Drugs In Urine Samples Research Articles

Silane–based modified papers and their extractive phase roles in a microfluidic platform

Herein, some (modified) paper–based substrates were prepared and utilized as extractive phases in a microfluidic device and their extraction performances examined for analytes with different polarities. Reagents including hexadecyltrimethoxysilane (HDTMS), phenyltrimethoxysilane (PTES), (3-aminopropyl) triethoxysilane (APTES) and 3–(2,3–epoxypropoxy) propyltrimethoxysilane (EPPTMOS) were implemented for the modification process. Due to the induction of different silane functional groups, it was anticipated to have various interactions for the tested analytes. Eventually, the prepared paper sheets were used as extractive phases for solid–phase extraction within a microfluidic system. The microfluidic setup not only improves the mechanical stability of the paper–based phases but also facilitated the mass transfer process and decreases the energy consumption. The selected analytes contained antidepressant drugs, organophosphates and triazine toxins, having different structures and polarities. Afterward, the existing interactions between the paper–based sorbents and the selected analytes were investigated by Principal Component Analysis (PCA) and Hierarchical Clustering Analysis (HCA) for data analysis. According to the obtained data, the PTES modified surface paper was selected for quantitative investigation and optimization. The major parameters associated with the extraction performance were studied. A linear range was obtained for amitriptyline and trimipramine, 0.05–200 μg L−1, and the calibration graph for clomipramine was linear in the range of 0.02–200 μg L−1. The employment of microfluidic device in conjunction with gas chromatography-mass spectrometry led to the limits of detections of 0.005–0.01 μgL−1 for the antidepressant drugs. The relative recoveries for the tested drugs in urine samples were in the range of 95–103%.

Preparation, characterization and evaluation of hydrophilic polymers containing magnetic nanoparticles and amine-modified carbon nanotubes for the determination of anti-inflammatory drugs in urine samples

Hydrophilic solids based on poly(2-hydroxyethyl methacrylate) (pHEMA) with embedded magnetic nanoparticles and amine-modified carbon nanotubes were synthesized by photopolymerization. For this purpose, an oil in water (O/W) emulsion with an aqueous/oil ratio of 60/40 was prepared where the polymerization reaction occurred in the aqueous phase due to the hydrophilicity of pHEMA and the selected nanoparticles. Variables affecting the stability and emulsion formation as well as the initiation and propagation of the polymerization were studied. The morphology of the obtained magnetic solids was characterized by SEM/EDAX in order to show the differences in presence and absence of nanoparticles within the structure. Finally, the synergic effect of both magnetic and carbon nanoparticles in the sorbent capacity of the final hydrophilic solids was evaluated through the determination of non-steroidal anti-inflammatory drugs (NSAIDs) in human urine samples. HPLC-UV was used as instrumental technique and detection limits ranged from 5 to 10 μg L−1. The precision was calculated both intra- and inter-solids (same and different synthesis batches) obtaining satisfactory RSD values of less than 13%, which indicated the robustness of the synthesis and the extraction procedure. Finally, a study with real and fortified urine samples was also carried out obtaining recovery values between 86% and 109% for target NSAIDs.

Preconcentration and Determination of Psychotropic Drugs in Urine Samples by Ion Mobility Spectrometry with Electrospray Ionization Coupling On-Line Single-Drop Liquid-Liquid-Liquid Microextraction

A method for analysis of psychotropic drugs in urine is investigated using a novel single-drop liquid-liquid-liquid microextraction (SDLLLME) apparatus as an electrospray emitter in ion mobility spectrometry (IMS). In this method, ketamine and pethidine are back-extracted into the acceptor phase (water and methanol) from the water and methanol immiscible organic phase. Sensitivity of extraction is improved as it does not require further methanol-adding procedure. Factors affecting the extraction of psychotropic drugs are characterized, including organic solvent type, extraction time, and concentration of NaOH/Ac in the donor/acceptor phase. The best extraction sensitivity is obtained with 600 μL dodecane as the organic phase, 30 minutes extraction time, and 10 mL donor phase with 0.01 M sodium hydroxide (NaOH) and 3 μL acceptor phase with 0.5 M acetic acid (Ac). Using this method, the two analytes can be extracted and analyzed simultaneously, showing this method is valuable for field application.

