ABSTRACT
Zwitterionic sulfobetaine-based monolithic stationary phases have attracted increasing attention for their use in hydrophilic interaction chromatography.In this study,a novel hydrophilic polymeric monolith was fabricated through photo-initiated copolymerization of 3-(3-vinyl-1-imidazolio)-l-propanesulfonate(SBVI)with pentaerythritol triacrylate using methanol and tetrahydrofuran as the porogenic system.Notably,the duration for the preparation of this novel monolith was as little as 5 min,which was significantly shorter than that required for previously reported sulfobetaine-based monoliths prepared via conventional thermally initiated copolymerization.Moreover,these monoliths showed good morphology,permeability,porosity(62.4％),mechanical strength(over 15 MPa),column efficiency(51,230 plates/m),and reproducibility(relative standard deviations for all analytes were lower than 4.6％).Mechanistic studies indicated that strong hydrophilic and negative electrostatic interactions might be responsible for the retention of polar analytes on the zwitterionic SBVI-based monolith.In particular,the resulting monolith exhibited good anti-protein adhesion ability and low nonspecific protein adsorption.These excellent features seem to favor its application in bioanalysis.Therefore,the novel zwitterionic sulfobetaine-based monolith was successfully employed for the highly selective separation of small bioactive compounds and the efficient enrichment of N-glycopeptides from complex samples.In this study,we prepared a novel zwitterionic sulfobetaine-based monolith with good performance and developed a simpler and faster method for preparation of zwitterionic monoliths.
ABSTRACT
The chemical constituents of classical prescription Danggui Buxue Decoction were analyzed by reversed-phase(RP) chromatography and hydrophilic interaction chromatography(HILIC) coupled with quadrupole time-of-flight mass spectrometry. RP separation of Danggui Buxue Decoction was performed on ACQUITY UPLC HSS T3(2.1 mm×100 mm, 1.8 μm), while HILIC separation was on Waters BEH Amide(2.1 mm×100 mm, 1.7 μm). Mass spectrometry(MS) data were acquired in both negative and positive ion modes. Chemical constituents of Astragali Radix and Angelicae Sinensis Radix were searched from Reaxys and thus the in-house library was established. MS data were further analyzed by MassLynx 4.1 combined with in-house library, HMDB, Reaxys, and comparison with reference substances. In conclusion, a total of 154 compounds were identified and characterized: 16 saponins, 44 flavonoids, 10 phthalides, 7 phenylpropanoids, 15 bases and the corresponding nucleosides, 30 oligosaccharides, and 32 other compounds. Among them, 65 compounds were detected by HILIC-MS/MS. This study provides experimental evidences for the material basis research, quality control, and preparation development of Danggui Buxue Decoction and a reference method for comprehensive characterization of Chinese medicine decoctions typified by classical prescriptions.
Subject(s)
Chromatography, High Pressure Liquid/methods , Drugs, Chinese Herbal/chemistry , Prescriptions , Tandem Mass SpectrometryABSTRACT
Objective: The characteristic maps and immunological activities of some acid hydrolysates of polysaccharides from Astragali Radix (APS) from different areas were compared. The quality evaluation method of Astragali Radix with oligosaccharide mixture as quality control index was established. Methods: In this study, the polysaccharides from the traditional Chinese medicine Astragali Radix were used as the research object. Firstly, the optimal partial acid hydrolysis conditions were selected by orthogonal test. The polysaccharide was hydrolyzed into oligosaccharides for analysis. The characteristic map of Astragalus oligosaccharides based on partial acid hydrolysis-hydrophilic interaction chromatography was established. Multivariate statistical analysis was performed on the data by SIMCA software to distinguish Mongolian Astragalus from different areas. The partial acid hydrolysis products of Astragalus polysaccharides were characterized by hydrophilic interaction chromatography and mass spectrometry, and the activity was evaluated by mouse peritoneal macrophage phagocytosis neutral red experiment. Results: The optimal hydrolysis conditions obtained by orthogonal experiment were temperature 90 ℃, trifluoroacetic acid concentration 1 mol/L, hydrolysis time 1 h. Under this condition, Astragalus polysaccharide