Chemical fingerprinting of Andrographis paniculata using HPLC, HPTLC and densitometry

An attempt has been made to develop a method by which to determine the chemical fingerprint of Andrographis paniculata (Acanthaceae). High-performance thin layer chromatography (HPTLC) was used to analyse hexane, chloroform, methanol and water extracts of leaves of A. paniculata. A computerised densitometer was applied to the two-dimensional spectrographic image analysis of the HPTLC plates. An HPLC equipped with a photodiode array detector was used for the analyses of these different extracts. The analyses showed that andrographolide and neoandrographolide are absent in the hexane extract but are present in greater amounts in the methanol extract as compared with the other extracts. These chromatograms may serve as a chemical fingerprint of the drug A. paniculata for quality control purposes and in the preparation of formulations based on the drug.     

Chemical fingerprinting of Lawsonia inermis L. using HPLC, HPTLC and densitometry

Introduction –  Lawsonia inermis L. is a natural red colouring agent, commonly named “Henna”, which is used to dye skin and hair. The aim of this study was to evaluate the quality of L. inermis that is commercially available as a raw plant material or preparation in order to guarantee good quality products. Objective ?  To develop a simple protocol for the qualification of different samples labelled as L. inermis by using the HPTLC densitometry method and to identify possible adulterations with other plants. Methodology ?  Samples of leaves of L. inermis were extracted with methanol. Two chromatographic methods were developed to determine the chemical fingerprinting of L. inermis. The first was based on HPTLC identification followed by densitometric measurements at 337 nm. The second was based on RP‐HPLC separation with gradient elution and photodiode array detection at 337 nm. Samples of Cassia obovata Collad., and Indigofera tinctoria L., were treated in the same way. Results –  The simplicity of the sample preparation, and the possibility of analysing several samples of herbal products simultaneously in a short time, make HPTLC the method of choice. The HPTLC method was feasible for the comprehensive quality evaluation of herbal products. From the comparison of their “fingerprint”, it was possible to detect substitution of plants that are different from those declared on the label. Conclusion ?  The HPTLC may be used as a rapid method by which to control the quality of raw plant materials and formulations based on the title plant. Copyright © 2008 John Wiley & Sons, Ltd.     

Stabilization of standard samples of levorin and nystatin with antioxidants]

To prolong the storage period of the reference samples of levorin and nystatin, polyenic macrolide antibiotics, their effect of 23 antioxidants was studied by using rapid inactivation. The antioxidants belonged to compounds of different classes. The inactivation was performed at 20, 37 and 50 degrees C in the presence of 1, 2 or 5 per cent of the antioxidants. An antioxidant of the class of partially hydrated oxyquinolines was shown to have the highest stabilizing action on levorin and nystatin. It may be recommended for stabilizing the levorin and nystatin reference samples.     

Sterol composition in yeast strains sensitive and resistant to nystatin

The sterol composition of nystatin-sensitive and nystatin-resistant strains of Saccharomyces cerevisiae was being studied by gas-liquid chromatography and mass-spectroscopy. The synthesis of ergosterol is completely suppressed in polyene-resistant mutants. Three sterols derived from cholesterol were identified in the mutants: cholesta-8,24-diene-3 beta-ol, cholesta-5,7,24-triene-3 beta-ol, and cholesta-5,7,22,24-tetraene-3 beta-ol.     

A study of regional ventilation of the lungs in patients with chronic bronchitis using scanning densitometry tomograms]

A new method for examination of regional ventilation of the lungs has been developed, that permits assessment of the ventilation in patients with chronic bronchitis and detect a latent respiratory insufficiency. The results of examinations with this method conform to the results of examinations of external respiration function.     

Use of antibiotics in selecting a nystatin producer]

The lethal and mutagenic effect of streptomycin and nystatin on Act. noursei, strain 408 producing nystatin was studied. The survival of the spores of strain 408 on the medium with streptomycin decreased with an increase in the antibiotic concentration. Streptomycin had a selective effect on the nystatin-producing organism decreasing the frequency of morphologically changed and low active variants and revealing highly active and antibiotic stable variants. The survival of the spores of strain 408 on the medium with nystatin (20,000 units/ml) amounted to 35 per cent. Nystatin had an inhibitory effect on the organism producing it which was evident from delayed growth and significant modification variation of the colonies, as well as from a marked increase in the number of the variants characterized by low antibiotic production.     

