Effects of γ-polyglutamic acid on available nutrients and enzyme activities in gangue-based soil

Effects of γ-polyglutamic acid on available nutrients and enzyme activities in gangue-based soil | 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 Effects of γ-polyglutamic acid on available nutrients and enzyme activities in gangue-based soil Jing Shi, Jianhong Li, Shuojiang Song, Houhuan Liu, Guangli Chen, and 2 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-7097342/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 Purpose his study aimed to explore the effects of polyglutamic acid on the physical-chemical properties and invertase activity of gangue-based soil, clarifying how different addition amounts impact soil indicators and enzyme activities for ecological restoration. Methods ​ Soil samples with varying weathering years were collected from a gangue mountain. A potted experiment involved five polyglutamic acid treatments (0–43.2 g dissolved in 300 mL water). Soil pH, conductivity, nutrient contents, aggregate composition, and enzyme activities (urease, sucrase) were measured, followed by redundancy analysis (RDA). Results ​ Polyglutamic acid rapidly increased soil pH and modified conductivity trends. Its influence on organic matter, nitrogen, phosphorus, and potassium was complex, depending on dosage and time; optimal addition boosted organic matter and nitrogen conversion, promoted potassium availability, and altered water-stable aggregate ratios. Enzyme activities were significantly affected by dosage, with varying initial levels and trends across weathering years; overall, certain enzymes showed increased activity. RDA indicated distinct soil properties under different treatments, with enzyme impacts evolving over time. Conclusions Polyglutamic acid significantly improves gangue-based soil, offering a theoretical basis and technical reference for ecological restoration and soil quality optimization. γ-polyglutamic acid coal gangue-based soil soil physical and chemical properties enzyme activity Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 Figure 9 Introduction China is the world's largest coal producer and consumer, and the destruction of mining ecosystems has also come with it (Li and Wang 2019 ). According to statistics, China's coal mine waste accounts for about 10%-15% of raw coal production(Long et al. 2019 )。There are more than 1,500 gangue mountains formed by coal mining and coal washing waste discharge in the country, with a cumulative accumulation of 3 billion tons, covering an area of ​​5,000 hm2, and its accumulation is increasing at a rate of hundreds of millions of tons per year(Dong et al. 2024 )。 In recent years, the comprehensive utilization rate of coal gangue in China has been greatly improved, but there is still a large amount of coal gangue accumulated to form gangue mountains (Song et al. 2022 )。These gangue mountains not only occupy precious land resources, but also easily spontaneously combust, causing serious air pollution, soil quality decline, biodiversity loss, natural landscape destruction and other ecological and environmental problems, seriously threatening human health and regional ecological and economic sustainable development (Wang et al. 2022 )。Therefore, strengthening the ecological restoration and reconstruction of coal gangue heaps in mining areas has important practical significance for the entire ecosystem. Soil physical and chemical properties such as pH, electrical conductivity, organic matter, and nutrient content such as nitrogen, phosphorus, and potassium are important indicators for measuring soil quality, which directly affect soil fertility and plant growth(Liu et al. 2024 )。Soil aggregates are formed by the cementation of soil particles with organic and inorganic substances. They can store various nutrients in the soil and provide habitats for microbial activities. The number and distribution of water-stable aggregates can reflect the quality of soil structure. Therefore, the composition and stability of water-stable aggregates have become one of the important indicators for evaluating soil quality(Wilpiszeski et al. 2019)。At the same time, soil invertase activity, as an important manifestation of soil biological activity, reflects the vigorous degree of material metabolism in the soil and plays a key role in the material cycle and energy flow of the soil ecosystem (Alkorta et al. 2003 ) 。Polyglutamic acid (γ-PGA) is a biopolymer with super hydrophilicity and water retention, showing great potential in the fields of agriculture and soil improvement. Studies have shown that polyglutamic acid can improve soil structure, increase soil water and fertilizer retention capacity, and promote plant growth (Hong et al. 2024 )。However, there is still a lack of research on the effects of polyglutamic acid on the physical and chemical properties and invertase activity of gangue-based soil. Therefore, this study aims to explore the effects of adding different concentrations of polyglutamic acid on the physical and chemical properties of gangue-based soil and soil invertase activity, and provide scientific basis and technical support for the ecological restoration and sustainable utilization of gangue-based soil. 1 Materials and methods 1.1 Soil culture experiment Gangue soil samples were collected from the Wangjiazhai Coal Mine in Liupanshui, Guizhou in 1997, 2007, and 2021. The samples were collected by the five-point method, and 5 kg of samples were collected at each sampling point and mixed evenly. When sampling, plant litter and gravel on the soil surface were first cleaned, avoiding plant roots, and the soil profile was excavated to collect soil samples. The samples were packed in nylon bags and transported back to the laboratory, and then laid flat in the laboratory for natural air drying to remove debris from the gravel in the gangue. The coal gangue was ground and passed through a 2mm sieve, and 6.5kg of soil was accurately taken for potted culture experiments, and γ-polyglutamic acid was sprayed (CK-blank group, T-experimental group: T1-added 10.8g of polyglutamic acid, T2-added 21.6g of polyglutamic acid, T3-added 32.4g of polyglutamic acid, T4-added 43.2g of polyglutamic acid; dissolved in 300mL of distilled water respectively), and each concentration was repeated 3 times, and 100mL of deionized water was sprayed every 3 days. The plants were placed in a dry and ventilated place for culture, and the moisture content was maintained at 5%; at 3d, 18d, 33d, and 48d respectively, part of the soil was placed flat in a nylon bag in the laboratory, naturally air-dried, ground and sieved, and the physical and chemical properties and enzyme activity were determined. 1.2 Determination of soil chemical properties The determination of soil pH value was carried out according to ISO ( 1994 ) standards(ISO 1994 ). Weigh the air-dried soil sample that passed through a 2mm sieve, add deionized water without carbon dioxide according to the specified soil-water ratio to make a suspension, let it stand for 30 minutes, use the standard buffer solution to calibrate the Leiji SJ-4F pH meter, insert the electrode into the suspension, and record the soil pH value after the reading stabilizes; the determination of conductivity(Corwin and Lesch 2003 ) also uses air-dried soil sample that passed through a 2mm sieve, add deionized water according to the specified soil-water ratio, shake and stand, take the supernatant for determination, use the standard solution to calibrate the Leiji DDS-308F conductivity meter, insert the electrode, and record the value after the reading stabilizes to obtain soil conductivity data; Determination of soil organic matter (Bremner 1965 ),weigh the air-dried soil sample that passed through a 0.25 mm sieve and place it in a hard test tube, add a specific oxidant mixture, boil it in an oil bath, transfer it to a conical flask after cooling, and titrate it with a standard ferrous sulfate solution. The organic matter content is calculated based on the titration volume; the method for determining soil hydrolyzable nitrogen is the diffusion method (Mulvaney and Khan 2001 ).Weigh the fresh soil sample with the same treatment and place it in the outer chamber of the diffusion dish. Add boric acid and an indicator to the inner chamber. Add sodium hydroxide to the outer chamber, seal it, and place it in a constant temperature box to convert hydrolyzable nitrogen into ammonia and absorb it with boric acid. Then use a microburette to absorb the standard acid solution for titration, and calculate its content based on the titration volume; the determination of soil total nitrogen adopts the Kjeldahl method (Sparks et al. 2020), weigh the air-dried soil sample that passed the 0.25 mm sieve and place it in a Kjeldahl flask. After digestion with concentrated sulfuric acid and catalyst, transfer it to an automatic Kjeldahl nitrogen analyzer, add alkali for distillation, and the ammonia is absorbed by boric acid and titrated with standard acid. The instrument can automatically read and calculate the total nitrogen content of the soil; Determination of soil available phosphorus (Han et al.), weigh the air-dried soil sample that passed the 0.149 mm sieve and digest it with acid to dissolve phosphorus in the solution. The digested solution is determined by ICP-OES (inductively coupled plasma emission spectrometer), and a standard curve is drawn in combination with the standard solution to calculate the phosphorus content; The determination method of soil potassium content is similar to the determination steps of phosphorus. After digestion, the sample is determined by ICP-OES, and its potassium content is calculated based on the standard curve. 1.3 Soil enzyme activity determination The Solarbio kit was used to determine the activities of catalase, acid phosphatase, alkaline phosphatase, sucrase, and urease in soil (Bear and Salter 1916 ): First, a certain amount of soil sample was weighed, and the corresponding buffer (acid buffer for acid phosphatase, alkaline buffer for alkaline phosphatase, and ordinary buffer for other enzymes) was added, and the supernatant was obtained as the enzyme solution after oscillation, grinding, and other operations, and then centrifuged. Next, the enzyme solution was mixed with the respective substrate solutions (hydrogen peroxide solution for catalase, disodium phenyl phosphate solution for acid and alkaline phosphatases, sucrose solution for sucrase, and urease for urea solution) at an appropriate temperature and specific acid-base conditions. After the reaction was completed, the corresponding stop solution or reagent was added (some enzyme assays required the reaction to be stopped). Finally, a color developer was added, and the absorbance was measured at a specific wavelength. The corresponding products (remaining amount of hydrogen peroxide, amount of phenol generated, amount of glucose generated, and amount of ammonia generated) were calculated based on the standard curve to obtain the activity of each enzyme. 1.4 Determination of soil enzyme aggregate ratio Soil microaggregates (Six et al. 2000 ) were determined by wet sieving, in which soil samples were placed on sieves of different apertures and oscillated or stirred in water to disperse the soil particles into aggregates of different sizes. The contents of microaggregates of different sizes were then determined by drying and weighing. The wet sieving method was also used for soil water-stable macroaggregates. The ability of soil macroaggregates to resist dispersion in water was utilized. Through a specific wet sieving operation, the macroaggregates in the soil samples were classified by particle size, and their composition and stability were then determined. 1.5 Data analysis Excel was used to organize and preliminarily calculate the experimental data. The Duncan method was used for multiple comparisons to analyze the significance of the data of different treatments. The vegan package in R software was used to perform redundancy analysis on soil physicochemical properties and enzyme activities. All software versions used in this paper were Microsoft Office 2016 and R 4.4.2. 2 Results and analysis 2.1 Effects on soil physicochemical properties 2.1.1 Effects of γ-glutamic acid on pH of gangue-based soil The results of pH analysis showed that the pH values of the blank group (CK) and the treatment groups (T1-T4) added with polyglutamic acid in each year were different at 3d, indicating that the soil pH increased rapidly after the addition of polyglutamic acid; the pH values of most treatment groups showed an overall upward trend from 3d to 48d, but some treatment groups changed more slowly, such as the T3 and T4 groups in some years, while the blank group had the opposite trend, with the pH value decreasing with time; in the soil of the same year, the different amounts of polyglutamic acid added had different effects on the pH value. Generally, the higher the amount added, the higher the pH value at certain time points. For example, in the soil of 1997, from T1-97 to T4-97, as the amount added increased from 10.8g to 43.2g, at 18d, 33d, and 48d At the same time point, the pH value tended to increase gradually; the initial pH values of the blank groups in different years were different. The results showed a pattern, that is, the pH of the blank group was the lowest in 2007, but after adding polyglutamic acid, the soil pH growth rate in 2007 was the largest under each treatment group. 