Comparison of the retention behavior of β-blockers using immobilized artificial membrane chromatography and lysophospholipid micellar electrokinetic chromatography

We have demonstrated that significant differences exist between the retention of eight β-blockers analyzed with immobilized artificial membrane (IAM) and lysophospholipid micellar electrokinetic capillary (LMEKC) chromatographic methods. The general retention trends are maintained with highly hydrophilic compounds such as atenolol eluting first and more hydrophobic compounds such as propranolol eluting last. The retention order, however, is different and would result in major ranking differences. LMEKC demonstrates a better correlation with liposomal partitioning (R²=0.95) than does IAM chromatography (R²=0.60). LMEKC, with its higher efficiency, can allow a more specific evaluation of lipophilicity than IAM chromatography and is useful in the analysis of pharmaceutical candidates, particularly for ranking purposes.     

Simple and rapid high-performance liquid chromatographic method for endogenous alpha-tocopherol determination in human plasma

A simple and rapid reversed-phase high-performance liquid chromatographic method was developed and validated for the determination of endogenous alpha-tocopherol in human plasma. Following addition of alpha-tocopheryl acetate as the internal standard, the plasma was deproteinized using acetonitrile and isopropanol mixture prior to HPLC analysis. Methanol was used as the mobile phase and the effluent was quantitated at 292 nm. By this developed method, the concentrations of alpha-tocopherol were linearly related to their responses in the range of 0.8-30 microg/mL. The relative standard deviations intra-day and inter-day for alpha-tocopherol in plasma were less than 10%. The percentage of bias was within +/-4%, which confirmed the accuracy of the method. The method has been successfully applied for determining endogenous alpha-tocopherol in healthy Thai male volunteers.     

Analysis of alkannin derivatives from Alkanna species by high-performance liquid chromatography/photodiode array/mass spectrometry

Alkannin, shikonin (A/S) and their derivatives are enantiomeric hydroxynaphthoquinone red pigments found in the roots of almost 150 species of the Boraginaceae family. A/S have been shown to exhibit strong wound healing, antimicrobial, anti-inflammatory and antioxidant activities and recent extensive research has well established their antitumor properties. A/S and their derivatives comprise the active ingredients of several pharmaceutical and cosmetic preparations. Although A/S have been efficiently synthesized and have been produced by cell tissue cultures in high yield, most of the pharmaceutical preparations worldwide contain A/S extracted from the roots of Boraginaceous species, found in nature. In the present study, a high-performance liquid chromatography/photodiode array/mass spectrometry (HPLC/PDA/MS) method was established to identify monomeric hydroxynaphthoquinones of the alkannin series and other metabolites from Boraginaceous root extracts. This method can be applied for the identification of alkannin derivatives and other metabolites from Boraginaceous cell cultures, and also to determine active ingredients in pharmaceutical preparations containing A/S derivatives. A phytochemical investigation of the Alkanna genus grown in Greece was also performed. Fifty-three root samples belonging to 10 species of the genus Alkanna (A. calliensis, A. corcyrensis, A. graeca, A. methanaea, A. orientalis, A. pindicola, A. primuliflora, A. sieberi, A. stribrnyi and A. tinctoria) were collected from several regions of the Greek flora and analyzed for their constituent hydroxynaphthoquinones and other metabolites. In most of the above Alkanna samples tested, the main hydroxynaphthoquinones were determined to be beta,beta-dimethylacrylalkannin, isovalerylalkannin + alpha-methyl-n-butylalkannin and acetylalkannin. The hydroxynaphthoquinone constituents and their proportions were found to vary among Alkanna species. Unknown metabolites (not monomeric hydroxynaphthoquinones) were detected by HPLC-PDA-MS, while in several Alkanna species hydroxynaphthoquinones were detected for the first time.     

Beta2-agonist extraction procedures for chromatographic analysis

Normally, different procedures were necessary to prepare sample matrices for chromatographic determination of β₂-agonists. The present review includes sampling, pre-treatment and extraction/purification for urine, plasma, liver, meat, feeds, hair and milk powder, as previous steps for chromatographic analysis of β₂-agonists. Six methodologies were especially revised for extraction/purification namely, liquid–liquid extraction, solid-phase extraction (SPE), matrix solid-phase dispersion, immunoaffinity chromatography, dialysis and supercritical fluid extraction. SPE was discussed in detail and five mechanisms were described: adsorption, apolar, polar, ion-exchange and mixed phase. A brief conclusion in this field was also outlined.     

