Separation of Ribose-5-phosphate,Ribose-1-phosphate,Deoxyribose-1-phosphate by High Performance Liquid Chromatography and Spectrophotometric Determination Using 2-Cyanoacetamide

Abstract

A simple procedure for separation of ribose-5-phosphate, deoxyribose-1-phosphate and ribose-1-phosphate is based on high performance liquid chromatography using reversed phase 4 × 300 mm “μ Bondapak/NH₂” column. The column is equilibrated with 0.13 M borate buffer (pH 7.5) followed by gradient elution of ribose-5-phosphate, deoxyribose-1-phosphate and ribose-1-phosphate using water, 0.05 M borate buffer containing 0.1 M MgCl₂ (pH 9.6) and 0.05 M sodium acetate-acetic acid buffer containing 0.1 M MgCl₂ (pH 5.0) as eluants respectively. Eluates of borate complex “μ Bondapak/NH₂” column are brought to pH 9.6 by the addition of 1 N KOH and enzymatically hydrolysed with alkaline phosphatase (EC 3.1.3.1) to release the free pentoses. The free pentoses are mixed with a reagent solution prepared from aqueous solution of 2% cyanoacetamide and 0.6 M borate buffer (pH 9.6), and the mixture is boiled for 10 minutes and the absorbance of the product is measured at 276 nm using a spectrophotometer.     

Review of Sub-Microliter (Nanoliter) Injection Techniques in Liquid Chromatography

Abstract

In liquid chromatography with “low-dispersion methods”, there is an increasing need to reproducibly inject nanoliter sample volumes. Low-dispersion methods produce very narrow peaks because of short column length, narrow column bore, small particle packing, low particle surface area, open tubular configuration, or combinations of these parameters.

This paper reviews methods used for sub-microliter injections for use with these low-dispersion liquid chromatography methods. Some nanoliter injection methods for open-tubular electrophoresis are also described.     

The Effect of Silanol Masking on the Recovery of Picloram and Other Solutes from a Hydrocarbonaceous Pre-analysis Extraction Column

Abstract

The recoveries of picloram, picloram-methylester, hexazinone, benzene, and acetophenone from aqueous samples were studied using a commercially available hydrocarbonaceous pre-analysis extraction cartridge, both with and without tetrabutylammonium hydrogen sulfate (TBAHS) in the eluent. Extraction efficiency was found to be dependent on sample loading volume. The results suggest a mixed mechanism of retention involving both “silanophilic” and “hydrophobic” interactions in the absence of tetrabutylammonium ion. The ability of TBAHS to mask surface silanol groups and/or ion-pair with counterionic solutes is invoked to explain the observations. Chromatograms of the solutes obtained on a C₁₈ bonded analytical column in both the presence and absence of TBAHS are also presented.     

Enzymatic Synthesis of an RNA Fragment Corresponding to the Anti-Codon Loop and Stem of tRNAPhe from Yeast

Abstract

A hexadecamer corresponding to the anticodon loop and stem of tRNA^Phe yeast has been prepared using T₄ RNA ligase and isolated by high performance liquid chromatography. The two oligonucleotides used in the ligation were isolated from a ribonuclease T₁ digest of the tRNA which was resolved by HPLC on an anion exchange column. To prepare the “acceptor” oligonucleotide for the RNA ligase reaction a 3′ terminal phosphate was removed. To prepare the “donor” oligomer a 5′ terminal phosphate was added. Analysis of the product hexadecamer was by nucleoside and nucleotide-3′-mono-phosphate composition.     

Determination of Some Bipyridylium Herbicides by Ring-Oven Techniques

Abstract

The development of a rapid and a simple “Weisz ring-oven” method is described for the semiquantative determination of diquat and paraquat. In this method column chromatography is used for the separation of herbicides from water and soil samples. Further evidence of the identity of individual residues is provided by thin layer chromatography.     

