The physiological response of the common reed plants to As toxicity and its possible relationships with the growth parameters and nutrient composition of the plants were evaluated through a PCA. The analysis resulted in a total of eight components, of which the first four explained 75.5% of the variance. The first component related positively the photosynthetic pigments levels in the leaves with plant yield,height, and P mass fraction,as well as with oxidative stress parameters in the roots and in the aerial part of the plants. This component therefore shows that the growth and photosynthetic system of the common reed plants were not affected by the increasing presence of As in the nutrient solution, likely as a consequence of the formation of those compounds that acted as an efficient defense mechanism against oxidative stress. The second component related the different As species determined in the plants positively with each other, and negatively with Cu and Zn in the aerial part of the plants and with P and K in the roots. This confirms that As accumulation did not affect the plant’s photosynthetic and oxidative status, but may have limited macro and micro-nutrients uptake in the roots and their transport to the aerial part of the plants. The third component simply related the values of Cu, Fe, Mn, and Zn in the roots,while the fourth component related the levels of Mn and Zn in the aerial part of the plants negatively with those of MDA and proline in the aerial part and with that of N in the roots. The latter component seems to relate the deficiency of certain nutrients, likely a consequence of As uptake and accumulation in the plants,with the formation of MDA and proline in the plants, which is also indicative of As toxicity.
When the regression factors generated in the PCA were plotted,the highest As dose was clearly differentiated from the rest of the treatments, linked to a high accumulation of As in the roots. The control replicates grouped together opposite those of treatment T10, associated with the levels of nutrients in the plants,hydroponic nft system while the rest of the treatments showed an elevated variability and no clear or consistent relationships with any of the parameters determined in the plants.Malachite Green,a triphenylmethane dye, is a multipleuse compound that is mainly used in textile industries and partly used in aquaculture in fungicides and ectoparasiticides. While the effects of MG on aquatic invertebrates and algae have been scarcely elucidated,Hidayah et al. reported that MG in wastewater from either industry or aquaculture has been widely reported to be toxic to many kinds of fish with lethal effects at a concentration of less than 1 mg/L, with the dye and its derivatives being accumulated in aquaculture products such as fish, prawn and crab. It also possesses carcinogenic and genotoxic properties which pose a potential risk to humans and therefore, this dye has been banned in Europe, the USA and several countries. However, MG is still being used in some parts of the world because it is highly effective and easily available at low cost. It is also used domestically as a treatment for diseases of tropical fish and can be readily obtained by the public ; hence, concern about its illegal use exists. In Asian countries such as Bangladesh, MG has been reported to be used for the eradication of external parasites and fungal diseases in fish farming. However, removal of MG from aquaculture wastewater has received little or no attention compared to other pollutants. Consequently, contamination of MG in aquaculture waste could be expected with harmful consequences to the surrounding environment. Effluents from aquaculture usually contain high amounts of nutrients such as nitrogen ,phosphorus and organic compounds that either potentially cause algal bloom in receiving water or, if high enough, can support vegetable production.
To reduce water pollution problems, fishery industries in many countries including Thailand have been forced to treat their effluent in proper ways such as by the rational use of water and by the recovery of substances from wastewater. Hence, effluents from aquaculture have been used for garden applications or the production of hydroponic plants as a secondary treatment in the waste management procedure. In some management practices, such as the study by Somboonchai and Chaibu,four vegetables were grown in a hydroponic system integrated with catfish culture. However, the effluents from aquaculture such as shrimp farming contain not only nutrients but also other chemical substances such as antibiotics, herbicides and fungicides that potentially impact on the environment. A review by Carvalho et al. indicated that pharmaceutical products, including antibiotics, hormones, analgesics and anti-inflammatory drugs, chemical compounds used for disinfection and cleaning, and endocrinedisrupting compounds can be assimilated by the plants. Therefore, while the potential for biomass production and nutrients recovery from wastewater are primary concerns in wastewater management systems,bioaccumulation of toxic substances is another aspect of concern. Additionally, increasing water scarcity in either dry regions of the world or in developing countries makes the reuse of wastewater in agriculture more important. Nevertheless, it is of interest to identify whether or not a practice is productive and safe for both the environment and human health. Several plant species can tolerate toxic substances by accumulating them in non-toxic forms or transforming them to either nontoxic or less toxic products. Most studies showed that textile dyes can be either adsorbed and accumulated or transformed to less or non-toxic substances by detoxifying enzymes, predominantly peroxidase, in plant cells. The dye MG was found to be transformed to 4-dimethylamino-cyclohexa2,4 dienone in Blumea malcolmii Hook. using enzyme laccase and the products had less toxicity toward Phaseolous mungo and Triticum aestivum when tested. Rai et al. found that biodegradation of MG by Aloe barbadensis resulted in nontoxic metabolites, suggesting the possibility of using treated, dye wastewater for irrigation. Torbati reported that activity of antioxidative enzymes, namely SOD, POD and CAT, in Spirodela polyrhiza L. was increased with increased MG in the bathing medium.
