Hydrolysis of β-casein (193-209) Fragment by Whole Cells and Fractions of Lactobacillus casei and Lactococcus lactis

Whole cells and fractions of Lactococcus lactis subsp. lactis IFPL 359 and Lactobacillus casei subsp. casei IFPL731 were studied. Hydrolysis products were separated by reversed-phase, high-performance liquid chromatography (RPHPLC). Under conditions, pH 5.2 and 3% NaCl, L. casei IFPL 731 was more active in hydrolysis of the b-casein (f193-209) peptide than was L. lactis IFPL 359. This hydrolyzing activity was attributed for L. casei IFPL 731 by the cell-wall proteinase. Hydrolysis of the peptide by the intracellular extract of L. casei IFPL731 was mainly located in the fraction that contained endopeptidase and Pep N aminopeptidase activities. Results may help provide approaches and treatments to control bitterness in cheese products.     

Peptidase and proteinase activity ofLactococcus lactis,Lactobacillus casei andLactobacillus plantarum

The proteolytic system of several non-commercial strains of lactococci and lactobacilli that were isolated directly from traditional-Spanish, semi-hard, goats' milk cheese was studied. The aminopeptidase, X-prolyldipeptidyl aminopeptidase, dipeptidase and proteinase activity of these new strains was measured for the cytoplasmic, cell-wall/membrane and spontaneously released fractions. The aminopeptidase activity was exclusively intracellular and higher forLactobacillus casei subsp.casei than forLactococcus lactis subsp.lactis. Lactobacillus plantarum showed higher dipeptidase activity thanL. casei. The highest level of proteinase activity was recorded for the cell-wallmembrane fraction ofLactococcus lactis subsp.lactis IFPL 359, and was higher on β-casein than on α_s-casein for all the strains studied. These results suggest some different contribution of these strains to the proteolysis of cheese during ripening and they seem to complement each other when used together in the starter culture.     

The accelerating effect of aLactococcus lactis subsp.lactis lactose-utilization/proteinase-deficient strain on proteolysis and flavour development

This study describes the accelerating effect exerted on proteolysis and flavour development in cheese-curd slurries by combined application ofLactococcus lactis subsp.lactis IFPL359 and high concentrations of its Lac^? Prt^? derivative, (i.e. that which has reduced capacity to metabolize lactose and reduced proteolytic activity, strain T1). Cells of strain T1 partially lysed by either sonication or incubation with lysozyme, were also used to ascertain how proteolysis was affected by release of intracellular enzymes in the initial stage of incubation of the strains in cheese-curd slurries. The presence of strain T1 produced higher levels of non-protein nitrogen (NPN) and amine nitrogen (AN) during the first weeks of incubation when partially lysed cells had been added. Addition of whole cells of strain T1 produced higher values of NPN and AN at the end of incubation of slurries, accelerating proteolysis by about 2 weeks with respect to the control, which only contained the parental strain IFPL359. At the end of the experimental period higher amino acid levels were detected by HPLC in the slurry containing whole T1 cells. Volatile fractions of the different cheese-curd slurries were also analysed. The higher level of proteolysis produced by addition of high levels of strain T1 appeared to be related to release of intracellular enzymes by this strain owing to its greater capacity for autolysis.     

Release and partial characterization of cell-envelope proteinases fromLactococcus lactis subsp.lactis IFPL 359 andLactobacillus casei subsp. casei IFPL 731 isolated from raw goat,s-milk cheese

Different methods of releasing the cell-envelope proteinase (CEP) fromLactococcus lactis IFPL 359 (Lc-CEP) andLactobacillus casei IFPL 731 (Lb-CEP) have been tested. Release of Lc-CEP was higher in Ca^2+-free buffer than in the presence of lysozyme and Ca^2+-. Lb-CEP was not soluble in Ca^2+--free buffer, making necessary the use of chelating agents such as ethylenediaminetetraacetate (EDTA) to attain release yields of 15–20%. Solubilizing the cell wall oflb.casei using lysozyme and mutanolysin improved CEP release yields, even in the presence of Ca^2+-. Two differently charged chromophoric peptides were degraded by whole cells and the soluble fractions studied at different hydrolysis rates in both the strains considered. Based on the specificity of these CEPs for the different substrates, the two proteinases can be placed in the same class as the CEP_I/III mixed-type variants that have been identified in lactococcal proteinases. In both strains ß-casein was hydrolysed more rapidly thanα _s-cascin.     

