Preparation of monolithic silica column with strong cation-exchange stationary phase for capillary electrochromatography

A monolithic silica based strong cation-exchange stationary phase was successfully prepared for capillary electrochromatography. The monolithic silica matrix from a sol-gel process was chemically modified by treatment with 3-mercaptopropyltrimethoxysilane followed by a chemical oxidation procedure to produce the desired function. The strong cation-exchange stationary phase was characterized by its substantial and stable electroosmotic flow (EOF), and it was observed that the EOF value of the prepared column remained almost unchanged at different buffer pH values and slowly decreased with increasing phosphate concentration in the mobile phase. The monolithic silica column with strong cation-exchange stationary phase has been successfully employed in the electrochromatographic separation of beta-blockers and alkaloids extracted from traditional Chinese medicines (TCMs). The column efficiencies for the tested beta-blockers varied from 210,000 to 340,000 plates/m. A peak compression effect was observed for atenolol with the mobile phase having a low phosphate concentration.     

Acetogenic isoquinoline alkaloids: CXII. Separation and identification of dimeric naphthylisoquinoline alkaloids by liquid chromatography coupled to electrospray ionization mass spectrometry

The atropodiastereomeric dimeric naphthylisoquinoline alkaloids, michellamines A (1a), B (1b) and C (1c), together with their monomers, korupensamines A (2a) and B (2b), were investigated using electrospray ionization tandem mass spectrometry coupled to liquid chromatography (LC–ESI-MS–MS). From the spectra obtained, characteristic product ions were chosen to monitor the chromatographic separation achieved on an RP-18 column. Under acidic conditions required for chromatographic analysis, the monomeric alkaloids 2a and 2b yielded protonated molecules [M+H]⁺, while the dimers, the michellamines, exhibited doubly protonated [M+2H]²⁺ molecules. In addition, the coeluting alkaloids 1b and 2b were identified unambiguously by means of tandem mass spectrometry. Thus, together with the retention times of the alkaloids, the product ion spectra allowed us the identification of michellamines in the presence of their presumed biogenetic monomeric precursors. Application of the HPLC–MS–MS method successfully proved the enzymatic formation of michellamine C (1c) by in vitro dimerization of korupensamine B (2b).     

Phytochemical investigations and evaluation of antimutagenic activity of the alcoholic extract of Glycosmis pentaphylla and Tabernaemontana coronaria by Ames test

Chemical investigation of root bark of Glycosmis pentaphylla and stem bark of Tabernaemontana coronaria led to the isolation of three carbazole alkaloids glycozoline, glycozolidine and methyl carbazole 3-carboxylate, two furoquinoline alkaloids skimmianine and dictamine, an acridone alkaloid arborinine, three monomeric indole alkaloids coronaridine, 10-methoxy coronaridine and tabernaemontanine, and two dimeric indole alkaloids voacamine and tabernaelegantine B. Their structures were established by detailed spectral analysis. Mutagenic and antimutagenic potential of methanol extract of both plant materials were evaluated by Ames test against known positive mutagens 2-aminofluorine, 4-nitro-O-phenylenediamine and sodium azide using Salmonella typhimurium TA 98 and TA 100 bacterial strains both in the presence and absence of S9. Both the extracts were non-mutagenic in nature. Both the extracts of G. pentaphylla and T. coronaria exhibited significant antimutagenic activity against NPD and sodium azide for S. typhimurium TA98 and TA100 strains. The results indicated that the extracts could counteract the mutagenicity induced by different genotoxic compounds.     

Letter: Influence of experimental conditions on electrospray ionization mass spectrometry of ferrocenylalkylazoles

Influence of experimental conditions on electrospray/ionization (ESI) mass spectra of ferrocene derivatives FcCHRAz (Fc = eta5-C5H5-Fe-eta5- C5H4; R = H, Az = benzimidazole; R = Ph, Az = 2-methylimidazole) has been investigated. The spectra of all the compounds revealed [M]+*, product of its fragmentation [FcCHR]+ as well as products of ion/molecular interactions (protonated molecule [MH]+, binuclear ion [(FcCHR)2 Az]+, dimeric ion [M2]+* and its protonated form [M2H]+). Relative abundances of these ions appreciably (more than one order) depend on experimental conditions: analyte concentration, temperature of heated capillary, spray voltage, flow rate of mobile phase and polarity of solvents. Established correlations allow the selection of optimum experimental conditions for registration of ESI mass spectra, as required by the application. If an unknown compound is to be identified, it is necessary to operate by using polar solvents, small concentration, low temperature of heated capillary, high spray voltage and flow rates. There are high-intensity binuclear and protonated dimeric ions in mass spectra under other conditions. It can give rise to wrong interpretation of the structure of investigated compound. At the same time, for study of ion-molecular processes by ESI-MS it is necessary to use concentrated samples in non-polar solvents. In this case the dependence of reaction products yields on temperature and flow rate of mobile phase must be investigated.     

