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Sample records for chromatography method validacion

  1. Freeze chromatography method and apparatus

    DOEpatents

    Scott, C.D.

    1987-04-16

    A freeze chromatography method and apparatus are provided which enable separation of the solutes contained in a sample. The apparatus includes an annular column construction comprising cylindrical inner and outer surfaces defining an annular passage therebetween. One of the surfaces is heated and the other cooled while passing an eluent through the annular passageway so that the eluent in contact with the cooled surface freezes and forms a frozen eluent layer thereon. A mixture of solutes dissolved in eluent is passed through the annular passageway in contact with the frozen layer so that the sample solutes in the mixture will tend to migrate either toward or away the frozen layer. The rate at which the mixture flows through the annular passageway is controlled so that the distribution of the sample solutes approaches that at equilibrium and thus a separation between the sample solutes occurs. 3 figs.

  2. Liquid chromatography detection unit, system, and method

    DOEpatents

    Derenzo, Stephen E.; Moses, William W.

    2015-10-27

    An embodiment of a liquid chromatography detection unit includes a fluid channel and a radiation detector. The radiation detector is operable to image a distribution of a radiolabeled compound as the distribution travels along the fluid channel. An embodiment of a liquid chromatography system includes an injector, a separation column, and a radiation detector. The injector is operable to inject a sample that includes a radiolabeled compound into a solvent stream. The position sensitive radiation detector is operable to image a distribution of the radiolabeled compound as the distribution travels along a fluid channel. An embodiment of a method of liquid chromatography includes injecting a sample that comprises radiolabeled compounds into a solvent. The radiolabeled compounds are then separated. A position sensitive radiation detector is employed to image distributions of the radiolabeled compounds as the radiolabeled compounds travel along a fluid channel.

  3. Chromatography.

    ERIC Educational Resources Information Center

    Brantley, L. Reed, Sr.; Demanche, Edna L.; Klemm, E. Barbara; Kyselka, Will; Phillips, Edwin A.; Pottenger, Francis M.; Yamamoto, Karen N.; Young, Donald B.

    This booklet presents some activities on chromatography. Directions for preparing leaf pigment extracts using alcohol are given, and paper chromatography and thin-layer chromatography are described as modifications of the basic principles of chromatography. (KHR)

  4. Chromatography

    MedlinePlus

    Chromatography is a way of separating two or more chemical compounds. Chemical compounds are chemicals that are ... of chemical compound. There are different kinds of chromatography. These include gas, high pressure liquid, or ion ...

  5. A Better Method for Filling Pasteur Pipet Chromatography Columns

    ERIC Educational Resources Information Center

    Ruekberg, Ben

    2006-01-01

    An alternative method for the preparation of Pasteur pipet chromatography columns is presented that allows the column to be filled with solvent without bubbles and allows greater control of fluid flow while the materials to be separated are added. Students are required to wear gloves and goggles and caution should be used while handling glass…

  6. Modified electrokinetic sample injection method in chromatography and electrophoresis analysis

    DOEpatents

    Davidson, J. Courtney; Balch, Joseph W.

    2001-01-01

    A sample injection method for horizontal configured multiple chromatography or electrophoresis units, each containing a number of separation/analysis channels, that enables efficient introduction of analyte samples. This method for loading when taken in conjunction with horizontal microchannels allows much reduced sample volumes and a means of sample stacking to greatly reduce the concentration of the sample. This reduction in the amount of sample can lead to great cost savings in sample preparation, particularly in massively parallel applications such as DNA sequencing. The essence of this method is in preparation of the input of the separation channel, the physical sample introduction, and subsequent removal of excess material. By this method, sample volumes of 100 nanoliter to 2 microliters have been used successfully, compared to the typical 5 microliters of sample required by the prior separation/analysis method.

  7. Rapid column heating method for subcritical water chromatography.

    PubMed

    Fogwill, Michael O; Thurbide, Kevin B

    2007-01-19

    A novel resistive heating method is presented for subcritical water chromatography (SWC) that provides higher column heating rates than those conventionally obtained from temperature-programmed gas chromatography (GC) convection ovens. Since the polarity of water reduces dramatically with increasing temperature, SWC employs column heating to achieve gradient elution. As such, the rate at which the mobile phase is heated directly impacts the magnitude of such gradients applied in SWC. Data from the current study demonstrate that the maximum column heating rate attainable in a typical SWC apparatus (i.e. using a GC convection oven) is around 10 degrees C/min, even at instrument oven settings of over three times this value. Conversely, by wrapping the separation column with ceramic insulation and a resistively heated wire, the column heating rates are increased five-fold. As a result, elution times can be greatly decreased in SWC employing gradients. Separations of standard alcohol test mixtures demonstrate that the retention time of the latest eluting component decreases by 35 to 50% using the prototype method. Additionally, solute retention times in this mode deviate by less than 1% RSD over several trials, which compares very well to those obtained using a conventional GC convection oven. Results suggest that the developed method can be a useful alternative heating technique in SWC.

  8. Are analysts doing method validation in liquid chromatography?

    PubMed

    Ruiz-Angel, M J; García-Alvarez-Coque, M C; Berthod, A; Carda-Broch, S

    2014-08-01

    Method validation is being applied in the reported analytical methods for decades. Even before this protocol was defined, authors already somehow validated their methods without full awareness. They wished to assure the quality of their work. Validation is an applied approach to verify that a method is suitable and rugged enough to function as a quality control tool in different locations and times. The performance parameters and statistical protocols followed throughout a validation study vary with the source of guidelines. Before single laboratory validation, an analytical method should be fully developed and optimized. The purpose of the validation is to confirm performance parameters that are determined during method development, and it should provide information on how the method will perform under routine use. An unstable method may require re-validation. Further method development and optimization will be needed if validation results do not meet the accepted performance standards. When possible, the validation protocol should also be conducted as a collaborative study by multiple laboratories, on different instruments, reagents, and standards. At this point, it would be interesting to know how people are validating their methods. Are they evaluating all defined validation parameters? Are they indicating the followed guidelines? Is re-validation really currently used? Is validation performed by a single laboratory, or is it a collaborative work by several laboratories? Is it an evolving discipline? In this survey, we will try to answer these questions focused to the field of liquid chromatography.

  9. New method for determining total dietary fiber by liquid chromatography.

    PubMed

    Ohkuma, K; Matsuda, I; Katta, Y; Tsuji, K; Ohkuma, K; Matsuda, I; Katta, Y; Tsuji, K

    2000-01-01

    The molecular weight limit of water-soluble dietary fiber (SDF) determined by the Prosky method was studied by liquid chromatography (LC). It was confirmed that only SDF with an average degree of polymerization of 12 or higher can be determined by the Prosky method. Total dietary fiber (TDF) was determined by 2 additional methods using LC. In the first method, the total quantity of insoluble dietary fiber (IDF) and high molecular weight SDF (HMSDF) was determined according to the modified Prosky method (MES-TRIS buffer-based). The quantitatively collected final filtrate was analyzed by LC for the quantity of low molecular weight SDF (LMSDF), and the 2 quantities were totaled to obtain TDF. TDF values thus determined for rice, polished or unpolished, soybean flour, and pressed barley were higher than those determined by the Prosky method by approximately 6, 3.5, and 3.5%, respectively. In the second method, direct determination by LC analysis was done on samples after enzymatic treatment according to the Prosky method. Results showed that the determination of LMSDF, in particular, was highly accurate and more effective. In both of these methods, the quantity of LMSDF was determined from its chromatographic peak area ratio to glucose as an internal standard, which was produced by hydrolysis.

  10. Liquid chromatography method to determine polyamines in thermosetting polymers.

    PubMed

    Dopico-García, M S; López-Vilariño, J M; Fernández-Martínez, G; González-Rodríguez, M V

    2010-05-14

    A simple, robust and sensitive analytical method to determine three polyamines commonly used as hardeners in epoxy resin systems and in the manufacture of polyurethane is reported. The studied polyamines are: one tetramine, TETA (triethylenetetramine), and two diamines, IPDA (Isophorone diamine) and TCD-diamine (4,7-methano-1H-indene-5,?-dimethanamine, octahydro-). The latter has an incompletely defined structure, and, as far as we know, has not been previously determined by other methods. All three polyamines contain primary amines; TETA also contains secondary amines. The analytical method involves derivatization with 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate, used for the first time for these compounds, followed by high performance liquid chromatography (HPLC) analysis with a fluorescence (FL) detector (lambda excitation 248nm, lambda emision 395nm). The HPLC-DAD-LTQ Orbitrap MS was used in order to provide structural information about the obtained derivatized compounds. The hybrid linear ion trap LTQ Orbitrap mass spectrometer has been introduced in recent years and provides a high mass accuracy. The structures of the derivatized analytes were identified from the protonated molecular ions [M+H](+) and corresponded to the fully labelled analytes. The following analytical parameters were determined for the method using the HPLC-FL: linearity, precision (2.5-10%), instrumental precision intraday (0.8-1.5%) and interday (2.9-6.3%), and detection limits (0.02-0.14mgL(-1)). The stability of stock solutions and derivatized compounds was also investigated. The method was applied to determine the amine free content in epoxy resin dust collected in workplaces.

  11. An improved method for analysis of biomass sugars and galacturonic acid by anion exchange chromatography

    Technology Transfer Automated Retrieval System (TEKTRAN)

    While the most accurate method for analysis of sugars in biomass is based on gas chromatography of trimethylsilane or alditol acetate derivitives of sugars, the derivation method is time consuming and laborious. In comparison, sample preparation for sugar analysis using liquid chromatography is a si...

  12. New methods and materials for solid phase extraction and high performance liquid chromatography

    SciTech Connect

    Dumont, Philip John

    1996-04-23

    This paper describes methods for solid phase extraction and high performance liquid chromatography (HPLC). The following are described: Effects of Resin Sulfonation on the Retention of Polar Organic Compounds in Solid Phase Extraction; Ion-Chromatographic Separation of Alkali Metals In Non-Aqueous Solvents; Cation-Exchange Chromatography in Non-Aqueous Solvents; and Silicalite As a Stationary Phase For HPLC.

  13. Comparison between high-performance liquid chromatography and gas chromatography methods for fatty acid identification and quantification in potato crisps.

    PubMed

    Sanches-Silva, A; Rodríguez-Bernaldo de Quirós, A; López-Hernández, J; Paseiro-Losada, P

    2004-04-02

    A reversed-phase high performance liquid chromatographic (RP-HLPC) method was compared with a gas chromatography-flame ionization detection (GC-FID) method for determining fatty acids in potato crisps. Different extraction procedures were used. Fatty acids were quantified by linear regression. Both methods presented good precision (R.S.D. < or = 5.88%) and recovery (> or = 82.31%). The precision using HPLC method was slightly better than for GC-FID method. There was good agreement between the fatty acid composition of potato crisps analysed by both methods. For most purposes the HPLC method would be better. However, when more fatty acids need to be analysed, GC is a more suitable method.

  14. Mixed-bed affinity chromatography: principles and methods.

    PubMed

    Boschetti, Egisto; Righetti, Pier Giorgio

    2015-01-01

    Mixed-bed chromatography is far from being a well-established technology within the panoply of bioseparation tools. Composed of an assembly of distinct sorbents that are mixed in a single bed, they have been mostly developed in the last decade for the reduction of dynamic concentration range where they allowed discovering many low-copy proteins within very complex proteomes. Other interesting preparative applications of mixed-bed chromatography have since been developed. In this chapter the basic concepts first and then detailed application recipes are described for (1) the reduction of protein dynamic concentration range, (2) the removal of impurity traces at the last stage of a biopurification process, and (3) the selection and use of sorbents as mixed bed in protein purification.

  15. Evaluation between ultrahigh pressure liquid chromatography and high-performance liquid chromatography analytical methods for characterizing natural dyestuffs.

    PubMed

    Serrano, Ana; van Bommel, Maarten; Hallett, Jessica

    2013-11-29

    An evaluation was undertaken of ultrahigh pressure liquid chromatography (UHPLC) in comparison to high-performance liquid chromatography (HPLC) for characterizing natural dyes in cultural heritage objects. A new UHPLC method was optimized by testing several analytical parameters adapted from prior UHPLC studies developed in diverse fields of research. Different gradient elution programs were tested on seven UHPLC columns with different dimensions and stationary phase compositions by applying several mobile phases, flow rates, temperatures, and runtimes. The UHPLC method successfully provided more improved data than that achieved by the HPLC method. Indeed, even though carminic acid has shown circa 146% higher resolution with HPLC, UHPLC resulted in an increase of 41-61% resolution and a decrease of 91-422% limit of detection, depending on the dye compound. The optimized method was subsequently assigned to analyse 59 natural reference materials, in which 85 different components were ascribed with different physicochemical properties, in order to create a spectral database for future characterization of dyes in cultural heritage objects. The majority of these reference samples could be successfully distinguished with one single method through the examination of these compounds' retention times and their spectra acquired with a photodiode array detector. These results demonstrate that UHPLC analyses are extremely valuable for the acquisition of more precise chromatographic information concerning natural dyes with complex mixtures of different and/or closely related physicochemical properties, essential for distinguishing similar species of plants and animals used to colour cultural heritage objects.

  16. Integration of gas chromatography mass spectrometry methods for differentiating ricin preparation methods.

    PubMed

    Wunschel, David S; Melville, Angela M; Ehrhardt, Christopher J; Colburn, Heather A; Victry, Kristin D; Antolick, Kathryn C; Wahl, Jon H; Wahl, Karen L

    2012-05-07

    The investigation of crimes involving chemical or biological agents is infrequent, but presents unique analytical challenges. The protein toxin ricin is encountered more frequently than other agents and is found in the seeds of Ricinus communis, commonly known as the castor plant. Typically, the toxin is extracted from castor seeds utilizing a variety of different recipes that result in varying purity of the toxin. Moreover, these various purification steps can also leave or differentially remove a variety of exogenous and endogenous residual components with the toxin that may indicate the type and number of purification steps involved. We have applied three gas chromatography-mass spectrometry (GC-MS) based analytical methods to measure the variation in seed carbohydrates and castor oil ricinoleic acid, as well as the presence of solvents used for purification. These methods were applied to the same samples prepared using four previously identified toxin preparation methods, starting from four varieties of castor seeds. The individual data sets for seed carbohydrate profiles, ricinoleic acid, or acetone amount each provided information capable of differentiating different types of toxin preparations across seed types. However, the integration of the data sets using multivariate factor analysis provided a clear distinction of all samples based on the preparation method, independent of the seed source. In particular, the abundance of mannose, arabinose, fucose, ricinoleic acid, and acetone were shown to be important differentiating factors. These complementary tools provide a more confident determination of the method of toxin preparation than would be possible using a single analytical method.

  17. Advanced high-performance liquid chromatography method development. Discovering unexpected choices in chromatography.

    PubMed

    Rieger, H J; Molnar, I

    2002-03-01

    The influence of some important experimental parameters on the resolution of compounds as well as the validity of widely used rules of thumb and of common expectations about how to improve resolution is discussed. It will be shown, on the basis of selected examples, that the general expectations about how the experimental parameters have to be adjusted for better resolution does not cover all chances for resolution improvement. The tool for understanding the method and to discover all chances for increasing selectivity is the resolution map of a method.

  18. Purification of flavonoids from licorice using an off-line preparative two-dimensional normal-phase liquid chromatography/reversed-phase liquid chromatography method.

    PubMed

    Fan, Yunpeng; Fu, Yanhui; Fu, Qing; Cai, Jianfeng; Xin, Huaxia; Dai, Mei; Jin, Yu

    2016-07-01

    An orthogonal (71.9%) off-line preparative two-dimensional normal-phase liquid chromatography/reversed-phase liquid chromatography method coupled with effective sample pretreatment was developed for separation and purification of flavonoids from licorice. Most of the nonflavonoids were firstly removed using a self-made Click TE-Cys (60 μm) solid-phase extraction. In the first dimension, an industrial grade preparative chromatography was employed to purify the crude flavonoids. Click TE-Cys (10 μm) was selected as the stationary phase that provided an excellent separation with high reproducibility. Ethyl acetate/ethanol was selected as the mobile phase owing to their excellent solubility for flavonoids. Flavonoids co-eluted in the first dimension were selected for further purification using reversed-phase liquid chromatography. Multiple compounds could be isolated from one normal-phase fraction and some compounds with bad resolution in one-dimensional liquid chromatography could be prepared in this two-dimensional system owing to the orthogonal separation. Moreover, this two-dimensional liquid chromatography method was beneficial for the preparation of relatively trace flavonoid compounds, which were enriched in the first dimension and further purified in the second dimension. Totally, 24 flavonoid compounds with high purity were obtained. The results demonstrated that the off-line two-dimensional liquid chromatography method was effective for the preparative separation and purification of flavonoids from licorice.

  19. A Static Method as an Alternative to Gel Chromatography: An Experiment for the Undergraduate Biochemistry Laboratory

    ERIC Educational Resources Information Center

    Burum, Alex D.; Splittgerber, Allan G.

    2008-01-01

    This article describes a static method as an alternative to gel chromatography, which may be used as an undergraduate laboratory experiment. In this method, a constant mass of Sephadex gel is swollen in a series of protein solutions. UV-vis spectrophotometry is used to find a partition coefficient, KD, that indicates the fraction of the interior…

  20. Neuere Chromatographie

    NASA Astrophysics Data System (ADS)

    Hostettmann, K.

    1983-04-01

    Besides high-performance liquid chromatography (HPLC) which is now a well-established and currently used technique, several emerging methods for the isolation and separation of natural products are receiving considerable attention. Centrifugal thin-layer chromatography is a very rapid technique, but limited in resolution. Of special interest are the recently developed support-free liquid-liquid chromatography methods such as droplet counter-current chromatography (DCCC) and rotation locular counter-current chromatography (RLCC). This latter method was applied to the separation of the enantiomers of (±)-norephedrine.

  1. A Simple Robust Orthogonal Background Correction Method for Two-Dimensional Liquid Chromatography

    PubMed Central

    Filgueira, Marcelo R.; Castells, Cecilia; Carr, Peter W.

    2012-01-01

    Background correction is a very important step that must be done before peak detection or any quantification procedure. When successful, this step greatly simplifies such procedures and enhances the accuracy of quantification. In the past, much effort has been invested to correct drifting baseline in one dimensional chromatography. In fast online comprehensive two-dimensional liquid chromatography (LC×LC) coupled with diode array detector (DAD), the change in the refractive index (RI) of the mobile phase in very fast gradients causes extremely serious baseline disturbances. The method reported here can be combined with many of existing baseline correction methods for one dimensional (1D) chromatography in two dimensional (2D) liquid chromatography and recreate the background structure for further correction. When such methods are applied orthogonally to the second dimension (2D), the background correction is dramatically improved. It results in an almost zero mean background level and it provides better background correction than does a simple subtraction of a blank. Indeed, the method proposed does not require running a blank sample. PMID:22702415

  2. Pre-staining paper chromatography method for quantification of gamma-aminobutyric acid.

    PubMed

    Li, Haixing; Qiu, Ting; Cao, Yusheng; Yang, Jiyan; Huang, Zhibing

    2009-06-19

    The routine method of paper chromatography includes five steps: spotting, separating, drying, spraying/immersing and color development. In this paper, a pre-staining paper chromatography which only consisted of spotting, separating and color development was developed for quantitative analysis of gamma-aminobutyric acid. Compared to the routine paper chromatography, the improved method is clean, rapid, inexpensive and reproducible. The effects of ninhydrin concentration, color temperature, color time and Cu(2+) concentration on the color yield in the ninhydrin reaction were optimized. And then the pre-staining paper chromatography coupled with vis spectrophotometry was applied to gamma-aminobutyric acid quantification. The results indicated that the limit of detection was 0.05 mg mL(-1) and the linear range was from 0.5 to 20.0 mg mL(-1). Furthermore, an excellent correlation coefficient was observed with an R(2)=0.998. The method is accurate (RSD<2.64%), and has good recoveries (102.7-103.9%). The validation of the modified technique was verified by a HPLC method.

  3. Downstream processing and chromatography based analytical methods for production of vaccines, gene therapy vectors, and bacteriophages

    PubMed Central

    Kramberger, Petra; Urbas, Lidija; Štrancar, Aleš

    2015-01-01

    Downstream processing of nanoplexes (viruses, virus-like particles, bacteriophages) is characterized by complexity of the starting material, number of purification methods to choose from, regulations that are setting the frame for the final product and analytical methods for upstream and downstream monitoring. This review gives an overview on the nanoplex downstream challenges and chromatography based analytical methods for efficient monitoring of the nanoplex production. PMID:25751122

  4. Retention modeling and method development in hydrophilic interaction chromatography.

    PubMed

    Tyteca, Eva; Périat, Aurélie; Rudaz, Serge; Desmet, Gert; Guillarme, Davy

    2014-04-11

    In the present study, the possibility of retention modeling in the HILIC mode was investigated, testing several different literature relationships over a wide range of different analytical conditions (column chemistries and mobile phase pH) and using analytes possessing diverse physico-chemical properties. Furthermore, it was investigated how the retention prediction depends on the number of isocratic or gradient trial or initial scouting runs. The most promising set of scouting runs seems to be a combination of three isocratic runs (95, 90 and 70%ACN) and one gradient run (95 to 65%ACN in 10min), as the average prediction errors were lower than using six equally spaced isocratic runs and because it is common in Method development (MD) to perform at least one scouting gradient run in the screening step to find out the best column, temperature and pH conditions. Overall, the retention predictions were much less accurate in HILIC than what is usually experienced in RPLC. This has severe implications for MD, as it restricts the use of commercial software packages that require the simulation of the retention of every peak in the chromatogram. To overcome this problem, the recently proposed predictive elution window shifting and stretching (PEWS(2)) approach can be used. In this computer-assisted MD strategy, only an (approximate) prediction of the retention of the first and the last peak in the chromatogram is required to conduct a well-targeted trial-and-error search, with suggested search conditions uniformly covering the entire possible search and elution space. This strategy was used to optimize the separation of three representative pharmaceutical mixtures possessing diverse physico-chemical properties (pteridins, saccharides and cocktail of drugs/metabolites). All problems could be successfully handled in less than 2.5h of instrument time (including equilibration).

  5. Comparison of different methods to estimate the uncertainty in composition measurement by chromatography.

    PubMed

    Ariza, Adriana Alexandra Aparicio; Ayala Blanco, Elizabeth; García Sánchez, Luis Eduardo; García Sánchez, Carlos Eduardo

    2015-06-01

    Natural gas is a mixture that contains hydrocarbons and other compounds, such as CO2 and N2. Natural gas composition is commonly measured by gas chromatography, and this measurement is important for the calculation of some thermodynamic properties that determine its commercial value. The estimation of uncertainty in chromatographic measurement is essential for an adequate presentation of the results and a necessary tool for supporting decision making. Various approaches have been proposed for the uncertainty estimation in chromatographic measurement. The present work is an evaluation of three approaches of uncertainty estimation, where two of them (guide to the expression of uncertainty in measurement method and prediction method) were compared with the Monte Carlo method, which has a wider scope of application. The aforementioned methods for uncertainty estimation were applied to gas chromatography assays of three different samples of natural gas. The results indicated that the prediction method and the guide to the expression of uncertainty in measurement method (in the simple version used) are not adequate to calculate the uncertainty in chromatography measurement, because uncertainty estimations obtained by those approaches are in general lower than those given by the Monte Carlo method.

  6. Fast and accurate numerical method for predicting gas chromatography retention time.

    PubMed

    Claumann, Carlos Alberto; Wüst Zibetti, André; Bolzan, Ariovaldo; Machado, Ricardo A F; Pinto, Leonel Teixeira

    2015-08-07

    Predictive modeling for gas chromatography compound retention depends on the retention factor (ki) and on the flow of the mobile phase. Thus, different approaches for determining an analyte ki in column chromatography have been developed. The main one is based on the thermodynamic properties of the component and on the characteristics of the stationary phase. These models can be used to estimate the parameters and to optimize the programming of temperatures, in gas chromatography, for the separation of compounds. Different authors have proposed the use of numerical methods for solving these models, but these methods demand greater computational time. Hence, a new method for solving the predictive modeling of analyte retention time is presented. This algorithm is an alternative to traditional methods because it transforms its attainments into root determination problems within defined intervals. The proposed approach allows for tr calculation, with accuracy determined by the user of the methods, and significant reductions in computational time; it can also be used to evaluate the performance of other prediction methods.

  7. Validation of a sensitive ion chromatography method for determination of monoethylsulfate in Indinavir sulfate drug substance.

    PubMed

    Prasanna, S John; Sharma, Hemant Kumar; Mukkanti, K; Sivakumaran, M; Pavan Kumar, K S R; Kumar, V Jagadeesh

    2009-12-05

    The present study relates to the optimization of an ion chromatography method to determine the content of monoethylsulfate at very low levels in Indinavir sulfate drug substance, and subsequent validation of the method to prove its suitability, reliability and sensitivity. Monoethylsulfate is a potential impurity of Indinavir sulfate, and may forms during the preparation as well as during storage. The ion chromatography method was developed in such a way that to enhance the detection level by introducing suppressor, and minimizing acquisition time by using suitable buffer of 3.2 mmole of sodium carbonate and 1 mmole of sodium hydrogen carbonate in water as eluent. The retention time of monoethylsulfate was about 9.5 min and the total acquisition time was 25 min. The optimized method was validated to prove its performance characteristics by demonstrating selectivity, sensitivity (limit of detection and quantification), linearity, precision and accuracy. The established limit of detection and quantification of monoethylsulfate in Indinavir sulfate by this method was found to be 24 ng/ml and 74 ng/ml respectively, and the overall percent accuracy (recovery) of samples evaluated at different concentration levels was found to be 97.1, indicating the sensitivity and accuracy of this optimized ion chromatography method.

  8. High-throughput multiclass method for antibiotic residue analysis by liquid chromatography-tandem mass spectrometry.

    PubMed

    Chico, J; Rúbies, A; Centrich, F; Companyó, R; Prat, M D; Granados, M

    2008-12-12

    A simple and rapid method has been developed for the residue analysis of 39 antibiotics (tetracyclines, quinolones, penicillins, sulfonamides and macrolides) in foodstuffs of animal origin. The method combines an effective extraction technique, which uses water-methanol as extracting solvent, with ultra-high-pressure liquid chromatography-tandem mass spectrometry, allowing both confirmation and quantification in a single chromatographic run. The multiresidue method has been validated in chicken muscle matrix according to European Union Decision 2002/657/EC. It has been implemented as a routine method in a Public Health Laboratory, instead of the five plates test and LC methods previously used.

  9. Minibodies and Multimodal Chromatography Methods: A Convergence of Challenge and Opportunity.

    PubMed

    Gagnon, Pete; Cheung, Chia-Wei; Lepin, Eric J; Wu, Anna M; Sherman, Mark A; Raubitschek, Andrew A; Yazaki, Paul J

    2010-02-01

    This case study describes early phase purification process development for a recombinant anticancer minibody produced in mammalian cell culture. The minibody did not bind to protein A. Cation-exchange, anion-exchange, hydrophobic-interaction, and hydroxyapatite (eluted by phosphate gradient) chromatographic methods were scouted, but the minibody coeluted with BSA to a substantial degree on each. Hydroxyapatite eluted with a sodium chloride gradient separated BSA and also removed a dimeric contaminant, but BSA consumed so much binding capacity that this proved impractical as a capture tool. Capto MMC media proved capable of supporting adequate capture and significant dimer removal, although both loading and elution selectivity varied dramatically with the amount of supernatant applied to the column. An anion-exchange step was included to fortify overall virus and DNA removal. These results illustrate the value of multimodal chromatography methods when affinity chromatography methods are lacking and conventional alternatives prove inadequate.

  10. Methods of analysis-Determination of pesticides in sediment using gas chromatography/mass spectrometry

    USGS Publications Warehouse

    Hladik, Michelle; McWayne, Megan M.

    2012-01-01

    A method for the determination of 119 pesticides in environmental sediment samples is described. The method was developed by the U.S. Geological Survey (USGS) in support of the National Water Quality Assessment (NAWQA) Program. The pesticides included in this method were chosen through prior prioritization. Herbicides, insecticides, and fungicides along with degradates are included in this method and span a variety of chemical classes including, but not limited to, chloroacetanilides, organochlorines, organophosphates, pyrethroids, triazines, and triazoles. Sediment samples are extracted by using an accelerated solvent extraction system (ASE®, and the compounds of interest are separated from co-extracted matrix interferences (including sulfur) by passing the extracts through high performance liquid chromatography (HPLC) with gel-permeation chromatography (GPC) along with the use of either stacked graphitized carbon and alumina solid-phase extraction (SPE) cartridges or packed Florisil®. Chromatographic separation, detection, and quantification of the pesticides from the sediment-sample extracts are done by using gas chromatography with mass spectrometry (GC/MS). Recoveries in test sediment samples fortified at 10 micrograms per kilogram (μg/kg) dry weight ranged from 75 to 102 percent; relative standard deviations ranged from 3 to 13 percent. Method detection limits (MDLs), calculated by using U.S. Environmental Protection Agency procedures (40 CFR 136, Appendix B), ranged from 0.6 to 3.4 μg/kg dry weight.

  11. Possibilities and limitations of the kinetic plot method in supercritical fluid chromatography.

    PubMed

    De Pauw, Ruben; Desmet, Gert; Broeckhoven, Ken

    2013-08-30

    Although supercritical fluid chromatography (SFC) is becoming a technique of increasing importance in the field of analytical chromatography, methods to compare the performance of SFC-columns and separations in an unbiased way are not fully developed. The present study uses mathematical models to investigate the possibilities and limitations of the kinetic plot method in SFC as this easily allows to investigate a wide range of operating pressures, retention and mobile phase conditions. The variable column length (L) kinetic plot method was further investigated in this work. Since the pressure history is identical for each measurement, this method gives the true kinetic performance limit in SFC. The deviations of the traditional way of measuring the performance as a function of flow rate (fixed back pressure and column length) and the isopycnic method with respect to this variable column length method were investigated under a wide range of operational conditions. It is found that using the variable L method, extrapolations towards other pressure drops are not valid in SFC (deviation of ∼15% for extrapolation from 50 to 200bar pressure drop). The isopycnic method provides the best prediction but its use is limited when operating closer towards critical point conditions. When an organic modifier is used, the predictions are improved for both methods with respect to the variable L method (e.g. deviations decreases from 20% to 2% when 20mol% of methanol is added).

  12. Caprylic acid precipitation method for impurity reduction: an alternative to conventional chromatography for monoclonal antibody purification.

    PubMed

    Brodsky, Yan; Zhang, Cheng; Yigzaw, Yinges; Vedantham, Ganesh

    2012-10-01

    We report the use of caprylic acid based impurity precipitation as (1) an alternative method to polishing chromatography techniques commonly used for monoclonal antibody purification and (2) an impurity reduction step prior to harvesting the bioreactor. This impurity reduction method was tested with protein A purified antibodies and with cell culture fluid. First, the operational parameters influencing precipitation of host cell proteins and high molecular weight aggregate in protein A pools were investigated. When used as a polishing step, the primary factor affecting purification and yield was determined to be pH. Caprylic acid precipitation was comparable to polishing IEX chromatography in reducing host cell protein and aggregate levels. A virus reduction study showed complete clearance of a model retrovirus during caprylic acid precipitation of protein A purified antibody. Caprylic acid mediated impurity precipitation in cell culture showed that the impurity clearance was generally insensitive to pH and caprylic acid concentration whereas yield was a function of caprylic acid concentration. Protein A purification of caprylic acid precipitated cell culture fluid generated less turbid product pool with reduced levels of host cell proteins and high molecular weight aggregate. The results of this study show caprylic acid precipitation to be an effective purification method that can be incorporated into a production facility with minimal cost as it utilizes existing tanks and process flow. Eliminating flow through chromatography polishing step can provide process intensification by avoiding the process tank volume constraints for high titer processes.

  13. Development and validation of ultra-high performance supercritical fluid chromatography method for determination of illegal dyes and comparison to ultra-high performance liquid chromatography method.

    PubMed

    Khalikova, Maria A; Šatínský, Dalibor; Solich, Petr; Nováková, Lucie

    2015-05-18

    A novel simple, fast and efficient ultra-high performance supercritical fluid chromatography (UHPSFC) method was developed and validated for the separation and quantitative determination of eleven illegal dyes in chili-containing spices. The method involved a simple ultrasound-assisted liquid extraction of illegal compounds with tetrahydrofuran. The separation was performed using a supercritical fluid chromatography system and CSH Fluoro-Phenyl stationary phase at 70°C. The mobile phase was carbon dioxide and the mixture of methanol:acetonitrile (1:1, v/v) with 2.5% formic acid as an additive at the flow rate 2.0 mL min(-1). The UV-vis detection was accomplished at 500 nm for seven compounds and at 420 nm for Sudan Orange G, Butter Yellow, Fast Garnet GBC and Methyl Red due to their maximum of absorbance. All eleven compounds were separated in less than 5 min. The method was successfully validated and applied using three commercial samples of chili-containing spices - Chili sauce (Indonesia), Feferony sauce (Slovakia) and Mojo sauce (Spain). The linearity range of proposed method was 0.50-9.09 mg kg(-1) (r ≥ 0.995). The detection limits were determined as signal to noise ratio of 3 and were ranged from 0.15 mg kg(-1) to 0.60 mg kg(-1) (1.80 mg kg(-1) for Fast Garnet) for standard solution and from 0.25 mg kg(-1) to 1.00 mg kg(-1) (2.50 mg kg(-1) for Fast Garnet, 1.50 mg kg(-1) for Sudan Red 7B) for chili-containing samples. The recovery values were in the range of 73.5-107.2% and relative standard deviation ranging from 0.1% to 8.2% for within-day precision and from 0.5% to 8.8% for between-day precision. The method showed potential for being used to monitor forbidden dyes in food constituents. The developed UHPSFC method was compared to the UHPLC-UV method. The orthogonality of Sudan dyes separation by these two methods was demonstrated. Benefits and drawbacks were discussed showing the reliability of both methods for monitoring of studied illegal dyes in real

  14. Method for trapping affinity chromatography of transcription factors using aldehyde-hydrazide coupling to agarose.

    PubMed

    Jia, Yinshan; Jarrett, Harry W

    2015-08-01

    The use of a method of coupling DNA was investigated for trapping and purifying transcription factors. Using the GFP-C/EBP (CAAT/enhancer binding protein) fusion protein as a model, trapping gives higher purity and comparable yield to conventional affinity chromatography. The chemistry used is mild and was shown to have no detrimental effect on GFP fluorescence or GFP-C/EBP DNA binding. The method involves introducing a ribose nucleotide to the 3' end of a DNA sequence. Reaction with mM NaIO4 (sodium metaperiodate) produces a dialdehyde of ribose that couples to hydrazide-agarose. The DNA is combined at nM concentration with a nuclear extract or other protein mixture, and DNA-protein complexes form. The complex is then coupled to hydrazide-agarose for trapping the DNA-protein complex and the protein eluted by increasing NaCl concentration. Using a different oligonucleotide with the proximal E-box sequence from the human telomerase promoter, USF-2 transcription factor was purified by trapping, again with higher purity than results from conventional affinity chromatography and similar yield. Other transcription factors binding E-boxes, including E2A, c-Myc, and Myo-D, were also purified, but myogenin and NFκB were not. Therefore, this approach proved to be valuable for both affinity chromatography and the trapping approach.

  15. A Size-Exclusion Chromatography Method for Analysis of Clostridium difficile Vaccine Toxins.

    PubMed

    Lancaster, Catherine; Rustandi, Richard R; Pannizzo, Paola; Ha, Sha

    2016-01-01

    High-performance size-exclusion chromatography (HPSEC or SEC) is a method that can be applied to measure size distribution of proteins, including aggregates, monomers, and fragments. In the biopharmaceutical industry the quantitation of aggregates contained in biotherapeutics and protein-based vaccines is critical given the potential impact on safety, immunogenicity, and efficacy. Hence, aggregation analysis of therapeutic proteins or protein-based vaccine products is almost always a requirement of regulatory agencies. SEC, also referred to as gel-filtration chromatography, separates molecules by size through a porous resin stationary phase. Under isocratic flow small molecules are retained on the column longer than large molecules. Here we describe the use of this SEC technique to characterize aggregation levels for four different protein antigens for a Clostridium difficile vaccine.

  16. An enzymatic method for assaying sulbactam in human serum: comparison with high performance liquid chromatography.

    PubMed

    Sotto, A; Peray, P; Geny, F; Brunschwig, C; Carrière, C; Galtier, M; Ramuz, M; Jourdan, J

    1995-03-01

    An enzymatic method using nitrocefin as substrate was developed to assay sulbactam in human serum. Serum containing sulbactam was incubated with purified titrated TEM-1 beta-lactamase and nitrocefin was then added to the mixture to determine the remaining beta-lactamase activity and consequently the concentration of sulbactam. Assays were carried out on five patients with pulmonary infections receiving sulbactam plus amoxycillin iv. The values for serum sulbactam concentrations determined by the enzymatic method were compared with those determined by high performance liquid chromatography (HPLC). The correlation coefficient was 0.990 for serum sulbactam concentrations below 15 mg/L.

  17. Comparison of photoacoustic radiometry to gas chromatography/mass spectrometry methods for monitoring chlorinated hydrocarbons

    SciTech Connect

    Sollid, J.E.; Trujillo, V.L.; Limback, S.P.; Woloshun, K.A.

    1996-03-01

    A comparison of two methods of gas chromatography mass spectrometry (GCMS) and a nondispersive infrared technique, photoacoustic radiometry (PAR), is presented in the context of field monitoring a disposal site. First is presented an historical account describing the site and early monitoring to provide an overview. The intent and nature of the monitoring program changed when it was proposed to expand the Radiological Waste Site close to the Hazardous Waste Site. Both the sampling methods and analysis techniques were refined in the course of this exercise.

  18. Development of Hydrophilic Interaction Liquid Chromatography Method for the Analysis of Moxonidine and Its Impurities

    PubMed Central

    2016-01-01

    Fast and simple hydrophilic interaction liquid chromatography (HILIC) method was developed and validated for the analysis of moxonidine and its four impurities (A, B, C, and D) in pharmaceutical dosage form. All experiments were performed on the Agilent Technologies 1200 high-performance liquid chromatography (HPLC) system using Zorbax RX-SIL, 250 mm × 4.6 mm, 5 μm column as stationary phase (T = 25°C, F = 1 mL/min, and λ = 255 nm), and mixture of acetonitrile and 40 mM ammonium formate buffer (pH 2.8) 80 : 20 (v/v) as mobile phase. Under the optimal chromatographic conditions, selected by central composite design, separation and analysis of moxonidine and its four impurities are enabled within 12 minutes. Validation of the method was conducted in accordance with ICH guidelines. Based on the obtained results selectivity, linearity (r ≥ 0.9976), accuracy (recovery: 93.66%–114.08%), precision (RSD: 0.56%–2.55%), and robustness of the method were confirmed. The obtained values of the limit of detection and quantification revealed that the method can be used for determination of impurities levels below 0.1%. Validated method was applied for determination of moxonidine and its impurities in commercially available tablet formulation. Obtained results confirmed that validated method is fast, simple, and reliable for analysis of moxonidine and its impurities in tablets. PMID:27847672

  19. Efficient methods for isolating five phytochemicals from Gentiana macrophylla using high-performance countercurrent chromatography.

    PubMed

    Rho, Taewoong; Jung, Mila; Lee, Min Won; Chin, Young-Won; Yoon, Kee Dong

    2016-12-01

    Efficient high-performance countercurrent chromatography methods were developed to isolate five typical compounds from the extracts of Gentiana macrophylla. n-Butanol-soluble extract of G. macrophylla contained three hydrophilic iridoids, loganic acid (1), swertiamarin (2) and gentiopicroside (3), and a chromene derivative, macrophylloside D (4) which were successfully isolated by flow rate gradient (1.5 mL/min in 0-60 min, 5.0 mL/min in 60-120 min), and consecutive flow rate gradient HPCCC using n-butanol/0.1% aqueous trifluoroacetic acid (1:1, v/v, normal phase mode) system. The yields of 1-4 were 22, 16, 122, and 6 mg, respectively, with purities over 97% in a flow rate gradient high-performance countercurrent chromatography, and consecutive flow rate gradient high-performance countercurrent chromatography gave 1, 2, 3 (54, 41, 348 mg, respectively, purities over 97%) and 4 (13 mg, purity at 95%) from 750 mg of sample. The main compound in methylene chloride soluble extract, 2-methoxyanofinic acid, was successfully separated by n-hexane/ethyl acetate/methanol/water (4:6:4:6, v/v/v/v, flow-rate: 4 mL/min, reversed phase mode) condition. The structures of five isolates were elucidated by (1) H, (13) C NMR and ESI-Q-TOF-MS spectroscopic data which were compared with previously reported values.

  20. A versatile noninvasive method for adsorber quantification in batch and column chromatography based on the ionic capacity.

    PubMed

    Huuk, Thiemo C; Briskot, Till; Hahn, Tobias; Hubbuch, Jürgen

    2016-05-01

    Within the Quality by Design (QbD) framework proposed by the International Conference on Harmonisation (ICH), high-throughput process development (HTPD) and mechanistic modeling are of outstanding importance for future biopharmaceutical chromatography process development. In order to compare the data derived from different column scales or batch chromatographies, the amount of adsorber has to be quantified with the same noninvasive method. Similarly, an important requirement for the implementation of mechanistic modeling is the reliable determination of column characteristics such as the ionic capacity Λ for ion-exchange chromatography with the same method at all scales and formats. We developed a method to determine the ionic capacity in column and batch chromatography, based on the adsorption/desorption of the natural, uv-detectable amino acid histidine. In column chromatography, this method produces results comparable to those of classical acid-base titration. In contrast to acid-base titration, this method can be adapted to robotic batch chromatographic experiments. We are able to convert the adsorber volumes in batch chromatography to the equivalent volume of a compressed column. In a case study, we demonstrate that this method increases the quality of SMA parameters fitted to batch adsorption isotherms, and the capability to predict column breakthrough experiments. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:666-677, 2016.

  1. The Generally Useful Estimate of Solvent Systems (GUESS) method enables the rapid purification of methylpyridoxine regioisomers by countercurrent chromatography

    PubMed Central

    Liu, Yang; Friesen, J. Brent; Klein, Larry L.; McAlpine, James B.; Lankin, David C.; Pauli, Guido F.; Chen, Shao-Nong

    2016-01-01

    The TLC-based Generally Useful Estimate of Solvent Systems (GUESS) method was employed for countercurrent chromatography solvent system selection, in order to separate the three synthetic isomers: 3-O-methylpyridoxine, 4′-O-methylpyridoxine (ginkgotoxin) and 5′-O-methylpyridoxine. The Rf values of the three isomers indicated that ChMWat +2 (chloroformmethanol-water 10:5:5, v/v/v) was appropriate for the countercurrent chromatography. The isomer separation was highly selective and demonstrated that the TLC-based GUESS method can accelerate solvent system selection for countercurrent chromatography. Accordingly, the study re-emphasizes the practicality of TLC as a tool to facilitate the rapid development of new countercurrent and centrifugal partition chromatography methods for this solvent system. Purity and structure characterization of all samples was performed by quantitative 1H NMR. PMID:26680272

  2. Chemistry and liquid chromatography methods for the analyses of primary oxidation products of triacylglycerols.

    PubMed

    Zeb, A

    2015-05-01

    Triacylglycerols (TAGs) are one of the major components of the cells in higher biological systems, which can act as an energy reservoir in the living cells. The unsaturated fatty acid moiety is the key site of oxidation and formation of oxidation compounds. The TAG free radical generates several primary oxidation compounds. These include hydroperoxides, hydroxides, epidioxides, hydroperoxy epidioxides, hydroxyl epidioxides, and epoxides. The presence of these oxidized TAGs in the cell increases the chances of several detrimental processes. For this purpose, several liquid chromatography (LC) methods were reported in their analyses. This review is therefore focused on the chemistry, oxidation, extraction, and the LC methods reported in the analyses of oxidized TAGs. The studies on thin-layer chromatography were mostly focused on the total oxidized TAGs separation and employ hexane as major solvent. High-performance LC (HPLC) methods were discussed in details along with their merits and demerits. It was found that most of the HPLC methods employed isocratic elution with methanol and acetonitrile as major solvents with an ultraviolet detector. The coupling of HPLC with mass spectrometry (MS) highly increases the efficiency of analysis as well as enables reliable structural elucidation. The use of MS was found to be helpful in studying the oxidation chemistry of TAGs and needs to be extended to the complex biological systems.

  3. Method transfer from high-pressure liquid chromatography to ultra-high-pressure liquid chromatography. II. Temperature and pressure effects.

    PubMed

    Åsberg, Dennis; Samuelsson, Jörgen; Leśko, Marek; Cavazzini, Alberto; Kaczmarski, Krzysztof; Fornstedt, Torgny

    2015-07-03

    The importance of the generated temperature and pressure gradients in ultra-high-pressure liquid chromatography (UHPLC) are investigated and compared to high-pressure liquid chromatography (HPLC). The drug Omeprazole, together with three other model compounds (with different chemical characteristics, namely uncharged, positively and negatively charged) were used. Calculations of the complete temperature profile in the column at UHPLC conditions showed, in our experiments, a temperature difference between the inlet and outlet of 16 °C and a difference of 2 °C between the column center and the wall. Through van't Hoff plots, this information was used to single out the decrease in retention factor (k) solely due to the temperature gradient. The uncharged solute was least affected by temperature with a decrease in k of about 5% while for charged solutes the effect was more pronounced, with k decreases up to 14%. A pressure increase of 500 bar gave roughly 5% increase in k for the uncharged solute, while omeprazole and the other two charged solutes gave about 25, 20 and 15% increases in k, respectively. The stochastic model of chromatography was applied to estimate the dependence of the average number of adsorption/desorption events (n) and the average time spent by a molecule in the stationary phase (τs) on temperature and pressure on peak shape for the tailing, basic solute. Increasing the temperature yielded an increase in n and decrease in τs which resulted in less skew at high temperatures. With increasing pressure, the stochastic modeling gave interesting results for the basic solute showing that the skew of the peak increased with pressure. The conclusion is that pressure effects are more pronounced for both retention and peak shape than the temperature effects for the polar or charged compounds in our study.

  4. Rapid method for determination of carbonyl groups in lignin compounds by headspace gas chromatography.

    PubMed

    Li, Jing; Hu, Hui-Chao; Chai, Xin-Sheng

    2015-07-24

    The paper reports on a novel method for rapid determination of carbonyl in lignins by headspace gas chromatography (HS-GC). The method involves the quantitative carbonyl reduction for aldehydes in 2min at room temperature or for acetones in 30min at 80°C by sodium borohydride solution in a closed headspace sample vial. After the reaction, the solution was acidified by injecting sulfuric acid solution and the hydrogen released to the headspace was determined by GC using thermal-conductivity detector. The results showed that with the addition of SiO2 powder, the reduction reaction of carbonyl groups can be greatly facilitated. The method has a good measurement precision (RSD<7.74%) and accuracy (relative error <10% compared with a reference method) in the carbonyl quantification. It is suitable to be used for rapid determination of carbonyl content in lignin and related materials.

  5. Rapid quantitative method for total brominated vegetable oil in soft drinks using ion chromatography.

    PubMed

    Yousef, Ashraf A; Abbas, Alaa B; Badawi, Bassam Sh; Al-Jowhar, Wafaa Y; Zain, Esam A; El-Mufti, Seham A

    2012-08-01

    A simple, quantitative and rapid method for total brominated vegetable oil (BVO) using ion chromatography (IC) with suppressed conductivity detection was developed and successfully applied to soft drinks with results expressed as inorganic bromide anion. The procedure involves extraction of BVO with diethyl ether and treatment with zinc dust in a solution of acetic acid, giving recoveries ranging between 92.5 and 98.5%. The calibration curves obtained were linear with correlation coefficients (r²) of 0.998, a coefficient of variation (CV) of less than 5% and limit of detection (LOD) and limit of quantification (LOQ) of 250 and 750 µg l⁻¹, respectively. The method was successfully applied to the determination of BVO in several commercial soft drinks which were found to contain BVO in the range 1.8-14.510 mg l⁻¹. The method has less sources of error compared to previously published methods.

  6. First screening method for the simultaneous detection of seven allergens by liquid chromatography mass spectrometry.

    PubMed

    Heick, J; Fischer, M; Pöpping, B

    2011-02-18

    The development of a multi-method for the detection of seven allergens based on liquid chromatography and triple-quadrupole tandem mass spectrometry in multiple reaction mode is described. It is based on extraction of the allergenic proteins from a food matrix, followed by enzymatic digestion with trypsin. The chosen marker peptides were implemented into one method that is capable of the simultaneous detection of milk, egg, soy, hazelnut, peanut, walnut and almond. This method has been used to detect all seven allergenic commodities from incurred reference bread material, which was baked according to a standard recipe from the baking industry. Detected concentrations ranged from 10 to 1000 μg/g, demonstrating that the mass spectrometric based method is a useful tool for allergen screening.

  7. Development of a size exclusion chromatography method for the determination of molar mass for poloxamers.

    PubMed

    Erlandsson, Bengt; Wittgren, Bengt; Brinkmalm, Gunnar

    2003-04-01

    An aqueous size exclusion chromatography (SEC) method for determination of the molar mass of poloxamers 188 and 407 has been developed as an alternative to the pharmacopoeia methods. During the development work two different columns and several different eluent compositions were investigated. With a PL-aquagel-OH column, non-exclusion behaviour was obtained. A TSKgel column gave good separation of both poloxamers. The best separation was obtained with an eluent consisting of sodium chloride (0.01 M)-methanol (90:10, v/v) on the TSKgel column. The method was shown to be linear within the elution time of the two poloxamers and to have acceptable precision. The results from the SEC method was compared to results obtained using SEC with online multi angle light scattering detection (MALS) and to results obtained with matrix-assisted laser desorption/ionisation mass spectrometry (MALDI-MS).

  8. Development and validation of a microemulsion electrokinetic chromatography method for patulin quantification in commercial apple juice.

    PubMed

    Murillo-Arbizu, M; González-Peñas, E; Hansen, S H; Amézqueta, S; Ostergaard, J

    2008-06-01

    A microemulsion electrokinetic chromatography (MEECK) method for patulin (PAT) quantification in apple juice samples has been developed. The effects of several important factors such as co-surfactant type, concentration of surfactant, acetonitrile percentage in the microemulsion, and running voltage and temperature were investigated to determine the optimum conditions. They resulted to be: a background electrolyte (BGE) composed of 25mM of sodium tetraborate, SDS (2.16%w/w), ethanol (6.49%w/w), n-octanol (0.82%w/w) and 2%v/v acetonitrile; applied voltage of +15kV; and a capillary temperature of 35 degrees C. PAT was detected at 276nm. Quantification and detection limits (LOQ and LOD) in apple juice samples were 8.0microgL(-1) and 3.2microgL(-1), respectively. Patulin was extracted from apple juice using ethyl acetate with a mean recovery value of 75.3% (RSD=4.5). This method was applied to the measurement of patulin in twenty commercial apple juice samples obtained from different Danish supermarkets. The PAT apple juice mean and median levels obtained were 35.9 and 10.9microgL(-1), respectively. The comparison with a previously validated micellar electrokinetic chromatography (MEKC) method for PAT analysis showed the suitability of using MEEKC for this mycotoxin analysis. However, the expectations of obtaining a higher efficiency and thus lower limits of detection and quantitation when using MEEKC were not met.

  9. Isotope dilution gas chromatography/mass spectrometry method for determination of pyrethroids in apple juice.

    PubMed

    Wong, Siu-kay; Yu, Kwok-chiu; Lam, Chi-ho

    2010-03-01

    This paper presents the development of a highly precise and accurate analytical method for the determination of three matrix-bound pyrethroids, namely, cypermethrin, permethrin, and bifenthrin, using an isotope dilution gas chromatography/mass spectrometry technique. Identification of the analytes was confirmed under selective ion monitoring mode by the presence of two dominant ion fragments within specific time windows and matching of relative ion intensities of the ions concerned in samples and calibration standards. Quantitation was based on the measurement of concentration ratios of the natural and isotope analogues in the sample and calibration blends. Intraday and interday repeatabilities of replicate analyses of the pyethroids in an apple juice sample were below 0.5%. The expanded relative uncertainty ranged from 3 to 6%, which was significantly lower than the range obtained using internal or external calibration methods. As a labeled analogue is not available for bifenthrin, bifenthrin was determined using labeled cis-permethrin as the internal standard. The results were counterchecked by a gas chromatography-electron capture detection technique using PCB 209 as the internal standard. The method developed was applied to a recent pilot study organized by CCQM and the results were consistent with those of other participants.

  10. Determination of vaporization enthalpies of the branched esters from correlation gas chromatography and transpiration methods

    SciTech Connect

    Verevkin, S.P.; Heintz, A.

    1999-12-01

    Vaporization enthalpies are indispensable for the assessment of the environmental fate and behavior of environmental contaminants. The temperature dependencies of retention indices of a set of 80 esters with branched molecular structures were measured on a nonpolar gas chromatographic column. The correlation gas chromatography method and reliable data set of 16 esters selected from the literature were used to derive a correlation for the prediction of the standard molar enthalpies of vaporization {Delta}{sub 1}{sup g}H{sub m}{sup {degree}} at the temperature T = 298.15 K. Experimental values of {Delta}{sub 1}{sup g}H{sub m}{sup {degree}} for 64 branched esters were obtained with the help of this correlation. The vaporization enthalpies of isopentyl acetate, ethyl hexanoate, and neopentyl pivalate were additionally obtained by the transpiration method from the temperature dependence of the vapor pressure measured in a flow system and used for checking the validity of the correlation gas chromatography method.

  11. Novel orthogonal liquid chromatography methods to dose neurotransmitters involved in Parkinson's disease.

    PubMed

    Sardella, Roccaldo; Scorzoni, Stefania; Conte, Carmela; Lisanti, Antonella; Ianni, Federica; Natalini, Benedetto

    2014-09-01

    Parkinson's disease is a multifactorial neurodegenerative disorder, characterized by a reduction of dopamine (DA) levels. The molecular pathways involved in the pathogenesis of disease have not yet been fully disclosed. Therefore, developing new diagnostic methods and tools to evaluate the depletion of DA and of some of its metabolites (3,4-dihydroxyphenylacetic acid, homovanillic acid, and 3-methoxytyramine) is of outstanding importance for biochemical evaluations. Moreover, neurons responsible for DA release also produce the neurotransmitter gamma-aminobutyric acid (GABA), thus, quantitative measurements of GABA levels can have a relevant impact for a further understanding of the biochemical processes involved in the neurodegenerative event. In the present study, two HPLC methods based on the reversed-phase ion-pairing chromatography (RP-IPC) and the hydrophilic liquid interaction chromatography (HILIC) concepts were developed to allow the quantification of DA and its metabolites as well as GABA levels in mouse striatal and cortical tissue homogenates. The two fairly orthogonal HPLC methods were directly applied to the biological samples, without preliminary derivatization of the compounds of interest. A high level of selectivity was obtained for DA metabolites and GABA by running the gradient RP-IPC method with a volatile ion-pairing reagent, which makes it suitable for the quantitative assay of four out of five compounds. Matrix deriving interferences unabled the base-line separation of DA which was instead successfully achieved with the HILIC-based method. To avail of HPLC methods providing distinct selectivity profiles, makes possible the correct species quantification and allows to compensate the intrinsic limits characterizing all chromatographic methods.

  12. A validated high performance thin layer chromatography method for determination of yohimbine hydrochloride in pharmaceutical preparations

    PubMed Central

    Badr, Jihan M.

    2013-01-01

    Background: Yohimbine is an indole alkaloid used as a promising therapy for erectile dysfunction. A number of methods were reported for the analysis of yohimbine in the bark or in pharmaceutical preparations. Materials and Method: In the present work, a simple and sensitive high performance thin layer chromatographic method is developed for determination of yohimbine (occurring as yohimbine hydrochloride) in pharmaceutical preparations and validated according to International Conference of Harmonization (ICH) guidelines. The method employed thin layer chromatography aluminum sheets precoated with silica gel as the stationary phase and the mobile phase consisted of chloroform:methanol:ammonia (97:3:0.2), which gave compact bands of yohimbine hydrochloride. Results: Linear regression data for the calibration curves of standard yohimbine hydrochloride showed a good linear relationship over a concentration range of 80–1000 ng/spot with respect to the area and correlation coefficient (R2) was 0.9965. The method was evaluated regarding accuracy, precision, selectivity, and robustness. Limits of detection and quantitation were recorded as 5 and 40 ng/spot, respectively. The proposed method efficiently separated yohimbine hydrochloride from other components even in complex mixture containing powdered plants. The amount of yohimbine hydrochloride ranged from 2.3 to 5.2 mg/tablet or capsule in preparations containing the pure alkaloid, while it varied from zero (0) to 1.5–1.8 mg/capsule in dietary supplements containing powdered yohimbe bark. Conclusion: We concluded that this method employing high performance thin layer chromatography (HPTLC) in quantitative determination of yohimbine hydrochloride in pharmaceutical preparations is efficient, simple, accurate, and validated. PMID:23661986

  13. Development of high performance liquid chromatography method for miconazole analysis in powder sample

    NASA Astrophysics Data System (ADS)

    Hermawan, D.; Suwandri; Sulaeman, U.; Istiqomah, A.; Aboul-Enein, H. Y.

    2017-02-01

    A simple high performance liquid chromatography (HPLC) method has been developed in this study for the analysis of miconazole, an antifungal drug, in powder sample. The optimized HPLC system using C8 column was achieved using mobile phase composition containing methanol:water (85:15, v/v), a flow rate of 0.8 mL/min, and UV detection at 220 nm. The calibration graph was linear in the range from 10 to 50 mg/L with r 2 of 0.9983. The limit of detection (LOD) and limit of quantitation (LOQ) obtained were 2.24 mg/L and 7.47 mg/L, respectively. The present HPLC method is applicable for the determination of miconazole in the powder sample with a recovery of 101.28 % (RSD = 0.96%, n = 3). The developed HPLC method provides short analysis time, high reproducibility and high sensitivity.

  14. Practical method for the definition of chromatographic peak parameters in preparative liquid chromatography.

    PubMed

    Jin, Gaowa; Guo, Zhimou; Xiao, Yuansheng; Yan, Jingyu; Dong, Xuefang; Shen, Aijin; Wang, Chaoran; Liang, Xinmiao

    2016-10-01

    A practical method was established for the definition of chromatographic parameters in preparative liquid chromatography. The parameters contained both the peak broadening level under different amounts of sample loading and the concentration distribution of the target compound in the elution. The parameters of the peak broadening level were defined and expressed as a matrix, which consisted of sample loading, the forward broadening and the backward broadening levels. The concentration distribution of the target compound was described by the heat map of the elution profile. The most suitable stationary phase should exhibit the narrower peak broadening and it was best to broaden to both sides to compare to the peak under analytical conditions. Besides, the concentration distribution of the target compounds should be focused on the middle of the elution. The guiding principles were validated by purification of amitriptyline from the mixture of desipramine and amitriptyline. On the selected column, when the content of the impurity desipramine was lower than 0.1%, the recovery of target compound was much higher than the other columns even when the sample loading was as high as 8.03 mg/cm(3) . The parameters and methods could be used for the evaluation and selection of stationary phases in preparative chromatography.

  15. A simple and simultaneous identification method for aloe, catechu and gambir by high performance liquid chromatography.

    PubMed

    Zhao, Yan; Kim, Young Ho; Lee, Wonjae; Lee, Young Keun; Kim, Kyung Tae; Kang, Jong Seong

    2016-01-05

    An effective and rapid method was developed for the simultaneous identification of aloe, catechu and gambir by high performance liquid chromatography-diode array detector (HPLC-DAD). Identification of three maker compounds presented in three medicinal materials was performed on high performance liquid chromatography-mass spectrometry (HPLC-MS). Under the optimal HPLC chromatographic conditions, sixty-two samples were processed on an Optimapak C18 column using a solvent system of acetonitrile (from 10% to 35%) and 0.1% phosphoric acid solution (from 90% to 65%) at a total flow rate of 1.0 mL/min and detected at 270 nm. All calibration curves exhibited good linear relationship (r(2)>0.9992). The relative standard deviation values of intra-day and inter-day precision were less than 1% and 2%, respectively. The recoveries of three analytes ranged from 99.48 to 100.97% with low RSDs (<2%). For the first time, this study demonstrates that the processed aloe, catechu and gambir are sold in local material markets in China and Korea without their correct identification. It indicates the existent of high potential medicinal risk by misuse of three medicinal materials. The developed HPLC method can be applied to prevent unexpected biological activity due to misapplication of medicinal materials.

  16. On-line coupled high performance liquid chromatography-gas chromatography for the analysis of contamination by mineral oil. Part 1: method of analysis.

    PubMed

    Biedermann, Maurus; Grob, Koni

    2012-09-14

    For the analysis of mineral oil saturated hydrocarbons (MOSH) and mineral oil aromatic hydrocarbons (MOAH), on-line coupled high performance liquid chromatography-gas chromatography-flame ionization detection (HPLC-GC-FID) offers important advantages: it separates MOSH and MOAH in robust manner, enables direct injection of large aliquots of raw extracts (resulting in a low detection limit), avoids contamination of the sample during preparation and is fully automated. This review starts with an overview of the technology, particularly the fundamentals of introducing large volumes of solvent into GC, and their implementation into various transfer techniques. The main part deals with the concepts of MOSH and MOAH analysis, with a thorough discussion of the choices made. It is followed by a description of the method. Finally auxiliary tools are summarized to remove interfering components, enrich the sample in case of a high fat content and obtain additional information about the MOSH and MOAH composition.

  17. Quantification of PSP toxins in toxic shellfish matrices using post-column oxidation liquid chromatography and pre-column oxidation liquid chromatography methods suggests post-column oxidation liquid chromatography as a good monitoring method of choice.

    PubMed

    Rey, Verónica; Botana, Ana M; Botana, Luis M

    2017-04-01

    Different shellfish samples were analyzed by Pre- and Post-Column Oxidation Liquid Chromatography to compare the toxins profiles and get information about the degree of accomplishment of both methods. Comparison of the results obtained, the linear correlation coefficient (r(2) = 0.94) and the paired t test (two tails, α = 0.05), indicated that there were not significant differences between both sets of data. Nevertheless, important differences related to toxins profiles were found: it was remarkable the difference in results for both Gonyautoxins 1 and 4 and Decarbamoylgonyautoxins 2 and 3, depending on the method of choice, due to an overestimation in the Pre-Column method. It was necessary to modify the elution conditions in the Post-Column method to avoid the interference of matrix peaks at retention times closer to the retention times of the calibrants, mostly when working with oyster and scallop matrices, although it is a good method to use routinely.

  18. Gas chromatography of safranal as preferable method for the commercial grading of saffron (Crocus sativus L.).

    PubMed

    Bononi, Monica; Milella, Paola; Tateo, Fernando

    2015-06-01

    We present a new extraction protocol, using ethyl alcohol as a solvent, to evaluate safranal by gas chromatography (GC). A linear response was obtained with R(2)=0.995 and a reproducibility standard deviation of 4.7-6.0%. The limit of detection and limit of quantitation were 0.05 and 0.25gkg(-1), respectively. The GC data for several samples of powdered saffron from different origins were compared to specific absorbance values measured according to the ISO Normative 3632-1:2011 method. The aroma strength of saffron samples quantitated by GC and the specific absorbance values of safranal by the UV method did not correlate. Quantitative evaluation of safranal by GC appears to be more specific and useful for commercial comparisons of saffron quality.

  19. A high performance liquid chromatography method for determination of furfural in crude palm oil.

    PubMed

    Loi, Chia Chun; Boo, Huey Chern; Mohammed, Abdulkarim Sabo; Ariffin, Abdul Azis

    2011-09-01

    A modified steam distillation method was developed to extract furfural from crude palm oil (CPO). The collected distillates were analysed using high performance liquid chromatography (HPLC) coupled with an ultraviolet diode detector at 284nm. The HPLC method allowed identification and quantification of furfural in CPO. The unique thermal extraction of CPO whereby the fresh fruit bunches (FFB) are first subjected to steam treatment, distinguishes itself from other solvent-extracted or cold-pressed vegetable oils. The presence of furfural was also determined in the fresh palm oil from FFB (without undergoing the normal extraction process), palm olein, palm stearin, olive oil, coconut oil, sunflower oil, soya oil and corn oil. The chromatograms of the extracts were compared to that of standard furfural. Furfural was only detected in CPO. The CPO consignments obtained from four mills were shown to contain 7.54 to 20.60mg/kg furfural.

  20. A novel thin-layer chromatography method to screen 1,3-propanediol producers.

    PubMed

    Anand, Pinki; Saxena, Rajendra Kumar

    2012-11-01

    To date, there is no established protocol for the screening of 1,3-propanediol producers. The proposed method has a wide applicability to harness the commercial potential of microorganisms which produce 1,3-propanediol as the end product. Glycerol fermentation broth of 50 bacteria spotted on thin-layer chromatography plates and run by appropriate solvent systems followed by colour development using vanillin reagent gave different coloured spots with most of the compounds present in the fermentation broth. The appearance of a purple-coloured spot of 1,3-propanediol with a retention factor (R(f)) of 0.62 forms the basis for the selection of 1,3-propanediol producers. Apart from being a rapid detection system the proposed method is pH independent and its authenticity was reconfirmed by HPLC.

  1. [Methods of nanoparticles control in food and biological objects. Report 1. Use of microscopic and chromatography investigation methods].

    PubMed

    Raspopov, R V; Gmoshinskiĭ, I V; Popov, K I; Krasnoiarova, O V; Khotimchenko, S A

    2012-01-01

    One of the promising applications of modern nanotechnology are food productions, which includes the improvement of food packaging, creation of new forms of nutrients that are characterized by improved assimilation and technological characteristics, quality control through the creation of compact and cheap test kits. All these applications of nanomaterials related to the risks of the possibility of receipt of potentially toxic nanoparticles in the diet. The task of regulation and hygienic standardization requires developing of the methods, their qualitative and quantitative analysis for such complex, multicomponent systems, which are the agricultural commodities and food products. The best hope in this plan are assigned to a group of approaches related to the microscopic visualization of artificial nanoparticles in the biological objects. While the typical size of nanoparticles (<100 nm) are below the theoretical maximum-resolution light optical methods, transmission electron microscopy often allows not only to identify nanoparticles on their size and shape, but also a qualitative and quantitative analysis their chemical composition with the use of additional analytical options. Another group of elaborate methods used in solving the problems of qualitative and quantitative analysis of nanoparticles are chromatographic methods, in particular, the exclusion, hydrodynamic, high-performance liquid chromatography, and the flow-field fractionation. Limitation of chromatographic approaches related with the need of complex sample preparation, as well as specific difficulties in nanoparticles detecting in chromatographic fractions. Transmission electron microscopy and high-performance liquid chromatography methods are officially recommended in Russia for the analysis of artificial nanoparticles in natural biological systems, including food products.

  2. High-performance thin-layer chromatography method for inositol phosphate analysis.

    PubMed

    Hatzack, F; Rasmussen, S K

    1999-12-24

    A simple and inexpensive high-performance thin-layer chromatography (HPTLC) method for the analysis of inositol mono- to hexakisphosphates on cellulose precoated plates is described. Plates were developed in 1-propanol-25% ammonia solution-water (5:4:1) and substance quantities as low as 100-200 pmol were detected by molybdate staining. Chromatographic mobilities of nucleotides and phosphorylated carbohydrates were also characterized. Charcoal treatment was employed to separate nucleotides from inositol phosphates with similar R(F) values prior to HPTLC analysis. Practical application of the HPTLC system is demonstrated by analysis of grain extracts from wild type and low-phytate mutant barley as well as phytate degradation products resulting from barley phytase activity.

  3. Hydrophilic interaction liquid chromatography method development and validation for the assay of HEPES zwitterionic buffer.

    PubMed

    Xu, Xiaolong; Gevaert, Bert; Bracke, Nathalie; Yao, Han; Wynendaele, Evelien; De Spiegeleer, Bart

    2017-02-20

    HEPES is a zwitterionic buffer component used as a raw material in the GMP-manufacturing of advanced therapy medicinal products (ATMPs), hence requiring an adequate assay method with sufficient selectivity toward related impurities. Therefore, a hydrophilic interaction chromatography (HILIC) method was developed. Different factors were investigated towards the retention behavior of HEPES, its analogue EPPS and its starting material isethionate: pH, ion concentration and organic solvent ratio of the mobile phase, as well as column temperature. Moreover, stress testing resulted in the N-oxide degradant, identified by high resolution MS. The final method consisted of an isocratic system with an aqueous (pH 2.0 with H3PO4) acetonitrile (35:65, v/v) mobile phase on a zwitterionic HILIC (Obelisc N) column with a flow rate of 0.5mL/min and UV detection at 195nm. The assay method of HEPES was validated, obtaining adequate linearity (R(2)=0.999), precision (RSD of 0.5%) and accuracy (recovery of 100.08%). Finally, the applicability of the validated method was demonstrated by analysis of samples from different suppliers.

  4. Comprehensive measurement of total nondigestible carbohydrates in foods by enzymatic-gravimetric method and liquid chromatography.

    PubMed

    Nishibata, Toyohide; Tashiro, Kouichi; Kanahori, Sumiko; Hashizume, Chieko; Kitagawa, Machiko; Okuma, Kazuhiro; Gordon, Dennis T

    2009-09-09

    Total nondigestible carbohydrate (NDC) in foods was determined by combining, not modifications, AOAC Official Methods 991.43, 2001.03, and 2002.02. Total NDC included insoluble dietary fiber (IDF) + high-molecular-weight soluble dietary fiber (HMWSDF), nondigestible oligosaccharides (NDO) not precipitated in ethanol solution, and resistant starch (RS). Eight sources of NDC (cellulose, wheat bran, gum arabic, resistant maltodextrin, polydextrose, fructooligosaccharide, galactooligosaccharides, and RS) were incorporated in different combinations into standard formula bread samples. All of the NDC sources and bread samples were analyzed for their (1) IDF + HMWSDF content with corrections for residual RS amount using AOAC Official Method 991.43, (2) NDO by liquid chromatography (LC) in AOAC Official Method 2001.03, and (3) RS by AOAC Official Method 2002.02. The correlation coefficient (R(2)) comparing calculated amounts versus measured amounts of total NDC in 11 bread samples was 0.92. Analysis of commercial food samples was also well matched with the DF + NDO value on their nutritional label. Consequently, we confirmed a single measurement of LC can determine all NDO in foods, and total NDC in foods can be determined by unifying existing AOAC Official Methods.

  5. Ceftriaxone pharmacokinetics by new simple and sensitive ultra-high-performance liquid chromatography method.

    PubMed

    Campos, Michel Leandro; de Moura Alonso, Juliana; Dos Santos Martins, Evelin; Oliveira, Jonata Augusto; Hussni, Carlos Alberto; Peccinini, Rosângela Gonçalves

    2017-02-27

    Ceftriaxone is a cephalosporin antibiotic with a potent antimicrobial activity and excellent penetration in most body fluids such as pleural, peritoneal, spinal and brain. These facts contribute to the application of ceftriaxone in the treatment of bacterial peritonitis, an abdominal disorder in veterinary medicine, with potential risk of death. The determination of ceftriaxone levels in plasma and peritoneal fluid may be used to assess the pharmacokinetic profile at various instances of administration and allows observing if the concentrations needed are being achieved. Therefore a method was developed and validated for the determination of ceftriaxone in plasma and peritoneal fluid which after was applied in a pharmacokinetic profile study. The bioanalytical method validation was performed according to widely acceptable experiments. Two horses were used as a model of the method applicability; ceftriaxone was intraperitoneally administered to these animals as a single dose. The plasma and peritoneal fluid analysis were performed using an UHPLC system in reverse phase chromatography mode in fully validated conditions. The methods have shown linearity between 0.49 and 500μg/mL for plasma, and between 0.24 and 500μg/mL for peritoneal fluid. The quantitative analysis of ceftriaxone in these matrices allows monitoring of the therapy. This method showed improved sensitivity as well as the quantitation in peritoneal fluid.

  6. Determination of hormonal growth promoters in bovine hair: comparison of liquid chromatography-mass spectrometry and gas chromatography-mass spectrometry methods for estradiol benzoate and nortestosterone decanoate.

    PubMed

    Duffy, Eleanor; Rambaud, Lauriane; Le Bizec, Bruno; O'Keeffe, Michael

    2009-04-01

    The detection of steroid residues in hair is a powerful strategy to demonstrate long-term administration of these growth promoters in meat production animals. Analysis of the ester form of administered steroids is an unambiguous approach to prove the illegal use of natural hormones. For detection, gas chromatography-mass spectrometry (GC-MS/MS) was generally used. However, recent advances in liquid chromatography-tandem mass spectrometry (LC-MS/MS) technology have improved the robustness and potential sensitivity of this method. This paper describes development and validation, according to Commission Decision 2002/657/EC, of LC-MS/MS and GC-MS/MS methods, in two separate laboratories, for determination of steroid esters in bovine hair. Bovine hair samples taken from animals treated with estradiol-3-benzoate and nortestosterone decanoate, as well as from an untreated animal, were used to evaluate the comparability of the results of the two validated methods. The results of the inter-comparison demonstrate that both the LC-MS/MS and the GC-MS/MS methods are fit for purpose and capable of determining steroid esters in hair samples from treated bovine animals.

  7. Assessment of the chemical changes during storage of phenol-formaldehyde resins pyrolysis gas chromatography mass spectrometry, inverse gas chromatography and Fourier transform infra red methods.

    PubMed

    Strzemiecka, B; Voelkel, A; Zięba-Palus, J; Lachowicz, T

    2014-09-12

    The chemical changes occurring in the phenol-formaldehyde resins (resol and novolac type) during their storage were investigated. In this paper the FT-IR, py-GCMS and inverse gas chromatography methods were applied for assessment of the changes occurring during storage of the phenolic resins. We have found that during storage some examined resins occurred partial curing. The results from all techniques applied are consistent. Py-GCMS is useful technique for screening the storage processes but IGC seems to be most sensitive one.

  8. Modern methods for vancomycin determination in biological fluids by methods based on high-performance liquid chromatography--A review.

    PubMed

    Javorska, Lenka; Krcmova, Lenka Kujovska; Solichova, Dagmar; Solich, Petr; Kaska, Milan

    2016-01-01

    Vancomycin is a glycopeptide antibiotic used in the therapy of severe bacterial infection. The monitoring of vancomycin levels is recommended because of its narrow therapeutic index and toxicity. This measurement is especially appropriate in patients with unstable renal functions, who receive high doses of vancomycin or present serious bacterial infections accompanied by important sequestration of liquids when it could be difficult to achieve the optimal therapeutic dose. Most of the methods for vancomycin determination in routine practice are immunoassays. However, chromatography-based techniques in combination with UV or mass spectrometry detection provide results with greater accuracy and precision also in complicated biological matrices. This review provides a detailed overview of modern approaches for the chromatographic separation of vancomycin in various biological samples and useful sample preparation procedures for vancomycin determination in various biological fluids.

  9. Integration of Gas Chromatography Mass Spectrometry Methods for Differentiating Ricin Preparation Methods

    SciTech Connect

    Wunschel, David S.; Melville, Angela M.; Ehrhardt, Christopher J.; Colburn, Heather A.; Victry, Kristin D.; Antolick, Kathryn C.; Wahl, Jon H.; Wahl, Karen L.

    2012-05-17

    The investigation of crimes involving chemical or biological agents is infrequent, but presents unique analytical challenges. The protein toxin ricin is encountered more frequently than other agents and is found in the seeds of the castor plant Ricinus communis. Typically, the toxin is extracted from castor seeds utilizing a variety of different recipes that result in varying purity of the toxin. Moreover, these various purification steps can also leave or differentially remove a variety of exogenous and endogenous residual components with the toxin that may indicate the type and number of purification steps involved. We have applied three gas chromatographic - mass spectrometric (GC-MS) based analytical methods to measure the variation in seed carbohydrates and castor oil ricinoleic acid as well as the presence of solvents used for purification. These methods were applied to the same samples prepared using four previously identified toxin preparation methods starting from four varieties of castor seeds. The individual data sets for seed carbohydrate profiles, ricinoleic acid or acetone amount each provided information capable of differentiating different types of toxin preparations across seed types. However, the integration of the data sets using multivariate factor analysis provided a clear distinction of all samples based on the preparation method and independent of the seed source. In particular the abundance of mannose, arabinose, fucose, ricinoleic acid and acetone were shown to be important differentiating factors. These complementary tools provide a more confident determination of the method of toxin preparation.

  10. Profiling methods to identify cold-regulated primary metabolites using gas chromatography coupled to mass spectrometry.

    PubMed

    Dethloff, Frederik; Erban, Alexander; Orf, Isabel; Alpers, Jessica; Fehrle, Ines; Beine-Golovchuk, Olga; Schmidt, Stefanie; Schwachtje, Jens; Kopka, Joachim

    2014-01-01

    This book chapter describes the analytical procedures required for the profiling of a metabolite fraction enriched for primary metabolites. The profiling is based on routine gas chromatography coupled to mass spectrometry (GC-MS). The generic profiling method is adapted to plant material, specifically to the analysis of single leaves from plants that were exposed to temperature stress experiments. The described method is modular. The modules include a rapid sampling and metabolic inactivation protocol for samples in a wide size range, a sample extraction procedure, a chemical derivatization step that is required to make the metabolite fraction amenable to gas chromatographic analysis, a routine GC-MS method, and finally the procedures of data processing and data mining. A basic and extendable set of standardizations for metabolite recovery and retention index alignment of the resulting GC-MS chromatograms is included. The method has two applications: (1) the rapid screening for changes of relative metabolite pools sizes under temperature stress and (2) the verification of cold-regulated metabolites by exact quantification using a GC-MS protocol with extended internal and external standardization.

  11. Histamine H1 receptor cell membrane chromatography online high-performance liquid chromatography with mass spectrometry method reveals houttuyfonate as an activator of the histamine H1 receptor.

    PubMed

    Guo, Ying; Han, Shengli; Cao, Jingjing; Zhang, Tao; He, Langchong

    2014-11-01

    Allergy is an abnormal reaction of the body to an allergen. Histamine is responsible for many of the acute symptoms of allergic diseases. Many of the allergic and inflammatory actions of histamine are mediated by the histamine H1 receptor. In the present study, we established a two-dimensional histamine H1 receptor/cell membrane chromatography with online high-performance liquid chromatography and mass spectrometry method for screening potential histamine-activating components in a traditional Chinese medicine injection. The specification of the method was validated by screening, separating, and identifying a mixed standard solution of diphenhydramine hydrochloride, gefitinib, tamsulosin, and nitrendipine. The Yujin injection, an example of traditional Chinese medicine injection, was screened and potential allergic components acting on the histamine H1 receptor were identified. A Ca(2+) flux assay showed that houttuyfonate and Yujin injection induced calcium release in a dose-dependent manner. This suggests that houttuyfonate is an activator of the histamine H1 receptor. The mechanism of houttuyfonate activation involves phosphorylation of the inositol-1,4,5-trisphosphate receptor. In conclusion, this two-dimensional method can rapidly detect and enrich target components isolated from the Yujin injection. This indicates that individuals with an overexpression of the histamine H1 receptor should be aware of possible allergic reactions when receiving the Yujin injection.

