Rapid screening and characterization of caffeic acid metabolites in rats by UHPLC-Q-TOF mass spectrometry

Purpose: To determine the metabolism of caffeic acid in rats. Methods: Sprague-Dawley rats were intragastrically administered caffeic acid in saline suspension, and biological samples collected. After sample pretreatment by solid phase extraction, ultra-high performance liquid chromatography combined with quadrupole-time of flight mass spectrometry system (UHPLC-Q-TOF-MS/MS) was established to rapidly screen and characterize caffeic acid metabolites in rats. Waters HSS T3 UPLC chromatographic column (2.1 mm × 100 mm, 1.7 μm) was applied for the gradient elution with aqueous solution of formic acid (A)-acetonitrile (B). Mass spectral data for the biological samples in electrospray positive and negative ion modes were collected and analyzed by SCIEX OS 1.3 workstation. Results: Based on their precise molecular weights and multistage mass spectrometry cleavage information, caffeic acid and 21 metabolites in vivo were identified. The results demonstrate that the biotransformation of caffeic acid in vivo was mainly achieved via hydrogenation, hydroxylation, methylation, sulfonation, glucuronidation, acetylation, and composite reactions. Conclusion: The metabolites and metabolic pathways of caffeic acid in rats have been rapidly elucidated, and its potential pharmacodynamics forms have been clarified. This provides a valuable and meaningful reference for the study of caffeic acid metabolites, biological activities, and its medicinal material basis in vivo.

Studies of the pharmacokinetics of morroniside and loganin in rat after oral administration of raw and wine-processed Corni fructus by UPLC-QqQ-MS/MS

Purpose: To study the pharmacokinetics of morroniside (MR) and loganin (LG) in rats after oral administration of raw and wine-processed Corni fructus by UPLC-QqQ-MS/MS. Methods: Arctiin (AT) was used as internal standard, and the plasma or tissue samples were extracted twice using ethyl acetate. Electrospray ionization (ESI) negative ion mode was used, and the multiple reaction monitoring mode (MRM) was set in acquisition mode. The extraction and detection method is supported by selectivity, sensitivity, precision, accuracy, stability, extraction, recovery and matrix effect. The non-atrioventricular model of das2.0 software was used to calculate the pharmacokinetic parameters. Results: The absorption rate of MR (PTmax=0.092) and LG (PTmax=0.092) in Corni Fructus after wine-processing was faster in rats. The mean residence time was longer, and distribution of MR (PMRT0~t = 0.294) and LG (PMRT0~t = 0.000) in wine-processed Corni Fructus group increased in liver and kidneys. Conclusion: The proposed method has been successfully validated and is suitable for studying the pharmacokinetics of the two analytes in rats.

Association of Differential Metabolites With Small Intestinal Microflora and Maternal Outcomes in Subclinical Hypothyroidism During Pregnancy

ObjectiveTo investigate the association of differential metabolites with small intestinal microflora and maternal outcomes in subclinical hypothyroidism (SCH) during pregnancy.MethodsThe plasma of pregnant women in the SCH group and control group was analyzed by liquid chromatography-mass spectrometry (LC-MS), obtaining differential metabolites. Then, methane and hydrogen breath tests were performed in both groups, and basic clinical data and maternal outcome information were collected. Finally, differential metabolites were analyzed for small intestinal bacterial overgrowth (SIBO) and pregnancy outcomes using Spearman correlation analysis.Results(1) Multivariate statistics: There were 564 different metabolites in positive ion mode and 226 different metabolites in negative ion mode. (2) The positive rate of the methane hydrogen breath test in the SCH group was higher than that in the control group (p<0.05). (3) KEGG pathway analysis revealed that differential metabolites were mainly involved in bile secretion, cholesterol metabolism, and other pathways. (4) Serum cholesterol (TC) and triglyceride (TG) levels and hypertensive disorder complicating pregnancy (HDCP) were higher in the SCH group (p<0.05), and newborn birth weight (BW) was lower than that in the control group (p<0.05). (5) SIBO was negatively correlated with glycocholic acid and BW, and positively correlated with TC. Glycocholic acid was negatively correlated with TG but positively correlated with BW. TG was positively correlated with HDCP.ConclusionDifferential metabolites in the SCH group during pregnancy were disordered with small intestinal bacteria, which may affect pregnancy outcomes, and bile acids and cholesterol may be potential biomarkers for studying their mechanism of action.

