Determination of potentially hallucinogenic N-dimethylated indoleamines in human urine by HPLC/ESI-MS-MS.

A new method for the determination of N,N-dimethyl-5-hydroxytryptamine (bufotenine), N,N-dimethyltryptamine (DMT)*, 5-methoxy-N,N-dimethyltryptamine (5-MeODMT), and N-methyltryptamine (NMT) was developed using high-performance liquid chromatography-mass spectrometry (HPLC-MS). Identification of the analytes is based on liquid chromatographic retention times of analytes and two fragment ions produced by a triple quadrupole mass spectrometer. Quantification is based on electrospray ionization (ESI), and multiple reaction monitoring (MRM) was also utilized for getting better selectivity. The analytes and internal standard were separated from the urine matrix by solid-phase extraction (SPE). The method was applied for the determination of these compounds in urine samples of patients from surgical, medical and psychiatric wards. Of the dimethylated amines, only bufotenine was found in significant amounts (up to 34 microg/L). In keeping with our earlier results, the bufotenine excretion of psychiatric patients was found to be higher than that of the somatic patients. Method, procedure, considerations, statistical evaluations and urine sample spectra are presented.     

HPLC法测定刺五加中刺五加苷B的含量

目的:对不同生长年限、不同药用部位的刺五加中刺五加苷B进行了含量测定,为合理开发、利用这一药用植物资源提供了理论依据。方法:采用高效液相色谱法,研究了不同生长年限、不同药用部位的刺五加中的刺五加苷B含量的变化规律。结果:3年生的刺五加中刺五加苷B的含量高于1年生的剌五加,且均以茎皮中含量较高,茎中含量次之,根中最低。结论:对刺五加药材的合理采收提供了参考依据。     

离子对-反相高效液相色谱法测定人血清中美托洛尔的含量

目的离子对-反相高效液相色谱(HPLC)法测定人血清中美托洛尔浓度的评价。方法血清样品经蛋白沉淀剂(N,N-二甲基甲酰胺)沉淀后,高速离心,取上清液进行分析。测定使用C18色谱柱,甲醇-水(50:50,水相中含0.14%磷酸二氢钾)为流动相,以VP-16为内标,荧光检测Ex=277nm,Em=299nm。结果美托洛尔的浓度检测范围是2～1000pg/ml,r=0.9988;样品平均回收率为94.15%,相对标准差RSD为3.62%(n=6)。结论本方法选择性强,灵敏,重现性好。     

HPLC 法测定间歇性高容量血液滤过患者血清、尿液及置换液中莫西沙星浓度

目的:建立高效液相色谱法(HPLC)测定间歇性高容量血液滤过(PHVHF)患者血清、尿液及置换液中莫西沙星浓度.方法:PHVHF联合莫西沙星治疗肾衰合并感染患者8例,测定肾功、电解质.采用依利特SinoChrom ODS-BP色谱柱(5 μm,4.6 mm × 150 mm)测定莫西沙星浓度,流动相为乙腈:(10 mmol/L磷酸二氢钾、1.5 mmol/L四丁基硫酸氢胺) ＝ 25:75(v/v),柱温 40℃,检测波长 290 nm,流速 1 mL/min.结果:血BUN 及SCr、血清K+ 水平显著下降(P ＜ 0.05).血清莫西沙星在0.1 ～ 10.0 mg/L范围的线性回归方程为:y ＝ 4.49x-0.257,r ＝ 0.999 86.准确度为98.7% ～ 101%,日内和日间RSD为1.12% ～ 5.7%.置换液在0.1 ～ 10.0 mg/L范围的线性回归方程为:y ＝ 3.877x-0.249,r ＝ 0.998 5.准确度为85.7% ～ 95.7%,日内和日间RSD 为1.34% ～ 8.9%.尿液在0.1 ～ 10.0 mg/L范围的线性回归方程为:y ＝ 3.565x-0.270,r ＝ 0.986 8.准确度为97.4% ～ 101.7%,日内和日间RSD为1.51% ～ 11.6%.结论:HPLC方法检测血滤患者血清、尿液及置换液中莫西沙星浓度,方法简便、快速、重复性好,可满足临床研究血滤时莫西沙星药代动力学变化的要求.     

