Journal of Chromatography B (v.799, #2)

OFC: Update (OFC).

News Section (N1-N2).

Quantitative liquid chromatographic determination of sanguinarine in cell culture medium and in rat urine and plasma by Henri Hoellinger; Micheline Re; Alain Deroussent; Ravindra Pratap Singh; Thierry Cresteil (195-200).
Sanguinarine is a quaternary benzo[c]phenanthridine alkaloid, extracted from the argemone oil, which produced severe human intoxications. To investigate the sanguinarine biotransformation, we develop a simple extraction process and a high performance liquid chromatographic separation coupled to a sensitive fluorometric detection of sanguinarine in cell culture medium, as well as in rat urine and plasma. After extraction with an acidified organic solvent, sanguinarine elution is performed within 15 min on a Nucleosil C18 column with a gradient using 0.2% formic acid/water/acetonitrile as mobile phase. Extracted and standard sanguinarine are characterized by mass spectrometry. The extraction recovery of sanguinarine is about 80% in cell culture medium and in rat urine, but lower in plasma. This convenient high performance liquid chromatography (HPLC) method allows to quantify sanguinarine over concentrations ranged 10–2000 ng ml−1. The limit of fluorometric detection is 0.5 ng. Under these conditions, the lower limit of quantification of sanguinarine is 50 ng ml−1 in cell culture medium and in rat urine and 100 ng ml−1 in rat plasma. This analytical HPLC method is specific, linear and reproducible in all media and is suitable for quantitative determination of sanguinarine in biological fluids.
Keywords: Sanguinarine;

Determination of plasma ziprasidone using liquid chromatography with fluorescence detection by Raymond F Suckow; Mira Fein; Christoph U Correll; Thomas B Cooper (201-208).
A liquid chromatographic procedure was developed for the determination of a new antipsychotic agent ziprasidone in plasma using fluorescence detection. A one-step liquid–liquid extraction from 1 ml of alkalinized plasma containing an internal standard α-ergocryptine using methyl-t-butyl ether afforded a greater than 84% recovery of ziprasidone. Chromatography was performed using a reversed-phase trimethylsilyl bonded silica column with a mobile phase of 72:28 phosphate buffer:acetonitrile at a flow rate of 1.5 ml/min. Detection of the eluted peaks was observed using excitation and emission wavelengths of 320 and 410 nm, respectively. Chromatographic run time did not exceed 14 min with no interference from endogenous material. The calibration curve was linear over the concentration range of 0.5 to 200 ng/ml and the inter- and intra-assay imprecision (CV) was less than 10%. The lower limit of quantitation was assessed at 0.5 ng/ml. Specificity of the method is demonstrated by the lack of interference from a large number of commonly used drugs and their metabolites in clinical use. The utility of the method is exemplified with the presentation of clinical data from patients receiving ziprasidone.
Keywords: Ziprasidone;

Determination of trivalent methylated arsenicals in rat urine by liquid chromatography–inductively coupled plasma mass spectrometry after solvent extraction by Masako Okina; Kaoru Yoshida; Koichi Kuroda; Hideki Wanibuchi; Shoji Fukushima; Ginji Endo (209-215).
A method for the determination of trivalent arsenicals in urine was examined. Trivalent arsenicals, extracted as complexes with diethylammonium diethyldithiocarbamate (DDDC) into carbon tetrachloride, were determined by liquid chromatography–inductively coupled plasma mass spectrometry (LC–ICP-MS). The trivalent methylated arsenicals monomethylarsonous acid (MMA(III)), dimethylarsinous acid (DMA(III)), and trimethylarsine (TMA) were detected in urine of rats that had received dimethylarsinic acid (DMA(V)) or monomethylarsonic acid (MMA(V)) at concentration of 200 μg ml−1 in drinking water for 24 weeks. This method is the first to permit quantification of trivalent methylated arsenicals in urine without significant changes in concentration during storage or pretreatment.
Keywords: Trivalent methylated arsenicals;