Analysis of wine phenolics by high-performance liquid chromatography using a monolithic type column

A polyacrylamide gel crosslinked with allyl-β-cyclodextrin can be used repeatedly for several weeks for the separation of DNA fragments, since bubbles are not generated during a run. Allyl-β-cyclodextrin can easily be synthesized in one step from allylglycidylether and β-cyclodextrin. The plate numbers for DNA fragments, up to about 1500 bp, are high: for the separation of pBR322/HaeIII fragments they were in the range 450 000–1 600 000 m⁻¹. The resolution was almost independent of the concentration of the crosslinker (allyl-β-cyclodextrin) — in sharp contrast to gels crosslinked with N,N′-methylenebisacrylamide.     

On the conformation of the propranolol molecule

The structure of the propranolol molecule has been optimized within the AM1 and PM3 semiempirical framework followed by ab initio HF/6-31G^* refinement. On each calculation level the conformational space was sampled to search for the lowest-energy conformer(s) from among a few hundreds of conformers at the semiempirical step and next from among a few dozens of conformers at the ab initio level. Finally, five stable conformers were found; each stabilized by one or two of the three possible hydrogen bonds. The geometrical and electronic parameters were established and found to differ only slightly in the structures with the hydrogen bond either present or not.     

Development and optimisation of a single extraction procedure for the LC/MS/MS analysis of two pharmaceutical classes residues in sewage treatment plant

A unique extraction procedure leading to the separation of 2 different pharmaceutical classes molecules has been developed and optimised by chemometric tools. From only one sampling, this analytical method allows the determination of 21 pharmaceuticals from corticosteroids and β-blockers classes. Performing the SPE on Oasis MCX (mixed-mode cation exchange), the sequential elution of each pharmaceutical class is achievable, allowing a high purity level of extracts as well as high recovery rates. Performing a unique sample preparation results in an important save of time. The extracts were then analysed by LC/MS/MS, using a Hibar Purospher Star column for β-blockers and an X-Bridge column for corticosteroids with formate buffer (pH 3.8)/AcN and water/AcN mobile phases, respectively. This work also includes a study of the chromatographic and mass spectrometric parameters in order to increase the analyte signal. The optimised SPE-LC/MS/MS method was then applied to environmental samples from sewage treatment plant (STP). β-Blockers and corticosteroids were detected, respectively, in concentrations up to 318 ng L⁻¹ (sotalol) and 174 ng L⁻¹ (cortisone), in STP influents. Moreover, both pharmaceutical classes have also been detected in STP effluents. As far as we know, this is the first paper reporting the detection of corticosteroids in environmental waters. The developed analytical method can be used in further studies to investigate the environmental contamination by these drugs.     

Evaluation of the interactions between lipids and γ-globulin protein at the air–liquid interface

The interactions between lipids (cholesterol, distearoylphosphatidylcholine, distearoylphosphatidylethanolamine and sphingomyelin) and the γ-globulin protein have been analyzed using the monolayer technique at the air–liquid interface. The analysis has been carried out using both state equations and an adequate thermodynamic formulation for the surface pressure (π)–molecular area (a) isotherms. Different parameters as the virial coefficients, have been estimated. For the uncharged lipid monolayers, the interactions between the molecules are of an attractive nature, at medium and long distance, and of a steric repulsive nature at short distance. At low surface pressures the lipid molecules form small domains. The net force between γ-Globulin molecules in the monolayers has been found to be attractive. Finally, it can be concluded that when the lipid monolayers are uncharged, there is practically no interaction between the protein and lipid molecules at the mentioned interface.     

5-雄烯-3β, 7α, 17β-三醇和5-雄烯-3β, 7β, 17β-三醇的合成

以5-雄烯二醇为原料,用微生物转化的方法合成了两个重要的神经甾体5-雄烯-3β, 7α, 17β-三醇和5-雄烯-3β, 7β, 17β-三醇。所用菌种总枝毛霉为我们自己筛选,并首次应用于5-雄烯-3β, 7α, 17β-三醇和5-雄烯-3β, 7β, 17β-三醇的合成中。     

A sensitive and selective liquid chromatographic/electrospray ionization tandem mass spectrometric assay for the simultaneous quantification of alpha-,beta-arteether and its metabolite dihydroartemisinin in plasma,useful for pharmacokinetic studies

