Simulated Gastrointestinal Tract Conditions Research Articles

Genetically manipulated phages with improved pH resistance for oral administration in veterinary medicine.

Orally administered phages to control zoonotic pathogens face important challenges, mainly related to the hostile conditions found in the gastrointestinal tract (GIT). These include temperature, salinity and primarily pH, which is exceptionally low in certain compartments. Phage survival under these conditions can be jeopardized and undermine treatment. Strategies like encapsulation have been attempted with relative success, but are typically complex and require several optimization steps. Here we report a simple and efficient alternative, consisting in the genetic engineering of phages to display lipids on their surfaces. Escherichia coli phage T7 was used as a model and the E. coli PhoE signal peptide was genetically fused to its major capsid protein (10 A), enabling phospholipid attachment to the phage capsid. The presence of phospholipids on the mutant phages was confirmed by High Performance Thin Layer Chromatography, Dynamic Light Scattering and phospholipase assays. The stability of phages was analysed in simulated GIT conditions, demonstrating improved stability of the mutant phages with survival rates 102–107 pfu.mL−1 higher than wild-type phages. Our work demonstrates that phage engineering can be a good strategy to improve phage tolerance to GIT conditions, having promising application for oral administration in veterinary medicine.

Commercial Dairy Cow Milk microRNAs Resist Digestion under Simulated Gastrointestinal Tract Conditions

Background: MicroRNAs are small, gene-regulatory noncoding RNA species present in large amounts in milk, where they seem to be protected against degradative conditions, presumably because of their association with exosomes.Objective: We monitored the relative stability of commercial dairy cow milk microRNAs during digestion and examined their associations with extracellular vesicles (EVs).Methods: We used a computer-controlled, in vitro, gastrointestinal model TNO intestinal model-1 (TIM-1) and analyzed, by quantitative polymerase chain reaction, the concentration of 2 microRNAs within gastrointestinal tract compartments at different points in time. EVs within TIM-1 digested and nondigested samples were studied by immunoblotting, dynamic light scattering, quantitative polymerase chain reaction, and density measurements.Results: A large quantity of dairy milk Bos taurus microRNA-223 (bta-miR-223) and bta-miR-125b (∼109–1010 copies/300 mL milk) withstood digestion under simulated gastrointestinal tract conditions, with the stomach causing the most important decrease in microRNA amounts. A large quantity of these 2 microRNAs (∼108–109 copies/300 mL milk) was detected in the upper small intestine compartments, which supports their potential bioaccessibility. A protocol optimized for the enrichment of dairy milk exosomes yielded a 100,000 × g pellet fraction that was positive for the exosomal markers tumor susceptibility gene-101 (TSG101), apoptosis-linked gene 2–interacting protein X (ALIX), and heat shock protein 70 (HSP70) and containing bta-miR-223 and bta-miR-125b. This approach, based on successive ultracentrifugation steps, also revealed the existence of ALIX−, HSP70−/low, and TSG101−/low EVs larger than exosomes and 2–6 times more enriched in bta-miR-223 and bta-miR-125b (P < 0.05).Conclusions: Our findings indicate that commercial dairy cow milk contains numerous microRNAs that can resist digestion and are associated mostly with ALIX−, HSP70−/low, and TSG101−/low EVs. Our results support the existence of interspecies transfer of microRNAs mediated by milk consumption and challenge our current view of exosomes as the sole carriers of milk-derived microRNAs.

