Extreme temperature fluctuations from climate change negatively affect the growth of medicinal and economic plants. In arid regions, these changes disrupt biochemical processes necessary for metabolic activities, impacting osmotic balance and protein levels, which influence thermostability. This study investigates the water output and metabolic responses of broad leaf plants in Arabian deserts.
Samples of broad succulent and non-succulent leaves from some native species were collected from different areas between latitudes N 25° and N 30° in the Egyptian deserts to assess their responses in water output and primary metabolites to temperature variations under controlled conditions.
Results indicate that increased temperatures above 45°C lead to excessive water output and metabolic changes in plants. Notably, Hyoscyamus muticus, along with Calotropis procera, showcases a capacity to endure extreme temperatures, contrasting with Cynanchum acutum, which shows poor resilience to heat stress. Investigations reveal significant correlations among parameters in plant species at latitude 27°, particularly in the Assiut region, influenced by extreme climatic conditions.
In agricultural projects, the succulent leaf characteristics of introduced species and native plants play a crucial role in managing climate changes by absorbing high insolation, even under extreme temperature conditions.
Introduction
In hot dryland ecosystems, plants face atmospheric aridity, high insolation, and extreme temperatures. Most desert species possess small and modified leaves as adaptations to these conditions. Leaf thermal resilience depends on the ability of leaves to regulate temperature, which differs among species and environments (Fauset et al., 2018). Climate fluctuations further increase leaf susceptibility to temperature extremes (Still et al., 2021). Evapotranspiration plays a key role in cooling leaves and shows greater plasticity than physical leaf traits (Uni et al., 2022), making the mitigation of physical warming essential for plant survival in extreme heat.
Evapotranspiration cooling regulates energy flow and leaf temperature (Kibler et al., 2023), while high leaf temperatures can cause biochemical stress (Vinod et al., 2022). Plants enhance thermostability through the accumulation of water-binding molecules and compatible solutes (Farghali & El-Aidarous, 2014), which increase cytoplasmic viscosity and osmolality (Rayan & Farghali, 2007). Heat tolerance in xerophytes is also supported by heat shock proteins (Hasanuzzaman, Nahar, Alam, Roychowdhury, & Fujita, 2013). Reduced osmotic potential promotes the accumulation of soluble sugars, free amino acids, and soluble proteins, improving tolerance to heat and drought stress (Jaleel et al., 2008). Investigating these soluble metabolic mechanisms is crucial for improving heat resistance in economic crops in hot desert regions.
This study hypothesizes that (1) metabolic compounds accumulate under heat stress, (2) species differ in physiological responses due to inherent adaptive strategies, and (3) environmental variations among locations influence plant thermal responses. It investigates changes in soluble sugars, free amino acids, and total soluble proteins in selected broad leaf desert plants, considering leaf succulence as a thermal avoidance trait and measuring water output under simulated temperatures. Key morphological traits, including plant height, leaf area, and leaf mass per area, are also analyzed to understand species-specific strategies for heat adaptation, providing insights into the physiological mechanisms underlying plant tolerance to extreme environments.
Study areas
Broad leaf species inhabiting three Egyptian desert regions in addition to one site in the Nile Valley, located between latitudes 25°–30° N, were investigated. The studied locations included:
Kharga Oasis (Kh. O.), located at N 25° E 30° in the Western Desert (Salman, Howari, El-Sankary, Wali, & Saleh, 2010), is characterized by a tropical arid climate. It experiences extreme temperature fluctuations, with daytime highs reaching 45–50°C in summer, while winter temperatures can drop to zero at night. Recognized as the driest area in the Eastern Sahara and possibly the driest region on Earth (Kehl & Bronkamm, 1993), Kharga Oasis has an annual mean relative humidity of approximately 39%, with very scarce atmospheric precipitation, such as rainfall.
Assiut Nile Valley (Ast. N.) is bounded by Eocene limestone plateaus to the east and west and lies between latitudes 26°50′–27°40′ N and longitudes 30°40′–31°32′ E. This area forms part of the Nile Valley within Egypt's stable shelf, where surface features reflect basement structures and major fault systems trending NW, NE, and N–S (El Shemi, Setto, Mauritish, & Abu Helelia, 1999).
Assiut Eastern Desert (Ast. D.) is located in a transitional region between the Arabian and El-Galala deserts, extending between latitudes 27°10′–27°25′ N and longitudes 31°15′–31°50′ E. Elevation ranges from approximately 350 m in the eastern upstream region to 70 m above sea level in the western downstream deltaic part (Farghali, 1998).
Cairo Eastern Desert (Cairo D.) is situated between latitudes 30°05′04″–30°35′09″ N and longitudes 31°14′30″–32°15′49″ E (Azer, 2018). In the Eastern Desert, the natural vegetation occurs mainly along wadi channels and mountain slopes receiving limited rainfall, with a mean annual precipitation of up to 60 mm (Zahran & El-Amier, 2014).