2.1.2 Effect of γ-glutamic acid on the conductivity of gangue-based soil The conductivity of the blank groups in the three years increased with time, indicating that when no polyglutamic acid was added, the conductivity of the gangue-based soil increased over time. After the addition of polyglutamic acid to each experimental group, the initial conductivity was much lower than that of the blank group in the corresponding year, indicating that the addition of polyglutamic acid significantly reduced the initial conductivity of the soil. Overall, the conductivity of each experimental group showed a downward trend or an upward trend followed by a downward trend with time, and was significantly lower than the blank group during the same period, indicating that the addition of polyglutamic acid changed the change pattern of the conductivity of the gangue-based soil over time and inhibited its upward trend. Table 1 Changes in soil conductivity Treatment 3d 18d 33d 48d CK-97 1023.33 1722.00 1844.67 1905.00 T1-97 3.82 2.86 2.73 2.74 T2-97 3.55 2.92 2.84 2.77 T3-97 3.52 2.94 2.83 2.79 T4-97 4.53 2.90 2.42 2.35 CK-07 432.33 1364.00 1846.67 1915.00 T1-07 7.77 6.55 5.35 2.88 T2-07 5.84 4.66 3.76 2.43 T3-07 4.36 3.89 2.87 2.63 T4-07 2.91 2.75 2.24 1.96 CK-21 1045.33 1790.33 1901.33 1958.67 T1-21 2.95 2.86 2.56 2.37 T2-21 2.84 2.66 2.34 2.15 T3-21 4.13 3.29 2.83 2.73 T4-21 5.18 3.86 2.90 2.86 2.1.3 Effect of γ-glutamic acid on the content of organic matter, nitrogen, phosphorus and potassium in gangue-based soil (1) Soil organic matter content The results of the analysis of organic matter content showed that in different treatment groups in the same year, the addition of polyglutamic acid would cause the soil organic matter content to differ from that of the blank group. For example, in the soil of 1997, the organic matter content of the T1-97 to T4-97 groups at each time point was different from that of the CK-97 group, and the content of the treatment groups with different addition amounts also varied. In the soil of the same year, the addition of different amounts of polyglutamic acid had different effects on the soil organic matter content. The organic matter content of most treatment groups showed a trend of first increasing and then decreasing with the increase of the addition of polyglutamic acid, indicating that the moderate addition of polyglutamic acid was beneficial to increase the organic matter content. From the perspective of overall time changes, the organic matter content of some treatment groups gradually increased over time; while the content of some treatment groups increased and decreased, or the change range was small, such as the CK-97 content of the soil in 2007. In group 07, the organic matter content decreased from the 3rd day to the 18th day, and then increased from the 18th day to the 33rd day. The initial soil organic matter content of the blank groups in different years was different, and the changes over time were also different. For example, the initial content of CK-97, CK-07 and CK-21 on the 3rd day was high and low, and the change range and trend in the subsequent time were also different. This may be related to the composition, properties and environmental factors of the gangue-based soil itself in different years. (2）Soil nitrogen content The analysis results of total nitrogen show that after adding polyglutamic acid, the total nitrogen content of the soil is different from that of the blank group. There is an obvious trend that the total nitrogen content increases with the increase of polyglutamic acid application. From the perspective of time change trend, the total nitrogen content of some treatment groups gradually increases over time, while some treatment groups first decrease and then increase, or first increase and then decrease, or the change range is small. The initial total nitrogen content of the blank group in different years is different, and the change over time is also different. For example, the initial total nitrogen content of CK-97, CK-07 and CK-21 at 48 days is high and low, and the change range and trend in the subsequent time are also different. This may be related to the nitrogen storage state, microbial activity and environmental factors of the gangue-based soil itself in different years. The analysis results of hydrolyzable nitrogen show that in the same year, the different amounts of polyglutamic acid added to the treatment group had a significant effect on the soil hydrolyzable nitrogen content compared with the blank group. In most cases, with the increase in the amount of addition, the soil hydrolyzable nitrogen content was relatively higher. From the time trend, there was no uniform pattern in the changes of soil hydrolyzable nitrogen content in each treatment group over time. Some decreased first and then increased, and some fluctuated. However, the hydrolyzable nitrogen content in most treatment groups was higher on 18 days and 33 days, indicating that the microbial conversion of nitrogen was strongest at these times. (3）Soil potassium content The results of soil total potassium analysis show that in the same year, the effect of different amounts of polyglutamic acid addition on soil total potassium content is not obvious. In most cases, the total potassium content between treatment groups with different addition amounts is not much different, and there is no trend of regular change with the increase or decrease of addition amount; from the perspective of time change trend, the soil total potassium content of each treatment group changes relatively steadily over time, without a particularly significant upward or downward trend. However, the total potassium content of treatment groups in different years at different time points still fluctuates. For example, in some treatment groups in 2007, the total potassium content has a small increase and decrease from 3 days to 48 days. The analysis of soil available potassium content showed that in the soil of the same year, the effects of different amounts of polyglutamic acid added on soil available potassium content were more complicated. Except for the 3-day time point, the available potassium at other time points showed obvious regularity, that is, the soil available potassium content increased with the increase of polyglutamic acid application rate. From the time change trend, there was no uniform pattern in the changes of soil available potassium content in each treatment group over time. The content of some treatment groups increased over time, while some first increased and then decreased, or fluctuated up and down. (4）Soil phosphorus content The results of soil phosphorus content analysis show that in the same year, the effect of different amounts of polyglutamic acid addition on soil total phosphorus content is relatively complex. At the beginning of application, soil phosphorus content increased with the increase of polyglutamic acid application, and this trend weakened over time. By 48 days, phosphorus content showed only a slight increase with the increase of polyglutamic acid application. Overall, in the same year, soil phosphorus content tended to decrease over time; from the perspective of time change trend, soil total phosphorus content of each treatment group did not show a uniform increase or decrease pattern over time. Some treatment groups increased in certain time periods, while others decreased or remained relatively stable. The results of soil available phosphorus content analysis showed that in the same year, there were differences in soil available phosphorus content between the treatment group with polyglutamic acid addition and the blank group. In the soil of the same year, there was no obvious pattern in the effect of different amounts of polyglutamic acid added on soil available phosphorus content. The content of soil available phosphorus increased significantly when polyglutamic acid was just applied, but this pattern did not appear in the subsequent time. The order of available phosphorus content in the treatment groups with different addition amounts at different time points was unstable. From the perspective of time change trend, there was no obvious pattern in the change trend of soil available phosphorus content in each treatment group over time. The content of some treatment groups increased over time, while that of others decreased or fluctuated. However, there was an obvious change that the content of soil available phosphorus was significantly higher when polyglutamic acid was just applied than that of soil available phosphorus at 18, 33 and 48 days after application. 2.1.4 Effect of γ-glutamic acid on aggregates in coal gangue-based soil For soil water-stable macroaggregates, in the soil of the same year, after adding polyglutamic acid, the proportion of water-stable macroaggregates in different particle size ranges changed. Some treatment groups with added amounts increased the proportion of aggregates, while others changed the proportion of aggregates, and there was no uniform change with the increase or decrease of the amount of addition; the blank groups of different years had different initial proportions of soil water-stable macroaggregates in each particle size range, and the proportions of aggregates in different particle size ranges were different among the treatment groups, but the particle size proportions did not show regularity; Overall, the aggregates with the smallest particle size (5mm) accounted for the largest proportion. For soil water-stable microaggregates, compared with different treatment groups in the same year, after adding polyglutamic acid, the proportion of soil water-stable microaggregates in each particle size range was different from that in the blank group. The treatment groups with some addition amounts would increase or decrease the proportion of microaggregates in certain particle size ranges, but did not show a uniform pattern of change with the addition amount. The blank groups in different years had different initial proportions of soil water-stable microaggregates in each particle size range, but did not show obvious differences between years. Overall, aggregates with a particle size range of 0.01 - 0.05 mm and > 0.25 mm accounted for the largest proportion. 2.2 Effect of γ-glutamic acid on the activity of invertase in coal gangue-based soil In different years, the urease activity of the CK group mostly showed a downward trend over time, but the degree of decline was different in each year; compared with the CK group, the changes in urease activity were different after adding polyglutamic acid, but the main manifestation was that except for the T1 treatment group, the soil urease activity of other treatment groups decreased over time, but the speed and magnitude of the decline were different in different treatment groups, indicating that the addition of polyglutamic acid would affect the soil urease activity; between different years, the initial value and change trend of soil urease activity in different treatment groups were different, but there was no regularity. In different years, the sucrase activity of the CK group generally showed an upward trend over time, but the increase and speed were different in each year. Compared with the CK group, after adding polyglutamic acid, the activity growth of each treatment group was different, and the activity differences between treatment groups with different addition amounts were also inconsistent at different time points, but overall, the soil sucrase activity increased over time, and the application of polyglutamic acid increased the activity of soil sucrase, indicating that the addition amount and time factors of polyglutamic acid will affect the soil sucrase activity; in different years, the initial activity of each treatment group was relatively low in 1997, and then the increase was more obvious. In 2007, the initial activity was unevenly distributed, and the subsequent changes were also different. The initial activity in 2021 was relatively medium, and the change trend was also different from the previous two years. In different years, there was no uniform pattern in the changes of catalase activity in the CK group. Compared with the CK group, after adding polyglutamic acid, the catalase activity changed in various ways. The trends and amplitudes of activity changes in each treatment group were different, and the differences between treatment groups with different addition amounts were different at different time points. The soil enzyme activity was the largest at 18 or 33 days, and except for 2021, the application of polyglutamic acid increased soil catalase activity in other years, indicating that the amount and time of polyglutamic acid addition would significantly affect the activity of soil catalase; the initial values and change trends of soil catalase activity were significantly different between different years. In 1997, the initial activity distribution of each treatment group was relatively scattered, and the amplitude of change was large; in 2007, the initial activity was relatively concentrated, and the changes were more