高效液相色谱法测定茶叶中的茶氨酸

建立了未衍生化高效液相色谱法(HPLC)测定茶叶中茶氨酸含量的方法。采用的色谱条件为:C 18 色谱柱,以0.05%(体积分数)三氟乙酸水溶液为流动相,流速1 mL/min ,进样量10 μL,检测波长203 nm。茶氨酸质量浓度在0.02～1 g/L 内,其浓度与峰面积呈良好的线性关系,最低检出限为1.75 ng(S/N=3),回收率为97.2%,相对标准偏差(RSD)为1.7%。同时以高效液相色谱-电喷雾离子化质谱对所分离的茶氨酸进行了纯度鉴定。方法具有精确、灵敏、流动相组成简单等特点。     

高效液相色谱-紫外检测法测定人体尿液中的蝶呤-6-羧酸

建立了人体尿液中蝶呤-6-羧酸的高效液相色谱-紫外分析新方法. 运用Lichrospher C18柱(250×4.6 mm,5 μm),甲醇-水(70∶30,V/V)为流动相,流速为0.4 mL/min,可较好地将尿液中蝶呤-6-羧酸与其它共存干扰物质分离.在355 nm检测波长下,蝶呤-6-羧酸在0.19～4.8 μg/mL范围内与色谱峰面积呈良好线性关系,相关系数为0.9999,方法检出限为0.015 μg/mL.尿液经0.45 μm滤膜过滤后,可直接进样分析,方法简便,应用于癌症病人和健康人尿样中蝶呤-6-羧酸测定,结果较好.方法的加标回收率为94.6%～100.2%,相对标准偏差为0.81%～ 5.07% .     

高效液相色谱法测定化妆品原料中N-乙酰神经氨酸

建立高效液相色谱法测定化妆品原料中N-乙酰神经氨酸含量的方法。以体积分数为50%的乙酸溶液为水解溶剂,样品于80℃水浴中加热30 min,以CAPCELL PAK SCX柱（250 mm×4.6 mm,5μm）为分析柱,流动相为乙腈-0.1%磷酸水溶液（体积比为80∶20）,等度洗脱,流量为1.0 mL/min,柱温为35℃,采用高效液相色谱二极管阵列检测器测定,以色谱峰面积外标法定量。检测波长为205 nm,进样体积为10μL。N-乙酰神经氨酸的色谱峰面积与质量浓度在10.0～500.0μg/ml范围内线性关系良好,相关系数为0.999 9,方法检出限为0.12μg/mL,定量限为0.4μg/mL。样品加标平均回收率为97.7%～98.9%,测定结果的相对标准偏差为0.37%～1.36%(n=6)。该方法样品处理简便、高效,适用于化妆品原料中N-乙酰神经氨酸的含量测定。     

气相色谱法测定乙酸阿比特龙中1,1,3,3-四甲基胍

建立气相色谱测定乙酸阿比特龙原料药中1,1,3,3-四甲基胍的方法。选择乙酸乙酯为溶剂溶解样品，以安捷伦CP-Volamine毛细管柱（30 m×0.32 mm,0.45μm）作为分离色谱柱，采用氢火焰离子化检测器（FID）进行检测，外标法定量分析。1,1,3,3-四甲基胍的质量浓度在10.22～204.38μg/mL范围内与色谱峰面积线性关系良好，相关系数为0.999 4，定量限为10.22μg/mL。样品加标回收率为97.9%～113.7%,9份样品测定结果的相对标准偏差为5.0%。该方法操作简单，可用于乙酸阿比特龙原料中1,1,3,3-四甲基胍的测定。     

柱前衍生-高效液相色谱法测定鱼类中组胺

建立测定鱼类中组胺含量的柱前衍生-高效液相色谱（HPLC）联合紫外检测分析方法。样品均质后先用三氯乙酸水溶液震荡超声提取,再用丹磺酰氯衍生,采用HPLC-紫外检测器在254 nm处对组胺衍生物进行检测,以色谱峰面积外标法定量。组胺的质量浓度在1.0～50.0μg/mL范围内与色谱峰面积线性关系良好,相关系数为0.999 7,方法检出限为7.2μg/kg,定量限为24μg/kg。样品加标回收率为96.8%～99.2%,测定结果的相对标准偏差为1.6%～3.7%(n=6)。该方法快速准确、灵敏度高、重复性好,可用于鱼类产品中组胺的定量分析。     