Hybrid simulation of instabilities in capacitively coupled RF CF4/Ar plasmas

Radio frequency capacitively coupled plasmas (RF CCPs) sustained in fluorocarbon gases or their mixtures with argon are widely used in plasma-enhanced etching. In this work, we conduct studies on instabilities in a capacitive CF4/Ar (1:9) plasma driven at 13.56 MHz at a pressure of 150 mTorr, by using a one-dimensional fluid/Monte-Carlo (MC) hybrid model. Fluctuations are observed in densities and fluxes of charged particles, electric field, as well as electron impact reaction rates, especially in the bulk. As the gap distance between the electrodes increases from 2.8 cm to 3.8 cm, the fluctuation amplitudes become smaller gradually and the instability period gets longer, as the driving power density ranges from 250 to 300 W/m2. The instabilities are on a time scale of 16-20 RF periods, much shorter than those millisecond periodic instabilities observed experimentally owing to attachment/detachment in electronegative plasmas. At smaller electrode gap, a positive feedback to the instability generation is induced by the enhanced bulk electric field in the highly electronegative mode, by which the electron temperature keeps strongly oscillating. Electrons at high energy are mostly consumed by ionization rather than attachment process, making the electron density increase and overshoot to a much higher value. And then, the discharge becomes weakly electronegative and the bulk electric field becomes weak gradually, resulting in the continuous decrease of the electron density as the electron temperature keeps at a much lower mean value. Until the electron density attains its minimum value again, the instability cycle is formed. The ionization of Ar metastables and dissociative attachment of CF4 are noticed to play minor roles compared with the Ar ionization and excitation at this stage in this mixture discharge. The variations of electron outflow from and negative ion inflow to the discharge center need to be taken into account in the electron density fluctuations, apart from the corresponding electron impact reaction rates. We also notice more than 20% change of the Ar+ ion flux to the powered electrode and about 16% difference in the etching rate due to the instabilities in the case of 2.8 cm gap distance, which is worthy of more attention for improvement of etching technology.

UHPLC-HRMS-Based Untargeted Lipidomics Reveal Mechanism of Antifungal Activity of Carvacrol against Aspergillus flavus

Aspergillus flavus is a common contaminant in grain, oil and their products. Its metabolite aflatoxin B1 (AFB1) has been proved to be highly carcinogenic. Therefore, it is of great importance to find possible antifungal substances to inhibit the growth and toxin production of Aspergillus flavus. Carvacrol (CV) was reported as a potent antifungal monoterpene derived from plants. In this paper, the antifungal effects and mechanism of CV on Aspergillus flavus were investigated. CV was shown good inhibition on the growth of Aspergillus flavus and the production of AFB1. CV used in concentrations ranging from 0, 50, 100 and 200 μg/mL inhibited the germination of spores, mycelia growth and AFB1 production dose-dependently. To explore the antifungal mechanism of CV on Aspergillus flavus, we also detected the ergosterol content of Aspergillus flavus mycelia, employed Scanning Electron Microscopy (SEM) to observe mycelia morphology and utilized Ultra-High-Performance Liquid Chromatography-High-Resolution Mass Spectrometry (UHPLC-HRMS) to explore the lipidome profiles of Aspergillus flavus. The results showed that the production of ergosterol of mycelia was reduced as the CV treatment concentration increased. SEM photographs demonstrated a rough surface and a reduction in the thickness of hyphae in Aspergillus flavus treated with CV (200 µg/mL). In positive ion mode, 21 lipids of Aspergillus flavus mycelium were downregulated, and 11 lipids were upregulated after treatment with 200-µg/mL CV. In negative ion mode, nine lipids of Aspergillus flavus mycelium were downregulated, and seven lipids upregulated after treatment with 200-µg/mL CV. In addition, the analysis of different lipid metabolic pathways between the control and 200-µg/mL CV-treated groups demonstrated that glycerophospholipid metabolism was the most enriched pathway related to CV treatment.

Pharmacokinetics Study of Rabdosia Rubescens Drop Pills Based on UPLC-MS/MS

Background: Rabdosia rubescens drop pills have the effects of clearing away heat and toxin, detumescence, relieving pain. Objective: A simple and sensitive method for simultaneous determination of oridonin, ponicidin, and rosmarinic acid in rat plasma was developed based on ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). Methods: Chromatographic separation was performed on a Waters ACQUITY UPLC BEH C18 column (2.1 × 100 mm, 1.7 µm) with a mobile phase consisting of water containing 0.2% formic acid (mobile phase A) and methanol (mobile phase B) at a flow rate of 0.3 mL/min over a total run time of 3.8 min. All analytes were measured with optimized multiple reaction monitoring (MRM) in positive and negative ion ESI mode. Results: The transitions of oridonin, ponicidin, rosmarinic acid, diphenhydramine, and chloramphenicol were 365.3→347.3, 363.3→345.2, 359.0→160.9, 256.0→167.2, and 321.1→151.9, respectively. The linear ranges were 1-256 ng/mL for ponicidin and rosmarinic acid and 2-512 ng/mL for oridonin. The validated method wasstable and reliable. There was no significant difference in the half-life (t1/2) of the three analytes at three doses. The area under the curve (AUC0-t) and peak concentration (Cmax) of the three analytes decreased linearly in each dose range, and the linear correlation R2 of each analyte under the three doses was greater than 0.95. Conclusion: This method was successfully applied to pharmacokinetic studies of oridonin, ponicidin, and rosmarinic acid in rat plasma after intragastric administration of Rabdosia rubescens drop pills.

