Huatao Feng

Determination of five toxic alkaloids in two common herbal medicines with capillary electrophoresis

A method was developed for the determination of five highly toxic alkaloids in two commonly used herbal medicines by capillary electrophoresis, which had not been applied to the determination of Aconitum alkaloids before. The buffer contained 40 mM ammonium acetate and 0.1% acetic acid in 80% methanol. Five alkaloids can be determined in 15 min by a single run. The calibration curves showed a linear range from 2 to 200 mg/l for these alkaloids with correlation coefficients (R2) between 0.9988 and 0.9999. Detection limits (SIN = 3) varied from 0.85 to 1.90 mg/l. Recoveries ranged from 95 to 108.8%. The method can provide an effective tool for the strict control of these fetal herbal medicine components.

Analysis of Chinese medicine preparations by capillary electrophoresis-mass spectrometry

In Chinese medicines, herbs are usually prepared before use by patients. Since the preparation procedures convert the original component into one or more products, study of the procedures is usually complex and involves several compounds. On-line coupling of capillary electrophoresis (CE) to mass spectrometry (MS) allows both the efficient separation of CE and the specific and sensitive detection of MS to be achieved. In this study, CE-MS was applied to the determination of alkaloids in Maqianzi (the seed of Strychnos pierrian) and Wutou (aconite root, Radix aconiti praeparata) during the preparation procedure. With optimal CE-MS conditions, alkaloids in both prepared and unprepared Maqianzi were determined successfully in the total ion current (TIC) mode. However, single ion monitoring (SIM) had to be applied for the separation of aconitum alkaloids and their hydrolysis products. Quantification data indicated that MS detection under SIM mode is more sensitive than UV detection. Based on the CE-MS method developed, the hydrolysis of aconitum alkaloids in water and methanol was also studied.

Sensitive determination of trace-metal elements in tea with capillary electrophoresis by using chelating agent 4-(2-pyridylazo) resorcinol (PAR)

A capillary electrophoresis method was developed for the determination of trace metals (Cu, Fe, Zn, Co and Ni) in tea based on the conversions of strong complexes between 4-(2-pyridylazo) resorcinol (PAR) and metals. Five metals can be determined in 10 min by a single run. The calibration curves showed a linear range from 50 μg/l to 5 mg/l for these metals with correlation coefficients (r2) being greater than 0.999. Detection limits (S/N=3) of Co, Fe, Cu, Zn and Ni varied from 6 to 30 μg/l. Recoveries of these metal ions ranged from 97.9 to 106.2%. The method is simple and separation time is short.

An efficient “off–on” carbon nanoparticle-based fluorescent sensor for recognition of chromium(VI) and ascorbic acid based on the inner filter effect

A simple approach based on calcination treatment of diethylenetriaminepentaacetic acid (DTPA) was developed to prepare water-soluble nitrogen doped carbon nanoparticles (N-CNPs) with a high quantum yield of approximately 53.7%. The fluorescence of N-CNPs could be quickly and efficiently quenched by Cr(vi) rather than Cr(iii) based on an inner filter effect (IFE) process. The addition of ascorbic acid (AA) can recover the intensity of fluorescence of the N-CNP-Cr(vi) system through the reduction of Cr(vi) to Cr(iii) and inhibit the IFE process between N-CNPs and Cr(vi) (turn-on). Accordingly, an efficient N-CNP based fluorescent probe for sensitive and selective sensing of Cr(vi) ions and l-ascorbic acid (AA) has been established. The proposed fluorescence sensor displays excellent performance for Cr(vi) determination in the range from 0.5 to 160 μmol L-1 (R2 = 0.998) with a detection limit down to 0.15 μmol L-1. Moreover, the observed linear response concentration range was from 1 to 400 μmol L-1 for AA with a detection limit as low as 0.13 μmol L-1. The fluorescent probe was successfully applied to detect Cr(vi) concentration in different water samples and AA concentration in human serum samples.

Indirect capillary electrophoresis with 8-anilino-1-naphthalenesulfonic acid as a fluorescence probe for determining the apparent stability constant of an inclusion complex formed between a cyclodextrin and a solute

An indirect capillary electrophoresis (CE) method was developed based on two competitive chemical equilibria for determining the stability constant of an inclusion complex formed between a cyclodextrin and a solute. 8-Anilino-1-naphthalenesulfonic acid was employed as a fluorescence probe. A linear relationship between mobility difference and concentration of uncomplexed ligand was theoretically established and experimentally verified. The principle of the method was explained using an example of determining stability constant of an inclusion complex formed between a ligand of hydroxypropyl-beta-cyclodextrin and a solute of amantadine. The stability constant was determined to be approximately 2 x 10(2) M(-1). It was calculated without knowledge of the mobility of the complex measured at saturating ligand concentrations. This indirect method can be applied to solutes and ligands lacking signal response on the selected detector in the CE. In addition, the indirect method is valid for both charged and neutral solutes and ligands.

Detection of Proteins by On‐Column, Non‐Covalent Labeling with NanoOrange During Capillary Zone Electrophoresis

Abstract To avoid prior derivatization of proteins with fluorescent reagents, on‐column labeling of non‐covalent dye NanoOrange on proteins was evaluated with sodium dodecyl sulfate (SDS) as an additive using laser induced fluorescence (LIF) monitoring in capillary electrophoresis (CE). Performance of NanoOrange in buffer solutions of various pH values was tested. Acidic buffer solution was found to be unsuitable for on‐column NanoOrange labeling. SDS enhanced the fluorescence intensities of basic proteins with high pI values. The fluorescence intensities of protein solutions with or without SDS additives were compared using a fluorescence spectrophotometer. The applicability of on‐column labeling and detection of staphylococcal enterotoxin B was also demonstrated. This on‐column labeling method provides a rapid, direct, widely applicable technique for protein studies.