complex; in 2021, the initial activity was different, and the activity of some treatment groups increased significantly in the later period, indicating that the intrinsic properties of different soil years had different effects on catalase activity. The initial values and changing trends of soil acid phosphatase activity in different years were quite different. The comparison between treatment groups showed that the changes in acid phosphatase activity in the CK group were different in different years. It showed a downward trend in 1997, first stabilized and then increased in 2007, and fluctuated in 2021, reflecting that the soil's own factors had different effects on the enzyme activity in different years. Compared with the CK group, the changing trends and amplitudes of the activities of each treatment group after adding polyglutamic acid were different, and the activity differences between treatment groups with different addition amounts were also inconsistent at different time points, but most treatment groups showed a trend of first increasing and then decreasing over time, and the enzyme activity of the T2 treatment group was relatively high, indicating that the addition of polyglutamic acid under the T2 condition of polyglutamic acid would significantly increase the enzyme activity. The results between different years showed that the initial activity of each treatment group was unevenly distributed in 1997, and the subsequent changes fluctuated significantly; the initial activity was low in 2007, and some treatment groups showed an upward trend afterwards; the initial activity was different in 2021, and the change trends of each treatment group in the later period were diverse, indicating that there were significant differences in soil phosphatase activity in different years. In different years, the alkaline phosphatase activity of the CK group changed differently. In 1997, it first increased and then decreased, in 2007 it was high at first and then low with large fluctuations, and in 2021 it first increased and then decreased, indicating that the soil's own characteristics in different years have different effects on the enzyme activity; compared with the CK group, after the addition of polyglutamic acid, the change trends and amplitudes of the activity of each treatment group were different, and the activity differences between treatment groups with different addition amounts were also different at each time point, but the alkaline phosphatase activity of the treatment group reached a peak at 18 or 33 days, but regardless of the application amount, the alkaline phosphatase activity of most treatment groups was lower than that of the blank group, indicating that the addition of polyglutamic acid will reduce the activity of alkaline phosphatase in the soil, and the effect of the addition amount of polyglutamic acid on enzyme activity will change over time. 2.3 Redundancy analysis of soil physicochemical properties and invertase activity RDA analysis was performed with soil physicochemical properties after adding different concentrations of γ-polyglutamic acid as the response variable and soil carbon, nitrogen and phosphorus cycle-related enzyme activities as the explanatory variables. The points of each treatment group are scattered in the figure, indicating that there are differences in the comprehensive properties of the soil under different treatments. The 3d RDA analysis results showed that there was a significant correlation between the enzymes urease, sucrase, catalase, and alkaline phosphatase; catalase and acid phosphatase had a greater impact on soil physicochemical properties, followed by alkaline phosphatase, and the other two enzymes had a lower explanatory rate; at the same time, the enzyme activity of the soil in 1997 and 2007 with polyglutamic acid added was higher, and the enzyme activity of the blank group and the treatment group in 2021 was lower. The results of the 18-day RDA analysis showed that, except for urease and alkaline phosphatase, there was a certain degree of correlation between other enzymes; acid phosphatase and alkaline phosphatase had a greater impact on soil physical and chemical properties, urease and sucrase had a certain impact, and catalase had the least impact; the enzyme activity of the 2007 treatment group and the 1997 T1 and T2 treatment groups was higher, and the enzyme activity of other treatment groups was lower. The results of the 33-day RDA analysis showed that there was a significant correlation between the five enzymes, especially sucrase and acid phosphatase had a very strong correlation; urease, catalase and acid phosphatase had similar effects on soil physical and chemical properties, and alkaline phosphatase and sucrase had less impact; the enzyme activity of the 2007 treatment group and the 1997 T1 and T2 treatment groups was higher, and the enzyme activity of other treatment groups was lower. The RDA analysis results of 48 days showed that there was a certain degree of correlation between the five enzymes; urease, sucrase and acid phosphatase had similar effects on the physical and chemical properties of the soil, while alkaline phosphatase and catalase had smaller effects; the enzyme activity was higher in the T1 and T2 treatment groups in 1997 and 2021. 3 Discussion 3.1 Effect of polyglutamic acid on soil physicochemical properties Polyglutamic acid can quickly increase soil pH after addition. The pH of most treatment groups showed an upward trend over time, while the blank group showed a downward trend. This may be due to the chemical properties of polyglutamic acid itself or its reaction with acid-base substances in the soil. At the same time, the addition of polyglutamic acid significantly reduced the initial conductivity of the soil, changed its change pattern over time, and inhibited the upward trend. This shows that polyglutamic acid can effectively regulate the acid-base balance and ion concentration of the soil (Skalski et al. 2024 ), which has positive significance for improving the chemical environment of coal gangue-based soil. Polyglutamic acid has a complex effect on the content of nutrients such as soil organic matter, nitrogen, phosphorus, and potassium. The addition of polyglutamic acid caused a difference in soil organic matter content compared with the blank group. The organic matter content of most treatment groups increased first and then decreased with the increase in the amount of addition, indicating that the appropriate amount of polyglutamic acid is beneficial to increase organic matter (Guo et al. 2024 ). It may be that polyglutamic acid promotes soil microbial activity and accelerates the decomposition and synthesis of organic matter(Guo et al. 2024 ). In terms of nitrogen, the total nitrogen and hydrolyzable nitrogen contents changed significantly after the addition of polyglutamic acid, and increased with the increase in the amount of addition, indicating that polyglutamic acid can improve the storage and conversion efficiency of soil nitrogen (Zhang et al. 2017a ). For potassium, the total potassium content was not significantly affected by the amount of polyglutamic acid added, but the available potassium content increased with the increase in the amount of addition except for 3 days, indicating that polyglutamic acid can promote the effectiveness of potassium(Guo et al. 2017 ). In terms of phosphorus, the total phosphorus and available phosphorus content are complexly affected by the amount and time of polyglutamic acid addition. Polyglutamic acid can increase the phosphorus content in the initial stage, but this promotion effect weakens over time, which may be related to the fixation and release mechanism of phosphorus in the soil (Zhang et al. 2017b ). The effect of γ-PGA on soil organic matter and nutrients such as nitrogen, phosphorus, and potassium is affected by many factors. It needs to be reasonably applied according to specific soil conditions to optimize nutrient utilization efficiency. The effect of γ-PGA on soil nutrients is complex due to different soil types, environmental conditions, and application methods. In some cases, γ-PGA can improve the effectiveness of soil nitrogen and plant absorption of nutrients, but its effects on soil organic matter, phosphorus, potassium and other nutrients may vary depending on specific conditions (Zhang et al. 2017a )。 3.2 Effect of polyglutamic acid on soil invertase activity Polyglutamic acid has a significant effect on the activities of soil urease, sucrase, catalase, acid phosphatase and alkaline phosphatase. Urease activity decreased over time in most treatment groups except the T1 treatment group, indicating that the amount of polyglutamic acid added would affect urease activity. It may be that polyglutamic acid changed the structure of soil microbial community and affected the growth and metabolism of urease-producing bacteria (Qiong et al. 2018). Sucrase activity increased over time, and polyglutamic acid could increase its activity, indicating that polyglutamic acid could promote the circulation and transformation of carbon in soil (O'Dowd and Hopkins 1998 ). Catalase activity varied. Except for 2021, polyglutamic acid could increase its activity, and the activity was the highest at 18 or 33 days, indicating that polyglutamic acid and time factors jointly affect the redox process of soil. The activity of acid phosphatase in most treatment groups increased first and then decreased, and the enzyme activity in the T2 treatment group was higher, indicating that a specific amount of polyglutamic acid added can significantly increase the activity of the enzyme, which is beneficial to the transformation and utilization of phosphorus in the soil (Zhang et al. 2024 ). After adding polyglutamic acid, the activity of alkaline phosphatase in most treatment groups was lower than that in the blank group, and reached a peak at 18 or 33 days, indicating that polyglutamic acid inhibits the activity of the enzyme, and its effect changes over time. The initial values ​​and change trends of soil enzyme activity in different years are significantly different, reflecting the important influence of soil properties on enzyme activity(Hou et al. 2018 )。 There are differences in the chemical composition, microbial community composition and structure of coal gangue-based soil in different years, which can affect the synthesis, stability and activity of enzymes (Sun et al. 2020 ). For example, differences in soil pH, organic matter content, etc. can affect the growth environment of microorganisms, and thus affect the enzyme activity produced by microorganisms. 3.3 Relationship between soil physicochemical properties and invertase activity RDA analysis showed that there were differences in the comprehensive properties of soil under different treatments, and the relationship between enzyme activities at different time points and between enzyme activities and soil physicochemical properties was complex (Deng et al. 2019 )。The degree of influence of different enzyme activities on soil physicochemical properties varies with time (Lan Yu et al. 2011 ). For example, catalase and acid phosphatase had a greater effect at 3d, while acid phosphatase and alkaline phosphatase had a prominent effect at 18d. There were also differences in enzyme activity between different years and treatment groups, indicating that soil physicochemical properties and invertase activity are interrelated and affect each other. Soil physicochemical properties such as nutrient content and pH will affect enzyme activity and stability, and changes in enzyme activity will react to soil material circulation and transformation processes, thereby affecting soil physicochemical properties. In summary, polyglutamic acid has a significant effect on the physicochemical properties and invertase activity of coal gangue-based soil, and is restricted by multiple factors such as addition amount, time and soil year. This study provides a theoretical basis and technical reference for the improvement and ecological restoration of coal gangue-based soils, but further in-depth research on the mechanism of interaction between polyglutamic acid and soil is still needed to optimize soil improvement measures. Conclusion This study explored the effects of polyglutamic acid on gangue-based soil from many aspects, comprehensively analyzed soil physical and chemical properties, invertase activity and the relationship between the two, and drew the following conclusions: polyglutamic acid rapidly increased soil pH, significantly reduced initial conductivity, and effectively improved soil acid-base and salinity; the appropriate addition of polyglutamic acid was conducive to the accumulation of organic matter, and the addition of polyglutamic acid could improve the storage and conversion efficiency of soil nitrogen and enhance the effectiveness of potassium. The fixation and release mechanism of soil phosphorus was jointly affected by polyglutamic acid and time; the addition of polyglutamic acid changed the structure of soil microbial community and increased soil enzyme activity; the properties of the soil itself also played a decisive role in the formation and change of enzyme activity. Polyglutamic acid has a significant improvement effect on gangue-based soil, but the improvement effect is synergistically restricted by multiple factors such as the amount of polyglutamic acid added, time and soil year. This study provides a solid theoretical basis for the ecological restoration and sustainable utilization of gangue-based soil. In the future, it is necessary to further explore the microscopic mechanism of the interaction between polyglutamic acid and soil, further optimize soil improvement strategies, and promote the effective restoration and sustainable development of the ecological environment in mining areas. Declarations Funding This study was funded by the Mountain Plateau Specialty Agricultural Products Technology Innovation Center (520202023018). Competing Interests On behalf of all authors, the corresponding author states that there is no conflict of interest. Author contribution Jing Shi, Houhuan Liu, Guangli Chen, and Xiaoxia Zhao were responsible for sample collection. Jing Shi also conducted the physical and chemical index analysis. Jianhong Li was in charge of the analysis of soil enzyme activities. Shuo Jiang Song performed the data analysis, with Lili Feng providing guidance throughout the data analysis process. Each author's efforts were integral to the completion of this study, ensuring the reliability and validity of the research findings. References Alkorta, I., A. Aizpurua, P. Riga, I. Albizu, I. Amezaga, and C. Garbisu. 