  12. Determination of nucleosides in Cordyceps sinensis and Ganoderma lucidum by high performance liquid chromatography method

    PubMed Central

    Khan, Masood Shah; Parveen, Rabea; Mishra, Kshipra; Tulsawani, Rajkumar; Ahmad, Sayeed

    2015-01-01

    Background: Nucleosides are supportive in the regulation and modulation of various physiological processes in body, they acts as precursors in nucleic acid synthesis, enhance immune response, help in absorption of iron and influence the metabolism of fatty acids. Cordyceps sinensis and Ganoderma lucidum are well-known for its use in traditional medicine of China, Nepal and India. They are rich in nucleosides such as adenine, adenosine, cordycepin, etc. Hence, a simple, economic and accurate high-performance liquid chromatography (HPLC) analytical method was proposed for determination of adenine and adenosine for the quality control of plants. Materials and Methods: Chromatographic experiments were conducted on YL9100 HPLC system (South Korea). Reversed-phase chromatography was performed on a C18 column with methanol and dihydrogen phosphate as the mobile phase in isocratic elution method at a flow rate of 1.0 mL/min. Detection was carried out at 254 nm, which gives a sharp peak of adenine and adenosine at a retention time of 6.53 ± 0.02 min and 12.41 ± 0.02, respectively. Results and Discussion: Linear regression analysis data for the calibration plot showed a good linear relationship between response and concentration in the range of 25–200 µg/mL for adenosine and 100–800 µg/mL for adenine with regression coefficient of 0.999 and 0.996, respectively. The adenine was found 0.16% and 0.71% w/w in G. lucidum and in C. sinensis, respectively, and adenosine was found to be 0.14% w/w in G. lucidum whereas absent in C. sinensis. Conclusion: The developed HPLC method for the quantification of adenosine and adenine can be used for the quality control and standardization of crude drug and for the different herbal formulations, in which adenine and adenosine are present as major constituents. The wide linearity range, sensitivity, accuracy, and simple mobile phase imply the method is suitable for routine quantification of adenosine and adenine with high precision and

  13. Development and validation of high-performance liquid chromatography and high-performance thin-layer chromatography methods for the quantification of khellin in Ammi visnaga seed

    PubMed Central

    Kamal, Abid; Khan, Washim; Ahmad, Sayeed; Ahmad, F. J.; Saleem, Kishwar

    2015-01-01

    Objective: The present study was used to design simple, accurate and sensitive reversed phase-high-performance liquid chromatography RP-HPLC and high-performance thin-layer chromatography (HPTLC) methods for the development of quantification of khellin present in the seeds of Ammi visnaga. Materials and Methods: RP-HPLC analysis was performed on a C18 column with methanol: Water (75: 25, v/v) as a mobile phase. The HPTLC method involved densitometric evaluation of khellin after resolving it on silica gel plate using ethyl acetate: Toluene: Formic acid (5.5:4.0:0.5, v/v/v) as a mobile phase. Results: The developed HPLC and HPTLC methods were validated for precision (interday, intraday and intersystem), robustness and accuracy, limit of detection and limit of quantification. The relationship between the concentration of standard solutions and the peak response was linear in both HPLC and HPTLC methods with the concentration range of 10–80 μg/mL in HPLC and 25–1,000 ng/spot in HPTLC for khellin. The % relative standard deviation values for method precision was found to be 0.63–1.97%, 0.62–2.05% in HPLC and HPTLC for khellin respectively. Accuracy of the method was checked by recovery studies conducted at three different concentration levels and the average percentage recovery was found to be 100.53% in HPLC and 100.08% in HPTLC for khellin. Conclusions: The developed HPLC and HPTLC methods for the quantification of khellin were found simple, precise, specific, sensitive and accurate which can be used for routine analysis and quality control of A. visnaga and several formulations containing it as an ingredient. PMID:26681890

  14. The analysis of carbohydrates in milk powder by a new "heart-cutting" two-dimensional liquid chromatography method.

    PubMed

    Ma, Jing; Hou, Xiaofang; Zhang, Bing; Wang, Yunan; He, Langchong

    2014-03-01

    In this study, a new"heart-cutting" two-dimensional liquid chromatography method for the simultaneous determination of carbohydrate contents in milk powder was presented. In this two dimensional liquid chromatography system, a Venusil XBP-C4 analysis column was used in the first dimension ((1)D) as a pre-separation column, a ZORBAX carbohydrates analysis column was used in the second dimension ((2)D) as a final-analysis column. The whole process was completed in less than 35min without a particular sample preparation procedure. The capability of the new two dimensional HPLC method was demonstrated in the determination of carbohydrates in various brands of milk powder samples. A conventional one dimensional chromatography method was also proposed. The two proposed methods were both validated in terms of linearity, limits of detection, accuracy and precision. The comparison between the results obtained with the two methods showed that the new and completely automated two dimensional liquid chromatography method is more suitable for milk powder sample because of its online cleanup effect involved.

  15. A novel fast gas chromatography method for higher time resolution measurements of speciated monoterpenes in air

    NASA Astrophysics Data System (ADS)

    Jones, C. E.; Kato, S.; Nakashima, Y.; Kajii, Y.

    2014-05-01

    Biogenic emissions supply the largest fraction of non-methane volatile organic compounds (VOC) from the biosphere to the atmospheric boundary layer, and typically comprise a complex mixture of reactive terpenes. Due to this chemical complexity, achieving comprehensive measurements of biogenic VOC (BVOC) in air within a satisfactory time resolution is analytically challenging. To address this, we have developed a novel, fully automated Fast Gas Chromatography (Fast-GC) based technique to provide higher time resolution monitoring of monoterpenes (and selected other C9-C15 terpenes) during plant emission studies and in ambient air. To our knowledge, this is the first study to apply a Fast-GC based separation technique to achieve quantification of terpenes in ambient air. Three chromatography methods have been developed for atmospheric terpene analysis under different sampling scenarios. Each method facilitates chromatographic separation of selected BVOC within a significantly reduced analysis time compared to conventional GC methods, whilst maintaining the ability to quantify individual monoterpene structural isomers. Using this approach, the C9-C15 BVOC composition of single plant emissions may be characterised within a 14.5 min analysis time. Moreover, in-situ quantification of 12 monoterpenes in unpolluted ambient air may be achieved within an 11.7 min chromatographic separation time (increasing to 19.7 min when simultaneous quantification of multiple oxygenated C9-C10 terpenoids is required, and/or when concentrations of anthropogenic VOC are significant). These analysis times potentially allow for a twofold to fivefold increase in measurement frequency compared to conventional GC methods. Here we outline the technical details and analytical capability of this chromatographic approach, and present the first in-situ Fast-GC observations of 6 monoterpenes and the oxygenated BVOC (OBVOC) linalool in ambient air. During this field deployment within a suburban forest

  16. Gas chromatography-mass spectrometric method for metabolic profiling of tobacco leaves.

    PubMed

    Li, Yong; Pang, Tao; Li, Yanli; Wang, Xiaolin; Li, Qinghua; Lu, Xin; Xu, Guowang

    2011-06-01

    A gas chromatography-mass spectrometric method was developed for profiling of tobacco leaves. The differentiation among tobacco leaves planted in two different regions was investigated. Prior to analysis, the extraction solvent formulation was optimized and a combination of water, methanol and acetonitrile with a volume ratio of 3:1:1 was found to be optimal. The reproducibility of the method was satisfactory. Kendall tau-b rank correlation coefficients were equal to 1 (p<0.05) for 82% of the resolved peaks (up to 95% of the overall peak areas), indicating the good response correlation. Forty-four compounds including 9 saccharides, 9 alcohols, 9 amino acids, 16 organic acids and phosphoric acid were identified based on standard compounds. The method was successfully applied for profiling of tobacco leaves from Zimbabwe and Yunnan of China. Our result revealed that levels of saccharides and their derivatives including xylose, ribose, fructose, glucose, turanose, xylitol and glyceric acid were more abundant while sucrose, glucitol and D-gluconic acid were less abundant in tobacco leaves from Yunnan as compared to those from Zimbabwe. Amino acids such as L-alanine, L-tyrosine and L-threonine were found to be richer in Zimbabwe tobacco than in Yunnan tobacco.

  17. Development of a new method for the determination of nitrosamines by micellar electrokinetic capillary chromatography.

    PubMed

    Filho, Pedro José Sanches; Rios, Angel; Valcárcel, Miguel; Caramao, Elina Bastos

    2003-09-01

    A new method was developed for separation and quantification of nitrosamines by micellar electrokinetic capillary chromatography (MEKC). The effects of composition of the buffer, concentration of sodium dodecyl sulfate (SDS), gamma-cyclodextrin (CD) and pH on the separation and migration times of the nitrosamines were investigated. The different instrumental variables affecting sensitivity and resolution, such as power supply, injection mode and wavelength for measurement, were carefully optimized. The direct application of this method to aqueous synthetic samples allowed the separation and determination of nitrosamine mixtures at the mg x L(-1) level. The calibration curves revealed a linear range between 0.25 and 10 mg x L(-1), with detection limit (DL) varying from 0.16 to 0.27 mg x L(-1), quantification limit (QL) varying from 0.52 to 0.90 mg x L(-1), and RSD relative standard deviation (RSD) from 4.5% to 8.5%. The proposed method was successfully applied to the determination of dimethylnitrosamine (DMN), diethylnitrosamine (DEN), N-nitrosopyrrolidine (NPYR), N-nitrosomorpholine (NMOR) and N-nitrosopiperidine (NPIP).

  18. Integrated liquid chromatography method in enantioselective studies: Biodegradation of ofloxacin by an activated sludge consortium.

    PubMed

    Maia, Alexandra S; Castro, Paula M L; Tiritan, Maria Elizabeth

    2016-09-01

    Ofloxacin is a chiral fluoroquinolone commercialized as racemate and as its enantiomerically pure form levofloxacin. This work presents an integrated liquid chromatography (LC) method with fluorescence detection (FD) and exact mass spectrometry (EMS) developed to assess the enantiomeric biodegradation of ofloxacin and levofloxacin in laboratory-scale microcosms. The optimized enantioseparation conditions were achieved using a macrocyclic antibiotic ristocetin A-bonded CSP (150×2.1mm i.d.; particle size 5μm) under reversed-phase elution mode. The method was validated using a mineral salts medium as matrix and presented selectivity and linearity over a concentration range from 5μgL(-1) (quantification limit) to 350μgL(-1) for each enantiomer. The method was successfully applied to evaluate biodegradation of ofloxacin enantiomers at 250μgL(-1) by an activated sludge inoculum. Ofloxacin (racemic mixture) and (S)-enantiomer (levofloxacin) were degraded up to 58 and 52%, respectively. An additional degradable carbon source, acetate, enhanced biodegradation up to 23%. (S)-enantiomer presented the highest extent of degradation (66.8%) when ofloxacin was supplied along with acetate. Results indicated slightly higher biodegradation extents for the (S)-enantiomer when supplementation was done with ofloxacin. Degradation occurred faster in the first 3days and proceeded slowly until the end of the assays. The chromatographic results from LC-FD suggested the formation of the (R)-enantiomer during levofloxacin biodegradation which was confirmed by LC-MS with a LTQ Orbitrap XL.

  19. Capture chromatography for Mo-99 recovery from uranyl sulfate solutions: minimum-column-volume design method.

    PubMed

    Ling, Lei; Chung, Pei-Lun; Youker, Amanda; Stepinski, Dominique C; Vandegrift, George F; Wang, Nien-Hwa Linda

    2013-09-27

    Molybdenum-99 (Mo-99), generated from the fission of Uranium-235 (U-235), is the radioactive parent of the most widely used medical isotope, technetium-99m (Tc-99m). An efficient, robust, low-pressure process is developed for recovering Mo-99 from uranyl sulfate solutions. The minimum column volume and the maximum column length for required yield, pressure limit, and loading time are determined using a new graphical method. The method is based on dimensionless groups and intrinsic adsorption and diffusion parameters, which are estimated using a small number of experiments and simulations. The design is tested with bench-scale experiments with titania columns. The results show a high capture yield and a high stripping yield (95±5%). The design can be adapted to changes in design constraints or the variations in feed concentration, feed volume, or material properties. The graph shows clearly how the column utilization is affected by the required yield, loading time, and pressure limit. The cost effectiveness of various sorbent candidates can be evaluated based on the intrinsic parameters. This method can be used more generally for designing other capture chromatography processes.

  20. Method for (236)U Determination in Seawater Using Flow Injection Extraction Chromatography and Accelerator Mass Spectrometry.

    PubMed

    Qiao, Jixin; Hou, Xiaolin; Steier, Peter; Nielsen, Sven; Golser, Robin

    2015-07-21

    An automated analytical method implemented in a flow injection (FI) system was developed for rapid determination of (236)U in 10 L seawater samples. (238)U was used as a chemical yield tracer for the whole procedure, in which extraction chromatography (UTEVA) was exploited to purify uranium, after an effective iron hydroxide coprecipitation. Accelerator mass spectrometry (AMS) was applied for quantifying the (236)U/(238)U ratio, and inductively coupled plasma mass spectrometry (ICPMS) was used to determine the absolute concentration of (238)U; thus, the concentration of (236)U can be calculated. The key experimental parameters affecting the analytical effectiveness were investigated and optimized in order to achieve high chemical yields and simple and rapid analysis as well as low procedure background. Besides, the operational conditions for the target preparation prior to the AMS measurement were optimized, on the basis of studying the coprecipitation behavior of uranium with iron hydroxide. The analytical results indicate that the developed method is simple and robust, providing satisfactory chemical yields (80-100%) and high analysis speed (4 h/sample), which could be an appealing alternative to conventional manual methods for (236)U determination in its tracer application.

  1. Micellar electrokinetic chromatography method development for simultaneous determination of thiabendazole, carbendazim, and fuberidazole.

    PubMed

    Soliman, Laiel C; Donkor, Kingsley K

    2014-01-01

    Thiabendazole (TBz), carbendazim (CBz), and fuberidazole (FBz) are systemic benzimidazole-type fungicides used for pre- and post-harvest treatment to control various types of fungal diseases on a variety of crops. Significant levels of these fungicides could alter the composition or flavour of crops, and being possible carcinogens, they could also pose risks for humans and the environment. A mode of capillary electrophoresis called micellar electrokinetic chromatography (MEKC) was investigated for the determination of these three benzimidazole fungicides. The study involved two kinds of surfactants in which several experimental conditions were optimized, i.e., buffer concentration, pH, micelle concentration, and percent organic modifier (methanol). Using the optimum experimental conditions, the fungicides were successfully separated by MEKC. The limits of detection and quantification were in the range of 0.6-0.7 and 2.1-2.5 mg L(-1), respectively, and the calibration curves were linear over the range of 5-60 mg L(-1) for the three fungicides. The potential of the proposed MEKC method was demonstrated by analyzing water samples which were fortified with the fungicides. The proposed method enabled simultaneous determination of the three benzimidazole fungicides and method validation with spiked water samples yielded satisfactory quantitative recoveries for all the three fungicides.

  2. Method for analysis of polar volatile trace components in aqueous samples by gas chromatography.

    PubMed

    Pettersson, Johan; Roeraade, Johan

    2005-05-15

    A new method has been developed for direct analysis of volatile polar trace compounds in aqueous samples by gas chromatography. Water samples are injected onto a short packed precolumn containing anhydrous lithium chloride. A capillary column is coupled in series with the prefractionation column for final separation of the analytes. The enrichment principle of the salt precolumn is reverse to the principles employed in conventional methods such as SPE or SPME in which a sorbent or adsorbent is utilized to trap or concentrate the analytes. Such methods are not efficient for highly polar compounds. In the LiCl precolumn concept, the water matrix is strongly retained on the hygroscopic salt, whereas polar as well as nonpolar volatile organic compounds show very low retention and are eluted ahead of the water. After transfer of the analytes to the capillary column, the retained bulk water is removed by backflushing the precolumn at elevated temperature. For direct injections of 120 microL of aqueous samples, the combined time for injection and preseparation is only 3.5 min. With this procedure, direct repetitive automated analyses of highly volatile polar compounds such as methanol or tetrahydrofuran can be performed, and a limit of quantification in the low parts-per-billion region utilizing a flame ionization detector is demonstrated.

  3. AMS radiocarbon age for fossil bone by XAD-2 chromatography method

    NASA Astrophysics Data System (ADS)

    Minami, Masayo; Nakamura, Toshio

    2000-10-01

    The XAD-2 chromatography method was examined for its ability to efficiently eliminate exogenous organic matter from fossil bones and to improve the accuracy of radiocarbon ( 14C) dating and stable isotope determinations on bone proteins. The fossil bones used in the experiment were animal fossil bones collected from the Awazu submarine archaeological site, Shiga, Japan. For comparison, the gelatin-extraction method was also applied to the same samples. It was found that the gelatin-extraction method is sufficient for 14C dating on well-preserved bones, but insufficient on poorly preserved bones, containing less than 1% extractable gelatin. The XAD-2 resin is useful for the clean up of proteins especially from poorly preserved bones. The carbon stable isotope fractionation of around 1‰ by XAD-2 treatment on modern collagen standards was larger than reported previously. The isotopic variation by sequential extraction of bones probably originates from changes in the amino acid composition and seems to be less sensitive to the indication of the removal of organic contamination.

  4. Chromatography as Method for Analytical Confirmation of Paracetamol in Postmortem Material Together with Psychoactive Substances

    PubMed Central

    Biscevic-Tokic, Jasmina; Tokic, Nedim; Ibrahimpasic, Elma

    2015-01-01

    Introduction: Paracetamol (Acetaminophen) in addition to aspirin is the most commonly used analgesic and antipyretic medication by millions of patients worldwide. It is an example that paracetamol as medicine that in the world is provided without a doctor’s prescription, can lead to death. Today paracetamol became an integral part of a heroin mixture and is very popular at the street market. The main reason for this is that it can be obtained without a prescription, it is cheap, and by most people well tolerated without side effects. It is probably used for “cutting” the pure heroin, as it says in the jargon, and in that manner from small amount of pure drug is obtained greater amount, which is then sold on the street. The goal is to identify presence of paracetamol, by analytical method of gas chromatography mass spectrometer (GC-MS) in postmortem material together with psychoactive substances. Material and methods: For chemical-toxicological analysis is used biological material collected trough autopsy of 20 deceased people, suspected to have died due to psychoactive substance overdose. All received samples are stored at -20 ° C until analysis at our laboratory. From processed 47 samples that were analyzed in the period from 2014 to 2015, 19 are blood samples, urine 19, 3 samples of stomach contents, and 6 samples of bile content. Deceased were middle-aged, of which only 7 were female. The tested samples were processed according to two methods of extraction. Extraction by XAD-2 resin, and the extraction by the method of salting out with sodium tungstate. Extracts of the samples were then dissolved in chloroform and continued analysis at the analytical instrument. Identification of the paracetamol presence, in the test biological samples is demonstrated by the technique of gas chromatography with mass spectometry (hereinafter referred to as GC-MS). The technique of GC-MS is a selective, sensitive and reliable, and is therefore considered a “gold standard

  5. Rapid, economical qualitative method for separation of aflatoxins B-1, B-2 & G-1, G-2 by dry column chromatography.

    PubMed

    Megalla, S E

    1983-12-01

    A good correlation of four components of aflatoxins was accomplished by using the dry column chromatography method. The decolorization process of interfering substances, by 0.01 N KOH and defatting the extract with petroleum ether yields a clean residue for DCC separation. It is clear that the dry column chromatography is a very simple and time-saving procedure for separation of aflatoxins. DCC columns are more economical than precoated 'thick layer' preparative plates and, in DCC, no large developing tanks need to be used. Hazards associated with the use of large volumes of flammable solvents are greatly reduced.

  6. Constructing a LabVIEW-Controlled High-Performance Liquid Chromatography (HPLC) System: An Undergraduate Instrumental Methods Exercise

    ERIC Educational Resources Information Center

    Smith, Eugene T.; Hill, Marc

    2011-01-01

    In this laboratory exercise, students develop a LabVIEW-controlled high-performance liquid chromatography system utilizing a data acquisition device, two pumps, a detector, and fraction collector. The programming experience involves a variety of methods for interface communication, including serial control, analog-to-digital conversion, and…

  7. CAPILLARY GAS CHROMATOGRAPHY-ATOMIC EMISSION DETECTION METHOD FOR THE DETERMINATION OF PENTYLATED ORGANOTIN COMPOUNDS: INTERLABORATORY STUDY

    EPA Science Inventory

    A capillary gas chromatography-atomic emission detection (GC-AED) method was developed for the U. S. Environmental Protection Agency's Environmental Monitoring Systems Laboratory in Las Vegas, NV, for determination of selected organotin compounds. Here we report on an interlabora...

  8. A new ultra-pressure liquid chromatography method for the determination of biogenic amines in cheese.

    PubMed

    Mayer, H K; Fiechter, G; Fischer, E

    2010-05-07

    A fast and reliable ultra-performance liquid chromatography (UPLC) method for the determination of biogenic amines (ethanolamine, methylamine, agmatine, histamine, dimethylamine, ethylamine, octopamine, pyrrolidine, dopamine, isopropylamine, propylamine, tyramine, putrescine, butylamine, cadaverine, tryptamine, 2-phenylethylamine, 3-methylbutylamine, spermidine, spermine) in cheese was established. After pre-column derivatization with 6-aminoquinolyl-N-hydroxy-succinimidyl carbamate (AQC), 20 primary and secondary biogenic amines were separated on an Acquity UPLC column (BEH C(18), 1.7 microm; 2.1 mm x 50 mm) within 9 min. Limits of detection (mg/100g cheese) ranged from 0.04 (ethanolamine) to 1.62 (spermine), and limits of quantification were between 0.16 (ethanolamine) and 6.09 (spermine). The UPLC method was applied to the analysis of 58 cheese samples as retailed in Austria. About 13.8% of samples had a histamine content above 10mg/100g, and 22.4% had a tyramine content above 10mg/100g. Moreover, 8.6% of samples had a putrescine or cadaverine content higher than 10mg/100g. The total concentration of biogenic amines in two cheese samples was about 194 mg/100g. Thus, obligatory monitoring of biogenic amines should be considered to ensure quality of cheese in future.

  9. Analysis of Fluconazole in Human Urine Sample by High Performance Liquid Chromatography Method

    NASA Astrophysics Data System (ADS)

    Hermawan, D.; Ali, N. A. Md; Ibrahim, W. A. Wan; Sanagi, M. M.

    2013-04-01

    A method for determination of fluconazole, antifungal drug in human urine by using reversed-phased high performance liquid chromatography (RP-HPLC) with ultraviolet (UV) detector was developed. Optimization HPLC conditions were carried out by changing the flow rate and composition of mobile phase. The optimum separation conditions at a flow rate 0.85 mL/min with a composition of mobile phase containing methanol:water (70:30, v/v) with UV detection at a wavelength 254 nm was able to analyze fluconazole within 3 min. The excellent linearity was obtained in the range of concentration 1 to 10 μg/mL with r2 = 0.998. The limit of detection (LOD) and limit of quantitation (LOQ) were 0.39 μg/mL and 1.28 μg/mL, respectively. Solid phase extraction (SPE) method using octadecylsilane (C18) as a sorbent was used to clean-up and pre-concentrated of the urine sample prior to HPLC analysis. The average recoveries of fluconazole in spiked urine sample was 72.4% with RSD of 3.21% (n=3).

  10. A novel method for the determination of guanfacine in urine by gas chromatography-mass spectrometry.

    PubMed

    Haglock, Carrie; Wolf, Carl; Poklis, Alphonse

    2008-10-01

    Guanfacine (Tenex), an antihypertensive available since 1975, has recently been indicated for the treatment of attention deficit hyperactivity disorder in children (Intuniv). Because of this new usage, a gas chromatography-mass spectrometry method was developed and validated for the determination of guanfacine in urine. Guanfacine and 100 ng of protriptyline (internal standard) were extracted from 1.0 mL urine with 0.5 mL of saturated carbonate/bicarbonate buffer and 2 mL of ethyl acetate. The solvent extract was evaporated and derivatized with heptaflurobutyric anhydride in n-butyl chloride. Chromatographic separation was achieved using a DB-5 capillary column (30 m x 0.32 mm, 0.25 microm). Ions monitored for guanfacine were m/z 86.1, 272.1, and 274.1, and ions monitored for protriptyline were m/z 191.1 and 189.1. Concentrations were determined using calibrators over the range of 0.1-2.0 mg/L. The linear regression for all calibration curves had r2 values > or = 0.99. The limit of detection was 0.05 mg/L; limit of quantitation was 0.1 mg/L; and upper limit of linearity was 10.0 mg/L. Percent recovery of guanfacine at 0.1 and 2.0 mg/L was 93% and 71%, respectively. The method was found acceptable for routine quantitative analysis of guanfacine in urine.

  11. Ultra performance liquid chromatography PDA method for determination of tigecycline in human plasma.

    PubMed

    D'Avolio, Antonio; Peila, Emanuela; Simiele, Marco; Pensi, Debora; Baietto, Lorena; Cusato, Jessica; Cinnirella, Giacoma; De Rosa, Francesco; Di Perri, Giovanni

    2013-12-01

    : A simple ultra performance liquid chromatography with photodiode array method for the quantification of human plasma concentrations of tigecycline was developed and validated. Quinaxoline, used as an internal standard, was added to 500 μL of plasma before adding 1 mL of protein precipitation solution. The extracts were dried in a vacuum centrifuge system at 60°C and reconstituted with 60 μL of water and acetonitrile (95:5, vol/vol), and 5 μL was injected onto an ACQUITY UPLC H-Class system. Chromatographic separation was performed on a C18 ACQUITY UPLC HSS T3 column using a gradient of potassium phosphate buffer (pH 3.2) and acetonitrile. Detection was performed using a photodiode array detector at 350 nm. Relative error at 3 quality control concentrations ranged from -2.49% to -8.74%. Intraday and interday (percent relative standard error) precision ranged from 3.93% to 12.27% and from 9.53% to 13.32%, respectively. Limit of quantification and limit of detection were 0.024 and 0.006 μg/mL, respectively. Mean recovery was 95%. The calibration curve was linear up to 6 μg/mL. This concentration range proved to be adequate to measure tigecycline concentrations in patients treated with the drug, therefore this method would be suitable for therapeutic drug monitoring.

  12. Multi-class method for biomonitoring of hair samples using gas chromatography-mass spectrometry.

    PubMed

    Martín, Julia; Möder, Monika; Gaudl, Alexander; Alonso, Esteban; Reemtsma, Thorsten

    2015-11-01

    Currently, non-invasive biomonitoring of human exposure to organic pollutants bases upon the analysis mainly of urine and human breast milk. While mostly persistent organic pollutants are the center of interest, the aim of our study was to develop a method for the determination of different chemical classes of emerging pollutants (organophosphorus flame retardants, plastic additives such as phthalates, bisphenol A, insecticides, antimicrobials, preservatives and musk fragrances) in hair by gas chromatography-mass spectrometry. The preferred sample preparation included hydrolysis of the hair with trifluoroacetic acid in methanol followed by a liquid-liquid extraction using hexane/ethyl acetate. The validated method is characterized by recoveries higher than 77 % for most analytes, relative standard deviations below 16 % and limits of detection between 2 pg mg(-1) (HHCB) and 292 pg mg(-1) (propylparaben) using 50 mg of dry hair. After respective blank corrections, bis-(2-ethylhexyl)phthalate (DEHP) and the musk fragrance HHCB were the predominant compounds determined in all hair samples at concentrations between 32 and 59 ng mg(-1) and 0.8-13 ng mg(-1), respectively. The bactericide triclosan and the insect repellent N,N-diethyl-3-methylbenzamide (DEET) were detected in selected hair samples at 2 and 0.8 ng mg(-1), respectively.

  13. Isotope-ratio-monitoring gas chromatography-mass spectrometry: methods for isotopic calibration

    NASA Technical Reports Server (NTRS)

    Merritt, D. A.; Brand, W. A.; Hayes, J. M.

    1994-01-01

    In trial analyses of a series of n-alkanes, precise determinations of 13C contents were based on isotopic standards introduced by five different techniques and results were compared. Specifically, organic-compound standards were coinjected with the analytes and carried through chromatography and combustion with them; or CO2 was supplied from a conventional inlet and mixed with the analyte in the ion source, or CO2 was supplied from an auxiliary mixing volume and transmitted to the source without interruption of the analyte stream. Additionally, two techniques were investigated in which the analyte stream was diverted and CO2 standards were placed on a near-zero background. All methods provided accurate results. Where applicable, methods not involving interruption of the analyte stream provided the highest performance (sigma = 0.00006 at.% 13C or 0.06% for 250 pmol C as CO2 reaching the ion source), but great care was required. Techniques involving diversion of the analyte stream were immune to interference from coeluting sample components and still provided high precision (0.0001 < or = sigma < or = 0.0002 at.% or 0.1 < or = sigma < or = 0.2%).

  14. System and method for chromatography and electrophoresis using circular optical scanning

    DOEpatents

    Balch, Joseph W.; Brewer, Laurence R.; Davidson, James C.; Kimbrough, Joseph R.

    2001-01-01

    A system and method is disclosed for chromatography and electrophoresis using circular optical scanning. One or more rectangular microchannel plates or radial microchannel plates has a set of analysis channels for insertion of molecular samples. One or more scanning devices repeatedly pass over the analysis channels in one direction at a predetermined rotational velocity and with a predetermined rotational radius. The rotational radius may be dynamically varied so as to monitor the molecular sample at various positions along a analysis channel. Sample loading robots may also be used to input molecular samples into the analysis channels. Radial microchannel plates are built from a substrate whose analysis channels are disposed at a non-parallel angle with respect to each other. A first step in the method accesses either a rectangular or radial microchannel plate, having a set of analysis channels, and second step passes a scanning device repeatedly in one direction over the analysis channels. As a third step, the scanning device is passed over the analysis channels at dynamically varying distances from a centerpoint of the scanning device. As a fourth step, molecular samples are loaded into the analysis channels with a robot.

  15. Quality Evaluation of Potentilla fruticosa L. by High Performance Liquid Chromatography Fingerprinting Associated with Chemometric Methods

    PubMed Central

    Liu, Wei; Wang, Dongmei; Liu, Jianjun; Li, Dengwu; Yin, Dongxue

    2016-01-01

    The present study was performed to assess the quality of Potentilla fruticosa L. sampled from distinct regions of China using high performance liquid chromatography (HPLC) fingerprinting coupled with a suite of chemometric methods. For this quantitative analysis, the main active phytochemical compositions and the antioxidant activity in P. fruticosa were also investigated. Considering the high percentages and antioxidant activities of phytochemicals, P. fruticosa samples from Kangding, Sichuan were selected as the most valuable raw materials. Similarity analysis (SA) of HPLC fingerprints, hierarchical cluster analysis (HCA), principle component analysis (PCA), and discriminant analysis (DA) were further employed to provide accurate classification and quality estimates of P. fruticosa. Two principal components (PCs) were collected by PCA. PC1 separated samples from Kangding, Sichuan, capturing 57.64% of the variance, whereas PC2 contributed to further separation, capturing 18.97% of the variance. Two kinds of discriminant functions with a 100% discrimination ratio were constructed. The results strongly supported the conclusion that the eight samples from different regions were clustered into three major groups, corresponding with their morphological classification, for which HPLC analysis confirmed the considerable variation in phytochemical compositions and that P. fruticosa samples from Kangding, Sichuan were of high quality. The results of SA, HCA, PCA, and DA were in agreement and performed well for the quality assessment of P. fruticosa. Consequently, HPLC fingerprinting coupled with chemometric techniques provides a highly flexible and reliable method for the quality evaluation of traditional Chinese medicines. PMID:26890416

  16. Improved liquid chromatography-tandem mass spectrometry method for the analysis of eptifibatide in human plasma.

    PubMed

    Zhou, Zhi-Ling; Yu, Xi-Yong; Yang, Min; Mai, Li-Ping; Lin, Qiu-Xiong; Deng, Chun-Yu; Shan, Zhi-Xin; Kuang, Su-Juan; Zhu, Ping; Huang, Xiao-Zhong; Li, Xiao-Hong; Chen, Tie-Feng; Lin, Shu-Guang

    2010-08-01

    A rapid, selective and highly sensitive high performance liquid chromatography-tandem mass spectrometry method (LC-MS/MS) was developed and validated for the determination and pharmacokinetic investigation of eptifibatide in human plasma. Eptifibatide and the internal standard (IS), EPM-05, were extracted from plasma samples using solid phase extraction. Chromatographic separation was performed on a C(18) column at a flow rate of 0.5 mL/min. Detection of eptifibatide and the IS was achieved by tandem mass spectrometry with an electrospray ionization (ESI) interface in positive ion mode. Traditional multiple reaction monitoring (MRM) using the transition of m/z 832.6-->m/z 646.4 and m/z 931.6-->m/z 159.4 was performed to quantify eptifibatide and the IS, respectively. The calibration curves were linear over the range of 1-1000 ng/mL with the lower limit of quantitation validated at 1 ng/mL. The intra- and inter-day precisions were within 13.3%, while the accuracy was within +/-7.6% of nominal values. The validated LC-MS/MS method was successfully applied for the evaluation of pharmacokinetic parameters of eptifibatide after intravenous (i.v.) administration of a 45 microg/kg bolus of eptifibatide to 8 healthy volunteers.

  17. A validated new method for nevirapine quantitation in human plasma via high-performance liquid chromatography.

    PubMed

    Silverthorn, Courtney F; Parsons, Teresa L

    2006-01-01

    A fully validated and clinically relevant assay was developed for the assessment of nevirapine concentrations in neonate blood plasma samples. Solid-phase extraction with an acid-base wash series was used to prepare subject samples for analysis. Samples were separated by high performance liquid chromatography and detected at 280 nm on a C8 reverse-phase column in an isocratic mobile phase. The retention times of nevirapine and its internal standard were 5.0 and 6.9 min, respectively. The method was validated by assessment of accuracy and precision (statistical values <15%), specificity, and stability. The assay was linear in the range 25-10,000 ng/mL (r2 > 0.996) and the average recovery was 93% (n = 18). The lower limit of quantification (relative standard deviation <20%) was determined to be 25 ng/mL for 50 microL of plasma, allowing detection of as little as 1.25 ng of nevirapine in a sample. This value represents an increase in sensitivity of up to 30-fold over previously published methods.

  18. A micellar liquid chromatography method for the quantification of abacavir, lamivudine and raltegravir in plasma.

    PubMed

    Peris-Vicente, Juan; Villareal-Traver, Mónica; Casas-Breva, Inmaculada; Carda-Broch, Samuel; Esteve-Romero, Josep

    2014-09-01

    An analytical methodology based on micellar liquid chromatography has been developed to quantify abacavir, lamivudine and raltegravir in plasma. These three antiretroviral drugs are prescribed as a set in highly active antiretroviral therapy to acquired immunodeficiency syndrome patients. The experimental procedure consists in the dilution of the sample in micellar media, followed by filtration and, without cleanup step. The analytes were resolved in less than 30min using a mobile phase of 0.05M sodium dodecyl sulphate at pH 7, running at 1mLmin(-1) under isocratic mode at room temperature through a C18 column (125×4.6mm, 5μm particle size). The UV detection wavelength was set at 260nm. The method was successfully validated following the requirements of ICH guidelines in terms of: linear range (0.25-2.5μgmL(-1)), linearity (r(2)>0.990), intra- and interday precision (<6.8%) and accuracy (92.3-104.2%) and robustness (<7.1%). To the extent of our knowledge, this is the first published method to quantify these three drugs in plasma. Several blood samples from AIDS patients taking this HAART set provided by a local hospital were analyzed with satisfactory results.

  19. Quality Evaluation of Potentilla fruticosa L. by High Performance Liquid Chromatography Fingerprinting Associated with Chemometric Methods.

    PubMed

    Liu, Wei; Wang, Dongmei; Liu, Jianjun; Li, Dengwu; Yin, Dongxue

    2016-01-01

    The present study was performed to assess the quality of Potentilla fruticosa L. sampled from distinct regions of China using high performance liquid chromatography (HPLC) fingerprinting coupled with a suite of chemometric methods. For this quantitative analysis, the main active phytochemical compositions and the antioxidant activity in P. fruticosa were also investigated. Considering the high percentages and antioxidant activities of phytochemicals, P. fruticosa samples from Kangding, Sichuan were selected as the most valuable raw materials. Similarity analysis (SA) of HPLC fingerprints, hierarchical cluster analysis (HCA), principle component analysis (PCA), and discriminant analysis (DA) were further employed to provide accurate classification and quality estimates of P. fruticosa. Two principal components (PCs) were collected by PCA. PC1 separated samples from Kangding, Sichuan, capturing 57.64% of the variance, whereas PC2 contributed to further separation, capturing 18.97% of the variance. Two kinds of discriminant functions with a 100% discrimination ratio were constructed. The results strongly supported the conclusion that the eight samples from different regions were clustered into three major groups, corresponding with their morphological classification, for which HPLC analysis confirmed the considerable variation in phytochemical compositions and that P. fruticosa samples from Kangding, Sichuan were of high quality. The results of SA, HCA, PCA, and DA were in agreement and performed well for the quality assessment of P. fruticosa. Consequently, HPLC fingerprinting coupled with chemometric techniques provides a highly flexible and reliable method for the quality evaluation of traditional Chinese medicines.

  20. Improved method for rapid detection of phthalates in bottled water by gas chromatography-mass spectrometry.

    PubMed

    Otero, Paz; Saha, Sushanta Kumar; Moane, Siobhan; Barron, John; Clancy, Gerard; Murray, Patrick

    2015-08-01

    An improved gas chromatography-mass spectrometry (GC-MS) method for simple, rapid and precise quantification of phthalates in drinking water is presented. This method was validated for bis (2-n-butoxyethyl) phthalate (DBEP), bis (2-n-ethylhexyl) phthalate (DEHP), butyl benzyl phthalate (BBP), di-butyl phthalate (DBP), diethyl phthalate (DEP), dihexyl phthalate (DHP), dimethyl phthalate (DMP), di-n-octyl phthalate (DNOP) and dinonyl phthalate (DINP). Linearity of 0.9984>r(2)>0.9975 in the range of 0.075-4.8μg/mL for the selected phthalates was obtained. Accuracy values were in the range of 93-114% and RSD% for the analysis of 1.2μg/mL of each phthalate was below 2.3% (n=9). This new method design has significantly improved the detection in terms of rapidity, specificity, repeatability and accuracy compared to available methods. The procedure has been applied to the analyses of three different brands of commercially available bottled mineral water and the corresponding plastic bottles. Phthalates were extracted with dichloromethane and re-constituted in cyclohexane prior to GC-MS analysis. When the validated GC-MS method was applied to the quantification of the selected phthalates in the samples, only DBP (up to 0.0675±0.0018μg/mL) and DEHP (up to 1.6848±0.1631μg/mL) were found. Furthermore, we provide specific data about the concentration of DBP and DEHP in bottled water attributable to migration of phthalates from respective plastic bottles.

  1. Ultra-high-performance liquid chromatography profiling method for chemical screening of proanthocyanidins in Czech hops.

    PubMed

    Olšovská, J; Kameník, Z; Čejka, P; Jurková, M; Mikyška, A

    2013-11-15

    Hops represent an important natural source of bioactive polyphenols, particularly proanthocyanidins, which can contribute to prevention of several civilization diseases, owing to their antioxidant and radical scavenging activity. We have developed a high-throughput ultra-high-performance liquid chromatography time-of-flight mass spectrometry profiling method, which can be used for monitoring of bioactive proanthocyanidins in hops. The method was applied for analysis of hops of four Czech varieties (Saaz, Sladek, Preminat and Agnus) from the 2011 crop (9 localities, 11 samples) and the 2012 crop (24 localities, 40 samples). Hop samples were extracted by acetone and the analytes were separated on the Acquity UPLC BEH Shield RP18 column. Partial validation of the method revealed a satisfactory intra-day repeatability of the method for retention times (relative standard deviation within 1.39%) as well as areas under the peaks (within 9.89%). Experimental data were evaluated using principal component analysis and cluster analysis. Significant amounts of di-, tri- and tetramer proanthocyanidins consisting of (epi)catechin and (epi)gallocatechin were found in the hop samples. The dependence of the proantocyanidin composition on both the variety and the growing locality was observed. Specifically, the traditional Saaz variety contained more frequently oligomers formed by (epi)catechin units only, whereas the varieties Premiant and Agnus produced oligomers consisting of (epi)catechin as well as (epi)gallocatechin units. The relative abundance of proanthocyanidins in studied hop varieties from the two crops, 2011 and 2012, did correspond to each other. In the further perspective, the method may also be used for prediction of qualitative marks or authenticity verification of hops.

  2. Characterization and classification of pseudo-stationary phases in micellar electrokinetic chromatography using chemometric methods.

    PubMed

    Fu, Cexiong; Khaledi, Morteza G

    2014-03-04

    Two types of chemometric methods, principal component analysis (PCA) and cluster analysis, are employed to characterize and classify a total of 70 pseudostationary phases (54 distinct systems and 16 decoy systems) in micellar electrokinetic chromatography (MEKC). PCA excels at removing redundant information for micellar phase characterization and retaining principal determinants for phase classification. While PCA is useful in the characterization of micelle selectivities, it is ineffective in defining the grouping of micellar phases. Hierarchical clustering yields a complete dendrogram of cluster structures but provides only limited cluster characterizations. The combination of these two chemometric methods leads to a comprehensive interpretation of the micellar phase classification. Moreover, the k-means analysis can further discern subtle differences among those closely located micellar phases. All three chemometric methods result in similar classifications with respect to the similarities and differences of the 70 micelle systems investigated. These systems are categorized into 3 major clusters: fluoro-surfactants represent cluster I, identified as strong hydrogen bond donors and dipolar but weak hydrogen bond acceptors. Cluster II includes sulfonated acrylamide/acrylate copolymers and surfactants with trimethylammonium head groups, characterized by strong hydrophobicity (v) and weak hydrogen bond acidity (b). The last cluster consists of two subclusters: clusters III and IV. Cluster III includes siloxane-based polymeric micelles, exhibiting weak hydrophobicity and medium hydrogen bond acidity and basicity (a), and the cluster IV micellar systems are characterized by their strong hydrophobicity and medium hydrogen bond acidity and basicity but rather weak dipolarity. Cluster III differs from cluster IV by its slightly weaker hydrophobicity and hydrogen bond donating capability. The classification by chemometric methods is in good agreement with the

  3. Rapid liquid chromatography for paralytic shellfish toxin analysis using superficially porous chromatography with AOAC Official Method 2005.06.

    PubMed

    Hatfield, Robert G; Turner, Andrew D

    2012-01-01

    The bioaccumulation of paralytic shellfish toxins in mussels, oysters, cockles, hard clams, razors, and king scallops is monitored in England, Scotland, and Wales by AOAC Official Method 2005.06 LC-with fluorescence detection (FLD). One of the commonly perceived disadvantages of using this method is the long turnaround time and low throughput in a busy laboratory environment. The chromatographic analysis of each sample typically utilizes a 15 min cycle time to achieve toxin oxidation product separation and column equilibration prior to subsequent analysis. A standard RP C18 analytical column, used successfully in recent years, achieves good separation with a long column lifetime. The analysis of a 40 sample qualitative screening batch takes approximately 18 h, including blanks, standards, and other QC samples. The availability of superficially porous column technology has offered the potential to reduce analysis time while retaining column performance on existing hardware. In this study, AOAC Official Method 2005.06 with LC-FLD was transferred to two different commercially available superficially porous columns, and the method performance characteristics were evaluated. Both columns separated all toxins adequately with cycle times less than half that of the existing method. Linearity for each toxin was acceptable up to two times the European maximum permitted limit of 800 microg di-HCl saxitoxin equivalent/kg flesh. LOD and LOQ values were substantially improved for the majority of toxins, with gonyautoxin 1&4 and neosaxitoxin showing up to a two- and fourfold improvement, respectively, depending on the column used. Quantification results obtained from parallel analysis of contaminated samples were acceptable on both columns. Comparative screen results gave a slight increase in the occurrence of contaminated samples, which was attributed to the improved detection limit for most toxins. Issues with rapidly increasing back pressure, however, were identified with both

  4. New methods for determination of cinnarizine in mixture with piracetam by spectrodensitometry, spectrophotometry, and liquid chromatography.

    PubMed

    Metwally, Fadia H; Elzeany, B A; Darwish, H W

    2005-01-01

    Four new methods were developed and validated for the determination of cinnarizine HCl in its binary mixture with piracetam in pure and pharmaceutical preparations. The first one was a densitometric analysis that provides a simple and rapid method for the separation and quantification of cinnarizine HCI. The method depends on the quantitative densitometric evaluation of thin-layer chromatograms of cinnarizine HCI at 252 nm over concentration range of 1-6 microg/spot, with a mean accuracy of 100.05 +/- 0.91%. The second method was determination of the drug using a colorimetric method that utilizes the reaction of 3-methyl-benzothiazolin-2-one in the presence of FeCl3 as an oxidant. The green color of the resulting product was measured at 630 nm over concentration range 10-40 microg/mL, with a mean accuracy of 100.10 +/- 1.13%. The third method was a direct spectrophotometric determination of cinnarizine HCI at 252 nm over the concentration range 7-20 microg/mL, while piracetam was determined by derivative ratio spectrophotometry at 221.6 nm over concentration range 5-30 microg/mL, with a mean accuracy of 100.14 +/- 0.79 and 100.26 +/- 1.24% for cinnarizine HCI and piracetam, respectively. The last method was a liquid chromatography analysis of both cinnarizine HCI and piracetam, depending on quantitative evaluation of chromatograms of cinnarizine HCI and piracetam at 252 and 212 nm, respectively, over the concentration range 10-200 microg/mL for cinnarizine HCI and 20-500 microg/mL for piracetam, with a mean accuracy of 100.03 +/- 0.89 and 100.40 +/- 0.94% for cinnarizine HCI and piracetam, respectively. The proposed procedures were checked using laboratory-prepared mixtures and successfully applied for the analysis of their pharmaceutical preparations. The validity of the proposed procedures was further assessed by applying the standard addition technique. Recoveries were quantitative, and the results obtained agreed with those obtained by other reported methods.

  5. Method transfer from high-pressure liquid chromatography to ultra-high-pressure liquid chromatography. I. A thermodynamic perspective.

    PubMed

    Åsberg, Dennis; Leśko, Marek; Samuelsson, Jörgen; Kaczmarski, Krzysztof; Fornstedt, Torgny

    2014-10-03

    This is the first investigation in a series that aims to enhance the scientific knowledge needed for reliable analytical method transfer between HPLC and UHPLC using the quality by design (QbD) framework. Here, we investigated the differences and similarities from a thermodynamic point of view between RP-LC separations conducted with 3.5μm (HPLC) and 1.7μm (UHPLC) C18 particles. Three different model solutes and one pharmaceutical compound were used: the uncharged cycloheptanone, the cationic benzyltriethylammonium chloride, the anionic sodium 2-naphatlene sulfonate and the pharmaceutical compound omeprazole, which was anionic at the studied pH. Adsorption data were determined for the four solutes at varying fractions of organic modifier and in gradient elution in both the HPLC and UHPLC system, respectively. From the adsorption data, the adsorption energy distribution of each compound was calculated and the adsorption isotherm model was estimated. We found that the adsorption energy distribution was similar, with only minor differences in degree of homogeneity, for HPLC and UHPLC stationary phases. The adsorption isotherm model did not change between HPLC and UHPLC, but the parameter values changed considerably especially for the ionic compounds. The dependence of the organic modifier followed the same trend in HPLC as in UHPLC. These results indicates that the adsorption mechanism of a solute is the same on HPLC and UHPLC stationary phases which simplifies design of a single analytical method applicable to both HPLC and UHPLC conditions within the QbD framework.

  6. Liquid chromatography/tandem mass spectrometry method for quantitation of cremophor el and its applications.

    PubMed

    Vijaya Bhaskar, V; Middha, Anil

    2013-01-01

    A rapid sensitive and selective MRM based method for the determination of Cremophor EL (CrEL) in rat plasma was developed using liquid chromatography/tandem mass spectrometry (LC-MS/MS). CrEL and polypropylene glycol (internal standard) were extracted from rat plasma with acetonitrile and analysed on C18 column (XBridge, 50 × 4.6 mm, 3.5  μ m). The most abundant molecular ions corresponding to PEG oligomers at m/z 828, 872, 916 and 960 with daughter ion at m/z 89 were selected for multiple reaction monitoring (MRM) in electrospray mode of ionisation. Plasma concentrations of CrEL were quantified after administration through oral and intravenous routes in male sprague dawley rats at a dose of 0.26 g/kg. The standard curve was linear (0.9972) over the concentration range of 1.00 to 200  μ g/mL. The lower limit of quantitation for CrEL was 1.00  μ g/mL using 50  μ L plasma. The coefficient of variation and relative error for inter and intra assay at three QC levels were 0.69 to 9.21 and -7.60 to 4.74 respectively. A novel proposal was conveyed to the scientific community, where formulation excipient can be analysed as qualifier in the analysis of NCEs to address the spiky plasma concentration profiles.

  7. A gas chromatography-mass spectrometry method to monitor detergents removal from a membrane protein sample.

    PubMed

    Shi, Chaowei; Han, Fang; Xiong, Ying; Tian, Changlin

    2009-12-01

    In membrane protein biochemical and structural studies, detergents are used to mimic membrane environment and maintain functional, stable conformation of membrane proteins in the absence of lipid bilayers. However, detergent concentration, esp. molar ratio of membrane protein to detergent is usually unknown. Here, a gas chromatography-mass spectrometry selected ion monitoring (GC-MS-SIM) method was developed to quantify four detergents which are frequently used in membrane protein structural studies. To remove excessive detergents, a filtered centrifugation using Centricon tubes was applied. A membrane protein Ig-Beta fragment in four different detergent micelles was exemplified. Detergent concentrations in the upper and lower fraction of the Centricon tube were measured after each round of centrifugation. The results were very consistent to basic properties of detergent micelles in aqueous solvents. Therefore, coupling of GC-MS-SIM and detergent removal by Centricon tubes, detergents concentration, esp. molar ratio of membrane protein to detergent could be controlled, which will expedite membrane protein structural and biochemical studies.

  8. Simple micellar electrokinetic chromatography method for the determination of hydrogen sulfide in hen tissues.

    PubMed

    Kubalczyk, Paweł; Borowczyk, Kamila; Chwatko, Grażyna; Głowacki, Rafał

    2015-04-01

    A new method for the determination of hydrogen sulfide in hen tissues has been developed and validated. For estimation of hydrogen sulfide content, a sample (0.1 g) of hen tissue was treated according to the procedure consisted of some essential steps: simultaneous homogenization of a tissue and derivatization of hydrogen sulfide to its S-quinolinium derivative with 2-chloro-1-methylquinolinium tetrafluoroborate, separation of so-formed derivative by micellar electrokinetic chromatography with sweeping, and detection and quantitation with the use of UV detector set to measure analytical signals at 375 nm. Effective electrophoretic separation was achieved using fused silica capillary (effective length 41.5 cm, 75 μm id) and 0.05 mol/L, pH 8 phosphate buffer with the addition of 0.04 mol/L SDS and 26% ACN. The lower limit of quantification was 0.12 μmol hydrogen sulfide in 1 g of tissue. The calibration curve prepared in tissue homogenate for hydrogen sulfide showed linearity in the range from 0.15 to 2.0 μmol/g, with the coefficient of correlation 0.9978. The relative standard deviation of the points of the calibration curve varied from 8.3 to 3.2% RSD.

  9. Determination of eptifibatide concentration in human plasma utilizing the liquid chromatography-tandem mass spectrometry method.

    PubMed

    Liu, Jia; Duan, Xiaotao; Chen, Xiaoyan; Zhong, Dafang

    2009-02-15

    A sensitive and selective liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed to determine the concentration of eptifibatide in human plasma. Following protein precipitation, the analyte was separated on a reversed-phase C(18) column. Acetonitrile:5mM ammonium acetate:acetic acid (30:70:0.1, v/v/v) was used at a flow-rate of 0.5mL/min with the isocratic mobile phase. An API 4000 tandem mass spectrometer equipped with a Turbo IonSpray ionization source was used as the detector and was operated in the positive ion mode. "Truncated" multiple reaction monitoring using the transition of m/z 832.6-->m/z 832.6 and m/z 931.3-->m/z 931.3 was performed to quantify eptifibatide and the internal standard (EPM-05), respectively. The method had a lower limit of quantification of 4.61ng/mL for eptifibatide. The calibration curve was demonstrated to be linear over the concentration range of 4.61-2770ng/mL. The intra- and inter-day precisions were less than 10.5% for each QC level, and the inter-day relative errors were 2.0%, 5.6%, and 2.8% for 9.22, 184, and 2490ng/mL, respectively. The validated method was successfully applied to the quantification of eptifibatide concentration in human plasma after intravenous (i.v.) administration of a 270-microg/kg bolus of eptifibatide and i.v. administration of eptifibatide at a constant rate of infusion of 2microg/(kgmin) for 18h in order to evaluate the pharmacokinetics.

  10. Evaluation of injection methods for fast, high peak capacity separations with low thermal mass gas chromatography.

    PubMed

    Fitz, Brian D; Mannion, Brandyn C; To, Khang; Hoac, Trinh; Synovec, Robert E

    2015-05-01

    Low thermal mass gas chromatography (LTM-GC) was evaluated for rapid, high peak capacity separations with three injection methods: liquid, headspace solid phase micro-extraction (HS-SPME), and direct vapor. An Agilent LTM equipped with a short microbore capillary column was operated at a column heating rate of 250 °C/min to produce a 60s separation. Two sets of experiments were conducted in parallel to characterize the instrumental platform. First, the three injection methods were performed in conjunction with in-house built high-speed cryo-focusing injection (HSCFI) to cryogenically trap and re-inject the analytes onto the LTM-GC column in a narrower band. Next, the three injection methods were performed natively with LTM-GC. Using HSCFI, the peak capacity of a separation of 50 nl of a 73 component liquid test mixture was 270, which was 23% higher than without HSCFI. Similar peak capacity gains were obtained when using the HSCFI with HS-SPME (25%), and even greater with vapor injection (56%). For the 100 μl vapor sample injected without HSCFI, the preconcentration factor, defined as the ratio of the maximum concentration of the detected analyte peak relative to the analyte concentration injected with the syringe, was determined to be 11 for the earliest eluting peak (most volatile analyte). In contrast, the preconcentration factor for the earliest eluting peak using HSCFI was 103. Therefore, LTM-GC is demonstrated to natively provide in situ analyte trapping, although not to as great an extent as with HSCFI. We also report the use of LTM-GC applied with time-of-flight mass spectrometry (TOFMS) detection for rapid, high peak capacity separations from SPME sampled banana peel headspace.

  11. Determination of Alternaria mycotoxins in wine and juice using ionic liquid modified countercurrent chromatography as a pretreatment method followed by high-performance liquid chromatography.

    PubMed

    Fan, Chen; Cao, Xueli; Liu, Man; Wang, Wei

    2016-03-04

    Alternariol (AOH), alternariol monomethyl ether (AME), and tenuazonic acid (TeA) are some of the main Alternaria mycotoxins that can be found as contaminants in food materials. The objective of this study was to develop a pretreatment method with countercurrent chromatography (CCC) for enrichment and cleanup of trace Alternaria mycotoxins in food samples prior to high-performance liquid chromatography (HPLC) analysis. An Analytical CCC instrument with a column volume 22.5mL was used, and a two-phase solvent system composed of ethyl acetate and water modified with 6% [HOOMIM][Cl] in mass to volume ratio was selected. Under the optimized CCC operation conditions, trace amounts of AOH, AME, and TeA in large volume of liquid sample were efficiently extracted and enriched in the stationary phase, and then eluted out just by reversing the stationary phase as mobile phase in the opposite flowing direction tail-to-head. The enrichment and elution strategies are unique and can be fulfilled online with high enrichment factors (87-114) and high recoveries (81.14-110.94%). The method has been successively applied to the determination of Alternaria mycotoxins in real apple juice and wine samples with the limits of detection (LOD) in the range of 0.03-0.14μgL(-1). Totally 12 wine samples and 15 apple juice samples from the local market were analyzed. The detection rate of AOH and AME in both kinds of the samples were more than 50%, while TeA was found in relatively high level of 1.75-49.61μgL(-1) in some of the apple juice samples. The proposed method is simple, rapid, and sensitive and could also be used for the analysis and monitoring of Alternaria mycotoxin in other food samples.

  12. Ultrafast gas chromatography method with direct injection for the quantitative determination of benzene, toluene, ethylbenzene, and xylenes in commercial gasoline.

    PubMed

    Miranda, Nahieh Toscano; Sequinel, Rodrigo; Hatanaka, Rafael Rodrigues; de Oliveira, José Eduardo; Flumignan, Danilo Luiz

    2017-04-01

    Benzene, toluene, ethylbenzene, and xylenes are some of the most hazardous constituents found in commercial gasoline samples; therefore, these components must be monitored to avoid toxicological problems. We propose a new routine method of ultrafast gas chromatography coupled to flame ionization detection for the direct determination of benzene, toluene, ethylbenzene, and xylenes in commercial gasoline. This method is based on external standard calibration to quantify each compound, including the validation step of the study of linearity, detection and quantification limits, precision, and accuracy. The time of analysis was less than 3.2 min, with quantitative statements regarding the separation and quantification of all compounds in commercial gasoline samples. Ultrafast gas chromatography is a promising alternative method to official analytical techniques. Government laboratories could consider using this method for quality control.

  13. [Measurement of free urinary cortisol and cortisone using liquid chromatography associated with tandem mass spectrometry method].

    PubMed

    Vieira, José Gilberto H; Nakamura, Odete H; Carvalho, Valdemir M

    2005-04-01

    Free urinary cortisol (UFF) measurement is one of the most useful screening tests for Cushing's syndrome. Immunoassays employed today by most clinical laboratories present limitations, specially concerning specificity. These limitations restrain a widespread application of the method, as well as the comparison of results obtained by the use of different methods. We present the development and characterization of a UFF and cortisone method based on liquid chromatography and tandem mass spectrometry (LC-MS/MS). A 200 microL aliquot from a 24 h urine sample is mixed with a solution containing a known quantity of deuterated cortisol and on-line extracted in solid phase (C18). The eluate is transferred to a second C18 column (Phenomenex Luna, 3 micro, 50 x 2 mm) and the isocratic mode elution profile is directly applied to a tandem mass spectrometer model Quattro Micro operating in positive mode atmospheric pressure chemical ionization (APCI). All process is automated and the quantification is performed by isotopic dilution, based on the analyte and the deuterated internal standard peak area ratios. The specificity study showed that all the steroids tested presented cross reactivity of <1% for cortisol and cortisone. Functional sensitivity is <1 microg/L for both steroids, and the interassay CV <8%. Recovery and linearity studies were satisfactory and comparison of results obtained using a RIA for UFF and the present method in 98 routine samples showed a correlation of r= 0.838, with the results obtained with LC-MS/MS significantly lower (medians of 22.0 vs. 49.4 microg/24 h for RIA) (P<0.0001). Reference values for cortisol were defined as values between 11 and 43 microg/24 h, compatible to those recently described for similar methods. The concomitant measurement of UF cortisone allows the study of the activity of the enzyme 11beta-HSD2 and the diagnosis of the apparent mineralocorticoid excess syndrome. The method represents the first steroid assay of a new generation

  14. Ultra-high-pressure liquid chromatography-tandem mass spectrometry method for the determination of alkylphenols in soil.

    PubMed

    Wang, Jing; Pan, Hefang; Liu, Zhengzheng; Ge, Fei

    2009-03-20

    A novel method has been developed for the determination of alkylphenols in soil by ultra-high-pressure liquid chromatography employing small particle sizes, combined with tandem mass spectrometry. Soil samples were extracted with pressurized liquid extraction (PLE) and then cleaned with solid-phase extraction (SPE). The extracts were separated on C18 column (1.7 microm, 50 mm x 2.1mm) with a gradient elution and a mobile phase consisting of water and acetonitrile, and then detected by an electrospray ionization tandem mass spectrometry in negative ion mode with multiple reaction monitoring (MRM). Compared with traditional liquid chromatography, it took ultra-high-pressure liquid chromatography much less time to analyze alkylphenols. Additionally, the ultra-high-pressure liquid chromatography/tandem mass spectrometry method produces satisfactory reliability, sensitivity, and accuracy. The average recoveries of the three target analytes were 74.0-103.4%, with the RSD<15%. The calibration curves for alkylphenols were linear within the range of 0.01-0.4 microg/ml, with the correlation coefficients greater than 0.99. When 10 g soil sample was used for analysis, the limits of quantification (LOQs) of the three alkylphenols were all 1.0 microg/kg.

  15. Influence of drying method on the surface energy of cellulose nanofibrils determined by inverse gas chromatography.

    PubMed

    Peng, Yucheng; Gardner, Douglas J; Han, Yousoo; Cai, Zhiyong; Tshabalala, Mandla A

    2013-09-01

    Research and development of the renewable nanomaterial cellulose nanofibrils (CNFs) has received considerable attention. The effect of drying on the surface energy of CNFs was investigated. Samples of nanofibrillated cellulose (NFC) and cellulose nanocrystals (CNC) were each subjected to four separate drying methods: air-drying, freeze-drying, spray-drying, and supercritical-drying. The surface morphology of the dried CNFs was examined using a scanning electron microscope. The surface energy of the dried CNFs was determined using inverse gas chromatography at infinite dilution and column temperatures: 30, 40, 50, 55, and 60 °C. Surface energy measurements of supercritical-dried NFCs were performed also at column temperatures: 70, 75, and 80 °C. Different drying methods produced CNFs with different morphologies which in turn significantly influenced their surface energy. Supercritical-drying resulted in NFCs having a dispersion component of surface energy of 98.3±5.8 mJ/m(2) at 30 °C. The dispersion component of surface energy of freeze-dried NFCs (44.3±0.4 mJ/m(2) at 30 °C) and CNCs (46.5±0.9 mJ/m(2) at 30 °C) were the lowest among all the CNFs. The pre-freezing treatment during the freeze-drying process is hypothesized to have a major impact on the dispersion component of surface energy of the CNFs. The acid and base parameters of all the dried CNFs were amphoteric (acidic and basic) although predominantly basic in nature.

  16. The Generally Useful Estimate of Solvent Systems (GUESS) method enables the rapid purification of methylpyridoxine regioisomers by countercurrent chromatography.

    PubMed

    Liu, Yang; Friesen, J Brent; Klein, Larry L; McAlpine, James B; Lankin, David C; Pauli, Guido F; Chen, Shao-Nong

    2015-12-24

    The TLC-based Generally Useful Estimate of Solvent Systems (GUESS) method was employed for countercurrent chromatography solvent system selection, in order to separate the three synthetic isomers: 3-O-methylpyridoxine, 4'-O-methylpyridoxine (ginkgotoxin), and 5'-O-methylpyridoxine. The Rf values of the three isomers indicated that ChMWat+2 (chloroform-methanol-water 10:5:5, v/v/v) was appropriate for the countercurrent separation. The isomer separation was highly selective and demonstrated that the TLC-based GUESS method can accelerate solvent system selection for countercurrent separation. Accordingly, the study re-emphasizes the practicality of TLC as a tool to facilitate the rapid development of new countercurrent and centrifugal partition chromatography methods for this solvent system. Purity and structure characterization of all samples was performed by quantitative (1)H NMR.

  17. Method developments approaches in supercritical fluid chromatography applied to the analysis of cosmetics.

    PubMed

    Lesellier, E; Mith, D; Dubrulle, I

    2015-12-04

    Analyses of complex samples of cosmetics, such as creams or lotions, are generally achieved by HPLC. These analyses are often multistep gradients, due to the presence of compounds with a large range of polarity. For instance, the bioactive compounds may be polar, while the matrix contains lipid components that are rather non-polar, thus cosmetic formulations are usually oil-water emulsions. Supercritical fluid chromatography (SFC) uses mobile phases composed of carbon dioxide and organic co-solvents, allowing for good solubility of both the active compounds and the matrix excipients. Moreover, the classical and well-known properties of these mobile phases yield fast analyses and ensure rapid method development. However, due to the large number of stationary phases available for SFC and to the varied additional parameters acting both on retention and separation factors (co-solvent nature and percentage, temperature, backpressure, flow rate, column dimensions and particle size), a simplified approach can be followed to ensure a fast method development. First, suited stationary phases should be carefully selected for an initial screening, and then the other operating parameters can be limited to the co-solvent nature and percentage, maintaining the oven temperature and back-pressure constant. To describe simple method development guidelines in SFC, three sample applications are discussed in this paper: UV-filters (sunscreens) in sunscreen cream, glyceryl caprylate in eye liner and caffeine in eye serum. Firstly, five stationary phases (ACQUITY UPC(2)) are screened with isocratic elution conditions (10% methanol in carbon dioxide). Complementary of the stationary phases is assessed based on our spider diagram classification which compares a large number of stationary phases based on five molecular interactions. Secondly, the one or two best stationary phases are retained for further optimization of mobile phase composition, with isocratic elution conditions or, when

  18. Ion chromatography to detect salts in stone structures and to assess salt removal methods

    NASA Astrophysics Data System (ADS)

    Alvarez de Buergo, M.; Lopez-Arce, P.; Fort, R.

    2012-04-01

    thorough previous inspection, we can select the most representative points by a drilling net - as minimum as possible- and make some profiles of the inner salt content of a structure. Moreover, this procedure is not only reliable for determining the nature and extent of salts damage, but to assess the efficacy of salts removal methods in cultural heritage. Here we present two case studies from relevant buildings of the Spanish cultural heritage in which this procedure was performed with successful and useful results, in both terms of understanding what types of salts were decaying the stones structures, and also whether the salts removal methods that were planned in the restoration project were efficient or not. It should be remarked that even ion chromatography is not a non destructive technique (can be considered as a minimally destructive one due to the few quantity it is needed for the analysis), the information it can provide is so useful that should not be ruled out from the beginning, depending of each specific case.

  19. EPA Method 525.3 - Determination of Semivolatile Organic Chemicals in Drinking Water by Solid Phase Extraction and Capillary Column Gas Chromatography/Mass Spectrometry (GC/MS)

    EPA Science Inventory

    Method 525.3 is an analytical method that uses solid phase extraction (SPE) and gas chromatography/mass spectrometry (GC/MS) for the identification and quantitation of 125 selected semi-volatile organic chemicals in drinking water.

  20. Ion Chromatography.

    ERIC Educational Resources Information Center

    Mulik, James D.; Sawicki, Eugene

    1979-01-01

    Accurate for the analysis of ions in solution, this form of analysis enables the analyst to directly assay many compounds that previously were difficult or impossible to analyze. The method is a combination of the methodologies of ion exchange, liquid chromatography, and conductimetric determination with eluant suppression. (Author/RE)

  1. Sensitive and rapid high-performance liquid chromatography tandem mass spectrometry method for estimation of fulvestrant in rabbit plasma.

    PubMed

    Balaram, Varanasi Murali; Parmar, Dharmesh; Teja, Bulusu B; Rathnam, Shivprakash; Rao, Jangala Venkateswara; Dasandi, Bhavesh

    2010-08-01

    A rapid and sensitive high-performance liquid chromatography and electrospray tandem mass spectrometry method was developed and validated for estimation of fulvestrant in rabbit plasma using liquid-liquid extraction. The separation and quantification of fulvestrant were achieved by reverse-phase chromatography on a Sunfire C18 column (50 x 2.1. i.d., 3.5 microm) with isocratic elution at a flow rate of 300 microL/min using norethistrone as an internal standard from 500 microL plasma sample. The method was validated over the concentration range from 0.092 to 16.937 ng/mL with a lower limit of detection of 0.023 ng/mL. The intra-day and inter-day accuracy and precision were within 10%. The recovery was 85 and 90% for fulvestrant and norethistrone respectively. The chromatographic run time was only 2.5 min.

  2. Identification of atmospheric organic sources using the carbon hollow tube-gas chromatography method and factor analysis

    SciTech Connect

    Cobb, G.P.; Braman, R.S.; Gilbert, R.A. )

    1989-04-15

    Atmospheric organics were sampled and analyzed by using the carbon hollow tube-gas chromatography method. Chromatograms from spice mixtures, cigarettes, and ambient air were analyzed. Principal factor analysis of row order chromatographic data produces factors which are eigenchromatograms of the components in the samples. Component sources are identified from the eigenchromatograms in all experiments and the individual eigenchromatogram corresponding to a particular source is determined in most cases. Organic sources in ambient air and in cigaretts are identified with 87% certainty. Analysis of clove cigarettes allows the determination of the relative amount of clove in different cigarettes. A new nondestructive quality control method using the hollow tube-gas chromatography analysis is discussed.

  3. Liquid chromatography-tandem mass spectrometry method for the determination of anthelmintics in alfalfa plants.

    PubMed

    Islam, M Dabalus; Haberhauer, G; Gerzabek, M; Cannavan, A

    2012-01-01

    A simple and inexpensive liquid chromatography-tandem mass spectrometric method for the determination of anthelmintics in alfalfa plants (Medicago sativa L.) was developed and validated. Anthelmintics in plant leaves and stems (green chops) were extracted with methanol/acetonitrile (7:3, v/v) followed by a concentration and clean-up step using solid-phase extraction (Strata-X, 500 mg, 6 ml cartridge). After drying with nitrogen gas, the adsorbed analytes were eluted with methanol/acetonitrile (50:50, v/v) mixture followed by 100% acetonitrile. Chromatographic separation was achieved on an Atlantis T-3 (2.1 × 100 mm × 3 µm) analytical column with a Phenomenex guard cartridge (C8, 4 × 3 mm) attached to a Waters triple quadrupole mass spectrometer operated in positive electrospray ionisation mode with selected reaction monitoring. Samples were analysed using gradient elution at a flow rate of 0.35 ml min⁻¹. The mobile phase consisted of a 10 mM ammonium formate solution in (A) water/acetonitrile (90:10, v/v) and (B) methanol/acetonitrile (50:50, v/v). The method was validated for levamisole, fenbendazole, fenbendazole sulphoxide and fenbendazole sulphone at 10, 20 and 40 µg kg⁻¹ and for eprinomectin at 20, 40 and 80 µg kg⁻¹. Limits of quantification (LOQ) were 10 µg kg⁻¹ for all analytes except eprinomectin, which had an LOQ of 20 µg kg⁻¹. The overall mean recovery in green plants was between 74.2% and 81.4% with repeatabilities ranging from 2.2% to 19.1% and reproducibilities in the range 3.8-8.7%. The validated method was applied to plant samples in a study on the behaviour of anthelmintic drugs in a soil, plant and water system.

  4. Use of thermal desorption/gas chromatography as a performance-based screening method for petroleum hydrocarbons

    SciTech Connect

    Slavin, P.J. |; Crandall, K.; Dawson, L.; Kottenstette, R.; Wade, M. |

    1996-08-01

    Thermal desorption/gas chromatography (TD/GC) was used to screen soil samples on site for total petroleum hydrocarbon (TPH) content during a RCRA Facility Investigation (RFI). It proved to be a rapid, cost- effective tool for detecting non-aromatic mineral oil in soil. The on- site TD/GC results correlated well with those generated at an off- site laboratory for samples analyzed in accordance with EPA Method 418.1.

  5. Comparison of standard methods and gas chromatography method in determination of formaldehyde emission from MDF bonded with formaldehyde-based resins.

    PubMed

    Kim, Sumin; Kim, Hyun-Joong

    2005-09-01

    Formaldehyde emissions from MDF bonded with urea-formaldehyde resin (UF), melamine-formaldehyde resin (MF) and the co-polycondensation resin of urea-melamine-formaldehyde (UMF) and melamine-formaldehyde, measured by the Japanese standard method of determining formaldehyde emission with a desiccator (JIS A 5908) and the DIN EN 120 (European Committee For Standardization, 1991) method using the perforator value, were used as the typical standard methods. While the UF resin showed a desiccator value of 7.05 ppm and a perforator value of 12.1 mg/100 g panel, the MF resin exhibited a desiccator value of 0.6 ppm and a perforator value of 2.88 mg/100 g panel. According to the Japanese industrial standard and the European standard, the formaldehyde emission level of the MDF panels made with UF resin in this study was E(2) grade. The formaldehyde emission level was dramatically reduced by the addition of MF resin. This is because the addition of formaldehyde to melamine occurs more easily and completely than its addition to urea, even though the condensation reaction of melamine with formaldehyde is similar to that between urea and formaldehyde. These two methods, the desiccator method and the perforator method, produced proportionally equivalent results. Gas chromatography, a more sensitive and advanced method, was also used. The samples used for gas chromatography were gathered during the experiment involving the perforator method. The formaldehyde emission levels obtained from gas chromatography were similar to those obtained from the perforator method. The formaldehyde contents measured by gas chromatography were directly proportional to the perforator values.

  6. Green chromatography.

    PubMed

    Płotka, Justyna; Tobiszewski, Marek; Sulej, Anna Maria; Kupska, Magdalena; Górecki, Tadeusz; Namieśnik, Jacek

    2013-09-13

    Analysis of organic compounds in samples characterized by different composition of the matrix is very important in many areas. A vast majority of organic compound determinations are performed using gas or liquid chromatographic methods. It is thus very important that these methods have negligible environmental impact. Chromatographic techniques have the potential to be greener at all steps of the analysis, from sample collection and preparation to separation and final determination. The paper summarizes the approaches used to accomplish the goals of green chromatography. While complete elimination of sample preparation would be an ideal approach, it is not always practical. Solventless extraction techniques offer a very good alternative. Where solvents must be used, the focus should be on the minimization of their consumption. The approaches used to make chromatographic separations greener differ depending on the type of chromatography. In gas chromatography it is advisable to move away from using helium as the carrier gas because it is a non-renewable resource. GC separations using low thermal mass technology can be greener because of energy savings offered by this technology. In liquid chromatography the focus should be on the reduction of solvent consumption and replacement of toxic and environmentally hazardous solvents with more benign alternatives. Multidimensional separation techniques have the potential to make the analysis greener in both GC and LC. The environmental impact of the method is often determined by the location of the instrument with respect to the sample collection point.

  7. Development of a rapid diagnostic method for identification of Staphylococcus aureus and antimicrobial resistance in positive blood culture bottles using a PCR-DNA-chromatography method.

    PubMed

    Ohshiro, Takeya; Miyagi, Chihiro; Tamaki, Yoshikazu; Mizuno, Takuya; Ezaki, Takayuki

    2016-06-01

    Blood culturing and the rapid reporting of results are essential for infectious disease clinics to obtain bacterial information that can affect patient prognosis. When gram-positive coccoid cells are observed in blood culture bottles, it is important to determine whether the strain is Staphylococcus aureus and whether the strain has resistance genes, such as mecA and blaZ, for proper antibiotic selection. Previous work led to the development of a PCR method that is useful for rapid identification of bacterial species and antimicrobial susceptibility. However, that method has not yet been adopted in community hospitals due to the high cost and methodological complexity. We report here the development of a quick PCR and DNA-chromatography test, based on single-tag hybridization chromatography, that permits detection of S. aureus and the mecA and blaZ genes; results can be obtained within 1 h for positive blood culture bottles. We evaluated this method using 42 clinical isolates. Detection of S. aureus and the resistance genes by the PCR-DNA-chromatography method was compared with that obtained via the conventional identification method and actual antimicrobial susceptibility testing. Our method had a sensitivity of 97.0% and a specificity of 100% for the identification of the bacterial species. For the detection of the mecA gene of S. aureus, the sensitivity was 100% and the specificity was 95.2%. For the detection of the blaZ gene of S. aureus, the sensitivity was 100% and the specificity was 88.9%. The speed and simplicity of this PCR-DNA-chromatography method suggest that our method will facilitate rapid diagnoses.

  8. Advances in analytical techniques for neutron capture therapy: thin layer chromatography matrix and track etch thin layer chromatography methods for boron-10 analysis

    SciTech Connect

    Schremmer, J.M.; Noonan, D.J.

    1987-09-01

    A new track etch autoradiographic technique for quantitating boron-10 containing compounds used for neutron capture therapy is described. Instead of applying solutions of Cs2B12H11SH and its oxidation products directly to solid-state nuclear track detectors, diethylaminoethyl cellulose thin layer chromatography (TLC) plates are utilized as sample matrices. The plates are juxtaposed with Lexan polycarbonate detectors and irradiated in a beam of thermal neutrons. The detectors are then chemically etched, and the resultant tracks counted with an optoelectronic image analyzer. Sensitivity to boron-10 in solution reaches the 1 pg/microliter level, or 1 ppb. In heparinized blood samples, 100 pg boron-10/microliter are detected. This TLC matrix method has the advantage that sample plates can be reanalyzed under different reactor conditions to optimize detector response to the boron-10 carrier material. Track etch/TLC allows quantitation of the purity of boron neutron capture therapy compounds by utilizing the above method with TLC plates developed in solvent systems that resolve Cs2B12H11SH and its oxidative analogs. Detectors irradiated in juxtaposition to the thin layer chromatograms are chemically etched, and the tracks are counted in the sample lane from the origin of the plate to the solvent front. A graphic depiction of the number of tracks per field yields a quantitative analysis of compound purity.

  9. [THE DEVELOPMENT AND VALIDATION OF THE GAS CHROMATOGRAPHY METHOD AND VALIDATION DEVELOPMENT FOR THE ORGANIC CYANIDE (CYANOETHYLENE) DETERMINATION IN EXPIRED AIR].

    PubMed

    Zaĭtseva, N V; Ulanova, T S; Nurislamova, T V; Popova, N A; Mal'tseva, O A

    2015-01-01

    There are presented results of experimental studies on the development of gas chromatography method for the cyan ethylene determination in expired air During the process of the study there was chosen and proved the capillary gas chromatography method; there were investigated and elaborated optimal parameters of the gas chromatography separation of cyanoethylene with associated hydrocarbons together with the sample preparation and quantitative measurement methods. There was achieved the optimal level of gas chromatography quantification method for the cyan ethylene determination at 0, 00012 mg/m3, with the method uncertainty not more than 25%. The method was tried during the medical and biological examination of groups of 6-8 years old children, living in the territory of exposition from the moment of the birth and in the control territory.

  10. The use of multivariate curve resolution methods to improve the analysis of muramic acid as bacterial marker using gas chromatography-mass spectrometry: an alternative method to gas chromatography-tandem mass spectrometry.

    PubMed

    Moazeni-Pourasil, Roudabeh Sadat; Piri, Farhad; Ghassempour, Alireza; Jalali-Heravi, Mehdi

    2014-02-15

    In analysis of muramic acid (MA) as bacterial marker, two dominant disturbing factors lead the researchers to use gas chromatography-tandem mass spectrometry (GC-MS/MS) technique instead of gas chromatography-mass spectrometry (GC-MS). These factors are the trace concentration of MA and fundamental disturbance of base line mass channels in GC-MS technique. This study aimed to utilize multivariate curve resolution (MCR) methods combined with GC-MS to improve the analysis of MA. First, the background and noise in GC-MS analysis were corrected and reduced using MCR methods. In addition, the MA overlapped peaks were resolved to its pure chromatographic and mass spectral profiles. Then the two-way response of each component was reconstructed by the outer product of the pure chromatographic and mass spectral profiles. The overall volume integration (OVI) method was used for quantitative determination. The MA peak area was decreased dramatically after the background correction and noise reduction. The findings severely ratify the appropriateness of using MCR techniques combined with GC-MS analysis as a simple, fast and inexpensive method for the analysis of MA in complex mixtures. The proposed method may be considered as an alternative method to GC-MS/MS for thorough analysis of the bacterial marker.

