Antioxidant capacity and identification of radical scavenging peptides from Crema de Chiapas, Fresco and Cocido cheeses.

Bioactive peptides may positively impact bodily functions. One of these are the antioxidant peptides which are well documented for a wide variety of food matrices, mostly from plant sources. Nevertheless, information of antioxidant milk-derived peptides is still a little-known field. The present study was aimed to evaluating the antioxidant capacity (AC) in vitro of water soluble extracts < 3 kDa (WSE) from three artisanal Mexican cheeses: Crema de Chiapas (CrC), Fresco (FC) and Cocido (CC). This study was carried out for cheeses of different days of storage (0, 5, 10, 15, and 20) at 4 °C. AC was assayed to the respective WSE by 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) diamoniun salt (ABTS) and oxygen radical absorbance capacity (ORAC) methods and those WSE that showed the most antioxidant capacity from each cheese were analyzed by using RP-HPLC/MS to identify and characterize the novel specific peptides. All the WSE analyzed show antioxidant capacity, especially those from CrC and CC which display the highest AC at 15 days of storage. Regarding to WSE from FC, the AC was constant during storage. Identified structures reveal that these novel peptides possess high content of specific amino acids, mainly proline, valine, leucine and phenylalanine, of which it has already been shown antioxidant properties. This study demonstrate that these artisanal Mexican cheeses are sources of potential antioxidant peptides.

Protective role of lactic acid bacteria and yeasts as dietary carcinogen-binding agents - a review.

The importance of food contaminants in the link between diet and cancer has been widely demonstrated. Therefore, different physical and chemical strategies for the control of human exposure to such dietary carcinogens has been explored; however, most of these strategies are complex, costly, and have low efficiency which limited their applications. Hence, microbiological methods have been receiving more attention. Recent in vitro and in vivo studies have indicated that lactic acid bacteria (LAB) and yeast may act as dietary carcinogen-binding agents. This review describes the promising protective role of strains belonging mainly to the Lactobacillus, Bifidobacterium and Saccharomyces genera by acting as dietary carcinogen-binding agents. This property suggests that these microorganisms may have a protective role by reducing the bioaccessibility of dietary carcinogens, thereby decreasing their toxic effects. The mechanisms by which the binding process takes place have not been completely elucidated; thus, the possible underlying mechanisms and factors influencing carcinogens-binding will be addressed.

Protective Effect of the Intracellular Content from Potential Probiotic Bacteria against Oxidative Damage Induced by Acrylamide in Human Erythrocytes.

The aim of this study was to assess the protective effect of the intracellular content obtained from potential probiotic bacteria against acrylamide-induced oxidative damage in human erythrocytes. First, the antioxidant properties of 12 potential probiotic strains was evaluated. Two commercial probiotic bacteria were included as reference strains, namely, Lactobacillus casei Shirota and Lactobacillus paracasei 431. Data showed that the intracellular content from four strains, i.e., Lactobacillus fermentum J10, Lactobacillus pentosus J24 and J26, and Lactobacillus pentosus J27, showed higher (P < 0.05) antioxidant capacity in most methods used. Thereafter, the intracellular content of such pre-selected strains was able to prevent the disturbance of the antioxidant system of human erythrocytes exposed to acrylamide, thereby reducing cell disruption and eryptosis development (P < 0.05). Additionally, the degree of oxidative stress in erythrocytes exposed to acrylamide was significantly (P < 0.05) reduced to levels similar to the basal conditions when the intracellular content of Lact. fermentum J10, Lact. pentosus J27, and Lact. paracasei 431 were employed. Hence, our findings suggest that the intracellular contents of specific Lactobacillus strains represent a potential source of metabolites with antioxidant properties that may help reduce the oxidative stress induced by acrylamide in human erythrocytes.

In Silico Prediction and In Vitro Assessment of Multifunctional Properties of Postbiotics Obtained From Two Probiotic Bacteria.