A sensitive, selective, specific and rapid liquid chromatographic/electrospray ionization tandem mass spectrometric assay method was developed and validated for the simultaneous quantitation of alpha-,beta-arteether (alpha-,beta-AE) and its metabolite alpha-dihydroartemisinin (DHA) in monkey plasma using the propyl ether analogue of beta-arteether (PE) as an internal standard. The method involves a simple two-step liquid-liquid extraction with hexane. The analytes were chromatographed on a C(18) reversed-phase chromatographic column by isocratic elution with methanol-ammonium acetate buffer (pH 4) (92 : 8, v/v) and analysed by mass spectrometry in the multiple reaction monitoring mode. The chromatographic run time was 7 min and the weighted (1/x(2)) calibration curves were linear over the range 0.78-200 ng ml(-1). The method was validated in terms of accuracy, precision, absolute recovery, freeze-thaw stability, bench-top stability and re-injection reproducibility. The limit of detection and lower limit of quantification in monkey plasma were 0.39 and 0.78 ng ml(-1) respectively for all the analytes. The intra- and inter-batch precision and accuracy were found to be well within acceptable limits (<15%). All three analytes were stable even after three freeze-thaw cycles (deviation < 15%). The average absolute recoveries of alpha-,beta-AE, DHA and PE, used as an internal standard, from spiked plasma samples were 85.85 +/- 6.56, 70.10 +/- 7.06, 54.37 +/- 3.39 and 93.90 +/- 6.9%, respectively. The assay method described here could be applied to study the pharmacokinetics of alpha-,beta-AE and DHA in rhesus monkeys.     

Ultrasensitive assays of trans- and cis-β-carotenes in vegetable oils by high-performance liquid chromatography–thermal lens detection

The fact that β-carotene might be the protective factor against various cancers, suggests the need for a rapid reliable assay for this potential marker. We have proposed the method for selective, precise and simple profiling of carotenoids as well as for simultaneous ultrasensitive assaying of trans-β-carotene (TBC) and cis-β-carotene(s) (CBC) in five vegetable oils. The oil samples diluted 20 or 100 times were directly injected and analysed by means of the isocratic non-aqueous reversed-phase high-performance liquid chromatography (HPLC) combined with ultrasensitive thermal lens detection (TLS).

Elution of TBC was followed by CBC; both were identified and determined in olive, safflower, sesame, wheat germ and linseed oil by standard addition method. The presence of lutein/zeaxanthin, some xanthophyll and/or early eluting carotene may be also presumed in the aforementioned oils. The examined oils showed different and characteristic carotenoid/carotene profiles and characteristic TBC-to-CBC ratios. Both analytes were selectively detected in the presence of palmitic, oleic, linoleic, linolenic and stearic acid, β-sitosterol and --tocopherol. This extends applicability of the method to other vegetable oils as well.

Favourable analytical performances (high sensitivity, low limits of detection (LODs) and wide linearity span) enabled the direct analyses of highly diluted oils. This resulted in several major benefits of the proposed method, among which (i) reduced risk of stationary phase deterioration, (ii) avoiding the risk of carotenoids transformations and (iii) substantial labour and time savings. The TBC and CBC in diluted vegetable oils were reliably measured at ultratrace level (1–26 ng ml⁻¹) with the S/N ratio ranging from 4 to 140 and precisely determined (imprecision 0.4–8.3%). The concentrations of TBC+CBC estimated in the original oils were as follows: 90.5+51.2 ng ml⁻¹ in sesame oil, 146.0+164 ng ml⁻¹in safflower oil, 464.6+206.1 ng ml⁻¹in linseed oil, 453.7+143.3 ng ml⁻¹ in olive oil and 2.31+2.63 μg ml⁻¹ in wheat germ oil. The characteristic and variable portion of TBC within total β-carotene may serve as a reliable indicator of both, quality and authenticity of the vegetable oil. The HPLC–TLS assay proposed may therefore be successfully applied in nutritional, agricultural and epidemiological studies.     

Screening and diagnosis of beta-ureidopropionase deficiency by gas chromatographic/mass spectrometric analysis of urine

Dihydropyrimidine dehydrogenase (DHPDase), dihydropyrimidinase (DHPase) and beta-ureidopropionase (betaUPase) are the enzymes that catalyze the first, second, and third steps of the degradation of pyrimidines, respectively. beta-Ureidopropionate (betaUP) and beta-ureidoisobutyrate (betaUIB) are increased in the urine of patients with betaUPase deficiency. The original case in which betaUPase deficiency was discovered by NMR spectroscopy was an 11-month-old patient who presented with hypotonia and dystonic movement. We detected a second but asymptomatic case during a pilot study of neonatal screening with filter-paper urine, urease pretreatment and gas chromatography/mass spectrometry (GC/MS). The urease pretreatment of urine without fractionation resulted in a high recovery of these polar ureide compounds and allowed the highly sensitive GC/MS detection and diagnosis of betaUPase deficiency. betaUP and betaUIB were identified using GC/MS techniques. In the urine of the neonate with betaUPase deficiency, betaUP and betaUIB were persistently increased. Thymine, 5,6-dihydrothymine and 5,6-dihydrouracil were increased only moderately but significantly. It is known that thymine and uracil increase markedly in DHPDase deficiency, and 5,6-dihydrothymine and 5,6-dihydrouracil increase in DHPase deficiency. Therefore, betaUPase deficiency can be differentially diagnosed from the first and second enzyme deficiencies. Application of this specific and sensitive diagnostic procedure will lead to an understanding of the clinical heterogeneity of betaUPase deficiency. Furthermore, the identification of patients with defects in pyrimidine metabolism will enable doctors to avoid cancer chemotherapy with pyrimidine analogues such as 5-fluorouracil, which could be dangerous for these patients.     

