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Environment

Preparation of DMMTAV and DMDTAV Using DMAV for Environmental Applications: Synthesis, Purification, and Confirmation

Published: March 9, 2018 doi: 10.3791/56603
Automatic Translation
1, 2, 1, 1
1Korea Basic Science Institute, Seoul Center, 2Natural Science Research Institute, Yonsei University

Summary

This article presents modified experimental protocols for dimethylmonothioarsinic acid (DMMTAV) and dimethyldithioarsinic acid (DMDTAV) synthesis, inducing dimethylarsinic acid (DMAV) thiolation through mixing of DMAV, Na2S, and H2SO4. The modified protocol provides an experimental guideline, thereby overcoming limitations of the synthesis steps that could have caused experimental failures in quantitative analysis.

Abstract

Dimethylated thioarsenicals such as dimethylmonothioarsinic acid (DMMTAV) and dimethyldithioarsinic acid (DMDTAV), which are produced by the metabolic pathway of dimethylarsinic acid (DMAV) thiolation, have been recently found in the environment as well as human organs. DMMTAV and DMDTAV can be quantified to determine the ecological effects of dimethylated thioarsenicals and their stability in environmental media. The synthesis method for these compounds is unstandardized, making replicating previous studies challenging. Furthermore, there is a lack of information about storage techniques, including storage of compounds without species transformation. Moreover, because only limited information about synthesis methods is available, there may be experimental difficulties in synthesizing standard chemicals and performing quantitative analysis. The protocol presented herein provides a practically modified synthesis method for the dimethylated thioarsenicals, DMMTAV and DMDTAV, and will help in the quantification of species separation analysis using high performance liquid chromatography in conjunction with inductively coupled plasma mass spectrometry (HPLC-ICP-MS). The experimental steps of this procedure were modified by focusing on the preparation of chemical reagents, filtration methods, and storage.

Introduction

Since dimethylarsinic acid (DMAV) has been demonstrated to exhibit both acute toxicity and genotoxicity due to undergoing methylation and thiolation upon ingestion1,2, the metabolic pathway of arsenic thiolation has been intensively studied both in vitro and in vivo3,4 as well as in environmental media (e.g., landfill leachate)5,6. Previous studies have found both reduced and thiolated analogs of DMAV in living cells, for example, dimethylarsinous acid (DMAIII), dimethylmonothioarsinic acid (DMMTAV), and dimethyldithioarsinic acid (DMDTAV)7,8,9, with dimethylated thioarsenicals such as DMMTAV exhibiting greater toxicity than other known inorganic or organic arsenicals10. The abundance of highly toxic thioarsenicals has serious environmental implications, since they may pose a risk to humans and the environment under highly sulfidic conditions11. However, the mechanisms of DMMTAV and DMDTAV (trans)formation and their fates in environmental media still require further study. Thus, the quantitative analysis of thioarsenicals is required to improve understanding of the environmental effects of DMMTAV and DMDTAV.

Although standard chemicals are the key requirement for quantitative analysis, the standards of DMMTAV and DMDTAV are difficult to obtain by replicating previous studies, owing to the high risk of species transformation into other species and unstandardized synthesis procedures12. Moreover, the methods referenced have limitations that may lead to practical difficulties in synthesizing the standard chemicals and performing quantitative analysis. DMMTAV and DMDTAV are commonly prepared by mixing DMAV, Na2S, and H2SO4 in a certain molar ratio1 or bubbling H2S gas through a solution of DMAV 13,14. The bubbling method features substitution of oxygen by sulfur using a direct supply of H2S gas, which, is highly toxic and difficult to control for an inexperienced user. Conversely, the above mixing method1, widely used for the qualitative analysis of DMMTAV and DMDTAV in environmental sudies5,6,12, features the thiolation of DMAV with H2S generated by mixing Na2S and H2SO4 and produces DMMTAV and DMDTAV, allowing easier stoichiometric control to produce target chemicals, as compared to the direct use of H2S gas.

