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Determination of Biodegradation Products from Benzene, Toluene, Ethylbenzene and Xylenes in Seawater by Purge and Trap Gas Chromatography
CHINESE JOURNAL OF ANALYTICAL CHEMISTRY Volume 34, Issue 10, October 2006 Online English edition of the Chinese language journal Cite this article as: Chin J Anal Chem, 2006, 34(10), 1361−1365.
RESEARCH PAPER
Determination of Biodegradation Products from Benzene, Toluene, Ethylbenzene and Xylenes in Seawater by Purge and Trap Gas Chromatography Han Dongqiang, Ma Wanyun*, Chen Dieyan Key Laboratory for Atomic and Molecular Nanosciences of Ministry of Education, Department of Physics, Tsinghua University, Beijing 100084, China
Abstract: Benzene, toluene, ethylbenzene and xylenes (BTEX) are commonly found in crud oil and used as the direct indicating substance in the Geochemical Investigation for Oil and Gas. BTEX are easily volatile and can be degraded by microorganism, which affects the precise measurement seriously. A method for the determination of the biodegradation process of BTEX in seawater is described. The determination is performed by dynamic headspace (purge and trap) gas chromatography with a photoionization detector (PID). The features of the described method are: detection limits of 7.3–13.2 ng l–1, no sample preparation required, and recovery rate of 92.84%–100.92%. The results of the BTEX biodegradation process are of great significance in the collection, transportation, preservation, and measurement of samples in the Geochemical Investigation for Oil and Gas. Key Words: Benzene; Toluene; Ethylbenzene; Xylenes; Purge and trap; Gas chromatography; Biodegradation; Seawater
1 Introduction Benzene, toluene, ethylbenzene, and three isomers (ortho-, meta- and para-) of xylene, which are collectively known as BTEX, are widely used as the direct indicating substance of oil and gas. Determining the concentration and distribution of BTEX in seawater and seabed’s sediment is very important in the geochemical investigation for oil and gas. However, BTEX are easily volatile and can be degraded by microorganism, which affects the precise measurement seriously. Thus, an understanding of the biodegradation of BTEX[1,2] in seawater environment is of great significance in the collection, transportation, preservation, and measurement of samples in the Geochemical Investigation for oil and gas. With the change of the seawater’s depth, the environment of the ocean such as temperature, pressure, light, and oxygen varies greatly. Accordingly, the kinds of the microorganism are changing continuously too. In the surface of the ocean, there are various kinds of aerobic, autotrophy microorganism;
and in the bottom of the ocean, there are various kinds of anaerobic microorganism[3]. The biodegradation of BTEX by microorganism varies with the change of the ocean environment. Therefore, it is vital to study the biodegradation process of BTEX by microorganism in the geochemical investigation for oil and gas. Previous BTEX studies usually focus on two aspects: one is the study of the biodegradation of the environment pollution[4–8] and this part is mainly concerned with the extent of the spread contamination and the effectiveness of intrinsic bioremediation cleanup operations; the other is the study of high-sensitive determination methods of BTEX[9–17]. This part centralizes on the technology of high-sensitive determination methods of BTEX. However, few works have been reported on the biodegradation of BTEX in seawater environment until now and considerable attention has been given to the toxicity of BTEX on halobios. The determination of BTEX by purge and trap gas chromatography is highly sensitive, effective, and rapid[18]. In this study, the biodegradation process of BTEX in
Received 18 January 2006; accepted 25 April 2006 * Corresponding author. Email: [email protected] This work was supported by the National High Technology Research and Development Program of China (No.2002AA615160). Copyright © 2006, Changchun Institute of Applied Chemistry, Chinese Academy of Sciences. Published by Elsevier Limited. All rights reserved.
HAN Dongqiang et al. / Chinese Journal of Analytical Chemistry, 2006, 34(10): 1361–1365
2 Experimental
The measurements of samples were carried out at 0 day, 1 day, 2 days, 4 days, 8 days, 15 days, 20 days, and 30 days (Total 8 time points). At each time point, 4 groups of sample as mentioned above were measured respectively and each sample was measured 3–4 times repeatedly.
