Determination of clofentezine in medical herb extracts by chromatographic methods combined with diode array scanning densitometry

The aim of this study is to report a new procedure for extraction, cleanup and determination of clofentezine in herb extracts by ultrasound‐assisted solvent extraction, SPE and multidimensional planar chromatography with diode array detector (MDPC‐DAD) and/or HPLC‐DAD. The application of various extraction solvents in SPE experiments conducted on octadecyl silane coupled with styrene‐divinylbenzene cartridges for fractionation and purification samples has been described. Normal‐phase systems were used in MDPC experiments on silica layer. The procedure described for the determination of compounds is inexpensive and can be applied to the routine analysis of analytes in plant extracts, after preliminary cleanup and concentration, e.g. by SPE. Application of MDPC‐DAD and HPLC‐DAD is especially useful for correct identification of components of difficult, complicated mixtures, e.g. analytes in medical herbs.     

Determination of four sulfonylurea herbicides in tea by matrix solid‐phase dispersion cleanup followed by dispersive liquid–liquid microextraction

Matrix solid‐phase dispersion combined with dispersive liquid–liquid microextraction has been developed as a new sample pretreatment method for the determination of four sulfonylurea herbicides (chlorsulfuron, bensulfuron‐methyl, chlorimuron‐ethyl, and pyrazosulfuron) in tea by high‐performance liquid chromatography with diode array detection. The extraction and cleanup by matrix solid‐phase dispersion was carried out by using CN‐silica as dispersant and carbon nanotubes as cleanup sorbent eluted with acidified dichloromethane. The eluent of matrix solid‐phase dispersion was evaporated and redissolved in 0.5 mL methanol, and used as the dispersive solvent of the following dispersive liquid–liquid microextraction procedure for further purification and enrichment of the target analytes before high‐performance liquid chromatography analysis. Under the optimum conditions, the method yielded a linear calibration curve in the concentration range from 5.0 to 10 000 ng/g for target analytes with a correlation coefficients (r²) ranging from 0.9959 to 0.9998. The limits of detection for the analytes were in the range of 1.31–2.81 ng/g. Recoveries of the four sulfonylurea herbicides at two fortification levels were between 72.8 and 110.6% with relative standard deviations lower than 6.95%. The method was successfully applied to the analysis of four sulfonylurea herbicides in several tea samples.     

Development and validation of a liquid chromatography method with electrospray ionization tandem mass spectrometry for the determination of brotizolam residues in beef and commercial whole milk

In this work, a liquid chromatography–tandem mass spectrometric detection technique was developed and validated for the determination of brotizolam residues in beef muscle and commercial whole milk. This procedure involves the extraction of the analyte from the samples via liquid–solid extraction, and caffeine was used as an internal standard. The analyte was successfully separated on an XTerra‐C₁₈ column, with a mobile phase composed of 0.01% formic acid in acetonitrile and 1 mm ammonium formate–0.01% formic acid in water. The one‐step extraction method evidenced good selectivity, precision (RSD = 9.87–26.47%), and the recovery of the extractable analyte was 92.61–115.98% in the matrices. The limits of quantification ranged between 0.4 and 0.5 µg/kg. The developed method is simple since it requires no additional cleanup procedures. Copyright © 2011 John Wiley & Sons, Ltd.     

Residue analysis of tebufenozide and indoxacarb in chicken muscle,milk, egg and aquatic animal products using liquid chromatography–tandem mass spectrometry

We developed an analytical method using liquid–liquid extraction (LLE) and liquid chromatography–tandem mass spectrometry (LC‐MS/MS) to detect and quantify tebufenozide (TEB) and indoxacarb (IND) residues in animal and aquatic products (chicken muscle, milk, egg, eel, flatfish, and shrimp). The target compounds were extracted using 1% acetic acid (0.1% acetic acid for egg only) in acetonitrile and purified using n‐hexane. The analytes were separated on a Gemini‐NX C₁₈ column using (a) distilled water with 0.1% formic acid and 5 mm ammonium acetate and (b) methanol with 0.1% formic acid as the mobile phase. All six‐point matrix‐matched calibration curves showed good linearity with coefficients of determination (R²) ≥0.9864 over a concentration range of 5–50 μg/kg. Intra‐ and inter‐day accuracy was expressed as the recovery rate at three spiking levels and ranged between 73.22 and 114.93% in all matrices, with a relative standard deviation (RSD, corresponding to precision) ≤13.87%. The limits of quantification (LOQ) of all target analytes ranged from 2 to 20 μg/kg, which were substantially lower than the maximum residue limits (MRLs) specified by the regulatory agencies of different countries. All samples were collected from different markets in Seoul, Republic of Korea, and tested negative for tebufenozide and indoxacarb residues. These results show that the method developed is robust and may be a promising tool to detect trace levels of the target analytes in animal products.     

