Effects of polymerised whey protein-based microencapsulation on survivability of Lactobacillus acidophilus LA-5 and physiochemical properties of yoghurt

Abstract

Lactobacillus acidophilus LA-5 has poor survivability in yoghurts. This study aims to microencapsulate L. acidophilus LA-5 using polymerised whey protein as wall material and evaluate its survivability and effects on physiochemical properties of yoghurts. The microencapsulation yield was 92.90% and average beads size was 744?μm. Microencapsulated L. acidophilus LA-5 showed higher survival at simulated gastrointestinal conditions. Microencapsulated L. acidophilus LA-5 had improved survivability (～10⁶ cfu/mL) than free cells (～10⁴ cfu/mL) after digesting by simulated intestinal juice (6?h) in yoghurts during 10-week storage. Microencapsulated L. acidophilus LA-5 improved the firmness and protein content and decreased the spontaneous whey separation and synaeresis of yoghurts significantly (p?<?0.01). Changes in pH and free microorganism population of yoghurts during storage were not affected by adding microencapsulated L. acidophilus LA-5. Microencapsulation of L. acidophilus LA-5 using polymerised whey protein as wall material improved its survivability and the physiochemical properties of the yoghurt.     

Hot-melt extrusion microencapsulation of quercetin for taste-masking

Besides its poor dissolution rate, the bitterness of quercetin also poses a challenge for further development. Using carnauba wax, shellac or zein as the shell-forming excipient, this work aimed to microencapsulate quercetin by hot-melt extrusion for taste-masking. In comparison with non-encapsulated quercetin, the microencapsulated powders exhibited significantly reduced dissolution in the simulated salivary pH 6.8 medium indicative of their potentially good taste-masking efficiency in the order of zein?>?carnauba wax?>?shellac. In vitro bitterness analysis by electronic tongue confirmed the good taste-masking efficiency of the microencapsulated powders. In vitro digestion results showed that carnauba wax and shellac-microencapsulated powders presented comparable dissolution rate with the pure quercetin in pH 1.0 (gastric) and 6.8 (intestine) medium; while zein-microencapsulated powders exhibited a remarkably slower dissolution rate. Crystallinity of quercetin was slightly reduced after microencapsulation while its chemical structure remained unchanged. Hot-melt extrusion microencapsulation could thus be an attractive technique to produce taste-masked bioactive powders.     

Enrichment of Bifidobacterium longum subsp. infantis ATCC 15697 within the human gut microbiota using alginate-poly-l-lysine-alginate microencapsulation oral delivery system: an in vitro analysis using a computer-controlled dynamic human gastrointestinal model

Abstract

This study evaluates alginate-poly-l-lysine-alginate Bifidobacterium longum subsp. infantis ATCC 15697-loaded microcapsules to enrich the human gut microbiota. The cell survival of alginate-poly-l-lysine-alginate microencapsulated B. infantis ATCC 15697 in gastric acid, bile, and through human gastrointestinal transit was investigated, as well as the formulation’s effect on the gut microbiota. Results show that microencapsulation increases B. infantis ATCC 15697 cell survival at pH1.0 (33.54?±?2.80% versus <1.00?±?0.00%), pH1.5 (41.15?±?2.06% versus <1.00?±?0.00%), pH2.0 (60.88?±?1.73% versus 36.01?±?2.63%), pH3.0 (75.43?±?1.23% versus 46.30?±?1.43%), pH4.0 (71.40?±?2.02% versus 47.75?±?3.12%) and pH5.0 (73.88?±?3.79% versus 58.93?±?2.26%) (p?<?0.05). In addition, microencapsulation increases cell survival at 0.5% (76.85?±?0.80% versus 70.77?±?0.64%), 1.0% (59.99?±?0.97% versus 53.47?±?0.58%) and 2.0% (53.10?±?1.87% versus 44.59?±?1.52%) (p?<?0.05) (w/v) bile. Finally, daily administration of alginate-poly-l-lysine-alginate microencapsulated B. infantis ATCC 15697 in a human gastrointestinal model induces a significant enrichment of B. infantis within the ascending (184.51?±?17.30% versus 53.83?±?17.82%; p?<?0.05), transverse (174.79?±?25.32% versus 73.17?±?15.30%; p?<?0.05) and descending (94.90?±?25.22% versus 46.37?±?18.93%; p?>?0.05) colonic microbiota.     

