Peak Resolution Research Articles

High throughput and cost-effective purification of small molecules using superficially porous stationary phase columns: An environment friendly approach

Over the past two decades, higher efficiencies have become the goal of most research organizations, and more so for high throughput synthesis. Consequently, there is an urgent need for an advanced purification approach to accommodate the large number of small molecules in a shorter turnaround time. To address this, an efficient and high throughput purification method was developed leveraging the superficially porous particle (SPP) column technology, which was optimized with respect to detector acquisition rate, flow rate, and gradient time. The experiments were performed using a sample mixture of six commercially available small molecules including Nicotinamide (NIC), 4-Amino benzophenone (ABP), Praziquantel (PRA), butyl-4-hydroxybenzoate (BHB), Warfarin sodium (WAR), and Ibuprofen (IBU). The study outcomes were promising and exhibited a good separation of all six analytes with ∼83% less turnaround time and ∼65% less solvent consumption than the regular methods. Furthermore, it showed significant improvement in peak symmetry, capacity factor, and resolution for all six analytes and afforded superior chromatographic performance than the regular method with fully porous particle (FPP) columns. The study concludes that SPP columns provide cost-effective, eco-friendly, and high throughput purification of small molecules.

Comparative analysis of SWIFT and Grid-SWIFT waveforms for improved ion excitation and isolation resolution

Miniaturized mass spectrometers, with their increasing portability and performance enhancements, play a crucial role in selectively analyzing target compounds within complex mixtures, particularly in situ environments. As part of our ongoing efforts to enhance the analytical capabilities of the custom-built "brick" mass spectrometer, this study compared and optimized methods for ion isolation using Stored Waveform Inverse Fourier Transform (SWIFT) and Grid-SWIFT waveforms. The efficacy of the SWIFT and Grid-SWIFT waveforms in selective ion isolation was investigated, and through optimization of both waveforms, improved ion isolation efficiency, and peak resolution were demonstrated by adjusting waveform duration and amplitude. The results conclusively showed that Grid-SWIFT outperforms traditional SWIFT waveforms in enhancing the isolation of target ions. Additionally, a novel simulation-based approach was developed to simulate the behavior of ion trajectories under the SWIFT and Grid-SWIFT waveforms within LIT to provide a theoretical basis for better performance of the Grid-SWIFT waveforms. Results show that longer waveform durations help reduce energy leakage, providing higher isolation efficiency. Grid-SWIFT waveforms exhibited significantly less energy leakage compared to SWIFT waveforms. Consequently, using Grid-SWIFT as isolation waveforms improved the mass spectra' intensity and resolution.

Tailoring glassy carbon electrode surface with FeBiO3 for electrochemical detection of dinitrophenol isomers

ABSTRACT This study presents a simultaneous, rapid, and highly sensitive electrochemical detection of 2,4- and 2,5-dinitrophenol isomers using an iron bismuth oxide (FeBiO3)-modified glassy carbon electrode (GCE). The FeBiO3-modified GCE exhibited superior electrochemical performance over bare GCE, Bi2O3-modified GCE, and Fe2O3-modified GCE, as demonstrated by electrochemical impedance spectroscopy (EIS) with the potassium ferricyanide (Fe(CN6)3-/4-) standard. The synthesised electrocatalyst FeBiO3 exhibited strong absorption both in the visible and UV regions of the spectrum, attributed to band gap transitions and charge transfer transitions linked to its Bi2O3 and Fe2O3 components. The morphology and the structure of the FeBiO3 were characterised using scanning electron microscopy (SEM), transmission electron microscopy (TEM), and X-ray diffraction (XRD), while Fourier transform infrared spectroscopy (FTIR) was employed to analyse the functional groups attached to the surface of FeBiO3. Cyclic voltammetry (CV) clearly demonstrated an increased current and improved peak resolution for the reduction of 2,4- and 2,5-dinitrophenols at the FeBiO3/GCE, compared to the bare GCE. The mechanism shows that the reduction of 2,4- and 2,5-dinitrophenols occurs through interactions with the surface oxygen functional groups of FeBiO3, resulting in the stepwise reduction of their NO2 groups to hydroxyamino compounds and then to nitrosophenols. Using square wave voltammetry (SWV), the reduction peak currents were significantly improved, enhancing detection sensitivity beyond that reported in previous works. A linear range from 1 µM to 1 mm for the dinitrophenol isomers was established, with detection limits of 0.07 µM for 2,4-dinitrophenol and 0.09 µM for 2,5-dinitrophenol. The FeBiO3/GCE electrode showed interferences with mono-nitrophenols and mono-/di-chlorophenols in solution but maintained excellent long-term stability and reproducibility, allowing it to effectively detect dinitrophenol isomers in drinking water, tap water, and wastewater samples.

