Quantitative determination of tilmicosin in canine serum by high performance liquid chromatography-tandem mass spectrometry

A highly sensitive and quantitative LC/MS/MS assay for the determination of tilmicosin in serum has been developed and validated. For sample preparation, 0.2 mL of canine serum was extracted with 3 mL of methyl tert-butyl ether. The organic layer was transferred to a new vessel and dried under nitrogen. The sample was then reconstituted for analysis by high performance liquid chromatography-tandem mass spectrometry. A Phenomenex Luna C8(2) analytical column was used for the chromatographic separation. The eluent was subsequently introduced to the mass spectrometer by electrospray ionization. A single range was validated for 50-5000 ng/mL for support of toxicokinetic studies. The inter-day relative error (inaccuracy) for the LLOQ samples ranged from -5.5% to 0.3%. The inter-day relative standard deviations (imprecision) at the respective LLOQ levels were < or =10.1%.     

Simultaneous determination of 1,5-dicaffeoylquinic acid and its active metabolites in human plasma by liquid chromatography-tandem mass spectrometry for pharmacokinetic studies

1,5-Dicaffeoylquinic acid (1,5-DCQA), a potent HIV-1 integrase inhibitor, is undergoing an evaluation as a promising novel HIV therapeutic agent. Here, we report a simple, rapid and robust LC-MS/MS method for simultaneous determination of 1,5-DCQA and its two active metabolites, 1-caffeoyl-5-feruoylquinic acid (1,5-CFQA) and 1,5-O-diferuoylquinic acid (1,5-DFQA) in human plasma. The quantitation of the target compounds was determined by selected reaction monitoring (SRM) mode using electrospray ionization (ESI). Good linearity was obtained in the 3-500 ng/ml range for each analyte and the analytical method was validated in terms of specificity, precision, accuracy, recovery, stability and matrix effect. These assays gave R.S.D.% values for precision always lower than 13.8% and R.E.% values for accuracy between -8.9 and 0.9%. In addition, the specificity, extraction recovery, stability and matrix effect were satisfactory too. Using the measured plasma concentrations of 1,5-DCQA and its active metabolites in five healthy volunteers, pharmacokinetic profiles of 1,5-DCQA and its active metabolites were evaluated, which supported the clinical pharmacokinetic studies successfully. Due to its high sensitivity, specificity and simplicity, the method could be used for pharmacokinetic studies of both 1,5-DCQA and its active metabolite, and for routine monitoring of their levels in human plasma.     

Determination and pharmacokinetic study of chiisanogenin in rat plasma by ultra performance liquid chromatography-tandem mass spectrometry

Introduction – Chiisanogenin existing in many Acanthopanax species has been reported to possess anti‐inflammatory, antibacterial and antiplatelet aggregatory activities. Objective – To develop and validate a rapid and sensitive ultra performance liquid chromatography‐tandem mass spectrometry method for the determination of chiisanogenin in rat plasma and to investigate its pharmacokinetics after oral administration of chiisanogenin or the extract of Acanthopanax sessiliflorus fruits. Methodology – The sample pretreatment involved a one‐step extraction of 0.2 mL plasma with diethyl ether. Acetaminophen was used as the internal standard. The separation was carried out on an ACQUITY UPLC? BEH C₁₈ column with a mobile phase of acetonitrile‐5 mM ammonium acetate (90:10, v/v) at a flow rate of 0.2 mL/min. The detection was performed on a triple quadrupole tandem mass spectrometer by multiple reaction monitoring (MRM) mode via electrospray ionization (ESI) source. Results – A high sample throughput was achieved with an analysis time of 1.1 min per sample. The calibration curve was linear (r² ≥ 0.99) over the concentration range of 5–500 ng/mL with a lower limit of quantification (LLOQ) of 5 ng/mL. The intra‐day and inter‐day precision (relative standard deviation, R.S.D.) values were below 11% and the accuracy (relative error, R.E.) was within 8% at all three quality control (QC) levels. Conclusion – The method was successfully applied to the pharmacokinetic study of chiisanogenin in rat after oral administration of chiisanogenin and the extract of Acanthopanax sessiliflorus fruits. Other constituents in the extract affected the pharmacokinetic behavior of chiisanogenin. Copyright © 2010 John Wiley & Sons, Ltd.     

