Survey
* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
DISCUSSION DISCUSSION Heavy metal pollution is a worldwide environmental crisis. At high concentration, heavy metals can accumulate in plant tissues that result in toxic effects affecting both physiological and biochemical processes in plants. Plant growth inhibition and biomass reduction are the basic phenomenon that occur in response to any stress and is true for heavy metal toxicity also (Shekhawat et al., 2010; Shanmugaraj et al., 2013). The addition of toxic heavy metals to the terrestrial communities as a result of anthropogenic activities is a serious concern as it threatens plant and animal life (Raza and Shafiq, 2013). Heavy metal toxic effect on the plants may cause alteration in their metabolic pathways, such as photosynthesis, respiration, growth and modifying plant anatomy. Grain is a developmental stage that is highly protective against external stresses in the plant life cycle (Sharma et al., 2011; Gowayed and. Almaghrabi, 2013). Cadmium (Cd) is one such environmental toxicant, which persists and prevails as toxic heavy metal among animals and plants (Sanita di Toppi and Gabbrielli, 1999; Raza and Shafiq, 2013). Cadmium (Cd) pollution is regarded as one of the most harmful or toxic environmental issues that mainly resulted from mining, use of phosphatic fertilizers, sewage sludge and untreated waste water, concentrations of Cd in agricultural soils have posed a significant threat to safe crop production and have therefore become a global concern (Nedjoud et al., 2012; 137 DISCUSSION Soudeh and Zarinkamar, 2012; Gowayed and Almaghrabi, 2013). Chromium (Cr) is the seventh most abundant metal in the earth’s crust and an important environmental contaminant released into the environment due to its huge industrial use. Chromium is mainly present in the environment as insoluble Cr (III) and Cr (VI) compounds (Surekha and Duhan, 2012). 5.1 Seed Germination Decrease in seed germination of plant can be attributed to the accelerated breakdown of stored nutrients in seed and alterations of selection permeability properties of cell membrane, due to negative effects of heavy metals. Under cadmium stress, seed germination percentage reduced significantly in Milk thistle as compared to control. Control showed the highest, whereas 600 mg L-1 of cadmium concentration resulted to the lowest value of seed germination (Khatamipour et al., 2011). Shafiq et al. (2008), Bahmani et al. (2012) indicated that Cd can cause significant reduction in seed germination percentage of Leucaena leucocephala and bean (Phaseolus vulgaris L.). Dubey (1997) noticed that Cd toxicity adversely affected germination of seeds and seedling vigour in rice by limiting water transport to growing tissues. The negative effect of cadmium on germination rate of has been reported in wheat, barley and rice (Titov et al., 1995; Rascio et al., 2008) and peanut (Shan et al., 2012). Increasing the concentration of CdCl2 during the germination stage had suppressed the seed germination of B. rapa as found also by Asgharipour et al. (2011), Shaimma et al. (2012), Heidari and Sarani (2011). Reduction in seed germination can be attributed to alterations of properties of cell membrane as Barcelo and Kahle (1992) indicated that Cd affected water relations not only by decreasing water absorption and transport, but also by lowering water stress tolerance. Cadmium (Cd) decreased pea seed germination by inhibiting respiration in cotyledons and altering mitochondrial enzyme activities and patterns of oligo-elements and coenzymes (Smiri et al., 2010; Gangwar et al., 2012) as the data shown in Fig. 21 and 22. The high levels (500 ppm) of Cr (VI) in soil reduced germination in the bush bean Phaseolus vulgaris (Parr and Taylor, 1982). Peralta et al. (2001) found that 40 ppm of Cr (VI) reduced ability of seeds of lucerne (Medicago sativa cv. Malone) to germinate and grow in the contaminated medium. Reduction of in sugarcane bud 138 DISCUSSION germination were observed with 20 and 80 ppm Cr, respectively (Jain et al., 2000). This type of toxicity may be due to the depression on oxygen uptake and physiological disturbance in mobilization of reserve food materials of seeds. Chromium which remains in soil for a long time can affect germination of seeds, plant growth, leaf water potential and turgor, and induces plasmolysis (Vazques et al., 1987; Sharma et al., 2003; Vernay et al., 