Formation of methional by Lactococcus lactis IFPL730 under cheese model conditions

The present work describes the capacity of Lactococcus lactis to produce methional and other sulphur compounds derived from methionine (Met) in a cheese model system. Cheese slurries were prepared from pasteurized ewes' skimmed milk, chemically acidified with glucono-'-lactone and homogenized aseptically adding Met, !-ketoglutarate, pyridoxal 5'-phosphate, thiamine pyrophosphate and NaCl. Slurries were incubated at 12 °C for 14 days with cellular suspensions of L. lactis IFPL359, L. lactis IFPL730 and L. lactis NCDO763 in different combinations. Slurries added with resting cells and the intracellular fraction from L. lactis IFPL730 showed the highest production of methional at the outset of incubation, which decreased during incubation along with a concomitant increase in 3-methylthiopropanol. The sensorial analysis of slurries indicated a characteristic methional aroma (cooked potato-like) in samples containing 4-methylthio-2-ketobutyrate and the intracellular fraction from L. lactis IFPL730. As incubation proceeded, the intensity of methional aroma decreased but samples were judged by the panel tasters as developing a cheese-like flavour.     

Production of beta-glucosidase and hydrolysis of isoflavone phytoestrogens by Lactobacillus acidophilus, Bifidobacterium lactis, and Lactobacillus casei in soymilk

ABSTRACT:  The study determined β-glucosidase activity of commercial probiotic organisms for hydrolysis of isoflavone to aglycones in fermenting soymilk. Soymilk made with soy protein isolate (SPI) was fermented with Lactobacillus acidophilus LAFTI^® L10, Bifidobacterium lactis LAFTI^® B94, and Lactobacillus casei LAFTI^® L26 at 37 °C for 48 h and the fermented soymilk was stored for 28 d at 4 °C. β-Glucosidase activity of organisms was determined using ρ-nitrophenyl β-D-glucopyranoside as a substrate and the hydrolysis of isoflavone glycosides to aglycones by these organisms was carried out. The highest level of growth occurred at 12 h for L. casei L26, 24 h for B. lactis B94, and 36 h for L. acidophilus L10 during fermentation in soymilk. Survival after storage at 4 °C for 28 d was 20%, 15%, and 11% greater ( P < 0.05) than initial cell counts, respectively. All the bacteria produced β-glucosidase, which hydrolyzed isoflavone β-glycosides to isoflavone aglycones. The decrease in the concentration of β-glycosides and the increase in the concentration of aglycones were significant ( P < 0.05) in the fermented soymilk. Increased isoflavone aglycone content in fermented soymilk is likely to improve the biological functionality of soymilk.     

Combined effects of concentrated thermophilic and mesophilic cultures and conditions of curd acidifications on the manufacture and quality of kasseri cheese

The effect offour process variables on the biological acidification of curd to be usedfor the manufacture of kasseri cheese was examined. The variables were (i) concentrated starter cultures: a thermophilic mixture of Streptococcus thermophilus and Lactobacillus delbrueckii subsp bulgar-icus compared with a blend of Lactococcus lactis subsp lactis, Lactococcus lactis subsp cremoris and Lactobacillus casei subsp casei, and (ii) acidification temperatures: 30 and 40° C for the thermophilic cultures and 20 and 30°C for the mesophiles. The use of concentrated starter cultures enabled the cheese to be made in one day as compared with three in the traditional system usually using raw milk and no culture. The mesophilic blend combined with a lower scalding temperature (35°C) gave a higher yield than the alternative culture, and the cheeses were higher in total solids, acidity andsalt; the same cheeses were also harder and less elastic. The use of mesophilic cultures and a low acidification temperature (20° C) is recommended.     

Peptidase and proteinase activity ofLactococcus lactis, Lactobacillus casei andLactobacillus plantarum

The proteolytic system of several non-commercial strains of lactococci and lactobacilli that were isolated directly from traditional-Spanish, semi-hard, goats' milk cheese was studied. The aminopeptidase, X-prolyldipeptidyl aminopeptidase, dipeptidase and proteinase activity of these new strains was measured for the cytoplasmic, cell-wall/membrane and spontaneously released fractions. The aminopeptidase activity was exclusively intracellular and higher forLactobacillus casei subsp.casei than forLactococcus lactis subsp.lactis. Lactobacillus plantarum showed higher dipeptidase activity thanL. casei. The highest level of proteinase activity was recorded for the cell-wallmembrane fraction ofLactococcus lactis subsp.lactis IFPL 359, and was higher on -casein than on _s-casein for all the strains studied. These results suggest some different contribution of these strains to the proteolysis of cheese during ripening and they seem to complement each other when used together in the starter culture.

