- Email: [email protected]
Development of the extraction method for the simultaneous determination of butyl-, phenyl- and octyltin compounds in sewage sludge
Talanta 80 (2010) 1945–1951
Contents lists available at ScienceDirect
Talanta journal homepage: www.elsevier.com/locate/talanta
Development of the extraction method for the simultaneous determination of butyl-, phenyl- and octyltin compounds in sewage sludge ˇ canˇcar a Tea Zuliani a,b,∗ , Gaetane Lespes b , Radmila Milaˇciˇc a , Janez Sˇ a b
Department of Environmental Sciences, Joˇzef Stefan Institute, Jamova 39, 1000 Ljubljana, Slovenia Laboratoire de Chimie Analytique, UMR 5254, Université de Pau, BP 1155, F-64013 PAU Cedex, France
a r t i c l e
i n f o
Article history: Received 6 July 2009 Received in revised form 9 October 2009 Accepted 23 October 2009 Available online 10 November 2009 Keywords: Organotin compounds Sewage sludge Extraction procedures GC–MS
a b s t r a c t The toxicity and bioaccumulation of organotin compounds (OTCs) led to the development of sensitive and selective analytical methods for their determination. In the past much attention was assigned to the study of OTCs in biological samples, water and sediments, coming mostly from marine environment. Little information about OTCs pollution of terrestrial ecosystems is available. In order to optimise the extraction method for simultaneous determination of butyl-, phenyl- and octyltin compounds in sewage sludge five different extractants (tetramethylammonium hydroxide, HCl in methanol, glacial acetic acid, mixture of acetic acid and methanol (3:1), and mixture of acetic acid, methanol and water (1:1:1)), the presence or not of a complexing agent (tropolone), and the use of different modes of extraction (mechanical stirring, microwave and ultrasonic assisted extraction) were tested. Extracted OTCs were derivatised with sodium tetraethylborate and determined by gas chromatography coupled with mass spectrometer. Quantitative extraction of butyl-, phenyl- and octyltin compounds was obtained by the use of glacial acetic acid as extractant and mechanical stirring for 16 h or sonication for 30 min. The limits of detection and quantification for OTCs investigated in sewage sludge were in the ng Sn g−1 range. © 2009 Elsevier B.V. All rights reserved.
1. Introduction Organotin compounds (OTCs) are organometallic derivatives of tin that have the highest number of commercial applications than any other element [1]. Mostly they have been used as stabilizers and catalysts in the production of poly(vinyl chloride) (PVC), polyurethanes and silicones, as antifouling paints for ship hulls, and as agricultural and wood preservative fungicides [2–4]. Due to their widespread use, OTCs may be found in different ecosystems. Today, their high toxicity for aquatic and terrestrial organisms is well documented [3,4]. Moreover, because of their toxicity and bioaccumulation potential, OTCs have been registered as priority pollutants by the European Union in the Pollutant Emission Register (2000/479/EC) and in the Water Framework Directive (2000/60/EC). During last decades, a variety of analytical methods for the speciation of OTCs has been developed. Analytical procedures for OTCs speciation generally attempt to preserve only the organic radical during extraction, whereas the counterion and other tin heteroatomic bonds are cleaved during extraction or derivatisa-
∗ Corresponding author at: Department of Environmental Sciences, Joˇzef Stefan Institute, Jamova 39, 1000 Ljubljana, Slovenia. Tel.: +386 1 4773900; fax: +386 1 2519385. E-mail address: [email protected] (T. Zuliani). 0039-9140/$ – see front matter © 2009 Elsevier B.V. All rights reserved. doi:10.1016/j.talanta.2009.10.050
tion [5]. Sample preparation techniques for speciation analyses of OTCs in solid samples consist of several steps: solid–liquid extraction, simultaneous derivatisation and extraction into an organic solvent, separation and detection. One of the most difficult parts of the analysis of OTCs in sewage sludge is their extraction from the complex matrix. Extraction of OTCs from solid matrices keeping their speciation preserved is not straightforward because the binding forces between OTCs and matrices are species and matrices dependant [6,7]. In addition, some OTCs are not stable during extraction. Species transformation can be considered to be one of the most serious problems during phenyltin speciation analysis [8–10]. From the discovery of the toxicity of OTCs towards numerous marine species in late 1970s, most efforts have been devoted to the development of analytical methods for their determination in marine environments (seawater, sediments and biota) [5,11,12]. There are few studies concerning tin speciation in samples from terrestrial ecosystem [10,13,14]. OTCs are entering the terrestrial environment via the leaching from products made of PVC and silicone, by the use of pesticides and the amendment of cultivated soil with contaminated sewage sludge [15,16]. OTCs were also found in leachates from landfills [17,18]. Sewage sludge is an important source of OTCs in the terrestrial ecosystems [15,16]. Thus, reliable analytical methods are needed in order to evaluate the contribution of contaminated sewage sludge to the OTCs pollution of terrestrial environment. The aim
1946
T. Zuliani et al. / Talanta 80 (2010) 1945–1951
Table 1 Mass/charge (m/z) ratios and retention times for studied OTCs. OTCs
Retention time (min)
m/z
MBT DBT TBT MPhT DPhT TPhT MOcT DOcT TOcT TPrT
6.7 8.9 10.8 10.1 15.0 17.9 11.9 16.6 19.9 7.6
235, 179, 149 263, 179, 151 291, 263, 179 255, 197 303, 275, 197 351, 197 291, 179, 151 375, 263, 151 375, 263, 151 249, 235, 193
Ions used for quantification are reported in bold.
of our investigation was to set up an analytical procedure for the speciation of OTCs in sewage sludge by gas chromatography mass spectrometry (GC–MS). The influence of the extractant, extraction technique and the presence of complexing agent on the extraction efficiency of butyl-, phenyl- and octyltin compounds from sewage sludge have been studied. The selected methods encompass five different extractants (tetramethylammonium hydroxide (TMAH), hydrochloric acid (HCl) in methanol, glacial acetic acid, mixture of acetic acid and methanol (3:1), and mixture of acetic acid, methanol and water (1:1:1)), the presence or not of a complexing agent (tropolone), and the use of different modes of extraction (mechanical stirring, microwave and ultrasonic assisted extraction). Extracted OTCs were derivatised with sodium tetraethylborate (NaBEt4 ) and determined by GC–MS. 2. Experimental 2.1. Instrumentation For the separation and detection a Hewlett-Packard 6890 gas chromatograph (GC) (Hewlett-Packard, Waldbronn, Germany) coupled to a Hewlett-Packard 5972A mass spectrometer (MS) was used. The GC was equipped with a 30 m × 25 mm HP-MS5 polydimethylsiloxane capillary column (film thickness 0.25 m). The following temperature program was applied for the separation of OTCs: the column temperature was held at 80 ◦ C for the first 4 min, raised to 180 ◦ C at the heating rate of 10 ◦ C min−1 , then to 220 ◦ C at 20 ◦ C min−1 , and to 270 ◦ C at 30 ◦ C min−1 (held at this temperature for 6 min). The splitless injector port was kept at a temperature of 240 ◦ C and the transfer line at 280 ◦ C. Helium (He) was used as a carrier gas (1 mL min−1 ). The volume of injected sample was 1 L. The MS operated with the electronic impact (70 eV) ion source at a temperature of 180 ◦ C in the total ionization mode. Selected Ion Monitoring signals (SIM) were recorded. In Table 1 are presented the mass/charge (m/z) ratios of OTCs and their retention times. For the quantification the most abundant ions were used. Mechanical shaking during the extraction procedure was performed on an elliptical table Vibramax 40 (Tehtnica, Zˇ elezniki, Slovenia). Microwave assisted extraction was performed with a CEM MARS 5 Microwave Acceleration Reaction System, CEM Corporation (Matthews, NC, USA). Ultrasonic-assisted extraction of samples was performed with an ultrasonic bath Model 550D, VWR International (West Chester, PA, USA). Centrifugation of sample extracts was performed on a Heraeus (Osterode, Germany) Model 17S Sepatech Biofuge centrifuge and on a LC 320 centrifuge (Tehtnica, Zˇ elezniki, Slovenia). 2.2. Reagents Monobutyltin trichloride (MBT, 95%), monophenyltin trichloride (MPhT, 98%) and diphenyltin dichloride (DPhT, 96%) were
purchased from Aldrich (Milwaukee, WI, USA). Dibutyltin dichloride (DBT, 97%), tributyltin chloride (TBT, 96%), triphenyltin chloride (TPhT, 95%) and tripropyltin chloride (TPrT, 98%) were obtained from Merck (Darmstadt, Germany). Monooctyltin trichloride (MOcT, 98%), dioctyltin dichloride (DOcT, 98%) were obtained from LGC Promochem (Wesel, Germany) and trioctyltin chloride (TOcT, 95%) from Fluka (Buchs, Switzerland). Organotin standard stock solutions (1000 mg Sn L−1 ) were prepared in methanol. Working standard solutions (10 mg Sn L−1 ) were prepared weekly from stock standard solutions by dilution in Milli-Q water (18.2 M cm) (Millipore, Bedford, MA, USA). Standard solutions of 100 g Sn L−1 were prepared daily. All the standards were stored in the dark at 4 ◦ C. Glacial acetic acid, ammonia solution and nitric acid were obtained from J.T. Baker (Phillipsburg, NJ, USA). Methanol was purchased from Merck (Darmstadt, Germany). Sodium tetraethylborate (NaBEt4 ) was purchased from Strem Chemicals (Newburyport, MA, USA). NaBEt4 was dissolved in Milli-Q water to provide a 2% (m/v) ethylating solution. 2.3. Sample preparation Sewage sludge samples were dried at 50 ◦ C in the dark for 3 days, homogenised and sieved through a 0.2 mm sieve. The samples were then frozen at −20 ◦ C. 2.4. Analytical procedure 2.4.1. Extraction from the sample Approximately 1 g of dried sewage sludge was weighted in a polyethylene (PE) extraction tube. Based on our previous experiences 0.2 g of TPrT in methanol was added as internal standard [19]. After 15 min the samples were subjected to different extraction methods. 2.4.1.1. Method 1. 20 mL of tetramethylammonium hydroxide (TMAH) was added to the sample. The tubes were subjected to ultrasound assisted extraction for 2 h and centrifuged for 15 min at 4200 rpm. 2.4.1.2. Method 2. Samples were supplemented with 15 mL of 0.1 mol L−1 HCl in methanol. The closed tubes were firstly mechanically shaken on an elliptic table for 2 h and then subjected to ultrasonic assisted extraction for 1 h and centrifuged for 15 min at 4200 rpm. 2.4.1.3. Method 3. Samples were supplemented with 20 mL of glacial acetic acid. The tubes were subjected to closed vessel microwave assisted extraction for 4, 7 and 16 min. Microwave extraction conditions were as follows: 1200 W, 1 min ramp to 50 ◦ C, hold at that temperature for 3, 6 and 15 min. After the extraction, the suspension was centrifuged for 15 min at 4200 rpm. 2.4.1.4. Method 4. To the solid sample 5 mL of methanol and 15 mL of glacial acetic acid were added. The tubes were subjected to closed vessel microwave assisted extraction for 7 min. The microwave extraction conditions were as described in Section 2.4.1.3. After the extraction, the suspension was centrifuged for 15 min at 4200 rpm. 2.4.1.5. Method 5. Samples were supplemented with 20 mL mixture of acetic acid, methanol and water (1:1:1). The tubes were subjected to closed vessel microwave assisted extraction for 7 min. The microwave extraction conditions were as described in Section 2.4.1.3. After the extraction, the suspension was centrifuged for 15 min at 4200 rpm.
