Simultaneous determination of 15 marker constituents in various radix Astragali preparations by solid-phase extraction and high-performance liquid chromatography

An improved quality control method was developed to simultaneously determine 15 major constituents (eight flavonoids and seven saponins) in various radix Astragali preparations, using SPE for pretreatment of samples, HPLC with diode-array and evaporative light scattering detectors (DAD-ELSD) for quantification in one run, and HPLC-ESI-TOF/MS for definite identification of compounds in preparations. Optimum separations were obtained with a ZORBAX C(18) column, using a gradient elution with 0.3% aqueous formic acid and ACN. This established method was fully validated with respect to linearity, precision, repeatability, and accuracy, and was successfully applied to quantify the 15 compounds in 19 commercial samples, including 3 dosage forms, i. e., oral solution, injection, concentrated granule, and its processed products of radix Astragali. The results demonstrated that many factors might result in significant differences in quality of the final preparations, including crude drugs, pretreatment processes, manufacturing procedure, storage conditions, etc. Then the developed method provided a reasonable and powerful manner to ensure the efficacy, safety, and batch-to-batch uniformity of radix Astragali products by standardizing each procedure, and thus should be proposed as quality control for the clinical use and modernization of herbal preparations.     

Determination of organophosphorus pesticides and metabolites in crops by solid-phase extraction followed by liquid chromatography/Diode array detection

Summary A multiresidue method for the analysis of 28 common organophosphorus pesticides and 3 of their main metabolites (paraoxon-ethyl, paraoxon-methyl and malaoxon) in a variety of crop samples has been developed. An aliquot of the chopped sample is homogenized with an organic solvent. The efficiency of extraction methods using methanol, acetone and acetonitrile was evaluated. The acetonitrile gives higher recoveries and minimizes co-extractives from the samples matrix. The resulting aqueous acetonitrile extract is filtered and cleaned by solid phase extraction (SPE). For SPE three different types of adsorption materials (Carbograph 1, LiChrobut-EN and Amberchrom CG-161m) were compared. The cleaned-up extract is injected into the LC system. Three different analytical columns were tested in conjunction with two mobile phase compositions of different polarity. The use of LC-DAD techniques allowed the identification of both organophosphorus pesticides and metabolites by means of standard and spectral comparison, respectively. The accuracy of the quantitative determination measured in terms of average percentage recovery of 31 compounds in crop samples was 61–96% with a relative standard deviation of 5–10%.     

Selective solid-phase extraction of amoxicillin and cephalexin from urine samples using a molecularly imprinted polymer

In this paper we describe, for the first time, a molecularly imprinted polymer (MIP) for the antibiotic amoxicillin (AMX), synthesised by a noncovalent molecular imprinting approach and used to extract AMX selectively from urine samples. The MIP was applied as a molecularly selective sorbent in molecularly imprinted SPE (MISPE) in an off-line mode, where it showed useful cross-selectivity for a structurally related antibiotic, cephalexin (CPX). By using a MISPE protocol, the MIP was able to selectively extract both AMX and CFX from 5 mL of water spiked with 10 mg/L with recoveries of 75 and 78% for AMX and CFX, respectively. When applied to real samples (urine) at clinically relevant concentrations, recoveries from 2 mL of human urine spiked with 20 mg/L decreased slightly to 65 and 63% for AMX and CFX, respectively. To demonstrate further the selectivity of the MIP obtained, a comparison with commercially available SPE cartridges was performed. Improvements in the retention of both AMX and CFX on the MIP were obtained relative to the commercially available cartridges, and the MISPE extracts were considerably cleaner, due to molecularly selective analyte binding by the MIP.     

