Tunnel construction-induced changes decrease chemical stability of soil organic carbon in the Eastern Tibetan Plateau

Tunnel construction-induced changes decrease chemical stability of soil organic carbon in the Eastern Tibetan Plateau | Authorea try { document.documentElement.classList.add('js'); } catch (e) { } var _gaq = _gaq || []; _gaq.push(['_setAccount', 'G-8VDV14Y67G']); _gaq.push(['_trackPageview']); (function() { var ga = document.createElement('script'); ga.type = 'text/javascript'; ga.async = true; ga.src = ('https:' == document.location.protocol ? 'https://ssl' : 'http://www') + '.google-analytics.com/ga.js'; var s = document.getElementsByTagName('script')[0]; s.parentNode.insertBefore(ga, s); })(); Skip to main content Preprints Collections Wiley Open Research IET Open Research Ecological Society of Japan All Collections About About Authorea FAQs Contact Us Quick Search anywhere Search for preprint articles, keywords, etc. Search Search ADVANCED SEARCH SCROLL This is a preprint and has not been peer reviewed. Data may be preliminary. 8 October 2025 V1 Latest version Share on Tunnel construction-induced changes decrease chemical stability of soil organic carbon in the Eastern Tibetan Plateau Authors : Jincheng Xiang , Yanting Xiong , Yuzhuo Chen , Yuqing Zhang , Xiaodong Wang , Yang Xiao , Xuejun Cheng , … Show All … , Shijun Wang , Pingfeng Li , Li Ma , Guo Chen , Longxi Cao 0000-0002-2734-0641 , Mengdi Xie , Xiangjun Pei , Ahmad Latif Virk 0000-0002-1558-9642 , Lin Li , and Xiaolu Tang 0000-0001-5624-1768 [email protected] Show Fewer Authors Info & Affiliations https://doi.org/10.22541/au.175992153.30018482/v1 221 views 143 downloads Contents Abstract Information & Authors Metrics & Citations View Options References Figures Tables Media Share Abstract Tunnels construction are widely used to facilitate large transport infrastructure projects, which may lead to a significant impact on soil organic carbon (SOC) dynamics and SOC chemical stability through depleting groundwater. However, the mechanisms of the effects tunnel construction on the chemical stability of SOC remain unexplored, which has become one of the most important but the least studied field. Therefore, this study aimed to investigate how tunnel construction impacts the chemical stability of SOC in shrublands along three altitude gradients (3240, 3420 and 3600 m above sea level) in the Eastern Tibetan Plateau. The results showed that tunnel construction did not significantly change SOC and its fractions regardless of altitude levels. However, tunnel construction increased O-alkyl carbon (C) by 10-15%, and decreased alkyl C (8-13%) and aromatic C (8-12%). These changes reduced SOC disintegration (alkyl C/O-alkyl C) and increased aliphaticity (aliphatic C/aromatic C) along three altitudes, causing a decrease in the chemical stability of SOC. This phenomenon was attributed to the reduced microbial activity caused by the decrease in soil water content after tunnel construction and an increase in active functional groups due to the increase of fine root biomass revealed by structural equation model. Our finding suggests that tunnel construction had no impact on SOC content, but the reduction in chemical stability of SOC may have a profound impact on long-term SOC sequestration. These findings offered new insights in predicting long-term SOC dynamics following giant construction engineering. Tunnel construction-induced changes decrease chemical stability of soil organic carbon in the Eastern Tibetan Plateau Jincheng Xiang a, b, c , Yanting Xiong c , Yuzhuo Chen a , Yuqing Zhang a , Xiaodong Wang a , Yang Xiao a , Xuejun Cheng d , Shijun Wang d , Pingfeng Li d , Li Ma e , Guo Chen b, c , Longxi Cao b, c , Mengdi Xie c , Xiangjun Pei a , Ahmad Latif Virk a , Lin Li c, * , Xiaolu Tang a, b, c, * a State Key Laboratory of Geohazard Prevention and Geoenvironment Protection, Chengdu University of Technology, Chengdu 610059, China b Tianfu Yongxing Laboratory, Chengdu 610213, China c College of Ecology and Environment, Chengdu University of Technology, Chengdu 610059, China d Huaneng Tibet Yarlung Zangbo River Hydropower Development and Investment Co., Ltd., Lhasa 850000, Xizang, China e Chengdu Shude High School, Chengdu 610031, China Corresponding author: Lin Li, [email protected] ; Xiaolu Tang ( [email protected] ) Abstract: Tunnels construction are widely used to facilitate large transport infrastructure