REVIEW

The Role of Enzymes

in Modern Detergency
Hans Sejr Olsen* and Per Falholt
Enzyme Development & Application, Enzyme Business, Novo Nordisk A/S, DK-2880 Bagsvaerd, Denmark

ABSTRACT: Enzymes have effectively assisted the develop- lulases, the foundations were already laid in 1913 for the
ment and improvement of modern household and industrial de- commercial use of enzymes that continues to be important
tergents. The major classes of detergent enzymes—proteases, li- today.
pases, amylases, and cellulases—each provide specific benefits Today the most widely used industrial enzymes are hy-
for application in laundry and automatic dishwashing. Histori- drolases, which remove soils based on proteins, lipids, and
cally, proteases were first to be used extensively in laundry de-
polysaccharides. Cellulolytic enzymes are another class of
tergents. In addition to raising the level of cleaning, they have
hydrolases that provide fabric care through selective reac-
also provided environmental benefits by reducing energy con-
sumption through shorter washing times, lower washing tem- tions not previously possible on fabrics. Research is cur-
peratures, and reduced water consumption. Today proteases are rently underway into the possibility of using redox en-
joined by lipases and amylases in improving detergent efficacy zymes—oxidases or peroxidases—for bleaching colored
especially for household laundering at lower temperatures and, components (2).
in industrial cleaning operations, at lower pH levels. Cellulases To support the 18–19-million -ton global annual market
contribute to overall fabric care by rejuvenating or maintaining for laundry and dishwashing detergents (3), the world-
the new appearance of washed garments. Enzymes are pro- wide consumption of detergent enzymes amounted to
duced by fermentation technologies that utilize renewable re- ca. U.S. $500 million in 1995 (4). The principal producers
sources. of enzymes are Novo Nordisk A/S (headquartered in
JSD 1, 555–567 (1998).
Bagsvaerd, Denmark) and Genencor International Inc.
(headquartered in Rochester, NY), serving the market with
KEY WORDS: Amylases, automatic dishwashing, cellulases,
enzymes in detergents, industrial cleaning, laundry, lipases, more than 90% of the total volume of detergent enzyme
proteases, storage stability, wash performance. products.

THE ROLES PLAYED BY ENZYMES IN DETERGENTS

The field of enzymology is an important branch of bio-
process technology. Developments in submerged fermen- Over the years, enzymes have been an important factor in
tation processes and genetic engineering have resulted in the development and improvement of detergent products.
great progress in the manufacture of industrial enzymes. In laundering, dishwashing, and in industrial and institu-
Enzymes find use as functional ingredients in deter- tional (I&I) cleaning, they have contributed to shortening
gents and contribute to cleaning of laundry and dishes in washing times, reducing energy and water consumption
an efficient, environmentally mild, and energy-saving by lowering washing temperatures, providing environ-
manner. This largest of all industrial enzyme applications mentally friendlier wash-water effluents, lowering pH lev-
began slowly in the early 1930s, based on Röhm’s 1913 els in wash liquors, and providing fabric care. Enzymes
patent of the use of pancreatic enzymes in presoak solu- themselves are environmentally attractive since they are
tions (1). Pancreatic enzymes include protease (trypsin and derived from renewable sources. They are also highly
chymotrypsin), carboxypeptidase, α-amylase, lactase, su- space-efficient and are thus of particular advantage in con-
crase, maltase, and lipase. Thus, with the exception of cel- centrated detergent formulations. Developments in genetic
and protein engineering have contributed to long-needed
*To whom correspondence should be addressed at Enzyme Develop-
ment & Application, Enzyme Business, Novo Nordisk A/S, Novo improvements in the stability, economy, specificity, and
Allé, DK-2880 Bagsvaerd, Denmark. E-mail: [email protected] overall potential of industrial enzyme products.

Copyright © 1998 by AOCS Press Journal of Surfactants and Detergents, Vol. 1, No. 4 (October 1998) 555
556 REVIEW

TABLE 1
Consumption During the Washing of Fabrics (European data) (Refs. 5–9)
Consumption/kg 1943–44 1965 1975 1985 1990–95
dry fabric (manual) (machine) (machine) (machine) (machine)

Chemicals (g)
Conventional 109 ca. 50 ca. 50 38–48 32
Compact 0 0 0 0 16–21
Water (L) 31 37 31 21 10–12
Energy for heating (kWh) 1.45 0.6 0.4 0.25 0.2
(to heat water)
Energy for motors
and pumps (kWh) 0 0.4 0.2 0.15 0.1

Detergent mechanism based on enzymes. From an enzyme zymes were introduced by Lion Corp. in 1979, and in the
point of view, detergents on the international market con- United States the resurrection of enzymes began in the
tain principal ingredients that operate by almost identical 1980s after health problems with factory workers had ar-
detergency mechanisms. Soil and stains are removed by rested the initial introduction in 1971 (3).
mechanical action assisted by surfactants, builders, and en- Throughout the period covered in Table 1, the develop-
zymes. Alkaline proteases, amylases, or lipases in heavy- ment of new ingredient combinations (builders and bleach
duty detergents hydrolyze and solubilize substrate soil at- systems) and machines was optimized in conjunction with
tached to fabrics or hard surfaces (e.g., dishes). Cellulases the improved cleaning properties offered by enzyme sys-
clean by hydrolysis of glycosidic bonds which removes tems. At the same time, the development of new enzyme
particulate soils attached to cotton microfibers. Key effects systems was influenced by the new compositions of deter-
of cellulases are to soften and improve the color brightness gent mixes.
of worn textile surfaces. Surfactants lower the surface ten- A comparison of calculated cost distributions for
sion at interfaces and enhance the repulsive force between washes from 1943 (5) and 1997 (10) reflects many dramatic
the original soil, enzymatically degraded soil, and the fab- changes in society and also indicates the benefits of new
ric. Builders act to chelate, precipitate or ion-exchange cal- developments. Table 2 shows the annual cost of washing
cium and magnesium salts, to provide alkalinity, to pre- on a family basis for the 2 yr. Interestingly, the proportion
vent soil redeposition, to provide buffering capacity, and of the costs attributable to the detergents has not changed.
to inhibit corrosion. The energy part of the washing costs has been reduced sig-
Laundry. Over the past 35 yr the role of enzymes in de- nificantly, but the costs for water have increased consider-
tergents has changed from one of a minor additive to be- ably due to the increasing shortage of clean water and
coming a key ingredient. The largest application in which heavy taxation of environmental resources.
the use of enzymes has grown is household laundry. Table 2 explains, to some degree, the developments seen
Before the introduction of modern detergent formula- in laundry technology over the years from 1943 to 1997.
tions, soap and sodium carbonate were the principal deter- New detergent compositions and enzymes made possible
gent constituents, and the cleaning effect in laundering was a lowering of washing temperatures and hence of energy
dependent mainly on mechanical action. Table 1 shows a consumption. Thus it is easy to understand the current
comparison of the consumption of chemicals, water, and trend of washing machine design heading in the direction
energy over the last 50 yr. The advent of washing machines of reducing water consumption.
brought a considerable reduction in the consumption of Dishwashing. Automatic dishwashing machines have
chemicals over that in manual washing. In the period of been in use for almost as long as washing machines but not
1967–1970 (5), a reduction in energy consumption of about as widely. Today, automatic dishwashers are found in
40% was brought about by the successful introduction of 50–60% of households in the United States, West Germany,
enzymatic detergents. In 1969, at least 50% of European de-
tergents were offering “biological attack” (proteolytic en-
zymes), which resulted in the energy-savings shown (3). TABLE 2
The move away from 95°C washes to 60°C washes, and Yearly Costs (DKK—Danish crowns) and Cost Distribution
for Washing in a Danish Model Family
later to 40°C washes became more common. Furthermore,
prewashes and overnight soaking could be eliminated. The 1943 (Ref. 5) 1997 (Ref. 10)
additional energy reduction resulted from a combination of Component DKK % distribution DKK % distribution
more diverse enzyme ingredients (cellulases, amylases, and Electricity 14.5 12 368 27
lipases), more mechanically efficient washing machines (jet- Fuel 47 39
spray and cascade systems), better-managed machine elec- Water 9.7 8 415 31
tronics, and compact detergents. In Japan, proteolytic en- Detergents 48.8 41 565 42