High‐performance liquid chromatography with diode‐array detection cotinine method adapted for the assessment of tobacco smoke exposure

Smoking is considered to be one of the main risk factors for cancer and other diseases and is the second leading cause of death worldwide. As the anti‐tobacco legislation implemented in Europe has reduced secondhand smoke exposure levels, analytical methods must be adapted to these new levels. Recent research has demonstrated that cotinine is the best overall discriminator when biomarkers are used to determine whether a person has ongoing exposure to tobacco smoke. This work proposes a sensitive, simple and low‐cost method based on solid‐phase extraction and liquid chromatography with diode array detection for the assessment of tobacco smoke exposure by cotinine determination in urine. The analytical procedure is simple and fast (20 min) when compared to other similar methods existing in the literature, and it is cheaper than the mass spectrometry techniques usually used to quantify levels in nonsmokers. We obtained a quantification limit of 12.30 μg/L and a recovery of over 90%. The linearity ranges used were 12–250 and 250–4000 μg/L. The method was successfully used to determine cotinine in urine samples collected from different volunteers and is clearly an alternative routine method that allows active and passive smokers to be distinguished.     

Stereoselective determination of hydroxychloro-quine and its major metabolites in human urine by solid-phase microextraction and HPLC

The enantioselective analysis of hydroxychloroquine (HCQ) and its major metabolites was achieved by HPLC and solid-phase microextraction. The chromatographic separation was performed on a Chiralcel OD-H column using hexane/methanol/ethanol (96:2:2, v/v/v) plus 0.2% diethylamine as the mobile phase, at the flow rate of 1.3 mL/min. The main extraction parameters were optimized. The best condition was achieved by the addition of 10% NaCl and 1 mL phosphate buffer 1 mol/L pH 11 to 3 mL human urine. The extraction was conducted for 40 min at 25 degrees C and the desorption time was 3 min using methanol (100%). PDMS-DVB 60 microm fiber was used in this study. The mean recoveries were 9.3, 9.2, and 14.4% for HCQ, desethylhydroxychloroquine (DHCQ), and desethylchloroquine (DCQ), respectively. The method was linear over the range of 50-1000 ng/mL for HCQ enantiomers and over the range of 42-416 ng/mL for DCQ and DHCQ enantiomers. Within-day and between-day precision and accuracy assays for HCQ and its metabolites were lower than 15%. The preliminary 48 h urinary excretion study performed in human urine showed to be stereoselective. The amount of (+)-(S)-enantiomer excreted was higher than its antipode.     

Simultaneous determination of seven phthalates and four parabens in cosmetic products using HPLC-DAD and GC-MS methods

Studies on the determination of seven kinds of phthalates, i.e. diethyl phthalate, dipropyl phthalate, dibutyl phthalate, benzyl butyl phthalate, dicyclohexyl phthalate, di-(2-ethylhexyl) phthalate, and dioctyl phthalate, and four parabens, i.e. methylparaben, ethylparaben, propylparaben, and butylparaben, in 15 kinds of cosmetic products, including hair sprays, perfumes, deodorants, cream, lotion, etc., by HPLC with diode array detection and GC-MS in electron impact ionization mode with selected-ion monitoring have been carried out. Methods have been developed for both qualitative and quantitative detection of phthalates and parabens. Extraction, clean-up, and analysis procedures have been optimized. HPLC and GC-MS determinations were performed after sonication-assisted extraction with methanol and clean-up with C18 SPE. These techniques permit detection of phthalates at a level of 10.0-100.0 microg/kg and of parabens at a level of 20.0-200.0 microg/kg. Overall recoveries were 85-108% with RSD values of 4.2-8.8%. Only one of the 15 examined samples was free from phthalates and parabens. The remaining 14 samples were found to contain at least three or more of these phthalates and/or parabens. The predominant phthalates and parabens detected in the studied samples were methylparaben, propylparaben, diethyl phthalate, dibutyl phthalate, dicyclohexyl phthalate, and di-(2-ethylhexyl) phthalate. The residue level is at 1.22-5289 mg/kg.     

Simultaneous extraction and determination of free and conjugated phytosterols in tobacco

Acid hydrolysis and alkaline saponification were incorporated into a microwave‐assisted extraction process for the simultaneous extraction of free and conjugated phytosterols from tobacco. The crude extract of the microwave‐assisted extraction was purified by C₁₈ solid‐phase extraction and then determined by high‐performance liquid chromatography. Phytosterols of cholesterol, ergosterol, stigmasterol, campesterol, and β‐sitosterol were determined by chromatographic quantification. The multiple parameters of microwave‐assisted extraction were optimized by a uniform design method. The optimal ratio of extraction ethanol solvent to tobacco mass was 30 mL/g. The microwave‐assisted extraction acid hydrolysis was carried out in sulfuric acid medium by heating for 10 min at 55°C. The microwave‐assisted extraction alkaline saponification was performed after adding excessive sodium hydroxide by heating another 10 min. The repeatability of the proposed method was acceptable with recoveries from 69.68 to 88.17% for the phytosterols. Five target phytosterols were all found in the tobacco samples, and the contents were significantly different in samples from different producing areas.     

