高效液相色谱法测定化妆品中5种季铵盐类防腐剂

建立了高效液相色谱法(HPLC)同时测定化妆品中5种季铵盐类防腐剂(西曲溴铵、西他氯铵、米他氯铵、劳拉氯铵和苄索氯铵)的检测方法。这5种季铵盐类防腐剂在15 min内能达到有效分离,在10~200μg/m L范围内线性良好,检出限在1.11~1.97μg/m L范围内,平均回收率为77.1%~110.2%。     

离子对色谱-抑制电导法测定6-溴己基三甲基溴化铵中相关物质北大核心CSCD

建立了离子对色谱法测定6-溴己基三甲基溴化铵中相关物质的分析方法。样品经超纯水溶解后过0.22μm滤膜进样分析,选用Shinesil C18色谱柱进行色谱分离,高氯酸和乙腈作为淋洗液进行梯度洗脱,采用电导检测器检测,外标法定量。结果表明,6-溴己基三甲基溴化铵和溴化六甲铵的峰面积与其质量浓度在一定范围内呈良好线性关系,检出限分别为1.076 mg/L、0.2327 mg/L,定量限分别为3.587 mg/L、0.7757 mg/L。以6-溴己基三甲基溴化铵样品进行加标回收试验,回收率在95.8%~103.3%之间,测定值的相对标准偏差(RSD,n=6)均小于1.10%。该方法简单快速,可用于6-溴己基三甲基溴化铵样品中主成分6-溴己基三甲基溴化铵和杂质溴化六甲铵的定性和定量分析。     

固相萃取结合超高效液相色谱-串联质谱法及气相色谱-负化学源质谱法测定人血清中的四溴双酚A、六溴环十二烷和多溴联苯醚

建立了固相萃取同时提取、净化血清中四溴双酚A(TBBPA)、α, β, γ-六溴环十二烷(HBCD)和8种多溴联苯醚(PBDEs)同系物的样本前处理方法,并结合色谱-质谱分离分析技术检测人血清样本中该类化合物的含量。试样在加入各自的同位素内标物后以甲基叔丁基醚/正己烷(1:1, v/v)混合溶剂进行萃取,再经浓硫酸去除脂肪后,以LC-Si固相萃取柱分离HBCD/TBBPA和PBDEs。采用分步检测的方式,在50 mm长BEH C18反相色谱柱上以超高效液相色谱-串联质谱(UPLC-MS/MS)的多反应监测模式(MRM)检测HBCD和TBBPA,在15 m长的毛细管柱上以气相色谱-负化学源质谱(GC-NCI/MS)的选择离子监测模式(SIM)检测PBDEs。以胎牛血清为空白基质,当HBCD、TBBPA和BDE-209的加标水平为0.5 ng/g和5 ng/g、三溴至七溴联苯醚的加标水平为0.05 ng/g和0.5 ng/g时,它们的平均加标回收率为80.3%～108.8%,相对标准偏差为1.02%～11.42%(n=5);以信噪比(S/N)为3计算,方法的检出限(LOD)为1.81～42.16 pg/g。采用该方法对实际样品进行测定,结果表明,本方法快速、准确、灵敏度高,能够满足血清中HBCD、TBBPA和PBDEs残留的同时提取及测定的要求。     

HPLC法分析复方异丙托溴铵溶液雾化气溶胶成分

建立HPLC法同时测定吸入用复方异丙托溴铵溶液雾化气溶胶中硫酸沙丁胺醇和异丙托溴铵两组分的含量。采用C₁₈(4.6mm×250 mm,5μm)色谱柱,流动相为1.2 g/L的1-庚烷磺酸钠溶液(磷酸调节pH 3.2左右)(A)-乙腈(B),等度洗脱,流速为1.0 mL/min,进样量为50μL,分析时间15 min,柱温40℃,检测波长为210 nm。结果表明,硫酸沙丁胺醇和异丙托溴铵分别在0.1~2.0 mg/L和0.075~2.4 mg/L浓度范围线性良好(r值均大于0.999)。复方异丙托溴铵溶液雾化气溶胶中硫酸沙丁胺醇和异丙托溴铵加样回收率分别为97.8%和102.3%。雾化气溶胶中硫酸沙丁胺醇和异丙托溴铵提取方法的回收率分别为96.1%和97.1%。在吸入暴露装置中经雾化5 min和35 min后,气溶胶中硫酸沙丁胺醇含量分别为22.39和3.21 mg/m³,异丙托溴铵含量分别为22.34和3.23 mg/m³,回收率均不低于99.5%,RSD不大于1.9%,表明雾化5 min即达到稳定。本方法适用于吸入用复方异丙托溴铵溶液雾化气溶胶的药物含量测定,也可为吸入制剂雾化气溶胶供试品分析提供参考。     

