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maximum of MV Chl b , elicited a Chl b emission maximum at 660 nm. Appearance
of Chl b fluorescence in vivo or in organello is usually an indication of a certain
degree of disruption of the structural relationship of Chl a and b in the thylakoid
membranes. Indeed, Chl b in healthy thylakoids, does not fluoresce but transfers its
excitation energy to Chl a .
Further evidence of disorganization of the aforementioned complexes, was
evidenced by the state of the 77 K fluorescence excitation spectra. In three excita-
tion spectra, recorded at F685, 696 and 739 nm, the normal three-banded fluores-
cence excitation profile with maxima at 440, 475 and 485 nm was replaced by one
Soret excitation maximum at 443 nm (Amindari et al. 1995 ). This Soret excitation
maximum corresponds to MV Chl a coordinated to two small ligands such as
pyridine at room temperature. The Soret excitation maximum at 424 nm is that
of membrane bound DV Mpe which transfers its excitation energy to the remnants
of the LHCII, LHCI-680, Chl a CP47 and/or CP29, and LHCI-730 complexes, as
evidenced by the sloping tail between 685 and 740 nm (Amindari et al. 1995 ).
17.4.2.4 Effect of Exogenous DV Pchlide a on Photodynamic
Damage in Isolated Cucumber Chloroplasts
As discussed in Chap. 8 , Divinyl Pchlide a is the precursor of DV Chlide a . It differs
from DV Mpe by the presence of a fifth ring, the cyclopentanone ring, at position
six and
of the macrocycle. In its native state it is bound to the plastid membranes
(Smith and Rebeiz 1979 ). Membrane-bound DV Pchlide a exhibits emission
maxima between 629 and 658 nm, depending on its state of aggregation and the
stage of greening of the tissue (Cohen and Rebeiz 1978 ).
After 2 h of incubation with isolated chloroplasts in the light, 20 % of the added
DV Pchlide a disappeared probably as a consequence of photodestruction (Rebeiz
et al. 1984a ). Added DV Pchlide caused considerable destruction of MV Chl a and b ,
which was accompanied by the formation of significant amounts of chlorophyllide
(Chlide) a and b probably by hydrolysis of the long chain fatty acid at position seven
of the macrocycle of the corresponding Chls. In three replicates, the 77 K fluores-
cence emission and excitation profiles after 2 h of incubation in the absence of added
DV Pchlide a , were indistinguishable from 0 h controls. However after incubation
with 5,493 nmol of DV Pchlide a per 100 mg plastid protein for 2 h in the light, the
77 K fluorescence emission and excitation profiles underwent profound changes.
Essentially the organized structure of the chloroplast was profoundly disrupted.
The 740 nm fluorescence emission maximum decreased considerably in magnitude
thus indicating disruption of the LHCI-730 protein-pigment complex (Amindari
et al. 1995 ). The fluorescence emission maximum at 696 nm disappeared completely,
thus indicating the complete disorganization of the CP47 and/or CP29 complex. The
686 nm fluorescence emission peak underwent a 3 nm blue shift which also indicated
a certain degree of disorganization of the LHCII and LHCI-680 complexes (Amindari
et al. 1995 ).
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