Microwave-assisted micellar extraction coupled with solid-phase extraction for preconcentration of pharmaceuticals in molluscs prior to determination by HPLC

A simple and specific analytical method was developed and tested for the determination of pharmaceuticals in mollusc samples. A combination of microwave-assisted micellar extraction (MAME) and solid-phase extraction (SPE) using a non-ionic surfactant, polyoxyethylene 10 lauryl ether, was examined to extract and determine simultaneously a group of pharmaceuticals such as carbamazepine, clorfibric acid, ketoprofen, naproxen, bezafibrate and ibuprofen by liquid chromatography using UV-diode array detector. The MAME extraction performance was evaluated by studying various parameters such as the volume and concentration of surfactant and microwave conditions. Finally, an OASIS HLB cartridge was used as an optimum SPE sorbent to clean up the extracts and preconcentrate the selected analytes. The proposed method showed satisfactory linearity and reproducibility (between 3 and 15%), as well as detection limits ranging from 30 to 220 ng/g. Finally, the method was successfully applied to the determination of the target pharmaceuticals in various kinds of mollusc samples. This study has demonstrated that microwave-assisted micellar extraction with solid-phase extraction may be used as a viable alternative to conventional methods for the extraction of pharmaceuticals in this type of matrices.     

固相萃取-高效液相色谱检测葡萄酒中赭曲霉毒素A

使用C18小柱固相萃取, C18反相柱(250 mm×4.6 mm i.d.)分离,V(乙腈):V(水):V(乙酸)＝99:99:2为流动相,荧光检测器(激发波长333 nm,发射波长460 nm)检测,测定葡萄酒中赭曲霉毒素A.其质量浓度在6.25～200 ng/mL范围内呈良好线性,相关系数为0.9997.样品经浓缩60倍后,方法检出限为0.027 ng/mL.对红葡萄酒、干红及白葡萄酒进行了加标回收实验,回收率为80.1%～109.8%.平行7份样品加标回收率相对标准偏差为5.9%.对市售6种葡萄酒进行了赭曲霉毒素A的测定.     

Simultaneous determination of pharmaceutically active compounds in wastewater samples by solid phase extraction and high-performance liquid chromatography with diode array and fluorescence detectors

In the present work, an analytical method for the simultaneous determination of five anti-inflammatory drugs (acetaminophen, diclofenac, ibuprofen, ketoprofen and naproxen), an antiepileptic drug (carbamazepine) and a nervous stimulant (caffeine) is proposed for the routine analysis of these pharmaceuticals in wastewater influents and effluents from WWTPs. The method involves pre-concentration and clean-up by solid phase extraction (SPE) using Oasis HLB extraction cartridges. Final analysis of the selected pharmaceutical compounds was carried out by high-performance liquid chromatography (HPLC) with diode array detector (DAD). Confirmation of the presence of the fluorescence compounds (ibuprofen and naproxen) was performed by on-line fluorescence detection. Recoveries were ranged from 71 to 103% with relative standard deviation below 15.1%. Limits of quantification were in the range 6.2–319.8 and 3.0–160.0 ng ml⁻¹ for influent and effluent wastewater samples, respectively. The described method was applied to the determination of the drugs in wastewater samples from four treatment plants in Seville.     