同系物季铵盐离子液体阳离子的离子交换色谱法分析

建立了离子交换色谱-直接电导检测法分离测定3种同系物季铵盐离子液体阳离子(四甲基铵、四乙基铵和四丙基铵阳离子)的方法。采用磺酸型阳离子交换色谱柱,以乙二胺-柠檬酸-乙腈为淋洗液,考察了淋洗液种类、浓度及色谱柱温度对3种阳离子保留的影响。并根据测定对象不同,调整乙二胺浓度及乙腈含量以改善分离效果。淋洗液中增加乙腈含量,可明显缩短四丙基铵阳离子的保留时间,并改善其色谱峰形。季铵阳离子同系物的保留符合碳数规律。优化的色谱条件为:流速1.0 mL/min,色谱柱温度40℃;以0.02 mmol/L乙二胺-0.12 mmol/L柠檬酸(pH 4.0)为淋洗液分离测定四甲基铵;以0.2 mmol/L乙二胺-0.4 mmol/L柠檬酸-1%乙腈(pH 4.0)为淋洗液分离测定四乙基铵和四丙基铵。所测阳离子的检出限(S/N=3)分别为0.015,0.22,1.88 mg/L,相对标准偏差(n=5)均小于2.3%。将方法应用于表面活性剂和实验室合成的离子液体的分析,加标回收率为99%~104%。本方法简单、准确、可靠,具有良好的实用性。     

离子色谱-非抑制电导法同时测定铵盐、四乙基铵、甲基三乙基铵

建立了离子色谱非抑制电导法同时分离测定铵根与两种季铵盐四乙基铵、甲基三乙基铵的方法。分别实验了在亲水性和疏水性阳离子交换色谱柱上三种铵类的分离效果,研究了使用不同淋洗液和流速情况下离子的分离情况,结果表明使用SH-Cation-101型疏水性阳离子色谱柱,淋洗液采用甲烷磺酸(5.0mmol/L),其中加入乙腈(7%),于0.8mL/min的流速条件下,三种铵类物质分离良好,其中结构极为相似的两种季铵盐四乙基铵和甲基三乙基铵分离度达到1.5以上,分离时间短,3种物质在13min内实现完全分离。采用国产离子色谱仪非抑制电导法检测,无需使用抑制器,成本低,操作简便可行。检测结果的灵敏度高,线性范围铵根为0.5～50mg/L,四乙基铵和甲基三乙基铵为5～500mg/L,相关系数均高于0.999,相对标准偏差均在3%以内,平均加标回收率在98.5%～101.2%。     

离子色谱-三重四极杆质谱法同时测定血浆和尿液中百草枯和敌草快

百草枯和敌草快是广泛使用的非选择性触杀型除草剂,中毒后会造成急性肺损伤,病死率高,同时监测血浆和尿液中百草枯和敌草快的浓度,可以为临床早期诊断和预后提供有价值的信息。血浆和尿液中百草枯和敌草快的主要检测方法为液相色谱-质谱法。百草枯和敌草快为强极性水溶性化合物,在反相色谱柱上难以保留,多采用离子对色谱法或亲水色谱法进行分离。采用离子对色谱法时,加入的离子对试剂有离子抑制作用,降低了质谱检测的灵敏度,还给质谱系统增加了额外的污染;亲水色谱法易受基质成分影响,保留时间不稳定。考虑到百草枯和敌草快在水溶液中以双电荷联吡啶离子状态存在,更适合采用阳离子交换色谱法,建立了离子色谱-三重四极杆质谱测定血浆和尿液中百草枯和敌草快的检测方法。血浆和尿液样品经水稀释后,直接过混合型聚合物反相吸附和弱阳离子交换固相萃取柱（Oasis WCX）净化,经IonPac CS 18型阳离子色谱柱（250 mm×2.0 mm,6.0 μm）分离,以自动在线产生的甲磺酸进行梯度洗脱,色谱柱流出液经阳离子抑制器抑制后进入质谱系统,在ESI⁺ 、多反应监测（MRM）模式下检测,稳定同位素内标法定量。百草枯和敌草快分别在1.0~150 μg/L和0.5~75 μg/L范围内线性关系良好,血浆中的平均基质效应分别为84.2%~89.3%和84.7%~91.1%,尿液中平均基质效应分别为50.3%~58.4%和51.9%~59.4%;血浆中百草枯和敌草快的平均加标回收率分别为93.5%~117%和91.7%~112%,相对标准偏差（RSD）分别为3.4%~16.7%和2.8%~13.2%;尿液中百草枯和敌草快的平均加标回收率分别为90.0%~118%和99.2%~116%,RSD分别为5.6%~14.9%和2.4%~17.3%（n =6）;血浆和尿液中百草枯和敌草快的检出限分别0.3 μg/L和0.2 μg/L,定量限分别为1.0 μg/L和0.5 μg/L。该法灵敏度高,准确性好,可用于血浆和尿液中百草枯和敌草快的中毒检测。     

