S-(+)-naproxen chloride as acylating agent for separating the enantiomers of chiral amines and alcohols

Recently, the strongly fluorescent benoxaprofen (BOP) molecule was described as chiral fluorescence label. Also S-naproxen (NAP), like BOP a non-steroidal antiinflammatory drug from the group of 2-arylpropionic acids, can be used as derivatization reagent for primary and secondary amines and alcohols, if activated to the corresponding acyl chloride. The synthesis of derivatives as well as chromatographic systems for the separation of the diastereomeric products are described.     

Fluorescence label of alcohols and amines with pyrene-1-carbonyl fluoride]

Pyrene-1-carbonyl fluoride (PCF) was synthesized as a precolumn fluorescent labeling reagent for alcohols and amines for use in high performance liquid chromatography (HPLC). PCF reacted with primary and phenolic hydroxyl groups in dichloromethane at 100 degrees C for 30 min in the presence of 4-dimethylaminopyridine (DMAP) to give the corresponding fluorescent pyrene esters and also reacted with primary amines in acetonitrile at room temperature for 2 min in the presence of DMAP to give the amides. The PCF esters of corticosteroids were separated by normal-phase chromatography on a Cosmosil 5 SL column with hexane-ethyl acetate (6:5, v/v) and the PCF amides of primary amines were separated by reversed-phase chromatography on a TSK gel ODS-80 TM column with methanol-water (10:3, v/v). The detection limits (S/N = 3) of cortisone and 2-phenylethylamine were 600 fmol and 800 fmol for an injection volume of 10 microliters, respectively.     

Novel reactions using organoselenium compounds

Organoselenium compounds are important tools in the field of synthetic chemistry because their specific reactivities have been used as the key reactions in the synthesis of various organic compounds. Nucleophilic organoselenium reagents are frequently utilized to introduce a selenium atom into organic molecules. However, there is no method available for the direct transformation of a hydroxy group into a selenayl group, except for the Grieco-Nishizawa reaction. In this review, novel methods for the conversion of alcohols into selenides in one step are described: both selenolate-aluminum chloride and TMSSePh-aluminum bromide are effective reagent systems for this transformation. Although benzylic alcohols could be efficiently converted to the corresponding selenides, these reagent systems were not applicable to non-benzylic alcohols. On the other hand, the reaction of cinnamyl alcohol with TMSSePh-aluminum bromide afforded 4-phenylselenochroman as a main product. The scope and limitations of this selenochroman formation were investigated using several types of allyl alcohols. To clarify the reaction path, the reaction of cinnamyl phenyl selenide with aluminum bromide was carried out. As expected, 4-phenylselenochroman was obtained in high yield. The transformation of various cinnamyl selenides was also successful in generating the corresponding selenochroman derivatives. Consequently, we developed a novel method for selenochroman synthesis.     

Fluorimetric determination of aluminum traces in hemodialysis solutions using Mordant Red 19

A sensitive and accurate method for the spectrofluorimetric determination of trace levels of aluminum in hemodialysis solutions using Mordant Red 19 as the complexation reagent has been developed. The optimal experimental conditions for the concentration of fluorimetric reagent, pH, temperature, and the specific type of matrix are reported. The emission of the fluorescent metal chelate was measured at 555 nm, excitation at 478 nm. Linearity between emission intensity and aluminum concentration was found in the 2-20 ppb range in standard aluminum solutions. Limit of detection was 0.4 ppb. The aluminum amounts in some commercial hemodialysis solutions were determined by means of the extrapolation method. The proposed method proved to be suitable in terms of sensitivity and accuracy for the determination of aluminum in dialysis fluids.     

