The Impacts of Ethylicin on Absorption, Transport, and Growth in Tomato Plants

Abstract

This study investigates the uptake, translocation, and pathogen control efficacy of ethylicin in tomato plants using a combination of indoor root irrigation, spraying, and field root irrigation experiments. The results indicate that ethylicin shows dual-directional translocation in tomato plants. On the third day after foliar spraying, ethylicin was detected in the roots at a concentration of 2.93 mg/kg, indicating downward movement. On the third day after root irrigation, ethylicin was detected in the leaves at a concentration of 3.44 mg/kg, confirming upward movement. In the field experiments, ethylicin was absorbed and transported to the upper leaves within six hours of root irrigation at a concentration of 3.85 mg/kg for a single-agent ethylicin and 5.87 mg/kg for an ethylicin–oligosaccharin compound. These results indicate that oligosaccharins enhance the absorption of ethylicin. Ethylicin residue dissipated by the fifth day. No ethylicin was detected in the untreated controls. Root irrigation during the growing period showed an effective reduction of Fusarium spp. and Phytophthora spp. populations in the soil and control of soil-borne diseases. These findings provide theoretical support for the efficient application of ethylicin in the field.

1. Introduction

Soil fumigation is widely used as an effective means of controlling soil-borne diseases, improving the physical properties of soil, and promoting crop growth [1,2,3]. Common fumigants include chloropicrin, dazomet, and metham sodium. However, methyl bromide, an effective fumigant, has been banned due to its harmful effects on the ozone layer [4], and chloropicrin is classified as a restricted-use pesticide due to its high toxicity [5]. Currently, there is an extreme shortage of registered soil fumigant and traditional fumigants are mainly applied before planting. This highlights the urgent need for environmentally friendly fumigants that can be used during the growing season to prevent and control soil-borne diseases.

A crucial aspect of developing new fumigants is understanding their uptake and translocation mechanisms in plants. When a pesticide is applied to a plant, it could pass through the plant’s surface into its interior in a process called endosorption [6]. This process involves not only passive diffusion of pesticides, but also active uptake and phloem transport, offering significant advantages for the control of root and vascular diseases, as well as pests with piercing-sucking mouthparts [7]. The efficiency of pesticide uptake and translocation in plants depends on the chemical properties of the compound [8] as well as a variety of other factors, such as plant species, growth stage, environmental conditions, and application techniques [9]. Studying the endosorption and accumulation of pesticides in crops not only helps to clarify the mechanism of pesticide action and provide a scientific basis for pesticide selection and application, but also helps to understand the residual behavior of pesticides and ensure the safety of agricultural products [10,11,12,13].

Ethylicin, a potential fumigant with the molecular formula C₄H₁₀O₂S₂, a boiling point of 283.71 °C, and a flash point of 125.38 °C, has a density of 1.191 g/cm³ and appears as a colorless or slightly yellow oily liquid. The chemical structural formula of ethylicin is shown in Figure S1. Owing to its high efficiency, broad-spectrum activity, low residue, and rapid degradation, ethylicin demonstrates a strong potential for the control of soil-borne diseases [14,15,16]. Compared to traditional fumigants, ethylicin has less of an impact on the environment, does not contribute to the depletion of the ozone layer, and effectively inhibits a wide range of pathogenic bacteria and nematodes [3]. When used at the recommended doses, it does not accumulate in the soil [17,18], making it a good agent for green agricultural production [19].

This study systematically investigates the mechanisms of ethylicin uptake in tomato plants, focusing on the variation in uptake rate over time. Additionally, the study examines the transport pathways of ethylicin within the plant, determining whether it primarily moves through the xylem, phloem, or other tissues. Its distribution in various plant parts, including stems, leaves, and fruits, is also evaluated. Furthermore, the study comprehensively assesses the effectiveness of ethylicin in controlling soil-borne diseases throughout the tomato plant’s growth cycle. By addressing these objectives, this research seeks to provide a robust theoretical foundation of and practical guidance for the safe and effective use of ethylicin as a fumigant during crop growth. Moreover, the findings of this study are expected to offer valuable insight in the optimization of agricultural disease control strategies and in the development of novel agrochemicals.

