Physiological and Metabolomic Analyses of Exogenously Applied Sorbitol-Chelated Potassium Enhancing Drought Tolerance in Wheat

Physiological and Metabolomic Analyses of Exogenously Applied Sorbitol-Chelated Potassium Enhancing Drought Tolerance in Wheat | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Physiological and Metabolomic Analyses of Exogenously Applied Sorbitol-Chelated Potassium Enhancing Drought Tolerance in Wheat Mingxia Zhang, Guohui Du, Huanyang Zhang, Ruili Zheng, Li Zhao, and 4 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-7498264/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Potassium fertilization is a strategy to alleviate the impact of drought stress on wheat production. However, the effects of chelated potassium remain to be verified. In this study, 10% PEG-6000 was used to simulate moderate drought stress on hydroponically grown XinHua818 wheat ( Triticum aestivum L. ) seedlings, and the physiological and biochemical parameters of wheat sprayed with water (CK 2 ), sorbitol (S), potassium chloride (K), potassium mixed with sorbitol (MK), and sorbitol-chelated potassium (SK) were monitored. Results showed that SK effectively alleviated the inhibitory effects of drought stress on seedling growth. The aboveground biomass of SK-treated seedlings was significantly higher than that of K and MK-treated seedlings, increasing by 15.66% and 20.00%, respectively. Compared to MK treatment, SK treatment significantly increased total chlorophyll content by 18.74% and reduced malondialdehyde levels by 16.02%. Superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) activity were also elevated in SK-treated seedlings compared to other treatments. Metabolomic analysis identified 51 differential metabolites in SK compared to CK 2 treatment, including sugars, amino acids, lipids, plant hormones, carotenoids, flavonoids, and their derivatives. These metabolites were enriched in 18 metabolic pathways, notably α-linolenic acid metabolism, histidine metabolism, plant hormone signal transduction, carotenoid biosynthesis, and flavonoid biosynthesis, suggesting their potential role in enhancing drought tolerance in wheat and their broader significance in drought resistance research. Wheat Drought stress Sorbitol-chelated potassium Antioxidant system Metabolomics Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 Introduction Drought stress disrupts key physiological processes in plants, impairing photosynthesis and antioxidant defenses [ 1 ], and is a major factor limiting wheat growth and yield [ 2 ]. As an early response to water deficit, photosynthesis is notably vulnerable, with drought-induced damage to photosynthetic pigments, reductions in net photosynthetic rate, stomatal conductance, and transpiration rate, these ultimately disrupt plant metabolism and suppress biomass accumulation [ 3 , 4 ]. In parallel, drought stress induces overproduction of reactive oxygen species (ROS), causing oxidative damage to plant cells and tissues. To mitigate the oxidative damage of ROS to cells and maintain redox balance, plants activate enzymatic antioxidant systems, which are composed of peroxidase (POD), superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPX), and non-enzymatic antioxidant systems, including secondary metabolites such as flavonoids (FC) and total phenols (TPC) compounds [ 5 , 6 ]. Plant responses to drought by profiling metabolite changes. Serving as a bridge between plant genotype and phenotypes, the metabolome provides insights into mechanisms underlying phenotypic variation under stress conditions [ 7 ]. Wang et al. [ 8 ] using LC -MS technology, four key types of differential metabolites, namely amino acids, organic acids, sugars, and alkaloids, such as gibberellin A4 (GA4), abscisic acid (ABA), and sucrose, have been identified. These metabolites are crucial for alfalfa to withstand drought stress. Bowne et al. [ 9 ] investigated a targeted GC-MS approach to monitor 103 structurally identified metabolites from leaf tissue of the drought-stressed wheat plants, predominantly amino and organic acids and sugars. However, most drought-stress studies have focused on individual plant systems, leaving the interactions among photosynthetic, antioxidant, and metabolomic responses underexplored. Critical for maintaining growth and yield stability. Potassium, a key macronutrient, plays a pivotal role in stomatal regulation, photosynthesis, ROS detoxification, and stress resistance [ 10 – 12 ]. Drought impairs root growth and limits potassium from the soil [ 13 ], making exogenous potassium application an effective strategy to compensate for the potassium deficiency and enhance drought tolerance. Sorbitol-chelated potassium, a novel chelated foliar potassium fertilizer, has demonstrated a positive regulatory role in improving wheat drought resilience. Compared to conventional inorganic (non-chelated) potassium fertilizers, sorbitol-chelated potassium enhances potassium uptake, stress resistance, and crop yield. In wheat, sorbitol-chelated potassium application has been reported to increase post-anthesis potassium accumulation and yield by 13.99%-35.48% [ 14 ]. Similar benefits have been observed in other crops, including enhanced dry matter accumulation and kernel potassium content in peanuts (average yield increase of 18.9%) [ 15 ] and improved growth, potassium uptake, and quality in celery [16.17]. Despite these findings, studies investigating the effects of chelated potassium on crops under drought stress remain limited [ 18 ]. Given the role of potassium in wheat growth and stress responses, elucidating how sorbitol-chelated potassium enhances drought resistance holds significant scientific and practical implications. Because drought affects multiple physiological pathways, including photosynthesis, antioxidant defense, and metabolism, a systematic analysis integrating these factors is necessary to uncover potential mechanisms by which sorbitol-chelated potassium alleviates drought-induced damage in wheat. Materials and methods Plant materials and seed preparation Plump XinHua818 wheat seeds of uniform size were selected and surface sterilized with 0.1% sodium hypochlorite for 10 minutes, followed by three rinses with ultrapure water. The sterilized seeds were incubated in the dark at 25℃ for 24 hours in a biological incubator to promote germination. Seedling cultivation and hydroponic growth conditions Germinated seeds were placed into seedling trays lined with three layers of gauze and grown in a greenhouse under controlled conditions: temperature 25℃, light intensity 5000lx, and a photoperiod of 16 h light/8 h dark. Once the seedlings reached 5–6 cm in height, they were transferred to plastic hydroponic containers (70 seedlings per container) containing Hoagland nutrient solution. The solution volume in each container was maintained consistently and replaced every 3 days. Experimental design and drought treatment At the two-leaf and one-heart stage, seedlings were transferred to hydroponic containers (8.5 cm diameter, 12 cm height) to commence experimental treatments. Six treatments were established, each with three biological replicates (48 seedlings per replicate). Drought stress was simulated using 10% polyethylene glycol (PEG-6000) in the nutrient solution for 7 days. Foliar sprays corresponding to each treatment were applied on days 1 and 3 (twice in total) of drought period. After 7 days of drought stress, seedling samples were collected and immediately frozen in liquid nitrogen for subsequent analyses. The normal control (CK 1 ) was treated with Hoagland nutrient solution, while the drought-stressed control (CK 2 ) was treated with Hoagland nutrient solution supplemented with 10% PEG-6000. The experimental design is detailed in Table 1 . Table 1 Experimental setup with different foliar spray treatments Treatment Growth environment Spraying concentration (mg·L − 1 ) Control (CK 1 ) Hoagland nutrient solution - Control (CK 2 ) Hoagland nutrient solution + 10%PEG-6000 Distilled water 10 (K + ) Sorbitol (S) Potassium Chloride (K) Potassium Mixed with Sorbitol (MK) Sorbitol-chelated Potassium (SK) Determination of growth indicators and leaf relative water content For each treatment, twelve wheat plants were selected, washed with pure water, and wiped dry before measuring plant height and stem diameter. Each whole plant was then divided into above-ground and below-ground parts, and immediately weighed, with the weights recorded as root fresh weight and leaf fresh weight (FW). The leaves were soaked in double-distilled water for 24 h and then removed. After removing residual water from the leaf surface with absorbent paper, the leaves were weighed and the weight recorded as turgid weight (TW). The leaves were then deactivated at 105°C for 30 min and dried at 60°C to a constant weight, which was recorded as dry weight (DW). The relative water content of the leaves was calculated via the following formula: RWC (%) =(FW-DW)/(TW-DW) ×100% [ 19 ]. Three biological replicates were analysed. Determination of chlorophyll content and gas exchange Following the methods of Lichtenthaler [ 20 ], with slight modifications, the first fully expanded leaf of wheat seedlings was selected, ground with a mortar, and extracted with acetone. The absorbance of chlorophyll a , chlorophyll b and carotenoids were measured at OD 663 , OD 645 and OD 480 nm using a spectrophotometer. Following drought stress, the net photosynthetic rate (Pn), stomatal conductance (Gs), transpiration rate (Tr), and intercellular CO 2 concentration (Ci) were determined using an Li-6800 portable photosynthesis system (Li-CORBiosciences, Lincoln, NE, USA), as previously described by He et al. [ 21 ]. The measurements were conducted on the last fully expanded leaf, employing a photon flux density of 1000 µmol photo m − 2 s − 1 and a CO 2 concentration of 400 µmol mol − 1 , within the time from of 9:00 a.m. to 11:00 a.m. Antioxidant enzyme activity and malondialdehyde content The activities of superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) were determined using assay kits (Boxbio, Beijing) according to the manufacturer’s instructions. One gram of leaf sample was homogenized in 1 mL of extraction buffer in an ice bath, followed by centrifugation at 8,000 g for 10 min at 4°C. The supernatant was then collected and used for enzyme activity assays. Following the method outlined by Tan et al. [ 22 ], a 1 g sample was homogenized with 10 mL of a mixed TCA-TBA solution, followed by centrifugation at 4000 rpm for 10 min. The mixture was kept in a boiling water for 15 minutes and centrifuged at 4,000 g for 10 min. Malondialdehyde (MDA) content in the reaction solution was determined using a spectrophotometer at OD 450 , OD 532 and OD 600 nm. Metabolite extraction Six wheat seedlings were randomly selected for each treatment group. Leaves from the same position on each seedling were collected, immediately flash-frozen in liquid nitrogen, and then stored in a -40 ℃ freezer. The metabolite extraction steps were as follows: (1) Weigh 50 mg of the sample into a pre-cooled mortar and grind it into powder. The powder is then transferred to an EP tube, and 1000 µL of a mixed extraction solution of methanol, acetonitrile, and water (2:2:1, containing internal standard at 20 mg L − 1 ) is added, followed by vortexing for 30 s. (2) Steel beads are added, and the mixture is processed using a grinding instrument (45 Hz) for 10 min, followed by sonication in an ice-water bath for 10 min. Subsequently, the sample is allowed to stand at -20°C for 1 h, and then centrifuged at 12000 rpm (4°C, 15 min). (3) 500 µL of the supernatant is transferred to an EP tube and dried in a vacuum concentrator. The extract is then reconstituted with 160 µL of acetonitrile/water (1:1, v/v), vortexed for 30 s, and sonicated in an ice-water bath for 10 min. (4) Collect 120 µL of the supernatant (12000 rpm, 4°C, 15 min) into a 2 mL vial. 10 µL from each sample is then pooled to create a QC sample for on-machine analysis. Chromatographic conditions: An Acquity UPLC HSS T3 column (1.8 µm, 2.1*100 mm, Waters, US) was used with mobile phase A (0.1% formic acid in water) and mobile phase B (0.1% formic acid in acetonitrile). Chromatographic data acquisition was performed on an Acquity I-Class PLUS ultra-high-performance liquid chromatograph (Waters, US) coupled to a Xevo G2-XS QTOF (Waters, US) high-resolution mass spectrometer, with detection in both positive and negative ion modes. Statistic analysis Normality and log-normality tests, along with one-way ANOVA, were conducted using SPSS 27.0 and GraphPad Prism 10.1.2 to determine differences between numerical values. Structural equation modeling (SEM) data processing was performed using SetupStata 18, and Power Point 2021 and Hiplot ( https://hiplot.cn/basic/ggscatterstats ) were used to generate figures. Raw metabolomics data were collected using MassLynx V4.2 software, and Progenesis QI software was used for data processing operations such as mass peak extraction and alignment [ 23 ]. PCA and heatmap images were generated using MetaboAnalyst 6.0 ( https://www.metaboanalyst.ca/ ) and Origin 2021. Commercial databases, including the online METLIN database (based on Progenesis QI software), KEGG (Kyoto Encyclopedia of Genes and Genomes) ( http://www.genome.jp/kegg/ ), HMDB ( https://hmdb.ca ), and Lipidmaps ( http://www.lipidmaps.org/ ), were used to search metabolic pathways for profiling and metabolite identification. Results Effect of sorbitol-chelated potassium on wheat growth and biochemical indices under drought stress Growth traits Different foliar spray treatments significantly affected the wheat seedling growth traits, including biomass, plant height, stem diameter, and leaf relative water content (RWC) (Table2). Notably, sorbitol-chelated potassium treatment effectively promoted biomass accumulation compared to other drought stress treatments. Under drought stress, aboveground biomass in SK-treated seedlings increased by 15.66% and 20.00% compared to K and MK treatments, respectively (both significant). Root biomass also increased under SK treatment by 3.23% and 10.34% compared with K and MK treatments, respectively, though the difference was not significant relative to K treatment. Plant height increased by 8.42% under SK compared to K treatment, whereas stem diameter did not differ significantly across treatments. Additionally, SK treatment increased leaf RWC by 15.25% compared to K treatment, indicating enhanced water retention under drought stress. Table 2 Growth indicators of wheat seedlings under normal conditions and drought stress with different fertilization treatments Treatment Aboveground biomass (g) Root biomass(g) Plant height (cm) Stem diameter (mm) Relative