Biology Reference
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In ether at 77
K, MV and DV Chl(ide)
a
exhibit similar fluorescence emission
bands with maxima at 673-674 nm (Wu et al.
1989
). Fortunately the Soret excita-
tion bands are very different. While MV Chl(ide)
a
exhibits a Soret excitation
maximum at 446-447 nm, DV Chl(ide)
a
exhibits a Soret excitation maximum
at458-459 nm(Wu et al.
1989
). Furthermore, at these wavelengths, the contribution
of the DV Chl(ide)
a
Soret signal to the MV Chl(ide)
a
Soret signal (about 1/3 at
447 nm) and the contribution of the MV Chl(ide)
a
signal to the DV Chl(ide)
a
signal (about 1/14 at 458 nm) were suitable for efficient correction with simulta-
neous equations. It was therefore considered that the Soret excitation maxima of
MV and DV Chl(ide)
a
at 447 and 458 nm, respectively, should be most suitable for
the best signal deconvolution possible in mixtures of the two compounds.
The choice of appropriate fluorescence wavelengths or the mathematical
Deconvolution of the DV Chl(ide)
b
signal from the MV Chl(ide)
b
signal proved
to be more involved. At 77
K, MV and DV Chl(ide)
b
exhibited emission maxima
at 659 and 666 nm, respectively (Wu et al.
1989
). Their Soret excitation profile was
more complicated, however. Monovinyl Chl(ide)
b
exhibited a major excitation
maximum at 475 nm and a pronounced shoulder at 486-489 nm (Wu et al.
1989
).
Such a splitting of Soret excitation bands into By(0-0) and Bx(0-0) components is
rather common in asymmetrical tetrapyrroles and was also observed in DV and MV
Pchlides (Tripathy and Rebeiz
1985
). Likewise, DV Chl(ide)
b
exhibited a split
Soret excitation band with a maximum at 489-490 nm and a shoulder at
498-500 nm (Wu et al.
1989
). Since the contribution of the DV Chl(ide)
b
signal
to the MV signal amounted to 1/3.5 at 475 nm, while the contribution of the MV Chl
(ide)
b
signal to the DV Chl(ide)
b
signal amounted to 1/6.6 at 498 nm, the two
aforementioned wavelengths were judged to be most suitable for the mathematical
deconvolution of the DV from the MV Chl(ide)
b
signals.
3.7.2 Calculation of the Net Fluorescence Amplitudes
at 447 and 458 nm of Monovinyl and Divinyl
Chlorophyll(ide) a, Respectively, in a Mixture
of the Two Compounds
Let
a
and
b
represent any two fluorescence excitation(
a
) and emission (
b
)
wavelengths of fluorescent compound X. Likewise, let
c
and
d
represent any two
fluorescence excitation (
c
) and emission (
d
) wavelengths of fluorescent compound
Y.
It has been demonstrated above (Sect.
3.5.1
) and elsewhere (Rebeiz et al.
1975a
)
that the deconvoluted net fluorescence signals,
X(E
a
F
b
)
and
Y(E
c
F
d
), generated by
compound
X
and Y, respectively, at the designated wavelengths
a, b, c,
and
d,
can
be determined from the equations described in Sect.
3.5.1
, namely
X
ð
E
a
F
b
Þ¼ ð
½
F
a
F
b
Þð
E
c
F
d
Þ=
k
2
ð
l
=
K
1
Þ
(3.37)
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