Simultaneous determination of morphine, oxycodone, morphine-3-glucuronide, and noroxycodone concentrations in rat serum by high performance liquid chromatography-electrospray ionization-tandem mass spectrometry

An assay using high performance liquid chromatography (HPLC)-electrospray ionization-tandem mass spectrometry (ESI-MS-MS) was developed for simultaneously determining concentrations of morphine, oxycodone, morphine-3-glucuronide, and noroxycodone, in 50 microl samples of rat serum. Deuterated (d(3)) analogues of each compound were used as internal standards. Samples were treated with acetonitrile to precipitate plasma proteins; acetonitrile was removed from the supernatant by centrifugal evaporation before analysis. Limits of quantitation (ng/ml) and their between-day accuracy and precision (%deviation and %CV) were--morphine, 3.8 (4.3% and 7.6%); morphine-3-glucuronide, 5.0 (4.5% and 2.9%); oxycodone, 4.5 (0.4% and 9.3%); noroxycodone, 5.0 (8.5% and 4.6%).     

Simultaneous determination of gemcitabine and gemcitabine-squalene by liquid chromatography-tandem mass spectrometry in human plasma

Gemcitabine-squalene is a new prodrug that self-organizes in water forming nanoassemblies. It exhibits better anti-cancer properties in vitro and in vivo than gemcitabine. A liquid chromatography/tandem mass spectrometry assay of gemcitabine-squalene and gemcitabine was developed in human plasma in order to quantitate gemcitabine and its squalene conjugate. After protein precipitation with acetonitrile/methanol (90/10, v/v), the compounds were analyzed by reversed-phase high performance liquid chromatography and detected by tandem mass spectrometry using multiple reaction monitoring. The method was linear over the concentration range of 10-10,000 ng/ml of human plasma for both compounds with an accuracy lower than 10.4% and a precision below 14.8%. The method showed a lower limit of quantitation of 10 ng/ml of human plasma for dFdC and dFdC-SQ. A preliminary in vivo study in mice was shown as application of the method as no significant difference between human and mice plasma for the analysis of dFdC and dFdC-SQ was demonstrated.     

Simultaneous determination of methylephedrine and noscapine in human plasma by liquid chromatography-tandem mass spectrometry

A selective and sensitive method has been developed and validated for simultaneous quantification of methylephedrine and noscapine in human plasma. Analytes were extracted from human plasma samples by liquid-liquid extraction, separated on a Diamonsil C18 column and detected by tandem mass spectrometer with an atmospheric pressure chemical ionization (APCI) interface. Diphenhydramine was used as the internal standard (I.S.). The method was found to be precise and accurate within the linear range 0.1-100 ng/ml for each analyte. The intra- and inter-day relative standard deviations (R.S.D.s) were below 5.2% for methylephedrine and 6.7% for noscapine. The inter-day relative error (RE) as determined from quality control samples (QCs) was less than 3.0% for each analyte. The assay was successfully employed in a pharmacokinetic study after an oral administration of a multicomponent formulation containing 20 mg DL-methylephedrine hydrochloride, 16 mg noscapine, 300 mg paracetamol and 1mg of chlorpheniramine maleate.     

Simultaneous determination of enalapril and enalaprilat in human plasma by liquid chromatography-tandem mass spectrometry

A rapid, sensitive, and highly selective liquid chromatography-tandem mass spectrometry method was developed and validated for simultaneous determination of enalapril and its major active metabolite enalaprilat in human plasma. The analytes were extracted from plasma samples by liquid-liquid extraction, separated on a Zorbax Extend-C(18) column, and detected by tandem mass spectrometry with a Turbo IonSpray ionization interface. The method has a lower limit of quantification (LLOQ) of 0.1 ng/ml for both enalapril and enalaprilat. The chromatographic run time was approximately 3.5 min. The standard calibration curves for both enalapril and enalaprilat were linear in the concentration ranges of 0.10-100.0 ng/ml in human plasma. The intra- and inter-run precisions, expressed as the relative standard deviation (R.S.D.), were less than 7.7 and 7.8%, determined from QC samples for enalapril and enalaprilat, and accuracy was within +/-3.9 and +/-2.7% in terms of relative error, respectively. The method was successfully applied for the evaluation of the pharmacokinetics of enalapril and enalaprilat in 20 volunteers after an oral dose of 10 mg enalapril maleate.     

