Foliar Nourishment with Different Zinc-Containing Forms Effectively Sustains Carrot Performance in Zinc-Deficient Soil

Abstract

Among the essential micronutrients, zinc (Zn) affects vital functions in crop plants. The influences of foliar nourishing with certain Zn-containing forms on the growth, productivity, and physiology of carrot plants (cv. Fire wedge F1) and their nutritional contents when grown in Zn-deficient soil were examined in both 2019/2020 and 2020/2021 field trials. Two doses of zinc oxide nanoparticles (ZnO-NPs(1) = 20 and ZnO-NPs(2) = 40 mg L⁻¹), zinc–EDTA (Zn–EDTA(1) = 1 and Zn–EDTA(2) = 2 g L⁻¹), or bulk zinc oxide (ZnO-B(1) = 200 and ZnO-B(2) = 400 mg L⁻¹) were applied three times. The data outputted indicated, in general, that ZnO-NPs(2) were the best treatment that conferred more acceptable plant growth (measured as shoot length and fresh and dry weights), physiology (measured as cell membrane stability index, SPAD readings, and nutrient uptake), and nutritional homeostasis (e.g., P, Ca, Fe, Mn, Zn, and Cu contents). All these positive attributes were reflected in the highest yield, which was measured as fresh weight, dry matter, length, diameter, volume, and total yield of carrot roots. However, there were some exceptions, including the highest membrane stability index in both seasons, the highest Cu uptake and Mn content in the first season, and root fresh weight in both seasons obtained with ZnO-NPs(1). Moreover, the maximum P uptake and root dry matter were obtained with ZnO-B(1) and the highest content of root P was obtained by ZnO-B(2). Based on the above data, foliar nourishing with ZnO-NPs(2) can be recommended for the sustainability of carrot cultivation in Zn-deficient soils.

1. Introduction

Most developing countries depend on agriculture as their main source of food and other essential uses. Therefore, agriculture is the backbone of the national income of these countries, including Egypt. In addition, the excessive growth of the world’s population, which is expected to approach 10 billion by 2050, requires an increase in agricultural productivity by at least 50% to face the problem of steadily increasing population and achieving food security [1]. In this context, among food crops, carrot (Daucus carota L.), as the most economically important vegetable crop for exportation and local consumption in Egypt in recent years, may contribute to food security. Worldwide, carrot production is approximately 24 million tons [2]. Attempts have been made to grow this crop successfully to achieve high yields through applying some strategies, including integrated management of fertilization, especially zinc (Zn), due to the large number of Zn-deficient soils in Mediterranean countries because of the abundance of calcareous soils and soils with a high pH [3,4]. Since Zn is highly recommended for all root vegetable crops, its deficiency greatly affects the yield of carrot and represents a widespread health risk influencing a large population in developing countries [5,6].

It is necessary to apply Zn using modern technologies that can support the agricultural field to increase the productivity of these crops [7]. Nanotechnology (NT) is an emerging field with huge potential for innovation in agriculture and related fields. NT in agriculture is currently focusing on targeted agriculture using nutrients as nanoparticles (-NPs) with unique properties to increase productivity [8]. Recently, Zn has been applied as a foliar nourishment for some crop plants (e.g., sorghum, soybean, and rice) in various forms, including zinc oxide nanoparticles (ZnO-NPs), bulk zinc oxide (ZnO-B), and zinc salts (Zn²⁺); however, the ZnO-NPs form outperformed the other forms in improving plant growth, physiology, biochemistry, antioxidant defense system, and productivity under adverse stress conditions [9,10,11].

Micronutrients play important roles in plant growth, physiological processes, yield, and quality. Moreover, all micronutrients are essential for human health, yet more than 3 billion people worldwide suffer from micronutrient deficiencies, such as Zn, iron (Fe), manganese (Mn), and copper (Cu), which have become performance-limiting factors, and deficiency of these micronutrients in human food is responsible for low nutrition. Generally, plants uptake Zn as a divalent cation (Zn²⁺). It is an essential micronutrient for metabolic activities such as carbohydrates, fats, nucleic acids, and protein synthesis, as well as degradation and the normal growth and development of plants [12]. It has fundamental functions in the synthesis of auxin or indole acetic acid (IAA) from the amino acid called “tryptophan” and also in the biochemical reactions needed to form the structure of chlorophyll and carbohydrates. In addition, the yield and quality of different crop plants can be affected by Zn deficiency [13]. Zn has been found to play an important role in controlling reactive oxygen species (ROS) and in protecting plant cells from oxidative stress [14]. It is also a nutrient that is integral to the structure of many enzymes and is the only one represented in all six classes of enzymes, i.e., oxide reductase, transferases, hydrolases, layases, isomerases, and ligases. Therefore, Zn deficiency in soils is a serious global problem. It is estimated that about 60% of the world’s soil is unfit for agriculture due to nutrient deficiency and the unavailability of some trace nutrients [15].

A reasonable number of studies have not been performed evaluating different doses of ZnO-NPs, Zn–EDTA, and ZnO-B in relation to carrot plant growth, physiology, nutrient absorption, and productivity under zinc deficiency conditions. Therefore, this study aimed at examining whether a Zn form (e.g., ZnO-NPs, Zn–EDTA, or ZnO-B) would outperform the other forms in improving the yield of carrot plants when grown in Zn-deficient soils. The potential enhancing influences of the best applied form of Zn on plant growth (e.g., shoot length and fresh and dry weights), physiology (e.g., cell membrane stability index, SPAD readings, and nutrient uptake), nutritional homeostasis (e.g., P, Ca, Fe, Mn, Zn, and Cu contents), and root yield and quality (e.g., fresh weight, dry matter, length, diameter, volume, and total yield) were examined.

2. Materials and Methods

2.1. Location, Climatic Conditions, and Plant Material for Experiments, and Soil Properties

The current study was accomplished at the field level utilizing a sandy loam soil located at Fayoum University Farm Station (29°170′ N; 30°530′ E), Egypt. The seasons of 2019/2020 and 2020/2021 were the two trial seasons, which started on 30 September 2019 and 23 September 2020, respectively, by sowing seeds (cv. Fire wedge F1, supplied from Takii Co., Kyoto, Japan) of carrot (Daucus carota L.) and terminated on 11 January 2020 and 20 January 2021, respectively, by harvesting carrot roots. Average weather data for the studied region for the two seasons from September to January are offered in

Table 1. Average weather data for the Fayoum region in Egypt during the carrot growing seasons.

Month	Day °C	Night °C	RH (%)	AWS (ms−1)	AM-PEC-A (mm d−1)	AP (mm d−1)
September	30.1	15.8	43.7	2.1	8.64	0.00
October	25.1	13.9	43.0	2.0	6.61	0.00
November	23.3	12.8	40.5	2.2	4.63	0.02
December	17.2	10.1	53.1	2.3	3.49	0.15
January	16.5	7.6	46.7	1.5	3.43	0.25

Day °C = Average day temperature, Night °C = Average night temperature, RH = Average relative humidity, AWS = Average wind speed, AM-PEC-A = Average of measured pan evaporation class A, and AP = Average precipitation.

.

