اثر كاربرد EDTA و اسيدسيتريك بر تغييرات فعاليت آنزيمي خاك، تنفس ميكروبي برانگيخته و فراهمي سرب در خاك

The Effect of EDTA and Citric acid on Soil Enzymes Activity, Substrate Induced Respiration and Pb Availability in a Contaminated Soil

افزودن EDTA به عنوان یك عامل كلات كننده به خاك‌های آلوده به فلزات سنگین ممكن است فراهمی و استخراج گیاهی فلزات را از خاك افزایش دهد اما می‌تواند اثرات منفی بر ویژگی‌های بیولوژیكی خاك داشته باشد. به منظور بررسی اثر كاربرد EDTA و اسید سیتریك بر فراهمی سرب، فعالیت دهیدروژناز، اوره‌آز و فسفومونواستراز قلیایی و همچنین تنفس میكروبی برانگیخته (SIR)، آزمایشی در قالب طرح كاملاً تصادفی با آرایش فاكتوریل و سه تكرار در شرایط گلخانه اجرا گردید. تیمارهای آزمایش شامل EDTAو اسید سیتریك در دو سطح 3 و 5 میلی¬مول بر كیلوگرم به همراه شاهد و زمان (7، 14، 21 و 28 روز) بودند. نتایج نشان داد كه به طور میانگین تیمارهای EDTA3 و EDTA5 به ترتیب سبب افزایش 17/2 % و 10 % غلظت فراهم سرب نسبت به شاهد شدند. این در حالی است كه بررسی اثر زمان نشان داد كه فراهمی سرب برای تیمار EDTA3 در زمان 28 در مقایسه با اولین زمان نمونه¬برداری (روز هفتم) 3/12 % كاهش یافت در حالی كه برای تیمار EDTA5 تغییر معنی‌داری در طی زمان مشاهده نشد. همچنین به طور میانگین تیمارهای CA3 و CA5 به ترتیب سبب كاهش 3/8 % و 7/15 % غلظت فراهم سرب نسبت به شاهد شدند به طوری¬كه در آخرین زمان نمونه¬برداری فراهمی سرب به ترتیب 9/15 % و 8/12 % نسبت به اولین زمان كاهش پیدا كرد. نتایج نشان دهنده اثر منفی EDTA بر فعالیت آنزیم‌های دهیدروژناز و اوره‌آز خاك بود. از طرف دیگر افزودن اسیدسیتریك اثر مثبتی بر فعالیت آنزیم‌های دهیدروژناز، اوره‌آز و فسفومونواستراز قلیایی خاك داشت. در تمام تیمارها به جز تیمار شاهد فعالیت آنزیم‌های مورد مطالعه با گذشت زمان افزایش پیدا كرد كه این افزایش در تیمارهای اسید سیتریك بیشتر بود. افزودن EDTA و اسیدسیتریك مقدار SIR خاك را در همه زمان‌های مورد مطالعه افزایش دادند كه این افزایش برای تیمارهای اسیدسیتریك بیشتر از EDTA بود.

Introduction: Application of EDTA may increase the heavy metal availability and phytoextraction efficiency in contaminated soils. In spite of that, it might also have some adverse effects on soil biological properties. Metals as freeions are considered to be severely toxic, whereas the complexed form of these metalswith organic compounds or Fe/Mn oxides may be less available to soil microbes. However, apart from this fact, some of these compounds like EDTA and EDTA-metal complexes have low bio- chemo- and photo-degradablity and high solubility in their own characteristics andable to cause toxicity in soil environment. So more attentions have been paid to use of low molecular weight organic acids (LMWOAs) such as Citric acid because of having less unfavorable effects to the environment. Citric acid increases heavy metals solubility in soils and it also improves soil microbial activity indirectly. Soil enzymes activity is a good indicator of soil quality, and it is more suitable for monitoring the soil quality compared to physical or chemical indicators. The aims of this research were to evaluate the changes of dehydrogenase, urease and alkaline phosphomonoesterase activities, substrate-induced respiration (SIR) and Pb availability after EDTA and citric acid addition into a contaminated soil with PbCl2. Materials and Methods: An experiment was conducted in a completely randomized design with factorial arrangement and three replications in greenhouse condition. The soil samples collected from surface horizon (0-20 cm) of the Typic haplocalsids, located in Mashhad, Iran. Soil samples were artificially contaminated with PbCl2 (500 mg Pb per kg of soil) and incubated for one months in 70 % of water holding capacity at room temperature. The experimental treatments included control, 3 and 5 mmol EDTA (EDTA3 and EDTA5) and Citric acid (CA3 and CA5) per kg of soil. Soil enzymes activity, substrate-induced respiration and Pb availability of soil samples were determined by standard methods after 7, 14, 21 and 28 days of chelates addition. Results and Discussion: The soil texture was loam and the indigenous Pb content was 25.55 mg kg-1. The soil pH was 7.4 and electrical conductivity of saturated extraction measured 2.5 dS m-1. The soil carbonate calcium was 14% and the content of organic carbon and essential nutrients were low. The results showed that EDTA3 and EDTA5 treatments increased Pb availability by 2.17% and 10% compared to control treatment but CA3 and CA5 treatments decreased it by 3.8% and 15.7% respectively. The Pb availability in control and EDTA5 treatments did not change during the incubation time. The available Pb concentration dropped sharply during the incubation time in EDTA3, CA3 and CA5 treatments. The reduction rates in CA3 and CA5 treatments were more than EDTA3 treatment. This may be due to the high stability and low biodegradability of EDTA than biodegradable chelators and low molecular weight organic acids. The results showed that urease and dehydrogenase activities were significantly reduced in EDTA3 and EDTA5 treatments compared to control treatment. Urease and dehydrogenase activities were decreased with the increase of EDTA concentration. Alkaline phosphomonoesterase activity was not affected by the EDTA3 and EDTA5 treatments. In CA3 and CA5 treatments, dehydrogenase and alkaline phosphomonoesterase activities significantly increased with increasing the concentration of citric acid. CA5 treatment showed a prominent effect on urease activity compare to CA3 treatment. The soil enzyme activities increased with incubation time. It seems that reduction in Pb availability causes an increase of soil enzymes activities. Significant negative relationships were found between soil enzymes activities and available Pb concentration (dehydrogenase activity (r=-0.906, P <0.05), alkalinephosphomonoesterase activity (r=-0.747, P <0.05) and urease activity (r=-0.539, P <0.05)). The results showed that soil enzyme activities were affected by Pb availability. The results also revealed that substrate induced respiration enhanced by adding EDTA and Citric acid but additive effect of Citric acid was more than EDTA. The CA3 and CA5 treatments increased substrate induced respiration by 60.14% and 60.5% compared to control treatment, respectively. The effects of EDTA on substrate induced respiration depended on the EDTA concentration. The application of 5 mmol EDTA per kg of soil did not significantly effect substrate induced respiration but the 3 mmol EDTA per kg of soil incresead it by 22.11% compared to control treatment. Conclusion: Based on our finding, EDTA application increased Pb availability as compared to control. In comparison with EDTA, Citric acid did not have ability to increase Pb availability. Activity of soil enzymes significantly were reduced by EDTA application. On the contrary, soil enzymes and microbial activity improved significantly by Citric acid applications. It is suggested that lower EDTA application and higher Citric acid concentrations can be considered for futures studies.