The reference mixing method procedures1,3,4,8,15 mentioned in this study exhibit limitations in some of their critical experimental steps, which might lead to experimental failure. For example, the details of specific solvent (i.e., deionized water) preparation and the extraction and crystallization of the synthesized arsenicals are over-abbreviated or not described in sufficient detail. Such dispersed and limited information on procedural steps might lead to the inconsistent formation of thioarsenicals and unreliable quantification analysis. Therefore, the modified protocol developed herein describes the synthesis of DMMTAV and DMDTAV stock solutions with quantitative species separation analysis.

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Protocol

1. Synthesis of DMMTAV

  1. Chemical preparation and molar ratio mixing of DMAV, Na2S, and H2SO4
    NOTE: DMAV:Na2S:H2SO4 = 1:1.6:1.6
    1. Dissolve 5.24 g of DMAV in 40 mL deionized and N2-purged (purged for at least 30 min) water in a 50 mL centrifuge tube.
    2. Prepare Na2S reagent by dissolving 14.41 g of Na2S·9H2O in 50 mL deionized and N2-purged water in a 250 mL flask.
    3. Prepare H2SO4 reagent by adding 3.3 mL of concentrated sulfuric acid (96%) to 40 mL of deionized and N2-purged water in a 50 mL centrifuge tube.
      NOTE: Final molar ratio of DMAV:Na2S:H2SO4 = 1:1.6:1.6 1,3,4,8,15.
    4. Add the prepared 40 mL of DMAV solution (step 1.1.1) to the 50 mL of Na2S solution contained in the 250 mL flask (step 1.1.2). Rinse the tube containing DMAV with 10 mL of purged water and add this to the 250 mL flask as well.
    5. Close the flask with a three-hole rubber stopper fitted with glass tubes. Use the glass tubes for N2 gas inflow, outflow, and H2SO4 solution inlet, respectively. Immediately after closing the flask, allow N2 gas flow into the flask.
      NOTE: Gas pressure should be maintained to flow over the surface of the reaction solution without splashing.
    6. Connect acid resistant tubing to the glass tube of the H2SO4 solution inlet with a 50 mL syringe to add the prepared 40 mL of H2SO4 solution (step 1.1.3). Add 40 mL of H2SO4 solution, slowly and in a stepwise manner.
      Caution: Upon adding sulfuric acid, white fumes will be generated; use a well-ventilated fume hood.
    7. Monitor color change of the reaction mixture in the flask while adding sulfuric acid at regular intervals. Maintain an interval of 4 - 5 mL drops of H2SO4; the mixture should be a white cloudy solution.
      NOTE: Instantaneous yellow precipitation might appear due to rapid addition of concentrated sulfuric acid.
    8. Ensure the reaction solution has been standing for 1 h since the start of step 1.1.4.
  2. DMMTAV extraction using liquid-liquid extraction method
    1. After 1 h, pour the reaction solution into a separatory funnel containing around 200 mL of diethyl ether.
    2. Shake the funnel for 5 - 10 min, releasing gas several times by switching the stopcock.
      NOTE: Synthesized DMMTAV will be transferred to the upper layer of diethyl ether (0.713 g·mL-1).
      Caution: Diethyl ether gas might be harmful; use a well-ventilated fume hood.
    3. Collect the reaction solution in a beaker, and separately collect the diethyl ether containing DMMTAV in a bottle. Place the reaction solution back in the same separatory funnel and add about 200 mL of fresh diethyl ether for reshaking. Repeat steps 1.2.2 - 1.2.3 three times.
    4. Pour the collected diethyl ether from step 1.2.3 into the same separatory funnel again, and add about 100 mL N2-purged deionized water. Shake for 5 - 10 min, and discard the N2-purged deionized water and a few mL of diethyl ether for purity purposes. Collect the remaining diethyl ether in a glass petri dish (minimum inner diameter of 160 mm and minimum height of 50 mm).
    5. Transfer the glass petri dish into a N2 atmosphere glove box to prevent species transformation.
      Caution: Ensure the solvent is not pulled into the vacuum pump through the outlet of the pass box.
    6. Dry until a white precipitate of dimethylmonothioarsinate (crystalized DMMTAV) is formed on the glass petri dish.
      NOTE: The protocol can be paused here.
  3. Verification of synthesized DMMTAV and storage
    1. Take the white precipitate of crystalized DMMTAV, and measure and record its total weight.
      Caution: Use a well ventilated fume hood or glove box to prevent inhalation of hydrogen sulfide gas.
    2. Dissolve crystalized DMMTAV in 50 mL of N2-purged deionized water, and filter the yellow precipitate through a 0.2-µm syringe filter.
    3. Assume the total As of used DMAV is converted into DMMTAV, i.e.≈9,649 mg As·L-1. Dilute DMMTAV stock solution to ≈1 mg·L-1 and ≈40 µg·L-1 for verification analysis using Electrospray ionization Mass Spectromtery (ESI-MS) and HPLC-ICP-MS, respectively.
    4. Analyze m/z of DMMTAV using ESI-MS11,16 and fragments at m/z 155 in the positive-ion mode or at m/z 153 in the negative-ion mode (Table 1).
      NOTE: See reference values of m/z (Table 1).
    5. Analyze chromatogram of DMMTAV in the stock solution using HPLC-ICP-MS11,16,17 with appropriate eluent conditions and confirm a major peak is found at the retention time described in the literature.
      NOTE: Purity of synthesized DMMTAV should be calculated using the analysis results from step 1.3.5.
    6. Analyze the total arsenic concentration using ICP-MS after acid digestion11 and calculate the true DMMTAV concentration in synthesized DMMTAV stock solution using dilution factors and purity, as in the following equation:
      Analyzed total As concentration (µg·L-1) · Dilution factor · Purity (%) = True DMMTAV concentration in DMMTAV stock solution (µg·L-1)
    7. Store DMMTAV stock solution at 4 °C in the dark for further quantitative speciation analysis18.