2007). Since seed germination is considered as a sensitive process compared to other stages of plant development, and it represents a limiting stage of plant life cycle (Smiri et al., 2010). Seed germination was reduced in Pisum sativum L. seedlings (Gangwar et al., 2012). The reduced germination of seeds under Cr stress would be due to the depressive effect of cron the subsequent transport of sugars to the embryo axis (Zeid, 2001) Protease activity increases simultaneously with the chromium treatment which couldalso contribute to the reduction in germination of chromium treated seeds (Zeid, 2001; Datta et al., 2011) as the data shown in Fig. 38 and 39. Avoidance and tolerance to metals ions in symplasm by complexation are the two mechanism reported for the tolerance to heavy metals in plants (Seregin and Ivano, 2001; Shi et al., 2009). Tolerance of plant to heavy metal toxicity is usually estimated on the basis of the degree of inhibition in growth by the metal present in a nutrient solution (Gajewska and Sklodowska, 2010) as shown in data in Fig. 22 and 39. 5.2 Phenotypic Characteristics In the present study cadmium exposure showed a significant alteration in the seed germination ratio, biomass content (moisture content and dry weight), root and shoot length of both the cultivars. The seeds grown indifferent concentrations of cadmium showed stunted growth and short lateral root formation (Shekhawat et al., 2010). The decrease in the shoot length and root length seen may be attributed to the accumulation of cadmium in the plant roots that reduces the water and mineral uptake, which will ultimately affect the plant metabolism as the shown in Fig. 29A,B. High levels of cadmium in the soil also damage the root tip, reduces the water transport in the growing tissues, that leads to the reduced transpiration rate and also inhibits photosynthesis by altering the enzymes involved in theCalvin cycle resulting in the stunted growth (Arduini et al., 1996; Chen et al., 2010; Bauddh and Singh, 2011). During stress, one fourth of the root growth was reduced and it positively correlates with the reduction in shoot growth. The results were in accordance with John et al. 139 DISCUSSION (2007), Bauddh and Singh (2011), Gill et al. (2011), as shown in Fig. 23 and 30. Similar results were also obtained in maize, sorghum, tomato, lentils, Albizia lebbeck, wheat and Glycyrrhiza uralensis (Stilborova et al., 1987; Pandit and Prasannakumar, 1999; Jing et al., 2005; Kiran and Shahin, 2006; Farooqi et al., 2009; Ci et al., 2010; Zheng et al., 2010; Shanmugaraj et al., 2013). The most common effects of Cd toxicity in plants are stunted growth, leaf chlorosis and altered activity of many key enzymes in various metabolic pathways (Arduini et al., 1996; Zhang et al., 2009; Zou et al., 2012). Drazic et al. (2004), Aksoy and Dinler (2012) reported that growth of plants under stress conditions were related with concentration, kind of treatment, duration and plant species. Shafi et al. (2009) reported that Cd and NaCl treatment decreased in shoot length of wheat species. Razıuddin et al. (2011) observed excess Cd accumulation in soil can reduce the growth of plants by disturbing the photosynthesis and chlorophyll synthesis. Shan et al. (2012) reported that high Cd concentrations resulted in a reduction in seedlings growth, expressed as shoot and root length, induced root browning in peanut. Normally, Cd ions are retained in the roots and only small amounts are transported to the shoots (Cataldo et al., 1983). Cd in cells gets associated with cell walls and middle lamella and increases the cross-linking between the cell wall components, resulting in the inhibition of the cell expansion growth (Poschenrieder et al., 1989). Moreover, Cd also alters the water relation in plants, causing a physiological drought (Barcelo and Poschenrieder, 1990) and causes metabolic functions such as production of reactive oxygen species (Asada, 1999), photosynthesis (Krupa et al., 1993; Chugh and Sawhney, 1999). Hasan et al. (2007a), Faizan et al. (2011) also reported that the presence of Cadmium in the soil decreases the growth of chickpea plants. The application of Cd reduced the shoot length in bean, wheat and alfalfa (Chaoui et al., 1997; Bhardwaj et al., 2009; Aydinalp and Marinova, 2009), Phaseolus vulgaris (Bahmani et al., 2012) and also Mihalescu et al. (2010) were of the opinion that the reduction in root length and height of the plant was due to Cd accumulation in Zea mays. The reduction in root length affected by Cd treatment in bean (Bhardwaj et al., 2009; Bahmani et al., 2012), Solanum melongena (Siddhu et al., 2008) and alfalfa (Aydinalp and Marinova, 2009) was recorded. 