Effect of cryoprotectants,prebiotics and microencapsulation on survival of probiotic organisms in yoghurt and freeze-dried yoghurt

The survival of probiotic microorganisms including Lactobacillus acidophilus, Lactobacillus casei, Lactobacillus rhamnosus and Bifidobacterium spp. was evaluated in yoghurt and freeze-dried yoghurt after processing and storage. The effectiveness of microencapsulating probiotic organisms as well as adding cryoprotectants and prebiotics in improving their viability was also investigated. The viability of Bifidobacterium infantis 17930 and L. rhamnosus GG was reduced by 0.07 log, while that of L. casei 1520 and Bifidobacterium longum 1941 was reduced by 0.28 and 0.39 log, respectively. There was a 7% improvement in the viability of L. casei 1520 when cryoprotectant ‘Unipectine™ RS 150’ was added at 2.5% (w/v). The prebiotic ‘Raftilose^®P95’ when added at 1.5% w/v to yoghurt improved the viability of the combined selected probiotic organisms by 1.42 log during four weeks of storage at 4 °C. Microencapsulation with alginate improved viability of combined selected probiotic organisms by 0.31 log in freeze-dried yoghurt stored at 21 °C.     

Incorporation of selected nutraceuticals and probiotic bacteria into a fermented milk

Yoghurts were produced from a base milk containing three important nutraceuticals, namely ω-3-fatty acids, isoflavones and phytosterols. The cultures employed to make the yoghurts were single probiotic strains of Lactobacillus gasseri or Bifidobacterium infantis and, to achieve a short production time, a two-stage fermentation procedure was used with Streptococcus thermophilus and Lactobacillus delbrueckii subsp. bulgaricus providing the rapid acidification. Yoghurts containing counts of >1.0×10⁸ cfu mL⁻¹ of the individual probiotics and high counts of the traditional species from yoghurt were awarded overall scores for sensory acceptability >4.0 out of 5.0; the nutraceuticals appeared to have no adverse effect on flavour. Storage trials at 5 °C showed that the viability of the probiotic cultures was retained over 15 days.     

Survival and activity of selected probiotic organisms in set-type yoghurt during cold storage

The growth and metabolism of two probiotic organisms (L. acidophilus LAFTI^® L10 and Lactobacillus casei LAFTI^® L26) and a regular yoghurt culture (L. delbrueckii ssp. bulgaricus Lb1466 and Streptococcus thermophilus St1342) were studied in yoghurt containing 0.5%, 1.0%, and 1.5% (w/v) of high amylose corn starch powder (Hi-maize^®) or inulin. Viable cell counts of probiotic organisms, their metabolites and proteolytic activities, and viscosity of the yoghurts were determined during refrigerated storage for 28 d at 4 ^oC. In the presence of inulin, cultures showed better retention of viability (8.0 log cfu g⁻¹) in comparison with that of Hi-maize, which had a reduction by one log cycle. Lower concentrations of 0.5–1.0% Hi-maize improved (P<0.05) the production of propionic acid and also increased proteolytic activity of probiotic organisms substantially. A greater release of free amino acids may have sustained better growth of the organisms in yoghurts. Supplementation with either Hi-maize or inulin increased the viscosity of probiotic yoghurts significantly (P<0.05).     

Properties of low-fat stirred yoghurts made from high-pressure-processed skim milk

