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Antimicrobial, bacteriocin, fermented foods, LAB, preservative
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Ogundipe A, Ogunnusi TA and Akpor OB. Isolation and Optimization of Bacteriocin-Producing Lactic Acid Bacteria from Nigerian Fermented Foods [version 1; peer review: awaiting peer review]. F1000Research 2025, 14:1467 (https://doi.org/10.12688/f1000research.174479.1)
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Research Article
Isolation and Optimization of Bacteriocin-Producing Lactic Acid Bacteria from Nigerian Fermented Foods
[version 1; peer review: awaiting peer review]
PUBLISHED 27 Dec 2025
OPEN PEER REVIEW
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Abstract
Corresponding author:
Abigael Ogundipe
Competing interests:
No competing interests were disclosed.
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The author(s) declared that no grants were involved in supporting this work.
Copyright:
© 2025 Ogundipe A et al.
This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
How to cite: Ogundipe A, Ogunnusi TA and Akpor OB. Isolation and Optimization of Bacteriocin-Producing Lactic Acid Bacteria from Nigerian Fermented Foods [version 1; peer review: awaiting peer review]. F1000Research 2025, 14:1467 (https://doi.org/10.12688/f1000research.174479.1)
First published: 27 Dec 2025, 14:1467 (https://doi.org/10.12688/f1000research.174479.1)
Latest published: 27 Dec 2025, 14:1467 (https://doi.org/10.12688/f1000research.174479.1)
1. Introduction
Lactic acid bacteria (LAB), such as Streptococcus, Enterococci, Lactococcus, Lactobaccilli and Leuconostoc are gram-positive, non-respiring, non-sporing rods or cocci that form fermentative end products of carbohydrates such as lactic acid. They do not have cytochromes, and derive their energy through substrate-level phosphorylation. They have been employed in the production of sorghum beer, all fermented milk, cassava (to make garri and fufu) and most fermented (so-called pickled) vegetables. They also ferment sugars and are nutritionally fastidious, thus requiring many vitamins, amino acids, purines, and pyrimidines.
Lactic acid bacteria are naturally found in various raw materials such as milk and meat. They are employed as either natural or chosen starter cultures in food fermentation, in which acidification is achieved through the synthesis of lactic acids. They prevent food spoilage and pathogens by producing organic acids. They constitute significant populations of microorganisms in good fermentation and have a positive impact on the nutritional organoleptic properties and shelf stability of products. They are commonly considered safe and can serve as natural competitive inhibitors or starter cultures under controlled conditions. They can synthesize minute organic compounds that add aroma and produce certain organoleptic properties in food products.
Silva et al. (2018) stated that bacteria produce proteins known as bacteriocins, which inhibit the growth of similar or closely related strains of bacteria. Numerous antimicrobial compounds, such as organic acids, diacetyl, acetone, hydrogen peroxide, reuterin, anti-fungal peptides, and bacteriocins, have been reported to be produced by LAB. The presence of antagonistic properties of lactic acid bacteria (LAB) in terms of their safe history of application in traditional fermented food products makes them highly attractive for use as biopreservatives.
Food processing using biotechnology can assist in enhancing food safety because it allows the replacement of food preservatives with non-toxic biopreservatives. In this regard, the production of bacteriocin, a natural preservative of non-pathogenic LAB obtained locally (indigenously) from fermented food, may be useful in the food manufacturing industry as an alternative to microbial spoilage and food safety, as well as for enhancing the microbial quality of food. Thus, the purpose of the current study was to produce and characterize bacteriocins from lactic acid bacteria obtained from locally fermented food samples (fufu, ogi, and iru). In this study, crude bacteriocin (crude supernatant) was produced under varying culture conditions from lactic acid bacteria isolated from Nigerian fermented food to preserve fruit juice.
A number of emerging spoilage microorganisms are also of great concern to the fruit juice industry, and Alicyclobacillus acidoterrestris has been reported to exist in various juices and juice products with reported ranges of 14.7 to 18.3. Propionibacterium cyclohexanicum and heat-resistant species of mycelial fungi, such as Byssochlamys fulva, B. nivea, and Neosartorya fischeri, and species of Talaromyces have also been noted to spoil fruit juices. Outbreaks of foodborne diseases linked to fruit juices are enormous. In consideration of the threat posed by spoilage and pathogenic microorganisms in the fruit juice industry, as well as in the general health authorities, various guidelines have been released by national food standard agencies.
Fruit juices and drinks are popular products that contain chemical preservatives that help them last longer in shelves, such as sodium benzoate and potassium sorbate. However, the demand for safe and fresh foods with no preservatives generated through chemical reactions is of increasing interest for the implementation of natural food preservatives. Some food products have been preserved using natural preservatives such as bacteriocins, organic acids, essential oils, and phenolic compounds.
Acid-tolerant bacteria (Bacillus subtilis, Bacillus proteolyticus, Klebsiella pneumoniae, Pseudomonas aeruginosa and Pseudomonas fuscoginae), yeast, and molds cause fruit juice spoilage with unacceptable odor and flavor. The use of natural preservatives is important because applying chemical preservatives to food may cause health issues (allergies and cancer risks). Isolation of bacteriocins from Nigerian local fermented foods can be difficult because of factors such as low stability and susceptibility to proteolytic enzymes. For LAB strains to be isolated from Nigerian local fermented foods to produce the most bacteriocins, it is important to optimize the culture conditions. Low production yields, unstable activity in acidic juice matrices, and unclear effects on sensory quality and shelf life hinder the isolation of LAB strains for the production of bacteriocin applied in fruit juice biopreservation when optimization is absent of optimization. The aim of this study was to optimize bacteriocin production by LAB from Nigerian fermented foods and evaluate its effectiveness in fruit juice preservation.
2. Materials and method
2.1 Isolation of lactic acid bacteria from Nigerian fermented foods
Three Nigerian traditional fermented foods (iru, fufu, and ogi) were used to isolation of lactic acid bacteria (LAB). The foods were purchased from a local market in Ado Ekiti, Ekiti State, Nigeria. The foods were aseptically sealed in food-grade sampling bags and transported carefully in containers with ice packs to maintain acceptable conditions for the transportation of samples to the laboratory for isolation.