생선 내장으로부터 분리된 프로바이오틱 유산균에 의한 히스타민 생산균의 제어

본 연구에서는 생선 내장으로부터 분리된 유산균의 프로바이오틱 특성과 아민 산화효소(diamine oxidase, DAO) 및 박테리오신 생산을 통한 히스타민 분해능을 조사하였다. 조기, 가자미, 명태 및 우럭 내장으로부터 분리된 총 97종의 유산균 중에서 CIL08, CIL16, FIL20, FIL31, PIL45, PIL49, PIL52 및 RIL60 균주는 인공 소화액에 대한 저항성이 강하고, HT-29 상피세포에 대해서도 높은 부착력을 보였으며, 항생제(amoxicillin, ampicillin, erythromycin, penicillin G, streptomycin, tetracycline 및 vancomycin)에 대한 내성도 강한 것으로 나타났다. 게다가 이들 균주들은 히스티딘이 함유된 탈카르복시화 액체배지 내에서 히스타민을 생산하지 않았다. 특히 DAO를 생산하는 것으로 추정되는 CIL08, FIL20, PIL52 및 RIL60 등의 4균주는 히스타민 분해능이 유의하게 높았다. FIL20, FIL31 및 PIL52 유산균이 생산한 박테리오신에 의해선 Enterococcus aerogenes CIH05, Serratia marcescens CIH09, Enterococcus faecalis FIH11, Pediococcus halophilus FIH15, Lactobacillus sakei PIH16, Enterococcus faecium PIH19, Leuconostoc mesenteroides RIH25 혹은 Aeromonas hydrophilia RIH28의 증식과 히스타민 생성량이 유의하게 감소되었다. 또한 생선 내장에서 분리된 히스타민 생성균과 히스타민 분해능 혹은 박테리오신 생산능을 가진 CIL08, FIL20, PIL52 및 RIL60 유산균과 혼합 배양에 의해 히스타민 축적량이 감소되었다. 히스타민 생성을 억제하는 프로바이오틱 유산균의 배양학적 특성과 16S rRNA 염기서열 분석을 통해 Pediococcus pentosaceus CIL08, Lactobacillus plantarum FIL20, Lactobacillus paracasei FIL31, Lactobacillus sakei PIL52 및 Leuconostoc mesenteroides RIL60으로 동정되었다. In this study, we examined in vitro the potential probiotic properties of lactic acid bacteria (LAB) obtained from the fish intestine and their ability to degrade histamine through the production of diamine oxidase (DAO) enzymes and bacteriocin. Among 97 LAB strains isolated from the intestine of croaker, flatfish, pollack, and rockfish, CIL08, CIL16, FIL20, FIL31, PIL45, PIL49, PIL52, and RIL60 isolates exhibited excellent survival rates under simulated gastrointestinal tract conditions, high adhesion ability to HT-29 epithelial cells, and resistance to the antibiotics such as amoxicillin, ampicillin, erythromycin, penicillin G, streptomycin, tetracycline, or vancomycin. In addition, these strains did not produce histamine in decarboxylating broth containing histidine. In particular, 4 strains (CIL08, FIL20, PIL52, and RIL60) that may produce DAO were significantly able to degrade histamine. The bacteriocins produced by FIL20, FIL31, and PIL52 LAB inhibited the growth and histamine production of Enterococcus aerogenes CIH05, Serratia marcescens CIH09, Enterococcus faecalis FIH11, Pediococcus halophilus FIH15, Lactobacillus sakei PIH16, Enterococcus faecium PIH19, Leuconostoc mesenteroides RIH25, or Aeromonas hydrophilia RIH28. Histamine-producing strains isolated from fish intestine were found to reduce histamine accumulation during co-culture with CIL08, FIL20, PIL52, and RIL60 LAB showing histamine degradation or bacteriocin production ability. The probiotic strains preventing histamine formation were identified as Pediococcus pentosaceus CIL08, Lactobacillus plantarum FIL20, Lactobacillus paracasei FIL31, Lactobacillus sakei PIL52, and Leuconostoc mesenteroides RIL60 with high similarity based on 16S rRNA gene sequencing.

