Assessment of Polycyclic Aromatic Hydrocarbons (PAHs) in Seawater and Sediments, Human and Ecological Risks, Northern Coastline of Persian Gulf.

This study aimed to assess 13 PAHs concentrations and induced human and ecological risks in seawater and surficial sediments of eight estuaries in the northern coastline of the Persian Gulf, Iran, 2021. The range of Σ13 PAHs concentration was 0.24-8.83 µg L-l and 3.1-11.46 µg g-1 dry weight, and the mean value was 4.99 µg L-l and 6.06 µg g-1 dry weight in seawater and sediment, respectively. Two, three and four rings PAHs were dominant with 29.33% and 41.33% of ΣPAHs in seawater and sediment, respectively. The primary source of PAHs was both pyrolytic and petrogenic. Most PAHs' calculated health (DCR, HQs, TEF, MEF) and ecological risks (SQGs) values in seawater were in the moderate range or high-risk values for damage to the marine environment. It is concluded that the pollution of PAHs should be carefully considered, and the government should make a proper action plan to minimize the pollution.

Assessment of incremental lifetime cancer risks of ambient air PM10-bound PAHs in oil-rich cities of Iran.

This study investigates the concentrations of PM10-bound PAHs and their seasonal variations in three cities of Ahvaz, Abadan, and Asaluyeh in Iran. The mean concentrations of PM10 in two warm and cold seasons in Ahvaz were higher and in Abadan and Assaluyeh were lower than the national standard of Iran and the guidelines of the World Health Organization. The Σ16 PAHs concentration in ambient air PM10 during the cold season in Ahvaz, Abadan and Asaluyeh was 244.6, 633, and 909 ng m- 3, respectively, and during the warm season in Ahvaz, Abadan, and Asaluyeh was 242.1, 1570 and 251 ng m- 3, respectively. The high molecular weight PAHs were the most predominant components. The most abundant PAHs species were Pyr, Chr, B [ghi] P, and Flt. The results showed that the total PAHs concentration in the cold and warm seasons was dependent on industrial activities, particularly the neighboring petrochemical units of the city, vehicular exhausts, traffic and use of oil, gas, and coal in energy production. The total cancer risk values as a result of exposure to PAHs in ambient air PM10 in all three cities for children and adults and in both cold and warm seasons were between 1 × 10- 6 and 1 × 10- 4, and this indicates a potential carcinogenic risk. Therefore, considering the various sources of air pollutants and its role on people's health, decision makers should adopt appropriate policies on air quality to reduce the ambient air PAHs and to mitigate human exposure.

Metal-coded hydrogel magnetic molecularly imprinted polymer for preconcentration and cleanup of sarcosine: Determination in urine; coupled to on-column capillary electrophoresis.

In this study, sarcosine metal-coded hydrogel magnetic molecularly imprinted polymer (Hydro-MeC-MMIP) has been fabricated and coupled to on-column derivatization capillary electrophoresis (CE). As a metal-coding approach, sarcosine-Cu2+-ligand (Sar-Cu2+-L) chelate complex was introduced as a template to overcome the problems associated with the fabrication of MMIP for a small molecule having limited functional groups such as sarcosine. To our best knowledge, it is the first time that methacrylamide (MA) coated Fe3O4 (Fe3O4@MA) with abounded reactive double-bound on the surface has been used as a magnetic core in the one-pot synthesis of MMIPs. As prepared, Hydro-MeC-MMIP was characterized by different microscopic, spectroscopic, and thermal gravimetric methods. Hydro-MeC-MMIP was used to extract and preconcentrate sarcosine in the urine sample with no treatment and dilution. Sarcosine was quantified by on-column derivatization capillary electrophoresis equipped with a photodiode array detector. A mixture of thirteen amino acids was separated with a total run time of 12 min. Three structural analogs, including alanine, sarcosine, and glycine, were significantly resolved. Under optimal experimental conditions, the method's detection and quantification limits were 9.93 and 33.10 ng mL-1, respectively. The linear range of 50-2000 ng mL-1 and 96% recovery, along with the relative standard deviation of 6.07% (n = 6) for the target amino acid, were obtained. This method provides a simple, low-cost, fast, and efficient tool for extracting and quantifying sarcosine in the urine. The present method can address inconsistency in evaluating sarcosine as a candidate biomarker for prostate cancer with a simple CE/UV; no need for a sophisticated detection system such as a mass spectrometer.