Microencapsulation of Lactobacillus plantarum (mtcc 5422) by spray-freeze-drying method and evaluation of survival in simulated gastrointestinal conditions

Spray-drying (SD) and freeze-drying (FD) are widely used methods for microencapsulation of heat-sensitive materials like probiotics for long-term preservation and transport. Spray-freeze-drying (SFD) is relatively a new technique that involves spraying a solution into a cold medium and removal of solvent (water) by conventional vacuum FD method. In this study, the SFD microencapsulated Lactobacillus plantarum powder (1:1 and 1:1.5 core-to-wall ratios of whey protein) is compared with the microencapsulated powders produced by FD and SD methods. The SFD and FD processed microencapsulated powder show 20% higher cell viability than the SD samples. In simulated gastrointestinal conditions, the SFD and FD cells show up to 4?h better tolerance than SD samples and unencapsulated cells in acidic and pepsin condition. The morphology of SFD samples shows particles almost in spherical shape with numerous fine pores, which in turn results in good rehydration behaviour of the powdered product.     

Microencapsulation of a probiotic bacteria with alginate–gelatin and its properties

Lactobacillus casei ATCC 393-loaded microcapsules based on alginate and gelatin had been prepared by extrusion method and the product could increase the cell numbers of L. casei ATCC 393 to be 10⁷ CFU g^?1 in the dry state of microcapsules. The microparticles homogeneously distributed with size of 1.1 ± 0.2 mm. Four kinds of microcapsules (S₁, S₂, S₃ and S₄) exhibited swelling in simulated gastric fluid (SGF) while the beads eroded and disintegrated rapidly in simulated intestinal fluid (SIF). Cells of L. casei ATCC 393 could be continuously released from the microcapsules during simulated gastrointestinal tract (GIT) and the release amounts and speeds in SIF were much higher and faster than that in SGF. Encapsulation in alginate–gelatin microcapsules successfully improved the survival of L. casei ATCC 393 and this approach might be useful in delivery of probiotic cultures as a functional food.     

Soy extract and maltodextrin as microencapsulating agents for Lactobacillus acidophilus: a model approach

Abstract

The present study aimed to optimise the microencapsulation of Lactobacillus acidophilus La-05 by spray drying, using soy extract and maltodextrin as encapsulants. Air inlet temperature, maltodextrin/soy extract ratio and feed flow rate were investigated through Central Composite Rotational Design (CCRD). Probiotic viability increased with increasing the proportion of soy extract. Temperature and feed flow rate had a negative effect. Particle diameter ranged from 4.97 to 8.82?μm, water activity from 0.25 to 0.52 and moisture from 2.30 to 7.01?g.100g^?1 Particles produced following the optimised conditions (air temperature of 87?°C, maltodextrin/soy extract ratio of 2:3 w.w^?1, feed flow rate of 0.54?L.h^?1) reached Encapsulation yield (EY) of 83%. Thermogravimetry and FTIR analysis suggested that microcapsules could protect L. acidophilus cells against dehydration and heating. During storage, microencapsulated probiotic had high cell viability (reductions ranged between 0.12 and 1.72 log cycles). Soy extract/maltodextrin presented well-encapsulating properties of Lactobacillus acidophilus La-05.     

