Optimization of cholesterol removal, growth and fermentation patterns of Lactobacillus acidophilus ATCC 4962 in the presence of mannitol, fructo-oligosaccharide and inulin: a response surface methodology approach

AIMS: To optimize cholesterol removal by Lactobacillus acidophilus ATCC 4962 in the presence of prebiotics, and study the growth and fermentation patterns of the prebiotics. METHODS AND RESULTS: Lactobacillus acidophilus ATCC 4962 was screened in the presence of six prebiotics, namely sorbitol, mannitol, maltodextrin, hi-amylose maize, fructo-oligosaccharide (FOS) and inulin in order to determine the best combination for highest level of cholesterol removal. The first-order model showed that the combination of inoculum size, mannitol, FOS and inulin was best for removal of cholesterol. The second-order polynomial regression model estimated the optimum condition of the factors for cholesterol removal by L. acidophilus ATCC 4962 to be 2.64% w/v inoculum size, 4.13% w/v mannitol, 3.29% w/v FOS and 5.81% w/v inulin. Analyses of growth, mean doubling time and short-chain fatty acid (SCFA) production using quadratic models indicated that cholesterol removal and the production of SCFA were growth associated. CONCLUSIONS: Optimum cholesterol removal was obtained from the fermentation of L. acidophilus ATCC 4962 in the presence of mannitol, FOS and inulin. Cholesterol removal and the production of SCFA appeared to be growth associated and highly influenced by the prebiotics. SIGNIFICANCE AND IMPACT OF THE STUDY: Response surface methodology proved reliable in developing the model, optimizing factors and analysing interaction effects. The results provide better understanding on the interactions between probiotic and prebiotics for the removal of cholesterol.     

Development of Bifidobacterium lactis Bb 12 on β‐(2,6)‐Linked Fructan‐Containing Substrate

β‐(2,1)‐linked fructan of plant origin (inulin) and the related oligosaccharides (FOS) as non‐digestible carbohydrates, i.e., potent prebiotics, can stimulate the growth of various probiotic lactic acid bacteria, including a number of bifidobacteria strains. The related β‐(2,6)‐linked fructans of microbial origin (levan and FOS), however, have scarcely been investigated in this respect. Therefore, the bifidogenic properties of various fructans, i.e., inulin, levan, fructooligosaccharides (FOS) and fructan syrup (FS), were tested as glucose substitutes in MRS media and were compared concerning their effect on the commercial strain Bifidobacterium lactis Bb 12. Although glucose was the preferred substrate for growth and biomass formation, FS exhibited a comparable cell growth (8.4 × 10⁷ counts/mL and 1.0 × 10⁷ counts/mL, respectively) and acidification power (84 °T and 74 °T, respectively) during 48 h of fermentation, as well as an increase in lactic acid and decrease in acetic acid formation. Bifidobacterium lactis Bb 12 did not utilize inulin as a sole carbon source as judged from the 60 % decrease in cell count and the insignificant (0.1 pH unit) acidification of the growth medium, whereas levan provided a noticeable increase in cell count and acidification (0.4 pH units) during 48 h of fermentation. FOS preparation appeared to be a satisfactory carbon source for this strain, but lower acidification power (56 °T) and cell counts were observed as compared to glucose‐ or FOS‐containing media (2.6 % and 22 %, respectively). The products obtained under conditions of mild lactic acid hydrolysis of levan (37 °C, pH 3.3, 24 h) enhanced the cell count (7–10 %) and acidification power (by a factor of 2.7) of Bifidobacterium lactis Bb 12.     

A comparative in vitro evaluation of the fermentation properties of prebiotic oligosaccharides

AIMS: Comparison of in vitro fermentation properties of commercial prebiotic oligosaccharides. METHODS AND RESULTS: Populations of predominant gut bacterial groups were monitored over 24 h of batch culture through fluorescent in-situ hybridization. Short-chain fatty acid and gas production were also measured. All prebiotics increased the numbers of bifidobacteria and most decreased clostridia. Xylo-oligosaccharides and lactulose produced the highest increases in numbers of bifidobacteria whilst fructo-oligosaccharides produced the highest populations of lactobacilli. Galacto-oligosaccharides (GOS) resulted in the largest decreases in numbers of clostridia. Short-chain fatty acid generation was highest on lactulose and GOS. Gas production was lowest on isomalto-oligosaccharides and highest on inulin. CONCLUSIONS: The oligosaccharides differed in their fermentation characteristics. Isomalto-oligosaccharides and GOS were effective at increasing numbers of bifidobacteria and lactate whilst generating the least gas. SIGNIFICANCE AND IMPACT OF THE STUDY: The study provides comparative data on the properties of commercial prebiotics, allowing targeting of dietary intervention for particular applications and blending of oligosaccharides to enhance overall functionality.     