was hydrolyzed into characteristic oligosaccharide fragments, and the method is reproducible. The characteristic map of Astragalus oligosaccharides based on partial acid hydrolysis-hydrophilic interaction chromatography showed that the maps of the same Astragalus oligosaccharides had good consistency, and the maps of different Astragalus oligosaccharides were quite different. PCA showed that three different kinds of Mongolian Astragalus can be distinguished. It was found by mass spectrometry that the extracted Astragalus polysaccharides were mainly 1→4 linear glucan, and gluco-oligosac-charides with the degrees of polymerization 3—8 were obtained after partial acid hydrolysis. The partial acid hydrolysate of the wild Astragalus polysaccharides from Shanxi Hunyuan had higher ability to enhance the phagocytic activity of macrophages than the transplanted Astragalus and higher than the unhydrolyzed total astragalus polysaccharide. Conclusion: This study showed that the characteristics of Astragalus polysaccharides based on partial acid hydrolysis-hydrophilic interaction chromatography and the effects on cellular immune function can be used to evaluate the quality of Astragali Radix in different habitats and different planting methods, and it is also an important supplement to the quality evaluation method of Astragali Radix. At the same time, it has a certain exemplary role in the characterization of other Chinese materia medica polysaccharides.
ABSTRACT
Aryloxypropanolamine is an essential structural scaffold for a variety of β-adrenergic receptor antago-nists such as metoprolol. Molecules with such a structural motif tend to degrade into α, β-hydroxypropanolamine impurities via a radical-initiated oxidation pathway. These impurities are typically polar and nonchromophoric, and are thus often overlooked using traditional reversed phase chromatography and UV detection. In this work, stress testing of metoprolol confirmed the generation of 3-isopropylamino-1,2-propanediol as a degradation product, which is a specified impurity of metoprolol in the European Pharmacopoeia (impurity N). To ensure the safety and quality of metoprolol drug products, hydrophilic interaction chromatography (HILIC) methods using Halo Penta HILIC column (150 mm×4.6 mm, 5 μm) coupled with charged aerosol detection (CAD) were developed and optimized for the separation and quantitation of metoprolol impurity N in metoprolol drug products including metoprolol tartrate injection, metoprolol tartrate tablets, and metoprolol succinate extended-release tablets. These HILIC-CAD methods were validated per USP validation guidelines with respect to speci-ficity, linearity, accuracy, and precision, and have been successfully applied to determine impurity N in metoprolol drug products.
ABSTRACT
A metabolic profiling analysis method for metabolomic studies of rice leaf was established based on HSS T3 combined with XBridge Amide Q-TOF LC/MS by comparing the influences of different extraction methods in rice leaves of metabolites. The extraction and separation of rice leaf metabolites using three different methods including methanol-chloroform-water,methanol-chloroform-ammonia,methanol-methyl tert-butyl ether -water and different chromatographic systems were compared by the numbers of peaks, identified metabolites and the metabolic pathways. The results showed that the method of methanol-chloroform-water reached the highest coverage rate of metabolites in rice leaves,and the maximum number of unique metabolites including prephenic acid, luteolin, α-linolenic acid, aconitic acid, gibberellin A12 aldehyde, isovitexin, L-Glutamate were detected. Metabolites with different polarity in rice leaf could be detected by HSS T3 and XBridge Amide. A total of 16 kinds of organic acids, 17 kinds of nucleotides, 21 kinds of amino acids, 66 kinds of fatty acids,11 kinds of phospholipids and 7 kinds of sphingolipids were identified. XBridge Amide had an absolute advantage in detecting phospholipids and sphingolipids. The metabolic pathways involved purine metabolism, pyrimidine metabolism, tricarboxylic acid cycle, arginine metabolism, fatty acid metabolism, phospholipid metabolism, sphingolipid metabolism, phenylalanine metabolism and vitamin B2 synthesis. It showed certain complementarity between the two columns in identifying metabolites and involved the metabolic pathways. The established method is expected to be useful for the metabolomic studies of rice.