In this study, a global metabolite profile using Raman spectroscopy analysis was obtained in order to predict, by an in silico prediction of activity spectra for substance approach, the bioactivities of the intracellular content (IC) and cell wall (CW) fractions obtained from Lactobacillus casei CRL 431 and Bacillus coagulans GBI-30 strains. Additionally, multifunctional in vitro bioactivity of IC and CW fractions was also assessed. The metabolite profile revealed a variety of compounds (fatty acids, amino acids, coenzyme, protein, amino sugars), with significant probable activities (Pa > 0.7) as immune-stimulant, anti-inflammatory, neuroprotective, antiproliferative, immunomodulator, and antineoplastic, among others. Moreover, in vitro assays exhibited that both IC and CW fractions presented angiotensin-converting enzyme-inhibitory (> 90%), chelating (> 79%), and antioxidant (ca. 22-57 cellular antioxidant activity units) activities. Our findings based on in silico and in vitro analyses suggest that L. casei CRL 431 and B. coagulans GBI-30 strains appear to be promising sources of postbiotics and may impart health benefits by their multifunctional properties.

Potential role of natural bioactive peptides for development of cosmeceutical skin products.

In recent years, consumers' demand for cosmeceutical products with protective and therapeutic functions derived from natural sources have caused this industry to search for alternative active ingredients. Bioactive peptides have a wide spectrum of bioactivities, which make them ideal candidates for development of these cosmeceutical products. In vitro studies have demonstrated that bioactive peptides (obtained as extracts, hydrolysates, and/or individual peptides) exhibit biological properties including antioxidant, antimicrobial, and anti-inflammatory activities, in addition to their properties of inhibiting aging-related enzymes such as elastase, collagenase, tyrosinase and hyaluronidase. Some studies report multifunctional bioactive peptides that can simultaneously affect, beneficially, multiple physiological pathways in the skin. Moreover, in vivo studies have revealed that topical application or consumption of bioactive peptides possess remarkable skin protection. These properties suggest that bioactive peptides may contribute in the improvement of skin health by providing specific physiological functions, even though the mechanisms underlying the protective effect have not been completely elucidated. This review provides an overview of in vitro, in silico and in vivo properties of bioactive peptides with potential use as functional ingredients in the cosmeceutical field. It also describes the possible mechanisms involved as well as opportunities and challenges associated with their application.

Modulatory Effect of the Intracellular Content of Lactobacillus casei CRL 431 Against the Aflatoxin B1-Induced Oxidative Stress in Rats.

It has been recognized that lactic acid bacteria exhibit antioxidant properties, which have been mainly endorsed to the intact viable bacteria. However, recent studies have shown that intracellular content (IC) may also be good sources of antioxidative metabolites, which may potentially contribute to oxidative homeostasis in vivo. Hence, the modulatory effect of the intracellular content of Lactobacillus casei CRL 431 (IC431) on aflatoxin B1 (AFB1)-induced oxidative stress in rats was evaluated on the basis of its influence on hepatic lipid peroxidation (LPO), antioxidant status-antioxidant capacity (TAC), catalase (CAT), and glutathione peroxidase (GPx) activities; and on the oxidative stress index (OSi). Results demonstrated that CAT and GPx activities, and TAC, determined in plasma samples, were significantly (P < 0.05) higher in rats treated with AFB1 plus IC431 (3.98 µM/min/mg protein, 1.88 µM/min/mg protein, and 238.7 µM Trolox equivalent, respectively) than AFB1-treated rats (3.47 µM/min/mg protein, 1.46 µM/min/mg protein, and 179.7 µM Trolox equivalent, respectively). Furthermore, plasma and liver tissue samples from rats treated with AFB1 plus IC431 showed significantly (P < 0.05) lower LPO values (52 and 51%, respectively) and OSi (59 and 51%, respectively) than AFB1-treated rats. Hence, our results proved that the intracellular content of Lact. casei CRL 431 contains metabolites that are capable to modulate the antioxidant defense systems in living organism, which may help to ameliorate the damage associated to AFB1-induced oxidative stress.

Assessment of multifunctional activity of bioactive peptides derived from fermented milk by specific Lactobacillus plantarum strains.