T. Zuliani et al. / Talanta 80 (2010) 1945–1951
1947
2.4.1.6. Method 6. To the solid sample 5 mL of 0.02% (w/v) tropolone in methanol and 15 mL of glacial acetic acid were added. The tubes were subjected to closed vessel microwave assisted extraction for 7 min. The microwave extraction conditions were as described in Section 2.4.1.3. After the extraction, the suspension was centrifuged for 15 min at 4200 rpm. 2.4.1.7. Method 7. Samples were supplemented with 20 mL of glacial acetic acid. The closed tubes were subjected to ultrasound assisted extraction for 5, 10, 20, 30 and 60 min. After the extraction, the suspension was centrifuged for 15 min at 4200 rpm. 2.4.1.8. Method 8. As Method 7. Acetic acid–methanol mixture (3:1) instead of glacial acetic acid as extractant was applied to the samples. 2.4.1.9. Method 9. To the solid sample 20 mL of glacial acetic acid was added. The closed tubes were mechanically shaken on an elliptic table for 1, 5, 15 and 24 h in the dark. Then, the suspension was centrifuged for 15 min at 4200 rpm. 2.4.2. Derivatisation and extraction Aliquots (1–4 mL) of the extract described above, 0.5 mL 2% (w/v) solution of NaBEt4 for the derivatisation and 1 mL of isooctane for the extraction of ethylated organotin species were added to 20 mL of sodium acetate-acetic acid buffer (pH 4.8). The mixture was shaken for 45 min at 300 rpm. The same procedure, with the exception that no sample was added, was applied to determine blanks. All the analyses were made in triplicate. 2.4.3. Spiking protocol The recovery of the studied OTCs from sewage sludge was investigated by spiking known amount of OTCs into samples of relatively low OTCs contents. For spiking of sewage sludge samples OTCs as chlorides dissolved in methanol were used. The equilibration step was carried out by shaking (350 rpm) the samples for 30 min. For each extraction three different spikes were prepared. One aliquot of the sewage sludge was spiked with 1000 ng Sn g−1 of monosubstituted butyl-, phenyl- and octyltin compounds (MBT, MPhT, MOcT). The second one was spiked with 1000 ng Sn g−1 of disubstituted butyl-, phenyl- and octyltin compounds (DBT, DPhT, DOcT) and the third with 1000 ng Sn g−1 of trisubstituted butyl-, phenyl- and octyltin compounds (TBT, TPhT, TOcT). The differences between recoveries depend only on the differences in extraction procedure, as the post-extraction steps of the analytical method remained the same. 3. Results and discussion Sewage sludge consists of a complex matrix. In order to release the molecular and ionic OTCs from organic matter in sewage sludge an appropriate digestion/extraction must be applied. In any speciation procedure, the extraction procedure must be carefully chosen so as to destroy the matrix but not to affect the authentic forms of the analyte molecules [20]. The removal of OTCs in wastewater treatment plants takes place in two steps. First, in the primary treatment mostly by sorption onto particulate matter of the wastewater [21]. Second, in the secondary biological treatment by sorption onto particulate organic sludge material and by assimilation of carbon from OTCs by microorganisms [15]. In bio-tissue the OTCs are strongly bound to the cellular structure [9]. Usually, for the extraction of OTCs from bio-tissues 25% water solution of TMAH or 0.1 mol L−1 HCl in methanol are applied [5,22,23]. Since sewage sludge from wastewater treatment plant is mostly composed of organic matter (dead biomass), those two extractants (Methods 1 and 2) were considered. On the other
Fig. 1. Extraction recoveries of butyl-, phenyl- and octyltin compounds from spiked sewage sludge sample (1000 ng Sn g−1 ) applying the extraction with 25% water solution of TMAH and 0.1 mol L−1 HCl in methanol.
hand, the bonds between OTCs and particulate organic matter are strong. So, extraction solvents that are usually applied to sediment samples were also verified. As glacial acetic acid and mixtures of acetic acid and methanol have been the most widely reported in the literature for the extraction of OTCs from sediments and soil [12,14,24], their performance on sewage sludge was examined (Methods 3–9). All the tested methods (1–9) are precisely described in Section 2. In Table 2 a schematic description of the selected methods is presented. 3.1. Methods 1 and 2 25% water solution of TMAH and 0.1 mol L−1 HCl in methanol are the most frequently applied solvents for the extraction of OTCs from biological samples [5,24]. As sewage sludge is mostly composed by organic matter, the extraction efficiency of those two solvents for the extraction of OTCs from sewage sludge was tested. In Fig. 1 the extraction recoveries of both solvents are presented. When TMAH was applied as extractant, an emulsion disturbing the analytical process during the derivatisation was formed. 25% water solution of TMAH was not efficient for the extraction of butly-, phenyl- and octyltin compounds from sewage sludge. The recoveries for all OTCs were around 10% (Fig. 1). As can be also seen from Fig. 1, when 0.1 mol L−1 HCl in methanol was used, the recoveries for butyltins, octyltins, DPhT and TPhT were between 40 and 60%, while MPhT was not extracted at all. High recovery values for DBT and DOcT, 100 and 105%, respectively, were observed. If OTCs in sewage sludge is bound to the cellular structure of the biomass TMAH would efficiently solubilise the matrix in order to release OTCs (as in fish and mussel tissue). From the results it may be deduced that OTCs present in the sewage sludge are not bound to the cellular structure of the biomass but are sorbed onto the particulate organic matter. Since the binding to particulate organic matter is stronger than that to biological tissue, TMAH was too weak for the extraction of OTCs from the complex matrix of sewage sludge. As highly polar mono- and disubstituted OTCs need acidification to increase the efficiency of extraction from the sample [7], the use of 0.1 mol L−1 HCl in methanol would favour their extraction. The results in Fig. 1 did not confirm that. It can be seen that monosubstituted OTCs were not efficiently extracted most probably due to too low acid concentration. Contrary, 0.1 mol L−1 HCl turned out to be too strong for trisubstituted OTCs provoking their degradation.