2. Materials and Methods

2.1. Experimental Materials and Reagents

2.1.1. Test Formulations

Dichloromethane was procured from Shanghai Macklin Biochemical Technology Co., Ltd. (Shanghai, China) while NaCl, MgSO₄, potassium phthalate, graphitized carbon black (GCB) and N,N-Dipropylethylamine (PSA) were sourced from Sinopharm Chemical Reagent Co., Ltd. (Shanghai, China) The tested formulations are listed in

Table 1. Description of formulations.

Name	Manufacturer	Abridge
80% Ethylicin	Hainan Zhengye Zhongnong High-Tech Co., Ltd. (Haikou, Hainan)	Single-agent formulation
80% Ethylicin + 5% oligosaccharins	Hainan Zhengye Zhongnong High-Tech Co., Ltd. (Haikou, Hainan)	Compound formulation

.

2.1.2. Soil Treatments

Soil was collected from two blank control areas of the following tomato plan greenhouses: Nanhe Village, Dashiwo Town, Fangshan District, Beijing, and Nanshan Xinnong, Miyun. The surface layer of soil (5–20 cm) was retrieved, adjusted to optimal moisture levels using sterilized water, and incubated indoors for one week. The physiochemical properties of the soils are shown in

Table 2. Physicochemical properties of tested soils.

Physicochemical Properties	NH4+-N (mg/kg)	NO3−-N (mg/kg)	Available K (mg/kg)	Organic Matter (g/kg)	pH (1:2.5)	Electrical Conductivity (ms/cm)
Fangshan	3.98	160	845.00	32.50	7.75	239.00
Miyun	4.08	42.82	1600.00	20.02	6.26	1692.25

. The soil from the greenhouses in the Fangshan District was characterized as “sandy loam” (sand: 35%, silt: 52%, clay: 13%). The soil from the greenhouses in the Miyun District was characterized as “sandy clay” (sand: 58%, silt: 6%, clay: 36%). The soils’ water-holding capacity was kept at 15% during indoor incubation and at 60% under field conditions, and greenhouse temperatures were maintained at 27–28 °C during the day and 15–28 °C at night.

2.2. Methods

2.2.1. Tomato Cultivation

Pre-cultivated tomato seedlings in 96-well black soil culture boxes were selected and transplanted into untreated soil. After 30 days of growth, healthy tomato plants of uniform size were selected for subsequent ethylicin root irrigation and foliar spraying.

2.2.2. Uptake and Translocation of Ethylicin in Indoor Tomato Plants

The sieved and prepared soil from Section 2.1.2 was used for this experiment. The soil was passed through a 2 cm sieve and uniformly mixed to remove any debris. A total of 240 g of the moist soil was placed into 2 L plastic nutrient pots (12 cm in diameter). Pre-cultivated tomato seedlings were then treated with a 1000× dilution of ethylicin via root irrigation and foliar spraying, with each treatment replicated four times. Samples of the soil and tomato plants were collected 3, 5, 7, 10, and 14 days after application. The tomato plants were carefully extracted, rinsed with deionized water to remove the soil, and dried with filter paper. The roots and shoots were separated, cut into pieces, sealed in bags, and stored at −80 °C until analysis.

2.2.3. Sample Pre-Treatment

Prior to testing, stem, leaf, and root samples from the tomato plants were removed from the freezer, thawed, and ground in liquid nitrogen for 10 min until fully pulverized. For each sample, 2 g (±0.01 g) of the ground material was precisely weighed into a 50 mL plastic centrifuge tube. The samples were vortexed for 3 min to ensure homogeneity. Subsequently, 1 g of NaCl, 10 mL of methylene chloride [20], and 0.5 mL of buffer (pH = 5) were added to each centrifuge tube [21]. The tubes were vortexed again at 2500 rpm for 10 min and left to stand for 10 min to allow for phase separation.

Next, 2 mL of the supernatant was carefully aspirated using a syringe and transferred into 2.5 mL centrifuge tubes containing a purifying mixture. For the root samples, the mixture consisted of 50 mg of PSA and 150 mg of anhydrous MgSO₄. For the stem and leaf samples, the mixture consisted of 20 mg of GCB, 50 mg of PSA, and 150 mg of anhydrous MgSO₄. The samples were vortexed for 1 min, centrifuged at 5000 rpm for 5 min, and the supernatant was filtered through a 0.22 μm microfiltration membrane. The final sample was collected in a 2 mL injection vial for subsequent GC–MS analysis.

