variants such as
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those described in WO 94/07998, EP 0 531 315, US 5,457,046, US 5,686,593, US 5,763,254,
WO 95/24471, WO 98/12307 and WO 99/01544.
Commercially available cellulases include Carezyme™, Celluzyme™, Celluclean™,
Celluclast™, Endolase™, Renozyme™, Whitezyme™ (Novozymes A/S); Clazinase™, Puradax,
Puradax HA, and Puradax EG (available from Genencor) and KAC-500(B)™ (Kao Corporation).
Peroxidases/Oxidases
Suitable peroxidases are comprised by the enzyme classification EC 1.11.1.7, as set out
by the Nomenclature Committee of the International Union of Biochemistry and Molecular
Biology (IUBMB), or any fragment derived therefrom, exhibiting peroxidase activity.
Suitable peroxidases include those of plant, bacterial or fungal origin. Chemically
modified or protein engineered mutants are included. Examples of useful peroxidases include
peroxidases from Coprinopsis, e.g., from C. cinerea (EP 179,486), and variants thereof as those
described in WO 93/24618, WO 95/10602, and WO 98/15257.
The peroxidases also include a haloperoxidase enzyme, such as chloroperoxidase,
bromoperoxidase and compounds exhibiting chloroperoxidase or bromoperoxidase activity.
Haloperoxidases are classified according to their specificity for halide ions. Chloroperoxidases
(E.C. 1.11.1.10) catalyze formation of hypochlorite from chloride ions.
In an embodiment, the haloperoxidase of the invention is a chloroperoxidase. Preferably,
the haloperoxidase is a vanadium haloperoxidase, i.e., a vanadate-containing haloperoxidase.
In a preferred method of the present invention the vanadate-containing haloperoxidase is
combined with a source of chloride ion.
Haloperoxidases have been isolated from many different fungi, in particular from the
fungus group dematiaceous hyphomycetes, such as Alternaria, Botrytis, Caldariomyces, e.g., C.
fumago, Curvularia, e.g., C. verruculosa and C. inaequalis, Drechslera, and Ulocladium.
Haloperoxidases have also been isolated from bacteria such as Pseudomonas, e.g., P.
pyrrocinia, and Streptomyces, e.g., S. aureofaciens.
In a preferred embodiment, the haloperoxidase is derivable from Curvularia sp., in
particular Curvularia inaequalis or Curvularia verruculosa, such as C. inaequalis CBS 102.42 as
described in WO 95/27046; or C. verruculosa CBS 147.63 or C. verruculosa CBS 444.70 as
described in WO 97/04102; Dendryphiella salina as described in WO 01/79458; Drechslera
hartlebii as described in WO 01/79459; Geniculosporium sp. as described in WO 01/79460; or
Phaeotrichoconis crotalarie as described in WO 01/79461.
Suitable oxidases include, in particular, any laccase enzyme comprised by the enzyme
classification EC 1.10.3.2, or any fragment derived therefrom exhibiting laccase activity, or a
compound exhibiting a similar activity, such as a catechol oxidase (EC 1.10.3.1), an o-
aminophenol oxidase (EC 1.10.3.4), ora bilirubin oxidase (EC 1.3.3.5).
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Preferred laccase enzymes are enzymes of microbial origin. The enzymes may be
derived from plants, bacteria or fungi (including filamentous fungi and yeasts).
Suitable examples from fungi include a laccase derivable from a strain of Aspergillus,
Botrytis, Collybia, Coprinopsis, e.g., C. cinerea, C. comatus, C. friesii, and C. plicatilis, Coriolus,
e.g., C. hirsutus (JP 2238885), Fomes, Lentinus, Myceliophthora, e.g., M. thermophila,
Neurospora, e.g., N. crassa, Panaeolus, e.g., P. papilionaceus, Phlebia, e.g., P. radiata (WO
92/01046), Pleurotus, Podospora, Polyporus, e.g., P. pinsitus, Psathyrella, e.g., P. condelleana,
Rhizoctonia, e.g., R. solani, Schytalidium, e.g., S. thermophilum, or Trametes, e.g., T. villosa
and T. versicolor.
Suitable examples from bacteria include a laccase derivable from a strain of Bacillus.
A laccase derived from Coprinopsis or Myceliophthora is preferred; in particular a
laccase derived from Coprinopsis cinerea, as disclosed in WO 97/08325; or from
Myceliophthora thermophila, as disclosed in WO 95/33836.
