Responses of C:N:P stoichiometry and non-structural carbohydrate contents of Prunus mume seedling to nitrogen addition

Introduction

The seedling stage represents a critical phase in the plant life cycle, underpinning physiological development and subsequent growth. Nutrient requirements vary across developmental stages (Yang et al., 2023), making appropriate fertilization essential for producing vigorous seedlings and establishing productive plantations. Nitrogen (N) is the primary growth-limiting nutrient in plants (LeBauer & Treseder, 2008), and soil N availability strongly influences elemental concentrations in terrestrial plants by regulating nutrient and water uptake (Sardans & Peñuelas, 2012). Fertilization must therefore be timely, balanced, and technically appropriate. Excessive N supply can be counterproductive, often delaying reproductive development, particularly in young trees with strong vegetative growth (Wang et al., 2024a). Hence, controlled early fertilization following established protocols is recommended. Irrigation and nutrient management practices can profoundly affect plant physiological and ecosystem functioning. Furthermore, anthropogenic N emissions have significantly increased atmospheric N deposition globally (Kanakidou et al., 2016). Both elevated deposition and top-dressing N fertilization can alter the stoichiometry of aboveground and root organs in plants (Kou et al., 2018), subsequently altering the chemical composition, architecture, and functional traits in plant species.

In response to N fertilizer application, plants adjust elemental balances to modify tissue carbon (C): N: phosphorus (P) ratios across roots, stems and leaves (Ågren, 2008). C constitutes the structural framework of plant tissues, typically accounting for 39–42% of dry mass, and can limit growth and development in plants (Körner, 2003). N enhances leaf area index, prolongs photosynthesis, improves nutrient uptake, and ultimately increases productivity (Tian et al., 2023; Wang et al., 2024a). P availability strongly influences the transport of photosynthetic products and is recognized as a key element in plant metabolism (Dissanayaka et al., 2021; Vance et al., 2003). A growing body of evidence indicated that N addition differentially affects the stoichiometry of roots, stems, and leaves across plant species, and that elemental composition varies substantially among organs (Jing et al., 2017; Li et al., 2021). Changes in C, N and P contents as well as their stoichiometric ratios are associated with physiological activities, growth rate and overall plant performance (Ågren, 2008; Yin et al., 2021). Extensive research has examined the response of N and P levels and their stoichiometric ratios to N addition in different plants (Minden & Kleyer, 2014), because plant growth and organ function depend not only on absolute nutrient supply, but also on the balance among multiple elements (Chen & Chen, 2021; Güsewell, 2004). However, the plasticity of organ-specific stoichiometry under environmental change, and which plant organs exhibit greater homeostasis, remain unclear (Minden & Kleyer, 2014). Non-structural carbohydrates (NSC) represent the C pool available to buffer external stresses, and serve as indicators of stress resistance and environmental tolerance in plants (Dietze et al., 2014; Li et al., 2023). Studies have revealed a positive correlation between leaf N concentration and NSC fixation capacity (Wang et al., 2019), whereas P deficiency impairs photosynthesis (Wang et al., 2023). The levels of NSC and C, N, and P in plant organs reflect nutrient availability and strongly influence plant growth and development (Liu et al., 2020). Thus, the individual and interactive impacts of these factors shape the survival, growth, and development strategies in plants.

Prunus mume (Siebold) Siebold & Zucc., known as Chinese plum, Qingmei, or sour plum in China, Japanese apricot in Japan, is a deciduous fruit tree species in the family Rosaceae, genus Prunus. Native to China, it has been cultivated for over 3,000 years throughout most of the country (Gao et al., 2026; Mitani et al., 2018). Its fruit has been used as a medicinal herb and an excellent natural health food in East Asian countries for more than 2,000 years (Gong et al., 2021; Hu et al., 2025). Beyond its traditional use as a fruit and medicinal plant, P. mume is increasingly recognized for its role in forestry and agroforestry systems. Its deep root system and moderate tolerance to poor soils also make it suitable for soil and water conservation in hilly areas, as well as for low value forest transformation and urban greening (Gao et al., 2026; Mitani et al., 2018). Nursery practices, especially fertilization, critically affect seedling morphology, nutrition, and physiology (Grossnickle & El-Kassaby, 2016; Timmer, 1997). Seedlings with adequate nutrient reserves and balanced C allocation exhibit higher post-planting survival, stress tolerance, and growth (Oliet et al., 2013; Villar-Salvador et al., 2015), whereas nutrient deficiencies or imbalances can impair root development, mycorrhization, and field establishment (Mason et al., 2000).

