Sensory evaluation of probiotic Minas fresh cheese with Lactobacillus acidophilus added solely or in co-culture with a thermophilic starter culture

Sensory acceptance of formulations of probiotic Minas fresh cheese was investigated. Cheeses were prepared and supplemented with Lactobacillus acidophilus (T1 – probiotic), Lactobacillus acidophilus + Streptococcus thermophilus (T2 – probiotic + starter) or produced with no addition of cultures (T3 – control). Sensory acceptance tests were performed after 7 and 14 days of storage at 5 °C, using a 9‐point hedonic scale (1 = dislike extremely; 9 = like extremely). After 7 days, no significant difference was detected among cheeses T1, T2 and T3 (P > 0.05). After 14 days, cheeses T1 and T2 presented higher acceptance and differed significantly from cheeses T3. Cheeses T3 presented significant difference between 7 and 14 days of storage (P < 0.05), whereas probiotic cheeses T1 and T2 were stable in the same period (P > 0.05). The addition of L. acidophilus, either solely or in co‐culture with a thermophilic starter culture, resulted in good acceptance of Minas fresh cheese, improving sensory performance of the product during storage.     

An excessively high Lactobacillus acidophilus inoculation level in yogurt lowers product quality during storage

The effect of manufacturing yogurt with a wide variation in Lactobacillus acidophilus inoculation level while holding the yogurt culture inoculation level constant on the properties of the resulting yogurt was determined to find out if any problems can occur if an excessively high level of L. acidophilus is used in yogurt production. Four batches of plain, set-style yogurt were manufactured with skim milk, nonfat dry milk, yogurt cultures, and with or without L. acidophilus (0, 0.0239, 0.238, or 2.33 g/100 g). After homogenization, pasteurization, and cooling, yogurt mixes were inoculated, poured into containers, incubated to pH 4.5, and cooled. Lactobacilli and L. acidophilus counts, pH, amount of syneresis, color, apparent viscosities, and sensory scores were determined during storage. The yogurt inoculated with 0.238 g/100 g L. acidophilus had the highest L. acidophilus counts from 4 to 7 wk. Yogurts inoculated with 2.33 g/100 g L. acidophilus generally had lower lactobacilli counts, L* values, apparent viscosities, and sensory scores but more syneresis and higher a* and b* values than the remaining yogurts. An excessively high inoculated level of L. acidophilus (2.33 g/100 g) resulted in an inferior quality yogurt.     

Synbiotic potential of fresh cream cheese supplemented with inulin and Lactobacillus paracasei in co-culture with Streptococcus thermophilus

The influence of the addition of Lactobacillus paracasei and Streptococcus thermophilus on the fructan content at the beginning and at the end of storage at 4 ± 1 °C of a potentially synbiotic fresh cream cheese manufactured with inulin was investigated. Three cheese-making trials were prepared, all supplemented with a lactic culture of S. thermophilus (T1, T2 and T3). L. paracasei subsp. paracasei was added in T1 and T2. Inulin was added in T2 and the fructan content was measured after 1 and 21 days of storage. Samples of T2 possessed similar mean concentrations of fructans after 1 and 21 days of storage, 7.32% and 7.27%, respectively, and no significant difference was observed. These results indicated that the metabolism of starter and probiotic bacteria did not degrade the fructans present in those cheeses. Additionally, synbiotic cheeses possessed a fructan content higher than 7 g per 100 g, sufficient to confer prebiotic potential during the entire storage period of these products.     

Incorporation of Lactobacillus acidophilus in Minas fresh cheese and its implications for textural and sensorial properties during storage

The effect of Lactobacillus acidophilus on instrumental texture profile and related properties of Minas fresh cheese during storage at 5 °C and on sensory performance was investigated. Four cheese-making trials were prepared, two supplemented with a mesophilic type O culture (T1, T2) and two with lactic acid (T3, T4). L. acidophilus was added in T2 and T3. The viability of L. acidophilus, instrumental texture profile analysis and related properties were monitored during storage for up to 21 days. Probiotic cheeses T3 were firmer by the end of storage, due to higher values of pH and hardness. Differences detected were attributed to the starter, rather than to L. acidophilus. Viability of L. acidophilus during storage ranged from 6.04 to 6.93 for T2 and from 5.46 to 6.53 log cfu g⁻¹ for T3, which performed better in sensory evaluation. Minas fresh cheese is a suitable food system for the delivery of L. acidophilus.     

