We prepared two kinds of fine particles by treating lactose monohydrate (Lac) with the same formulation in a fluidized-bed granulator, which differed in the spraying air pressure. Raman intensities ...of treated Lac during processing were measured using a handheld-type Raman spectrometer and plotted against particle size. As particle size increased, Raman intensity decreased in both operating conditions. A linear relationship between them was observed, and regression lines were obtained with good correlation coefficients as calibration curves in both operating conditions. We also prepared two other types of fine particles by treating Lac with the same formulation in a fluidized-bed granulator, which differed in the scale or processing mechanism. The particle size could be successfully predicted using the calibration curve obtained taking powder porosity into consideration. Based on this study, we present a new at-line approach in process analytical technology to monitor and predict particle size using a handheld-type Raman spectrometer.
In this study, an at-line nanofractionation (ANF) platform was successfully fabricated in parallel with mass spectrometry and trypsin inhibitory bioactivity assessment for rapid screening of trypsin ...inhibitors (TIs) from natural products for the first time. After systematic optimization, the ANF platform was applied to screen and identify TIs in the extract of a traditional Chinese herb, i.e., Cotinus coggygria Scop. The semi-preparative reverse-phase liquid chromatography was used subsequently to further simplify and enrich the insufficiently separated components. After comprehensive evaluation and validation, the ANF platform successfully identified 12 compounds as potential TIs, including 8 flavonoids and 2 organic acids. Additionally, a comparison study was conducted using two other ligand fishing approaches, i.e., capillary monolithic and magnetic beads-based trypsin-immobilized enzyme microreactors, which successfully identified 8 identical flavonoids as TIs. Importantly, the molecular docking study showed the molecular interactions between enzymes and inhibitors, thus strongly supporting the experimental results. Overall, this work has fully demonstrated the feasibility of the established ANF platform for screening TIs from Cotinus coggygria Scop., and proved its great prospects for screening bioactive components from natural products.
Introduction
Chromatographic techniques coupled with bioassays are popularly used for the detection of bioactive compounds in natural products. In this study phytochemicals responsible for showing ...Phosphodiesterase type 5 (PDE5) inhibitory activity in Derris scandens were studied using at‐line method.
Objective
The objective of this study was to develop an at‐line liquid chromatography quadrupole time‐of‐flight mass spectrometry (LC‐QTOF‐MS) micro‐fractionation method for rapid separation and identification of PDE5A1 inhibitors in 95% ethanolic extract of D. scandens.
Methodology
Initially, the correlation between LC‐MS and PDE5A1 inhibitory activity was studied using three concentrations of 1:1 mixture of sildenafil and derrisisoflavone A; PDE5A1 inhibitors. The mixture was separated by high‐performance liquid chromatography (HPLC) column and the eluent was split into two flows in the ratio of 1:9. The major part was collected in a 96‐well plate, in each well consecutively every 30 s. The minor part was fed into an electrospray ionisation (ESI)‐QTOF‐MS system. After subsequent solvent removal, the collected micro‐fractions were subjected to radioassay to determine PDE5A1 inhibition.
Results
The result showed, PDE5A1 inhibitory activities of the micro‐fractions were observed in a dose response manner and found to be in agreement with an off‐line study. Similarly, 95% ethanolic extract of D. scandens was subjected to the at‐line LC‐QTOF‐MS micro‐fractionation developed, resulting in separation and tentative identification of 25 compounds with PDE5A1 inhibitory activity. Most of the compounds contained prenylated isoflavone skeleton. Additionally, the active micro‐fractions also showed selectivity on PDE5A1 over PDE6 and PDE1B.
Conclusion
Our results demonstrated that the at‐line coupled LC‐QTOF‐MS micro‐fractionation with PDE5A1 inhibitory assay is a valuable tool for identifying PDE5A1 inhibitors from complex extracts.
Chromatographic techniques coupled with bioassays are popularly used for the detection of bioactive compounds in natural products. In this study, at‐line liquid chromatography quadrupole time‐of‐flight mass spectrometry (LC‐QTOF‐MS) micro‐fractionation method was developed for rapid separation and identification of PDE5A1 inhibitors from 95% ethanolic extract of Derris scandens. Initially, the at‐line correlation between LC‐MS and PDE5A1 inhibitory activity was studied using positive controls, after that D. scandens extract was subjected to micro‐fractionation which resulted in separation and tentative identification of 25 compounds with PDE5A1 inhibitory activity.