Perhydrolase
Suitable perhydrolases are capable of catalyzing a perhydrolysis reaction that results in
the production of a peracid from a carboxylic acid ester (acyl) substrate in the presence of a
source of peroxygen (e.g., hydrogen peroxide). While many enzymes perform this reaction at
low levels, perhydrolases exhibit a high perhydrolysis:hydrolysis ratio, often greater than 1.
Suitable perhydrolases may be of plant, bacterial or fungal origin. Chemically modified or protein
engineered mutants are included.
Examples of useful perhydrolases include naturally occurring Mycobacterium
perhydrolase enzymes, or variants thereof. An exemplary enzyme is derived from
Mycobacterium smegmatis. Such enzyme, its enzymatic properties, its structure, and variants
thereof, are described in WO 2005/056782, WO 2008/063400, US 2008/0145353, and US
2007/0167344.
Water filtration membranes
A variety of membrane types and configurations can be used in water or wastewater
treatment processes. Types of membrane configurations include capillary tube, tubular, hollow
fiber, multi-tube, plat-and-frame/flat sheet, pleated cartridge filter, spiral wound, and ceramic
including ceramic disc. Membranes can be made from one or more materials including, for
example, chlorinated polyethylene, polyacrylonitrile, polysulfone, polyethersulfone,
polyvinylalcohol, cellolose acetate, regenerated cellulose, polyvinylidene difluoride,
polyethlysulphone, polyethylene, polypropylene, and ceramic material. Other characteristics of
the membranes that can vary based on the application include, for example, the membrane
pore size. The size of the membrane pores may be larger or smaller depending upon the size of
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particulate or impurity being removed from the water or wastewater. Membrane types, according
to the present invention, include those utilized for ultrafiltration, microfiltration, and nanofiltration.
Membrane Bioreactor Systems
Membrane bioreactor (MBR) systems typically combine two basic processes: biological
degradation and membrane separation, into a single process where suspended solids and
microorganisms responsible for biodegradation are separated from the treated water by a
membrane filtration unit. See, for example, Water Treatment Membrane Processes, McGraw-
Hill, 1996, p. 17.2. The entire biomass is confined within the system, providing for both control of
the residence time for the microorganisms in the reactor (sludge age) and the disinfection of the
effluent.
In a typical MBR unit, influent wastewater is pumped or gravity fed into an aeration tank
where it is brought into contact with the biomass which biodegrades organic material in the
wastewater. Aeration means such as blowers provide oxygen to the biomass. The resulting
mixed liquor is pumped or gravity fed from the aeration tank into the membrane module where it
is mechanically or gravitationally filtered through a membrane under pressure or is drawn
through a membrane under low vacuum. In some systems, the aeration tank and the membrane
tank are the same tank. The effluent is discharged from the system while the concentrated
mixed liquor is returned to the bioreactor. Excess sludge is pumped out in order to maintain a
constant sludge age, and the membrane is regularly cleaned by backwashing, chemical
washing, air scouring, or any combination of these mechanisms.
MBR systems have multiple configurations. Two main MBR process configurations
include submerged/immersed and sidestream. There are also two primary mechanisms of
hydraulic operation including pumping and airlifting. These configurations and bulk liquid
transfer modes are typically referred to as conventional biomass rejection MBRs. Other
configurations include extractive and diffusive process modes which employ membranes for
purposes other than separating biomass from the treated water. All of these process
configurations include one or more membrane units comprising membranes such as those
described in the “Membranes” section above.
In one embodiment, the membranes are present in a membrane bioreactor. In another
embodiment, the wastewater treatment process occurs in a membrane bioreactor in which the
membrane flat-sheet cassette unit, or hollow-fiber unit, itself is typically immersed.
In one embodiment, the wastewater is pretreated prior to entering the membrane
bioreactor. Pretreatment can occur at the source of the wastewater, at a pretreatment plant, or
as part of the overall MBR system. Such pretreatments can include a bar screen, grit chamber,
or rotary drum screen to achieve coarse solids removal. Other pretreatments may include
removal of substances such as harmful pollutants, oils or fuels, or other toxic substances.
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Water treatment processes
One or more water treatment processes are contemplated by the present invention.