Journal of Surfactants and Detergents, Vol. 1, No. 4 (October 1998)

 REVIEW 557

Norway, Sweden, and Denmark, and in ca. 30% of the from 2.8 to 1.5 kWh (13). Chlorine or hypochlorite-releas-
households in France and Italy, while in other European ing bleach compounds (e.g., chloroisocyanurates) have
countries like The Netherlands and Spain distribution is been deemed essential to remove certain stains, e.g., tea
less than 20% (4,11). In Japan, automatic dishwashers are stains in cups.
found in fewer than 5% of domestic households (4). In the United States, most of the ADD market is still
In manual dishwashing, the removal of food residues is based on the traditional chlorine-type ADD although some
mainly dependent on surfactants and mechanical energy. enzymatic products have been available for a number of
In machine dishwashing, cleaning performance is depen- years. Efforts to introduce enzymatic low-alkaline ADD in
dent on the complete detergent, the temperature, and me- the United States have been ineffective to date. Unlike Eu-
chanical energy. Mechanical energy is delivered by the ropean dishwashing machines, their U.S. counterparts lack
pump, which circulates the wash water. The chemical con- both an ion exchange water softener and a built-in water
tribution of the detergents must therefore be high. The heater. This places the main burden of cleaning on the de-
most severe cleaning problem is encountered when food tergent and does not favor mild detergent formulations (3).
items coat tableware—porcelain plates, cups, cutlery, and As with laundry detergent formulations, the introduc-
glasses—with a film. Food may be dried-on, baked-on, or tion of new ADD with enzymes (amylases and proteases)
even burned-on and forms complex compounds from car- has made it possible to replace harsh chemicals like
bohydrates, proteins, and lipids. Vegetable pigments or tea bleaches and alkalis. Since hypochlorite in low concentra-
scum consisting of Ca-tannin compounds—in brewed tions will degrade enzymes, peroxygen/activator bleach
tea—also demand considerable attention. In addition, systems based on the combination of sodium perborate or
combinations of polyphenols and carbohydrates—from percarbonate with activators (TAED, etc.) that oxidize pol-
coffee—are considered difficult. Thus, dishwashing is con- yaromatics from fruit, coffee, tea, wine, and vegetables are
sidered successful if tea and coffee stains have been com- used instead. At the reduced pH in chlorine-free ADD, the
pletely removed. Complex compounds based on proteins, hydrolytic swelling needed to remove food residues from
polysaccharides, and lipids require hydrolytic treatment surfaces is supported by the action of respectively starch-
that causes swelling and degradation of the soil and thus and protein-degrading enzymes. These systems are widely
releases it from the surface. Bleach systems that oxidize used in Europe in compact ADD powders and tablets.
polyaromatics are also needed to remove fruit, coffee, tea, I&I laundry. The application of enzymes in I&I laundry,
wine, and vegetable stains. institutional dishwash, and cleaning in the food industry
The combination of short washing times, high tempera- has grown substantially (14).
ture, and the need for avoiding attack on dishware calls for Textiles treated under I&I laundry conditions include
chemicals with special properties. Highly alkaline-oxidiz- hospital bed linen, coats and uniforms, workers overalls
ing chemicals are needed to ensure the hydrolysis and ox- and white coats from slaughterhouses, and tablecloths and
idative hydrolysis of food compounds and to disperse napkins from hotels and restaurants.
them into the washing liquor without foaming. Effective Machines used can be washer–extractors having capaci-
binding of hardness ions to prevent lime deposits and pro- ties of 120 kg dry cloth per load or they can be tunnel
tective agents for glass and on-glaze decorations are also washers with several compartments and capacities of 550
needed. kg per hour.
Before enzyme products were introduced in Europe in In washer–extractors a prewash at low temperature
1990, automatic dishwashing detergents (ADD) were for- (e.g., 35°C) and pH = 11 permit the use of enzymes within
mulated on the basis of strongly alkaline, strong complex- a time period of 8–10 min. The main wash is usually car-
ing agents [sodium triphosphate (20–30%), sodium meta- ried out for 15 min at 85ºC, pH = 11.5. In order to permit
silicate (40–70%), and sodium carbonate, 0–10%)] oxidiz- enzymes to work, this temperature can be reduced to 60°C.
ing compounds, usually di- or trichloroisocyanurate Usually, the last bleach process is a treatment with active
(1–3%), and nonionic surfactants (0–2%; pH in 1% solution chlorine in cold water.
= 12.0–12.5; typical dosages = 60–80 g per wash. In tunnel washers the holding time in each compart-
At a pH of 12–13, rinsing to remove chemicals must be ment is 4–5 min. During prerinse and first wash, enzymes
very effective. This is the reason why large volumes of may be added followed by bleaching steps with chlorine
water were used in dishwashers in the past. Although or hydrogen peroxide.
glass is chemically reasonably resistant, its silicate struc- Institutional dishwashing. In a traditional institutional
ture is destroyed by hydrolytic cleavage of Si–O–Si bonds dishwashing process, the dishware is placed vertically in
under highly alkaline conditions, resulting in a cloudy sur- trays and then washed with 60°C wash water including
face appearance (12). Disilicates were tested and found to 0.5–5 g/L of a detergent containing sodium hydroxide,
be effective in preventing glass weight loss (12). potassium hydroxide, phosphonate, polycarboxylate, and
Over the last 20 yr, water consumption was reduced sodium EDTA. In the absence of enzyme, the pH normally
from 60 to 15 L per dishwashing cycle, detergent consump- is 13–14. The washing process with detergent normally
tion from 80 to 30 g per wash, and energy consumption lasts 60–90 s followed by a rinsing process lasting 20 s at

Journal of Surfactants and Detergents, Vol. 1, No. 4 (October 1998)

