Journal of Chromatography B (v.961, #C)
Editorial Board (i).
LC–MS analysis of polyclonal IgGs using IdeS enzymatic proteolysis for oxidation monitoring by Yann Leblanc; Marie Romanin; Nicolas Bihoreau; Guillaume Chevreux (1-4).
Susceptibility of IgGs to oxidation is a significant issue for intravenous immunoglobulin preparations (IVIG) in liquid solution and raises both safety and efficacy concerns. Here we present an optimized chromatography method coupled to mass spectrometry (MS) to determine the oxidation of Fc/2 fragments derived from polyclonal IgGs after IdeS treatment. Separation of the four IgG subclasses was achieved using a diphenyl column and UV/MS detections were used for quantification and characterization. Several oxidized Fc/2 fragments generated by stress conditions were resolved and oxidized methionines were identified. This procedure can be used to monitor the oxidative status of IVIG preparations during formulation or stability studies.
Keywords: IdeS; Mass spectrometry; IVIG; Oxidation; IgG;
Ionic liquid-based aqueous two-phase system extraction of sulfonamides in milk by Mingyuan Shao; Xuli Zhang; Na Li; Jiayuan Shi; Huijie Zhang; Zhibing Wang; Hanqi Zhang; Aimin Yu; Yong Yu (5-12).
A simple method for the determination of six sulfonamides (SAs) in milk samples was developed. 1-Butyl-3-methylimidazolium tetrafluoroborate and trisodium citrate dihydrate were used to form aqueous two-phase system. The aqueous two phase system was applied to the extraction of the SAs and the determination of the analytes was performed by high-performance liquid chromatography. To achieve optimum extraction performance, several experimental parameters, including the type and the amount of salt, the type and amount of ionic liquid, ultrasonic time and pH of sample solution, were investigated and optimized. Under the optimal experimental conditions, good linearity was observed in the range of 8.55–1036.36 ng mL−1. The limits of detection and quantification were in the range of 2.04–2.84 and 6.73–9.37 ng mL−1, respectively. The present method was successfully applied to the determination of SAs in milk samples, and the recoveries of analytes were in the range of 72.32–108.96% with relative standard deviations ranging from 0.56 to 12.20%. The results showed that the present method was rapid, feasible and environmentally friendly.
Keywords: Aqueous two-phase system; Trisodium citrate dihydrate; Sulfonamides; Ionic liquid; High-performance liquid chromatography; Dairy milk;
High performance liquid chromatography–tandem mass spectrometric assay of dexmedetomidine in plasma, urine and amniotic fluid samples for pregnant ewe model by Z. Cui; D.S.-L. Chow; L. Wu; D.A. Lazar; R. Rodrigo; O.O. Olutoye; O.A. Olutoye (13-19).
Dexmedetomidine (DEX; Precedex®), approved by the Food and Drug Administration (FDA) in 1999 as a sedative for use in the intensive care unit, is a potent and highly selective α2-adrenoceptor agonist with significant sedative, analgesic and anxiolytic effects. However, the research of DEX use during pregnancy is limited and the impact of DEX on the fetal development is unclear. This article describes a high performance liquid chromatography–tandem mass spectrometry (HPLC–MS/MS) assay suitable for various biomatrices of plasma, urine and amniotic fluid, as a prerequisite for pharmacokinetic characterization of DEX in the pregnant ewe model. DEX and testosterone (internal standard; IS) were extracted from 200 μL of plasma, urine or amniotic fluid with ethyl acetate. The HPLC resolution was achieved on an Agilent ZORBAX SB-CN column with a gradient elution at a flow rate of 0.5 mL/min using a mobile phase of 5–100% of acetonitrile with 0.5% formic acid (mobile phase B) in water (mobile phase A). The detection was performed by a triple quadrupole tandem mass spectrometer with positive electrospray ionization. The precursor/product transitions (m/z) in the positive ion mode [M+H]+ were m/z 201.5 → 95.4 for DEX and m/z 289.2 → 109.1 for IS. The method was validated in the concentration range of 25 (lower limit of quantification; LLOQ)–5000 pg/mL for both maternal and fetal plasma, and of 50 (LLOQ)–5000 pg/mL for urine and amniotic fluid, respectively. The intra- and inter-day precision and accuracy were within ±9%. The overall recoveries of DEX were 82.9–87.2%, 85.7–88.4%, 86.2–89.7% and 83.7–88.1% for maternal plasma, urine, fetal plasma and amniotic fluid, respectively. The percentage matrix factors in different biomatrices were less than 120%. Stability studies demonstrated that DEX was stable after three freeze/thaw cycles, in the autosampler tray at 20 °C for 24 h and during the 3 h sample preparation at room temperature. The validated HPLC–MS/MS method has been successfully employed for pharmacokinetic evaluation of DEX in pregnant ewes and fetuses.