140 DISCUSSION The germination percentage, root length, shoot length, fresh weight, and dry weight of blackgram gradually decreased with the increase in chromium concentrations. This may be due to the accumulation of chromium in seeds (Sankar Ganesh et al., 2006; Chidambaram et al., 2009). Jan et al. (2010), Hamid et al. (2012) who reported that Cr toxicity in B. juncea leads to stunted growth, leaf chlorosis and alteration in the activity of many enzymes of various pathways. Cr (VI) seems to act principally on plant roots, resulting in intense growth inhibition. Increasing concentration of Cr caused significant reduction in root length and shoot length (Gani, 2011) , as shown in Fig. 46A,B. Adverse effects of Cr on plant height and shoot growth have been reported earlier by Rout et al. (1997). Chromium was found to be more toxic affecting root, shoot length. The reduction in the plant height might be mainly due to the reduced root growth and consequent lesser nutrient and water transport to the above parts of the plant. Chromium transport to the aerial part of the plant can have a direct impact on cellular metabolism of shoots contributing to the reduction of plant height (Shankar et al., 2005). Root was found to be more affected than shoot. This is due to the fact that heavy metals (Cr-VI) accumulated on root due to binding of metals (Cr-VI) on the cell wall of root and retard cell division and cell elongation (Woolhouse, 1983). General decreased root growth due to chromium toxicity could be due to inhibition of root cell division/root elongation or to the extension of cell cycle in the roots. The reason of the high accumulation in roots of the plants could be because chromium is immobilized in the vacuoles of the root cells, thus rendering it less toxic, which may be a natural toxicity response o plant (Shanker et al., 2004). Datta et al. (2011) reported that Cr was reduced the growth in Triticum aestivum L.). The changes of root growth extrapolate in succession to the other phytotoxic symptoms on shoot under Cr contamination (Liu et al., 2008). In general, Cr-induced alteration of physiological activities in all plants starts with disease development and malfunctioning of root physiology (Dhir et al., 2009), which is manifested, to impair photosynthetic activities, water relation, translocation of organic solutes and so on as reported in cereal crop (Panda, 2007). A distinct reduction in root growth as well as in allied attributes has been documented in the maize (Maiti et al., 2012) as shown in Fig. 40 and 47. 141 DISCUSSION Higher concentration of Cd significantly reduced number of leaves in pea as similar to chickpea (Faizan et al., 2011) as shown in Fig. 23 and 30. This may be attributed to be due to the senescence and death of older leaves and appearance of injury symptoms on the younger leaves at high Cd stress, thereby reducing number of leaves and leaf area per pot (Ghani and Wahid, 2007).Increasing concentration of Chromium decreased number of leaves in Miscanthus (Arduini et al., 2006) as shown in Fig. 40 and 47. 5.3 Photosynthetic Pigment Chlorophyll is the main pigment which can help plants to photosynthesize. When plants exposure to stress, their photosynthesis will be inhibited and the concentrations of chlorophyll can directly indicate the extent of stress-induced damage in plants (Lin et al., 2012). Hou et al. (2007), Katazyna and Smolik (2011) obtained a decreased amount of chlorophyll a, chlorophyll b and carotenoids in Lemna minor after the application of a Cd treatment. Erdei et al. (2002) recorded a higher decrease in the content of chlorophyll in barley after the application of cadmium as the shown in Fig. 25 and 33. The chlorophyll pigments are present in thylakoid within chloroplast, and any damage brought to these structures can lead to denaturation of these pigments. It may be suggested that observed decrease in chlorophyll content at higher concentration of chromium may be due to breakdown of thylakoid and chloroplast envelope as was previously reported (Dodge and Law, 1974). Reducing in amount of chlorophyll and photosynthesis by heavy metals like Cr has been reported in plants like bean, wheat and