Isolation of LAB isolates was performed using de Man Rogosa Sharpe Agar (MRS) supplemented with 0.05 g fluconazole. Fifteen grams of each food sample was dissolved in 150 mL sterile normal saline (0.85 w/v NaCl) and serially diluted 10-fold in sterile distilled water. LAB were isolated on a standard pour plate. Fifteen milliliters of sterile MRS Agar, in a known dilution in a 1 mL sterile Petri dish, was precooled at 45°C. The plates were left to dry and were placed in an incubator at 35°C ± 2°C for 48 h. Colonies of various morphologies were randomly selected after incubation with a flamed platinum wire loop, streak plated, and subcultured on MRS agar plates to obtain pure colonies. The respective pure colonies were kept as slants in bottles with sterile MRS agar and kept in a refrigerator (4°C ± 2°C) until required.
2.2 Preliminary antibacterial screening
Preliminary antibacterial screening was carried out using (48 h old broth) cultures of the LAB strains against selected pathogens (Salmonella typhi, Bacillus subtilis, Bacillus proteolyticus, Xanthomonas Campestris, Pseudomonas aeroginosa, Klebsiella pneumonia, and Pseudomonas fuscoginae) using the agar-well diffusion method as reported by (Akpor et al., 2022; Reuben et al., 2019). To 200 mL sterile molten nutrient agar in a 250 mL capacity Erlenmeyer flask, 1 mL of 24 h old broth culture of the respective test pathogen was inoculated and swirled gently and mixed thoroughly several times. Following mixing, 20 mL of cooled molten agar with the inoculated pathogen was dispensed in sterile petri dishes and left to solidify. Following solidification, four holes were bored on each plate, using a sterile cork borer (5 mm diameter) before adding enough inoculum of the 48 h old broth cultures of LAB to fill a bored hole and left to disperse. The plates were left to dry and set in an incubator at 35°C ±2°C for 24 h to observe clear zones. The antibacterial potential of the isolate was observed in the presence of a clear zone around the bored hole, without any growth. The degree of inhibition was measured using a meter ruler in millimeters.
2.3 Pathogenicity screening of isolates
Pathogenicity screening of isolates was performed by plating the LAB strains on sterile nutrient agar with 5% of human blood been added. The plates were incubated at 37°C for 24 h and the red blood cell lytic activity surrounding the LAB colonies was reported. The zones that were green around the colonies were regarded as alpha hemolysis, clear zones around the colonies were regarded as β-hemolysis, and those with no zones around were regarded as gamma hemolysis on the blood agar plates. Only strains with gamma hemolysis were considered safe for further use (Mangia et al., 2019).
2.4 Characterization of isolates
All isolates that passed pathogenicity screening were characterized using 16 S rRNA polymerase chain reaction (PCR) and Sanger partial gene sequencing.
DNA was extracted according to the protocol described by Wang et al. (2011). Briefly, single colonies of a particular isolate to 1.5 mL of liquid medium and allowed to grow in a shaker after 48 h at 28°C. Centrifugation of the cultures was performed at 4600 g for 5 min, and the resulting pellets were resuspended in 520 μL of Tris-EDTA buffer (10 mMTris-HCl, 1 mM EDTA, pH 8.0). Then, 15 μL of Sodium Dodecyl Sulfate (20% solution) and 3 μl of Proteinase K (20 mg/ml) were added to the suspension, the suspension was incubated at 37°C for 1 h, followed by the addition of 100 uL of 5M M sodium chloride and 80 μL of 10% cetyltrimethylammonium bromide solution in 0.7M Sodium chloride, which was vortexed. The mixture was then incubated at 65°C for 10 min and placed on ice for 15 min. The mixture was incubated on ice (5 min) and centrifuged at 7200 g (adding an equal amount of chloroform: isoamyl alcohol (24:1)). This was then mixed with aqueous phase in another tube and isopropanol (1: 0.6) was added and the DNA precipitated at -20°C for 16 h. Centrifugation of the DNA was undertaken for 10 min at 13000g and g, followed by DNA washing with 500 μL of 70% ethanol, left to air dry at room temperature for 3 h, and then by the final miscible addition of 50 μL Tris-EDTA buffer.
The PCR sequencing cocktail mixture comprised 10 μL 5x GoTaq colourless reaction, 3 μL of 25mM MgCl2, 1 μL of 10 mM mix of dNTPs, 1 μL of 27F 5′- AGA GTT TGA TCM TGG CTC AG-3′ and - 1525R, 5′-AAGGAGGTGATCCAGCC-3′ primers and 0.3 units of Taq DNA polymerase (Promega, USA). Up to 42 μL, the mixture was mixed with 8 μL of sterile distilled water DNA template.
PCR was carried out in a GeneAmp 9700 PCR System Thermalcycler (Applied Biosystem Inc., USA) with a PCR profile that consisted of an initial denaturation at 94°C denaturation for 5 min. This was followed by 30 cycles which included of 94°C/30 for s, 50°C for 60 s and 72°C/1 90 s, and termination at 72°C/10 for min before chilling at 4°C GEL (Innis et al., 2012).
Determination of the integrity of the amplified gene was determined by placing the fragment on a 1% agarose gel that was run to confirm amplification. The buffer (1XTAE buffer) was prepared and it was then applied to the preparation of 1.5% agarose gel by boiling the suspension in a microwave oven for 5 min. The molten agarose was left to dry and cool to 60°C and then stained with 3 ml of 0.5 g/mL ethidium bromide, which is invisible ultraviolet (UV) radiation and emits visible orange radiation. The slots of the casting tray were loaded with a comb, molten agarose was poured into the casting tray, and the gel was left to harden for 20 min to establish the wells. The gel tank was filled gradually by pouring × 1XTAE buffer into the gel. Each of the PCR products (4 mm) was mixed with 2 mL of 10X blue gel loading dye and loaded into the wells when the 100 bp DNA ladder was loaded into well 1. The gel was subjected to electrophoresis for 45 min at 120 V, followed by UV transillumination, and photographs were taken. The PCR products were assessed by comparing the mobility of a 100 bp molecular weight ladder that was stained and placed into a gel with the experimental samples.
To eliminate the PCR reagents when the gel integrity was guaranteed, 7.6 μL of sodium acetate 3M and 240 μL of 95% ethanol were added to each approximately 40 μL of PCR amplified product in a new sterile 1.5 μl tube Eppendorf, mixed thoroughly by vortexing, and stored at -20°C at least 30 min. The suspension was centrifuged at 13000 × g and 4°C for 10 min, the supernatant was removed (invert tube on trash once), and the pellets were washed by placing 150 μL of 70 ethanol into the mixture and centrifuging at 7500 G 15 min at 4°C. The resulting supernatants were removed, inverted tubes were incubated on tissue paper, allowed to dry in a fume hood for 10-15 min at room temperature, resuspended in 20 μL of sterile distilled water, and stored at -20°C prior to sequencing. To verify the existence of the purified product, the purified fragment was resolved on a 1.5% agarose gel that was employed to run at 110 V for 1 h before quantification with Nanodrop (model 2000-thermo scientific).