Despite extensive studies on N addition effects on crop and model species, the organ-specific responses of C:N:P stoichiometry and NSC pools in P. mume seedlings under varying N supply remain poorly understood. Specifically, it remains unclear whether N addition consistently increases N content across organs, whether high N exacerbates P limitation, and whether C allocation shifts between starch (storage) and soluble sugars. To address these gaps, we formulated three hypotheses: (1) N addition increases total N but decreases P in all organs, raising N:P ratios most markedly in stems, (2) moderate N supply enhances biomass and soluble NSC, while excessive N reduces starch reserves and inhibits growth, and (3) roots exhibit greater stoichiometric sensitivity than leaves due to direct soil N exposure. Herein, this study aimed to investigate the effects of N addition on the growth, elemental accumulation, and NSC dynamics of P. mume seedlings. These results are expected to provide theoretical and practical information for adjusting nutrient management of P. mume seedlings. From a forestry perspective, understanding how N availability affects C:N:P stoichiometry and NSC dynamics is essential for producing robust seedlings for field planting. These findings are intended to support evidence-based fertilization protocols for P. mume nurseries and young plantations, thereby improving reforestation success and long-term productivity of P. mume based agroforestry systems.

Materials and Methods

Seedling culture and treatment

P. mume seeds were stratified in moist sand for approximately three months and subsequently sown via drill sowing. Following germination, the seedlings were cultivated until they reached a height of 50 cm (approximately 30–40 days), at which point poorly growing individuals were removed. The pot experiment was conducted in a greenhouse at Sichuan Agricultural University, located in Wenjiang District, Chengdu, China. Each pot (non-woven fabric, 21 cm upper diameter × 20 cm lower diameter × 15 cm height) was filled with 10 kg of soil. On April 16, 2023, the pre-cultivated seedlings were transplanted, with one seedling per pot. Based on the pot surface area (0.0314 m²) and a typical field planting density for P. mume orchards (≈ 833 plants ha⁻¹, corresponding to 3 m × 4 m spacing), the approximate field-equivalent N rates were: CK = 0, T1 ≈ 147, T2 ≈ 294, T3 ≈ 588, and T4 ≈ 1176 kg N ha⁻¹. The highest level (T4) was deliberately designed as supra-optimal to investigate the threshold of N, and is not intended as a practical fertilization recommendation. Consequently, four N addition levels per plant were established: CK (0 g N plant⁻¹), T1 (5 g N plant⁻¹), T2 (10 g N plant⁻¹), T3 (20 g N plant⁻¹), and T4 (40 g N plant⁻¹) (Figure 1). Each treatment comprised 10 pots, arranged in a completely randomized design. After transplanting, the seedlings were allowed a 45-day acclimatization period, after which fertilization was initiated. Fertilizer was applied in four equal split doses over a 60-day period (once every 15 days, Figure 1). For each application, the pre-scribed amount of urea was dissolved in 500 ml of tap water, and after thorough mixing, 25 ml of the solution was evenly poured around the seedling root zone (≈ 5 cm from the stem). The entire experimental period was conducted under a rain-out shelter to prevent N leaching from rain-fall. Daily watering was performed to maintain optimal soil moisture, and routine disease, pest, and weed control measures were carried out. At the end of the experiment, destructive sampling was performed on three randomly selected pots per treatment. From each selected seedling, roots, stems, and leaves were separately harvested. All chemical analyses were conducted on three biological replicates (n = 3), each derived from a single pot.

Figure 1.