water content of leaves CK 1 3.04±0.09a 0.31±0.00ab 23.46±0.98a 2.46±0.04a 0.88±0.03a CK 2 2.42±0.03b 0.28±0.01c 22.23±0.03ab 2.27±0.04c 0.56±0.03cd S 2.47±0.10b 0.28±0.01c 22.29±0.24ab 2.44±0.03ab 0.47±0.01d K 2.49±0.08b 0.31±0.02ab 21.37±0.26b 2.40±0.00ab 0.59±0.06bc MK 2.40±0.08b 0.29±0.00bc 22.42±0.16ab 2.39±0.02b 0.63±0.06bc SK 2.88±0.06a 0.32±0.01a 23.17±0.80a 2.40±0.03ab 0.68±0.05b Note: Different lowercase letters in the same column indicate significant differences between treatment groups for the same indicator ( p <0.05). (Above ground and root biomass measured as fresh weight) Photosynthetic pigments and gas exchange Different foliar spray treatments significantly affected the accumulation of chlorophyll a , chlorophyll b , carotenoids, and total chlorophyll content in wheat seedlings under drought stress (Fig. 1). Drought stress reduced photosynthetic pigment content, whereas foliar spraying alleviated this effect. Compared with the ionic potassium treatment, the sorbitol-chelated potassium treatment effectively increased chlorophyll a and total chlorophyll content, with significant differences. Chlorophyll a increased by 11.60% and 19.54% in SK treatment compared to K and MK treatments, respectively. Similarly, chlorophyll b and carotenoid content increased by 15.94% and 10.68%, respectively, compared with MK treatment. Carotenoid levels in SK treatment decreased by 1.52% and 4.11% relative to K and S treatments, respectively, though these differences were not statistically significant. Total chlorophyll content in SK treatment increased by 18.74% compared to MK treatment, a difference that was statistically significant. Under drought stress, different spraying treatments significantly affected the net photosynthetic rate (Pn), transpiration rate (Tr), intercellular CO 2 concentration (Ci), and stomatal conductance (Gs) of wheat seedlings (Fig. 2). Drought stress decreased leaf gas exchange parameters relative to the control treatment (CK 1 ). However, these parameters improved following foliar treatments. Compared with the MK, K, and S treatments, SK treatment significantly increased Pn by 62.90%, 31.98%, and 65.00%, respectively, and Ci by 14.55%, 23.28%, and 14.00%, respectively. Tr decreased by 18.64% and 25.00% under SK treatment compared to K and S treatments, respectively. However, these differences were not statistically significant. Gs decreased by 15.39% and 26.75% compared to MK and S treatments, respectively, with no significant differences. Antioxidant System Under drought stress, the levels of superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT), as well as malondialdehyde (MDA) content in wheat seedlings, were significantly affected by different foliar spray treatments (Fig. 3). Drought stress significantly reduced SOD and POD activities. However, foliar application with sorbitol-chelated potassium (SK), a mixture of sorbitol and potassium chloride (MK), sorbitol (S), potassium chloride (K) significantly increased SOD and POD activity compared to control treatment (CK 2 ). SOD activity increased by 46.07% and 19.58% in SK treatment relative to MK and S treatments, respectively. CAT activity increased by 64.00% in SK treatment compared to S treatment. POD activity followed a similar trend, increasing by 7.23% and 19.46% in SK compared to K and MK treatments, respectively, but decreasing by 19.92% compared to S treatment. Drought stress induced the accumulation of malondialdehyde (MDA) levels, enhancing membrane lipid peroxidation and causing damage to wheat seedlings (Fig. 3D). The MDA levels was decreased by 16.02% in SK treatment compared to MK treatment. Metabolomic profiles of wheat under drought stress Principal component analysis (PCA) PCA was performed to compare metabolic differences among the treatment groups (SK, K, MK, S, and CK 2 ) and assess the overall metabolomic changes in wheat seedling leaves under drought stress. Additionally, the repeatability and stability of metabolites within each treatment group were evaluated. Figure 4A and B presents the PCA results for positive and negative ion modes. The total explained variances were 51.5% and 44.4%, respectively, indicating satisfactory clustering of samples. In both modes, the metabolite profiles of potassium chloride (K) and sorbitol-chelated potassium (SK) treatments were clearly separated, suggesting distinct metabolic responses. While partial overlap was observed between chelated potassium (SK) and non-chelated potassium (MK) treatments, overall significant differences persisted, indicating that the three potassium formulations exerted differential effects on the metabolism of wheat seedling leaves. These metabolic distinctions were consistent with the variations in agronomic traits, physiological parameters, and biochemical indices described as described above. Identification of differentially expressed metabolites A total of 418 differentially expressed metabolites (DEMs) were identified from 1788 metabolites (Supplemental Material 1). Compared with CK 2 , SK showed 11 DEMs upregulated and 40 downregulated; K had 30 upregulated and 17 downregulated; MK had 28 upregulated and 231 downregulated; and S presented 27 upregulated and 34 downregulated (Fig. 5). It was found that the number of downregulated DEMs was greater than that of upregulated DEMs in response to drought stress when each treatment was compared with CK 2 . To clarify the characteristics of DEM expression in sorbitol-chelated potassium, upregulated metabolites (FC≥2) in SK.VS.CK 2 were found to include Prolyl-Histidine, Lariciresinol, Chelirubine, L-Sepiapterin, 3,5-Diprenyl-4-hydroxybenzaldehyde,, Etherolenic acid and 4-Amino-4-deoxychorismate, while downregulated metabolites (FC≤0.5) included (-)-Jasmonoyl-L-isoleucine, N-Acetyl-D-glucosamine, (+)-Abscisic acid, 5-Hydroxypseudobaptigenin, and D-Glucosaminate-6-phosphate. Functional annotation and enrichment analysis of differential metabolites based on KEGG To identify the metabolic pathways involved, DEMs from different comparison groups were matched against the KEGG database to obtain information on as many metabolites as possible in each extract (Supplementary Material 2, Fig. S1). Enrichment analysis was then performed on the annotated results based on the P-values of the metabolic pathways, revealing a total of 119 metabolic pathways in both positive and negative ion modes. Compared with CK 2 treatment, 17, 16, 25, and 109 DEMs were identified for sorbitol-chelated potassium, potassium, sorbitol-mixed potassium, and sorbitol, respectively, and subsequently annotated to 18, 16, 24, and 61 metabolic pathways, respectively (Supplementary Material 3). The top ten most significantly enriched metabolic pathways of differential metabolites were selected (Fig. 6). Compared to CK 2 , foliar spraying with SK, K, MK, and S activated a broad range of metabolites, particularly those involved "antioxidation" and regulating "osmotic potential" [24]. These findings indicate that foliar application of osmoprotectants mitigates drought-induced damage in wheat seedlings. In contrast, the SK, K, MK, and S mobilized the most diverse set of metabolites. Specifically, in SK.VS.CK 2 , upregulated metabolites were primarily enriched in metabolic pathways such as alpha-Linolenic acid metabolism, Folate biosynthesis, and Isoquinoline alkaloid biosynthesis, while downregulated metabolites were mainly enriched in pathways such as the Pentose phosphate pathway, Flavonoid biosynthesis, and Plant hormone signal transduction. Metabolic pathway enrichment analysis revealed that SK application modulated key metabolic pathways associated with intracellular water balance, ROS scavenging, and overall seedling vigor, thereby improving drought tolerance in wheat. The correlation of wheat growth with physiological and biochemical indices, and differential metabolites Differential metabolites in wheat exhibited a close relationship with growth morphology, photosynthetic performance, and antioxidant enzymes (Fig. 7). Correlation analysis revealed that N-Acetyl-D-glucosamine and (-)-Jasmonoyl-L-isoleucine were extremely significantly positively correlated with wheat growth indicators, while 5-Methoxytryptamine showed a significant positive correlation. Furthermore, (-)-Jasmonoyl-L-isoleucine and 5-Methoxytryptamine were extremely significantly positively correlated with wheat photosynthetic performance, and 9,10-Dihydroxy-12,13-epoxyoctadecanoic acid, (-)-Maackiain, and N-Acetyl-D-glucosamine were significantly positively correlated. Etherolenic acid was significantly positively correlated with antioxidant enzymes, suggesting that these metabolites interact within the plant metabolic network to cooperatively maintain normal plant physiological functions under drought stress. Additionally, 9,10-Dihydroxy-12,13-epoxyoctadecanoic acid and (-)-Maackiain were positively correlated with wheat growth morphology; D-Glucosaminate-6-phosphate, Betaine, Rhodopinal, GibberellinA4, and Hydroxychlorobactene were positively correlated with wheat photosynthetic performance; Chlorophylla, Rhodopinal, 5-Methoxytryptamine, Hydroxychlorobactene, Astaxanthin, (-)-Maackiain, and GibberellinA4 were positively correlated with the antioxidant system. Under drought stress, foliar spraying induced the accumulation of sugars, amino acids, lipids, plant hormones, carotenoids, and flavonoids, which collectively regulated wheat growth, physiology, and biochemical responses. To further elucidate the mechanisms underlying these effects, structural equation modeling (SEM) (Fig. 8A) was employed to quantify the direct and indirect influences of five factors, growth morphology, photosynthetic performance, antioxidant enzyme activity, primary metabolites, and secondary metabolites, on biomass accumulation. SEM results indicated a good model fit, with the model χ² value at 77.01, RMSEA at 0.00, CFI at 1.00, and AIC at 324.68, all fitting indices being within the standard range. SEM findings revealed that all five factors had either direct or indirect effects on biomass accumulation. Growth morphology, photosynthetic performance, antioxidant enzymes, and secondary metabolites exerted direct positive effects on biomass (direct effect value: 0.665, 0.211, 0.306, and 0.353), with growth morphology showing the strongest most significant influence. In contrast, primary metabolites exerted a direct negative effect on wheat biomass (-0.620, nonsignificant), but their indirect effect on biomass accumulation was substantial, primarily mediated through their influence on secondary metabolites. Overall, growth morphology exhibited the greatest total effect (0.685) (Fig. 8B) on biomass, followed by photosynthetic performance, antioxidant enzymes, and secondary metabolites (all positive total effects), whereas primary metabolites had a negative total effect. Discussion Exogenous sorbitol-chelated potassium affects growth and photosynthetic performance of wheat seedlings under drought stress Drought stress inhibits wheat seedling growth and biomass accumulation, reduces photosynthetic pigment content, and directly impairs net photosynthetic efficiency and stomatal conductance. This study demonstrates that exogenous sorbitol-chelated potassium application positively influences wheat seedling growth under drought conditions. Drought stress impedes the synthesis of photosynthetic pigments in wheat seedling leaves, thereby reducing photosynthetic capacity. Chlorophyll, a key pigment for light energy absorption, plays a key role in regulating photosynthetic efficiency [ 25 ]. In this study, SK treatment significantly increased total chlorophyll content compared with non-chelated MK treatment, and this change was positively correlated with the net photosynthetic rate. These findings suggest that SK helps inhibit chlorophyll degradation, thereby maintaining photosynthetic stability under drought stress. To cope with water deficit, plants typically adjust their stomatal aperture to reduce transpiration, limiting gas exchange and preventing excess water loss [ 24 ]. In this study, SK treatment reduced stomatal conductance, indicating enhanced drought resistance through partial stomatal closure. This effect may reflect sorbitol-chelated potassium induced regulation of stomatal behavior near its physiological threshold. While stomatal closure conserves water, it also limits CO₂ entry, potentially limiting photosynthesis. Research indicates that a decline in intercellular carbon dioxide concentration reduces photosynthetic activity, consequently impairing plant growth and development [ 26 ]. By converting free potassium ions into organically chelated forms, sorbitol-chelated potassium enhances photosynthetic efficiency and facilitates the transport of photosynthetic products, thereby promoting crop growth and development. Chelated potassium penetrates the waxy cuticular layer of leaves more effectively than free ionic forms. The lipophilic nature of the chelate allows more efficient passage through the lipid bilayer of the cuticle and enhances its mobility within the phloem. In this study, sorbitol-chelated potassium not only regulated wheat seedling growth but also significantly improved photosynthetic performance and activated antioxidant enzyme activity. These findings provide an important foundation for a deeper understanding of the comprehensive physiological mechanisms by which sorbitol-chelated potassium alleviates drought stress. Exogenous sorbitol-chelated potassium mitigate oxidative stress by enhancing antioxidant defense Drought stress leads to increased malondialdehyde (MDA) accumulation and disrupts ROS homeostasis in wheat seedlings. MDA, a well-established marker of lipid peroxidation, was significantly elevated in CK 2 treatment, consistent with the findings of Aksu [ 27 ]. Compared to MK treatment, the application of SK significantly reduced MDA levels, suggesting that SK enhances ROS scavenging capacity. This effect may be attributed to the coordination properties of sorbitol, which converts potassium ions from free ionic state into chelated form. Following foliar absorption, chelated potassium is more readily translocated within wheat plants and experiences reduced fixation in the phloem. This mechanism is comparable to how sorbitol facilitates calcium transport [ 28 ], contributing to improved drought stress mitigation in wheat plants. Moreover, drought stress stimulates ROS-mediated redox signaling, which activates plant defense mechanisms. The enzymatic antioxidant system, which includes SOD, POD, CAT and other enzymes, plays a crucial role in detoxifying ROS and preventing oxidative damage [ 29 ]. In this study, sorbitol-chelated potassium application significantly enhanced SOD, POD, and CAT activity. SOD catalyzes the dismutation of