Effect of calcium alginate and resistant starch microencapsulation on the survival rate of Lactobacillus acidophilus La5 and sensory properties in Iranian white brined cheese

Two types of probiotic cheese, with free and microencapsulated bacteria, were manufactured in triplicate under the same conditions. The number of viable cells during 182 days of storage in refrigerated conditions was evaluated. The number of viable cells of Lactobacillus acidophilus was reduced significantly from day 28 to day 182 of storage period in both types of cheese, but reduction in the cheese containing free cells (5.1 ± 0.67 log cfu g⁻¹) was significantly p < 0.05 higher than the cheese containing microencapsulated cells (11.00 ± 0.58 log cfu g⁻¹). The results showed that, microencapsulation in calcium alginate gel and resistant starch was able to increase the survival rate of L. acidophilus La5 in Iranian white brined cheese after 6 months of storage.     

Temporal gene expression and probiotic attributes of Lactobacillus acidophilus during growth in milk

Lactic acid bacteria have been used as starter strains in the production of fermented dairy products for centuries. Lactobacillus acidophilus is a widely recognized probiotic bacteria commonly added to yogurt and used in dietary supplements. In this study, a whole genome microarray was employed to monitor gene expression of L. acidophilus NCFM cells propagated in 11% skim milk during early, mid and late logarithmic phase, and stationary phase. Approximately 21% of 1,864 open reading frames were differentially expressed at least in one time point. Genes differentially expressed in skim milk included several members of the proteolytic enzyme system. Expression of prtP (proteinase precursor) and prtM (maturase) increased over time as well as several peptidases and transport systems. Expression of Opp1 (oligopeptide transport system 1) was highest at 4 h, whereas gene expression of Opp2 increased over time reaching its highest level at 12 h, suggesting that the 2 systems have different specificities. Expression of a 2-component regulatory system, previously shown to regulate acid tolerance and proteolytic activity, also increased during the early log and early stationary phases of growth. Expression of the genes involved in lactose utilization increased immediately (5 min) upon exposure to milk. The acidification activity, survival under storage conditions, and adhesion to mucin and Caco-2 tissue culture cells of selected mutants containing insertionally inactivated genes differentially expressed in the wild-type strain during growth in milk were examined for any potential links between probiotic properties and bacterial growth and survival in milk. Some of the most interesting genes found to be expressed in milk were correlated with signaling (autoinducer-2) and adherence to mucin and intestinal epithelial cells, in vitro.     

Elaboration of a probiotic oblea from whey fermented using Lactobacillus acidophilus or Bifidobacterium infantis

A novel probiotic product was developed, which was formulated as an oblea (wafer-type dehydrated traditional Mexican dessert) using goat sweet whey fermented with Bifidobacterium infantis or Lactobacillus acidophilus. To obtain the probiotic oblea, the fermented whey was formulated with prebiotic carbohydrates (inulin and resistant starch) and gelatin, and the preparation was poured onto a polytetrafluoroethylene-coated nonstick baking pan, dried in a convection oven, and finally dehydrated at a low relative humidity and room temperature (23 ± 2°C). The amounts of prebiotic carbohydrates and gelatin to be used in the formulation were determined by a factorial experimental design. An untrained sensory panel evaluated 3 quality characteristics (film formation, homogeneity, and smoothness) in the final product. Three different drying temperatures were tested, namely, 40, 55, and 70°C. Bacterial survival at each temperature was determined by viable plate-counting. The best formulation, based on the quality characteristics tested, consisted of 58.33% (vol/vol) of fermented whey, 8.33% (vol/vol) of 6% (wt/vol) resistant starch dispersion, 16.66% (vol/vol) of 15% (wt/vol) inulin solution, and 16.66% (vol/vol) of a 10% (wt/vol) gelatin solution. Drying at 55 ± 2°C for 2.66 ± 0.22 h allowed for concentrations of probiotic bacteria above 9 log₁₀ cfu/g, which is above the minimum concentration required in a probiotic product.     