Stability of microencapsulated B. lactis (BI 01) and L. acidophilus (LAC 4) by complex coacervation followed by spray drying

Microcapsules containing Bifidobacterium lactis (BI 01) and Lactobacillus acidophilus (LAC 4) were produced by complex coacervation using a casein/pectin complex as the wall material, followed by spray drying. The aim of this study was to evaluate the resistance of these microorganisms when submitted to the spray drying process, a shelf-life of 120 days at 7–37°C and the in vitro tolerance after being submitted to acid pH (pH 1.0 and 3.0) solutions besides morphology of microcapsules. Microencapsulated microorganisms were shown to be more resistant to acid conditions than free ones. Microencapsulated L. acidophilus maintained its viability for a longer storage period at both temperatures. The microcapsules presented a spherical shape with no fissures. The process used and the wall material were efficient in protecting the microorganisms under study against the spray drying process and simulated gastric juice; however, microencapsulated B. lactis lost its viability before the end of the storage time.     

Effect of natural polymers on the survival of Lactobacillus casei encapsulated in alginate microspheres

Linseed and okra mucilages, the fungal exopolysaccharide botryosphaeran, and commercial fructo-oligosaccharides (FOS) were used to microencapsulate Lactobacillus casei LC-01 and L. casei BGP 93 in sodium alginate microspheres by the extrusion technique in calcium chloride. The addition of carbohydrate biopolymers from linseed, okra and the fungal exocellular (1 → 3)(1 → 6)-β-D-glucan, named botryosphaeran provided higher encapsulation efficiency (EE) (>93% and >86%) for L. casei LC 01 and L. casei BGP 93, respectively. The use of linseed, okra and botryosphaeran improved the stability of probiotics encapsulated in the microspheres during the storage period over 15 d at 5?°C when compared to microspheres formulated with sodium alginate alone as the main encapsulating agent (p?≤?0.05). In in vitro gastrointestinal simulation tests, the use of FOS combined with linseed mucilage was shown to be more effective in protecting L. casei cells LC-01 and L. casei BGP 93.     

Improved viability of Lactobacillus acidophilus NRRL-B 4495 during freeze-drying in whey protein-pullulan microcapsules

Abstract

In this research, pullulan was incorporated in protein-based encapsulation matrix in order to assess its cryoprotective effect on the viability of freeze-dried (FD) probiotic Lactobacillus acidophilus NRRL-B 4495. This study demonstrated that pullulan in encapsulation matrix resulted in a 90.4% survival rate as compared to 88.1% for whey protein (WPI) encapsulated cells. The protective effects of pullulan on the survival of FD-encapsulated cells in gastrointestinal conditions were compared. FD WPI-pullulan capsules retained higher survived cell numbers (7.10?log CFU/g) than those of FD WPI capsules (6.03?log CFU/g) after simulated gastric juice exposure. Additionally, use of pullulan resulted in an increased viability after bile exposure. FD-free bacteria exhibited 2.18?log CFU/g reduction, while FD WPI and FD WPI-pullulan encapsulated bacteria showed 0.95 and 0.49?log CFU/g reduction after 24?h exposure to bile solution, respectively. Morphology of the FD microcapsules was visualized by scanning electron microscopy.     

Acacia-Gelatin Microencapsulated Liposomes: Preparation,Stability, and Release of Acetylsalicylic Acid

Liposomes of dipalmitoylphosphatidylcholine (DPPC) containing acetylsalicylic acid (ASA) have been microencapsulated by acacia-gelatin using the complex coacervation technique as a potential oral drug delivery system. The encapsulation efficiency of ASA was unaltered by the microencapsulation process. The stability of the microencapsulated liposomes in sodium cholate solutions at pH 5.6 was much greater than the corresponding liposomes. The optimum composition and conditions for stability and ASA release were 3.0% acacia-gelatin and a 1- to 2-hr formaldehyde hardening time. Approximately 25% ASA was released in the first 6 hr from microencapsulated liposomes at 23°C and the kinetics followed matrix-controlled release (Q t ^l/2). At 37°C, this increased to 75% released in 30 min followed by a slow constant release, likely due to lowering of the phase transition temperature of DPPC by the acacia-gelatin to near 37°C. At both temperatures, the release from control liposomes was even more rapid. Hardening times of 4 hr and an acacia-gelatin concentration of 5% resulted in a lower stability of liposomes and a faster release of ASA. It is concluded that under appropriate conditions the microencapsulation of liposomes by acacia-gelatin may increase their potential as an oral drug delivery system.     

