Acute and Delayed Effects of Melatonin Pretreatment Against Cold Stress in Leek (Allium ampeloprasum L. var. porrum)
1
Department of Horticulture, Faculty of Agriculture, Erciyes University, 38030 Kayseri, Türkiye
2
Graduate School of Natural and Applied Sciences, Erciyes University, 38030 Kayseri, Türkiye
*
Author to whom correspondence should be addressed.
Horticulturae 2025, 11(10), 1208; https://doi.org/10.3390/horticulturae11101208
Submission received: 31 August 2025
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Revised: 6 October 2025
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Accepted: 7 October 2025
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Published: 7 October 2025
(This article belongs to the Special Issue Tolerance of Horticultural Plants to Abiotic Stresses)
Abstract
Although known as a cool-season vegetable, leeks (Allium ampeloprasum L. var. porrum) are particularly susceptible to sudden temperature drops during the seedling stage. While melatonin’s mitigating effect on abiotic stresses has been observed in many plants, very few studies have investigated its effects on leek seedlings under cold stress. In this study, leek seedlings grown under ideal conditions were sprayed with melatonin at concentrations of 0, 1, 2, 4, and 8 µM on the 45th and 60th days. Morphological data were recorded on the 74th day, and the following day, cold stress was applied at 0 °C for 0, 2, 4, 6, and 24 h. On the 76th day, the first biochemical analyses (antioxidant capacity, total phenolic content, total flavonoid content, and total soluble protein content) were performed. After a three-week growth period under ideal conditions, morphological measurements and biochemical analyses were repeated. According to the findings, treatments with 4 and 8 μM melatonin prevented cold-induced changes in the plants’ morphological data. It was determined that concentrations of 1 and 2 µM melatonin were more effective on biochemical contents. According to the study’s findings, melatonin treatment mitigated the harmful effects of cold stress on leeks. The results obtained indicate that melatonin is a promising tool for increasing plants’ resistance to cold stress.
1. Introduction
Melatonin is a fundamental phytohormone that acts as a “master regulator” in diverse directions of plant development [1]. Melatonin was initially identified as a hormone-like substance crucial for regulating growth and development in animals, as well as for signal transmission, by scientists who isolated it from the pineal glands of cattle. As technology advances and makes progress in identification, plant physiology, and cell biology, molecular biology, sequencing technology, and other research methods have also evolved. As a result, scientists have conducted more in-depth studies on melatonin production, its location, and its biological effects on plants [2].
The natural defense mechanisms of plants do not provide adequate protection under extreme stress conditions; therefore, exogenous bio-stimulants can be used to improve plant stress tolerance. Among these regulators, melatonin (N-acetyl-5-methoxytryptamine) has become a frequently tried agent among plants recently [3]. As a potent antioxidant, melatonin regulates stress-responsive genes, promotes water use and photosynthetic efficiency, and interacts synergistically with other plant hormones to support growth under adverse abiotic stress conditions [4]. Additionally, melatonin is reported to play a crucial role in the biosynthesis pathways of gibberellic acid and abscisic acid by regulating gene expression [5]. Melatonin modulates plant defense mechanisms in response to various abiotic stresses, including low-temperature stress [6,7,8].
Leek (Allium ampeloprasum L. var. porrum) is a cool-season vegetable that is known and consumed in many countries around the world. World leek production in 2022 was listed as 2.109.066 tons. According to the same year’s data, Indonesia ranks first in leek production quantity at 638.734 tons. Türkiye ranks second with 168.710 tons of production. In addition to these production values, Türkiye has a broad genetic diversity of leeks [9,10]. The breeding period lasts a considerable amount of time. Depending on the season, the time from sowing seeds to planting seedlings may take up to 3 months in some cases. Long-term cold stress, particularly during the seedling stage, results in various losses in leeks [11]. Although the mitigating effect of melatonin on abiotic stress has been observed in many plants, the number of studies investigating the effects of cold stress on leeks is very few. Leeks, a cold-climate vegetable, are tolerant of low temperatures. However, sudden drops in temperature following seed sowing, especially in October and November, can cause severe losses in plants.
In this study, we investigated several morphological and biochemical parameters in leaf seedlings treated with varying concentrations of exogenous melatonin. The central hypothesis of this study is that the effects of melatonin application on specific biochemical properties in leek seedlings exposed to cold stress for different durations change over time (H1). In the experiment, morphological and biochemical changes were measured at two time points, allowing for a comparison of the effects of cold stress and melatonin in different developmental periods. The study’s objective is to investigate the morphological and biochemical effects of different durations of cold stress on leek seedlings at the early stage of development. Additionally, it aimed to investigate the potential effects of exogenous melatonin treatment over two weeks.
2. Materials and Methods
2.1. Plant Growth and Melatonin Treatments
The experiment was carried out in the laboratories and greenhouses of Erciyes University. Seeds of leek cultivar (Allium ampeloprasum L var. porrum, cv İnegöl-92), packaged by Arzuman Seed Company, were obtained from Güçlü Supply Electronic Services LLC. Konya-Türkiye. The seeds were sown in vials in a climate chamber under controlled conditions (16/8 h of day/night at +22 °C and +16 °C, with cool fluorescent lamps providing approximately 225 μmol m−2 s−1 PAR) in February 2022.
The study conducted by Yağmur and Hanci [12] on onions was referenced in determining melatonin concentrations. Plants were sprayed twice with melatonin solutions at concentrations of 0, 1, 2, 4, and 8 µM when the number of leaves was approximately two to three (45th day) and four to five (60th day). The volumes used were 10 and 20 mL per plant, respectively.
2.2. Stress Conditions
This experiment was designed to observe the protective effects of melatonin on morphological characteristics in response to cold stress and to evaluate its effects on biochemical contents immediately and three weeks later. Therefore, the stress level that could have killed the plants was avoided. Cold stress treatments were initiated on the 75th day of the study. The reason for starting the stress treatments 15 days after the last melatonin application is to allow sufficient time for the plant’s defense mechanisms to activate and for the synthesis of protective compounds. Cold stress was applied at 0 °C in a fully controlled climate chamber for four different durations: 2, 4, 6, and 24 h. During cold stress, conditions encountered in conventional cultivation were simulated by adjusting the light/dark conditions. In this context, since cold stress is more likely to occur during nighttime hours, stress periods of 2, 4, and 6 h were applied under dark conditions. However, since there is also a possibility of cold stress occurring throughout the entire day, and keeping plants in darkness for 24 h is not feasible in real-life conditions, the 24 h stress was applied under 8 h of darkness and 16 h of light (provided by cold fluorescent lamps delivering approximately 225 μmol m−2s−1 of light).
2.3. Morphological Measurements
Before the stress treatments, the first morphological measurements and observations were made two weeks after the completion of melatonin treatments (74th day of the experiment). The second set of morphological measurements and observations was made three weeks after the cold stress treatment (97th day of the experiment). The following features were examined within the scope of morphological measurements: whole plant length (from soil level to the tip of the longest leaf), diameter of white pseudostem (cm); leaf crown development (upright: 1, semi-erect: 3, broad: 5); leaf color (yellow-green: 1, green: 3, gray-green: 5, blue-green: 7); plant fresh weight (g); length of longest leaf (cm); the width of the broadest part of the longest leaf (mm); and white pseudostem length (cm).
2.4. Biochemical Analysis
The first biochemical analysis was conducted the day following the cold stress (day 76 of the experiment), and the subsequent analysis occurred three weeks later (day 97). The ferric reducing antioxidant power (FRAP) method was used to determine the antioxidant capacity. [13]. The results were computed as equivalent to Trolox based on the curve derived from the calibration formula established with Trolox.
The total phenolic content of the samples was determined using Folin–Ciocalteu’s method, as developed by Singleton and Rossi [14]. The absorbance results of the samples were quantified as gallic acid equivalents in mg/100 g of dry matter by plotting a graph with standard solutions prepared at various concentrations of gallic acid (mg/mL) and applying the derived formula.
The technique established by Zhishen et al. [15] was employed to quantify the total flavonoid content. The total flavonoid content of the plants was expressed as mg quercetin equivalent (QUE) per gram of extract.
Lowry et al. [16] devised a method for quantifying soluble protein content. Soluble protein concentrations in mg BSA/mL buffer were determined using the specified formula.
2.5. Statistical Analysis
The study employed a completely randomized design (CRD) with three replications. In each replicate, three plants were measured. Using the Shapiro–Wilk test, normality was determined prior to data processing. A two-way analysis of variance (ANOVA) was employed to assess the significance of variations in treatment concerning melatonin concentrations, stress conditions, and their interactions across all attributes. Tukey’s Honest Significant Difference (HSD) test was used as a post hoc test for intergroup comparisons at a significance level of p < 0.05. The data were analyzed using the SAS 9.1 statistical package program.
