High-performance liquid chromatographic determination of forphenicine in mouse serum and muscle by pre-column fluorescence derivatization using 1,2-diamino-4,5-ethylenedioxybenzene as fluorogenic reagent

A simple and sensitive high-performance liquid chromatographic method has been developed for the determination of forphenicine in biological samples. Forphenicine in the deproteinized sample is converted by reaction with 1,2-diamino-4,5-ethylenedioxybenzene into a fluorescent derivative. The derivative is separated on a reversed-phase column (TSK gel ODS-120T) by isocratic elution with acetonitrile-30 mM phosphate buffer (pH 6.5) (5:1, v/v) and monitored fluorimetrically. The method allows the quantification of forphenicine in serum (100 microliters) and muscle (0.1 g) of mice dosed with forphenicine or forphenicinol. The limits of detection (signal-to-noise ratio of 3) are 7.35 pmol/ml in serum and 5.36 pmol/g in muscle. The distribution of forphenicine and forphenicinol in the mouse serum and muscle after oral administration of these compounds is also described.     

4,5-Diaminophthalhy drazide as a highly sensitive chemiluminescence reagent for α-keto acids in liquid chromatography

4,5-Diaminophthalhydrazide dihydrochloride is studied as a highly sensitive and selective chemiluminescence derivatization reagent for α-keto acids in liquid chromatography (LC). The reagent reacts selectively with α-keto acids in dilute hydrochloric acid to give derivatives which produce chemiluminescence by reaction with hydrogen peroxide and potassium hexacyanoferrate(III). The derivatives in the reaction mixture of eight biologically important α-keto acids are separated within 50 min by reversed-phase LC with isocratic elution, followed by chemiluminescence detection. The detection limits for the acids are in the range 4–50 fmol for a 20-μl injection.     

Fluorimetric determination of aromatic aldehydes with 4,5-dimethoxy-1,2-diaminobenzene

A sensitive fluorimetric method for the determination of aromatic aldehydes is described based on their reaction in dilute acid with 4,5-dimethoxy-1,2-diaminobenzene to give a compound which fluoresces intensely in alkaline solution. The fluorescence is stabilized by β-mercaptoethanol. The method is simple, selective for aromatic and arylaliphatic aldehydes, and sensitive; almost all the aldehydes can be determined at concentrations of 10^-8–10^-7 M.     

1,2-bis(4-methoxyphenyl)ethylenediamine as a fluorogenic reagent for reducing carbohydrates

Eight 1,2-diarylethylenediamines in the meso- or DL-form produce fluorescence when heated with reducing carbohydrates in an alkaline medium. Of the diamines, meso-1,2-bis(4-methoxyphenyl)ethylenediamine is the most favourable reagent for reducing carbohydrates, including 2- deoxy sugars, amino sugars and sialic acids. The reagent permits the fluorimetric determination of reducing carbohydrates at concentrations as low as 0.2–0.9 nmol ml^?1.     

Selective and sensitive determination of peptides using 3,4-dihydroxyphenylacetic acid as a fluorogenic reagent

A novel fluorescence (FL) reaction for N-terminal Gly-containing peptides has been developed using 3,4-dihydroxyphenylacetic acid (3,4-DHPAA). The reaction of the peptides with 3,4-DHPAA was carried out in borate buffer (pH 8.0) in the presence of sodium periodate at 37 °C for 10 min, and the FL was measured with a spectrofluorimeter at the excitation and emission wavelengths of 370 nm and 465 nm, respectively. The 3,4-DHPAA reagent generated particularly strong FL for peptides containing Gly at their N-termini. When various other bio-substances, such as amino acids, sugars, nucleic bases, nucleotides, and proteins, were reacted with 3,4-DHPAA, no FL was observed. Under optimized reaction conditions, the lower detection limit of 0.25 μmol L⁻¹ was obtained for the N-terminal Gly-containing peptides of Gly-Pro (GP) and Gly-Pro-Pro (GPP), which gave 3 times greater FL intensity than that observed for the reagent blank. The proposed reaction with 3,4-DHPAA as a fluorogenic reagent is selective and sensitive for the detection of N-terminal Gly-containing peptides, and therefore, this method could be a useful tool for the determination of these particular oligopeptides.     

