ABSTRACT
There is a significant rate of therapeutic failure in rheumatoid arthritis (RA) patients treated with leflunomide (LEF). This study investigates the utility values of teriflunomide levels (A77 1726) in identifying RA patients who remained with moderate or severe disease activity after the treatment with LEF. In this cross-sectional study, we compared: (a) RA patients who achieved a DAS28-ESR ≤ 3.2, and (b) RA patients who maintained a DAS28-ESR > 3.2 after treatment. ROC curves determined the cut-off of A77 1726 with the better performance to identify patients achieving a DAS28-ESR ≤ 3.2. Of the 115 patients treated with LEF, 69 (60%) remained with moderate/severe disease activity and 46 (40%) achieved low disease activity/remission. Higher A77 1726 levels showed a negative correlation with DAS28-ESR (r = - 0.42, p < 0.001) and other parameters of disease activity. We obtained the following utility values with the cut-off of A77 1726 > 10 µg/mL to identify RA patients who achieved a DAS28-ESR ≤ 3.2: sensitivity of 91.31%; specificity of 73.91%; positive predictive value of 70.00%; and negative predictive value of 92.73%. Serum A77 1726 discriminated between RA patients who remained with moderate/severe disease activity despite the treatment with LEF both as monotherapy and LEF as combo therapy.
Subject(s)
Antirheumatic Agents/therapeutic use , Arthritis, Rheumatoid/drug therapy , Crotonates/therapeutic use , Hydroxybutyrates/therapeutic use , Leflunomide/therapeutic use , Nitriles/therapeutic use , Toluidines/therapeutic use , Adult , Aged , Antirheumatic Agents/adverse effects , Antirheumatic Agents/blood , Arthritis, Rheumatoid/blood , Arthritis, Rheumatoid/diagnosis , Cross-Sectional Studies , Crotonates/adverse effects , Crotonates/blood , Drug Monitoring , Drug Therapy, Combination , Female , Humans , Hydroxybutyrates/adverse effects , Hydroxybutyrates/blood , Leflunomide/adverse effects , Leflunomide/blood , Male , Middle Aged , Nitriles/adverse effects , Nitriles/blood , Predictive Value of Tests , Remission Induction , Severity of Illness Index , Time Factors , Toluidines/adverse effects , Toluidines/blood , Treatment OutcomeABSTRACT
OBJECTIVE: Leflunomide is a commonly used disease-modifying drug in the treatment of rheumatoid arthritis (RA). Its effects are mediated via inhibition of dihydroorotate dehydrogenase (DHODH) by its active metabolite teriflunomide, and the pharmacokinetics of teriflunomide are highly variable. Our objective was to examine the association between the DHODH haplotype and plasma teriflunomide concentration with response to leflunomide in patients with RA where leflunomide was added to an existing disease-modifying drug regimen after failure to achieve an adequate response with conventional triple therapy. METHODS: Patients with RA who were taking, or were about to initiate, leflunomide were included. Participant characteristics, including the DHODH haplotype, were determined. Up to 5 plasma samples were collected after leflunomide was initiated for assays of total and free teriflunomide concentration. Disease activity was determined via the 28-joint Disease Activity Score (DAS28). The association between DAS28 scores and patient covariates was determined by linear mixed-effects modeling. RESULTS: A total of 67 patients were included in the study. The DAS28 score after initiation of leflunomide was associated with the baseline DAS28 score (ß = 0.70, P < 0.001) and was higher in those who carried the DHODH haplotype 2 (ß = 0.56. P = 0.01) and did not carry the shared epitope (ß = 0.56, P = 0.013). As total and free plasma teriflunomide concentration increased, the DAS28 score was significantly lower (P < 0.001 and P = 0.001, respectively). When considering threshold concentrations, teriflunomide concentrations >16 mg/liter were associated with a DAS28 score that was 0.33 lower, and when free teriflunomide concentration was >35 µg/liter, the DAS28 score was 0.32 lower. CONCLUSION: Teriflunomide concentration and carriage of the DHODH haplotype 2 are associated with response to leflunomide in patients with RA, and a total plasma teriflunomide concentration of at least 16 mg/liter is needed to maximize the likelihood of response.
