**3. Organomercury, organotin, and organolead detection from complex environmental and biota samples: local case study**

Considering the extensive use from past and their improper disposal, as well the lack of evaluation of possible contaminated sites (from past activities) made that even in our days many of such sites to still being used either for agriculture or for pasture. Without a proper evaluation of contaminants, such as mercury, lead, or tin distribution and speciation in soils, and without an assessment of their risks to animals and humans, exposure to such contaminants could be occurred nowadays. To assess such risks for environment, biota, and public health protection purposes, it is imperative to consider their speciation both in soils, water, and in biota plant.

### **3.1. Environmental and biota sampling for organomercury, organotin, and organolead monitoring**

Soil, water, and vegetable samples were collected from Turda region, Cluj district from the northwestern part of Transylvania (46°34′ and 23°47′E) including Turda town, nearest rural regions and industrial zones—banned chemical factory, Romania (see **Figure 1**). Soil, water, Concerning Organometallic Compounds in Environment: Occurrence, Fate, and Impact http://dx.doi.org/10.5772/67755 51

those to inorganic forms of lead [31]. According to Gallert and Winter [30] and Pyrzynska [32], the toxicity of alkyl lead compounds decreases with a decreasing number of ethyl or methyl moieties or with a decreasing number of carbon atoms (ethyl lead → methyl lead) according to

Organotin compounds are organometallic compounds in which carbons are bonded directly to

forms of tin have been used as active agents in a wide range of applications in industry, such as stabilizers in the polyvinyl chloride industry, plastic additives production, industrial catalysts, antifouling paints, wood preservatives, and in agriculture as biocide products (insecti-

Nowadays, use of organotin compounds as anti-foulant has been banned due to their severe toxic effects on the aquatic organisms [36]. Moreover, use of tributyl tin and triphenyl tin compounds in various industrial applications has raised a great concern in the last decades owing to their serious toxic effects on nontarget organisms when leached into environment even at very low concentrations (ng·L−1) [37]. Besides the fact that they are considered as endocrine disruptors among organometallic compounds, they also possess teratogenic properties and can cause disruption to the reproductive function in mammals, as well as could act as hepa-

In the following sections of this chapter, we will present quantitative and qualitative data about the presence of these compounds in different environmental compartments and biota samples.

**3. Organomercury, organotin, and organolead detection from complex** 

Considering the extensive use from past and their improper disposal, as well the lack of evaluation of possible contaminated sites (from past activities) made that even in our days many of such sites to still being used either for agriculture or for pasture. Without a proper evaluation of contaminants, such as mercury, lead, or tin distribution and speciation in soils, and without an assessment of their risks to animals and humans, exposure to such contaminants could be occurred nowadays. To assess such risks for environment, biota, and public health protection purposes, it is imperative to consider their speciation both in soils, water, and in biota plant.

**3.1. Environmental and biota sampling for organomercury, organotin, and organolead** 

Soil, water, and vegetable samples were collected from Turda region, Cluj district from the northwestern part of Transylvania (46°34′ and 23°47′E) including Turda town, nearest rural regions and industrial zones—banned chemical factory, Romania (see **Figure 1**). Soil, water,

Pb2+, with R being either –CH<sup>3</sup>

, where *n* is between 1 and 4, and R is an alkyl or aryl group) [33]. Organometallic

 or –C2 H5 [27].

the following sequence: R<sup>4</sup>

50 Recent Progress in Organometallic Chemistry

tin (R*<sup>n</sup>*

**monitoring**

SnX4-*<sup>n</sup>*

**2.3. Tin and its organic derivatives**

cides, fungicides, and bactericides) [33–35].

Pb > R3

toxins, immunotoxins, neurotoxins, and obesogens [35, 38].

**environmental and biota samples: local case study**

Pb+ > R2

**Figure 1.** Sampling site map—rectangle corresponds to industrial sites; oval corresponds to inhabited areas; rhomb corresponds to agricultural sites.

and vegetable samples were collected in a period of March, July, and October for 2 years consecutively. Vegetables included for study were selected based on their edible part contact with different environmental compartments: leafy vegetables (lettuce, spinach, and cabbage), "root" and "bulb" vegetables (carrot, parsnip, onion, and garlic), and fruit vegetables (peas, tomato, and eggplant).

Soil and vegetable sample were collected with metallic collectors, returned to the laboratory in polyethylene bags and stored at −20°C. Before analysis, the samples were spread and dried at ambient temperature, and after drying samples were homogenized and shifted through a 2-mm stainless steel sieve.