Determination of fungal glucosamine using HPLC with 1-napthyl isothiocyanate derivatization and microwave heating

A rapid method for the determination of fungal glucosamine (GlcN) from Aspergillus sp BCRC 31742 was developed. The hydrochlorination process using microwave effectively reduced reaction time needed for GlcN analysis. The analytical method consisted of two steps: (1) hydrochlorination of fungal cells and (2) derivatization process. Fungal GlcN hydrochloride was reacted with 1-napthyl isothiocyanate (1-NITC) as the derivatizing agent to enhance the sensitivity of GlcN and so to achieve high resolution. This method was specific for quantification of GlcN hydrochloride at the wavelength of 230 nm. The standard deviation and relative error of the analytical results were less than 5%. By using microwave heating, the reaction time of hydrochlorination process was shortened from 24 h to 3 min. Thus, the overall time needed for analyzing GlcN from fungal sources was reduced from 5 h (thermal method) to 2 h (microwave method).     

Determination of catecholamines in tissue and body fluids using microbore HPLC with amperometric detection

Performance of microbore reverse phase HPLC coupled with amperometric detection is detailed for the analysis of catecholamines in small tissue samples and human blood plasma and cerebrospinal fluid. Extraction procedures for pre-concentration and clean-up of these samples are described. Marked signal enhancement is observed due to the smaller column volume as well as the increased coulometric yield which results from the lower flow rates used with this technique. Detection limits of 0.2 to 0.5 picograms are obtained allowing analysis of catecholamines in extremely small tissue samples or small volumes of cerebrospinal fluid or plasma.     

Determination of reactor biomass by acoustic resonance densitometry

Acoustic resonance densitometry (ARD) is reported as a method suitable not only for precise investigations into changes of specific gravity in bioreactor media but also as a technique able to provide an accurate wide range and direct determination of cellular mass in fermentation processes. It is further shown that this method can replace present optical procedures, minimizing dilution errors and operator involvement and is suitable for development as an on-line biomass monitoring system.     

Determination of plasmid copy number by fluorescence densitometry

A simple and reliable method for the determination of plasmid copy numbers by direct fluorescence densitometry of ethidium bromide-stained electrophoretic gels was developed. In developing the method, the following parameters were evaluated and controlled: plasmid DNA trapping in the linear chromosomal DNA, staining-destaining kinetics for ethidium bromide, linearity of the fluorescence response, and the effect of the molecular topology of DNA on ethidium bromide binding to DNA in agarose.     

薄芝糖肽的单糖组成分析

目的对灵芝属薄树芝的活性成分薄芝糖肽的单糖进行研究.方法采用薄层色谱法和高效液相色谱-蒸发光散射检测器(HPLC-ELSD)联用技术.结果薄芝糖肽的单糖组成及摩尔比为葡萄糖∶D-甘露糖∶半乳糖∶木糖=1∶0.074∶0.067∶0.0178.结论建立了薄芝糖肽单糖组成的检测方法.     

HPLC法检测苦杏仁苷及其酶解产物

本文建立了一个快速高效的HPLC方法同时检测苦杏仁苷及其4个体外酶解产物,色谱分析系统为安捷伦液相色谱系统,自动进样器,迪马ODS C18色谱柱(250 mm×4.6 mm,5μm),柱温25℃,流动相30%甲醇,检测波长210 nm,进样量10μL。结果表明,苦杏仁苷、phenyl-(3,4,5-trihydroxy-6-methyl-tetrahydro-pyran-2-yloxy)-acetonitrile、野樱花苷、苯甲醛和杏仁腈的保留时间分别为5.087、13.836、16.357、22.775和3.307 min。该HPLC方法的重现性好,精确度和准确度高,当柱温低于30℃、pH值介于6.0~7.0时该方法结果稳定、重现性好。     