Milk-derived bioactive peptides with a single activity (e.g., antioxidant, immunomodulatory, or antimicrobial) have been previously well documented; however, few studies describe multifunctional bioactive peptides, which may be preferred over single-activity peptides, as they can simultaneously trigger, modulate, or inhibit multiple physiological pathways. Hence, the aim of this study was to assess the anti-inflammatory, antihemolytic, antioxidant, antimutagenic, and antimicrobial activities of crude extracts (CE) and peptide fractions (<3 and 3-10 kDa) obtained from fermented milks with specific Lactobacillus plantarum strains. Overall, CE showed higher activity than both peptide fractions (<3 and 3-10 kDa) in most of the activities assessed. Furthermore, activity of <3 kDa was generally higher, or at least equal, to the 3 to 10 kDa peptide fractions. In particular, L. plantarum 55 crude extract or their fractions showed the higher anti-inflammatory (723.68-1,759.43µg/mL of diclofenac sodium equivalents), antihemolytic (36.65-74.45% of inhibition), and antioxidant activity [282.8-362.3µmol of Trolox (Sigma-Aldrich, St. Louis, MO) equivalents]. These results provide valuable evidence of multifunctional role of peptides derived of fermented milk by the action of specific L. plantarum strains. Thus, they may be considered for the development of biotechnological products to be used to reduce the risk of disease or to enhance a certain physiological function.

Evaluation of acrylamide-removing properties of two Lactobacillus strains under simulated gastrointestinal conditions using a dynamic system.

The aim of this study was to evaluate the capability of Lactobacillus reuteri NRRL 14171 and Lactobacillus casei Shirota to remove dietary acrylamide (AA) under simulated gastrointestinal conditions using a dynamic system. The effects of different AA levels or bacteria concentration on toxin removal by Lactobacillus strains were assessed. Thereafter, AA-removing capability of bacteria strains under either fasting or postprandial simulated gastrointestinal conditions was evaluated. Commercial potato chips were analyzed for their AA content, and then used as a food model. Average AA content (34,162µg/kg) in potato chips exceeded by ca. 34-fold the indicative values recommended by the EU. Toxin removal ability was dependent on AA content and bacterial cell concentration. A reduction on bacterial viability was observed in the food model and at the end of both digestive processes evaluated. However, bacteria survived in enough concentrations to remove part of the toxin (32-73%). Both bacterial strains were able to remove AA under different simulated gastrointestinal conditions, being L. casei Shirota the most effective (ca. 70% removal). These findings confirmed the risk of potato chips as dietary AA exposure for consumers, and that strains of the genus Lactobacillus could be employed to reduce the bioavailability of dietary AA.

Invited review: Fermented milk as antihypertensive functional food.

Over the past decade, interest has risen in fermented dairy foods that promote health and could prevent diseases such as hypertension. This biological effect has mainly been attributed to bioactive peptides encrypted within dairy proteins that can be released during fermentation with specific lactic acid bacteria or during gastrointestinal digestion. The most studied bioactive peptides derived from dairy proteins are antihypertensive peptides; however, a need exists to review the different studies dealing with the evaluation of antihypertensive fermented milk before a health claim may be associated with the product. Thus, the objective of this overview was to present available information related to the evaluation of fermented milk containing antihypertensive peptides by in vitro and in vivo studies, which are required before a fermented functional dairy product may be introduced to the market. Although commercial fermented milks with antihypertensive effects exist, these are scarce and most are based on Lactobacillus helveticus. Thus, a great opportunity is available for the development of functional dairy products with new lactic acid bacteria that support heart health through blood pressure- and heart rate-lowering effects. Hence, the consumer may be willing to pay a premium for foods with important functional benefits.

Screening of Lactobacillus strains for their ability to produce conjugated linoleic acid in milk and to adhere to the intestinal tract.

Conjugated linoleic acid (CLA) has been shown to provide beneficial effects on health; however, the amount consumed in food is far from that required for the desired effects. Thus, increasing the CLA content in dairy foods through milk fermentation with specific lactic acid bacteria (LAB) offers an interesting alternative. Moreover, some LAB may be able to adhere to the intestinal mucosa and produce CLA through endogenous synthesis. Therefore, the objective of this study was to screen LAB isolates for their ability to produce CLA in skim milk and in simulated gastrointestinal conditions. Additionally, the ability of selected CLA-producing LAB to adhere to the intestinal mucosa in a murine model was assessed. Results showed that of 13 strains of Lactobacillus tested, only 4 were able to produce CLA in skim milk supplemented with linoleic acid (13.44 ± 0.78 to 50.9 ± 0.26 µg/mL). Furthermore, these 4 Lactobacillus strains were able to survive and produce CLA in simulated gastrointestinal conditions and to adhere to the intestinal mucosa of Wistar rats after 7 d of oral inoculation with fluorescently labeled bacteria. Accordingly, these 4 Lactobacillus strains may be used to manufacture fermented dairy foods to increase CLA content, and consumption of these fermented milks may result in CLA produced endogenously by these LAB.