Free and immobilized Lactobacillus casei ATCC 393 on whey protein as starter cultures for probiotic Feta-type cheese production

The use of free and immobilized Lactobacillus casei ATCC 393 on whey protein as starter culture in probiotic Feta-type cheese production was evaluated. The probiotic cultures resulted in significantly higher acidity; lower pH; reduced counts of coliforms, enterobacteria, and staphylococci; and improved quality characteristics compared with cheese with no culture. Microbiological and strain-specific multiplex PCR analysis showed that both free and immobilized L. casei ATCC 393 were detected in the novel products at levels required for conferring a probiotic effect at the end of the ripening. The effect of starter culture on production of volatile compounds was investigated by the solid-phase microextraction gas chromatography-mass spectrometry analysis technique. The immobilized cells resulted in an improved profile of aroma-related compounds and the overall high quality of the novel products was ascertained by the preliminary sensory test. Finally, the high added value produced by exploitation of whey, which is an extremely polluting industrial waste, was highlighted and assessed.     

A synbiotic containing Lactobacillus acidophilus CHO-220 and inulin improves irregularity of red blood cells

This randomized, double-blind, placebo-controlled, and parallel-design study was conducted to investigate the effect of a synbiotic product containing Lactobacillus acidophilus CHO-220 and inulin on the irregularity in shape of red blood cells (RBC) in hypercholesterolemic subjects. The subjects (n = 32) were randomly allocated to 2 groups, a treatment group (synbiotic product) and a control group (placebo), and received 4 capsules of either synbiotic or placebo daily for 12 wk. Morphological representation via scanning electron microscopy showed that the occurrence of spur RBC was improved upon supplementation of the synbiotic. In addition, the supplementation of synbiotic reduced the cholesterol:phospholipids ratio of the RBC membrane by 47.02% over 12 wk, whereas the control showed insignificant changes. Our present study also showed that supplementation of the synbiotic reduced the concentration of saturated fatty acids (SFA), increased unsaturated fatty acids (UFA), and increased the ratio of UFA:SFA over 12 wk, whereas the control showed inconspicuous changes. The alteration of RBC membrane was assessed using fluorescence anisotropy (FAn) and fluorescence probes with different affinities for varying sections of the membrane phospholipid bilayer. A noticeable decrease in FAn of three fluorescent probes was observed in the synbiotic group compared with the control over 12 wk, indicative of increased membrane fluidity and reduced cholesterol enrichment in the RBC membrane.     

The effect of probiotics (Lactobacillus rhamnosus HN001, Lactobacillus paracasei LPC-37, and Lactobacillus acidophilus NCFM) on the availability of minerals from Dutch-type cheese

The use of probiotic cultures in the production of Dutch-type cheeses did not lead to significant changes in their chemical composition but it lowered their acidity. The availability of calcium and magnesium analyzed by in vitro enzymatic hydrolysis was 19 and 35%, respectively; the availability of phosphorus was significantly higher, at >90%. The use of probiotic cultures significantly increased the availability of calcium (~2.5%), phosphorus (~6%), and magnesium (~18%). The in vitro method supports accurate determination of the effect of the Lactobacillus spp. cultures on the availability of mineral compounds ingested with Dutch-type cheese.     

Survival of human derived Lactobacillus plantarum in fermented cereal extracts during refrigerated storage

The aim of this study was to investigate the survival of a potentially probiotic Lactobacillus plantarum strain in barley, wheat and barley malt extracts. The extracts were produced from three flour/water suspensions, i.e., 5%, 20%, 30% w/w. After inoculation, the cultures were incubated for 24 h at 37 °C, and were subsequently stored at 4 °C for up to seventy days. The lactic acid and reducing sugar concentrations at the beginning of storage were significantly different between the fermented media, ranging from 0.5 g/L to 17 g/L and from 0.8 g/L to 6.5 g/L respectively, while the pH ranged between 2.9 and 3.4. It was observed that the cells survived much better in the malt extracts compared to barley and wheat extracts during refrigerated storage. Based on the results from a study using model media and supplemented cereal extracts it was derived that this was most likely due to their higher sugar concentration and the presence of protective unidentified compounds, albeit the fact that the malt extracts contained higher amounts of lactic acid.     