注射用丹参(冻干)中糖和糖醇类成分的定性定量分析

目的:研究建立注射用丹参(冻干)中糖和糖醇成分定性、定量分析的方法。方法:以l-甲基咪唑作溶剂和催化剂,盐酸羟胺和乙酸酐为肟化和乙酰化试剂,将注射用丹参(冻干)样品中的糖类成分乙酰化后,再利用GC-MS联用技术进行定性和定量分析。结果:对注射用丹参(冻干)中所含8个糖和糖醇成分进行了结构确证,并对其中6个成分的含量进行了测定,6个成分的线性关系良好(r≥0.9995),平均回收率在95.77%～104.4%之间。结论:本法操作简便、灵敏、准确,可用于注射用丹参(冻干)中糖和糖醇成分的质量控制。     

Evaluation of brilliance and visibility of fluorescence and chemiluminescence solution for training of preparing injections

Personnel who prepare and administer chemotherapeutic agents have been reported to develop untoward effects. The use of appropriate techniques for preparing these agents is encouraged, and educational training systems that involve the use of a fluorescent or chemiluminescence reagent as placebos have been established to minimize potential exposure to these agents. However, the optimum conditions for the use and visibility of these placebos remain obscure. In this study, our results indicated that the fluorescence intensity of fluorescent reagent decreased when it was used at a concentration greater than 0.01%. Because drops created due to splashes and leaks are extremely small and easily evaporate, it is possible that the fluorescence resulting from such drops readily disappears despite using an anti-evaporation reagent. We also developed a method to evaluate the visibility of the small drop; using this method, we determined the distance at which the drop present on the pin could be seen by the observer. The distance at which the drop was clearly recognized as a pinpoint by using the fluorescence method was almost comparable to that for the chemiluminescence method. In the chemiluminescence method, the drop on the pin was faintly visible as a slightly bright area because of low background when observed at a certain distance that was much greater than that at which the drop was clearly visible; however, such an area was not observed in the fluorescence method. The results of our study will help in the selection of a training method depending on the situation.     

Peroxidation of polyunsaturated fatty acid methyl esters catalyzed by N-methyl benzohydroxamic acid: a new and convenient method for selective synthesis of hydroperoxides and alcohols

A new catalytic system, based on N-methyl benzohydroxamic acid (NMBHA), was developed for the selective synthesis of lipid hydroperoxides and alcohols by aerobic oxidation of lipid precursors. The reactions were carried out under mild conditions with air at atmospheric pressure and 37 degrees C. The products were isolated in much better yields (50-60%) as compared to previously reported procedures. NMBHA can be recovered at the end of the reaction and recycled. The putative reactive intermediate, a nitroxyl radical, readily abstracts a hydrogen atom from polyunsaturated lipid precursors such as linoleate esters, and NMBHA itself is an excellent hydrogen atom donor, thus conferring high selectivity to oxidations catalyzed by this reagent.     

HPLC-fluorescence determination of amino acids in pharmaceuticals after pre-column derivatization with phanquinone

4,7-Phenanthroline-5,6-dione (phanquinone) was used as a fluorogenic labeling reagent in pre-column derivatization for the quality control of amino acids in pharmaceuticals. The amino acid adducts were efficiently separated by C12 RP high-performance liquid chromatography (HPLC) using a ternary mixture of triethylamine (TEA) phosphate buffer (pH 2.5, 0.05 M)-methanol- tetrahydrofuran (THF) as mobile phase by varying composition gradient elution and detected fluorometrically. The results obtained by the proposed method were compared statistically, by means of the Student's t-test and the variance ratio F-test, with those obtained by a rapid reference method, which involved o-phthaldialdehyde (OPA) as pre-column reagent; no significant difference was found. The stronger derivatization conditions (60 degrees C, pH 8, 60 min) required for the method with phanquinone are compensated by the major stability of derivatives and by the absence of fluorescent degradation products.     