改进QuEChERS法结合超高效液相色谱-串联质谱法同时测定冷冻食品中7种季铵盐化合物

近年来,季铵盐类消毒剂的广泛使用,导致其残留在食品或环境中,并经食物链进入人体,引起呼吸系统和生殖系统的不良反应,威胁人类身体健康。为此,建立了QuEChERS-超高效液相色谱-串联质谱(QuEChERS-UPLC-MS/MS)同时测定冷冻食品中6种常规季铵盐化合物和1种新型季铵盐化合物残留的分析方法。对样品前处理方法进行改良,色谱条件进行优化,样品经20 mL 90%甲醇水溶液(含0.5%甲酸)涡旋提取20 min后,超声10 min, 10000 r/min离心10 min,取1 mL上清液放入装有100 mg PSA净化剂的15 mL离心管中净化,涡旋1 min后,10000 r/min离心5 min,取上清液直接上机测定。以甲醇与5 mmol/L乙酸铵溶液作为流动相进行梯度洗脱,采用ACQUITY UPLC BEH C₈色谱柱(50 mm×2.1 mm, 1.7 μm)分离,在ESI⁺模式,多反应监测(MRM)方式下采集数据,基质匹配曲线外标法定量。结果表明,7种待测物在C₈色谱柱上完全基线分离,在0.1~100.0 μg/L的范围内线性关系良好,相关系数(r²)为0.9971~0.9983,该方法的检出限(LOD)为0.5~1.0 μg/kg,定量限(LOQ)为1.5~3.0 μg/kg。以阴性三文鱼、鸡肉为基质样品,在低、中、高3个水平(3.0、10.0、100.0 μg/kg)下进行加标回收试验,各待测物的平均回收率为65.4%~101%,相对标准偏差(RSD)为0.64%~16.8%。该检测方法灵敏度高,选择性强,稳定性好,结果准确可靠,适用于冷冻食品中7种季铵盐化合物的批量快速测定,同时,弥补了新型季铵盐化合物检测方法的空白。     

凝胶渗透色谱-分散固相萃取法同时测定鸡蛋中多溴联苯醚及其衍生物、四溴双酚A和六溴环十二烷

建立了一种同时检测鸡蛋中四溴双酚A(TBBP A)、六溴环十二烷(HBCD)和多溴联苯醚(PBDEs)及其衍生物羟基多溴联苯醚(OH-PBDEs)和甲氧基多溴联苯醚(MeO-PBDEs)的凝胶渗透色谱(GPC)-分散固相萃取(DSPE)-液相色谱-串联质谱(HPLC-MS/MS)和气相色谱-负化学源质谱(GC-NCI/MS)的检测方法。样品经正己烷、二氯甲烷(1∶1,V/V)加速溶剂萃取,凝胶渗透色谱净化后,经100 mg十八烷基键合硅胶(C18)分散固相萃取吸附剂去除杂质,液相色谱-串联质谱和气相色谱-负化学源质谱方法测定,外标法定量。在蛋白和蛋黄样品中添加1.0或5.0μg/kg的目标物,其回收率分别为64.5%~97.2%和65.6%~109.2%(除BDE-85为54.8%,OH-BDE-137为47.4%外),相对标准偏差小于20.2%,定量限为0.01~0.2μg/kg。     