高效液相色谱法测定氢醌脂质体的氢醌含量与包封率

目的为减轻氢醌的刺激性和毒性,增加药物在表皮内的浓度和滞留时间,制备氢醌脂质体并了解其氢醌含量与包封率。方法采用旋转蒸发超声法制备氢醌脂质体,高效液相色谱(HPLC)法检测氢醌脂质体含量及包封率。结果色谱条件下氢醌脂质体与辅料及溶剂峰分离良好,且线性关系良好。3组氢醌脂质体包封率分别为80.2%、85.2%、81.7%。结论氢醌脂质体质量控制方法简单、可靠、准确。     

Multi-analyte procedures for screening for and quantification of drugs in blood, plasma, or serum by liquid chromatography-single stage or tandem mass spectrometry (LC-MS or LC-MS/MS) relevant to clinical and forensic toxicology

This paper reviews multi-analyte procedures for screening and quantification of drugs in blood, plasma, or serum using liquid chromatography coupled with a single stage or tandem mass spectrometer (LC-MS, LC-MS/MS). These procedures are relevant tools in clinical and forensic toxicology, and cover analysis of amphetamines, cocaine, hallucinogens, opioids, anesthetics, hypnotics, benzodiazepines, antidepressants, neuroleptics, antihistamines, sulfonylurea-type antidiabetics, beta-blockers, and other cardiac drugs. Basic information on the procedures is given in two tables and multi-analyte screening, identification, and quantification are illustrated in three figures. A critical discussion on the pros and cons of such LC-MS procedures is also included.     

用高效液相色谱法测定尿和血清多胺的改进及应用

本文建立了人尿及血清多肢的固相草取和高效液相色谱测定法。回收率：尿99．35～101．3％，血清：92．11～94．36％，日内和田间变异系数：1．5～4．2％，检测限：0．1μmol／L。探讨了常用抗癌药物对测定的干扰和此法在临床癌症病人多胶测定及疗效评价中的应用。本法简便、准确、重复性好、效率高，适宜临床检测。     

应用反相高压液相色谱内标法测定血清和尿中肌酐

目的评价以5-氟尿嘧啶（5-Fluorouracil，5-Fu）作内标的反相高压液相色谱（highpressure liquid chromatography，HPLC）测定血清和尿中肌酐（ereatinine）的方法，拟初步推荐其作为肌酐测定的候选参考方法，同时对3种常规肌酐测定试剂进行评定。方法首先对以5-Fu作内标的反相HPLC测定血清和尿中肌酐的方法进行了方法学评价。进一步以该方法为对比方法，分别采用Jaffe动力学法、酶法、干化学法测定试剂对85份新鲜血清和94份新鲜尿液标本进行测定，对测定结果进行相关和偏差分析。结果以5-Fu作内标的反相HPLC，其线性范围达2210μmol／L，批内变异系数（n=5）〈2．5％，日间变异系数（n=10）〈4．5％，回收率96．3％～102．4％。血清肌酐含量〈150μmol／L时（n=53），HPLC与各种方法间的相关系数分别为0．9314、0．9421、0．9284。血清肌酐含量〉150μmol／L时（n=32），HPLC与各种方法间的相关系数分别为0．9877、0．9841、0．9934。结论拟推荐以5-Fu作内标的反相HPLC作为肌酐测定的候选参考方法；肌酐酶偶联肌氨酸氧化酶法可作为常规测定方法。     