Physical properties of stirred yoghurt made from reconstituted skim milk that was high-pressure (HP)-treated at 100, 250 or 400 MPa, at 25, 70 or 90 °C, for 10 min, prior to inoculation with yoghurt cultures, were studied; portions of milk HP-treated at 25 °C were also heat-treated at 90 °C for 10 min before or after pressure treatment. Control yoghurts were made from skim milk given a heat treatment at 90 °C for 10 min. Fermentation time was not affected by treatment applied to the milk. HP treatment of skim milk at 25 °C, before or after heat treatment, gave stirred yoghurts of similar viscosities to that made from conventionally heat-treated milk. Lower viscosities were obtained when stirred yoghurts were made with milk HP-treated at elevated temperatures. A model is proposed to correlate properties of yoghurt with HP/heat-induced changes in interactions and structures of protein in the milk samples.Industrial relevanceTo meet end user expectations, the dairy industry needs to diversify its product range by tailoring specific functionalities. To meet these expectations, new processing methods such as high-pressure processing are of interest for their potential to achieve specific and/or novel functionalities and/or improve efficiencies, including reduced chemical and water use. In this paper, an investigation of the use simultaneous pressurization and heating of milk before the manufacture of stirred yoghurt is presented.     

Flavour profiles and survival of starter cultures of yoghurt produced from high-pressure homogenized milk

Volatile carbonyl compounds, organic acids and yoghurt bacteria counts were investigated in yoghurts made from ultra-high pressure homogenized milk. Yoghurts were manufactured from milk treated using ultra-high pressure homogenization at 200 or 300 MPa and at 30 °C or 40 °C, and compared with those produced from heat-treated milk with 3% added skim milk powder. To study the evolution of these parameters, samples were analysed after days 1, 14 and 28 of storage. Yoghurts from milk heat-treated or treated at 300 MPa had very similar profiles of organic acids and volatile compounds, as well as similar bacterial counts of both starter cultures. In comparison, yoghurts from milk treated at 200 MPa at either 30 °C or 40 °C gave different profiles, together with a sharp decrease in counts of lactobacilli. During storage, only slight differences in flavour compounds and yoghurt bacteria counts were detected, except in those samples from milk treated at 200 MPa.     

Characteristics of stirred low-fat yoghurt as affected by high pressure

The effects of high pressure on the physicochemical, chemical, microbiological and sensory characteristics of stirred low-fat yoghurt were studied. Laboratory-made yoghurts were treated at high pressure (100–400 MPa) for 15 min at 20°C. No significant changes in pH and total organic acids were observed after pressuring the yoghurt. Pressures over 200 MPa prevented post-acidification of the yoghurt during chilled storage. Pressurized yoghurts exhibited higher viscosity and amino acid contents than did the untreated controls, and the differences were maintained after chilled storage. High-pressure treatments at 300 and 400 MPa reduced the number of viable cells of lactobacilli to below the legal minimum permitted in many countries. Significant differences in sensory characteristics between untreated and pressurized yoghurts (200–300 MPa) were detected after chilled storage.     

Survival of Lactobacillus casei (LC-1) adhered to prebiotic vegetal fibers

This study evaluated the survival of Lactobacillus casei (LC-1) when adhered to four different dehydrated prebiotic fibers. After vacuum drying in oat bran with 9% β-glucan and green banana flour, LC-1 viability was 79% and 76%, respectively. Scanning electron microscopy images revealed no morphological changes in the cells adhered to these fibers, and the addition of trehalose as a cell protectant had a positive and significant effect on the survival of LC-1. At different storage temperatures, oat bran had the highest stability as well as in simulated gastrointestinal conditions, where LC-1 that adhered to the oat bran had greater viability (7.1 log CFU g^? 1) than in the free form (2.4 log CFU g^? 1). In the sensory evaluation, the probiotic oat bran added to a dairy fruit beverage was well accepted by consumers.Industrial relevanceThe diversification of food products and the growing interest in health life requires from food industry innovations and new products for this. Probiotic cultures into non-dairy products is still an innovation and a challenge to the functional food segment. The development of an oat bran with LC-1 adhered is important for the functional food industries because the probiotic can be viable during dehydration and storage at room temperature. In addition the LC-1 is protected under simulated gastrointestinal conditions without compromising sensory acceptability.     