3. Results
3.1. Results of Morphological Measurements
3.1.1. Results of First and Second Measurements
In the first measurement results, since the plants had not yet experienced any cold stress, only the effects of the melatonin concentration on development were evaluated (Table 1). At this measurement stage, the effect of melatonin was found to be statistically significant on all parameters except for whole plant length and width on the longest leaf, according to the ANOVA test (p < 0.05).
After the first morphological measurements and observations, the plants were exposed to cold stress for durations of 0, 2, 4, 6, and 24 h. Then, the climate chamber was adjusted to ideal conditions again, and the plants were grown under these conditions for another 21 days. In the second measurement results, the effects of melatonin concentrations on plants, the duration of cold stress, and their interaction were evaluated together, and the differences between the results were examined using the ANOVA test (p < 0.05). According to the results obtained, only the duration of cold stress had a statistically significant effect on the length of the longest leaf data; only the effect of melatonin concentrations had a statistically significant effect on leaf crown development and diameter of pseudo white stem (mm) data; and the interaction effect of melatonin concentrations x cold stress duration had a statistically significant effect on plant fresh weight. Neither melatonin concentrations, cold duration, nor their interaction effect had a statistically significant effect on the other parameters. (p < 0.05).
In the first and second measurement results, the effect of melatonin concentrations on whole plant length was not statistically significant (Table 2). In contrast to the first measurements, this difference in the second measurement shows that the effect of melatonin on whole plant length in leek seedlings occurs at a later stage. Among the other concentrations, 4 µM and 8 µM melatonin were less effective, with low increase rates of 7.0% and 7.7% at 24 h of cold for the whole plant length.
As melatonin concentrations increased, plant white pseudostem lengths showed a tendency to decrease (Table 2). In the initial measurement results, the longest white stem length was measured in plants without melatonin supplementation (6.3 cm), while this value decreased to 2.3 cm at the highest melatonin concentration.
High-dose melatonin treatments (4 and 8 µM) caused the length of the longest leaf of the plants to increase at the first measurement (Table 2). In this parameter, 2 µM melatonin + 0 h of cold treatment provided the highest increase with 100.8%, according to the second measurement results. This indicates that melatonin promotes leaf growth at low concentrations. In the measurement results for the length of the longest leaf, the difference between the effects of melatonin concentrations in the first measurement results was found to be statistically significant. The highest values were obtained at high concentrations (4 and 8 µM) (29.8 and 30.2 cm, respectively). In the second measurement, the difference between melatonin doses was found to be statistically insignificant.
In the second measurement, 2 µM melatonin was the most effective dose in increasing the width of the longest leaf in plants exposed to cold stress (Table 2). Higher melatonin concentrations (4 µM and 8 µM) showed limited positive effects on the width of the longest leaf. For prolonged cold treatments (24 h), the increases in width of the longest leaf reached the lowest values.
In the first measurement, where only the effects of melatonin were monitored without any stress condition, it was observed that the effect of the 2 µM melatonin concentration was more pronounced on plant fresh weight values (15.8 g) (Table 3). Melatonin treatments above the 2 µM dose led to partial reductions in this value. The second measurement monitored the effects of cold stress and melatonin; 4 µM and 8 µM melatonin concentrations provided more limited increases; even these concentrations caused negative results within 24 h of cold treatment. In the 24 h cold treatment, the fresh weight of plants without melatonin decreased to as low as 2.1%.
In the first measurement, the effects of melatonin on leaf color were statistically significant. At this stage, the leaf colors of non-melatonin-treated plants were evaluated as being darker than those of the others. In the second measurement and the rate of change between the two measurements, leaf color changes were minimal.
3.1.2. Results of Changes Between Two Measurements
For a more comprehensive data analysis, the percentages of change between the results obtained from measurements made in two different periods were calculated, and the differences between the results were examined using the ANOVA test (p < 0.05) (Table 1). When evaluating the percentage changes in results between the two measurement periods, it was determined that melatonin concentrations alone had a statistically significant effect on the parameters of length of the longest leaf, leaf crown development, and diameter of white stem; melatonin concentrations and cold stress duration separately had a statistically significant effect on white pseudostem length; and melatonin concentrations and cold duration interaction had a statistically significant effect on the parameters of plant length and plant fresh weight. No significant difference was detected between the width of the longest leaf and the length of the white pseudostem, resulting in this measurement period (p < 0.05) (Table 2 and Table 3).
The most significant percentage change in whole plant length values was observed in samples treated with 2 μM melatonin without exposure to cold stress, reaching 56.8%. The lowest increase in this regard was observed in plants treated with 1 μM melatonin and maintained at cold conditions for 24 h (2.1%). The most important data regarding the percentage change in whole plant length values is that the effect of 24 h cold stress on plants not treated with melatonin was more positive than the results obtained with other melatonin doses (Table 2).
When examining the percentage changes in white pseudostem length values, statistically significant differences were observed based on melatonin concentration and average stress duration (Table 2). The most tremendous change for this parameter was observed in samples exposed to 24 h of cold stress (−50.4%). As melatonin concentrations increased, the percentage changes in white pseudostem length decreased. In this sense, the most protective effect was obtained at an 8 μM melatonin concentration (−11.7%).
No statistically significant differences were observed in the values or percentage changes when examining the width of the longest leaf. In general, increasing stress durations reduced the amount of increase in this value (Table 2).
In the values of the longest leaf length, only the effect of melatonin concentrations on percentage changes was found to be statistically significant. In general, increasing melatonin concentrations reduced the amount of increase in this value. This was observed based on melatonin concentration and the average duration of stress. The most tremendous change between the first and second measurement results was observed in plants treated with a 2 μM concentration of melatonin (64.9%) (Table 2).
In plant fresh weight values, the interaction effect of melatonin concentration × cold stress duration on percentage changes was found to be statistically significant (Table 3). For this parameter, the most significant change was observed in samples supplemented with 2 µM melatonin and not exposed to cold stress (104.2%). In samples with a melatonin concentration of 1 µM that were supplemented and exposed to 6 and 24 h of cold stress, losses were observed in the second measurement compared to the first measurement (−14.8% and −34.3%, respectively).
In leaf color values, only the effect of melatonin concentrations on percentage changes was found to be statistically significant (Table 3). The most tremendous change for this parameter was observed in samples supplemented with 1 µM of melatonin (40.0%).
In leaf crow development values, only the effect of melatonin concentrations on percentage changes was found to be statistically significant (Table 3). In general, as melatonin concentrations increased, percentage changes also increased negatively. The lowest change was detected in the control group samples (0.2%). At melatonin concentrations of 4 and 8 μM, greater changes were observed (−50.0% and −55.3%, respectively).
3.2. Results of Biochemical Analysis
Biochemical analyses were performed on the 75th day (immediately after cold stress treatment) and on the 97th day. Unlike morphological properties, the interaction between melatonin and cold stress has a significant impact on the results obtained for the four parameters examined. Therefore, the average melatonin or cold duration values were ignored when evaluating the results.
On day 97, the second test showed the highest total flavonoid content (83.762 mg quercetin/g) in 8 µM melatonin and plants that had never been exposed to cold stress (Table 4).
On the 75th day, the total flavonoid content was determined to be 53.762 mg quercetin/g in plants grown under the same conditions (8 µM melatonin and no stress applied). Another interesting finding about the total flavonoid content is that the lowest value (22.333 mg quercetin/g) was found in plants that had not been sprayed with melatonin and were not under any stress. However, these plants had a considerably higher total flavonoid content than the others, as measured after 22 days (85.190 mg quercetin/g). Exposure to cold for 24 h resulted in reductions in total flavonoids in the second measurement in all melatonin groups except for the 1 µM melatonin treatment. When examining the total phenolic content results, the five lowest values were predominantly observed in the second measurement, which occurred on the 97th day, and were typically found in plants that had been exposed to cold stress for 24 h (Table 4).
These plants were given either no melatonin or extreme limits (1 µm and 8 µM) of melatonin. When the three highest values of the total phenolic content were examined, it was determined that there was a balance between the melatonin dose and the cold time, regardless of the measurement time. Accordingly, 6 h of cold in plants without melatonin, cold stress for 4 h in plants given 4 µM melatonin, and zero hours in plants given 8 µM melatonin increased the total amount of phenolic substances. As a result of the antioxidant capacity (FRAP) determination, the highest values were determined in the first and second measurements of the plants that were not given melatonin and were exposed to cold stress for 2–4 h (Table 4).
This situation suggests that the low-temperature period of 2–4 h, without any external treatment, naturally increases the antioxidant level (FRAP) in leek plants, a vegetable native to cool climates. When the lowest 10 values regarding antioxidant capacity are examined, it is seen that 6 and 24 h cold stress is effective regardless of melatonin concentration. These values were generally obtained in the second measurement. When the rate of change between the first and second measurements is considered, all melatonin concentrations omitted for 24 h have significantly lower antioxidant capacities. The highest increase rate (32.051%) was found in plants in the 8 µM melatonin group without stress. The second measurement of soluble protein, similar to the FRAP results, showed an increase in 2 h cold stress in plants without melatonin (Table 5).