Probiotic characteristics of Bacillus strains isolated from Korean traditional soy sauce

The objective of this work was to isolate Bacillus strains from Korean traditional soy sauces, and evaluate them for probiotic potential and safety. Three Bacillus strains, MKSK-E1, MKSK-J1 and MKSK-M1, were selected which were highly resistant to simulated gastrointestinal tract conditions, and showed antimicrobial activity against Bacillus cereus, Listeria monocytogenes, Staphylococcus aureus and Escherichia coli. MKSK-M1 possessed the highest relative survival rate, with 96.0% survival in 0.1% pepsin solution (pH 2.0) and 99.3% survival in 3% bile salt solution for 3 h. All three strains exhibited amylase and protease activity, and utilized more than 19 carbohydrates among 49 tested carbohydrates. MKSK-E1 inhibited the growth of 11 of 13 tested foodborne pathogens. All three selected strains possessed strong antimicrobial activity against E. coli and B. cereus. They had antibiotic susceptibility against 8 tested antibiotics. Also, all strains were non-hemolytic on sheep blood, and non-biogenic amine producers. The results of this work indicate that MKSK-E1, J1 and M1 could be used as probiotic cultures for human consumption and animal feeds and as probiotic starter cultures in food industries.

In vitro screening of lactic acid bacteria isolated from gastrointestinal tract of Atlantic Salmon (Salmo salar) as probiont candidates

A total of 20 lactic acid bacteria (LAB) were isolated from the gastrointestinal tracts of Atlantic salmon (Salmo salar) and screened for antimicrobial activity against six fish pathogens using either a microtiter plate assay or a well diffusion assay. The results indicated that three LAB exhibited antimicrobial activity against at least two pathogens. Based on their partial 16S ribosomal deoxyribonucleic acid (16S rDNA) sequences, these isolates were identified as Lactobacillus farraginis, Pediococcus acidilactici, and P. pentosaceus. In addition, these LAB were able to tolerate the simulated gastrointestinal tract conditions (stomach and intestine), had a good adhesion capacity to intestinal mucus as well as were able to grow in intestinal mucus of Atlantic salmon. These results suggest that these LAB are potential probionts in aquaculture. However, further studies are required to evaluate probiotic properties in aquatic animals.

Technological and safety properties of newly isolated GABA-producing Lactobacillus futsaii strains.

To evaluate the technological and safety properties of Lactobacillus futsaii CS3 and CS5 isolated from Thai fermented shrimp products (Kung-Som) in order to develop a valuable gamma-aminobutyric acid (GABA)-producing starter culture. Both strains showed a high GABA-producing ability (>8mgml(-1) ) in MRS broth containing 20mgml(-1) monosodium glutamate (MSG) for 120h. They also exhibited inhibitory activity against foodborne pathogens and spoilage bacteria. Cell surface hydrophobicity and proteolytic activity were observed in both strains. Strain CS3 survived better under simulated gastrointestinal tract conditions with only 1·5 log-units cell decrease over 8h. Both strains showed the ability to deconjugate taurocholate and taurodeoxycholate acid. Neither virulence genes nor biogenic amine production was detected. Strain CS3 exhibited susceptibility to all tested antibiotics with the exception of vancomycin, while strain CS5 showed resistance to vancomycin, ampicillin and chloramphenicol. Based on the results obtained, Lact.futsaii CS3 is very promising as a GABA-producing and potentially probiotic starter culture strain for applications in functional fermented foods. This study focuses on the technological and safety characteristics of Lact.futsaii CS3 and CS5 including their high GABA-producing capacity for the first time. This provides a way of replacing chemical GABA by natural GABA using a GABA-producing starter culture candidate, at the same time offering the consumer new attractive food products.