A study of the behaviour of some new column materials in the chromatographic analysis of Cinchona alkaloids

Summary A major problem in the HPLC analysis of alkaloids is the poor peak shape and consequently low resolution, due to the interactions of the basic alkaloids with the residual acidic silanol groups of most reversed phase materials. The performance of new packing materials specially designed for the separation of basic compounds has been studied using mobile phases without the special additives commonly applied in the analysis of alkaloids. Strongly basic Cinchona alkaloids were used as test compounds. Retention characteristics and selectivities of each material were studied, after mobile phase optimisation for the column. The influence of the major factors (nature and content of the organic modifier, pH value, salt concentration) affecting resolution was studied. The mobile phases were chosen so that they could be used in thermospray LC-MS. The addition of salts to the mobile phase improves separation but in general the modification of the mobile phase gave little change in selectivity. The performance of silicabased C₁₈ material proved superior to the polymer materials tested.     

A study of the behaviour of some new column materials in the chromatographic analysis of Cinchona alkaloids

Summary A major problem in the HPLC analysis of alkaloids is the poor peak shape and consequently low resolution, due to the interactions of the basic alkaloids with the residual acidic silanol groups of most reversed phase materials. The performance of new packing materials specially designed for the separation of basic compounds has been studied using mobile phases without the special additives commonly applied in the analysis of alkaloids. Strongly basic Cinchona alkaloids were used as test compounds. Retention characteristics and selectivities of each material were studied, after mobile phase optimisation for the column. The influence of the major factors (nature and content of the organic modifier, pH value, salt concentration) affecting resolution was studied. The mobile phases were chosen so that they could be used in thermospray LC-MS. The addition of salts to the mobile phase improves separation but in general the modification of the mobile phase gave little change in selectivity. The performance of silica-based C₁₈ material proved superior to the polymer materials tested.     

Development and validation of a reversed-phase HPLC method for determination of alkaloids from Papaver somniferum L. (Papaveraceae)

An HPLC method for the separation of six target alkaloids from Papaver somniferum L. (morphine, codeine, oripavine, thebaine, papaverine, and noscapine) was developed, optimized, and validated. The chromatographic behavior of these alkaloids was investigated using a reversed-phase chromatography at acidic and alkaline pH. The effects of ion-pairing agents, pH value of the mobile phase, concentration of the buffer components, mobile phase organic modifier, and column temperature were studied. Regardless of the large differences in their pKa values, all alkaloids were separated within a close retention window, and good peak shape was achieved for each of the six alkaloids. The proposed method has adequate selectivity, linearity, accuracy, precision, and reproducibility and is applicable for poppy straw.     

Secologanin,a Biogenetic Key Compound—Synthesis and Biogenesis of the Iridoid and Secoiridoid Glycosides

The monoterpene glycoside secologanin is a key intermediate in the biosynthesis of most indole, cinchona, ipecacuanha, and pyrroloquinoline alkaloids, as well as of simple monoterpene alkaloids. More than a thousand alkaloids are formed from secologanin in vivo; this represents almost a quarter of this large group of natural products. It is also the parent compound of the secoiridoids. Many of the compounds derived from secologanin display a high degree of biological activity and are employed as pharmaceuticals, e.g., the dimeric indole alkaloid leurocristine (vincristine) which is used very successfully in the treatment of acute leukemia. A knowledge of the biosynthesis and biological reactions of secologanin provides a sound basis for the biosynthesis-orientated classification of numerous natural products and the taxonomy of many plants. Secologanin and structurally related substances can be synthesized in a few steps by stereocontrolled photochemical and thermal cycloadditions. Its biomimetic reaction with amines and amino acids yields other natural products and compounds of pharmacological interest.     