The aim of this work was to compare two near infrared spectrometers (960–1650 nm) to assess coffee matrices for real-time characterization during processing. A benchtop spectrophotometer built for ...process measurement, equipped with a rotating acquisition system suitable for in-line analyses (device 1), and a portable spectrophotometer for at-line analyses (device 2) were tested. The experimentation was conducted on green, roasted bean, and ground coffee, relating to three coffee blends, employing samples collected at different steps from the process. A total of 399 (247 green, 76 roasted, and 76 ground coffee samples) and 229 (142 green, 43 roasted and 44 ground coffee samples) samples were analyzed using device 1 and device 2, respectively. Principal component analysis was applied to the spectra of the coffee samples to evaluate the feasibility of real-time characterization for each matrix type, during the different steps of the process. Partial least square regression analysis was applied to develop calibrations to predict moisture content, tap density, and powder granulometry on ground coffee and to estimate moisture on roasted bean with a view of real-time measurements. The Passing and Bablok regression method and Bland–Altman analysis were performed to compare results obtained from the tested devices. Considering the models built on different datasets were based on different blends, quality parameters, and matrices, the results obtained from partial least square models, in cross-validation, gave R2 values between 0.21 and 0.98 and between 0.18 and 0.84 for devices 1 and 2, respectively. However, considering the comparison of model results derived from the two devices, no significant differences are noticeable for the most part of the dataset analyzed. Hence, based on the strategy to be undertaken by coffee industry operators, a future real-scale application could be envisaged directly in-line using device 1 or at-line using device 2.
•A novel nanofractionation screening platform was established to screen α-glucosidase inhibitors.•Two orthogonal separation strategies (HILIC & RPLC) were performed in parallel for accurately ...identifying inhibitors.•The sensitivity of the screening platform was improved by multiple nanofractionations.•Ten α-glucosidase inhibitors were successfully screened out from natural products.
In this study, a novel at-line nanofractionation screening platform was successfully developed for the rapid screening and identification of α-glucosidase inhibitors from natural products. A time-course bioassay based on high density well-plates was performed in parallel with high resolution mass spectrometry (MS), providing a straightforward and rapid procedure to simultaneously obtain chemical and biological information of active compounds. Through multiple nanofractionations into the same well-plate and comparisons of the orthogonal separation results of hydrophilic interaction liquid chromatography (HILIC) and reversed-phase liquid chromatography (RPLC), the α-glucosidase inhibitors can be accurately identified from co-eluates. The screening platform was comprehensively evaluated and validated, and was applied to the screenings of green tea polyphenols and Ginkgo folium flavonoids. After accurate peak shape and retention time matching between the bioactivity chromatograms and MS chromatograms, ten α-glucosidase inhibitors were successfully screened out and identified. The proposed screening method is rapid, effective and can avoid ignoring low abundant/active inhibitors.
The paper presents a method for checking the geometry of stamped car body parts using a 3D optical measurement system. The analysis focuses on the first forming operation due to the deformation and ...material flow associated with stall thresholds. An essential element of the analysis is determining the actual gap occurring between the forming surfaces based on the die and punch geometry used in the first stamping operation. The geometry of car body elements at individual production stages was analyzed using an optical laser scanner. The control carried out in this way allowed one to correctly position the tools (punch and die), thus introducing the correction of technological parameters, having a fundamental influence on the specific features of the final product. This type of approach has not been used before to calibrate the technological line and setting of shaping tools. The influence of the manufactured product geometry in intermediate operations on the final geometry features was not investigated.