  11. Basic Principles of Chromatography

    NASA Astrophysics Data System (ADS)

    Ismail, Baraem; Nielsen, S. Suzanne

    Chromatography has a great impact on all areas of analysis and, therefore, on the progress of science in general. Chromatography differs from other methods of separation in that a wide variety of materials, equipment, and techniques can be used. [Readers are referred to references (1-19) for general and specific information on chromatography.]. This chapter will focus on the principles of chromatography, mainly liquid chromatography (LC). Detailed principles and applications of gas chromatography (GC) will be discussed in Chap. 29. In view of its widespread use and applications, high-performance liquid chromatography (HPLC) will be discussed in a separate chapter (Chap. 28). The general principles of extraction are first described as a basis for understanding chromatography.

  12. Development of a liquid chromatography-tandem mass spectrometry method for the determination of sulfite in food.

    PubMed

    Robbins, Katherine S; Shah, Romina; MacMahon, Shaun; de Jager, Lowri S

    2015-06-03

    Sulfites are widely used food preservatives that can cause severe reactions in sensitive individuals. As a result, the U.S. FDA requires that sulfites be listed on the label of any food product containing >10 mg/kg (ppm) sulfite (measured as sulfur dioxide). Currently, the optimized Monier-Williams (MW) method (AOAC Official Method 990.28) is the most common approach for determining sulfite concentrations in food samples. However, this method is time-consuming and lacks specificity in certain matrices. An improved rapid, sensitive, and selective method has been developed using electrospray ionization (ESI) high-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) for the determination of sulfite in various food matrices. A total of 12 different types of foods were evaluated. These included dried fruits and vegetables, frozen seafood, sweeteners, and juices. The matrix is extracted with a buffered formaldehyde solution, converting free and reversibly bound sulfite to the stable formaldehyde adduct, hydroxymethylsulfonate (HMS). Extracts are prepared for injection using a C18 SPE cartridge to remove any lipophilic compounds. HMS is then separated from other matrix components using hydrophilic interaction chromatography (HILIC) and detected using multiple reaction monitoring (MRM). The method was validated at 5 concentrations in 12 food matrices. Accuracy data showed spiked recoveries ranging from 84 to 115% in representative foods. Six commercially available sulfited products were analyzed using the LC-MS/MS method, as well as the MW method, to determine if differences exist.

  13. Evaluation of a liquid chromatography method for quality control of methylated cyclodextrins.

    PubMed

    Fougère, L; Elfakir, C; Lafosse, M

    2013-02-15

    Halo C18 column (fused core particles) and Chromolith RP18 column (monolith) were evaluated in liquid chromatography in order to analyze methylated-β-cyclodextrins (Me-β-CD) with various degrees of substitution, DS such as the number of methyl groups per cyclodextrin ring. Chromolith RP18 enables a performing analysis of Me-β-CD with low DS but is not suitable for dimethyl-β-cyclodextrins (DM-β-CD). On the other hand, Halo C18 column allows an improved fingerprint of CDs having a DS from 4.9 up to a value major than 14 and avoiding the use of various chromatographic systems. Thus, liquid chromatography performed with this column and an evaporative light scattering detector can be used as a generic system for methylated CD analysis. Moreover, fused core particles of Halo C18 column enables a rapid analysis and liquid chromatography coupled with electrospray-mass spectrometry appears as a powerful tool to determine co-elution and to characterize various isomers of complex methylated-β-cyclodextrin mixtures.

  14. A reverse-phase liquid chromatography/mass spectrometry method for the analysis of high-molecular-weight fructooligosaccharides.

    PubMed

    Harrison, Scott J; Fraser, Karl; Lane, Geoffrey A; Villas-Boas, Silas; Rasmussen, Susanne

    2009-12-01

    Many important crop and forage plants accumulate polymeric water-soluble carbohydrates as fructooligosaccharides (or fructans). We have developed an improved method for the analysis of the full fructan complement in plant extracts based on porous graphitized carbon chromatography coupled to negative electrospray ionization mass spectrometry. By the use of profile data collection and multiple charge state ions, the effective mass range of the ion trap was extended to allow for the analysis of very high-molecular-weight oligosaccharides. This method allows the separation and quantification of isomeric fructan oligomers ranging from degree of polymerization (DP) 3 to DP 49.

  15. Rapid method for hydrocarbon-type analysis of heavy oils and synthetic fuels by pyrolysis thin layer chromatography

    SciTech Connect

    Poirier, M.A.; George, A.E.

    1982-09-01

    This work describes a rapid method for hydrocargon-type analysis applying thin layer chromatography (TLC) to the pentane-soluble fraction *malthenes) of the petroleum and synthetic fuels boiling above 200/sup 0/C. The principal component types encountered in this paper are saturates (SA), aromatics (AR), (mono and di together) polynuclear aromatics (PNA) and polar material (PO). The method uses a Iatroscan TLC pyrolyzer which combines the resolution capabilities of TLC with the possibility of quantification by using a flame-ionization detector (FID). Comparison of the results with those obtained by the API-60 procedure is presented.

  16. Development of a liquid chromatography-mass spectrometry method for the high-accuracy determination of creatinine in serum.

    PubMed

    Stokes, Peter; O'Connor, Gavin

    2003-08-25

    An LC-MS method for the high-accuracy determination of creatinine in serum has been developed and used to provide results for an international measurement evaluation programme (IMEP) and the Comité Consultatif pour la Quantité de Matière (CCQM) international inter-laboratory studies. An assessment of different sample preparation methods, including ion-exchange chromatography, solid-phase extraction, plasma ultrafiltration and ethanol protein precipitation, revealed that no bias or reduced precision was associated with the quicker less extensive clean-up methods, when using liquid chromatography-isotope dilution mass spectrometry (LC-IDMS) for quantitation. A number of different calibration regimes were also investigated. External calibration was shown to provide adequate calibration for most routine analysis with a relative associated expanded uncertainty (k=2) of 6% at the 95% confidence level. The use of a non-isotopically labelled internal standard was shown to improve the relative expanded uncertainty (k=2) to 4%. However, the difference in retention time between the internal standard and the creatinine was such that a matrix interferent produced an observed bias of over 16%. The use of an isotopically labelled internal standard was shown to reduce any bias to less than 0.2% with an expanded uncertainty (k=2) of less than 0.3%. The developed method was then used, in a blind trial organised jointly by IMEP and CCQM, to determine the amount of creatinine in human serum. The method performed well against the established reference method of ion-exchange chromatography followed by derivatisation gas chromatography (GC)-IDMS. The observed difference between the values determined by LC-IDMS and the key comparison reference value (average of all the submitted results) was less than 0.3%. The biggest advantage of the described method is in the speed of analysis. With a chromatographic run time of less than 10 min and sample preparation consisting of a simple protein

  17. PDZ Affinity Chromatography: A general method for affinity purification of proteins based on PDZ domains and their ligands

    PubMed Central

    Walkup, Ward G.; Kennedy, Mary B.

    2014-01-01

    PDZ (PSD-95, DiscsLarge, ZO1) domains function in nature as protein binding domains within scaffold and membrane-associated proteins. They comprise ~ 90 residues and make specific, high affinity interactions with complementary C-terminal peptide sequences, with other PDZ domains, and with phospholipids. We hypothesized that the specific, strong interactions of PDZ domains with their ligands would make them well suited for use in affinity chromatography. Here we describe a novel affinity chromatography method applicable for the purification of proteins that contain PDZ domain-binding ligands, either naturally or introduced by genetic engineering. We created a series of affinity resins comprised of PDZ domains from the scaffold protein PSD-95, or from neuronal nitric oxide synthase (nNOS), coupled to solid supports. We used them to purify heterologously expressed neuronal proteins or protein domains containing endogenous PDZ domain ligands, eluting the proteins with free PDZ domain peptide ligands. We show that Proteins of Interest (POIs) lacking endogenous PDZ domain ligands can be engineered as fusion products containing C-terminal PDZ domain ligand peptides or internal, N- or C-terminal PDZ domains and then can be purified by the same method. Using this method, we recovered recombinant GFP fused to a PDZ-domain ligand in active form as verified by fluorescence yield. Similarly, chloramphenicol acetyltransferase (CAT) and β-Galactosidase (LacZ) fused to a C-terminal PDZ domain ligand or an N-terminal PDZ domain were purified in active form as assessed by enzymatic assay. In general, PDZ domains and ligands derived from PSD-95 were superior to those from nNOS for this method. PDZ Domain Affinity Chromatography promises to be a versatile and effective method for purification of a wide variety of natural and recombinant proteins. PMID:24607360

  18. PDZ affinity chromatography: a general method for affinity purification of proteins based on PDZ domains and their ligands.

    PubMed

    Walkup, Ward G; Kennedy, Mary B

    2014-06-01

    PDZ (PSD-95, DiscsLarge, ZO1) domains function in nature as protein binding domains within scaffold and membrane-associated proteins. They comprise ∼90 residues and make specific, high affinity interactions with complementary C-terminal peptide sequences, with other PDZ domains, and with phospholipids. We hypothesized that the specific, strong interactions of PDZ domains with their ligands would make them well suited for use in affinity chromatography. Here we describe a novel affinity chromatography method applicable for the purification of proteins that contain PDZ domain-binding ligands, either naturally or introduced by genetic engineering. We created a series of affinity resins comprised of PDZ domains from the scaffold protein PSD-95, or from neuronal nitric oxide synthase (nNOS), coupled to solid supports. We used them to purify heterologously expressed neuronal proteins or protein domains containing endogenous PDZ domain ligands, eluting the proteins with free PDZ domain peptide ligands. We show that Proteins of Interest (POIs) lacking endogenous PDZ domain ligands can be engineered as fusion products containing C-terminal PDZ domain ligand peptides or internal, N- or C-terminal PDZ domains and then can be purified by the same method. Using this method, we recovered recombinant GFP fused to a PDZ domain ligand in active form as verified by fluorescence yield. Similarly, chloramphenicol acetyltransferase (CAT) and β-Galactosidase (LacZ) fused to a C-terminal PDZ domain ligand or an N-terminal PDZ domain were purified in active form as assessed by enzymatic assay. In general, PDZ domains and ligands derived from PSD-95 were superior to those from nNOS for this method. PDZ Domain Affinity Chromatography promises to be a versatile and effective method for purification of a wide variety of natural and recombinant proteins.

  19. METHOD 332.0: DETERMINATION OF PERCHLORATE IN DRINKING WATER BY ION CHROMATOGRAPHY WITH SUPPRESSED CONDUCTIVITY AND ELECTROSPRAY IONIZATION MASS SPECTROMETRY

    EPA Science Inventory

    This method is applicable to the identification and quantitation of perchlorate in raw and finished drinking waters. The approach used is ion chromatography with suppressed conductivity and electrospray ionization mass spectrometry (IC-ESI/MS)

  20. EPA Method 507: Determination of Nitrogen- and Phosphorus-Containing Pesticides in Water by Gas Chromatography with a Nitrogen-Phosphorus Detector

    EPA Pesticide Factsheets

    Method 507 describes procedures for sample preparation and analysis using solvent extraction of organophosphate pesticides in drinking water samples which are analyzed using a gas chromatography –nitrogen-phosphorus detector (GC-NPD).

  1. EPA Method 8321B (SW-846): Solvent-Extractable Nonvolatile Compounds by High Performance Liquid Chromatography-Thermospray-Mass Spectrometry (HPLC-TS-MS) or Ultraviolet (UV) Detection

    EPA Pesticide Factsheets

    Method 8321B describes procedures for preparation and analysis of solid, aqueous liquid, drinking water and wipe samples using high performance liquid chromatography and mass spectrometry for extractable non-volatile compounds.

  2. Thermal desorption-gas chromatography-mass spectrometry method to determine phthalate and organophosphate esters from air samples.

    PubMed

    Aragón, M; Borrull, F; Marcé, R M

    2013-08-16

    A method based on thermal desorption-gas chromatography-mass spectrometry (TD-GC-MS) has been developed to determine four organophosphate esters, seven phthalate esters, and bis(2-ethylhexyl) adipate in the gas phase from harbour and urban air samples. The method involves the sampling of 1.5L of air in a Tenax TA sorbent tube followed by thermal desorption (using a Tenax TA cryogenic trap) coupled to gas chromatography-mass spectrometry. The repeatability of the method expressed as %RSD (n=3) is less than 15% and the MQLs are between 0.007μgm(-3) (DMP, TBP, BBP, TPP and DnOP) and 6.7μgm(-3) (DEHP). The method was successfully applied in two areas (urban and harbour) testing two and three points in each one, respectively. Some of these compounds were found in both urban and harbour samples. Di-(2-ethylhexyl)phthalate was the most abundant compound found in both areas at concentration levels between 6.7μgm(-3) and 136.4μgm(-3). This study demonstrates that thermal desorption is an efficient method for the determination of these semi-volatile compounds in the gas phase fraction of air samples.

  3. A Simple and Sensitive Method for Quantitative Measurement of Methylmalonic Acid by Turbulent Flow Chromatography and Tandem Mass Spectrometry

    PubMed Central

    Tecleab, AG; Schofield, RC; Ramanathan, LV; Carlow, Dean C

    2016-01-01

    A simple and sensitive method for the detection of methylmalonic acid in serum without derivatization has been developed. This method implements protein precipitation using methanol followed by additional sample clean up by turbulent flow liquid chromatography (TFLC). The sample was directly injected into the turbulent flow liquid chromatography tandem mass spectrometry system (TFLC-MS/MS) for online extraction followed by HPLC separation. The eluent was transferred to the mass spectrometer and ionized by heated electrospray negative ionization (HESI) and the analyte was quantified using a six-point calibration curve. The validated analytical measurement range (AMR) is 30–1,000 nMol/L. Dilutions of 10 and 200-fold were validated giving a clinical reportable range (CRR) of 30–200,000 nMol/L. The between-day and within-day imprecision values at concentrations spanning the AMR were less than 15%. This method was compared to an established LC-MS/MS method at a CLIA certified national reference laboratory and shows an excellent correlation with our TFLC-MS/MS method. PMID:27833786

  4. Determination of (R)-timolol in (S)-timolol maleate active pharmaceutical ingredient: validation of a new supercritical fluid chromatography method with an established normal phase liquid chromatography method.

    PubMed

    Marley, Adrian; Connolly, Damian

    2014-01-17

    An enantioselective supercritical fluid chromatography (SFC) method was developed and validated to meet the current European Pharmacopoeia requirements of a limit test for the determination of S-timolol maleate enantiomeric purity in timolol maleate drug substance. The developed method is presented as an alternative to the current normal phase high performance liquid chromatography (NP-HPLC) method described in the European Pharmacopoeia (Timolol Maleate Monograph). Using a 4.6mm×250mm Chiralcel OD-H (dp: 5μm) column and a mobile phase of (93:7) CO2/0.1% (v/v) TEA in MeOH delivered at 4.0mLmin(-1) resolution of 2.0 was achieved within 5min, representing a 3-fold reduction in run-time and an 11-fold reduction in solvent consumption relative to the NP-HPLC method. Method robustness was examined by the variation of flow rate (±0.5mLmin(-1)), column temperature (±5°C) and column back-pressure (±10bar) and resolution was maintained at ≥1.9 in all cases. R-timolol was resolved from all potential impurities and the limit of detection was improved by increasing the sample concentration threefold compared to the NP-HPLC method such that the method could detect the R-timolol enantiomer at 0.5% (w/w) with respect to S-timolol maleate. Additional validation parameters demonstrated that the potential of the method to be used for routine release testing of timolol maleate raw material for drug product manufacturing in which the quantitation of R-timolol impurity in S-timolol maleate drug substance would be a requirement.

  5. Stability-indicating High-performance Liquid Chromatography Method for Simultaneous Determination of Aminophylline and Chlorpheniramine Maleate in Pharmaceutical Formulations

    PubMed Central

    Ali, A.; Ahmed, M.; Mahmud, T.; Qadir, M. A.; Nadeem, K.; Saleem, A.

    2015-01-01

    The present work deals with the development and validation of method for simultaneous determination of antihistaminic drugs in pharmaceutical formulations. A precise, specific and accurate reverse phase-high-performance liquid chromatography method for the simultaneous measurement of aminophylline and chlorpheniramine maleate was developed. The separation of drugs was achieved on C-18 (5 μm, 250×4.6 mm) high-performance liquid chromatography column. The runtime for analysis was 10 min. Mobile phase is mixture containing dilute H2SO4:methanol (60:40% v/v) with flow rate adjusted at 1.5 ml/min. The detection of components was performed at a wavelength of 264 nm. Retention times of aminophylline and chlorphinramine maleate were found to be 2.00 and 3.25 min, respectively. Linearity was found in the range of 16-24 μg/ml for chlorpheniramine maleate and 102.4-153.6 μg/ml for aminophylline with a correlation coefficient of 0.9998 and 0.9996, respectively. High peak purity index of 99.99% indicated the complete separation of analytes in the presence of degradation products is justification of method stability. Linearity, accuracy, specificity, precision and robustness studies were performed for method validation. PMID:26798164

  6. Ion chromatography as highly suitable method for rapid and accurate determination of antibiotic fosfomycin in pharmaceutical wastewater.

    PubMed

    Zeng, Ping; Xie, Xiaolin; Song, Yonghui; Liu, Ruixia; Zhu, Chaowei; Galarneau, Anne; Pic, Jean-Stéphane

    2014-01-01

    A rapid and accurate ion chromatography (IC) method (limit of detection as low as 0.06 mg L(-1)) for fosfomycin concentration determination in pharmaceutical industrial wastewater was developed. This method was compared with the performance of high performance liquid chromatography determination (with a high detection limit of 96.0 mg L(-1)) and ultraviolet spectrometry after reacting with alizarin (difficult to perform in colored solutions). The accuracy of the IC method was established in the linear range of 1.0-15.0 mg L(-1) and a linear correlation was found with a correlation coefficient of 0.9998. The recoveries of fosfomycin from industrial pharmaceutical wastewater at spiking concentrations of 2.0, 5.0 and 8.0 mg L(-1) ranged from 81.91 to 94.74%, with a relative standard deviation (RSD) from 1 to 4%. The recoveries of effluent from a sequencing batch reactor treated fosfomycin with activated sludge at spiking concentrations of 5.0, 8.0, 10.0 mg L(-1) ranging from 98.25 to 99.91%, with a RSD from 1 to 2%. The developed IC procedure provided a rapid, reliable and sensitive method for the determination of fosfomycin concentration in industrial pharmaceutical wastewater and samples containing complex components.

  7. Stability-indicating High-performance Liquid Chromatography Method for Simultaneous Determination of Aminophylline and Chlorpheniramine Maleate in Pharmaceutical Formulations.

    PubMed

    Ali, A; Ahmed, M; Mahmud, T; Qadir, M A; Nadeem, K; Saleem, A

    2015-01-01

    The present work deals with the development and validation of method for simultaneous determination of antihistaminic drugs in pharmaceutical formulations. A precise, specific and accurate reverse phase-high-performance liquid chromatography method for the simultaneous measurement of aminophylline and chlorpheniramine maleate was developed. The separation of drugs was achieved on C-18 (5 μm, 250×4.6 mm) high-performance liquid chromatography column. The runtime for analysis was 10 min. Mobile phase is mixture containing dilute H2SO4:methanol (60:40% v/v) with flow rate adjusted at 1.5 ml/min. The detection of components was performed at a wavelength of 264 nm. Retention times of aminophylline and chlorphinramine maleate were found to be 2.00 and 3.25 min, respectively. Linearity was found in the range of 16-24 μg/ml for chlorpheniramine maleate and 102.4-153.6 μg/ml for aminophylline with a correlation coefficient of 0.9998 and 0.9996, respectively. High peak purity index of 99.99% indicated the complete separation of analytes in the presence of degradation products is justification of method stability. Linearity, accuracy, specificity, precision and robustness studies were performed for method validation.

  8. Chromatographic fingerprint analysis of metabolites in natural and artificial agarwood using gas chromatography-mass spectrometry combined with chemometric methods.

    PubMed

    Gao, Xiaoxia; Xie, Mingrong; Liu, Shaofeng; Guo, Xiaoling; Chen, Xiaoying; Zhong, Zhaojian; Wang, Lei; Zhang, Weimin

    2014-09-15

    Agarwood is a resinous material formed in wounded Aquilaria sinensis in China, which is widely used as an effective traditional Chinese medicine (TCM). This study is aimed to use gas chromatography-mass spectrometry combined with chemometric methods to create reliable criteria for accurate identification of natural agarwood and artificial agarwood, as well as for quality evaluation of artificial agarwood. Natural agarwood and artificial agarwood (stimulated by formic acid or formic acid plus fungal inoculation) were used as standards and controls for the gas chromatography-mass spectrometry (GC-MS) and multivariate analysis. The identification criteria developed were applied to commercial agarwood. A reliable criteria including correlation coefficient of GC-MS fingerprint of natural agarwood and 22 markers of metabolism in natural and artificial agarwood was constructed. Compared with chemically stimulated agarwood (formic acid) and in terms of the 22 markers, artificial agarwood obtained by formic acid stimulation and fungal inoculation were much closer to natural agarwood. The study demonstrates that the chemical components of artificial agarwood obtained by comprehensive stimulated method (formic acid plus fungal inoculation) are much closer to the natural agarwood than those obtained by chemically stimulated method (formic acid), as times goes by. A reliable criteria containing correlation coefficient of GC-MS fingerprint of natural agarwood and 22 metabolism markers can be used to evaluate the quality of the agarwood. As an application case, three samples were identified as natural agarwood from the 25 commercial agarwood by using the evaluation method.

  9. A novel derivatization-free method of formaldehyde and propylene glycol determination in hydrogels by liquid chromatography with refractometric detection.

    PubMed

    Isakau, Henadz; Robert, Marielle; Shingel, Kirill I

    2009-04-05

    The paper describes the development and validation of a new derivatization-free liquid chromatography method for simultaneous determination of propylene glycol and formaldehyde in the formulations containing formaldehyde-releasing preservative. Highly swollen hydrogel made of poly(ethylene glycol)-protein conjugates was taken as a model formulation for integration of the propylene glycol and the diazolydinyl urea as formaldehyde releaser. The method is shown to be simple and selective and, more importantly, allows determining an existing level of formaldehyde at the moment of analysis instead of all available formaldehyde that might be released during chemical derivatization. After liquid extraction the propylene glycol (PG) and formaldehyde (FA) amounts are determined chromatographically on a Shodex SH 1011 ligand-exchange column using 0.01 M sulfuric acid mobile phase, a flow rate of 1.0 ml/min and RI detection. The assay is validated showing good linearity, precision, and accuracy. The limits of detection of formaldehyde and propylene glycol in the analyzed solutions were estimated to be 25 ng and 87 ng, respectively. This analytical assay is considered useful for product stability studies and in developing new formaldehyde releaser-containing formulations where the concentration of formaldehyde is a presumable subject of labeling requirements. This method can also provide a rapid and convenient alternative to gas chromatography method of propylene glycol quantification.

  10. Methods of Analysis - Determination of Pyrethroid Insecticides in Water and Sediment Using Gas Chromatography/Mass Spectrometry

    USGS Publications Warehouse

    Hladik, Michelle; Smalling, Kelly L.; Kuivila, Kathryn

    2009-01-01

    A method for the determination of 14 pyrethroid insecticides in environmental water and sediment samples is described. The method was developed by the U.S. Geological Survey in response to increasing concern over the effects of pyrethroids on aquatic organisms. The pyrethroids included in this method are ones that are applied to many agricultural and urban areas. Filtered water samples are extracted for pyrethroids using solid-phase extraction (SPE) with no additional cleanup steps. Sediment and soil samples are extracted using a microwave-assisted extraction system, and the pyrethroids of interest are separated from co-extracted matrix interferences by passing the extracts through stacked graphitized carbon and alumina SPE cartridges, along with the use of high-performance liquid chromatography and gel-permeation chromatography (HPLC/GPC). Quantification of the pyrethroids from the extracted water and sediment samples is done using gas chromatography with mass spectrometry (GC/MS) or gas chromatography with tandem mass spectrometry (GC/MS/MS). Recoveries in test water samples fortified at 10 ng/L ranged from 83 to 107 percent, and recoveries in test sediment samples fortified at 10 ug/kg ranged from 82 to 101 percent; relative standard deviations ranged from 5 to 9 percent in the water samples and 3 to 9 percent in the sediment samples. Method detection limits (MDLs), calculated using U.S. Environmental Protection Agency procedures (40 CFR 136, Appendix B), in water ranged from 2.0 to 6.0 ng/L using GC/MS and 0.5 to 1.0 ng/L using GC/MS/MS. For sediment, the MDLs ranged from 1.0 to 2.6 ug/kg dry weight using GC/MS and 0.2 to 0.5 ug/kg dry weight using GC/MS/MS. The matrix-spike recoveries for each compound, when averaged for 12 environmental water samples, ranged from 84 to 96 percent, and when averaged for 27 environmental sediment samples, ranged from 88 to 100 percent.

  11. Portable system and method combining chromatography and array of electrochemical sensors

    DOEpatents

    Zaromb, Solomon; Stetter, Joseph R.

    1989-01-01

    A portable system for analyzing a fluid sample includes a small, portable, low-pressure and low-power chromatographic analyzer and a chemical parameter spectrometry monitor including an array of sensors for detecting, identifying and measuring the concentrations of a variety of components in the eluent from the chromatographic analyzer. The monitor includes one or more operating condition controllers which may be used to change one or more of the operating conditions during exposure of the sensors to the eluent from the chromatography analyzer to form a response pattern which is then compared with a library of previously established patterns. Gas and liquid chromatographic embodiments are disclosed. In the gas embodiment, the operating condition controllers include heated filaments which may convert electrochemically inactive components to electrochemically active products. In the liquid chromatography embodiment, low-power, liquid-phase equivalents of heated filaments are used with appropriate sensors. The library response patterns may be divided into subsets and the formed pattern may be assigned for comparison only with the patterns of a particular subset.

  12. Mathematical method for the prediction of retention times of fatty acid methyl esters in temperature-programmed capillary gas chromatography.

    PubMed

    Torres, Alexandre G; Trugo, Nádia M F; Trugo, Luiz C

    2002-07-17

    An accurate method for identification of fatty acids in complex mixtures analyzed by temperature-programmed capillary gas chromatography is described. The method is based on a mathematical approach using regression curves obtained by plotting the relative retention times of fatty acid methyl esters (FAMEs) analyzed in isothermal and gradient temperature conditions. The method was applied to a complex biological sample (human milk), and it was possible to identify 64 fatty acids, including branched-chain and other fatty acids for which reference standards were not readily available. The identities of the majority of the peaks were confirmed by mass spectrometry. The relative residuals and the relative differences between estimated and measured relative retention times of individual FAMEs varied from 0.03 to 3.15% and from 0.0 to 2.9%, respectively. The method is useful for identification of fatty acids in routine analysis.

  13. /Chromatography+RECOVERY=superresolution chromatography

    NASA Astrophysics Data System (ADS)

    Kosarev, E. L.; Muranov, K. O.

    2003-04-01

    A method for improving the resolution of the chromatographic analysis based on deriving the point-spread function of a chromatographic column, i.e., a chromatogram of an individual compound, is described. The system of two data sets, namely, a chromatogram of a substance analyzed and a point-spread function of a chromatographic column in combination with the noise statistics, makes it possible to use the RECOVERY signal-reconstruction software package described in paper by Gelfgat et al. (Comp. Phys. Commun. 74 (1993) 335). The proposed method has been tested by chromatography of bovine serum albumin using gel filtration. The resultant resolution exceeds that reached using high-performance liquid chromatography (with the cost of the instruments being lower by a factor of 15-20).

  14. Novel rapid liquid chromatography tandem masspectrometry method for vemurafenib and metabolites in human plasma, including metabolite concentrations at steady state.

    PubMed

    Vikingsson, Svante; Strömqvist, Malin; Svedberg, Anna; Hansson, Johan; Höiom, Veronica; Gréen, Henrik

    2016-08-01

    A novel, rapid and sensitive liquid chromatography tandem-mass spectrometry method for quantification of vemurafenib in human plasma, that also for the first time allows for metabolite semi-quantification, was developed and validated to support clinical trials and therapeutic drug monitoring. Vemurafenib was analysed by precipitation with methanol followed by a 1.9 min isocratic liquid chromatography tandem masspectrometry analysis using an Acquity BEH C18 column with methanol and formic acid using isotope labelled internal standards. Analytes were detected in multireaction monitoring mode on a Xevo TQ. Semi-quantification of vemurafenib metabolites was performed using the same analytical system and sample preparation with gradient elution. The vemurafenib method was successfully validated in the range 0.5-100 μg/mL according to international guidelines. The metabolite method was partially validated owing to the lack of commercially available reference materials. For the first time concentration levels at steady state for melanoma patients treated with vemurafenib is presented. The low abundance of vemurafenib metabolites suggests that they lack clinical significance. Copyright © 2016 John Wiley & Sons, Ltd.

  15. Fast, simple, and sensitive high-performance liquid chromatography method for measuring vitamins A and E in human blood plasma.

    PubMed

    Yuan, Chao; Burgyan, Maria; Bunch, Dustin R; Reineks, Edmunds; Jackson, Raymond; Steinle, Roxanne; Wang, Sihe

    2014-09-01

    Vitamins A and E are fat-soluble vitamins that play important roles in several physiological processes. Monitoring their concentrations is needed to detect deficiency and guide therapy. In this study, we developed a high-performance liquid chromatography method to measure the major forms of vitamin A (retinol) and vitamin E (α-tocopherol and γ-tocopherol) in human blood plasma. Vitamins A and E were extracted with hexane and separated on a reversed-phase column using methanol as the mobile phase. Retinol was detected by ultraviolet absorption, whereas tocopherols were detected by fluorescence emission. The chromatographic cycle time was 4.0 min per sample. The analytical measurement range was 0.03-5.14, 0.32-36.02, and 0.10-9.99 mg/L for retinol, α-tocopherol, and γ-tocopherol, respectively. Intr-aassay and total coefficient of variation were <6.0% for all compounds. This method was traceable to standard reference materials offered by the National Institute of Standards and Technology. Reference intervals were established using plasma samples collected from 51 healthy adult donors and were found to be 0.30-1.20, 6.0-23.0, and 0.3-3.2 mg/L for retinol, α-tocopherol, and γ-tocopherol, respectively. In conclusion, we developed and validated a fast, simple, and sensitive high-performance liquid chromatography method for measuring the major forms of vitamins A and E in human plasma.

  16. A fast liquid chromatography-mass spectrometry (LC-MS) method for quantification of major polar metabolites in plants.

    PubMed

    Liu, Zhiqian; Rochfort, Simone

    2013-01-01

    Current liquid chromatography (LC) based methods for the analysis of polar plant metabolites require multiple runs using complex mobile phases and a combination of different columns. Here we describe a fast liquid chromatography-mass spectrometry (LC-MS) method for the determination of major polar metabolites in plants that requires only a single run using a single column. The method takes advantage of the ability to acquire both positive and negative data in an ion trap mass spectrometer (MS) and also the accurate mass capability of the orbitrap MS. The separation of polar compounds is achieved with a polar, reversed-phase column (Synergi Hydro-RP). A single analysis with a 25min runtime is able to reliably determine the level of nearly all essential amino acids, several major organic acids and several major sugars in plant materials, as exemplified by analysis of a perennial ryegrass extract. The level of detection on column was below 0.1ng (average 0.03ng) for most amino acids, below 5ng (average 2.3ng) for organics acids and below 1ng (average 0.64ng) for sugars. The levels of quantified metabolites in ryegrass varied from 22μg/g dry weight for histidine to 41mg/g dry weight for sucrose.

  17. Dating of seals produced with stamp-pad ink using gas chromatography method.

    PubMed

    Li, Biao

    2014-09-01

    Seals are often placed on important documents, and determining the date when a seal was affixed can be important to assess the authenticity of a document. In this study, thirty-seven different brands of stamp-pad inks and three different types of paper were used to make diachronic samples of the seals. The volatile components of stamp-pad ink were determined, and the stamp-pad ink was classified by gas chromatography with a flame ionization detector. Calibration curves were created to show the relationship between the content of volatile stamp-pad ink components and the age of stamp-pad ink entries stored under natural aging, UV-induced aging, and heating aging conditions. The experimental results showed that GC was useful in the analysis of stamp-pad ink and applicable for determining the relative age of stamp-pad ink entries in some cases where the seal was placed on the document within 3 months.

  18. Development and Validation of an Affinity Chromatography-Protein G Method for IgG Quantification

    PubMed Central

    Paradina Fernández, Lesly; Calvo, Loany; Viña, Lisel

    2014-01-01

    Nimotuzumab, an IgG that recognizes the epidermal growth factor receptor (EGF-R) overexpressed in some tumors, is used in the treatment of advanced head and neck cancer. For the quantification of this protein in cell culture supernatants, protein G-HPLC affinity chromatography is used due to its high affinity and specificity for antibodies of this class. The technique relies on the comparison of the area under the curve of the elution peak of the samples to be evaluated versus to a calibration curve of well-known concentrations and was validated by assessment of its robustness, specificity, repeatability, intermediate precision, accuracy, linearity, limit of detection, limit of quantification, and range. According to results of the study all validation parameters fulfilled the preestablished acceptance criteria and demonstrated the feasibility of the assay for the analysis of samples of cell culture supernatant as well as drug product. PMID:27379284

  19. Novel materials and methods for solid-phase extraction and liquid chromatography

    SciTech Connect

    Ambrose, Diana

    1997-06-24

    This report contains a general introduction which discusses solid-phase extraction and solid-phase micro-extraction as sample preparation techniques for high-performance liquid chromatography, which is also evaluated in the study. This report also contains the Conclusions section. Four sections have been removed and processed separately: silicalite as a sorbent for solid-phase extraction; a new, high-capacity carboxylic acid functionalized resin for solid-phase extraction; semi-micro solid-phase extraction of organic compounds from aqueous and biological samples; and the high-performance liquid chromatographic determination of drugs and metabolites in human serum and urine using direct injection and a unique molecular sieve.

  20. [Affinity chromatography and proteomic screening as the effective method for S100A4 new protein targets discovery].

    PubMed

    Koshelev, Iu A

    2014-01-01

    Affinity chromatography followed by a selective binding proteins identification can be using as effective method for a biological impotent interactions discovery. The molecular structure and their surface charge as and conformational regulation possibilities, which change their surface hydrophobic properties, all they should to taken in account during method optimization process. With the same' method we had identify some new S100A4 target proteins such as cytoskeleton proteins Sept2, Sept7, Sept11 and this interaction would can to highlight as S100A4 would regulate cell motility. Even we had identify the transcription cofactor Ddx5 and through such complex formation a S100A4 protein would can to regulate E-cadherin, p21 Waf1/Cip1), Bnip3 gene expression. The same protocol can be using for a target proteins search with another S100 protein family members, because their molecules demonstrate a high homology level in amino aside sequences and 3D structures.

  1. Liquid chromatography-tandem mass spectrometry screening method for the simultaneous detection of stimulants and diuretics in urine.

    PubMed

    Hsu, Ku Fu; Chien, Kuei-Yu; Chang-Chien, Guo Ping; Lin, Su Fan; Hsu, Pei Hsuan; Hsu, Mei-Chich

    2011-11-01

    This study established a simultaneous screening method based on solid-phase extraction and liquid chromatography-tandem mass spectrometry (LC-MS-MS) for the detection of 23 stimulants and 23 diuretics in human urine. An electrospray ionization source and multiple reaction monitoring were used for data acquisition. All stimulants and diuretics were separated in less than 12.52 min. The limits of detection were in the range of 25-500 ng/mL for stimulants and 25-125 ng/mL for diuretics. To evaluate the performance of this method, urine samples were collected from 1627 athletes in Taiwan, and 7 positive samples were found. This LC-MS-MS method not only meets the minimum required performance limits set by the World Anti-Doping Agency but also provides a fast way to analyze the authentic urine samples in doping control laboratories.

  2. Rapid and sensitive method for analysis of nitrate in meat samples using ultra performance liquid chromatography-mass spectrometry.

    PubMed

    Siddiqui, Masoom Raza; Wabaidur, Saikh Mohammad; ALOthman, Zeid A; Rafiquee, M Z A

    2015-12-05

    A sensitive and selective ultra performance liquid chromatography-mass spectrometric method has been developed for the quantitative analysis of nitrate in meat samples. Selected ion reaction (SIR) mode was adopted to identify and quantify the nitrate. Chromatographic analyses were performed on a BEH C-18 column with a mobile phase consisting of a surfactant (Cetylpyridinium chloride) and acetonitrile in equal ratio (50/50, v/v) at a flow rate of 0.4 mL min(-1). The limit of detection and limit of quantitation of the developed method was found to be 0.0599 and 0.1817 mg kg(-1), respectively. The linearity of the proposed method was checked in the concentration range of 0.5-10 mg kg(-1) with an excellent correlation coefficient (r) of 0.997. The recovery of the nitrate in the meat samples were in the range of 98.02-98.99%.

  3. A Supercritical Fluid Chromatography/Tandem Mass Spectrometry Method for the Simultaneous Quantification of Metformin and Gliclazide in Human Plasma

    PubMed Central

    Agrawal, Y. K.; Gogoi, P. J.; Manna, K.; Bhatt, H. G.; Jain, V. K.

    2010-01-01

    Present study reports the development and validation of a simultaneous estimation of metformin and gliclazide in human plasma using supercritical fluid chromatography followed by tandem mass spectrometry. Acetonitrile:water (80:20) mixture was used as a mobile phase along with liquid CO2 in supercritical fluid chromatography and phenformin as an internal standard. The modified plasma samples were analyzed by electro-spray ionization method in selective reaction monitoring mode in tandem mass spectrometry. Supercritical fluid chromatographic separation was performed using nucleosil C18 containing column as a stationary phase. The separated products were identified by characteristic peaks and specific fragments peaks in tandem mass spectrometry as m/z 130 to 86 for metformin, m/z 324 to 110 for gliclazide and m/z 206 to 105 for phenformin. The present method was found linear in the concentration ranges of 6.0-3550 ng/ml and 7.5-7500 ng/ml for metformin and gliclazide, respectively. Pharmacokinetic study was performed after an oral administration of dispersible tablets containing 500 mg of metformin and 80 mg of gliclazide using same techniques. PMID:20582190

  4. A convenient method for epichlorohydrin determination in water using headspace-solid-phase microextraction and gas chromatography.

    PubMed

    Lasa, M; Garcia, R; Millán, E

    2006-08-01

    A simple procedure for epychlorohydrin determination in water is presented. In order to optimize the epichlorohydrin extraction conditions in water using headspace (HS)-solid-phase microextraction (SPME), followed by gas chromatography, an experimental design in two steps is performed. Firstly, a 2(5-2) fractional factorial design for screening the significant variables is used. Secondly, a central composite design for optimizing them is carried out. The best experimental conditions are the followings: poly(dimethysiloxane)-divinylbenzene coating fiber; 20 min extraction time; 5 degrees C extraction temperature; 300 g/L sodium chloride; and 20 mL HS volume in a 40-mL vial. Using the previous extraction conditions with gas chromatography (GC)-flame ionization detection equipment, a limit of detection (LOD) of 1.8 microg/L and a relative standard deviation (RSD) of 3.8% (for 25 microg/L) are obtained. With a GC electron capture detection equipment the RSD is 6.6% (for 5 microg/L), and the LOD found is lower (0.08 microg/L). The method is applied to the analysis of water from four treatment plants at the entrance and effluent stream. The standard addition method is used to quantitate the epichlorohydrin that is found in the raw water of the three wastewater treatment plants.

  5. Proteomics-based method for the assessment of marine pollution using liquid chromatography coupled with two-dimensional electrophoresis.

    PubMed

    Amelina, Hanna; Apraiz, Itxaso; Sun, Wei; Cristobal, Susana

    2007-06-01

    Using a proteomic approach, we have developed a new method for the assessment of marine pollution that generates highly reproducible protein expression patterns and it is simple and scalable. The protocol is based on applying liquid chromatography (LC) coupled with two-dimensional electrophoresis (2-DE) to analyze changes in the protein expression pattern after exposure to marine pollution. The digestive gland of the sentinel "blue mussel" (Mytilus edulis) was batch-processed through a simple cell fractionation followed by ion-exchange chromatography and 2-DE. The selection of ligands, elution method, and small volume design was carefully considered to define a protocol that could be mainly robotized. A pilot study with samples collected from different Gothenburg harbor areas indicated that the clean area could be distinguished from the polluted ones based on a protein expression pattern (PES) composed of 13 proteins. Principal component analysis (PCA) and hierarchical clustering confirmed that the PES was sufficient to discriminate polluted and unpolluted areas and to provide a spatial gradient from the polluted source. Several proteins from the PES were identified by electrospray ionization tandem mass spectrometry (ESI-MS/MS), and they are involved in beta-oxidation, amino acid metabolism, detoxification, protein degradation, organelle biogenesis, and protein folding. In the near future, this methodology could show potential advantages to assess marine pollution and could become a stable platform to elucidate ecotoxicological questions.

  6. Retention mechanism assessment and method development for the analysis of iohexol and its related compounds in hydrophilic interaction liquid chromatography.

    PubMed

    Jovanović, Marko; Rakić, Tijana; Jančić-Stojanović, Biljana; Ivanović, Darko; Medenica, Mirjana

    2014-07-01

    Hydrophilic interaction liquid chromatography (HILIC) has emerged in recent years as a valuable alternative to reversed-phase liquid chromatography in the analysis of polar compounds. Research in HILIC is divided into two directions: the assessment of the retention mechanism and retention behavior, and the development of HILIC methods. In this work, four polar neutral analytes (iohexol and its related compounds A, B, and C) were analyzed on two silica and two diol columns in HILIC mode with the aim to investigate thoroughly the retention mechanisms and retention behavior of polar neutral compounds on these four columns. The adsorption and partition contribution to the overall HILIC retention mechanism was investigated by fitting the retention data to linear (adsorption and partition) and nonlinear (mixed-retention and quadratic) theoretical models. On the other hand, the establishment of empirical second-order polynomial retention models on the basis of D-optimal design made possible the estimation of the simultaneous influence of several mobile-phase-related factors. Furthermore, these models were also used as the basis for the application of indirect modeling of the selectivity factor and a grid point search approach in order to achieve the optimal separation of analytes. After the optimization goals had been set, the grids were searched and the optimal conditions were identified. Finally, the optimized method was subjected to validation.

  7. A Method for the Quantitation of Trace Levels of Dimethyl Sulfoxide in Urine by High Performance Liquid Chromatography

    DTIC Science & Technology

    1989-05-01

    HIGH PERFORMANCE LIQUID CHROMATOGRAPHY by...for the sample cleanup and concentration, followed by separation by reversed phase high performance liquid chromatography . EXPERIMENTAL Materials...DIMETHYL SULFOXIDE IN URINE BY HIGH PERFORMANCE LIQUID CHROMATOGRAPHY 4. AUTHORS (Last name, first name, middle initial. If military, show rank, e.g.

  8. Biomonitoring method for bisphenol A in human urine by ultra-high-performance liquid chromatography-tandem mass spectrometry.

    PubMed

    Anderson, David J; Brozek, Eric M; Cox, Kyley J; Porucznik, Christina A; Wilkins, Diana G

    2014-03-15

    An ultra-high-performance liquid chromatography-tandem mass spectrometry method for the measurement of total bisphenol A in human urine was developed and validated. The method utilized liquid/liquid extraction with 1-chlorobutane and a human urine aliquot size of 800μL. Chromatography was performed on an Acquity UPLC(®) system with a Kinetex(®) Phenyl-Hexyl column. Mass spectrometric analysis was with negative electrospray ionization on a Quattro Premier XE™. The surrogate matrix method was used for the preparation of calibration standards in synthetic urine due to the presence of BPA in control human urine. The validated calibration range was 0.75-20ng/mL with a limit of detection of 0.1ng/mL. The internal standard was d16-bisphenol A. Method validation utilized quality control samples at three concentrations in both synthetic urine and human urine. Bisphenol A mono-glucuronide was fortified in synthetic urine in each analytical run to monitor the enzymatic conversion of the glucuronide conjugate to BPA by β-glucuronidase. Validated method parameters included linearity, accuracy, precision, integrity of dilution, selectivity, re-injection reproducibility, recovery/matrix effect, solution stability, and matrix stability in human urine. Acceptance criteria for analytical standards and QCs were ±20% of nominal concentration. Matrix stability in human urine was validated after 24h at ambient temperature, after three freeze/thaw cycles, and after frozen storage at -20°C and -80°C for up to 218 days. The method has been applied to the analysis of over 1750 human urine samples from a biomonitoring study. The median and mean urine BPA concentrations were 2.71ng/mL and 4.75ng/mL, respectively.

  9. A fast, simple, and reliable hydrophilic interaction liquid chromatography method for the determination of ascorbic and isoascorbic acids.

    PubMed

    Barros, Ana I R N A; Silva, Ana P; Gonçalves, Berta; Nunes, Fernando M

    2010-03-01

    A reliable method for the determination of total vitamin C must be able to resolve ascorbic acid (AA) and the epimeric isoascorbic acid (IAA) and determine the sum of AA and its oxidized form dehydroascorbic acid. AA and IAA are polar molecules with a low retention time in conventional reversed phase systems, and hence of difficult resolution. Hydrophilic interaction chromatography using a TSKgel Amide-80 stationary phase with isocratic elution was successful in resolving the two epimers. The column was compatible with injections of high concentrations of metaphosphoric acid, tris(2-carboxyethyl)-phosphine, and EDTA without drift of baseline and retention time. Total AA and IAA were extracted, stabilized, and reduced in one step at 40 °C, using 5% m-phosphoric acid, 2 mM of EDTA, and 2 mM of tris(2-carboxyethyl)-phosphine as reducing agent. This simple, fast, and robust hydrophilic interaction chromatography-DAD method was applied for the analysis of food products namely fruit juices, chestnut, and ham and also in pharmaceutical and multivitamin tablets. Method validation was performed on the food products, including parameters of precision, accuracy, linearity, limit of detection, and quantification (LOQ). The absence of matrix interferences was assessed by the standard addition method and Youden calibration. The method was fast, accurate, and precise with a LOQ(AA) of 1.5 mg/L and LOQ(IAA) of 3.7 mg/L. The simple experimental procedure, completed in 1 h, the possibility of using IAA as an internal standard, and low probability of artifacts are the major advantages of the proposed method for the routine determination of these compounds in a large number of samples.

  10. Applying Chromatography.

    ERIC Educational Resources Information Center

    Klein, Jessie W.; Patev, Paul

    1998-01-01

    Presents three experiments to introduce students to different kinds of chromatography: (1) paper chromatography; (2) gel filtration chromatography; and (3) reverse-phase liquid chromatography. Written in the form of a laboratory manual, explanations of each of the techniques, materials needed, procedures, and a glossary are included. (PVD)

  11. A method to assess genomic DNA methylation using high-performance liquid chromatography/electrospray ionization mass spectrometry.

    PubMed

    Friso, Simonetta; Choi, Sang-Woon; Dolnikowski, Gregory G; Selhub, Jacob

    2002-09-01

    Eukaryotic DNA is methylated at some cytosine residues, and this epigenetic feature performs critical functions. We developed a method for quantitative determination of 5-methyl-2'-deoxycytidine in human DNA using liquid chromatography/electrospray ionization mass spectrometry (LC/ESI-MS). The DNA was enzymatically hydrolyzed by sequential digestion with three enzymes. DNA hydrolyzates were subsequently separated by reversed-phase high-performance liquid chromatography in isocratic mode. The four major DNA bases and 5-methyl-2'-deoxycytidine were resolved and eluted in 13 min. Identification of 2'-deoxycytidine and 5-methyl-2'-deoxycytidine was obtained by combined diode array UV spectra analysis and mass spectra of chromatographic peaks. The isotopomers [15N3]-2'-deoxycytidine and (methyl-d3,ring-6-d1)-5-methyl-2'-deoxycytidine were used as internal standards. Ions of m/z 126 and 130 were used to detect 5-methyl-2'-deoxycytidine and its isotopomer, and ions of m/z 112 and 115 were used to detect 2'-deoxycytidine and its stable isotopomer, respectively. The DNA methylation status was calculated on the basis of the amount of 5-methyl-2'-deoxycytidine per microgram of DNA with percent relative standard deviations (%RSD) for a method precision of 7.1 (within-day) and 5.7 (day-to-day). This method also allows the measurement of 5-methyl-2'-deoxycytidine expressed as a percentage of total deoxycytidine residues in genomic DNA with %RSD for method precision of 1.9 (within-day) and 1.7 (day-to-day). This LC/MS method for quantitative determination of genomic DNA methylation status is rapid, sensitive, selective, and precise.

  12. Gas chromatography.

    PubMed

    Eiceman, G A; Hill, H H; Gardea-Torresdey, J

    1998-06-15

    This review of the fundamental developments in gas chromatography (GC) includes articles published from 1996 and 1997 and an occasional citation prior to 1996. The literature was reviewed principally using CA Selects for Gas Chromatography from Chemical Abstracts Service, and some significant articles from late 1997 may be missing from the review. In addition, the online SciSearch Database (Institute for Scientific Information) capability was used to abstract review articles or books. As with the prior recent reviews, emphasis has been given to the identification and discussion of selected developments, rather than a presentation of a comprehensive literature search, now available widely through computer-based resources. During the last two years, several themes emerged from a review of the literature. Multidimensional gas chromatography has undergone transformation encompassing a broad range of activity, including attempts to establish methods using chromatographic principles rather than a totally empirical approach. Another trend noted was a comparatively large effort in chromatographic theory through modeling efforts; these presumably became resurgent with inexpensive and powerful computing tools. Finally, an impressive level of activity was noted through the themes highlighted in this review, and this was particularly true with detectors and field instruments.

  13. Preliminary validation of high performance liquid chromatography method for detection of methyl-testosterone residue in carp muscle

    NASA Astrophysics Data System (ADS)

    Jiang, Jie; Lin, Hong; Fu, Xiaoting; Li, Mingming

    2005-07-01

    The use of synthetic anabolic steroid methyltestosterone (MT) as growth promoter is prohibited in China. Validations of analytical methods for MT residue in food and the results obtained have become indispensable. The high performance liquid chromatography (HPLC) method for the detection of MT with liquid-liquid extraction by trichloromethane-methanol in carp muscle tissue was preliminarily validated with reference to the following parameters: recovery (accuracy) at the 1, 5 and l0 mgkg-1 level, between-run and within-run CV values (repeatability, also called relative standard deviation (RSD)) and limit of detection. The recoveries were above 80% and the between-run and within-run CV values below 10% for muscle tissue. The limit of detection was 0.05 mgkg-1.

  14. Determination of aflatoxins in hazelnuts by various sample preparation methods and liquid chromatography-tandem mass spectrometry.

    PubMed

    Bacaloni, Alessandro; Cavaliere, Chiara; Cucci, Francesca; Foglia, Patrizia; Samperi, Roberto; Laganà, Aldo

    2008-02-01

    A sensitive and reliable liquid chromatography-tandem mass spectrometric with electrospray ionization method for determining aflatoxins in hazelnuts has been developed. Three different extraction techniques, such as homogenization, ultrasonic extraction, and matrix solid phase dispersion have been tested and compared in terms of recovery, matrix effect, accuracy and precision. Ultrasound extraction was the most performing sample preparation method. Absolute recoveries for analytes and I.S. ranged from 93 to 101%. Accuracy and precision were calculated using matrix matched calibration, and ranged 91-102% and 2-11%, respectively. CC alpha and CC beta for aflatoxin B1 (EU limit=2 microg/kg) were 2.15 and 2.33 microg/kg, respectively. A ruggedness test performed on three other matrices demonstrated that sonication time was critical and a matrix matched calibration must be constructed for every sort of matrix.

  15. A new liquid chromatography method with charge aerosol detector (CAD) for the determination of phospholipid classes. Application to milk phospholipids.

    PubMed

    Kiełbowicz, Grzegorz; Micek, Piotr; Wawrzeńczyk, Czesław

    2013-02-15

    A new rapid method for the quantitative analysis of five classes of phospholipids (PLs) (phosphatidylcholine--PC, lysophosphatidylcholine--LPC, phosphatidylethanolamine--PE and phosphatidylserine--PS and phosphatidylinositol--PI) using liquid chromatography with charge aerosol detector (CAD) is described. The separation of the compounds of interest was achieved on a diol stationary phase with a mobile phase consisting of 13% HCOOH, hexane and 2-propanol in 19 min elution program, including 10 min equilibration of the column. The method was applied to characterize the phospholipid fractions of cow milk. PLs present in cow milk were separated by solid-phase extraction (SPE) procedure with Si cartridges before LC analysis with recovery ranging from 95.3% to 104.4%. The use of CAD detection of the eluted compounds was precise, linear and sensitive.

  16. Alternative method for the quantification by gas chromatography triacylglycerol class analysis of cocoa butter equivalent added to chocolate bars.

    PubMed

    Guyon, François; Destouesse, Sarah; Moustirats, Joëlle; Esclapez, Maryse; Salagoity, Marie-Hélène; Medina, Bernard

    2004-05-19

    Directive 2000/36/EC allows chocolate makers to add up to 5% of only six specific cocoa butter equivalents (CBEs) to cocoa butter (CB). A quantification method based on triacylglycerol (TAG) class analysis by gas chromatography with an unpolar column was set up for routine control purposes of chocolate bars. Mixtures of CBEs/CB were elaborated according to a Placket-Burman experiment design and analyzed by gas chromatography. A matrix was built with the normalized values of TAG classes (C50, C52, C54, and C56) of pure CBs of various origins, homemade CB/CBE mixtures (1 CB type), and mixtures containing CBE with CBs of various origins. A multivariate calibration equation was computed from this matrix using a partial least-squares regression technique. CBE addition can be detected at a minimum level of 2%, and the mathematical model allows its quantification with an uncertainty of 2% with respect to the cocoa butter fats. The model has also been applied for deconvolution and quantification of each CBE of a CBE mixture in chocolate bars.

  17. PEG precipitation coupled with chromatography is a new and sufficient method for the purification of botulinum neurotoxin type B [corrected].

    PubMed

    Zhao, Yao; Kang, Lin; Gao, Shan; Gao, Xing; Xin, Wenwen; Wang, Jinglin

    2012-01-01

    Clostridium botulinum neurotoxins are used to treat a variety of neuro-muscular disorders, as well as in cosmetology. The increased demand requires efficient methods for the production and purification of these toxins. In this study, a new purification process was developed for purifying type B neurotoxin. The kinetics of C.botulinum strain growth and neurotoxin production were determined for maximum yield of toxin. The neurotoxin was purified by polyethylene glycol (PEG) precipitation and chromatography. Based on design of full factorial experiment, 20% (w/v) PEG-6000, 4 °C, pH 5.0 and 0.3 M NaCl were optimal conditions to obtain a high recovery rate of 87% for the type B neurotoxin complex, as indicated by a purification factor of 61.5 fold. Furthermore, residual bacterial cells, impurity proteins and some nucleic acids were removed by PEG precipitation. The following purification of neurotoxin was accomplished by two chromatography techniques using Sephacryl™ S-100 and phenyl HP columns. The neurotoxin was recovered with an overall yield of 21.5% and the purification factor increased to 216.7 fold. In addition, a mouse bioassay determined the purified neurotoxin complex possessed a specific toxicity (LD(50)) of 4.095 ng/kg.

  18. Development of analytical method for catechol compounds in mouse urine using hydrophilic interaction liquid chromatography with fluorescence detection.

    PubMed

    Kanamori, Takahiro; Isokawa, Muneki; Funatsu, Takashi; Tsunoda, Makoto

    2015-03-15

    An analytical method for catecholamines and related compounds using hydrophilic interaction liquid chromatography (HILIC) with native fluorescence detection has been developed. We found that ZIC-cHILIC with phosphorylcholine was suitable for the separation of catechol compounds with good peak shapes among six different HILIC columns (Inertsil SIL, Inertsil Amide, Inertsil Diol, TSKgel NH2-100, ZIC-HILIC, and ZIC-cHILIC). Using ZIC-cHILIC, eight catechol compounds (dopamine, epinephrine, norepinephrine, 3,4-dihydroxyphenylalanine, 3,4-dihydroxyphenylacetic acid, 3,4-dihydroxyphenylglycol, 3,4-dihydroxymandelic acid, and internal standard 3,4-dihydroxybenzylamine) were separated within 15min. The limit of detection at a signal to noise ratio of 3 was 3-28nM. An improved sensitivity was obtained as compared to that of reversed-phase liquid chromatography. This was partly attributed to the increase in the fluorescence intensity of the catechol compounds in the acetonitrile-rich mobile phase. Solid phase extraction using a monolithic silica disk-packed spin column with phenylboronate moieties, which have affinity to catechol compounds, was performed for the selective extraction of catechol compounds from mouse urine. Dopamine, epinephrine, norepinephrine, 3,4-dihydroxyphenylalanine, and 3,4-dihydroxyphenylglycol were successfully quantified in mouse urine.

  19. A novel liquid chromatography method using diode-array detector for the determination of oleuropein in dietary supplements.

    PubMed

    Bertolini, Tiziana; Vicentini, Lorenza; Boschetti, Silvia; Andreatta, Paolo; Gatti, Rita

    2016-09-10

    A simple and fast chromatographic method using ultraviolet diode-array detector (UV-DAD) was developed for the automatic high performance liquid chromatography (HPLC) determination of the title of oleuropein in a new dietary supplements in form of effervescent granules. The chromatographic separations were performed on a C18 core-shell column with detection at λ=232nm. The mobile phase consisted of deionized water with 0.1% TFA and acetonitrile under gradient conditions at a flow-rate of 0.8mL/min. Oleuropein and oleuroside present in the raw material were characterized by high performance liquid chromatography-electrospray ionization tandem mass spectrometry (HPLC-ESI-MS/MS). The validation of the analytical procedure has been performed determining the following parameters: specificity, linearity, repeatability, reproducibility, accuracy, limit of quantification (LOQ), stability of the standard and sample solutions. Linear response was observed in fortified placebo solutions (determination coefficient: 0.9998). Intra-day precision (relative standard deviation, RSD) was ≤5.0% for peak area and for retention times (tR) without significant differences between intra- and inter-day data. The limits of quantitation (LOQ) was about 5μg/mL and 9pmol/inject. Oleuropein recovery studies gave good results (99.9%) with a R.S.D. of 0.5%. The speed of analysis and the stability of the solutions with a fluctuation Δ (%) ≤2.0 at room temperature means an undoubted advantage of the method allowing the simultaneous preparation of many samples and consecutive chromatographic analyses by using an autosampler. The developed method is suitable for the quality control of oleuropein in raw material and industrial products. The method can be applied in any analytical laboratory not requiring a sophisticated instrumentation.

  20. Determination of perchlorate in drinking water by ion chromatography using macrocycle-based concentration and separation methods.

    PubMed

    Lamb, John D; Simpson, David; Jensen, Bryce D; Gardner, Joseph S; Peterson, Quinn P

    2006-06-16

    Macrocycle-based ion chromatography provides a convenient, reliable method for the determination of perchlorate ion, which is currently of great interest to the environmental community. This study shows that effective perchlorate determinations can be made using standard conductimetric detection by combining an 18-crown-6-based mobile phase with an underivatized reversed-phase mobile phase ion chromatography (MPIC) column. One unique feature of this method is the flexibility in column capacity that is achieved through simple variations in eluent concentrations of 18-crown-6 and KOH, facilitating the separation of target analyte anions such as perchlorate. Using a standard anion exchange column as concentrator makes possible the determination of perchlorate as low as 0.2 ug/L in low ionic strength matrices. Determination of perchlorate at the sub-ug/L level in pure water and in spiked local city hard water samples with high background ion concentrations can be achieved this way. However, like other IC techniques, this method is challenged to achieve analyses at the ug/L level in the demanding high ionic strength matrix described by the United States Environmental Protection Agency (EPA) (1,000 mg/L chloride, sulfate and carbonate). We approached this challenge by use of the Cryptand C1 concentrator column, provided by Dionex Corporation, to effectively preconcentrate perchlorate while reducing background ion concentrations in the high ionic strength matrix. The retention characteristics of the concentrator column were studied in order to maximize its effectiveness for perchlorate determinations. The method makes possible the determination of perchlorate at the 5 ug/L level in the highest ionic strength matrix described by the EPA.

  1. From analytical methods to large scale chiral supercritical fluid chromatography using chlorinated chiral stationary phases.

    PubMed

    Wu, Dauh-Rurng; Yip, Shiuhang Henry; Li, Peng; Sun, Dawn; Mathur, Arvind

    2016-02-05

    While traditional non-chlorinated Cellulose- and Amylose-derivatized phases have been used successfully in supercritical fluid chromatography (SFC) to resolve a broad variety of chiral compounds, some chiral pharmaceutical compounds are not well resolved on these traditional chiral stationary phases (CSP) due to the lack of chiral selectivity. Since there are no universal CSP to resolve all chiral compounds, chlorinated CSP can be complementary to the non-chlorinated CSP. Chlorinated CSP such as 4-Chloro-3-methylphenyl-carbamatecellulose (Lux-Cellulose-4), 3-Chloro-4-methylphenyl-carbamatecellulose (Lux-Cellulose-2), 5-Chloro-2-methylphenyl-carbamateamylose (Lux-Amylose-2) and immobilized 3,5-dichlorophenyl-carbamatecellulose (Chiralpak IC) have provided a range of chiral recognition mechanisms which have allowed the authors to successfully achieve chiral SFC resolution on several structurally diverse compounds, which are not well resolved in the non-chlorinated CSP. In addition, chlorinated Lux-Cellulose-4, Chiralpak IC and Lux-Amylose-2 have enabled us to utilize non-alcohol solvents as sample diluents and as co-solvents to significantly improve compound solubility and selectivity. This article will discuss the challenges associated with several SFC applications on both coated and immobilized chlorinated CSP to deliver high-quality drug candidates in large quantity. The use of dichloromethane in both sample preparation and as co-solvent in CO2 to increase sample solubility will be presented in preparative example #2 and #3.

  2. A novel method for rapid determination of total solid content in viscous liquids by multiple headspace extraction gas chromatography.

    PubMed

    Xin, Li-Ping; Chai, Xin-Sheng; Hu, Hui-Chao; Barnes, Donald G

    2014-09-05

    This work demonstrates a novel method for rapid determination of total solid content in viscous liquid (polymer-enriched) samples. The method is based multiple headspace extraction gas chromatography (MHE-GC) on a headspace vial at a temperature above boiling point of water. Thus, the trend of water loss from the tested liquid due to evaporation can be followed. With the limited MHE-GC testing (e.g., 5 extractions) and a one-point calibration procedure (i.e., recording the weight difference before and after analysis), the total amount of water in the sample can be determined, from which the total solid contents in the liquid can be calculated. A number of black liquors were analyzed by the new method which yielded results that closely matched those of the reference method; i.e., the results of these two methods differed by no more than 2.3%. Compared with the reference method, the MHE-GC method is much simpler and more practical. Therefore, it is suitable for the rapid determination of the solid content in many polymer-containing liquid samples.

  3. Development of thermal desorption gas chromatography/mass spectrometry as a rapid method for ambient particulate characterization

    NASA Astrophysics Data System (ADS)

    Sheya, Sue Anne N.

    A direct thermal desorption gas chromatography/mass spectrometry (TD GC/MS) method for air particulate matter (PM) analysis of volatile and semivolatile organic compounds was investigated. This technique uses a specially designed microdesorption GC inlet utilizing an inductively heated ferromagnetic foil with a Curie point temperature suitable for desorption, which can accommodate microgram amounts of material deposited on a thin strip of quartz fiber filter. Liquid or solid samples can be rapidly desorbed within 10 s at 315°C, followed by 30--40 min of chromatography time. The results obtained by this technique were found to be statistically equivalent to those obtained by the conventional solvent extraction gas chromatography/mass spectrometry (SX GC/MS) method for analysis of aromatic and n alkane standards, single source soot particles, and PM 10 filter samples. Correlations between injecting an extract, desorbing an extract, and desorbing particles averaged R = 0.94, with a three way correlation averaging R = 0.97. High volume sampling conducted at 12 spatially distributed sites located along the US/Mexican border of the El Paso/Juarez metroplex supplied 24h PM 10 filters for an investigation combining thermal desorption with a rapid online chemical derivatization procedure, and multivariate methods of source attribution using principal component and canonical correlation analysis of the resultant chemical markers. Four major combustion related PM emission sources were revealed at these sites: automotive, waste burning, biomass burning and meat cooking. A second investigation conducted in the same area used mediumvolume sampling to collect 2 h timeresolved PM 10 receptor samples for TD GC/MS analysis. Additionally, 2 h samples for inorganic analysis, multichannel particle size distribution measurements, and meteorological data were collected enabling generation of circadian PM multicharacterization profiles. Factor analysis based receptor modeling using

  4. Development and validation of a gas chromatography-flame ionization detection method for quantifying sucrose in equine serum.

    PubMed

    Hewetson, Michael; Aaltonen, Kaisa; Tulamo, Riitta-Mari; Sankari, Satu

    2014-03-01

    A simple and accurate method for quantifying sucrose in equine serum that can be applied to sucrose permeability testing in the horse was developed and validated using gas chromatography with flame ionization detection. The assay provided an acceptable degree of linearity, accuracy, and precision at concentrations of sucrose as low as 2.34 μmol/l and as high as 20.45 μmol/l. Percentage recovery of sucrose from serum ranged from 89% to 102%; repeatability and intermediate precision (relative standard deviation) ranged from 3.6% to 6.7% and 4.1% to 9.3%, respectively. The limit of detection was 0.73 μmol/l. No interfering peaks were observed except lactose, which gave 2 peaks, one of which overlapped partially with sucrose. To evaluate the suitability of the method for quantifying sucrose in serum samples from horses with naturally occurring gastric ulceration, 10 horses with and without naturally occurring gastric ulceration were subjected to sucrose permeability testing. All horses demonstrated an increase in serum sucrose concentration over time following oral administration of sucrose; however, the increase from baseline was significant for horses with gastric ulceration at 45 min (P = 0.0082) and 90 min (P = 0.0082) when compared with healthy horses. It was concluded that gas chromatography with flame ionization detection is a valid method for quantifying sucrose in equine serum and can be applied directly to the analysis of sucrose in equine serum as part of a larger validation study aimed at developing a blood test for the diagnosis of gastric ulcers in horses.

  5. Determination of soluble immunoglobulin G in bovine colostrum products by Protein G affinity chromatography-turbidity correction and method validation.

    PubMed

    Holland, Patrick T; Cargill, Anne; Selwood, Andrew I; Arnold, Kate; Krammer, Jacqueline L; Pearce, Kevin N

    2011-05-25

    Immunoglobulin-containing food products and nutraceuticals such as bovine colostrum are of interest to consumers as they may provide health benefits. Commercial scale colostrum products are valued for their immunoglobulin G (IgG) content and therefore require accurate analysis. One of the most commonly used methods for determining total soluble IgG in colostrum products is based on affinity chromatography using a Protein G column and UV detection. This paper documents improvements to the accuracy of the Protein G analysis of IgG in colostrum products, especially those containing aggregated forms of IgG. Capillary electrophoresis-sodium dodecyl sulfate (CE-SDS) analysis confirmed that aggregated IgG measured by Protein G does not contain significant amounts of casein or other milk proteins. Size exclusion chromatography identified the content of soluble IgG as mainly monomeric IgG and aggregated material MW > 450 kDa with small amounts of dimer and trimer. The turbidity of the eluting IgG, mainly associated with aggregated IgG, had a significant effect on the quantitative results. Practical techniques were developed to correct affinity LC data for turbidity on an accurate, consistent, and efficient basis. The method was validated in two laboratories using a variety of colostrum powders. Precision for IgG was 2-3% (RSD(r)) and 3-12% (RSD(R)). Recovery was 100.2 ± 2.4% (mean ± RSD, n = 10). Greater amounts of aggregated IgG were solubilized by a higher solution:sample ratio and extended times of mixing or sonication, especially for freeze-dried material. It is concluded that the method without acid precipitation and with turbidity correction provides accurate, precise, and robust data for total soluble IgG and is suitable for product specification and quality control of colostrum products.

  6. Development and validation of a liquid chromatography-tandem mass spectrometry method to determine intact glucosinolates in bee pollen.

    PubMed

    Ares, Ana M; Nozal, María J; Bernal, José

    2015-09-01

    A new method was developed to determine twelve intact-glucosinolates (GLSs) (glucoiberin, GIB; glucoraphanin, GRA; glucoerucin GER; gluconapin, GNA; glucotropaeolin, GTL; glucobrassicin, GBC; gluconasturtiin, NAS; glucoalyssin, ALY; 4-hydroxyglucobrassicin, 4OH; 4-methoxyglucobrassicin, 4ME; neoglucobrassicin, NEO; sinigrin, SIN) in bee pollen, by means of liquid chromatography tandem mass spectrometry (LC-MS/MS) with electrospray ionization (ESI). An efficient extraction procedure was proposed (average analyte recoveries were between 85% and 96%); this involved a solid-liquid extraction (SLE) with heated water, followed by a solid phase extraction (SPE) with a weak anion exchange (NH2) sorbent. Chromatography was performed on a Gemini(®) C18 analytical column with a mobile phase of formic acid in water (0.5%,v/v) and formic acid in acetonitrile (0.5%,v/v), in gradient elution mode at 1mL/min, resulted in baseline-separated peaks and a run time of 30min. The method was fully validated in terms of selectivity, limits of detection (LOD) and quantification (LOQ), linearity, carry-over effect, reinjection reproducibility, precision and accuracy. A good selectivity, low LODs and LOQs, ranging from 1 to 16μg/kg, wide linear ranges from LOQ to 1000μg/kg, and satisfactory reinjection reproducibility, precision and accuracy with relative standard deviation and relative error values lower than or equal to 9%; meanwhile, results indicates a negligible carry-over effect. The proposed method was applied to analyze intact-GLSs in bee pollen. Nine of the GLSs studied were identified in certain samples analyzed over a wide concentration range (LOQ-2226μg/kg), and significant differences in GLS content were observed among the samples.

  7. Sensitive method for detection of cocaine and associated analytes by liquid chromatography-tandem mass spectrometry in urine.

    PubMed

    Langman, Loralie J; Bjergum, Matthew W; Williamson, Christopher L; Crow, Frank W

    2009-10-01

    Cocaine (COC) is a potent CNS stimulant that is metabolized to benzoylecgonine (BE) and further metabolized to minor metabolites such as m-hydroxybenzoylecgonine (m-HOBE). COC is also metabolized to norcocaine (NC). Cocaethylene (CE) is formed when cocaine and ethyl alcohol are used simultaneously. Anhydroecgonine methyl ester (AEME) is a unique marker following smoked cocaine, and anhydroecgonine ethyl ester (AEEE) is found in cocaine smokers who also use ethyl alcohol. We developed a liquid chromatography-tandem mass spectrometry (LC-MS-MS) method for the detection and quantitation of COC, BE, NC, CE, m-HOBE, AEME, and AEEE in urine. Two hundred samples previously analyzed by gas chromatography (GC) coupled with MS were extracted using solid-phase extraction. Chromatographic separation was achieved using a gradient consisting of mobile phase A [20 mM ammonium formate (pH 2.7)] and mobile phase B (methanol/acetonitrile, 50:50), an XDB-C(8) (50 x 2.1 mm, 1.8 microm) column and a flow rate of 270 microL/min. Concentrations were calculated by comparing the peak-area with the internal standard and plotted against a standard curve. The assay displayed linearity from 1.0 to 100 ng/mL. Within- and between-run coefficients of variation were < 10% throughout the linear range. A method comparison between GC-MS and LC-MS-MS showed good correlation for COC (r(2) = 0.982) and BE (r(2) = 0.955). We report here on a sensitive method to identify clinically and forensically relevant cocaine and associated analytes at concentrations as low as 1.0 ng/mL.

  8. Method development and validation study for quantitative determination of nifedipine and related substances by ultra-high-performance liquid chromatography.

    PubMed

    Galan-Rodriguez, Cristobal; González-Álvarez, Jaime; Valls-Remolí, Màrius

    2015-02-01

    A novel stability-indicating reversed phase ultra-high performance liquid chromatography (UPLC) coupled photodiode array gradient method was developed for determination of the nifedipine and related compounds. Furthermore, based on the chromatographic conditions and forced degradation studies performed through the development of the related substances method a UPLC isocratic method was validated for the determination of the assay of this active substance. An Acquity Shield RP18 (50 × 3.0 mm 1.7 µm) column was used for separation of nifedipine and its five potential impurities within 11 min, which is 5-fold less than the official method. A mobile phase consisting of 10 mm ammonium formate (pH 4.5) and methanol, delivered at a flow rate 0.5 mL/min, was employed to achieve a minimum resolution of 2.0 for all consecutive pairs of compounds. The precision value expressed as percentage relative standard deviation for method repeatability and reproducibility was <5.0%. The recoveries for all the related compounds were in the range of 99-105.0%. Linearity was found to be acceptable over the concentration range of 0.25-1.5 µg/mL for nifedipine and its impurities. The limit of quantification for nifedipine was 0.05 µg/mL, which is much less than the European Pharmacopoeia method.

  9. Predicting gas chromatography relative retention times for polychlorinated biphenyls using chlorine substitution pattern contribution method.

    PubMed

    Li, An; Gao, Jie; Freels, Sally; Huang, Jun; Yu, Gang

    2016-01-04

    Various quantitative structure retention relationships have been published in an effort to understand and predict chromatographic retention times. This work presents a chlorine substitution pattern contribution (Cl-SPC) model for relative retention times (RRT) of polychlorinated biphenyls (PCBs), using 27 sets of previously published gas chromatography RRT data. The Cl-SPC model calculates the contribution factors (βk) for each of 19 chlorine substitution "patterns" (such as 2-, 2,4-, 2,3,6-, 2,3,4,5,6-, etc.) using multiple linear regression (MLR). The 27 separate MLRs had R(2) values ranging from 0.961 to 1.000; the average absolute errors were 0.55% for the training sets and 0.95% for the test sets. Cross-validation of the model was carried out by splitting each data set into training and test sets for groupings based on nine PCB congener mixes commercialized by AccuStandard. No weakening of the model performance was observed when the size of data set used to develop the model was decreased from 209 to 39 congeners. In addition to the separate models, a single mixed model was fit combining all 27 data sets. The estimated random effects, which reflect the impact of GC configuration and operational conditions on RRTs, are minor compared with the fixed effects estimated for the βk values. The major advantages of the Cl-SPC model are its unmatched simplicity and equally excellent robustness when compared with other quantitative structure retention relationship models.

  10. Atmospheric-pressure laser ionization: a novel ionization method for liquid chromatography/mass spectrometry.

    PubMed

    Constapel, M; Schellenträger, M; Schmitz, O J; Gäb, S; Brockmann, K J; Giese, R; Benter, Th

    2005-01-01

    We report on the development of a new laser-ionization (LI) source operating at atmospheric pressure (AP) for liquid chromatography/mass spectrometry (LC/MS) applications. APLI is introduced as a powerful addition to existing AP ionization techniques, in particular atmospheric-pressure chemical ionization (APCI), electrospray ionization (ESI), and atmospheric pressure photoionization (APPI). Replacing the one-step VUV approach in APPI with step-wise two-photon ionization strongly enhances the selectivity of the ionization process. Furthermore, the photon flux during an ionization event is drastically increased over that of APPI, leading to very low detection limits. In addition, the APLI mechanism generally operates primarily directly on the analyte. This allows for very efficient ionization even of non-polar compounds such as polycyclic aromatic hydrocarbons (PAHs). The APLI source was characterized with a MicroMass Q-Tof Ultima II analyzer. Both the effluent of an HPLC column containing a number of PAHs (benzo[a]pyrene, fluoranthene, anthracene, fluorene) and samples from direct syringe injection were analyzed with respect to selectivity and sensitivity of the overall system. The liquid phase was vaporized by a conventional APCI inlet (AP probe) with the corona needle removed. Ionization was performed through selective resonance-enhanced multi-photon ionization schemes using a high-repetition-rate fixed-frequency excimer laser operating at 248 nm. Detection limits well within the low-fmol regime are readily obtained for various aromatic hydrocarbons that exhibit long-lived electronic states at the energy level of the first photon. Only molecular ions are generated at the low laser fluxes employed ( approximately 1 MW/cm(2)). The design and performance of the laser-ionization source are presented along with results of the analysis of aromatic hydrocarbons.

  11. Chromatography resin support

    DOEpatents

    Dobos, James G.

    2002-01-01

    An apparatus and method of using an improved chromatography resin support is disclosed. The chromatography support platform is provided by a stainless steel hollow cylinder adapted for being inserted into a chromatography column. An exterior wall of the stainless steel cylinder defines a groove for carrying therein an "O"-ring. The upper surface of the stainless steel column is covered by a fine stainless steel mesh welded to the edges of the stainless steel cylinder. When placed upon a receiving ledge defined within a chromatography column, the "O"-ring provides a fluid tight seal with the inner edge wall of the chromatography cylinder. The stainless steel mesh supports the chromatography matrix and provides a back flushable support which is economical and simple to construct.

  12. Separation techniques: Chromatography

    PubMed Central

    Coskun, Ozlem

    2016-01-01

    Chromatography is an important biophysical technique that enables the separation, identification, and purification of the components of a mixture for qualitative and quantitative analysis. Proteins can be purified based on characteristics such as size and shape, total charge, hydrophobic groups present on the surface, and binding capacity with the stationary phase. Four separation techniques based on molecular characteristics and interaction type use mechanisms of ion exchange, surface adsorption, partition, and size exclusion. Other chromatography techniques are based on the stationary bed, including column, thin layer, and paper chromatography. Column chromatography is one of the most common methods of protein purification. PMID:28058406

  13. New background correction method for liquid chromatography with diode array detection, infrared spectroscopic detection and Raman spectroscopic detection.

    PubMed

    Boelens, Hans F M; Dijkstra, Reyer J; Eilers, Paul H C; Fitzpatrick, Fiona; Westerhuis, Johan A

    2004-11-19

    A new method to eliminate the background spectrum (EBS) during analyte elution in column liquid chromatography (LC) coupled to spectroscopic techniques is proposed. This method takes into account the shape and also intensity differences of the background eluent spectrum. This allows the EBS method to make a better estimation of the background eluent spectrum during analyte elution. This is an advantage for quantification as well as for identification of analytes. The EBS method uses a two-step procedure. First, the baseline spectra are modeled using a limited number of principal components (PCs). Subsequently, an asymmetric least squares (asLS) regression method is applied using these principal components to correct the measured spectra during elution for the background contribution. The asymmetric least squares regression needs one parameter, the asymmetry factor p. This asymmetry factor determines relative weight of positive and negative residuals. Simulations are performed to test the EBS method in well-defined situations. The effect of spectral noise on the performance and the sensitivity of the EBS method for the value of the asymmetry factorp is tested. Two applications of the EBS method are discussed. In the first application, the goal is to extract the analyte spectrum from an LC-Raman analysis. In this case, the EBS method facilitates easy identification of unknown analytes using spectral libraries. In a second application, the EBS method is used for baseline correction in LC-diode array detection (DAD) analysis of polymeric standards during a gradient elution separation. It is shown that the EBS method yields a good baseline correction, without the need to perform a blank chromatographic run.