Effect of ß-lactoglobulin A and B whey protein variants on cheese yield potential of a model milk system.

Cheese yield mainly depends on the amount and proportion of milk constituents; however, genetic variants of the proteins present in milk may also have an important effect. The objective of this research was to study the effect of the variants A and B of ß-lactoglobulin (LG) on cheese yield using a model system consisting of skim milk powder fortified with different levels of a mixture containing α-lactalbumin and ß-LG genetic variants (A, B, or A-B) in a 1:2 ratio. Fortified milk samples were subjected to pasteurization at 65 °C for 30 min. Miniature cheeses were made by acidifying (pH=5.9) fortified milk and incubating with rennet for 1h at 32 °C. The clot formed was cut, centrifuged at 2,600 × g for 30 min at 20 °C and drained for determining cheese yield. Cheese-yielding capacity was expressed as actual yield (grams of cheese curd per 100g of milk) and dry weight yield (grams of dried cheese curd per 100g of milk). Free-zone capillary electrophoresis was used for determining ß-LG A or B recovery in the curd during rennet-induced coagulation. The presence of ß-LG variant B resulted in a significantly higher actual and dried weight cheese yield than when A or A-B were present at levels ≤ 0.675% of whey protein (WP) addition. Results of free-zone capillary electrophoresis allowed us to infer that ß-LG B associates with the casein micelles during renneting, as shown by an increase in the recovery of this variant in the curd when ß-LG B was added up to a maximum at 0.45% (equivalent to 0.675% WP). In general, actual or dried weight cheese yield increased as WP addition was increased from 0.225 to 0.675%. However, when WP addition ranged from 0.675 to 0.90%, a drastic drop in cheese yield was observed. This behavior may be because an increase in the aggregation of casein micelles with a concomitant inclusion of whey protein in the gel occurs at low levels of WP addition, whereas once the association of WP with the casein micelles reach a saturation point at addition levels higher than 0.675%, rearrangements of the gel network result in larger whey expulsion and syneresis. This knowledge is expected to be useful to maximize cheese yield and optimize processing conditions during cheese and cheese analogs manufacturing.

Antihypertensive and hypolipidemic effect of milk fermented by specific Lactococcus lactis strains.

The antihypertensive and hypolipidemic effects of milk fermented by specific Lactococcus lactis strains in spontaneously hypertensive rats (SHR) were investigated. The SHR were fed ad libitum milk fermented by Lc. lactis NRRL B-50571, Lc. lactis NRRL B-50572, Captopril (40mg/kg of body weight, Sigma-Aldrich Co., St. Louis, MO) or purified water for 4 wk. Results suggested that Lc. lactis fermented milks presented a significant blood pressure-lowering effect. No significant difference was noted among milk fermented by Lc. lactis NRRL B-50571 and Captopril by the second and third week of treatment. Additionally, milk fermented by Lc. lactis strains modified SHR lipid profiles. Milk fermented by Lc. lactis NRRL B-50571 and B-50572 were able to reduce plasma low-density lipoprotein cholesterol and triglyceride contents. Thus, milk fermented by Lc. lactis strains may be a coadjuvant in the reduction of hypertension and hyperlipidemia and may be used as a functional food for better cardiovascular health.

Hypotensive and heart rate-lowering effects in rats receiving milk fermented by specific Lactococcus lactis strains.