Such water treatment processes include, but are not limited to, reverse osmosis, water
desalination and drinking water purification, and wastewater treatment processes. The water or
wastewater, according to the present invention, can be from any source including human waste,
cesspit leakage, septic tank discharge, sewage plant discharge, washing water such as
greywater or sullage, collected rainwater, groundwater, surplus manufactured liquids, seawater,
river water, manmade liquid disposal, highway drainage, storm drains, blackwater, process
water from industries manufacturing products including electronics, transportation vehicles,
pharmaceuticals, paints, lubricants, plastics, crude oil and gas, and thereof derived refined
products, feed, food and beverages, textile, non-wowens, paper and pulp, processed grains and
vegetable oils, chemicals, proteins (e.g., enzymes), industrial waste, industrial site wastewater
or drainage such as cooling or process waters, and agricultural wastewater or drainage.
Liquid cleaning composition
The liquid cleaning composition used according to the invention has a physical form,
which is not solid (or gas). It may be a pourable liquid, a pourable gel or a non-pourable gel. It
may be either isotropic or structured, preferably isotropic.
The liquid cleaning composition may be aqueous, typically containing at least 20% by
weight and up to 95% water, such as up to 70% water, up to 50% water, up to 40% water, up to
% water, or up to 20% water. Other types of liquids, including without limitation, alkanols,
amines, diols, ethers and polyols may be included in an aqueous liquid composition. An
aqueous liquid composition may contain from 0-30% organic solvent. A liquid composition may
even be non-aqueous, wherein the water content is below 10%, preferably below 5%.
The cleaning composition may take the form of a unit dose product. A unit dose product
is the packaging of a single dose in a non-reusable container. It is increasingly used in
detergents for laundry and dish wash. A detergent unit dose product is the packaging (e.g., in a
pouch made from a water soluble film) of the amount of detergent used for a single wash.
Pouches can be of any form, shape and material which is suitable for holding the
composition, e.g., without allowing the release of the composition from the pouch prior to water
contact. The pouch is made from water soluble film which encloses an inner volume. Said inner
volume can be divided into compartments of the pouch. Preferred films are polymeric materials
preferably polymers which are formed into a film or sheet. Preferred polymers, copolymers or
derivates thereof are selected polyacrylates, and water soluble acrylate copolymers, methyl
cellulose, carboxy methyl cellulose, sodium dextrin, ethyl cellulose, hydroxyethyl cellulose,
hydroxypropyl methyl cellulose, malto dextrin, poly methacrylates, most preferably polyvinyl
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alcohol copolymers and, hydroxypropyl methyl cellulose (HPMC). Preferably the level of
polymer in the film for example PVA is at least about 60%. Preferred average molecular weight
will typically be about 20,000 to about 150,000. Films can also be a blend compositions
comprising hydrolytically degradable and water soluble polymer blends such as polyactide and
polyvinyl alcohol (known under the Trade reference M8630 as sold by Chris Craft In. Prod. Of
Gary, Ind., US) plus plasticizers like glycerol, ethylene glycerol, Propylene glycol, sorbitol and
mixtures thereof. The pouches can comprise a solid laundry cleaning composition or part
components and/or a liquid cleaning composition or part components separated by the water
soluble film. The compartment for liquid components can be different in composition than
compartments containing solids (see e.g., US 2009/0011970).
The choice of cleaning components may include the consideration of the type of
membrane to be cleaned, the type and/or degree of soiling, the temperature at which cleaning is
to take place, and the formulation of the cleaning product. Although components mentioned
below are categorized by general header according to a particular functionality, this is not to be
construed as a limitation, as a component may comprise additional functionalities as will be
appreciated by the skilled artisan.
The choice of additional components is within the skill of the artisan and includes
conventional ingredients, including the exemplary non-limiting components set forth below.
Surfactants
The cleaning composition may comprise one or more surfactants, which may be anionic
and/or cationic and/or non-ionic and/or semi-polar and/or zwitterionic, or a mixture thereof. In a
particular embodiment, the composition includes a mixture of one or more nonionic surfactants
and one or more anionic surfactants. The surfactant(s) is typically present at a level of from
about 0.1% to 60% by weight, such as about 1% to about 40%, or about 3% to about 20%, or
about 3% to about 10%. The surfactant(s) is chosen based on the desired cleaning application,
and includes any conventional surfactant(s) known in the art. Any surfactant known in the art for
use in cleaning compositions may be utilized.