Optimization of simultaneously enzymatic fructo- and inulo-oligosaccharide production using co-substrates of sucrose and inulin from Jerusalem artichoke

Prebiotic substances are extracted from various plant materials or enzymatic hydrolysis of different substrates. The production of fructo-oligosaccharide (FOS) and inulo-oligosaccharide (IOS) was performed by applying two substrates, sucrose and inulin; oligosaccharide yields were maximized using central composite design to evaluate the parameters influencing oligosaccharide production. Inulin from Jerusalem artichoke (5–15% w/v), sucrose (50–70% w/v), and inulinase from Aspergillus niger (2–7 U/g) were used as variable parameters for optimization. Based on our results, the application of sucrose and inulin as co-substrates for oligosaccharide production through inulinase hydrolysis and synthesis is viable in comparative to a method using a single substrate. Maximum yields (674.82?mg/g substrate) were obtained with 5.95% of inulin, 59.87% of sucrose, and 5.68 U/g of inulinase, with an incubation period of 9?hr. The use of sucrose and inulin as co-substrates in the reaction simultaneously produced FOS and IOS from sucrose and inulin. Total conversion yield was approximately 67%. Our results support the high value-added production of oligosaccharides using Jerusalem artichoke, which is generally used as a substrate in prebiotics and/or bioethanol production.     

Effects of the prebiotics inulin and lactulose on intestinal immunology and hematology of preruminant calves

Prebiotics are suggested as an alternative to antibiotics in animal rearing. Fermentable substances such as inulin or lactulose have been proposed to stimulate the immune system and health by modulation of the intestinal flora and its fermentation products. In this study, effects of inulin and lactulose on the intestinal health and hematology of calves have been investigated. Both prebiotics significantly decreased thrombocyte counts in peripheral blood. Only inulin was able to increase hemoglobin concentration and hematocrit. Total leukocyte count was decreased by lactulose while both prebiotics tended to lower monocyte proportions. mRNA expression of inflammation-related markers in the intestine was also affected by both prebiotics hinting at a decreased inflammatory status. This may be due to a possible decrease in intestinal pathogen load that remains to be verified. Only mRNA amounts of interleukin 8 were increased by lactulose in mesenteric lymph nodes. In the ileum, expression of a proliferation marker was increased by inulin while an apoptosis-related gene was increased by both prebiotics. The results of this study show a clear effect of prebiotics on certain parameters associated with animal health and performance that remain to be studied in detail in future investigations.     

几种益生元制剂对肠道菌群作用效果的研究

目的探讨几种益生元制剂对肠道菌群作用效果。方法通过体外实验和体内试验。结果低聚果糖、水苏糖、低聚木糖、低聚异麦芽糖均能促进双歧杆菌和乳杆菌的增殖,并能够酸化肠道的pH;能够显著提高肠道的B／E值（肠道内双歧杆菌和肠杆菌数量log值的比值）,增加肠道中有益菌的比例,有益于稳定肠道的微生态平衡。结论这几种益生元制剂对肠道菌群有较好的调节作用。     

Prebiotic effectiveness of inulin extracted from edible burdock

To investigate the prebiotic potential of burdock inulin (B-INU), the in vitro and in vivo effects of B-INU on bacterial growth were studied. B-INU significantly stimulated the growth of bifidobacteria in Man-Rogosa-Sharp (MRS) medium, anaerobically. Compared with chicory inulin (C-INU), long-chain inulin (L-INU) and fructooligosaccharides (FOS), 1% (w/v) B-INU promoted the specific growth rate of beneficial bacteria. The decreases of media pH with B-INU were almost the same as that with C-INU and FOS. In vivo, B-INU significantly increased the number of lactobacilli and bifidobacteria (P<0.05) in cecal content. Mice fed with B-INU, C-INU and FOS for 14 days had greater number of cecal beneficial bacteria population than those fed with L-INU for 14 days. In addition, all fructans did not cause any side effects, such as eructation and bloating. Results indicated that inulin extracted from edible burdock showed prebiotic properties that could promote health.     

