Quantification of sugars and organic acids in hygroscopic pharmaceutical herbal dry extracts

Three chromatographic methods have been employed for the determination of hydrophilic compounds, namely carbohydrates and organic acids in herbal dry extracts of Eschscholtzia californica Cham. The hydrophilic compounds were separated from the other components of the dry extracts by solid-phase extraction methods, which were optimised with respect to recovery rates. Carbohydrates were quantified using high-performance anion-exchange chromatography with pulsed amperometric detection. Organic acids were analysed by ion-exclusion chromatography with evaporative light scattering detection and gas chromatography-mass spectrometry. Using the latter method, large amounts of glyceric acid were separated from the extracts of Eschscholtzia californica Cham. This substance together with sugars may be responsible for the increased hygroscopicity and the poor processing behaviour of the extracts.     

Speciation of aluminum in soil extracts using cation and anion exchangers followed by a flow-injection system with fluorescence detection using lumogallion.

Flow-injection analysis (FIA) with fluorescence detection of aluminum using lumogallion was applied to the chemical speciation of aluminum in soil extracts after the separation of aluminum species with ionic exchangers. Aluminum complexes with organic substances (anion species) can be specified from other species by using a strongly acidic cation exchanger in the pH range of 3 to 5. Furthermore, aluminum can be separated into three categories, namely, (i) the Al3+ and Al-OH complex, (ii) aluminum organic complexes (cation species), and (iii) its anion species by using a strongly acidic and a weakly acidic cation exchanger at around pH 5. A considerable percentage of water-soluble aluminum in soils was found to be complexes with humic substances.     

Electroactivity of nitriles on platinum electrodes and its use in chromatographic detection

Acetonitrile yields two oxidative peaks, first at ca. +0.30 and second at ca. +1.15 V vs. Ag/AgCl in cyclic voltammetry with platinum electrodes in 0.10 M methanesulfonic acid (MSA) containing 0.05–5 mM concentrations of acetonitrile. This electroactivity of the nitrile group was used for a direct detection of nitriles after their chromatographic separation. Three organic nitriles (acetonitrile, propionitrile and butanenitrile) were separated with an IonPac ICE-AS 1 column, eluted with 0.10 M MSA and detected on a platinum electrode via pulsed amperometric detection. Analytical performance was evaluated with a three potential waveform (+0.30 V, +1.15 V, −0.30 V vs. Ag/AgCl, current integration at +1.15 V). Numerical values of detection limits, linearity of calibration and reproducibility are reported for all three organic nitriles.     

Selective dissolution and one step separation of terpene trilactones in ginkgo leaf extracts for GC-FID determination

Under ultrasonication, the ginkgo terpene trilactones, ginkgolides and bilobalide, in ginkgo extracts can be selectively dissolved in 10% aqueous NaH(2)PO(4) solution at a temperature of 50-60 degrees C and separated from the solution by extraction with a mixture of ethyl acetate/tetrahydrofuran in a capped vial. After derivatization, these terpene trilactones can be quantified using GC-FID. This method has a detection limit of 10 ng, and the RSD was 6% (n=5). Twelve commercial GBE products in powder, liquid, tablet and capsule forms were analyzed. The total time required for analyzing these samples from sample preparation to final data processing was less than 6 h, and the total organic solvent consumption was less than 40 ml. This procedure proves to be a simple, fast, safe, and effective method for all types of Ginkgo biloba extracts (GBE) including the "complex" or "advanced" formulas.     

Hydrocarbon type analysis by thin-layer chromatography with flame-ionization detection: vacuum gas oils, heavy feeds, and hydroprocessed products

Thin-layer chromatography with flame-ionization detection (TLC-FID) provides quantitative hydrocarbon type data as well as distribution of aromatics by ring number. This method has been applied to obtain amounts of saturates, aromatics, and polars in heavy oil distillates such as light vacuum gas oils and heavy vacuum gas oils derived from different crude sources. TLC-FID chromatograms and resultant quantitative hydrocarbon type data show that these distillates vary markedly in aromatic contents and aromatic ring types. Similar observations are made with several fluid catalytic cracking feeds. Effects of process parameters such as operating pressure and temperature on hydroconversion of aromatics and polars from a heavy oil are assessed by TLC-FID. It has been demonstrated that there is a preferential reduction of higher polycyclic aromatic hydrocarbons and polars with an increase of both hydrogen partial pressure and reactor temperature.     

Characterization and determination of organic compounds in the mutagenic XAD-2 extracts of drinking water.