When included therein the cleaning composition will usually contain from about 1% to
about 40% by weight, such as from about 5% to about 30%, including from about 5% to about
%, or from about 20% to about 25% of an anionic surfactant. Non-limiting examples of
anionic surfactants include sulfates and sulfonates, in particular, linear alkylbenzenesulfonates
(LAS), isomers of LAS, branched alkylbenzenesulfonates (BABS), phenylalkanesulfonates,
alpha-olefinsulfonates (AOS), olefin sulfonates, alkene sulfonates, alkane-2,3-diylbis(sulfates),
hydroxyalkanesulfonates and disulfonates, alkyl sulfates (AS) such as sodium dodecyl sulfate
(SDS), fatty alcohol sulfates (FAS), primary alcohol sulfates (PAS), alcohol ethersulfates (AES
or AEOS or FES, also known as alcohol ethoxysulfates or fatty alcohol ether sulfates),
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secondary alkanesulfonates (SAS), paraffin sulfonates (PS), ester sulfonates, sulfonated fatty
acid glycerol esters, alpha-sulfo fatty acid methyl esters (alpha-SFMe or SES) including methyl
ester sulfonate (MES), alkyl- or alkenylsuccinic acid, dodecenyl/tetradecenyl succinic acid
(DTSA), fatty acid derivatives of amino acids, diesters and monoesters of sulfo-succinic acid or
soap, and combinations thereof.
When included therein the cleaning composition will usually contain from about 0.1% to
about 10% by weight of a cationic surfactant. Non-limiting examples of cationic surfactants
include alklydimethylethanolamine quat (ADMEAQ), cetyltrimethylammonium bromide (CTAB),
dimethyldistearylammonium chloride (DSDMAC), and alkylbenzyldimethylammonium, alkyl
quaternary ammonium compounds, alkoxylated quaternary ammonium (AQA) compounds, and
combinations thereof.
When included therein the cleaning composition will usually contain from about 0.2% to
about 40% by weight of a non-ionic surfactant, for example from about 0.5% to about 30%, in
particular from about 1% to about 20%, from about 3% to about 10%, such as from about 3% to
about 5%, or from about 8% to about 12%. Non-limiting examples of non-ionic surfactants
include alcohol ethoxylates (AE or AEO), alcohol propoxylates, propoxylated fatty alcohols
(PFA), alkoxylated fatty acid alkyl esters, such as ethoxylated and/or propoxylated fatty acid
alkyl esters, alkylphenol ethoxylates (APE), nonylphenol ethoxylates (NPE), alkylpolyglycosides
(APG), alkoxylated amines, fatty acid monoethanolamides (FAM), fatty acid diethanolamides
(FADA), ethoxylated fatty acid monoethanolamides (EFAM), propoxylated fatty acid
monoethanolamides (PFAM), polyhydroxy alkyl fatty acid amides, or A/-acyl A/-alkyl derivatives
of glucosamine (glucamides, GA, or fatty acid glucamide, FAGA), as well as products available
under the trade names SPAN and TWEEN, and combinations thereof.
When included therein the cleaning composition will usually contain from about 0.1% to
about 20% by weight of a semipolar surfactant. Non-limiting examples of semipolar surfactants
include amine oxides (AO) such as alkyldimethylamineoxide, N-(coco alkyl)-A/,A/-dimethylamine
oxide and A/-(tallow-alkyl)-A/,A/-bis(2-hydroxyethyl)amine oxide, fatty acid alkanolamides and
ethoxylated fatty acid alkanolamides, and combinations thereof.
When included therein the cleaning composition will usually contain from about 0.1% to
about 10% by weight of a zwitterionic surfactant. Non-limiting examples of zwitterionic
surfactants include betaine, alkyldimethylbetaine, sulfobetaine, and combinations thereof.
Methods of Use
As described in the above paragraphs, the present invention provides a method for
cleaning a water filtration membrane by contacting the membrane with a liquid cleaning
composition comprising a DNase. Preferably, the water flow rate through the membrane (flux) is
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improved. The liquid cleaning composition may further include the ingredients as described in
the paragraphs above.
In an embodiment, the cleaning process is Cleaning-In-Place (CIP) or Cleaning-Out-of-
Place (COP).
In an embodiment, the DNase treatment of the water filtration membrane is preceded or
followed by contacting the membrane with a biocidal composition.
In an embodiment, the water filtration membrane comprises a biofilm. Preferably, the
biofilm comprises one or more bacteria selected from the group consisting of Acinetobacter,
Bacillus, Comamonas, Escherichia, Pseudomonas, and Sphingomonas species. More
preferably, the biofilm comprises one or more bacteria selected from the group consisting of
Acinetobacter calcoaceticus, Bacillus amyloliquefaciens SC 100, Bacillus amyloliquefaciens
SC168, Comamonas denitrificans, Escherichia coli K-12, Pseudomonas aeruginosa, and
Sphingomonas mucosissima,׳ preferably the bacteria are Pseudomonas aeruginosa.