corn. There were also significant decreases in chlorophylls a and b content of the B. oleracea var. acephala plants treated with Cr (Ozdener et al., 2011). Levels of assimilatory pigments viz. chl a and b are often related to tissue damage in higher plants. Decrease in total chlorophyll, chlorophyll a, b and carotenoids have been well documented under Cr stress in plants (Panda and Patra, 1998, 2000; Tripati and Smith, 2000; Panda, 2003; Panda and Khan, 2003; Panda et al., 2003) as the shown in Fig. 42 and 50. Cr possesses the capacity to degrade aminolevulinic acid dehydrates, an important enzyme involved in chlorophyll biosynthesis reactions (Shi and Dalal, 1990). Chlorophyll content reduced in Brassica juncea L. (Hamid et al., 2012) and Brassica napus L. (Najafian et al., 2012) with increasing concentration of Cr. 142 DISCUSSION Decrease in chlorophyll may be due to the inhibition in the activity of aminolevulinic acid dehydratase and proto-chlorophyllide reductase activities and breakdown of pigment or their precursors as reported for other stresses (Ouzounidou, 1995). Carotenoids are known to be indispensable constituents of the photosynthetic apparatus, being essential not only for antioxidative protection but also for the efficient synthesis and accumulation of photosynthetic proteins and especially that of PSII antenna subunits (Sozer et al., 2010) therefore, significant decrease in carotenoids could result in adverse consequences on photosynthetic efficiency of the plants. Roots and shoots highly varied in their ability to accumulate Cr. Chlorophyll decreased in Pisum sativum L. seedlings (Gangwar et al., 2012). Total chlorophyll content decreased with increasing concentration of Cr (VI). The formation of chlorophyll pigment depends on the adequate supply of iron as it is the main component of the protoporphyrin, a precursor of chlorophyll synthesis. An excessive supply of Chromium seems to prevent the incorporation of iron into the protoporphyin molecule, resulting in the reduction of chlorophyll pigment in Triticum aestivum L. (Datta et al., 2011). 5.4 Antioxidative Enzymes Heavy metals cause oxidative damage to plants either directly or indirectly through the formation of ROS, which cause further severe oxidative damage to different cell organelles and biomolecules (Radotic et al., 2000). To scavenge ROS, plants possess a well-organized antioxidative defense system comprising enzymatic antioxidants (e.g., SOD, POD, GR, GPX, and CAT) and non-enzymatic antioxidants (GSH, NP-SH, PCs). The cooperative functioning of these antioxidants plays an important role in scavenging ROS and maintaining the physiological redox status of organisms (Cho and Seo, 2005; Zou et al., 2012). Antioxidant enzymes are produced to protect the cells from the accumulation of oxygen free radicals, superoxide radicals and hydroxyl radicals, which causes the deleterious effects to the organisms (Shanmugaraj et al., 2013).One of the basic mechanisms under lying Cd toxicity to plants is increased production of ROS (Romero-Puertas et al., 2004). The increased activity of SOD, CAT, POD and GR caused by Cd2+ has been observed in several plant species and is considered to be an adjustment response to stress (Xu et al. 2008). 143 DISCUSSION Both enzymatic and non-enzymatic antioxidants were increased in Cd treated plants in comparison to the control. The increase in the antioxidant enzymes is one of the defense mechanism in plants to survive from the toxic metal pollutants (Arleta et al. 2001; El-Beltagi et al. 2010). The peroxides produced in response to oxidative stress are converted to water by the antioxidant enzyme catalase, thus preventing membrane damage (Polidoros and Scandalios, 1999; Gill et al., 2011). The increase in peroxidase can develop a physical barrier in the cell by increasing lignin biosynthesis resulting in thickening of tissues, hence protecting the cell from ROS damage (Hegedus et al., 2001; Shanmugaraj et al., 2013). Our result clearly correlated with the results reported in Brassica juncea (Gill et al., 2011; Shanmugaraj et al., 2013), sunflower cotyledons (Gallego et al., 1996) and radish (El-Beltagi et al., 2010) as the shown in Fig. 26 and 34. Cadmium chloride treatments effectively increased the activity of the peroxidase enzyme (POD) in B. rapa leaves (Shaimma et al., 2012). Dinakar et al. (2009), Martinez et al. (2010), Semane et al. (2010) reported increase in POD activities in