Sensory and rheological screening of exopolysaccharide producing strains of bacterial yoghurt cultures

Yoghurts were produced with 24 different cultures differing in their ability to produce exopolysaccharides (EPS). Rheological and sensory analyses showed large differences in texture properties in the yoghurt samples. EPS production was found to have a major effect on the texture properties, but varying textures within the EPS±groups were also found. Yoghurts fermented with EPS-producing cultures showed increased mouth thickness and ropiness and tended to be creamier than yoghurts without these cultures; in contrast, these yoghurts had the lowest syneresis and highest gel firmness (initial gel strength before the yoghurt was subjected to shear). Correlations between rheological parameters and sensory texture attributes included G* correlating with gel firmness, while viscosity measured at 241 s⁻¹ correlated with mouth thickness. An interesting viscometry relationship was found between ropiness and hysteresis loop area.     

Rheology,spontaneous whey separation,microstructure and sensorial characteristics of probiotic yoghurts enriched with passion fruit fiber

Beyond demonstrated beneficial health attributes, passion fruit rinds are a by-product of the fruit pulp industry, rich in total dietary fiber, particularly pectin. The aim of this study was to evaluate the influence of the addition of passion fruit fiber on the whey formation, rheological parameters, microstructure and sensorial characteristics of probiotic yoghurts. Skim milk bases enriched with 1% of passion fruit fiber or not were heat treated and inoculated with Streptococcus thermophilus and Lactobacillus delbrueckii subsp. bulgaricus, and divided into four groups according to the probiotic strain added — Lactobacillus acidophilus strains L10 and NCFM and Bifidobacterium animalis subsp. lactis strains Bl04 and B94. Fermentations were performed until the pH reached 4.5. Rheological characteristics of yoghurts were determined by a rotational rheometer in two cycles of shear rate ranging from 0 to 15 s^? 1 in both upward and downward curves. Sensorial analysis of passion fruit fiber yoghurts, either without any probiotic or co-fermented by L. acidophilus L10 or B. animalis subsp. lactis Bl04, was evaluated against a control yoghurt without fiber. Photomicrographs of freeze-dried yoghurts were made by field-emission scanning electron microscope (SEM). Thixotropy of enriched yoghurts was higher than that of their respective controls in the two cycles of shear rate. Apparent viscosity was significantly higher in fiber yoghurts co-fermented by the lactobacilli than in their controls at the end of cold storage. Photomicrographs demonstrated that in passion fruit fiber yoghurts the casein gel was more compact and overlaid the fiber, while filaments of exopolysaccharides were more frequent in control yoghurts. Appearance, odor and color of the passion fruit fiber yoghurts received scores as ‘good’, and the intensity of the passion fruit flavor was considered weak by the sensory assessors. Results indicate that the passion fruit fiber is an almost neutral ingredient for the design of new high value-added yoghurt.     

Rheological,microstructural and sensory characterization of low-fat and whole milk set yoghurt as influenced by inulin addition

The effect of inulin addition (0–4%) upon texture and microstructure of set yoghurt with different levels of fat (0.2%–3.5%) was investigated. A two-factor experimental design with four treatments was used for data analysis. Skimmed milk with various inulin and cream concentrations was standardized to 4% protein content, homogenized, heated to 92 °C and fermented at 42 °C until a pH of 4.6 was reached. The chemical composition, pH, consistency and microstructure properties of the yoghurts were analysed after 6 days of storage at 5 °C. The statistical analysis showed that inulin and fat significantly affected the rheological and sensory results. Higher yield stress, “firmness” and “creaminess” values were observed in yoghurt produced with higher inulin additions, whereas the pH value was not affected. A significant correlation was found between yield stress and sensory determined firmness (r = 0.91). The microstructure examined by confocal laser scanning microscopy (CLSM) was only slightly affected by the concentrations of inulin in the range studied, possibly due to weak protein interactions between the inulin and the milk protein network.     