Graphene oxide-Fe3O4 nanocomposite magnetic solid phase extraction followed by UHPLC-MS/MS for highly sensitive determination of eight psychoactive drugs in urine samples

Graphene oxide–Fe3O4 (GO–Fe3O4) nanocomposite was synthesized by a facile chemical co-precipitation method. The GO–Fe3O4 was used as magnetic sorbent to extract the eight psychoactive drugs from urine samples. The analytes are morphine (MOR), 6-monoacetylmorphine (6-MAM), amphetamine (AMP), methamphetamine (MAMP), codeine, cocaine, dolantin and benzoylecgonine (BZE), which were determined by ultrahigh performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). This method has high selectivity for the target analytes. The limit of detection (LOD) and limit of quantification (LOQ) were 0.02–0.2 μg L−1 and 0.05–0.5 μg L−1, respectively. The Mandel's fitting test revealed good linearity within all linear ranges. The linear ranges were calculated as 0.05–1000 μg L−1 for AMP, MAMP, cocaine and dolantin; 0.1–1000 μg L−1 for 6-MAM and codein; and 0.5–1000 μg L−1 for MOR and BZE. The recoveries ranged in 80.4–105.5%. The intra-day and inter-day RSDs are in the range of 2.7–13.1% and 3.9–13.7%, respectively. Magnetic solid phase extraction (MSPE) with GO–Fe3O4 provides a convenient, rapid and green sample pretreatment method for extracting the target psychoactive drugs from urine. This methodology can be used for simultaneous or individual detection of eight major psychoactive drugs with high sensitivity. This method has high potential in clinical and forensic areas for psychoactive drugs analysis.

Cardiovascular Medication Stability in Urine for Non-Adherence Screening by LC-MS-MS.

Biochemical testing in urine is a powerful new tool in the investigation of non-adherence to cardiovascular medications Drug testing using liquid chromatography tandem mass spectrometry (LC-MS-MS) is the mainstay of the laboratory test but may be subject to pre-analytical factors that could impact on test results. The stability of cardiovascular medications in urine is one such factor that has not been fully explored in non-adherence testing and has the potential to result in patients appearing falsely non-adherent to their therapy. The stability of 29 cardiovascular medications in patients' urine samples were assessed at room temperature (RT) and at -80°C using a LC-MS-MS screening method. All drugs and drug metabolites were found to be stable under the storage conditions studied. The findings imply that the medication stability in urine samples does not have any impact on non-adherence results and thus allowing samples to be taken and transported without the need for specialist sample handling procedures. The stability of cardiovascular drugs in urine samples will allow adherence testing to be utilized more widely into routine clinics and research.

Magnetic Solid Phase Extraction Based on Modified Magnetite Nanoparticles Coupled with Dispersive Liquid–Liquid Microextraction as an Efficient Method for Simultaneous Extraction of Hydrophobic and Hydrophilic Drugs

In this work, surfactant-coated Fe3O4@decanoic acid nanoparticles was synthesized as a viable nanosorbent for coextraction of drugs with different polarities (hydrophobic, hydrophilic). To reach desirable enrichment factors, efficient clean-up and low limits of detection (LODs), the method was combined with dispersive liquid–liquid microextraction (DLLME). The coupling of these extraction methods with GC-FID detection was applied to simultaneous extraction and quantification of venlafaxine (VLF) as a hydrophilic model drug and desipramine (DESI) and clomipramine (CLO) as hydrophobic model drugs in urine samples. The effect of sample pH, nanosorbent amount, sorption time, surfactant concentration, eluent type, eluent volume, salt content, elution time in magnetic solid phase extraction step and extraction solvent and its volume along with sample pH in DLLME step were optimized. Under the selected conditions, linearity was achieved within the range of 5–5000 µg L−1. The LOD values were obtained in the range of 1.5–3.0 µg L−1 for DESI, 1.2–2.5 µg L−1 for VLF and 2.0–4.0 µg L−1 for CLO, respectively. The percent of extraction recoveries and relative standard deviations (n = 5) were in the range of 82.4–95.9 and 6.1 for DESI, 60.5–92.8 and 6.9 for VLF and 57.2–58.0 and 5.5 for CLO, respectively. Ultimately, the applicability of the new method was successfully confirmed by the extraction and quantification of DESI, VLF and CLO from human urine samples.

Sensitive determination of psychotropic drugs in urine samples using continuous liquid-phase microextraction with an extraction solvent lighter than water

A novel extraction vessel was employed, for the first time, in continuous liquid-phase microextraction (CLPME) with an extraction solvent lighter than water for the extraction of psychotropic drugs from urine samples.

