The Effect of Late Frost Damage on the Growth and Development of Flower Organs in Different Types of Peach Varieties

Abstract

Late frost damage in spring is a significant limiting factor in peach industry development, with the flowering period being the most vulnerable to late frost. This study aimed to observe the flower organ state and physiological changes of two peach varieties under various temperature treatments and to provide a theoretical basis for selecting frost-resistant varieties. By analyzing the supercooling points of ‘Longyoutao 1’ (Y1) and ‘Longmi 15’ (L15), we simulated late frost at five temperatures, 4 °C, 2 °C, 0 °C, −2 °C, and −4 °C, and observed the flower organ changes at these five temperature stages during the flowering period. The contents of flower hormones (IAA, GA, ABA), membrane lipid peroxidation products (MDA), antioxidant enzymes (POD, SOD, CAT), and osmoregulatory substances (Pro, SS) were analyzed under various low-temperature stress conditions. The results showed no significant difference in flower morphology between Y1 and L15 at 4 °C, 2 °C, and 0 °C. At −2 °C, the anthers of Y1 turned brown and dried, the ovary froze, and water stains appeared on the sepals and the center. At −4 °C, the water stain on the ovary intensified, and the ovule froze. Moreover, by integrating the differences in the contents of IAA, GA, ABA, MDA, POD, SOD, and SS of the two varieties at the critical temperature of 0 °C, L15 showed the strongest ability to resist late frost. This study provides a physiological foundation for researching frost resistance during the flowering period.

1. Introduction

As an important fruit resource, peach (Prunus persica L.) is widely planted in China. According to the statistics of the Food and Agriculture Organization of the United Nations (FAO), in 2022, China had a peach planting area of 83.59 × 10⁴ hectares (https://www.sohu.com/a/762002012_121275473, accessed on 16 February 2025). Gansu Province is located in the northwest of China between the Loess Plateau and the Qilian Mountains, with unique climatic and geographical conditions and rich and diverse peach resources. It has become a key area for the preservation, evolution research, and cultivation practice of peach germplasm resources in China. According to statistics, as of 2019, the peach tree cultivation area in Gansu Province has expanded to 2.13 × 10⁴ hectares, becoming one of the important pillars to promote local economic development [1]. However, the spring temperature is changeable, the temperature difference between day and night is significant, and it is susceptible to unstable factors, especially frequent late frost damage, which affects the flowering of peach trees, resulting in a significant decline in the fruit setting rate, which seriously restricts the development of the peach industry [2].

Frost refers to the phenomenon in early spring when the temperature warms up and the short-term, near-ground temperature drops below 0 °C, causing water vapor in the air to condense into frost or water to freeze on the surface of plants, resulting in crop damage or death [3]. In the northwest of China, the spring climate is unstable, and early spring frost easily occurs during the critical period of peach flowering and fruit setting, resulting in reduced production or even no production [4]. The flowering period is the beginning of reproductive growth in peach trees, and the formation and opening of flowers are directly related to the development of later fruits, whose frost resistance temperature ranges from −2 °C to −1 °C [5]. The fruiting period is a critical stage for peach fruit formation, and the occurrence of frost may cause damage to flowers or fruit shedding, affecting the final yield [6]. The northern region is most affected by late frost damage. In the spring of 2023, there were four large-scale cooling processes in Gansu Province from April to May, with a long duration of low temperatures and a wide range of impacts. Especially in early April, the economic forests and fruits in the Hexi, Longzhong, and southeast regions were affected to varying degrees by multiple periods of low temperatures [7]. According to Yang [8] et al.’s research, there were five severe late frost damage incidents in Tianshui area from 2001 to 2018, each lasting 2–3 days, causing significant damage to the flower organs of fruit trees. From April to May 2022, severe late frost damage occurred in the Zhongning area of Ningxia, with varying degrees of frost damage observed in all five peach varieties. The total area of frost damage reached 11.0 hm², accounting for 59.5% of the peach cultivation area that year [9]. Therefore, it is extremely important to select and breed varieties that are resistant to late frost damage.