The activity of these enzymes allowed the species to tolerate MG at concentrations of 10 mg/L and 20 mg/L. However, knowledge on the degradation of synthetic dyes by vegetable plants is scarce since phytotransformation has been studied mainly in non-edible plants. Nowadays, the trend toward eco-friendly and sustainable production of any kind of product strongly influences consumers. The current study investigated the application of wastewater containing MG from aquaculture for the production of pak choy,a vegetable that is produced commercially in many Asian countries. It was hypothesized that being a member of the genus Brassica whose species usually have high antioxidant enzyme activity upon exposure to toxic substances,B. chinensis may have ability to degrade MG dye and, hence, tolerate the dye at the low concentration used in aquaculture. If so, the reuse of water contaminated with MG could be applied. However, some Brassica species such as cabbage and Wisconsin fast plants could take up and accumulate some toxic substances in their tissue, especially in the roots. Therefore, on the other hand, the MG dye in water may be accumulated in plant tissue and inhibit growth of the plant. Thus, the aims of this study were: 1) to study the effects of MG on the growth of B. chinensis and 2) to evaluate the accumulation of toxic substances in the edible parts of B. chinensis. The findings from this study will be useful for consideration in a wastewater management strategy, particularly for the reuse of aquaculture wastewater in crop irrigation.When the seedlings were age 7 d, the nutrient mixed solution was applied replacing water, and the seedlings were allowed to grow under ambient conditions to age 14 d before being transferred to the growth medium used in the growth experiment. Fourteen-day-old seedlings with 3e5 leaves and an average height of 8 cm were selected for the growth experiment. The seedlings were grown in nutrient mixed solution for 1 wk to allow for acclimation to the hydroponic growth conditions. The nutrient mixed solution was prepared from tap water and 1 ml/L of commercial A and B nutrient solution for hydroponic planting. The pH and electrical conductivity of the nutrient mixed solution were monitored and maintained at 6.0e6.5 and 1.5e2.5 ms/cm, respectively.After 1 wk acclimation, 48 seedlings were distributed to four levels of MG concentration treatments: 0 mg/L,1 mg/L, 2 mg/L and 4 mg/L. The basal part of each seedling was fitted in a small plastic basket to hold the plant in an upright position and the baskets were fixed on the lids of 5 L plastic tub containers. One container with 12 seedlings was used for each treatment. The chemical formula of the MG used was C23H25N2Cl. The concentration of each treatment was obtained by adding the appropriate volume to make up 500 mg/L of MG stock solution to the nutrient mixed solution which hereafter is called the growth solution.
The pH and EC of the growth solution were monitored and maintained as mentioned above and the growth solutions were renewed weekly. The experiment was maintained under ambient conditions with the air temperature 24e29 C,nft channel relative humidity 41e60% and natural sunlight. Decolorization of MG in the growth solution at day 7 was detected spectrophotometrically using an ultravioletevisible spectrophotometer. The solution from each treatment was sampled and measured for absorbance at 400e800 nm compared with the absorbance of freshly prepared solution at the same concentration. After 4 wk of growing, growth parameters were measured. Then, all plants were harvested and each plant was separated into root and shoot parts, and the shoots were stored at 70 C in a freezer for further tissue analysis. The roots were abandoned since it is normally a non-used part of this vegetable and it was impossible to separate the plant roots from the supporting sponge. The weight of the shoot was measured after drying in a hot-air oven at 60 C for 48 h and the final dry mass was determined. The experiment was conducted between February and March.The effects of MG on plant species have been mostly tested using seed germination and the plant seedling stage, with germination and seedling development being generally inhibited. The current study found that MG contamination in water also caused negative effects to B. chinensis, particularly at concentrations greater than 1 mg/L. From the results, the negative effects of MG were strongly evidenced on root growth which was reduced by 50% upon exposure to MG of 2 mg/L and 4 mg/L compared to the control or 1 mg/L MG treatments. Similar effects on root growth were found in Arabidopsis thaliana grown on medium supplied with 4 mg/L of Crystal Violet and MG. The stunted roots may contribute to the overall reduction in plant growth since the uptake of water and nutrients could occur only via root transport under the hydroponic growth conditions used in this study. Nevertheless, the overall growth of B. chinensis in the current study indicated that the plant tolerates MG at a concentration of 1 mg/L. MG has been suggested to be toxic to plants as it could be strongly absorbed on the surface of cellulose and taken up through the roots and accumulated in plant tissues. Saranya et al. found that the chlorophyll contents in Hydrilla verticillata decreased with increasing Basic Violet 14 dye concentrations from 5 mg/L to 25 mg/ L, although the difference was not significant at 5 mg/L and 10 mg/L, and this result supported the inhibition of the dye on chlorophyll biosynthesis. In the current study, there was no evidence of chlorosis in the B. chinensis leaves at the concentrations applied. However, the effect of MG on chlorophyll biosynthesis in this plant species should be better explained by pigment analysis. Dye contamination in either the water or soil usually causes a reduction in the total content of macromolecules such as proteins and carbohydrates whilst it usually induces the activity of several enzymes used for dye degradation in exposed organisms. Triphenylmethane dyes such as Crystal Violet and Methyl Violet could cause lower protein synthesis which consequently inhibited cell growth in Bacillus subtilis. Jayanthy et al. found that soil contaminated with dyes from dyeing industries caused decreasing protein, total free amino acid and carbohydrate contents in Vigna radiata, whilst in the same plant, there were increases in the proline, glutathione and methyl glyoxal contents in either leaf or root tissue which indicated a response to abiotic stress. Moreover, the activities of lignin peroxidase, veratryl alcohol oxidase, laccase, tyrosinase and DCIP reductase were induced in Aster amellus Linn. and Glandularia pulchella Tronc. upon exposure to the dye Remazol Orange 3R.