The Establishment And Application Of Methods For Toxicity Detection Of Human Fecal Water

There are a huge amount of bacterial cells which co-exchange and co-metabolize with the host in human gut. Studies suggest that the gut microbiota can produce compounds which are cytotoxic and genotoxic to intestinal epithelium cells. Fecal water (FW) directly reflects the diet composition and the metabolite of the host and gut microbiota, and indirectly reflects the status of gut microbial community. Studies indicate that fecal water has many biological activities, such as cytotoxicity and genotoxicity. Thus, fecal water can reflect the toxic compounds caused by the diet composition and the gut microbiota.Using Caco-2 cells as the model, Comet assay and MTT assay were performed to detect the genotoxicity and cytotoxicity of human fecal water to intestinal epithelial cells, respectively. In order to establish the best reaction system for Comet assay, series experiments were designed, results showed the best Caco-2 cell concentration was 5×10~5cell/ml, the best lysis time was 60 min, and the best fecal water concertation was 80%. The MTT assay was also refined by establishing the best Caco-2 cell concentration.Using the methods for toxicity detection of human fecal water established, the effects of the gut microbiota targeted nutrition intervention on 80 obesity volunteers'fecal water toxicity were studied. Previous work showed that this intervention can modulate the gut microbiota structure of the obesity, and significantly reduce their weight and improve their metabolic syndrome indications. Our results showed that 9w nutrition intervention can significantly reduce the FW cytoxicity (p<0.05) and genotoxicity (p<0.05) of the obesity; the cytotoxicity data was associated with 454 bar-coded pyrosequencing data of gut microbiota 16S rRNA genes, and 13 OTUs, which had significant changes after 9w intervention, were correlated to the FW cytotoxicity.Then the effects of oral administration of probiotic (LAB) product on healthy people'fecal water toxicity were studied. DGGE Profile showed that the LAB stain can get to human gut after drinking the probiotic product for 3 weeks, but it didn't change the gut microbiota structure. However, the probiotic intervention can significantly reduce the FW cytoxicity of the volunteers (p<0.05), and the FW genotoxicity also decresed, but with no significance.In summary, in this paper the methods for genotoxicity (Comet assay) and cytotoxicity (MTT assay) detection of human fecal water were established; Our nutrition intervention can significantly reduce FW toxicity of the obesity; Oral administration of probiotic product can reduce the FW cytotoxicity of the volunteers. The methods can effectively reflect the FW toxicity, thus they can act as a new biologic marker, and be used to evaluate the dietary intervention or drug therapy.