Quantitative Chromatographic Analysis Research Articles

Chemodiversity of Arctic Plant Dryas oxyodonta: LC-MS Profile and Antioxidant Activity

Dryas oxyodonta Yuz. is a perennial evergreen shrub from the Rosaceae family. D. oxyodonta thrives in subalpine and subarctic regions, as well as in highlands spanning from Central Asia to Siberia and Mongolia. Owing to a lack of information on its chemical composition, we conducted qualitative and quantitative chromatographic analyses on extracts from the leaves and flowers of D. oxyodonta sourced from various Siberian habitats. Employing high-performance liquid chromatography with photodiode-array detection and electrospray ionization triple-quadrupole mass spectrometric detection, we identified 40 compounds, encompassing gallotannins, hydroxycinnamates, procyanidins, catechins, flavonoids, and triterpenes. All Siberian populations of D. oxyodonta exhibited a notable abundance of phenolic compounds. Furthermore, we identified rare glycosides, such as sexangularetin and corniculatusin, as potential markers of the chemodiversity within the Dryas genus. Extracts from the flowers and leaves were effective scavengers of free radicals, including DPPH•, ABTS•+−, O2•−, and •OH radicals. Our findings unequivocally establish D. oxyodonta as a rich source of phenolic compounds with potent antioxidant activity, suggesting its potential utility in developing novel functional products.

(Invited) Deeply Rechargeable Zinc Anodes for High-Energy Rechargeable Aqueous Batteries

Zinc-based aqueous batteries are promising alternative to many mainstream battery technologies today, due to the superior safety and broad availability of raw materials. However, existing zinc anodes suffer from either poor cycle life or low utilization in both neutral and alkaline aqueous electrolytes. Passivation, dissolution, and hydrogen evolution are three main reasons for irreversibility of zinc anodes in alkaline electrolytes, which limits the rechargeability and usable energy density. In this talk, I will present our recent works on using nanoscale material design to overcome passivation, dissolution, and hydrogen evolution issues of zinc anode, towards a deeply rechargeable zinc-based battery. I will also introduce the battery-gas chromatography quantitative analysis, as well as in situ microscopy methodologies we have developed, to quantify gas evolution side reaction, as well as visualize the reaction on electrodes during operation.

A novel process for developing a quantitative chromatographic fingerprint analysis method based on analytical quality by design: A case study of Xiaochaihu capsules

In this study, an improved Analytical Quality by Design (AQbD) process was proposed to establish quantitative fingerprints of Chinese medicines. A quantitative fingerprint method for the strong polar components in Xiaochaihu capsules was developed as an example. Different method evaluation indicators were used to characterize this method, and these indicators were measured at different stages of quantitative fingerprint development. Through this method, the difficulty in determining common peaks in the early stage of quantitative fingerprint development is eliminated. The method parameters that needed to be optimized were the mobile phase gradient conditions, column temperature and flow rate through the Ishikawa diagram. Single-factor experiments were used to determine some parameters. Definitive screening design (DSD) experiments were used to optimize critical method parameters (CMPs). The common peaks were determined, and indices were optimized. The critical method attributes (CMAs) were the resolution of guanosine, the percentage of common peak to the total peak area, retention time of the last peak and the common peak number. A stepwise regression method was used to build quantitative models for critical method attributes and critical method parameters. The method operable design region (MODR) was calculated by the experimental error simulation method and successfully verified. After the analysis conditions were optimized, four components, uridine, adenine, 5-hydroxymethylfurfural, and guanosine, were quantitatively determined. There were 7 common peaks in the fingerprints. The common peak area accounted for 90.64%. Both fingerprint and quantitative detection analysis methods were methodologically validated. The new AQbD process proposed in this study can be potentially applied to develop other Chinese medicines fingerprint methods.