Bio-detoxification of aflatoxin B1 in artificially contaminated maize grains using lactic acid bacteria

Aflatoxins are a group of carcinogenic mycotoxins produced by Aspergillus flavus, A. parasiticus, and A. nomius. Due to the ubiquitous occurrence of aflatoxins, preventive and remediative measures are necessary including detoxification techniques. Physical and chemical decontamination strategies are inconvenient. In this study a biological detoxification strategy was tested using bacteria of the Lactobacillus species collected from the biotechnology laboratory at University of Ibadan, Nigeria. Maize grains with moisture content of 17% were artificially inoculated with toxigenic A. flavus (LA 32G_28) and atoxigenic A. flavus (LA32_79) at ambient temperature and four samples of bulk maize grains were prepared at aflatoxin B1 concentrations of 50, 100, 200, and 500?ng/g. To evaluate the detoxifying potential of lactic acid bacteria five different cultures consisting of Lactobacillus acidophilus, L. brevis, L. casei, L. delbruekii, and L. plantarum were used to inoculate the aflatoxin B1–contaminated maize samples at 37°C. After 5 days, the residual aflatoxin B1 on maize was determined. All treatments showed significant reductions (P<?0.05) in aflatoxin B1. Lactic acid bacteria decreased the pH of the medium from 5.0 to 4.0. Pronounced aflatoxin B1 reduction was observed in maize contaminated at 50?ng/g (44.5%), while maize contaminated at 500ng/g was the least reduced (29.9%). L. plantarum was the most efficient organism in degrading aflatoxin B1. Use of lactic acid bacteria, which already has Generally Regarded As Safe (GRAS) status, should be encouraged for use as a bio-detoxification agent for aflatoxins.     

Screening of Lactobacillus casei strains for their ability to bind aflatoxin B1

It has been proposed that the consumption of lactic acid bacteria capable of binding or degrading foodborne carcinogens would reduce human exposure to these deleterious compounds. In the present study, the ability of eight strains of Lactobacillus casei to bind aflatoxin B₁ in aqueous solution was investigated. Additionally, the effect of addition of bile salts to the growth medium on aflatoxin B₁ binding was assessed. The eight strains tested were obtained from different ecological niches (cheese, corn silage, human feces, fermented beverage). The strains exhibited different degrees of aflatoxin binding; the strain with the highest AFB₁ binding was L. casei L30, which bound 49.2% of the available aflatoxin (4.6 μg/mL). In general, the human isolates bound the most aflatoxin B₁ and the cheese isolates the least. Stability of the bacterial–aflatoxin complex was assessed by repeated washings. Binding was to a limited degree (0.6–9.2% release) reversible; the L. casei 7R1–aflatoxin B₁ complex exhibited the greatest stability. L. casei L30, a human isolate, was the strain least sensitive to the inhibitory effects of bile salts. Exposure of the bacterial cells to bile significant increased aflatoxin B₁ binding and the differences between the strains was reduced.     

Application of whey protein micro-bead coatings for enhanced strength and probiotic protection during fruit juice storage and gastric incubation

Context: Coated whey protein micro-beads may improve probiotic protection and provide delayed cell-release mechanisms.

Objective: Lactobacillus rhamnosus GG was encapsulated in whey protein micro-beads by droplet extrusion with coating via electrostatic deposition: primary-polysaccharide and secondary-whey protein.

Materials and methods: Storage studies were performed in cranberry and pomegranate juice (pH 2.4; 28 days; 4 and 25°C) followed by simulated ex vivo porcine gastric (pH 1.6) and intestinal (pH 6.6) digestion.

Results and discussion: After storage and simulated gastro-intestinal digestion, free cells, cells suspended in protein and cells encapsulated in alginate micro-beads, illustrated complete probiotic mortality, while coated micro-beads enhanced probiotic viability after juice storage (8.6?±?0.1 log₁₀CFUmL^?1). Beads also showed significant binding of hydrophobic molecules. Coated micro-beads illustrated high gastric survival (9.5?±?0.1 log₁₀CFUmL^?1) with 30?min delayed intestinal release relative to non-coated micro-beads.

