双氟沙星及其代谢产物在中华绒螯蟹体内药物代谢及残留消除规律

采用高效液相色谱法,在口灌给药途径下,研究双氟沙星及其代谢产物沙拉沙星在中华绒螯蟹(Eriocheir sinensis)体内的药代动力学.中华绒螯蟹以20 mg/kg剂量给药双氟沙星后,其血淋巴、肌肉、肝胰腺、精巢和卵巢中药物-时间曲线关系符合开放性二室模型.双氟沙星在中华绒螯蟹体内吸收迅速,在不同组织中分布较广,达峰时间短.血淋巴、肌肉、肝胰腺、精巢和卵巢中的Vd分别为3.170 L/kg、2.122 L/kg、1.045 L/kg、1.051 L/kg和0.203 L/kg;双氟沙星在中华绒螯蟹体内消除缓慢,在血淋巴、肌肉、肝胰腺、精巢和卵巢中的消除半衰期t1/2β分别为96.316 h、88.228 h、137.524 h、67.021 h和124.679 h;总体清除率CLa分别为0.783 L/h·kg、0.040 L/h·kg、0.013 L/h·kg、0.011 L/h·kg和0.008 L/h·kg.代谢产物沙拉沙星在中华绒螯蟹血淋巴、肌肉、肝胰腺、精巢和卵巢中药物水平的变化趋势与双氟沙星相似,都呈现多峰现象.但肌肉、肝胰腺、精巢和卵巢4种组织中代谢产物沙拉沙星出现药峰的时候恰好是这4种组织中双氟沙星下降缓慢时期.鉴于双氟沙星及其代谢产物沙拉沙星在中华绒螯蟹体内消除缓慢,而可食组织中脂肪含量较高,因此若规定可食组织中双氟沙星的最大残留限量以脂肪中最大残留限量(100μg/kg)为标准,沙拉沙星的最大残留限量为30μg/kg,则建议双氟沙星在中华绒螯蟹中的休药期大于24d,才能保障食用者的安全.     

盐酸氯苯胍在斑点叉尾鮰体内的药代动力学及残留消除规律

在水温(28±1)℃条件下, 以20 mg/kg剂量的盐酸氯苯胍口灌斑点叉尾鮰(Ictalurus punctatus), 采用高效液相色谱-串联质谱法研究盐酸氯苯胍在斑点叉尾鮰体内的药代动力学和残留消除规律。结果显示, 在单次口灌给药后, 盐酸氯苯胍在斑点叉尾鮰血浆中的药时曲线符合二室模型特征, 其药动学方程为C= 7.69e–0.02t+ 0.13e–0.01t–7.82e–0.27t。盐酸氯苯胍在斑点叉尾鮰血浆、肌肉、皮、鳃、肝脏和肾脏中的T_(peak)分别为10.03、15.79、11.10、2.61、12.89、7.87h; C_max分别为5.76 μg/mL、2.91、2.90、3.05、3.04、0.42 mg/kg, 消除半衰期分别为58.63、23.57、35.37、19.74、29.34、43.30h; 药-时曲线下面积AUC分别为326.74 (μg/mL)/h、157.58、183.72、95.09、174.82、29.85 (mg/kg)/h。在连续口灌给药5d后, 停药后盐酸氯苯胍在斑点叉尾鮰肠道中的浓度最高, 肌肉中浓度最低, 盐酸氯苯胍在体内各组织中的消除速度由高到低依次为血浆、鳃、脑、肌肉、皮肤、肝脏、肾脏、肠。若将10 μg/kg作为盐酸氯苯胍的最高残留限量, 在试验条件下, 建议盐酸氯苯胍在斑点叉尾鮰体内的休药期至少应为23d。     

盐酸氯苯胍在美国红鱼体内的药代动力学和残留消除规律研究

在水温(28±2)℃、盐度28条件下,盐酸氯苯胍(robenidine hydrochloride,ROBH)按30 mg/kg的剂量口灌实验鱼,用HPLC-MS/MS法研究盐酸氯苯胍在美国红鱼体内的药代动力学和残留消除规律。结果显示,单剂量口灌给药后,美国红鱼血浆中ROBH的药时数据符合一级吸收二室模型,药物在血浆中的达峰时间(tp)、血药浓度峰值(Cmax)、药时曲线下面积(AUC_(0-∞))和消除半衰期(t_(1/2β))分别为2.39 h、958.78μg/L、33 247.57μg/(L·h)和19.24 h;ROBH在肌肉、肝脏和肾脏的Cmax分别为156.72μg/kg、227.68μg/kg和553.44μg/kg,tp分别为2.0 h、1.5 h、2.0 h;AUC_(0-∞)分别4 664.04μg/(kg·h)、4 897.74μg/(kg·h)、17 228.19μg/(kg·h);t_(1/2β)分别为19.68 h、24.33 h和22.81 h。按30 mg/kg剂量连续5 d口灌给药后,美国红鱼肌肉、肝脏、肾脏中的药物消除半衰期(t1/2):24.46 h、35.39 h、39.60 h和33.94 h。若以10μg/kg为最高残留限量,肌肉作为食用靶组织,在本试验条件下,建议休药期不少于7 d。     