Spartina densiflora, Arachis hypogaea and Arabidopsis even in low concentrations. The increased activities of POD by cadmium stress suggest that the plant depends on this antioxidative enzyme for elimination of H2O2 under Cd stress. Prodanovic et al., (2012) reported that POD activity was also shown to increase during late germination and early seedling growth in some annual herbaceous species like Chenopodium rubrum (Ducic et al., 2003) and tomato (Morohashi., 2002), biannual species like Brassica oleracea (Bellani et al., 2002), and perennial species like Viola carnuta (Mitchell and Barrett, 2000). Under stress conditions, combined action of enzymatic antioxidants is critical for mitigating the damages induced by ROS (Salin, 1988; Shanker et al., 2004). The importance of antioxidants in preventing oxidative stress in plants is based on the fact that level of one or more antioxidants increased under stress which is generally related with the increased stress tolerance (Gangwar et al., 2012). According to Shanker et al. (2005), the increase in antioxidant enzymes observed might have been in direct response to the generation of superoxide radical by Cr-induced blockage of the electron transport chain in the mitochondria .Activity of peroxidase in general increased with the increasing treatment of chromium in 144 DISCUSSION radish and in Pisum sativum L. Seedlings (Gangwar et al., 2012) as shown in Fig. 43 and 51. Cadmium chloride treatments effectively increased the activity of the catalase enzyme in pea similar to B. rapa leaves (Shaimma et al., 2012) as shown in Fig. 26 and 34. CAT activity increased as heavy metals concentration increased and decreased at higher concentration for long-term exposure (Arleta et al., 2001; Salama et al., 2009; Liu et al., 2011). The increase in CAT activity after Cd treatments may be due to the scavenging role of CAT to H2O2, which could be quenched by the induction of specific enzymes like CAT (Elstner et al., 1988). CAT is one of the major antioxidant enzymes that eliminate hydrogen peroxide by converting it into oxygen and water (Miller et al., 2008). Many reports indicated that CAT activity was significantly influenced by Cadmium stress. This recommended that CAT activity, plays a very important role in the protection against oxidative damage caused by cadmium (Scebba et al., 2006; Mobin and Khan, 2007; Zhang et al., 2009; Foroozesh et al., 2012). The effects of cadmium on catalase activity were assessed and the obtained results were found as shown in Fig. 26 and 34, similar to Foroozesh et al. (2012). Katazyna and Smolik (2011) reported that applied doses of cadmium (0.025-5 mM) caused a significant increase in the activity of catalase in all the growth phases of wheat. Amongst various enzymes involved in the antioxidant metabolism of ROS, the GST, CAT and GR activities was found higher at all the Cr (VI) concentrations than their controls (Malmir, 2011) as shown in Fig. 43 and 51. The ROS generation during the oxidative damage results in the release of hydrogen from unsaturated fatty acids that, eventually elevates the aldehyde and lipid radicals inside the cell that cause the membrane damage by disrupting the lipid bilayer (Reinheckel et al., 1998). As a result there is a marked increase in the level of cellular antioxidants for scavenging the free radicals formed during the metal stress. This indicates that the plants have developed an efficient defense system to cope with the metal stress (Shanmugaraj et al., 2013). Shafi et al. (2010) reported that MDA content was increased under salt and Cd stress in wheat plants Soybean (Aksoy and Dinler, 2012) as the shown in Fig. 26 and 34. Although Cd does not generate ROS directly, it generates oxidative stress via interference with the antioxidant defense system (Shi et al., 2010) and increases MDA content in plants due to increased lipid peroxidation (Zhang et al., 2009). An increase in the level of MDA was also reported in safflower 145 DISCUSSION (Ahmed et al. 2010), barley (Hegedus et al., 2001) and in Wolffia arrhiza (Piotrowska et al., 2010) during heavy metal treatment. Hegedus et al. (2001) mentioned Cd toxicity also increased MDA content of barley plants. Panda et al. (2003), Dey et al. (2007), Malmir (2011) reported that compared to control, concentration of MDA was found increased in Cr (VI) concentrations in the roots, stem and leaves of the Cr (VI) treated sorghum plants.MDA was found increased