高效液相色谱法测定动物组织中四环素类抗生素残留量的研究

建立了动物组织中四环素、金霉素、土霉素、强力霉素、去甲基金霉素、甲烯土霉素和二甲胺四环素等7种四环素类抗生素残留量的液相色谱同时测定方法。方法采用Inertsil C8-3(5μm,250 mm×4.0 mm i.d)反相色谱柱,以pH 4.0的EDTA-Mcllvaine缓冲溶液为提取溶液,以HLB固相萃取柱为净化柱,流动相为甲醇+乙腈+0.01mol/L三氟乙酸(梯度洗脱),流速1.5 mL/min,检测波长350 nm,进样量100μL。方法的检出限为1.5~5.0μg/kg,测定低限为50μg/kg,线性范围为50~1200μg/kg,加标回收率为73.8%~103%,相对标准偏差为0.5%~8.5%。方法适用于动物肌肉、肝脏和肾脏组织中7种四环素类抗生素残留量的同时检测。     

Introduction of the HPLC method for the determination of quinolone residues in various muscle tissues

For use in veterinary sanitary control of foodstuffs and raw materials of animal origin in Slovenia, we developed a routine and confirmation analytical method for determining the residues of enrofloxacin, ciprofloxacin and flumequine in the muscle tissue of cattle, pigs and poultry. For the muscle tissue of freshwater fish, the determination of the flumequine residues was introduced. The results obtained through simultaneous determination of the residues of enrofloxacin, ciprofloxacin and flumequine showed that the values for the examined antibiotics were up to 600 times lower than the prescribed maximum residue levels (MRL). Another advantage of this method is that it covers a wide range of different fluoroquinolones.     

Chromatographic determination of herbicide residues in various matrices

The newest results in the use of various extraction techniques and chromatographic methods such as gas-liquid and high-performance liquid chromatography used for the assessment of herbicide residues in various matrices have been compiled and critically evaluated. Practical employments in water and soil research, environmental protection, clinical and food chemistry are presented.     

Measurement of beta-agonist residues in retinal tissue of food producing animals

This report presents the results of an investigation on the accumulation of beta-agonist residues in the retinal tissue of food producing animals. Three different species, calf, pig and turkey, were treated with six different beta-agonists and analysed for beta-agonist residues in retinal tissue applying a newly developed retina preparation procedure which provides sufficient sample material for multiple analyses. The results show that all selected beta-agonists accumulate in the retina, though in varying concentrations. The results are discussed on the basis of existing binding theories and with regard to their impact on the existing residue control strategy for beta-agonists.     

Automatic chemiluminescence-based determination of carbaryl in various types of matrices

Carbaryl, a modern pesticide widely used for both agricultural and non-agricultural purposes, was determined from the chemiluminescence produced in its reaction with Ce(IV) in a nitric acid medium containing rhodamine 6G as sensitizer, using flow-injection techniques. A straightforward automatic method based on measurements peak height and peak area, which are directly proportional to the carbaryl concentration, was thus developed. Calibration graphs are linear over the concentration range from 50 to 2000 ng mL⁻¹. The limit of detection, as determined according to Clayton, is 45.6 and 28.7 ng mL⁻¹ for peak height and peak area measurements, respectively. The relative standard deviation for 10 samples was less than 1.4% with both types of measurements. Two commercial formulations containing carbaryl were analysed using both types of measurements, which provided acceptable recovery values. Solid-phase extraction was used to concentrate and separate the analyte from the matrix. The method was successfully applied to the analysis of spiked water samples as well as in soil and grain samples. The proposed method exhibited a high selectivity no other pesticide containing the naphthalene group such as antu, napropamide or naftalam, etc., was found to interfere with the determination of carbaryl.     

An HPLC method for determination of azadirachtin residues in bovine muscle

A high-performance liquid chromatography (HPLC) method for the determination of azadirachtin (A and B) residues in bovine muscle has been developed. Azadirachtin is a neutral triterpene and chemotherapeutic agent effective in controlling some pest flies in horses, stables, horns and fruit. The actual HPLC method uses an isocratic elution and UV detection. Liquid-liquid extraction and solid-phase purification was used for the clean-up of the biological matrix. The chromatographic determination of these components is achieved using a C18 analytical column with water-acetonitrile mixture (27.5:72.5, v/v) as mobile phase, 1 mL/min as flow rate, 45 °C column temperature and UV detector at 215 nm. The azadirachtin peaks are well resolved and free of interference from matrix components. The extraction and analytical method developed in this work allows the quantitation of azadirachtin with precision and accuracy, establishing a lower limit of quantitation of azadirachtin, extracted from the biological matrix.     

