Quantitation of DNA adduct of thymidylyl(3'-5')thymidine methyl phosphotriester by liquid chromatography/negative electrospray tandem mass spectrometry

A rapid and selective method based on liquid chromatography/electrospray tandem mass spectrometry (LC/ESI-MS/MS) has been developed for the direct quantitation of a methyl phosphotriester DNA adduct, thymidyl (3'-5') thymidine [dTp(Me)dT] from enzymatic hydrolysates of DNA (either in vitro DNA or in cell culture) treated with MNU (N-methyl-N-nitrosourea) or MMS (methyl methane sulfonate). The lower limit of quantitation was 2 ng/mL. Linearity of the calibration curve was greater than 0.999 from 2 to 1000 ng/mL. Intraday precision for four levels of quality controls ranged from 2.8 to 20.1%, and interday precision ranged from 2.9 to 5.6%. This method was used to quantify the levels of dTp(Me)dT in enzymatic hydrolysates of DNA obtained from a series of incubations of salmon testis DNA or mouse lymphoma cells with either MNU or MMS.     

Development and validation of an improved method for the quantitation of sertraline in human plasma using LC-MS-MS and its application to bioequivalence studies

A rapid and sensitive LC-MS-MS method for the quantitation of sertraline in human plasma was developed and validated. Sertraline and the internal standard, telmisartan, were cleaned up by protein precipitation from 100 μL of plasma sample, and analyzed on a TC-C18 column (5 μm, 150 × 4.6 mm i.d.) using 70% acetonitrile and 30% 10 mM ammonium acetate (0.1% formic acid) as mobile phase. The method was demonstrated to be linear from 0.1 ng/mL to 50 ng/mL with the lower limit of quantitation of 0.1 ng/mL. Intra- and inter-day precision were below 4.40% and 3.55%. Recoveries of sertraline at low, medium, and high levels were 88.0 ± 2.3%, 88.2 ± 1.9%, and 90.0 ± 2.0%, respectively. The method was successfully applied to a bioequivalence study of sertraline after a single oral administration of 50 mg sertraline hydrochloride tablets.     

Quantitation of 8-hydroxydeoxyguanosine in DNA by liquid chromatography/positive atmospheric pressure photoionization tandem mass spectrometry

A methodology has been developed and validated for quantifying 8-hydroxydeoxyguanosine (8-OHdG) in both commercial DNA and DNA isolated from livers of male Sprague-Dawley rats by liquid chromatography/positive atmospheric pressure photoionization tandem mass spectrometry. The analytical method conditions, including conditions for stabilizing 8-OHdG during complex nuclease P1 enzymatic digestion, were also evaluated. The limit of detection for 8-OHdG was 1.0 ng/mL (17.6 fmol on-column), and the linearity of the calibration curve was greater than 0.998 from 1.0 to 500 ng/mL. The intraday assay precision relative standard deviation (RSD) value for quality control (QC) samples was < or =5.59% with accuracies ranging from 91.84 to 117.61%. The interday assay precision (RSD) value was < or =1.76% with accuracies ranging from 91.84 to 116.67%. This method, combined with the LC/UV analysis of deoxyguanosine (dG), was used for determination of the levels of 8-OHdG/10(6) dG in DNA nuclease P1 enzymatic hydrolysates from both commercial DNA and rat liver DNA.     

On-line solid phase extraction using the Prospekt-2 coupled with a liquid chromatography/tandem mass spectrometer for the determination of dextromethorphan, dextrorphan and guaifenesin in human plasma

An on-line liquid chromatography/tandem mass spectrometry (LC-MS/MS) procedure, using the Prospekt- 2 system, was developed and used for the determination of the levels of the active ingredients of cough/cold medications in human plasma matrix. The experimental configuration allows direct plasma injection by performing on- line solid phase extraction (SPE) on small cartridge columns prior to elution of the analyte(s) onto the analytical column and subsequent MS/MS detection. The quantitative analysis of three analytes with differing polarities, dextromethorphan (DEX), dextrorphan (DET) and guaifenesin (GG) in human plasma presented a significant challenge. Using stable-isotope-labeled internal standards for each analyte, the Prospekt-2 on-line methodology was evaluated for sensitivity, suppression, accuracy, precision, linearity, analyst time, analysis time, cost, carryover and ease of use. The lower limit of quantitation for the on-line SPE procedure for DEX, DET and GG was 0.05, 0.05 and 5.0 ng mL(-1), respectively, using a 0.1 mL sample volume. The linear range for DEX and DET was 0.05-50 ng mL(-1) and was 5-5,000 ng mL(-1) for GG. Accuracy and precision data for five different levels of QC samples were collected over three separate days. Accuracy ranged from 90% to 112% for all three analytes, while the precision, as measured by the %RSD, ranged from 1.5% to 16.0%     