Comparative characterization of different variants of quantitative chromatographic analysis using external and internal standards

Chromatographic analysis of a series of two-component samples (solutions of a target analyte and a standard) allows simultaneous comparison of the possibilities and features of six variants of quantitative analysis using the external and internal standards techniques according to the criteria of results precision and repeatability (random and systematic errors values). These variants include the simplest version of the external standard method (I); its modified version (II) that implies the application of an additional standard and averaging not the absolute, but relative peak areas; the commonly used version of the internal standard method (III); and its version modified in a similar manner (IV). Besides, two variants of using a homologue of the target analyte as the internal standard without determining the calibration coefficients are considered (V) and (VI). This topic is of interest for optimization of practical works on chromatography and teaching the subject in general. The requirements to the additional and internal standards are not identical. Any compound (both present in the samples or added to them) can be selected as the additional standards, and precise setting of their concentrations is not required. It is only necessary to ensure their equal concentrations in the analyzed and reference solutions. It is shown that the modified versions of the external and internal standard methods are characterized by the equal relative standard deviations of the results. The minimal relative standard deviations of the results are typical for the variants implying the use of an additional (II) or an internal (IV) standard and the averaging the ratio of peak areas of target analytes and such standards. The systematic errors of determinations appeared to be minimal for the same variants. Controlling the values of the calibration coefficients is informative for revealing the possible distortions of the composition of samples due to the partial evaporation of volatile constituents in the course of handling such samples.

Development of a Quantitative Chromatographic Fingerprint Analysis Method for Sugar Components of Xiaochaihu Capsules Based on Quality by Design Concept

Background: Xiaochaihu capsule is composed of seven traditional Chinese medicines. The pharmacopoeia only focuses on the quantitative detection of baicalin, which cannot fully reflect the quality of the preparation. Some medium polar components were used to establish the fingerprint of Xiaochaihu capsule, but there was no report on the strong polar components. Methods: A high performance liquid chromatography-corona charged aerosol detection technology was used to establish a fingerprint analysis method for Xiaochaihu capsules following an analytical quality by design approach. Definitive screening designed experiments were used to optimize the method parameters. A stepwise regression method was used to build quantitative models. The method operable design region was calculated using the experimental error simulation method. Plackett–Burman designed experiments were carried out to test robustness. Results: The contents of four components were simultaneously determined. There were seven common peaks in the fingerprint. The common peak area accounted for 91.72%. Both fingerprint and quantitative analysis methods were validated as applicable in the methodology study. The quantitative fingerprint analysis method for sugar components can fill the gap in the detection of strong polar components in the existing methods. It provides a new technology for the comprehensive overall evaluation of Xiaochaihu capsule.

(Invited) Deeply Rechargeable Zinc Anodes for High-Energy Rechargeable Aqueous Batteries

Zinc-based aqueous batteries are promising alternative to many mainstream battery technologies today, due to the superior safety. However, existing zinc anodes suffer from either poor cycle life or low utilization in both neutral and alkaline aqueous electrolytes. Passivation, dissolution, and hydrogen evolution are three main reasons for irreversibility of zinc anodes in alkaline electrolytes, which limits the rechargeability and usable energy density. In this talk, I will present our recent works on using nanoscale material design to overcome passivation, dissolution, and hydrogen evolution issues of zinc anode, towards a deeply rechargeable zinc-based battery. I will also introduce the battery-gas chromatography quantitative analysis, as well as in situ microscopy methodologies we have developed, to quantify gas evolution side reaction, as well as visualize the reaction on electrodes during operation.

Comprehensive analysis of pyrolysis oil obtained from waste tire in a water vapor environment

Determination of the composition of pyrolysis products of organic compounds is a very difficult task due to their multicomponent nature [1–3]. Pyrolysis oils formed from rubber products and organic polymer wastes contain various classes of organic compounds with a huge spread in molecular weight and polarity: hydrocarbons, amides, phenols, amines, organosulfur compounds, etc. [1]. At the same time, the results of qualitative and quantitative chromatographic analysis of pyrolysis oils are often ambiguous: there are problems of overlapping peaks and misinterpretation of the data obtained due to the complexity of the matrix and the multicomponent composition [2–5]. Pyrolysis products can be used as valuable hydrocarbons or additives to hydrocarbon fuels. Therefore, the analysis of the chemical composition of pyrolysis oils is important both for assessing the degree of their toxicity and for finding an effective method for their purification.

