► Different 2D-LC modes were combined for the analysis of
Scutellaria barbata D. Don. ► 206 peaks were detected in polar fraction by 2-D HILIC
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HILIC. ► 543 peaks were detected in medium-polar ...fraction by 2-D HILIC
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RP-LC.
Two-dimensional liquid chromatography (2-D LC) has been widely used for the analysis of complex samples owing to its great improvement in separation selectivity and peak capacity. However, one 2-D LC system may not be enough to meet the separation requirements due to the complexity of certain samples and respective limitations of two separation modes. In this work, water extract of
Scutellaria barbata D. Don, a traditional Chinese medicine, was fractionated into polar fraction and medium-polar fraction by means of solid phase extraction (SPE). The fraction preparation made it easy to select the corresponding combination of 2-D LC method from hydrophilic interaction chromatography (HILIC) and reversed-phase liquid chromatography (RP-LC). An off-line 2-D HILIC
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HILIC to analyze the polar fraction and an off-line 2-D HILIC
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RP-LC to analyze the medium-polar fraction were developed, respectively. In total, 749 peaks were detected: 206 peaks from the polar fraction by the 2-D HILIC
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HILIC and 543 from the medium-polar fraction by the 2-D HILIC
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RP-LC. The practical peak capacities obtained in both systems were 2698 and 2879, and the orthogonality reached 63.18% and 90.62%, respectively. The results demonstrated that the two systems were both highly orthogonal, and the peak capacities greatly increased.
In this study, an uncommon enzymatic-fingerprinting workflow, was proposed for characterization and discrimination of mushroom polysaccharides (MPs) by hydrophilic interaction liquid ...chromatography-negative electrospray mass spectrometry (HILIC-ESI–-MS). Firstly, the HILIC-ESI–-MS was used to screen and identify the enzymatic digestion products of MPs using HILIC-Orbitrap based on full scan and MS/MS modes. Secondly, a targeted structural-fingerprinting of polysaccharides (SFP) was built in a multiple-ion monitoring (MIM) mode using the same HILIC separation with a triple quadrupole MS. Thirdly, a case study of polysaccharides in Hericium erinaceus fruiting bodies (HEP) was performed to obtain the expected SFP based on dextranase digestion that allows for visual discrimination of polysaccharides from other five edible mushrooms attributed to Agrocybe cylindracea, Arimillaria mellea, Flammulina velutipes, Pleurotus eryngii, and Lentinula edodes. Furthermore, a major structural backbone of HEP was unveiled by occurrence of → 6(Hex)1 → along with multiple possible substitutions including of terminal GalA, Fuc, acetyl, → 4Hex1 →, and → 3Hex1 →. Finally, the similarity analysis, hierarchical cluster analysis (HCA), and partial least squares discriminant analysis (PLS-DA) were performed to visualize various MPs. As a result, the enzymatic-fingerprinting workflow presents an effective example for quality evaluation of fungi polysaccharides using a SFP strategy.
•An enzymatic-fingerprinting workflow was designed for discrimination of various MPs.•The expected SFP was established by full scan MS screening to targeted MIM profiling.•The main structural backbone of HEP was proposed using HILIC-ESI--MS/MS.•Similarity analysis, HCA, and PLS-DA were performed to visualize various MPs.
•The preparation strategy of DAC-based mixed-mode stationary phases was proposed.•The both columns were prepared by utilizing the easy functionalization of DAC.•The HILIC/IEC column and ...HILIC/IEC/chiral separation column were developed.•The both columns had good chromatographic performance in each separation mode.
A new strategy for the preparation of mixed-mode chromatographic stationary phases based on modified dialdehyde cellulose was proposed. Two novel mixed-mode chromatographic stationary phases, dicarboxyl cellulose-modified silica (DCC/SiO2) and (S)-α-phenylethylamine-bonded DCC/SiO2 ((S)-α-PEA/DCC/SiO2), were prepared by utilizing the easy functionalization characteristics of dialdehyde cellulose. The chromatographic evaluation showed that DCC/SiO2 column could be used in hydrophilic interaction liquid chromatography (HILIC) and ion exchange chromatography (IEC) modes, (S)-α-PEA/DCC/SiO2 column could be used in HILIC, IEC and chiral separation modes. The DCC/SiO2 column and (S)-α-PEA/DCC/SiO2 column exhibited excellent chromatographic performance by separating strongly polar compounds, phenylamines and chiral compounds in the above separation modes. The preparation method of modified dialdehyde cellulose-based mixed-mode chromatographic stationary phases was simple, and also provided a new idea for the development of the subsequent novel mixed-mode chromatographic stationary phases.
•Quantitative determination of 19 paralytic shellfish toxins in shellfish.•Novel shellfish matrix clean-up using non-porous graphitised carbon SPE.•Rapid (11min) UPLC HILIC–MS/MS method for PSP ...monitoring.•Chromatographic resolution of toxin epimer pairs such as GTX1 and GTX4.