聚苯乙烯荧光微球的制备及性能考察

目的制备一种应用于流式细胞术的绝对计数试剂。方法以乙醇为分散介质,聚乙烯吡咯烷酮(polyvinyl pyrrolidone,PVP)为分散剂,偶氮二异丁腈(azodiisobutyronitrile)为引发剂,采用分散聚合法,制备聚苯乙烯(polystyrene,PS)微球。以三氯甲烷和异丙醇为溶胀剂,吖啶橙(acridine orange,AO)和罗丹明101(rhodamine 101,R101)为荧光素,通过溶剂逐渐挥发法对PS微球染色。结果制备了粒径范围在1～5μm的单分散PS微球,染色后的微球在荧光显微镜下可观察到被激发出红、绿、蓝等多种颜色的荧光。结论制备的PS荧光微球荧光发射光谱宽,能满足流式细胞仪对不同荧光微球的检测。     

Application of 2-acetylbutyrolactone to spectrofluorimetry: fluorescence properties of Schiff bases derived from 2-acetylbutyrolactone and spectrofluorimetric determination of primary amine-containing compounds

2-Acetylbutyrolactone (ABL) has been characterized for use as a fluorogenic reagent for the spectrofluorimetric determination of primary amines. The reagent forms strongly fluorescent Schiff bases upon the reaction with primary amines in acid-catalized aqueous solutions or in dimethylformamide (DMF). Sulfamethoxazole (SMX) and ampicillin sodium (AMP Na) were used as model amines of type ArNH₂ and RNH₂, respectively. The reaction conditions, fluorescence spectral properties and the stability of the derivatives have been investigated. The chemistry and the pathway of the reaction have been discussed. Calibration data, accuracy, precision, limits of detection, limits of quantification and other aspects of analytical merit were presented in the text. The utility of ABL for the analysis of the model drugs in pharmaceutical preparations was demonstrated. The results indicated that the proposed methods are equally accurate and precise as the official or other reported methods.     

利用Pirkle型手性固定相柱拆分对映体药物:用异氰酸-α萘酯作衍生化试剂拆分胺类、醇类药物

本文首次介绍了利用异氰酸-α萘酯(NI)作衍生化试剂,在Pirkle型DNBPG柱上对五种含胺基、醇基的药物对映体进行了拆分。其中拆分美西律(mexiletine)、氯胺酮(ketamine)、本芴醇(benflumelol)和苯丙醇(phenylpropanol)药物尚未见报道。结果表明,用异氰酸-α萘酯作衍生化试剂的方法,具体简便易行、分离度好、几乎没有衍生化带来的杂质、不需分离出未反应的异氰酸-α萘酯便可直接进行拆分实验等优点。可以预计,NI将可能成为一种很有发展前途的新衍生化试剂,而且为Pirkle型DNBPG柱开发了新的领域。     

A method for determination of Ukrain in blood plasma for monitoring and pharmacokinetic study

We developed a method using high-performance liquid chromatography for the determination of the main fluorescent component of Ukrain, a novel antitumor and immune-stimulating drug. Our method was based on ion-pair separation of Ukrain from perchloric acid extracts using reversed-phase column, buffer with high molarity (0.5 M potassium phosphate, pH 2.65), high concentration of ion-pair reagent in the mobile phase (10 mM octylsulfonic acid), controlled temperature of the separation (45 degrees C) and detection by fluorescence (360/455 nm). Under the above conditions a peak of the main Ukrain compound was resolved from fluorescent peaks of the sum of alkaloids of Chelidonium majus L. although several peaks of alkaloids were retained in Ukrain as traces. The height of this main peak was nearly constant, while the alkaloid peaks varied depending on the series of the preparation; chelidonine and thio-triethylenephosphoramide gave no peaks. Analytical recovery for Ukrain from human plasma was 98.0 +/- 4.5%. Therefore, Ukrain possesses neither significant stable binding to plasma proteins nor adsorption in blood cells.     

一种新的异氰酸酯类手性衍生化试剂用于药物对映体高效液相色谱拆分性能的研究

A new chiral derivatizing reagent (s) -α-methoxybenzyl isocyanate (MIB) was prepared from (s)-(+)-mandelic acid with a practical method, and its ability of liquid chromatographic rasolution for drug enantiomers by pre-column chiral derivatization was evaluated with racemic amphetamine and mexiletine as well as phenylpropanol. The operation is simple and rapid. Separation is good for the amine isomers, but poor for alcohols and extensive work remains to be done. The solution of derivatization was directly injected into the chromatograph. Conventional silica gel column was used with light petroleum as the mobile phase.     

