Pharmacokinetic studies
Mice were randomly caged in groups of 3 of each sex where each data
point (dose or time) was derived from 6 mice. Stocks of sildenafil and
malonyl-sildenafil were prepared in diH20 to deliver
different doses by gavage of 0.1 ml per mouse, or 9 mg/kg for the time
course. Blood was drawn from mice by submandibular bleed (Golde et
al. , 2005), and plasma was isolated using BD microtainer lithium tubes
(Becton, Dickinson and Company, Franklin Lakes, NJ). Fecal pellets were
collected over a 2 h period and the PDE5i were extracted by mixing with
1 ml methanol for 3 seconds in a Pro-200 homogenizer (Cole-Parmer,
Vernon Hills, IL), followed by centrifugation at 10,000 x g for 10 min.