projects, which may lead to a significant impact on soil organic carbon (SOC) dynamics and SOC chemical stability through depleting groundwater. However, the mechanisms of the effects tunnel construction on the chemical stability of SOC remain unexplored, which has become one of the most important but the least studied field. Therefore, this study aimed to investigate how tunnel construction impacts the chemical stability of SOC in shrublands along three altitude gradients (3240, 3420 and 3600 m above sea level) in the Eastern Tibetan Plateau. The results showed that tunnel construction did not significantly change SOC and its fractions regardless of altitude levels. However, tunnel construction increased O-alkyl carbon (C) by 10-15%, and decreased alkyl C (8-13%) and aromatic C (8-12%). These changes reduced SOC disintegration (alkyl C/O-alkyl C) and increased aliphaticity (aliphatic C/aromatic C) along three altitudes, causing a decrease in the chemical stability of SOC. This phenomenon was attributed to the reduced microbial activity caused by the decrease in soil water content after tunnel construction and an increase in active functional groups due to the increase of fine root biomass revealed by structural equation model. Our finding suggests that tunnel construction had no impact on SOC content, but the reduction in chemical stability of SOC may have a profound impact on long-term SOC sequestration. These findings offered new insights in predicting long-term SOC dynamics following giant construction engineering. Keywords: tunnel construction; SOC fractions; SOC functional groups; SEM Introduction Soil organic carbon (SOC) is the largest carbon pool in terrestrial ecosystems, which is equivalent to two to three times of carbon (C) reserve in the atmosphere and plant biomass (Scharlemann et al., 2014). SOC have an important role in Earth’s ecosystem functioning and mitigation of the greenhouse effect. Small changes in SOC stocks can lead to significant fluctuations in atmospheric carbon dioxide concentrations (Lal, 2004; Paustian et al., 2016). On-field understanding of SOC changes is critical for accurately forecasting regional and global C cycle under environmental changes (Jackson et al., 2017) (Angst et al., 2021). SOC stability is defined as the resistance ability of SOC to edaphic (physicochemical and biological properties of soil) environmental changes i.e., rainfall and temperature anomalies (Yang et al., 2020) (Abdalla et al., 2018; Amelung et al., 2020). Sollins et al. (1996) proposed three mechanisms of SOC stability: (1) the recalcitrance SOC form, which is a molecular characteristic and highly stable; (2) surface interactions and adsorption; and (3) aggregate protection, protects SOC from decomposition. A significant number of investigations have indicated that SOC stability is affected by a variety of anthropogenic drivers, such as revegetation (Zhang et al., 2022), biochar application (Sarker et al., 2018), nitrogen input (Chen et al., 2022), and human engineering activities (Zeng et al., 2020). However, how human activities, such as tunnel construction, can affect SOC stability has not been fully understood. The SOC chemical functional groups are key indicator for chemical stability (Panettieri et al., 2015; Wang et al., 2012), and closely associated with SOC stock changes (Liu et al., 2023; Wang et al., 2012). Previous studies demonstrated that alkyl C is a relatively stable and recalcitrant component of SOC, deriving from suberin and cutin (Ussiri and Johnson, 2003). Conversely, O-alkyl C constitutes a significant proportion of SOC and derives from easily decomposable substances like cellulose (Fang et al., 2017). Aromatic C, usually associated with aromatic structures, represents a recalcitrant fraction and difficult for microbes to degrade (Liu et al., 2023). Wang et al. (2004) showed that the main structural changes during disintegration reduces O/N-alkyl C and di-O-alkyl C, and enhances alkyl C, aromatic C, phenolic C and COO/NC = O groups, indicating the loss of proteinaceous and carbohydrate materials, and the accumulation of recalcitrant materials containing a high proportion of aromatic C. Conservation tillage has been observed to enhance the relative abundance of labile carbon functional groups, including O-alkyl C and di-alkyl C, while reduced the comparative content of alkyl and aromatic carbons (Zhang et al., 2017). Other studies found