Solid Phase Extraction – Voltammetric Coupled Detection of Caffeine in Acetonitrile

Caffeine (CAF) in aqueous solutions was extracted into acetonitrile (MeCN) using solid phase extraction (SPE). The voltammetric detection of CAF in MeCN was performed in a microcentrifuge tube after SPE in order to minimise the required amount of MeCN and reagents used during the analysis. The voltammetric determination of CAF from a range of beverages extracted into MeCN was in good agreement with the results obtained from HPLC measurements on the same samples. The SPE‐coupled voltammetric method gave more accurate results than those obtained via the direct voltammetric detection of CAF in real beverage samples.     

植物激素的反相高效液相色谱法分离和测定

探讨了分离和测定８种植物生长激素的反相高效液相色谱法。结果表明：以含有０．８％醋酸的甲醇和水作为流动相,采用梯度洗脱,可在１６ｍｉｎ内将８种物质完全分离并可进行定量测定,线性关系良好。方法具有快速、简便、准确的特点。     

Comparison of different HPLC systems for analysis of galantamine and lycorine in various species of Amaryllidaceae family

Retention parameters of galantamine and lycorine standards were determined on different columns, i.e., octadecyl silica, SM C18, and strong cation-exchange (SCX) columns with different aqueous mobile phases. Retention of alkaloids was investigated on C18, SM C18 columns with mobile phase containing 5% MeCN, 20% acetate buffer at pH 3.5, and 0.025 ML^?1 diethylamine (DEA), and on SCX column with mobile phase containing 8% MeCN and phosphate buffer at pH 2.5. Better results were also obtained in ion-exchange chromatographic system. On the basis of results obtained in different chromatographic systems, simple, rapid, and sensitive high-performance liquid chromatography methods were developed for determining lycorine and galantamine in plant extracts from various species belonging to Amaryllidaceae family. Extracts were prepared from various parts of plants collected at different times of the growing season.     

Simultaneous Analysis Of Estradiol Dienanthate,Estradiol 3-Benzoate And Testosterone Enanthate Benzilic Acid Hydrazone In Oily Formulations Bygradient-Hplc

Abstract

A procedure is described for the analysis of estradiol-3-benzoate, testosterone enanthate benzylic acid hydrazone and estradiol dienanthate in oily solution. The multi-step gradient reversed phase ion-pairing HPLC method uses 1-pentane sulfonic acid sodium salt (0.0025 M in 0.1% acetic acid methanolic solution): acetonitrile and water (55 : 30 to 45 : 15 to 0) mobile phase gradient and a 250 × 4.6 mm, 5 micron octadecyl silane column, with 1,2,4,5-tetrachlorobenzene as internal standard. The time required for chromatography is 31 minutes. The method gives relative standard deviations (RSD) of .94% or better for the assay of active ingredients in commercial formulations.     

Rapid determination of 95 pesticides in soybean oil using liquid-liquid extraction followed by centrifugation, freezing and dispersive solid phase extraction as cleanup steps and gas chromatography with mass spectrometric detection

A multi-residue method using liquid-liquid extraction (LLE) followed by centrifugation, freezing and dispersive solid phase extraction (dispersive SPE) as clean up steps and gas chromatography with mass spectrometric detection has been developed for the determination of trace levels of 95 pesticides in soybean oil. LLE has been optimized to extract these pesticide residues from soybean oil by studying the effect of different partitions between (i) acetonitrile (MeCN) saturated with petroleum ether and a soybean oil solution dissolved in petroleum ether saturated with MeCN, (ii) partition between MeCN and a soybean oil solution dissolved in petroleum ether saturated with MeCN, (iii) partition between MeCN and a soybean oil solution dissolved in petroleum ether, (iv) partition between MeCN saturated with n-hexane and a soybean oil solution dissolved in n-hexane saturated with MeCN, (v) partition between MeCN and a soybean oil solution, (vi) partition between MeCN and a soybean oil solution dissolved in n-hexane and (vii) partition between MeCN and a soybean oil solution dissolved in mixture of acetone and n-hexane (3:2) to the highest recovery yield of pesticides and the lowest co-extract fat residue in the final extract. Experiments were carried out in order to study the efficiency of using centrifugation and freezing steps as well as the used of primary secondary amine (PSA), florisil, graphite carbon black (GCB) and C18 for dispersive SPE on clean up stages to minimize the co-extract fat. The recoveries obtained ranged from 80 to 114% and the relative standard deviation (RSDs) from 2 to 14% for spiking levels of 0.040, 0.080 and 0.160 mg kg^− 1. The limits of quantification (LOQs) of almost all compounds were below the maximum residue limits (MRLs) established by the Korean legislations for soybean oil.     