HPLC法测定精浆L-肉毒碱的研究及其临床意义

目的 研究建立人精浆中L-肉毒碱HPLC测定方法,分析正常生育男性及弱精症不育患者精浆中L-肉毒碱水平差异及其临床意义.方法 精浆样品经乙腈沉淀蛋白、衍生化反应后,以Lichrospher SiO2为固定相,乙腈-柠檬酸缓冲液(内含三乙胺12 mmol/L;pH 5.0)为流动相进行色谱等度分离,260 nm波长下定量检测,并对30例正常生育男性和87例不育患者进行精浆中L-肉毒碱水平分析.结果 在所建立的分析条件下,L-肉毒碱衍生化产物的色谱保留时间约为13 min,峰形清晰对称,与样品中其余内源性物质分离完全,定量准确.L-肉毒碱在0～1 000 μmol/L浓度范围线性良好.样品测定批间(n=6)及平均批内(n=5)测定的相对标准偏差分别为1.36%和1.23%,回收率为91.6%～96.5%.对30名正常生育男性及87例不育患者精浆中L-肉毒碱含量测定结果显示:正常生育组(n=30)为(392.7±107.2) μmol/L;弱精不育组(n=29)为(270.0±83.9) μmol/L;少精不育组(n=19)为(187.9±43.9) μmol/L;弱、少精不育组(n=39)为(175.7±67.1) μmol/L.统计学分析表明正常生育组与各不育组之间的差异均有统计学意义(P＜0.01);各不育组组间差异无统计学意义(P＞0.05).结论 精浆L-肉毒碱水平与精液中精子活力、精子密度等参数以及生育状况密切相关.其含量测定可作为男性不育症检查时一项有用的生化指标,为临床诊治和进行相关男性生殖功能机制研究提供参考.     

HPLC法测定尿白蛋白方法的建立和初步临床应用

目的 建立HPLC法测定尿白蛋白(Alb)的方法并做初步临床应用.方法 采用Agilent 4.6 mm×250 mm、5 μm的C18反相柱,优化流动相并选择合适的检测波长,评价该法的各项性能指标;测定46例糖尿病患者尿Alb.结果标准品和尿液Alb均在13.1 min出峰,在1 820 mg/L范围内标准曲线的直线性非常好,r值达到0.999 7,生物检测限为4.2 mg/L;24.5 mg/L和546.9 mg/L浓度时的批内和批间变异系数(CV)分别为3.36%、4.12%和1.93%、1.97%;回收率为96.3%、98.2%和97.5%.在46例糖尿病患者尿中,HPLC法能检出25例(54.3%)的微量白蛋白尿,比免疫散射比浊法(12例,占26.1%)检出更多.结论 HPLC法比免疫法能检出更多的微量白蛋白尿,在目前尚无方便可靠的方法测定尿总自蛋白前,HPLC法可用于常规尿微量白蛋白的测定,也可用于有关糖尿病肾病等的科研用途.     

高效液相色谱法检测人血清游离和总肉毒碱水平

目的 研究人血清中游离和总肉毒碱HPLC测定方法,建立成年体检人群血清中游离和总肉毒碱水平参考值.方法 血清样品经乙腈沉淀蛋白、衍生化反应后,以Lichrospher SiO2为固定相,乙腈-柠檬酸-三乙胺为流动相进行色谱等度分离,260 nm波长下定量检测,并对347名成年体检人群进行血清中游离和总肉毒碱水平测定.结果 在所建立的分析条件下,肉毒碱衍生化产物的色谱保留时间约为10 min,峰形清晰对称,与样品中其余内源性物质分离完全,定量准确.肉毒碱在0～400 μmol/L浓度范围线性良好.血清中游离肉毒碱和总肉毒碱批间(n=7)及平均批内(n=5)测定的相对标准偏差分别为3.04%、3.36%和1.77%、1.97%,测定平均回收率分别为98.2%和96.3%.对347名成年体检人群血清中肉毒碱水平测定结果显示:男182名总肉毒碱(52.2±8.6)μmol/L,游离肉毒碱(42.3±8.3)μmol/L,酯酰肉毒碱(9.9±2.9)μmol/L;女165名,总肉毒碱(48.2±9.9)μmol/L,游离肉毒碱(37.9±8.7)μmol/L,酯酰肉毒碱(10.3±3.5)μmol/L.统计学分析显示,男性血清中游离肉毒碱及总肉毒碱水平与女性组相比明显偏高,差异具有统计学意义(t=4.88、3.98,P<0.01).两组间酯酰肉毒碱浓度相比,差异无统计学意义(t=-1.32,P>0.05).结论 HPLC方法可同时检测血清游离和总肉毒碱含量,且灵敏度、特异性、重复性好,为有关肉毒碱在临床的合理应用及相关疾病的研究建立了有效的参考指标和检测方法.     

