Simple and rapid characterization of soybean cultivars by perfusion reversed-phase HPLC: application to the estimation of the 11S and 7S globulin contents

A perfusion RP HPLC method enabling the separation of soybean proteins in an analysis time lower than 3 min has been used to obtain the chromatographic profiles of different soybean cultivars. The chromatograms obtained for each soybean variety presented clear differences that justified the potential use of this method for cultivar characterization. The area percentages obtained were employed as variables for cluster and principal components analysis of these soybeans. The application of these multivariate methods enabled the grouping of the soybeans in different categories. The protein fractions obtained from these soybeans by the application of a fractionation method were also analyzed. The chromatographic profiles obtained enabled the assignment of peaks to the main soybean proteins (7S and 11S globulins). These data were used for the estimation, for the first time, of the 7S and 11S globulin contents in soybean cultivars.     

Micro-scale extraction procedure for standardized screening of fungal metabolite production in cultures

A simple and rapid standardized micro-scale extraction procedure has been developed to prepare extracts from fungal cultures for high-performance liquid chromatographic (HPLC) analysis. The method is based on ultrasonic extraction of three 6-mm plugs cut from a culture using 0.5 ml of solvent followed by a simple solvent change, filtration and injection. Approximately 5 min of work is involved in the extraction and work-up process and the extract can prepared for HPLC analysis within 60–70 min. The method has been used for determination of chromatographic metabolite profiles from 395 fungal isolates, including all terverticillate Penicillium species, cultivated on both Czapek Yeast Autolysate agar and Yeast Extract Sucrose agar. The concentration of the extracts proved to be sufficient to determine all secondary metabolites reported to be produced by these species using HPLC with diode array detection. These findings were confirmed by analyses of 132 pure metabolite standards.     

Classification of polymer batch variations by dynamic headspace/capillary gas chromatography/multivariate data analysis

Gas chromatographic profiles have been generated from different batches of a polypropylene/polyethylene copolymer. The profiles generated from polymer pellets have been obtained by dynamic headspace/capillary gas chromatography analysis. Initially 10 to 40 peaks were chosen at random from the quantitative reports and transferred to a data table. After appropriate scaling the table has been analyzed by a multivariate statistic program, SIMCA (Soft Independent Modeling of Class Analogy) a pattern recognition technique. The method has been used to differentiate batches according to sensory qualities of the final packaging product and changes in polymer peilet production.     

LC Analysis of Oxytetracycline and Chlortetracycline: Application for In Vitro Bio-Equivalence Study of Veterinary Medicines

A liquid chromatographic method has been applied for the analysis of two antibiotics widely used in the veterinary field, oxytetracycline dihydrate and chlortetracycline hydrochloride in premixes for medicated feeding stuffs for veterinary use. In particular, the validated method was employed to study the releasing profile of each drug from two formulations, a commercially available and a new formulation, having different excipient composition. The dissolution profiles obtained from the chromatographic analysis allowed to verify the in vitro bio-equivalence of the commercial and the new formulations for oxytetracycline and chlortetracycline.     

On-site field sampling and analysis of fragrance from living lavender (Lavandula angustifolia L.) flowers by solid-phase microextraction coupled to gas chromatography and ion-trap mass spectrometry

Solid-phase microextraction coupled to gas chromatography and mass spectrometry has been applied as a simple alternative method for the analysis of essential oil directly from lavender intact flowering spikes and genuine oils. All recognised major oil constituents were detected by this procedure, with results comparable to those given by a conventional method (organic solvent extraction). Distinctive chromatographic profiles were found for various species.     

Determination of triphenyltin hydroxide derivatives by capillary GC and tin-selective FPD

A new method for the determination of triphenyltin hydroxide using capillary column gas chromatography with a tin-selective flame photometric detector has been developed. Triphenyltin hydroxide and its potential metabolites are converted to methyl derivatives and separated on glass capillary columns coated with OV-101. Derivatization of triphenyltin hydroxide, triphenyltin chloride, diphenyltin dichloride, phenyltin trichloride, and bis-triphenyltin oxide is nearly quantitative with a minimum of redistribution products. The selectivity of the flame photometric detector is cearly demonstrated by the comparison of chromatographic profiles obtained from using both the flame photometric and flame ionization detectors. The use of this chromatographic system in the analysis of triphenyltin hydroxide in a fortified water sample demonstrates the potential use of this system in organotin residue chemistry.     

