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Determination of a highly selective mixed-affinity sigma receptor ligand, in rat plasma by ultra performance liquid chromatography mass spectrometry and its application to a pharmacokinetic study
23 March 2012,
10:56:29
Publication year:
2012
Source:Journal of Chromatography B, Volumes 891–892
Seshulatha Jamalapuram, Pradeep K. Vuppala, Christophe Mesangeau, Christopher R. McCurdy, Bonnie A. Avery
A selective, rapid and sensitive ultra performance liquid chromatography mass spectrometry (UPLC/MS) method was developed and validated to quantitate a highly selective mixed-affinity sigma receptor ligand, CM156 (3-(4-(4-cyclohexylpiperazin-1-yl)butyl)benzo[d] thiazole-2(3H)-thione), in rat plasma. CM156 and the internal standard (aripiprazole) were extracted from plasma samples by a single step liquid–liquid extraction using chloroform. The analysis was carried out on an ACQUITY UPLC™ BEH HILIC column (1.7μm, 2.1mm×50mm) with isocratic elution at flow rate of 0.2mL/min using 10mM ammonium formate in 0.1% formic acid and acetonitrile (10:90) as the mobile phase. The detection of the analyte was performed on a mass spectrometer operated in selected ion recording (SIR) mode with positive electrospray ionization (ESI). The validated analytical method resulted in a run time of 4min and the retention times observed were 2.6±0.1 and 2.1±0.1min for CM156 and the IS, respectively. The calibration curve exhibited excellent linearity over a concentration range of 5–4000ng/mL with the lower limit of quantification of 5ng/mL. The intra- and inter-day precision values were below 15% and accuracy ranged from −6.5% to 5.0%. The mean recovery of CM156 from plasma was 96.8%. The validated method was applied to a pilot intravenous pharmacokinetic study in rats.
Source:Journal of Chromatography B, Volumes 891–892
Seshulatha Jamalapuram, Pradeep K. Vuppala, Christophe Mesangeau, Christopher R. McCurdy, Bonnie A. Avery
A selective, rapid and sensitive ultra performance liquid chromatography mass spectrometry (UPLC/MS) method was developed and validated to quantitate a highly selective mixed-affinity sigma receptor ligand, CM156 (3-(4-(4-cyclohexylpiperazin-1-yl)butyl)benzo[d] thiazole-2(3H)-thione), in rat plasma. CM156 and the internal standard (aripiprazole) were extracted from plasma samples by a single step liquid–liquid extraction using chloroform. The analysis was carried out on an ACQUITY UPLC™ BEH HILIC column (1.7μm, 2.1mm×50mm) with isocratic elution at flow rate of 0.2mL/min using 10mM ammonium formate in 0.1% formic acid and acetonitrile (10:90) as the mobile phase. The detection of the analyte was performed on a mass spectrometer operated in selected ion recording (SIR) mode with positive electrospray ionization (ESI). The validated analytical method resulted in a run time of 4min and the retention times observed were 2.6±0.1 and 2.1±0.1min for CM156 and the IS, respectively. The calibration curve exhibited excellent linearity over a concentration range of 5–4000ng/mL with the lower limit of quantification of 5ng/mL. The intra- and inter-day precision values were below 15% and accuracy ranged from −6.5% to 5.0%. The mean recovery of CM156 from plasma was 96.8%. The validated method was applied to a pilot intravenous pharmacokinetic study in rats.
Determination of landiolol, an ultra-short-acting β1-receptor antagonist, in human plasma by liquid chromatography–tandem mass spectrometry
23 March 2012,
10:56:29
Publication year:
2012
Source:Journal of Chromatography B, Volumes 891–892
Qun He, Meiyun Shi, Xidong Liu, Yantong Sun, Lianghai Hu, Yan Yang, J. Paul Fawcett, Jingkai Gu, Limei Zhao
A method for the determination of landiolol, an ultra-short-acting β1-adrenoreceptor antagonist, in human plasma has been developed and validated. With the addition of pyridostigmine bromide to stabilize landiolol in the blood/plasma samples, and bisoprolol as internal standard, plasma samples were subjected to liquid–liquid extraction with diethyl ether:dicholoromethane (60:40, v/v) prior to assay by liquid chromatography–tandem mass spectrometry. Separation was performed on a TC-C18 column (150mm×4.6mm, 5μm) using a mobile phase of methanol:10mM ammonium acetate containing 1% formic acid (65:35, v/v) in a run time of 3.5min. Detection involved electrospray ionization in the positive ion mode followed by multiple reaction monitoring of the precursor-to-product ion transitions of landiolol at m/z 510.1→157.2 and bisoprolol at m/z 326.3→116.1. The method was linear over the concentration range 0.5–500ng/ml with a lower limit of quantitation of 0.5ng/ml. Intra- and inter-day precisions (as relative standard deviation, RSD) were <4.4% and <10.0%, respectively, with accuracy (as relative error, RE) <10.0%. The method was successfully applied to a clinical pharmacokinetic study involving a continuous infusion of landiolol hydrochloride to healthy Chinese volunteers.
Source:Journal of Chromatography B, Volumes 891–892
Qun He, Meiyun Shi, Xidong Liu, Yantong Sun, Lianghai Hu, Yan Yang, J. Paul Fawcett, Jingkai Gu, Limei Zhao
A method for the determination of landiolol, an ultra-short-acting β1-adrenoreceptor antagonist, in human plasma has been developed and validated. With the addition of pyridostigmine bromide to stabilize landiolol in the blood/plasma samples, and bisoprolol as internal standard, plasma samples were subjected to liquid–liquid extraction with diethyl ether:dicholoromethane (60:40, v/v) prior to assay by liquid chromatography–tandem mass spectrometry. Separation was performed on a TC-C18 column (150mm×4.6mm, 5μm) using a mobile phase of methanol:10mM ammonium acetate containing 1% formic acid (65:35, v/v) in a run time of 3.5min. Detection involved electrospray ionization in the positive ion mode followed by multiple reaction monitoring of the precursor-to-product ion transitions of landiolol at m/z 510.1→157.2 and bisoprolol at m/z 326.3→116.1. The method was linear over the concentration range 0.5–500ng/ml with a lower limit of quantitation of 0.5ng/ml. Intra- and inter-day precisions (as relative standard deviation, RSD) were <4.4% and <10.0%, respectively, with accuracy (as relative error, RE) <10.0%. The method was successfully applied to a clinical pharmacokinetic study involving a continuous infusion of landiolol hydrochloride to healthy Chinese volunteers.