Previous studies have demonstrated that milk fermented by specific Lactococcus lactis strains significantly inhibits the activity of angiotensin I-converting enzyme (ACE). However, the relationship between the ACE inhibitor and its in vivo action has revealed discrepancies. Therefore, the aim of the present study was to investigate the antihypertensive and heart rate (HR)-lowering effect of milk fermented by specific L. lactis in a murine model. Spontaneously hypertensive male rats (271 (SD14) g) were randomised into four treatment groups that were orally administered with milk fermented by L. lactis NRRL B-50 571 or L. lactis NRRL B-50 572 at 35 or 50 mg protein/kg body weight (BW), respectively. Further, two more groups were fed with different solutions as controls: a saline solution as the negative control and Captopril (40 mg/kg BW), a proven ACE inhibitor, as the positive control. Blood pressure and HR were monitored by the tail-cuff method before the treatments and at 2, 4, 6 and 24 h post-oral administration. The results demonstrated that milk fermented by L. lactis NRRL B-50 571 as well as by L. lactis NRRL B-50 572 presented an important systolic and diastolic blood pressure- and HR-lowering effect. Thus, milk fermented by specific L. lactis strains may present potential benefits in the prevention and treatment of CVD associated with hypertension in humans.

Novel angiotensin I-converting enzyme inhibitory peptides produced in fermented milk by specific wild Lactococcus lactis strains.

The ability of specific wild Lactococcus lactis strains to hydrolyze milk proteins to release angiotensin I-converting enzyme (ACE) inhibitory peptides was evaluated. The peptide profiles were obtained from the <3 kDa water-soluble extract and subsequently fractionated by reversed-phase HPLC. The fractions with the lowest half-maximal inhibitory concentration estimated values (peptide concentration necessary to inhibit ACE activity by 50%) were Lc. lactis NRRL B-50571 fraction (F)1 (0.034 ± 0.002 µg/mL; mean ± SD) and Lc. lactis NRRL B-50572B F 0005 (0.041 ± 0.003 µg/mL; mean ± SD). All peptide fractions were analyzed by reversed-phase HPLC tandem mass spectrometry. Twenty-one novel peptide sequences associated with ACE inhibitory (ACEI) activity were identified. Several novel ACEI peptides presented peptides encrypted with proven hypotensive activity. In conclusion, specific wild Lc. lactis strains were able to hydrolyze milk proteins to generate potent ACEI peptides. However, further studies are necessary to find out the relationship between Lc. lactis strain proteolytic systems and their ability to biogenerate hypotensive peptides.

Molecular traceability of beef from synthetic Mexican bovine breeds.

Traceability ensures a link between carcass, quarters or cuts of beef and the individual animal or the group of animals from which they are derived. Meat traceability is an essential tool for successful identification and recall of contaminated products from the market during a food crisis. Meat traceability is also extremely important for protection and value enhancement of good-quality brands. Molecular meat traceability would allow verification of conventional methods used for beef tracing in synthetic Mexican bovine breeds. We evaluated a set of 11 microsatellites for their ability to identify animals belonging to these synthetic breeds, Brangus and Charolais/Brahman (78 animals). Seven microsatellite markers allowed sample discrimination with a match probability, defined as the probability of finding two individuals sharing by chance the same genotypic profile, of 10(-8). The practical application of the marker set was evaluated by testing eight samples from carcasses and pieces of meat at the slaughterhouse and at the point of sale. The DNA profiles of the two samples obtained at these two different points in the production-commercialization chain always proved that they came from the same animal.

Angiotensin-converting enzyme inhibitory activity in Mexican Fresco cheese.

The objective of this study was to evaluate if Mexican Fresco cheese manufactured with specific lactic acid bacteria (LAB) presented angiotensin I-converting enzyme inhibitory (ACEI) activity. Water-soluble extracts (3 kDa) obtained from Mexican Fresco cheese prepared with specific LAB (Lactococcus, Lactobacillus, Enterococcus, and mixtures: Lactococcus-Lactobacillus and Lactococcus-Enterococcus) were evaluated for ACEI activity. Specific peptide fractions with high ACEI were analyzed using reverse phase-HPLC coupled to mass spectrometry for determination of amino acid sequence. Cheese containing Enterococcus faecium or a Lactococcus lactis ssp. lactis-Enterococcus faecium mixture showed the largest number of fractions with ACEI activity and the lowest half-maximal inhibitory concentration (IC(50); <10 µg/mL). Various ACEI peptides derived from ß-casein [(f(193-205), f(193-207), and f(193-209)] and α(S1)-casein [f(1-15), f(1-22), f(14-23), and f(24-34)] were found. The Mexican Fresco cheese manufactured with specific LAB strains produced peptides with potential antihypertensive activity.