Modifying the microstructure of low-fat yoghurt by microfluidisation of milk at different pressures to enhance rheological and sensory properties

The effects of microfluidisation of milk at different pressures, prior to heat treatment, on structural and sensory properties of low-fat stirred yoghurt, were investigated. Low-fat yoghurts prepared from microfluidised milk were compared with low-fat (1.5%) and full-fat (3.5%) control yoghurts made with homogenised (20/5 MPa) milk. The microstructure of low-fat yoghurts prepared with microfluidised milk consisted of smaller and more uniform fat globules, well incorporated into more interconnected fat-protein gel networks, compared with those of control yoghurts. This modification in microstructure caused significant changes in gel particle size, sensory profile and rheological behaviour. Microfluidisation increased the gel particle size, gel strength and viscosity; marked beneficial effects were found at higher pressures (50–150 MPa). Microfluidising milk at 50–150 MPa increased the gel strength by 171–195% and viscosity by 98–103%, creating low-fat yoghurts with creaminess and desirable texture properties similar to, or better than, full-fat conventional yoghurt.     

Fermentation of beet juice by beneficial lactic acid bacteria

Red beets were evaluated as a potential substrate for the production of probiotic beet juice by four species of lactic acid bacteria (Lactobacillus acidophilus, Lactobacillus casei, Lactobacillus delbrueckii, Lactobacillus plantarum). All the lactic cultures were found capable of rapidly utilizing beet juice for cell synthesis and lactic acid production. However, L. acidophilus and L. plantarum produced a greater amount of lactic acid than other cultures and reduced the pH of fermented beet juice from an initial value of 6.3 to below 4.5 after 48 h of fermentation at 30°C. Although the lactic cultures in fermented beet juice gradually lost their viability during cold storage, the viable cell counts of these lactic acid bacteria except for L. acidophilus in the fermented beet juice still remained at 10⁶–10⁸ CFU/ml after 4 weeks of cold storage at 4°C.     

Development of probiotic beads similar to fish eggs

Probiotic foods are mainly restricted to dairy and soy products. This study aimed to develop a new probiotic beads similar to fish eggs, commonly used in oriental cuisine. Beads were produced by the extrusion encapsulation technique with calcium alginate, added to one of the following cultures: Lactobacillus rhamnosus GG ATCC 53103 and Bifidobacterium animalis DN-173 010 and stored for 30 days at 4 °C. The beads were characterized by the size, weight, morphology and viability of the probiotic strains in different storage temperatures and in simulated gastric juice adjusted to different pH values. The beads were also evaluated by a sensorial affective hedonic scale. The beads present a 2.8 mm diameter and a weight of 0.01 g (p > 0.05). Free and encapsulated cells were tolerant to pH 3.0. At pH 2.5 only of the encapsulated cells presented counts above 6 Log colony-forming units per gram (CFU/g). Beads containing L. rhamnosus showed higher viability 10⁷ CFU/g in storage for 30 days under refrigeration. The beads may be stored at abusive temperature for 5 h without loss of viability cells. The probiotic product developed showed an 82.2% acceptability index of overall characteristics and good market potential as a new probiotic product.     

Apple,grape or orange juice: Which one offers the best substrate for lactobacilli growth? — A screening study on bacteria viability,superoxide dismutase activity,folates production and hedonic characteristics

Fermentation can contribute to improve functional aspects of foods. The first goal of this study was to determine amongst apple, grape and orange juices, the one with the best bacterial growth performance during fermentation by Lactobacillus strains from commercial and artisanal food origins, at 40 °C for 48 h. The juice with the highest bacterial growth was evaluated for bacteria viability during 4 weeks of cold storage, superoxide dismutase (SOD) activity and folates production analyzed through HPLC/fluorimetry. Acceptability of fermented juice was appraised through hedonic analysis. Lactobacilli counts were the highest in apple and the lowest in orange juices at t = 48 h. In most cases, bacteria counts were higher in fermented (5.5 to 9.5 log CFU/ml) than in supplemented apple juices (4.2 to 5.7 log CFU/ml), at the 4th week of cold storage. SOD activity was significantly increased in all apple juices fermented by commercial Lactobacilli strains. Folates were produced in apple juices fermented by Lactobacillus plantarum and Lactobacillus rhamnosus. Apple juice was the best substrate for Lactobacillus growth and, considering bacterial viability and overall acceptance by the panelists, Lactobacillus acidophilus L10 was the most suitable strain for apple juice fermentation.     