that aromaticity showed no significant trend with soil depth, while both aliphaticity and the combined index of aliphaticity and aromaticity in NMR spectra gradually increased with increasing soil depth (Hou et al., 2019). The lower microbial activity and biomass decreased the degradation of alkyl C, while increases in microbial biomass favored the production and accumulation of O-alkyl C (Chen et al., 2004). It has been found that different human activities and land types affect SOC functional groups (Carvalho et al., 2009; Deng et al., 2019). However, the changes in different SOC chemical functional groups (as abovementioned alkyl C and O-alkyl C, etc.) in response to human activities i.e., tunnel construction, remain unknown, which has become one of the most importance but the least study field in SOC stability. Tunnels construction is an integral part of China’s transportation industry (Ma et al., 2022). Meanwhile, tunnel construction is complex, costly, and has many environmental impact (Kaewunruen et al., 2020), including water erosion (Xu et al., 2022), soil water content (SWC) depletion (Liu et al., 2019), and overflow capacity decline in rivers and tributaries along the tunnels (Liu et al., 2015). Changes in SWC may affect microbial activity and vegetation growth, which are closely linked to SOC formation and transformation (Lindahl and Tunlid, 2015). The Tibetan Plateau, as the “Third Pole”, encompasses the largest alpine permafrost on earth (Mu et al., 2015). This permafrost harbors substantial quantity of SOC within terrestrial ecosystems (Ding et al., 2016). However, climatic variation and human activities have impacted structure and functionality transformations of alpine grassland ecosystem (Li et al., 2021). In recent years, rapid development of the western region, giant construction projects, such as the Qinghai-Tibet Railway and the West-East Natural Gas Pipeline, have been intensively launched on the Tibetan Plateau (Zhang et al., 2015) and no study findings have been reported on SOC chemical stability under tunnel construction . To fill this knowledge gap, we hypothesized that tunnel construction changes SWC and affects vegetation growth, which would reduce the chemical stability by decreasing plant driven inputs of SOC. The specific objective of this study was to identify the effects of tunnel construction on chemical stability of SOC by measuring SOC fractions and chemical functional groups. Moreover, a structural equation model was used to explore the key drivers to affect SOC chemical stability after tunnel construction. This research finding can provide an indication for ecological conservation through SOC stabilization in tunnel construction affected areas. 2. Materials and methods 2.1. Site description The study area is located at the eastern edge of Tibetan Plateau, in the transition zone between the mountainous area on the western edge of the Sichuan Basin and the Tibetan Plateau, with a dominant mounded plateau and mountainous plain landscape. Regional mean annual precipitation is 824.7 mm (mainly from July to September) and the average annual temperature is 5.6℃. The soil is mainly classified as brown soil and subalpine meadow soil with dominant vegetation of alpine meadows and shrublands ( Quercus aquifolioides Rehde and Rhododendron lapponicum ). Tunnel construction was started in 2020 and scheduled to be completed within 10 years. The length of the tunnel is about 19.8 km. Shotcrete and anchor rods are always used as supporting materials in the tunnel construction in China (He et al., 2022). Fig. 1 . Location of study area and distribution of the sample plots, which is on the eastern edge of the Tibetan Plateau, TA1, TA2, TA3 and CK1, CK2, CK3 represent tunnel-affected and control areas at different altitudes, respectively. Maps source from Ovi Interactive Map (V9.7.1 X64). 