毛细管电泳中移动反应界面法富集和检测尿样中的氧化苦参碱

采用建立在移动反应界面理论上的体系进行尿样中氧化苦参碱的富集与定量检测。与传统的毛细管电泳相比,体系中引入了富集缓冲溶液(富集相)和分离缓冲溶液(分离相)。优化的条件如下:样品缓冲溶液为20 mmol/L 甲酸钠(用氨水调节pH至10.70),富集缓冲溶液为40 mmol/L 甲酸-甲酸钠(pH 2.60),分离缓冲溶液为100 mmol/L 甲酸-甲酸钠(pH 4.80);样品相压力进样1.4 kPa×3 min,富集相压力进样1.4 kPa×7 min,紫外检测波长210 nm,电压21 kV。氧化苦参碱在2.2~65 mg/L的质量浓度范围内呈良好的线性关系(r=0.9991),检出限为0.74 mg/L,灵敏度比常规毛细管电泳方法提高约70倍,重现性良好。该方法已经成功地应用于尿样中氧化苦参碱的检测。     

高效毛细管电泳安培法测定肉苁蓉中松果菊苷的含量

采用毛细管电泳安培法建立了肉苁蓉中松果菊苷含量的检测方法,探讨了缓冲溶液种类、工作电位、分离电压、进样时间等对分离和检测的影响.在15 mmol/L硼砂缓冲溶液(pH 9.5),25 kV电压,0.6 V工作电位的条件下,松果菊苷的质量浓度在0.5～50.0 mg/L范围内与峰面积呈良好线性,相关系数为0.9994,检...     

Direct tris(2,2'-bipyridyl)ruthenium (II) electrochemiluminescence detection of polyamines separated by capillary electrophoresis

Capillary electrophoresis (CE) with tris(2,2'-bipyridyl) ruthenium (II) (Ru(bpy)3(2+)) electrochemiluminescence (ECL) detection technique was developed for the analysis of four polyamines (putrescine (Put), cadaverine (Cad), spermidine (Spd), and spermine (Spm)) analysis. The four polyamines contain different amine groups, which have different ECL activity. There are several parameters which influence the resolution and ECL peak intensities, including the buffer pH and concentrations, separation voltage, sample injection, electrode materials, and Ru(bpy)3(2+) concentrations. Polyamines are separated by capillary zone electrophoresis in an uncoated fused-silica capillary (50 cmx25 micro m (ID) filled with acidic phosphate buffer (200 mmol/L phosphate, pH 2.0) - 1mol/L phosphoric acid (9:1 v/v) and a separation voltage of 5 kV (25 micro A), with end-column Ru(bpy)3(2+) ECL detection. A 5 mmol/L Ru(bpy)3(2+) solution plus 200 mmol/L phosphate buffer (pH 11.0) is added into the reagent reservoir. The calibration curve is linear over a concentration range of two or three orders of magnitude for the polyamines. The analysis time is less than 25 min. Detection limits for Put and Cad are 1.9x10(-7) mol/L and 7.6x10(-9) mol/L for Spd and Spm, respectively. Intraday and interday relative standard deviations of ECL peak intensities are less than 8%. The main advantages of this CE-ECL detection technique for polyamines analysis presented herein are the omission of chemical derivatization of the analytes and the high selectivity.     

Quantification of polyamines in human erythrocytes using a new near-infrared cyanine 1-(epsilon-succinimidyl-hexanoate)-1'-methyl-3,3,3',3'-tetramethyl-indocarbocyanine-5,5'-disulfonate potassium with CE-LIF detection