2.1 Instrument and reagents
2.6
Benzene, toluene, ethylbenzene, p-xylene, m-xylene, o-xylene, and iso-propylbenzene were dissolved in methanol (Benzenes in methanol, State Environmental Protection Administration of China) and the concentration of BTEX was 201–206 mg l–1 with the uncertain extent about (±7–±14) mg l–1; Wahaha pure water (Hangzhou Wahaha, Ltd., China); Seawater samples of the South China Sea (Collected depth: about 90 m); O.I. Analytical Eclipse model 4660; Tenex trap and Gas Chromatography 500 with 10.2 eV PID Samples preparation (PerkinElmer, USA).
Purge gas: N2 (high purity of 99.999%); the flux of the purge gas was 40 ml min–1. The properties of the purge and trap processes were set as follows: sample temperature: 80℃ and purge 10 min; the bake temperature of water management: 210oC and bake for 8 min; the trap temperature: 20oC; the desorb temperature: 180oC and desorb time 2 min. Carrier gas: N2 (high purity of 99.999%) with a flux of 1ml min–1 (18.4 cm s–1). All tech capillary columns (stationary phase: ATTM-1, film thickness: 0.25 μm; length: 60 m, and inner diameter: 0.32 mm). The distributary rate was 10:1. The injector temperature was 180oC and the temperature of the PID was 250oC. The oven temperature program was that the initial temperature was 40oC holding for 5 min, then ramping up to 180oC at 10oC min–1 and holding for 20 min. Under the conditions employed, the system of purge and trap gas chromatography has high-sensitivity and BTEX can be separated effectively. The chromatography of the abbreviation of benzene, toluene, ethylbenzene, and xylenes of 120 ng l–1 standard sample refers to Fig.1.
seawater samples is studied by dynamic headspace (purge and trap) gas chromatography with a photoionization detector (PID).
2.2
Preparation of seawater samples
First, the benzenes in methanol standard sample with concentration of 200 mg l–1 were used to prepare the solution with concentration of 200 μg l–1 for further use (‘the second BTEX standard sample’). The method was as follows: 100 μl benzenes in methanol standard sample were taken and then diluted by 1000 times with pure water. Next, the seawater sample with concentration 1 μg l–1 was prepared. In this study, 5 ml of the second BTEX standard sample was taken precisely and then diluted by 200 times with the South China Sea water that was collected.
Instrument conditions
2.3 Preparation of control samples To distinguish the biodegradation from the volatilization of BTEX, a new method, known as the ‘control method’, was adopted; the pure water sample, control sample, was prepared using pure water (containing no microorganism) and 1 μg l–1 of the second BTEX standard sample. The method of preparation was the same as that for the seawater sample. 2.4
Samples grouping and preservation
Fig.1 Chromatograms of BTEX in 120 ng l–1 standard sample 1. benzene; 2. toluene; 3. ethylbenzene; 4. m-, p-xylene; 5. o-xylene;
In the interest of studying the biodegradation process of BTEX in seawater sample at different temperatures such as the room temperature, low temperature (0oC), 4 groups of sample were prepared: seawater sample preserved at low temperature (0oC); control sample preserved at low temperature (0oC); seawater sample preserved at room temperature; control sample preserved at room temperature. 2.5
Samples measurements
6. iso-propylbenzene
3
Results and discussion
3.1 Analysis of pure water and the South China Sea water First, the samples of pure water and the South China Sea water were measured and the Chromatograms of the abbreviation of BTEX (refer to Fig.2) were obtained. Fig.2 showed that the concentration of BTEX in pure water and the
HAN Dongqiang et al. / Chinese Journal of Analytical Chemistry, 2006, 34(10): 1361–1365
South China Sea water was extremely small (<10 ng l–1) and it was suitable for preparing the seawater sample and the control sample, which have the same concentration of 1 μg l–1.
from the measurement was calculated using the calibration of the system and thus the recovery rate of seawater samples was obtained, i.e., benzene 100.92%; toluene 99.85%; ethylbenzene 98.70%; p,m-xylene 98.41%; o-xylene 92.84%. 3.4
Detection limits
The 10 ng l–1 of sample was prepared using 200 mg l–1 benzenes in the methanol standard sample and pure water were measured more than 10 times. The detection limits were then calculated using the triple Signal-to-Noise method, the confidence coefficient of which can reach up to 95%. The detection limits of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene were 7.3 ng l–1, 8.1 ng l–1, 11.4 ng l–1, 8.3 ng l–1 and 13.2 ng l–1, respectively. 3.5
Fig.2 Chromatograms of BTEX in pure water and the South China Sea water A. the South China Sea water; B. pure water; a. 0 day; b. 4 days. Peaks 1– 6 are the same as in Fig.1.