Keywords: Dexmedetomidine; HPLC–MS/MS; Pregnant ewe; Plasma; Urine; Amniotic fluid;
A sensitive liquid chromatography–tandem mass spectrometry method for pharmacokinetics and tissue distribution of nuciferine in rats by Shengying Gu; Guanhua Zhu; Yuzhu Wang; Qin Li; Xin Wu; Jigang Zhang; Gaolin Liu; Xiaoyu Li (20-28).
Nuciferine is an important drug candidate for the treatment of obesity-related diseases. However, few investigations have been conducted about the pharmacokinetics and tissue distribution of nuciferine to better understand its behavior and action mechanism in vivo. Thus, a sensitive and reliable liquid chromatography with tandem mass spectrometry (HPLC–MS/MS) method was established and validated for the quantification of nuciferine in rat plasma and tissue samples. The validated method was successfully applied to the pharmacokinetic and tissue distribution study of nuciferine in rats. One-compartmental pharmacokinetic parameters indicated that nuciferine had rapid distribution, extensive tissue uptake, and poor absorption into systemic circulation. The values of absolute bioavailability were (3.8 ± 1.4)%, (4.2 ± 1.3)% and (3.9 ± 1.0)% after oral administration of 2.0, 5.0 and 10.0 mg/kg nuciferine and intravenous administration of 0.2 mg/kg nuciferine in rats. The results of the tissue distribution study suggested that nuciferine was distributed into the brain, liver and adipose tissue after intravenous administration. In conclusion, the present study may provide a material basis for study of the pharmacological action of nuciferine in the treatment of obesity, and meaningful insights into further study on dosage modification.
Keywords: Nuciferine; LC–MS/MS; Pharmacokinetics; Tissue distribution;
Simultaneous determination of four sulfur mustard–DNA adducts in rabbit urine after dermal exposure by isotope-dilution liquid chromatography–tandem mass spectrometry by Yajiao Zhang; Lijun Yue; Zhiyong Nie; Jia Chen; Lei Guo; Bidong Wu; Jianlin Feng; Qin Liu; Jianwei Xie (29-35).
Sulfur mustard (SM) is a classic vesicant agent, which has been greatly employed in several wars or military conflicts. The most lesion mechanism is its strong alkylation of DNAs in vivo. Until now there are four specific DNA adducts of SM identified for further retrospective detection, i.e., N 7-(2-hydroxyethylthioethyl)-2′-guanine (N 7-HETEG), bis(2-ethyl-N 7-guanine)thioether (Bis-G), N 3-(2-hydroxyethylthioethyl)-2′-adenine (N 3-HETEA) and O 6-(2-hydroxyethylthioethyl)-2′-guanine (O 6-HETEG), respectively. Here, a novel and sensitive method of isotope-dilution ultrahigh performance liquid chromatography–tandem mass spectrometry (UPLC–MS/MS) combining with solid phase extraction was reported for the simultaneous determination of four SM–DNA adducts. A lower limit of detection of 2–5 ng L−1, and a lower limit of quantitation of 5–10 ng L−1 were achieved, respectively, and the recoveries ranged from 87% to 116%. We applied this method in the determination of four SM–DNA adducts in rabbit urine after dermal exposure by SM in three dose levels (2, 5, 15 mg kg−1), so as to investigate the related metabolic behavior in vivo. For the first time, in SM exposed rabbit urine, our results revealed the relative accumulation abundance of four SM–DNA adducts, i.e., 67.4% for N 7-HETEG, 22.7% for Bis-G, 9.8% for N 3-HETEA, 0.1% for O 6-HETEG, and significant dose and time dependent responses of these SM–DNA adducts. The four adducts were detectable after 8 h, afterwards, their contents continuously increased, achieved maximum in the first two or three days and then gradually decreased till the end of one month. Meanwhile, the amounts of SM–DNA adducts were positively correlated with the exposure doses.