The average weather data for Kh. O. indicates a maximum temperature of 37.4°C, a minimum of 13.6°C, irradiance of 22 MJ/m2/day, relative humidity of 30.6%, wind speed of 4.9 m/s, and no precipitation. For Ast. N., the averages are a maximum temperature of 35.4°C, a minimum of 11.5°C, irradiance of 21.5 MJ/m2/day, relative humidity of 40.0%, wind speed of 4.3 m/s, and light precipitation of 0.02 mm/day. In Ast. D., the average maximum temperature is 37.5°C, with a minimum of 12.5°C, irradiance of 22.9 MJ/m2/day, relative humidity of 36.2%, wind speed of 4.3 m/s, and 0.02 mm/day precipitation. Lastly, Cairo D. reports a maximum temperature of 34.3°C, a minimum of 10.9°C, irradiance of 21.6 MJ/m2/day, relative humidity of 55.6%, wind speed of 3.6 m/s, and 0.5 mm/day precipitation, according to Link to the website, (2023–2025).
Materials and methods
The present study was conducted on three wild medicinal plant species inhabiting four locations representing different ecological conditions in the Egyptian desert and Nile Valley (Figure 1). The investigated locations were:
A map of Egypt showing the locations of studied areas in the Western and Eastern Deserts, as well as the Nile Valley and Cairo Eastern Desert. The map includes international boundaries, roads, and specific study areas marked with dashed lines. Key locations such as Siwa, Bahariya Oasis, Farafra Oasis, Dakhla Oasis, Kharga Oasis, Assiut, Cairo, Suez, and Aswan are labeled. The map also highlights the Nile River, the Red Sea, and the Mediterranean Sea. The study areas are numbered and include specific regions in the Western Desert, Assiut Nile Valley, Assiut Eastern Desert, and Cairo Eastern Desert. The map provides a detailed view of the geographic layout and key points of interest in these regions.Locations of studied areas at Kharga Oasis, Assiut Nile Valley, Assiut Eastern Desert, and Cairo Eastern Desert between latitudes 25° and 30° N (Salman et al., 2010)
A map of Egypt showing the locations of studied areas in the Western and Eastern Deserts, as well as the Nile Valley and Cairo Eastern Desert. The map includes international boundaries, roads, and specific study areas marked with dashed lines. Key locations such as Siwa, Bahariya Oasis, Farafra Oasis, Dakhla Oasis, Kharga Oasis, Assiut, Cairo, Suez, and Aswan are labeled. The map also highlights the Nile River, the Red Sea, and the Mediterranean Sea. The study areas are numbered and include specific regions in the Western Desert, Assiut Nile Valley, Assiut Eastern Desert, and Cairo Eastern Desert. The map provides a detailed view of the geographic layout and key points of interest in these regions.Locations of studied areas at Kharga Oasis, Assiut Nile Valley, Assiut Eastern Desert, and Cairo Eastern Desert between latitudes 25° and 30° N (Salman et al., 2010)
Kharga Oasis in the Western Desert (25° N, 30° E; Kh. O.).
Assiut Nile Valley (27° N, 30° E; Ast. N.).
Assiut Eastern Desert (27° N, 31° E; Ast. D.).
Cairo Eastern Desert (30° N, 32° E; Cairo D.).
The studied species included Calotropis procera (Aiton) W.T. Aiton, Cynanchum acutum L. (Family: Apocynaceae), and Hyoscyamus muticus L. (Family: Solanaceae). Plant samples were collected during summer (2023–2025) (Plate 1). Species identification was carried out according to Täckholm (1974), El-Hadidi and Fayed (1995), and Boulos (2000). Leafy branches were collected from healthy mature plants in the field during the morning hours. For each species, five branches were collected within each study location. The samples were immediately placed in tightly sealed plastic containers to minimize water loss and transported to the laboratory in insulated ice boxes. All samples were processed within a short time after collection to minimize physiological changes before the experiment.
The image contains three sets of photos, each set showing a different species of plants collected from four distinct locations. The first set, labeled I, shows Calotropis procera from Kharga Oasis, Assiut Nile Valley, Assiut Eastern Desert, and Cairo Eastern Desert. The second set, labeled II, displays Hyoscyamus muticus from the same locations. The third set, labeled III, features Cynanchum acutum also from the same locations. Each set includes four photos, one from each location, showcasing the plants in their natural habitats.The studied species (I. Calotropis procera, II. Hyoscyamus muticus, and III. Cynanchum acutum) were collected from different locations: (A) Kharga Oasis, (B) Assiut Nile Valley, (C) Assiut Eastern Desert, and (D) Cairo Eastern Desert. The species were sampled during summer 2023–2025
The image contains three sets of photos, each set showing a different species of plants collected from four distinct locations. The first set, labeled I, shows Calotropis procera from Kharga Oasis, Assiut Nile Valley, Assiut Eastern Desert, and Cairo Eastern Desert. The second set, labeled II, displays Hyoscyamus muticus from the same locations. The third set, labeled III, features Cynanchum acutum also from the same locations. Each set includes four photos, one from each location, showcasing the plants in their natural habitats.The studied species (I. Calotropis procera, II. Hyoscyamus muticus, and III. Cynanchum acutum) were collected from different locations: (A) Kharga Oasis, (B) Assiut Nile Valley, (C) Assiut Eastern Desert, and (D) Cairo Eastern Desert. The species were sampled during summer 2023–2025
Determination of morphological traits
Plant height (cm) was measured from the base to the apex of five plants of each species at each location using a ruler. Leaf area (cm2) was determined from five fully expanded mature leaves of each species at each location by tracing the leaf outline on paper of known weight per unit area, cutting it out, weighing it, and calculating the area based on the paper mass per unit area. Leaf mass per area (mg/cm2) was calculated for the same five leaves as the ratio of leaf dry mass to leaf area. Mean values were then calculated for all morphological traits. All measurements were performed following standardized protocols (Cornelissen et al., 2003).