1948
T. Zuliani et al. / Talanta 80 (2010) 1945–1951
Table 2 Schematic description of tested extraction methods. Method
Extractant
Complexing agent
Mode of extraction
1 2 3 4 5 6 7 8 9
25% water solution TMAH 0.1 mol L−1 HCl in methanol Glacial acetic acid Glacial acetic acid:methanol (3:1) Glacial acetic acid:methanol:water (1:1:1) Glacial acetic acid:methanol (3:1) Glacial acetic acid Glacial acetic acid:methanol (3:1) Glacial acetic acid
– – – – – Tropolone – – –
Ultrasound assisted extraction Mechanical stirring and ultrasonication Microwave assisted extraction Microwave assisted extraction Microwave assisted extraction Microwave assisted extraction Ultrasound assisted extraction Ultrasound assisted extraction Mechanical stirring
Table 3 Extraction recoveries (%) of butyl-, phenyl- and octyltin compounds for spiked sewage sludge (1000 ng Sn g−1 ) applying MWAE with glacial acetic acid. OTCs
Extraction times 4 min
MBT DBT TBT MPhT DPhT TPhT MOcT DOcT TOcT
99 105 102 81 20 27 86 92 90
± ± ± ± ± ± ± ± ±
7 min 6 6 6 8 3 4 8 9 7
107 105 97 100 10 10 105 107 100
± ± ± ± ± ± ± ± ±
16 min 5 6 4 11 2 2 5 6 4
44 81 78 101 13 11 100 87 98
± ± ± ± ± ± ± ± ±
5 9 7 10 2 2 7 5 6
3.2. Methods from 3 to 6 Microwave assisted extraction (MWAE) was investigated in order to substantially reduce the extraction time for the determination of OTCs in sewage sludge. Extraction times of 4, 7 and 16 min were tested. Glacial acetic acid was used as extractant (Method 3). The results are presented in Table 3. It is evident from Table 3 that butyltin compounds were efficiently extracted with acetic acid after 4 and 7 min of MWAE. At longer extraction time the extraction efficiency of butyltin com-
pounds was appreciably reduced. In order to find out whether the degradation of butyltin compounds took place, an additional experiment was done. Two sewage sludge samples were spiked with DBT and TBT (1000 ng Sn g−1 ), respectively, and subjected to MWAE with glacial acetic acid for 16 min. Results are presented in Fig. 2(A) and (B). Data from Fig. 2(A) and (B) indicate that no degradation processess occured during the course of the experiment. In addition, no increase of the Sn(IV) peak was observed. Therefore, the degradation of MBT or DBT and TBT direct to Sn(IV) may be excluded. From the results, it may be presumed that lower extraction efficiencies of butyltin compounds after 16 min of MWAE are most probably related to more efficient extraction of the organic matter from sewage sludge. The co-extracted organic matter inhibits ethylation/extraction steps, resulting in poor butyltin compounds recoveries. It can be seen from Table 3 that MPhT was efficiently extracted after 7 min. The recoveries for DPhT and TPhT were about 20% after 4 min and below 10% at longer extraction times. To check if the degradation of phenyltin compounds occurred, two aliquots of sewage sludge were spiked with DPhT and TPhT (1000 ng Sn g−1 ), respectively, and subjected to MWAE with glacial acetic acid for 4 min, Fig. 3(A) and (B). From data of Fig. 3, it is obvious that TPhT was almost completely degraded via DPhT to MPhT. Also 90% of DPhT was degraded to MPhT.
Fig. 2. Degradation of (A) TBT and (B) DBT in spiked sewage sludge (1000 ng Sn g−1 of TBT and/or DBT) during MWAE with glacial acetic acid (16 min).
Fig. 3. Degradation of (A) TPhT and (B) DPhT in spiked sewage sludge (1000 ng Sn g−1 of TPhT and/or DPhT) during MWAE with glacial acetic acid (4 min).
T. Zuliani et al. / Talanta 80 (2010) 1945–1951
1949
Fig. 4. Extraction recoveries for spiked sewage sludge with OTCs (1000 ng Sn g−1 ) applying MWAE with glacial acetic acid, acetic acid and methanol (3:1), and acetic acid, methanol and water (1:1:1).
Organometallic degradation during MWAE has been previously reported by Abalos et al. [5] and Huang et al. [10]. The fast temperature increase of extracting solution induces dealkylation and dearylation of analytes [14], especially the most thermolabile, i.e. trisubstituted ones, which is in agreement with the presented data. Octyltin compounds were efficiently extracted after 7 min (Table 3). When subjected to longer extraction times only DOcT exhibits slightly lower extraction efficiency. Since there was no evidence of the degradation of DOcT, the lower extraction efficiency for DOcT after 16 min of MWAE may be related to the influence of the co-extracted organic matter. Based on results from Table 3 and Figs. 2 and 3, the optimal extraction time for butyl- and octyltin compounds applying MWAE and glacial acetic acid as extractant was found to be 7 min. The exceptions were phenyltin compounds, for which MWAE was not applicable. To examine the potential of other extractants, mixtures of acetic acid and methanol (3:1) as well as acetic acid, methanol and water (1:1:1), were subjected to MWAE for 7 min. The comparison between extraction efficiencies of OTCs from spiked sewage sludge samples (1000 ng Sn g−1 ) of the above-mentioned extractants is given in Fig. 4. Namely, the addition of methanol to the glacial acetic acid should enhance the extraction efficiency for TBT [6], while the extraction efficiency of phenyltin and octyltin compounds should not be affected [14,25]. From comparison of results (Fig. 4) it can be seen that, as expected, the extraction recovery of TBT using acetic acid–methanol (3:1) was slightly higher than with glacial acetic acid. For MBT, DBT and octyltin compounds the addition of methanol had no great influence on the extraction efficiency, while the extraction efficiency for phenyltin compounds was even worse. When water was added to glacial acetic acid and methanol quantitative extraction efficiency for TBT and moderate (85%) for DBT was obtained. The addition of water seemed to severely reduce the extraction efficiency of MBT, MPhT and octyltin compounds, most probably due to lower extractant strength. The most beneficial influence of water addition was observed for TPhT, where the extraction recovery raised to 100%. The addition of water and methanol to glacial acetic acid decreased the strength of the acid. This resulted in optimal condition for the extraction of TPhT, while for the efficient extraction of DPhT and MPhT the acid strength was too low. It was experimentally proven that MWAE using 0.02% tropolone as a component of the extractant solution (Method 6) was not appropriate for OTCs analyses. Tropolone enhanced the extraction of organic matter that resulted in significant increase of the back-
ground signal during the chromatographic separation of OTCs (see data from Fig. 5). 3.3. Methods 7 and 8 Ultrasound assisted extraction (USAE) has become a widely used extraction technique. In combination with glacial acetic acid and acetic acid and methanol, high recoveries for sediment and soil samples were obtained [12,14]. For the simultaneous extraction of butyl-, phenyl- and octyltin compounds from sewage sludge sample glacial acetic acid USAE was applied for 5, 10, 20, 30 and 60 min. In Table 4 the obtained extraction efficiencies are presented. It can be observed from Table 4 that the time of sonication has a great influence on the extraction efficiency. MBT and DBT were quantitatively extracted after 30 min, while for TBT 20 min of sonication gave quantitative recoveries. Longer extraction times did not have significant influence on the recovery of butyltin compounds.
Fig. 5. Chromatograms of butyl-, phenyl- and octyltin compounds from spiked sewage sludge sample (1000 ng Sn g−1 ) using MWAE with (A) methanol and acetic acid and (B) 0.02% (w/v) tropolone in methanol and acetic acid.
1950
T. Zuliani et al. / Talanta 80 (2010) 1945–1951
Table 4 Extraction recoveries of butyl-, phenyl- and octyltin compounds for spiked sewage sludge sample (1000 ng Sn g−1 ) applying USAE with glacial acetic acid. OTCs
Extraction time 5 min
MBT DBT TBT MPhT DPhT TPhT MOcT DOcT TOcT
Table 5 Extraction recoveries (%) of butyl-, phenyl- and octyltin compounds from spiked sewage sludge sample (1000 ng Sn g−1 ) applying MSAE with glacial acetic acid.
31 86 89 44 72 90 54 83 89
± ± ± ± ± ± ± ± ±
OTCs
10 min
6 8 7 10 9 10 8 7 9
80 83 99 67 81 81 82 83 100
± ± ± ± ± ± ± ± ±
20 min
7 6 10 12 8 9 7 6 9
63 85 100 63 82 85 80 108 99
± ± ± ± ± ± ± ± ±
30 min
6 9 10 5 7 7 8 10 9
100 97 100 93 92 100 92 99 96
± ± ± ± ± ± ± ± ±
Extraction time
60 min
5 4 7 5 4 7 5 4 5
98 101 99 86 88 97 102 100 100
± ± ± ± ± ± ± ± ±
1h
9 10 10 12 10 15 17 15 16
MBT DBT TBT MPhT DPhT TPhT MOcT DOcT TOcT
67 68 71 59 58 74 72 75 72
± ± ± ± ± ± ± ± ±
5h 7 10 8 6 7 10 7 15 10
74 79 75 67 74 78 84 80 80
± ± ± ± ± ± ± ± ±
16 h 7 1 4 7 6 4 8 2 3
97 92 109 100 98 97 104 97 95
± ± ± ± ± ± ± ± ±
24 h 5 4 5 6 5 6 4 3 5
82 80 75 40 28 62 90 87 85
± ± ± ± ± ± ± ± ±
8 4 3 3 4 6 9 2 3
3.4. Method 9
Fig. 6. Extraction recoveries of butyl-, phenyl- and octyltin compounds from spiked sewage sludge sample (1000 ng Sn g−1 ) applying USAE with acetic acid and methanol (3:1).