When the duration of cold stress exceeded 2 h, however, the amount of soluble protein observed in the second measurement decreased compared to the first melatonin measurement, and this reversal only occurred in the results for the 4 h stress duration. However, no dose of melatonin was effective in either the 6 or 24 h treatments. The most significant change between the two analysis periods occurred in total flavonoid content (237%) in plants that were not sprayed with melatonin and were not exposed to cold stress. In the control group (0 µM melatonin), the total flavonoid content generally decreased between the two measurement periods, especially at 24 h of cold stress.
In 4 and 8 µM melatonin treatments, flavonoid content increased significantly, especially after 6 h of stress. In the 8 µM treatment, the highest increases were detected at 4–6 h. In the control group, phenolic content showed mostly negative changes between the two measurement periods. In 4–8 µM melatonin treatments, phenolic content increased between the two periods, especially in the 6 h stress period. At the 8 µM concentration, the highest increases were also measured at 24 h. Between the two measurement periods, significant increases in antioxidant capacity were measured in the 4 and 8 µM melatonin treatments at the 4–6 h stress period.
4. Discussion
Decreased growth was the most recognizable indication of cold stress in the leek plants compared with plants maintained at normal conditions (22/16 °C). Melatonin supplementation significantly reduced the limited growth caused by cold stress. These findings demonstrate that exogenous melatonin treatment improved the cold stress tolerance of leeks. Exogenous melatonin supplementation in plants enhances cold tolerance by improving biochemical defense mechanisms. This supplementation increases the total soluble protein content, enriches the osmolyte content in cells, and enhances the synthesis of antifreeze proteins that protect cells from low-temperature stress.
Furthermore, melatonin, a potent antioxidant, slightly increases FRAP antioxidant capacity, rendering reactive oxygen species (ROS) caused by low-temperature stress harmless and reducing oxidative damage. Melatonin promotes the production of secondary metabolites in plants, thereby increasing the total phenolic and flavonoid content. These compounds also contribute to antioxidant activity and stabilize cell membranes, thereby enhancing the plant’s physiological resistance to cold weather conditions. Recently, it was reported that melatonin-added plants, such as strawberries [17], tomatoes [18], Arabidopsis thaliana [19], Bermuda grass [20], and cucumbers [21], grew better and were less stressed by cold weather. Numerous concentrations have been examined in studies assessing the effects of melatonin on plants subjected to cold stress. A 1 µM dose of melatonin has been demonstrated to be efficacious in barley [22]. It was also successful at 5 µM of melatonin in pepper and eggplant [23,24]. It was effective at 200 µM of melatonin in cucumber [21] and 150 µM in rice [25]. In this study, we preferred using the lower concentrations that have been tried on onion plants [12]. Melatonin and its metabolites, among the best-known antioxidants, are classified as both hydrophilic and hydrophobic because they are soluble in both water and oil. These features allow them to freely cross cell membranes and disperse in any aqueous compartment, such as the cytosol, nucleus, and mitochondria [26,27]. Forming the first line of defense against oxidative stress and promoting growth are among the primary functions of melatonin [28,29].
This study indicated that exogenous melatonin treatment can partially recover cold-induced declines in young leek plants. However, the reducing effect varied in some parameters based on the melatonin concentrations and the duration of the cold. While the effects of cold stress can sometimes appear immediately, they can also manifest themselves in later developmental periods. The effects of external growth regulators on plants can be observed similarly, albeit gradually. Based on these facts, in our study, the effects of melatonin and cold stress were measured twice, three weeks apart. Melatonin promotes plant growth by increasing cellular antioxidant defenses.
Additionally, it promotes cell division through synergistic effects with hormones, such as auxin. Melatonin has been reported to regulate gene expression that increases cell elongation, especially at low concentrations [30]. In our study, while melatonin promoted growth at low concentrations (2 µM), it showed limited or adverse effects on growth at high concentrations (6–8 µM). This suggests that melatonin synergizes with growth hormones in the optimum dose range, but oxidative balance may be disrupted at high concentrations. It has been reported that melatonin promotes plant growth under abiotic stress; however, high concentrations can have adverse effects on plant growth. In the study investigating the contribution of melatonin to plant elongation by reducing oxidative stress, growth inhibition was observed at high concentrations [31]. Similar results were obtained in our study. High melatonin concentrations had limited effects on plant length. Changes in the width of the longest leaf depend on melatonin dose and cold duration. 2 µM melatonin was the most effective dose in increasing this width. Prolonged cold treatments significantly reduced the width of the longest leaf. The finding suggests that melatonin may play a role in maintaining cellular wall structure and water balance in leaf cells. Arnao and Hernández-Ruiz [32] reported that melatonin maintains turgor pressure in leaf cells by preventing water loss. Plant fresh weights were significantly increased by 2 µM melatonin. High concentrations of melatonin may have impaired photosynthetic processes. Prolonged cold treatments caused a significant decrease in fresh weight. Tan et al. [26] showed that melatonin increases biomass production by regulating carbon fixation and sugar metabolism. The fresh weight increases in our data are consistent with these findings.
Several studies have found that melatonin treatment can increase antioxidant capacity and delay postharvest senescence in sweet cherries [33], fresh-cut broccoli [34], and peaches [35]. The phenylpropanoid pathway activity, which is mediated by phenylalanine ammonia-lyase and chalcone synthase activity, contributes to the accumulation of phenols, flavonoids, and anthocyanins in fruits and vegetables by providing phenylalanine [25]. Furthermore, Yang et al. [36] suggest that attenuating phenol oxidation with low polyphenol oxidase expression and activity may lead to a higher accumulation of phenols, flavonoids, and anthocyanins in fruits and vegetables. Additionally, it has been previously reported that exogenous melatonin treatment increases FRAP values in cotton [37] and wheat [38] under drought stress conditions. These results are consistent with our findings. We found that cold stress in limited periods naturally increases the FRAP level in leeks without needing external treatment. The phenomenon can explain that leeks are a cold-season vegetable and that cold stress in limited periods can have beneficial effects due to their nature. The positive effect of melatonin was evident in the damage caused by cold stress that lasted for 24 h.
Total flavonoid amounts are generally increased in plants exposed to cold stress, indicating that flavonoids play a critical role in how plants cope with cold stress through their antioxidant properties [39]. Flavonoids protect cells against oxidative damage by neutralizing cold stress-induced reactive oxygen species (ROS). However, melatonin supplementation may further enhance this process. In the literature, melatonin has been reported to promote plant flavonoid production by increasing the activity of antioxidant enzymes and regulating defense metabolism [6]. For example, melatonin treatment has been shown to alleviate the adverse effects of cold stress and increase plant resilience by activating the flavonoid biosynthesis pathway [40]. Increases in the carbohydrate content of plants subjected to cold stress have been reported previously [41].
Additionally, it was reported that the amount of soluble protein and phenolic compounds increased significantly in the leaves of wheat seedlings under cold stress (5 °C day/night for 3 days) [29]. While the increase in soluble protein content was approximately 12% in plants subjected to cold stress alone, the treatment of melatonin and cold stress resulted in a higher (up to 20%) increase in soluble protein content. Using melatonin has also been shown to increase soluble protein in rapeseed [42] and watermelon plants when subjected to cold weather stress [43]. In our study, similar to the FRAP results, we found that 2 h cold stress increased soluble protein levels in the second measurement, which did not receive any melatonin. However, as the duration of the cold stress increased, the amount of soluble protein observed in the second measurement decreased compared to the first measurement. The measurement of melatonin reversed this reduction in the outcomes of the 4 h stress period. However, no melatonin dose was effective in either the 6 or 24 h treatments.
5. Conclusions
The findings indicated that external melatonin treatment significantly mitigated the impact of cold stress in leek seedlings. This effect, nevertheless, fluctuated based on the duration of cold exposure, the dosage of melatonin, and the timing of measurement. These results validate the study’s initial hypothesis. According to the results, 4 and 8 µM melatonin were the most effective concentrations for all morphological characters before the stress treatment (74-day-old plants). Morphological measurements 21 days after the cold stress treatment showed that 2 µM melatonin treatment positively affected morphological data. Concentrations of 1 and 2 µM melatonin had the highest positive effect on the total flavonoid content and FRAP value of the plants after one day of continuous cold stress. A similar effect was observed for total phenolic matter at a 4 µM dose and soluble protein content at a 2 µM melatonin dose. Findings suggest that melatonin may be a promising agent for enhancing plant resilience to cold stress. The research elucidates the formulation of novel strategies to alleviate the detrimental impacts of cold stress on agricultural output. It provides a basis for investigating the potential benefits of melatonin treatments in other plants. Future studies under different environmental conditions and in various plant species will contribute to a broader understanding of melatonin’s effects and expand its treatment areas. However, our study examined only the effects of specific melatonin concentrations (0, 1, 2, 4, and 8 µM). Evaluating a wider range of doses and different treatment times may provide more comprehensive results.