Simultaneous extraction and determination of antibiotics in soils using a method based on quick,easy, cheap,effective, rugged,and safe extraction and liquid chromatography with tandem mass spectrometry

The increasing use of antibiotics has caused substantial environmental problems, which are a matter of great concern. The aim of this work was to develop a quick, easy, cheap, effective, rugged, and safe method for 20 antibiotic residues in soil. The developed method is based on extraction with acetonitrile and phosphate buffer, clean up with dispersive solid‐phase extraction adsorbent using primary secondary amine, octadecylsilane, followed by liquid chromatography with tandem mass spectrometry determination. We optimized different extraction methods and the ratio of cleanup adsorbents to achieve good recoveries at seven spiking levels that ranged from 61.4 to 118.9% with a relative standard deviation below 20% (n  = 5). The method quantification limit was in the range of 2–5 μg/kg for most analytes. Good linear regression coefficients greater than 0.990 were obtained. This method was applied for the analysis of real agricultural soil samples, confirming the feasibility of the method.     

Simple and rapid determination of N6‐(Δ2‐isopentenyl)adenine,zeatin, and dihydrozeatin in plants using on‐line cleanup liquid chromatography coupled with hybrid quadrupole‐Orbitrap high‐resolution mass spectrometry

A simple and rapid method was developed for the determination of three free cytokinins, namely, N⁶‐(Δ²‐isopentenyl)adenine, zeatin, and dihydrozeatin, in plants using TurboFlow on‐line cleanup liquid chromatography combined with hybrid quadrupole‐Orbitrap high‐resolution mass spectrometry. The samples were extracted using acetonitrile, and then the extract was purified on a C₁₈‐p column, in which the sample matrix was removed and the analytes were retained. Subsequently, the analytes were eluted from the extraction column onto the analytical column (Hypersil Gold C₁₈ column) prior to chromatographic separation and hybrid Q‐Orbitrap detection using the targeted‐MS² scan mode. The linearity was satisfactory with a correlation coefficient of >0.999 at concentrations ranging from 5–5000 pg/mL. The limits of quantification for the analytes ranged from 4.2–5.2 pg/mL. The intra‐ and inter‐day average recoveries of analytes fortified at three levels ranged from 85.4–108.2%, and the intra‐ and inter‐day relative standard deviations ranged from 4.04–8.57%. The method was successfully applied for the determination of free cytokinins in different tissue samples of Oryza sativa and Arabidopsis thaliana.     

Single‐step multiresidue determination of ten multiclass veterinary drugs in pork,milk, and eggs using liquid chromatography with tandem mass spectrometry

A multiclass, multiresidue determination method is reported for the detection of ten veterinary drugs, including scopolamine, metoclopramide, acriflavine, berberine, tripelennamine, diphenhydramine, acrinol, triamcinolone, loperamide, and roxithromycin in pork, milk, and eggs. The method involves a simple extraction using 0.1% formic acid in acetonitrile, followed by defatting with n‐hexane, centrifugation, and filtration prior to liquid chromatography with tandem mass spectrometric analysis. As ion suppression and enhancement effects are reported, matrix‐matched calibrations are used for quantification, with determination coefficients ≥0.9765. For the majority of the tested analytes, the intra‐ and interday accuracy (expressed as recovery %) range from 70.6 to 94.6% and from 70.1 to 93.3%, respectively, and the precision (expressed as relative standard deviation) ranges from 0.5 to 19.8% and from 2.8 to 18.4% in all matrices. The limits of quantification range between 0.5 and 10 ng/g. The validated tandem mass spectrometry method is successfully applied to market samples; the target analytes are not detected in any of the tested samples. In terms of accuracy, no extract cleanup is deemed necessary. The developed method is feasible for the simultaneous detection of the tested analytes in pork, milk, and eggs.     