Molecular characterization of Ribosomal DNA (ITS2) of hard ticks in Iran: understanding the conspecificity of Dermacentor marginatus and D. niveus.

OBJECTIVES: Hard ticks (Acari: Ixodidae) are ectoparasites of medical and veterinary importance. They are obligate blood-feeding vectors with the ability to transmit a wide variety of pathogens. Standard morphological keys are normally used for the identification of tick species. However, considering the importance of accurate species identification and the determination of bio-ecological characteristics of species, relying on morphological keys alone can be questionable. In this study, two DNA fragments (ITS2 and COI) were selected for phylogenetic evaluation of Iranian hard tick species belonging to the genera Dermacentor, Hyalomma, and Rhipicephalus. RESULTS: 1229 specimens of Dermacentor marginatus, D. niveus, Hyalomma anatolicum, Rhipicephalus bursa, and R. sanguineus s.l constituting 11 populations were collected from three different climatic and zoogeographical zones in Iran. Morphological studies revealed notable differences in important morphological characteristics between different populations of D. marginatus. The results of ITS2 sequence analysis provided additional evidence which supports the conspecificity of D. niveus and D. marginatus. Contrary to this finding, the sequence analysis of COI and phylogeny favored the separation of the two species. Given the greater importance of COI in identifying and discriminating species, a possibility heterospecificity between the two species should be considered.

Effect of chicken raw materials on physicochemical and microbiological properties of mechanically deboned chicken meat.

Excessive consumption of red meat is associated with various diseases including coronary heart diseases and cancer. Lower health-related problems of chicken meat, consumption of chicken meat, and mechanically deboned chicken meat (MDCM) have been increased due to their cheaper prices. Thereby, chemical, microbial, and physical causes of chicken meat losses and the safety aspects are needed to be fully considered to save food by improved application of chicken meat and its by-product. This study investigated the effects of chicken classes, layer, and broiler, and different carcass cuts, fillet, skeleton, and the whole carcass, on physicochemical, protein, fat, ash, moisture, pH, and peroxide, and microbiological, total plate counts, Escherichia coli, Staphylococcus aureus, Campylobacter, and Salmonella, characteristics as well as introducing content changes of metal elements iron, calcium (Ca), lead, cadmium, and arsenic in MDCM. The highest values of physicochemical characteristics, calcium, iron, and heavy metals were observed in deboned layer chicken carcass and deboned broiler skeleton (p<0.05). Although Escherichia coli was detected in all of the treatments, Salmonella, Staphylococcus aureus, and Campylobacter contamination were found only in the deboned layer and broiler skeleton. In conclusion, the application of MDCM by-products in meat products without thermal processing is not recommended. The broiler and layer skeleton MDCMs are not suitable for human consumption due to the high contents of heavy metals. However, the whole carcasses of layer chickens are suitable to be mutually used in MDCM at the end of the egg laying period.

Fabrication of UMCM-1 based monolithic and hollow fiber - Metal-organic framework deep eutectic solvents/molecularly imprinted polymers and their use in solid phase microextraction of phthalate esters in yogurt, water and edible oil by GC-FID.

In this study, for the first time, hollow fiber and monolithic fiber were fabricated based on metal-organic framework deep eutectic solvents/molecularly imprinted polymers (MOF- DES/MIPs) and were used for microextraction of phthalate esters under termed hollow fiber liquid membrane-protected solid-phase microextraction (HFLMP-SPME) followed by gas chromatography- flame ionization detection. Several parameters influencing extraction recoveries of phthalate esters including adsorption and desorption parameters were investigated and optimized using fabricated MOF- DES/MIPs monolithic fiber. Under optimal conditions, detection limits (S/N = 3) of the method were in a range of 0.008-0.03 µg L-1 and limits of quantification (S/N = 10) were between 0.028 and 0.12 µg L-1. RSD (%) for intra-day and inter-day precisions were between 2.4-4.7% and 2.6-3.4%, respectively. Subsequently, this procedure was successfully applied with satisfactory results in the determination of phthalate esters in yogurt, water, and soybean oil samples. The R (%) ranged from 95.5 to 100.0% in different samples.

PHBV/PLGA nanoparticles for enhanced delivery of 5-fluorouracil as promising treatment of colon cancer.

