Increasing Soil Organic Carbon but Decoupling of Ecological Attributes after Loss of Dominant Functional Groups in Alpine Meadow

Sampling was conducted from August to late September 2023. Within each plot, a 0.25 m×0.25 m quadrat was used for sampling. To ensure randomness and minimize edge effects, two sampling points were randomly selected within each quadrat, avoiding the edges. All plants within the quadrat were clipped with scissors and placed in paper bags. The aboveground biomass samples were transported to the laboratory, where they were initially heated at 105°C for 30 minutes to stop enzyme activity, then dried at 65°C for 48 hours until a constant weight was achieved. The samples were weighed using a high-precision balance to obtain accurate aboveground biomass data. In addition to the aboveground biomass collection, soil cores with a diameter of 5 cm were taken from the 0-10 cm depth to include both roots and soil. These samples were immediately sealed and transported to the laboratory for further analysis. In the laboratory, roots were separated from the soil samples using a 2 mm stainless steel sieve. The roots were washed under running water several times until no visible soil adhered. The cleaned roots were then dried at 65°C for 48 hours, until a constant weight was achieved, and weighed to calculate belowground biomass. During the root separation process, soil samples were divided into two portions: one portion was air-dried in a well-ventilated area for subsequent analysis of soil physical and chemical properties, while the other portion was stored at 4°C for analysis of organic carbon and other sensitive active components.Soil water content (SWC, g·kg⁻¹) was measured using the gravimetric method by drying fresh soil samples at 105°C for 24 hours until a constant weight was achieved(Xie et al. 2017). The water loss was calculated based on the initial weight of the soil sample； Soil pH was measured using a calibrated pH meter in a soil-to-water suspension with a ratio of 2.5:1. Deionized water was boiled before use to ensure the removal of dissolved gases, which could otherwise affect pH measurements (Haghverdi and Kooch 2019); Total nitrogen (TN, g·kg-1) was determined using the Kjeldahl digestion method, which involves the conversion of organic nitrogen to ammonium. The ammonium was then quantified using a spectrophotometer at the appropriate wavelength (Barbano et al. 1990); Soil peroxidase (PX, U·g⁻¹) activity was determined by measuring the enzyme-catalyzed oxidation of organic substrates to quinones, which exhibit a characteristic absorption peak at 430 nm. This method provides insight into the oxidative capacity of the soil and its potential to decompose organic matter (Saiya-Cork, Sinsabaugh, and Zak 2002); Soil organic matter was determined using the potassium dichromate oxidation method. Fe³⁺ and Al³⁺ (mg·kg⁻¹) were determined by extracting soil samples with 0.1 M FeCl3 or AlCl3. After filtration, spectrophotometric analysis was conducted: Fe³⁺ was measured using 1,10-phenanthroline at 595 nm, and Al³⁺ was measured using pyrocatechol violet at 430 nm (Sharpley and Smith 1994).The soil organic matter values were then converted to soil organic carbon (SOC, g·kg⁻¹) using a standard conversion factor of 0.58(Krieger 2001); Microbial biomass carbon (MBC, mg·kg⁻¹) was determined by the fumigation-extraction method (Brookes et al. 1985). Approximately 4 grams of fresh soil were fumigated with chloroform for 24 hours, then extracted with 0.5 M K₂SO₄ using an overhead shaker at room temperature for 1.5 hours. The extracts were then filtered through paper filters and analyzed for soluble organic carbon. Dissolved organic carbon (DOC, mg·kg⁻¹) was measured by shaking a mixture of soil and deionized water at a 1:4 soil-to-water ratio for one hour. The mixture was then filtered through 0.45 μm cellulose acetate filters, and DOC content was quantified using a multi N/C 2100 automatic analyzer (Jones and Willett 2006). Additionally, we calculated the contribution rates of active fractions of soil organic carbon (SOC) to total SOC. Specifically, the contribution of dissolved organic carbon (DSOC, %) to SOC was determined by dividing the content of dissolved organic carbon by the total SOC content. Similarly, the contribution of microbial biomass carbon (MSOC, %) to SOC was calculated by dividing the microbial biomass carbon content by total SOC. Furthermore, the combined contribution of microbial biomass carbon and dissolved organic carbon (MDSOC, %) to SOC was derived by dividing the sum of these two fractions by the total SOC content.