Simultaneous determination of four immunosuppressants by means of high speed and robust on-line solid phase extraction-high performance liquid chromatography-tandem mass spectrometry

In this study immunosuppressants, i.e. cyclosporin A (CyA), tacrolimus (TRL), sirolimus (SRL) and everolimus (RAD) were quantified in whole blood samples from immunosuppressant treated transplant recipients by an integrated on-line solid phase extraction-high performance liquid chromatography-tandem mass spectrometry (SPE-HPLC-MS/MS) system. This method has been developed to improve the following characteristics: speed, robust analysis, simultaneous determination and low cost. This can be achieved by the use of a perfusion column as an extraction cartridge in combination with a short HPLC column and highly selective and sensitive atmospheric pressure ionisation tandem mass spectrometry (API-MS/MS) in the multiple reaction monitoring (MRM) detection mode. This high throughput technique is perfectly appropriate for routine therapeutic drug monitoring (TDM) of organ transplanted patients.     

Simultaneous determination of hydrochlorothiazide, quinapril and quinaprilat in human plasma by liquid chromatography-tandem mass spectrometry

A rapid and sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) method has been developed and validated for the simultaneous estimation of hydrochlorothiazide, quinapril and its metabolite quinaprilat in human plasma. After solid phase extraction (SPE), the analytes and IS were chromatographed on a hypurity C8 (100mmx2.1mm i.d., 5mum particle size) column using 2muL injection volume with a run time of 2.8min. An isocratic mobile phase consisting of 0.5% (v/v) formic acid:acetonitrile (25:75, v/v) was used to separate all these drugs. The precursor and product ions of these drugs were monitored on a triple quadrupole mass spectrometer, operating in the multiple reaction monitoring mode (MRM) without polarity switch. The proposed method was validated over the range of 5-500ng/mL for hydrochlorothiazide method and 5-1500ng/mL for quinapril and quinaprilat. Inter-batch and intra-batch precision (coefficient of variation - % CV) across five validation runs lower limit of quantitation (LLOQ), lower quality control (LQC), middle quality control (MQC), higher quality control (HQC) and upper limit of quantitation (ULOQ) was less than 15. The accuracy determined at these levels was within +/-13% in terms of relative percentage error.     

Simultaneous determination of doxifluridine and 5-fluorouracil in monkey serum by high performance liquid chromatography with tandem mass spectrometry

A reverse-phase high performance liquid chromatography method with electrospray ionization and detection by mass spectrometry is described for the simultaneous determination of doxifluridine and its active metabolite 5-fluorouracil in monkey serum. A liquid/liquid extraction with ethyl acetate (90%) and isopropyl alcohol (10%) was used to extract simultaneously doxifluridine and 5-FU which have considerable difference in the polarity. Optimum chromatographic separation was achieved on a Agilent Zorbax C(18) (100 mm x 2.1mm, 3.5 microm) column with a mobile phase of methanol-water (20:80, v/v). The flow rate was 0.2 mL/min with total cycle time of 5 min. The lower limit of quantification (LLOQ) was validated at 10.0 ng/mL of serum for both doxifluridine and 5-FU. Accuracy and precision of quality control (QC) samples for both compounds met FDA Guidance criteria of +/-15% with average QC accuracy of 95.5-105.0% and coefficients of variation of 1.1-9.5% in the 10-2000 ng/mL concentration range. This method demonstrated adequate sensitivity, specificity, accuracy, precision, stability to support the analysis of monkey serum samples.     

Simultaneous determination of thienorphine and its active metabolite thienorphine glucuronide in rat plasma by liquid chromatography-tandem mass spectrometry and its application to pharmacokinetic studies

A simple, sensitive and reliable method was developed to determine simultaneously the concentrations of thienorphine and its metabolite thienorphine glucuronide conjugate in rat plasma by liquid chromatography-tandem mass spectrometry (LC-MS/MS). The metabolite was identified by MS: thienorphine glucuronide conjugate. Sample preparation involved protein precipitation with methanol. Analytes were separated on Finnigan BetaBasic-18 column (150 mm x 2.1mm i.d., 5 microm) using methanol: water: formic acid (56:44:0.1, v/v/v) as mobile phase at a flow rate of 0.2 ml/min. The method had a linear calibration curve over the concentration range of 0.1-50 ng/ml for thienorphine and 2-1000 ng/ml for thienorphine glucuronide conjugate, respectively. LOQ of thienorphine and thienorphine glucuronide conjugate was 0.1 and 2 ng/ml, respectively. The intra- and inter-batch precisions were less than 12% and their recoveries were greater than 80%. Pharmacokinetic data of thienorphine and its metabolite thienorphine glucuronide conjugate obtained with this method following a single oral dose of 3mg/kg thienorphine to rats were also reported for the first time.     