ABSTRACT
In this study, a novel tetrapeptide hydrophilic interaction chromatography (HILIC) material was synthesized, and corresponding enrichment method for glycopeptides was developed.The tetrapeptide modified silica gel materials (denoted as Poly-DAPD) were synthesized by atom-transfer radical-polymerization (ATRP) and characterized by N2 adsorption desorption and thermometer, thermal gravimetric analyzer (TGA) and X-ray photoelectron spectrometer (XPS).The characterization results indicated that tetrapeptide had been successfully synthesized on silica gel.Poly-DAPD materials showed high enrichment selectivity toward fetuin glycopeptides under solid-phase extraction (SPE) mode.Comparing with commercialized ZIC-HILIC in glycopeptides enrichment of fetuin digest which mixed with 5 mole ratio of albumin bovine (BSA), the as-prepared materials showed higher selectivity in both aspects of the identified number of glycopeptides and anti-interference property.The SPE results demonstrated that the tetrapeptide-based HILIC materials could be a potential tool in large-scale glycosylation analysis.
ABSTRACT
2-(Dimethylamino)ethyl methacrylate and sodium monochloroacetate were employed to synthesize [2-(Methacryloyloxy)ethyl] dimethyl ammonium acetate (CBMA) functional monomer.CBMA was grafted on the surface of silica by surface initiated atom transfer radical polymerization (SI-ATRP) to obtain silica-CBMA hydrophilic interaction stationary phase.Three silica-CBMA stationary phases with different grafted density of CBMA monomer were synthesized in SI-ATRP progress by changing the concentration of CBMA under the same conditions.The ability to separate organic acid compounds of the synthesized silica-CBMA stationary phases was evaluated under different conditions, including effects of pH value, salt concentration and content of water of mobile phase on retention of solutes.The results showed that the stationary phases could effectively separate organic acid compounds in HILIC mode, which followed a mixed mode of chromatography of ion exchange and hydrophilic interaction.The retention of solutes decreased with the increases of salt concentration of mobile phase, which consistent with the characteristics of ion exchange;the pH value of mobile phase had significant influence on ionization of the stationary phase and solutes, i.e., the retention of solutes increased as the increasing of pH value of mobile phase.However, the retention of solutes decreased with the increasing of the content of water in mobile phase, which was the typical characteristic of HILIC.The method of hydrophilic interaction chromatography combined with silica-CBMA stationary phases could conveniently determinate the content of vitamin C and rutin in rutin tablets, providing a new method for the separation and determination of strong polar samples.
ABSTRACT
Organic acids are widely distributed in plants and related products, and participate in a wide range of metabolic pathways (e.g. tricarboxylic acid cycle), showing diverse pharmacological activities. As a widely used Chinese patent medicine, its adverse reactions are often reported. Therefore, we should further clarify the chemical components of Shenfu injection, and prepare strict quality standards to ensure the safety and effectiveness of its clinical use. Shenfu injection is prepared from red ginseng (steamed roots of Panax ginseng) and black prepared lateral roots of Aconitum carmichaelii (Heishunpian) by using modern extraction process, and organic acids are regarded as one of its main components. In current study, a hydrophilic interaction chromatography (HILIC) coupled with mass spectrometric method (HILIC-LC-MS) was developed and validated for the simultaneous determination of 14 organic acids, including cinnamic acid, ferulic acid, 4-hydroxylbenzoic acid, L-(+)-lactic acid, adipic acid, fumaric acid, caffeic acid, succinic acid, maleic acid, malonic acid, D-malic acid, (-)-shikimic acid, D-tartaric acid, and quinic acid in Shenfu injection. Satisfactory retention and separation were achieved for all organic acids on HILIC chromatographic column. Except cinnamic acid (231 μg•L⁻¹), lactic acid (113 μg•L⁻¹) and malonic acid (32.5 μg•L⁻¹), the limit of quantitation for the remaining 11 compounds were less than 10 μg•L⁻¹. D-Malic acid, malonic acid, quinic acid, L-(+)-lactic acid, and cinnamic acid were observed to have higher contents in Shenfu injection (>1.89 mg•L⁻¹), whereas caffeic acid and adipic acid were undetectable in all batches. Above all, the developed method is suitable for the simultaneous determination of organic acids in Shenfu and some other traditional Chinese medicine injections.