气相色谱/离子阱串联质谱法检测大气中的多溴二苯醚

运用气相色谱-离子阱质谱(ion trap mass spectrometry,ITMS)建立了大气中8种多溴二苯醚的二级质谱(MS/MS)检测方法。优化后的MS/MS方法对7种三溴至七溴二苯醚(BDE28、-47、-99、-100、-153、-154、-183)和十溴二苯醚(BDE209)的仪器检出限分别为0.04~0.23 pg和9.38 pg;方法检出限为6.0~27.1 pg/m3和211.6 pg/m3;6点标准曲线相关系数(r)为0.9980~0.9999。方法对8种目标物的加标回收率为79.1%~101.0%,相对标准偏差(RSD)为4.3%~14.3%(n=6)。对6份南京实际大气样品的检测结果表明:离子阱质谱的选择反应监测(selected reaction monitoring,SRM)技术能有效的降低背景干扰。优化后的MS/MS方法对目标化合物有较高的灵敏度,可应用于大气中痕量多溴二苯醚的定量检测。     

Analysis of steroids. XLI. Ion-pair high-performance liquid chromatographic separation of quaternary ammonium steroids on silica.

A normal-phase ion-pair chromatographic system has been developed for the high-performance liquid chromatographic investigation of pipecuronium bromide (2 beta,16 beta-bis-(N'-dimethyl-l-piperazinyl)-3 alpha,17 beta-diacetoxy-5 alpha-androstane dibromide) and related quaternary ammonium steroids. The use of silica as the stationary phase and a 96:4 mixture of acetonitrile and water containing 0.1 mol/dm3 sodium perchlorate as the eluent with detection at 213 nm enable the potential impurities as well as the hydrolytic and oxidative degradation products of pipecuronium bromide to be separated and detected down to the 0.01% level. The above system is also applicable to the high-performance liquid chromatographic investigation of other quaternary ammonium steroids (pancuronium bromide, vecuronium bromide).     

Mass Spectrometric and Contactless Conductivity Detection Approaches in the Determination of Muscle Relaxants by Capillary Electrophoresis

Novel and rapid capillary electrophoresis-coupled tandem mass spectrometry (CE-MS/MS) and capillary electrophoresis with capacitively coupled contactless conductivity detection (CE-C⁴D) methods have been developed for the separation and determination of three neuromuscular blocking agents: pancuronium, vecuronium, and rocuronium. In both cases, the separation was conducted in background electrolytes based on acidic acetate-ammonium buffers to avoid possible decomposition of the analytes that are known to be unstable in alkaline media. Baseline resolution of the analytes was achieved in the presence of modified γ-cyclodextrin by CE with C⁴D detection. The two detection techniques were compared with regard to analytical figures of merit including linear dynamic range, limit of detection, limit of quantification, precision, and accuracy. The calibration curves showed good linearity for both detection methods examined (characterized by r² ≥ 0.9908). The LODs of the CE-MS/MS and the CE-C⁴D methods differed at least by two orders of magnitude considering all analytes. The differences in precision and accuracy of these methods were evaluated and discussed. The assays of pancuronium, vecuronium, and rocuronium in commercial injection solutions by CE-MS/MS and CE-C⁴D were performed and the results compared.     

Detection of Quaternary Ammonium Drugs in Equine Urine by Liquid Chromatography-Mass Spectrometry

Quaternary ammonium drugs are anticholinergic agents and some of which have been known to be abused in equine sports. A general screening method for this class of drugs in equine urine by liquid chromatography-mass spectrometry (LC-MS) has not been reported. The paper describes an efficient LC-MS-MS method for the simultaneous detection and confirmation of twenty quaternary ammonium drugs at low ng mL^?1 in equine urine after solid-phase extraction. Quaternary ammonium drugs were extracted from equine urine by solid-phase extraction using ISOLUTE^® CBA SPE Columns and analysed by LC-MS-MS in the positive electrospray ionisation mode. Separation of twenty quaternary ammonium drugs (the quaternary ammonium ions of edrophonium chloride, pyridostigmine bromide, neostigmine bromide, bretylium tosylate, ipratropium bromide, tubocurarine chloride, N-butylscopolammonium bromide, mepenzolate bromide, rocuronium bromide, clidinium bromide, pipenzolate bromide, isopropamide iodide, glycopyrronium bromide, alcuronium chloride, oxyphenonium bromide, propantheline bromide, tridihexethyl chloride, vecuronium bromide, mivacurium chloride and pancuronium bromide) was achieved in a reversed phase column with a mixture of aqueous ammonium formate (pH 3.0, 10 mM) and acetonitrile as the mobile phase. Detection and confirmation of the twenty quaternary ammonium drugs at about 5 ng mL^?1 in equine urine could be achieved within 22 min using product-ion scan MS-MS. The target analytes were detected by examination of extracted-ion chromatograms of their product ions. Drugs spiked in different equine urine (n = 15) were consistently detected. Negative samples (n = 30) of normal post-race equine urine have also been analysed, no matrix interference at the targeted masses and retention times was observed. The method was successfully applied to the analyses of drug-administration samples. Other method validation data including reproducibility and recovery will also be presented. An LC-MS-MS method for the simultaneous detection and confirmation of twenty quaternary ammonium drugs in equine urine was developed. The methodology should be applicable to other biological matrices such as human urine.     