Combination of ultracentrifugation and solid‐phase extraction with subsequent chromatographic analysis of α‐tocopherol in erythrocyte membranes

A novel and rapid sample pretreatment technique based on a combination of ultracentrifugation and solid‐phase extraction for the determination of α‐tocopherol in human erythrocyte membranes by high‐performance liquid chromatography with ultraviolet detection is presented in this work. Red blood cell samples were ultracentrifuged (288 000 × g, 3 min, 4°C) in the presence of d ‐mannitol, 4‐(2‐hydroxyethyl)‐1‐piperazineethanesulfonic acid and calcium chloride. The α‐tocopherol was then extracted from the erythrocyte membranes by solid‐phase extraction with n‐hexane in the presence of ascorbic acid. Tocopherol acetate was used as the internal standard. The extract was dissolved in methanol and separated on the monolithic column Chromolith Performance RP‐18e (100 × 4.6 mm) using 100% methanol as the mobile phase. The absorbance of α‐tocopherol was measured at a wavelength of 295 nm. The method was validated and showed sufficient accuracy and precision, ranging from 96.4 to 100.8% and from 4.5 to 6.3%, respectively. Moreover, the developed method was applied to the determination of erythrocyte α‐tocopherol in real samples from patients. The combined ultracentrifugation and solid‐phase extraction technique substantially decreased the time for the sample pretreatment step compared to liquid–liquid extraction and could be applicable for the quantitation of other analytes in erythrocyte membranes.     

GC�CMS Determination of PAHs in Fish Samples Following Salting-out-Assisted Solvent Extraction-Gel Permeation Chromatography

A fast and effective sample cleanup procedure for the quantification of polycyclic aromatic hydrocarbons (PAHs) in fish samples is presented. The procedure involved extraction of fish samples using acetonitrile and cleanup by an automated gel-permeation chromatography (GPC) following liquid?Cliquid partition into n-hexane. The extracted samples were analyzed by gas chromatography-mass spectrometry (GC?CMS). Electron ionization was employed in a single analysis for the determination of PAHs in the selected ion monitoring mode. Three different solvents were studied for the extraction step: acetonitrile/n-hexane, methanol/n-hexane and acetone/n-hexane. The best solvent was found to be acetonitrile/n-hexane. The cleanup technique resulted in a good separation of analytes from co-extractive matrix compounds.     

Simultaneous and rapid determination of deoxynivalenol and its acetylate derivatives in wheat flour and rice by ultra high performance liquid chromatography with photo diode array detection

A simple and reliable method of ultra high performance liquid chromatography coupled with photo‐diode array detection has been proposed for the simultaneous determination of deoxynivalenol and its acetylated derivatives in wheat flour and rice, especially focusing on the optimization of sample extraction, cleanup, and chromatographic separation conditions. Sample pretreatment consisted of a first step using a quick, easy, cheap, effective, rugged, and safe based extraction procedure and a subsequent cleanup step based on solid‐phase extraction. The method was extensively validated in wheat flour and rice, obtaining satisfactory analytical performance with good linearity (R² ≥ 0.999), acceptable recoveries (80.0–104.4%), and repeatability (RSDs 1.3–10.7%). The limits of detection (21.7–57.4 μg/kg) and quantitation (72.3–191.4 μg/kg) for deoxynivalenols were lower than those usually permitted by various countries’ legislation in these food matrices. The method was applied to 34 wheat and rice samples. The results were further compared with results of ultra high performance liquid chromatography with electrospray ionization tandem mass spectrometry.     

Magnetic mixed hemimicelles solid‐phase extraction coupled with high‐performance liquid chromatography for the extraction and rapid determination of six fluoroquinolones in environmental water samples

In this study, a mixed hemimicelle solid‐phase extraction method based on Fe₃O₄ nanoparticles coated with sodium dodecyl sulfate was applied for the preconcentration and fast isolation of six fluoroquinolones in environmental water samples before high‐performance liquid chromatography determination. The main factors affecting the extraction efficiency of the analytes, such as amount of surfactant, amount of Fe₃O₄ nanoparticles, extraction time, sample volume, sample pH, ionic strength, and desorption conditions, were investigated and optimized. The method has detection limits from 0.05 to 0.1 ng/mL and good linearity (r ≥ 09948) in the range 0.1–200 ng/mL depending on the fluoroquinolone. The enrichment factor is ～200. The recoveries (at spiked levels of 1, 5, and 50 ng/mL) are in the range of 79–120%.     