Magnetic solid phase extraction of rizatriptan in human urine samples prior to its spectrofluorimetric determination

A method has been developed and validated for the determination of rizatriptan drug in human urine samples. Magnetic solid phase extraction (MSPE) utilizing a molecularly imprinted nanoadsorbent was used as the sample preparation method for selective extraction of the drug molecules from the urine matrix prior to the spectrofluorometric determination of the drug at λem=365nm (λex=280nm). The molecularly imprinted polymer (MIP) technique was used to improve the nanosorbent selectivity toward the drug molecules. The size, morphology, and structure of the synthesized MIP adsorbent were characterized by transmission electronic microscope, X-ray diffractometer, vibrating sample magnetometer, and FT-IR spectrometer. The nanoadsorbent was utilized to develop an MSPE method for selective extraction of the drug molecules in human urine samples prior to spectrofluorometric determination. The results showed that utilization of the MSPE step could provide a sensitive (limit of detection of 1.1ngmL−1), precise (coefficients of variation <2.21%), and accurate (recoveries >97%) method for the spectrofluorometric determination rizatriptan drug in urine samples under the optimized conditions (pH, 8.0; adsorbent amount, 0.02g; sorption time, 15min; eluent, methanol: 1.0molL−1 acetic acid (1:1v/v); desorption time, 10min).

UHPLC–MS/MS method with sample dilution to test therapeutic adherence through quantification of ten antihypertensive drugs in urine samples

Nowadays, hypertension represents an important health problem, particularly in developed countries. In some cases the standard therapeutic approaches are not able to reestablish the normal blood pressure values: this condition is called “resistant hypertension”. However, a fraction of cases of resistant hypertension are actually due to poor adherence to the prescribed therapy. Therapeutic Drug Monitoring could represent a direct and useful tool to correctly identify non-compliant patients. Nevertheless, high throughput methods for the simultaneous monitoring of a wide panel of drugs in the same analysis are lacking and, furthermore, there is not a generally acknowledged “standard” matrix for this test (plasma or urine).In this work, we validated a UHPLC–MS/MS assay to quantify ten among the most used antihypertensive agents in urine samples, covering all the current classes: amlodipine, atenolol, clonidine, chlortalidone, doxazosin, hydrochlorothiazide, nifedipine, olmesartan, ramipril and telmisartan.Both standards and quality controls were prepared in urine matrix. Only 100μL of each sample were added with 40μL of internal standard and 860μL of water:acetonitrile 90:10, acidified with 0.05% formic acid. Chromatographic separation was performed on an Acquity® UPLC HSS T3 1.8μm 2.1×150mm column, with a gradient of water and acetonitrile, both added with 0.05% formic acid.Accuracy, intra-day and inter-day precision fitted FDA guidelines for all analytes, while matrix effects resulted reproducible among different urine lots. Method performances were tested on urine samples from hypertensive patients with good results. This simple analytical method could represent a useful tool for the management of antihypertensive therapy.

Simultaneous determination of opioid drugs in urine with high‐performance liquid chromatography–ultraviolet after supramolecular based magnetic NP solid‐phase extraction

A simple and rapid extraction/preconcentration procedure using supramolecular based magnetic nanoparticle (MNP) solid-phase extraction method by sodium dodecyl sulphate (SDS)-coated MNPs has been developed for both removal and preconcentration of three important opioid drugs named morphine, codeine and methadone before their high-performance liquid chromatography–ultraviolet determination. To get insight configuration of SDS molecules on the surface of the MNPs, zeta potential measurements were performed in different SDS (mg)/mg of MNPs ratios. Parameters affecting the extraction and preconcentration efficiency of the opioids were optimised as recovery by one variable at-a-time methodology and the obtained recoveries were 91, 94 and 98% for morphine, codeine and methadone, respectively. Under optimum conditions (such as pH = 3.5; adsorbent amount = 60 mg; eluent solvent volume = 2 ml of acetonitrile, sample volume = 200 ml etc.), the method was linear in the range of 1–200 ng ml−1 with enrichment factors (EFs) of 91, 94 and 98 for morphine, codeine and methadone, respectively, and the limits of detection were <0.27 ng ml−1. Finally, the proposed method was successfully applied for both removal and trace determination of the targeted drugs in urine samples.

