Search

1-Methylxanthine is a high-value derivative of caffeine of limited natural availability with many potential pharmaceutical applications. Unfortunately, production of 1-methylxanthine through purely chemical methods of synthesis are unfavorable due to lengthy chemical processes and the requirement of hazardous chemicals, ultimately resulting in low yields. Here, we describe a novel biosynthetic process for the production of 1-methylxanthine from theophylline using engineered Escherichia coli whole-cell biocatalysts and reaction optimization. When scaled-up to 1590 mL, the simple biocatalytic reaction produced approximately 1188 mg 1-methylxanthine from 1444 mg theophylline, constituting gram-scale production of 1-methylxanthine in as little as 3 hours. Following HPLC purification and solvent evaporation, 1163 mg of dried 1-methylxanthine powder was collected, resulting in a 97.9 wt% product recovery at a purity of 97.8%. This is the first report of a biocatalytic process designed specifically for the production and purification of the high-value biochemical 1-methylxanthine from theophylline. This process is also the most robust methylxanthine N -demethylation process featuring engineered E. coli to date, capable of gram-scale production. • Novel, biosynthetic pathway for production of 1-methylxanthine at gram scale. • Engineered E. coli for high-value biochemical production. • High performance liquid chromatography (HPLC) separation for high purity. • Co-factor (NADH) recycle through by-product (formaldehyde) degradation. [ABSTRACT FROM AUTHOR]

Tritium‐labeled compounds are generally less stable than their non‐labeled counterparts. This requires storage at low temperatures, a constant workflow of quality checks, and subsequent re‐purifications. As the amount of tritium‐labeled material is typically purified in the μg range, repeated injections on analytical‐scale ultra high‐performance liquid chromatography systems can provide high‐resolution re‐purification results. Yet, degradants can be undesirably included in the compound isolation because the amount of decomposition can vary dramatically depending on the structure. We report a case of a sensitive molecule that could not be isolated in pure form even though the chromatographic separation was successful. In this case, the use of a small‐scale two‐dimensional preparative liquid chromatography approach with a direct transfer interface to a second (trapping) column resulted in a highly pure compound (>98% radiochemical purity). This approach combines high chromatographic resolution, accurate control over the re‐purification process, minimal sample manipulation, and higher overall safety for the handling of radioactive samples. [ABSTRACT FROM AUTHOR]

Copyright of Science & Technology of Cereals, Oils & Foods is the property of Science & Technology of Cereals, Oils & Foods Editorial Office and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

• Optimisation of the separation based on global models was carried out. • A green tea sample was used as representative of the teas in the optimisation. • Expected and actual separations matched well. • Optimised chromatogram is used to classify 40 samples of 6 tea varieties. • Satisfactory results with supervised and unsupervised classificatory techniques. As a result of their metabolic processes, medicinal plants produce bioactive molecules with significant implications for human health, used directly for treatment or for pharmaceutical development. Chromatographic fingerprints with solvent gradients authenticate and categorise medicinal plants by capturing chemical diversity. This work focuses on optimising tea sample analysis in HPLC, using a model-based approach without requiring standards. Predicting the gradient profile effects on full signals was the basis to identify optimal separation conditions. Global models characterised retention and bandwidth for 14 peaks in the chromatograms across varied elution conditions, facilitating resolution optimisation of 63 peaks, covering 99.95 % of total peak area. The identified optimal gradient was applied to classify 40 samples representing six tea varieties. Matrices of baseline-corrected signals, elution bands, and band ratios, were evaluated to select the best dataset. Principal Component Analysis (PCA), k -means clustering, and Partial Least Squares-Discriminant Analysis (PLS-DA) assessed classification feasibility. Classification limitations were found reasonable due to tea processing complexities, involving drying and fermentation influenced by environmental conditions. [ABSTRACT FROM AUTHOR]

Background: Nowadays, in Nuclear Medicine, clinically applied radiopharmaceuticals must meet quality release criteria such as high radiochemical purity and radiochemical yield. Many radiopharmaceuticals do not have marketing authorization and have no dedicated monograph within European Pharmacopeia (Ph. Eur.); therefore, general monographs on quality controls (QCs) have to be applied for clinical application. These criteria require standardization and validation in labeling and preparation, including quality controls measurements, according to well defined standard operation procedures. However, QC measurements are often based on detection techniques that are specific to a certain chromatographic system. Several radiosyntheses of [68Ga]Ga-radiopharmaceuticals are more efficient and robust when they are performed with 2-[4-(2-hydroxyethyl)piperazin-1-yl] ethanesulfonic acid (HEPES) buffer, which is considered as an impurity to be assessed in the QC procedure, prior to clinical use. Thus, Ph. Eur. has introduced a thin-layer chromatography (TLC) method to quantify the HEPES amount that is present in [68Ga]Ga-radiopharmaceuticals. However, this is only qualitative and has proven to be unreliable. Here we develop and validate a new high-performance liquid chromatography (UV-Radio-HPLC) method to quantify the residual amount of HEPES in 68Ga-based radiopharmaceuticals. Method: To validate the proposed UV-Radio-HPLC method, a stepwise approach was used, as defined in the guidance document that was adopted by the European Medicines Agency (CMP/ICH/381/95 2014). The assessed parameters are specificity, linearity, precision (repeatability), accuracy, and limit of quantification. A range of concentrations of HEPES (100, 80, 60, 40, 20, 10, 5, 3 μg/mL) were analyzed. Moreover, to test the validity and pertinence of our new HPLC method, we analyzed samples of [68Ga]Ga-DOTATOC; [68Ga]Ga-PSMA; [68Ga]Ga-DOTATATE; [68Ga]Ga-Pentixafor; and [68Ga]Ga-NODAGA-Exendin-4 from different batches that were prepared for clinical use. Results: In the assessed samples, HEPES could not be detected by the TLC method that was described in Ph. Eur. within 4 min incubation in an iodine-saturated chamber. Our developed HPLC method showed excellent linearity between 3 and 100 μg/mL for HEPES, with a correlation coefficient (R2) for calibration curves that was equal to 0.999, coefficients of variation (CV%) < 2%, and percent deviation value of bias from 100% to 5%, in accordance with acceptance criteria. The intra-day and inter-day precision of our method was statistically confirmed and the limit-of-quantification (LOQ) was 3 μg/mL, confirming the high sensitivity of the method. The amount of HEPES that was detected with our developed HPLC method in the tested [68Ga]Ga-radiopharmaceuticals resulted well below the Ph. Eur. limit, especially for [68Ga]Ga-NODAGA-Exendin-4. Conclusions: The TLC method that is described in Ph. Eur. to assess residual HEPES in [68Ga]-based radiopharmaceuticals may not be sufficiently sensitive and thus unsuitable for QC release. Our new HPLC method was sensitive, quantitative, reproducible, and rapid for QCs, allowing us to exactly determine the residual HEPES amount in [68Ga]Ga-radiopharmaceuticals for safe patient administration. [ABSTRACT FROM AUTHOR]