Surface and well water were collected in polyethylene bottles excepting the cases when organic mercury species were the target analytes, the case when the samples were collected in Teflon containers in order to avoid metallic compound reaction with the bottle surface. Before all sampling campaign, the sampling bottles were subjected to acid cleaning with HNO<sup>3</sup> in order to remove possible metal impurities from the bottle's wall and to prevent further metal adsorption [39]. All water samples were stored in dark at 4°C until analysis and analyzed in less than 7 days from sampling time.

#### **3.2. Organomercury compounds analysis from soil, water, and vegetable samples**

As previous work had shown that the organic species of mercury that were found to be important from hazard and toxicological point of view and those are prevalent in the environment are as follows methylmercury (CH<sup>3</sup> Hg<sup>+</sup> ), ethylmercury (C<sup>2</sup> H5 Hg<sup>+</sup> ) [23, 24], phenylmercury (C<sup>6</sup> H5 Hg<sup>+</sup> ) [25], and dimethylmercury ((CH<sup>3</sup> ) 2 Hg) [26].

#### *3.2.1. Water analysis*

According to Cai et al. [40] and with minor modifications, extraction and derivatization of organic forms of mercury were conducted using 20 mL of water sample that was placed in 40 mL amber glass vials sealed with screw caps with polytetrafluoroethylene (PTFE)-coated silicon rubber septum. Noted that 2 g of NaCl with 150 µL of 0.4% NaBEt<sup>2</sup> was added to the sample and the pH was set to 4.5 using acid acetic, and then the vials were immediately closed tightly. The derivatization step was acquired during 15 min at 70°C.

Afterward, a 50 µm/30 µm divinylbenzene/carboxen/polydimethylsiloxane (DVB/CAR/ PDMS) fiber was exposed to the solution headspace for 20 min, maintaining the same temperature (40°C) and assuring a continuous agitation with a rate of 175 rpm. Finally, the fiber was introduced in the chromatographic injector and the target compounds were thermally desorbed at 260°C for 5 min.

#### *3.2.2. Soil and vegetal sample analysis*

According to the method presented by Korbas et al. [41], 5 g of homogenized soil and their respective dried vegetable samples were put in an extraction tube with 1 mL of aqueous H<sup>2</sup> SO4 (14 M, saturated with cupric sulfate), 1 mL of 4 M KBr solution, and 20 mL of toluene. The mixture with samples was shacked for 30 min after that subjected to centrifugation for 15 min at 2000 rpm. The procedure was repeated once under the same condition, after that the collected supernatant organic phases were combined and back extracted with 20 mL of L-cysteine solution (1.5% w/v). The organic phase was separated once again after shaking and centrifugation process (2000 rpm for 15 min). From the obtained organic layer, water was removed using anhydrous Na<sup>2</sup> SO4 and from that 1 µL was injected into gas chromatograph inlet.

#### *3.2.3. Gas chromatography-mass spectrometric analysis*

Gas chromatography-mass spectrometric analysis was carried out on a GC Focus DSQ II equipment (Thermo Finnigan) using a TR-5MS capillary column with the following characteristics: 30 m × 0.32 mm i.d., with a 0.25-µm film thickness. The mass spectrometer was operated in an electron impact ionization mode at 70 eV ionizing energy. The GC injection port temperature was set at 280°C while the detector source temperature was set at 250°C. Splitless mode was used for injection of 1 µL volume of extracts. Applied temperature program for column oven started from 80°C (3 min) to 150°C·min−1 with a rate of 5°C·min−1 and maintained at 150°C for 5 min followed by an increase of 10°C·min−1 until 280°C and maintained at this final temperature for 5 min. Identification of the target compounds was done through full scan monitoring mode ranging between 50 and 600 m/z.

## **3.3. Organolead compounds analysis from soil, water, and vegetable samples**

Organolead compounds are found in major environmental compartments not only as a consequence of their use in anthropogenic activity, but also via naturally as a consequence of biomethylation processes. As mentioned earlier, the toxicity of these groups of compounds was widely demonstrated, it is known that tetraethyllead (TEL) is much more toxic to animals [16] while ionic allkylead compound was found to be more toxic to plants [42], with both showing higher toxicity than inorganic lead, mainly due to their liposolubility [43]. Generally, it is accepted that the toxicity of organolead compounds increases with the degree of alkylation, respecting the following sequence tetraethyllead > triethyllead > diethyllead > monoethyllead [30].