The determination of peach blossom organs, supercooling points, antioxidant enzymes, and hormones is of great significance for studying peach varieties that have suffered from late frost damage. The supercooling point is the temperature at which plant tissue begins to freeze. The lower the supercooling point, the stronger the plant tissue’s tolerance to low temperatures. For areas with frequent late frost damage, choosing peach varieties with lower supercooling points for planting can help reduce frost damage losses. The activity of antioxidant enzymes reflects the adaptability of plants to low-temperature stress. Hormones, as signaling molecules, participate in the response and adaptation process of plants to low-temperature stress, and changes in their content can reflect the physiological regulatory mechanisms within the plant. A series of advances have been made in research on the mechanism of peach late frost damage, such as the determination of the supercooling point and osmotic regulating substances of peaches under simulated late frost conditions [10]. The effect of low-temperature stress on membrane lipid peroxidation and protective enzyme activity in peach blossom organs has been studied [11]. These studies contribute to understanding the physiological response and adaptation mechanisms of peach varieties under low-temperature stress, as well as predicting the risk of frost damage.

According to climate observation regulations and the characteristics of late frost damage climates, a low-temperature day is defined when the temperature is ≤2.0 °C, a final frost day is defined when the ground temperature in spring is ≤0 °C or the last white frost appears in spring, and a heavy frost is defined when the lowest temperature on that day is ≤−2.0 °C [12]. There are differences in frost tolerance among different peach varieties. Studying the growth performance of different types of peach varieties under late frost conditions can provide a basis for variety improvement. By comparing and analyzing the flowering and fruit setting of different types of peach varieties under frost conditions, we can understand the physiological differences of different peach varieties and then selectively plant them to improve the stress resistance and economic benefits of peach trees. Therefore, during the late flowering period, we used a laboratory climate chamber to simulate late frost conditions (4 °C, 2 °C, 0 °C, −2 °C, −4 °C) and measured physiological and biochemical indicators during the flowering and fruiting stages of peach varieties (Longyoutao 1) and common peach varieties (Longmi 15). We compared their sensitivity differences to late frost damage, hoping to provide a reference and inspiration for research in related fields.

2. Materials and Methods

2.1. Test Materials

Two peach varieties, ‘Longyoutao 1’ (Y1) and ‘Longmi 15’ (L15), were selected as experimental materials at 10 days post-flowering. The registration numbers are, respectively, GPD Peach (2018) 620010 and Gan Ren Guo 2010001. These varieties are local resources of Gansu Province and preserved in the Peach Germplasm Bank of the Gansu Academy of Agricultural Sciences located in Gansu Province, China. All specimens were grafted onto red mountain peach rootstock (P. davidiana Franch). The region experiences a continental climate characterized by its geographical coordinates, longitude 103°41′ E and latitude 36°6′ N, with an altitude of 1530 m above sea level. The annual average temperature recorded was approximately 9.6 °C.

2.2. Test Methods

In April 2024, the flower organs in the middle of the branches of three trees with the same growth trend of each variety were used for independent biological duplication. After 10 days of the peak flowering period, 80 branches of equal length and uniform diameter (0.8–1.0 cm) were collected from four sides of a 10-year-old tree at a height of 1.8 m. Each variety was divided into 5 groups, and, in subsequent processing, each group was randomly assigned to 5 adjustable-temperature incubators, with temperatures set at 4 °C (control), 2 °C, 0 °C, −2 °C, and −4 °C. Cold treatment lasted for 2 h, with a cooling or heating rate of 1 °C/0.5 h [13,14,15]. The relative humidity recorded by the local meteorological station in Anning District, Lanzhou City, in early April 2024 fluctuated between 90% and 59%. To ensure the consistency of the influence of humidity on the test, the average relative humidity (65 ± 8% during the day and 82 ± 5% at night) was taken as the reference value. A humidification system was placed inside of the incubator to maintain consistent humidity between the chambers (the set value was 75 ± 3%RH, and the measured fluctuation range was 72–78%RH), ensuring that the critical humidity conditions for frost formation (>70%RH) were met. After cold treatment, we took photos of the freezing conditions of representative flower vases. Then, we immediately sampled the flower organs, froze them in liquid nitrogen, and stored them at −80 °C for further use.

