Abstract
Metformin, an anti-diabetes drug, has been recently emerging as a potential "anti-aging" intervention based on its reported beneficial actions against aging in preclinical studies. Nonetheless, very few metformin studies using mice have determined metformin concentrations and many effects of metformin have been observed in preclinical studies using doses/concentrations that were not relevant to therapeutic levels in human. We developed a liquid chromatography-tandem mass spectrometry protocol for metformin measurement in plasma, liver, brain, kidney, and muscle of mice. Young adult male and female C57BL/6 mice were voluntarily treated with metformin of 4 mg/ml in drinking water which translated to the maximum dose of 2.5 g/day in humans. A clinically relevant steady-state plasma metformin concentrations were achieved at 7 and 30 days after treatment in male and female mice. Metformin concentrations were slightly higher in muscle than in plasma, while, ~3 and 6-fold higher in the liver and kidney than in plasma, respectively. Low metformin concentration was found in the brain at ~20% of the plasma level. Furthermore, gender difference in steady-state metformin bio-distribution was observed. Our study established steady-state metformin levels in plasma, liver, muscle, kidney, and brain of normoglycemic mice treated with a clinically relevant dose, providing insight into future metformin preclinical studies for potential clinical translation.
Highlights
Metformin is the first-line drug for the treatment of type 2 diabetes mellitus with a favorable risk/benefit profile
All metformin-treated tissues and blank tissues for preparation of standard curve and quality control samples were homogenized in 1:15 Milli-Q water using GLH-01 homogenizer and Sonic Ruptor 250 ultrasonic homogenizer (OMNI international the homogenizer company, Kennesaw, GA)
We adopted two ion pairs in monitoring metformin and used metformin-D6 to improve the precision of the analytical method
Summary
Metformin is the first-line drug for the treatment of type 2 diabetes mellitus with a favorable risk/benefit profile. Many effects of metformin have been observed in mouse studies using doses that were not relevant to therapeutic levels in human. Metformin is widely distributed into different organs, including intestine, liver, skeletal muscle, and brain, and excreted unchanged mainly through kidney [12]. Bio-distribution of metformin has been studied in rodents using 11C-metformin PET imaging in which metformin was found to accumulate in intestine, kidney and liver at much higher concentrations than in plasma after single intravenous administration [14, 15]. We treated C57BL/6J normoglycemic mice with metformin in a clinically relevant paradigm and metformin bio-distribution in the plasma, liver, kidney, muscle, and brain was determined by a LC-MS/MS methods established and verified in the laboratory
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