Gas chromatography–mass spectrometry determination of pharmacologically active substances in urine and blood samples by use of a continuous solid-phase extraction system and microwave-assisted derivatization
23 March 2012,
10:56:29
Publication year:
2012
Source:Journal of Chromatography B, Volumes 891–892
Abdelmonaim Azzouz, Evaristo Ballesteros
A sensitive method based on gas chromatography–mass spectrometry was used to determine 22 pharmacologically active substances (frequently used in the treatment of human and animal's diseases) including analgesics, antibacterials, anti-epileptics, antiseptics, β-blockers, hormones, lipid regulators and non-steroidal anti-inflammatories in blood and urine samples. Samples were subjected to continuous solid-phase extraction in a sorbent column (Oasis HLB), and then the target analytes were eluted with ethyl acetate and derivatized in a household microwave oven at 350W for 3min. Finally, these products were determined in a gas chromatograph–mass spectrometer equipped with a DB-5 fused silica capillary column. The analyte detection limits thus obtained ranged from 0.2 to 1.3ngL−1 for urine samples and 0.8–5.6ngL−1 for blood samples. Recoveries from both blood and urine ranged from 85 to 102%, and within-day and between-day relative standard deviations were all less than 7.5%. The proposed method offers advantages in reduction of the exposure danger to toxic solvents used in conventional sample pretreatment, simplicity of the extraction processes, rapidity, and sensitivity enhancement. The method was successfully used to quantify pharmacologically active substances in human and animal (lamb, veal and pig) blood and urine. The hormones estrone and 17β-estradiol were detected in virtually all samples, and so were other analytes such as acetylsalicylic acid, ibuprofen, ketoprofen and triclosan in human samples, and florfenicol, pyrimethamine and phenylbutazone in animal samples.
Source:Journal of Chromatography B, Volumes 891–892
Abdelmonaim Azzouz, Evaristo Ballesteros
A sensitive method based on gas chromatography–mass spectrometry was used to determine 22 pharmacologically active substances (frequently used in the treatment of human and animal's diseases) including analgesics, antibacterials, anti-epileptics, antiseptics, β-blockers, hormones, lipid regulators and non-steroidal anti-inflammatories in blood and urine samples. Samples were subjected to continuous solid-phase extraction in a sorbent column (Oasis HLB), and then the target analytes were eluted with ethyl acetate and derivatized in a household microwave oven at 350W for 3min. Finally, these products were determined in a gas chromatograph–mass spectrometer equipped with a DB-5 fused silica capillary column. The analyte detection limits thus obtained ranged from 0.2 to 1.3ngL−1 for urine samples and 0.8–5.6ngL−1 for blood samples. Recoveries from both blood and urine ranged from 85 to 102%, and within-day and between-day relative standard deviations were all less than 7.5%. The proposed method offers advantages in reduction of the exposure danger to toxic solvents used in conventional sample pretreatment, simplicity of the extraction processes, rapidity, and sensitivity enhancement. The method was successfully used to quantify pharmacologically active substances in human and animal (lamb, veal and pig) blood and urine. The hormones estrone and 17β-estradiol were detected in virtually all samples, and so were other analytes such as acetylsalicylic acid, ibuprofen, ketoprofen and triclosan in human samples, and florfenicol, pyrimethamine and phenylbutazone in animal samples.
A LC–MS/MS method to evaluate the hepatic uptake of the liver-specific magnetic resonance imaging contrast agent gadoxetate (Gd-EOB-DTPA) in vitro and in humans
23 March 2012,
10:56:29
Publication year:
2012
Source:Journal of Chromatography B, Volumes 891–892
Jia Jia, Markus Keiser, Ali Nassif, Werner Siegmund, Stefan Oswald
Gadoxetate (Gd-EOB-DTPA, Primovist®) is a frequently used liver-specific magnetic resonance imaging (MRI) contrast agent which disposition is so far not fully understood in humans. Here, we describe the development and validation of a selective and sensitive quantification method to measure cellular in vitro concentrations as well as human serum concentrations of gadoxetate. The drug was measured after protein precipitation with acetonitrile and ethyl acetate-mediated sample concentration using amoxicillin as internal standard and liquid chromatography coupled with tandem mass spectrometry (LC–MS/MS) for detection. Hydrophilic interaction chromatography (HILIC) was performed by using the column Atlantis® HILIC Silica (2.1mm×100mm), a step-elution gradient with acetonitrile and ammonium acetate (5mM, pH 3.8) as mobile phases and a flow rate of 200μl/min. The MS/MS detection was done in the negative multiple reaction monitoring (MRM) mode by monitoring the m/z transitions 681.3/635.2 for gadotrexate and 363.8/222.7 for the internal standard. The method was validated between 5 and 4000ng/ml in serum and between 1.25 and 500ng/ml in cell lysates. The method was shown to possess sufficient specificity, accuracy, precision and stability without any matrix effects, thereby fulfilling current bioanalytical guidelines. The developed assay was successfully applied to quantify gadoxetate in cellular uptake studies in OATP1B1-transfected cell lines and to monitor serum concentrations-time profiles from a clinical pilot study performed in healthy volunteers carrying the wild-type or the functionally relevant variants T521C (*5) and A388G (*1b) of the hepatic uptake transporter OATP1B1.
Source:Journal of Chromatography B, Volumes 891–892
Jia Jia, Markus Keiser, Ali Nassif, Werner Siegmund, Stefan Oswald
Gadoxetate (Gd-EOB-DTPA, Primovist®) is a frequently used liver-specific magnetic resonance imaging (MRI) contrast agent which disposition is so far not fully understood in humans. Here, we describe the development and validation of a selective and sensitive quantification method to measure cellular in vitro concentrations as well as human serum concentrations of gadoxetate. The drug was measured after protein precipitation with acetonitrile and ethyl acetate-mediated sample concentration using amoxicillin as internal standard and liquid chromatography coupled with tandem mass spectrometry (LC–MS/MS) for detection. Hydrophilic interaction chromatography (HILIC) was performed by using the column Atlantis® HILIC Silica (2.1mm×100mm), a step-elution gradient with acetonitrile and ammonium acetate (5mM, pH 3.8) as mobile phases and a flow rate of 200μl/min. The MS/MS detection was done in the negative multiple reaction monitoring (MRM) mode by monitoring the m/z transitions 681.3/635.2 for gadotrexate and 363.8/222.7 for the internal standard. The method was validated between 5 and 4000ng/ml in serum and between 1.25 and 500ng/ml in cell lysates. The method was shown to possess sufficient specificity, accuracy, precision and stability without any matrix effects, thereby fulfilling current bioanalytical guidelines. The developed assay was successfully applied to quantify gadoxetate in cellular uptake studies in OATP1B1-transfected cell lines and to monitor serum concentrations-time profiles from a clinical pilot study performed in healthy volunteers carrying the wild-type or the functionally relevant variants T521C (*5) and A388G (*1b) of the hepatic uptake transporter OATP1B1.