superoxide radicals (·O ⁻2 ) into hydrogen peroxide (H 2 O 2 ) and oxygen (O 2 ). Subsequently, POD and CAT detoxify H 2 O 2 by reducing it to water, further mitigating oxidative damage [ 30 ]. Therefore, the foliar application of SK alleviates drought stress in plants by enhancing antioxidant enzyme activity and improving ROS detoxification. Building on these findings, metabolomic analysis provides additional insights into the molecular mechanisms through which SK modulates redox balance and enhances drought tolerance at the metabolic regulation level. Effects of exogenous sorbitol-chelated potassium on differential metabolite expression of wheat seedlings under drought stress KEGG enrichment analysis revealed that both metabolic pathways and biosynthetic pathways of secondary metabolites were highly enriched with differential metabolites, suggesting a close association between these pathways and wheat growth under water-limited conditions. In this study, a comparison with the CK 2 treatment identified a total of 167 up-regulated and 170 down-regulated metabolites in the metabolic and biosynthetic pathways of secondary metabolites in the SK, K, MK, and S treatments. Compared with CK 2 treatment, the K exhibited a higher number of upregulated differential metabolites than downregulated ones. Conversely, SK, MK, and S treatments showed a greater number of downregulated than upregulated metabolites. This pattern suggests that exogenous application of sorbitol-containing treatments primarily mediates drought stress responses through metabolite downregulation, whereas potassium applied alone (K) may mitigate drought-induced damage by promoting metabolite upregulation. The upregulation of differential metabolites likely reflects an active adaptive response of wheat seedlings to environmental stress, enabling the enhancement of protective biochemical processes. Conversely, the downregulation of metabolites may represent a strategy for reducing overall metabolism, suppressing growth, and facilitating adaptation to unfavorable conditions. To mitigate drought stress, wheat synthesizes significant quantities of primary and secondary metabolites, including soluble sugars, amino acids, lipids, flavonoids, and other antioxidant compounds, while also activating detoxification enzymes [ 31 ]. These differential metabolites highlight the mechanisms through which exogenous sorbitol-chelated potassium application mobilizes metabolic pathways in wheat leaves to coordinate physiological and biochemical responses, thereby alleviating drought-induced damage in wheat. Primary Metabolites Sugars serve as not only as a primary energy source for plants but also as key signaling molecules that regulate various physiological processes during growth and stress adaptation [ 32 ]. Varshney et al. [ 33 ] revealed that celery and many Rosaceae species exhibit drought stress resistance primarily due to the accumulation of sorbitol and mannitol, which function as osmotic protectants and antioxidants. Soluble sugars undergo rapid interconversion; sucrose is hydrolyzed into glucose and fructose, while these hexoses, in turn, promote sucrose resynthesis, thus integrating into multiple metabolic pathways [ 34 ]. The results of this study indicate that in the SK, MK, and S treatments, D-glucosamine-6-phosphate and xylitol participates in glycolysis and gluconeogenesis via the pentose phosphate pathway, yet its levels were downregulated. This downregulation may indicate an adaptive strategy to reduce energy consumption, as gluconeogenesis is energetically costly, thereby limiting energy loss under stress conditions. Previous studies suggest that sugar metabolism plays a dual role in regulating both energy balance and osmotic adjustment during drought. Furthermore, accumulating evidence indicates that amino acids also play a crucial role in plant stress resistance by functioning as osmotic regulators and signaling molecules, reinforcing their significance in drought tolerance mechanisms. Amino acids function as osmolytes, precursors of secondary metabolites, ROS scavengers, and potential regulatory and signaling molecules that help plants to cope with stress. Elevated levels of specific amino acids are considered functionally significant in enhancing stress resistance [ 35 ]. Drought stress typically induces a significant accumulation of free amino acids in wheat seedlings, which may contribute to improved osmotic stress tolerance. These findings align with previous reports in other plant species. In this study, we observed significant changes in the metabolic pathways of several amino acids in wheat leaves under drought stress. Notably, pathways related to tryptophan, glutamate, valine, leucine, and isoleucine were significantly downregulated, while the free amino acid pools of proline, tryptophan, and branched-chain amino acids (valine, leucine, and isoleucine) were elevated [ 36 , 37 ]. In our study, we found that the metabolic pathways of several high-abundance amino acids and most low-abundance amino acids in wheat leaves were significantly downregulated during drought stress, including those of tryptophan, glutamate, valine, leucine, and isoleucine, suggesting that wheat seedlings mitigate drought stress by accumulating free amino acids. Levels of amino acids, most notably proline, tryptophan, and the branched chain amino acids leucine, isoleucine, and valine were increased under drought stress in all cultivars. These findings suggest that wheat seedlings mitigate drought stress through the accumulation of amino acids, serving both as osmoprotectants and regulators of metabolic homeostasis. Lipids and lipid-like molecules in plant cells are critical not only for membrane structure and energy storage but also for diverse biological functions. They serve as signaling molecules and as precursors for the synthesis of defense-related phytohormones, such as jasmonic acid [ 38 ]. In this study, linolenic acid and α-linolenic acid metabolic pathways were significantly affected by the SK, MK, and S treatments. Compared with potassium application alone, sorbitol-containing treatments promoted greater lipid accumulation in wheat leaves, suggesting enhanced membrane integrity and stress signaling capacity. In addition, purine and pyrimidine metabolism, key components of plant energy metabolism [ 39 ], were modulated by SK application, contributing to the maintenance of energy homeostasis under drought conditions. Collectively, these findings demonstrate that exogenous SK application significantly enhances wheat drought tolerance by regulating carbohydrates, amino acid, and lipid metabolism, thereby improving energy metabolism, osmotic regulation, and cell defense mechanisms. These findings reveal the complexity and synergy of plant metabolic networks and provide an important foundation for further exploration of the role of secondary metabolites in drought resistance mechanisms in wheat seedlings. Secondary Metabolites Plant hormones are central regulators enabling plants to cope with environmental stress, integrating external drought signals with internal physiological responses. In this study, the formation of the jasmonoyl-isoleucine conjugate in the SK treatment likely contributed to stress resistance. Jasmonoyl-isoleucine levels typically increase under dehydration stress [ 40 ] and interact with abscisic acid (ABA) accumulation to participate in the dehydration signal transduction pathway [ 41 , 42 ]. Therefore, sorbitol-chelated potassium may alleviate drought stress by mobilizing jasmonoyl-isoleucine and modulating hormone-mediated signaling in wheat leaves. Beyond hormonal signaling, carotenoids play a crucial role in drought tolerance as photoprotective agents, antioxidants, and precursors for ABA biosynthesis. Carotenoid accumulation not only provides precursors for the biosynthesis of the plant hormone ABA [ 43 , 44 ], but also enhances drought tolerance by supporting ROS scavenging and improving photoprotection [ 45 ]. In this study, Rhodopinal, Astaxanthin, and Hydroxychlorobactene were detected in the MK, S, and K treatments. As a carotenoid derivative, Rhodopinal and Hydroxychlorobactene possess characteristic chromophores that assist photosynthetic pigments and scavenge free radicals [ 46 ]. Additionally, astaxanthin, a highly efficient antioxidant, was identified and may contribute to enhanced antioxidant capacity and stress adaptation [ 47 ]. These findings suggest that carotenoids, as key components of the non-enzymatic antioxidant system, mitigate drought stress by scavenging ROS modulating detoxification enzyme activity. Similarly, flavonoids, another class of secondary metabolites, play multifaceted roles in plant responses to drought stress. Flavonoids regulate hormone signal transduction and act as inhibitors of auxin transport [ 48 ]. Recent studies have shown that flavonoids, such as flavones, flavanones, flavonols, isoflavones, and anthocyanins, play an essential role as ROS scavengers, contributing significantly to oxidative stress mitigation [ 49 ]. Furthermore, flavonoids and anthocyanins enhance plant drought resistance and support growth under adverse conditions [ 50 ]. In this study, anthocyanin biosynthesis was observed in MK-treated plants, suggesting a potential role for these compounds in drought tolerance mechanisms. Notably, the expression of some flavonoids was downregulated under drought stress, indicating a dynamic regulatory adjustment. This pattern suggests that the production of flavonoid-related metabolites contributes to plant defense against drought stress and positively influences growth and performance. Plant hormones, carotenoids, and flavonoids, key regulators in plant metabolic networks, work in coordination to mediate drought stress responses. Plant hormones regulate physiological processes primarily through signal transduction pathways, while carotenoids and flavonoids, as secondary metabolites, enhance stress resistance through photoprotection, antioxidation, and regulation of hormone signaling. Collectively, these integrated processes highlight the synergistic regulatory role of primary and secondary metabolites in improving plant drought stress tolerance, orchestrating these metabolic and signaling adjustments to optimize plant responses to water deficit. Mechanism Analysis of Exogenous Sorbitol-Chelated Potassium on Biomass of Wheat Seedlings under Drought Stress Wheat drought resistance is mediated by the synergistic regulation of photosynthetic performance, antioxidant defense systems, and metabolite expression. However, most existing studies have not comprehensively integrated these key factors, including growth morphology, photosynthesis, antioxidant defense, and differential metabolites, into a unified framework. In this study, we applied correlation analysis and structural equation modeling (SEM) to investigate these interactions. Correlation analysis revealed that the exogenous application of sorbitol-chelated potassium orchestrates a cascade response by synergistically regulating N-Acetyl-D-glucosamine, (-)-Jasmonoyl-L-isoleucine, and growth morphology, and (-)-Maackiain, N-Acetyl-D-glucosamine, (-)-Jasmonoyl-L-isoleucine, 5-Methoxytryptamine, and 9,10-Dihydroxy-12,13-epoxyoctadecanoic acid were significantly correlated with wheat photosynthetic performance. The results indicated that the exogenous application of sorbitol-chelated potassium significantly mitigates the inhibitory effects of drought stress by synergistic regulation of growth morphology and photosynthetic performance of wheat seedlings. SEM analysis further revealed that growth morphology exerted the strongest total effect on wheat seedling biomass accumulation (0.685), directly dominating biomass formation. Photosynthetic performance (direct effect 0.211) and the antioxidant enzyme system (direct effect 0.306) positively regulated biomass, acting through independent pathways. Specifically, photosynthetic performance mediated an indirect effect of 0.320 through antioxidant enzymes and secondary metabolites, while the antioxidant enzyme system exerted an indirect effect of 0.351 via growth morphology (Fig. 8B). These results highlight growth morphology as the primary driver of biomass accumulation, while photosynthetic performance and antioxidant defense form a synergistic regulatory network through direct effects and multiple indirect pathways (such as secondary metabolite regulation and growth morphology feedback), jointly regulating biomass accumulation. This mechanism highlights the critical role of integrated regulation between growth morphology and physiological metabolism in driving wheat biomass accumulation. Therefore, the synergistic mechanism by which exogenous application of sorbitol-chelated potassium enhances drought resistance involves growth morphology serving as the primary driver of biomass accumulation, and photosynthetic performance, the antioxidant enzyme system, and secondary metabolites, such as (-)-jasmonoyl-L-isoleucine and Etherolenic acid, form a synergistic regulatory network that collectively alleviates the inhibitory effects of drought stress across multiple functional dimensions. This integrative framework offers a novel perspective for understanding wheat seedling growth and metabolic adaptation under water-limited conditions and provides valuable insights for improving drought resilience in wheat. Conclusion The exogenous application of sorbitol-chelated potassium (SK) alleviates drought-induced growth inhibition in wheat seedlings through a multi-faceted physiological mechanism. This mitigation is achieved by enhancing photosynthetic pigment content and gas exchange capacity, upregulating antioxidant enzyme activity, and regulating accumulation of key metabolites. Drought stress significantly limits wheat growth and biomass accumulation. Compared to CK 2 , SK application significantly mitigated biomass reduction (19.2%) under drought stress conditions. Exogenous SK significantly increased photosynthetic pigment content and gas exchange parameters in wheat flag leaves, while enhancing the activities of antioxidant enzymes and reducing malondialdehyde levels (45.66%), significantly alleviating oxidative damage caused by drought stress. Untargeted metabolomics analysis revealed that SK treatment induced the relative accumulation of differential metabolites, thereby inhibiting sugar metabolism to minimize energy loss, accumulating free amino acids to enhance osmotic adjustment, and promoting lipid metabolism to maintain cellular energy homeostasis. Additionally, SK mitigated drought-induced growth inhibition in wheat by modulating plant hormone signaling, enhancing carotenoid and flavonoid biosynthesis, and activating detoxification enzyme system to scavenge ROS. In summary, structural equation modeling (SEM) demonstrated that exogenous application of sorbitol chelated potassium, with growth morphology as the core driving factor, synergistically regulates key physiological processes, including photosynthetic performance, antioxidant defense systems, differential metabolite. This forms a coordinated multi-system network of "morphology-physiology-metabolism" to regulate drought resistance, which promotes biomass accumulation and significantly enhances the drought tolerance of wheat seedlings. Abbreviations ABA Abscisic CAT Catalase Ci Intercellular CO 2 concentration Gs Stomatal conductance MDA Malondialdehyde Pn Photosynthetic rate POD Peroxidase RWC Relative water content ROS Reactive oxygen species SOD Superoxide dismutase Tr Transpiration rate Declarations Funding This study was supported by the projects of the National Natural Science Foundation of China (31972516), the Key Research and Development Program of Shandong Province, China (2017GNC11116). Authors ’ Contributions Guohui Du conceived the research. Mingxia Zhang and Huanyang Zhang conducted the experiments, analyzed the data, and then wrote the paper. Ruili Zheng and Li Zhao revised the paper in English. Mingli Huang, Xiaocui Wang, and Kezhong Liu revised the manuscript. Yan Dongyun performed the final review. All the authors contributed to the article and approved the submitted version. Data availability Data will be made available on request. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-7498264","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":512445011,"identity":"8769114f-00a5-4d04-a799-4dd656085471","order_by":0,"name":"Mingxia Zhang","email":"","orcid":"","institution":"Qingdao University","correspondingAuthor":false,"prefix":"","firstName":"Mingxia","middleName":"","lastName":"Zhang","suffix":""},{"id":512445012,"identity":"6b18c331-7a7d-4dc7-904f-2c9a95933b2b","order_by":1,"name":"Guohui Du","email":"","orcid":"","institution":"Shandong Institute of Commerce and Technology","correspondingAuthor":false,"prefix":"","firstName":"Guohui","middleName":"","lastName":"Du","suffix":""},{"id":512445013,"identity":"56b98c63-788b-4f0f-93dc-089429555120","order_by":2,"name":"Huanyang Zhang","email":"","orcid":"","institution":"Qingdao University","correspondingAuthor":false,"prefix":"","firstName":"Huanyang","middleName":"","lastName":"Zhang","suffix":""},{"id":512445014,"identity":"1e85d9d7-a322-4def-a14b-b48ea5359d96","order_by":3,"name":"Ruili Zheng","email":"","orcid":"","institution":"Qingdao University","correspondingAuthor":false,"prefix":"","firstName":"Ruili","middleName":"","lastName":"Zheng","suffix":""},{"id":512445015,"identity":"28bfdca3-202a-4c1a-9b1d-83377dfc5efc","order_by":4,"name":"Li Zhao","email":"","orcid":"","institution":"Qingdao University","correspondingAuthor":false,"prefix":"","firstName":"Li","middleName":"","lastName":"Zhao","suffix":""},{"id":512445016,"identity":"7c2083eb-69f6-46db-842e-095f06f5c21c","order_by":5,"name":"Mingli Huang","email":"","orcid":"","institution":"Qingdao University","correspondingAuthor":false,"prefix":"","firstName":"Mingli","middleName":"","lastName":"Huang","suffix":""},{"id":512445017,"identity":"6495000c-8ab6-4cc7-961c-e541f135e2f8","order_by":6,"name":"Xiaocui Wang","email":"","orcid":"","institution":"Qingdao University","correspondingAuthor":false,"prefix":"","firstName":"Xiaocui","middleName":"","lastName":"Wang","suffix":""},{"id":512445018,"identity":"01343894-57da-4b94-bf12-8d0a18389464","order_by":7,"name":"Kezhong Liu","email":"","orcid":"","institution":"Shandong Agricultural University","correspondingAuthor":false,"prefix":"","firstName":"Kezhong","middleName":"","lastName":"Liu","suffix":""},{"id":512445019,"identity":"022ebecc-c3a8-4aa5-95bc-dc8be7a052f7","order_by":8,"name":"Dongyun Yan","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAABAklEQVRIiWNgGAWjYJACCQjFfPDBh4oDYOaBB8RpYUs2nHHmAAMPSEsCcVp41IQ52yBaGPBpMTh+9uCNjztqEzfc7mFjZpx3R85e7PBDoC12croNOLScyUu2nHnmeOKGO2ePPS7c9syYRzrNAKgl2djsAA4tB3LMpHnbjiVuuJGXbjxz2+HEHukEkJYDidtwaTn/BqYFpHcOSEv6B/xawCrbaqBaGkBacvDbInnjjbHlzLYDxjNvpAED+dhhY57bOQUHEgxw+4XvfI7hjY9tdbJ9N5KBUVlzWI59dvrmDx8q7ORwaVGAiB92bEBzMHblICAPUVpnj1vJKBgFo2AUjHgAAD8WcABdg86UAAAAAElFTkSuQmCC","orcid":"","institution":"Qingdao University","correspondingAuthor":true,"prefix":"","firstName":"Dongyun","middleName":"","lastName":"Yan","suffix":""}],"badges":[],"createdAt":"2025-08-31 04:08:02","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-7498264/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-7498264/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":91178892,"identity":"b5c42094-1d3f-4dd5-897c-0fa52d012fc1","added_by":"auto","created_at":"2025-09-12 12:28:56","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":106360,"visible":true,"origin":"","legend":"\u003cp\u003eChanges in photosynthetic pigment content of wheat seedlings. (A) changes in chlorophyll a content. (B) changes in chlorophyll b content. (C) changes in carotenoid content. (D) changes in total chlorophyll content. Note: Values are expressed as mean ± standard deviation (SD) (n=3, biological independent replicate experiments). The p-values were determined using one-way ANOVA and Brown-Forsythe test（*\u003cem\u003ep\u003c/em\u003e \u0026lt;0.05;**\u003cem\u003ep\u003c/em\u003e\u0026lt;0.01;***\u003cem\u003ep\u003c/em\u003e \u0026lt;0.001）.\u003c/p\u003e","description":"","filename":"1.png","url":"https://assets-eu.researchsquare.com/files/rs-7498264/v1/a17b01be547a86a4fccb9488.png"},{"id":91179181,"identity":"d40db836-ba0f-49e2-86cb-44dfc3287b4e","added_by":"auto","created_at":"2025-09-12 12:36:55","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":118692,"visible":true,"origin":"","legend":"\u003cp\u003eChanges in gas exchange of wheat seedling leaves. (A) Net photosynthetic rate (Pn). (B) Transpiration rate (Tr). (C) Stomatal conductance (Gs). (D) Intercellular CO\u003csub\u003e2\u003c/sub\u003e concentration (Ci).\u003c/p\u003e","description":"","filename":"2.png","url":"https://assets-eu.researchsquare.com/files/rs-7498264/v1/b8ad51d3b1a5185e5fb67147.png"},{"id":91179196,"identity":"641f6584-ff2d-4075-8677-c864d48a5fd7","added_by":"auto","created_at":"2025-09-12 12:36:58","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":108189,"visible":true,"origin":"","legend":"\u003cp\u003eAntioxidant enzyme activity and malondialdehyde content in wheat seedlings. (A) Superoxide Dismutase (SOD). (B) Peroxidase (POD). (C) Catalase (CAT). (D) Malondialdehyde (MDA).\u003c/p\u003e","description":"","filename":"3.png","url":"https://assets-eu.researchsquare.com/files/rs-7498264/v1/13d9023b29b3c2791c4674be.png"},{"id":91180131,"identity":"558af967-52a8-4047-92d9-9db026f12e38","added_by":"auto","created_at":"2025-09-12 12:44:56","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":112075,"visible":true,"origin":"","legend":"\u003cp\u003eThe PCA results of metabolites in wheat leaves. (A) Positive ion mode. (B) Negative ion mode.\u003c/p\u003e","description":"","filename":"4.png","url":"https://assets-eu.researchsquare.com/files/rs-7498264/v1/1fcf2df6421d1a8048b015c2.png"},{"id":91178925,"identity":"69009ac2-fcdf-4dca-8867-90633ca9b4a1","added_by":"auto","created_at":"2025-09-12 12:29:01","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":839543,"visible":true,"origin":"","legend":"\u003cp\u003eHeat map analysis of metabolites identified under drought stress compared with CK\u003csub\u003e2\u003c/sub\u003e. (A)SK.VS.CK\u003csub\u003e2\u003c/sub\u003e; (B) K.VS.CK\u003csub\u003e2\u003c/sub\u003e; (C) MK.VS.CK\u003csub\u003e2\u003c/sub\u003e; (D) S.VS.CK\u003csub\u003e2\u003c/sub\u003e.\u003c/p\u003e","description":"","filename":"5.png","url":"https://assets-eu.researchsquare.com/files/rs-7498264/v1/5294fa8ddf162e28d643316c.png"},{"id":91178888,"identity":"1740115f-c3bf-4681-9861-59a6d392b698","added_by":"auto","created_at":"2025-09-12 12:28:56","extension":"png","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":197794,"visible":true,"origin":"","legend":"\u003cp\u003eKEGG analysis of major metabolic pathways under drought stress compared to CK\u003csub\u003e2\u003c/sub\u003e. The size and shade of the circles represent the number of differentially expressed metabolites and P-values, respectively. （A） SK.VS.CK2;（B） K.VS.CK2;（C） MK.VS.CK2;（D） S.VS.CK\u003csub\u003e2\u003c/sub\u003e.\u003c/p\u003e","description":"","filename":"6.png","url":"https://assets-eu.researchsquare.com/files/rs-7498264/v1/a2b22a7511044f295c6f103e.png"},{"id":91178887,"identity":"37e5520f-7164-41da-a3c0-c78ca64becb2","added_by":"auto","created_at":"2025-09-12 12:28:56","extension":"png","order_by":7,"title":"Figure 7","display":"","copyAsset":false,"role":"figure","size":505525,"visible":true,"origin":"","legend":"\u003cp\u003eHeatmap of growth morphology, photosynthetic performance, and correlations between antioxidant enzymes and differential metabolites in wheat.\u003c/p\u003e","description":"","filename":"7.png","url":"https://assets-eu.researchsquare.com/files/rs-7498264/v1/8b6f59c57d730d90ac26a7cb.png"},{"id":91180130,"identity":"8b305749-ad50-409b-8ad1-032342189494","added_by":"auto","created_at":"2025-09-12 12:44:56","extension":"png","order_by":8,"title":"Figure 8","display":"","copyAsset":false,"role":"figure","size":153969,"visible":true,"origin":"","legend":"\u003cp\u003e(A) SEM relationship between biomass and wheat plant, photosynthetic performance, antioxidant enzymes and metabolites. (B) SEM standardized effect values of biomass and wheat indexes. Note: *, **, *** indicate significant difference at the level of 0.05, 0.01, 0.001, respectively.\u003c/p\u003e","description":"","filename":"8.png","url":"https://assets-eu.researchsquare.com/files/rs-7498264/v1/09dfd5a13739685aa093c48b.png"},{"id":92876054,"identity":"d4b66094-9d6b-499e-ba5b-05553356099b","added_by":"auto","created_at":"2025-10-06 14:39:08","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":2717978,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-7498264/v1/2f7ae9ba-3429-4343-9550-08b07f78e540.pdf"},{"id":91180132,"identity":"ed6db53c-79fc-48c1-bee5-d2e6d8dee556","added_by":"auto","created_at":"2025-09-12 12:44:56","extension":"docx","order_by":0,"title":"","display":"","copyAsset":false,"role":"supplement","size":155006,"visible":true,"origin":"","legend":"","description":"","filename":"SupplementaryMaterial.docx","url":"https://assets-eu.researchsquare.com/files/rs-7498264/v1/5c1e8a5c2714f35b72a9b391.docx"},{"id":91179182,"identity":"21dc0b10-6cff-4906-a1b5-63e40494ddb4","added_by":"auto","created_at":"2025-09-12 12:36:56","extension":"xlsx","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":42648,"visible":true,"origin":"","legend":"","description":"","filename":"SupplementaryMaterial.xlsx","url":"https://assets-eu.researchsquare.com/files/rs-7498264/v1/155f9b51512de57ef75ec85a.xlsx"}],"financialInterests":"No competing interests reported.","formattedTitle":"Physiological and Metabolomic Analyses of Exogenously Applied Sorbitol-Chelated Potassium Enhancing Drought Tolerance in Wheat","fulltext":[{"header":"Introduction","content":"\u003cp\u003eDrought stress disrupts key physiological processes in plants, impairing photosynthesis and antioxidant defenses [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e], and is a major factor limiting wheat growth and yield [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e]. As an early response to water deficit, photosynthesis is notably vulnerable, with drought-induced damage to photosynthetic pigments, reductions in net photosynthetic rate, stomatal conductance, and transpiration rate, these ultimately disrupt plant metabolism and suppress biomass accumulation [\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e, \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e]. In parallel, drought stress induces overproduction of reactive oxygen species (ROS), causing oxidative damage to plant cells and tissues. To mitigate the oxidative damage of ROS to cells and maintain redox balance, plants activate enzymatic antioxidant systems, which are composed of peroxidase (POD), superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPX), and non-enzymatic antioxidant systems, including secondary metabolites such as flavonoids (FC) and total phenols (TPC) compounds [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e, \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e].\u003c/p\u003e\u003cp\u003ePlant responses to drought by profiling metabolite changes. Serving as a bridge between plant genotype and phenotypes, the metabolome provides insights into mechanisms underlying phenotypic variation under stress conditions [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e]. Wang et al. [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e] using LC -MS technology, four key types of differential metabolites, namely amino acids, organic acids, sugars, and alkaloids, such as gibberellin A4 (GA4), abscisic acid (ABA), and sucrose, have been identified. These metabolites are crucial for alfalfa to withstand drought stress. Bowne et al. [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e] investigated a targeted GC-MS approach to monitor 103 structurally identified metabolites from leaf tissue of the drought-stressed wheat plants, predominantly amino and organic acids and sugars. However, most drought-stress studies have focused on individual plant systems, leaving the interactions among photosynthetic, antioxidant, and metabolomic responses underexplored.\u003c/p\u003e\u003cp\u003eCritical for maintaining growth and yield stability. Potassium, a key macronutrient, plays a pivotal role in stomatal regulation, photosynthesis, ROS detoxification, and stress resistance [\u003cspan additionalcitationids=\"CR11\" citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e]. Drought impairs root growth and limits potassium from the soil [\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e], making exogenous potassium application an effective strategy to compensate for the potassium deficiency and enhance drought tolerance. Sorbitol-chelated potassium, a novel chelated foliar potassium fertilizer, has demonstrated a positive regulatory role in improving wheat drought resilience. Compared to conventional inorganic (non-chelated) potassium fertilizers, sorbitol-chelated potassium enhances potassium uptake, stress resistance, and crop yield. In wheat, sorbitol-chelated potassium application has been reported to increase post-anthesis potassium accumulation and yield by 13.99%-35.48% [\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e]. Similar benefits have been observed in other crops, including enhanced dry matter accumulation and kernel potassium content in peanuts (average yield increase of 18.9%) [\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e] and improved growth, potassium uptake, and quality in celery [16.17].