3.2
Calibration
To calibrate the system of purge and trap gas chromatography, an external standard was established, that is, 400 ng l–1, 800 ng l–1, and 1200 ng l–1 standard samples were prepared and each sample was measured 3–5 times repeatedly at each time point. The peak area was averaged, obtained from the chromatography of BTEX of benzene, toluene, ethylbenzene, o-xylene, and p,m-xylene of 8 time points and the peak area (μV s) vs concentration (ng l–1) linear correlation was obtained. respectively. The linear correlation coefficients of benzene, toluene, ethylbenzene, o-xylene, and p,m-xylene are 0.999–1.000, which satisfy the requirements of the quantitative analysis of the seawater sample and the control sample. 3.3
Recovery rate of seawater samples
The peak areas of benzene, toluene, ethylbenzene, o-xylene, and p,m-xylene were averaged after more than 10 times’ measurement of the South China Sea water repeatedly and were regarded as the background peak areas, respectively. Then, 1 μg l–1 seawater sample was prepared using the South China Sea water and 200 mg l–1 benzenes in the methanol standard sample, and measured using the purge and trap gas chromatography. Finally, the actual concentration obtained
Biodegradation of BTEX in seawater samples
The biodegradation process of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene depends on the temperature. Figs.2A and 2B represented chromatograms of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene at 0 day and 4 days in 1 μg l–1 seawater sample at room temperature and low temperature (0oC), respectively. By comparing the chromatograms of BTEX at 0 day with the chromatograms at 4 days, it can be concluded that the peaks of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene at 0 day and 4 days in seawater samples at room temperature changed apparently and thus the biodegradation was strong, but the peaks at low temperature (0oC) changed insignificantly and thus the biodegradation was faint. According to the calibration equation of the system established by external standard, the concentration of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene was determined at 0 day, 1 day, 2 days, 4 days, 15 days, 20 days, and 30 days in seawater sample and control sample at room temperature, respectively and similarly for the seawater sample and control sample at low temperature (0oC). The volume of the sample was 0.025 l uniformly and thus when the international unit of mass (ng) was adopted, the concentration of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene was determined using the following formula: M=C×V Where C represents the concentration of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene obtained using the calibration equation of the system, and V represents the volume of the sample, that is, 0.025 l. The complete experiment was carried out 2 times repeatedly following the same procedures as mentioned above. The concentrations of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene at 8 time points in seawater sample and control sample were then averaged at room temperature respectively and similarly for the seawater sample and the control sample
HAN Dongqiang et al. / Chinese Journal of Analytical Chemistry, 2006, 34(10): 1361–1365
at low temperature (0oC). Finally, sketch maps of the mass-time of BTEX were made respectively (refer to Fig.3 and Fig.4). The concentration change of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene in the control sample is ascribed to the volatility of BTEX because pure water
basically contains no microorganism. Therefore, the biodegradation effect of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene can be obtained by comparing the concentration of the seawater sample with the concentration of the control sample at the same time point.
Fig. 3 The concentration change of BTEX in seawater sample and control sample at room temperature a: control sample; b: seawater sample
Fig. 4 The concentration change of BTEX in seawater sample and control sample at 0℃ a: control sample; b: seawater sample
The concentration of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene in the seawater sample at room temperature descends apparently and the falling trend can be classified into three periods. The concentration of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene declines slowly in 2 days (basically at the same level) but falls quickly from 2 days to 8 days(the biodegradation effect is evident). After 8 days, it is difficult to detect benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene. On the contrary, the trend of the concentration change of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene in the seawater
sample is basically in accord with that in the control sample at low temperature(0oC) and thus the biodegradation of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene by microorganism is faint. The volatilization, degradation, and the total percentage concentration change of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene at room temperature at special time point (8 days) refers to Table 1.