Keywords: Sulfur mustard; DNA adducts; Isotope-dilution ultrahigh performance liquid chromatography–tandem mass spectrometry; Solid-phase extraction; Metabolism;
Determination of homocysteine thiolactone in urine by field amplified sample injection and sweeping MEKC method with UV detection by Paulina Furmaniak; Paweł Kubalczyk; Rafał Głowacki (36-41).
Homocysteine thiolactone (Hcy-thiolactone), an intramolecular thioester, easily acylates free-amino groups in proteins, which impairs or alters the protein's biological function. Here, we describe new capillary electrophoresis assay for the determination of Hcy-thiolactone in human urine based on a field amplified sample injection and sweeping MEKC with UV detection. The two steps procedure relies on sample liquid–liquid extraction followed by CE separation and UV detection at 240 nm. The Hcy-thiolactone standard added to the urine before the extraction step shows that the response of the detector is linear within the range studied, from 0.1 to 1 μmol L−1 urine. The intra- and interday precision and recovery were 3.2–14.4% (average 5.1% and 9.3%) and 92.5–112.6% (average 99.8% and 99.1%), respectively. The lower limit of quantification was 0.09 nmol Hcy-thiolactone in 1 mL of urine. The proposed method was applied for the analysis of 15 urine samples donated by apparently healthy volunteers. The average concentration of the analyte was 0.170 ± 0.029 μmol L−1.
Keywords: Capillary electrophoresis; Homocysteine thiolactone; Ultraviolet detection, Urine;
Quantitative determination of the anti-tumor agent tasquinimod in human urine by liquid chromatography–tandem mass spectrometry by Nico C. van de Merbel; Peter Walland; Mikael Tiensuu; Carl J. Sennbro (42-48).
Tasquinimod is an anti-tumor drug that is currently in clinical development for the treatment of solid cancers. After oral administration, tasquinimod and a number of its metabolites are excreted in the urine. The quantitative determination of tasquinimod in urine is challenging because of the required sensitivity (down to 0.1 nM or 40 pg/mL), the highly variable nature of this biological matrix and the presence of potentially unstable metabolites, which may convert back to the parent drug. In this article, an LC–MS/MS method is described for the determination of tasquinimod in human urine in the concentration range 0.1–200 nM. Liquid–liquid extraction with n-chlorobutane was used to extract tasquinimod from 100 μL human urine and to remove interfering endogenous urinary constituents. Reversed-phase liquid chromatography coupled to a triple quadrupole mass spectrometer equipped with an ESI source was used for quantification of tasquinimod in a 2.5-min run. A stable-isotope labeled internal standard was used for response normalization. The intra- and inter-day coefficients of variation (precision) as well as the bias (accuracy) of the method were below 7%. Although considerable conversion of conjugated tasquinimod metabolites back to parent drug was observed when incurred samples were stored at 37 °C for a prolonged time, tasquinimod as well as its metabolites were sufficiently stable under all relevant sampling, storage and analysis conditions. The method was successfully applied to determine the urinary excretion of tasquinimod in healthy volunteers and patients with renal impairment after a 0.5-mg oral dose.
Keywords: Tasquinimod; Prostate cancer; Urine analysis; LC–MS/MS; Incurred sample stability;
A chiral high-performance liquid chromatography–tandem mass spectrometry method for the stereospecific determination of morinidazole in human plasma by Kan Zhong; Zhiwei Gao; Qin Li; Dafang Zhong; Xiaoyan Chen (49-55).