Determination of leaf succulence
Leaf succulence was calculated according to Delf's index (Delf, 1912). Fresh leaf weight was measured, and leaf area was estimated using the weighing method. Leaf succulence was expressed as mg/cm2 according to the following equation:
Preparation of samples
Healthy, fully expanded leaves were carefully detached from the collected branches. The leaves were cleaned from dust and debris and gently blotted with tissue paper. Three replicates were used for each species, location, and temperature treatment.
Leaves were cut at the petiole, and the cut end was immediately sealed with a thin layer of melted paraffin wax to prevent latex exudation and water loss from the cutting surface. Each leaf was weighed to determine its fresh weight. The leaves were then incubated in controlled-temperature chambers at constant temperatures of 25°C, 35°C, 45°C, and 55°C for one hour to simulate the high temperature conditions recorded in the studied habitats. After incubation, the leaves were reweighed to determine water loss. Water output was calculated using the following equation:
This parameter represents the relative water loss from leaves under temperature stress.
Analysis of plant extracts
Immediately after the temperature treatments, leaves were rapidly rinsed with distilled water and gently blotted dry using filter paper. To prevent further metabolic activity, all extraction procedures were performed quickly under cold conditions. Leaf samples from three replicates of each treatment were used for extract preparation. A known fresh weight of leaf tissue from each replicate was homogenized in 10 cm3 of ice-cold distilled water using a pre-chilled mortar and pestle. The homogenate was then centrifuged at 7,000 rpm for 10 minutes. The resulting supernatant was carefully collected and stored at −20°C until subsequent biochemical analyses.
Determination of water-soluble metabolites
Total soluble sugars (SS) were measured colorimetrically using the phenol–sulfuric acid method (Dubois, Gilles, Hamilton, Rabers, & Smith, 1956), where sugars react with phenol and concentrated sulfuric acid to form a colored complex, with absorbance read at 490 nm.
Free amino acids (AA) were quantified via the ninhydrin reaction (Lee & Takahashi, 1966), producing a purple complex measured at 570 nm.
Total soluble proteins (SP) were determined using the Lowry method (Lowry, Rosebrough, Farr, & Randall, 1951), in which proteins react with Folin–Ciocalteu reagent to form a blue complex, measured at 750 nm.
Statistical analysis
The study utilized factorial analysis (two-way method) of variance (F test) to evaluate the effects of single factors (location, temperature, or species) on leaf succulence, water output, and metabolites across different species. The relative influence of each factor and their interactions on the overall response was assessed using the coefficient of determination (share %) (Ostle, 1963; Ploxinski, 1969). A simple linear correlation coefficient (r) between all parameters for each plant species separately and the correlation between water output, leaf succulence and metabolites were calculated by using SPSS program (2020).
Results
Morphological traits
C. acutum recorded the highest plant height (64.95 cm), whereas H. muticus showed the lowest value (37.44 cm). Leaf area was largest in C. procera (82.71 cm2) and smallest in C. acutum (12.89 cm2). Leaf mass per area was highest in C. acutum (6.18 mg/cm2) and lowest in H. muticus (3.24 mg/cm2). These clear variations in morphological traits, particularly leaf area and leaf mass per area, likely contribute to their differential responses to heat stress (Table 1).
Leaf succulence
In the leaf succulence study across different locations, H. muticus showed the highest value at 98.8 mg/cm2 (Ast. D.), followed by C. procera at 74.0 mg/cm2 (Kh. O.) and C. acutum at 44.3 mg/cm2 (Cairo D.) (Figure 2). At Kh. O., H. muticus decreased to 77.8 mg/cm2, while C. procera and C. acutum declined to 50.5 and 16.9 mg/cm2, respectively (Ast. N.). H. muticus exhibited superior water storage, reducing evapotranspiration, similar to C. procera, whereas C. acutum showed low succulence with high water output. Species, location, and their interaction significantly affected leaf succulence, with species being the dominant factor (Table 2).
Water output (as % of leaf fresh weight)
Water output was strongly influenced by temperature across all species (Figure 3). Elevated temperatures (45–55°C) caused substantial water loss, with the highest observed in C. acutum (75.3%, Ast. N.), while temperatures below 45°C minimized water output, with the lowest values in C. procera (4.6%, Cairo D.) and H. muticus (4.6%, Kh. O.). Plants from Ast. D. generally showed higher water output than those from other locations. Regardless of temperature, low water loss also varied among species depending on location, with reduced values in H. muticus and C. acutum at Cairo D. and in C. procera at Kh. O. Statistical analysis indicated that temperature was the main factor increasing water output, location had a secondary effect in C. procera and C. acutum, and the temperature × location interaction had an equivalent influence in H. muticus (Table 3).
Metabolic constituents
Total soluble sugars (SS)
The investigation revealed that total soluble sugars (SS) are influenced by temperature and location, with a notable increase at 55°C (Figure 4). C. acutum had the highest SS content (38.4 mg/g leaf fresh weight [leaf f. wt.], Ast. N.), while H. muticus had the lowest (1.2 mg/g leaf f. wt.). Between 25°C and 45°C, SS variation was minimal, with C. acutum at Cairo D. showing the highest values. The study indicated that temperature significantly affects SS in C. procera and C. acutum, while location primarily influences H. muticus, playing a secondary role for C. acutum, and its interaction with temperature had the same role in the case of the rest of the species (Table 4).
