Heavy metal:

Heavy metals are defined as metallic elements that have a relatively high density compared to water

(Erric, 2023). With the assumption that heaviness and toxicity are inter-related, heavy metals also

include metalloids, such as arsenic, that are able to induce toxicity at low level of exposure (Duffus,

2002). In recent years, there has been an increasing ecological and global public health concern

associated with environmental contamination by these metals. Also, human exposure has risen

dramatically as a result of an exponential increase of their use in several industrial, agricultural,

domestic and technological applications.

Heavy metals are conventionally defined as elements with metallic properties and an atomic number >20. The

most common heavy metal contaminants are Cd, Cr, Cu, Hg, Pb, and Zn. Metals are natural components in

soil(Gaur et,.al 2004). Some of these metals are micro-nutrients necessary for plant growth, such as Zn, Cu, Mn,

Ni, and Co, while others have unknown biological function, such as Cd, Pb, and Hg.

Heavy metals, which are defined as metals with relatively large densities and atomic weights, have a

variety of uses that have caused major environmental and possibly health issues.Chromium is a

significant heavy metal that is involved in biological metabolism, however exposure to Cr can have a

serious negative influence on the health of the general population oremployees. Skin contact,

inhalation, and ingestion are used as three exposure routes to examine the harmful consequences of Cr

exposure. The metabolism of proteins, lipids, and carbohydrates all contain the micro nutrient

chromium (Aris Ismanto et al., 2022).

An essential metal, Cr is used in the production of refractory materials, metallurgy, painting, metal

plating, wood treatment and leather tanning, among many other uses. The detrimental consequences

of Cr exposure differ according on various modes of exposure, including dermal contact, inhalation,

and ingestion, according to the “National Institute for Occupational Safety and Health”(NIOSH)

(Tiina Santonen et al., 2022). The general public is exposed to Cr by the consumption of food or
water that has been polluted with the substance, usually in or near an area that has been contaminated

with it for a long period (MahdiBalali-Moodetal.,2021).In contrast to exposure by eating, exposure to

Cr through inhalation and skin contact happens during work, for example in tanneries or the lumber

industry (Oginawati et al., 2021). The hazardous consequences are strongly influenced by the

exposure path in addition to the exposure amount.

Reduction of abiotic stress:

The most popular strategy for reducing abiotic stress, particularly the stress caused byheavy metals in

crop plants, in recent years has been the use of organic fertilizer and important micro-nutrient

minerals (Etesami et al., 2018). Essential minerals such as silicon, magnesium, calcium, selenium, and

others can be applied exogenously to treat metal stress (Hayatetal.,2012). Additionally, adding

organic amendments like aloe vera gel, compost, biochar, vermicompost, etc. might help plants to

grow healthy and lessen the harmful effects of many environmental factors(Sharmaet al., 2019;

Pierre-Louis et al., 2021).

Additionally, aloe vera has enzymes and plant hormones that lessen transplant shock andincrease the

plant's resistance to disease, stress, and drought. For instance, aloe vera contains significant amounts

of acemannan and saponin, both of which have antibacterial, antifungal, and

Antiviral effects. This aids in defending plants against infections such as dangerous microorganisms,

fungi, yeast, mold, or blight (Bashyal, Bishnu Maya et al., 2022).

Low doses of selenium help a variety of physiological processes in plants, including growth

promotion, an improvement in antioxidant capacity, lipid peroxidation, a decrease in electrolyte

leakage and an increase in the sugars accumulation and starches that improve quality and yield, and

adelayinsene scence and ripening (Saha et al.,2019). According to Chi et al.(2017),selenium may have

positive impacts on crop growth, quality, and production potential for crops grown in difficult
environments. Se may limit heavy metal absorption and contamination by absorbing heavy metals in

growth conditions containing heavy metals, as well as inhibit their transmission from root to shoot

(Zhao et al., 2019). Selenium has great attention due to its ability to neutralizetheharmful effects of

heavy metals(Huanget al.,2018).The group of metallic elements known as heavy metals that have

high densities and atomic weights. They can negatively impact both human beings and the

environment due to their hazardous qualities (Klaassen, 2020).

Due to their persistent presence in the environment and capacity to build up in living things, heavy

metals including lead, mercury, cadmium, and arsenic cause special concern. There are many ways that

these metals can get into the food chain, including through polluted water, soil, and air. Heavy metals,

once ingested by organisms, can result in a variety of health issues, such as neurological illnesses, renal

damage, and aberrant growth patterns (Jarup, 2003).

Heavy metal pollution is mostly caused by industrial processes including mining, smelting, and

production. In addition, poor agricultural practices, such as the indiscriminate use of pesticides and

fertilizers, and incorrect electronic waste disposal all contribute to the release of heavy metals into the

environment (Bose-O'Reilly et al., 2018).

The impact of heavy metal pollution is being lessened by enacting stronger laws on industrial

emissions, supporting environmentally friendly waste management techniques, and developing cleaner

production methods. In order to pinpoint contamination hotspots and guarantee public safety,

monitoring and testing of environmental samples and food products are also essential (Abrahams,

2019).

In conclusion, heavy metals provide serious dangers to the environment and to human health. It is

crucial to spread knowledge about the causes and effects of heavy metal pollution and to take

preventative steps to limit and avoid their release into the environment.
Biodata of chromium:

Chromium is the 21st most abundant element in the Earth's crust with a mean concentration in United

States soils of about 40 mg/kg. Although it exists in several oxidation states, the zero, trivalent, and

hexavalent states are the most important in commercial products and the environment. Nearly all

naturally occurring chromium is in the trivalent state, usually in combination with iron or other metal

oxides. Although only about 15% of the chromium mined is used in the manufacture of chemicals,

most applications of chromium utilize the chemistry of chromium. For instance, the “stainless” nature

of stainless steel is due to the chemical properties of the chromium oxides which form on the surface of

the alloy. Similarly, the protective properties of chrome plating of metals, chromated copper arsenate

(CCA) treatment of wood, and chrome tanning of leather are all dependent on chromium chemistry.

The key to these uses is that under typical environmental and biological conditions of pH and

oxidation–reduction potential, the most stable form of chromium is the trivalent oxide. This form has

very low solubility and low reactivity resulting in low mobility in the environment and low toxicity in

living organisms. In this paper the chemical properties of chromium are discussed for the major

commercial products in the context of the Eh–pH diagram for chromium.

Sources of chromium:

Waste disposal into the environment generates adverse effects by changing the normal physiochemical

properties of soil and water(Nigam et al., n.d.). However, several conventional physicochemical methods

are currently being practiced but biological methods are better alternatives, because they are

economical as well as eco-friendly.

Naturally chromium occurs in various oxidation states, but Cr (III) and Cr (VI) are of remarkable

worry biologically. Chromium is a necessary metal that is involved in the metabolism of glucose in
humans and animals, but its hexavalent form is very toxic and carcinogenic (Raziyeh Rahmaty & Khara,

2017). Demand for Cr (VI) is increasing day by day due to its extensive use in numerous industrial and

chemical processes such as film and photography, galvanometric and electrical procedures, metal

cleaning, plating and electroplating, leather, and mining (Unal et al., 2010).These industrial processes

produce toxic waste in a large amount that contains hexavalent chromium with concentrations ranging

from tens to hundreds of milligrams per liter along with other forms of chromium (JAMIL & NAEEM,

2013). Hexavalent chromium is often found in soil and ground water due to its wide spread industrial

use in several process industries(El-Shamy, n.d.), such as tannery, electroplating, steel industries etc.

Cr(VI) are highly toxic(Zhang, 2022) pollutant even at very low concentration(Gu & Cheung, 2001). As

many aerobic and anaerobic microorganisms are capable of reducing Cr(VI) to Cr(III), bioremediation

may play an important role for the detoxification from Cr (VI) even at very low (ppm or ppb) level. It

has already been reported that because of the presence of some enzymes called chromium reductases,

completely different microorganisms belonging particularly to the genus, Pseudomonas can reduce Cr

(VI) to Cr (III). The reduction of transformation capacity of Cr (VI) by microorganisms at higher (Hsu,

2011) initial concentration of Cr(VI) has been observed by other researchers and the phenomenon has

been explained by the presence of inhibitory effect of Cr(VI) at high concentration level (Turick & Apel,

1997).

Anthropogenic source of chromium

Cr and its compounds have multifarious industrial uses. They are extensively employed in leather

processing and finishing (Nriagu, 1988) in the production of refractory steel, drilling muds,

electroplating cleaning agents, and catalytic manufacture and in the production of chromic acid.

Hexavalent chromium (VI) compounds are used in industry for metal planting, cooling tower water

treatment, hide tanning and until recently, wood preservation. These anthropogenic activities have led
to the wide spread contamination that Cr shows in the environment and have increased its

bioavailability and bio mobility. (Kotas and Stasicka, 2000).

The leather industry is the major cause for the high influx of Cr to the biosphere, accounting for 40%

of the total industrial use (Barnhart, 1997). In India, about 2000 to 32000 tons of elemental Cr annually

escape into the environment from tanning industries. Even if the recommended limit for Cr

concentration in water are set differently for Cr (III) (8 μg L -1) and Cr (VI) (1 mg L-1), it ranges from 2

to 5μ g L-1 in the effluents of these industries (Chandra et al. 1997).

