**Tolerance to high copper ions concentration in the nutrient medium of
some Bulgarian barley cultivars**

J. Stoinova^1^, S. Phileva^1^, M. Merakchyiska^2^ and S. Paunova^2^

^1^ D. Kostov Institute of Genetics, Bulgarian Academy of Sciences, 1113
Bulgaria

^2^ M. Popov Institute of Plant Physiology, Bulgarian Academy of\
Sciences, 1113 Bulgaria

**Abstract.** The winter two-rowed barley cultivars Obzor, Krasii 2,
Vihren and Karan were examined for tolerance to environmental copper
ions pollution. The plants were treated with 10^-6^ M and 10^-5^ M
CuSO~4~.5H~2~O for 7 days. It was found that under 10^-6^ M the roots of
cultivars Krassi 2 and Vihren were longer by 9% and 16% respectively,
while plant green part was identical to that of the control. The root of
cultivar Karan was severely inhibited its length reaching 77% that of
the control. The green part was less affected. The data suggest that
cultivars Krasii 2 and Vihren are tolerant while cultivar Karan is
susceptible to increased copper ions concentration in the environment.

**Key words:** barley, tolerance, copper ions

Barley is an economically important crop being efficiently used in
animal husbandry and brewing. Resistant to abiotic and biotic stress
barley cultivars are a reliable tool to produce high quality stuff
avoiding diverse chemical substances for pest control and other
agricultural practices. Acevedo and Fereres (1993 ) concluded that
breeding for abiotic stress resistance is becoming more promising with
the recognition that selection showed be carried out in the target
environment and that it is related to narrow adaption.

During the last decades barley resistance to certain heavy metals has
been examined in different aspects. It was shown that increased boron
concentration in the environment reduce plant growth rate, while
relative susceptibility of the genotypes to boron toxicity was not
affected ( Nable et al.,1999). The effect of high boron concentration on
barley was studied on cell level ( Jenkin et al.,1993). Data from
studies with leaf protoplasts revealed that lack of cell walls prevents
manifestation of differences between tolerant and susceptible barley
genotypes. Treatment of *H*. *vulgare* seeds with nickel sulphate
resulted into morphological alterations and increased chlorophyll and
protein percentage, as well as that of the aberrant cells (Mishra and
Singh, 1999). Addition of copper (2-4 kg/ha) led to higher grain yield
from wheat, rye and oats (Piening et al., 1989). Tang et al. (2000)
studied the location of genes for tolerance to aluminium and found that
the latter is disposed to the long arm of 4H chromosome, while according
to Rigin and Yakovleva (2001) the tolerance is coded by two polygenes.

The purpose of this investigation was to examine the tolerance to
increased copper ions concentration in the nutrient medium of four
Bulgarian barley cultivars.

**Materials and Methods**

The winter two-rowed cultivars Obzor, Krassi 2, Vihren and Karan were
studied. The seeds were germinated in moist filter paper rols in a
semi-dark chamber at 18° C. After germination (2.5 -- 3.5 cm root
length) the shoots were transffered on a solution containing 10^-6^ M or
10^-5^ M CuSO~4~.5H~2~O at a temperature of while the control plants
were grown in distilled water. All plants were grown at 12 h
illumination/12 h dark and 20°- 24° C. After 7 days treatment biometric
data were collected. To determine copper tolerance of the plants usually
the tolerance index (Simon, 1978) was used, i.e. the plant growth in a
heavy metal polluted medium was compared to that under control
conditions:

![](media/image1.wmf)

The data of copper influence on length of root and shoot have been
processed by the variation statistical method.

**Results and Discussion**

A three-fold treatment with 10^-6^ M copper ions concentration of the
root (cultivar Obzor) caused inhibition (93-94% as compared to the
control ) or stimulation up to 103%. In the case of the cultivar Karan
the roots were strongly inhibited their length varying from 62% to 94%
of the control. The plants of the cultivar Krassi 2 and cultivar Vihren
responded to two of the treatments through stimulated root growth from
122% to 135% and from 116% to 129%, respectively. The shoot length was
less affected, some inhibition varying from 1% to 6% or stimulation up
to 2 -- 4% being manifested by the cultivars Obzor, Krassi 2 and Vihren.
More significant inhibition was manifested by cultivar Karan.

The higher (10^-5^ M) copper ions concentration severely inhibited both
root and shoot of the plants from all cultivars after the three
treatment accomplished.