Morinidazole is a novel 5-nitroimidazole derivative used for the treatment of amoebiasis, trichomoniasis, and anaerobic bacterial infections. Morinidazole possesses a chiral carbon and is clinically administered as a racemate. In the present study, an enantioselective and sensitive liquid chromatography–tandem mass spectrometry method of determining morinidazole enantiomers in human plasma was developed and validated to characterize the stereoselective pharmacokinetics. Plasma samples were processed by liquid–liquid extraction using tert-butyl methyl ether. Chiral separation was optimized within 8.5 min on a cellulose column using an isocratic mobile phase of methanol/water (80:20, v/v). Detection was using mass spectrometry in multiple reaction monitoring mode, using the transitions of m/z 271 → 144 for morinidazole enantiomers, and m/z 275 → 148 for d4-morinidazole (internal standard). The calibration curves were linear over 5.00–6000 ng/mL for each enantiomer. The lower limit of quantification for each enantiomer was established at 5.00 ng/mL. Intra- and inter-day precisions were less than 6.4% for each enantiomer in terms of relative standard deviation, and accuracies were between −2.5% and 6.4% in terms of relative error for each enantiomer. No chiral inversion was observed during sample storage, preparation procedure and analysis. Major glucuronide and sulfate conjugates were not observed to interfere with the determination of morinidazole enantiomers. The method was applied to study the stereoselective pharmacokinetics of morinidazole in humans. Moderate stereoselectivity was observed in healthy subjects and patients with severe renal impairment.
Keywords: Morinidazole enantiomer; Liquid chromatography–tandem mass spectrometry; Stereoselective pharmacokinetics; 5-Nitroimidazole antimicrobial drug;
Screening for selective inhibitors of xanthine oxidase from Flos Chrysanthemum using ultrafiltration LC–MS combined with enzyme channel blocking by Hui-Peng Song; Hui Zhang; Yu Fu; Hua-yan Mo; Mu Zhang; Jun Chen; Ping Li (56-61).
In this study, a new method based on ultrafiltration liquid chromatography-mass spectrometry (UF-LC–MS) combined with enzyme channel blocking (ECB) was developed to discover bioactive components from herbal medicines. Xanthine oxidase (XOD), a critical enzyme for treating gout, was employed as the target protein for screening. By comparing chromatographic profiles of the compounds binding to XOD before and after the ECB experiment, the selective ligands could be distinguished from the non-selective binders. In this experiment, febuxostat bound to the channel entering into the active site of the enzyme and prevented potential ligands from binding. Finally, four compounds, namely, luteolin-7-O-glucoside, apigenin-7-O-glucoside, luteolin and apigenin were screened and identified as the candidate XOD inhibitors based on the ultrafiltration chromatogram of Flos Chrysanthemum, a famous traditional Chinese medicine used in many prescriptions for gout treatment. To verify the compounds screened further, a microplate method was applied to evaluate their enzyme inhibitory activities. The IC50 values of the above 4 compounds were 23.61, 38.80, 1.54 and 1.96 μM, respectively. The structure-function relationship was also estimated according to the in vitro assay. The results were in favor of the hypothesis that the Flos Chrysanthemum extract might be used for gout treatment by inhibiting XOD.
Keywords: Xanthine oxidase; Ultrafiltration; Liquid chromatography/mass spectrometry; Flos Chrysanthemum; Febuxostat;
Development and validation of an LC–MS/MS based method for quantification of 25 hydroxyvitamin D2 and 25 hydroxyvitamin D3 in human serum and plasma by Stanley (Weihua) Zhang; Wenying Jian; Sheryl Sullivan; Banu Sankaran; Richard W. Edom; Naidong Weng; David Sharkey (62-70).