基于液相色谱电化学法的鸡肉中磺胺类药物残留量测定

目的:建立了鸡肉中7种磺胺类药物残留量的液相色谱测定方法。方法:样品中加入乙腈、无水硫酸钠,均质、离心,再加入乙腈,提取合并乙腈提取液。在提取液中加入乙腈饱和正己烷溶液净化,用高效液相色谱法测定。结果:7种磺胺类药物标准曲线的线性范围10~800μg/kg,检出限为10μg/kg,回收率为69.8%~95.7%,相对标准偏差为5.6%~12.7%。结论:液相色谱法适合对鸡肉中磺胺类药物进行检测,结果准确。     

双氟沙星对异育银鲫血脑屏障渗透性及消除规律简

以异育银鲫(Carassais auratus gibebio)为研究对象,采用组织匀浆法和高效液相色谱法,研究了双氟沙星(Difloxacin,DIF)通过异育银鲫血脑屏障情况,并比较分析了大脑和外周组织中DIF消除差异。结果显示,根据DIF 96h半数致死剂量(2840 mg/kg b.W)给药后,第96h时异育银鲫大脑组织匀浆中DIF的含量为(10.49±0.35)μg/g;同时在临床推荐用药剂量(20 mg/kg)给药后的15个时间点(0?960h)上均能从大脑组织匀浆中检测出DIF。上述结果表明DIF能渗透通过血脑屏障而进入异育银鲫大脑组织。另外,在大脑和外周组织消除过程上,以大脑组织中的DIF消除过程最为平缓(按照20 mg/kg给药)。到试验第960h,大脑组织中DIF含量最高,为(0.392±0.007)μg/g,且大脑中的消除半衰期最长,为1157.713h。因此,异育银鲫大脑组织可作为DIF药物残留分析的靶组织。另根据欧盟关于食品中DIF最大残留限量(MRL)之规定,实验条件下DIF休药期至少为25d。结果为研究鱼类血脑屏障作用,DIF神经毒性及其在水产养殖上的临床应用提供了参考。     

天可力氨糖胶囊中氨基葡萄糖硫酸钾测定方法比较

目的:选择一种能准确测定天可力氨糖胶囊中氨基葡萄糖硫酸钾含量的方法。方法:参照GB/T 20365-2006硫酸软骨素和盐酸氨基葡萄糖含量的方法进行液相色谱法测定以及利用盐酸氨基葡萄糖Fdson—Morgan反应后在525 nm左右波长处有吸收峰,测定其吸收值与标准品比较进行比色定量。结果:液相色谱法测定时胶囊中的其它中药成分干扰很大,数据偏差太大;而比色法测定时,氨基葡萄糖盐酸盐含量在175μg/mL浓度范围内,呈良好线性关系:Y(μg/mL)=0.196X-O.004,R=0.997,平均回收率102.09%。结论:该比色法测定方法灵敏、准确、简便,可供天可力氨糖胶囊中氨基葡萄糖硫酸钾含量测定。     

高效液相色谱法测定大鼠血浆中来那度胺的浓度及其药动学(英文)

目的:建立高效液相色谱法测定大鼠血浆中来那度胺的浓度。方法:色谱柱采用VenusilASBC18column(4.6 mm×250mm,5μm)购自于博纳艾杰尔科技有限公司。样品采用梯度洗脱,流动相为乙腈和0.05%甲酸溶液,流速为1.0 mL/min,检测波长为254 nm,以沙利度胺为内标。结果:来那度胺血药浓度的线性范围为0.1-5μg/mL,最低检测限为0.1μg/mL,三个浓度的QC样品(0.2,1 and 5μg/mL)的提取回收率分别为78.8±3.8,80.1±3.2 and 79.1±7.6%。结论:此方法准确、简便、灵敏度高,对来那度胺的血药浓度测定和药物代谢动力学研究极具价值。     