Results

Growth responses to nitrogen addition

The variations in plant height and ground diameter across treatments were presented in Figure 2. Figure 2a showed that the initial seedling height was comparable among the five groups. After four fertilization cycles, the final plant height of the CK, T1, T2, T3 and T4 groups were 113.43 cm, 111.23 cm, 128.35 cm, 121.63 cm, and 122.40 cm, respectively. The T4 group had the highest growth rate, closely followed by the T2 group. With the exception of T1, plant height in all other treatment groups was significantly greater than that in the CK. The CK group showed greater variability in height increment compared to the other groups. Figure 2b showed that the initial ground diameter ranged primarily from 2.60 to 2.85 cm. The CK group had an initial diameter of 2.85 cm, which exceeded that of the other treatment groups. Following the fourth fertilization, the ground diameter of the CK group decreased after the fourth fertilization, relative to the other experimental groups, whereas the ground diameter of the T2 group increased following the fourth fertilization. Overall, throughout the entire fertilization period, the lowest diameter growth rate was observed in the CK group, whereas the highest value was recorded in the T2 group.

Figure 2.

Responses of C, N and P contents to nitrogen addition

The C, N and P contents in roots, stems and leaves under N addition conditions were shown in Figure 3. As shown in Figure 3a, C content 307.66 to 389.34 mg g⁻¹ in leaves, 371.61 to 412.87 mg g⁻¹ in stems, and 367.96 to 408.79 mg g⁻¹ in roots, respectively. Among all treatments, the T1 group exhibited the highest C content in both leaves and roots, significantly surpassing that of the T4 group. Conversely, the T4 group showed a markedly lower C content across all three organs compared to the other treatment groups. Figure 3b showed that N content ranged from 31.44 to 35.83 mg g⁻¹ in leaves, 4.83 to 8.71 mg g⁻¹ in stems, and 10.78 to 15.31 mg g⁻¹ in roots, respectively. N content was highest in T4 roots and stems (15.31 and 8.71 mg g⁻¹, respectively), representing a 42% and 80% increase over the control. Figure 3c showed that P content ranged from 0.487 to 0.558 mg g⁻¹ in leaves, 0.300 to 0.383 mg g⁻¹ in stems, and 0.360 to 0.551 mg g⁻¹ in roots, respectively. Root P content in the control group was significantly higher than that in all four N-addition treatments. Stem P content in CK and T1 was significantly higher than in T2, T3, and T4. Overall, C, N, and P contents varied significantly among organs and across N addition levels when analyzed separately by organ.

Figure 3.

C, N and P accumulation responses to nitrogen addition

Figure 4 showed the impact of varying N application rates on C, N, and P accumulation in P. mume seedlings. Figure 4a showed that the range of C accumulation was 348.81–398.71 mg plant⁻¹ under different N addition levels. From group T1 to T4, C accumulation declined significantly with increasing N supply. Only the T4 group had a lower C accumulation than that in the CK group. Figure 4b showed that N accumulation ranged from 16.44 to 20.66 mg plant⁻¹. N accumulation increased significantly in all N-supplied groups (T1–T4), and the highest value (20.66 mg plant⁻¹) was observed in T4, compared to 19.01 mg plant⁻¹ in CK. As shown in Figure 4c, P accumulation ranged from 0.385 to 0.471 mg plant⁻¹. P accumulation in T4 was notably lower than in the other groups. Across root, stem, and leaf fractions, P accumulation initially increased with N supply (from T1 onward) and then declined, with the decreasing trend consistent across all three organs.

Figure 4.

Stoichiometric ratios of C, N, P responses to nitrogen addition

Figure 5 showed the C, N, and P stoichiometric ratios across treatments. In Figure 5a, the C:N ratios in the leaves, roots, stems, and the total plant ranged from 8.57 to 12.32, 25.18 to 37.95, 42.79 to 83.28, and 16.88 to 24.03, respectively. The whole-plant C:N ratio was between the roots' and leaves' C:N ratios. The T1 group showed significantly higher C:N ratios in leaves, roots, and whole plant than the other groups. In stems, the C:N ratio in T1 group was significantly higher than in the T3, T4 and CK groups. As depicted in Figure 5b, under N addition conditions, the C:P ratios in the leaves ranged from 614.82 to 699.42, while the C:P ranges in the roots and stems were 701.44 to 1070.51 and 1017.89 to 1237.68, respectively. The C:P ratios in the seedlings fluctuated between 776.83 and 906.06, closely resembling the root C:P. Figure 5c showed that the N:P ratios in the leaves, roots, stems, and total plant ranged from 56.67 to 73.64, 22.11 to 42.56, 13.81 to 29.00, and 35.25 to 53.66, respectively. The whole-plant N:P ratio was slightly higher than the stems' N:P ratio.