Carrier assisted electromembrane extraction based on nonionic lipophilic surfactants for the determination of basic drugs in urine samples

Herein, an efficient carrier assisted electromembrane extraction coupled with capillary electrophoresis has been proposed for the quantitation of basic drugs (pseudoephedrine and tramadol) in urine samples.

Magnetic field assisted μ-solid phase extraction of anti-inflammatory and loop diuretic drugs by modified polybutylene terephthalate nanofibers

A magnetic nanocomposite consisting of nanoparticles–polybutylene terephthalate (MNPs–PBT) was electrospun and used as an extracting medium for an on-line μ-solid phase extraction (μ–SPE)–high performance liquid chromatography (HPLC) set–up with an ultraviolet (UV) detection system. Due to the magnetic property of the prepared nanofibers, the whole extraction procedure was implemented under an external magnetic field to enhance the extraction efficiencies. The developed method along with the synthesized nanocomposite were found to be appropriate for the determination of trace levels of selected drugs including furosemide, naproxen, diclofenac and clobetasol propionate in the urine sample. The prepared MNPs-PBT electrospun nanocomposite was characterized using the scanning electron microscopy (SEM), energy dispersive spectroscopy (EDX) and Fourier transform infrared (FT–IR) spectroscopy. The prepared magnetic fibers showed high porosity, which was another driving force for the extraction efficiency enhancement. Major parameters affecting the extraction efficiency of the selected drugs were optimized. The limits of detections (LOD) of the studied drugs were in the range of 0.4–1.6 μg L−1 and the limits of quantification (LOQ) were 1–4 μg L−1 under the optimized conditions. Relative standard deviation (RSD%) for three replicates at three concentration levels of 6, 100 and 400 μg L−1 were 5.9–8.0% while acceptable linear range with two orders of magnitude was obtained (R2 = 0.99). The method was validated by the determination of the selected drugs in urine samples and the results indicated that this method has sufficient potential for enrichment and determination of the desired drugs in the urine sample. The relative recovery values were found to be in the range of 78–91%. Implementing the developed on–line μ–SPE method under the external magnetic field induction, led to higher extraction efficiencies for the selected drugs with various diamagnetic properties.

LC of high to moderately polar basic drugs in urine with water and detergent, and direct injection.

Micellar LC was first proposed as a 'green' mode using mobile phases of water and surfactant. However, in most procedures a small amount of organic solvent is required to decrease the retention to convenient values. Results & methodology: Mixed micellar mobile phases prepared with both cationic (sodium dodecyl sulphate) and nonionic surfactant (Brij-35) modulate the retention of high to moderately polar basic drugs to practical times, eliminating the need of organic solvent. While the mobile phase is continuously recycled through the system, the stationary phase performance is maintained after repetitive injection of the samples. Through an extensive validation, the approach is shown to be appropriate to determine these drugs in urine samples without previous pretreatment.

Application of electromembrane extraction followed by corona discharge ion mobility spectrometry analysis as a fast and sensitive technique for determination of tricyclic antidepressants in urine samples

In the paper, electromembrane extraction (EME) method coupled with corona discharge ion mobility spectrometry (CD-IMS) was introduced for determination of antidepressant drugs (desipramine, sertraline, clomipramine, citalopram) in urine samples. IMS is a well-known, comparatively inexpensive, robust and easy to operate analytical instrumentation. This combination would provide a selective and sensitive determination. In order to achieve the best extraction efficiency, optimization of the variables affecting these methods was carried out. Optimal extractions were accomplished with 2-nitrophenyl octyl ether as the supported liquid membrane, with 190V as the driving force, and with pH4 in donor and pH1 in acceptor solutions. Extractions were obtained after 30min of operation with the whole assembly agitated at 1000rpm.Under the optimized conditions, the proposed technique provided good linearity (>0.9974), repeatability (RSD<4.5%), low limits of detection (0.9–1.5ngmL−1), excellent preconcentration factor (PF=158–190) and high recoveries (79–95%). Finally, developed method was applied to quantification of antidepressant drugs in urine samples.