The soil texture tested in both seasons was sandy loam. Soil samples were collected from the upper surface layer with a depth of 30 cm to analyze the most important properties of soil physics and chemistry [16] as offered in

Table 2. Some soil physical and chemical characteristics of the experimental farm.

Soil Property		2019/2020 Season	2020/2021 Season
Particle size distribution
Sand (%)		66.8	65.6
Silt (%)		16.5	17.8
Clay (%)		16.7	16.6
Soil texture		Sandy loam	Sandy loam
pH in soil paste		7.77	7.59
ECe (dS m−1) in soil paste extracted		4.24	3.95
Organic matter (%)		0.98	0.90
CaCO3 (%)		10.50	10.80
Soluble ions (mmol L−1)
CO32−		-	-
HCO3−		2.70	2.03
Cl−		25.60	21.10
SO42−		18.30	20.30
Ca++		7.47	7.11
Mg++		6.98	4.03
Na+		31.30	31.6
K+		0.88	0.65
Total N (mg kg−1)		515	450
Available-P mg kg−1 (Extractable with NaHCO3, pH = 8.5)		4013	3424
Available-K mg kg−1 (Extractable with NH4AOC, pH = 7.0)		1237	1816
Fe	(mg kg−1) (Extractable with DPTA)	4.15	6.03
Mn		10.7	18.2
Zn		0.04	0.07
Cu		0.40	0.68

.

2.2. Treatments, Design of the Experiments, Sowing, and Intercultural Operations

Two doses of three forms of Zn were prepared for foliar nourishment of Zn deficiency-stressed carrot plants. Zinc oxide nanoparticles were applied at 20 mg L⁻¹ (ZnO-NPs(1)) and 40 mg L⁻¹ (ZnO-NPs(2)), zinc–EDTA was applied at 1 g L⁻¹ (Zn–EDTA(1)) and 2 g L⁻¹ (Zn–EDTA(2)), and bulk zinc oxide was applied at 200 mg L⁻¹ (ZnO-B(1)) and 400 mg L⁻¹ (ZnO-B(2)). ZnO-NPs, Zn–EDTA, and ZnO-B were secured from Sigma-Aldrich, St. Louis, MO, USA. Plants were foliarly nourished with all spraying solutions three times; 28, 48, and 68 days after sowing (DaS) using a 20-L back sprayer. All plants (n = 140) in each plot were sprayed to runoff with 4.25, 5.10, and 6.80 L of spraying solution (e.g., 25, 30, and 40 mL plant⁻¹) in the three spraying times, respectively. The description of all experimental treatments is offered in

Table 3. The description of the experimental treatments.

Treatment	Dose	Replications	Spraying Times
Control	0 mg L−1 (spraying with distilled water)	Three times in three plots of 10.5 m2 each.	All sprays were performed three times at 28, 48, and 68 days after sowing.
ZnO-NPs(1)	20 mg L−1
ZnO-NPs(2)	40 mg L−1
Zn–EDTA(1)	1 g L−1
Zn–EDTA(2)	2 g L−1
ZnO-B(1)	200 mg L−1
ZnO-B(2)	400 mg L−1

. ZnO-NPs were subjected to transmission electron microscopy (TEM), and the morphological structure of ZnO-NPs (≤100 nm) is illustrated in Figure 1.

The area assigned to the experiments was divided into 21 plots (3.0 m × 3.5 m = 10.5 m² for each) that were allocated to 7 treatments (Table 3), and each treatment was repeated three times in three plots. There were four rows of 3.0 m length in each plot and 42–43 hills for sowing seeds in each row. Three seeds were sown in each designated hill with a distance of about 7 cm between each two hills. Four weeks after sowing, the plants were thinned to one hill⁻¹. The carrot plants were provided with different fertilizers (N, P, and K) based on the recommendations of the Egyptian Ministry of Agriculture. N, P, and K fertilizers were added in the form of (NH₄)₂SO₄ (20.6% N), CaH₆O₉P₂ (15.5% P₂O₅), and K₂SO₄ (48–50% K₂O) at levels of 200, 300, and 400 kg ha⁻¹, respectively. All fertilizers were applied at planting, 30, and 60 DaS in three equal portions in the two growing seasons.

2.3. Evaluation of Growth and Physiological Parameters

Average plant height and root length were measured (in cm) using a graduated ruler. Leaf fresh and dry weights plant⁻¹ (g) were assessed using a digital electric balance. For the rapid, accurate, and non-destructive measurement of leaf chlorophyll concentration, the SPAD-502-meter handheld device was utilized. Measurements with the SPAD-502 meter produce relative SPAD meter values that are proportional to the amount of chlorophyll present in the leaf. Relative SPAD meter values were evaluated using uniform leaves on five selected plants from each treatment.

After excluding leaf midrib, a duplicate 0.2 g leaf sample was taken in test tubes with 10 mL of deionized water to determine leaf membrane stability index (MSI) [18]. At 40 °C, a sample was heated for 30 min with a water bath. Solution electrical conductivity (EC₁) was taken. At 100 °C, the other sample was boiled for 10 min. Solution conductivity (EC₂) was also measured. The percentage of membrane stability index (MSI) was calculated by using the following equation:

MSI (%) = [1 − (EC₁/EC₂)] × 100

(1)

All previous measurements were performed using five randomly selected plants.

2.4. Evaluation of Leaf and Root Nutrient Contents

Random samples of the upper fourth leaf and root were collected using five selected plants from each treatment, washed with distilled water, and oven dried at 70 °C to constant weight to determine the contents of P, Ca, Fe, Mn, Zn, and Cu. Inductively coupled plasma–optical emission spectrometry (ICP-OES, Perkin-Elmer OPTIMA-2100 DV, Norwalk, CT, USA) was used to determine those elements in the Laboratory of Soil Chemistry, Faculty of Agriculture and Natural Resources, Aswan University, Aswan, Egypt.

Anti-radical power (ARP) content of roots was determined according to the widely used decolorization assay of Brand-Williams et al. [19]. An appropriate volume of the root extract was mixed with 90 M DPPH^• in methanol, making up final volume of 3.0 mL. The mixtures were shaken vigorously and were stored in the dark for 30 min at room temperature. The decrease in absorbance of the reaction mixtures regarding the control was monitored spectrophotometrically at 515 nm. To determine β-carotene content, the root samples were thawed in the dark at 4 °C to avoid carotenoid oxidation. A total of 2 mL of acetone: hexane (4:6) solvent was added to 0.1 g of root tissue homogenate and mixed in a test tube; then, the mixture was centrifuged at 10,000 rpm for 15 min at 40 °C. Automatically, 2 phases separated and an aliquot was taken from the upper solution, and the absorbance readings (A) were taken at 663, 645, 505 and 453 nm using UV–Vis spectrophotometry. β-Carotene content was calculated using the following equation [20]:

β-carotene (mg 100 mL^–1 extract) = 0.216 × A₆₆₃ − 1.22 × A₆₄₅ − 0.304 × A₅₀₅ + 0.452 × A₄₅₃

(2)

All previous measurements were performed using five randomly selected plants.

2.5. Measurement of Yield and Its Components

Average root fresh and dry weights (g) were measured using a digital electric balance. Average root diameter (cm) was assessed using a precision graduated ruler. Average root volume (cm³) was evaluated with the help of a graduated glass cylinder and water. All previous measurements were performed using five randomly selected plants. At the time of harvest, the total root yield was calculated (ton ha⁻¹) using all plants in all plots of each treatment.

