Determination of Iodate by HPLC-UV after On-Line Electrochemical Reduction to Iodide

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1 Journal of Chromatographic Science 2015;53: doi: /chromsci/bmu053 Advance Access publication July 6, 2014 Article Determination of Iodate by HPLC-UV after On-Line Electrochemical Reduction to Iodide Tao Wang 1, Weimei Lin 2, Xueliang Dai 2, Lijun Gao 1, Bing Wang 2 and Dongqin Quan 1 * 1 Beijing Institute of Pharmacology and Toxicology, Beijing , People s Republic of China, and 2 Beijing Techmate Technology Corporation Limited, Beijing , People s Republic of China *Author to whom correspondence should be addressed. qdqwzb@163.com (D. Q.) These authors contributed equally to this work. Received 6 April 2013; revised 1 May 2014 In this study, a novel on-line pre-column electrochemical instrument (PECI) coupled with high-performance liquid chromatography (HPLC) was developed, and a novel method based on PEC HPLC-UV for amplifying the ultraviolet (UV) response of iodate (IO 3 2 ) was studied. Iodate undergoes reduction in the PECI, and the resulting I 2 was injected to an HPLC system and detected by a UV detector. For IO 3 2 analysis, conditions that can influence the reduction efficiency, including applied potential, ph value and salt concentration, were investigated in detail. In an appropriate condition, the UV response of iodate after passing through PECI was almost 10 times more than that of the initial form with good precision (relative standard deviation %). The detection limit and quantity limit were 9 and 20 ng, respectively. It can be concluded that the proposed method is simple and highly sensitive. reversed-phase or ion chromatography mode column and direct UV detection has been developed for the separation and quantification of periodate, iodate and iodide (8, 9). It is also aimed to develop a new method based on improving the UV absorption of iodate. The purpose of this approach was to develop a novel instrument including pre-column electrochemical instrument (PECI) and coupling it with HPLC-UV to determine IO 3 in iodized salt. In this study, IO 3 undergoes reduction in PECI and produces iodide (I 2 ): IO 3 þ 6e þ 6H þ! I þ 3H 2 O. Then, the resulting I 2 was retained and isolated by a column and detected by an UV detector. The UV response of I 2 is nearly 10 times more than that of IO 3. Therefore, the proposed method is simple and highly sensitive. Introduction Iodine deficiency is the greatest single cause of preventable brain damage and mental retardation (1, 2). Remarkable success has been achieved by common use of iodized salt in China since However, occasional adverse effects occurred. The principal effect is iodine-induced hyperthyroidism (3). Therefore, the China National Standard decided that iodized salt must contain: no less than 25 mg kg 21, and no more than 50 mg kg 21 of iodine. At the very beginning, salt was iodized by the addition of potassium iodide (KI); nowadays, the most common form of iodine in iodized salt is potassium iodate (KIO 3 ). Many methods based on different principles have been proposed for determination of iodate (IO 3 Þ, including spectrophotometry (4, 5), ion chromatography (6) and high-performance liquid chromatography (HPLC) (7). In recent years, ion chromatography has been used to determine iodide in seawater, urine and other natural samples. At the very beginning, ion chromatography equipped with an ultraviolet (UV) detector was developed in which the electrochemical detector (ED) was used to detect the iodide. However, some challenges still exit, in particular the instability of ED. On the other hand, formation of large amount of matrix ions (chloride, sulfate and other organic ions) impedes the determination of the target analysts by the way of saturating the active sites of ion-exchange column; and high price also hinders from spreading the ion chromatography in Chinese laboratories. HPLC with UV detector becomes a more significant method among all the methods of IO 3 analysis and it is more commonly used in conventional analysis laboratories. An HPLC system with a Experimental Apparatus The HPLC-UV system consisted of a 3001 high-pressure pump equipped with a 3010 degasser, a 3002 UV-visible detector, a 3006 autosampler and a 3004 column oven (Shiseido, Tokyo, Japan). A TSK-GEL-NH column (Tosoh, Tokyo, Japan) was used for analysis. The PECI system comprised an HTEC500 high-pressure pump equipped with a dependant degasser, a PEC-500 ED (Eicom, Kyoto, Japan), which was used as the reduction reactor, and a 3011 high-voltage switching six-way valve (Shiseido), which overcomes the high back pressure of the column and protect the electrode and cell in the PECI. The operating conditions for PECI-HPLC-UV are given in Table I. The samples were introduced by the autosampler (Shiseido), transferred by a mobile phase into the reduction reactor and then accommodated in the loop before detecting by the HPLC-UV system. Standard solution and reagents Reverse osmosis-milli Q water (18 MV) (Millipore Corp., Bedford, USA) was used for all solutions and dilutions. The iodide and iodate stock solutions were 1.0 mg ml 21, which were prepared by dissolving g of potassium iodide (Sigma, USA) and g of potassium iodate (Sigma, Milwaukee, USA) in 100 ml of water, respectively. The stock solutions were stored under dark condition at 48C. The working standard solutions were prepared by suitable dilution of the stock solutions with water. # The Author Published by Oxford University Press. All rights reserved. For Permissions, please journals.permissions@oup.com