高效液相色谱法定量分析奥曲肽

建立奥曲肽的高效液相色谱定量分析方法。色谱柱为Eclipse plus C18柱(4.6 mm×250 mm,5 μm),流动相为乙腈-0.25%高氯酸水溶液(体积比为30∶70),流量为1.0 mL/min,检测波长为210 nm,柱温为25℃。奥曲肽的质量浓度在4.38~219 μg/mL范围内与色谱峰面积成良好的线性关系,相关系数为0.9999,检出限为1.1 ng,定量限为2.19 ng。测定结果的相对标准偏差为0.26%~0.46% (n=5),加标回收率为97.41%~100.26%。该方法简便、快速、准确,适用于奥曲肽原料药与制剂的定量分析。     

固相萃取-高效液相色谱法测定啤酒中的黄腐酚

采用Waters Sep-Pak C18固相萃取小柱对啤酒样品进行分离纯化,建立了啤酒中黄腐酚的固相萃取-高效液相色谱检测方法。选用色谱柱Zorbax Eclipse XDB-C18柱(250 mm×4.6 mm,5 μm),以甲醇和0.1%甲酸水溶液为流动相进行梯度洗脱,柱温25 ℃,流速0.4 mL/min,检测波长370 nm。在此条件下,黄腐酚分离良好且无杂质峰干扰,在0.5～500 μg/L的范围内线性关系良好(r21),在高、中、低浓度下的加标回收率为91.21%～95.58%,相对标准偏差小于2%。方法的检出限为0.24 μg/L,定量限为0.80 μg/L。该方法简便快速、结果准确、重现性好,是检测啤酒中黄腐酚含量的有效方法。     

气相色谱法测定医疗防护用品中环氧乙烷残留量

建立气相色谱法检测口罩、防护服等医疗防护用品中环氧乙烷的残留量,为生产厂家改进灭菌工艺提供数据支持。气相色谱法采用顶空进样、FID检测器,环氧乙烷的质量浓度在0.5～8 μg/mL范围内与色谱峰面积线性关系良好,检出限为0.10 μg/g,定量限为0.35 μg/g。环氧乙烷测定结果的相对标准偏差为1.73%(n=6),平均加标回收率为94.3%～99.3%。该方法操作简便,精密度、准确度较高,适用于医疗防护用品中环氧乙烷的测定。     

液相色谱-电喷雾串联质谱法测定禽类产品中克球酚的残留

建立了禽类产品中克球酚残留的液相色谱-电喷雾串联质谱(LC-ESI-MS/MS)检测方法。用甲醇对样品进行提取,提取液用正己烷萃取去油脂,然后用LC-18柱和阴离子交换柱净化,LC-ESI-MS/MS测定。利用基质校正曲线对克球酚准确定量。在2,5,10,20 μg/kg 4个添加水平下,克球酚的平均回收率稳定在55.38%～132.44%之间,日内精密度小于9.54%,日间精密度小于15.27%。在1～40 μg/kg范围内色谱峰面积与克球酚含量呈良好的线性关系,检出限为0.5 μg/kg,定量限为2.0 μg/kg。该方法选择性好,抗干扰能力强,可作为禽类产品中克球酚残留检测的确证方法。     

高效液相色谱法测定冷面、米线中乙二胺四乙酸二钠

建立高效液相色谱法测定冷面和米线中乙二胺四乙酸二钠的分析方法。样品用水提取后,经三氯化铁络合,用Shimpack VP-ODS C₁₈色谱柱(250 mm×4.6 mm,5μm)分离,以四丁基溴化铵-乙酸钠混合溶液(pH 4.0)-甲醇(体积比为85∶15)为流动相,等度洗脱,采用紫外检测器检测,检测波长为254 nm,用色谱峰面积外标法定量。乙二胺四乙酸二钠的质量浓度在0.5～50μg/mL范围内与色谱峰面积线性关系良好,相关系数为0.999 8,方法检出限为1.5mg/kg,定量限为5.0 mg/kg。冷面和米线样品加标回收率为90.7%～103.5%,测定结果的相对标准偏差为1.2%～5.0%(n=6)。该方法操作简单快速。     