By adapting previously reported reaction conditions, [18F]FMISO was synthesized using a TracerLab FX‐FDG module in 31 min from no carrier added [18F]fluoride in 56% radiochemical yield. A novel and simple purification setup based on disposable cartridges was developed allowing the radiopharmaceutical to be obtained in high radiochemical purity (>99.5%) and with chemical impurities far below the Ph.Eur.'s thresholds. Moreover, the study pinpointed some shortcomings of the high‐performance liquid chromatography (HPLC) method reported in the dedicated Ph.Eur's monograph. [ABSTRACT FROM AUTHOR]

The basis for the release of [18F]FDG for use is the results of quality control studies within the scope of the specification, based on the recommendations of the Pharmacopoeia (FP XI and Ph. Eur. IX). The object of this study is the influence of the [18F]FDG hydrolysis time on the quality parameters, with particular regard to radiochemical purity. The synthesis was performed at three different hydrolysis times: 8, 10, and 12 minutes. 21 syntheses were performed for each time. For the [18F]FDG solution, the radiochemical purity was assessed with the TLC method, the radiochemical and chemical purity with the HPLC method, and the pH value with the potentiometric method. Extending the hydrolysis time significantly increased the radiochemical purity of the final [18F]FDG product, improving its quality parameters (mainly reducing the amount of acetyl derivatives of [18F]FDG while maintaining chemical purity and pH value but with a slight loss of activity. A result at the accept/reject limit may negatively affect the stability test. Increasing the radiochemical purity allows us to avoid the invalidation of this test due to the influence of device measurement error and possible operator error (human). Minimizing contamination reduces impact GLUT independent tumor cell accumulation of [18F]FDG, which its detail of character is still unknown. The possibility of a false-positive result in an imaging examination and minimizes the impact of radiation from contaminants on the patient's tissues. Additional biological studies should be performed to investigate the metabolism of the acetyl derivative of [18F]FDG. [ABSTRACT FROM AUTHOR]

[Objectives] This study was conducted to isolate and screen the bacteria that can convert trans-anethole to anisic acid from star anise and its environmental samples, and identify the bacteria. [Methods] According to the traditional microbial culture method, with trans-anethole as the sole carbon source, through enrichment culture and separation and purification, preliminary screening by thin layer chromatography and re-screening by high-performance liquid chromatography, strains that degraded trans-anethole to produce anisic acid were obtained, and 16 S rDNA sequencing and phylogenetic tree construction were performed for genetic analysis. [Results] Eleven strains that degraded trans-anethole to produce anisic acid were obtained, among which strain NT2 that produced anisic acid with a relatively high efficiency was initially identified as Pseudomonas sp. The strain's trans-anethole degradation rate was 45.41%, and the molar production rate and cumulative concentration of anisic acid were 21.80% and 1.96 g/L, respectively. [Conclusions] Strain NT2 has a strong ability to degrade trans-anethole to produce anisic acid, and can enrich strain resources for degradation of trans-anethole to anisic acid through microbial conversion. [ABSTRACT FROM AUTHOR]

Saponins extracted from Sapindus mukorossi are natural nonionic surfactants with various biological activities and strong surface activities, which can be widely used in biomedicine and the daily cleaning chemical industry. In this paper, Sapindus pericarp was used as the raw material for the extraction of saponins. Effects of major extraction variables, including solvents, extraction methods, material-to-liquid ratios, and extraction time were investigated. A yield of 30.48% and purity of 61.38% were obtained by using water extraction for 4 h with a material-to-liquid ratio of 1 : 6. Fermentation was used to purify the saponins, and a high sugar resistant yeast performed better with an optimal amount of 5%. The saponins concentration slightly decreased from 38.86 g L−1 to 37.72 g L−1 after fermentation, and the purity of the saponins increased to 78.97%. Structure and performance tests of homemade saponins were also carried out. The structure of Sapindus saponins was found to be similar to that of mukurozi-saponin using FTIR and HPLC, and the foam half-life of homemade saponins was 25.82% longer than that of unfermented saponins. On comparing four chemical surfactants, the foam half-life of saponins after fermentation was found to be much longer. These results provide useful information for the development of industrial purification and application of Sapindus saponins. [ABSTRACT FROM AUTHOR]

Cannabis indica obtained from the Office of the Narcotics Control Board (ONCB) of Thailand was examined for medical purposes. The ONCB cannabis was seized and distributed to our research group for investigation of chemical compositions and purification of major cannabinoids. Its essential oil was analysed and aromadendrene was identified as the major terpene. Small amounts of cannabinoids were also observed. The components of the seized cannabis were extracted using ethanolic extraction and supercritical CO2 extraction. All crude extracts were analysed using high-performance liquid chromatography coupled with a cannabinoid analyser. All extracts contained cannabidiol (CBD), cannabinol (CBN), and Δ99-tetrahydrocannabinol (Δ9-THC) as the major constituents. A preliminary study showed notable anticancer activities of CBD, CBN and Δ99-THC against cervix carcinoma cell and epithelial human breast cancer cell. [ABSTRACT FROM AUTHOR]

