Determination of ammonium and potassium extracted from soil samples by capillary electrophoresis with capacitively coupled contactless conductivity detection (ce-C4d) - Van Anh Nguyen

135 Tạp chí phân tích Hóa, Lý và Sinh học - Tập 21, Số 2/2016 DETERMINATION OF AMMONIUM AND POTASSIUM EXTRACTED FROM SOIL SAMPLES BY CAPILLARY ELECTROPHORESIS WITH CAPACITIVELY COUPLED CONTACTLESS CONDUCTIVITY DETECTION (CE-C4D) Đến tòa soạn 23 - 3 - 2016 Van Anh Nguyen Department of Environmental Sciences and Technologies, Hanoi Metropolitan University Thi Trang Do, Van Lau Ha, Hung Viet Duong, Tien Duc Pham, Thi Anh Huong Nguyen* Faculty of Chemistry - Hanoi University of Science - Vietnam National University, Hanoi, Thu Hien Le Faculty of Transport Economics, University of Transport Technology TÓM TẮT XÁC ĐỊNH AMONI VÀ KALI TRONG DỊCH CHIẾT CỦA MẪU ĐẤT BẰNG PHƯƠNG PHÁP ĐIỆN DI MAO QUẢN SỬ DỤNG DETECTOR ĐỘ DẪN KHÔNG TIẾP XÚC (CE-C4D) Bài báo nghiên cứu quy trình phân tích hai ion NH4+ và K+ trong dịch chiết mẫu đất nông nghiệp bằng kĩ thuật điện di mao quản sử dụng detector độ dẫn không tiếp xúc. Các điều kiện phân tích đã được tối ưu bao gồm: Dung dịch đệm điện di sử dụng hỗn hợp đệm histidine/axit acetic (His/Ace) (pH = 4), 2mM 18-crown-6; mao quản Silica: đường kính ngoài (O.D.) 365 μm, đường kính trong (I.D.) 50 μm, chiều dài tổng Lt= 50cm và chiều dài hiệu dụng Leff = 40cm; điện thế tách: 20kV; bơm mẫu theo phương pháp thủy động lực theo kiểu xiphong ở độ cao 15cm, trong thời gian 20s. Quy trình được áp dụng để phân tích dịch chiết 01 mẫu đất và 01 mẫu nước lấy tại cùng một khu canh tác nông nghiệp. Các kết quả phân tích được kiểm chứng bằng các phương 136 pháp quang phổ hấp thụ nguyên tử (kiểm chứng kết quả phân tích K+) và phương pháp quang phổ hấp thu phân tử (kiểm chứng kết quả phân tích NH4+). Sai số tương đối giữa phương pháp phân tích và các phương pháp đo kiểm chứng đều dưới 15%. Kết quả cho thấy hàm lượng NH4-N và K+ trong mẫu đất lần lượt là 29,6 và 1055 mg/kg đất khô. Hàm lượng NH4-N và K+ trong mẫu nước lần lượt là 2,68 và 33,7 mg/L. Hàm lượng NH4-N trong mẫu nước ruộng vượt hơn 5 lần quy chuẩn kĩ thuật Quốc gia về chất lượng nước mặt bảo vệ đời sống thủy sinh. Key words: ammonium, potassium, contactless conductivity detector, agricultural soil 1. INTRODUCTION Nitrogen and potassium are essential nutrients for plant growth, survival, development and reproduction. However, there are not always enough of these nutrients in the soil for a plant to grow healthily so that many farmers and gardeners use fertilizers as the nutrients for the soil. Despite of the fact that fertilizers are beneficial to agriculture, excessive or improper fertilizer applying would induce to environmental and human health risks. The excess of nutrients in environment would cause soil pollution, surface water eutrophication, groundwater contamination etc. Elevated levels of nutrients accumulated in crop products could make unexpected symptoms and diseases to consumers [1]. For better agricultural land use, it is necessary to quantify the nutrient residues that are available to be released to environment. The extractions of the exchangeable forms of K+ and NH4+ can be performed by using salt solutions. Because NH4+ and K+ exhibit nearly identical characteristics, common extracting reagents for exchangeable potassium are ammonium salts. On the other hand, ammonium adsorbed on the exchange complex is normally removed by using potassium salts such as KCl or K2SO4 [2]. Thus, measurements of soil extracts for K+ and NH4+ would be strongly interfered by the matrix effects. Capillary Electrophoresis (CE) with Capacitively Coupled Contactless Conductivity Detection (C4D) is a simple and inexpensive method that can be applicable to simultaneously determine NH4+ and K+ in aqueous samples. However, the separations of two cations solely based on differences in their electrophoretic mobility seem to be a challenge. Numerous attempts were investigated to separate NH4+ and K+ in aqueous solutions. One of the most successful approach was the employment of complex-forming reactions. Among comlexing agents, 18-crown-6 ether 137 was widely used to