Combined solid-phase extraction and gas chromatography-mass spectrometry used for determination of chloropropanols in water

A sensitive and rapid derivatization method for the simultaneous determination of 1,3-dichloro-2-propanol (1,3-DCP) and 3-chloropropane-1,2-diol (3-MCPD) in water samples has been developed. The aim was to research the optimal conditions of the derivatization process for two selected reagents. A central composite design was used to determine the influence of derivatization time, derivatization temperature and reagent volume. A global desirability function was applied for multi-response optimization. The analysis was performed by gas chromatography-mass spectrometry. During the optimization of the extraction procedure, four different types of solid-phase extraction (SPE) columns were tested. It was demonstrated that the Oasis HLB cartridge produced the best recoveries of the target analytes. The pH value and the salinity were investigated using a Doehlert design. The best results for the SPE of both analytes were obtained with 1.5 g of NaCl and pH 6. The proposed method provides high sensitivity, good linearity (R(2)≥0.999) and repeatability (relative standard deviations % between 2.9 and 3.4%). Limits of detection and quantification were in the range of 1.4-11.2 ng/mL and 4.8-34.5 ng/mL, respectively. Recoveries obtained for water samples were ca. 100% for 1,3-DCP and 3-MCPD. The method has been successfully applied to the analysis of different samples including commercially bottled water, an influent and effluent sewage.     

Determination of amphetamines in human urine by liquid chromatography with fluorimetric detection using a solid-phase extraction procedure

A precise and feasible HPLC method has been developed for the analysis of amphetamine (AMPH), methamphetamine (MAMPH) and methylenedioxymethamphetamine (MDMA, ecstasy) in human urine. A chromatographic run on a C8 Genesis (150 mm x 4.6 mm, 5 microm) column maintained at 30 degrees C lasts about 17 min, using a mobile phase composed of ACN (12%) and a pH 2.5 phosphate buffer (88%) containing 0.3% triethylamine. Mirtazapine was used as the internal standard. Good linearity was found in the 100-2000 ng/mL concentration range for AMPH and MAMPH and in the 12-2000 ng/mL concentration range for MDMA. The pretreatment of urine samples was carried out by means of a careful SPE procedure on C2 cartridges. The extraction yields were very satisfactory for all analytes, with average values greater than 97%. The leading conditions allowed the determination of AMPH, MAMPH and MDMA with satisfactory precision and accuracy. The method has been successfully applied to the determination of the analytes in urine of AMPH users.     

Flow-injection determination of phenylephrine hydrochloride in pharmaceutical dosage forms with on-line solid-phase extraction and spectrophotometric detection

A flow injection method is proposed for the determination of phenylephrine hydrochloride in pharmaceutical dosage forms. The method involves the use of on-line solid-phase extraction by means of a microcolumn containing Dowex 50W X8 ion-exchange resin for the separation of the analyte prior to colour development and spectrophotometric detection in the visible region.

The influence of preconcentration flow, preconcentration pH and elution volume was studied.

The method exhibits appropriate linearity (r² = 0.9999) which was proved statistically by means of the “F”-test. When applied to commercial samples containing several active ingredients and excipients, a significant reduction of interferences was found. Accuracy, evaluated by means of the spike recovery method was in the range 99.7–100.8%, with precision (R.S.D., %) better than 1%.

In order to achieve the automation the system was controlled from a notebook computer by means of a program written in QuickBASIC language. Under these conditions, a sampling frequency of 40 samples per hour could be attained.     

Effective cleanup for the determination of six quinolone residues in shrimp before HPLC with diode array detection in compliance with the European Union Decision 2002/657/EC

A high‐performance liquid chromatographic method was developed for the determination of six quinolone residues (ciprofloxacin, enrofloxacin, sarafloxacin, oxolinic acid, nalidixic acid, and flumequine) in shrimp tissue samples. Separation was carried out by a LiChrospher® 100 RP‐8e column, running at a 22 min gradient elution program, and the mobile phase consisted of citric acid (0.4 mol/L), acetonitrile and methanol. Detection was achieved by a diode array detector, monitoring at 255 and 275 nm. Sample preparation included initial extraction with citric acid solution and further clean‐up by solid‐phase extraction, employing Lichrolut RP‐18 cartridges. Validation was performed according to the European Union Decision 2002/657/EC. The detection capability was 127.2 μg/kg for ciprofloxacin, 115.2 μg/kg for enrofloxacin, 126.2 μg/kg for sarafloxacin, 113.1 μg/kg for oxolinic acid, 125.2 μg/kg for nalidixic acid, and 239.0 μg/kg for flumequine. Recoveries ranged between 83.0 and 121.6%. The Youden test was applied to study the method ruggedness.     