High-performance liquid chromatographic method for the simultaneous measurement of floxuridine and fluorouracil in human body fluids.

A method for the simultaneous measurement of floxuridine (5-fluorodeoxyuridine) and fluorouracil in human plasma and peritoneal fluid has been developed. This method utilizes high-performance liquid chromatographic analysis with a Zorbax RX column (25 cm x 4.6 mm I.D.) plus a guard cartridge of the same material. The sensitivity limits for this assay are 0.25 mumol/l in a 20-microliters sample. The detection limit at a signal-to-noise ratio of 3 is 2.5 nmol/l. This procedure has been used to quantitatively measure concentration versus time profiles of floxuridine and fluorouracil in plasma and peritoneal fluid of human patients after receiving intraperitoneal administration of floxuridine.     

Rapid analysis of multicomponent pesticide mixture by GC-MS with the aid of chemometric resolution

Applications of hyphenated chromatographic techniques, especially GC-MS technique, have been reported in chemical, biological, environmental, agricultural and medical analysis. The complexity of the samples in these fields is still an obstacle for the technique to be practical and the overlapping of the multicomponent signals induces chemometric methods widely employed. In this work, taking the rapid analysis of pesticide mixture as an example, a chemometric approach was proposed for resolution of multicomponent overlapping GC-MS signal. In the method, a mass spectral library of pesticides was organized at first, then target factor analysis (TFA) was employed for testing the existence of a specific pesticide in the multicomponent overlapping GC-MS signal, and finally the chromatographic information of the pesticide was extracted by a non-negative immune algorithm (IA). A GC-MS signal of a 40-component pesticide mixture eluted within 9 min was analyzed by the method. It was found that the mass spectra and chromatographic profiles of almost all the pesticides can be obtained.     

Underivatized steroids on persilanized capillary columns with non-vaporizing on-column injection

Recent developments in the preparation of capillary columns by persilanization of the glass surfaces have resulted in highly inactive, heat-stable columns. With this type of column certain steroid hormone classes can be analyzed without derivatization; these include the 17-keto steroids, 11-oxo compounds, estrogens, and progesterone metabolites. It has been found that good quantitative analysis calls for non-vaporizing, on-column injection, as normal vaporizing injection results in breakdown of thermolabile compounds such as estriol and 17-OH progesterone metabolites. The gas chromatographic method described was used to obtain urinary steroid profiles.     

Chemical fingerprinting of Hoodia species and related genera: chemical analysis of oxypregnane glycosides using high-performance liquid chromatography with UV detection in Hoodia gordonii

Hoodia gordonii, family Asclepiadaceae, is a succulent plant and is traditionally used in southern Africa for its appetite-suppressant properties. A high-performance liquid chromatographic (HPLC) method with UV detection for analysis of 11 oxypregnane glycosides from H. gordonii has been developed. The simultaneous analysis of 11 oxypregnane glycosides was achieved with a Phenomenex (Torrance, CA) reversed-phase C18 column using gradient mobile phase of water and acetonitrile, both containing 0.025% trifluoroacetic acid. The developed method was applied to the identification of oxypregnane glycosides in 3 different species of Hoodia and 23 related genera. The HPLC profiles of various plant samples were compared for the presence of oxypregnane glycosides.     

Simple high-performance liquid chromatographic method for the measurement of toloxatone in rabbit cerebrospinal fluid and plasma

A simplified, rapid high-performance liquid chromatographic procedure has been developed for the measurement of toloxatone in rabbit plasma and cerebrospinal fluid. The method involves a single-step extraction of the alkalinized sample with diethyl ether and analysis of the evaporated extract on a C8 column. Detection was performed by ultraviolet absorbance monitored at 240 nm. The overall run-time of the assay was 8 min at a flow-rate of 1 ml min-1. The sensitivity limit of toloxatone was 70 ng ml-1 at a signal-to-noise ratio of 3:1 in rabbit cerebrospinal fluid and plasma. The assay has been used to define plasma toloxatone concentration-time profiles and to quantitate cerebrospinal fluid toloxatone levels after a single intravenous injection in rabbits.     