Determination of eight selected organophosphorus insecticides in postmortem blood samples using solid-phase extraction and gas chromatography/mass spectrometry

A simple, rapid and sensitive method is described for the determination of omethoate, dimethoate, diazinon, chlorpyrifos, parathion‐ethyl, chlorfenvinphos, quinalphos and azinphos‐ethyl in postmortem whole blood samples. The analytes and internal standard (ethion) were isolated from the matrix by solid‐phase extraction, and were analysed by gas chromatography/mass spectrometry in the selected ion monitoring mode. The method has shown to be selective after analysis of postmortem samples of 40 different origins. Calibration curves were established between 0.05 (0.1 for omethoate) and 25 µg/mL, and the values obtained for intra‐ and interday precision and accuracy were within the criteria usually accepted for bioanalytical method validation. Lower limits of quantitation were 50 ng/mL for all compounds, except for omethoate (100 ng/mL); the limits of identification of the method were 25 ng/mL for all analytes, except for omethoate, for which 50 ng/mL was obtained. Absolute recovery was determined at three concentration levels, and ranged from 31 to 108%. The proposed method is simple and fast, and can be routinely applied in the determination of these compounds in postmortem whole blood samples within the scope of forensic toxicology. In addition, mass spectrometry has demonstrated to be a powerful and indispensable tool for the unequivocal identification of the analytes, since the acceptance criteria were accomplished even at very low levels, thus allowing obtaining forensically valid and sound results. Copyright © 2010 John Wiley & Sons, Ltd.     

High-throughput quantification of isoflavones, biochanin A and genistein, and their conjugates in female rat plasma using LC-ESI-MS/MS: Application in pharmacokinetic study

Isoflavones containing foods and dietary supplements are widely consumed for putative health benefits (e.g. cancer chemoprevention, beneficial effects on serum lipids associated with cardiovascular health, reduction of osteoporosis, relief of menopausal symptoms). This paper describes the development and validation of a sensitive high throughput LC‐ESI‐MS/MS method for quantifying biochanin A (BCA) and genistein (GEN), and their conjugates in rat plasma. The analytes were separated on a Supelco Discovery C18 (4.6×50 mm, 5.0 μm) column under isocratic condition using acetonitrile/methanol (50:50, v/v) and 0.1% acetic acid in the ratio of 90:10 v/v as a mobile phase. The intra‐ and inter‐day assay precision ranged from 2.66 to 8.34% and 4.40 to 8.10% (RSD %), respectively, and intra‐ and inter‐day assay accuracy was between 90.67–109.25% and 95.86–106.32%, respectively, for both the analytes. The lowest quantitation limit for BCA and GEN was 0.5 ng/mL in 0.1 mL of rat plasma. The method was successfully applied to the estimation of BCA, GEN and their conjugates in rat plasma following oral administration of BCA. Circulating conjugates (glucuronides/sulfates) of BCA and GEN were quantified using enzymatic hydrolysis of plasma samples. The levels of isoflavones glucuronides/sulfates were found to be much greater than the corresponding aglycones.     

Quantitation of glutathione by liquid chromatography/positive electrospray ionization tandem mass spectrometry

Glutathione (GSH) is a tripeptide composed of glutamate, cysteine, and glycine. It is present in practically all cells and has several important roles, such as preventing the oxidation of the sulfhydryl groups of proteins within a cell. Evidence for GSH deficiency or depletion has been found in a variety of diseases and toxicity-related studies, including diabetes and induction of oxidative stress to form reactive oxygen species which cause DNA, lipid, and protein oxidations. A simple, selective, and sensitive analytical method for measuring low levels of GSH in biological fluids would therefore be desirable to conduct GSH deficiency or depletion-related mechanistic toxicity studies. Here a method for both low- and high-level quantitation of GSH from cultured cells and rat liver tissues via liquid chromatography/positive electrospray ionization tandem mass spectrometry (LC/ESI-MS/MS) has been developed. The lower limit of quantitation (LOQ) of the method was 5 ng/mL. The method is linear over a wide dynamic concentration range of 5.0 to 5000.0 ng/mL, with a correlation coefficient R2 > 0.99. The intra-day assay precision relative standard deviation (RSD) values for all quality control (QC) samples were < or =16.31%, with accuracy values ranging from 94.13 to 97.80%. The inter-day assay precision RSD values for all QC samples were < or =15.94%, with accuracy values ranging from 94.51 to 100.29%. With this method, low levels of GSH from diethyl maleate (DEM)-treated mouse lymphoma cells, and GSH in rat liver tissues, were quantified.     