Figure 5.

NSCs content responses to nitrogen addition

The concentrations of NSCs in the roots, stems, and leaves were presented in Figure 6. As shown in Figure 6a, glucose content in the leaves, roots and stems ranged from 32.41 to 37.68 mg g⁻¹, 28.25 to 39.81 mg g⁻¹, and 21.32 to 28.55 mg g⁻¹, respectively. The highest root glucose concentration (39.81 mg g⁻¹) was in the CK group. As shown in Figure 6b, fructose content in the leaves, roots and stems ranged from 3.83 to 9.21 mg g⁻¹, 2.17–7.95 mg g⁻¹ and 2.32–4.95 mg g⁻¹, respectively. The maximum leaf fructose (9.21 mg g⁻¹) was in the T4 group. As depicted in Figure 6c, sucrose content in the leaves, roots and stems ranged from 19.73 to 25.96 mg g⁻¹, 11.12 to 23.24 mg g⁻¹, and 8.21 to 16.93 mg g⁻¹, respectively. The highest leaf sucrose (25.96 mg g⁻¹) was in the T1 group. As illustrated in Figure 6d, starch content in the leaves, roots and stems ranged from 30.04 to 40.96 mg g⁻¹, 42.34 to 81.73 mg g⁻¹, and 46.40 to 53.42 mg g⁻¹, respectively. Starch content in roots, especially in CK and T1, was notably higher than in the other groups.

Figure 6.

Correlation analysis, PCA, and linear fitting

Figure 7a showed the correlation and cluster analysis of morphological indicators and C, N, and P accumulation under the N addition. Positive correlations were observed between biomass and root length, root length and plant height, plant height and ground diameter, root-shoot ratio and ground diameter, C and P accumulation. In contrast, N accumulation had significant negative correlations with biomass, C accumulation, and P accumulation. Cluster analysis showed that N accumulation contrasts with both C and P accumulation, whereas C accumulation aligned with P accumulation. Biomass with root length, and seedling height with ground diameter, formed smaller clusters. These findings indicated that increasing N addition leads to significant differences in element accumulation among treatments relative to CK, yet the relationships between C, N, P accumulation and morphological indicators remained non-significant. As shown in Figure 7b, PCA was performed to explore multivariate relationships among C, N, P, soluble sugars (glucose, fructose, sucrose), starch, and biomass across all organs and treatments. All variables were standardized (z-score) prior to analysis. Each point in the PCA biplot represents an individual biological replicate, with organs distinguished by colors and treatments by symbols. The first principal component (PC1) explained 46.2% of the total variance and was characterized by positive loadings for soluble sugars and negative loadings for starch and biomass, suggesting a trade-off between immediate metabolic soluble sugars and storage starch. The second principal component (PC2) explained 22.7% of the variance and was primarily associated with N and P contents. In the ordination space, roots from T3 and T4 treatments showed more pronounced shifts toward higher soluble sugar and N content, whereas stems and leaves exhibited less separation across treatments. This descriptive pattern suggests that roots are more responsive to N addition in terms of C-nutrient reallocation, which is consistent with the organ-specific ANOVA results showing significant N-induced changes in root NSC and stoichiometry. However, the PCA does not provide formal statistical evidence for organ differences; rather, it serves as a complementary visualization of the multivariate trends. Figure 7c showed the linear correlations between N content and C, P, and NSC. Negative correlations between N content and C content as well as starch were revealed, respectively. In contrast, N content and P content showed a positive correlation (K = 0.006). Furthermore, positive correlations were observed between N content and glucose, fructose, sucrose, with K values of 0.292, 0.075, and 0.366, respectively.

