Kefir yeasts enhance probiotic potentials of Lactobacillus paracasei H9: The positive effects of coaggregation between the two strains

This study investigated pure Lactobacillus paracasei H9 tolerance to simulated gastrointestinal juices and adhesion to intestinal mucosa cells without yeasts, with viable yeasts (VY) and with different pretreated yeasts. Three models including gastric secretion tolerance (GST), intestinal juice tolerance (IJT) and sequential gastrointestinal tolerance (SGT) were respectively employed to assay the tolerance of L. paracasei H9, whilst Caco-2 cell line was used to investigate the bacterial adhesion. Particularly, the co-aggregation ability of the two strains at pH values of 2.0, 8.0 and 7.2 was originally carried out to study relations to the bacterial probiotic potentials. Results showed that yeast counts in the range from 3.0 to 5.0 log CFU mL^? 1 could gradually increase the viability of L. paracasei H9 in SGT. The bacterial viability in the three tolerance models and the adherent number to Caco-2 cells were significantly improved with addition of VY (P < 0.05). The L. paracasei H9 with VY in gastric juice at pH 2.0 and intestinal juice at pH 8.0, respectively, exhibited higher aggregation percentage compared with that of single L. paracasei H9 at 37 °C (P < 0.05). The aggregation ability of L. paracasei H9 with VY at pH 7.2, which might contribute to increase the adhesion of the bacteria, also excelled that of L. paracasei H9 (P < 0.05). It is deduced that proteins of the bacterial cell surface and polysaccharides in yeast cell walls play important roles in co-aggregation of the two strains and the microbial adhesion specificity to Caco-2 cells. The co-aggregation of the two strains also contributes to enhancing probiotic potentials of L. paracasei H9.     

Prenylation preserves antioxidant properties and effect on cell viability of the natural dietary phenol curcumin

Curcumin (CRM) is a naturally occurring phenolic compound with a variety of biological and pharmacological activities. We have investigated the effect of aromatic C-prenylation on the antioxidant activity of this natural compound. The protective effect of CRM and its diprenyl derivative (PCRM) was investigated against neat cholesterol degradation (at 140 °C) and Cu^2 +-induced oxidation (at 37 °C) of liposomes and human low density lipoproteins. The activity of two simplified vanilloid analogs (vanillin and vanillyl alcohol) was also compared in the same systems. Cytotoxicity and cell permeation of both curcuminoids were also assessed using differentiated Caco-2 cell monolayers. PCRM, like CRM, significantly inhibited the oxidative degradation of polyunsaturated fatty acids and cholesterol, and the formation of their oxidation products in the oxidative stress systems, acting as scavenger of peroxyl radicals, without toxic effect (in the range 10–100 μM) on differentiated Caco-2 cell viability. Nevertheless, the structural modification of the lead compound severely affected membrane permeation through the Caco-2 monolayers, with apparent permeability coefficient (P_app) values in the apical-to-basolateral direction (2-h incubation) of 2.93 ± 0.94 × 10^− 6 cm/s and < 10^− 7 cm/s for CRM and PCRM, respectively. Taken together, our observations reveal a surprising bioactivity of PCRM, and qualify this compound as an interesting probe to explore the antioxidant pharmacophore of curcuminoids.     

Milk enhances intestinal absorption of green tea catechins in in vitro digestion/Caco-2 cells model

The effect of milk on the absorption of polyphenols is still controversial so far. In order to determine the impact of milk addition on green tea catechins bioaccessibility and intestinal absorption an in vitro digestion/Caco-2 cell model was applied. Green tea extract (GTE) was solubilized in distilled water at 23 °C and 100 °C, combined with skimmed milk (GTE + 10% milk and GTE + 25% milk) and subjected to simulated gastric and intestinal digestion, followed by transepithelial absorption in Caco-2 cells monolayers. In the mixture with milk, gallated catechins: ECG and EGCG showed binding to milk proteins while EC and EGC seemed to have weaker affinity. Catechins were stable during gastric incubation and very sensitive to intestinal digestion. Bioaccessibility of green tea catechins brewed at 100 °C was higher than brewed at 23 °C. Catechins from digested GTE with 10% and 25% milk exhibited enhanced intestinal permeability in Caco-2 model in comparison to non-digested GTE and digested GTE without milk. Apparent permeability coefficients (P_app) of EGCG and ECG in digested GTE with 25% milk were significantly higher compared to those in GTE with 10% milk, and amounted to 2.41 × 10^− 6 cm/s and 1.39 × 10^− 6 cm/s. The recoveries of all catechins in GTE with milk in Caco-2 cells after 2 h incubation were significantly higher than that without milk. To summarize, these data suggest that milk addition may increase catechin bioavailability by enhancing their transepithelial absorption and uptake from green tea extract.     