2.2. Experimental design and sampling The long-term fixed experimental sample sites were established in 2022 at elevations of 3240, 3420, and 3600 m directly above the tunnel, designated as TA1, TA2, and TA3, respectively. (Fig. 1). To analyze the full extent of potential effects, each sample plot was set at a vertical distance of 60, 240 and 420 meters from the tunnel entrance. Meanwhile, three control plots, named CK1, CK2, and CK3, in tunnel unaffected areas were established on a separate hill with similar altitude, vegetation type, and plot characteristics (Fig. 1). Each experimental plot was divided into three replicated sub-plots with a radius of 5 meters. A buffer zone of 5-10 meters were set between two plots. To ensure comprehensive sampling, three samples were collected at a distance of 2.5 meters from plot center at direction of 0° (starting from the north direction), 120° and 240°. In August 2022, bulk soil samples were randomly collected from 0-10 cm and 10-30 cm soil layers from each plot using stainless steel soil samplers for the soil physical properties. Subsequently, soil samples from each plot were uniformly mixed to create a representative sample following the removal of roots and other impurities. The collected samples were stored in polyethylene bags. The fresh soil samples were sieved through a 2.5 mm mesh, and subsequently stored at -4°C. Prior to analysis, a portion of the sample was dried at 105°C for soil moisture determination. The remaining soil samples were air dried at room temperature, and grounded to pass through a 0.15 mm sieve for subsequent laboratory analyses. 2.3 SOC and its fractions SOC content was determined using the K 2 Cr 2 O 7 volumetric method (Bao, 2000). Briefly, air-dried soil samples weighing between 0.1-1.0 g and passing through a 0.15 mm sieve were subjected to digestion using a mixture of 5 ml of K 2 Cr 2 O 7 (0.8 mol L -1 ) and 5 ml of H 2 SO 4 (1.84 g ml -1 ). Subsequently, the digested solution underwent titration with a standardized 0.2 mol L -1 FeSO 4 solution, which was prepared by mixing with 15 ml concentrated H 2 SO 4 per liter to prevent oxidation (Zhang et al., 1999). Soil reactive organic carbon (ROC) was analyzed with the acid hydrolysis method (Belay-Tedla et al., 2009). Soil microbial biomass carbon (MBC) was determined by using chloroform fumigation extraction method (Joergensen et al., 2011). Dissolved organic carbon (DOC) content was extracted through high purity deionized water and measured by total organic carbon analyzer (Alburquerque et al., 2013). Excretion of organic carbon (EOC) was determined using 333 mmol L −1 K 2 SO 4 oxidation-colorimetry method (Blair et al., 1995). At the same time, we assessed SWC and pH . The collected soil stored in plastic bags at low temperature, thawed and washed immediately, retained fine roots < 2 mm, and dried in an oven at 65°C until constant weight to estimate soil fine root biomass (FRB) (Tang et al., 2016). 2.4. Solid-state 13 CNMR spectroscopy Solid-state 13 CNMR spectroscopy was conducted to characterize chemical composition of SOC. Prior to NMR analysis, soil samples were pretreated with hydrofluoric acid (HF) and the samples were shaken, centrifuged, and the supernatant was removed and dried before being sieved for machine determination (Liu et al., 2023; Schmidt et al., 1997). Chemical shift refers to difference in resonance frequencies in organic compounds under the nuclear magnetic resonance conditions to determine the different SOC chemical functional groups (Li et al., 2015). In our study, the spectrum was divided into four chemical shift regimes assigned as follows: 0-45 ppm, alkyl C; 45-110 ppm, O-alkyl C; 110-165 ppm, aromatic C; 165-220 ppm, carboxyl C (Wang et al., 2022). SOC chemical stability was estimated by eq (1) and eq (2) using SOC functional groups (Wagai et al., 2013). \begin{equation} \begin{matrix}\frac{Aliphatic\ C}{\text{Aromatic\ C}}=\frac{Alkyl\ C+O‐alkyl\ C}{\text{Aromatic\ C}}\#\left(1\right)\\ \end{matrix}\nonumber \\ \end{equation}\begin{equation} \begin{matrix}\frac{Hydrophobic\ C}{Hydrophilic\ C}=\frac{Alkyl\ C+Aromatic\ C}{O‐akyl\ C+Carbonyl\ C}\#\left(2\right)\\ \end{matrix}\nonumber \\ \end{equation} 2.5. Statistical analyses Statistical analyses were conducted using R 4.2.3 (R Core Team, Auckland, NZL) software package with the inclusion of ”ggplot2”, ”pheatmap”, and ”corrplot” packages. Significance between treatment groups was determined through a three-way ANOVA, followed by least significant difference (LSD) test (p < 0.05). To understand the hypothetical pathways links of soil physicochemical properties, SOC fractions, and SOC chemical composition; structural equation modeling (SEM) was employed in R. The goodness of fit of the model was evaluated using χ2-test, comparative fit indices (CFI), probability level (p), and mean errors of approximation (RSMEA). Results 3.1. Soil properties Tunnel construction decreased SWC (p = 0.001; Fig. 2a) and increased FRB (p = 0.022; Fig. 2c), while