Figure 7.

Discussion

Fertilization is a cornerstone agronomic practice for cultivating robust seedlings and ensuring their subsequent growth and development (Jiang et al., 2024; Yang et al., 2023). Consequently, identifying an appropriate N addition regime is of paramount importance for the successful expansion of P. mume cultivation areas. In this study, P. mume seedlings exhibited viable growth across a gradient of N supply levels, however, significant changes were observed in morphological indicators, C, N, P and NSC concentrations, accumulation, distribution, and stoichiometric ratios across roots, stems, and leaves. Furthermore, correlation analysis, PCA, and linear fitting collectively revealed multiple significant positive and negative relationships among growth indicators, C:N:P ratios, and NSC contents in response to differential N addition (Figure 7). These multifaceted responses highlight that both deficiency and excess in N supply can disrupt internal nutrient homeostasis, with implications for stress resilience and post-transplant establishment. These findings provide both theoretical foundations and practical guidelines for refining nutrient management strategies during the nursery stage of P. mume cultivation. Moreover, they provide a basis for optimizing field transplantation protocols and tailoring N application schedules to specific soil conditions, ultimately contributing to enhanced orchard productivity and sustainable cultivation practices.

Morphological indicators are fundamental in seedling research, as they directly reflect growth status and provide actionable insights for optimizing cultivation practices. Changes in these indicators can be effectively employed to assess the supply-demand balance between seedlings and N fertilizer. In the present study, plant height reached a maximum at a low N addition level and then decreased when N supply either increased or decreased, demonstrating a dual (unimodal) response to N fertilizer. A similar trend was observed for several biomass parameters, with peak values occurring at either T1 (5 g N plant⁻¹) or T3 (20 g N plant⁻¹). The root-to-shoot ratio peaked under low N addition but also showed a moderate increase at the high N level. In contrast, ground diameter showed no significant change across N gradients, whereas root length increased steadily with rising N supply (Table 1 and Figure 2). The inhibitory effect observed under the highest N addition (40 g N plant⁻¹) may be attributed to multiple factors, including altered soil pH, shifts in soil microbial community composition and activity, reduced nitrogen use efficiency (NUE), and intensified competitive partitioning between aboveground and belowground organs (Jia et al., 2024). Comparable unimodal responses have been documented in other woody species. For instance, Cunninghamia lanceolata (Lamb.) Hook. seedlings also exhibit biomass promotion under low to moderate N but inhibition under excessive N (Xu & Timmer, 1999). Importantly, the deliberate inclusion of our highest dose (T4, 40 g N plant⁻¹) as a supra-optimal treatment was specifically designed to capture the declining phase of the response curve. This level was never intended to replicate realistic field fertilization, but rather to define the upper threshold beyond which N becomes phytotoxic. The observed growth suppression in T4, together with the marked reduction in root starch reserves (Figure 6d), confirms that this level exceeds the physiological optimum for P. mume seedlings and provides a mechanistic rationale for avoiding excessive N application in nursery practice. Unlike certain coniferous species in which high N supply suppresses root elongation, we found a steady increase in P. mume root length with rising N addition. This interspecific difference may reflect the higher N demand of fruit trees or alternatively, a compensatory foraging response under confined pot conditions. The observed root length increase could also result from accelerated fine-root growth and a greater root node length at lower N levels. Overall, the growth suppression under high N in this experiment likely results from diminished NUE, osmotic imbalance, and the accumulation of metabolic by-products that exert toxic effects on seedling physiology. Taken together, these morphological and allocation patterns indicated that moderate N (10–20 g N plant⁻¹) constitutes the optimal dosage for promoting biomass accumulation in P. mume seedlings.