Phenyltins are also efficiently extracted after 30 min of USAE. The extraction recoveries for phenyltin compounds remained constant after 30 min of sonication. MOcT showed the same behaviour as phenyltin compounds, while DOcT was efficiently extracted after 20 and TOcT after 10 min. The extending of extraction time did not significantly influence the recoveries of these two octyltin compounds. The studied OTCs in the spiked sewage sludge sample were extracted also with a mixture of acetic acid and methanol (3:1) and subjected to USAE. The results are presented in Fig. 6. As it was the case of MWAE, the mixture of acetic acid and methanol lowered the extraction efficiencies for phenyltin compounds by USAE, while it did not have significant influence on the extraction of butyl- and octyltin compounds.
The most frequently used extraction mode for OTCs determination in solid samples is mechanical stirring (MSAE). For the simultaneous extraction of butyl-, phenyl- and octyltin compounds from sewage sludge sample MSAE using glacial acetic acid as extractant was applied for 1, 5, 16 and 24 h. In Table 5 the obtained extraction efficiencies are presented. As can be seen from Table 5 the extraction time has a significant influence on the extraction efficiency of MSAE. It is evident that butyl-, phenyl- and octyltin compounds are quantitatively extracted after 16 h and no degradation occurred. At longer extraction time (24 h) lower extraction efficiencies of butyl- and octyltin compounds are observed. Additional experiments where sewage sludge sample was spiked either with TBT and TOcT or with DBT and DOcT confirmed that no degradation occurred. Therefore, lower extraction efficiencies are related most probably to re-adsorption on particulate matter and/or highest amount of co-extracted matter from the sample. From Table 5, it is further evident that poor extraction recoveries after 24 h are obtained for phenyltin compounds. In the additional experiment sewage sludge sample was spiked with either TPhT or DPhT. Under these conditions important degradation of TPhT and DPhT occurred. In addition, no increase of the Sn(IV) peak was observed. Therefore, the degradation of MPhT to Sn(IV) may be excluded. Lower extraction recovery for MPhT after 24 h of MSAE with glacial acetic acid may be due to the influence of the coextracted matter from the sample and re-adsorption on particulate matter. Since glacial acetic acid applying MSAE for 16 h quantitatively extracted butyl-, phenyl- and octyltin compounds from sewage sludge other extracting agents (acetic acid and methanol (3:1), and acetic acid, methanol and water (1:1:1)) were not tested.
Table 6 Limits of detection (LOD), quantification (LOQ) and repeatability (as RSD) for MSAE and USAE of butyl-, phenyl- and octyltin compounds in sewage sludge. OTCs
MSAE a
USAE −1
LOD (ng Sn g MBT DBT TBT MPhT DPhT TPhT MOcT DOcT TOcT
27 28 40 42 9 5 19 13 16
)
b
−1
LOQ (ng Sn g 53 54 73 82 18 12 39 25 31
Concentrations are expressed in ng Sn g−1 (dry mass basis). a Limits of detection. b Limits of quantification.
)
RSD (%)
LOD (ng Sn g−1 )
LOQ (ng Sn g−1 )
RSD (%)
8 3 4 5 2 3 3 4 2
24 26 39 43 11 7 17 12 17
49 53 71 83 23 15 35 24 32
6 5 3 5 3 3 9 7 4
T. Zuliani et al. / Talanta 80 (2010) 1945–1951
3.5. Analytical performances of MSAE and USAE Limits of detection (LOD) and quantification (LOQ) for MSAE and USEA were calculated on the basis of the standard addition method calibration curves [26]. The results for LOD, LOQ and repeatability (as relative standard deviation, RSD) for both methods are presented in Table 6. Based on results from Tables 4 and 5 it can be concluded that quantitative extraction of butyltin compounds (MBT, DBT, TBT), phenyltin compounds (MPhT, DPhT, TPhT) and octyltin compounds (MOcT, DOcT, TOcT) is obtained by the use of glacial acetic acid as extractant and MSAE for 16 h or USAE for 30 min. As can be seen from Table 6, LOD and LOQ for the two methods are similar. Therefore, the choice of the extraction mode depends on the availability of the laboratory equipment. The experiment also confirmed that MWAE and extractants as TMAH, 0.1 mol L−1 HCl in methanol, mixtures of acetic acid and methanol (3:1) and acetic acid, methanol and water (1:1:1) were not applicable for the simultaneous determination of butyl-, phenyl- and octyltin compounds in sewage sludge. 4. Conclusions For the extraction of OTCs from sewage sludge five extraction solvents (TMAH, HCl diluted with methanol, glacial acetic acid, acetic acid with methanol (3:1), and acetic acid, methanol and water (1:1:1)), the use of complexing agent (tropolone), and three extraction modes (MSAE, USAE and MWAE) were tested. Regardless of extractants applied, MWAE was not appropriate for simultaneous determination of butyl-, phenyl- and octyltin compounds in sewage sludge. Quantitative extraction of butyl-, phenyl- and octyltin compounds was obtained by the use of glacial acetic acid as extractant and MSAE for 16 h or sonication for 30 min. Both extraction procedures represent reliable analytical tool for the determination of the content of OTCs in sewage sludge. Acknowledgements The authors would like to acknowledge the financial support of the Ministry of Higher Education, Science and Technology
1951
of the Republic of Slovenia within the research programme P10143, French Ministry of Education and PROTEUS bilateral project: “Organotin compounds in the Slovenian and French environment”. References [1] A. Chapman, S.J. Blunden, in: P.J. Craig (Ed.), Organometallic Compounds in the Environment: Principles and Reactions, Longman, Essex, 1986, pp. 111– 150. [2] L.A. Hobbs, S.J. Blunden, P.J. Smith, in: H.J. Bowen (Ed.), Environmental Chemistry, The Royal Society of Chemistry, London, 1984, pp. 49–77. [3] M. Hoch, Appl. Geochem. 16 (2001) 719–743. [4] H. Rüdel, Ecotox. Environ. Safe. 56 (2003) 180–189. [5] M. Abalos, J.M. Bayona, R. Compano, M. Granados, C. Leal, M.D. Prat, J. Chromatogr. A 788 (1997) 1–49. [6] C. Brunori, I. Ipolyi, P. Massanisso, R. Morabito, Handbook of Environmental Chemistry. Water Pollution, vol. 5, 2006, pp. 51–70. [7] F. Smedes, A.S. De Jong, I.M. Davies, J. Environ. Monitor. 2 (2000) 541–549. [8] R.B. Rajendran, H. Tao, T. Nakazato, A. Miyazaki, Analyst (2000) 1757–1763. [9] M. Abalos, J.M. Bayona, P. Quevauviller, Appl. Organomet. Chem. 12 (1998) 541–549. [10] J.H. Huang, G. Ilgen, E. Matzner, Anal. Chim. Acta 493 (2003) 23–34. [11] Y. Cai, S. Rapsomanikis, M.O. Andreae, Anal. Chim. Acta 274 (1993) 243–251. ˇ canˇcar, Int. J. Environ. Anal. Chem. 87 (2007) [12] T.M. Nemaniˇc, R. Milaˇciˇc, J. Sˇ 615–625. [13] C. Marcic, I. Le Hecho, L. Denaix, G. Lespes, Chemosphere 65 (2006) 2322–2332. [14] J. Heroult, T. Zuliani, M. Bueno, L. Denaix, G. Lespes, Talanta 75 (2008) 486–493. [15] K. Fent, Sci. Total Environ. 185 (1996) 151–159. [16] I. Heninger, M. Potin-Gautier, M. Astruc, L. Galvez, V. Vignier, Chem. Spec. Bioavailab. 10 (1998) 1–10. [17] P. Pinel-Raffaitin, D. Amouroux, I. LeHecho, P. Rodriguez-Gonzalez, M. PotinGautier, Water Res. 42 (2008) 987–996. [18] P. Pinel-Raffaitin, P. Rodriguez-Gonzalez, M. Ponthieu, D. Amouroux, I. Le Hecho, L. Mazeas, O.F.X. Donard, M. Potin-Gautier, J. Anal. Atom. Spectrom. 22 (2007) 258–266. [19] T. Zuliani, G. Lespes, R. Milacic, J. Scancar, M. Potin-Gautier, J. Chromatogr. A 1188 (2008) 281–285. [20] R. Morabito, P. Massanisso, P. Quevauviller, TrAC—Trends Anal. Chem. 19 (2000) 113–119. [21] O.F.X. Donard, P. Quevauviller, A. Bruchet, Water Res. 27 (1993) 1085–1089. [22] I. Rodriguez, M. Santamarina, M.H. Bollain, M.C. Mejuto, R. Cela, J. Chromatogr. A 774 (1997) 379–387. ˇ canˇcar, Acta Chim. Slov. 54 (2007) [23] T.M. Nemaniˇc, L. Zupanˇciˇc-Kralj, R. Milaˇciˇc, J. Sˇ 40–48. [24] B. Radke, L. Leczynski, A. Wasik, J. Namiesnik, J. Bolalek, Chemosphere 73 (2008) 407–414. [25] C. Montigny, G. Lespes, M. Potin-Gautier, J. Chromatogr. A 819 (1998) 221–230. [26] J. Mocak, A.M. Bond, S. Mitchell, G. Scollary, Pure Appl. Chem. 69 (1997) 298–320.