Overcoming the Strong Sample Solvent Effect for Sustainability Measurement Challenges in Liquid Chromatography. Example for Bisphenol-A.

This paper describes an approach to achieve low parts per billion (ppb) concentration level detection using a reversed-phase ultrahigh-performance liquid chromatographic ultraviolet absorbance detection method with large-volume feed injection (FI) for analytes in dichloromethane (DCM). FI is a novel technology that allows sample injection at a defined speed into the LC mobile phase. We demonstrate this approach for a mixture of bisphenol A and its diglycidyl ether derivatives in DCM. DCM is a very strong injection solvent at reversed-phase LC conditions and is typically immiscible with a starting reversed-phase gradient mobile phase containing a high percentage of water. As a result, reduced separation performance and even peak splitting are seen with classic flow-through injection. The method setup comprised an octyl-bonded core-shell stationary phase (150 × 4.6 mm i.d., 2.7 μm particle size) with a water/acetonitrile mobile phase gradient and an exceptionally high injection volume of 45 μL of DCM solution using FI. Optimization of feed speed (1%) and isocratic hold duration (4 min) was crucial for final separation conditions. FI delivered more than a 20-fold improvement in the limit of detection (LOD) compared to standard flow-through injection while maintaining peak resolution. The LOD of the final method ranged between 1 and 10 ppb in the polymer resin. This methodology has high potential for trace analysis in a large variety of applications that suffer from the "strong solvent effect".

Evaluating the reliability of solid phase extraction techniques for hydrocarbon analysis by GC–MS

Saturate and aromatic compounds are essential in the petroleum industry for assessing the thermal maturity of source rocks and oils, which is critical for basin modeling and sweet-spot mapping. These compounds also play a role in environmental applications, such as oil spill fingerprinting and biogeochemistry. However, the analysis of these compounds by gas chromatography-mass spectrometry (GC–MS) requires meticulous and time-consuming separation processes. Traditional methods like normal-phase liquid column chromatography (LCC) involve large volumes of harmful solvents. This study evaluates the effectiveness of five different sorbents using solid-phase extraction (SPE) techniques—neutral Si, SiOH, Ag-ion, neutral Al, and Ag-ion mixed with activated silica—compared to LCC. The goal was to discern differences in peak resolution, concentration, and isomer ratios of saturate and aromatic compounds for thermal maturity and source rock assessments. The results show that SiOH, neutral Si, and neutral Al do not fully separate aromatic compounds from the saturate fraction, sometimes leaving 40–100% of aromatics within the saturate fraction. Ag-ion mixed with activated silica provided the best separation, resulting in up to 23 times higher aromatic concentration than SiOH. This method is more reliable for quantifying both saturate and aromatic compounds, increases the efficiency of hydrocarbon evaluations, and reduces solvent consumption by 63%, offering a more sustainable approach to hydrocarbon analysis.

Simultaneous Estimation of Atazanavir & Ritonavir in API & Marketed Formulations by Using RP-HPLC

Development and validation of RP-HPLC method for simultaneous estimation of Atazanavir and Ritonavir in their combined tablet dosage form. Atazanavir and Ritonavir are antiviral agents used in treatment of HIV. A simple, precise, rapid, accurate and cost-effective high- performance liquid chromatography (HPLC) method was successfully developed and validated for simultaneous estimation of Atazanavir and Ritonavir in their combined tablet dosage form. The selected mobile phase was Methanol: Phosphate Buffer in proportion 65:35 v/v respectively. The optimized columns used are C18 column, Symmetry and Zodiac column. X bridge C18 (4.6×150 mm, 5 µm) particle size was found to be ideal as it gave good peak shape and resolution at 1ml/min flow for Atazanavir and Ritonavir. In this study, the validation of Atazanavir and Ritonavir in API and marketed formulations were performed keeping in accordance with the parameters like system suitability, specificity, linearity, accuracy, precision (reproducibility & repeatability), robustness. The developed stability indicating method is capable for determination of impurities of Atazanavir and Ritonavir in combined tablet dosage form as well as individual dosage forms also. The method has been successfully validated according to ICH guidelines and the results obtained by using RP – HPLC are rapid, accurate and precise. The proposed methods were successfully applied for the analysis of synthetic mixtures and pharmaceutical formulations of Atazanavir and Ritonavir.

