Analele Ştiinţifice ale Universităţii „Al. I. Cuza” Iaşi
s. II a. Biologie vegetală, 2014, 60, 2: 5-10
http://www.bio.uaic.ro/publicatii/anale_vegetala/anale_veg_index.html
ISSN: 1223-6578, E-ISSN: 2247-2711
HISTO-ANATOMICAL ASPECTS IN MAIZE (ZEA MAYS L.) SEEDLINGS
DEVELOPING UNDER INFLUENCE OF DEUTERIUM DEPLETED WATER
Corneliu TĂNASE1*, Irina BOZ2, Valentin I. POPA3
Abstract. The paper presents histo-anatomical aspects, evidenced in seedlings of maize (Zea mays L.)
grown under the influence of deuterium depleted water (DDW). The aim of this study was to identify the
internal structure changes in maize seedlings resulted in this case. For histo-anatomical characterization,
the vegetal material was fixed and preserved, following to be cut by hand microtome with botanical razor.
The sections obtained were stained using double staining reagents. As compared with the control sample
it was obsereved that deuterium depleted water determines a better development of the absorbents bristle
and increase leading bundles number and central vessels in metaxilem.
Keywords: deuterium depleted water, maize, histo-anatomical investigations.
Introduction
DDW or light water, is a distilled water that is micro-biologically pure, containing
an isotopic concentration of 25 ppm, obtained by isotopic distillation in vacuum of natural
water with an isotopic concentration of 145 ppm D/(D + H) (Somlyai, 1998). In Romania,
DDW is obtained in two centres: the heavy water plant at Halânga, where daily is discharge
as waste and in National Institute of Research and Development for Cryogenic and Isotopic
Technologies of Ramnicu Valcea in a pilot plant. The properties of deuterium depleted
water are similar to those of normal water excepting the isotopic concentration, which is 25
ppm, compared to 145 ppm for normal water (Tanase et al. 2013a),.
On the other work (Somlyai et al. 1993; Tanase et al., 2013a; Tanase et al., 2013b;
Tanase et al. 2014) it is known that the DDW has a great influence on living organisms,
namely in developing cells and tissues, and changes that occur in normal water features
which lead to significant changes in fundamental processes of the cells. Thus, was observed
that DDW influenced the development and multiplication of cells, cellular transport, DNA
synthesis and also antioxidant properties (Somlyai et al., 1993; Olariu et al., 2007). In case
of vegetal systems, DDW stimulates germination energy, radicle and primary leaves
elongation, biomass accumulation, and inhibits photoassimilating pigment synthesis in
soybean (Tanase et al., 2012; Tanase et al. 2014). Since the studies carried out on the
influence of DDW in plant systems are not considered representative, as yet further we
need to develop this subject to obtain supplementary information.
Thus, the aim of this study was to identify the internal structure changes in maize
seedlings, under the influence of deuterium depleted water compared with control.
University of Medicine and Pharmacy of Tirgu Mureș, Faculty of Pharmacy, Botanical Pharmaceutic
Department, Gheorghe Marinescu, 38, 540139, Tirgu Mures, Mures, Romania
2
“Alexandru Ioan Cuza” University of Iasi, Faculty of Biology, Vegetal Biology Department, Bulevardul Carol I,
Nr. 11, 700506 Iasi, Romania
3
Gheorghe Asachi Technical University of Iasi, Chemical Engineering and Environmental Protection Faculty, 71
Bd. Prof. D. Mangeron, 700050, Iasi, Romania
*corresponding author: E-mail: [email protected], phone: +00400744215543
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Tanase, C. et al. 2014/ Analele Stiint. Univ. Al. I. Cuza Iasi, Sect. II a. Biol. veget., 60, 2: 5-10
Materials and methods
Maize seeds were purchased from Unisem Company, Romania. Deuterium depleted
water was provided by Romag Prod, Severin (Halânga), a manufacturer of heavy water
used for the Cernavodă Nuclear Power Station. Tons of deuterium depleted water are
discharged daily.
Germination tests were carried out going through a standard procedure, using
increments of 10 Petri dishes for each solution studied. On a filter paper were placed every
five maize seeds, carefully selected without any major damages. For the begining, the
vegetal material has undergone a pre-sterilization process, which consisted of submerged
seed in absolute ethanol for 10 seconds, following the sterilization in the presence of
sodium hypochlorite 10% for 20-30 minutes (Cachița et al., 2004). The volume of DDW
added was 10 mL / dishes meanwhile for the reference samples distilled water were used.
Petri dishes thus prepared were incubated in the dark in a thermostat set at 28 º C. After a
period of seven days Petri dishes were taken out and the roots, stems and leaves separated
for histo-anatomic analysis.
In order to perform the histo-anatomical characterization, the vegetal material was
fixed and preserved in alcohol 70%, following to be cut by hand microtome with botanical
razor.