5-Fluorouracil (5-FU) is one of the most widely used agents in the first-line chemotherapy for colon cancer. However, clinical use of 5-FU is limited because of the low efficacy of drug uptake and systemic toxic effects. Therefore, there is a critical need to find better drug delivery systems in order to improve the efficacy of the drug. In the present study, we have developed a novel combination drug delivery system based on PHBV/PLGA NPs for delivery of 5-FU to cancer cells. NPs were prepared by the double emulsion method and their optimization of preparation was evaluated using Box-Behnken design (BBD) of response surface methodology (RSM). 5-FU loaded NPs were characterized by scanning electron microscope (SEM), differential scanning calorimetry (DSC), thermogravimetry analysis (TGA), and Fourier transformed infra-red spectroscopy (FT-IR). SEM image implied that NPs were spherical in shape and the results of DSC, TGA, and FT-IR suggest that 5-FU was encapsulated into NPs. The obtained results revealed that 5-FU loaded PHBV/PLGA NPs induced significant higher cell death at concentration much lower than free 5-FU. Results of hemolysis assay indicated that the NPs were hemo-compatible. In vivo anti-tumor studies showed that 5-FU loaded NPs reduced tumor volume significantly in comparison with free 5-FU. As the first example of using PHBV/PLGA as nano-drug delivery system with enhanced anti-tumor activities, this study establishes PHBV/PLGA as a novel promising drug delivery platform for treatment of colon cancer.

Anti-leishmanial activity and identification of chemical constituents of Scrophularia striata essential oil by head space solid phase micro extraction gas chromatography mass spectrometry (HS-SPME GC-MS) / Actividad anti-leishmanial e identificación de componentes químicos del aceite esencial de Scrophularia striata mediante microextracción en fase sólida en el espacio de cabeza seguido de cromatografía gaseosa acoplada a espectrometría de masas (HS-SPME GC-MS)

The aim of the present study was to determine the main constituents of Scrophularia striata essential oil and to evaluate in vitro effect of essential oil on Leishmania tropica and Leishmania major promastigotes and axenic amastigotes. Chemical constituents of the extracted essential oil were separated by headspace solid-phase microextraction (HS-SPME) equipped with a PDMS/DVB fiber. The fiber was injected to gas chromatogram- mass spectroscopy (GC-MS) to determine their identity. Finally, after exposure of parasites to different concentrations of water soluble fraction of essential oil, viability of promastigotes and axenic amastigotes were investigated. Based on the HS-SPME results, 47 compounds representing 95.6% of the total oil, were identified in essential oil. Essential oil analysis showed that nonane (19.7%), α-terpineol (17.4%) and linalool (10.2%) were the most abundant compounds. This study indicates that water soluble fraction of S. striata essential oil has promising anti-leishmanial activity.

El objetivo del presente estudio fue determinar los principales componentes del aceite esencial de Scrophularia striata y evaluar el efecto in vitro del aceite esencial en promastigotes y amastigotes axénicos de Leishmania tropica y Leishmania major. Los componentes químicos del aceite esencial extraído se separaron mediante microextracción de fase sólida en el espacio superior (HS-SPME) equipado con una fibra PDMS/DVB. Para determinar su identidad la fibra se inyectó en un cromatógrafo de gases acoplado un espectrómetro de masas (GC-MS). Finalmente, después de la exposición de los parásitos a diferentes concentraciones de fracción soluble del aceite esencial en agua, se investigó la viabilidad de los promastigotes y los amastigotes axénicos. En base a los resultados de HS-SPME, se identificaron 47 compuestos que representan el 95.6% del aceite total en el aceite esencial. El análisis de aceites esenciales mostró que el nonano (19.7%), el α-terpineol (17.4%) y el linalol (10.2%) fueron los compuestos más abundantes. Este estudio indica que la fracción soluble en agua del aceite esencial de S. striata tiene una actividad antileishmanial prometedora.

New folate receptor targeted nano liposomes for delivery of 5-fluorouracil to cancer cells: Strong implication for enhanced potency and safety.