2.3. Determination of Peach Blossom Supercooling Point

The floral organs were selected after 10 days of the peak flowering period to measure the supercooling point and freezing point. Each new shoot was 30 cm long and of equal length and consistent diameter. There were approximately 20 flowering organs on each new shoot, with 10 new shoots being used for each variety. The peach branches to be tested were placed in advance using hydroponics in an artificial experimental frost chamber, and a PT-100 thermocouple temperature sensor probe (The DHlihua; Shenzhen, China) was used to measure the flower organs after the peak flowering period of peach blossoms. The probe was placed at the ovary and then covered with plastic film. The natural frost cooling process was artificially simulated, with the cooling rate set to 1 °C/0.5 h. A total of 9 temperature levels were set up for the experiment, ranging from −4 to 4 °C. When the temperature dropped to the set value, it was maintained for 0.5 h. Then, the warming process of sunny weather in the same season in nature was simulated, with a heating rate of 1 °C/0.5 h. The freezing point and the supercooling point were recorded during the process.

2.4. Physiological Index Testing

Hormone levels were measured using the Enzyme Linked Immunoassay (ELISA) kit (96T), and the auxin (IAA) ELISA kit, gibberellin (GA) ELISA kit, and abscisic acid (ABA) ELISA kit (Shanghai Keaibo Biological, Shanghai, China) were used to determine the levels of plant hormones IAA, ABA, and GA. First, 0.1 g of floral samples was weighed and frozen in liquid nitrogen, and the mortar was pre-cooled. Then, the peach blossom container was placed into the mortar from liquid nitrogen, quickly ground, and placed in a pre-cooled centrifuge tube. Then, 1 mL of PBS buffer (pH 7.4) was added and vortexed for 30 s to ensure thorough mixing of the sample. We centrifuged it at 3000 r·min⁻¹ for 30 min, collected the supernatant, divided it, and determined the content of different hormones. The experimental method followed the instructions of the kit and was analyzed using an enzyme-linked immunosorbent assay (ELISA) reader Rayto RT-6100 (Rayto, Shanghai, China). Each sample was repeated three times. The level of lipid peroxidation is an indicator of oxidative stress, and the content of malondialdehyde (MDA) is determined by the thiobarbituric acid (TBA) reaction [16]. We used the reagent kit to measure the enzyme activities of peroxidase (POD), superoxide dismutase (SOD), and catalase (CAT) (provided by Suzhou Keming Biotechnology Co., Ltd., Suzhou, China) [17]. The concentration of proline (Pro) is a signal for osmotic regulation, which was evaluated using the Sulfosalicylate-acid ninhydrin method [18]. We determined the soluble sugar levels indicating carbohydrate metabolism using the anthrone colorimetric method [19].

2.5. Data Processing

Excel 2010 and SPSS Statistics 24.0 software packages were used for data organization and preliminary statistical analysis, and Origin 2018 64Bit was used for plotting. Each experiment used three independent biological replicates.

3. Results

3.1. Changes in Supercooling and Freezing Points of Flower Organs

The overcooling point (T1) and the freezing point (T2) can be found from the temperature curve. When the flower tissue is stressed to a certain extent by low temperature, the flower organ cell solution will change from a liquid to a solid state and release latent heat at the same time. The temperature curve shows a sudden rise in temperature and a jump in peak value. The ice crystal core is formed, the crystal grows, the temperature stops rising and begins to fall, the heat release and heat absorption reach an equilibrium state, and the temperature point is the freezing point (T2). As can be seen from Figure 1, with the change of the scale value of the galvanometer, the temperature change of Y1 and L15 is basically the same, but, in the range of the 1000–1500 scale, Y1 has a peak value, the highest value of the supercooling point T1 is −3.7 °C, and the lowest value is −4.6 °C. T2 is −3.7 °C. In the range of 1500~2000, L15 has a peak value, T1 is −7.2 °C, and T2 is −4.8 °C. The supercooling point temperature and freezing point temperature of L15 are lower than those of Y1 (

Table 1. Changes in supercooling point and freezing point of peach floral organ.

Tissues and Organs	Supercooling Point/°C	Freezing Point/°C
Y1	−4.6~−3.7	−6.3~−3.7
L15	−7.2~−4.8	−8.5~−4.8

).

3.2. Changes in Floral Organs After Freezing

It can be seen from Figure 2 that the ovaries of Y1 and L15 are normally developed when the temperature is above 0 °C. The anthers of Y1 are browning and shrinking under the low temperature of −2 °C, the ovaries are frozen, and the sepals and the center are water stained. The ovaries were watered, and the ovules were frozen at −4 °C. At −2 °C in L15, the ovary also suffered slight freezing injury, but the degree was lighter than that in Y1. At −4 °C, the anthers were browning, drying, and shrinking, the ovaries were severely frozen, the ovules were frozen, and the sepals and receptacles were watery.