The determination of raloxifene in rat tissue using HPLC

We report a rapid and reliable HPLC-UV method for determination of raloxifene, a kind of selective estrogen receptor modulator (SERM), in rat tissue. Proteins were precipitated by adding 200 microL of acetonitrile and 50 microL of methanol to 100 microL of the tissue homogenates, following vortex mixing and centrifugation. Separation was carried out on a reversed-phase C(18) column (150 x 4.6 mm, 5 microm) with a mobile phase of acetonitrile:0.05 m ammonium acetate (pH 4.0 +/- 0.1; 33:67, v/v) at a flow rate of 1.0 mL/min. The UV detection wavelength was set at 289 nm and the temperature of column was kept at 23 degrees C, without interference from endogenous tissue compounds. The calibration curve was linear from 0.0125 to 10.0 microg/mL with correlation coefficient of over 0.994, while the limit of quantification was 0.008 microg/mL. The intra- and inter-day coefficients of variation were less than 10% (RSD). The recovery of assay was between 95.8 and 104.5%. Furthermore, the method was used to measure the concentration of raloxifene in rat tissue after a simple oral dose. The highest level was observed in liver, lung, spleen, then heart and kidney. The lowest level was found in brain. These results suggest that raloxifene distributes rapidly and moderately into tissues such as liver, lung and spleen.     

Post-harvest HPLC determination of chlorfluazuron residues in pears treated with different programs

The present study was conducted to monitor the level of chlorfluazuron residues in pear samples in order to assess the risk posed by the presence of such residues to the consumer. Chlorfluazuron was sprayed onto pear trees at the recommended dose rate at two different times at 30 and 21 days prior to harvesting in one treatment, at 21 and 14 days prior to harvesting in another treatment, and three times at 30, 21 and 14 days prior to harvesting in a third treatment. Chlorfluazuron residues were extracted with acetonitrile and partitioned into ethyl acetate. The residue determination was performed on an Apollo C(18) column using HPLC with a UV detection of 254 nm following the clean-up of the extract by open column chromatography with Florisil. The versatility of this method was evidenced by its good linearity (>0.995) in the concentration range between 0.2 and 10 microg/mL. The majority of the mean recoveries at two different fortification levels, 0.05 and 0.25 ppm, ranged from 84.9 +/- 3.2 to 94.3 +/- 10.6, and the repeatability (as the relative standard deviation) from three repetitive determinations of recovery was between 3.8 and 11%. The calculated limit of detection (LOD) was 0.008 ppm and the limit of quantitation was 0.03 ppm. Trace amounts of chlorfluazuron were detectable when it was applied onto the pear trees at two or three times prior to harvesting; however, the levels of chlorfluazuron were not quantified. The excellent sensitivity and selectivity of this method allowed for quantitation and identification at low levels with a run time of less than 12 min. Chlorfluazuron can be used safely to protect pears when sprayed two or three times at 14 days prior to harvesting.     

高效液相色谱法同步检测牛奶中替米考星、泰乐菌素和螺旋霉素残留量

研究了牛奶中替米考星、泰乐菌素和螺旋霉素残留量的液相色谱同步测定方法。方法采用ZORBAX Eclipse XDB C18(5μm,150 mm×4.6 mmi.d)反相色谱柱,以甲醇为提取液,以SCX因相萃取柱为净化柱,流动相为0.05 mol/L磷酸二氢钠溶液 乙腈,梯度洗脱,流速1 mL/min,用二极管阵列检测器检测,替米考星和泰乐菌素的检测波长285 nm,螺旋霉素的检测波长232 nm,进样量100μL。替米考星、泰乐菌素和螺旋霉素的检出限分别为:30、20、40μg/kg,线性范围为20~800μg/kg,加标回收率为88.8%~99.4%,相对标准偏差为2.2%~8.9%。方法适用于牛奶中替米考星、泰乐菌素和螺旋霉素残留量的同步检测。     

HPLC determination of cefotaxime and cephalexine residues in milk and cephalexine in veterinary formulation