The amplified fragments were sequenced on a Genetic Analyzer 3130xl sequencer (Applied Biosystems) in accordance with the manufacturer’s instructions. The sequencing kit used was the BigDye terminator v3.1 cycle sequencing Kit. To carry out the genetic analysis, MEGA 6 and BioEdit software were used.
2.5 Optimization of cell-free supernatant and crude bacteriocins
2.5.1 Effect of incubation time
Pure cultures of LAB strains were transferred into MRS broth and incubated at 37°C for (24, 48, 72, 96, 120, 144, or 168) h, respectively. After incubation, centrifugation was done on the broth culture at 4,000 rpm for 10 min. The cell-free supernatant (CFS) was then transferred into sterile containers for antibacterial analysis against five selected bacterial pathogens (Bacillus subtilis, Bacillus proteolyticus, Pseudomonas aeruginosa, Klebsiella pneumonia, and Pseudomonas fuscoginae) using the agar well diffusion procedure and measurement of zones of inhibition, as described in section 2.2.
2.5.2 Effect of pH
To determine the effect of pH, CFS/crude bacteriocin was adjusted to various pH levels of 3, 4, 5, 6, 7, 8, 9 and 10, respectively, using 1 M HCl and/or I M NaOH, and a pH meter was used to measure the pH level. An overnight broth culture of the bacterial pathogen was mixed with the prepared nutrient agar and poured into sterile petri dishes. A sterile cork borer was used to make wells of 6 mm diameter and loaded with 50 μL of the CFS or crude bacteriocin at various pH, then the plates were incubated for 24 h at 37°C, using the agar well diffusion procedure and measurement of zones of inhibition as described in section 2.2.
2.5.3 Effect of organic solvents
The CFS/crude bacteriocins of each LAB strain were incubated with respective organic solvents (10%) which include acetone, ethanol, ethyl acetate and methanol. The antibacterial activity of CFS and crude bacteriocins was determined using the agar well diffusion procedure and measurement of zones of inhibition, as described in section 2.2.
2.5.4 Effect of temperature
For The effect of temperature on CFS/crude bacteriocins was investigated by subjecting them to heat treatment at various temperatures at (30°C, 40°C, 50°C, 60°C, 70°C, 80°C, and 121°C) for 30 min, respectively. Following the treatment, all the CFS and crude bacteriocin samples went for a 24 h incubation period at 37°C. The antibacterial activity of CFS and crude bacteriocins was determined using the agar well diffusion procedure and measurement of zones of inhibition, as described in section 2.2.
2.5.5 Effect of ultra-violet radiation
To determine the effect of UV radiation, cell-free supernatants/crude bacteriocins were exposed to ultraviolet irradiation for different durations (20, 40, 60, and 80 min, and no radiation). The antibacterial activity of CFS and crude bacteriocins was determined using the agar well diffusion procedure and measurement of zones of inhibition, as described in section 2.2.
2.5.6 Effect of storage condition
The stability of the CFS/crude bacteriocins of LAB strains was observed under different storage conditions (freezing, refrigeration, and ambient conditions). The antibacterial potential of CFS and crude bacteriocins was determined using the agar well diffusion procedure and measurement of zones of inhibition, as described in section 2.2.
2.5.7 Effect of proteases
The CFS/crude bacteriocins were treated with proteases (trypsin and pepsin) in order to verify their protein structure as described by Zdolec et al. (2007). The proteases were prepared according to the manufacturer’s instructions. Following the treatment with pepsin and trypsin, the treated CFS was placed in an incubator for 30 min at 80°C. The antibacterial activity of CFS and crude bacteriocins was observed using the agar well diffusion procedure and measurement of zones of inhibition, as described in section 2.2.
2.5.8 Effect of concentration
Crude bacteriocins were prepared at concentrations ranging from 10 to 100 mg/mL. Preparation of concentration: For 10 mg/mL, 420 μL of crude bacteriocin was mixed with 4.58 mL of sterile distilled water. The higher the concentration, the higher is the mixture utilized. The antibacterial activity of CFS and crude bacteriocins was determined using the agar well diffusion procedure and measurement of zones of inhibition, as described in section 2.2.
3. Results
3.1 Test bacterial species
A total of 33 lactic acid bacteria (LAB) were isolated from Nigerian fermented foods (Iru, Ogi and fufu). Preliminary screening of the isolates for bacteriocin production yielded five LAB strains for a detailed study. Polymerase chain reaction and partial gene sequencing of the LAB strains revealed three strains of Lactobacillus plantarum, one strain of Lactobacillus paracasei and one strain of Lactobacillus acidophilus ( Table 1 and Figure 1).
Table 1. NCBI Blast showing the identity of the sequenced LAB strains.
Figure 1. Gel electrophoresis of the PCR amplified fragments of the bacterial isolates. MK represents ‘marker’ while ‘PSE’, ‘IRF’, ‘FUE’, ‘IPE’ and ‘PMD’ indicate the LAB strains.
3.2 Effect on cell free supernatant (CFS)
Culture age
Generally, peak inhibitory effects occur between 48 h and 120 h, after which activity wanes. Early cultures (24 h) produced moderate inhibition, whereas very old cultures (144–168 h) generally showed sharp declines or complete loss of activity ( Table 2).
Table 2. Effect of culture age on bacteriocin activity by cell-free supernatant of test LAB strains.
Among the test pathogens, K. pneumoniae and P. aeruginosa were the most frequently inhibited. In the case of B. subtilis, inhibition was intermittent and often zero beyond early time points, whereas P. fuscoginae and B. proteolyticus responses varied widely by LAB strain; for the LAB strains, L. plantarum (PV937063) showed the broadest spectrum, with strong inhibition (16–21 mm) against K. pneumoniae and P. aeruginosa at 24–72 h, with activity dropping markedly at 96 h but rebounding at 120 h for several of the pathogens ( Table 2).
In the case of L. paracasei (PV937064), a moderate early effect (24 h) with B. subtilis and B. proteolyticus was recorded with marked increases at 48–120 h, reaching up to 17 mm against P. aeruginosa and sustained activity against P. fuscoginae over 144 h. For the setup with L. plantarum (PV937062), a more selective activity was observed, showing the strongest and most consistent inhibition of P. fuscoginae and P. aeruginosa at 48 h and again at 120 h with little or no activity towards K. pneumoniae ( Table 2).
