Bacteriocins and Their Applications in Food Preser
Bacteriocins and Their Applications in Food Preser
Bacteriocins and Their Applications in Food Preser
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To cite this article: Ramith Ramu, Prithvi S Shirahatti, Aishwarya T Devi, Ashwini Prasad, Kumuda J., Lochana M. S., Zameer
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Reviews in Food Science and Nutrition, DOI: 10.1080/10408398.2015.1020918
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Bacteriocins applications
Ramith Ramu1, Prithvi S Shirahatti1, Aishwarya T Devi1, Ashwini Prasad2, Kumuda J1, Lochana
1
Department of Biotechnology, Sri Jayachamarajendra College of Engineering, Mysore 570 006,
India
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2
Department of Microbiology, Faculty of Life Sciences, JSS University, Mysore, India
3
Mahajana Life Science Research Laboratory, Department of Biotechnology, Microbiology and
Biochemistry, Pooja Bhagavat Memorial Mahajana Post Graduate Centre, Metagalli, Mysore -
ABSTRACT
produced by bacterial strains. They are generally effective in inhibiting the growth of similar or
closely related bacterial strains. A high diversity of various bacteriocins is produced by many
lactic acid bacteria (LAB) and is found in numerous fermented and non-fermented foods. Several
bacteriocins from LAB extend potential applications in food preservation, thus help foods to be
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naturally preserved and richer in organoleptic and nutritional properties. Though chemical
preservatives for the preservation of food are successful to some extent, their quality is not as
satisfying as fresh food. Hence, an alternative is required and bacteriocins serve the purpose.
Nisin is currently the only bacteriocin widely used as a food preservative. Numerous bacteriocins
have been characterized chemically, biochemically, genetically and also at the molecular level to
understand their basic mode of action. This article gives an overview of classification of
bacteriocins, isolation & characterization, and mode of action. Besides, article highlights the
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optimized parameters for growth of bacteria in the production of bacteriocins and various
bioassays for their determination. Special emphasis has been provided on explaining the
Keywords
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INTRODUCTION
All organisms produce organic compounds that are not directly involved in their normal growth,
development, or reproduction and are termed as secondary metabolites (Fraenkel 1959). They are
are also produced by some bacteria and, hence, along with bacteriocins they should be
considered as different antimicrobial compounds (Davidson and Hoover 1993). A large group of
functionally diverse antimicrobial compounds found in all major lineages of bacteria and archaea
constitute the bacteriocin family (Riley and Wertz 2002). While some possess a narrow
bactericidal spectrum, many others exhibit a broader activity against some of the distantly related
species (Klaenhammer 1993; Adam and Moss 1995). Over-all, specific immunity proteins
responsible for this are encoded in the bacteriocin operon (McAuliffe et al., 2000; Cintas et al.,
2001). Majority of them exert their action through the formation of transitory poration complexes
or ionic channels which cause reduction or dissipation of the proton motive force leading to the
formation of pores in the cytoplasmic membrane of sensitive cells (Cintas et al., 1998; Herranz et
al., 1999). They also employ other killing mechanisms like cell wall interference and nuclease
them and antibiotics under the same class but since bacteriocins are bactericidal peptides, while
antibiotics are produced by multi-enzyme complexes, there remains a demarcation between the
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two bactericidal agents. Most bacteriocins kill a narrow spectrum of bacteria, as compared to the
traditional antibiotics (Tu et al., 2002). Some bacteriocins produced by lactic acid bacteria (LAB)
are called lantibiotics and they have potential applications in the food industry. Currently
existing synthetic food preservatives raise some concern with regard to the quality of food and
are associated to several health hazards. In this view, bacteriocins prove safer alternatives and
efficiently produced from isolated and purified bacteria. Bacteriocins are categorized and
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discussed in detail as shown in Figure 1. They are characterized based on different principles
viz., chemical, biochemical, molecular, and genetic characteristics (Mian et al., 2012).