Chitosan based nanoparticles as protein carriers for efficient oral antigen delivery

This study aimed to investigate the efficacy of nanoparticles based on chitosan as a vehicle for oral antigen delivery in fish vaccination. Carboxymethyl chitosan/chitosan nanoparticles (CMCS/CS-NPs) loaded extracellular products (ECPs) of Vibrio anguillarum were successfully developed by ionic gelation method. The prepared ECPs-loaded CMCS/CS-NPs were characterized for various parameters including morphology, particle size (312±7.18nm), zeta potential (+17.4±0.38mV), loading efficiency (57.8±2.54%) and stability under the simulated gastrointestinal (GI) tract conditions in turbot. The in vitro profile showed that the cumulative release of ECPs from nanoparticles was higher in pH 7.4 (58%) than in pH 2.0 (37%) and pH 4.5 (29%) after 48h. Fluorescein isothiocyanate-labeled bovine serum albumin (FITC-BSA) was used as model protein antigen and encapsulated in CMCS/CS-NPs for investigating the biodistribution of antigen after oral delivery to turbot in 24h. Oral immunization of ECPs-loaded CMCS/CS-NPs group in turbot showed elevated specific antibody and higher concentrations of lysozyme activity and complement activity in fish serum than ECPs solution. CMCS/CS-NPs loaded with ECPs could enhance both adaptive and innate immune responses than the group treated with ECPs solution and suggested to be a potential antigen delivery system.

Influence of Lipid Phase Composition of Excipient Emulsions on Curcumin Solubility, Stability, and Bioaccessibility

The influence of oil type on the ability of excipient emulsions to improve the solubility, stability, and bioaccessibility of curcumin was examined. Oil-in-water emulsions were prepared using coconut, sunflower, corn, flaxseed, or fish oils. These excipient emulsions were then mixed with powdered curcumin and incubated at 30 or 100 °C. For all oils, more curcumin was transferred from powder to excipient emulsion at 100 °C (190–200 μg/mL) than at 30 °C (30–36 μg/mL), which was attributed to increased curcumin solubility with temperature. Oil type influenced the stability and bioaccessibility of curcumin when excipient emulsions were exposed to simulated gastrointestinal tract conditions, which was attributed to differences in the molecular composition and physicochemical properties of the oils. Overall, the use of fish oil led to the highest effective curcumin bioavailability (38 %). This study provides valuable information for optimizing excipient emulsions to increase curcumin bioavailability in food, supplement, or pharmaceutical applications.

Entrapment in food-grade transglutaminase cross-linked gelatin–maltodextrin microspheres protects Lactobacillus spp. during exposure to simulated gastro-intestinal juices

Dietary intake of probiotic bacteria has been shown to impart health effects, however, maintaining viable cells in foods and during passage of the adverse conditions in the upper gastro-intestinal tract is often a problem. The objective of this research was to develop and characterize novel food-grade phase-separated gelatin–maltodextrin (G-MD) microspheres, where the gelatin was cross-linked with transglutaminase (TGase), to determine if encapsulated probiotic lactic acid bacteria were protected during exposure to simulated upper gastro-intestinal tract conditions. The stability, size, structure and protective ability of G-MD microspheres as a function of different TGase concentrations and gelatin bloom strengths were tested. The G-MD microspheres made with gelatin A 300 bloom and a TGase concentration of 10U/g prevented pepsin-induced degradation of the microspheres in simulated gastric juice (pH2.0, 2h, 37°C), resulting in significantly (p<0.05) higher numbers of survivors due to the buffering effect of intact microspheres (average diameter 46μm). After sequential incubation in simulated gastric (1h) and intestinal juices (pH7.4, 4h, 37°C), survivor levels of each of the three encapsulated Lactobacillus sp. (3C2–10, 21C2–10 and 21C2–12) were reduced by 0.2–1log(CFU/g) as compared to 3–4log(CFU/g) for the free non-encapsulated cells. This study presents a new protein based microencapsulation method, which using all food-grade ingredients protects probiotic lactic acid bacteria during exposure to adverse environmental conditions.

Feruloyl esterase activity is influenced by bile, probiotic intestinal adhesion and milk fat.

