The intracellular concentrations of fluoroquinolones determined the antibiotic resistance response of Escherichia coli.

The extensive consumption of antibiotics has been reported to significantly promote the generation of antibiotic resistance (ABR), however, a quantitative causal relationship between antibiotic exposure and ABR response is absent. This study aimed to pinpoint the accurate regulatory concentration of fluoroquinolones (FQs) and to understand the biochemical mechanism of the mutual action between FQ exposure and FQ resistance response. Highly sensitive analytical methods were developed by using UPLC-MS/MS to determine the total residual, extracellular residual, total intracellular, intracellular residual and intracellular degraded concentration of three representative FQs, including ciprofloxacin (CIP), ofloxacin (OFL) and norfloxacin (NOR), with detection limits in the range of 0.002-0.057 µg/L, and recoveries in the range of 80-93%. The MICs of Escherichia coli (E. coli) were 7.0-31.4-fold of the respective MIC0 after 40-day FQ exposure, and significant negative associations were discovered between the intracellular (residual, degraded or the sum) FQ concentrations and FQ resistance. Transcriptional expression and whole-genome sequencing results indicated that reduced membrane permeability and enhanced multi-drug efflux pumps contributed to the decreasing intracellular concentration. These results unveiled the pivotal role of intracellular concentration in triggering FQ resistance, providing important information to understand the dose-response relationship between FQ exposure and FQ resistance response, and ascertain the target dose metric of FQs for eliminating FQ resistance crisis.

Uncovering an Emerging Group of Halogenated Nucleobase-Derived Disinfection Byproducts in Drinking Water: Prioritization of the Highly Cytotoxic 2-Chloroadenine.

Constant efforts have been devoted to exploring new disinfection byproducts in drinking water causally related to adverse health outcomes. In this study, five halogenated nucleobases were identified as emerging disinfection byproducts in drinking water, including 5-chlorouracil, 6-chlorouracil, 2-chloroadenine, 6-chloroguanine, and 5-bromouracil. We developed a solid phase extraction-ultraperformance liquid chromatography-tandem mass spectrometry method with the limits of detection (LOD) and recoveries ranging between 0.04-0.86 ng/L and 54-93%, respectively. The detection frequency of the five halogenated nucleobases ranged from 73 to 100% with a maximum concentration of up to 65.3 ng/L in the representative drinking water samples. The cytotoxicity of the five identified halogenated nucleobases in Chinese hamster ovary (CHO-K1) cells varied with great disparity, in which the cytotoxicity of 2-chloroadenine (IC50 = 9.4 µM) is appropriately three times higher than emerging DBP 2,6-dichloro-1,4-benzoquinone (IC50 = 42.4 µM), indicating the significant toxicological risk of halogenated nucleobase-DBPs. To the best of our knowledge, this study reports the analytical method, occurrence, and toxicity of halogenated nucleobase-DBPs for the first time. These findings will provide a theoretical basis for further research on probing the relationship between its mutagenicity and human health risk.

Chlorinated nucleotides and analogs as potential disinfection byproducts in drinking water.

Identification of emerging disinfection byproducts (DBPs) of health relevance is important to uncover the health risk of drinking water observed in epidemiology studies. In this study, mutagenic chlorinated nucleotides were proposed as potential DBPs in drinking water, and the formation and transformation pathways of these DBPs in chlorination of nucleotides were carefully investigated. A total of eleven chlorinated nucleotides and analogs were provisionally identified as potential DBPs, such as monochloro uridine/cytidine/adenosine acid and dichloro cytidine acid, and the formation mechanisms involved chlorination, decarbonization, hydrolysis, oxidation and decarboxylation. The active sites of nucleotides that reacted with chlorine were on the aromatic heterocyclic rings of nucleobases, and the carbon among the two nitrogen atoms in the nucleobases tended to be transformed into carboxyl group or be eliminated, further forming ring-opening or reorganization products. Approximately 0.2-4.0 % (mol/mol) of these chlorinated nucleotides and analogs finally decomposed to small-molecule aliphatic DBPs, primarily including haloacetic acids, trichloromethane, and trichloroacetaldehyde. Eight intermediates, particularly chlorinated imino-D-ribose and imino-D-ribose, were tentatively identified in chlorination of uridine. This study provides the first set of preliminary evidence for indicating the promising occurrence of chlorinated nucleotides and analogs as potential toxicological-relevant DBPs after disinfection of drinking water.