An ion-pairing liquid chromatography/tandem mass spectrometric method for the determination of ethephon residues in vegetables

A rapid, sensitive and selective method has been developed for the direct determination of ethephon residues in vegetables (apple, cherry and tomato). Given the anionic character of ethephon, the use of ion-pairing liquid chromatography (LC) in combination with tandem mass spectrometry (MS/MS, triple quadrupole) allowed its direct determination in these matrices avoiding a derivatisation step and favouring the automation of the method. Samples were extracted with a mixture of dichloromethane/aqueous formic acid (pH 3) (1:1). Then, tetrabutylammonium acetate (TBA) was added as an ion-pairing reagent, and an aliquot of the aqueous extract was directly injected into the LC/MS/MS system. Quantification was performed with matrix-matched standards prepared from blank sample extracts. MS/MS measurements were made in the selected reaction monitoring (SRM) mode, using the most sensitive transition (m/z 107 > 79) for quantification, and up to four additional transitions for confirmation. Quantitative recoveries were obtained for all matrices (between 83% and 96%) at two concentration levels tested (0.05 and 0.5 mg/kg), with relative standard deviations lower than 9% in all cases. The addition of TBA directly into the sample extract contained in the injection vial was found sufficient to obtain satisfactory LC retention for the analyte. Under these conditions, the absence of ion-pairing reagent in the mobile phase minimised the ionisation suppression for ethephon in the MS source, leading to an increase in the sensitivity of the method and reaching limits of detection of 0.02 mg/kg for all matrices investigated. The acquisition of five specific MS/MS transitions for ethephon allowed the simultaneous and reliable quantification and confirmation of the analyte in the samples.     

Effects of ion-pairing reagents on the electrospray signal suppression of sulphonated dyes and intermediates

High-performance liquid chromatography/mass spectrometry (HPLC/MS) analysis of anionic species such as sulphonic acid dyes and intermediates requires volatile ion-pairing mobile phase additives. Six di- and trialkylammonium acetates were compared with tetraalkylammonium salts and ammonium acetate in the concentration range 0-20 mmol l(-1) as mobile phase additives for HPLC/MS of polysulphonated compounds. The effects of the structure and concentration of the ion-pairing reagents on the electrospray response of mono-, di- and tetrasulphonic aromatic acids and acid dyes were studied in detail. Further, five different mass analysers and instrument geometries were compared. A higher signal decrease is observed with linear geometry instruments in comparison to orthogonal or even Z-spray geometry mass spectrometers. The concentration of mobile phase additives has a significant influence on the abundance ratios of multiply charged ions in the mass spectra of polysulphonated compounds. The competing ions of sulphonic acids may also cause significant signal suppression.     

Arsenic speciation in Chinese brake fern by ion-pair high-performance liquid chromatography-inductively coupled plasma mass spectroscopy

Ion-pair reverse-phase HPLC-inductively coupled plasma (ICP) MS was employed to determine arsenite [As(III)], dimethyl arsenic acid (DMA), monomethyl arsenic (MMA) and arsenate [As(V)] in Chinese brake fern (Pteris vittata L.). The separation was performed on a reverse-phase C18 column (Haisil 100) by using a mobile phase containing 10 mM hexadecyltrimethyl ammonium bromide (CTAB) as ion-pairing reagent, 20 mM ammonium phosphate buffer and 2% methanol at pH 6.0. The detection limits of arsenic species with HPLC-ICP-MS were 0.5, 0.4, 0.3 and 1.8 ppb of arsenic for As(III), DMA, MMA, and As(V), respectively. MMA has been shown for the first time to experimentally convert to DMA in the Chinese brake fern, indicating that Chinese brake fern can convert MMA to DMA by methylation.     