Cinnamoyl esterases (CE) are microbial and mammalian intestinal enzymes able to release antioxidant hydroxycinnamic acids from their non-digestible ester-linked forms naturally present in vegetable foods. Previous findings showed that oral administration of Lactobacillus fermentum CRL1446 increased intestinal CE activity and improved oxidative status in mice. The aim of this work was to evaluate the in vitro CE activity of L. fermentum CRL1446 and the effect of bile on this activity, as well as strain resistance to simulated gastrointestinal tract (GIT) conditions and its ability to adhere to intestinal epithelium and influence its basal CE activity. L. fermentum CRL1446 and L. fermentum ATCC14932 (positive control for CE activity) were able to hydrolyse different synthetic hydroxycinnamates, with higher specificity toward methyl ferulate (3,853.73 and 899.19 U/g, respectively). Feruloyl esterase (FE) activity was mainly intracellular in L. fermentum CRL1446 and cell-surface associated in L. fermentum ATCC14932. Both strains tolerated simulated GIT conditions and were able to adhere ex vivo to intestinal epithelium. Pre-incubation of L. fermentum strains with bile increased FE activity in both whole cells and supernatants (~2-fold), compared to controls, suggesting that cells were permeabilised by bile, allowing more substrate to enter the cell and/or leakage of FE enzymes. Three-fold higher FE activities were detected in intestinal tissue fragments with adhered L. fermentum CRL1446 cells compared to control fragments (without bacteria), indicating that this strain provides exogenous FE activity and could stimulate esterase activity in the intestinal mucosa. Finally, we found that milk fat had a negative effect on FE activity of intestinal tissue, in absence or presence of adhered L. fermentum. These results help explaining the increase in intestinal FE activity previously observed in mice fed with L. fermentum CRL1446, and support the potential use of this strain for the development of new functional foods directed to oxidative stress-related ailments.

Dairy milk microRNAs resist in vitro digestion in a gastro‐intestinal tract model

MicroRNAs are small, gene regulatory non‐coding RNA species that are present in large amounts in milk where they seem to be protected against degradative conditions, presumably because of their association with exosomes. We monitored the relative stability of commercial dairy milk microRNAs, under the gastrointestinal (GI) tract conditions that prevails during digestion, and examined their association with extracellular vesicles (EVs). To do so, we used a computer‐controlled TNO in vitro GI model (TIM‐1) and analyzed, by quantitative PCR (qPCR), the level of two milk microRNAs within all GI tract compartments at different time points. MicroRNA‐containing EVs within digested and non‐digested samples were studied by immunoblotting, dynamic light scattering, qPCR and density measurements. We observed that an important proportion of dairy milk bta‐miR‐223 and bta‐miR‐125b withstand digestion under simulated GI tract conditions, with the stomach causing the greater loss in microRNAs. A significant amount of these two microRNAs (~108–109 copies/300 ml milk) was detected in the upper small intestine compartments, which supports their bioavailability. A protocol optimized for the enrichment of dairy milk exosomes yielded a 100,000 g pellet fraction positive for the exosomal marker Tumor susceptibility gene‐101 (TSG‐101) and containing bta‐miR‐223 and bta‐miR‐125b. This approach, based on successive ultracentrifugation steps, also revealed the existence of TSG‐101 negative EVs that are larger than exosomes and more enriched in bta‐miR‐223 and bta‐miR‐125b. Our findings indicate that commercial dairy milk contains numerous microRNAs that can resist digestion and are associated mostly with non‐exosomal EVs. Our results strengthen the idea that milk microRNAs are bioactive compounds in dairy milk, support the existence of interspecies transfer of microRNAs based on milk consumption, and challenge our current view of exosomes as the sole carriers of milk‐derived microRNAs.Support or Funding InformationThis work was supported by Grants No. 319618 and 327522 from the Canadian Institutes of Health Research (CIHR), through the Institute of Genetics (to P.P.).

