InternationalJournal of Environment, Agriculture and Biotechnology (IJEAB)
https://dx.doi.org/10.22161/iieab.46.29
Vol-A, Issue-6, Nov-Dec- 2019
ISSN: 2456-1878
Antibacterial Activities of Lactic Acid Bacteria
from Langsat Fruit {Lansiumdomesticum) against
Phatogenic Bacteria and Spoilage Bacteria
Helen J. Lawalata*, Mariana Rengkuan, Utari Satiman
Laboratory of Biology, Faculty of Mathematics and Natural Science, State University of Manado, Indonesia
‘Corresponding author : Lawalata_helen@yahoo.com
Abstract — Langsat fruit (Lansiumdomesticum) sample collected from 4 district in Minahasa, North Celebes. Lactic
Acid Bacteria (LAB) isolates were found at that samples. The study aim to examinedthe antibacterial activity of
Lactic acid bacteria (LAB) from Langsat fruit sample against pathogenic bacteria Escherichia coli (Gram Negatrive
Bacteria) and spoilage bacteria Staphylococcus aureus (Gram Positive Bacteria) using agar well diffusion
method.The isolation result obtained 42 isolates of LAB namely 10 isolate were obtained from Southeast Minahasa
district, 8 isolates were obtained from the southern district of Minahasa, 12 isolates were obtained from from the
Minahasa district, and 12 isolates were obtained from North Minahasa district. Fourty two isolates LAB identified
as Lactobacillus genera based on profile matching method were effective against bacteria indicator (pathogenic
and spoilage bacteria). The antibacterial activity showed through the formation of a clear zone around well that
containing LAB isolates. The diameter inhibition of LAB isolates ranged from 3,0 - 17,0 mm. Overall, the isolated
LAB showed the remarkable inhibitory effect against both Gram positive (spoilage bacteria) and Gram negative
(pathogenic bacteria). However, the spectrum of inhibition was different for the isolates tested. These results
suggest that this potent isolates could be used as a natural biopreservativesagent.
Keyword —Langsat fruit, LAB, Antibacterial, iMctobacillus, well diffusion.
1. INTRODUCTION
Langsat Fruit (Lansiumdomesticum) is a tropical fruit
that will only grow well in tropical regions such as Indonesia
and is an endemic fruit in Minahasa, North Sulawesi
Province. Langsat fruit has benefits and uses for the human
body because it has a high nutritional content. Fresh fruits
have a limited shelf life and are easily damaged. This is
caused by spoilage microorganisms found in these fruits [16].
Likewise with Langsat fruit, although it has benefits for the
human body, this fruit decomposes more quickly when
picked from the tree.
Fruits are a substrate for bacterial growth because they
have high carbohydrate content and high organic acids. LAB
are found in environments that are rich in nutrients such as
meat, vegetables, fish, fruits, fermented products and so on.
Some studies have succeeded in obtaining LAB in fruits and
vegetables including Markissa Fruit Fermentation [9], Dried
Fruit (Raisins and Fig) from Morocco [1], Ackee fruit from
Nigeria [6], Cabbage Juice [15],Mango fruit [4];[5], and
Cayene pepper fermentation[l 1].
Lactic acid bacteria also produce metabolites and
some of them have antibacterial activity including organic
acids (lactic acid, acetic acid, propionic acid, sorbic acid,
benzoic acid), hydrogen peroxide, bacteriocin and
diacetyl[3], but some strains capable of synthesizebacteriocin
antimicrobial compounds [2]. Antimicrobial compounds
produced by lactic acid bacteria have the potential as natural
preservatives to improve the safety of food products.
This study aims to isolate LAB which has antibacterial
activity fromLangsat Fruits from Minahasa, North Sulawesi.
II. MATERIALS AND METHODS
Sampling : Langsat fruit samples were collected from
4 districts in Minahasa, North Sulawesi Province, namely
Southeast Minahasa districts (Mitra), South Minahasa
districts (Minsel), Minahasa districts (MI), and North
Minahasa districts (Minut).