3.3. Changes in Flower Organs’ Hormone Content

As shown in Figure 3A, the overall change in IAA content over five time periods (4 °C, 2 °C, 0 °C, −2 °C, and −4 °C) showed a trend of first decreasing and then increasing. At 0 °C, the IAA content reached the lowest point, and the Y1 and L15 content were 0.0137 μmol·g⁻¹ and 0.0141 μmol·g⁻¹, respectively. The content of Y1 was 0.026 μmol·g⁻¹ at 4 °C and 0.028 μmol·g⁻¹ at −4 °C, and the difference was significant. The IAA content of Y1 was 17.63% higher than that of L15 at −4 °C.

As shown in Figure 3B, the change trend of GA content is similar to that of IAA, and the overall change of IAA content shows a trend of first decreasing and then increasing. At 0 °C, the IAA content reaches the lowest level, and at 0 °C Y1 and L15 content are 94.14 pmol·g⁻¹ and 87.69 pmol·g⁻¹, respectively, with significant differences between them.

As shown in Figure 3C, the ABA content showed a trend of first increasing and then decreasing in the five periods, and it reached the highest level at 0 °C. At 2 °C, 0 °C, and −4 °C, the ABA content of L15 was always higher than that of Y1, but the difference was not significant.

3.4. Changes in MDA Content in Floral Organs

The accumulation of MDA will cause further damage to the cell membrane, so its content can reflect the degree of aging and stress injury of plants. In this experiment, MDA content showed a trend of first increasing and then decreasing, and it reached the highest level at 0 °C, indicating that the degree of lipid peroxidation increased (Figure 4). At 4 °C and 2 °C, the MDA content of Y1 was 282.09 and 333.77 nmol·g⁻¹, respectively, which was significantly different from that of L15. At 2 °C, Y1 and L15 contents were 404.83 nmol·g⁻¹ and 425.82 nmol·g⁻¹, respectively, with significant differences.

3.5. The Contents of Antioxidant Enzymes (POD, SOD, CAT) in Floral Organs Were Changed

In Figure 5A, the POD content shows a trend of decreasing first and then increasing. At 4 °C, the Y1 content was significantly higher than L15, and at 2 °C the Y1 content decreased, and the difference was significant. At this time, the POD content of L15 was 1.13 times that of Y1. At 2 °C, 0 °C, and −2 °C, the L15 content is higher than that of Y1.

In Figure 5B, the SOD content exhibited a trend of initially increasing, subsequently decreasing, and then increasing again. At 4 °C, the SOD content in Y1 was notably lower than that in L15, with a highly significant difference between the two. Following the temperature decrease to 2 °C, 0 °C, −2 °C, and −4 °C, the SOD content in Y1 was consistently and significantly higher than that in L15, being 1.51, 1.22, 1.22, and 1.44 times higher, respectively.

CAT catalyzes the decomposition of hydrogen peroxide into oxygen and water, serving as a crucial antioxidant enzyme. As illustrated in Figure 5C, the CAT content in Y1 consistently decreases with decreasing temperature, whereas the CAT content in L15 initially increases, then decreases, and subsequently increases again as the temperature drops. At 4 °C, the CAT content in Y1 was significantly higher than that in L15, with a difference of 82.12 μmol·min⁻¹·g⁻¹. Conversely, at −4 °C, the CAT content in L15 was notably higher than that in Y1, exceeding it by 375.24 μmol·min⁻¹·g⁻¹, which is 1.25 times greater than the CAT content in Y1.

3.6. Changes in the Content of Osmoregulatory Substances (Pro, Soluble Sugars) in Floral Organs

With the decrease in temperature, the Pro content of Y1 and L15 peach blossoms showed a trend of first decreasing, then increasing, and then decreasing (Figure 6A). When the temperature was −2 °C, the Pro content of both varieties reached the highest value, and the Pro content was significantly different from that at 4 °C. At 2 °C, the Pro content of L15 was 259.31 μg·g⁻¹, which was significantly higher than that of Y1 (229.22 μg·g⁻¹).