The effect of installing different lipid chains (C 6 , C 8 , C 10 , and C 16) on the N -terminus of an octapeptide derived from the antiparallel β-interface of the diaminopimelate decarboxylase protein homotetramer has been investigated. Notably, the C 8 peptide conjugate assembled into wide twisted nanoribbons and formed hydrogels, which to the best of our knowledge constitutes the first example of a peptide containing an eight carbon alkyl chain that demonstrates these properties, a space typically occupied by peptide amphiphiles with long lipid chains. Furthermore, this self-assembling lipopeptide exhibited pH and temperature stability with shear thinning properties suitable for biomedical applications. Importantly, in this work the application of the polystyrene-based sorbent Diaion™ HP20SS for the simple large-scale purification of self-assembling peptides is presented as an alternative to the use of time-consuming and labor-intensive reverse-phase high-performance liquid chromatography. Peptides that can self-assemble into defined nanostructures are highly attractive for many biomedical applications given their unique physical and chemical properties. It is recognized that self-assembling peptides derived from naturally occurring proteins offer an unlimited source of functionalities and structures, which are hard to uncover with designed sequences. In this study, we have investigated the effect of installing different lipids chains on the N-terminus of an octapeptide derived from the antiparallel β-interface of the diaminopimelate decarboxylase protein homo tetramer. We also reported the use of polymeric DiaionⓇ HP20SS beads as an alternative solid support to purify self-assembling peptides. Image, graphical abstract [ABSTRACT FROM AUTHOR]

The marine microalga Tisochrysis lutea, a Haptophyta with a thin cell wall and currently used mainly in aquaculture is a potential source of several bioactive compounds of interest such as carotenoids. In the present study, the simultaneous extraction and purification of fucoxanthin, the main carotenoid from T. lutea, was optimized using pressurized fluid extraction followed by in‐cell purification. An experimental design was employed to maximize carotenoids' extraction; the experimental factors chosen were: (i) percentage of ethanol/ethyl acetate (0–100 %), (ii) temperature (40–150°C), and (iii) number of static extraction cycles (1–3). The maximum carotenoids' recovery, mainly fucoxanthin, was obtained with pure ethyl acetate at 40°C using one extraction cycle, achieving values of 132.8 mg of carotenoids per gram of extract. Once the optimum extraction conditions were confirmed, in‐cell purification strategies using different adsorbents were developed to obtain fucoxanthin‐enriched extracts. Activated charcoal showed potential retention of chlorophylls allowing an effective purification of fucoxanthin in the obtained extracts. Chemical characterization of extracts was carried out by reversed‐phase high‐performance liquid chromatography with diode array detection. Therefore, a selective fractionation of high value compounds was achieved using the proposed green downstream platform based on the use of compressed fluids. [ABSTRACT FROM AUTHOR]

The extraction condition of curcumin from Curcuma longa L was optimized through four factors and three levels orthogonal experiment based on the results of single factor tests. Under the optimal conditions: the concentration of ethanol 80%, extraction temperature 70°C, the ratio of liquid to material 20, and extraction time 3 h, a crude extract with the yield of curcumin 56.8 mg/g could be obtained. The isolation and purification of curcuminoids from the crude extract was performed on high performance counter current chromatography employing an optimized solvent system n‐hexane/ethyl acetate/methanol/water (2/3/3/1, v/v/v/v). From 97 mg crude sample (in which the purity of curmumin was 68.56%), 67 mg curmumin, 18 mg demethoxycurcumin, and 9.7 mg bisdemethoxycurcumin with a high‐performance liquid chromatography purity of 98.26, 97.39, and 98.67%, respectively, were obtained within 70 min. The antioxidant activities and cytotoxicity of purified curcumin was comparable to that of the commercial product, indicating that the biological activity of curcumin could be maintained by this method. [ABSTRACT FROM AUTHOR]

A novel method based on acid–base-induced cloud point extraction (AB-CPE) and magnetic solid-phase extraction (MSPE) coupled with high performance liquid chromatography (HPLC) for extraction and purification of 3-acetyl-2,5-dimethylthiophene (ADP) in the fast food noodle samples was established for the first time. In MSPE-AB-CPE, Fe3O4@SiO2 nanoparticles are used to purify samples, medium-chain saturated fatty acids (n-octanoic acid), and nonionic surfactant (TMN-6) were investigated as a pH-dependent phase based on acid–base reaction caused solely by adjusting pH. The calibration curves demonstrated linearity in the range 0.5–100 µg mL−1 (R2 = 0.9969), with relative recoveries (95.7–101.2%) and relative standard deviations (2.6–4.8%, n = 5). [ABSTRACT FROM AUTHOR]

The ever-increasing speed of biotherapeutic drug discovery has driven the development of automated and high throughput purification capabilities. Typically, purification systems require complex flow paths or third-party components that are not found on a standard fast protein liquid chromatography instrument (FPLC) (e.g., Cytiva's ÄKTA) to enable higher throughput. In early mAb discovery there is often a trade-off between throughput and scale where a high-throughput process requires miniaturized workflows necessitating a sacrifice in the amount of material generated. At the interface of discovery and development, flexible automated systems are required that can perform purifications in a high-throughput manner, while also generating sufficient quantities of preclinical material for biophysical, developability, and preclinical animal studies. In this study we highlight the engineering efforts to generate a highly versatile purification system capable of balancing the purification requirements between throughput, chromatographic versatility, and overall product yields. We incorporated a 150 mL Superloop into an ÄKTA FPLC system to expand our existing purification capabilities. This allowed us to perform a range of automated two-step tandem purifications including primary affinity captures (protein A (ProA)/immobilized metal affinity chromatography (IMAC)/antibody fragment (Fab)) followed by secondary polishing with either size exclusion (SEC) or cation exchange (CEX) chromatography. We also integrated a 96 deep-well plate fraction collector into the ÄKTA FPLC system with purified protein fractions being analyzed by a plate based high performance liquid chromatography instrument (HPLC). This streamlined automated purification workflow allowed us to process up to 14 samples within 24 h, enabling purification of ∼1100 proteins, monoclonal antibodies (mAbs), and mAb related protein scaffolds during a 12-month period. We purified a broad range of cell culture supernatant volumes, between 0.1 and 2 L, with final purification yields up to 2 g. The implementation of this new automated, streamlined protein purification process greatly expanded our sample throughput and purification versatility while also enabling the accelerated production of greater quantities of biotherapeutic candidates for preclinical in vivo animal studies and developability assessment. • Comprehensive characterization of manual vs automated two-step protein purification. • Superloop enabled automated purification of ∼1100 molecules in 12 months. • Automation of tandem two-step purifications through incorporation of a Superloop. • Enabling preclinical studies through automated two-step purifications. • Versatility of automated two-step purification imparted by use of a Superloop. [ABSTRACT FROM AUTHOR]