Rapid determination of endogenous cytokinins in plant samples by combination of magnetic solid phase extraction with hydrophilic interaction chromatography–tandem mass spectrometry
23 March 2012,
10:56:29
Publication year:
2012
Source:Journal of Chromatography B, Volumes 891–892
Zhao Liu, Bao-Dong Cai, Yu-Qi Feng
A 2-acrylamido-2-methyl-1-propanesulfonic acid-co-ethylene glycol dimethacrylate (Fe3O4/SiO2/P(AMPS-co-EGDMA)) copolymer was prepared and used as a magnetic solid phase extraction (MSPE) medium for recovery of endogenous cytokinins (CKs) from plant extracts. This magnetic porous polymer was characterized by electron microscopy, nitrogen sorption experiments, elemental analysis and Fourier-transformed infrared spectroscopy. It was demonstrated to have high extraction capacity toward CKs in plants due to its specificity, surface area and porous structure. Coupled with hydrophilic interaction chromatography–tandem mass spectrometry (HILIC–MS/MS), a rapid, simple, and effective MSPE–HILIC–MS/MS analytical method for the quantitative analysis of endogenous CKs in Oryza sativa (O. sativa) roots was successfully established. Good linearities were obtained for all CKs investigated with correlation coefficients (R 2)>0.9975. The results showed that LODs (S/N=3) were ranged from 0.18 to 3.65pgmL−1. Reproducibility of the method was obtained with intra-day and inter-day relative standard deviations (RSDs) less than 16.1% and the recoveries in plant samples ranged from 72.8% to 115.5%. Finally, the MSPE–HILIC–MS/MS method was applied to several plant samples, and the amounts of endogenous CKs in O. sativa roots, leaves and Arabidopsis thaliana (A. thaliana) were successfully determined.
Source:Journal of Chromatography B, Volumes 891–892
Zhao Liu, Bao-Dong Cai, Yu-Qi Feng
A 2-acrylamido-2-methyl-1-propanesulfonic acid-co-ethylene glycol dimethacrylate (Fe3O4/SiO2/P(AMPS-co-EGDMA)) copolymer was prepared and used as a magnetic solid phase extraction (MSPE) medium for recovery of endogenous cytokinins (CKs) from plant extracts. This magnetic porous polymer was characterized by electron microscopy, nitrogen sorption experiments, elemental analysis and Fourier-transformed infrared spectroscopy. It was demonstrated to have high extraction capacity toward CKs in plants due to its specificity, surface area and porous structure. Coupled with hydrophilic interaction chromatography–tandem mass spectrometry (HILIC–MS/MS), a rapid, simple, and effective MSPE–HILIC–MS/MS analytical method for the quantitative analysis of endogenous CKs in Oryza sativa (O. sativa) roots was successfully established. Good linearities were obtained for all CKs investigated with correlation coefficients (R 2)>0.9975. The results showed that LODs (S/N=3) were ranged from 0.18 to 3.65pgmL−1. Reproducibility of the method was obtained with intra-day and inter-day relative standard deviations (RSDs) less than 16.1% and the recoveries in plant samples ranged from 72.8% to 115.5%. Finally, the MSPE–HILIC–MS/MS method was applied to several plant samples, and the amounts of endogenous CKs in O. sativa roots, leaves and Arabidopsis thaliana (A. thaliana) were successfully determined.
Measurements of polybrominated diphenyl ethers and polychlorinated biphenyls in a single drop of blood
23 March 2012,
10:56:29
Publication year:
2012
Source:Journal of Chromatography B, Volumes 891–892
Dasheng Lu, Dongli Wang, Ho Sai Simon Ip, Frank Barley, Robert Ramage, Jianwen She
A quantitative method that requires only a small volume (50μL) of blood has been developed for the determination of polybrominated diphenyl ethers (PBDEs) and polychlorinated biphenyls (PCBs). Target analytes in both plasma sample (DBSV) and dried blood spot (DBS) were analyzed by a gas chromatography/high resolution mass spectrometer (GC/HRMS). Measurements of standard reference materials by the developed method were in agreement with those certified values. Linear correlation coefficients were found to be 0.9984 and 0.9965 for DBS and DBSV analysis, respectively. Other analytical criteria, such as limits of detection, recoveries, precision, accuracy and linearity of the proposed method are also reported. From recovery studies, the addition of formic acid to the extraction solvent was found to be effective in extracting PBDEs and PCBs from filter paper. The PBDE and PCB levels in spiked DBS were monitored at room temperature for up to 30 days and the variations of target analytes were found to be insignificant. Our results suggest that DBS sampling technique is feasible for PBDE and PCBs biomonitoring in human population.
Source:Journal of Chromatography B, Volumes 891–892
Dasheng Lu, Dongli Wang, Ho Sai Simon Ip, Frank Barley, Robert Ramage, Jianwen She
A quantitative method that requires only a small volume (50μL) of blood has been developed for the determination of polybrominated diphenyl ethers (PBDEs) and polychlorinated biphenyls (PCBs). Target analytes in both plasma sample (DBSV) and dried blood spot (DBS) were analyzed by a gas chromatography/high resolution mass spectrometer (GC/HRMS). Measurements of standard reference materials by the developed method were in agreement with those certified values. Linear correlation coefficients were found to be 0.9984 and 0.9965 for DBS and DBSV analysis, respectively. Other analytical criteria, such as limits of detection, recoveries, precision, accuracy and linearity of the proposed method are also reported. From recovery studies, the addition of formic acid to the extraction solvent was found to be effective in extracting PBDEs and PCBs from filter paper. The PBDE and PCB levels in spiked DBS were monitored at room temperature for up to 30 days and the variations of target analytes were found to be insignificant. Our results suggest that DBS sampling technique is feasible for PBDE and PCBs biomonitoring in human population.