Conclusions: Micro-bead coatings could be applied in delayed cell-release for targeted intestinal probiotic delivery.     

Effect of microencapsulation of Lactobacillus salivarus 29 into alginate/chitosan/alginate microcapsules on viability and cytokine induction

Harsh gastric condition causes low bioavailability of probiotics when supplied orally. Polymeric encapsulation has successfully protected bacteria from harsh gastric condition and ultimately increased persistency and multiplication at the targeted region. In this study, we encapsulated LS29 into ACA microcapsules and characterized them. The survivability and release of LS29 from LS29-loaded ACA microcapsules in SGF and SIF were studied. Encapsulation efficiency of LS29 in ACA microcapsules was 99.9%. Approximately 70% of bacteria survived at pH 2 by 120?min after encapsulation. Although not much difference of the survivability of LS29 encapsulated into ACA and FDACA was observed, freeze-drying improved the controlled-release of LS29 in SIF and also showed better storage survivability at 4°C for 8 weeks. Furthermore, investigation of in vitro production of cytokines in RAW264.7 showed high level of induction of TNF-α and IL-10. These in vitro results support that the LS29 might have a balanced immunomodulatory effect.     

Microencapsulation of Lactobacillus plantarum (mtcc 5422) by spray-freeze-drying method and evaluation of survival in simulated gastrointestinal conditions

Spray-drying (SD) and freeze-drying (FD) are widely used methods for microencapsulation of heat-sensitive materials like probiotics for long-term preservation and transport. Spray-freeze-drying (SFD) is relatively a new technique that involves spraying a solution into a cold medium and removal of solvent (water) by conventional vacuum FD method. In this study, the SFD microencapsulated Lactobacillus plantarum powder (1:1 and 1:1.5 core-to-wall ratios of whey protein) is compared with the microencapsulated powders produced by FD and SD methods. The SFD and FD processed microencapsulated powder show 20% higher cell viability than the SD samples. In simulated gastrointestinal conditions, the SFD and FD cells show up to 4?h better tolerance than SD samples and unencapsulated cells in acidic and pepsin condition. The morphology of SFD samples shows particles almost in spherical shape with numerous fine pores, which in turn results in good rehydration behaviour of the powdered product.     

Improving functional properties of pea protein isolate for microencapsulation of flaxseed oil

Unhydrolysed pea protein (UN) forms very viscous emulsions when used at higher concentrations. To overcome this, UN was hydrolysed using enzymes alcalase, flavourzyme, neutrase, alcalase–flavourzyme, and neutrase–flavourzyme at 50?°C for 0?min, 30?min, 60?min, and 120?min to form hydrolysed proteins A, F, N, AF, and NF, respectively. All hydrolysed proteins had lower apparent viscosity and higher solubility than UN. Foaming capacity of A was the highest, followed by NF, N, and AF. Hydrolysed proteins N60, A60, NF60, and AF60 were prepared by hydrolysing UN for 60?min and used further for microencapsulation. At 20% oil loading (on a total solid basis), the encapsulated powder N60 had the highest microencapsulation efficiency (ME?=?56.2). A decrease in ME occurred as oil loading increased to 40%. To improve the ME of N60, >90%, UN and maltodextrin were added. Flowability and particle size distribution of microencapsulated powders with >90% microencapsulation efficiency and morphology of all powders were investigated. This study identified a new way to improve pea protein functionality in emulsions, as well as a new application of hydrolysed pea protein as wall material for microencapsulation.     