Methoxylated Modification of Glutathione-Mediated Metabolism of Halobenzoquinones In Vivo and In Vitro.

Xenobiotics were generally detoxified in organisms through interaction with endogenous molecules, which may also generate metabolites of increased toxicity. Halobenzoquinones (HBQs), a group of highly toxic emerging disinfection byproducts (DBPs), can be metabolized by reacting with glutathione (GSH) to form various glutathionylated conjugates (SG-HBQs). In this study, the cytotoxicity of HBQs in CHO-K1 cells showed a wavy curve as a function of increased GSH dosage, which was inconsistent with the commonly recognized progressive detoxification curve. We hypothesized that the formation and cytotoxicity of GSH-mediated HBQ metabolites contribute to the unusual wave-shaped cytotoxicity curve. Results showed that glutathionyl-methoxyl HBQs (SG-MeO-HBQs) were identified to be the primary metabolites significantly correlated with the unusual cytotoxicity variation of HBQs. The formation pathway was initiated by stepwise metabolism via hydroxylation and glutathionylation to produce detoxified hydroxyl HBQs (OH-HBQs) and SG-HBQs, followed by methylation to generate SG-MeO-HBQs of potentiated toxicity. To further verify the occurrence of the aforementioned metabolism in vivo, SG-HBQs and SG-MeO-HBQs were detected in the liver, kidney, spleen, testis, bladder, and feces of HBQ-exposed mice, with the highest concentration quantified in the liver. The present study supported that the co-occurrence of metabolism can be antagonistic, which enhanced our understanding of the toxicity and metabolic mechanism of HBQs.

Fabrication of ionic liquid-mesoporous silica/platinum electrode with high hydroelectric stability for electric-field-assisted particle separation.

Surface chemistry of electrodes plays a critical role in the fields of electrochemistry and electric-field-assisted separation. In this study, making ingenious use of the ordered mesoporous structure of silica materials and the electrochemical stability of ionic liquids (ILs) when integrated with polyvinylpyrrolidone (PVP), the PVP-modified IL-mesoporous silica/platinum wire (Pt/PVP@meso-SiO2@IL) was fabricated to increase hydroelectric stability and avoid the problem of electrode polarization. The effect of different amounts of mesoporous silica material used to modify the surface of the Pt electrode was systematically investigated. As a result, we successfully obtained a highly ordered mesoporous Pt/PVP@meso-SiO2 material with smooth surface. Because pentyl triethylamine bis(trifluoromethylsulfonyl) imide exhibits a wide electrochemical window between -3 to 3 V, this IL was chosen to modify mesopores under vacuum. Even after repeatedly applying electric field on Pt/PVP@meso-SiO2@IL 100 times, this working electrode remained stable and showed high hydroelectric stability. After verifying the feasibility of this method, it was successfully applied in the electric-field-assisted separation of 2.0 and 3.0 µm polystyrene particles without any impediment from electrode polarization problems. This work provides a brand-new insight for resolving the problem of electrode polarization by developing a versatile tool for the electroseparation of micro-objects.

Fast removal of phenolic compounds from water using hierarchical porous carbon nanofibers membrane.

Water pollution is becoming increasingly serious and seriously endangering human health, especially the direct emissions of phenolic compounds. An integrated sample pre-treatment and derivatization method based on a biopolymers/TEOS-based carbon nanofibers microextraction that allows rapid extraction (5 min), followed by separation and highly sensitive detection of phenolic compounds by gas chromatography‒mass spectrometry (GC-MS) analysis, is described. The developed methodology, coupled with GC-MS, allowed low detection limits (0.03-0.32 ng mL‒1), good linearities (0.5-200 ng mL‒1) and recoveries (73.58-85.76%) to be achieved in a few steps and short time. Based on the high adsorption properties of materials, the on-line removal device of phenolic compounds in water was designed. The elimination rate of phenolic compounds in water was higher than 73.58% at 1 mL min‒1 and three cycles. The elimination strategy for the phenolic compounds is very versatile and is easy to apply.

NLow matrix effect pretreatment method based on gas-liquid micro-extraction technique for determining multi-class pesticides in crops.

