**Abstract**

Sodium tetra (n-propyl) borate was used as derivatizing agent to measure methyl and ethylmercury compounds. This study investigated the artifact formation of methyl and ethylmercury compounds during derivatization using NaBPr4, simultaneously with the influence of this artifact on methylmercury analysis in biological samples (chlor alkali hair samples). The artifact methylmercury and ethylmercury compounds during derivatization using NaBPr4 were evident and depended strongly on the amount of inorganic mercury (Hg2+) present in the sample solution for derivatization and depended on the purity of sodium tetra (n-propyl) borate reagent. The high formation rate of artifact Et-Hg (0.76–0.81% of high-level Hg2+ present) interferes strongly with the ethylmercury analysis. The rate of artifact formation of Me-Hg is small and constant at the different concentration ranges of In-Hg (0.012% of In-Hg present) and does not affect on Me-Hg analysis and it can be subtracted from this Me-Hg artifact ratio from the measured value of Me-Hg in the biological samples. However, the mathematical correction for Me-Hg measurement can be done only when the Et-Hg peak is already appearing in the chromatogram samples.

**Keywords:** inorganic mercury, monomethyl ethylmercury, sodium tetra (n-propyl) borate, artifacts derivatization, mercury compounds

### **1. Introduction**

Mercury has been well known as an environmental toxin and pollutant for several decades. The environmental cycling of mercury is a very complex distribution, involving a large variety of physical and chemical processes that affect its toxicity and mobility [1–4]. The lengthy mercury transport cycle in the atmosphere, it is deposition, bioaccumulation, and the concentration of extremely hazardous methylmerury

methylmercury (Me-Hg) molecules in the aquatic food chain represent a severe environmental concern, even in distant places, poisoning people [3–7].

Analytical techniques for the separation of methylmercury (Me-Hg) are well documented [8]. After extraction from solid matrices and derivatization, the methylmercury (Me-Hg) is frequently measured using hyphenated techniques [8]. Mercury speciation analysis is usually performed by resorting to hyphenated techniques, based on the coupling of an effective separation technique to a sensitive element-specific detector. Capillary gas chromatography (CGC), liquid chromatography (LC), or more recently capillary electrophoresis (EC) can be interfaced with specific atomic detection including atomic absorption spectrometry (AAS), atomic fluorescence spectrometry (AFS), electron capture, or inductively coupled plasma mass spectrometry (ICPMS) [9–17]. Recently research has shown that the coupling of GC to ICP-MS appears to be a more suitable hyphenated technique to carry out the mercury speciation analysis because of its high sensitivity, multi-isotopic, and multi-elemental capabilities [8, 9, 15]. However, the GC-ICP-MS is only suitable for volatile species like mercury species [11, 15]. The isotope dilution ICP-MS is a new powerful approach and offers great potential for very small uncertainties since quantitative recoveries are not required and rearrangement reactions are easily detected [10, 17–21]. The main advantage of this technique (IDMS) is that chemical separation if required for accurate ratio determination need not be quantitative. Moreover, concentrations of chemical species can be measured very precisely because ratios can be measured very reproducibly [8, 11, 16].

Quality results are sometimes associated with sample pre-treatment; the analysis of solids such as biological and environmental samples requires leaching (alkaline or acid)/digestion step to liberate mercury species from the sample matrix before detection with GC-ICP-MS. However, for ionic mercury species, derivatization reactions are required to achieve good results [11, 22].

In earlier studies, monomethyl mercury (Me-Hg) was the most investigated organomercury compound, and measurement of monomethylmercury (Me-Hg) in environmental samples using sodium tetraethylborate (NaBEt4) was one of the most used methods for methylmerury analysis [23, 24]. However, in some cases during the ethylation (Eth) with sodium tetraethyl borate (NaBEt4), the Hg2+ is transformed to HgEt2, while MeHg forms MeHgEt Eqs. (1) and (2).

