The root length and shoot length of paddy were measured 120 days after sowing by using a centimetre scale and it is expressed in cm/plant.

The length and breadth of each and every leaf was measured and the total leaf area was calculated by using the formula L × B × K and it is expressed in cm²/plant, where, L and B are the length and breadth, respectively, and K is the Kemp's constant, (for Monocot 0.9).

The collected plant samples were washed thoroughly and separated into root and shoot. Their fresh weight was measured using an electronic balance. Then, the samples were kept in a hot air oven at 80 °C for 24 hours, and their dry weight measured using an electrical single pan balance.

The yield parameters of 1000 grains weight and yield/ha were taken and recorded for both the treated and control plants.

Two hundred milligrams of dried powdered plant sample was taken in a 100 ml Kjeldahl flask. Two hundred milligrams of salt mixture (potassium sulphate, cupric sulphate and selenium powder mixed in the ratio of 50:10:1) and 3 ml of concentrated sulphuric acid was added. After digestion, 10 ml of distilled water was added and cooled. The diluted sample was decanted into the micro-Kjeldahl distillation flask. To that, 10 ml of 40% sodium hydroxide was added and distilled. The distillate was collected in a conical flask containing 10 ml of 4% boric acid and three drops of mixed indicator (0.3 g bromocresol green and 0.2 g methyl red in 400 ml of 90% ethanol). This solution was titrated against 0.05 N HCl. Nitrogen content was calculated and recorded.

One gram of dried powdered plant material was digested with 10 ml of acid mixture (nitric acid 750 ml, sulphuric acid 150 ml and perchloric acid 300 ml). The digest was cooled and made up to 50 ml and filtered. One millilitre of the digest was mixed with 2 ml of 2 N nitric acid and diluted to 8 ml. One millilitre of molybdovanadate reagent (25 g of ammonium molybdate in 500 ml of water, 1.25 g of ammonium vanadate in 500 ml of 1 N nitric acid, both were mixed in equal volume) was added, shaken and the absorbance was measured after 20 min at 420 nm in UV-Spectrophotometer. A calibration curve was prepared using potassium dihydrogen phosphate as standard.

Dried and ground tissue of paddy (0.5 g) were digested in a 100 ml Kjeldahl flask using 10 ml of concentrated nitric acid, 0.5 ml of 60% perchloric acid and 0.5 ml of sulphuric acid. The inorganic residue was cooled and diluted with 15 ml of distilled water and filtered through Whatmann No. 42 filter paper. The filtrate was made up to 50 ml with distilled water. The filtrate was used for potassium estimation by Flame photometer and standards were prepared with potassium chloride.

One millilitre of sulphuric acid and 15 ml of double distilled water were added to a Kjeldahl flask containing 0.5 g of dried and powdered material was incubated overnight at 80 °C. After that, 5 ml of acid mixture (nitric acid and perchloric acid in the ratio of 3:1) was added and then digested. The digested material was cooled, made up to 50 ml and filtered through Whatmann No. 42 filter paper. The sample was aspirated to an Atomic Absorption Spectrophotometer with air/acetylene flame and the readings were taken for manganese (530 nm), copper (324.6 nm), zinc (214 nm) and iron (568 nm).

Five hundred milligrams of the dried sample was digested in an acid mixture (HNO₃ and HCl), filtered and brought up to 100 ml with distilled water. To 25 ml of the solution, 5 ml of diphenylcarbazide reagent was added and made to 50 ml. The solution was read in an Atomic Absorption Spectrophotometer at 540 nm. A standard curve was prepared by using a known amount of chromium.

Experimental results presented are the mean values of the experiments done in triplicates. The data was statistically analysed using a two-tailed T-test (SPSS 14.0) to compare paired means between different chromium treatments at 5% and 1% level of probability.

The root length and shoot length of paddy decreased gradually with the increase of chromium concentrations (2.5, 5, 10, 25, 50, 75 and 100 mg/l). The root length of paddy varied from 19.52 to 10.76 cm/plant respectively between the control and treatments (200 mg/l). Similarly, the shoot length significantly varied from 107.2 to 46.5 cm/plant between control and the treatments (200 mg/l) respectively.