Determination of moniliformin using SAX column clean-up and HPLC/DAD-detection

This work describes a method for the determination of theFusarium mycotoxin moniliformin (MON) in cereals. In addition to the optimization of the clean-up and the HPLC determination the most efficient extraction mode was investigated on natural contaminated samples. The method was validated for maize and wheat using a calibration range from 57 to 2300 μg/kg. Due to the ionic nature of the toxin the clean-up of the extracts was carried out with strong-anion-exchange columns. Moniliformin was separated by reversed phase ion-pair-chromatography (RP-Ion pair-HPLC) and detected by DAD. The validated method yielded recoveries of 76%±9% (maize) and 87%±5% (wheat) and detection limits of 39 μg/kg and 30 μg/kg, respectively. The suitability of the developed method was demonstrated on natural contaminated samples. Presented at the 25^th Mykotoxin Workshop in Giessen, Germany, May 19–21, 2003     

Effect of nystatin,amphotericin B and amphotericin B methyl ester on Saccharomyces cerevisiae with different lipid composition

Saccharomyces cerevisiae was cultured under anaerobiosis in semi-complete medium to which either palmitoleic or oleic acid was added. Cells were grown at 20 °C or 30 °C. The levels of total lipids, total sterols, and phospholipids were higher in cells grown at 20 °C than at 30 °C. The effects of nystatin (NYS), amphotericin B (AMB), and amphotericin B methyl ester (AME) were evaluated by determining cell viability and liberation of intracellular compounds. The loss of cell viability is higher in the first 30 minutes of incubation with the drugs and is the same regardless of the type of cells obtained. Low molecular weight compounds and ions such as K⁺ are liberated a few minutes after incubation with the drugs whereas proteins and substances absorbing at 260 nm are liberated later. Phosphate liberation comes after K⁺ and before compounds of higher molecular weights.     

Correlation between serum glutathione reductases and bone densitometry values]

Free radicals, because of their marked chemical activity, have often been found to be involved in many human diseases. Enzymatic antioxidant systems, namely glutathione-reductase, present both in red blood cells and in serum, have been demonstrated to play a key role as free radicals scavengers. The present work has been carried out in order to evaluate the possible role played by free radicals in the demineralization process of the bone matrix. Glutathione-reductase activity, assayed by a slightly modified Horn's method, was related to bone density measurements. All the subjects with reduced densitometric values showed reduced glutathione-reductase levels. Our results seem to support the hypothesis of a strict relationship between low activity of antioxidant systems and demineralization process of the bone, in consequence of enhanced free radical levels.     

Determination of the enantiomeric composition of a new insulin sensitizer in plasma samples from non-clinical and clinical investigations using chiral HPLC with electrospray tandem mass spectrometric detection

Two drug assays were developed and applied to assess the enantiomeric composition of an insulin sensitizer drug in plasma after administration of its racemate to man, and in human and animal plasma and serum samples generated after in vitro experiments. The sample preparation for the assays consisted either of protein precipitation and column-switching, or liquid-liquid extraction and direct injection. Subsequently, both assays employed chiral HPLC coupled to atmospheric pressure ionization mass spectrometry. An interconversion of the racemate to a mixture enriched with the (+)-enantiomer could be confirmed for all species and biological matrices. The individual enantiomers could be quantified in the concentration range 0.5-500 ng/ml, starting with a 100-microl plasma aliquot. Inter- and intra-assay precision and accuracy were in the range 0.1-7.9 and 88.8-106.0%, respectively. Run times of 5 min for a single sample allows the analysis of more than 200 samples overnight.     

Simultaneous Determination of Bulky Imides and Their Hydrolyzed Products by D-Hydantoinase with Cyclic-imide-hydrolyzing Activity using HPLC

Applied Biochemistry and Microbiology - A novel HPLC method for simultaneous detecting bulky imides and their hydrolyzed products by D-hydantoinase was established. Phthalimide (PI) and...