Oxidation State of Chromium:

The oxidation state of Cr exists in a wide range of forms, from bivalent chromium [Cr(II)] to Cr(VI),

but the most common forms are Cr(III) and Cr(VI) (Tariq et al. 2019; Carlos et al. 2016), which can

form chromate (CrO42-), potassium dichromate (Cr2O72-), H2CrO4, HCrO4- (Jobby et al. 2018),

chromic hydroxide (Cr(OH)3) or chromic oxide (Cr2O3).

The oxidation/reduction of Cr (VI) and Cr (III) in soil is thermodynamically spontaneous and can

occur simultaneously. Cr from different sources in the soil will eventually form different valences in

this redox reaction (Shahid et al. 2017). There are many factors that affect the bioavailability of heavy

metals in soil, including soil pH, soil texture, redox status (Shahid et al. 2017), clay mass, organic

matter content (Zeng et al. 2011), oxidized reducing substances, and the presence of other cations in

the soil (Narendrula-Kotha et al. 2019).

Main heading Chromium toxicity in plants

Toxicity of Cr to plants depends on its valence state, Cr (VI) is highly toxic and mobile whereas Cr

(III) is less toxic. Toxic effects of Cr on plant growth and development include alterations in the

germination process as well as in the growth of roots, stems and leaves, which may affect total dry
matter production and yield. Cr also causes deleterious effects of plant physiological processes such as

photosynthesis, water relation and minerals nutrition.

Physiological effect of chromium on plants:

Although there is no conclusive evidence of the essentiality of Cr in plant metabolism, some studies

have shown that small additions of Cr have stimulating effects on growth and plant productivity

(Zayed and Terry, 2003). Beneficial effects, such as antifungal and growth stimulation have also been

observed in plants grown in soil or in solutions containing Cr (Barceló et al., 1993). For example, in

one study, the presence of 1 µmol L−1 Cr was found to stimulate plant growth (Ghosh and Singh,

2005), and, according to El-Bassam (1978), low concentrations of Cr3+ stimulate chlorophyll synthesis

and photosynthesis activity. Increases in carbon assimilation were also observed in a study conducted

with E. crassipes collected from the river Imbé, in Southeastern Brazil and exposed to 1 mmol L−1

Cr2O3 (Paiva et al., 2009). Toxic effects of Cr on the growth and development of plants, however, are

more apparent, and have been increasingly investigated. These include several metabolism changes,

such as modifications to the germinating process and the growth of the roots, stems and leaves

(Shanker et al., 2005). Cr also causes harmful effects on physiological processes, such as

photosynthesis, water relations and mineral nutrition (Nagajyoti et al., 2010; Shanker et al., 2005), and

can also generate morphological changes (Daud, 2014; Rodriguez et al., 2012; Singh et al., 2013).

In plans, high levels of Cr supply can inhibit seed germination and subsequent seedling growth. Peralta

et al., (2001) found that 40 ppm of Cr (VI) reduced by 23% the ability of seeds of lucerne ( Medicago

sativa) to germinate and grow in the contaminated medium. Reductions of 32-57% in sugarcane bud

germination were observed with 20 & 80 ppm Cr, respectively (Jain et al. 2000).

Effect of chromium on plant morphology:

Plant exposure to Cr has been shown to reduce certain root parameters, such as diameter, surface area,

and number of hairs, and has also been determined as the cause of wilting and plasmolysis in root cells

(Ali et al., 2011; Moral et al., 1995; Panda and Choudhury, 2005). Cr6+ has been proven to reduce the

number of root cells as it enters the cell through the cellular membrane, and can be responsible for

cytotoxic effects and damage to DNA (Chidambaram et al., 2009). Cr may also cause alterations in the

ultrastructure of chloroplasts in cell membranes and necrosis in plant leaves (Chidambaram et al.,

2009; Choudhury and Panda, 2005; Ghani, 2011; Tiwari et al., 2009). For example, in a study

conducted with E. crassipes after 2 and 4 days exposure to 1 and 10 mmol L−1 Cr2O3 and K2Cr2O7,

it was observed that after 2 days of exposure to 10 mmol L−1 plants exposed to Cr6+ plants were

completely necrosed, while plants subjected to 1 mmol L−1 Cr6+ showed necrosis only after 4 days of

metal exposure, indicating lesser toxic effects to lower Cr concentrations. After 4 days of exposure to

10 mmol L−1 of Cr3+, plants showed healthier conditions when compared to the control plants and the

plants exposed to 1 mmol L−1 Cr3+(Paiva et al., 2009). It has also been demonstrated that Cr6+ is

more phytotoxic than Cr3+, since, at high Cr6+ concentrations (1 mmol L−1 ), complete distortion of

the chloroplatinic membrane alongside severe thylakoid disarrangement were observed (Choudhury

and Panda, 2005). Cr has also been shown to be toxic to bean plants, causing changes in morphology

compared to control plants (Azmat and Khanum, 2005), including reduced root length with increasing

Cr concentrations ( 0, 5, 10, 50, 100, 150, 200 and 250 mg kg−1 ). Canopy size, however, was less

affected. Changes in morphology of corn leaves after one day of exposure to 300 mg L−1 of K2Cr2O7

have also been observed, where young leaves suffered epinasty after 6 h of exposure, and significant

wilting was observed after 12 h, possibly due to water stress caused by Cr (Wang et al., 2013).

Toxicity symptoms of Cr3+ were also observed in Ocimum basilicum plants exposed to CrCl3·6H2O

concentrations of 0, 2, 4, 6, and 8 mg L−1 . Alterations were also observed in the cytoplasm, such as

cytoplasm de organization, alterations in the ultrastructure of chloroplasts, an underdeveloped lamellar

structure with widely spaced thylakoids and less amounts of granna (Bishehkolaei et al., 2011). These

morphological changes can, in turn, severely affect photosynthetic pigments and photosynthesis

(Bishehkolaei et al., 2011), discussed in the following section.

Effects of chromium on photosynthesis metabolism:

Photosynthesis inhibition during stress caused by metals is one of the main consequences in plants,

since these elements invariably, directly or indirectly, affect the photosynthetic apparatus (Sytar et al.,

2013). Metals alter the functions of the chloroplast membrane and components of the electron transport

chain in mitochondria (Ventrella et al., 2011) and, thus, inhibit part of the energy transfer from one

level to another (Sytar et al., 2013). According to Dixit et al. (2002), the change in redox reactions of

Cu and Fe carriers (Fig. 2), allows for Cr6+ to be transferred via cytochrome mitochondria, allowing

this element to bind to the heme group of cytochrome, interfering with the transport electrons. Cr6+

can also bind to the cytochrome a3, as well as the Complex IV of cytochrome oxidase (E.C. 1.9.3.1.),

thus causing a severe inhibition of the activity of this enzyme (Dixit et al., 2002). Metals can also

affect the activity of the photosystem I (PSI) and the photosystem II (PSII), located in the thylakoid

membranes. PSII has been shown to be more susceptible to toxic metal effects in comparison to PSI

(Sytar et al., 2013). For example, in thylakoids isolated from Brassica juncea individuals exposed to

200 and 400 mmol L−1 Cr6+, greater activity of PSII was observed when compared to control plants

(Gupta et al., 2009). The PSI activity in thylakoids of seedlings exposed to 200 µmol L−1 Cr6+ was

similar to controls, whereas the PSI activity in plants exposed to over 200 mmol L−1 Cr6+

concentrations were lower compared to control seedlings (Gupta et al., 2009). In research conducted

with chloroplasts isolated from Beta vulgaris L. submitted to Cr6+ exposure, a significant inhibition of

electron transport activity in both PSI and PSII was observed. Within the PSII, the pheophytin and

plastoquinone regions were more affected (Fig. 2) (Pandey et al., 2013). The water oxidation complex,
however, was unaffected by exposure to Cr6+. Studies have also demonstrated that Cr interferes with

gas exchange parameters consisting of CO2 assimilation (A), evapotranspiration (E), stomatal

conductance (gs) and internal carbon (Ci) (Rodriguez et al., 2012). In this context, Cr3+ causes water

imbalance in plants and affects stomatal opening, leading to gs changes (Barbosa et al., 2007). Cr3+ in

high concentrations also affects photosynthesis in terms of carbon assimilation, electron transport,

photophosphorylation and alterations in Rubisco activity (Pandey and Sharma, 2003). Additionally,

several enzymes are inhibited at high Cr concentrations (Vazques et al., 1987), which leads to

reduction of photossynthetic yield (Nagajyoti et al., 2010). When comparing the effects of Cr3+ and

Cr6+ on photosynthetic parameters, Paiva et al. (2009) found that the Cr3+ can increase carbon

assimilation in E. crassipes. However, the same study observed that Cr6+ caused a decrease in carbon

assimilation and chlorophyll a content, and fluorescence parameters. One explanation for the reduction

in photossynthetic yield caused by Cr6+ can be ascribed to the disorganization of the ultrastructure of

chloroplasts (Van Assche and Clijsters, 1983) and inhibition of the electron transport processes due to

a deviation of electrons from the PSI electron donor side (Shanker et al., 2005). It is possible that the

electrons produced by the photochemical process are not used for carbon sequestration, as evidenced

by the low photosynthetic yield observed in plants exposed to Cr6+. Thus, it is hypothesized that part

of the electrons may be used for the reduction of oxygen molecules, which may explain oxidative

stress caused by Cr6+(Shanker et al., 2005). A decrease in photosynthetic rates, transpiration and

stomatal conductance of Oryza sativa L. cultured with 50, 100, 150, 200, 300, 400 and 500 mg kg−1 of

Cr6+ was observed, compared to control plants. The results indicate that gas exchange parameters and

chlorophyll a, b and carotenoids were reduced with increasing concentration of Cr6+(Ahmad et al.,

2011). The reduction of photosynthetic pigments by metals occurs by inhibition of the activity of

enzymes involved in the biosynthesis of chlorophyll, as well as by substitution of the central Mg ion of

the chlorophyll molecule by the metal, impairing the reception of light and leading to the collapse of
photosynthetic activity (Küpper et al., 2002; Prasad and Strzałka, 1999). For example, Dhir et al.