Medium pressure liquid chromatography and semipreparative high-performance liquid chromatography, followed by recycling high-performance liquid chromatography, were used for the isolation and purification of oligostilbenes from the acetone extracts ofDipterocarpus semivestitusandNeobalanocarpus heimii. Two stilbene dimers, ε-viniferin andcis-ε-viniferin, and a stilbene trimer,cis-miyabenol C were isolated fromD. semivestitus. A trimer, ampelopsin C, and two tetramers, hemsleyanol D and isohopeaphenol, were isolated fromN. heimii. The purities of the compounds were determined by an ultrahigh-performance liquid chromatography. [ABSTRACT FROM PUBLISHER]

This instalment in our series on ultrahigh-pressure liquid chromatography (UHPLC) highlights its benefits in fast analysis, high-resolution separations, high performance liquid chromatography (HPLC) method development, reduced solvent and sample usage, and enhanced sensitivity and precision performance. [ABSTRACT FROM AUTHOR]

The current in-solution analysis of N -glycans suffers from several disadvantages including tedious de-glycosylation time and multi-step pre-treatment procedures. Here, an ultra-simple flowing on-line analysis of labeled N -glycans for high-performance liquid chromatography with fluorescence detection (HPLC-FLD) was developed for eliminating the deficiencies. This on-line analysis consisted of an immobilized enzyme reactor (IMER) of PNGase F for efficient release of N -glycans, labeling of released N -glycans and following purification of derivatives on microfluidic chip. Notably, efficient preparations for all type of N -glycans were completed within ∼30 min. To our best knowledge, this is the first time to integrated the whole preparation of N -glycan deglycosylation, labeling and purification only by a simple fluidic flow with our developed device. Good reproducibility and stability were achieved with the relative standard deviation (RSD) less than 10%. Furthermore, the glycome studies with human serum revealed a good adaptability for biological samples. Our work provides an efficient N -glycomic strategy that can be applied to further multilayered clinical analysis. [Display omitted] • Flowing on-line preparation of N -glycan based on IMER and microfluidic chip was first time developed. • Efficient preparations for all types of N -glycans were completed within 30 min. • Good repeatability, reproducibility, stability and adaptability were achieved. • Providing an efficient and high-throughput N -glycomic strategy for further multilayered clinical analysis. [ABSTRACT FROM AUTHOR]

The glyceride in oil food simulant usually causes serious interferences to target analytes and leads to failure of the normal function of the RP-HPLC column. In this work, a convenient HPLC-UV method for the determination of the total specific migration of nine ultraviolet (UV) absorbers in food simulants was developed based on 1,1,3,3-tetramethylguanidine (TMG) and organic phase anion exchange (OPAE) SPE to efficiently remove glyceride in olive oil simulant. In contrast to the normal ion exchange carried out in an aqueous solution or aqueous phase environment, the OPAE SPE was performed in the organic phase environments, and the time-consuming and challenging extraction of the nine UV absorbers from vegetable oil with aqueous solution could be readily omitted. The method was proved to have good linearity (r ≥ 0.99992), precision (intra-day RSD ≤ 3.3%), and accuracy(91.0% ≤ recoveries ≤ 107%); furthermore, the lower limit of quantifications (0.05–0.2 mg/kg) in five types of food simulants(10% ethanol, 3% acetic acid, 20% ethanol, 50% ethanol and olive oil) was observed. The method was found to be well suited for quantitative determination of the total specific migration of the nine UV absorbers both in aqueous and vegetable oil simulant according to Commission Regulation (EU) No. 10/2011. Migration levels of the nine UV absorbers were determined in 31 plastic samples, and UV-24, UV-531, HHBP and UV-326 were frequently detected, especially in olive oil simulant for UV-326 in PE samples. In addition, the OPAE SPE procedure was also been applied to efficiently enrich or purify seven antioxidants in olive oil simulant. Results indicate that this procedure will have more extensive applications in the enriching or purification of the extremely weak acidic compounds with phenol hydroxyl group that are relatively stable in TMG n -hexane solution and that can be barely extracted from vegetable oil. [ABSTRACT FROM AUTHOR]

A standard-free and sensitive method was developed for analysis of sucrose acetates in a crude 6- O -acetyl sucrose (S-6-a) product by on-line hydrolysis-high-performance liquid chromatography with pulsed amperometric detection (PAD). Sucrose, three regio-isomers of acetyl sucrose and five regio-isomers of diacetyl sucrose were separated on a C18 column using 3% (v/v) acetonitrile in water as eluent within 25 min. After purification with LC followed by semi-preparative HPLC, their chemical structures were identified by 1D, 2D NMR and LC–MS. Moreover, quantification of those regio-isomers was achieved by on-line alkaline hydrolysis to liberate sucrose using a post-column delivery system, and then detected by PAD for indirect estimation of the sucrose acetate content. Under optimal conditions, the linear ranges were from 0.03 to 150 μmol L −1 for sucrose corresponding to sucrose acetates with coefficient of determination as 0.9997 and detection limit as 0.01 μmol L −1 ( S/N = 3). Good repeatability was obtained (RSD < 3%, n = 6). Furthermore, this method has been successfully applied to the analysis of sucrose and sucrose acetates in a crude S-6-a product during synthesis, purification and structure elucidation studies. The recoveries were from 94.89% to 102.31% for sucrose and sucrose acetates. [ABSTRACT FROM AUTHOR]

Herein, we report a short, three-step synthesis of d-[1-14C]-serine (4) in high enantiomeric purity. Starting from [14C]-KCN and 2-(benzyloxy)acetaldehyde, Strecker reaction using ( R)-1-phenylethylamine as the chiral auxiliary gave two diastereomeric aminonitriles 1 and 2 in the ratio of 4:3, which were conveniently separated and purified chromatographically. Following hydrolysis and subsequent hydrogenolysis, the purified major diastereomer 1, was smoothly converted to d-[1-14C]-serine (4) in an enantiomeric excess of >99%, thus circumventing time intensive chiral HPLC enantiomeric resolution. [ABSTRACT FROM AUTHOR]