558 REVIEW

75–85°C. The dishware is then dried either in the machine for cleaning spiral-wound ultrafiltration modules that are
or outside. very sensitive to harsh chemicals and are generally diffi-
In large institutional dishwashers, trays are placed on a cult to clean with traditional cleaning products (18).
conveyor belt which pulls the dishware through the above The combination of protease and lipase in a pure en-
treatments in separate chambers. zyme cleaning process of plate heat exchangers used for
In institutional dishwashing processes, the use of en- high pasteurization of sweet milk surfactants (soaps) leads
zymes provides cleaning effects as in household usage. to the in situ formation of emulsifiers and foaming agents
With a low-alkaline detergent system without bleach, a from the fat and proteins. This is illustrated schematically
preferable temperature range is 45–65°C. Very often a two- in Figure 1.
component liquid detergent system is used. One compo- Regulatory aspects and quality assurance of detergent en-
nent is a liquid detergent containing, e.g., sodium hydrox- zymes. In most countries, the regulatory status, classifica-
ide, a calcium binder like phosphonate, amphoteric or non- tion, and labeling are determined in accordance with exist-
ionic surfactant, and polycarboxylate. Typical dosages may ing schemes for chemicals. Many enzyme types are listed
be about 2–4 g/L of wash liquid, resulting in a pH of 8.5–9 on chemical inventories, e.g., EINECS in the European
of the cleaning solution. The second component may be Union and TSCA in the United States. In some cases, en-
starch- and protein-degrading enzymes. The enzymes are zymes are considered natural substances exempt from list-
dosed from a separate container directly into the washing ing or are regulated by specific biotechnology products
chamber. legislation.
Cleaning in the food industry. For many years, proteases The Association of Manufacturers of Fermentation En-
have been used as minor functional ingredients in formu- zyme Products has defined Good Manufacturing Practice
lated detergent systems for cleaning reverse-osmosis mem- for microbial food enzymes. This practice is generally fol-
branes. The requirements and dosages of enzymes are sim- lowed for detergent enzymes also. The most important ele-
ilar to those for laundry detergents and ADD. Enzymes are ment is to ensure a pure culture of the production organism.
now also used for cleaning processes in the dairy (15) and Commercial enzyme products are usually formulated
in the brewing industries for cleaning microfiltration and in aqueous solutions or processed to dry nondusting gran-
ultrafiltration membranes (16). Enzymatic cleaning is also ulates.
being used for membranes used in fruit juice processing Safety. Like many other proteins foreign to the human
(17). Both organic and inorganic soils are removed by body, enzymes are potential inhalation allergens. Inhala-
cleaning-in-place (CIP). In CIP plants, the cleaning tion of even small concentrations of a foreign protein in the
medium is normally 0.5–1% NaOH (± surfactants and form of dust or aerosols can stimulate the body’s immune
EDTA) at 75–85°C followed by rinsing and 0.5–1% HNO3 system to produce antibodies. In some individuals, in-
(± surfactant) or “acid sulfate.” creased concentrations of antibody–enzyme protein com-
In dairies, the most difficult soil to remove from hard plexes can trigger increased concentrations of histamine.
surfaces is “burnt-on milk” from the fouling of heating sur- The latter compounds can cause hay fever-like symptoms
faces, e.g., on heat exchangers or evaporator tubes. This such as watery eyes, running nose, and a sore throat. When
substrate is a kind of gel assumed to be a complex formed exposure ceases, these effects cease also.
by Maillard-type reactions between protein, lactose, and Enzymes must be inhaled in order to present a risk of
fat. Milkstone (a calcium phosphate protein complex) may causing sensitization, which may lead to an allergic reac-
also be involved. For this application, an enzymatic CIP tion. Working environments where enzymes are used are
system based on protease and lipase has proven effective therefore subjected to extensive monitoring to confirm that

FIG. 1. In situ cleaning effects of protease + lipase.

Journal of Surfactants and Detergents, Vol. 1, No. 4 (October 1998)

 REVIEW 559

threshold limit values (TLV) for atmospheric enzymes are source of origin (animal, plant, microbial), their catalytic
not exceeded. In many countries, the TLV for enzymes is action (endopeptidase or exopeptidase), and the nature of
based on the proteolytic enzyme subtilisin and is stated the catalytic site (active site). They are characterized by
0.00006 mg/cu.m of pure crystalline subtilisin in air (19). common names and trade names, typical pH ranges, and
Successful experience in controlling enzyme exposure preferential specificity. Based on a comparison of active
and protecting workers’ health has been reported by Proc- sites, catalytic residues, and three-dimensional structures,
ter & Gamble (20). Already in 1971 a National Research four major protease families are recognized: serine, thiol,
Council report had concluded that consumers did not de- aspartic, and metalloproteases. The serine protease family
velop respiratory allergies from the use of enzyme-contain- contains two subgroups: chymotrypsin-like and subtilisin-
ing laundry products (21). Further studies over the years like. The latter is the most important group for detergent
have confirmed that enzyme-containing laundry and dish- applications.
washing detergents are safe for consumer use. Examples of thiol (or cysteine) proteases are plant pro-
teases from papain latex (Papain), from pineapple stem
(Bromelain), and from fig latex (Ficin). These enzymes are
DETERGENT ENZYMES IN USE
not used in detergents.
Manufacture. The production of microbial enzymes repre- The characteristics of a number of commercial enzyme
sents a significant part of today’s industrial biotechnology. products used for industrial detergents or food protein
The most significant industrial enzyme products are pro- products are shown in Table 3. The enzymes Alcalase®, Es-
duced by aerobic batch or continuous fermentation in fer- perase®, Savinase®, and Trypsin are serine proteases, while
mentors with volumes ranging from 20–1,000 m3. Fermen- the enzyme called Neutrase® is a metalloprotease with
tation processes are carried out on sterilized nutrients Zn2+ in its active site. Furthermore, this enzyme is stabi-
based on renewable raw materials like corn starch, various lized by Ca2+. Durazym® and Everlase™ are protein-engi-
sugars, and soy grits in the presence of various added salts neered variants of Savinase® and mainly used in deter-
to provide basic elements. gents containing bleach. The development of selected pro-
Most industrial enzymes are secreted from selected mi- teases has been guided by factors like oxidation stability,
croorganisms into the fermentation medium in order to temperature optima, autoproteolysis, denaturation, water
break down the carbon and the nitrogen source. hardness, alkalinity, and economy. Thus detergent pro-
The enzyme activity is harvested from the fermentation teases have been categorized in types for uses within alka-
broth by removing insoluble products and the biomass line, high alkaline, high bleach stability, and cold-water ap-
produced, usually by filtration or centrifugation. Enzyme plications (3). Table 4 is an attempt to show the status of to-
in the solution is then concentrated by evaporation, mem- day’s merchant market. Practical selections of proteases for
brane processes, and crystallization. Depending on the specific detergents are made on the basis of performance
specific application, the enzyme product is posttreated. For tests and stability tests carried out under certain standard
solid products, such as detergent powders, nondusting conditions chosen in conjunction with formulators.
granulated enzyme formulations are manufactured (22). Protein hydrolysis in detergency. Proteases catalyze the
For use in liquid detergents, liquid or encapsulated formu- hydrolytic cleavage of the peptide chain, as shown for re-
lations are easy to handle and dose in a factory. actants and products in Scheme 1.
Proteases. Proteases are the most widely used enzymes.
In laundry detergents, protein stains such as grass, blood,
egg, and human sweat are removed through proteolysis.
In ADD, proteases secure the removal of proteinaceous
food films, which are a particular problem with glassware
and cutlery. Proteases are classified according to their SCHEME 1

