ABSTRACT
Background Tricresyl phosphate (TCP) is mainly used as a flame retardant. Studies have confirmed that it has cytotoxicity and neurotoxicity, but its reproductive toxicity is not clear. Objective To investigate the reproductive toxicity and potential mechanism of TCP subacute exposure on Caenorhabditis elegans. Methods Caenorhabditis elegans were exposed to solvent control and 0.1, 1, 10, 100, and 1000 μg·L−1 TCP respectively for 72 h. Brood size and number of fertilized eggs in the uterus were detected to evaluate reproductive ability. The number of total germline cells and the relative area of gonad arm were measured to evaluate the development of gonads. The body length and body width of Caenorhabditis elegans were detected to evaluate growth and development. The activities of reactive oxygen species (ROS) and superoxide dismutase (SOD) in Caenorhabditis elegans, and the mitochondrial active oxygen metabolism genes (mev-1 and gas-1) of N2 nematodes were detected by real-time fluorescence quantitative polymerase chain reaction (qRT-PCR) to evaluate oxidative stress. WS1433 transgenic nematodes and wild-type nematodes N2 were exposed to solvent control or TCP (0.1, 1, 10, 100, and 1000 μg·L−1) respectively. DNA damage in germ cells of WS1433 transgenic nematodes was detected, the relative expressions of DNA damage-related genes (hus-1, clk-2, cep-1, and egl-1) in N2 nematodes were detected by qRT-PCR to evaluate the effect of TCP exposure on genetic damage. Results Compared with the solvent control group (217.00 ± 12.20), the brood size of N2 nematodes in the 100 μg·L−1 and 1000 μg·L−1 TCP groups decreased (170.80 ± 11.51, 169.60 ± 10.52, P < 0.05). Compared with the solvent control group (18.43 ± 1.69), the number of fertilized eggs of N2 nematodes in the 100 μg·L−1 and 1000 μg·L−1 TCP groups decreased (13.47 ± 0.81, 11.95 ± 0.90, P < 0.05). Compared with the solvent control group (312.46 ± 77.4), the number of total germline cells of N2 nematodes in the 100 μg·L−1 and 1000 μg·L−1 TCP groups decreased (281.80 ± 12.98, 273.50 ± 8.53, P < 0.05). Compared with the solvent control group, the relative area of gonads of N2 nematodes in the 100 μg·L−1 and 1000 μg·L−1 TCP groups decreased by 13.83% and 17.25% respectively (P<0.05). Compared with the solvent control group [(1058.10±80.12) μm, (78.21±14.69) μm], the body length and body width of N2 nematodes in the 100 μg·L−1 and 1000 μg·L−1 TCP groups decreased (P<0.05). Compared with the solvent control group, the relative fluorescence intensity of ROS in nematodes in the 10, 100, and 1000 μg·L−1 TCP groups increased significantly (107.60%±1.02%, 105.90%±1.40%, and 106.40%±1.85%, respectively, P<0.05), and the activities of SOD were reduced (by 20.66%, 15.88%, and 16.44%, respectively, P<0.05). Compared with the solvent control group (1.3±1.3), the number of DNA-damaged germ cells of WS1433 nematodes in the 100 and 1000 μg·L−1 TCP groups increased significantly (2.4±0.3, 2.7±0.3, P<0.05); the expressions of mev-1 and gas-1 genes in N2 nematodes in the 10, 100 and 1000 μg·L−1 TCP groups decreased significantly (P<0.05); the expressions of hus-1 in the 0.1-1000 μg·L−1 TCP groups significantly increased (P<0.05); the expressions of clk-2 and egl-1 in the 100 and 1000 μg·L−1 TCP groups increased significantly (P<0.05); the expressions of cep-1 in the 1, 10, and 100 μg·L−1 TCP groups increased significantly (P<0.05). Conclusion TCP may cause reproductive damage to nematodes through oxidative stress and germ cell DNA damage.
ABSTRACT
A method was developed for determination of 12 kinds of phosphate compounds in water and sediment by high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) coupled with solid phase extraction (SPE) and ultrasonic extraction.The water samples were concentrated by HLB solid-phase extraction (SPE) column and eluted twice with ethyl acetate, ultrasonic solvent extraction for sediment samples and then repeated the operation of water samples after diluted with deionized water.The sample were separated on a ZORBAX Eclipse Plus C18 (150 mm × 2.1 mm, 3.5 μm) column by a gradient elution with 0.2% formic acid aqueous solution and methanol as the mobile phase.Ion mode analysis was monitored by high performance liquid chromatography mass spectrometer (MRM).The target compounds were quantified by external standard method.At the spiked levels (0.05, 0.1 and 0.5 μg/L), the average recoveries of 12 kinds of OPEs in water samples ranged from 66.4% to 115%, except for TMP (28.5%-47.8%) and TEHP (22.4%-73.8%).The relative standard deviation RSD (n=3) was 0.5%-9.09%, and the method quantification (MOQ) was 0.001-0.05 μg/L, However at the spiked levels of 5, 10 and 50 μg/kg, the average recoveries of 12 kinds of OPEs in sediment samples ranged from 65.4% to 120.0%, except for TMP (35.7%-44.9%) and TCEP (31.2%-48.9%).The relative standard deviation RSD (n=3) was 0.01%-9.54%, and the MOQ for sediment was 0.02-2.0 μg/kg dw.Based on the above methods, the detection and analysis of the targets in the water and sediments samples of Taihu Lake were carried out.The results showed that the concentrations of ΣOPEs were 0.1-1.7 μg/L and 8.1-420 μg/(kg dw), respectively.