  14. Ambient formic acid in southern California air: A comparison of two methods, Fourier transform infrared spectroscopy and alkaline trap-liquid chromatography with UV detection

    SciTech Connect

    Grosjean, D. ); Tuazon, E.C. ); Fujita, E. )

    1990-01-01

    Formic acid is an ubiquitous component of urban smog. Sources of formic acid in urban air include direct emissions from vehicles and in situ reaction of ozone with olefins. Ambient levels of formic acid in southern California air were first measured some 15 years ago by Hanst et al. using long-path Fourier transform infrared spectroscopy (FTIR). All subsequent studies of formic acid in the Los Angeles area have involved the use of two methods, either FTIR or collection on alkaline traps followed by gas chromatography, ion chromatography, or liquid chromatography analysis with UV detection, ATLC-UV. The Carbon Species Methods Comparison Study (CSMCS), a multilaboratory air quality study carried out in August 1986 at a southern California smog receptor site, provided an opportunity for direct field comparison of the FTIR and alkaline trap methods. The results of the comparison are presented in this brief report.

  15. Suppressed ion chromatography methods for the routine determination of ultra low level anions and cations in ice cores.

    PubMed

    Curran, M A; Palmer, A S

    2001-06-01

    The concentration of trace ionic species in snow and ice samples was determined using suppressed ion chromatography (IC) with conductivity detection and ultra-clean sample preparation techniques. Trace anion species were determined in a single 24-min run by combining sample preconcentration with gradient elution using Na2B4O7 eluent. The detection limits (ranging from 0.001 to 0.006 microM) are the lowest reported in the literature. Cation species were analysed by direct injection of 0.25 ml and isocratic elution with a H2SO4 eluent. The clean preparation techniques showed no evidence of a difference (Student's t-test) between Milli-Q water samples analysed directly and processed Milli-Q ice samples. These robust, ultra-clean IC methods were routinely applied to the analysis of large number of samples to produce a high-resolution trace ion ice core record from Law Dome, East Antarctica.

  16. Development of a size exclusion chromatography-electrochemical detection method for the analysis of total organic and inorganic chloramines.

    PubMed

    Amiri, Fariba; Andrews, Susan

    2008-08-01

    A size exclusion chromatography (SEC) followed by post-column reaction with iodide and electrochemical detection method is developed for analysis of total organic and inorganic chloramines. Ammonium chloride and a group of test compounds (Glycine, Tyrosine, DL-Alanyl-DL-Alanine, Arg-Gly-Asp-Ser, Bradykinin, Aprotinin, and alpha-Lactalbumin) are selected and chlorinated to represent inorganic chloramines and different sizes of organic N-chloramines. An analytical SEC column with pore size of 60A is used and chromatographic conditions including the working electrode potential and flow rate are optimized to gain optimum resolution and sensitivity. The detection limits are estimated to be 0.12 mg/L and 0.05 mg/L, respectively, for tested inorganic and organic chloramines.

  17. Development of a high sensitivity bioanalytical method for alprazolam using ultra-performance liquid chromatography/tandem mass spectrometry.

    PubMed

    Mather, Joanne; Rainville, Paul D; Potts, Warren B; Smith, Norman W; Plumb, Robert S

    2010-01-01

    A rapid, specific, assay was developed for the benzodiapine alprazolam in rat plasma using sub-2 µm particle liquid chromatography (LC) and tandem quadrupole mass spectrometry (MS/MS). The limit of quantification using protein precipitation was determined to 10 pg/mL, whereas the limit of quantification using solid-phase extraction (SPE) was determined to be 1.0 pg/mL. The assay was optimized for throughput and resolution of the analyte of interest from the hydroxy metabolite. During the method development process the plasma matrix signal was monitored, for lipids and other endogenous metabolites, to maximize signal response and minimize ion suppression. This was achieved by using a tandem quadrupole mass spectrometer equipped with a novel collision cell design which allowed for the simultaneous collection of full scan MS and multiple reaction monitoring (MRM) data. The lipid profile from the SPE process was significantly less than obtained with the protein precipitation approach.

  18. Evaluation of low-pressure gas chromatography-tandem mass spectrometry method for analysis of greater than 140 pesticides in fish

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A multi-residue method for analysis of 143 pesticide residues in fish was developed and evaluated using fast, low pressure gas chromatography triple quadrupole tandem mass spectrometry (LP-GC/MS-MS). The method was based on a QuEChERS (quick, easy, cheap, effective, rugged, safe) extraction with ace...

  19. METHOD 544. DETERMINATION OF MICROCYSTINS AND NODULARIN IN DRINKING WATER BY SOLID PHASE EXTRACTION AND LIQUID CHROMATOGRAPHY/TANDEM MASS SPECTROMETRY (LC/MS/MS)

    EPA Science Inventory

    Method 544 is an accurate and precise analytical method to determine six microcystins (including MC-LR) and nodularin in drinking water using solid phase extraction and liquid chromatography tandem mass spectrometry (SPE-LC/MS/MS). The advantage of this SPE-LC/MS/MS is its sensi...

  20. Packing of large-scale chromatography columns with irregularly shaped glass based resins using a stop-flow method

    PubMed Central

    Siu, Sun Chau; Chia, Celeste; Mok, Yanglin; Pattnaik, Priyabrata

    2014-01-01

    Rigid chromatography resins, such as controlled pore glass based adsorbents, offer the advantage of high permeability and a linear pressure-flow relationship irrespective of column diameter which improves process time and maximizes productivity. However, the rigidity and irregularly shaped nature of these resins often present challenges in achieving consistent and uniform packed beds as formation of bridges between resin particles can hinder bed consolidation. The standard flow-pack method when applied to irregularly shaped particles does not yield well-consolidated packed beds, resulting in formation of a head space and increased band broadening during operation. Vibration packing methods requiring the use of pneumatically driven vibrators are recommended to achieve full packed bed consolidation but limitations in manufacturing facilities and equipment may prevent the implementation of such devices. The stop-flow packing method was developed as an improvement over the flow-pack method to overcome these limitations and to improve bed consolidation without the use of vibrating devices. Transition analysis of large-scale columns packed using the stop-flow method over multiple cycles has shown a two- to three-fold reduction of change in bed integrity values as compared to a flow-packed bed demonstrating an improvement in packed bed stability in terms of the height equivalent to a theoretical plate (HETP) and peak asymmetry (As). PMID:25080096

  1. A validated micellar electrokinetic chromatography method for the quantitation of dexamethasone, ondansetron and aprepitant, antiemetic drugs, in organogel.

    PubMed

    Bourdon, Florence; Lecoeur, Marie; Duhaut, Marion; Odou, Pascal; Vaccher, Claude; Foulon, Catherine

    2013-12-01

    A micellar electrokinetic chromatography (MEKC) method was developed for the determination of three anti-vomiting drugs (aprepitant, dexamethasone and ondansetron) in pharmaceutical formulations. The method was optimized using a central composite design (CCD). Four main factors (borate buffer concentration, pH, methanol content and sodium dodecyl sulfate concentration) were optimized in order to obtain best resolutions and peak efficiencies in a minimum runtime. The separation was performed in a fused-silica capillary. After optimization, the background electrolyte consisted of a borate buffer (62.5mM, pH 8.75) containing sodium dodecyl sulfate (77.5mM) and methanol (3.75%). Under these conditions, a complete separation of each antiemetic drug and its respective internal standards was achieved in 38min. The method was validated with trueness values from 94.9 to 107.2% and precision results (repeatability and intermediate precision) lower than 5.9%. MEKC-UV was the first method allowing the separation of aprepitant, dexamethasone and ondansetron and was suitable for the quantitation of these three antiemetic drugs in organogel formulations. The rapid sample preparation coupled with an automated separation technique make this method convenient for quality control of extemporaneous magistral ready-to-use formulation.

  2. A stability-indicating reversed-phase high-performance liquid chromatography method for Ambrisentan: an endothelin receptor antagonist.

    PubMed

    Satheeshkumar, Nanjappan; Naveenkumar, Gandham

    2014-09-01

    A stability-indicating reversed-phase high-performance liquid chromatography (HPLC) method was developed for the determination of ambrisentan, a drug used in the treatment of pulmonary hypertension. The desired chromatographic separation was achieved on a Kromasil C18 column (250 × 4.6 mm, 5 µm) using the mobile phase acetonitrile-ammonium formate (pH 3.0; 0.02 M) in gradient mode. The flow rate was set at 1.0 mL/min, and chromatograms were extracted at 262 nm using a photodiode array detector. The method was successfully validated in accordance to International Conference on Harmonization (ICH) guidelines acceptance criteria for linearity, accuracy, precision, robustness and forced degradation studies, which further proved the stability-indicating power. Linearity of ambrisentan peak area responses was demonstrated within the concentration range of 25-200 µg/mL. The limits of detection and quantitation were 0.2 and 0.6 µg/mL, respectively. Forced degradation studies were performed on ambrisentan bulk drug samples as per ICH guidelines to demonstrate the stability-indicating power of the HPLC method. Significant degradation was observed during acidic hydrolysis, neutral hydrolysis and oxidative stress. The degradation products were well resolved from primary peak of ambrisentan, indicating that the method is specific and stability indicating. The newly developed method is applicable for assay determination of active pharmaceutical ingredient.

  3. A novel ultra performance liquid chromatography-tandem mass spectrometry method for the determination of sucrose octasulfate in dog plasma.

    PubMed

    Ke, Yuyong; Li, Steve Lianghong; Chang, Linda Dongxia; Kapanadze, Theo

    2015-01-26

    A novel, specific and sensitive bioanalytical method has been developed for the determination of sucrose octasulfate (SOS) in dog plasma and urine using ion-pair reversed-phase ultraperformance liquid chromatography coupled with electrospray triple quadruple mass spectrometry (IPRP-UPLC ESI MS/MS). (13)C-labeled sucrose octasulfate-(13)C12 sodium salt is used as the internal standard. 200 μL of plasma or serum sample is extracted using weak anion exchange solid phase cartridge. In this method, a polar amide column is employed for the liquid chromatograph (LC) separation while the diethylamine and formic acid buffer is used as the ion-pairing reagent. The low limitation of quantitation of sucrose octasulfate is 0.20 ng on the column with a signal to noise ratio larger than 50. Parameters such as linearity, accuracy and precision have been validated in full compliance with the FDA guidelines for the bioanalytical method development and validation. A linear regression model fit the calibration curve very well with R>0.99. The bias and coefficient of variation of all levels of QCs are within the range of 15%. The selectivity, matrix effect and stabilities of analytes in solution and matrix have also been evaluated and the results met the acceptance criteria according to the guidelines. Based on these results, the method has qualified to analyze sucrose octasulfate in dog plasma for clinic research. This method has been applied to 1000 preclinical samples.

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

    PubMed

    Bartolomé, Mónica; Gallego-Picó, Alejandrina; Huetos, Olga; Castaño, Argelia

    2014-06-01

    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.

  5. Sensitive and robust method for anabolic agents in human urine by gas chromatography-triple quadrupole mass spectrometry.

    PubMed

    Delgadillo, Miguel A; Garrostas, Lorena; Pozo, Oscar J; Ventura, Rosa; Velasco, Benjamín; Segura, Jordi; Marcos, Josep

    2012-05-15

    A rapid, sensitive and robust gas chromatography-triple quadrupole mass spectrometry method was developed for the determination of seven anabolic agents in human urine. The selection of analytes includes the main metabolites of all anabolics with higher sensitivity requirements. After optimizing the fragmentation conditions for each compound, a validation procedure for qualitative analysis was performed. The selectivity of the method showed that no interfering peaks were observed at the retention time of the compound. Adequate intermediate precision, below 14%, was observed for all of the compounds at the lower concentration tested. The concentrations assayed were in accordance with the performance limits required by the World Anti-Doping Agency (WADA). Unlike a previously published GC/QqQ method, detection of 17α-methyl-5β-androstane-3α,17β-diol (the main metabolites of methyltestosterone) at 2 ng/mL was accomplished under routine conditions. The qualitative method was applied to the analysis of 1367 samples in the span of 2 weeks, as part of the doping control of the XVI Pan American Games which took place in Mexico (14th-30th October, 2011). The high sensitivity was maintained during the analysis of all analytical batches, proving for the first time the excellent ruggedness of GC/QqQ methods.

  6. Improvements and application of a modified gas chromatography atomic fluorescence spectroscopy method for routine determination of methylmercury in biota samples.

    PubMed

    Gorecki, Jerzy; Díez, Sergi; Macherzynski, Mariusz; Kalisinska, Elżbieta; Golas, Janusz

    2013-10-15

    Improvements to the application of a combined solid-phase microextraction followed by gas chromatography coupled to pyrolysis and atomic fluorescence spectrometry method (SPME-GC-AFS) for methylmercury (MeHg) determination in biota samples are presented. Our new method includes improvements in the methodology of determination and the quantification technique. A shaker instead of a stirrer was used, in order to reduce the possibility of sample contamination and to simplify cleaning procedures. Then, optimal rotation frequency and shaking time were settled at 800 rpm and 10 min, respectively. Moreover, the GC-AFS system was equipped with a valve and an argon heater to eliminate the effect of the decrease in analytical signal caused by the moisture released from SPME fiber. For its determination, MeHg was first extracted from biota samples with a 25% KOH solution (3h) and then it was quantified by two methods, a conventional double standard addition method (AC) and a modified matrix-matched calibration (MQ) which is two times faster than the AC method. Both procedures were successfully tested with certified reference materials, and applied for the first time to the determination of MeHg in muscle samples of goosander (Mergus merganser) and liver samples of white-tailed eagle (Haliaeetus albicilla) with values ranging from 1.19 to 3.84 mg/kg dry weight (dw), and from 0.69 to 6.23 mg kg(-1) dw, respectively.

  7. Determination of methamphetamine enantiomer composition in human hair by non-chiral liquid chromatography-tandem mass spectrometry method.

    PubMed

    Shu, Irene; Alexander, Amy; Jones, Mary; Jones, Joseph; Negrusz, Adam

    2016-08-15

    Chiral separation is crucial for investigating methamphetamine positive cases. While (S)-(+)-enantiomer of methamphetamine (S-MAMP) is a schedule II controlled substance, (R)-(-)-enantiomer (R-MAMP) is an active ingredient of a few over-the-counter drugs in the United States. Among biological specimen types, hair provides greater detection window than blood, urine or oral fluid, and are therefore regarded with particular interest. Herein we describe a novel non-chiral liquid chromatography-tandem mass spectrometry (LC-MS/MS) method to directly determine methamphetamine enantiomeric composition (percentage) in hair specimens. Hair samples were washed once with acetone, powdered, incubated overnight at 53°C in 0.1M hydrochloric acid (HCl), and subjected to a solid phase extraction (SPE). The extracts were derivatized using Marfey's reagent at 53°C for 60min. The final mixture was analyzed by LC-MS/MS. Chromatographic separation was achieved using a C18 Kinetex analytical column and 60% (v/v) aqueous methanol as mobile phase (isocratic). Triple quadrupole mass spectrometer was equipped with an electro-spray ionization (ESI) source operating in negative mode and the chromatograms were acquired using a multiple-reaction monitoring (MRM) approach. The results were expressed as ratio of R- to S-MAMP and then derived to composition percentages without requiring quantitating each enantiomer. The method was precise and accurate across 0-100% S-composition at a range of 80-18,000pg/mg. The performance of the new method was compared with an (S)-(-)-N-trifluoroacetylprolyl chloride (S-TPC) derivatization and gas chromatography-mass spectrometry (GC-MS) method on authentic methamphetamine-positive hair samples. Not only the new Marfey's reagent approach presented satisfactory correlation with the S-TPC approach, but it also exhibited significantly improved quality (e.g., S/N) of the chromatograms. In summary, our protocol employs cost effective and minimally hazardous Marfey

  8. A novel reverse phase high-performance liquid chromatography method for standardization of Orthosiphon stamineus leaf extracts

    PubMed Central

    Saidan, Noor Hafizoh; Aisha, Abdalrahim F.A.; Hamil, Mohd Shahrul Ridzuan; Majid, Amin Malik Shah Abdul; Ismail, Zhari

    2015-01-01

    Background: Orthosiphon stamineus Benth. (Lamiaceae) is a traditional medicinal plant which has been used in treating various ailments such as kidney diseases, bladder inflammation, arthritis and diabetes. The leaves contain high concentration of phenolic compounds, thus, rosmarinic acid (RA), 3’-hydroxy-5, 6, 7, 4’-tetramethoxyflavone (TMF), sinensetin (SIN) and eupatorin (EUP) were chosen as a marker compounds for standardization of various O. stamineus leaf extracts. Objective: The aim was to develop and validate a new high-performance liquid chromatography (HPLC) method for quantification of 4 marker compounds (RA, TMF, SIN, EUP) in various O. stamineus leaf extracts. Materials and Methods: The method was developed and validated using RP-HPLC-diode-array detection at 320 nm for accuracy, precision and limits of detection and was applied for quantification of it markers in five different extracts prepared in solvents with increasing polarity, using a gradient mobile phase 0.1% formic acid: Acetonitrile at a flow rate of 1 ml/min on reverse phase acclaim polar advantage II C18 column (3 μm, 3 × 150 mm) with 18 min separation time. Results: The developed method provided satisfactory precision, and the accuracy of this method was in the range of 90.2% to 105.5%. All of 4 compounds showed good linearity at R2 > 0.999. Conclusion: The developed method is a simple, cost effective with shorter run time (18 min) in comparison to previous methods (30 min) and utilization of environmental-friendly solvents system. Therefore, this method has the potential to replace currently used methods in the routine standardization work of O. stamineus extracts, raw materials and its commercial products. PMID:25598631

  9. Method development for the characterization of biofuel intermediate products using gas chromatography with simultaneous mass spectrometric and flame ionization detections.

    PubMed

    Sťávová, Jana; Stahl, Danese C; Seames, Wayne S; Kubátová, Alena

    2012-02-10

    Accurate analytical methods are required to develop and evaluate the quality of new renewable transportation fuels and intermediate organic liquid products (OLPs). Unfortunately, existing methods developed for the detailed characterization of petroleum products, are not accurate for many of the OLPs generated from non-petroleum feedstocks. In this study, a method was developed and applied to the detailed characterization of complex OLPs formed during triacylglyceride (TG) pyrolysis which is the basis for generating one class of emerging biofuels. This method uses gas chromatography coupled simultaneously with flame ionization and mass spectrometry detectors (GC-FID/MS). The FID provided accurate quantification of carbonaceous species while MS enabled identification of unknown compounds. A programed temperature vaporizer using a 25 °C, 0.1 min, 720 °C min(-1), 350 °C, 5 min temperature program is employed which minimizes compound discrimination better than the more commonly utilized split/splitless injector, as verified with injections at 250 and 350 °C. Two standard mixtures featuring over 150 components are used for accurate identification and a designed calibration standard accounts for compound discrimination at the injector and differing FID responses of various classes of compounds. This new method was used to identify and quantify over 250 species in OLPs generated from canola oil, soybean oil, and canola methyl ester (CME). In addition to hydrocarbons, the method was used to quantify polar (upon derivatization) and unidentified species, plus the unresolved complex mixture that has not typically been determined in previous studies. Repeatability of the analytical method was below 5% RSD for all individual components. Using this method, the mass balance was closed for samples derived from canola and soybean oil but only ca. 77 wt% of the OLP generated from CME could be characterized. The ability to close the mass balance depended on sample origin

  10. A novel method for the determination of black liquor viscosity by multiple headspace extraction gas chromatography.

    PubMed

    Hu, Hui-Chao; Chai, Xin-Sheng

    2013-12-13

    This work demonstrates a novel method for the determination of viscosity in the concentrated black liquors from pulp mill recovery process. The method is based on the kinetic release of methanol (a vapor tracer) to the headspace in a sample closed vial by a multiple headspace extraction gas chromatographic technique. Both theoretical and empirical models were proposed for establishing the correlation with the reference method. The results showed that the correlation using either of the models is excellent for the tested black liquor samples (at 110°C). The presented method is simple and practical and can be a valuable tool for black liquor viscosity related research and applications.

  11. Effects of Hemoglobin Variants on Hemoglobin A1c Values Measured Using a High-Performance Liquid Chromatography Method

    PubMed Central

    De-La-Iglesia, Silvia; Ropero, Paloma; Nogueira-Salgueiro, Patricia; Santana-Benitez, Jesus

    2014-01-01

    Hemoglobin A1c (HbA1c) is routinely used to monitor long-term glycemic control and for diagnosing diabetes mellitus. However, hemoglobin (Hb) gene variants/modifications can affect the accuracy of some methods. The potential effect of Hb variants on HbA1c measurements was investigated using a high-performance liquid chromatography (HPLC) method compared with an immunoturbimetric assay. Fasting plasma glucose (FPG) and HbA1c levels were measured in 42 371 blood samples. Samples producing abnormal chromatograms were further analyzed to characterize any Hb variants. Fructosamine levels were determined in place of HbA1c levels when unstable Hb variants were identified. Abnormal HPLC chromatograms were obtained for 160 of 42 371 samples. In 26 samples HbS was identified and HbA1c results correlated with FPG. In the remaining 134 samples HbD, Hb Louisville, Hb Las Palmas, Hb N-Baltimore, or Hb Porto Alegre were identified and HbA1c did not correlate with FPG. These samples were retested using an immunoturbidimetric assay and the majority of results were accurate; only 3 (with the unstable Hb Louisville trait) gave aberrant HbA1c results. Hb variants can affect determination of HbA1c levels with some methods. Laboratories should be aware of Hb variants occurring locally and choose an appropriate HbA1c testing method. PMID:25355712

  12. Screening of extraction methods for glycoproteins from jellyfish ( Rhopilema esculentum) oral-arms by high performance liquid chromatography

    NASA Astrophysics Data System (ADS)

    Ren, Guoyan; Li, Bafang; Zhao, Xue; Zhuang, Yongliang; Yan, Mingyan; Hou, Hu; Zhang, Xiukun; Chen, Li

    2009-03-01

    In order to select an optimum extraction method for the target glycoprotein (TGP) from jellyfish ( Rhopilema esculentum) oral-arms, a high performance liquid chromatography (HPLC)-assay for the determination of the TGP was developed. Purified target glycoprotein was taken as a standard glycoprotein. The results showed that the calibration curves for peak area plotted against concentration for TGP were linear ( r = 0.9984, y = 4.5895 x+47.601) over concentrations ranging from 50 to 400 mgL-1. The mean extraction recovery was 97.84% (CV2.60%). The fractions containing TGP were isolated from jellyfish ( R. esculentum) oral-arms by four extraction methods: 1) water extraction (WE), 2) phosphate buffer solution (PBS) extraction (PE), 3) ultrasound-assisted water extraction (UA-WE), 4) ultrasound-assisted PBS extraction (UA-PE). The lyophilized extract was dissolved in Milli-Q water and analyzed directly on a short TSK-GEL G4000PWXL (7.8 mm×300 mm) column. Our results indicated that the UA-PE method was the optimum extraction method selected by HPLC.

  13. Purification method development for chiral separation in supercritical fluid chromatography with the solubilities in supercritical fluid chromatographic mobile phases.

    PubMed

    Gahm, Kyung H; Tan, Helming; Liu, Jodi; Barnhart, Wesley; Eschelbach, John; Notari, Steve; Thomas, Samuel; Semin, David; Cheetham, Janet

    2008-04-14

    A comprehensive approach was applied to develop a chiral purification method for an analyte that was found to be unusually difficult to scale-up in supercritical fluid chromatography (SFC). This was performed by studying major factors such as the solubility of an analyte in SFC mobile phases, impurity profiles, and cycle time. For this case study, the solubility in SFC mobile phase was measured by a packed column technique, coupled with a novel trapping mechanism to enhance measurement precision in SFC conditions. The solubility studies in SFC mobile phases suggested a couple of possible SFC mobile phases, in which the analyte would potentially be most soluble. The SFC methods were developed to purify a sample containing 15% of an impurity, after considering impurity profiles and cycle times of several potential methods in addition to SFC mobile phase solubility. An equal volume mixture of acetonitrile and ethanol was chosen for the final purification method, since this mixture demonstrated the relatively high SFC solubility among all solvent combinations with enhanced resolution between the analyte and the impurity as well as the shortest run time. The solubility of the compound was also determined in various organic solvents using a high throughput solubility screening system to better understand relative change of solubility from neat solution to SFC mobile phases.

  14. Gas chromatography-high-resolution mass spectrometric method for determination of methamphetamine and its major metabolite amphetamine in human hair.

    PubMed

    Kim, Jin Young; Suh, Sung Ill; In, Moon Kyo; Chung, Bong Chul

    2005-01-01

    Gas chromatography-high-resolution mass spectrometric (GC-HRMS) method is presented for the qualitative and quantitative analysis of methamphetamine (MA) and its major metabolite, amphetamine (AMP), in human hair. The method procedure involves decontamination of hair with distilled water and acetone, acidic hydrolysis and extraction in the presence of the internal standard, and GC-HRMS selective ion monitoring (SIM) analysis. The limits of detection (LOD) were 9 pg/mg for MA and 21 pg/mg for AMP using a 30-mg hair sample, and the SIM responses were linear with coefficients of correlation ranged from 0.9998 to 0.9999. The recoveries were found to be 91.1-92.3%. By using HRMS (resolution of 5000), detection sensitivity is improved because of the elimination of the biological background, and the LODs for MA and AMP were 2.4-4.4 times lower than those of low-resolution MS. The GC-HRMS method was successfully applied to the analysis of cosmetically treated hair, which is difficult to analyze with the conventional method.

  15. New method for determination of epichlorohydrin in epoxy-coated cans by oxolane derivatization and gas chromatography-mass spectrometry.

    PubMed

    Sung, Jun Hyun; Lee, Young Ja; Park, Hyun Jin

    2008-08-01

    A new method was developed for the determination of epichlorohydrin (ECH) in food contact surface of epoxy-coated cans. The oxolane derivative, which produced by reaction of epoxy moiety in ECH with cyclopentanone in the presence of borontrifluoride-diethyletherate, was analyzed by gas chromatography (GC)/mass spectrometry (MS). 1,2-Epoxyhexane was used as internal standard (IS), which produced an oxolane derivative under the same reaction mechanism as ECH. The developed method was validated with 1 ng ml(-1) of limit of detection (LOD, surface area related 20 ng dm(-2)), >0.999 of linearity. Good precision, which was tested both in terms of intra-day repeatability and inter-day reproducibility, and 97.3-102.7% of good recoveries were obtained on three spiked levels of 5.2, 40.3 and 149.1 ng ml(-1). The excellent validation data suggests that this method is more simple, quick and effective than the official method in European Committee for Standardization (CEN) to determine the residual amount of ECH in food contact materials for food law compliance test. The residual amount of ECH for 13 epoxy-coated can samples was analyzed, and none of the samples was found to be detectable levels of ECH in epoxy-coated cans.

  16. A new liquid chromatography-tandem mass spectrometry method for determination of parabens in human placental tissue samples.

    PubMed

    Jiménez-Díaz, I; Vela-Soria, F; Zafra-Gómez, A; Navalón, A; Ballesteros, O; Navea, N; Fernández, M F; Olea, N; Vílchez, J L

    2011-05-15

    Endocrine disruptors are a group of organic compounds widely used, which are ubiquitous in the environment and in biological samples. The main effect of these compounds is associated with their ability to mimic or block the action of natural hormones in living organisms, including humans. Parabens (esters of p-hydroxybenzoic acid) belong to this group of compounds. In this work, we propose a new liquid chromatography-tandem mass spectrometry (LC-MS/MS) method to asses the presence of parabens most commonly used in industrial applications (methyl-, ethyl-, propyl- and butyl-paraben) in samples of human placental tissue. The method involves the extraction of the analytes from the samples using ethyl acetate, followed by a clean-up step using centrifugation prior to their quantification by LC-MS/MS using an atmospheric pressure chemical ionization (APCI) interface in the negative mode. Deuterated bisphenol A (BPA-d(16)) was used as surrogate. Found detection limits (LOD) ranged from 0.03 to 0.06 ng g(-1) and quantification limits (LOQ) from 0.1 to 0.2 ng g(-1), while inter- and intra-day variability was under 13.8%. The method was validated using standard addition calibration and a spike recovery assay. Recovery rates for spiked samples ranged from 82% to 108%. This method was satisfactorily applied for the determination of parabens in 50 placental tissue samples collected from women who live in the province of Granada (Spain).

  17. Development of a liquid chromatography-tandem mass spectrometry method for quantitative analysis of trace d-amino acids.

    PubMed

    Nakano, Yosuke; Konya, Yutaka; Taniguchi, Moyu; Fukusaki, Eiichiro

    2017-01-01

    d-Amino acids have recently attracted much attention in various research fields including medical, clinical and food industry due to their important biological functions that differ from l-amino acid. Most chiral amino acid separation techniques require complicated derivatization procedures in order to achieve the desirable chromatographic behavior and detectability. Thus, the aim of this research is to develop a highly sensitive analytical method for the enantioseparation of chiral amino acids without any derivatization process using liquid chromatography-tandem mass spectrometry (LC-MS/MS). By optimizing MS/MS parameters, we established a quantification method that allowed the simultaneous analysis of 18 d-amino acids with high sensitivity and reproducibility. Additionally, we applied the method to food sample (vinegar) for the validation, and successfully quantified trace levels of d-amino acids in samples. These results demonstrated the applicability and feasibility of the LC-MS/MS method as a novel, effective tool for d-amino acid measurement in various biological samples.

  18. Facile method of quantification for oxidized tryptophan degradants of monoclonal antibody by mixed mode ultra performance liquid chromatography.

    PubMed

    Wong, Cintyu; Strachan-Mills, Camille; Burman, Sudhir

    2012-12-28

    Oxidation in therapeutic monoclonal antibody is a common occurrence and it may affect potency. Thus controlling and monitoring the amount of oxidized variant in the drug product sample is important since it may impact the purity. Here, we present the development of a fast and easy method utilizing size exclusion - ultra performance liquid chromatography (SE-UPLC) run under moderate hydrophobic conditions (mixed mode) to monitor the heterogeneity in drug product samples. The best separation was obtained using Waters Acquity BEH200 size exclusion column along with a mobile phase consisting of sodium acetate and sodium sulfate that separates IgG into aggregate, monomer, and fragment. The moderate salt concentration resulted in a second mode of separation based on hydrophobicity, resolving a monomer pre-peak from the monomer main peak. Multi-angle light scattering (MALS) determined the pre-peak has a similar mass as the IgG monomer. Characterization of the purified pre-peak fraction using mass spectrometry (MS), and bioactivity revealed this degradant to be a Trp-oxidized IgG monomer with significantly reduced bioactivity. Method qualification of the mixed mode UPLC method showed good recovery for the spiked monomer pre-peak and Fab fragment. However, the recovery of spiked dimer was low. This method is suitable for determining the relative distribution of the oxidized monomer and the native monomer species.

  19. Simultaneous determination of spinetoram residues in tomato by high performance liquid chromatography combined with QuEChERS method.

    PubMed

    Malhat, Farag Mahmoud

    2013-02-01

    A sensitive and simple method for simultaneous analysis of spinetoram residues and its dissipation in tomato were studied. Spinetoram residues were extracted from tomato samples with acetonitrile. The extract was cleaned-up with QuEChERS method by dispersive solid-phase extraction with primary secondary amine sorbent to remove co-extractives, prior to analysis by high performance liquid chromatography coupled with diode array detector (HPLC-DAD). This method is characterized by recovery >97 %, relative standard division (RSD) <12.3 %, and limit of quantification (LOQ) of 0.04 mg kg(-1), in agreement with directive for method validation in residue analysis. Also, the results showed that spinetoram dissipation pattern followed the first order kinetics with the half-life of 2.6 days, in tomato. The spinetoram residues in tomato were below the codex maximum residue level (0.06 mg kg(-1)) after 10 days of application. This study suggests that spinetoram is acceptable to apply for tomato under the recommended dosage.

  20. An improved validated ultra high pressure liquid chromatography method for separation of tacrolimus impurities and its tautomers.

    PubMed

    Subasranjan, Acharya; C, Srinivasulu; Hemant, Raoutray

    2010-03-01

    A selective, specific and sensitive ultra high pressure liquid chromatography (UHPLC) method was developed for determination of tacrolimus degradation products and tautomers in the preparation of pharmaceuticals. The chromatographic separation was performed on Waters ACQUITY UPLC system and BEH C₈ column using gradient elution of mobile phase A (90:10 v/v of 0.1% v/v triflouroacetic acid solution and Acetonitrile) and mobile phase B (90:10 v/v acetonitrile and water) at a flow rate of 0.6 mL min⁻¹. Ultraviolet detection was performed at 210 nm. Tacrolimus, tautomers and impurities were chromatographed with a total run time of 25 min. Calibration showed that the response of impurity was a linear function of concentration over the range 0.3-6 µg mL⁻¹ (r² ≥ 0.999) and the method was validated over this range for precision, intermediate precision, accuracy, linearity and specificity. For precision study, percentage relative standard deviation of each impurity was < 15% (n = 6). The method was found to be precise, accurate, linear and specific. The proposed method was successfully employed for estimation of tacrolimus impurities in pharmaceutical preparations.

  1. Fingerprint Analysis of Desmodium Triquetrum L. Based on Ultra Performance Liquid Chromatography with Photodiode Array Detector Combined with Chemometrics Methods.

    PubMed

    Zhang, Meiling; Zhao, Cui; Liang, Xianrui; Ying, Yin; Han, Bing; Yang, Bo; Jiang, Cheng

    2016-01-01

    A fingerprinting approach was developed by means of ultra high-performance liquid chromatography with photodiode array detector for the quality control of Desmodium triquetrum L., an herbal medicine widely used for clinical purposes. Ten batches of raw material samples of D. triquetrum were collected from different regions of China. All UPLC analyses were carried out on a Waters ACQUITY UPLC BEH shield RP18 column (2.1 × 50 mm, 1.7 µm particle size) at 60°C, with a gradient mobile phase composed of 0.1% aqueous formic acid and acetonitrile at a flow rate of 0.45 mL/min. The method validation results demonstrated the developed method possessing desirable reproducibility, efficiency, and allowing fingerprint analysis in one chromatographic run within 13 min. The quality assessment was achieved by using chemometrics methods including similarity analysis, hierarchical clustering analysis and principal component analysis. The developed method can be used for further quality control of D. triquetrum.

  2. Rapid screening method for determination of Ecstasy and amphetamines in urine samples using gas chromatography-chemical ionisation mass spectrometry.

    PubMed

    Pellegrini, M; Rosati, F; Pacifici, R; Zuccaro, R; Romolo, F S; Lopez, A

    2002-04-05

    The need for analytical screening tests more reliable and valid to detect amphetamine and related "designer drugs" in biological samples is becoming critical, due to the increasing diffusion of these drugs on the European illegal market. The most common screening procedures based on immunoassays suffer a number of limitations, including low sensitivity, lack of specificity and limited number of detectable substances. This paper describes a screening method based on gas-chromatography-mass-spectrometry (GC/MS) using positive chemical ionisation (PCI) detection. Methanol was used as reactant gas in the ionisation chamber. Molecular ions of different compounds were monitored, allowing a sensitivity of 5-10 ng/ml with high selectivity. The sensitivity of the method gives positive results in samples taken 48-72 h after intake of one dose of 50-100 mg. The method is simple and rapid. Sample preparation was limited to one liquid-liquid extraction, without any hydrolysis and derivatisation. Hydrolysis is critical to identify metabolites excreted as conjugates. Blank urine samples spiked with known amounts of amphetamine (AM), methylamphetamine (MA), methylenedioxyamphetamine (MDA), methylenedioxymethylamphetamine (MDMA), methylenedioxyethylamphetamine (MDEA) and methylenedioxyphenyl-N-methyl-2-butanamine (MBDB) were analysed. The method was successfully tested on real samples of urine from people, whose use of amphetamine was suspected, and results were compared with results obtained with immunoassays.

  3. Sample-directed pseudotargeted method for the metabolic profiling analysis of rice seeds based on liquid chromatography with mass spectrometry.

    PubMed

    Zhang, Junjie; Zhao, Chunxia; Zeng, Zhongda; Luo, Ping; Zhao, Yanni; Zhao, Jieyu; Li, Lili; Lu, Xin; Xu, Guowang

    2016-01-01

    Rice is one of the most important food crops in the world. Metabolite composition in rice seeds varies significantly depending on genetic variety, climatic alternation and agricultural practice. Metabolomics is a powerful tool to reveal the metabolic response of rice to various conditions. In this work, a rice seed sample-directed pseudotargeted metabolomics method was first established and validated based on ultra high performance liquid chromatography with triple quadrupole mass spectrometry in the multiple reaction monitoring mode. A total of 749 and 617 ion pairs in positive and negative modes were achieved, respectively. Among them, about 200 metabolites were identified or tentatively identified. The developed method showed better linearity and repeatability than those of non-targeted metabolomics method. Good intra-day and inter-day precisions, recoveries and wide linear range were also obtained. Furthermore, the method was applied for the investigation of metabolic variation of rice seeds with two wild cultivars and their transgenic lines that were grown in two locations. Principal component analysis indicated that the effects of cultivar and location on metabolic variations were far more than those of gene modification. The nonparametric Mann-Whitney U test revealed that most metabolites were influenced by cultivar, location and gene modifications together.

  4. Development and validation of the liquid chromatography-tandem mass spectrometry method for quantitative estimation of candesartan from human plasma

    PubMed Central

    Prajapati, Shailesh T.; Patel, Pratik K.; Patel, Marmik; Chauhan, Vijendra B.; Patel, Chhaganbhai N.

    2011-01-01

    Introduction: A simple and sensitive liquid chromatography-tandem mass spectrometry method was developed and validated for estimation of candesartan in human plasma using the protein precipitation technique. Materials and Methods: The chromatographic separation was performed on reverse phase using a Betasil C8 (100 × 2.1 mm) 5-μm column, mobile phase of methanol:ammonium tri-floro acetate buffer with formic acid (60:40 v/v) and flow rate of 0.45 ml/min. The protonated analyte was quantitated in positive ionization by multiple reaction monitoring with a mass spectrometer. The mass transitions m/z 441.2 → 263.2 and 260.2 → 116.1 were used to measure candesartan by using propranolol as an internal standard. Results: The linearity of the developed method was achieved in the range of 1.2–1030 ng/ml (r2 ≥ 0.9996) for candesartan. Conclusion: The developed method is simple, rapid, accurate, cost-effective and specific; hence, it can be applied for routine analysis in pharmaceutical industries. PMID:23781443

  5. Development of a Gas Chromatography-Mass Spectrometry Method for the Quantification of Glucaric Acid Derivatives in Beverage Substrates

    PubMed Central

    Craig, Ana Paula; Fields, Christine C.; Simpson, John V.

    2014-01-01

    A gas chromatography-mass spectrometry (GC-MS) method using the standard addition methodology was developed for the determination of glucuronolactone (GL) and glucuronic acid (DGuA) in four beverages categorized as detoxification, recovery, or energy drinks. The method features a precolumn derivatization step with a combination of BSTFA (N,O-bis(trimethylsilyl)trifluoroacetamide) and TMCS (trimethylchlorosilane) to silylate the analytes. The sample pretreatment required no extraction, filtration, or reduction step prior to the injection. The quantification of the analytes was performed using a five-point standard addition protocol. The proposed method presented excellent intraday precision (%RSD < 10) and linearity for GL calibration curves (correlation coefficients > 0.995) and acceptable linearity for DGuA calibration curves (correlation coefficients > 0.97). The estimated limits of detection (LOD) and quantification (LOQ) for GL ranged from 0.006 ppm to 0.14 ppm, and 0.02 ppm to 0.47 ppm, respectively. The estimated LOD and LOQ for DGuA determination ranged, respectively, from 0.06 ppm to 1.1 ppm and 0.2 ppm to 3.8 ppm. The results demonstrated that the method should be regarded as a reliable alternative to the simultaneous determination of GL and DGuA. PMID:25024704

  6. An online coupled peritoneal macrophage/cell membrane chromatography and high-performance liquid chromatography/mass spectrometry method to screen for anti-inflammatory components from the Chinese traditional medicine Chloranthus multistachys Pei.

    PubMed

    Li, Weifeng; Xing, Wei; Wang, Sicen; Fan, Ting; Huang, Huimin; Niu, Xiaofeng; He, Langchong

    2013-11-01

    Cell membrane chromatography (CMC) is a chromatographic biological affinity method that uses specific cell membranes as the stationary phase. In this study, a novel peritoneal macrophage/cell membrane chromatography (PM/CMC)-online-high performance liquid chromatography/mass spectrometry (HPLC/MS) method was established to screen for the anti-inflammatory components from traditional Chinese medicines using hydrocortisone and dexamethasone as standards. The stationary phase of the CMC employed mouse peritoneal macrophage cell membranes. This method was applied to the purification and identification of components in extracts of Chloranthus multistachys Pei. The major component retained by CMC was identified as isofraxidin by HPLC/MS. In vitro experiments revealed that IF was able to inhibit the production of nitric oxide and tumor necrosis factor-α in lipopolysaccharide-stimulated mice and peritoneal macrophages in a dose-dependent manner. The results demonstrated that the PM/CMC-online-HPLC/MS is an effective screening system for the rapid detection, enrichment, and identification of target components from complex samples.

  7. Evaluation of the quantitative performances of supercritical fluid chromatography: from method development to validation.

    PubMed

    Dispas, Amandine; Lebrun, Pierre; Ziemons, Eric; Marini, Roland; Rozet, Eric; Hubert, Philippe

    2014-08-01

    Recently, the number of papers about SFC increased drastically but scientists did not truly focus their work on quantitative performances of this technique. In order to prove the potential of UHPSFC, the present work discussed about the different steps of the analytical life cycle of a method: from development to validation and application. Moreover, the UHPSFC quantitative performances were evaluated in comparison with UHPLC, which is the main technique used for quality control in the pharmaceutical industry and then could be considered as a reference. The methods were developed using Design Space strategy, leading to the optimization of robust method. In this context, when the Design Space optimization shows guarantee of quality, no more robustness study is required prior to the validation. Then, the methods were geometrically transferred in order to reduce the analysis time. The UHPSFC and UHPLC methods were validated based on the total error approach using accuracy profile. Even if UHPLC showed better precision and sensitivity, UHPSFC method is able to give accurate results in a dosing range larger than the 80-120% range required by the European Medicines Agency. Consequently, UHPSFC results are valid and could be used for the control of active substance in a finished pharmaceutical product. Finally, UHPSFC validated method was used to analyse real samples and gave similar results than the reference method (UHPLC).

  8. Development Of Ion Chromatography Methods To Support Testing Of The Glycolic Acid Reductant Flowsheet In The Defense Waste Processing Facility

    SciTech Connect

    Wiedenman, B. J.; White, T. L.; Mahannah, R. N.; Best, D. R.; Stone, M. E.; Click, D. R.; Lambert, D. P.; Coleman, C. J.

    2013-10-01

    Ion Chromatography (IC) is the principal analytical method used to support studies of Sludge Reciept and Adjustment Tank (SRAT) chemistry at DWPF. A series of prior analytical ''Round Robin'' (RR) studies included both supernate and sludge samples from SRAT simulant, previously reported as memos, are tabulated in this report.2,3 From these studies it was determined to standardize IC column size to 4 mm diameter, eliminating the capillary column from use. As a follow on test, the DWPF laboratory, the PSAL laboratory, and the AD laboratory participated in the current analytical RR to determine a suite of anions in SRAT simulant by IC, results also are tabulated in this report. The particular goal was to confirm the laboratories ability to measure and quantitate glycolate ion. The target was + or - 20% inter-lab agreement of the analyte averages for the RR. Each of the three laboratories analyzed a batch of 12 samples. For each laboratory, the percent relative standard deviation (%RSD) of the averages on nitrate, glycolate, and oxalate, was 10% or less. The three laboratories all met the goal of 20% relative agreement for nitrate and glycolate. For oxalate, the PSAL laboratory reported an average value that was 20% higher than the average values reported by the DWPF laboratory and the AD laboratory. Because of this wider window of agreement, it was concluded to continue the practice of an additional acid digestion for total oxalate measurement. It should also be noted that large amounts of glycolate in the SRAT samples will have an impact on detection limits of near eluting peaks, namely Fluoride and Formate. A suite of scoping experiments are presented in the report to identify and isolate other potential interlaboratory disceprancies. Specific ion chromatography inter-laboratory method conditions and differences are tabulated. Most differences were minor but there are some temperature control equipment differences that are significant leading to a recommendation of

  9. Validation of a two-dimensional liquid chromatography method for quality control testing of pharmaceutical materials.

    PubMed

    Yang, Samuel H; Wang, Jenny; Zhang, Kelly

    2017-04-07

    Despite the advantages of 2D-LC, there is currently little to no work in demonstrating the suitability of these 2D-LC methods for use in a quality control (QC) environment for good manufacturing practice (GMP) tests. This lack of information becomes more critical as the availability of commercial 2D-LC instrumentation has significantly increased, and more testing facilities begin to acquire these 2D-LC capabilities. It is increasingly important that the transferability of developed 2D-LC methods be assessed in terms of reproducibility, robustness and performance across different laboratories worldwide. The work presented here focuses on the evaluation of a heart-cutting 2D-LC method used for the analysis of a pharmaceutical material, where a key, co-eluting impurity in the first dimension ((1)D) is resolved from the main peak and analyzed in the second dimension ((2)D). A design-of-experiments (DOE) approach was taken in the collection of the data, and the results were then modeled in order to evaluate method robustness using statistical modeling software. This quality by design (QBD) approach gives a deeper understanding of the impact of these 2D-LC critical method attributes (CMAs) and how they affect overall method performance. Although there are multiple parameters that may be critical from method development point of view, a special focus of this work is devoted towards evaluation of unique 2D-LC critical method attributes from method validation perspective that transcend conventional method development and validation. The 2D-LC method attributes are evaluated for their recovery, peak shape, and resolution of the two co-eluting compounds in question on the (2)D. In the method, linearity, accuracy, precision, repeatability, and sensitivity are assessed along with day-to-day, analyst-to-analyst, and lab-to-lab (instrument-to-instrument) assessments. The results of this validation study demonstrate that the 2D-LC method is accurate, sensitive, and robust and is

  10. Alternative purification method for recombinant measles viral nucleoprotein expressed in insect cells by ion-exchange chromatography.

    PubMed

    Lee, Han Saem; Kim, You-Jin; Yang, Jeongsun; Yoon, Hee Sook; Kim, Seung Tae; Kim, Kisoon

    2014-03-01

    Recombinant measles virus nucleoproteins (rMeV N) and fusion (F) proteins were characterized as major antigenic proteins expressed in insect cells mediated by recombinant baculoviruses (rBVs). Band intensities were analyzed by Western blotting to recognize IgG and IgM antibodies against the rMeV N and F proteins in human sera and cerebrospinal fluids (CSFs) from patients with measles infections. Positive results from the blots using the rMeV N were consistent with the results of enzyme-linked immunosorbent assays (ELISAs) in which whole viral proteins were used as antigens. Human sera and CSFs reacted more strongly with the rMeV N than with the rMeV F proteins prepared in an identical expression system. For efficient and reliable purification, ion-exchange chromatography using Source Q anion resin was applied, and high-purity rMeV N protein was harvested. To characterize the similarity with the native viral protein to purified N protein, structural mimicry of purified recombinant proteins with intact rMeV N was shown through transmission electron microscopy, and the truncation and the phosphorylation status of the expressed protein were analyzed. These results suggest that the rMeV N purified by ion-exchange chromatography has features similar to those of naïve N including a self-assembled structure, phosphorylation and antigenic function. Thus, these expression and purification methods can be applied to the large-scale production of the rMeV N, which is essential for the development of new diagnostic tools and vaccines for acute and chronic MeV infections.

  11. Development of an offline bidimensional high-performance liquid chromatography method for analysis of stereospecific triacylglycerols in cocoa butter equivalents.

    PubMed

    Kadivar, Sheida; De Clercq, Nathalie; Nusantoro, Bangun Prajanto; Le, Thien Trung; Dewettinck, Koen

    2013-08-21

    Acyl migration is a serious problem in enzymatic modification of fats and oils, particularly in production of cocoa butter equivalent (CBE) through enzymatic acidolysis reaction, which leads to the formation of non-symmetrical triacylglycerols (TAGs) from symmetrical TAGs. Non-symmetrical TAGs may affect the physical properties of final products and are therefore often undesired. Consequently, an accurate method is needed to determine positional isomer TAGs during the production of CBE. A bidimentional high-performance liquid chromatography (HPLC) method with combination of non-aqueous reversed-phase HPLC and silver ion HPLC joining with an evaporative light scattering detector was successfully developed for the analysis of stereospecific TAGs. The best separation of positional isomer standards was obtained with a heptane/acetone mobile-phase gradient at 25 °C and 1 mL/min. The developed method was then used in multidimensional determination of the TAG positional isomers in fat and oil blends and successfully identified the TAGs and possible isomers in enzymatically acidolyzed CBE.

  12. Screening method for the determination of tetracyclines and fluoroquinolones in animal drinking water by liquid chromatography with diode array detector.

    PubMed

    Patyra, E; Kowalczyk, E; Grelik, A; Przeniosło-Siwczyńska, M; Kwiatek, K

    2015-01-01

    A liquid chromatography - diode array detector (HPLC-DAD) procedure has been developed for the determination of oxytetracycline (OTC), tetracycline (TC), chlorotetracycline (CTC), doxycycline (DC), enrofloxacin (ENR), ciprofloxacin (CIP), sarafloxacin (SAR) and flumequine (FLU) residues in animal drinking water. This method was applied to animal drinking water. Solid-phase extraction (SPE) clean-up on an Oasis HLB cartridge allowed an extract suitable for liquid chromatographic analysis to be obtained. Chromatographic separation was carried out on a C18 analytical column, using gradient elution with 0.1% trifluoroacetic acid - acetonitrile - methanol at 30°C. The flow-rate was 0.7 mL/min and the eluate was analysed at 330 nm. The whole procedure was evaluated according to the requirements of the Commission Decision 2002/657/EC, determining specificity, decision limit (CCα), detection capacity (CCβ), limit of detection (LOD), limit of quantification (LOQ), precision and accuracy during validation of the method. The recoveries of TCs and FQs from spiked samples at the levels of 10, 100 and 1000 μg/L were higher than 82%. The developed method based on HPLC-DAD has been applied for the determination of four tetracyclines and four fluoroquinolones in animal drinking water samples.

  13. A fast gas chromatography/mass spectrometry method for the determination of stimulants and narcotics in urine.

    PubMed

    Strano Rossi, Sabina; de la Torre, Xavier; Botrè, Francesco

    2010-05-30

    A fast method has been developed for the simultaneous determination of 52 stimulants and narcotics excreted unconjugated in urine by gas chromatography/mass spectrometry (GC/MS). The procedure involves the liquid/liquid extraction of the analytes from urine at strong alkaline pH and the injection of the extract into a GC/MS instrument with a fast GC column (10 m x 0.18 mm i.d.); the short column allows the complete separation of the 52 analytes in a chromatographic run of 8 min. The method has been fully validated giving lower limits of detection (LLODs) satisfactory for its application to antidoping analysis as well as to forensic toxicology. The repeatability of the concentrations and the retention times are good both for intra- and for inter-day experiments (%CV of concentrations always lower than 15 and %CV of retention times lower than 0.6). In addition, the analytical bias is satisfactory (A% always >15%). The method proposed here would be particularly useful whenever there are time constraints and the analyses have to be completed in the shortest possible time.

  14. Validation of a liquid chromatography method for the simultaneous quantification of ochratoxin A and its analogues in red wines.

    PubMed

    Remiro, Rebeca; Ibáñez-Vea, María; González-Peñas, Elena; Lizarraga, Elena

    2010-12-24

    A validated high-performance liquid chromatography (HPLC) method with fluorescence detection for the simultaneous quantification of ochratoxin A (OTA) and its analogues (ochratoxin B (OTB), ochratoxin C (OTC) and methyl ochratoxin A (MeOTA)) in red wine at trace levels is described. Before their analysis by HPLC-FLD, ochratoxins were extracted and purified with immunoaffinity columns from 50 mL of red wine at pH 7.2. Validation of the analytical method was based on the following parameters: selectivity, linearity, robustness, limits of detection and quantification, precision (within-day and between-day variability), recovery and stability. The limits of detection (LOD) in red wine were established at 0.16, 0.32, 0.27 and 0.17 ng L(-1) for OTA, OTB, MeOTA and OTC, respectively. The limit of quantification (LOQ) was established as 0.50 ng L(-1) for all of the ochratoxins. The LOD and LOQ obtained are the lowest found for OTA in the reference literature up to now. Recovery values were 93.5, 81.7, 76.0 and 73.4% for OTA, OTB, MeOTA and OTC, respectively. For the first time, this validated method permits the investigation of the co-occurrence of ochratoxins A, B, C and methyl ochratoxin A in 20 red wine samples from Spain.

  15. Stepwise elusion method in micellar electrokinetic chromatography via sequential use of lithium perfluorooctadecyl sulfonate and lithium dodecyl sulfate.

    PubMed

    Esaka, Yukihiro; Rin, Fumiaki; Kobayashi, Miki; Osako, Ryohei; Murakami, Hiroya; Uno, Bunji

    2014-09-05

    An effective stepwise micellar electrokinetic chromatography (MEKC) elution method was developed using lithium perfluorooctadecyl sulfonate (LPFOS) and lithium dodecyl sulfate (LDS). The hydrogen-bonding property of LPFOS micelles differs from that of LDS micelles, which leads to remarkably different selectivity in the transfer of solutes to the micelles. The present stepwise method is performed by replacing the inlet reservoir of a first running solution containing LPFOS with that of a second running solution containing LDS during a single separation run in the absence of electroosmotic flow under acidic conditions, where LPFOS micelles work as carriers in first and then LDS micelles turn over. Effective separation of 15 nonionic aromatic compounds was controlled well by adjusting the time in the inlet reservoir, which could not be accomplished with systems using only LPFOS or only LDS, with significant changes in the elution order where necessary. Furthermore, separations with the present stepwise method were easily simulated, and the replacement time was optimized for 3.1min from a 70.0mM LPFOS solution to a 67.5mM LDS solution with nearly complete separation within 15min using the simulated parameters.

  16. Novel multiresidue method for determination of pesticides in red wine using gas chromatography-mass spectrometry and solid phase extraction.

    PubMed

    Pelajić, Maja; Peček, Gorana; Mutavdžić Pavlović, Dragana; Vitali Čepo, Dubravka

    2016-06-01

    A new multiresidue method was developed for determination of 25 pesticide residues in red wine by gas chromatography coupled to mass spectrometry with a single run of 23.63 min. Samples were extracted from wine with solid phase extraction using Oasis HLB. Mixture of methanol and water was used for rinsing, while acetonitrile and n-hexane were used as elution solvents. Method was validated according to SANCO/12571/2013 criteria in wide linearity range (limit of quantification - 400 μg L(-1)). Limits of quantification (LOQ) were well below 10 μg L(-1) for most pesticides and recoveries at 2×LOQ and 10×LOQ concentration levels were in range 70-120%. Precision, expressed as a relative standard deviation, was always under 14%. The method was applied to 32 red wine samples from Croatia. Pesticides were detected in 30 samples with a total of 15 pesticides found, 7 of which were at a high concentration.

  17. A simple and rapid method for detection of paraquat in human plasma by high-performance liquid chromatography

    PubMed Central

    Sun, Bin; Chen, Yuguo

    2015-01-01

    We have developed an effective analytical method to determine the concentration of paraquat in human plasma by high-performance liquid chromatography (HPLC), which can provide methodological support for diagnosis, therapy, and prognostic evaluation of acute poisoning related to paraquat. The plasma samples were deproteinized with 35% perchloric acid. Then the paraquat was extracted from the samples and separated on Ultimate XB-C18 column with a mobile phase consisting of 0.1 M phosphate buffer (containing 75 mM sodium heptane sulfonate) and methyl cyanides (PH 3.0), flow rate of 1 mL/min, column temperature of 28°C and a detection wave length on 258 nm. A good linearity was obtained within the range of 0.2 to 500 g/ml in human plasma. The average rate of absolute and relative recovery were about 100.6% and 101.31%, accompanied by the variations less than 3% and 6% respectively. The within- and between-day relative standard deviations were all less than 6%. After repeated freeze and thaw of plasma samples, no significant difference was observed for the concentration of paraquat (P>0.05). The method was validated by successfully applying in one patient with acute paraquat poisoning. Due to the celerity, accuracy and no-interference by other components of blood sample, this method was suitable for determination the concentration of paraquat in human plasma. PMID:26770298

  18. Simultaneous determination of methamphetamine and its metabolite, amphetamine, in urine using a high performance liquid chromatography column-switching method.

    PubMed

    Kumihashi, Mitsuru; Ameno, Kiyoshi; Shibayama, Takayuki; Suga, Keisuke; Miyauchi, Hiroshi; Jamal, Mostofa; Wang, Weihuan; Uekita, Ikuo; Ijiri, Iwao

    2007-01-01

    We describe here a simple, precise, and highly sensitive method for the simultaneous determination of methamphetamine (MA) and amphetamine (AM) in urine using a high performance liquid chromatography (HPLC) column-switching method. A PK-2A (Shodex) column was used for extraction and deproteinization, and a CAPCELL PAK SCX semi-micro, polymer-coated cation-exchange column was employed for separation. The urine sample was mixed with an equal volume of borate buffer (0.1M, pH 9.4), and then 100 microl of the mixture was injected into the HPLC column. The column was switched for 6 min, and then 10 min later detection was performed at 210 nm. Recovery yields of the MA and AM spiked in the urine were 93.0-100.4% with a coefficient of variation of less than 1%. The calibration curves of MA and AM were in the range of 0.1-10 microg/ml with good linearity (r(2)=0.999), with the limit of qualification being 0.005 microg/ml. This method of using HPLC with column-switching can be used for both qualification and quantification of MA and its metabolite, AM, in urine, especially in forensic cases.

  19. Methods for analysis of size-exclusion chromatography-small-angle X-ray scattering and reconstruction of protein scattering.

    PubMed

    Malaby, Andrew W; Chakravarthy, Srinivas; Irving, Thomas C; Kathuria, Sagar V; Bilsel, Osman; Lambright, David G

    2015-08-01

    Size-exclusion chromatography in line with small-angle X-ray scattering (SEC-SAXS) has emerged as an important method for investigation of heterogeneous and self-associating systems, but presents specific challenges for data processing including buffer subtraction and analysis of overlapping peaks. This paper presents novel methods based on singular value decomposition (SVD) and Guinier-optimized linear combination (LC) to facilitate analysis of SEC-SAXS data sets and high-quality reconstruction of protein scattering directly from peak regions. It is shown that Guinier-optimized buffer subtraction can reduce common subtraction artifacts and that Guinier-optimized linear combination of significant SVD basis components improves signal-to-noise and allows reconstruction of protein scattering, even in the absence of matching buffer regions. In test cases with conventional SAXS data sets for cytochrome c and SEC-SAXS data sets for the small GTPase Arf6 and the Arf GTPase exchange factors Grp1 and cytohesin-1, SVD-LC consistently provided higher quality reconstruction of protein scattering than either direct or Guinier-optimized buffer subtraction. These methods have been implemented in the context of a Python-extensible Mac OS X application known as Data Evaluation and Likelihood Analysis (DELA), which provides convenient tools for data-set selection, beam intensity normalization, SVD, and other relevant processing and analytical procedures, as well as automated Python scripts for common SAXS analyses and Guinier-optimized reconstruction of protein scattering.

  20. An improved method for cyanide determination in blood using solid-phase microextraction and gas chromatography/mass spectrometry.

    PubMed

    Frison, Giampietro; Zancanaro, Flavio; Favretto, Donata; Ferrara, Santo Davide

    2006-01-01

    A new method is described for the qualitative and quantitative analysis of cyanide, a very short-acting and powerful toxic agent, in human whole blood. It involves the conversion of cyanide into hydrogen cyanide and its subsequent headspace solid-phase microextraction (HS-SPME) and detection by gas chromatography/mass spectrometry (GC/MS) in selected ion monitoring (SIM) mode. Optimizing the conditions for the GC/MS (type of column, injection conditions, temperature program) and SPME (choice of SPME fiber, effect of salts, adsorption and desorption times, adsorption temperature) led to the choice of a 75-microm carboxen/polydimethylsiloxane SPME fiber, with D3-acetonitrile as internal standard, and a capillary GC column with a polar stationary phase. Method validation was carried out in terms of linearity, precision and accuracy in both aqueous solutions and blood. The limit of detection (LOD) and limit of quantitation (LOQ) were determined only in aqueous solutions. The assay is linear over three orders of magnitude (water 0.01-10, blood 0.05-10 microg/mL); and the LOD and LOQ in water were 0.006 and 0.01 microg/mL, respectively. Good intra- and inter-assay precision was obtained, always <8%. The method is simple, fast and sensitive enough for the rapid diagnosis of cyanide intoxication in clinical and forensic toxicology.

  1. Development and validation of stability indicating method for the determination of exemestane by reverse phase high performance liquid chromatography.

    PubMed

    Konda, Bharath; Tiwari, Ravi N; Fegade, Harshal

    2011-09-01

    Exemestane is an aromatase inhibitor used in the treatment of breast cancer. A selective stability-indicating reversed-phase high performance liquid chromatography (RP-HPLC) method has been developed which can separate and accurately quantitate low levels of exemestane. The stability-indicating capability of the method was demonstrated by adequate separation of exemestane and all the degradation product peaks from exemestane peak and also from each other in stability samples of exemestane. Chromatographic separation of exemestane and its degraded products were achieved by using isocratic elution at a flow rate of 1.0 mL/min on a C18 reverse phase column (Phenomenex, size: 250 × 4.60 mm, particle size 5 μm) at ambient temperature. The mobile phase used for the analysis was acetonitrile-water (60:40, %v/v) with UV visible detection at 242 nm. The proposed method was used to study the degradation behavior of drug under various stress conditions as per ICH recommended guidelines.

  2. A reversed-phase high performance liquid chromatography method for quantification of methotrexate in cancer patients serum.

    PubMed

    Li, Yuan-dong; Li, Yan; Liang, Ning-sheng; Yang, Fan; Kuang, Zhi-peng

    2015-10-01

    A simple, rapid and sensitive reversed-phase high performance liquid chromatography (HPLC) method has been developed for the determination of methotrexate in human serum. After deproteinization of the serum with 40% silver nitrate solution, methotrexate and internal standard (IS) were separated on a reversed-phase column with a mobile phase consisting of 10mM sodium phosphate buffer (pH6.40)-methanol (78:22%, v/v) and ultraviolet detection at 310nm. The linearity is evaluated by a calibration curve in the concentration range of 0.05-10.0μg/mL and presented a correlation coefficient of 0.9995. The absolute recoveries were 97.52±3.9% and 96.87±3.7% for methotrexate and ferulic acid (internal standard), respectively. The intra- and inter-day precision were less 6.19 and 5.89%, respectively (n=6). The limit of quantitation was 0.02μg/mL and the limit of detection was 0.006μg/mL. The complete analysis was achieved less than 10min with no interference from endogenous components or 22 examined drugs. This method was validated by using serum samples from high-dose methotrexate treated patients with osteosarcoma, breast cancer, acute leukemia and lymphoma. The method was demonstrated to be a simple, rapid and reliable approach in quantification of methotrexate in serum samples from patients with high-dose methotrexate therapy.

  3. Simple column-switching ion chromatography method for determining eight monosaccharides and oligosaccharides in honeydew and nectar.

    PubMed

    Ni, Chengzhu; Zhu, Binhe; Wang, Nani; Wang, Muhua; Chen, Suqing; Zhang, Jiajie; Zhu, Yan

    2016-03-01

    Honeydew is excreted by aphids as a sweet waste and nectar is floral honey. Honeydew and nectar are complicated samples which consist of various sugars and amino acids. In this work, a simple ion chromatography with column-switching method was developed for the simultaneous analysis of 8 monosaccharides and oligosaccharides in honeydew and nectar. A reversed-phase column was used as a pretreatment column to eliminate organics on-line and sugars were eluted from a collection loop to analytical column by using column-switching technique. This method showed good linearity (r⩾0.9994) and afforded low limits of detection ranging from 1.55 to 10.17μgL(-1) for all the analytes. Recoveries ranged from 95% to 105% and repeatability results were acceptable with relative standard deviation of less than 3.21% (n=6). This method was successfully applied to quantification of these sugars in honeydew and nectar. These results showed honeydew had much more oligosaccharides than nectar.

  4. Solid phase microextraction and gas chromatography-mass spectrometry methods for residual solvent assessment in seized cocaine and heroin.

    PubMed

    Cabarcos, Pamela; Herbello-Hermelo, Paloma; Álvarez-Freire, Iván; Moreda-Piñeiro, Antonio; Tabernero, María Jesús; Bermejo, Ana María; Bermejo-Barrera, Pilar

    2016-09-01

    A simple sample pre-treatment method based on solid phase microextraction (SPME) and gas chromatography-mass spectrometry (GC-MS) has been optimized and validated for the assessment of 15 residual solvents (2-propanol, 2-methylpentane, 3-methylpentane, acetone, ethyl acetate, benzene, hexane, methylcyclohexane, methylcyclopentane, m-xylene, propyl acetate, toluene, 1,2,4-trimethylbenzene, dichloromethane, and ethylbenzene) in seized illicit cocaine and heroin. DMSO and DMF as sample diluents were found to offer the best residual solvent transference to the head space for further adsorption onto the SPME fiber, and the developed method therefore showed high sensitivity and analytical recovery. Variables affecting SPME were fully evaluated by applying an experimental design approach. Best conditions were found when using an equilibration time of 5 min at 70 °C and headspace sampling of residual solvents at the same temperature for 15 min. Method validation, performed within the requirements of international guidelines, showed excellent sensitivity, as well as intra- and inter-day precision and accuracy. The proposed methodology was applied to 96 cocaine samples and 14 heroin samples seized in Galicia (northwestern Spain) within 2013 and 2014.

  5. Improved method for the quantification of lysophospholipids including enol ether species by liquid chromatography-tandem mass spectrometry.

    PubMed

    Bollinger, James G; Ii, Hiromi; Sadilek, Martin; Gelb, Michael H

    2010-02-01

    LC/ESI-MS/MS has been previously demonstrated to be a powerful method to detect and quantify molecular species of glycerophospholipids including lysophospholipids. In this study, we provide an improved pre-mass spectrometry lipid extraction procedure that avoids the acid-catalyzed decomposition of plasmenyl phospholipids that is problematic with previously reported methods. We show that the use of lysophospholipid internal standards with perdeuterated fatty acyl chains avoids isobar problems associated with the use of internal standards containing odd carbon number fatty acyl chains. We also show that LC prior to MS is required to avoid numerous problems associated with isobars and with MS in-source decomposition of lysophosphatidylserine. The reported method of using normal phase chromatography/ESI-MS is used to quantify lysophospholipids in serum and to quantify lysophospholipids produced in mammalian cells by human group X secreted phospholipase A(2). The latter shows that group X phospholipase A(2) added exogenously to cells generates a different set of lysophospholipids compared with enzyme produced endogenously in cells, which supports earlier studies showing that this phospholipase A(2) can act on cell membranes prior to externalization from cells.

  6. Ultra-performance liquid chromatography-tandem mass spectrometry method for the analysis of amphetamines in plasma.

    PubMed

    Fernández, María del Mar Ramírez; Samyn, Nele

    2011-10-01

    A fast and selective ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS-MS) method for the determination of amphetamines (amphetamine, methamphetamine, methylenedioxymethamphetamine, methylenedioxyamphetamine, methylenedioxyethylamphetamine, ephedrine, and p-methoxyamphetamine) in plasma has been developed and validated. Sample preparation was performed by liquid-liquid extraction using ethyl acetate. For optimized chromatographic performance with repeatable retention times, narrow and symmetrical peaks, and focusing all analytes at the column inlet, a gradient start, with acid mobile phase consisting of 0.1% formic acid and methanol was chosen. Positive electrospray ionization MS-MS detection was performed with two multiple reaction monitoring transitions for each analyte. Deuteriumlabeled internal standards were used for five of the analytes. The limit of detection was in the range 0.25-1.25 ng/mL, and the limit of quantification was fixed at the lowest calibrator of 2.5 ng/mL for all of the compounds. The RSD values of the intra- and interassay precision and accuracy were lower than 11% at four concentration levels, including two external quality controls. No or only minor matrix effects were observed, and the extraction method presented recoveries higher than 93% for all the compounds. Total run time, including equilibration, was 12 min. The method is routinely used at the National Institute of Criminalistics and Criminology for quantitative determination of the main amphetamines in plasma from forensic and driving under the influence cases.

  7. Magnetic solid phase extraction and static headspace gas chromatography-mass spectrometry method for the analysis of polycyclic aromatic hydrocarbons.

    PubMed

    Cai, Ying; Yan, Zhihong; Wang, Lijia; NguyenVan, Manh; Cai, Qingyun

    2016-01-15

    A magnetic solid phase extraction (MSPE) protocol combining a static headspace gas chromatography coupled to mass spectrometry (HS-GC-MS) method has been developed for extraction, and determination of 16 polycyclic aromatic hydrocarbons (PAHs) in drinking water samples. Magnetic nanoparticles (MNPs) were coated with 3-aminopropyltriethoxysilane and modified by cholesterol chloroformate. Transmission electron microscope, vibrating sample magnetometer, Fourier transform infrared spectrometry and X-ray photoelectron spectroscopy were used to characterize the cholesterol-functionalized sorbents, and the main parameters affecting the extraction as well as HS sampling, such as sorbent amount, extraction time, oven temperature and equilibration time have been investigated and established. Combination with HS sampling, the MSPE procedure was simple, fast and environmentally friendly, without need of any organic solvent. Method validation proved the feasibility of the developed sorbents for the quantitation of the investigated analytes at trace levels obtaining the limit of detection (S/N=3) ranging from 0.20 to 7.8 ng/L. Good values for intra and inter-day precision were obtained (RSDs ≤ 9.9%). The proposed method was successfully applied to drinking water samples.

  8. An alternative derivatization method for the analysis of amino acids in cerebrospinal fluid by gas chromatography-mass spectrometry.

    PubMed

    de Paiva, Maria José Nunes; Menezes, Helvécio Costa; Christo, Paulo Pereira; Resende, Rodrigo Ribeiro; Cardeal, Zenilda de Lourdes

    2013-07-15

    The determination of the concentrations of l-amino acids in cerebrospinal fluid (CSF) has been used to gain biochemical insight into central nervous system disorders. This paper describes a microwave-assisted derivatization (MAD) method using N,O-bis-(trimethylsilyl)trifluoroacetamide (BSTFA) as a derivatizing agent for determining the concentrations of l-amino acids in human CSF by gas chromatography with mass spectrometry (GC/MS). The experimental design used to optimize the conditions showed that the optimal derivatization time was 3min with a microwave power of 210W. The method showed good performance for the validation parameters. The sensitivity was very good, with limits of detection (LODs) ranging from 0.01μmolL(-1) to 4.24μmolL(-1) and limits of quantification (LOQs) ranging from 0.02 to 7.07μmolL(-1). The precision, measured using the relative standard deviation (RSD), ranged from 4.12 to 15.59% for intra-day analyses and from 6.36 to 18.71% for inter-day analyses. The coefficients of determination (R(2)) were above 0.990 for all amino acids. The optimized and validated method was applied to the determination of amino acid concentrations in human CSF.

  9. A liquid chromatography-tandem mass spectrometry method for the detection of economically motivated adulteration in protein-containing foods.

    PubMed

    MacMahon, Shaun; Begley, Timothy H; Diachenko, Gregory W; Stromgren, Selen A

    2012-01-13

    A new analytical method was developed to determine the presence of six (6) compounds with the potential to be used in economic adulteration to enhance the nitrogen content in milk products and bulk proteins. Residues were extracted from the matrix with 2% formic acid, after which acetonitrile (ACN) was added to induce precipitation of the proteins. Extracts were analyzed by liquid chromatography using a ZIC-HILIC column with tandem mass spectrometry (LC-MS/MS) using electrospray ionization (ESI). Single-laboratory method validation data was collected in six matrices fortified at concentrations down to 1.0 μg/g (ppm). Average recoveries and average relative standard deviations (RSD) using spiked matrix calibration standard curves were the following: cyromazine (CY) 95.9% (7.5% RSD), dicyandiamide (DC) 98.1% (5.6% RSD), urea 102.5% (8.6% RSD), biuret (BU) 97.2% (6.6% RSD), triuret (TU) 97.7% (5.7% RSD), and amidinourea (AU) 93.4% (7.4% RSD). This method provides a rapid and effective approach to proactively combat economically motivated adulteration in protein-containing products.

  10. Validation of a pH gradient-based ion-exchange chromatography method for high-resolution monoclonal antibody charge variant separations.

    PubMed

    Rea, Jennifer C; Moreno, G Tony; Lou, Yun; Farnan, Dell

    2011-01-25

    Ion-exchange chromatography is widely used for profiling the charge heterogeneity of proteins, including monoclonal antibodies. Despite good resolving power and robustness, ionic strength-based ion-exchange separations are product-specific and time-consuming to develop. We have previously reported a novel pH-based separation of proteins by cation exchange chromatography that was multi-product, high-resolution, and robust against variations in sample matrix salt concentration and pH. In this study, a pH gradient-based separation method using cation exchange chromatography was evaluated in a mock validation. This method was shown to be robust for monoclonal antibodies and suitable for its intended purpose of charge heterogeneity analysis. Simple mixtures of defined buffer components were used to generate the pH gradients that separated closely related antibody species. Validation characteristics, such as precision and linearity, were evaluated. Robustness to changes in protein load, buffer pH and column oven temperature was demonstrated. The stability-indicating capability of this method was determined using thermally stressed antibody samples. In addition, intermediate precision was demonstrated using multiple instruments, multiple analysts, multiple column lots, and different column manufacturers. Finally, the precision for this method was compared to conventional ion-exchange chromatography and imaged capillary isoelectric focusing. These results demonstrate the superior precision and robustness of this multi-product method, which can be used for the high-throughput evaluation of in-process and final product samples.

  11. Critical assessment of three high performance liquid chromatography analytical methods for food carotenoid quantification.

    PubMed

    Dias, M Graça; Oliveira, Luísa; Camões, M Filomena G F C; Nunes, Baltazar; Versloot, Pieter; Hulshof, Paul J M

    2010-05-21

    Three sets of extraction/saponification/HPLC conditions for food carotenoid quantification were technically and economically compared. Samples were analysed for carotenoids alpha-carotene, beta-carotene, beta-cryptoxanthin, lutein, lycopene, and zeaxanthin. All methods demonstrated good performance in the analysis of a composite food standard reference material for the analytes they are applicable to. Methods using two serial connected C(18) columns and a mobile phase based on acetonitrile, achieved a better carotenoid separation than the method using a mobile phase based on methanol and one C(18)-column. Carotenoids from leafy green vegetable matrices appeared to be better extracted with a mixture of methanol and tetrahydrofuran than with tetrahydrofuran alone. Costs of carotenoid determination in foods were lower for the method with mobile phase based on methanol. However for some food matrices and in the case of E-Z isomer separations, this was not technically satisfactory. Food extraction with methanol and tetrahydrofuran with direct evaporation of these solvents, and saponification (when needed) using pyrogallol as antioxidant, combined with a HPLC system using a slight gradient mobile phase based on acetonitrile and a stationary phase composed by two serial connected C(18) columns was the most technically and economically favourable method.

  12. Direct liquid chromatography method for the simultaneous quantification of hydroxytyrosol and tyrosol in red wines.

    PubMed

    Piñeiro, Zulema; Cantos-Villar, Emma; Palma, Miguel; Puertas, Belen

    2011-11-09

    A validated HPLC method with fluorescence detection for the simultaneous quantification of hydroxytyrosol and tyrosol in red wines is described. Detection conditions for both compounds were optimized (excitation at 279 and 278 and emission at 631 and 598 nm for hydroxytyrosol and tyrosol, respectively). The validation of the analytical method was based on selectivity, linearity, robustness, detection and quantification limits, repeatability, and recovery. The detection and quantification limits in red wines were set at 0.023 and 0.076 mg L(-1) for hydroxytyrosol and at 0.007 and 0.024 mg L(-1) for tyrosol determination, respectively. Precision values, both within-day and between-day (n = 5), remained below 3% for both compounds. In addition, a fractional factorial experimental design was developed to analyze the influence of six different conditions on analysis. The final optimized HPLC-fluorescence method allowed the analysis of 30 nonpretreated Spanish red wines to evaluate their hydroxytyrosol and tyrosol contents.

  13. A robust method for determining water-extractable alkylphenol polyethoxylates in textile products by reaction-based headspace gas chromatography.

    PubMed

    Zhang, Shu-Xin; Chai, Xin-Sheng; Huang, Bo-Xi; Mai, Xiao-Xia

    2015-08-07

    Alkylphenol polyethoxylates (APEO), surfactants used in the production of textiles, have the potential to move from the fabric to the skin of the person wearing the clothes, posing an inherent risk of adverse health consequences. Therefore, the textile industry needs a fast, robust method for determining aqueous extractable APEO in fabrics. The currently-favored HPLC methods are limited by the presence of a mixture of analytes (due to the molecular weight distribution) and a lack of analytical standards for quantifying results. As a result, it has not been possible to reach consensus on a standard method for the determination of APEO in textiles. This paper addresses these limitations through the use of reaction-based head space-gas chromatography (HS-GC). Specifically, water is used to simulate body sweat and extract APEO. HI is then used to react the ethoxylate chains to depolymerize the chains into iodoethane that is quantified through HS-GC, providing an estimate of the average amount of APEO in the clothing. Data are presented to justify the optimal operating conditions; i.e., water extraction at 60°C for 1h and reaction with a specified amount of HI in the headspace vial at 135°C for 4h. The results show that the HS-GC method has good precision (RSD<10%) and good accuracy (recoveries from 95 to 106%) for the quantification of APEO content in textile and related materials. As such, the method should be a strong candidate to become a standard method for such determinations.

  14. Method development for Lawsone estimation in Trichup herbal hair powder by high-performance thin layer chromatography.

    PubMed

    Patel, Maunang M; Solanki, Bhavna R; Gurav, Nilesh C; Patel, Prateek H; Verma, Shweta S

    2013-07-01

    A simple, specific, accurate, precise and robust high-performance thin-layer chromatographic method has been developed and validated for estimation of Lawsone in Trichup herbal hair powder (coded as a THHP), polyherbal formulation. The chromatographic development was carried out on aluminum plates pre-coated with silica gel 60F254 and good resolution was achieved with Toluene: Ethyl acetate: Glacial acetic acid (8:1:1 v/v/v) as mobile phase. Lawsone detection was carried out densitometrically at 277 nm and obtained retardation factor value was 0.46 ± 0.02. The method was validated with respect to specificity, linearity, accuracy, precision and robustness. The calibration curve was achieved to be linear over a range of 5-60 μg/ml and regression coefficient was obtained 0.998. Accuracy of chromatographic method was evaluated by standard addition method; recovery was obtained 99.25 ± 0.61%. The peak purity of Lawsone was achieved 0.999 r. Relative standard deviation for intraday and inter-day precision was 0.37-0.56% and 0.42-0.55%, respectively. The limit of detection and limit of quantification of the Lawsone were found to be 1.08 μg/m land 3.28 μg/ml, respectively. This result shows that the method was well validated. In the present study, the Lawsone content was found 0.322 ± 0.014% in THHP. This study reveals that the proposed high performance thin layer chromatography method is accurate, fast and cost- effective for routine estimation of Lawsone in polyherbal formulation.