2.6. Statistical Analysis

All experimental data were analyzed applying ANOVA, and Fisher’s LSD test was applied to identify whether the differences between means (±SE) of all treatments were significant. The analysis software (SAS ver. 9.2; SAS Institute Inc., Cary, NC, USA) was applied, and differences were identified at p ≤ 0.05.

3. Results

3.1. Length and Fresh and Dry Matter of Plant Shoot

The tested soil was a native alkaline sandy loam (pH 7.77 and 7.59) with available zinc (Zn) of 0.04 and 0.07 mg kg⁻¹ (Table 2) in the 2019/2020 and 2020/2021 seasons, respectively. Shoot length (ShL), shoot fresh weight (SFrW), and shoot dry matter (SDrM) of carrot plants were appreciably influenced by the maximum dose (40 mg L⁻¹) of zinc oxide nanoparticles (ZnO-NPs(2)), which produced incremental increases and the highest values of the above-mentioned growth parameters; however, the lowest values were produced by the untreated plants (control) (

Table 4. Influence of foliar nourishment with some zinc (Zn) forms on some vegetative growth and physiological parameters of carrot plants grown under Zn deficiency stress in two growth seasons.

Treatments	ShL	SFrW	SDrM	SPAD Readings	MSI
	(cm)	(g plant−1)			(%)
	2019/2020 Season
Control	41.9 b ± 5.2	15.1 e ± 0.7	9.2 e ± 0.5	26.0 e ± 1.5	85.6 d ± 2.2
ZnO-NPs(1)	45.1 a,b ± 1.6	33.4 c ± 2.3	19.3 b ± 0.3	60.2 b ± 2.6	93.7 a ± 0.4
ZnO-NPs(2)	52.5 a ± 2.8	51.2 a ± 0.5	21.0 a ± 0.2	72.8 a ± 0.3	92.6 a,b ± 1.2
Zn–EDTA(1)	50.1 a,b ± 1.1	22.6 d ± 1.2	13.2 d ± 0.6	52.4 b–d ± 1.9	89.1 b–d ± 0.6
Zn–EDTA(2)	46.6 a,b ± 0.5	41.6 b ± 1.4	17.1 c ± 0.5	58.1 b,c ± 3.1	89.1 b–d ± 1.3
ZnO-B(1)	49.7 a,b ± 2.5	22.1 d ± 3.4	12.9 d ± 0.6	47.4 c,d ± 6.2	88.3 c,d ± 1.2
ZnO-B(2)	47.3 a,b ± 1.2	22.7 d ± 1.7	13.2 d ± 0.4	46.1 d ± 4.4	92.6 a–c ± 0.8
	2020/2021 Season
Control	46.3 c ± 2.1	18.7 c ± 0.9	9.9 b ± 0.7	27.0 d ± 1.6	87.0 b ± 2.0
ZnO-NPs(1)	47.5 c ± 0.8	40.8 a ± 2.3	21.0 a ± 5.4	59.9 b ± 3.8	91.8 a ± 1.3
ZnO-NPs(2)	53.5 a ± 0.4	51.6 a ± 0.7	21.3 a ± 3.9	77.5 a ± 3.6	91.2 a ± 0.4
Zn–EDTA(1)	53.1 a ± 1.6	29.3 c ± 1.6	13.9 a,b ± 0.1	58.2 b,c ± 2.6	91.1 a ± 0.1
Zn–EDTA(2)	49.2 b,c ± 0.3	38.6 b ± 3.7	21.0 a ± 1.7	58.3 b,c ± 5.4	87.0 b ± 2.6
ZnO-B(1)	52.7 a,b ± 0.5	19.8 c ± 3.2	11.2 b ± 0.6	51.9 b,c ± 2.3	91.5 a ± 1.7
ZnO-B(2)	51.0 a,b ± 0.3	31.4 b ± 0.5	18.2 a,b ± 0.5	46.1 c ± 3.9	91.2 a ± 2.3

Mean values (±SE) with different letters in each column are significant (p ≤ 0.05). ZnO-NPs(1) = Zinc oxide nanoparticles applied as foliar spraying at 20 mg L⁻¹, ZnO-NPs(2) = Zinc oxide nanoparticles applied as foliar spraying at 40 mg L⁻¹, Zn–EDTA(1) = Zinc–EDTA applied as foliar spraying at 1 g L⁻¹, Zn–EDTA(2) = Zinc–EDTA applied as foliar spraying at 2 g L⁻¹, ZnO-B(1) = bulk zinc oxide applied as foliar spraying at 200 mg L⁻¹, and ZnO-B(2) = bulk zinc oxide applied as foliar spraying at 400 mg L⁻¹. ShL = Shoot length, SFrW = Shoot fresh weight, SDrM = Shoot dry matter, SPAD = Relative chlorophyll content, and MSI = Membrane stability index.

). Values of 52.5 cm and 53.5 cm were recorded as the highest ShL in the two seasons, respectively. The corresponding SFrW and SDrM were 51.2 and 21.0 g plant⁻¹, respectively, in the 2019/2020 season and 52.6 and 21.3 g plant⁻¹, respectively, in the 2020/2021 season. It was noticed that the increasing percentages of the greatest and lowest values were 25.3 vs. 15.5%, 240.3 vs. 175.6%, and 127.5 vs. 115.2% for the above-mentioned growth parameters, respectively.

3.2. Relative Chlorophyll Content and Membrane Stability Index

The data shown in Table 4 display that the relative chlorophyll content (SPAD) was improved by all Zn form treatments compared to the control, and ZnO-NPs(2) conferred the highest significant increase in SPAD in both seasons. ZnO-NPs(2) contributed to the SPAD reading with values of 72.8 vs. 77.5 in both seasons, respectively. The increasing percentages amounted to 180.1 vs. 187.2% compared to the comparison treatment, which gave the lowest values (26.0 vs. 27.0) in the growing seasons 2019/2020 and 2020/2021, respectively. The behavior of the membrane stability index (MSI) was the same as that of SPAD with Zn form treatments (Table 4). However, the percentages of increase in MSI were 9.46 and 5.30% in the two seasons, respectively, which were obtained using ZnO-NPs(1) that produced the maximum values (93.7 vs. 91.8) in both seasons, respectively, compared to the control.

3.3. Leaf Nutrient Contents

As shown in Table 2, the experimental soil is characterized as calcareous containing 10.5 vs. 10.8% CaCO₃ in both seasons, respectively. It is characterized by low nutrient availability. The results related to

Table 5. Influence of foliar nourishment with some zinc (Zn) forms on leaf nutrient contents of carrot plants grown under Zn deficiency stress in two growth seasons.