Particle size by design: Standardizing measurements for complex topical drug product assessment

The physicochemical and biopharmaceutical properties of drug substances and dosage forms can be significantly influenced by particle size. However, the diversity of equivalent particle diameters generated by different methods poses a fundamental challenge in particle size analysis. This study aimed to develop an Analytical Quality by Design (AQbD) approach to accurately assess the particle size of a complex formulation – clobetasol propionate (CP) 0.5 mg/g cream – through automated microscopy (AM) and laser light diffraction (LD). Additionally, Raman spectroscopy was utilized to determine the chemical composition of the formulation particles. In the AQbD approach, prior knowledge was considered for the construction of the Ishikawa diagram and estimate failure mode and effects analysis (FMEA). The methods were developed following the ICH Q8-Q10, and ICH Q14 guidelines, and validated according to ICH Q2, ISO 13320:2020, and EP2.9.31./USP<429>. Results indicate that a trade-off between the techniques must be established for a particle size by design: while LD offers higher throughput and more precise values at the expense of peak resolution and broadening, AM has higher variability but more reliable information in terms of size and shape analysis. The validated methods successfully demonstrated the implementation of an AQbD method in the definition of particle size methods.

Comprehensive characterization of volatile compounds in Iranian black teas using chemometric analysis of GC-MS fingerprints

Black tea, a widely popular non-alcoholic beverage, is renowned for its unique aroma and has attracted significant attention due to its complex composition. However, the chemical profile of Iranian tea remains largely unexplored. In this research, black tea samples from key tea cultivation regions in four geographical areas in northern Iran were firstly analyzed using headspace solid-phase microextraction followed by gas chromatography–mass spectrometry (HS-SPME-GC–MS) to separate, identify, and quantify their volatile organic compounds. Subsequently, employing a robust investigative strategy, we utilized for the first time the well-known multivariate curve resolution-alternating least square (MCR-ALS) method as a deconvolution technique to analyze the complex GC–MS peak clusters of tea samples. This approach effectively addressed challenges such as severe baseline drifts, overlapping peaks, and background noise, enabling the identification of minor components responsible for the distinct flavors and tastes across various samples. The MCR-ALS technique significantly improved the resolution of spectral and elution profiles, enabling both qualitative and semi-quantitative analysis of tea constituents. Qualitative analysis involved comparing resolved peak profiles to theoretical spectra, along with retention indices, while semi-quantification was conducted using the overall volume integration (OVI) approach for volatile compounds, providing a more accurate correlation between peak areas and concentrations. The application of chemometric tools in GC–MS analysis increased the number of recognized components in four tea samples, expanding from 54 to 256 components, all with concentrations exceeding 0.1 %. Among them, 32 volatile compounds were present in every tea sample. Hydrocarbons (including alkenes, alkanes, cycloalkanes, monoterpenes and sesquiterpenes), esters and alcohols were the three major chemical classes, comprising 78 % of the total relative content of volatile compounds. Analyzing black teas from four distinct regions revealed variations not only in their volatile components but also in their relative proportions. This integrated approach provides a comprehensive understanding of the volatile chemical profiles in Iranian black teas, enhances knowledge about their unique characteristics across diverse geographical origin, and lays the groundwork for quality improvement.

QbD-Driven Approach to Cleaning Method Development and Validation for Darunavir Analysis in Oral Films.