Free amino acids (AA)
C. acutum had the highest free amino acid content (55.2 mg/g leaf f. wt.), while H. muticus had the lowest (7.3 mg/g leaf f. wt.) (Figure 5). Free amino acids increased with temperature, especially at Ast. D, where C. procera and H. muticus reached maximum levels of 13.4 and 7.3 mg/g leaf f. wt., respectively, at 55°C. ANOVA showed that temperature, location, and their interaction significantly affected amino acid content: temperature dominated in C. procera, location in C. acutum, and the interaction effect in H. muticus (Table 5).
Total soluble proteins (SP)
Morphological traits of the studied species expressed as mean ± standard deviation (SD)
| Species | Plant height (cm) | Leaf area (cm2) | Leaf mass per area (mg/cm2) |
|---|---|---|---|
| C. procera | 42.77 ± 0.03 | 82.71 ± 0.07 | 5.23 ± 0.01 |
| H. muticus | 37.44 ± 0.19 | 34.90 ± 0.02 | 3.24 ± 0.01 |
| C. acutum | 64.95 ± 0.08 | 12.89 ± 0.01 | 6.18 ± 0.01 |
A box-and-whisker plot compares leaf succulence in three plant species: Calotropis procera, Hyoscyamus muticus, and Cynanchum acutum across four locations in Egypt: Kharga Oasis, Assiut Nile Valley, Assiut Eastern Desert, and Cairo Eastern Desert. The x-axis represents the locations, and the y-axis represents leaf succulence in milligrams per cubic centimeter, ranging from 0 to 120. The plot includes three vertical box plots for each location, color-coded in blue for Calotropis procera, red for Hyoscyamus muticus, and gray for Cynanchum acutum. Each box represents the interquartile range (IQR), with the horizontal line within each box indicating the median. Whiskers extend to the data range. For Kharga Oasis, Calotropis procera shows a median around 70, Hyoscyamus muticus around 80, and Cynanchum acutum around 15. In Assiut Nile Valley, Calotropis procera has a median around 50, Hyoscyamus muticus around 85, and Cynanchum acutum around 20.Leaf succulence (mg/cm2) in Calotropis procera, Hyoscyamus muticus, and Cynanchum acutum under four locations in Egypt (Kharga Oasis, Assiut Nile Valley, Assiut Eastern Desert, and Cairo Eastern Desert). The species were sampled during summer 2023–2025. Boxes represent the interquartile range (IQR), the horizontal line within each box indicates the median, and whiskers represent the data range
A box-and-whisker plot compares leaf succulence in three plant species: Calotropis procera, Hyoscyamus muticus, and Cynanchum acutum across four locations in Egypt: Kharga Oasis, Assiut Nile Valley, Assiut Eastern Desert, and Cairo Eastern Desert. The x-axis represents the locations, and the y-axis represents leaf succulence in milligrams per cubic centimeter, ranging from 0 to 120. The plot includes three vertical box plots for each location, color-coded in blue for Calotropis procera, red for Hyoscyamus muticus, and gray for Cynanchum acutum. Each box represents the interquartile range (IQR), with the horizontal line within each box indicating the median. Whiskers extend to the data range. For Kharga Oasis, Calotropis procera shows a median around 70, Hyoscyamus muticus around 80, and Cynanchum acutum around 15. In Assiut Nile Valley, Calotropis procera has a median around 50, Hyoscyamus muticus around 85, and Cynanchum acutum around 20.Leaf succulence (mg/cm2) in Calotropis procera, Hyoscyamus muticus, and Cynanchum acutum under four locations in Egypt (Kharga Oasis, Assiut Nile Valley, Assiut Eastern Desert, and Cairo Eastern Desert). The species were sampled during summer 2023–2025. Boxes represent the interquartile range (IQR), the horizontal line within each box indicates the median, and whiskers represent the data range
ANOVA test showed the effects of species (Sp.), location (L.) and their interaction on the leaf succulence of investigated species
| Source | df | F | Share % |
|---|---|---|---|
| Species | 2 | 226.82** | 87.12 |
| Location | 3 | 8.32** | 4.79 |
| Species × Location | 6 | 7.02** | 8.09 |
Note(s): **Significant at 0.01 level *Significant at 0.05 level
Three box plots depict water output as a percentage of leaf fresh weight for three plant species under different temperature treatments from four locations in Egypt. Panel A shows data for Calotropis procera, Panel B for Hyoscyamus muticus, and Panel C for Cynanchum acutum. Each panel includes temperature on the horizontal axis, ranging from 25 to 55 degrees Celsius, and water output on the vertical axis, ranging from 0 to 100 percent. The box plots represent data from four locations: Kharga Oasis, Assiut Nile Valley, Assiut Eastern Desert, and Cairo Eastern Desert, distinguished by different colors: blue, red, gray, and green respectively. Boxes represent the interquartile range (IQR), the horizontal line within each box indicates the median, and whiskers represent the data range. In Panel A, water output increases with temperature for all locations, with the highest output at 55 degrees Celsius.Water output (as % of leaf fresh weight) in the three studied species: (A) Calotropis procera, (B) Hyoscyamus muticus, and (C) Cynanchum acutum under four increasing temperature treatments from four locations in Egypt (Kharga Oasis, Assiut Nile Valley, Assiut Eastern Desert, and Cairo Eastern Desert). The species were sampled during summer 2023–2025. Boxes represent the interquartile range (IQR), the horizontal line within each box indicates the median, and whiskers represent the data range