双氟沙星对异育银鲫血脑屏障渗透性及消除规律

以异育银鲫（Carassais auratus gibebio）为研究对象,采用组织匀浆法和高效液相色谱法,研究了双氟沙星（Difloxacin,DIF）通过异育银鲫血脑屏障情况,并比较分析了大脑和外周组织中DIF消除差异。结果显示,根据DIF 96h 半数致死剂量（2840 mg/kg b.W）给药后,第96h时异育银鲫大脑组织匀浆中DIF的含量为（10.490.35） g/g;同时在临床推荐用药剂量（20 mg/kg）给药后的15个时间点（0960h）上均能从大脑组织匀浆中检测出DIF。上述结果表明DIF能渗透通过血脑屏障而进入异育银鲫大脑组织。另外,在大脑和外周组织消除过程上,以大脑组织中的DIF消除过程最为平缓（按照20 mg/kg给药）。到试验第960h,大脑组织中DIF含量最高,为（0.3920.007） g/g,且大脑中的消除半衰期最长,为1157.713h。因此,异育银鲫大脑组织可作为DIF药物残留分析的靶组织。另根据欧盟关于食品中DIF最大残留限量（MRL）之规定,实验条件下DIF休药期至少为25d。结果为研究鱼类血脑屏障作用,DIF神经毒性及其在水产养殖上的临床应用提供了参考。       

高效液相法测定盐酸乐卡地平片含量

使用高效液相色谱法测定乐卡地平片含量,流动相为乙腈-0.O1 mol/L乙酸铵溶液—三乙胺(650:350:1)(pH 6.0).结果显示盐酸乐卡地平在浓度为2.05～404.0μg/mL范围内具有良好的线性关系,盐酸乐卡地平片剂的平均标示量含量为100.3％,符合要求.研究表明HPLC色谱法对乐卡地平片剂进行含量测定...     

拔毒消炎软膏质量控制研究

目的研究拔毒消炎软膏的质量控制方法。方法用薄层色谱法(HPLC)对制剂中大黄、黄柏进行定性鉴别,高效液相色谱法测定大黄酚的含量。结果定性鉴别薄层色谱斑点特征明显;高效液相色谱法测定含量,大黄酚在0.061~0.304μg/ml范围内呈良好的线性关系(r=0.9954),平均加样回收率为98.68%,RSD=1.39%。结论采用薄层色谱法定性、高效液相色谱法定量,简便准确、重现性良好,可有效控制该制剂质量。     

Multiresidue determination of quinolone antibacterials in eggs of laying hens by liquid chromatography with fluorescence detection

An analytical method for the simultaneous determination of seven quinolones (ciprofloxacin, enrofloxacin, danofloxacin, difloxacin, flumequine, oxolinic acid and sarafloxacin) in egg samples of laying hens was developed. Their use is totally prohibited in animals from which eggs are produced for human consumption. Protein precipitation was achieved by addition of acetonitrile and ammonia, removal of acetonitrile with dichloromethane, the quinolones remaining in the basic aqueous extract. The aqueous extract was analysed by liquid chromatography with fluorescence detection (LC-FD). The mobile phase was composed of acetonitrile and 10 mM citrate buffer solution of pH 4.5, with an initial composition of acetonitrile-water (12:88, v/v) and using linear gradient elution. Norfloxacin was used as an internal standard. The limits of detection found were 4-12 ng g(-1). These values were lower than the maximum residue limits (MRLs) established by the European Union for these compounds in different tissues of eggs-producing animals.     

Simultaneous determination of fluoroquinolones and tetracyclines in chicken muscle using HPLC with fluorescence detection

A multiresidue method has been developed which allows for the simultaneous determination of both fluoroquinolones and tetracyclines in chicken muscle. Samples were extracted with a mix of acetonitrile and 0.1 M citrate, 150 mM MgCl(2), pH 5.0. After centrifugation and evaporation, the extracts could be analyzed by liquid chromatography with fluorescence detection. Good recoveries (63-95%) were obtained from samples fortified with a mix of five fluoroquinolones and three tetracyclines, with satisfactory relative standard deviations. Limits of detection were 0.5 ng/g (danofloxacin), 1 ng/g (oxytetracycline, ciprofloxacin, enrofloxacin), 1.5 ng/g (tetracycline), 2 ng/g (difloxacin) and 5 ng/g (sarafloxacin, chlortetracycline). Enrofloxacin and its metabolite ciprofloxacin, as well as oxytetracycline were determined in enrofloxacin and oxytetracycline incurred chicken muscle using this method.     

High-performance liquid chromatographic analysis of ascorbyl-2-phosphate in fish tissues

l-Ascorbic-2-phosphate magnesium salt (APM) in fish tissues was determined by high-performance liquid chromatography. APM extracted with 5% metaphosphoric acid was separated by a LiChrospher 100 RP18 column within 20 min. The detection limit was 0.1 μg/g tissue.     