Chemical discrimination of compounds possessing an amino group

Cyclic 1,3-amino alcohols have been successfully resolved via lipase-catalysed O-acylation of their Z derivatives, using vinyl butyrate as acyl donor in different ether solvents. A direct HPLC method was developed for the monitoring of the resolutions. Most of the screened lipases preferred the S enantiomer. Both alcohol and ester enantiomers were obtained in ee > 90% after gram-scale resolutions. A new isothiocyanato-type CDA, (1S,2S)- or (1R,2R)-1,3-diacetoxy-1-(4-nitrophenyl)-2-propyl isothiocyanate [(R,R)- or (S,S)-DANI], was designed for the indirect HPLC enantioseparation of primary and secondary amino compounds. The new reagent can be readily synthetized in optically pure form (> or = 99.5%) after a straightforward two-step synthesis. It is stable chemically and stereochemically both in the solid and in the solution phase. It is available in both enantiomeric forms, allowing the appropriate selection of the elution sequence. The kinetics of derivatization was studied in detail for alpha-amino acids and for beta-blockers. The quantitativeness of reactions was ensured by optimization of reaction conditions (pH, reagent excess, temperature). As concerns the HPLC analyses of the thiourea derivatives formed, significant differences were found between selectivities of the organic modifiers applied; in most cases, MeOH proved much more effective than MeCN with respect to both retention times and resolutions. In conclusion, the new CDA was capable of the resolution of most of the investigated analytes.     

Liquid chromatographic determination of ivermectin in animal plasma with trifluoroacetic anhydride and N-methylimidazole as the derivatization reagent

Ivermectin is a potent anthelmintic agent which was detected at low concentrations in cattle plasma by LC after conversion to a fluorescent derivative. This was accomplished by reaction with acetic anhydride (AA) and pyridine for 24 h at 100 degrees C or with AA and N-methylimidazole (NMIM) for 1 h at 95 degrees C. Substituting trifluoroacetic anhydride (TFAA) for AA reduced the reaction time to less than 30 s at 25 degrees C, yielding an intensely fluorescent derivative with substantially fewer reagent by-products. The need for further sample preparation after derivatization with TFAA-NMIM was thereby eliminated, and detection limits of less than 20 pg ml-1 ivermectin could be achieved with 1 ml of plasma by a considerably simpler analytical procedure.     

Cellular uptake,stability, visualization by ‘Naturstoff reagent A’, and multidrug resistance protein 1 gene-regulatory activity of cyanidin in human keratinocytes

There is increasing interest in the role of anthocyanidins as potential skin protective phytochemicals. However, little is known if and to what extent anthocyanidins are taken up by the human skin. In the present study cellular uptake (as determined by HPLC), stability, and gene-regulatory activity of cyanidin were determined in human HaCaT keratinocytes in culture. Using the fluorescent dye Naturstoff reagent A cyanidin was visualized in order to determine its cellular accumulation via flow cytometry and fluorescence microscopy.Cyanidin was rapidly taken up by HaCaT cells at relatively low concentrations. Following incubation, cellular cyanidin levels decreased time-dependently most likely due to degradation into protocatechuic acid and phloroglucinol aldehyde. Confocal laser scanning microscopy data demonstrated that cyanidin was mainly present in the cytoplasm. Cellular uptake of cyanidin was accompanied by an inhibition of multidrug resistance protein 1 (involved in cellular efflux of flavonoids) mRNA-levels indicating its gene-regulatory activity.Naturstoff reagent A seems to be a promising fluorescent dye to visualize cyanidin in keratinocytes.     

Activation of environmental carcinogenic N-nitrosodialkylamines by model systems for metabolic oxidation

Nitrosamines are environmental carcinogens, and their relevance to human cancer is highly suspected. Dialkylnitrosamines require activation through metabolizing enzymes before they become ultimate electrophilic active species. This review summarizes two model systems for metabolic oxidation of dialkylnitrosamines. One system utilizes porphyrin and oxidant as a model for shunt pathway in the metabolizing pathway of cytochrome P450, and the other one utilizes Fenton reagent. Porphyrin and oxidant activated nitrosodialkylamines into direct acting mutagen by releasing aldehydes. During the process the alkylating activity was observed and alcohols are formed from the alkylation of water. Fenton reagent, consisting of iron salt and hydrogen peroxide supplemented with copper salt, activated dialkylnitrosamines into mutagens of a novel type containing an oxadiazine ring as their proposed structures. These works with model systems open a new aspect into the elucidation of the mechanism of xenobiotic metabolism.     