A low matrix effect pretreatment method by Gas-liquid microextraction technique (GLME) combined with ultrasound assisted extraction (UAE-GLME) has been developed for fast analysis of 41 multi-class pesticides in different crop samples. Extraction and cleanup procedures were completed within 20 min, and the impact of matrix effects in potato, lentil, corn and soybean matrices were systematically evaluated. As high as 97.6% pesticides treated by UAE-GLME experienced low matrix effect, and up to 99% pesticides except parathion-methyl in lentil samples were categorized as medium matrix effect. Good trueness (recoveries of 61.3-115.8%) and precision (RSD≤20%) were demonstrated by the UAE-GLME method, with MLOQs ranged from 0.005 to 0.02 mg kg-1. This method detected 2-phenylphenol, pyrimethanil, triadimefon and tebuconazole in the range of 0.008-0.012 mg kg-1 in real food samples. The multiresidue analysis feature of GLME has been validated, which displays further potential for high-throughput determination of organic pollutants in foods with distinctive properties.

Halohydroxybenzonitriles as a new group of halogenated aromatic DBPs in drinking water: Are they of comparable risk to halonitrophenols?

Halogenated aromatic disinfection byproducts (DBPs) exhibited similar total organic halogen levels in chlorinated drinking water samples as compared with aliphatic ones, and they predominantly accounted for the overall toxicity of the samples. Among the reported halogenated aromatic DBPs, halonitrophenols (HNPs) have received particular attention in recent years due to the relatively high risk in drinking water. In this study, a new group of halogenated aromatic DBPs were detected and then proposed to be halohydroxybenzonitriles (HHBNs) by employing the ultra-performance liquid chromatography/tandem mass spectrometers. Thereafter, the specific HHBN species in drinking water were theoretically speculated and then thoroughly identified with standard compounds. Their occurrence in drinking water was investigated, their cytotoxicity was evaluated, and their stability in the presence of chlorine was assessed. Seven newly identified HHBNs, including 3,5-dichloro-4-hydroxybenzonitrile, 3,5-dichloro-2-hydroxybenzonitrile, 5-bromo-3-chloro-4-hydroxybenzonitrile, 5-bromo-3-chloro-2-hydroxybenzonitrile, 3,5-dibromo-4-hydroxybenzonitrile, 3,5-dibromo-2-hydroxybenzonitrile, and 3,5-diiodo-4-hydroxybenzonitrile, showed 100% detection frequency in the collected drinking water samples with concentrations up to 36 ng/L. HHBNs exhibited significantly higher cytotoxicity in Chinese hamster ovary cells than regulated DBPs (e.g., trihalomethanes and haloacetic acids), which might be contributed by their cellular uptake efficiency and nucleophilicity. The seven HHBNs were proved to undergo transformation during chlorination following pseudo-first-order decay with half-lives in the range of 9-63 h. More importantly, in comparison to HNPs, which showed relatively high toxicity and strong stability among the halogenated aromatic DBPs, HHBNs presented comparable concentration-cytotoxicity contribution (50%) and slightly weaker stability (43%), suggesting that HHBNs should be a new group of DBPs of concern in drinking water.

Halonaphthoquinones: A group of emerging disinfection byproducts of high toxicity in drinking water.

Aromatic halogenated disinfection byproducts (DBPs) have received particular attention in recent years due to their high toxicity. However, most relevant researches at present focused merely on halo-monocyclic DBPs, while halo-polycyclic DBPs were scarcely explored. In this study, a new group of halo-bicyclic DBPs termed as halonaphthoquinones (HNQs) was systematically studied. By coupling with vacuum centrifugal concentrator, a SPE-UPLC-MS/MS method with high accuracy and sensitivity was developed to detect five semi-volatile HNQs in drinking water, which achieved the detection limits in the range of 0.05-0.24 ng/L. Five HNQs were identified using this method with 100% detection frequency at concentrations up to 136.7 ng/L in drinking water originated from seven water treatment plants. The cytotoxicity of the five tested HNQs in CHO-K1 cells (IC50 from 3.17 to 13.18 µM) was comparable to the most toxic known carbonaceous DBP in drinking water, iodoacetic acid (IC50=2.95 µM). Meanwhile, the cytotoxicity of five tested HNQs were also higher than 2,6-dichloro-1,4-benzoquinone (IC50=21.73 µM) which is hundreds to thousands of times more toxic than regulated DBPs, indicating the significant toxicity risk of HNQ DBPs. To the best of our knowledge, this study presents the first analytical method for analysis of HNQ DBPs, and the first set of data on the occurrence and cytotoxicity of HNQ DBPs in drinking water. These findings are meaningful for probing deeply into the presence of varied halo-polycyclic DBPs in the aqueous environment.