2. Synthesis of DMDTAV

  1. Chemical preparation and molar ratio mixing of DMAV , Na2S, and H2SO4
    NOTE: DMAV:Na2S:H2SO4 = 1:7.5:7.5
    1. Dissolve 1.38 g of DMAV in 40 mL deionized water in a 50 mL centrifuge tube.
    2. Prepare Na2S reagent by dissolving 18.01 g of Na2S·9H2O in a 50 mL deionized water in a 250 mL flask.
    3. Prepare H2SO4 reagent by adding 4 mL of concentrated sulfuric acid (96%) to 40 mL of deionized water contained in a 50 mL centrifuge tube.
      NOTE: Final molar ratio of DMAV:Na2S:H2SO4 = 1:7.5:7.51,3,4,8,15.
    4. Add the prepared 40 mL of DMAV solution (step 2.1.1) to the 50 mL of Na2S solution contained in the 250 mL flask (step 2.1.2). Rinse the tube containing DMAV with 10 mL of deionized water and add this to the 250 mL flask as well.
    5. Add the prepared 40 mL of H2SO4 solution (step 2.1.3), slowly and in a stepwise manner, into the flask.
    6. Monitor the color change of the reaction mixture in the flask while adding sulfuric acid at regular intervals. Maintain an interval of 4 - 5 mL drops of H2SO4; the mixture should be a white/yellow cloudy solution.
      NOTE: Instantaneous yellow precipitation might appear due to a rapid addition of concentrated sulfuric acid.
      Caution: Upon adding sulfuric acid, white fumes will be generated; use a well ventilated fume hood.
    7. Maintain the reaction solution in the flask overnight without covering.
      NOTE: The protocol can be paused here.
  2. DMDTAV extraction using solid-phase extraction (SPE) method
    1. After the overnight standing reaction, filter the reaction solution using a C18 syringe type silica-based SPE in order to trap synthesized DMDTAV on the resin.
      Caution: Use a well-ventilated fume hood.
    2. Prepare 10 mM ammonium acetate (pH 6.3) by dissolving 0.77 g ammonium acetate in 1 L of deionized water. Elute a sufficient volume of 10 mM ammonium acetate through the C18 syringe (step 2.2.1) to extract the adsorbed DMDTAV. Collect the filtered ammonium acetate in a glass Petri dish (minimum inner diameter of 160 mm and minimum height of 50 mm).
    3. Transfer the glass petri dish into a N2 atmosphere glove box to prevent species transformation.
      Caution: Ensure solvent is not pulled into the vacuum pump through the outlet of the pass box.
    4. Dry until a white precipitate of dimethyldithioarsinate is formed (crystalized DMDTAV) on the glass petri dish.
      NOTE: The protocol can be paused here.
  3. Verification of synthesized DMDTAV and storage
    1. Take the white precipitate of crystalized DMDTAV, and measure its total weight, recording the measurement.
      Caution: Use a well-ventilated fume hood or glove box to prevent inhalation of hydrogen sulfide gas.
    2. Dissolve crystalized DMDTAV in 50 mL of N2-purged deionized water in a 50 mL centrifuge tube, and filter any precipitate through a 0.2-µm syringe filter.
    3. Assume that the total As of used DMAV is converted into DMDTAV, i.e.≈2,539 mg As·L-1. Dilute DMDTAV stock solution to ≈1 mg·L-1 and ≈40 µg·L-1 for verification analysis using ESI-MS and HPLC-ICP-MS, respectively.
    4. Analyze m/z of DMDTAV using ESI-MS11,16 and fragments at m/z 171 in the positive-ion mode or at m/z 169 in the negative-ion mode (Table 1).
      NOTE: See reference values of m/z (Table 1).
    5. Analyze the chromatogram of DMDTAV in the stock solution using HPLC-ICP-MS11,16,17 with appropriate eluent conditions and confirm that a peak is found at the retention time described in the literature.
      NOTE: Purity of synthesized DMDTAV should be calculated using the analysis results from step 2.3.5.
    6. Analyze the total arsenic concentration using ICP-MS after acid digestion11 and calculate the true DMDTAV concentration in synthesized DMDTAV stock solution using dilution factors and purity as the following equation:
      Analyzed total As concentration (µg·L-1) · Dilution factor · Purity (%) = True DMDTAV concentration of DMDTAV in stock solution (µg·L-1)
    7. Store DMDTAV stock solution at 4 °C in the dark for further quantitative speciation analysis18.