ABSTRACT
Objective: Daidzein was efficiently purified by agar gel microspheres bonded β-cyclodextrin (AG-β-CD). Methods: Using agar as raw material, after emulsification, crosslinking, and bonding β-CD as functional group, AG-β-CD was synthesized for the purification of daidzein, and the purification process was determined and proved with mobile phase, flow rate, and loading capacity of microspheres. The structure of daidzein was identified by MS and NMR, AG-β-CD was chromatographically evaluated with daidzein, EGCG, and puerarin as the following tripartition such as difference of retention behavior on C18 reversed phase column chromatography, molecular simulation by autoDOCK4.0, and retention time curves on AG-β-CD with different contents of acetonitrile. Results: The main component of soybean isoflavone was daidzein (57.14%). The loading quantity of AG-β-CD was 1.33 mg/mL, flow rate was 2 BV/h, eluted by 2 BV of 20% ethanol, 1.33 BV of 40% ethanol, and 6-7 BV of 70% ethanol, the content was ≥ 95%, purity of daidzein (96.98%) was obtained with 97.86% yield. Chromatographic mechanism research showed that AG-β-CD had hydrophilic interaction chromatography and reversed-phase chromatography. Conclusion: AG-β-CD is capable of highly efficient purification of daidzein.
ABSTRACT
Lipidomics is a new branch discipline of metabolism. The hydrophilic interaction chromatography-mass spectrometry (HILIC-MS) technology is the new study method of lipidomics. The application, operation points, special advantages, and development potential for HILIC-MS were clarified from the angles of biological sample processing method, chromatography, mass spectrometry conditions establishment, and full analysis on quality control. The feasibility of using HILIC-MS technology to solve some lipid metabolism diseases problem in clinical and scientific research of TCM has been investigated.
ABSTRACT
A method for rapid determination of semicarbazide in water by hydrophilic interaction chromatography-quadrupole / electrostatic field orbitrap high resolution mass spectrometry was developed. The sample was extracted with acetonitrile after 0. 1 mol/ L NaOH was added in the sample and then excessive amounts of Na2 SO4 was added to stratify acetonitrile from the mix solution. The acetonitrile extraction solution was dehydrated with anhydrous sodium sulfate. The preparation was separated by an amide column using as hydrophilic interaction column, and gradient elution program was employed by using water and acetonitrile containing 0. 1% formic acid as mobile phase, then it was detected in positive and selected ion monitoring mode by a quadrupole / electrostatic field orbitrap high resolution mass spectrometry. Internal standard method was used for quantitative analysis. The linear correlation coefficient of semicarbazide was 0. 997 in the concentration range of 0. 2 -20 μg / L under the optimal conditions. The limit of detection was 0. 09 μg / L, while the limit of quantitation was 0. 30 μg / L. The recoveries were 82. 3% to 92. 0% , and the relatively standard deviations were less than 7. 6% at the spiked levels of 0. 5, 1. 0 and 5. 0 μg / L using river water and sea water as blank samples. The developed method is suitable for the analysis of trace semicarbazide in environment water samples.
ABSTRACT
Objective: To develop a new method based on hydrophilic interaction chromatography-electrospray ionization-time of flight-mass spectrometry (HILIC-DAD-ESI-TOF/MS) for the rapid identification of the active components in Syngnathus acus and the development of their specific fingerprint chromatograms. Methods: Samples were extracted by accelerated solvent extraction, and the extraction conditions were optimized. The developed HILIC-DAD-ESI-TOF/MS method was used to identify the components in water extract from S. acus, and a chromatographic fingerprint based on HILIC analysis was established. Results: Ten compounds in S. acus extract could be primarily identified by HILIC-DAD-ESI-TOF/MS on-line detection, in which seven nucleosides were determined. The HPLC characteristic fingerprint was established on the basis of analysis on the multi batches of S. acus, which could be used to evaluate the quality of S. acus combined with similarity calculation. Conclusion: This method is simple and rapid, and is a powerful tool for the identification of S. acus.