四种钙离子拮抗剂的高效液相色谱测定

介绍采用ＢｏｎｄＥｌｕｔＣ１８小柱提取以反相高效液相色谱法同时测定尼弗地平、尼卡地平、尼群地平和尼莫地平４种钙离子拮抗剂的方法。应用Ｗａｔｅｒｓ高效液相色谱仪，Ｃ１８分析柱及乙腈／ＫＨ＿２ＰＯ＿４＝４６／５４流动相，在１．５ｍ１／ｍｉｎ流速下分离上述４种药物。回收率分别为１０１％～１０９％，９５％～１０１％，９９％～１０４％和１０３％～１０８％，批内变异系数（ＣＶ）＜７．４％，批间ＣＶ＜８．７％。本方法特异性好，操作简便、迅速，适合于临床和药代动力学研究应用。     

HPLC法在小儿尿液新蝶呤和生物蝶呤检测中的应用

目的 建立HPLC法测定小儿高苯丙氨酸血症(HPA)患者尿液中新蝶呤(N)和生物蝶呤(B)含量的方法,以帮助临床对经典型苯丙酮尿症(PKU)和四氢生物蝶呤(BH4)缺乏症的诊断和鉴别诊断.方法 选用Cosmosil 5C18-AR-Ⅱ色谱柱(4.6 I.D.×250 mm),甲醇-水(8∶92)为流动相,用三维荧光检测器检测.结果 新蝶呤线性范围为1.60～40.0 ng/μl,生物蝶呤线性范围为1.73～43.2 ng/μl;最小检测限为0.16 ng/μl和0.1728 ng/μl.样品测定的变异系数(CV)值分别为2.18%和1.70%;原尿样品前处理时氧化的最佳时间为30 min;原尿样品不便于保存,氧化后可于室温或4℃冰箱保存3 d.结论 本方法准确、可靠、简便易行,灵敏度高,方法学实验结果满意,适用于临床HPA患儿的鉴别诊断.     

Determination of ciprofloxacin concentrations in human serum and urine by HPLC with ultraviolet and fluorescence detection

OBJECTIVE: To develop a simple and sensitive high performance liquid chromatography method for the determination of ciprofloxacin concentrations in human serum and urine. METHOD: Serum proteins were removed by ultrafiltration through a filtering device after the addition of a displacing reagent. Urine samples were diluted with mobile phase prior to injection. Separation was achieved with a C18 reverse-phase column and using ultraviolet (UVD) and fluorescence detection (FD) for serum samples and UVD for urine samples. RESULTS: The quantitation limits of the assay were 20 ng/ml (FD) and 100 ng/ml (UVD) in serum and 1 microg/ml in urine. The assay was successfully applied to a pharmacokinetic study of ciprofloxacin in healthy volunteers. CONCLUSION: The method presented for ciprofloxacin assay in human serum and urine requires less sample clean up and is more sensitive than those reported in the literature.     

Determination of sinomenine in human plasma by HPLC/ESI/ion trap mass spectrum

BACKGROUND: Sinomenine is a pure alkaloid with a variety of pharmacological actions including anti-inflammation, immuno-suppression, analgesia, antihypertension, and antiarrhythmia. Methods have been developed to measure sinomenine in rats and rabbits through high-performance liquid chromatography (HPLC). But only one report described a method for determination of sinomenine in humans. METHOD: We developed a method to quantitate sinomenine in human plasma based on liquid chromatography-ion trap mass spectrometry coupled with electrospray ionization (HPLC/ESI/ion trap mass spectrum). RESULTS: The calibration curve (r2=0.9993) of sinomenine was established with standard solutions. The mean inter-day and intra-day CVs<15%. The limit of detection for sinomenine was 0.5 ng/ml. The relative recovery of high, middle and low concentration was 99%, 103%, and 118%, respectively. CONCLUSION: The method is simple, rapid, sensitive, specific, and accurate enough for determination of sinomenine in human plasma and may be used to investigate the metabolism and pharmacokinetics of sinomenine.     