Selection and optimization procedure of synbiotic for cholesterol removal

A selection and optimization procedure for the synbiotic combination of probiotic and prebiotics was established to optimize its cholesterol removal in vitro. In light of fermentability, prebiotics utilization by probiotics was highly variable and interspecies differences existed. Based on the results of fermentability, L. plantarum LS12, Ls31, LP529 and L. ruminis La3 could be the better candidates for symbiotic research. The bile tolerance of all the tested strains could be improved by the strain-specific prebiotics comparing to the control carbon source (glucose). The strain LS12 was finally selected to form the symbiotic according to its better ability to ferment prebiotics and bile tolerance, while the five prebiotics (FOS, stachyose, GOS, IMO and mannitol) were selected to make their synbiotic combination because of their better enhancement of bile tolerance and growth support to LS12. The synbiotic combination for cholesterol removal was optimized by use of response surface methodology. The first-order model showed that the selected prebiotics mannitol and GOS were significant factors. Then through the second-order polynomial regression model, the optimum conditions of the two factors for cholesterol removal by the synbiotic were suggested.     

Investigation on the precaecal and faecal digestibility of lactulose and inulin and their influence on nutrient digestibility and microbial characteristics

This study was conducted to determine the pre-caecal and faecal digestibility of lactulose and inulin and the influence of these substances on nutrient digestibility and microbial characteristics. In metabolic trials three of six male growing pigs (German Landrace x Pietrain) were fitted with an ileo-rectal anastomosis (IRA) in end-to-end technique with preserved ileo-caeco-colic valve. The metabolic trials were conducted from day 21-63 after surgery. The remaining pigs were used as intact partners (IN) for the IRA pigs. The experimental diets, based on corn, wheat, barley and soybean meal, were supplemented with either 1.5% lactulose or 2% inulin in replacement of diatomaceous earth (control). Pre-caecal digestibility of lactulose and inulin was assessed to be 79 and 98%, respectively. faecal digestibility was determined as 100%. The supplementation of lactulose and inulin had only minor effects on the pre-caecal and faecal digestibility of nutrients. Significant differences in nutrient digestibility were obvious between IRA and IN pigs, whereas the IRA pigs showed lower digestibility values with the exception of ether extracts (EE). Bacterial population in the digesta of IRA and IN pigs were not affected by the experimental diets except the concentration of gram-negative anaerobes, which inclined when the IRA pigs received the lactulose diet. The pH of chyme was significantly lower than the pH of faeces, however the pH was unaffected by the different supplemented diets. The concentration of volatile fatty acids (VFA) in pre-caecal chyme decreased significantly when IRA pigs received the lactulose supplemented diet whereas VFA in faeces were unaffected by the supplementation. IRA pigs administered with lactulose excreted more N via the urine, but the nitrogen balance remained unaffected. From the present investigation it can be concluded that lactulose and inulin did only partly or scarcely fulfill the expectation of acting as prebiotics in pigs.     

Investigation on the precaecal and faecal digestibility of lactulose and inulin and their influence on nutrient digestibility and microbial characteristics

This study was conducted to determine the pre-caecal and faecal digestibility of lactulose and inulin and the influence of these substances on nutrient digestibility and microbial characteristics. In metabolic trials three of six male growing pigs (German Landrace?×?Pietrain) were fitted with an ileo-rectal anastomosis (IRA) in end-to-end technique with preserved ileo-caeco-colic valve. The metabolic trials were conducted from day 21?–?63 after surgery. The remaining pigs were used as intact partners (IN) for the IRA pigs. The experimental diets, based on corn, wheat, barley and soybean meal, were supplemented with either 1.5% lactulose or 2% inulin in replacement of diatomaceous earth (control). Pre-caecal digestibility of lactulose and inulin was assessed to be 79 and 98%, respectively, faecal digestibility was determined as 100%. The supplementation of lactulose and inulin had only minor effects on the pre-caecal and faecal digestibility of nutrients. Significant differences in nutrient digestibility were obvious between IRA and IN pigs, whereas the IRA pigs showed lower digestibility values with the exception of ether extracts (EE). Bacterial population in the digesta of IRA and IN pigs were not affected by the experimental diets except the concentration of gram-negative anaerobes, which inclined when the IRA pigs received the lactulose diet. The pH of chyme was significantly lower than the pH of faeces, however the pH was unaffected by the different supplemented diets. The concentration of volatile fatty acids (VFA) in pre-caecal chyme decreased significantly when IRA pigs received the lactulose supplemented diet whereas VFA in faeces were unaffected by the supplementation. IRA pigs administered with lactulose excreted more N via the urine, but the nitrogen balance remained unaffected. From the present investigation it can be concluded that lactulose and inulin did only partly or scarcely fulfill the expectation of acting as prebiotics in pigs.     