The current investigations involve developing a high-performance liquid chromatographic method that’s simple to operate, fast to establish, accurate, precise, and economical through design-enabled methods. A positive experimental design is offered to utilize the central composite design of pH and the mobile phase, two crucial components of the RP-HPLC technology. The chromatographic conditions were optimized with the release of Design Expert software version 13.0. Symmetry C18 column (4.6×150mm, 5µm) was used in the procedure, along with acetonitrile (20:80 v/v) and potassium dihydrogen orthophosphate buffer pH 4.0 as the mobile phase. There was a 0.70 ml/min flow rate. 263 nm is the wavelength of detection. Darunavir’s sharp, resolved peak was seen 2.3 minutes after swabbed samples from the 10*10 cm SS plate were injected. A linear calibration curve with an R2 of 0.9991 was observed in the concentration range of 0.25 to 25µg/ml. For precision %RSD is 1.03,1.48. Accuracy % concentration 50%,100%,150% mean recovery 99.43-100.53.LOD &amp; LOQ 1.12µg/ml, 3.39µg/ml. Robustness was tested with modest modifications to the wavelength and flow rate. Forced degradation studies were carried out and found to be within the limits. the degradation parameters such as Acid, Base, Oxidative, Photolytic, and thermal degradation parameters are according to the ICH guidelines. % Assay was done for oral films and it was found to be within the limits. Thus, the outcomes unequivocally demonstrated that the QbD technique could be effectively used to enhance the HPLC method for the measurement of darunavir in production settings.

An Untargeted Lipidomics Workflow Incorporating High-Resolution Demultiplexing (HRdm) Drift Tube Ion Mobility-Mass Spectrometry.

Global discovery lipidomics can provide comprehensive chemical information toward understanding the intricacies of metabolic lipid disorders such as dyslipidemia; however, the isomeric complexity of lipid species remains an analytical challenge. Orthogonal separation strategies, such as ion mobility (IM), can be inserted into liquid chromatography-mass spectrometry (LC-MS) untargeted lipidomic workflows for additional isomer separation and high-confidence annotation, and the emergence of high-resolution ion mobility (HRIM) strategies provides marked improvements to the resolving power (Rp > 200) that can differentiate small structural differences characteristic of isomers. One such HRIM strategy, high-resolution demultiplexing (HRdm), utilizes multiplexed drift tube ion mobility spectrometry (DTIMS) with post-acquisition algorithmic deconvolution to access high IM resolutions while retaining the measurement precision inherent to the drift tube technique; however, HRdm has yet to be utilized in untargeted studies. In this manuscript, a proof-of-concept study using ATP10D dysfunctional murine models was investigated to demonstrate the utility of HRdm-incorporated untargeted lipidomic analysis pipelines. Total lipid features were found to increase by 2.5-fold with HRdm compared to demultiplexed DTIMS as a consequence of more isomeric lipids being resolved. An example lipid, PC 36:5, was found to be significantly higher in dysfunctional ATP10D mice with two resolved peaks observed by HRdm that were absent in both the functional ATP10D mice and the standard demultiplexed DTIMS acquisition mode. The benefits of utilizing HRdm for discerning isomeric lipids in untargeted workflows have the potential to enhance our analytical understanding of lipids related to disease complexity and biologically relevant studies.

Radioxenon beta-gamma coincidence efficiency for a SAUNA detector by Geant4 simulation

A simulation for SAUNA II detector was carried out using the GEANT4 platform. The results demonstrated the accuracy of the simulation, which was well consistent with the experimental results. The simulation was used to calculate the coincidence detection efficiency of each radioxenon isotope, as well as to calculate the resolution for conversion electron peaks. Furthermore, using a 137Cs point source, the simulation was also utilized to perform the beta detector calibration procedure in two scenarios: first, the 137Cs was placed close to the middle of the plastic scintillator, and second, the 137Cs was placed outside the NaI(Tl) detector. The results showed that the spectrum in the first case is better; the source is closer to the plastic detector, leading to maximum Compton scattering. In the second case, a line in the spectrum disappears, so a high-intensity source is needed. Several methods exist to determine the coincidence detection efficiency of the detector. The recommended approach is to determine the total detection efficiency for different regions of interest. The simulation was also used to investigate the effect of replacing helium with nitrogen as a xenon-carrier gas, with the nitrogen resulting in a slight improvement in coincidence detection efficiencies and conversion electron resolution.

