1021-4437/02/4902- $27.00 © 2002
Russian Journal of Plant Physiology, Vol. 49, No. 2, 2002, pp. 242–249. Translated from Fiziologiya Rastenii, Vol. 49, No. 2, 2002, pp. 272–279.
Original Russian Text Copyright © 2002 by Nikolaev, Tropin, Goryachev.
Over 60% of the earth’s surface experiences average
annual temperatures below 0
C . Benthic seaweeds
inhabiting the intertidal zone (littoral) of the Barents
Sea are exposed to subzero temperatures for more than
half of the year. At high tide, the seaweeds are sub-
merged in water with temperature from 0 to –1.8
low tide, the algae become exposed to air at tempera-
tures below 0
C. Air temperatures from –15 to –20
are quite common for these areas. The duration of
freezing exposure varies, depending on the location of
the algae on the littoral with respect to the low-water
line. The algae inhabiting the upper littoral zone are
subjected to subzero temperatures at regular intervals
during high and low ebb tides (up to 10–12 h a day).
The algae inhabiting the low littoral zone are exposed
to air only for 2–4 h during neap tides (4–6 times during
29 days). Despite these “severe” environmental condi-
tions, the seaweeds not only survive but also develop
reproductive organs initiated in autumn [2, 3].
The adaptive potential of aquatic plants to freezing
temperatures has come under study since the work by
Kylin . The majority of studies dealt with the deteri-
orative action of freezing temperatures at a whole plant
level [5–8], as well as at the cellular [9, 10] and bio-
chemical  levels of thallus organization. However,
the intracellular mechanisms of seaweed resistance to
freezing temperatures still remain unknown.
The exposure to freezing temperatures impairs mor-
phological, physiological, and biochemical characteris-
tics of plant cells. The cells would need speciﬁc mech-
anisms to restore their viability after freezing treatment.
In this context, the role of natural cryoprotectants, i.e.,
polysaccharides, is particularly emphasized in the liter-
ature on benthic macroalgae . One of the feasible
and important approaches to unveiling the mechanisms
of algal tolerance to freezing temperatures consists in
studying the phasic transitions of intracellular water in
algae during cold treatment.
In this work, we studied the changes in physical
state of water in vegetative apices of intertidal macroal-
gae of the Barents Sea at freezing temperatures.
MATERIALS AND METHODS
Collection, delivery, and maintenance of algae.
Undamaged 3- to 5-year-old shoots of
(L.) Le Jol.  were collected at the end of
March and beginning of April 2000 at the intertidal
zone (water temperature –1.5
C) of the Barents Sea
(Dalnie Zelentsy Biological Station; 69
mansk Marine Biological Institute of the Kola Science
Center, Russian Academy of Sciences). The collected
plants were cleaned of visible epiphytes and epibionts,
placed in polyethylene bags, and transported to Mos-
cow within 75 h in a thermally insulated bag (5
algae were placed in an aquarial system (
= 200 l)
reproducing the main climatic and hydrochemical
parameters of the Barents Sea (photon ﬂux density
s), temperature 3
C, salinity 34‰, pH 8.2).
Algae were kept in aquarium for ten weeks prior to
Culturing of algae in experiments.
In the low-tem-
perature experiment, vegetative apices (shoot segments
Water Status Changes in Shoots of a Seaweed
at Subzero Temperatures
G. M. Nikolaev, I. V. Tropin, and S. N. Goryachev
Faculty of Biology, Moscow State University, Vorob’evy gory 1-12, Moscow, 119899 Russia;
fax: 7 (095) 939-1115; e-mail: firstname.lastname@example.org
Received December 8, 2000
—The gradient freezing and NMR spectroscopy were used to study the physical state of water in api-
ces of the intertidal seaweed
at freezing temperatures. In the apices exposed to temper-
atures below –10
C, two fractions of bound water were revealed. The slow (T
~ 50 ms) fraction of bound water
was completely frozen at –25
C, and its freezing rate was temperature-sensitive. This fraction was apparently
associated with protoplasmic water and cell-wall polysaccharides. The fast fraction (T
< 10 ms) of bound water
was presumably due to water-soluble globular proteins. The freezing rate for this fraction depended on neither
the temperature nor the amount of water. The presence of unfrozen water in apical cells at –40
C was demon-
strated. The role of this water fraction in maintaining the native structure of biomacromolecules and apex sur-
vival is discussed.
Key words: Ascophyllum nodosum - NMR spectroscopy - gradient freezing - algae - subzero temperatures -
water - biomacromolecules