Dietary fructooligosaccharides alter the cultivable faecal population of rats but do not stimulate the growth of intestinal bifidobacteria

The effect of fructans on the cultivable faecal community of Bio Breeding rats fed diets containing 5% (m/v) food-grade fructooligosaccharide (FOS) was investigated. Culturing of faecal material using chicory inulin as the sole carbohydrate source revealed the presence of a greater diversity of inulin-utilizing bacterial species in FOS-fed rats as compared with the control rats, although both contained species which effectively utilized inulin. The majority of cultivable inulin-utilizing species fell within the Clostridium coccoides group and Clostridium leptum subgroup, some of which were related to previously cultured butyrate-producing bacteria from the intestines of various animals. The impact of FOS on the growth of the indigenous bifidobacteria community and three inulin-utilizing isolates was assessed using real-time polymerase chain reaction. While dietary FOS was found to stimulate the growth of all three inulin-utilizing isolates, no growth stimulation of the indigenous bifidobacteria community occurred over the duration of the feeding trial.     

Effects of inulin and lactulose on the intestinal morphology of calves

For some time now prebiotics have been proposed to improve health by stimulation of beneficial bacteria in the intestine of humans and animals. The current study is aiming to show effects of feeding of either 2% inulin or 2% lactulose in milk replacer on performance and intestinal morphology of male Holstein-Friesian calves. After 20 weeks of feeding inulin led to significantly higher daily weight gains than lactulose while control animals ranged between the experimental feedings. Ingestion of milk replacer was reduced in lactulose treated animals. Additionally differences of villus height in jejunum (P = 0.07) and ileum (P = 0.03) could be found with an increase for lactulose treated animals and a decrease for inulin treated animals. In ileum the density of proliferative epithelial cells tended to be lower in inulin treated and higher in lactulose treated animals (P = 0.08). Both inulin and lactulose tended to decrease the quantity of goblet cells in the tips of ileal villi (P = 0.07). Both prebiotics can affect performance and intestinal morphology of calves and may as such affect animal health. But effects differ between substances.     

Bacterial metabolism and health-related effects of galacto-oligosaccharides and other prebiotics

Most studies involving prebiotic oligosaccharides have been carried out using inulin and its fructo-oligosaccharide (FOS) derivatives, together with various forms of galacto-oligosaccharides (GOS). Although many intestinal bacteria are able to grow on these carbohydrates, most investigations have demonstrated that the growth of bifidobacteria, and to a lesser degree lactobacilli, is particularly favoured. Because of their safety, stability, organoleptic properties, resistance to digestion in the upper bowel and fermentability in the colon, as well as their abilities to promote the growth of beneficial bacteria in the gut, these prebiotics are being increasingly incorporated into the Western diet. Inulin-derived oligosaccharides and GOS are mildly laxative, but can result in flatulence and osmotic diarrhoea if taken in large amounts. However, their effects on large bowel habit are relatively minor. Although the literature dealing with the health significance of prebiotics is not as extensive as that concerning probiotics, considerable evidence has accrued showing that consumption of GOS and FOS can have significant health benefits, particularly in relation to their putative anti-cancer properties, influence on mineral absorption, lipid metabolism, and anti-inflammatory and other immune effects such as atopic disease. In many instances, prebiotics seem to be more effective when used as part of a synbiotic combination.     

Prebiotics enhance survival and prolong the retention period of specific probiotic inocula in an in vivo murine model

AIM: To identify novel prebiotics that could be used to maintain persistence of three representative probiotic strains in vivo. METHODS AND RESULTS: Test mice were treated with prebiotics soybean oligosaccharide (SOS), fructooligosaccharide (FOS) or inulin, followed by probiotics Lactobacillus acidophilus LAFTI L10 (L10), Bifidobacterium lactis LAFTI B94 (B94) or Lactobacillus casei L26 LAFTI (L26). Faecal samples were then collected and analysed using selective medium and PCR analysis to determine the presence of the probiotic strains. In contrast to the control groups, in mice fed prebiotics, the survival and retention time of the test probiotics was increased extensively. SOS and FOS prolonged the retention period of L10 from 24 to 30 h. Of the three prebiotics, FOS gave the best result with B94, prolonging the retention period from 3 to > or =10 days. Of the three prebiotics, inulin gave the best result for L26, prolonging the retention period from 2 to > or =6 days. CONCLUSIONS: The prebiotics SOS, FOS and inulin significantly enhance survival and prolong the retention period of L10, B94 and L26 in vivo. SIGNIFICANCE AND IMPACT OF THE STUDY: Our results demonstrate the potential use of FOS, inulin and SOS as prebiotics in conjunction with the probiotic strains L10, B94 and L26 for new synbiotic products.     