Furthermore, the fact that the study was conducted only under controlled conditions may limit the estimation of the results obtained under real field conditions. Therefore, it is recommended that similar studies be conducted under different environmental conditions and in different plant species. This study serves as an important reference, demonstrating that using melatonin as a stress management tool against cold stress can be beneficial for leeks and other plants.
Author Contributions
Conceptualization, F.H.; methodology, F.H. and A.H.H.H.; software, F.H.; validation, F.H. and A.H.H.H.; formal analysis, F.H.; investigation, F.H. and A.H.H.H.; resources, F.H.; data curation, F.H.; writing—original draft preparation, F.H.; writing—review and editing, F.H.; visualization, F.H.; supervision, F.H. All authors have read and agreed to the published version of the manuscript.
Funding
This study was not supported by any sponsor or funder.
Data Availability Statement
The original contributions presented in this study are included in the article. Further inquiries can be directed to the corresponding author.
Acknowledgments
We would like to thank the Proofreading & Editing Office of the Dean for Research at Erciyes University.
Conflicts of Interest
The authors declare no conflicts of interest.
Abbreviations
The following abbreviations are used in this manuscript:
| FRAP | Ferric-reducing antioxidant power method |
| MEL | Melatonin |
| ROS | Reactive oxygen species |
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Table 1.
Summary of the results of the ANOVA test (p < 0.05).
| Observations | Factors | Whole Plant Length | White Pseudostem Length | Width of the Longest Leaf | Length of the Longest Leaf | Plant Fresh Weight | Leaf Color | Leaf Crow Development | Diameter of White Pseudostem |
|---|---|---|---|---|---|---|---|---|---|
| 1st Measurement | Melatonin concentrations | ns | ns | ns | ns | * | * | * | * |
| 2nd Measurement | Melatonin concentrations | ns | ns | ns | ns | ns | ns | ns | * |
| Stress conditions | ns | ns | ns | * | ns | ns | ns | ns | |
| Interaction effect | ns | ns | ns | ns | * | ns | ns | ns | |
| Change ratio | Melatonin concentrations | ns | * | ns | * | ns | * | * | ns |
| Stress conditions | ns | * | ns | ns | ns | ns | ns | ns | |
| Interaction effect | * | ns | ns | ns | * | ns | ns | ns |
*: Significant at the p < 0.05 level according to the ANOVA test. ns: non-significant.
Table 2.
Some morphological measures result from melatonin and cold treatments on the 75th and 97th days *.
Table 2.
Some morphological measures result from melatonin and cold treatments on the 75th and 97th days *.
| Melatonin (µM) | Cold Duration (Hours) | Whole Plant Length (cm) | White Pseudo Stem Length (cm) | The Width of the Broadest Part of the Longest Leaf (mm) | Length of the Longest Leaf (cm) | ||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| 1st Measurement | 2nd Measurement | Change (%) | 1st Measurement | 2nd Measurement | Change (%) | 1st Measurement | 2nd Measurement | Change (%) | 1st Measurement | 2nd Measurement | Change (%) | ||
| 0 | 0 | 29.81 ± 0.36 ** ns | 41.41 ± 0.51 | 39.21 ± 0.47 b | 6.31 ± 0.08 | 3.91 ± 0.05 | −36.90 ± 0.47 | 5.30 ± 0.06 | 6.32 ± 0.08 | 20.01 ± 0.24 | 24.11 ± 0.29 | 39.60 ± 0.45 | 64.51 ± 0.72 |
| 2 | 37.90 ± 0.47 | 27.40 ± 0.33 c | 3.62 ± 0.04 | −43.11 ± 0.52 | 6.10 ± 0.05 | 16.72 ± 0.22 | 37.32 ± 0.45 | 54.81 ± 0.66 | |||||
| 4 | 33.42 ± 0.41 | 12.23 ± 0.17 e | 3.43 ± 0.04 | −45.02 ± 0.54 | 5.81 ± 0.07 | 10.02 ± 0.12 | 32.81 ± 0.39 | 35.90 ± 0.43 | |||||
| 6 | 35.11 ± 0.43 | 18.14 ± 0.24 d | 2.71 ± 0.03 | −56.91 ± 0.65 | 5.70 ± 0.04 | 9.21 ± 0.14 | 43.30 ± 0.52 | 79.70 ± 0.96 | |||||
| 24 | 34.40 ± 0.40 | 15.52 ± 0.19 d | 1.94 ± 0.02 | −70.01 ± 0.84 | 5.50 ± 0.07 | 5.00 ± 0.06 | 30.01 ± 0.36 | 24.30 ± 0.29 | |||||
| 1 | 0 | 29.50 ± 0.35 | 33.10 ± 0.41 | 12.13 ± 0.17 e | 3.01 ± 0.04 | 2.02 ± 0.02 | −33.31 ± 0.42 | 5.50 ± 0.07 | 7.72 ± 0.10 | 40.01 ± 0.45 | 23.80 ± 0.29 | 37.50 ± 0.45 | 57.93 ± 0.69 |
| 2 | 40.53 ± 0.49 | 37.31 ± 0.45 b | 2.13 ± 0.03 | −29.20 ± 0.35 | 6.61 ± 0.08 | 19.22 ± 0.25 | 37.61 ± 0.45 | 58.42 ± 0.71 | |||||
| 4 | 37.52 ± 0.46 | 27.01 ± 0.32 c | 1.81 ± 0.02 | −39.62 ± 0.45 | 6.10 ± 0.07 | 10.80 ± 0.13 | 35.61 ± 0.43 | 50.12 ± 0.63 | |||||
| 6 | 33.13 ± 0.42 | 12.10 ± 0.17 e | 1.24 ± 0.02 | −58.31 ± 0.71 | 6.01 ± 0.04 | 8.30 ± 0.12 | 30.31 ± 0.36 | 27.71 ± 0.33 | |||||
| 24 | 30.12 ± 0.36 | 2.12 ± 0.03 e | 1.1 ± 0.01 | −64.61 ± 0.72 | 5.80 ± 0.07 | 5.80 ± 0.07 | 30.20 ± 0.36 | 27.10 ± 0.33 | |||||
| 2 | 0 | 28.53 ± 0.34 | 44.74 ± 0.54 | 56.83 ± 0.65 a* | 2.50 ± 0.03 | 2.12 ± 0.03 | −16.70 ± 0.23 | 6.02 ± 0.07 | 8.71 ± 0.16 | 44.40 ± 0.53 | 23.52 ± 0.28 | 47.20 ± 0.57 | 100.81 ± 1.21 |
| 2 | 34.81 ± 0.42 | 22.12 ± 0.27 d | 2.31 ± 0.04 | −8.01 ± 0.01 | 8.30 ± 0.11 | 38.61 ± 0.46 | 45.10 ± 0.54 | 91.92 ± 1.14 | |||||
| 4 | 38.63 ± 0.46 | 35.63 ± 0.43 b | 2.14 ± 0.03 | −16.71 ± 0.22 | 7.42 ± 0.10 | 23.92 ± 0.29 | 40.51 ± 0.49 | 72.51 ± 0.87 | |||||
| 6 | 33.91 ± 0.41 | 18.82 ± 0.25 d | 1.50 ± 0.02 | −41.72 ± 0.53 | 6.81 ± 0.08 | 13.31 ± 0.16 | 36.30 ± 0.47 | 54.60 ± 0.66 | |||||