Three box plots depict water output as a percentage of leaf fresh weight for three plant species under different temperature treatments from four locations in Egypt. Panel A shows data for Calotropis procera, Panel B for Hyoscyamus muticus, and Panel C for Cynanchum acutum. Each panel includes temperature on the horizontal axis, ranging from 25 to 55 degrees Celsius, and water output on the vertical axis, ranging from 0 to 100 percent. The box plots represent data from four locations: Kharga Oasis, Assiut Nile Valley, Assiut Eastern Desert, and Cairo Eastern Desert, distinguished by different colors: blue, red, gray, and green respectively. Boxes represent the interquartile range (IQR), the horizontal line within each box indicates the median, and whiskers represent the data range. In Panel A, water output increases with temperature for all locations, with the highest output at 55 degrees Celsius.Water output (as % of leaf fresh weight) in the three studied species: (A) Calotropis procera, (B) Hyoscyamus muticus, and (C) Cynanchum acutum under four increasing temperature treatments from four locations in Egypt (Kharga Oasis, Assiut Nile Valley, Assiut Eastern Desert, and Cairo Eastern Desert). The species were sampled during summer 2023–2025. Boxes represent the interquartile range (IQR), the horizontal line within each box indicates the median, and whiskers represent the data range
ANOVA test showed the effect of temperature (T.), location (L.) and their interaction on the water output of different species investigated
| Species | ||||||||
|---|---|---|---|---|---|---|---|---|
| Source | C. procera | H. muticus | C. acutum | |||||
| df | F | Share% | F | Share% | F | Share% | ||
| Water output % | T. | 3 | 466.17** | 88.61 | 193.31** | 70.04 | 126.22** | 67.68 |
| L. | 3 | 36.29** | 6.90 | 22.67** | 8.21 | 43.60** | 23.38 | |
| T. × L. | 9 | 7.88** | 4.50 | 20.01** | 21.75 | 5.56** | 8.95 | |
Note(s): **Significant at 0.01 level *Significant at 0.05 level
Three box plots depict total soluble sugar contents in milligrams per gram of leaf fresh weight for three plant species under different temperature treatments from four locations in Egypt. Panel A shows data for Calotropis procera, Panel B for Hyoscyamus muticus, and Panel C for Cynanchum acutum. Each panel includes temperature on the horizontal axis in degrees Celsius, ranging from 25 to 55 degrees Celsius, and total soluble sugars on the vertical axis in milligrams per gram of leaf fresh weight, ranging from 0 to 50. The box plots represent data from four locations: Kharga Oasis, Assiut Nile Valley, Assiut Eastern Desert, and Cairo Eastern Desert, indicated by different colors. Boxes represent the interquartile range, the horizontal line within each box indicates the median, and whiskers represent the data range. In Panel A, total soluble sugars generally increase with temperature, with notable variations among locations.Total soluble sugar contents (mg/g leaf fresh weight) in the three studied species: (A) Calotropis procera, (B) Hyoscyamus muticus, and (C) Cynanchum acutum under four increasing temperature treatments from four locations in Egypt (Kharga Oasis, Assiut Nile Valley, Assiut Eastern Desert, and Cairo Eastern Desert). The species were sampled during summer 2023–2025. Boxes represent the interquartile range (IQR), the horizontal line within each box indicates the median, and whiskers represent the data range
Three box plots depict total soluble sugar contents in milligrams per gram of leaf fresh weight for three plant species under different temperature treatments from four locations in Egypt. Panel A shows data for Calotropis procera, Panel B for Hyoscyamus muticus, and Panel C for Cynanchum acutum. Each panel includes temperature on the horizontal axis in degrees Celsius, ranging from 25 to 55 degrees Celsius, and total soluble sugars on the vertical axis in milligrams per gram of leaf fresh weight, ranging from 0 to 50. The box plots represent data from four locations: Kharga Oasis, Assiut Nile Valley, Assiut Eastern Desert, and Cairo Eastern Desert, indicated by different colors. Boxes represent the interquartile range, the horizontal line within each box indicates the median, and whiskers represent the data range. In Panel A, total soluble sugars generally increase with temperature, with notable variations among locations.Total soluble sugar contents (mg/g leaf fresh weight) in the three studied species: (A) Calotropis procera, (B) Hyoscyamus muticus, and (C) Cynanchum acutum under four increasing temperature treatments from four locations in Egypt (Kharga Oasis, Assiut Nile Valley, Assiut Eastern Desert, and Cairo Eastern Desert). The species were sampled during summer 2023–2025. Boxes represent the interquartile range (IQR), the horizontal line within each box indicates the median, and whiskers represent the data range
