Different yield and properties of mitochondria from skeletal muscle of normal hamsters

Yield and properties of mitochondria, isolated from hind leg muscles of normal hamsters, are different, when 4 preparation procedures are applied simultaneously: 1. mechanical homogenization of muscle with a glass-Teflon homogenizer followed by rehomogenization of the isolated nuclear fraction for 1--6 times. The procedure yields impure mitochondria showing the highest amounts of mitochondrial enzymes (50--65% of total), protein and DNA; the isolated mitochondria exhibit disrupted outer membranes and loosened inner membranes causing a better supply with substrates of enzymes located at the inner membrane and in the matrix. 2. Single mechanical homogenization of muscle for increasing periods (2--40 min) yields lower amounts of all constituents (25--40% of total). 3. The yields of Nagarse treatment with one step of 2 min mechanical homogenization range between 20--35% of total. Mitochondria isolated with the latter procedure exhibit compact membranes and are very pure, but still contaminated with Nagarse, causing destructions of mitochondrial membranes already after 24 h storage in buffer with sucrose and mannitol at 2 degrees C. 4. One single Nagarse treatment of muscle for 40 min produces a minor yield of these these constituents. Therefore, at least two different isolation procedures are proposed for optimum investigation of skeletal muscle mitochondria.