Fermentation of Fructooligosaccharides and Inulin by Bifidobacteria: a Comparative Study of Pure and Fecal Cultures

The utilization of fructooligosaccharides (FOS) and inulin by 55 Bifidobacterium strains was investigated. Whereas FOS were fermented by most strains, only eight grew when inulin was used as the carbon source. Residual carbohydrates were analyzed by high-performance anion-exchange chromatography with pulsed amperometric detection after batch fermentation. A strain-dependent capability to degrade fructans of different lengths was observed. During batch fermentation on inulin, the short fructans disappeared first, and then the longer ones were gradually consumed. However, growth occurred through a single uninterrupted exponential phase without exhibiting polyauxic behavior in relation to the chain length. Cellular β-fructofuranosidases were found in all of the 21 Bifidobacterium strains tested. Four strains were tested for extracellular hydrolytic activity against fructans, and only the two strains which ferment inulin showed this activity. Batch cultures inoculated with human fecal slurries confirmed the bifidogenic effect of both FOS and inulin and indicated that other intestinal microbial groups also grow on these carbon sources. We observed that bifidobacteria grew by cross-feeding on mono- and oligosaccharides produced by primary inulin intestinal degraders, as evidenced by the high hydrolytic activity of fecal supernatants. FOS and inulin greatly affected the production of short-chain fatty acids in fecal cultures; butyrate was the major fermentation product on inulin, whereas mostly acetate and lactate were produced on FOS.     

Synbiotics growth optimization of Bifidobacterium pseudocatenulatum G4 with prebiotics using a statistical methodology

Aims:  This study demonstrated the optimum growth of Bifidobacterium pseudocatenulatum G4 with prebiotics via statistical model.
Methods and Results:  Commercial prebiotics [inulin and fructooligosaccharide (FOS)], together with sorbitol, arabinan and inoculum rate, were tested by fractional factorial design to determine their impact on growth of Bif. pseudocatenulatum G4 in skim milk. At 48 h incubation, bacterial growth was mainly influenced by FOS and inoculum rate. Growth reduction was observed in all samples incubated for 72 h. Central composite design (CCD) was adopted using FOS and inoculum rate at 48 h incubation to develop the statistical model for optimization. The model predicted that 2·461 log CFU ml⁻¹ produced the optimum growth increase of Bif. pseudocatenulatum G4. The combination that produced the optimum point was 2·86% FOS (g/v) and 0·67% inoculum rate (v/v).
Conclusion:  At optimum combination of inoculum rate and FOS, validation experiments recorded 2·40 ± 10·02 log CFU ml⁻¹. The application in 1-l bioreactor for 24 h showed higher growth increase of 2·95 log CFU ml⁻¹.
Significant and Impact of the Study:  Response surface methodology approach is useful to develop optimum synbiotics combination for strain G4 with FOS.     

In Vitro and In Vivo Evaluation of the Prebiotics GroBiotic®-A, Inulin, Mannanoligosaccharide, and Galactooligosaccharide on the Digestive Microbiota and Performance of Hybrid Striped Bass (Morone chrysops × Morone saxatilis)

Two separate experiments were conducted with hybrid striped bass to evaluate four potential prebiotics: GroBiotic®-A (partially autolyzed brewer’s yeast, dairy ingredient components, and fermentation products), mannanoligosaccharide (MOS), galactooligosaccharide (GOS), and inulin. In the in vitro experiment, intestinal contents were incubated with the individual prebiotics (0.5% by weight) at 25°C for 24 and 48 h. Analysis of volatile fatty acids in the supernatant showed that GroBiotic®-A, MOS, and GOS tended to produce lower acetate levels but higher butyrate levels at 48 h compared to diet alone. However, denaturing gradient gel electrophoresis (DGGE) analysis failed to detect any differences in the composition of the microbial community among treatments. DNA sequencing of a common band for all inoculated samples revealed close similarity to the anaerobic Fusobacteria bacterium. An 8-week feeding trial also was conducted to evaluate the four prebiotics looking at growth performance; weight gain, feed efficiency ratio, protein efficiency ratio, whole-body ash, moisture, and lipid did not vary among fish fed the various diets. However, DGGE analysis revealed that all prebiotics produced a different type of microbial community in the intestinal tract of hybrid striped bass compared to fish fed the basal diet. Thus, GroBiotic®-A, FOS, GOS, and MOS exhibited prebiotic effects in hybrid striped bass.     

