CSA PAKISTAN

Determination of phytotoxicity threshold on wheat crop by using biogenic zinc, copper nanoparticles

Muhammad Waseem1, Muhammad Qasim1, Qudrat Ullah1, Syed Raza Hussain1, Irfan Haidri1, Memoona1,  Sikander Hayat2, Usman Shafqat1*

1Department of Environmental Sciences, Government College University, Faisalabad 38000, Pakistan

2Fodder Research Institute

Abstract

Nanotechnology is the 5th revolutionary technology of the century after biotechnology. Nanotechnology is an emerging technology that brings revolution to the different scientific fields including agriculture. In this study, we used zinc and copper nanoparticles to investigate their positive or toxic effect on maize crops. A pot experiment was conducted to test the phytotoxicity of green synthesized zinc and copper nanoparticles in wheat crops. 500g of soil was filled in pots. 5 seeds were sown in each pot. Priming of seeds was done before the sowing. Different concentrations of zinc and copper nanoparticles ie.0, 50, 100, 150, 200, 250, 300, 350, and 400ppm were used for priming the seeds. After harvesting, crop samples were analyzed for physical growth parameters of shoot length, root length, fresh shoot weight, fresh root weight, dry shoot weight, and dry root weight. Results showed that both biosynthesized nanoparticles showed different threshold levels for toxicity at different concentrations. The shoot length of wheat showed toxicity at 350 ppm of copper nanoparticle, however, for zinc nanoparticle wheat crop did not show any toxicity up to 400 ppm.

Introduction

Nanotechnology has gained great attention [1]. In agriculture and many other fields, the use of nanoparticles increases day by day all over the world. The use of nanomaterials is increasing rapidly worldwide, generating interest in their behavior within biological systems, including agricultural systems [2]. Due to their cost-effectiveness and eco-friendliness compared to chemically produced nanoparticles (NPs), bioengineered nanoparticles (BNPs) have recently been widely used in a variety of sectors around the world, including the apparel, pharmaceutical, biomedical, and food industries [3, 4]. The successful use of BNPs in these sectors opened the door for their implementation in the agricultural sector. It was discovered that the BNPs, particularly environmental stress, had greater impacts on the health and vigor of plants growing in complex ecosystems [4, 5]. Thus, it is important to comprehend how nanomaterials interact with plants, including any possible uptake and translocation of these materials within plants [6].

Nanotechnology has different applications in the agriculture field like Nano fertilizers, Nano pesticides [7]. The use of nanoparticles reduces the negative effect on the ecosystem [8]. The most widely grown crop in the world, wheat, requires a lot of nitrogen-rich fertilizers, which affects the agricultural ecosystem. This situation has been upgraded by the use of nanoparticles and intelligent delivery methods in this crop [9].

Due to their distinct physicochemical characteristics, ZnO NPs are used in a variety of consumer products; however, as their production and use rise, it is important to consider how these NPs may come into contact with living things [10]. Zinc oxide (ZnO) nanoparticles possess unique semiconducting, and optical properties. That is why, ZnO nanomaterials have been studied for wider applications. One of the most important features of ZnO nanoparticles is low toxicity and biodegradability [11].  Nanoparticles (NPs) of micronutrients are beneficial to increase crop yield and reduce abiotic stresses like drought [12]. Nanoparticles (NPs) induce changes both positive and negative according to plant species, doses, exposure time, and methods [13].

Researchers have recently become more interested in the green synthesis of nanoparticles since it is an easy, practical, and non-toxic alternative to chemical synthesis for producing nanomaterials [14]. Different micro-macro organisms, including plants, bacteria, fungi, seaweeds, and microalgae, are involved in the creation of nanoparticles [15]. Due to the presence of phytochemicals and other nutritional components, plants were more attractive for the non-hazardous synthesis of nanoparticles [16], and they also serve as reducing and stabilizing agents for the creation of novel nanoparticles [17]. Successful attempts have been made to synthesize nanoparticles of cobalt, copper, silver, gold, palladium, zinc-oxide, and platinum from plants like Conocarpus erectus [18].

In this study we used the biogenic nanoparticles of zinc and copper to investigate their positive and negative impact on maize growth, therefore the objectives of this study are;

  • Investigate the effect of zinc and copper nanoparticles on the growth parameters of wheat crops.
  • Determination of toxicity threshold level for growth parameters of wheat crop.