In addition, L. acidophilus (PV937061) showed the weakest overall activity with no detectable activity until 120 h, when modest zones (11 mm) appeared against K. pneumoniae, B. proteolyticus, and P. aeruginosa, then quickly declined. A narrow spectrum was observed with L. plantarum (PV937065), with early inhibition of K. pneumoniae and B. subtilis at 24 h (13 mm), and B. proteolyticus at 72 h ( Table 2).
pH
In general, the medium pH had a significant impact on the antimicrobial activity of the cell-free supernatant, with acidic to nearly neutral environments (pH 3–6) exhibiting higher inhibitory effects. Activity frequently decreased at alkaline levels (≥8), with some strains exhibiting abrupt decreases in inhibition against specific pathogens or total loss of inhibition ( Table 3).
Table 3. Effect of medium pH on bacteriocin activity by cell-free supernatant of test LAB strains.
Across the test pathogens, K. pneumoniae and P. aeruginosa were most consistently inhibited at the respective pH ranges, with zones of inhibition greater than 15 mm. When tested against P. fuscoginae and B. proteolyticus, the most varied and strain-dependent responses were observed at the respective pH values. However, substantial inhibition of B. subtiliswas observed at acidic and neutral pH.
Among the LAB strains, L. plantarum (PV937063) displayed the highest activity against the test pathogens, with zones of inhibition ranging from 13 to 22 mm. The observed inhibition in the presence of L. plantarum (PV937063) was evident across all pathogens and pH ranges, although the most stable activity was evident in acidic environments (pH 3–6). In addition, in terms of specific inhibition against a test pathogen, the highest activity was L. paracasei (PV937064) against K. pneumonia with maximal effects at pH 4-6, reaching 24 mm ( Table 3).
Organic solvents
Generally, all cell-free supernatants consistently showed no inhibitory activity against the test pathogens treated with ethyl acetate. The highest inhibitory activity was observed for ethanol-treated CFS, followed by methanol and, to a lesser extent, acetone ( Table 4).
Table 4. Effect of organic solvents treatment on bacteriocin activity by cell-free supernatant of test LAB strains.
Among the different CFS of the test LAB strains, K. pneumoniae was only inhibited by ethanol treatment. The highest levels of inhibition were observed in L. plantarum (PV937063, PV937062, and PV937065) (14.0 – 15 mm). Based on the activity of L. acidophilus (PV937061) and L. paracasei (PV937064) (12–14 mm), ethanol-treated CFS was observed to be the most potent inhibitor ( Table 4). Pseudomonas aeruginosa was inhibited by almost all LAB strains, particularly those treated with ethanol treatments of Lactobacillus plantarum (PV937063) and L. paracasei (PV937064), with inhibition zones of 19 mm ( Table 4).
Incubation temperature
In general, temperature affected the patterns of activity, with broad thermostability observed in some strains, whereas others displayed restricted or specific inhibition against the test pathogens. Among the test pathogens, P. fuscoginae, P. aeruginosa, and B. subtilis all showed similar inhibitory activities against L. plantarum (PV937063), with zones of inhibition that ranged from 10–15 mm at 30–60°C.
Generally, highest heat tolerance was observed at between 70 and 80°C, where inhibition against P. fuscoginae (14.5–15.0 mm) and P. aeruginosa (14.0 mm) was highest, although no inhibition against B. proteolyticus or K. pneumoniae. Among the CFS, the most selective trend was observed in L. paracasei (PV937064), with inhibition only against P. fuscoginae at the different temperature treatments, with highest zone of inhibition at 40°C (27.5 mm) and modest inhibition (10–13 mm) in other treatments ( Table 5).
Table 5. Effect of incubation temperature on bacteriocin activity by cell-free supernatant of test LAB strains.
On the other hand, L. plantarum (PV937062) showed strong inhibition against P. fuscoginae, P. aeruginosa, and B. proteolyticus at different temperature treatments, maintaining significant activity at sterilization (13.5–17.5 mm at 121°C). However, no activity was observed against B. subtilis. The highest inhibition was recorded for at 70°C and 80oC treatment with zones of inhibition 25.5 mm and 20 mm against K. pneumoniae and P. aeruginosa, respectively.
For L. acidophilus (PV937061), consistent and moderate inhibition was observed against most tested pathogens whileL. Plantarum (PV937065) showed lower range of antimicrobial activity at the respective temperature treatments, with strong activity against P. fuscoginae at 80°C (22.5 mm) and K. pneumoniae at 40°C (20 mm). However, activity was however less consistent at 121°C ( Table 5).
Ultra-violet (UV) radiation
Overall, the effect of ultra-violet (UV) irradiation on the antimicrobial activity of CFS showed distinct strain- and pathogen-specific responses. The highest inhibition was recorded in the presence of L. plantarum (PV937063), which showed strong activity against K. pneumoniae (20–30 mm), P. aeruginosa (20–30 mm), and B. subtilis (20–22 mm) after exposure to UV light for 60–80 min. However, no activity was observed against B. proteolyticusat after 40 min of UV exposure ( Table 6).
Table 6. Effect of ultra-violet radiation on bacteriocin activity by cell-free supernatant of test LAB strains.
In contrast, L. paracasei (PV937064) showed extreme sensitivity to UV treatment, with pathogen inhibition only noticeable during the first 40 min. In addition, L. plantarum (PV937062) showed remarkable inhibition against K. pneumoniae and P. aeruginosa (up to 21 mm), although no effect was observed against B. subtilis and B. proteolyticus. After 60 min of exposure, decreased activity was observed for L. plantarum (PV937065) against P. aeruginosa, whereas moderate inhibition was maintained against K. pneumoniae and B. proteolyticus (19–22 mm) ( Table 6).
Storage condition
Freezing and refrigerated storage conditions often resulted in partial or total loss of action, while ambient storage maintained the highest inhibition across the test pathogens ( Table 7).
Table 7. Effect of storage antibacterial activity of the cell free supernatants of the LAB strains.
Among the test pathogens, P. aeruginosa and P. fuscoginae were the most consistently inhibited, with inhibition zones sometimes exceeding 20 mm. This was followed by B. subtilis and K. pneumoniae, then B. proteolyticus which showed lesser or more varied inhibition ( Table 7).
Proteases
In general, treatment with pepsin completely eliminated the antibacterial activity against all strains and test pathogens. Although activity was also typically lost for most of the CFS, L. plantarum (PV937063) and L. paracasei (PV937064) retained little inhibition (10.5 mm) against P. aeruginosa ( Table 8).
Table 8. Effect of protease antibacterial activity of the cell free supernatants of the LAB strains.