4
Conclusions We conclude with the following comments. Firstly, the
HAN Dongqiang et al. / Chinese Journal of Analytical Chemistry, 2006, 34(10): 1361–1365
biodegradation of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene are similar to each other in seawater sample at room temperature, that is, the concentration of BTEX remains well within 2 days and most of these are degraded by microorganism at 8 days. However, the concentration of BTEX remains well all the time and the biodegradation is faint in the seawater sample at low temperature (0ºC). Secondly, the biodegradation of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene is predominant when compared with the volatilization in the seawater sample at room temperature. However, the biodegradation and the volatilization of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene are not very evident in the seawater sample at low temperature (0ºC). Therefore, the biodegradation process of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene
depends on the temperature. Thirdly, the seawater sample must be measured immediately or saved at low temperature when collected in the geochemical investigation for oil and gas, which uses BTEX as the indicating substance. In conclusion, the concentration of benzene, toluene, ethylbenzene, p,m-xylene and o-xylene changes slowly within 2 days but most of these are biodegraded and volatilized at 8 days in seawater at room temperature, which attracts attention to the collection, transportation, preservation, and measurement of samples. Whether the sample is measured immediately or not affects the correlation between the abnormity of BTEX and the oil or gas field seriously. Therefore, to ensure the precision of the measurement results, it is better to measure the samples on the spot when collected.
Table 1 The volatilization, degradation, and total percentage change of BTEX concentration at room temperature at special time points (8 days) Component
Benzene (%)
Toluene (%)
Ethylbenzene (%)
p, m -Xylene (%)
Volatilization
16.48
15.91
18.13
17.35
9.91
Degradation
81.32
82.29
75.86
76.36
85.86
Total
97.80
98.20
93.99
93.71
95.77
Acknowledgements
[9]
o-Xylene (%)
Fu J M, Sheng G Y, Chen Y, Wang X M, Min Y S, Li S H, Chen L Y, Wang Z S. Climatic and Environmental Research, 1997, 2(1):
The authors express their thanks to Lab assistants Liang Xiao, Li Hongyan for their efforts in seawater samples’ measurement and postgraduate Yin Hailian for her thoughtful English translation discussions.
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RESEARCH PAPER
Determination of Biodegradation Products from Benzene, Toluene, Ethylbenzene and Xylenes in Seawater by Purge and Trap Gas Chromatography Han Dongqiang, Ma Wanyun*, Chen Dieyan Key Laboratory for Atomic and Molecular Nanosciences of Ministry of Education, Department of Physics, Tsinghua University, Beijing 100084, China
Abstract: Benzene, toluene, ethylbenzene and xylenes (BTEX) are commonly found in crud oil and used as the direct indicating substance in the Geochemical Investigation for Oil and Gas. BTEX are easily volatile and can be degraded by microorganism, which affects the precise measurement seriously. A method for the determination of the biodegradation process of BTEX in seawater is described. The determination is performed by dynamic headspace (purge and trap) gas chromatography with a photoionization detector (PID). The features of the described method are: detection limits of 7.3–13.2 ng l–1, no sample preparation required, and recovery rate of 92.84%–100.92%. The results of the BTEX biodegradation process are of great significance in the collection, transportation, preservation, and measurement of samples in the Geochemical Investigation for Oil and Gas. Key Words: Benzene; Toluene; Ethylbenzene; Xylenes; Purge and trap; Gas chromatography; Biodegradation; Seawater
1 Introduction Benzene, toluene, ethylbenzene, and three isomers (ortho-, meta- and para-) of xylene, which are collectively known as BTEX, are widely used as the direct indicating substance of oil and gas. Determining the concentration and distribution of BTEX in seawater and seabed’s sediment is very important in the geochemical investigation for oil and gas. However, BTEX are easily volatile and can be degraded by microorganism, which affects the precise measurement seriously. Thus, an understanding of the biodegradation of BTEX[1,2] in seawater environment is of great significance in the collection, transportation, preservation, and measurement of samples in the Geochemical Investigation for oil and gas. With the change of the seawater’s depth, the environment of the ocean such as temperature, pressure, light, and oxygen varies greatly. Accordingly, the kinds of the microorganism are changing continuously too. In the surface of the ocean, there are various kinds of aerobic, autotrophy microorganism;