Amberlite XAD-2 extracts, which exhibit mutagenicity in the Ames assays, of drinking water sampled each month during the period from April 1988 to March 1989 were studied in order to characterize and determine the organic pollutants. The major organic pollutants were phthalate ester plasticizers such as dibutyl and di(2-ethylhexyl) phthalate. Several polynuclear aromatic hydrocarbons (PAHs) and the organocholorine pesticide oxadiazon were also identified to be present at low concentrations. The XAD-extractable and chromatographable organic pollutants were found to be composed of PAHs with a mean concentration of 0.136 micrograms/l(ca. 10% of the total amount of organic compounds detected), phthalates with a mean value of 0.405 micrograms/l(ca. 30%) and other compounds with a mean value of 0.845 micrograms/l(ca. 60%). The concentrations and compositions of these organic pollutants were correlated with the effective rainfall content of the river and with the water temperature.     

Determination of delta 9-tetrahydrocannabinol in plasma using solid-phase extraction and high-performance liquid chromatography with electrochemical detection

A method for the determination of nanogram amounts of delta 9-tetrahydrocannabinol (THC) in plasma and serum is described. THC was quantitatively isolated by solid-phase extraction after addition of an aqueous solution of urea and methanol to the sample. The extracts were analysed by high-performance liquid chromatography with electrochemical detection in the oxidizing mode. The detection limit of THC is ca. 100 pg for a signal-to-noise ratio of 3. With this method, levels of 2 ng/ml of THC in plasma can be measured.     

Analysis of vinca alkaloids in plasma and urine using high-performance liquid chromatography with electrochemical detection

The reversed-phase high-performance liquid chromatography with electrochemical detection was used to quantify plasma and urine levels of vinblastine, vincristine, vindesine and a metabolite of vinblastine, desacetylvinblastine. Sample clean-up consisted of solid-phase extraction with a Bond Elut CN column. The extracts were separated on a Hypersil ODS column. The mobile phase consisted of a mixture of methanol and 10 mM phosphate buffer (pH 7.0). The limit of sensitivity using electrochemical detection was 100 pg on-column for all compounds with a signal-to-noise ratio of 3. Quantification of the compounds in human plasma and urine was possible down to 1 ng/ml (ca. 1 pmol). Pharmacokinetic results show that the sensitivity of the method is adequate for drug monitoring in clinical research.     

Separation of glucuronides from urine by coupled-column separation using underivatized silica as precolumn.

Glucuronides are separated from urine by coupled-column separations (CCSs). The fraction containing the glucuronide(s) is transferred on-line from a silica precolumn to the analytical column (octadecyl derivatized silica), enriched, and separated by ion-pair chromatography. The retention and selectivity on the precolumn are controlled by pH, buffer components, organic modifier, and ion-pair agent. After the injection of filtered urine samples, glucuronides with different chemical properties can be separated. The total analysis of morphine-3-glucuronide and morphine-6-glucuronide is accomplished in less than six minutes, with UV detection at 210 nm.     

Separation and characterization of basic barley seed proteins

Basic proteins in barley starchy endosperm from developing seeds were separated by two-dimensional (2-D) nonequilibrium pH gel electrophoresis. Total as well as partial extracts were analyzed. Edman degradation sequencing and immunological detection were performed after transfer of separated proteins onto membranes. Only one protein could be analyzed by N-terminal sequencing of blotted and separated proteins from the total extract. Fractionation of extracts was done using cation exchange chromatography, concanavalin A and heparin affinity chromatography. Internal sequences were determined after in-gel cleavage of proteins using trypsin or cyanogen bromide and separation of the fragments by reversed-phase chromatography or in a gel electrophoresis system for peptide separation. This resulted in a new protocol for obtaining internal sequences from proteins separated by 2-D electrophoresis. A total of 16 sequences, including nine internal sequences, were analyzed, permitting the identification of ten proteins, including five that appeared to have a blocked N-terminus. An additional protein was identified using immunological detection. Three protein sequences remained unidentified. Separated proteins were also analyzed with a glycan detection method.     

Determination of mono- and disaccharides by capillary electrophoresis with contactless conductivity detection

The separation and detection of common mono- and disaccharides by capillary electrophoresis (CE) with contactless conductivity detection (CCD) is presented. At high values of pH, the sugars are converted to anionic species that can be separated by CE and indirectly detected by CCD. The main anionic species present in the running electrolytes are hydroxide and phosphate, which have greater mobility than the ionized sugars, and, thus, the indirect detection is possible. The method was applied to analysis of glucose, fructose, and sucrose in soft drinks, isotonic beverages, fruit juice, and sugarcane spirits. Galactose was used as internal standard in all cases. Plate numbers range from ca. 70,700 to 168,200 and the limits of detection from 13 to 31 microM.     