separate the NH4+ and alkaline metal cations as it forms a stable complex with alkaline metal cations [3, 4, 5, 6, 7]. Adam J. Gaudry et al. separated an aqueous mixture comprising three inorganic cations (NH4+, Na+ and Li+) in 50 mM acetic acid/10 mM L-histidine /2.5 mM18- crown-6 ether electrolyte at pH 4.2. The limit of detections (LODs) of NH4+, Na+, and Li+ were 1.54, 2.26, and 3.06mg/L, respectively [3]. Another separation of alkaline metal cations (K+, Na+, Li+) in aqueous solutions was obtained by using the background electrolyte (BGE) of 12 mM histidine adjusted to pH 4 with acetic acid and 2 mM 18-crown-6. The LOD for K+ was of 1.5 μM [8]. Solutions containing NH4+, K+ and other cations were determined by Thanh Duc Mai et al. [5]. Background electrolyte solutions were His 12 mM (adjusted to pH 4 with acetic acid) in the presence of 2 mM of 18-crown-6; capillary was of 50 μm I.D., 36 cm effective length, and 50 cm total length, while separation voltage was 15 kV. The LODs were in the lower micromolar range and varied depending on species. Baseline separation between NH4+ and K+ was achieved at the concentration of less than100 μM for each cation [5]. Nevertheless, to our best knowledge, a simultaneous separation of NH4+ and K+ aqueous solutions extracted from soils by CE-C4D has not been reported. In the present study, operation parameters were set up in our laboratory according to previously published papers for measuring NH4+ and K+ in aqueous solutions. The optimizing amount of 18-crown-6 was systematically investigated. Appropriate extracting solutions used to remove exchangeable faction of each ion from soil were also examined. The preliminary conditions were applied to detect NH4+ and K+ in a water sample and the extracts of a soil sample collected from agricultural land in Vietnam. 2. MATERIALS AND METHODS 2.1. Materials All the chemicals such as NH4Cl, K2SO4, His, C6H9N302, HCl, NaOH etc were of analytical grade and purchased from Fluka or Merck. Ultrapure water system (Labconco, USA) with resistivity 18.2 MΩ was used to produce ultrapure water in all solutions and measurements. Stock solutions of ammonium and potassium ions were daily prepared. The background electrolyte (BGE) solutions were prepared with the buffer of histidine and acetic acid (His/Ace) (pH=4) in the presence of 18-Crown-6. The pH of solutions was controlled by using an HI 2215 Hanna Instruments 138 pH meter (Woonsocket, RI, USA). Fused silica capillaries of 50 μm I.D. and 365 μm O.D. with total length (Lt) of 50 cm and effective length (Leff) of 40 cm (purchased from Polymicro, Phoenix, AZ, USA) were used for separations. The capillaries were preconditioned with 1 M NaOH for 10minutes and then, deionized water for 10minutes prior to flushing with buffer solutions. Experiments were performed using the portable semi-automated CE system supported by 3Sanalysis JSC. ( (Figure 1). More detailed information about the instrument was given elsewhere in T. A. H. Nguyen et. al. [9]. Figure 1. Portable semi-automated CE-C4D instrument 2.2. Methods According to former references reviewed earlier, the BGE with the buffer adjusted to pH 4 was commonly used for the separations of the two cations using CE -C4D. In the present study, the buffer solution of His/Ace (pH=4) was selected. The high voltage of 20kV was applied for separation. Hydrodynamic injection of samples was carried out by setting the high-voltage end of the capillary at 15cm height. The injection time was 20 seconds. Table 1 summarized the operating parameters used in this research. 