ANOVA test showed the effects of temperature (T.), location (L.) and their interaction on total soluble sugars of different investigated species
| Species | ||||||||
|---|---|---|---|---|---|---|---|---|
| Source | C. procera | H. muticus | C. acutum | |||||
| df | F | Share% | F | Share% | F | Share% | ||
| Total Soluble sugars | T. | 3 | 13.65** | 51.18 | 14.13** | 13.73 | 10.09** | 38.49 |
| L. | 3 | 3.78* | 14.17 | 69.50** | 67.51 | 8.49** | 32.39 | |
| T. × L. | 9 | 3.08** | 34.66 | 6.44** | 18.76 | 2.55* | 29.12 | |
Note(s): **Significant at 0.01 level *Significant at 0.05 level
The image contains three box plots labeled A, B, and C, each representing different plant species: Calotropis procera, Hyoscyamus muticus, and Cynanchum acutum, respectively. The x-axis of each plot represents temperature in degrees Celsius, with values at 25, 35, 45, and 55 degrees. The y-axis represents free amino acid content in milligrams per gram of leaf fresh weight, ranging from 0 to 80. Each box plot includes data from four locations in Egypt: Kharga Oasis, Assiut Nile Valley, Assiut Eastern Desert, and Cairo Eastern Desert, represented by different colors: blue, red, gray, and green, respectively. Boxes represent the interquartile range (IQR), the horizontal line within each box indicates the median, and whiskers represent the data range. For Calotropis procera, the amino acid content remains relatively low across all temperatures and locations, with slight increases at higher temperatures. All values are approximated.Free amino acid contents (mg/g leaf fresh weight) in the three studied species: (A) Calotropis procera, (B) Hyoscyamus muticus, and (C) Cynanchum acutum under four increasing temperature treatments from four locations in Egypt (Kharga Oasis, Assiut Nile Valley, Assiut Eastern Desert, and Cairo Eastern Desert). The species were sampled during summer 2023–2025. Boxes represent the interquartile range (IQR), the horizontal line within each box indicates the median, and whiskers represent the data range
The image contains three box plots labeled A, B, and C, each representing different plant species: Calotropis procera, Hyoscyamus muticus, and Cynanchum acutum, respectively. The x-axis of each plot represents temperature in degrees Celsius, with values at 25, 35, 45, and 55 degrees. The y-axis represents free amino acid content in milligrams per gram of leaf fresh weight, ranging from 0 to 80. Each box plot includes data from four locations in Egypt: Kharga Oasis, Assiut Nile Valley, Assiut Eastern Desert, and Cairo Eastern Desert, represented by different colors: blue, red, gray, and green, respectively. Boxes represent the interquartile range (IQR), the horizontal line within each box indicates the median, and whiskers represent the data range. For Calotropis procera, the amino acid content remains relatively low across all temperatures and locations, with slight increases at higher temperatures. All values are approximated.Free amino acid contents (mg/g leaf fresh weight) in the three studied species: (A) Calotropis procera, (B) Hyoscyamus muticus, and (C) Cynanchum acutum under four increasing temperature treatments from four locations in Egypt (Kharga Oasis, Assiut Nile Valley, Assiut Eastern Desert, and Cairo Eastern Desert). The species were sampled during summer 2023–2025. Boxes represent the interquartile range (IQR), the horizontal line within each box indicates the median, and whiskers represent the data range
ANOVA test showed the effects of temperature (T.), location (L.) and their interaction on free amino acids of different species investigated
| Species | ||||||||
|---|---|---|---|---|---|---|---|---|
| Source | C. procera | H. muticus | C. acutum | |||||
| df | F | Share% | F | Share% | F | Share% | ||
| Free amino acids | T. | 3 | 21.28** | 46.77 | 4.96** | 19.98 | 13.76** | 27.15 |
| L. | 3 | 11.05** | 24.29 | 9.76** | 39.31 | 26.39** | 52.09 | |
| T. × L. | 9 | 4.39** | 28.94 | 3.37** | 40.71 | 3.51** | 20.76 | |
Note(s): **Significant at 0.01 level *Significant at 0.05 level
C. acutum had the highest soluble protein content (137.9 mg/g leaf f. wt.) at Kh. O., while H. muticus had the lowest (12.9 mg/g leaf f. wt.) at Ast. D. (Figure 6). An ambient temperature of 55°C generally increased SP across species. C. procera also showed significant SP levels (22.7 mg/g leaf f. wt.) at Kh. O. Statistical analysis indicated that temperature significantly affected soluble protein in all species, with location and its interaction with temperature additionally influencing C. acutum (Table 6).