Treatments	P	Ca	Fe	Mn	Zn	Cu
	(%)		(mg kg−1)
	2019/2020 Season
Control	4.99 e ± 0.01	1.52 a ± 0.01	173.1 g ± 0.49	52.0 b ± 0.49	175.7 e ± 0.21	397.6 d ± 2.75
ZnO-NPs(1)	5.23 c ± 0.00	1.22 c ± 0.00	201.9 e ± 0.40	43.6 c ± 0.17	626.6 c ± 1.27	437.6 b ± 1.62
ZnO-NPs(2)	5.15 d ± 0.01	1.08 e ± 0.00	917.6 a ± 0.81	56.5 a ± 0.72	964.0 a ± 1.62	264.2 e ± 0.39
Zn–EDTA(1)	4.35 g ± 0.02	1.18 d ± 0.01	468.2 b ± 0.78	38.4 e ± 0.38	918.5 b ± 0.04	422.0 c ± 1.85
Zn–EDTA(2)	4.69 f ± 0.03	1.01 f ± 0.00	365.3 c ± 0.98	38.2 e ± 0.29	104.0 g ± 0.16	470.2 a ± 1.50
ZnO-B(1)	5.40 b ± 0.04	1.21 c ± 0.00	183.3 f ± 0.68	41.2 d ± 0.52	279.1 d ± 0.73	267.3 e ± 0.20
ZnO-B(2)	5.48 a ± 0.00	1.33 b ± 0.00	207.2 d ± 0.64	50.3 b ± 0.74	145.9 f ± 0.06	399.8 d ± 1.96
	2020/2021 Season
Control	5.07 e ± 0.01	1.53 a ± 0.00	203.6 c ± 0.58	52.0 b ± 0.45	176.4 d ± 0.21	394.1 c ± 0.72
ZnO-NPs(1)	5.26 c ± 0.00	1.15 b ± 0.03	172.9 g ± 0.40	42.8 d ± 0.31	629.2 b ± 0.23	439.6 a,b ± 2.77
ZnO-NPs(2)	5.17 d ± 0.01	1.07 c ± 0.01	916.1 a ± 0.09	56.9 a ± 0.49	961.4 a ± 0.12	324.9 d ± 35.45
Zn–EDTA(1)	4.39 g ± 0.01	1.18 b ± 0.00	467.2 b ± 0.20	37.2 f ± 0.32	176.8 d ± 0.17	418.8 b,c ± 0.00
Zn–EDTA(2)	4.74 f ± 0.01	1.03 d ± 0.00	366.8 c ± 0.12	37.8 f ± 0.04	104.3 e ± 0.04	473.8 a ± 0.58
ZnO-B(1)	5.46 b ± 0.01	1.15 b ± 0.01	183.1 f ± 0.00	42.0 e ± 0.07	280.5 c ± 0.10	265.9 c ± 0.61
ZnO-B(2)	5.49 a ± 0.00	1.19 b ± 0.01	206.3 d ± 0.12	49.2 c ± 0.08	201.9 d ± 32.23	402.6 b,c ± 0.35

Mean values (± SE) with different letters in each column are significant (p ≤ 0.05). ZnO-NPs(1) = Zinc oxide nanoparticles applied as foliar spraying at 20 mg L⁻¹, ZnO-NPs(2) = Zinc oxide nanoparticles applied as foliar spraying at 40 mg L⁻¹, Zn–EDTA(1) = Zinc–EDTA applied as foliar spraying at 1 g L⁻¹, Zn–EDTA(2) = Zinc–EDTA applied as foliar spraying at 2 g L⁻¹, ZnO-B(1) = bulk zinc oxide applied as foliar spraying at 200 mg L⁻¹, and ZnO-B(2) = bulk zinc oxide applied as foliar spraying at 400 mg L⁻¹. P = Phosphorus, Ca = Calcium, Fe = Iron, Mn = Manganese, Zn = Zinc, and Cu = Copper.

indicate that ZnO-NPs(2) were the best treatment for carrot leaf iron (Fe), manganese (Mn), and zinc (Zn) contents. This treatment produced the highest contents of 917.6 vs. 916.1 mg kg⁻¹ for Fe, 56.5 vs. 56.9 mg kg⁻¹ for Mn, and 964.0 vs. 961.4 mg kg⁻¹ for Zn in the 2019/2020 and 2020/2021 seasons, respectively, while the plant leaves nourished with 2 g L⁻¹ of zinc-chelated EDTA (Zn–EDTA(2)) collected the maximum content of Cu (437.6 vs. 402.6 mg kg⁻¹ in both seasons, respectively). In contrast, the maximum leaf contents of phosphorus (P) and calcium (Ca) were 5.48 vs. 5.47% and 1.52 vs. 1.53% in both seasons, respectively, which were obtained with the higher dose of bulk zinc oxide, 400 mg L⁻¹ (ZnO-B(2)), and the comparison treatment in both seasons, respectively, while the lowest contents of P (4.35 vs. 4.39%) were recorded with 1 g L⁻¹ of zinc-chelated EDTA (Zn–EDTA(1)) and the lowest contents of Ca (1.01 vs. 1.03%) were produced with Zn–EDTA(2) treatment.

3.4. Root Nutrient Contents

All treatments had a significant effect on the contents of root nutrients such as P, Ca, Fe, Mn, Zn, and Cu, as depicted in

Table 6. Influence of foliar nourishment with some zinc (Zn) forms on root nutrient contents of carrot plants grown under Zn deficiency stress in two growth seasons.

Treatments	P	Ca	Fe	Mn	Zn	Cu
	(%)		(mg kg−1)
	2019/2020 Season
Control	0.39 f ± 0.02	0.17 b ± 0.01	34.4 e ± 0.03	9.5 b ± 0.21	17.3 e ± 0.68	11.1 c ± 0.36
ZnO-NPs(1)	0.44 f ± 0.03	0.12 e ± 0.01	30.5 f ± 0.19	52.1 a ± 24.64	17.7 e ± 0.01	15.7 b ± 1.03
ZnO-NPs(2)	0.48 e ± 0.03	0.21 a ± 0.02	49.8 b ± 0.05	11.5 b ± 0.06	30.5 a ± 0.01	12.8 b,c ± 0.51
Zn–EDTA(1)	0.69 b ± 0.04	0.15 d ± 0.01	23.0 g ± 0.02	19.9 b ± 0.29	19.3 d ± 0.01	20.1 a ± 0.17
Zn–EDTA(2)	0.78 a ± 0.05	0.16 c ± 0.02	63.4 a ± 0.42	16.8 b ± 0.03	27.8 c ± 0.06	19.9 a ± 0.01
ZnO-B(1)	0.59 d ± 0.03	0.12 e ± 0.01	43.6 c ± 0.16	10.0 b ± 0.58	20.0 d ± 0.01	12.2 c ± 1.94
ZnO-B(2)	0.63 c ± 0.04	0.15 d ± 0.01	38.7 d ± 0.07	10.6 b ± 0.06	29.3 b ± 0.00	13.2 b,c ± 1.11
	2020/2021 Season
Control	0.52 e ± 0.03	0.13 e ± 0.01	139.3 e ± 0.02	12.3 c ± 0.40	16.7 b ± 0.02	15.6 a,b ± 2.11
ZnO-NPs(1)	0.70 c ± 0.04	0.18 c ± 0.02	135.2 f ± 0.02	57.1 b ± 21.99	20.7 b ± 1.62	15.8 a,b ± 0.14
ZnO-NPs(2)	0.53 d ± 0.03	0.62 a ± 0.05	154.9 b ± 0.01	93.0 a ± 12.56	38.4 a ± 1.12	13.2 b ± 0.69
Zn–EDTA(1)	0.53 d ± 0.02	0.15 d ± 0.01	127.9 g ± 0.04	16.7 c ± 2.69	31.4 a,b ± 0.52	17.8 a,b ± 2.26
Zn–EDTA(2)	0.50 f ± 0.02	0.15 d ± 0.01	168.6 a ± 0.50	15.1 c ± 1.98	22.9 b ± 2.75	16.0 a,b ± 0.19
ZnO-B(1)	0.77 a ± 0.05	0.30 b ± 0.03	148.8 c ± 0.00	20.1 c ± 4.79	21.3 b ± 10.60	30.2 a ± 11.19
ZnO-B(2)	0.74 b ± 0.04	0.14 d ± 0.01	144.3 d ± 0.29	13.5 c ± 0.27	26.2 a,b ± 1.78	17.8 a,b ± 1.29