Simultaneous determination of four trace estrogens in feces,leachate, tap and groundwater using solid–liquid extraction/auto solid‐phase extraction and high‐performance liquid chromatography with fluorescence detection

A simple and selective high‐performance liquid chromatography method coupled with fluorescence detection was developed for the simultaneous measurement of trace levels of four estrogens (estrone, estradiol, estriol and 17α‐ethynyl estradiol) in environmental matrices. For feces samples, solid–liquid extraction was applied with a 1:1 v/v mixture of acetonitrile and ethyl acetate as the extraction solvent. For liquid samples (e.g., leachate and groundwater), hydrophobic/lipophilic balanced automated solid‐phase extraction disks were selected due to their high recoveries compared to conventional C₁₈ disks. Chromatographic separations were performed on a reversed‐phase C₁₈ column gradient‐eluted with a 45:55 v/v mixture of acetonitrile and water. The detection limits were down to 1.1 × 10^?2 (estrone), 4.11 × 10^?4 (estradiol), 5.2 × 10^?3 (estriol) and 7.18 × 10^?3 μg/L (17α‐ethynyl estradiol) at excitation/emission wavelengths of 288/310 nm, with recoveries in the range of 96.9 ± 3.2–105.4 ± 3.2% (n = 3). The method was successfully applied to determine estrogens in feces and water samples collected at livestock farms and a major river in Northeast China. We observed relatively high abundance and widespread distribution of all four estrogens in our sample collections, implying the urgency for a comprehensive and intricate investigation of estrogenic fate and contamination in our researched area.     

Simultaneous determination of albiflorin and paeoniflorin in rat urine by solid-phase extraction and high-performance liquid chromatography following oral administration of Si-Wu decoction

A high performance liquid chromatographic method (HPLC), together with solid phase extraction (SPE), was developed for simultaneous determination of albiflorin and paeoniflorin in rat urine after oral administration of Si-Wu decoction. The samples were pretreated with solid phase extraction using Extract-Cleantrade mark cartridges. Analysis of the extract was performed on a reversed-phase C18 column and a mobile phase made up of acetonitrile and 0.03% formic acid (17:83, v/v). UV detection was set at 230 nm. The assay was linear over the range 2.625-52.50 mg/mL for albiflorin and 3.875-77.50 microg/mL for paeoniflorin. The average percentage recoveries of three spiked urines were 97.01 +/- 3.32 and 102.32 +/- 6.97 for albiflorin and paeoniflorin, respectively. The intra-day precision (RSD) ranged from 0.21 to 1.79% at concentrations of 4.20, 10.50, 26.25 and 39.375 microg/mL of albiflorin and 0.12 to 2.92% at concentrations of 3.875, 10.85, 23.25 and 58.125 microg/mL of paeoniflorin, and inter-day precision (RSD) was from 1.02 to 1.86% for albiflorin and 0.94 to 3.30% for paeoniflorin, at the same four concentrations. This method was applied in order to analyze albiflorin and paeoniflorin in rat urine following oral administration of traditional Chinese medicinal preparation of Si-Wu decoction.     

Simultaneous detection of three antiviral and four antibiotic compounds in source‐separated urine with liquid chromatography

An analytical method for the simultaneous screening of three antiviral agents (nevirapine, zidovudine, lamivudine), four antibiotics (sulfamethoxazole, trimethoprim, ciprofloxacin, rifampicin) and one reference compound (carbamazepine) in human urine was developed. Separation was achieved with a Kinetex XB‐C₁₈ (75 × 4.6 mm, 2.6 μm) column after the extraction of pharmaceuticals from urine with SPE. Gradient elution with a mobile phase consisting of acetonitrile and 10 mM KH₂PO₄ (pH 2.5), and diode array detection with monitoring at 210 and 264 nm was applied. The developed method was validated in terms of selectivity, linearity, stability and sensitivity. Repeatability (n = 3) and between‐day precision (n = 3) revealed RSD <5%. The detection limits were estimated as 0.02–0.54 g/L (depending on compound). The method was validated for human urine and successfully applied to the simultaneous quantification of selected compounds. Strata‐X cartridges provided good recoveries ranging from 81 to 109%. The limits of detection for urine varied between 0.04 and 1.61 g/L. The method is suitable for the fast determination of selected pharmaceuticals from source‐separated urine samples for further environmental risk assessment and degradation potential evaluation. It provides a way to enhance safe nutrient recycling from wastewater streams and promotes the safe use of urine as fertiliser.     