A three-stage linear gradient strategy using reverse-phase high-performance liquid chromatography (HPLC) was optimized for rapid, high-quality, and simultaneous purification of the lipopeptide isoforms of iturin, fengycin, and surfactin, which may differ in composition by only a single amino acid and/or the fatty acid residue. Matrix-assisted laser desorption ionization time-of-flight tandem mass spectrometry (MALDI-TOF-MS/MS) was applied to detect the lipopeptides harvested from each reversed-phase HPLC peak. Amino acid analysis based on phenyl isothiocyanate derivatization was further used for confirmation of the amino acid species and molar ratio in a certain HPLC fraction. By this MALDI-TOF-MS/MS coupled with amino acid analysis, it was revealed that iturin at m/ z 1,043 consists of a circular Asn-Tyr-Asn-Gln-Pro-Asn-Ser peptide and C β-OH fatty acid. Surfactin homologs from Bacillus subtilis THY-7 at m/ z 1,030, 1,044, 1,058, and 1,072 possess a circular Glu-Leu-Leu-Val-Asp-Leu-Leu peptide and the β-OH fatty acid with a different length (C-C). Fengycin species at m/ z 1,463 and 1,477 are homologs possessing the circular peptide Glu-Orn-Tyr-Thr-Glu-Ala-Pro-Gln-Tyr-Ile linked to a C or C γ-OH fatty acid, whereas fengycin at m/ z 1,505 contains a Glu-Orn-Tyr-Thr-Glu-Val-Pro-Gln-Tyr-Ile sequence with a Val instead of Ala at position 6. The method developed in this work provided an efficient approach for characterization of diverse lipopeptide isoforms from the iturin, fengycin, and surfactin families. [ABSTRACT FROM AUTHOR]

Radix isatidishas been widely used as a Chinese traditional medicine for its antivirus and anticancer activities where the minor components may contribute to these beneficial pharmaceutical effects. In order to enrich the target minor compounds effectively and rapidly, extraction, medium-pressure liquid chromatography (MPLC), high-speed countercurrent chromatography (HSCCC), and preparative high-performance liquid chromatography (pre-HPLC) were integratively used for separation and purification of two target minor compounds indole-3-acetonitrile-6-O-β-D-glucopyranoside (target 1) and clemastanin B (target 2) in the present study.Radix isatidiswas dried, pulverized, and extracted with 50% methanol at room temperature, then concentrated, and subjected to pretreatment with D-101 macroporous resin chromatography and extraction by MPLC. The first target compound was separated by MPLC at the purity raised to 70–80%, but without the second minor compounds which were irreversibly adsorbed by C18 solid support. Therefore, the second target compound in the crude extract was directly separated by HSCCC at a purity of 80–90%. Finally, these refined samples were further separated by pre-HPLC to obtain a high purity at 98–99%. The chemical structure identification of each target compound was carried out by infrared radiation (IR), electrospray ionization mass spectrometry (ESI-MS), and1H-NMR. [ABSTRACT FROM AUTHOR]

ABSTRACT The concept of Process Analytical Technology ( PAT) introduced by the US FDA has received a lot of attention lately. The use of such a scheme ensures robust process performance over the entire life cycle of the product and consistent product quality at the end of the manufacturing process. Successful implementation of PAT requires use of high resolution, orthogonal analytical tools that are able to measure the critical quality attributes ( CQA) of the raw materials and in-process materials with the aim of ensuring final product quality. This article presents a review of PAT applications for monitoring commonly used downstream biotech unit operations. Focus of the review will be on recent advancements (last 5 years). Analytical tools that have been used in these approaches have also been discussed with reference to their speed of analysis, robustness and sensitivity. Also discussed is their potential to replace the traditional high performance liquid chromatography ( HPLC) which continues to be the gold standard for analysis of biotech therapeutics. © 2014 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

A methodology to purify a Ho solution has been developed by a combination of activity and mass concentration measurements in order to further determine the Ho half-life. The isobaric interference at m/ q ≃ 166 requires Ho purification from non-natural Er with a high purification degree due to the large amount of Ho as opposed to Er. The Ho/Er separation was achieved using high-performance liquid chromatography on a semi-preparative column followed by purification on gravitational chromatography. The efficiency of the separation was evaluated after precise determination of the Er isotopic composition. The purification methodology enabled to separate Ho from Er. [ABSTRACT FROM AUTHOR]

Highlights: [•] A new preparative recycle HPLC system with a multifunctional trap was built up. [•] The system was designed for separating a low abundant compound from a complex mixture. [•] Absorption capability of the trap can be modified by online mobile phase adjusting. [•] The system could work against the problems of volume overloading and solvent effect. [•] The system could speed up the drying procedure of the final product. [Copyright &y& Elsevier]

Highlights: [•] CE/UV was assessed as a chemical purity analysis platform for PET tracers. [•] Two samples of PET tracers and their impurities were analyzed as examples. [•] The performance of CE was compared with the accepted performance of HPLC. [•] CE provided shorter analysis times, μ-molar LOD and sufficient reproducibility. [•] CE/UV was introduced as a novel method for the quality control of Kryptofix2.2.2. [Copyright &y& Elsevier]

Abstract: Cyclodextrins (CDs) do not disturb HPLC purification and do not form lather, which makes them superior to commonly used surfactants. In this paper, the authors describe the utility of CDs, specifically α-CD, β-CD, and γ-CD, for the synthesis of neoglycolipids from insoluble peptide-linked substrates. γ-CD was found to be well matched to the bulky benzyl group and accelerated the reaction for the substrate having Phe (F), while β-CD was well matched to the methyl group and accelerated the reaction for the substrate having Ala (A). Galactosylation to water-insoluble peptide-linked glucosaminide substrate by galactosyltransferase proceeded 63% at the most, but using either β-CD or γ-CD gave quantitative yield within 10 h. [Copyright &y& Elsevier]

Highlights: [•] A new fucoidan from kelp crude polysaccharide was purified. [•] GPC–HPLC profiles showed KPS-2-1 was a single component. [•] The FT-IR spectrum showed that the purified fucoidan was a sulfated polysaccharide. [•] The purified fucoidan is a strong competitive inhibitor of the tyrosinase. [•] The study lays a good foundation for further developing the fucoidan from kelp. [ABSTRACT FROM AUTHOR]

Forsythoside A and suspensaside A were isolated and purified fromForsythia suspensaby one step of high-speed counter-current chromatography for the first time using a solvent system of ethyl acetate-n-butanol-methanol-water (4:0.5:0.5:5,v/v). The purities of the two compounds were 98.19% and 98.05%, respectively, as determined by HPLC. Their structures were identified by MS, UV, and NMR analysis. Such a simple and effective method was fairly useful to prepare pure compounds as reference substances for related study onForsythia suspensa. [ABSTRACT FROM PUBLISHER]