Cimicifoetones A and B,Dimeric Prenylindole Alkaloids as Black Pigments of Cimicifuga foetida

Two dimeric prenylindole alkaloids with a unique indole‐benzoindolequinone skeleton, cimicifoetones A ( 1 ) and B ( 2 ), were isolated as black pigments from the rhizomes of Cimicifuga foetida . The structures were elucidated by spectroscopic methods including HRMS and 2D NMR, as well as single‐crystal X‐ray diffraction and computational chemical modeling techniques. Cimicifoetones A and B represented the first examples of dimeric indole alkaloids generated through [4+2] Diels–Alder cycloaddition between the prenyl side chain in one 3‐prenylindole and the aromatic ring in another. The two compounds showed promising anti‐proliferative activity on seven tumor cell lines with IC₅₀ values in the range of 1.36–21.09 μm . Flow cytometric and western blot analysis revealed that compound 2 induced cell apoptosis via death receptor‐mediated extrinsic and mitochondrial‐mediated intrinsic pathways.     

Determination of organic and inorganic mercury compounds by reverse-phase high-performance liquid chromatography after extraction of the compounds as their dithizonates

A sensitive and selective method is described for the simultaneous determination of inorganic and organic mercury compounds. The mercury compounds are extracted into toluene or chloroform with dithizone, and the dithizonates are separated by liquid chromatography on an ODS column. Complete resolution was obtained between methylethyl-, phenyl- and inorganic mercury with a mobile phase of THF/methanol (2:1) with 0.05 M acetate buffer pH 4 (62 + 38), containing 50 μM EDTA. The mercury chelates were detected spectrophotometrically at 475 nm. The detection limits were at the subnanogram level. The method is applicable to human urine, tap water and tomatoes.     

Chemical constituents of Glycosmis citrifolia (Willd.) Lindl. Structures of four new acridones and three new quinolone alkaloids

The structures of two new dimeric acridone alkaloids, glycobismine-D (1) and -E (2), having a novel linkage as binary acridones, three monomeric acridones, glycocitrine-IV (3), -V (4), and -VI (5), and three quinolone alkaloids, glycocitlone-A (6), -B (7), and -C (8) from Glycosmis citrifolia (Willd.) Lindl. (Rutaceae) have been elucidated by spectrometric studies.     

Voafrine A and Voafrine B,New Dimeric Indole Alkaloids from Cell Suspension Cultures of Voacanga africana Stapf

The structures of voafrine A and voafrine B, two novel dimeric indole alkaloids isolated from Voacanga africana Stapf cell suspension cultures, were established and the medium-dependent formation of both alkaloids was investigated.     

Comparison of peak shapes obtained with volatile (mass spectrometry-compatible) buffers and conventional buffers in reversed-phase high-performance liquid chromatography of bases on particulate and monolithic columns

Retention factor, column efficiency and asymmetry factor were recorded for nine basic compounds on a number of RP-HPLC columns using phosphate and a variety of (MS-compatible) volatile mobile phase buffers of acid and neutral pH, in order to assess any effects of the buffer on performance. With formic or acetic acid, some phases gave partial or complete solute exclusion effects (reduced or negative k) compared with results using phosphate buffers at low pH. Despite its possible suppression of mass spectrometer sensitivity, trifluoroacetic acid was useful in enhancing retention times of relatively hydrophilic protonated bases, due to ion-pair effects. Peak shape was relatively poor on some pure silica-based ODS phases at pH 7 compared with results at acid pH. At low pH and at pH 7, ammonium and potassium phosphate gave very similar k, but the former may be preferable due to its volatile cation. Improved peak shapes, attributed to superior silanol masking effects, were obtained with ammonium phosphate at pH 7, but not at acid pH. Ammonium acetate gave acceptable peak shape at pH 7, but due to very limited buffer capacity, poor results were obtained for solutes having a pKa close to the mobile phase pH. Due to its instability, ammonium hydrogen carbonate is not a viable alternative buffer at pH 7.     

Study of the selectivity,retention mechanisms and performance of alternative silica-based stationary phases for separation of ionised solutes in hydrophilic interaction chromatography

The separation of a mixture of neutral, strongly acidic and strongly basic compounds was studied in hydrophilic interaction chromatography using a bare silica phase, and bonded silica phases with diol, zwitterionic, amide and hydrophilic/hydrophobic groups. The mobile phase was acetonitrile–ammonium formate buffer at low pH. Differences in selectivity between these various columns indicate that the stationary phase cannot function merely as an inert support for a water layer into which the solutes partition from the bulk mobile phase. Attempts to fit the retention data to equations which describe either partition or adsorption mechanisms were inconclusive. Ion exchange was a significant contributor to the retention of ionised bases on all columns studied. Van Deemter plots indicated that the efficiency as a function of flow rate varied between the columns, which might be attributable in part to the presence of either monomeric or polymeric bonded phase layers.     