高效液相色谱法测定人血浆中盐酸替扎尼定含量

目的建立用高效液相色谱法快速检测盐酸替扎尼定血药浓度的方法。方法取血浆加1 mol/L NaOH,混匀,加氯仿提取吹干,用流动相复溶进样;色谱柱SUNTEK Krom asiL C185μm粒径,150×4.6 mm;色谱条件:A∶乙睛=100∶25[A:水-甲酸-氨水(28%)=100∶5∶10(V/V,pH 8.50)];流速1.0 m l/m in;紫外检测波长318 nm,内标物为雷尼替丁。结果盐酸替扎尼定的保留时间为6.9 m in,雷尼替丁的保留时间为8.2 m in,检测的线性范围0.3867~49.5000 ng/m l(r=0.9982)方法回收率大于90.78%(n=5)日内RSD=7.74%,日间RSD=3.79%(n=5)。结论本方法简便快速,定量准确,适用于盐酸替扎尼定人体临床药代动力学和生物利用度的研究。     

反相高效液相色谱法测定人血浆及尿样中普卢利沙星活性代谢物

目的建立分析人血浆及尿样中普卢利沙星活性代谢物浓度的反相高效液相色谱方法。方法样本用甲醇-乙腈(1 :1)沉淀后离心取上清液进行色谱分析。分析柱为Diamonsil C_(18)(250 mm×4．6 mm,5μm)。流动相为乙腈—0．05 mol/L磷酸二氢钾(pH2．2,含1%的四丁基溴化铵);血浆和尿样分析分别用20:80(V/V)、流速1．0 mL/min和12:88(V/V)、流速1．6 mL/mln。激发和发射波长分别为280 nm和425 nm。结果血浆和尿样测定的线性范围分别为0．005～5 mg/L(r= 0．9999)和0．05～5 mg/L(r=0．9999),最低检测浓度分别为0．002 mg/L和0．01mg/L。血浆5．00、0．50和0．05 mg/L浓度的相对回收率在100．64%～101．00%,日内和日间相对标准差(RSD)分别小于2．5%和4．6%。尿样2．50、0．50和0．10 mg/ L浓度的相对回收率在97．20%～100．20%,日内和日间RSD分别小于1．3%和4．3%。方法成功用于健康人口服普卢利沙星的药动学研究。结论本方法具有简便、快速、灵敏度高、重现性好等特点,适用于普卢利沙星的药动学研究。     

Determination of beta 2-microglobulin in human urine and serum by latex immunoassay

This highly sensitive method for determination of beta 2-microglobulin (beta 2-m) in human urine or serum is based on direct agglutination by beta 2-m of latex particles on which an antibody against beta 2-m is adsorbed. The agglutination is quantified by counting the remaining unagglutinated particles, or by turbidimetry. A novel aspect of this method is the capability to prevent nonspecific agglutination of the antibody-coated particles by diluting them with an albumin solution of well-defined characteristics (pH, freshness, concentration) just before the assay. The assayable concentration range is 1--32 micrograms/L, the detection limit 0.5 micrograms/L. Within-assay CV, based on 10 determinations of beta 2-m in urine and serum at two different dilutions, ranged from 4.6 to 8.7%. Between-assay CV, calculated from 10 determinations of beta 2-m in urine and serum, was 10 and 8.4%, respectively. Analytical recovery of beta 2-m in urine averaged 97% and in serum 104% (n = 10). No component of urine or serum interfered. Coefficients of correlation for beta 2-m in urine or serum as measured by radioimmunoassay and latex immunoassay were 0.97 and 0.93, respectively. Concentrations of beta 2-m in serum and urine from 33 healthy men (ages 20 to 67 years) averaged 1.5 mg/l and 54 micrograms/g of creatine, respectively.     