and in the bottom of the ocean, there are various kinds of anaerobic microorganism[3]. The biodegradation of BTEX by microorganism varies with the change of the ocean environment. Therefore, it is vital to study the biodegradation process of BTEX by microorganism in the geochemical investigation for oil and gas. Previous BTEX studies usually focus on two aspects: one is the study of the biodegradation of the environment pollution[4–8] and this part is mainly concerned with the extent of the spread contamination and the effectiveness of intrinsic bioremediation cleanup operations; the other is the study of high-sensitive determination methods of BTEX[9–17]. This part centralizes on the technology of high-sensitive determination methods of BTEX. However, few works have been reported on the biodegradation of BTEX in seawater environment until now and considerable attention has been given to the toxicity of BTEX on halobios. The determination of BTEX by purge and trap gas chromatography is highly sensitive, effective, and rapid[18]. In this study, the biodegradation process of BTEX in
Received 18 January 2006; accepted 25 April 2006 * Corresponding author. Email: [email protected] This work was supported by the National High Technology Research and Development Program of China (No.2002AA615160). Copyright © 2006, Changchun Institute of Applied Chemistry, Chinese Academy of Sciences. Published by Elsevier Limited. All rights reserved.
HAN Dongqiang et al. / Chinese Journal of Analytical Chemistry, 2006, 34(10): 1361–1365
2 Experimental
The measurements of samples were carried out at 0 day, 1 day, 2 days, 4 days, 8 days, 15 days, 20 days, and 30 days (Total 8 time points). At each time point, 4 groups of sample as mentioned above were measured respectively and each sample was measured 3–4 times repeatedly.
2.1 Instrument and reagents
2.6
Benzene, toluene, ethylbenzene, p-xylene, m-xylene, o-xylene, and iso-propylbenzene were dissolved in methanol (Benzenes in methanol, State Environmental Protection Administration of China) and the concentration of BTEX was 201–206 mg l–1 with the uncertain extent about (±7–±14) mg l–1; Wahaha pure water (Hangzhou Wahaha, Ltd., China); Seawater samples of the South China Sea (Collected depth: about 90 m); O.I. Analytical Eclipse model 4660; Tenex trap and Gas Chromatography 500 with 10.2 eV PID Samples preparation (PerkinElmer, USA).
Purge gas: N2 (high purity of 99.999%); the flux of the purge gas was 40 ml min–1. The properties of the purge and trap processes were set as follows: sample temperature: 80℃ and purge 10 min; the bake temperature of water management: 210oC and bake for 8 min; the trap temperature: 20oC; the desorb temperature: 180oC and desorb time 2 min. Carrier gas: N2 (high purity of 99.999%) with a flux of 1ml min–1 (18.4 cm s–1). All tech capillary columns (stationary phase: ATTM-1, film thickness: 0.25 μm; length: 60 m, and inner diameter: 0.32 mm). The distributary rate was 10:1. The injector temperature was 180oC and the temperature of the PID was 250oC. The oven temperature program was that the initial temperature was 40oC holding for 5 min, then ramping up to 180oC at 10oC min–1 and holding for 20 min. Under the conditions employed, the system of purge and trap gas chromatography has high-sensitivity and BTEX can be separated effectively. The chromatography of the abbreviation of benzene, toluene, ethylbenzene, and xylenes of 120 ng l–1 standard sample refers to Fig.1.
seawater samples is studied by dynamic headspace (purge and trap) gas chromatography with a photoionization detector (PID).
2.2
Preparation of seawater samples
First, the benzenes in methanol standard sample with concentration of 200 mg l–1 were used to prepare the solution with concentration of 200 μg l–1 for further use (‘the second BTEX standard sample’). The method was as follows: 100 μl benzenes in methanol standard sample were taken and then diluted by 1000 times with pure water. Next, the seawater sample with concentration 1 μg l–1 was prepared. In this study, 5 ml of the second BTEX standard sample was taken precisely and then diluted by 200 times with the South China Sea water that was collected.