Mean values (± SE) with different letters in each column are significant (p ≤ 0.05). ZnO-NPs(1) = Zinc oxide nanoparticles applied as foliar spraying at 20 mg L⁻¹, ZnO-NPs(2) = Zinc oxide nanoparticles applied as foliar spraying at 40 mg L⁻¹, Zn–EDTA(1) = Zinc–EDTA applied as foliar spraying at 1 g L⁻¹, Zn–EDTA(2) = Zinc–EDTA applied as foliar spraying at 2 g L⁻¹, ZnO-B(1) = bulk zinc oxide applied as foliar spraying at 200 mg L⁻¹, and ZnO-B(2) = bulk zinc oxide applied as foliar spraying at 400 mg L⁻¹. P = Phosphorus, Ca = Calcium, Fe = Iron, Mn = Manganese, Zn = Zinc, and Cu = Copper.

. The overall trend of results indicates that the 2019/2020 season outperformed the 2020/2021 season in terms of root nutrient contents. The application of ZnO-NPs(2) produced significant maximum values (0.21 vs. 0.62%) for Ca in the first and second seasons, respectively, whereas Zn–EDTA(2) and ZnO-B(1) conferred maximum contents (0.78 vs. 0.77%) for P in both seasons, respectively. The greatest Mn contents (63.4 vs. 168.6 mg kg⁻¹) were recorded with Zn–EDTA(1), but the lowest Mn contents (9.5 vs. 12.3) were produced with the comparison treatment in both seasons, respectively. For Fe, the highest contents (63.4 vs. 168.6 mg kg⁻¹) were produced with Zn–EDTA(2), while the lowest contents (23.0 vs. 127.9 mg kg⁻¹) were obtained with Zn–EDTA(1) in both seasons, respectively. Regarding Cu, the maximum contents (20.1 vs. 17.8 mg kg⁻¹) were produced with Zn–EDTA(1), while the minimum contents (11.1 vs. 15.6 mg kg⁻¹) were obtained with the comparison treatment in both seasons, respectively. With respect to Zn nutrients, the highest contents (30.5 vs. 38.4 mg kg⁻¹) were produced with ZnO-NPs(2), while the minimum contents (17.3 vs. 16.7 mg kg⁻¹) were obtained with the comparison treatment in both seasons, respectively.

3.5. Anti-Radical Power (ARP) and β-Carotene

The results of the anti-radical power (ARP) and β-carotene in the root of carrot plants are shown in Figure 2 and Figure 3. The application of ZnO-NPs(1) did not affect ARP, but the other treatments significantly reduced ARP, and the largest values (233.0 vs. 224.8) were recorded with the control treatment. ZnO-B(1) treatment recorded the lowest values (142.3 vs. 140.5) with decreasing percentages of 38.9 vs. 37.5% compared to the control in both seasons, respectively. The maximum content of β-carotene (3.37 vs. 3.32) was obtained with ZnO-NPs(1) (20 mg L⁻¹) and ZnO-B(1) (200 mg L⁻¹) in the 2019/2020 and 2020/2021 seasons, respectively, while the lowest values (2.54 vs. 2.28) were recorded with ZnO-B(2) (400 mg L⁻¹) and ZnO-NPs(2) (40 mg L⁻¹) in both seasons, respectively. The increasing percentages reached 32.7 vs. 45.6% in both seasons, respectively, compared to the lowest ones.

3.6. Root Yield and Its Components

The results regarding the effect of ZnO-NPs, Zn–EDTA, and ZnO-B on the root quality of carrot plants are depicted in

Table 7. Influence of foliar nourishment with some zinc (Zn) forms on yield and its components of carrot plants grown under Zn deficiency stress in two growth seasons.

Treatments	Root Fresh Weight	Root Dry Matter	Root Length	Root Diameter	Root Volume	Total Yield
	(g plant−1)		(cm)	(cm2)	(cm3)	(t ha−1)
	2019/2020 Season
Control	193.4 d ± 12.55	107.8 c ± 15.86	21.0 a ± 1.53	4.22 a ± 0.46	176.7 d ± 16.67	28.1 e ± 5.50
ZnO-NPs(1)	347.4 a ± 20.84	139.3 a,b ± 6.22	21.1 a ± 0.59	5.33 a ± 0.51	223.3 b,c ± 12.02	49.2 a,b ± 0.82
ZnO-NPs(2)	332.4 a,b ± 27.55	159.4 a ± 11.80	22.8 a ± 0.78	5.44 a ± 0.20	313.3 a ± 13.33	52.1 a ± 0.65
Zn–EDTA(1)	314.9 a–c ± 5.14	145.2 a,b ± 2.54	20.9 a ± 0.29	4.81 a ± 0.51	246.7 b ± 6.67	43.2 b,c ± 1.43
Zn–EDTA(2)	269.1 c ± 4.52	134.5 a–c ± 5.53	21.0 a ± 0.33	5.33 a ± 0.48	220.0 b–d ± 11.55	33.1 d,e ± 1.77
ZnO-B(1)	290.7 b,c ± 7.06	125.7 b,c ± 4.41	21.8 a ± 0.55	5.01 a ± 0.53	240.0 b,c ± 30.55	42.2 b,c ± 1.47
ZnO-B(2)	311.9 a–c ± 23.88	118.4 b,c ± 2.50	21.6 a ± 0.40	4.50 a ± 0.25	200.0 c,d ± 11.55	38.0 c,d ± 2.71
	2020/2021 Season
Control	214.7 d ± 16.59	102.4 b ± 16.93	25.7 a ± 0.88	4.52 b ± 0.17	193.3 b ± 6.67	31.8 d ± 2.61
ZnO-NPs(1)	324.7 a,b ± 5.26	127.6 a,b ± 11.01	22.8 b ± 0.77	6.33 b ± 0.19	226.7 b ± 13.33	56.8 a ± 4.21
ZnO-NPs(2)	313.7 a,b ± 6.25	156.1 a ± 5.42	25.6 a,b ± 1.16	6.44 a ± 0.39	330.0 a ± 30.66	59.0 a ± 3.57
Zn–EDTA(1)	332.9 a ± 12.80	121.5 a,b ± 7.17	24.1 a,b ± 0.91	5.14 b ± 0.44	193.3 b ± 24.04	51.4 a,b ± 1.11
Zn–EDTA(2)	288.7 c ± 4.30	140.9 a,b ± 14.45	24.0 a,b ± 0.88	5.41 a,b ± 0.42	233.3 b ± 35.28	38.2 c,d ± 1.53
ZnO-B(1)	281.0 c ± 10.92	127.4 a,b ± 13.38	24.8 a,b ± 0.55	5.35 a,b ± 0.34	186.7 b ± 6.67	45.5 b,c ± 1.47
ZnO-B(2)	301.2 b,c ± 1.57	120.4 a,b ± 3.75	24.9 a,b ± 0.29	5.17 b ± 0.10	186.7 b ± 6.67	41.8 c ± 0.81