Determination of new derivatives of genistein in culture media by liquid chromatography by Hanna Ksycińska; Barbara Sobik; Joanna Popiołkiewicz; Krzysztof Polkowski; Piotr Krzeczyński; Jan Ramza; Wiesław Pucko; Grzegorz Grynkiewicz (217-231).
Methods for determination of genistein and its four new analogues in culture media have been developed to support studies on their potential anticancer activities. The investigated compounds were extracted from the media using liquid–liquid extraction with appropriate solvent. After evaporation of organic solvents each of the dry extracts was reconstituted in appropriate mobile phase. Reversed-phase HPLC was applied to quantitative determining of tested compounds. The methods are specific, sensitive and technically simple. They were used to evaluate concentration level of investigated compounds in experiments with human promyelocytic leukemia cells (HL-60 cell line).
Keywords: Genistein;

A simple and sensitive reversed-phase isocratic HPLC method for the determination of albendazole and its metabolites has been developed. The mobile phase consisting of acetonitrile–water–perchloric acid (70%) (30:110:0.06 (v/v/v)) was pumped at a flow rate of 0.80 ml/min on a 5 μm, reverse phase, Discovery® RPamide C16 column with UV detection at 290 nm. The calibration graphs were linear in the range of 0.05–1 μg/ml for albendazole, albendazole sulphoxide and albendazole sulphone. The limit of quantification was 50 ng/ml for albendazole, 25 ng/ml for albendazole sulphoxide and 30 ng/ml for albendazole sulphone. The within-day and day-to-day coefficient of variation averaged 4.98 and 6.95% for albendazole, 3.83 and 6.83% for albendazole sulphoxide and 3.44 and 5.51% for albendazole sulphone, respectively. The mean extraction recoveries of albendazole, albendazole sulphoxide and albendazole sulphone were 79.25, 93.03 and 88.78%, respectively. The method was applied to determine the plasma levels of albendazole sulphoxide in endemic normals administered with albendazole during pharmacokinetic studies.
Keywords: Albendazole sulphoxide;

Alcohol dehydrogenase (ADH) was extracted from a crude bovine liver homogenate by dye–ligand affinity counter-current chromatography (CCC) using a cross-axis coil planet centrifuge (x-axis CPC). The purification was performed using two types of polymer phase systems composed of 4.4% polyethylene glycol (PEG) 8000–7.0% dextran T500–0.1 M potassium phosphate buffers and 16% PEG 1000–12.5% potassium phosphate buffers, both containing a procion red dye as an affinity ligand at various pH values. The best purification was achieved using the PEG 1000–potassium phosphate system at pH 7.3 containing 0.05% procion red as a ligand. The upper PEG-rich phase containing procion red was used as the stationary phase and a crude bovine liver homogenate was eluted with the potassium phosphate-rich lower phase at 0.5 ml/min. After elution of bovine liver proteins in the homogenate, ADH still retained in the stationary phase was collected from the column by eluting with the PEG 1000-rich upper phase. Collected fractions were analyzed by ADH enzymatic activity and by sodium dodecyl sulfate (SDS)–polyacrylamide gel electrophoresis (PAGE) to detect contaminant proteins in the ADH fractions. The ADH was purified directly from crude bovine liver extract within 6 h with minimum loss of its enzymatic activity.
Keywords: Purification; Dye–ligand affinity counter-current chromatography; Alcohol dehydrogenase;

We developed an isotope dilution HPLC–atmospheric pressure chemical ionization–tandem mass spectrometry (HPLC–APCI–MS/MS) method for the simultaneous determination of p-tyrosine, phenylalanine, o,o′-dityrosine, m-tyrosine, o-tyrosine, 3-chlorotyrosine and 3-nitrotyrosine and 8-hydroxy-2′-deoxyguanosine (8-OHdG) that requires no extensive sample pre-treatment. p-[2H4]Tyrosine and o,o′-[2H6]dityrosine were used as internal standards. Calibration curves of the method were linear (r 2=0.990–0.999) over a concentration range of 0.03–10 μM for o-tyrosine; 0.04–10 μM for 3-nitrotyrosine and 3-chlorotyrosine; 0.05–10 μM for o,o′-dityrosine; and for m-tyrosine; 1.0–100 μM for p-tyrosine and for phenylalanine; and 0.01–10 μM for 8-OHdG. The detection limits were from 0.025 to 0.05 μM for the tyrosine derivatives; 0.01 μM for 8-OHdG; and 0.5 μM for p-tyrosine and for phenylalanine, respectively. Within-day coefficients of variation (CV) for spiked human urine samples ranged from 2.7 to 7.0%, except for 8-OHdG (13.7%). Between-day variations ranged from 7.9 to 13.0%, except for o-tyrosine (CV = 18.2%), and for 8-OHdG (CV = 24.7%).The background levels of p-tyrosine, phenylalanine, o,o′-dityrosine, and o-tyrosine in morning urine of eight healthy volunteers were 3890±590, 3420±730, 5.8±0.3, and 9.2±1.5 μmol/mol creatinine, respectively. Using the present HPLC–APCI–MS/MS method, the urinary background levels of m-tyrosine, 3-chlorotyrosine, 3-nitrotyrosine and 8-OHdG were below the limit of detection.
Keywords: Non-invasive biomarkers; Tyrosine; Phenylalanine; Deoxyguanosine;