  15. METHOD FOR THE DETERMINATION OF PERCHLORATE ANION IN PLANT AND SOLID MATRICES BY ION CHROMATOGRAPHY

    EPA Science Inventory

    A standardized method for the analysis of perchlorate in plants was developed, based on dry weight, and applied to the analysis of plant organs, foodstuffs, and plant products. The procedure greatly reduced the ionic interferences in water extracts of plant materials. Ion chro...

  16. Application of liquid chromatography method with electrochemical detection for bioequivalence study of trimetazidine in human plasma.

    PubMed

    Grabowski, Tomasz; Swierczewska, Anna; Borucka, Beata; Sawicka, Renata; Sasinowska-Motyl, Małgorzata; Gumułka, Stanisław Witold

    2012-01-01

    A method to estimate trimetazidine (CAS: 13171-25-0) levels in human plasma by means of HPLC with electrochemical detection was developed. Trimethoprim (CAS: 26807-65-8) was used as an internal standard. This method of analysis was fully validated according to the guidelines of the United States Food and Drug Administration, European Medicines Agency and Organization for Economic Co-operation and Development and Good Laboratory Practice rules. The accuracy and precision of the developed method were found to be satisfactory and stability studies showed acceptable variation (below 15%) of trimetazidine concentrations when samples were stored frozen at -75 degrees C for 54 days. The developed method was successfully used for a comparative 2 x 2 period, crossover bioequivalence study of two extended-release preparations of trimetazidine performed on 24 healthy volunteers at the steady state after multiple dosing of 35 mg twice daily for 4 days and a single 35 mg dose on the 5th day and after a single dose of 35 mg under fasting or postprandial conditions.

  17. Matrix elimination ion chromatography method for trace level azide determination in irbesartan drug.

    PubMed

    Subramanian, Narayanan Harihara; Babu, V R Sankar; Jeevan, R Ganesh; Radhakrishnan, Ganga

    2009-08-01

    Ultra-trace analysis of azide in complicated Irbesartan sample matrix is achieved by the in-line sample preparation technique. Sodium azide is the precursor of Irbesartan, which is used as an anti-hypertensive drug. Due to the toxic nature of sodium azide, reliable determination of azide in Irbesartan is necessary. Irbesartan when analyzed for sodium azide, as per the USP 31-NF26 method, gets adsorbed to the analytical column, leading to reduction in column capacity and irreproducible retention time. The retained drug has to be removed with special rinsing solution, followed by re-equilibration with the mobile phase. This process takes at least 3 to 4 h for each sample analysis. The new method developed overcomes the limitations of the USP 31-NF26 method. This method is validated for specificity, linearity, accuracy, precision, sample solution stability, and robustness as per International Conference on Harmonization guidelines. The relationship between peak response and concentration is found to be linear between 5 to 80 ng/mL of sodium azide, with the correlation coefficient (r(2)) of 0.9995. The limits of detection and quantification for sodium azide are 0.532 and 1.61 microg/gm with respect to the sample weight.

  18. Simple Method for Assaying Colistin Methanesulfonate in Plasma and Urine Using High-Performance Liquid Chromatography

    PubMed Central

    Li, Jian; Milne, Robert W.; Nation, Roger L.; Turnidge, John D.; Coulthard, Kingsley; Valentine, Jason

    2002-01-01

    A simple and sensitive high-performance liquid chromatographic method is described for the determination of colistimethate sodium in plasma and urine. The accuracy and reproducibility was within 10.1 and 11.2% with rat plasma and urine, respectively. Several commonly coadministered antibacterial agents do not interfere with the assay. PMID:12234867

  19. Comparison of ultra-high performance methods in liquid and supercritical fluid chromatography coupled to electrospray ionization - mass spectrometry for impurity profiling of drug candidates.

    PubMed

    Lemasson, Elise; Bertin, Sophie; Hennig, Philippe; Lesellier, Eric; West, Caroline

    2016-11-11

    Impurity profiling of organic products synthesized as possible drug candidates represents a major analytical challenge. Complementary analytical methods are required to ensure that all impurities are detected. Both high-performance liquid chromatography (HPLC) and supercritical fluid chromatography (SFC) can be used for this purpose. In this study, we compared ultra-high performance HPLC (UHPLC) and ultra-high performance SFC (UHPSFC) using a large dataset of 140 pharmaceutical compounds. Four previously optimized methods (two on each technique) were selected to ensure fast high-resolution separations. The four methods were evaluated based on response rate, peak capacity, peak shape and capability to detect impurities (UV). The orthogonality between all methods was also assessed. The best UHPLC method and UHPSFC methods provided comparable quality for the 140 compounds included in this study. Moreover, they were found to be highly orthogonal. At last, the potential of the combined use of UHPLC and UHPSFC for impurity profiling is illustrated with practical examples.

  20. Size exclusion chromatography with evaporative light scattering detection as a method for speciation analysis of polydimethylsiloxanes. II. Validation of the method for analysis of pharmaceutical formulations.

    PubMed

    Mojsiewicz-Pieńkowska, Krystyna

    2011-12-05

    The aim of this study was to demonstrate the usefulness of the size exclusion chromatography with evaporative light scattering detection (SEC-ELSD) method in the identification and quantitative analysis of polydimethylsiloxanes (PDMS). The process of validation for the method was conducted, and the values obtained were compared with the acceptance criteria. Particularly important was the conclusion that SEC-ELSD method showed a high specificity for PDMS. PDMS is an organosilicon polymer and for this reason, it does not exist as a concrete chemical species. Depending on the length of the chain, PDMS can be toxic for organism. So far, the SEC-ELSD method has not been applied for the control of pharmaceutical products containing such PDMS as dimeticone or simeticone. The safety of use and effectiveness of such pharmaceutical products relies on the control of their quality. Therefore, the analytical methods and procedures that meet acceptance criteria for qualitative and quantitative analysis of the PDMS should be used. In the case of the analysis of pharmaceutical products, the acceptance criteria are established and recommended by, for example, the Pharmacopoeias, the U.S. Food and Drug Administration (FDA), the International Conference on Harmonisation (ICH) and the World Health Organization (WHO). The progress of knowledge, however, requires the development of new analytical tools which are able to solve incoming problems. In the case of pharmaceutical formulations containing PDMS, which are used not only by adults but also by children, it is necessary to use analytical methods which are characterized by a high specificity.

  1. Comparison of sample preparation methods for reliable plutonium and neptunium urinalysis using automatic extraction chromatography.

    PubMed

    Qiao, Jixin; Xu, Yihong; Hou, Xiaolin; Miró, Manuel

    2014-10-01

    This paper describes improvement and comparison of analytical methods for simultaneous determination of trace-level plutonium and neptunium in urine samples by inductively coupled plasma mass spectrometry (ICP-MS). Four sample pre-concentration techniques, including calcium phosphate, iron hydroxide and manganese dioxide co-precipitation and evaporation were compared and the applicability of different techniques was discussed in order to evaluate and establish the optimal method for in vivo radioassay program. The analytical results indicate that the various sample pre-concentration approaches afford dissimilar method performances and care should be taken for specific experimental parameters for improving chemical yields. The best analytical performances in terms of turnaround time (6h) and chemical yields for plutonium (88.7 ± 11.6%) and neptunium (94.2 ± 2.0%) were achieved by manganese dioxide co-precipitation. The need of drying ashing (≥ 7h) for calcium phosphate co-precipitation and long-term aging (5d) for iron hydroxide co-precipitation, respectively, rendered time-consuming analytical protocols. Despite the fact that evaporation is also somewhat time-consuming (1.5d), it endows urinalysis methods with better reliability and repeatability compared with co-precipitation techniques. In view of the applicability of different pre-concentration techniques proposed previously in the literature, the main challenge behind relevant method development is pointed to be the release of plutonium and neptunium associated with organic compounds in real urine assays. In this work, different protocols for decomposing organic matter in urine were investigated, of which potassium persulfate (K2S2O8) treatment provided the highest chemical yield of neptunium in the iron hydroxide co-precipitation step, yet, the occurrence of sulfur compounds in the processed sample deteriorated the analytical performance of the ensuing extraction chromatographic separation with chemical

  2. Ruggedness testing and validation of a practical analytical method for > 100 veterinary drug residues in bovine muscle by ultrahigh performance liquid chromatography – tandem mass spectrometry

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In this study, optimization, extension, and validation of a streamlined, qualitative and quantitative multiclass, multiresidue method was conducted to monitor great than100 veterinary drug residues in meat using ultrahigh-performance liquid chromatography – tandem mass spectrometry (UHPLC-MS/MS). I...

  3. A NEW SW-846 METHOD FOR THE ANALYSIS OF TOXAPHENE AND TOXAPHENE CONGENERS IN SOLID AND AQUEOUS SAMPLES USING GAS CHROMATOGRAPHY / NEGATIVE ION MASS SPECTROMETRY

    EPA Science Inventory

    US EPA SW-846 methods have typically relied on dual column gas chromatography coupled with electron capture detection (GC-ECD) for analysis of low concentrations of organochlorine pesticides, including toxaphene, in environmental samples. Toxaphene is one of the most widely appl...

  4. METHOD 530 DETERMINATION OF SELECT SEMIVOLATILE ORGANIC CHEMICALS IN DRINKING WATER BY SOLID PHASE EXTRACTION AND GAS CHROMATOGRAPHY/ MASS SPECTROMETRY (GC/MS)

    EPA Science Inventory

    1.1. This is a gas chromatography/mass spectrometry (GC/MS) method for the determination of selected semivolatile organic compounds in drinking waters. Accuracy and precision data have been generated in reagent water, and in finished ground and surface waters for the compounds li...

  5. A method for determination of selenium in organic tissues using microwave digestion and liquid chromatography.

    PubMed

    Govasmark, Espen; Grimmett, Mark G

    2007-01-01

    An existing laboratory procedure for selenium analysis using open-vessel wet digestion and liquid chromatographic fluorescence determination was modified for use with microwave digestion. The proposed microwave digestion method eliminated the hazards associated with the use of HClO4 while maintaining excellent recoveries of selenium. A 2-step HNO3/H2O2 digestion procedure was developed. Digested samples were derivatized with 2,3-diaminonaphthalene, and the resultant piazselenol complex was measured fluorometrically using a liquid chromatograph. Measured values were in agreement with 9 different certified reference materials. The detection limit for this method was 0.54 ng Se/g tissue (3 sigma), and the calibration curve remained linear (r2 = 0.9968) up to 2 microg Se/g.

  6. Development and validation of a liquid chromatography-tandem mass spectrometry method for the quantification of opiorphin in human saliva.

    PubMed

    Brkljačić, Lidija; Sabalić, Maja; Salarić, Ivan; Jerić, Ivanka; Alajbeg, Ivan; Nemet, Ina

    2011-12-15

    Opiorphin, QRFSR-peptide, is a mature product of the PROL1 (proline rich, lacrimal 1) protein that showed beneficial effects in pain management, antidepressant-like actions as well as involvement in colonic motility and erectile physiology. Using opiorphin as a potential biomarker of different pathological states requires the development of robust and sensitive methods. We report a highly sensitive and specific liquid chromatography with tandem mass spectrometric detection (LC-MS/MS) analytical method for the analysis of opiorphin in human saliva. Quantification was based on multiple reaction monitoring using characteristic transitions (m/z 347/120 - as quantifying ion; 347/175 and 347/268 as qualifying ions). The assay was linear in the range of 0-110 ng/ml and the lower limit of quantification reached was 1.0 ng/ml. The intra-day precision and accuracy were between 2.7-5.6% and -2.3 to 3.2%, respectively. The inter-day precision and accuracy were between 10.8-13.7% and -11.0 to 52%, respectively. Mean recovery was 106% and mean matrix effect was 0.97. Opiorphin in TFA treated saliva samples was stable for at least 12h at room temperature and up to 30 days at -20°C. Opiorphin levels in human saliva samples collected from young healthy individuals ranged from 2.8 to 25.9 ng/ml.

  7. [Development of a gas chromatography-mass spectrometry method for the metabolomic study of rice (Oryza sativa L.) grain].

    PubMed

    Zhou, Jia; Wang, Shuangyuan; Chang, Yuwei; Zhao, Yanni; Lu, Xin; Zhao, Chunxia; Xu, Guowang

    2012-10-01

    An analytical strategy for the metabolic profiling of rice grain was developed based on gas chromatography-mass spectrometry (GC-MS). For the purpose of obtaining abundant metabolite information, sample preparation step prior to instrumental analysis is necessary to be optimized. D-optimal experimental design was applied to optimize the extraction solvent. Four solvents, including water, methanol, isopropanol and acetonitrile, and their combinations were evaluated for the extraction efficiency using multivariate statistical analysis (partial least square regression). The count of resolved peaks and the sum of peak areas were taken as the evaluation indexes. Methanol/water (80:20, v/v) mixture was highly efficient for rice metabolites and was selected as the suitable solvent formulation. Then, the analytical characteristics of the method were measured. More than 90% of the metabolites had satisfactory precisions, reproducibilities and stabilities (relative standard deviations (RSDs) < 30%). Most of the detected metabolites (about 88.0% of total peak area) showed good linear responses. With the optimized analytical protocol, 315 metabolites were detected in rice and 86 of which were structurally identified by searching in the NIST 08/Wiley standard mass spectral library, covering carbohydrates, amino acids, organic acids, steroids and so on which showed a broad coverage of metabolite data. The established method is expected to be useful for the metabolomic studies of rice.

  8. Investigation of calcium antagonist-L-type calcium channel interactions by a vascular smooth muscle cell membrane chromatography method.

    PubMed

    Du, Hui; He, Jianyu; Wang, Sicen; He, Langchong

    2010-07-01

    The dissociation equilibrium constant (K(D)) is an important affinity parameter for studying drug-receptor interactions. A vascular smooth muscle (VSM) cell membrane chromatography (CMC) method was developed for determination of the K(D) values for calcium antagonist-L-type calcium channel (L-CC) interactions. VSM cells, by means of primary culture with rat thoracic aortas, were used for preparation of the cell membrane stationary phase in the VSM/CMC model. All measurements were performed with spectrophotometric detection (237 nm) at 37 degrees C. The K(D) values obtained using frontal analysis were 3.36 x 10(-6) M for nifedipine, 1.34 x 10(-6) M for nimodipine, 6.83 x 10(-7) M for nitrendipine, 1.23 x 10(-7) M for nicardipine, 1.09 x 10(-7) M for amlodipine, and 8.51 x 10(-8) M for verapamil. This affinity rank order obtained from the VSM/CMC method had a strong positive correlation with that obtained from radioligand binding assay. The location of the binding region was examined by displacement experiments using nitrendipine as a mobile-phase additive. It was found that verapamil occupied a class of binding sites on L-CCs different from those occupied by nitrendipine. In addition, nicardipine, amlodipine, and nitrendipine had direct competition at a single common binding site. The studies showed that CMC can be applied to the investigation of drug-receptor interactions.

  9. A High-Throughput Size Exclusion Chromatography Method to Determine the Molecular Size Distribution of Meningococcal Polysaccharide Vaccine

    PubMed Central

    Khan, Imran; Rahman, K. M. Taufiqur; Siraj, S. M. Saad Us; Karim, Mahbubul; Muktadir, Abdul; Maheshwari, Arpan; Kabir, Md Azizul; Nahar, Zebun

    2016-01-01

    Molecular size distribution of meningococcal polysaccharide vaccine is a readily identifiable parameter that directly correlates with the immunogenicity. In this paper, we report a size exclusion chromatography method to determine the molecular size distribution and distribution coefficient value of meningococcal polysaccharide serogroups A, C, W, and Y in meningococcal polysaccharide (ACWY) vaccines. The analyses were performed on a XK16/70 column packed with sepharose CL-4B with six different batches of Ingovax® ACWY, a meningococcal polysaccharide vaccine produced by Incepta Vaccine Ltd., Bangladesh. A quantitative rocket immunoelectrophoresis assay was employed to determine the polysaccharide contents of each serogroup. The calculated distribution coefficient values of serogroups A, C, W, and Y were found to be 0.26 ± 0.16, 0.21 ± 0.11, 0.21 ± 0.11, and 0.14 ± 0.12, respectively, and met the requirements of British Pharmacopeia. The method was proved to be robust for determining the distribution coefficient values which is an obligatory requirement for vaccine lot release. PMID:27688770

  10. Repeatability of gradient ultrahigh pressure liquid chromatography-tandem mass spectrometry methods in instrument-controlled thermal environments.

    PubMed

    Grinias, James P; Wong, Jenny-Marie T; Kennedy, Robert T

    2016-08-26

    The impact of viscous friction on eluent temperature and column efficiency in liquid chromatography is of renewed interest as the need for pressures exceeding 1000bar to use with columns packed with sub-2μm particles has grown. One way the development of axial and radial temperature gradients that arise due to viscous friction can be affected is by the thermal environment the column is placed in. In this study, a new column oven integrated into an ultrahigh pressure liquid chromatograph that enables both still-air and forced-air operating modes is investigated to find the magnitude of the effect of the axial thermal gradient that forms in 2.1×100mm columns packed with sub-2μm particles in these modes. Temperature increases of nearly 30K were observed when the generated power of the column exceeded 25W/m. The impact of the heating due to viscous friction on the repeatability of peak capacity, elution time, and peak area ratio to an internal standard for a gradient UHPLC-MS/MS method to analyze neurotransmitters was found to be limited. This result indicates that high speed UHPLC-MS/MS gradient methods under conditions of high viscous friction may be possible without the negative effects typically observed with isocratic separations under similar conditions.

  11. Chiral liquid chromatography-mass spectrometry (LC-MS/MS) method development for the detection of salbutamol in urine samples.

    PubMed

    Chan, Sue Hay; Lee, Warren; Asmawi, Mohd Zaini; Tan, Soo Choon

    2016-07-01

    A sequential solid-phase extraction (SPE) method was developed and validated using liquid chromatography-electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS) for the detection and quantification of salbutamol enantiomers in porcine urine. Porcine urine samples were hydrolysed with β-glucuronidase/arylsulfatase from Helix pomatia and then subjected to a double solid-phase extraction (SPE) first using the Abs-Elut Nexus SPE and then followed by the Bond Elut Phenylboronic Acid (PBA) SPE. The salbutamol enantiomers were separated using the Astec CHIROBIOTIC™ T HPLC column (3.0mm×100mm; 5μm) maintained at 15°C with a 15min isocratic run at a flow rate of 0.4mL/min. The mobile phase constituted of 5mM ammonium formate in methanol. Salbutamol and salbutamol-tert-butyl-d9 (internal standard, IS) was monitored and quantified with the multiple reaction monitoring (MRM) mode. The method showed good linearity for the range of 0.1-10ng/mL with limit of quantification at 0.3ng/mL. Analysis of the QC samples showed intra- and inter-assay precisions to be less than 5.04%, and recovery ranging from 83.82 to 102.33%.

  12. Simultaneous Determination of Tramadol and Its Metabolite in Human Urine by the Gas Chromatography-Mass Spectrometry Method.

    PubMed

    Yilmaz, Bilal; Erdem, Ali Fuat

    2015-08-01

    A sensitive and efficient method was developed for determination of tramadol and its metabolite (O-desmethyltramadol) in human urine by gas chromatography-mass spectrometry. Tramadol, O-desmethyltramadol and medazepam (internal standard) were extracted from human urine with a mixture of ethylacetate and diethylether mixture (1 : 1, v/v) at basic pH with liquid-liquid extraction. The calibration curves were linear (r = 0.99) over tramadol and O-desmethyltramadol concentrations ranging from 10 to 200 ng/mL and 7.5 to 300 ng/mL, respectively. The method had an accuracy of >95% and intra- and interday precision (relative standard deviation %) of ≤4.93 and ≤4.62% for tramadol and O-desmethyltramadol, respectively. The extraction recoveries were found to be 94.1 ± 2.91 and 96.3 ± 3.46% for tramadol and O-desmethyltramadol, respectively. The limit of quantification using 0.5 mL human urine was 10 ng/mL for tramadol and 7.5 ng/mL for O-desmethyltramadol. After oral administration of 100 mg of tramadol hydrochloride to a patient, the urinary excretion was monitored during 24 h. About 15% of the dose was excreted as unchanged tramadol.

  13. Schinus terebinthifolius countercurrent chromatography (Part II): Intra-apparatus scale-up and inter-apparatus method transfer.

    PubMed

    Costa, Fernanda das Neves; Vieira, Mariana Neves; Garrard, Ian; Hewitson, Peter; Jerz, Gerold; Leitão, Gilda Guimarães; Ignatova, Svetlana

    2016-09-30

    Countercurrent chromatography (CCC) is being widely used across the world for purification of various materials, especially in natural product research. The predictability of CCC scale-up has been successfully demonstrated using specially designed instruments of the same manufacturer. The reality is that the most of CCC users do not have access to such instruments and do not have enough experience to transfer methods from one CCC column to another. This unique study of three international teams is based on innovative approach to simplify the scale-up between different CCC machines using fractionation of Schinus terebinthifolius berries dichloromethane extract as a case study. The optimized separation methodology, recently developed by the authors (Part I), was repeatedly performed on CCC columns of different design available at most research laboratories across the world. Hexane - ethyl acetate - methanol - water (6:1:6:1, v/v/v/v) was used as solvent system with masticadienonic and 3β-masticadienolic acids as target compounds to monitor stationary phase retention and calculate peak resolution. It has been demonstrated that volumetric, linear and length scale-up transfer factors based on column characteristics can be directly applied to different i.d., volume and length columns independently on instrument make in an intra-apparatus scale-up and inter-apparatus method transfer.

  14. Development of supercritical fluid extraction and supercritical fluid chromatography purification methods using rapid solubility screening with multiple solubility chambers.

    PubMed

    Gahm, Kyung H; Huang, Ke; Barnhart, Wesley W; Goetzinger, Wolfgang

    2011-01-01

    Rapid solubility screening in diverse supercritical fluids (SCFs) was carried out via multiple solubility chambers with a trapping device and online ultraviolet (UV) detection. With this device, it was possible to rapidly study the solubility variations of multiple components in a mixture. Results from solubility studies have been used to develop efficient supercritical fluid extraction (SFE) and supercritical fluid chromatography (SFC) methods. After the investigation of solubilities of theophylline and caffeine in several neat organic solvents and SCFs, advantages of SFE over conventional organic solvent extraction were demonstrated with a model mixture of theophylline and caffeine. The highest solubility ratio of 1:40 (theophylline:caffeine) was observed in the SCF with 20% acetonitrile (MeCN), where a ratio of 1:11 was the highest in the neat organic solvents. A model mixture of theophylline:caffeine (85:15 w/w, caffeine as an impurity) was successfully purified by SFE by leveraging the highest solubility difference. The SCF with 20% MeCN selectively removed caffeine and left theophylline largely intact. Rapid SCF solubility screening was applied to development of SFE and SFC methods in a drug discovery environment. Two successful applications were demonstrated with proprietary Amgen compounds to either remove an achiral impurity before chiral purification or enhance chiral chromatographic throughput.

  15. Improved highly sensitive method for determination of nicotine and cotinine in human plasma by high-performance liquid chromatography.

    PubMed

    Nakajima, M; Yamamoto, T; Kuroiwa, Y; Yokoi, T

    2000-05-26

    A highly sensitive and reliable method for the determination of nicotine and its metabolite cotinine in human plasma by high-performance liquid chromatography was developed. Nicotine and cotinine were extracted from alkalinized plasma with dichloromethane and the volatility of nicotine was prevented by the addition of conc. HCl to the organic solvent during evaporation. The sensitivity of quantification at 260 nm absorption was improved by using a noise-base clean Uni-3 to 0.2 ng/ml nicotine and 1.0 ng/ml cotinine. The method was validated over linear ranges of 0.2-25.0 ng/ml for nicotine and 1.0-80.0 ng/ml for cotinine. The intra-day precision and accuracy were < or = 15.9% relative standard variation (RSD) and 89.9-103.5% for nicotine and < or = 8.0% RSD and 98.7-103.0% for cotinine. The inter-day precision and accuracy were < or = 17.0% RSD and 94.2-100.9% for nicotine and < or = 8.2% RSD and 98.0-105.1% for cotinine.

  16. Gas chromatography-mass spectrometric method-based urine metabolomic profile of rats with pelvic inflammatory disease

    PubMed Central

    ZOU, WEI; WEN, XIAOKE; SHENG, XIAOQI; ZHENG, YI; XIAO, ZUOQI; LUO, JIEYING; CHEN, SHUQIONG; WANG, YICHAO; CHENG, ZENENG; XIANG, DAXIONG; NIE, YICHU

    2016-01-01

    Pelvic inflammatory disease (PID) can lead to a poor outcome of severe sequelae, and the current methods of clinical diagnosis are not satisfactory. Metabolomics is an effective method for the identification of disease-related metabolite biomarkers to facilitate disease diagnosis. However, to the best of our knowledge, no PID-associated metabolomic study has yet been carried out. The metabolomic changes of rats with PID were investigated in the present study. A PID model was constructed by the multi-pathogenic infection of the upper genital tract in rats. Infiltration of inflammatory cells and elevated expression levels of the cytokines interleukin (IL)-1β and IL-6 in the uterus and fallopian tubes validated the disease model. Gas chromatography-mass spectrometry coupled with derivatization was used to determine the urine metabolomic profile. Principal component analysis and partial least squares-discriminant analysis of the data sets showed a clear separation of metabolic profiles between rats with PID and control rats. Eighteen differentiating metabolites were found, including four amino acids, three fatty acids, nine organic acids, and two sugars, which indicated alterations in sugar metabolism, the citric acid cycle, amino acid metabolism and fatty acid metabolism. These metabolites could be potential biomarkers of PID, and this research may offer a new approach to evaluate the effect of anti-PID drugs in pre-clinical or clinical trials. PMID:27168785

  17. New High-performance Liquid Chromatography-DAD Method for Analytical Determination of Arbutin and Hydroquinone in Rat Plasma

    PubMed Central

    Gallo, F. R.; Pagliuca, G.; Multari, G.; Panzini, G.; D’amore, E.; Altieri, I.

    2015-01-01

    Natural substances present in herbal preparations should be carefully used because they can give toxic or therapeutic effects despite of their amount or the way of administration. The safety of products of vegetable origin must be assessed before commercialisation by monitoring the active ingredients and their metabolites. This study was therefore designed to identify and quantify arbutin and its metabolite hydroquinone, naturally present in Arctostaphylos uva-ursi (L.) Spreng plant in rat plasma, after an acute and subacute administration of aqueous arbutin solution in Wistar rats. For this purpose a reversed-phase high-performance liquid chromatography coupled with photodiode array detection was developed to assess the pharmacokinetic of arbutin and hydroquinone in plasma of female rats treated with aqueous arbutin solutions. The detection (arbutin: 0.0617 µg/ml and hydroquinone 0.0120 µg/ml) and quantification (arbutin: 0.2060 µg/ml and hydroquinone: 0.0400 µg/ml) limits were determined. At the arbutin concentration level of 10.7 µg/ml repeatability was 13.33% and its recovery 93.4±6.93%, while at the hydroquinone concentration level of 10.6 µg/ml repeatability was 11.66% and its recovery 92.9±7.75%. Furthermore the method was fully validated and the obtained data indicate that the new method provides good performances. PMID:26798166

  18. Optimized ultra performance liquid chromatography tandem high resolution mass spectrometry method for the quantification of paraquat in plasma and urine.

    PubMed

    Lu, Haihua; Yu, Jing; Wu, Linlin; Xing, Jingjing; Wang, Jun; Huang, Peipei; Zhang, Jinsong; Xiao, Hang; Gao, Rong

    2016-08-01

    A simple, sensitive and specific ultra performance liquid chromatography coupled to electrospray tandem high resolution mass spectrometry (UPLC-ESI-HRMS/MS) method has been developed and validated for quantification of paraquat in plasma and urine. The sample preparation was carried out by one-step protein precipitation with acetonitrile. The paraquat was separated with a HILIC column in 10min. Detection was performed using Q Exactive Orbitrap mass spectrometer by Targeted-MS/MS scan mode. Methodological parameters, such as ammonium formate concentration, formic acid concentration, spray voltage, capillary temperature, heater temperature and normalized collision energy were optimized to achieve the highest sensitivity. The calibration curve was linear over the concentration range of LOQ-1000ng/mL. LOD was 0.1 and 0.3ng/mL, LOQ was 0.3 and 0.8ng/mL for urine and plasma, respectively. The intra- and inter-day precisions were <7.97% and 4.78% for plasma and urine. The accuracies were within the range 93.51-100.90%. The plasma and urine matrices had negligible relative matrix effect in this study. This method was successfully applied to determine paraquat concentration in plasma samples with hemoperfusion from 5 suspected paraquat poisoning patients.

  19. Development of a comprehensive analytical method for phosphate metabolites in plants by ion chromatography coupled with tandem mass spectrometry.

    PubMed

    Sekiguchi, Yoko; Mitsuhashi, Naoto; Kokaji, Tetsuo; Miyakoda, Hidekazu; Mimura, Tetsuro

    2005-08-26

    This paper describes the development of a practical method for the analysis of phosphorus compounds with a focus on sugar phosphates from the model higher plant Arabidopsis thaliana by ion chromatography coupled to electrospray ionization tandem mass spectrometry (IC-ESI-MS-MS). After the analytical separation, the potassium hydroxide eluent was converted to water with an anion suppressor allowing the effluent from the IC to be connected to the mass spectrometer directly. In the optimized method, 17 phosphorous compounds (adenosine diphosphate (ADP), fructose 1,6-bisphosphate, fructose 2,6-bisphosphate, fructose 6-phosphate, galactose 1-phosphate, glucose 1-phosphate, glucose 1,6-bisphosphate, glucose 6-phosphate, mannose 6-phosphate, phosphoenol pyrvate, 3-phosphoglyceric acid, ribulose 1,5-bisphosphate, ribulose 5-phosphate, ribose 5-phosphate, sucrose 6-phosophate and uridine 5'-diphosphate-glucose (UDPG)) were determined. The linearity of response for these phosphorous compounds over the concentration range of 0 and 10 microM was better than 0.9993 in all cases. The minimum detection limit was between 0.01 and 2.50 microM for a 25 microL injection, and recovery rates for standard addition to the sample were within the range from 93% to 110%.

  20. Rapid and sensitive method for the analysis of carbon monoxide in blood using gas chromatography with flame ionisation detection.

    PubMed

    Sundin, Anna-Maja; Larsson, Jan Erik

    2002-01-05

    In order to measure changes in physiological CO concentrations in blood with good accuracy, a method was developed using gas chromatography with flame ionisation detection (250 degrees C). A nickel catalyst system was fitted to convert CO to methane at 375 degrees C after separation with a molecular sieve column at 35 degrees C. Helium was used as carrier at 30 ml/min. Porcine or human blood (400 microl) was sampled in gastight tubes and treated with sulfuric acid and saponin (800 microl). Accuracy was 1.4% and 1.5% (RSD), respectively. Precision was 2.8% (porcine blood). Limit of detection was 0.01 nmol/ml gas and limit of quantification 12 nmol/ml blood. Calibration was made in the interval 12-514 nmol/ml blood (corresponding to 0.1-6% COHb). Samples were stable for at least a month at +4 degrees C. This paper describes a method with high sensitivity and good accuracy, suitable for analysis of low CO concentrations.

  1. A novel method for analysing key corticosteroids in polar bear (Ursus maritimus) hair using liquid chromatography tandem mass spectrometry.

    PubMed

    Weisser, Johan J; Hansen, Martin; Björklund, Erland; Sonne, Christian; Dietz, Rune; Styrishave, Bjarne

    2016-04-01

    This paper presents the development and evaluation of a methodology for extraction, clean-up and analysis of three key corticosteroids (aldosterone, cortisol and corticosterone) in polar bear hair. Such a methodology can be used to monitor stress biomarkers in polar bears and may provide as a useful tool for long-term and retrospective information. We developed a combined pressurized liquid extraction (PLE)-solid phase extraction (SPE) procedure for corticosteroid extraction and clean-up followed by high pressure liquid chromatography tandem mass spectrometry (HPLC-MS/MS) analysis. This procedure allows for the simultaneous determination of multiple steroids, which is in contrast to previous polar bear studies based on ELISA techniques. Absolute method recoveries were 81%, 75% and 60% for cortisol, corticosterone and aldosterone, respectively. We applied the developed method on a hair sample pooled from four East Greenland polar bears. Herein cortisol and corticosterone were successfully determined in levels of 0.32±0.02ng/g hair and 0.13±0.02ng/g hair, respectively. Aldosterone was below limit of detection (LOD<0.17ng/g). The cortisol hair concentration found in these East Greenland polar bears was consistent with cortisol levels previously determined in the Southern Hudson Bay and James Bay in Canada using ELISA kits.

  2. Development of a thermal desorption-gas chromatography-mass spectrometry method for determining personal care products in air.

    PubMed

    Ramírez, Noelia; Marcé, Rosa Maria; Borrull, Francesc

    2010-06-25

    This study describes the development of a new analytical method for determining 14 personal care products (PCPs) - nine synthetic musks, four parabens and one insect repellent - in air samples. The method is based on active sampling on sorbent tubes and thermal desorption-gas chromatography-mass spectrometry analysis, and is rapid, sensitive and drastically reduces the risk of sample contamination. Three kinds of tubes and traps were tested, those filled with Tenax TA being the most suitable for this study. Method validation showed good repeatability and reproducibility, low detection limits (between 0.03 ng m(-3) for DPMI and 12.5 ng m(-3) for propyl paraben) and good linearity for all compounds. Stability during storage indicated that samples must be kept refrigerated at 4 degrees C and analysed within 1 week of collection. The applicability of the technique to real samples was tested in different indoor and outdoor atmospheres. The total PCP values for indoor air ranged from 135 ng m(-3) in a pharmacy to 2838 ng m(-3) in a hairdresser's, whereas the values for outdoor air ranged from 14 ng m(-3) for a suburban environment to 26 ng m(-3) for an urban environment. In general, the most abundant synthetic musks were galaxolide (5.9-1256 ng m(-3)), musk xylene (1.6-766 ng m(-3)) and tonalide (1.1-138 ng m(-3)). Methyl and ethyl paraben (2.4-313 ng m(-3) and 1.8-117 ng m(-3), respectively) were the most abundant parabens. Although thermal desorption methods have been widely used for determining volatile organic compounds, they are rarely used with semi-volatile compounds. This study thus demonstrates that the thermal desorption method performs well with semi-volatile compounds and, for the first time, that it can be used for determining PCPs.

  3. Method development and validation for the analysis of didanosine using micellar electrokinetic capillary chromatography.

    PubMed

    Mallampati, Swapna; Leonard, Stefanie; De Vulder, Sabine; Hoogmartens, Jos; Van Schepdael, Ann

    2005-11-01

    A selective MEKC method was developed for the analysis of didanosine in bulk samples. Successful separation of didanosine from 13 of its potential impurities, derived from the various synthetic preparation procedures, was achieved. As CZE gave poor separation selectivity, MEKC was preferable. The use of EKC allowed achievement of the separation in a significantly shorter time than conventional HPLC. An anionic long-chain surfactant, lithium dodecyl sulfate (LiDS), was used as the pseudostationary phase and sodium tetraborate buffer as the aqueous phase. In order to obtain the optimal conditions and to test the method robustness, a central composite response surface modeling experiment was performed. The optimized electrophoretic conditions include the use of an uncoated fused-silica capillary with a total length of 40 cm and an ID of 50 microm, a BGE containing 40 mM sodium tetraborate and 110 mM LiDS at pH 8.0, an applied voltage of 18.0 kV, and the capillary temperature maintained at 15 degrees C. The method was found to be robust. The parameters for validation such as linearity, precision, and sensitivity are also reported. Three commercial bulk samples were analyzed with this system.

  4. Method for the Determination of Ammonia in Mainstream Cigarette Smoke Using Ion Chromatography

    PubMed Central

    Watson, Christina Vaughan; Feng, June; Valentin-Blasini, Liza; Stanelle, Rayman; Watson, Clifford H.

    2016-01-01

    Ammonia in mainstream smoke is present in both the particulate and vapor phases. The presence of ammonia in the cigarette filler material and smoke is of significance because of the potential role ammonia could have in raising the “smoke pH.” An increased smoke pH could shift a fraction of total nicotine to free-base nicotine, which is reportedly more rapidly absorbed by the smoker. Methods measuring ammonia in smoke typically employ acid filled impingers to trap the smoke. We developed a fast, reliable method to measure ammonia in mainstream smoke without the use of costly and time consuming impingers to examine differences in ammonia delivery. The method uses both a Cambridge filter pad and a Tedlar bag to capture particulate and vapor phases of the smoke. We quantified ammonia levels in the mainstream smoke of 50 cigarette brands from 5 manufacturers. Ammonia levels ranged from approximately 1μg to 23μg per cigarette for ISO smoking conditions and 38μg to 67μg per cigarette for Canadian intense smoking conditions and statistically significance differences were observed between brands and manufacturers. Our findings suggest that ammonia levels vary by brand and are higher under Canadian intense smoking conditions. PMID:27415766

  5. A model free method for estimation of complicated adsorption isotherms in liquid chromatography.

    PubMed

    Forssén, Patrik; Fornstedt, Torgny

    2015-08-28

    Here we show that even extremely small variations in the adsorption isotherm can have a tremendous effect on the shape of the overloaded elution profiles and that the earlier in the adsorption isotherms the variation take place, the larger its impact on the shape of the elution profile. These variations are so small that they can be "hidden" by the discretization and in the general experimental noise when using traditional experimental methods, such as frontal analysis, to measure adsorption isotherms. But as the effects of these variations are more clearly visible in the elution profiles, the Inverse Method (IM) of adsorption isotherm estimation is an option. However, IM usually requires that one selects an adsorption isotherm model prior to the estimation process. Here we show that even complicated models might not be able to estimate the adsorption isotherms with multiple inflection points that small variations might give rise to. We therefore developed a modified IM that, instead of fixed adsorption isotherm models, uses monotone piecewise interpolation. We first validated the method with synthetic data and showed that it can be used to estimate an adsorption isotherm, which accurately predicts an extremely "strange" elution profile. For this case it was impossible to estimate the adsorption isotherm using IM with a fixed adsorption model. Finally, we will give an example of a real chromatographic system where adsorption isotherm with inflection points is estimated by the modified IM.

  6. The novel assay method for nicotine metabolism to cotinine using high performance liquid chromatography.

    PubMed

    Miyazawa, Mitsuo; Kawauchi, Yumi; Okuno, Yoshiharu; Oda, Yoshimitsu

    2011-01-01

    Nicotine is the primary psychoactive component in tobacco. It is taken into the body by tobacco smoking, and mainly metabolized to cotinine in the hepatic cytochrme P450 (CYP) 2A6. The objective of this study was to develop a sensitive method for the determination of nicotine metabolism to cotinine using HPLC. The internal standard, trans-4'-carboxycotinine methyl ester was synthesized with a simple method. The nicotine and cotinine were separated completely and detected by C(18) 5-µm analytical column (L-column Octa decyl silyl (ODS), 150 mm × 4.6 mm i.d.) equipped with a C(18) 5-µm guard column (L-column ODS, 10 mm × 4.6 mm i.d.) and ultraviolet detection at 260 nm. The detection limit of the assay was 0.05 µM for cotinine (n=5, R.S.D) and 0.1 µM for nicotine. Thus the present results provided a sensitive and useful method for the determination of nicotine metabolism catalyzed by CYP2A6.

  7. A hydrophilic interaction chromatography-tandem mass spectrometry method for amino acid profiling in mussels.

    PubMed

    Tsochatzis, Emmanouil D; Begou, Olga; Gika, Helen G; Karayannakidis, Panayotis D; Kalogiannis, Stavros

    2017-03-15

    A UHPLC-HILIC-tandem MS method has been developed and validated for the quantification of 21 amino acids (20 protein amino acids and cystine) in their free form (FAA) and as protein constituents (total amino acids, TAA) in a rich protein food matrix such as lyophilized mussels (Mytilus galloprovincialis) samples. FAA were analyzed after suspending the samples in the presence of trichloroacetic acid in order to prevent dissolving the proteins, while TAA were determined after acid hydrolysis with 6M HCl in the presence of 4% v/v thioglycolic acid as a reducing agent. In hydrolysed samples 17 amino acids could be determined since tryptophan, cysteine, cystine and asparagine were degraded during acid hydrolysis. Linear regression coefficients (R(2)) were above 0.99 for all amino acids. Accuracy and precision, expressed as recovery (%) and relative standard deviation (RSD, %) were in acceptable levels, ranging from 78.2 to 123.3% and below 15%, respectively for both FAA and TAA. Uncertainty was also below 12% for FAA and below 22% for TAA. Sensitivity of the method was high with LOD values ranging from 0.003 to 0.034g/100g for FAA and 0.001 to 0.004g/100g for TAA, while LOQ ranged from 0.009 to 0.104g/100g for FAA and 0.002 to 0.011g/100g for TAA. The method proved to be a fast and reliable tool for acquiring information on free and total amino acids profile in high protein content foodstuffs such as mussels.

  8. Method development and validation for isoflavones in soy germ pharmaceutical capsules using micellar electrokinetic chromatography.

    PubMed

    Micke, Gustavo Amadeu; Fujiya, Neide Mitsue; Tonin, Fernando Gustavo; de Oliveira Costa, Ana Carolina; Tavares, Marina Franco Maggi

    2006-08-28

    The separation of six soy isoflavones (Glycitein, Daidzein, Genistein, Daidzin, Glycitin and Genistin) was approached by a 3(2) factorial design studying MEKC electrolyte components at the following levels: methanol (MeOH; 0-10%) and sodium dodecylsulfate (SDS; 20-70 mmol L(-1)); sodium tetraborate buffer (STB) concentration was kept constant at 10 mmol L(-1). Nine experiments were performed and the apparent mobility of each isoflavone was computed as a function of the electrolyte composition. A novel response function (RF) was formulated based on the production of the mobility differences, mobility of the first and last eluting peaks and the electrolyte conductance. The inspection of the response surface indicated an optimum electrolyte composition as 10 mmol L(-1) STB (pH 9.3) containing 40 mmol L(-1) SDS and 1% MeOH promoting baseline separation of all isoflavones in less than 7.5 min. The proposed method was applied to the determination of total isoflavones in soy germ capsules from four different pharmaceutical laboratories. A 2h extraction procedure with 80% (v/v) MeOH under vortexing at room temperature was employed. Peak assignment of unknown isoflavones in certain samples was assisted by hydrolysis procedures, migration behavior and UV spectra comparison. Three malonyl isoflavone derivatives were tentatively assigned. A few figures of merit for the proposed method include: repeatability (n=6) better than 0.30% CV (migration time) and 1.7% CV (peak area); intermediate precision (n=18) better than 6.2% CV (concentration); recoveries at two concentration levels, 20 and 50 microg mL(-1), varied from 99.1 to 103.6%. Furthermore, the proposed method exhibited linearity in the concentration range of 1.6-50 microg mL(-1) (r(2)>0.9999) with LOQ varying from 0.67 to 1.2 microg mL(-1). The capsules purity varied from 93.3 to 97.6%.

  9. Development of a Method for Crustacean Allergens Using Liquid Chromatography/Tandem Mass Spectrometry.

    PubMed

    Nagai, Hiroyuki; Minatani, Tomiaki; Goto, Kotaro

    2015-01-01

    An LC/MS/MS analysis method was developed for crustacean allergens, tropomyosin, and arginine kinase. A protein extract from shrimp was reduced, alkylated, and digested by trypsin. Peptide spectra were obtained using full scan analysis by LC/MS/MS, and we determined a sequence through a protein search. 22ADTLEQQNK30, 92IQLLEEDLER101, 113LAEASQAADESER125, 134SLSDEER140, 153FLAEEADR160, and 190IVELEEELR198 of tropomyosin and 152VSSTLSSLEGELK164 and 217TFLVWVNEEDHLR229 of arginine kinase were selected as the specific peptides, and optimal multiple-reaction monitoring conditions were used. The results obtained through the LC/MS/MS analysis correlated well with those using the ELISA method for various crustacean samples (r2>0.9). Moreover, unregulated species, such as krill or insects, which produce positive results in some crustacean ELISA assays, can be differentiated by LC/MS/MS. These findings suggest that LC/MS/MS analysis may be effective for crustacean food allergen analysis.

  10. Validated high-performance thin-layer chromatography method for steviol glycosides in Stevia rebaudiana.

    PubMed

    Jaitak, Vikas; Gupta, A P; Kaul, V K; Ahuja, P S

    2008-08-05

    A high-performance thin-layer chromatographic (HPTLC) method was developed and validated as per ICH (International Conferences on Harmonization) guidelines for simultaneous quantification of three steviol glycosides, i.e. steviolbioside, stevioside and rebaudioside-A in Stevia rebaudiana leaves. For achieving good separation, mobile phase of ethyl acetate-ethanol-water (80:20:12, v/v/v) on pre-coated silica gel 60 F254 HPTLC plates were used. The densitometric quantification of steviol glycosides was carried out at lambda=510 nm in reflection-absorption mode after spraying with acetic anhydride:sulphuric acid:ethanol reagent. The calibration curves were linear in the range of 160-960 ng/spot for steviolbioside, 1-6 microg/spot for stevioside and 0.5-3 microg/spot for rebaudioside-A with good correlation coefficients (0.998-0.999). The method was found to be reproducible for quantitative analysis of steviol glycosides in S. rebaudiana leaves collected from ten different locations and will serve as a quality control indicator to monitor the commercial production of stevioside and its allied molecules during different stages of its processing.

  11. Validation of a high performance liquid chromatography method for the stabilization of epigallocatechin gallate.

    PubMed

    Fangueiro, Joana F; Parra, Alexander; Silva, Amélia M; Egea, Maria A; Souto, Eliana B; Garcia, Maria L; Calpena, Ana C

    2014-11-20

    Epigallocatechin gallate (EGCG) is a green tea catechin with potential health benefits, such as anti-oxidant, anti-carcinogenic and anti-inflammatory effects. In general, EGCG is highly susceptible to degradation, therefore presenting stability problems. The present paper was focused on the study of EGCG stability in HEPES (N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid) medium regarding the pH dependency, storage temperature and in the presence of ascorbic acid a reducing agent. The evaluation of EGCG in HEPES buffer has demonstrated that this molecule is not able of maintaining its physicochemical properties and potential beneficial effects, since it is partially or completely degraded, depending on the EGCG concentration. The storage temperature of EGCG most suitable to maintain its structure was shown to be the lower values (4 or -20 °C). The pH 3.5 was able to provide greater stability than pH 7.4. However, the presence of a reducing agent (i.e., ascorbic acid) was shown to provide greater protection against degradation of EGCG. A validation method based on RP-HPLC with UV-vis detection was carried out for two media: water and a biocompatible physiological medium composed of Transcutol®P, ethanol and ascorbic acid. The quantification of EGCG for purposes, using pure EGCG, requires a validated HPLC method which could be possible to apply in pharmacokinetic and pharmacodynamics studies.

  12. Improved gas chromatography-flame ionization detector analytical method for the analysis of epoxy fatty acids.

    PubMed

    Mubiru, Edward; Shrestha, Kshitij; Papastergiadis, Antonios; De Meulenaer, Bruno

    2013-11-29

    In this study an improved method for analysis of epoxy fatty acids is reported. Data obtained from analysis of polar fatty acids has previously been presented, but due to the high number of compounds that co-elute in the polar fraction, the resultant chromatograms are complex which may lead to compromising the accuracy of the data. A three steps separation of fatty acid methyl esters (FAMEs) by solid-phase extraction (SPE) on a silica gel column to remove hydroxy fatty acid interferences was proposed. This approach is opposed to a two step separation procedure that has been often used to prevent analytical interferences caused by non-altered fatty acids. A gas chromatograph with a flame ionization detector (GC-FID) equipped with a polar CP-Sil 88™ column was used. Quantification was based on the use of methyl nonadecanoate (C19:0), as an internal standard. Individual mono epoxy fatty acids were well separated without co-eluting compounds. The optimized method was finally applied to screen epoxy fatty acids in 37 fresh oil samples. Results obtained for the total epoxy fatty acids were in the range 0.03-2mgg(-1) of oil with repeatability coefficient of variation (CV) ranging from 2.8 to 9.9% for duplicate analysis showing that the results obtained are repeatable.

  13. Comparative Analysis of Volatile Defensive Secretions of Three Species of Pyrrhocoridae (Insecta: Heteroptera) by Gas Chromatography-Mass Spectrometric Method

    PubMed Central

    Krajicek, Jan; Havlikova, Martina; Bursova, Miroslava; Ston, Martin; Cabala, Radomir; Exnerova, Alice; Stys, Pavel; Bosakova, Zuzana

    2016-01-01

    The true bugs (Hemiptera: Heteroptera) have evolved a system of well-developed scent glands that produce diverse and frequently strongly odorous compounds that act mainly as chemical protection against predators. A new method of non-lethal sampling with subsequent separation using gas chromatography with mass spectrometric detection was proposed for analysis of these volatile defensive secretions. Separation was performed on Rtx-200 column containing fluorinated polysiloxane stationary phase. Various mechanical irritation methods (ultrasonics, shaking, pressing bugs with plunger of syringe) were tested for secretion sampling with a special focus on non-lethal irritation. The preconcentration step was performed by sorption on solid phase microextraction (SPME) fibers with different polarity. For optimization of sampling procedure, Pyrrhocoris apterus was selected. The entire multi-parameter optimization procedure of secretion sampling was performed using response surface methodology. The irritation of bugs by pressing them with a plunger of syringe was shown to be the most suitable. The developed method was applied to analysis of secretions produced by adult males and females of Pyrrhocoris apterus, Pyrrhocoris tibialis and Scantius aegyptius (all Heteroptera: Pyrrhocoridae). The chemical composition of secretion, particularly that of alcohols, aldehydes and esters, is species-specific in all three pyrrhocorid species studied. The sexual dimorphism in occurrence of particular compounds is largely limited to alcohols and suggests their epigamic intraspecific function. The phenetic overall similarities in composition of secretion do not reflect either relationship of species or similarities in antipredatory color pattern. The similarities of secretions may be linked with antipredatory strategies. The proposed method requires only a few individuals which remain alive after the procedure. Thus secretions of a number of species including even the rare ones can be

  14. A high performance liquid chromatography with ultraviolet method for Eschweilera nana leaves and their anti-inflammatory and antioxidant activities

    PubMed Central

    Outuki, Priscila M.; Lazzeri, Nides S.; de Francisco, Lizziane M. B.; Bersani-Amado, Ciomar A.; Ferreira, Izabel C. P.; Cardoso, Mara Lane C.

    2015-01-01

    Background: Eschweilera nana Miers is a tree widely distributed in Cerrado, Brazil. Objective: In this study, we aimed to describe its phytochemical properties and antioxidant and topical anti-inflammatory effects for the first time, as well validate an high performance liquid chromatography with ultraviolet/visible (HPLC-UV-Vis) method for the separation and quantification of the main components (hyperoside and rutin) in the hydroalcoholic extract of E. nana leaves. Materials and Methods: Structural identification of compounds in E. nana extract was performed by analysis of spectral data by 1H nuclear magnetic resonance, 13C nuclear magnetic resonance and/or ESI/EM. The HPLC-UV-Vis method was validated according International Conference on Harmonization (ICH) parameters. The 2,2’-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) and 1,1-diphenyl-2-picrylhydrazyl (DPPH) method were used for determination of in vitro antioxidant activities and the croton oil-induced inflammation for evaluation of in vivo anti-inflammatory effects. Results: Hyperoside, rutin, α-amirin, β-amirin, β-sitosterol, and stigmasterol were identified in the hydroalcoholic extract of E. nana leaves. HPLC-UV-Vis was validated according to ICH parameters. Furthermore, in vitro and in vivo assays demonstrated that the hydroalcoholic extract and methanol fraction showed significant antioxidant and topical anti-inflammatory effects, as they were able to reduce ear edema induced by croton-oil application. Conclusions: This research showed the first phytochemical study of E. nana extract and their biological activities may be associated with the presence of flavonoids in the extracts. PMID:26246741

  15. Determination of total and unbound concentrations of lopinavir in plasma using liquid chromatography-tandem mass spectrometry and ultrafiltration methods.

    PubMed

    Illamola, S M; Labat, L; Benaboud, S; Tubiana, R; Warszawski, J; Tréluyer, J M; Hirt, D

    2014-08-15

    Lopinavir is an HIV protease inhibitor with high protein binding (98-99%) in human plasma. This study was designed to develop an ultrafiltration method to measure the unbound concentrations of lopinavir overcoming the non-specific binding issue. A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the determination of total concentrations of lopinavir in plasma was developed and validated, and an adaptation was also optimized and validated for the determination of unbound concentrations. The chromatographic separation was performed with a C18 column (100 mm × 2.1mm i.d., 5 μm particle size) using a mobile phase containing deionized water with formic acid, and acetonitrile, with gradient elution at a flow-rate of 350 μL min(-1). Identification of the compounds was performed by multiple reaction monitoring, using electrospray ionization in positive ion mode. The method was validated over a clinical range of 0.01-1 μg/mL for human plasma ultrafiltrate and 0.1-15 μg/mL in human plasma. The inter and intra-assay accuracies and precisions were between 0.23% and 11.37% for total lopinavir concentrations, and between 3.50% and 13.30% for plasma ultrafiltrate (unbound concentration). The ultrafiltration method described allows an accurate separation of the unbound fraction of lopinavir, circumscribing the loss of drug by nonspecific binding (NSB), and the validated LC-MS/MS methodology proposed is suitable for the determination of total and unbound concentrations of lopinavir in clinical practice.

  16. Method for enhanced accuracy in predicting peptides using liquid separations or chromatography

    DOEpatents

    Kangas, Lars J.; Auberry, Kenneth J.; Anderson, Gordon A.; Smith, Richard D.

    2006-11-14

    A method for predicting the elution time of a peptide in chromatographic and electrophoretic separations by first providing a data set of known elution times of known peptides, then creating a plurality of vectors, each vector having a plurality of dimensions, and each dimension representing the elution time of amino acids present in each of these known peptides from the data set. The elution time of any protein is then be predicted by first creating a vector by assigning dimensional values for the elution time of amino acids of at least one hypothetical peptide and then calculating a predicted elution time for the vector by performing a multivariate regression of the dimensional values of the hypothetical peptide using the dimensional values of the known peptides. Preferably, the multivariate regression is accomplished by the use of an artificial neural network and the elution times are first normalized using a transfer function.

  17. Improved resins and novel materials and methods for solid phase extraction and high performance liquid chromatography

    SciTech Connect

    Freeze, Ronald

    1997-10-08

    Solid-phase extraction (SPE) has grown to be one of the most widely used methods for isolation and preconcentration of a vast range of compounds from aqueous solutions. By modifying polymeric SPE resins with chelating functional groups, the selective uptake of metals was accomplished. The resin, along with adsorbed metals, was vaporized in the ICP and detection of the metals was then possible using either mass or emission spectroscopy. Drug analyses in biological fluids have received heightened attention as drug testing is on the increase both in sports and in the work environment. By using a direct-injection technique, biological fluids can be injected directly into the liquid chromatographic system with no pretreatment. A new surfactant, a sulfonated form of Brij-30 (Brij-S) is shown to prevent the uptake of serum proteins on commercial HPLC columns by forming a thin coating on the silica C18 surface. Excellent separations of eight or more drugs with a wide range of retention times were obtained. The separations had sharper peaks and lower retention times than similar separations performed with the surfactant sodium dodecylsulfate (SDS). Quantitative recovery of a number of drugs with limits of detection near 1 ppm with a 5 μl injection volume were obtained. Finally, a method for solid-phase extraction in a syringe is introduced. The system greatly reduced the volume of solvent required to elute adsorbed analytes from the SPE bed while providing a semi-automated setup. SPE in a syringe consists of a very small bed of resin-loaded membrane packed into a GC or HPLC syringe. After extraction, elution was performed with just a few μl of solvent. This small elution volume allowed injection of the eluent directly from the syringe into the chromatographic system, eliminating the handling problems associated with such small volumes.

  18. A simple, rapid and validated high-performance liquid chromatography method suitable for clinical measurements of human mercaptalbumin and non-mercaptalbumin.

    PubMed

    Yasukawa, Keiko; Shimosawa, Tatsuo; Okubo, Shigeo; Yatomi, Yutaka

    2017-01-01

    Background Human mercaptalbumin and human non-mercaptalbumin have been reported as markers for various pathological conditions, such as kidney and liver diseases. These markers play important roles in redox regulations throughout the body. Despite the recognition of these markers in various pathophysiologic conditions, the measurements of human mercaptalbumin and non-mercaptalbumin have not been popular because of the technical complexity and long measurement time of conventional methods. Methods Based on previous reports, we explored the optimal analytical conditions for a high-performance liquid chromatography method using an anion-exchange column packed with a hydrophilic polyvinyl alcohol gel. The method was then validated using performance tests as well as measurements of various patients' serum samples. Results We successfully established a reliable high-performance liquid chromatography method with an analytical time of only 12 min per test. The repeatability (within-day variability) and reproducibility (day-to-day variability) were 0.30% and 0.27% (CV), respectively. A very good correlation was obtained with the results of the conventional method. Conclusions A practical method for the clinical measurement of human mercaptalbumin and non-mercaptalbumin was established. This high-performance liquid chromatography method is expected to be a powerful tool enabling the expansion of clinical usefulness and ensuring the elucidation of the roles of albumin in redox reactions throughout the human body.

  19. Methods of analysis by the U.S. Geological Survey Organic Geochemistry Research Group; determination of chloroacetanilide herbicide metabolites in water using high-performance liquid chromatography-diode array detection and high-performance liquid chromatography/mass spectrometry

    USGS Publications Warehouse

    Zimmerman, L.R.; Hostetler, K.A.; Thurman, E.M.

    2000-01-01

    Analytical methods using high-performance liquid chromatography-diode array detection (HPLC-DAD) and high-performance liquid chromatography/mass spectrometry (HPLC/MS) were developed for the analysis of the following chloroacetanilide herbicide metabolites in water: acetochlor ethanesulfonic acid (ESA), acetochlor oxanilic acid (OXA), alachlor ESA, alachlor OXA, metolachlor ESA, and metolachlor OXA. Good precision and accuracy were demonstrated for both the HPLC-DAD and HPLC/MS methods in reagent water, surface water, and ground water. The mean HPLC-DAD recoveries of the chloroacetanilide herbicide metabolites from water samples spiked at 0.25, 0.50, and 2.0 mg/L (micrograms per liter) ranged from 84 to 112 percent, with relative standard deviations of 18 percent or less. The mean HPLC/MS recoveries of the metabolites from water samples spiked at 0.05, 0.20, and 2.0 mg/L ranged from 81 to 125 percent, with relative standard deviations of 20 percent or less. The limit of quantitation (LOQ) for all metabolites using the HPLC-DAD method was 0.20 mg/L, whereas the LOQ using the HPLC/MS method was 0.05 mg/L. These metabolite-determination methods are valuable for acquiring information about water quality and the fate and transport of the parent chloroacetanilide herbicides in water.

  20. Methods of analysis by the U.S. Geological Survey National Water Quality Laboratory; determination of chlorinated pesticides in aquatic tissue by capillary-column gas chromatography with electron-capture detection

    USGS Publications Warehouse

    Leiker, Thomas J.; Madsen, J.E.; Deacon, J.R.; Foreman, W.T.

    1995-01-01

    A method for the determination of chlorinated organic compounds in aquatic tissue by dual capillary-column gas chromatography with electron-capture detection is described. Whole-body-fish or corbicula tissue is homogenized, Soxhlet extracted, lipid removed by gel permeation chromatography, and fractionated using alumina/silica adsorption chromatography. The extracts are analyzed by dissimilar capillary-column gas chromatography with electron-capture detection. The method reporting limits are 5 micrograms per kilogram (μg/kg) for chlorinated compounds, 50 μg/kg for polychlorinated biphenyls, and 200 μg/kg for toxaphene.

  1. Enantiomeric separation of the antiuremic drug colchicine by electrokinetic chromatography. Method development and quantitative analysis.

    PubMed

    Menéndez-López, Nuria; Valimaña-Traverso, Jesús; Castro-Puyana, María; Salgado, Antonio; García, María Ángeles; Marina, María Luisa

    2017-05-10

    Two analytical methodologies were developed by CE enabling the enantiomeric separation of colchicine, an antiuremic drug commercialized as pure enantiomer. Succinyl-γ-CD and Sulfated-γ-CD were selected as chiral selectors after a screening with different anionic CDs. Under the optimized conditions, chiral resolutions of 5.6 in 12min and 3.2 in 8min were obtained for colchicine with Succinyl-γ-CD and Sulfated-γ-CD, respectively. An opposite enantiomeric migration order was observed with these two CDs being S-colchicine the first-migrating enantiomer with Succinyl-γ-CD and the second-migrating enantiomer with Sulfated-γ-CD. H NMR experiments showed a 1:1 stoichiometry for the enantiomer-CD complexes in both cases. However, the apparent and averaged equilibrium constants for the enantiomer-CD complexes could be calculated only for Succinyl-γ-CD. The developed methods were applied to the analysis of pharmaceutical formulations but only the use of Succinyl-γ-CD enabled to detect a 0.1% of enantiomeric impurity in colchicine formulations.

  2. Evaluation of fast enantioselective multidimensional gas chromatography methods for monoterpenic compounds: Authenticity control of Australian tea tree oil.

    PubMed

    Wong, Yong Foo; West, Rachel N; Chin, Sung-Tong; Marriott, Philip J

    2015-08-07

    This work demonstrates the potential of fast multiple heart-cut enantioselective multidimensional gas chromatography (GC-eGC) and enantioselective comprehensive two-dimensional gas chromatography (eGC×GC), to perform the stereoisomeric analysis of three key chiral monoterpenes (limonene, terpinen-4-ol and α-terpineol) present in tea tree oil (TTO). In GC-eGC, separation was conducted using a combination of mid-polar first dimension ((1)D) column and a chiral second dimension ((2)D) column, providing interference-free enantioresolution of the individual antipodes of each optically active component. A combination of (1)D chiral column and (2)D polar columns (ionic liquid and wax phases) were tested for the eGC×GC study. Quantification was proposed based on summation of two major modulated peaks for each antipode, displaying comparable results with those derived from GC-eGC. Fast chiral separations were achieved within 25min for GC-eGC and<20min for eGC×GC, while ensuring adequate interference-free enantiomer separation. The suitability of using these two enantioselective multidimensional approaches for the routine assessment of chiral monoterpenes in TTO was evaluated and discussed. Exact enantiomeric composition of chiral markers for authentic TTOs was proposed by analysing a representative number of pure TTOs sourced directly from plantations of known provenance in Australia. Consistent enantiomeric fractions of 61.6±1.5% (+):38.4±1.5% (-) for limonene, 61.7±1.6% (+):38.3±1.6% (-) for terpinen-4-ol and 79.6±1.4% (+):20.4±1.4% (-) for α-terpineol were obtained for the 57 authentic Australian TTOs. The results were compared (using principle component analysis) with commercial TTOs (declared as derived from Melaleuca alternifolia) obtained from different continents. Assessing these data to determine adulteration, or additives that affect the enantiomeric ratios, in commercially sourced TTOs is discussed. The proposed method offers distinct advantages over e

  3. Simultaneous determination of some ultraviolet-absorbing chemicals in sunscreen cosmetics using a high-performance liquid chromatography method.

    PubMed

    Liu, T; Wu, D

    2011-10-01

    A method of gradient elution high-performance liquid chromatography (HPLC) for simultaneous determination of 11 different ultraviolet-absorbing chemicals of phenylbenzlmldazole sulphonic acid, 4-aminobenzoic acid, benzophenone-4, benzophenone-3, isoamyl p-methoxycinnamate, 4-methylbenzylidene camphor, octocrylene, ethylhexyl methoxycinnamate, homosalate, ethylhexyl salicylate, methylene bis-benzotriazolyl tetramethylbutyl phenol was developed for the application to sunscreen cosmetic products. In this study, an Agilent SB-C18 analytical column (250 × 4.6 mm, 5 μm) was utilized and methanol, tetrahydrofuran and perchloric acid aqueous solution (0.2 mL HClO(4) + 300 mL H(2)O) were used for gradient elution at a total flow rate of 1.0 mL min(-1). The optimum conditions for 11 different ultraviolet-absorbing chemicals analyses were investigated. All calibration curves showed good linear regression with UV detection (311 nm) within test ranges. The correlation coefficients were better than 0.999 in all cases. The assay was simple, selective, convenient and reproducible and is suitable for the determination of ultraviolet-absorbing chemicals in commercial sunscreen cosmetic products. The use frequency of 11 different ultraviolet absorbents in 100 sunscreen cosmetics was investigated and statistically analysed. The ultraviolet absorbent of maximum use frequency was ethylhexyl methoxycinnamate.

  4. Therapeutic monitoring of albendazole: a high-performance liquid chromatography method for determination of its active metabolite albendazole sulfoxide.

    PubMed

    Zeugin, T; Zysset, T; Cotting, J

    1990-03-01

    A sensitive and specific reversed-phase high-performance liquid chromatography (HPLC) method is described for the quantitative determination of albendazole sulfoxide (ASOX); since albendazole sulfone (ASON) appears only in small amounts and albendazole (ABZ) normally does not appear in human plasma, only a qualitative determination of ASON and ABZ was made in human plasma. Plasma samples were extracted three times using ethylacetate and petroleum benzine; this yielded optically clear samples which after evaporation were dissolved in the HPLC solvent and injected onto an RP-C18 column, with ultraviolet detection at 290 nm. The detection limit of the main metabolite ASOX was 50 nM and that of ASON was 100 nM. The intraday coefficient of variation for ASOX was 3.3% at a concentration of 2.2 microM, and the interday coefficients of variation were 14.5, 7.3, and 9.1% at ASOX concentrations of 0.5, 2.5, and 5.0 microM, respectively. Calibration was linear in a concentration range of 0.05-12 microM for ASOX and 0.1-8 microM for ASON, respectively. Pharmacokinetic data of a patient with echinococcosis are presented.

  5. [Determination of partition coefficient of dissolved gases in transformer oil using phase ratio variation method and static headspace gas chromatography].

    PubMed

    Zhao, Jinghong; Wang, Hailong; Liu, Wenmin; Zhou, Yansheng; Guan, Yafeng

    2004-05-01

    The partition coefficients of dissolved gases in transformer oil were determined using a phase ratio variation method and static headspace gas chromatography (GC). A pressure balancing and gas volume-metering device was connected to the vent of a sample loop on a six-port injection valve of the GC. The gas phase sample from the headspace vial of 25 mL was transferred to an 80 microL sample-loop through a fused silica capillary of 0.53 mm i.d., and then separated and determined quantitatively by GC. A 2 m x 1 mm i.d. GDX502 micro-packed column was used for the separation. Five different gas-liquid volume ratios in the headspace vials were measured at different equilibrium concentrations. The partition coefficients of hydrocarbon gases including methane, acetylene, ethylene, ethane and propane dissolved in transformer oil were determined by using linear regression analysis at 20 degrees C and 50 degrees C separately. The errors between the real values and regression values from experimental data were less than 4.14% except methane. Fundamental data for on-line measurement of dissolved gases in transformer oil are provided by GC.

  6. Chromatography of carbon nanotubes separated albumin from other serum proteins: a method for direct analysis of their interactions.

    PubMed

    Kuboki, Yoshinori; Koshikawa, Takamitu; Takita, Hiroko; Fujisawa, Ryuichi; Lee, Min-ho; Abe, Shigeaki; Akasaka, Tsukasa; Uo, Motohiro; Watari, Fumio; Sammons, Rachel

    2010-08-01

    Chromatography technology was employed to clarify the mechanism of interaction between multi-wall carbon nanotubes (MWCNT) and proteins. A column (16x100 mm) was packed with purified MWCNT, and various proteins were eluted with phosphate buffered saline (PBS) with and without gradient systems. It was found that albumin in bovine serum was eluted immediately from the column without any adsorption to MWCNT. Conversely, the non-albumin proteins, including a protein of 85 kDa molecular mass and a group of proteins with molecular masses higher than 115 kDa, exhibited considerably high affinity towards MWCNT. A sample of pure bovine serum albumin was also eluted immediately from the column, while lysozyme did not elute as a peak with PBS, but eluted with 0.6 M NaCl. Fundamentally, carbon nanotubes are devoid of any electrical charge. Therefore, other forces including the hydrogen bonds, hydrophilic interactions, and van der Waals forces were most probably responsible for the differential elution behaviors. In conclusion, this chromatographic method provided a simple and direct analysis of the interactions between carbon nanotubes and the various proteins.

  7. Vortex and air assisted liquid-liquid microextraction as a sample preparation method for high-performed liquid chromatography determinations.

    PubMed

    Hosseini, Mohammad; Heydari, Rouhollah; Alimoradi, Mohammad

    2014-12-01

    A novel, simple and sensitive method based on vortex and air assisted liquid-liquid microextraction (VAALLME) technique coupled with high-performance liquid chromatography (HPLC) has been developed for quantitative analysis of β-naphthol, naphthalene and anthracene as model analytes. Unlike the dispersive liquid-liquid microextraction (DLLME), dispersive solvent and centrifuging step were eliminated in proposed technique. In this technique, extraction solvent was dispersed into the aqueous sample solution by using vortex. Phase separation was achieved via motion of air bubbles from the bottom to top of the extraction tube, which promoted the analytes transfer into the supernatant organic phase. Influential parameters on the extraction efficiency such as type and volume of extraction solvent, salt type and its concentration, vortex and aeration times, and sample pH were evaluated and optimized. The calibration curves showed good linearity (r(2)>0.9947) and precision (RSD<5.0%) in the working concentration ranges. The limit of detection (LOD) for β-naphthol, naphthalene and anthracene were 10, 5.0 and 0.5 ng mL(-1), respectively. The recoveries were in the range of 97.0-102.0% with RSD values ranging from 2.2 to 5.2%.

  8. Micellar electrokinetic chromatography method for the determination of several natural red dyestuff and lake pigments used in art work.

    PubMed

    Maguregui, M I; Alonso, R M; Barandiaran, M; Jimenez, R M; García, N

    2007-06-22

    The identification of organic colorants used in artistic paintings is an important information source for reconstructing the working techniques found in a particular work and for defining a programme for the restoration and conservation of the painting. In this work, sodium dodecyl sulfate (SDS) was used as a surfactant in micellar electrokinetic chromatography (MEKC) for separating a broad range of red organic pigments, based on their colouring matters: madder (colouring matters: alizarin, quinizarin and purpurin), cochineal (colouring matter: carminic acid), red sandalwood (colouring matter: santalin), brazilwood (colouring matter: brazilin), lac dye (colouring matter: laccaic acid) and dragon's blood (colouring matter: dracorhodin). The running electrolyte used was 20 mM borax (pH 9), containing 20 mM SDS and 10% acetonitrile as organic modifier. Separation was carried out by applying a +20 kV voltage at the injection end, 25 degrees C and 214 nm/254 nm as detection wavelengths. All colorants were separated within less than 13 min with a good baseline resolution. The method was applied to the analysis of paint samples obtained from the Diocesan Museum of Holy Art of Bilbao.

  9. Characterization of volatile compounds in fermented milk using solid-phase microextraction methods coupled with gas chromatography-mass spectrometry.

    PubMed

    Dan, T; Wang, D; Jin, R L; Zhang, H P; Zhou, T T; Sun, T S

    2017-04-01

    Lactic acid bacteria (LAB) are industrially important bacteria that are widely used in the fermented food industry, especially in the manufacture of yogurt. Characteristic flavors are produced by LAB during fermentation and storage that affect the quality and acceptability of fermented milk products. In this study, the volatile compounds in milk fermented by Streptococcus thermophilus IMAU80842 alone, Lactobacillus delbrueckii ssp. bulgaricus IMAU20401 alone, or both species together were identified using solid-phase microextraction methods coupled with gas chromatography-mass spectrometry. A total of 53, 43, and 32 volatile compounds were identified in milk fermented by S. thermophilus alone, L. delbrueckii ssp. bulgaricus alone, or both species together, respectively. The presence of some important flavor compounds was confirmed: acetic acid, acetaldehyde, acetoin, 2,3-butanedione, ethanol, and 1-heptanol. Our results demonstrate that the composition of the volatile compounds in fermented milk depends on the species of LAB used and whether they are used alone or in combination. This is important for the selection of appropriate starter cultures for the production of different types of fermented milk product with particular flavors.

  10. A rapid liquid chromatography tandem mass spectrometry-based method for measuring propranolol on dried blood spots.

    PubMed

    Della Bona, Maria Luisa; Malvagia, Sabrina; Villanelli, Fabio; Giocaliere, Elisa; Ombrone, Daniela; Funghini, Silvia; Filippi, Luca; Cavallaro, Giacomo; Bagnoli, Paola; Guerrini, Renzo; la Marca, Giancarlo

    2013-05-05

    Propranolol, a non-selective beta blocker drug, is used in young infants and newborns for treating several heart diseases; its pharmacokinetics has been extensively evaluated in adult patients using extrapolation to treat pediatric population. The purpose of the present study was to develop and validate a method to measure propranolol levels in dried blood spots. The analysis was performed by using liquid chromatography/tandem mass spectrometry operating in multiple reaction monitoring mode. The calibration curve in matrix was linear in the concentration range of 2.5-200 μg/L with correlation coefficient r=0.9996. Intra-day and inter-day precisions and biases were less than 8.0% (n=10) and 11.5% (n=10) respectively. The recoveries ranged from 94 to 100% and the matrix effect did not result in a severe signal suppression. Propranolol on dried blood spot showed a good stability at three different temperatures for one month. This paper describes a micromethod for measuring propranolol levels on dried blood spot, which determines a great advantage in neonates or young infants during pharmacokinetic studies because of less invasive sampling and small blood volume required.

  11. Development and Validation of a Method for Alcohol Analysis in Brain Tissue by Headspace Gas Chromatography with Flame Ionization Detector.

    PubMed

    Chun, Hao-Jung; Poklis, Justin L; Poklis, Alphonse; Wolf, Carl E

    2016-10-01

    Ethanol is the most widely used and abused drug. While blood is the preferred specimen for analysis, tissue specimens such as brain serve as alternative specimens for alcohol analysis in post-mortem cases where blood is unavailable or contaminated. A method was developed using headspace gas chromatography with flame ionization detection (HS-GC-FID) for the detection and quantification of ethanol, acetone, isopropanol, methanol and n-propanol in brain tissue specimens. Unfixed volatile-free brain tissue specimens were obtained from the Department of Pathology at Virginia Commonwealth University. Calibrators and controls were prepared from 4-fold diluted homogenates of these brain tissue specimens, and were analyzed using t-butanol as the internal standard. The chromatographic separation was performed with a Restek BAC2 column. A linear calibration was generated for all analytes (mean r(2) > 0.9992) with the limits of detection and quantification of 100-110 mg/kg. Matrix effect from the brain tissue was determined by comparing the slopes of matrix prepared calibration curves with those of aqueous calibration curves; no significant differences were observed for ethanol, acetone, isopropanol, methanol and n-propanol. The bias and the CVs for all volatile controls were ≤10%. The method was also evaluated for carryover, selectivity, interferences, bench-top stability and freeze-thaw stability. The HS-GC-FID method was determined to be reliable and robust for the analysis of ethanol, acetone, isopropanol, methanol and n-propanol concentrations in brain tissue, effectively expanding the specimen options for post-mortem alcohol analysis.

  12. Development and validation of an ion chromatography method for the determination of phosphate-binding of lanthanum carbonate.

    PubMed

    Samy, Raghu; Faustino, Patrick J; Adams, Wallace; Yu, Lawrence; Khan, Mansoor A; Yang, Yongsheng

    2010-04-06

    Lanthanum carbonate is indicated to reduce serum phosphate in patients with end stage renal disease (ESRD). When given orally, lanthanum carbonate dissociates in the acid environment of the upper gastrointestinal tract to release lanthanum ions. The free lanthanum ions bind with dietary phosphate released from food during digestion to form highly insoluble lanthanum-phosphate complexes which prevent the absorption of phosphate, consequently reduce the serum phosphate. In order to evaluate the in vitro binding capacity of lanthanum carbonate, a simple and efficient ion chromatography (IC) method was developed and validated for determination of phosphate across the pH range encountered in the gastrointestinal tract. Chromatographic separation was achieved on a Dionex ICS-2000 IC system using a Dionex AS16, IonPac (4mmx250mm) analytical column and Dionex AG16, IonPac (4mmx50mm) guard column. Column temperature was maintained at 30 degrees C. Injection volume was 10microL. The compounds were eluted isocratically at a flow rate of 1mL/min and detected by suppressed conductivity. The analytical method was validated according to USP Category I requirements. The validation characteristics included accuracy, precision, quantification limit, linearity, and stability. The intra-day accuracy ranged from 89% to 103% for the solutions of pH 1.2-6.8. The intra-day precision (RSD) ranged from 0.6% to 3.7% for the solutions of pH 1.2-6.8. The analytical range was linear from 2 to 200ppm (mg/L). The R(2) ranged from 0.9998 to 1.0. This method was found to be simple, robust, sensitive, specific, and accurate. It has been successfully applied for determination of phosphate binding to lanthanum carbonate over the human gastrointestinal pH range at different time-points (from 0.5 to 24h).

  13. Method for quantification of opioids and their metabolites in autopsy blood by liquid chromatography-tandem mass spectrometry.

    PubMed

    Al-Asmari, Ahmed I; Anderson, Robert A

    2007-09-01

    A method using liquid chromatography-electrospray ionization-tandem mass spectrometry was developed and validated for the determination of morphine, codeine, hydromorphone, dihydrocodeine, oxycodone, buprenorphine, and naloxone with their metabolites morphine-3-glucuronide, morphine-6-glucuronide, normorphine, 6-acetylmorphine, 6-acetylcodeine, codeine-6-glucuronide, norcodeine, hydromorphine-3-glucuronide, dihydrocodeine-6-glucuronide, dihydromorphine, dihydromorphine-3-glucuronide, dihydromorphine-6-glucuronide, oxymorphone, norbuprenorphine, buprenorphine-3-glucuronide, norbuprenorphine-3-glucuronide, and naloxone-3-glucuronide in human whole blood. Polar metabolites (glucuronides) and other analytes were extracted by SPE using Bond Elut C18. Chromatographic separation was performed on a Phenomenex Synergi reversed-phase column with gradient elution based on a mobile phase consisting of 10mM ammonium formate adjusted to pH 3 and acetonitrile. Intraday and interday precision for all analytes were between 0.6% and 13.8%, and recoveries were between 80.3% and 101.4%. Calibration curves were linear for all analytes over the concentration range 5-400 ng/mL, and correlation coefficients (R(2)) were better than 0.999. Limits of detection and quantitation were 0.16-1.2 ng/mL and 0.5-4.09 ng/mL, respectively. The method described consolidates previous work on opioids and their metabolites published in the literature and is the first to include the detection of naloxone-3-glucuronide. The method has been applied in routine postmortem cases after opiate overdose with the threefold purpose of providing interpretive information on the cause and type of death (rapid, sub-acute, or delayed death) and to distinguish heroin, morphine, and codeine users.