A solid phase extraction method for the screening and determination of pyrethroid metabolites and organochlorine pesticides in human urine.

A screening method has been developed for the determination of 23 organochlorine pesticides (OCPs) and 3-pyrethroid metabolities [cis- and trans-3-(2,2-dichlorovinyl)-2,2-dimethyl-(1-cyclopropane) carboxylic acid, cis-3-(2,2-dibromovinyl)-2,2-dimethyl-(1-cyclopropane) carboxylic acid and 3-phenoxybenzoic acid] from human urine. OCPs were directly detected in urine samples while pyrethroid metabolites required acid-induced hydrolysis to convert their conjugates into free acids; all compounds were then cleaned-up/preconcentrated using solid phase extraction. Determination and quantitation was achieved by gas chromatography with a mass spectrometer detector operating in selected ion monitoring mode. Limits of detection varied between 0.1 and 0.3 ng/mL with linear ranges from 0.3 to 700 ng/mL; the precision of the method was high (4.3-7.2%). Recoveries of all analytes from urine samples fortified at levels of 30 ng/mL for each OCP and 15 ng/mL for each pyrethroid metabolite ranged from 88 to 101% (captan gave the lowest recovery). The results obtained from the analysis of real urine samples show the suitability of the proposed method for monitoring people exposed to organochlorine and pyrethroid pesticides.     

Determination of seven selected antipsychotic drugs in human plasma using microextraction in packed sorbent and gas chromatography–tandem mass spectrometry

A method using microextraction by packed sorbent (MEPS) and gas chromatography–tandem mass spectrometry (GC-MS/MS) is described for the determination of seven antipsychotic drugs in human plasma. The studied compounds were chlorpromazine (CPZ), haloperidol (HAL), cyamemazine, quetiapine, clozapine, olanzapine (OLZ), and levomepromazine; promazine, protriptyline, and deuterated CPZ were used as internal standards. The validation parameters included selectivity, linearity and limits of detection and quantitation, intra- and interday precision and trueness, recovery, and stability and were studied according to internationally accepted guidelines. The method was found to be linear between the lower limit of quantitation and 1000 ng/mL, except for OLZ and HAL (200 ng/mL), with determination coefficients higher than 0.99 for all analytes, and extraction efficiencies ranged from 62 to 92 %. Intra- and interday precision ranged from 0.24 to 10.67 %, while trueness was within a ±15 % interval from the nominal concentration for all analytes at all studied levels. MEPS has shown to be a rapid procedure for the determination of the selected antipsychotic drugs in human plasma, allowing reducing the handling time and the costs of analysis. Furthermore, GC-MS/MS has demonstrated to be a powerful tool for the simultaneous quantitation of the studied compounds, enabling obtaining adequate selectivity and sensitivity using a sample volume of as low as 0.25 mL.     

Determination of several synthetic cathinones and an amphetamine‐like compound in urine by gas chromatography with mass spectrometry. Method validation and application to real cases

Most routine practices for drugs‐of‐abuse testing do not include screening procedures for new psychoactive substances, despite their increasing diffusion, preventing clear knowledge of the real consumption of these drugs in the populations. To make up for this shortcoming, a gas chromatography with mass spectrometry method was developed for the simultaneous determination of 18 synthetic cathinones and one amphetamine‐like compound in human urine. The sample preparation was based on liquid–liquid extraction under alkaline condition followed by derivatization with trifluoroacetic anhydride. The separation of the 19 analytes was achieved in less than 10 min. The whole methodology was validated according to national and international guidelines. Selectivity, linearity range, limit of detection and limit of quantitation, precision and accuracy were evaluated. For all the analytes, the calibration curve was linear in the 100–1000 ng/mL concentration range. The limits of detection ranged from 10 to 30 ng/mL and limits of quantitation from 30 to 100 ng/mL. Precisions were in the ranges 0.1–10.4%, and 1.0–12.1% for low (100 ng/mL) and high (1000 ng/mL) concentration, respectively. The accuracy, expressed as bias% was within ±20% for all the analytes. The present method was successfully applied to urine samples originating from autopsies, drug abuse/withdrawal controls, clinical investigations, roadside controls, driving re‐licensing, and workplace testing.     