Nano‐structured gemini‐based supramolecular solvent for the microextraction of cyhalothrin and fenvalerate

A novel supramolecular solvent‐based microextraction followed by high‐performance liquid chromatography with ultraviolet detection method has been developed for the extraction and determination of two pyrethroid analytes, cyhalothrin and fenvalerate, in water and soil samples. The liquid–liquid‐phase separation of surfactants has been used in analytical extraction. The surfactant‐rich phase is a nano‐structured liquid, recently named as a supramolecular solvent, generated from the amphiphiles. The alkyl carboxylic acid based supramolecular solvents were introduced before. Coacervates made up of gemini surfactant, consisting of two amphiphilic moieties, were first used as solvent. The effective parameters on extraction (i.e., type of organic solvent, the amount of surfactant and volume of tetrahydrofuran, sample solution pH, salt addition, ultrasonic and centrifugation time) were investigated and optimized. Under the optimum conditions, preconcentration factors of 110 and 145 were obtained for the analytes. The linearity was 0.5–200.0 μg/L with the correlation of determination of (R²) ≥ 0.9984. The limit of detection of the method was (S/N = 3) 0.2 μg/L, and precisions in the range of 6.3–10.3% (RSDs, n = 5) were obtained. This method has been successfully applied to analyze real samples, and good recoveries in the range of 101.2–108.8% were obtained.     

Ultrasound‐assisted extraction and solid‐phase extraction as a cleanup procedure for organochlorinated pesticides and polychlorinated biphenyls determination in aquatic samples by gas chromatography with electron capture detection

The feasibility of developing a quick, easy, efficient procedure for the simultaneous determination of organochlorinated pesticides and polychlorinated biphenyls in aquatic samples using gas chromatography with electron capture detection based on solid‐phase extraction was investigated. The extraction solvent (n‐hexane/acetone, cyclohexane/ethyl acetate, n‐hexane/dichloromethane, n‐hexane) for ultrasound‐assisted solid–liquid extraction and solid‐phase extraction columns (florisil, neutral alumina, acidic alumina, aminopropyl trimethoxy silane, propyl ethylenediamine, aminopropyl trimethoxy silane/propyl ethylenediamine, graphitized carbon black and silica) for cleanup procedure were optimized. The gas chromatography with electron capture detection method was validated in terms of linearity, sensitivity, reproducibility, and recovery. Mean recoveries ranged from 75 to 115% with relative standard deviations <13%. Quantification limits were 0.20–0.40 ng/g for organochlorinated pesticides and polychlorinated biphenyls. The satisfactory data demonstrated the good reproducibility of the method with relative standard deviations lower than 13%. In comparison to other related methods, this method requires less time and solvent and allows for rapid isolation of the target analytes with high selectivity. This method therefore allows for the screening of numerous samples and can also be used for routine analyses.     

Determination of Phenolics in Water and Arthrospira (Spirulina) platensis by Concentrated Sulfuric Acid and Ultrasound-Assisted Surfactant-Enhanced Emulsification Microextraction and High Performance Liquid Chromatography

A novel, practical, and environmentally friendly technique, termed concentrated sulfuric acid cleanup and ultrasound-assisted surfactant-enhanced emulsification microextraction (CSAC-UASEM), was combined with HPLC for the preconcentration and determination of five phenolics in water and Arthrospira (Spirulina) platensis samples. The main advantages include that the concentrated sulfuric acid is used to decrease macromolecular interferences prior to microextraction and, unlike dispersive liquid–liquid microextraction procedures, no dispersive organic solvent is required. Chloroform and sodium dodecyl sulfate were used as the extraction solvent and emulsifier, respectively. The algal cell preparation and CSAC-UASEM procedure parameters, including selection of cleanup method, ultrasound power, cell cytocylasis time, type and volume of extraction solvent, extraction temperature, ultrasound-extracted time, and sample pH, were optimized. At the fortification levels of 1.0 and 10.0 µg/L, the enrichment factors of analytes were in the range of 201.38 to 269.24. The percent extraction ranged from 71.57% to 107.42% in environmental Arthrospira-350, -793, and -834 samples, whereas the range was from 74.17% to 106.72% in water samples. The limits of detection (at S/N = 3) were 0.02 to 0.04 µg/L (except for 4-bromobisphenol A of 0.10 µg/L). These values indicate an approximately ten-fold improvement compared with the values reported by other techniques. In summary, the CSAC-UASEM sample preparation technique has great potential in the routine determination of trace phenolics in environmental waters and aquatic biological samples.     