Classification
Bacteriocins are an important part of the defence system of bacteria and are produced by both
Gram-positive and Gram-negative species. Bacteriocins from Gram-positive bacteria are broadly
classified into 4 groups. The presence of sulfide (such as lanthionine) and disulfide bonds has
been taken as the major distinguishing factor since it also entails their spectrum of activity (Jack
et al., 1995). However, the molecular mass, thermo-stability, enzymatic sensitivity, presence of
post translational modified amino acids, and mode of action also play a role, especially in the
classification into sub-groups (Klaenhammer 1993). The groups may be stated as:
i. Class I-Bacteriocins
Class I-Bacteriocins
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Class I bacteriocins are post-translationally modified peptides containing unusual amino acids,
such as lanthionine or methyl lanthionine residues and are called lantibiotics (Nissen and Nes
1997). They are produced by LAB to attack other Gram-positive bacteria and are smaller than 5
kDa (Boman 1995). Nisin, subtilin, cytolysin and variacin 8 are a few examples (Gross and
Class II-Bacteriocins
Class II bacteriocins are heat-stable, unmodified peptides, and do not contain the unusual amino
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acid and hence they are called non-lantibiotics. They are small peptide molecules which are
smaller than 10 kDa and produced by LAB (Bierbaum and Sahl 2009). Class II bacteriocins are
further divided into 3 sub-classes, namely IIa, IIb, and IIc (Ennahar et al., 2000). Pediocin-like
bacteriocins, two-component bacteriocins, and thiol-activated bacteriocins are grouped under the
IIa, IIb, and IIc sub-classes, respectively. Some of the examples are pediocin PA-1, lactacin F,
enterocin AS-48 (Nissen et al., 2009). The major difference between Class I and Class II
bacteriocins is that Class I bacteriocins undergo post-translational modifications and thus give
rise to the unusual amino acid lanthionine, whereas Class II bacteriocins do not undergo any such
modifications.
Class III-Bacteriocins
Class III bacteriocins are heat-labile and larger (>10 kDa) protein molecules (Savadogo et al.,
2006). They are sub-classified into 2 sub-classes: Class IIIa or bacteriolysins and Class IIIb or
Class IV-Bacteriocins
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Class IV bacteriocins are complex proteins that contain lipid or carbohydrate moieties (Oman et
al., 2011; Upreti and Hinsdill 1975). This group was added after the observation that bacteriocin
activities obtained in a cell free supernatant were abolished not only by protease treatments, but
also by glycolytic and lipolytic enzymes (Jimenez et al., 1993; Tagg et al., 1976; Nes et al.,
2000; Garneau et al., 2002). Plantaricin S and leuconocin S are major examples of Class IV
Further, according to the scheme of Cotter et al., (2005), bacteriocins are classified into 2 major
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groups, namely lantibiotics and unmodified peptides. The authors eliminated classes IIa, III, and
IV and restructured class II. Thus, the class of unmodified peptides includes pediocin-like
both the schemes, a universal bacteriocin classification was established, as shown in Figure 1.
While Gram-positive bacteria are classified as detailed in the above section, Gram-negative
bacteriocins are categorized based on their size. These bacteriocins are produced from Gram-
negative bacteria (arise mainly from enterobacteriaceae) and are heat-stable molecules. They are
ii. Low-molecular-mass peptides (1 kDa-10 kDa) called microcins (Drider and Ribuffat
2011).
Since the beginning of 21 stcentury, bacteriocins have been classified in several other ways, based
on their specific adsorption and immunity patterns (Frederic 1957). Reeves named and grouped
bacteriocins into 16 classes based on the species of the producing strain, like the colicins which
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Bacteriocins affect different essential functions of the living cell viz., transcription, translation,
replication, and cell wall biosynthesis due to great variety of chemical structures, but most of
them destroy the energy potential of sensitive cells by forming membrane channels or pores
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(Juan et al., 2000; Ganzle 2004). The best-described mechanism is pore formation. The presence
of molecular receptors in the membrane of the target cell is suggested by the relatively small
action spectrum of some bacteriocins, although this has not been demonstrated (Van and Stiles
2000).