5-Amino-4-sulfanylphthalhydrazide as a chemiluminescence derivatization reagent for aromatic aldehydes in liquid chromatography

5-Amino-4-sulfanylphthalhydrazide (ASPH) was synthesized as a chemiluminescence derivatization reagent for aromatic aldehydes in liquid chromatography (LC). Benzaldehyde, 4-tolualdehyde, 4-chlorobenzaldehyde, 4-formylbenzoic acid, 4-hydroxybenzaldehyde and vanillin were used as model compounds to optimize the derivatization conditions. This reagent, ASPH, reacts selectively with aromatic aldehydes in the presence of sodium sulfite and disodium hydrogenphoshite in acidic medium at 100 degrees C to give the corresponding highly chemiluminescent 2-arylbenzothiazole derivatives. The resulting derivatives generated intense chemiluminescence by reaction with hydrogen peroxide and potassium hexacyanoferrate(III) in alkaline solution. The ASPH derivatives of aromatic aldehydes were separated by reversed-phase liquid chromatography with isocratic elution, and detected chemiluminometrically after mixing with oxidizing agents. The detection limits (signal-to-noise ratio = 3) for aromatic aldehydes are in the range 0.2-4.0 fmol for a 20-microl injection volume. Currently, the method is not effective for aliphatic aldehydes because of interfering LC peaks.     

4-Carbomethoxybenzaldehyde as a highly sensitive pre-column fluorescence derivatization reagent for 9,10-phenanthrenequinone

9,10-Phenanthrenequinone (PQ) is harmful environmental pollutant that is detected in airborne particulates. The measurement of PQ in the air should be necessary to evaluate the potential adverse effects of PQ on human health. We have recently developed a determination method for PQ based on the fluorescence derivatization of PQ using benzaldehyde and ammonium acetate as a reagent. In this study, in order to obtain more sensitive and selective fluorescence derivatization reaction, we measured the fluorescence of the reaction mixture of PQ with 21 kinds of aromatic aldehydes in the presence of ammonium acetate. Among the tested aldehydes, 4-carbomethoxybenzaldehyde was found to be the best reagent in regard to fluorescence intensity and emission wavelength maximum. Based on the fluorescence derivatization with 4-carbomethoxybenzaldehyde, a highly sensitive chromatographic method was developed for the determination of PQ with the detection limit (S/N=3) of 1.2 fmol/injection.     

pH sensitive fluorogenic spray reagents I. General discussion and investigation of 1,2-dichloro-4,5-dicyanobenzoquinone

Summary Studies were done on the reaction between bromine vapour and sulfur-containing compounds such as pesticides and amino acids on silica gel TLC plates. Results indicate that the sulfur is oxidized and that acid is formed. A pH sensitive compound, 1,2-dichloro-4,5-dicyanobenzoquinone (DDQ), was found suitable as a fluorogenic spray reagent for mineral acids, yielding blue fluorescent spots on a non-fluorescing background. The combination of bromine vapour and DDQ spaying has enabled the in situ fluorescent detection of 37 out of 44 sulfur compounds examined. The detection limits of a number of these substrates were found to be smaller than 0.1 g.

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pH-empfindliche Fluoreszenz-Sprühreagenzien I. Allgemeine Diskussion und Untersuchung von 1,2-Dichlor-4,5-Dicyanobenzochinon

Zusammenfassung Untersuchungen wurden durchgeführt über die Reaktion von Bromdämpfen mit schwefelhaltigen Verbindungen vom Typ der Pestizide und Aminosäuren auf Kieselgel-Dünnschicht-Chromatogrammen. Die Ergebnisse zeigen, daß offenbar Schwefel oxidiert wird und in der Fleckenzone Säure entsteht. Die pH-empfindliche Verbindung 1,2-Dichlor-4,5-Dicyanobenzochinon (DDQ) eignete sich aus diesem Grund als Fluoreszenz-Sprühreagenz. Blau fluoreszierende Flecken wurden auf nicht fluoreszierendem Untergrund erhalten. Die Kombination von Bromdampf und DDQ als Sprühreagenz ermöglichte die in situ Lokalisierung von 37 aus 44 schwefelhaltigen Testsubstanzen. Die Empfindlichkeitsgrenze lag für einige dieser Stoffe unter 0,1 g pro Flecken.

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Reactif pulvérisable fluorogène sensible au pH I. Discussion générale et recherches sur la 1,2-dichloro-4,5-dicyanobenzoquinone

Résumé Des études ont été faites sur la réaction, dans des couches minces de silice, entre les vapeurs de brome et les composés contenant du soufre: pesticides, acides aminés. Les résultats indiquent que le soufre est oxydé et que de l'acidité apparait. Un composé sensible au pH, la 1,2-dichloro-4,5-dicyanobenzoquinone (DDQ), a été montré satisfaisant comme réactif pulvérisable devenant fluorescent au contact des acides minéraux; il donne des taches fluorescentes bleues sur fond non-fluorescent. La combinaison des vapeurs de brome et de la pulvérisation de DDQ a permis de détecter 37 composés soufrés sur les 44 étudiés. La limite de détection d'un certain nombre de ces substances est inférieure à 0,1 g.