LC–ESI-MS/MS determination of in vivo metabolites of almotriptan in rat plasma, urine and feces: Application to pharmacokinetics
23 March 2012,
10:56:29
Publication year:
2012
Source:Journal of Chromatography B, Volumes 891–892
R. Nageswara Rao, K. Guruprasad, Ch. Gangu Naidu, B. Raju, R. Srinivas
A highly sensitive and specific liquid chromatography–electrospray ionization tandem mass spectrometric (LC–ESI-MS/MS) method for investigating the in vivo metabolites of almotriptan in rat plasma, feces and urine was developed. Chromatographic separation was achieved on a Lichrospher RP-18 column (250mm×4.6mm, 5μm), using 20mM ammonium acetate (pH 3.5) and acetonitrile (60:40, v/v) as a mobile phase at 25°C. MS/MS detection was performed by positive ion electrospray ionization using target ions at m/z 336 [M+H]+, m/z 368 and m/z 282 [M+H]+ for almotriptan and its two metabolites, respectively. Two metabolites viz., γ-aminobutyric acid and sulfonamide were detected in plasma as well as feces after 24h of oral administration of almotriptan, while only γ-aminobutyric acid was found in urine. The method was sensitive with a lower limit of quantification of 1.43ng/mL and linear over the range of 1.43–5000ng/mL in plasma. The method was validated and successfully applied to a pharmacokinetic study of almotriptan in rat plasma using sumatriptan as an internal standard. The peak plasma concentration (C max) after 0.3h of 5mg/kg oral dose of almotriptan was determined to be 69.85ng/mL.
Source:Journal of Chromatography B, Volumes 891–892
R. Nageswara Rao, K. Guruprasad, Ch. Gangu Naidu, B. Raju, R. Srinivas
A highly sensitive and specific liquid chromatography–electrospray ionization tandem mass spectrometric (LC–ESI-MS/MS) method for investigating the in vivo metabolites of almotriptan in rat plasma, feces and urine was developed. Chromatographic separation was achieved on a Lichrospher RP-18 column (250mm×4.6mm, 5μm), using 20mM ammonium acetate (pH 3.5) and acetonitrile (60:40, v/v) as a mobile phase at 25°C. MS/MS detection was performed by positive ion electrospray ionization using target ions at m/z 336 [M+H]+, m/z 368 and m/z 282 [M+H]+ for almotriptan and its two metabolites, respectively. Two metabolites viz., γ-aminobutyric acid and sulfonamide were detected in plasma as well as feces after 24h of oral administration of almotriptan, while only γ-aminobutyric acid was found in urine. The method was sensitive with a lower limit of quantification of 1.43ng/mL and linear over the range of 1.43–5000ng/mL in plasma. The method was validated and successfully applied to a pharmacokinetic study of almotriptan in rat plasma using sumatriptan as an internal standard. The peak plasma concentration (C max) after 0.3h of 5mg/kg oral dose of almotriptan was determined to be 69.85ng/mL.
Directly suspended droplet microextraction coupled with high performance liquid chromatography: A rapid and sensitive method for acetaldehyde assay in peritoneal dialysis fluids
23 March 2012,
10:56:29
Publication year:
2012
Source:Journal of Chromatography B, Volumes 891–892
Zarrin Es’haghi, Freshte Babazadeh
The aim of present study was to develop and validate a rapid, sensitive, inexpensive and reliable method for the detection of trace levels of acetaldehyde in peritoneal dialysis fluids (PDFs) by 2,4-dinitrophenylhydrazine (DNPH) derivatization and extraction. Separation and analysis of acetaldehyde via 2,4-dinitrophenylhydrazone (DNPH) was by reverse phase high performance liquid chromatography (HPLC). In order to remove co-eluting interferences and to pre-concentrate acetaldehyde, the extraction and clean-up of the sample has been performed using a liquid phase microextraction technique. In this research directly suspended droplet microextraction technique (DSDME) coupled with HPLC was used to determine acetaldehyde in PDFs. In DSDME method a free suspended droplet of an organic solvent (1-octanol) used as extraction phase. Important factors such as organic solvent, extraction time, droplet volume, sample and reagent solution volumes and rate of stirring were optimized. After extraction under optimal conditions the samples were analyzed by HPLC with UV detection at 360nm. The linearity ranged from 0.01 to 100mgL−1 with a relative standard deviation (RSD%; n =3) 5.6 Enrichment factor and limit of detection (LOD; n =5) were 54 and 1.12μgL−1, respectively.
Source:Journal of Chromatography B, Volumes 891–892
Zarrin Es’haghi, Freshte Babazadeh
The aim of present study was to develop and validate a rapid, sensitive, inexpensive and reliable method for the detection of trace levels of acetaldehyde in peritoneal dialysis fluids (PDFs) by 2,4-dinitrophenylhydrazine (DNPH) derivatization and extraction. Separation and analysis of acetaldehyde via 2,4-dinitrophenylhydrazone (DNPH) was by reverse phase high performance liquid chromatography (HPLC). In order to remove co-eluting interferences and to pre-concentrate acetaldehyde, the extraction and clean-up of the sample has been performed using a liquid phase microextraction technique. In this research directly suspended droplet microextraction technique (DSDME) coupled with HPLC was used to determine acetaldehyde in PDFs. In DSDME method a free suspended droplet of an organic solvent (1-octanol) used as extraction phase. Important factors such as organic solvent, extraction time, droplet volume, sample and reagent solution volumes and rate of stirring were optimized. After extraction under optimal conditions the samples were analyzed by HPLC with UV detection at 360nm. The linearity ranged from 0.01 to 100mgL−1 with a relative standard deviation (RSD%; n =3) 5.6 Enrichment factor and limit of detection (LOD; n =5) were 54 and 1.12μgL−1, respectively.
Quantitative determination of oseltamivir and oseltamivir carboxylate in human fluoride EDTA plasma including the ex vivo stability using high-performance liquid chromatography coupled with electrospray ionization tandem mass spectrometry
23 March 2012,
10:56:29
Publication year:
2012
Source:Journal of Chromatography B, Volumes 891–892
W. Kromdijk, H. Rosing, M.P.H. van den Broek, J.H. Beijnen, A.D.R. Huitema
Oseltamivir, the ethyl ester prodrug of the neuramidase inhibitor oseltamivir carboxylate, is licensed for the treatment of patients with influenza virus infection. Here we describe the development and validation of an assay for the simultaneous quantification of oseltamivir and oseltamivir carboxylate in human fluoride EDTA plasma including the ex vivo stability using liquid chromatography coupled to tandem mass spectrometry. Sample pretreatment consisted of protein precipitation with 8% (v/v) trichloroacetic acid in water using only 50μL plasma. Chromatographic separation was performed on a reversed phase C18 column (150mm×2.0mm ID, particle size 4μm) with a stepwise gradient using 0.1% formic acid and methanol at a flow rate of 250μL/min. A triple quadrupole mass spectrometer operating in the positive ionization mode was used for detection and drug quantification. The method was validated over a range of 3–300ng/mL for oseltamivir and 10–10,000ng/mL for oseltamivir carboxylate. Deuterated oseltamivir and oseltamivir carboxylate were used as internal standards. The intra-assay accuracies and precisions for oseltamivir were between −8.8 and 16.3% at the LLOQ level, whereas for all other concentration levels this was −8.6 and 14.5%. For oseltamivir carboxylate the intra-assay accuracies and precisions were between −10.9 and 10.7% at all levels. Furthermore, oseltamivir was stable in plasma and whole blood ex vivo in commercially available fluoride EDTA tubes for at least 24h at 2–8°C. This method is now applied for the determination of both compounds in specific patient populations to evaluate current dosing guidelines.