Validated UHPLC–MS/MS assay for quantitative determination of etoposide,gemcitabine, vinorelbine and their metabolites in patients with lung cancer

A fully valid UHPLC–MS/MS method was developed for the determination of etoposide, gemcitabine, vinorelbine and their metabolites (etoposide catechol, 2′,2′‐difluorodeoxyuridine and 4‐O ‐deacetylvinorelbine) in human plasma. The multiple reaction monitoring mode was performed with an electrospray ionization interface operating in both the positive and negative ion modes per compound. The method required only 100 μL plasma with a one‐step simple de‐proteinization procedure, and a short run time of 7.5 min per sample. A Waters ACQUITY UPLC HSS T3 column (2.1 × 100 mm, 1.8 μm) provided chromatographic separation of analytes using a binary mobile phase gradient (A, 0.1% formic acid in acetonitrile, v /v; B, 0.1% formic acid in water, v /v). Linear coefficients of correlation were >0.995 for all analytes. The relative deviation of this method was <10% for intra‐ and inter‐day assays and the accuracy ranged between 86.35% and 113.44%. The mean extraction recovery and matrix effect of all the analytes were 62.07–105.46% and 93.67–105.87%, respectively. This method was successfully applied to clinical samples from patients with lung cancer.     

Dispersive solid‐phase extraction followed by vortex‐assisted dispersive liquid–liquid microextraction based on the solidification of a floating organic droplet for the determination of benzoylurea insecticides in soil and sewage sludge

A novel dispersive solid‐phase extraction combined with vortex‐assisted dispersive liquid–liquid microextraction based on solidification of floating organic droplet was developed for the determination of eight benzoylurea insecticides in soil and sewage sludge samples before high‐performance liquid chromatography with ultraviolet detection. The analytes were first extracted from the soil and sludge samples into acetone under optimized pretreatment conditions. Clean‐up of the extract was conducted by dispersive solid‐phase extraction using activated carbon as the sorbent. The vortex‐assisted dispersive liquid–liquid microextraction based on solidification of floating organic droplet procedure was performed by using 1‐undecanol with lower density than water as the extraction solvent, and the acetone contained in the solution also acted as dispersive solvent. Under the optimum conditions, the linearity of the method was in the range 2–500 ng/g with correlation coefficients (r) of 0.9993–0.9999. The limits of detection were in the range of 0.08–0.56 ng/g. The relative standard deviations varied from 2.16 to 6.26% (n = 5). The enrichment factors ranged from 104 to 118. The extraction recoveries ranged from 81.05 to 97.82% for all of the analytes. The good performance has demonstrated that the proposed methodology has a strong potential for application in the multiresidue analysis of complex matrices.     

Development of multiple monolithic fiber solid‐phase microextraction and liquid chromatography–tandem mass spectrometry method for the sensitive monitoring of aminoglycosides in honey and milk samples

A simple and sensitive method for the simultaneous extraction and determination of six aminoglycosides in honey and milk samples was developed using multiple monolithic fiber solid‐phase microextraction and liquid chromatography with tandem mass spectrometry. The multiple monolithic fibers based on poly(methacrylic acid‐co‐ethylenedimethacrylate) monolith as the extraction medium was used to concentrate target analytes. Because there were abundant carboxyl groups in the monolith, the monolithic fibers could extract aminoglycosides effectively through cation‐exchange and hydrophobic interactions. To obtain optimum extraction performance, several extraction parameters including desorption solvent, adsorption and desorption time, pH value and ionic strength in sample matrix, were investigated in detail. Under the optimized extraction conditions, the limits of detection of the proposed method were 0.10–0.30 and 0.23–0.59 μg/kg for honey and milk samples, respectively. Satisfactory linearity was achieved for analytes with the coefficients of determination above 0.99. At the same time, the developed method showed acceptable method repeatability and reproducibility. Finally, the proposed method was successfully applied to the determination of aminoglycosides in real honey and milk samples. Recoveries obtained for the determination of six target analytes in spiking samples ranged from 67.9 to 110%, and the relative standard deviations were in the range of 1.2–11%.     