MATERIALS AND METHOD

Extract formation

Leaves of Monocarps at 80g were taken and washed with distilled water so that undesired particles like dust particles were removed. Then for drying, it is kept in a shady place till the elimination of moisture. Then it was ground well toymaker the substance powdery. This powdery substance was taken in falcon till used for extraction. 5gileavesipowder was dispensed with 50 mL deionized water in 250 m of conical flask and heated for 20 minutes at 75oC.  Then Leaf extract was filtered with Whatman filter paper Grade 1 [19], and the extract was kept in the freezer at 4oC for future use[20].

Preparation of 1mM ZnSO4;

0.29 g of 1 mM solution of Zinc sulfate heptahydrate was prepared and kept in tanned bottles to avoid auto-oxidation of zinc ions.

Zinc & copper nanoparticle synthesis

80 mL Zinc sulfate was added into 20 mL leaf extract by using a method developed by [21]. This solution was then heated on a stirring hot plate at 75oC for 25 minutes [22]. Production of zinc nanoparticles was confirmed by a color change from green cloudy to light dark. The solution was centrifuged at 10,000 rpm for 20 minutes. Similarly, 80ml of copper sulfate pentahydrate solution was added into 20ml leaf extracts by using the method developed by [21]. This solution was then heated on a stirring hot plate at 75oC for 25 minutes [22]. Production of copper nanoparticles was confirmed by a color change from green cloudy to high dark.

 Soil samples preparation for pot experiment

 The pot experiment was operated from 5 December to 5 March at the research area of the Government college university, Faisalabad. sand samples were cleared to erase any live and dead roots and pebbles. the sand was sieved through a 2mm sieve After completing the sieving process, each pot was filled with 1 kg of sieved sand

Treatments for zinc and copper nanoparticles

Table.1

0 ppmT1R1 Z, CT1R2 Z, CT1R3 Z, C
50 ppmT2R1 Z, CT2R2 Z, CT2R3 Z, C
100 ppmT3R1 Z, CT3R2 Z, CT3R3 Z, C
150 ppmT4R1 Z, CT4R2 Z, CT4R3 Z, C
200 ppmT5R1 Z, CT5R2 Z, CT5R3 Z, C
250 ppmT6R1 Z, CT6R2 Z, CT6R3 Z, C
300 ppmT7R1 Z, CT7R2 Z, CT7R3 Z, C
350 ppmT8R1 Z, CT8R2 Z, CT8R3 Z, C
400 ppmT9R1 Z, CT9R2 Z, CT9R3 Z, C

Seed preparation and growing of maize crop

Seeds of wheat witan 2019 taken from main gala mandi near Chenab chock Faisalabad for an experiment. Priming of seed was conducted. Seeds of wheat were dipped into the zinc nanoparticles for 12 hours.5 seeds were sowed in each pot. Priming seeds were sown in the pots.

Results:

Effect of nanoparticles on plant growth:

The zinc oxide and copper nanoparticles showed significant effects on the growth parameters of wheat crops. The application of zinc oxide nanoparticle NPs improved all the growth parameters.

Shoot Length

Data presented in figure 01 showed that the shoot length of the wheat crop was significantly increased with seed priming of zinc oxide and copper nanoparticles (NPs). The result revealed that zinc oxide nanoparticles showed a maximum shoot length (10.5cm) at 400 ppm and a minimum shoot length was observed at 0 ppm. Similarly, seed priming with copper nanoparticles (Cu NPs) also increased the shoot length of wheat crops. Fig (01) showed that the maximum shoot length of the wheat crop (20.1cm) was observed at 300ppm and the minimum shoot length was observed at 0ppm. Overall, zinc oxide nanoparticles showed more promising results in terms of crop shoot length as compared to copper nanoparticles. The result showed that the threshold for zinc oxide nanoparticles (ZnO NPs) is higher than the copper nanoparticle. Fig 02 showed that the toxicity of copper nanoparticles started at 300 ppm however for zinc oxide nanoparticles, there was no toxicity up to 400 ppm.