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Peptidase- und Proteinase-Aktivität vonLactococcus lactis, Lactobacillus casei undLactobacillus plantarum

Zusammenfassung Das proteolytische System von mehreren nicht kommerziellenLactococcus- undLactobacillus-Stämmen wurde direkt vom traditionellen spanischen halbfesten Ziegenmilchkäse isoliert und untersucht. Die Aktivität von Aminopeptidase, X-Prolyldipeptidylaminopeptidase, Dipeptidase and Proteinase dieser neuen Stämme wurde in cytoplasmatischen, Zellwand-Membran- und spontan freigesetzten Fraktionen gemessen. Die Aminopeptidase-Aktivität erfolgte ausschließlich intracellular und war höher fürLactobacillus casei subsp. casei als fürLactococcus lactis subsp.lactis. Lactobacillus plantarum zeigte höhere Dipeptidase Aktivität alsL. casei. Die höchsten Werte für die Proteinase-Aktivität wurden für die Zellwand-Membran-Fraktion vonLactococcus lactis subsp.lactis IFPL359 gemessen. Für alle untersuchten Stämme war die Aktivität höher bei -Casein als bei _s-Casein. Dieses Ergebnis weist auf den unterschiedlichen Einfluß dieser Stämme bei der Proteolyse von Käse während der Reifung hin. Die Stämme scheinen sich gegenseitig zu ergänzen, wenn sie gemeinsam in Starterkulturen verwendet werden.

     

Cell membrane damage induced by lacticin 3147 enhances aldehyde formation in Lactococcus lactis IFPL730

Amino acid catabolism is mainly initiated in Lactococcus lactis by a transamination reaction that leads to the formation of alpha-keto acids. In addition, a novel alpha-keto acid decarboxylase enzyme, rare in lactic acid bacteria, responsible for the conversion of alpha-keto acids into aldehydes has been reported in L. lactis IFPL730. The effect of lacticin 3147-induced cell damage on both amino acid transamination and alpha-keto acid decarboxylation by L. lactis IFPL730 leading to the formation of aldehydes from amino acids was investigated. Cell membrane permeabilization induced by lacticin 3147 facilitated the diffusion of amino acids into the cells and thus, enhanced amino acid transamination and formation of alpha-keto acids. However, alpha-keto acid decarboxylation was not affected by cell membrane permeabilization since decarboxylation of alpha-keto acids in both control and lacticin 3147-treated cells were similar, suggesting that these substrates could freely diffuse inside the cells. Nevertheless, the formation of 2-methylbutyraldehyde from isoleucine was enhanced in lacticin 3147-treated cells. The increase in alpha-keto acids formation rate by L. lactis IFPL730 due to lacticin 3147-induced cell damage, led to a concomitant increase in the subsequent decarboxylation reaction that complete the metabolic pathway to aldehyde production from amino acids. The present study points out to the use of the food grade lacticin 3147 along with L. lactis IFPL730 as a valuable tool in the development of cheese flavour.     

Characterization of the lactic acid bacteria in ewe's milk and cheese from northwest Argentina

Indigenous lactic acid bacteria in ewe's milk and artisanal cheese were studied in four samples of fresh raw milk and four 1-month-old cheeses from the provinces of northwest Argentina. Mean growth counts on M17, MRS, and MSE agar media did not show significant differences (P < 0.05) in raw milk and cheeses. Isolates of lactic acid bacteria from milk were identified as Enterococcus (48%), lactococci (14%), leuconostocs (8%), and lactobacilli (30%). All lactococci were identified as Lactococcus lactis (subsp. lactis and subsp. cremoris). Lactobacilli were identified as Lactobacillus plantarum (92%) and Lactobacillus acidophilus (8%). Enterococci (59%) and lactobacilli (41%) were isolated from cheeses. L. plantarum (93%), L. acidophilus (5%), and Lactobacillus casei (2%) were most frequently isolated. L. lactis subsp. lactis biovar diacetylactis strains were considered as fast acid producers. L. lactis subsp. cremoris strains were slow acid producers. L. plantarum and L. casei strains identified from the cheeses showed slow acid production. The majority of the lactobacilli and Lactococcus lactis strains utilized citrate and produced diacetyl and acetoin in milk. Enzyme activities (API-ZYM tests) of lactococci were low, but activities of L. plantarum strains were considerably higher. The predominance of L. plantarum in artisanal cheese is probably important in the ripening of these cheeses due to their physiological and biochemical characteristics.     