Vitamin D deficiency is increasing in the general population and has become a serious public health risk globally. As a reliable clinical indicator of vitamin status, 25 hydroxyvitamin D (25(OH)D) has been measured by various methods. However, the accuracy of these measurements has been the subject of considerable debate. Here, we report the development and validation of a liquid chromatography–triple quadrupole mass spectrometry based method for the quantification of 25(OH)D2 and 25(OH)D3 in human serum and plasma samples. Samples were first processed by protein precipitation to release the analytes from the vitamin D binding protein (DBP), followed by a liquid–liquid extraction procedure. Analysis was performed on an LC–MS/MS system which utilized an AB Sciex API 3000 mass spectrometer. A six point calibration curve ranging from 2.5 to 100 ng/mL was established for both 25(OH)D2 and 25(OH)D3. A complete method validation was conducted, including intra- and inter-assay accuracy and precision, LLOQ, dilution QC, specificity, recovery, matrix effect, and a thorough stability profile of stock solutions and QC samples. Matching samples of serum and plasma (containing either heparin or EDTA anticoagulant) generated from the same blood samples were tested, and no significant differences in 25(OH)D2 and 25(OH)D3 concentrations were found in these sample matrices. In method comparison, we analyzed 10 serum samples obtained from the Vitamin D External Quality Assessment Scheme (DEQAS), and the total 25(OH)D concentrations measured by our method were very close to the LC–MS/MS Method Mean values provided by DEQAS (average 0.17% bias, R 2 = 0.99). However, comparison with the DiaSorin Liaison 25(OH)D TOTAL Assay demonstrated limited correlation between these two methods (R 2 = 0.54). In general, concentrations measured by our LC–MS/MS method were roughly 9% higher than those measured by the DiaSorin Liaison assay. The correlation with DiaSorin Liaison measurement was better for samples in the lower concentration range. In summary, we developed and validated an LC–MS/MS based method that can be reliably applied in routine quantification of 25(OH)D2 and 25(OH)D3 in human serum and plasma samples. This method is not suitable for pediatric determinations due to the potential interference of 3-epi 25(OH)D3.
Keywords: 25-Hydroxyvitamin D; LC–MS/MS; Validation; Matrix comparison; Method comparison; Stability;
Determination of meropenem in bacterial media by LC–MS/MS by Liusheng Huang; Janus Haagensen; Davide Verotta; Patricia Lizak; Francesca Aweeka; Katherine Yang (71-76).
To support the development of a dynamic in vitro human pharmacokinetic/pharmacodynamic simulation model for biofilm-mediated infections and study stability of meropenem, an LC-MS/MS method for the determination of meropenem in Luria Bertani (LB) media was developed and validated in an API2000 LC-MS/MS system. A partial validation was also performed in M9 media. Sample aliquots of 100 μL (or 25 μL for M9 media) were mixed with the internal standard (IS) ceftazidime and filtered. The filtrate was directly injected onto a C8 column eluted with ammonium formate (10 mM, pH 4) and acetonitrile (0.1% formic acid) in a gradient mode. ESI+ and MRM with ion pair m/z 384→68 for meropenem and m/z 547→468 for the IS were used for quantification. The calibration curve concentration range was 50 to 25,000 ng/mL. The recovery was over 98%. In LB media, significant signal suppression was observed throughout the time period of detection when compared with mobile phase solvents, but the matrix effect was compensated well with the IS. In M9 media, much less signal suppression was observed. The method is simple, fast, and reliable. Using the method, stability of meropenem in LB and M9 media were tested. No significant degradation was observed for at least 8 h in both LB media (37 °C) and M9 media (30 °C), but more than 15% degradation was observed overnight (∼20 h). The method was transferred to an API5000 LC–MS/MS system using meropenem-d6 as the IS.
Keywords: LC–MS/MS; Meropenem; M9; LB media; Stability;
Liquid chromatography/tandem mass spectrometry method for simultaneous determination of cocaine and its metabolite (−)ecgonine methyl ester in human acidified stabilized plasma samples by Yongzhen Liu; Bo Zheng; Stephanie Strafford; Ravi Orugunty; Michael Sullivan; Jeffrey Gus; Christian Heidbreder; Paul J. Fudala; Azmi Nasser (77-85).