     

1,2-Diarylethylenediamines as fluorogenic reagents for catecholamines

Twenty-eight 1,2-diarylethylenediamines were investigated in a search for highly sensitive fluorogenic reagents for catecholamines (norepinephrine, epinephrine and dopamine). They all react with epinephrine used as a model catecholamine under mild conditions (pH 6.5–6.8, 37–50°C) in the presence of potassium hexacyanoferrate (III) to produce fluorescence. Of the compounds tested, 1,2-bis(3,4-dimethoxyphenyl)ethylenediamine, 1,2-bis(4-methoxyphenyl)ethylenediamine and 1,2-bis (4-ethoxyphenyl)ethylenediamine, all in the meso-form, are the most sensitive for all the catecholamines. 1,2-Diphenylethylenediamine, 1,2-bis(4-methoxyphenyl)ethylenediamine and 1,2-bis (4-methylphenyl) ethylenediamine in the dl-form are more sensitive to dopamine than the corresponding compounds in the meso-form. Catecholamines can be determined at concentrations as low as 10–15 pmol ml^?1 by using those compounds as reagents. The reactions of other catechol compounds are reported. Other types of biologically important compounds do not interfere.     

1,2-Cycloheptanedionedioxime as a micro-gravimetric reagent for nickel

Summary 1,2-Cycloheptanedionedioxime (heptoxime), previously reported as a water-soluble gravimetric reagent for nickel, has been applied succesfully to the micro-gravimetric determination of this ion. The procedure, modified from the macro-determination, consists of precipitating nickelheptoxime in a micro-filter beaker and weighing the precipitate after drying at 110–120° C. The commonly encountered cations and metallic ions do not interfere or may be masked by complexing agents. The method has been tested on standard nickel solutions and standard steel samples.

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Zusammenfassung Über die Eignung des 1,2-Cycloheptandiondioxims (Heptoxim) als wasserlösliches Reagens für die gravimetrische Bestimmung von Nickel wurde in einer früheren Arbeit berichtet. Nun wird gezeigt, daß es sich auch für die Mikrobestimmung des Nickels eignet. Das Nickelheptoxim wird in einem Filterbecher gefällt und nach Trocknen bei 110 bis 120° C gewogen. Die gewöhnlich das Nickel begleitenden Metalle hindern die Bestimmung entweder nicht oder können durch Überführung in geeignete Komplexe zufriedenstellend maskiert werden. Das Mikroverfahren wurde an Nickellösungen bekannten Gehaltes und an Stahlproben des National Bureau of Standards geprüft.

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Résumé On a rapporté dans un travail précédent, la possibilité d'emploi de la cyclo-1,2 heptanedionedioxime (heptoxime) comme réactif hydrosoluble pour le dosage gravimétrique du nickel. On a montré maintenant qu'elle est convenable pour le microdosage du nickel. L'heptoximate de nickel est précipité au bécher-filtre et pesé après séchage à 110–120°. Les métaux qui accompagnent habituellement le nickel, ou bien ne gênent pas ou bien peuvent être masqués d'une façon satisfaisante. Le procédé microchimique a été essayé sur des solutions de nickel de titre connu ou sur des échantillons d'acier du National Bureau of Standards.

     

A highly sensitive fluorogenic chemodosimeter for rapid visual detection of phosgene

A highly sensitive chemodosimeter was identified from a panel of rhodamine derivatives for rapid and visual detection of phosgene with a detection limit of 50 nM triphosgene. Visual detection of gaseous phosgene with chemodosimeter absorbed paper strips was demonstrated.     

Fluorometric determination of aromatic aldehydes with 1,2-Diaminonaphthalene

A new fluorometric method for the determination of aldehydes is presented. 1,2-Diaminonaphthalene reacts with aldehyde in dilute sulphuric acid to give a compound which fluoresces intensely in alkaline medium. The fluorescences produced from aromatic aldehydes in this method are fairly characteristic of individual aldehydes and their intensities are generally higher than those of fluorescences from aliphatic aldehydes. The only interference is from 2-oxo acids. The method may be suitable for the determination of aldehyde in complex samples.     

Nickel-catalyzed 1,2-addition of arylboroxines to aromatic aldehydes

Development of Ni-Et-Duphos-catalyzed 1,2-addition of arylboroxines to aromatic aldehydes is described. The dramatic effect of boron reagent and phosphine ligand is observed. This method with a phosphine ligand allows asymmetric arylation of aromatic aldehydes (up to 78% ee).     