Use of buffering and other means to improve results of problematic pesticides in a fast and easy method for residue analysis of fruits and vegetables

A modification that entails the use of buffering during extraction was made to further improve results for certain problematic pesticides (e.g., folpet, dichlofluanid, chlorothalonil, and pymetrozine) in a simple, fast, and inexpensive method for the determination of pesticides in produce. The method, known as the quick, easy, cheap, effective, rugged, and safe (QuEChERS) method for pesticide residues in foods, now involves the extraction of the sample with acetonitrile (MeCN) containing 1% acetic acid (HAc) and simultaneous liquid-liquid partitioning formed by adding anhydrous MgSO4 plus sodium acetate (NaAc). The extraction method is carried out by shaking a centrifuge tube which contains 1 mL of 1% HAc in MeCN plus 0.4 g anhydrous MgSO4 and 0.1 g anhydrous NaAc per g sample. The tube is then centrifuged, and a portion of the extract is transferred to a tube containing 50 mg primary secondary amine sorbent plus 150 mg anhydrous MgSO4/mL of extract. After a mixing and centrifugation step, the extract is transferred to autosampler vials for concurrent analysis by gas chromatography/mass spectrometry (GC/MS) and liquid chromatography/tandem mass spectrometry. Independent of the original sample pH, the use of buffering during the extraction yields pH <4 in the MeCN extract and >5 in the water phase, which increases recoveries of both acid- and base-sensitive pesticides. The method was evaluated for 32 diverse pesticides in different matrixes, and typical percent recoveries were 95 +/- 10, even for some problematic pesticides. Optional solvent exchange to toluene prior to GC/MS analysis was also evaluated, showing equally good results with the benefit of lower detection limits, but at the cost of more time, material, labor, and expense.     

Analytical and semi‐preparative enantioresolution of (RS)‐ketorolac from pharmaceutical formulation and in human plasma by HPLC

An efficient, simple, validated, analytical and semi‐preparative HPLC method has been developed for direct enantioresolution of (RS)‐Ketorolac (Ket) using monochloro‐methylated derivatives of cellulose and amylose, i.e. cellulose (tris‐3‐chloro‐4‐methylphenylcarbamate) and amylose (tris‐5‐chloro‐2‐methylphenylcarbamate) as chiral stationary phases (CSPs) with photo diode array detection at 320 nm. Enantioresolution was carried out in samples of human plasma spiked with (RS)‐Ket under normal and reversed‐phase elution modes with suitable mobile phase compositions. The effect of nature of alcohols (MeOH, EtOH, PrOH and n‐BuOH) and other solvents (MeCN and MeOH) as organic modifiers in the mobile phase was investigated on the separation performance of two CSPs in terms of retention and separation of enantiomers. The best resolution was observed on cellulose‐based CSP using EtOH, while using 2‐PrOH (15%) and amylose‐based CSP obtained the highest retention. Under reversed‐phase elution mode the best enantioseparation was observed using 30% MeCN with ammonium formate buffer. The elution order of enantiomers was ascertained by determining specific rotations. The limit of detection and quantitation values were 5 and 15.5 ng/mL for each enantiomer of (RS)‐Ket, respectively. Copyright © 2016 John Wiley & Sons, Ltd.     

An improved sample preparation method for the sensitive detection of peptides by MALDI‐MS

We describe here an optimization study of the sample preparation conditions for sensitive detection of peptides by matrix‐assisted laser desorption/ionization mass spectrometry (MALDI‐MS). Among many factors in the conditions, we varied the percent acetonitrile in the peptide solution, the percent acetonitrile in the matrix solution and the α‐cyano‐4‐hydroxycinnamic acid (CHCA) concentration in the matrix solution. CHCA was chosen because it is the most frequently used matrix for analyzing peptides. The well‐established dried‐droplet method was employed for sample deposition. The examined range of the concentration of CHCA was from 0.01 to 10 mg/ml, and the MeCN content of the solvent for matrix/analyte was 10% to 50%. The indicator for the detection sensitivity was the S/N ratio of the peaks of peptides used. Highly increased sensitivity (100‐ to 1000‐fold) was observed for the optimal CHCA concentration of 0.1 mg/ml in 20% MeCN/0.1% aq. trifluoroacetic acid (TFA), as compared with the conventional concentration (10 mg/ml) in 50% MeCN/0.1% aq. TFA. For example, the limit of detection of human ACTH 18–39 was 10 amol/well for the optimal condition but 10 fmol/well for the conventional condition. The optimal condition (0.1 mg/ml CHCA in 20% MeCN/0.1% aq. TFA) was verified with five model peptides and provided significant improvement in sensitivity (by two to three orders of magnitude) compared with the conventional conditions. Optimizing the CHCA concentration and solvent composition significantly improved the detection sensitivity in the analysis of peptides by MALDI‐MS. Copyright © 2013 John Wiley & Sons, Ltd.     