Enantioselective determination of lercanidipine in human plasma for pharmacokinetic studies by normal-phase liquid chromatography-tandem mass spectrometry

We describe here the first method for the enantioselective analysis of the calcium antagonist lercanidipine in human plasma by high performance liquid chromatography (HPLC) employing tandem mass spectrometric (MS) detection. Routine determination of lercanidipine enantiomers in human plasma in the working range of 0.025-50.0 ng ml(-1) plasma for each enantiomer with an accuracy and precision less than 15% was possible. Application of the method to a stereospecific study of the pharmacokinetics showed that plasma levels after an oral dose of rac-lercanidipine administered to a healthy volunteer were found to be higher for the (S)-enantiomer.     

Simultaneous determination of six urinary porphyrins using liquid chromatography-tandem mass spectrometry

A liquid chromatographic-tandem mass spectrometric (LC-MS-MS) method without sample pretreatment was developed and validated for determination of porphyrins in samples of canine urine. Acidified urine samples were directly injected into the LC-MS system and a gradient elution program was applied. The mass spectrometer was operated in the multi-reaction monitoring (MRM) mode and six porphyrins were detected with excellent sensitivity and selectivity. The lower limits of quantification were 0.014 nmol/mL for mesoporphyrin IX, coproporphyrin I, 5-carboxylporphyrin, 6-carboxylporphyrin and 7-carboxylporphyrin, and 0.029 nmol/mL for uroporphyrin I. Good ln-quadratic responses of calibration standards over the range 0.01 to 1.0 nmol/mL for mesoporphyrin IX, coproporphyrin I, 5-carboxylporphyrin, 6-carboxylporphyrin and 7-carboxylporphyrin, and 0.02 to 1.0 nmol/mL for uroporphyrin I were demonstrated. This method should be easily adapted through cross-validation for use in determining the effects of chemicals and pharmaceuticals on the urinary excretion profile of porphyrins in preclinical studies with other species, and in assisting the diagnosis of porphyria in clinical studies.     

Simultaneous quantification of nicotine and metabolites in rat brain by liquid chromatography-tandem mass spectrometry

A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for simultaneous quantification of nicotine (NIC), cotinine (COT), nornicotine (NNIC), norcotinine (NCOT), nicotine-N-β-D-glucuronide (NIC GLUC), cotinine-N-β-D-glucuronide (COT GLUC), nicotine-1'-oxide (NNO), cotinine-N-oxide (CNO), trans-3'-hydroxycotinine (3-HC), anabasine (AB) and anatabine (AT) was modified and validated for quantification of these selected analytes in rat brain tissue. This analytical method provides support for preclinical NIC pharmacokinetic and toxicological studies after controlled dosing protocols. After brain homogenization and solid-phase extraction, target analytes and corresponding deuterated internal standards were chromatographically separated on a Discovery(?) HS F5 HPLC column with gradient elution and analyzed by LC-MS/MS in positive electrospray ionization (ESI) mode with multiple reaction monitoring (MRM) data acquisition. Method linearity was assessed and calibration curves were determined over the following ranges: 0.1-7.5 ng/mg for NIC, COT GLUC and AB; and 0.025-7.5 ng/mg for COT, NNIC, NCOT, NIC GLUC, NNO, CNO, 3-HC and AT (R(2)≥0.99 for all analytes). Extraction recoveries ranged from 64% to 115%, LC-MS/MS matrix effects were ≤21%, and overall process efficiency ranged from 57% to 93% at low and high quality control concentrations. Intra- and inter-assay imprecisions and accuracy for all analytes were ≤12.9% and ≥86%, respectively. The method was successfully applied to quantification of NIC and metabolites in the brain of post-natal day 90 rats that were sacrificed 2-h after a single 0.8 mg/kg s.c. administration of (-)NIC. In these tissues, striatal concentrations were 204.8±49.4, 138.2±14.2 and 36.1±6.1 pg/mg of NIC, COT and NNIC, respectively. Concentrations of NIC, COT and NNIC in the remaining whole brain (RWhB) were 183.3±68.0, 130.0±14.1 and 46.7±10.3 pg/mg, respectively. Quantification of these same analytes in plasma was also performed by a previously validated method. NIC, COT, NNIC, NCOT, NNO and CNO were detected in plasma with concentrations comparable to those reported in previous studies. However, and in contrast to brain tissues, COT concentrations in plasma were significantly higher than were those of NIC (194.6±18.6 ng/mL versus 52.7±12.9 ng/mL). Taken together, these results demonstrate that a sensitive and selective method has been developed for the determination of NIC biomarkers in rat brain.     