A fast,sensitive, and high‐throughput method for the simultaneous quantitation of three ellagitannins from Euphorbiae pekinensis Radix in rat plasma by ultra‐HPLC–MS/MS

A fast, sensitive, and high‐throughput ultra‐HPLC–MS/MS method has been developed and validated for the simultaneous determination of three main active constituents of Euphorbiae pekinensis Radix in rat plasma. After addition of the internal standard, plasma samples were extracted by liquid–liquid extraction with ethyl acetate/isopropanol (1:1, v/v) and separated on a CAPCELL PAK C₁₈ column (100 × 2.0 mm, 2 μm, Shiseido, Japan), using a gradient mobile phase system of methanol/water. The detection of the analytes was performed on a 4000Q UHPLC–MS/MS system with turbo ion spray source in the negative ion and multiple reaction‐monitoring mode. The linear range was 1.0–1000 ng/mL for 3,3′‐di‐O‐methyl ellagic acid‐4′‐O‐β‐d ‐glucopyranoside (i), 1.5–1500 ng/mL for 3,3′‐di‐O‐methyl ellagic acid‐4′‐O‐β‐d ‐xylopyranoside (ii), and 5.0–5000 ng/mL for 3,3′‐di‐O‐methyl ellagic acid (iii). The intra‐ and interday precision and accuracy of all the analytes were within 15%. The extraction recoveries of the three analytes and internal standard from plasma were all more than 80%. The validated method was first successfully applied to the evaluation of pharmacokinetic parameters of compounds 1 , 2 , and 3 in rat plasma after intragastric administration of the Euphorbiae pekinensis Radix extract.     

Enrichment and low‐level determination of glyphosate,aminomethylphosphonic acid and glufosinate in drinking water after cleanup by cation exchange resin

For the determination of glyphosate, aminomethylphosphonic acid and glufosinate in drinking water, different procedures of enrichment and cleanup were examined using anion exchange or SPE. In many cases interactions of, e.g. alkaline earth metal ions especially calcium could be observed during enrichment and cleanup resulting in loss of analytes. For that reason, a novel cleanup and enrichment procedure for the determination of these phosphonic acid herbicides has been developed in drinking water using cation‐exchange resin. In summary, the cleanup procedure with cation‐exchange resin developed in this study avoids interactions as described above and is applicable to calcium‐rich drinking water samples. After derivatization with 9‐fluorenylmethylchloroformate followed by LC with fluorescence detection, LOD of 12, 14 and 12 ng/L and mean recoveries from real‐world drinking water samples of 98±9, 100±16 and 101±11% were obtained for glyphosate, aminomethylphosphonic acid and glufosinate, respectively. The low LODs and the high precision permit the analysis of these phosphonic acid herbicides according to the guidelines of the European Commission.     

Rapid selective accelerated solvent extraction and simultaneous determination of herbicide atrazine and its metabolites in fruit by ultra high performance liquid chromatography

A selective accelerated solvent extraction procedure achieved one step extraction and cleanup for analysis of herbicide atrazine and its metabolites in fruit. Using a BEH C18 analytical column and the gradient mode with 2 mM ammonium acetate aqueous solution/acetonitrile as a mobile phase achieved effective chromatographic separation of the five analytes within 4 min. The calibration curves were linear over two orders of magnitude of concentration with correlation coefficients (r) of 0.9996?0.9999. The method limit of quantification was 1, 2, 1.5, 3, and 2 μg/kg for atrazine, desethylatrazine, desisopropylatrazine, desethyldesisopropylatrazine, and hydroxyatrazine, respectively, in the case of atrazine it is at least two orders of magnitude lower than the maximum residue limit (0.25 mg/kg). The intra‐day and inter‐day precisions of the five analytes were in the range of 2.1–3.5 and 3.1–4.8 %, respectively. The recoveries of the five analytes at three spiked levels varied from 85.9 to 107% with a relative standard deviation of 1.8–4.9% for pear and apple samples. The ultra high performance liquid chromatography with diode array detection method was proved to be fast, inexpensive, selective, sensitive, and accurate for the quantification of the analytes in pear and apple samples.