The depression in growth was more marked at the 200 mg/l concentration of chromium. The general response of decreased root growth due to chromium toxicity could be due to inhibition of root cell division or cell elongation or combination of both to the extension of cell cycle in the roots. It resulted in the limited exploration of the soil volume, uptake and translocation of nutrients and water, and induced mineral deficiency in plants [31]. It has been already reported that chromium causes retardation of growth up to 80% in both the root and shoot of tomato and brinjal [9]. The plant height reduced significantly in wheat cv. UP 2003 grown in sand with 0.5 M sodium dichromate [32]. The reduction in plant height might be mainly due to the reduced root growth and consequent less nutrients and water transport to the aerial parts of the plant.

The total leaf area is a component of length, breadth and number of leaves, which reflects in the process of photosynthesis. The highest total leaf area was recorded in control plants and the lowest total leaf area was recorded in chromium-treated plants. The total leaf area gradually decreased with the increase in 200 mg/l chromium concentrations. The total leaf area (970.35 and 288.10 cm²/plant) varied significantly from control plant to 200 mg/l concentrations of chromium treated crop. A steady decrease in leaf area at higher concentrations can be attributed either to reduction in the number of cells in the leaves stunted by salinisation or reduction in cell size [33]. The reduced cell size and decreased intercellular spaces responsible for reduction in leaf area. The variation in size of areoles is also another factor which is responsible for reduction in total leaf area by chromium treatment [34,9].

The fresh weight and dry weight of crops are mainly based on their growth performance of a particular crop. Growth directly influences the weight of plants. The highest dry weight (5.116 g/plant) in shoot of paddy was registered in control and the lowest dry weight (0.968 g/plant) was registered in 200 mg/l chromium treated paddy root. The total dry weight of root and shoot of paddy decreased gradually with the increase in chromium concentrations. It was mainly due to the inhibition of water uptake and enlargement of root cells. There was a distinct reduction in dry biomass at flowering stage in white mustard when Cr(VI) was given at the rates of 200 or 400 mg/kg soil [35,36]. The decreased biomass might also be due to disturbed carbohydrate and nitrogen metabolisms and reduction in protein synthesis or low photosynthetic reactions under excess Cr conditions [35,37].

Most physiological and biochemical processes are severely affected by chromium and, as a consequence, the yield of crop are equally affected [38]. The highest 1000 seed weight (24.78 g/plant) and yield (8035 kg/ha) were recorded in the control plants. The lowest seed weight (11.0 g/plant) and yield (1735 kg/ha) were recorded in the crop irrigated with 200 mg/l of chromium.

The increase in chromium supply from 2.5 to 200 mg/l, markedly decreased the grain weight and their economic yield (kg/ha). An almost 80% reduction in grain weight was observed at 200 mg/l chromium when compared to control plants. The depression in grain yield as a result of differential chromium supply was very high and ranged between 82% and 92% from 0.05 to 1 mM Cr [38]. Nitrogen, one of the primary essential nutrients involved as a constituent of biomolecules such as nucleic acids, nitrogen bases, coenzymes and proteins. Any deviation in nitrogen content would severely inhibit the growth and yield of plants [39].

Nitrogen is an important constituent of protein and protoplasm and in many essential compounds. Nitrogen content in shoot and root showed a decreasing trend with increasing concentrations (2.5, 5, 10, 25, 50, 75 and 100 mg/l) of chromium (Fig. 1). The highest nitrogen content in shoot (192.3 mg/g dry wt.) and root (176.0 mg/g dry wt.) were recorded in control plants. Similarly, the lowest nitrogen content in shoot (130.0 mg/g dry wt.) and root (126.0 mg/g dry wt.) were registered in the 200 mg/l chromium-treated crop. Nitrogen content of paddy decreased significantly with the increase of chromium concentration. Similar inhibition of nitrogen metabolism due to excess of metals was also observed [42]. Chromium treatment affects nitrogen uptake and assimilation, which is evident from the decline in the concentration of nitrogen in wheat leaves. The uptake of nitrogen from soil was also inhibited by the presence of heavy metals [43].

Phosphorus forms an important component of nuclear protein. A decrease in phosphorus content of paddy shoot and root were observed with increasing concentrations of chromium (Fig. 2). Phosphorus content of root and shoot of paddy varied significantly with chromium treatments. Similar differences in phosphorus accumulation due to metal stress were reported in soybean [44], greengram [45], groundnut [46] and Convolvulus arvensis [47]. The reduction in phosphorus content in the paddy may be due to: (i) blockage in mobilization of phosphorus from soil and root system to the aerial system due to chromium [48]; (ii) the toxic effects of chromium on root cells which impaired phosphorus absorption process, or to decrease availability of phosphorus with the addition of chromium in soil [49]; and (iii) chromium can competitively inhibit either the uptake or transport of phosphorus by the roots [50].