Analysis of derivatized and underivatized theanine enantiomers by high-performance liquid chromatography/atmospheric pressure ionization-mass spectrometry

Theanine, a naturally occurring non-proteinic amino acid found in tea leaves, has demonstrated wide-ranging physiological activity, from lowering blood pressure to enhancing the anti-tumor activity of chemotherapeutic drugs. The chiral nature of theanine suggests that enantiospecificity plays a significant role in its various pharmacological functions. Using the Chirobiotic T (teicoplanin) chiral stationary phase, native and derivatized theanine enantiomers were separated and detected via high-performance liquid chromatography (HPLC) coupled to atmospheric pressure ionization mass spectrometry (API-MS). With the use of flow rates compatible with each ionization source, native theanine standards achieved excellent sensitivity and detection limits (10 ng/mL) for both atmospheric pressure chemical ionization (APCI) and electrospray ionization (ESI). Optimum sensitivity and detection limits for derivatized theanine standards were achieved using ESI-MS. The enantiomeric composition of six commercially available L-theanine samples was evaluated using the high-flow APCI-MS method and confirmed with photodiode array detection. Five of the six products contained significant amounts of D-theanine. Only one product, SunTheanine, appeared to contain only the L-theanine enantiomer.     

Isotachophoretic determination of theanine

Free amino acids, the key quality components of tea, are contained in higher amounts in higher grade teas. Among free amino acids in tea, the content of theanine, which is the amino acid of the highest amount, shows high correlation to the price of green tea. A capillary isotachophoresis (CITP) method was developed for the analysis of L-theanine in tea and food supplements. The optimized electrolyte system was following: 0.01 mol/l HCl + 0.02 mol/l TRIS + 0.05% HEC (leading electrolyte), 0.01 mol/L-valine + barium hydroxide to pH 10 (terminating electrolyte). Good separation of L-theanine from other components of sample was achieved within 20 min. Method characteristics, i.e., linearity (0–200 mg/l), accuracy (99 ± 2 %), intra-assay (1.5 %), quantification limit (2 mg/l), and detection limit (0.7 mg/l) were determined. Sufficient sensitivity, low labouriousness (extraction only) and low running cost are important attributes of this method. It was proved that the developed method is suitable for the routine analysis of L-theanine in green tea and food supplements containing green tea extract.     

柱前衍生高效液相色谱法测定茶叶中茶氨酸及γ-氨基丁酸含量

对茶叶中的茶氨酸和γ-氨基丁酸(GABA)建立了柱前衍生方法,并利用高效液相色谱法(HPLC)对衍生物进行了测定。采用邻苯二甲醛(OPA)及N-乙酰-L-半胱氨酸(NAC)为衍生试剂,考察了脯氨酸(Pro)、茶多酚(Tp)及维生素C(Vc)等对衍生体系的影响,并对色谱分离条件进行了优化。结果表明,茶氨酸与GABA衍生物在色谱柱上的保留行为与分离效率主要受流动相pH及缓冲盐浓度的影响;Pro、Tp及Vc对茶氨酸的衍生产率影响较小。而Vc可以提高GABA检测的灵敏度,并能稳定衍生用储备液。GABA与茶氨酸衍生后的方法检出限(LOD)分别为3.01×10~5mmol/L和7.98×10~5mmol/L;定量限(LOQ)分别为9.99×10~5mmol/L和2.658×10~4mmol/L;线性范围分别为0.01～0.4 mmol/L(相关系数为0.996)和0.05～0.8 mmol/L(相关系数为0.995);方法的回收率分别为99.29%～119.60%和93.18%～141.06%(除本身含茶氨酸量高的绿茶中回收率为62.88%外)。这说明经优化的柱前衍生-高效液相色谱体系可用于茶叶中两种特征性氨基酸的测定。     

Liquid chromatography determination of citalopram enantiomers using beta-cyclodextrin as a chiral mobile phase additive