Direct infusion electrospray ionization mass spectra of crude cell extracts for microbial characterizations: influence of solvent conditions on the detection of proteins

Direct infusion electrospray ionization mass spectrometry (DIES-MS) of crude bacterial extracts is a rapid method that can be used to characterize microbial cells. Phospholipids, metabolites, and proteins can be detected rapidly with minimal sample preparation. However, several factors influence the detection of signals in such high-throughput analyses. We studied the influence of solvent conditions, including the organic content and pH of the solvent, on the extraction and subsequent detection of signals in DIES-MS, with a view to improving the detection of protein signals. Unfractionated cell extracts from three strains of the Gram-negative Escherichia coli (including one encoding a recombinant green fluorescence protein), and the Gram-positive Bacillus sphaericus and B. subtilis were investigated. Both pH and the organic content of the solvent were found to influence the spectral information as observed from principal component analysis of the spectral data. A polar solvent with higher organic content resulted in the extraction of phospholipids that overtly dominate the spectral information. Decreasing the organic content of the extraction solvent resulted in the improved detection of protein peaks. Altering the pH of the extraction solvent resulted in different protein profiles from the same bacterium, as observed after spectral deconvolution. In addition, the protein profiles were also different when using different organic solvents. Spectral deconvolution showed several protein peaks that had mass-based homology with those in protein databases for the (sequenced) organisms studied. These results suggest that a combination of solvent conditions can be used to generate protein profiles rapidly that when combined can provide additional valuable proteomic information.     

Speciation of dissolved phosphorus in environmental samples by gel filtration and flow-injection analysis

A study of the factors affecting separation and detection of dissolved organic and inorganic phosphorus species found in waters sediments is reported. The system involved the use of gel filtration and flow injection analysis (FIA). Orthophosphate and myo-inositol hexakisphosphate, as model solutes representative of low molecular weight P (LMWP) and high molecular weight P (HMWP), were separated on a Sephadex G25 column incorporated into a flow-injection manifold which utilized photo-oxidation and spectrophotometry for detection of dissolved reactive phosphorus (DRP) and dissolved organic phosphorus (DOP). The influence of eluent pH and ionic strength on adsorption and anionic exclusion of the model solutes is described, and the optimum eluent composition and sample size are described. The method was used to determine LMWP and HMWP in natural and waste waters, and in sediment extracts. Potential limitations of this approach are discussed.     

The identification of the sign and strength of disclinations in the schlieren (nucleated domain) texture of the nematic phase,by optical microscopy

ABSTRACT

The optical texture of the nematic phase, variously known as the schlieren, structure à noyuax or nucleated domain texture, was identified over a century ago as being an array of point singularities. When viewed between crossed polars, patterns of dark brushes radiate from each point nucleus. The sign and strength of each nucleus can be uniquely determined from the changes in the orientation of these brushes when either the sample or the crossed polars are rotated, from two formulae given by Chadrasekhar in 1977. However, these were given with little exemplification and have been largely overlooked. Consequently, the majority of the discussions given in current literature are either incomplete and confusing or, in some cases, incorrect. Here, we provide a detailed explanation of the textures and their behaviour as viewed with the most commonly used experimental geometry (i.e. with a rotating sample and stationary polars).     

Potential of Capillary Electrophoresis for the Profiling of Propolis

The usefulness of capillary electrophoresis (CE) with diode array detection for the profiling of Propolis, a hive product, is investigated. Water extracts of Propolis were analyzed with both capillary zone electrophoresis (CZE) at pH 7.0 and 9.3, and micellar electrokinetic chromatography (MEKC) with sodium dodecyl sulfate at pH 9.3. Characteristic profiles were obtained and several organic acids and preservatives could be identified by means of library comparison of the recorded UV spectra combined with addition of reference compounds to the extracts. The selectivity of the CZE and MEKC system differed considerably but the information obtained with both methods was similar. The dry residues of the water extraction were extracted with ethanol-water (70 : 30, v/v) and analyzed with the MEKC system to enable the separation of the more hydrophobic constituents of the Propolis samples. Complex profiles containing various well separated peaks were obtained allowing the identification of some interesting flavonoids. On the basis of the recorded CZE and MEKC profiles, the Propolis samples could be divided into two clearly different groups which are probably from a different origin.     