TABLE 3
Some Commercial Proteolytic Enzymes (Novo Nordisk A/S, Bagsvaerd, Denmark)
Product names Microorganism or other origin State of product Practical application range (pH) Practical application range (°C)

Alcalase® Bacillus spp. Liquid or granulate 6–10 10–80

Esperase® Bacillus spp. Liquid or granulate 7–12 10–80
Everlase™ Bacillus GMO (engineered) Liquid or granulate 8–11 15–80
Savinase® Bacillus GMO Liquid or granulate 8–11 15–75
Durazym® Bacillus spp. GMO (engineered) Liquid or granulate 8–11 15–70
Neutrase® Bacillus spp. Liquid or granulate 6–8 10–65
Protamex™ Bacillus spp. Microgranulate 6–8 10–65
Flavourzyme™ Aspergillus spp. Liquid or granulate 4–8 10–55
Trypsin Pancreas Granulate 7–9 10–55

Journal of Surfactants and Detergents, Vol. 1, No. 4 (October 1998)

560 REVIEW

TABLE 4
Merchant Market Detergent Proteasesa
Producer Alkaline High alkaline High bleach stability Cold water

Genencor International Maxatase Purafect Maxapem Properase

(Rochester, NY) Optimase Maxacal Purafect OxP
Opticlean Opticlean plus
Novo Nordisk A/S Alcalase® Esperase® Durazym® New developments
Savinase® Everlase
a
See Table 3 for other producer’s location.

The most important parameters for the hydrolysis reac- Some kinetic aspects of protein hydrolysis. The pH-stat tech-
tion are surface-available substrate S (percentage protein nique was utilized to study the proteolytic degradation of
for the reaction), E/S (enzyme–substrate ratio in activity hemoglobin in solution and to understand the reaction
units per kg protein), pH, reaction time, and temperature. mechanism (24).
Together with the specificity and properties of the enzyme In the discussions of initial proteolysis, Linderstrøm-
itself, these parameters are responsible for the course of re- Lang (25) referred to the reaction of a protease on native
action on a given protein stain. The quantitative criterion hemoglobin molecules as “one-by-one,” indicating that a
for a proteolytic reaction is the degree of hydrolysis (DH) particular protease molecule degraded one substrate mol-
or hydrolytic activity (HA). DH is calculated by determin- ecule at a time. No appreciable amounts of intermediary
ing the number of peptide bonds cleaved and the total products were present. The reaction mixture consisted of
number of peptide bonds in the intact protein molecule. native proteins and end-products only.
HA is here defined as milliequivalent (meq) NaOH con- In another reaction mechanism referred to, the native
sumed in a pH-stat/mass of substrate for a given time. protein molecules were rapidly converted into intermedi-
The DH of enzyme-treated protein is proportional to the ary forms, which degraded more slowly to end-products
molecular size of degraded proteins. Therefore measure- (“Zipper reaction”).
ment of the DH is important. Only in this way is it possi-
ble to optimize the reactions and to study the mechanisms
in detail (23). For laundry or dishwashing applications, no
estimates of this parameter have been reported, presum-
ably because the substrate concentration is very low and
difficult to measure.
However, when sufficient substrate is available, the rel-
atively simple pH-stat technique can be applied directly as
an analytical tool during the reaction. For the standard test
textile swatches, EMPA 116 (cotton) or EMPA 117 (cotton +
polyester), containing blood, milk, and carbon black, Fig-
ures 2 and 3 show examples of hydrolytic activity as a
function of time. Different detergents, enzyme dosages,
and temperatures were tested. The protease is added in
dosages relevant to the application, but Figures 2 or 3 do
not, as such, represent state of the art with regard to maxi-
mal performance under Japanese or European washing
conditions. The reaction is carried out at fixed temperature
in a 1-L reaction vessel held under an N2 atmosphere with
sufficient stirring to keep the swatches floating. The pH-
stat technique can be used as a quick and nonlabor-inten-
sive method for evaluating the effects of some washing pa-
rameters. When a simultaneous solubilization of protein
occurs as an effect of the protein hydrolysis, measurements FIG. 2. Example of a pH-stat trial on EMPA 116 swatches—Japanese de-
of soluble nitrogen can be made. After rinsing and drying, tergent. HA, hydrolytic activity. Reaction conditions: temperature,
the reflectance can be measured (see Washing performance 15°C; concentration, 6.5 g EMPA 116 in 800 g wash; detergent, 0.67
evaluations—laundry section). These trials cannot fully g/L Japanese detergent, pH 10.5; water hardness, 3° dH (German de-
grees hardness); enzyme, Savinase 12 T; activity dosages, 0–0.20
substitute for machine washing trials because mechanical kNPU/L, where kNPU is the proteolytic activity in kilo Novo Protease
effects and temperature profiles are different, and the ef- Units measured as the hydrolysis rate of dimethyl casein and compared
fects of rinsing are not measured. with a standard reference protease preparation.

Journal of Surfactants and Detergents, Vol. 1, No. 4 (October 1998)

 REVIEW 561

ing emulsification, foaming properties, reducing surface

tension and redeposition of degraded protein material.
Protease inhibitors. In detergent powders where pro-
teases are granulated and encapsulated, all enzymes are
stable in the absence of water. In liquid detergents [more
than 40% of the U.S. market (4)], the aqueous environment
is inimical to enzyme stability and inhibitors are necessary
to prevent autodigestion of the proteases themselves or to
prevent the degrading of other enzymes. Polyols such as
glycerin, propylene glycol, and polyethylene glycol have
been used in combination with boric acid to inhibit pro-
tease activity and reduce autodigestion (4,26). Borate
works as a reversible protease inhibitor by interacting with
groups in the active site of the enzymes. Once in the wash
water, these inhibitors are diluted and enzyme activity in-
creases (4). Protein hydrolysate products and amino acids
are also known to be able to stabilize proteases by binding
FIG. 3. Example of a pH-stat trial on EMPA 117 swatches—European to the active site of the enzyme (27).
detergent. Reaction conditions: temperature, 40°C; concentration, 6.5 g
Amylases. Native starch is only slowly degraded by
EMPA 117 in 800 g wash; detergent, 4 g/L European (bleach-contain-
ing) detergent, pH 9.8; water hardness, 18° dH; enzyme, Savinase 12 T; α-amylases. Gelatinization and swelling are needed to
activity dosage, 0–0.3 kNPU/L. See Figure 2 for abbreviations. make the starch susceptible to enzymatic breakdown. For
most foods, various degrees of gelatinization result from
cooking. Therefore, in detergents for laundry and auto-
In Figure 4, the slight bending in the hydrolysis curve matic dishwashing, amylases facilitate the removal of
at low DH values indicates a thorough initial degradation starch-containing stains, e.g., pasta, potato, gravy, choco-
to small peptides (a zero-order reaction). This degradation late, and baby food. Amylases also prevent swollen starch
is close to the ideal one-by-one reaction. Hydrolysis pro- from adhering to the surface of laundry and dishes that
ceeds rapidly with little decrease in velocity to the stage may otherwise act as a glue for particulate soiling. Com-
where the substrate concentration is so low that first-order plexes or reaction products between protein, starch,
kinetics take over. and/or fat are usually found in prepared foods. In such
Proteolysis of proteins improves cleaning of fibers and cases, enzyme synergy effects make it possible to remove
hard surfaces by increasing the solubility of soils, promot- soil even more efficiently than with single enzyme systems
(22). If the food processes have not exceeded the starch
gelatinization temperature, the starch may be in the form
of partly or nongelatinized granules, or it may be partly
retrograded. Such starch may be amorphous and usually
difficult to remove from surfaces without cooking. Pres-
ence of an amylase renders cooking superfluous.
Industrially important α-amylases are made from Bacil-
lus species and Aspergillus species. Table 5 shows some ap-
plication conditions and a few comparative characteristics
for amylases.
During enzymatic liquefaction, the α-1,4-linkages in
starch (amylose and amylopectin) are hydrolyzed at ran-
dom, thus reducing the viscosity of the gelatinized starch
and increasing the solubility of attached starch, which is
converted to water-soluble dextrins and oligosaccharides.
Therefore, Termamyl®, Duramyl®, and BAN are often re-
ferred to as “liquefying amylases.” Fungamyl is a fungal
exo-amylase, which also hydrolyzes the α-1,4-linkages in
liquefied starch. A prolonged reaction results in the forma-
FIG. 4. The pH-stat hydrolysis curve for a hemoglobin in an automatic tion of large amounts of maltose. Fungamyl is relevant in
dishwashing detergent with Savinase. DH, degree of hydrolysis. Reac- detergents for use at low pH levels in industrial cleaning
tion conditions: temperature, 50°C; concentration, 0.5% protein (N ×
6.25); detergent, 4.5 g/L European dishwashing detergent (pH 10.3); en-
tasks. Duramyl® was developed to achieve increased oxi-
zyme, Savinase 16 L; activity dosage, 0.75 kNPU/L. See Figure 2 for dation stability in the humid environment of detergent
other abbreviation. powders containing moisture (28). In practice, amylases