| 24 | 29.43 ± 0.35 | 3.04 ± 0.04 e | 1.30 ± 0.02 | −50.02 ± 0.64 | 6.60 ± 0.08 | 10.30 ± 0.12 | 24.61 ± 0.3 | 4.81 ± 0.06 | |||||
| 4 | 0 | 32.50 ± 0.39 | 36.22 ± 0.43 | 11.41 ± 0.14 e | 2.82 ± 0.03 | 2.21 ± 0.03 | −20.81 ± 0.25 | 6.01 ± 0.07 | 7.20 ± 0.10 | 20.20 ± 0.24 | 29.81 ± 0.36 | 38.61 ± 0.46 | 29.61 ± 0.36 |
| 2 | 36.12 ± 0.40 | 11.22 ± 0.13 e | 1.72 ± 0.04 | −39.60 ± 0.45 | 7.21 ± 0.10 | 19.22 ± 0.25 | 34.92 ± 0.42 | 17.30 ± 0.21 | |||||
| 4 | 39.51 ± 0.47 | 21.41 ± 0.26 d | 1.41 ± 0.03 | −47.91 ± 0.57 | 6.72 ± 0.08 | 11.90 ± 0.14 | 35.91 ± 0.43 | 20.52 ± 0.25 | |||||
| 6 | 35.53 ± 0.43 | 9.13 ± 0.14 e | 1.51 ± 0.02 | −45.81 ± 0.55 | 6.62 ± 0.09 | 9.20 ± 0.14 | 34.02 ± 0.41 | 14.30 ± 0.17 | |||||
| 24 | 35.01 ± 0.42 | 7.70 ± 0.10 e | 1.14 ± 0.01 | −60.40 ± 0.71 | 6.30 ± 0.07 | 4.20 ± 0.05 | 32.10 ± 0.39 | 7.81 ± 0.10 | |||||
| 8 | 0 | 32.32 ± 0.39 | 36.32 ± 0.47 | 12.70 ± 0.17 e | 2.31 ± 0.03 | 2.12 ± 0.03 | −8.30 ± 0.11 | 6.02 ± 0.07 | 7.90 ± 0.10 | 31.50 ± 0.38 | 30.21 ± 0.36 | 37.60 ± 0.45 | 24.41 ± 0.29 |
| 2 | 42.01 ± 0.53 | 30.23 ± 0.36 c | 2.11 ± 0.03 | −8.31 ± 0.12 | 6.91 ± 0.08 | 14.92 ± 0.16 | 37.11 ± 0.45 | 22.90 ± 0.27 | |||||
| 4 | 38.62 ± 0.46 | 19.81 ± 0.24 d | 1.73 ± 0.04 | −25.01 ± 0.33 | 6.90 ± 0.06 | 15.51 ± 0.19 | 34.81 ± 0.42 | 15.20 ± 0.16 | |||||
| 6 | 37.53 ± 0.45 | 16.32 ± 0.23 d | 2.12 ± 0.03 | −8.30 ± 0.12 | 6.51 ± 0.08 | 8.02 ± 0.12 | 33.51 ± 0.43 | 10.29 ± 0.13 | |||||
| 24 | 34.53 ± 0.41 | 7.01 ± 0.08 e | 2.11 ± 0.03 | −8.30 ± 0.10 | 6.51 ± 0.05 | 8.30 ± 0.14 | 32.80 ± 0.39 | 8.51 ± 0.15 | |||||
| Means (hours) | 0 | 38.31 ± 0.46 | 26.41 ± 0.32 | 2.51 ± 0.03 | −23.22 ± 0.28 A | 7.61 ± 0.10 | 31.21 ± 0.37 | 40.10 ± 0.45 A | 55.41 ± 0.66 | ||||
| 2 | 38.32 ± 0.46 | 25.63 ± 0.29 | 2.40 ± 0.03 | −24.03 ± 0.29 A | 7.01 ± 0.09 | 21.70 ± 0.26 | 38.41 ± 0.46 A | 49.11 ± 0.59 | |||||
| 4 | 37.52 ± 0.45 | 23.21 ± 0.28 | 2.13 ± 0.03 | −34.83 ± 0.42 B | 6.61 ± 0.08 | 14.42 ± 0.17 | 35.92 ± 0.43 AB | 38.82 ± 0.47 | |||||
| 6 | 35.03 ± 0.42 | 14.91 ± 0.16 | 1.82 ± 0.04 | −42.23 ± 0.51 B | 6.30 ± 0.08 | 9.61 ± 0.12 | 35.51 ± 0.43 AB | 37.51 ± 0.45 | |||||
| 24 | 32.70 ± 0.39 | 7.11 ± 0.10 | 1.54 ± 0.02 | −50.60 ± 0.61 B | 6.10 ± 0.07 | 6.70 ± 0.08 | 29.90 ± 0.36 B | 14.51 ± 0.17 | |||||
| Means (Melatonin) | 0 | 36.41 ± 0.47 | 22.50 ± 0.27 | 3.13 ± 0.04 | −50.41 ± 0.61 b | 5.92 ± 0.07 | 12.20 ± 0.17 | 36.61 ± 0.47 | 51.81 ± 0.62 ab | ||||
| 1 | 34.82 ± 0.42 | 18.11 ± 0.24 | 1.62 ± 0.02 | −45.01 ± 0.54 b | 6.41 ± 0.09 | 16.82 ± 0.24 | 34.30 ± 0.41 | 44.20 ± 0.53 ab | |||||
| 2 | 36.30 ± 0.47 | 27.21 ± 0.33 | 1.91 ± 0.04 | −25.02 ± 0.31 ab | 7.62 ± 0.10 | 26.10 ± 0.29 | 38.81 ± 0.47 | 64.92 ± 0.71 a | |||||
| 4 | 36.42 ± 0.47 | 12.10 ± 0.17 | 1.62 ± 0.03 | −26.62 ± 0.51 ab | 6.80 ± 0.08 | 12.91 ± 0.17 | 35.12 ± 0.42 | 17.91 ± 0.21 b | |||||
| 8 | 37.83 ± 0.45 | 17.23 ± 0.21 | 2.04 ± 0.05 | −11.72 ± 0.14 a | 6.91 ± 0.09 | 15.70 ± 0.19 | 35.11 ± 0.42 | 16.40 ± 0.23 b | |||||
* Letters were assigned based on the ANOVA test results. Accordingly, the interaction effect between melatonin dose and cold duration on the rate of change in the whole plant length parameter; the separate effects of melatonin doses and cold durations on the rate of change in the white pseudo stem length parameter; cold durations in the second measurement of the length of the longest leaf parameter; and melatonin doses on the rates of change in the length of the longest leaf parameter were lettered because they were statistically significant (p < 0.05). Uppercase/lowercase letters for each measurement parameter and measurement period indicate a statistically significant difference according to Tukey’s Honest Significant Difference (HSD) test (p < 0.05). ** ns: Non-significant.
Table 3.
Some morphological measures result from melatonin and cold treatments on the 75th and 97th days.
Table 3.
Some morphological measures result from melatonin and cold treatments on the 75th and 97th days.
| Melatonin (µM) | Duration (hours) | Plant Fresh Weight (g) | Leaf Color | Leaf Crow Development | Diameter of White Pseudo Stem (mm) | ||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| 1st Measurement | 2nd Measurement | Change (%) | 1st Measurement | 2nd Measurement | Change (%) | 1st Measurement | 2nd Measurement | Change (%) | 1st Measurement | 2nd Measurement | Change (%) | ||
| 0 | 0 | 10.60 ± 0.14 e | 14.91 ± 0.15 c | 40.70 ± 0.44 g | 2.01 ± 0.02 a | 2.20 ± 0.03 | 8.30 ± 0.11 | 3.40 ± 0.04 b | 2.01 ± 0.02 | −43.31 ± 0.52 | 2.21 ± 0.03 b | 2.31 ± 0.03 | 2.51 ± 0.03 |
| 2 | 13.30 ± 0.15 c | 25.01 ± 0.33 h | 1.72 ± 0.02 | −16.70 ± 0.23 | 4.41 ± 0.05 | 24.30 ± 0.29 | 2.30 ± 0.04 | 2.30 ± 0.04 | |||||
| 4 | 12.61 ± 0.13 d | 19.21 ± 0.25 i | 1.70 ± 0.02 | −16.71 ± 0.22 | 3.40 ± 0.04 | −3.31 ± 0.04 | 2.31 ± 0.03 | 2.30 ± 0.03 | |||||
| 6 | 12.42 ± 0.13 d | 16.50 ± 0.20 k | 1.31 ± 0.02 | −33.31 ± 0.40 | 3.51 ± 0.04 | 0.01 ± 0.01 | 2.32 ± 0.02 | 2.32 ± 0.04 | |||||
| 24 | 11.02 ± 0.14 d | 3.62 ± 0.04 m | 1.32 ± 0.02 | −33.30 ± 0.41 | 4.42 ± 0.05 | 23.31 ± 0.28 | 2.20 ± 0.03 | 2.20 ± 0.03 | |||||