2.2.4. Sample Addition Recovery

Samples were spiked with ethylicin at concentrations of 1 mg/kg, 10 mg/kg, and 100 mg/kg. A blank control (without the addition of ethylicin) was included in all treatments, with five replicates per treatment. After thorough mixing, the samples were left undisturbed for 30 min before undergoing the extraction procedure described in Section 2.2.3. The final extracts were collected in 2 mL injection vials and analyzed by gas chromatography to determine ethylicin recovery across different matrices.

2.2.5. Uptake and Translocation of Ethylicin in Field-Grown Tomato Plants

To determine the absorption time of ethylicin in field-grown tomato plants, two treatments were applied: one with ethylicin alone and one with a compound formulation in which ethylicin/oligosaccharins/deionized water = 4:1:5. The concentrations used were 400 mg/L and 800 mg/L. Each treatment received 100 mL of solution, with three replications per treatment. Samples of the upper, middle, and lower leaves were collected at 1, 2, 4, 6, and 24 h after irrigation to track the absorption of ethylicin.

Subsequently, 1000× and 2000× dilutions of ethylicin alone and the ethylicin–oligosaccharin complex were applied to field-grown tomatoes via root irrigation, with three replications per treatment. Samples were collected 1, 3, 5, and 7 days after treatment. All samples were stored at −80 °C for analysis using the same extraction method outlined in Section 2.2.3.

2.2.6. Effects on Fusarium spp. and Phytophthora spp. After Field Root Irrigation

Seven days after root irrigation, tomato plants were sampled from the 1000× and 2000× dilution treatments. The samples were collected at 5 cm above the base of the plants at soil depths of 5, 10, and 15 cm, with three replications per treatment. The samples were then transported to the laboratory for analysis.

A soil suspension was made by weighing 5 g of soil sample and adding it to 95 mL of 0.7‰ sterilized agar water, followed by shaking and mixing. In a sterile environment, 1 mL of the soil suspension was then placed onto Petri dishes containing Komada’s medium [22] and Masago’s medium [23] for the isolation of Fusarium spp. and Phytophthora spp., respectively, and incubated for 2 to 3 days in a dark environment at 28 °C. The number of colonies of Fusarium spp. and Phytophthora spp. was then recorded.

2.3. Testing Conditions

Based on a study by Wenjing Li [3], the detection method was optimized as follows: The inlet temperature was set to 140 °C. The column temperature started at 50 °C, was held for 1 min, then increased to 145 °C at a rate of 10 °C/min. The sample was injected using a split injection with a split ratio of 10:1. The temperature of the mass spectrometry ionization source and the transmission line was set to 230 °C and 150 °C, respectively. Detection was performed using Selected Ion Monitoring (SIM) at m/z values of 61, 62, 75, 95, and 154. The solvent delay was set to 9.8 min. Under these conditions, the ethylicin peak appeared at 9.9 min, with a lowest limit of detection (LOQ) of 20 μg/kg.

2.4. Data Statistics

Data obtained from the GC–MS analysis were processed using Microsoft Excel 2010. The recovery rates, absorption, and translocation quantities of ethylicin in tomato plant tissues and soil, as well as their relative standard deviations, were calculated. Correlation curves were plotted using Origin8 software (version 8.0). One-way analysis of variance (ANOVA) and Duncan’s new complex polarity method were used to test significance of difference (α = 0.05).

In order to observe pesticide uptake and translocation patterns, root concentration factor (RCF), translocation factor (TF), and transpiration stream concentration factor (TSCF) were used [24].

The RCF was used to describe pesticide uptake by plant roots. A higher RCF indicates stronger uptake. The formula is as follows:

$$\mathrm{R}\mathrm{C}\mathrm{F}={\displaystyle \frac{C_{root}}{C_{soil}}},$$

where C_root refers to the concentration of compounds in the root and C_soil refers to the concentration of compounds in the soil, measured in mg/kg.

The TF describes the ability of a compound to move from the root system to the stems and leaves. It is calculated as follows:

$$\mathrm{T}\mathrm{F}={\displaystyle \frac{C_{shoot}}{C_{root}}}$$

The higher the TF, the greater the ability of the compound to move from the underground part of the plant to the aboveground part of the plant.