\u003c/p\u003e\u003cp\u003eDespite these findings, studies investigating the effects of chelated potassium on crops under drought stress remain limited [\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e]. Given the role of potassium in wheat growth and stress responses, elucidating how sorbitol-chelated potassium enhances drought resistance holds significant scientific and practical implications. Because drought affects multiple physiological pathways, including photosynthesis, antioxidant defense, and metabolism, a systematic analysis integrating these factors is necessary to uncover potential mechanisms by which sorbitol-chelated potassium alleviates drought-induced damage in wheat.\u003c/p\u003e"},{"header":"Materials and methods","content":"\u003cp\u003ePlant materials and seed preparation\u003c/p\u003e\u003cp\u003ePlump XinHua818 wheat seeds of uniform size were selected and surface sterilized with 0.1% sodium hypochlorite for 10 minutes, followed by three rinses with ultrapure water. The sterilized seeds were incubated in the dark at 25℃ for 24 hours in a biological incubator to promote germination.\u003c/p\u003e\u003cp\u003eSeedling cultivation and hydroponic growth conditions\u003c/p\u003e\u003cp\u003eGerminated seeds were placed into seedling trays lined with three layers of gauze and grown in a greenhouse under controlled conditions: temperature 25℃, light intensity 5000lx, and a photoperiod of 16 h light/8 h dark. Once the seedlings reached 5\u0026ndash;6 cm in height, they were transferred to plastic hydroponic containers (70 seedlings per container) containing Hoagland nutrient solution. The solution volume in each container was maintained consistently and replaced every 3 days.\u003c/p\u003e\u003cp\u003eExperimental design and drought treatment\u003c/p\u003e\u003cp\u003eAt the two-leaf and one-heart stage, seedlings were transferred to hydroponic containers (8.5 cm diameter, 12 cm height) to commence experimental treatments. Six treatments were established, each with three biological replicates (48 seedlings per replicate). Drought stress was simulated using 10% polyethylene glycol (PEG-6000) in the nutrient solution for 7 days. Foliar sprays corresponding to each treatment were applied on days 1 and 3 (twice in total) of drought period. After 7 days of drought stress, seedling samples were collected and immediately frozen in liquid nitrogen for subsequent analyses. The normal control (CK\u003csub\u003e1\u003c/sub\u003e) was treated with Hoagland nutrient solution, while the drought-stressed control (CK\u003csub\u003e2\u003c/sub\u003e) was treated with Hoagland nutrient solution supplemented with 10% PEG-6000. The experimental design is detailed in Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e.\u003c/p\u003e\u003cp\u003e\u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e\u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e\u003cdiv class=\"CaptionContent\"\u003e\u003cp\u003eExperimental setup with different foliar spray treatments\u003c/p\u003e\u003c/div\u003e\u003c/caption\u003e\u003ccolgroup cols=\"4\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u003cp\u003eTreatment\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u003cp\u003eGrowth environment\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colspan=\"2\" nameend=\"c4\" namest=\"c3\"\u003e\u003cp\u003eSpraying concentration (mg\u0026middot;L\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e)\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eControl (CK\u003csub\u003e1\u003c/sub\u003e)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eHoagland nutrient solution\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e-\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colspan=\"1\" nameend=\"c4\" namest=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eControl (CK\u003csub\u003e2\u003c/sub\u003e)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\" morerows=\"4\" rowspan=\"5\"\u003e\u003cp\u003eHoagland nutrient solution\u0026thinsp;+\u0026thinsp;10%PEG-6000\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\" morerows=\"4\" rowspan=\"5\"\u003e\u003cp\u003eDistilled water\u003c/p\u003e\u003cp\u003e10 (K\u003csup\u003e+\u003c/sup\u003e)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colspan=\"1\" nameend=\"c4\" namest=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eSorbitol (S)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colspan=\"1\" nameend=\"c4\" namest=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003ePotassium Chloride (K)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colspan=\"1\" nameend=\"c4\" namest=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003ePotassium Mixed with Sorbitol (MK)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colspan=\"1\" nameend=\"c4\" namest=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eSorbitol-chelated Potassium (SK)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colspan=\"1\" nameend=\"c4\" namest=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/colgroup\u003e\u003c/table\u003e\u003c/div\u003e\u003c/p\u003e\u003cp\u003eDetermination of growth indicators and leaf relative water content\u003c/p\u003e\u003cp\u003eFor each treatment, twelve wheat plants were selected, washed with pure water, and wiped dry before measuring plant height and stem diameter. Each whole plant was then divided into above-ground and below-ground parts, and immediately weighed, with the weights recorded as root fresh weight and leaf fresh weight (FW). The leaves were soaked in double-distilled water for 24 h and then removed. After removing residual water from the leaf surface with absorbent paper, the leaves were weighed and the weight recorded as turgid weight (TW). The leaves were then deactivated at 105\u0026deg;C for 30 min and dried at 60\u0026deg;C to a constant weight, which was recorded as dry weight (DW). The relative water content of the leaves was calculated via the following formula: \u003cem\u003eRWC (%) =(FW-DW)/(TW-DW) \u0026times;100%\u003c/em\u003e [\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e]. Three biological replicates were analysed.\u003c/p\u003e\u003cp\u003eDetermination of chlorophyll content and gas exchange\u003c/p\u003e\u003cp\u003eFollowing the methods of Lichtenthaler [\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e], with slight modifications, the first fully expanded leaf of wheat seedlings was selected, ground with a mortar, and extracted with acetone. The absorbance of chlorophyll \u003cem\u003ea\u003c/em\u003e, chlorophyll \u003cem\u003eb\u003c/em\u003e and carotenoids were measured at OD\u003csub\u003e663\u003c/sub\u003e, OD\u003csub\u003e645\u003c/sub\u003e and OD\u003csub\u003e480\u003c/sub\u003e nm using a spectrophotometer.\u003c/p\u003e\u003cp\u003eFollowing drought stress, the net photosynthetic rate (Pn), stomatal conductance (Gs), transpiration rate (Tr), and intercellular CO\u003csub\u003e2\u003c/sub\u003e concentration (Ci) were determined using an Li-6800 portable photosynthesis system (Li-CORBiosciences, Lincoln, NE, USA), as previously described by He et al. [\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e]. The measurements were conducted on the last fully expanded leaf, employing a photon flux density of 1000 \u0026micro;mol photo m\u003csup\u003e\u0026minus;\u0026thinsp;2\u003c/sup\u003e s\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e and a CO\u003csub\u003e2\u003c/sub\u003e concentration of 400 \u0026micro;mol mol\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e, within the time from of 9:00 a.m. to 11:00 a.m.\u003c/p\u003e\u003cp\u003eAntioxidant enzyme activity and malondialdehyde content\u003c/p\u003e\u003cp\u003eThe activities of superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) were determined using assay kits (Boxbio, Beijing) according to the manufacturer\u0026rsquo;s instructions. One gram of leaf sample was homogenized in 1 mL of extraction buffer in an ice bath, followed by centrifugation at 8,000 g for 10 min at 4\u0026deg;C. The supernatant was then collected and used for enzyme activity assays.\u003c/p\u003e\u003cp\u003eFollowing the method outlined by Tan et al. [\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e], a 1 g sample was homogenized with 10 mL of a mixed TCA-TBA solution, followed by centrifugation at 4000 rpm for 10 min. The mixture was kept in a boiling water for 15 minutes and centrifuged at 4,000 g for 10 min. Malondialdehyde (MDA) content in the reaction solution was determined using a spectrophotometer at OD\u003csub\u003e450\u003c/sub\u003e, OD\u003csub\u003e532\u003c/sub\u003e and OD\u003csub\u003e600\u003c/sub\u003e nm.\u003c/p\u003e\u003cp\u003eMetabolite extraction\u003c/p\u003e\u003cp\u003eSix wheat seedlings were randomly selected for each treatment group. Leaves from the same position on each seedling were collected, immediately flash-frozen in liquid nitrogen, and then stored in a -40 ℃ freezer. The metabolite extraction steps were as follows:\u003c/p\u003e\u003cp\u003e(1) Weigh 50 mg of the sample into a pre-cooled mortar and grind it into powder. The powder is then transferred to an EP tube, and 1000 \u0026micro;L of a mixed extraction solution of methanol, acetonitrile, and water (2:2:1, containing internal standard at 20 mg L\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e) is added, followed by vortexing for 30 s.\u003c/p\u003e\u003cp\u003e(2) Steel beads are added, and the mixture is processed using a grinding instrument (45 Hz) for 10 min, followed by sonication in an ice-water bath for 10 min. Subsequently, the sample is allowed to stand at -20\u0026deg;C for 1 h, and then centrifuged at 12000 rpm (4\u0026deg;C, 15 min).\u003c/p\u003e\u003cp\u003e(3) 500 \u0026micro;L of the supernatant is transferred to an EP tube and dried in a vacuum concentrator. The extract is then reconstituted with 160 \u0026micro;L of acetonitrile/water (1:1, v/v), vortexed for 30 s, and sonicated in an ice-water bath for 10 min.\u003c/p\u003e\u003cp\u003e(4) Collect 120 \u0026micro;L of the supernatant (12000 rpm, 4\u0026deg;C, 15 min) into a 2 mL vial. 10 \u0026micro;L from each sample is then pooled to create a QC sample for on-machine analysis.\u003c/p\u003e\u003cp\u003eChromatographic conditions: An Acquity UPLC HSS T3 column (1.8 \u0026micro;m, 2.1*100 mm, Waters, US) was used with mobile phase A (0.1% formic acid in water) and mobile phase B (0.1% formic acid in acetonitrile). Chromatographic data acquisition was performed on an Acquity I-Class PLUS ultra-high-performance liquid chromatograph (Waters, US) coupled to a Xevo G2-XS QTOF (Waters, US) high-resolution mass spectrometer, with detection in both positive and negative ion modes.\u003c/p\u003e\u003cp\u003eStatistic analysis\u003c/p\u003e\u003cp\u003eNormality and log-normality tests, along with one-way ANOVA, were conducted using SPSS 27.0 and GraphPad Prism 10.1.2 to determine differences between numerical values. Structural equation modeling (SEM) data processing was performed using SetupStata 18, and Power Point 2021 and Hiplot (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://hiplot.cn/basic/ggscatterstats\u003c/span\u003e\u003cspan address=\"https://hiplot.cn/basic/ggscatterstats\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e) were used to generate figures. Raw metabolomics data were collected using MassLynx V4.2 software, and Progenesis QI software was used for data processing operations such as mass peak extraction and alignment [\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e]. PCA and heatmap images were generated using MetaboAnalyst 6.0 (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://www.metaboanalyst.ca/\u003c/span\u003e\u003cspan address=\"https://www.metaboanalyst.ca/\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e) and Origin 2021. Commercial databases, including the online METLIN database (based on Progenesis QI software), KEGG (Kyoto Encyclopedia of Genes and Genomes) (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttp://www.genome.jp/kegg/\u003c/span\u003e\u003cspan address=\"http://www.genome.jp/kegg/\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e), HMDB (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://hmdb.ca\u003c/span\u003e\u003cspan address=\"https://hmdb.ca\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e), and Lipidmaps (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttp://www.lipidmaps.org/\u003c/span\u003e\u003cspan address=\"http://www.lipidmaps.org/\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e), were used to search metabolic pathways for profiling and metabolite identification.\u003c/p\u003e"},{"header":"Results","content":"\u003cp\u003eEffect of sorbitol-chelated potassium on wheat growth and biochemical indices under drought stress\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eGrowth traits\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eDifferent foliar spray treatments significantly affected the wheat seedling growth traits, including biomass, plant height, stem diameter, and leaf relative water content (RWC) (Table2). Notably, sorbitol-chelated potassium treatment effectively promoted biomass accumulation compared to other drought stress treatments. Under drought stress, aboveground biomass in SK-treated seedlings increased by 15.66% and 20.00% compared to K and MK treatments, respectively (both significant). Root biomass also increased under SK treatment by 3.23% and 10.34% compared with K and MK\u0026nbsp;treatments, respectively, though the difference was not significant relative to K treatment. Plant height increased by 8.42% under SK compared to K treatment, whereas stem diameter did not differ significantly across treatments. Additionally, SK\u0026nbsp;treatment increased leaf RWC by 15.25% compared to K\u0026nbsp;treatment, indicating enhanced water retention under drought stress.