Routine regulatory monitoring of paralytic shellfish toxins (PST) commonly employs oxidative derivitisation and complex liquid chromatography fluorescence detection methods (LC-FL). The pre-column oxidation LC-FL method is currently implemented in New Zealand and the United Kingdom. When using this method positive samples are fractionated and two different oxidations are required to confirm the identity and quantity of each PST analogue present. There is a need for alternative methods that are simpler, provide faster turnaround times and have improved detection limits. Hydrophilic interaction liquid chromatography (HILIC) HPLC–MS/MS analysis of PST has been used for research purposes, but high detection limits and substantial sample matrix issues have prevented it from becoming a viable alternative for routine monitoring purposes. We have developed a HILIC UPLC–MS/MS method for paralytic shellfish toxins with an optimised desalting clean-up procedure on inexpensive carbon solid phase extraction cartridges for reduction of matrix interferences. This represents a major technical breakthrough and allows sensitive, selective and rapid analysis of paralytic shellfish toxins from a variety of sample types, including many commercially produced bivalve molluscan shellfish species. Additionally, this analytical approach avoids the need for complex calculations to determine sample toxicity, as unlike other methods each PST analogue is able to be quantified as a single resolved peak. This article presents the method development and optimisation information. A thorough single laboratory validation study has subsequently been performed and this data will be presented elsewhere.
•Fe3O4-CG nanospheres was synthesized via one-step hydrothermal strategy.•Fe3O4-CG nanospheres showed high selectivity, sensitivity and stable reusability.•180 glycopeptides were identified by ...Fe3O4-CG nanospheres.•Fe3O4-CG nanospheres had fine potential for early diagnosis of Alzheimer's disease.
A novel ultra-hydrophilic zwitterionic-HILIC (ZIC-HILIC) nanosphere (Fe3O4-CG) was synthesized via a one-step hydrothermal strategy, which significantly simplified the conventional multi-step procedures for the preparation of ZIC-HILIC materials. The dual-functional Fe3O4-CG nanosphere exhibited excellent selectivity (molar ratio BSA:HRP = 5000:1), low detection limit (0.05 fmol/μL), satisfactory reusability (at least 5 times) and recovery rate (93.7 ± 2.1%). The binding constant of Fe3O4-CG for HRP is 2.45 ± 0.32 × 10−6 M and the theoretical binding capacity is 330 mg g−1. In addition, the Fe3O4-CG microsphere showed excellent performance in the detection of glycopeptides from real biological samples. Furthermore, 131 glycopeptides related to 71 glycoproteins were selectively enriched from healthy human serum and 180 glycopeptides related to 82 glycoproteins were captured from Alzheimer's disease patients' serum analyzed by Nano-LC-MS/MS. Gene ontology analysis of the biological process and molecular function showed that 21 primitive glycoproteins in glycopeptides captured from Alzheimer's disease patients' serum were meaningfully involved in a variety of neurodegenerative disease-related events, including serine-type endopeptidase inhibitor activity, receptor binding, positive regulation of B cell activation, and platelet activation.
Paralytic shellfish toxins (PST) and tetrodotoxin (TTX) are naturally-occurring toxins that may contaminate the food chain, inducing similar neurological symptoms in humans. They are co-extracted ...under the same conditions and thus their combined detection is desirable. Whilst PST are regulated and officially monitored in Europe, more data on TTX occurrence in bivalves and gastropods are needed before meaningful regulations can be established. In this study, we used three separate analytical methods - pre-column oxidation with liquid chromatography and fluorescence detection, ultrahigh performance hydrophilic interaction liquid chromatography (HILIC) tandem mass spectrometry (MS/MS) and HILIC high resolution (HR) MS/MS - to investigate the presence of PST and TTX in seawater and shellfish (mussels, clams) collected in spring summer 2015 to 2017 in the Mediterranean Sea. Samples were collected at 10 sites in the Syracuse Bay (Sicily, Italy) in concomitance with a mixed bloom of Alexandrium minutum and A. pacificum. A very high PST contamination in mussels emerged, unprecedentedly found in Italy, with maximum total concentration of 10851 μg saxitoxin equivalents per kg of shellfish tissue measured in 2016. In addition, for the first time TTX was detected in Italy in most of the analysed samples in the range 0.8–6.4 μg TTX eq/kg. The recurring blooms of PST-producing species over the 3-year period, the high PST levels and the first finding of TTX in mussels from the Syracuse bay, suggest that monitoring programmes of PST and TTX in seafood should be activated in this geographical area.
•Recurring blooms of Alexandrium minutum and Alexandrium pacificum in Syracuse bay.•Toxin profile and content of seawater and shellfish collected in 2015, 2016, 2017.•Paralytic shellfish toxins contaminating mussels at levels up to 10851 μg STX eq/Kg.•First finding in Italy of tetrodotoxin in mussels at levels up to 6.4 μg TTX eq/Kg.•Comparison among PreCOX-HPLC-FLD, UHPLC-HILIC-MS/MS, and HPLC-HILIC-HRMS for PST.
