Micelle-mediated extraction of tanshinones from Salvia miltiorrhiza bunge with analysis by high-performance liquid chromatography

The feasibility of employing non-ionic surfactant oligoethylene glycol monoalkyl ether (Genapol X-080) as an alternative and effective solvent for the extraction of tanshinones from Salvia miltiorrhiza bunge was studied for the first time. Various experimental conditions were investigated to optimize the extraction. Under optimum conditions, i.e. 10% Genapol X-080 (w/v), liquid/solid ratio of 20:1 (ml g⁻¹), ultrasonic-assisted extraction for 45 min, the extraction recovery of the tanshinones reached the highest value. When compared with commonly used solvents, 10% Genapol X-080 yielded almost the same extraction efficiency as methanol and dichloromethane-methanol (1:4). For the pre-concentration of tanshinones by cloud-point extraction (CPE), sodium chloride was added to the solution to facilitate the phase separation and increase the pre-concentration factor by reducing the volume of the surfactant-rich phase.     

Determination of the isoflavone genistein in soybeans by high-performance liquid chromatography following cloud point extraction

A simple and sensitive method has been developed for preconcentration and determination of genistein in soybeans. This method is based on cloud point extraction (CPE) of genistein from soybeans using ethylene glycol monoalkyl ether (Genapol X-080) as a nonionic surfactant. The concentration of extracted genistein was determined by HPLC with a UV detector. Optimum experimental conditions were established. With 5% Genapol X-080 (v/v), a liquid/solid ratio of 25:1 mL/g, and ultrasonic-assisted extraction at 40 degrees C for 45 min, the extraction percentage of genistein reached its highest value. The preconcentration factor for genistein was about 16.5. The RSD for seven replicate measurements and the LOD were +/- 4.45% and 15.0 ng/mL, respectively. CPE is simple, inexpensive, and suitable for extraction of genistein from soybean. It uses environmentally friendly surfactants and offers a convenient alternative to more conventional extraction systems.     

Application of non-ionic surfactant in the microwave-assisted extraction of alkaloids from Rhizoma Coptidis

The feasibility of employing non-ionic surfactant oligoethylene glycol monoalkyl ether (Genapol X-080) as an alternative and effective solvent for the microwave-assisted extraction of alkaloids from Rhizoma Coptidis was demonstrated. When compared with commonly used solvents, 5% Genapol X-080 enhanced the extraction efficiency. Under optimal conditions, i.e. 5% acidified Genapol X-080 (v/v), microwave-assisted extraction for 10 min at 100 °C, the extraction recovery of alkaloids reached 92.8% in a one-step extraction. The efficiency of cloud-point preconcentration of three alkaloids was in the range 93.6-94.7% with relative standard deviation lower than 3.3% by the proposed procedure. Furthermore, the combination of microwave-assisted extraction and cloud-point extraction was shown to be a green, rapid and effective approach for alkaloids preconcentration of Rhizoma Coptidis samples.     

Combination of microwave assisted micellar extraction and liquid chromatography for the determination of organophosphorous pesticides in soil samples

A new methodology based on the microwave assisted micellar extraction (MAME) technique has been optimised, using soil samples, to extract and determine a mixture of the eight organophosphorous pesticides mainly used in agriculture. The pesticides under study have been extracted using the non-ionic surfactants polyoxyethylene 10 lauryl ether (POLE) and oligoethylene glycol monoalkyl ether (Genapol X-080). The optimal extraction variables, such as surfactant concentration, pH, radiation time and microwave power were determined for each surfactant. The results show the advantage of using POLE instead of Genapol X-080 for the extraction of the organophosphorus pesticides with recoveries higher than 70% for most of the compounds and relative standard deviations (RSD) below 2.6%. This method was successfully applied to fresh samples as well as to aged samples for the analysis of soils with different characteristics and compared with the traditional Soxhlet technique.     