Formation and Cytotoxicity of Halophenylacetamides: A New Group of Nitrogenous Aromatic Halogenated Disinfection Byproducts in Drinking Water.

Nitrogenous aromatic halogenated disinfection byproducts (DBPs) in drinking water have received considerable attention recently owing to their relatively high toxicity. In this study, a new group of nitrogenous aromatic halogenated disinfection byproducts, halophenylacetamides (HPAcAms), were successfully identified for the first time in both the laboratory experiments and realistic drinking water. The formation mechanism of HPAcAms during chlorination of phenylalanine in the presence of Br- and I-, occurrence frequencies, and concentrations in authentic drinking water were investigated, and a quantitative structure-activity relationship (QSAR) model was developed based on the acquired cytotoxicity data. The results demonstrated that HPAcAms could be formed from phenylalanine in chlorination via electrophilic substitution, decarboxylation, hydrochloric acid elimination, and hydrolysis. The HPAcAm yields from phenylalanine were significantly affected by contact time, pH, chlorine dose, and temperature. Nine HPAcAms with concentrations in the range of 0.02-1.54 ng/L were detected in authentic drinking water samples. Most tested HPAcAms showed significantly higher cytotoxicity compared with dichloroacetamide, which is the most abundant aliphatic haloacetamide DBP. The QSAR model demonstrated that the cellular uptake efficiency and the polarized distributions of electrons of HPAcAms play essential roles in their cytotoxicity mechanisms.

Analytical characterization of nucleotides and their concentration variation in drinking water treatment process.

Nucleotides, as the basic building blocks of nucleic acids, widely exist in aqueous environment. In this study, we developed a solid phase extraction-high performance liquid chromatography-tandem mass spectrometry (SPE-UPLC-MS/MS) method for the analysis of 5'-adenosine monophosphate (AMP), 5'-uridine monophosphate (UMP), 5'-cytidine monophosphate (CMP) and 5'-guanosine monophosphate (GMP). The method achieved limits of detection (LODs) of 0.1-1.0 ng/L, and recoveries of 85-95% for the four tested nucleotides. The occurrence and concentrations of the four nucleotides in water from eight representative drinking water treatment and distribution systems in China were determined using this method. All four nucleotides were detectable in water treatment plant (WTP) influent and effluent, at concentrations of up to 30 ng/L and with occurrence frequency of around 90%. The concentrations of identified nucleotides increased 3-10 times after 10 km of water age in the water distribution system. Biological filters and coagulation increased the concentrations of nucleotides, conversely, active carbon, ozonation, and ultrafiltration membrane removed nucleotides in water. The effects of active carbon and coagulation were further confirmed using laboratory-controlled experiment. In addition, monochlorinated nucleotides were identified as the chlorination products of nucleotides.

Tracing historical changes, degradation, and original sources of airborne polycyclic aromatic hydrocarbons (PAHs) in Jilin Province, China, by Abies holophylla and Pinus tabuliformis needle leaves.

Due to their wide distribution and availability, plant leaves can be considered interesting candidates as biomonitoring substrates for the evaluation of atmospheric pollution. In addition, some species can also retain historical information, for example, related to environmental pollution, due to their leaf class age. In this study, the content of polycyclic aromatic hydrocarbons (PAHs) in Abies holophylla and Pinus tabuliformis needle samples in the function of their class age has been investigated to obtain information regarding the degradation constant for each PAH under investigation (α values ranging from 0.173 to 1.870) and to evaluate the possibility to correlate the presence of PAHs in needles with some important pollution environmental factors. Considering air pollutant variables registered in Jilin Province, interesting correlations (at 95% confidence level) have been found between coal consumption per year and anthracene contents in needles, while fluorene, phenanthrene, and anthracene results correlated with coal consumption. Furthermore, it has been demonstrated that the total PAH concentration in needles, for both species, increased with their age (from 804 to 3604 ng g-1 dry weight), showing a general tendency to accumulate these substances through years. PAH degradation rates increased instead with molecular complexity. This study could be considered a first trial to obtain historical environmental information by pine needles biomonitoring.

One-step integrated sample pretreatment technique by gas-liquid microextraction (GLME) to determine multi-class pesticide residues in plant-derived foods.