We previously showed that folate liposomes of 5FU made from Dipalmitoylphosphatidylcholine (DPPC) induced cell death in HT-29 and HeLa cells more potently than bulk 5FU. Also, a primary 5FU liposomal formulation with phosphatidyl choline (PC) exhibited higher cytotoxicity in murine colon cancer cells. In the present study, optimization of 5FU PC liposome, mechanism of cell death induction in human cancer cell lines and its safety along with other assays have been employed for targeted PC liposomes of 5FU. Liposomes were prepared using thin layer method and optimization of preparation was assessed using central composite design (CCD) of response surface methodology (RSM). Folic acid (FA) was employed as the targeting ligand. Morphology of 5FU loaded liposomes and changes in their thermal behavior were assessed by transmission electron microscopy (TEM) and differential scanning calorimetry (DSC), respectively. In vitro cytotoxicity was explored using MTT assay in HT-29, Caco-2, HeLa and MCF-7 cell lines. Cytotoxicity mechanism of the targeted delivery system was searched through the evaluation of reactive oxygen species (ROS) overproduction, mitochondrial membrane potential (∆Ψm), the release of cytochrome c, the activity of caspase 3/7 and apoptosis and necrosis rate. Liposomes were spherical in shape and 5FU was successfully encapsulated into liposomes rather in an amorphous state. Our interesting results showed that in HT-29 cells targeted liposomes triggered the mitochondrial apoptotic pathway by decreasing the mitochondrial membrane potential, releasing of cytochrome c and promoting the substantial activity of caspase 3/7. In HeLa cells, however, targeted liposomes particularly activated necrosis pathway through the overproduction of ROS. Folate-liposomal 5FU showed significantly higher antitumor efficiency compared to free drug. The results of this study offer new prospects for cancer therapy with reducing systemic drug exposure and associated toxicities.

A nanocomposite consisting of graphene oxide, zeolite imidazolate framework 8, and a molecularly imprinted polymer for (multiple) fiber solid phase microextraction of sterol and steroid hormones prior to their quantitation by HPLC.

A method is described for multiple monolithic fiber solid-phase microextraction of five sterol and steroid hormones from complex food samples. A composite was prepared from graphene oxide, a metal-organic framework (ZIF-8) and a molecularly imprinted polymers was deposited on a single thin fiber. Four thin fibers were combined to obtain a fiber bundle. The nanocomposite was characterized by Fourier transform infrared spectroscopy, powder X-ray diffraction, scanning electron microscopy (SEM) and energy-dispersive X-ray spectroscopy. The parameters affecting the extraction were optimized by Box-Behnken design. Following elution with methanol, the analytes progesterone, testosterone, ß-sitosterol, cholesterol and campesterol were quantified via HPLC. Response is linear in the 0.01-1000 µg L-1 concentration range, and limits of detection range from 3 to 5 ng L-1. The method was successfully applied to the determination of the five analytes in spiked samples of white meat, egg yolks and vegetables. The relative mean recoveries ranged from 95.0% to 101.0%. Graphical abstract Schematic presentation of a monolith molecularly imprinting polymer (MIP) fiber fabrication for solid phase microextraction (SPME) of sterol and steroid hormones. The fiber was synthesis using graphene oxide and metal-organic framework, ZIF-8, composite by imprinting technique and progesterone as template. Four fibers were combined to obtain a fiber bundle. Then, multiple fiber solid-phase microextraction was employed for determination of analytes by coupling with HPLC/UV detection.

A mechanistic study of the effect of transferrin conjugation on cytotoxicity of targeted liposomes.

This study was performed to prepare 5-fluorouracil (5FU) containing targeted liposomes for the safety and efficacy enhancement. Liposomes were prepared using thin layer method and transferrin (Tf) was employed as the targeting ligand. Morphology of 5FU-loaded liposomes was assessed by transmission electron microscopy (TEM). The in vitro cytotoxicity was investigated via MTT assay on HT-29, CT26 and fibroblast cells. Mitochondrial membrane and cell death evaluations were also investigated. Resulted showed that the encapsulation efficiency (EE%) and particle size of the liposomes were 40.12% and 130 nm, respectively. TEM image implied that liposomes were spherical in shape. In cancer cells, targeted liposomes triggered the mitochondrial apoptotic pathway by lower production of reactive oxygen species (ROS) (63.58 vs 84.95 fluorescence intensity), reduced mitochondrial membrane potential and releasing of cytochrome c (68.66 vs 51.13 ng/mL). The results of this study indicated that Tf-targeted 5FU liposomes can be employed as promising nanocarrier for the delivery of drugs to cancer cells.

A novel 5-Fluorouracil targeted delivery to colon cancer using folic acid conjugated liposomes.