Enhanced Bacterial Tumor Delivery by Modulating the EPR Effect and Therapeutic Potential of Lactobacillus casei

Bacteria of micrometer size could accumulate in tumor based on enhanced permeability and retention (EPR) effect. We report here Lactobacillus casei (L. casei), a nonpathogenic facultatively anaerobic bacterium, preferentially accumulated in tumor tissues after intravenously (i.v.) injection; at 24 h, live bacteria were found more in the tumor, whereas the bacteria in normal tissues including the liver and spleen were cleared rapidly. The tumor‐selective accumulation and growth of L. casei is probably due to the EPR effect and the hypoxic tumor environment. Moreover, the bacterial tumor delivery was significantly increased by a nitric oxide (NO) donor nitroglycerin (NG, 10–70 times) and an angiotensin II converting enzyme inhibitor, enalapril (6–18 times). Consequently significant suppression of tumor growth was found in a colon cancer C26 model, and more remarkable antitumor effect was achieved when L. casei was combined with NG, probably by modulating the host nonspecific immune responses; tumor necrosis factor‐α significantly increased in tumor after the treatment, as well as NO synthase activity and myleoperoxidase activity. These findings suggest the potential of L. casei as a candidate for targeted bacterial antitumor therapy, especially in combine with NG or other vascular mediators. © 2014 Wiley Periodicals, Inc. and the American Pharmacists Association J Pharm Sci 103:3235–3243, 2014     

Stability of microencapsulated B. lactis (BI 01) and L. acidophilus (LAC 4) by complex coacervation followed by spray drying

Microcapsules containing Bifidobacterium lactis (BI 01) and Lactobacillus acidophilus (LAC 4) were produced by complex coacervation using a casein/pectin complex as the wall material, followed by spray drying. The aim of this study was to evaluate the resistance of these microorganisms when submitted to the spray drying process, a shelf-life of 120 days at 7-37 degrees C and the in vitro tolerance after being submitted to acid pH (pH 1.0 and 3.0) solutions besides morphology of microcapsules. Microencapsulated microorganisms were shown to be more resistant to acid conditions than free ones. Microencapsulated L. acidophilus maintained its viability for a longer storage period at both temperatures. The microcapsules presented a spherical shape with no fissures. The process used and the wall material were efficient in protecting the microorganisms under study against the spray drying process and simulated gastric juice; however, microencapsulated B. lactis lost its viability before the end of the storage time.     

Microencapsulation of Theobroma cacao L. waste extract: optimization using response surface methodology

The cocoa extract (Theobroma cacao L.) has a significant amount of polyphenols (TP) with potent antioxidant activity (AA). This study aims to optimise microencapsulation of the extract of cocoa waste using chitosan and maltodextrin. Microencapsulation tests were performed according to a Box–Behnken factorial design, and the results were evaluated by response surface methodology with temperature, maltodextrin concentration (MD) and extract flowrate (EF) as independent variables, and the fraction of encapsulated TP, TP encapsulation yield, AA, yield of drying and solubility index as responses. The optimum conditions were: inlet temperature of 170?°C, MD of 5% and EF of 2.5?mL/min. HPLC analysis identified epicatechin as the major component of both the extract and microparticles. TP release was faster at pH 3.5 than in water. These results as a whole suggest that microencapsulation was successful and the final product can be used as a nutrient source for aquatic animal feed.

• Highlights

• Microencapsulation is optimised according to a factorial design of the Box–Behnken type.

• Epicatechin is the major component of both the extract and microcapsules.

• The release of polyphenols from microcapsules is faster at pH 3.5 than in water.

     

Oral Immunization Against Candidiasis Using Lactobacillus casei Displaying Enolase 1 from Candida albicans

Candidiasis is a common fungal infection that is prevalent in immunocompromised individuals. In this study, an oral vaccine against Candida albicans was developed by using the molecular display approach. Enolase 1 protein (Eno1p) of C. albicans was expressed on the Lactobacillus casei cell surface by using poly-gamma-glutamic acid synthetase complex A from Bacillus subtilis as an anchoring protein. The Eno1p-displaying L. casei cells were used to immunize mice, which were later challenged with a lethal dose of C. albicans. The data indicated that the vaccine elicited a strong IgG response and increased the survival rate of the vaccinated mice. Furthermore, L. casei acted as a potent adjuvant and induced high antibody titers that were comparable to those induced by strong adjuvants such as the cholera toxin. Overall, the molecular display method can be used to rapidly develop vaccines that can be conveniently administered and require minimal processing.