In another embodiment, the pH of the cleaning composition is 6-9; preferably the pH is
about neutral.
In an embodiment, the membrane is contacted with the cleaning composition at a
temperature between 30-100°C, preferably at a temperature between 50-80°C.
In yet another embodiment, the membrane is simultaneously or separately contacted
with one or more non-DNase enzyme(s) selected from the group consisting of protease, lipase,
cutinase, amylase, carbohydrase, cellulase, pectinase, pectate lyase, mannanase, arabinase,
galactanase, xylanase, perhydrolase, and oxidoreductase; preferably the non-DNase enzyme is
a protease.
In an embodiment, the DNase used in the cleaning composition is an Aspergillus oryzae
DNase or a derivative thereof. Preferably, the DNase is a polypeptide having DNase activity,
which comprises or consists of an amino acid sequence having at least 80% sequence identity
to the amino acid sequence shown as SEQ ID NO: 1. More preferably, the DNase is a
polypeptide having DNase activity, which comprises or consists of the amino acid sequence
shown as SEQ ID NO: 1.
In another embodiment, the DNase is a Bacillus licheniformis DNase or a derivative
thereof. Preferably, the DNase is a polypeptide having DNase activity, which comprises or
consists of an amino acid sequence having at least 80% sequence identity to the amino acid
sequence shown as SEQ ID NO: 2.
Assay I: DNase activity
DNase activity was determined on DNase Test Agar with Methyl Green (BD, Franklin
Lakes, NJ, USA), which was prepared according to the manual from supplier. Briefly, 21 g of the
DNase Test Agar was dissolved in 500 ml water and then autoclaved for 15 min at 121°C.
-18-WO 2017/066510 PCT/US2016/056955
Autoclaved agar was temperated to 48°C in water bath, and 20 ml of agar was poured into
petridishes with and allowed to solidify by incubation 0/n at room temperature. On solidified agar
plates, 5 pi of enzyme solutions are added, and DNase activity are observed as colorless zones
around the spotted enzyme solutions.
EXAMPLES
Chemicals used as buffers and substrates were commercial products of at least reagent
grade. The DNase used in Example 1 was obtained from Aspergillus oryzae and is the mature
polypeptide of the amino acid sequence shown in SEQ ID NO: 1.
EXAMPLE 1
Treatment of water filtration membranes with DNase
Bacteria and Culture Media
Seven microbes commonly found in wastewater and water treatment facilities were used
to create a multispecies biofilm for this study. The following microbes were chosen:
Pseudomonas aeruginosa (ATCC 19429)
Acinetobacter calcoaceticus (ATCC 23055)
Comamonas denitrificans (ATCC 700937)
Escherichia coli K-12 (ATCC 10798)
Sphingomonas mucosissima (NRRL B-59454)
Bacillus amyloliquefaciens SC 100
Bacillus amyloliquefaciens SC 168
The bacteria were cultivated in Luria Broth (LB; 15.5 g/L; Beckton Dickenson, Franklin
Lakes, NJ) at 30°C at 200 RPM overnight for the biofilm assays performed. All biofilm assays
were performed with LB.
Filter Plate Flux Assay Calibration Curve
To assess flux changes, an indicator dye, Brilliant Green (Clariant Corp, Switzerland),
was used to determine the volume of liquid able to pass through the membranes of an EMD
Millipore MultiScreenH1s 96-Well Filter Plate. A calibration curve was established for Brilliant
Green at a 0.05% (wt/vol) concentration in distilled water. A Costar® 96-well black assay plate
with a clear flat bottom was inoculated with volumes of 0.05% Brilliant Green dye ranging from
-240 pL. These volume series were run in triplicate and measured on the Biotek Synergy H4
Hybrid Reader Kinetic Plate Reader (KPR) at an absorbance at 610 nm. Based on the
absorbance readings from each of the corresponding volumes, a standard curve was created in
excel by plotting the absorbance vs. the volume corresponding to that absorbance reading. This
-19-WO 2017/066510 PCT/US2016/056955
standard curve generated a linear equation that was used to assess the volume of liquid able to
pass through the membranes of the filter plate.