Fig.1. Effect of zinc and copper nanoparticles on shoot length of wheat crop

Root length

Data presented in figure 2 showed that the shoot length of the wheat crop was significantly increased with seed priming of zinc oxide and copper nanoparticles. The result revealed that zinc oxide nanoparticles showed a maximum shoot length (7.5cm) at 400 ppm and a minimum shoot length was observed at 0 ppm. Similarly, seed priming with copper nanoparticles (Cu NPs) also increased the shoot length of wheat crops. Fig (02) showed that the maximum shoot length of wheat crop (7.1cm) was observed at 300ppm and the minimum shoot length was observed at 0ppm. Overall, zinc oxide nanoparticles showed more promising results in terms of crop shoot length as compared to copper nanoparticles. The result showed that the threshold for zinc oxide nanoparticles (ZnO NPs) is higher than the copper nanoparticle. Fig 02 showed that the toxicity of copper nanoparticles started at 300 ppm however for zinc oxide nanoparticles, there was no toxicity up to 400 ppm.

 Fig. 2. Effects of zinc and copper oxide nanoparticles on root length of wheat crop                                                                    

Shoot fresh weight

Data presented in figure 1 showed that the shoot length of the wheat crop was significantly increased with seed priming of zinc oxide and copper nanoparticles. The result revealed that zinc oxide nanoparticles (ZnO NPs) showed a maximum shoot length (11.9cm) at 400 ppm and a minimum shoot length was observed at 0 ppm. Similarly, seed priming with copper nanoparticles (Cu NPs) also increased the shoot length of wheat crops. Fig (03) showed that the maximum shoot length of the wheat crop (8.9cm) was observed at 300ppm and the minimum shoot length was observed at 0ppm. Overall, zinc oxide nanoparticles (ZnO NPs) showed more promising results in terms of crop shoot length as compared to copper nanoparticles. The result showed that the threshold for zinc oxide nanoparticles is higher than for copper nanoparticles. Fig 03 showed that toxicity of copper nanoparticles (Cu NPs) was started at 300 ppm however for zinc oxide nanoparticles, there was no toxicity up to 400 ppm.

Fig. 3. Effects of copper and zinc oxide nanoparticles on shoot fresh weight of the wheat crop

Root fresh weight

Data presented in figure 4 showed that the shoot length of the wheat crop was significantly increased with seed priming of zinc oxide and copper nanoparticles. The result revealed that zinc oxide nanoparticles (ZnO NPs) showed a maximum shoot length (7.3cm) at 400 ppm and a minimum shoot length was observed at 0 ppm. Similarly, seed priming with copper nanoparticles Cu NPs) also increased the shoot length of wheat crops. Fig (04) showed that the maximum shoot length of the wheat crop (7.1cm) was observed at 300ppm and the minimum shoot length was observed at 0ppm. Overall, zinc oxide nanoparticles (ZnO NPs) showed more promising results in terms of crop shoot length as compared to copper nanoparticles. The result showed that the threshold for zinc oxide nanoparticles is higher than for copper nanoparticles. Fig 04 showed that toxicity of copper nanoparticles (Cu NPs) was started at 300 ppm however for zinc oxide nanoparticles, there was no toxicity up to 400 ppm.

Fig. 4. Effects of copper and zinc oxide nanoparticles on root fresh weight of the wheat crop

Shoot dry weight

Data presented in figure 5 showed that the shoot length of the wheat crop was significantly increased with seed priming of zinc oxide and copper nanoparticles. The result revealed that zinc oxide nanoparticles (ZnO NPs) showed maximum shoot length (5cm) at 400 ppm and minimum shoot length was observed at 0 ppm. Similarly, seed priming with copper nanoparticles (Cu NPs) also increased the shoot length of wheat crops. Fig (05) showed that the maximum shoot length of the wheat crop (3.9cm) was observed at 300ppm and the minimum shoot length was observed at 0ppm. Overall, zinc oxide nanoparticles showed more promising results in terms of crop shoot length as compared to copper nanoparticles (Cu NPs). The result showed that the threshold for zinc oxide nanoparticles (Cu NPs) is higher than the copper nanoparticle. Fig 05 showed that toxicity of copper nanoparticles was started at 300 ppm however for zinc oxide nanoparticles (ZnO NPs), there was no toxicity up to 400 ppm.