Source:Journal of Chromatography B, Volumes 891–892
W. Kromdijk, H. Rosing, M.P.H. van den Broek, J.H. Beijnen, A.D.R. Huitema
Oseltamivir, the ethyl ester prodrug of the neuramidase inhibitor oseltamivir carboxylate, is licensed for the treatment of patients with influenza virus infection. Here we describe the development and validation of an assay for the simultaneous quantification of oseltamivir and oseltamivir carboxylate in human fluoride EDTA plasma including the ex vivo stability using liquid chromatography coupled to tandem mass spectrometry. Sample pretreatment consisted of protein precipitation with 8% (v/v) trichloroacetic acid in water using only 50μL plasma. Chromatographic separation was performed on a reversed phase C18 column (150mm×2.0mm ID, particle size 4μm) with a stepwise gradient using 0.1% formic acid and methanol at a flow rate of 250μL/min. A triple quadrupole mass spectrometer operating in the positive ionization mode was used for detection and drug quantification. The method was validated over a range of 3–300ng/mL for oseltamivir and 10–10,000ng/mL for oseltamivir carboxylate. Deuterated oseltamivir and oseltamivir carboxylate were used as internal standards. The intra-assay accuracies and precisions for oseltamivir were between −8.8 and 16.3% at the LLOQ level, whereas for all other concentration levels this was −8.6 and 14.5%. For oseltamivir carboxylate the intra-assay accuracies and precisions were between −10.9 and 10.7% at all levels. Furthermore, oseltamivir was stable in plasma and whole blood ex vivo in commercially available fluoride EDTA tubes for at least 24h at 2–8°C. This method is now applied for the determination of both compounds in specific patient populations to evaluate current dosing guidelines.
Simultaneous determination of 1-(2′-deoxy-2′-fluoro-β-d-arabinofuranosyl) uracil (FAU) and 1-(2′-deoxy-2′-fluoro-β-d-arabinofuranosyl) 5-methyluracil (FMAU) in human plasma by liquid chromatography/tandem mass spectrometry
23 March 2012,
10:56:29
Publication year:
2012
Source:Journal of Chromatography B, Volumes 891–892
Richard Wiegand, Jianmei Wu, Anthony F. Shields, Patricia LoRusso, Jing Li
A liquid chromatography coupled with tandem mass spectrometry (LC–MS/MS) assay was developed and validated for simultaneous determination of 1-(2′-deoxy-2′-fluoro-β-d-arabinofuranosyl) uracil (FAU) and its active metabolite 1-(2′-deoxy-2′-fluoro-β-d-arabinofuranosyl) 5-methyluracil (FMAU) in human plasma. FAU and FMAU were extracted from plasma samples using solid-phase extraction with Waters Sep-Pak® Vac C18 cartridge. Chromatographic separation was achieved on a Waters Atlantis T3 C18 column with a gradient mobile phase consisting of methanol and water with 0.45% formic acid (v/v) running at a flow rate of 0.2ml/min. The analytes were monitored by triple quadrupole mass spectrometer under positive ionization mode. The lower limit of quantitation (LLOQ) was 10 and 2ng/ml for FAU and FMAU in plasma, respectively. Calibration curves were linear over FAU and FMAU plasma concentration range of 10–2000 and 2–1000ng/ml, respectively. The intra-day and inter-day accuracy and precision were within the generally accepted criteria for bioanalytical method (<15%). The method has been successfully employed to characterize the plasma pharmacokinetics of FAU and FMAU in cancer patients receiving 1-h intravenous infusion of FAU 50mg/m2.
Source:Journal of Chromatography B, Volumes 891–892
Richard Wiegand, Jianmei Wu, Anthony F. Shields, Patricia LoRusso, Jing Li
A liquid chromatography coupled with tandem mass spectrometry (LC–MS/MS) assay was developed and validated for simultaneous determination of 1-(2′-deoxy-2′-fluoro-β-d-arabinofuranosyl) uracil (FAU) and its active metabolite 1-(2′-deoxy-2′-fluoro-β-d-arabinofuranosyl) 5-methyluracil (FMAU) in human plasma. FAU and FMAU were extracted from plasma samples using solid-phase extraction with Waters Sep-Pak® Vac C18 cartridge. Chromatographic separation was achieved on a Waters Atlantis T3 C18 column with a gradient mobile phase consisting of methanol and water with 0.45% formic acid (v/v) running at a flow rate of 0.2ml/min. The analytes were monitored by triple quadrupole mass spectrometer under positive ionization mode. The lower limit of quantitation (LLOQ) was 10 and 2ng/ml for FAU and FMAU in plasma, respectively. Calibration curves were linear over FAU and FMAU plasma concentration range of 10–2000 and 2–1000ng/ml, respectively. The intra-day and inter-day accuracy and precision were within the generally accepted criteria for bioanalytical method (<15%). The method has been successfully employed to characterize the plasma pharmacokinetics of FAU and FMAU in cancer patients receiving 1-h intravenous infusion of FAU 50mg/m2.