\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 2\u003c/strong\u003e Growth indicators of wheat seedlings under normal conditions and drought stress with different fertilization treatments\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\" width=\"586\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eTreatment\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 99px;\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003eAboveground biomass\u003c/strong\u003e\u003cstrong\u003e(g)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 90px;\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003eRoot biomass(g)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 99px;\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003ePlant height\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003e(cm)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 105px;\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003eStem diameter (mm)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 113px;\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003eRelative water content of leaves\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eCK\u003csub\u003e1\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 99px;\"\u003e\n \u003cp\u003e3.04\u0026plusmn;0.09a\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 90px;\"\u003e\n \u003cp\u003e0.31\u0026plusmn;0.00ab\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 99px;\"\u003e\n \u003cp\u003e23.46\u0026plusmn;0.98a\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 105px;\"\u003e\n \u003cp\u003e2.46\u0026plusmn;0.04a\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 113px;\"\u003e\n \u003cp\u003e0.88\u0026plusmn;0.03a\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eCK\u003csub\u003e2\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 99px;\"\u003e\n \u003cp\u003e2.42\u0026plusmn;0.03b\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 90px;\"\u003e\n \u003cp\u003e0.28\u0026plusmn;0.01c\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 99px;\"\u003e\n \u003cp\u003e22.23\u0026plusmn;0.03ab\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 105px;\"\u003e\n \u003cp\u003e2.27\u0026plusmn;0.04c\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 113px;\"\u003e\n \u003cp\u003e0.56\u0026plusmn;0.03cd\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eS\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 99px;\"\u003e\n \u003cp\u003e2.47\u0026plusmn;0.10b\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 90px;\"\u003e\n \u003cp\u003e0.28\u0026plusmn;0.01c\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 99px;\"\u003e\n \u003cp\u003e22.29\u0026plusmn;0.24ab\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 105px;\"\u003e\n \u003cp\u003e2.44\u0026plusmn;0.03ab\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 113px;\"\u003e\n \u003cp\u003e0.47\u0026plusmn;0.01d\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eK\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 99px;\"\u003e\n \u003cp\u003e2.49\u0026plusmn;0.08b\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 90px;\"\u003e\n \u003cp\u003e0.31\u0026plusmn;0.02ab\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 99px;\"\u003e\n \u003cp\u003e21.37\u0026plusmn;0.26b\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 105px;\"\u003e\n \u003cp\u003e2.40\u0026plusmn;0.00ab\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 113px;\"\u003e\n \u003cp\u003e0.59\u0026plusmn;0.06bc\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eMK\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 99px;\"\u003e\n \u003cp\u003e2.40\u0026plusmn;0.08b\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 90px;\"\u003e\n \u003cp\u003e0.29\u0026plusmn;0.00bc\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 99px;\"\u003e\n \u003cp\u003e22.42\u0026plusmn;0.16ab\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 105px;\"\u003e\n \u003cp\u003e2.39\u0026plusmn;0.02b\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 113px;\"\u003e\n \u003cp\u003e0.63\u0026plusmn;0.06bc\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eSK\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 99px;\"\u003e\n \u003cp\u003e2.88\u0026plusmn;0.06a\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 90px;\"\u003e\n \u003cp\u003e0.32\u0026plusmn;0.01a\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 99px;\"\u003e\n \u003cp\u003e23.17\u0026plusmn;0.80a\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 105px;\"\u003e\n \u003cp\u003e2.40\u0026plusmn;0.03ab\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 113px;\"\u003e\n \u003cp\u003e0.68\u0026plusmn;0.05b\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003eNote: Different lowercase letters in the same column indicate significant differences between treatment groups for the same indicator (\u003cem\u003ep\u003c/em\u003e\u0026lt;0.05). (Above ground and root biomass measured as fresh weight)\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cem\u003ePhotosynthetic pigments and gas exchange\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eDifferent foliar spray treatments significantly affected the accumulation of chlorophyll \u003cem\u003ea\u003c/em\u003e, chlorophyll \u003cem\u003eb\u003c/em\u003e, carotenoids, and total chlorophyll content in wheat seedlings under drought stress (Fig. 1). Drought stress reduced photosynthetic pigment content, whereas foliar spraying alleviated this effect. Compared with the ionic potassium treatment, the sorbitol-chelated potassium treatment effectively increased chlorophyll \u003cem\u003ea\u003c/em\u003e and total chlorophyll content, with significant differences. Chlorophyll \u003cem\u003ea\u0026nbsp;\u003c/em\u003eincreased by 11.60% and 19.54% in SK treatment compared to K and MK treatments, respectively. Similarly, chlorophyll \u003cem\u003eb\u003c/em\u003e and carotenoid content increased by 15.94% and 10.68%, respectively, compared with MK treatment. Carotenoid levels in SK treatment decreased by 1.52% and 4.11% relative to K and S treatments, respectively, though these differences were not statistically significant. Total chlorophyll content in SK treatment increased by 18.74% compared to MK treatment, a difference that was statistically significant.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eUnder drought stress, different spraying treatments significantly affected the net photosynthetic rate (Pn), transpiration rate (Tr), intercellular CO\u003csub\u003e2\u003c/sub\u003e concentration (Ci), and stomatal conductance (Gs) of wheat seedlings (Fig. 2). Drought stress decreased leaf gas exchange parameters relative to the control treatment (CK\u003csub\u003e1\u003c/sub\u003e). However, these parameters improved following foliar treatments. Compared with the MK, K, and S treatments, SK treatment significantly increased Pn by 62.90%, 31.98%, and 65.00%, respectively, and Ci by 14.55%, 23.28%, and 14.00%, respectively. Tr decreased by 18.64% and 25.00% under SK treatment compared to K and S treatments, respectively. However, these differences were not statistically significant. Gs decreased by 15.39% and 26.75% compared to MK and S treatments, respectively, with no significant differences. \u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eAntioxidant System\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eUnder drought stress, the levels of superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT), as well as malondialdehyde (MDA) content in wheat seedlings, were significantly affected by different foliar spray treatments (Fig. 3). Drought stress significantly reduced SOD and POD activities. However, foliar application with sorbitol-chelated potassium (SK), a mixture of sorbitol and potassium chloride (MK), sorbitol (S), potassium chloride (K) significantly increased SOD and POD activity compared to control treatment (CK\u003csub\u003e2\u003c/sub\u003e). SOD activity increased by 46.07% and 19.58% in SK treatment relative to MK and S treatments, respectively. CAT activity increased by 64.00% in SK treatment compared to S treatment. POD activity followed a similar trend, increasing by 7.23% and 19.46% in SK compared to K and MK treatments, respectively, but decreasing by 19.92% compared to S treatment. Drought stress induced the accumulation of malondialdehyde (MDA) levels, enhancing membrane lipid peroxidation and causing damage to wheat seedlings (Fig. 3D). The MDA levels was decreased by 16.02% in SK treatment compared to MK treatment.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eMetabolomic profiles of wheat under drought stress\u003c/p\u003e\n\u003cp\u003e\u003cem\u003ePrincipal component analysis (PCA)\u0026nbsp;\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003ePCA was performed to compare metabolic differences among the treatment groups (SK, K, MK, S, and CK\u003csub\u003e2\u003c/sub\u003e) and assess the overall metabolomic changes in wheat seedling leaves under drought stress. Additionally, the repeatability and stability of metabolites within each treatment group were evaluated. Figure 4A and B presents the PCA results for positive and negative ion modes. The total explained variances were 51.5% and 44.4%, respectively, indicating satisfactory clustering of samples. In both modes, the metabolite profiles of potassium chloride (K) and sorbitol-chelated potassium (SK) treatments were clearly separated, suggesting distinct metabolic responses. While partial overlap was observed between chelated potassium (SK) and non-chelated potassium (MK) treatments, overall significant differences persisted, indicating that the three potassium formulations exerted differential effects on the metabolism of wheat seedling leaves. These metabolic distinctions were consistent with the variations in agronomic traits, physiological parameters, and biochemical indices described as described above.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eIdentification of differentially expressed metabolites\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eA total of 418 differentially expressed metabolites (DEMs) were identified from 1788 metabolites (Supplemental Material 1). Compared with CK\u003csub\u003e2\u003c/sub\u003e, SK showed 11 DEMs upregulated and 40 downregulated; K had 30 upregulated and 17 downregulated; MK had 28 upregulated and 231 downregulated; and S presented 27 upregulated and 34 downregulated (Fig. 5). It was found that the number of downregulated DEMs was greater than that of upregulated DEMs in response to drought stress when each treatment was compared with CK\u003csub\u003e2\u003c/sub\u003e. To clarify the characteristics of DEM expression in sorbitol-chelated potassium, upregulated metabolites (FC\u0026ge;2) in SK.VS.CK\u003csub\u003e2\u003c/sub\u003e were found to include Prolyl-Histidine, Lariciresinol, Chelirubine, L-Sepiapterin, 3,5-Diprenyl-4-hydroxybenzaldehyde,, Etherolenic acid and 4-Amino-4-deoxychorismate, while downregulated metabolites (FC\u0026le;0.5) included (-)-Jasmonoyl-L-isoleucine, N-Acetyl-D-glucosamine, (+)-Abscisic acid, 5-Hydroxypseudobaptigenin, and D-Glucosaminate-6-phosphate.\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eFunctional annotation and enrichment analysis of differential metabolites based on KEGG\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eTo identify the metabolic pathways involved, DEMs from different comparison groups were matched against the KEGG database to obtain information on as many metabolites as possible in each extract (Supplementary Material 2, Fig. S1). Enrichment analysis was then performed on the annotated results based on the P-values of the metabolic pathways, revealing a total of 119 metabolic pathways in both positive and negative ion modes. Compared with CK\u003csub\u003e2\u0026nbsp;\u003c/sub\u003etreatment, 17, 16, 25, and 109 DEMs were identified for sorbitol-chelated potassium, potassium, sorbitol-mixed potassium, and sorbitol, respectively, and subsequently annotated to 18, 16, 24, and 61 metabolic pathways, respectively (Supplementary Material 3).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThe top ten most significantly enriched metabolic pathways of differential metabolites were selected (Fig. 6). Compared to CK\u003csub\u003e2\u003c/sub\u003e, foliar spraying with SK, K, MK, and S activated a broad range of metabolites, particularly those involved \u0026quot;antioxidation\u0026quot; and regulating \u0026quot;osmotic potential\u0026quot; [24].\u0026nbsp;These findings indicate that foliar application of osmoprotectants mitigates drought-induced damage in wheat seedlings. In contrast, the SK, K, MK, and S mobilized the most diverse set of metabolites. Specifically, in SK.VS.CK\u003csub\u003e2\u003c/sub\u003e, upregulated metabolites were primarily enriched in metabolic pathways such as alpha-Linolenic acid metabolism, Folate biosynthesis, and Isoquinoline alkaloid biosynthesis, while downregulated metabolites were mainly enriched in pathways such as the Pentose phosphate pathway, Flavonoid biosynthesis, and Plant hormone signal transduction. Metabolic pathway enrichment analysis revealed that SK application modulated key metabolic pathways associated with intracellular water balance, ROS scavenging, and overall seedling vigor, thereby improving drought tolerance in wheat.\u003c/p\u003e\n\u003cp\u003eThe correlation of wheat growth with physiological and biochemical indices, and differential metabolites\u003c/p\u003e\n\u003cp\u003eDifferential metabolites in wheat exhibited a close relationship with growth morphology, photosynthetic performance, and antioxidant enzymes (Fig. 7). Correlation analysis revealed that N-Acetyl-D-glucosamine and (-)-Jasmonoyl-L-isoleucine were extremely significantly positively correlated with wheat growth indicators, while 5-Methoxytryptamine showed a significant positive correlation. Furthermore, (-)-Jasmonoyl-L-isoleucine and 5-Methoxytryptamine were extremely significantly positively correlated with wheat photosynthetic performance, and 9,10-Dihydroxy-12,13-epoxyoctadecanoic acid, (-)-Maackiain, and N-Acetyl-D-glucosamine were significantly positively correlated. Etherolenic acid was significantly positively correlated with antioxidant enzymes, suggesting that these metabolites interact within the plant metabolic network to cooperatively maintain normal plant physiological functions under drought stress. Additionally, 9,10-Dihydroxy-12,13-epoxyoctadecanoic acid and (-)-Maackiain were positively correlated with wheat growth morphology; D-Glucosaminate-6-phosphate, Betaine, Rhodopinal, GibberellinA4, and Hydroxychlorobactene were positively correlated with wheat photosynthetic performance; Chlorophylla, Rhodopinal, 5-Methoxytryptamine, Hydroxychlorobactene, Astaxanthin, (-)-Maackiain, and GibberellinA4 were positively correlated with the antioxidant system.