Table 3. Inhibitory effect of metformin on the isolated mitochondrial complex 1 in permeabilized KB cells.

Permeabilization of KB cells was achieved after intact cells had been previously preincubated in the absence or presence of 10 mM or 100 μM metformin. Digitonin was added to intact cells in a medium containing 250 mM sucrose, 40 mM KCl, 2 mM EGTA and 20 mM Tris, pH 7.2 (see the Materials and methods section). Complex 1 activity was measured after hypo-osmotic shock-induced mitochondrial membrane rupture, and NADH oxidation was then monitored in the simultaneous presence of 150 μM NADH and 100 μM decylubiquinone before and after the addition of 6 μM rotenone. Citrate synthase activity was measured after the lysis of permeabilized KB cells in a buffer (100 mM KH₂PO₄, 2 mM EDTA and 1 mM dithiothreitol, pH 7.3) containing 0.1% Triton X-100. Each enzyme activity and the corresponding ratio of both were calculated and are expressed as means±S.E.M. (n=4 independent experiments, each performed in duplicate). *P<0.05; **P<0.01 versus control.

		Metformin
	Control	10 mM for 30 min	100 μM for 24 h
Rotenone-sensitive activity of complex 1 [nmol of NADH·min−1·(mg of protein)−1]	3.8±0.5	2.5±0.5**	2.9±0.5*
Citrate synthase activity [pmol·min−1·(mg of protein)−1]	42.6±8.5	41.7±8	40.4±11.3
Enzyme activities ratio (nmol of NADH/unit of citrate synthase)	90.3±7.4	59.9±4.1**	73.2±7.5*