96-Well Filter Plate Flux Assay
Cultures were grown separately overnight in LB at 30°C, shaking at 200 RPM. Culture
growth was measured by optical density at 600 nm using a Thermo Scientific Spectronic 20D+
spectrophotometer. Each of the cultures was diluted to an OD 600 of 0.5. Diluted cultures were
added together in equal 1 mL volumes in a separate tube and gently vortexed. An EMD
Millipore MultiScreenH1s 96-Well Filter Plate was loaded with the mixed cultures. A multi-channel
pipette dispensed 100 pL aliquots of the mixed culture into columns 1-4 and 9-12. An LB media
control was added in 100 pL aliquots to columns 5-8. Once the plate was set-up, a “breathe-
easy” film was applied to the plate. The plate was taped inside a Tupperware container along
with a damp WYPALL towel to maintain 99% relative humidity to ensure the plate did not dry
out. The Tupperware was placed on a shaker at 200 RPM for 7 days at 30°C.
After the 7 day incubation of the 96-well filter plate, the “breathe-easy” film was removed
in the BSL II hood. The plate was inverted over a WYPALL towel, allowing the majority of the
liquid in the wells to be removed. Each well was washed with 200 pL of phosphate buffer. The
plate was immediately inverted and the buffer was removed from the wells of the plate. To
ensure all of the liquid was removed from the wells, the inverted plate was gently tapped on the
towel.
From here, the following paths were taken to determine the effectiveness of DNase
treatment on bio-fouled membranes:
Effect of Temperature
An ultrafiltrate DNase sample was diluted to a final concentration of 1 ppm in sterile
water on the tested plates, with treated wells being chosen semi-randomly. Sterile water was
added to wells not receiving DNase treatment. Two plates were incubated at respectively room
temperature (22°C) and optimum temperature (60°C) for one hour. After incubation, the plates
were inverted and the liquid was removed from the wells. The plates were rinsed once again
with phosphate buffer and the liquid was removed.
Effect of Enzyme Concentration
An ultrafiltrate DNase sample was diluted to a final concentration of respectively 0.1 ppm
and 1 ppm in sterile water on two separate plated plates, with treated wells being chosen semi-
randomly. Sterile water was added to wells not receiving DNase treatment. The two plates were
incubated at room temperature (22°C) for 20 minutes. After incubation, the plates were inverted
-20-WO 2017/066510 PCT/US2016/056955
and the liquid was removed from the wells. The plates were rinsed once again with phosphate
buffer and the liquid was removed.
Effect of Time
An ultrafiltrate DNase sample was diluted to a final concentration of 1 ppm in sterile
water on the tested plates, with treated wells being chosen semi-randomly. Sterile water was
added to wells not receiving DNase treatment. Five plates were incubated at room temperature
(22°C) for respectively 0, 5, 10, 20 and 60 minutes. After incubation, the plates were inverted
and the liquid was removed from the wells. The plates were rinsed once again with phosphate
buffer and the liquid was removed.
Once each of the plates was rinsed with phosphate buffer, a 200 pL sample of 0.05%
Brilliant Green indicator dye was added to each well. The filter plate was placed over a Costar®
96-well black with a clear flat bottom and used as a collection plate. This combination was
placed in an Eppendorf Centrifuge 5810 along with a blank counter-balance. The centrifuge was
set for one minute at 150 ref at room temperature (22°C). After centrifugation, the collection
plate was removed from the centrifuge and the volume of the indicator dye that passed through
the membranes into each of the respective wells was measured on the Biotek Synergy H4
Hybrid Reader Kinetic Plate Reader (KPR) at an absorbance at 610 nm. Using the equation
generated from the calibration curve (described above), a volume could be calculated from each
well of the collection plate. Flux percentage was calculated by dividing the treated wells by their
corresponding un-fouled controls. This was performed with un-treated fouled wells as well, to
show flux comparisons between membranes treated with DNase and those that were not.
96-Well Filter Plate Flux Assay Pre-treated With DNase
The seven microorganisms previously mentioned were grown separately overnight in LB
at 30°C, shaking at 200 RPM. Culture growth was measured by optical density at 600 nm using
a Thermo Scientific Spectronic 20D+ spectrophotometer. Each culture was diluted to an OD 600
of 0.5. Diluted cultures were added together in equal 1 mL volumes in a separate tube and
gently vortexed. An EMD Millipore MultiScreenHTs 96-Well Filter Plate was loaded with the
mixed cultures as well as ultrafiltrate DNase of three species (Aspergillus oryzae, Bacillus
licheniformis and Bacillus subtilis) in selected wells. Treated wells were chosen randomly, while
sterile water was added to wells not receiving DNase treatment. Once the plate was set-up, a
“breathe-easy” film was applied to the plate. The plate was taped inside a Tupperware container
along with a damp WYPALL towel to maintain 99% relative humidity to ensure the plate did not
dry out. The Tupperware was placed on a shaker at 200 RPM for 7 days at 30°C.