Systematic evaluation of supported liquid extraction in reducing matrix effect and improving extraction efficiency in LC–MS/MS based bioanalysis for 10 model pharmaceutical compounds
23 March 2012,
10:56:29
Publication year:
2012
Source:Journal of Chromatography B, Volumes 891–892
Hongliang Jiang, Huachuan Cao, Yang Zhang, Douglas M. Fast
In past a few years, there has been a large increase in the application of supported liquid extraction (SLE) for LC–MS/MS based bioanalysis due to its distinct practical advantage in reduced time cost, ease of operation and the feasibility for automation. The main purpose of this study was to systematically evaluate supported liquid extraction in reducing matrix effect and improving extraction efficiency/recovery under various extraction conditions with 10 model pharmaceutical compounds in liquid chromatography coupled to electrospray tandem mass spectrometry (LC–ESI-MS/MS) analysis. Selected compounds have diverse physicochemical properties where log P ranges from 0.1 to 6.24 and pK a ranges from 4.0 to 11.1. The factors that may have the impact on the recovery of analytes and phospholipids (PL) were assessed. Over 75% recovery was achieved for every analyte under its respectively optimized extraction conditions where the selection of the polarity of extraction solvent and buffered pH can be critical for efficient recovery. Furthermore, the matrix effect was assessed by postextraction spike and postcolumn infusion method. The matrix effect was considerably reduced for all analytes under most extraction conditions evaluated for SLE, compared with protein precipitation (PPT) method. The correlation between matrix effect and residual phospholipids in sample extract was clearly shown. Although analyte-dependent matrix effect was observed prominently in sample extract prepared by PPT, it was minimized by SLE sample preparation process that effectively removes the majority of phospholipids. Sample extracted by ethyl acetate contained more phospholipids and demonstrated stronger matrix effect than by other organic solvents. Water-miscible organic content, such as methanol and acetonitrile in samples prior to loading has significant impact on PL recovery when eluting with methyl tert-butyl ether. However, isopropanol does not enhance the recovery of PL when adding to dichloromethane for elution. In addition, the compromise between improved extraction efficiency by SLE and reduced matrix effect is sometimes necessary to yield clean extract with acceptable recovery. The effective removal of phospholipids and reduction of matrix effect, while achieving good recovery for all pharmaceutical compounds with diverse physicochemical properties, demonstrated that SLE is a valuable alternative technique to liquid–liquid extraction (LLE) in high throughput LC–MS/MS based bioanalysis.
Source:Journal of Chromatography B, Volumes 891–892
Hongliang Jiang, Huachuan Cao, Yang Zhang, Douglas M. Fast
In past a few years, there has been a large increase in the application of supported liquid extraction (SLE) for LC–MS/MS based bioanalysis due to its distinct practical advantage in reduced time cost, ease of operation and the feasibility for automation. The main purpose of this study was to systematically evaluate supported liquid extraction in reducing matrix effect and improving extraction efficiency/recovery under various extraction conditions with 10 model pharmaceutical compounds in liquid chromatography coupled to electrospray tandem mass spectrometry (LC–ESI-MS/MS) analysis. Selected compounds have diverse physicochemical properties where log P ranges from 0.1 to 6.24 and pK a ranges from 4.0 to 11.1. The factors that may have the impact on the recovery of analytes and phospholipids (PL) were assessed. Over 75% recovery was achieved for every analyte under its respectively optimized extraction conditions where the selection of the polarity of extraction solvent and buffered pH can be critical for efficient recovery. Furthermore, the matrix effect was assessed by postextraction spike and postcolumn infusion method. The matrix effect was considerably reduced for all analytes under most extraction conditions evaluated for SLE, compared with protein precipitation (PPT) method. The correlation between matrix effect and residual phospholipids in sample extract was clearly shown. Although analyte-dependent matrix effect was observed prominently in sample extract prepared by PPT, it was minimized by SLE sample preparation process that effectively removes the majority of phospholipids. Sample extracted by ethyl acetate contained more phospholipids and demonstrated stronger matrix effect than by other organic solvents. Water-miscible organic content, such as methanol and acetonitrile in samples prior to loading has significant impact on PL recovery when eluting with methyl tert-butyl ether. However, isopropanol does not enhance the recovery of PL when adding to dichloromethane for elution. In addition, the compromise between improved extraction efficiency by SLE and reduced matrix effect is sometimes necessary to yield clean extract with acceptable recovery. The effective removal of phospholipids and reduction of matrix effect, while achieving good recovery for all pharmaceutical compounds with diverse physicochemical properties, demonstrated that SLE is a valuable alternative technique to liquid–liquid extraction (LLE) in high throughput LC–MS/MS based bioanalysis.
Plasma persistence of 2-aminothiazoline-4-carboxylic acid in rat system determined by liquid chromatography tandem mass spectrometry
23 March 2012,
10:56:29
Publication year:
2012
Source:Journal of Chromatography B, Volumes 891–892
Ilona Petrikovics, Jorn C.C. Yu, David E. Thompson, Prashanth Jayanna, Brian A. Logue, Jessica Nasr, Raj K. Bhandari, Steven I. Baskin, Gary Rockwood
2-Aminothiazoline-4-carboxylic acid (ATCA) was intravenously injected to rats in order to investigate its plasma distribution. ATCA was extracted from plasma samples by solid phase extraction (SPE) and molecularly imprinted polymer stir bar sorption extraction (MIP-SBSE). Detection and quantification of ATCA were achieved by using liquid chromatography–tandem mass spectrometry (LC–MS/MS). It was found that the intravenously injected ATCA concentration quickly decreased to half within 2.5h in the rat system. However, after 2.5h, the concentration of ATCA in plasma stayed constant at least 5 folds above the endogenous ATCA level for more then 48h. This finding can be used for evaluating ATCA's diagnostic and forensic value as a biomarker for cyanide exposure.
Source:Journal of Chromatography B, Volumes 891–892
Ilona Petrikovics, Jorn C.C. Yu, David E. Thompson, Prashanth Jayanna, Brian A. Logue, Jessica Nasr, Raj K. Bhandari, Steven I. Baskin, Gary Rockwood
2-Aminothiazoline-4-carboxylic acid (ATCA) was intravenously injected to rats in order to investigate its plasma distribution. ATCA was extracted from plasma samples by solid phase extraction (SPE) and molecularly imprinted polymer stir bar sorption extraction (MIP-SBSE). Detection and quantification of ATCA were achieved by using liquid chromatography–tandem mass spectrometry (LC–MS/MS). It was found that the intravenously injected ATCA concentration quickly decreased to half within 2.5h in the rat system. However, after 2.5h, the concentration of ATCA in plasma stayed constant at least 5 folds above the endogenous ATCA level for more then 48h. This finding can be used for evaluating ATCA's diagnostic and forensic value as a biomarker for cyanide exposure.
Detection of allantoin in clinical samples using hydrophilic liquid chromatography with stable isotope dilution negative ion tandem mass spectrometry
23 March 2012,
10:56:29
Publication year:
2012
Source:Journal of Chromatography B, Volumes 891–892
Rufus Turner, Lisa K. Stamp, Anthony J. Kettle
Allantoin is the major oxidation product of urate in humans and is a potential biomarker of oxidative stress. Several methods are used to measure allantoin in biological samples but they have inherent issues that can include lack of specificity and sensitivity, difficulty in sample preparation, or artefactual generation of allantoin. We have developed a method for measuring allantoin using hydrophilic liquid chromatography with stable isotope dilution tandem mass spectrometry (HILIC–MS/MS). It was validated for measuring allantoin in plasma, synovial fluid and urine from human subjects. The limit of quantification was determined to be 10fmol and the assay displayed excellent linearity for the wide range of concentrations found in clinical samples. Relative standard deviations were <5% for between-day and <7% for within-day variation. Accuracy was between 100% and 104%. Concentrations of allantoin in plasma of healthy controls (2.0μM; interquartile range 1.4–3.6μM, n =35) was significantly lower (p <0.001) than that in plasma from patients with rheumatoid arthritis (3.7μM; IQR 3.0–5.6μM, n =43) and in synovial fluid of patients with gout (3.3μM; IQR 2.8–5.8μM, n =10). This newer HILIC–MS/MS method is a simple and highly sensitive assay for detection of allantoin. It can be used to assess the level of oxidative stress in human pathologies.