Phytochemical Analysis of Polyphenols in Leaf Extract from Vernonia amygdalina Delile Plant Growing in Uganda

Due to the presence of phytochemicals, plants have been known to be used in the treatment and management of various diseases. Vernonia amygdalina, belonging to the Asteraceae family, is a plant known for its many applications in traditional medicine for various purposes. Previous studies on the methanolic leaf extract of this plant have proved the antibacterial, cytotoxic, anticancer and antioxidant effects indicative of promising therapeutic potentials. In this work, chromatographic and spectroscopic techniques along with high-performance liquid chromatography quantitative analysis were adopted to isolate, identify and quantify polyphenolic compounds in V. amygdalina leaf extract. UHPLC-DAD-ESI-MS/MS and UHPLC-DAD methods were adopted for qualitative and quantitative analysis, respectively. In the case of polyphenol separation, some reference substances were isolated by preparative HPLC. Seven polyphenols were identified and quantified in this study: 5-O-caffeoylquinic acid, luteolin hexoside, 3,4-O-dicaffeoylquinic acid, 1,5-O-dicaffeoylquinic acid, 3,5-O-dicaffeoylquinic acid, 4,5-O-dicaffeoylquinic acid and luteolin dihexoside, with 3,5-O-dicaffeoylquinic acid being isolated in the highest quantity of 27.49 mg g−1 extract.

High-performance liquid chromatography quantitative analysis of ephedrine alkaloids in Ephedrae Herba on a perfluorooctyl stationary phase.

Ephedrae Herba is one of the most commonly used herbal medicines, and it has been shown that most of the clinical efficacy for cold and asthma is exerted by its alkaloidal components. A simple and sensitive high-performance liquid chromatography method was developed using a perfluorooctyl column for the simultaneous determination of five alkaloids (norephedrine, norpseudoephedrine, ephedrine, pseudoephedrine, and methylephedrine) in Ephedrae Herba. The mobile phase comprising acetonitrile and 15mM ammonium trifluoroacetate was used to elute the targets in isocratic elution mode. The method was validated for linearity (R2 >0.999), repeatability, intraday and interday precision, recoveries with trueness (93.87-110.99%), limits of detection (5.35-5.76µg/mL), and limits of quantification (20µg/mL). The quantitative results revealed that the developed method was precise and accurate. Then it was successfully applied to determine the difference in the contents ofthree batches of Ephedrae Herba from three pharmaceutical companies.

Phase structure characterization and compatibilization mechanism of epoxy asphalt modified by thermoplastic elastomer (SBS)

Epoxy resin is widely used in thermosetting polymer modified asphalt. However, the delamination and segregation will occur owing to the poor compatibility between epoxy resin and asphalt. The rigid molecular structure of epoxy resin also leads to insufficient toughness of epoxy asphalt (EA) at low temperature. In order to solve these problems of EA, styrene butadiene styrene block copolymer (SBS) is used to prepare modified epoxy asphalt (SBS-EA). The phase separation microstructure, rheological properties, mechanical properties and compatibilization mechanism of SBS-EAs are investigated. The initial viscosity of EA is increased by introducing SBS, but it still satisfies the requirement of pavement. The results of fluorescence microscopy and micro-computed tomography reveal that the compatibility of EA can be significantly improved by 3 wt% SBS. The average particle size of dispersed phase asphalt can be reduced with the content of SBS. Micro-infrared and gel permeation chromatography quantitative analysis turn out that SBS will migrate from the base asphalt to epoxy resin crosslinking network. The tensile strength, elongation at break and toughness of 3%SBS-EA are 12%, 67% and 78% higher than those of EA due to the good phase separation structure. Furthermore, the dispersed asphalt phase size is closely correlated with the mechanical properties of SBS-EA.