(2009) found a significant decrease in the activity of ribulose bisphosphate carboxylase oxygenase

(RuBisCO, E.C. 4.1.1.39.) induced by exposure to wastewater rich in Cr in Salvinianatans, and

suggested that this result may be explained by the substitution of Mg2+ in the active site of

RuBisCOby subunits metal ions. This would, in turn deplete chlorophyll content as suggested by other

authors (Vajpayee et al., 2000).

Oxidative stress induction:

Plants have developed strategies to tolerate adverse conditions and their negative effects (Liu and Yao,

2007), and excess of Cr and other metals can initiate a variety of metabolic responses leading to

changes in plant development (Hayat et al., 2012). Redox metals, such as Cr, can directly generate

oxidative injury via the Haber-Weiss and Fenton reactions, which in turn lead to the production of

reactive oxygen species (ROS) in plants (Flora, 2009). Reactive oxygen species (ROS) such as

superoxide radicals (O2 −•), hydrogen peroxide (H2O2), hydroxyl radicals (OH• ) and oxygen singlets

(1 O2) are normally produced in small amounts in aerobic organisms. However, under stress

conditions, their production is dramatically increased (Miller et al., 2010). This overproduction of ROS

in plants can result in cell homeostasis disruption, DNA strand breakage, defragmentation of proteins,

or cell membrane and damage to photosynthetic pigments, which may trigger cell death (Flora, 2009).

Production of ROS in plants exposed to Cr has been demonstrated, and has been shown to result in

oxidative stress leading to DNA, protein and pigment damage, as well as the initiation of lipid

peroxidation (Choudhury and Panda, 2005; Panda, 2003). Absorption of Cr is facilitated by a carrier

membrane, thereby ROS generation and their impact on the plasma membrane are very important

(Maiti et al., 2012). ROS are usually eliminated in their production sites by antioxidant compounds

(Hossain et al., 2012), such as antioxidant enzymes. The main antioxidant enzymes studied in plants

include catalase (CAT, E.C. 1.11.1.6), guaicol peroxidase (E.C. 1.11.1.7), glutathione reductase (E.C.
1.8.1.7), ascorbate peroxidase (E.C. 1.11.1.11) and superoxide dismutase (SOD, E.C. 1.15.1.1) (Gill

and Tuteja, 2010; Panda and Choudhury, 2005). Besides enzymes, increases in the synthesis of other

compounds have also been reported in response to metal-induced stress, such as polyamines (PAs)

(Hussain et al., 2011), proline (Pro) (Kishor et al., 2005), nitric oxide (NO) (Delledonne, 2005; Liu and

Yao, 2007) and metallothionein (MT) (Teixeira et al., 2013). Interestingly, despite the harmful effects

of ROS, it has been proposed that H2O2, in itself a ROS species, can act in signaling mechanisms in

response to stress (Mittler et al., 2004; Sharma and Dietz, 2009), and has been proposed as a key

molecule to elicit signal transduction for metal tolerance in plants, since it is immediately produced

under stress by metals (Seth et al., 2012). Further discussion in this regard shall follow. Many

antioxidant enzymes have been demonstrated as significantly altered in plants exposed to metals. SOD,

for example, acts as a first line of defense against oxidative stress in all aerobic organisms and all

subcellular compartments prone to oxidative stress mediated by ROS (Gill and Tuteja, 2010). This

enzyme catalyzes the dismutation O2 •− to H2O2 and O2 (Gill and Tuteja, 2010), and is found in

almost all cell compartments, as well as in the water and ascorbate-glutathione cycles in chloroplasts

and in the cytosol, mitochondria, peroxisomes and apoplasts (Bhaduri and Fulekar, 2012). CAT is also

an essential enzyme for the detoxification of ROS in plants (Panda and Choudhury, 2005).

Effect of chromium on the nutrient uptake:

Heavy metal stress affects nutrient uptake in plants by interacting with other essential minerals.

Chromium restricts the uptake of nutrients in soil by forming insoluble compounds(Chigonum et al:

2019). Nutrient uptake is thereby inhibited by the metal toxicity especially when the concentration of

the metal exceeds its permissible limits (Osu et al;2016). For instance, excessive Cr had been observed

to reduce the uptake of essential minerals like iron (Fe), magnesium (Mg), phosphorus (P) and calcium

(Ca) by masking the sorption sites and forming insoluble complexes(Antônio et al., 2011). However,
Cr transport to different parts of Citrullus plants had increased leading to enhancement in the

concentrations of manganese (Mn) and P, and reduction in Sulphur (S), copper (Cu), zinc (Zn) and iron

(Fe) contents in the leaves, suggesting that Cr disturbs the nutrient balance(Dube et al., 2003). Turner

and Rust ((P. Sundaramoorthy et al., 2010)) also suggested the similar effects of Cr on the uptake of

various nutrients under Cr toxicity. A gradual decrease in the uptake of micronutrients like Zn, Cu, Fe,

Mn and macronutrients like potassium (K), P and nitrogen (N) had been noticed in the paddy plants

(Oryza sativa L.) under excessive Cr exposure. This reduced nutrient uptake may occur due to decline

in the root growth and impairment of the root penetration under Cr toxicity, or may be due to the

decrease in essential element translocation because of the displacement of nutrients from the

physiologically important binding sites(Shahzad et al., 2018).

Effect of chromium on enzymatic antioxidative system:

Activities of various antioxidative enzymes drastically change in plants when subject to Cr toxicity.

ROS leads to oxidative stress, which may affect different subcellular compartments sensitive to ROS.

Superoxide dismutase (SOD) is considered as a first line of defense against various stresses in almost

all the aerobic organisms (Shahzad et al., 2018). Dismutation of superoxide ion is catalyzed by the

SOD enzyme, which is localized in almost all the cellular compartments, leading to the production of

hydrogen peroxide and release of oxygen (Sarvajeet Singh Gill & Narendra Tuteja, 2010). SOD is

involved in the Asada–Halliwell cycle in chloroplasts and also present in cytosol, apoplasts,

mitochondria and peroxisomes (Bhaduri & Fulekar, 2011). For the removal of ROS, catalase (CAT)

enzyme also plays crucial role, hence considered as important antioxidant enzyme (Sanjib Kumar

Panda & Choudhury, 2005). Dismutation of H2O2 into O2 and H2O is undergone by this enzyme (Amna

Mhamdi et al., 2010).

When exposed to Cr (III) stress for seven days, chamomile plants showed increased accumulation of

Cr mainly in the roots of the plants, which contained high concentrations of ROS, nitric oxide and

thiols. At higher concentration of Cr (III), SOD activity specifically was increased in the roots, while

level of H2O2 showed irregular trend under different concentrations of Cr due to the altered activities of

various peroxidases (Jozef Kováčik et al., 2013). Different concentrations of Cr (VI) (50, 100, 200 and

300 µM L−1) escalated the production of H 2O2 leading to the lipid peroxidation and triggered the

activities of antioxidative enzymes like SOD and guaiacol peroxidase (GPX) in comparison to control

plants (Swatilekha Maiti et al., 2012). In contrast, activities of peroxidase (POD), SOD and CAT were

decreased when subjected to Cr (III) in a dose dependent manner (Tang et al., 2014).

Maintaining metabolic functions under stress conditions is crucial for plants to survive. Therefore, a

balance between generation and scavenging of ROS is required, which is achieved by regulating the

production of enzymatic and non-enzymatic antioxidants. Hence, the ability of plants to cope with

oxidative stress is characterized by the degree of antioxidant activities (Shahzad et al., 2018).

However, activities of these antioxidants may vary with the duration, crop species and tissues under

any stress condition [135]. For instance, Echinochloacolona plants showed increased activities of POD

and CAT in tolerant calluses in comparison to non-tolerant ones [29]. At 0.5 mg L−1 concentration of

Cr (VI), CAT activity was increased, however decreased at higher concentrations (1.0–2.0 mg L −1).

Activity of CAT was measured highest at 2.0 mg L −1 concentration of Cr in the roots of K. candel but

decreased at higher concentration [30]. As CAT is an iron-porphyrine biomolecule, reduction in CAT

activity indicates that Cr has the potential to interact with iron in metabolic pool or it may influence the

presence of active form of iron [136]. Cr toxicity has detrimental effects on antioxidant enzymes such

as POD, GPX, glutathione reductase (GR) and ascorbate peroxidase (APX), hence resulting in the

inhibition of enzyme activities [129,137]. Plants equipped with an efficient antioxidant system are

more capable to withstand and tolerate higher Cr concentrations. Failure to do so results in the
breakdown of the plant defense system hence activities of antioxidants are jeopardized leading to

reduced plant growth or even leading to plant death.