Human monoamine oxidase B (hMAO-B) has emerged as a pivotal therapeutic target for Parkinson's disease. Due to adverse effects and shortage of commercial drugs, there is a need for novel, highly ...selective, and reversible hMAO-B inhibitors with good blood-brain barrier permeability. In this study, a high-throughput at-line nanofractionation screening platform was established with extracts from Chuanxiong Rhizoma, which resulted in the discovery of 75 active compounds, including phenolic acids, volatile oils, and phthalides, two of which were highly selective novel natural phthalide hMAO-B inhibitors that were potent, selective, reversible and had good blood‒brain permeability. Molecular docking and molecular dynamics simulations elucidated the inhibition mechanism. Sedanolide (IC50 = 103 nmol/L; SI = 645) and neocnidilide (IC50 = 131 nmol/L; SI = 207) demonstrated their excellent potential as hMAO-B inhibitors. They offset the limitations of deactivating enzymes associated with irreversible hMAO-B inhibitors such as rasagiline. In SH-SY5Y cell assays, sedanolide (EC50 = 0.962 μmol/L) and neocnidilide (EC50 = 1.161 μmol/L) exhibited significant neuroprotective effects, comparable to the positive drugs rasagiline (EC50 = 0.896 μmol/L) and safinamide (EC50 = 1.079 μmol/L). These findings underscore the potential of sedanolide as a novel natural hMAO-B inhibitor that warrants further development as a promising drug candidate.
Two hMAO-B natural inhibitors were screened out using a novel at-line nanofractionation platform, demonstrating nanomolar inhibitory activity, highly selective, reversible, BBB permeability, and neuroprotective effects comparable to current drugs. Display omitted
•Two new pharmaceutical co-crystal lamivudine:theophylline polymorphs I and II are synthetized.•Application of FT-IR spectroscopy with MCR-ALS as an at-line process analytical technology is ...demonstrated.•MCR-ALS is applied in FT-IR spectroscopic data for direct quantification of co-crystal polymorph I among its APIs.
This paper, reports for the first time the green synthesis of the polymorphs I and II of new pharmaceutical co-crystals lamivudine:theophylline in solid-phase, through the mixture between lamivudine and theophylline (both active pharmaceutical ingredients-APIs) in the proportion of 1:1 by neat grinding and liquid assisted grinding (10 μL ethanol). Fourier transform-infrared (FT-IR) spectroscopy and multivariate curve resolution with alternating least-squares (MCR-ALS) were employed as non-invasive analytical methodology for the at-line green synthesis monitoring of the novels lamivudine:theophylline co-crystals. By MCR-ALS it was possible to identify each component present in a complex matrix, with strong spectral overlapping, containing lamivudine, theophylline, and the novel lamivudine:theophylline co-crystal with high confidence based on the comparison of the pure and recovered spectral and concentration profiles. This model allowed to identify the end of the reaction and understand the mechanism involved in the synthesis through the identification of the intermediates present in the synthesis process. Also, MCR-ALS model estimated the concentration of co-crystal polymorph I with a root mean square error of prediction (RMSEP) and the percentage relative error of prediction (REP%) equal to 3.323 (w/w) and 9.9%, respectively. These were good results since the spectral profile of cocrystal and the physical mixture of its APIs present strong spectral overlapping in their spectral domain. Therefore, the quantification of the co-crystal between its APIs (lamivudine and theophylline) certified that the co-crystal as final product was obtained, collaborating with the results obtained by differential scanning calorimetry (DSC) and X-ray powder diffraction (XRPD).
•In-line monitoring of the degradation kinetics of hydrolysable drug exploiting the great opportunities offered by ISEs.•ISEs exceptionally provide straightforward degradation profiles and kinetic ...parameters in reference to UV spectrophotometry.•Promising approach to the FDA regulatory requirements for stressing tests.•ISE excels in real-time observation for continuous profile of the hydrolysis behavior under various conditions.
Pharmaceutical product quality is of vital importance for the patients’ safety. In spite of the long history of development of spectroscopic methods for monitoring pharmaceutical products degradation kinetics; this work investigates the opportunities offered by electroanalytical methods (particularly, ion selective electrodes) for in-line monitoring the degradation kinetics compared to at-line monitoring offered by conventional spectroscopic methods. For a meaningful comparison, pyridostigmine bromide (PB) was chosen as hydrolysable anti-cholinesterase drug and two novel strategies for monitoring of PB degradation kinetics catalyzed by hydroxyl ions are presented. The first strategy is achieved by continuous measurement of the decrease in the produced emf over time by incorporation of an in-site PB selective electrode constructed using PVC membrane containing calix6arene as an ionophore. The second strategy utilizes UV spectrophotometry for at-line tracking of either the decrease of PB peak at 269nm or the increase of THMP peak at 320nm over time. The use of these new methods provides real time observation and yields a continuous profile of the hydrolysis behavior of PB under various pH and temperature conditions. Moreover, a great advantage of these proposed in- and at-line systems is their higher accuracy for rate constant estimation relative to other off-line methods.
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