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T h e u p t a k e o f c o p p e r a n d l e a d s u p p l i e d i n n u t r i e n t s o l u t i o n ( 5 0 0 M ) a n d t h e i r d e t r i m e n t a l i m p a c t o n p h o t o s y n t h e t i c p i g m e n t s c o n c e n t r a t i o n a n d d r y matter accumulation in three lettuce cultivars were investigated. The concentration of copper and lead in lettuce leaf ranged from 8.62 to 14.20 mg kg-1 leaf dry weight for copper, and from 1.97 to 4.05 mg kg-1 leaf dry weight for lead, after two days of exposure. In copper and lead treated plants significant differences in leaf and root dry weight accumulation were not established, but concentrations of chlorophyll b and carotenoids showed significant reduction in cultivar Triatlon. In other words, tolerant genotype (cultivar Nadine) accumulated less copper and lead in leaves, the edible parts of lettuce, therefore such genotypes could be recommended for planting on potentially polluted soils.
Keywords: copper, dry matter content, lead, heavy metal stress, lettuce, photosynthetic pigments
Introduction
Vegetables are very important source of beneficial organic compounds and minerals in human diet. When they are planted on urban and suburban contaminated soils, they can be potentially dangerous carriers of heavy metals, like cooper and lead, in to the food chain. In the research of Hibben et al. (1984), lettuce planted in contaminated soils accumulated more lead than other vegetables. High heavy metals uptake causes oxidative stress in plants which is manifested through induction of peroxidases (POX or POD; EC 1.11.1.7, Singh et al., 1997). In the research of El-Tayeb (2006), oxidative stress can also cause decrease of photosynthetic pigments concentration (chl a, chl b, carotenoids) in wheat under drought cond i t i o n s . T o x i c i t y o f h e a v y m e t a l s i n p l a n t o r g a n i s m c a n c a u s e d a m a g e o f p h o t o s y n t h e t i c p i g m e n t s t h a t a c c o r d i n g t o V e r e s e t a l . ( 2 0 0 7 ) i n d u c e r e m i s s i o n o f d r y m a t t e r p r o d u c t i o n , w h i c h i s v e r y i m p o r t a n t i n a g r o n o m y . K i s i e t a l . ( 2 0 0 7 ) , r e p o r t e d t h a t a i r a n d water pollution have been broadly addressed and included in teaching curriculum and science researches, while the heavy metal soil contamination has been highly neglected. Blint et al. (2007) stated that it would be recommendable to avoid planting on heavy metal contaminated soils. The aim of the research presented here is to evaluate short term copper and lead toxicity in lettuce using photosynthetic pigments concentration as well as dry matter accumulation as stress parameters.
Material and methods
Seeds of three lettuce (Lactuca sativa L.) cultivars were planted for three weeks in glasshouse conditions in commercial substrate, till the four to five leaf stages. After washing thoroughly with running deionised water they were fixed on perforated polystyrene fasteners of glass pots containing nutrient solution for leafy vegetables after Lee and Park (2001). The nutrient medium was renewed every 3 days and aerated for one hour each day. The experiment was settled in four replicates and each replicate contai n e d f o u r p l a n t l e t s . T h e p o t s w e r e k e p t f o r t h r e e w e e k s i n a g r o w t h c h a m b e r a t 2 0 o C , w i t h 7 0 % r e l a t i v e h u m i d i t y a n d 1 2 - h p h o t o p e r i o d . T h e f l u o r e s c e n t l a m p s w e r e u s e d t o p r o v i d e t h e p h o t o s y n t h e t i c p h o t o n f l u x d e n s i t y 1 2 0 m o l / m 2 / s a t t h e l e a f l e v e l . S u b s e q u ently, the plantlets were treated with 500 M Cu(SO4) or Pb(NO3)2 (final concentration) in nutrient solution for 2 days.
Thereafter, plantlets were divided in roots and leaves. The amount of tissue dry weight was determined by drying at 70oC for 24 h. Dry samples of leaves were subtracted to wet digestion using of 4% HClO4 acid in concentrated H2SO4, with addition of H2O2. Copper and lead concentration was measured by ICP-AAS.
Leaf chloroplast pigments were extracted from the most developed leaf using 100% acetone. The absorbance of the extracts was measured spectrophotometrycally (Wettstein, 1957). The individual levels of chlorophyll a (Chl a), chlorophyll b (Chl b), total chlorophylls (Chl a+b) and total carotenoids (Car) were calculated. The results of plant analysis were statistically analysed using ANOVA with F-test for the evaluation of significance of the applied treatments, as well as t-test in single regression analyses for the evaluation of the significance of established correlations among tested p a r a m e t e r s ( * P d"0 . 0 5 ; * * P d"0 . 0 1 ) .
R e s u l t s a n d d i s c u s s i o n
T h e c h l o r o p h y l l c o n t e n t o f t h e l e a v e s c a n i n f l u e n c e o n d r y m a t t e r i n c o r p o r a t i o n , w h i c h a f f e c t s o n y i e l d f a c t o r s ( S z l e s , 2 0 0 7 ) . H i g h e r c h l o r o p h y l l c o n t e n t s m a y e n h a n c e p h o t o s y n t h e s i s a n d r e s u l t i n h igher amount of dry matter (Veres et al., 2007). As stated by Vassilev et al. (2007) at low degree of metal contamination the visible symptoms, such as chlorosis, necrosis of leaf tips, etc. are less pronounced or even could be absent, although some reduction of plant quality as well as biomass inhibition persist. Here, copper treatment slightly reduced fresh weight and dry weight of the leaf in all three tested cultivars (Table 1) while plantlets treated with lead showed appreciable higher values concerning control plants. The influence of heavy metals on root dry weight was diverse, regarding cultivars. Significant decline in root dry weight was observed in cooper treated Marcord, whereas root dry weight was higher in copper-treated Nadine and lead-treated Triatlon ( P d"0 . 0 5 ) .