Gas-liquid microextraction technique (GLME) has been integrated with dispersive solid phase extraction to establish a one-step sample pretreatment approach for rapid analysis of multi-class pesticides in different plant-derived foods. A 50 µL of organic solvent plus 40 mg of PSA were required throughout the 5-minute pretreatment procedure. Good trueness (recoveries of 67.2 - 105.4%) and precision (RSD ≤ 18.9%) were demonstrated by the one-step GLME method, with MLOQs ranged from 0.001 to 0.011 mg kg-1. As high as 93.6% pesticides experienced low matrix effect through this method, and the overall matrix effects (ME%) were generally better or comparable to QuEChERS. This method successfully quantified 2-phenylphenol, quintozene, bifenthrin and permethrin in the range of 0.001 - 0.008 mg kg-1 in real food samples. The multiresidue analysis feature of GLME has been validated, which displays further potential for on-site determination of organic pollutants in order to safeguard food safety and human health.

Carbon Nanofibers-Based Nanoconfined Liquid Phase Filtration for the Rapid Removal of Chlorinated Pesticides from Ginseng Extracts.

A rapid nanoconfined liquid phase filtration system (NLPF) based on solvent-confined carbon nanofibers/carbon fiber materials (CNFs/CFs) was proposed to effectively remove chlorinated pesticides from ginsenosides-containing ginseng extracts. A series of major parameters that may affect the separation performance of the CNFs-NLPF method were extensively investigated, including the water solubility of nanoconfined solvents, filtration rate, ethanol content of the ginseng extracts, and reusability of the material for repeated adsorption. The developed method showed a high removal efficiency of pesticides (85.5-97.5%), high retainment rate of ginsenosides (95.4-98.9%), and consistent reproducibility (RSD < 11.8%). Furthermore, the feasibility of the CNFs-NLPF technique to be scaled-up for industrial application was systematically explored by analyzing large-volume ginseng extract (1 L), which also verified its excellent modifiable characteristic. This filtration method exhibits promising potential as a practical tool for removing pesticide residues and other organic pollutants in food samples to assure food quality and safeguard human health.

Simple and rapid analysis of phthalate esters in marine sediment using ultrasound-assisted extraction combined with gas purge microsyringe extraction followed by GC-MS.

Phthalate esters (PAEs) are a class of the emerging pollutants that pose a potential environmental threat to marine ecosystems. In this study, a simple analytical method using ultrasound-assisted extraction combined with gas purges microsyringe extraction (GP-MSE) coupled with GC-MS was utilized for the reliable and rapid determination of PAEs in different types of marine sediment. The analytical results showed that the method exhibited excellent reproducibility, linear responses, and detection limits, which verified the suitability of the method for the determination of PAEs in marine sediment. This approach requires minimal reagents, solvents, and sample pretreatment procedures as well as a short analysis time; thus, procedural blanks can be kept to a minimum. This method was demonstrated to be a highly efficient and sensitive quantitative analytical method for the simple detection of PAEs in marine sediment.

Simultaneous determination of multiple phytohormones in tomato by ionic liquid-functionalized carbon fibers-based solid-phase microextraction coupled with liquid chromatography-mass spectrometry.

Phytohormones are interrelated by synergistic or antagonistic crosstalk and play important roles in the regulation of plant growth and development. In order to understand the interaction between phytohormones in the plant physiological network, it is necessary to determine trace levels of multiple phytohormones simultaneously in a complex matrix. Here, we synthesized ionic liquids containing different functional groups and modified the surface of carbon fibers with them. Based on these carbon fibers-ionic liquid (CFs-IL) materials, a solid phase microextraction method was developed to enable the simultaneous extraction of phytohormones. The adsorption specificity of multiple phytohormones was studied by identifying the hydrophobic, electrostatic, and π-π interactions, as well as hydrogen bonds, which favor simultaneous extraction of the relevant acidic, alkaline and neutral phytohormones by improving compatibility. The proposed method, coupled with liquid chromatography-tandem mass spectrometry, was applied to the simultaneous determination of 13 acidic, alkaline and neutral phytohormones in tomato. The limits of quantification were found to be in the range of 0.32-54.05 ng mL-1 and 4.6-185.8 pg mL-1, respectively, when measured by QQQ and Q-TOF. All of the relative recoveries were in the range of 94.40-113.37% with RSDs ≤15.36% (n = 3) for spiked tomato samples. This method is expected to be widely applied to multiple phytohormones analysis for in-depth researches concerning the physiological networks of plants.