Angiotensin-converting enzyme inhibitory activity of milk fermented by wild and industrial Lactococcus lactis strains.

Angiotensin I-converting enzyme inhibitory (ACEI) activity was evaluated and compared in <3 KDa water-soluble extracts (WSE) isolated from milk fermented by wild and commercial starter culture Lactococcus lactis strains after 48 h of incubation. The highest ACEI activities were found in WSE from milk inoculated with wild L. lactis strains isolated from artisanal dairy products and commercial starter cultures. On the other hand, the lowest ACEI activities were found in WSE from milk inoculated with wild strains isolated from vegetables. Moreover, the IC(50) values (concentration that inhibits 50% activity) of WSE from artisanal dairy products were the lowest, indicating that these fractions were the most effective in inhibiting 50% of ACE activity. In fact, a strain isolated from artisanal cheese presented the lowest IC(50) (13 µg/mL). Thus, it appears that wild L. lactis strains isolated from artisanal dairy products and commercial starter cultures showed good potential for the production of fermented dairy products with ACEI properties.

Evaluation of aroma generation of Lactococcus lactis with an electronic nose and sensory analysis.

There is an increased interest in exploring the potential of new Lactococcus lactis strains isolated from different natural ecosystems for the production of aroma compounds. Thus, the objective of this study was to screen the aroma generation of Lactococcus lactis strains isolated from different sources by an electronic nose and sensory evaluation for their potential use in starter cheese cultures. Twenty-three strains of Lactococcus lactis were isolated from dairy sources such as artisanal raw-milk cheeses, nondairy sources, and commercial starter cultures (industrial). All the strains were assessed for their ability to produce aromas by an electronic nose and sensory analysis after their incubation in milk. Some phenotypic characteristics of technological importance such as lactose fermentation, proteolytic activity, and citrate utilization were also evaluated. Lactococcus lactis strains showed clear phenotypic differences related to their isolation source. Strains isolated from raw-milk dairy products or commercial starter cultures presented faster lactose fermentation and proteolytic activity than those presented by strains isolated from nondairy sources. Additionally, strains isolated from dairy and nondairy sources presented better citrate utilization than strains isolated from commercial dairy starters. On the other hand, there was not a clear relationship between the source of isolation and the ability of lactococci strains to produce aroma. Principal components analysis of electronic nose data revealed 4 distinctive groups based on aroma profiles. Additionally, odor intensity scores (yogurt-like and Fresco cheese-like) for these 4 groups revealed the nature of their differences. In general, strains from dairy products presented intense yogurt-like and Fresco cheese-like aromas, with the latter being the most intense for one specific strain. On the other hand, the majority of wild strains from nondairy sources presented a stronger yogurt-like aroma, whereas industrial strains presented low intensity for both aroma descriptors. Additionally, an aroma potentiation effect was observed when mixtures of 2 lactococci strains isolated from different sources acted together.

alpha-Ketoglutarate biosynthesis in wild and industrial strains of Lactococcus lactis.

AIMS: This study was carried out to explore the ability of wild and industrial strains of Lactococcus lactis to produce alpha-ketoglutarate (alpha-KG), which is essential during the conversion of amino acids to flavour compounds. METHODS AND RESULTS: Two pathways in alpha-KG biosynthesis were explored in strains of L. lactis isolated from dairy products, vegetables and commercial dairy starter cultures. Half of the strains efficiently converted glutamine to glutamate (Glu) and grew in Glu-free medium. Strains did not present isocitrate dehydrogenase and aconitase activities. However, half of the strains presented glutamate dehydrogenase (GDH) activity. CONCLUSIONS: The ability of L. lactis to synthesize either alpha-KG or Glu via GDH was confirmed. However, L. lactis strains were not able to biosynthesize alpha-KG by the citrate-isocitrate pathway. NADP-GDH activity was mainly found in strains isolated from vegetables, whereas NAD-GDH activity was mainly found in strains isolated from dairy products. SIGNIFICANCE AND IMPORTANCE OF THE STUDY: The origin of isolation highly influenced NAD or NADP-GDH activities. These enzymatic activities may be correlated to the flavour production capacity of the different strains.