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Representative Results

Since DMMTAV has been mistakenly prepared by the DMAIII synthesis method19, verification of synthesized DMMTAV and DMDTAV is a critical step for synthesis and extraction and determining the ideal standard chemical materials. Synthesized chemicals can be verified by the peak of DMMTAV (MW 154 g·mol-1) and DMDTAV (MW 170 g·mol-1) mass-to-charge ratio (m/z) using either the positive or negative ion mode of electrospray ionization-mass spectrometer (ESI-MS) through real-time injection. The reference values of m/z are listed in Table 119. Additional verification of the successful synthesis of DMMTAV and DMDTAV was conducted by comparing the species separation analysis results of the retention time (RT) of major peaks to reference data using HPLC-ICP-MS. Figure 1 shows similar RT of major and minor peaks, DMAV and DMMTAV (Figure 1a) or DMDTAV (Figure 1b), using 1.0 mL·min-1 of 5 mM formic acid as an eluent and a C18 Liquid Chromatography (LC) column as described in17. Note that the RT of the major peaks may vary depending on the instrumental and eluent conditions, and which LC column is used. Species included in the stock solutions of DMMTAV and DMDTAV should be examined prior to every analysis, although this protocol suggests storage conditions of 4 ˚C in the dark, which maintains synthesized DMMTAV and DMDTAV with, respectively, 2.2% and 5.8% transformation during the 13 weeks of analysis (Figure 2).

Figure 1
Figure 1: HPLC-ICP-MS chromatogram of the synthesized DMMTAV and DMDTAV. 1: DMAV, 2: DMDTAV, and 3: DMMTAV were measured as major peaks at 3.8 min, 5.9 min, and 8.0 min in each stock solutions of a: DMMTAV and b: DMDTAV, which were corresponded to those reported by Li et al. in 201017. Instrumental conditions of ICP-MS were 1550 V RF power and 50 µL injection volume. A C18 column was used with 5 mM formic acid as an eluent17. Please click here to view a larger version of this figure.

Figure 2
Figure 2: Stability of DMMTAV and DMDTAV at 4 ˚C in the dark. Percentage changes of As species distribution in each of the stock solutions DMMTAV (a) and DMDTAV (b) for 13 weeks. Please click here to view a larger version of this figure.