Tentative qualitative and quantitative analysis of phenolic compounds in leaf extract from Carica papaya Linn. plant growing in Uganda

Summary Introduction Carica papaya L. belongs to Caricaceae family. It is an evergreen tree or shrub with usually unbranched trunk. Previous studies on the methanolic leaf extract of this plant proved the antibacterial, cytotoxic, anticancer and antioxidant effects indicative of promising therapeutic potentials. Methods Chromatographic and spectroscopic techniques along with high performance liquid chromatography quantitative analysis were performed to isolate, tentatively identify and quantify polyphenolic compounds in MeOH extract of C. papaya leaves. UHPLC-DAD-ESI-MS/MS and UHPLC-DAD methods were adopted for qualitative and quantitative analysis respectively. In the case of polyphenol separation, some reference substances were isolated by preparative HPLC. Results Six polyphenols were isolated and quantified from C. papaya leaf extract, including three phenolic acids: derivatives of caffeic acid, p-coumaric acid, ferulic acid and three flavonoids: quercetin dirhamnosylhexoside, kaempferol dirhamnosyl-hexoside, quercetin 3-O-rutinoside. Among them quercetin dirhamnosyl-hexoside was significantly higher (1.9 mg/g extract) than other identified compounds. Conclusion The results of present study provide a new data on polyphenols composition of C. papaya plant derived from Uganda and promote research on its possible standardization and use as a dietary supplement and/or herbal medicine.

Quantitative phytochemical and chromatographic analysis of phenolic compounds in methanolic leaf extract of Costus pictus D. Don

Quantitative phytochemical and chromatographic analysis of phenolic compounds in methanolic leaf extract of Costus pictus D. Don

INSIGHT INTO BASELINE TROUBLES AND PROBLEM RESOLUTIONS IN HIGH-PERFORMANCE LIQUID CHROMATOGRAPHY

For pharmaceutical industry, HPLC is more than 40 years old. Even after instrumental advancements in HPLC problems still arise, troubleshooting still comes. HPLC is the main stay of analytical section nowadays in pharmaceutical industry whether it is API industry or formulation plant. Best approach for troubleshooting HPLC problems is going with systematic way. At the start of quantitative chromatographic analysis, the first parameter of paramount importance is baseline, especially for measuring the area of any given peak. Baseline usually suffers from these errors, namely, high baseline drift, periodic baseline fluctuation, and spikes. This article aims at suggesting some working methodologies which will resolve as well as avoid cases of such errors and failures.

Quality Evaluation of Corydalis yanhusuo by High-Performance Liquid Chromatography Fingerprinting Coupled with Multicomponent Quantitative Analysis

Corydalis Rhizoma is the tuber of Corydalis yanhusuo W. T. Wang, which has been long used in traditional Chinese medicine. Herein, the quality of C. yanhusuo samples collected from 23 regions of three provinces in China is evaluated through high-performance liquid chromatography fingerprinting coupled with similarity, hierarchical clustering, and principal component analyses. Sample similarities are evaluated according to the State Food and Drug Administration requirements by selection of 18 characteristic chromatographic fingerprint peaks and are found to vary between 0.455 and 0.999. Moreover, common patterns of a typical local variety of C. yanhusuo sourced in the Panan County are established. The obtained results show that the combination of quantitative analysis and chromatographic fingerprint analysis can be readily utilized for quality control purposes, offering a comprehensive strategy for quality evaluation of C. yanhusuo and related products.