Contents lists available at ScienceDirect
Talanta journal homepage: www.elsevier.com/locate/talanta
Development of the extraction method for the simultaneous determination of butyl-, phenyl- and octyltin compounds in sewage sludge ˇ canˇcar a Tea Zuliani a,b,∗ , Gaetane Lespes b , Radmila Milaˇciˇc a , Janez Sˇ a b
Department of Environmental Sciences, Joˇzef Stefan Institute, Jamova 39, 1000 Ljubljana, Slovenia Laboratoire de Chimie Analytique, UMR 5254, Université de Pau, BP 1155, F-64013 PAU Cedex, France
a r t i c l e
i n f o
Article history: Received 6 July 2009 Received in revised form 9 October 2009 Accepted 23 October 2009 Available online 10 November 2009 Keywords: Organotin compounds Sewage sludge Extraction procedures GC–MS
a b s t r a c t The toxicity and bioaccumulation of organotin compounds (OTCs) led to the development of sensitive and selective analytical methods for their determination. In the past much attention was assigned to the study of OTCs in biological samples, water and sediments, coming mostly from marine environment. Little information about OTCs pollution of terrestrial ecosystems is available. In order to optimise the extraction method for simultaneous determination of butyl-, phenyl- and octyltin compounds in sewage sludge five different extractants (tetramethylammonium hydroxide, HCl in methanol, glacial acetic acid, mixture of acetic acid and methanol (3:1), and mixture of acetic acid, methanol and water (1:1:1)), the presence or not of a complexing agent (tropolone), and the use of different modes of extraction (mechanical stirring, microwave and ultrasonic assisted extraction) were tested. Extracted OTCs were derivatised with sodium tetraethylborate and determined by gas chromatography coupled with mass spectrometer. Quantitative extraction of butyl-, phenyl- and octyltin compounds was obtained by the use of glacial acetic acid as extractant and mechanical stirring for 16 h or sonication for 30 min. The limits of detection and quantification for OTCs investigated in sewage sludge were in the ng Sn g−1 range. © 2009 Elsevier B.V. All rights reserved.
1. Introduction Organotin compounds (OTCs) are organometallic derivatives of tin that have the highest number of commercial applications than any other element [1]. Mostly they have been used as stabilizers and catalysts in the production of poly(vinyl chloride) (PVC), polyurethanes and silicones, as antifouling paints for ship hulls, and as agricultural and wood preservative fungicides [2–4]. Due to their widespread use, OTCs may be found in different ecosystems. Today, their high toxicity for aquatic and terrestrial organisms is well documented [3,4]. Moreover, because of their toxicity and bioaccumulation potential, OTCs have been registered as priority pollutants by the European Union in the Pollutant Emission Register (2000/479/EC) and in the Water Framework Directive (2000/60/EC). During last decades, a variety of analytical methods for the speciation of OTCs has been developed. Analytical procedures for OTCs speciation generally attempt to preserve only the organic radical during extraction, whereas the counterion and other tin heteroatomic bonds are cleaved during extraction or derivatisa-
∗ Corresponding author at: Department of Environmental Sciences, Joˇzef Stefan Institute, Jamova 39, 1000 Ljubljana, Slovenia. Tel.: +386 1 4773900; fax: +386 1 2519385. E-mail address: [email protected] (T. Zuliani). 0039-9140/$ – see front matter © 2009 Elsevier B.V. All rights reserved. doi:10.1016/j.talanta.2009.10.050
tion [5]. Sample preparation techniques for speciation analyses of OTCs in solid samples consist of several steps: solid–liquid extraction, simultaneous derivatisation and extraction into an organic solvent, separation and detection. One of the most difficult parts of the analysis of OTCs in sewage sludge is their extraction from the complex matrix. Extraction of OTCs from solid matrices keeping their speciation preserved is not straightforward because the binding forces between OTCs and matrices are species and matrices dependant [6,7]. In addition, some OTCs are not stable during extraction. Species transformation can be considered to be one of the most serious problems during phenyltin speciation analysis [8–10]. From the discovery of the toxicity of OTCs towards numerous marine species in late 1970s, most efforts have been devoted to the development of analytical methods for their determination in marine environments (seawater, sediments and biota) [5,11,12]. There are few studies concerning tin speciation in samples from terrestrial ecosystem [10,13,14]. OTCs are entering the terrestrial environment via the leaching from products made of PVC and silicone, by the use of pesticides and the amendment of cultivated soil with contaminated sewage sludge [15,16]. OTCs were also found in leachates from landfills [17,18]. Sewage sludge is an important source of OTCs in the terrestrial ecosystems [15,16]. Thus, reliable analytical methods are needed in order to evaluate the contribution of contaminated sewage sludge to the OTCs pollution of terrestrial environment. The aim
1946
T. Zuliani et al. / Talanta 80 (2010) 1945–1951
Table 1 Mass/charge (m/z) ratios and retention times for studied OTCs. OTCs
Retention time (min)
m/z
MBT DBT TBT MPhT DPhT TPhT MOcT DOcT TOcT TPrT
6.7 8.9 10.8 10.1 15.0 17.9 11.9 16.6 19.9 7.6
235, 179, 149 263, 179, 151 291, 263, 179 255, 197 303, 275, 197 351, 197 291, 179, 151 375, 263, 151 375, 263, 151 249, 235, 193
Ions used for quantification are reported in bold.
of our investigation was to set up an analytical procedure for the speciation of OTCs in sewage sludge by gas chromatography mass spectrometry (GC–MS). The influence of the extractant, extraction technique and the presence of complexing agent on the extraction efficiency of butyl-, phenyl- and octyltin compounds from sewage sludge have been studied. The selected methods encompass five different extractants (tetramethylammonium hydroxide (TMAH), hydrochloric acid (HCl) in methanol, glacial acetic acid, mixture of acetic acid and methanol (3:1), and mixture of acetic acid, methanol and water (1:1:1)), the presence or not of a complexing agent (tropolone), and the use of different modes of extraction (mechanical stirring, microwave and ultrasonic assisted extraction). Extracted OTCs were derivatised with sodium tetraethylborate (NaBEt4 ) and determined by GC–MS. 2. Experimental 2.1. Instrumentation For the separation and detection a Hewlett-Packard 6890 gas chromatograph (GC) (Hewlett-Packard, Waldbronn, Germany) coupled to a Hewlett-Packard 5972A mass spectrometer (MS) was used. The GC was equipped with a 30 m × 25 mm HP-MS5 polydimethylsiloxane capillary column (film thickness 0.25 m). The following temperature program was applied for the separation of OTCs: the column temperature was held at 80 ◦ C for the first 4 min, raised to 180 ◦ C at the heating rate of 10 ◦ C min−1 , then to 220 ◦ C at 20 ◦ C min−1 , and to 270 ◦ C at 30 ◦ C min−1 (held at this temperature for 6 min). The splitless injector port was kept at a temperature of 240 ◦ C and the transfer line at 280 ◦ C. Helium (He) was used as a carrier gas (1 mL min−1 ). The volume of injected sample was 1 L. The MS operated with the electronic impact (70 eV) ion source at a temperature of 180 ◦ C in the total ionization mode. Selected Ion Monitoring signals (SIM) were recorded. In Table 1 are presented the mass/charge (m/z) ratios of OTCs and their retention times. For the quantification the most abundant ions were used. Mechanical shaking during the extraction procedure was performed on an elliptical table Vibramax 40 (Tehtnica, Zˇ elezniki, Slovenia). Microwave assisted extraction was performed with a CEM MARS 5 Microwave Acceleration Reaction System, CEM Corporation (Matthews, NC, USA). Ultrasonic-assisted extraction of samples was performed with an ultrasonic bath Model 550D, VWR International (West Chester, PA, USA). Centrifugation of sample extracts was performed on a Heraeus (Osterode, Germany) Model 17S Sepatech Biofuge centrifuge and on a LC 320 centrifuge (Tehtnica, Zˇ elezniki, Slovenia). 2.2. Reagents Monobutyltin trichloride (MBT, 95%), monophenyltin trichloride (MPhT, 98%) and diphenyltin dichloride (DPhT, 96%) were
purchased from Aldrich (Milwaukee, WI, USA). Dibutyltin dichloride (DBT, 97%), tributyltin chloride (TBT, 96%), triphenyltin chloride (TPhT, 95%) and tripropyltin chloride (TPrT, 98%) were obtained from Merck (Darmstadt, Germany). Monooctyltin trichloride (MOcT, 98%), dioctyltin dichloride (DOcT, 98%) were obtained from LGC Promochem (Wesel, Germany) and trioctyltin chloride (TOcT, 95%) from Fluka (Buchs, Switzerland). Organotin standard stock solutions (1000 mg Sn L−1 ) were prepared in methanol. Working standard solutions (10 mg Sn L−1 ) were prepared weekly from stock standard solutions by dilution in Milli-Q water (18.2 M cm) (Millipore, Bedford, MA, USA). Standard solutions of 100 g Sn L−1 were prepared daily. All the standards were stored in the dark at 4 ◦ C. Glacial acetic acid, ammonia solution and nitric acid were obtained from J.T. Baker (Phillipsburg, NJ, USA). Methanol was purchased from Merck (Darmstadt, Germany). Sodium tetraethylborate (NaBEt4 ) was purchased from Strem Chemicals (Newburyport, MA, USA). NaBEt4 was dissolved in Milli-Q water to provide a 2% (m/v) ethylating solution. 2.3. Sample preparation Sewage sludge samples were dried at 50 ◦ C in the dark for 3 days, homogenised and sieved through a 0.2 mm sieve. The samples were then frozen at −20 ◦ C. 2.4. Analytical procedure 2.4.1. Extraction from the sample Approximately 1 g of dried sewage sludge was weighted in a polyethylene (PE) extraction tube. Based on our previous experiences 0.2 g of TPrT in methanol was added as internal standard [19]. After 15 min the samples were subjected to different extraction methods. 2.4.1.1. Method 1. 20 mL of tetramethylammonium hydroxide (TMAH) was added to the sample. The tubes were subjected to ultrasound assisted extraction for 2 h and centrifuged for 15 min at 4200 rpm. 2.4.1.2. Method 2. Samples were supplemented with 15 mL of 0.1 mol L−1 HCl in methanol. The closed tubes were firstly mechanically shaken on an elliptic table for 2 h and then subjected to ultrasonic assisted extraction for 1 h and centrifuged for 15 min at 4200 rpm. 2.4.1.3. Method 3. Samples were supplemented with 20 mL of glacial acetic acid. The tubes were subjected to closed vessel microwave assisted extraction for 4, 7 and 16 min. Microwave extraction conditions were as follows: 1200 W, 1 min ramp to 50 ◦ C, hold at that temperature for 3, 6 and 15 min. After the extraction, the suspension was centrifuged for 15 min at 4200 rpm. 2.4.1.4. Method 4. To the solid sample 5 mL of methanol and 15 mL of glacial acetic acid were added. The tubes were subjected to closed vessel microwave assisted extraction for 7 min. The microwave extraction conditions were as described in Section 2.4.1.3. After the extraction, the suspension was centrifuged for 15 min at 4200 rpm. 2.4.1.5. Method 5. Samples were supplemented with 20 mL mixture of acetic acid, methanol and water (1:1:1). The tubes were subjected to closed vessel microwave assisted extraction for 7 min. The microwave extraction conditions were as described in Section 2.4.1.3. After the extraction, the suspension was centrifuged for 15 min at 4200 rpm.