Determination of benzimidazole fungicides in soil samples using microwave-assisted micellar extraction and liquid chromatography with fluorescence detection

A simple and fast analytical method was developed for the determination of benzimidazole fungicides (benomyl, carbendazim, thiabendazole, and fuberidazole) in soil samples. The analytes were extracted from the soil samples by means of conventional microwave-assisted extraction, using the non-ionic surfactants polyoxyethylene 10 lauryl ether (POLE) and oligoethylene glycol monooalkyl ether (Genapol X-080) as extractants. Determinations were made by using liquid chromatography with direct fluorescence detection. The use of an analytical column Symmetry C-18 offered short retention times of analytes without the need of any pH regulators with mobile phase methanol-water (50 + 50, v/v). The best results were obtained using 5% (v/v) POLE as extractant with recoveries of the fungicides in spiked soil samples between 71 and 105%. The results were compared with those obtained when Soxhlet extraction was applied to the same soil samples.     

Development and application of high-performance liquid chromatography for the study of ampelopsin pharmacokinetics in rat plasma using cloud-point extraction

A simple, rapid and specific method based on cloud-point extraction (CPE) was developed to determine ampelopsin in rat plasma after oral administration by reversed-phase high-performance liquid chromatography. The non-ionic surfactant Genapol X-080 was chosen as the extract solvent. Some important parameters affecting the CPE efficiency, such as the nature and concentration of surfactant, extraction temperature and time, centrifuge time and salt effect, were investigated and optimized. Separation was accomplished using a C(18) column by gradient elution with a acetonitrile-phosphate buffer solution as the mobile phase. The detection wavelength was set at 290 nm. Under optimum conditions, the linear range of ampelopsin in rat plasma was 20-2000 ng/mL (r(2)=0.9996). The limit of detection was 6 ng/mL (S/N=3) with the limit of quantification being 20 ng/mL (S/N=10). The proposed method has been successfully applied for pharmacokinetic studies of ampelopsin from rat plasma after oral administration.     

Micelle-mediated extraction and cloud-point preconcentration of osthole and imperatorin from Cnidium monnieri with analysis by high performance liquid chromatography

A new method based on micelle-mediated extraction and cloud-point preconcentration was developed for the separation and determination of hydrophobic compounds osthole and imperatorin from Cnidium monnieri by high performance liquid chromatography with photodiode array detection. The non-ionic surfactant C(13)E(8) (Genapol X-080) was chosen as the extract solvent. Various experimental conditions were investigated to evaluate and optimize the extraction and preconcentration process. The chromatographic separation was accomplished on a Zorbax SB-C(18) analytical column (150 mm x 4.6mm i.d., 5 microm particle diameter) maintained at 30 degrees C and detected by UV absorption at 320 nm. The gradient elution was achieved with a mobile phase composed of 0.1% phosphoric acid and acetonitrile at a flow rate of 1.0 mL min(-1). Under the optimum conditions, the calibration curve for both analytes was linear in the range of 0.52-33.5 microg mL(-1) with the correlation coefficients greater than 0.9996. The intra-day and inter-day precision (RSD) is below 5.3% and the limits of detection (LOD) for the analytes are 93 and 124 ng mL(-1)(S/N=3). The proposed technique is a low cost, simple and sensitive method with high clean-up effect. Finally, the method was successfully applied to separate and determine osthole and imperatorin from C. monnieri, respectively.     

Determination of Benzimidazole Fungicides by HPLC with Fluorescence Detection After Micellar Extraction

An analytical method was developed to determine the benzimidazole fungicides and their residues (benomyl, carbendazim, thiabendazole and fuberidazole) in real water samples. Analyses were performed by reverse phase (RP) HPLC with direct fluorescence detection with mobile phase methanol:water, 40:60 (v/v) with 0.6% (v/v) ammonia. The extraction of analytes from water samples was performed with the use of micellar systems. Specifically, oligoethylene glycol monoalkyl ether (Genapol X-080) and polyoxyethylene 10 lauryl ether (POLE) were used as extractants. The recoveries of fungicides obtained in spiked water samples ranged from 68% to 94% for Genapol and from 68% to 96% for POLE. The limit of detection (LOD) was lower than 6 g L^–1 for carbendazim, 7 g L^–1 benomyl, 0.15 g L^–1 for thiabendazole and 0.01 g L^–1 for fuberidazole in both surfactants.     

