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Research Article
Article History
Received : 24.04.2015
Revised : 02.05.2015
Accepted : 15.05.2015
1. Introduction
J. Bhuvaneswari/ Life Science Archives (LSA), Volume 1, Issue 4, Page 257 - 263, 2015
stolonifer,
Penicillium
chrysogenum
and
Salmonella typhi have been identified as the
causing agents of the major food borne diseases,
food borne illness and food spoilage (Betts et al.,
1999; Solomakos et al., 2008).
A number of microorganisms belongs to
Lactobacillus species are able to produce
biologically active antimicrobial peptides which is
a proteinaceous substance named bacteriocins
which have a broad spectrum activity, target
attained mode of action, appropriate molecular
weight and biochemical properties. Bacteriocins
are bacteriostatic or bactericidal to pathogenic
bacteria, especially those are close in
taxonomically related, but also bacteria confined
within the food spoilage causing group. The
producer strain is usually immune to the produced
bacteriocin (Abee et al., 1995; OKeeffe et al.,
1999; Van Reenen et al., 2002). Bacteriocins
produced by food-associated microorganisms such
as lactic acid bacteria, in particular Lactobacillus
lactis and Lactobacillus acidophilus is attracting
increasing attention as food biopreservatives
(Abee et al., 1995). Because of its protein nature,
bacteriocins are readily degraded by the protein
degrading enzyme protease in the gastrointestinal
tract of humans and most of the bacteriocin
producing lactic acid bacteria has Generally
Regarded as Safe (GRAS) status. Because of that
reasons bacteriocins are considered as the natural
biopreservatives (Schillinger et al., 2001;
Aymerich et al., 2000). Bacteriocins may also be
used as part of a multiple hurdle preservation
system (Cleveland et al., 2001), the bacteriocin
produces may be applied as bacteriocinogenic
cultures to non-fermented foods or even be used as
starter culture for fermented foods facilitate to the
improvement of quality and safety and to control
the bacterial spoilage or pathogenic bacterial
isolates (Zhang et al., 1999; Nilsson et al., 2000).
Several activities of bacteriocins have been
reported by various researchers and they proposed
its potential in the food industry (Cleveland et al.,
2001). However, it was important that applied
studies be done to confirm the effectiveness of the
addition of bacteriocins to food systems as it has
258
J. Bhuvaneswari/ Life Science Archives (LSA), Volume 1, Issue 4, Page 257 - 263, 2015
distilled water for 10% concentration and stored
in refrigerator. The antagonistic activity of crude
bacteriocin from Lactobacillus lactis and
Lactobacillus acidophilus was tested by Well
diffusion assay and Minimum inhibitory
concentration test was followed by Tagg and
McGiven (1971) and Darwina et al. (2012).
2.3. Effect of Lactobacillus lactis and
Lactobacillus acidophilus crude bacteriocin on
the inhibition of growth of the dairy product
spoilage bacterial strains by Well diffusion
assay method
One ml of the cell suspension (107 cell ml) of the dairy product spoilage bacterial strains
viz., Listeria monocytogenes (MPB - 1), Yersinia
enterocolitica (MPB - 2), Campylobacter jejuni
(MPB - 3), Staphylococcus aureus (MPB - 4) and
Pseudomonas aeruginosa (MPB - 5) were
prepared separately, mixed with the 100 ml
Mueller Hinton agar medium (seeded medium)
and plated. On the surface of the medium, well
were made by using sterile cork borer (6 mm in
size). Ten percent of crude bacteriocin at different
levels viz., 10, 20, 30, 40 and 50 l mixed with
sterile water to make upto total volume to 100 l
was discharged into the well. The well with sterile
water served as control. The plates were incubated
for 2 days at 30C. The inhibition zone (in mm)
was measured around the well using antibiotic
zone scale. Three replications were maintained in
each treatment. The test for antagonistic activity
was performed as per by Tagg and McGiven
(1971) and Jageethadevi et al. (2012).
1
259
J. Bhuvaneswari/ Life Science Archives (LSA), Volume 1, Issue 4, Page 257 - 263, 2015
260
Table 1: Inhibitory effect of Lactobacillus lactis crude bacteriocin on Dairy product spoilage bacteria
by Agar well diffusion method
Concentration of
Diameter of zone of inhibition (mm)
Lactobacillus
Listeria
Yersinia
Campylobacter
Staphylococcus
lactis crude
monocytogenes
enterocolitica
jejuni
aureus
bacteriocin (l)
Control
NZ
NZ
NZ
NZ
10
13.66
12.00
10.00
13.00
20
14.66
13.00
11.66
14.00
30
16.00
14.66
12.00
15.33
40
17.66
15.00
13.66
16.66
50
23.00
21.00
18.33
22.00
S.Ed
1.64
1.56
1.42
1.57
CD (P = 0.05)
3.29
3.13
2.85
3.15
NZ No zone of inhibition; *Values are average of three replications
Pseudomonas
aeruginosa
NZ
11.66
12.66
13.00
14.00
19.66
1.41
2.83
Pseudomonas
aeruginosa
+++
+++
++
+
+
-
Table 3: Inhibitory effect of Lactobacillus acidophilus crude bacteriocin on Dairy product spoilage bacteria
by Agar well diffusion method
Concentration of
Lactobacillus
Diameter of zone of inhibition (mm)
acidophilus crude
Listeria
Yersinia
Campylobacter
Staphylococcus
bacteriocin (l)
monocytogenes
enterocolitica
jejuni
aureus
Control
NZ
NZ
NZ
NZ
10
14.78
13.10
11.18
14.06
20
15.69
14.25
12.73
15.18
30
17.22
15.78
13.20
16.42
40
18.73
16.08
14.80
17.60
50
24.15
22.16
19.40
23.26
S.Ed
1.65
1.56
1.40
1.60
CD (P = 0.05)
3.04
3.13
2.81
3.03
NZ No zone of inhibition ; *Values are average of three replications
Pseudomonas
aeruginosa
NZ
12.70
13.58
14.26
15.18
20.72
1.41
2.83
J. Bhuvaneswari/ Life Science Archives (LSA), Volume 1, Issue 4, Page 257 - 263, 2015
261
Pseudomonas
aeruginosa
+++
++
++
+
+
-
J. Bhuvaneswari/ Life Science Archives (LSA), Volume 1, Issue 4, Page 257 - 263, 2015
Yersinia enterocolitica, Campylobacter jejuni,
Flavobacterium sp., Shewanell sp. and
Enterobacter aerogenes (Dilabaghi and Sharma,
1996).