Two simple, sensitive and rapid liquid chromatography/electrospray ionization tandem mass spectrometry (LC–ESI-MS/MS) methods (low range and high range) were developed and validated for the quantification of cocaine and its metabolite (−)ecgonine methyl ester (EME) in human acidified stabilized plasma samples. In the low range assay, cocaine and the internal standard, cocaine-D3, were extracted using a single step liquid–liquid extraction from human acidified stabilized plasma. For the high range assay, human acidified stabilized plasma containing cocaine, EME, and the internal standards, cocaine-D3 and EME-D3, was mixed with acetonitrile, and the protein precipitate was separated by centrifugation. Both cocaine and EME extracted from both assays were separated on a HILIC column and detected in positive ion mode using multiple reaction monitoring (MRM). Both methods were validated and the specificity, linearity, lower limit of quantitation (LLOQ), precision, accuracy, recoveries and stability were determined. The linear range for the low range assay was 0.01–5 ng/mL for cocaine; in the high range assay values were 5–1000 ng/mL for cocaine and 1–200 ng/mL for EME. The correlation coefficient (R 2) values for both assays were 0.993 or greater. The precision and accuracy for intra-day and inter-day were better than 13.0%. The recovery was above 85% and matrix effects were low with the matrix factor ranging from 0.817 to 1.10 for both analytes in both assays. The validated methods were successfully used to quantify the plasma concentrations of cocaine and EME in clinical pharmacokinetic and pharmacodynamic studies.
Keywords: Cocaine; Ecgonine methyl ester; LC–MS/MS; Human acidified stabilized plasma;
Determination of domperidone in human plasma using high performance liquid chromatography with fluorescence detection for clinical application by Tsuneaki Yoshizato; Kimiko Tsutsumi; Tsutomu Kotegawa; Hiromitsu Imai; Shigeyuki Nakano (86-90).
A simple and reliable method for the determination of domperidone in human plasma has been developed. Plasma samples (1 mL) were pre-purified by a solid-phase extraction with Bond Elut® C18. The separation was achieved with XBridge™ C18 column (150 mm × 4.6 mm i.d., 5 μm) at 40 °C. The mobile phase was a mixture of acetonitrile and 10 mM ammonium acetate buffer (36:64, v/v), adjusted to pH 9.4 with 20% ammonium solution at a flow rate of 1.0 mL/min. The peak was detected using fluorescence detector at excitation 282 nm and emission 328 nm. Retention times for domperidone and internal standard (propranolol) were 8.3 min and 11.2 min, respectively. The method showed a good linearity (r > 0.999), precision (relative standard deviations <10.6%), and extraction recovery (85.7–99.7%) over a concentration of 1–100 ng/mL. The lower limit of quantification (LLOQ) was 1.0 ng/mL. This proposed method was successfully applied to a pharmacokinetic interaction study of domperidone in healthy Japanese volunteers.
Keywords: Domperidone; Itraconazole; HPLC; Solid phase extraction; Fluorescence detection; MDR1;
Highly sensitive, selective and rapid LC–MS method for simultaneous quantification of diadenosine polyphosphates in human plasma by Anna Schulz; Vera Jankowski; Walter Zidek; Joachim Jankowski (91-96).
Diadenosine polyphosphates (Ap n As) are endogenous mediators involved in large number of physiologic and pathophysiologic processes. The quantification of diadenosine polyphosphates in plasma and biological matrices is still challenging. Therefore, there is an urgent need for a simple and reliable quantification method suitable for clinical studies. The classical quantification of diadenosine polyphosphates is based on chromatographic separation and UV adsorption of the resulting fractions. These procedures are associated with low selectivity due to co-eluting plasma components. Therefore, we developed and validated a highly sensitive, selective and rapid LC–ESI–MS method for simultaneous quantification of Ap n As (with n = 3–6) in human plasma within this study. The identities of the endogenous Ap n As (with n = 3–6) were revealed by comparison of ESI-MS/MS fragment spectra of isolated endogenous compounds with those of authentic Ap n As.Diadenosine polyphosphates were extracted from 100 μl human plasma using weak anion-exchange extraction cartridges. The separation of Ap n As was achieved using capillary C18 columns. ESI-HCT mass spectrometer (Bruker Daltonik, Germany) operated in negative ion mode was used for detection and quantification of Ap n As.A calibration curve was established for diadenosine polyphosphate free plasma in the concentration range 1.9–125 nM (r 2 > 0.998) for all analytes. The intra- and inter-day accuracies were in the range of 91.4% and 110.9%. The intra- and inter-day precisions were determines as 0.1% and 11.4%, respectively. The mean plasma concentrations of Ap n As were quantified as 31.9 ± 5.9 nM for Ap3A, 40.4 ± 6.6 nM for Ap4A, 10.7 ± 1.5 nM for Ap5A and 10.0 ± 18.9 nM for Ap6A.The developed and validated ESI MS-based method for quantification of diadenosine polyphosphates in human plasma was successfully evaluated within the study. Conclusion Since the quantification is based on a volume of 100 μl plasma, this method is highly applicable for clinical applications aiming at the validation of the impact of highly physiological and pathophysiological active diadenosine polyphosphates.