Isolation and Purification Lactic Acid Bacteria
The initial stage of LAB isolation from Langsat fruit
was enrichment. 10 g of Langsat fruit sample was mashed
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InternationalJournal of Environment, Agriculture and Biotechnology (IJEAB)
https://dx.doi.org/10.22161/iieab.46.29
Vol-A, Issue-6, Nov-Dec- 2019
ISSN: 2456-1878
and put in a 100 ml erlenmeyer containing MRS broth
medium pH (5.5), then grown at room temperature for 2
days.
LAB isolation was carried out using the pour plate
method. Take 1 ml of fermented Langsat fruit then put into a
test tube containing 9 ml of sterile Aquades (10 ' dilution).
Dilution is continued until the 10'^ dilution series, then from
each series of dilutions 10'^, 10’® and 10'^, 1 ml of sample is
taken and poured into a petri dish and added with MRSA +
1% CaC03. Incubation is carried out at 37 ° C for 2-3 days
until a clear zone is formed around the bacterial colony. Each
colony which has different morphological appearance and
clear zone formed is isolated and then purified [7]. LAB
colonies were purified on the MRS medium so that by the
quadrant streak method until separate colonies were obtained
[ 8 ].
Cell morphology. Gram staining and catalase test,
motility, non-spore forming were performed as a preliminary
screening forLAB.The selectedLAB were maintained as
stock cultures at -80 °C in 10% skim milk and 20% glycerol.
Antibacterial Activity of Lactic Acid Bacteria
The antibacterial activity test uses the well diffussion
assay against test bacteria (£’.co//ATCC35218 andS.aureus
ATCC 25923). Test bacteria were taken as much as 15 ml
and then inoculated on Nutrien soft agar (0,75% agar) for
each bacterium of 50 pi in petridish inoculated by pour plate.
The solid medium was perforated using a 6 mm diameter
perforator.MRS hard agar poured on petri dish and allow to
solidify, then overlaid with nutrient broth were prepared
previously and then in place at a temperature of 4 ° C for 1
hour.Culture of LAB was taken as much as 50 pi and
inserted into the wellbore. Petridish that have been inoculated
with testbacterial were incubated at 37°C for 24 hours. After
that, the inhibitory zone produced and measured based on the
diameter of the antimicrobial area [8].
III. RESULT AND DISCUSSION
Isolation of Lactic Acid Bacteria
Samples of LangsatFruit were collect from 4 district
of Minahasawere used for isolation of LAB. 50 isolates
ofbacteria-acid producing that produce clear zones around
the colony. The formation of a clear zone occurs because
CaCOsin the MRS medium is to he dissolved by acids
produced of bacteria acid production. After being confirmed
as LAB, 42 isolates of LAB were obtained(10 isolates were
obtained from Southeast Minahasa district, 8 isolates were
obtained from the southern district of Minahasa, 12 isolates
were obtained from from the Minahasa district, and 12
isolates were obtained from North Minahasa district).All
these isolates were gram positive, rods or cocci, appeared
singly, in pair, chain, tetrad. Cell were non motile and
nonsporing, they gave negative reaction for catalase. These
strains were then classified into genus level using profile
matching method. Based on the profile matching method that
42 isolates were putatively identified as genus Lactobacillus.
Table 1. Screening of lactic acid bacteria isolates into
genera level by profile matching method.
Character
Isolate
Lactobacillus*
Amount of isolates
42
Gram stain
-H
+
Shape
Rod
Rod
Cell arrangement
Single, pair and
chain.
Single, pair and
chain
Production gas
from glucose
-
-
Catalase
-
-
Spore formation
-
-
Motility
-
-
Homofermentati
Homofermentati
Fermentation type
ve
ve
*) Bergey’s Mannual of Systematic Bacteriology
AntibacterialActivity of Lactic Acid Bacteria
The antibacterial activity of LAB isolates (culture)
were tested against pathogenic bacteria and spoilage bacteria
are summarized in (Table 2) by using agar well diffusion
assay, and figure (1) illustrate the zones of inhibition against
pathogenic bacteria and spoilage bacteria under study. The
diameters of the inhibitionzones were varied it ranged
between 2,0 to 17,0 mm. All of LAB isolate haveinhibitory
activity againts pathogenic bacteria and spoilage bacteria.
IsolatesLMT2, LMS8, LMI7, and LMU2 have the highest
diameter of inhibition zones 16,5-17,0 mm (S.aureus).