$$\text{Hg}^{2+} + 2\text{NaB} \left( \text{C}\_2\text{H}\_5 \right)\_4 \rightarrow \text{Hg} \left( \text{C}\_2\text{H}\_5 \right)\_2 + 2\text{Na}^+ + 2\text{B} (\text{C}\_2\text{H}\_5)\_3 \tag{1}$$

$$\text{CH}\_3\text{Hg}^+ + \text{NaB} \left( \text{C}\_2\text{H}\_5 \right)\_4 \rightarrow \text{CH}\_2\text{HgC}\_2\text{H}\_5 + \text{Na}^+ + \text{B(C}\_2\text{H}\_5)\_3 \tag{2}$$

As mentioned above, isotope dilution ICP-MS is a new powerful approach to solving the problems with the matrix and non-quantitative derivatization. A drawback of the ethylation (Eth) procedure is the impossibility to distinguish between Hg<sup>2</sup> <sup>+</sup> and EtHg<sup>+</sup> , both species that often coexist in the environment [25]. It was observed that derivatization using ethylation reagent (NaBEt4) induced the formation of MeHg from inorganic mercury (InHg) if inorganic mercury was present at high concentrations and also the presence of dissolved organic matrix in the sample strongly interferes with ethylation process [18, 26, 27]. Therefore, ignoring this effect of artifact formation may lead to systematic errors in methylmercury analysis. Recently, an alternative is the use of the propylation as a derivatization technique with sodium tetra-propyl borate as the derivatizing agent which is more tolerant to interferences from chlorides [11, 18, 26, 28]. However, it was found that the artifact of methylmercury (Me-Hg) and ethylmercury (Et-Hg) compounds during NaBPr4 derivatization

*A Study on the Methyl and Ethylmercury Artifacts in Biological Samples Using Sodium… DOI: http://dx.doi.org/10.5772/intechopen.110050*

was evident and depended strongly on the concentration of inorganic mercury (Hg+2) presence in the solution for derivatization. For example, Jen-How Huang [26] observed a transformation of In-Hg into ethylmercury (Et-Hg) and methylmerury (Me-Hg) during derivatization using NaBPr4, and he reported that the artifact formation rates of EtHg and MeHg are 0.99–2.9% and 0.03–0.28%, respectively. This conclusion may ignore the artifact formation of monomethylmercury (Me-Hg) and monoethylmercury (Et-Hg) during derivatization by NaBPr4 similar to NaBEt4. Therefore, without taking this effect of artifact formation into account, the artifact may lead to an overestimation of organomercury species concentrations and a false impression of organomercury speciation.

This study aims to investigate the formation of Me-Hg and Et-Hg artifacts in hair samples with the high level of In-Hg in hair workers of ICL factory in Pakistan by comparing the Hg artifacts in un-spiked and spiked blank samples, different concentrations of normal abundance In-Hg solution, enriched 199In-Hg solution and hair sample (normal hair) with low level (0.98 mg/kg) of In-Hg during the derivatization step.

The objectives are [1] to examine the artifact formation of methyl and ethylmercury from inorganic mercury (Hg2+) during propylation using NaBPr4, [2] to identify the factors which govern the artifact formation of MeHg and EtHg, and [3] to evaluate the influence of MeHg and EtHg artifact information on the determination of actual monomethylmercury (Me-Hg) concentrations in chlor alkali hair samples with high inorganic mercury concentrations (Up to 0.9%).

### **2. Materials and methods**

### **2.1 Devices and instrument**

Microwave digestion oven model MARS-5 from CEM Instrument, UK, was used for digestion and decomposition of hair samples. The microwave operating conditions are listed in **Table 1**. A gas chromatograph (GC) model HP 6850 outfitted with a capillary column was connected to an Agilent model HP-7500 ICP mass spectrometer through a heated steel transfer capillary for speciated isotope dilution analysis (SIDMS). The heated steel transfer capillary was inserted into the ICP torch injector, and connection to the torch was realized through a glass T-piece. A conventional Meinhard concentric nebulizer and low volume water-cooled cyclonic spray chamber


### **Table 1.**

*Microwave operating conditions for hair samples digestion.*

were connected to the heated steel transfer capillary line connected ICP torch, and this enabled continuous aspiration of a standard thallium solution (25μgl<sup>1</sup> ). This configuration allowed optimization of instrument performance and simultaneous measurement of 203Tl and 205Tl for mass bias correction during the chromatographic run [9]. Operating conditions for the GC-ICP-MS coupling system are listed in **Table 2**.