High-performance liquid chromatographic assay of a new non-steroidal anti-inflammatory agent, 2-(10,11-dihydro-10-oxodibenzo[b,f]thiepin-2-yl)propionic acid, in human plasma and urine

A high-performance liquid chromatographic method for the quantitation of a new anti-inflammatory agent, 2-(10,11-dihydro-10-oxodibenzo[b,f]thiepin-2-yl)propionic acid (CN-100; I), has been developed. The assay consists in extracting samples containing I and mefenamic acid, the internal standard, under acidic conditions and analysis by reversed-phase chromatography using ultraviolet detection at 330 nm. Preliminary plasma concentration-time and cumulative urinary excretion profiles from a healthy subject following oral administration of the tablet formulation are presented. This method is simple, sensitive and reproducible and is applicable to studies of the pharmacokinetic behaviour of I in humans.     

Continuous melting and ion chromatographic analyses of ice cores

A new method for determining concentrations of organic and inorganic ions in ice cores by continuous melting and contemporaneous ion chromatographic analyses was developed. A subcore is melted on a melting device and the meltwater produced is collected in two parallel sample loops and then analyzed simultaneously by two ion chromatographs, one for anions and one for cations. For most of the analyzed species, lower or equal blank values were achieved with the continuous melting and analysis technique compared to the conventional analysis. Comparison of the continuous melting and ion chromatographic analysis with the conventional analysis of a real ice core segment showed good agreement in concentration profiles and total amounts of ionic species. Thus, the newly developed method is well suited for ice core analysis and has the advantages of lower ice consumption, less time-consuming sample preparation and lower risk of contamination.     

Validation of the concentration profiles obtained from the near infrared/multivariate curve resolution monitoring of reactions of epoxy resins using high performance liquid chromatography as a reference method

This paper reports the validation of the results obtained by combining near infrared spectroscopy and multivariate curve resolution-alternating least squares (MCR-ALS) and using high performance liquid chromatography as a reference method, for the model reaction of phenylglycidylether (PGE) and aniline. The results are obtained as concentration profiles over the reaction time. The trueness of the proposed method has been evaluated in terms of lack of bias. The joint test for the intercept and the slope showed that there were no significant differences between the profiles calculated spectroscopically and the ones obtained experimentally by means of the chromatographic reference method at an overall level of confidence of 5%. The uncertainty of the results was estimated by using information derived from the process of assessment of trueness. Such operational aspects as the cost and availability of instrumentation and the length and cost of the analysis were evaluated. The method proposed is a good way of monitoring the reactions of epoxy resins, and it adequately shows how the species concentration varies over time.     

Comprehensive description of the photodegradation of bromophenols using chromatographic monitoring and chemometric tools

A general procedure for the study of complex photodegradation processes of environmental pollutants based on chromatographic monitoring and chemometric method is proposed. The procedure consists of multiset data analysis of aliquots collected at different reaction times and injected in High-Performance Liquid Chromatography coupled to diode array detection and mass spectrometry (HPLC-DAD-MS). In this study, photodegradation of six bromophenols with different degrees of bromination has been investigated in order to find out their photodegradation pathways and kinetics and to show the potential of the procedure proposed. Multivariate curve resolution-alternating least squares (MCR-ALS) has been used to resolve chromatographic elution profiles and pure spectra of species involved in the photodegradation process and, hence, to elucidate the photodegradation mechanism and to propose the chemical structure of the main photoproducts. This study shows that chromatographic monitoring is the preferred option when photochemical systems with large number of components with similar spectra and kinetic evolution are analyzed. This work reveals the advantages of the double DAD and MS detection to provide mechanistic and structural information about these complex photodegradation processes.     

Instrumental method for automated on-line fraction analysis and peak deconvolution in multicomponent-overloaded high-performance liquid chromatography

A bi-dimensional (non-orthogonal) chromatographic system made of two columns of different length (longer along the first direction and shorter on the second) but packed with identical (or similar) materials has been employed to perform automated on-line fraction analysis and peak deconvolution of multicomponent overloaded band profiles. Multicomponent overloaded peaks, eluted from the first column, are sampled through a remotely-controlled switching valve and fractions are injected, under analytical conditions, directly into the second direction. This set-up serves a twofold purpose. First of all, it has been possible to use linear calibration curves for detector calibration (DC). Secondly, since single component peaks were resolved on the second direction, individual concentration profiles have been obtained beneath the envelop of overlapping bands. All the information necessary to operate the bi-dimensional system can be achieved by analyzing the chromatographic behavior of the mixture on the shorter column under linear conditions. Therefore, solutions of unknown concentration can be prepared and their composition reconstructed with a simple chromatographic run. Two- and three-component overloaded mixtures have been employed to validate the system.     