Three box plots depict total soluble protein contents in three plant species under different temperature treatments from four locations in Egypt. Panel A shows the data for Calotropis procera, Panel B for Hyoscyamus muticus, and Panel C for Cynanchum acutum. Each panel includes box plots for four temperature treatments: 25 degrees Celsius, 35 degrees Celsius, 45 degrees Celsius, and 55 degrees Celsius. The x-axis represents temperature in degrees Celsius, and the y-axis represents total soluble protein content in mg/g leaf fresh weight. Each box plot represents data from four locations: Kharga Oasis, Assiut Nile Valley, Assiut Eastern Desert, and Cairo Eastern Desert. Boxes represent the interquartile range (IQR), the horizontal line within each box indicates the median, and whiskers represent the data range. In Panel A, the protein content for Calotropis procera remains relatively low and consistent across all temperatures and locations.Total soluble protein contents (mg/g leaf fresh weight) in the three studied species: (A) Calotropis procera, (B) Hyoscyamus muticus, and (C) Cynanchum acutum under four increasing temperature treatments from four locations in Egypt (Kharga Oasis, Assiut Nile Valley, Assiut Eastern Desert, and Cairo Eastern Desert). The species were sampled during summer 2023–2025. Boxes represent the interquartile range (IQR), the horizontal line within each box indicates the median, and whiskers represent the data range
Three box plots depict total soluble protein contents in three plant species under different temperature treatments from four locations in Egypt. Panel A shows the data for Calotropis procera, Panel B for Hyoscyamus muticus, and Panel C for Cynanchum acutum. Each panel includes box plots for four temperature treatments: 25 degrees Celsius, 35 degrees Celsius, 45 degrees Celsius, and 55 degrees Celsius. The x-axis represents temperature in degrees Celsius, and the y-axis represents total soluble protein content in mg/g leaf fresh weight. Each box plot represents data from four locations: Kharga Oasis, Assiut Nile Valley, Assiut Eastern Desert, and Cairo Eastern Desert. Boxes represent the interquartile range (IQR), the horizontal line within each box indicates the median, and whiskers represent the data range. In Panel A, the protein content for Calotropis procera remains relatively low and consistent across all temperatures and locations.Total soluble protein contents (mg/g leaf fresh weight) in the three studied species: (A) Calotropis procera, (B) Hyoscyamus muticus, and (C) Cynanchum acutum under four increasing temperature treatments from four locations in Egypt (Kharga Oasis, Assiut Nile Valley, Assiut Eastern Desert, and Cairo Eastern Desert). The species were sampled during summer 2023–2025. Boxes represent the interquartile range (IQR), the horizontal line within each box indicates the median, and whiskers represent the data range
ANOVA test showed the effects of temperature (T.), location (L.) and their interaction on soluble proteins of different species investigated
| Species | ||||||||
|---|---|---|---|---|---|---|---|---|
| Source | C. procera | H. muticus | C. acutum | |||||
| df | F | Share% | F | Share% | F | Share% | ||
| Total soluble proteins | T. | 3 | 5.92** | 58.33 | 2.98* | 28.06 | 52.02** | 67.54 |
| L. | 3 | 2.70 | 26.66 | 1.93 | 18.14 | 5.34** | 6.93 | |
| T. × L. | 9 | 0.51 | 15.00 | 1.91 | 53.80 | 6.55** | 25.52 | |
Note(s): **Significant at 0.01 level *Significant at 0.05 level
Correlation coefficient between water output, leaf succulence, and metabolic compounds
The study investigated how temperature affects correlations among water output, leaf succulence, and metabolites across locations (Tables 7 and 8). Most correlations were positive, except for negative correlations between soluble sugars and leaf succulence in H. muticus at Ast. N. and C. acutum at Cairo D. In H. muticus at Ast. N., soluble sugars were also negatively correlated with water output. C. acutum displayed numerous correlations between metabolites and water output under heat stress. Plants located between latitudes N 25° and N 27° showed a greater number of significant correlations compared with those at latitude N 30°, suggesting a stronger association between the studied parameters under conditions of higher heat stress and atmospheric aridity. Leaf succulence generally exhibited weak or non-significant correlations with the other measured parameters across the different species and locations.
Correlation coefficient between water output, leaf succulence and metabolic compounds in species studied at different locations
| Species | |||||
|---|---|---|---|---|---|
| Locations | Parameters Water output % | ||||
| Total soluble proteins | Amino acids | Total soluble sugars | Leaf succulence | ||
| C. procera | Kh. O. | 0.860 | 0.993** | 0.593 | −0.409 |
| Ast. N. | 0.834 | 0.924* | 0.977** | −0.264 | |
| Ast. D. | 0.971** | 0.908* | 0.929* | −0.138 | |
| Cairo D. | 0.953* | 0.839 | 0.687 | −0.450 | |
| H. muticus | Kh. O. | 0.957* | 0.896* | 0.716 | −0.716 |
| Ast. N. | −0.777 | −0.334 | −0.974** | 0.840 | |
| Ast. D. | 0.998** | 0.879* | 0.978** | −0.291 | |
| Cairo D. | 0.286 | 0.873 | 0.692 | −0.783 | |
| C. acutum | Kh. O. | 0.987** | 0.894* | 0.992** | −0.045 |
| Ast. N. | 0.891* | 0.906* | 0.907* | −0.551 | |
| Ast. D. | 0.931* | 0.912* | 0.825 | −0.182 | |
| Cairo D. | 0.858 | 0.450 | −0.160 | 0.500 | |
Note(s): **Significant at 0.01 level *Significant at 0.05 level
Correlation coefficient between metabolic compounds (total soluble proteins, S. P.; amino acids, A. A.; soluble sugars, S. S.) and leaf succulence (L. Succ.) in species studied at different locations
| Species | |||||
|---|---|---|---|---|---|
| Locations | Parameters S. P. × A. A. | S. P. × S. S. | A. A. × S. S. | S. S. x L. Succ. | |
| C. procera | Kh. O. | 0.858 | 0.496 | 0.684 | −0.822 |
| Ast. N. | 0.969** | 0.932* | 0.981** | −0.067 | |
| Ast. D. | 0.790 | 0.823 | 0.949* | 0.075 | |
| Cairo D. | 0.639 | 0.741 | 0.508 | 0.105 | |
| H. muticus | Kh. O. | 0.798 | 0.597 | 0.951* | −0.637 |
| Ast. N. | 0.616 | 0.892* | 0.504 | −0.891* | |
| Ast. D. | 0.876 | 0.987** | 0.897* | −0.419 | |
| Cairo D. | 0.141 | −0.153 | 0.350 | −0.711 | |
| C. acutum | Kh. O. | 0.813 | 0.959** | 0.944* | −0.102 |
| Ast. N. | 0.925* | 0.980** | 0.982** | −0.180 | |
| Ast. D. | 0.996** | 0.937* | 0.963** | −0.278 | |
| Cairo D. | 0.825 | 0.250 | 0.739 | −0.924* | |
Note(s): **Significant at 0.01 level *Significant at 0.05 level
Discussion
The study examined the impact of temperature on leaf physiology and metabolites in desert plants. Leaf temperature closely correlates with ambient air temperature, often exceeding it and causing thermal stress that can harm leaf biochemistry (Kibler et al., 2023). Leaf succulence varied among species, crucial for survival under heat stress. H. muticus exhibited the highest succulence, followed by C. procera, enhancing their heat resistance, while C. acutum had the lowest succulence, indicating less water storage. Plants use morphological, anatomical, and biochemical defenses under thermal stress, with species-specific differences affecting leaf succulence (Egigu et al., 2014).