Determination of p-aminohippuric acid in rat plasma by liquid chromatography-tandem mass spectrometry

A rapid, simple and sensitive method was developed for the determination of para-aminohippuric acid (PAH) in rat plasma using liquid chromatography tandem mass spectrometry (LC-MS-MS). Acetaminophen was used as the internal standard. Chromatographic separation was performed using a Symmetry C₁₈ column and the mobile phase was composed of A: 2 mM ammonium formate and 0.1% formic acid in water and B: 2 mM ammonium formate and 0.1% formic acid in acetonitrile (ACN) (A:B, 30:70, v/v). Detection was performed on a triple–quadrupole tandem mass spectrometer using positive ion mode electrospray ionization (ESI) in the multiple reaction monitoring (MRM) mode. The MS/MS ion transitions monitored were m/z 195.2 → 120.2 and 152.1 → 110.1 for PAH and acetaminophen, respectively. Good linearity is observed over the concentration range of 0.1–500 μg/ml. The method was proved to be accurate and reliable and was applied to a pharmacokinetic study in rat.     

Simultaneous determination of triazolam and its metabolites in human plasma by liquid chromatography-tandem mass spectrometry

A sensitive and selective liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the determination of triazolam and its metabolites, alpha-hydroxytriazolam (alpha-OHTRZ) and 4-hydroxytriazolam (4-OHTRZ), was developed and validated. Triazolam-D4 was used as the internal standard (IS). This analysis was carried out on a Thermo((R)) C(18) column and the mobile phase was composed of acetonitrile:H(2)O:formic acid (35:65:0.2, v/v/v). Detection was performed on a triple-quadrupole tandem mass spectrometer using positive ion mode electrospray ionization (ESI) and quantification was performed by multiple reaction monitoring (MRM) mode. The MS/MS ion transitions monitored were m/z 343.1-->308.3, 359.0-->308.3, 359.0-->111.2 and 347.0-->312.0 for triazolam, alpha-OHTRZ, 4-OHTRZ and triazolam-D4, respectively. LLOQ of the analytical method was 0.05ng/mL for triazolam and 0.1ng/mL for alpha-OHTRZ and 4-OHTRZ. The within- and between-run precisions were less than 15.26% and accuracy was -8.08% to 13.33%. The method proved to be accurate and specific, and was applied to the pharmacokinetic study of triazolam in healthy Chinese volunteers.     

Determination of mycophenolic acid glucuronide in microsomal incubations using high performance liquid chromatography-tandem mass spectrometry

A sensitive and specific HPLC-MS/MS method was developed for the analysis of mycophenolic acid glucuronide (MPAG) in incubations with human liver microsomes. Incubation samples were processed by protein precipitation with acetonitrile. MPAG and the internal standard phenolphthalein glucuronide were chromatographed on a C18 Synergi Fusion-RP column (100 mm x 2 mm, 4 microm) using gradient elution with a mixture of 1mM acetic acid in deionized water and 1mM acetic acid in acetonitrile at a flow rate of 0.22 mL/min. The mass spectrometer was operated with negative electrospray ionization and analysis was carried out in the single reaction monitoring (SRM) mode using the mass transitions of m/z 495-->319 and m/z 493-->175 for MPAG and phenolphthalein glucuronide, respectively. The MPAG calibration curve was linear over the concentration range of 1.0-20 microM. The within-day and between-day relative standard deviations ranged from 1.1 to 7.9% and accuracy was within 8%. The simple and reproducible method is suitable for measuring mycophenolic acid glucuronidation in microsomal incubations.     