139 Table 1. Separation conditions of CE-C4D system for determine NH4+ and K+. BGE composition His/Ace buffer adjusted to pH 4 18-Crown-6 High voltage 20kV Detection C4D Capillary 365 μm O.D., 50 μm I.D.; Lt= 50cm; Leff= 40cm; Preconditioned Hydrodynamic injection 20s at 15cm height The BGE solutions with differences amount of 18-crown-6 were prepared in order to get optimum BGE composition for the separation of two cations. The concentrations of 18-crown-6 varied from 1 to 3mM. The concentrations of both NH4+ and K+ were 0,1mM. Appropriate soil extraction procedure was investigated. Simulated soil samples contaminated by 20mM of K+ and 2.2mM of NH4+ were prepared by adding certain amount of K+ and NH4+ solutions to uncontaminated soils. The samples were left for 1 day to reach to equilibrium, and then dried in desiccator before any further preparation. For K+ extraction, the CH3COONH4 solutions of 0.01M and 0.1M were used. The ion NH4+ was extracted with K2SO4 solutions of 0.001 and 0.1 M. The amount of 1.000 g dried soil and precisely 20ml of extracting agent were put into 50ml- falcon tubes and shaken for 1 hour using a shaker (Cole Parmer, 51704 Series). Finally, the mixture was centrifuged and filtered through 0.45μm membrane. The filtrates were used for the K+ and NH4+ measurements using CE-C4D. The levels of K+ and NH4+ were compared with Flame - Atomic Absorption Spectroscopy (F-AAS) and Ultraviolet– Visible Spectroscopy ((UV-Vis), respectively. One soil and one water samples were collected in agricultural regions Thuong Tin, Hanoi, Vietnam. The soil sample was immediately stored at 4oC and then dried in desiccator. The water sample was kept at 4oC and filtered through 0.45μm membrane before analysis. 3. RESULTS AND DISCUSSION It was known that potassium forms a stable complex with 18-crown-6 [10]. Consequently, the presence of 18- crown-6 in the BGE increases the migration time of K+ while the migration time of NH4+ stays unchanged. As clearly shown in Figure 2, the best separation was performed at the concentration of 2mM 18-crown-6. Lower concentrations than that of 2 mM 140 18-crown-6 would not be enough for reasonable separation of the two cations while higher concentrations seem to be not necessary. Thus, for further experiments the BGE solutions contain Tris/Ace buffer (pH=4); 2 mM 18- Crown-6. 15014013012011010090 10 mV Migration time (s) 1 2 3 4 5 6 Figure 2. Separation of NH4+ and K+ with BGE solutions containing different levels of 18- crown-6; Concentration of NH4+ and K+: 10-4M; Concentration of 18-crown-6: (1): Non 18-crown-6 contained; (2) 1.0 mM; (3) 1.5 mM; (4) 2.0 mM; (5): 2.5 mM. (6): 3 mM The calibration curves (six points) were extrapolated using standard addition method in order to avoid the matrix interference of the sample (Table 2). Linear range extended to the concentration of 200 μM for each cations with a correlation coefficient (r2) of at least 0.99. The LOD and LOQ values were calculated from peak areas corresponding to 3 and 10 times the baseline noise (S/N= 3 and S/N= 10), respectively. The LOD values (1.0 µM for NH4+ and 3.0 µM for K+) are comparable to previously reported results [5, 8]. Table 2. Calibration curve extrapolated by standard addition method Analyte Rangea (μM) r2 LODb (μM) LOQc (μM) NH4+ 50.0-200 0.9988 1.0 5.0 K+ 50.0-200 0.9909 3.0 10.0 a Six points b S/N: 3 c S/N: 10 As can be seen in Table 3, the best extraction efficiency was achieved when using 0.1M CH3COONH4 and 0.001M K2SO4 for extraction K+ and 141 NH4+, respectively. Higher concentrations of extractants are unfavorable due to the strong matrix effect to the separation of the two cations by CE, while lower ones are not enough to completely extract the exchangeable solutes. Table 3. Extraction of the simulated soil sample for K+ and NH4+ Extractant Extraction efficiency (%) Extraction for K+ CH3COONH4 0.01M 25.0 CH3COONH4 0.1M 99.2 Extraction for NH4+ K2SO4 0.001M 109.0 K2SO4 0.1M -a aunable to detect Figure 3 and 4 indicate good separation performances in the soil extract and water sample. Obtained results from CE-C4D are in good agreement with those from the confirmation method (F- AAS for K+ analysis and UV-Vis for NH4+ analysis) (Table 4). This suggests that the conditions found are applicable for detection of NH4+ and K+ in environmental samples. Levels of NH4- N and K+ determined in the real soil sample were of 29.6 and 1055 mg/kg dried soil, respectively. Those highly exceeded the normal level of NH4-N (10 mg/kg) and the excessive level of K (> 800mg/kg) defined by D.A. Horneck et al. [11]. The concentration of NH4-N in water sample was more than 5 times higher than the National technical regulation on surface water quality for protection of aquatic lives [12]. It should be noted that sufficiently high levels of NH4+ and K+ in agricultural run-off and soil would cause nutrient imbalance and serious environmental problem. 