Cellular membranes from a cell line expressing P-glycoprotein (Pgp(+)) and from a cell line that does not express Pgp (Pgp(−)) were immobilized on the surface of glass capillaries (25  cm×100  μm i.d.) by non-covalent interactions using the avidin–biotin coupling system to create two open tubular columns, Pgp(+)-OT and Pgp(−)-OT. Frontal displacement chromatography on the Pgp(+)-OT demonstrated that the immobilized Pgp retained its ability to specifically bind the known Pgp substrates vinblastin and ketoconazole. The calculated affinities, expressed as K d, for vinblastin and ketoconazole were 97 nM and 12.1 μM, which were comparable with previously reported K d values of 37 nM and 8.6 μM, respectively. The results confirm that the Pgp(+)-OT can be used to quantitatively estimate binding affinities for the Pgp. Frontal displacement chromatography on the Pgp(−)-OT demonstrated that the immobilized membranes retained the ability to bind some Pgp substrates, but that the binding was not due to specific binding to Pgp. A cohort of compounds containing high affinity Pgp substrates (vinblastin, prazosin) and moderate-low affinity Pgp substrates (doxorubicin, verapamil, ketoconazole) and a non-substrate (nicotine) were chromatographed on the Pgp(+)-OT and Pgp(−)-OT using fast frontal analysis and mass spectrometric detection. The results demonstrated that when the retention on the Pgp(+)-OT was corrected by subtraction of the retention on the Pgp(−)-OT, the test compounds could be accurately sorted into high, moderate-low and non-substrate categories. The data from the study indicates that a single 30-min parallel chromatographic experiment can be used to rank a compound based upon its relative affinity for the immobilized Pgp.
Keywords: Substrates; ABC transporters; Screening; P-glycoprotein;

A sensitive analytical method for gabapentin [1-(aminomethyl) cyclohexaneacetic acid] (GBP) in human plasma based on capillary electrophoretic separation and laser-induced fluorescence (LIF) detection has been developed. 6-Carboxyfluorescein succinimidyl ester (CFSE) was used for precolumn derivatization of the non-fluorescent drug in plasma. Optimal separation and detection were obtained with an electrophoretic buffer of 50 mM sodium borate (pH 9.5) and an air-cooled argon-ion laser (excitation at 488 nm, emission at 520 nm). A calibration curve ranging from 0.3 to 150 μM was shown to be linear. The concentration limit of detection (LOD) in plasma was 60 nM. We also demonstrate how the detection limit can be enhanced by using acetonitrile stacking technique. With stacking, the limit of detection for gabapentin in plasma was 4.8 nM. A calibration curve ranging from 0.03 to 15 μM was shown to be linear. Both the within-day and day-to-day reproducibility and accuracy were ≤10.8 and 6.0%, respectively.
Keywords: Acetonitrile stacking technique; Gabapentin;

A rapid, sensitive, specific, accurate, and reproducible automated liquid chromatography–tandem mass spectrometric (LC–MS/MS) method for the quantitative determination of 1′-(2-amino-3-methylbenzoyl)-4-[[[(3-chlorophenyl)sulfonyl]phenyl]methyl]-1,4′-bipiperidine hydrochloride (SCH 211803) in plasma has been developed. The method was validated in rat and monkey plasma over the concentration range of 0.5–250 ng/ml using 2 H 4 -SCH 211803 as the internal standard (IS). Automated 96-well plate protein precipitation (PP) with acetonitrile (ACN) was used for sample processing. The method employed a Betasil C18 column with a fast gradient for the separation of analyte and internal standard from the plasma matrix and a triple quadrupole mass spectrometer operated in positive ion multiple reaction monitoring (MRM) mode for detection. The method was used for the determination of SCH 211803 plasma concentrations to support pre-clinical studies.
Keywords: Protein precipitation; SCH 211803;