\u003c/p\u003e\n\u003cp\u003eUnder drought stress, foliar spraying induced the accumulation of sugars, amino acids, lipids, plant hormones, carotenoids, and flavonoids, which collectively regulated wheat growth, physiology, and biochemical responses. To further elucidate the mechanisms underlying these effects, structural equation modeling (SEM) (Fig. 8A) was employed to quantify the direct and indirect influences of five factors, growth morphology, photosynthetic performance, antioxidant enzyme activity, primary metabolites, and secondary metabolites, on biomass accumulation. SEM results indicated a good model fit, with the model \u0026chi;\u0026sup2; value at 77.01, RMSEA at 0.00, CFI at 1.00, and AIC at 324.68, all fitting indices being within the standard range. SEM findings revealed that all five factors had either direct or indirect effects on biomass accumulation. Growth morphology, photosynthetic performance, antioxidant enzymes, and secondary metabolites exerted direct positive effects on biomass (direct effect value: 0.665, 0.211, 0.306, and 0.353), with growth morphology showing the strongest most significant influence. In contrast, primary metabolites exerted a direct negative effect on wheat biomass (-0.620, nonsignificant), but their indirect effect on biomass accumulation was substantial, primarily mediated through their influence on secondary metabolites. Overall, growth morphology exhibited the greatest total effect (0.685) (Fig. 8B) on biomass, followed by photosynthetic performance, antioxidant enzymes, and secondary metabolites (all positive total effects), whereas primary metabolites had a negative total effect.\u0026nbsp;\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eExogenous sorbitol-chelated potassium affects growth and photosynthetic performance of wheat seedlings under drought stress\u003c/p\u003e\u003cp\u003eDrought stress inhibits wheat seedling growth and biomass accumulation, reduces photosynthetic pigment content, and directly impairs net photosynthetic efficiency and stomatal conductance. This study demonstrates that exogenous sorbitol-chelated potassium application positively influences wheat seedling growth under drought conditions. Drought stress impedes the synthesis of photosynthetic pigments in wheat seedling leaves, thereby reducing photosynthetic capacity. Chlorophyll, a key pigment for light energy absorption, plays a key role in regulating photosynthetic efficiency [\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e]. In this study, SK treatment significantly increased total chlorophyll content compared with non-chelated MK treatment, and this change was positively correlated with the net photosynthetic rate. These findings suggest that SK helps inhibit chlorophyll degradation, thereby maintaining photosynthetic stability under drought stress.\u003c/p\u003e\u003cp\u003eTo cope with water deficit, plants typically adjust their stomatal aperture to reduce transpiration, limiting gas exchange and preventing excess water loss [\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e]. In this study, SK treatment reduced stomatal conductance, indicating enhanced drought resistance through partial stomatal closure. This effect may reflect sorbitol-chelated potassium induced regulation of stomatal behavior near its physiological threshold. While stomatal closure conserves water, it also limits CO₂ entry, potentially limiting photosynthesis. Research indicates that a decline in intercellular carbon dioxide concentration reduces photosynthetic activity, consequently impairing plant growth and development [\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e]. By converting free potassium ions into organically chelated forms, sorbitol-chelated potassium enhances photosynthetic efficiency and facilitates the transport of photosynthetic products, thereby promoting crop growth and development. Chelated potassium penetrates the waxy cuticular layer of leaves more effectively than free ionic forms. The lipophilic nature of the chelate allows more efficient passage through the lipid bilayer of the cuticle and enhances its mobility within the phloem. In this study, sorbitol-chelated potassium not only regulated wheat seedling growth but also significantly improved photosynthetic performance and activated antioxidant enzyme activity. These findings provide an important foundation for a deeper understanding of the comprehensive physiological mechanisms by which sorbitol-chelated potassium alleviates drought stress.\u003c/p\u003e\u003cp\u003eExogenous sorbitol-chelated potassium mitigate oxidative stress by enhancing antioxidant defense\u003c/p\u003e\u003cp\u003eDrought stress leads to increased malondialdehyde (MDA) accumulation and disrupts ROS homeostasis in wheat seedlings. MDA, a well-established marker of lipid peroxidation, was significantly elevated in CK\u003csub\u003e2\u003c/sub\u003e treatment, consistent with the findings of Aksu [\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e]. Compared to MK treatment, the application of SK significantly reduced MDA levels, suggesting that SK enhances ROS scavenging capacity. This effect may be attributed to the coordination properties of sorbitol, which converts potassium ions from free ionic state into chelated form. Following foliar absorption, chelated potassium is more readily translocated within wheat plants and experiences reduced fixation in the phloem. This mechanism is comparable to how sorbitol facilitates calcium transport [\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e], contributing to improved drought stress mitigation in wheat plants. Moreover, drought stress stimulates ROS-mediated redox signaling, which activates plant defense mechanisms. The enzymatic antioxidant system, which includes SOD, POD, CAT and other enzymes, plays a crucial role in detoxifying ROS and preventing oxidative damage [\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e]. In this study, sorbitol-chelated potassium application significantly enhanced SOD, POD, and CAT activity. SOD catalyzes the dismutation of superoxide radicals (\u0026middot;O\u003csup\u003e⁻2\u003c/sup\u003e) into hydrogen peroxide (H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e) and oxygen (O\u003csub\u003e2\u003c/sub\u003e). Subsequently, POD and CAT detoxify H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e by reducing it to water, further mitigating oxidative damage [\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e]. Therefore, the foliar application of SK alleviates drought stress in plants by enhancing antioxidant enzyme activity and improving ROS detoxification. Building on these findings, metabolomic analysis provides additional insights into the molecular mechanisms through which SK modulates redox balance and enhances drought tolerance at the metabolic regulation level.\u003c/p\u003e\u003cp\u003eEffects of exogenous sorbitol-chelated potassium on differential metabolite expression of wheat seedlings under drought stress\u003c/p\u003e\u003cp\u003eKEGG enrichment analysis revealed that both metabolic pathways and biosynthetic pathways of secondary metabolites were highly enriched with differential metabolites, suggesting a close association between these pathways and wheat growth under water-limited conditions. In this study, a comparison with the CK\u003csub\u003e2\u003c/sub\u003e treatment identified a total of 167 up-regulated and 170 down-regulated metabolites in the metabolic and biosynthetic pathways of secondary metabolites in the SK, K, MK, and S treatments. Compared with CK\u003csub\u003e2\u003c/sub\u003e treatment, the K exhibited a higher number of upregulated differential metabolites than downregulated ones. Conversely, SK, MK, and S treatments showed a greater number of downregulated than upregulated metabolites. This pattern suggests that exogenous application of sorbitol-containing treatments primarily mediates drought stress responses through metabolite downregulation, whereas potassium applied alone (K) may mitigate drought-induced damage by promoting metabolite upregulation.\u003c/p\u003e\u003cp\u003eThe upregulation of differential metabolites likely reflects an active adaptive response of wheat seedlings to environmental stress, enabling the enhancement of protective biochemical processes. Conversely, the downregulation of metabolites may represent a strategy for reducing overall metabolism, suppressing growth, and facilitating adaptation to unfavorable conditions. To mitigate drought stress, wheat synthesizes significant quantities of primary and secondary metabolites, including soluble sugars, amino acids, lipids, flavonoids, and other antioxidant compounds, while also activating detoxification enzymes [\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e]. These differential metabolites highlight the mechanisms through which exogenous sorbitol-chelated potassium application mobilizes metabolic pathways in wheat leaves to coordinate physiological and biochemical responses, thereby alleviating drought-induced damage in wheat.\u003c/p\u003e\u003cdiv id=\"Sec11\" class=\"Section2\"\u003e\u003ch2\u003ePrimary Metabolites\u003c/h2\u003e\u003cp\u003eSugars serve as not only as a primary energy source for plants but also as key signaling molecules that regulate various physiological processes during growth and stress adaptation [\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e]. Varshney et al. [\u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e33\u003c/span\u003e] revealed that celery and many Rosaceae species exhibit drought stress resistance primarily due to the accumulation of sorbitol and mannitol, which function as osmotic protectants and antioxidants. Soluble sugars undergo rapid interconversion; sucrose is hydrolyzed into glucose and fructose, while these hexoses, in turn, promote sucrose resynthesis, thus integrating into multiple metabolic pathways [\u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e]. The results of this study indicate that in the SK, MK, and S treatments, D-glucosamine-6-phosphate and xylitol participates in glycolysis and gluconeogenesis via the pentose phosphate pathway, yet its levels were downregulated. This downregulation may indicate an adaptive strategy to reduce energy consumption, as gluconeogenesis is energetically costly, thereby limiting energy loss under stress conditions. Previous studies suggest that sugar metabolism plays a dual role in regulating both energy balance and osmotic adjustment during drought. Furthermore, accumulating evidence indicates that amino acids also play a crucial role in plant stress resistance by functioning as osmotic regulators and signaling molecules, reinforcing their significance in drought tolerance mechanisms.\u003c/p\u003e\u003cp\u003eAmino acids function as osmolytes, precursors of secondary metabolites, ROS scavengers, and potential regulatory and signaling molecules that help plants to cope with stress. Elevated levels of specific amino acids are considered functionally significant in enhancing stress resistance [\u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e35\u003c/span\u003e]. Drought stress typically induces a significant accumulation of free amino acids in wheat seedlings, which may contribute to improved osmotic stress tolerance. These findings align with previous reports in other plant species. In this study, we observed significant changes in the metabolic pathways of several amino acids in wheat leaves under drought stress. Notably, pathways related to tryptophan, glutamate, valine, leucine, and isoleucine were significantly downregulated, while the free amino acid pools of proline, tryptophan, and branched-chain amino acids (valine, leucine, and isoleucine) were elevated [\u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e, \u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e]. In our study, we found that the metabolic pathways of several high-abundance amino acids and most low-abundance amino acids in wheat leaves were significantly downregulated during drought stress, including those of tryptophan, glutamate, valine, leucine, and isoleucine, suggesting that wheat seedlings mitigate drought stress by accumulating free amino acids. Levels of amino acids, most notably proline, tryptophan, and the branched chain amino acids leucine, isoleucine, and valine were increased under drought stress in all cultivars. These findings suggest that wheat seedlings mitigate drought stress through the accumulation of amino acids, serving both as osmoprotectants and regulators of metabolic homeostasis.\u003c/p\u003e\u003cp\u003eLipids and lipid-like molecules in plant cells are critical not only for membrane structure and energy storage but also for diverse biological functions. They serve as signaling molecules and as precursors for the synthesis of defense-related phytohormones, such as jasmonic acid [\u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e38\u003c/span\u003e]. In this study, linolenic acid and α-linolenic acid metabolic pathways were significantly affected by the SK, MK, and S treatments. Compared with potassium application alone, sorbitol-containing treatments promoted greater lipid accumulation in wheat leaves, suggesting enhanced membrane integrity and stress signaling capacity. In addition, purine and pyrimidine metabolism, key components of plant energy metabolism [\u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e39\u003c/span\u003e], were modulated by SK application, contributing to the maintenance of energy homeostasis under drought conditions. Collectively, these findings demonstrate that exogenous SK application significantly enhances wheat drought tolerance by regulating carbohydrates, amino acid, and lipid metabolism, thereby improving energy metabolism, osmotic regulation, and cell defense mechanisms. These findings reveal the complexity and synergy of plant metabolic networks and provide an important foundation for further exploration of the role of secondary metabolites in drought resistance mechanisms in wheat seedlings.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec12\" class=\"Section2\"\u003e\u003ch2\u003eSecondary Metabolites\u003c/h2\u003e\u003cp\u003ePlant hormones are central regulators enabling plants to cope with environmental stress, integrating external drought signals with internal physiological responses. In this study, the formation of the jasmonoyl-isoleucine conjugate in the SK treatment likely contributed to stress resistance. Jasmonoyl-isoleucine levels typically increase under dehydration stress [\u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e40\u003c/span\u003e] and interact with abscisic acid (ABA) accumulation to participate in the dehydration signal transduction pathway [\u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e41\u003c/span\u003e, \u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e42\u003c/span\u003e]. Therefore, sorbitol-chelated potassium may alleviate drought stress by mobilizing jasmonoyl-isoleucine and modulating hormone-mediated signaling in wheat leaves. Beyond hormonal signaling, carotenoids play a crucial role in drought tolerance as photoprotective agents, antioxidants, and precursors for ABA biosynthesis.