Higher plant plasma membranes contain a b-type cytochrome that is rapi
dly reduced by ascorbic acid. The affinity towards ascorbate is 0.37 m
M and is very similar to that of the chromaffin granule cytochrome b(5
61). High levels of cytochrome b reduction are reached when ascorbic a
cid is added either on the cytoplasmic or cell wall side of purified p
lasma membrane vesicles. This result points to a transmembrane organis
ation of the heme protein or alternatively indicates the presence of a
n effective ascorbate transport system. Plasma membrane vesicles loade
d by ascorbic acid are capable of reducing extravesicular ferricyanide
. Addition of ascorbate oxidase or washing of the vesicles does not el
iminate this reaction, indicating the involvement of the intravesicula
r electron donor. Absorbance changes of the cytochrome b alpha-band su
ggest the electron transfer is mediated by this redox component. Elect
ron transport to ferricyanide also results in the generation of a memb
rane potential gradient as was demonstrated by using the charge-sensit
ive optical probe oxonol VI. Addition of ascorbate oxidase and ascorba
te to the vesicles loaded with ascorbate results in the oxidation and
subsequent re-reduction of the cytochrome b. It is therefore suggested
that ascorbate free radical (AFR) could potentially act as an electro
n acceptor to the cytochrome-mediated electron transport reaction. A w
orking model on the action of the cytochrome as an electron carrier be
tween cytoplasmic and apoplastic ascorbate is discussed.