An approach to the application of microwave-assisted micellar extraction and liquid chromatography with ultraviolet detection to the extraction and determination of organophosphorus pesticides in tomato samples

Microwave-assisted micellar extraction methodology was applied to extract a mixture of 8 organophosphorus pesticides from the cuticle of tomato samples prior to analysis by liquid chromatography with ultraviolet detection. This technique provided very good results and was simple, fast, and environmentally friendly. The pesticides under study were extracted using the nonionic surfactants polyoxyethylene 10 lauryl ether (POLE) and oligoethylene glycol monoalkyl ether (Genapol X-080). The optimal extraction variables to be applied were determined for each surfactant and then compared. POLE proved to be the most suitable for the extraction, with recoveries over 70% in the majority and relative standard deviation values under 4.8%. After validation using a tomato sample enriched with a certified mixture, the proposed method was applied to the analysis of organophosphorus pesticides in lettuce and pepper samples.     

Determination of antihyperglycemic drugs in nanomolar concentration levels by micellar electrokinetic chromatography with non-ionic surfactant

A method for the separation of six selected antihyperglycemic (antidiabetic) drugs (tolbutamide, gliclazide, glimepiride, glibenclamide, repaglinide, and glipizide) was developed with use of micellar electrokinetic chromatography. Two non-ionic poly(ethylene glycol)-based surfactants Genapol X-080 and Triton X-114 (reduced) were studied as neutral pseudostationary phases. High alkaline pH 10.0 was used to obtain negative charges of separated antidiabetic drugs and non-ionic surfactants were employed for selectivity alteration. Both non-ionic surfactants provided good selectivity at concentration 0.2% (v/v) in sodium borate buffer and the separation of six drugs was obtained within 5 min. An on-line preconcentration method based on reversed electrode polarity switching was employed for the determination of antihyperglycemic drugs in blood serum after acetonitrile protein precipitation. The limits of detection ranged from 20.8 nmol L⁻¹ for tolbutamide to 6.5 nmol L⁻¹ for glibenclamide, respectively.     

Effect of short-chain alcohols on surfactant-mediated reversed-phase liquid chromatographic systems

The behaviour of β-blockers in a reversed-phase liquid chromatographic (RPLC) column with mobile phases containing a short-chain alcohol (methanol, ethanol or 1-propanol), with and without the surfactant sodium dodecyl sulphate (SDS), was explored. Two surfactant-mediated RPLC modes were studied, where the mobile phases contained either micelles or only surfactant monomers at high concentration. Acetonitrile was also considered for comparison purposes. A correlation was found between the effects of the organic solvent on micelle formation (monitored by the drop weight procedure) and on the nature of the chromatographic system (as revealed by the retention, elution strength and peak shape of β-blockers). When SDS is added to the mobile phase, the free surfactant monomers bind the C18 bonded chains on the stationary phase, forming an anionic layer, which attracts strongly the cationic β-blockers. The retention is modified as a consequence of the solving power of the organic solvent, micelles and surfactant monomers. The molecules of organic solvent bind the micelles, modify their shape, and may avoid their formation. They also bind the monomers of surfactant, desorbing them from the stationary phase, which affects the retention. The remaining surfactant covers the free silanols on the siliceous support, avoiding the interaction with the cationic solutes. The retention of β-blockers results from a combination of electrostatic and hydrophobic interactions, the latter being weaker compared to the hydro-organic system. The peak efficiencies and asymmetries are excellent tools to probe the surfactant layer on the stationary phase in an SDS/organic solvent system. The peaks will be nearly symmetrical wherever enough surfactant coats the stationary phase (up to 60% methanol, 40% ethanol, 35% 1-propanol, and 50% acetonitrile).     