Abstract: The present study aims to provide new in vitro and in vivo biochemical information about a novel Kunitz trypsin inhibitor purified from Piptadenia moniliformis seeds. The purification process was performed using TCA precipitation, Trypsin–Sepharose and reversed-phase C18 HPLC chromatography. The inhibitor, named PmTKI, showed an apparent molecular mass of around 19 kDa, visualized by SDS-PAGE, which was confirmed by mass spectrometry MALDI-ToF demonstrating a monoisotopic mass of 19.296 Da. The inhibitor was in vitro active against trypsin, chymotrypsin and papain. Moreover, kinetic enzymatic studies were performed aiming to understand the inhibition mode of PmTKI, which competitively inhibits the target enzyme, presenting Ki values of 1.5 × 10−8 and 3.0 × 10−1 M against trypsin and chymotrypsin, respectively. Also, the inhibitory activity was assayed at different pH ranges, temperatures and reduction environments (DTT). The inhibitor was stable in all conditions maintaining an 80% residual activity. N-terminal sequence was obtained by Edman degradation and the primary sequence presented identity with members of Kunitz-type inhibitors from the same subfamily. Finally after biochemical characterization the inhibitory effect was evaluated in vitro on insect digestive enzymes from different orders, PmTKI demonstrated remarkable activity against enzymes from Anthonomus grandis (90%), Plodia interpuncptella (60%), and Ceratitis capitata (70%). Furthermore, in vivo bioinsecticidal assays of C. capitata larvae were also performed and the concentration of PmTKI (w/w) in an artificial diet required to LD50 and ED50 larvae were 0.37 and 0.3% respectively. In summary, data reported here shown the biotechnological potential of PmTKI for insect pest control. [Copyright &y& Elsevier]

Silica gel column combined with high-speed counter-current chromatography separation was successfully applied to the separation of schizandrin (I), ange-loylgomisin H (II), gomisin A (III), schisantherin C (IV), deoxyschizandrin (V), γ-schisandrin (VI) and schisandrin C (VII) from the fruits of Schisandra chinensis (Turcz.) Baillon. The petroleum ether extracts of the fruits of S. chinensis were pre-separated first on a silica gel column and divided into two fractions as sample 1 and sample 2. 260 mg of sample 1 was separated by HSCCC using petroleum ether-ethyl acetate-methanol-water (10:8:10:8, v/v) as the two-phase solvent system and 18.2 mg of schizandrin, 15.7 mg of angeloylgomisin H, 16.5 mg of gomisin A and 16.7 mg of schisantherin C were obtained. 230 mg of sample 2 was separated using petroleum ether-ethyl acetate-methanol-water (10:0.5:10:1, v/v) as the two-phase solvent system and 19.7 mg of deoxyschizandrin, 23.4 mg of γ-schisandrin and 18.2 mg of schisandrin C were obtained. The purities of the separated compounds were all over 94% as determined by HPLC. The chemical structures of these compounds were confirmed by ESI-MS and ¹H-NMR. [ABSTRACT FROM AUTHOR]

The present study describes an efficient hyphenated flash chromatographic (FC) for the purification of coumarins and polymethoxy flavones (PMF's) from cold pressed grapefruit peel oil (GFO), a byproduct from citrus juice processing plant. GFO was subjected to silica column chromatography using sequential isocratic elution by hexane and chloroform to separate the waxes and phytochemicals enriched fractions, respectively. The chloroform extract was used for hyphenated FC separation using a gradient mobile phase of hexane and acetone. The peak separations were monitored at dual wavelengths of λ210 nm and λ340 nm. The individual fractions were analyzed by HPLC, pooled and concentrated resulting in isolation of two coumarins (auraptene and bergapten) and three PMF's (tangeretin, heptamethoxyflavone, and nobiletin). The purity and identity of the isolated compounds was confirmed by spectral analysis using high performance liquid chromatography, matrix-assisted laser desorption/Ionization-time of flight-mass spectrometry, and nuclear magnetic resonance. The hyphenated FC separation method was further validated by test of repeatability resulting in low RSD for the yield of the phytochemicals. Utilization of the developed separation method for isolation of value added products from grapefruit byproducts could be beneficial to the citrus processing industry.Supplemental materials are available for this article. Go to the publisher's online edition ofSeparation Science & Technologyto view the supplemental file. [ABSTRACT FROM PUBLISHER]

A 17-run Box-Behnken design (BBD) with three factors was used to improve the conditions for extracting astaxanthin from shrimp waste. The astaxanthin level was determined by high performance liquid chromatography (HPLC) using a C18column and a UV detector at 476 nm. The mean extraction yield of astaxanthin at the improved conditions (extraction time 1.4 h, extraction temperature 72.4°C, and liquid-solid ratio 27.3) was 98.7 ± 2.6 µg g−1. A solid-phase extraction (SPE)-HPLC method was developed with a new hybrid organic-inorganic hybrid monolith for further purification of astaxanthin from shrimp waste. The SPE recoveries were ranging from 81.3 ± 2.4% to 86.5 ± 3.3%, and the intra-day and inter-day relative standard deviations (RSDs) of the proposed method were less than 4.3 ± 0.5% and 4.8 ± 0.2%, respectively. BBD increased the extraction efficiency and shortened extraction times significantly. SPE-HPLC with hybrid monolith showed good selectivity and purified astaxanthin from shrimp waste. [ABSTRACT FROM PUBLISHER]

An effective high–speed counter–current chromatography (HSCCC) method was established for semi-preparative isolation and purification of bioactive chemical constituents from the Tibetan medicinal plantPedicularis longifloravar.tubiformis(Klotzsch) P. C. Tsoong. With a two-phase solvent system composed of chloroform–methanol–water (8:4:5, v/v), 40 mg of extract was separated to yield luteolin (12.5 mg), apigenin (9.6 mg), and chrysoeriol (4.8 mg), with purities of 99.3, 98.2, and 98.6%, respectively, as determined by high-performance liquid chromatography (HPLC). The chemical structures of these three components were identified by1H NMR and13C NMR. [ABSTRACT FROM PUBLISHER]

In this study, a preparative high performance centrifugal partition chromatography (CPC) method for isolation and purification of the bioactive component astilbin from the crude extract ofSmilax glabrawas successfully established by usingn-hexane-ethyl acetate-methanol-water as the two-phase solvent system. The upper phase ofn-hexane-ethyl acetate-methanol-water (1:10:1:10, v/v) was used as the stationary phase of CPC. Under the optimum conditions, 88 mg of astilbin at 98.5% purity, as determined by HPLC, was yielded from 500 mg of the crude extract in a single CPC separation. The peak fraction of CPC was identified by1H NMR and13C NMR. [ABSTRACT FROM AUTHOR]