Mean values (± SE) with different letters in each column are significant (p ≤ 0.05). ZnO-NPs(1) = Zinc oxide nanoparticles applied as foliar spraying at 20 mg L⁻¹, ZnO-NPs(2) = Zinc oxide nanoparticles applied as foliar spraying at 40 mg L⁻¹, Zn–EDTA(1) = Zinc–EDTA applied as foliar spraying at 1 g L⁻¹, Zn–EDTA(2) = Zinc–EDTA applied as foliar spraying at 2 g L⁻¹, ZnO-B(1) = bulk zinc oxide applied as foliar spraying at 200 mg L⁻¹, and ZnO-B(2) = bulk zinc oxide applied as foliar spraying at 400 mg L⁻¹.

.

ZnO-NPs(2) treatment recorded the highest root length (22.8 cm) in the 2019/2020 season and the largest root volume (330.0 cm³) in the 2020/2021 season, along with the highest root diameter (5.44 vs. 6.44 cm) in both seasons, respectively. Moreover, ZnO-B(1) and ZnO-B(2) produced the lowest root volume (186.7 cm³) in the 2020/2021 season, while the lowest root volume (176.7 cm³) in the 2019/2020 season was produced with the comparison treatment. For root length, the maximum values (22.8 vs. 25.6 cm) were obtained with ZnO-NPs(2) treatment in both seasons, respectively. The increasing percentages were 8.5 vs. 12.7% for root length, 35.8 vs. 76.8% for root volume, and 28.9 vs. 42.5% for root diameter in both seasons, respectively. In general, the incremental rates in the yield of carrot roots and yield components in the second season were better than those in the first season. The results of variables associated with total root production are also shown in Table 7. For all the treatments tested, significant differences were obtained. ZnO-NPs(2) treatment produced the highest root yield, surpassing ZnO-NPs(1) by 5.9 vs. 3.8%, ZnO-B(2) by 37.0 vs. 41.3%, ZnO-B(1) by 23.7 vs. 29.8%, and the control by 85.3 vs. 85.4% in the 2019/2020 and 2020/2021 seasons, respectively. The largest root yields (52.1 vs. 59.0 t ha⁻¹) were obtained with ZnO-NPs(2), while the untreated plants produced the lowest values (28.1 vs. 31.8 t ha⁻¹) in both seasons, respectively.

3.7. Correlation and Regression Analysis

Pearson’s correlation coefficients in

Table 8. Pearson’s correlation coefficients between total root yield (TRY) with some selected attributes of Zn-stressed carrot plants nourished by foliar spraying with different doses of zinc oxide nanoparticles in two seasons.

Character	TRY (t ha−1)
	2019/2020		2020/2021
	Person r	Probability	Person r	Probability
Vegetative Growth Characters
Shoot length (SL)	0.577 **	0.003	0.437 *	0.033
Leaf fresh weight (LFW)	0.410 *	0.046	0.658 **	0.000
Leaf dry matter (LDM)	0.503 *	0.012	0.710 **	0.000
Physiological Characters
MSI%	0.728 **	0.000	0.478 *	0.018
SPAD reading	0.344	0.100	0.219	0.305
Yield Component Characters
Root length (RL)	0.204	0.339	−0.088	0.683
Root volume (RV)	0.563 **	0.004	0.351	0.093
Root diameter (RD)	0.548 **	0.006	0.668 **	0.000
Root fresh weight (RFW)	0.747 **	0.000	0.742 **	0.000
Root dry matter (RDM)	0.737 **	0.000	0.461 *	0.023
Leaf nutrients content
Phosphorus (P)	0.128	0.550	−0.037	0.865
Calcium (Ca)	−0.404	0.051	−0.381	0.066
Iron (Fe)	0.539 **	0.007	0.561 **	0.004
Manganese (Mn)	0.199	0.350	0.263	0.215
Zinc (Zn)	−0.388	0.061	−0.179	0.402
Copper (Cu)	0.668 **	0.000	0.637 **	0.001

* and ** indicate, respectively, differences at p ≤ 0.05 and p ≤ 0.01 probability level.

between total root yield (TRY) and each of all the tested growth and physiological parameters (ShL, SFrW, SDrM, MSI, and SPAD readings), tested yield component parameters (RL, RV, RD, RFrW, and RDrM), and tested leaf and root nutrient contents (P, Ca, Fe, Mn, Zn, and Cu) were computed to highlight the efficacious TRY. Highly significant (p ≤ 0.01) positive correlations (r = 0.747 **, 0.737 **, 0.728 **, 0.668 **, 0.577 **, 0.563 **, 0.548 **, and 0.539 **) were obtained between TRY and each of RFrW, RDrM, MSI, Cu, SL, RV, RD, and Fe, respectively, while significant (p ≤ 0.05) positive correlations (r = 0.503 * and 0.410 *) were recorded between TRY and both SDrM and SFrW, respectively, in the 2019/2020 growth season. Highly positive correlations (r = 0.742 **, 0.710 **, 0.668 **, 0.658 **, 0.637 **, and 0.561 **) were obtained between TRY and each of RFW, LDM, RD, LFW, Cu, and Fe, respectively, and significant (p ≤ 0.05) positive correlations (r = 0.478 *, 0.461 *, and 0.437 *) were recorded between TRY and each of MSI, RDM, and SL, respectively, in the 2020/2021 season.

Outputs of stepwise regression analysis in

Table 9. Proportional contribution in predicting total root yield (TRY) using stepwise multiple linear regression for Zn deficiency-stressed carrot plants nourished by foliar spraying with different doses of zinc oxide nanoparticles in two seasons.

Year	r	R2	Adjusted R2	SEE	Significance	Fitted Equation
2019/2020	0.959	0.920	0.898	2.705	***	TRY = −98.073 + 1.056Fe + 0.848LFW + 2.513RD + 0.294Mn + 0.011Cu
2020/2021	0.957	0.915	0.897	3.367	***	TRY = −26.395 + 0.862LDM + 0.130RFW + 0.387Mn + 0.008Cu

*** indicates differences at p ≤ 0.001 probability level.

clarify that there existed five studied parameters (i.e., Fe, LFW, RD, Mn, and Cu) and four studied parameters (DM, RFW, Mn, and Cu) in the 2019/2020 and 2020/2021 seasons, respectively. Both seasons had a highly significant contribution to TRY.