3.3 Effect on crude bacteriocin
pH
Overall, acidic conditions (pH 3–4) did not affect the inhibitory activity of crude bacteriocins against the tested pathogens. However, as the pH increased towards neutral and alkaline conditions, inhibition generally decreased, and in several cases, the activity was terminated completely ( Table 9).
Table 9. Effect of pH on antibacterial activity of the test bacteriocins.
Among the tested strains, bacteriocins from L. plantarum (PV937063) showed the highest and most potent inhibitory activities, especially at pH 3 and 4, where the zones of inhibition reached 28–32 mm against K. pneumoniae and P. fuscoginae. The activity remained evident at near-neutral pH (pH 7). However, total loss of inhibition was observed against P. aeruginosa and B. subtilis, whereas at pH ≥ 9, activity against B. proteolyticus and B. subtilis was entirely decreased ( Table 9).
For the bacteriocin from L. paracasei (PV937064), medium but consistent inhibition against the test pathogens was observed, with the largest inhibition against B. proteolyticus at pH 3 (27.5 mm). For bacteriocin from L. plantarum (PV937062), more selective activity, particularly strong inhibition of P. fuscoginae (25 mm), was observed at pH 9. However, beyond pH 7, the activity against P. aeruginosa was absent ( Table 9).
In the presence of the bacteriocin from L. acidophilus (PV937061), the weakest overall activity was observed, with inhibitory effects restricted mainly to acidic pH, where zones reached 27.5 mm against B. proteolyticus and B. subtilis at pH 3. For L. plantarum (PV937065), medium and variable activity was observed against the test pathogens at the different pH treatments, with high activity at pH 8 against B. subtilis (25 mm) ( Table 9).
Organic solvents
In the presence of crude bacteriocin from L. plantarum (PV937063), ethyl acetate, methanol, and acetone treatments did not affect the antibacterial activity against K. pneumoniae, P. fuscoginae and P. aeruginosa. However, no inhibition was observed against B. pretolyticus when bacteriocin was treated with ethyl acetate, acetone, or methanol. Except for acetone treatment, all other solvent-treated bacteriocins from L. plantarum (PV937063) demonstrated inhibitory activity against B. subtilis ( Table 10).
Table 10. Effect of organic solvents on antibacterial activity of the test bacteriocins.
In the case of the bacteriocin from L. plantarum (PV937063), the highest inhibition was observed against K. pneumoniae (24 mm) and P. aeruginosa (22.5 mm), while there was a complete loss of activity against B. proteolyticus. In contrast, ethanol treatment substantially inhibited B. proteolyticus (14.5 mm) and B. subtilis (16.5 mm). In addition, medium but consistent activity was observed for bacteriocin from L. paracasei (PV937064) in the different treatments. Although ethyl acetate yielded little or no inhibition against P. fuscoginae and P. aeruginosa, methanol and acetone increased its activity against P. fuscoginae and P. aeruginosa (17 mm), with activity observed against B. subtilis (16.5 mm) following ethanol treatment ( Table 10).
Incubation temperature
Generally, temperature significantly influenced the bacteriocin activity, with the greatest inhibition occurring between 30 and 40°C and a steady decrease at higher temperatures. Some bacteriocins from some of the strains showed thermostable features, retaining remarkable inhibition up to 121°C ( Table 11).
Table 11. Effect of temperature antibacterial activity of the test bacteriocins.
Among the LAB strains, bacteriocins from Lactobacillus plantarum (PV937063) consistently produced the most dominant and largest inhibition, especially against Klebsiella pneumoniae and Pseudomonas fuscoginae (16–19 mm at 30–40°C), with retained activity at 121°C (10–18 mm). Bacteriocin from L. paracasei (PV937064) also showed stable activity across most pathogens at the different temperature treatments, maintaining inhibition up to 121°C. In contrast, although bacteriocin from L. plantarum (PV937062) was highly effective against Bacillus subtilis (27.5 mm at 30°C), less stable inhibition was observed against other pathogens in the other treatments. Bacteriocin from L. acidophilus (PV937061) showed the lowest stability, with strong initial inhibition at 30°C against K. pneumoniae (17.5 mm) and B. proteolyticus (23 mm), but significantly reduced activity pathogens at elevated temperatures. For bacteriocin from L. plantarum (PV937065), activity was observed following treatment at 70°C (18–19 mm against K. pneumoniae and B. proteolyticus) and consistent at 121°C ( Table 11).
Ultra-violet radiation
In general, ultraviolet (UV) light exposure had a time-dependent effect on the test bacteriocins, with the extent of inhibition against pathogens occurring with longer exposure times. After 20 to 40 min of exposure, the inhibitory activity reached a maximum, followed by a significant decrease. Among the test pathogens, K. pneumoniae and P. aeruginosa were the most consistently inhibited across LAB strains, while activity against B. subtilis and B. proteolyticus varied more widely. The inhibition of P. fuscoginae was particularly unstable, showing a pronounced reduction with extended UV exposure ( Table 12).
Table 12. Effect of ultra violet radiation on antibacterial activity of the test bacteriocins.
Bacteriocin fromL. Plantarum (PV937063) showed the largest spectrum of activity, maintaining high inhibition against K. pneumoniae (19.5 mm at 20 min) and B. proteolyticus (22.5 mm at 20 min) before progressive decline in activity with exposure to UV. Notably, the inhibition of P. aeruginosa was the highest at 60 min (20 mm). In addition, L. paracasei (PV937064) demonstrated stable activity, with strong inhibition against P. fuscoginae (17.5 mm at 20 min) and B. subtilis (15–19 mm across treatments), though responses diminished significantly at 80 min ( Table 12).
In contrast, L. plantarum (PV937062) showed a limited but steady profile, showing moderate activity (10–16 mm) across most pathogens with less drastic fluctuations over time. For L. acidophilus (PV937061), the loss of inhibition of P. fuscoginae and P. aeruginosa beyond was 40 min. Although high inhibition of B. subtilis (22.5 mm at 20–60 min) and B. proteolyticus (16–18 mm) was observed for L. plantarum (PV937065) at shorter UV exposure times, the activity was reduced after 40 min of exposure ( Table 12).
Ultra-violet radiation
In general, ultraviolet (UV) light exposure had a time-dependent effect on the test bacteriocins, with the extent of inhibition against pathogens occurring with longer exposure times. After 20 to 40 min of exposure, the inhibitory activity reached a maximum, followed by a significant decrease. Among the test pathogens, K. pneumoniae and P. aeruginosa were the most consistently inhibited across LAB strains, while activity against B. subtilis and B. proteolyticus varied more widely. The inhibition of P. fuscoginae was particularly unstable, showing a pronounced reduction with extended UV exposure ( Table 12).