Subject(s)
Arthritis, Rheumatoid/drug therapy , Crotonates/pharmacokinetics , Enzyme Inhibitors/pharmacokinetics , Hydroxybutyrates/pharmacokinetics , Immunosuppressive Agents/pharmacokinetics , Leflunomide/pharmacokinetics , Nitriles/pharmacokinetics , Oxidoreductases Acting on CH-CH Group Donors/antagonists & inhibitors , Oxidoreductases Acting on CH-CH Group Donors/genetics , Pharmacogenomic Variants , Polymorphism, Single Nucleotide , Toluidines/pharmacokinetics , Adult , Arthritis, Rheumatoid/diagnosis , Arthritis, Rheumatoid/genetics , Crotonates/blood , Dihydroorotate Dehydrogenase , Drug Monitoring , Enzyme Inhibitors/administration & dosage , Enzyme Inhibitors/blood , Female , Haplotypes , Humans , Hydroxybutyrates/blood , Immunosuppressive Agents/administration & dosage , Immunosuppressive Agents/blood , Leflunomide/administration & dosage , Leflunomide/blood , Male , Middle Aged , Nitriles/blood , Oxidoreductases Acting on CH-CH Group Donors/metabolism , Pharmacogenetics , Precision Medicine , Recovery of Function , Remission Induction , Toluidines/blood , Treatment OutcomeABSTRACT
Teriflunomide is an oral first-line disease modifying treatment (DMT) for patients with relapsing-remitting multiple sclerosis (RRMS). It can take up to two years to achieve systemic clearance of teriflunomide to an acceptable level, but this washout period may be accelerated by administration of cholestyramine. Relapse of multiple sclerosis (MS) during washout of teriflunomide or other first-line DMT is not as common. We report two patients with RRMS who experienced a relapse after the accelerated elimination period (AEP) of teriflunomide and confirmation of negative plasmatic levels (<0.02 µg/ml). In cases of risk of MS activity, we should not wait for teriflunomide negative plasmatic levels confirmation before starting the next DMT to reduce the risk of relapse.
Subject(s)
Crotonates/pharmacokinetics , Immunologic Factors/pharmacokinetics , Multiple Sclerosis, Relapsing-Remitting/drug therapy , Multiple Sclerosis, Relapsing-Remitting/physiopathology , Toluidines/pharmacokinetics , Adult , Anion Exchange Resins/administration & dosage , Cholestyramine Resin/administration & dosage , Crotonates/blood , Female , Humans , Hydroxybutyrates , Immunologic Factors/blood , Male , Nitriles , Recurrence , Toluidines/bloodABSTRACT
In the present study, a new graphene based nanofibers material (Polyacrylonitrile/Graphene Oxide (PAN/GO)) was used for microextraction by packed sorbent (MEPS). The PAN/GO nanofiber was synthesized using the electrospinning technique. MEPS online with liquid chromatography-tandem mass spectrometry (LC-MS/MS) was utilized for the extraction and determination of two local anesthetic drugs (lidocaine, prilocaine) and their major metabolites (2,6-xylidine, o-toluidine) in human plasma samples. Parameters affecting the extraction efficiency were investigated and optimized (including sample pH, washing solution and elution solution). The validation of the method was based on FDA (Food and Drug Administration) guidelines for bioanalytical methods. The calibration curve ranged from 2.00 to 2000â¯nmol/L for lidocaine and prilocaine, and from 10.0 to 2000â¯nmol/L for 2,6-xylidine and o-toluidine. The coefficient of determination (R2) values were 0.996, 0.995, 0.995, 0.996 (nâ¯=â¯3) for lidocaine, prilocaine, 2,6-xylidine and o-toluidine, respectively. The extraction recovery was 93.0% for lidocaine, 96.0% for prilocaine, 68.0% for 2,6-xylidine and 69.0% for o-toluidine. The limits of detection (LODs) were 0.25, 0.50, 2.50, 1.25â¯nmol/L for lidocaine, prilocaine, 2,6-xylidine and o-toluidine, respectively. The lower limits of quantification (LLOQs) were 2.0â¯nmol/L for lidocaine and prilocaine, and 10â¯nmol/L for 2,6-xylidine and o-toluidine, respectively. The accuracy values for the quality control (QC) samples were in the range of 91.0-111% for lidocaine, 92.0-118% for prilocaine, 84.0-98.0% for 2,6-xylidine and 82.0-90.0% for o-toluidine. The inter-day precisions for QC samples ranged from 7.0% to 11.8% for lidocaine, from 8.6% to 11.7% for prilocaine, from 8.0% to 10.0% for 2,6-xylidine and from 8.0% to 9.0% for o-toluidine. The matrix effect values were in the range of -2.3% to -8.6% for lidocaine, -2.7% to -10.2% for prilocaine, 4.8%-5.2% for 2, 6-xylidine and -8.2% to 9.4% for o-toluidine.