  14. Development and validation of a gas chromatography/mass spectrometry method for the assessment of genomic DNA methylation.

    PubMed

    Rossella, Federica; Polledri, Elisa; Bollati, Valentina; Baccarelli, Andrea; Fustinoni, Silvia

    2009-09-01

    A method for the determination of DNA global methylation, taken as the ratio (%) of 5-methylcytosine (5mCyt) versus the sum of cytosine (Cyt) and 5mCyt, via gas chromatography/mass spectrometry (GC/MS), was developed and validated. DNA (2.5 microg) was hydrolyzed with aqueous formic acid 88%, spiked with cytosine-2,4-(13)C(2),(15)N(3) and 5-methyl-(2)H(3)-cytosine-6-(2)H(1) as internal standards, and derivatized with N-methyl-N-(tert-butyldimethylsilyl)trifluoroacetamide and 1% tert-butyldimethylchlorosilane, in the presence of acetonitrile and pyridine. GC/MS, operating in single ion monitoring mode, separated and specifically detected all nucleobases as tert-butyldimethylsilyl derivatives, without interferences, with the exception of guanosine. The method was linear throughout the range of clinical interest and had good sensitivity, with a limit of quantification of 3.2 pmol for Cyt and 0.056 pmol for 5mCyt, the latter corresponding to a methylation level of 0.41%. Intra- and inter-day precision and accuracy were below 4.0% for both analytes and methylation. The matrix absolute effect, process efficiency and coefficient of variation ranged from 96.5 to 101.2%. The matrix relative effect was below 1%. The method was applied to the analysis of different human DNAs, including: nonmethylated DNA from PCR (methylation 0.00%), hypermethylated DNA prepared using M.SssI CpG methyltransferase (methylation 18.05%), DNA from peripheral blood leukocytes of healthy subjects (N = 6, median methylation 5.45%), DNA from bone marrow of leukemia patients (N = 5, 3.58%) and DNA from myeloma cell lines (N = 4, 2.74%).

  15. A capillary liquid chromatography method for benzalkonium chloride determination as a component or contaminant in mixtures of biocides.

    PubMed

    Prieto-Blanco, M C; Argente-García, A; Campíns-Falcó, P

    2016-01-29

    A method for quantifying benzalkonium chloride (BAK), an alkyl dimethyl benzyl ammonium compound, in several biocides formulations is proposed. A tertiary amine like N-(3-aminopropyl)-N-dodecyl-1,3-propanediamine (TA) and a straight-chain alkyl ammonium compound like trimethyl-tetradecyl ammonium chloride (TMTDAC), have been employed as trade surfactants besides BAK. Two capillary analytical columns with different polarities are tested: inertsil CN-3 capillary column (150mm×0.5mm i.d., 3μm particle diameter) and a non endcapped Zorbax C18 capillary column (35mm×0.5mm i.d., 5μm particle diameter). This latter column provided the best separation of the BAK homologues in less than 12min using acetonitrile:acetate buffer (50mM, pH 5) 85:15 at 20μLmin(-1). The proposed method combines on-line in-tube solid-phase microextraction (IT-SPME) coupled to capillary liquid chromatography (CapLC) and UV diode array detection. Matrix effect was present when TA were in excess to BAK. If TMTDAC is the co-biocide, matrix effect is always present. A decreasing of analytical response mainly for C12-BAK homologue was found using both chromatographic columns. The charged amount of mixture in the system was the most important parameter for obtaining reliable results. 1mL was the on line processed sample volume optimum for concentrations lower than 35μgmL(-1) of total surfactants. LODs were 0.03μgmL(-1) and 0.006μgmL(-1) for C12-BAK and C14-BAK, respectively. This method is also of use to evaluate the unwanted presence of BAK in biocide formulations due to industrial processes.

  16. Universal method for online enrichment of target compounds in capillary gas chromatography using in-oven cryotrapping.

    PubMed

    Chin, Sung-Tong; Maikhunthod, Bussayarat; Marriott, Philip J

    2011-09-01

    A method is described that permits automated online enrichment of injected compounds in multidimensional gas chromatography by using a microfluidic heart-cut (H-C) device to direct target compounds into a cryogenically cooled internal trap (cryotrap, CT). By performing multiple injections of a sample, selected compounds or regions of a primary column separation can be collected in the CT. Remobilizing the trapped species allows elution and further resolution on the second column. Using a well-balanced H-C device, compounds can be fully excluded from the collection step or quantitatively transferred to the CT. Peak areas of the remobilized compound correlate well with the number of sample injections. Trapping on various column phases shows the method is suited to quantitative trapping of alkanes of mass greater than about dodecane and fatty acid methyl esters greater than the C8 homologue. Caffeine and menthol standards of concentration 100 μg mL(-1) gave peak area correlation coefficients for 1-10 and 1-50 replicate split injections of 1 μL volume of 0.999 and 0.996, respectively. Peak height correlations were less favorable as a result of peak broadening on the second column, presumably due to overloading at greater collected mass. The method was applied to 0.2% solutions of peppermint oil (menthol; a major component; 44%) and 1.0% lavender oil (α-terpineol and neryl acetate; minor components of 1.05 and 0.42% abundance). The minor components gave good area and height correlations, and good recovery around 90% was observed for menthol compounds recovered from 15 accumulations. Response amplification was further demonstrated for menthol from mint oil headspace sampling using solid phase microextraction. This approach should be a valuable adjunct for improved detection specificity, for detectors of low sensitivity, and when prior sample concentration provides insufficient response of selected target analytes.

  17. Analytical performance of a new liquid chromatography/tandem mass spectrometric method for determination of everolimus concentrations in whole blood.

    PubMed

    McMillin, Gwendolyn A; Johnson-Davis, Kamisha; Dasgupta, Amitava

    2012-04-01

    The immunosuppressant everolimus was recently approved for prophylactic use in the United States, to prevent organ rejection in adult kidney transplant recipients. The currently accepted therapeutic range for everolimus is 3-8 ng/mL. Therapeutic drug monitoring (TDM) using predose EDTA whole blood samples is required to optimize dose. We describe a simple extraction method and analysis by liquid chromatography tandem mass spectrometry (LC-MS/MS) to support routine TDM of everolimus. Samples were prepared by protein precipitation and filtration. The first quadrupole was set to select the ammonium adducts (Equation is included in full-text article.)of everolimus (m/z 975.62) and rapamycin-d3 (m/z 934.70), the internal standard. The second quadrupole was used as a collision chamber, and the third quadrupole was then used to select characteristic product ions of everolimus (m/z 908.50 and 890.50) and rapamycin-d3 (m/z 864.60 and 846.50). The method had an analytical measurement range of 2.0-150 ng/mL. Total imprecision, expressed as percent coefficient of variation (mean concentration), was 19.1% (3.3 ng/mL), 10.6% (5.9 ng/mL), 8.1% (19.2 ng/mL), 5.7% (25.8 ng/mL), and 9.1% (34.2 ng/mL). The new method was compared with 2 other everolimus methods also based on LC-MS/MS, with 64 residual patient specimens. Agreement, based on simple linear regression, was excellent. Method A comparison: y = 0.96x - 1.12 (r = 0.99), n = 20, 2.5-44.7 ng/mL. Method B comparison: y = 0.96x + 0.49 (r = 0.99), n = 44, 2.1-85.6 ng/mL. We conclude that this method could support TDM of everolimus for a wide range of clinical indications.

  18. Screening anti-allergic components of Astragali Radix using LAD2 cell membrane chromatography coupled online with UHPLC-ESI-MS/MS method.

    PubMed

    Lv, Yanni; Sun, Yueming; Fu, Jia; Kong, Liyun; Han, Shengli

    2017-02-01

    Huangqi (Astragali Radix), a traditional Chinese herb, is widely used in clinical therapy in China. In addition, an anti-allergic effect of constituents in Huangqi has been reported in the scientific literature. In the present study, cell membrane chromatography coupled online with UHPLC-ESI-MS/MS method was developed to screen, analyze and identify the anti-allergic components of Huangqi. The Laboratory of Allergic Disease 2 (LAD2) cell was used to establish cell membrane chromatography, which was combined with UHPLC-ESI-MS/MS. The coupled system was then used to screen anti-allergic components from Huangqi. Effects of active components were verified by histamine release assay. A component retained on the LAD2 cell membrane chromatography was identified as formononetin. Bioactivity of formononetin was investigated by histamine release assay in LAD2 cells, and it was found that formononetin could inhibit histamine release in a dose-dependent manner from 1 to 100 μm. The LAD2 cell membrane chromatography online with UHPLC-ESI-MS/MS method is an effective technique for screening the anti-allergic components of Huangqi.

  19. Development of a method based on on-line reversed phase liquid chromatography and gas chromatography coupled by means of an adsorption-desorption interface for the analysis of selected chiral volatile compounds in methyl jasmonate treated strawberries.

    PubMed

    de la Peña Moreno, Fernando; Blanch, Gracia Patricia; Flores, Gema; Ruiz Del Castillo, Maria Luisa

    2010-02-12

    A method based on the use of the through oven transfer adsorption-desorption (TOTAD) interface in on-line coupling between reversed phase liquid chromatography and gas chromatography (RPLC-GC) for the determination of chiral volatile compounds was developed. In particular, the method was applied to the study of the influence of methyl jasmonate (MJ) treatment on the production and enantiomeric composition of selected aroma compounds in strawberry. The compounds studied were ethyl 2-methylbutanoate, linalool and 4-hydroxy-2,5-dimethyl-3(2H)-furanone (i.e. furaneol), which were examined on days 3, 6 and 9 after treatment. The method developed resulted in relative standard deviations (RSDs) of 21.6%, 8.1% and 9.8% and limits of detection (LD) of 0.04, 0.07 and 0.02mg/l for ethyl 2-methylbutanoate, linalool and furaneol, respectively. The application of the RPLC-TOTAD-GC method allowed higher levels of ethyl 2-methylbutanoate, linalool and furaneol to be detected, particularly after 9 days of treatment. Besides, MJ demonstrated to affect the enantiomeric distribution of ethyl 2-methylbutanoate. On the contrary, the enantiomeric composition of linalool and furaneol kept constant in both control and MJ-treated strawberries throughout the study. These results are discussed.

  20. Method development and validation for rapid quantification of hydroxychloroquine in human blood using liquid chromatography-tandem mass spectrometry.

    PubMed

    Wang, Ling-Zhi; Ong, Rina Yue-Ling; Chin, Tan-Min; Thuya, Win-Lwin; Wan, Seow-Ching; Wong, Andrea Li-Ann; Chan, Sui-Yung; Ho, Paul C; Goh, Boon-Cher

    2012-03-05

    A novel and specific liquid chromatography-tandem mass spectrometric method (LC-MS/MS) was developed and validated for the quantification of hydroxychloroquine in human blood using its stable labeled isotope, hydroxychloroquine-d4 as the internal standard. Chromatographic separation of analytes was achieved using an Agilent ZORBAX Eclipse XDB - C8 analytical HPLC column (50 mm × 2.1 mm, 5 μm). The mobile phase comprising water containing 0.1% formic acid-acetonitrile (94:6, v/v) was delivered isocratically at a flow rate of 0.5 mL/min. The column effluent was detected by API 4000 triple quadrupole mass spectrometer using electrospray ionization (ESI) and monitored by multiple reaction monitoring with positive mode. The precursor to product ion transitions of m/z 336 → 247 and m/z 340 → 251 were used to measure the analyte and IS, respectively. The assay demonstrated a good linear dynamic range of 5-2000 ng/mL for hydroxychloroquine in human blood, with coefficient of determination (r(2)) of =0.9999. The values for intra-day and inter-day precisions of hydroxychloroquine were ≤ 7.86% with the accuracies ranged from 93.8% to 107.6%. The chromatographic run time was 3 min, making it possible to achieve a high throughput analysis. This method was used as a bio-analytical tool in a phase I clinical trial to quantify blood hydroxychloroquine concentrations in patients with non-small cell lung cancer receiving both hydroxychloroquine and gefitinib in their treatment.

  1. Aroma composition of red wines by different extraction methods and Gas Chromatography-SIM/MASS spectrometry analysis.

    PubMed

    Genovese, Alessandro; Dimaggio, Rosa; Lisanti, Maria Tiziana; Piombino, Paola; Moio, Luigi

    2005-06-01

    One hundred and one volatile compounds, reported in literature as powerful odorants of wine, were quantified by Gas Chromatography-Selective Ion Monitoring/Mass Spectrometry (GC-SIM/MS) in Primitivo, Aglianico, Merlot and Cabernet Sauvignon red wines. Wine samples were extracted by 3 different extraction methods: 1) separation of the alcoholic fraction from the aqueous phase by salting-out and subsequent extraction by liquid-liquid micro-extraction with 1,1,2-trichlorotrifluoroethane (Freon 113); 2) extraction by liquid-liquid micro-extraction with dichloromethane; 3) solid phase extraction (SPE cartridge: 800 mg of LiChrolut EN resin) with pentane-dichloromethane (20:1) and dichloromethane. The selection of the ion fragments used for quantification was directly performed on a red wine sample. For each compound the area of the corresponding peak was normalized respect to the peak of the internal standard and then interpolated in a calibration curve obtained analysing a model wine solution (water, ethanol, tartaric acid and known amounts of analytes and of internal standard). The methods showed a good linearity: r2>0.990, except for farnesol (isomer a and c), octanal, decanal, furaneol and phenylacetic acid with 0.966 < or = r2 < or = 0.990. The 7 most powerful odorants were: beta-damascenone, acetaldehyde, maltol, ethyl 2-methylbutanoate, ethyl 3-methylbutanoate, 3-methylbutanoic acid and acetal; 7 other slightly less important were: ethyl hexanoate, ethyl acetate, 1-octen-3-ol, butanoic acid, rose oxide, furaneol and sotolon. The Aglianico wines were characterised by the major fermentation compounds (esters, fatty acids and 2-phenylethanol), beta-damascenone, beta-ionone and linalool. The Primitivo wines were characterized by furaneol, methoxypyrazine, gamma-nonalactone and acetaldehyde, while Cabernet Sauvignon and Merlot wines principally by cask derivates (vanillin, (Z) 3-methyl-gamma-octalactone [(Z) wiskylactone], maltol and eugenol), some aldehydes and 3

  2. Development of an analytical method coupling cell membrane chromatography with gas chromatography-mass spectrometry via microextraction by packed sorbent and its application in the screening of volatile active compounds in natural products.

    PubMed

    Li, Miao; Wang, Sicen; He, Langchong

    2015-01-01

    Natural products (NPs) are important sources of lead compounds in modern drug discovery. To facilitate the screening of volatile active compounds in NPs, we have developed a new biochromatography method that uses rat vascular smooth muscle cells (VSMC), which are rich in L-type calcium channels (LCC), to prepare the stationary phase. This integrated method, which couples cell membrane chromatography (CMC) with gas chromatography-mass spectrometry (GC-MS) via microextraction by packed sorbent (MEPS) technology, has been termed VSMC/CMC-MEPS-GC-MS. Methodological validation confirmed its specificity, reliability and convenience. Screening results for Radix Angelicae Dahuricae and Fructus Cnidii obtained using VSMC/CMC-MEPS-GC-MS were consistent with those obtained using VSMC/CMC-offline-GC-MS. MEPS connection plays as simplified solid-phase extraction and replaces the uncontrollable evaporation operation in reported offline connections, so our new method is supposed to be more efficient and reliable than the offline ones, especially for compounds that are volatile, thermally unstable or difficult to purify. In application, senkyunolide A and ligustilide were preliminary identified as the volatile active components in Rhizoma Chuanxiong. We have thus confirmed the suitability of VSMC/CMC-MEPS-GC-MS for volatile active compounds screening in NP.

  3. Enhanced high-performance liquid chromatography method for the determination of retinoic acid in plasma. Development, optimization and validation.

    PubMed

    Teglia, Carla M; Gil García, María D; Galera, María Martínez; Goicoechea, Héctor C

    2014-08-01

    When determining endogenous compounds in biological samples, the lack of blank or analyte-free matrix samples involves the use of alternative strategies for calibration and quantitation. This article deals with the development, optimization and validation of a high performance liquid chromatography method for the determination of retinoic acid in plasma, obtaining at the same time information about its isomers, taking into account the basal concentration of these endobiotica. An experimental design was used for the optimization of three variables: mobile phase composition, flow rate and column temperature through a central composite design. Four responses were selected for optimization purposes (area under the peaks, quantity of peaks, analysis time and resolution between the first principal peak and the following one). The optimum conditions resulted in a mobile phase consisting of methanol 83.4% (v/v), acetonitrile 0.6% (v/v) and acid aqueous solution 16.0% (v/v); flow rate of 0.68 mL min(-1) and an column temperature of 37.10 °C. Detection was performed at 350 nm by a diode array detector. The method was validated following a holistic approach that included not only the classical parameters related to method performance but also the robustness and the expected proportion of acceptable results lying inside predefined acceptability intervals, i.e., the uncertainty of measurements. The method validation results indicated a high selectivity and good precision characteristics that were studied at four concentration levels, with RSD less than 5.0% for retinoic acid (less than 7.5% for the LOQ concentration level), in intra and inter-assay precision studies. Linearity was proved for a range from 0.00489 to 15.109 ng mL(-1) of retinoic acid and the recovery, which was studied at four different fortification levels in phuman plasma samples, varied from 99.5% to 106.5% for retinoic acid. The applicability of the method was demonstrated by determining retinoic acid and

  4. Determination of vegetable oils and fats adulterants in diesel oil by high performance liquid chromatography and multivariate methods.

    PubMed

    Brandão, Luiz Filipe Paiva; Braga, Jez Willian Batista; Suarez, Paulo Anselmo Ziani

    2012-02-17

    The current legislation requires the mandatory addition of biodiesel to all Brazilian road diesel oil A (pure diesel) marketed in the country and bans the addition of vegetable oils for this type of diesel. However, cases of irregular addition of vegetable oils directly to the diesel oil may occur, mainly due to the lower cost of these raw materials compared to the final product, biodiesel. In Brazil, the situation is even more critical once the country is one of the largest producers of oleaginous products in the world, especially soybean, and also it has an extensive road network dependent on diesel. Therefore, alternatives to control the quality of diesel have become increasingly necessary. This study proposes an analytical methodology for quality control of diesel with intention to identify and determine adulterations of oils and even fats of vegetable origin. This methodology is based on detection, identification and quantification of triacylglycerols on diesel (main constituents of vegetable oils and fats) by high performance liquid chromatography in reversed phase with UV detection at 205nm associated with multivariate methods. Six different types of oils and fats were studied (soybean, frying oil, corn, cotton, palm oil and babassu) and two methods were developed for data analysis. The first one, based on principal component analysis (PCA), nearest neighbor classification (KNN) and univariate regression, was used for samples adulterated with a single type of oil or fat. In the second method, partial least square regression (PLS) was used for the cases where the adulterants were mixtures of up to three types of oils or fats. In the first method, the techniques of PCA and KNN were correctly classified as 17 out of 18 validation samples on the type of oil or fat present. The concentrations estimated for adulterants showed good agreement with the reference values, with mean errors of prediction (RMSEP) ranging between 0.10 and 0.22% (v/v). The PLS method was

  5. Gas Chromatography.

    ERIC Educational Resources Information Center

    Karasek, Francis W.; And Others

    1984-01-01

    This review covers fundamental developments in gas chromatography during 1982 and 1983. Literature is considered under these headings: columns; liguid phases; solid supports; sorption processes and solvents; open tubular column gas chromatography; instrumentation; high-resolution columns and applications; other techniques; qualitative and…

  6. Application of Design of Experiment and Simulation Methods to Liquid Chromatography Analysis of Topical HIV Microbicides Stampidine and HI443.

    PubMed

    Agrahari, Vivek; Meng, Jianing; Zhang, Tao; Youan, Bi-Botti C

    2014-02-01

    This study intended to determine if experimental design and Monte Carlo simulation methods can be utilized to optimize the liquid chromatography (LC) analysis of active molecules. The method was applied for the simultaneous analysis of two topical microbicides, stampidine (STP) and HI443 in bulk and nanoformulations. The Plackett-Burman design was used for screening; whereas, Box-Behnken design was used to evaluate the main and interaction effects of the selected factors on the responses, namely peak area of STP (Y1), HI443 (Y2), tailing of STP (Y3), and HI443 (Y4). The Monte Carlo simulation was applied to get the minimum defect rate (DR) of the process. The optimized LC conditions were found to be X1; flow rate: 0.6 mL/min, X2; injection volume: 18 μL, and X3; initial gradient acetonitrile ratio: 92% v/v with a minimal DR of 0.077%. The optimized method was applied to determine the percent encapsulation efficiency (%EE) and in vitro release profile of STP and HI443 from solid lipid nanoparticles (SLNs). The %EE of STP and HI443 in SLNs was found to be 30.56 ± 9.44 and 94.80 ± 21.90% w/w, respectively, (n=3). It was observed that the release kinetics of STP followed the first order, whereas, HI443 followed the Peppas kinetic model in SLNs. The LC method was also applied for the estimation of molar extinction coefficients (ε270 ) of both drugs for the first time. These values were estimated to be 7,569.03 ± 217.96 and 17,823.67 ± 88.12 L/mol/cm for STP and HI443, respectively, (n=3). The results suggest that experimental design and Monte Carlo simulation can be effectively used to reduce the DR of a process and to optimize the chromatographic conditions for the analysis of bio-active agents as applied in this study.

  7. A simultaneous quantitative method for vitamins A, D and E in human serum using liquid chromatography-tandem mass spectrometry.

    PubMed

    Albahrani, Ali A; Rotarou, Victor; Roche, Peter J; Greaves, Ronda F

    2016-05-01

    Non-classical roles of fat-soluble vitamins (FSVs) in many pathologies including cancer have been identified. There is also evidence of hormonal interactions between two of these vitamins, A and D. As a result of this enhanced clinical association with disease, translational clinical research and laboratory requests for FSV measurement has significantly increased. However there are still gaps in the analytical methods available for the measurement of these vitamins. This study aimed to develop a method for simultaneous quantification of 25-hydroxyvitamin-D2 (25-OHD2), 25-hydroxyvitamin-D3 (25-OHD3) and its 3-epimer (epi-25-OHD3), retinol and α-tocopherol in human serum using liquid chromatography-tandem mass spectrometry (LC-MS/MS). The procedure was developed and validated across two LC-MS/MS platforms, using commercial calibrators referenced to certified reference materials, controls, and deuterated internal standards. The samples were prepared by liquid-liquid extraction prior to injection and LC separation (using a Pursuit-PFP column) on two Agilent MS/MS systems (6410 and 6490) in electrospray ionisation positive mode with multiple reaction monitoring. Identification and quantification of 25-OHD3 from its 3-epimer as well as 25-OHD2, retinol and α-tocopherol were achieved. The dynamic ranges were 4-160 nmol/L for 25-OHD2 and epi-25-OHD3, 4-200 nmol/L for 25-OHD3, 0.1-4.0μmol/L for retinol and 4-70μmol/L for α-tocopherol with correlation (r(2)) of 0.997-0.998. Based on participation in an external quality assurance program, the overall performance of the simultaneous methods were: imprecision (CV%) and inaccuracy (average bias) 3.0% and 3.2 nmol/L, respectively, for 25-OHD3; 5.0% and 0.04μmol/L, respectively, for retinol; and 4.7% and 0.2μmol/L, respectively, for α-tocopherol. In summary, two simple LC-MS/MS methods were successfully developed and validated for the simultaneous quantification of the three vitamin D metabolites (25-OHD2, 25-OHD3 and 3

  8. Serum bilirubin fractionation using multilayer film method in hepatobiliary diseases in comparison with high performance liquid chromatography.

    PubMed

    Adachi, Y; Inufusa, H; Yamashita, M; Ozaki, K; Kanbe, A; Nanno, T; Ohba, Y; Nakamura, H; Yamamoto, T

    1987-10-01

    The Ektachem multilayer film method (Ektachem) and high performance liquid chromatography (HPLC) were employed to fractionate and evaluate serum bilirubin species in 45 serum samples. The false-positive or false-high levels of bilirubin close-bonded with albumin (i.e. the delta bilirubin fraction (B delta] was obtained by Ektachem in sera of cases with normal bilirubin concentration and cases with unconjugated hyperbilirubinemia when compared with the results of HPLC. In the sera of cases with conjugated hyperbilirubinemia, Ektachem gave comparable levels of total bilirubin (TB), and unconjugated bilirubin (Bu) to those of HPLC, but underestimated conjugated bilirubin (Bc) and slightly overestimated B delta. To investigate the clinical significance of B delta in 113 cases of various hepatobiliary diseases with conjugated hyperbilirubinemia, the ratios of B delta to TB (B delta/TB) and to directly-reacting bilirubin fractions (B delta/(Bc + B delta] and that of Bc to B delta (Bc/B delta) were calculated based on the results of Ektachem and compared with each other during the course of jaundice. The mean B delta/TB was below 40% in various hepatobiliary diseases but became as high as approximately 60% in the convalescence stage. The mean B delta/(Bc + B delta) was below 50% in acute hepatitis (the serum bilirubin-elevating stage) and obstructive jaundice, and it increased to above 80% in the recovery stage. In decompensated liver cirrhosis and intrahepatic cholestasis the mean B delta/(Bc + B delta) was about 60%, indicating continuous backflow of Bc from liver cells. The changes in B delta/(Bc + B delta) were much greater than in B delta/TB.(ABSTRACT TRUNCATED AT 250 WORDS)

  9. Influence of Different Shellfish Matrices on the Separation of PSP Toxins Using a Postcolumn Oxidation Liquid Chromatography Method

    PubMed Central

    Rey, Verónica; Alfonso, Amparo; Botana, Luis M.; Botana, Ana M.

    2015-01-01

    The separation of PSP toxins using liquid chromatography with a post-column oxidation fluorescence detection method was performed with different matrices. The separation of PSP toxins depends on several factors, and it is crucial to take into account the presence of interfering matrix peaks to produce a good separation. The matrix peaks are not always the same, which is a significant issue when it comes to producing good, reliable results regarding resolution and toxicity information. Different real shellfish matrices (mussel, scallop, clam and oyster) were studied, and it was seen that the interference is not the same for each individual matrix. It also depends on the species, sampling location and the date of collection. It was proposed that separation should be accomplished taking into account the type of matrix, as well as the concentration of heptane sulfonate in both solvents, since the mobile phase varies regarding the matrix. Scallop and oyster matrices needed a decrease in the concentration of heptane sulfonate to separate GTX4 from matrix peaks, as well as dcGTX3 for oysters, with a concentration of 6.5 mM for solvent A and 6.25 mM for solvent B. For mussel and clam matrices, interfering peaks are not as large as they are in the other group, and the heptane sulfonate concentration was 8.25 mM for both solvents. Also, for scallops and oysters, matrix interferences depend not only on the sampling site but also on the date of collection as well as the species; for mussels and clams, differences are noted only when the sampling site varies. PMID:25884908

  10. Validation of the Mass-Extraction-Window for Quantitative Methods Using Liquid Chromatography High Resolution Mass Spectrometry.

    PubMed

    Glauser, Gaétan; Grund, Baptiste; Gassner, Anne-Laure; Menin, Laure; Henry, Hugues; Bromirski, Maciej; Schütz, Frédéric; McMullen, Justin; Rochat, Bertrand

    2016-03-15

    A paradigm shift is underway in the field of quantitative liquid chromatography-mass spectrometry (LC-MS) analysis thanks to the arrival of recent high-resolution mass spectrometers (HRMS). The capability of HRMS to perform sensitive and reliable quantifications of a large variety of analytes in HR-full scan mode is showing that it is now realistic to perform quantitative and qualitative analysis with the same instrument. Moreover, HR-full scan acquisition offers a global view of sample extracts and allows retrospective investigations as virtually all ionized compounds are detected with a high sensitivity. In time, the versatility of HRMS together with the increasing need for relative quantification of hundreds of endogenous metabolites should promote a shift from triple-quadrupole MS to HRMS. However, a current "pitfall" in quantitative LC-HRMS analysis is the lack of HRMS-specific guidance for validated quantitative analyses. Indeed, false positive and false negative HRMS detections are rare, albeit possible, if inadequate parameters are used. Here, we investigated two key parameters for the validation of LC-HRMS quantitative analyses: the mass accuracy (MA) and the mass-extraction-window (MEW) that is used to construct the extracted-ion-chromatograms. We propose MA-parameters, graphs, and equations to calculate rational MEW width for the validation of quantitative LC-HRMS methods. MA measurements were performed on four different LC-HRMS platforms. Experimentally determined MEW values ranged between 5.6 and 16.5 ppm and depended on the HRMS platform, its working environment, the calibration procedure, and the analyte considered. The proposed procedure provides a fit-for-purpose MEW determination and prevents false detections.

  11. Application of counter-current chromatography as a new pretreatment method for the determination of polycyclic aromatic hydrocarbons in environmental water.

    PubMed

    Cao, Xueli; Yang, Chunlei; Pei, Hairun; Li, Xinghong; Xu, Xiaobai; Ito, Yoichiro

    2012-02-01

    Counter-current chromatography (CCC) was investigated as a new sample pretreatment method for the determination of trace polycyclic aromatic hydrocarbons (PAHs) in water environmental samples. The experiment was performed with a non-aqueous binary two-phase solvent system composed of n-heptane and acetonitrile. The CCC column was first filled with the upper stationary phase, and then a large volume of water sample was pumped into the column while the CCC column was rotated at 1600 rpm. Finally, the trace amounts of PAHs extracted and enriched in the stationary phase were eluted out by the lower mobile phase and determined by gas chromatography-flame ionization detector (GC-FID) or gas chromatography-mass spectrometry (GC-MS). The enrichment and cleanup of PAHs can be fulfilled online by this method with high recoveries (84.1-103.2%) and good reproducibility (RSDs: 4.9-12.2%) for 16 EPA PAHs under the optimized CCC pretreatment conditions. This method has been successfully applied to determine PAHs in lake water where 8 PAHs were detected in the concentration of 40.9-89.9 ng/L. The present method is extremely suitable for the preparation of large volume of environmental water sample for the determination of trace amounts of organic pollutants including PAHs as studied in this paper.

  12. Quality by Design: Multidimensional exploration of the design space in high performance liquid chromatography method development for better robustness before validation.

    PubMed

    Monks, K; Molnár, I; Rieger, H-J; Bogáti, B; Szabó, E

    2012-04-06

    Robust HPLC separations lead to fewer analysis failures and better method transfer as well as providing an assurance of quality. This work presents the systematic development of an optimal, robust, fast UHPLC method for the simultaneous assay of two APIs of an eye drop sample and their impurities, in accordance with Quality by Design principles. Chromatography software is employed to effectively generate design spaces (Method Operable Design Regions), which are subsequently employed to determine the final method conditions and to evaluate robustness prior to validation.

  13. Simple Detection Methods for Antinutritive Factor β-ODAP Present in Lathyrus sativus L. by High Pressure Liquid Chromatography and Thin Layer Chromatography.

    PubMed

    Ghosh, Bidisha; Mitra, Joy; Chakraborty, Saikat; Bhattacharyya, Jagannath; Chakraborty, Anirban; Sen, Soumitra Kumar; Neerathilingam, Muniasamy

    2015-01-01

    Lathyrus sativus L. (Grass pea) is the source for cheap and nutritious food choice in drought and famine susceptible zones in greater part of North India and Africa. The non-protein amino acid β-N-oxalyl-L-α,β-diaminopropionic acid (β-ODAP) has been known for decades for its potent neurotoxic effect, causing irreversible neurodegenerative disease "neurolathyrism", present in both seed and leaf of Lathyrus sativus L. and other species in varying proportions. It is crucial to establish a rapid as well as reliable detection methodology for β-ODAP content in various Lathyrus plants. Currently available HPLC based methods involve multi-step derivatization of the sample. To overcome this, we have developed β-ODAP analysis method by HPLC without any prior derivatization. This method is statistically significant in the range of 2 to 100μg/ml and exhibited linear response with r2 > 0.99. Limit of detection and quantitation of the later method was determined to be 5.56 μg/ml and 16.86 μg/ml, respectively. In addition to this, a TLC based method has also been developed. The limit of detection of β-ODAP is 0.6μg and for its substrate, L-1,2-diaminopropionic acid is 5μg. Both HPLC and TLC methods were validated by conducting in-vitro bioconversion test to detect the presence of biocatalyst in plant extract. This method is economical, rapid and simple.

  14. [Comparison of standard methods for determination of pseudocumene in urine using gas chromatography with the headspace technique and a new method using a headspace automatic sampler].

    PubMed

    Kostrzewski, P; Wiaderna-Brycht, A; Czerski, B

    1996-01-01

    The biological indicators that have been proposed for monitoring occupational exposure are: concentration of the solvent or metabolized compounds in alveolar or expired air samples, in venous or arterial capillary blood samples and in urine samples. Recently, many researches have reported significant relationships between the time-weighted average exposure and the urinary concentrations for various solvents. The aim of our study was to compare two methods in which urinary concentrations of pseudocumene were determined by gas chromatography using headspace technique. The standard method was based on determining concentration of organic solvents in 100 mm3 or 1 cm3 samples of urine. The incubation conditions were as follows: equilibration temperature and time: 70 degrees C, 30 min., respectively. 1 cm3 of gas phase was sampled with a gas-tight syringe and injected into a gas chromatograph. The new method using Headspace Sampler was based on determining concentrations of solvents in 10 cm3 samples of urine. The operating conditions were: equilibration time 30 min.; equlibration temperature 80 degrees C; pressurization time 0.1 min.; loop fill time 0.1 min.; loop equilibration 0.1 min.; loop equilibration time 0.05 min.; inject time 1 min.; loop temperature 150 degrees C, transfer line temperature 150 degrees C. HP 7694 Headspace Sampler minimizes sample degradation with a chemically inert pathway extending from the sample loop to the column head. The analytical parameters of both methods (linearity, precision, reproducibility, stability and sensitivity) are fully compatible with the principles of biological monitoring. Application of the headspace autosampler eliminated interference from the biological matrix and made it possible to achieve very low detection limit.

  15. Method validation and application of a liquid chromatography-tandem mass spectrometry method for drugs of abuse testing in exhaled breath.

    PubMed

    Stephanson, Niclas; Sandqvist, Sören; Lambert, Marjan Shafaati; Beck, Olof

    2015-03-15

    A mass spectrometric method for drugs of abuse testing in exhaled breath employing a sampling device collecting aerosol particles was developed and applied in routine use. Analytes covered were amphetamine, methamphetamine, 6-acetylmorphine, morphine, cocaine, benzoylecgonine, diazepam, oxazepam and tetrahydrocannabinol. The method involved eluting drugs from the collection filter with methanol, quantification using deuterated analogs as internal standards, reversed phase chromatography with gradient elution, positive electrospray ionization and monitoring of two product ions per analyte in selected reaction monitoring mode. The measuring range was 6.0-1000pg/filter. The intra- and inter-assay imprecision expressed as the coefficient of variation was less than 7%. Influence from matrix was noted for most compounds but was compensated for the use of co-eluting internal standards. The LLOQ was 6.0pg/filter with intra-assay CV <5% and accuracy within 99-102% for all analytes. No chromatographic interference was observed in 20 negative control samples. The LC-MS/MS method was successfully applied for measuring drugs in unknown samples collected for the purpose of drug testing. Among the 1096 analyzed samples analytical findings were made in breath in 39 cases (3.6%). Most frequently found substances were the following: amphetamine (25 cases) methamphetamine (10 cases), THC (8 cases), cocaine (4 cases), benzoylecgonine (2 cases) and diazepam (2 cases). In conclusion, a fully validated and robust screening method suitable for the routine measurement of drugs of abuse in exhaled breath with a simple procedure for specimen collection and sample preparation was successfully developed.

  16. Ultra-high-pressure liquid chromatography tandem mass spectrometry method for the determination of 9 organophosphate flame retardants in water samples.

    PubMed

    Lorenzo, María; Campo, Julián; Picó, Yolanda

    2016-01-01

    Few methods are available for comprehensive organophosphate flame retardants (PFRs) detection in water and wastewater. Gas chromatography has been employed previously, but this approach is less selective, not amenable for use with deuterated standards and can suffer unfavorable fragmentation. Ultra-high-pressure liquid chromatography tandem mass spectrometry (UHPLC-QqQ-MS/MS) has become the most promising platform, already applied successfully for analysis of selected PFRs in some environmental matrices like water and wastewater. However, the presence of some interferences from the dissolvent, the equipment and the used materials should be taken into account. The procedure involves: •The first determination of PFRs by UHPLC-QqQ-MS/MS using a trap column to distinguish the interferences coming from the instrument and mobile phases.•The optimization of the LC separation to distinguish all target compounds and their interferences.•This method coupled to a solid-phase extraction (SPE) improve the detection and quantification of PFRs.

  17. Evaluation of automated sample preparation, retention time locked gas chromatography-mass spectrometry and data analysis methods for the metabolomic study of Arabidopsis species.

    PubMed

    Gu, Qun; David, Frank; Lynen, Frédéric; Rumpel, Klaus; Dugardeyn, Jasper; Van Der Straeten, Dominique; Xu, Guowang; Sandra, Pat

    2011-05-27

    In this paper, automated sample preparation, retention time locked gas chromatography-mass spectrometry (GC-MS) and data analysis methods for the metabolomics study were evaluated. A miniaturized and automated derivatisation method using sequential oximation and silylation was applied to a polar extract of 4 types (2 types×2 ages) of Arabidopsis thaliana, a popular model organism often used in plant sciences and genetics. Automation of the derivatisation process offers excellent repeatability, and the time between sample preparation and analysis was short and constant, reducing artifact formation. Retention time locked (RTL) gas chromatography-mass spectrometry was used, resulting in reproducible retention times and GC-MS profiles. Two approaches were used for data analysis. XCMS followed by principal component analysis (approach 1) and AMDIS deconvolution combined with a commercially available program (Mass Profiler Professional) followed by principal component analysis (approach 2) were compared. Several features that were up- or down-regulated in the different types were detected.

  18. Rapid and accurate liquid chromatography and tandem mass spectrometry method for the simultaneous quantification of ten metabolic reactions catalyzed by hepatic cytochrome P450 enzymes.

    PubMed

    Shi, Rong; Ma, Bingliang; Wu, Jiasheng; Wang, Tianming; Ma, Yueming

    2015-10-01

    The hepatic cytochrome P450 enzymes play a central role in the biotransformation of endogenous and exogenous substances. A sensitive high-throughput liquid chromatography with tandem mass spectrometry assay was developed and validated for the simultaneous quantification of the products of ten metabolic reactions catalyzed by hepatic cytochrome P450 enzymes. After the substrates were incubated separately, the samples were pooled and analyzed by liquid chromatography with tandem mass spectrometry using an electrospray ionization source in the positive and negative ion modes. The method exhibited linearity over a broad concentration range, insensitivity to matrix effects, and high accuracy, precision, and stability. The novel method was successfully applied to study the kinetics of phenacetin-O deethylation, coumarin-7 hydroxylation, bupropion hydroxylation, taxol-6 hydroxylation, omeprazole-5 hydroxylation, dextromethorphan-O demethylation, tolbutamide-4 hydroxylation, chlorzoxazone-6 hydroxylation, testosterone-6β hydroxylation, and midazolam-1 hydroxylation in rat liver microsomes.

  19. Rapid method for the preparation of 125I-labelled human growth hormone for receptor studies, using reverse-phase high performance liquid chromatography

    SciTech Connect

    Ilondo, M.M.; Dehart, I.; De Meyts, P.

    1986-01-29

    Human growth hormone was labelled with 125 Iodine by the stoichiometric modification of the chloramine-T method to a specific activity of 50-80 microCi/microgram, and the iodinated mixture was purified by reverse-phase high performance liquid chromatography using a C18 column (SynChropak RP-P) and a linear gradient. Compared with the usual Sephadex G-100 chromatography, HPLC gave a much better separation, with a higher yield and a considerably reduced analysis time (30 min vs 5 h). The (125I)-labelled preparation had normal binding to IM-9 lymphocyte receptors. The maximum bindability of the HPLC-purified preparation approximated 90%, which is the best value so far reported for human growth hormone. It is concluded that HPLC is a fast, convenient and reproducible method for obtaining an improved (125I)-labelled human growth hormone for receptor studies.

  20. A liquid chromatography and tandem mass spectrometry method for the determination of potential biomarkers of cardiovascular disease.

    PubMed

    Magiera, Sylwia; Baranowska, Irena; Kusa, Jacek; Baranowski, Jacek

    2013-03-01

    A simple, accurate and sensitive liquid chromatography tandem mass spectrometry (LC-MS/MS) method was developed and validated for the quantitation of α-ketoglutaric acid (α-KG), L-carnitine (L-CAR) and acetyl-L-carnitine (acetyl-L-CAR) in human urine as potential biomarkers of cardiovascular disease. The separation was performed using an isocratic elution of 0.1% formic acid in water and acetonitrile (97:3, v/v) on an Acclaim 120 C8 column (150 mm × 4.6 mm, 3.0 μm). The flow rate of the mobile phase was 1.2 mL/min and the total assay run time was 3 min. Detection was performed on a triple-quadrupole mass spectrometer in selected reaction monitoring (SRM) mode via an electrospray ionization (ESI) source in positive and negative ion modes. This method covered a linearity range of 0.1-500 ng/mL for L-CAR and acetyl-L-CAR and 1-1000 ng/mL for α-KG with lower limits of quantification (LLOQ) of 0.08 ng/mL for L-CAR, 0.04 ng/mL for acetyl-L-CAR and 0.8 ng/mL for α-KG. The intra-day and inter-day precision and accuracy of the quality control samples exhibited relative standard deviations of less than 5.54% and relative error values from -5.95% to 3.11%. Analyte stability was evaluated under various sample preparation, analysis and storage conditions and varied from -9.89% to -0.47%. A two-step solid-phase extraction (SPE) procedure using silica gel and quaternary amine cartridges was used for urine sample cleanup. The average recoveries for all analyzed compounds were better than 86.64% at three concentrations. The method was successfully applied for the quantitation of α-KG, L-CAR and acetyl-L-CAR in human urine samples.

  1. Gas chromatography-mass spectrometry method for the determination of methadone and 2-ethylidene-1,5-dimethyl-3, 3-diphenylpyrrolidine (EDDP).

    PubMed

    Snozek, Christine L H; Bjergum, Matthew W; Langman, Loralie J

    2010-01-01

    Methadone is a synthetic opioid used to relieve pain, treat opioid withdrawal, and wean heroin addicts. Measurement of methadone and its major metabolite EDDP in urine is useful for assessing compliance with addiction rehabilitation and pain management programs. This method quantitatively measures methadone and its metabolite EDDP in urine. Methadone and EDDP are recovered from urine by solid phase extraction at pH 6.0. Analysis is performed by gas chromatography with mass spectrometry detection, using selective ion monitoring.

  2. Simple Detection Methods for Antinutritive Factor β-ODAP Present in Lathyrus sativus L. by High Pressure Liquid Chromatography and Thin Layer Chromatography

    PubMed Central

    Ghosh, Bidisha; Mitra, Joy; Chakraborty, Saikat; Bhattacharyya, Jagannath; Chakraborty, Anirban; Sen, Soumitra Kumar; Neerathilingam, Muniasamy

    2015-01-01

    Lathyrus sativus L. (Grass pea) is the source for cheap and nutritious food choice in drought and famine susceptible zones in greater part of North India and Africa. The non-protein amino acid β-N-oxalyl-L-α,β-diaminopropionic acid (β-ODAP) has been known for decades for its potent neurotoxic effect, causing irreversible neurodegenerative disease “neurolathyrism”, present in both seed and leaf of Lathyrus sativus L. and other species in varying proportions. It is crucial to establish a rapid as well as reliable detection methodology for β-ODAP content in various Lathyrus plants. Currently available HPLC based methods involve multi-step derivatization of the sample. To overcome this, we have developed β-ODAP analysis method by HPLC without any prior derivatization. This method is statistically significant in the range of 2 to 100μg/ml and exhibited linear response with r2 > 0.99. Limit of detection and quantitation of the later method was determined to be 5.56 μg/ml and 16.86 μg/ml, respectively. In addition to this, a TLC based method has also been developed. The limit of detection of β-ODAP is 0.6μg and for its substrate, L-1,2-diaminopropionic acid is 5μg. Both HPLC and TLC methods were validated by conducting in-vitro bioconversion test to detect the presence of biocatalyst in plant extract. This method is economical, rapid and simple. PMID:26524073

  3. A new method for rapid determination of indole-3-carbinol and its condensation products in nutraceuticals using core-shell column chromatography method.

    PubMed

    Fibigr, Jakub; Šatínský, Dalibor; Havlíková, Lucie; Solich, Petr

    2016-02-20

    Indole-3-carbinol is a natural glucosinolate known for prevention of human breast, prostate and other types of cancer and it started to be used in commercial preparations, as food supplements. However no analytical method has been proposed for quality control of nutraceuticals with this substance yet. In this paper a new high-performance liquid chromatography (HPLC) method using core-shell column for separation of indole-3-carbinol and its condensation/degradation products was developed and used for the quantitative determination of indole-3-carbinol in nutraceuticals. Separation of indole-3-carbinol, its condensation/degradation products and internal standard ethylparaben was performed on the core-shell column Kinetex 5μ XB-C18 100A (100×4.6mm), particle size 5.0μm, with mobile phase acetonitrile/water according to the gradient program at a flow rate of 1.25mLmin(-1) and at temperature 50°C. The detection wavelength was set at 270nm. Under the optimal chromatographic conditions good linearity of determination was achieved. Available commercial samples of nutraceuticals were extracted with 100% methanol using ultrasound bath. A 5-μL sample volume of the supernatant was directly injected into the HPLC system. The developed method provided rapid and accurate tool for quality control of nutraceuticals based on cruciferous vegetable extracts with indole-3-carbinol content. The presented study showed that the declared content of indole-3-carbinol significantly varied in the different nutraceuticals available on the market. Two analyzed preparations showed the presence of condensation/degradation products of indole-3-carbinol which were not officially declared by the manufacturer. Moreover, further two analyzed nutraceutical preparations showed absolutely no content of declared amount of indole-3-carbinol.

  4. Rapid quantitative analysis of individual anthocyanin content based on high-performance liquid chromatography with diode array detection with the pH differential method.

    PubMed

    Wang, Huayin

    2014-09-01

    A new quantitative technique for the simultaneous quantification of the individual anthocyanins based on the pH differential method and high-performance liquid chromatography with diode array detection is proposed in this paper. The six individual anthocyanins (cyanidin 3-glucoside, cyanidin 3-rutinoside, petunidin 3-glucoside, petunidin 3-rutinoside, and malvidin 3-rutinoside) from mulberry (Morus rubra) and Liriope platyphylla were used for demonstration and validation. The elution of anthocyanins was performed using a C18 column with stepwise gradient elution and individual anthocyanins were identified by high-performance liquid chromatography with tandem mass spectrometry. Based on the pH differential method, the high-performance liquid chromatography peak areas of maximum and reference absorption wavelengths of anthocyanin extracts were conducted to quantify individual anthocyanins. The calibration curves for these anthocyanins were linear within the range of 10-5500 mg/L. The correlation coefficients (r(2)) all exceeded 0.9972, and the limits of detection were in the range of 1-4 mg/L at a signal-to-noise ratio ≥5 for these anthocyanins. The proposed quantitative analysis was reproducible with good accuracy of all individual anthocyanins ranging from 96.3 to 104.2% and relative recoveries were in the range 98.4-103.2%. The proposed technique is performed without anthocyanin standards and is a simple, rapid, accurate, and economical method to determine individual anthocyanin contents.

  5. Application of high-speed counter-current chromatography as a new pretreatment method for analysis of polycyclic aromatic hydrocarbons in environmental water samples

    PubMed Central

    Cao, Xueli; Yang, Chunlei; Pei, Hairun; Li, Xinghong; Xu, Xiaobai; Ito, Yoichiro

    2011-01-01

    High-speed counter-current chromatography (HSCCC) was investigated as a new sample pretreatment method for the analysis of trace polycyclic aromatic hydrocarbons (PAHs) in water environmental samples. The experiment was performed with a nonaqueous binary two-phase solvent system composed of heptane-acetonitrile. The HSCCC column was first entirely filled with the upper stationary phase of the and a large volume of water sample was pumped into the column while the CCC column was rotated at 1600 rpm. Finally, the trace amount of PAHs extracted and enriched in the stationary phase were eluted out by the lower mobile phase. and analyzed by gas chromatography-flame ionization detector (GC-FID) or gas chromatography-mass spectrometry (GC-MS) after concentration. The enrichment and cleanup of PAHs can be fulfilled online by this methodwithhigh recoveries (84.1%–103.2%) and good reproducibility (RSDs 4.9–12.2%) for 16 EPA PAHs under the optimized HSCCC pretreatment conditions. This method has been successfully applied to determine PAHs in lake waterwhere 8 PAHs were detected in the concentration of 40.9–89.9 ng/L. The present method is extremely suitable in the preparation of large volume of environmental water sample for the determination of a trace amount of organic pollutants including PAHs as studied in this paper. PMID:22282420

  6. An Eco-Friendly Direct Injection HPLC Method for Methyldopa Determination in Serum by Mixed-Mode Chromatography Using a Single Protein-Coated Column.

    PubMed

    Emara, Samy; Masujima, Tsutomu; Zarad, Walaa; Kamal, Maha; Fouad, Marwa; El-Bagary, Ramzia

    2015-09-01

    A simple, rapid and environment-friendly direct injection HPLC method for the determination of methyldopa (MTD) in human serum has been developed and validated. The method was based on cleanup and separation of MTD from serum by mixed-mode liquid chromatography using a single protein-coated TSK gel ODS-80 TM analytical column (50 × 4.0 mm i.d., 5 µm). The protein-coated column exhibited excellent resolution, selectivity and functioned in two chromatographic modes: size-exclusion chromatography [i.e., solid-phase extraction (SPE) for serum proteins] and reversed-phase chromatography for the final separation of MTD. SPE and HPLC separation were carried out simultaneously with a green mobile phase consisting of acetate buffer (0.1 M, pH 2.4) at a flow rate of 1 mL/min and at room temperature (23 ± 1°C). The eluent was monitored at emission and excitation wavelengths of 320 and 270 nm, respectively. A calibration curve was linear over the range of 0.1-30 µg/mL with a detection limit of 0.027 µg/mL. This online SPE method was successfully applied to real samples obtained from patients receiving MTD therapy.

  7. On-line stop-flow two-dimensional liquid chromatography-mass spectrometry method for the separation and identification of triterpenoid saponins from ginseng extract.

    PubMed

    Wang, Shuangyuan; Qiao, Lizhen; Shi, Xianzhe; Hu, Chunxiu; Kong, Hongwei; Xu, Guowang

    2015-01-01

    A method based on stop-flow two-dimensional liquid chromatography coupled with electrospray ionization mass spectrometry (2D LC-ESI MS) was established and applied to analyze triterpenoid saponins from the main root of ginseng. Due to the special structure of triterpenoid saponins (they contain polar sugar side chains and nonpolar aglycones), hydrophilic interaction chromatography (HILIC) and reversed-phase liquid chromatography (RPLC) were used for the two dimensions, respectively. A trap column was used to connect the two dimensions. The dilution effect, which is one of the main shortcomings of traditional comprehensive 2D LC methods, was largely avoided. The peak capacity of this system was 747 and the orthogonality was 56.6 %. Compared with one-dimensional HILIC or RP LC MS analysis, 257 and 185 % more mass spectral peaks (ions with intensities that were higher than 1,000) were obtained from the ginseng main root extracts, and 94 triterpenoid saponins were identified based on MS(n) information and summarized aglycone structures. Given its good linearity and repeatability, the established method was successfully applied to classify ginsengs of different ages (i.e., years of growth), and 19 triterpenoid saponins were found through statistical analysis to vary in concentration depending on the age of the ginseng.

  8. Exploring Liquid Sequential Injection Chromatography to Teach Fundamentals of Separation Methods: A Very Fast Analytical Chemistry Experiment

    ERIC Educational Resources Information Center

    Penteado, Jose C.; Masini, Jorge Cesar

    2011-01-01

    Influence of the solvent strength determined by the addition of a mobile-phase organic modifier and pH on chromatographic separation of sorbic acid and vanillin has been investigated by the relatively new technique, liquid sequential injection chromatography (SIC). This technique uses reversed-phase monolithic stationary phase to execute fast…

  9. Method for the quantification of diamorphine and its metabolites in pediatric plasma samples by liquid chromatography-tandem mass spectrometry.

    PubMed

    Al-Asmari, Ahmed; Anderson, Robert A; Kidd, Susan; Thomson, Alison H

    2010-05-01

    In recent years, intranasal diamorphine (DIM) has been recommended as an alternative to intravenous administration for the treatment of acute-to-severe pain in children. This provides a rapid and less painful route of administration without decreasing the effectiveness of the analgesic properties. A sensitive technique for the detection and quantitation of DIM and its metabolites is essential because of the low concentrations of DIM and metabolites in children's plasma, which are a result of the low dose of DIM given and the limited sample volume obtained from children (0.25 mL or less). DIM can be easily hydrolyzed to 6-monoacetylmorphine (6-MAM) during sample preparation and extraction, so this must be considered when developing a solid-phase extraction (SPE) method to prevent the hydrolysis of DIM. This work was aimed at validating and developing a liquid chromatography-tandem mass spectrometry (LC-MS-MS) method with electrospray ionization for the identification and quantification of DIM and its metabolites, namely 6-MAM, MOR, M3G, M6G, and NMOR in plasma samples obtained from children who are under treatment for acute-to-severe pain. Following the addition of deuterated internal standards, analytes were extracted by SPE with Bond Elut C(18) cartridges followed by LC-MS-MS analysis. Intraday and interday precision for all analytes were determined at five concentration (1, 5, 25, 50, and 200 ng/mL), and these were found to be 2.5-13.4% and 1.8-15%, respectively. Recoveries of analytes of interest were between 81 and 109%. Calibration curves were linear for all analytes over the concentration range 0.1-50 ng/mL, and correlation coefficients were better than 0.999. Limits of detection and quantitation were 0.08-0.37 ng/mL and 0.28-1.22 ng/mL, respectively. DIM and metabolites were detected in all case samples with the exception of NMOR, which tested negative in all cases. The pharmacokinetics of DIM and its metabolites following INDIM and IVDIM administration in

  10. Convex hull: a new method to determine the separation space used and to optimize operating conditions for comprehensive two-dimensional gas chromatography.

    PubMed

    Semard, Gaëlle; Peulon-Agasse, Valerie; Bruchet, Auguste; Bouillon, Jean-Philippe; Cardinaël, Pascal

    2010-08-13

    It is important to develop methods of optimizing the selection of column sets and operating conditions for comprehensive two-dimensional gas chromatography. A new method for the calculation of the percentage of separation space used was developed using Delaunay's triangulation algorithms (convex hull). This approach was compared with an existing method and showed better precision and accuracy. It was successfully applied to the selection of the most convenient column set and the geometrical parameters of second column for the analysis of 49 target compounds in wastewater.

  11. Gas Chromatography.

    ERIC Educational Resources Information Center

    Cram, Stuart P.; And Others

    1980-01-01

    Selects fundamental developments in theory, methodology, and instrumentation in gas chromatography (GC). A special section reviews GC in the People's Republic of China. Over 1,000 references are cited. (CS)

  12. Analytical Method for Sugar Profile in Pet Food and Animal Feeds by High-Performance Anion-Exchange Chromatography with Pulsed Amperometric Detection.

    PubMed

    Ellingson, David J; Anderson, Phillip; Berg, Daniel P

    2016-01-01

    There is a need for a standardized, accurate, rugged, and consistent method to measure for sugars in pet foods and animal feeds. Many traditional standard sugar methods exist for other matrixes, but when applied in collaborative studies there was poor agreement and sources of error identified with those standard methods. The advancement in technology over the years has given us the ability to improve on these standard methods of analysis. A method is described here that addresses these common issues and was subjected to a single-laboratory validation to assess performance on a wide variety of pet foods and animal feeds. Of key importance to the method performance is the sample preparation before extraction, type of extraction solvent, postextraction cleanup, and, finally, optimized chromatography using high-performance anion exchange chromatography with pulsed amperometric detection. The results obtained from the validation demonstrate how typical issues seen with these matrixes can influence performance of sugar analysis. The results also demonstrate that this method is fit-for-purpose and can meet the challenges of sugar analysis in pet food and animal feeds to lay the foundation for a standardized method of analysis.

  13. Quantification of F2-isoprostanes as a reliable index of oxidative stress in vivo using gas chromatography-mass spectrometry (GC-MS) method.

    PubMed

    Liu, Wei; Morrow, Jason D; Yin, Huiyong

    2009-10-15

    Free radical-induced lipid peroxidation has been implicated in a number of human diseases including atherosclerosis, cancer, and neurodegenerative diseases. F(2)-Isoprostanes (IsoPs) are isomers of prostaglandin PGF(2alpha) that are generated in vivo from the free radical-initiated peroxidation of arachidonic acid independent of cyclooxygenase enzymes. Since the discovery of the IsoPs in the early 1990s, a large body of evidence has been accumulated to indicate that quantification of these F(2)-IsoPs represents the most reliable biomarker to assess oxidative stress in vivo. A variety of analytical approaches have been developed for the quantification of these novel compounds; these methods include mass spectrometry (MS) detection coupled to gas chromatography (GC) or liquid chromatography (LC) separation, and detection using immunological approaches. This article summarizes our current methodology to quantify F(2)-IsoPs in biological fluids and tissues using GC-MS. This method includes solid-phase extraction (SPE), thin-layer chromatography (TLC) purification, chemical derivatization, and MS detection using negative ion chemical ionization (NICI) coupled with GC. The protocol described herein has been optimized and validated to provide the best sensitivity and selectivity for quantification of F(2)-IsoPs from a variety of biological sources.

  14. Development and evaluation of a hydrophilic interaction liquid chromatography-MS/MS method to quantify 19 nucleobases and nucleosides in rat plasma.

    PubMed

    Du, Yan; Li, Yin-Jie; Hu, Xun-Xiu; Deng, Xu; Qian, Zeng-Ting; Li, Zheng; Guo, Meng-Zhe; Tang, Dao-Quan

    2017-04-01

    As essential endogenous compounds, nucleobases and nucleosides fulfill various functions in living organisms. This study presents the development and validation of a new hydrophilic interaction liquid chromatography tandem mass spectrometry method for simultaneous quantification of 19 nucleobases and nucleosides in rat plasma. For the sample preparation, 15 kinds of protein precipitants were evaluated according to the chromatographic profile and ion response of analytes. The optimization of chromatographic separation was respectively performed using reversed-phase liquid chromatography and hydrophilic interaction liquid chromatography mode; each separation mode included two test columns with different stationary phases. The chromatographic profile and parameters such as half-width (W1/2 ), capacity factor (K') and tailing factor (ft ) were used to evaluate the separation efficiencies. Furthermore, the adopted composition of two mobile phase systems and the concentrations of the additives in the optimum buffer system were also investigated. The developed method was fully validated and successfully applied quantitatively to determine 19 nucleobases and nucleosides in plasma from normal and diabetic nephropathy (DN) rats. Significant differences between normal and DN rats were found in plasma levels of cytosine, xanthine, thymidine, adenosine, guanosine, inosine and 8-hydroxy-2'-deoxyguanosine. This information may provide a useful reference for the discovery of potential biomarkers of DN.

  15. Cell membrane chromatography coupled with UHPLC-ESI-MS/MS method to screen target components from Peucedanum praeruptorum Dunn acting on α1A adrenergic receptor.

    PubMed

    Han, Shengli; Li, Chunlei; Huang, Jing; Wei, Fen; Zhang, Yu; Wang, Sicen

    2016-02-01

    Peucedanum praeruptorum Dunn (BaiHuaQianHu in Chinese) is a traditional Chinese medicine that has a long history of use in China. In this study, HEK 293 α1A adrenergic cell membrane chromatography was coupled with UHPLC-ESI-MS/MS and successfully used to identify active components from Peucedanum praeruptorum Dunn. Paeruptorin A, paeruptorin B, and paeruptorin C were identified with α1A adrenergic receptor activity. Pharmacological assays showed that tamsulosin hydrochloride, paeruptorin A, paeruptorin B, and paeruptorin C in concentrations of 1×10(-8) to 1×10(-4)mol/mL could relax prostate strips pre-contracted with adrenalin in a concentration dependent manner. Therefore, the HEK293 α1A cell membrane chromatography coupled UHPLC-ESI-MS/MS system may be a potentially useful drug discovery method for screening for medicinal herbal components with α1A adrenergic receptor inhibitory activity.

  16. A simple, sensitive, and rapid method for the determination of cotinine in urine by high-performance liquid chromatography with UV detection.

    PubMed

    Rabbaa-Khabbaz, Lydia; Abi Daoud, Rita; Karam-Sarkis, Dolla

    2006-10-01

    Cotinine levels in biological fluids are a reliable indicator of the presence of nicotine. In this paper, a simple and sensitive high-performance liquid chromatography (HPLC) procedure for the determination of cotinine in urine following liquid-liquid extraction with dichloromethane in an alkaline medium is described. Calibration curves show linearity over the 50 to 3000 ng/mL range with low intra- and interday variability as well as good selectivity and specificity. No solid-phase extraction is performed because the liquid dichloromethane extraction step yields excellent results. This method is a good alternative for routine analysis of urinary cotinine in laboratories where gas chromatography or HPLC-mass spectrometry is not available.

  17. Quantification of two polyacetylenes in Radix Ginseng and roots of related Panax species using a gas chromatography-mass spectrometric method.

    PubMed

    Liu, Ji-Hua; Lee, Che-Sum; Leung, Kit-Ming; Yan, Zhong-Kai; Shen, Bai-Hua; Zhao, Zhong-Zhen; Jiang, Zhi-Hong

    2007-10-31

    A sensitive method for quantitating the pharmacologically active polyacetylenes panaxynol and panaxydol in Radix Ginseng was developed using a capillary gas chromatography-mass spectrometric (GC-MS) method. The detection mode of selected ion monitoring (SIM) allowed sensitive and selective quantitation of the two compounds in ginseng. Method validation showed that the GC-MS method has much lower detection and quantitation limits than the high-performance liquid chromatography (HPLC)-UV method. This indicates that GC-MS is particularly useful for the analysis of polyacetylene compounds, which have relatively low abundances compared with ginsenosides in ginseng. Based on the quantitative results of different types of ginseng herbs, it was found that the panaxydol and panaxynol contents were higher in forest ginseng than in cultivated ginseng. This method was further applied to the quantitative analyses of panaxynol and panaxydol in Radix Notoginseng and American ginseng. The ratio of panaxydol to panaxynol can be utilized as a marker for differentiating ginseng, notoginseng, and American ginseng. This study introduces the first GC-MS method for the quantitative analysis of polyacetylenes in herbs of the Panax genus.

  18. Validation of a fast liquid chromatography-UV method for the analysis of drugs used in combined cardiovascular therapy in human plasma.

    PubMed

    Iriarte, Gorka; Gonzalez, Oskar; Ferreirós, Nerea; Maguregui, Miren Itxaso; Alonso, Rosa Maria; Jiménez, Rosa Maria

    2009-10-01

    Ultra-performance liquid chromatography (UPLC) was investigated as a faster alternative to high-performance liquid chromatography (HPLC) for the simultaneous analysis of drugs usually prescribed in cardiovascular therapy. Upon a previously developed and validated solid phase extraction (SPE)-HPLC-photodiode array (PDA)-fluorescence (FLR) method, separation of chlorthalidone (CLTD; diuretic), valsartan and its metabolite (VAL and VAL-M1 respectively; angiotensin II receptor antagonist drugs) and fluvastatin (FLUV; statin) was performed in human plasma using an RP C18 column (50mmx2.1mm, 1.7microm, Waters Acquity UPLC (BEH)) and a tunable UV-vis (TUV) detector. After method transfer, different system variables were modulated to study the evolution of responses of the analytes and the endogenous interferences. The improved method was fully validated and the results were compared with its precursor HPLC method relating to analysis time, efficiency and sensitivity. The studied compounds were separated in less than 8min and the method showed good linearity (20-3000microg/L for chlorthalidone, 110-1100microg/L for valsartan-M1, 67-1900microg/L for valsartan and 48-1100microg/L for fluvastatin), precision and accuracy. The proposed method was found to be reproducible (RSD<10%), accurate (RE<15%), robust and suitable for quantitative analysis of the studied drugs in plasma obtained from patients under combined cardiovascular treatment.

  19. Simultaneous quantification of five proteins and seven additives in dairy products with a heart-cutting two-dimensional liquid chromatography method.

    PubMed

    Hou, Xiaofang; Sun, Meng; He, Xiaoshuang; Chen, Liang; Zhang, Ping; He, Langchong

    2015-09-01

    A heart-cutting two-dimensional high-performance liquid chromatography method was developed to simultaneously quantify five major proteins and seven food additives (maltol, ethyl maltol, vanillin, ethyl vanillin, benzoic acid, sorbic acid, and saccharin sodium) in milk and milk powders. In this two-dimensional system, a Venusil XBP-C4 column was selected in the first dimension for protein separation, and a Hypersil ODS-2 C18 column was employed in the second dimension for additive separation; a two-position, six-port switching valve was used to transfer the targets (additives) from the first dimension to the second dimension. Method validation consisted of selectivity, response function, linearity, precision, sensitivity, and recovery. In addition, a conventional one-dimensional high-performance liquid chromatography method was also tested for comparison. The two-dimensional method resulted in significantly improved recovery of the food additives compared to the conventional method (90.6-105.4% and 65.5-86.5%, respectively). Furthermore, this novel method has a simple one-step sample preparation procedure, which shortens the analysis time, resulting in more efficient analysis and less solvent usage.

  20. Chemometric approach to open validation protocols: Prediction of validation parameters in multi-residue ultra-high performance liquid chromatography-tandem mass spectrometry methods.

    PubMed

    Alladio, Eugenio; Pirro, Valentina; Salomone, Alberto; Vincenti, Marco; Leardi, Riccardo

    2015-06-09

    The recent technological advancements of liquid chromatography-tandem mass spectrometry allow the simultaneous determination of tens, or even hundreds, of target analytes. In such cases, the traditional approach to quantitative method validation presents three major drawbacks: (i) it is extremely laborious, repetitive and rigid; (ii) it does not allow to introduce new target analytes without starting the validation from its very beginning and (iii) it is performed on spiked blank matrices, whose very nature is significantly modified by the addition of a large number of spiking substances, especially at high concentration. In the present study, several predictive chemometric models were developed from closed sets of analytes in order to estimate validation parameters on molecules of the same class, but not included in the original training set. Retention time, matrix effect, recovery, detection and quantification limits were predicted with partial least squares regression method. In particular, iterative stepwise elimination, iterative predictors weighting and genetic algorithms approaches were utilized and compared to achieve effective variables selection. These procedures were applied to data reported in our previously validated ultra-high performance liquid chromatography-tandem mass spectrometry multi-residue method for the determination of pharmaceutical and illicit drugs in oral fluid samples in accordance with national and international guidelines. Then, the partial least squares model was successfully tested on naloxone and lormetazepam, in order to introduce these new compounds in the oral fluid validated method, which adopts reverse-phase chromatography. Retention time, matrix effect, recovery, limit of detection and limit of quantification parameters for naloxone and lormetazepam were predicted by the model and then positively compared with their corresponding experimental values. The whole study represents a proof-of-concept of chemometrics potential to

  1. A review on development of analytical methods to determine monitorable drugs in serum and urine by micellar liquid chromatography using direct injection.

    PubMed

    Esteve-Romero, Josep; Albiol-Chiva, Jaume; Peris-Vicente, Juan

    2016-07-05

    Therapeutic drug monitoring is a common practice in clinical studies. It requires the quantification of drugs in biological fluids. Micellar liquid chromatography (MLC), a well-established branch of Reverse Phase-High Performance Liquid Chromatography (RP-HPLC), has been proven by many researchers as a useful tool for the analysis of these matrices. This review presents several analytical methods, taken from the literature, devoted to the determination of several monitorable drugs in serum and urine by micellar liquid chromatography. The studied groups are: anticonvulsants, antiarrhythmics, tricyclic antidepressants, selective serotonin reuptake inhibitors, analgesics and bronchodilators. We detail the optimization strategy of the sample preparation and the main chromatographic conditions, such as the type of column, mobile phase composition (surfactant, organic solvent and pH), and detection. The finally selected experimental parameters, the validation, and some applications have also been described. In addition, their performances and advantages have been discussed. The main ones were the possibility of direct injection, and the efficient chromatographic elution, in spite of the complexity of the biological fluids. For each substance, the measured concentrations were accurate and precise at their respective therapeutic range. It was found that the MLC-procedures are fast, simple, inexpensive, ecofriendly, safe, selective, enough sensitive and reliable. Therefore, they represent an excellent alternative for the determination of drugs in serum and urine for monitoring purposes.

  2. Development and validation of a multiclass method for the quantification of veterinary drug residues in honey and royal jelly by liquid chromatography-tandem mass spectrometry.

    PubMed

    Jin, Yue; Zhang, Jinzhen; Zhao, Wen; Zhang, Wenwen; Wang, Lin; Zhou, Jinhui; Li, Yi

    2017-04-15

    The aim of this study was to develop an analytical method for the analysis of a wide range of veterinary drugs in honey and royal jelly. A modified sample preparation procedure based on the quick, easy, cheap, effective, rugged and safe (QuEChERS) method was developed, followed by liquid chromatography tandem mass spectrometry determination. Use of the single sample preparation method for analysis of 42 veterinary drugs becomes more valuable because honey and royal jelly belong to completely different complex matrices. Another main advantage of the proposed method is its ability to identify and quantify 42 veterinary drugs with higher sensitivity than reference methods of China. This work has shown that the reported method was demonstrated to be convenient and reliable for the quick monitoring of veterinary drugs in honey and royal jelly samples.

  3. EPA Air Method, Toxic Organics - 15 (TO-15): Determination of Volatile Organic Compounds (VOCs) in Air Collected in Specially-Prepared Canisters and Analyzed by Gas Chromatography/Mass Spectrometry (GC/MS)

    EPA Pesticide Factsheets

    Method T)-15 describes procedures for for preparation and analysis of air samples containing volatile organic compounds collected in specially-prepared canisters, using gas chromatography-mass spectrometry.

  4. EPA Method 551.1: Determination of Chlorination Disinfection Byproducts, Chlorinated Solvents, and Halogenated Pesticides/Herbicides in Drinking Water by Liquid-Liquid Extraction and Gas Chromatography With Electron-Capture Detection

    EPA Pesticide Factsheets

    SAM lists this method as a gas chromatography with electron capture detection applicable to the determination of halogenated analytes in finished drinking water, drinking water during intermediate stages of treatment and raw source water.

  5. Methods of analysis by the U.S. Geological Survey National Water Quality Laboratory; determination of organochlorine pesticides and polychlorinated biphenyls in bottom sediment by dual capillary-column gas chromatography with electron-capture detection

    USGS Publications Warehouse

    Foreman, William T.; Connor, Brooke F.; Furlong, Edward T.; Vaught, Deborah G.; Merten, Leslie M.

    1995-01-01

    A method for the determination of 30 individual organochlorine pesticides, total toxaphene, and total polychlorinated biphenyls (PCBs) in bottom sediment is described. The method isolates the pesticides and PCBs by solvent extraction with dichlorobenzene, removes inorganic sulfur, large naturally occurring molecules, and other unwanted interferences by gel permeation chromatography, and further cleans up and class fractionates the extract using adsorption chromatography. The com- pounds then are instrumentally determined using dual capillary-column gas chromatography with electron-capture detection. Reporting limits range from 1 to 5 micrograms per kilogram for 30 individual pesticides, 50 micrograms per kilogram for total PCBs, and 200 micrograms per kilogram for total toxaphene. The method also is designed to allow the simultaneous isolation of 79 other semivolatile organic compounds from the sediment, which are separately quantified using gas chromatography with mass spectrometric detection. The method was developed in support of the U.S. Geological Survey's National Water-Quality Assessment program.

  6. Micellar Electrokinetic Chromatography

    NASA Astrophysics Data System (ADS)

    Bald, Edward; Kubalczyk, Paweł

    Since the introduction of micellar electrokinetic chromatography by Terabe, several authors have paid attention to the fundamental characteristics of this separation method. In this chapter the theoretical and practical aspects of resolution optimization, as well as the effect of different separation parameters on the migration behavior are discussed. These among others include fundamentals of separation, retention factor and resolution equation, efficiency, selectivity, and various surfactants and additives. Initial conditions for method development and instrumental approaches such as mass spectrometry detection are also mentioned covering the proposals for overcoming the difficulties arising from the coupling micellar electrokinetic chromatography with mass spectrometry detection.

  7. [A novel method for the identification of illegal cooking oil (1): detection of three capsaicinoids with liquid chromatography-mass spectrometry].

    PubMed

    Ang, Longxing; Jin, Jing; Wang, Shuqiu; Wang, Xingfu; Tian, Yuzeng; Chen, Jiping

    2012-11-01

    Illegal cooking oil (ICO, also named swill-cooked dirty oil) has recently become a serious food safety problem in China. Now, the identification method of ICO is also a hot research area. Owning to the special eating habits of Chinese people, cayenne is widely used in catering business. Capsaicinoids are main spicy compounds in cayenne. So, they are potential evaluation indices for the identification of ICO. In this study, a solid phase extraction-liquid chromatography-mass spectrometry (SPE-LC-MS) method has been developed to detect the trace residues of three capsaicinoids (capsaicin, dihydrocapsaicin and nonylic acid vanillylamide) in cooking oil. The oil sample was first extracted with 20 g/L sodium hydroxide, the C18 SPE cartridge was then used to clean-up the sample and enrich the analytes before the liquid chromatography-mass spectrometry (LC-MS) detection. With this method, sixty seven blind samples provided by China National Center for Food Safety Risk Assessment were analyzed. The results showed that the capsaicinoids are good evaluation indices for the identification of ICO. In all the 48 ICO samples, 36 samples were successfully recognized. All the 19 normal oil samples were accurately identified. This method has been chosen and authorized as one of the four standard instrumental identification methods for ICO by the National Ministry of Health of China.

  8. Application of Solid Phase Microextraction Coupled with Gas Chromatography/Mass Spectrometry as a Rapid Method for Field Sampling and Analysis of Chemical Warfare Agents and Toxic Industrial Chemicals

    DTIC Science & Technology

    2003-01-01

    PHASE MICROEXTRACTION COUPLED WITH GAS CHROMATOGRAPHY/MASS SPECTROMETRY AS A RAPID METHOD FOR FIELD SAMPLING AND ANALYSIS OF CHEMICAL WARFARE AGENTS...4. TITLE AND SUBTITLE APPLICATION OF SOLID PHASE MICROEXTRACTION COUPLED WITH GAS CHROMATOGRAPHY/MASS SPECTROMETRY AS A RAPID METHOD FOR FIELD...unclassified Standard Form 298 (Rev. 8-98) Prescribed by ANSI Std Z39-18 iii ABSTRACT APPLICATION OF SOLID PHASE MICROEXTRACTION COUPLED WITH

  9. Development of a qualitative liquid chromatography/tandem mass spectrometric method for the detection of narcotics in urine relevant to doping analysis.

    PubMed

    Deventer, Koen; Pozo, Oscar J; Van Eenoo, Peter; Delbeke, Frans T

    2007-01-01

    A new screening procedure for 18 narcotics in urine for anti-doping purposes has been developed using liquid chromatography/triple quadrupole mass spectrometry (LC/MS). Electrospray ionization (ESI) was used as interface. Infusion experiments were performed for all substances to investigate their mass spectrometric behaviour in terms of selecting product specific ions. These product ions were then used to develop a tandem mass spectrometric method using selected reaction monitoring (SRM). For the LC/MS analysis, chromatography was performed on an octadecylsilane column. The total run time of the chromatographic method was 5.5 min. For the sample preparation prior to LC/MS analysis, the urine samples were liquid-liquid extracted at pH 9.5 after overnight enzymatic hydrolysis. Two extraction solvents were evaluated: dichloromethane/methanol 9/1 (v/v), which is currently used for the extraction of narcotics, and diethyl ether, used for the extraction of steroids. With diethyl ether the detection limits for all compounds ranged between 0.5 and 20 ng/mL and with the mixture containing dichloromethane the detection limits ranged between 0.5 and 10 ng/mL. Taking into account the minimum required performance limits of the World Anti-Doping Agency of 200 ng/mL for narcotics, diethyl ether can also be considered as extraction solvent for narcotics. Finally, the described method was applied to the analysis of urine samples previously found to contain narcotics by our routine gas chromatography/mass spectrometry (GC/MS) method.

  10. Improved solvent extraction procedure and high-performance liquid chromatography-evaporative light-scattering detector method for analysis of polar lipids from dairy materials.

    PubMed

    Le, Thien Trung; Miocinovic, Jelena; Nguyen, Tuyet Mai; Rombaut, Roeland; van Camp, John; Dewettinck, Koen

    2011-10-12

    A normal-phase high-performance liquid chromatography-evaporative light-scattering detector method employing dichloromethane, methanol, and acetic acid/triethylamine buffer as the mobile phase was developed for analysis of polar lipids (PLs). This method was applicable for analysis of PLs from both dairy materials and soy lecithin. All of the PLs of interest such as glycolipids, phospholipids, and sphingomyelin were well separated with a total run time of 22.5 min and without necessitating the removal of neutral lipids beforehand. Peak retention times were stable, and the method was reproducible. In this study, a modified method of using solvents for extraction of PLs from dairy matrices was also investigated. The modified method offered higher extraction efficiency, consumed less time, and in some cases saved solvent use.

  11. A high-performance liquid chromatography-electronic circular dichroism online method for assessing the absolute enantiomeric excess and conversion ratio of asymmetric reactions.

    PubMed

    Zhang, Xiang; Wang, Mingchao; Li, Li; Yin, Dali

    2017-03-02

    Asymmetric reactions often need to be evaluated during the synthesis of chiral compounds. However, traditional evaluation methods require the isolation of the individual enantiomer, which is tedious and time-consuming. Thus, it is desirable to develop simple, practical online detection methods. We developed a method based on high-performance liquid chromatography-electronic circular dichroism (HPLC-ECD) that simultaneously analyzes the material conversion ratio and absolute optical purity of each enantiomer. In particular, only a reverse-phase C18 column instead of a chiral column is required in our method because the ECD measurement provides a g-factor that describes the ratio of each enantiomer in the mixtures. We used our method to analyze the asymmetric hydrosilylation of β-enamino esters, and we discussed the advantage, feasibility, and effectiveness of this new methodology.

  12. Validation and uncertainty analysis of a multi-residue method for pesticides in grapes using ethyl acetate extraction and liquid chromatography-tandem mass spectrometry.

