3. FOOD ChEMISTRy & bIOTEChNOLOGy 3.1. Lectures

Chem. Listy, 102, s265–s1311 (2008) Food Chemistry & Biotechnology
lene-bis-acrylamide, 0.37M Tris-HCl (pH 8.8) 0.1 % (w/v)
SDS, 0.04 % (v/v) n,n,n’,n’-tetramethylethylene-diamine
(TEMED), and 0.1 % (w/v) ammonium persulphate. Stacking
gels contained 5 % (w/v) acrylamide, 0.13 % (w/v)
n,n´-methylene-bis-acrylamide, 0.19M Tris, 0.02 % (v/v)
TEMED, and 0.1 % (w/v) ammonium persulphate.
A 5 μl of sample solution were loaded on polyacrylamide
gel. Electrophoresis ran 60 min at the constant voltage
140 V. The large subunit of Rubisco was detected by staining
with Coomassie brilliant blue and its identity was confirmed
by co-electrophoresis of Rubisco standard (Sigma-Aldrich)
(Fig.1). Quantification of individual bands was performed on
HP Scanjet 5590P with programme Advanced Image Data
Analyzer, ver. 3.23.001 (Raytest, Germany).
Results
Fig. 1. A representative SDS-PAGE gel with separated proteins
from Norway spruce needles (six left columns) and Rubisco
standard 1 mg ml –1 (the first column from the right). The upper
arrow indicates the large subunit of Rubisco protein and the
lower arrow indicates the small one
Both initial and total activities in E were lesser than
those in A with the exception of autumn. It is in good accordance
with data of other authors 14–16 . Maximum of activities
we measured in May and then they gradually decreased.
Rubisco initial activity in A decreased about 57 % while in
E dropped about 41 % during the growing season. Total activity
decreased about 66 % in A and about 44 % in E. Rubisco
content dropped about 55 % in A and about 50 % in E. The
sharp diminution of Rubisco content in E in July could be
caused by enormous drought in the first half of summer in
combination with the phenomenon that elevated CO 2 concentrations
accelerate the senescence of leaves 6,8,10 . Both initial
and total activities as well as content decreased less in E than
in A (Fig. 2.). We hypothesised that down-regulation in E – if
there was any – proceeded in the first year of needles life.
s658
Fig. 2. Rubisco activity and content in Norway spruce, A –
ambient [CO 2 ], E – elevated [CO 2 ], n = 10
Conclusions
We confirmed a decrease both Rubisco initial and total
activities as well as Rubisco content in norway spruce during
the growing season. According to our measurements, there is
not down-regulation of Rubisco activity and its content in last
year needles in norway spruce during the growing season.
The work forms a part of the research supported by
grants No. 522/06/0930 (GA CR) and by the Research Intention
of ISBE AS CR AV0Z60870520.