2003. Soil Enzyme Activities as Biological Indicators of Soil Health. Reviews on environmental health 18 :65-73. Bear, F. E., and R. M. Salter. 1916. Methods in soil analysis. W. Va. Agr. Exp. Sta., Tech. Bul 159 :1-24. Bremner, J. M. 1965. Total Nitrogen. Pages 1149-1178 Methods of Soil Analysis. Corwin, D. L., and S. M. Lesch. 2003. Application of Soil Electrical Conductivity to Precision Agriculture. Agronomy Journal 95 :455-471. Deng, J., Y. Chong, D. Zhang, C. Ren, F. Zhao, X. Zhang, X. Han, and G. Yang. 2019. Temporal variations in soil enzyme activities and responses to land-use change in the loess plateau, China. Applied Sciences 9 :3129. Dong, Y., H. Lu, and H. Lin. 2024. Comprehensive study on the spatial distribution of heavy metals and their environmental risks in high-sulfur coal gangue dumps in China. Journal of Environmental Sciences 136 :486-497. Guo, Z., J. Wang, T. Chen, H. Zhang, X. Hou, and J. Li. 2024. Effects of γ-polyglutamic acid supplementation on alfalfa growth and rhizosphere soil microorganisms in sandy soil. Sci Rep 14 :6440. Guo, Z., N. Yang, C. Zhu, and L. Gan. 2017. Exogenously applied poly-γ-glutamic acid alleviates salt stress in wheat seedlings by modulating ion balance and the antioxidant system. Environmental Science and Pollution Research 24 :6592-6598. Han, C.-w., D.-h. Kim, and R. Bradford. ICP-OES Analysis of Phosphorus in Soils Extracted using the Lancaster Leachate Method. Hong, L., L. Wei, G. Fanglan, L. Jiao, T. Shiheng, Y. Hong, R. Yao, G. Xinyue, and Y. Can. 2024. Unveiling the regulatory mechanism of poly-γ-glutamic acid on soil characteristics under drought stress through integrated metagenomics and metabolomics analysis. Front Microbiol 15 :1387223. Hou, H., C. Wang, Z. Ding, S. Zhang, Y. Yang, J. Ma, F. Chen, and J. Li. 2018. Variation in the soil microbial community of reclaimed land over different reclamation periods. Sustainability 10 :2286. ISO. 1994. ISO 10390: 1994 Soil Quality Determination of pH. Lan Yu, L. Y., H. X. Han XiaoRi, Y. J. Yang JinFeng, W. Y. Wang Yue, F. D. Fang DaWei, and L. N. Li Na. 2011. Temporal and spatial dynamics of enzyme activities under long-term fertilization in a maize growing brown soil. Li, J., and J. Wang. 2019. Comprehensive utilization and environmental risks of coal gangue: A review. Journal of Cleaner Production 239 :117946. Liu, J., D. Wang, X. Yan, L. Jia, N. Chen, J. Liu, P. Zhao, L. Zhou, and Q. Cao. 2024. Effect of nitrogen, phosphorus and potassium fertilization management on soil properties and leaf traits and yield of Sapindus mukorossi. Frontiers in Plant Science 15 . Long, J., S. Zhang, and K. Luo. 2019. Selenium in Chinese coal gangue: Distribution, availability, and recommendations. Resources, Conservation and Recycling 149 :140-150. Mulvaney, R. L., and S. A. Khan. 2001. Diffusion Methods to Determine Different Forms of Nitrogen in Soil Hydrolysates. Soil Science Society of America Journal 65 :1284-1292. O'Dowd, R., and D. Hopkins. 1998. Mineralization of carbon from D-and L-amino acids and D-glucose in two contrasting soils. Soil Biology and Biochemistry 30 :2009-2016. Qiong, X., W. Qi-qi, W. Lei, C. An-dong, W. Chuan-jie, Z. Wen-ju, and X. Ming-gang. 2018. Fertilization impacts on soil microbial communities and enzyme activities across China’s croplands: A meta-analysis. Journal of Plant Nutrition and Fertilizers 24 :1598-1609. Six, J., E. T. Elliott, and K. Paustian. 2000. Soil macroaggregate turnover and microaggregate formation: a mechanism for C sequestration under no-tillage agriculture. Soil Biology and Biochemistry 32 :2099-2103. Skalski, T., E. Zając, E. Jędrszczyk, K. Papaj, J. Kohyt, A. Góra, A. Kasprzycka, D. Shytum, B. Skowera, and A. Ziernicka-Wojtaszek. 2024. Effects of γ-polyglutamic acid on grassland sandy soil properties and plant functional traits exposed to drought stress. Sci Rep 14 :3769. Song, W., J. Zhang, M. Li, H. Yan, N. Zhou, Y. Yao, and Y. Guo. 2022. Underground Disposal of Coal Gangue Backfill in China. Applied Sciences 12 :12060. Sparks, D. L., A. L. Page, P. A. Helmke, and R. H. Loeppert. 2020. Methods of soil analysis, part 3: Chemical methods. John Wiley & Sons. Sun, J., L. Yang, J. Wei, J. Quan, and X. Yang. 2020. The responses of soil bacterial communities and enzyme activities to the edaphic properties of coal mining areas in Central China. PLOS ONE 15 :e0231198. Wang, Q., Y. Zhao, W. Xiao, Z. Lin, and H. Ren. 2022. Assessing Potential Spontaneous Combustion of Coal Gangue Dumps after Reclamation by Simulating Alfalfa Heat Stress Based on the Spectral Features of Chlorophyll Fluorescence Parameters. Remote Sensing 14 :5974. Wilpiszeski, R. L., J. A. Aufrecht, S. T. Retterer, M. B. Sullivan, D. E. Graham, E. M. Pierce, O. D. Zablocki, A. V. Palumbo, and D. A. Elias. 2019. Soil Aggregate Microbial Communities: Towards Understanding Microbiome Interactions at Biologically Relevant Scales. Appl Environ Microbiol 85 . Zhang, L., D. Gao, J. Li, N. Fang, L. Wang, and Y. Shi. 2017a. Effects of poly-γ-glutamic acid on soil nitrogen and carbon leaching and CO2 fluxes in a sandy clay loam soil. Canadian Journal of Soil Science 97 :319-328. Zhang, L., X. Yang, D. Gao, L. Wang, J. Li, Z. Wei, and Y. Shi. 2017b. Effects of poly-γ-glutamic acid (γ-PGA) on plant growth and its distribution in a controlled plant-soil system. Scientific Reports 7 . Zhang, Y., S. Zhang, B. Zhao, Y. Li, M. Xu, Y. g. Yan, J. Jing, and L. Yuan. 2024. Glutamic Acid-Enhanced Phosphate Fertilizer Increases Phosphorus Availability in Fluvo-Aquic Soil via Phosphamide (O = P-N) Formation, Decreasing Phosphate Fixation and Increasing Soil Microbial Diversity. Journal of soil science and plant nutrition 24 :2748-2760. Additional Declarations No competing interests reported. Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. 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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-7097342","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":511381976,"identity":"1502eac1-01f9-4ecf-9394-ab6620081099","order_by":0,"name":"Jing Shi","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA3ElEQVRIie3RsQrCMBCA4QuFdIl0jQj6BEJKQRBEXyVFcIq7m5GCLoKrg+AziCCOB4XgILh2VAQnBd0VXXVq3ATz7x93xwG4XD9YECSYPu6N/txP0I4UpybeF3SHLMZG2hGBKhIFnRKdKWG5GWKNF9eeR6bqlp2hWa7qHEES3eHhllKfXZb1GbSjGuYQD9BwSRkjo+6qxADjVR6hEA85Us4B1cmOMGh74WAoBGwUtSOcG3KErZTh2ET1mbC4pbWbXFPoPWXFTw7Zudcs55LPkcz2NW/kW+FyuVx/0QtzT0WVKXOkpAAAAABJRU5ErkJggg==","orcid":"","institution":"China University of Mining \u0026 Technology","correspondingAuthor":true,"prefix":"","firstName":"Jing","middleName":"","lastName":"Shi","suffix":""},{"id":511381979,"identity":"af2f39bd-6f31-43f9-bcab-03744a69f252","order_by":1,"name":"Jianhong Li","email":"","orcid":"","institution":"Liupanshui Normal University","correspondingAuthor":false,"prefix":"","firstName":"Jianhong","middleName":"","lastName":"Li","suffix":""},{"id":511381982,"identity":"a3949998-3473-4ad6-a8db-9e98251afab4","order_by":2,"name":"Shuojiang Song","email":"","orcid":"","institution":"Liupanshui Normal University","correspondingAuthor":false,"prefix":"","firstName":"Shuojiang","middleName":"","lastName":"Song","suffix":""},{"id":511381984,"identity":"ba335955-9351-4681-99c5-480e24a48d52","order_by":3,"name":"Houhuan Liu","email":"","orcid":"","institution":"Liupanshui Vocational and Technical College","correspondingAuthor":false,"prefix":"","firstName":"Houhuan","middleName":"","lastName":"Liu","suffix":""},{"id":511381987,"identity":"887bf797-b733-4742-a960-b9b962c75389","order_by":4,"name":"Guangli Chen","email":"","orcid":"","institution":"Liupanshui Vocational and Technical College","correspondingAuthor":false,"prefix":"","firstName":"Guangli","middleName":"","lastName":"Chen","suffix":""},{"id":511381989,"identity":"6fb8a180-6537-4bde-a23f-b92dc115a2fa","order_by":5,"name":"Xiaoxia Zhao","email":"","orcid":"","institution":"Liupanshui Vocational and Technical College","correspondingAuthor":false,"prefix":"","firstName":"Xiaoxia","middleName":"","lastName":"Zhao","suffix":""},{"id":511381991,"identity":"9d9ebd41-ce71-4574-9c63-044ac0031e88","order_by":6,"name":"Li Feng","email":"","orcid":"","institution":"China University of Mining \u0026 Technology","correspondingAuthor":false,"prefix":"","firstName":"Li","middleName":"","lastName":"Feng","suffix":""}],"badges":[],"createdAt":"2025-07-11 03:38:18","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-7097342/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-7097342/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":91016585,"identity":"923414c4-d3cc-4a12-921e-267c5287e0ee","added_by":"auto","created_at":"2025-09-10 17:01:09","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":882020,"visible":true,"origin":"","legend":"\u003cp\u003eStatus of base soil appearance\u003c/p\u003e","description":"","filename":"1.png","url":"https://assets-eu.researchsquare.com/files/rs-7097342/v1/ef71966261a992c517002f3b.png"},{"id":91016583,"identity":"93816819-1850-43e1-8816-faa2bbedaca6","added_by":"auto","created_at":"2025-09-10 17:01:09","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":52030,"visible":true,"origin":"","legend":"\u003cp\u003epH content of coal gangue soil in different treatment groups\u003c/p\u003e\n\u003cp\u003eNote: Y-97 is coal gangue soil from 1997, 97, 07, 21 represent years, CK represents blank group, T1-T4 represent experimental groups, which means 10.8g, 21.6g, 32.4g, 43.2g of polyglutamic acid were added respectively, and Treatment represents different treatment groups, the same below.\u003c/p\u003e","description":"","filename":"2.png","url":"https://assets-eu.researchsquare.com/files/rs-7097342/v1/f114fcd0fb6ced2303305fb6.png"},{"id":91016584,"identity":"ea53e61e-7493-4821-b66a-d72a819586d4","added_by":"auto","created_at":"2025-09-10 17:01:09","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":76759,"visible":true,"origin":"","legend":"\u003cp\u003eOrganic matter content of coal gangue soil in different treatment groups\u003c/p\u003e","description":"","filename":"3.png","url":"https://assets-eu.researchsquare.com/files/rs-7097342/v1/e2109ee3886ec2a2c7968a32.png"},{"id":91016586,"identity":"4101a394-f1bc-43bb-b18a-6a03431f8f95","added_by":"auto","created_at":"2025-09-10 17:01:09","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":65753,"visible":true,"origin":"","legend":"\u003cp\u003eNitrogen content in coal gangue soil of different treatment groups\u003c/p\u003e","description":"","filename":"4.png","url":"https://assets-eu.researchsquare.com/files/rs-7097342/v1/5c2a1171dcc733d1fefa8e24.png"},{"id":91016589,"identity":"c2b5270c-0aee-4e89-9dcc-b748b1f4aaac","added_by":"auto","created_at":"2025-09-10 17:01:09","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":67125,"visible":true,"origin":"","legend":"\u003cp\u003ePotassium content of coal gangue soil in different treatment groups\u003c/p\u003e","description":"","filename":"5.png","url":"https://assets-eu.researchsquare.com/files/rs-7097342/v1/86208778877557c7ac7b8b9a.png"},{"id":91016596,"identity":"fd8994eb-f643-4766-8bcf-64f5a4a67d93","added_by":"auto","created_at":"2025-09-10 17:01:09","extension":"png","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":65569,"visible":true,"origin":"","legend":"\u003cp\u003ePhosphorus content of coal gangue soil in different treatment groups\u003c/p\u003e","description":"","filename":"6.png","url":"https://assets-eu.researchsquare.com/files/rs-7097342/v1/5b0d670f6a0df8ec35057242.png"},{"id":91017074,"identity":"77d18154-b013-4ac6-83b5-0778aa1b91b8","added_by":"auto","created_at":"2025-09-10 17:09:09","extension":"png","order_by":7,"title":"Figure 7","display":"","copyAsset":false,"role":"figure","size":173999,"visible":true,"origin":"","legend":"\u003cp\u003eRatio of aggregates in coal gangue soil in different treatment groups\u003c/p\u003e","description":"","filename":"7.png","url":"https://assets-eu.researchsquare.com/files/rs-7097342/v1/dea58c764eba5788d43db7bb.png"},{"id":91017071,"identity":"e8a5531c-16ca-472a-8c84-55874a987b03","added_by":"auto","created_at":"2025-09-10 17:09:09","extension":"png","order_by":8,"title":"Figure 8","display":"","copyAsset":false,"role":"figure","size":246852,"visible":true,"origin":"","legend":"\u003cp\u003eChanges in enzyme activity in coal gangue soil in different treatment groups\u003c/p\u003e","description":"","filename":"8.png","url":"https://assets-eu.researchsquare.com/files/rs-7097342/v1/fb89dd0a36248c37e9334951.png"},{"id":91016592,"identity":"e8978ab2-cdf6-43ea-ba72-98bc09faf998","added_by":"auto","created_at":"2025-09-10 17:01:09","extension":"png","order_by":9,"title":"Figure 9","display":"","copyAsset":false,"role":"figure","size":198894,"visible":true,"origin":"","legend":"\u003cp\u003eRedundancy analysis of soil physicochemical properties and invertase activity at different times\u003c/p\u003e","description":"","filename":"9.png","url":"https://assets-eu.researchsquare.com/files/rs-7097342/v1/3b83388ebde7187324095400.png"},{"id":93024490,"identity":"1e6b119c-c348-43c2-afbd-286b7d71088b","added_by":"auto","created_at":"2025-10-08 09:17:17","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":2625648,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-7097342/v1/ca215289-fd03-4449-b4f6-bee33cda32cf.