Effect of Chromium on non-enzymatic antioxidative enzyme:

Apart from the enzymatic antioxidants, plants are also comprised of a complex non-enzymatic

antioxidant defense system to avoid the toxic effects of ROS. These non-enzymatic antioxidants

consist of low molecular weight molecules such as ascorbic acid, glutathione (GSH), phenolic acids,

carotenoids, flavonoids, etc. [94,138] and some high molecular weight secondary metabolites such as

tannins [139]. Biosynthesis and accumulation of these non-enzymatic antioxidants by plants could be

due to two main reasons. First, plants have an innate ability to synthesize a variety of phytochemicals

to carry out their normal physiological functioning or to protect them from any pathogenic or

herbivores. Second, plants also synthesize phytochemicals to respond to the environmental factors

which could be due to their natural tendency of defense against any biotic and abiotic stress [ 140].

Therefore, these lower molecular weight antioxidants are synthesized and act as a redox buffer to

interact with cellular components and directly influence plant growth and development by modulating

different processes from mitosis to cell elongation and to senescence. Hence, it is crucial for plants to

synthesize these antioxidants under stressed conditions. Glutathione is a redoxactive molecule that can

be present in a reduced form GSH or an oxidized form GSSG. It plays important roles in the plant

defensive system including biosynthetic pathways, detoxification, antioxidant biochemistry and redox

homeostasis [141,142]. GSH acts as an antioxidant by quenching ROS and is involved in the

ascorbate-glutathione cycle, which eliminates damaging peroxides [143]. In poplar trees, glutathione

(GSH) biosynthesis was stimulated under Cr toxicity [144]. In the leaf extracts of tomato

(Lycopersicon esculentum), maize (Zea mays) and cauliflower (Brassica oleracea) plants, GSH level

increased subjected to Cr toxicity [145]. Alterations were observed in the glutathione pool dynamics
where individual level of GSSG and GSH and GSH/GSSG ratio was affected, however sorghum

(Sorghum biclor) plants showed potential to scavenge the free radicals generated under Cr toxicity

[146].Under sub-optimal conditions, level of antioxidants may decrease or increase depending on the

severity of stress. For instance, a sharp decline in the GSH pool was observed under Cr stress, which

severely affected the roots of the plants. Many reports suggested the oxidation of various cellular thiols

like GSH and cysteine in plants subjected to Cr (VI) stress in in vitro conditions [147]. Therefore, to

maintain the redox homeostasis of the cell and for scavenging of free radicals, the interconversion of

reduced and oxidized forms of glutathione (GSH and GSSG respectively) is required.

A non-enzymatic antioxidant “carotenoid” contributes in providing protection to the chlorophylls

against stress conditions by replacing peroxides and scavenging of photodynamic reactions [148]. For

instance, carotenoid level increased in Capsicum annuum plants subjected to industrial effluents

specifically containing Cr [27], acting as a defensive mechanism for capsicum plants to scavenge the

free radicals. Other non-enzymatic antioxidants such as cysteine, proline, nonprotein thiol, etc. may

also contribute in modulating resistance against Cr toxicity and protect the macromolecules from the

free radicals generated during the oxidative burst [77]. In one of the reports, carotenoids, non-protein

thiol (NP-SH) and cysteine level increased in the plants subjected to Cr [72].For providing protection

against different types of stresses, accumulation of compatible osmolytes like proline (Pro) occurs in

most of the plants for providing membrane stability and osmotic adjustment [94,149]. Proline contents

can increase against different types of biotic and abiotic stresses such as salinity, drought, temperature,

heavy metal and pathogen attack [77]. For instance, contents of Pro increased

in Ocimumtenuiflorum L. under Cr stress, which acted as an antioxidant by providing protection

against the hazardous effects of metal [31]. Reports suggested that proline is the only amino acid that

accumulates in the leaves of plants under stress conditions [150]. It starts accumulating even at low

doses of stress and increases in a dose dependent manner. Hence, accumulation of proline contributes
to osmotic adjustment when it gets accumulated in tissues and acts as a dependent marker for

genotypes for the stress tolerance [150]. Polyamines are other non-enzymatic Osmo protectants, which

increase under different abiotic stress conditions [151], and also associated with the boosting up of

plant defensive mechanisms [92,152]. Polyamines including putrescine, spermine and spermidine have

been investigated to increase Cr tolerance in plants under Cr toxicity [38].

Effect of chromium on the endogenous levels of plant hormones:

Plant hormones control and regulate plant growth and development through different biochemical and

physiological process. These hormones may act either close to or remote from their synthesis site to

regulate responses to environmental stimuli or genetically programmed developmental changes [153].

Hormones thus have a vital role in plant adaptation to abiotic stress, from which the plant may attempt

to escape or survive under stressful conditions [154]. Thus, abiotic stresses often alter the production,

distribution or signal transduction of growth as well as stress hormones, which may promote specific

protective mechanisms. The phytohormone abscisic acid (ABA) plays a vital role against abiotic

stresses, thus it is considered as a stress hormone. Under different types of abiotic stress conditions,

endogenous level of ABA are increased drastically in plants, which boosts up the signaling pathways

and activates expression of ABA-responsive genes [155]. For instance, strong expression of ABA

biosynthesis genes (OsNCED2 and OsNCED3) and four ABA signaling genes were upregulated in

response to heavy metal stress [156]. Salicylic acid (SA) also provides protection under Cr stress. SA

plays a significant positive role in growth and development of plants as well as ripening and abiotic

stress tolerance [157]. Interaction of ABA and SA also contributes in boosting the defensive strategies

of plants against Cr toxicity [158]. Apart from this, SA also plays essential role in combination with

jasmonic acid (JA) and ethylene, where the interaction of these hormones provides tolerance against Cr

stress [159,160]. The biosynthesis, transportation and accumulation of these plant hormones boost the
signaling pathways, activating certain antioxidant gene expressions and stimulating the production of

Osmo protectants such as proline, soluble sugars, amino acids, etc. [161].

Indole acetic acid (IAA), a plant hormone of auxins contributes to the growth and development of

plants under ideal as well as stressed conditions [162]. IAA plays a key role in plant adaptation to

heavy metal stress by either increasing the membrane permeability or by increasing the concentration

of osmotically active solutes [16,163]. The level of IAA usually increases in the plants under metal

toxicity by affecting different metabolic activities of plants including growth and hormonal balance

[164]. Gibberellins (GAs) are considered as comprehensive class of tetracyclic diterpenoid carboxylic

acid compounds. Different forms of GAs have potential to play the role of growth hormone in higher

plants such as GA1 and GA3 [165]. Generally, at low concentration of metal like Cr leads to the

elevated level of GA3, although its high concentration reduces the GA3 content [166]. Apart from

these, plant hormone like cytokinins (CKs), contributes in the regulation of plant development by

stimulating cell division and elongation. Cr stress alters endogenous level of CKs suggesting that CKs

are also involved in tolerating the stress [155]. A few reports also indicated the reduction in the

synthesis of CKs and their transport from roots to other aerial parts of plants during Cr toxicity and

they are also found to interact with other plant hormones [167].

Genetic mechanisms to control Cr toxicity in plants:

A significant problem is avoiding and reducing the harmful effects of heavy metals contamination in

soil (Zeeshan et al., 2021). Genetic engineering can significantly improve a plant’s ability to transform,

translocate, and lessen the adverse impacts of heavy metals (Raza et al., 2021). Omic tools have gained

a lot of interest recently for their use in plant development and programs to mitigate agricultural

production challenges, specially to mitigate heavy metal stress (Khan et al., 2021). To identify target

genes, proteins, and metabolites linked to Cr detoxification and stress tolerance responses in plants,
genomics, proteomics, and metabolomics have become effective methods (Chaudhary et al., 2019). It

is possible to modify the Cr stress-responsive genes, proteins, and metabolites to either increase plant

tolerance to Cr stress or decrease Cr accumulation (Thakur et al., 2019). Tools for genetic engineering

that are particularly effective at changing the genes involved in the acquisition, transport, and

accumulation of Cr inside the plant are necessary for this type of manipulation (Khan et al., 2021). The

main goal of genetic engineering is the creation of tolerant varieties using either a transgenic approach

or genome editing (Raza et al., 2021). Anwar and Kim (2020) reported that through genome editing

active participation in the control of plant metabolism, essential genes important for increased metal

tolerance have been developed into transgenics, which provide insights into how to understand and

improve the tolerance capacity of plants. A successful method for creating resistant cultivars is to

transfer candidate genes from plants with a high tendency for HM hyper-accumulation (Rahman et al.,

2022).

The best way to reduce metal toxicity within cellular locations is to use transgenic plants with altered

efficiencies for metal transport into vacuoles (Khan et al., 2021). Heavy metals (HM) transporter genes

are thought to be potential candidates for genetic engineering to improve metal tolerance in plants

(Zhang et al., 2018). OsMTP1 in cultivated tobacco (Nicotiana tabacum) and PgIREG1

in Arabidopsis are two examples of metal transporter genes that have been genetically modified

(Merlot et al., 2014; Das et al., 2016). Other metal transporter genes include those that encode metal

chelators, metallothionein (MTs) (Peng et al., 2017), and genes associated with antioxidant machinery

(Peng et al., 2017; Raza et al., 2021). The use of transgenic techniques to increase resistance to metal

oxidation has also been documented. Transgenic hyperaccumulators may be created by manipulating

the antioxidant system to maintain redox equilibrium to avoid the destruction of biomolecules such as

DNA, proteins, and lipids and to maintain the structural and functional stability of cellular structures of

plant under Cr stress (Du et al., 2019).