Development of rice as potential carriers for probiotic Lactobacillus amylovorus

SummaryAn experiment using a standard strain of Lactobacillus amylovorus TISTR1110 to test for the capability of carbohydrate utilisation of nine cultivars of rice found that L. amylovorus TISTR1110 could utilise carbohydrate from all cultivars and also exhibit nonhaemolytic properties. After testing the viability of the strain under simulated gastrointestinal tract conditions by enumeration after 5 h, the results showed that the amount of L. amylovorus TISTR1110 present was 4.0 × 105 CFU/mL or a 73.78% survival rate. From the evaluation of Thai rice cultivars as carriers in probiotic products, the highest amount of probiotic bacteria was found in baked and encapsulated glutinous Luem Pua Thai sticky rice up to 4.9 × 106 CFU/g or a survival rate of 69.39 ± 0.06%. The second highest rate (4.8 × 105 CFU/g) was found in heat–moisture steamed rice of Riceberry rice cultivar or a survival rate of 59.74 ± 0.11%. After testing the capability of adherence using SEM, the results showed that glutinous Luem Pua sticky rice resulted in considerably high survival rate of probiotic bacteria compared with other rice. Additionally, the microbiological safety test reported that the amounts of contaminants were acceptable.

Probiotic Potential of Lactic Acid Bacteria from Traditional Fermented Dairy and Meat Products: Assessment by In Vitro Tests and Molecular Characterization

The aim of the present study was to evaluate the probiotic potential of lactic acid bacteria (LAB) isolated from Greek traditional fermented products. A series of In vitro tests that included survival in simulated gastrointestinal conditions (resistance to low pH, bile salts resistance and bile salts hydrolysis) and safety assessment (resistance to antibiotics, haemolytic and antimicrobial activity) were performed to select potential probiotic candidates, while Lactobacillus rhamnosus GG and Lactobacillus casei Shirota were used as reference strains. Initially, a total of 255 isolates of LAB have been recovered and screened for their survival in simulated gastrointestinal tract conditions and 133 isolates that exhibited moderate or good behavior in these tests were subsequently differentiated and characterized at species level with molecular tools. Pulsed Field Gel Electrophoresis was applied for strain differentiation, while species differentiation was based on restriction analysis of the amplified 16S rRNA gene. Specific multiplex PCR assay targeting the recA genes was applied to resolve the species level of the isolates, belonged to Lb. plantarum group. From the 133 isolates, 47 different strains were recovered and were assigned to Lactobacillus sakei(14), Lactobacillus curvatus (4), Leuconostoc mesenteroides (4), Lactococcus lactis (4), Lactobacillus casei group (1), Lactobacillus brevis (1), Lb. plantarum (10), Lb. pentosus (7) and Lb. paraplantarum (2). The identified strains with good behavior to the gastrointestinal tract tests were selected and further evaluated for their safety aspect. In conclusion, 19 out of the 47 identified strains were assessed as well-behaved, under simulated gastrointestinal conditions and also considered as safe, possessing thus desirable In vitro probiotic properties similar or better to that of the reference strains. These strains may be considered as good candidates for further investigation at in vivo and in situ studies to assess their potential health benefits and their performance as novel probiotic starters or adjunct cultures.

Characterization and in vitro properties of potential probiotic Bifidobacterium strains isolated from breast-fed infant feces

Probiotics are live microorganisms that when administered in adequate amounts, confer health benefits on the host. In this study, 13 strains of Bifidobacterium were isolated from three samples of breast-fed infant feces. The isolates were identified based on conservative gene sequencing and phylogenetic analysis. In vitro tests included survival under simulated gastrointestinal tract conditions, aggregation, hydrophobicity, intestinal epithelial cell adhesion, antimicrobial activity, and antibiotic resistance according to international guidelines for probiotics. The results suggest that B. bifidum, B. adolescentis, and B. breve had high adhesive ability compared with B. longum and B. catenulatum/B. pseudocatenulatum group strains. In particular, B. bifidum IF3-211 has a highest adhesion index (8273 ± 247 and 18,009 ± 1476 adhering bacteria per 100 HT-29 and Caco-2 cells, respectively), far higher than the two reference strains, B. lactis Bb12 and B. longum BBMN68. B. adolescentis IF1-11 showed highest autoaggregation (82.52 ± 0.24 %) and coaggregation (45.59 ± 4.16 %) with L. monocytogenes among isolates. In conclusion, B. bifidum IF3-211 and B. adolescentis IF1-11 showed promising characteristics as probiotic candidates that have good potential for application in food industry.