with Cr (VI) concentrations in the roots, stem and leaves of the Cr (VI) treated sorghum plants (Malmir, 2011), as the shown in Fig. 43 and 52. Panda et al. (2003) also reported increase in MDA in the metal treated Triticum aestivumplants. Plants have shown proline accumulation under environmental stress (Ahmad and John, 2005; Ahmad et al., 2006; Ahmad et al., 2008). It has been often suggested that proline accumulation may contribute to osmotic adjustment at the cellular level and enzyme protection stabilizes the structure of macromolecules and organelles. Increase in proline content may be either due to de-novo synthesis or decreased degradation or both (Kasai et al., 1998). Proline accumulation in B. juncea, Triticum aestivum and Vigna radiata in response to Cd++ toxicity has been demonstrated by Dhir et al. (2004) and in chickpea (Faizan et al., 2011). Arabidopsis plants, proline content was increased under combined of Cd and salt stress (Xu et al., 2010) and in soybean (Aksoy and Dinler, 2012). During stress, proline accumulates in the cell as a result of water deficit (Nikolic et al., 2008) in order to protect the cell from osmotic damage (John et al., 2007) and also it involves in detoxification mechanism by binding to cadmium ions, converting them into non-toxic forms (Sharma et al., 1998). It was also suggested that proline accumulation may also be due to the reduced electron transport process in mitochondria (Saradhi et al., 1995). The increase in proline content is associated with osmotic adjustment, water balance maintenance, ROS scavenging activity thereby providing the better tolerance against the stress induced oxidative damage (Siripornadulsil et al., 2002). Shanmugaraj et al. (2013) reported that proline content was increased in all the parts of the plant uniformly in Brassica juncea. Liu et al. (2011) had reported increased levels of proline in Lonicera japonica, Bassi and Sharma (1993) in wheat and Rastgoo and Alemzadeh (2011), in Aeluropus littoralis in response to different metal treatments as shown in Fig. 35. Proline content increased in tea plant with increasing concentration of Cr (Tang et al., 2012). Increase 146 DISCUSSION of proline was occurred due to increase of chromium in root and aerial part of Brassica napus L. plant. Increase of proline in plants is a defense mechanism (Najafian et al., 2012) as shown in Fig. 52. 5.5 Amylase Seed germination relies almost exclusively on seed reserves for the supply of metabolites for respiration as well as other anabolic reactions. Starch is quantitatively the most abundant storage material in seeds and available evidence indicates that in germinating seed starch is degraded predominantly via the amylolytic pathway (Juliano and Varner, 1969). The average total amylolytic, α amylase and β amylase activities were significantly depressed by the higher concentrations of Cd as shown in Fig. 27. Amylase activity was the most sensitive parameter, α amylase is the major enzyme involved in the initial degradation of starch into more soluble forms while phosphorylase and β amylase assist in the further conversion to free sugars which affords the nutrition of seed germination (Juliano and Varner, 1969). Reduction of amylase activity may therefore be the major factor involved inthe depression of seed germination amyase activity decrease in wheat with increase Cd (Amirjani, 2012). Amylase depressed with increasing concentration of Cd in pea seeds. No remarkable change in amylase activity was observed at lesser Cr supply except a decrease in activity at greater Cr levels (Sharma et al., 2003), as shown in Fig. 44. 5.6 Biomass Shanmugaraj et al. (2013) reported that the overall growth of the plant biomass is commonly attributed by the accumulation of biomass and moisture content of the plant as shown in Fig. 24, 31 and 32. The biomass reduction is a cumulative effect of reduction in moisture content, reduced water and mineral uptake in Brassica juncea. Bhardwaj et al. (2009) in their investigation on the bean plant observed a reduction in biomass. The reduction in fresh weight may be referred to toxicity of CdCl2, thereby this toxic material can breakdown normal physiological mechanisms and finally have negative influences on biomass. Bahmani et al. (2012) reported that Mean fresh weight under Cd treatment reduced as comparison with control in Phaseolus vulgaris L. The plant processes during early growth and development