Multiplexing N-glycan analysis by DNA analyzer

Analysis of N‐glycan structures has been gaining attentions over the years due to their critical importance to biopharma‐based applications and growing roles in biological research. Glycan profiling is also critical to the development of biosimilar drugs. The detailed characterization of N‐glycosylation is mandatory because it is a nontemplate driven process and that significantly influences critical properties such as bio‐safety and bio‐activity. The ability to comprehensively characterize highly complex mixtures of N‐glycans has been analytically challenging and stimulating because of the difficulties in both the structure complexity and time‐consuming sample pretreatment procedures. CE‐LIF is one of the typical techniques for N‐glycan analysis due to its high separation efficiency. In this paper, a 16‐capillary DNA analyzer was coupled with a magnetic bead glycan purification method to accelerate the sample preparation procedure and therefore increase N‐glycan assay throughput. Routinely, the labeling dye used for CE‐LIF is 8‐aminopyrene‐1,3,6‐trisulfonic acid, while the typical identification method involves matching migration times with database entries. Two new fluorescent dyes were used to either cross‐validate and increase the glycan identification precision or simplify sample preparation steps. Exoglycosidase studies were carried out using neuramididase, galactosidase, and fucosidase to confirm the results of three dye cross‐validation. The optimized method combines the parallel separation capacity of multiple‐capillary separation with three labeling dyes, magnetic bead assisted preparation, and exoglycosidase treatment to allow rapid and accurate analysis of N‐glycans. These new methods provided enough useful structural information to permit N‐glycan structure elucidation with only one sample injection.

Rapid detection of an anthrax biomarker based on the recovered fluorescence of carbon dot–Cu(II) systems

The sensitive detection of dipicolinic acid (DPA) is strongly associated with the sensing of bacterial organisms in environmental and biological samples. In this work, we developed a novel and sensitive label-free sensor for the detection of dipicolinic acid (DPA), a biomarker of the biological threat-agent anthrax, using an “off–on” fluorescent carbon dot (CD)–Cu(II) system. Fluorescent CDs with a quantum yield of 24.7% were prepared by a simple one-pot hydrothermal treatment of dopamine in acidic conditions (pH 5.0) under nitrogen gas. As the prepared CDs have two fluorescent centers at a single-wavelength excitation of 315 nm at low concentration, a novel ratiometric fluorescent sensor for the determination of DPA was designed to facilitate self-calibration by using one center as the internal reference and another center as the signal reporter. The fluorescence of the CDs was found to be effectively quenched by Cu(II), and subsequently restored with high sensitivity in the presence of DPA via an “off–on” fluorescence response due to the strong chelate-conjugation between Cu(II) and DPA. Under optimal conditions, good linearity for DPA detection was obtained over the concentration range from 0.25 to 20 μmoL L−1, with a detection limit of 0.079 μmoL L−1. The proposed simple, fast and convenient method was applied for DPA detection in different types of environmental water samples and fetal bovine serum samples, and the results are in agreement with the results of standard additions in an acceptable recovery range.

Parallel analysis and orthogonal identification of N-glycans with different capillary electrophoresis mechanisms

The deep involvement of glycans or carbohydrate moieties in biological processes makes glycan patterns an important direction for the clinical and medicine researches. A multiplexing CE mapping method for glycan analysis was developed in this study. By applying different CE separation mechanisms, the potential of combined parallel applications of capillary zone electrophoresis (CZE), micellar electrokinetic chromatography (MEKC) and capillary gel electrophoresis (CGE) for rapid and accurate identification of glycan was investigated. The combination of CZE and MEKC demonstrated enhancing chromatography separation capacity without the compromises of sample pre-treatment and glycan concentration. The separation mechanisms for multiplexing platform were selected based on the orthogonalities of the separation of glycan standards. MEKC method exhibited promising ability for the analysis of small GU value glycans and thus complementing the unavailability of CZE. The method established required only small amount of samples, simple instrument and single fluorescent labelling for sensitive detection. This integrated method can be used to search important glycan patterns appearing in biopharmaceutical products and other glycoproteins with clinical importance.

Extraction, separation and characterization of endotoxins in water samples using solid phase extraction and capillary electrophoresis-laser induced fluorescence.

This study focuses on one of the key environmental threats, endotoxins, also known as lipopolysaccharides (LPS). A capillary electrophoresis method in combination with laser induced fluorescence (LIF) detection was developed for the analysis of endotoxins from 16 different bacterial strains. LPSs were derivatized with the amino-reactive fluorescent dye, fluorescein isothiocyanate (FITC), separated by capillary zone electrophoresis (CZE) under the optimized conditions with the use of 50 mM sodium tetraborate buffer (pH 9.30), and detected by LIF detector. To improve the sensitivity of CZE-LIF detection for the determination of trace amounts of endotoxins and to remove possible interference materials in environmental samples, a solid phase extraction (SPE) pre-concentration technique was applied successfully. The SPE targeted at polysaccharide moieties of LPSs and showed LPS enrichment effects too. CE migration time could also reveal the O-antigen chain lengths of LPSs. This CE method and SPE pretreatment showed linearity at 99.84%, and repeatabilities at 8.44% and 11.0% for endotoxins from E. Coli O55:B5 and E. Coli O26:B6. The limit of detection (LOD) could reach around 5 ng/mL at optimized condition. The method was applied successfully to the determination of LPS levels in tap water and wastewater, and demonstrated sensitive, reproducible and reliable results.