Rapid enumeration of Bifidobacterium lactis in milk powders using impedance

An impedance method was evaluated to enumerate Bifidobacterium lactis, a probiotic, added to milk powder. Impedance changes were measured at 40 °C and recorded using the BacTrac™ 4100 microorganism growth analyser. Five different media were compared for the optimum impedance response. A raffinose-based medium (B. lactis medium) produced the fastest and most reproducible results. Good correlations were obtained between cell numbers from pure cultures of B. lactis (DR 10™) on reinforced clostridial agar plates and the impedance changes in the B. lactis medium. Enumeration of these bacteria in milk powder using the BacTrac™ 4100 impedance system showed no significant difference when compared with agar plate count results. The impedance counts estimated the cell counts of 10⁶ cells g⁻¹ within 15 h and was faster than the 3 days required to obtain a result using the agar plate count method.     

Transport of bovine milk-derived opioid peptides across a Caco-2 monolayer

The transepithelial transport of μ-opioid (MOP) receptor agonists, i.e., bovine β-casomorphin-5 and -7 (derived from β-casein) and an antagonist, i.e., casoxin-6 (derived from κ-casein) were investigated using a human intestinal cell (Caco-2) monolayer. To determine the transport pathway of the peptides across Caco-2 monolayers, two directions of the transport (apical-to-basolateral and basolateral-to-apical) were studied. All investigated peptides were transported across Caco-2. The cumulative fractions of transported peptides were increased linearly in time in each case. The permeability coefficients (P_aap) calculated for all peptides in two transport directions ranged between 0.57 and 9.21 × 10^?6 cm min^?1. These data suggest the possibility of transport of opioid peptides derived from food across human intestinal mucosa.     

Temperature effect on the biological activity of Bifidobacterium longum CRL 849 and Lactobacillus fermentum CRL 251 in pure and mixed cultures grown in soymilk

The influence of temperature on the growth and biological activity of two probiotic strains (Bifidobacterium longum CRL 849 and Lactobacillus fermentum CRL 251) as pure and mixed cultures in soymilk (SM) were evaluated. Maximum growth was observed at 37°C in both mixed and pure cultures. In a product prepared with the mixed culture (1:1) at 37°C, the amount of lactic acid produced was approximately 55 mmol l⁻¹ after 24 h with a slow production rate (2.8 mmol l⁻¹ h⁻¹); the formation of acetic acid was higher with respect to pure cultures (82.01 mmol l⁻¹ after 24 h), and final pH (24 h) was 5.0. About 85% of the total amount of sugars in SM was reduced, mainly sucrose. Stachyose was reduced (71%) after 4 h of incubation. Maximum activity of alpha-galactosidase (alpha-gal) (13.2 U ml⁻¹) was observed after 6 h. At 37°C the bifidobacterium strain was viable in mixed culture throughout the period assayed. At lower (30°C) or higher (42°C) temperatures, mixed culture showed slower growth and lower acid production in SM but the alpha-gal activity was stimulated at 30°C.     

Influence of pH and NaCl on monolaurin inactivation of Streptococcus iniae

Streptococcus iniae, a Gram-positive bacterium, has been recognized as a cause of human and fish streptococcosis. Aquacultured fish are susceptible to infection and have been epidemiologically linked with wound infections of fish handlers. Therefore, there is a need to develop methods of prevention and/or control of this pathogen. This study determined the minimum inhibitory concentration of monolaurin (glycerol monolaurate) against S. iniae strains PW and BU and utilized gradient plates at 37°C to determine the combined influence of pH and NaCl on monolaurin inactivation of S. iniae. The minimum inhibitory concentration of monolaurin against strain PW was 12.4±0.3 μg ml⁻¹ and against strain BU was 12.1±0 μg ml⁻¹. Compared to growth in the absence of monolaurin, the incorporation of 6 μg ml⁻¹ monolaurin into the salt (1.12–6.95%)–pH (4.26–8.88) gradient plates prevented growth of strain PW at pH values <6.00 and strain BU at pH values <6.20, regardless of the salt concentration. The combination of monolaurin with environmental variables such as pH and salt proved to be an effective tool for in vitro control of S. iniae.     