A model demonstrated using nisin suggests that, using C-terminal, nisin primarily binds to
bacteria (Breukink and Kruijff 2006). Hence a complete loss of activity could be observed if the
C-terminal is removed (Giffard et al., 1997). Binding of nisin is followed by insertion and
penetration of a part of peptide into the cytoplasmic membrane. Fluorescence studies indicate a
parallel orientation of nisin molecule with respect to the membrane surface having N-terminal
inserted slightly deeper than C-terminal (Breukink et al., 1998). This initiates the process of
membrane insertion and pore formation, leading to rapid cell death (Cotter et al., 2005; Breukink
In general, the helical amphiphilic structure of class II peptides allows them to get inserted into
the membrane of the target cell, leading to depolarization and death (Cotter et al., 2005; Drider et
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al., 2006). At the hydrophilic N-terminus of the peptides, the initial interaction takes place with
the heads of the anionic membrane phospholipids. The C-terminus of the peptide is thought to be
involved in hydrophobic interactions with the membrane as it is more hydrophobic than the N-
terminal (Alonso et al., 2000).Class II bacteriocins allow the insertion of the peptide into the
membrane of sensitive microorganism, using its amphiphilic structure, and causes depolarization
On the contrary, the large bacteriolytic proteins, such as lysostaphin (Class III bacteriocins), lead
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to death and cell lysis by directly acting on the cell wall of Gram-positive target cells (Cotter et
al., 2005). Under the influence of various mechanisms, bacteriocins cause microbial cell killing
either in an isolated or most consorted manner (Paker et al., 1989; Daw and Falkiner 1996).
with protein synthesis and changing the cell translator mechanism lead to bacteriocins killing a
microbial cell (Nes and Tagg 1996). Due to the development of specific immunity mediated by a
protein, bacteriocin-producing cells are not affected by the action of their own bacteriocin
(Hancock and Chapple 1999). Bacteriocins have been noteworthy in detections and causing
fatality to competitor bacteria, with little or no damage to their producing cell (Tadashi and
Schnneewind 1998).
Microorganisms have competitive superiority when they dispute ecological niches in the
environment where they are developing their metabolic activities (Stockewell et al., 1993).
Based on this narrow action, bacteriocins play a role as intra-specific mediators or promoters of
microorganisms are present in an environment and adversely interfere with growth and survival
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bacteria on the other bacteria in the same environment (Schillinger 1990; Ray 1996).
The majority of bacteriocin producers are natural food isolates and hence they are suited for food
applications (Raja et al., 2010). An increased health-conscious public now tries to avoid foods
with chemical preservatives or which have undergone extensive processing. The most powerful
means for obtaining useful cultures for scientific and commercial purposes are isolation and
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screening of microorganisms from natural sources (Vanden et al., 1993). Isolation procedures
employed by different researchers were relatively similar, but for the culture conditions and the
growth media required they differed between different food sources. The principle involved in
According to the procedure employed by Arokiyamary and Shivakumar (2011) on milk products,
isolation of bacteriocins from LAB was standardized. The bacterial count was performed to
make appropriate dilutions on the most accepted medium, MRS agar, to maintain optimal
growth. The LAB grown in an aerobic environment, utilizing the nutrients from MRS agar, were
Vijai et al., (2005) standardized a procedure for the extraction of bacteriocins from South Indian
special dosa (commonly known as appam) batter. Saline dilutions of appam batter, cultured on
MRS agar anaerobically, was transferred to MRS broth and checked for purity. For the purpose
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was utilized.