Production of traditional Greek yoghurt using Lactobacillus strains with probiotic potential as starter adjuncts

Lactobacillus plantarum ACA-DC 146 and L. paracasei subsp. tolerans ACA-DC 4037 were examined for their potential application as adjuncts in the production of traditional Greek set-type yoghurt. Both strains displayed low milk acidification activity, while no inhibition was observed towards or from the yoghurt starters used. Yoghurt produced with L. paracasei subsp. tolerans ACA-DC 4037 exhibited the best sensory properties, with a rich traditional smooth taste, and the strain was selected for further trials. Yoghurt produced with this strain as an adjunct had good physicochemical properties. After 2 weeks of refrigerated storage, microbial loads (>7.0 log cfu g⁻¹) were in accordance with international recommendations and guidelines for probiotic and starter cultures in milk products. Increasing the microbial load further, using concentrated and encapsulated inocula (10–11 log cfu g⁻¹), gave yoghurt with long fermentation times and poor organoleptic properties.     

Improving ambient temperature stability of probiotics with stress adaptation and fluidized bed drying

Stabilization efficiency in terms of long term ambient temperature storage viability of the probiotic strain Lactobacillus casei CRL 431 was compared using freeze and fluidized bed drying techniques. Fluidized bed drying was able to retain 2.5 log cfu/g higher viability after 52 weeks of storage at 25 °C. A combination of fluidized bed drying and osmotic stress adaptation to the probiotic cells yielded further improvement of 0.83 log cfu/g higher viability compared to the unstressed cells. The findings were validated with other two lactobacilli and two bifidobacterium strains with probiotic characteristics and significant improvements in storage stability over freeze dried samples were observed. Fortification of vitamin E in the stabilization matrix as an antioxidant improved the stability by 0.18 log cfu/g during 20 weeks storage period at 25 °C, whereas any similar benefit of fortifying inulin as a prebiotic was not observed.     

Microencapsulation of bifidobacteria by spray drying in the presence of prebiotics

This study was conducted to evaluate the viability and the physical properties of Bifidobacterium BB-12 microencapsulated by spray drying with partial replacement of reconstituted skim milk (RSM), as encapsulating agent, with the prebiotics inulin, oligofructose, and oligofructose-enriched inulin (at a ratio of 1:1, 200 g L^?1 total concentrations). The viable cell counts of the microcapsules were determined during storage for 180 days at 4 °C and at ? 18 °C. The physical characterization included analysis of morphology, particle size, moisture content, water activity, dissolution, hygroscopicity, color, and thermal properties. All the microcapsules produced in this study showed a high survival rate of bifidobacteria during storage at the temperatures evaluated. The microcapsules produced with inulin and those produced with oligofructose-enriched inulin showed higher initial counts. However, blending oligofructose-enriched inulin with RSM and blending oligofructose with RSM resulted in better protection of bifidobacteria during storage. All microcapsules showed similar morphologies and particle sizes, between 14.45 and 18.78 μm. The partial replacement with prebiotics decreased moisture content and water activity of the microcapsules. The time of dissolution in water was higher for the microcapsules produced with inulin, while the microcapsules produced with oligofructose were more hygroscopic. The value of a* increased in the microcapsules produced with prebiotics. The results of the thermal analysis suggest a higher stability of the microcapsules produced with prebiotics than those produced with RSM only.     