Subject(s)
Acrylic Resins/chemistry , Anesthetics, Local/blood , Aniline Compounds/blood , Graphite/chemistry , Nanofibers/chemistry , Solid Phase Microextraction/methods , Acrylic Resins/chemical synthesis , Chromatography, Liquid/instrumentation , Chromatography, Liquid/methods , Graphite/chemical synthesis , Humans , Hydrogen-Ion Concentration , Lidocaine/blood , Limit of Detection , Prilocaine/blood , Tandem Mass Spectrometry/methods , Toluidines/bloodABSTRACT
A simple high-performance liquid chromatography coupled with tandem mass spectrometry method was developed and fully validated to simultaneously determine teriflunomide (TER) and its metabolite 4-trifluoro-methylaniline oxanilic acid (4-TMOA) in human plasma and urine. Merely 50 µL plasma and 20 µL urine were employed in sample preparation using protein precipitation and direct dilution method, respectively. An Agilent Zorbax eclipse plus C18 column was selected to achieve rapid separation for TER and 4-TMOA within 3 min. Electrospray ionization under multiple reaction monitoring was used to monitor the ion transitions for TER (m/z 269.0 â 159.9), 4-TMOA (m/z 231.9 â 160.0), internal standard teriflunomide-d4 (m/z 273.0 â 164.0) and 2-amino-4-trifluoromethyl benzoic acid (m/z 203.8 â 120.1), operating in the negative ion mode. This method proved to have better accuracy and precision over concentration range of 10-5000 ng/mL in plasma as well as 10-10,000 ng/mL in urine. After a full validation, this method was successfully applied in a pharmacokinetic study of teriflunomide sodium and leflunomide in Chinese healthy volunteers.
Subject(s)
Chromatography, High Pressure Liquid/methods , Crotonates/blood , Crotonates/urine , Leflunomide/blood , Leflunomide/urine , Tandem Mass Spectrometry/methods , Toluidines/blood , Toluidines/urine , Crotonates/chemistry , Crotonates/pharmacokinetics , Drug Stability , Humans , Hydroxybutyrates , Leflunomide/chemistry , Leflunomide/pharmacokinetics , Limit of Detection , Linear Models , Nitriles , Reproducibility of Results , Toluidines/chemistry , Toluidines/pharmacokineticsABSTRACT
In this study, a reliable ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method coupled with an easy, fast and effective sample pretreatment procedure was developed for simultaneous determination of amitraz, chlordimeform, formetanate and their metabolites in human blood. With the procedures of protein precipitation and a phospholipid-removal step, the endogenous compound interference was significantly reduced, and matrix effects were significantly reduced. The linear ranges of matrix-matched standard curves were from 0.5 to 1000 ng/mL with coefficients of determination >0.996. Very low limits of detection (0.05-0.12 ng/mL) and limits of quantitation (0.15-0.4 ng/mL) were achieved. Reasonable recoveries ranging from 88.1 to 103.5% were obtained. The intra-day RSDs ranging from 3.2 to 8.6% and inter-day RSDs ranging from 4.8 to 9.2% indicated good precision. With the introduction of a phospholipid-removal step, the ME ranged from 90.1 to 98.5%. The established method was successfully applied to the analysis of a blood sample from a formetanate poisoning case. This method possesses the advantages of high sensitivity, reduced matrix effects and rapidity.