2 Iodized salt was prepared by adding potassium iodate to sodium chloride (China National Pharmaceutical Group, Beijing, China). Acetonitrile (ACN) was purchased from Fisher Scientific (HPLC grade, Fair lawn, NJ, USA). Analytical grade sodium dihydrogen phosphate and phosphoric acid were bought from China Table I The Operating Conditions for PECI-HPLC-UV PECI system Mobile phase 1 Flow rate 0.2 ml min 21 Applied potential 2600 to 21,700 mv Peek loop volume 200 ml Switch internal time 30 s Cell temperature 358C HPLC-UV system Mobile phase 2 Stationary phase TSK-GEL-NH 2 Flow rate 0.5 ml min 21 Column temperature 358C Detected wavelength 215 nm Sodium dihydrogen phosphate buffer solution (ph , salt concentration mmol L 21 ) Acetonitrile 50 mmol L 21 sodium dihydrogen phosphate buffer solution (ph 3.0) (50 : 50, v/v) National Pharmaceutical Group (Beijing, China). All mobile phases were degassed prior to use either by vacuum or by ultrasonic wave. Procedure A schematic diagram of PECI-HPLC-UV system was illustrated in Figure 1. A PECI, consisting of a pump, a sampler, a high-voltage switching six-way valve and a coulometric ED, was used for online coupling with the HPLC-UV system. As shown in Figure 1, the analyzed chemicals were sent to the HPLC-UV system and detected by the UV detector after passing through the PECI system undergoing a reduction or oxidation. The designed analytical programs included two steps. Initially, when the six-way valve was in LOAD position, and IO 3 was delivered to the cell of the coulometric ED by pump 1 and reduced to I 2 at the electrode in PECI cell filled with mobile phase 1, then the resulting I 2 passed through PCEI and had been collected in the polyether ether ketone (peek) loop (Figure 1A). The process would take 30 s to get ready for the next stage. At step 2, the six-way was immediately switched to INJECT position and then pump 2 transferred the Figure 1. Schematic diagram of PECI-HPLC-UV system: (A) step 1 and (B) step 2. Determination of Iodate by HPLC-UV 281

3 mobile phase 2 to inject I 2 from opposite end of the peek loop to HPLC-UV system for subsequent analysis. I 2 was isolated by the column and detected by the UV detector. The high-voltage switching six-way valve made the cell and electrode of PECI link to atmosphere all the time. Therefore, the PECI on-line coupling with HPLC could avoid the high back pressure of the column (Figure 1B). Operating conditions of the PECI system To meet the requirement of determination, the high reduction efficiency (RE) ofio 3 in PECI was considered as a key target. Three operating conditions that can influence the RE of the PECI system, including applied potential, ph value and salt concentration of mobile phase 1, were investigated in detail. First, 10 mgml 21 of I 2 and IO 3 standard solutions were prepared by diluting their stock solutions with water. Then, IO 3 was analyzed by the PECI-HPLC-UV system in the form of I 2 under different conditions listed in Table II. Then, the RE value was calculated by the following equation: Reduction efficiency (RE) ¼ Amount of I produced Amount of total IO 3 100% where the amount of I 2 produced is calculated as moles of I 2 produced by IO 3 reduction and the amount of total IO 3 is the moles of IO 3 in the standard solution. Result Operating conditions of the PECI system Effect of ph The effect of ph on the RE of IO 3 is shown in Figure 2A. The figure shows that the RE values of IO 3 were very low within ph When ph was adjusted from 4.0 to 2.0, the RE was increased obviously and then kept unchanged when the ph further reduced from 2.0 to 1.0. Effect of potential and salt concentration Figure 2B shows the effect of salt concentration on RE of IO 3. When the salt concentration was changed in the range of mmol L 21, the RE of IO 3 increased from to 70.00%. High salt concentration is not necessary because it can increase the viscosity of electrolysis solution and leading to the slow release of I 2 from the electrode. High salt concentration may also suppress molecule or ion diffusion from the working electrode. The effect of the applied potential on RE was also Table II The Operation Condition Screening of PECI System Factor Level Other condition retained same ph 7.5, 5.0, 4.0, 3.0, 2.0 and mmol L 21 of salt concentration; 21,500 mv of applied potential Salt concentration 50, 120 and 200 mmol L 21 ph 3.0; 21,500 mv of applied potential Applied potential 2600, 2700, 2900, 21,100, 21,500 and 21,700 mv 120 mmol L 21 of salt concentration; ph 3.0 Figure 2. The effect of ph (A), salt concentration (B) and applied potential (C) on reduction deficiency of IO 3 (10.0 mg ml 21 ). 282 Wang et al.