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Effects of γ-polyglutamic acid on available nutrients and enzyme activities in gangue-based soil","fulltext":[{"header":"Introduction","content":"\u003cp\u003eChina is the world's largest coal producer and consumer, and the destruction of mining ecosystems has also come with it (Li and Wang \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e2019\u003c/span\u003e). According to statistics, China's coal mine waste accounts for about 10%-15% of raw coal production(Long et al. \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e2019\u003c/span\u003e)。There are more than 1,500 gangue mountains formed by coal mining and coal washing waste discharge in the country, with a cumulative accumulation of 3\u0026nbsp;billion tons, covering an area of ​​5,000 hm2, and its accumulation is increasing at a rate of hundreds of millions of tons per year(Dong et al. \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e2024\u003c/span\u003e)。 In recent years, the comprehensive utilization rate of coal gangue in China has been greatly improved, but there is still a large amount of coal gangue accumulated to form gangue mountains (Song et al. \u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e2022\u003c/span\u003e)。These gangue mountains not only occupy precious land resources, but also easily spontaneously combust, causing serious air pollution, soil quality decline, biodiversity loss, natural landscape destruction and other ecological and environmental problems, seriously threatening human health and regional ecological and economic sustainable development (Wang et al. \u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e2022\u003c/span\u003e)。Therefore, strengthening the ecological restoration and reconstruction of coal gangue heaps in mining areas has important practical significance for the entire ecosystem. Soil physical and chemical properties such as pH, electrical conductivity, organic matter, and nutrient content such as nitrogen, phosphorus, and potassium are important indicators for measuring soil quality, which directly affect soil fertility and plant growth(Liu et al. \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e2024\u003c/span\u003e)。Soil aggregates are formed by the cementation of soil particles with organic and inorganic substances. They can store various nutrients in the soil and provide habitats for microbial activities. The number and distribution of water-stable aggregates can reflect the quality of soil structure. Therefore, the composition and stability of water-stable aggregates have become one of the important indicators for evaluating soil quality(Wilpiszeski et al. 2019)。At the same time, soil invertase activity, as an important manifestation of soil biological activity, reflects the vigorous degree of material metabolism in the soil and plays a key role in the material cycle and energy flow of the soil ecosystem (Alkorta et al. \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e2003\u003c/span\u003e) 。Polyglutamic acid (γ-PGA) is a biopolymer with super hydrophilicity and water retention, showing great potential in the fields of agriculture and soil improvement. Studies have shown that polyglutamic acid can improve soil structure, increase soil water and fertilizer retention capacity, and promote plant growth (Hong et al. \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e2024\u003c/span\u003e)。However, there is still a lack of research on the effects of polyglutamic acid on the physical and chemical properties and invertase activity of gangue-based soil. Therefore, this study aims to explore the effects of adding different concentrations of polyglutamic acid on the physical and chemical properties of gangue-based soil and soil invertase activity, and provide scientific basis and technical support for the ecological restoration and sustainable utilization of gangue-based soil.\u003c/p\u003e"},{"header":"1 Materials and methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e\u003ch2\u003e1.1 Soil culture experiment\u003c/h2\u003e\u003cp\u003eGangue soil samples were collected from the Wangjiazhai Coal Mine in Liupanshui, Guizhou in 1997, 2007, and 2021. The samples were collected by the five-point method, and 5 kg of samples were collected at each sampling point and mixed evenly. When sampling, plant litter and gravel on the soil surface were first cleaned, avoiding plant roots, and the soil profile was excavated to collect soil samples. The samples were packed in nylon bags and transported back to the laboratory, and then laid flat in the laboratory for natural air drying to remove debris from the gravel in the gangue. The coal gangue was ground and passed through a 2mm sieve, and 6.5kg of soil was accurately taken for potted culture experiments, and γ-polyglutamic acid was sprayed (CK-blank group, T-experimental group: T1-added 10.8g of polyglutamic acid, T2-added 21.6g of polyglutamic acid, T3-added 32.4g of polyglutamic acid, T4-added 43.2g of polyglutamic acid; dissolved in 300mL of distilled water respectively), and each concentration was repeated 3 times, and 100mL of deionized water was sprayed every 3 days. The plants were placed in a dry and ventilated place for culture, and the moisture content was maintained at 5%; at 3d, 18d, 33d, and 48d respectively, part of the soil was placed flat in a nylon bag in the laboratory, naturally air-dried, ground and sieved, and the physical and chemical properties and enzyme activity were determined.\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec4\" class=\"Section2\"\u003e\u003ch2\u003e1.2 Determination of soil chemical properties\u003c/h2\u003e\u003cp\u003eThe determination of soil pH value was carried out according to ISO (\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e1994\u003c/span\u003e) standards(ISO \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e1994\u003c/span\u003e). Weigh the air-dried soil sample that passed through a 2mm sieve, add deionized water without carbon dioxide according to the specified soil-water ratio to make a suspension, let it stand for 30 minutes, use the standard buffer solution to calibrate the Leiji SJ-4F pH meter, insert the electrode into the suspension, and record the soil pH value after the reading stabilizes; the determination of conductivity(Corwin and Lesch \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e2003\u003c/span\u003e) also uses air-dried soil sample that passed through a 2mm sieve, add deionized water according to the specified soil-water ratio, shake and stand, take the supernatant for determination, use the standard solution to calibrate the Leiji DDS-308F conductivity meter, insert the electrode, and record the value after the reading stabilizes to obtain soil conductivity data; Determination of soil organic matter (Bremner \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e1965\u003c/span\u003e),weigh the air-dried soil sample that passed through a 0.25 mm sieve and place it in a hard test tube, add a specific oxidant mixture, boil it in an oil bath, transfer it to a conical flask after cooling, and titrate it with a standard ferrous sulfate solution. The organic matter content is calculated based on the titration volume; the method for determining soil hydrolyzable nitrogen is the diffusion method (Mulvaney and Khan \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e2001\u003c/span\u003e).Weigh the fresh soil sample with the same treatment and place it in the outer chamber of the diffusion dish. Add boric acid and an indicator to the inner chamber. Add sodium hydroxide to the outer chamber, seal it, and place it in a constant temperature box to convert hydrolyzable nitrogen into ammonia and absorb it with boric acid. Then use a microburette to absorb the standard acid solution for titration, and calculate its content based on the titration volume; the determination of soil total nitrogen adopts the Kjeldahl method (Sparks et al. 2020), weigh the air-dried soil sample that passed the 0.25 mm sieve and place it in a Kjeldahl flask. After digestion with concentrated sulfuric acid and catalyst, transfer it to an automatic Kjeldahl nitrogen analyzer, add alkali for distillation, and the ammonia is absorbed by boric acid and titrated with standard acid. The instrument can automatically read and calculate the total nitrogen content of the soil; Determination of soil available phosphorus (Han et al.), weigh the air-dried soil sample that passed the 0.149 mm sieve and digest it with acid to dissolve phosphorus in the solution. The digested solution is determined by ICP-OES (inductively coupled plasma emission spectrometer), and a standard curve is drawn in combination with the standard solution to calculate the phosphorus content; The determination method of soil potassium content is similar to the determination steps of phosphorus. After digestion, the sample is determined by ICP-OES, and its potassium content is calculated based on the standard curve.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec5\" class=\"Section2\"\u003e\u003ch2\u003e1.3 Soil enzyme activity determination\u003c/h2\u003e\u003cp\u003eThe Solarbio kit was used to determine the activities of catalase, acid phosphatase, alkaline phosphatase, sucrase, and urease in soil (Bear and Salter \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e1916\u003c/span\u003e): First, a certain amount of soil sample was weighed, and the corresponding buffer (acid buffer for acid phosphatase, alkaline buffer for alkaline phosphatase, and ordinary buffer for other enzymes) was added, and the supernatant was obtained as the enzyme solution after oscillation, grinding, and other operations, and then centrifuged. Next, the enzyme solution was mixed with the respective substrate solutions (hydrogen peroxide solution for catalase, disodium phenyl phosphate solution for acid and alkaline phosphatases, sucrose solution for sucrase, and urease for urea solution) at an appropriate temperature and specific acid-base conditions. After the reaction was completed, the corresponding stop solution or reagent was added (some enzyme assays required the reaction to be stopped). Finally, a color developer was added, and the absorbance was measured at a specific wavelength. The corresponding products (remaining amount of hydrogen peroxide, amount of phenol generated, amount of glucose generated, and amount of ammonia generated) were calculated based on the standard curve to obtain the activity of each enzyme.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec6\" class=\"Section2\"\u003e\u003ch2\u003e1.4 Determination of soil enzyme aggregate ratio\u003c/h2\u003e\u003cp\u003eSoil microaggregates (Six et al. \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e2000\u003c/span\u003e) were determined by wet sieving, in which soil samples were placed on sieves of different apertures and oscillated or stirred in water to disperse the soil particles into aggregates of different sizes. The contents of microaggregates of different sizes were then determined by drying and weighing. The wet sieving method was also used for soil water-stable macroaggregates. The ability of soil macroaggregates to resist dispersion in water was utilized. Through a specific wet sieving operation, the macroaggregates in the soil samples were classified by particle size, and their composition and stability were then determined.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec7\" class=\"Section2\"\u003e\u003ch2\u003e1.5 Data analysis\u003c/h2\u003e\u003cp\u003eExcel was used to organize and preliminarily calculate the experimental data. The Duncan method was used for multiple comparisons to analyze the significance of the data of different treatments. The vegan package in R software was used to perform redundancy analysis on soil physicochemical properties and enzyme activities. All software versions used in this paper were Microsoft Office 2016 and R 4.4.2.