Simultaneous spectrophotometric determination of carbamazepine and phenytoin in serum by PLS regression and comparison with HPLC

Carbamazepine (CBZ) and phenytoin (PHT) are two antiepileptic drugs which are used simultaneously. In this paper a partial least-squares (PLS) calibration method is described for the simultaneous spectrophotometric determination of CBZ and PHT in plasma. Standard binary mixtures of CBZ and PHT have been resolved by application of PLS-1 to their UV spectra. Then, the binary standard solutions, spiked to plasma, were prepared and after the extraction of the drugs, their corresponding UV spectrum were analyzed by PLS regression to calculate the concentration of drugs in unknown plasma. A leave one out cross-validation procedure was employed to find the optimum numbers of latent variables using PRESS. A HPLC method was also applied for simultaneous determination of two drugs in the plasma and in methanol. The mean recoveries obtained by PLS were 98.4 and 98.2 for CBZ and PHT and those obtained by HPLC were 100.1 and 101.7, respectively. Although, the HPLC method showed better performance than PLS, it was found that the results obtained by PLS were comparable with those obtained by HPLC method.     

A new molecularly imprinted polymer for selective extraction of cotinine from urine samples by solid-phase extraction

Cotinine, the main metabolite of nicotine in human body, is widely used as a biomarker for assessment of direct or passive exposure to tobacco smoke. A method for molecularly imprinted solid-phase extraction (MISPE) of cotinine from human urine has been investigated. The molecularly imprinted polymer (MIP) with good selectivity and affinity for cotinine was synthesized using cotinine as the template molecule, methacrylic acid as the functional monomer, and ethylene glycol dimethacrylate as the cross-linker. The imprinted polymer was evaluated for use as a SPE sorbent, in tests with aqueous standards, by comparing recovery data obtained using the imprinted form of the polymer and a non-imprinted form (NIP). Extraction from the aqueous solutions resulted in more than 80% recovery. A range of linearity for cotinine between 0.05 and 5 μg mL⁻¹ was obtained by loading 1 mL blank urine samples spiked with cotinine at different concentrations in acetate buffer of pH 9.0, and by using double basic washing and acidic elution. The intra-day coefficient of variation (CV) was below 7% and inter-day CV was below 10%. This investigation has provided a reliable MISPE–HPLC method for determination of cotinine in human urine from both active smokers and passive smokers. Figure     

Tandem use of solid-phase extraction and dispersive liquid-liquid microextraction for the determination of mononitrotoluenes in aquatic environment

Solid‐phase extraction (SPE) in tandem with dispersive liquid–liquid microextraction (DLLME) has been developed for the determination of mononitrotoluenes (MNTs) in several aquatic samples using gas chromatography‐flame ionization (GC‐FID) detection system. In the hyphenated SPE‐DLLME, initially MNTs were extracted from a large volume of aqueous samples (100 mL) into a 500‐mg octadecyl silane (C₁₈) sorbent. After the elution of analytes from the sorbent with acetonitrile, the obtained solution was put under the DLLME procedure, so that the extra preconcentration factors could be achieved. The parameters influencing the extraction efficiency such as breakthrough volume, type and volume of the elution solvent (disperser solvent) and extracting solvent, as well as the salt addition, were studied and optimized. The calibration curves were linear in the range of 0.5–500 μg/L and the limit of detection for all analytes was found to be 0.2 μg/L. The relative standard deviations (for 0.75 μg/L of MNTs) without internal standard varied from 2.0 to 6.4% (n=5). The relative recoveries of the well, river and sea water samples, spiked at the concentration level of 0.75 μg/L of the analytes, were in the range of 85–118%.