Target organolead compounds of this study were tetraethyllead (TEL), followed by its transformation products in environment, as triethyllead (TREL), diethyllead (DEL), and monoethyllead (MEL) resulted from dealkylation reactions having as standard their chlorinated forms.

#### *3.3.1. Water analysis*

**3.2. Organomercury compounds analysis from soil, water, and vegetable samples**

Hg<sup>+</sup>

silicon rubber septum. Noted that 2 g of NaCl with 150 µL of 0.4% NaBEt<sup>2</sup>

tightly. The derivatization step was acquired during 15 min at 70°C.

are as follows methylmercury (CH<sup>3</sup>

52 Recent Progress in Organometallic Chemistry

) [25], and dimethylmercury ((CH<sup>3</sup>

(C<sup>6</sup> H5 Hg<sup>+</sup>

*3.2.1. Water analysis*

desorbed at 260°C for 5 min.

anhydrous Na<sup>2</sup>

*3.2.2. Soil and vegetal sample analysis*

SO4

*3.2.3. Gas chromatography-mass spectrometric analysis*

monitoring mode ranging between 50 and 600 m/z.

As previous work had shown that the organic species of mercury that were found to be important from hazard and toxicological point of view and those are prevalent in the environment

According to Cai et al. [40] and with minor modifications, extraction and derivatization of organic forms of mercury were conducted using 20 mL of water sample that was placed in 40 mL amber glass vials sealed with screw caps with polytetrafluoroethylene (PTFE)-coated

sample and the pH was set to 4.5 using acid acetic, and then the vials were immediately closed

Afterward, a 50 µm/30 µm divinylbenzene/carboxen/polydimethylsiloxane (DVB/CAR/ PDMS) fiber was exposed to the solution headspace for 20 min, maintaining the same temperature (40°C) and assuring a continuous agitation with a rate of 175 rpm. Finally, the fiber was introduced in the chromatographic injector and the target compounds were thermally

According to the method presented by Korbas et al. [41], 5 g of homogenized soil and their respective dried vegetable samples were put in an extraction tube with 1 mL of aqueous H<sup>2</sup>

(14 M, saturated with cupric sulfate), 1 mL of 4 M KBr solution, and 20 mL of toluene. The mixture with samples was shacked for 30 min after that subjected to centrifugation for 15 min at 2000 rpm. The procedure was repeated once under the same condition, after that the collected supernatant organic phases were combined and back extracted with 20 mL of L-cysteine solution (1.5% w/v). The organic phase was separated once again after shaking and centrifugation process (2000 rpm for 15 min). From the obtained organic layer, water was removed using

and from that 1 µL was injected into gas chromatograph inlet.

Gas chromatography-mass spectrometric analysis was carried out on a GC Focus DSQ II equipment (Thermo Finnigan) using a TR-5MS capillary column with the following characteristics: 30 m × 0.32 mm i.d., with a 0.25-µm film thickness. The mass spectrometer was operated in an electron impact ionization mode at 70 eV ionizing energy. The GC injection port temperature was set at 280°C while the detector source temperature was set at 250°C. Splitless mode was used for injection of 1 µL volume of extracts. Applied temperature program for column oven started from 80°C (3 min) to 150°C·min−1 with a rate of 5°C·min−1 and maintained at 150°C for 5 min followed by an increase of 10°C·min−1 until 280°C and maintained at this final temperature for 5 min. Identification of the target compounds was done through full scan

) 2

), ethylmercury (C<sup>2</sup>

Hg) [26].

H5 Hg<sup>+</sup>

) [23, 24], phenylmercury

was added to the

SO4

The extraction and derivatization of organic forms of lead is similar to the extraction of organotin species. Shortly, 10 mL of water sample was placed in 40 mL amber glass vials sealed with screw caps with PTFE-coated silicon rubber septum. Noted that 2 g of NaCl with 500 µL of 0.4% NaBEt<sup>2</sup> was added to sample and the pH was set at 4.5 using acid acetic, after that the vials were immediately tightly closed. The derivatization step was acquired during 100 min at 40°C.

Afterward, 100-µm polydimethylsiloxane (PDMS) fiber was exposed to the solution headspace for 20 min, maintaining the same temperature (40°C) and ensuring a continuous agitation with a rate of 175 rpm. Finally, the fiber was introduced in the chromatographic injector and the target compounds were thermally desorbed at 260°C for 5 min.