Reaction of 11C‐benzoyl chlorides with metalloid reagents: 11C‐labeling of benzyl alcohols,benzaldehydes, and phenyl ketones from [11C]CO

In this article, we describe the carbon‐11 (¹¹C, t_1/2 = 20.4 minutes) labeling of benzyl alcohols, benzaldehydes, and ketones using an efficient 2‐step synthesis in which ¹¹C‐carbon monoxide is used in an initial palladium‐mediated reaction to produce ¹¹C‐benzoyl chloride as a key intermediate. In the second step, the obtained ¹¹C‐benzoyl chloride is further treated with a metalloid reagent to furnish the final ¹¹C‐labeled product. Benzyl alcohols were obtained in moderated to high non‐isolated radiochemical yields (RCY, 35%–90%) with lithium aluminum hydride or lithium aluminum deuteride as metalloid reagent. Changing the metalloid reagent to either tributyltin hydride or sodium borohydride, allowed for the reliable syntheses of ¹¹C‐benzaldehydes in RCYs ranging from 58% to 95%. Finally, sodium tetraphenylborate were utilized to obtain ¹¹C‐phenyl ketones in high RCYs (77%–95%). The developed method provides a new and efficient route to 3 different classes of compounds starting from aryl iodides or aryl bromides.     

Determination of phenylenediamines in hair colors derivatizated with 5-(4, 6-dichlorotriazinyl)aminofluorescein via micellar electrokinetic chromatography

Phenylenediamines (PDs), which are reported to cause allergic dermatitis and possess genotoxicity and carcinogenicity, are the ingredients used in permanent hair dyes. The fluorescent derivatization strategy coupled with micellar electrokinetic chromatography (MEKC) were established to analyze four PDs, including o-phenylenediamine (OPD), m-phenylenediamine (MPD), p-phenylenediamine (PPD) and toluene-2,5-diamine (PTD). Additionally, 5-(4, 6-dichlorotriazinyl) aminofluorescein (DTAF) was used as a fluorescent reagent derived at amino groups of PDs and underwent nucleophilic substitution reaction to improve the detection sensitivity. The derivatization condition reacted at 90 °C for 10 min in alkaline conditions. The optimized separation conditions were 20 mM borate (pH 8.0) containing 10 mM Brij 35 and 35% (v/v) methanol. The limits of detection (S/N = 3) for MPD, PTD, PPD and OPD were 25, 25, 50 and 100 nM, respectively. Compared to MEKC-UV, the sensitivity enhancements were 30- to 81-fold when PDs were derived with DTAF. The high-sensitivity MEKC-LIF method was successfully established and applied to determine PDs in commercial hair colors for quality control and in real hair samples for evaluating the location of PDs in dyed hair samples, as well as in percutaneous absorption samples for evaluating the ability of PDs to penetrate skin.     

Fluorometric determination of gastric acidity with 2-fluorenecarboxaldehyde hydrazone

Fluorenecarboxaldehyde hydrazone (FCH) was synthesized as a new fluorescent reagent for the determination of acidity. FCH had no fluorescence in itself, however, it strongly fluoresced under acidic conditions at excitation maximum of 392 nm and emission maximum of 447 nm, respectively. It showed good correlation with pH in the range from pH 0.60 to pH 3.60 in strong acids. As an application, the acidity of gastric juice in rats and humans was determined. In comparison with pH-metry, the acidity measured by the developed method showed generally increased values of about 0.7–1.3 unit and 0.8–1.2 unit in rats and humans, respectively. However, these results had statistically close correlation with those of pH-metry and the correlation coefficients were 0.806 and 0.887 in rats and humans between two methods.