4. Discussion

From setting seeds in seedbeds to harvesting, plants suffer from one or more of several abiotic environmental adversaries, such as salinity [21,22,23,24,25,26,27,28,29,30,31,32], drought [33,34,35,36,37], heavy metals [38,39,40,41,42], weed invasion [43,44,45], nutrient deficiency [46,47], high soil CaCO₃ content [4,17,48], or even more than one stress at the same time [18,49,50,51,52,53,54]. Therefore, lots of attempts have been made by many researchers to explore effective solutions that lead to a successful adaptation of different plant species, especially those sensitive to those abiotic adversaries, which are increasing with climate change conditions that we did not realize before. This led to the emergence of a major problem for the agricultural sector, which began to threaten food security and agricultural sustainability as a result of declining crop productivities. One of these agricultural issues that greatly influences crop growth and productivity is nutrient deficiencies in agricultural soils, especially micronutrients, including zinc (Zn) deficiency [46,47,55].

A wide range of different soil types in many parts of the world suffer from Zn deficiency, especially in sandy-textured soils with high pH and calcium carbonate (CaCO₃), as shown in Table 2. The uptake of Zn by plants is related to the soil carbonate content. Calcium and soil pH are two of the main factors that limit the availability of this nutrient. Thus, soil Zn deficiency reduces the Zn content of edible foodstuffs [56], and thus Zn deficiency affects more than a third of the world’s population [57]. Therefore, this study was planned to address the problem of Zn deficiency in Zn-deficient soils by treating carrot plants using a foliar spray strategy with different forms of Zn such as ZnO nanoparticles (ZnO-NPs), Zn–EDTA, and ZnO-B in an attempt to identify which one of these Zn forms could be a solution for the successful adaptation of plants to coexist with the problem of Zn deficiency in defective soils. The application of ZnO-NPs, Zn–EDTA, and ZnO-B as a foliar nourishing strategy for carrot plants has rarely been studied at the field level under Zn-deficient soil conditions.

For carrot plants grown in Zn-deficient soil, ZnO-B was applied at both levels of 200 and 400 mg L⁻¹, Zn–EDTA was applied at both levels of 1 and 2 g L⁻¹, and ZnO-NPs was applied at both levels of 20 and 40 mg L⁻¹ compared to the control, in which Zn was not applied. Among all these treatments, ZnO-NPs applied at 40 mg L⁻¹ (ZnO-NPs(2)) were the best treatment that collected more acceptable growth and physiological parameters and nutrient homeostasis, which was reflected in the highest yields. The best nutrient homeostasis in plant tissues obtained due to the balanced nutrient uptake with applying ZnO-NPs(2) leads to acceptable plants benefiting from the crucial roles of nutrients in noticeably improving photosynthesis and plant growth in favor of plant productivity [58,59]. Therefore, foliar use of ZnO-NPs(2) due to limited nutrient availability can be resolved by reducing the loss of Zn added to the soil [60].

In our study, the resulting data indicate that the application of ZnO-NPs(2) improved all studied growth variables since Zn plays a key role in maintaining the plants’ physiological health (Table 3). This result was obtained because ZnO-NPs(2) improved nutrient usability by plants to enhance photosynthetic pigments, the rate of photosynthesis that increases total carbohydrate accumulation, leaf dry matter production, and finally the outcome of plant growth parameters [59]. The Marschner [58] study reported an enhanced effect of ZnO-NPs on chlorophyll content, which was explained on the basis of micronutrients playing critical roles in the synthesis of chloroplast proteins and thus perhaps interfering with chlorophyll synthesis. It has been revealed that micronutrient deficiency inhibits chlorophyll formation through the inhibition of protein synthesis [58]. These findings can be explained in more detail by Mahmoud et al. [61], who indicated that Zn assimilation is more efficient when applied at a nano-metric size. Duhan et al. [62] showed increases in the growth of seedlings treated with ZnO-NPs as a foliar application, which is more effective than ZnO-B, as its release can be slow and gradual [63], but it has not been identified whether this effect is due to ZnO-NPs uptake or due to the dissolution of its products [64]. With results regarding relative chlorophyll content (SPAD values), Pullagurala et al. [65] attributed the increase in SPAD values to the fact that Zn plays an essential and vital role in plant metabolism, affecting the activation of important enzymes such as carbonic anhydrase containing a Zn atom that catalyzes the hydration of H₂O and CO₂, facilitating the diffusion of carbon dioxide to the carboxylation sites in plants [66]. These results are in good agreement with the findings of Atteya et al. [67] and Gheith et al. [68] regarding jojoba and maize plants, attributing the positive findings obtained to the fact that the foliar application of ZnO-NPs leads to a large acquisition of Zn by plants.

This study demonstrated that the role played by ZnO-NPs is not only to improve plant growth and physiological (membrane stability index and SPAD values) parameters, but also to enhance nutrient uptake and plant nutrient content (Table 5 and Table 6). In this connection, the use of Zn in the form of ZnO-NPs is to overcome the adverse effects of Zn-deficient conditions and improve plant nutritional quality. However, the performance of nutrient nanoparticles depends on several properties such as particle size, chemical structure, service covering, and application dosages [69]. In this study, among all Zn treatments, ZnO-NPs applied at 40 mg L⁻¹ (ZnO-NPs(2)) made carrot plants perform well under Zn-deficient soil conditions (Table 3, Table 4, Table 5, Table 6, Table 7 and Table 8). This desirable output can be attributed to the fact that Zn plays an important role in increasing protein synthesis, membrane function, cell elongation, and encouraging plant roots to positively exchange cations, which helps plants absorb more nutrients. In addition, Zn modifies auxin influences by regulating the synthesis of the amino acid tryptophan, which is a precursor to the synthesis of indole-3-acetic acid (IAA) auxin, which is very important for cell elongation and plant growth. It also acts as a cofactor for many enzymes such as superoxide and dehydrogenases [70,71].

This study revealed that the Cu content does not match that of Fe, Zn, and Mn, which can be attributed to the antagonistic effect between Cu and other nutrients, including Zn. This antagonism (between Zn and Cu) may lead to maintaining the Cu content at the appropriate level to prevent Cu toxicity, as Cu activates many enzymes related to photosynthesis and respiratory systems as an essential metal cofactor [72]. The general trend of Zn, Fe, and Mn contents was stable due to the synergistic effect, and the trend of the results obtained is in good agreement with those obtained by Mahmoud et al. [61]. As expected, outperforming the other forms of Zn, the application of ZnO-NPs to carrot plants noticeably increased the Zn content. Zn can be absorbed primarily in the leaf wax layer [11]. Size is probably a limiting factor in the diffusion of Zn in Zn–EDTA and ZnO-B across stomata (19.1 to 71.5 µm) [73], which only allows particles with smaller sizes to pass through [11]. In contrast, our data showed a decrease in calcium (Ca) uptake due to the application of ZnO-NPs, which can be attributed to the fact that Zn competes with Ca for plant uptake. The increase in the uptake of other nutrients may be attributed to the fact that ZnO-NPs enhance the cation exchange capacity (CEC) of the roots, which in turn enhances the absorption of nutrients, which is positively reflected in the improved plant growth and satisfactory productivity. Supporting plant growth and productivity, Zn also plays a key role in controlling the production of IAA hormone, as well as in carbohydrate and protein metabolism and synthesis [70,71].