it did not affect soil pH. SWC exhibited an increasing trend with altitude (p = 0.003; Fig. 2a). FRB decreased with increasing soil depth (Fig. 2c). Soil pH demonstrated significantly higher levels at an altitude of 3420 m compared to 3240 m and 3600 m altitudes (p = 0.001; Fig. 2b). Furthermore, soil pH was lower at 0-10 cm than 10-30 cm depth (p = 0.036; Fig. 2b). Fig. 2 . Changes in soil water content (SWC), pH and fine root biomass (FRB) under tunnel construction. TA and CK indicate the treatment area and control area, and TA1, TA2 and TA3 indicate sites at 3240m, 3420 m and 3600 m, respectively. *: p < 0.05, **: p < 0.01, ***: p < 0.001. Error bars represent standard errors (n = 3). Table 1 . P-values of treatment (T), altitude (A) and soil depth (D) on soil physicochemical properties, SOC fractions, soil functional groups, and soil chemical index using three-way ANOVA. Factor A D T A×D A×T D×T A×D×T SWC 0.003 0.420 0.001 0.504 0.419 0.931 0.919 pH 0.001 0.036 0.994 0.971 0.597 0.658 0.789 FRB 0.006 ＜0.001 0.022 0.946 0.595 0.892 0.950 SOC ＜0.001 ＜0.001 0.281 0.453 0.451 0.870 0.740 ROC 0.004 ＜0.001 0.423 0.409 0.497 0.402 0.986 DOC ＜0.001 0.008 0.322 0.584 0.420 0.455 0.501 EOC ＜0.001 0.002 0.839 0.824 0.352 0.671 0.582 MBC 0.001 ＜0.001 0.852 0.620 0.723 0.496 0.240 Alkyl C ＜0.001 0.297 0.013 0.141 0.002 0.391 0.708 O- Alkyl C 0.026 0.217 < 0.001 0.444 < 0.001 0.550 0.862 Aromatic C 0.685 0.907 < 0.001 0.511 0.725 0.314 0.678 Carboxyl C 0.004 0.885 0.386 0.861 0.680 0.318 0.581 Alkyl C/O-alkyl C 0.002 0.146 ＜0.001 0.095 < 0.001 0.753 0.938 Aromatic C/O-alkyl C 0.331 0.641 < 0.001 0.931 0.058 0.277 0.661 Aliphatic C/Aromatic C 0.466 0.854 < 0.001 0.504 0.781 0.278 0.586 Hydrophobic C/Hydrophilic C ＜0.001 0.163 < 0.001 0.225 ＜0.001 0.937 0.980 Note: SWC: soil water content, FRB: fine root biomass, SOC: soil organ carbon, EOC: easily oxidizable carbon, DOC: dissolved organic carbon, MBC: microbial biomass carbon, ROC: recalcitrant organic carbon. A: altitude, D: depth, T: treatment. 3.2. SOC and its labile fractions Tunnel construction did not impact on SOC and its fractions (Table 1). However, variations in soil depth and altitude had a significant influence on changes in SOC and its fractions. Specifically, SOC and its fraction content at 0-10 cm soil layer was significantly higher than that of 10-30cm soil layer (Table 1; Fig. 3). Additionally, SOC and its fraction content at an altitude of 3420 m was significantly lower than that at altitudes of 3240 m and 3600 m (Table 1; Fig. 3). Fig. 3 . Changes in SOC: soil organic carbon, EOC: easily oxidizable carbon, DOC: dissolved organic carbon, MBC: microbial biomass carbon, ROC: recalcitrant organic carbon. TA and CK indicate the treatment area and control area, and TA1, TA2 and TA3 indicate sites at 3240m, 3420 m and 3600 m, respectively. *: p < 0.05, **: p < 0.01, ***: p < 0.001. Error bars represent standard errors (n = 3). 3.3. Functional forms of SOC Soil alkyl C (p = 0.013; Fig. 4a) and aromatic C (p < 0.001; Fig. 4c) contents of TA were significantly lower than that of CK, while the O-alkyl C content was significantly higher than that of CK (p < 0.001; Fig. 4b). Altitude significantly impacted soil alkyl C (p < 0.001; Fig. 4a), O-alkyl C (p = 0.026; Fig. 4b) and carboxyl C (p = 0.004; Fig. 4d). There were no significant differences in SOC functional groups across different soil layers. Notably, the interactions between tunnel construction and altitude had a significant effect on soil alkyl C (p = 0.002; Fig. 4a) and O-alkyl C contents (p < 0.001; Fig. 4b), but no significant effect on soil aromatic C (Fig. 4; Table 1) and carboxyl C contents (Fig. 4; Table 1). Fig. 4 . Changes in SOC functional groups. Where TA and CK indicate the treatment area and control area, and TA1, TA2 and TA3 indicate sites at 3240m, 3420 m and 3600 m, respectively. *: p < 0.05, **: p < 0.01, ***: p < 0.001. Error bars represent standard errors (n = 3). 3.4. Chemical stability index of SOC Tunnel construction had a significant effect on soil Alkyl C/O-alkyl C (p < 0.001; Fig. 5a), Aromatic C/O-alkyl C (p < 0.001; Fig. 5b), Aliphatic C/Aromatic C (p < 0.001; Fig. 5c) and Hydrophobic C/Hydrophilic C (p < 0.001; Fig. 5d). Altitude significantly affected Alkyl C/O-alkyl C (p = 0.002; Fig. 5a), Hydrophobic C/Hydrophilic (p < 0.001; Fig. 5d). The interactions between elevation and tunnel construction significantly, decreased soil Alkyl C/O-alkyl C (p < 0.001; Fig. 5a) and Hydrophobic C/Hydrophilic C (p < 0.001; Fig. 5d). Fig. 5 . Changes in chemical index of carbon. Where TA and CK indicate the treatment area and control area, and TA1, TA2 and TA3 indicate sites at 3240m, 3420 m and 3600 m, respectively. *: p < 0.05, **: p < 0.01, ***: p < 0.001. Error bars represent standard errors (n = 3). 