Determination of trace tin in foods by single-sweep polarography

A new method for the determination of trace tin in a solution of oxalic acid–methylene blue by single-sweep polarography is developed. The mechanism for the polarographic waves and the conditions for determining tin are discussed in this report. The peak height is directly proportional to the concentration of tin (IV) over the range 2.0×10⁻⁸–1.0×10⁻⁶ g ml⁻¹. The detection limit and the recovery of the method are 1.0×10⁻⁸ g ml⁻¹ and 93–98%, respectively. The method was successfully employed for the determination of tin in juices of canned fruits.     

Features and performance of edible films,obtained from whey protein isolate formulated with antimicrobial compounds

The goal of this research effort was to assess the efficacy of edible films produced from whey protein isolate (WPI) and glycerol, including incorporation of lactic acid (LA) and propionic acid (PRO), chitooligosaccharides with nominal MW of 3 kDa (COS) and natamycin (NA) as antimicrobial agents. Their features were evaluated in vitro via agar diffusion and viable cell counting, against spoilage microflora often found contaminating cheese surfaces. The effect of incorporating the aforementioned compounds upon thickness, moisture content (MC), solubility (S), density (ρ^s), water activity (a_w) and water vapor permeability (WVP), as well as upon tensile and optical properties of those films were also evaluated. Films formulated with LA, PRO or COS exhibited antimicrobial activity against all microorganisms tested, yet the viable cell count assay was more sensitive and reproducible. COS was the most active against Gram-negative bacteria, whereas LA was the most active against Gram-positive ones. NA was not active against bacteria, but displayed the strongest effect against yeasts. Incorporation of said antimicrobial compounds did not significantly (p > 0.05) affect film thickness, yet it significantly (p < 0.05) reduced tensile strength (TS). Incorporation of LA and NA in particular did not significantly (p < 0.05) affect MC, S, ρ^s, WVP, elongation at break (EB) and Young's modulus (YM) values; however, a statistically significant increase (p < 0.05) of MC, S and WVP, together with a statistically significant decrease (p < 0.05) of ρ^s were attained upon incorporation of PRO or COS. Moreover, PRO produced the highest variation (p < 0.05) in EB, TS and YM, whereas COS produced the highest change (p < 0.05) in optical properties.     

Growth-inhibition of hiochi bacteria in namazake (raw sake) by bacteriocins from lactic acid bacteria

The bacteriocins produced by Lactococcus lactis subsp. lactis C101910 (C101910) and NBRC 12007 (NBRC 12007) were used to prevent the growth of sake spoiling hiochi bacteria (Lactobacillus hilgardii, Lactobacillus fructivorans, and Lactobacillus paracasei) in namazake, which is raw (unpasteurized) sake. The bacteriocin concentrations required for decreasing the viable cell concentrations of L. hilgardii and L. fructivorans below the detection limit (1.0 × 10² cells/ml) in 24 h from the initial concentration of 4.0–9.5 × 10⁵ cells/ml in the namazake at pH 4.5 and at 4°C, were 18–35 U/ml and 5.6 U/ml for the bacteriocin from C101910 and NBRC 12007, respectively. To decrease the viable cell concentration of L. paracasei from the initial concentration of 7.5 × 10⁵ cells/ml to below the detection limit (1.0 × 10² cells/ml) in 24 h, 350 U/ml bacteriocin from C101910 and 140 U/ml bacteriocin from NBRC 12007 were required. In experiments using McIlvaine buffer (pH 4.5) with 15% ethanol instead of namazake as the medium, the viable cell concentrations of L. hilgardii and L. paracasei decreased to less than 1.0 × 10² cells/ml, whereas those of L. fructivorans decreased to less than 1.0 × 10³ cells/ml, when bacteriocins were added at the concentrations that had proven effective in namazake. The membrane depolarization assay using a fluorescent probe showed that the presence of ethanol stimulated the collapse of the membrane potential induced by bacteriocins. The ethanol induced collapse of the membrane potential suggests that the application of bacteriocins at the storage stage of namazake is more beneficial than when used in other stages of the sake brewing process.     