Simple and sensitive determination of five quinolones in food by liquid chromatography with fluorescence detection

A simple and sensitive high-performance liquid chromatographic (HPLC) method has been developed for the determination of five different quinolones: enrofloxacin, ciprofloxacin, sarafloxacin, oxolinic acid and flumequine in pork and salmon muscle. The method includes one extraction and clean-up step for the five quinolones together which are detected in two separated HPLC runs by means of their fluorescence. The proposed analytical method involves homogenizing of the tissue sample with 0.05 M phosphate buffer, pH 7.4 and clean-up by Discovery DS-18 cartridges. For chromatographic separation a Symmetry C(18) column is used in two different runs: (1) ciprofloxacin, enrofloxacin and sarafloxacin with acetonitrile-0.02 M phosphate buffer pH 3.0 (18:82) as mobile phase and the detector at excitation wavelength: 280 nm and emission wavelength 450 nm; and (2) oxolinic acid and flumequine with acetonitrile-0.02 M phosphate buffer pH 3.0 (34:66) as mobile phase and excitation wavelength: 312 nm and emission wavelength: 366 nm. Detection limit was as low as 5 ng g(-1), except for sarafloxacin which had a limit of 10 ng g(-1). Standard curves using blank muscle tissues spiked at different levels showed a good linear correlation coefficient, r(2) higher than 0.999 for all quinolones.     

Simultaneous determination of danofloxacin and N-desmethyldanofloxacin in cattle and chicken edible tissues by liquid chromatography with fluorescence detection

A rugged, simple, and selective method for the determination of danofloxacin and its primary metabolite, N-desmethyldanofloxacin, in cattle (liver, muscle, kidney, and fat) and chicken (liver and muscle) tissues was developed. The method is selective for danofloxacin and N-desmethyldanofloxacin over other veterinary important fluoroquinolones, such as enrofloxacin, ciprofloxacin, norfloxacin, and ofloxacin. Selectivity is achieved through a combination of extraction, chromatography, and fluorescence detection. The analytes were extracted from homogenized tissues using a methanolperchloric-phosphoric acid solution. After centrifugation, direct injection of extraction supernate was possible. The limit of quantitation was 20 pg on column. Separation was achieved on an Inertsil C₈ (5 μm, 100 Å) column with dimensions of 250×4.6 mm I.D. The mobile phase consisted of 0.05 M phosphate buffer (pH 3.5)-acetonitrile (88:12). A fluorescence detector was utilized with an excitation wavelenght of 280 nm and an emission wavelength of 440 nm. The assay was accurate and reproducible within the range of 10 to 500 ng/g for both danofloxacin and N-desmethyldanofloxacin. Intra-assay accuracy was between 98 and 101%, and precision was less than 4%. Inter-assay accuracy was between 99 and 102%, while precision was less than 2%. Recoveries for both analytes over the dynamic range were greater than 90% for all the tissues.     

Multiresidue analysis of fluoroquinolone antibiotics in chicken tissue using liquid chromatography-fluorescence-multiple mass spectrometry

An efficient liquid chromatographic method for the multiresidue analysis of fluoroquinolone antibiotics in chicken tissue has been developed in which quantitation using fluorescence and confirmation with multiple mass spectrometry (MS(n)) was achieved simultaneously. Using this method, eight fluoroquinolones were analyzed in fortified samples of chicken liver and muscle tissue with recoveries at levels of 10-200 ng/g generally in the range of 60-93%, except for desethylene ciprofloxacin, which consistently gave recoveries >or=45%. Relative standard deviations were excellent in all cases, and the limits of detection in ng/g were determined as follows in liver and (muscle): desethylene ciprofloxacin 0.3 (0.1), norfloxacin 1.2 (0.2), ciprofloxacin 2 (1.5), danofloxacin 0.2 (0.1), enrofloxacin 0.3 (0.2), orbifloxacin 1.5 (0.5), sarafloxacin 2 (0.6), difloxacin 0.3 (0.2). Confirmation of the identities of the fluoroquinolones was achieved by monitoring the ratios of two prominent product ions in MS(2) (desethylene ciprofloxacin) or MS(3) (all others). Levels of confirmation as related to ion ratio variability criteria were established. Enrofloxacin and ciprofloxacin were also determined in enrofloxacin incurred chicken liver and muscle using this method.     