3.5. Interactions between soil properties, SOC fractions, SOC functional groups and SOC chemical stability SEM analysis indicated a significant negative correlation between soil pH and SOC fractions, while SWC exhibited a significant positive correlation with SOC functional groups. Additionally, FRB was positively correlated with SOC, with pH and SWC together explained 82% of the variation in SOC content. Furthermore, SOC fractions and functional groups were negatively (p<0.05) correlated with SOC chemical stability index. and SOC displayed a negative correlation with the chemical index. The interactions of pH, FRB, SWC, SOC fractions and SOC functional groups explained 76% of the variation in the chemical stability index of SOC. Fig. 6 . Structural equation model (SEM) analysis of soil pH, FRB, SWC, SOC fractions, SOC functional groups and SOC chemical stability mechanisms and their relationships. Standardized path coefficients. Solid arrows indicate significant positive correlations, and dashed arrows indicate negative correlations. R ² values associated with response variables indicate the proportion of variation explained by relationships with other variables in correlation analysis. SWC: soil water content, FRB: fine root biomass, SOC: soil organ carbon, EOC: easily oxidizable carbon, DOC: dissolved organic carbon, MBC: microbial biomass carbon, ROC: recalcitrant organic carbon, OAC: O-alkyl C, ARC: Aromatic C, CAC: carboxyl C, A/O: Alkyl C/O-alkyl C, AR/O: Aromatic C/O-alkyl C, A/AR: Aliphatic C/Aromatic C, H/HY: Hydrophobic C/Hydrophilic C. *: p < 0.05, **: p < 0.01, ***: p < 0.001. 4. Discussion 4.1. Driving factors of SOC changes after tunnel construction Maintaining SOC balance is the interplay between carbon inputs from litter, root exudates, and root debris, as well as carbon losses from soil respiration and leaching (Benbi et al., 2015). In contrast to the findings of Liu et al. (2016) and Benbi et al. (2015), our study observed no significant changes in SOC and its labile fraction contents after tunnel construction (Fig. 2; Table 1). Studies have suggested that tunnel construction can disrupt soil structure, aquifer structure, modify water resource distribution and groundwater flow patterns, thereby affecting SOC distribution and loss (Lv et al., 2020). The tunnels excavation depletes water levels, causing a decline in SWC and plant water uptake, transitioning from predominantly relying on soil water sources during the rainy season to primarily utilizing groundwater sources during the dry season (Liu et al., 2019). A decrease in groundwater levels has the potential to reduce plant growth and even mortality. Concurrently, SOC may decrease with groundwater loss due to leaching. Additionally, land subsidence incurs damage to plant root systems, resulting in decreased growth rates and even mortality (Lei et al., 2010). These mechanisms contribute to a reduction in plant-derived SOC sources, ultimately decreasing SOC content. However, in our study, increasing FRB (p = 0.022; Fig. 2) due to the shrubland maintained SOC content despite reduced SWC (Sandoval et al., 2019). This can be attributed to the high-water stress tolerance of shrubland in our study area. Consequently, the short-term decrease in SWC did not negatively affect plant growth but instead stimulated the development of deeper root systems, improving access to water resources and leading to an increase in FRB (Sainju et al., 2017). This observation is supported by the strong positive correlation between FRB and SOC in our structural equation model analysis (path coefficient = 0.32, Fig. 6). Therefore, our study results showed that the combined effects of SWC depletion and FRB promotion may not affect SOC. Soil depth and altitude are also important factors influencing SOC content (Sun et al., 2023). Our findings are consistent with those reported by Hou et al. (2019) that SOC content and its labile fraction decrease significantly with the increasing soil depth (Fig. 3, Table 1). This can be partially attributed to the different distribution patterns of root growth and biomass allocation between above-ground and below-ground (Pang et al., 2019). It is reported that plant biomass and FRB are mainly concentrated in the top layer of the soil (Liang et al., 2021), which results in less carbon input in deeper soils than in surface layers (Jobbagy and Jackson, 2000). These findings can also be supported by a significant positive correlation between the SOC fractions content and FRB in our study (Fig. 6). Our results show a significant increasing trend of SOC with increasing altitude (Fig. 3; Table 1), which may be attributed to increasing FRB (Fig. 2; Table 1) (Bangroo et al., 2017) and decreasing temperature. 