HELP法在中草药分析中的应用研究-冬虫夏草的化学成分测定

基于直观推导式演进特征投影（ＨＥＬＰ）法,对冬虫夏草子座和虫体分别进行了多组分同时定性定量测定．结果表明,ＨＥＬＰ法能减少样本提取分离的步骤,降低色谱分离条件的要求,提高检测准确度．联用色谱检测与化学计量学解析法相结合将为复杂中草药分析提供一种全新手段     

免疫算法用于复杂样品的GC-MS快速分析

建立了一种基于免疫算法的复杂样品气相色谱-质谱联用(GC-MS)快速分析方法, 该方法采用快速温度程序测定复杂样品的GC-MS信号, 再通过快速解析得到各组分的信息. 计算过程中, 采用各种可能存在于混合物中的组分质谱作为算法的输入, 按照保留时间的顺序逐一对重叠的质谱信号进行解析, 得到各组分的色谱信息. 对于混合物中实际存在的组分可得到该组分的色谱流出曲线, 而对于混合物中不存在的组分所得色谱流出曲线几乎为零. 采用所建立的方法对含有16个组分的有机磷农药混合物进行了分析, 在保留时间10 min内得到了所有组分的色谱信息.     

Suitability of selected chromatographic columns for analysis of fatty acids in dialyzed patients

Gas chromatography–mass spectrometry is a preferred method for fatty acid (FA) analysis in biofluids from patients with metabolic diseases. Complex characteristics of FAs make their analysis particularly challenging. Selection of an appropriate chromatographic column is particularly important component of the process as it provides optimal separation and detection of possibly all FAs present in the sample. However, no accurate protocol for comparative evaluation of capillary columns for the analysis of whole serum FA profile in patients with chronic kidney disease (CKD) has been developed thus far. Therefore, in the present study four columns were examined to select the one providing optimal separation and determination of FA profiles in this group of patients. Moreover, serum FA profiles obtained with the selected column in CKD patients subjected to peritoneal dialysis and healthy controls were compared. Thirty‐seven component FAME Mix and sera from CKD patients were used to optimize chromatographic conditions and to select the most appropriate column. The ZB‐5 column turned out to be the most appropriate for the analysis of whole FA profile in CKD patients' sera. Then, this column was used to compare FA profiles in patients subjected to peritoneal dialysis and in healthy controls. The analysis demonstrated many abnormalities in the FA profile of CKD patients. Further studies involving larger groups of patients presenting with other stages of CKD are required to explain the impact of the disease progression on composition of serum FAs.     

Time-resolved cryogenic modulation reveals isomer interconversion profiles in dynamic chromatography.

The dynamic chromatographic study of interconversion of E and Z forms of oximes has been investigated by using a novel cryogenic modulation method in a two-dimensional gas chromatographic array. The primary column is a conventional capillary GC column on which the molecular interconversion proceeds. In this case, the molecular dynamical process leads to a peak profile describing the kinetics and thermodynamics of the interconverting molecules during its chromatographic elution. Thus an interconversion region intercedes the elution of the individual stereoisomers of the reaction. Since the molecules are isomers, classical molecular identification methods such as gas chromatography-mass spectrometry are unable to study the individual instantaneous amounts of each of the compounds. Hence the infinitesimal profiles of interconversion along the entire column have never been experimentally observed; rather the total profile is normally subjected to mathematical modelling studies in order to match experiment with theory, and to gain the kinetic parameters of the process. In the present study, an instantaneous ratio of the individual isomers can be found during the chromatographic elution by direct measurement. This is achieved by using a cryogenic zone focussing process, with rapid longitudinal modulation of a cold trap and continual pulsing of collected zones into a fast-analysis high-resolution capillary column on which isomer interconversion is minimized. The data can be displayed as a two-dimensional contour plot to demonstrate the individual isomer profiles. The two-dimensional analysis also allows easy measurement of the peak ratios of the two isomers which is an indicator of the extent of interconversion that has taken place. Two model systems, acetaldoxime and butyraldoxime, were chosen to illustrate the use of the cryogenic modulation procedure. It is anticipated that the procedure could be applied to other molecules which exhibit gas-phase isomerizations or reactions.