A novel near-infrared (NIR) cyanine 1-(epsilon-succinimidyl-hexanoate)-1'-methyl-3,3,3',3'-tetramethyl-indocarbocyanine-5,5'-disulfonate potassium (MeCy5-OSu) has been developed in our laboratory. Simultaneous determination of MeCy5-OSu-derivatized polyamines spermine (Spm), spermidine (Spd), cadaverine (Cad), and putrescine (Put) based on the separation by CE combined with diode LIF detection has been accomplished. The highest derivatization efficiency was achieved in 0.2 mol/L borate buffer (pH 8.8) for 20 min at 25 degrees C. Polyamine derivatives were separated within 14 min in the phosphate running buffer (pH 3) containing 50 mmol/L phosphoric acid, 40 mmol/L SDS, and 35% methanol v/v. Linearity of response was obtained in the range of 10-200 nmol/L. The detection limits (S/N = 3) for Spm, Spd, Cad, and Put were 0.8, 1, 3, and 2 nmol/L, respectively. The proposed method has been successfully applied to the analysis of polyamines in erythrocytes of two healthy persons and one cancer patient. Average recoveries for erythrocyte samples were 93.6-106% and coefficients of variation ranged from 1.8 to 5.4%. The analysis of polyamines in erythrocytes can be used for studying the relationship between their changes and the carcinogenesis process involved in erythrocytes.     

毛细管胶束电动色谱法分离测定中药半枝莲中的7种有效成分

建立了毛细管胶束电动色谱同时分析检测中药半枝莲药材及其膏剂中黄芩素、柚皮素、汉黄芩素、野黄芩苷、芹菜素、木犀草素和原儿茶酸7种有效成分的方法。半枝莲样品中7种有效成分经甲醇超声提取。实验考察了运行缓冲溶液的pH值和浓度、添加剂、检测波长、分离电压和进样时间等重要参数对目标物分离的影响。得到的优化条件为: 运行缓冲液50 mmol/L硼砂-0.20 mol/L硼酸溶液(pH 8.4),含8.5 mmol/L十二烷基硫酸钠(SDS),分离电压25 kV,检测波长260 nm和335 nm。在此条件下,7种组分于12 min内达到基线分离。各组分在8×10~6～3.2×10~4mol/L范围内呈良好的线性关系,相关系数(r2)为0.9965～0.9999;检出限为7.0×10~8～2.0×10~6mol/L;回收率均大于85%。该方法提取简便、准确可靠、重复性好、灵敏度高,可以用于中药半枝莲中7种有效成分的定量检测。     

Urine polyamines determination using dansyl chloride derivatization in solid-phase extraction cartridges and HPLC

The derivatization of biogenic amines such as putrescine, cadaverine, spermidine and spermine with dansyl chloride in solid phase extraction cartridges is described. Different types of filling materials were tested in order to have the highest retention of the different analytes. The best results were obtained by using C18 cartridges. The optimal conditions were: amine solution buffered at pH 12, 2 mM dansyl chloride (acetone-bicarbonate solution 20 mM (pH 9-9.5), 2 + 3 v/v) as reagent concentration, room temperature and 30 min reaction time. The developed procedure was applied to the determination of these polyamines in urine samples from healthy controls and cancer patients using HPLC with 1,7-diaminoheptane as internal standard. The concentrations ranged from 0.5 to 5 micrograms mL-1 and the detection limits were 10 ng mL-1 for all polyamines. By concentrating the urine extracts, the detection limits were improved down to 2 ng mL-1. The accuracy and the precision of the method were tested. The proposed dansylation method is advantageous with respect to solution dansylation. It improves the total analysis time, avoids high temperatures that can affect the thermal stability of the derivatives and could make possible the automation of the procedure.     

毛细管电泳法高压快速分离分析菠菜中的水溶性维生素

在毛细管电泳法中,通过双模对接高压电源可以获得0～40 kV甚至40 kV以上的超高电压。本研究在40 kV的超高电压下,以纯电解质水溶液为缓冲液,实现了蔬菜中通常含有的8种水溶性维生素(VB1、VB2、VB6、VC、D-泛酸钙、D-生物素、烟酸和叶酸)的快速分离及菠菜样品的定量分析。通过考察电压、缓冲溶液浓度、pH值等因素对分离的影响,确定了优化的实验条件。结果表明,在40 kV高压下,采用25 mmol/L硼砂-硼酸溶液缓冲液(pH 8.8),菠菜中上述8种水溶性维生素在2.2 min内获得了较好的基线分离。用此方法对菠菜中的水溶性维生素进行定量分析,得到了令人满意的结果。水溶性维生素的线性相关系数范围为0.9981～0.9999,检出限为0.2～0.3 mg/L,在菠菜中的平均加标回收率为88.0%～100.6%,峰面积的相对标准偏差(RSD)为1.15%～4.13%。     