Isolates LMT2, LMS8, LMI7, and LMU7 have the highest
diameter of inhibition zones 7,0-8,0 mm (E.coli)
thisrevealed that the LAB inhibited all the pathogenic
bacteria and spoilage bacteria tested.
Previous studies have shown that most of the
selected strains of lactic acid bacteria had good antagonistic
activity against pathogens including Escherichia coli [12].
According to [13] whose mentioned that inhibition was
scored positive if the width of the clear zone around the
colonies of the producer strain was 0.5 mm or larger.
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InternationalJournal of Environment, Agriculture and Biotechnology (IJEAB)
https://dx.doi.org/10.22161/iieab.46.29
Vol-A, Issue-6, Nov-Dec- 2019
ISSN: 2456-1878
Generally the antibacterial components produced by LAB negative and the same was stated by [14].
can inhibit the growth of gram-positive bacteria and gram-
Tabel 5. Selection of LAB Isolates based on antagonistic activity (diameter of the clear zonaproduced againstindicatot bacteria)
No
Isolate Code
S.coh'ATCC 35218 (mm)
St. aureus ATTC 25923 (mm)
Southeast Minahasa
1
LMTl
3,0
13,2
2
LMT2
7,0*
16,5*
3
LMT3
6,0
6,0
4
LMT4
3,0
16,0
5
LMT5
2,0
12,5
6
LMT6
3,0
11,2
7
LMT7
5,0
6,0
8
LMT8
5,0
14,5
South Minahasa
9
LMSl
3,0
15,7
10
LMS2
2,0
16,3
11
LMS3
5,0
13,5
12
LMS4
2,0
7,5
13
LMS5
3,0
11,2
14
LMS6
3,0
11,2
15
LMS7
6,0
13,7
16
LMS8
7,0*
17,0*
17
LMS9
5,0
15,7
18
LMSIO
3,0
11,5
Minahasa
19
LMIl
2,0
10,0
20
LMI2
3,0
3,0
21
LMI3
5,0
10,0
22
LMI4
4,0
11,0
23
LMI5
4,0
13,7
24
LMI6
3,0
13,7
25
LMI7
7,0*
15,0*
26
LMI8
6,0
11,0
27
LMI9
4,0
12,0
28
LMIIO
5,0
14,0
29
LMIll
3,0
6,0
30
LMIl 2
3,0
6,0
North Minahasa
31
LMUl
6,0
16,0
32
LMU2
5,0
17,0*
33
LMU3
3,0
10,0
34
LMU4
3,0
11,0
35
LMU5
2,0
11,7
36
LMU6
6,0
11,7
37
LMU7
8,0*
12,0
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InternationalJournal of Environment, Agriculture and Biotechnology (IJEAB)
https://dx.doi.org/10.22161/iieab.46.29
Vol-A, Issue-6, Nov-Dec- 2019
ISSN: 2456-1878
38
LMU8
6,0
15,0
39
LMU9
4,0
12,0
40
LMUIO
3,0
12,0
41
LMUll
4,0
10,0
42
LMU12
3,0
10,0
Fig.l: Antibacterial activity of LAB against pathogenicbacteria and spoilage bacteria.
(1). Escherichiacoli, (2). Staphylococcus aureus
The activity on inhibition of variety of bacteria by LAB due
toa combination of many factorsproduced by LAB e.g.
production of lactic acid whichreduce pH of Langsat fruit
and also other inhibitory substances such as bacteriocins,
hydrogen peroxide, diasetylwhich are responsible for the
mostantimicrobial activity [10].
IV. CONCLUSION
The results were obtained42isolates
oflacticacidbacteriawhichcan begroupedinto
thegeneiaLactobacillus. All ofisolates
LABhaveantibacterial activityagainstthe growth
ofpathogenic bacteriaandspoilage bacteria. LMT2, LMS8,
LMI7,LMU2, and LMU7 have the highest antibacterial
activity.
REFERENCES
[1] Askari, Al, Galal.,Kahouadji, A., Khedid, K., Charof, R, and
Mennane, Z. 2012. Screening of Lactic Acid Bacteria
Isolated from Dried Fruit and Study of Antibacterial
Activity. Middle-East Journal of Scientific Research, 11(2) :
209-212.