ABSTRACT
A simple method was developed for simultaneous determination of seven urinary metabolites of organophosphate esters by liquid chromatography-tandem mass spectrometry (LC-MS / MS). Based on different physical and chemical properties of these OPs metabolites, the solid phase extraction cartridges and the washing and eluting solvents were optimized in details. Furthermore, the mobile phase and mass spectrometric parameters were also investigated. The results showed that Oasis WAX cartridge was the best SPE column in this study, and 2 mL of NH3 ·H2 O (5% ) in methanol and 2 mL of methanol were chosen as the eluting solvents. The recoveries of six analytes were ranged from 60. 5% to 104. 0% , whereas DEP ranged from 17. 8% to 36. 2% . Seven analytes could be baseline separated from each other under the optimized chromatographic conditions. The limits of detection and quantification of seven analytes ranged from 0. 005 to 0. 2 μg / L and 0. 02 to 0. 5 μg / L, respectively. The standard deviations of response repeatability for intra-day and inter-day period were lower than 15. 4% . This method was finally applied to determination of metabolites of OPs from 10 urines from general population in Guangzhou city. The concentrations of total OPs metabolites in urine samples ranged from 0. 5 to 6. 7 μg / L.
ABSTRACT
A gel permeation chromatography ( GPC ) coupled with solid phase extraction and gas chromatography-mass spectrometric ( GPC-SPE-GC/MS ) method was developed to analyze seven kinds of organophosphate esters ( OPEs ) , including tri-n-butylphosphate, tri ( 2-chloroethyl ) phosphate, triphenyl phosphate, tris ( 2-butoxyethyl ) phosphate, tri-o-tolylphosphate, tri-m-tolylphosphate, and tri-p-tolylphosphate) in human serum. The recoveries of cleanup methods between GPC-silica/alumina column and H2 SO4-silica/sulfuric acid gel column were compared. The purification method with the GPC-silica/alumina column didn’t destroy the structure of organophosphate esters ( OPEs ) and could effectively remove some protein and lipid matrix influence in serum. The developed method was verified using the spiked blank and the spiked serum, the good recoveries and reproducibilities were achieved. The recoveries of all of OPEs in spiked blank (n=3) were all more than 75%. The recoveries of d12-TCEP and d15-TPhP in human serum samples (n=9) were 86. 3%±21. 6% and 103. 1%±16. 5%, respectively. In human serum samples (n=9), the detection ratios for TnBP, TCEP, TPhP, TBEP and m-TTP were more than 90% in all of the serum samples, p-TTP was only 30%, o-TTP was not detectable. The concentrations of TnBP, TCEP, TPhP, TBEP and m-TTP in serum were 3. 4-46. 5 ng/g lipid, 248. 6-958. 2 ng/g lipid, n. d. -4. 2 ng/g lipid, n. d. -49. 9 ng/g lipid and n. d.-23. 1 ng/g lipid, respectively.
ABSTRACT
Background contamination is a major problem in the analysis of organophosphate esters (OPEs). In this study, the possible sources of OPEs pollution were screened and several different ways were applied to minimize the blank contamination. Under the strict quality control measures, the cleanup efficiency of different solid phase extraction (SPE) was investigated for OPEs in different environmental matrices. A method was developed for the detection of 7 OPEs in dust, soil and sediment samples by gas chromatograph coupled with mass spectrometry ( GC / MS). Target compounds were extracted by hexane:dichloromethane (1 : 1, V/ V) followed by aminopropyl silica gel SPE column cleanup for dust, and target compounds in soil and sediment were Soxhlet extracted and cleanuped by two-step SPE. The results showed that the aminopropyl silica gel SPE column displayed the best purification performance among the three employed columns. Instrumental detection limits among the 7 OPEs ranged from 2. 5 to 25. 8 μg / L, and the method limits of quantification (MLOQs) in dust and soil sample ranged from 1. 4 to 15. 7 ng / g and 0. 3 to 2. 9 ng / g, respectively. The average recoveries of 7 OPEs in different matrices ( dust and soil) at two spiked concentration levels ranged from 67. 9% to 117. 4% . The proposed method was successfully applied to detect OPEs in different environmental matrices collected in Shanghai.