#### *3.3.2. Soil and vegetable sample analysis*

The extraction of organolead compounds from soil and vegetable samples was acquired ultrasound assisted for 20 min using 10 g of samples and 50 mL of n-hexane. The supernatant was collected and the extraction was repeated once again under the same conditions. Collected supernatants were rotary evaporated until 1 mL. The concentrate was mixed with 300 µL of NaBET<sup>4</sup> (2 g NaBET4 in 10 mL ethanol), used as a derivatization agent for detection and quantification of the target organic lead compounds. The obtained extract mix was subjected to gas chromatography-mass spectrometric (GC-MS) analysis.

#### *3.3.3. Gas chromatography-mass spectrometric analysis*

This was carried out on a GC Focus DSQ II equipment (Thermo Finnigan) using a TR-5MS capillary column with the following characteristics: 30 m × 0.25 mm i.d. with a 0.25-µm film thickness. The mass spectrometer was operated in an electron impact ionization mode at 70 eV ionizing energy. The GC injector was set at 260°C while the detector source temperature was set at 280°C. Splitless mode was used for injection of 1 µL volume of extracts. Applied temperature program for column oven was started from 50°C (5 min) to 100°C·min−1 with a rate of 7°C·min−1 and maintained at 100°C for 2 min followed then by an increase of 15°C·min−1 until 280°C and maintained at this final temperature for 10 min also. The identification of target compounds was done through a full-scan monitoring mode between the range of 50 and 600 m/z.

#### **3.4. Organotin compounds analysis from soil, water, and vegetable samples**

In this work, a field study was conducted investigating the pathways of organotins in soilwater environment and their uptake potential in vegetables grown on possible contaminated areas.

Monitored organotin compounds were as follows: monobutyltin trichloride (MBT), monophenyltin trichloride (MPT), diphenyltin dichloride (DPT), dibutyltin dichloride (DBT), tributyltin chloride (TBT), and triphenyltin chloride (TPT).

#### *3.4.1. Water samples analysis*

Water samples preparation for analysis were done as presented by Kovacs et al. [39] and Hu et al. [44] with minor modifications as follows: about 20 mL of water sample was placed in 40 mL amber glass vials sealed with screw caps with PTFE-coated silicon rubber septum. Noted that 1.5 g of NaCl, 2 mL of acetate buffer (acetate buffer: 82 g·L−1 sodium acetate in water adjusted to pH 5 with acetic acid), and 100 µL of NaBEt<sup>4</sup> solution (2 g NaBEt<sup>4</sup> in 10 mL ethanol) had been added to water sample and the vials were immediately closed and stirred on a magnetic stirrer at 100 rpm and 20°C. The optimal SPME fiber for organotin compounds extraction was found to be 100 µm polydimethylsiloxane (PDMS)-coated fiber. SPME was performed in the headspace. Use of NaBEt<sup>4</sup> solution allowed the organotin compounds to be derivatized *in-situ* and simultaneously extracted into the PDMS phase. The fiber was exposed to the headspace for 30 min under continuous stirring, after that the fiber was withdrawn into the needle of the holder and the SPME was placed into the GC injector at 240°C for 10 min in order to allow target compound desorption.

#### *3.4.2. Soil and vegetal sample analysis*

The extraction of organotin compounds from soil and vegetable samples was done according to the method presented by Hu et al. [44] with minor modifications as follows: 5 g of soil and vegetables, respectively, were extracted with 50 mL methanol that contains 2 mL of hydrochloric acid (37%) by ultrasonic shaking at 50°C for 30 min. The extraction was repeated twice with 30 mL methanol containing 1.2 mL hydrochloric acid (37%). The supernatants were combined and transferred to a separation funnel containing 100 mL of 30% (w/v) sodium chloride salt and 50 mL of dichloromethane and shacked manually for 10 min. The extraction procedure was repeated in the same manner, after that the collected organic layers were united and subjected for concentration through a rotary evaporator almost until to dryness. The concentrate was mixed with a pH 5.0 buffer solution (acetate buffer: 82 g·L−1 sodium acetate in water adjusted to pH 5 with acetic acid) and 120 °L of NaBET<sup>4</sup> (2 g NaBET4 in 10 mL ethanol) was used as a derivatization agent for the target organic tin compounds detection and quantification. The ethylated organotin compounds were extracted with 5 mL of hexane three times after that the extracts were cleaned up using florisil column. The collected organic fraction was concentrated at 1 mL after that the obtained extracts were subjected to gas chromatography-mass spectrometric (GC-MS) analysis.