culminates in total dry matter as a consequence of poor production, translocation and partitioning of assimilates to the economic parts of the plant. The negative effect on 147 DISCUSSION dry matter acquisition may be essentially an indirect effect of Cr on plants. The overall adverse effect of Cr on growth and development of plants could be serious impairment of uptake of mineral nutrients and water leading to deficiency in the shoot. Chen et al. (2001) reported that the total root weight and root length of wheat was affected by 20 mg Cr (VI)/kg soil as K2Cr2O7 as compare to control plants. Vajpayee et al. (2001) studied that the Cr accumulation and toxicity in relation to biomass production found that dry matter production was severely affected by Cr concentrations in Vallisneria spiralis as shown in Fig. 41, 48 and 49. Singh (2001) in a study on the effect of Cr (III) and Cr (VI) on spinach reported that Cr applied at 60 mg kg-1 and higher levels reduced the leaf size, caused burning of leaf tips or margin and slowed leaf growth rate resulting in reduced dry biomass. Vernay et al. (2008) reported that D. innoxia plants grown in presence of Cr(VI) showed reduced growth leading to reduction in root and shoot biomass in rice (Ahmad, 2011). Sinha et al. (2005) reported the restricted biomass in cauliflower with chromium. The moisture content (%) decreased upon treat with CdCl2 in Brassica juncea (Shanmugaraj et al., 2013). 5.7 Protein The reduction in total soluble protein under CdCl2 stress could be due to protein reaction with oxygen free radicals resulting in a change in specific amino acid, polypeptide splitting and increased susceptibility of protein to proteolysis as shown in Fig. 28. The functionality of protein can also be affected by ROS either by oxidation of amino acidside chains or by secondary reaction with aldehydes products of lipid peroxidation (Romero-Puertas et al., 2002, Ayoughi et al., 2011). Protein decline have been reported in Brassica juncea (Mobin and Khan, 2007). A reduction in protein content was observed in all cultivars treated with different CdCl2 concentration over controls in Brassica napus (Touiserkani and Haddad, 2012). The protein content in the leaves of wheat plants decreased with increase in Cr which may be due to decrease in the nitrogen content, nitrogen is the precursor for the synthesis of amino acids which are the building blocks of protein in case of rice plants (Nag et al., 1981) and also in Triticum aestivum L. (Datta et al., 2011) as shown in Fig. 45. 148 DISCUSSION 5.8 Sugars Tandon and Gupta (2002) have also reported decrease sugar contents at increased doses of heavy metals as shown in Fig. 28, 35, 45 and 53. In many plant species, the accumulation of soluble sugars has been observed in response to various environmental stresses. Total carbohydrate found inhibited if cadmium concentration applied more than 5 kg-1 soil (Saleh and Al-Garni, 2006). 5.9 Harvest Index The harvest index value significantly decreased with increase of cadmium and chromium over control (Fig. 37 and 56). Higher harvest index indicates higher translocation ability from source to sink. The decrease in shoot dry weight registered under Cd stress was concomitant with a significant reduction in both length and diameter of the internodes as well as in leaf area and thickness (Djebali et al., 2010). Shoot and root biomasses of corn and wheat decreased with the increase in Cd concentrations in the solutions (Zhao, 2011). Reduction in pod setting may be formation of non functional flower possibly at fag end of flowering, which abscised without being converted into pods (Dhingra, 2002). Manisha and Dhingra (2003) indicated that the reduction in seed yield in pea has been associated with decline in number of flower, number of seed and seed size. Biological and harvest index of plant was decreased with increasing concentration of cadmium in term of reduction in number of pod and leaf, root and stem growth (Siddhu et al., 2008, 2012). Physiology of seed and fruit yield has been affected by cadmium in term of decrease in number of seed/fruit, weight of 1000 seeds, number and fresh weight of fruit /plant (Masih et al., 2004). Root and shoot dry weight were also significantly decreased with increasing concentration of chromium (Ghani, 2011). The weight of dry root and aerial parts of Brassica napus L. decreased with increasing concentration of chromium (Najafian et al., 2012). 149