Journal of Surfactants and Detergents, Vol. 1, No. 4 (October 1998)

562 REVIEW

TABLE 5
Some Commercial Starch-Degrading Enzymesa
Product name Microorganisms State of product Practical application range (pH) Practical application range (°C)

Termamyl® Bacillus spp. (GMO) Liquid or granulate 6–11 25–100

Duramyl® Bacillus spp. (GMO) Liquid or granulate 6–10 25–100
BAN Bacillus spp. Liquid or granulate 5–8 15–90
Fungamyl® Aspergillus spp. Liquid or granulate 4–7 15–60
a
GMO, genetically modified organism.

for detergent applications are selected on the basis of per- tending from either the C-terminal or the N-terminal of the
formance and stability tests in specific detergents. core. The nature of the core determines catalytic properties
Gelatinized starch may form a film on fabric that can re- such as endo-activity vs. exo-activity, substrate specificity,
sult in an increased pick-up of particulate soil after wash- and the type of reaction products that are formed. The
ing (29). Starch stains, combined with particulate soiling, presence of a CBD is of particular importance for binding
are more difficult to remove than starch alone. As a result on insoluble and crystalline cellulose and for hydrolytic ef-
white laundry items turn increasingly gray after repeated fects (31). Both EG and CBH can contain linkers and cellu-
wash cycles, an effect that has been demonstrated by lose-binding domains. The cellulase products in Table 6 are
adding about 0.5 g starch/kg cotton fabric (30). Starches composed as follows: Celluzyme®: A complex mixture of
may react differently depending on their amylose content, at least seven cellulases: Humicola CBH I, CBH II, EG I, EG
which is thought to be the film-forming component of the II, EG III, EG V, and EG VI; and Carezyme®: mono-compo-
starch. Film formation is favored under European condi- nent Humicola. These cellulases can be used in bleach-con-
tions where the temperature may be closest to the starch taining detergents and are used in a number of “Color”
gelatinization temperature. In laundry detergents, amy- and compact detergent powders.
lases may maintain or even contribute to increased whiten- Extremely high dosages of “color clarification cellu-
ing of dingy fabrics (29) and inhibit the graying of white lases” can inflict fabric damage in some cotton products
fabrics resulting from a combination of starch and particu- after repeated launderings. Damage may appear as loss of
late soiling (29,30). fabric strength and excessive softening of the mechanically
Cellulases. Cellulases cleave β-1,4-glucosidic bonds in exposed parts of laundry items, such as hems and edges.
cellulose and operate directly on the natural cotton fibers These effects may be eliminated by balancing the dosage
or cotton/flax blends and on the cellulose portion in syn- to manage the desired benefits. Application tests include
thetic fibers. This enzyme class is divided into endo-cellu- small-scale laundering in Terg-O-tometers and full-scale
lases (endo-glucanase = EG) and exo-cellulases (cellobio- multicycle laundering in commercial washing machines
hydrolase = CBH). Table 6 shows some application condi- (32).
tions and a few comparative characteristics for cellulases. Lipases. Because of their strong hydrophobicity, fats and
Celluzyme® and Carezyme® (Novo Nordisk) are “color oils (triglycerides) are difficult to remove from laundry at
clarification cellulases,” which are applied in detergents to low temperatures. Lipases hydrolyze triglyceride to more
make cotton fabrics regain and maintain clear colors, a hydrophilic mono- and diglycerides, free fatty acids, and
smooth surface, and softness. Cellulases provide these ef- glycerol. These hydrolysis products are all soluble in alka-
fects by shaving off the fuzz and pills of cotton fibrils that line conditions. At pH >8 the hydrolysis reaction may be
are generated on the fabric by normal wear and washing. favored by small amounts of free Ca ions due to the forma-
Cellulases are unique in providing these effects. tion of Ca soap, although lipases are effective also at low
The cellulase molecule is composed of up to three types free calcium levels.
of functionally different domains: the core, which is a large, The first commercial lipase for detergent application
spherical, catalytic domain; a linker, which is an elongated was Lipolase®, which was introduced in 1988 and used at
and flexible spacer; and a small spherical cellulose-binding once in Japanese detergents. Leading brands in the United
domain (CBD) (Fig. 5). States and Europe included lipase from 1990/91 (33). Lipo-
Some cellulases consist of a core domain only, others lase was first isolated from the fungus Humicola lanuginosa
comprise a core domain plus one or two linkers + CBD ex- from where the genetic coding for the lipase was trans-

TABLE 6
Cellulose-Degrading Enzymes for Detergentsa
Product name Microorganisms and enzyme type State of product Practical application range (pH) Practical application range (°C)

Celluzyme® Humicola spp. 5 EG + 2 CBH Liquid or granulate 4–10 25–70

Carezyme® Humicola spp. Liquid or granulate 5–10.5 25–70
a
EG, endo-glucanase; CBH, cellobiohydrolase. Product names from Novo Nordisk (Bagsvaerd, Denmark).

Journal of Surfactants and Detergents, Vol. 1, No. 4 (October 1998)

 REVIEW 563

ator, and hydrogen peroxide. The oxidized form of the me-

diator is the actual oxidizing agent. The dye (the colored
form) released from the fabric will be oxidized by the en-
zymatically oxidized mediator and become colorless. After
intensive screening of mediators, phenothiazine-10-propi-
onic acid was found useful under practical conditions in a
detergent system (2,38).