Development and validation of an analytical method for the quantification of cytochrome c in skin transport studies

A simple isocratic HPLC method for the quantification of Cytochrome c in skin permeation samples was developed and validated. The mobile phase comprised a 41 : 59 mixture of an organic phase A (0.1% trifluoroacetic acid in a 90 : 10 mixture of MeCN–H₂O) and an aqueous phase B (0.1% trifluoroacetic acid in H₂O). The Cytochrome c retention and run times were 2.62 and 8.0 min, respectively—much shorter than those for existing gradient methods. The response was accurate, precise and linear from 2.5 to 25 μg/mL. The mean recoveries for intra‐day and inter‐day analysis ranged from 88.5 to 103.8% and the RSD varied from 0.05 to 1.55%. The assay was used to quantify transport of Cytochrome c across intact and laser‐microporated porcine skin in vitro. Cytochrome c permeation and the amount of protein retained within the membrane over 24 h were quantified as a function of the number of micropores. Although no Cytochrome c permeation was observed across intact skin, laser microporation enabled delivery of 22.9 ± 3.3 and 56.0 ± 15.9 μg/cm² of the protein across skin samples with 300 and 1800 micropores, respectively. In conclusion, the HPLC method provided a fast, efficient means to quantify Cytochrome c in samples from skin transport studies. Copyright © 2009 John Wiley & Sons, Ltd.     

Measurement of Cefixime in Serum and Cerebrospinal Fluid by High-Performance Liquid Chromatography

Abstract

Cefixime is a new cephalosporin antibiotic for oral administration. A high-performance liquid chromatographic (HPLC) method was developed to measure cefixime in small volumes of serum and cerebrospinal fluid (CSF) to conduct a pharmacokinetics study in pediatric patients. The assay involved precipitation of serum proteins with 6% trichloroacetic acid, using 7-hydroxycoumarin as an internal standard. Chromatographic separation was accomplished using ultrasphere C8 column and mobile phase containing 15% acetonitrile in a buffer at a detection wave length of 280 nm. The retention time of cefixime and 7-hydroxycoumarin was about 5 and 9 minutes, respectively. The method was suitable for quantitation of cefixime at a concentration ranging from 0.05 to 10 μg/ml. The coefficient of variation was less than 3%. The technique was used successfully to measure cefixime in serum and CSF obtained from an infant receiving cefixime.     

Ion Chromatographic Determination of Dibutylphosphoric Acid in Nuclear Fuel Reprocessing Streams

Abstract

A rapid method has been developed for the determination of dibutylphosphoric acid (DBP), a degradation product of tributylphosphate (TBP), which is used in a solvent extraction process for recovery of uranium. DBP along with any monobutylphosphoric acid (MBP) and phosphoric acid are extracted from the organic phase into dilute sodium hydroxide. DBP is separated from MBP and phosphoric acid by ion chromatography and is determined on a peak height ratio basis. The method requires only 30 minutes per analysis as compared to the conventional alumina column separation-colorimetric determination procedure which requires 8 h to complete. DBP has been quantified to a lower limit of 1.5 mg/L. Relative standard deviations ranging from 5.7% to 0.4% were obtained for DBP concentrations ranging from 1.5 to 500 mg/L, respectively.     

Asymmetric cross-aldol reaction of isatin and ketones catalyzed by crude earthworm extract as efficient biocatalyst

ABSTRACT

The asymmetric cross-aldol reaction of simple ketones (acetone, cyclohexanone) with isatin derivatives in the presence of crude extract from earthworms as green and effective biocatalyst proceeds easily in MeCN/H₂O (1:1) as solvent to afford 3-hydroxy-2-oxindoles derivatives. Ten compounds were synthesized in high yields (62–88%) and moderate ee (29–42%). Structure of the synthesized compounds has been characterized on the basis of NMR spectra and CHN analysis. The ee of the obtained compounds was determined by chiral phase HPLC analysis.     