150140130120110100 10 mV Migration time (s) NH4 + K + 1 2 Figure 3. Separation of NH4+ and K+ in the extracts of soil sample: (1) Soil extract for NH4+; (2) Soil extract for K+, dilution factor (DF) was 50 times 142 150140130120110100 10 mV Migration time (s) NH4 + K + Figure 4. Simultaneous determination of NH4+ and K+ in water sample (DF was 6 times) Table 4. Concentrations of NH4+ and K+ in soil and water samples Type of sample Analyte Concentration* Confirmation (% difference) Soil sample K+ 1055 5,7a NH4-N 29.6 10.8b Water sample K+ 33,7 12,9a NH4-N 2.68 0.50b * mg/kg for soil sample; mg/L for water sample a confirmation by F-AAS method b confirmation by UV-Vis method 4. CONCLUSIONS The conditions for simultaneous determination of NH4+ and K+ by CE- C4D were optimized as follows: BGE: His/Ace buffer, 2 mM 18-crown-6; silica capillary: 365 μm O.D., 50 μm I.D.; Lt = 50cm; Leff = 40cm; voltage: 20V; hydrodynamic injection: 20s at 15cm height. The procedure are applicable to examine NH4+ and K+ in both soil and water samples. Excessive levels of NH4+ and K+ were detected in soil. The concentration of NH4+ in water sample was significantly higher than the National technical regulation on surface water quality for protection of aquatic lives. ACKNOWLEDGEMENT This research was partly funded by Hanoi Metropolitan University under the project number C.2015-19. REFERENCE [1] Stanley E. Manahan, (2009), Environmental chemistry, CRC Press. [2] Marc Pansu, Jacques Gautheyrou, (2006), Handbook of Soil Analysis: Mineralogical, Organic and Inorganic Methods, Springer. [3] Adam J. Gaudry, Michael C. Breadmore, and Rosanne M. Guijt (2013), In-plane alloy electrodes for 143 capacitively coupled contactless conductivity detection in poly (methylmethacrylate) electrophoretic chips, Electrophoresis, 34, 2980–2987. [4] Marko Stojkovic, Boris Schlensky, Peter C. Hauser, (2013), Referenced Capacitively Coupled Conductivity Detector for Capillary Electrophoresis, Electroanalysis, 25, No. 12, 2645 – 2650 [5] Thanh Duc Mai, Thi Thanh Thuy Pham, Hung Viet Pham, Jorge Saiz, Carmen García Ruiz and Peter C. Hauser, (2013), Portable capillary electrophoresis instrument with automated injector and contactless conductivity detection, Anal. Chem., 85, 2333−2339 [6] Thanh Duc Mai, Stefan Schmida, Beat Müller, Peter C. Hauser (2010), Capillary electrophoresis with contactless conductivity detection coupled to asequential injection analysis manifold for extended automated monitoring applications, Analytica Chimica Acta, 665, 1–6 [7] Pavel Kubáň and Peter C. Hauser (2009), Ten years of axial capacitively coupled contactless conductivity detection for CZE - a review. Electrophoresis, 30, pp.176-188. [8] Adam J. Gaudrya, Rosanne M. Guijtb, MirekMackaa, Joseph P. Hutchinsona, CameronJohnsa, Emily F. Hildera, Greg W. Dicinoski, Pavel N. Nesterenkoa, Paul R. Haddada, Michael C. Breadmorea, (2013), On- line simultaneous and rapid separation of anions and cations from a single sample using dual-capillary sequential injection-capillary electrophoresis, Analytica Chimica Acta, 781, 80– 87. [9] Thi Anh Huong Nguyen, Thi Ngoc Mai Pham, Thi Tuoi Doan, Thi Thao Ta, Jorge Sáizc, Thi Quynh Hoa Nguyen, Peter C. Hauser, Thanh Duc Mai, (2014), Simple semi-automated portable capillary electrophoresis instrument with contactless conductivity detection for the determination of β-agonists in pharmaceutical and pig-feed samples, Journal of Chromatography A, 1360, 305–311. [10] Harald Heiland, John A. Ringseth and Thorvald S. Brun (1979), Cation- Crown Ether Complex Formation in Water. II. Alkali and Alkaline Earth Cations and 12-Crown-4, 15-Crown-5, and 18-Crown-6, Journal of Solution Chemistry, Vol. 8, No. 11. [11] D. A. Horneck, D.M. Sullivan, J.S. Owen, and J.M. Hart (2011), Soil Test Interpretation Guide, EC 1478. Corvallis. [12] Ministry of Science, Technology and Environment (2011) QCVN 38:2011/BTNMT, National technical regulation on surface water quality for protection of aquatic lives.