Procarbazine is a cytotoxic chemotherapeutic agent used in the treatment of lymphomas and brain tumors. Its pharmacokinetic behavior remains poorly understood even though more than 30 years have elapsed since the drug was approved for clinical use. To characterize the pharmacokinetics of procarbazine in brain cancer patients during a phase I trial, a method for determining the drug in human plasma by reversed-phase high-performance liquid chromatography (HPLC) with electrospray ionization mass spectrometry (ESI–MS) was developed and thoroughly validated. Plasma samples were prepared for analysis by precipitating proteins with trichloroacetic acid and washing the protein-free supernatant with methyl tert-butyl ether to remove excess acid. The solution was separated on a Luna C-18 analytical column using methanol-25 mM ammonium acetate buffer, pH 5.1 (22:78, v/v) as the mobile phase at 1.0 ml/min. A single-quadrupole mass spectrometer with an electrospray interface was operated in the selected-ion monitoring mode to detect the [M+H]+ ions at m/z 222.2 for procarbazine and at m/z 192.1 for the internal standard (3-dimethylamino-2-methylpropiophenone). Procarbazine and the internal standard eluted as sharp, symmetrical peaks with retention times (mean±S.D.) of 6.3±0.1 and 9.9±0.3 min, respectively. Calibration curves of procarbazine hydrochloride in human plasma at concentrations ranging from 0.5 to 50 ng/ml exhibited excellent linearity. The mean absolute recovery of the drug from plasma was 102.9±1.0%. Using a sample volume of 150 μl, procarbazine was determined at the 0.5 ng/ml (1.9 nM) lower limit of quantitation with a mean accuracy of 105.2% and an interday precision of 3.60% R.S.D. on 11 different days over 5 weeks. During this same time interval, the between-day accuracy for determining quality control solutions of the drug in plasma at concentrations of 2.0, 15 and 40 ng/ml ranged from 97.5 to 98.2% (mean±S.D., 97.9±0.4%) and the precision was 3.8–6.2% (mean±S.D., 5.1±1.2%). Stability characteristics of the drug were thoroughly evaluated to establish appropriate conditions to process, store and prepare clinical specimens for chromatographic analysis without inducing significant chemical degradation. The sensitivity achieved with this assay permitted the plasma concentration–time profile of the parent drug to be accurately defined following oral administration of standard doses to brain cancer patients.
Keywords: Procarbazine;

In order to develop possible correlations to predict partioning behaviour of proteins, five mammalian albumins (goat, bovine, equine, human and pig ones) with similar physico-chemical properties (molecular mass and isoelectrical point) were chosen. Evaluation of the relationship between hydrophobicity and partitioning coefficient (Kr) in polyethylenglycol–dextran (PEG–DxT500) systems formed by polyethyleneglycols of different molecular mass (3350, 6000 and 10,000) was investigated by estimating relative surface hydrophobicity (So) with a fluorescent probe, 1 anilino-8-naphthalene sulfonate. No relationship between Kr and So was found for systems formed by PEG3350, while aqueous two-phase systems with PEG6000 and PEG10,000 gave better correlations. The results obtained may be explained on the basis of an increase in the interaction between the latter PEGs and the protein due to their higher hydrophobic character which increases as the PEG molecular mass does so. In this way, systems with PEGs of higher molecular mass give the highest resolution to exploit hydrophobicity in partitioning.
Keywords: Protein surface hydrophobicity; Partitioning; Aqueous two-phase systems; Polyethyleneglycol–dextran;

Analytical derivatization–a tool for determination of orotic acid by Renata Hušková; Petr Barták; Lubomı́r Čáp; David Friedecký; Tomáš Adam (303-309).
Derivatization of orotic acid (OA) into various forms (trimethylsilylderivate, alkyl ester and per-methylated derivate) and their evaluation by GC/MS is described. The tested approach includes ion-exchange SPE clean-up, evaporation and chemical reaction with different types of derivatization agents (N,O-bis-(trimethylsilyl)trifluoroacetamide with trimethylchlorosilane, butanol with acetylchloride and ethereal solution of diazomethane). Derivate originated in the reaction with diazomethane was used for determination of urinary orotic acid by GC/MS. Detection limit of 0.28 μmol l−1 was reached using the ion 82  m/z in single ion monitoring (SIM) mode. Linearity of the method was tested within the range of 3.4–2503.4 μmol l−1 covering physiological and pathological levels of orotic acid in urine sample. Recoveries were within the range 93.7–110.6%. Application of the method on the patient with defect of ornithine transcarbamylase (OTC) was demonstrated as well.
Keywords: Derivatization, GC; Orotic acid;