Rapid and highly sensitive liquid chromatography/electrospray ionization tandem mass spectrometry method for the quantitation of buspirone in human plasma: application to a pharmacokinetic study

A rapid and sensitive method for the quantitation of buspirone in human plasma by liquid chromatography/electrospray ionization tandem mass spectrometry (LC/ESI-MS/MS) was developed. Plasma samples were treated by liquid-liquid extraction with methyl tert-butyl ether (MTBE). The chromatographic separation was performed isocratically on a reversed-phase Shiseido C18 column (50 mm x 2.0 mm, 3 microm) with a mobile phase of acetonitrile/0.1% acetic acid (1:1, v/v). The acquisition was performed in multiple reaction monitoring (MRM) mode, monitoring the transitions m/z 386 --> 122 for buspirone and m/z 409 --> 238 for amlodipine (the internal standard). The method was validated to determine its specificity, recovery, limit of quantitation, accuracy and precision. The lower limit of quantitation was 0.02 ng/mL with a relative standard deviation of less than 10%. The present method provides an accurate, precise and sensitive tool for buspirone and was successfully applied to a pharmacokinetic study in eight subjects.     

Liquid chromatographic determination of nicarbazin in feeds

A new liquid chromatographic method has been developed for determination of nicarbazin in feeds. Approximately 40 g feed is extracted with 200 mL acetonitrile-water (80 + 20, v/v). An aliquot of the extract is filtered and assayed using a reversed-phase isocratic method that measures the 4,4'-dinitrocarbanilide moiety of nicarbazin at a wavelength of 340 nm. For medicated feeds, the method uses a standard linear range of 5 to 100 microg/mL. For lower levels, a linear range of 50 to 150 ng/mL can be used. The method has a limit of detection of 250 ng/g and a limit of quantitation of 500 ng/g in a 40 g feed sample. Recovery was 99.1%, with a range of 95.2 to 101.8%. In the typical U.S. dosing range of 27 to 113.5 g/ton, the precision of the method based on one analyst, one day, and 2 weighings ranged from 2.8% (113.5 g/ton) to 4.7% (27 g/ton).     

Enantiomeric determination of praziquantel and its main metabolite trans-4-hydroxypraziquantel in human plasma by cyclodextrin-modified micellar electrokinetic chromatography

A capillary electrophoresis method for the simultaneous quantitation of praziquantel and its main metabolite trans-4-hydroxypraziquantel enantiomers in human plasma was developed and validated using cyclodextrin-modified micellar electrokinetic chromatography. Sample clean-up involved a single-step liquid-liquid extraction of plasma with toluene after the addition of NaCl. The complete enantioselective analysis was obtained in less than 7 min using 2% w/v sulfated beta-cyclodextrin as chiral selector and 20 mmol/L sodium deoxycholate as surfactant, in 20 mmol/L sodium borate buffer, pH 10. A 50 microm x 42 cm uncoated fused-silica capillary was used for the analysis, performed at a voltage of 18 kV and at 20 degrees C. The calibration curves were linear over the 125-625 ng/mL concentration range. The mean recoveries for praziquantel and trans-4-hydroxypraziquantel were up to 96 and 71%, respectively, with good precision. All four enantiomers were quantified at two concentration levels (200 and 600 ng/mL) with precision and accuracy below 15%. The quantitation limit was 50 ng/mL for (-)-(R)- and (+)-(S)-praziquantel and 62.5 ng/mL for (-)-(R)- and (+)-(S)-trans-4-hydroxypraziquantel, using 1 mL of human plasma.     