3. Results

3.1. Translocation of Ethylicin Uptake in Potted Tomato Plants

3.1.1. Additive Recovery Rate

The added concentration, average recovery rate, and coefficient of variation for ethylicin in the three matrices are shown in

Table 3. The recovery, coefficient of variation, and matrix standard curve for the addition of ethylicin to the three matrices.

Substrate	Fortified Level (mg/kg)	Average Recovery Rate (%, n = 3)	Coefficient of Variation (%, n = 3)
Root	1	85.94	7.43
	10	94.87	4.71
	100	93.64	2.64
Stem	1	115.29	8.60
	10	105.01	3.15
	100	104.07	3.33
Leaf	1	83.78	7.90
	10	101.85	3.77
	100	102.75	10.45

. Recovery ranged from 80% to 120%; ethylicin was not detected in the blank control samples. The coefficient of variation in all spiked concentrations was below 10%, indicating the pesticide residue detection method used in this experiment meets the necessary accuracy and sensitivity requirements for pesticide residue analysis.

3.1.2. Uptake and Translocation of Ethylicin in Tomato Plants

After a 500-fold dilution (500×) root irrigation of the tomato plants, symptoms of pesticide damage, such as wilting, yellowing, and dead leaves, appeared within 14 days. The concentration of ethylicin in the roots, stems, and leaves of the tomato plants within 14 days of the 500× application is shown in Figure 1. The detected concentration of ethylicin in the roots ranged from 0.03 to 0.14 mg/kg, in the stems from 0.05 to 0.12 mg/kg, and in the leaves from 0.001 to 0.22 mg/kg. The transfer factor of the stems and leaves was less than 1. Due to damage caused by the 500× treatment, the absorption and translocation capacity of ethylicin was low.

After spraying and root irrigation with the 1000× dilution, no visible symptoms of pesticide damage, such as wilting, yellowing, or dead leaves, were observed in the tomato plants within 14 days and no significant inhibitory effects were noted. The concentration of ethylicin in the roots, stems, and leaves of the plants during this period is shown in Figure 2A. Ethylicin was absorbed and translocated rapidly after foliar application. In the leaves, the concentration of ethylicin decreased quickly, while in the stems, it gradually declined and reached equilibrium by day 5. In the roots, the ethylicin concentration peaked at 2.93 mg/kg on day 3, then gradually decreased and stabilized by day 5. Over the 14-day period, the ethylicin concentration in the roots ranged from 1.02 to 2.93 mg/kg, in the stems from 1.10 to 2.36 mg/kg, and in the leaves from 0.83 to 2.20 mg/kg.

As shown in Figure 2B, after root application the concentration of ethylicin in the roots reached its maximum value of 5.07 mg/kg on day 5, then slowly decreased. In the stems, the concentration first decreased and then increased, reaching a peak of 3.95 mg/kg on day 14. The concentration in the leaves gradually increased and stabilized by day 5. After root application, the concentration of ethylicin in the roots, stems, and leaves ranged from 4.08 to 5.07 mg/kg, 3.10 to 3.95 mg/kg, and 3.44 to 3.68 mg/kg, respectively. The transport factor (TF), calculated as shown in Figure 2C, remained above 1, indicating strong translocation of ethylicin within the plant, with equilibrium reached by day 5.

3.2. The Absorption of Ethylicin in Tomato Plants in the Field

3.2.1. Field Uptake of Ethylicin After Root Irrigation

When ethylicin was applied to tomato roots at a concentration of 400 mg/kg, both the single-agent formulation and the compound formulation were absorbed by the plants within 6 h, as shown in Figure 3. The concentration of ethylicin in the upper leaves was higher for the compound formulation (5.87 mg/kg) compared to the single-agent formulation (3.85 mg/kg). These results indicate that the addition of oligosaccharins enhances the translocation of ethylicin to the upper leaves. Complete absorption was achieved within 24 h, with the ethylicin concentration in the upper leaves reaching 6.22 mg/kg for the single agent and 13.57 mg/kg for the compound formulation. No ethylicin was detected in the control group after application. These results confirm that tomato plants can rapidly absorb ethylicin without any damage within the first 6 h.