4.2. SOC stability and distribution of C functional groups Our results showed that tunnel construction decreased soil alkyl C and aromatic C contents, while increased soil O-alkyl C content (Fig. 4). As a result, the chemical stability of SOC was indicated by lower values of alkyl C/O-alkyl C, aromatic/O-alkyl C, and hydrophobic C/hydrophilic C in TA compared to CK (Fig. 5). These findings suggested that TA exhibited a lower degree of chemical stability compared to CK. Thevenot et al. (2010) have previously reported that the chemical stability of SOC is attributed to the presence of biochemically complex plant-derived carbon sources. Similarly, Carvalho et al. (2009) found that different land management practices can alter the composition of SOC functional groups. Microbial decomposition of more resistant molecules leads to an increase in more easily decomposable and smaller molecules, resulting in an increased proportion of O-alkyl C. In our study, decreased chemical stability of SOC may be explained by several factors. Firstly, the increased FRB can improve plant-derived carbon supply in the soil (Xie et al., 2012). Some studies have reported that root exudates can stimulate microbial decomposition of vegetation residue carbon, consequently influencing the composition of SOC functional groups (Liu et al., 2021), this mechanism likely serves as the predominant factor contributing to the observed increment in O-alkyl C content (Hasegawa et al., 2021). Secondly, the lower SWC in TA might have affected microbial activities associated with the breakdown of SOC functional groups, ultimately reducing SOC decomposition (Chen et al., 2004; Ma et al., 2020). The decomposition process of SOC starts with the breakdown of hydrocarbons, such as cellulose (O-alkyl C), followed by the decomposition of recalcitrant C, including alkyl C and aromatic C. Therefore, the higher soil alkyl C and aromatic C contents in CK can be attributed to lipids, fatty acids, plant aliphatic polymers (alkyl C), as well as lignin and tannins (aromatic C) (Du et al., 2014; Liu et al., 2023). These components are important indicators of SOC that exhibit high resistance to degradation and relative stability (Bonanomi et al., 2013). Overall, our findings suggest that tunnel construction does not immediately alter SOC and its labile fraction contents, but it reduces the chemical stability of SOC, which is important implications for the long-term preservation of SOC after tunnel construction. Altitude is a critical factor that influences the chemical stability of SOC. Soil alkyl C and carboxyl C contents at varying altitudes exhibit notable variations, with alkyl C concentrations decreasing and carboxyl C concentrations increasing with increasing altitude (Fig. 4; Table 1). Notably, tunnel construction can intensify these variations(Fig. 4; Table 1). These findings are in line with previous studies by Du et al. (2014) and Xu et al. (2014), while aromatic C and O-alkyl C remain stable across different elevations. This phenomenon may be attributed to lower temperatures, higher precipitation (Xie et al., 2023), and lower pH values at higher altitudes (Fig. 2), which impede decomposition rates at those elevations and result in the accumulation of recalcitrant carbon (alkyl C) at lower elevations. These findings are supported by SEM in our study (Fig. 6; Table 1). Additionally, a decline in soil hydrophobicity was observed with increasing elevation (Fig. 5; Table 1), possibly due to that tunnel construction induced changes (Fig. 5; Table 1). This phenomenon can be attributed to the predominant presence of alkyl C and aromatic C in the hydrophobic C functional groups (Zhang et al., 2013). The variations in soil hydrophobicity align with the observed changes in alkyl C along the altitude gradient (Fig. 4; Table 1). Ultimately, these findings suggest that SOC demonstrates enhanced chemical stability and resilience in low-altitude areas (Liu et al., 2021). The results suggests that stability dynamics of SOC during the construction of tunnels should be noticed at different altitudes, which is crucial to the ecological environment. 