Nanofiltration membranes in asymmetric flow field-flow fractionation for improved organic matter size fractionation

Size fractionation of organic matter (OM) by asymmetric flow field-flow fractionation (FFFF) with ultrafiltration (UF) is limited by solute loss and peak resolution, particularly for low molecular weight fractions (<10 kDa). Nanofiltration (NF) with a molecular weight cut-off (MWCO) of ≤1 kDa can achieve significant OM retention and may improve OM fractionation. NF membranes with MWCOs ranging from 0.3 to 3 kDa were used to evaluate the retention and fractionation of nine OMs in the humic, polyphenol, biopolymer, and low molecular weight organic classes in stirred cell NF and FFFF. Membranes with an MWCO of 0.2–0.3 kDa (with a pure water permeability of 5–16 L.m−2.h−1.bar−1) with low OM interaction exhibited high peak recovery and resolution and hence improved OM fractionation. Loose NF (MWCO 3 kDa) exhibited a poor peak recovery because of a high OM loss due to both adhesion and permeation, particularly at high ionic strengths (>10 mM). A mixture of nine types of OM and natural organic matter samples from rivers, swamps and lakes achieved good fractionation with a 0.3 kDa (NF270) membrane at low ionic strength. The performance of NF for the analysis of OM and colloids in the smallest size range (≤1 kDa MW) could be further improved by optimizing the salt composition of the mobile phase and by overcoming the pressure limitations of FFFF channels.

Development and Application of HPLC and UFLC Methods for the Analysis of Cardiovascular Drugs

Separation and identification of amiloride, metoprolol, hydrochlorothiazide, Carvedilol and amlodipine, in human plasma using the presented SPE and HPLC procedures was selective, efficient, robust, economical, environmentally friendly, and repeatable. Because plasma samples showed no evidence of a secondary peak, it was determined that the claimed drug-drug interaction between cardiovascular medications did not occur. Furthermore, no new peak was generated, indicating no metabolic product. This is the first report on isolating and identifying these nine cardiovascular medicines. Thus, SPE and HPLC techniques may be used to investigate these 5 cardiovascular medications. Successful monitoring of these medications in human plasma was achieved using the established SPE and HPLC technologies. The results of this inquiry were contrasted with those that had already been released. In comparison to other works in the literature, it was determined that the current study had the best complete baseline separation and lowest detection limit. The retention, separation, and resolution factors were discovered to have ranges of 0.07-9.14, 1.44-4.21, and 2.15-18.66, respectively. The SPE and UFLC techniques created and verified for this purpose were used to examine human plasma samples for the presence of cardiovascular drugs. Peak retention, separation, resolution, and symmetry all matched the reference samples' values. The results for symmetry, separation, and resolution were all in agreement with the reference samples. The methods used to separate and identify amiloride, metoprolol, hydrochlorothiazide; Carvedilol and Amlodipine in human plasma were found to be selective, effective, hardy, affordable, environmentally friendly, and reproducible, according to the authors. The selectivity of the SPE technique was validated by the absence of any secondary signal in the plasma samples. Additionally, there was no evidence of these drugs degrading in the plasma sample. This is the initial report on the simultaneous isolation and identification of these eight medications. The well-known SPE and UFLC techniques were successfully used to monitor these drugs in human plasma. Therefore, these drugs can be detected in any plasma sample using SPE or UFLC methods

Development and Application of HPLC and UFLC Methods for the Analysis of Anti Histaminic Drugs

Research focused on medications used to treat allergic reactions. The results of this inquiry were contrasted with those that had already been released. In comparison to other works in the literature, it was determined that the current study had the best complete baseline separation and lowest detection limit. The retention, separation, and resolution factors were discovered to have ranges of 0.07-9.14, 1.44-4.21, and 2.15-18.66, respectively. The SPE and UFLC techniques created and verified for this purpose were used to examine human plasma samples for the presence of antihistaminic drugs. Peak retention, separation, resolution, and symmetry all matched the reference samples' values. The results for symmetry, separation, and resolution were all in agreement with the reference samples. The methods used to separate and identify Phenylephrine, cetirizine, loratadine, montelukast, and Ebastine in human plasma were found to be selective, effective, hardy, affordable, environmentally friendly, and reproducible, according to the authors. The selectivity of the SPE technique was validated by the absence of any secondary signal in the plasma samples. Additionally, there was no evidence of these drugs degrading in the plasma sample. This is the initial report on the simultaneous isolation and identification of these eight medications. The well-known SPE and UFLC techniques were successfully used to monitor these drugs in human plasma. Therefore, these drugs can be detected in any plasma sample using SPE or UFLC methods.

