Prebiotic functions and mechanisms of natural polysaccharides on different bacterial species

Abstract

Natural polysaccharides (PS) from various sources have been increasingly recognized as potential pharmaceutical and nutraceutical materials due to their notable bioactivities and health benefits. The natural PS with complex structures and high molecular weights (MWs) usually have a strong resistance to digestion in the human gastrointestinal system and can reach the large intestine. In other words, these PS may perform prebiotic functions on the gut bacteria, resulting in various health benefits to the human host. Therefore, the various health benefits of natural and bioactive PS may be connected to their prebiotic functions. The aim of this project was to investigate the potential prebiotic functions of three natural PS from fungal and plant sources in pure cultures of probiotic bacteria. In the first part of this study, the bifidogenic effects of an exopolysaccharide (EPS) from a medicinal fungus (Cordyceps sinensis) and two well-known food PS, konjac glucomannan (KGM) and arabinoxylan (AX), with different MW ranges were evaluated in liquid cultures of Bifidobacterium. The preliminary results showed that the native EPS and KGM could not be well utilized as a carbon source by the bifidobacteria for growth due probably to their high MW and complex structures. Therefore, the native EPS and KGM were partially degraded with high intensity ultrasound (US) to increase the water solubility and lower the viscosity, and to much lower MW by acid hydrolysis with trifluoroacetic acid (TFA). Only the acid-degraded fractions (EPS-AH and KGM-AH) were able to significantly (p < 0.05) increase the growth of all five bifidobacterial species compared to the control without any carbon source, but the effects were much less than that of glucose or galacto-oligosaccharide (GOS). The US-degraded high MW fractions, EPS-US and KGM-US, could slightly support the growth of some bacterial species. All EPS fractions increased the acetic acid production of most bacterial species. Very interestingly, the high MW EPS-US and KGM-US fractions significantly enhanced the cell viability, giving rise to much higher CFU counts than the cultures with glucose or the prebiotic reference GOS as the carbon source, indicating a strong protective effect for the survival of bifidobacteria. These results suggested that EPS and KGM could be used as prebiotic fibres for a healthy gut microbiota. Further study was carried out to evaluate the protective effects of high MW EPS fractions on the probiotic bacteria including Lactobacillus and Bifidobacterium. The EPS at 5 g/L significantly increased the survival rate of the probiotic bacteria during cold storage (4 °C) and in simulated gastric acid, reducing the death rate of different bacterial species by 50% to 70%. The protective effect of EPS was weaker when the concentration was decreased to 3 g/L or when the MW of EPS was reduced by partial degradation with power US. EPS also showed significant protective effect on the four bacterial species in bile juice. Compared with EPS, two commercial prebiotic fibers including inulin and GOS showed much less or no significant protective effect on probiotic bacteria in these conditions. In addition, the EPS had a total dietary fiber content about 70%, which was close to its total carbohydrate content; it was resistant to artificial gastric acid (pH2) with no more than 4% hydrolysis in 6 hours. The results have demonstrated the potential value of Cs-HK1 EPS as a novel prebiotic fiber for the formulation of synbiotic products with probiotic bacteria. Arabinoxylan (AX) is an important dietary fiber which is abundant in many cereal grains such as rye, wheat, barley, oats and rice. In addition to its multiple health benefits, AX has shown prebiotic function, stimulating the growth of beneficial bacteria such as Bifidobacterium in the gut. However, the mechanism for utilization of AX by the bifidobacteria is still unclear due to the substrate specificity of AX hydrolases and the complex structures of native AX as well as the different metabolism processes in different bifidobacterial species. In the preliminary experiments, it was interesting to find that AX could only be consumed effectively by B.longum (CICC6186) but barely used by another four species of bifidobacteria. Further study was performed to investigate the enzymatic metabolism of wheat AX by B.longum. Based on the enzyme activities and consumption rates of AX and its low MW hydrolysates, a strategy was proposed involving extracellular cleavage of xylose backbone and intracellular degradation of both backbone and arabinose substitution. Another interesting finding was that the B.longum had a preference on the AX with a single substitution and a relatively higher degree of polymerization. This suggested that the polymer chain structure of AX played a key role in the uptake and metabolism of AX by the bifidobacterial cell. Genomic analysis showed the lack of the well-known β-xylosidase in the bifidobacterial strain, suggesting the existence of a novel or unknown enzyme or metabolic pathway. According to these results, a hypothetical model was proposed to describe the pathway for uptake and metabolism of AX by the B.longum cell. In summary, following are the chief findings from this project: (1) In pure cultures of bifidobacteria, the high MW EPS produced by the Cs-HK1 fungus or the high MW KGM could not be well utilized by the bacteria as a carbon source for growth, even after acid hydrolysis to much lower MW fractions. (2) The high MW PS especially the EPS produced by the Cs-HK1 fungus could provide protective effects on bifidobacteria and lactobacilli in harsh conditions. (3) Arabinoxylan (AX), a well-known dietary fiber from plant foods, could be selectively utilized by certain bifidobacterial species for growth, e.g. B.longum in this study, and the utilization and bacterial growth was dependant strongly on the AX chain structure. These findings are new and useful for understanding the probiotic functions of natural PS and the relationships to their structure and properties and for development and application of natural PS as prebiotics in food and pharmaceutical industry.