The best way to reduce metal toxicity within cellular locations is to use transgenic plants with altered

efficiencies for metal transport into vacuoles (Khan et al., 2021). Heavy metals (HM) transporter genes

are thought to be potential candidates for genetic engineering to improve metal tolerance in plants

(Zhang et al., 2018). OsMTP1 in cultivated tobacco (Nicotiana tabacum) and PgIREG1

in Arabidopsis are two examples of metal transporter genes that have been genetically modified

(Merlot et al., 2014; Das et al., 2016). Other metal transporter genes include those that encode metal

chelators, metallothioneins (MTs) (Peng et al., 2017), and genes associated with antioxidant machinery

(Peng et al., 2017; Raza et al., 2021). The use of transgenic techniques to increase resistance to metal

oxidation has also been documented. Transgenic hyperaccumulators may be created by manipulating

the antioxidant system to maintain redox equilibrium to avoid the destruction of biomolecules such as

DNA, proteins, and lipids and to maintain the structural and functional stability of cellular structures of

plant under Cr stress (Du et al., 2019). Transgenic plants that overexpress antioxidant genes for SOD,

CAT, and APX with reduced ROS generation under Cr stress have been created to prevent metal

toxicity-induced oxidative stress (Gao et al., 2016). Additionally, enhanced antioxidant systems in

transgenic lines are associated with higher growth performance in terms of photosynthesis, mineral

uptake, maintenance of redox homeostasis, and enzyme activity (Khan et al., 2021). Although

transgenic lines created for over-expression traits do not always show the expected benefits, they can

nevertheless have positive consequences by influencing the alternative tolerance mechanisms.

Techniques use for Chromium Remediation:

The concentration of metals in polluted soils is affected by multiple chemical and biological

attributes (Alengebawy et al., 2021). Soils preserve heavy metals by adsorbent, crystallization,

and chelation; nevertheless, such interactions restrict their mobility and bioavailability (Yan
 et al., 2020). However, the implementation of chemical processes, such as organic and

inorganic modifications in field can complement this natural attenuation process (Mench et al.,

2006; Shahid et al., 2017). These technologies generally minimized the availability of Cr, boost

the fertility of the soil, and increase plant growth (Gavrilescu, 2022). Organic amendments

(compost) possess a significant proportion of humified organic material and may restrict the

availability of Cr in the soil, even though they allow vegetation to be regenerated (Lwin et al.,

2018). On the other hand, phosphate fertilizers are useful in metal inactivation through the

creation of stable mineral phosphate within the inorganic amendments (Ahmad et al., 2019).

Biological options, particularly phytoremediation, have been considered reliable, ecologically

acceptable, and cost-effective replacement to physicochemical approaches for the restoration of

depleted environments. Various physicochemical activities that can be used to eliminate Cr-

polluted environments include ionization, precipitation, reverse osmosis, evaporation, and

chemical reduction (Roy and Bharadvaja, 2021). Moreover, there are numerous issues linked

with these processes, like permeate flux, inflated prices, high energy consumption, and low

extraction efficiency shows that these are less significant in industry. In general, the main

considerations in choosing an acceptable treatment to eliminate metals are technological

applicability, eco-friendly, and cost-effectiveness (Acheampong et al., 2010).

Phytoremediation:

Phytoremediation is a process in which plants are used for remediation of polluted soils

and considered an eco-friendly and green approach (Ali H. et al., 2013; Srivastava et al., 2021).

There are various strategies associated with phytoremediation techniques including

phytoextraction, rhizofiltration, phytovolatilization, biotransformation, rhizodegradation,

phytostabilization, and Phyto restoration (Yan et al., 2020). Phytoextraction is focused on the
 ‘hyperaccumulation’ process, and phytostabilization is focused on the surface complexation

mechanism and both are involved in metal affinity phenomena (Xu et al., 2012).

Phytoextraction and Phyto stabilization are two of those practically and economically viable

solutions for treating metal-polluted soils (Kuiper et al., 2004). Biotransformation is another

term for Phyto-transformation. That is the separation of pollutants absorbed by

plants via internal metabolic pathways or the segmentation of pollutants just outside of the

plant because of plant-generated chemicals (such as enzymes). Plant absorption and

metabolism are the primary components, which result in plant deterioration. The uptake of

contaminants by plant roots and its conversion to a gaseous state, and release into the

atmosphere is referred as phytovolatilization. Volatilization through leaves (ITRC, 2009) is the

phytovolatilization process. Degradation by plant rhizosphere microorganisms is the method

referred as rhizodegradation (Mench et al., 2009). This ecologically accepted technology is

successfully used to fix soils that are polluted by various contaminants. Furthermore,

phytoremediation is increasingly used as a technical alternative to treat contaminated water in

various forms of wetland treatment (Zhang et al., 2010). In crux, phytoremediation is a feasible,

socially, and economically suitable, and eco-friendly solution for the soils polluted with Cr.

Nonetheless, to counteract the health risks due to Cr concentration in edible parts of food crops,

the proportion of Cr in edible parts of food crops should be closely scrutinized.

Microbe associated phytoremediation:

Microbial remediation depends on the ubiquity and diversity of microorganisms, which are able to adapt to a wide

range of adverse conditions and can transform a variety of contaminants into their nutrient source and convert

these contaminants into non-toxic or less toxic substances (Mereuta et al. 2020). Relative studies have shown that

indigenous microorganisms are more effective in remediation of Cr(VI) contamination because they live in Cr-

containing environments for a long time (Carlos et al. 2016; Jalili et al. 2020)
Several methods of metal remediation have been used to address the harmful impacts of metal

contamination, including physical, chemical, and biological processes, to inactive specific

hazardous metals from the atmosphere (Marques et al., 2011). Microbial remediation has

gained significant attention among different biological remediation methods because of its cost-

effectiveness, higher efficacy, and non-expendable technologies (Malaviya and Singh,

2014; Fernandez et al., 2018). Some of the microbes that tolerate Cr establish ability to

minimize the toxicity of Cr(VI) concentration from the atmosphere and thus play a prominent

role in the remediation of Cr(VI) (Table 3; Figure 3). Many investigations on the collection and

profiling of distinct Cr-lowering microbial strains of bacteria have been published in last few

years (Pseudomonas spp., Bacillus spp., Enterobacter spp., Acinetobacter spp,.), fungi

(Aspergillus spp., Penicillium spp., Rhizopus spp.), and yeast

(Candida spp., Saccharomyces spp.) (Chen et al., 2016).

The use of plant growth-promoting bacteria (PGPB) in plants, is also regarded as a

significant and environmentally acceptable method for the removal of heavy metals from soil

(Fahad et al., 2014). These bacteria encourage plants to endure extreme stress and improve

plant nutrition to stimulate plant growth (N, P, Fe) and release different metabolites related to

stresssuch as the production of phytohormones, solubilization of phosphates, and production of

siderophores (Dodd and Perez-Alfocea, 2012). Several studies have documented the use of

plant growth promotion (PGP) rhizobacteria for heavy metal bioremediation,

like Bacillus sp., Pseudomonas sp., etc. (Ndeddy-Aka and Babalola, 2016). Microorganisms

have been found to reduce hexavalent Cr through various means, either by using hexavalent Cr

as the final acceptor of electrons or by releasing some dissolving enzymes (Table 4; Ahemad,

2015). In an experiment, Karthik and Arulselvi (2017) evaluate the effect of Cr(VI) on the plant

growth-promoting properties of potential rhizobacterial strain isolated from the rhizosphere of a

 common bean (Phaseolus vulgaris). The strain AR8 was chosen from 36 rhizobacterial strains

when compared to uninoculated Cr(VI) treated plants, the inoculation of Cellulose

microbiomesfunkei strain AR8 significantly improved the root length of test crops in both the

presence and absence of Cr(VI).

Chemical remediation:

In-situ or ex-situ complex formation through chelating substances has been used for metal extraction

(Di-Palma et al., 2005; Finzgar and Lestan, 2007). The efficacy of extraction depends upon the

availability of readily exchangeable ions in the soil matrix capable of forming strong complexes with

minimum specific chelating agents (Di-Palma, 2009). For removal of maximum amounts of metals

found in polluted soils, phytoextraction may be used, with some portion of the soil metal content freely

available to plants. There are various synthetic chelating components, such as EDTA (ethylene diamine

tetra acetic acid), diethylenetrinitrilepentaacetic acid (DTPA), nitrile triacetic acid (NTA), pyridine-

2,6-dicarboxylic acid (PDA), trans-l,2-diaminocyclohexane-N,N,N0,N0-tetraacetate (CDTA), or

ethylenediaminedisuccinate (EDDS) used for remediation of soil polluted with organic and inorganic

contaminants. To increase the accessibility of metals in soil and the transference of metals from root to

shoot, several ideas have been proposed (Meers et al., 2005). Application of chelating agents

substantially improved the Cr uptake in above-ground biomass of many crops.