The
biopreservative
activity
of
Lactobacillus acidophilus strains was reported by
Kanatani et al. (1995). They reported that
Lactobacillus acidophilus produced bacteriocin
that was active against different species of
Enterococcus casseliflavus, Lactobacillus lactis,
Lactobacillus acidophilus, Pediococcus sp.,
Streptococcus
pyogenes
and
Listeria
monocytogenes. The inoculation of Listeria
monocytogenes in dairy foods as starter culture
can be used to prevent the proliferation of dairy
product spoilage bacteria which may cause
degradation of milk products that was exposed to
high temperature.
The inhibitory effect of heat stability of
crude bacteriocin (extracted from Lactobacillus
lactis and Lactobacillus acidophilus) at various
time intervals and different temperatures against
dairy product spoilage bacteria were studied. The
inhibitory activity was unaffected by heating upto
120 C for 40 min and 70 % of activity retained
after heat treatment at 120 C for 15 min. More
inhibition was noted in 15 min time interval at 20
C followed by 40 C, 60 C, 80 C, 100 C and
120 C. Savitha Janakiraman et al. (2012) reported
that the activity of crude bacteriocin in
Lactobacillus acidophilus and Lactobacillus lactis
after heat treatment at 121 0C for 15 min.
Therefore, it could be grouped under heat stable
low molecular weight bacteriocin. The finding of
the present research was matched with the results
which were proposed by Savitha Janakiraman et
al. (2012).
4. References
1) Abee, T. 1995. Pore-forming bacteriocins
of Gram-positive bacteria and selfprotection mechanisms of producer
organisms. FEMS Microbiological Letters,
19: 1-10.
2) Aymerich, T., M.G. Artigas, M. Garriga,
J.M. Monfort and M. Hugas. 2000.
262
J. Bhuvaneswari/ Life Science Archives (LSA), Volume 1, Issue 4, Page 257 - 263, 2015
11) Nilsson, L., Y. Chen, M. L. Chikindas, H.
H. Huss, L. Gram and T. J. Montville.
2000. Carbon dioxide and nisin act
synergistically on Listeria monocytogenes.
Applied and Environmental Microbiology,
66: 769 - 774.
12) OKeeffe, T., H. John and C. Hill. 1999.
Bacteriocins:
Potential
in
Food
Preservation.http://www.foodsci.cornell.ed
u/fs406/BACTERIOCINS.doc. 20 Feb.,
2004.
13) Ogunbanwo, S. T and B. M. Okanlawon.
2008.
Bio-preservative activities of
Lactobacillus
acidophilus
U1during
fermentation of fresh minced goat meat.
Journal of Applied Biosciences,12: 650 656.
14) Ray, B and M. Daeschel. 1992. Food Biopreservatives of Microbial Origin. C R C
Press Boca Ratron Ann Arbor London
Tokyo Florida: 3-172.
15) Saranraj, P and M. Geetha. 2012.
Microbial spoilage of Bakery products and
its control by preservatives. International
Journal of Pharmaceutical and Biological
Archives, 3 (1): 204 - 214.
16) Savitha Janakiraman and Malini Maria.
2012. Detection of heat stable bacteriocin
from
Lactobacillus
acidophilus
NCIM5426 by liquid chromatography/
mass spectrometery, 3: 2325 - 2332.
17) Schillinger, U., B. Becker, G. Vignolo and
W.H. Holzapfel. 2001. Efficiency of nisin
in combination with cultures against
Listeria monocytogenes Scott A in tofu.
International
Journal
of
Food
Microbiology, 71: 159-168.
18) Solomakos, N., A. Govaris, P. Koidis and
N. Botsoglou. 2008. The antimicrobial
effect of thyme essential oil, nisin and their
combination against Escherichia coli
O157:H7 in minced beef during
refrigerated storage. Meat Science, 80: 159
- 166.
19) Tagg, J.R and M.C. Given. 1971. Assay
system
for
bacteriocins.
Applied
Microbiology, 21(5): 943.
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