Nutrient availability is a primary determinant of seedling performance, and stoichiometric analysis offers a powerful lens for examining the internal balance among elements and its modulation by N supply. Across a gradient of N addition, the concentrations of C, N, and P contents and their stoichiometric ratios in leaves, stems, and roots, collectively reflect the plant's resource allocation strategies, growth strategy, and stress resistance (Scharler et al., 2015). Our results showed substantial organ-specific shifts in C, N, and P contents under varying N addition. With increasing N supply, N content rose markedly in all three organs, whereas P content decreased significantly. Notably, N content in the CK group was equal to or even higher than that in the three lowest N-treated groups, indicating that the relationship between N addition and tissue N accumulation is not strictly linear, particularly at low doses. Elevated N application was associated with a clear reduction in C content in both roots and leaves (Figure 3). These patterns are consistent with previous studies in other woody species. For instance, N addition significantly increased N content in the roots, stems, and leaves of Pinus tabuliformis Carrière while decreasing P content (Jing et al., 2017). Similarly, trends were reported for Pinus massoniana Lamb. seedlings, in which fertilization elevated N content relative to controls (Wu et al., 2019). In our experiment, total P accumulation decreased substantially with escalating N fertilizer, whereas N accumulation showed a significant upward trend. Among all treatments, the T4 group (40 g N plant⁻¹) had the highest N accumulation but the lowest P accumulation across all organs (Figure 4). Figure 7 illustrated significant negative correlations between N accumulation and both C accumulation and P accumulation, as well as between P accumulation and biomass and root length. These findings suggested that N- and P-related stoichiometric changes under N addition are closely linked to morphological adjustments in plant parts.

The sensitivity of C:N:P stoichiometry to N addition was also evident. Our results indicated that the C:N ratio in seedling roots, stems, and leaves decreased significantly with increasing N supply, while the N:P ratio increased markedly. Comparable findings have been reported for Pleioblastus amarus (Keng) Keng f. and Pinus koraiensis Siebold & Zucc. (Li et al., 2021; Yang et al., 2019). Additionally, N application coincided with a pronounced increase in the C:P ratio in roots and stems. It is widely accepted that C:N and C:P ratios reflect C sequestration capacity and nutrient-use efficiency, whereas the N:P ratio serves as an effective indicator of plant response to environmental change and growth limitation. According to the established threshold, N:P values between 14 and 16 suggest co-limitation by N and P, while values exceeding 16 points to P limitation (Güsewell, 2004). In this study, only stems under low N addition fell within the 14–16 range, all other plant parts had N:P ratios above 16 (Figure 5). This suggests that under low N conditions, stems are co-limited by N and P, possibly reflecting lower sensitivity to N availability or a higher N demand compared to roots and leaves. Indeed, roots have been reported to respond particularly strongly to N addition (Yin et al., 2021), and our root N:P data are consistent with that pattern. The fact that both roots and leaves showed N:P ratios exceeding 16, coupled with the significant increases in C:P and N:P ratios and the sharp decline in P content under high N, supports the interpretation that excessive N supply may exacerbate P scarcity or reduce P utilization efficiency.

N is indispensable for photosynthesis, which drives the synthesis and accumulation of various nutrients, particularly NSCs. Soluble NSCs serve immediate functions and act as rapid C sinks, whereas starch functions as a long-term C sink (Dietze et al., 2014). Consequently, N addition can directly or indirectly affect NSC content in P. mume seedlings. Our results showed that high N application was associated with a marked reduction in starch content, especially in roots (Figure 6). This aligns with previous studies on Quercus mongolica Fisch. ex Ledeb. and Acer pseudoplatanus L. (Mackie-Dawson et al., 1994; Wang et al., 2024b). The underlying mechanisms remain unclear, but possible explanations include shifts in C allocation priorities, altered source–sink relationships, or accelerated starch turnover. In contrast, N addition significantly increased fructose content in all three organs, and increased sucrose content in stems and roots (Figure 6), implying a redistribution of C among NSC fractions under varying N supply. Whether the increase in soluble NSCs reflects enhanced photosynthetic output, reduced conversion to starch, or accelerated starch turnover cannot be determined from our current data. Notably, we found positive correlations between N and P contents and soluble NSCs, while soluble NSC showed a negative correlation with C content, indicating coordinated shifts in nutrient and C status, although causal links remain speculative. This pattern reflects the possibility that C availability constrains seedling growth (Wiley & Helliker, 2012). Resolving these relationships would require direct measurements of C fluxes, photosynthetic rates, and activities of enzymes involved in NSC metabolism. Furthermore, prior research indicated that the growth-promoting effect of N addition on seedlings diminishes with sustained high N supply, as excessive N impairs multiple aspects of seedling performance. This further explains why morphological indicators did not improve under high N in our experiment. The apparent shift from starch to soluble NSCs under high N supply has potential ecological implications. Starch reserves are important for stress tolerance and resprouting capacity following disturbance (Chapin et al., 1990; Villar-Salvador et al., 2015). The observed reduction in starch, particularly in roots, may compromise seedling resilience to transplant shock, drought, or other adverse conditions, although we did not directly measure stress tolerance or post-planting survival. Whether such compositional changes affect subsequent establishment in the field remains an open question that warrants further investigation.