T. Zuliani et al. / Talanta 80 (2010) 1945–1951
1947
2.4.1.6. Method 6. To the solid sample 5 mL of 0.02% (w/v) tropolone in methanol and 15 mL of glacial acetic acid were added. The tubes were subjected to closed vessel microwave assisted extraction for 7 min. The microwave extraction conditions were as described in Section 2.4.1.3. After the extraction, the suspension was centrifuged for 15 min at 4200 rpm. 2.4.1.7. Method 7. Samples were supplemented with 20 mL of glacial acetic acid. The closed tubes were subjected to ultrasound assisted extraction for 5, 10, 20, 30 and 60 min. After the extraction, the suspension was centrifuged for 15 min at 4200 rpm. 2.4.1.8. Method 8. As Method 7. Acetic acid–methanol mixture (3:1) instead of glacial acetic acid as extractant was applied to the samples. 2.4.1.9. Method 9. To the solid sample 20 mL of glacial acetic acid was added. The closed tubes were mechanically shaken on an elliptic table for 1, 5, 15 and 24 h in the dark. Then, the suspension was centrifuged for 15 min at 4200 rpm. 2.4.2. Derivatisation and extraction Aliquots (1–4 mL) of the extract described above, 0.5 mL 2% (w/v) solution of NaBEt4 for the derivatisation and 1 mL of isooctane for the extraction of ethylated organotin species were added to 20 mL of sodium acetate-acetic acid buffer (pH 4.8). The mixture was shaken for 45 min at 300 rpm. The same procedure, with the exception that no sample was added, was applied to determine blanks. All the analyses were made in triplicate. 2.4.3. Spiking protocol The recovery of the studied OTCs from sewage sludge was investigated by spiking known amount of OTCs into samples of relatively low OTCs contents. For spiking of sewage sludge samples OTCs as chlorides dissolved in methanol were used. The equilibration step was carried out by shaking (350 rpm) the samples for 30 min. For each extraction three different spikes were prepared. One aliquot of the sewage sludge was spiked with 1000 ng Sn g−1 of monosubstituted butyl-, phenyl- and octyltin compounds (MBT, MPhT, MOcT). The second one was spiked with 1000 ng Sn g−1 of disubstituted butyl-, phenyl- and octyltin compounds (DBT, DPhT, DOcT) and the third with 1000 ng Sn g−1 of trisubstituted butyl-, phenyl- and octyltin compounds (TBT, TPhT, TOcT). The differences between recoveries depend only on the differences in extraction procedure, as the post-extraction steps of the analytical method remained the same. 3. Results and discussion Sewage sludge consists of a complex matrix. In order to release the molecular and ionic OTCs from organic matter in sewage sludge an appropriate digestion/extraction must be applied. In any speciation procedure, the extraction procedure must be carefully chosen so as to destroy the matrix but not to affect the authentic forms of the analyte molecules [20]. The removal of OTCs in wastewater treatment plants takes place in two steps. First, in the primary treatment mostly by sorption onto particulate matter of the wastewater [21]. Second, in the secondary biological treatment by sorption onto particulate organic sludge material and by assimilation of carbon from OTCs by microorganisms [15]. In bio-tissue the OTCs are strongly bound to the cellular structure [9]. Usually, for the extraction of OTCs from bio-tissues 25% water solution of TMAH or 0.1 mol L−1 HCl in methanol are applied [5,22,23]. Since sewage sludge from wastewater treatment plant is mostly composed of organic matter (dead biomass), those two extractants (Methods 1 and 2) were considered. On the other
Fig. 1. Extraction recoveries of butyl-, phenyl- and octyltin compounds from spiked sewage sludge sample (1000 ng Sn g−1 ) applying the extraction with 25% water solution of TMAH and 0.1 mol L−1 HCl in methanol.
hand, the bonds between OTCs and particulate organic matter are strong. So, extraction solvents that are usually applied to sediment samples were also verified. As glacial acetic acid and mixtures of acetic acid and methanol have been the most widely reported in the literature for the extraction of OTCs from sediments and soil [12,14,24], their performance on sewage sludge was examined (Methods 3–9). All the tested methods (1–9) are precisely described in Section 2. In Table 2 a schematic description of the selected methods is presented. 3.1. Methods 1 and 2 25% water solution of TMAH and 0.1 mol L−1 HCl in methanol are the most frequently applied solvents for the extraction of OTCs from biological samples [5,24]. As sewage sludge is mostly composed by organic matter, the extraction efficiency of those two solvents for the extraction of OTCs from sewage sludge was tested. In Fig. 1 the extraction recoveries of both solvents are presented. When TMAH was applied as extractant, an emulsion disturbing the analytical process during the derivatisation was formed. 25% water solution of TMAH was not efficient for the extraction of butly-, phenyl- and octyltin compounds from sewage sludge. The recoveries for all OTCs were around 10% (Fig. 1). As can be also seen from Fig. 1, when 0.1 mol L−1 HCl in methanol was used, the recoveries for butyltins, octyltins, DPhT and TPhT were between 40 and 60%, while MPhT was not extracted at all. High recovery values for DBT and DOcT, 100 and 105%, respectively, were observed. If OTCs in sewage sludge is bound to the cellular structure of the biomass TMAH would efficiently solubilise the matrix in order to release OTCs (as in fish and mussel tissue). From the results it may be deduced that OTCs present in the sewage sludge are not bound to the cellular structure of the biomass but are sorbed onto the particulate organic matter. Since the binding to particulate organic matter is stronger than that to biological tissue, TMAH was too weak for the extraction of OTCs from the complex matrix of sewage sludge. As highly polar mono- and disubstituted OTCs need acidification to increase the efficiency of extraction from the sample [7], the use of 0.1 mol L−1 HCl in methanol would favour their extraction. The results in Fig. 1 did not confirm that. It can be seen that monosubstituted OTCs were not efficiently extracted most probably due to too low acid concentration. Contrary, 0.1 mol L−1 HCl turned out to be too strong for trisubstituted OTCs provoking their degradation.