FACTORS AFFECTING ENZYME PERFORMANCE

IN HOUSEHOLD DETERGENTS
Enzyme performance is influenced by factors like deter-
FIG. 5. Cellulase domains. gent pH, ionic strength, wash temperature, washing time,
detergent composition, and mechanical handling. Both
performance and stability are influenced by some deter-
ferred to a host organism, Aspergillus spp., which is used gent surfactants and some bleach systems.
in production (34). In laundering, the effects of lipases are To illustrate the reaction conditions under which en-
seen only after several wash cycles (“multicycle wash per- zymes have to operate, Table 7 shows the components of
formance”). Further engineering was carried out to de- household detergents in very broad terms. Heavy-duty
velop first-wash effects in laundry usage: one of the 269 powdered detergent formulations may vary by geographic
amino acids in the Lipolase molecule was changed (in regions, with a tendency for higher levels of builder in
amino acid position no. 96 aspartic acid was substituted North America than in Europe and Japan (9). The ratio of
with leucine) to make the active site more hydrophobic so anionic to nonionic surfactant may also vary from region
that the affinity to a lipid contact zone on the textile sur- to region. In Japan, surfactant levels are higher than else-
face was improved. For cold washing, this effect was im- where (9). These variations between regions should be
proved considerably. The new lipase is produced by Novo seen in connection with the laundry practices common in
Nordisk as Lipolase Ultra. Further efforts are now being North America, Europe, and Japan. Table 8 shows in broad
made to develop “first-wash lipases” (35). terms some parameters in laundry conditions in the three
The delayed cleaning efficacy of lipases is attributed to principal geographical areas. Worth noting are the differ-
increased activity when the moisture concentration during ent washing machine types which dominate in each re-
the drying process is reduced. Thus, the activity of Lipo- gion. The ratio of wash liquor to fabric load, temperatures,
lase vs. the moisture content was found to be optimal at the usual dosage of detergents, and water hardness around
20–40% water content on fabric in a line-drying process at the world are significantly different—as are detergent com-
room temperature (36). positions. Most of these parameters may influence the de-
In testing lipase, three consecutive washing/drying cy- tergency of enzymes in a laundry washing machine or in
cles are usually employed. Washing temperature, time, automatic dishwashing. In laundry, particularly, the water
and Ca2+ level depend on the geographical area where the hardness (i.e., the calcium concentration) is of concern. Ca-
detergent is used. After washing and drying, the re- ions are known to stabilize some enzymes and destabilize
flectance of the colored, lard-containing test swatches is others. Usually sufficient amounts of enzymes are present
measured and the remaining lard may be extracted for in the powders to secure the desired detergency power.
quantitative determination. Performance. Evaluation of detergency is divided into
Redox enzymes. Redox enzymes, mainly peroxidases and two steps. (i) “Primary washing effects” (cleaning efficacy)
oxidases (including laccases), are being researched as po- refer to the removal of soil and stains after one wash. Test-
tential components of novel bleaching systems that—apart ing is carried out using either artificially soiled test fabrics
from utilizing less harsh chemicals than current bleaching or naturally soiled laundry. (ii) “Secondary washing ef-
systems—might even provide bleaching effects at low fects” (effects after repeat washing) refer to the detection
washing temperatures (down to 5–10°C). This would pro- of damage such as loss of tear strength, incrustations (ash
vide energy savings and also ensure less overall wear of residues), and graying. The latter evaluations are usually
garments being washed (37). based on 25 or 50 washes, including a control fabric.
Until now a peroxidase from Coprinus cinereus has been In the evaluation of dishwashing performance, “pri-
developed for use in dye transfer inhibition (DTI) in com- mary washing effects” are usually tested on soiled plates
bination with a mediator substance (2). This peroxidase or test pieces. “Secondary washing effects” in dishwashing
(Guardzyme™) can bleach dye material that is released refer mainly to the appearance of cloudiness and weight
from colored laundry items during the wash (38). loss (13).
Guardzyme™ is a hem-containing enzyme. The DTI (i) Washing performance evaluations—laundry. Laboratory
system consists of three components: the enzyme, a medi- washing performance evaluations are carried out at rele-

Journal of Surfactants and Detergents, Vol. 1, No. 4 (October 1998)

564 REVIEW

TABLE 7
General Composition of Detergents for Laundry and Dishwashinga
Contents (w/w%)
Raw
materials Functions in general Type of compounds Laundry Dishwash

Builders Washing alkalis Phosphates (sodium 30–60 10–40

Water softening triphosphate)
Buffer action Phosphonates
Stabilizers Na-citrates
Corrosion protection Zeolites
Soda
Silicates
Complex formers
(NTA, etc.)
Surfactants Emulsify particles Surfactants: 10–30 1–4
interfacial activity Anionics
Reduce surface Nonionics
tension Cationics
Soap Prevents overfoaming Soap 1–5 0
in the machine
Bleach—as Oxidize polyphenolics Na-perborates 0–25 4–15
peroxygen/activator from, e.g., fruits, coffee, Na-percarbonates
tea, wine, and Activators (TAED, etc.)
vegetables
Enzymes Removal of starch- Amylases 0.4–1 1–3
based stains
Removal of protein Proteases 0.4–2 0.5–2
complexes
Removal of fat stains Lipases 0.2–1 0.5
Textile color brightening, Cellulases 1–3 0
softening, soil
removal, whiteness
maintenance
Salt Process aid to promote Na-sulfate Balance to Balance to
flow and loading 100 100
properties (powders)
Perfume To add a pleasant Various esters 0.1–0.5 0–0.1
smell to the fabric
Other organics Graying inhibitors Carboxy methyl 0.5–5 0–1
cellulose,
polycarboxylates
Optical brighteners Alcohols
Solvents
a
Data from general brochures, package information, encyclopedias, and other general literature, e.g., Reference 9.

vant temperatures using the Terg-O-tometer, which simu- − Runwashed is a measure of the total detergency. ∆REnz =
lates the top-loading U.S.-type of washing machine, and Rwashed − Rwashed without enzyme reflects the contribution of the
enzyme.
the Launder-O-meter, which simulates the European
drum-type machines. These “-O-meters” simulate to some Detailed washing data have been published on many
degree washing mechanics and washing temperatures, in- occasions. Such data are used to inform about the perfor-
cluding initial temperatures, heating rates, and maximum mance of new enzymes or as studies of the influences of
temperatures. Dose-response trials may be carried out in particular surfactants, builders, or effect of design of wash-
ordinary washing machines with ballast laundry and arti- ing machines (39). Several of the references cited contain
ficially soiled fabrics. A sufficient number of “standard” such information.
swatches may be fixed onto the ballast cloth. (ii) Washing performance evaluation—ADD. A standard
Assessment of washed test pieces is done visually or instru- laboratory procedure for evaluating the cleaning efficacy
mentally, the latter by measuring the reflectance of light re- of enzymes, mainly proteases and amylases, has been pub-
mitted at 460 nm. The intensity of the reflected light, % R, lished (28):
(% remission) can thus be measured. The value ∆R = Rwashed To test the efficiency of proteases, stainless steel plates are