A reliable and specific method for the determination of citalopram enantiomers was developed and validated. Chromatographic resolution of citalopram enantiomers was made on a Shim-pack (5 microm particle size) cyanopropyl column with beta-cyclodextrin (beta-CD) as an effective chiral mobile phase additive. The composition of the mobile phase was (90 + 10, v/v) aqueous 0.1% triethylammonium acetate buffer, pH 4.0 (adjusted with acetic acid), and acetonitrile, containing 12 mM beta-CD. The flow rate was 0.8 mL/min with ultraviolet detection at 240 nm. The effects of the mobile phase composition, concentration of beta-CD, and pH of the triethylammonium acetate buffer on peak shape and resolution of the enantiomers were investigated. The calibration graphs were linear (r = 0.9999, n = 8) in the range of 1-40 microg/mL for S(+) citalopram and R-(-) citalopram. The limit of detection values were 5.51 x 10(-3) and 4.35 x 10(-3) pg/mL, while the limit of quantification values were found to be 1.84 x 10(-2) and 1.45 x 10(-2) microg/mL for S-(+) citalopram and R-(-) citalopram, respectively.     

高效液相色谱-质谱法测定比格犬血浆中的艾普拉唑及其药代动力学

运用高效液相色谱-电喷雾质谱(HPLC-ESI-MS)技术,建立了快速、简单、灵敏的比格犬静脉滴注艾普拉唑钠盐后血药浓度的检测方法。血浆样品采用蛋白沉淀法,以丁螺环酮作为内标,色谱柱为Teknokroma Kromasil C18(100 mm×2.1 mm, 5 μm),流动相为水-甲醇-乙腈(69:8:23, v/v/v)(含0.1%的甲酸),流速0.2 mL/min,采用电喷雾(ESI)离子源以正离子方式检测。绘制血药浓度-时间曲线,并采用DAS 2.0计算药代动力学参数。方法学实验结果表明内源性杂质不干扰艾普拉唑和内标的测定,线性范围为5～10000 μg/L (r=0.994),最低定量限为5 μg/L,精密度和准确度均符合生物样品测定的要求。低、中、高3个浓度的绝对回收率在106%左右,基质效应小于142.0%,表明该方法适合比格犬血浆中艾普拉唑浓度的测定及药代动力学研究。比格犬静脉滴注艾普拉唑钠盐3个剂量(0.2 mg/kg、0.8 mg/kg和3.2 mg/kg)后的药-时曲线下面积(AUC(0~∞))分别为(2.4×104±3×103)、(8.8×104±1.6×104)和(5.4×105±8×104) μg/L•min,呈线性药物代谢动力学过程。     

Bupropion hydrochloride: the development of a chiral separation using an ovomucoid column

The separation of bupropion enantiomers on an ovomucoid stationary phase was investigated. The mobile- and stationary-phase parameters that may influence the separation were identified. The parameters that were studied include: type and concentration of organic modifier, mobile phase pH, ionic strength, type of buffer, and column temperature, as well as the effect that the amount of sample injected had on the separation. The optimized chiral separation baseline-resolved the enantiomers in less than 10 min. Calibration curves for a standard were linear over a range of 0.27-53.0 microg/g (ppm) with a correlation coefficient of 0.999 for both enantiomers. A detection limit of 0.13 microg/g and a quantitation limit of 0.27 microg/g were also found. The system precision of the method was 0.2%.     

On-line preconcentration and chiral separation of propiconazole by cyclodextrin-modified micellar electrokinetic chromatography

A method for the chiral separation of propiconazole using cyclodextrin-modified micellar electrokinetic chromatography (CD-MEKC) with hydroxypropyl-gamma-cyclodextrin (HP-gamma-CD) as chiral selector is reported. The use of a mixture of 30 mM HP-gamma-CD, 50mM SDS, methanol-acetonitrile 10%:5% (v/v) in 25 mM phosphate buffer solution was able to separate two enantiomeric pairs of propiconazole. Stacking- and sweeping-CD-MEKC under neutral pH (pH 7) and under acidic condition (pH 3.0) were used as two on-line preconcentration methods to increase detection sensitivity of propiconazole. Good repeatabilities in the migration time, peak area and peak height were obtained in terms of relative standard deviation (RSD). A sensitivity enhancement factor of 100-fold was achieved using sweeping-CD-MEKC at acidic pH. This is the first report on the separation of two pairs of propiconazole enantiomers and all the enantiomers of fenbuconazole and tebuconazole using sweeping-CD-MEKC. The limit of detection (S/N=3) for the three triazole fungicides ranged from 0.09 to 0.1 microg/mL, which is well below the maximum residue limits (MRL) set by Codex Alimentarius Commission (CAC). Combination of solid-phase extraction (SPE) pretreatment and sweeping-CD-MEKC procedure was applied to the determination of selected triazole fungicides in grapes samples spiked at concentration 10-40 times lower than the MRL established by the CAC. The average recoveries of the selected fungicides in spiked grapes samples were good, ranging from 73% to 109% with RSD of 9-12% (n=3).     