1948
T. Zuliani et al. / Talanta 80 (2010) 1945–1951
Table 2 Schematic description of tested extraction methods. Method
Extractant
Complexing agent
Mode of extraction
1 2 3 4 5 6 7 8 9
25% water solution TMAH 0.1 mol L−1 HCl in methanol Glacial acetic acid Glacial acetic acid:methanol (3:1) Glacial acetic acid:methanol:water (1:1:1) Glacial acetic acid:methanol (3:1) Glacial acetic acid Glacial acetic acid:methanol (3:1) Glacial acetic acid
– – – – – Tropolone – – –
Ultrasound assisted extraction Mechanical stirring and ultrasonication Microwave assisted extraction Microwave assisted extraction Microwave assisted extraction Microwave assisted extraction Ultrasound assisted extraction Ultrasound assisted extraction Mechanical stirring
Table 3 Extraction recoveries (%) of butyl-, phenyl- and octyltin compounds for spiked sewage sludge (1000 ng Sn g−1 ) applying MWAE with glacial acetic acid. OTCs
Extraction times 4 min
MBT DBT TBT MPhT DPhT TPhT MOcT DOcT TOcT
99 105 102 81 20 27 86 92 90
± ± ± ± ± ± ± ± ±
7 min 6 6 6 8 3 4 8 9 7
107 105 97 100 10 10 105 107 100
± ± ± ± ± ± ± ± ±
16 min 5 6 4 11 2 2 5 6 4
44 81 78 101 13 11 100 87 98
± ± ± ± ± ± ± ± ±
5 9 7 10 2 2 7 5 6
3.2. Methods from 3 to 6 Microwave assisted extraction (MWAE) was investigated in order to substantially reduce the extraction time for the determination of OTCs in sewage sludge. Extraction times of 4, 7 and 16 min were tested. Glacial acetic acid was used as extractant (Method 3). The results are presented in Table 3. It is evident from Table 3 that butyltin compounds were efficiently extracted with acetic acid after 4 and 7 min of MWAE. At longer extraction time the extraction efficiency of butyltin com-
pounds was appreciably reduced. In order to find out whether the degradation of butyltin compounds took place, an additional experiment was done. Two sewage sludge samples were spiked with DBT and TBT (1000 ng Sn g−1 ), respectively, and subjected to MWAE with glacial acetic acid for 16 min. Results are presented in Fig. 2(A) and (B). Data from Fig. 2(A) and (B) indicate that no degradation processess occured during the course of the experiment. In addition, no increase of the Sn(IV) peak was observed. Therefore, the degradation of MBT or DBT and TBT direct to Sn(IV) may be excluded. From the results, it may be presumed that lower extraction efficiencies of butyltin compounds after 16 min of MWAE are most probably related to more efficient extraction of the organic matter from sewage sludge. The co-extracted organic matter inhibits ethylation/extraction steps, resulting in poor butyltin compounds recoveries. It can be seen from Table 3 that MPhT was efficiently extracted after 7 min. The recoveries for DPhT and TPhT were about 20% after 4 min and below 10% at longer extraction times. To check if the degradation of phenyltin compounds occurred, two aliquots of sewage sludge were spiked with DPhT and TPhT (1000 ng Sn g−1 ), respectively, and subjected to MWAE with glacial acetic acid for 4 min, Fig. 3(A) and (B). From data of Fig. 3, it is obvious that TPhT was almost completely degraded via DPhT to MPhT. Also 90% of DPhT was degraded to MPhT.
Fig. 2. Degradation of (A) TBT and (B) DBT in spiked sewage sludge (1000 ng Sn g−1 of TBT and/or DBT) during MWAE with glacial acetic acid (16 min).
Fig. 3. Degradation of (A) TPhT and (B) DPhT in spiked sewage sludge (1000 ng Sn g−1 of TPhT and/or DPhT) during MWAE with glacial acetic acid (4 min).
T. Zuliani et al. / Talanta 80 (2010) 1945–1951
1949
Fig. 4. Extraction recoveries for spiked sewage sludge with OTCs (1000 ng Sn g−1 ) applying MWAE with glacial acetic acid, acetic acid and methanol (3:1), and acetic acid, methanol and water (1:1:1).
Organometallic degradation during MWAE has been previously reported by Abalos et al. [5] and Huang et al. [10]. The fast temperature increase of extracting solution induces dealkylation and dearylation of analytes [14], especially the most thermolabile, i.e. trisubstituted ones, which is in agreement with the presented data. Octyltin compounds were efficiently extracted after 7 min (Table 3). When subjected to longer extraction times only DOcT exhibits slightly lower extraction efficiency. Since there was no evidence of the degradation of DOcT, the lower extraction efficiency for DOcT after 16 min of MWAE may be related to the influence of the co-extracted organic matter. Based on results from Table 3 and Figs. 2 and 3, the optimal extraction time for butyl- and octyltin compounds applying MWAE and glacial acetic acid as extractant was found to be 7 min. The exceptions were phenyltin compounds, for which MWAE was not applicable. To examine the potential of other extractants, mixtures of acetic acid and methanol (3:1) as well as acetic acid, methanol and water (1:1:1), were subjected to MWAE for 7 min. The comparison between extraction efficiencies of OTCs from spiked sewage sludge samples (1000 ng Sn g−1 ) of the above-mentioned extractants is given in Fig. 4. Namely, the addition of methanol to the glacial acetic acid should enhance the extraction efficiency for TBT [6], while the extraction efficiency of phenyltin and octyltin compounds should not be affected [14,25]. From comparison of results (Fig. 4) it can be seen that, as expected, the extraction recovery of TBT using acetic acid–methanol (3:1) was slightly higher than with glacial acetic acid. For MBT, DBT and octyltin compounds the addition of methanol had no great influence on the extraction efficiency, while the extraction efficiency for phenyltin compounds was even worse. When water was added to glacial acetic acid and methanol quantitative extraction efficiency for TBT and moderate (85%) for DBT was obtained. The addition of water seemed to severely reduce the extraction efficiency of MBT, MPhT and octyltin compounds, most probably due to lower extractant strength. The most beneficial influence of water addition was observed for TPhT, where the extraction recovery raised to 100%. The addition of water and methanol to glacial acetic acid decreased the strength of the acid. This resulted in optimal condition for the extraction of TPhT, while for the efficient extraction of DPhT and MPhT the acid strength was too low. It was experimentally proven that MWAE using 0.02% tropolone as a component of the extractant solution (Method 6) was not appropriate for OTCs analyses. Tropolone enhanced the extraction of organic matter that resulted in significant increase of the back-
ground signal during the chromatographic separation of OTCs (see data from Fig. 5). 3.3. Methods 7 and 8 Ultrasound assisted extraction (USAE) has become a widely used extraction technique. In combination with glacial acetic acid and acetic acid and methanol, high recoveries for sediment and soil samples were obtained [12,14]. For the simultaneous extraction of butyl-, phenyl- and octyltin compounds from sewage sludge sample glacial acetic acid USAE was applied for 5, 10, 20, 30 and 60 min. In Table 4 the obtained extraction efficiencies are presented. It can be observed from Table 4 that the time of sonication has a great influence on the extraction efficiency. MBT and DBT were quantitatively extracted after 30 min, while for TBT 20 min of sonication gave quantitative recoveries. Longer extraction times did not have significant influence on the recovery of butyltin compounds.
Fig. 5. Chromatograms of butyl-, phenyl- and octyltin compounds from spiked sewage sludge sample (1000 ng Sn g−1 ) using MWAE with (A) methanol and acetic acid and (B) 0.02% (w/v) tropolone in methanol and acetic acid.
1950
T. Zuliani et al. / Talanta 80 (2010) 1945–1951
Table 4 Extraction recoveries of butyl-, phenyl- and octyltin compounds for spiked sewage sludge sample (1000 ng Sn g−1 ) applying USAE with glacial acetic acid. OTCs
Extraction time 5 min
MBT DBT TBT MPhT DPhT TPhT MOcT DOcT TOcT
Table 5 Extraction recoveries (%) of butyl-, phenyl- and octyltin compounds from spiked sewage sludge sample (1000 ng Sn g−1 ) applying MSAE with glacial acetic acid.
31 86 89 44 72 90 54 83 89
± ± ± ± ± ± ± ± ±
OTCs
10 min
6 8 7 10 9 10 8 7 9
80 83 99 67 81 81 82 83 100
± ± ± ± ± ± ± ± ±
20 min
7 6 10 12 8 9 7 6 9
63 85 100 63 82 85 80 108 99
± ± ± ± ± ± ± ± ±
30 min
6 9 10 5 7 7 8 10 9
100 97 100 93 92 100 92 99 96
± ± ± ± ± ± ± ± ±
Extraction time
60 min
5 4 7 5 4 7 5 4 5
98 101 99 86 88 97 102 100 100
± ± ± ± ± ± ± ± ±
1h
9 10 10 12 10 15 17 15 16
MBT DBT TBT MPhT DPhT TPhT MOcT DOcT TOcT
67 68 71 59 58 74 72 75 72
± ± ± ± ± ± ± ± ±
5h 7 10 8 6 7 10 7 15 10
74 79 75 67 74 78 84 80 80
± ± ± ± ± ± ± ± ±
16 h 7 1 4 7 6 4 8 2 3
97 92 109 100 98 97 104 97 95
± ± ± ± ± ± ± ± ±
24 h 5 4 5 6 5 6 4 3 5
82 80 75 40 28 62 90 87 85
± ± ± ± ± ± ± ± ±
8 4 3 3 4 6 9 2 3
3.4. Method 9
Fig. 6. Extraction recoveries of butyl-, phenyl- and octyltin compounds from spiked sewage sludge sample (1000 ng Sn g−1 ) applying USAE with acetic acid and methanol (3:1).