Subject(s)
Carbamates/blood , Chlorphenamidine/blood , Chromatography, High Pressure Liquid/methods , Mass Spectrometry/methods , Pesticides/blood , Toluidines/blood , Adult , Carbamates/chemistry , Carbamates/poisoning , Chlorphenamidine/chemistry , Chlorphenamidine/poisoning , Humans , Limit of Detection , Linear Models , Male , Pesticides/chemistry , Pesticides/poisoning , Phospholipids/chemistry , Phospholipids/isolation & purification , Reproducibility of Results , Toluidines/chemistrySubject(s)
Colesevelam Hydrochloride/administration & dosage , Colesevelam Hydrochloride/blood , Crotonates/administration & dosage , Crotonates/blood , Metabolic Clearance Rate/drug effects , Toluidines/administration & dosage , Toluidines/blood , Adult , Anticholesteremic Agents/administration & dosage , Anticholesteremic Agents/blood , Drug Administration Schedule , Female , Healthy Volunteers , Humans , Hydroxybutyrates , Male , Metabolic Clearance Rate/physiology , Nitriles , Treatment OutcomeABSTRACT
Leflunomide's active metabolite teriflunomide inhibits dihydro-oroate dehydrogenase, an enzyme essential to proliferation of T lymphocytes. As teriflunomide must reach the target site to have this effect, this study assessed the distribution of teriflunomide into T lymphocytes, as intracellular concentrations may be a superior response biomarker to plasma concentrations. CD3 MicroBeads (Miltenyi Biotec, Bergisch Gladbach, Germany) were used to extract CD3+ T cells from the peripheral blood of patients with rheumatoid arthritis who were taking a stable dose of leflunomide. Unbound plasma and intra-CD3+ T cell teriflunomide concentrations were quantified using liquid chromatography-mass spectrometry. Concentration (log transformed) and partition differences were assessed through paired Student t tests. Sixteen patients provided plasma steady-state teriflunomide samples, and eight provided a sample 6-12 weeks later. At time-point one, the geometric mean teriflunomide concentration (range) in CD3+ T cells was 18.12 µg/L (6.15-42.26 µg/L) compared with 69.75 µg/L (32.89-263.1 µg/L) unbound in plasma (P < 0.001). The mean partition coefficient (range) for unbound plasma teriflunomide into CD3+ T cells was 0.295 (0.092-0.632), which was significantly different from unity (P < 0.001). The median (range) change in teriflunomide concentration between the two time points was 14% (-10% to 40%) in unbound plasma and -29% (-69 to 138%) for CD3+ T cells. Because teriflunomide concentrations in CD3+ T cells were lower and displayed a higher intraindividual variability than the unbound plasma concentrations, its applicability as a therapeutic drug-monitoring marker may be limited.
Subject(s)
Antirheumatic Agents/blood , Arthritis, Rheumatoid/blood , CD3 Complex/immunology , Crotonates/blood , T-Lymphocytes/metabolism , Toluidines/blood , Aged , Arthritis, Rheumatoid/immunology , Chromatography, Liquid , Dose-Response Relationship, Drug , Female , Humans , Hydroxybutyrates , Male , Middle Aged , Nitriles , T-Lymphocytes/immunology , Tandem Mass SpectrometryABSTRACT
This paper describes the use of UPHLC-QTOF for the analysis of pesticide amitraz and its metabolites in human blood. UPHLC-QTOF working in full scan mode simultaneously with a single MS/MS scan at 10 eV was employed to analyze in solid-support liquid-liquid extracts (SLE) of the human blood. Detection and quantification were carried out using full scan data (protonated molecules [M+H]+) while MS/MS data were used for fragment identification only. MS/MS scans were not found to have any negative influence on quantitation under the applied conditions. Verification studies were accomplished at low concentrations (1 ng/mL), and accuracy errors lower than 5 ppm were achieved. The human blood extracts spiked at LOQ and three QC fortification levels produced average recoveries in the range of 79.3-92.5% with relative standard deviations smaller than 10% for all analytes. Limits of detection (LODs) were between 0.1 and 0.5 ng/mL. Finally, the proposed method was successfully applied to a case of human poisoning.