    PubMed

    Banerjee, Kaushik; Oulkar, Dasharath P; Dasgupta, Soma; Patil, Shubhangi B; Patil, Sangram H; Savant, Rahul; Adsule, Pandurang G

    2007-11-30

    A method was validated for the multi-residue analysis of 82 pesticides in grapes at chromatography-tandem mass spectrometry. Reduction in sample size and proportion of ethyl acetate for extraction did not affect accuracy or precision of analysis when compared to the reported methods and was also statistically similar to the QuEChERS technique. The method was rugged (HorRat < 0.5) with <20% measurement uncertainties. Limit of quantification was <10 ng/g with recoveries 70-120% for most pesticides. The method offers cheaper and safer alternative to typical multi-residue analysis methods for grape.

  13. A high-performance liquid chromatography-electronic circular dichroism online method for assessing the absolute enantiomeric excess and conversion ratio of asymmetric reactions

    PubMed Central

    Zhang, Xiang; Wang, Mingchao; Li, Li; Yin, Dali

    2017-01-01

    Asymmetric reactions often need to be evaluated during the synthesis of chiral compounds. However, traditional evaluation methods require the isolation of the individual enantiomer, which is tedious and time-consuming. Thus, it is desirable to develop simple, practical online detection methods. We developed a method based on high-performance liquid chromatography-electronic circular dichroism (HPLC-ECD) that simultaneously analyzes the material conversion ratio and absolute optical purity of each enantiomer. In particular, only a reverse-phase C18 column instead of a chiral column is required in our method because the ECD measurement provides a g-factor that describes the ratio of each enantiomer in the mixtures. We used our method to analyze the asymmetric hydrosilylation of β-enamino esters, and we discussed the advantage, feasibility, and effectiveness of this new methodology. PMID:28252028

  14. A high-performance liquid chromatography-electronic circular dichroism online method for assessing the absolute enantiomeric excess and conversion ratio of asymmetric reactions

    NASA Astrophysics Data System (ADS)

    Zhang, Xiang; Wang, Mingchao; Li, Li; Yin, Dali

    2017-03-01

    Asymmetric reactions often need to be evaluated during the synthesis of chiral compounds. However, traditional evaluation methods require the isolation of the individual enantiomer, which is tedious and time-consuming. Thus, it is desirable to develop simple, practical online detection methods. We developed a method based on high-performance liquid chromatography-electronic circular dichroism (HPLC-ECD) that simultaneously analyzes the material conversion ratio and absolute optical purity of each enantiomer. In particular, only a reverse-phase C18 column instead of a chiral column is required in our method because the ECD measurement provides a g-factor that describes the ratio of each enantiomer in the mixtures. We used our method to analyze the asymmetric hydrosilylation of β-enamino esters, and we discussed the advantage, feasibility, and effectiveness of this new methodology.

  15. Development, validation, and application of a method for selected avermectin determination in rural waters using high performance liquid chromatography and fluorescence detection.

    PubMed

    Lemos, Maria Augusta Travassos; Matos, Camila Alves; de Resende, Michele Fabri; Prado, Rachel Bardy; Donagemma, Raquel Andrade; Netto, Annibal Duarte Pereira

    2016-11-01

    Avermectins (AVM) are macrocyclic lactones used in livestock and agriculture. A quantitative method of high performance liquid chromatography with fluorescence detection for the determination of eprinomectin, abamectin, doramectin and ivermectin in rural water samples was developed and validated. The method was employed to study samples collected in the Pito Aceso River microbasin, located in the Bom Jardim municipality, Rio de Janeiro State, Brazil. Samples were extracted by solid phase extraction using a polymeric stationary phase, the eluted fraction was re-concentrated under a gentle N2 flow and derivatized to allow AVM determination using liquid chromatography with fluorescence detection. The excitation and emission wavelengths of the derivatives were 365 and 470nm, respectively, and a total chromatographic run of 12min was achieved. Very low limits of quantification (22-58ngL(-1)) were found after re-concentration using N2. Recovery values varied from 85.7% to 119.2% with standard deviations between 1.2% and 10.2%. The validated method was applied in the determination of AVM in 15 water samples collected in the Pito Aceso River microbasin, but most of them were free of AVM or showed only trace levels of these compounds, except for a sample that contained doramectin (9.11µgL(-1)). The method is suitable for routine analysis with satisfactory recovery, sensitivity, and selectivity.

  16. High-resolution gas chromatography/mass spectrometry method for characterization and quantitative analysis of ginkgolic acids in Ginkgo biloba plants, extracts, and dietary supplements.

    PubMed

    Wang, Mei; Zhao, Jianping; Avula, Bharathi; Wang, Yan-Hong; Avonto, Cristina; Chittiboyina, Amar G; Wylie, Philip L; Parcher, Jon F; Khan, Ikhlas A

    2014-12-17

    A high-resolution gas chromatography/mass spectrometry (GC/MS) with selected ion monitor method focusing on the characterization and quantitative analysis of ginkgolic acids (GAs) in Ginkgo biloba L. plant materials, extracts, and commercial products was developed and validated. The method involved sample extraction with (1:1) methanol and 10% formic acid, liquid-liquid extraction with n-hexane, and derivatization with trimethylsulfonium hydroxide (TMSH). Separation of two saturated (C13:0 and C15:0) and six unsaturated ginkgolic acid methyl esters with different positional double bonds (C15:1 Δ8 and Δ10, C17:1 Δ8, Δ10, and Δ12, and C17:2) was achieved on a very polar (88% cyanopropyl) aryl-polysiloxane HP-88 capillary GC column. The double bond positions in the GAs were determined by ozonolysis. The developed GC/MS method was validated according to ICH guidelines, and the quantitation results were verified by comparison with a standard high-performance liquid chromatography method. Nineteen G. biloba authenticated and commercial plant samples and 21 dietary supplements purported to contain G. biloba leaf extracts were analyzed. Finally, the presence of the marker compounds, terpene trilactones and flavonol glycosides for Ginkgo biloba in the dietary supplements was determined by UHPLC/MS and used to confirm the presence of G. biloba leaf extracts in all of the botanical dietary supplements.

  17. Liquid chromatography-tandem mass spectrometry method for determination of panel of neurotransmitters in cerebrospinal fluid from the rat model for tauopathy.

    PubMed

    Kovac, Andrej; Somikova, Zuzana; Zilka, Norbert; Novak, Michal

    2014-02-01

    Alzheimer's disease (AD) is still being recognized today as an unmet medical need. Currently, there is no cure and early preclinical diagnostic assay available for AD. Therefore much attention is now being directed at the development of novel methods for quantitative determination of AD biomarkers in the cerebrospinal fluid (CSF). Here, we describe the liquid chromatography-tandem mass spectrometry method for determination of 5-hydroxytryptamine (SER), 5-hydroxyindoleacetic acid (5-HIAA), homovanilic acid (HVA), noradrenaline (NADR), adrenaline (ADR), dopamine (DA), glutamic acid (Glu), γ-aminobutyric acid (GABA), 3,4-dihydroxyphenylacetic acid (DOPAC) and histamine (HIS) in cerebrospinal fluid (CSF) from the rat model for human tauopathy. The benzoyl chloride was used as pre-column derivatization reagents. Neurotransmitters and metabolites were analysed on ultra performance liquid chromatography (UPLC) on C18 column in combination with tandem mass spectrometry. The method is simple, highly sensitive and showed excellent linearity with regression coefficients higher than 0.99. The accuracy was in a range of 93-113% for all analytes. The inter-day precision (n=5 days), expressed as %RSD, was in a range 2-10% for all analytes. Using this method we detected significant changes of CSF levels of two important neurotransmitters/metabolites, ADR and 5-HIAA, which correlates with progression of neurodegeneration in our animal model.

  18. High-throughput sample preparation and simultaneous column regeneration liquid chromatography-tandem mass spectrometry method for determination of nitrogen mustard metabolites in human urine.

    PubMed

    Reddy, Muntha K; Mills, Grier; Nixon, Christopher; Wyatt, Shane A; Croley, Timothy R

    2011-08-15

    Nitrogen mustards (NMs) are known to have DNA alkylation and strong vesicant properties. Their availability to terrorist organizations makes them a potential choice for chemical attacks on civilian populations. After an exposure, it is difficult to measure NMs directly because of their rapid metabolism in the human body. Therefore to determine an individual's level of exposure to NMs, it is necessary to analyze for NM metabolites being excreted by the body. The metabolites of NMs are generated by a hydrolysis reaction, and are easily detectable by liquid chromatography tandem mass spectrometry (LC-MS/MS). This work is focused on the development of a high-throughput assay for the quantitation of N-ethyldiethanolamine (EDEA) and N-methyldiethanolamine (MDEA) metabolites of bis (2-chloroethyl) ethylethanamine (HN1) and bis (2-chloroethyl) methylethanamine (HN2), respectively. The method uses automated 96-well plate sample preparation of human urine samples and a 2-position 10-port switching valve to allow for simultaneous regeneration of the liquid chromatography (LC) columns. Using this method, over 18 h was saved through the reduction of sample preparation and analysis time when compared to a conventional method for 96 samples. The validated method provided excellent accuracy for both EDEA (100.9%) and MDEA (100.6%) with precision better than 5.27% for each analyte.

  19. Review: Recent application of chiral liquid chromatography-tandem mass spectrometric methods for enantiomeric pharmaceutical and biomedical determinations.

    PubMed

    Nie, Yanfang; Liu, Xiumei; Yang, Xinmei; Zhao, Zhongxi

    2013-09-01

    The technology of analyzing the pharmacological and toxicological properties of chiral drugs has become more commonly utilized since the knowledge of the biological actions of enantiomers has been gradually acquired during the last few decades. This work reviews the applications of chiral liquid chromatography coupled to tandem mass spectrometry (LC-MS) for the analysis of chiral pharmaceuticals since 2005. The enantioselective determinations in pharmaceutical and biomedical fields with LC-MS are classified based on three mobile phase modes, including normal phase, reversed phase and polar organic solvent, in terms of their compatibility with various ionization sources and specific applications.

  20. Gas chromatography-mass spectrometry screening methods for select UV filters, synthetic musks, alkylphenols, an antimicrobial agent, and an insect repellent in fish.

    PubMed

    Mottaleb, Mohammad A; Usenko, Sascha; O'Donnell, John G; Ramirez, Alejandro J; Brooks, Bryan W; Chambliss, C Kevin

    2009-01-30

    Two screening methods have been developed for simultaneous determination of ten extensively used personal care products (PCPs) and two alkylphenol surfactants in fish. The methods consisted of extraction, clean-up, derivatization and analysis by gas chromatography-mass spectrometry with selected ion monitoring (GC-SIM-MS) or gas chromatography-tandem mass spectrometry (GC-MS/MS) techniques. Among solvents tested to assess recovery of target compounds from 1-g tissue homogenates, acetone was selected as optimal for extracting compounds with dissimilar physicochemical properties from fish tissue. Initial experiments confirmed that GC-SIM-MS could be applied for analysis of lean fillet tissue (<1% lipid) without gel-permeation chromatography (GPC), and this approach was applied to assess the presence of target analytes in fish fillets collected from a regional effluent-dominated stream in Texas, USA. Benzophenone, galaxolide, tonalide, and triclosan were detected in 11 of 11 environmental samples at concentrations ranging from; 37 to 90, 234 to 970, 26 to 97, and 17 to 31 ng/g, respectively. However, performance of this analytical approach declined appreciably with increasing lipid content of analyzed tissues. Successful analysis of samples with increased lipid content was enabled by adding GPC to the sample preparation protocol and monitoring analytes with tandem mass spectrometry. Both analytical approaches were validated using fortified fillet tissue collected from locations expected to be minimally impacted by anthropogenic influences. Average analyte recoveries ranged from 87% to 114% with RSDs <11% and from 54% to 107% with RSDs <20% for fish tissue containing <1% and 4.9% lipid, respectively. Statistically derived method detection limits (MDLs) for GC-SIM-MS and GC-MS/MS methodologies ranged from 2.4 to 16 ng/g, and 5.1 to 397 ng/g, respectively.

  1. Analytical approach to determining human biogenic amines and their metabolites using eVol microextraction in packed syringe coupled to liquid chromatography mass spectrometry method with hydrophilic interaction chromatography column.

    PubMed

    Konieczna, Lucyna; Roszkowska, Anna; Synakiewicz, Anna; Stachowicz-Stencel, Teresa; Adamkiewicz-Drożyńska, Elżbieta; Bączek, Tomasz

    2016-04-01

    Analysis of biogenic amines (BAs) in different human samples provides insight into the mechanisms of various biological processes, including pathological conditions, and thus may be very important in diagnosing and monitoring several neurological disorders and cancerous tumors. In this work, we developed a simple and fast procedure using a digitally controlled microextraction in packed syringe (MEPS) coupled to liquid chromatography mass spectrometry (LC-MS) method for simultaneous determination of biogenic amines, their precursors and metabolites in human plasma and urine samples. The separation of 12 low molecular weight and hydrophilic molecules with a wide range of polarities was achieved with hydrophilic interaction chromatography (HILIC) column without derivatization step in 12 min. MEPS was implemented using the APS sorbent in semi-automated analytical syringe (eVol(®)) and small volume of urine and plasma samples, 5 0µL and 100 μL, respectively. We evaluated important parameters influencing MEPS efficiency, including stationary phase selection, sample pH and volume, number of extraction cycles, and washing and elution volumes. In optimized MEPS conditions, the analytes were eluted by 3 × 50 μL of methanol with 0.1% formic acid. The chromatographic separation of analytes was performed on XBridge Amide™ BEH analytical column (3.0mm × 100 mm, 3.5 µm) using gradient elution with mobile phase consisting of phase A: 10mM ammonium formate buffer in water pH 3.0 and phase B: 10mM ammonium formate buffer in acetonitrile pH 3.0. The LC-HILIC-MS method was validated and, in optimum conditions, presented good linearity in concentration range within 10-2000 ng/mL for all the analytes with a determination coefficient (r(2)) higher than 0.999 for plasma and urine samples. Method recovery ranged within 87.6-104.3% for plasma samples and 84.2-98.6% for urine samples. The developed method utilizing polar APS sorbent along with polar HILIC column was applied for

  2. An Improved RP-HPLC (Reversed-Phase, High-Performance Liquid Chromatography) Method for Determining Nitroaromatics and Nitramines in Water

    DTIC Science & Technology

    1988-11-01

    high - performance liquid chromatography on an LC-18 column. The method employs dilution of an aqueous sample 1:1 with methanol, filtration through a 0.5 micro meter Millex-Sr filter, separation on the LC-18 column using a 1:1 water-methanol eluent, and determination by UV-254 nm. A careful comparison was made with an earlier standard protocol, which uses separation on an LC-8 column with a 50:38:12 water-methanol-acetonitrile eluent. overall, the new procedure provides better separation for a wider range of analytes and equivalent recovery for all

  3. Development and comparison of two multi-residue methods for the analysis of select pesticides in honey bees, pollen, and wax by gas chromatography-quadrupole mass spectrometry.

    PubMed

    Li, Yuanbo; Kelley, Rebecca A; Anderson, Troy D; Lydy, Michael J

    2015-08-01

    One of the hypotheses that may help explain the loss of honey bee colonies worldwide is the increasing potential for exposure of honey bees to complex mixtures of pesticides. To better understand this phenomenon, two multi-residue methods based on different extraction and cleanup procedures have been developed, and compared for the determination of 11 relevant pesticides in honey bees, pollen, and wax by gas chromatography-quadrupole mass spectrometry. Sample preparatory methods included solvent extraction followed by gel permeation chromatography (GPC) cleanup and cleanup using a dispersive solid-phase extraction with zirconium-based sorbents (Z-Sep). Matrix effects, method detection limits, recoveries, and reproducibility were evaluated and compared. Method detection limits (MDL) of the pesticides for the GPC method in honey bees, pollen, and wax ranged from 0.65 to 5.92 ng/g dw, 0.56 to 6.61 ng/g dw, and 0.40 to 8.30 ng/g dw, respectively, while MDLs for the Z-Sep method were from 0.33 to 4.47 ng/g dw, 0.42 to 5.37 ng/g dw, and 0.51 to 5.34 ng/g dw, respectively. The mean recoveries in all matrices and at three spiking concentrations ranged from 64.4% to 149.5% and 71.9% to 126.2% for the GPC and Z-Sep methods, with relative standard deviation between 1.5-25.3% and 1.3-15.9%, respectively. The results showed that the Z-Sep method was more suitable for the determination of the target pesticides, especially chlorothalonil, in bee hive samples. The Z-Sep method was then validated using a series of field-collected bee hive samples taken from honey bee colonies in Virginia.

  4. Development of reversed-phase high performance liquid chromatography methods for quantification of two isomeric flavones and the application of the methods to pharmacokinetic studies in rats.

    PubMed

    Whitted, Crystal L; Palau, Victoria E; Torrenegra, Ruben D; Harirforoosh, Sam

    2015-09-15

    Isomers 5,7-dihydroxy-3,6,8-trimethoxy-2-phenyl-4H-chromen-4-one (5,7-dihydroxy-3,6,8 trimethoxy flavone) (flavone A) and 3,5-dihydroxy-6,7,8-trimethoxy-2-phenyl-4H-chromen-4-one (3,5-dihydroxy-6,7,8-trimethoxy flavone) (flavone B) have recently demonstrated differential antineoplastic activities against pancreatic cancer in vitro. These studies also indicated that these compounds target highly tumorigenic cells while sparing normal cells. The in vivo antitumor activities of these flavones have not been determined, and detection protocols for these compounds are needed to conduct pre-clinical assays following intravenous dosing. Here, we report methods developed using acetonitrile to extract two flavone isomers and corresponding internal standards, celecoxib and diclofenac, from rat plasma. Separation was achieved using a Shimadzu liquid chromatography system with a C18 column and mobile phase acetonitrile/water (60:40 and 70:30 for flavones A and B, respectively) containing 0.2% acetic acid and 0.05% triethylamine at a flow rate of 0.4mL/min and detection at 245nm. Calibration curves ranging from 250 to 2500ng/mL and 2500 to 100,000ng/mL for both flavones were linear (r(2)≥0.99) with the lower limits of quantification being 250ng/mL. Recovery of concentrations 250, 1000, 2500, 5000, and 100,000ng/mL ranged from 87 to 116% and 84 to 103% (n=3) for flavone A and B, respectively. Stability of both flavones after a freezing/thawing cycle yielded a mean peak ratio ≥0.92 when compared to freshly extracted samples. Intravenous administration of a 20mg/kg dose in rats yielded half-lives of 83.68±56.61 and 107.45±53.31min with clearance values of 12.99±13.78 and 80.79±35.06mL/min/kg for flavones A and B, respectively.

  5. Development and validation of high-throughput liquid chromatography-tandem mass spectrometric method for simultaneous quantification of loratadine and desloratadine in human plasma.

    PubMed

    Srinubabu, G; Patel, Rajaram S; Shedbalkar, Vinay P; Rao, Allam Appa; Rao, M Narasimha; Bandaru, Veera Venkata Ratnam

    2007-12-15

    As a continuation of effort to improve our high flow on-line bioanalytical approach for high-throughput quantification of drugs and metabolites in plasma by high-throughput liquid chromatography tandem mass spectrometry (HTLC-MS/MS), we have developed a simple, sensitive and reliable method for simultaneous quantification of loratadine and desloratadine in human plasma. We have performed on-line coupling of extraction with Cyclone P 50 mm x 0.5 mm 50 microm HTLC column and chromatographic separation is performed with Zorbax XDB C18 50 mm x 2.1 mm 5 microm, followed by quantification with mass detector. The method is validated and showed good performances in terms of linearity, sensitivity, precision, accuracy and stability. A marked improvement in sample throughput efficiency is realized with this method and the proposed method will be useful for pharmacokinetic and/or bioequivalence studies.

  6. High-performance liquid chromatography method for determination of hydrogen peroxide in aqueous solution and application to simulated Martian soil and related materials.

    PubMed

    Steinberg, Spencer M

    2013-05-01

    A new method for determination of hydrogen peroxide (H2O2) is described. H2O2 reacts with iodide in the presence of ammonium molybdate and vanillic acid resulting in the formation of iodovanillic acid which is quantified by reversed-phase high-performance liquid chromatography (HPLC) and detected by UV absorption at 280 nm. The method provides a detection limit of ~0.1 μM and is easy to implement. Iodovanillic acid is stable, allowing multiple measurements of the same sample, and the separation power of the HPLC yields high selectivity against potential interferences. This method will be useful for many environmental applications. The method has been applied to quantify the photochemical production of H2O2 by aqueous suspensions of various minerals and soils.

  7. Development and validation of sensitive method for determination of serum cotinine in smokers and nonsmokers by liquid chromatography/atmospheric pressure ionization tandem mass spectrometry.

    PubMed

    Bernert, J T; Turner, W E; Pirkle, J L; Sosnoff, C S; Akins, J R; Waldrep, M K; Ann, Q; Covey, T R; Whitfield, W E; Gunter, E W; Miller, B B; Patterson, D G; Needham, L L; Hannon, W H; Sampson, E J

    1997-12-01

    We describe a sensitive and specific method for measuring cotinine in serum by HPLC coupled to an atmospheric pressure chemical ionization tandem mass spectrometer. This method can analyze 100 samples/day on a routine basis, and its limit of detection of 50 ng/L makes it applicable to the analysis of samples from nonsmokers potentially exposed to environmental tobacco smoke. Analytical accuracy has been demonstrated from the analysis of NIST cotinine standards and from comparative analyses by both the current method and gas chromatography/high-resolution mass spectrometry. Precision has been examined through the repetitive analysis of a series of bench and blind QC materials. This method has been applied to the analysis of cotinine in serum samples collected as part of the Third National Health and Nutrition Examination Survey (NHANES III).

  8. Development and application of a liquid chromatography-mass spectrometry method to evaluate the glyphosate and aminomethylphosphonic acid dissipation in maize plants after foliar treatment.

    PubMed

    Bernal, José; Martin, María T; Soto, María E; Nozal, María J; Marotti, Ilaria; Dinelli, Giovanni; Bernal, José L

    2012-04-25

    A simple and fast method has been developed and validated to measure glyphosate (GLYP) and aminomethylphosphonic acid (AMPA), which were previously derivatized with 9-fluorenylmethylchloroformate (FMOC-Cl), in maize plants using liquid chromatography (LC) coupled to fluorescence (FLD) and electrospray ionization mass spectrometry (ESI-MS) detection. The method has shown to be consistent, reliable, precise, and efficient. Moreover, the limits of detection (LOD) and quantification (LOQ) reached with the proposed method for GLYP and AMPA are lower than the established maximum residue levels (MRLs). The validated method was applied to quantify GLYP and AMPA in genetically modified (GM) maize foliar treated with the herbicide. It has been found that the GLYP dissipation was mainly due to the progressive dilution effect after herbicide treatment. Finally, it was also observed that the GLYP residue dissipation trend in maize shoot (leaves and stem) tissue determined by LC-ESI-MS matched that determined by liquid scintillation.

  9. Development of a Stationary Phase Optimised Selectivity Liquid Chromatography based screening method for adulterations of food supplements for the treatment of pain.

    PubMed

    Deconinck, E; Kamugisha, A; Van Campenhout, P; Courselle, P; De Beer, J O

    2015-06-01

    Illegally adulterated dietary supplements are an increasing problem worldwide. One of the important groups of often adulterated products are the dietary supplements, sold for the treatment of pain. These often contain analgesics, a heterogeneous group of molecules, containing both hydrophilic and hydrophobic compounds. The development of a screening method for these components, especially when mass spectrometric detection is not available, necessitates chromatographic separation, difficult to achieve with traditional chromatographic columns. In this paper Stationary Phase Optimised Selectivity Liquid Chromatography was used for the development of a screening method for nine analgesics, codeine and caffeine, often present in this type of dietary supplements. The method shows a good separation of all the compounds, allowing the screening to be performed with diode array detection and is fully compatible with mass spectrometry. The method was validated for its selectivity following the guidelines as described for the screening of pesticide residues and residues of veterinary medicines in food.

  10. Method development and validation for the quantification of dasatinib, erlotinib, gefitinib, imatinib, lapatinib, nilotinib, sorafenib and sunitinib in human plasma by liquid chromatography coupled with tandem mass spectrometry.

    PubMed

    Lankheet, N A G; Hillebrand, M J X; Rosing, H; Schellens, J H M; Beijnen, J H; Huitema, A D R

    2013-04-01

    To support pharmacokinetic-guided dosing in individual patients, a fast and accurate method for simultaneous determination of anticancer tyrosine kinase inhibitors (TKIs) dasatinib, erlotinib, gefitinib, imatinib, lapatinib, nilotinib, sorafenib and sunitinib in human plasma was developed using high-performance liquid chromatography and detection with tandem mass spectrometry (HPLC-MS/MS). Stable isotopically labeled compounds of the eight different TKIs were used as internal standards. Plasma proteins were precipitated and an aliquot of supernatant was directly injected onto a reversed phase chromatography system consisting of a Gemini C18 column (50 × 2.0 mm i.d., 5.0 µm particle size) and then compounds were eluted with a gradient. The outlet of the column was connected to a triple quadrupole mass spectrometer with electrospray interface. Ions were detected in the positive multiple reaction monitoring mode. This method was validated over a linear range from 20.0 to 10,000 ng/mL for erlotinib, gefitinib, imatinib, lapatinib, nilotinib and sorafenib, and from 5.00 to 2500 ng/mL for dasatinib and sunitinib. Results from the validation study demonstrated good intra- and inter-assay accuracy (<13.1%) and precision (10.0%) for all analytes. This method was successfully applied for routine therapeutic drug monitoring purposes in patients treated with the investigated TKIs.

  11. Development and validation of a turbulent flow chromatography and tandem mass spectrometry method for the quantitation of methotrexate and its metabolites 7-hydroxy methotrexate and DAMPA in serum.

    PubMed

    Schofield, Ryan C; Ramanathan, Lakshmi V; Murata, Kazunori; Grace, Marie; Fleisher, Martin; Pessin, Melissa S; Carlow, Dean C

    2015-10-01

    A rapid and simple turbulent flow liquid chromatography (TFC-LC) method implementing positive heated electrospray ionization (HESI) for the accurate and precise determination of methotrexate (MTX), 7-hydroxy methotrexate (7-OH MTX), and 4-amino-4-deoxy-N(10)-methylpteroic acid (DAMPA) concentrations in serum was developed. MTX was isolated from serum samples (100μL) after protein precipitation with methanol containing formic acid and internal standard (MTX-D3) followed by centrifugation. The supernatant was injected into the turbulent flow liquid chromatography which is followed by electrospray positive ionization tandem mass spectrometry (TFC-LC-MS/MS) and quantified using a six-point calibration curve. For MTX and DAMPA the assays were linear from 10 to 1000nmol/L and for 7-OH MTX from 20 to 2000nmol/L. Dilutions of 10, 100 and 1000-fold were validated giving a clinically reportable range of 10nmol/L to 5×10(5)nmol/L. Within-day and between-day precisions at concentrations spanning the analytical measurement ranges were less than 10% for all three analytes. MTX, DAMPA and 7-OH MTX were sufficiently stable under all relevant analytical conditions. No significant matrix effect was observed during the method validation. The TFC-LC-MS/MS MTX method was also compared with three other clinically validated MTX assays: a dihydrofolate reductase (DHFR) inhibition assay, an immunoassay based on fluorescence polarization and a previously developed LC-MS/MS assay.

  12. Environmentally friendly method for the determination of acrylamide and trimethylolpropane in paper packaging materials by liquid chromatography with tandem mass spectrometry.

    PubMed

    Yang, Fei; Li, Zhonghao; Bian, Zhaoyang; Tang, Gangling; Fan, Ziyan; Wang, Ying; Liu, ShanShan; Zhang, Hongfei

    2014-12-01

    A simple, rapid, and environmentally friendly method was developed for the determination of acrylamide and trimethylolpropane in paper packaging materials. No organic solvent was used and the matrix effect was investigated. The extract was directly analyzed by liquid chromatography with tandem mass chromatography for quantification and confirmation. The chromatographic separations were performed on a ZORBAX HILIC Plus (2.1 mm × 150 mm, 3μm; Agilent, USA) column with only one mobile phase (100% water). Calibration curves for acrylamide and trimethylopropane were achieved with concentrations ranging from 0.4 to 20 mg/kg and the corresponding r(2) values were 0.998 and 0.999, respectively. The recoveries were >85% with relative standard deviations <10%. The validated method was applied to the analysis of 50 real samples, and positive results were obtained for 30 samples. The result indicated that trimethylolpropane is associated with inks and printing activity and acrylamide is widely used as a papermaking additive in many paper packages. The concentrations of acrylamide and trimethylolpropane ranged from 0.41 to 7.5 and 0.50 to 8.8 mg/kg, respectively. The results of this study revealed that this method could be used accurately and precisely.

  13. Multi-residue analysis method for analysis of pharmaceuticals using liquid chromatography-time of flight/mass spectrometry (LC-TOF/MS) in water sample

    NASA Astrophysics Data System (ADS)

    Al-Qaim, Fouad Fadhil; Abdullah, Md Pauzi; Othman, Mohamed Rozali

    2013-11-01

    In this work, a developed method using solid - phase extraction (SPE) followed by liquid chromatography - time of flight mass spectrometry (LC-ESI-TOF/MS) was developed and validated for quantification and confirmation of eleven pharmaceuticals with different therapeutic classes in water samples, Malaysia. These compounds are caffeine (CAF), prazosin (PRZ), enalapril (ENL), carbamazepine (CBZ), nifedipine (NFD), levonorgestrel (LNG), simvastatin (SMV), hydrochlorothiazide (HYD), gliclazide (GLIC), diclofenac-Na (DIC-Na) and mefenamic acid (MEF). LC was performed on a Dionex Ultimate 3000/LC 09115047 (USA) system. Chromatography was performed on a Thermo Scientific C18 (250 mm × 2.1 mm, i.d.: 5μm) column. Several parameters were optimised such as; mobile phase, gradient elution, collision energy and solvent elution for extraction of compounds from water. The recoveries obtained ranged from 30-148 % in river water. Five pharmaceutical compounds were detected in the surface water samples: caffeine, prazosin, enalpril, diclofenac-Na and mefenamic acid. The developed method is precise and accepted recoveries were got. In addition, this method is suitable to identify and quantify trace concentrations of pharmaceuticals in surface water.

  14. Application of a fast and cost-effective in situ derivatization method prior to gas chromatography with mass spectrometry to monitor endocrine disruptors in water matrices.

    PubMed

    Melo, Armindo; Ferreira, Isabel M P L V O; Mansilha, Catarina

    2015-06-01

    This work deals with the optimization of a rapid, cost-effective, and eco-friendly gas chromatography with mass spectrometry method for the simultaneous determination of four endocrine disruptor compounds in water matrices: estrone, 17β-estradiol, 17α-ethinylestradiol, and bisphenol A, that are currently considered to be of main concern in the field of water policy and that could became candidates for future regulations. The method involves simultaneous derivatization and extraction of compounds by dispersive liquid-liquid microextraction followed by gas chromatography with mass spectrometry analysis. Derivatization and extraction parameters were optimized with the aid of experimental design approach. An excellent linear response was achieved for all analytes (r(2) ≥ 0.999). Limits of detection and quantification are 0.003-0.005 and 0.0094-0.0164 μg/L, respectively. Intraday precision ranged between 1.1 and 12.6%, whereas interday precision ranged between 0.5 and 14.7%. For accuracy, bias values varied between -15.0 and 13.7%. Recoveries at three concentration levels ranged from 86.4 to 118.2%. The proposed method can be applied to the routine analysis of groundwater, river, sea, tap, and mineral water samples with excellent sensitivity, precision, and accuracy.

  15. Liquid chromatography-particle beam electron ionization mass spectrometry method for analysis of botanical extracts: evaluation of ephedrine alkaloids in standard reference materials.

    PubMed

    Castro, Joaudimir; Krishna, M V Balarama; Marcus, R Kenneth

    2010-01-01

    The preliminary validation of a high-performance liquid chromatography particle beam mass spectrometry method (HPLC-PB/MS) with electron impact ionization source for analysis of botanical extracts is presented. The LC-PB/MS system was evaluated for the analysis of ephedrine alkaloids using ephedra-containing National Institute of Standards and Technology dietary supplement standard reference materials (SRMs) 3241 Ephedra Sinica Stapf Native Extract and 3242 Ephedra Sinica Stapf Commercial Extract. The ephedrine alkaloids were separated by reversed-phase chromatography using a phenyl column at room temperature. A linear gradient method with a mobile phase composition varying from 5:95 [MeOH:0.1% trifluoroacetic acid (TFA) in water] to 20:80 (MeOH:0.1% TFA in water) at a flow rate of 1.0 ml/min, with an analysis time of less than 20 min, was used. The source block temperature was evaluated to determine the optimal operating conditions by monitoring the intensities and fragmentation patterns of the ephedrine alkaloids. Ephedrine and N-methylephedrine were taken as a representative of the test alkaloids. The LODs on the sub-nanogram level were achieved, with ephedrine, pseudoephedrine, and methylephedrine in the SRMs quantified by a standard addition method with recoveries of > or = 86% and RSDs of < or = 14% (n = 3).

  16. Critical points in the evaluation of analytical methods based on liquid chromatography separation for the determination of doramectin in different environmental samples.

    PubMed

    Wagil, Marta; Białk-Bielińska, Anna; Maszkowska, Joanna; Stepnowski, Piotr; Kumirska, Jolanta

    2015-01-01

    In recent years, the number of analytical methods of target compound residues (such as pharmaceuticals) has grown rapidly. Most of them are based on high performance liquid chromatography (HPLC). From the economic point of view, it is usual to apply the conditions of available HPLC methods or to design extraction and chromatographic separation conditions using HPLC and transfer them subsequently to a more sensitive technique like liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS). However, if such a transfer is planned, it is important to assess the quality of the newly-designed LC-MS/MS method. The determination of parameters like matrix effects (ME), extraction efficiency (EE) and absolute recovery (AR) is mandatory. These parameters can visualise the weakest step in the analytical method and enable methods based on different techniques to be compared. The aim of this work was to show how quality assessment should be carried out in order to transfer an optimised method from one technique to another. The representative compound used in our investigation was doramectin (DOR), an anthelmintic drug used in veterinary medicine. The quality of the suggested methods for determining this drug in three environmental matrices (water, sediment and fish tissue) using HPLC-UV and LC-MS/MS was evaluated on the basis of known values of absolute recovery (HPLC-UV) and matrix effect, extraction efficiency and absolute recovery (all LC-MS/MS). Finally, the suggested methods for determining DOR in water, sediment and fish tissue based on LC-MS/MS measurements were validated and applied to the analysis of real environmental samples.

  17. A novel two-dimensional liquid-chromatography method for online prediction of the toxicity of transformation products of benzophenones after water chlorination.

    PubMed

    Li, Jian; Ma, Li-Yun; Xu, Li; Shi, Zhi-Guo

    2015-08-01

    Benzophenone-type UV filters (BPs) are ubiquitous in the environment. Transformation products (TPs) of BPs with suspected toxicity are likely to be produced during disinfection of water by chlorination. To quickly predict the toxicity of TPs, in this study, a novel two-dimensional liquid-chromatography (2D-LC) method was established in which the objective of the first dimension was to separate the multiple components of the BPs sample after chlorination, using a reversed-phase liquid-chromatography mode. A biochromatographic system, i.e. bio-partitioning micellar chromatography with the polyoxyethylene (23) lauryl ether aqueous solution as the mobile phase, served as the second dimension to predict the toxicity of the fraction from the first dimension on the basis of the quantitative retention-activity relationships (QRARs) model. Six BPs, namely 2,4-dihydroxybenzophenone, oxybenzone, 4-hydroxybenzophenone, 2-hydroxy-4-methoxybenzophenone-5-sulfonic acid, 2,2'-dihydroxy-4,4'-dimethoxybenzophenone and 2,2'-dihydroxy-4-methoxybenzophenone, were the target analytes subjected to chlorination. The products of these BPs after chlorination were directly injected to the 2D-LC system for analysis. The results indicated that most TPs may be less toxic than their parent chemicals, but some may be more toxic, and that intestinal toxicity of TPs may be more obvious than blood toxicity. The proposed method is time-saving, high-throughput, and reliable, and has great potential for predicting toxicity or bioactivity of unknown and/or known components in a complex sample. Graphical Abstract The scheme for the 2D-LC online prediction of toxicity of the transformation products of benzophenone-type UV filters after chlorination.

  18. [Determination of 51 carbamate pesticide residues in vegetables by liquid chromatography-tandem mass spectrometry based on optimization of QuEChERS sample preparation method].

    PubMed

    Wang, Lianzhu; Zhou, Yu; Huang, Xiaoyan; Wang, Ruilong; Lin, Zixu; Chen, Yong; Wang, Dengfei; Lin, Dejuan; Xu, Dunming

    2013-12-01

    The raw extracts of six vegetables (tomato, green bean, shallot, broccoli, ginger and carrot) were analyzed using gas chromatography-mass spectrometry (GC-MS) in full scan mode combined with NIST library search to confirm main matrix compounds. The effects of cleanup and adsorption mechanisms of primary secondary amine (PSA) , octadecylsilane (C18) and PSA + C18 on co-extractives were studied by the weight of evaporation residue for extracts before and after cleanup. The suitability of the two versions of QuEChERS method for sample preparation was evaluated for the extraction of 51 carbamate pesticides in the six vegetables. One of the QuEChERS methods was the original un-buffered method published in 2003, and the other was AOAC Official Method 2007.01 using acetate buffer. As a result, the best effects were obtained from using the combination of C18 and PSA for extract cleanup in vegetables. The acetate-buffered version was suitable for the determination of all pesticides except dioxacarb. Un-buffered QuEChERS method gave satisfactory results for determining dioxacarb. Based on these results, the suitable QuEChERS sample preparation method and liquid chromatography-positive electrospray ionization-tandem mass spectrometry under the optimized conditions were applied to determine the 51 carbamate pesticide residues in six vegetables. The analytes were quantified by matrix-matched standard solution. The recoveries at three levels of 10, 20 and 100 microg/kg spiked in six vegetables ranged from 58.4% to 126% with the relative standard deviations of 3.3%-26%. The limits of quantification (LOQ, S/N > or = 10) were 0.2-10 microg/kg except that the LOQs of cartap and thiofanox were 50 microg/kg. The method is highly efficient, sensitive and suitable for monitoring the 51 carbamate pesticide residues in vegetables.

  19. High Performance Liquid Chromatography-Diode Array Detector Method for the Simultaneous Determination of Five Compounds in the Pulp and Seed of Sea Buckthorn

    PubMed Central

    Zhao, Lu; Wen, E; Upur, Halmuart; Tian, Shuge

    2017-01-01

    Context: Sea buckthorn (Hippophae rhamnoides L.) as a traditional Chinese medicinal plant has various uses in Xinjiang. Objective: A reversed-phase rapid-resolution liquid-chromatography method with diode array detector was developed for simultaneous determination of protocatechuic acid, rutin, quercetin, kaempferol, and isorhamnetin in the pulp and seed of sea buckthorn, a widely used traditional Chinese medicine for promoting metabolism and treating scurvy and other diseases. Settings and design: Compounds were separated on an Agilent ZORBAX SB-C18 column (4.6 mm × 250 mm, 5 μm; USA) with gradient elution using methanol and 0.4% phosphoric acid (v/v) at 1.0 mL/min. Detection wavelength was set at 280 nm. Materials and Methods: The fruits of wild sea buckthorn were collected from Wushi County in Aksu, Xinjiang Province. Statistical performances: The RSD of precision test of the five compounds were in the range of 0.60-2.22%, and the average recoveries ranged from 97.36% to 101.19%. Good linearity between specific chromatographic peak and component qualities were observed in the investigated ranges for all the analytes (R2 > 0.9997). Results: The proposed method was successfully applied to determine the levels of five active components in sea buckthorn samples from Aksu in Xinjiang. Conclusions: The proposed method is simple, fast, sensitive, accurate, and suitable for quantitative assessment of the pulp and seed of sea buckthorn. SUMMARY Quantitative analysis method of protocatechuic acid, rutin, quercetin, kaempferol, and isorhamnetin in the extract of sea buckthorn pulp and seed is developed by high-performance liquid chromatography (HPLC) diode array detection.This method is simple and accurate; has strong specificity, good precision, and high recovery rate; and provides a reliable basis for further development of the substances in the pulp and seed of sea buckthorn.The method is widely used for content determination of active ingredients or physiologically

  20. A liquid chromatography/mass spectrometry-based generic detection method for biochemical assay and hit discovery of histone methyltransferases.

    PubMed

    Li, Shu; Gu, X Justin; Hao, Qin; Fan, Hong; Li, Ling; Zhou, Shaolian; Zhao, Kehao; Chan, Ho Man; Wang, Y Karen

    2013-12-15

    Epigenetic modifications of the genome, such as DNA methylation and posttranslational modifications of histone proteins, contribute to gene regulation. Growing evidence suggests that histone methyltransferases are associated with the development of various human diseases, including cancer, and are promising drug targets. High-quality generic assays will facilitate drug discovery efforts in this area. In this article, we present a liquid chromatography/mass spectrometry (LC/MS)-based S-adenosyl homocysteine (SAH) detection assay for histone methyltransferases (HMTs) and its applications in HMT drug discovery, including analyzing the activity of newly produced enzymes, developing and optimizing assays, performing focused compound library screens and orthogonal assays for hit confirmations, selectivity profiling against a panel of HMTs, and studying mode of action of select hits. This LC/MS-based generic assay has become a critical platform for our methyltransferase drug discovery efforts.

  1. Easy Extraction Method To Evaluate δ13C Vanillin by Liquid Chromatography-Isotopic Ratio Mass Spectrometry in Chocolate Bars and Chocolate Snack Foods.

    PubMed

    Bononi, Monica; Quaglia, Giancarlo; Tateo, Fernando

    2015-05-20

    An easy extraction method that permits the use of a liquid chromatography-isotopic ratio mass spectrometry (LC-IRMS) system to evaluate δ(13)C of vanillin in chocolate products and industrial flavorings is presented. The method applies the determination of stable isotopes of carbon to discriminate between natural vanillin from vanilla beans and vanillin from other sources (mixtures from beans, synthesis, or biotechnology). A series of 13 chocolate bars and chocolate snack foods available on the Italian market and 8 vanilla flavorings derived from industrial quality control processes were analyzed. Only 30% of products considered in this work that declared "vanilla" on the label showed data that permitted the declaration "vanilla" according to European Union (EU) Regulation 1334/2008. All samples not citing "vanilla" or "natural flavoring" on the label gave the correct declaration. The extraction method is presented with data useful for statistical evaluation.

  2. Sensitive and rapid reversed-phase liquid chromatography-fluorescence method for determining bisphenol A diglycidyl ether in aqueous-based food simulants.

    PubMed

    Paseiro Losada, P; López Mahía, P; Vázquez Odériz, L; Simal Lozano, J; Simal Gándara, J

    1991-01-01

    A method has been developed for determination of bisphenol A diglycidyl ether (BADGE) in 3 aqueous-based food simulants: water, 15% (v/v) ethanol, and 3% (w/v) acetic acid. BADGE is extracted with C18 cartridges and the extract is concentrated under a stream of nitrogen. BADGE is quantitated by reversed-phase liquid chromatography with fluorescence detection. Relative precision at 200 micrograms/L was 3.4%, the detection limit of the method was 0.1 micrograms/L, and recoveries of spiking concentrations from 1 to 8 micrograms/L were nearly 100%. Relative standard deviations for the method ranged from 3.5 to 5.9%, depending on the identity of the spiked aqueous-based food simulant.

  3. Using second-order calibration method based on trilinear decomposition algorithms coupled with high performance liquid chromatography with diode array detector for determination of quinolones in honey samples.

    PubMed

    Yu, Yong-Jie; Wu, Hai-Long; Shao, Sheng-Zhi; Kang, Chao; Zhao, Juan; Wang, Yu; Zhu, Shao-Hua; Yu, Ru-Qin

    2011-09-15

    A novel strategy that combines the second-order calibration method based on the trilinear decomposition algorithms with high performance liquid chromatography with diode array detector (HPLC-DAD) was developed to mathematically separate the overlapped peaks and to quantify quinolones in honey samples. The HPLC-DAD data were obtained within a short time in isocratic mode. The developed method could be applied to determine 12 quinolones at the same time even in the presence of uncalibrated interfering components in complex background. To access the performance of the proposed strategy for the determination of quinolones in honey samples, the figures of merit were employed. The limits of quantitation for all analytes were within the range 1.2-56.7 μg kg(-1). The work presented in this paper illustrated the suitability and interesting potential of combining second-order calibration method with second-order analytical instrument for multi-residue analysis in honey samples.

  4. Performance characterization of a quantitative liquid chromatography-tandem mass spectrometric method for 12 macrolide and lincosamide antibiotics in salmon, shrimp and tilapia.

    PubMed

    Dickson, Leslie C

    2014-09-15

    This paper describes an extension and performance characterization of a quantitative confirmatory multi-residue liquid chromatography-tandem mass spectrometric method for residues of macrolide and lincosamide antibiotics, originally validated for application to bovine kidney tissues, to tissues of salmon, shrimp and tilapia. The 12 analytes include clindamycin, erythromycin A, gamithromycin, josamycin, lincomycin, neospiramycin 1, oleandomycin, pirlimycin, spiramycin 1, tildipirosin, tilmicosin and tylosin A. The limit of detection was 0.5 μg/kg. Within-laboratory precision evaluated over the analytical range of 5.0-50.0 μg/kg ranged from 4 to 17%. The accuracy of the method ranged from 80 to 112%. Recoveries ranged from 47 to 99% with all but one recovery above 60%. This is the first report of a quantitative confirmatory method for gamithromycin, pirlimycin and tildipirosin in fish and shrimp.

  5. Development of a fast screening and confirmatory method by liquid chromatography-quadrupole-time-of-flight mass spectrometry for glucuronide-conjugated methyltestosterone metabolite in tilapia.

    PubMed

    Amarasinghe, Kande; Chu, Pak-Sin; Evans, Eric; Reimschuessel, Renate; Hasbrouck, Nicholas; Jayasuriya, Hiranthi

    2012-05-23

    This paper describes the development of a fast method to screen and confirm methyltestosterone 17-O-glucuronide (MT-glu) in tilapia bile. The method consists of solid-phase extraction (SPE) followed by high-performance liquid chromatography-mass spectrometry. The system used was an Agilent 6530 Q-TOF with an Agilent Jet stream electrospray ionization interface. The glucuronide detected in the bile was characterized as MT-glu by comparison with a chemically synthesized standard. MT-glu was detected in bile for up to 7 days after dosing. Semiquantification was done with matrix-matched calibration curves, because MT-glu showed signal suppression due to matrix effects. This method provides a suitable tool to monitor the illegal use of methyltestosterone in tilapia culture.

  6. Ion chromatography electrospray ionization mass spectrometry method development and investigation of lithium hexafluorophosphate-based organic electrolytes and their thermal decomposition products.

    PubMed

    Kraft, Vadim; Grützke, Martin; Weber, Waldemar; Winter, Martin; Nowak, Sascha

    2014-08-08

    A method based on the coupling of ion chromatography (IC) and electrospray ionization mass spectrometry (ESI-MS) for the separation and determination of thermal decomposition products of LiPF6-based organic electrolytes is presented. The utilized electrolytes, LP30 and LP50, are commercially available and consist of 1mol/l LiPF6 dissolved in ethylene carbonate/dimethyl carbonate and ethylene carbonate/ethyl methyl carbonate, respectively. For the separation method development three ion chromatographic columns with different capacity and stationary phase were used and compared. Besides the known hydrolysis products of lithium hexafluorophosphate, several new organophosphates were separated and identified with the developed IC-ESI-MS method during aging investigations of the electrolytes. The chemical structures were elucidated with IC-ESI-MS/MS.

  7. Validation of a method for the determination of sterols and triterpenes in the aerial part of Justicia anselliana (Nees) T. Anders by capillary gas chromatography.

    PubMed

    Kpoviéssi, Dossou Sika Salomé; Gbaguidi, Fernand; Gbénou, Joachim; Accrombessi, Georges; Moudachirou, Mansourou; Rozet, Eric; Hubert, Philippe; Quetin-Leclercq, Joëlle

    2008-12-01

    An accurate and sensitive method, combining soxhlet extraction, solid phase-extraction and capillary gas chromatography is described for the quantitative determination of one triterpene (lupeol) and three sterols (stigmasterol, campesterol and beta-sitosterol) and the detection of another triterpene (alpha-amyrin) from the aerial part of Justicia anselliana. This is the first method allowing the quantification of sterols and triterpenes in this plant. It has been fully validated in order to be able to compare the sterol and triterpene composition of different samples of J. anselliana and therefore help to explain the allelopathic activity due to these compounds. This method showed that the aerial part of J. anselliana contained (292+/-2)mg/kg of lupeol, (206+/-1)mg/kg of stigmasterol, (266+/-2)mg/kg of campesterol and (184+/-9)mg/kg of beta-sitosterol.

  8. Determination of benzene at trace levels in air by a novel method based on solid-phase microextraction gas chromatography/mass spectrometry.

    PubMed

    Saba, A; Cuzzola, A; Raffaelli, A; Pucci, S; Salvadori, P

    2001-01-01

    A new method for the determination of benzene at trace levels in air is presented. The method consists of the collection of air samples on adsorbent cartridges with simultaneous adsorption of pre-established amounts of D6-labeled internal standard. Desorption from the cartridge is performed by solid-phase microextraction (SPME) with analysis by gas chromatography/mass spectrometry (GC/MS) using an ion trap mass spectrometer. The influence of several parameters (type of SPME fiber, temperature, time, for example) was investigated, and good linearity in the range 10-400 ng of C6D6, with a coefficient of variance (CV) around 3-5%, was obtained. The method was tested by sampling air in a town center in Italy, and a benzene concentration of approximately 50 microg/m(3) was determined. The maximum limit recommended by the European Community is 10 microg/m(3).

  9. Validation of a headspace trap gas chromatography and mass spectrometry method for the quantitative analysis of volatile compounds from degraded rapeseed oil.

    PubMed

    Sghaier, Lilia; Cordella, Christophe B Y; Rutledge, Douglas N; Watiez, Mickaël; Breton, Sylvie; Sassiat, Patrick; Thiebaut, Didier; Vial, Jérôme

    2016-05-01

    Due to lipid oxidation, off-flavors, characterized by a fishy odor, are emitted during the heating of rapeseed oil in a fryer and affect the flavor of rapeseed oil even at low concentrations. Thus, there is a need for analytical methods to identify and quantify these products. To study the headspace composition of degraded rapeseed oil, and more specifically the compounds responsible for the fishy odor, a headspace trap gas chromatography with mass spectrometry method was developed and validated. Six volatile compounds formed during the degradation of rapeseed oil were quantified: 1-penten-3-one, (Z)-4-heptenal, hexanal, nonanal, (E,E)-heptadienal, and (E)-2-heptenal. Validation using accuracy profiles allowed us to determine the valid ranges of concentrations for each compound, with acceptance limits of 40% and tolerance limits of 80%. This method was then successfully applied to real samples of degraded oils.

  10. Rapid and sensitive method for determining free amino acids in honey by gas chromatography with flame ionization or mass spectrometric detection.

    PubMed

    Nozal, Ma J; Bernal, J L; Toribio, M L; Diego, J C; Ruiz, A

    2004-08-20

    This paper describes a rapid, sensitive and specific method for determination of free amino acids in honey involving a new reaction of derivatization and gas chromatography (GC) with flame ionization (FID) and mass spectrometric (MS) detection. The method allows the determination of 22 free amino acids in honey samples in a short time: 8 and 5 min for GC-FID and GC-MS, respectively. Quantitation was performed using Norvaline as internal standard, with detection limits ranging between 0.112 and 1.795 mg/L by GC-FID and between 0.001 and 0.291 mg/L by GC-MS in the selected-ion monitoring mode. The method was validated and applied to a set of 74 honey samples belonging to four different botanical origins: eucaliptus, rosemary, orange and heather. The statistical treatment of data shows a correct classification of different origins over 90%.

  11. Improved liquid chromatography-tandem mass spectrometric method for the determination of ethyl glucuronide concentrations in hair: applications to forensic cases.

    PubMed

    Imbert, Laurent; Gaulier, Jean-Michel; Dulaurent, Sylvain; Morichon, Julien; Bevalot, Fabien; Izac, Paul; Lachâtre, Gérard

    2014-01-01

    Ethyl glucuronide (EtG) is a direct marker of ethanol consumption, and its assay in hair is an efficient tool for chronic alcoholism diagnosis. In 2012, the Society of Hair Testing proposed a new consensus for hair concentrations interpretation, strongly advising the use of analytical methods providing a limit of quantification of less than 3 pg/mg. The present work describes the optimization and validation of a previously developed liquid chromatography-tandem mass spectrometric method in order to comply with this recommendation. The concentration range of this improved method is from 3 to 1,000 pg/mg. Some cases are then described to illustrate the usefulness of hair EtG: a forensic post-mortem case and two cases of suspension of driving licences. Finally, hair samples of some teetotallers (n = 10) have been analyzed, which allowed neither to quantitate nor to detect any trace of EtG.

  12. Development and validation of a supercritical fluid chromatography method for the direct determination of enantiomeric purity of provitamin B5 in cosmetic formulations with mass spectrometric detection.

    PubMed

    Khater, Syame; West, Caroline

    2015-01-01

    A rapid and efficient chiral supercritical fluid chromatography (SFC) method has been developed for the quantitative determination of panthenol enantiomers in cosmetic formulations (cream, lotion, wipe, and exfoliant). Indeed, the pharmacological effect only depends on the D form (Dexpanthenol) thus accurate measurement of its enantiomeric purity in formulated cosmetic products is of interest. The samples were prepared with liquid-liquid extraction followed by solid-phase extraction on Adsorbex amino cartridges. After testing several enantioselective columns in an attempt at reversing the elution order to have the minor enantiomer eluted first, the best separation of enantiomers and internal standard (N-acetyl-L-alanine) was achieved on a 3 μm-amylose-type immobilized polysaccharide chiral stationary phase (Chiralpak IA) in less than 6 min with a simple mobile phase comprising carbon dioxide and 11% methanol pumped at 2.3 mL/min, 25°C and 150 bar backpressure. Supercritical fluid chromatography coupled to both an optical diode-array detector and a user-friendly single-quadrupole mass spectrometer (Waters QDa) equipped with electrospray ionization source has been used. The on-line coupling ensures the technique to be more informative and improves detection sensitivity, as underivatized panthenol has a poor UV absorption. The limit of quantification (LOQ) achieved with single-ion recording was 0.5 μg/mL. The method was validated in terms of linearity, precision and accuracy and satisfactory results were obtained.

  13. Epitope detection chromatography: a method to dissect the structural heterogeneity and inter-connections of plant cell-wall matrix glycans.

    PubMed

    Cornuault, Valérie; Manfield, Iain W; Ralet, Marie-Christine; Knox, J Paul

    2014-05-01

    Plant cell walls are complex, multi-macromolecular assemblies of glycans and other molecules and their compositions and molecular architectures vary extensively. Even though the chemistry of cell-wall glycans is now well understood, it remains a challenge to understand the diversity of glycan configurations and interactions in muro, and how these relate to changes in the biological and mechanical properties of cell walls. Here we describe in detail a method called epitope detection chromatography analysis of cell-wall matrix glycan sub-populations and inter-connections. The method combines chromatographic separations with use of glycan-directed monoclonal antibodies as detection tools. The high discrimination capacity and high sensitivity for the detection of glycan structural features (epitopes) provided by use of established monoclonal antibodies allows the study of oligosaccharide motifs on sets of cell-wall glycans in small amounts of plant materials such as a single organ of Arabidopsis thaliana without the need for extensive purification procedures. We describe the use of epitope detection chromatography to assess the heterogeneity of xyloglucan and pectic rhamnogalacturonan I sub-populations and their modulation in A. thaliana organs.

  14. Quantitative measurement of vitamin K2 (menaquinones) in various fermented dairy products using a reliable high-performance liquid chromatography method.

    PubMed

    Manoury, E; Jourdon, K; Boyaval, P; Fourcassié, P

    2013-03-01

    We evaluated menaquinone contents in a large set of 62 fermented dairy products samples by using a new liquid chromatography method for accurate quantification of lipo-soluble vitamin K(2), including distribution of individual menaquinones. The method used a simple and rapid purification step to remove matrix components in various fermented dairy products 3 times faster than a reference preparation step. Moreover, the chromatography elution time was significantly shortened and resolution and efficiency were optimized. We observed wide diversity of vitamin K(2) contents in the set of fermented dairy products, from undetectable to 1,100 ng/g of product, and a remarkable diversity of menaquinone forms among products. These observations relate to the main microorganism species currently in the different fermented product technologies. The major form in this large set of fermented dairy products was menaquinone (MK)-9, and contents of MK-9 and MK-8 forms were correlated, that of MK-9 being around 4 times that of MK-8, suggesting that microorganisms able to produce MK-9 also produce MK-8. This was not the case for the other menaquinones, which were produced independently of each other. Finally, no obvious link was established between MK-9 content and fat content or pH of the fermented dairy products.

  15. A quick, easy, cheap, effective, rugged, and safe sample pretreatment and liquid chromatography with tandem mass spectrometry method for the simultaneous quantification of 33 mycotoxins in Lentinula edodes.

    PubMed

    Han, Zheng; Feng, Zhihong; Shi, Wen; Zhao, Zhihui; Wu, Yongjiang; Wu, Aibo

    2014-08-01

    Lentinula edodes, one of the most cultivated edible fungi in the world, are usually neglected for mycotoxins contamination due to the initial thinking of its resistance to mycotoxingenic molds. In the present study, a sensitive and reliable liquid chromatography with tandem mass spectrometry method was developed for the simultaneous quantification of 33 mycotoxins in L. edodes. Targeted mycotoxins were extracted using a quick, easy, cheap, effective, rugged, and safe procedure without any further clean-up step, and analyzed by liquid chromatography with tandem mass spectrometry on an Agilent Poroshell 120 EC-C18 column (100 × 3 mm, 2.7 μm) with a linear gradient elution program using water containing 5 mM ammonium acetate and methanol as the mobile phase. After validation by determining linearity (R(2) > 0.99), sensitivity (LOQ ≤ 20 ng/kg), recovery (73.6-117.9%), and precision (0.8-19.5%), the established method has been successfully applied to reveal the contamination states of various mycotoxins in L. edodes. Among the 30 tested samples, 22 were contaminated by various mycotoxins with the concentration levels ranging from 3.3-28,850.7 μg/kg, predicting that the edible fungus could be infected by the mycotoxins-producing fungi. To the best of our knowledge, this is the first report about real mycotoxins contamination in L. edodes.

  16. Hydrophilic interaction liquid chromatography-tandem mass spectrometry quantitative method for the cellular analysis of varying structures of gemini surfactants designed as nanomaterial drug carriers.

    PubMed

    Donkuru, McDonald; Michel, Deborah; Awad, Hanan; Katselis, George; El-Aneed, Anas

    2016-05-13

    Diquaternary gemini surfactants have successfully been used to form lipid-based nanoparticles that are able to compact, protect, and deliver genetic materials into cells. However, what happens to the gemini surfactants after they have released their therapeutic cargo is unknown. Such knowledge is critical to assess the quality, safety, and efficacy of gemini surfactant nanoparticles. We have developed a simple and rapid liquid chromatography electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) method for the quantitative determination of various structures of gemini surfactants in cells. Hydrophilic interaction liquid chromatography (HILIC) was employed allowing for a short simple isocratic run of only 4min. The lower limit of detection (LLOD) was 3ng/mL. The method was valid to 18 structures of gemini surfactants belonging to two different structural families. A full method validation was performed for two lead compounds according to USFDA guidelines. The HILIC-MS/MS method was compatible with the physicochemical properties of gemini surfactants that bear a permanent positive charge with both hydrophilic and hydrophobic elements within their molecular structure. In addition, an effective liquid-liquid extraction method (98% recovery) was employed surpassing previously used extraction methods. The analysis of nanoparticle-treated cells showed an initial rise in the analyte intracellular concentration followed by a maximum and a somewhat more gradual decrease of the intracellular concentration. The observed intracellular depletion of the gemini surfactants may be attributable to their bio-transformation into metabolites and exocytosis from the host cells. Obtained cellular data showed a pattern that grants additional investigations, evaluating metabolite formation and assessing the subcellular distribution of tested compounds.

  17. A salting out-acetonitrile homogeneous extraction coupled with gas chromatography-mass spectrometry method for the simultaneous determination of thirteen N-nitrosamines in skin care cosmetics.

    PubMed

    Dong, Hao; Guo, Xindong; Xian, Yanping; Luo, Haiying; Wang, Bin; Wu, Yuluan

    2015-11-27

    A sensitive gas chromatography-mass spectrometry method was established for the simultaneous determination of thirteen N-nitrosamines (NAs) in skin care cosmetics. The cosmetics samples were firstly dispersed by water and subsequently extracted and purified using salting out-acetonitrile homogeneous extraction method. Finally, the extracting solution was concentrated by slow nitrogen gas blowing. All of the samples were separated by INNOWAX capillary chromatographic column, and detected by selected ion monitoring (SIM) mode of gas chromatography-mass spectrometry (GC-MS) and quantified by isotope internal standard method. The method was validated for linearity and range, accuracy, precision and sensitivity. Under the optimized condition, the calibration curves were linear over the selected concentration ranges of 2-500μg/L for all the thirteen analytes, with calculated coefficients of determination (R(2)) of greater than 0.996. The limits of detection (LODs) and the limits of quantitation (LOQs) of the method were 3-15μg/kg and 10-50μg/kg, respectively. Recoveries were calculated at three levels of concentration spiked in two kinds of cosmetics (skin care cream and water). The values were found between 93.8% and 121.0% with relative standard deviation (RSD) values of 2.5-7.2% for intra-day precision (n=6) and 3.3-6.7% for inter-day precision (n=5). The method was successfully applied to analyze twenty-two cosmetics samples and N-nitrosodimethylamine (NDMA) was detected in one sample with the concentration of 207μg/kg.

  18. A column switching ultrahigh-performance liquid chromatography-tandem mass spectrometry method to determine anandamide and 2-arachidonoylglycerol in plasma samples.

    PubMed

    Marchioni, Camila; de Souza, Israel Donizeti; Grecco, Caroline Fernandes; Crippa, José Alexandre; Tumas, Vitor; Queiroz, Maria Eugênia Costa

    2017-03-23

    This study reports a fast, sensitive, and selective column switching ultrahigh-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method to determine the endocannabinoids (eCBs), anandamide (AEA), and 2-arachidonoylglycerol (2-AG) in plasma samples. This bidimensional system used a restricted access media column (RP-8 ADS, 25 mm × 4 mm × 25 μM) in the first dimension and a core-shell Kinetex C18 (100 mm × 2, 1.7 mm × 1 μM) column in the second dimension, followed by detection in a mass spectrometer triple quadrupole (multiple reactions monitoring mode) operating in the positive mode. RP-8 ADS was used for trace enrichment of eCBs (reverse phase partitioning) and macromolecular matrix size exclusion; the core-shell column was used for the chromatographic separation. The column switching UHPLC-MS/MS method presented a linear range spanning from 0.1 ng mL(-1) (LOQ) to 6 ng mL(-1) for AEA and from 0.04 ng mL(-1) (LOQ) to 10 ng mL(-1) for 2-AG. Excluding the LLOQ values, the precision assays provided coefficients of variation lower than 8% and accuracy with relative standard error values lower than 14%. Neither carryover nor matrix effects were detected. This high-throughput column switching method compared to conventional methods is time saving as it involves fewer steps, consumes less solvent, and presents lower LLOQ. The column switching UHPLC-MS/MS method was successfully applied to determine AEA and 2-AG in plasma samples obtained from Alzheimer's disease patients. Graphical abstract A column switching ultra high-performance liquid chromatography-tandem mass spectrometry method using RP-8 ADS column and core shell column to determine endocannabinoids in plasma samples.

  19. Development and validation of an ultra-performance liquid chromatography quadrupole time of flight mass spectrometry method for rapid quantification of free amino acids in human urine.

    PubMed

    Joyce, Richard; Kuziene, Viktorija; Zou, Xin; Wang, Xueting; Pullen, Frank; Loo, Ruey Leng

    2016-01-01

    An ultra-performance liquid chromatography quadrupole time of flight mass spectrometry (UPLC-qTOF-MS) method using hydrophilic interaction liquid chromatography was developed and validated for simultaneous quantification of 18 free amino acids in urine with a total acquisition time including the column re-equilibration of less than 18 min per sample. This method involves simple sample preparation steps which consisted of 15 times dilution with acetonitrile to give a final composition of 25 % aqueous and 75 % acetonitrile without the need of any derivatization. The dynamic range for our calibration curve is approximately two orders of magnitude (120-fold from the lowest calibration curve point) with good linearity (r (2) ≥ 0.995 for all amino acids). Good separation of all amino acids as well as good intra- and inter-day accuracy (<15 %) and precision (<15 %) were observed using three quality control samples at a concentration of low, medium and high range of the calibration curve. The limits of detection (LOD) and lower limit of quantification of our method were ranging from approximately 1-300 nM and 0.01-0.5 µM, respectively. The stability of amino acids in the prepared urine samples was found to be stable for 72 h at 4 °C, after one freeze thaw cycle and for up to 4 weeks at -80 °C. We have applied this method to quantify the content of 18 free amino acids in 646 urine samples from a dietary intervention study. We were able to quantify all 18 free amino acids in these urine samples, if they were present at a level above the LOD. We found our method to be reproducible (accuracy and precision were typically <10 % for QCL, QCM and QCH) and the relatively high sample throughput nature of this method potentially makes it a suitable alternative for the analysis of urine samples in clinical setting.

  20. New process for purifying high purity α1-antitrypsin from Cohn Fraction IV by chromatography: A promising method for the better utilization of plasma.

    PubMed

    Huangfu, Chaoji; Zhang, Jinchao; Ma, Yuyuan; Jia, Junting; Lv, Maomin; Zhao, Xiong; Zhang, Jingang

    2017-03-01

    α1-antitrypsin (AAT) is a 52kDa serine protease inhibitor that is abundant in plasma. It is synthesized mainly by hepatic cells, and widely used to treat patients with emphysema due to congenital deficiency of AAT. A new isolation method for the purification of AAT from Cohn Fraction IV (Cohn F IV) is described. Cohn F IV is usually discarded as a byproduct from Cohn process. Using Cohn F IV as starting material does not interfere with the production of other plasma proteins and the cost of purification could be reduced greatly. Parameters of each step during purification were optimized, 15% polyethyleneglycol (PEG) concentration and pH 5.2 for PEG precipitation, elution with 0.05M sodium acetate and pH 4.7 for ion-exchange chromatography, and two steps blue sepharose affinity chromatography were chosen for AAT purification. The final protein with purity of 98.17%, specific activity of 3893.29 IU/mg, and yield of 28.35%, was achieved. Western blotting was applied for qualitative identification of final product, which specifically reacted with goat anti-human AAT antibody. LC-ESI-MS/MS was also employed to confirm the final protein. High performance liquid chromatography was used to analyze the composition of purified protein suggesting that pure protein was achieved. The molecular weight of AAT is 51062.77Da which was identified by LC-MS-MS. The manufacturing process described here may make better use of human plasma with Cohn F IV as starting material. The simple process described in this study is simple and inexpensive, it has a potential value for large scale production.

  1. Comparison of two derivatization methods for the analysis of fatty acids and trans fatty acids in bakery products using gas chromatography.

    PubMed

    Salimon, Jumat; Omar, Talal A; Salih, Nadia

    2014-01-01

    Two different procedures for the methylation of fatty acids (FAs) and trans fatty acids (TFAs) in food fats were compared using gas chromatography (GC-FID). The base-catalyzed followed by an acid-catalyzed method (KOCH3/HCl) and the base-catalyzed followed by (trimethylsilyl)diazomethane (TMS-DM) method were used to prepare FA methyl esters (FAMEs) from lipids extracted from food products. In general, both methods were suitable for the determination of cis/trans FAs. The correlation coefficients (r) between the methods were relatively small (ranging from 0.86 to 0.99) and had a high level of agreement for the most abundant FAs. The significant differences (P = 0.05) can be observed for unsaturated FAs (UFAs), specifically for TFAs. The results from the KOCH3/HCl method showed the lowest recovery values (%R) and higher variation (from 84% to 112%), especially for UFAs. The TMS-DM method had higher R values, less variation (from 90% to 106%), and more balance between variation and %RSD values in intraday and interday measurements (less than 4% and 6%, resp.) than the KOCH3/HCl method, except for C12:0, C14:0, and C18:0. Nevertheless, the KOCH3/HCl method required shorter time and was less expensive than the TMS-DM method which is more convenient for an accurate and thorough analysis of rich cis/trans UFA samples.

  2. Multi-residue method for the determination of pesticides and pesticide metabolites in honeybees by liquid and gas chromatography coupled with tandem mass spectrometry--Honeybee poisoning incidents.

    PubMed

    Kiljanek, Tomasz; Niewiadowska, Alicja; Semeniuk, Stanisław; Gaweł, Marta; Borzęcka, Milena; Posyniak, Andrzej

    2016-02-26

    A method for the determination of 200 pesticides and pesticide metabolites in honeybee samples has been developed and validated. Almost 98% of compounds included in this method are approved to use within European Union, as active substances of plant protection products or veterinary medicinal products used by beekeepers to control mites Varroa destructor in hives. Many significant metabolites, like metabolites of imidacloprid, thiacloprid, fipronil, methiocarb and amitraz, are also possible to detect. The sample preparation was based on the buffered QuEChERS method. Samples of bees were extracted with acetonitrile containing 1% acetic acid and then subjected to clean-up by dispersive solid phase extraction (dSPE) using a new Z-Sep+ sorbent and PSA. The majority of pesticides, including neonicotionoids and their metabolites, were analyzed by liquid chromatography tandem mass spectrometry (LC-MS/MS) but some of pesticides, especially pyrethroid insecticides, were analyzed by gas chromatography tandem mass spectrometry (GC-MS/MS). The procedure was validated according to the Guidance document SANCO/12571/2013 at four concentration levels: 1, 5, 10 and 100 ng/g bees and verified in the international proficiency test. The analysis of bee samples spiked at the limit of quantification (LOQ) showed about 98% mean recovery value (trueness) and 97% of analytes showed recovery in the required range of 70-120% and RSDr (precision) below 20%. Linearity and matrix effects were also established. The LOQs of pesticides were in the range of 1-100 ng/g. The developed method allows determination of insecticides at concentrations of 10 ng/g or less, except abamectin and tebufenozide. LOQ values are lower than the median lethal doses LD50 for bees. The method was used to investigate more than 70 honeybee poisoning incidents. Data about detected pesticides and their metabolites are included.

  3. Optimization and validation of a fast supercritical fluid chromatography method for the quantitative determination of vitamin D3 and its related impurities.

    PubMed

    Andri, B; Lebrun, P; Dispas, A; Klinkenberg, R; Streel, B; Ziemons, E; Marini, R D; Hubert, Ph

    2017-03-31

    In the uprising context of green analytical chemistry, Supercritical Fluid Chromatography (SFC) is often suggested as an alternative to Normal Phase Liquid Chromatography. Indeed, SFC provides fast, efficient and green separations. In this report, the quantitative performances of SFC were challenged on a real-life case study: the Quality Control (QC) of vitamin D3. A rapid and green SFC method was optimized thanks to the Design of Experiments-Design Space (DoE-DS) methodology. It provided robust and high quality separation of the compounds within a 2min timeframe, using a gradient of ethanol as co-solvent of the carbon dioxide. The analytical method was fully validated according to the total error approach, demonstrating the compliance of the method to the specifications of U.S. Pharmacopeia (USP: 97.0-103.0%) and European Pharmacopeia (EP: 97.0-102.0%) for an interval of [50-150%] of the target concentration. In order to allow quantification of impurities using vitamin D3 as an external standard in SFC-UV, correction factors were determined and verified during method validation. Thus, accurate quantification of impurities was demonstrated at the specified levels (0.1 and 1.0% of the main compound) for a 70.0-130.0% dosing range. This work demonstrates the validity of an SFC method for the QC of vitamin D3 raw material and its application to real samples. Therefore, it supports the switch to a greener and faster separative technique as an alternative to NPLC in the pharmaceutical industry.

  4. A novel liquid chromatography/tandem mass spectrometry method for the quantification of glycine as biomarker in brain microdialysis and cerebrospinal fluid samples within 5min.

    PubMed

    Voehringer, Patrizia; Fuertig, René; Ferger, Boris

    2013-11-15

    Glycine is an important amino acid neurotransmitter in the central nervous system (CNS) and a useful biomarker to indicate biological activity of drugs such as glycine reuptake inhibitors (GRI) in the brain. Here, we report how a liquid chromatography/tandem mass spectrometry (LC-MS/MS) method for the fast and reliable analysis of glycine in brain microdialysates and cerebrospinal fluid (CSF) samples has been established. Additionally, we compare this method with the conventional approach of high performance liquid chromatography (HPLC) coupled to fluorescence detection (FD). The present LC-MS/MS method did not require any derivatisation step. Fifteen microliters of sample were injected for analysis. Glycine was detected by a triple quadrupole mass spectrometer in the positive electrospray ionisation (ESI) mode. The total running time was 5min. The limit of quantitation (LOQ) was determined as 100nM, while linearity was given in the range from 100nM to 100μM. In order to demonstrate the feasibility of the LC-MS/MS method, we measured glycine levels in striatal in vivo microdialysates and CSF of rats after administration of the commercially available glycine transporter 1 (GlyT1) inhibitor LY 2365109 (10mg/kg, p.o.). LY 2365109 produced 2-fold and 3-fold elevated glycine concentrations from 1.52μM to 3.6μM in striatal microdialysates and from 10.38μM to 36μM in CSF, respectively. In conclusion, we established a fast and reliable LC-MS/MS method, which can be used for the quantification of glycine in brain microdialysis and CSF samples in biomarker studies.

  5. Coordinating fingerprint determination of solid-phase microextraction/gas chromatography-mass spectrometry and chemometric methods for quality control of oxidized tallow.

    PubMed

    Song, Shiqing; Zhang, Xiaoming; Hayat, Khizar; Xiao, Zuobing; Niu, Yunwei; Eric, Karangwa

    2013-02-22

    Based on optimized solid-phase microextraction/gas chromatography-mass spectrometry (SPME/GC-MS) and chemometric methods, simple, reliable and reproducible methods were described for the first time for developing a chromatographic fingerprint of oxidized tallow. Eight optimal oxidized tallow samples were used to establish the chromatographic fingerprint. Spectral correlative chromatogram was adopted to identify 33 "common components". The validation of fingerprint analysis was performed based on the relative retention time, the relative peak area of common peaks, sample stability and similarity analysis. The correlation coefficient of similarity of eight optimal oxidized tallow samples was more than 0.962, which showed that samples from different batches were consistent to some extent in spite of slightly different chemical indexes. Through principal component analysis (PCA), 14 constituents were further screened out to be the main chemical markers, which could be applied to more accurate quantitative discrimination and quality control of oxidized tallow.

  6. A High-Performance Liquid Chromatography-Based Screening Method for the Analysis of Atrazine, Alachlor, and Ten of Their Transformation Products

    USGS Publications Warehouse

    Schroyer, B.R.; Capel, P.D.

    1996-01-01

    A high-performance liquid Chromatography (HPLC) method is presented for the for the fast, quantitative analysis of the target analytes in water and in low organic-carbon, sandy soils that are known to be contaminated with the parent herbicides. Speed and ease of sample preparation was prioritized above minimizing detection limits. Soil samples were extracted using 80:20 methanol:water (volume:volume). Water samples (50 ??L) were injected directly into the HPLC without prior preparation. Method quantification limits for soil samples (10 g dry weight) and water samples ranged from 20 to 110 ng/g and from 20 to 110 ??g/L for atrazine and its transformation products and from 80 to 320 ng/g and from 80 to 320 ??g/L for alachlor and its transformation products, respectively.

  7. A rapid monitoring method for inorganic arsenic in rice flour using reversed phase-high performance liquid chromatography-inductively coupled plasma mass spectrometry.

    PubMed

    Narukawa, Tomohiro; Chiba, Koichi; Sinaviwat, Savarin; Feldmann, Jörg

    2017-01-06

    A new rapid monitoring method by means of high performance liquid chromatography-inductively coupled plasma mass spectrometry (HPLC-ICP-MS) following the heat-assisted extraction was developed for measurement of total inorganic arsenic species in rice flour. As(III) and As(V) eluted at the same retention time and completely separated from organoarsenic species by an isocratic elution program on a reversed phase column. Therefore, neither ambiguous oxidation of arsenite to arsenate nor the integration of two peaks were necessary to determine directly the target analyte inorganic arsenic. Rapid injection allowed measuring 3 replicates within 6min and this combined with a quantitative extraction of all arsenic species from rice flour by a 15min HNO3-H2O2 extraction makes this the fastest laboratory based method for inorganic arsenic in rice flour.

  8. Liquid chromatography-electrospray ionization-mass spectrometry method in multiple reaction monitoring mode to determine 17alpha-ethynylestradiol residues in cattle hair without previous digestion.

    PubMed

    Pedreira, S; Lolo, M; Vázquez, B I; Franco, C M; Cepeda, A; Fente, C

    2007-11-14

    A liquid chromatography-mass spectrometric (LC-MS/MS) method has been developed for determination of ethynylestradiol residues in cattle hair. Hair samples were pulverized with a cryogenic mill followed by a simple extraction with acetonitrile. A dansyl derivatization procedure to improve ethynylestradiol detection was used before the LC-MS/MS analysis in multiple reaction monitoring (MRM) mode using alpha-estradiol as an internal standard. The method was validated following the latest EU guidelines using blank hair samples spiked at 2 ng g(-1). The detection capability (CCbeta) was less than 2 ng g(-1), and the decision limit (CCalpha) was 1 ng g(-1). Incurred samples obtained 56 days after cow treatment with ethynylestradiol were analyzed, and the presence of ethynylestradiol in the hair was confirmed in all cases.

  9. Rapid liquid chromatography/tandem mass spectrometer (LCMS) method for clozapine and its metabolite N-desmethyl clozapine (norclozapine) in human serum.

    PubMed

    Rao, L V; Snyder, M L; Vallaro, G M

    2009-01-01

    Clozapine is indicated for the treatment of schizophrenia and related psychotic disorders. Several methods have been developed for monitoring Clozapine levels; however, they possess limited specificity and are often laborious. This study describes a simple liquid chromatography/tandem mass spectrometer (LCMS) method in human serum. The ion transitions monitored were m/z 327, 270, 296 for Clozapine, m/z 313, 192, 227 for Norclozapine and m/z 328, 271 for Loxapine. The assay is linear (25-1000 ng/ml) and showed a good correlation (r=0.98) within the analytical range of 79-1210 ng/ml in human serum. This assay is highly specific and sensitive for the simultaneous measurements of Clozapine and Norclozapine. The simplification of this assay makes it ideal for high throughput analyses of the patient samples in a routine clinical laboratory staffed with general medical technologists.

  10. Extractionless and sensitive method for high-throughput quantitation of cetirizine in human plasma samples by liquid chromatography-tandem mass spectrometry.

