On-line sample cleanup in the liquid chromatographic analysis of pharmaceuticals for citrate content

Matrix interferents are removed from pharmaceutical samples via an on-line, automated column switching process in which the difference in hydrophobicity between the analyte and the interferents allows these species to be isolated in different parts of the chromatographic system. In this case, the interferents are trapped on a cleanup column and are flushed to waste as the analyte undergoes additional separation on an analytical column. The utility of this approach is demonstrated by the quantitation of citrate in pharmaceutical samples by ion suppression, reversed-phase liquid chromatography. The performance of this system is statistically equivalent to that of a manual pretreatment method employing disposable, solid-phase extraction cartridges.     

High-performance liquid chromatographic determination of ambroxol in human plasma.

Ambroxol has been determined in biological fluids using a rapid and sensitive high-performance liquid chromatographic method. The samples prepared from plasma by liquid-liquid extraction were analysed on reversed-phase silica gel by competing-ion chromatography with ultraviolet detection. The method was applied to the determination of ambroxol levels in twelve healthy volunteers after oral administration of 90 mg of ambroxol in tablets of Mucosolvan and Ambrosan.     

中性溶质在MEKC中柱内富集的理论研究

毛细管电动力学色谱(MEKC)中通过“扫”的技术可以使样品组分在线富集。本研究基于柱分离过程弛豫理论的基本方法,对溶质在MEKC中柱内输运过程加以研究,得到了描述进样长度及样品区带和运行区带性质差别对富集效果影响的理论表达式。通过对进样长度及溶质在两区带中的容量因子与富集效果之间关系的进一步探讨,证实溶质在胶束中的溶解度是影响富集的最重要因素;在基本不影响分离效果的情况下,适当加大进样长度一般对富集有利。     

On-line SFE-GC for determination of PCBs in human milk and blood serum

On-line coupled supercritical fluid extraction and gas chromatography (SFE-GC) has been utilized for the determination of PCBs and other organochlorine compounds in human milk and blood serum. The procedure involved preconcentration of the sample on C₁₈-silica sorbent in an extraction cell: after precipitation of the proteins up to 20 ml of human milk was concentrated on 0.5 g of sorbent. Serum (up to 5 ml) was applied to the C₁₈ material without pretreatment. Basic alumina was utilized as a selective adsorbent for lipids in the on-line SFE-GC system. The method was used to analyze milk and serum spiked with 0.5 and 10 ng of Aroclor 1260 and the results compared with those obtained by liquid–liquid extraction of serum.     

Instrumental planar chromatographic method for determination of carbamazepine in human serum

An instrumental planar chromatographic (HPTLC) method for quantification of carbamazepine in human serum was developed using liquid‐liquid extraction with dichloromethane, fluorescence activation with perchloric acid 60%/ethanol/water (1:1:1, v/v) and fluorescence detection. Planar chromatographic separation was performed on precoated silica gel F254 HPTLC plates using a mixture of ethyl acetate/toluene/methanol/acetic acid glacial (5:4:0.5:0.5, v/v) as mobile phase. Densitometric detection was done at 366 nm. The method was validated for linearity, precision and accuracy. Linear calibration curves in the range of 3 and 20 ng/μL showed correlation coefficient of 0.998. The intra‐assay and inter‐assay precision, expressed as the RSD, were in the range of 0.41–1.24% (n = 3) and 2.17–3.17% (n = 9), respectively. The LOD was 0.19 ng, and the LOQ was 0.57 ng. Accuracy, calculated as percentage recovery, was between 98.98 and 101.96%, with a RSD not higher than 1.52%. The method was selective for the active principle tested. In conclusion, the method is useful for quantitative determination of carbamazepine in human serum.     

On-capillary sample cleanup method for the electrophoretic determination of carbohydrates in juice samples

On many occasions, sample treatment is a critical step in electrophoretic analysis. As an alternative to batch procedures, in this work, a new strategy is presented with a view to develop an on-capillary sample cleanup method. This strategy is based on the partial filling of the capillary with carboxylated single-walled carbon nanotube (c-SWNT). The nanoparticles retain interferences from the matrix allowing the determination and quantification of carbohydrates (viz glucose, maltose and fructose). The precision of the method for the analysis of real samples ranged from 5.3 to 6.4%. The proposed method was compared with a method based on a batch filtration of the juice sample through diatomaceous earth and further electrophoretic determination. This method was also validated in this work. The RSD for this other method ranged from 5.1 to 6%. The results obtained by both methods were statistically comparable demonstrating the accuracy of the proposed methods and their effectiveness. Electrophoretic separation of carbohydrates was achieved using 200 mM borate solution as a buffer at pH 9.5 and applying 15 kV. During separation, the capillary temperature was kept constant at 40 degrees C. For the on-capillary cleanup method, a solution containing 50 mg/L of c-SWNTs prepared in 300 mM borate solution at pH 9.5 was introduced for 60 s into the capillary just before sample introduction. For the electrophoretic analysis of samples cleaned in batch with diatomaceous earth, it is also recommended to introduce into the capillary, just before the sample, a 300 mM borate solution as it enhances the sensitivity and electrophoretic resolution.     