Determination of phosphorus by electrothermal atomic absorption spectrometry with a phosphorus hollow-cathode lamp as light source

Graphite tubes are pre-heated with a 10% (w/v) zirconium solution, and a 1% (w/v) zirconium solution is injected prior to each phosphorus sample solution. The calibration graph is linear up to ca. 100 ng P, and the detection limit is 5 ng. Phosphorus in water-soluble organic and inorganic compounds can be determined directly by the proposed method, which is also suitable for the determination of phosphorus in NBS standard reference plant materials and in green tea leaves.     

Determination of 2,4-D and Dicamba in food crops by MEKC

Summary The determination of 2,4-D (2,4-dichlorophenoxyacetic acid) and Dicamba (2-methoxy-3,6-dichlorobenzoic acid) residues in sugar cane, rice and corn was performed by a supercritical fluid extraction (SFE) method using CO₂/acetone as extraction mix and an SFE apparatus developed in our laboratory. The extracts were cleaned up after extraction by both liquid-liquid partition and a Florisil column. Micellar electrokinetic capillary chromatography (MEKC) coupled with ultraviolet on-column detection was used for the analysis of these pesticides. The detection limits were improved by the preparation of a special detection cell with an increased pathlength that gave detection limits of ca. 0.6 pg for 2,4-D and Dicamba. Our results demonstrated that capillary electrophoresis can be a powerful new analytical tool for pesticide residue analysis.     

A hybrid microdevice for electrophoresis and electrochromatography using UV detection

We have designed a new class of microdevices composed of a supporting plastic (polyvinyl chloride, PVC) plate integrated with a groove for a piece of fused silica capillary (the separation channel), a slit for on-tube detection, an "islet" for the application of sample, electrode vessels and platinum electrodes. The design permits electrophoretic, electrochromatographic and chromatographic separations with on-tube UV detection. The efficient heat dissipation allows relatively high field strengths. This article is the first one dealing with microdevices where polymer solutions are replaced by homogeneous gels. A new type of gels synthesized from acrylamide and 2-hydroxy-3-allyloxy-propyl-beta-cyclodextrin (allyl-beta-CD) as a cross-linker was employed for electrophoresis and electrochromatography. 2-Acrylamido-2-methylpropanesulfonic acid was added to the monomer solution to create a high electroendosmotic flow in electrochromatographic runs. These gels have excellent electrochromatographic and electrophoretic properties for low-molecular-weight compounds and DNA, as shown previously, namely high resolution combined with high stability. The unique cross-linker can be used for specific interaction with the alkyl and phenyl groups. The tripeptide glutathione (gamma-L-glutamyl-L-cysteinyl-glycine) and its benzyl conjugates were selected as model compounds to study the resolving power of the gel because they are difficult to separate by free zone electrophoresis. The limit of detection (LOD) for S-benzyl-glutathione was determined (ca. 7 microM). Run-by-run reproducibility was high (the separation factor of glutathione in the gel was 0.3 with 2.5% coefficient of variation, CV). Neutral compounds (acetone, acetophenone, propiophenone and butyrophenone) were separated electrochromatographically in the gel. The influence of organic solvent (acetonitrile) on the electroendosmotic mobility was similar to that in reversed-phase separations, although the separation mechanism is different. ATP, ADP and AMP were separated in less than 10 s by free-zone electrophoresis.     

Determination of organic acids, amino acids and saccharides by high-performance liquid chromatography and a postcolumn enzyme reactor with amperometric detection.

A technique for the determination of organic acids, amino acids and sugars is described. The compounds of interest are separated by high-performance liquid chromatography (HPLC) and converted on-line by immobilized enzymes. The enzymes employed are covalently bound to a synthetic carrier. Hydrogen peroxide, which is produced in the reaction with oxidases, makes possible the application of an electrochemical detector. This arrangement combines the separation efficiency of HPLC, the substrate specificity of enzymes and the high sensitivity of electrochemical detection. The enzymes act according to known reaction mechanisms, but coupling with HPLC leads to a promising extension in the field of biosensors. The simple pretreatment of the samples (often a dilution step is sufficient) allows a rapid analysis of foodstuffs and biological or clinical extracts. The examples presented demonstrate the very high sensitivity of the method with detection limits in the nano- to picomolar range and a wide field of application.     

Headspace gas chromatography-flame ionization detector method for organic solvent residue analysis in dietary supplements

An analytical method has been developed for the identification and quantification of 20 organic solvent residues in dietary supplements. The method utilizes a headspace sampler interfaced with gas chromatography and flame ionization detection. With split injection (5:1) and a DB-624 column, most of the organic solvents are separated in 9 min. The method has been validated and was found to be relatively simple and fast, and it can be applied to most common organic solvent residues. With the mass detector, the method was able to identify organic solvents beyond the 20 standards tested.