Source:Journal of Chromatography B, Volumes 891–892
Rufus Turner, Lisa K. Stamp, Anthony J. Kettle
Allantoin is the major oxidation product of urate in humans and is a potential biomarker of oxidative stress. Several methods are used to measure allantoin in biological samples but they have inherent issues that can include lack of specificity and sensitivity, difficulty in sample preparation, or artefactual generation of allantoin. We have developed a method for measuring allantoin using hydrophilic liquid chromatography with stable isotope dilution tandem mass spectrometry (HILIC–MS/MS). It was validated for measuring allantoin in plasma, synovial fluid and urine from human subjects. The limit of quantification was determined to be 10fmol and the assay displayed excellent linearity for the wide range of concentrations found in clinical samples. Relative standard deviations were <5% for between-day and <7% for within-day variation. Accuracy was between 100% and 104%. Concentrations of allantoin in plasma of healthy controls (2.0μM; interquartile range 1.4–3.6μM, n =35) was significantly lower (p <0.001) than that in plasma from patients with rheumatoid arthritis (3.7μM; IQR 3.0–5.6μM, n =43) and in synovial fluid of patients with gout (3.3μM; IQR 2.8–5.8μM, n =10). This newer HILIC–MS/MS method is a simple and highly sensitive assay for detection of allantoin. It can be used to assess the level of oxidative stress in human pathologies.
Recovery of active anti TNF-α ScFv through matrix-assisted refolding of bacterial inclusion bodies using CIM monolithic support
23 March 2012,
10:56:29
Publication year:
2012
Source:Journal of Chromatography B, Volumes 891–892
Krishnan Sushma, Chuvappumkal Joseph Bilgimol, Mookambeswaran A. Vijayalakshmi, Padikara Kutty Satheeshkumar
Anti TNF-α molecules are important as therapeutic agents for many of the autoimmune diseases in chronic stage. Here we report the expression and purification of a recombinant single chain variable fragment (ScFv) specific to TNF-α from inclusion bodies. In contrast to the conventional on column refolding using the soft gel supports, an efficient methodology using monolithic matrix has been employed. Nickel (II) coupled to convective interaction media (CIM) support was utilized for this purpose with 6M guanidine hydrochloride (GuHCl) as the chaotropic agent. The protein purified after solubilization and refolding proved to be biologically active with an IC50 value of 15μg. To the best of our knowledge, this is the first report showing the application of methacrylate based chromatographic supports for matrix-assisted refolding and purification of Escherichia coli inclusion bodies. The results are promising to elaborate the methodology further to exploit the potential positive features of monoliths in protein refolding science.
Source:Journal of Chromatography B, Volumes 891–892
Krishnan Sushma, Chuvappumkal Joseph Bilgimol, Mookambeswaran A. Vijayalakshmi, Padikara Kutty Satheeshkumar
Anti TNF-α molecules are important as therapeutic agents for many of the autoimmune diseases in chronic stage. Here we report the expression and purification of a recombinant single chain variable fragment (ScFv) specific to TNF-α from inclusion bodies. In contrast to the conventional on column refolding using the soft gel supports, an efficient methodology using monolithic matrix has been employed. Nickel (II) coupled to convective interaction media (CIM) support was utilized for this purpose with 6M guanidine hydrochloride (GuHCl) as the chaotropic agent. The protein purified after solubilization and refolding proved to be biologically active with an IC50 value of 15μg. To the best of our knowledge, this is the first report showing the application of methacrylate based chromatographic supports for matrix-assisted refolding and purification of Escherichia coli inclusion bodies. The results are promising to elaborate the methodology further to exploit the potential positive features of monoliths in protein refolding science.
Separation of nucleobases and their derivatives with organic-high ionic strength aqueous phase systems by spiral high-speed counter-current chromatography
23 March 2012,
10:56:29
Publication year:
2012
Source:Journal of Chromatography B, Volumes 891–892
Yoichi Shibusawa, Akio Yanagida, Atsushi Ogihara, Ying Ma, Xiaoyuan Chen, Yoichiro Ito
A set of nucleic acid constituents were separated with ultra polar two-phase solvent systems by a spiral multilayer coil mounted on the rotary frame of a type-J coil planet centrifuge. These two-phase systems were composed of 1-butanol/ethanol/50% saturated aqueous ammonium sulfate at various volume ratios. Nucleobases including adenine, cytosine, uracil, and thymine; nucleosides including adenosine, guanosine, cytidine, and uridine; and nucleotides including, AMP, GMP, CMP, UMP, and TMP are partitioned in each group with suitable solvent ratios. Adenine derivatives such as adenosine, AMP, ADP, and ATP were well resolved in the most polar solvent system composed of ethanol/50% saturated aqueous ammonium sulfate at a volume ratio of 1:2. It was found that cytosine and cytidine peaks showed some irregular two peaks probably due to their keto and enol isomers, while the separation of AMP forms two peaks especially when TMP was added in the sample solution, the mechanism of which is now under investigation in our laboratory.
Source:Journal of Chromatography B, Volumes 891–892
Yoichi Shibusawa, Akio Yanagida, Atsushi Ogihara, Ying Ma, Xiaoyuan Chen, Yoichiro Ito
A set of nucleic acid constituents were separated with ultra polar two-phase solvent systems by a spiral multilayer coil mounted on the rotary frame of a type-J coil planet centrifuge. These two-phase systems were composed of 1-butanol/ethanol/50% saturated aqueous ammonium sulfate at various volume ratios. Nucleobases including adenine, cytosine, uracil, and thymine; nucleosides including adenosine, guanosine, cytidine, and uridine; and nucleotides including, AMP, GMP, CMP, UMP, and TMP are partitioned in each group with suitable solvent ratios. Adenine derivatives such as adenosine, AMP, ADP, and ATP were well resolved in the most polar solvent system composed of ethanol/50% saturated aqueous ammonium sulfate at a volume ratio of 1:2. It was found that cytosine and cytidine peaks showed some irregular two peaks probably due to their keto and enol isomers, while the separation of AMP forms two peaks especially when TMP was added in the sample solution, the mechanism of which is now under investigation in our laboratory.