Apart from food products, isolation of bacteriocins has been reported in marine environments,
rock sediments, and chicken intestine. Narayanapillai et al., (2012) have designed an isolation
method from chicken intestine. Briefly, pure cultures obtained from inoculation of crushed
pieces of chicken intestine were grown on MRS agar at optimum temperature to test the
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bacteriocin production. The strains from the pure culture were Gram-stained to examine
homogeneity become a very complex task. Although bacteriocins are generally extracted after
centrifugation using the ammonium sulfate precipitation method, there are 3 other major
methods of purification which have been widely accepted. The first is a conventional method and
includes ammonium sulfate precipitation, ion exchange, hydrophobic interaction, gel filtration,
and reverse phase high-pressure liquid chromatography (RP-HPLC). A second method is much
simpler and generally used for the production of bacteriocins at laboratory-scale. The steps
then RP-HPLC. The third bacteriocin purification method requires maximization of the
unit operation protocol, namely hydrophobic interaction gel or expanded bed adsorption, can be
employed. This method is used for the production of bacteriocins at bulk scale (Eduardo et al.,
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2013). Isolation of bacteriocins has also been done from a variety of food systems like dhokla,
By definition, bacteriocins can also be designated as antibiotics due to their bactericidal mode of
action against closely related species (De Man et al., 1960; Klement et al., 1990; Kuryolowcz
1991; Davis 1999). The most abundant and diversified molecules with antibiotic activity
produced by different bacteria including Gram-positive and Gram -negative species are
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bacteriocins (Riley 1998). A major difference between bacteriocins and antibiotics is found in
their activity of inhibiting specific microorganisms. Bacteriocins restrict their activity to strains
of related species whereas antibiotics have a wider activity spectrum and do not show any
Based on their molecular size, antibacterial spectrum, stability, physical and chemical properties,
and action mode, bacteriocins are formed into heterogeneous groups (Davidson and Hoover
1993; De Vuyst and Vandamme 1994, b). Most bacteriocins of low molecular weight are
cationic and have greater antimicrobial activity at low pH (Sinell 1989; Bendjeddou et al., 2012).
maximum adsorption at or above pH 6. The mode of action, effect of heat, pH, proteolytic
enzymes, salt, and detergents on bacteriocin activity, determination of molecular mass, amino
acid composition and sequence, and determination of the genetic organization, production, and
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According to the experiments conducted by Maja et al., (2010) bacteriocin BacUB9 is a heat-
paracasei BGUB9.Among the 2 bacteriocins characterized from this bacterium, BacUB9 is the
one which has a very narrow anti-microbial spectrum. Its molecular mass is 4 kDa and it is
susceptible to the activity of proteolytic enzymes. Its activity decreases at 100ºC for 60 min and
its activity is completely abolished at 100ºC for 120 min. Bacteriocin BacUB9 retains its activity
in the pH range of 1-10 when it is present in a cell-free supernatant of the producer strain.
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Antimicrobial activity is lost at pH 11. Treatment of bacteriocin UB9 with various proteolytic
enzymes (pepsin, trypsin, α-chymotrypsin, pronase E, and proteinase K) results in the loss of its
antibacterial activity. Bacteriocin UB9 shows that it is of a proteinaceous nature and it is strongly
Carotovoricin and carocin S1 are 2 different bacteriocins which are found in the Gram-negative
plants (Eunjung et al., 2010). Carocin D is the third bacteriocin found in the same producing
species. An antibacterial activity of carocin D against the indicator strain Pcc3 is carried by a
particular strain Pcc21 (Chan et al., 2009). Carocin D is encoded by the caroDK gene located in
the genomic DNA, which is an immunity protein providing protection. The Edman degradation
process determined the N-terminal amino acid sequences of purified carocin D. In a comparison
with the amino acid sequence of caroDK, it was found that 8 amino acids are missing at the N
terminus. Mora et al., (2008) and Bendjeddou et al., (2012) proved that 8 amino acids are
removed from the N terminus of carocin D during maturation and that it is synthesized as a
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precursor peptide. Bacteria became resistant to carocin D, when caroDK and caroDI genes were
transformed into carocin D-sensitive bacteria such as Pcc3. This bacteriocin is slight resistant to
heat but not to proteases. CarocinD sensitive, non-pathogenic bacteria readily express this
Carnobacterium piscicola LV17B produces carnobacteriocins BM1 and B2, which are thermo
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et al., 1995). Using the information from the N-terminal amino acid sequences for the purified
bacteriocins, probes were synthesized to locate structural genes for the carnobacteriocins
(Saucier et al., 1995). A HindIII fragment of 1.9-kilobase (kb) from a plasmid (pCP40) of 61-kb
contains the carnobacteriocin B2 structural gene. Presence of the 61-kb plasmid is necessary for
the expression of chromosomal bacteriocin and its immunity function. These bacteriocins are
synthesized as pre-bacteriocins (Jack et al., 1995). Mature 43-amino acid carnobacteriocin BM1
(4524.6 kDa) and the mature 48-amino acid carnobacteriocin B2 (4969.9 kDa) are obtained by
-2 and -1). These 2 peptides show significant amino acid homology to each other and with those
class II bacteriocins containing the YGNGV amino acid motif near the N terminus.