Stability-indicating liquid chromatographic method development and validation for quantification of trifarotene in pure and topical drug product

BackgroundTrifarotene is effective for treating acne and other skin issues. To ensure its quality and meet regulatory standards, a reverse phase liquid chromatography (RP-LC) stability-indicating method was developed and validated for its quantification in pure and topical dosage forms. An isocratic elution chromatographic method was employed, using an octadecylsilyl silica gel-packed column (150 mm × 4.6 mm, 3 µm particle size). The mobile phase was a mixture of phosphate buffer and acetonitrile (40:60 v/v). Chromatographic conditions included a flow rate of 0.5 mL/min, column temperature of 40 °C, detection at 265 nm, and injection volume of 20 µL.ResultsThe developed analytical method reports the retention time of trifarotene 11.2 min, higher theoretical plate count, asymmetric peak, and good resolution between the peaks of trifarotene, phenoxyethanol, and environmentally generated impurities.ConclusionThe analytical method has been found to be linear, accurate, robust, specific, and selective for impurities produced during forced degradation studies. The proposed analytical method can be utilized for routine pharmaceutical analysis of trifarotene to judge its quality and safety.

Quantitative Analysis of Chondroitin Sulfate and Dermatan Sulfate Using Capillary Electrophoresis With Gaussian Distribution‐Based Peak Fitting for Improved Peak Resolution

ABSTRACTIn this work, capillary electrophoresis with Gaussian distribution‐based peak fitting for improved peak resolution for simultaneous quantitative analysis of chondroitin sulfate (CS) and dermatan sulfate (DS) was developed for the first time. The effects of different pH, voltage, and peak shapes on the peak fitting were investigated. The results indicate that separation conditions that affect peak shape have a significant impact on peak fitting. Under optimized separation conditions, the peak fitting achieves high accuracy and precision, the relative standard deviation (n = 5) of peak area and migration time of CS and DS were 1.1%, 1.8% and 0.94%, 0.84%, respectively. Furthermore, the results obtained by peak fitting were close to real values. The developed method was used to analyze the purity of CS. The results had a high consistency with the labeled value; furthermore, the peak fitting could point out the number of possible impurities. The developed method has good potential for the analysis of other glycosaminoglycans with peak overlap.

Racetrack FAIMS for High-Resolution and High-Sensitivity Characterization of Peptide Conformers.

A racetrack field asymmetric waveform ion mobility spectrometry (r-FAIMS) device, which consists of both cylindrical FAIMS (c-FAIMS) and planar FAIMS (p-FAIMS) sections with a 1 mm gap width, was developed and applied for high-resolution and high-sensitivity exploration of conformational diversity for peptides. The optimal operating conditions of r-FAIMS were systemically studied, and the performance of the fully optimized r-FAIMS was compared to a previously developed p-FAIMS in detail by using pure nitrogen as the FAIMS carrier gas. Relying on the ion focusing effect in the c-FAIMS section, the intensity of the FAIMS spectrum for doubly charged bradykinin ions acquired by using r-FAIMS is ∼8.5-fold higher than that acquired by using p-FAIMS under the same resolving power/resolution condition, implying about an order of magnitude better sensitivity of r-FAIMS. In addition, the peak separation resolution of r-FAIMS was ∼1.70-fold higher than p-FAIMS under a similar sensitivity condition for doubly charged bradykinin ions. Due to a reduced gap width of the newly designed r-FAIMS (1 mm) as compared to the previously developed p-FAIMS (1.88 mm), r-FAIMS can operate at a much higher separation field with a similar FAIMS dispersion voltage (DV) to gain significantly higher resolving power. For triply charged syntide 2 ions, the resolving power of r-FAIMS can easily exceed 120 at -3.5 kV DV by using pure nitrogen as the FAIMS carrier gas as compared to 44.2 resolving power obtained by using p-FAIMS at -4.0 kV DV. All of the experimental results have confirmed that r-FAIMS can perform structural characterization of biomolecules with both high resolution and high sensitivity.

Development of a high-performance liquid chromatography method for simultaneous quantification of sixteen polyphenols and application to tomato