Insights into the protective role of solid lipid nanoparticles on rosmarinic acid bioactivity during exposure to simulated gastrointestinal conditions

The evaluation of the digestion effects on bioactive solid lipid nanoparticles (SLN) was performed. For this purpose, witepsol and carnauba SLN loaded with rosmarinic acid (RA) were exposed to the simulated gastrointestinal tract (GIT) conditions prevailing in stomach and small intestine. The simulation of intestinal epithelium was made with a dialysis bag and intestinal cell culture lines. Changes on SLN physical properties, RA release and absorption profiles were followed at each step. Combination of digestion pH and enzymes showed a significant effect upon SLN physical properties. Zeta potential values increased at stomach conditions and decreased at small intestine simulation. Also, at intestine, SLN increased their sizes and released 40–60% of RA, maintaining its initial antioxidant activity values. Sustained release of 40% of RA from SLN was also observed in dialysis tube. At CaCo-2 cell line, both types of SLN showed similar absorbed RA % (ca. 30%). Nevertheless, in CaCo-2/HT29x mix cell lines, for carnauba SLN a lower adsorption RA % was observed than for witepsol SLN. Solid lipid nanoparticles protected RA bioactivity (in terms of antioxidant activity) until reaching the intestine. A controlled release of RA from SLN was achieved and a significant absorption was observed at intestinal cells. Overall, SLN produced with witepsol showed a higher stability than carnauba SLN.

Screening for potential novel probiotic Lactobacillus strains based on high dipeptidyl peptidase IV and α-glucosidase inhibitory activity

Lactobacillus strains were screened for inhibition of dipeptidyl peptidase IV (DPP-IV) and α-glucosidase. Cell-free excretory supernatants (CFS) and cell-free extracts (CFE) of 21 strains were prepared, and most of them showed DPP-IV inhibition in a concentration-dependent manner, which started at the exponential phase and peaked at the stationary phase of the Lactobacillus. The CFS bioactivity was heat-resistant, stable to acid–alkali and glucosidase treatment. Trypsin treatment specifically elevated CFS's DPP-IV inhibition. Lactobacillus plantarum strains CFS displayed different protein patterns and trypsin sensitive bands on SDS-PAGE, demonstrating specific peptide-cleavage contributes to higher inhibition. Seven strains with highest DPP-IV inhibition also displayed α-glucosidase inhibitory activity, of which, three strains of L. plantarum and one of Lactobacillus brevis showed tolerance to simulated gastrointestinal tract conditions and high HT-29 cell adhesion. This initial report of lactobacilli secreting strong DPP-IV and α-glucosidase inhibition components will be beneficial for selection and application of anti-diabetic probiotics.

Effects of milk components and food additives on survival of three bifidobacteria strains in fermented milk under simulated gastrointestinal tract conditions.

BackgroundIn the dairy industry, probiotic strains of Bifidobacterium are introduced into the composition of traditional starter cultures intended for the production of fermented foods, or sometimes are the sole microflora responsible for the fermentation process. In order to be able to reach the intestines alive and fulfil their beneficial role, probiotic strains must be able to withstand the acidity of the gastric juices and bile present in the duodenum.ObjectiveThe paper reports effects of selected fermented milk components on the viability of three strains of bifidobacteria in fermented milk during subsequent incubation under conditions representing model digestive juices.DesignThe viability of the bifidobacterial cells was examined after a 3-h incubation of fermented milk under simulated gastric juice conditions and then after 5-h incubation under simulated duodenum juice conditions. The Bifidobacterium strains tested differed in their sensitivity to the simulated conditions of the gastrointestinal juices.ResultsBifidobacterial cell viability in simulated intestinal juices was dependent on the strain used in our experiments, and product components acted protectively towards bifidobacterial cells and its dose.ConclusionsBifidobacterial cells introduced into the human gastrointestinal tract as food ingredients have a good chance of survival during intestinal transit and to reach the large intestine thanks to the protective properties of the food components and depending on the strain and composition of the food.