Nisin is the first lantibiotic (Class I - Bacteriocins) mentioned in the scientific literature; it is
produced by certain strains of Lactococcus lactis (Jung 1991; Murugesh 2003). It has a
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molecular weight of 3354 and is composed of 34 amino acids and displays a bactericidal mode of
activity and is said to be nontoxic to animals and humans (Hurst 1983). Rogers and Whittier
observed its activity for the first time in 1928, and it was studied as a discrete antimicrobial
substance in 1944 (Mattick and Hirsch 1947). Nisin is the best-studied bacteriocin produced by
some Lactococcus lactis strains and has been used for food preservation in over 50 countries.
The corresponding study describes characterization of nisin Z-producing L. lactis strain isolated
from Boza traditional cereal-based fermented food from Turkey. In this study, bacteriocin by
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Lactococcus lactis subsp. lactis GYL32 strain inhibited not only related strains but also Gram-
positive bacteria, such as Listeria monocytogenes, Staphylococcus aureus, and Bacillus cereus.
heat-stable when treated at 100 ºC for 20 min. Different pH, enzyme, and heat treatments
concluded that the bacteriocin produced by GYL32 is nisin Z. The sequence analysis result
showed that it is nisin Z, and its genetic determinants are encoded on genomic DNA. The
results of this study suggested that nisin Z producer L. lactis subsp. lactis GYL32 strain may be
used as a starter culture for improving the safety of fermented products (Gozde and Yasin 2012).
Optimization
Different ecological niches from which strains of bacteriocin have been isolated, including meat,
fish, fruits, vegetables, milk, and cereal products (Ammor et al., 2006). Factors affecting the
bacteriocin production include type of medium used, glucose concentration, temperature, pH,
and agitation (De Vuyst and Vandamme 1994, b); Sanchez et al., 2002). These bacteriocins
differ widely in molecular weight, pH, presence and number of particular groups of amino-acids.
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However, the differences in antimicrobial activity cannot be solely attributed to the amino acids
Some of the widely used bacteriocins have been well researched and their production has been
well optimized. For example, in the production of nisin and pediocin PA-1 from P. acidilactici,
Yang and Ray (1994) identified that high cell density, optimal pH, and specific nutrients in the
media resulted in an increased yield. Specifically for nisin, the pH optimum was identified to be
pH 4 – 6 and temperature optimum at 150C. Also, Coventry et al., (1996) established that
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calcium silicate and several desorbing agents in comparison to ammonium sulfate precipitation.
Universally, Class I and Class II bacteriocins are stable at acidic pH. Rodriguez et al., (2002) and
Jack et al., (1996) effectively demonstrated this for pediocin PA-1 and piscicolin 126 which
remained stable at pH range between pH 4-6 and pH 2-3, respectively. According to Usmiati
(2009), the optimum conditions for bacteriocin production on S. typhimurium, E. Coli, and L.
monocytogenes (Holo et al., 1991; Sullivan 2002) are at 330 C and pH 5. The trials involved 2
steps:
The parameters to be observed are turbidity of bacterial culture in nutrient broth and the number
of activated colonies of the isolate (Vaughan et al., 2001). Svetoslav et al.(2012) observed that,
bacteriocin ST22Ch belonging to Class II group, had a narrow spectrum of activity, was heat-
resistant and stable at pH 2-10. He also observed that different levels of bacteriocin ST22Ch are
produced during the stationary phase of fermentation in the presence of 2% (w/v) glucose and a
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combination of tryptone, meat extract, and yeast extract. These findings determined the varying
For the detection of bacteriocin production and its activity, many methods have been described.