Influence of short peptides with aromatic amino acid residues on aggregation properties of serum amyloid A and its fragments

Serum amyloid A variant 1.1 (SAA1.1) is an acute phase protein. In response to injury, inflammation or infection its production increases highly, which may lead to aggregation of the protein and accumulation of its deposits in various organs. Due to the cellular toxicity of the aggregates, as well as the fact that accumulated deposits are a burden that obstructs proper functioning of the affected tissues, it is vital to find a way to suppress the process of pathological aggregates formation. To make this possible, it is necessary to investigate thoroughly the oligomerization process and recognize factors that may influence its course. Some previous studies showed that aromatic interactions are important to the potential of an inhibitor to suppress the aggregation process. In our research we had proved that a five-residue peptide RSFFS (saa1-5) is an efficient inhibitor of aggregation of the most amyloidogenic fragment of SAA1.1, SAA1-12. In the present work the oligomerization and aggregation propensity of SAA1-12 was compared to that of SAA1-27, in order to determine the contribution of the sequence which extends beyond the most amyloidogenic region but encompasses residues reportedly involved in the stabilization of the SAA native conformation. Thioflavin T fluorescence assay, quantitative chromatographic analysis of the insoluble fraction and transmission electron microscopy allowed for a deeper insight into the SAA aggregation process and the morphology of aggregates. Substitutions of Phe3 and/or Phe4 residues in saa1-5 sequence with tryptophan, tyrosine, homophenylalanine, naphthylalanine and β,β-diphenylalanine allowed to study the influence of different aromatic systems on the aggregation of SAA1-12 and SAA1-27, and evaluate these results in relation to hSAA1.1 protein. Our results indicate that compounds with aromatic moieties can affect the course of the aggregation process and change the ratio between the soluble and insoluble aggregates.

Quantitative chromatographic fingerprint analysis of Sanye Tangzhiqing Decoction based on quality by design concept

In this work,a high performance liquid chromatography-ultraviolet( HPLC-UV) detection technology was used to establish fingerprint analysis method for Sanye Tangzhiqing Decoction following an analytical quality by design( AQb D) approach. Firstly,column temperature,flow rate,and gradient elution conditions were determined as the method parameters needing to be optimized. Then according to the results of definitive screening design,three critical method attributes( CMAs) were identified,including peak number,the percentage of common peak area to total peak area,and retention time of the last peak. A stepwise regression method was used then to build quantitative models between CMAs and method parameters. Probability-based design space was calculated and successfully verified using the experimental error simulation method. After the analysis conditions were optimized,the contents of six components,namely chlorogenic acid,paeoniflorin,rutin,hyperoside,quercetin-3-O-β-D-glucuronide,and salvianolic acid B were simultaneously determined. There were 19 common peaks in the fingerprint and their common peak area accounted for 96% of the total peak area. Both fingerprint and quantitative analysis methods were validated applicable in methodology study,and they can be applied to determine new samples.

Determination of polioksin B in the air environment and in washouts from skin of operators by HPLC

Polyoxin B being an effective inhibitor of synthesis of chitin of the cell wall of many phytopathogenic fungi, is recommended as a fungicide for control of phytopathogenic organisms that cause damage to crop. For the determination of the exposure of employees working with pesticides there was developed the method of the measurement of concentrations of polyoxin B in air of working area, atmospheric air of populated areas and washouts from the operators ’ integuments, based on high performance liquid chromatography with ultraviolet detector (detection wavelength of270 nm), including sampling air environment in the sorption tube ORBO-44, filled with sorbent XAD-2, extraction of the sorbent with polyoxin by a mixture of carbinol-water (in a ratio of 95:5,on volume), washout from the surface of the skin with ethyl alcohol by way of washing, concentrating, quantitative chromatographic analysis. Lower limits of the quantification ofpolyoxin B in the air ofworking area - 0.2 mg/m at the aspiration of 2 dm of air, atmospheric air - 0.016 mg/m at the aspiration of 25 dm of air, in washouts from the operators’ integuments - 0.4 pg/wash, the linear range of the defined concentrations accounted for of 0.2 - 2.4 pg/cm, the total error of measurement of the concentrations of polyoxin B in air is 17%; in washouts from the operators’ integuments - 16%. The developed method was approbated for the determination of polyoxin in samples of air of working zone, atmospheric air within the sanitary gap, washouts from the operators ’ integuments and air drift samples taken under processing of roses in the hothouse and in the monitoring of the phytosanitary condition of the plants every other day after treatment.