The detrimental effect of heterogeneity on the efficiency of particle packed columns has been a long standing issue in chromatographic separations. This heterogeneity arises from the difference in packing density between the wall and central regions of the chromatographic bed and imparts radial variance in mobile phase velocity through the column. In conjunction with the uneven distribution of the sample band through frits and distributors at the head of the column, the resultant effect is a decrease in efficiency due to the formation of a parabolic-“like” sample band. We introduce the concept of “Active Flow Management” to counteract the effects of column heterogeneity via the separation of the flow eluting from the center of the column from that of the flow eluting near the peripheral wall region. The flow streams from the two aforementioned regions are separated by the use of a specialized column outlet fitting. By varying the ratio of flow eluting from the center to the peripheral region it was possible to increase the efficiency of a 100 × 21 mm column by up to 57% compared to a column operated in the conventional manner. [ABSTRACT FROM AUTHOR]

Reversed phase-high performance liquid chromatography (RP-HPLC) was used to analyze fucoxanthin content of two species of Malaysian brown seaweeds,Turbinaria turbinate, andSargassum plagyophyllum. The fucoxanthin contents ofT. turbinataandS. plagyophyllumwere 0.59 ± 0.08 and 0.71 ± 0.01 mg/g dry-weight, respectively. Ultraviolet spectrum of fucoxanthin showed the maximum absorbance at 450 nm. Furthermore, the HPLC chromatograms of purified all-trans-fucoxanthin from showed one major peak with a retention time of 7.8 min. [ABSTRACT FROM AUTHOR]

In this study, a simple, rapid, and efficient purification method of five major flavonoids from crudeFlaveria bidentisextracts was established. Five flavonoids components were initially obtained by ethanol-water extraction ofFlaveria bidentis(L.) Kuntze, followed by using D4020 resin-based column chromatography and semi-preparative high performance liquid chromatography. 9.3 mg hyperoside, 6.5 mg patuletin-3-O-glucoside, 1.7 mg isorhamnetin 3-sulphate, 8.4 mg astragalin, and 4.9 mg 6-methoxykaempferol-3-O-galactoside at high purity of over 94% were obtained from 580 mgFlaveria bidentisextracts. This method is more efficient than HSCCC comparing with the results of these two methods regarding analysts’ purity, recovery, and running time. [ABSTRACT FROM PUBLISHER]

Abstract: A (1→6)-β-d-glucan (WPLE-N-1) and two mannogalactoglucans (WPLE-N-2 and WPLE-N-3) were isolated from the basidiocarps of Lentinus edodes by hot water extraction, ethanol precipitation, anion exchange chromatography, and further purified by gel-permeation chromatography (GPC). Their structural features were investigated by high performance liquid chromatography (HPLC), high performance gel-permeation chromatography (HPGPC), methylation analysis, periodate oxidation–Smith degradation, Fourier transform-infrared (FT-IR) and NMR spectroscopy, including two-dimensional (2D) NMR. HPLC analysis revealed that WPLE-N-1 was mainly composed of glucose (92%) with small amount of galactose (3.9%) and mannose (4.1%), WPLE-N-2 and WPLE-N-3 contained mannose–galactose–glucose in the molar ratio of 10:27:63 and 5:12:83, respectively. GPC and HPGPC showed that WPLE-N-1, WPLE-N-2 and WPLE-N-3 are homogeneous fractions and their molecular weights were estimated to be 757.5kDa, 20.9kDa and 4.7kDa, respectively. Chemical and spectroscopic studies indicated that WPLE-N-1 consisted of (1→6)-β-d-glucopyranosyl residues; while WPLE-N-2 and WPLE-N-3 were found to contain (1→6)-, (1→4)- and (1→3)-α-d-glucopyranosyl residues, (1→6)-α-d-galactopyranosyl residues, (1→3,6)- and (1→2,4)-α-d-mannopyranosyl residues and terminal residues of d-glucopyranosyl residues. On a preliminary bioactivity test, these three polysaccharides exhibited antitumor activity against Sarcoma S-180, Carcinoma HCT-116 and HT-29 in vitro. This finding suggests that mannogalactoglucan should be explored as potential antitumor agents and utilized as tumor cell growth inhibitors for food and pharmaceutical industries. [Copyright &y& Elsevier]

Abstract: Echinacoside and verbascoside possess a spectrum of significant beneficial bioactivities. A new method of preparative high-speed counter-current chromatography for rapid separation and purification of verbascoside from the leaves of Penstemon barbatus (Cav.) Roth was introduced. The solvent system employed here was n-butanol–water (1:1, v/v). The structure was confirmed by IR, 1H NMR and 13C NMR. According to the quantification results, echinacoside and verbascoside were found to be as much as 5.25±0.13mg/g and 11.21±0.16mg/g, respectively, in the leaves of P. barbatus. This indicates that P. barbatus might be an ideal alternative resource for large scale preparation of echinacoside and verbascoside. [Copyright &y& Elsevier]

Abstract: The confirmation by GC/C/IRMS of the exogenous origin of pseudo-endogenous steroids from human urine samples requires extracts of adequate purity. A strategy based on HPLC sample purification prior to the GC/C/IRMS analysis of human urinary endogenous androgens (i.e. testosterone, androsterone and/or androstenediols), is presented. A method without any additional derivatization step is proposed, allowing to simplify the urine pretreatment procedure, leading to extracts free of interferences permitting precise and accurate IRMS analysis, without the need of correcting the measured delta values for the contribution of the derivatizing agent. The HPLC extracts were adequately combined to both reduce the number of GC/C/IRMS runs and to have appropriate endogenous reference compounds (ERC; i.e. pregnanediol, 11-keto-etiocholanolone) on each GC–IRMS run. The purity of the extracts was assessed by their parallel analysis by gas chromatography coupled to mass spectrometry, with GC conditions identical to those of the GC/C/IRMS assay. The method has been validated according to ISO17025 requirements (within assay precision below 0.3‰13C delta units and between assay precision below 0.6‰13C delta units for most of the compounds investigated) fulfilling the World Anti-Doping Agency requirements. [Copyright &y& Elsevier]