Nanotechnology: intro plus importance in different sectors mainly in agriculture

where different nanoparticles are using:

Nanoparticles: intro plus imp (like used as fertilizer, insecticide etc) in differ

sectors and move from general to selenium nanoparticle:

Nanotechnology:

Nanotechnology can be defined as the science and engineering involved in the design, synthesis,
characterization, and application of materials and devices whose smallest functional organization, in at

least one dimension, is on the nanometer scale or one billionth of a meter (Saini et al., 2010). At these

scales, consideration of individual molecules and interacting groups of molecules in relation to the bulk

macroscopic properties of the material or device becomes important, as it has a control over the

fundamental molecular structure, which allows control over the macroscopic chemical and physical

properties (Silva, 2004).

Nanotechnology, contrary to its name, has massively revolutionized industries around the world. This

paper predominantly deals with data regarding the applications of nanotechnology in the

modernization of several industries. A comprehensive research strategy is adopted to incorporate the

latest data driven from major science platforms. Resultantly, a broad-spectrum overview is presented

which comprises the diverse applications of nanotechnology in modern industries. This study reveals

that nanotechnology is not limited to research labs or small-scale manufacturing units of

nanomedicine, but instead has taken a major share in different industries (Malik et al., 2023). Companies

around the world are now trying to make their innovations more efficient in terms of structuring,

working, and designing outlook and productivity by taking advantage of nanotechnology. From small-

scale manufacturing and processing units such as those in agriculture, food, and medicine industries to

larger-scale production units such as those operating in industries of automobiles, civil engineering,

and environmental management, nanotechnology has manifested the modernization of almost every

industrial domain on a global scale (Malik et al., 2023). With pronounced cooperation among

researchers, industrialists, scientists, technologists, environmentalists, and educationists, the more

sustainable development of nano-based industries can be predicted in the future.

Importance of Nanotechnology:

After thorough and careful analyses, a wide range of industries—in which nanotechnology is producing

remarkable applications—have been studied, reviewed, and selected to be made part of this review. It should be
notified that multiple subcategories of industrial links may be discussed under one heading to elaborate upon the

wide-scale applications of nanotechnology in different industries.

Nanotechnology and Computer Industry

Nanotechnology has taken its origins from microengineering concepts in physics and material sciences (Hulla et

al., 2015). Nano scaling is not a new concept in the computer industry, as technologists and technicians have

been working for a long time to design such modified forms of computer-based technologies that require

minimum space for the most efficient work. Resultantly, the usage of nanotubes instead of silicon chips is being

increasingly experimented upon in computer devices. Feynman and Drexler’s work has greatly inspired

computer scientists to design revolutionary nano computers from which wide-scale advantages could be

attained (M.S. El Naschie, 2006). A few years ago, it was an unimaginable to consider laptops, mobiles, and

other handy gadgets as thin as we have today, and it is impossible for even the common man to think that with

the passage of time, more advanced, sophisticated, and lighter computer devices will be commonly used.

Nanotechnology holds the potential to make this possible.

Nanotechnology and Bioprocessing Industries

Scientific and engineering rigor is being carried out to the link fields of nanotechnology with contributions to
the bioprocessing industry. Researchers are interested in how the basics of nanomaterials could be used for the
high-quality manufacturing of food and other biomaterials (Hulla et al., 2015). Pathogenic identification, food
monitoring, biosensor devices, and smart packaging materials, especially those that are reusable and
biodegradable, and the nanoencapsulation of active food compounds are only a few nanotechnological
applications which have been the prime focus of the research community in recent years. Eventually, societal
acceptability and dealing with social, cultural, and ethical concerns will allow the successful delivery of nano-
based bio-processed products into the common markets for public usage (Palit, 2020).
With the increasing population worldwide, food requirements are increasing in addition to the concerns
regarding the production of safe, healthy, and recurring food options. Sensors and diagnostic devices will help
improve the sensitivity in food quality monitoring (Yadav et al., 2021). Moreover, the fake industrial application of
food products could be easily scanned out of a system with the application of nanotechnology which could
control brand protection throughout bio-processing (McNeil, 2005). The power usage in food production might
also be controlled after a total nanotechnological application in the food industry. The decrease in power
consumption would ultimately be positive for the environment. This could directly bring in the interplay of
environment, food, and nanotechnology and would help to reduce environmental concerns in future.
Nanotechnology and Food Industry
The applications of nanotechnology in the food industry are immense and include food manufacturing,

packaging, safety measures, drug delivery to specific sites (Yu et al., 2018), smart diets, and other modern

preservatives. Nanomaterials such as polymer/clay nanocomposites are used in packing materials due to their

high barrier properties against environmental impacts. Similarly, nanoparticle mixtures are used as antimicrobial

agents to protect stored food products against rapid microbial decay, especially in canned products. Similarly,
several nanosensor and nano-assembly-based assays are used for microbial detection processes in food

storage and manufacturing industries (Biswas et al., 2022).

Nanotechnology and Agri-Industries

Agriculture is the backbone of the economies of various nations around the globe. It is a major contributing
factor to the world economy in general and plays a critical role in population maintenance by providing nutritional
needs to them. As global weather patterns are changing owing to the dramatic changes caused by global
warming, it is accepted that agriculture will be greatly affected (Saini & Bhatt, 2020). Under this scenario, it is
always better to take proactive measures to make agricultural practices more secure and sustainable than
before. Modern technology is thus being employed worldwide. Nanotechnology has also come to play an
effective role in this interplay of sustainable technologies. It plays an important role during the production,
processing, storing, packaging, and transport of agricultural industrial products (Usman et al., 2020).
Nanotechnology has introduced certain precision farming techniques to enhance plant nutrients’
absorbance, alongside better pathogenic detection against agricultural diseases. Fertilizers are being improved
by the application of nano clays and zeolites which play effective roles in soil nutrient broths and in the
restoration soil fertility (Usman et al., 2020). Modern concepts of smart seeds and seed banks are also
programmed to germinate under favorable conditions for their survival; nano polymeric mixtures are used for
coating in these scenarios (Vijayakumar et al., 2022). Herbicides, pesticides, fungicides, and insecticides are
also being revolutionized through nanotechnology applications. It has also been considered to upgrade linked
fields of poultry and animal husbandry via the application of nanotechnology in treatment and disinfection
practices.
Comparison between selenium in bulk and Se as nanoparticle:

Remediation by the nanoparticles:

Nano-remediation is an eco-friendly and cost-effective method of detoxifying heavy metals in soil and

other environments using nanoparticles (NPs) (Ahmed et al., 2021a; Wei et al., 2022a; Table 6). By

absorbing heavy metals, lowering the hazardous valence to a stable metallic state, and accelerating the

reaction, this unique remediation strategy has been demonstrated to be efficient in the removal of toxic

heavy metals (Mondal et al., 2020). Synthesis of nZVI NPs in colloidal solution using green tea extract

having an average particle diameter of 5-10 nm with polyphenol coating (which served as a capping

and reducing agent) was significantly effective in remediating Cr(VI) from groundwater passing

through porous soil beds (Mystrioti et al., 2014). Synthesis of NPs by using various rose apple

(Syzgiumjambos L.), candlenut tree (Aleuritesmoluccanus L.), and oolong-tea leaves extracts were

significantly remediate Cr(VI) from aqueous medium up to 90% at initial 5 minutes, due to its

maximum NPs antioxidant property, but complete removal took after 60 minutes (Xiao et al., 2016).
The removal effectiveness of Cr(VI) was greatly influenced by factors i.e., Cr(VI) initial concentration,

NPs dosage, solution pH, and temperature. For a constant concentration of Cr, the availability of active

sites rises with increasing NPs dosage, which improves the removal rate (Xiao Z. et al., 2017).

Use of organic amendments for remediation

Organic material is facilitated in soil deposition of Cr, according to Branzini and Zubillaga (2012). We

postulated that soil comprising most of the humidified organic material had a lesser Cr accessibility,

which would minimize Cr deposition in plants. The use of organic modifications in Cr polluted soils

and their impact on reducing Cr absorption in plants have been reported in several studies.

Biochar is produced in a low oxygen atmosphere through the combustion of carbonaceous material

collected from a range of sources (Tomczyk et al., 2020). Biochar has a higher porosity, extensive

functional groups containing oxygen over its microscopic layer, and acts as an adsorbent to sequester

heavy metals in soil (Xu and Fang, 2015). It has a larger surface area, a higher negative and stronger

surface charge, biochar has higher absorption properties as compared to raw organic soil materials.

Thus, biochar application enhances water holding capacity, reduce nutrient losses, and improve soil

structure. In addition, biochar-containing soils have resemblance to organic pollutants (Yu et al.,

2009; Haider et al., 2022a). Integrating biochar with other soil amendments before tillage activity, such

as manure fertilizer, compost, or lime, will enhance sustainability by cutting down the amount of

tillage practices needed (Haider et al., 2022b; Haider et al., 2022c). Biochar improves nutrient uptake

by preventing their loss by leaching (Major, 2009). The incorporation of biochar minimizes the

availability of Cr and its accumulation and toxicity in plants.

Add about mulching process then add about aloe vera:

Selenium/ Selenium Nanoparticles and heavy metals:

Selenium is a vital ingredient for plants but harmful to many other living things, including people,

bacteria, and the majority of chlorophyte species (Lobanov et al., 2009). Even Se is essential for the

prevention of heavy metal toxicity, the promotion of immunity, and the resistance to oxidative stress

(Conte et al., 2015). Metalloid poisoning is now becoming a major issue for people and the environment

(MohdAsgher et al., 2022). Metal toxicity in agricultural soils is mostly brought about by anthropogenic

activities on a global scale. As they build up at higher trophic levels of the food chain

(biomagnification), heavy metals are not biodegradable and pose a harm to biotic forms and their

environment (Guo et al., 2013).