An Improved Assay for Measuring Heparin Binding to Bull Sperm

The binding of heparin to sperm has been used to study capacitation and to rank relative fertility of bulls. Previous binding assays were laborious, used 10⁷ sperm per assay point, and required large amounts of radiolabeled heparin. A modified heparin-binding assay is described that used only 5 × 10⁴ cells per incubation well and required reduced amounts of [³H] heparin. The assay was performed in 96-well Millititer plates, enabling easy incubation and filtering. Dissociation constants and concentrations of binding sites did not differ if analyzed by Scatchard plots, Woolf plots, or by log-logit transformed weighted nonlinear least squares regression, except in the case of outliers. In such cases, Scatchard analysis was more sensitive to outliers. Nonspecific binding was insignificant using nonlinear logistic fit regression and a proportion graph. The effects were tested of multiple freeze-thawing of sperm in either a commercial egg yolk extender, 40 mM Tris buffer with 8% glycerol, or 40 mM Tris buffer without glycerol. Freeze-thawing in extender did not affect the dissociation constant or the concentration of binding sites. However, freeze-thawing three times in 40 mM Tris reduced the concentration of binding sites and lowered the dissociation constant (raised the affinity). The inclusion of glycerol in the 40 mM Tris did not significantly affect the estimated dissociation constant or the concentration of binding sites as compared to 40 mM Tris without glycerol.     

Antibiotic resistance and susceptibility to some food preservative measures of spoilage and pathogenic micro-organisms from spices

Antibiogram of 84 strains of Bacillus cereus, 26 strains of Clostridium perfringens, four strains of Staphylococcus aureus, 51 strains of Enterobacteriaceae, two strains of each of Salmonella and Shigella; isolated from spices, were studied against 20 different antibiotics that are commonly used against foodborne diseases, mainly gastroenteritis. All the tested strains of B. cereus, Cl. perfringens, Staph. aureus, Escherichia coli, Salmonella and Shigella were found resistant to at least 3, 4, 7, 6, 10 and 9 antibiotics, respectively. In brain–heart infusion broth supplemented with glucose, the D_100°C-values for B. cereus were 3.5–5.9 min, and the z-values were 17–18°C. The D_100°C-values for Cl. perfringens in fluid thioglycolate medium were higher (10.0–19.8 min) than those of B. cereus. The minimum inhibitory concentrations (MICs) of sodium chloride were 45–80 mg ml⁻¹. While the MIC of benzoic acid for Cl. perfringens, tested on perfringens agar (pH 7.3) plates by incubating anaerobially at 35°C for 24 h, was 1.9–2.2 mg ml⁻¹, for others, tested on nutrient agar (pH 6.8) plates by incubating at 35°C for 18 h in static aerobic condition, it was much less. Similarly, the MIC of sorbic acid for all the tested isolates, excepting Cl. perfringens, was 0.6–1.1 mg ml⁻¹. Of the eight isolates of Cl. perfringens, only three were inhibited at 2.0 mg sorbic acid ml⁻¹, while others were resistant. Sixty percent and 75% of the respective strains of B. cereus and Cl. perfringens were resistant to 5000 IU Nisaplin ml⁻¹, whereas the MIC values of Staph. aureus were between 3000 and 5000 IU ml⁻¹. While studying combined effect of selected hurdles on the growth of enterotoxigenic Cl. perfringens 16-C2, the judicious combination considered was low acid (pH 6.0), 30 mg sodium chloride ml⁻¹ and 1.25 mg benzoic acid ml⁻¹.     

Development of an immunosensor based on surface plasmon resonance for enumeration of Escherichia coli in water samples

An immunosensor for the rapid, sensitive and selective detection of generic Escherichia coli in water samples was developed on the basis of surface plasmon resonance (SPR). Antibodies against E. coli were immobilized on a sensor surface and water samples with different concentrations of E. coli were injected to the flow cell. The changes in the response unit (RU) due to the binding of different concentrations of bacteria were computed and a linear correlation (R² = 0.976) was obtained between log E. coli concentration and the change in the RU with a working range of 9 × 10¹ and 1.8 × 10⁵ cfu ml⁻¹. The complete regeneration of the sensor surface was realized by using 1% SDS solution. The selectivity of the developed sensor was examined with Enterobacter aerogenes and Enterobacter dissolvens which did not produce any significant response. The ability of the developed sensor to detect E. coli in real water samples was investigated and the results were closed to that obtained from plate counting method. Based on these results, the developed immunosensor can be employed for rapid, label-free, sensitive and selective detection of E. coli in water samples.     