| 1 | 0 | 11.70 ± 0.14 d | 18.03 ± 0.22 c | 53.81 ± 0.63 d | 1.00 ± 0.01 b | 1.52 ± 0.02 | 50.01 ± 0.60 | 3.01 ± 0.04 c | 1.51 ± 0.02 | −50.01 ± 0.62 | 2.11 ± 0.03 b | 2.20 ± 0.02 | 2.60 ± 0.03 |
| 2 | 12.71 ± 0.13 d | 8.41 ± 0.13 m | 1.50 ± 0.02 | 50.01 ± 0.62 | 3.51 ± 0.04 | 16.72 ± 0.23 | 2.10 ± 0.03 | 2.01 ± 0.02 | |||||
| 4 | 12.20 ± 0.13 d | 3.92 ± 0.05 m | 1.50 ± 0.02 | 50.00 ± 0.62 | 2.52 ± 0.03 | −16.72 ± 0.22 | 2.11 ± 0.04 | 1.71 ± 0.03 | |||||
| 6 | 10.01 ± 0.12 d | −14.80 ± 0.15 n | 1.51 ± 0.02 | 50.01 ± 0.61 | 3.51 ± 0.04 | 16.71 ± 0.21 | 2.11 ± 0.03 | 1.60 ± 0.02 | |||||
| 24 | 7.71 ± 0.09 e | −34.30 ± 0.41 o | 1.01 ± 0.01 | 0.01 ± 0.01 | 3.00 ± 0.04 | 0.01 ± 0.01 | 2.11 ± 0.01 | 1.62 ± 0.01 | |||||
| 2 | 0 | 15.83 ± 0.19 a | 32.20 ± 0.33 a* | 104.20 ± 1.25 a | 1.50 ± 0.02 b | 2.02 ± 0.02 | 33.30 ± 0.41 | 3.40 ± 0.04 b | 4.60 ± 0.06 | 30.01 ± 0.36 | 2.30 ± 0.03 b | 2.40 ± 0.03 | 2.51 ± 0.03 |
| 2 | 29.33 ± 0.35 a | 85.90 ± 1.03 b | 2.01 ± 0.02 | 33.30 ± 0.42 | 5.61 ± 0.07 | 60.02 ± 0.72 | 2.41 ± 0.01 | 2.31 ± 0.03 | |||||
| 4 | 28.21 ± 0.34 a | 78.81 ± 0.92 b | 1.22 ± 0.01 | −16.71 ± 0.22 | 2.10 ± 0.03 | −40.00 ± 0.48 | 2.41 ± 0.03 | 2.61 ± 0.03 | |||||
| 6 | 22.32 ± 0.27 b | 41.41 ± 0.52 f | 1.20 ± 0.01 | −16.71 ± 0.21 | 2.71 ± 0.03 | −23.31 ± 0.28 | 2.41 ± 0.04 | 3.20 ± 0.04 | |||||
| 24 | 20.33 ± 0.24 b | 28.80 ± 0.35 h | 1.21 ± 0.01 | −16.72 ± 0.22 | 2.23 ± 0.03 | −36.70 ± 0.44 | 2.31 ± 0.03 | 2.02 ± 0.02 | |||||
| 4 | 0 | 13.32 ± 0.15 c | 20.02 ± 0.24 b | 50.40 ± 0.60 d | 1.51 ± 0.02 b | 1.2 ± 0.01 | −16.71 ± 0.22 | 4.52 ± 0.05 a | 2.40 ± 0.03 | −46.71 ± 0.56 | 2.91 ± 0.03 a | 3.01 ± 0.04 | 3.51 ± 0.04 |
| 2 | 18.91 ± 0.25 b | 41.81 ± 0.50 g | 1.21 ± 0.01 | −16.71 ± 0.23 | 2.71 ± 0.03 | −40.01 ± 0.48 | 3.02 ± 0.03 | 3.41 ± 0.04 | |||||
| 4 | 16.41 ± 0.23 c | 23.01 ± 0.28 h | 1.20 ± 0.01 | −16.70 ± 0.20 | 1.90 ± 0.02 | −56.70 ± 0.68 | 3.02 ± 0.05 | 3.41 ± 0.05 | |||||
| 6 | 16.30 ± 0.22 c | 22.63 ± 0.27 h | 1.21 ± 0.01 | −16.71 ± 0.21 | 1.51 ± 0.02 | −66.70 ± 0.80 | 3.02 ± 0.04 | 3.20 ± 0.04 | |||||
| 24 | 14.30 ± 0.17 c | 7.82 ± 0.09 l | 1.21 ± 0.01 | −16.71 ± 0.21 | 2.71 ± 0.03 | −40.01 ± 0.48 | 3.01 ± 0.03 | 2.91 ± 0.03 | |||||
| 8 | 0 | 13.81 ± 0.17 b | 22.41 ± 0.27 d | 62.61 ± 0.75 c | 1.01 ± 0.01 b | 1.01 ± 0.01 | 0.01 ± 0.01 | 4.01 ± 0.05 a | 2.50 ± 0.03 | −36.71 ± 0.44 | 2.20 ± 0.03 b | 2.30 ± 0.03 | 2.61 ± 0.03 |
| 2 | 20.32 ± 0.24 d | 47.92 ± 0.53 d | 1.52 ± 0.02 | 50.0 ± 0.61 | 1.11 ± 0.01 | −73.31 ± 0.88 | 2.21 ± 0.04 | 1.90 ± 0.02 | |||||
| 4 | 20.03 ± 0.24 d | 45.11 ± 0.54 e | 1.01 ± 0.01 | 0.01 ± 0.01 | 1.51 ± 0.02 | −63.32 ± 0.76 | 2.20 ± 0.04 | 2.31 ± 0.03 | |||||
| 6 | 15.30 ± 0.15 c | 11.50 ± 0.14 k | 1.51 ± 0.02 | 50.01 ± 0.60 | 2.12 ± 0.03 | −46.72 ± 0.56 | 2.21 ± 0.03 | 2.20 ± 0.04 | |||||
| 24 | 14.41 ± 0.17 c | 4.62 ± 0.06 m | 1.02 ± 0.01 | 0.01 ± 0.01 | 1.71 ± 0.02 | −56.71 ± 0.68 | 2.21 ± 0.02 | 2.21 ± 0.03 | |||||
| Means (hours) | 0 | 21.53 ± 0.26 ** | 62.31 ± 0.75 | 1.63 ± 0.02 | 15.01 ± 0.15 | 2.60 ± 0.03 | −29.32 ± 0.35 | 2.40 ± 0.03 | 2.71 ± 0.03 | ||||
| 2 | 18.92 ± 0.25 | 41.80 ± 0.52 | 1.60 ± 0.02 | 20.00 ± 0.24 | 3.51 ± 0.04 | −2.53 ± 0.03 | 2.41 ± 0.04 | 2.41 ± 0.03 | |||||
| 4 | 17.90 ± 0.21 | 34.02 ± 0.41 | 1.32 ± 0.02 | 0.01 ± 0.01 | 2.32 ± 0.03 | −36.01 ± 0.46 | 2.41 ± 0.03 | 2.40 ± 0.02 | |||||
| 6 | 16.61 ± 0.22 | 15.42 ± 0.15 | 1.41 ± 0.02 | 6.70 ± 0.08 | 2.72 ± 0.03 | −24.01 ± 0.29 | 2.41 ± 0.03 | 2.50 ± 0.03 | |||||
| 24 | 15.01 ± 0.15 | 2.11 ± 0.03 | 1.20 ± 0.01 | −13.30 ± 0.15 | 2.81 ± 0.03 | −22.02 ± 0.26 | 2.40 ± 0.02 | 2.21 ± 0.02 | |||||
| Means (Melatonin) | 0 | 12.80 ± 0.13 | 21.01 ± 0.25 | 1.41 ± 0.02 | −18.30 ± 0.22 b | 3.50 ± 0.04 a | 0.21 ± 0.01 a | 2.31 ± 0.03 ab | 2.31 ± 0.03 | ||||
| 1 | 12.13 ± 0.13 | 22.01 ± 0.26 | 1.41 ± 0.02 | 40.01 ± 0.48 a | 2.81 ± 0.03 ab | −6.71 ± 0.08 ab | 2.10 ± 0.02 b | 1.90 ± 0.02 | |||||
| 2 | 26.51 ± 0.32 | 67.80 ± 0.77 | 1.20 ± 0.01 | 3.31 ± 0.04 ab | 3.41 ± 0.04 bc | −2.00 ± 0.02 ab | 2.41 ± 0.03 ab | 2.50 ± 0.03 | |||||
| 4 | 17.20 ± 0.21 | 29.10 ± 0.35 | 1.00 ± 0.01 | −16.70 ± 0.21 b | 2.22 ± 0.03 bc | −50.01 ± 0.62 b | 3.01 ± 0.04 a | 3.31 ± 0.04 | |||||
| 8 | 18.53 ± 0.22 | 34.32 ± 0.41 | 1.21 ± 0.01 | 20.00 ± 0.24 ab | 1.80 ± 0.02 c | −55.31 ± 0.66 b | 2.21 ± 0.03 b | 2.21 ± 0.03 | |||||
* Letters were assigned based on the ANOVA test results. Accordingly, the effects of melatonin doses on the first measurements of the plant fresh weight, leaf color, leaf crow development, and diameter of white pseudostem parameters; the interaction effect between melatonin dose and cold duration on the second measurement and change ratio of the plant fresh weight parameter; melatonin doses on the change ratios of the leaf color and leaf crow development parameters; and melatonin doses on the second measurement of the diameter of white pseudostem were lettered because they were statistically significant (p < 0.05). Upper-case/lowercase letters for each measurement parameter and measurement period indicate a statistically significant difference according to Tukey’s Honest Significant Difference (HSD) test (p < 0.05). **: Non-significant.
Table 4.