In vitro enantioselective metabolism of TJ0711 hydrochloride by human liver microsomes using a novel chiral liquid chromatography–tandem mass spectrometry method
23 March 2012,
10:56:29
Publication year:
2012
Source:Journal of Chromatography B, Volumes 891–892
Jiangeng Huang, Lei Hu, Li Xu, Minghui Sun, Zhaoze Fan, Jun Qiu, Gao Li, Luqin Si
A novel liquid chromatography–tandem mass spectrometry (LC–MS/MS) method employing chiral analytical techniques was developed and validated for in vitro enantioselective metabolic stability study of racemic 1-[4-(2-methoxyethyl) phenoxy]-3-[[2-(2-methoxyphenoxy) ethyl]amino]-2-propanol hydrochloride (TJ0711 HCl), a newly developed vasodilatory β-blocker. Robust enantiomeric separations were achieved on a chiral SUMICHIRAL OA-2500 column using ethanol and hexane (40:60, v/v) as a mobile phase. Metabolic stability results demonstrated that both TJ0711 enantiomers underwent a rapid phase I metabolism, but preferential metabolism of R-TJ0711 was observed. Our previously reported ultra-performance liquid chromatography-multiple reaction monitoring-information dependent acquisition-enhanced product ion (UPLC-MRM-IDA-EPI) method was finally chosen for metabolite profiling study of TJ0711 enantiomers, because the newly developed HPLC-based method resulted in compromised chromatographic separation, particularly for TJ0711 metabolites. A number of metabolic products were detected and the structures of formed metabolites were predicted. Similar to racemic TJ0711 HCl, demethylation and hydroxylation were proposed to be the principle metabolism pathways during in vitro incubations of each enantiomer with human liver microsomes.
Source:Journal of Chromatography B, Volumes 891–892
Jiangeng Huang, Lei Hu, Li Xu, Minghui Sun, Zhaoze Fan, Jun Qiu, Gao Li, Luqin Si
A novel liquid chromatography–tandem mass spectrometry (LC–MS/MS) method employing chiral analytical techniques was developed and validated for in vitro enantioselective metabolic stability study of racemic 1-[4-(2-methoxyethyl) phenoxy]-3-[[2-(2-methoxyphenoxy) ethyl]amino]-2-propanol hydrochloride (TJ0711 HCl), a newly developed vasodilatory β-blocker. Robust enantiomeric separations were achieved on a chiral SUMICHIRAL OA-2500 column using ethanol and hexane (40:60, v/v) as a mobile phase. Metabolic stability results demonstrated that both TJ0711 enantiomers underwent a rapid phase I metabolism, but preferential metabolism of R-TJ0711 was observed. Our previously reported ultra-performance liquid chromatography-multiple reaction monitoring-information dependent acquisition-enhanced product ion (UPLC-MRM-IDA-EPI) method was finally chosen for metabolite profiling study of TJ0711 enantiomers, because the newly developed HPLC-based method resulted in compromised chromatographic separation, particularly for TJ0711 metabolites. A number of metabolic products were detected and the structures of formed metabolites were predicted. Similar to racemic TJ0711 HCl, demethylation and hydroxylation were proposed to be the principle metabolism pathways during in vitro incubations of each enantiomer with human liver microsomes.
Simultaneous determination of pimpinellin, isopimpinellin and phellopterin in rat plasma by a validated UPLC–MS/MS and its application to a pharmacokinetic study after administration of Toddalia asiatica extract
23 March 2012,
10:56:29
Publication year:
2012
Source:Journal of Chromatography B, Volumes 891–892
Zhigang Liu, Minyan Jiang, Xiumei Lu, Feng Qin, Yang Song, Jing Wen, Famei Li
A rapid and selective ultra performance liquid chromatography–tandem mass spectrometry (UPLC–MS/MS) method was developed for simultaneous determination of three bioactive coumarins of Toddalia asiatica extract including pimpinellin, isopimpinellin and phellopterin in rat plasma for the first time. Phenacetin was used as the internal standard (IS). Plasma samples were extracted by liquid–liquid extraction with methyl tert-butyl ether. The chromatographic separation was carried out on an ACQUITY UPLC™ BEH C18 column with an isocratic mobile phase consisting of methanol-5mmol/L ammonium acetate (65:35, v/v). The detection was performed on a triple quadrupole tandem mass spectrometer by multiple reaction monitoring (MRM) via electrospray ionization (ESI) source with positive ionization mode. The method was linear for all analytes over investigated range with all correlation coefficients greater than 0.9942. The lower limits of quantification (LLOQ) were 25.0ng/mL for pimpinellin, 10.0ng/mL for isopimpinellin and 5.00ng/mL for phellopterin. The intra- and inter-day precision (RSD%) was within 12% and the accuracy (RE%) ranged from −2.3% to 5.5%. The rapid and sensitive method was fully validated and successfully applied to the pharmacokinetic study of pimpinellin, isopimpinellin and phellopterin in rats following oral administration of Toddalia asiatica extract.
Source:Journal of Chromatography B, Volumes 891–892
Zhigang Liu, Minyan Jiang, Xiumei Lu, Feng Qin, Yang Song, Jing Wen, Famei Li
A rapid and selective ultra performance liquid chromatography–tandem mass spectrometry (UPLC–MS/MS) method was developed for simultaneous determination of three bioactive coumarins of Toddalia asiatica extract including pimpinellin, isopimpinellin and phellopterin in rat plasma for the first time. Phenacetin was used as the internal standard (IS). Plasma samples were extracted by liquid–liquid extraction with methyl tert-butyl ether. The chromatographic separation was carried out on an ACQUITY UPLC™ BEH C18 column with an isocratic mobile phase consisting of methanol-5mmol/L ammonium acetate (65:35, v/v). The detection was performed on a triple quadrupole tandem mass spectrometer by multiple reaction monitoring (MRM) via electrospray ionization (ESI) source with positive ionization mode. The method was linear for all analytes over investigated range with all correlation coefficients greater than 0.9942. The lower limits of quantification (LLOQ) were 25.0ng/mL for pimpinellin, 10.0ng/mL for isopimpinellin and 5.00ng/mL for phellopterin. The intra- and inter-day precision (RSD%) was within 12% and the accuracy (RE%) ranged from −2.3% to 5.5%. The rapid and sensitive method was fully validated and successfully applied to the pharmacokinetic study of pimpinellin, isopimpinellin and phellopterin in rats following oral administration of Toddalia asiatica extract.
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