Synthesized stock solution  ion mode Fragments  m/z References
DMMTAV Positive [Me2As(SH)OH]+ 155 13,14,19,17,21,24
[Me2As(OH)2]+ 139 19,17
[Me2AsS]+ 137 13,19,17,24
[Me2As(S)SAsMe2]+ 275 13
Negative [Me2AsOS]- 153 17,5,23
[MeAsSO]- 138 17,5
[AsSO]- 123 17,5
DMDTAV Positive [Me2As(SH)2]+ 171 19,17
[Me2As(SH)OH]+ 155 19
[Me2AsS]+ 137 19,17
Negative [Me2AsS2]- 169 20,17,5,22
[MeAsS2]- 154 20,17,5
[AsS2]- 139 17,5,22

Table 1: Suggested structure of synthesized DMMTAV and DMDTAV, and minor fragment ions by positive and/or negative ion mode of ESI-MS. The list of DMMTAV, DMDTAV, and minor fragments m/z measured by ESI-MS was reproduced from literature5,13,14,17,19,20,21,22,23,24. Note that m/z peaks may vary with instrumental conditions and/or matrices of the stock solutions.

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Discussion

The developed protocol has clarified critical steps that previous studies1,3,4,8,15 omitted or abbreviated, which may have led to difficulties with or failure during DMMTAV and DMDTAV synthesis. As DMMTAV is oxidation-sensitive1,5, chemical reagents for its synthesis were prepared using N2-purged deionized water (steps 1.1.1 - 1.1.3) to prevent the possible retardation of DMAV thiolation and the oxidation of DMMTAV. DMMTAV prepared using this protocol had a purity of 92%. The reference DMDTAV extraction method featured silica-based C18 column extraction with ammonium acetate or phosphate buffer solution as an eluent,3,4,7,8 with the purity of the thus prepared DMDTAV varying depending on the column compartments used, which is not described in references studies3,4,8. In contrast, the use of disposable SPE in the developed protocol (step 2.2.1) allowed the extraction of 88% pure DMDTAV. In addition, previous methods of crystallizing DMDTAV referenced only a freeze-drying procedure20, whereas in this protocol, simple drying of the extracted DMDTAV solution in an atmosphere of N2 inside the glove box (step 2.2.3) produced a white residue of crystalized dimethyldithioarsinate.

Identity verification of the synthesized DMMTAV and DMDTAV is a critical step for determining ideal standard chemical. In our previous work16 based on this protocol, major fragments at m/z 155 in the positive-ion mode and m/z 169 in the negative-ion mode were detected by ESI-MS analysis of as-synthesized DMMTAV and DMDTAV, respectively. The latter fragment was ascribed to [CH3]2AsS(=S)S]- (i.e., [M-H]-), in good agreement with previous results5,17,20,22, whereas the fragment at m/z 155 was assigned to [CH3]2As(=S)(OH) + H]+ (i.e., [M+H]+), again, in agreement with references13,14,17,19,21,24. Since ESI-MS analysis cannot be used as the only verification method, major peaks in the chromatograms of DMMTAV and DMDTAV stock solutions obtained by HPLC-ICP-MS were compared to those reported by Li et al.17 (Figure 1). Although DMAIII is known as an intermediate produced in the initial stage of DMMTAV formation1,7,8,25, it exhibits low stability, disappearing within 70 min19 and therefore is not detectable by this procedure (Figure 1).

Another goal of this study was to suggest storage conditions for DMMTAV and DMDTAV stock solutions with fewer impurities to prevent sulfide-to-oxide conversion and thus achieve greater stability18. The storage conditions used here (i.e., 4 ˚C in the dark) allowed the preservation of the synthesized DMMTAV and DMDTAV as major species in stock solutions (Figure 2) for four weeks, with no drastic decomposition observed even after 13 weeks. Although the solvent pH (i.e., that of deionized water) may affect the transformation of species during storage due to the presence of native10 and excess sulfide species originating from chemical reagents such as HS- or H2S, the stock solutions maintained neutral pH without significant transformation of the species contained therein (Figure 2). Therefore, stock solutions could be stored at 4 ˚C in the dark for 13 weeks prior to quantitative speciation analysis.