\u003c/p\u003e\u003c/div\u003e"},{"header":"2 Results and analysis","content":"\u003ch2\u003e2.1 Effects on soil physicochemical properties\u003c/h2\u003e\n\u003ch3\u003e2.1.1 Effects of \u0026gamma;-glutamic acid on pH of gangue-based soil\u003c/h3\u003e\n\u003cp\u003eThe results of pH analysis showed that the pH values of the blank group (CK) and the treatment groups (T1-T4) added with polyglutamic acid in each year were different at 3d, indicating that the soil pH increased rapidly after the addition of polyglutamic acid; the pH values of most treatment groups showed an overall upward trend from 3d to 48d, but some treatment groups changed more slowly, such as the T3 and T4 groups in some years, while the blank group had the opposite trend, with the pH value decreasing with time; in the soil of the same year, the different amounts of polyglutamic acid added had different effects on the pH value. Generally, the higher the amount added, the higher the pH value at certain time points. For example, in the soil of 1997, from T1-97 to T4-97, as the amount added increased from 10.8g to 43.2g, at 18d, 33d, and 48d At the same time point, the pH value tended to increase gradually; the initial pH values of the blank groups in different years were different. The results showed a pattern, that is, the pH of the blank group was the lowest in 2007, but after adding polyglutamic acid, the soil pH growth rate in 2007 was the largest under each treatment group.\u003c/p\u003e\n\u003ch3\u003e2.1.2 Effect of \u0026gamma;-glutamic acid on the conductivity of gangue-based soil\u003c/h3\u003e\n\u003cp\u003eThe conductivity of the blank groups in the three years increased with time, indicating that when no polyglutamic acid was added, the conductivity of the gangue-based soil increased over time. After the addition of polyglutamic acid to each experimental group, the initial conductivity was much lower than that of the blank group in the corresponding year, indicating that the addition of polyglutamic acid significantly reduced the initial conductivity of the soil. Overall, the conductivity of each experimental group showed a downward trend or an upward trend followed by a downward trend with time, and was significantly lower than the blank group during the same period, indicating that the addition of polyglutamic acid changed the change pattern of the conductivity of the gangue-based soil over time and inhibited its upward trend.\u003c/p\u003e\n\u003cp\u003eTable 1 Changes in soil conductivity\u003c/p\u003e\n\u003ctable border=\"0\" cellspacing=\"0\" cellpadding=\"0\" width=\"440\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 64px;\"\u003e\n \u003cp\u003eTreatment\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 94px;\"\u003e\n \u003cp\u003e3d\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 94px;\"\u003e\n \u003cp\u003e18d\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 94px;\"\u003e\n \u003cp\u003e33d\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 94px;\"\u003e\n \u003cp\u003e48d\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eCK-97\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e1023.33\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e1722.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e1844.67\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e1905.00\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eT1-97\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e3.82\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.86\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.73\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.74\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eT2-97\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e3.55\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.92\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.84\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.77\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eT3-97\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e3.52\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.94\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.83\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.79\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eT4-97\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e4.53\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.90\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.42\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.35\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eCK-07\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e432.33\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e1364.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e1846.67\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e1915.00\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eT1-07\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e7.77\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e6.55\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e5.35\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.88\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eT2-07\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e5.84\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e4.66\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e3.76\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.43\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eT3-07\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e4.36\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e3.89\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.87\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.63\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eT4-07\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.91\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.75\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.24\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e1.96\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eCK-21\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e1045.33\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e1790.33\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e1901.33\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e1958.67\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eT1-21\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.95\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.86\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.56\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.37\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eT2-21\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.84\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.66\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.34\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.15\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eT3-21\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e4.13\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e3.29\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.83\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.73\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eT4-21\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e5.18\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e3.86\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.90\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.86\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003ch3\u003e2.1.3 Effect of \u0026gamma;-glutamic acid on the content of organic matter, nitrogen, phosphorus and potassium in gangue-based soil\u003c/h3\u003e\n\u003cp\u003e(1) Soil organic matter content\u003c/p\u003e\n\u003cp\u003eThe results of the analysis of organic matter content showed that in different treatment groups in the same year, the addition of polyglutamic acid would cause the soil organic matter content to differ from that of the blank group. For example, in the soil of 1997, the organic matter content of the T1-97 to T4-97 groups at each time point was different from that of the CK-97 group, and the content of the treatment groups with different addition amounts also varied. In the soil of the same year, the addition of different amounts of polyglutamic acid had different effects on the soil organic matter content. The organic matter content of most treatment groups showed a trend of first increasing and then decreasing with the increase of the addition of polyglutamic acid, indicating that the moderate addition of polyglutamic acid was beneficial to increase the organic matter content. From the perspective of overall time changes, the organic matter content of some treatment groups gradually increased over time; while the content of some treatment groups increased and decreased, or the change range was small, such as the CK-97 content of the soil in 2007. In group 07, the organic matter content decreased from the 3rd day to the 18th day, and then increased from the 18th day to the 33rd day. The initial soil organic matter content of the blank groups in different years was different, and the changes over time were also different. For example, the initial content of CK-97, CK-07 and CK-21 on the 3rd day was high and low, and the change range and trend in the subsequent time were also different. This may be related to the composition, properties and environmental factors of the gangue-based soil itself in different years.\u003c/p\u003e\n\u003cp\u003e(2）Soil nitrogen content\u003c/p\u003e\n\u003cp\u003eThe analysis results of total nitrogen show that after adding polyglutamic acid, the total nitrogen content of the soil is different from that of the blank group. There is an obvious trend that the total nitrogen content increases with the increase of polyglutamic acid application. From the perspective of time change trend, the total nitrogen content of some treatment groups gradually increases over time, while some treatment groups first decrease and then increase, or first increase and then decrease, or the change range is small. The initial total nitrogen content of the blank group in different years is different, and the change over time is also different. For example, the initial total nitrogen content of CK-97, CK-07 and CK-21 at 48 days is high and low, and the change range and trend in the subsequent time are also different. This may be related to the nitrogen storage state, microbial activity and environmental factors of the gangue-based soil itself in different years.\u003c/p\u003e\n\u003cp\u003eThe analysis results of hydrolyzable nitrogen show that in the same year, the different amounts of polyglutamic acid added to the treatment group had a significant effect on the soil hydrolyzable nitrogen content compared with the blank group. In most cases, with the increase in the amount of addition, the soil hydrolyzable nitrogen content was relatively higher. From the time trend, there was no uniform pattern in the changes of soil hydrolyzable nitrogen content in each treatment group over time. Some decreased first and then increased, and some fluctuated. However, the hydrolyzable nitrogen content in most treatment groups was higher on 18 days and 33 days, indicating that the microbial conversion of nitrogen was strongest at these times.\u003c/p\u003e\n\u003cp\u003e(3）Soil potassium content\u003c/p\u003e\n\u003cp\u003eThe results of soil total potassium analysis show that in the same year, the effect of different amounts of polyglutamic acid addition on soil total potassium content is not obvious. In most cases, the total potassium content between treatment groups with different addition amounts is not much different, and there is no trend of regular change with the increase or decrease of addition amount; from the perspective of time change trend, the soil total potassium content of each treatment group changes relatively steadily over time, without a particularly significant upward or downward trend. However, the total potassium content of treatment groups in different years at different time points still fluctuates. For example, in some treatment groups in 2007, the total potassium content has a small increase and decrease from 3 days to 48 days.\u003c/p\u003e\n\u003cp\u003eThe analysis of soil available potassium content showed that in the soil of the same year, the effects of different amounts of polyglutamic acid added on soil available potassium content were more complicated. Except for the 3-day time point, the available potassium at other time points showed obvious regularity, that is, the soil available potassium content increased with the increase of polyglutamic acid application rate. From the time change trend, there was no uniform pattern in the changes of soil available potassium content in each treatment group over time. The content of some treatment groups increased over time, while some first increased and then decreased, or fluctuated up and down.\u003c/p\u003e\n\u003cp\u003e(4）Soil phosphorus content\u003c/p\u003e\n\u003cp\u003eThe results of soil phosphorus content analysis show that in the same year, the effect of different amounts of polyglutamic acid addition on soil total phosphorus content is relatively complex. At the beginning of application, soil phosphorus content increased with the increase of polyglutamic acid application, and this trend weakened over time. By 48 days, phosphorus content showed only a slight increase with the increase of polyglutamic acid application. Overall, in the same year, soil phosphorus content tended to decrease over time; from the perspective of time change trend, soil total phosphorus content of each treatment group did not show a uniform increase or decrease pattern over time. Some treatment groups increased in certain time periods, while others decreased or remained relatively stable.