A Validated Normal Phase LC Method for Enantiomeric Separation of Rasagiline Mesylate and Its (S)‐Enantiomer on Cellulose Derivative‐Based Chiral Stationary Phase

A simple, sensitive, and robust normal‐phase isocratic HPLC‐UV method was developed and validated for the enantiomeric separation of rasagiline mesylate and its (S)‐enantiomer. The rasagiline and its (S)‐enantiomer were resolved on a Chiralcel‐OJ‐H (4‐methylbenzoate cellulose coated on silica) column using a mobile phase consisting of n‐hexane:isopropyl alcohol:ethanol:diethyl amine (96:2:2:0.01) at a flow rate of 1.0 ml/min. The column temperature was maintained at 27 °C and elution was monitored at 215 nm. The resolution (R_s) between the enantiomers was found to be more than 2.0. The limit of detection and the limit of quantification of the (S)‐enantiomer were found to be 0.35 and 1.05 µg/ml, respectively. The developed method was validated as per ICH guidelines with respect to linearity, limit of detection and quantification, accuracy, precision, and robustness—and satisfactory results were obtained. The sample solution and mobile phase were found to be stable up to 48 h. The method is useful for routine evaluation of the quality of rasagiline mesylate in bulk drug‐manufacturing units. Chirality 25:324–327, 2013. © 2013 Wiley Periodicals, Inc.     

Determination of the enantiomers of a novel 20,21-dinoreburnamenine derivative in rat plasma and brain by high-performance liquid chromatography using a chiral stationary phase

A high-performance liquid chromatographic method with solid-phase extraction was developed for the assay of the enantiomers of a novel 20,21-dinoreburnamenine derivative (RU 49041) in rat plasma and brain using a chiral stationary phase (Nucleosil Chiral 2) and ultraviolet detection. The limit of detection was 10 ng/ml (or ng/g) in both tissues and the intra-assay precision was satisfactory (plasma, ca. 5%; brain, ca. 1%). The pharmacokinetic profiles of the two enantiomers were determined following oral administration of the racemate (10 mg/kg). The results show that their pharmacokinetics are very different: whereas both enantiomers appear in the brain, only the 3α,16β-enantiomer is detected in plasma.     

金鱼胰蛋白酶原基因的克隆及镉和过氧化氢暴露对其基因表达的影响

为深入研究胰蛋白酶在鱼类中的蛋白结构和生理功能, 利用RT-PCR和RACE方法, 从金鱼肝胰脏中成功克隆获得了一种全长864 bp的胰蛋白酶原cDNA序列(gfTryp)。gfTrypc DNA包含21 bp的5′-非翻译区、114 bp的3′-非翻译区和729 bp的开放读码框, 编码242个氨基酸组成的胰蛋白酶原(gfTryp)。gfTryp含有15个氨基酸的信号肽和5个氨基酸(LDDDK)的激活肽。氨基酸序列分析表明, gfTryp具备胰蛋白酶原的保守结构特征, 如含有催化三联体氨基酸(His-57、Asp-102和Ser-195), 12个半胱氨酸, 位于底物结合口袋底部Asp-189和口袋开口处的Gly-216、Gly-226等, 提示其可能具有保守的蛋白消化功能。RT-PCR结果显示, gfTryp mRNA在所检测的各个组织中均有表达, 其中在肝胰脏、肠和脂肪中表达量为最高。进一步研究发现, 相较于摄食前, 肝胰脏gfTryp mRNA在金鱼摄食后显著升高。在0.5和5 μg/mL镉暴露处理后, 肝胰脏gfTryp mRNA显著升高(与未处理组相比, 分别约为3.2 和 4.7倍)。随着镉浓度增加到10 μg/mL后, gfTryp mRNA表达量下降。经100 μmol/L过氧化氢处理3h、6h、12h和24h后, 金鱼肝胰脏gfTryp mRNA的表达水平均显著下降, 在6h达到最大效应(约为对照组的0.21倍)。研究结果证实了重金属镉和过氧化氢处理能调控胰蛋白酶原基因表达, 为进一步探讨鱼类消化生理提供了新的视角。     

High-performance liquid chromatographic determination of gossypol and gossypolone enantiomers in fish tissues using simultaneous electrochemical and ultraviolet detectors

This paper describes a method for residue analysis of difloxacin and sarafloxacin in chicken muscle. Clean-up and preconcentration of the samples are effected by solid-phase extraction (C18) and the determination is carried out by capillary electrophoresis using a photodiode array detection system. The method was validated with satisfying results. The calibration graphs are linear for difloxacin and sarafloxacin from 50 to 300 microg/kg. The limit of detection obtained for difloxacin and sarafloxacin are 10 and 25 microg/kg, respectively, which allows the detection of positive muscle samples at the required maximum residue limits of European Union.