Theory of Open Capillary Chrcmatography Using Electrical Diffuse Layer as the Virtual Stationary Phase for Charged Macro Mxecular Separations

Abstract

Electrical diffuse layers established by arrangement of ions along a metal capillary column can be used as the virtual “stationary phase” for charged macromolecular separations and determinations. Preliminary studies have indicated that resolution depends on solvent flow rate, pH, column diameter, etc., as well as the applied potential. In this communication we report a theory which underlies the mechanism of retention of charged macro-molecules, e.g., proteins, on the column. The utility and difficulties of this novel fractionation technique will also be discussed.     

Comparison of Two Novel Techniques for Eliminating Liquid Chromatography Gradient Ghost Peaks

Abstract

Distilled water to organic gradients in reversed phase liquid chromatography are often plagued with UV-detectable “ghost” peaks that can obscure sample peaks and complicate interpretation of results.^3,4 These contaminants usually come from the distilled water³ although it is possible for them to originate in the organic eluent⁴. The ghost-peak problem can be eliminated by using specially prepared “LC-grade” water, although this is expensive. In addition water can be cleaned with the Milli-Q ion-exchange system, containing a carbon absorption column⁵ but not where these systems contain buffers or organic amine eluting agents such as triethylamine phosphate. Gurkin and Ripphahn have shown that a C-8 silica column can be used off-line in a low- pressure mode to clean water⁶.     

Cleanup of Samples for High Performance Chromatography and Instrumental Analysis

Abstract

Cleanup methods have been developed which allow utilization of modern chromatographic techniques and instrumental readouts. A cellulose cleanup for organo-phosphorus pesticides and carbamates has already been reported. A particulate carbon column has been described for Rumensin. These columns maybe combined to improve the cleanup for organophosphorus and carbamate pesticides. This same column cleanup maybe used for Ochratoxin. Powder charcoal maybe used for the mycotoxins aflatoxin, and zearalencne. It maybe used for T-2 toxin and trichothecenes with some loss (50-75% recovery).

The thin layer plates themselves provide a background reference for spectroscopy. This background maybe removed by “annealing” the plates at 400°C for 2 hours.

Sweepco distillation maybe used to cleanup biological samples so that carbamates and organophosphorus pesticides maybe analyzed by thin layer chromatography. Trichothecenes mycotoxins maybe cleanuped with about 40% loss.     

Experimental Tests of Parity Violation at Helical Polysilylene Level

The author would like to announce the following corrections: On page 671, right column, line 2, the sentence should read “For a certain polysilylene, ΔE_ST is reported to be 0.32 eV”. The insets in Figure 3d should read “S‐1” and “R‐1” instead of “S‐1L” and “R‐1L” . In the reference section, [34] reads correctly “Y. Ishimaru, S. Sumida, T. Iida, Chem. Commun. 1997 , 2187‐‐2188”.     

An Improved Method Fòr Extraction,Partial Purification,Separation and Isolation of Chlorophyll from Spinach Leaves

Abstract

Powdered sugar column chromatographic separation and isolation of chlorophyll were studied. A wet column was employed using a Pharmacia R25 chromatographic tube connected, through a UV monitor, to a fraction collector. Chlorophyll extracted from spinach leaves with acetone was selectively precipitated with dioxane and water, and then was washed with 80% (v/v) aqueous methanol. Use of the further purified chlorophyll as a starting material for chromatographic separation and isolation of chlorophyll, made column chromatographic procedures become extremely effective and easier. Thin-layer chromatographic and high-performance liquid chromatographic tests revealed that chlorophyll preparations in this study did not contain any other photosynthetic pigments and their degradation products.     