Simultaneous determination of three naturally occurring estrogens in environmental waters by high-performance liquid chromatography

A simple, sensitive and accurate reversed-phase high-performance liquid chromatographic (HPLC) method for simultaneous determination of three naturally occurring estrogenic steroids including estrone (E1), 17β-estradiol (E2) and estriol (E3) in environmental water samples was developed. Analytes were extracted with ethyl acetate solvents and preconcentrated prior to HPLC analysis. Separations were accomplished in <20 min using a reversed-phase C(18) column (4.6×250 mm id, 5 μm) with a gradient elution of mobile phase containing 3.0 mM ammonium acetate/acetonitrile mixtures (flow rate, 1.0 mL/min). UV light absorption responses at 205 nm were linear over a wide concentration range from 100,000 μg/L to the detection limits of 0.96 μg/L E1, 0.64 μg/L E2 and 0.78 μg/L E3. Quantitation was carried out by the peak area method. The relative standard deviation for the analysis of three estrogens was <3.0%. This method was applied for the simultaneous determination of estrogens in environmental water samples collected in Zhejiang, China. The higher concentrations of both E2 and E3 were found in Tang River and West Lake waters, and E1 was detected in lake water only. All three estrogens were below the detection limits in rain waters.     

Structural determination of a new 2(3 --> 20)abeotaxane with an unusual 13beta-substitution pattern and a new 6/8/6-ring taxane from Taxus cuspidata

A new 2(3 --> 20)abeotaxane with an unusual 13beta-substitution pattern and a new 6/8/6-ring taxane were isolated from the methanol extract of the needles of Taxus cuspidata. The structures were established as 2alpha,7beta-diacetoxy-5alpha,10beta,13beta-trihydroxy-2(3 --> 20)abeotaxa-4(20), 11-dien-9-one (1) and 2alpha,5alpha,7beta,9alpha,13alpha-pentahydroxy-10beta-acetoxytaxa-4(20),11-diene (2) on the basis of 1D and 2D NMR spectral data and high-resolution FAB-MS analyses.     

Use of an amino-silica column for the high-performance liquid chromatographic analysis of synthetic oligodeoxy-nucleotides.

The use of an amino-silica column in the chromatographic analysis of synthetic oligodeoxyribonucleotides and their derivatives from different stages of oligonucleotide synthesis has been investigated. By eluting with 0.10 M potassium phosphate solution of pH 3.30, the nucleotide composition of oligonucleotides can be established within 15 min. In a linear gradient of phosphate buffer (0.10-0.75 M) at neutral pH, the separation of oligonucleotides by length and in an acidic medium pH 3.30-4.30) by composition is possible; the oligonucleotides may be in the free form or modified by the various protecting groups used in synthetic oligonucleotide chemistry. The analysis of some reaction mixtures from different stages of oligonucleotide synthesis and of a number of synthetic oligodeoxyribonucleotides and their derivatives has been performed.     

Synthesis of γγγ-trifluorocarbonyl compounds

γγγ-Trifluorocarbonyl compounds are easily obtained in a good yield by introduction of the 1,1,1-trifluoroethyl moiety (CF₃-CH₂-) on the -methylene group of a ketone.     

SmI2－Mediated Addition Reaction of α－Halomethylsulfones to Carbonyl Compounds. A Convenient Synthesis of β－Hydroxysulfones

Due to the chemoselective dehalogenation by SmI2, the addition of a-halomethylsulfones to carbonyl compounds afforded ,β-hydroxysulfones. Those reactions with α-bromomethylsulfones gave the products in moderate to good yields. The SmI2-mediated addition of gem-dihalomethylsulfones to ketones also afforded α-halo-β-hydroxysulfones in moderate yields.     

Facile Preparation of Homoallyl β′，γ′—Unsaturated Amines via 1,2—Addition of α，β—Unsaturated Imines with Allylsamarium Bromide

A series of homoallyl β′，γ′-unsaturated amines were synthe sized via 1,2-addition of α，β-unsaturated imines with allylsamarium bromide in excellent yields under mild and neutral conditions.