\u003c/p\u003e\n\u003cp\u003eThe results of soil available phosphorus content analysis showed that in the same year, there were differences in soil available phosphorus content between the treatment group with polyglutamic acid addition and the blank group. In the soil of the same year, there was no obvious pattern in the effect of different amounts of polyglutamic acid added on soil available phosphorus content. The content of soil available phosphorus increased significantly when polyglutamic acid was just applied, but this pattern did not appear in the subsequent time. The order of available phosphorus content in the treatment groups with different addition amounts at different time points was unstable. From the perspective of time change trend, there was no obvious pattern in the change trend of soil available phosphorus content in each treatment group over time. The content of some treatment groups increased over time, while that of others decreased or fluctuated. However, there was an obvious change that the content of soil available phosphorus was significantly higher when polyglutamic acid was just applied than that of soil available phosphorus at 18, 33 and 48 days after application.\u003c/p\u003e\n\u003ch3\u003e2.1.4 Effect of \u0026gamma;-glutamic acid on aggregates in coal gangue-based soil\u003c/h3\u003e\n\u003cp\u003eFor soil water-stable macroaggregates, in the soil of the same year, after adding polyglutamic acid, the proportion of water-stable macroaggregates in different particle size ranges changed. Some treatment groups with added amounts increased the proportion of aggregates, while others changed the proportion of aggregates, and there was no uniform change with the increase or decrease of the amount of addition; the blank groups of different years had different initial proportions of soil water-stable macroaggregates in each particle size range, and the proportions of aggregates in different particle size ranges were different among the treatment groups, but the particle size proportions did not show regularity; Overall, the aggregates with the smallest particle size (\u0026lt;0.25mm) and the largest particle size (\u0026gt;5mm) accounted for the largest proportion.\u003c/p\u003e\n\u003cp\u003eFor soil water-stable microaggregates, compared with different treatment groups in the same year, after adding polyglutamic acid, the proportion of soil water-stable microaggregates in each particle size range was different from that in the blank group. The treatment groups with some addition amounts would increase or decrease the proportion of microaggregates in certain particle size ranges, but did not show a uniform pattern of change with the addition amount. The blank groups in different years had different initial proportions of soil water-stable microaggregates in each particle size range, but did not show obvious differences between years. Overall, aggregates with a particle size range of 0.01 - 0.05 mm and \u0026gt; 0.25 mm accounted for the largest proportion.\u003c/p\u003e\n\u003ch2\u003e2.2 Effect of \u0026gamma;-glutamic acid on the activity of invertase in coal gangue-based soil\u003c/h2\u003e\n\u003cp\u003eIn different years, the urease activity of the CK group mostly showed a downward trend over time, but the degree of decline was different in each year; compared with the CK group, the changes in urease activity were different after adding polyglutamic acid, but the main manifestation was that except for the T1 treatment group, the soil urease activity of other treatment groups decreased over time, but the speed and magnitude of the decline were different in different treatment groups, indicating that the addition of polyglutamic acid would affect the soil urease activity; between different years, the initial value and change trend of soil urease activity in different treatment groups were different, but there was no regularity.\u003c/p\u003e\n\u003cp\u003eIn different years, the sucrase activity of the CK group generally showed an upward trend over time, but the increase and speed were different in each year. Compared with the CK group, after adding polyglutamic acid, the activity growth of each treatment group was different, and the activity differences between treatment groups with different addition amounts were also inconsistent at different time points, but overall, the soil sucrase activity increased over time, and the application of polyglutamic acid increased the activity of soil sucrase, indicating that the addition amount and time factors of polyglutamic acid will affect the soil sucrase activity; in different years, the initial activity of each treatment group was relatively low in 1997, and then the increase was more obvious. In 2007, the initial activity was unevenly distributed, and the subsequent changes were also different. The initial activity in 2021 was relatively medium, and the change trend was also different from the previous two years.\u003c/p\u003e\n\u003cp\u003eIn different years, there was no uniform pattern in the changes of catalase activity in the CK group. Compared with the CK group, after adding polyglutamic acid, the catalase activity changed in various ways. The trends and amplitudes of activity changes in each treatment group were different, and the differences between treatment groups with different addition amounts were different at different time points. The soil enzyme activity was the largest at 18 or 33 days, and except for 2021, the application of polyglutamic acid increased soil catalase activity in other years, indicating that the amount and time of polyglutamic acid addition would significantly affect the activity of soil catalase; the initial values and change trends of soil catalase activity were significantly different between different years. In 1997, the initial activity distribution of each treatment group was relatively scattered, and the amplitude of change was large; in 2007, the initial activity was relatively concentrated, and the changes were more complex; in 2021, the initial activity was different, and the activity of some treatment groups increased significantly in the later period, indicating that the intrinsic properties of different soil years had different effects on catalase activity.\u003c/p\u003e\n\u003cp\u003eThe initial values and changing trends of soil acid phosphatase activity in different years were quite different. The comparison between treatment groups showed that the changes in acid phosphatase activity in the CK group were different in different years. It showed a downward trend in 1997, first stabilized and then increased in 2007, and fluctuated in 2021, reflecting that the soil\u0026apos;s own factors had different effects on the enzyme activity in different years. Compared with the CK group, the changing trends and amplitudes of the activities of each treatment group after adding polyglutamic acid were different, and the activity differences between treatment groups with different addition amounts were also inconsistent at different time points, but most treatment groups showed a trend of first increasing and then decreasing over time, and the enzyme activity of the T2 treatment group was relatively high, indicating that the addition of polyglutamic acid under the T2 condition of polyglutamic acid would significantly increase the enzyme activity. The results between different years showed that the initial activity of each treatment group was unevenly distributed in 1997, and the subsequent changes fluctuated significantly; the initial activity was low in 2007, and some treatment groups showed an upward trend afterwards; the initial activity was different in 2021, and the change trends of each treatment group in the later period were diverse, indicating that there were significant differences in soil phosphatase activity in different years.\u003c/p\u003e\n\u003cp\u003eIn different years, the alkaline phosphatase activity of the CK group changed differently. In 1997, it first increased and then decreased, in 2007 it was high at first and then low with large fluctuations, and in 2021 it first increased and then decreased, indicating that the soil\u0026apos;s own characteristics in different years have different effects on the enzyme activity; compared with the CK group, after the addition of polyglutamic acid, the change trends and amplitudes of the activity of each treatment group were different, and the activity differences between treatment groups with different addition amounts were also different at each time point, but the alkaline phosphatase activity of the treatment group reached a peak at 18 or 33 days, but regardless of the application amount, the alkaline phosphatase activity of most treatment groups was lower than that of the blank group, indicating that the addition of polyglutamic acid will reduce the activity of alkaline phosphatase in the soil, and the effect of the addition amount of polyglutamic acid on enzyme activity will change over time.\u003c/p\u003e\n\u003ch2\u003e2.3 Redundancy analysis of soil physicochemical properties and invertase activity\u003c/h2\u003e\n\u003cp\u003eRDA analysis was performed with soil physicochemical properties after adding different concentrations of \u0026gamma;-polyglutamic acid as the response variable and soil carbon, nitrogen and phosphorus cycle-related enzyme activities as the explanatory variables.\u003c/p\u003e\n\u003cp\u003eThe points of each treatment group are scattered in the figure, indicating that there are differences in the comprehensive properties of the soil under different treatments. The 3d RDA analysis results showed that there was a significant correlation between the enzymes urease, sucrase, catalase, and alkaline phosphatase; catalase and acid phosphatase had a greater impact on soil physicochemical properties, followed by alkaline phosphatase, and the other two enzymes had a lower explanatory rate; at the same time, the enzyme activity of the soil in 1997 and 2007 with polyglutamic acid added was higher, and the enzyme activity of the blank group and the treatment group in 2021 was lower. The results of the 18-day RDA analysis showed that, except for urease and alkaline phosphatase, there was a certain degree of correlation between other enzymes; acid phosphatase and alkaline phosphatase had a greater impact on soil physical and chemical properties, urease and sucrase had a certain impact, and catalase had the least impact; the enzyme activity of the 2007 treatment group and the 1997 T1 and T2 treatment groups was higher, and the enzyme activity of other treatment groups was lower. The results of the 33-day RDA analysis showed that there was a significant correlation between the five enzymes, especially sucrase and acid phosphatase had a very strong correlation; urease, catalase and acid phosphatase had similar effects on soil physical and chemical properties, and alkaline phosphatase and sucrase had less impact; the enzyme activity of the 2007 treatment group and the 1997 T1 and T2 treatment groups was higher, and the enzyme activity of other treatment groups was lower. The RDA analysis results of 48 days showed that there was a certain degree of correlation between the five enzymes; urease, sucrase and acid phosphatase had similar effects on the physical and chemical properties of the soil, while alkaline phosphatase and catalase had smaller effects; the enzyme activity was higher in the T1 and T2 treatment groups in 1997 and 2021.\u003c/p\u003e"},{"header":"3 Discussion","content":"\u003cdiv id=\"Sec16\" class=\"Section2\"\u003e\u003ch2\u003e3.1 Effect of polyglutamic acid on soil physicochemical properties\u003c/h2\u003e\u003cp\u003ePolyglutamic acid can quickly increase soil pH after addition. The pH of most treatment groups showed an upward trend over time, while the blank group showed a downward trend. This may be due to the chemical properties of polyglutamic acid itself or its reaction with acid-base substances in the soil. At the same time, the addition of polyglutamic acid significantly reduced the initial conductivity of the soil, changed its change pattern over time, and inhibited the upward trend. This shows that polyglutamic acid can effectively regulate the acid-base balance and ion concentration of the soil (Skalski et al. \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e2024\u003c/span\u003e), which has positive significance for improving the chemical environment of coal gangue-based soil.