Journal of Surfactants and Detergents, Vol. 1, No. 4 (October 1998)

 REVIEW 565

TABLE 8
Laundry Washing Conditions and Procedures by Regiona
Conditions United States/Canada Japan Europe

Machine type Top-loading Top-loading Front-loading

Vertical axis Vertical axis Horizontal axis
Agitator Impeller/pulsator Drum type
Fabric load (kg) 2–3 1–1.5 3–5
Wash liquor (L) 35–80 30–45 15–25
Wash temperature (°C) 20–50 10–30 30–95
Washing time (min) 10–15 15–35 40–60
Water hardness (ppm CaCO3) 100 50 250
(simplified picture)
Detergent dosage levels (g/L)
Powder 1–5 0.5–1.5 5–10
Liquid 2–4 0.75–1.0 7.5–10
a
Data from general brochures, package information, encyclopedias, and other general literature,
e.g., Reference 9.

soiled with a baked egg–milk film. To test amylases, porce- into a detergent formulation may be based on washing tri-
lain plates are soiled with a gelatinized starch solution, als, model tests or machine tests, and on the results of stor-
which is dried overnight at room temperature. After wash-
age tests. Based on these data, a “minimum task value” for
ing, light reflectance values R are measured directly for
protein film or for starch films, after staining with iodine the user can be defined for the enzyme detergency perfor-
(KI/I2). Calculations of % removed film—RPF% for protein mance for the last wash out of a given package.
or RSF% for starch—can be as described in the formula: The factors affecting enzyme stability in formulated de-
RPF(%) [or RSF(%)]
tergents are summarized in Table 9.

= [Rafter wash − Rbefore wash]/[Rclean plate − Rbefore wash] · 100%

FUTURE POSSIBILITIES FOR BIOTECHNOLOGY
IN DETERGENTS
For protein, especially, measured soil removal values
are affected by the temperature at which the film has been Although enzymes have contributed to more environmen-
baked. Removal of starch film is dependent on film thick- tally friendly washing and cleaning, the process still re-
ness, water hardness, and pretreatment of the porcelain quires large quantities of chemicals, energy, and water.
plates (acid or alkaline). Past developments have clearly shown that detergent for-
Storage stability. Average storage stability curves for for- mulations can be optimized based on biotechnological sys-
mulated detergents stored at a relative humidity of 55% in tems. Supported by further development of enzyme sys-
an open vial simulate the storage of an opened package in tems, this trend may continue toward development of ef-
the household. In open packages, detergents may absorb fective detergent systems that use substantially smaller
humidity while bleaches like percarbonate or perborate re- quantities of chemicals and require less water and energy
lease small amounts of hydrogen peroxide that can oxidize to attain maximal washing or cleaning performance. One
the enzyme (14). The measured loss of enzyme activity possibility is the development of special dosing techniques
after several weeks of storage is a useful measure of en- that add active ingredients when they are needed at a par-
zyme stability. ticular stage in the washing or cleaning cycle and thus en-
Storage in a closed vial simulates the storage of a com- hance their performance.
mercial product in the original closed package. Ther- Continued development of new enzymes through mod-
mostated chambers at 30°C are used for both tests. The ern biotechnology may yield functional compounds with
enzyme activities are measured after 1, 2, 6, and 8 wk of considerable and sufficient cleaning effects at low temper-
storage. atures, with low water consumption, at a reasonable price,
Storage stability of the enzymes in detergent formula- and without creating problems for the environment.
tions and wash performance go hand in hand. Storage sta-
bility of enzymes in activated bleach systems has been im-
CONCLUSION
proved significantly in recent years by using genetic engi-
neering. By such techniques, oxidation-sensitive regions of The first development of submerged fermentation tech-
amino acids in an enzyme molecule may be replaced by niques for industrial protease production resulted in the
more stable amino acids (14). creation and maturation of an entire field, and the process
The dosage and selection of enzymes for incorporation is no doubt set to continue. Enzymes have contributed to

Journal of Surfactants and Detergents, Vol. 1, No. 4 (October 1998)

566 REVIEW

TABLE 9
Some Experience of the Effects of Detergent Compounds on the Stability of Enzymes
Compounds Powder detergents Liquid detergents

Water Humidity decreases stability of nonbleach-stable Decreases stability.

enzymes
Proteases Proteolysis is usually no problem in dry products Proteases must be inhibited and recovered during dilution
in the washing process.
Surfactants Usually no problem in dry products Anionics denature. Nonionic or amphoteric surfactants
stabilize most enzymes in water-free systems.
O2-bleach Many conventional types of enzymes are affected Bleaches are usually not used in combination with enzymes.
by oxidation when humidity is present. Exceptions are found when the bleach is kept completely
encapsulated.
Builders Usually no problem in dry products Binding or sequestering of Ca may reduce stability of
proteases, lipase, and amylase. Free Ca2+ may stabilize most
enzymes. The pH should be around neutral.