Abiotic stressors, including as nutrient deficiencies, excessive light (UV-B), drought, high carbon

dioxide (CO2), ozone (O3), heat, cold, and chilling, are widely known flooding, soil compaction,

salinity, freezing, and Heavy metals can have disastrous or negative effects influences plant growth,

yield, and consequently the agricultural output (Suzuki et al., 2014).

Selenium/Selenium nanoparticles effects on plants under heavy metal stress:

One of the primary abiotic factors affecting crop yield is heavy metal (HM) contamination, which

also poses a concern to human health when consumed as food. Due to their expanding use for

commercial purposes, negative effects on plants and other life forms, and threat to human existence,

HMs have attracted a lot of attention in recent years (MohdAsgher et al., 2023).

But a number of strategies have been used recently to counteract the negative effects of HMs.

Researchers have more recently looked at the usage of mineral supplements like selenium (Se) in the

reduction of HM stress after phytohormones (MohdAsgher et al., 2023).

The antioxidant effects of selenium in both plants and animals make it an essential mineral.

Exogenous Se enhances the antioxidant system, reduces metal uptake and translocation, and confers

resistance to HM toxicity in plants (MohdAsgher et al., 2023).

Additionally, Se controls the synthesis of different osmolytes in cells, aiding in the development of

cell osmolarity. Various secondary metabolites are also produced by selenium as part of the plant's

defensive systems under various conditions. One of the most important processes for plant growth

and development is the uptake of mineral nutrients, which is also favorably associated with Se under

metalloid toxicity (MohdAsgher et al., 2023).

One of the major barriers to assessing the yield of crops globally is heavy metal (HM) pollution (Md.

Mahadi Hasan et al., 2019). Around the world, the issue of HM stress has emerged as a major concern

in a number of soil and aquatic ecosystems (Alsafran et al., 2022). Inorganic substances known as

heavy metals are non-biodegradable, persistent, and have mutagenic, cytotoxic, and genotoxic effects

on people, plants, and animals. They penetrate food chains and contaminate ecosystems of soil and

water (Jitendra Kumar Sharma et al., 2023).

25 selenoproteins have been found, and their physiological functions for lower plants (including

bacteria and algae), mammals, and human metabolism have been discovered (Kumar and Priyadarsini

2014). In order to maintain the redox potential of cells, se has been identified as a component of

selenoenzymes such as glutathione peroxidase (GPx), thioredoxin reductases (TR), and proteins with

unidentified roles (Rayman 2000). There have been several experimental attempts to identify these

selenoproteins in higher plants, but this is still an unanswered question (Mora et al. 2015).

In addition to mineral elements, plants also need a small number of HMs for regular metabolic

processes. Se is a crucial element that has been created to protect crops from metal toxicity because of

its many advantageous functions and capacity to make plants more tolerant to heavy metals. One of

the major mechanisms directly influencing the growth and development of plants is mineral nutrition.

Under stressful circumstances, plants create a variety of signaling molecules to combat the stress(Md.

Mahadi Hasan et al., 2021). One essential ingredient for plants is selenium. A widely dispersed trace

element called selenium has both hazardous and advantageous impacts on people, plants, and
animals. In both favorable and unfavorable environments, se may be beneficial for plant growth and

development at low concentrations(Masanori Tamaoki et al., 2008). Se can clump together on the

surface of water or soil, which can make it poisonous to plants. Because of this, the majority of plant

Se resistance research has focused on Se interaction with sulfur (S) metabolisms(Masanori Tamaoki

et al., 2008). By synthesizing numerous growth regulators, the addition of Se restricts the uptake of

metals by roots and their transfer to shoots, one of plants' essential stress-tolerance

processes(MirzaHasanuzzaman et al., 2022).

A suitable concentration of Se can stimulate auxin levels, postpone senescence, enhance

photosynthesis, increase antioxidant capacity, and promote plant development (ÖzgürÇakır et al.,

2015). By detoxifying ROS-induced damages through control of the activities of several antioxidant

enzymes, selenium administration at the optimal concentration enhanced the development and

biomass of Cucumis sativus plants under heavy metals stress (Cd, Pb, Ni). In Al-treated

Loliumperenne, application of 2 M Se increased root development and lowered oxidative

stress(Cartes et al., 2010). By preventing As from being absorbed, exogenous treatment with Se

decreased the toxicity that As caused in Phaseolus aureus, and by activating the antioxidant system

under toxic conditions, it enhanced plant development(Malik et al., 2010). However, the application

of Se to Viciafaba plants under Pb stress decreased the production of ROS through an increase in the

activity of several antioxidant enzymes (Mroczek-Zdyrska et al., 2016).

Se/Se nanoparticles reduces intake of metal/metalloids:

By recovering their transportation from lower plant to upper regions, selenium limits the

uptake and metabolism of harmful metal ions (Alyemeni et al., 2018; Hasanuzzaman et al.,

2020). For instance, such Se-induced lower Cd absorption by roots, together with reduced

translocation into shoots and leaves, had been found in Cd-stressed tomato seedlings
(Alyemeni et al., 2018).

Additionally, Se modifies the translocation capacity of many rice cultivars by decreasing root

to stem and stem to grain translocation and enhancing stem to leaves translocation (Gao et al.,

2018). By enhancing pectin and hemicellulose, supplemental selenium can improve the

binding of metal ions to the root cell wall. It can also change the subcellular distribution of

roots, with metal contents being decreased in plastids and mitochondria and increased in

vacuolar sap and ribosomes, for example (Zhao et al., 2019). Additionally, Se's protective role

influences phytochelatin (PC) production, the development of harmless Se-metal complexes,

and the reduction of metal transport from roots to aerial plant parts to reduce metal toxicity

(Hawrylak-Nowak et al., 2014).

By changing the root shape of B. campestris sp. Pekinensis, exogenous application of Se

decreased the concentration and influx of Cr (Zhao et al., 2019). Ulhassan et al. (2019) found

that applying Se to B. napus caused a reduction in the accumulation of Cr in plant tissues.

According to (Hawrylak-Nowak et al. (2014), Se-treated plants showed enhanced

translocation toward shoots and a decrease in Cd accumulation and phytochelatins in

cucumber roots. Contrarily, several investigations have shown that adding Se does not

significantly affect the accumulation of Pb in beans grown in the field (Mroczek-Zdyrska and

Wójcik, 2012).

Heavy metal absorption is influenced by se application in a time-dependent way. For instance,

Se supplementation increased Cd absorption in rice cultivated under Cd stress, but Se uptake

decreased with increased Cd exposure (Wan et al., 2016). Even among the cultivars of the

same plant species, Se can cause a reduction in the uptake of heavy metals in plants. In three

rice cultivars, WYHZ, NJ5055, and ZF1Y, Si and Se alone and in combination were tested by

Gao et al. (2018) for their effects on Cd uptake.

 Se/Se nanoparticles enhances the physiology of plants and growth in

metal/metalloid toxicity:

Selenium is essential for defending plants against the harmful effects of metals and metalloids

(Feng et al., 2020; Riaz et al., 2021). Plants exposed to metals or metalloids frequently exhibit

a tendency to slow their growth. Numerous studies have shown that Se, delivered in a variety

of ways and at various concentrations to crop plants grown under heavy metal/metalloid

stress, promotes plant growth and enhances physiological processes. The effects of 2 or 6 M

of various forms of Se (selenite IV and selenate VI administered as Na2SeO3 or Na2SeO4) on

lettuce plants exposed to Ni (5 and 10 M) were assessed by Hawrylak-Nowak and Matraszek-

Gawron (2020).

Se increased photosynthesis and changed anatomical characteristics, according to Alves et al.

(2020). As a result of reduced phytotoxicity in B. napus, increased growth, biomass,

photosynthetic pigments, leaf gas exchange, and Fv/Fm were observed (Ulhassan et al., 2019).

Following the application of Se to wheat plants, Khan et al. (2015) showed a reduction in Cd-

induced oxidative stress. According to Sun et al. (2016), Se administration reduced the

phytotoxic effects of Cd on cucumber plants by controlling proteins and processes associated

with the stress response, such as the glycolysis pathway and the nitrate absorption pathway,

which may promote Se-induced Cd tolerance. According to Chauhan et al. (2020), increased

activity of glutathione S-transferases (GST), glutaredoxins, peroxiredoxins, and heat shock

proteins improved As-induced oxidative stress in mice.

Selenium/Se Nanoparticle Uptake and Translocation:

Selenium is a metalloid occurring naturally in sedimentary rocks in dry areas of the world.

The inorganic forms of Se i.e., Selenite (SeO32-) and Selenate (SeO42-), are the two main
forms of Se which remain available to plants through soil. Selenite and selenate are present in

anoxic and toxic soils (White, 2018). Selenium chemically resembles sulfur(S), hence taken up

inside the plants via S transporters located in the root plasma membrane (Gupta & Gupta, 2017).