OPA柱前衍生反相高效液相色谱法测定氨基酸含量

建立了邻苯二甲醛（ＯＰＡ）手动柱前衍生反相高效液相色谱法测定样品中氨基酸含量的方法。以邻苯二甲醛（ＯＰＡ）／３－巯基丙酸（３－ＭＰＡ）为衍生试剂进行衍生,ＯＤＳ柱分离,３４０ｎｍ检测,在４０ｍｉｎ内１８种氨基酸全部得到基线分离。测定牛血清白蛋白（ＢＳＡ）的氨基酸组成和小鼠血清中的游离氨基酸,取得满意的结果。     

Enantioselective determination of acylamino acid fungicides in vegetables and fruits by chiral liquid chromatography coupled with tandem mass spectrometry

An efficient and sensitive enantioselective method for simultaneous determination of three acylamino acid fungicides in vegetables and fruits was presented by high-performance liquid chromatography (HPLC) coupled with tandem mass spectrometry. The three fungicides (benalaxyl, furalaxyl, and metalaxyl) residues in samples were extracted with acetonitrile containing 1% acetic acid and an aliquot was cleaned up with Si-(CH(2) )(3) -NH-(CH(2) )(2) -NH(2) and C(18) sorbent. Complete enantioseparation of three acylamino acid fungicides enantiomers was obtained under reversed-phase conditions on a cellulose tris (4-chloro-3-methylphenylcarbamate) column at 25°C using acetonitrile-0.1% formic acid solution (45:55, v/v) as a mobile phase. The elution orders of the eluted enantiomers were determined by a circular dichroism (CD) detector. The linearity, matrix effect, recovery, and precision were evaluated. Good linearity was obtained over the concentration range of 0.5-250 μg/L for each enantiomer in the standard solution and sample matrix calibration curves. There was no significant matrix effect for three fungicides determination based on the method. The inter-day mean recoveries, intra-day repeatability, and inter-day reproducibility varied from 81.3 to 95.7%, 2.2 to 9.4%, and 2.3 to 9.6%, respectively. The method provided high selectivity and sensitivity, and limits of quantification for enantiomers of three fungicides in vegetables and fruits were both 1 μg/kg.     

高效液相色谱法测定鼠脑谷氨酸脱羧酶活性

应用高效液相色谱-电化学检测技术建立了鼠脑谷氨酸脱羧酶(GAD)活性的测定方法.方法快速,灵敏.总分析时间为9min.γ-氨基丁酸的最低柱上检测限为50fmol.天内及天间相对标准差分别小于4.3%和9.8%.测得CAD最大催化速率为0.723nmol/min/mg蛋白.讨论了最佳衍生化pH及色谱条件.     

Evaluation of common organic solvents for gas chromatographic analysis and stability of multiclass pesticide residues

In this study, we evaluated the suitability of six common organic solvents for gas chromatographic (GC) analysis of pesticides. Three of these, acetone, acetonitrile (MeCN) and ethyl acetate (EtAc), represent extraction solvents commonly used in multiresidue methods for determination of pesticides in produce. The other three, isooctane, hexane and toluene, often serve as exchange solvents before a GC analysis. An ideal solvent for GC analysis of multiclass pesticide residues should be compatible with: the analytes, sample preparation, and GC analysis. This study addresses each aspect with emphasis placed on stability of selected pesticides in the given solvents. In this respect, the exchange solvents proved to be superior to the more polar extraction solvents. Degradation of N-trihalomethylthio fungicides (e.g., captan, folpet, dichlofluanid) in MeCN was observed only in certain lots of the tested MeCN, but even if it occurred, the stability of these analytes as well as that of dicofol and chlorothalonil was dramatically improved by the addition of 0.1% (v/v) acetic acid. Dicofol and chlorothalonil were also unstable in acetone, and pesticides with a thioether group (e.g., fenthion, disulfoton) degraded in the tested EtAc. Formation of isomers of certain pyrethroids (deltamethrin, lambda-cyhalothrin) was recorded in the chromatograms from MeCN and acetone solutions, but this effect more likely occurred during the GC injection than in solution. For several reasons, MeCN was found to be the most suitable solvent for extraction of a wide polarity range of pesticide residues from produce. After acidification, the stability of problematic pesticides in MeCN is acceptable, and MeCN can also serve as a medium for GC injection; therefore solvent exchange is generally not required before GC analysis. If sensitivity is an issue in splitless injection, then toluene was demonstrated to be the best exchange solvent due to its miscibility with MeCN and stronger responses of relatively more polar pesticides (e.g., acephate, methamidophos) as compared to hexane and isooctane.     