After the 7 day incubation of the 96-well filter plate, the “breathe-easy” film was removed
in a BSL II hood. The plate was inverted over a WYPALL towel, allowing all of the liquid in the
-21 -WO 2017/066510 PCT/US2016/056955
wells to be removed. Wells were washed with 200 |jL of phosphate buffer. The plate was
immediately inverted and the buffer was removed from the wells of the plate. To ensure all of
the liquid was removed from the wells, the inverted plate was gently tapped on the towel.
Once the filter plates were rinsed with phosphate buffer, 200 pL of 0.05% Brilliant Green
indicator dye was added to each well. The filter plate was placed over a Costar® 96-well black
with a clear flat bottom assay plate used as a collection plate. This combination was placed in
an Eppendorf Centrifuge 5810 along with a blank counter-balance. The centrifuge was set for a
minute at 150 ref at room temperature (22°C). After centrifugation, the collection plate was
removed from the centrifuge and the volume of the indicator dye that passed through the
membranes into the collection plate was measured on the Biotek Synergy H4 Hybrid Reader
Kinetic Plate Reader (KPR) at an absorbance at 610 nm. The volume passed through each well
was calculated using the calibration curve (described above). By dividing the treated wells by
their corresponding un-fouled controls, a flux percentage was generated. This was performed
with un-treated fouled wells as well, to show flux comparisons between membranes treated with
DNase and those that were not.
Results
Effect of Temperature
Temperature showed little effect on DNase efficacy, as DNase treatments administered
at room temperature showed very little to no difference in flux compared to the DNase
treatments administered at the elevated temperatures. The flux of biofouled membranes treated
with DNase was still greater than 88% compared to un-fouled controls (see Table 1).
Effect of Enzyme Concentration
When comparing DNase concentrations of 1 ppm and 0.1 ppm, there was little to no
difference between the dosages.
Effect of Time
At each of the timepoints (5 minutes, 10 minutes, 20 minutes, 60 minutes), each of the
DNase treatments showed a significant increase in the flux compared to the water treated wells.
However, little improvement in flux was noted after the treatment was applied for longer than 20
minutes. At this point in time, the DNase treated membranes reach over 90% of the flux to that
of the unfouled membranes (see Table 1).
Conclusion
The percent flow through volume passed through membranes of a 96-well filter plate
fouled by a multi-species biofilm pre-treated with the ultrafiltrate DNase of Aspergillus oryzae at
-22-WO 2017/066510 PCT/US2016/056955
1 ppm concentrations. Wells treated with the ultrafiltrates showed a significant increase in flux
compared to those that went untreated. Flux was restored in fouled membranes to over 88% of
the flux of un-fouled membranes.
Table 1. Overview of 96-Well filter plate flux assay results.
Aspergillus oryzae Ultrafiltrate
Flux Volume/Control Volume (Percent Flux)
1ppm Optimum Temperature 155.88uL/169.05pL(91.7%)
Untreated 120.72pL/167.69pL (72.0%)
1ppm Room Temperature 155.04pL/174.03pL (89.6%)
Untreated 105.52pL/172.95pL (61.0%)
0.1 ppm Room Temperature 20 minutes 150.52pL/158.59pL (94.9%)
Untreated 106.3pL/162.0pL (65.6%)
1ppm 5 minutes Room Temperature 101.08pL/168.84pL (59.9%)
Untreated 30.8pL/169.3pL(12.3%)
1ppm 10 minutes Room Temperature 136.12pL/170.74pL (79.7%)
Untreated 45.25pL/169.20pL (26.7%)
1ppm 20 minutes Room Temperature 152.76pL/159.26pL (95.9%)
Untreated 82.82pL/155.17pL (53.4%)
1ppm 60 minutes Room Temperature 155.04pL/174.03pL (89.6%)
Untreated 105.52pL/172.95pL (61.0%)
-23-258550/3
Claims (15)
1. A method for cleaning a water filtration membrane wherein the water filtration membrane comprises a biofilm by i) contacting the membrane with a liquid cleaning composition comprising a DNase and optionally further comprising one or more non-DNase enzyme(s) selected from the group consisting of arabinase, carbohydrase, cellulase, cutinase, galactanase, lipase, mannanase, oxidoreductase; pectinase, pectate lyase, perhydrolase, protease, and xylanase; or by ii) contacting the membrane with a liquid cleaning composition comprising a DNase and optionally separately contacting the membrane with one or more non-DNase enzyme(s) selected from the group consisting of amylase, arabinase, carbohydrase, cellulase, cutinase, galactanase, lipase, mannanase, oxidoreductase; pectinase, pectate lyase, perhydrolase, protease, and xylanase.