5. Conclusions Our study found that short-term tunnel construction did not significantly affect SOC content and its labile fractions content. However, tunnel construction resulted in a decrease in soil alkyl C and aromatic C content, while increasing the content of reactive O-alkyl C. The reduction in alkyl C/O-alkyl C and aromatic/O-alkyl C, causing a decline in the SOC chemical stability in tunnel area. This reduction is possibly attributed to a decrease in soil water contents and an increase in fine root biomass. These findings emphasize that SOC stability of SOC pools can be increased by optimizing the tunnel construction indced changes such as water content depletion and root biomass of shrublands. Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. Data availability Data will be made available on request. Acknowledgement This study was funded by Organized Scientific Research Project of Tianfu Yongxing Laboratory (2023KJGG06), Base and Talent Project of Tibet (XZ202401JD0003), Ongoing Engineering Project in Tibet of Huaneng, China (JC2022/D01), the Everest Scientific Research Program of Chengdu University of Technology (80000-2023ZF11410). Reference Abdalla, M., Hastings, A., Chadwick, D.R., Jones, D.L., Evans, C.D., Jones, M.B., Rees, R.M., Smith, P., 2018. 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Accumulation of organic components and its association with macroaggregation in a sandy loam soil following conservation tillage. Plant and Soil 416, 1-15. 10.1007/s11104-017-3183-3. Information & Authors Information Version history V1 Version 1 08 October 2025 Copyright This work is licensed under a Non Exclusive No Reuse License. Keywords sem soc chemical stability soc fractions soc functional groups tunnel construction Authors Affiliations Jincheng Xiang Chengdu University of Technology State Key Laboratory of Geohazard Prevention and Geoenvironment Protection View all articles by this author Yanting Xiong Chengdu University of Technology College of Ecology and Environment View all articles by this author Yuzhuo Chen Chengdu University of Technology State Key Laboratory of Geohazard Prevention and Geoenvironment Protection View all articles by this author Yuqing Zhang Chengdu University of Technology State Key Laboratory of Geohazard Prevention and Geoenvironment Protection View all articles by this author Xiaodong Wang Chengdu University of Technology State Key Laboratory of Geohazard Prevention and Geoenvironment Protection View all articles by this author Yang Xiao Chengdu University of Technology State Key Laboratory of Geohazard Prevention and Geoenvironment Protection View all articles by this author Xuejun Cheng China Huaneng Group Co Ltd View all articles by this author Shijun Wang China Huaneng Group Co Ltd View all articles by this author Pingfeng Li China Huaneng Group Co Ltd View all articles by this author Li Ma Chengdu Shude High School View all articles by this author Guo Chen Tianfu Yongxing Laboratory View all articles by this author Longxi Cao 0000-0002-2734-0641 Tianfu Yongxing Laboratory View all articles by this author Mengdi Xie Chengdu University of Technology College of Ecology and Environment View all articles by this author Xiangjun Pei Chengdu University of Technology State Key Laboratory of Geohazard Prevention and Geoenvironment Protection View all articles by this author Ahmad Latif Virk 0000-0002-1558-9642 Chengdu University of Technology State Key Laboratory of Geohazard Prevention and Geoenvironment Protection View all articles by this author Lin Li Chengdu University of Technology College of Ecology and Environment View all articles by this author Xiaolu Tang 0000-0001-5624-1768 [email protected] Chengdu University of Technology State Key Laboratory of Geohazard Prevention and Geoenvironment Protection View all articles by this author Metrics & Citations Metrics Article Usage 221 views 143 downloads .FvxKWukQNSOunydq8rnd { width: 100px; } Citations Download citation Jincheng Xiang, Yanting Xiong, Yuzhuo Chen, et al. 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