It is a fact that bacteriocins can diffuse in solid or semisolid culture media, which can be
subsequently inoculated with a suitable indicator strain. Making use of this fact, authors Kekessy
and Piguet (1970) conducted experiments on the detection of bacteriocins using agar. Indicator
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and producing strains of bacteria were used for the screening purpose (Nicolle and Prunet 1964).
Using the micro-titration method involving polystyrene micro-plate wells containing MRS broth,
Daba et al., (1994) made serial 2-fold dilutions of bacteriocin preparations. The diluted culture
was incubated at optimum temperature and the activity was further detected. For the experiment,
Bouksaim et al., (1999) utilized ELISA for the detection of bacteriocins. They mainly used
affinity-purified anti-nisin immunoglobulin for the binding of nisin and anti-nisin peroxidise
which was linked with the substrate. A carbonate-bicarbonate buffer of basic pH was used for
incubation. To reduce the nonspecific binding, the microtiter wells were coated with a blocking
reagent, Tween-20. The bound enzyme was determined which implied the amount of
bacteriocins. ELISA and agar diffusion assays, even though were more sensitive for pure nisin,
they failed to differentiate nisin from subtilin and also the closest structural analogue of nisin
(Fowler et al., 1975; Falahee et al., 1990). Later, competitive direct ELISA assays were
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published by Suarez et al., (1996) where the detection limit for nisin was based on polyclonal
mouse antibodies, and the above limit for nisin was measured utilizing the monoclonal mouse
antibodies.
It was used for detection of nisin by Simone et al., (2003) by the use of a special media
containing meat extract, yeast extract, tryptone, glucose, NaCl, Na 2HPO4 with neutral pH. At
specific time intervals, samples were collected, their optical densities were measured
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spectrophotometrically and the growth curve of the test microorganism was obtained. To stop the
further growth of microorganism, the samples were injected with thiomersalate solution.
Electrophoretic method
The only electrophoretic method for nisin quantification has been published by Rossano et al.,
(1998). Their capillary zonal electrophoretic assay was used to analyze nisin from milk. The
bioluminescence genes from Xenorhabdus luminescens species were placed under the nisin
inducible promoter in the same plasmid and were transformed into the non-nisin producer L.
lactis. This led to the extra cellular nisin stimulation and a subsequent addition of substrate for
the luciferase, resulted in bioluminescence by the indicator strain (Wahlstrom and Saris 1999).
Very recently Immonen and Karp (2007) introduced an improved luciferase assay.
It was described by Bouksaim et al., (1999) in which serum from rabbit immunized with nisin Z
was used to detect nisin. Dadoudi et al., (2001) developed the first method to distinguish between
nisin A and nisin Z. The monoclonal antibodies raised against nisin Z could be used to quantify
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nisin Z with detection limits in pure solution, in fermentation broth, in milk and in whey and the
It is a fact that bacteriocins can diffuse in solid or semi-solid culture media, which can be
subsequently inoculated with suitable indicator strain like phage (FredericQ 1957). Before
proceeding to any steps of assay, chloroform vapour was used to ensure the sterility of agar
surface. As chloroform vapours attack plastic, the assay does not allow the use of plastic Petri-
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dishes (Nicolle and Prunet 1964). Kekessy and Piguet (1970) described a new technique based
on the tri-dimensional diffusion of bacteriocins. The detection of bacteriocins was done on the
reverse side of agar plate which serves as culture surface for the indicator strain. There is no
direct contact between producer and indicator strains of bacteria. As in the case of other
methods, where it is not easy to decide whether a zone of inhibition is due to either producing
strain or indicator strain, in this method, no inhibition area due to indicator strain are formed.
