LACTOPEROXIDASE
Review of Biological Properties
through 1997. (Adapted from Harper, 2000).
The structure, function and antimicrobial
properties of lactoperoxidase have been reviewed recently by de Wit and
Hooydonk (1996). Next to xanthine
oxidase, lactoperoxidase is the most abundant enzyme in milk and is found
almost exclusively in the whey after cheese making.
Lactoperoxidase is a glycoprotein consisting of
a single peptide chain with a molecular weight of 78, 431 Dal. It has 15 half-systemic residues and a much
higher isoelectric point (pH 9.2) than most of the other whey proteins.
The carbohydrate content is about 10%,
structured into four or five potential binding sites. Partial loss of some of the glycosidic
components during isolation has been attributed to its electrophoretic
heterogeneity. The enzyme contains a
haeme structure, with 1 iron molecule per mole of lactoperoxidase. The conformation of the protein is stabilized
by a strongly chelated calcium ion.
Biological activity:
Lactoperoxidase has been identified as an
antimicrobial agent in milk, saliva and tears. Lactoperoxidase is a natural
bacterial defence system through the
oxidation of thiocyanate ions (SCN-) by hydrogen peroxide. These are both present in biological fluids
and together with lactoperoxidase are termed the lactoperoxidase system
(LP-s). LP-s has proven to be both
bactericidal and bacteriostatic to a wide variety of microorgnisms, while
having no effect on the proteins and enzymes of the organisms producing it
(Ekstrand (1994.)
The mechanism of action of the lactoperoxidase
system has been explained in detail by DeWit and Hooydonk (1996).
Lactoperoxidase in more active at acidic pH levels ( Wever, etal. 1982), but is less stable under
acidic conditions ( Carmen, etal. 1990)
Pruitt, etal.
(1982) showed that the lactoperoxidase-catalysed reactions yield short lived intermediary oxidation
products of SCN-, providing antibacterial activity. The major intermediary oxidation product is
hypothiocyante (OSCN-), which is produced in an amount of about 1
mol per mol of hydrogen peroxide. At
the pH optimum of 5.3, the OSCN- is in equilibrium with HOSCN. The unchanged HOSCN is considered to be more
bactericidal of the two forms ( Thomas, etal., 1983). The action of lactoperoxidase against
bacteria is reported to be caused by sulfydryl (-SH) oxidation ( Aune and
Thomas, 1978); Ekstrand, et.al., 1985)
The oxidation of -SH groups in
the bacterial cytoplasmic membrane results in loss of the ability to transport
glucose and also in leaking of potassium ions, amino acids and peptides .
The microbial specificity of LPO has been
reviewed by Kurhonen (1980). Gram
negative, catalase positive organisms are more readily inhibited by LPO than
are gram positive, catalase negative bacteria. Gram negative, catalase positive
organism, (coliforms, salmonella, etc)
are not only inhibited, but are killed if sufficient hydrogen peroxide
is provided -- chemically, eznymatically
or by hydrogen peroxide producing microorganism (Bjorck, 1992; Prudy, et.al
(1983); Reiter, et. al, 1976.) . On the
other hand the action of lactoperoxidase against gram + organisms is generally
bacteriostatic and not lethal (Oram and Reiter, 1966)
Applications:
Lactoperoxidase has been recognized as an effective
antimicrobial agent for many years and has been used extensively as an
antibacteriostatic agent in reducing microflora in milk and in milk for cheese
making(Reiter, 1985a, 1995b). Activation
of the LP-s by addition of hydrogen peroxide and SCN- , both at
concentration of 0.25 mM, extended the shelf life of raw milk at 10oC
for at least three days (Zajac, et.al., 1983). Other dairy uses have
included control of post-culturing
acidification of yogurt (Dosako, 1991); preservation of HTST pasteurized milk
with additional hydrogen peroxide and SCN- added (de Wit and
Hooydonk, 1996); stabillization of caseinate stabilized emulsions at room
temperature ( de Wit and Hooydonk, 1996). When I-was used instead of
SCN-, a shelf life of 10 days was obtained. The
effect was bactericidal, with a decrease in bacterial numbers during storage.
Applications are being found in addition to
use as antibacterial agents and the use of the LPs in cosmetics, ophthalmic
solutions, dental and wound treatment,
and as anti-tumor and anti viral agents are of particular interest.
Godfrey, et.al. (1990) found that there was a
critical combination of LPO, glucose, glucose oxidase (GO), iodide and
thiocyanate to be effective in cosmetics.
The treatment was effective against a range of yeasts, fungi and
viruses, as well as bacteria for periods of up to 4 months.
Poulson (1986)
patented a process for using a lactoperoxidase system for dental and
wound treatment. Hoogedoorn ( 1985) used LPs in toothpaste or a mouthrinse to
reduced acid formation by oral microoganisms. Clinical studies have supported
the possibility that plaque accumulation, gingivitis and early onset carries
may be reduced by appropriate LPs preparations.
LPO, together with glucose oxidase and
monoclonal antibodies, have been applied in tumor therapy (Stanilawski, etlal., 1989;Lefkowitz, et.al.,
1990). Such treatments may also possibly
delay or eliminate the human virus (HSV I) (Courtois, et.al., 1990) -- or
possibly decrease the transcription of human immunodeficiency virus (HIV)-coded
protein (Pourtois, et.al. , 1990)
There has been a suggestion the lactoperoxidase
acts synergistically with lactoferrin (Reiter, 1985); secretory IgA (Tenovuo,
1985) and lysozyme (Roger, etal.,
1994)
References 1999-2001
The lactoperoxidase system (LP) is a part of the natural host
defense system against invading
microorganisms. It’s use as a
preservative for food and pharmaceuticals is of continuing interest. During the
past several years’ additional work has been published, especially as related
to synergistic effects with other agents
and additional information concerning the action of the lactoperoxidase system
against both E. coli O157-H7 and
Listeria.