Subject(s)
Insecticides/blood , Toluidines/blood , Chromatography, Liquid , Forensic Toxicology , Humans , Poisoning/blood , Poisoning/diagnosisABSTRACT
BACKGROUND: Teriflunomide, a once-daily oral immunomodulator approved for treatment of relapsing-remitting multiple sclerosis, is eliminated slowly from plasma. If necessary to rapidly lower plasma concentrations of teriflunomide, an accelerated elimination procedure using cholestyramine or activated charcoal may be used. The current bioanalytical assay for determination of plasma teriflunomide concentration requires laboratory facilities for blood centrifugation and plasma storage. An alternative method, with potential for greater convenience, is dried blood spot (DBS) methodology. Analytical and clinical validations are required to switch from plasma to DBS (finger-prick sampling) methodology. METHODS: Using blood samples from healthy subjects, an LC-MS/MS assay method for quantification of teriflunomide in DBS over a range of 0.01-10 mcg/mL was developed and validated for specificity, selectivity, accuracy, precision, reproducibility, and stability. Results were compared with those from the current plasma assay for determination of plasma teriflunomide concentration. RESULTS: Method was specific and selective relative to endogenous compounds, with process efficiency â¼88%, and no matrix effect. Inaccuracy and imprecision for intraday and interday analyses were <15% at all concentrations tested. Quantification of teriflunomide in DBS assay was not affected by blood deposit volume and punch position within spot, and hematocrit level had a limited but acceptable effect on measurement accuracy. Teriflunomide was stable for at least 4 months at room temperature, and for at least 24 hours at 37°C with and without 95% relative humidity, to cover sampling, drying, and shipment conditions in the field. The correlation between DBS and plasma concentrations (R = 0.97), with an average blood to plasma ratio of 0.59, was concentration independent and constant over time. CONCLUSIONS: DBS sampling is a simple and practical method for monitoring teriflunomide concentrations.
Subject(s)
Crotonates/blood , Plasma/chemistry , Toluidines/blood , Blood Specimen Collection/methods , Chromatography, Liquid/methods , Dried Blood Spot Testing/methods , Drug Stability , Hematocrit/methods , Humans , Hydroxybutyrates , Nitriles , Reproducibility of Results , Sensitivity and Specificity , Tandem Mass Spectrometry/methodsABSTRACT
This study describes a comparison between LC-UV and LC-MS method for the simultaneous analyses of a few disease-modifying agents of multiple sclerosis. Quantitative determination of fampridine (FAM), teriflunomide (TFM) and dimethyl fumarate (DMF) was performed in human plasma with the recovery values in the range of 85-115%. A reversed-phase high-performance liquid chromatography (HPLC) with UV as well as MS detection is used. The method utilizes an XBridge C18 silica column and a gradient elution with mobile phase consisting of ammonium formate and acetonitrile at a flow rate of 0.5 mL min(-1) . The method adequately resolves FAM, TFM and DMF within a run time of 15 min. Owing to low molecular weights, the estimation of DMF and FAM is more versatile in UV than MS detection. With LC-UV, the detection limits of FAM, TFM and DMF were 0.1, 0.05, 0.05 µg and the quantification limit for all the analytes was 1 µg. With LC-MS, the detection and quantification limits for all of the analytes were 1 and 5 ng, respectively. The two techniques were completely validated and shown to be reproducible and sensitive. They were applied to a pharmacokinetic study in rats by a single oral dose. Copyright © 2016 John Wiley & Sons, Ltd.
Subject(s)
4-Aminopyridine/blood , Chromatography, Liquid/methods , Crotonates/blood , Dimethyl Fumarate/blood , Mass Spectrometry/methods , Spectrophotometry, Ultraviolet/methods , Toluidines/blood , 4-Aminopyridine/pharmacokinetics , Animals , Crotonates/pharmacokinetics , Dimethyl Fumarate/pharmacokinetics , Humans , Hydroxybutyrates , Nitriles , Rats , Reference Standards , Reproducibility of Results , Toluidines/pharmacokineticsABSTRACT
Leflunomide is a prodrug used primarily for treatment of rheumatoid arthritis. The active metabolite, teriflunomide (A77 1726), inhibits the enzyme dihydroorotate dehydrogenase and thereby reduces the synthesis of pyrimidine ribonucleotides. Teriflunomide is also administered directly and finds use in treating multiple sclerosis. Therapeutic concentrations are generally in the tens of µg/mL serum or plasma and, due to adverse effects and the time required to reach steady state, therapeutic drug monitoring is beneficial. The drug is also a potential teratogen. A method was developed and validated to quantify the drug teriflunomide over a 40,000-fold concentration range of 5 ng/mL to 200 µg/mL in serum or plasma. This is accomplished by dividing the quantitative range into two separate but overlapping regions; a high curve and a low curve range. Samples are evaluated first against the high curve after a 100-fold dilution of the sample extract. Samples falling below the upper curve region are evaluated again without dilution and quantified, if possible, against the low curve calibration standards. Appropriate choice of a concentration for the deuterated internal standard (D4-teriflunomide) allows for a single, identical, extraction procedure to be performed for both curve regions but with the dilution performed for high curve samples. The method is rugged and reliable with good accuracy and precision statistics.