In context to food, it isn’t enough to only improve production but it is also necessary to store the
produce for distribution and later use. But there are many criteria considered, in order to safely
store food products (Brul and Coote 1999). The currently existing preservatives are not entirely
satisfactory to the consumers, food laws or the producers. Food preservation basically requires
the maintenance of the functional properties of the product, and also other physico-chemical
properties so that it remains appealing to the consumer. However, higher the preservative
treatment, higher is the damage caused to the food product (Ray 1992). Thus, continued
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Bacteriocins as biopreservatives
In recent times, there is a continuous raise of awareness in the public regarding the amount of
chemical intake, which curbs the amount of chemical preservatives that may be used in food
(Harris et al., 1997). Thus, barely any variety exists without controversies of the multifarious
health problems associated with them. In view of the above problems, bio-preservatives are in
very high commercial demand at present, in the form of protective cultures or their metabolites
i.e. enzymes and bacteriocins (Holzapfel et al., 1995). Bio-preservation is defined as extension of
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shelf life and food safety by use of natural or controlled microbiota and/or their antimicrobial
compounds. The destruction of the pathogens depends on improved food safety by search of
more efficient chemical preservative and drastic physical treatment (high temperature treatment).
These have various drawbacks such as toxicity of chemicals used, nutritional alteration of food
and to overcome these disadvantages, LAB and their bacteriocins are being popularized.
Lactic acid bacteria have been widely used for thousands of years in the production of fermented
products. Since they are present in several consumed products, they have been deemed safe for
human consumption and been provided the GRAS (Generally Regarded As Safe) status
(Jeevaratnam et al., 2005). Due to this, most of the research on bacteriocins so far has been
concentrated exclusively from LAB. The bacteriocins from food grade Lactic Acid Bacteria
ii. Does not alter the nutritional properties of the food product
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iv. Active under refrigerated storage (Sinell 1989; Daeschel and Ray 1992; Gould 1995).
Most of the Gram-positive bacteria are especially susceptible to Nisin (Class I-Bacteriocin), but
have little or no effect on Gram-negative bacteria, yeast or fungi (De Vuyst and Vandamme
1994, a). In the year 1969, nisin was stated to be safe and natural food additive by FAO/WHO
expert committee on food additive (FAO/WHO 1969). Nearly 15 years later, nisin was used
commercially in around 39 countries (Hurst 1983). It was given the designation E234 in 1983
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after being incorporated to the EEC food additive list (EEC 1983). Nisin was GRAS (Generally
Recognized As Safe) certified in USA in 1988 (Federal Register 1988) and by Food and Drug
Administration in the year 2001 (FDA 2001). It was allowed as food additive in more than 50
countries including Europe, China and the US by the year 1996 (Delves-Broughtan et al., 1996).
Its potential application was first demonstrated in 1981 for food preservation and elaborated its
use since 1990 (Delves-Broughtan 1990). Nisin concentrate was commercialized as ‘Nisaplin’ by
Alpin and Barrett which is now used as a preservative in dairy products, cured meat, canned food
and other fermentation industry divisions. Nisin was first discovered in 1928 as a result of
difficulties faced in cheese making where storage of milk had supported the growth of
contaminant organisms to produce the inhibitor (Hirsch et al., 1951). A partially purified form of
nisin was commercially made (De Vuyst and Vandamme 1994 , a; Twomey et al., 2002) and a
al., 1999).