Electrophoretic separation of tryptophan enantiomers in biological samples.

A method for the determination of D- and L-tryptophan (Trp) in biological samples is described. The amino acid enantiomers were precolumn-derivatized with a fluorescence tagging reagent, naphthalene-2,3-dialdehyde (NDA). In the presence of hydroxypropyl-gamma-cyclodextrin (HP-gamma-CD) as the chiral selector, NDA-tagged Trp enantiomers were well resolved by micellar electrokinetic chromatography (MEKC). Using laser induced fluorescence (LIF) detection, a detection limit of 3.3 x 10(-8) M Trp was obtained. The method was applied to the determination of Trp enantiomers in biological samples including human urine and cerebrospinal fluid (CSF), rat brain tissue, and Aplysia ganglia. No interference from other amino acids or the endogenous compounds in the sample matrices was observed. D-Trp was found at the sub-microM level in human urine samples collected from several healthy subjects. Further, the determination of DL-Trp residues in small quantities (10 microg) of peptides after acid hydrolysis is demonstrated.     

Simultaneous chiral analysis of methamphetamine and related compounds by capillary electrophoresis/mass spectrometry using anionic cyclodextrin.

A capillary electrophoresis/mass spectrometry method for the simultaneous chiral analysis of enantiomers of methamphetamine (MA), amphetamine (AP), dimethylamphetamine (DMA), ephedrine (EP), norephedrine (NE) and methylephedrine (ME) in urine has been developed. The background electrolyte was 1 M formic acid (pH 1.7). Using 0.85 mM heptakis(2,6-diacethyl-6-sulfato)-beta-cyclodextrin as the chiral selector, the 12 enantiomers were completely separated within 25 min. The detection limits were 0.01 microg mL(-1) for the enantiomers of MA, AP, DMA, EP and ME, and 0.02 microg mL(-1) for the enantiomers of NE using selected ion monitoring. The reproducibilities of within-run (n = 4) for the migration times and peak areas of the standard mixture were under 0.58% and 7.83%, respectively. The calibration curves of the peak areas of the 12 enantiomers were linear in the range of 0.05 - 10 microg mL(-1). This method was applicable to the analysis of urine samples.     

Enantiomeric determination of azole antifungals in wastewater and sludge by liquid chromatography–tandem mass spectrometry

A sensitive and reliable liquid chromatographic-tandem mass spectrometric method for enantiomeric determination of five chiral azole antifungals (econazole, ketoconazole, miconazole, tebuconazole, and propiconazole) in wastewater and sludge has been established and validated. An isotope-labeled internal standard was used for quantification. Recovery of the individual enantiomers was usually in the range of 77-102 % for wastewater and 71-95 % for sludge, with relative standard deviations within 20 %. No significant difference (p>0.05) was observed between recovery of pairs of enantiomers of the chiral azole antifungals except for those of tebuconazole. Method quantification limits for individual enantiomers were 0.3-10 ng L(-1) and 3-29 ng g(-1) dry weight for wastewater and sludge, respectively. The method was used to investigate the enantiomeric composition of the azole pharmaceuticals in wastewater and sludge samples from a sewage treatment plant in China. Enantiomers of miconazole, ketoconazole, and econazole were widely detected. The results showed that the azole antifungals in wastewater and sludge were generally racemic or marginally non-racemic. The method is a useful tool for investigation of the enantiomeric occurrence, behavior, and fate of the chiral azole antifungals in the environment.