Probiotic Cheddar Cheese: Influence of Ripening Temperatures on Proteolysis and Sensory Characteristics of Cheddar Cheeses

ABSTRACT:  Bifidobacterium longum 1941, B. animalis subsp. lactis LAFTI^® B94, Lactobacillus casei 279, Lb. casei LAFTI L26, Lb. acidophilus 4962, or Lb. acidophilus LAFTI L10 were used as an adjunct in the production of Cheddar cheeses, which were ripened at 4 and 8 °C for 24 wk. Effects of ripening temperatures and probiotic adjuncts on proteolysis and sensory evaluation of the cheeses were examined. Higher ripening temperature increased the level of proteolysis in the cheeses. Product of proteolysis and organic acids released during ripening were shown to be important for the flavor of Cheddar cheeses. There were positive and significant correlations between the levels of soluble nitrogen, lactic, acetic, and butyric acids, percentage hydrolysis of α_s1-CN and β-CN to the scores of cheddary flavor ( P < 0.05). Scores for sour-acid and vinegary flavors were higher in cheeses with the addition of Bifidobacterium sp. or Lb. casei 279 ripened at 8 °C. The scores were positively and significantly correlated to the level of lactic, acetic, and free amino acids in the cheeses ( P < 0.05). The results show that both 4 and 8 °C have potential for use in the ripening of probiotic Cheddar cheeses.     

Is thermotolerance correlated to heat-shock protein synthesis in Lactococcus lactis subsp. lactis?

Exposure of Lactococcus lactis subsp. lactis cells to a heat shock at 40 degrees C for 30 min induces thermotolerance, the increased ability of bacterial cells to survive exposure to lethal temperature (52 degrees C for 25 min). This transient state of thermal resistance is accompanied, as in Escherichia coli, by the synthesis of a new set of specific proteins termed heat-shock proteins (Hsps). Pre-treatment of the bacterial cells by antibiotics (streptomycin, spiramycin, kanamycin and erythromycin) known to act on translation, induces the major Hsps synthesis but no thermal protection; conversely, puromycin and amino acid analogues treatments, known to produce abnormal and incomplete peptides, triggers the thermotolerance state without inducing significant Hsps synthesis. These results demonstrate that heat-shock response and induced thermotolerance are not tightly correlated phenomena in L. lactis subsp. lactis.     

The accelerating effect of aLactococcus lactis subsp.lactis lactose-utilization/proteinase-deficient strain on proteolysis and flavour development

This study describes the accelerating effect exerted on proteolysis and flavour development in cheese-curd slurries by combined application ofLactococcus lactis subsp.lactis IFPL359 and high concentrations of its Lac^– Prt^– derivative, (i.e. that which has reduced capacity to metabolize lactose and reduced proteolytic activity, strain T1). Cells of strain T1 partially lysed by either sonication or incubation with lysozyme, were also used to ascertain how proteolysis was affected by release of intracellular enzymes in the initial stage of incubation of the strains in cheese-curd slurries. The presence of strain T1 produced higher levels of non-protein nitrogen (NPN) and amine nitrogen (AN) during the first weeks of incubation when partially lysed cells had been added. Addition of whole cells of strain T1 produced higher values of NPN and AN at the end of incubation of slurries, accelerating proteolysis by about 2 weeks with respect to the control, which only contained the parental strain IFPL359. At the end of the experimental period higher amino acid levels were detected by HPLC in the slurry containing whole T1 cells. Volatile fractions of the different cheese-curd slurries were also analysed. The higher level of proteolysis produced by addition of high levels of strain T1 appeared to be related to release of intracellular enzymes by this strain owing to its greater capacity for autolysis.     