Subject(s)
Chromatography, Liquid/methods , Crotonates/blood , Drug Monitoring/methods , Prodrugs/pharmacokinetics , Tandem Mass Spectrometry/methods , Toluidines/blood , Arthritis, Rheumatoid/drug therapy , Dihydroorotate Dehydrogenase , Humans , Hydroxybutyrates , Isoxazoles/blood , Leflunomide , Limit of Detection , Nitriles , Oxidoreductases Acting on CH-CH Group Donors/antagonists & inhibitorsABSTRACT
The intracellular transcription factor aryl hydrocarbon receptor (AHR) is bound and activated by xenobiotics, thereby promoting their catabolism by inducing expression of cytochrome P450 oxidase (CYP) genes through binding xenobiotic response elements (XRE) in their promoter region. In addition, it is involved in several cellular pathways like cell proliferation, differentiation, regeneration, tumor invasiveness and immune responses. Several pharmaceutical compounds like benzimidazoles activate the AHR and induce their own metabolic degradation. Using newly generated XRE-reporter mice, which allow in vivo bioluminescence imaging of AHR activation, we show here that the AHR is activated in vivo by teriflunomide (TER), which has recently been approved for the treatment of multiple sclerosis. While we did not find any evidence that the AHR mediates the immunomodulatory effects of TER, AHR activation led to metabolism and detoxification of teriflunomide, most likely via CYP. Mice deficient for the AHR show higher blood levels of teriflunomide, suffer from enhanced thrombo- and leukopenia and elevated liver enzymes as well as from severe gastrointestinal ulcers and bleeding which are lethal after 8-11 days of treatment. Leukopenia, acute liver damage and diarrhea have also been described as common side effects in human trials with TER. These data suggest that the AHR is relevant for detoxification not only of environmental toxins but also of drugs in clinical use, with potential implications for the application of AHR-modifying therapies in conjunction to TER in humans. The XRE-reporter mouse is a useful novel tool for monitoring AHR activation using in vivo imaging.
Subject(s)
Crotonates/toxicity , Encephalomyelitis, Autoimmune, Experimental/physiopathology , Receptors, Aryl Hydrocarbon/physiology , Toluidines/toxicity , Animals , Crotonates/blood , Hydroxybutyrates , Mice , Mice, Inbred C57BL , Multiple Sclerosis/physiopathology , Nitriles , Receptors, Aryl Hydrocarbon/genetics , Toluidines/bloodABSTRACT
BACKGROUND: A liquid chromatography tandem mass spectrometry (LC-MS/MS) method has been validated for use in therapeutic monitoring of the drug leflunomide in human serum and plasma. Because of concerns of teratogenicity, it is recommended that women who want to become pregnant have concentrations below 0.02 mcg/mL, although therapeutic levels are generally greater than 20 mcg/mL. Consequently, the method required a 40,000-fold dynamic range, which was achieved by dividing the curve range into 2 separate regions but with a single extraction procedure used for both. METHODS: A chromatographic separation was achieved between the parent drug and the active metabolite, teriflunomide (A77 1726), and the latter was quantified across a quantitative range of 0.005-200 mcg/mL. Samples were evaluated in an upper curve region first, with dilution, to determine whether the drug concentrations were in an appropriate therapeutic range. Samples that fell below the upper region were then reevaluated in the lower region without dilution. RESULTS: The method was shown to be reliable, with good accuracy and precision statistics, and acceptable quantitation using 4 different collection tube types. Mean accuracy over 6 control concentrations was within 5.4%, over 5 validation runs, whereas %coefficient of variation (CV) was within 8.15%. Evaluation of sodium heparin, KEDTA, NaF/K oxalate, and plain serum tubes from 6 separate individuals at the lower limit of quantification (LLOQ) showed no influence on the ability to quantify teriflunomide accurately. Regression equations for a curve range of 0.005-1 mcg/mL gave R values of 0.998 or better, whereas the range 0.8-200 mcg/mL had R values of 0.997 or better. CONCLUSIONS: The authors have developed and validated a method that allows quantification of leflunomide across a 40,000-fold range of 0.005-200 mcg/mL.