the development of detergency and improvements in 8. Kemna, R., in Ibid., 1996, pp. 22–27.
household and industrial detergents. In compact detergent 9. Showell, M.S., in Powdered Detergents, edited by M.S. Showell,
and E.J. Baas, Marcel Dekker Inc., New York, 1998, Surfactant
formulations, enzymes have also provided significant en-
Science Series, Vol. 71, pp. 1–19.
vironmental benefits by reducing energy consumption 10. Andersen, M., and H. Lundsgaard, Vaskemaskiner in Rad &
through shorter washing times, lower washing tempera- Resultater, Forbrugerstyrelsen, Copenhagen, 1997, Vol. 37. pp.
tures, reduced water consumption, and reduced chemical 2–7.
load. Proteases are now supported by lipases and amylases 11. Härer, J., Geschirrspülen von Hand und in der Maschine—
ein Systemvergleich, in The 37th International WFK Deter-
in further increasing these benefits, especially by increas-
gency Conference, Proceedings, wfk-Forschungsinstitute für
ing cleaning effects at lower temperatures in household Reinigungstechnologie e.V., D-47798 Krefeld, 1996, pp.
laundering, and at lower pH levels in industrial cleaning. 156–158.
Cellulases have contributed to overall fabric care by reju- 12. Buchmeier, W., H. Andree, P. Jeschke, J. Härer, and R. Sorg,
venating or maintaining the new appearance of washed Corrosion of Tableware in Machine Dishwashing, Henkel-
Referate 33:76–84 (1997).
garments.
13. Gutzschebauch, C., J. Härer, P. Jeschke, F.R. Schröder, and D.
Zaika, Dishwashing by Hand and by Machine—a System
ACKNOWLEDGMENTS Comparison, Ibid. 33:69–75 (1997).
14. Eriksen, N., Detergents in Industrial Enzymology—Second Edi-
We are grateful for help and advice during the preparation of the tion, edited by T. Godfrey, and S. West, Stockton Press, New
manuscript from colleagues at Novo Nordisk A/S and at Novo York and Macmillan Press Ltd., London, 1996, pp. 189–200.
Nordisk BioChem North America, Inc. In particular the following 15. Potthoff, A., W. Serve, and P. Macharis, The Cleaning Revo-
have offered us valuable challenges, critical and well-meaning lution, in Dairy Industries 62:25–29, 1997.
corrections to improve our circulating drafts for internal ap- 16. Krack, R., and H. Manners, Patent Anmeldung, WO 96/23579
proval: Erik Gormsen, Sonja Salmon, Ture Damhus, Tina (PCT/EP96/00258) (Henkel-Ecolab GmbH & Co.) (1996).
Sejersgård Jakobsen, Marianne Thellersen, and Morten Birket 17. Stutz, Chr., The Use of Enzymes in Ultrafiltration, in Fruit Pro-
Andersen. cessing 7, 1993, pp. 1–4.
18. Olsen, H.S., PCT/WO 97/02753 (Novo Nordisk A/S) (1997).
REFERENCES 19. American Conference of Governmental Industrial Hygienists, Doc-
umentation of the threshold limit values, 5th edn., 1986, pp.
1. Röhm, O., German Patent DE 283 923 (1913). 540–541.
2. Conrad, L.S., T. Damhus, O. Kirk, and P. Schneider, Enzy- 20. Peters, G., and D.P. Mackenzie, in Enzymes in Detergency,
matic Inhibition of Dye Transfer, INFORM 8:950–957, 1997. edited by J.H. van Ee, O. Misset, and E.J. Baas, Marcel Dekker
3. Houston, J.H., in Enzymes in Detergency, edited by J.H. van Ee, Inc., New York, 1997, Surfactant Science Series, Vol. 69, pp.
O. Misset, and E.J. Baas, Marcel Dekker Inc., New York, 1997, 327–340.
Surfactant Science Series, Vol. 69, pp. 11–21. 21. National Research Council (NRC), Enzyme-Containing Laun-
4. Krawczyk, T., Detergent Enzymes, INFORM 8:7–14 (1997). dering Compounds and Consumer Health, Report of the ad
5. Braae, K. (ed.), Dansk Husmoder Leksikon, Standard Forlaget, hoc Committee on Enzyme Detergents, Division of Medical
Copenhagen, 1954, pp. 560–569. Sciences, National Academy of Sciences-National Research
6. Rieck, H.-P., in New Horizons. An AOCS/CSMA Detergent In- Council, Report PB 204 118, Washington, DC, 1971, pp. 1–31.
dustry Conference, edited by R.T. Coffey, AOCS Press, Cham- 22. Gormsen, E., E. Marcussen, and T. Damhus, in Powdered De-
paign, 1996, pp. 23–36. tergents, edited by M.S. Showell, and E.J. Baas, Marcel Dekker
7. Hloch, H.G., Tendenzen im Bereich Wasch-und Spülmaschi- Inc., New York, 1998, Surfactant Science Series, Vol. 71, pp.
nen, in The 37th International WFK Detergency Conference, Pro- 137–163.
ceedings, wfk-Forschungsinstitute für Reinigungstechnologie 23. Adler-Nissen, J., and H.S. Olsen, The Influence of Peptide
e.V., D-47798 Krefeld, 1996, pp. 8–14. Chain Length on Taste and Functional Properties of Enzymat-

Journal of Surfactants and Detergents, Vol. 1, No. 4 (October 1998)

 REVIEW 567

ically Modified Soy Protein, American Chemical Society Sympo- 37. Hagen, H.A., in New Horizons. An AOCS/CSMA Detergent In-
sium Series 92, pp. 125–146, 1979. dustry Conference, edited by R.T. Coffey, AOCS Press, Cham-
24. Adler-Nissen, J., Enzymic Hydrolysis of Food Proteins, Applied paign, 1996, pp. 57–62.
Science Publishers, London, 1986, pp. 25–43, 170–186. 38. Damhus, T., L.S. Conrad, and T.T. Hansen, An Enzymatic
25. Linderstrøm-Lang, K., Proteins and Enzymes. III. The Initial System for Dye Transfer Inhibition in Laundry Detergents,
Stages in the Breakdown of Proteins by Enzymes, Lane Medical Lec- Presented at the 88th AOCS Annual Meeting & Expo, Seattle,
tures, Stanford University Press, California, Vol. VI, 1952, pp. Washington, May 11–14, 1997.
53–72. 39. Fleuren, R., P. Bauditz, S. Schløsler, and H. Bodenhoff, Paper
26. Eilertsen, J.H., A.D. Fog, and K. Gibson, U.S. Patent 4,497,897 presented at the 88th AOCS Annual Meeting, Seattle, Wash-
(Novo Nordisk) (1982). ington, May 14–18, 1997.
27. Eilertsen, J.H., A.D. Fog, and K. Gibson, U.S. Patent 4,543,333
(Novo Nordisk) (1982). [Received February 9, 1998; accepted June 30, 1998]
28. Jensen, G., The Development and Application of a New Sta-
ble Detergent Amylase, SÖFW-J. 123:723–731 (1997).
29. Baeck, A.C., T.W. Horner, R. Ohtani, K. Pramod, and S. Rai,
Patent WO9704067-A1 (1997).
30. Gibson, K., The Importance of Detergent Amylases for White-
ness Maintenance, Paper Presented at CID, Genoa, October Dr. Hans Sejr Olsen joined the enzyme application research of
22–24, 1997 (available from Novo Nordisk A/S as A-6506). Novo Nordisk A/S in 1977 as a research scientist. He was head
31. Grime, J.K., Laundry Technology Trends in the Americas, 3rd of Enzyme Process Development Pilot Plant 1980–1993 and has
World Conference and Exhibition on Detergents: Global Perspec- worked as application technologist with detergent enzymes
tives, edited by A. Cahn, AOCS Press, Champaign, 1993, pp.
64–71. mainly for hard-surface cleaning, automatic dishwashing, and
32. Jakobsen, T.S., P. Lindegaard, and M. Chan, Colour Care Cel- industrial cleaning the last few years. He holds several patents
lulases, INFORM 9:788–792 (1998). and over the years has published many papers mainly within the
33. Nielsen, P.H., Development of New Detergent Enzymes, area of enzymes for food processing. He received his M.Sc. in
Paper Presented at ISSD96, China, 1996 (available from Novo chemical engineering from the Technical University of Denmark
Nordisk A/S as A-6457).
34. Gormsen, E., and H. Malmos, Lipolase™—a Lipase for the De- in 1971 and his Ph.D. from the same university in 1980.
tergent Industry, The Soap and Detergent Association, 1991 Per Falholt was born in 1958 and graduated with a M.Sc. in
Annual Convention, Boca Raton, Florida, February 2, 1991. chemical engineering from the Technical University of Denmark
35. Gormsen, E., New Detergent Enzymes Based on Biotechnology, in 1983. He has worked for Novo Nordisk A/S as a research sci-
JOCS/AOCS Conference in Kuala Lumpur, Malaysia, Sep- entist, manager, and director primarily within development of
tember 1997 (available from Novo Nordisk A/S as A-6537).
36. Aaslyng, D., E. Gormsen, and H. Malmos, Mechanistic Stud- detergent enzymes, but lately also personal care, textile, pulp and
ies of Proteases and Lipases for the Detergent Industry, J. paper and food enzymes. Currently he is working out of Novo
Chem. Tech. Biotechnol. 50:321–330 (1991). Nordisk US, NC.

Journal of Surfactants and Detergents, Vol. 1, No. 4 (October 1998)