Analysis of free amino acids in fermented shrimp waste by high-performance liquid chromatography

This work presents an HPLC method for the quantification of free amino acids in lyophilized protein fraction from shrimp waste hydrolysate which is obtained by acid lactic fermentation and analyzed using pre-column derivatization with 9-fluorenylmethyl-chloroformate. The amino acids were separated in a Hypersil ODS 5 microm column (250 mm x 4.6 mm) at 38 degrees C. The mobile phase was a mixture of phase A: 30 mM ammonium phosphate (pH 6.5) in 15:85 (v/v) methanol/water; phase B: 15:85 (v/v) methanol/water; and phase C: 90:10 (v/v) acetonitrile/water, with flow rate 1.2 ml/min. Fluorescence detection was used at an excitation wavelength of 270 nm and an emission wavelength of 316 nm. Method precisions for the different amino acids were between 4.4 and 7.1% (relative standard deviation, RSD); detection limits were between 23 and 72 ng/ml; and the recoveries were between 89.0 and 95.0%. The amino acid present at the highest concentration was tyrosine.     

Chiral separation of beta-adrenergic antagonists, profen non-steroidal anti-inflammatory drugs and chlorophenoxypropionic acid herbicides using teicoplanin as the chiral selector in capillary liquid chromatography

Three groups of structurally diverse chiral compounds were used to study the interaction mechanism responsible for stereoselective recognition with teicoplanin as chiral selector in capillary liquid chromatography. Teicoplanin-based chiral stationary phase (CSP) was used. The effect of the variation of mobile phase composition on retention and enantioselective separation was studied. The mobile phase composition suitable for enantioresolution of the various chiral compounds differed according to the interaction forces needed for chiral recognition. Mobile phases with high buffer portion (70-90 vol.%) were preferred for separation of enantiomers of profen non-steroidal anti-inflammatory drugs and chlorophenoxypropionic acid herbicides that require hydrophobic interactions, inclusion and pi-pi interactions for stereoselective recognition with teicoplanin. Higher concentration triethylamine in the buffer (0.5-1.0%) increased resolution of these acids. On the other hand, H-bonding and electrostatic interactions are important in stereoselective interaction mechanism of beta-adrenergic antagonists with teicoplanin. These interaction types predominate in the reversed phase separation mode with high organic modifier content (95% methanol) and in polar organic mobile phases. For this reason beta-adrenergic antagonists were best enantioresolved in the polar organic mode. The mobile phase composed of methanol/acetic acid/triethylamine, 100/0.01/0.01 (v/v/v), provided enantioresolution values of all the studied beta-adrenergic antagonists in the range 1.1-1.9. Addition of teicoplanin to the mobile phase, which was suitable for enantioseparation of certain compounds on the CSP, was also investigated. This system was used to dispose of nonstereoselective interactions of analytes with silica gel support that often participate in the interaction with CSPs. Very low concentration of teicoplanin in the mobile phase (0.1 mM) resulted in enantioselective separation of 2,2- and 2,4-chlorophenoxypropionic acids.     

High-performance liquid chromatographic determination of diuretics in urine by micellar liquid chromatography.

The use of micellar liquid chromatography for the determination of diuretics in urine by direct injection of the sample into the chromatographic system is discussed. The retention of the urine matrix at the beginning of the chromatograms was observed for different sodium dodecyl sulphate (SDS) mobile phases. The eluent strengths of a hybrid SDS-methanol micellar mobile phase for several diuretics were compared and related to the stationary phase/water partition coefficient with a purely micellar mobile phase. The urine band was appreciably narrower with a mobile phase of 0.05 M SDS-5% methanol (v/v) at 50 degrees C (pH 6.9). With this mobile phase the determination of bendroflumethiazide and chlorthalidone was adequate. Acetazolamide, ethacrynic acid, furosemide, hydrochlorothiazide and probenecid were overlapped by the urine matrix, and the retention of amiloride and triamterene was too long.     