Development and validation of a high performance liquid chromatography-tandem mass spectrometry for the determination of etodolac in human plasma

A simple and specific method using a one-step liquid-liquid extraction (LLE) with butyl acetate followed by high performance liquid chromatography (HPLC) coupled with positive ion electrospray ionization tandem mass spectrometry (ESI-MS/MS) detection was developed for the determination of etodolac in human plasma, using indomethacin as an internal standard (IS). Chromatographic separation was performed isocratically using a Capcellpak MGII C(18) column with 65% acetonitrile and 35% water containing 10mM ammonium formate (adjusted to pH 3.5 with formic acid). Acquisition was performed in multiple reaction monitoring (MRM) mode by monitoring the transitions: m/z 287.99>172.23 for etodolac and m/z 357.92>139.01 for IS. The method was validated to determine its selectivity, linearity, sensitivity, precision, accuracy, recovery and stability. The limit of quantitation (LLOQ) was 0.1microg/mL with a relative standard deviation of less than 15%. The devised method provides an accurate, precise and sensitive tool for determining etodolac levels in plasma.     

Simultaneous determination of sphingosine and sphingosine 1-phosphate in biological samples by liquid chromatography-tandem mass spectrometry

D-erythro-sphingosine (Sph) and its phosphorylated product, d-erythro-sphingosine 1-phosphate (S1P) are sphingolipids mediating numerous cellular processes. Imbalance of Sph/S1P levels contributes to many diseases. Given the interconversion of these two opposing signaling molecules, it is essential to examine their levels simultaneously. In the present study, we developed a rapid and sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) method to simultaneously quantify the levels of Sph and S1P in biological samples using C17-Sph and C17-S1P as internal standards. With one step of methanol-induced protein precipitation, each sample was subjected to LC-MS/MS analysis using positive electrospray ionization under selected reaction monitoring mode. The running time was within 4 min with a simple mobile phase consisting of methanol-0.1% formic acid (95:5, v/v) at a flow rate of 0.2 mL/min. Standard curves were linear over ranges of 1-100 ng/mL for Sph and 0.1-10 ng/mL for S1P with correlation coefficient (r2) greater than 0.997. The lower limit of quantifications (LLOQs) were 1 ng/mL for Sph and 0.1 ng/mL for S1P. The intra-batch and inter-batch precision was less than 15% for all quality control samples. The recoveries of the method were found to be 76.36-89.84%. The method was applied to simultaneously determine the Sph and S1P levels in mouse kidney, human plasma, and HEK 293 cells treated with tumor necrosis factor-α (TNF-α) and N,N-dimethylsphingosine (DMS). The S1P levels increased in cells treated with TNF-α whereas decreased in cells treated with DMS. These results indicated that this new LC-MS/MS method was rapid, sensitive, specific and reliable to quantify Sph and S1P levels in biological samples simultaneously.     

Determination of aldosterone in serum by liquid chromatography-tandem mass spectrometry

Measurement of serum aldosterone is clinically important in the diagnosis of hypertension. While isotope dilution gas chromatography-mass spectrometry (ID-GC-MS) provides reliable results, it requires derivatization and is lengthy and time-consuming. Detection by liquid chromatography-mass spectrometry (LC-MS) is a potentially superior method. The analysis utilizes 0.5mL of serum. The samples were extracted with dichloromethane-ether. The extract was evaporated to dryness and aldosterone was analyzed by LC-MS/MS operating in the negative mode ESI after separation on a reversed-phase column. Aldosterone was also measured by RIA. The calibration curves for analysis of serum aldosterone exhibited consistent linearity and reproducibility in the range of 60-3000pmol/L. Interassay CVs were 4.3-7.5% at aldosterone concentrations of 97-993pmol/L. The lower limit of quantitation (LOQ) was 30pmol/L (signal to noise ratio=10). The mean recovery of the analyte added to serum ranged from 95 to 102%. The regression equation by LC-MS/MS (x) and RIA (y) method was: y=1.33x+185 (r=0.95; n=124). Sensitivity and specificity of the LC-MS/MS method for serum aldosterone offer advantages over GC-MS by eliminating derivatization. The novel method is rapid, reliable and simple to perform with a routine LC-MS/MS spectrometer. The sensitivity is adequate for patient samples. Aldosterone concentrations reported by nonextraction RIA were consistently higher than those produced by LC-MS/MS.