Instrument conditions
2.3 Preparation of control samples To distinguish the biodegradation from the volatilization of BTEX, a new method, known as the ‘control method’, was adopted; the pure water sample, control sample, was prepared using pure water (containing no microorganism) and 1 μg l–1 of the second BTEX standard sample. The method of preparation was the same as that for the seawater sample. 2.4
Samples grouping and preservation
Fig.1 Chromatograms of BTEX in 120 ng l–1 standard sample 1. benzene; 2. toluene; 3. ethylbenzene; 4. m-, p-xylene; 5. o-xylene;
In the interest of studying the biodegradation process of BTEX in seawater sample at different temperatures such as the room temperature, low temperature (0oC), 4 groups of sample were prepared: seawater sample preserved at low temperature (0oC); control sample preserved at low temperature (0oC); seawater sample preserved at room temperature; control sample preserved at room temperature. 2.5
Samples measurements
6. iso-propylbenzene
3
Results and discussion
3.1 Analysis of pure water and the South China Sea water First, the samples of pure water and the South China Sea water were measured and the Chromatograms of the abbreviation of BTEX (refer to Fig.2) were obtained. Fig.2 showed that the concentration of BTEX in pure water and the
HAN Dongqiang et al. / Chinese Journal of Analytical Chemistry, 2006, 34(10): 1361–1365
South China Sea water was extremely small (<10 ng l–1) and it was suitable for preparing the seawater sample and the control sample, which have the same concentration of 1 μg l–1.
from the measurement was calculated using the calibration of the system and thus the recovery rate of seawater samples was obtained, i.e., benzene 100.92%; toluene 99.85%; ethylbenzene 98.70%; p,m-xylene 98.41%; o-xylene 92.84%. 3.4
Detection limits
The 10 ng l–1 of sample was prepared using 200 mg l–1 benzenes in the methanol standard sample and pure water were measured more than 10 times. The detection limits were then calculated using the triple Signal-to-Noise method, the confidence coefficient of which can reach up to 95%. The detection limits of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene were 7.3 ng l–1, 8.1 ng l–1, 11.4 ng l–1, 8.3 ng l–1 and 13.2 ng l–1, respectively. 3.5
Fig.2 Chromatograms of BTEX in pure water and the South China Sea water A. the South China Sea water; B. pure water; a. 0 day; b. 4 days. Peaks 1– 6 are the same as in Fig.1.
3.2
Calibration
To calibrate the system of purge and trap gas chromatography, an external standard was established, that is, 400 ng l–1, 800 ng l–1, and 1200 ng l–1 standard samples were prepared and each sample was measured 3–5 times repeatedly at each time point. The peak area was averaged, obtained from the chromatography of BTEX of benzene, toluene, ethylbenzene, o-xylene, and p,m-xylene of 8 time points and the peak area (μV s) vs concentration (ng l–1) linear correlation was obtained. respectively. The linear correlation coefficients of benzene, toluene, ethylbenzene, o-xylene, and p,m-xylene are 0.999–1.000, which satisfy the requirements of the quantitative analysis of the seawater sample and the control sample. 3.3
Recovery rate of seawater samples
The peak areas of benzene, toluene, ethylbenzene, o-xylene, and p,m-xylene were averaged after more than 10 times’ measurement of the South China Sea water repeatedly and were regarded as the background peak areas, respectively. Then, 1 μg l–1 seawater sample was prepared using the South China Sea water and 200 mg l–1 benzenes in the methanol standard sample, and measured using the purge and trap gas chromatography. Finally, the actual concentration obtained
Biodegradation of BTEX in seawater samples
The biodegradation process of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene depends on the temperature. Figs.2A and 2B represented chromatograms of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene at 0 day and 4 days in 1 μg l–1 seawater sample at room temperature and low temperature (0oC), respectively. By comparing the chromatograms of BTEX at 0 day with the chromatograms at 4 days, it can be concluded that the peaks of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene at 0 day and 4 days in seawater samples at room temperature changed apparently and thus the biodegradation was strong, but the peaks at low temperature (0oC) changed insignificantly and thus the biodegradation was faint. According to the calibration equation of the system established by external standard, the concentration of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene was determined at 0 day, 1 day, 2 days, 4 days, 15 days, 20 days, and 30 days in seawater sample and control sample at room temperature, respectively and similarly for the seawater sample and control sample at low temperature (0oC). The volume of the sample was 0.025 l uniformly and thus when the international unit of mass (ng) was adopted, the concentration of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene was determined using the following formula: M=C×V Where C represents the concentration of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene obtained using the calibration equation of the system, and V represents the volume of the sample, that is, 0.025 l. The complete experiment was carried out 2 times repeatedly following the same procedures as mentioned above. The concentrations of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene at 8 time points in seawater sample and control sample were then averaged at room temperature respectively and similarly for the seawater sample and the control sample
HAN Dongqiang et al. / Chinese Journal of Analytical Chemistry, 2006, 34(10): 1361–1365
at low temperature (0oC). Finally, sketch maps of the mass-time of BTEX were made respectively (refer to Fig.3 and Fig.4). The concentration change of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene in the control sample is ascribed to the volatility of BTEX because pure water
basically contains no microorganism. Therefore, the biodegradation effect of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene can be obtained by comparing the concentration of the seawater sample with the concentration of the control sample at the same time point.