Liquid chromatography/mass spectrometry for the quantitation of muraglitazar in monkey plasma

A high-performance liquid chromatographic-mass spectrometric (LC/MS) assay was developed and validated for the determination of muraglitazar, a novel alpha/gamma, dual PPAR activator, in monkey plasma. The method utilized trazodone as the internal standard (IS). The extraction scheme involved a simple protein precipitation procedure with the use of a mixture of acetonitrile and methylene chloride. Separation was carried out on a BDS Hypersil C(18) analytical column (2 x 50 mm, 3 microm) and an effective chromatographic separation of muraglitazar (3.31 min) and trazadone (2.27 min) was achieved at a ssow rate of 0.3 mL/min. The mobile phase, used in an isocratic mode, consisted of 90% A (acetonitrile: 0.1% formic acid, 50:50 v/v) and 10% B (acetonitrile: 0.1% formic acid, 95:5 v/v). Detection of muraglitazar and trazodone was by positive ion turbo-ion spray mass spectrometry in the SIM mode. The mass spectrometer was programmed to admit the protonated molecules at m/z 372.0 (IS) and m/z 517.1 (muraglitazar). The standard curve, which ranged from 2 to 500 ng/mL, was fitted to a 1/x weighted linear regression model. The between run precision and within-run precision values of the assay was within 6.2% RSD. The assay accuracy was within 10.0% of the nominal values of the range of QC samples (6.0-400 ng/mL). At the lower limit of quantitation (LLQ) of 2 ng/mL, the deviation of the predicted concentrations from the nominal value of LLQ samples (n = 6) were within +/-16.6%. Muraglitazar was stable in monkey K(3)EDTA plasma for at least three freeze-thaw cycles. The processed samples (spiked samples) were stable for 48 h in auto-sampler at 10 degrees C. The average extraction recoveries of muraglitazar and IS were 83.3 and 91.9%, respectively. The assay was applied to delineate the pharmacokinetic disposition of muraglitazar in monkeys following a single oral dose.     

A new HPLC-UV validated method for therapeutic drug monitoring of tyrosine kinase inhibitors in leukemic patients

Development and validation of simple, rapid, and reliable high-performance liquid chromatography (HPLC)-UV method for quantification of major tyrosine kinase inhibitors, imatinib, dasatinib, and nilotinib, in human plasma is presented. Chromatographic separation of the drugs is achieved on an RP-C(18) column at flow rate of 0.9 mL/min at 35°C; eluate is monitored at 267 nm. Mean intra-day and inter-day precision for all compounds are 2.5 and 13.3%; mean accuracy is 13.9%; extraction recovery ranges within 40.24 and 81.81%. Calibration curves range from 10 to 0.005 μg/mL. Limits of detection are 10 ng/mL for imatinib and nilotinib, 50 ng/mL for dasatinib; limits of quantitation are 50 ng/mL for imatinib and nilotinib, 100 ng/mL for dasatinib. Although this method allows the detection of dasatinib, levels found in patients plasma are close to the limit of detection, then below the limit of quantitation. Quantification with HPLC-mass spectrometry, then, is required for dasatinib to give a correct evaluation. In conclusion, the sensitivity of this new method is sufficient to perform therapeutic monitoring and pharmacokinetic studies of imatinib and nilotinib but not dasatinib in CML patients.     

市售虾中磺胺甲噁唑的LC-MS-MS定量分析

当前各国对食品安全都极为重视,我国以及发达国家相继制定了相关兽药用量的法规和标准,尽管中国已经加入到了WTO,但我国的畜禽产品出口还是遇到了很多困难,农药、兽药、饲料添加剂等非法滥用,造成药物残留而直接影响到人类的健康。如引发癌症,婴儿畸形,代谢紊乱以及抗药性等诸多问题。其中磺胺类兽药是一类受到严格限制使用的兽药,本文研究了市售虾中磺胺甲(口恶)唑定量分析方法,检出限绝对量可达到0.5pg,完全满足发达国家的最低检出规定的苛刻标准。     

Determination of naphthodianthrones and phloroglucinols from Hypericum perforatum extracts by liquid chromatography/tandem mass spectrometry

Hypericum perforatum L. (St. John's Wort) has long been known as a medicinal plant, and has been used for the treatment of depression and neuralgic disorders. Its main active constituents are believed to be a naphthodianthrone, hypericin, and a phloroglucinol, hyperforin. A sensitive high performance liquid chromatography (HPLC)/electrospray tandem mass spectrometric method for fast simultaneous determination of six major naphthodianthrones and phloroglucinols of Hypericum perforatum extract has been developed. The method, based on multiple dissociation reaction monitoring (MRM), allows the analysis of hypericin, protohypericin, pseudohypericin, protopseudo-hypericin, hyperforin and adhyperforin from the extract in less than 5 min. Good linearity over the range 0.1-1000 ng/mL for hyperforin and 2-500 ng/mL for hypericin was observed. Intra-assay accuracy and precision varied from 2 to 19% within these ranges. Lower levels of quantitation for hyperforin were 0.5 ng/mL and 2 ng/mL for hypericin.     