Results of total flavonoid and total phenolic content of leek seedlings.
| Melatonin (µM) | Cold Duration (Hours) | Total Flavonoid (mg quercetin/g) | Total Phenolic (GA mg/g) | ||||
|---|---|---|---|---|---|---|---|
| Before | After | Change (%) | Before | After | Change (%) | ||
| 0 | 0 | 22.333 ± 0.510 q | 75.190 ± 0.521 c | 236.679 ± 5.321 a | 26.397 ± 0.301 l | 27.931 ± 0.471 k | 5.813 ± 0.171 d |
| 2 | 51.857 ± 0.510 i | 62.810 ± 0.530 e | 21.121 ± 0.940 g | 27.614 ± 0.330 k | 32.376 ± 0.621 def | 17.245 ± 0.350 a | |
| 4 | 55.667 ± 0.521 f | 58.048 ± 0.910 f | 4.277 ± 0.500 m | 31.159 ± 0.611 f–i | 27.931 ± 0.533 k | −10.358 ± 0.332 m | |
| 6 | 81.381 ± 0.510 a* | 72.333 ± 0.901 d | −11.118 ± 0.522 o | 36.450 ± 0.522 a | 29.730 ± 0.562 ij | −18.435 ± 0.323 q | |
| 24 | 46.143 ± 0.501 kl | 39.476 ± 0.521 k | −14.448 ± 0.502 p | 27.614 ± 0.322 k | 16.767 ± 0.272 o | −39.280 ± 0.343 t | |
| 1 | 0 | 54.238 ± 0.521 fgh | 56.810 ± 0.520 f | 4.741 ± 0.511 lm | 31.741 ± 0.452 fg | 31.460 ± 0.640 fgh | −0.883 ± 0.101 i |
| 2 | 63.762 ± 0.522 e | 71.857 ± 0.510 d | 12.696 ± 0.521 i | 31.106 ± 0.344 f–i | 32.751 ± 0.501 cde | 5.290 ± 0.324 e | |
| 4 | 69.952 ± 0.523 c | 80.322 ± 1.113 b | 14.824 ± 0.532 h | 31.529 ± 0.422 fgh | 33.270 ± 0.471 cd | 5.521 ± 0.231 de | |
| 6 | 47.571 ± 0.511 k | 51.355 ± 0.541 h | 7.953 ± 0.513 jk | 30.630 ± 0.540 h | 22.481 ± 0.320 m | −26.602 ± 0.654 r | |
| 24 | 47.571 ± 0.501 k | 48.555 ± 0.532 i | 2.068 ± 0.564 n | 30.894 ± 0.343 ghi | 21.476 ± 0.323 n | −30.485 ± 0.514 s | |
| 2 | 0 | 25.190 ± 0.501 p | 54.714 ± 0.822 g | 117.202 ± 1.521 b | 30.894 ± 0.423 ghi | 30.947 ± 0.613 gh | 0.171 ± 0.130 h |
| 2 | 53.286 ± 0.542 ghi | 56.619 ± 0.921 fg | 6.256 ± 0.221 kl | 32.799 ± 0.540 de | 34.693 ± 0.681 b | 5.775 ± 0.261 d | |
| 4 | 44.714 ± 0.522 l | 48.888 ± 0.750 i | 9.334 ± 0.234 j | 29.042 ± 0.350 j | 30.661 ± 0.402 hi | 5.575 ± 0.254 de | |
| 6 | 39.000 ± 0.510 n | 42.810 ± 0.531 j | 9.768 ± 0.502 j | 30.365 ± 0.412 i | 27.614 ± 0.322 k | −9.061 ± 0.320 l | |
| 24 | 71.857 ± 0.573 b | 61.857 ± 0.514 e | −13.917 ± 0.400 p | 34.122 ± 0.551 b | 29.942 ± 0.360 ij | −12.250 ± 0.312 n | |
| 4 | 0 | 35.667 ± 0.531 o | 54.714 ± 0.544 g | 53.405 ± 0.502 d | 33.540 ± 0.510 bcd | 33.566 ± 0.750 c | 0.079 ± 0.133 h |
| 2 | 40.905 ± 0.520 m | 56.143 ± 0.511 fg | 37.253 ± 0.541 f | 29.307 ± 0.440 j | 32.481 ± 0.441 de | 10.832 ± 0.270 b | |
| 4 | 65.667 ± 0.521d | 71.666 ± 0.902 d | 9.136 ± 0.194 j | 33.910 ± 0.402 bc | 36.175 ± 0.421 a | 6.678 ± 0.130 c | |
| 6 | 49.476 ± 0.550 j | 51.571 ± 0.522 h | 4.234 ± 0.563 m | 33.963 ± 0.521 bc | 31.778 ± 0.651 efg | −6.434 ± 0.214 k | |
| 24 | 52.810 ± 0.542 hi | 39.286 ± 0.530 k | −25.609 ± 0.500 q | 30.683 ± 0.541 hi | 29.048 ± 0.562 j | −5.329 ± 0.181 j | |
| 8 | 0 | 53.762 ± 0.531 gh | 83.762 ± 0.524 a | 55.802 ± 0.440 c | 34.492 ± 0.590 b | 35.339 ± 0.621 ab | 2.454 ± 0.151 f |
| 2 | 55.667 ± 0.521 f | 79.857 ± 0.521 b | 43.456 ± 0.404 e | 32.958 ± 0.424 cde | 33.566 ± 0.562 c | 1.846 ± 0.142 g | |
| 4 | 45.667 ± 0.522 l | 61.857 ± 0.542 e | 35.454 ± 0.411 f | 32.111 ± 0.344 ef | 33.016 ± 0.630 cd | 2.818 ± 0.122 f | |
| 6 | 54.714 ± 0.553 fg | 62.333 ± 0.522 e | 13.925 ± 0.413 hi | 34.439 ± 0.461 b | 29.466 ± 0.420 j | −14.442 ± 0.313 o | |
| 24 | 55.667 ± 0.530 f | 40.429 ± 0.500 k | −27.374 ± 0.210 q | 31.423 ± 0.301 fgh | 26.450 ± 0.411 l | −15.828 ± 0.383 p | |
| Mean (hours) | 0 | 38.238 ± 0.521 | 65.038 ± 0.821 | 93.565 ± 1.130 | 31.412 ± 0.591 | 31.848 ± 0.620 | 1.526 ± 0.133 |
| 2 | 53.095 ± 0.650 | 65.457 ± 0.711 | 24.1561 ± 0.434 | 30.756 ± 0.562 | 33.173 ± 0.432 | 8.197 ± 0.252 | |
| 4 | 56.333 ± 0.714 | 64.156 ± 0.732 | 14.604 ± 0.454 | 31.550 ± 0.642 | 32.210 ± 0.560 | 2.046 ± 0.221 | |
| 6 | 54.428 ± 0.521 | 56.080 ± 0.500 | 4.952 ± 0.221 | 33.169 ± 0.601 | 28.213 ± 0.544 | −14.994 ± 0.321 | |
| 24 | 54.809 ± 0.520 | 45.920 ± 0.414 | −15.855 ± 0.331 | 30.947 ± 0.442 | 24.736 ± 0.432 | −20.634 ± 0.324 | |
| Mean (melatonin) | 0 | 51.476 ± 0.622 | 61.571 ± 0.640 | 47.302 ± 0.531 | 29.846 ± 0.423 | 26.947 ± 0.314 | −9.003 ± 0.380 |
| 1 | 56.619 ± 0.732 | 61.779 ± 0.622 | 8.456 ± 0.250 | 31.179 ± 0.561 | 28.287 ± 0.304 | −9.431 ± 0.271 | |
| 2 | 46.809 ± 0.510 | 52.977 ± 0.524 | 25.728 ± 0.422 | 31.444 ± 0.532 | 30.771 ± 0.241 | −1.958 ± 0.154 | |
| 4 | 48.904 ± 0.523 | 54.676 ± 0.444 | 15.683 ± 0.324 | 32.280 ± 0.601 | 32.609 ± 0.531 | 1.165 ± 0.120 | |
| 8 | 53.095 ± 0.641 | 65.647 ± 0.733 | 24.252 ± 0.531 | 33.084 ± 0.611 | 31.567 ± 0.470 | −4.630 ± 0.242 | |
* For each measurement parameter and measurement period, different lowercase letters in each column indicate a statistically significant difference according to Tukey’s Honest Significant Difference (HSD) test (p < 0.05).
Table 5.