Fig. 5. Effects of zinc and copper oxide nanoparticles on shoot dry weight of the wheat crop

Root dry weight

Data presented in figure 6 showed that the shoot length of the wheat crop was significantly increased with seed priming of zinc oxide and copper nanoparticles. The result revealed that zinc oxide nanoparticles (ZnO NPs) showed a maximum shoot length (4.3cm) at 400 ppm and a minimum shoot length was observed at 0 ppm. Similarly, seed priming with copper nanoparticles (Cu NPs) also increased the shoot length of wheat crops. Fig (06) showed that the maximum shoot length of the wheat crop (3.1cm) was observed at 300ppm and the minimum shoot length was observed at 0ppm. Overall, zinc oxide nanoparticles showed more promising results in terms of crop shoot length as compared to copper nanoparticles (Cu NPs). The result showed that the threshold for zinc oxide nanoparticles (ZnO NPs) is higher than for copper nanoparticles. Fig 06 showed that toxicity of copper nanoparticles was started at 300 ppm however for zinc oxide nanoparticles (ZnO NPs), there was no toxicity up to 400 ppm.

Fig. 6. Effects of biogenic zinc and copper nanoparticles on root dry weight of the wheat crop

Microscopy of zinc & copper

The UV-spectroscopy result showed the absorbed wavelength of copper nanoparticles (Cu NPs). The copper nanoparticle was maximum as compared to zinc oxide nanoparticle or the absorption of copper nanoparticle 410 (nm) and the absorption of zinc oxide nanoparticle 376 (nm). So, results showed that copper nanoparticles absorbed the maximum wavelength as compared to zinc oxide nanoparticles (ZnO NPs).

Fig.7. Showed the microscopic property of zinc & copper nanoparticles

Discussion

One study revealed that different plant species are highly toxic to nanoparticles [23, 24] and on the other hand, it is essential for reducing heavy metal stress and fostering plant development [25]. Prior research on wheat found that after priming, the Zn concentration of seed tissues increased [26].

 The pot experiment showed a positive result of zinc oxide on the wheat crop at 400 ppm. And copper nanoparticles (Cu NPs) also showed positive results at 300ppm but at 350ppm to 400ppm copper nanoparticles showed a negative effect on wheat crops. Zinc oxide nanoparticles showed a phytotoxicity threshold at 400ppm nanoparticle concentration but copper showed a phytotoxicity threshold at 350ppm to 400ppm concentration. So, zinc oxide is less toxic than copper. The shoot length of the wheat crop attain a maximum of 400pm of zinc nanoparticles and minimum results were obtained at 0ppm [27]. While at 300ppm of copper maximum shoot length was observed. Experiment results also showed maximum shoot fresh weight at 400ppm of copper nanoparticles. While at 400ppm zinc nanoparticles showed the maximum shoot fresh weight of wheat attained at 400ppm of zinc nanoparticles. While at 300ppm copper nanoparticles showed maximum root fresh weight [28]. Above their threshold plant growth retinted because of metal stress. For plant growth minute quantity of micronutrients is required. The enzymatic metabolism of plants is sieved without micronutrients.

The phytotoxicity threshold [29] of zinc nanoparticles was started at 400ppm while the phytotoxicity threshold of copper nanoparticles [30] started at 300ppm. Increasing the threshold growth of crops was an infected mental stress. A minute quantity of micronutrients [31] is required for the growth of crops. When the micronutrients off enzymatic metabolism stop. These micronutrients enhance crop growth and growth parameters.

But the main concern is regarding the micronutrient threshold because after the threshold they cause metal stress and reduce crop growth. So, metal stress in plants is also contaminated in the food chain [32].

Conclusion

The synthesis of nanoparticles by using biological methods is eco-friendly and low-cost. Biologically synthesized nanoparticles also showed the least toxic effects on crops and plants. The synthesis of zinc and copper oxide nanoparticles for biological methods to use plant extract Conocarpus erectus L. was eco-friendly and less toxic. In this experiment (ZnO NPs) germination up to the threshold which was 400ppm for wheat crops. Copper nanoparticles (Cu NPs) also increase plant growth but the threshold of copper nanoparticles (Cu NPs) was narrow in comparison with zinc oxide (ZnO NPs) nanoparticles at 300ppm. above 300ppm copper nanoparticles (ZnO NPs) was toxic and deteriorated plant growth parameters in wheat crops.

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