Purification Of Target Compounds Research Articles

Chromatographic separation of itaconic anhydride recycling products with 5-amino-1-phenylpyrazoles

An important role in the creation of the entire variety of modern physiologically active compounds is played by the synthesis of complexly constructed poly–substituted molecules – analogues of natural substances. Effective design methods involve a significant increase in molecular complexity with a minimum number of stages. However, such an approach often involves complex processes involving multifunctional reagents, which, ultimately, can lead to difficult-to-separate mixtures of the target compound and a number of by-products, both undesirable and promising. This problem remains very significant in the design of various polyazaheterosystems, including those with a pyrazolopyridine skeleton, which is one of the most common fragments in the structures of natural and synthetic biologically active compounds. Therefore, an urgent task is to develop effective methods for the separation of such mixtures and purification of target compounds. Currently, column chromatography is used to solve it, which is one of the most important methods for separating liquid or solid mixtures of organic substances on a preparative scale. In this paper, the reactions of itacone anhydride with substituted 5-amino-1-phenylpyrazoles are investigated, a chromatographic method for separating alternative interaction products and their identification is proposed.

Benzothiazolium ionic liquid-induced crystallization of active alkaloid in its alcoholic solutions

Some ionic liquids have good potential to play the role of anti-solvents in the crystallization of natural active compounds after their initial extraction. In order to prove it and explore suitable candidates, a novel separation method of berberine hydrochloride was developed based on benzothiazolium ionic liquid induced crystallization in this study. Ionic liquids [C6Bth]PF6 and [HBth]BF4 were selected to conduct the separation process. The effects of various conditions on IL performance were investigated, under which the separation efficiency could achieve above 90% just by using a small amount of these ionic liquids. Moreover, the separation process was greatly simplified due to the different solubility of ionic liquids and berberine hydrochloride, so the further purification of target compounds and recycling process of the ILs could be avoided. The whole process is very easy to be scaled up and has been practiced for actual herbal sample.

Development of advanced biorefinery concepts using magnetically responsive materials

Utilization of magnetic materials and particles brings a considerable simplification into biorefinery technologies, based on their fast, simple and selective separation even from various difficult-to-handle environments and conditions. Additionally, application of magnetically responsive (bio)catalysts can significantly improve the productivity, economic feasibility, sustainability and product quality during the biorefinery processes. This review is focused on the presentation of typical examples of magnetic materials applicable in biorefinery, especially for the preparation of magnetically responsive enzymatic and whole-cell biocatalysts, as well as solid acid/base catalysts, magnetically responsive adsorbents or magnetically separable materials with suitable properties for isolation and purification of target compounds from complex mixtures. This brief overview emphasises remarkable possibilities of magnetic materials and magnetic techniques and their application potential in biorefinery technologies.

Polyglycidyl methacrylate based immunoaffinity cryogels for insulin adsorption.

Immunoaffinity chromatography (IAC) is a kind of bioaffinity chromatography which used antibodies or antibody-related molecules as the stationary phase. IAC is used by many applications for analytical, clinical and diagnostic purposes, particularly preferring in analytical purposes on one-step separation and purification of target compounds. Moreover, immunoaffinity chromatography is used in antibody enrichment and separation of cells. IAC columns are usually applied in the antibody experiments due to powerful and selective binding of antibodies and/or their target antigens. Antigen or antibody molecules could be immobilized to the solid support. Therefore, target antibody or cell is purified. Specific bioligands can be immobilized directly on glycidyl based polymeric material with simple acid-base catalyst. In this study, polyglycidyl methacrylate based therefore cryogels were prepared and anti-insulin antibodies were immobilized on porous surface of cryogels. Swelling test, Fourier transform infrared spectroscopy (FTIR), Raman spectroscopy, and scanning electron microscopy (SEM) were conducted to characterize cryogels developed. To optimize separation conditions, effects of pH, initial insulin concentration, flow rate, salt concentration, contact time and temperature on insulin adsorption capacity were examined. The results indicated that the immunoaffinity cryogel developed here could be classified as good alternative with prominent properties such as high reusability and cost-friendly adsorbent and would be one of the primary reports for immunoaffinity purification of insulin molecules in not only lab-scale but also for industrial purposes.