•A sensitive HILIC-ESI-MS/MS method was developed for aqueous environmental samples.•Freeze-drying was used for the first time as a sample preparation in HILIC.•Eluent and diluent composition must be ...strictly controlled to ensure reproducibility.•Co-elution of chloride or nitrate with the analytes can cause positive matrix effects.
Benefits and limitations of HILIC were studied for the analysis of extreme polar organic contaminants in aqueous environmental matrices. A sensitive analytical method was developed and validated for the detection of 11 pharmaceuticals, 15 pharmaceutical metabolites and transformation products and the artificial sweetener acesulfame in aqueous environmental samples. The analytical method consisted of a simple and non-specific sample preparation based on freeze-drying followed by detection with large injection volume (70 μL) zwitterionic HILIC-ESI-MS/MS. Robustness studies showed a high sensitivity of the retention times and peak shapes to variations of the acetonitrile/water ratio of both the eluent and the diluent. Thus, a thorough sample and eluent preparation is required to obtain reproducible results. Extreme matrix effects of >200% were observed for emtricitabine and acyclovir, which could be traced to the co-elution of nitrate and chloride, respectively. These matrix effects and those of other analytes could be efficiently compensated by using deuterated, 13C and 15N-labeled internal standards. The developed method was able to detect the selected 27 analytes in treated wastewater, surface water and groundwater down to limit of quantification (LOQ) in the lower ng/L range. Appreciable concentrations were detected, ranging up to 110 μg/L (guanyl urea) in treated wastewater, up to 5.1 μg/L (oxipurinol) in surface water and up to 6.1 μg/L (acesulfame) in groundwater. In a German drinking water, only the X-ray contrast medium diatrizoate and the artificial sweetener acesulfame were quantified above LOQ with 0.19 μg/L and 0.35 μg/L, respectively.
Hydrophilic interaction liquid chromatography (HILIC) has gained widespread acceptance as an alternative to reversed-phase liquid chromatography (RPLC) for the retention and separation of polar ...compounds. Because a great number of emerging organic contaminants are polar, this represents progress in terms of solving most of the problems and limitations encountered in the conventional methods developed for their determination. In this review, recent HILIC applications developed in the environmental field are described, which are normally coupled with mass spectrometry, in order to reach the limits required to quantify contaminants present at trace levels and benefit from its capacity for confirmation. Special attention is paid to the stationary and mobile phases commonly used in terms of the advantages that they offer compared to reversed-phase columns and the high-water content mobile phases traditionally used in RPLC. The most important features related to the matrices and contaminants normally studied are also discussed, as well as the contribution of the HILIC mode towards reducing the matrix effect.
•Recent trends in the applications of HILIC-MS for environmental samples analysis.•Discussion of commonly used stationary phases and mobile phase conditions.•Advantages of HILIC vs RPLC. Combination of HILIC and RPLC phases.•Examples of HILIC-MS in environmental field covering different analytes.
•The benefits of comprehensive two-dimensional LC are discussed.•Analytical challenges relative to polyphenols determination are addressed.•RP-LC × RP-LC vs. HILIC × RP-LC platforms are ...compared.•Past and recent applications for LC × LC are reported.
Comprehensive two-dimensional liquid chromatography is a well-established method for the unraveling of very complex real-world samples. With regard to food and natural products such a technique turned out to be a very promising approach due to its high resolving power and improved identification capability, especially in combination with mass spectrometry. In this context, polyphenols comprise a particular complex class of bioactive compounds, due to their nature and content in commonly consumed foodstuffs, making their analysis challenging. The present contribution shows an overview of the two commonly employed approaches used for polyphenol analysis, viz. RP-LC × RP-LC and HILIC × RP-LC. Furthermore, the latest implementations as well as limitations and future perspectives are critically reported.
Two pectin-polysaccharides, ASP-B2 (Mw, 5.32 kDa) and B3 (Mw, 30.51 kDa), were obtained from Acanthopanax senticosus leaves. An HILIC-UPLC-ESI−-HCD-MS/MS technique was introduced for structural ...elucidations of skeletal structure of pectin-polysaccharides in addition to empolying SEC-MALLS, GC–MS, UPLC-ESI+-MS, methylation analysis, FT-IR and NMR methods. Skeletal structure of ASP-B2 was characterized by combining HG smooth region and RG hairy region with a 2:1 chain length ratio, wheres ASP-B3 had a distinguishing 3:1 chain length ratio (HG:RG). HG were defined as repeated →4GalA1→, and RG were featured by repeated →4GalA1 → 2Rhap1 → with branched points at C-3, C-4 and C-3,4 of rhamnosyl. Side chains mainly consisted of galactans, arabinans and arabinogalactans. Both showed a random coil conformation and no triple-helix conformation. ASP-B2 and B3 possess stronger DPPH and hydroxyl radical scavenging activities, with a relatively low reducing power. Thus, two polysaccharides can be further explored as novel natural antioxidant.
•Two complex pectins, ASP-B2 and B3, were comparatively studied in structures.•ASP-B2 and B3 showed a random coil conformation and no triple-helix conformation.•ASP-B2 and B3 exerted strong DPPH and OH scavenging effects, low reducing power.