多肽在金纳米粒子表面偶联反应效率研究

通过1-(3-二甲氨基丙基)-3-乙基碳二亚胺(EDC)与N-羟基琥珀酰亚胺(NHS)活化反应将多肽偶联到金纳米粒子表面,采用荧光光谱研究其形成的酰胺键的反应偶联效率.考察了实验条件,包括缓冲液的种类(HEPES、Tris-HCl、硼酸、PBS缓冲液)、pH值(pH 6.5~9.0)、缓冲液浓度(10, 25, 40和50 mmol/L)、NHS和EDC的浓度(NHS 0.2~1.0 mol/L,EDC 0.01~0.5 mol/L)及二者比例(0,0.5,1.0,2.0和2.5)、偶联反应时间(4, 8, 12, 24和36 h)等对偶联效率的影响,筛选出最佳实验条件.实验结果表明, 25 mmol/L 4-羟乙基哌嗪乙磺酸(HEPES)缓冲液(pH 7.0), NHS/EDC浓度为0.4 mol/L/0.2 mol/L和24 h的反应时间为EDC-NHS活化反应将多肽偶联到金纳米粒子上的最佳实验条件.本研究结果可为相关研究提供技术参考.     

Determination of vitamin C and preservatives in beverages by conventional capillary electrophoresis and microchip electrophoresis with capacitively coupled contactless conductivity detection

The separation and detection of commonly used preservatives (benzoate, sorbate) and vitamin C by both conventional CE and microchip electrophoresis with capacitively coupled contactless conductivity detection is presented. The separation was optimized by adjusting the pH-value of the buffer and the use of hydroxypropyl-beta-CD (HP-beta-CD) and CTAB as additives. For conventional CE, optimal separation conditions were achieved in a histidine/tartrate buffer at pH 6.5, containing 0.025% HP-beta-CD and 0.1 mM CTAB. LOD ranged from 0.5 to 3 mg/L (S/N = 3) and the RSDs for migration time and peak area were less than 0.1 and 2%, respectively. A considerable reduction of analysis time can be accomplished by using microchip electrophoresis without significant loss in sensitivity under optimal separation conditions. A histidine/tartrate buffer at pH 6.5, incorporating 0.06% HP-beta-CD and 0.25 mM CTAB, gave detection limits ranging between 3 and 10 mg/L and satisfactory reproducibilities of < or =0.4% for the migration time and < or =3.5% for the peak area. The methods developed are useful for the quantitative determination of food additives in real samples such as soft drinks and vitamin C tablets.     

芯片毛细管电泳激光诱导荧光快速分离检测麻黄碱类兴奋剂

研究用芯片毛细管电泳激光诱导荧光检测系统分离测定经7-chloro-4-n itrobenzo-2-oax-1,3-d iazole(NBD-C l)衍生的麻黄碱和伪麻黄碱的实验条件。采用胶束毛细管电动色谱分离体系(12 mmol/L SDS 10mmol/L硼砂缓冲液,pH 9.0),在45 mm长的通道上实现了麻黄碱和伪麻黄碱的快速分离,一次分离小于1.5m in。10~100 mg/L范围内,峰高与浓度呈良好的线性关系,麻黄碱、伪麻黄碱的检出限分别是0.83 mg/L和1.10 mg/L。所建立的方法应用于尿中麻黄碱和伪麻黄碱的分离测定,取得满意的结果。     

Determination of Nicotine in Tobacco by Capillary Electrophoresis with Electrochemical Detection

A sensitive, simple and low-cost method based on capillary electrophoresis(CE) with electrochemical(EC) detection at a carbon fiber microdisk electrode(CFE) was developed for the determination of nicotine. Effects of detection potential, concentration and pH value of the phosphate buffer, and injection time as well as separation voltage were investigated. Under the optimized conditions: a detection potential of 1.20 V, 40 mmol/L phosphate buffer(pH 2.0), a sample injection time of 10 s at 10 kV and a separation voltage of 16 kV, the linear range obtained was from 5.0×10^-7 mol/L to 1.0×10^-4 mol/L with a correlation coefficient of 0.9989 and the limit of detection(LOD, S/N=3) obtained was 5.0×10^-8 mol/L. The method was also used to determine the nicotine in cigarettes. Nicotine amount ranged from 0.211 mg/g to 0.583 mg/g in the pipe tobacco of seven brands of cigarette and the amount in one cigarette varied from 0.136 mg/cigarette to 0.428 mg/cigarette.     