#### *3.4.3. Gas chromatography-mass spectrometric analysis*

rate of 7°C·min−1 and maintained at 100°C for 2 min followed then by an increase of 15°C·min−1 until 280°C and maintained at this final temperature for 10 min also. The identification of target compounds was done through a full-scan monitoring mode between the range of 50

In this work, a field study was conducted investigating the pathways of organotins in soilwater environment and their uptake potential in vegetables grown on possible contaminated

Monitored organotin compounds were as follows: monobutyltin trichloride (MBT), monophenyltin trichloride (MPT), diphenyltin dichloride (DPT), dibutyltin dichloride (DBT), tribu-

Water samples preparation for analysis were done as presented by Kovacs et al. [39] and Hu et al. [44] with minor modifications as follows: about 20 mL of water sample was placed in 40 mL amber glass vials sealed with screw caps with PTFE-coated silicon rubber septum. Noted that 1.5 g of NaCl, 2 mL of acetate buffer (acetate buffer: 82 g·L−1 sodium acetate in water adjusted

been added to water sample and the vials were immediately closed and stirred on a magnetic stirrer at 100 rpm and 20°C. The optimal SPME fiber for organotin compounds extraction was found to be 100 µm polydimethylsiloxane (PDMS)-coated fiber. SPME was performed in the

and simultaneously extracted into the PDMS phase. The fiber was exposed to the headspace for 30 min under continuous stirring, after that the fiber was withdrawn into the needle of the holder and the SPME was placed into the GC injector at 240°C for 10 min in order to allow

The extraction of organotin compounds from soil and vegetable samples was done according to the method presented by Hu et al. [44] with minor modifications as follows: 5 g of soil and vegetables, respectively, were extracted with 50 mL methanol that contains 2 mL of hydrochloric acid (37%) by ultrasonic shaking at 50°C for 30 min. The extraction was repeated twice with 30 mL methanol containing 1.2 mL hydrochloric acid (37%). The supernatants were combined and transferred to a separation funnel containing 100 mL of 30% (w/v) sodium chloride salt and 50 mL of dichloromethane and shacked manually for 10 min. The extraction procedure was repeated in the same manner, after that the collected organic layers were united and subjected for concentration through a rotary evaporator almost until to dryness. The concentrate was mixed with a pH 5.0 buffer solution (acetate buffer: 82 g·L−1 sodium acetate in water adjusted

derivatization agent for the target organic tin compounds detection and quantification. The

solution (2 g NaBEt<sup>4</sup>

solution allowed the organotin compounds to be derivatized *in-situ*

(2 g NaBET4 in 10 mL ethanol) was used as a

in 10 mL ethanol) had

**3.4. Organotin compounds analysis from soil, water, and vegetable samples**

tyltin chloride (TBT), and triphenyltin chloride (TPT).

to pH 5 with acetic acid), and 100 µL of NaBEt<sup>4</sup>

and 600 m/z.

54 Recent Progress in Organometallic Chemistry

areas.

*3.4.1. Water samples analysis*

headspace. Use of NaBEt<sup>4</sup>

target compound desorption.

*3.4.2. Soil and vegetal sample analysis*

to pH 5 with acetic acid) and 120 °L of NaBET<sup>4</sup>

It was carried out on a GC Focus DSQ II equipment (Thermo Finnigan) using a TR-5MS capillary column with the following characteristics: 60 m × 0.25 mm i.d. with a 0.25-µm film thickness. The mass spectrometer was operated in an electron impact ionization mode at 70 eV ionizing energy. The GC injector was set at 270°C while the detector source temperature was set at 280°C. Splitless mode was used for injection of 1 µL volume of extracts. Applied temperature program for column oven was started from 50°C (5 min) to 130°C·min−1 with a rate of 15°C·min−1 and maintained at 130°C for 10 min followed then by an increase of 15°C·min−1 until 280°C and maintained at this final temperature for 10 min.

Identification of target compounds was done through a selective ion-monitoring mode, and the fragment ions were those most abundant in each oligomers. Their values are presented in **Table 1**.


**Table 1.** Monitored ions in selective ion monitoring mode (SIM) of target organometallic compounds.