Water-compatible graphene oxide/molecularly imprinted polymer coated stir bar sorptive extraction of propranolol from urine samples followed by high performance liquid chromatography-ultraviolet detection

Due to the high selectivity and stability, molecularly imprinted polymers (MIPs) have been successfully applied in stir bar sorptive extraction (SBSE) as a special coating to improve the selective extraction capability for target analytes. However, traditional MIPs usually suffer from incompatibility in aqueous media and low adsorption capacity, which limit the application of MIP coated stir bar in aqueous samples. To solve these problems, a water-compatible graphene oxides (GO)/MIP composite coated stir bar was prepared in this work by in situ polymerization. The prepared water-compatible GO/MIP coated stir bar presented good mechanical strength and chemical stability, and its recognition ability in aqueous samples was improved due to the polymerization of MIP in water environment, the adsorption capacity for target analytes was also increased by the addition of GO in MIP pre-polymer solution. Based on it, a method of water-compatible GO/MIP coated stir bar sorptive extraction combined with high performance liquid chromatography-ultraviolet detector (HPLV-UV) was proposed for the analysis of propranolol (PRO) in aqueous solution. The influencing factors of SBSE, such as sample pH, salt effect, stirring rate, extraction time, desorption solvent and desorption time, were optimized, and the analytical performance of the developed SBSE-HPLC-UV method was evaluated under the optimized conditions. The limit of detection (LOD) of the proposed method for PRO was about 0.37μgL−1, and the enrichment factor (EF) was 59.7-fold (theoretical EF was 100-fold). The reproducibility was also investigated at concentrations of 5μgL−1 and the relative standard deviation (RSD) was found to be 7.3% (n=7). The proposed method of GO/MIP coating-SBSE-HPLC-UV was successfully applied for the assay of the interested PRO drug in urine samples, and further extended to the investigation of the excretion of the drugs by monitoring the variation of the concentration of PRO in urine within 10h after drug-taking.

Simultaneous determination of trace amounts of anti-hypertensive drugs in urine using magnetic mixed hemimicelles solid-phase extraction combined with HPLC-UV

Hypertension is the leading cause of morbidity and mortality in developed and developing countries. The combination of amlodipine and carvedilol is very effective for reducing blood pressure and improving treatment compliance. Determination of drugs in biological fluids is vital in therapeutic efficacy. In the present study, a simple, rapid and efficient magnetic solid phase extraction procedure based on mixed hemimicelles is described and validated for the simultaneous determination of trace amounts of anti-hypertensive drugs in urine samples using sodium dodecyl sulfate-coated magnetite nanoparticles as extractant. Various factors, which could affect the extraction efficiency were investigated and optimized. Under the optimum conditions, the limits of detection (S/N = 3) for amlodipine and carvedilol were 5 and 2 ng/mL, respectively. The relative standard deviation for six measurements of 50 ng/mL of amlodipine and carvedilol were 4.7 and 5.2%, respectively. The developed method was applied to the analysis of patients' urine samples, and satisfactory results were obtained in the range between 81.6 and 92.7%.

Combination of counter current salting-out homogenous liquid-liquid extraction and dispersive liquid-liquid microextraction as a novel microextraction of drugs in urine samples.

The counter current salting-out homogenous liquid-liquid extraction (CCSHLLE) joined with the dispersive liquid-liquid microextraction based on solidification of floating organic drop (DLLME-SFO) has been developed as a high preconcentration technique for the determination of different drugs in urine samples. Amphetamines were employed as model compounds to assess the extraction procedure and were determined by high performance liquid chromatography-ultraviolet detection (HPLC-UV). In this method, initially, NaCl as a separation reagent is filled into a small column and a mixture of urine and acetonitrile is passed through the column. By passing the mixture, NaCl is dissolved and the fine droplets of acetonitrile are formed due to salting-out effect. The produced droplets go up through the remained mixture and collect as a separated layer. Then, the collected acetonitrile is removed with a syringe and mixed with 30.0μL 1-undecanol (extraction solvent). In the second step, the 5.00mLK2CO3 solution (2% w/v) is rapidly injected into the above mixture placed in a test tube for further DLLME-SFO. Under the optimum conditions, calibration curves are linear in the range of 1-3000μgL(-1) and limit of detections (LODs) are in the range of 0.5-2μgL(-1). The extraction recoveries and enrichment factors ranged from 78 to 84% and 157 to 168, respectively. Repeatability (intra-day) and reproducibility (inter-day) of method based on seven replicate measurements of 100μgL(-1) of amphetamines were in the range of 3.5-4.5% and 4-5%, respectively. The method was successfully applied for the determination of amphetamines in the actual urine samples. The relative recoveries of urine samples spiked with amphetamine and methamphetamine are 90-108%.