Keywords: Dinucleoside polyphosphates; Mediators; Quantification; Biomolecule; Hypertension;
Determination of free vancomycin, ceftriaxone, cefazolin and ertapenem in plasma by ultrafiltration: Impact of experimental conditions by Alexander Kratzer; Uwe Liebchen; Michael Schleibinger; Martin G. Kees; Frieder Kees (97-102).
Ultrafiltration is a rapid and convenient method to determine the free concentrations of drugs. In the present work, we aimed to develop an ultrafiltration method which is appropriate for routine determination of the free fraction of vancomycin and highly protein bound beta-lactams such as ertapenem, ceftriaxone and cefazolin in plasma from intensive care unit patients. Different filter types and experimental conditions (molecular weight cut-off, centrifugal force and time, pH, temperature) were evaluated and found to have influence on the result. In the final protocol, serum or plasma was buffered to pH 7.4–7.5, ultrafiltered at 1000 × g at 37 °C for 20 min using Nanosep Omega 10 K filters and subsequently analysed for the antibiotics by RP-HPLC with UV detection. The data from our investigation suggest to aim physiological conditions, i.e. 37 °C and pH 7.4, and low to moderate relative centrifugal forces in order to get reliable results. With regard to the chromatographic separation, modulation of the pH in the range of 2.5–7.0 allows to determine several beta-lactams isocratically and/or to avoid interferences by co-administered drugs.
Keywords: Protein binding; Beta-lactam; pH; Temperature; Relative centrifugal force; HPLC;
A simple and accurate liquid chromatography–tandem mass spectrometry method for quantification of zonisamide in plasma and its application to a pharmacokinetic study by Kamal M. Matar (103-109).
Zonisamide (ZNM) is an antiepileptic drug that is used as an adjunctive therapy in the treatment of adults with partial seizures. An LC–MS/MS method for quantification of ZNM in human and rabbit plasma using 2H4,15N-Zonisamide as an internal standard (IS) has been developed and validated. The drug and IS were extracted by ether and analyzed on Symmetry® C18 column. Quantitation was achieved using ESI-interface employing MRM mode. The method was validated over the concentration range of 0.5–50 μg/mL and 0.5–30 μg/mL (r 2 > 0.99) in human and rabbit plasma samples, respectively. Intra- and inter-run precision of ZNM assay in human and rabbit plasma samples ranged from 0.8 to 8.5% with accuracy (bias) varied from −11.3 to 14.4% indicating good precision and accuracy. Stability of ZNM in human and rabbit plasma samples at various conditions showed that the drug was stable under the studied conditions. Analytical recoveries of ZNM and IS from spiked human and rabbit plasma samples were in the range of 70.8–77.3% and 85.6–110.4%, respectively. Matrix effect study showed a lack of matrix effect on mass ions of ZNM and IS. The developed method was successfully applied for a pharmacokinetic study by measuring ZNM in rabbit plasma samples. Moreover, the method is routinely utilized for TDM of ZNM.
Keywords: Zonisamide; LC–MS/MS; Human; Rabbit; Plasma; Pharmacokinetics;
Corrigendum to “Liquid chromatography and ion trap mass spectrometry for simultaneous and multiclass analysis of antimicrobial residues in feed water” [J. Chromatogr. B 945–946 (2014) 31–38] by Chusak Ardsoongnearn; Ongart Boonbanlu; Sunan Kittijaruwattana; Leena Suntornsuk (110).
Corrigendum to “Potentially antioxidant compounds indicated from Mallotus and Phyllanthus species fingerprints” [J. Chromatogr. B 910 (2012) 114–121] by S. Thiangthum; B. Dejaegher; M. Goodarzi; C. Tistaert; A.Y. Gordien; N. Nguyen Hoai; M. Chau Van; J. Quetin-Leclercq; L. Suntornsuk; Y. Vander Heyden (111).