The mean data (Table 1) show that 10^-6^ M ions stimulate (9-16%) root
growth of the plants from cultivars Krassi 2 and Vihren while the roots
of the plants from Karan reaced only 77% of the control. The green part
of the plants was equal to that of the control ones. The variation
coefficients revealed slight heterogenic variation, while the plant
green part length variation was homogenic, exept that for cultivar
Karan. Higher (10^-5^ M) copper ions concentration highly reduced root
growth, root length reaching 32-41% that of the control. The aboveground
part of the plants from cultivar Krassi 2 and cultivar Vihren was
identical to that of the control while those of cultivar Karan and Obzor
were 69-75% that of the control. The variation coefficients revealed
slight heterogenic variation.

Under 10^-6^M copper ions concentration root mass of cultivar Obzor and
cultivar Karan reaching 90-91% that of the control, the negative effect
being stronger than that exerted on cultivars Krassi 2 and Vihren. The
green part mass increased (1-3%) in the case of cultivar Obzor, cultivar
Krassi 2 and cultivar Vihren, while that of cultivar Karan was slightly
inhibited.

10^-5^ M copper ions concentration redused plant green part and root
mass to 75-79% and 52-61%, respectively.

To conclude, the cultivars Krassi 2 and Vihren manifested tolerance to
increased copper ions concentration in the environment as under 10^-6^ M
the root was 9-16% longer than that of the control combined with less
reduction of the mass (94%) of the control. The grain yield was 672
kg/da and 677 kg/da, respectively. Therefore, they are suitable for
cultivation in highly polluted regions.

Cultivar Karan proved to be susceptible to copper pollution its root
being severely reduced.

**Reference**

Avecedo,E. and Fereres, E. 1993. Resistance to abiotic stress. In: Plant
Breeding: principles and prospects (ed. by Hayward, M .D; Bosemark, N.
O., Romagoza, I.). London, UK, Chapman and Hall Ltd, 406-421.

Jenkin, M., Hu, H. N.,Brown, P., Graham, R., Lance, R. and Sparrow, D.
1993. Investigation on boron uptake at the cellular level. Plant and
Soil, 155/56, 143-146.

Mishra, K. and Singh, R. J. 1999. Nickel genotoxicity assessment in
*Hordeum vulgare*. J of Environmental Biology, 20, 71-72.

Nable, R. O., Lance, R. C. H. and Cartwright, B. 1990. Uptake of boron
and silicon by barley genotypes with differing susceptibilities to boron
toxicity. Annals of Botany, 66, 83-90.

Piening, L. J., Mac Pherson, D. J. and Malhi, S. S. 1989. Stem melanosis
of some wheat, barley and oat cultivars on a copper deficient soil. Can.
J of Plant Pathology, 11, 65-67.

Rigin, B.V. and Yakovleva, O. V. 2001. Genetic aspects of barley
tolerance to toxic aluminium ions. International research-practic
conference Sankt-Peterburg, 13-16 november, 2001, p. 397.

Simon, E. 1978. Heavy metals in soils, vegetation development and heavy
metal tolerance in plant populations from metalliferous areas. New.
Phytol., 81, 175-188.

Tang, Y., Sorre, M. E., Kohain, L.V. and Garvin, D. F. 2000.
Identification of RFLP markers linked to the barley aluminium tolerance
gene Alp. Crop science, 40, 778-782.

Table 1. Influence of copper ions on the length of root and shoot

  --------- ---------- ---------- ----------- ------- ---------- ------------ -------
  Variant   Cultivar   Length                         Length                  
                       root                           shoot                   

                       %control   M ± m       Vc%     %control   M ± m        Vc%

  Control   Obzor      100        12.0 ± 0.13 10.57   100        14.30 ± 0.13 8.93

  10^-6^M              97         11.7 ± 0.18 12.84   99         14.20 ±.0.13 7.88

  10^-5^M              38         4.6 ±.0.36  14.82   75         10.67 ±.0.13 11.91

  Control   Krassi 2   100        12.8 ± 0.21 15.28   100        14.40 ±.0.13 9.62

  10^-6^M              116        14.8 ± 0.23 15.51   100        14.40 ± 0.12 8.49

  10^-5^M              41         5.3 ± 0.25  12.40   73         10.50 ± 0.11 11.11

  Control   Vihren     100        13.7 ± 0.29 19.65   100        16.40 ± 0.15 8.40

  10^-6^M              109        15.0 ± 0.23 15.29   99         16.20 ±.0.16 9.47

  10^-5^M              40         5.5 ± 0.08  15.75   72         11.80 ± 0.16 12.85

  Control   Karan      100        14.5 ± 0.23 14.65   100        15.40 ± 0.18 10.26

  10^-6^M              77         11.2 ± 0.22 16.50   98         15.10 ± 0.21 12.16

  10^-5^M              32         4.6 ± 0.08  16.49   69         10.60 ± 0.19 16.15
  --------- ---------- ---------- ----------- ------- ---------- ------------ -------