A reversed-phase high-performance liquid chromatographic (HPLC) method with ultraviolet (UV) detection was developed and validated for the quantification of 6-deoxy-6-demethyl-4-dedimethylamino-tetracycline (COL-3), a matrix metalloproteinase (MMPs) inhibitor, in rat serum. This simple, sensitive, rapid and reproducible assay involved a preliminary serum deproteinization by adding a mixture of acetonitrile–methanol–0.5 M oxalic acid (70:20:10 (v/v)), as the combined precipitant and metal blocking agent, into serum samples (2:1 (v/v)). An aliquot (20 μl) of the supernatant was injected into the HPLC system linked to a Waters XTerra™ RP18 column (150  mm×4.6  mm i.d., particle size 5 μm). The compound was eluted by a mixture of acetonitrile–methanol–0.01 M oxalic acid (40:10:50 (v/v), pH 2.00), as the mobile phase, and detected at the wavelength of 350 nm. The total running time was 10 min. The low and high concentration calibration curves were linear in the range of 50–1200 ng/ml and 1200–12,000 ng/ml, respectively. The intra- and inter-day coefficients of variation at three quality control concentrations of 100, 1200, and 12,000 ng/ml were all less than 6%, while the percent error ranged from −2.5 to 6.6%. The limit of quantitation (LOQ) for COL-3 in serum was 50 ng/ml. This assay was successfully employed to study the serum concentration–time profiles of COL-3 after its intravenous and oral administration in rats. The method with some minor modifications in sample pretreatment was also applicable to the determination of the concentrations of COL-3 in rat bile, urine and feces.
Keywords: COL-3; Metalloproteinase inhibitor;

The double coating principle of CEofix® buffers was evaluated for the analysis of some basic drugs by capillary electrophoresis–diode-array detection (CE–DAD) and capillary electrophoresis–mass spectrometry (CE–MS). The involatile phosphate present in original low pH CEofix®, was replaced with formic acid for hyphenation of CE with MS. The double coating produces a substantial and highly reproducible electroosmotic flow (EOF), even at low pH. The rinsing procedure and electrolyte composition were optimized for both CE–DAD and CE–MS. The system was evaluated with the analysis of a mixture of basic drugs and a spiked urine sample enriched by solid-phase extraction (SPE). The R.S.D. values on the migration time and peak area measured for 28 analyses with CE–DAD were below 0.25 and 2.40%, respectively. For CE–MS, the R.S.D. on the migration time was 0.85% or less and the area precision ranged from 5.65 to 14.33% (for seven injections). The LOD with the developed CE–MS method was below 50 ppb for all five drug standards tested.
Keywords: Dynamic coating; Basic drugs;

Rapid and sensitive static headspace gas chromatography–mass spectrometry method for the analysis of ethanol and abused inhalants in blood by Ibrahim A Wasfi; Ahmed Hassan Al-Awadhi; Zainat Naser Al-Hatali; Fatima Juma Al-Rayami; Nawal Abdulla Al Katheeri (331-336).
A sensitive and specific method using static headspace gas chromatography coupled with mass spectrometry (GC/MS) has been developed for the quantitative determination of ethanol in biological fluids using n-propanol as internal standard. Gas chromatography was performed in isothermal mode with a GC run time of 2.6 min. The quantification was performed using scan mode abstracting a quantitative ion and a qualifier ion for ethanol and for the internal standard. The method was linear (r 2, 0.999, in the concentration range of 5–200 mg/dl), specific (no interference from methanol acetaldehyde, acetone or from endogenous materials), sensitive (limit of quantification and limit of detection of 0.2 and 0.02 mg/dl, respectively) and robust (less than 5% inter- and intra-assay coefficient of variation). A slightly modified method was also developed for the quantification of five commonly abused inhalants (dichloromethane, ethyl acetate, benzene, toluene and xylene) in blood. The method used a gradient GC program with a run time of 8 min. The quantification was performed using scan mode and integrating the area under the peak using trichloroethane as an internal standard. Without optimization, the method was linear (from 5 to 100 mg/l) and sensitive.
Keywords: Ethanol; Dichloromethane; Ethyl acetate; Benzene; Toluene; Xylene;