Revealing the inner workings of the power function algorithm in Charged Aerosol Detection: A simple and effective approach to optimizing power function value for quantitative analysis

In recent years, charged aerosol detection (CAD) has become a valuable tool for fast and efficient quantitative chromatographic analysis of drug substances with weak UV absorption. In analytical method development using CAD, the power function settings available in the instrument software are key for linearization of the signal response with respect to analyte concentration. However, the relatively poor understanding of the power function algorithm has limited a more widespread use of CAD for quantitative assays, especially in the late stage of method validation and GMP laboratories. Herein, we present an approach to understand the inner workings of the power function value (PFV), the PFV optimization algorithm, as well as a method to determine the optimum PFV based on the signals acquired at PFV = 1 (default CAD settings). The exponent and the constant in the PFV equation used for modeling follow a trend as a function of PFV. The CAD signal at any PFV was modeled based on the signal acquired at PFV = 1, the modelling was successful for two analytes at different concentration levels on two different CAD detectors of the same model. This method reveals the functionality of the PFV which substantially simplifies the workflow needed to optimize the detector signal. The accuracy between the experimental and theoretical results showed high correlation and always resulted in the same optimum PFV determined by both ways. The approach described in this investigation simplifies the selection of the optimum PFV at which the signal is more linear, the signal-to-noise is higher, and the area reproducibility is better. The power function algorithm elucidated herein enables determination of optimum PFV from minimal experimental output and excellent overall accuracy. This paper provides an approach that includes no data transformation outside the vendor software, a very important requirement to easily validate and report results in a GMP environment.

Validated method for trace impurities analysis in bulk gas using Gas Chromatography with Pulse Discharge Helium Ionization Detection

Pulse discharge helium ionization detector (PDHID) is developed to respond the need of many applications in the area of chromatographic quantitative analysis. The PDHID is a general type of detector with very high detectability property covering a wide range of chemical compounds. Based on ISO/IEC 17025, a reliable and accurate of measurement result can be obtained by applying a validated method. This paper reports a validation study with regard to the application of GC-PDHID for trace impurity analysis in bulk gases. An optimized operating condition of GC was used to evaluate the method performance with parameters including precision (repeatability and reproducibility), linearity, limit of detection (LoD), and limit of quantification (LoQ). In this study, six trace compounds (CO2, H2, Ar, O2, CH4, and CO) in bulk gas were analyzed. The results show that the validated of the GC-PDHID method was repeatable and reproducible for the analysis of trace impurities in bulk gas. The %RSD of repeatability and reproducibility were found to be < 3.00% and < 10.50%, respectively. Good linearity of the validated method was obtained with the correlation coefficient (R2) higher than 0.99 for all six trace compounds. In addition, the calculated LOD and LOQ of the GC-PDHID were found to be < 0.60 and < 2.00 μmol mol−1 respectively. In conclusion, validated method of the GC-PDHID for the measurement of the six trace compounds in bulk gas is reliable and fit for its purpose, implying that the validated method can be used in daily measurement in the testing laboratory.

Quality attributes and chemical composition of commercial cinnamon oils

Cinnamon oil is commercially available and has been used by the food, fragrance and pharmaceutical industries. In present work, 22 batches of commercial cinnamon oils from different brands were investigated. There are no significant differences in the appearance profiles and physicochemical properties between the samples, but great variation in the content of cinnamaldehyde was found by gas chromatography quantitative analysis. They were further chemically characterised by gas chromatography-mass spectrometry (GC-MS), and 54 volatile compounds were identified. According to principal component analysis of the GC-MS data set, the samples were clustered into three distinct groups with higher concentrations of cinnamaldehyde, diethyl phthalate, and eugenol as their markers, respectively. Moreover, coumarin, a hepatotoxic natural compound, was detected in some samples, which indicated that the samples with higher level of cinnamaldehyde and limited amount of coumarin would be better choice for efficacious and safe use.