The soluble sugar content of Y1 and L15 varieties decreased first and then increased (Figure 6B). The soluble sugar content of Y1 at 4 °C was 10.18%, which was significantly different from that at 2 °C, 0 °C, −2 °C, and −4 °C (7.26%, 7.13%, 7.72%, and 8.64%). The soluble sugar content of L15 at 4 °C was 8.96%, which was significantly different from that at 2 °C, 0 °C, and −2 °C (8.28%, 7.67%, and 8.15%).

3.7. Correlation Analysis of Resistance of Peach Varieties to Late Frost Damage

According to Figure 7A, a correlation analysis was conducted on the IAA, GA, ABA, MDA, POD, SOD, CAT, Pro, and SS of the Y1 variety under five temperature conditions, and it was found that MDA was significantly positively correlated with ABA and IAA was significantly positively correlated with GA content. Notably, the correlation coefficient between MDA and GA was −0.99, indicating a highly significant negative correlation; furthermore, the correlation coefficient between MDA and IAA was −1, reflecting a completely negative relationship. Conversely, ABA content demonstrated a significant negative correlation with both GA and IAA. In contrast to the Y1 variety’s findings in Figure 7A, the analysis for L15 under five temperature conditions (Figure 7B) revealed that ABA had a significantly negative correlation with SS as well as with both GA and IAA. That is to say, with the change in ABA content, the content of SS, GA, and IAA showed the opposite trend; SS is significantly positively correlated with GA, with SS being completely positively correlated with IAA and GA being significantly positively correlated with IAA. Upon comparing Y1 with L15, the changes in ABA and MDA content in Y1 showed a positive trend. The changes in ABA content and SS content in L15 showed a reverse trend.

4. Discussion

The effects of low temperatures on trees and their internal mechanisms are complex, involving various aspects, such as physiological responses, organizational structure, and biochemistry [20]. When plants are exposed to low temperatures, a key part of their response focuses on the cell membrane. The cell membrane is composed of a phospholipid bilayer and a variety of proteins, which together maintain the structural integrity and functional activity of the cell membrane, and the fluidity of the membrane is the basis for its normal operation. However, a decrease in temperature weakens membrane fluidity, promoting ice crystal formation in cells and slowing biochemical reactions, thereby disrupting substance transport and signal transmission between cells [21]. Studies have shown that late spring frost, a short period of freezing damage, causes water in plant cells to freeze, damaging cell structures and disrupting normal growth and development [22]. It also damages flowers and young fruits, leading to fruit drop and reduced yield [23]. Various preventive measures have been implemented to mitigate the impact of late spring frost, including the use of cold-resistant varieties, late-maturing varieties, mulch, and fans to enhance air circulation [24]. Cold-resistant varieties can continue growing after temperatures rise, compensating for frost-induced growth stagnation and reducing physiological damage. Thus, they are one of the most effective ways to mitigate late frost damage [25].

The supercooling point is closely linked to a plant’s cold resistance [26,27]. When the temperature drops below 0 °C, the water in plant tissues and organs remains liquid, entering a supercooled state, instead of freezing immediately [28]. As the supercooling point temperature decreases, plants freeze more quickly at lower temperatures, indicating stronger cold resistance [29]. Y1 and L15, representing two distinct local peach varieties, exhibited anisotropy under low-temperature stress. Trend analysis revealed that the peak supercooling point of Y1 occurred before L15, with its freezing point around −3.7 °C, while Y1’s supercooling point was higher, at about −4.6 °C. The freezing point of L15 is approximately −4.8 °C, while its supercooling point can reach as low as −7.2 °C. The low supercooling point of L15 enables it to retain liquid water effectively under short-term, low-temperature stress, preventing cell water from freezing and reducing the risk of cell damage. This means that Y1 is more prone to freezing at low temperatures, leading to cell damage and growth arrest. Therefore, the difference in supercooling mechanisms between Y1 and L15 reflects their respective abilities to adapt to low-temperature environments.