Bacteriocin from L. Plantarum (PV937063) showed the largest spectrum of activity, maintaining high inhibition against K. pneumoniae (19.5 mm at 20 min) and B. proteolyticus (22.5 mm at 20 min) before progressive decline in activity with exposure to UV. Notably, the inhibition of P. aeruginosa was the highest at 60 min (20 mm). In addition, L. paracasei (PV937064) demonstrated stable activity, with strong inhibition against P. fuscoginae (17.5 mm at 20 min) and B. subtilis (15–19 mm across treatments), though responses diminished significantly at 80 min ( Table 12).
In contrast, L. plantarum (PV937062) showed a limited but steady profile, showing moderate activity (10–16 mm) across most pathogens with less drastic fluctuations over time. For L. acidophilus (PV937061), the loss of inhibition of P. fuscoginae and P. aeruginosabeyond was 40 min. Although high inhibition of B. subtilis (22.5 mm at 20–60 min) and B. proteolyticus (16–18 mm) was observed for L. plantarum (PV937065) at shorter UV exposure times, the activity was reduced after 40 min of exposure ( Table 12).
Storage condition
Generally, the antibacterial activity of bacteriocins is significantly affected by storage conditions, with ambient and refrigerated conditions typically maintaining stronger inhibition than freezing conditions. While refrigerated conditions maintained potency and occasionally increased activity with freezing, activity was either decreased or lost ( Table 13).
Table 13. Effect of storage condition on activity of the test bacteriocins.
For bacteriocin from L. plantarum (PV937063) highest spectrum, with strong inhibition against B. subtilis (27.0 mm refrigerated) and K. pneumoniae (26.5 mm refrigerated, 24.0 mm ambient) were recorded. In addition, significant activity against B. subtilis was maintained during ambient and refrigerated storage, while freezing resulted in a significant loss of activity. Under the respective storage conditions, L. paracasei (PV937064) consistently inhibited P. aeruginosa (16–22 mm) and P. fuscoginae (16–19 mm). However, a more selective activity was observed for bacteriocin from L. plantarum (PV937062), which inhibited P. fuscoginae (17–19 mm) and P. aeruginosa (13–21 mm), but showed little to no inhibition against K. pneumoniae. Under the respective storage conditions, L. acidophilus (PV937061) showed limited overall activity, losing all inhibition against K. pneumoniae and B. proteolyticus, while recording activity against P. aeruginosa (15–21 mm) and P. fuscoginae (19–20 mm). Although there was no activity against K. pneumoniae, bacteriocin from L. plantarum (PV937065) showed remarkable inhibition against P. aeruginosa (19–22 mm) and B. subtilis (21 mm ambient) ( Table 13).
Proteases
Following treatment with proteases, the inhibitory activity of the bacteriocins against the majority of pathogens was lost after pepsin treatment. The two exceptions were L. plantarum (PV937065), which demonstrated a narrow inhibitory impact against K. pneumoniae (10.0 mm), and L. plantarum (PV937063), which maintained moderate activity against B. proteolyticus(15.5 ± 6.36 mm) and B. subtilis (12.5 ± 3.54 mm). Following exposure to pepsin, L. paracasei (PV937064) also retained residual inhibition against B. subtilis (15.5 ± 6.36 mm) ( Table 14).
Table 14. Effect of proteases on antibacterial activity of the test bacteriocins.
In contrast, more selective retention of activity across the strains was observed after trypsin treatment. For bacteriocin from L. acidophilus (PV937061) inhibition against K. pneumoniae (12.5 ± 3.54 mm), P. aeruginosa (16.0 ± 5.66 mm), and B. subtilis (12.5 ± 3.54 mm) was observed. In the case of L. plantarum (PV937063), inhibition against K. pneumoniae and P. aeruginosa was completely eliminated, but inhibition against P. fuscoginae (17.5 ± 3.54 mm) and B. subtilis (12.5 ± 3.54 mm) following trypsin treatment was observed. L. plantarum (PV937062) and (PV937065) also showed resistance to enzymatic degradation, retaining action only against some pathogens. This trend revealed that most of the protease-resistant bacteriocins were produced by L. plantarum (PV937063) and L. acidophilus (PV937061). In contrast, L. paracasei (PV937064) maintained a pathogen-limited action, whereas L. plantarum (PV937062) and (PV937065) showed a limited inhibitory range.
Bacteriocin concentration
The inhibition of K. pneumoniae steadily increased from 14.5 mm at 10 mg/mL to 24.0 mm at 100 mg/mL, with a similar gradual trend against P. aeruginosa (10.5–20.0 mm) for L. plantarum (PV937063). L. paracasei (PV937064) showed delayed but essential activity, particularly against P. fuscoginae, P. aeruginosa, and B. subtilis, with inhibition increasing sharply above 50 mg/mL and peaking at 19–24 mm at 100 mg/mL. In addition, L. plantarum (PV937062) showed a narrower spectrum, maintaining only steady inhibition (10–19 mm) against K. pneumoniae, P. fuscoginae, and B. subtilis ( Table 4.15). In addition, the widest concentration-dependent responses were observed for L. acidophilus (PV937061), which showed activity against all tested pathogens. In the case of L. plantarum (PV937065), a smaller but potent effect was observed, especially against K. pneumoniae (21 mm) and P. aeruginosa (18 mm) at 100 mg/mL ( Table 15).
Table 15. Effect of bacteriocin concentration on antibacterial activity.
4. Discussion
Three species of Lactobacillus (L. plantarum, L. acidophilus and L. paracasei) were isolated from fermented foods (ogi, iru, and fufu) used in this study. Previous studies have reported the isolation of L. plantarum and L. acidophilusfrom fermented milk (Goa et al., 2022). Similarly, Jamuna and Jeevaratnam (2004) reported the isolation of L. acidophilus from fermented appam batter and pickles. All isolates used in this study showed bacteriocin-producing ability. Bacteriocin production by L. plantarum has been reported previously (Todorov, 2009; Wang et al., 2023).
In the present study, peak inhibitory effects occurred mainly in cultures aged between 48 and 120 h, while mild inhibition was observed in early cultures (24 h). Relatively old cultures (144–168 h) generally show a sharp decline or complete loss of inhibitory activity. This observation corroborates the report of Wang et al. (2021), who in a related study indicated peak inhibitory effects of LAB isolates occurring in young cultures, with activity decreasing with older cultures.