The aim of this study was to develop and characterize 5-Fluorouracil (5FU) containing targeted liposomes in order to enhance the efficacy and safety of the drug. Folic acid (FA) was used as a targeting ligand. The in vitro cytotoxicity of formulation against HT-29, Caco-2, CT26, HeLa and MCF-7 cell lines was evaluated using MTT assay. Mechanism of cell death induced by targeted liposomes was further investigated via the production of reactive oxygen species (ROS), change in mitochondrial membrane potential (ΔΨm), release of cytochrome c and activity of caspase 3/7. The in vivo tumor inhibition study was also performed after administration of drug and targeted 5FU liposome. The encapsulation efficiency (EE%) of the optimized formulation was 39.71%. Particle size of liposomes was around 174 nm and the nanoparticles were found to be spherical in shape. Differential Scanning Calorimetry (DSC) results indicated that the drug remained in an amorphous state in liposomes. According to the MTT results, targeted liposomes exhibited higher cytotoxicity than 5FU and liposomal 5FU. Targeted liposomes were found to trigger necrosis in HT-29 cells; while, in HeLa cells, targeted liposomes activated apoptotic pathway by collapse of ΔΨm, increased activity of cytochrome c as well as caspases activity. in vivo results showed that targeted liposomes reduced tumor volume significantly in comparison with 5FU (169.00 mm3 tumor volume vs 326.40 mm3). From these findings, it can be concluded that folic acid targeted liposomes may provide a new platform for selective delivery of drugs to cancer cells.

A novel micro-extraction strategy for extraction of bisphosphonates from biological fluids using zirconia nanoparticles coupled with spectrofluorimetry and high performance liquid chromatography.

Extraction of bisphosphonates from biological fluids is important and time consuming step in sample preparation procedure. This paper describes a simple and green sample preparation technique for dispersive micro solid phase extraction (DMSPE) of alendronate sodium (ALS) from urine and serum samples prior to direct spectrofluorimetry (DSFL) and high performance liquid chromatography with fluorescence detection (HPLC-FLD), respectively. The DMSPE strategy is based on the selective chemisorption of ALS on zirconia nanoparticles (ZNPs) as an adsorbent followed by derivatization of the eluted analyte using o-phthalaldehyde (OPA) in the presence of 2-mercaptoethanol (2ME) at basic medium to form fluorescent species. The chemical and instrumental influencing parameters on DMSPE and measuring methods were optimized for the efficient extraction and determination of ALS. The presented methods were capable of extracting ALS from human urine and serum samples and determining over the wide ranges of 5-1000 and 5-2500 µg L-1 with limits of detection (LOD) of 1.5 and 1.4 µg L-1 for DSFL and HPLC methods, respectively. The relative recoveries for the three spiked standard levels of ALS in urine and serum samples ranged from 89.0% to 107.0%, and the intra-day relative standard deviations (%RSDs) were in the range of 2.9-7.9%.

Folic acid-modified liposomal drug delivery strategy for tumor targeting of 5-fluorouracil.

The aim of this study was to develop a liposomal formulation to selectively target cancer cells. Liposomes were prepared using thin layer method and folic acid (FA) was applied for targeted delivery of 5FU to cancer cells. Liposomes prepared were characterized for encapsulation efficiency (EE%), morphology and their particle size. Cellular uptake, cytotoxicity study and ROS production were evaluated using CT26 cell line. Hemolysis test was performed on rat red blood cells (RBCs). Moreover, the efficacy of targeted liposomes were investigated by in vivo antitumor activity and tissue toxicities were studied by histological examination. The EE% and average particle size of liposomes were 67.88±1.84% and 114.00±4.58nm, respectively. TEM image revealed that liposomes were spherical in shape. Targeted liposomes showed higher cellular uptake, lower IC50 (12.02µM compared to 39.81µM for liposomal 5FU and 39.81µM for free 5FU) and higher ROS production than free drug (62,271.28 vs 2369.55 fluorescence intensity) on cancer cells. Results of hemolysis assay confirmed the blood biocompatibility of the liposomes. Moreover, folate targeted liposomes showed better tumor inhibition than free drug (88.75mm3 tumor volume vs 210.00mm3) and no tissue abnormalities were found in histological examination. It can be concluded that folate targeted liposomes provide an effective and safe strategy for colon cancer targeted chemotherapy.

Transferrin targeted liposomal 5-fluorouracil induced apoptosis via mitochondria signaling pathway in cancer cells.