Proteolysis development in enzyme-modified Cheddar cheese using natural and recombinant enzymes of Lactobacillus rhamnosus S93

Proteolysis in enzyme-modified cheese was investigated with natural crude enzyme or recombinant aminopeptidase, both derived from Lactobacillus rhamnosus S93 in the presence of a commercial proteinase, Neutrase. For production of enzyme-modified cheeses, a cheese slurry was produced and pre-incubated with Neutrase. Natural enzyme or recombinant aminopeptidase (50 units 200 g⁻¹ slurry) was added alone or in combination to the cheese slurries, which were then incubated anaerobically under vacuum at 37 °C for 1, 3 and 6 d. The greatest levels of phosphotungstic acid soluble nitrogen and free amino acids were observed in the enzyme-modified cheese containing natural enzyme followed by the one treated with a combination of the natural enzyme and recombinant aminopeptidase. The enzyme-modified cheese containing the recombinant aminopeptidase alone resulted in the complete disappearance of proline after 1 d of maturation time.     

The influence of Bifidobacterium Bb-12 and lactic acid incorporation on the properties of Minas Frescal cheese

The effects of a probiotic bacterium (Bifidobacterium Bb-12) and lactic acid on the microbiological, physicochemical, rheological and microstructural proprieties of Minas Frescal cheese were evaluated after 1 day and after 28 days of storage (5 ± 1 °C). The cheese was produced with four different treatments: with no lactic acid (C1 and C2), with lactic acid (C3 and C4) and with bifidobacteria (C2 and C3). The cheese formulations containing bifidobacteria were classified as probiotic. The addition of bifidobacteria to the cheese did not influence its yield, protein or lipid levels one day after production. Moreover, after 28 days of storage, the cheese samples with no lactic acid showed lower moisture levels compared to the samples containing lactic acid (P < 0.05). The absence of lactic acid had an influence on the rheological and microstructural behavior, making them more elastic, firm and compact, however all the cheese samples evaluated showed a higher tendency to viscosity than to elasticity. The Minas Frescal cheese with Bifidobacterium Bb-12 and lactic acid showed great potential as a functional food, with possible industrial and commercial application.     

Quantitative profiling of bacteriocins present in dairy-free probiotic preparations of Lactobacillus acidophilus by nanoliquid chromatography-tandem mass spectrometry

Bacteriocins are a heterogeneous group of ribosomally synthesized peptides or proteins with antimicrobial activity, produced predominantly by lactic acid bacteria, with potential applications as biopreservatives and probiotics. We describe here a novel strategy based on a bottom-up, shotgun proteomic approach using nanoliquid chromatography-tandem mass spectrometry (nanoLC-MS/MS) with multiple fragmentation techniques for the quantitative profiling of bacteriocins present in the probiotic preparations of Lactobacillus acidophilus. A direct LC-MS/MS analysis with alternate collision-induced dissociation, high-energy collision dissociation, and electron-transfer dissociation fragmentation following a filter-assisted size-exclusion sample prefractionation has resulted in the identification of peptides belonging to 37 bacteriocins or related proteins. Peptides from lactacin F, helveticin J, lysin, avicin A, acidocin M, curvaticin FS47, and carocin D were predominant. The process of freeze drying under vacuum was observed to affect both the diversity and abundance of bacteriocins. Data acquisition using alternating complementary peptide fragmentation modes, especially electron-transfer dissociation, has significantly enhanced the peptide sequence coverage and number of bacteriocin peptides identified. Multi-enzyme proteolytic digestion was observed to increase the sample complexity and dynamic range, lowering the chances of detection of low-abundant bacteriocin peptides by LC-MS/MS. An analytical platform integrating size exclusion prefractionation, nanoLC-MS/MS analysis with multiple fragmentation techniques, and data-dependent decision tree-driven bioinformatic data analysis is novel in bacteriocin research and suitable for the comprehensive bioanalysis of diverse, low-abundant bacteriocins in complex samples.     