Rhodamine B amine as a highly sensitive fluorescence derivatization reagent for saccharides in reversed-phase liquid chromatography

6-Rhodamine B amine functions as a highly sensitive fluorescence derivatization reagent for mono- and oligosaccharides; it reacts with the reducing end of saccharides under acidic conditions. The fluorescent derivatives of five monosaccharides can be separated within 25 min by reversed-phase liquid chromatography with isocratic elution. The detection limits (S/N = 3) for mono-, di-, and oligosaccharides are 7-51, 13, and 9-35 fmol/20 microl injection, which correspond to analyte concentrations of 35-255, 65, 45-175 nM, respectively. We have applied this derivatization method successfully to the analysis of the components of oligosaccharides in glycoproteins (ribonuclease B and fetuin) following their acidic or enzymatic hydrolysis. The results from these analyses are in good agreements with the reported values established previously.     

Reaction of 1,2-Diamino-4,5-diphenylimidazole with 1,3-Diarylpropenones and Their Dibromo Derivatives

The reaction of 1,2-diamino-4,5-diphenylimidazole with 1,3-diarylpropenones and 1,3-diaryl-2,3-dibromopropenones gives dihydro- and heteroaromatic derivatives of 6-hydroxyimidazopyrimidines and also imidazopyrimidines which do not contain the 6-hydroxy group. It was found that the structure of the products depended on the conditions for carrying it the reaction.     

BINOL-Salen-catalyzed highly enantioselective alkyne additions to aromatic aldehydes

[structure: see text] The BINOL-Salen compound (-)-1 can catalyze the addition of both aryl- and alkylalkynes to aromatic aldehydes at room temperature with high enantioselectivity (86-97% ee). The conditions for this catalytic process are both mild and simple. Unlike most other BINOL-based catalysts, using ligand (-)-1 not only avoids heating or cooling but also does not require the addition of Ti(O(i)Pr)(4).     

1-Cyanoisoindole derivative as a highly sensitive fluorescence labelling reagent for steroidal primary alcohols in liquid chromatography

A sensitive and reactive labelling reagent for steroidal primary alcohols, m-(1-cyano-2-isoindole)benzoyl azide, was prepared from the corresponding carboxylic acid, which was synthesized from o-phthalaldehyde and m-aminobenzoic acid in the presence of potassium cyanide in one step. A hydroxysteroid, such as cortisol, can react with the reagent, when kept in benzene at 80 °C for 40 min, resulting in the formation of a urethane derivative which shows a single peak on the chromatogram and provides high sensitivity. The detection limit of the cortisol in reversed-phase chromatography is 80 fmol.     

Evaluation ofo-phthalaldehyde as bifunctional fluorogenic post-column reagent for glutathione in LC

Summary A method for the assay of various forms of glutathione in human plasma has been developed. The reduced form was measured after direct injection of deproteinized plasma with perchloric acid. The unbound oxidized forms were assayed in deproteinized plasma after reduction with dithiothreitol. The total amount of glutathione was determined by reduction before protein precipitation. Separation of reduced glutathione from other endogenous thiols was by ion-pair, reversed-phase, high-performance liquid chromatography. Post-column derivatization withortho-phthalaldehyde made the assay more selective than when using pyrenemaleimide as thiol fluorogenic reagent. The linear range was 0.02 to 2.0 g mL^–1 with good repeatability (relative standard deviation less than 5% for the lowest concentration measured).     

Fusaric acid as a novel proton-affinitive derivatizing reagent for highly sensitive quantification of hydroxysteroids by LC-ESI-MS/MS

A highly sensitive derivatization method for liquid chromatography (LC)-electrospray ionization (ESI) tandem mass spectrometry of dehydroepiandrosterone (DHEA), testosterone (T), pregnenolone (P5), and 17α-OH-pregnenolone (17-OHP5) was developed based on the use of fusaric acid as a reagent. DHEA, P5, and 17-OHP5 were rapidly and quantitatively converted to the 3-fusarate esters by treatment with fusaric acid and 2-methyl-6-nitrobenzoic anhydride. The positive ESI-mass spectra of the fusarate esters of each steroid were dominated by the appearance of [M + H]⁺ as base peaks. The fusarate derivatization of these steroids showed 17.6-fold (DHEA), 11.9-fold (P5), 3.3-fold (17-OHP5), and 1.8-fold (T) higher sensitivity to those of the corresponding picolinate derivatives in LC-selected reaction monitoring.