An HPLC method was developed and validated for the determination of the cephalosporins cefotaxime and cephalexine in skimmed bovine milk. The analytical column, Kromasil C₁₈ (250 mm × 4.0 mm, 5 μm) was operated at ambient temperature. Mobile phase consisted of CH₃OH-acetate buffer (pH = 4.0) and it was delivered isocratically at a flow rate of 1.0 mL · min⁻¹. Total analysis time was less than 5 min. Caffeine was used as internal standard (5 ng · μL⁻¹). UV detection was performed at 265 nm. Method validation was performed by means of intra-day (n = 5) and inter-day accuracy and precision (n = 8), sensitivity and linearity. Limits of detection (LOD) and limits of quantification (LOQ) were 0.1 and 0.3 ng · μL⁻¹, respectively. The method was applied to the analysis of a veterinary drug (CEPOREX) containing cephalexine. The results were quite accurate with the relative error varying from −8.0 to −3.5%. Solid-phase extraction was applied to remove all matrix interference from milk samples. High extraction recoveries (average 84–121%) were achieved by using Abselut NEXUS cartridges with acetonitrile as eluent and a rinsing step with water and n-butanol. A pre-concentration step was necessary in a 1/10 level to reach the EU MRL concentration level (100 μg · kg⁻¹). RSD values were less than 7% for both cephalosporins. Correspondence: Ioannis N. Papadoyannis, Laboratory of Analytical Chemistry, Department of Chemistry, Aristotle University of Thessaloniki, GR-54124 Thessaloniki, Greece     

Screening and determination of melamine residues in tissue and body fluid samples

Melamine is a chemical product that was sporadically mixed into animal feeds to boost protein content. Excessive melamine in animal feed can induce renal failure and even death in animals. The residue of melamine in edible animal products also threatens human health. Currently, there is no real-time and high throughput method to detect residual melamine in animal tissues. Successful development of such methods is very important for fast and on-site screening of melamine residue in animal tissues to eliminate the potential threat to human health. Here we demonstrate the detection of residual melamine from swine and chicken tissues and body fluids using indirect competitive enzyme-linked immunosorbent assay (ELISA) method. A detection sensitivity of 0.5 μg mL⁻¹ and a limit of detection of 0.05 μg mL⁻¹ were achieved with this method. A gas chromatography-mass spectrometry (GC-MS) method was also developed to act as a confirmatory and quantitative procedure for the ELISA results. The limits of quantitation (LOQ) of were 0.01 μg g⁻¹ and 0.005 μg mL⁻¹ for tissues and body fluids, respectively. The two methods showed good agreement (r² > 0.992). The method developed was performed on samples of tissues from chickens fed with melamine-spiked feed.     

Simultaneous determination of nine types of phthalate residues in commercial milk products using HPLC-ESI-MS-MS

A multi-residue method was developed for the confirmation and quantitation of nine types of phthalates in milk using high-performance liquid chromatography electrospray ionization tandem mass spectrometry. The samples were extracted with acetonitrile. The analytes were separated using a 0.1% formic acid-methanol system as the mobile phase, and a linear gradient elution program. Mass spectral acquisition was achieved by selectively monitoring the ions in electro-spray ionization mode. Qualitative analysis was based on the retention time and the mass spectrum results, and the quantity was carried out by comparison with the external standard. The mean recoveries for each analyte ranged from 65.2% to 98.3%, with relative standard deviations below 11.2%. The limits of detection were 5～25 μg/kg, and the limits of quantitation were 17～83 μg/kg, depending on the compounds. This method has the merits of convenient operation, high sensitivity, and good repeatability, making it an effective method for analysis of phthalates in milk. And the proposed analytical method has been applied to the analysis of phthalates presented in four commercial milk products. The main phthalate residues were DBP and DMP. And the amount of DBP was found to be more than 100 μg/kg in all this milk products.     

Trace determination of chromium in various water types by adsorption differential pulse voltammetry

Summary A new sensitive voltammetric method is presented for the determination of trace amounts of total chromium [Cr(III) and Cr(VI)] in natural waters. The method is based on the preconcentration of the Cr(III)-DTPA complex by adsorption at the HMDE at the potential of –1.0 V. The adsorbed complex is then reduced producing a response with a peak potential of –1.22 V and the peak height of the Cr(III) reduction is measured. The catalytic action of nitrate and bromate ions on the Cr(III)-DTPA reduction has been elucidated using cyclic voltammetry. The adsorption of chromium complexes at the HMDE was investigated using out-of-phase a. c. voltammetry and the potential range of adsorption was determined. Based on these investigations optimal conditions for the determination of the total chromium concentration in the range 20–2,000 ng/l have been established. The determination limit is 20 ng/l and the RSD is 5% for chromium concentrations 200 ng/l.The usefulness and wide scope of this new voltammetric method for reliable and highly sensitive chromium analysis down to the natural ultra trace levels existing in various types of natural waters is demonstrated by determinations of the total dissolved chromium content in river, lake, sea and rain water.