In this study, the reference molar ratio mixing method of synthesizing DMMTAV and DMDTAV 1,3,4,8,15 was modified to produce stable DMMTAV and DMDTAV stock solutions for HPLC-ICP-MS quantitative analysis. Due to a lack of critical step details, including not only chemical reagent preparation, extraction, and crystallization steps, but also the storage conditions of DMMTAV and DMDTAV, stock solutions had to be modified and optimized. Therefore, stock solutions prepared using this protocol are sufficiently applicable for quantitative analysis of DMMTAV and DMDTAV for environmental monitoring purposes.

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Disclosures

The authors have nothing to disclose.

Acknowledgments

This research was supported by Basic Science Research program (Project number: 2016R1A2B4013467) through the National Research Foundation of Korea (NRF) funded by the Ministry of Science, ICT & Future Planning 2016 and also supported by Korea Basic Science Institute Research Program (Project number: C36707).

Materials

Name Company Catalog Number Comments
Cacodylic acid Sigma-Aldrich 20835-10G-F
Sodium sulfide nonahydrate Sigma-Aldrich S2006-500G
Sulfuric acid 96% J.T.Baker 0000011478
Ammonium acetate Sigma-Aldrich A7262-500G
Formic acid 98% Wako Pure Chemical Industries, Ltd. 066-00461
Diethyl ether (Extra Pure) Junsei Chemical 33475-0380
Adapter cap for 60 mL Bond Elut catridges Agilent Technologies 12131004 Syringe type of SPE
Bond Elut C18 cartridge Agilent Technologies 14256031 Syringe type of SPE
HyPURITY C-18 Thermo Scientific 22105-254630 5 um, 125 x 4.6 mm
Glovebox Chungae-chun, Rep. of Korea Customized 
Agilent 1260 Infinity Bio-inert LC Agilent Technologies DEAB600252, DEACH00245
Agilent Technologies 7700 Series ICP-MS Agilent Technologies JP12031510
Finnigan LCQ Deca XP MAX Mass Spectrometer System Thermo Electron Corporation LDM10627