As depicted in Table 4, all treatments containing Zn nutrients in either nano or bulk forms caused an improvement in the cell membrane stability index (MSI) compared to the control, in which Zn was not applied. The enhancement in the cell MSI could be due to the inclusion of Zn in the defensive enzyme superoxide dismutase and peroxidases, which helps mitigate and minimize the harmful influences of oxidative stress and the accumulation of malondialdehyde and toxic ions such as Na⁺, which cause electrolyte leakage, and also helps to accelerate the synthesis of chlorophyll and the metabolic processes in the plant. In addition, it has been reported that ZnO-NPs notably help in the increase in soil organic carbon due to the release of root exudates from the roots of ZnO-NPs-treated plants, possibly increasing net photosynthetic rates because root exudates are considered to be part of the excess carbon from photosynthesis [74]. Therefore, Zn protects cell membranes from damage caused by Zn deficiency due to its rapid penetration into the plant cells when it is applied in the form of ZnO-NPs to fulfill its vital functions in the plant [61,75].

The positive effect of ZnO-NPs on enhancing the uptake of nutrients and their positive reflection on growth parameters and yield and its components (Table 7) can be due to the role of Zn in controlling plant hormonal (IAA) content and its relationship to cell elongation and plant growth, and also due to its role in protein synthesis and carbohydrate metabolism in addition to its excellent role in gene expression related to abiotic stress. These results are confirmed by Sadeghzadeh and Rengel [76] and Quary et al. [77], who reported that ZnO-NPs manipulate all plant growth parameters, resulting in useful changes in productivity traits. The possible reason for each positive role of Zn is the enhancement in the activity of bio-substances and/or the activity of the photosynthetic system, or might be due to the activating role of Zn in the metabolic processes of plants and the photosynthetic rate, which are reflected in the improvement in the yield components. Compared with other forms of Zn (Zn–EDTA and ZnO-B), with the use of nano-fertilizers (1–100 nm), nutrient uptake is greatly improved due to their very small size, which in turn can improve the absorption and interaction of other micronutrients in plant tissues [78]. In addition, the greatest positive influence of Zn in the form of ZnO-NPs may be due to its most easy and rapid assimilation into plant tissues, which improves photosynthesis and leads to the efficient growth of carrot plants. It has been previously explained that sprayed ZnO-NPs are mostly retained in the leaf waxy layer [79], leading to an increase in Zn content in leafy tissues. This may be due to the fact that Zn binds to the hydrophilic, histidine-rich amino acid stretch in the middle of the ZAT1p protein, an uptake system found in cytoplasmic membranes [80]. Thus, ZAT1 expression leads to the accumulation of Zn²⁺ in plant tissues. In addition, Zn transport does not require an energy source such as ATP or the proton motive force; thus, the Zn concentration gradient is sufficient to drive Zn transport by ZAT1p. It could be argued that the 25-amino-acid N-terminus of ZAT1p may be essential for energy coupling. Zn transport into cells by ZAT1p fits to a DY-driven uniport of Zn²⁺. Zn-transporting P-type ATPases seem to transport Zn²⁺ cations bound to thiol groups, e.g., to glutathione. Even in the presence of high-activity efflux systems, ZAT1p functions as an uptake system for Zn²⁺, while CzcD and ZRC1p act as Zn-efflux systems. The COT1p protein is also an uptake system, but only at an outside concentration of 1 μM Zn²⁺ [80].

Zn is taken up mainly as a divalent cation (Zn²⁺ ion) by plant roots. However, in some cases, organic ligand–Zn complexes are also absorbed by plant roots. Depending upon the ligand secreted by plant roots, two physiological strategies are involved in the uptake of divalent cations such as Zn²⁺ [81]. For a greater understanding of nutrient absorption, roots are not just static organs. Plant roots release various organic acids, amino acids, sugars, protons, and even some mineral ions, etc., in the rhizosphere that facilitate their adequate functioning and growth. Zn is absorbed as divalent metal ion Zn²⁺ through mass flow and diffusion mechanisms by roots. In the case of the foliar application of Zn, as in the current study, passive Zn uptake by these mechanisms may be involved in the participation of water (solvent) molecules and the difference in Zn concentrations across the leaf cell plasma membrane. The main driving force in Zn²⁺ uptake (cation uptake) may be the hyperpolarization of the leaf cell plasma membrane, which is mediated through the activity of the leaf cell plasma membrane H⁺-ATPase system. The leaf cell plasma membrane H⁺-ATPase system actively pumps H⁺ ion extracellularly at the expense of ATP hydrolysis. The release of H⁺ ions in the rhizosphere causes the hyper-polarization of the leaf cell plasma membrane on one hand, while it reduces the medium pH on the other hand, which results in an increased cation uptake rate. However, unlike water, charged Zn ions are not able to cross cell membranes freely, so these divalent cations are transported by specific transporter proteins. These proteins are not in close association with ATP breakdown, which confirms the passive uptake of Zn rather than active. Furthermore, Zn²⁺ uptake also occurs by non-selective cation channels associated with the passive flux of diverse groups of cations. This additional driving force in the uptake of many metal cations is likely due to their very low cytoplasmic activity, which is a result of metal sequestration and their binding to intracellular sites (i.e., Zn finger proteins, organic acids, enzymes, etc.) [81].

Nanoparticles have increasing applications and the transportation and separation of nanoparticles have attracted many interesting investigations. Nanoscale manipulation refers to the controllable transportation, separation, and classification of nano-objects, which can be further used to design more complex nano-devices and nano-instruments with nano-elements, and should be very useful for development in nanosciences and nano-industries. Some of these technologies are still in the development stage and can only be performed at the laboratory level, which still has a long way to go for industrial applications. The precise manipulation of small objects or particles remains a challenge, not to mention the problem of classifying nanoparticles of different sizes [82].

Nanoparticles can start to move and the subsequent movement of the nanoparticles depends mainly on the competition between the driving force and the damping force, considering the effect of nanoparticle sizes and thicknesses, as well as the velocity of sliding blocks on transport behavior. The transportation of nanoparticles would be easier if the sliding block velocity was lower. Critical velocity is put forward as a key parameter to describe the transport behavior, under which nanoparticles can be made. In general, the critical velocity decreases with increasing nanoparticle size and thickness, and as a result, transport is easier for smaller and thinner nanoparticles. Moreover, transport would also be easier with a lower viscous damping force and a higher temperature. The initial compatibility of the interface has no obvious effect on the transport behavior, especially in the case of a pre-stretched substrate. Nanotechnology can be further applied with the design of suitable transporters from which nanoparticles of different sizes can be selected [82].

5. Conclusions

Our investigation displayed that foliar nourishment with different forms of zinc (ZnO-NPs, Zn–EDTA, and ZnO-B) positively affected carrot plant growth, physiology (including zinc content), and productivity to varying degrees based on the form of zinc applied, and more acceptable results were obtained with ZnO-NPs applied at 40 mg L⁻¹. A future investigation should be carried out to explore more about how and why ZnO-NPs are more successful than other forms of zinc (e.g., Zn–EDTA and ZnO-B) in influencing different aspects of plant physiology and biochemistry, which are reflected in plant growth and productivity. In addition, it should be explored how and why zinc in ZnO-NPs outperformed Zn in other forms in performing its functions in plants. It is clear, undoubtedly, that the growing population poses many challenges to all countries; therefore, the strategy of nano-size foliar fertilizers, including ZnO-NPs, at low concentrations can be applied to be one of the most important tools for achieving food security effectively and economically.