The same steps were repeated at a concentration of 800 mg/kg (Figure 4); again, both the single-agent and compound formulations were fully absorbed within 6 h. The concentration of ethylicin in the upper leaves was 1.38 mg/kg for the single-agent formulation and 3.24 mg/kg for the compound formulation, further confirmation that the presence of oligosaccharins enhanced the translocation of ethylicin to the upper leaves.

3.2.2. Leaf Absorption of Ethylicin 1000× and 2000× After Root Irrigation

The single-agent formulation and compound formulation of ethylicin were applied to tomato roots and the concentration of ethylicin in the upper leaves was measured at a 1000× dilution (Figure 5). It was noted that the presence of both treatment methods was detected in the upper leaves. On the first day, ethylicin was detected in the upper leaves at a concentration of 6.87 mg/kg for the single-agent formulation and 8.97 mg/kg for the compound formulation. The presence of oligosaccharins in the compound formulation enhanced the absorption of ethylicin, resulting in a higher concentration in the upper leaves.

Over time, ethylicin from the single-agent formulation was primarily concentrated in the middle and lower leaves, and its concentration gradually decreased, with complete dissipation by day 5. No ethylicin was detected in the control plants. All treated tomato plants absorbed ethylicin quickly without any visible damage.

The single-agent formulation and compound formulation of ethylicin were applied to tomato roots at a concentration of 2000× and the concentration of ethylicin in the upper leaves was measured (Figure 6). On the first day, ethylicin was detected in the upper leaves at a concentration of 10.71 mg/kg for the single-agent formulation and 23.57 mg/kg for the compound formulation.

The oligosaccharins in the compound promote ethylicin uptake; therefore, a higher concentration of ethylicin was detected in the upper leaves. Post-application monitoring revealed that ethylicin from the single-agent formulation was mainly concentrated in the middle and lower leaves and gradually dissipated over time, with complete dissipation by day 5.

3.2.3. Control Effects of Root Irrigation at Different Soil Depths

Seven days after root drenching with the 1000× diluted ethylicin solution, the soil samples were collected at soil depth. The best control of Fusarium spp. was observed at 5–10 cm, where the number of Fusarium spp. decreased to 1.67 CFU/g. At 0–5 cm, the number was 7.67 CFU/g, likely due to ethylicin’s volatility, which allowed it to be absorbed by the roots and diffused into the soil. At 10–15 cm, fewer Fusarium spp. were detected due to limited penetration at this depth. Therefore, a depth of 5–10 cm has the best control effect on Fusarium spp. The compound formulation (1000× dilution) achieved an even better effect, reducing the number of Fusarium spp. to 0 at 5–10 cm; the addition of oligosaccharins reduced the number of Fusarium spp.

The control effect on Phytophthora spp. was slightly weaker than on Fusarium spp., but the number of Phytophthora spp. decreased in all treatment groups compared to the control (as shown in Figure 7).

4. Discussion

In recent years, with the vigorous development of green agriculture, ethylicin is expected to become a new fumigant by virtue of its low-toxicity and low-residue properties. It can be used during the growing period of crops. Ethylicin can be rapidly degraded in crops without leaving pesticide residue, providing a new strategy for disease control during the growing period of crops. Studies have shown that the half-life of ethylicin in paddy water is only 0.3–1.1 days, with no residue detected in the soil, rice grains, or rice husks at the time of harvest [25]. This property eliminates the need for prolonged open aeration after soil fumigation, which is required for other fumigants, such as dazomet, metham sodium, and chloropicrin (e.g., dazomet decomposes more slowly in the soil and requires a longer period of open aeration to ensure safe planting of the next crop) [26,27]. Therefore, ethylicin can quickly initiate the planting process of the next crop, effectively addressing the challenge of time constraints and narrow operational windows. The present study further demonstrates that ethylicin is safe and reliable for use during the growing season and its use as a soil fumigation treatment does not cause damage to subsequent crops. This gives it a significant advantage in continuous cropping, especially for agricultural production patterns that require frequent crop rotation.