Results of antioxidant capacity and total soluble protein content of leek seedlings.
| Melatonin (µM) | Cold Duration (Hours) | FRAP Antioxidant Capacity (mM Tloroks/g) | Total Soluble Protein (mg/mL) | ||||
|---|---|---|---|---|---|---|---|
| Before | After | Change (%) | Before | After | Change (%) | ||
| 0 | 0 | 14.982 ± 0.401 g | 14.921 ± 0.321 f | −0.410 ± 0.141 jk | 12.878 ± 0.361 k | 23.077 ± 0.570 gh | 79.202 ± 0.912 a |
| 2 | 32.000 ± 0.611 a* | 31.995 ± 0.662 a | −0.016 ± 0.154 lj | 19.110 ± 0.443 gh | 27.279 ± 0.603 de | 42.747 ± 0.720 b | |
| 4 | 31.912 ± 0.423 a | 31.702 ± 0.452 a | −0.660 ± 0.130 k | 23.758 ± 0.570 de | 21.833 ± 0.322 i | −8.102 ± 0.243 n | |
| 6 | 13.667 ± 0.310 j | 9.895 ± 0.321 j | −27.599 ± 0.614 o | 29.944 ± 0.553 a | 27.844 ± 0.560 d | −7.013 ± 0.201 m | |
| 24 | 9.105 ± 0.230 n | 6.112 ± 0.234 m | −32.871 ± 0.541 r | 14.122 ± 0.360 jk | 9.251 ± 0.331 p | −34.491 ± 0.522 s | |
| 1 | 0 | 23.316 ± 0.441 b | 23.509 ± 0.43 b | 0.828 ± 0.13 gh | 28.477 ± 0.580 b | 31.723 ± 0.620 a | 11.401 ± 0.232 fg |
| 2 | 17.877 ± 0.402 cd | 18.825 ± 0.311 c | 5.299 ± 0.204 d | 24.122 ± 0.531 cde | 30.343 ± 0.530 b | 25.788 ± 0.411 c | |
| 4 | 13.754 ± 0.331 j | 14.807 ± 0.342 f | 7.653 ± 0.334 c | 24.016 ± 0.452 cde | 26.892 ± 0.510 e | 11.973 ± 0.230 f | |
| 6 | 9.193 ± 0.211 n | 9.333 ± 0.202 k | 1.527 ± 0.102 f | 23.383 ± 0.301 e | 22.362 ± 0.46 hi | −4.367 ± 0.103 l | |
| 24 | 11.737 ± 0.321 m | 10.544 ± 0.334 i | −10.164 ± 0.32 l | 16.270 ± 0.333 i | 13.765 ± 0.370 o | −15.395 ± 0.350 p | |
| 2 | 0 | 13.404 ± 0.443 jk | 13.982 ± 0.414 g | 4.319 ± 0.224 e | 15.038 ± 0.410 ij | 17.056 ± 0.461 l | 13.425 ± 0.342 e |
| 2 | 18.404 ± 0.343 c | 19.123 ± 0.511 c | 3.908 ± 0.143 e | 25.061 ± 0.604 cd | 28.601 ± 0.531 c | 14.125 ± 0.330 e | |
| 4 | 11.386 ± 0.353 m | 12.719 ± 0.354 h | 11.710 ± 0.330 b | 16.423 ± 0.324 i | 20.601 ± 0.410 j | 25.443 ± 0.443 c | |
| 6 | 14.368 ± 0.310 hi | 9.982 ± 0.343 j | −30.525 ± 0.450 q | 25.131 ± 0.441 cd | 23.106 ± 0.431 gh | −8.061 ± 0.262 n | |
| 24 | 13.316 ± 0.401 jkl | 9.561 ± 0.324 jk | −28.195 ± 0.423 p | 29.779 ± 0.603 ab | 23.650 ± 0.462 fg | −20.582 ± 0.471 q | |
| 4 | 0 | 14.807 ± 0.351 gh | 14.912 ± 0.441 f | 0.711 ± 0.120 gh | 17.913 ± 0.450 h | 19.042 ± 0.472 k | 6.303 ± 0.232 i |
| 2 | 14.895 ± 0.432 gh | 15.061 ± 0.531 f | 1.119 ± 0.101 fg | 20.460 ± 0.450 fg | 22.667 ± 0.563 h | 10.785 ± 0.241 g | |
| 4 | 16.211 ± 0.522 f | 16.053 ± 0.404 e | −0.974 ± 0.141 k | 25.214 ± 0.520 c | 29.768 ± 0.631 b | 18.062 ± 0.333 d | |
| 6 | 12.789 ± 0.314 l | 9.456 ± 0.124 jk | −26.063 ± 0.512 n | 20.026 ± 0.484 g | 17.538 ± 0.321 l | −12.425 ± 0.202 o | |
| 24 | 12.965 ± 0.402 kl | 9.263 ± 0.351 kk | −28.552 ± 0.534 p | 19.838 ± 0.341 g | 13.462 ± 0.221 o | −32.138 ± 0.643 r | |
| 8 | 0 | 17.789 ± 0.424 d | 23.491 ± 0.620 b | 32.051 ± 0.451 a | 21.469 ± 0.400 f | 23.488 ± 0.400 g | 9.403 ± 0.242 h |
| 2 | 16.825 ± 0.510 e | 16.842 ± 0.510 d | 0.104 ± 0.104 ij | 24.580 ± 0.571 cde | 24.309 ± 0.550 f | −1.100 ± 0.173 k | |
| 4 | 12.789 ± 0.440 l | 12.825 ± 0.404 h | 0.274 ± 0.133 hi | 20.002 ± 0.440 g | 20.862 ± 0.421 j | 4.295 ± 0.220 j | |
| 6 | 16.123 ± 0.601 f | 13.667 ± 0.414 g | −15.234 ± 0.361 m | 19.228 ± 0.302 gh | 16.120 ± 0.460 m | −16.164 ± 0.243 p | |
| 24 | 13.842 ± 0.451 ij | 8.754 ± 0.251 l | −36.755 ± 0.411 s | 23.934 ± 0.492 cde | 15.277 ± 0.312 n | −36.171 ± 0.742 t | |
| Mean (hours) | 0 | 16.859 ± 0.332 | 18.163 ± 0.351 | 7.499 ± 0.371 | 19.154 ± 0.240 | 22.877 ± 0.342 | 23.946 ± 0.5323 |
| 2 | 20.000 ± 0.312 | 20.369 ± 0.413 | 2.083 ± 0.164 | 22.666 ± 0.464 | 26.639 ± 0.470 | 18.468 ± 0.383 | |
| 4 | 17.210 ± 0.241 | 17.621 ± 0.444 | 3.600 ± 0.103 | 21.882 ± 0.310 | 23.991 ± 0.410 | 10.334 ± 0.202 | |
| 6 | 13.228 ± 0.253 | 10.466 ± 0.203 | −19.578 ± 0.434 | 23.542 ± 0.404 | 21.393 ± 0.341 | −9.606 ± 0.222 | |
| 24 | 12.192 ± 0.204 | 8.847 ± 0.131 | −27.307 ± 0.510 | 20.788 ± 0.344 | 15.081 ± 0.231 | −27.755 ± 0.504 | |
| Mean (melatonin) | 0 | 20.333 ± 0.314 | 18.924 ± 0.451 | −12.311 ± 0.421 | 19.962 ± 0.270 | 21.856 ± 0.314 | 14.468 ± 0.344 |
| 1 | 15.175 ± 0.342 | 15.404 ± 0.343 | 1.028 ± 0.183 | 23.253 ± 0.471 | 25.016 ± 0.584 | 5.880 ± 0.2232 | |
| 2 | 14.175 ± 0.211 | 13.073 ± 0.334 | −7.756 ± 0.250 | 22.286 ± 0.304 | 22.602 ± 0.532 | 4.870 ± 0.230 | |
| 4 | 14.333 ± 0.232 | 12.949 ± 0.203 | −10.751 ± 0.353 | 20.690 ± 0.390 | 20.495 ± 0.420 | −1.882 ± 0.111 | |
| 8 | 15.473 ± 0.351 | 15.115 ± 0.310 | −3.911 ± 0.141 | 21.842 ± 0.404 | 20.011 ± 0.340 | −7.947 ± 0.253 | |
* For each measurement parameter and measurement period, different lowercase letters in each column indicate a statistically significant difference according to Tukey’s Honest Significant Difference (HSD) test (p < 0.05).
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Hanci, F.; Hasan, A.H.H. Acute and Delayed Effects of Melatonin Pretreatment Against Cold Stress in Leek (Allium ampeloprasum L. var. porrum). Horticulturae 2025, 11, 1208. https://doi.org/10.3390/horticulturae11101208
AMA Style
Hanci F, Hasan AHH. Acute and Delayed Effects of Melatonin Pretreatment Against Cold Stress in Leek (Allium ampeloprasum L. var. porrum). Horticulturae. 2025; 11(10):1208. https://doi.org/10.3390/horticulturae11101208
Chicago/Turabian StyleHanci, Fatih, and Abbas Hussein Hasan Hasan. 2025. "Acute and Delayed Effects of Melatonin Pretreatment Against Cold Stress in Leek (Allium ampeloprasum L. var. porrum)" Horticulturae 11, no. 10: 1208. https://doi.org/10.3390/horticulturae11101208
APA StyleHanci, F., & Hasan, A. H. H. (2025). Acute and Delayed Effects of Melatonin Pretreatment Against Cold Stress in Leek (Allium ampeloprasum L. var. porrum). Horticulturae, 11(10), 1208. https://doi.org/10.3390/horticulturae11101208
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