Isolation of halotolerant Lactococcus lactis subsp. lactis from intestinal tract of coastal fish

We isolated lactic acid bacteria from the intestinal tract of the pufferfish Takifugu niphobles caught in Shimoda, Shizuoka, Japan by using MRS broth prepared with 50% seawater. Additional screening was carried out using phenotypic tests such as Gram staining, cell morphology, catalase, oxidase and fermentation of glucose. Subsequently 227 isolates screened by the phenotypic tests were subjected to species-specific PCR for Lactococcus lactis, resulting in four positive isolates. The 16S rRNA gene sequences from three isolates were highly similar to that of L. lactis subsp. lactis (DNA database accession number M58837), while that of one isolate was identical to that of Leuconostoc mesenteroides (AB023246). These isolates were characterized by API 50 CH for carbohydrate fermentation and other phenotypic criteria for salt tolerance, and the characteristics were compared with those of L. lactis subsp. lactis from a cheese starter culture. The carbohydrate fermentation profiles of these isolates were characteristic of L. lactis subsp. lactis strains, whereas the tolerance of these isolates to salt was higher than that of L. lactis subsp. lactis from the cheese starter culture: the new L. lactis isolates showed high salt tolerance in MRS-agar plates containing 200% seawater or 6% sodium chloride. This is the first report of the isolation of halotolerant strains of L. lactis subsp. lactis from a marine environment.     

Comparison of newly isolated strains of Lactobacillus delbrueckii subsp. lactis for hydrogen peroxide production at 5 degrees C

Isolates of Lactobacillus delbrueckii subsp. lactis obtained from raw milk samples were compared for the ability to produce hydrogen peroxide (H2O2) at 5 degrees C. Nineteen out of 101 lactobacilli isolated were identified as L. delbrueckii subsp. lactis. The isolates of L. delbrueckii subsp. lactis from most raw milk samples produced more H2O2 than did isolates of other species of lactobacilli from the same samples. Seven isolates of L. delbrueckii subsp. lactis, which produced the highest levels of H2O2 at 5 degrees C were selected for comparison with a laboratory strain, L. delbrueckii subsp. lactis I. In 24 h, isolate RM2-5 produced 7.0 microg/10(9) cfu in buffer containing 5 mM sodium lactate and 4.4 microg/10(9) cfu in buffer containing 5 mM glucose. Three other isolates also produced more H2O2 on sodium lactate than on glucose. However, three remaining new isolates produced more H2O2 on glucose than on sodium lactate. All seven of the most active new isolates of L. delbrueckii subsp. lactis produced significantly higher concentrations of H2O2 than did L. delbrueckii subsp. lactis I in both solutions. Strain RM2-5 produced more H2O2 than did the other six most active newly isolated strains of L. delbrueckii subsp. lactis in this comparison.     

Viability of probiotic micro-organisms (Lactobacillus acidophilus La-5 and Bifidobacterium animalis subsp. lactis Bb-12) in ice cream

The purpose of this research was to determine the survival of two probiotic micro-organisms in ice creams (4% fat). The micro-organisms were Lactobacillus acidophilus La-5 and Bifidobacterium animalis subsp . lactis Bb-12 . To meet this objective, an ice cream mixture was formulated and subjected to three treatments. Treatment 1 was inoculated with L. acidophilus , treatment 2 with B. lactis and the third treatment was inoculated with a mixture of both bacteria inoculated in 1 : 1 proportions. The inoculation was with 4% culture for each treatment. The final products were stored at −25°C for 60 days. The ice cream inoculated with L. acidophilus had a final concentration of 2 × 10 ⁶ cfu/g and the survival rate was 87%. The treatment inoculated with B. lactis had a final concentration of 9 × 10 ⁶ cfu/g, with a logarithmic decrease of 10%. When both micro-organisms were inoculated together, the survival rate was 86%.     

镉胁迫下泡菜乳酸乳球菌的形态变化

通过扫描电镜和透射电镜分别观察不同质量浓度水平的Cd2+对泡菜乳酸乳球菌（Lactococcus lactis subsp.lactis）细胞的影响,扫描电镜结果显示：Cd2+质量浓度在0、10 mg/L时,泡菜乳酸乳球菌呈椭圆形、表面光滑、菌体生长繁殖旺盛,随着Cd2+质量浓度的增加菌体细胞表面产生白色颗粒状物质、菌体细胞存活数量大幅下降（OD600 nm值由1.336下降到0.515）。当添加200 mg/L Cd2+时,几乎没有见到明显的菌体、显示有少量棱形晶状物。透射电镜结果显示：当Cd2+质量浓度为0～50 mg/L时泡菜乳酸乳球菌结构完整、细胞内容物分布均、菌体生长较为正常,当菌体暴露于100、200 mg/L Cd2+时菌体细胞出现异常现象,如细胞破裂、内容物从薄膜穿孔中释放、质壁分离等。两类电镜结果均表明：在低质量浓度Cd2+（≤50 mg/L）胁迫下,对泡菜乳酸乳球菌的生长几乎不产生影响,添加Cd2+质量浓度上升到100、200 mg/L时泡菜乳酸乳球菌正常生长受到抑制。     