Selective screening for inborn errors of metabolism and secondary methylmalonic aciduria in pregnancy at high risk district of neural tube defects: a human metabolome study by GC-MS in China

ObjectiveUrease pretreatment-gas chromatography-mass spectrometry (UP-GC-MS) has become a valuable tool in the field of metabolome research, including analysis of inborn errors of metabolism (IEMs) and acquired metabolic disturbances secondary to nutrition or drugs. This research aims to screen IEMs in Chinese patients and to explore the cause of neural tube defects (NTDs), a congenital malformation very common in North China.Design and methodsUrine samples from 618 patients at high risk of IEMs in China were collected, and UP-GC-MS was performed in the selective screening. Urinary methylmalonate (MMA) levels in pregnancy with and without NTDs fetus, respectively, at Luliang district, a countryside region with NTDs incidence 227/10 000, Shanxi Province, North China, were analyzed by GC-MS-selective ion monitoring, and compared with that from control region.ResultsAmong the 618 patients, 22 kinds and 59 cases of IEM were found. Methylmalonic aciduria (MMAuria) is on top of the list, followed by neonatal intrahepatic cholestasis caused by citrin deficiency (NICCD), phenylketonuria (PKU), multiple carboxylase deficiency (MCD), etc. Satisfactory therapeutic effects have been achieved in patients such as NICCD, MCD, and galactosemia. At Luliang district, urinary MMA levels in pregnancy, no matter NTDs-affected or unaffected, are both significantly (P < 0.01) higher than that in normal control, while serum B₁₂ levels in NTDs-affected pregnancy are significantly lower than that both in NTDs-unaffected group (P < 0.01) and in normal control (P < 0.01). Furthermore, B₁₂ < 52.5 pmol/L is associated with a 7.78-fold increased NTDs risk (P < 0.01) at Luliang district.ConclusionsSelective screening for IEMs by UP-GC-MS provides valuable evidences for the diagnosis of IEMs. MMAuria secondary to B₁₂ deficiency is quite common at Luliang district, suggesting B₁₂ deficiency is involved in the development of NTDs in the specific population. This metabolome research by UP-GC-MS provides valuable epidemiological information that helps to understand the prevalence and the possible intervention strategy of NTDs and IEMs, especially in Chinese population.     

高效液相色谱——紫外、荧光双检测器同时测定人尿中犬尿氨酸和犬尿喹啉酸

目的建立高效液相色谱（HPLC）同时测定人尿中犬尿氨酸（KYN）和犬尿喹啉酸（KYNA）方法。用KYNA／KYN比值评价人体犬尿氨酸氨基转移酶（KAT）活性。方法采用HPLC测定KYN、KYNA,分析柱为安捷伦HCC18反相色谱柱;流动相为10mmol／L乙酸钠-乙酸缓冲液：7％乙腈=93：7;流速1mL／min;检测器波长：紫外365nm,荧光344nm（激发）／398nm（发射）。结果KYN和KYNA保留时间分别为6．9和11．5min。水溶液标准与标准加入工作曲线斜率差异无统计学意义（KYN：P=0．4068;KYNA：P=0．6462）。KYN和KYNA的平均回收率分别为98．74％±7．53％、95．17％±8．17％;稳定性试验的变异系数（CV）分别为3．35％和0．88％;精密度CV分别为2．73％和2．79％;最低检测限分别为0．4和0．2μmol／L。30份健康成年人尿样的KYNA／KYN比值为2．45±1．54。尿中KYNA与KYN含量呈正相关关系（r2=0．3758,P〈0．001）。结论测定方法灵敏、准确、稳定。KYNA／KYN比值评测KAT活性具有简便、实用等特点,可用于相关疾病的临床辅助诊断和基础研究。