High performance liquid chromatography-mass spectrometric analysis of sulphonated dyes and intermediates

This paper reports our results in the analysis of polysulphonated anionic dyes and their intermediates using high-performance liquid chromatography-mass spectrometry (HPLC-MS). Negative-ion electrospray ionization is the most suitable ionization technique for the molecular mass determination of polysulphonated dyes or other dyes carrying a negative charge. From the series of [M-xH]x- ions and their sodiated adducts [M-(x + y)H+yNa]x-, the molecular mass and the number of sulphonic and carboxylic groups can be determined. The mobile phase should be compatible with the mass spectrometric detection, which rules out non-volatile tetraalkylammonium salts usually used as ion-pair mobile phase additives for the HPLC of sulphonated compounds. Some mono- and disulphonated dyes and intermediates can be separated with aqueous-organic mobile phases containing 5 mM ammonium acetate, which is the most suitable additive as far as compatibility with MS detection is concerned. However, the retention of compounds with two or more sulphonic groups is too low for a successful separation both with this mobile phase additive and with ion-pair additives with short alkyl chains. The dihexylammonium acetate ion-pairing reagent offers a reasonable compromise in terms of sufficient volatility and adequate retention and separation selectivity for the HPLC-MS analysis of polysulphonated dyes.     

Determination of amphetamine, methamphetamine, 3,4-methylenedioxyamphetamine, 3,4-methylenedioxyethylamphetamine, and 3,4-methylenedioxymethamphetamine in urine by online solid-phase extraction and ion-pairing liquid chromatography with detection by electrospray tandem mass spectrometry

A method using an online solid-phase extraction (SPE) and ion-pairing liquid chromatography with electrospray tandem mass spectrometry (LC/ES-MS/MS) was developed for determination of amphetamine (Amp), methamphetamine (mAmp), 3,4-methylenedioxyamphetamine (MDA), 3,4-methylenedioxyethylamphetamine (MDEA), and 3,4-methylenedioxymethamphetamine (MDMA) in urine samples. A SPE cartridge column with both hydrophilic and lipophilic functions was utilized for online extraction. A reversed-phase C18 LC column was employed for LC separation and MS/MS was used for detection. Trifluoroacetic acid was added to the mobile phase as an ion-pairing reagent. This method was fully automated and the extraction and analysis procedures were controlled by a six-port switch valve. Recoveries ranging from 85-101% were measured. Good linear ranges (10-500 ng/mL) for Amp and mAmp were determined. For MDA, MDMA and MDEA, dual linear ranges were obtained from 5-100 and 100-500 ng/mL, respectively. The detection limit of each analytical compound, based on a signal-to-noise ratio of 3, ranged from 1-3 ng/mL. The applicability of this newly developed method was examined by analyzing several urine samples from drug users. Good agreement was obtained between the results from this method and a literature GC/MS method.     

液相色谱中的一个新的表征参数Z Ⅲ.反相高效液相色谱中的生物大分子分离体系

研究了在反相高效液相色谱中的烷基醇-甲酸-水、烷基醇-三氟醋酸-水和烷基醇-磷酸盐-水三种流动相,9种生物大分子的分离体系中Z的表征。发现当蛋白质完全变性时,Z值与其分子量呈正比,而与置换剂分子大小呈反比。Z值会因流动相中离子对试剂的存在而减小,且浓度愈大,减小程度愈甚。对于在异丙醇-水溶液中的不同种类离子对试剂而言,对蛋白构象变性影响的减小顺序为44％(V/V)甲酸>0.05mol/L磷酸盐缓冲液(pH<2.5)>0.1％(V/V)三氟醋酸。     

Determination of amphetamine in rat brain by in vivo microdialysis and ion-pairing liquid chromatography with electrospray tandem mass spectrometry

An ion-pairing liquid chromatography/electrospray tandem mass spectrometry (LC/ES-MS/MS) method with in vivo microdialysis for the determination of amphetamine in rat brain has been developed. A microdialysis probe was surgically implanted into the striatum of the rat and artificial cerebrospinal fluid (aCSF) was used as the perfusion medium. Samples were collected and then analyzed off-line by LC/ES-MS/MS. A reversed-phase C18 column was employed for LC separation and MS/MS was utilized for detection. Trifluoroacetic acid (TFA) was added to the mobile phase (acetonitrile/water) as an ion-pairing reagent. Detection was by ES-MS/MS directly, and no post-column addition of organic modifier was needed. Dual linear ranges were determined from 0.1-0.5 microg/mL and 0.005-0.1 microg/mL, respectively. The detection limit, based on a signal-to-noise ratio of 3, was 0.001 microg/mL (5 nM). Good precision and accuracy were obtained. The applicability of this newly developed method was demonstrated by continuous monitoring of amphetamine concentrations in rat brain. Amphetamine reached a maximum concentration of 0.086 +/- 0.017 microg/mL over 20-40 min after a single 3.0 mg/kg intraperitoneal administration.