Increased leaf temperature significantly affects water output, particularly in succulent leaves, as smaller leaf area reduces evapotranspiration under heat stress. C. acutum showed increased water output at 45–55°C across locations, whereas lower temperatures reduced water output in plants from Cairo D. and Kh. O., and species from Ast. D. exhibited higher water output under arid thermal stress. C. procera and H. muticus maintained controlled water loss below 45°C due to smaller leaf area, which aids physiological functions (Hultine et al., 2020). Statistical analysis indicated that temperature plays an important role in influencing water output in the studied species.
Under thermal stress, plants adopt protective strategies such as producing compatible solutes and colloidal particles, crucial for their metabolism, development, and growth (Admas, Shu, Shalmani, Pan, & Zhang, 2025). Enhanced production of soluble sugars (SS) under high temperatures was observed in various plant species, particularly C. acutum in Cairo D., which showed maximum SS levels comparable to those in other locations, except in Ast. N., where the SS peaked at supra-optimal temperatures of 55°C. The increased SS concentration in C. acutum is linked to the plants' defense against temperature and oxidative stresses (De Abreu & Mazzafera, 2005), exacerbated by restricted leaf water potential due to excessive evapotranspiration (Kohila & Gomathi, 2018). In contrast, H. muticus and C. procera had lower SS levels but managed to survive challenging thermal conditions due to a better water supply. Abdul Wahid (2007) concluded that the reduction in leaf water potential from thermal effects is compensated by synthesizing primary metabolites, enhancing survival by maintaining water status (Mansinhos et al., 2022). The study underscores significant effects of temperature and location on SS content, highlighting location's critical role for H. muticus and temperature's impact on C. procera and C. acutum.
Free amino acids act as protective antioxidants under heat stress (Shamloo et al., 2017). C. acutum accumulated the highest levels, while C. procera and H. muticus were lower. Plants at Ast. D. showed enhanced accumulation. Proline, a compatible osmolyte, accumulates under extreme stress and supports osmotic balance (El-Sharkawi, Farghali, Rayan, & Abdel, & Dalia, 2018). Temperature dominated amino acid accumulation in C. procera, location in C. acutum, and their interaction in H. muticus.
Thermal stress disrupts cellular volume and solute composition, promoting accumulation of water-binding proteins (Zhou, Ma, & Sun, 2008). Elevated temperatures stimulated soluble protein (SP) synthesis, particularly in C. acutum at Kh. O. Heat shock proteins protect enzymes, membranes, and structures from denaturation (Abbas, Saggu, Rehman, Al Thbiani, & Ansari, 2017). SP increases at 55°C serve as nitrogen reserves and osmotic regulators (Hanif et al., 2021), with low molecular weight proteins enhancing thermostability (El-Sharkawi, Farghali, Rayan, & Tammam, 2016). H. muticus and C. procera produce less SP due to higher leaf water storage. Temperature was the main factor affecting SP accumulation.
The study observed rare correlations between leaf succulence and other parameters in plants, particularly in Cairo D. Only H. muticus and C. acutum showed a negative correlation between soluble sugars and leaf succulence, and soluble sugars were negatively correlated with water output in H. muticus. Other significant correlations were positive, especially among plants in extreme environments at latitude N 27°. C. acutum exhibited more correlations than other species, suggesting that the interplay between metabolic compounds and water output is vital for thermal resistance in plants from hot drylands.
Measured morphological traits reflected species-specific adaptations. C. procera displayed the largest leaf area, which may enhance photosynthesis but also increase heat load and water loss (Taiz, Zeiger, Møller, & Murphy, 2015). C. acutum had the smallest leaf area and highest leaf mass per area, suggesting thicker leaves that support water conservation (Reich, 2014). H. muticus showed intermediate values, indicating a different structural strategy for maintaining water balance. Leaf succulence observations suggest that H. muticus and C. procera store more water than C. acutum. Overall, these data suggest that desert plants integrate physiological and structural traits to optimize water conservation and thermal regulation under extreme conditions.
Conclusion
Elevated ambient temperatures (45–55°C) significantly impacted the water output and metabolic compounds in broad leaf plants. C. acutum exhibited increased levels of soluble sugars, free amino acids, and soluble proteins to mitigate water loss. In contrast, C. procera and H. muticus displayed low levels of metabolic osmolytes but high leaf succulence, which enhances water use efficiency during thermal stress. This study provides insights for cultivating succulent broad leaf economical plants in hot deserts and highlights their role in regulating climate change and receiving high insolation in extreme environment conditions.