---

Spurenbestimmung von Chrom in verschiedenen Wassertypen durch Adsorptions-Differentialpuls-Voltammetrie

Zusammenfassung Eine neue voltammetrische Methode zur Spurenbestimmung von Chrom [als Summe von Cr(III) und Cr(VI)] in natürlichen Gewässern wurde entwickelt. Die Methode beruht auf einer Anreicherung des Cr(III)-DTPA-Komplexes durch Adsorption an der hängenden Queck-silbertropfenelektrode beim Potential –1.0 V. Der adsorbierte Komplex wird anschließend im differentiellen Pulsmodus reduziert und die Peakhöhe beim Peakpotential –1.22 V gemessen. Die katalytische Wirkung von Nitrat- und Bromationen auf die Cr(III)-DTPA-Reduktion wurde mit der cyclischen Voltammetrie geklärt. Die Adsorption der Cr-Komplexe wurde zusätzlich mit der a.c.-Voltammetrie (kapazitive Komponente) untersucht und der Potentialbereich der Adsorption ermittelt. Aufgrund der Untersuchungen wurden die optimalen Bedingungen zur Chrombestimmung im Konzentrationsbereich 20–2000 ng/l festgelegt. Die Bestimmungsgrenze liegt bei 20 ng/l und die relative Standardabweichung beträgt 5% für Konzentrationen 200 ng/l. Die weite Anwendbarkeit der Methode für die zuverlässige und hochempfindliche Analyse von Chromspuren bis zu den natürlichen Ultraspurengehalten in verschiedenen Typen natürlicher Wässer wird an Beispielen der Analyse des gelösten Gesamtgehaltes von Chrom in Flußwasser, Seewasser, Meerwasser und Regenwasser aufgezeigt.

---

---

Dedicated to Prof. Dr. H. Monien on the occasion of his 60th birthday

---

Attached from Department of Chemistry of Warsaw University, Poland, within the scope of the joint research project on Eco-toxic Metals in the Environment     

A rapid and sensitive HPLC procedure for the determination of atrazine residues in soil-water extracts

An automated procedure is described for the rapid determination of the residual herbicide atrazine at ppb levels in soil-water extracts. The system consists of an autosampler connected to a C18 precolumn linked via a switching valve to a C18 analytical column and spectrophotometer. Peak height is superior to peak area for quantifying results with coefficients of variation of 0.48% and 0.89%, respectively, for 7.5 ng standard injections. Precolumn concentration eliminates the need for sample cleanup and achieves up to a 37% increase in peak height compared to an isocratic system. In a black earth soil, atrazine is identified as the principal phytotoxic constituent. The minimum detectable level is one-tenth the critical no-effect threshold for wheat. The method is well suited for predicting possible atrazine injury to susceptible crops and the measurement of other nonpolar analytes in aqueous solution.     

Simultaneous separation of nine surfactants of various types by HPLC with evaporative light scattering detection

A simultaneous separation of cationic, amphoteric and nonioinc nine surfactants (DMDS, DMDP, DMDM, DMDL, BZC, CDE, A/O, SUNC, IMD) has been performed by a reverse phase-HPLC method utilizing a single J'sphere ODS (250 mm × 4.6 mm, 4 μm) column and a methanol-water containing 0.2% TFA eluent system within 60 min. The observed precision in determination of concentration was less than 5% R.S.D., which revealed that ELSD was an effective tool to detect the various studied surfactants of low volatility without chromophore. In addition, the detection limits were in the concentration range of 3.5-10 μg/mL, and the calibration curves, i.e. the log-log plots, were linear in the working range of 5-4600 μg/mL with the slopes of 1.321-1.668. The application of the analytical procedure to three household products without pretreatment supported that the presented chromatographic method was simple to be practical for a routine analysis of commercial products.     

An efficient procedure for extraction and determination of steroids in the tissue of laboratory animals

Summary A high-performance liquid chromatographic method for analysis of steroids in the tissue of laboratory animals is described. A two-stage solid extraction procedure, utilizing Sep-pak C₁₈ and graphitized carbon black, gives a purified product. Steroids in this extract were separated by multistep gradient elution with a reversedphase liquid chromatography system, using acetonitrilewater as solvent. Several columns with different geometry and phase loading were investigated. Optimum resolution was obtained with a medium covered and spherical particle.The applicability of this method was successfully demonstrated in the analysis of the testis and adrenal gland of rat.Dedicated to Prof. Dr. A. Liberti on the occasion of his 70th birthday.     

高效液相色谱法测定小鼠血浆和组织中的阿昔洛韦

改良HPLC用于测定小鼠血浆和组织中阿昔洛韦浓度。使用Beckinan Gold系列高效液相色谱仪,ODS柱,流动相为甲醇－水（5：95,V／V）,流速1．0mL/min,检测波长254nm,样品采用12％三氯醋酸去蛋白。血浆和组织中阿昔洛韦浓度在0．05－20mg/L范围内与峰高呈良好线性关系,最低检测浓度分别为0.02.0.05mg/L,天间精密度均优于6％。