Development of a Method to Extract and Purify Target Compounds from Medicinal Plants in a Single Step: Online Hyphenation of Expanded Bed Adsorption Chromatography and Countercurrent Chromatography

Pure compounds extracted and purified from natural sources are crucial to lead discovery and drug screening. This study presents a novel two-dimensional hyphenation of expanded bed adsorption chromatography (EBAC) and high-speed countercurrent chromatography (HSCCC) for extraction and purification of target compounds from medicinal plants in a single step. The EBAC and HSCCC were hyphenated via a six-port injection valve as an interface. Fractionation of ingredients of Salvia miltiorrhiza and Rhizoma coptidis was performed on the hyphenated system to verify its efficacy. Two compounds were harvested from Salvia miltiorrhiza, one was 52.9 mg of salvianolic acid B with an over 95% purity and the other was 2.1 mg of rosmarinic acid with a 74% purity. Another two components were purified from Rhizoma coptidis, one was 4.6 mg of coptisine with a 98% purity and one was 4.1 mg of berberine with a 82% purity. The processing time was nearly 50% that of the multistep method. The results indicate that the present method is a rapid and green way to harvest targets from medicinal plants in a single step.

Conceptual Process Synthesis for Recovery of Natural Products from Plants: A Case Study of Artemisinin from Artemisia annua

A systematic method of conceptual process synthesis for recovery of natural products from their biological sources is presented. This methodology divides the task into two major subtasks namely, isolation of target compound from a chemically complex solid matrix of biological source (crude extract) and purification of target compound(s) from the crude extract. Process analytical technology (PAT) is used in each step to understand the nature of material systems and separation characteristics of each separation method. In the present work, this methodology is applied to generate process flow sheet for recovery of artemisinin from the plant Artemisia annua (A. annua). The process flow sheet is evaluated on the basis of yield and purity of artemisinin obtained in bench scale experiments. Yields of artemisinin obtained in individual unit operations of maceration, flash column chromatography, and crystallization are 90.0%, 87.1% and 47.6%, respectively. Results showed that the crystallization step is dominant to the overall yield of the process which was 37.3%.

Separation of Two Cytotoxic Saponins from the Roots of Adenophora triphylla var. japonica by High‐speed Counter‐current Chromatography

The roots of Adenophorae species have been reported to exhibit anti-obese, anti-oxidant, anti-cancer, and anti-bacterial activities. However, there has been no single report regarding the preparative isolation and biological activities of the chemical components from Adenophora triphylla. To develop an efficient method for the determination of the active fraction from the methanol extract from the roots of Adenophora triphylla and for the preparative isolation and purification of target compounds having cytotoxicity on carcinoma cells from the active fraction by high-speed counter-current chromatography (HSCCC). The Plant (5 kg, dry weight) was extracted with methanol. Three hundred grams of the dried methanol extract (885 g) were fractionated by open-column chromatography with a stepwise gradient of water-methanol. Preparative isolation of bioactive components was performed by HSCCC with a two-phase solvent system composed of ethyl acetate-n-butanol-0.2% trifluoroacetic acid in water (5:5:10, v/v). The cytotoxicity of column fractions and isolated compounds was evaluated by 2-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide assay. The 70% MeOH column fraction showed inhibitory effects against three human carcinoma cells A549, AGS and HepG2. Two saponins were separated from 400 mg of the active fraction by HSCCC. After further purification with solid phase extraction column, 25 mg of peak fraction 1 and 20 mg of peak fraction 2 were obtained. Their structures were identified by ¹H-NMR, ¹³C-NMR, Fourier transform infrared, fast atom bombardment-MS and electrospray ionisation-MS/MS. They exhibited strong cytotoxic effects against three cancer cells. Two cytotoxic saponins were isolated for the first time from the roots of Adenophora triphylla by HSCCC.

A new reversed-phase/strong anion-exchange mixed-mode stationary phase based on polar-copolymerized approach and its application in the enrichment of aristolochic acids

A novel silica-based reversed-phase/strong anion-exchange mixed-mode stationary phase named C18SAX was synthesized based on the polar-copolymerized approach. C18SAX stationary phase showed excellent compatibility with 100% aqueous mobile phase and comparable performance with commercial SunFire™ C18 column in terms of column efficiency and methylene selectivity. And the results indicated that hydrophobic interaction and anion-exchange mechanism dominate the separation process of aristolochic acids. The utility of C18SAX material-based SPE was demonstrated for the enrichment of aristolochic acids extracted from Caulis Aristochiae Manshuriensis. The SPE method was developed rapidly and simply by HPLC evaluation. Under the optimized SPE conditions, the average recoveries of aristolochic acid I and aristolochic acid II in crude sample were 72.36% and 105.59%, with RSDs of 5.77% and 1.13%, respectively. The complexity of sample matrices was significantly simplified after the SPE procedure and further preparation and purification of target compounds was performed on a preparative C18TDE column. Four aristolochic acids, including aristolochic acid I (31.8mg), II (8.9mg), IIIa (6.1mg) and IVa (4.3mg) were purified with HPLC purities all above 93%. The results proved that the aristolochic acids could be efficiently enriched by C18SAX material and easily eluted in acidic conditions, indicating its great potential in the enrichment of acidic compounds from complex matrices.