Ultra-pressure liquid chromatography-electrospray tandem mass spectrometry for multiresidue determination of pesticides in water

A multiresidue analysis method has been developed for the determination of pesticides in water by ultra-performance liquid chromatography (UPLC) combined with tandem mass spectrometry (MS/MS). The selected pesticides represent a broad range of polarity and volatility [benzoylcyclohexanedione (mesotrione and sulcotrione); chloroacetamide (acetochlor, alachlor, dimethenamide, and metolachlor); phenoxyacetic acid (2,4-D and MCPA); phenoxypropionic (dichloprop and mecoprop); phenylurea (chlortoluron, diuron, isoproturon, linuron, and metoxuron); sulfonylurea (foramsulfuron, iodosulfuron, and nicolsulfuron); triazine (atrazine, cyanazine, desethylatrazine (DEA), desisopropylatrazine (DIA), simazine, and terbutylazine)]. The analytes were extracted using solid-phase extraction (SPE). The separation was carried out on an acquity UPLC BEH C18 column (1.7 microm, 50 mm x 1 mm ID) using a gradient elution profile and mobile phase consisting of 0.1% formic acid in water and acetonitrile. The pesticides were detected with a tandem mass spectrometer after being ionised positively or negatively (depending on the molecule) using an electrospray ionisation (ESI) source. To achieve the suitable extraction conditions for sample preparation, several parameters affecting the efficiency of SPE such as the nature of the sorbent and the eluent, extractant volume and pH were studied. The best recovery was obtained by the extraction with an Oasis HLB cartridge and 3 mL of a solution of acetonitrile/dichloromethane (1:1, v/v) at pH 2. The average recoveries of the pesticides in different samples ranged from 82 to 109%. The weight least squares (WLS) linear regression was used to calculate the limits of detection and quantification (LOD and LOQ) because the dispersion was heteroskedastic. All the pesticides could be correctly quantified at a concentration level of 50 ng L(-1) and most of them could be detected at a concentration inferior or equal to 8 ng L(-1). Efficiency and robustness of this method were evaluated by the analysis of several samples of real natural water.     

Combination of micellar extraction of polycyclic aromatic hydrocarbons from aqueous media with detection by synchronous fluorescence

Summary The application of different non-ionic surfactants for the micellar extraction and enrichment of PAHs from aqueous media was tested. Recoveries were up to 100%. A spectroscopic method for the simultaneous detection of PAH-mixtures by synchronous fluorescence in the micellar phase was developed, with detection limits of 6.8 and 2.6 ng/l for benzo(k)fluoranthene and benzo(a)pyrene, respectively. The method was applied to the extraction and detection of benzo(k)fluoranthene/benzo(a)pyrene mixtures from aqueous solutions and soil suspensions. Genapol X-80 was found to suppress PAH-adsorption on bentonite at surfactant concentrations above 0.1%.Dedicated to Prof. Dr. V. Krivan on the occasion of his 60th birthday.     

Effect of the ionic strength of salts on retention and overloading behavior of ionizable compounds in reversed-phase liquid chromatography I. XTerra-C18

The influence of the salt concentration (potassium chloride) on the retention and overloading behavior of the propranolol cation (R'-NH2+ -R) on an XTerra-C18 column, in a methanol:water solution, was investigated. The adsorption isotherm data were first determined by frontal analysis (FA) for a mobile phase without salt (25% methanol, v/v). It was shown that the adsorption energy distribution calculated from these raw adsorption data is bimodal and that the isotherm model that best accounts for these data is the bi-Moreau model. Assuming that the addition of a salt into the mobile phase changes the numerical values of the parameters of the isotherm model, not its mathematical form, we used the inverse method (IM) of chromatography to determine the isotherm with seven salt concentrations in the mobile phase (40% methanol, v/v; 0, 0.002, 0.005, 0.01, 0.05, 0.1 and 0.2 M). The saturation capacities of the model increase, q(s,1) by a factor two and q(s,2) by a factor four, with increasing salt concentration in the range studied while the adsorption constant b1 increases four times and b2 decreases four times. Adsorbate-adsorbate interactions vanish in the presence of salt, consistent with results obtained previously on a C18-Kromasil column. Finally, besides the ionic strength of the solution, the size, valence, and nature of the salt ions affect the thermodynamic as well as the mass transfer kinetics of the adsorption mechanism of propranolol on the XTerra column.     