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References

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  2. Kuroda, K., et al. Microbial metabolite of dimethylarsinic acid is highly toxic and genotoxic. Toxicol. Appl. Pharmacol. 198, 345-353 (2004).
  3. Naranmandura, H., Iwata, K., Suzuki, K. T., Ogra, Y. Distribution and metabolism of four different dimethylated arsenicals in hamsters. Toxicol. Appl. Pharmacol. 245, 67-75 (2010).
  4. Naranmandura, H., et al. Comparative toxicity of arsenic metabolites in human bladder cancer EJ-1 cells. Chem. Res. Toxicol. 24, 1586-1596 (2011).
  5. Wallschlager, D., London, J. Determination of methylated arsenic-sulfur compounds in groundwater. Environ. Sci. Technol. 42, 228-234 (2008).
  6. Zhang, J., Kim, H., Townsend, T. Methodology for assessing thioarsenic formation potential in sulfidic landfill environments. Chemosphere. 107, 311-318 (2014).
  7. Shimoda, Y., et al. Proposal for novel metabolic pathway of highly toxic dimethylated arsenics accompanied by enzymatic sulfuration, desulfuration and oxidation. Trace Elem. Med. Biol. 30, 129-136 (2015).
  8. Naranmandura, H., Suzuki, T. K. Formation of dimethylthioarsenicals in red blood cells. Toxicol. Appl. Pharmacol. 227, 390-399 (2008).
  9. Leffers, L., Ebert, F., Taleshi, S. M., Francesconi, A. K., Schwerdtle, T. In vitro toxicological characterization of two arsenosugars and their metabolites. Mol. Nutr. Food Res. 57, 1270-1282 (2013).
  10. Wang, Q. Q., Thomas, J. D., Naranmandura, H. Important of being thiomethylated: Formation, Fate and Effects of methylated thioarsenicals. Chem. Res. Toxicol. 25, 281-289 (2015).
  11. Kim, Y. T., Lee, H., Yoon, H. O., Woo, N. C. Kinetics of dimethylated thioarsenicals and the formation of highly toxic dimethylmonothioarsinic acid in environment. Environ. Sci. Technol. 50, 11637-11645 (2016).
  12. Cullen, W. R., et al. Methylated and thiolated arsenic species for environmental and health research - A review on synthesis and characterization. J. Environ. Sci. 49, 7-27 (2016).
  13. Fricke, M., et al. Chromatographic separation and identification of products form the reaction of dimethylarsinic acid with hydrogen sulfide. Chem. Res. Toxicol. 18, 1821-1829 (2005).
  14. Fricke, M., Zeller, M., Cullen, W., Witkowski, M., Creed, J. Dimethylthioarsinic anhydride: a standard for arsenic speciation. Anal. Chim. Acta. 583, 78-83 (2007).
  15. Suzuki, K. T., Iwata, K., Naranmandura, H., Suzuki, N. Metabolic differences between twon dimethylthioarsenicals in rats. Toxicol. Appl. Pharmacol. 218, 166-173 (2007).
  16. Jeong, S., et al. Development of a simultaneous analytical method to determine arsenic speciation using HPLC-ICP-MS: Arsenate, arsenite, monomethylarsonic acid, dimethylarsinic acid, dimethyldithioarsinic acid, and dimethylmonothioarsinic acid. Microchem. J. 134, 295-300 (2017).
  17. Li, Y., Low, C. -K., Scott, A. J., Amal, R. Arsenic speciation in municipal landfill leachate. Chemosphere. 79, 794-801 (2010).
  18. Conklin, D. S., Fricke, W. M., Creed, A. P., Creed, J. T. Investigation of the pH effects on the formation of methylated thio-arsenicals, and the effects of pH and temperature on their stability. J. Anal. At. Spectrom. 23, 711-716 (2008).
  19. Hansen, H. R., Raab, A., Jaspara, M., Milne, F. B., Feldmann, J. Sulfur-containing arsenical mistaken for dimethylarsinous acid [DMA(III)] and identified as a natural metabolite in urine: major implications for studies on arsenic metabolism and toxicity. Chem. Res. Toxicol. 17, 1086-1091 (2004).
  20. Mandal, B. K., Suzuki, K. T., Anzai, K., Yamaguchi, K., Sei, Y. A SEC-HPLC-ICP-MS hyphenated technique for identification of sulfur-containing arsenic metabolites in biological samples. J. Chromatogr. B. 874, 64-76 (2008).
  21. Bartel, M., Ebert, F., Leffers, L., Karst, U., Schwerdtle, T. Toxicological characterization of the inorganic and organic arsenic metabolite thio-DMAV in cultured human lung cells. J. Toxicol. 2011, (2011).
  22. An, J., et al. Formation of dimethyldithioarsinic acid in a simulated landfill leachate in relation to hydrosulfide concentration. Environ. Geochem. Health. 38, 255-263 (2016).
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  24. Alava, P., et al. HPLC-ICP-MS method development to monitor arsenic speciation changes by human gut microbiota. Biomed. Chromatogr. 26, 524-533 (2012).
  25. Kurosawa, H., et al. A novel metabolic activation associated with glutathione in dimethylmonoarsinic acid (DMMTAV)-induced toxicity obtained from in vitro reaction of DMMTAV with glutathione. J. trace Elem. Med. Biol. 33, 87-94 (2016).

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DMMTAV DMDTAV DMAV Environmental Applications Synthesis Purification Confirmation Speciation Analysis Thioarsenicals Experimental Guideline Modified Technique Chemical Standards Fume Hood Dimethylmonothioarsinic Acid Sodium Sulfide Sulfuric Acid Deionized Water Nitrogen Gas Rinse Flask Rubber Stopper Glass Tubes Nitrogen Gas Line
Preparation of DMMTA<sup>V</sup> and DMDTA<sup>V</sup> Using DMA<sup>V</sup> for Environmental Applications: Synthesis, Purification, and Confirmation
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Lee, H., Kim, Y. T., Jeong, S.,… More

Lee, H., Kim, Y. T., Jeong, S., Yoon, H. O. Preparation of DMMTAV and DMDTAV Using DMAV for Environmental Applications: Synthesis, Purification, and Confirmation. J. Vis. Exp. (133), e56603, doi:10.3791/56603 (2018).

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