In vivo inulin fermentation in the bowel and linking SCFA production to microbial species using Stable Isotope Probing
Polymeric plant components may pass undigested to the large bowel where they become carbon substrates for bacteria. Inulin in the diet positively influenced large bowel microbial fermentation (Butts et al., 2016). The large bowel accommodates numerous species occupying a wide range of ecological niches where they practice their often specific functions. Identification of the bowel bacteria involved in fermentation processes will lead to a better insight how inulin promotes short chain fatty acids (SCFA’s) production. SCFA’s improve the intestinal barrier function, reduce inflammation processes, and improve metabolism in obese persons (Beek et al., 2018, Deroover et al., 2017).
Stable Isotope Solution: 13C-labelled biomarkers
One of the problems when investigating the effects of prebiotics like inulin is how to follow the dynamics of its metabolites e.g. in the blood plasma and to distinguish these from other internal or external sources. The use of uniformly labelled 13C-inulin enables tracing of metabolites and linking SCFA production to bacterial species, either in vivo in the bowel or in vitro systems like TIM-2 (TNO gastro-Intestinal Models).
Results from publications with IsoLife’s U-13C Inulin
Deroover et al. (2017) demonstrated in healthy volunteers that intake of 13C-inulin resulted in increased SCFA-concentration in plasma for at least 8 hours. These findings were confirmed by Beek et al., 2018). Knowledge about the transformation of inulin into SCFA’s is important, e.g. for understanding how SCFA’s improves fat oxidation in obese men (Beek et al., 2018) and how one of these fatty acids, acetate, plays an important in the central appetite regulation (Frost et al., 2014).
The active microbial species involved in the fermentation process of inulin can be identified by extracting and analyzing 13C-RNA from faeces. This method, called RNA-stable isotope probing (RNA-SIP), has been successfully used in vitro with 13C-labelled potato starch (Kovatcheva et al. 2009) and in vivo in the study by Tannock et al. (2014). During the latter study, rats were fed with a diet containing IsoLife‘s uniformly labelled 13C-Inulin. Several hours after administration, RNA extracted from cecal samples and centrifuged over a density gradient to separate 12C-RNA from 13C-enriched RNA. Most of the RNA obtained was present in the heavier fractions, indicating an active population of 13C–Inulin consumers. It was shown that Bacteroides uniformis was able to directly use inulin as a carbon source (trophic level 1), whereas Blautia glucerasea, Clostridium indolis, and Bifidobacterium animalis fermented the fructo-oligosaccharides derived from inulin degradation (trophic level 2) (Figure 1).
Figure 1. Abundances of 16S 501 rRNA gene sequences from bacterial groups (Tannock et al, 2014).
Beek, van der CM, EE Canfora, AM Kip, SHM Gorissen, SWM Olde Damink, HM van Eijk, JJ Holst, EE Blaak, CHC Dejong, K Lenaerts. 2018.
The prebiotic inulin improves substrate metabolism and promotes short-chain fatty acid production in overweight to obese men.
Metabolism Clinical and Experimental 87: 25-35.
Butts CA, G Paturi, MH Tavendale, D Hedderley, H Stoklosinski, T Herath, D Rosendale, N Roy, JA Monro, J Ansell. 2016.
The fate of 13C-labelled and non-labelled inulin predisposed to large bowel fermentation in rats.
Food and Function 7: 1825-1832.
Deroover L, J Verspreet, A Luypaerts, G Vandermeulen, CM Courtin, K Verbeke. 2017.
Wheat bran does not affect postprandial plasma short-chain fatty acids from 13C-inulin fermentation in healthy subjects.
Nutrients 9: 83.
Tannock GW, B Lawley, K Munro, IM. Sims, J Lee, CA. Butts, N Roy. 2014.
RNA-stable isotope probing (RNA-SIP) shows carbon utilization from inulin by 2 specific bacterial populations in the large bowel of rats.
Applied Environmental Microbiology doi:10.1128/AEM.03799-13.
Frost G, ML Sleeth, M Sahuri-Arisoylu, B Lizarbe, S Cerdan, L Brody, J Anastasovska, S Ghourab, M Hankir, S Zhang, D Carling, JR Swann, G Gibson, A Viardot, D Morrison, EL Thomas, JD Bell. 2014.
The short-chain fatty acid acetate reduces appetite via a central homeostatic mechanism.
Nature Communications 5. DOI: 10.1038/ncomms4611.
Kovatcheva-Datchary P, M Egbert, A Maathuis, M Rajilic-Stojanovic, AA de Graaf, H Smidt, WM de Vos, K Venema. 2009.
Linking phylogenetic identities of bacteria to starch fermentation in an in vitro model of the large intestine by RNA-based stable isotope probing.
Environmental Microbiology 11: 914-926.