Lactulose production by β-galactosidase in permeabilized cells of<Emphasis Type="Italic"> Kluyveromyces lactis</Emphasis>

Lactulose production from lactose and fructose was investigated with several commercial -galactosidases. The enzyme from Kluyveromyces lactis exhibited the highest lactulose productivity among the -galactosidases tested. The reaction conditions for lactulose production were optimized using cells that had been permeabilized by treatment with 50% (v/v) ethanol: cell concentration, 10.4 g l^–1; concentration of substrates, 40% (w/v) lactose and 20% (w/v) fructose; temperature, 60^°C; pH 7.0. Under these conditions, the permeabilized cells produced approximately 20 g l^–1 lactulose in 3 h with a lactulose productivity of 6.8 g l^–1 h^–1. These results represent 1.3- and 2.1-fold increases in lactulose concentration and productivity compared with untreated washed cells. This is the first reported trial of enzymatic synthesis of lactulose using permeabilized yeast cells.     

In vitro study of prebiotic properties of levan-type exopolysaccharides from Lactobacilli and non-digestible carbohydrates using denaturing gradient gel electrophoresis

Batch cultures inoculated with human faeces were used to study the prebiotic properties of levan-type exopolysaccharides (EPS) from Lactobacillus sanfranciscensis as well as levan, inulin, and fructooligosaccharide (FOS). Denaturing gradient gel electrophoresis of 16S rDNA fragments generated by PCR with universal primers was used to analyse the cultures. Characteristic changes were revealed in the composition of the gut bacteria during fermentation of the carbohydrates. An enrichment of Bifidobacterium spp. was found for the EPS and inulin but not for levan and FOS. The bifidogenic effect of the EPS was confirmed by culturing on selective medium. In addition, the use of EPS and FOS resulted in enhanced growth of Eubacterium biforme and Clostridium perfringens, respectively.     

Prebiotic Oligosaccharides: Comparative Evaluation Using In Vitro Cultures of Infants' Fecal Microbiomes

The objective of this study was to systematically assess the bifidogenic effect of three commonly used prebiotic products using in vitro cultures of infant fecal samples. Fresh stool samples collected from six term infants, each exclusively fed human milk (n = 3) or infant formula (n = 3), at 28 days of age were used as inocula. The following prebiotic products were added at concentrations applicable to infant formula: Vivinal GOS 15 (containing 28.5% galacto-oligosaccharide [GOS]) at 7.2 g/liter, Beneo HP (99.5% long-chain inulin [IN]) at 0.8 g/liter, Beneo Synergy 1 (enriched oligofructose and inulin [OF-IN]) at 4 g/liter, and a combination of Vivinal GOS 15 (7.2 g/liter) and Beneo HP (0.8 g/liter) (GOS-IN). The growth of total bacteria, Bifidobacterium, Bacteroides, Bifidobacterium longum, and Escherichia coli was quantified using specific quantitative PCR (qPCR). Bifidobacterium was also enumerated on selective Beerens agar plates, with representative colonies identified by sequencing of their 16S rRNA genes. Volatile fatty acids (VFA) and pH in the cultures were also determined. Irrespective of the feeding methods, the GOS product, either alone or in combination with Beneo HP, resulted in substantially higher growth of total bifidobacteria, and much of this growth was attributed to growth of B. longum. Beneo Synergy 1 also increased the abundance of total bifidobacteria and B. longum. Corresponding to the increases in these two bacterial groups, acetic acid concentrations were higher, while there was a trend of lower E. coli levels and pH. The lower pH and higher acetic acid concentration might be directly responsible for the lower E. coli population. At the concentrations studied, the GOS product was more bifidogenic and potent in inhibiting E. coli than the other products tested. These results suggest that supplementation of infant formula with GOS may increase intestinal bifidobacteria and benefit infant health.