General
Kussendrager KD & Hooijdank ACM van (2000)
Lactoperoxidase: physico-chemical properties, occurrence, mechanism of action
and applications. British Journal of Nutrition 84 (Supplement 1):
19-25.
Preservative
effects
Atamer M, Kocak C, Cimer A, Odabasi S, Tamucay B & Yamaner N (1999) Some quality characteristics of Kasar cheese
manufactured from milk preserved by activation of
lactoperoxidase/thiocyanate/hydrogen peroxide (LP) system. Milchwissenschaft 54(10):553-556.
Barrett NE, Grandison AS & Lewis MJ (1999)
Contribution of the lactoperoxidase system to the keeping quality of
pasteurized milk. Journal of Dairy Research 66(1):73-80.
Bosch EH, Doorne H van Vries S de (2000) The
lactoperoxidase system: the influence of iodide and the chemical and
antimicrobial stability over the period of about 18 months. Journal of Applied Microbiology;
89(2):215-224.
Buikstra FPM (1999) The
lactoperoxidase system in milk and the way in which its activation can lead to
extended shelf life are described. Voedingsmiddelentechnologie
32(5):32-33, 35-36.
Doorne H van, De Vries S & Bosch EH (2000) The
lactoperoxidase system: the influence of iodide and the antimicrobial stability
over the period of 18 months. Journal of
Applied Microbiology 89(1): 215-224.
Girgis ES, Abd-El-Ghany IHI, Yousef LM & Mohammed LM (1999) Bactericidal effect of some pretreatments of
raw milk on its keeping quality. Egyptian Journal of Dairy Science21(1): 59-70.
Gwan ChangLin & Chi FaChow (2000) Studies on the lactoperoxidase system and its
use in extending the storage period of cow's raw milk. Journal
of the Chinese Society of Animal Science 29(1):89-99.
Jacob BM, Anthony KE,
Sreekumar B & Haridas M (2000) Thiocyanate mediated antifungal and
antibacterial property of goat milk. Life Sciences Including Pharmacology
Letter 66 (25):2433-2439.
Kennedy M, O’Rourke AL, McKay J & Simmonds R (2000)
Use of a ground beef model to assess
the effect of the lactoperoxidase system on the growth of Escherichia coli
O157-H7, Listeria monocytogenes and Staphylococcus aureus in red meat. International Journal of Food Microbiology
57(3):147-158.
Ming CH & Chi FC (1999) Studies on
the lactoperoxidase system and the use of it to extend the storage period of
goat's raw milk. Journal of the Chinese Society of Animal Science
28(3):401-414, 33.
Mehanna NM & Moussa MAM (1999)
Preparation and use of lactoperoxidase system capsules to preserve milk at
different temperatures. Egyptian Journal of Dairy Science
27(2):245-254.
Moussa MAM, Al-Ahwall RIH & Mehanna NM (2000) A comparative study on impact of thermization
and activation of LP-system on quality of milk kept at different temperatures. Egyptian Journal of Dairy Science 28(1):13-22.
Odabasi S, Gursoy A, Cimer A & Atamer M (1999) A study
on some quality criteria of white pickled cheese produced from milk preserved
by the activation of lactoperoxidase/thiocyanate/hydrogen peroxide (LP) system.
Gida 24(5):327-335.
Zhang S, Zhao X & Zheng X (1999) Lactoperoxidase system in milk and its use. China Dairy Industry 27 (2):28-29.
Antimicrobial effects
Doyle ME & Mazzotta AS (2000) Review
of studies on the thermal resistance of salmonellae.
Journal of Food Protection
63(6):779-795.
Heuvelink AE, Bleumink B, Biggelaar FLAM van den, Te-Giffel MC, Beumer RR & Boer E de (1998) Occurrence and survival of
verocytotoxin-producing Escherichia coli O157 in raw cow's milk in the
Netherlands. Journal of Food-Protection
61 (12) 1597-1601.
Sadhana Ravishankar, Harrison MA & WickerL (2000) Protein profile changes in acid adapted Listeria
monocytogenes exhibiting cross-protection against an activated lactoperoxidase
system in tryptic soy broth.
Journal of Food Safety
20(1):27-42.
Shin K, Tomita M & Lonnerdal B (2000) Identification of lactoperoxidase in mature
human milk.
Journal of Nutritional Biochemistry
11(2):94-102.
Boussouel N, Mathieu F, Revol-Junelles AM & Milliere JB (2000) Effects of combinations of lactoperoxidase
system and nisin on the behaviour of Listeria monocytogenes ATCC 15313 in skim
milk.
International-Journal-of-Food-Microbiology;
61 (2/3) 169-175
Garcia-Graells C Valckx C & Michiels CW (2000)
Inactivation of Escherichia coli and Listeria innocula in milk by combined treatment
with high hydrostatic pressure and the lactoperoxidase system. Applied and Environmental
Microbiology
66(10):4173-4179.
Zapicao P, Madeina M, Gaya P & Nnez M (1998) Synergistic
effect of nisin and the lactoperoxidasse system on Listeria monocytogenes in
skim milk. International Journal of Microbiology 11(2):94-102.