Retention of ionizable compounds on high-performance liquid chromatography. VII. Characterization of the retention of ionic solutes in a C18 column by mass spectrometry with electrospray ionization

The elution of ions from a C₁₈ column with mobile phases containing methanol (60%, v/v) and aqueous buffers is studied by mass spectrometry. It is demonstrated that the anions are excluded from the stationary phase by the ionized silanols. However, the ionized silanols interact strongly with cations, which are retained in the column. These cations are later eluted from the column by ion exchange with the cations present in the pH buffered mobile phase. The size of the ions, the mobile phase cation concentration and the mobile phase pH are the main parameters that affect elution of the retained cations. It is also demonstrated that there are at least two different types of ionizable silanols, with different acidities, that contribute to the retention of cations. An estimate of the pK_a values of these two groups of silanols in 60% methanol is given.     

Quantitative determination of clenbuterol enantiomers in human plasma by high-performance liquid chromatography using the macrocyclic antibiotic chiral stationary phase teicoplanin

We report a method for the high-performance liquid chromatographic (HPLC) chiral separation of racemic clenbuterol in human plasma. Human plasma was spiked with stock solutions of clenbuterol hydrochloride and practolol as the internal standard. Following a liquid-liquid extraction procedure with 10% (+/-)-2-butanol/isopropyl ether under alkaline conditions, the dried samples were reconstituted in methanol and chromatographed using a macrocyclic antibiotic chiral stationary phase (CSP) known as Chirobiotic T(trade mark) (teicoplanin). The mobile phase composition was methanol:acetonitrile (70:30, v/v), containing 0.3% (v/v) acetic acid and 0.2% (v/v) triethylamine. The resulting chromatogram achieved baseline separation for the clenbuterol enantiomers. Calibration curves (peak area ratio vs plasma concentration, n = 10) were constructed for the (-)-R-and (+)-S-clenbuterol enantiomers with a plasma concentration range of 0. 25-10 microM. The correlation coefficient (r) range was 0.99988-0. 99999 (mean = 0.99999). The lowest concentration measured was 0.25 microM. Inter- and intra-assay variation was determined for the lowest, medium and highest plasma concentration (0.25, 2 and 10 microM) by calculating the analytical recoveries with a range of 96-104%. The percentage recoveries for the clenbuterol enantiomers were 88.4-102% over the concentration range used. Detailed methodology is presented.     

Influence of solvent effects on retention in reversed-phase liquid chromatography and solid-phase extraction using a cyanopropylsiloxane-bonded,silica-based sorbent

Summary The solvation parameter model is used to characterize the retention properties of a cyanopropylsiloxanebonded, silica-based sorbent with methanol, acetonitrile, tetrahydrofuran, and isopropanol in water as mobile phases. The system constants over the composition range 1 to 50 % (v/v) organic solvent indicate that retention occurs because of the relative ease of cavity formation in the solvated stationary phase compared to the same process in the predominantly aqueous mobile phase as well as from more favorable stationary phase interactions with solutes containing π- and n-electrons. The capacity of the solute for dipole-type interactions is not important whereas all hydrogen-bond-type interactions result in reduced retention. Graphing the system constants as a function of mobile phase composition provides a simple mechanism for interpreting the change in capacity of the chromatographic system for retention in terms of changes in the relative weighting of fundamental intermolecular interactions. A comparison is also made with the retention properties of an octadecylsiloxane-bonded, silica-based sorbent with 30 % (v/v) methanol in water as the mobile phase and the extraction characteristics of a porous polymer sorbent with 1 % (v/v) methanol, acetonitrile, tetrahydrofuran, and isopropanol in water as the sample processing solvent. Changes in sorbent selectivity due to selective uptake of the processing solvent are much smaller for the cyanopropylsiloxane-bonded sorbent than the results found for a porous polymer sorbent.