Probiotic-loaded microcapsule system for human in situ folate production: Encapsulation and system validation

This study focused on the use of a new system, an alginate | Ɛ-poly-l-lysine | alginate | chitosan microcapsule (APACM), able to immobilize a folate-producing probiotic, Lactococcus lactis ssp. cremoris (LLC), which provides a new approach to the utilization of capsules and probiotics for in situ production of vitamins. LLC is able to produce 95.25 ± 26 μg·L^− 1 of folate, during 10 h, and was encapsulated in the APACM. APACM proved its capacity to protect LLC against the harsh conditions of a simulated digestion maintaining a viable concentration of 6 log CFU·mL^− 1of LLC. A nutrients exchange capacity test, was performed using Lactobacillus plantarum UM7, a high lactic acid producer was used here to avoid false negative results. The production and release of 2 g·L^− 1 of lactic acid was achieved through encapsulation of L. plantarum, after 20 h. The adhesion of APACM to epithelial cells was also quantified, yielding 38% and 33% of capsules adhered to HT-29 cells and Caco-2 cells, respectively.     

Immunoglobulins,growth factors and growth hormone in bovine colostrum and the effects of processing

In colostrum collected 0–80 h postpartum the contents of immunoglobulins (Igs), transforming growth factor beta-2 (TGF-β2), insulin-like growth factor-1 (IGF-1) and growth hormone (GH) were analysed. Colostrum initially contained 90 mg mL⁻¹ IgG1, 2.8 mg mL⁻¹ IgG2, 1.6 mg mL⁻¹ IgA, 4.5 mg mL⁻¹ IgM, and these concentrations declined by 92%, 87%, 93% and 84%, respectively, in the samples collected later. Of the growth factors, colostrum initially contained 289–310 ng mL⁻¹ TGF-β2 and the concentration diminished to 66 ng mL⁻¹. The content of IGF-1 and GH postpartum decreased from 870 to 150 ng mL⁻¹, and from 0.17 to <0.03 ng mL⁻¹, respectively. Heat treatment and freeze-drying of colostral whey decreased the content of Igs to 75%, while the contents of IGF-1 and TGF-β2 were unaffected. A similar processing, including filtration steps reduced also the IGF-1 and TGF-β2 by 25%. IgM seems to be the most sensitive of the Igs to processing.     

Survival and activity of selected probiotic organisms in set-type yoghurt during cold storage

The growth and metabolism of two probiotic organisms (L. acidophilus LAFTI^® L10 and Lactobacillus casei LAFTI^® L26) and a regular yoghurt culture (L. delbrueckii ssp. bulgaricus Lb1466 and Streptococcus thermophilus St1342) were studied in yoghurt containing 0.5%, 1.0%, and 1.5% (w/v) of high amylose corn starch powder (Hi-maize^®) or inulin. Viable cell counts of probiotic organisms, their metabolites and proteolytic activities, and viscosity of the yoghurts were determined during refrigerated storage for 28 d at 4 ^oC. In the presence of inulin, cultures showed better retention of viability (8.0 log cfu g⁻¹) in comparison with that of Hi-maize, which had a reduction by one log cycle. Lower concentrations of 0.5–1.0% Hi-maize improved (P<0.05) the production of propionic acid and also increased proteolytic activity of probiotic organisms substantially. A greater release of free amino acids may have sustained better growth of the organisms in yoghurts. Supplementation with either Hi-maize or inulin increased the viscosity of probiotic yoghurts significantly (P<0.05).     