The cold resistance index is a key indicator for assessing a plant’s ability to withstand low-temperature stress [30]. The cold resistance index of peaches helps to further investigate the mechanisms of low-temperature stress damage, as well as the plant’s ability to tolerate and respond to such stress, thereby mitigating the effects of late frost damage. Late frost damage typically occurs during the flowering and fruit setting stages of peaches; during this stage, changes in the types and levels of hormones in the floral organs interact to enhance survival rates [31]. Studies have shown that both nectarine and common wild peach flower organs exhibit changes in hormone content in response to low-temperature stress, with increased ABA levels being a shared characteristic [32]. Optimal levels of IAA contribute to flower formation and development, enhancing resistance to low temperatures [33]. Liu et al. [34] studied hormone and polyamine content in low-temperature grape branches and found that IAA content negatively correlates with cold resistance, suggesting that lower IAA levels impair IAA catabolic enzyme activity, reduce physiological processes like cell elongation, growth, and vascular bundle differentiation, and affect pollen mother cell division. This enhances the cold resistance of floral organs [35]. In this study, IAA content reached its lowest at 0 °C before increasing again. This was due to a decrease in cell metabolic rate following the physiological stress response, which hindered IAA synthesis and transport, leading to a reduction in its content; once the adaptive response was activated, IAA biosynthesis resumed, leading to an increase in its content. GA content promotes plant cell division and expansion, which is associated with the flowering transition [36]. In this study, the variation trend of GA content is the same as that of IAA. Y1’s GA content reached its lowest at 0 °C and was significantly higher than that of L15, indicating that L15 is more responsive to environmental changes and adjusts its internal mechanisms by rapidly decreasing GA content. ABA, a key stress hormone, plays a crucial role in plant responses to stress, such as low temperatures [37]. Increased ABA content enhances plant cold resistance and helps maintain physiological activity in floral organs under low-temperature conditions [38]. In this study, ABA content accumulated early in the simulated late frost damage phase, which was closely linked to the cold resistance of peaches and consistent with previous studies [39]. However, hormonal regulation is complex and varied, with hormones interacting to maintain balance.

MDA, a marker of oxidative stress, plays a role in the antioxidant response and cellular protection mechanisms in plants [40]. Yang et al. [41] investigated the effects of low-temperature stress on membrane lipid peroxidation products and enzyme activities in oil peach blossom organs. Wang et al. [42] demonstrated that the increased levels of lipid peroxidation products in common peach blossom organs reflect membrane damage under low-temperature stress, along with the plant’s adaptive and resistance mechanisms. In this study, the MDA content in the simulated early stage of late frost (4 °C, 2 °C, 0 °C) showed an increasing trend, and the MDA content in Y1 was significantly lower than that in L15, indicating that as the critical condition of late frost freezing damage was approaching, the plant enzyme system was limited and exceeded its self-regulation ability, which eventually led to the increase in MDA content, further indicating that L15 was sensitive to environmental response. After the critical condition of late frost, the plant’s damage was also reduced by reducing the MDA content. Antioxidant enzymes play an important role in plant protection by clearing ROS, maintaining REDOX balance, and promoting cell repair. POD can remove excess free radicals in the body, prevent the synthesis of the hydroxyl group (OH⁻), promote cell wall formation, and improve plant stress resistance [43]. In this study, POD content initially decreased before increasing. At temperatures of −2 °C and −4 °C, the trend of POD content mirrored that observed by Yu Dan et al. [44] in their study of pistil growth and development in apricot varieties, showing a gradual increase. SOD catalyzes the conversion of superoxide anions (O₂⁻) into hydrogen peroxide (H₂O₂) and oxygen (O₂), serving as the first line of antioxidant defense. Increased SOD activity effectively reduces intracellular O₂⁻ accumulation, mitigating oxidative damage [45]. CAT catalyzes the breakdown of H₂O₂ into water and oxygen, further lowering intracellular H₂O₂ levels. In this study, SOD content in Y1 was consistently higher than in L15 at 2 °C, 0 °C, −2 °C, and −4 °C, whereas the opposite was true for CAT content, with L15 exhibiting higher levels than Y1 at these temperatures. This suggests a difference in physiological adaptability between the two varieties, with L15’s higher CAT levels potentially exacerbating its sensitivity to environmental stress. In addition, studies have shown that the thawing process that follows late frost damage is often a critical stage in exacerbating oxidative damage. During thawing, enzyme activity (such as respiratory enzymes and oxidases) is rapidly activated, but, at this time, the electron transport chain has not fully recovered, promoting the generation of ROS, such as superoxide anions (O₂⁻) and hydrogen peroxide (H₂O₂), further damaging cells. This study did not cover the dynamic changes during lower temperatures or thawing processes, which to some extent limits the comprehensive understanding of the oxidative damage mechanisms throughout the entire process of late frost damage [46].