Potassium is essential for the formation of sugars and starch absorbed by the root and is generally transported to the shoot and this transport seems to be controlled by shoot growth [51]. The content of potassium in root and shoot of paddy is gradually decreased with the increasing concentrations of chromium (Fig. 3). The same pattern of response of heavy metal in potassium uptake was also observed in Trifolium repens [52], in Pisum sativum [53], in corn [54] and in Oryza sativa [55]. Changes in the conducting tissue and blockage of the xylem elements may also contribute to a limited translocation of potassium from the root. It may be due to competition of chromium ions with potassium, which in turn exercised a regulatory control on potassium uptake [56].

Manganese functions as an enzyme activator in several reactions of respiration and nitrogen metabolism. The chromium concentration markedly decreased the manganese content of root and shoot of paddy. The higher concentration of chromium (200 mg/l) treated paddy showed lower manganese content (Fig. 4). The chromium-induced manganese deficiencies are comparable to the works of Wallace and Abau-Zam Zam [57]. The decrease in manganese content may be due to the competition of chromium with manganese for transport sites in plasma lemma. The increase in chromium concentration might have displaced manganese from the absorption sites in cell walls [58,59].

Copper is required in very small quantity and acts as catalyst in oxidation-reduction reaction. The copper content of root and shoot of paddy decreased with a steady increase in chromium concentration (Fig. 5). The higher copper content was recorded in control sets while the lowest content was observed in paddy irrigated with 200 mg/l concentration of chromium. The results are in conformity with the reports of Gardea-Torresday et al. [47]. The decrease in copper content may be due, to some extent, to the close association of copper with nitrogen ligands and the close parallel relation in the movement of copper and nitrogen out of the leaves [60,61].

Zinc functions as activator of certain enzymes and it is essential for the biosynthesis of plant auxin Indole-3-acetic acid. The zinc content of root and shoot of paddy decreased gradually with concomitant increase in chromium concentration (Fig. 6). The highest zinc content was recorded in control and the lowest content was recorded in 200 mg/l chromium treated crop. The paddy crop showed marked differences in their ability to absorb, translocate and accumulate zinc in various parts and this can be compared with the findings of Gardea-Torresday et al. [47]. The reduction in zinc content to due metal stress was also observed in raddish [62], blackgram [63] and Convolvulus arvensis [47]. The zinc concentration in the root was found to be lower when compared with the shoot. Hence, it is noteworthy that zinc uptake was affected by the chromium treatments but not the translocation of zinc within the plant [40]. Plants subjected to zinc deficiency were reported to undergo changes in diverse metabolism [64].

Iron is essential for the synthesis of chlorophyll and also plays an important role in respiratory mechanism. Iron content of root and shoot of paddy varied significantly with different concentrations of chromium. There was a gradual decline in iron content with increase in chromium concentrations. The iron content of root is found to be higher than shoot (Fig. 7). Chromium can substitute to some extent for iron in biological system [65]. These findings agreed with the results in white beans [65], in tomato [66]. The reduction in iron content may be due to: (i) the increasing concentration of chromium leading to iron deficiency, because of the competitive occupation of active sites of iron [67]: (ii) the chromium may be competing with iron, which leads to the lowered translocation of iron from the roots to the shoots that resulted in chlorosis [68], and (iii) the reduction in the uptake of iron could be mainly due to the chemical similarity of these ions in solution.

Chromium uptake by plants is mainly nonspecific probably as a result of uptake of essential nutrients and water. The chromium accumulation in the paddy root and shoot varied significantly due to chromium treatment. The accumulation of chromium increased positively with the concentrations (2.5 to 200 mg/l). The accumulation of chromium in root and shoot of paddy are gradually increased with the increase in chromium concentrations (Fig. 8). It has been reported that translocation of chromium from roots to tops was inhibited in the presence of toxic levels of the metals [33]. Thus, major portion of the element remained in the roots and only a small part was translocated to tops due to metal stress. It has been already reported that Cr distribution in crops had a stable character which did not depend on soil properties and concentration of this element. The maximum quantity of element contaminant was always found in roots and a minimum in the vegetative and reproductive organs [69] reported that Cr was poorly taken up into the aerial tissues, but was held predominantly in the root. The reason of the high accumulation in roots of the plants could be because chromium is immobilized in the vacuoles is less toxic, which may be a natural toxicity response of the plant [70].