Ethylicin exhibits favorable uptake and translocation properties in crops, as it is readily absorbed by plants through the leaves, stems, and roots, and is rapidly distributed throughout the plant. Pesticide transport—specifically, the pathways and mechanisms by which a pesticide moves through a plant—is a key determinant of its effectiveness [28]. For ethylicin, the transport factor (TF) remained above 1 for 14 days, indicating efficient translocation from the roots to the shoots, where it accumulates. This strong upward mobility likely contributes to ethylicin’s potential to resist plant disease infection [29]. After foliar application, ethylicin is rapidly absorbed by leaves and distributed systemically, aligning with Han Ping’s findings of similar migration behaviors among different pesticides [30]. Like spirotetramat, ethylicin moves bidirectionally through the xylem and phloem, providing effective protection to new growth against pest eggs and larvae [31,32]. Similar observations have been made with glyphosate in non-target tea plants, where it is transported from roots to shoots via the vascular system [13]. Thus, ethylicin’s application, whether through root irrigation or foliar spraying, ensures thorough distribution throughout the plant, providing efficient pest and disease control during the growing season.

Oligosaccharides, acting as plant growth regulators, significantly enhance the absorption of ethylicin in plants. This finding provides a solid theoretical basis for the development of novel pesticide enhancers. Given the promoting effect of oligosaccharides on ethylicin absorption, it is reasonable to speculate that they may similarly affect the uptake of other pesticides. This indicates that oligosaccharides play an important role in improving pesticide absorption and distribution within plant tissues.

Yang Suping [33] found that spraying oligosaccharins reduces pest and disease incidence, promotes cherry tomato growth, decreases organic acid levels, increases vitamin C content, and improves overall tomato quality. Similarly, Zhu Yuyong [34] demonstrated that a treatment combining 5% oligosaccharins with 80% ethylicin effectively controls cotton yellow wilt, achieving a control rate exceeding 70%. Fu Xin [35] reported that the combination of ethylicin and oligosaccharins inhibits melon bacterial fruit blotch, where an optimal ratio of 4:1 provides the best control.

Collectively, these studies highlight the synergistic effects of oligosaccharins and ethylicin in enhancing plant health and controlling disease. Traditional fumigants are typically applied before planting; however, this study confirms that when combined with oligosaccharides, ethylicin can be effectively absorbed and transported within plants for the robust control of soil-borne diseases.

5. Conclusions

This study presents a sample pre-treatment and GC–MS method for the detection of ethylicin in the roots, stems, and leaves of tomato seedlings. This method is operationally straightforward and meets the sensitivity and accuracy requirements for pesticide analysis.

With this method, we elucidate the dynamic transmission pattern of ethylicin in tomato plants and the strong impact of application techniques on its distribution. The findings indicate that foliar application enables rapid ethylicin migration from leaves to roots, and root application markedly elevates the agent’s concentration throughout the plant, peaking at 5.07 mg/kg in the root system. Notably, the stems and leaves demonstrate strong conductivity, with a transport factor (TF) consistently exceeding 1 and rapidly reach equilibrium, confirming an efficient upward transport mechanism. From a practical standpoint, root application provides longer-lasting systemic protection and maintains stem and leaf concentrations between 3.10 and 3.95 mg/kg, while foliar application is better suited for short-term, targeted control. These results support the pivotal theoretical foundation for the optimization of ethylicin field-application strategies.

In this study, we investigated the absorption and degradation dynamics of ethylicin in plants and its effect on the number of pathogenic bacteria through root irrigation experiments in the field. The experimental results show that ethylicin is characterized by its rapid absorption and timely elimination. It can be absorbed by plants within 6 h and eliminated within 5 days. These findings provide a key reference for its rational application in practical agricultural production. Moreover, we found that ethylicin exhibits the most excellent inhibitory effect at a distance of 5–10 cm from plant roots. It effectively inhibits the growth of two common soil-borne pathogens, Fusarium spp. and Phytophthora spp., clarifying its target point in the prevention and control of soil-borne diseases. Further studies reveal a synergistic relationship between oligosaccharins and ethylicin. This relationship significantly promotes the absorption of ethylicin, enhances its inhibitory effect, and effectively reduces the populations of Fusarium spp. and Phytophthora spp. This finding provides a new strategy for the prevention and control of soil-borne diseases during plant growing season. These important findings not only provide a strong basis for the rational application of pesticides but also help to improve pesticide use efficiency and reduce the risk of pesticide residue. Additionally, they open up new ways for the prevention and control of soil-borne diseases, enriching their availability for this purpose.