Phenyltins are also efficiently extracted after 30 min of USAE. The extraction recoveries for phenyltin compounds remained constant after 30 min of sonication. MOcT showed the same behaviour as phenyltin compounds, while DOcT was efficiently extracted after 20 and TOcT after 10 min. The extending of extraction time did not significantly influence the recoveries of these two octyltin compounds. The studied OTCs in the spiked sewage sludge sample were extracted also with a mixture of acetic acid and methanol (3:1) and subjected to USAE. The results are presented in Fig. 6. As it was the case of MWAE, the mixture of acetic acid and methanol lowered the extraction efficiencies for phenyltin compounds by USAE, while it did not have significant influence on the extraction of butyl- and octyltin compounds.
The most frequently used extraction mode for OTCs determination in solid samples is mechanical stirring (MSAE). For the simultaneous extraction of butyl-, phenyl- and octyltin compounds from sewage sludge sample MSAE using glacial acetic acid as extractant was applied for 1, 5, 16 and 24 h. In Table 5 the obtained extraction efficiencies are presented. As can be seen from Table 5 the extraction time has a significant influence on the extraction efficiency of MSAE. It is evident that butyl-, phenyl- and octyltin compounds are quantitatively extracted after 16 h and no degradation occurred. At longer extraction time (24 h) lower extraction efficiencies of butyl- and octyltin compounds are observed. Additional experiments where sewage sludge sample was spiked either with TBT and TOcT or with DBT and DOcT confirmed that no degradation occurred. Therefore, lower extraction efficiencies are related most probably to re-adsorption on particulate matter and/or highest amount of co-extracted matter from the sample. From Table 5, it is further evident that poor extraction recoveries after 24 h are obtained for phenyltin compounds. In the additional experiment sewage sludge sample was spiked with either TPhT or DPhT. Under these conditions important degradation of TPhT and DPhT occurred. In addition, no increase of the Sn(IV) peak was observed. Therefore, the degradation of MPhT to Sn(IV) may be excluded. Lower extraction recovery for MPhT after 24 h of MSAE with glacial acetic acid may be due to the influence of the coextracted matter from the sample and re-adsorption on particulate matter. Since glacial acetic acid applying MSAE for 16 h quantitatively extracted butyl-, phenyl- and octyltin compounds from sewage sludge other extracting agents (acetic acid and methanol (3:1), and acetic acid, methanol and water (1:1:1)) were not tested.
Table 6 Limits of detection (LOD), quantification (LOQ) and repeatability (as RSD) for MSAE and USAE of butyl-, phenyl- and octyltin compounds in sewage sludge. OTCs
MSAE a
USAE −1
LOD (ng Sn g MBT DBT TBT MPhT DPhT TPhT MOcT DOcT TOcT
27 28 40 42 9 5 19 13 16
)
b
−1
LOQ (ng Sn g 53 54 73 82 18 12 39 25 31
Concentrations are expressed in ng Sn g−1 (dry mass basis). a Limits of detection. b Limits of quantification.
)
RSD (%)
LOD (ng Sn g−1 )
LOQ (ng Sn g−1 )
RSD (%)
8 3 4 5 2 3 3 4 2
24 26 39 43 11 7 17 12 17
49 53 71 83 23 15 35 24 32
6 5 3 5 3 3 9 7 4
T. Zuliani et al. / Talanta 80 (2010) 1945–1951
3.5. Analytical performances of MSAE and USAE Limits of detection (LOD) and quantification (LOQ) for MSAE and USEA were calculated on the basis of the standard addition method calibration curves [26]. The results for LOD, LOQ and repeatability (as relative standard deviation, RSD) for both methods are presented in Table 6. Based on results from Tables 4 and 5 it can be concluded that quantitative extraction of butyltin compounds (MBT, DBT, TBT), phenyltin compounds (MPhT, DPhT, TPhT) and octyltin compounds (MOcT, DOcT, TOcT) is obtained by the use of glacial acetic acid as extractant and MSAE for 16 h or USAE for 30 min. As can be seen from Table 6, LOD and LOQ for the two methods are similar. Therefore, the choice of the extraction mode depends on the availability of the laboratory equipment. The experiment also confirmed that MWAE and extractants as TMAH, 0.1 mol L−1 HCl in methanol, mixtures of acetic acid and methanol (3:1) and acetic acid, methanol and water (1:1:1) were not applicable for the simultaneous determination of butyl-, phenyl- and octyltin compounds in sewage sludge. 4. Conclusions For the extraction of OTCs from sewage sludge five extraction solvents (TMAH, HCl diluted with methanol, glacial acetic acid, acetic acid with methanol (3:1), and acetic acid, methanol and water (1:1:1)), the use of complexing agent (tropolone), and three extraction modes (MSAE, USAE and MWAE) were tested. Regardless of extractants applied, MWAE was not appropriate for simultaneous determination of butyl-, phenyl- and octyltin compounds in sewage sludge. Quantitative extraction of butyl-, phenyl- and octyltin compounds was obtained by the use of glacial acetic acid as extractant and MSAE for 16 h or sonication for 30 min. Both extraction procedures represent reliable analytical tool for the determination of the content of OTCs in sewage sludge. Acknowledgements The authors would like to acknowledge the financial support of the Ministry of Higher Education, Science and Technology
1951
of the Republic of Slovenia within the research programme P10143, French Ministry of Education and PROTEUS bilateral project: “Organotin compounds in the Slovenian and French environment”. References [1] A. Chapman, S.J. Blunden, in: P.J. Craig (Ed.), Organometallic Compounds in the Environment: Principles and Reactions, Longman, Essex, 1986, pp. 111– 150. [2] L.A. Hobbs, S.J. Blunden, P.J. Smith, in: H.J. Bowen (Ed.), Environmental Chemistry, The Royal Society of Chemistry, London, 1984, pp. 49–77. [3] M. Hoch, Appl. Geochem. 16 (2001) 719–743. [4] H. Rüdel, Ecotox. Environ. Safe. 56 (2003) 180–189. [5] M. Abalos, J.M. Bayona, R. Compano, M. Granados, C. Leal, M.D. Prat, J. Chromatogr. A 788 (1997) 1–49. [6] C. Brunori, I. Ipolyi, P. Massanisso, R. Morabito, Handbook of Environmental Chemistry. Water Pollution, vol. 5, 2006, pp. 51–70. [7] F. Smedes, A.S. De Jong, I.M. Davies, J. Environ. Monitor. 2 (2000) 541–549. [8] R.B. Rajendran, H. Tao, T. Nakazato, A. Miyazaki, Analyst (2000) 1757–1763. [9] M. Abalos, J.M. Bayona, P. Quevauviller, Appl. Organomet. Chem. 12 (1998) 541–549. [10] J.H. Huang, G. Ilgen, E. Matzner, Anal. Chim. Acta 493 (2003) 23–34. [11] Y. Cai, S. Rapsomanikis, M.O. Andreae, Anal. Chim. Acta 274 (1993) 243–251. ˇ canˇcar, Int. J. Environ. Anal. Chem. 87 (2007) [12] T.M. Nemaniˇc, R. Milaˇciˇc, J. Sˇ 615–625. [13] C. Marcic, I. Le Hecho, L. Denaix, G. Lespes, Chemosphere 65 (2006) 2322–2332. [14] J. Heroult, T. Zuliani, M. Bueno, L. Denaix, G. Lespes, Talanta 75 (2008) 486–493. [15] K. Fent, Sci. Total Environ. 185 (1996) 151–159. [16] I. Heninger, M. Potin-Gautier, M. Astruc, L. Galvez, V. Vignier, Chem. Spec. Bioavailab. 10 (1998) 1–10. [17] P. Pinel-Raffaitin, D. Amouroux, I. LeHecho, P. Rodriguez-Gonzalez, M. PotinGautier, Water Res. 42 (2008) 987–996. [18] P. Pinel-Raffaitin, P. Rodriguez-Gonzalez, M. Ponthieu, D. Amouroux, I. Le Hecho, L. Mazeas, O.F.X. Donard, M. Potin-Gautier, J. Anal. Atom. Spectrom. 22 (2007) 258–266. [19] T. Zuliani, G. Lespes, R. Milacic, J. Scancar, M. Potin-Gautier, J. Chromatogr. A 1188 (2008) 281–285. [20] R. Morabito, P. Massanisso, P. Quevauviller, TrAC—Trends Anal. Chem. 19 (2000) 113–119. [21] O.F.X. Donard, P. Quevauviller, A. Bruchet, Water Res. 27 (1993) 1085–1089. [22] I. Rodriguez, M. Santamarina, M.H. Bollain, M.C. Mejuto, R. Cela, J. Chromatogr. A 774 (1997) 379–387. ˇ canˇcar, Acta Chim. Slov. 54 (2007) [23] T.M. Nemaniˇc, L. Zupanˇciˇc-Kralj, R. Milaˇciˇc, J. Sˇ 40–48. [24] B. Radke, L. Leczynski, A. Wasik, J. Namiesnik, J. Bolalek, Chemosphere 73 (2008) 407–414. [25] C. Montigny, G. Lespes, M. Potin-Gautier, J. Chromatogr. A 819 (1998) 221–230. [26] J. Mocak, A.M. Bond, S. Mitchell, G. Scollary, Pure Appl. Chem. 69 (1997) 298–320.