The In Vitro Analysis of Prebiotics to Be Used as a Component of a Synbiotic Preparation

Abstract

Prebiotics are food components that are selectively fermented by beneficial microbiota and which confer a health benefit. The aim of the study was to select a prebiotic for the chosen probiotic strains to create a synbiotic. The impact of prebiotics (inulin, maltodextrin, corn starch, β-glucan, and apple pectin) on five Lactobacillus spp. strains' growth and metabolites synthesis (lactic, acetic, propionic, and butyric acids, ethanol, and acetaldehyde) was tested by the plate count method and by high-performance liquid chromatography, respectively. Moreover, the differences in the ratio of D(-) and L(+) lactate isomers produced by Lactobacillus spp., as well as variations in the probiotics' enzymatic profiles associated with the prebiotic used for cultivation, were determined with a Megazyme rapid assay kit and API^® ZYM assay, accordingly. Finally, the influence of the carbon source (prebiotic) used on the antagonistic activity of the probiotic strains towards pathogenic bacteria, such as Salmonella spp. or Listeria monocytogenes was analyzed in the co-cultures. The results showed that the growth, metabolic profile, and antagonistic activity of the probiotics towards selected pathogens were the most favorable when 2% (w/v) of inulin was used. Therefore, the combination of inulin with selected probiotics is a promising synbiotic mixture.

Keywords: antagonism; inulin; metabolic activity; microbial growth; prebiotics; probiotics; synbiotics.

Figure 1

Metabolites concentration produced by probiotic strains, namely, (a) Lb. rhamnosus ŁOCK 1087, (c) Lb. paracasei ŁOCK 1091, (e) Lb. reuteri ŁOCK 1092, (g) Lb. plantarum ŁOCK 0860, and (i) Lb. pentosus ŁOCK 1094 in dependence on the used prebiotic (MD–maltodextrin, INU–inulin, CS–corn starch, AP–apple pectin, BG–β-glucan) or glucose (GLU) comparatively as a carbon source in the cultivation medium. Moreover, the correlations between used carbohydrates, as well as their concentrations, and the synthesized amount of metabolites, were presented as the PCA biplots, namely, plots (b,d,f,h,j) for above-mentioned strains, respectively.

Figure 1

Metabolites concentration produced by probiotic strains, namely, (a) Lb. rhamnosus ŁOCK 1087, (c) Lb. paracasei ŁOCK 1091, (e) Lb. reuteri ŁOCK 1092, (g) Lb. plantarum ŁOCK 0860, and (i) Lb. pentosus ŁOCK 1094 in dependence on the used prebiotic (MD–maltodextrin, INU–inulin, CS–corn starch, AP–apple pectin, BG–β-glucan) or glucose (GLU) comparatively as a carbon source in the cultivation medium. Moreover, the correlations between used carbohydrates, as well as their concentrations, and the synthesized amount of metabolites, were presented as the PCA biplots, namely, plots (b,d,f,h,j) for above-mentioned strains, respectively.

Figure 2

Heatmap presenting the Lactobacillus spp. strains enzymatic profile in relation to carbohydrates used in the culture: MD–maltodextrin, INU–inulin, CS–corn starch, AP–apple pectin, BG–β-glucan, and GLU–glucose. The tested enzymes were enumerated as follows: 1–alkaline phosphatase, 2-esterase (C4), 3-lipase esterase (C8), 4-lipase (C14), 5-leucine arylamidase, 6-valine arylamidase, 7-cystine arylamidase, 8-trypsin, 9-chymotrypsin, 10-acid phosphatase, 11-naphthol-AS-BI-phosphohydrolase, 12-α-galactosidase, 13-β-galactosidase, 14-β-glucuronidase, 15-α-glucosidase, 16-β-glucosidase, 17-n-acetyl-β-glucosaminidase, 18-α-mannosidase, 19-α-fucosidase.

Figure 3

Antagonistic activity of a mixture of the probiotic strains, namely, Lb. rhamnosus ŁOCK 1087, Lb. paracasei ŁOCK 1091, Lb. reuteri ŁOCK 1092, Lb. plantarum ŁOCK 0860, and Lb. pentosus ŁOCK 1094 on the pathogenic bacteria: (a) L. monocytogenes, (b) S. Enteritidis, (c) S. Typhimurium, and (d) S. Choleraesuis. The co-cultures were conducted in the presence of each carbohydrate (MD–maltodextrtin, INU–inulin, AP–apple pectin, CS–corn starch, BG–β-glucan, or GLU–glucose).