Many species of plants have been identified as Se hyperaccumulators which can accumulate

high concentrations of Se, such as Stanleyapinnata(Parker et al., 2003). Selenium uptake differs

from species to species, soil type, soil concentrations, and form of Se supplied (Zayed et al.,

1998). Selenate absorption by plants is a well-established mechanism. The high-affinity

sulphate transporters help in the absorption of selenate by roots. However, the selectivity of

various plant species for sulphate and selenate varies (White, 2004). On the other hand, nothing

is understood about the process behind selenite uptake by plants. It has been observed that

plant roots absorb selenite through passive diffusion (Arvy, 1993). Several species of plants

belonging to the families Asteraceae, Brassicaceae and Fabaceae absorb higher concentrations

of Se in hair-like epidermal appendages called trichomes (Freeman et al., 2006). The

metabolism of Se in higher plants is closely associated with that of S due to their chemical

analogy. Both ions SeO32- and SeO42- are absorbed by the plants via roots, and SeO42- is

taken via sulphate transporters (SULTRs). This relationship is demonstrated by the fact that

Se is taken up by the plants via sulphate transporters and SeO32- via aquaporins and

phosphate transporters (Brown & Shrift, 1982). However, considerable differences in the uptake

and transport mechanisms of inorganic forms are found in plants. Selenate is uptake by plant

roots in an active manner and leads to increased Se concentrations in the roots.

Chapter 3:

Material and methods:

3.1 Experimental Site and Design

A pot experiment will be performed to evaluate the sole and synergetic effectiveness of soil
applied Aloe Vera gel and selenium will be applied as foliar to reduce the toxicity of Cr in spinach at
the Department of Environmental Sciences, University of Lahore, Pakistan. Completely randomized
design (CRD) under factorial arrangement will be applied with triplicates

3.2 Experimental Treatments

The spinach plants will be grown on Cr-contaminated soil. The experiment will be comprised of the
following treatments:

Factor A: Aloe vera gel (Seed priming)

Without Aloe vera gel (Half pots)

With Aloe vera gel (Half pots)

Factor B: Se NPs (Foliar)

Se NPs (0, 0.5, 1.0, 1.5 mg/L)

3.3 Preparation of Aloe vera gel

Fresh leaves of the Aloe Vera will cut and washed to remove dirt from surface of the leaves.
Then cut leaves with knife and collect internal gel of aloe Vera in a beaker. Peel of aloe Vera leaves
and blend the gel in a blender. After blending boil the gel at high temperature and place it at room
temperature to cold. Now aloe Vera gel is ready to use. Mix this blended gel in to soil and place it for a
week.

3.4 Plant Material and Growth Conditions

Random soil samples will be taken from the study site at a depth of 0 to 30 cm (clay sandy
loam), will be collected air dried or then passed via a 2 mm sieve. Mixed culture of soil and aloe vera
gel treatment, aloe vera gel (10% w/w) will also be prepared separately and transferred into plastic
bottles as per treatments. Spinach seeds will be dip in diluted solution of aloe vera gel and remain for
2-3 days until spinach seeds start to germinate.
Spinach seeds will be dip in diluted solution of aloe vera gel of different concentrations and
placed in petri plates for 2-3 days. All petri plates will be kept moist until the spinach seeds germinate.
Petri plates having no aloe vera gel treatments will be considered as control. Five seeds will be dip in
one petri plate and thinned to 5 seedlings per petri plate when shoots are about 5 cm tall. All petri
plates will keep in the open space under normal circumstances until the growth of the spinach seeds.
Manual hoeing will be done to homogenize the Cr contamination right before applying the aloe vera
gel solution and again two days later. Every petri plate will be checked every day, and when necessary,
deionized water and other cross-cultural tasks (such as weeding) were done. After seed germination the
grown seeds will be transferred to pots that have Cr-contaminated soil. Recommended dose of NPK
will be applied as source of nutrients. Foliar applications of Se (as prescribed) will be used, using
solution of 500 mL in each of the two sprays at a regular interval (Khosa et al., 2022). After harvesting,
data of biochemical morphological and physiological attributes will be recorded.

3.5 Data Collection Methods

3.5.1. Growth Parameters

Plants will be harvested and will be separated into roots and shoots in order to evaluate the
growth traits. The total number of leaves will be estimated before separating the leaves and roots. The
measuring scale will be used to find the plant's height, width of leaf, and length of leaf. The area of the
leaf will be computed by multiplying the leaf width and leaf length. The soil that has adhered to the
plants will be washed off with distilled water before they are allowed to air dry. Using an analytical
balance, the fresh weight of the shoot and roots will be determined. For 48 hours, the roots and leaves
will be dried in an oven at 70 °C. For estimating the individual dry weights of the root and leaves.

3.5.2. Photosynthetic parameters

According to Emanuil et al. (2020), 35 days after sowing, photosynthetic characteristics will be
calculated from the uppermost leaves of plant by using a transportable infrared gas analyzer (ADC
Bioscientific, Hoddesdon, UK) at increase the intensity of absorption from 09:00 to 12:00.

3.5.3. Oxidative Stress Parameters

 For the purpose of extracting pigment, a crushed leaf sample (5 g) will be introduced to the test
tube that contains 85% of acetone (v/v) then left for 24 hours. The sample will then undergo a 10-
minute centrifugation at 4000 g and 4 °C. The chlorophyll contents of the supernatant will be
determined by using the spectrophotometer (Halo DB-20/DB-20S; Dynamica Scientific, Newport
Pagnell, UK) between different wavelengths of 470, 647, and 664.5 nm, in accordance with
Lichtenthaler (1987). Electrolyte leakage (EL) level will be measured after tiny parts of leaves are
dipped into the deionized water. First reading of EL will be noted right after the incubation of the
sample at 32 °C for 2 h, and the second reading will take after incubation at 121 °C for 20 min
(Dionisio-Sese & Tobita, 1998).

EL = EC1/EC2 × 100
Where EC1 is electrical leakage at initial stage and EC2 at final stage (Dionisio Sese and Tobita,
1998).
MDA concentration in the leaves of both crop varieties were measured using grinded leave samples
(0.5 g) into the TCA solution. The readings were measured at 532 nm and 600 nm on pectrophotometer
for the measurement of MDA in plant leaves.
Hydrogen peroxide (H2O2) contents in the leaves of both rice and wheat varieties were measured. First
of all, grinded leaves samples (0.25 g) were put into 5 ml TCA solution. Afterward, samples were
centrifuged at 12,000 rpm for 15 min and cooled down for 20 min and following solutions were added
in 0.5 ml supernatant solution putted in the test tubes. Then 0.5 ml buffer having pH 7 was added to the
test tubes with 1 ml KCl (1 M) solution. The reading of hydrogen peroxide was estimated at 390 nm
using spectrophotometer.
3.5.4. Enzymatic Oxidation Activities
The fresh leaves of spinach will be extracted in phosphate buffer (pH 7.8), homogenized, and
centrifuged to determine the enzyme activity. For this experiment a supernatant will be used. Measure
the activities of peroxidase (POD), superoxide dismutase (SOD) and catalase (CAT), will be made
accordance with those methods that are published by Velikova et al. (2000), Aebi (1974), and
Beauchamp and Fridovich, (1971), respectively.
Measurement of the activities of enzymatic antioxidants (SOD, POD, CAT and APX):
Buffer solution was made using potassium di-hydrogen phosphate (6.8 g per liter) and Di-potassium
hydrogen phosphate (8.7 g per liter) to grind the leaves samples (0.5 g) of both crop varieties and pH
was maintained up to 7.8. It was put into falcon tubes and centrifuged at 4000 rpm for 20 min. The
reading of catalase enzymes in leaves of both crop varieties were taken at different time periods 0 S, 30
S, 60 S, 90 S, 120 S, 150 S and 180 S by following Aebi (1984). Similarly, SOD values in the leaves of
different crop varieties were measured by taking 400 μl hydrogen peroxide, 25 lL buffer, 100 μl
Tritoen, 50 μl NBT, 50 μL sample and 50 μl riboflavin into the test tubes and reading was measured at
560 nm using spectrophotometer by following Zhang and Kirkham (1994).
The POD values in the leaves were measured by taking 13.4 μl hydrogen peroxide into 20 ml buffer
and 33.3 μl of glycerol into 15 ml buffer. After the preparation of these solutions, 100 μl from these
two solutions were taken and put into the test tubes. About 50 μl of centrifuge solution of leaves of
both crop varieties were put into the prepared test tube solutions. Then, phosphate buffer (750 μl) was
put in each test tube. After the addition of all these solutions the reading of POD enzyme was measured
at 470 nm following Zhang and Kirkham (1994).
APX values were measured by using spectrophotometer. Leave samples of both crop varieties were
grinded into liquid nitrogen. Phosphate buffer (0.05 M) having pH 7.8 was used to standardized the
samples. It was centrifuged at 12,000 x rpm for 10 min at 4 ◦C. The APX activities were noticed as
reported by Nakano and Asada (1981).
CAT

3.5.5. Chromium Accumulation

A blend of di-acid (HNO3: HClO4 proportion of 2:1) will be used for the wet digestion of
spinach shoot samples. To analyze the concentration of chromium content in the shoot samples the
Atomic absorption spectrophotometer (Perkin-Elmer, Model 3300) will be used.

3.5.6. Statistical Analysis

Data will be analyzed statistically through Fisher’s ANOVA and compare the means of all
treatments via highest significant differences (HSD) test using Statistix 8.1 software.
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