Generally, L. plantarum (PV937063) showed the broadest spectrum, with strong inhibition (16–21 mm) against K. pneumoniae and P. aeruginosa at 24–72 h, with markedly decreased activity at 96 h but rebounded at 120 h for several pathogens. In a related study by Darbandi et al. (2022), two different strains of L. plantarum showed inhibitory activity against gram-negative bacteria, including P. aeruginosa, E. coli, and A. baumannii. The findings of this study, however, contradict the observation of Bolivar-Jacobo et al. (2023), who reported that culture age, growth medium, ultrasound amplitude, and time of exposure influenced the kinetic growth of L. acidophilus (PV937061), and reported higher inhibitory activity with older cultures seven times greater than the inhibition of early culture of L. acidophilus.
In this study, medium pH (pH 3–6) was observed to show higher inhibitory effects on cell-free supernatant from L. plantarum, as activity frequently decreased at alkaline levels (≥8), and some strains showed a great decline in inhibition against specific pathogens or total loss of inhibition. This finding is similar to the report of Omotayo et al. (2019) who in a study on process parameters optimization and molecular studies on bacteriocin-producing lactic acid bacteria from ‘Kati’ reported that antibacterial efficacy of LAB isolates was stable at acidic pH ranged from 2 to 6, while at pH 8 to 12, there was observed decrease in the antibacterial activity and stability. This also aligns with the reports of Hasannejad et al. (2017) and Mohankumar and Murugalatha (2011), who reported higher inhibition between pH 4 and 7, but reduced activities in the alkaline range (8 to 10). In the report of Oh et al. (2000), Lactobacillus species were found to exhibit the highest antibacterial activity at pH 7, with a considerable decrease in inhibitory activity at both acidic and alkaline pH. Also, Yang et al. (2018), when investigating the influence of culture media, pH, and temperature on growth and bacteriocin production by bacteriocinogenic lactic acid bacteria, found that the optimal pH of crude bacteriocin was between 7.4 to 8.5, with reduced activity observed at acidic pH. An optimal pH range of 4-7 was however reported by Najim et al. (2012), who observed decreased activity under alkaline pH conditions.
With respect to temperature, broad thermostability was observed in some strains, whereas others displayed restricted or specific inhibition against the test pathogens. L. plantarum (PV937062) showed strong inhibition against P. fuscoginae, P. aeruginosa and B. proteolyticus at different temperature treatments, maintaining significant activity during sterilization (13.5–17.5 mm at 121°C). A similar observation was reported by Ehrmann et al. (2000), who observed the production of thermostable class two bacteriocin by L. plantarum TMW1.25, isolated from sausage fermentation. Also, Wang et al. (2018), in a study on purification and characterization of plantaricinLPL-1, bacteriocin produced by L. plantarumisolated from fermented fish, indicated stability of inhibitory activity after treatment at 60, 80, and 100°C. In addition, this present finding relates with a previous study by Ihum et al. (2024), of the impact of temperature variations on bacteriocin activity against spoilage bacteria in melon and its fermented product ’Ogiri, it was reported that over 80% of the bacteriocin activity were retained between 30 to 70°C. The L. plantarum (PV937062) strain isolated in the present study showed stability even at sterilization range of 121°C, showing strong inhibition against some test pathogens, such as P. fuscoginae, P. aeruginosa, and B. proteolyticus. This corresponds with the study of Melia et al. (2022), who reported inhibitory activity of cell-free supernatant of L. plantarum SN13T at sterilization temperatures.
In the case of the crude bacteriocin, the findings of this study showed inhibitory activity influenced by temperature, with the greatest inhibition occurring between 30 and 40°C and a steady decrease at higher temperatures. Some bacteriocins from some of the LAB strains [L. plantarum (PV937063), L. paracasei (PV937064), and L. acidophilus (PV937061)], showed thermostable features, retaining remarkable inhibition up to 121°C. When compared to the study on characterization and profiling of bacteriocin-like substances produced by lactic acid bacteria from cheese samples by Afrin et al. (2021), it was observed that the activities of extracted bacteriocins from all the LAB isolates were stable within temperature range between 40–70 °C and following heat treatment at 121 °C. Also, Vijayendra et al. (2010) reported that bacteriocins isolated from LAB strains did not lose their antibacterial activity when treated at high temperatures between 100°C and 121°C.
Based on the findings of this study, treatment with pepsin completely eliminated the antibacterial activity across all strains and test pathogens. In the case of trypsin, although the activity was also typically lost in most CFS, L. plantarum (PV937063) and L. paracasei (PV937064) retained little inhibition (10.5 mm) against P. aeruginosa. A related study by Showpanish et al. (2022) reported that the inhibitory activity of L. plantarum RB01-SO was completely inactivated by proteolytic enzymes, such as pepsin, trypsin, and chymotrypsin, which showed the proteinaceous nature of bacteriocins after treatment with proteolytic enzymes. Another study reported the isolation, purification, and partial characterization of plantaricin 423, a bacteriocin produced by L. plantarum that LAB strains activity was inactivated when treated with proteases such as pepsin, papain, alpha-chymotrypsin, trypsin, and Proteinase K (van Reenen et al., 1998).
The L. plantarum (PV937065) used in the study demonstrated a narrow inhibitory effect against K. pneumoniae (10.0 mm), while L. plantarum (PV937063) maintained moderate activity against B. proteolyticus (15.5 ± 6.36 mm) and B. subtilis (12.5 ± 3.54 mm) following treatment with trypsin. Following exposure to pepsin, L. paracasei (PV937064) also retained residual inhibition against B. subtilis (15.5 ± 6.36 mm). In a previous study on the discovery and characterization of the circular bacteriocin plantacyclin B21AG from Lactiplantibacillus plantarum B21, bacteriocin was reported to be more resistant to trypsin than to pepsin (Golneshin et al., 2020). In addition, Bendjeddou et al. (2012), in a study on the characterization and purification of a bacteriocin from L. paracasei subsp. paracasei BMK2005, reported that bacteriocin showed stable antibacterial activity after treatment with trypsin, whereas its activity markedly diminished when treated with pepsin and α-chymotrypsin. It is hypothesized that the resistance of bacteriocins to pepsin and trypsin is linked to circularization (Perez et al., 2018).