The effect of chaotropic mobile phase additives on the separation of selected alkaloids in reversed-phase high-performance liquid chromatography

The retention behavior of selected alkaloids from different classes was studied. The effect of chaotropic salts additives to the mobile phase on chromatographic parameters of protonated basic analytes was investigated on Zorbax Extend-C18 column. The influence of the type of salts and their concentration on retention, efficiency, peak symmetry and separation selectivity of investigated alkaloids was established. Buffered acetonitrile-water mobile phase was chosen because of significant retention of added liophilic ions due to strong dispersive pi-pi interactions. These conditions are responsible for great contribution of electrostatic forces in the retention of protonated bases. The addition of salt, such as hexafluorophosphate, perchlorate, trifluoroacetate leads to the increase in retention, efficiency and separation selectivity of examined analytes. The influence of added salts on increase in retention parameters could be expressed as follows: H2PO4- < CF3COO- < ClO4- < PF6-. This order is in agreement with ability of salts to "salting-in" effect according to Hofmeister series. Obtained chromatograms of alkaloids mixture illustrate suitability of chaotropic effect to improve their separation selectivity.     

Influence of inorganic mobile phase additives on the retention, efficiency and peak symmetry of protonated basic compounds in reversed-phase liquid chromatography

Inorganic eluent additives affect the retention of protonated basic analytes in reversed-phase HPLC. This influence is attributed to the disruption of the analyte solvation-desolvation equilibria in the mobile phase, also known as "chaotropic effect". With an increase of counteranion concentration analyte retention increases with concomitant decrease in the tailing factor. Different inorganic counteranions at equimolar concentrations affect protonated basic analyte retention and peak symmetry to varying degrees. The effect of the concentrations of four different inorganic mobile phase additives (KPF6, NaClO4, NaBF4, NaH2PO4) on the analyte retention, peak symmetry, and efficiency on a C8-bonded silica column has been studied. The analytes used in this study included phenols, toluene, benzyl amines, beta-blockers and ophthalmic drugs. The following trend in increase of basic analyte retention factor and decrease of tailing factor was found: PF6- > ClO4- approximately BF4- > H2PO4-. With the increase of the counteranion concentration greater analyte loading could be achieved and consequently an increase in the apparent efficiency was observed until the maximum plate number for the column was achieved. At the highest concentration of counteranions, the peak efficiency for most of the basic compounds studied was similar to that of the neutral markers. In contrast, the neutral markers, such as phenols, showed no significant changes in retention, efficiency or loading capacity as counteranion concentration was increased.     

Enantioselective Formal Syntheses of 11 Nuphar Alkaloids and Discovery of Potent Apoptotic Monomeric Analogues

Concise, scalable, and enantioselective formal syntheses of eight dimeric and three monomeric nuphar alkaloids were achieved, along with the construction of a stereochemically diverse collection of the first known monomeric analogues having apoptotic activity. The syntheses involved the development of highly enantioselective Brønsted acid catalyzed vinylogous Mukaiyama–Mannich reactions, which feature the unprecedented use of a supersilyl group to control the regio‐, enantio‐ and diastereoselectivity. Biological studies reveal that several of these novel nuphar analogues are even more potent than their dimeric natural product counterparts.     

Investigation on liquid chromatographic separation of basic compounds using silica column with aqueous/organic mobile phase containing triethylamine and acetic acid

A high-performance liquid chromatography (HPLC) method using silica column eluted with aqueous solvent mobile phase containing triethylamine (TEA) and acetic acid (ACH) at trace percentages was characterized for the analysis of basic compounds. The key mechanism of this system is ion-exchange accompanying interaction of silanol groups. The increase in the ACH concentration in the mobile phase minimizes the ionization of the silanol group, leading to reduced retention time. However, the greater extent of ionization of silanol caused by the increase of TEA concentration helps to retain basic compounds in the column. Further, the protonated TEA that is positively charged also competes for the ionized silanol group with basic compounds, resulting in the modification of retention time. On the other hand, the retention becomes longer with increasing proportion of either organic or aqueous solvent in mobile phase, and partial replacement of methanol with acetonitrile.     

邻苯二酚紫-示波计时电位法测定天然水中不同形态铝

报道邻苯二酚紫-示波计时电位法分别在酸性和碱性条件下直接检测天然水中的无机单核铝和总单核铝浓度。并用该法测定了水样中的总铝,有机单核铝和酸溶态铝,从而实现了天然水中5种Al形态的电化学测定。测定了20多个实际水样,与Driscoll方法进行了对照,结果基本一致。