[2] Cizeikiene, D.,Juodeikiene, G.,Paskevicius, A.,Bartkiene, E.
2013. Antimicrobial Activity of Lactic Acid Bacteria
Againts Pathogenic and Spoilage Microorganism Isolated
from Pood and their Control in Wheat Bread. Journal Pood
Control, 31 : 539-545.
[3] Dalie, D,K,D., Deschamps, A,M., and Richard-Porget. 2010.
Lactic Acid Bacteria-Potencial for Control of Mould Growth
and Mycotoxin: review, Food Control, 21: 370-380.
[4] Fitriyani, I. 2010. Isolasi,
KarakterisasidanIdentifikasiBakteriAsamLaktat (BAL)
dariBuahMatang yang
berpotensiMenghasilkanAntimikrobia. Skripsi. UIN,
Yogyakarta.
[5] Ibrahim, A., FridayantidanDelvia, F. 2015.
IsolasidanIdentifikasiBakteriAsamLaktat (BAL)
daribuahMangga (Mangiferaindica L.)
JumalllmiahManuntung, 1(2):159-163.
[6] Lawal, R.T., Oyeleke, G.O.,Ishola, A.D.,andAkinsurojo,
M.O. 2018. Determination of Physiochemical Properties and
Lactic Acid Bacteria Presence in Ackee (Blighiasapida)
Fruit. International Journal of Environment, Agriculture and
Biotechnology (IJEAB), 3(3): 1079-1082.
[7] Lawalata V.N. 2012. Rekayasa proses
ekstrasikulitbuahlangsat {LansiumdomesticumvarLansat)
sebagaibahanantibakteridanantioksidan [disertasi]. Bogor
(ID): Institut Pertanian Bogor .
[8] Lawalata H.J. 2012.
Keanekaragamanbakteriasamlaktatpenghasilantimikrobiasela
ma proses fermentasibakasang [disertasi].
UniversitasGadj ahMada. Yogyakarta.
[9] Nurisva Y,M,S. 2013. Isolasi, Karakterisasi, Dan Identifikasi
DNA bakteriAsamLaktat (BAL) Yang
BerpotensiSebagaiAntimiki'oba Dari
FermentasiMarkisaKuning (Passifloraedulisvar.flavicarva).
J.KimiaUniversitasAndalas, 2(2) : 81-91.
[10] Ogunbanwo ST. Functional properties of lactic acid bacterial
isolated from ogi and fufu. Two Nigerian fermented foods. J
food Sci. 2005;27:14-21
[11] Rustan, I.Reskia. 2013.
StudilsolasidanIdentifikasiBakteriAsamLaktatdariFermentas
iCabaiRawit (Capsicum frutencens L.). Skripsi
,JurusanTeknologiPertanian, UniversitasHasanuddin,
Makassar.
www.iieab.com
Page I 1809
InternationalJournal of Environment, Agriculture and Biotechnology (IJEAB)
https://dx.doi.org/10.22161/iieab.46.29
Vol-A, Issue-6, Nov-Dec- 2019
ISSN: 2456-1878
[12] Savino,F., Cordisco, L., Tarasco V, Locatelli E, Di Gioia D,
Roberto Oggero R and Matteuzzi D. 2011. Antagonistic
Effect of Lactobacillus strains against Gas-
producingColiform Isolated from Colicky Infants. BMC
Microbial II: 157.
[13] Schillinger U, and Lucke F. Antimicrobial activity of
Lactobacillusssks isolated from meat. J Appl Environ
Microbiol. 1989;55 :1901-1906.
[14] Theron MM, and Lues JER (2011) Organic Acids and Food
Preservation. United State: CRC Press. Him: 273.
[15] Utama, C.S., Zuprizal.,Chusnul Hanim.,Wihandoyo. 2018.
Isolasidan Identifikasi Bakteri Asam Laktat Selulolitik yang
berasaldari Jus Kubis Terfermentasi. Jurnal Aplikasi
Teknologi Pangan, 7(1) :l-6.
[16] Wahyu, N. 2012. Peranan Bakteri Asam Laktat Terhadap
Buahdan Sa 5 Tjran. http://apriliana.wahvu.blogspot.com [25
Agustus 2019].
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