NEW HETEROCYCLIC SKELETONS DERIVED FROM THE APORPHINE ALKALOID BOLDINE

ABSTRACT

The abundant aporphine alkaloid (S)-(+)-boldine (1) was selectively nitrosated with sodium nitrite in acetic acid affording 8-nitrosoboldine (2) which was hydrogenated catalytically to give 8-aminoboldine (3). The latter was used as the starting material for annulations with ethyl ortho-formate to afford the corresponding oxazole (“boldine-9,8-oxazole”, 4), and with methyl benzoylformate giving the phenyl-oxazinone (“boldine-9,8-phenyloxazinone”, 5). This later product was treated with KOH/EtOH at room temperature and converted quickly into the ring-contracted phenyloxazole (“boldine-9,8-phenyloxazole”, 6) in moderate yield.     

Supramolecular Influences on Metal Ion Coordination: Lead(II) under Eight-coordination

Crystal structure determinations on the “acid salt” of Pb(II) and dipicolinic acid (pyridine-2,6-dicarboxylic acid, dipicH²), regarded as composed of dimer entities, [Pb²(dipic)²(dipicH²)²(OH²)⁶], 1, infinitely cross-linked, and on the complex of 2,4-dinitrophenol (2,4-DNPH), a simpler coordination polymer, [Pb(2,4-DNP)²]n, 2, show both solid state structures to be influenced by π-stacking of the ligands. Although the Pb(II) coordination environment in 1 can be regarded as “hemidirected” and that in 2 as “holodirected”, it is argued that inter-dimer stacking interactions, rather than lone pair effects, are responsible for the distorted 9-coordination of Pb in 1. In 2, Pb adopts close-to-cubic 8-coordination.     

A simple and convenient protocol for the synthesis of seven- and eight-membered phosphorus heterocycles

A simple procedure for the synthesis of eight-membered 6-(2-chloroethyl)/bis(2-chloroethyl)-amino-12-oxo-dibenzo[d,g][1,3,2]dioxaphosphocin 6-oxides (3a–b) and seven-membered 6-(2-chloroethyl)/bis-(2-chloroethyl)aminodibenzo[d,f][1,3,2]dioxaphosphepin 6-oxides (5a–b) from cyclocondensation of equimolar ratios of 2,2′-dihydroxybenzophenone (1) and 2,2′-dihydroxybiphenol (4), respectively with 2-chloroethylphosphonicdichloride (2a) and bis(2-chloroethyl)phosphoramidic dichloride (2b) in dry toluene in the presence of triethylamine at 45–50 °C is described. All synthesized compounds possessed significant growth inhibition for their antibacteria against ‘Bacillus subtilis’ and ‘Klebsiella pneumonia’ and antifungi activity on “Curvularia lunata” and “Aspergillus niger.”     

On the metabolites produced by Colletotrichum gloeosporioides a fungus proposed for the Ambrosia artemisiifolia biocontrol; spectroscopic data and absolute configuration assignment of colletochlorin A

Ambrosia artemisiifolia L. is responsible for serious allergies induced on humans. Different approaches for its control were proposed during the COST Action FA1203 “Sustainable management of Ambrosia artemisiifolia in Europe” (SMARTER). Fungal secondary metabolites often show potential herbicidal activity. Three phytotoxins were purified from the fungal culture filtrates of Colletotrichum gloeosporioides, isolated from infected leaves of A. artemisiifolia. They were identified by spectroscopic and chemical methods as colletochlorin A, orcinol and tyrosol (1, 2 and 3). The absolute configuration 6’R to colletochlorin A was assigned for the first time applying the advanced Mosher’s method. When assayed by leaf-puncture on A. artemisiifolia only 1 caused the appearance of large necrosis. The same symptoms were also induced by 1 on ambrosia plantlets associated with plant wilting. On Lemna minor, colletochlorin A caused a clear fronds browning, with a total reduction in chlorophyll content.     

Separation of Proteins on Polar Bonded Phases by Hydrophobic Interaction Chromatography

Abstract

Hydrophobic interaction chromatography (HIC) with a polar bonded phase (“Acetamide”) developped for size exclusion chromatography (SEC) is described. Retention of proteins depends on the surface area of the stationary phase, the pH and ionic strength of the eluent. For efficient separation the pore diameter should be 25 nm or more. The surface area should be large to achieve retention even at low ionic strength. Separation is only possible with a gradient from high to low ionic strength. Gradient volumes of 10 empty column volumes with column lengths above 15 cm are recommended. Selectivity can be optimized via pH adjustment. The advantage of this column packing is its applicability for two different separation modes: SEC and HIC.     