Subject(s)
Crotonates/blood , Toluidines/blood , Calibration , Chromatography, Liquid/standards , Humans , Hydroxybutyrates , Limit of Detection , Nitriles , Reference Standards , Reproducibility of Results , Tandem Mass Spectrometry/standardsABSTRACT
BACKGROUND: Carcinogenic aromatic amines derived from hair dyes have recently received new attention. One of these is ortho (o)-toluidine, which is classified as carcinogenic to humans. OBJECTIVES: To clarify exposure of hairdressers to potentially carcinogenic aromatic amines, including o-toluidine. METHODS: We measured eight potentially carcinogenic aromatic amines in the blood of 295 hairdressers, 32 users of hair dyes and 60 controls. The study was restricted to female non-smokers. Lifestyle data were collected for all participants using self-administered questionnaires. Blood samples were taken for analysis of ortho-, meta (m)- and para (p)-toluidine; 2-, 3- and 4-ethylaniline, 2,3- and 3,4-dimethylaniline as haemoglobin adducts. The samples were analysed with gas chromatography-tandem mass spectrometry. RESULTS: Generally, adduct concentrations were in the range of 0-200â pg/g haemoglobin. A comparison of the adduct concentrations found in hairdressers, consumers and controls showed no statistically significant differences. However, for hairdressers, o- and m-toluidine concentrations increased significantly with the weekly number of hair waving (p=0.020) and permanent hair dyeing treatments (p=0.026), respectively. o-Toluidine and m-Toluidine concentrations also tended (p=0.076 and 0.080, respectively) to increase with the frequency of light-colour permanent hair dye treatments. CONCLUSIONS: Hairdressers who use light-colour permanent hair dyes, other permanent hair dyes and hair waving treatments seem to be exposed to o- and m-toluidine as indicated by associations with the number of treatments performed. Analyses of hair waving and hair dye products should be performed to identify the possible sources of exposure to o- and m-toluidine.
Subject(s)
Barbering , Hair Dyes/chemistry , Hemoglobins/metabolism , Occupational Exposure/analysis , Toluidines/blood , Adult , Aniline Compounds/blood , Carcinogens/analysis , Female , Gas Chromatography-Mass Spectrometry , Hair Dyes/metabolism , Humans , Middle AgedABSTRACT
A method was developed for determination of amitraz and its metabolites, N-[2,4-(dimethylphenyl)-N'-methylformamidine (DMPF), 2,4-dimethylformamidine (DMF), 2,4-dimethylaniline (DMA) in whole blood. The analytes were extracted by solid-phase extraction (SPE) using dichloromethane, acetonitrile and methanol (2:1:1) mixture as elute solution. Analysis was performed by liquid chromatography-tandem mass spectrometry (LC-MS/MS) in the positive ion mode using multiple reaction monitoring (MRM) technique. Collision-induced dissociation (CID) of amitraz at the electrospray source in MS/MS was observed in the analytic conditions. The method was validated in human whole blood spiked at three concentration levels. The low limit of detection (LOD) and the low limit of quantification (LOQ) for all the analytes were below 0.5µg/L and 2µg/L, respectively. Recoveries were between 90.2% and 104.5%, Bias and relative standard deviation (RSD) were below 15% (n=6). The good linear relationships were obtained in certain concentration ranges of amitraz and its metabolites. The results demonstrated the method is exclusive, sensitive and accurate, and can be applied in forensic toxicology.
Subject(s)
Chromatography, Liquid/methods , Solid Phase Extraction/methods , Tandem Mass Spectrometry/methods , Toluidines/blood , Child, Preschool , Female , Humans , Limit of Detection , Linear Models , Reproducibility of Results , Toluidines/chemistry , Toluidines/isolation & purification , Toluidines/poisoningABSTRACT
Leflunomide is a disease-modifying antirheumatic drug. The purpose of this study was to evaluate the bioequivalence of a test drug (CJ leflunomide) and a commercially available reference drug (Arava®) at 2 doses (10 and 20 mg) in healthy Korean volunteers. This was a single-dose (28 individuals enrolled at each dose group), randomized, open-label, 2-way crossover study. The 2 treatment periods were separated by a 56-day wash-out interval. Blood sampling was conducted until 672 h after drug administration. Plasma teriflunomide (active metabolite of leflunomide) concentrations were determined, and pharmacokinetic parameters were calculated. Bioequivalence was evaluated using an ANOVA model, based on the AUCt and the Cmax after administration of leflunomide tablets. Bioequivalence was defined as the 90% confidence intervals (CIs) of the geometric mean ratios (GMRs) of AUCt and Cmax for the test and reference drugs being within the range of 0.80-1.25. The GMRs (90% CI) for AUCt and Cmax were 0.9506 (0.9091-0.9941) and 0.9861 (0.9360-1.0389), respectively, in the 10 mg study, and 0.9524 (0.9101-0.9968) and 0.9740 (0.9314-1.0186), respectively, in the 20 mg study. The 90% CIs of AUCt and Cmax at each dose were within the accepted range for bioequivalence. Based on the results, the test drug (CJ leflunomide) was bioequivalent to the commercially available reference drug (Arava®) at both doses.