The working protocol was carried out in the following steps:
1. First, were performed cross sections, using botanical razor, hand microtome and
marrow elderberry through the vegetative organs of plants and seedlings. Obtained
sections were collected on a watch glass with water.
2. The sections obtained were stained using double staining reagents: green - iodine and
ruthenium red. After removing water from the glass plate, the sections were stained
for several tens of seconds (with a few drops of methylene blue) or 1 minute (with a
few drops of iodine green). After removing the dye, the sections were washed with
70% ethanol and then once with tap water and were then stained for several tens of
seconds with ruthenium red and subsequently washed twice with tap water.
3. Subsequently stained sections were mounted between slide and slide in a few drops of
water and analyzed with a microscope Novex (Holland) and photographed with Nikon
Coolpix L22.
4. In this experiment were investigated ten sections for every sample.
Results and discussions
To highlight the effect of deuterium depleted water some histo-anatomical aspects of
the internal structure of maize seedlings were analysed. Thus, in this paper, histoanatomical elements, regarding the internal structure of primary roots, adventitious roots,
mezocotyl, epicotyl and leaves will be presented.
The primary roots of seedlings that have been grown in the presence of deuteriumdepleted water (DDW) have a rizodermis with numerous root hairs per unit area (Fig. 1).
Aerial cavities from cortex are being formed by local disruption of cortical parenchyma.
The endodermis is tertiary type, with cells having internal wall and partial side walls thicker
than the outside. In contrast with the control samples, endodermis is in the stage of
transition from primary type (with Caspary thickening in radial wall) to the tertiary. The
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Tanase, C. et al. 2014/ Analele Stiint. Univ. Al. I. Cuza Iasi, Sect. II a. Biol. veget., 60, 2: 5-10
number of vascular bundles is constant (15-16), also as that of metaxylem (6) central
vessels; more numerous than those found in control samples. Metaxylem central vessels are
surrounded by a layer of lignified parenchyma cells.
At the adventitious root level (Fig. 2) root hairs are long and numerous, in contrast
to the control where the root hairs are few and have different lengths. The exodermis is
barely visible, with cells higher than the endodermis, but with thin cellulose walls. The
aeriferous cavities from the cortical parenchyma are in training, 5-6 are already formed, but
small. The endodermis is tertiary type, in contrast with the control, where the endodermis is
in stage of transition to tertiary type. The number of vascular bundles is very large (19-20)
compared to control (11-12). The central vessels of metaxylem (10-12) are more numerous
compared with the control (4-5).
The mezocotyl has a transition structure between the root and the stem. The
epidermis has cells with slightly thickened external wall than the other and covered with a
very thin cuticle. In the cortical parenchyma are formed aeriferous cavities with different
size and shape (Fig. 3). The central cylinder is much thinner than in the control and the
endodermis is tertiary type. The root xylem vessels are disposed more irregular and the
vascular bundles are more numerous, than in the control. The pith is thick, parenchymaticcellulose type, the cells from the central area is much larger, all with very thin cellulose
walls.
The coleoptile (Fig. 4) has elliptic cross-section contour. External epidermis shows
isodiametric cells of different size, with external wall thicker than the other and covered
with a thin cuticle. Internal epidermis has larger cells and visibly elongated tangentially to
the walls thin. In clorenchimatic mesophyll it outlines some aeriferous cavities. Apparently
the two vascular bundles are of leptocentric types, although in the central area of phloem a
few parenchyma cells can be distinguished (Fig. 4). The phloem includes sieve elements
and companion cells and xylem vessels are arranged in a uniform ring uni- or bilayer,
separated by xylem parenchyma cells.
The leaf (Fig. 5) has both epidermises with isodiametrical cell, with strong external
wall bulging and slightly thickened. From place to place is observed stomata present in both
the epidermis. In contrast to the controls which bulliform cells are present, in the present
case, the cell of the leaf present bulliform group cells, visible in the upper epidermis. The
hypodermic mesophyll from the upper epidermis presents slightly elongated cells. At the
top of the epidermis starts to draw ribs and valeculs.
Conclusions
The addition of deuterium-depleted water in the growth of maize seedlings causes
the appearance of aeriferous cavities in internal structure of the root. The number of
vascular bundles is higher compared to the control.
In the adventitious root, the root hairs are more numerous compared to the control.
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a
b
Figure 1. Internal structure of root (DDW): a – general aspects; b –central cylinder
bb
a
Figure 2. Internal structure of adventitious root (DDW): a – general aspects; b –central cylinder and
cortex
a
b
Figure 3. Internal structure of mezocotyl (DDW): a – general aspects; b – vascular bundles
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Tanase, C. et al. 2014/ Analele Stiint. Univ. Al. I. Cuza Iasi, Sect. II a. Biol. veget., 60, 2: 5-10
b
a
Figure 4. Internal structure of coleoptiles (DDW): a – general aspects; b – vascular bundle
b
a
Figure 5. Internal structure of leaf (DDW): a – general aspects; b – vascular bundles
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