Stability of bioactive solid lipid nanoparticles loaded with herbal extracts when exposed to simulated gastrointestinal tract conditions

ABSTRACT Solid lipid nanoparticles (SLN) can be used as vehicles for phenolic compounds rich extracts. In the present work two types of waxes — witepsol and carnauba were tested for the first time in the production of solid lipid nanoparticles (WSLN and CSLN, respectively) loaded with sage and savoury extracts. Physical characterization and association efficiencies calculation were performed. Discrimination of loaded phenolic compounds from each extract was made using HPLC assays. Antioxidant activities of SLN were characterized using two different methods — ABTS and ORAC. Finally, the phenolic compound release profile from SLN and stability when exposed to simulated gastrointestinal tract (GIT) conditions were also evaluated. Different phenolic compounds from sage and savoury extracts were entrapped in SLN. The highest antioxidant activity was obtained for the SLN loaded with savoury extract. Stomach simulated condition provokes a partial release of rosmarinic acid from SLN, whereas at small intestine simulation step, all SLN showed a release of ca. 100%. Witepsol SLN were the ones that best maintained their physical integrity during digestion, showing to be the most stable vehicles for sage and savoury extracts. These SLN show to be suitable for the production of food functional ingredients bearing antioxidant activity.

Characterization of clinical and food Listeria monocytogenes isolates with different antibiotic resistance patterns through simulated gastrointestinal tract conditions and environmental stresses

Thirty-three Listeria monocytogenes isolates previously collected from two sources, food (n = 18) and human patients suffering from listeriosis (n = 15), with variable antibiotic susceptibility profiles (sensitive/resistant) to erythromycin, ciprofloxacin and nitrofurantoin, were studied for their ability to survive (i) the environmental stress provided by sequential conditions that simulate the digestive tract, and (ii) extreme pH values (1.5–12). The results showed a response that was only strain dependent. There were no variability in survival results based on type of stress (low or high pH), source (food or clinical), or sensitivity/resistance to antibiotics (p > 0.01). Some strains of L. monocytogenes are able to survive extreme acid and alkaline conditions, and conditions that mimic the sequential stressors found in the gastro-intestinal tract. The resistance to the antibiotics tested in this study by some L. monocytogenes strains did not confer any cross-protection to acid or alkaline stressors.

Influence of sub-lethal stresses on the survival of lactic acid bacteria after spray-drying in orange juice

The demand for new functional non-dairy based products makes the production of a probiotic orange juice powder an encouraging challenge. However, during drying process and storage, loss of viability of the dried probiotic cultures can occur, since the cells are exposed to various stresses. The influence of sub-lethal conditions of temperature, acidic pH and hydrogen peroxide on the viability of Pediococcus acidilactici HA-6111-2 and Lactobacillus plantarum 299v during spray drying in orange juice and subsequent storage under different conditions was investigated. At the end of storage, the survival of both microorganisms through simulated gastro-intestinal tract (GIT) conditions was also determined. The viability of cells previously exposed to each stress was not affected by the drying process. However, during 180 days of storage at room temperature, unlike P. acidilactici HA-6111-2, survival of L. plantarum 299v was enhanced by prior exposure to sub-lethal conditions. Previous exposure to sub-lethal stresses of each microorganism did not improve their viability after passage through simulated GIT. Nevertheless, as cellular inactivation during 180 days of storage was low, both microorganisms were present in numbers of ca. 107 cfu/mL at the end of GIT. This is an indication that both bacteria are good candidates for use in the development of an orange juice powder with functional characteristics.