Spherical Versus Irregular-Shaped Silica Gel Particles in HPLC

Abstract

With optimised packing procedures, spherical shaped silica gel particles produce 1.5 to 2 times more plates in HPLC than irregular shaped silica gel particles. The lowest reduced plate height obtained by us so far is for 5 μm ROSiL-C₁₈-HL-D and is h: 1.62 for k': 4.5. It is suggested to transform h into 100/h% and to name this the “Chromatographic efficiency”, or a % of the ideal 100% limit. This limit would be an h value equal to the mean particle diameter. Spherical and irregular silica gel particles of 5 and 10 μm particle diameter and with similar physical characteristics have the same permeability in HPLC columns.

Whether a correct column packing procedure is used can be shown by the constancy of plate number and column permeability in function of different packing pressures.     

High-Performance Liquid Chromatographic Behavior of 2-,4- and 6-Hydroxylated Bile Acid Stereoisomers

Abstract

High-performance liquid chromatographic behavior has been investigated for 40 stereoisomeric 2-, 4- and 6-hydroxylated bile acids, most of which possess a vicinal glycol structure, as their 4-nitrophthalimidomethyl (NPM) ester derivatives on a reversed-phase column using methanol-water systems as a mobile phase. The bile acid NPM esters were further derivatized to the so-called “mixed” acetonide-NPM esters, whereby the stereochemical relationships (α, α-and β,β-cis, diaxial trans or diequatorial trans) of 2,3-, 3,4-and 6,7-diol groups in the molecules were reflected to changes in the capacity factors of bile acids.     

A new fluorescence chemosensor based on benzothiazole derivative for Zn2+ and its logic gate behavior

A novel benzothiazole derivative J as a fluorescent chemosensor for Zn²⁺ has been designed and synthesized. The chemosensor J exhibits high selectivity and sensitivity to Zn²⁺ as fluorescence “turn-on” behavior, which the other cations do not make sense. The detection limit calculated by fluorescence titration method was 1.64 × 10^?8 M. Furthermore, the generated J-Zn²⁺ ensemble could recover the enhanced fluorescence upon the addition of EDTA generating an “on-off-on” recycle. In addition, the test strip based on J was fabricated, which could act as a convenient and efficient Zn²⁺ test kit.     

Analyse des composes fluores corrosifs par chromatographie gazeuse : Controle du fluor libre

A purely chromatographie and sufficiently general method is described for the analysis of molecular fluorine in permanent inert gases or in other corrosive fluorine compounds (UF₆, HF). The method is also applied to the analysis of permanent inert gases (even in trace quantities) in the same fluorine compounds. In this method, one or several chemical “precolumns” are placed in series with a gas-liquid chromatography column. The “precolumn” system retains or destroys the corrosive compounds (UF₆, HF, F₂) and the chromatography column separates the components formed.     

Reconstructed Mass Spectra,A Novel Approach for the Utilization of Gas Chromatograph—Mass Spectrometer Data

Abstract

Peak profile analysis of each of the few hundred mass chromatograms generated during a complete GC-MS experiment identifies all m/e values which maximize at any of the few hundred consecutive mass spectra recorded during the gas chromatogram. The resulting sets of data correspond to the mass spectrum of each eluting component (even very minor ones) practically free of the contributions of unresolved companion substances, tailing fractions, column bleed, etc. These “reconstructed mass spectra” are therefore more easily interpreted or automatically identified than the mass spectra originally recorded. A plot of the summed abundance of the ions that maximize at a given spectrum index number (“mass resolved gas chromatogram”) results in a gas chromatogram of dramatically improved apparent resolution.