Quantitative Metabolite Analysis in Mouse Gut

Samples for LC-MS analysis were prepared as we described previously (19 (link), 20 (link)). Briefly, aliquots of the mucosa of mouse stomach, small intestine, cecum, and colon were homogenized with 50% methanol by Bead Ruptor Homogenizer (Omni International, Kennesaw GA). All sulfated and glucuronide metabolites were measured by enzymatic hydrolysis of the processed samples with β-glucuronidase and sulfatase as described (20 (link), 21 (link)). Then the homogenates were extracted with equal volume of ethyl acetate for 3 times. The ethyl acetate extracts were combined and dried using a vacuum concentrator (Model: SVC 100H, Thermo Fisher Scientific Inc.), and then resuspend in 50% methanol for LC-MS (Model 2020, Shimadzu, Kyoto, Japan) analysis. Quantitation by LC-MS was performed by SIM, using ESI mode as described previously (22 (link)).

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Wu X., Li Z., Sun Y., Li F., Gao Z., Zheng J, & Xiao H. (2020). Identification of Xanthomicrol as a Major Metabolite of 5-Demethyltangeretin in Mouse Gastrointestinal Tract and Its Inhibitory Effects on Colon Cancer Cells. Frontiers in Nutrition, 7, 103.

Publication 2020

Bead Ruptor Homogenizer Model 2020

Cecum Colon Enzymatic Ethyl acetate Glucuronidase Glucuronide Hydrolysis Methanol Mouse Mucosa of stomach Small intestine Sulfatase Vacuum

Corresponding Organization : University of Massachusetts Amherst

Other organizations : Miami University, Anhui Agricultural University, Institute of Food Science and Technology, Chinese Academy of Agricultural Sciences

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Variable analysis

independent variables

Homogenization of aliquots of the mucosa of mouse stomach, small intestine, cecum, and colon with 50% methanol using Bead Ruptor Homogenizer

dependent variables

Levels of sulfated and glucuronide metabolites measured by enzymatic hydrolysis of the processed samples with β-glucuronidase and sulfatase

control variables

Enzymatic hydrolysis of the processed samples with β-glucuronidase and sulfatase as described in previous studies (20, 21)

controls

Positive control: Not explicitly mentioned
Negative control: Not explicitly mentioned

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