Evaluation of the sweep co-distillation cleanup technique for the determination of environmental contaminants in human adipose tissue

Beef and pork fat, corn, peanut, rapeseed and paraffin oils, as well as vegetable shortening were used to investigate their suitability as fortification media for environmental chemicals in the evaluation of the sweep co-distillation technique. The animal fats produced considerable gas chromatographic background interference, while the oils were partly carried over during the sweep co-distillation process, except for rapeseed and peanut oil. Residue free rapeseed oil was fortified with 26 environmental chemicals in several groups at 20 and 200 ng/g of oil. Recoveries for most compounds were greater than 80% with a coefficient of variation of less than or equal to 10. At the 20 ppb fortification level, recoveries for Aroclor 1260, Mirex and pentachlorobenzene were only 70-80%. A similar low recovery was observed for p,p'-DDT, Mirex, hexachloro-1,3-butadiene, while at the 200 ppb level, 1,2,4-trichlorobenzene was only 33% recovered. The sweep co-distillation technique was further evaluated by using rendered human fat and the same fat diluted with residue free peanut oil. Residue levels in diluted and non-diluted fat were in good agreement, except for hexachlorobenzene. These residue levels were further compared with those obtained by two other cleanup procedures: Florisil-silicic acid column chromatography and low temperature precipitation. In general the sweep co-distillation technique compared favourably with these other cleanup procedures. There was evidence, however, that p,p'-DDT broke down into p,p'-TDE and varying operating procedures did not completely remedy this situation.     

On-line solid-phase extraction and high-performance liquid chromatographic determination of nortriptyline and amitriptyline in serum.

An isocratic reversed-phase high-performance liquid chromatographic method with on-line solid-phase extraction for the simultaneous determination of amitriptyline and nortriptyline in serum has been developed. A 250-microliters serum sample is injected directly onto a commercially available CN cartridge and, after a washing step, the retained solutes are backflushed onto a bonded-phase CN column using a column-switching technique and a mobile phase composed of acetonitrile (26%) and 0.05 M phosphate buffer with diethylamine. Serum is diluted with 0.1 M sodium lauryl sulphate and centrifuged before the injection. Detection at 210 nm ensures sufficient sensitivity. The recovery is almost quantitative and the relative standard deviation ranges from 2.8 to 8.0% for concentrations of 200-40 ng/ml. Being rapid and simple, the method is convenient for routine use.     

Micellar liquid chromatographic method for the determination of ephedrine and pseudoephedrine in human serum by direct inject of the sample with simple dilution

A micellar high-performance liquid chromatographic method was developed to simultaneously determine ephedrine and pseudoephedrine in human serum. The serum sample pretreatment was a simple dilution in a micellar solution, filtration, and direct injection, thus avoiding time-consuming and tedious steps. Hence, there is no need to use an internal standard. The serum samples were analyzed using a mobile phase containing 1.50?×?10^?1?mol/L sodium dodecyl sulfate and 0.02?mol/L sodium dihydrogen phosphate with 7.5% (v/v) 1-propanol at pH 3.0, running at 1.0?mL/min by an Inertsil C18 (150?×?4.6?mm, 5?µm) column at 30°C. The UV wavelength was set at 210?nm. The developed method was validated by linearity (r?>?0.9990) and intra- and inter-day precisions of ephedrine and pseudoephedrine (relative standard deviation; RSD%, 0.04–10.40, and RSD %, 0.30–10.25, respectively), LODs for ephedrine and for pseudoephedrine was 2.63 and 2.70?µg/mL, respectively; lower limit of quantification for ephedrine and for pseudoephedrine was 4.38 and 4.51?µg/mL, respectively. Finally, the proposed method was applied to investigate ephedrine and pseudoephedrine in real human serum samples after oral administration of Kechuanning Koufye including Ephedra herb. It is environmentally friendly, easy-to-handle, and feasible method for routine analysis in clinical laboratory.     

