Berberine derivatives as cationic fluorescent probes for the investigation of the energized state of mitochondria.

The interaction of rat liver and bovine heart mitochondria with a series of fluorescent, cationic berberine derivatives varying in the length of alkyl chain has been investigated. An increase in the hydrophobicity of the derivative was accompanied by a larger value of the partition coefficient and by binding to a more hydrophobic region of the inner mitochondrial membrane. It was found that berberines could be used as sensitive indicators of processes which take place on the outer surface of the mitochondrial membrane; the greatest (15-fold) increase in fluorescence was obtained with 13-methylberberine in the energized state of mitochondria. The fluorescence increase was due to the increase in fluorescence quantum yield although a small increase in the amount of bound derivative could also be detected upon energization. The fluorescence was linearly dependent on the magnitude of the membrane potential. In parallel with an observed fluorescence enhancement a considerable decrease in rotational mobility was found. We suggest that berberines move in the inner membrane according to the polarity of the membrane potential; consequently, deeper immersion in the less polar region in the energized state brings about a larger fluorescence increase. More hydrophobic derivatives inhibited NAD-linked respiration in rat liver mitochondria but exerted no effect on succinate oxidation up to 10 microM concentration.