Nonintrusive Indicator for Rapid Assessment of In Vivo Photosynthesis
51
In addition, variations in energy distribution between the two photo systems
will also influence fluorescence yield. As PS I is essentially nonfluorescent,
any increase in energy transfer from PS II to PS I may be considered
equivalent to nonradiative dissipation. Fluorescence emission competes with
photochemistry and heat dissipation. Therefore, two basic types of fluorescence quenching, photochemical and nonphotochemical, can be distinguished
(see Sect. 3.5).
3.3 Rapid Fluoresceuce Induction Kinetics
The existence of two fundamentally different types of fluorescence quenching
has until recently prevented the practical use of chlorophyll fluorescence
in steady-state investigations. However, provided that changes in heat dissipation are relatively slow, the rapid fluorescence induction transients occurring upon a dark-light transition can be interpreted in terms of changes in
photochemical quenching only, which is determined by the concentration of
open reaction centers (Kautsky et al. 1960; Duysens and Sweers 1963). When
all reaction centers are open (QA fully oxidized), the minimal fluorescence
yield, F o , is observed, whereas the maximal fluorescence yield, Fm, is found
when all centers are closed (QA fully reduced). The difference between Fo
and Fm is called variable fluorescence, Fv. For a wide variety of darkadapted plants the Fm/Fo ratio amounts to values of 5-6, corresponding to
FjFm values of 0.8"':'0.833 (Bjorkman and Demmig 1987).
When a dark-adapted sample is illuminated with actinic light, QA reduction rate initially is higher than the rate of reoxidation by plastoquinone
and by PS I activity. The resulting fluorescence rise reflects the exhaustion
of the PS II acceptor pool (Fig. 3.2). A measure for the acceptor pool size is
given by the area between the induction curve and the Fm-line (Murata et al.
1966). In the presence of DeMU, which blocks electron transfer from Q A to
QB, a steep fluorescence rise with a complementary area corresponding to
one electron is observed. In a control sample, the area amounts to approximately 20 electron equivalents.
The relationship between QA reduction and variable fluorescence yield is
nonlinear (loliot and loliot 1964), presumably due to cooperativity between
PS II units at the antenna level. The increase in fluorescence yield is smaller
than predicted from the extent of QA reduction, as excitons absorbed in
closed units may be transferred to neighboring open units (see Fig. 3.3). The
degree of cooperativity between PS II units in vivo is under debate (Keuper
and Sauer 1989). The issue is complicated by the existence of PS II heterogeneity, with PS IIa units displaying a high extent of cooperativity and PS II
~ operating as separate units (Melis and Schreiber 1979; see review by Melis
1991).
51
In addition, variations in energy distribution between the two photo systems
will also influence fluorescence yield. As PS I is essentially nonfluorescent,
any increase in energy transfer from PS II to PS I may be considered
equivalent to nonradiative dissipation. Fluorescence emission competes with
photochemistry and heat dissipation. Therefore, two basic types of fluorescence quenching, photochemical and nonphotochemical, can be distinguished
(see Sect. 3.5).
3.3 Rapid Fluoresceuce Induction Kinetics
The existence of two fundamentally different types of fluorescence quenching
has until recently prevented the practical use of chlorophyll fluorescence
in steady-state investigations. However, provided that changes in heat dissipation are relatively slow, the rapid fluorescence induction transients occurring upon a dark-light transition can be interpreted in terms of changes in
photochemical quenching only, which is determined by the concentration of
open reaction centers (Kautsky et al. 1960; Duysens and Sweers 1963). When
all reaction centers are open (QA fully oxidized), the minimal fluorescence
yield, F o , is observed, whereas the maximal fluorescence yield, Fm, is found
when all centers are closed (QA fully reduced). The difference between Fo
and Fm is called variable fluorescence, Fv. For a wide variety of darkadapted plants the Fm/Fo ratio amounts to values of 5-6, corresponding to
FjFm values of 0.8"':'0.833 (Bjorkman and Demmig 1987).
When a dark-adapted sample is illuminated with actinic light, QA reduction rate initially is higher than the rate of reoxidation by plastoquinone
and by PS I activity. The resulting fluorescence rise reflects the exhaustion
of the PS II acceptor pool (Fig. 3.2). A measure for the acceptor pool size is
given by the area between the induction curve and the Fm-line (Murata et al.
1966). In the presence of DeMU, which blocks electron transfer from Q A to
QB, a steep fluorescence rise with a complementary area corresponding to
one electron is observed. In a control sample, the area amounts to approximately 20 electron equivalents.
The relationship between QA reduction and variable fluorescence yield is
nonlinear (loliot and loliot 1964), presumably due to cooperativity between
PS II units at the antenna level. The increase in fluorescence yield is smaller
than predicted from the extent of QA reduction, as excitons absorbed in
closed units may be transferred to neighboring open units (see Fig. 3.3). The
degree of cooperativity between PS II units in vivo is under debate (Keuper
and Sauer 1989). The issue is complicated by the existence of PS II heterogeneity, with PS IIa units displaying a high extent of cooperativity and PS II
~ operating as separate units (Melis and Schreiber 1979; see review by Melis
1991).