A fast and selective gas liquid microextraction of semiochemicals for quantitative analysis in plants.

A trapping-based gas liquid microextraction (GLME) method coupled with gas chromatography-mass spectrometry (GC-MS) was utilized to qualitatively and quantitatively characterize semiochemicals in plants. The main GLME extraction efficiency associated parameters (heating temperature and extraction time) were optimized. The results obtained from GLME process were compared with those of steam distillation and ultrasonic extraction, and the recovery, peak number and reproducibility were evaluated by using Thuja koraiensis Nakai as a representative plant. Furthermore, the quantitative performances of the GLME in terms of sample amount, recoveries of spiked standards and correlation were systematically evaluated using standard addition method, which gave a good quantitative ability for all the compounds with squares of correlation coefficient (r2) of higher than 0.99. Finally, the contents of α-pinene, camphene, linalool, α-terpinenol, ß-caryophyllene, α-caryophyllene, and totarol in Thuja koraiensis Nakai samples were quantified, and their concentrations (SD, n = 3) were; 0.65 (0.06), 0.62 (0.05), 4.12 (0.15), 0.99 (0.08), 1.11 (0.07), 0.63 (0.04), and 21.91 (0.25) µg g-1, respectively. It was demonstrated that GLME is a powerful sample preparation technique for quantitative and qualitative analysis of plant semiochemicals.

Open-tubular radially cyclical electric field-flow fractionation (OTR-CyElFFF): an online concentric distribution strategy for simultaneous separation of microparticles.

An open-tubular radially cyclical electric field-flow fractionation technique which achieves the online separation of microparticles in a functional annular channel is proposed in this study. The system was set up by using a stainless steel tube and a platinum wire modified with ionic liquid/mesoporous silica materials as the external and internal electrodes. The feasibility for online separation of various particles was experimentally demonstrated. Particles in the channel were affected by a radial electric field and field-flow fractionation (FFF). On the cross section, different particles showed distinctive migration distances depending on their own properties and the different magnitudes of forces being exerted. The same kind of particles form an annular distribution within the same annulus while different particles form annular distributions at varied concentric annuli through electrophoresis. Under a laminar flow of FFF, different sizes of particles formed a conical arrangement within the annular separation channel. With the joint influence of electric field and flow field, different trajectories were obtained and the particles were eventually separated. Voltage, frequency and duty cycle value are the main parameters affecting the separation of particles. By adjusting these parameters, particles migrate in a zigzag trajectory on one side of the electrodes (mode I) and reach both sides of the electrodes (mode II). Six polystyrene particles were completely separated with high resolution within several minutes. Our system offers numerous advantages of label-free, high-resolution and online separation without tedious operations, and it is a promising tool for the effective separation of various micro-objects.

A traceless clean-up method coupled with gas chromatography and mass spectrometry for analyzing polycyclic aromatic hydrocarbons in complex plant leaf matrices.

This study developed a traceless clean-up method by combining solid phase extraction (SPE) with gas purge-microsyringe extraction (GP-MSE) to purify sample extracts for the determination of polycyclic aromatic hydrocarbons (PAHs) in plant leaves. SPE exhibited good purification performance for the removal of polar lipids, while the GP-MSE technique effectively eliminated less-volatile lipids hence realizing zero damage to the instrument, and significantly improved the peak tailings. After ultrasonic extraction, the combined two-step clean-up procedure successfully removed over 99% of lipids from nineteen types of tree leaves, and PAHs in tree leaves were determined by GC-MS. The relative standard deviations (RSDs) for intra-day (n = 3) and inter-day (n = 3) analyses of PAHs in spiked willow samples were in the range of 0.8%-12.1% and 4.7%-15.3%, respectively. The recoveries of PAHs from spiked willow extracts ranged from 74 to 90%, with an average of 86%. The method detection limit (MDL) of PAHs in tree leaves ranged from 0.1 to 4.9 ng g-1 dry weight. In conclusion, the clean-up method in this study realized the analysis of PAHs in plant leaves with high accuracy, sensitivity and reproducibility. Most importantly, the two-step purification method significantly minimizes damage to the GC-MS system particularly to the column and ion source, which is beneficial to ensure continuous analysis of a large number of samples with good performance.