In order to isolate large amounts of pure lutonarin and saponarin, a high performance method for isolation and purification of these two flavonoids from barley (Hordeum vulgare) seedlings was successfully established by utilizing high-speed counter-current chromatography (HSCCC). Separation was performed with a two-phase solvent system: ethyl acetate/n-butanol/water (3:2:5, v/v/v). A total of 24 mg lutonarin and 14 mg saponarin was obtained in one-step separation from 100 mg sample. High performance liquid chromatography (HPLC) analysis showed that the purity of each of the two compounds was over 98%. Their chemical structures were confirmed by UV, 1H NMR, and 13C NMR data. [ABSTRACT FROM AUTHOR]

Abstract: The enzymatic synthesis of esters is well documented in the literature. A number of parameters have been revised to optimize the operating conditions, in view of a more industrial approach. The experimental conditions presented here are now superior to those of literature and the yields of esters show 78% of fructose laurate. Analytical methods have been perfected and rapid methods (based on different TLC conditions depending on the desired information), accurate methods (based on HPLC) and several purification methods were developed. [Copyright &y& Elsevier]

A centrifugal partition chromatography (CPC) was applied to separate and purify flavonoids from the extract of Pterocarpus santalinus. Three flavonoids, taxifolin (21 mg), dihydrokaempferol (18 mg), and naringenin (16 mg) were successfully separated from 500 mg of ethyl acetate fraction of P. santalinus using centrifugal partition chromatography with a two-phase solvent system composed of n-hexane-ethyl acetate-methanol-water (4:5:4:5, v/v) in single CPC run. Eryodictyol (8 mg) and quercetin (5 mg) were further purified by HPLC. The purities of the isolated compounds were determined to be over 90% by HPLC analysis and their structures were elucidated by 1H NMR, 13C NMR, and ESI-MS. [ABSTRACT FROM AUTHOR]

Abstract: A sensitive and specific ion-pair reversed-phase high performance liquid chromatography (HPLC) method for urinary iodine analysis is described. This method is based on pulsed amperometric detection (PAD) using a silver working electrode (HPLC–PAD), which improves peak shape, electrode stability as well as linearity and reproducibility. A two-step extraction process consisting of solid phase extraction (SPE) and liquid–liquid extraction with dichloromethane was added in order to improve sample purification which is essential with the use of PAD. Treated samples were eluted on a C18 column, using a phosphate buffer containing ion-pairing reagent tetrabutylammonium and 5% MeOH. The calibration standard curves were linear up to 500μg/L and within-run and between-run coefficients of variation (CVs) were <6% with the quantification limit fixed at 6μg/L. Accuracy, expressed as recovery, ranged from 94% to 104%. Comparison with the Technicon AutoAnalyzer acid digestion (AA) method resulted in a high correlation (r=0.9916). Due to a low quantification limit and high sample throughput, the proposed technique appears suitable for both epidemiological and clinical follow-up studies. [Copyright &y& Elsevier]

Abstract: Derivatisation, solid–liquid extraction, solid-phase purification and HPLC–DAD separation is a simple, fast, and accurate method developed for the determination of dithiocarbamate (DTC) fungicides in fruits and vegetables. Quantification is based on external standard calibration curves made with DTCs-spiked blank-matrices. Limits of detection (LODs) and limits of quantitation (LOQs) are in the range of 0.01–0.3 and 0.02–0.5mg/kg, respectively. Recoveries higher than 78% were obtained for the target DTCs in all the commodities studied. The aim of this research was to evaluate the quality and determine the concentrations of DTC fungicide residues in plant material harvested from September to November of 2010. Several fruits and vegetables produced in Spain were collected at small shops and large food chains distributed by the four provinces of Galicia (North-Western Spain). In total, 150 samples were studied (32 apples, 12 wine grapes, 32 lettuces, 32 peppers, 32 tomatoes, and 10 strawberries). Residues of DTCs were found in all the products analysed with the exception of strawberries, being ethylenebis(dithiocarbamates) (EBDCs) vs. propylenebis (dithiocarbamates) (PBDCs) the main contributors to the total pesticide load. Moreover, fungicide residues exceeding maximum residue limits (MRLs) were identified in 6% of the samples analysed, specifically on lettuces and peppers. [Copyright &y& Elsevier]

Abstract: Huperzine A (HupA) and Huperzine B (HupB) are natural alkaloids existed in Lycopodium plants. They both have potential clinical application for treating Alzheimer''s Disease (AD). For the purpose of better utilizing the limited plant resources, a quick and low cost method to separate and purify HupA and HupB from Huperzia serrata (Thunb. ex Murray) was established in this paper. Low polarity macroporous resin SP850 was selected from eight kinds of resins during initial purification. Trifluoroacetic acid (TFA) was proved to be the best acid modifier reagent among all acids used in our experiment for improving separation. HupA and HupB were baseline separated on a C18 column by preparative high performance liquid chromatography (Preparative HPLC), the optimal gradient mobile phase system contained methanol increasing from 15% (v/v) to 35% (v/v) and 0.1% (v/v) TFA within the water. The purity of HupA and HupB obtained was 99.1% and 98.6%, respectively, and the total recovery for them was 83.0% and 81.8%, respectively. [Copyright &y& Elsevier]

Abstract: This study describes an optimization of [6]-, [8]- and [10]-gingerol isolation and purification in semi-preparative HPLC scale and their anti-proliferative activity. The gingerols purification was carried out in HPLC system using a Luna-C18 and the best mobile phase evaluated was MeOH/H2O (75:25, v/v). This new methodology for the gingerols isolation was very effective, since considerable amounts (in the range of milligrams) with a good purity degree (∼98%) were achieved in 30min of chromatographic run. [6]-, [8]- and [10]-Gingerol purified by this methodology inhibited the proliferation of MDA-MB-231 tumor cell line with IC50 of 666.2±134.6μM, 135.6±22.6μM and 12.1±0.3μM, respectively. These substances also inhibited human fibroblasts (HF) cell proliferation, however in concentrations starting from 500μM. In conclusion, our results demonstrate an optimization of gingerols isolation and their specific anti-proliferative activities against tumor cells, suggesting their use as important models for drug design in an attempt to develop new compounds with fewer side effects when compared to conventional chemotherapy. [Copyright &y& Elsevier]

Abstract: 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. [Copyright &y& Elsevier]