T h e a p p l i e d c o p p e r a n d l e a d c o n c e n t r a t i o n i n n u t r i e n t s o l u t i o n , a l t h o u g h t h e e x p o s u r e l a s t e d o n l y t w o d a y s , r e s u l t e d w i t h t h e i r m o s t l y s i g n i f i c a n t a c c u m u l a t i o n i n t h e l e a v e s o f l e t t u c e . H e a v y m e t a l c o n t e n t w a s i n t h e r a n g e f r o m 1 2 . 7 7 ( N a d i n e ) t o 1 5 . 2 2 m g k g - 1 D W l e a f ( T r i a t l o n ) f o r c o p p e r a n d f r o m 1 . 9 7 ( N a d i n e ) t o 4 . 5 8 m g k g - 1 D W l e a f ( T r i a t l o n ) f o r l e a d ( T a b l e 1 . ) . T h e r e s u l t s s h o w e d s i g n i f i c a n t r e m i s s i o n o f c h l o r o p h y l l a ( d a t a n o t s h o w n ) a n d b u n d e r t r e a t m e n t w i t h 5 0 0 M P b i n c u l t i v a r T r i a t l o n . Treatment with copper had not been statistically justified for chlorophyll a, carotenoids and total chlorophylls, but chlorophyll b was notably reduced in all cultivars.
Regarding the ratios of chl a/chl b as well as chl/car, the influence of the applied heavy metal stress was not significant, except in chl a/chl b in cultivar Triatlon, where it was lowered by copper treatment ( P d"0 . 0 5 ) .
T a b l e 1 . T h e i m p a c t o f l e a d a n d c o o p e r t r e a t m e n t s ( 4 8 h ) o n c o n c e n t r a t i o n a n d r a t i o s o f p h o t o s y n t h e t i c p i g m e n t s a s w e l l a s t h e a c c u m u l a t i o n o f d r y m a t t e r a n d h e a v y m e t a l s i n l e t t u c e g r o w n i n n u t r i e n t s o l u t i o n
( d a t a a r e m e a n s o f f o u r r e p l i c a t e s ; the concentration of lead (Pb) and cooper (Cu) in nutrient solution 500 M Cu(SO4) or Pb(NO3)2 and control; chl b - chlorophyll b; chl a- chlorophyll a; car - carotenoids; FW fresh weight; DW dry weight).
CultivarHeavy metal treatmentchl bcarchl a/
chl bchl/
carDW
rootDW
leafCu
leafPb
leafmg g-1 FWmg g-1 FWmg plant-1mg kg-1 DWTriatloncontrol0.1230.1002.03.5242768.301.88Cu0.0830.0953.03.12526015.22-Pb0.0790.0802.83.429292-4.58Marcordcontrol0.1090.0922.23.5242328.502.55Cu0.0970.0862.13.31918914.63-Pb0.0880.0952.42.522257-3.92Nadinecontrol0.1140.1042.03.2322889.051.78Cu0.1160.1261.92.73827812.77-Pb0.1710.1061.13.634288-3.65meancontrol0.1150.0992.13.4272658.621.97Cu0.0990.1032.43.02824214.20-Pb0.1130.0942.13.229279-4.05LSD P d"0 . 0 5 0 . 0 3 3 0 . 0 1 9 1 . 0 0 . 7 4 . 2 4 9 . 6 2 . 2 1 1 . 7 6 L S D P d"0 . 0 1 0 . 0 4 5 0 . 0 2 6 1 . 4 1 . 0 5 . 8 6 7 . 9 3 . 1 8 2 . 5 2
F i g u r e 1 . T h e c o r r e l a t i o n b e t w e e n c a r o t e n o i d s c o n c e n t r a t i o n a n d l e a f f r e s h w e i g h t i n l e t t u c e c u l t i v a r s w i t h o u t h e a v y m e t a l t r e a t m e n t ( * * P d"0 . 0 1 ) F i g u r e 2 . T h e d i f f e r e n c e s i n c o r r e l a t i o n s a m o n g c h l o r o p h y l l s a n d c a r o t e n o i d s c o n c e n t r a t i o n s i n c o n t r o l ( y 1 ) , c o p p e r ( y 2 ) a n d l e a d ( 3 ) t r e a t e d l e t t u c e ( * P d"0 . 0 5 , * * P d"0 . 0 1 )
W i t h a n i n c r e a s e o f c a r o t e n o i d s c o n c e n t r a t i o n , f r e s h w e i g h t c o n t e n t w a s v e r y s i g n i f i c a n t l y increased in control plants, as shown in Figure 1 (r = 0.861**). In all treatments, highly significant correlations between total chlorophylls and carotenoids concentration were established. The highest correlation coefficient was obtained in control treatment (r = 0.831**, y1), where mostly higher concentrations of chlorophyll and carotenoids were observed. Excessive lead supply also gave high correlation between chlorophyll and carotenoids content in lettuce leaves (r = 0.785**, y3), while in copper stressed plants the lowest significant correlation was obtained (r = 0.634*, y2), that started from lowest level of photosynthetic pigments concentration.
Conclusions
The accumulation of Cu and Pb in lettuce was accompanied with the reduction of photosynthetic pigments concentration in general. A short term treatment with high concentration of the applied heavy metals in nutrient solution resulted with non significant differences between cultivars in accumulation of dry weight, ( 4 J X Z p v
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Vol. 36, 2008, Suppl. Cereal Research Communications
VII. Alps-Adria Scientific Workshop Stara Lesna, Slovakia, 2008
PAGE 2
PAGE 1
DOI: 10.1556/CRC.36.2008.Suppl.1
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