\u003c/p\u003e\u003cp\u003ePolyglutamic acid has a complex effect on the content of nutrients such as soil organic matter, nitrogen, phosphorus, and potassium. The addition of polyglutamic acid caused a difference in soil organic matter content compared with the blank group. The organic matter content of most treatment groups increased first and then decreased with the increase in the amount of addition, indicating that the appropriate amount of polyglutamic acid is beneficial to increase organic matter (Guo et al. \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e2024\u003c/span\u003e). It may be that polyglutamic acid promotes soil microbial activity and accelerates the decomposition and synthesis of organic matter(Guo et al. \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e2024\u003c/span\u003e). In terms of nitrogen, the total nitrogen and hydrolyzable nitrogen contents changed significantly after the addition of polyglutamic acid, and increased with the increase in the amount of addition, indicating that polyglutamic acid can improve the storage and conversion efficiency of soil nitrogen (Zhang et al. \u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e2017a\u003c/span\u003e). For potassium, the total potassium content was not significantly affected by the amount of polyglutamic acid added, but the available potassium content increased with the increase in the amount of addition except for 3 days, indicating that polyglutamic acid can promote the effectiveness of potassium(Guo et al. \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e2017\u003c/span\u003e). In terms of phosphorus, the total phosphorus and available phosphorus content are complexly affected by the amount and time of polyglutamic acid addition. Polyglutamic acid can increase the phosphorus content in the initial stage, but this promotion effect weakens over time, which may be related to the fixation and release mechanism of phosphorus in the soil (Zhang et al. \u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e2017b\u003c/span\u003e). The effect of γ-PGA on soil organic matter and nutrients such as nitrogen, phosphorus, and potassium is affected by many factors. It needs to be reasonably applied according to specific soil conditions to optimize nutrient utilization efficiency. The effect of γ-PGA on soil nutrients is complex due to different soil types, environmental conditions, and application methods. In some cases, γ-PGA can improve the effectiveness of soil nitrogen and plant absorption of nutrients, but its effects on soil organic matter, phosphorus, potassium and other nutrients may vary depending on specific conditions (Zhang et al. \u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e2017a\u003c/span\u003e)。\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec17\" class=\"Section2\"\u003e\u003ch2\u003e3.2 Effect of polyglutamic acid on soil invertase activity\u003c/h2\u003e\u003cp\u003ePolyglutamic acid has a significant effect on the activities of soil urease, sucrase, catalase, acid phosphatase and alkaline phosphatase. Urease activity decreased over time in most treatment groups except the T1 treatment group, indicating that the amount of polyglutamic acid added would affect urease activity. It may be that polyglutamic acid changed the structure of soil microbial community and affected the growth and metabolism of urease-producing bacteria (Qiong et al. 2018). Sucrase activity increased over time, and polyglutamic acid could increase its activity, indicating that polyglutamic acid could promote the circulation and transformation of carbon in soil (O'Dowd and Hopkins \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e1998\u003c/span\u003e). Catalase activity varied. Except for 2021, polyglutamic acid could increase its activity, and the activity was the highest at 18 or 33 days, indicating that polyglutamic acid and time factors jointly affect the redox process of soil. The activity of acid phosphatase in most treatment groups increased first and then decreased, and the enzyme activity in the T2 treatment group was higher, indicating that a specific amount of polyglutamic acid added can significantly increase the activity of the enzyme, which is beneficial to the transformation and utilization of phosphorus in the soil (Zhang et al. \u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e2024\u003c/span\u003e). After adding polyglutamic acid, the activity of alkaline phosphatase in most treatment groups was lower than that in the blank group, and reached a peak at 18 or 33 days, indicating that polyglutamic acid inhibits the activity of the enzyme, and its effect changes over time. The initial values ​​and change trends of soil enzyme activity in different years are significantly different, reflecting the important influence of soil properties on enzyme activity(Hou et al. \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2018\u003c/span\u003e)。 There are differences in the chemical composition, microbial community composition and structure of coal gangue-based soil in different years, which can affect the synthesis, stability and activity of enzymes (Sun et al. \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e2020\u003c/span\u003e). For example, differences in soil pH, organic matter content, etc. can affect the growth environment of microorganisms, and thus affect the enzyme activity produced by microorganisms.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec18\" class=\"Section2\"\u003e\u003ch2\u003e3.3 Relationship between soil physicochemical properties and invertase activity\u003c/h2\u003e\u003cp\u003eRDA analysis showed that there were differences in the comprehensive properties of soil under different treatments, and the relationship between enzyme activities at different time points and between enzyme activities and soil physicochemical properties was complex (Deng et al. \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2019\u003c/span\u003e)。The degree of influence of different enzyme activities on soil physicochemical properties varies with time (Lan Yu et al. \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e2011\u003c/span\u003e). For example, catalase and acid phosphatase had a greater effect at 3d, while acid phosphatase and alkaline phosphatase had a prominent effect at 18d. There were also differences in enzyme activity between different years and treatment groups, indicating that soil physicochemical properties and invertase activity are interrelated and affect each other. Soil physicochemical properties such as nutrient content and pH will affect enzyme activity and stability, and changes in enzyme activity will react to soil material circulation and transformation processes, thereby affecting soil physicochemical properties. In summary, polyglutamic acid has a significant effect on the physicochemical properties and invertase activity of coal gangue-based soil, and is restricted by multiple factors such as addition amount, time and soil year. This study provides a theoretical basis and technical reference for the improvement and ecological restoration of coal gangue-based soils, but further in-depth research on the mechanism of interaction between polyglutamic acid and soil is still needed to optimize soil improvement measures.\u003c/p\u003e\u003c/div\u003e"},{"header":"Conclusion","content":"\u003cp\u003eThis study explored the effects of polyglutamic acid on gangue-based soil from many aspects, comprehensively analyzed soil physical and chemical properties, invertase activity and the relationship between the two, and drew the following conclusions: polyglutamic acid rapidly increased soil pH, significantly reduced initial conductivity, and effectively improved soil acid-base and salinity; the appropriate addition of polyglutamic acid was conducive to the accumulation of organic matter, and the addition of polyglutamic acid could improve the storage and conversion efficiency of soil nitrogen and enhance the effectiveness of potassium. The fixation and release mechanism of soil phosphorus was jointly affected by polyglutamic acid and time; the addition of polyglutamic acid changed the structure of soil microbial community and increased soil enzyme activity; the properties of the soil itself also played a decisive role in the formation and change of enzyme activity. Polyglutamic acid has a significant improvement effect on gangue-based soil, but the improvement effect is synergistically restricted by multiple factors such as the amount of polyglutamic acid added, time and soil year. This study provides a solid theoretical basis for the ecological restoration and sustainable utilization of gangue-based soil. In the future, it is necessary to further explore the microscopic mechanism of the interaction between polyglutamic acid and soil, further optimize soil improvement strategies, and promote the effective restoration and sustainable development of the ecological environment in mining areas.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis study was funded by the Mountain Plateau Specialty Agricultural Products Technology Innovation Center (520202023018).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting Interests \u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eOn behalf of all authors, the corresponding author states that there is no conflict of interest.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthor contribution \u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eJing Shi, Houhuan Liu, Guangli Chen, and Xiaoxia Zhao were responsible for sample collection. Jing Shi also conducted the physical and chemical index analysis. Jianhong Li was in charge of the analysis of soil enzyme activities. Shuo Jiang Song performed the data analysis, with Lili Feng providing guidance throughout the data analysis process. Each author\u0026apos;s efforts were integral to the completion of this study, ensuring the reliability and validity of the research findings.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eAlkorta, I., A. Aizpurua, P. Riga, I. Albizu, I. Amezaga, and C. Garbisu. 2003. Soil Enzyme Activities as Biological Indicators of Soil Health. Reviews on environmental health \u003cstrong\u003e18\u003c/strong\u003e:65-73.\u003c/li\u003e\n\u003cli\u003eBear, F. E., and R. M. Salter. 1916. Methods in soil analysis. W. Va. Agr. Exp. Sta., Tech. Bul \u003cstrong\u003e159\u003c/strong\u003e:1-24.\u003c/li\u003e\n\u003cli\u003eBremner, J. M. 1965. Total Nitrogen. 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Yuan. 2024. Glutamic Acid-Enhanced Phosphate Fertilizer Increases Phosphorus Availability in Fluvo-Aquic Soil via Phosphamide (O\u0026thinsp;=\u0026thinsp;P-N) Formation, Decreasing Phosphate Fixation and Increasing Soil Microbial Diversity. Journal of soil science and plant nutrition \u003cstrong\u003e24\u003c/strong\u003e:2748-2760.\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":"γ-polyglutamic acid, coal gangue-based soil, soil physical and chemical properties, enzyme activity","lastPublishedDoi":"10.21203/rs.3.rs-7097342/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-7097342/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003e\u003cstrong\u003ePurpose\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003ehis study aimed to explore the effects of polyglutamic acid on the physical-chemical properties and invertase activity of gangue-based soil, clarifying how different addition amounts impact soil indicators and enzyme activities for ecological restoration.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMethods\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e​\u003c/strong\u003eSoil samples with varying weathering years were collected from a gangue mountain. A potted experiment involved five polyglutamic acid treatments (0–43.2 g dissolved in 300 mL water). Soil pH, conductivity, nutrient contents, aggregate composition, and enzyme activities (urease, sucrase) were measured, followed by redundancy analysis (RDA).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eResults\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e​\u003c/strong\u003ePolyglutamic acid rapidly increased soil pH and modified conductivity trends. Its influence on organic matter, nitrogen, phosphorus, and potassium was complex, depending on dosage and time; optimal addition boosted organic matter and nitrogen conversion, promoted potassium availability, and altered water-stable aggregate ratios. Enzyme activities were significantly affected by dosage, with varying initial levels and trends across weathering years; overall, certain enzymes showed increased activity. RDA indicated distinct soil properties under different treatments, with enzyme impacts evolving over time.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConclusions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003ePolyglutamic acid significantly improves gangue-based soil, offering a theoretical basis and technical reference for ecological restoration and soil quality optimization.\u003c/p\u003e","manuscriptTitle":"Effects of γ-polyglutamic acid on available nutrients and enzyme activities in gangue-based soil","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-09-10 17:01:04","doi":"10.21203/rs.3.rs-7097342/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":"ebe1e2b2-7313-4376-a4b2-2b13d405ba14","owner":[],"postedDate":"September 10th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2025-10-08T09:09:07+00:00","versionOfRecord":[],"versionCreatedAt":"2025-09-10 17:01:04","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-7097342","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-7097342","identity":"rs-7097342","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}