Physical characteristics of set yoghurt made with altered casein to whey protein ratios and EPS-producing starter cultures at 9 and 14% total solids

This study investigated firmness and syneresis of set yoghurts made at the CN to WP ratios of 4:1, 3:1, 2:1 and 1:1 using non-EPS-, capsular EPS- and ropy EPS-producing starter cultures. The yoghurts were made at 9 and 14% (w/w) total solids. The total solids, total protein, the concentration of lactose and the ratios of CN to WP as well as the protein's profiles (native- polyacrylamide gel electrophoresis) in heated and unheated milk blends were investigated. The level of soluble denatured whey protein aggregates in heated milk was also determined. The concentration of EPS, firmness and level of spontaneous syneresis in set yoghurt were monitored weekly throughout 28 days of storage. The microstructure of the set yoghurt made with milk blends at the CN to WP ratio of 4:1 and using three types of starter cultures was carried out after 1 day of storage. There was no difference in the total solids, total protein and lactose concentration of liquid milk blends, except the CN to WP ratios. There was no difference in whey protein denaturation between milk blends. The level of soluble denatured whey protein aggregates in heated milk blends decreased with reducing CN to WP ratio. The firmness and the level of spontaneous syneresis in set yoghurts decreased as the CN to WP ratios were reduced. The use of EPS-producing starter cultures reduced firmness and syneresis and changed the protein matrix in the microstructure of set yoghurts made at 9% (w/w) total solids compared to the control products. These were not observed in set yoghurts made at 14% (w/w) total solids.     

Kinetic parameters of Escherichia coli O157:H7 survival during fermentation of milk and refrigeration of home-made yoghurt

The survival parameters of Escherichia coli O157:H7 during milk fermentation (carried out by the LIM or “longer incubation method” at 30 °C, or by the SIM or “short incubation method” at 43 °C) and storage of home-made yoghurt at refrigeration temperatures (2, 4, or 8 °C) were studied. The E. coli O157:H7 counts increased slightly during fermentation by the LIM, from 5.1 to 5.4 log cfu mL⁻¹, and it was not found after 21 d of storage at 2 or 4 °C, and after 10 d at 8 °C. The microorganism counts increased from 4.8 to 5.4 log cfu mL⁻¹ during the SIM, and it was not detected after 7 d stored at 8 °C. The microorganism grew faster at 43 °C (generation time=0.93 h) than at 30 °C (4.12 h) during the fermentation period. The death time decreased with the increase of the storage temperature (from 38.1 h at 2 °C to 30.1 h at 8 °C) in the yoghurt produced by fermentation at 30 °C; however, a clear relationship between death time and storage temperature was not evident at 43 °C. The pH values of the yoghurt ranged from 4.0 to 4.7.     

Temperature effect on the biological activity of Bifidobacterium longum CRL 849 and Lactobacillus fermentum CRL 251 in pure and mixed cultures grown in soymilk

The influence of temperature on the growth and biological activity of two probiotic strains (Bifidobacterium longum CRL 849 and Lactobacillus fermentum CRL 251) as pure and mixed cultures in soymilk (SM) were evaluated. Maximum growth was observed at 37°C in both mixed and pure cultures. In a product prepared with the mixed culture (1:1) at 37°C, the amount of lactic acid produced was approximately 55 mmol l⁻¹ after 24 h with a slow production rate (2.8 mmol l⁻¹ h⁻¹); the formation of acetic acid was higher with respect to pure cultures (82.01 mmol l⁻¹ after 24 h), and final pH (24 h) was 5.0. About 85% of the total amount of sugars in SM was reduced, mainly sucrose. Stachyose was reduced (71%) after 4 h of incubation. Maximum activity of alpha-galactosidase (alpha-gal) (13.2 U ml⁻¹) was observed after 6 h. At 37°C the bifidobacterium strain was viable in mixed culture throughout the period assayed. At lower (30°C) or higher (42°C) temperatures, mixed culture showed slower growth and lower acid production in SM but the alpha-gal activity was stimulated at 30°C.