Routine and sensitive SPE-HPLC method for quantitative determination of pheophytin a and pyropheophytin a in olive oils

A sensitive and specific HPLC method with tandem diode array-fluorescence detection (DAD-FL) has been developed and validated for the simultaneous determination of pheophytin a (phy a) and pyropheophytin a (pyrophy a) in olive oils. Pigments were extracted with reverse phase solid phase extraction (RP-SPE) and subsequently analysed by HPLC-DAD-FL. The chromatographic analysis was carried out isocratically on ODS2 RP column using methanol–acetone (1:1 v/v) at flow-rate of 2.0 ml min⁻¹. Specificity of the method was assured by the simultaneous detection by UV–visible (410 nm) and FL (λ_Ex: 410 nm; λ_Em: 672 nm). Both compounds could be baseline separated within 7 min. The method was validated and applied in olive oil samples recently extracted as well as stored during 12 months. The limit of detection (LOD) defined at a signal-to-noise ratio of about 3 was ∼21.6 ng g⁻¹ for pyrophy a and ∼24.6 ng g⁻¹ for phy a under FL detection, and ∼148.0 ng g⁻¹ for both analytes under UV–visible detection. The calibration graphs were linear (r² > 0.9999; p < 0.01) between 0.25–14.00 ng μl⁻¹ for pyrophy a and 0.25–19.00 ng μl⁻¹ for phy a, under both fluorescence and UV–visible detection conditions. Recoveries of phy a and pyrophy a were over 94% as estimated by the standard addition method. Relative standard deviation for the intra-day and inter-day determination of phy a and pyrophy a were lower than 3.7% and 8.0%, respectively.     

Production of volatile aroma compounds by kefir starter cultures

Production of carbonyl compounds by single-strain cultures, kefir starter (Lactobacillus delbrueckii subsp. bulgaricus HP1+Lb. helveticus MP12+Lactococcus lactis subsp. lactis C15+Streptococcus thermophilus T15+Saccharomyces cerevisiae A13) and kefir grains during fermentation and storage of kefir was studied. The content of carbonyl compounds produced by kefir starter was greater than that produced by kefir grains. The maximum acetaldehyde concentration (18.3 μg g⁻¹) in kefir with starter culture was mainly due to the metabolic activity of Lb. delbrueckii subsp. bulgaricus HP1 isolated from kefir grains. The highest diacetyl production activity was recorded in the starter culture (1.87 μg g⁻¹) and the single-strain culture St. thermophilus T15 (1.62 μg g⁻¹), followed by Lb. helveticus MP12 (0.85 μg g⁻¹) and Lc. lactis subsp. lactis C15 (0.42 μg g⁻¹). The lactobacilli Lb. delbrueckii subsp. bulgaricus HP1 and Lb. helveticus MP12 produced acetone, which was not found in the cocci cultures. The presence of 2-butanone was related to the production ability of Lb. helveticus MP12. In comparison, Lc. lactis subsp. lactis C15 synthesized ethyl acetate more actively than the other single-strain cultures included in the starter. S. cerevisiae A13 produced ethanol and CO₂ in amounts (3975 μg g⁻¹; 1.80 g L⁻¹) that lent cultured kefir distinctive flavour and aroma characteristic of authentic kefir.     

Antimicrobial activity of pediocin-producing Lactococcus lactis on Listeria monocytogenes,Staphylococcus aureus and Escherichia coli O157:H7 in cheese

The antimicrobial activity of two pediocin-producing transformants obtained from wild strains of Lactococcus lactis on the survival of Listeria monocytogenes, Staphylococcus aureus and Escherichia coli O157:H7 during cheese ripening was investigated. Cheeses were manufactured from milk inoculated with the three pathogens, each at approximately 6 log cfu mL⁻¹. Pediococcus acidilactici 347 (Ped⁺), Lc. lactis ESI 153, Lc. lactis ESI 515 (Nis⁺) and their respective pediocin-producing transformants Lc. lactis CL1 (Ped⁺) and Lc. lactis CL2 (Nis⁺, Ped⁺) were added at 1% as adjuncts to the starter culture. After 30 d, L. monocytogenes, S. aureus and E. coli O157:H7 counts were 5.30, 5.16 and 4.14 log cfu g⁻¹ in control cheese made without adjunct culture. On day 30, pediocin-producing derivatives Lc. lactis CL1 and Lc. lactis CL2 lowered L. monocytogenes counts by 2.97 and 1.64 log units, S. aureus by 0.98 and 0.40 log units, and E. coli O157:H7 by 0.84 and 1.69 log units with respect to control cheese. All cheeses made with nisin-producing LAB exhibited bacteriocin activity throughout ripening. Pediocin activity was only detected throughout the whole ripening period in cheese with Lc. lactis CL1. Because of the antimicrobial activity of pediocin PA-1, its production in situ by strains of LAB growing efficiently in milk would extend the application of this bacteriocin in cheese manufacture.