Synthesis of a αMan(1→3)αMan(1→2)αMan Glycocluster Presented on aβ‐Cyclodextrin Scaffold

AbstractApplication of 6‐thio‐α‐ and β‐cyclodextrins as the core component for the construction of multivalent carbohydrate structures is described. The method employed for the attachment of monomeric glycosides to a cyclodextrin core is based on the efficient nucleophilic displacement of bromide from an N‐bromoacetamido functionality. The N‐bromoacetyl group was positioned at the end of a spacer arm of glycosides and reacted with thiol groups present on the primary face of thiolated cyclodextrins. This coupling reaction led to perfunctionalised cyclodextrins which were thiother‐linked to appended saccharides through a short spacer arm. Both components of the key conjugation step — 6‐thiocyclodextrins and monomeric glycosides — were used without protecting groups, thus facilitating isolation and purification of target compounds. Glycoclusters incorporating six and seven α‐D‐mannopyranose or β‐D‐glucopyranose residues linked to cyclodexrin cores were synthesized in 57–75 % yield. Using the same technique, 3‐(2‐bromoacetamido)propyl trioside, incorporating a synthetic Man‐α‐(1→3)‐Man‐α‐(1→2)‐Man fragment of antigenic yeast mannan, was attached to per‐6‐thio‐β‐cyclodextrin to afford a heptavalent glycocluster. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2005)

Novel approach for the purification and analysis of methotrexate-branched polymeric polypeptide conjugates by HPLC

Methotrexate (MTX) has been coupled to various structurally related, polycationic synthetic branched polypeptides containing a poly[L-Lys] backbone. We describe here a novel, rapid and simple RP-HPLC approach and its application for a) the monitoring of MTX-macromolecule conjugate synthesis, b) analysis of the composition of conjugates samples, c) semi-preparative purification of target compounds from reagents and side products, and d) stability studies of MTX-branched polypeptide conjugate samples.

Liquid chromatography coupled with high-field proton NMR for profiling human urine for endogenous compounds and drug metabolites

A number of applications of high-field NMR spectroscopy have previously been described for the analysis of biological fluids (e.g. blood plasma, urine, bile, cerebrospinal fluid) both for endogenous compounds and for xenobiotics and their metabolites [l-4]. In the field of xenobiotic metabolism it has been amply demonstrated that multinuclear NMR spectroscopy of such biological fluids can provide a rapid and convenient approach to the detection of metabolites [l-8]. Similarly ‘H NMR spectroscopy has proved to be of value in the study of clinical samples and in studies on the mechanisms and time course of toxicity [9-111. However, the complexity of the biological fluid spectra obtained has often been such that identification of sample components from the single-pulse ‘H NMR spectrum has not been straightforward. In such circumstances two orthogonal approaches to the identification of unknowns can be used, singly, or in combination. In the first, sophisticated multipulse (and multidimensional) NMR methods can be used to extract structural information directly. In the second, the problem can be simplified by the selective removal of endogenous contaminants followed, if necessary, by the separation and isolation of a particular analyte. To this end strategies have been devised based on solidphase extraction-chromatography monitored by NMR spectroscopy (SPEC-NMR) [12-141. The extraction and purification of target compounds is achieved by stepwise gradient elution from a suitable SPE phase (e.g. octadecylsilane bonded silica). This methodology has proved to be robust, convenient, inexpensive and rapid. However, despite the practical success of the SPEC-NMR approach it remains a ‘low resolution’ technique (in terms of chromatographic separation), the deficiencies of which are compensated for, to a significant extent, by the NMR spectrometer which serves as the ‘detector’. For very complex mixtures, such as intact biological fluids, the hyphenation of NMR with liquid chromatography might be expected to offer clear benefits. Such systems have already been described [15-201 but, due to the technical difficulties involved, such as insufficient dynamic range and sensitivity and problems with solvent suppression, LC-NMR has not been widely exploited. However, given the significant recent technical advances in this field including increases in NMR spectrometer field strengths, methods for ensuring adequate solvent suppression together with the design of dedicated LC-NMR probes, the authors have