Synchronized extraction and purification of L-lactic acid from fermentation broth by emulsion liquid membrane technique

This study aims to investigate the recovery of L-lactic acid from fermentation broth by an emulsion liquid membrane (ELM), made up of sunflower oil as the diluent, Sorbitan monooleate (Span 80) as the surfactant, Aliquat 336 as the carrier, and sodium hydroxide (NaOH) solution as the internal aqueous phase. Particularly, the ELM process was properly set up, through the identification of the optimal ELM operating parameters on the final extraction efficiency of L-lactic acid, including Span 80 concentration, NaOH concentration, Aliquat 336 concentration, stirring speed, phase ratio, and treatment ratio. The obtained results showed that the extraction efficiency of L-lactic acid reached up to 99% under the following optimal conditions: 10 minutes after contact time, 4% w/w Span 80, 3% w/w Aliquat 336, 0.1?N solution of NaOH, stirring speed of 300?rpm, phase ratio 1, and treatment ratio 0.25. A stable system without considerable emulsion swelling and breakage was monitored using a dynamic light scattering (DLS) apparatus for the selected optimal ELM operating parameters.     

Evaluation of two fast and easy methods for pesticide residue analysis in fatty food matrixes

Two rapid methods of sample preparation and analysis of fatty foods (e.g., milk, eggs, and avocado) were evaluated and compared for 32 pesticide residues representing a wide range of physicochemical properties. One method, dubbed the quick, easy, cheap, effective, rugged, and safe (QuEChERS) method for pesticide residue analysis, entailed extraction of 15 g sample with 15 mL acetonitrile (MeCN) containing 1% acetic acid followed by addition of 6 g anhydrous magnesium sulfate and 1.5 g sodium acetate. After centrifugation, 1 mL of the buffered MeCN extract underwent a cleanup step (in a technique known as dispersive solid-phase extraction) using 50 mg each of C18 and primary secondary amine sorbents plus 150 mg MgSO4. The second method incorporated a form of matrix solid-phase dispersion (MSPD), in which 0.5 g sample plus 2 g C18 and 2 g anhydrous sodium sulfate was mixed in a mortar and pestle and added above a 2 g Florisil column on a vacuum manifold. Then, 5 x 2 mL MeCN was used to elute the pesticide analytes from the sample into a collection tube, and the extract was concentrated to 0.5 mL by evaporation. Extracts in both methods were analyzed concurrently by gas chromatography/mass spectrometry and liquid chromatography/tandem mass spectrometry. The recoveries of semi-polar and polar pesticides were typically 100% in both methods (except that basic pesticides, such as thiabendazole and imazalil, were not recovered in the MSPD method), but recovery of nonpolar pesticides decreased as fat content of the sample increased. This trend was more pronounced in the QuEChERS method, in which case the most lipophilic analyte tested, hexachlorobenzene, gave 27 +/- 1% recovery (n=6) in avocado (15% fat) with a<10 ng/g limit of quantitation.     

高效液相色谱-串联质谱法同时测定水溶性迷迭香提取物中迷迭香酸、阿魏酸和咖啡酸的含量

本文建立了一种同时快速测定水溶性迷迭香提取物中迷迭香酸、阿魏酸和咖啡酸含量的高效液相色谱-串联质谱(HPLC/MS/MS)分析方法。以甲醇和0.1%乙酸铵溶液为HPLC的流动相;MS/MS使用多反应监测(MRM)扫描方式,在3 min内可完成迷迭香酸、阿魏酸和咖啡酸三种化合物的分离分析。上述三种分析物在5~500 ng/mL范围内线性良好(r>0.999);检出限均低于5.0 ng/mL。本法操作简便、分析速度快、灵敏度高,可用于水溶性迷迭香提取物中迷迭香酸、阿魏酸和咖啡酸含量的测定。     

爱滋病病毒中肽段的酶促合成

为了进一步研究酶促合成在多肽合成中的实际应用,选择合成了爱滋病病毒(人类免疫缺损病毒,HIV-I)的gp41中氨基酸序列598-609的三个肽段,该部分是HIV-I中的2个抗原决定簇部分,H-Leu-Glg-Leu-Trp-Glg-cgs-Ser-Glg-Lgs-Leu-Ile-Cgs-OH可以作为抗原来检测HIV抗体.