Impact of autolytic, proteolytic, and nisin-producing adjunct cultures on biochemical and textural properties of cheddar cheese

The effect of incorporating a highly autolytic strain (Lactobacillus delbrueckii subsp. bulgaricus UL12) a proteolytic strain (Lactobacillus casei subsp. casei L2A), or a nisin Z-producing strain (Lactococcus lactis, subsp. lactis biovar diacetylactis UL719) into Cheddar cheese starter culture (Lactococcus lactis KB and Lactococcus cremoris KB) on physicochemical and rheological properties of the resultant cheeses was examined. Cheeses were ripened at 7 degrees C and analyzed over a 6-mo period for viable lactococcal and lactobacilli counts, pH, titratable acidity (TA), lipolysis, proteolysis, and textural characteristics. The combination of the nisin-producing strain and autolytic adjuncts significantly increased the production of water-soluble nitrogen, free amino acids, and free fatty acids. The effect of Lc. diacetylactis UL719 alone or of Lb. casei L2A on water-soluble nitrogen and free amino acid contents were also significant, whereas their effect on free fatty acids was not. Viable counts of Lb. bulgaricus UL12 were significantly reduced in the presence of Lc. diacetylactis UL719. Lactobacilli-containing cheeses showed significantly lower values for hardness, fracturability, and springiness. It could be concluded that the addition of Lb. bulgaricus UL12 together with a nisin-producing strain produces a greater increase in cheese proteolysis and an improvement in Cheddar cheese texture.     

Characterization of bacteriocins from two Lactococcus lactis subsp. lactis isolates

In this study, bacteriocins from two Lactococcus lactis subsp. lactis isolates from raw milk samples in Turkey designated OC1 and OC2, respectively, were characterized and identified. The activity spectra of the bacteriocins were determined by using different indicator bacteria including Listeria, Bacillus and Staphylococcus spp. Bacteriocins were tested for their sensitivity to different enzymes, heat treatments and pH values. Loss of bacteriocin activities after alpha-amylase treatment suggested that they form aggregates with carbohydrates. Molecular masses of the purified bacteriocins were determined by SDS-PAGE. PCR amplification was carried out with specific primers for the detection of their structural genes. As a result of these studies, the two bacteriocins were characterized as nisin and lacticin 481, respectively. Examination of plasmid contents of the isolates and the results of plasmid curing and conjugation experiments showed that in L. lactis subsp. lactis OC1 strain the 39.7-kb plasmid is responsible for nisin production, lactose fermentation and proteolytic activity, whereas the 16.0-kb plasmid is responsible for lacticin 481 production and lactose fermentation in L. lactis subsp. lactis OC2 strain.     

Impact of Autolytic and Proteolytic Lactobacilli and Nisin-Producing Culture on Proteolysis and Sensory Characteristics in Cheddar Cheese

ABSTRACT: Cheddar cheeses were made using a nisin-tolerant starter culture with either Lactobacillus delbrueckii subsp. bulgaricus UL12 (autolytic strain), Lactobacillus casei subsp. casei L2A (proteolytic strain), Lactococcus lactis subsp. lactis biovar. diacetylactis UL719 (nisin producer), or of Lb. bulgaricus UL12 and Lc. diacetylactis UL719. Lb. bulgaricus UL12 produced more trichloroacetic acid-soluble nitrogen than did Lb. casei L2A, which produced more phosphotungstic acid-soluble nitrogen than did Lc. diacetylactis UL719. High-performance liquid chromatography analyses showed that either lactobacilli or Lc. diacetylactis UL719 increased the hydrophilic and hydrophobic peptide contents. Cheeses containing both Lb. bulgaricus UL12 and Lc. diacetylactis UL719 had the most intense old Cheddar cheese flavor after 6 mo of ripening.