\u003c/p\u003e\u003cp\u003eCarotenoid accumulation not only provides precursors for the biosynthesis of the plant hormone ABA [\u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e43\u003c/span\u003e, \u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e44\u003c/span\u003e], but also enhances drought tolerance by supporting ROS scavenging and improving photoprotection [\u003cspan citationid=\"CR45\" class=\"CitationRef\"\u003e45\u003c/span\u003e]. In this study, Rhodopinal, Astaxanthin, and Hydroxychlorobactene were detected in the MK, S, and K treatments. As a carotenoid derivative, Rhodopinal and Hydroxychlorobactene possess characteristic chromophores that assist photosynthetic pigments and scavenge free radicals [\u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e46\u003c/span\u003e]. Additionally, astaxanthin, a highly efficient antioxidant, was identified and may contribute to enhanced antioxidant capacity and stress adaptation [\u003cspan citationid=\"CR47\" class=\"CitationRef\"\u003e47\u003c/span\u003e]. These findings suggest that carotenoids, as key components of the non-enzymatic antioxidant system, mitigate drought stress by scavenging ROS modulating detoxification enzyme activity. Similarly, flavonoids, another class of secondary metabolites, play multifaceted roles in plant responses to drought stress.\u003c/p\u003e\u003cp\u003eFlavonoids regulate hormone signal transduction and act as inhibitors of auxin transport [\u003cspan citationid=\"CR48\" class=\"CitationRef\"\u003e48\u003c/span\u003e]. Recent studies have shown that flavonoids, such as flavones, flavanones, flavonols, isoflavones, and anthocyanins, play an essential role as ROS scavengers, contributing significantly to oxidative stress mitigation [\u003cspan citationid=\"CR49\" class=\"CitationRef\"\u003e49\u003c/span\u003e]. Furthermore, flavonoids and anthocyanins enhance plant drought resistance and support growth under adverse conditions [\u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e50\u003c/span\u003e]. In this study, anthocyanin biosynthesis was observed in MK-treated plants, suggesting a potential role for these compounds in drought tolerance mechanisms. Notably, the expression of some flavonoids was downregulated under drought stress, indicating a dynamic regulatory adjustment. This pattern suggests that the production of flavonoid-related metabolites contributes to plant defense against drought stress and positively influences growth and performance. Plant hormones, carotenoids, and flavonoids, key regulators in plant metabolic networks, work in coordination to mediate drought stress responses. Plant hormones regulate physiological processes primarily through signal transduction pathways, while carotenoids and flavonoids, as secondary metabolites, enhance stress resistance through photoprotection, antioxidation, and regulation of hormone signaling. Collectively, these integrated processes highlight the synergistic regulatory role of primary and secondary metabolites in improving plant drought stress tolerance, orchestrating these metabolic and signaling adjustments to optimize plant responses to water deficit.\u003c/p\u003e\u003cp\u003eMechanism Analysis of Exogenous Sorbitol-Chelated Potassium on Biomass of Wheat Seedlings under Drought Stress\u003c/p\u003e\u003cp\u003eWheat drought resistance is mediated by the synergistic regulation of photosynthetic performance, antioxidant defense systems, and metabolite expression. However, most existing studies have not comprehensively integrated these key factors, including growth morphology, photosynthesis, antioxidant defense, and differential metabolites, into a unified framework. In this study, we applied correlation analysis and structural equation modeling (SEM) to investigate these interactions. Correlation analysis revealed that the exogenous application of sorbitol-chelated potassium orchestrates a cascade response by synergistically regulating N-Acetyl-D-glucosamine, (-)-Jasmonoyl-L-isoleucine, and growth morphology, and (-)-Maackiain, N-Acetyl-D-glucosamine, (-)-Jasmonoyl-L-isoleucine, 5-Methoxytryptamine, and 9,10-Dihydroxy-12,13-epoxyoctadecanoic acid were significantly correlated with wheat photosynthetic performance. The results indicated that the exogenous application of sorbitol-chelated potassium significantly mitigates the inhibitory effects of drought stress by synergistic regulation of growth morphology and photosynthetic performance of wheat seedlings.\u003c/p\u003e\u003cp\u003eSEM analysis further revealed that growth morphology exerted the strongest total effect on wheat seedling biomass accumulation (0.685), directly dominating biomass formation. Photosynthetic performance (direct effect 0.211) and the antioxidant enzyme system (direct effect 0.306) positively regulated biomass, acting through independent pathways. Specifically, photosynthetic performance mediated an indirect effect of 0.320 through antioxidant enzymes and secondary metabolites, while the antioxidant enzyme system exerted an indirect effect of 0.351 via growth morphology (Fig.\u0026nbsp;8B). These results highlight growth morphology as the primary driver of biomass accumulation, while photosynthetic performance and antioxidant defense form a synergistic regulatory network through direct effects and multiple indirect pathways (such as secondary metabolite regulation and growth morphology feedback), jointly regulating biomass accumulation. This mechanism highlights the critical role of integrated regulation between growth morphology and physiological metabolism in driving wheat biomass accumulation. Therefore, the synergistic mechanism by which exogenous application of sorbitol-chelated potassium enhances drought resistance involves growth morphology serving as the primary driver of biomass accumulation, and photosynthetic performance, the antioxidant enzyme system, and secondary metabolites, such as (-)-jasmonoyl-L-isoleucine and Etherolenic acid, form a synergistic regulatory network that collectively alleviates the inhibitory effects of drought stress across multiple functional dimensions. This integrative framework offers a novel perspective for understanding wheat seedling growth and metabolic adaptation under water-limited conditions and provides valuable insights for improving drought resilience in wheat.\u003c/p\u003e\u003c/div\u003e"},{"header":"Conclusion","content":"\u003cp\u003eThe exogenous application of sorbitol-chelated potassium (SK) alleviates drought-induced growth inhibition in wheat seedlings through a multi-faceted physiological mechanism. This mitigation is achieved by enhancing photosynthetic pigment content and gas exchange capacity, upregulating antioxidant enzyme activity, and regulating accumulation of key metabolites. Drought stress significantly limits wheat growth and biomass accumulation. Compared to CK\u003csub\u003e2\u003c/sub\u003e, SK application significantly mitigated biomass reduction (19.2%) under drought stress conditions. Exogenous SK significantly increased photosynthetic pigment content and gas exchange parameters in wheat flag leaves, while enhancing the activities of antioxidant enzymes and reducing malondialdehyde levels (45.66%), significantly alleviating oxidative damage caused by drought stress. Untargeted metabolomics analysis revealed that SK treatment induced the relative accumulation of differential metabolites, thereby inhibiting sugar metabolism to minimize energy loss, accumulating free amino acids to enhance osmotic adjustment, and promoting lipid metabolism to maintain cellular energy homeostasis. Additionally, SK mitigated drought-induced growth inhibition in wheat by modulating plant hormone signaling, enhancing carotenoid and flavonoid biosynthesis, and activating detoxification enzyme system to scavenge ROS.\u003c/p\u003e\u003cp\u003eIn summary, structural equation modeling (SEM) demonstrated that exogenous application of sorbitol chelated potassium, with growth morphology as the core driving factor, synergistically regulates key physiological processes, including photosynthetic performance, antioxidant defense systems, differential metabolite. This forms a coordinated multi-system network of \"morphology-physiology-metabolism\" to regulate drought resistance, which promotes biomass accumulation and significantly enhances the drought tolerance of wheat seedlings.\u003c/p\u003e"},{"header":"Abbreviations","content":"\u003ctable border=\"0\" cellspacing=\"0\" cellpadding=\"0\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 94px;\"\u003e\n \u003cp\u003eABA\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 312px;\"\u003e\n \u003cp\u003eAbscisic\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 94px;\"\u003e\n \u003cp\u003eCAT\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 312px;\"\u003e\n \u003cp\u003eCatalase\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 94px;\"\u003e\n \u003cp\u003eCi\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 312px;\"\u003e\n \u003cp\u003eIntercellular CO\u003csub\u003e2\u003c/sub\u003e concentration\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 94px;\"\u003e\n \u003cp\u003eGs\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 312px;\"\u003e\n \u003cp\u003eStomatal conductance\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 94px;\"\u003e\n \u003cp\u003eMDA\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 312px;\"\u003e\n \u003cp\u003eMalondialdehyde\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 94px;\"\u003e\n \u003cp\u003ePn\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 312px;\"\u003e\n \u003cp\u003ePhotosynthetic rate\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 94px;\"\u003e\n \u003cp\u003ePOD\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 312px;\"\u003e\n \u003cp\u003ePeroxidase\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 94px;\"\u003e\n \u003cp\u003eRWC\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 312px;\"\u003e\n \u003cp\u003eRelative water content\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 94px;\"\u003e\n \u003cp\u003eROS\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 312px;\"\u003e\n \u003cp\u003eReactive oxygen species\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 94px;\"\u003e\n \u003cp\u003eSOD\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 312px;\"\u003e\n \u003cp\u003eSuperoxide dismutase\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 94px;\"\u003e\n \u003cp\u003eTr\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 312px;\"\u003e\n \u003cp\u003eTranspiration rate\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis study was supported by the projects of the National Natural Science Foundation of China (31972516), the Key Research and Development Program of Shandong Province, China (2017GNC11116).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthors\u003c/strong\u003e\u003cstrong\u003e\u0026rsquo;\u003c/strong\u003e\u003cstrong\u003eContributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eGuohui Du conceived the research. Mingxia Zhang and Huanyang Zhang conducted the experiments, analyzed the data, and then wrote the paper. Ruili Zheng and Li Zhao revised the paper in English. Mingli Huang, Xiaocui Wang, and Kezhong Liu revised the manuscript.\u0026nbsp;Yan Dongyun performed the final review. All the authors contributed to the article and approved the submitted version.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eData availability\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eData will be made available on request.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eDeclarations\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEthics approval and consent to participate\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent for publication\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare that they have no conflict of interest\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eClinical trial number\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eQiu YQ, Zhang LX, Yang DD, Chen JY, Zhang XS, Zhang HR. 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Tree Physiol.2016;36(2):129-132. https://doi.org/10.1093/treephys/tpv128 \u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Wheat, Drought stress, Sorbitol-chelated potassium, Antioxidant system, Metabolomics","lastPublishedDoi":"10.21203/rs.3.rs-7498264/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-7498264/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003ePotassium fertilization is a strategy to alleviate the impact of drought stress on wheat production. However, the effects of chelated potassium remain to be verified. In this study, 10% PEG-6000 was used to simulate moderate drought stress on hydroponically grown XinHua818 wheat (\u003cem\u003eTriticum aestivum L.\u003c/em\u003e) seedlings, and the physiological and biochemical parameters of wheat sprayed with water (CK\u003csub\u003e2\u003c/sub\u003e), sorbitol (S), potassium chloride (K), potassium mixed with sorbitol (MK), and sorbitol-chelated potassium (SK) were monitored. Results showed that SK effectively alleviated the inhibitory effects of drought stress on seedling growth. The aboveground biomass of SK-treated seedlings was significantly higher than that of K and MK-treated seedlings, increasing by 15.66% and 20.00%, respectively. Compared to MK treatment, SK treatment significantly increased total chlorophyll content by 18.74% and reduced malondialdehyde levels by 16.02%. Superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) activity were also elevated in SK-treated seedlings compared to other treatments. Metabolomic analysis identified 51 differential metabolites in SK compared to CK\u003csub\u003e2\u003c/sub\u003e treatment, including sugars, amino acids, lipids, plant hormones, carotenoids, flavonoids, and their derivatives. These metabolites were enriched in 18 metabolic pathways, notably α-linolenic acid metabolism, histidine metabolism, plant hormone signal transduction, carotenoid biosynthesis, and flavonoid biosynthesis, suggesting their potential role in enhancing drought tolerance in wheat and their broader significance in drought resistance research.\u003c/p\u003e","manuscriptTitle":"Physiological and Metabolomic Analyses of Exogenously Applied Sorbitol-Chelated Potassium Enhancing Drought Tolerance in Wheat","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-09-12 12:28:50","doi":"10.21203/rs.3.rs-7498264/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"d39484ac-3495-4e56-bf72-97824dd23254","owner":[],"postedDate":"September 12th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2025-10-06T14:38:42+00:00","versionOfRecord":[],"versionCreatedAt":"2025-09-12 12:28:50","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-7498264","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-7498264","identity":"rs-7498264","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}