2. The method of claim 1, wherein the cleaning process is Cleaning-In-Place (CIP) or Cleaning- Out-of-Place (COP).
3. The method of claim 1 or 2, wherein the water flow rate through the membrane (flux) is improved.
4. The method of any one of claims 1 to 3, which is preceded or followed by contacting the membrane with a biocidal composition.
5. The method of claim 1, wherein the biofilm comprises one or more bacteria selected from the group consisting of Acinetobacter, Bacillus, Comamonas, Escherichia, Pseudomonas, and Sphingomonas species.
6. The method of claim 1, wherein the biofilm comprises one or more bacteria selected from the group consisting of Acinetobacter calcoaceticus, Bacillus amyloliquefaciens SC100, Bacillus 24 258550/3 amyloliquefaciens SC168, Comamonas denitrificans, Escherichia coli K-12, Pseudomonas aeruginosa, and Sphingomonas mucosissima.
7. The method of claim 1, wherein the biofilm comprises Pseudomonas aeruginosa.
8. The method of any one of claims 1 to 7, wherein the pH of the cleaning composition is 6-9; preferably the pH is about neutral.
9. The method of any one of claims 1 to 8, wherein the membrane is contacted with the cleaning composition at a temperature between 30-100°C.
10. The method of any one of claims 1 to 9, wherein the non-DNase enzyme is a protease.
11. The method of any one of claims 1 to 10, wherein the DNase is an Aspergillus oryzae DNase.
12. The method of any one of claims 1 to 11, wherein the DNase is a polypeptide having DNase activity, which comprises or consists of an amino acid sequence with at least 80% sequence identity to the amino acid sequence shown as SEQ ID NO: 1.
13. The method of any one of claims 1 to 12, wherein the DNase is a polypeptide having DNase activity, which comprises or consists of the amino acid sequence shown as SEQ ID NO: 1.
14. The method of any one of claims 1 to 10, wherein the DNase is a Bacillus licheniformis DNase. 25 258550/3
15. The method of any one of claims 1 to 10, wherein the DNase is a polypeptide having DNase activity, which comprises or consists of an amino acid sequence with at least 80% sequence identity to the amino acid sequence shown as SEQ ID NO: 2. 26
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PCT/US2016/056955 WO2017066510A1 (en) | 2015-10-14 | 2016-10-14 | Cleaning of water filtration membranes |
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Citations (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2006031554A2 (en) * | 2004-09-10 | 2006-03-23 | Novozymes North America, Inc. | Methods for preventing, removing, reducing, or disrupting biofilm |
WO2009061380A2 (en) * | 2007-11-05 | 2009-05-14 | Danisco Us Inc., Genencor Division | VARIANTS OF BACILLUS sp. TS-23 ALPHA-AMYLASE WITH ALTERED PROPERTIES |
WO2014087011A1 (en) * | 2012-12-07 | 2014-06-12 | Novozymes A/S | Preventing adhesion of bacteria |
-
2018
- 2018-04-08 IL IL258550A patent/IL258550B/en unknown
- 2018-04-12 SA SA518391349A patent/SA518391349B1/en unknown
Patent Citations (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2006031554A2 (en) * | 2004-09-10 | 2006-03-23 | Novozymes North America, Inc. | Methods for preventing, removing, reducing, or disrupting biofilm |
WO2009061380A2 (en) * | 2007-11-05 | 2009-05-14 | Danisco Us Inc., Genencor Division | VARIANTS OF BACILLUS sp. TS-23 ALPHA-AMYLASE WITH ALTERED PROPERTIES |
WO2014087011A1 (en) * | 2012-12-07 | 2014-06-12 | Novozymes A/S | Preventing adhesion of bacteria |
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