On-line introduction of high-pressure gas-liquid sample for capillary gas chromatographic analysis

An on-line sample introduction technique in capillary gas chromatograph (CGC) for the analysis of high-pressure gas-liquid mixtures has been designed and evaluated. A sample loop of 0.05 microL and a washing solvent loop of 0.5 microL are mounted on a 10-port switching valve, which serves as the injection valve. A capillary resistor was connected to the vent of sample loop in order to maintain the pressure of the sample. Both the sample and the washing solvent are transferred into the split-injection port through a narrow bore fused silica capillary inserted into the injection liner through a septum. The volume of the liner is used both as the pressure-release damper and evaporation chamber of the sample. On-line analysis of both reactants and resultants in ethylene olimer reaction mixture at 5 MPa was carried out, which demonstrated the applicability of the technique.     

Sensitive high-performance liquid chromatographic method for the determination of chlorhexidine in human serum and urine

A high-performance liquid chromatographic method for the analysis of chlorhexidine in human serum and urine was developed. Chlorhexidine and the internal standard, chlorpheniramine, were extracted into chloroform, containing 5% 2-propanol, and back-extracted into dilute sulfuric acid. Chromatographic separation was achieved on a C18 column equilibrated with methanol-water (70:30, v/v), containing 0.005 M sodium heptane-sulfonate. The sensitivity of the assay was 20 ng/ml of biological matrix, using 0.5-ml samples. The application of this method to monitor chlorhexidine levels in burn patients treated topically with a chlorhexidine-containing burn cream was demonstrated.     

On-line solid-phase extraction-HPLC-fluorescence detection for simultaneous determination of puerarin and daidzein in human serum

Response surface methodology (RSM) was applied to the optimization of on-line solid-phase extraction (SPE) parameters, and an automated system of on-line SPE coupled with high-performance liquid chromatography (HPLC) with fluorescence detection was developed for the determination of puerarin and daidzein in human serum. The human serum sample of 50 μL was injected into a conditioned C18 SPE cartridge, and the matrix was washed out with acetonitrile-KH₂PO₄-triethylamine buffer (0.01 M, pH 7.4) (3:97, v/v) for 3 min at a flow rate of 0.25 mL/min. Then the target analytes were eluted and transferred to the analytical column. A chromatographic gradient elution was programmed with the mobile phase consisting of acetonitrile and KH₂PO₄-triethylamine buffer, and the analytes were determined with a fluorescence detector at excitation wavelength of 350 nm and emission wavelength of 472 nm, respectively. The proposed method presented good linear relations (0.85-170 μg/mL for puerarin and 0.2-40 μg/mL for daidzein), satisfactory precision (RSD < 8%), and accredited recovery (92.5-107.8%).     

Sequential injection chromatographic determination of ambroxol hydrochloride and doxycycline in pharmaceutical preparations

A new separation method based on a novel reversed-phase sequential injection chromatography (SIC) technique was used for simultaneous determination of ambroxol hydrochloride and doxycycline in pharmaceutical preparations in this contribution.The coupling of short monolith with SIA system results in an implementation of separation step to until no-separation low-pressure method.A Chromolith^® Flash RP-18e, 25-4.6 mm column (Merck, Germany) and a FIAlab^® 3000 system (USA) with a six-port selection valve and 5 ml syringe were used for sequential injection chromatographic separations in our study. The mobile phase used was acetonitrile-water (20:90, v/v), pH 2.5 adjusted with 98% phosphoric acid, flow rate 0.48 ml min⁻¹, UV detection was at 213 nm.The validation parameters have shown good results: linearity of determination for both compounds including internal standard (ethylparaben) >0.999; repeatability of determination (R.S.D.) in the range 0.5-5.4% at three different concentration levels, detection limits in the range 0.5-2.0 μg ml⁻¹, and recovery from the pharmaceutical preparation in the range 99.3-99.9%. The chromatographic resolution between peak compounds was >5.0 and analysis time was <9 min under the optimal conditions. The method was found to be applicable for routine analysis of the active compounds ambroxol hydrochloride and doxycycline in various pharmaceutical preparations.     

Multiresidue method for the gas chromatographic determination of pesticides in honey after solid-phase extraction cleanup

A new multiresidue method is described for the determination of pesticides in honey. The method involves dissolution of the honey in a methanol-water mixture, followed by solid-phase extraction cleanup and gas chromatographic determination. Twenty-six pesticides used on flowering field crops, on flowering fruit and vegetables, or as acaricides to control Varroa jacobsoni in beehives are determined by the method. Recoveries from honey, spiked at 0.02-1.6 mg/kg, ranged from 85 to 127% with a relative standard deviation (RSD) of 2-16%, except for the RSD of 27% for captan at 0.05 mg/kg.