Determination of polyamines by flow injection analysis with a chemiluminescence detector based on their complexation with copper(II).

Through the flow injection analysis experiments, we discovered that an unsaturated complex of Cu(II) and polyamines (spermine, spermidine, putrescine) had a strongly catalytic effect on luminol-H(2)O(2) chemiluminescence (CL) reaction, and that the CL intensity is proportional to the concentrations of polyamines. Based on the automatic formation of an unsaturated complex of polyamines and Cu(II) when the solution containing polyamines passed through a column packed with solid Cu(OH)(2), a new flow injection chemiluminescence analysis method was proposed for the determination of polyamines. The effects of pH, buffer concentration, the concentration of chemiluminescence reagent, and the influence of mixing coil length were examined. Under optimal conditions, the linear range was from 1.0 x 10(-7) mol L(-1) to 1.0 x 10(-5) mol L(-1), and the detection limits were 0.17, 0.38, 0.44 pmol for spermine, spermidine, and putrescine, respectively. Compared with other methods, the advantages of this method include convenience, time-saving and low cost.     

毛细管电泳电致化学发光检测血浆中布比卡因

布比卡因是一种外科局部麻醉剂,使用过量会导致中枢神经系统和心脏血管系统中毒^[1],可引起心脏停博.高效液相色谱和毛细管电泳(CE)^[2]是该药常用的检测方法.     

Determination of catecholamines and serotonin by micellar electrokinetic chromatography with laser-induced fluorescence detection

6-Oxy-(N-succinimidyl acetate)-9-(2'-methoxycarbonyl) fluorescein, a new synthesized fluorescent reagent, was established for the first time as a label for the sensitive analysis of catecholamines (CAs) and serotonin (5-HT) by micellar electrokinetic chromatography (MEKC) with laser-induced fluorescence detection. After careful study on the derivatization conditions such as pH value, reagent concentration, temperature and reaction time, the labeling reaction was accomplished as quickly as 7 min with stable yield. The separation parameters for the CAs and 5-HT were also optimized in detail. The derivatives were baseline separated in a running buffer containing 30 mM boric acid and 15 mM sodium dodeculsulfate at pH 9.0. The detection limits ranged from 5 x 10(-10) to 2 x 10(-9) M (signal-to-noise ratio = 3). The rapid and sensitive method was also applied to the determination of the CAs and 5-HT of urine samples.     

液相微萃取/离子色谱测定牛奶中的氨

以水为微滴萃取溶剂,采用顶空液相微萃取/离子色谱检测了牛奶中的氨.优化了顶空液相微萃取的实验条件:pH=12,萃取温度为35 ℃,萃取时间为15 min,搅拌速率为800 r/min,萃取溶剂体积为5 μL.测定氨的线性范围为10 ～300 μg·L-1(R2=0.998),检出限达1.8 μg·L-1,回收率为92% ～105%.     

月桂酸胶束电动毛细管色谱快速测定氨基甲酸脂类农药的研究

以月桂酸表面活性剂和氨水组成缓冲溶液,建立了毛细管胶束电动色谱快速测定七种氨基甲酸脂类农药的方法。对电泳介质的种类及浓度、pH值、操作电压和进样时间等影响因素进行了优化,以45 mmol/L月桂酸(乙醇溶)和120 mmol/L氨水(用0.1 mol/L NaOH调至pH=9.5)为电泳介质,在4.5 min内实现对速灭威、残杀威、克百威、抗蚜威、异丙威、硫双威、双氧威的分离检测。结合固相萃取(SPE)对农田水样进行测定,各农药检出限为2.0～15μg/L,回收率为85%～107%,相对标准偏差为4.3%～8.3%。     