The purpose of this study was to prepare transferrin (Tf) targeted liposomal 5-Fluorouracil (5FU) to improve the safety and efficacy of the drug. Liposomes were prepared using thin layer method. Morphology of liposomes was characterized by transmission electron microscopy (TEM) and their particle size was also determined. The in vitro cytotoxicity was investigated via MTT assay on HT-29 (as cancer cell) and fibroblast (as normal cell). Moreover, cytotoxicity mechanism of targeted liposomes was determined through the production of reactive oxygen species (ROS), mitochondrial membrane potential (∆Ψm) and release of cytochrome c. Results showed that encapsulation efficiency (EE%) was 58.66±0.58 and average size of liposomes was 107nm. Also, nano-particles were spherical as shown by TEM. MTT assay on HT-29 cells revealed the higher cytotoxic activity of targeted liposomes in comparison to free drug and non-targeted liposome. In contrast, comparing with cancer cells, targeted liposomes had no cytotoxic effect on normal cells. In addition, targeted liposomes induced apoptosis through activation of mitochondrial apoptosis pathways, as evidenced by decreased mitochondrial membrane potential and release of cytochrome c. Results of the study indicated that targeted liposomes would provide a potential strategy to treat colon cancer by inducing apoptosis via mitochondria signaling pathway with reducing dose of the drug and resulting fewer side-effects.

Preparation and Evaluation of Taste Masking Iron Suspension: Taking Advantage of Weak Cationic Exchange Resin.

The objective of this study is to use weak acid cation exchange resin to mask the taste and eliminate the teeth staining problems of ferrous sulfate liquid preparations for use in pediatrics. Amberlite IPR64 was loaded by Fe(II). Then, different suspensions of the iron-resin complex were prepared using various polymers as the suspending agents, and sorbitol as well as sucrose as sweeteners. Physical stability, rheological assessment, kinetics of Fe(II) release, and taste evaluation of suspensions were studied. The results implied that the prepared iron suspension was more stable when xanthan gum is used as the suspending agent. It was also shown that iron release in 0.7% NaCl (similar to saliva) was very negligible compared to relatively quick release in acidic medium (resembled the stomach). Overall, the volunteers confirmed that the formulations were successful in iron taste masking. The pH-dependent process of ion exchange by weak cationic resins opens an attractive approach to access a taste-masked iron suspension for infants.

Effects of coating layer and release medium on release profile from coated capsules with Eudragit FS 30D: an in vitro and in vivo study.

The aim of the present research was to evaluate the impact of coating layers on release profile from enteric coated dosage forms. Capsules were coated with Eudragit FS 30D using dipping method. The drug profile was evaluated in both phosphate buffer and Hank's solutions. Utilization X-ray imaging, gastrointestinal transmission of enteric coated capsules was traced in rats. According to the results, no release of the drug was found at pH 1.2, and the extent of release drug in pH 6.8 medium was decreased by adding the coating layers. The results indicated single-layer coated capsules in phosphate buffer were significantly higher than that in Hank's solution. However, no significant difference was observed from capsules with three coating layers in two different dissolution media. X-ray imaging showed that enteric coated capsules were intact in the stomach and in the small intestine, while disintegrated in the colon.

Urinary Trans, Trans-Muconic Acid is Not a Reliable Biomarker for Low-level Environmental and Occupational Benzene Exposures.

BACKGROUND: Benzene is a known occupational and environmental pollutant. Its urinary metabolite trans, trans-muconic acid (tt-MA) has been introduced by some environmental and occupational health regulatory associations as a biological index for the assessment of benzene exposure; however, recently, doubts have been raised about the specificity of tt-MA for low-level benzene exposures. In the present study, we investigated the association between urinary levels of tt-MA and inhalational exposure to benzene in different exposure groups. METHODS: Benzene exposure was assessed by personal air sampling. Collected benzene on charcoal tube was extracted by carbon disulfide and determined by a gas chromatograph (gas chromatography with a flame ionization detector). Urinary tt-MA was extracted by a strong anion-exchange column and determined with high-performance liquid chromatography-UV. RESULTS: Urinary levels of tt-MA in intensive benzene exposure groups (chemical workers and police officers) were significantly higher than other groups (urban and rural residents), but its levels in the last two groups with significant different exposure levels (mean = 0.081 ppm and 0.019 ppm, respectively) showed no significant difference (mean = 388 µg/g creatinine and 282 µg/g, respectively; p < 0.05). Before work shift, urine samples of workers and police officers showed a high amount of tt-MA and its levels in rural residents' samples were not zero. CONCLUSION: Our results suggest that tt-MA may not be a reliable biomarker for monitoring low-level (below 0.5 ppm) benzene exposures.