The main purpose of the current work was to develop a new method to evaluate and quantify sixteen polyphenol compounds from tomato fruit using high-performance liquid chromatography (HPLC). The separation of 16 polyphenols from tomato fruit was achieved in < 60 min by using a Waters Symmetry C18 column (250 × 4.6 mm i. d, 5 µm particle sizes) with a gradient system of ultrapure water (1 % acetic acid) and 100 % methanol, a temperature of 30 °C, an injection volume of 10 μL and a flow rate of 1.1 mL/min, respectively. The analytical characteristics of evaluation method provide sufficient sensitivity for all tomato polyphenols compounds within normal range 0.1–20 μg·mL−1 (R2≥0.999) with 0.069–0.365 μg·mL−1 LOD, and 0.171–1.106 μg·mL−1 LOQ, with good system suitability (<2 % RSD of retention time, peak area, and tailing factor, 6,000–1,336,000 N, and >1.5 peak resolution), <10 % RSD of precision, stability, repeatability, and robustness, and 99.2 - 105.0 % of recovery. The applicability of this method was demonstrated by the determination of polyphenols in nine cultivars of tomatoes. The results showed that ‘184′ possessed the highest content of total polyphenols (1249.53 μg·g−1 DW) followed by ‘Disease resistance 184′ (1064.93 μg·g−1 DW). The main polyphenol components were rutin, quercetin, gallic acid, chlorogenic acid, 2,5-dihydroxy benzoic acid, caffeic acid and benzoic acid in tomato fruits. In conclusion, this novel HPLC method is useful and acceptable to analyze polyphenols in tomato fruit.

Impact of the degree of substitution on the enantioseparation of anionic β-cyclodextrin in liquid‒liquid extraction and countercurrent chromatography: Insights from molecular docking simulations

Five types of sulfobutylether-β-cyclodextrin (SBE-β-CD) and carboxymethyl-β-cyclodextrin (CM-β-CD) with different degrees of substitution were synthesized, and six and five racemates were respectively chosen to study the influence of the degree of substitution on the enantioseparation factor. The synthesized SBE-β-CD and CM-β-CD were characterized using 1H NMR spectroscopy and mass spectrometry. The results indicated that the influence of the degree of substitution on enantioseparation for distinct racemates exhibited significant variability. Increasing the degree of substitution of CM-β-CD led to an increasing enantioseparation factor, while SBE-β-CD with a specific degree of substitution provided the optimum enantioseparation factor for some racemates. The optimum enantioseparation factors of N-methyl duloxetine and duloxetine were obtained when a relatively low degree of substitution (DS = 3.5) was selected. And the optimum enantioseparation factor was obtained with a relatively high degree of substitution (DS = 7.5). SBE-β-CD with a low substitution degree of 3.5 was chosen to optimize the countercurrent chromatographic enantioseparation of N-methyl-duloxetine, which resulted in a significant improvement in peak resolution from 0.51 to 0.83. Molecular docking was used to construct three SBE-β-CDs with different degrees and distributions of substitution, and a good agreement was found between the docking results and the experimental results.

Resolution Enhancement of Metabolomic J-Res NMR Spectra Using Deep Learning.

J-Resolved (J-Res) nuclear magnetic resonance (NMR) spectroscopy is pivotal in NMR-based metabolomics, but practitioners face a choice between time-consuming high-resolution (HR) experiments or shorter low-resolution (LR) experiments which exhibit significant peak overlap. Deep learning neural networks have been successfully used in many fields to enhance quality of natural images, especially with regard to resolution, and therefore offer the prospect of improving two-dimensional (2D) NMR data. Here, we introduce the J-RESRGAN, an adapted and modified generative adversarial network (GAN) for image super-resolution (SR), which we trained specifically for metabolomic J-Res spectra to enhance peak resolution. A novel symmetric loss function was introduced, exploiting the inherent vertical symmetry of J-Res NMR spectra. Model training used simulated high-resolution J-Res spectra of complex mixtures, with corresponding low-resolution spectra generated via blurring and down-sampling. Evaluation of peak pair resolvability on J-RESRGAN demonstrated remarkable improvement in resolution across a variety of samples. In simulated plasma data, 100% of peak pairs exhibited enhanced resolution in super-resolution spectra compared to their low-resolution counterparts. Similarly, enhanced resolution was observed in 80.8-100% of peak pairs in experimental plasma, 85.0-96.7% in urine, 94.4-98.9% in full fat milk, and 82.6-91.7% in orange juice. J-RESRGAN is not sample type, spectrometer or field strength dependent and improvements on previously acquired data can be seen in seconds on a standard desktop computer. We believe this demonstrates the promise of deep learning methods to enhance NMR metabolomic data, and in particular, the power of J-RESRGAN to elucidate overlapping peaks, advancing precision in a wide variety of NMR-based metabolomics studies. The model, J-RESRGAN, is openly accessible for download on GitHub at https://github.com/yanyan5420/J-RESRGAN.