Subject(s)
Antirheumatic Agents/pharmacokinetics , Crotonates/blood , Isoxazoles/pharmacokinetics , Toluidines/blood , Adult , Analysis of Variance , Antirheumatic Agents/administration & dosage , Area Under Curve , Cross-Over Studies , Dose-Response Relationship, Drug , Humans , Hydroxybutyrates , Isoxazoles/administration & dosage , Leflunomide , Middle Aged , Nitriles , Republic of Korea , Therapeutic Equivalency , Young AdultABSTRACT
Pharmacokinetic data of disease modifying antirheumatic drugs during hemodialysis are limited to sulfasalazine, methotrexate, and cyclosporine. Only respective anecdotal data have been reported on leflunomide. We repeatedly measured teriflunomide (A77-1726), the active metabolite of leflunomide, during standard hemodialysis sessions and calculated teriflunomide clearances in five patients with rheumatoid arthritis (RA) and end-stage renal disease. The calculated teriflunomide clearances during a standardized dialysis session of 3-4.5 h at a blood flow rate of 160-300 ml/min were between 0 and 4.3 ml/min, the mean clearances of the total dialysis ranged between 1.1 and 3.4 ml/min. Total amount of teriflunomide removed was 5.8-8.8 µg per dialysis session. Dialytic removal of the active metabolite of leflunomide, teriflunomide (A77-1726), is negligible. Leflunomide can be used for RA patients on chronic dialysis without any dosage modification.
Subject(s)
Antirheumatic Agents/pharmacokinetics , Arthritis, Rheumatoid/drug therapy , Isoxazoles/pharmacokinetics , Kidney Failure, Chronic/therapy , Renal Dialysis , Adult , Aged , Aged, 80 and over , Antirheumatic Agents/blood , Antirheumatic Agents/chemistry , Arthritis, Rheumatoid/complications , Crotonates/blood , Crotonates/chemistry , Female , Humans , Hydroxybutyrates , Isoxazoles/blood , Isoxazoles/chemistry , Kidney Failure, Chronic/complications , Leflunomide , Male , Middle Aged , Models, Biological , Nitriles , Retrospective Studies , Toluidines/blood , Toluidines/chemistrySubject(s)
Artifacts , Blood Chemical Analysis/methods , Calcium/blood , Calcium/chemistry , Crotonates/blood , Crotonates/metabolism , Isoxazoles/metabolism , Toluidines/blood , Toluidines/metabolism , False Positive Reactions , Humans , Hydroxybutyrates , Kidney Transplantation , Leflunomide , NitrilesABSTRACT
The pharmacokinetics of teriflunomide [CAS No. 163451-81-8], the metabolite of leflunomide [CAS No. 75706-12-6] has been evaluated in adult human volunteers after oral administration of tablet formulation. However, no published data is available regarding the bioavailability of this in the Indian population. In light of the above, a study was designed to carry out a bioequivalence study of 2 preparations of leflunomide 20 mg in healthy Indian male volunteers.24 healthy male volunteers (age, 25±4.1 years; weight, 57.58±7.01 kg) were enrolled in this study. Each subject received a test and reference formulation in a single dose, fasting 2 period, 2 way crossover study with a wash out period of 4 weeks. Analysis of teriflunomide from plasma samples was done by a simple and sensitive HPLC method using UV detection developed in our laboratory. An analysis of variance was performed on the pharmacokinetic parameters Cmax, AUC0-t, AUC0-∞ using GLM procedures in which sources of variation were subject, formulation, and period.The results indicated that there are no statistically significant differences between the 2 products in either the mean concentration-time profiles or in the obtained pharmacokinetic parameters. 90% confidence limits for the log transformed data of Cmax, AUC0-t, AUC0-∞. were within the acceptable range of 0.80-1.25.The results indicate that the 2 products are bioequivalent in terms of rate and extent of drug absorption. Both the preparations were well tolerated with no adverse reactions throughout the study.