Separation of glucooligosaccharides and polysaccharide hydrolysates by gradient elution hydrophilic interaction chromatography with pulsed amperometric detection

Commercial glucooligosaccharide mixtures (Polycose) and polysaccharide hydrolysates (acid and enzymatic) were fractionated by hydrophilic interaction chromatography and observed by pulsed amperometric detection. Seven peaks were observed when 625 ng of glucose oligomers in Polycose were fractionated. The between-run precision of retention times (n = 10, 100 μg, 15 peaks) ranged from a relative standard deviation (R.S.D.) of 0.09 to 0.40%; between-run precision of peak areas (n = 10) for the same separations had values that ranged from 2.66 to 14.4%. Injection-to-injection time was 48 min. When polysaccharide hydrolysates were fractionated using a gradient program capable of resolving all of the oligosaccharide species, dextran-derived -(1→6)- glucooligosaccharides were retained to a greater degree than amylose-derived -(1→4)-glucooligosaccharides, which were retained to a greater degree than β-(2→1)-fructooligosaccharides derived from inulin. Excluding the peaks that eluted before glucose or fructose, 25 to 35 peaks were observed after fractionation of the hydrolysates. Differences in elution profiles were observed between acid and enzymatic hydrolysis products of the same polysaccharide as well as between hydrolysis products of different polysaccharides. In conjunction with high-performance size-exclusion chromatography, the method demonstrated the effect of preheating starch before hydrolysis with isoamylase.     

Development of a liquid chromatography/tandem mass spectrometry assay for the quantification of Aplidin,a novel marine-derived antineoplastic agent,in human plasma

A rapid and sensitive liquid chromatography/tandem mass spectrometry (LC/MS/MS) assay was developed and validated to quantify a novel marine-derived depsipeptide, Aplidin, in human plasma. The method was validated to demonstrate the specificity, recovery, limit of quantitation (LOQ), accuracy, and precision of measurements. The calibration range for Aplidin was established using Aplidin standards from 0.05-50 ng/mL in blank human plasma. The multiple reaction monitoring, based on the transition m/z 1110.7 --> 295.3, was specific for Aplidin, and that based on the transition m/z 1112.6 --> 297.3 was specific for didemnin B (the internal standard); no endogenous materials interfered with the analysis of Aplidin and didemnin B from blank human plasma. The assay was linear over the concentration range 0.05-50.0 ng/mL. The correlation coefficients for the calibration curves ranged from 0.9979 to 0.9999. The mean intra- and interday accuracies for all calibration standards (n = 12) ranged from 97 to 106% (相似文献   

An improved ultra‐high‐performance liquid chromatography–tandem mass spectrometric method for the quantitation of dexmedetomidine in small volume of pediatric plasma

Dexmedetomidine (Dex), a highly selective α₂‐adrenergic agonist, is used primarily for the sedation and anxiolysis of adults and children in the intensive care setting. A sensitive and selective assay for Dex in pediatric plasma was developed by employing ultra‐high‐performance liquid chromatography–tandem mass spectrometry with d4‐Dex as an internal standard. Dex was extracted from 0.1 mL of plasma by micro‐elution solid‐phase extraction. Separation was achieved with a Waters XBridge C₁₈ column with a flow rate of 0.3 mL/min using a mobile phase comprising 5 mm ammonium acetate buffer with 0.03% formic acid in water and methanol–acetonitrile (50:50, v/v). The intra‐day precision (coefficient of variation) and accuracy for quality control samples ranged from 1.32 to 8.91% and from 92.8 to 108%, respectively. The inter‐day precision and accuracy ranged from 2.13 to 8.45% and from 97.0 to 104%, respectively. The analytical method showed excellent sensitivity using a small sample volume (0.1 mL) with a lower limit of quantitation of 5 pg/mL. This method is robust and has been successfully employed in a pharmacokinetic study of Dex in neonates and infants postoperative from cardiac surgery.