4 investigated, and results were showed in Figure 2C. The more negative applied potential, the more intensive reduction takes place at the electrode. Six levels were designed by us for the applied potential in the study such as 2600, 2700, 2800, 2110, 21,500 and 21,700 mv. The RE value with 21,500 and 21,700 mv was and 67.44%, respectively. Recovery and linearity AsshowninTableIII, the recovery of the method was in the range of % and the relative standard deviation (RSD) was in the range of %. The curves (y ¼ x ) were linear in the range of mg ml 21 for IO 3 with a correlation coefficient of or greater. The detection limit and quantity limit were 9 and 20 ng, respectively. The IO 3 in iodized salt was determined by PECI on-line coupling with high-performance liquid chromatograph (PECI-HPLC), and results are given in Table III (8). Discussion An appropriate ph value can improve the reduction and oxidation at an electrode. In particular, acidic ph is beneficial to reduction and alkaline ph is beneficial to oxidation. According to Faraday s law of electrolysis, the mass of produced substance at an electrode by an electrochemical reaction is proportional to the number of mass of electrons transferred at that electrode. When a molecule or ion undergoes oxidation or reduction in a solution, it may undergo a three-step process: diffusion to the electrode surface, oxidation or reduction and diffusion away from the vicinity of the working electrode. For the first step, IO 3 transferred from the bulk solution to the electrode surface was affected by the flow rate, diffusion coefficient and viscosity of the mobile phase 1. In general, diffusion of ion or molecule in liquid at room temperature is relatively slow, and either too fast or too slow flow of mobile phase 1 is not appropriate for diffusion. Therefore, 358C cell temperature and flow rate 0.2 ml min 21 were chosen in the present study. In the second step, electron transfer is primarily determined by applied potential of the electrode. As shown in Figure 3, the reduction of IO 3 was very weak with applied potential ranging from 2600 to 21,100 mv. With the increasing in applied potential from 21,100 to 21,700 mv, the amount of I 2 increased significantly. To obtain high reduction of IO 3, 200 mmol L 21 sodium dihydrogen phosphate (ph 2.0), mobile phase 1, 21,700 mv applied potential, 0.2 ml min 21 flow rate and 378C cell temperature were selected as operating conditions in this work. The UV response of iodide after reduction is almost 10 times more than that of the initial form (Figure 3). On the other hand, as the iodate was detected through the formation of iodide and this ion can be effectively isolated from chloride in the sample medium, a low interference exists during the analysis. It can be concluded that the method is specific and sensitive by using the PECI instrument under the selected conditions. Conclusion A novel PECI on-line coupled with HPLC was developed, and a novel method based on PEC-HPLC-UV for amplifying the UV response of iodate (IO 3 Þ was described. The proposed method was simple and highly sensitive with good precision. The PECI will be used widely in the field of HPLC analysis in the future. Table III Determination of Potassium Iodate in Salt Iodized (n ¼ 3) Sample Added iodate (mg kg 21 ) Round iodate (mg kg 21 ) Recovery% RSD% Acknowledgments We acknowledge Tijun Zhou of Shiseido China Co., Ltd for his assistance in equipments repair. We also thank Peng Tan and Shangyi Chen of Beijing Techmate Technology Corporation Limited and Haiyan Li of Beijing Institute of Pharmacology and Toxicology for helpful discussions and technical assistance. Figure 3. The chromatography diagram of IO 3 (10.0 mg ml 21 ), initial form (about RT 13.5 min) and after reduction (about RT 8.0 min). References 1. Ranganathan, S., Reddy, V.; Human requirements for iodine & safe use of iodised salt; Indian Journal of Medical Research, (1995); 102: Delange, F., Lecomte, P.; Iodine supplementation: benefits outweigh risks; Drug Safety, (2000); 22: Delange, F., de Benoist, B., Alnwick, D.; Risks of iodine-induced hyperthyroidism after correction of iodine deficiency by iodized salt; Thyroid, (1999); 9: May, W., Wu, D., Eastman, C.P., Maberly, G.; Evaluation of automated urinary iodine methods: problems of interfering substances identified; Clinical Chemistry, (1990); 36: Ford, H.C., Johnson, L.A.; Ascorbic acid interferes with an automated urinary iodide determination based on the ceric-arsenious acid reaction; Clinical Chemistry, (1991); 37: Kumar, S.D., Maiti, B., Mathur, P.K.; Determination of iodate and sulphate in iodized common salt by ion chromatography with conductivity detection; Talanta, (2001); 53: Determination of Iodate by HPLC-UV 283

5 7. Li, H.B., Chen, F., Xu, X.R.; Determination of iodide in seawater and urine by size exclusion chromatography with iodine starch complex; Journal of Chromatography A, (2001); 918: Huang, Z.P., Subhani, Q., Zhu, Z.Y., Guo, W.Q., Zhu, Y.; A single pump cycling-column-switching technique coupled with homemade high exchange capacity columns for the determination of iodate in iodized edible salt by ion chromatography with UV detection; Food Chemistry, (2013); 139(1 4): Sajonz, P.; Separation of periodate, iodate and iodide on a C-18 stationary phase. Dependence of the retention on the temperature and solvent composition. Monitoring of an oxidative cleavage reaction; Chromatographia, (2006); 64: Wang et al.

High Performance Liquid Chromatography

High Performance Liquid Chromatography What is HPLC? It is a separation technique that involves: Injection of small volume of liquid sample Into a tube packed with a tiny particles (stationary phase).