Poly(2-aminobenzothiazole)-coated graphene oxide/magnetite nanoparticles composite as an efficient sorbent for determination of non-steroidal anti-inflammatory drugs in urine sample

In this study, for the first time, 2-aminobenzothiazole monomer was polymerized on Fe3O4 NPs, graphene oxide/Fe3O4 (GO/Fe3O4) and graphene/Fe3O4 (G/Fe3O4) nanocomposites. The synthesized magnetic nanosorbents were characterized by various techniques. The extraction ability of these nanosorbents including Fe3O4, GO/Fe3O4, G/Fe3O4, Fe3O4@poly(2-aminobenzothiazole) (Fe3O4@PABT), GO/Fe3O4@PABT and G/Fe3O4@PABT were compared for dispersive-micro-solid phase extraction of three non-steroidal anti-inflammatory drugs. The results revealed that GO/Fe3O4@PABT nanocomposite demonstrates higher extraction efficiency for naproxen, diclofenac and ibuprofen as selected model analytes. Following the sorption and elution steps, the model analytes were quantified by high performance liquid chromatography-photo diode array detection. Afterwards, a central composite design methodology combined with desirability function approach was applied to find out the optimal experimental conditions. Under the optimized conditions, the limits of detection and linear dynamic ranges were achieved in the range of 0.07-0.3μgL−1 and 0.25–2000μgL−1, respectively. The percent of extraction recovery was 87.4, 85.5 and 90.5% for naproxen, diclofenac and ibuprofen, respectively. The obtained relative standard deviation (n=5) was 7.2, 5.4 and 6.4% for naproxen, diclofenac and ibuprofen, respectively. Ultimately, this method was employed for urinary monitoring of the target analytes and satisfactory results were obtained.

Determination of Methadone in Human Urine Using Salting-Out Effect and Dispersive Liquid-Liquid Microextraction Followed by HPLC-UV

The salting-out effect combined with the dispersive liquid-liquid microextraction based on solidification of floating organic drop (DLLME-SFO) has been developed as a high preconcentration technique for the determination of drug in urine samples. Methadone was employed as model compound to assess the extraction procedure and were determined by high performance liquid chromatography-ultraviolet detection (HPLC-UV). In this method, initially, NaCl as a separation reagent is filled into a small column and a mixture of urine and acetonitrile is passed through the column. By passing the mixture, NaCl is dissolved and the fine droplets of acetonitrile are formed due to saltingout effect. The produced droplets go up through the remained mixture and collect as a separated layer. Then, undecanol (extraction solvent). In the second step, the 5.00 mL K2CO3 solution (2% w/v) is rapidly injected into the above mixture placed in a test tube for further DLLME-SFO. Under the optimum conditions, calibration curves are linear in the range of 2-2000 μg L-1 and limit of detection (LOD) is 0.7 μg L-1. the collected acetonitrile is removed with a syringe and mixed with 30.0 μL-1 .The extraction recovery and enrichment factor were 83% and 140%, respectively. Repeatability (intra-day) and reproducibility (inter-day) of method based on seven replicate measurements of 100 μg L-1 of methadone were 4.1% and 5.3%, respectively. The relative recoveries of urine samples spiked with methadone are 92%-106%.

Two-phase and three-phase liquid-phase microextraction of hydrochlorothiazide and triamterene in urine samples.

This paper reports the applicability of two-phase and three-phase hollow fiber based liquid-phase microextraction (HF-LPME) for the extraction of hydrochlorothiazide (HYD) and triamterene (TRM) from human urine. The HYD in two-phase HF-LPME is extracted from 24 mL of the aqueous sample into an organic phase with microliter volume located inside the pores and lumen of a polypropylene hollow fiber as acceptor phase, but the TRM in three-phase HF-LPME is extracted from aqueous donor phase to organic phase and then back-extracted to the aqueous acceptor phase, which can be directly injected into HPLC for analysis. Under optimized conditions preconcentration factors of HYD and TRM were obtained as 128 and 239, respectively. The calibration curves were linear (R(2) ≥ 0.995) in the concentration range of 1.0-100 µg/L for HYD and 2.0-100 µg/L for TRM. The limits of detection for HYD and TRM were 0.5 µg/L. The intra-day and inter-day RSD based on four replicates were obtained as ≤5.8 and ≤9.3%, respectively. The methods were successfully applied for determining the concentration of the drugs in urine samples. Copyright © 2015 John Wiley & Sons, Ltd.