The present study revealed that bacteriocin efficacy was positively associated with concentration, with all strains showing increased activity at higher concentrations and L. plantarum (PV937063) and L. acidophilus (PV937061), which showed broad-spectrum and high-intensity inhibition across the test pathogens. Similarly, a previous study reported that bacteriocins produced by L. acidophilus strains exhibited antimicrobial activity against various pathogens, including food-borne microorganisms (Bharal et al., 2013). In another study on the antimicrobial potential and stability of L. acidophilus-derived bacteriocins against multidrug-resistant common foodborne pathogens, it was reported that bacteriocin showed potent antimicrobial activity at low concentrations; however, against some pathogens, higher concentrations were needed for effective inhibition (Hasan et al., 2025). Papagianni et al. (2006) in a related study reported that only 22% of the test pathogens investigated were sensitive to bacteriocin produced by L. curvatus ATCC 51436 and Pediococcus acidilactici ATCC 2574 at very low concentrations of the bacteriocins.
Based on the results of this study, the effect of UV irradiation on the antimicrobial activity of CFS showed distinct strain- and pathogen-specific responses. The highest inhibition was observed in the presence of L. plantarum (PV937063), which showed strong activity against K. pneumoniae (20–30 mm), P. aeruginosa (20–30 mm), and B. subtilis (20–22 mm) after 60–80 min of exposure to UV light, while the bacteriocins in CFS from L. paracasei (PV937064) and L. acidophilus (PV937061) were observed to be more susceptible to UV degradation. In a previous study, Mahreen et al. (2020), in a study on characterisation of bacteriocins produced by Lactobacillus spp. isolated from the traditional Pakistani yoghurt and their antimicrobial activity against common foodborne pathogens, it was reported that bacteriocin from LAB strains retained activity following 30 min of UV exposure. In addition, Shokri et al. (2014) also reported that after 30 min of exposure to UV light, bacteriocin-like inhibitory substances from LAB strains retained their activity. Afrin et al. (2021) have reported similar observation under similar exposure to UV.
With respect to the effect of storage conditions, freezing and refrigerated storage often resulted in partial or total loss of action, while ambient storage maintained the highest inhibition across the CFS of L. plantarum (PV937063) showed a more stable and wide-ranging spectrum and maintained activity in both ambient and frozen storage. L. acidophilus (PV937061) also showed freeze-stable efficacy, L. plantarum (PV937065) showed a more limited but somewhat stable inhibitory range. In contrast, in this study, L. paracasei (PV937064) and L. plantarum (PV937062) were more condition-dependent and retained considerable activity during ambient storage. It was reported that under refrigerated (4°C) and normal conditions, bacteriocins are generally very stable (Halimi et al., 2010). In a related study, Guo et al. (2024) reported that bacteriocin was variably affected by different low-temperature environments and at −20°C and −80°C. Their report indicated that inside the protein, the molecular motion is slowed, and there is a great reduction in the surroundings or halts enzymatic reactions and other structural-altering processes.
The antibacterial activity of the crude bacteriocins was significantly affected by storage conditions, with ambient and refrigerated conditions typically maintaining stronger inhibition than freezing, while refrigerated condition-maintained potency and occasionally increased activity, and with freezing, activity was either decreased or lost. Halimi et al. (2010), in a similar study, indicated that under refrigerated (4°C) and normal conditions, bacteriocins are generally very stable. When treated with different organic solvents, none of the CFS showed inhibitory activity against the test pathogens treated with ethyl acetate. The highest inhibitory activity was observed for ethanol-treated CFS, followed by methanol and, to a lesser extent, acetone. The inhibitory activity of L. paracasei was observed to be retained at 97% when treated with methanol, ethanol, acetone and benzene (Ge et al., 2016).
This study revealed L. plantarum (PV937063), ethyl acetate, methanol, and acetone treatments did not affect antibacterial activity against K. pneumonia, P.fuscoginae and P. aeruginosa and a medium but consistent activity was observed for bacteriocin from L. paracasei (PV937064) at the different treatments. In a similar study by Abanoz and Kunduhoglu (2018), bacteriocin KT11 was found to be stable after treatment with surfactants and organic solvents such as chloroform, propanol, methanol, ethyl alcohol, acetone, hexane, and ethyl ether. Bacteriocin from L. plantarum (PV937062) showed selective inhibition with acetone and ethanol. This finding is also in agreement with the study by Otunba et al. (2022), who reported that when treated with organic solvents such as ethanol, phenol, acetone, chloroform, and isoamyl alcohol, the activity of bacteriocin was stable. L.acidophilus (PV937061) showed the lowest inhibitory effect, with inhibition generally below 16 mm, except under ethanol treatment.
5. Conclusion
A total of 33 lactic acid bacteria (LAB) species isolated from Iru, Ogi, and fufu were preliminarily screened for bacteriocin production, of which five isolates [Lactobacillus plantarum (PV937063, PV937062, PV937065)], Lactobacillus paracasei (PV937064), and Lactobacillus acidophilus (PV937061)] were used for the study. Among the LAB strains, Lactobacillus plantarum (PV937063) and Lactobacillus paracasei (PV937064) had the widest antibacterial range and largest zones of inhibition, suggesting their possible application as natural antimicrobial agents.
Optimization studies of cell-free supernatants and bacteriocins from LAB showed that culture age strongly influenced bacteriocin activity, with maximum inhibition observed at mid-incubation stages. In addition, optimal antimicrobial activity was observed in mildly acidic environments, similar to natural fruit juice pH.
Stability studies in the presence of the organic acids methanol and acetone were moderately effective, while stability was lost for most isolates in the ethyl acetate-treated samples. Among the solvents, ethanol was the best for preserving bacteriocin activity, indicating that its polarity enhanced compound extraction.
Fort temperature stability, bacteriocin activity were stable between 30–80°C, with peak inhibition around 70–80°C, while activity reduced at sterilization temperature (121°C). In addition, when exposed to varying UV exposure times, a duration greater than 40 min significantly reduced the antibacterial activity. Furthermore, all bacteriocins were most stable under refrigeration, while ambient storage caused a gradual loss of activity. However, freezing preserved bacteriocin activity for longer periods. Regarding the effect of proteases on stability, enzymatic digestion reduces or eliminates antimicrobial activity, thus confirming the proteinaceous nature of the active compounds.
In addition, both LAB inocula and bacteriocins were stable under moderate heat and acidic conditions. In addition, the presence of bacteriocins as preservatives did not remarkably affect the sensory qualities and physicochemical properties of the juice; thus, they could serve as reliable natural preservatives suitable for the juice industry.
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Ogundipe A, Ogunnusi TA and Akpor OB. Isolation and Optimization of Bacteriocin-Producing Lactic Acid Bacteria from Nigerian Fermented Foods [version 1; peer review: awaiting peer review]. F1000Research 2025, 14:1467 (https://doi.org/10.12688/f1000research.174479.1)
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