In this investigation, addition of Cr (VI) led to the reduction in the levels of nutrients in root and shoot of paddy. It indicates that the Cr (VI) toxicity causes malfunction in the uptake of mineral nutrient, which leads to mineral nutrition deficiency [71].

The data for the uptake and accumulation of chromium using water plants are presented in Fig. 9. In this experiment, certain aquatic macrophytes such as Eichhornia crassipes, Pistia stratiotes and Salvinia natans were employed in the biological treatment. They were allowed to grow in chromium-contaminated water (100 mg/l) for 30 days. As a result, the roots of Eichhornia accumulated 75% chromium than in the leaves (60%). Similarly, lower chromium content was recorded in Pistia root (65%) and leaves (58%) than the Eichhornia. Salvinia accumulate 60% chromium and 55% in root and leaves from the chromium contaminated water. The accumulation of chromium increased with the number of days exposed. Among the plants, Eichhornia accumulated more chromium than the other plants tested. Water hyacinth (Eichhornia crassipes) is a flowering macrophyte whose appetite for uptake of nutrients and explosive growth rate has been put to use in cleaning up municipal and agricultural wastewater. The macrophytic species such as Potamogeton crisspus, Trapa bispinosa, Vallisneria spirallis Eichhornia crassipes, Azolla pinnata and Lemna minor [72,73] have been already reported in the removal of chromium from the aquatic medium.

The data related to uptake and accumulation of chromium using grasses and weeds are given in Fig. 10. The phytoremediation performance of a plant is not only determined by its ability to achieve high metal concentrations within its tissues, but also by its ability to translocate the metal to aerial organs and to produce a high biomass. The efficiency of phytoextraction for remediation of heavy metals from contaminated soil is highly dependent on the bioavailability of metals. In this experiment, two grasses (Cyperus rotundus, Cyperus kylinga)and two weeds (Marselia quadrifolia, Ludwigia parviflora) were used for the phytoremediation study. All the species were allowed to grow up to 30 days in 100 mg/l chromium contaminated soil. These plants were harvested periodically (5, 10, 20 and 30 DAS) and separated into root and shoot to determine the chromium accumulation. The higher accumulation of chromium was recorded in the root of Cyperus rotundus (275.5 μg/g), Ludwigea parviflora (242.0 μg/g), Cyperus kylinga (251.6 μg/g) and Marsilea quadrifolia (214.0 μg/g) than in shoot (248.4, 212.0, 182.0 and 174.50 μg/g) at 30 days of exposure. Among the plants tested, the grass Cyperus rotundus and Ludwigea sp. accumulated more amount of chromium from contaminated soil followed by Cyperus kylinga and Marsilea sp. because of their natural physiological and biochemical abilities. The root tissues accumulate significantly greater concentrations of metals than shoots including high plant availability of the substrate metals as well as its limited mobility once inside the plant [30]. Moreover, a 60% reduction of chromium from the contaminated soil was recorded by using the plants such as Cyperus rotundus and Ludwigea sp. Plants growing in polluted soil absorb nourishments from polluted soil and entry of pollutants to the plant system during the process of absorption may not be ruled out. It was assumed that uptake of pollutants by plants may cause the lowering of pollution load of the soil.

Brassica juncea and B. nigra have a better ability to take up heavy metals from soil substrates and to transport and concentrate these metals in their shoots. The plants for the phytoextraction process should tolerate and accumulate high levels of heavy metals in its harvestable parts. It must have a rapid growth rate and a potentiality to produce a high biomass in the field [74].

Besides hyperaccumulator plant species like Salix viminalis, Brassica juncea, Lolium perenne, Zea mays and Helianthus annuus, characterized by high content of heavy metals in biomass and have good remediation capacity [25].

The naturally growing dominant herbaceous species can also be used for this purpose because they have the adaptability to survive under stress of industrial effluents. The dominance of herbaceous flora in contaminated soil may be due to the fact that shrubs are more stable and less influenced by any small environmental conditions. The plants must have the ability to translocate chromium from the root to the shoot in order to continuous absorption of chromium from substrates. It can reduce chromium concentration and thus reduce toxic potential to the root and translocation to the shoot is one of the mechanisms of resistance to high Cr concentration [75]. The mechanisms involved in heavy metal tolerance may range from exclusion, inclusion and accumulation of heavy metals depending on the plant species [76,77]. It was found to be economically valuable to grow naturally growing weeds and grasses in order to soil effectively at larger scale in shorter time [78].