Effects of ginsenoside compound K on colitis-associated colorectal cancer and gut microbiota profiles in mice.

Background: Ginsenoside compound K (GC-K), generated from ginseng saponins bioconverted by gut microbiota, has potential anti-colorectal cancer (CRC) effects. Meanwhile, GC-K may interact with gut microbiota, playing important roles in the occurrence and development of CRC. However, the effects of gut microbiota on the preventive and therapeutic effects of GC-K in CRC remain to be elucidated. Methods: The anti-CRC effects of GC-K were evaluated in an azoxymethane/dextran sulfate sodium (AOM/DSS)-induced colitis-associated CRC Balb/c mice model under the dosage of 30 and 60 mg/kg. Stool samples were collected during the experiments for profiling gut microbiota by 16S rRNA sequencing. Correlative analysis between gut microbiota and anti-CRC effect of GC-K was also assessed. Finally, the anti-CRC effect of Akkermansia muciniphila (A. muciniphila) was validated in CRC cell lines. Results: GC-K could significantly suppress tumor growth in vivo at the dosage of 60 mg/kg without exogenous interference of gut microbiota. Moreover, dysbiosis of gut microbiota was observed in the CRC model group, which could be recovered by GC-K treatment. In particular, A. muciniphila, which could inhibit the proliferation of HCT-116, HT-29, and LOVO cells, was significantly up-regulated by GC-K. Conclusions: GC-K was verified to suppress the tumor growth of AOM/DSS-induced colitis-associated CRC through the modulation of gut microbiota, partially by up-regulation of A. muciniphila.

Bioconversion variation of ginsenoside CK mediated by human gut microbiota from healthy volunteers and colorectal cancer patients.

BACKGROUND: Ginsenoside CK (GCK) serves as the potential anti-colorectal cancer (CRC) protopanaxadiol (PPD)-type saponin, which could be mainly bio-converted to yield PPD by gut microbiota. Meanwhile, the anti-CRC effects of GCK could be altered by gut microbiota due to their different diversity in CRC patients. We aimed to investigate the bioconversion variation of GCK mediated by gut microbiota from CRC patients by comparing with healthy subjects. METHODS: Gut microbiota profiled by 16S rRNA gene sequencing were collected from healthy volunteers and CRC patients. GCK was incubated with gut microbiota in vitro. A LC-MS/MS method was validated to quantify GCK and PPD after incubation at different time points. RESULTS: The bioconversion of GCK in healthy subjects group was much faster than CRC group, as well as the yield of PPD. Moreover, significant differences of PPD concentration between healthy subjects group and CRC group could be observed at 12 h, 48 h and 72 h check points. According to 16S rRNA sequencing, the profiles of gut microbiota derived from healthy volunteers and CRC patients significantly varied, in which 12 differentially abundant taxon were found, such as Bifidobacterium, Roseburia, Bacteroides and Collinsella. Spearman's correlation analysis showed bacteria enriched in healthy subjects group were positively associated with the biotransformation of GCK, while bacteria enriched in CRC group displayed non correlation character. Among them, Roseburia which could secrete ß-glycosidase showed the strongest positive association with the bioconversion of GCK. CONCLUSIONS: The bioconversion of GCK in healthy subjects was much faster than CRC patients mediated by gut microbiota, which might alter the anti-CRC effects of GCK.

In Vivo Metabolic Profiles of Panax notoginseng Saponins Mediated by Gut Microbiota in Rats.

Panax notoginseng saponins (PNSs) are the major health-beneficial components of P. notoginseng with very low oral bioavailability, which could be biotransformed by gut microbiota in vitro. However, in vivo biotransformation of PNS mediated by gut microbiota is not well known. This study aimed to characterize the in vivo metabolic profiles of PNS mediated by gut microbiota. The saponins and yielded metabolites in rat feces were identified and relatively quantified by ultra-performance liquid chromatography tandem/quadrupole time-of-flight mass spectrometry. Seventy-three PNS metabolites had been identified in the normal control group, but only 11 PNS metabolites were determined in the pseudo germ-free (GF) group. In addition, the main biotransformation pathway of PNS metabolism was hydrolytic and dehydration reactions. The results indicated that a significant metabolic difference was observed between the normal control group and pseudo GF group, while gut microbiota played a profound role in the biotransformation of PNS in vivo.

Quantification of Panax notoginseng saponins metabolites in rat plasma with in vivo gut microbiota-mediated biotransformation by HPLC-MS/MS.

Panax notoginseng saponins (PNS) are the major components of Panax notoginseng, with multiple pharmacological activities but poor oral bioavailability. PNS could be metabolized by gut microbiota in vitro, while the exact role of gut microbiota of PNS metabolism in vivo remains poorly understood. In this study, pseudo germ-free rat models were constructed by using broad-spectrum antibiotics to validate the gut microbiota-mediated transformation of PNS in vivo. Moreover, a high performance liquid chromatography-electrospray ionization tandem mass spectrometry (HPLC-ESI-MS/MS) was developed for quantitative analysis of four metabolites of PNS, including ginsenoside F1 (GF1), ginsenoside Rh2 (GRh2), ginsenoside compound K (GCK) and protopanaxatriol (PPT). The results showed that the four metabolites could be detected in the control rat plasma, while they could not be determined in pseudo germ-free rat plasma. The results implied that PNS could not be biotransformed effectively when gut microbiota was disrupted. In conclusion, gut microbiota plays an important role in biotransformation of PNS into metabolites in vivo.

[Design and validation of primers for housekeeping genes of streptomycetes].

Multigene-based phylogenetic analyses are becoming more widely used in molecular taxonomy because of its lab-to-lab portability and reproducibility. The first step that needs to be settled for this approach is to amplify and sequence several housekeeping genes. Four housekeeping genes atpD, recA, rpoB and trpB were chosen for streptomycetes, which are representatives of high G + C mol % Gram-positive bacteria. Primer pairs for amplification and sequencing of the gene fragments were designed according to the known genome sequences of two streptomycetes and three mycobacteria, as well as the available recA gene sequences of another two streptomycetes in NCBI database, by using software packages Primer premier 5.0, Oligo 6.0 and SPCR 3.0, and NCBI BLAST program. Performance of the primer sets were validated by specific amplification of all gene fragments from the 55 streptomycete tested strains under optimized PCR reaction conditions, and by successful sequencing of the amplification products. It is concluded that the primer sets designed here are effective, and the primer design procedure and guidelines are valuable for other housekeeping genes of high G + C mol % bacteria.