**4. Discussion**

### *4.1. Patterns of Leaf Stoichiometry of C. paliurus across China*

The ranges of both leaf N and P concentrations and N:P ratio of *C. paliurus* populations were smaller than those species of Juglandaceae family that have been studied previously (*Carya cathayensis* Sarg. in China and *Juglans nigra* Linn. in the USA) [38,39], and are also smaller than those of China and the global flora (Table 4) [6,7]. Leaf mean N concentration was also smaller than that of China and the global flora, which might be due to the relatively lower nitrogen resorption capacity in woody plants than that of herbs [7,40]. Previous studies have reported that leaf P concentration in China was much lower than that at the global level, which was confirmed in this study [6,7]. Lower leaf P concentrations

in China have been demonstrated to be related to the lower soil available P concentration (3.83 mg kg−<sup>1</sup> for China vs 7.65 mg kg−<sup>1</sup> for the globe) [7,41,42]. Thus, soil conditions in *C. paliurus* populations, especially soil available P, needs to be further studied to confirm this internal relationship. Leaf N concentrations were significantly and positively correlated with leaf P concentrations in *C. paliurus* populations, which was inconsistent with previous studies [43]. This is mainly because N and P elements have similar biochemical pathways in plants, of which both are necessary in the basic biological processes such as protein synthesis, cell division and photosynthesis [44].

**Table 4.** Leaf N and P concentrations and N:P ratio for *C. paliurus* in the present study and other species of Juglandaceae family studied for China's and the global flora.


Leaf N:P is an important index that reflects the limitation of nutrition in plants and ecosystem. For different plants and vegetation in different regions, the threshold of element restriction shown by N:P value is always distinct [9,45]. Based on the effects of N:P ratios on chlorophyll yield, Koerselman and Meuleman have proposed that a leaf N:P <14 indicates N limitation, and leaf N:P >16 indicates P limitation in plants [46]. The mean value of leaf N:P (10.51) over all *C. paliurus* populations suggested that in general *C. paliurus* forests were limited by N supply. However the LWS (Laowangshan) site had an leaf N:P ration of 18.39, which may indicate that it was limited by the supply of P although the concentration of total P in soil was not low compared to other sites (Table 2). The unusually high N:P ration at the LWS site might be due to the differences of plant growth rate caused by microclimate in *C. paliurus* forests.

### *4.2. Leaf N and P Stoichiometry in Relation to Soil Properties, Geographic Origins and Climate Factors*

The results from our study showed that leaf stoichiometry of *C. paliurus* populations displayed significant relationships with geographic and climate factors. Meanwhile, results indicated that there were no significant correlations between leaf stoichiometry and the soil properties studied (including soil C, N and P concentrations). Our findings were quite different from those of Aerts and Chapin; they demonstrated that there was a significant correlation between leaf P and soil P concentrations [47]. This might be due to the differences in the species studied and their geographical distributions. Leaf N concentration increased with increasing latitude and decreasing MAT (Figures 3 and 4), which followed similar trends found in China's and the global flora [6,7,43]. Our study further demonstrated that leaf N concentration and N:P ratios increased with decreasing mean temperature in January (MTmin) (Figure 4C) and mean annual frost-free period (MAF) (Figure 5B). Together, these results confirmed that temperature-physiological stoichiometry with a latitudinal trend hold true at both global and regional levels [6]. Plants from colder habitats usually have higher values of leaf nutrient (N and P concentrations included) [11], which has been considered an adaptation mechanism to enhance their growth rates under lower temperatures [48,49].

A longitude-P relationship was also observed in *C. paliurus* populations across China (Figure 3E). Our study indicated that leaf P concentration increased with decreasing longitude. This result confirmed the biogeochemical hypothesis, which assumes that soil nutrient conditions, which are affected by leaching effects, drive the variation of plant nutrients [14]. Soil P concentration in China has been reported to decrease from northwest to southeast due to the variation of water distribution caused by precipitation [50,51]. Leaf N and P concentrations have been found strongly correlated with MAP at the global level [12]. However, a weak relationship between leaf P concentration and MAP

was found in our study, which might be due to the small range of MAP (1093–2371 mm) in *C. paliurus* populations. Wu et al. also demonstrated that leaf N concentration and N:P ratios of *Quercus acutissima* Caruth. increased with increasing longitude in China [52]; however, this was not observed in our study. This difference further reveals that the responses of leaf stoichiometry to climate factors in plants are species specific. Overall, these results indicated that variation in leaf stoichiometry for a given species was more complicated, and these differences were mainly driven by mean annual temperature and water availability associated with geographic variation.