Fig. 3 The concentration change of BTEX in seawater sample and control sample at room temperature a: control sample; b: seawater sample
Fig. 4 The concentration change of BTEX in seawater sample and control sample at 0℃ a: control sample; b: seawater sample
The concentration of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene in the seawater sample at room temperature descends apparently and the falling trend can be classified into three periods. The concentration of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene declines slowly in 2 days (basically at the same level) but falls quickly from 2 days to 8 days(the biodegradation effect is evident). After 8 days, it is difficult to detect benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene. On the contrary, the trend of the concentration change of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene in the seawater
sample is basically in accord with that in the control sample at low temperature(0oC) and thus the biodegradation of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene by microorganism is faint. The volatilization, degradation, and the total percentage concentration change of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene at room temperature at special time point (8 days) refers to Table 1.
4
Conclusions We conclude with the following comments. Firstly, the
HAN Dongqiang et al. / Chinese Journal of Analytical Chemistry, 2006, 34(10): 1361–1365
biodegradation of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene are similar to each other in seawater sample at room temperature, that is, the concentration of BTEX remains well within 2 days and most of these are degraded by microorganism at 8 days. However, the concentration of BTEX remains well all the time and the biodegradation is faint in the seawater sample at low temperature (0ºC). Secondly, the biodegradation of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene is predominant when compared with the volatilization in the seawater sample at room temperature. However, the biodegradation and the volatilization of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene are not very evident in the seawater sample at low temperature (0ºC). Therefore, the biodegradation process of benzene, toluene, ethylbenzene, p,m-xylene, and o-xylene
depends on the temperature. Thirdly, the seawater sample must be measured immediately or saved at low temperature when collected in the geochemical investigation for oil and gas, which uses BTEX as the indicating substance. In conclusion, the concentration of benzene, toluene, ethylbenzene, p,m-xylene and o-xylene changes slowly within 2 days but most of these are biodegraded and volatilized at 8 days in seawater at room temperature, which attracts attention to the collection, transportation, preservation, and measurement of samples. Whether the sample is measured immediately or not affects the correlation between the abnormity of BTEX and the oil or gas field seriously. Therefore, to ensure the precision of the measurement results, it is better to measure the samples on the spot when collected.
Table 1 The volatilization, degradation, and total percentage change of BTEX concentration at room temperature at special time points (8 days) Component
Benzene (%)
Toluene (%)
Ethylbenzene (%)
p, m -Xylene (%)
Volatilization
16.48
15.91
18.13
17.35
9.91
Degradation
81.32
82.29
75.86
76.36
85.86
Total
97.80
98.20
93.99
93.71
95.77
Acknowledgements
[9]
o-Xylene (%)
Fu J M, Sheng G Y, Chen Y, Wang X M, Min Y S, Li S H, Chen L Y, Wang Z S. Climatic and Environmental Research, 1997, 2(1):
The authors express their thanks to Lab assistants Liang Xiao, Li Hongyan for their efforts in seawater samples’ measurement and postgraduate Yin Hailian for her thoughtful English translation discussions.
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