Effect of microencapsulation on viability and other characteristics in Lactobacillus acidophilus ATCC 43121

Lactobacillus acidophilus ATCC 43121 were microencapsulated with sodium alginate by dropping method. The effects of microencapsulation on the changes in survival rate of the L. acidophilus ATCC 43121 during exposure to artificial gastrointestinal and on the change in heat susceptibility of L. acidophilus ATCC 43121 during the heat treatment were studied. In addition, cholesterol assimilation and intestinal adhesion of non-encapsulated and encapsulated L. acidophilus ATCC 43121 were also investigated to explore the effect of microencapsulation on health beneficial effect of lactic acid bacteria. Non-encapsulated cells were completely destroyed when exposed to artificial gastric juice (AGJ) of pH 1.2 and 1.5, while the treatment declined the viable count of encapsulated samples only by 3 log. Encapsulated cells exhibited a significantly higher resistance to artificial intestinal juice (AIJ) and heat treatment than non-encapsulated samples. The assimilative reductions of cholesterol by non-encapsulated and encapsulated L. acidophilus ATCC 43121 were 35.98% and 32.84%, respectively. However, encapsulation did not significantly (P>0.05) affect the adherence of L. acidophilus ATCC 43121 onto the human intestinal epithelial cell lines HT-29. The microencapsulation effectively protected the microorganisms from heat and acid treatment in delivering the viable cells to intestine without any significant adverse effect on their functionalities.     

Evaluation of proteolytic and ACE-inhibitory activity of Lactobacillus acidophilus in soy whey growth medium via response surface methodology

Soy whey is a rich by-product of tofu-manufacturing industries. We have previously optimized the growth of Lactobacillus acidophilus FTCC 0291 in soy whey upon supplementation of meat extract, vegetable extract and peptone using response surface methodology (RSM). The present study evaluated the proteolytic and angiotensin-I converting enzyme (ACE)-inhibitory activities of L. acidophilus FTCC 0291 in the optimized soy whey medium. The probiotic-fermented soy whey exhibited growth-associated proteolysis and ACE-inhibitory activity. Proteolysis was highly correlated with ACE-inhibitory activity, indicating that peptides liberated via fermentation may have exerted in vitro antihypertensive properties. Of the three nitrogen sources studied, peptone was found to have the highest influence on growth performance and ACE-inhibitory activity. Our results strongly indicated that probiotic-fermented soy whey produced in vitro antihypertensive bioactivity, and hence could be further developed into a carrier for probiotics with enhanced functional properties.     

Identification of seven species of the Lactobacillus acidophilus group by FT-IR spectroscopy

Fourier transform infrared (FT-IR) spectra of 102 strains of the seven species of the Lactobacillus acidophilus group were collected and investigated for their potential use in classification and identification on species level. The database built contains more than 370 spectra. Various procedures of pre-processing and classification methods have been compared with respect to their predictive ability. The most encouraging results were achieved with linear discriminant analysis (LDA) of the absorbance values of normalized spectra at selected wavenumbers. The rate of correct species assignment in cross-validation (Jackknife procedure with one spectrum left out for model building) were 95%, 95%, 69%, 100%, 88%, 100%, and 91% for L. acidophilus, L. amylovorus, L. crispatus, L. gallinarum, L. gasseri, L. helveticus, and L. johnsonii, respectively. Very distinct grouping was found for L. gallinarum and L. helveticus, the most difficult differentiation in LDA was between the pairs L. crispatus/L. amylovorus and L. gasseri/L. johnsonii.     

Evolution of aroma volatiles during storage of sourdough breads made by mixed cultures of Kluyveromyces marxianus and Lactobacillus delbrueckii ssp. bulgaricus or Lactobacillus helveticus

Two mixed starter cultures were used for sourdough bread making to evaluate their ability to improve quality and increase bread shelf-life: Lactobacillus delbrueckii ssp. bulgaricus or Lactobacillus helveticus mixed with the lactose fermenting yeast Kluyveromyces marxianus as alternative baker’s yeast. Control sourdough breads (K. marxianus) without the addition of bacteria, were also prepared. The changes on the headspace aroma volatiles during storage were assessed using solid-phase microextraction (SPME) GC–MS analysis. The effect of these changes on bread flavour was evaluated by consumer preference evaluations and the results were co-evaluated with those from the GC–MS analysis. The obtained results showed differences in the volatile composition of the different types of breads examined, as well as dramatic decreases of the number and the amount of volatiles after five days of storage. The sourdough breads made with K. marxianus and L. bulgaricus, had a more complex aroma profile, longer shelf-life and achieved the highest scores in the sensory tests.