Bacteriocins have been found to inhibit food spoilage by pathogenic bacteria such as
Staphylococcus aureus (Chang and Chang 2011), Escherichia coli (Foulds and Shemin 1969),
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Bacillus cereus (Kabore et al., 2013), Listeria monocytogenes (Pucci et al., 1988) and
Clostridium perfringens (Gamal 2006) which are recalcitrant to food preservation methods. The
production and application of bacteriocin could effectively combat nearly all of the problems of
the food industry apropos preservation. Bacteriocins are well suited for this purpose due to their
There are so many different types of food ranging from spicy ethnic food to down home
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favourites. The options are endless, even within a particular recipe or type of food. India being
the country of diversified geographical and cultural origin is also known for its many traditional
cuisines. Many of these products, apart from their cultural heritage also have been known to
possess many beneficial qualities. Few examples of such products are dosa (Mallesha et al.,
2010), idli (Vijayendra et al., 2010), kokum (Khurana and Kanawjia 2007) and lassi (Amit et al.,
2012). They are traditionally known for their effects such as cooling the body, aiding digestion,
Indian traditional products provide a promising array of sources for obtaining bacteriocin
producing bacteria. Lactic acid bacteria isolates from appam batter, capable of producing good
amount of bacteriocins have been identified by Vijai et al., (2005). Isolates of bacteriocin
producing microorganisms were also isolated from curd, dosa batter and idli batter and the
bacteria were identified as a species of Lactobacillus while the effect of physical and cultural
parameters were tested on them (Sourav and Arijit 2010). Lactobacillus species was also recently
isolated from different milk products such as curd, milk peda, butter and ghee and the bacteriocin
from them were found effective against many common food pathogens (Arokiyamary and
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Shivakumar 2011). So it is sufficiently established that Indian traditional products are rich in
lactic acid bacteria that produce bacteriocins with the potential to be used as food preservatives.
However, most of the work was started only very recently and there are still many unexplored
CONCLUSION
covering a broad range of applications in the food and medical industry. They employ a variety
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of killing mechanisms that contain genes which are either chromosomally or plasmid encoded
with certain toxins. Bacteriocins possess relatively narrow killing spectrum that includes
cytoplasmic membrane pore formation, cell wall interference and nuclease activity that alters the
cell membrane permeability properties. Most of the bacteriocins were originally isolated from
organisms which are involved in food fermentation. Considering its wide applications,
bacteriocins also known as Lantibiotics, were regarded as GRAS and hence is one of the most
researched types of bacteriocins. The most potent lantibiotic is nisin which is produced by
Lactococcus lactis and has been regarded as the only bacteriocin used as a food preservative that
has been approved by the FDA. Various bioassays are designed most of which are with regard to
Nisin which also finds applications in pharmaceuticals. Bacteriocins are potentially used in food,
human and animal health applications. The use of bacteriocins is of great interest in the food
system since they are recognized as safe and can also be used as bio-preservatives. Thus it is
desirable to expand the understanding of the activity of bacteriocins in order to determine their
efficacy more accurately for future applications in food model systems. Food safety is deemed as
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the paramount of international concern. Since bacteriocins are considered as natural products,
usage of bacteriocins in food systems will surely gain good acceptance from customers who
demand for more natural and safer food products that are free from chemicals.
Declaration of Interest
ACKNOWLEDGEMENTS
Ramith Ramu thanks TEQIP New Delhi, India for awarding the Research fellowship. The
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authors are grateful to the Principal, Sri Jayachamarajendra College of Engineering, Mysore, and
the Head of the Department of Biotechnology for their encouragement and support. DBL thanks
Jain University for constant encouragement and support given to the progress in research.
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Enterocin
AS-48,
Listeria Ananou and
Class II Enterocin A, Meat
monocytogens others 2005
Pediocin
Gram
ACH
positive
Staphylococcus Wu and others
Pasteurized
Class III aureus and 2003; Maria
Lysostaphin milk, cheese,
Staphylococcus and others
sausages
epidermidis 2004
Carr and
L. paraplantarum Milk and others 2002;
Class IV Plantaricin S
and L. pentosus cheese Pepe and
others 2004
Brenda, 2007;
Obi and
Colicins Colicin E1 E.coli O157:H7 Meat
Campbell,
Gram
1978
negative
Probiotic Anke and
Microcins Microcin S E. coli E2348/69 drug others 2012
Symbioflor 2
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