Under low-temperature stress caused by late frost damage, plants activate self-protection mechanisms by actively accumulating various organic and inorganic substances. The accumulation of these substances increases cell fluid concentration, lowering the osmotic potential and enabling cells to better retain water under low-temperature-induced water loss, thus preventing damage from excessive dehydration [47]. Proline (Pro) and soluble sugars (SS) are osmotic regulators that maintain osmotic balance between the protoplasm and the environment during thawing and freezing processes [48]. The two peach varieties differ in genetic background and physiological response mechanisms to environmental stress, such as temperature fluctuations, and the content of osmotic regulators also varies between the two varieties. In this study, Pro content showed minimal variation, but L15 consistently exhibited higher Pro levels than Y1, with a significant difference observed at 2 °C. The lowest SS content occurred at 0 °C, with L15 consistently showing higher SS levels than Y1 at temperatures above 2 °C. Therefore, increased Pro and SS levels indicate enhanced plant adaptability to low-temperature stress, potentially improving survival rates.

After flowering, plant growth priorities gradually shift towards the development and maturation of flower organs [49]. Under cold conditions at this stage, ABA synthesis is typically negatively correlated with auxin and gibberellin synthesis [50]. Correlation analysis revealed that ABA content in Y1 and L15 was negatively correlated with GA and IAA content across five temperature stages. These findings are consistent with previous studies. Studies have shown that under adverse conditions (e.g., low temperature, salt, and alkali), high ABA levels inhibit cell expansion and differentiation, reduce the biological activity of growth hormones, and, consequently, slow plant growth. This mechanism serves as an adaptive response, prioritizing protection and survival under environmental stress [51]. In this study, ABA content increased at 4 °C, 2 °C, and 0 °C, reflecting an adaptive response to short-term frost damage. At 0 °C, Y1 exhibited significantly higher GA content than L15, and, at −4 °C, Y1 had significantly higher IAA content than L15. This suggests that L15 may exhibit greater activity in the ABA signaling pathway, making its response to ABA more pronounced after flowering, thereby inhibiting the synthesis of IAA and GA. In addition to its relationship with growth hormones, the negative correlation between ABA content and antioxidant enzymes (POD, SOD, CAT) as well as biomolecules (Pro and SS) is noteworthy. It was found that ABA content was negatively correlated with POD, SOD, CAT, Pro, and SS content, with Y1 showing a higher negative correlation coefficient than L15. This phenomenon may be linked to ABA’s role in regulating the physiological state and metabolic pathways of plants. High ABA concentrations may inhibit the synthesis of antioxidant enzymes, exacerbating oxidative damage in plants under stress [52]. This negative correlation suggests that while ABA may enhance short-term plant resistance to low temperatures (e.g., 0 °C), its sustained high levels could inhibit the antioxidant system, affecting long-term plant viability. At the same time, the negative correlation between ABA and SS/Pro suggests that high ABA levels may inhibit the synthesis of these protective substances, either by reducing SS and Pro synthesis through gene expression inhibition or by enhancing respiration, leading to the consumption of SS for stress resistance. This process makes SS more available for energy production and metabolism, resulting in a temporary reduction in SS content [53]. However, the exact mechanisms require further verification.

In the future, our research will incorporate more years of data validation and be combined with practical applications, focusing more on monitoring temperature gradients and thawing stages to more accurately analyze the comprehensive effects of slow cooling and thawing on oxidative damage to flower organ cells. Furthermore, by observing the microstructure of ice crystal formation, the relationship between ice crystal size and distribution and the degree of cell damage can be further elucidated.

5. Conclusions

This study compared the changes in the cross-section of floral organs and physiological indicators of Y1 and L15 varieties under various simulated low-temperature conditions, highlighting the impact of late frost on floral organ development. The results showed that the supercooling point of L15 was lower than that of Y1, indicating greater frost tolerance. Further analysis revealed that at the same temperature, the surface structure of the floral organ in Y1 was affected by frost damage earlier than in L15. At −2 °C, the anthers turned brown, the ovaries froze, and water stains appeared on the sepals and the center. The frost damage in L15 was evident only at −4 °C. The L15 variety exhibited the strongest resistance to late frost. The results of this study are helpful for us to better understand how different types of peach blossoms cope with environmental pressure and provide a reference for the study and cultivation of peaches under the same longitude, latitude, and climatic conditions in the northwest region.