    PubMed

    de Jager, A D; Hundt, H K L; Swart, K J; Hundt, A F; Els, J

    2002-06-25

    Following a single 10-mg oral dose of cetirizine dihydrochloride to 24 healthy volunteers, the analyte was quantified in human plasma. Protein precipitation using acetonitrile (ACN) was followed by reversed-phase liquid chromatography and tandem mass spectrometry. The MS/MS method was optimised using a PE Sciex API 2000 triple quadrupole mass spectrometer in selected reaction monitoring (SRM) mode, using electrospray with positive ionisation. Oxybutynin was used as the internal standard. The assay method represents a robust, high-throughput, highly specific and sensitive quantitative assay procedure, with 0.5 ng/ml being the lowest plasma concentration that could be reliably quantified. The procedure involves minimal sample preparation, and is well suited to clinical studies of the drug involving large numbers of generated samples. Pre-dose as well as post-dose samples up to and including 48 h were quantified, and the data generated were used to determine the pharmacokinetic profile of the drug.

  11. Anatomical study of secondary tuberized roots of Harpagophytum procumbens DC and quantification of harpagoside by high-performance liquid chromatography method

    PubMed Central

    Babili, Fatiha El; Fouraste, I.; Rougaignon, C.; Moulis, C.; Chatelain, C.

    2012-01-01

    Aim and Background: A botanical study is conducted to provide a standard diagnostic tool. In order to improve the quality assurance of the secondary tuberized roots of Harpagophytum procumbens, derived extract and phytomedicine, a simple, rapid, and accurate high-performance liquid chromatography (HPLC) method was developed to assess the harpagoside. Material and Mehods: This HPLC assay was performed on a reversedphase C18 column with methanol and water (50/50–V/V) as the mobile phase with a flow rate of 1.5 mL/min and using a monitoring wavelength at 278 nm. Results and Conclusion: This method was successfully applied to quantify these bioactive iridoid in an aqueous extract of H. procumbens and in its related phytomedicine “harpagophyton”. The result demonstrated that the quantification of harpagoside, indicating that the quality control of the bioactive ingredient in H. procumbens, derived extract and phytomedicine, is critical to ensure its clinical benefits. PMID:22701294

  12. Development and validation of automatic HS-SPME with a gas chromatography-ion trap/mass spectrometry method for analysis of volatiles in wines.

    PubMed

    Paula Barros, Elisabete; Moreira, Nathalie; Elias Pereira, Giuliano; Leite, Selma Gomes Ferreira; Moraes Rezende, Claudia; Guedes de Pinho, Paula

    2012-11-15

    An automated headspace solid-phase microextraction (HS-SPME) combined with gas chromatography-ion trap/mass spectrometry (GC-IT/MS) was developed in order to quantify a large number of volatile compounds in wines such as alcohols, ester, norisoprenoids and terpenes. The procedures were optimized for SPME fiber selection, pre-incubation temperature and time, extraction temperature and time, and salt addition. A central composite experimental design was used in the optimization of the extraction conditions. The volatile compounds showed optimal extraction using a DVB/CAR/PDMS fiber, incubation of 5 ml of wine with 2g NaCl at 45 °C during 5 min, and subsequent extraction of 30 min at the same temperature. The method allowed the identification of 64 volatile compounds. Afterwards, the method was validated successfully for the most significant compounds and was applied to study the volatile composition of different white wines.

  13. Quantitative and chemical fingerprint analysis for the quality evaluation of Isatis indigotica based on ultra-performance liquid chromatography with photodiode array detector combined with chemometric methods.

    PubMed

    Shi, Yan-Hong; Xie, Zhi-Yong; Wang, Rui; Huang, Shan-Jun; Li, Yi-Ming; Wang, Zheng-Tao

    2012-01-01

    A simple and reliable method of ultra-performance liquid chromatography with photodiode array detector (UPLC-PDA) was developed to control the quality of Radix Isatidis (dried root of Isatis indigotica) for chemical fingerprint analysis and quantitative analysis of eight bioactive constituents, including R,S-goitrin, progoitrin, epiprogoitrin, gluconapin, adenosine, uridine, guanosine, and hypoxanthine. In quantitative analysis, the eight components showed good regression (R > 0.9997) within test ranges, and the recovery method ranged from 99.5% to 103.0%. The UPLC fingerprints of the Radix Isatidis samples were compared by performing chemometric procedures, including similarity analysis, hierarchical clustering analysis, and principal component analysis. The chemometric procedures classified Radix Isatidis and its finished products such that all samples could be successfully grouped according to crude herbs, prepared slices, and adulterant Baphicacanthis cusiae Rhizoma et Radix. The combination of quantitative and chromatographic fingerprint analysis can be used for the quality assessment of Radix Isatidis and its finished products.

  14. Improved high-performance liquid chromatography-diode array detection method for the determination of phenolic compounds in leaves and peels from different apple varieties.

    PubMed

    Kindt, Maddalena; Orsini, M Cristina; Costantini, Barbara

    2007-09-01

    Hydroxycinnamic acids derivatives, monomeric and oligomeric flavan-3-ols, flavonols, and dihydrocalcones are four of the major polyphenolic groups found in apples leaves and peels. A simple extraction with minimal pre-treatment and a high-performance liquid chromatography-diode array detection determination are optimized and validated, in order to identify and quantitate the polyphenolic profile of leaves and peels of four apples varieties (Gala, Topaz, Golden Delicious, and Florina). The improved chromatographic method has led to better separation of some known polyphenols in a single course, and diode-array detection has been used for the previsional identification of some polyphenolic compounds not available as standards. Because the mobile phase and the chromatographic column are compatible with a mass spectrometer, this method could investigate the unknown flavanols, flavonols, hydrocinnamic acid derivatives, and chalcone-related compounds found in apple leaves and peel extracts analyzed.

  15. Optimization and single-laboratory validation study of a high-performance liquid chromatography (HPLC) method for the determination of phenolic Echinacea constituents.

    PubMed

    Brown, Paula N; Chan, Michael; Betz, Joseph M

    2010-07-01

    Three species of Echinacea (Echinacea purpurea, Echinacea angustifolia, and Echinacea pallida) are commonly used for medicinal purposes. The phenolic compounds caftaric acid, cichoric acid, echinacoside, cynarin, and chlorogenic acid are among the phytochemical constituents that may be responsible for the purported beneficial effects of the herb. Although methods for the analysis for these compounds have been published, documentation of their validity was inadequate as the accuracy and precision for the detection and quantification of these phenolics was not systematically determined and/or reported. To address this issue, the high-performance liquid chromatography method, originally developed by the Institute for Nutraceutical Advancement (INA), was reviewed, optimized, and validated for the detection and quantification of these phenolic compounds in Echinacea roots and aerial parts.

  16. A rapid method for chemical fingerprint analysis of Pan Panax notoginseng powders by ultra performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry.

    PubMed

    Liu, Peng; Yu, He-Shuil; Zhang, Li-Juan; Song, Xin-Bo; Kang, Li-Ping; Liu, Jing-Yuan; Zhang, Jie; Cao, Man; Yu, Kate; Kang, Ting-Guo; Ma, Bai-Ping

    2015-06-01

    A method coupling ultra-performance liquid chromatography (UPLC) with quadrupole time-of-flight mass spectrometer (Qtof MS) using the electrospray ionization (ESI) source was developed for the identification of the major saponins from Panax notoginseng powder (PNP). Ten different PNP samples were analyzed and evaluated for their quality by similarity evaluation and principle component analysis (PCA). Based on the accurate mass, summarized characteristic fragmentation behaviors, retention times of different types of saponins, related botanical biogenesis, and reported chromatographic behavior of saponins, fifty-one common peaks were effectively separated and identified, including 28 protopanaxadiol saponins and 18 protopanaxatriol saponins. Simultaneously, 15 significant discrepancy compounds were identified from the disqualified PNP samples. The established UPLC/Qtof MS fingerprint method was successfully applied for profiling and identifying the major saponins of PNP, providing a fast quality evaluation tool for distinguishing the authentic PNP and the adulterated products.

  17. Identification and characterization of vinylpyrrolidone-vinylimidazolium chloride copolymers in cosmetic products by pyrolysis-gas chromatography-mass spectrometry method.

    PubMed

    Gmahl, E; Ruess, W

    1993-04-01

    Synopsis Commercially available copolymers of 1-vinyl-2-pyrrolidone and 1-vinyl-3-methylimidazolium chloride, known as 'Luviquat' types in the cosmetic industry, were analysed for their composition using a combination of pyrolysis-gas chromatography-mass spectrometry method. This is a report on the determined pyrolytic products and the fast identification of the analysed polymers both in raw materials and cosmetic products. Calibration with defined material ensures the determination of monomer ratios with good reproducibility. Résumé Les copolymères de chlorure de 1-vinyle-2-pyrolidone et de 1-vinyle-3-methylimidazolium disponibles dans le commerce, connus dans l'industrie cosmétique sous la dénomination de copolymères de vinyle, ont été analysés à laide d'une méthode combinant la pyrolyse, la spectrométrie de mass et la chromatographie en phase gazeuse. Cet article constitue un rapport sur les produits déterminés par pyrolyse et sur la rapidité d'identification des polymères analysés à la fois dans des matières premières et dans des produits cosmétiques. Le calibrage avec un matériel défini assure une bonne détermination des taux de monomères dotés d'une reproductibilité.

  18. Pseudotargeted metabolomics method and its application in serum biomarker discovery for hepatocellular carcinoma based on ultra high-performance liquid chromatography/triple quadrupole mass spectrometry.

    PubMed

    Chen, Shili; Kong, Hongwei; Lu, Xin; Li, Yong; Yin, Peiyuan; Zeng, Zhongda; Xu, Guowang

    2013-09-03

    Untargeted analysis performed using full-scan mass spectrometry (MS) coupled with liquid chromatography (LC) is commonly used in metabolomics. Although they are commonly employed, full-scan MS methods such as quadrupole-time-of-flight (Q-TOF) MS have been restricted by various factors including their limited linear range and complicated data processing. LC coupled with triple quadrupole (QQQ) MS operated in the multiple reaction monitoring (MRM) mode is the gold standard for metabolite quantification; however, only known metabolites are generally quantified, limiting its applications in metabolomic analysis. In this study, a pseudotargeted approach was proposed to perform serum metabolomic analysis using an ultra high-performance liquid chromatography (UHPLC)/QQQ MS system operated in the MRM mode, for which the MRM ion pairs were acquired from the serum samples through untargeted tandem MS using UHPLC/Q-TOF MS. The UHPLC/QQQ MRM MS-based pseudotargeted method displayed better repeatability and wider linear range than the traditional UHPLC/Q-TOF MS-based untargeted metabolomics method, and no complicated peak alignment was required. The developed method was applied to discover serum biomarkers for patients with hepatocellular carcinoma (HCC). Patients with HCC had decreased lysophosphatidylcholine, increased long-chain and decreased medium-chain acylcarnitines, and increased aromatic and decreased branched-chain amino acid levels compared to healthy controls. The novelty of this work is that it provides an approach to acquire MRM ion pairs from real samples, is not limited to metabolite standards, and it provides a foundation to achieve pseudotargeted metabolomic analysis on the widely used LC/QQQ MS platform.

  19. A rapid method for the simultaneous determination of 25 anti-hypertensive compounds in dietary supplements using ultra-high-pressure liquid chromatography.

    PubMed

    Heo, Seok; Yoo, Geum Joo; Choi, Ji Yeon; Park, Hyoung Joon; Park, Sung-Kwan; Baek, Sun Young

    2016-11-01

    A novel, stable, simple and specific ultra-performance liquid chromatography method with ultraviolet detection (205 nm) for the simultaneous analysis of 25 anti-hypertensive substances was developed. The method was validated according to the International Conference of Harmonisation guidelines with respect to linearity, accuracy, precision, limit of detection (LOD), limit of quantitation (LOQ) and stability. From the ultra-performance liquid chromatography results, we identified the LOD and LOQ of solid samples to be 0.20-1.00 and 0.60-3.00 μg ml(-1), respectively, while those of liquid samples were 0.30-1.20 and 0.90-3.60 μg ml(-1), respectively. The linearity exceeded 0.9999, and the intra- and inter-day precisions were 0.15-6.48% and 0.28-8.67%, respectively. The intra- and inter-day accuracies were 82.25-111.42% and 80.70-115.64%, respectively, and the stability was lower than 12.9% (relative standard deviation). This method was applied to the monitoring of 97 commercially available dietary supplements obtained in Korea, such as pills, soft capsules, hard capsules, liquids, powders and tablets. The proposed method is accurate, precise and of high quality, and can be used for the routine, reproducible analysis and control of 25 anti-hypertensive substances in various dietary supplements. The work presented herein may help to prevent incidents related to food adulteration and restrict the illegal food market.

  20. A fast and sensitive method for the separation of carotenoids using ultra-high performance supercritical fluid chromatography-mass spectrometry.

    PubMed

    Jumaah, Firas; Plaza, Merichel; Abrahamsson, Victor; Turner, Charlotta; Sandahl, Margareta

    2016-08-01

    In this study, a rapid and sensitive ultra-high performance supercritical fluid chromatography-mass spectrometry (UHPSFC-MS) method has been developed and partially validated for the separation of carotenoids within less than 6 min. Six columns of orthogonal selectivity were examined, and the best separation was obtained by using a 1-aminoanthracene (1-AA) column. The length of polyene chain as well as the number of hydroxyl groups in the structure of the studied carotenoids determines their differences in the physiochemical properties and thus the separation that is achieved on this column. All of the investigated carotenoids were baseline separated with resolution values greater than 1.5. The effects of gradient program, back pressure, and column temperature were studied with respect to chromatographic properties such as retention and selectivity. Electrospray ionization (ESI) and atmospheric pressure chemical ionization (APCI) were compared in both positive and negative mode, using both direct infusion and hyphenated with UHPSFC. The ESI in positive mode provided the highest response. The coefficient of determination (R (2)) for all calibration curves were greater than 0.998. Limit of detection (LOD) was in the range of 2.6 and 25.2 ng/mL for α-carotene and astaxanthin, respectively, whereas limit of quantification (LOQ) was in the range of 7.8 and 58.0 ng/mL for α-carotene and astaxanthin, respectively. Repeatability and intermediate precision of the developed UHPSFC-MS method were determined and found to be RSD < 3 % and RSD < 6 %, respectively. The method was applied in order to determine carotenoids in supercritical fluid extracts of microalgae and rosehip. Graphical Abstract Ultra-high performance supercritical fluid chromatography-a rapid separation method for the analysis of carotenoids in rosehip and microalgae samples.

  1. A broad-spectrum equine urine screening method for free and enzyme-hydrolysed conjugated drugs with ultra performance liquid chromatography/tandem mass spectrometry.

    PubMed

    Wong, Colton H F; Tang, Francis P W; Wan, Terence S M

    2011-07-04

    The authors' laboratory at one time employed four liquid chromatography/mass spectrometric (LC/MS) methods for the detection of a large variety of drugs in equine urine. Drug classes covered by these methods included anti-diabetics, anti-ulcers, cyclooxygenase-2 (COX-2) inhibitors, sedatives, corticosteroids, anabolic steroids, sulfur diuretics, xanthines, etc. With the objective to reduce labour and instrumental workload, a new ultra performance liquid chromatography/tandem mass spectrometric (UPLC/MS/MS) method has been developed, which encompasses all target analytes detected by the original four LC/MS methods. The new method has better detection limits than the superseded methods. In addition, it covers new target analytes that could not be adequately detected by the four LC/MS methods. The new method involves solid-phase extraction (SPE) of two aliquots of equine urine using two Abs Elut Nexus cartridges. One aliquot of the urine sample is treated with β-glucuronidase before subjecting to SPE. A second aliquot of the same urine sample is processed directly using another SPE cartridge, so that drugs that are prone to decomposition during enzyme hydrolysis can be preserved. The combined eluate is analysed by UPLC/MS/MS using alternating positive and negative electrospray ionisation in the selected-reaction-monitoring mode. Exceptional chromatographic separation is achieved using an UPLC system equipped with a UPLC(®) BEH C18 column (10 cm L×2.1 mm ID with 1.7 μm particles). With this newly developed UPLC/MS/MS method, the simultaneous detection of 140 drugs at ppb to sub-ppb levels in equine urine can be achieved in less than 13 min inclusive of post-run equilibration. Matrix interference for the selected transitions at the expected retention times is minimised by the excellent UPLC chromatographic separation. The method has been validated for recovery and precision, and is being used regularly in the authors' laboratory as an important component of the

  2. A NEW METHOD OF PEAK DETECTION FOR ANALYSIS OF COMPREHENSIVE TWO-DIMENSIONAL GAS CHROMATOGRAPHY MASS SPECTROMETRY DATA.

    PubMed

    Kim, Seongho; Ouyang, Ming; Jeong, Jaesik; Shen, Changyu; Zhang, Xiang

    2014-06-01

    We develop a novel peak detection algorithm for the analysis of comprehensive two-dimensional gas chromatography time-of-flight mass spectrometry (GC×GC-TOF MS) data using normal-exponential-Bernoulli (NEB) and mixture probability models. The algorithm first performs baseline correction and denoising simultaneously using the NEB model, which also defines peak regions. Peaks are then picked using a mixture of probability distribution to deal with the co-eluting peaks. Peak merging is further carried out based on the mass spectral similarities among the peaks within the same peak group. The algorithm is evaluated using experimental data to study the effect of different cut-offs of the conditional Bayes factors and the effect of different mixture models including Poisson, truncated Gaussian, Gaussian, Gamma, and exponentially modified Gaussian (EMG) distributions, and the optimal version is introduced using a trial-and-error approach. We then compare the new algorithm with two existing algorithms in terms of compound identification. Data analysis shows that the developed algorithm can detect the peaks with lower false discovery rates than the existing algorithms, and a less complicated peak picking model is a promising alternative to the more complicated and widely used EMG mixture models.

  3. A general method for fractionation of plasma proteins. Dye-ligand affinity chromatography on immobilized Cibacron blue F3-GA.

    PubMed

    Gianazza, E; Arnaud, P

    1982-01-01

    The chromatographic behaviour of 27 different plasma proteins on fractionation of human plasma on immobilized Cibacron Blue F3-GA was studied. The column was eluted by using a three-step procedure. First, a low-molarity buffer (30 mM-H3PO4/Na3PO4, pH 7.0, I0.053) was used, then a linear salt gradient (0-1 M-NaCl in the buffer above) was applied, followed by a wash with two bed volumes of 1.0 M-NaCl. Finally, bound proteins were 'stripped' with 0.5 M-NaSCN. Up to 1 ml of whole plasma could be loaded per 5 ml bed volume. No denaturation of proteinase inhibitors or complement fractions was observed. The recovery of individual proteins ranged between 52 and greater than 95%. Enrichment of four individual plasma components (alpha 1-antitrypsin, caeruloplasmin, antithrombin III and haemopexin) was between 10-fold and 75-fold. These results indicate that chromatography on immobilized Cibacron Blue F3-GA can be a useful initial step in the purification of plasma proteins.

  4. Inverse Gas Chromatography with Film Cell Unit: An Attractive Alternative Method to Characterize Surface Properties of Thin Films.

    PubMed

    Klein, Géraldine L; Pierre, Guillaume; Bellon-Fontaine, Marie-Noëlle; Graber, Marianne

    2015-09-01

    Inverse gas chromatography (IGC) is widely used for the characterization of surfaces. The present work describes a novel IGC tool, the recently developed film cell module, which measures monolithic thin solid film surface properties, whereas only samples in powder or fiber state or polymer-coated supports can be studied by classic IGC. The surface energy of four different solid supports was measured using both classic IGC with columns packed with samples in the powder state, and IGC with the new film cell module or the sessile drop technique, using samples in the film state. The total surface energy and its dispersive and specific components were measured for glass, polyethylene, polyamide and polytetrafluoroethylene. Similar results were obtained for the four materials using the three different techniques. The main conclusion is that the new film cell module for IGC is an attractive alternative to the sessile drop technique as it gives very accurate and reproducible results for surface energy components, with significant savings in time and the possible control of sample humidity and temperature. This film cell module for IGC extends the application field of IGC to any thin solid film and can be used to study the effect of any surface treatment on surface energy.

  5. Determination of rosmarinic acid in sage and borage leaves by high-performance liquid chromatography with different detection methods.

    PubMed

    Bandoniene, Donata; Murkovic, Michael; Venskutonis, Petras R

    2005-08-01

    Rosmarinic acid is separated and identified on the basis of high-performance liquid chromatography (HPLC)-UV-mass spectrometry data in 80% methanol in water extracts from the leaves of Salvia species (S. officinalis, S. glutinosa, S. aethiopis, S. sclarea, and Borago officinalis) as a dominant radical scavenger towards the 2,2'-diphenyl-1-picrylhydrazyl (DPPH*) stable radical in HPLC-DPPH* system. The content of rosmarinic acid in the plants is calibrated and quantitated from chromatograms obtained by UV detection at 280 nm. The concentration ranges from 13.3 to 47.3 mg of the phenolic acid per gram dried leaves of all plants is tested. S. glutinosa and S. sclarea have the highest concentration of rosmarinic acid. The amount of rosmarinic acid in borage leaves is similar compared with Salvia officinalis (15 mg/g). The HPLC-DPPH* system is calibrated for quantitative DPPH* scavenging assessment of rosmarinic acid. The results reveal excellent correlation (r2 = 0.98) between the rosmarinic acid concentration and antiradical activity.

  6. [Studies on the occurrence of furan in food for infants by gas chromatography with mass spectrometry method].

    PubMed

    Minorczyk, Maria; Starski, Andrzej; Jedra, Małgorzata; Gawarska, Halina; Sawilska-Rautenstrauch, Dorota

    2011-01-01

    Furan is an organic compound formed during heat treatment. It has been shown to be carcinogenic in animal laboratory studies. The aim of this study was to determine the content of furan in vegetables and vegetable-meat products intended for infants. The testing system used during this study was gas chromatography coupled with mass spectrometry (GC/MS). The content of furan in 48 samples of processed food ready to eat has been determined. In all samples furan was detected within the range from 13.2 to 91.1 microg/kg, and its average value was 43.3 microg/kg. The paper estimate the exposure assessment of infants to furan found in food. The calculated exposure ranged from 0.23 to 1.77 microg/kg bw/day with the average content of furan in ready to eat products ranged from 35.3 to 52.2 microg/kg. Exposure did not exceed the ADI value 2 microg/kg bw/day.

  7. A hemagglutinin quantification method for development of an influenza pandemic vaccine using size exclusion high performance liquid chromatography.

    PubMed

    Roh, Hang Sik; Song, Hye Min; Yun, Bo Reum; Kang, Hyun Kyung; Choi, Keum Suk; Park, Yun Ju; Kim, Dong Sub; Kim, Seung Hee; Mo, In Pil; An, Beum-Soo; Ahn, Chi Young

    2015-04-01

    Single radial immunodiffusion (SRID) assay requires a reference antigen and an antibody to the hemagglutinin (HA) of an influenza vaccine. As it takes 2‑3 months to develop the reference antigen, vaccine development is delayed in cases of an influenza pandemic. In the present study, the measurement of the HA content of influenza vaccines was assessed using size exclusion high performance liquid chromatography (SE‑HPLC) for the rapid development of a pandemic vaccine. When the 2009 H1N1 reference antigen, pandemic 2009 H1N1 vaccine and 2010 seasonal influenza vaccines were analyzed by SE‑HPLC, the HA of the reference antigen and vaccines was specifically separated. The presence and specificity of HA were evidenced with immunoprecipitation and ELISA assays. For the influenza vaccines, the chromatogram pattern and retention time of HA were similar among the antigen types (2009 H1N1, 2010 H3N2 and 2010 B). In addition, when SE‑HPLC was applied, the ratio of HA chromatogram to peak area revealed a significant correlation with HA concentration for the reference antigen and vaccine. The result of the HA content calculation based on SE‑HPLC exhibited 99.91‑100% similarity, compared with that of SRID. These findings suggest that the measurement of peak area ratio/HA content using SE‑HPLC may be a substitute for SRID and rapidly measure HA content to enable faster development of a vaccine during an influenza pandemic.

  8. Comparison of four extraction/methylation analytical methods to measure fatty acid composition by gas chromatography in meat.

    PubMed

    Juárez, M; Polvillo, O; Contò, M; Ficco, A; Ballico, S; Failla, S

    2008-05-09

    Four different extraction-derivatization methods commonly used for fatty acid analysis in meat (in situ or one-step method, saponification method, classic method and a combination of classic extraction and saponification derivatization) were tested. The in situ method had low recovery and variation. The saponification method showed the best balance between recovery, precision, repeatability and reproducibility. The classic method had high recovery and acceptable variation values, except for the polyunsaturated fatty acids, showing higher variation than the former methods. The combination of extraction and methylation steps had great recovery values, but the precision, repeatability and reproducibility were not acceptable. Therefore the saponification method would be more convenient for polyunsaturated fatty acid analysis, whereas the in situ method would be an alternative for fast analysis. However the classic method would be the method of choice for the determination of the different lipid classes.

  9. Development of liquid chromatography methods coupled to mass spectrometry for the analysis of substances with a wide variety of polarity in meconium.

    PubMed

    Meyer-Monath, Marie; Chatellier, Claudine; Cabooter, Deirdre; Rouget, Florence; Morel, Isabelle; Lestremau, Francois

    2015-06-01

    Meconium is the first fecal excretion of newborns. This complex accumulative matrix allows assessing the exposure of the fetus to xenobiotics during the last 6 months of pregnancy. To determine the eventual effect of fetal exposure to micropollutants in this matrix, robust and sensitive analytical methods must be developed. This article describes the method development of liquid chromatography methods coupled to triple quadrupole mass spectrometry for relevant pollutants. The 28 selected target compounds had different physico-chemical properties from very polar (glyphosate) to non-polar molecules (pyrethroids). Tests were performed with three different types of columns: reversed phase, ion exchange and HILIC. As a unique method could not be determined for the simultaneous analysis of all compounds, three columns were selected and suitable chromatographic methods were optimized. Similar results were noticed for the separation of the target compounds dissolved in either meconium extract or solvent for reversed phase and ion exchange columns. However, for HILIC, the matrix had a significant influence on the peak shape and robustness of the method. Finally, the analytical methods were applied to "real" meconium samples.

  10. Compound-Specific Chlorine Isotope Analysis of Tetrachloromethane and Trichloromethane by Gas Chromatography-Isotope Ratio Mass Spectrometry vs Gas Chromatography-Quadrupole Mass Spectrometry: Method Development and Evaluation of Precision and Trueness.

    PubMed

    Heckel, Benjamin; Rodríguez-Fernández, Diana; Torrentó, Clara; Meyer, Armin; Palau, Jordi; Domènech, Cristina; Rosell, Mònica; Soler, Albert; Hunkeler, Daniel; Elsner, Martin

    2017-03-21

    Compound-specific chlorine isotope analysis of tetrachloromethane (CCl4) and trichloromethane (CHCl3) was explored by both, gas chromatography-isotope ratio mass spectrometry (GC-IRMS) and GC-quadrupole MS (GC-qMS), where GC-qMS was validated in an interlaboratory comparison between Munich and Neuchâtel with the same type of commercial GC-qMS instrument. GC-IRMS measurements analyzed CCl isotopologue ions, whereas GC-qMS analyzed the isotopologue ions CCl3, CCl2, CCl (of CCl4) and CHCl3, CHCl2, CHCl (of CHCl3), respectively. Lowest amount dependence (good linearity) was obtained (i) in H-containing fragment ions where interference of (35)Cl- to (37)Cl-containing ions was avoided; (ii) with tuning parameters favoring one predominant rather than multiple fragment ions in the mass spectra. Optimized GC-qMS parameters (dwell time 70 ms, 2 most abundant ions) resulted in standard deviations of 0.2‰ (CHCl3) and 0.4‰ (CCl4), which are only about twice as large as 0.1‰ and 0.2‰ for GC-IRMS. To compare also the trueness of both methods and laboratories, samples from CCl4 and CHCl3 degradation experiments were analyzed and calibrated against isotopically different reference standards for both CCl4 and CHCl3 (two of each). Excellent agreement confirms that true results can be obtained by both methods provided that a consistent set of isotopically characterized reference materials is used.

  11. Single solid phase extraction method for the simultaneous analysis of polar and non-polar pesticides in urine samples by gas chromatography and ultra high pressure liquid chromatography coupled to tandem mass spectrometry.

    PubMed

    Cazorla-Reyes, Rocío; Fernández-Moreno, José Luis; Romero-González, Roberto; Frenich, Antonia Garrido; Vidal, José Luis Martínez

    2011-07-15

    A new multiresidue method has been developed and validated for the simultaneous extraction of more than two hundred pesticides, including non-polar and polar pesticides (carbamates, organochlorine, organophosphorous, pyrethroids, herbicides and insecticides) in urine at trace levels by gas and ultra high pressure liquid chromatography coupled to ion trap and triple quadrupole mass spectrometry, respectively (GC-IT-MS/MS, UHPLC-QqQ-MS/MS). Non-polar and polar pesticides were simultaneously extracted from urine samples by a simple and fast solid phase extraction (SPE) procedure using C(18) cartridges as sorbent, and dichloromethane as elution solvent. Recovery was in the range of 60-120%. Precision values expressed as relative standard deviation (RSD) were lower than 25%. Identification and confirmation of the compounds were performed by the use of retention time windows, comparison of spectra (GC-amenable compounds) or the estimation of the ion ratio (LC-amenable compounds). For GC-amenable pesticides, limits of detection (LODs) ranged from 0.001 to 0.436 μg L(-1) and limits of quantification (LOQs) from 0.003 to 1.452 μg L(-1). For LC-amenable pesticides, LODs ranged from 0.003 to 1.048 μg L(-1) and LOQs ranged from 0.011 to 3.494 μg L(-1). Finally, the optimized method was applied to the analysis of fourteen real samples of infants from agricultural population. Some pesticides such as methoxyfenozide, tebufenozide, piperonyl butoxide and propoxur were found at concentrations ranged from 1.61 to 24.4 μg L(-1), whereas methiocarb sulfoxide was detected at trace levels in two samples.

  12. Validation of a multi-residue method to determine deltamethrin and alpha-cypermethrin in mosquito nets by gas chromatography with electron capture detection (GC-μECD)

    PubMed Central

    2013-01-01

    Background Nowadays long-lasting insecticidal mosquito nets (LNs) are frequently used around the world to protect people against malaria vectors. As they contain insecticide, laboratory control is needed to check whether the content of the active ingredient follows the conditions of the manufacturer and also if the active ingredient is still present after some time of use. For this purpose, an analytical method had to be developed. The fact that LNs include a range of polymers for the yarn and use coated or incorporated technologies for the active ingredient, it is a challenge to find only one analytical method determining the active ingredient in LNs, which takes into account both impregnation technologies. Some methods are provided by international organizations but are limited by the determination of only one pesticide per method. The aim of this study was to optimize a short time extraction method for deltamethrin and alpha-cypermethrin from coated and incorporated mosquito nets and also to detect both insecticides in one analytical run, using gas chromatography with electron capture detection (GC-μECD). Methods Based on the literature, the most suitable solvent and the adequate extraction process for the insecticides used for net making were identified and adapted for the new multi-residue method. Results The validation data of the multi-residue method to determine deltamethrin and alpha-cypermethrin in mosquito nets by GC-μECD are given. Depending on the concentration of the active ingredient spiked on the nets, the mean recovery for alpha-cypermethrin ranged between 86% and 107% with a relative standard deviation below 3.5%. For deltamethrin it ranged between 90% and 108% with a relative standard deviation also below 3.5%. The limit of detection is 0.009 g.a.i/kg of net (0.3 mg a.i./m2 of net) both for alpha-cypermethrin and deltamethrin. Conclusions Data obtained are excellent. A 30 minutes reflux extraction method with xylene was developed to determine

  13. Chromatography in Industry

    NASA Astrophysics Data System (ADS)

    Schoenmakers, Peter

    2009-07-01

    This review focuses on the chromatography research that has been carried out within industry or in close cooperation with industry and that has been reported in the scientific literature between 2006 and mid-2008. Companies in the health care sector, such as pharmaceutical and biotechnology companies, are the largest contributors. Industrial research seems to take place in an open environment in cooperation with academia, peer companies, and institutions. Industry appears ready to embrace new technologies as they emerge, but they focus strongly on making chromatography work robustly, reliably, rapidly, and automatically. “Hyphenated” systems that incorporate on-line sample-preparation techniques and mass-spectrometric detection are the rule rather than the exception. Various multidimensional separation methods are finding numerous applications. Strategies aimed at speeding up the development of new chromatographic methods remain the focus of attention. Also, there is a clear trend toward exploring chromatographic methods for parallel processing along with other strategies for high-throughput analysis.

  14. Sensitive high-performance liquid chromatography method for the simultaneous determination of low levels of dichloroacetic acid and its metabolites in blood and urine.

    PubMed

    Narayanan, L; Moghaddam, A P; Taylor, A G; Sudberry, G L; Fisher, J W

    1999-06-11

    Dichloroacetic acid (DCA) is a contaminant found in treated drinking water due to chlorination. DCA has been shown to be a complete hepatocarcinogen in both mice and rats. In this study we developed a rapid and sensitive high-performance liquid chromatography (HPLC) method to simultaneously detect DCA and its metabolites, oxalic acid, glyoxylic acid and glycolic acid in blood and urine samples of animals sub-chronically administered with DCA (2 g/l) in drinking water. Both urine and plasma samples were treated minimally before HPLC analysis. Separation and detection of DCA and its metabolites were achieved using an anion-exchange column and a conductivity detector. The mobile phase consisted of an initial concentration of 0.01 mM sodium hydroxide in 40% methanol followed by a linear gradient from 0.01 mM to 60 mM sodium hydroxide in 40% methanol for 30 min. The lower detection limit for DCA and each of its three major metabolites was 0.05 microg/ml. DCA and its metabolites gave a linear response range from 0.05 to 100 microg/ml. Plasma DCA was also analyzed by gas chromatography (GC), and the results obtained correlated with those from the HPLC method (correlation coefficient=0.999). While available HPLC techniques offer sensitive procedures to detect either glycolic acid or oxalic acid, the described HPLC method has the unique advantage of determining simultaneously the parent compound (DCA) and its three major metabolites (oxalic acid, glyoxylic acid and glycolic acid) in biological samples, without complex sample preparation.

  15. Development of a supercritical fluid chromatography-tandem mass spectrometry method for the determination of azacitidine in rat plasma and its application to a bioavailability study.

    PubMed

    Li, Dongpo; Zhang, Tianhong; Kou, Longfa; Zhang, Youxi; Sun, Jin; He, Zhonggui

    2013-12-27

    Azacitidine is widely used for the treatment of myelodysplastic syndromes (MDS) and acute myelogenous leukaemia (AML). The analysis of azacitidine in biological samples is subject to interference by endogenous compounds. Previously reported high-performance liquid chromatography/tandem mass spectrometric (HPLC-MS/MS) bioanalytical assays for azacitidine suffer from expensive sample preparation procedures or from long separation times to achieve the required selectivity. Herein, supercritical fluid chromatography with tandem mass spectrometry (SFC-MS/MS) was explored as a more promising technique for the selective analysis of structure-like or chiral drugs in biological matrices. In this study, a simple, rapid and specific SFC/MS/MS analytical method was developed for the determination of azacitidine levels in rat plasma. Azacitidine was completely separated from the endogenous compounds on an ACQUITY UPLC™ BEH C18 column (100 mm×3.0 mm, 1.7 μm; Waters Corp., Milford, MA, USA) using isocratic elution with CO2/methanol as the mobile phase. The single-run analysis time was as short as 3.5 min. The sample preparation for protein removal was accomplished using a simple methanol precipitation method. The lower limit of quantification (LLOQ) of azacitidine was 20 ng/mL. The intra-day and inter-day precisions were less than 15%, and the relative error (RE) was within ±15% for the medium- and high-concentration quality control (QC) samples and within ±20% for the low-concentration QC samples. Finally, the developed method was successfully applied to a pharmacokinetic study in rats following the intravenous administration of azacitidine.

  16. Determination of low-molecular-weight organic acids in non-small cell lung cancer with a new liquid chromatography-tandem mass spectrometry method.

    PubMed

    Klupczynska, Agnieszka; Plewa, Szymon; Dyszkiewicz, Wojciech; Kasprzyk, Mariusz; Sytek, Natalia; Kokot, Zenon J

    2016-09-10

    As compared to other classes of metabolites, determination of organic acids is an underrepresented field in cancer research and till now there has been a lack of appropriate analytical procedure for determination of serum levels of organic acids potentially associated with cancer development. The aim of the study was to develop a new rapid liquid chromatography-tandem mass spectrometry method for the quantification of six low-molecular-weight organic acids in human serum and to apply this method in an analysis of samples collected from non-small cell lung cancer (NSCLC) patients and a matched control group. The samples were prepared by solid phase extraction (Clean-up CUQAX, UCT). Chromatography was conducted on a Synergi Hydro-RP column (Phenomenex) and a gradient run of 15min. Detection was performed using a negative multiple reaction monitoring mode. The calibration ranges were as follows: 0.24-38.42μmol/L for 2-hydroxybutyric acid, 0.09-17.23μmol/L for fumaric acid, 0.08-15.13μmol/L for glutaric acid, 0.11-2.22mmol/L for lactic acid, 0.39-30.98μmol/L for pyroglutamic acid, and 0.08-16.93μmol/L for succinic acid. Mean relative recovery range was 85.99-114.42% and the determined intra- and inter day coefficients of variation were ≤14%. Among the studied acids, pyroglutamic acid showed the best discriminating potential and enabled to identify accurately NSCLC patients and control subjects regardless of the cancer stage. Further investigations of serum organic acids may allow us to better understand the underlying mechanisms involved in NSCLC and develop novel means of its detection and treatment. The developed method may be also a valuable tool to study metabolic changes associated with other types of cancer.

  17. Determination of drugs in surface water and wastewater samples by liquid chromatography-mass spectrometry: Methods and preliminary results including toxicity studies with Vibrio fischeri

    USGS Publications Warehouse

    Farre, M.; Ferrer, I.; Ginebreda, A.; Figueras, M.; Olivella, L.; Tirapu, L.; Vilanova, M.; Barcelo, D.

    2001-01-01

    In the present work a combined analytical method involving toxicity and liquid chromatography-electrospray ionization-mass spectrometry (LC-ESI-MS) was developed for the determination of pharmaceutical compounds in water samples. The drugs investigated were the analgesics: ibuprofen, ketoprofen, naproxen, and diclofenac, the decomposition product of the acetyl salicylic acid: salicylic acid and one lipid lowering agent, gemfibrozil. The selected compounds are acidic substances, very polar and all of them are analgesic compounds that can be purchased without medical prescription. The developed protocol consisted, first of all, on the use Microtox?? and ToxAlert??100 toxicity tests with Vibrio fischeri for the different pharmaceutical drugs. The 50% effective concentration (EC50) values and the toxicity units (TU) were determined for every compound using both systems. Sample enrichment of water samples was achieved by solid-phase extraction procedure (SPE), using the Merck LiChrolut?? EN cartridges followed by LC-ESI-MS. Average recoveries loading 1 l of samples with pH=2 varied from 69 to 91% and the detection limits in the range of 15-56 ng/l. The developed method was applied to real samples from wastewater and surface-river waters of Catalonia (north-east of Spain). One batch of samples was analyzed in parallel also by High Resolution Gas Chromatography coupled with Mass Spectrometry (HRGC-MS) and the results have been compared with the LC-ESI-MS method developed in this work. ?? 2001 Elsevier Science B.V. All rights reserved.

  18. Determination and kinetics of enrofloxacin and ciprofloxacin in Tra catfish (Pangasianodon hypophthalmus) and giant freshwater prawn (Macrobrachium rosenbergii) using a liquid chromatography/mass spectrometry method.

    PubMed

    Danyi, S; Widart, J; Douny, C; Dang, P K; Baiwir, D; Wang, N; Tu, H T; Tung, V T; Phuong, N-T; Kestemont, P; Scippo, M-L

    2011-04-01

    Determination and kinetics of enrofloxacin and ciprofloxacin in Tra catfish (Pangasianodon hypophthalmus) and giant freshwater prawn (Macrobrachium rosenbergii) using a liquid chromatography/mass spectrometry method. J. vet. Pharmacol. Therap. 34, 142-152. The fluoroquinolones enrofloxacin (EF) and ciprofloxacin (CF) residues were investigated in the edible tissues of two important Asian aquacultured species such as Tra catfish (Pangasianodon hypophthalmus) and giant freshwater prawn (Macrobrachium rosenbergii) using a sensitive liquid chromatography-electrospray ionization-tandem mass spectrometry method. Fish and prawn were treated with medicated feed with multiple doses of EF, in field conditions. A validation study of the analytical method was realized in terms of linearity, specificity, precision (repeatability and within-laboratory reproducibility), recovery and decision limit (CCα). The time needed before the antibiotic disappears from animal tissues or reach the maximum residue limit (MRL, 100μg/kg) was assessed. The concentration values of EF detected in Tra catfish tissue were between the MRL and 2×MRL concentrations, according to the fish density, 7days following the end of the enrofloxacin treatment (20mg/kg body weight per day, for seven consecutive days). The concentration value of ER in prawn tissue was lower than the MRL and the limit of quantification (LOQ, 14μg/kg) 5 and 7days after the stop of the EF treatment (50mg/kg body weight per day, for five consecutive days), respectively. The mean detected levels of CF was much lower in comparison with that of EF, indicating that only a small part of EF is metabolized into CF (<5%) in both Tra catfish and prawn.

  19. Standardisation of Gymnema sylvestre R.Br. by high-performance thin-layer chromatography: an improved method.

    PubMed

    Raju, Valivarthi S R; Kannababu, S; Subbaraju, Gottumukkala V

    2006-01-01

    An improved high-performance thin-layer chromatographic (HPTLC) method for the standardisation of Gymnema sylvestre is reported. The method involves the initial hydrolysis of gymnemic acids, the active ingredients, to a common aglycone followed by the quantitative estimation of gymnemagenin. The present method rectifies an error found in an HPTLC method reported recently.

  20. Development of a perfusion ion-exchange chromatography method for the separation of soybean proteins and its application to cultivar characterization.

    PubMed

    Heras, J M; Marina, M L; García, M C

    2007-06-15

    A perfusion ion-exchange chromatography method has been designed, for the first time, for the separation of soybean proteins and its application to the characterization of soybean cultivars. For that purpose, the gradient, the mobile phase composition (buffer concentration, buffer pH, and elution salt), and the temperature were optimized. The method consisted of a two-step gradient (0% B for 2 min and from 0 to 50% B in 10 min) being mobile phase A a 2 0mM borate buffer (pH 9) and mobile phase B a 20 mM borate buffer (pH 9) containing 1M sodium chloride. The procedure used for the preparation of sample solutions was significantly simpler than that proposed by other authors and basically consisted of dissolving in water. This method enabled the separation of soybean proteins from a soybean protein isolate in 11 peaks in about 9 min. The method was used to separate soybean proteins in different commercial soybeans. In general, the 11 peaks yielded by the soybean protein isolate were also observed in the chromatograms of all soybeans. However, the area percentages of every peak in every soybean enabled the differentiation between soybeans. Moreover, the method was also used to separate soybean proteins in the proteic fractions obtained from every soybean. Multivariate methods were used for patterns recognition and the classification of samples.

  1. Ion pair-based dispersive liquid-liquid microextraction followed by high performance liquid chromatography as a new method for determining five folate derivatives in foodstuffs.

    PubMed

    Nojavan, Yones; Kamankesh, Marzieh; Shahraz, Farzaneh; Hashemi, Maryam; Mohammadi, Abdorreza

    2015-05-01

    A novel technique for simultaneous determination of five folate derivatives in various food matrices was developed by ion pair-based dispersive liquid-liquid microextraction (IP-DLLME) combined with high-performance liquid chromatography (HPLC). In the proposed method, N-methyl-N,N-dioctyloctan-1-ammonium chloride (aliquat-336) was used as an ion-pair reagent. Effective variables of microextraction process were optimized. Under optimum conditions, the method yielded a linear calibration curve ranging from 1-200 ng g(-1) with correlation coefficients (r(2)) higher than 0.98. The relative standard deviation for the seven analyses was 5.2-7.4%. Enrichment factors for the five folates ranged between 108-135. Limits of detection were 2-4.1 ng g(-1). A comparison of this method with other methods described that the new proposed method is rapid and accurate, and gives very good enrichment factors and detection limits for determining five folate derivatives. The newly developed method was successfully applied for the determination of five folate derivatives in wheat flour, egg yolk and orange juice samples.

  2. A versatile, stability-indicating and high-throughput ultra-fast liquid chromatography method for the determination of isoflavone aglycones in soybeans, topical formulations, and permeation assays.

    PubMed

    Nemitz, Marina C; Yatsu, Francini K J; Bidone, Juliana; Koester, Letícia S; Bassani, Valquiria L; Garcia, Cássia V; Mendez, Andreas S L; von Poser, Gilsane L; Teixeira, Helder F

    2015-03-01

    There is a growing interest in the pharmaceutical field concerning isoflavones topical delivery systems, especially with regard to their skin care properties and antiherpetic activity. In this context, the present work describes an ultra-fast liquid chromatography method (UFLC) for determining daidzein, glycitein, and genistein in different matrices during the development of topical systems containing isoflavone aglycones (IA) obtained from soybeans. The method showed to be specific, precise, accurate, and linear (0.1 to 5 µg mL(-1)) for IA determination in soybean acid extract, IA-rich fraction obtained after the purification process, IA loaded-nanoemulsions, and topical hydrogel, as well as for permeation/retention assays in porcine skin and porcine esophageal mucosa. The matrix effect was determined for all complex matrices, demonstrating low effect during the analysis. The stability indicating UFLC method was verified by submitting IA to acidic, alkaline, oxidative, and thermal stress conditions, and no interference of degradation products was detected during analysis. Mass spectrometry was performed to show the main compounds produced after acid hydrolysis of soybeans, as well as suggest the main degradation products formed after stress conditions. Besides the IA, hydroxymethylfurfural and ethoxymethylfurfural were produced and identified after acid hydrolysis of the soybean extract and well separated by the UFLC method. The method's robustness was confirmed using the Plackett-Burman experimental design. Therefore, the new method affords fast IA analysis during routine processes, extract purification, products development, and bioanalytical assays.

  3. Selective and sentivive method based on capillary liquid chromatography with in-tube solid phase microextraction for determination of monochloramine in water.

    PubMed

    Pla-Tolós, J; Moliner-Martínez, Y; Molins-Legua, C; Herráez-Hernández, R; Verdú-Andrés, J; Campíns-Falcó, P

    2015-04-03

    Due to the difficulties of working with chloramines, a critical examination of monochloramine standard preparation has been performed in order to select the best synthesis conditions. The analyte has been determined by in-tube solid phase extraction coupled to capillary liquid chromatography with UV detection (IT-SPME Capillary LC DAD). Potential factors affecting the response of monochloramine such as the pH of mobile phase and the volume of sample processed by IT-SPME Capillary LC DAD have been investigated and optimized. According to the results of the study, 0.1 mL or 4.0 mL of sample at neutral pH were loaded in the chromatographic system. A sensitive and selective method has been developed for the determination of monochloramine in water. Validation of the method has been performed. The linear range was 0.09-5mg/L with linear regression coefficients (R(2)) greater than 0.995. Method reproducibility expressed as relative standard deviation (RSD, %), was lower than 15%. The limits of detection (LODs) were 0.029 and 0.01 mg/L by processing 0.100mL or 4 mL of the samples, respectively, being below the maximum residues levels allowed for this compound. The sensitivity achieved by the developed method was better than that obtained by the reference method. The developed method was applied to water samples (tap and swimming pool water).

  4. Development and validation of a reversed-phase high-performance liquid chromatography method for routine identification and purity assessment of high-purity steviol glycoside sweeteners.

    PubMed

    Bililign, Tsion; Moore, Jeffrey C; Tan, Shane; Leeks, Allan T

    2014-02-12

    The widespread application of stevia-based sweeteners in food products has resulted in the need for reliable analytical methods for measuring the purity and identity of high-purity steviol glycoside ingredients. The objective of this research was to develop and validate a new reversed-phase separation method capable of separating and quantifying nine steviol glycosides present in typical high-purity stevia extract ingredients. Results of the study established the linearity of the method at a correlation factor of 1.000 for the two major components and other minor components of this food ingredient. Method accuracy values were in the range of 99.1-100.9%. The percent relative standard deviation for six independent assay determinations was 1.0%. The method was determined to be robust for minor changes in column temperature, initial acetonitrile content, flow rate, and wavelength. The validated high-performance liquid chromatography method was found to be suitable to be included by USP as a Food Chemicals Codex compendial standard for steviol glycosides.

  5. Quality-by-design-based ultra high performance liquid chromatography related substances method development by establishing the proficient design space for sumatriptan and naproxen combination.

    PubMed

    Patel, Prinesh N; Karakam, Vijaya Saradhi; Samanthula, Gananadhamu; Ragampeta, Srinivas

    2015-10-01

    Quality-by-design-based methods hold greater level of confidence for variations and greater success in method transfer. A quality-by-design-based ultra high performance liquid chromatography method was developed for the simultaneous assay of sumatriptan and naproxen along with their related substances. The first screening was performed by fractional factorial design comprising 44 experiments for reversed-phase stationary phases, pH, and organic modifiers. The results of screening design experiments suggested phenyl hexyl column and acetonitrile were the best combination. The method was further optimized for flow rate, temperature, and gradient time by experimental design of 20 experiments and the knowledge space was generated for effect of variable on response (number of peaks ≥ 1.50 - resolution). Proficient design space was generated from knowledge space by applying Monte Carlo simulation to successfully integrate quantitative robustness metrics during optimization stage itself. The final method provided the robust performance which was verified and validated. Final conditions comprised Waters® Acquity phenyl hexyl column with gradient elution using ammonium acetate (pH 4.12, 0.02 M) buffer and acetonitrile at 0.355 mL/min flow rate and 30°C. The developed method separates all 13 analytes within a 15 min run time with fewer experiments compared to the traditional quality-by-testing approach.

  6. Validation of an ultra-performance liquid chromatography-tandem mass spectrometry method for the determination of flecainide in human plasma and its clinical application.

    PubMed

    Mano, Yuji; Asakawa, Yoshiki; Kita, Kenji; Ishii, Takuho; Hotta, Koichiro; Kusano, Kazutomi

    2015-09-01

    A simple and reproducible bioanalytical method for the determination of flecainide in human plasma was developed and validated using an ultra-performance liquid chromatography with tandem mass spectrometry (UPLC-MS/MS) to obtain higher sensitivity than the current available methods. After simple protein precipitation, flecainide and a stable isotope-labeled internal standard (IS) were chromatographed on an Acquity UPLC BEH C18 column (2.1 × 100 mm, 1.7 µm) with isocratic elution of mobile phase consisting of 45% methanol containing 0.1% formic acid at a flow rate 0.25 mL/min. Detection was performed in positive electrospray ionization by monitoring the selected ion transitions at m/z 415.4/301.1 for flecainide and m/z 419.4/305.1 for the IS. The method was validated according to current bioanalytical method validation guidelines. The calibration standard curve was linear from 2.5 to 1000 ng/mL using 0.1 mL of plasma. No significant interferences were detected in blank human plasma. Accuracy and precision in the intra- and inter-batch reproducibility study were within acceptance criteria. Neither hemolysis effects nor matrix effects were observed. The UPLC-MS/MS method developed was successfully applied to determine plasma flecainide concentrations to support clinical studies and incurred sample reanalysis also ensured the reproducibility of the method.

  7. Development and validation of a fast high pressure liquid chromatography method for the analysis of lignocellulosic biomass hydrolysis and fermentation products.

    PubMed

    Scarlata, Christopher J; Hyman, Deborah A

    2010-04-02

    A simple, precise, and accurate 10-min high pressure liquid chromatography (HPLC) method was developed and validated for the analysis of organic acids, alcohols, and furans from processing biomass into renewable fuels. The method uses an H(+) form cation-exchange resin stationary phase that has a five-fold shorter analysis time versus that in the traditional method. The new method was used for the analysis of acetic acid, ethanol, 5-hydroxymethyl furfural, and furfural. Results were compared with a legacy method that has historically has been used to analyze the same compounds but with a 55 min run time. Linearity was acceptable on the new method with r(2)>0.999 for all compounds using refractive index detection. Limits of detection were between 0.003 and 0.03 g/L and limits of quantification were between 0.1 and 0.01 g/L. The relative standard deviations for precision were less than 0.4% and recoveries ranged from 92% to 114% for all compounds.

  8. Comparison of three development approaches for Stationary Phase Optimised Selectivity Liquid Chromatography based screening methods Part II: A group of structural analogues (PDE-5 inhibitors in food supplements).

    PubMed

    Deconinck, E; Ghijs, L; Kamugisha, A; Courselle, P

    2016-02-01

    Three approaches for the development of a screening method to detect adulterated dietary supplements, based on Stationary Phase Optimised Selectivity Liquid Chromatography were compared for their easiness/speed of development and the performance of the optimal method obtained. This comparison was performed for a heterogeneous group of molecules, i.e. slimming agents (Part I) and a group of structural analogues, i.e. PDE-5 inhibitors (Part II). The first approach makes use of primary runs at one isocratic level, the second of primary runs in gradient mode and the third of primary runs at three isocratic levels to calculate the optimal combination of segments of stationary phases. In each approach the selection of the stationary phase was followed by a gradient optimisation. For the PDE-5 inhibitors, the group of structural analogues, only the method obtained with the third approach was able to differentiate between all the molecules in the development set. Although not all molecules are baseline separated, the method allows the identification of the selected adulterants in dietary supplements using only diode array detection. Though, due to the mobile phases used, the method could also be coupled to mass spectrometry. The method was validated for its selectivity following the guidelines as described for the screening of pesticide residues and residues of veterinary medicines in food.

  9. The Verification of the Usefulness of Electronic Nose Based on Ultra-Fast Gas Chromatography and Four Different Chemometric Methods for Rapid Analysis of Spirit Beverages

    PubMed Central

    Śliwińska, Magdalena; Namieśnik, Jacek; Wardencki, Waldemar; Dymerski, Tomasz

    2016-01-01

    Spirit beverages are a diverse group of foodstuffs. They are very often counterfeited which cause the appearance of low quality products or wrongly labelled products on the market. It is important to find a proper quality control and botanical origin method enabling the same time preliminary check of the composition of investigated samples, which was the main goal of this work. For this purpose, the usefulness of electronic nose based on ultra-fast gas chromatography (fast GC e-nose) was verified. A set of 24 samples of raw spirits, 33 samples of vodkas, and 8 samples of whisky were analysed by fast GC e-nose. Four data analysis methods were used. The PCA was applied for the visualization of dataset, observation of the variation inside groups of samples, and selection of variables for the other three statistical methods. The SQC method was utilized to compare the quality of the samples. Both the DFA and SIMCA data analysis methods were used for discrimination of vodka, whisky, and spirits samples. The fast GC e-nose combined with four statistical methods can be used for rapid discrimination of raw spirits, vodkas, and whisky and in the same for preliminary determination of the composition of investigated samples. PMID:27446633

  10. Method using gas chromatography to determine the molar flow balance for proton exchange membrane fuel cells exposed to impurities

    NASA Astrophysics Data System (ADS)

    Bender, G.; Angelo, M.; Bethune, K.; Dorn, S.; Thampan, T.; Rocheleau, R.

    An understanding of the potentially serious performance degradation effects that trace level contaminants can cause in proton exchange membrane fuel cells (PEMFCs) is crucial for the successful deployment of PEMFC for commercial applications. An experimental and analytic methodology is described that employs gas chromatography (GC) to accurately determine the concentration of impurity species in the fuel and oxidant streams of a PEMFC. In this paper we further show that the accurate determination of the contaminant concentrations at the anode and cathode inlets and outlets provides a means to quantify reactions of contaminants within the cell and to identify diffusive mass transport across the membrane. High data accuracy down to sub-ppm contaminant levels is required and was achieved by addressing several challenges pertaining to experimental setup and data analysis which are both discussed in detail. The application of the methodology is demonstrated using carbon monoxide and toluene which were injected into the cell at concentrations between 1 and 10 ppm and 20 and 60 ppm, respectively. Both impurities were observed to react in the fuel cell: carbon monoxide to carbon dioxide, and toluene to methylcyclohexane. For both contaminants closure of the molar flow balances to within 3% was achieved even at the low contaminant concentrations. This allowed the extent of both reactions at the applied operating conditions to be quantified. The presented methodology is shown to be a valuable tool for investigating the effects and reactions of trace contaminants in fuel cells and for providing critical insights into the mechanisms responsible for the associated performance degradation.

  11. New Method for Development of Carbon Coated Silica Phases for Liquid Chromatography Part I. Preparation of Carbon Phases

    PubMed Central

    Paek, Changyub; McCormick, Alon V.; Carr, Peter W.

    2011-01-01

    Owing to its combination of unique selectivity and mechanical strength, commercial carbon clad zirconia (C/ZrO2) has been widely used for many applications, including fast two-dimensional liquid chromatography (2DLC). However, the low surface area available (only 20 - 30 m2/g for commercial porous ZrO2) limits its retentivity. We have recently addressed this limitation by developing a carbon phase coated on the high surface area of HPLC grade alumina (C/Al2O3). This material provides higher retentivity and comparable selectivity, but its use is still limited by how few HPLC quality types of alumina particles (e.g., particle size, surface area, pore size) are available. In this work, we have developed useful carbon phases on silica particles, which are available in various particle sizes, pore sizes and forms of HPLC grade. To make the carbon phase on silica, we first treat the silica surface with a monolayer or less of metal cations that bind to deprotonated silanols to provide catalytic sites for carbon deposition. After Al (III) treatment, a carbon phase is formed on the silica surface by chemical vapor deposition at 700 °C using hexane as the carbon source. The amount of Al (III) on the surface was varied to assess its effect on carbon deposition, and the carbon loading was varied at different Al (III) levels to assess its effect on the chromatographic properties of the various carbon adsorbents. We observed that use of a concentration of Al (III) corresponding to a full monolayer leads to the most uniform carbon coating. A carbon coating sufficient to cover all the Al (III) sites, required about 4 – 5 monolayers in this work, provided the best chromatographic performance. The resulting carbon phases behave as reversed phases with reasonable efficiency (50,000 – 79,000 plates/meter) for non-aromatic test species. PMID:21295308

  12. Development, optimization, validation and application of faster gas chromatography - flame ionization detector method for the analysis of total petroleum hydrocarbons in contaminated soils.

    PubMed

    Zubair, Abdulrazaq; Pappoe, Michael; James, Lesley A; Hawboldt, Kelly

    2015-12-18

    This paper presents an important new approach to improving the timeliness of Total Petroleum Hydrocarbon (TPH) analysis in the soil by Gas Chromatography - Flame Ionization Detector (GC-FID) using the CCME Canada-Wide Standard reference method. The Canada-Wide Standard (CWS) method is used for the analysis of petroleum hydrocarbon compounds across Canada. However, inter-laboratory application of this method for the analysis of TPH in the soil has often shown considerable variability in the results. This could be due, in part, to the different gas chromatography (GC) conditions, other steps involved in the method, as well as the soil properties. In addition, there are differences in the interpretation of the GC results, which impacts the determination of the effectiveness of remediation at hydrocarbon-contaminated sites. In this work, multivariate experimental design approach was used to develop and validate the analytical method for a faster quantitative analysis of TPH in (contaminated) soil. A fractional factorial design (fFD) was used to screen six factors to identify the most significant factors impacting the analysis. These factors included: injection volume (μL), injection temperature (°C), oven program (°C/min), detector temperature (°C), carrier gas flow rate (mL/min) and solvent ratio (v/v hexane/dichloromethane). The most important factors (carrier gas flow rate and oven program) were then optimized using a central composite response surface design. Robustness testing and validation of model compares favourably with the experimental results with percentage difference of 2.78% for the analysis time. This research successfully reduced the method's standard analytical time from 20 to 8min with all the carbon fractions eluting. The method was successfully applied for fast TPH analysis of Bunker C oil contaminated soil. A reduced analytical time would offer many benefits including an improved laboratory reporting times, and overall improved clean up

  13. High-performance Thin-layer Chromatography Method Development, Validation, and Simultaneous Quantification of Four Compounds Identified in Standardized Extracts of Orthosiphon stamineus

    PubMed Central

    Hashim, Suzana; Beh, Hooi Kheng; Hamil, Mohamad Shahrul Ridzuan; Ismail, Zhari; Majid, Amin Malik Shah Abdul

    2016-01-01

    Context: Orthosiphon stamineus is a medicinal herb widely grown in Southeast Asia and tropical countries. It has been used traditionally as a diuretic, abdominal pain, kidney and bladder inflammation, gout, and hypertension. Aims: This study aims to develop and validate the high-performance thin layer chromatography (HPTLC) method for quantification of rosmarinic acid (RA), 3'-hydroxy-5,6,7,4'-tetramethoxyflavone (TMF), sinensitin (SIN) and eupatorin (EUP) found in ethanol, 50% ethanol and water extract of O. stamineus leaves. Materials and Methods: HPTLC method was conducted using an HPTLC system with a developed mobile phase system of toluene: ethyl acetate: formic acid (3:7:0.1) performed on precoated silica gel 60 F254 TLC plates. The method was validated based on linearity, accuracy, precision, limit of detection, limit of quantification (LOQ), and specificity, respectively. The detection of spots was observed at ultraviolet 254 nm and 366 nm. Results: The linearity of RA, TMF, SIN, and EUP were obtained between 10 and 100 ng/spot with high correlation coefficient value (R2) of more than 0.986. The limit of detection was found to be 122.47 ± 3.95 (RA), 43.38 ± 0.79 (SIN), 17.26 ± 1.16 (TMF), and 46.80 ± 1.33 ng/spot (EUP), respectively. Whereas the LOQ was found to be 376.44 ± 6.70 (RA), 131.45 ± 2.39 (SIN), 52.30 ± 2.01 (TMF), and 141.82 ± 1.58 ng/spot (EUP), respectively. Conclusion: The proposed method showed good linearity, precision, accuracy, and high sensitivity. Hence, it may be applied in a routine quantification of RA, SIN, TMF, and EUP found in ethanol, 50% of ethanol and water extract of O. stamineus leaves. SUMMARY HPTLC method provides rapid estimation of the marker compound for routine quality control analysis.The established HPTLC method is rapid for qualitative and quantitative fingerprinting of Orthosiphon stamineus extract used for commercial product.Four identified markers (RA, SIN, EUP and TMF) found in three a different type of O

  14. Gas Chromatography

    NASA Astrophysics Data System (ADS)

    Qian, Michael C.

    Gas chromatography (GC) has many applications in the analysis of food products. GC has been used for the determination of fatty acids, triglycerides, cholesterol, gases, water, alcohols, pesticides, flavor compounds, and many more. While GC has been used for other food components such as sugars, oligosaccharides, amino acids, peptides, and vitamins, these substances are more suited to analysis by high performance liquid chromatography. GC is ideally suited to the analysis of volatile substances that are thermally stable. Substances such as pesticides and flavor compounds that meet these criteria can be isolated from a food and directly injected into the GC. For compounds that are thermally unstable, too low in volatility, or yield poor chromatographic separation due to polarity, a derivatization step must be done before GC analysis. The two parts of the experiment described here include the analysis of alcohols that requires no derivatization step, and the analysis of fatty acids which requires derivatization. The experiments specify the use of capillary columns, but the first experiment includes conditions for a packed column.

  15. A Binary-Like Approach for the Computer Assisted Method Development of Isocratic and Programmed Ternary Solvent Elutions in Reversed-Phase Liquid Chromatography

    PubMed Central

    García-Lavandeira, J.; Martinez-Pontevedra, J.A.; Cela, R.

    2012-01-01

    A specific tool to enable exploration of multisolvent isocratic and programmed elutions for the computer assisted method development of reversed-phase liquid chromatography separations is described. The tool is purposely identical to those used in the optimization of binary solvent systems, which are by far the most commonly used by chromatographers. Existing data from failed binary solvent optimization processes are reused to explore ternary solvent systems with a few additional isocratic and programmed runs. This allows the development of efficient retention models for ternary systems, although the work of the chromatographer remains identical to that for optimization of binary systems. The retention models are used to develop an unattended optimization process and finally, the chromatographer selects the most satisfactory solution for testing and implementing in routine analysis. The process is exemplified with a mixture of 12 compounds that cannot be separated satisfactorily in aqueous binary solvent systems with methanol and acetonitrile as modifiers. PMID:22291054

  16. Development and validation of a method for the quantitation of Delta9 tetrahydrocannabinol in human plasma by high performance liquid chromatography after solid-phase extraction.

    PubMed

    Abbara, Chadi; Galy, Romain; Benyamina, Amine; Reynaud, Michel; Bonhomme-Faivre, Laurence

    2006-06-07

    A high performance liquid chromatography (HPLC) procedure for the determination of Delta9 tetrahydrocannabinol (THC) in human plasma is described. A two-step solid-phase extraction on CN cartridges was coupled with a reversed phase HPLC system. THC was eluted using a mobile phase composed of methanol, acetonitrile and tetrabutylammonium perchlorate solution (0.005 M, pH 3.2), through a C18 Nucleosil column and detected at a wavelength of 215 nm. Calibration curve was linear over the range 5-100 ng/ml with a lower limit of quantification validated at 5 ng/ml. Extraction recovery using the developed extraction procedure was higher than 85%. This method is presently used for the quantification of THC in plasma samples from regular cannabis smokers.

  17. Methods for protecting silica sorbents used in high-performance liquid chromatography from strongly adsorbed impurities during purification of human recombinant insulin.

    PubMed

    Gusarov, D; Lasman, V; Bayramashvili, D

    2007-06-15

    One of the main stages of human recombinant insulin (HRI) production is the hormone purification by means of reversed phase high-performance liquid chromatography (RP HPLC). The optimization of this stage determines the increase of the total manufacturing yield. Therefore, the cost of the sorbent used in HPLC influences the cost of the manufacturing product, i.e. HRI substance. However, resolution between HRI and its admixtures decreases with time. The reason for this is the sorbent contamination with strongly adsorbed impurities (SAI) which are accumulated during elution. In the following research several methods for sorbent protection are studied. The opinion that SAI are mainly high-molecular weight compounds was examined using gel filtration. Different sorbent types were tested for the use in guard column. The results obtained were applied and improved at preparative level.

  18. Development of On-Line High Performance Liquid Chromatography (HPLC)-Biochemical Detection Methods as Tools in the Identification of Bioactives

    PubMed Central

    Malherbe, Christiaan J.; de Beer, Dalene; Joubert, Elizabeth

    2012-01-01

    Biochemical detection (BCD) methods are commonly used to screen plant extracts for specific biological activities in batch assays. Traditionally, bioactives in the most active extracts were identified through time-consuming bio-assay guided fractionation until single active compounds could be isolated. Not only are isolation procedures often tedious, but they could also lead to artifact formation. On-line coupling of BCD assays to high performance liquid chromatography (HPLC) is gaining ground as a high resolution screening technique to overcome problems associated with pre-isolation by measuring the effects of compounds post-column directly after separation. To date, several on-line HPLC-BCD assays, applied to whole plant extracts and mixtures, have been published. In this review the focus will fall on enzyme-based, receptor-based and antioxidant assays. PMID:22489144

  19. Development of on-line high performance liquid chromatography (HPLC)-biochemical detection methods as tools in the identification of bioactives.

    PubMed

    Malherbe, Christiaan J; de Beer, Dalene; Joubert, Elizabeth

    2012-01-01

    Biochemical detection (BCD) methods are commonly used to screen plant extracts for specific biological activities in batch assays. Traditionally, bioactives in the most active extracts were identified through time-consuming bio-assay guided fractionation until single active compounds could be isolated. Not only are isolation procedures often tedious, but they could also lead to artifact formation. On-line coupling of BCD assays to high performance liquid chromatography (HPLC) is gaining ground as a high resolution screening technique to overcome problems associated with pre-isolation by measuring the effects of compounds post-column directly after separation. To date, several on-line HPLC-BCD assays, applied to whole plant extracts and mixtures, have been published. In this review the focus will fall on enzyme-based, receptor-based and antioxidant assays.

  20. Validation of a method for the analysis of quinolones residues in bovine muscle by liquid chromatography with electrospray ionisation tandem mass spectrometry detection.

    PubMed

    Rubies, A; Vaquerizo, R; Centrich, F; Compañó, R; Granados, M; Prat, M D

    2007-04-15

    A liquid chromatography-tandem mass spectrometry method for the determination and confirmation of nine quinolones was optimised and validated according to Commission Decision 2002/657/EC. Analytes were extracted from veal muscle with water and extracts purified with 96-well plates Oasis HLB cartridges. Separation was carried out in a silica-based C(18) column (50mmx2.1mm) with mobile phases consisting of water/acetonitrile mixtures containing acetic acid. Linear calibration curves in the ranges 4-400 and 50-800ngg(-1), with correlation coefficients at least 0.995, were obtained for all the analytes. At concentration levels above 10ngg(-1), quantification errors were lower than 10% and repeatability and within-laboratory reproducibility standard deviations below 6% and 10%, respectively. Decision limits and detection capabilities are reported.

  1. Method validation and dissipation kinetics of four herbicides in maize and soil using QuEChERS sample preparation and liquid chromatography tandem mass spectrometry.

    PubMed

    Pang, Nannan; Wang, Tielong; Hu, Jiye

    2016-01-01

    A versatile liquid chromatography tandem mass spectrometry method with modified QuEChERS (quick, easy, cheap, effective, rugged, and safe) sample preparation was developed for the determination of rimsulfuron, mesotrione, fluroxypyr-meptyl, and fluroxypyr. By adjusting the amount of graphitized carbon black, the herbicide analytes could be quantified with satisfactory recoveries in the range of 80-110%. A dissipation kinetics study conducted under open field conditions at two sites during 2014 showed first order equations with half-lives between 0.6d and 3.6d, illustrating an appropriate degree of stability and safety. The dissipation kinetics were different in the different matrices. Although the herbicides had higher initial residues in straw than those in soil, they degraded faster in straw. The terminal residues for the herbicides formulated in two water dispersible granules were all below maximum residue limits. These results not only gave insights about the analytes but also contributed to environmental protection and food safety.

  2. Multi-residue and multi-class method for the determination of antibiotics in bovine muscle by ultra-high-performance liquid chromatography tandem mass spectrometry.

    PubMed

    Freitas, Andreia; Barbosa, Jorge; Ramos, Fernando

    2014-09-01

    A multi-residue quantitative screening method covering 41 antibiotics from 7 different families, by ultra-high-performance-liquid-chromatography tandem mass spectrometry (UHPLC-MS/MS), is described. Sulfonamides, trimethoprim, tetracyclines, macrolides, quinolones, penicillins and chloramphenicol are simultaneously detected after a simple sample preparation of bovine muscle optimized to achieve the best recovery for all compounds. A simple sample treatment was developed consisting in an extraction with a mixture of acetonitrile and ethylenediaminetetraacetic acid (EDTA), followed by a defatting step with n-hexane. The methodology was validated, in accordance with Decision 2002/657/EC by evaluating the required parameters: decision limit (CCα), detection capability (CCβ), specificity, repeatability and reproducibility. Precision in terms of relative standard deviation was under 20% for all compounds and the recoveries between 91% and 119%. CCα and CCβ were determined according the maximum residue limit (MRL) or the minimum required performance limit (MRPL), when required.

  3. New evidences on efficacy of boronic acid-based derivatization method to identify sugars in plant material by gas chromatography-mass spectrometry.

    PubMed

    Faraco, Marianna; Fico, Daniela; Pennetta, Antonio; De Benedetto, Giuseppe E

    2016-10-01

    This work presents an analytical procedure based on gas chromatography-mass spectrometry which allows the determination of aldoses (glucose, mannose, galactose, arabinose, xylose, fucose, rhamnose) and chetoses (fructose) in plant material. One peak for each target carbohydrate was obtained by using an efficient derivatization employing methylboronic acid and acetic anhydride sequentially, whereas the baseline separation of the analytes was accomplished using an ionic liquid capillary column. First, the proposed method was optimized and validated. Successively, it was applied to identify the carbohydrates present in plant material. Finally, the procedure was successfully applied to samples from a XVII century painting, thus highlighting the occurrence of starch glue and fruit tree gum as polysaccharide materials.

  4. EPA Method 524.2: Measurement of Purgeable Organic Compounds in Water by Capillary Column Gas Chromatography/Mass Spectrometry

    EPA Pesticide Factsheets

    SAM lists this method for preparation and analysis of water samples. This method determines carbon disulfide and 1,2-dichloroethane in drinking water and acrylonitrile and methyl acrylonitrile in drinking and aqueous/liquid samples.

  5. EPA Method 300.1, Revision 1.0: Determination of Inorganic Anions in Drinking Water by Ion Chromatography

    EPA Pesticide Factsheets

    SAM lists this method for preparation and analysis of aqueous liquid and drinking water samples for fluoride. This method can be adapted for analysis of solid and air samples when appropriate sample preparation techniques have been applied.

  6. Hydrophilic interaction liquid chromatography/positive ion electrospray ionization mass spectrometry method for the quantification of alprazolam and α-hydroxy-alprazolam in human plasma.

    PubMed

    Kalogria, Eleni; Pistos, Constantinos; Panderi, Irene

    2013-12-30

    A hydrophilic interaction liquid chromatography/positive ion electrospray-mass spectrometry (HILIC-ESI/MS) has been developed and fully validated for the quantification of alprazolam and its main metabolite, α-hydroxy-alprazolam, in human plasma. The assay is based on 50μL plasma samples, following liquid-liquid extraction. All analytes and the internal standard (tiamulin) were separated by hydrophilic interaction liquid chromatography using an X-Bridge-HILIC analytical column (150.0mm×2.1mm i.d., particle size 3.5μm) under isoscratic elution. The mobile phase was composed of a 7% 10mM ammonium formate water solution in acetonitrile and pumped at a flow rate of 0.20mLmin(-1). Running in positive electrospray ionization and selected ion monitoring (SIM) the mass spectrometer was set to analyze the protonated molecules [M+H](+) at m/z 309, 325 and 494 for alprazolam, α-hydroxy-alprazolam and tiamulin (ISTD) respectively. The assay was linear over the concentration range of 2.5-250ngmL(-1) for alprazolam and 2.5-50ngmL(-1) for α-hydroxy alprazolam. Intermediate precision was less than 4.1% over the tested concentration ranges. The method is the first reported application of HILIC in the analysis benzodiazepines in human plasma. With a small sample size (50μL human plasma) and a run time less than 10.0min for each sample the method can be used to support a wide range of clinical studies concerning alprazolam quantification.

  7. Dispersive liquid-liquid microextraction method based on solidification of floating organic drop combined with gas chromatography with electron-capture or mass spectrometry detection.

    PubMed

    Leong, Mei-I; Huang, Shang-Da

    2008-11-21

    A simple dispersive liquid-liquid microextraction (DLLME) method based on solidification of a floating organic drop (DLLME-SFO) technique combined with gas chromatography/electron-capture detection (GC/ECD) or gas chromatography/mass spectrometry (GC/MS) has been developed. The proposed method is simple, low in cost, and of high precision. It overcomes the most important problem in DLLME, the high-toxic solvent used. Halogenated organic compounds (HOCs) in water samples were determined as the model compounds. The parameters optimized for the DLLME-SFO technique were as follows: A mixture of 0.5 mL acetone, containing 10 microL 2-dodecanol (2-DD-OH), was rapidly injected by syringe into the 5 mL water sample. After centrifugation, the fine 2-DD-OH droplets (8+/-0.5 microL) were floated at the top of the screwcap test tube. The test tube was then cooled in an ice bath. After 5 min the 2-DD-OH solvent had solidified and was then transferred into a conical vial; it melted quickly at room temperature and 3 microL (for GC/ECD) or 2 microL (for GC/MS) of it was injected into a gas chromatograph for analysis. The limit of detection (LOD) for this technique was 0.005-0.05microgL(-1) for GC/ECD and was 0.005-0.047 microgL(-1) for GC/MS, respectively. The linear range of the calibration curve of DLLME-SFO was from 0.01 to 500 microgL(-1) with a coefficient of estimation (r2)>0.996 for GC/ECD and was from 0.02 to 500 microgL(-1) with a coefficient of estimation (r2)>0.996 for GC/MS.

  8. Universal method to determine acidic licit and illicit drugs and personal care products in water by liquid chromatography quadrupole time-of-flight.

    PubMed

    Andrés-Costa, María Jesús; Carmona, Eric; Picó, Yolanda

    2016-01-01

    Pharmaceuticals, illicit drugs and personal care products are emerging contaminants widely distributed in water. Currently, a number of solid-phase extraction (SPE) procedures followed by liquid chromatography tandem mass spectrometry (LC-MS/MS) have been reported. However, target analysis of selected compounds is commonly used whereas other related contaminants present in the sample remain invisible. Carmona et al. [1] described a method for determining 21 emerging contaminants by LC-MS/MS with improved mobile phases. We tested this protocol in combination with high resolution mass spectrometry using a quadrupole time-of-flight (QqTOF) instrument to get a wide non-target screening approach in order to have a broader scope and more practical method for detecting licit and illicit drugs and personal care products than traditional target methods. The essential points in the method are: •The screening capabilities of QqTOF (ABSciex Triple TOF™) are used for detecting and identifying non-target pharmaceuticals and a large number of other emerging contaminants in water.•The quantitative features of the instrument, the Achilles heel of the QqTOF mass spectrometers, are established for few selected compounds.•The method may be applied to identify a large number of emerging contaminants in water. However, pre-validation will be needed to quantify them.

  9. Development of a liquid chromatography-electrospray ionization-tandem mass spectrometry method for the simultaneous analysis of intact glucosinolates and isothiocyanates in Brassicaceae seeds and functional foods.

    PubMed

    Franco, P; Spinozzi, S; Pagnotta, E; Lazzeri, L; Ugolini, L; Camborata, C; Roda, A

    2016-01-08

    A new high pressure liquid chromatography-electrospray ionization-tandem mass spectrometry method for the simultaneous determination of glucosinolates, as glucoraphanin and glucoerucin, and the corresponding isothiocyanates, as sulforaphane and erucin, was developed and applied to quantify these compounds in Eruca sativa defatted seed meals and enriched functional foods. The method involved solvent extraction, separation was achieved in gradient mode using water with 0.5% formic acid and acetonitrile with 0.5% formic acid and using a reverse phase C18 column. The electrospray ion source operated in negative and positive mode for the detection of glucosinolates and isothiocyanates, respectively, and the multiple reaction monitoring (MRM) was selected as acquisition mode. The method was validated following the ICH guidelines. Replicate experiments demonstrated a good accuracy (bias%<10%) and precision (CV%<10%). Detection limits and quantification limits are in the range of 1-400ng/mL for each analytes. Calibration curves were validated on concentration ranges from 0.05 to 50μg/mL. The method proved to be suitable for glucosinolates and isothiocyanates determination both in biomasses and in complex matrices such as food products enriched with glucosinolates, or nutraceutical bakery products. In addition, the developed method was applied to the simultaneous determination of glucosinolates and isothiocyanates in bakery product enriched with glucosinolates, to evaluate their thermal stability after different industrial processes from cultivation phases to consumer processing.