Validation of a simplified method for determination of cimetidine in human plasma and urine by liquid chromatography with ultraviolet detection by Tahira Iqbal; Chetan S Karyekar; Minori Kinjo; Gilbert C Ngan; Thomas C Dowling (337-341).
A HPLC method was developed for determination of cimetidine in human plasma and urine. Plasma samples were alkalinized followed by liquid extraction with water-saturated ethyl acetate then evaporated under nitrogen. The extracts were reconstituted in mobile phase and injected onto a C18 reversed-phase column; UV detection was set at 228 nm. Urine samples were diluted with an internal standard/mobile phase mixture (1:9) prior to injection. The lower limit of quantification in plasma and urine were 100 ng/ml and 10 μg/ml, respectively; intra- and inter-day coefficients of variation were ≤4.2%. Advantages of this validated assay include a readily available internal standard, simplified plasma extraction and urine dilution methods, and applicability to clinical studies investigating the renal handling of cimetidine.
Keywords: Cimetidine;

Different chromatographic methods including chromatofocusing are used for separation of manganese peroxidase (MnP) isoforms and their isolation from the fungal growth medium. We tested strong anion exchange methacrylate based monolithic columns as a stationary phase for fast separation of MnP’s. Sodium acetate buffers of two different pH values (6 and 4) were used for formation of reproducible pH gradient. The entire cycle, involving analysis and column regeneration, was completed in 3 min. Use of pH gradient showed better MnP isoform separation comparing to the salt gradient, while application of combined pH–salt gradient, resulted in further improvement.
Keywords: pH gradient; Salt gradient; Manganese peroxidase isoenzymes;

We investigated high-performance liquid chromatographic (HPLC) determination of 1-adamantanamine hydrochloride (1-ADA) and 2-adamantanamine hydrochloride (2-ADA) in human plasma after the derivatization with o-phthalaldehyde (OPA) and 1-thio-β-d-glucose (TG). Extracted human plasma samples were mixed with OPA and TG at room temperature for 6 min and injected onto HPLC. Retention times of 1-ADA and 2-ADA derivatives were 12.6 and 14.1 min, respectively. The lower limits of detection of 1-ADA and 2-ADA were 0.02 and 0.008 μg/ml, and the lower limits of quantitation of 1-ADA and 2-ADA were 0.025 and 0.01 μg/ml, respectively. The coefficients of variation for intra-day and inter-day assay of 1-ADA and 2-ADA were less than 4.4 and 6.0%, respectively. l-Dopa and dopamine were not found to interfere with the peaks of 1-ADA and 2-ADA derivatives. Human plasma unbound fraction (f p) values of 1-ADA varied between 0.32 and 0.48, while those of 2-ADA varied between 0.38 and 0.68. These results indicate that HPLC assay of 1-ADA and 2-ADA by derivatization with OPA and TG is simple, rapid, sensitive and reproducible for determining 1-ADA and 2-ADA in human plasma.
Keywords: Derivatization; LC; Adamantanamine;

A simple, sensitive and reproducible HPLC method is presented for the simultaneous determination of mycophenolic acid (MPA) and its metabolites phenolic MPA-glucuronide (MPAG) and acyl glucuronide (AcMPAG) in human plasma. Sample purification requires protein precipitation with 0.1 M phosphoric acid/acetonitrile in the presence of Epilan D as an internal standard (IS). Separation was performed by reversed-phase HPLC, using a Zorbax SB-C18 column, 32% acetonitrile and a 40 mM phosphoric acid buffer at pH 3.0 as mobile phase; column temperature was 50 °C, flow rate 1.4 ml/min, and measurement by UV detection was at 215 nm (run time 12 min). The method requires only 50 μl plasma. Detection limits were 0.1 μg/ml for MPA and AcMPAG, and 2.0 μg/ml for MPAG, respectively. Mean absolute recovery of all three analytes was >95%. This analytical method for the determination of MPA and its metabolites is a reliable and convenient procedure that meets the criteria for application in routine clinical drug monitoring and pharmacokinetic studies.
Keywords: Mycophenolic acid;