Determination of galanthamine in Bulbus Lycoridis Radiatae by coupling capillary electrophoresis with end‐column electrochemiluminescence detection

A novel method for the determination of galanthamine (GAL) in Bulbus Lycoridis Radiatae has been developed based on coupling CE with an end‐column tris(2,2′‐bipyridyl)ruthenium(II) electrochemiluminescence (ECL). Parameters affecting CE separation and ECL detection were investigated and optimized. Baseline separation of GAL from other components in the Bulbus Lycoridis Radiatae sample was achieved with an 18 mmol/L phosphate running buffer at pH 9.0. Under the optimized conditions: 12 kV CE‐separation voltage, ECL detection potential at 1.25 V with 5 mmol/L and 50 mmol/L phosphate buffer at pH 7.5 in the detection reservoir, the linear range of GAL concentration was from 0.8 ng/mL to 2 μg/mL, whereas the detection limit was 0.25 ng/mL (S/N=3). The proposed method was successfully demonstrated for the determination of GAL in Bulbus Lycoridis Radiatae.     

毛细管区带电泳法测定卷烟中的生物碱

在磷酸缓冲体系中采用毛细管区带电泳法测定卷烟中的生物碱时,检测灵敏度低,分离度差。考察了卷烟中生物碱的 提取条件,分离缓冲溶液的类型、pH值和浓度,卷烟中生物碱测定方法的线性范围、检出限、重现性和回收率。结果发 现,当采用410 mmol/L的酒石酸溶液（pH 2.8）为缓冲体系时,卷烟中生物碱的检测灵敏度和分离度均有明显改善,烟碱 的线性范围为0.06~0.80 mg/L（其他生物碱为0.006~0.10 mg/L）,检出限为0.002~0.01 mg/L,相对标准偏差为2.2%~10%,回收率为87.6%~102%。     

Analysis of dideoxyadenosine triphosphate by CE with fluorescence detection. I. Derivatization through the phosphate group

A CZE method was developed, which separates 2',3'-dideoxyadenosine-5'-triphosphate (ddATP) from other metabolites and endogenous nucleotides at high concentrations (20-200 microg/mL) to allow UV detection. To enhance sensitivity, fluorescence detection which requires prior derivatization of compounds was examined. Precapillary derivatization of ddATP in the presence of N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (EDAC) with dansyl ethylenediamine (dansyl EDA) was faster and stable compared to that of 4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacene-3-propionyl ethylenediamine (BODIPY FL EDA). Reaction conditions, reagent concentrations and detection parameters were optimized and highest derivatization efficiency was achieved in 0.1 M 1-methylimidazole buffer (pH 8.0) with 140 mM EDAC in 1-methylimidazole buffer and 30 mM dansyl EDA in DMF for 90 min at 60 degrees C. Dansyl EDA derivatives of ddATP, 2'-deoxyadenosine-5'-triphosphate (dATP) and ATP were comigrating with the CZE method; therefore, a MEKC method was developed and optimized for repeatable separations. Upon dansylation, sensitivity of ddATP with fluorescence detection (LOQ = 12 ng/mL) was 160 times higher than UV detection (LOQ = 1.9 microg/mL).     

高效液相色谱法测定化妆品中氯噻酮和吩噻嗪

建立了反相高效液相色谱法测定化妆品中氯噻酮和吩噻嗪的分析方法。化妆品中氯噻酮和吩噻嗪用丙酮超声提取,采用Kromasil C18色谱柱(5μm,250 mm×4.6 mm i.d.),以30 mmol/L NaH2PO4缓冲液(pH 5.6)和甲醇为流动相进行梯度洗脱,得到了理想的分离效果。氯噻酮和吩噻嗪在0.2~100 mg/L范围内,质量浓度与其峰面积呈良好的线性关系。在低、中、高3个添加水平下,氯噻酮的平均回收率为94.8%~104.1%,RSD为0.7%~4.4%,吩噻嗪的平均回收率为92.5%~103.1%,RSD为1.0%~6.9%。方法可用于化妆品中氯噻酮和吩噻嗪的检测。