From a practical standpoint, our results offer preliminary guidance for nursery management of P. mume seedlings intended for afforestation or agroforestry systems. A unimodal growth response was observed, with moderate nitrogen supply (10–20 g N plant⁻¹) associated with enhanced biomass accumulation, whereas supra-optimal N (≥ 40 g N plant⁻¹) conferred no additional growth gains and coincided with a substantial depletion of root starch reserves. This starch depletion, coupled with elevated N:P ratios (> 16) indicative of progressive P limitation under high N, suggests that excessive fertilization may compromise seedling quality and stress tolerance. However, these interpretations should be viewed with caution. We did not quantify C assimilation rates, dark respiration, or the enzymatic activities governing starch–sugar interconversion, nor did we assess soil microbial activity, pH shifts, or rhizosphere processes – all of which could mediate plant responses to N fertilization. Importantly, our study was conducted under controlled greenhouse conditions using a single soil type, and the pot environment inherently restricts root development and may alter nutrient dynamics compared to open-field situations. Consequently, the generalizability of our findings is limited, and the tentative recommendations (10–20 g N plant⁻¹) require rigorous validation under operational nursery and field conditions before practical implementation. Long-term field trials across diverse soil types and climatic regimes, incorporating comprehensive physiological and edaphic measurements are essential to confirm these thresholds, establish mechanistic causality, and provide a robust basis for refining nutrient management in P. mume nursery production.

Conclusion

This study comprehensively analyzed the effects of five N levels (0–40 g N plant⁻¹) on morphological traits, C:N:P stoichiometry, and NSC pools in P. mume seedlings, with particular attention to organ-specific responses in roots, stems and leaves. Moderate N supply (10–20 g N plant⁻¹) maximized total biomass (18.2–19.6 g plant⁻¹), whereas supra-optimal N (40 g N plant⁻¹) reduced root starch by up to 48% without further growth gains. Across all organs, N addition consistently increased N content and N:P ratios, but decreased P content. In T3 and T4, leaf and root N:P ratios exceeded 16, indicating progressive P limitation. Soluble sugars (glucose, fructose, sucrose) increased with N supply, while starch content declined, particularly in roots, suggesting a reallocation of C away from long-term storage toward immediate metabolic demands. Correlation and cluster analyses further revealed that root length and total biomass clustered together at low dissimilarity, indicating coordinated growth between above- and below-ground organs under low to moderate N additions. For nursery production of P. mume seedlings intended for afforestation or agroforestry, we tentatively suggest that N application within 10–20 g N plant⁻¹ may be beneficial, provided that this is validated under actual nursery and field conditions before practical implementation. Routine soil P monitoring is also recommended to avoid nutritional imbalance. High N levels (≥ 40 g N plant⁻¹) appeared detrimental in this experimental setting and should be avoided. These findings are necessarily constrained by the controlled greenhouse conditions using a single soil type, field trials and longer-term observations are essential to confirm their applicability. Moreover, direct measurements of C flux, soil processes, and key physiological parameters are required to establish causal relationships. Nevertheless, the consistent stoichiometric patterns observed across the N gradient provide a useful framework for improving seedling quality of P. mume seedlings and for predicting growth responses in cultivation systems.