Negative Ion Purifier Effects on Indoor Particulate Dosage to Small Airways

Indoor air quality is an important health factor as we spend more than 80% of our time indoors. The primary type of indoor pollutant is particulate matter, high levels of which increase respiratory disease risk. Therefore, air purifiers are a common choice for addressing indoor air pollution. Compared with traditional filtration purifiers, negative ion air purifiers (NIAPs) have gained popularity due to their energy efficiency and lack of noise. Although some studies have shown that negative ions may offset the cardiorespiratory benefits of air purifiers, the underlying mechanism is still unclear. In this study, we conducted a full-scale experiment using an in vitro airway model connected to a breathing simulator to mimic inhalation. The model was constructed using computed tomography scans of human airways and 3D-printing technology. We then quantified the effects of NIAPs on the administered dose of 0.5–2.5 μm particles in the small airway. Compared with the filtration purifier, the NIAP had a better dilution effect after a 1-h exposure and the cumulative administered dose to the small airway was reduced by 20%. In addition, increasing the negative ion concentration helped reduce the small airway exposure risk. NIAPs were found to be an energy-efficient air purification intervention that can effectively reduce the small airway particle exposure when a sufficient negative ion concentration is maintained.

Lipidomic profile of seminal plasma in non-obstructive azoospermia with sperm maturation arrest

Introduction. The difference between obstructive and non-obstructive azoospermia with sperm maturation arrest is important for the choice of treatment tactics and adequate counseling of a married couple.Purpose of the study. The study aimed to assess the semen lipid profile in patients with sperm maturation arrest. Materials and methods. Samples of seminal plasma for lipid composition of 24 men with normozoospermia and 64 men with azoospermia were studied. Patients with azoospermia underwent microdissection testicular biopsy followed by the detection of testicular tissue pathology. Lipid extracts were analyzed by liquid chromatography with mass spectrometry. Lipid data were compared with the results of pathomorphological studies.Results. Comparison of two groups revealed a statistically significant concentration differences for 22 lipids detected in positive-ion mode and 11 lipids detected in negative-ion mode. Those lipids mainly belong to the classes hexosylceramides, sphingomyelins and phosphatidylcholines — simple ethers and oxidized lipids. In multivariate analysis, the following lipids were found to be statistically significant predictors of sperm maturation arrest: PC 16: 0_22: 6 lipid (β-coefficient: -0.73; 95% confidence interval (95% CI): -1.42 to -0.27; odds ratio (OR): 0.48; OR CI: 0.24 to 0.76; Wald's test: -2.58; p = 0.01), SM d20: 1/22:2 lipid (β-coefficient 4.96; 95% CI 2.29 to 9.13; OR: 142.31; OR CI: 9.90 to 9.22^103; Wald's test: 2.93; p = 0.003); PG 20:3_22: 6 lipid (β-coefficient 2.52; 95% CI 1.13 to 4.49; OR: 12.37; OR CI: 3.10 to 89.27; Wald's test: 3.02; p = 0.002); PC O- 16: 1/16:0 lipid (β-coefficient 1.96; 95% CI -4.12 to 0.27; OR: 0.14; OR CI: 0.02 to 0.76; Wald's test: -2.05; p = 0.04). The prediction model characteristics of sperm maturation arrest, obtained during cross-validation in the positiveion mode composed: sensitivity 91%, specificity 85%; in negative-ion mode: sensitivity 75%; specificity 81%.Conclusions. Even though early stages of spermatogenesis are equally preserved in both fertile men and men with homogeneous sperm maturation arrest, the semen in the studied group of patients differed in its lipid profile. Patients with non-obstructive azoospermia, associated with meiosis arrest, may have unique lipidomic characteristics of seminal plasma, which in the future will make it possible to differentiate various variants of severe male infertility using non-invasive methods.

Alfvén cascade eigenmodes above the TAE-frequency and localization of Alfvén modes in D- 3 He plasmas on JET

Various types of Alfvén Eigenmodes (AEs) have been destabilized by fast ions over a broad frequency range from ~80 kHz to ~700 kHz in a series of JET experiments in mixed D-3He plasmas heated with the three-ion ICRF scenario [M. Nocente et al., Nucl. Fusion 60, 124006 (2020)]. In this paper, we identify the radial localization of AEs using an X-mode reflectometer, a multiline interferometer and soft X-ray diagnostics. The analysis is focused on the most representative example of these measurements in JET pulse #95691, where two different types of Alfvén cascade (AC) eigenmodes were observed. These modes originate from the presence of a local minimum of the safety factor qmin. In addition to ACs with frequencies below the frequency of toroidal Alfvén eigenmodes (TAEs), ACs with frequencies above the TAE frequency were destabilized by energetic ions. Both low- (f ≈80-180 kHz) and high-frequency (f ≈ 330-450 kHz) ACs were localized in the central regions of the plasma. The characteristics of the high-frequency ACs are investigated in detail numerically using HELENA, CSCAS and MISHKA codes. The resonant conditions for the mode excitation are found to be determined by passing ions of rather high energy of several hundred keV and similar to those established in JT-60U with negative-ion-based NBI [M. Takechi et al., Phys. Plasmas 12, 082509 (2005)]. The computed radial mode structure is found to be consistent with the experimental measurements. In contrast to low-frequency ACs observed most often, the frequency of the high-frequency ACs decreases with time as the value of qmin decreases. This feature is in a qualitative agreement with the analytical model of the high-frequency ACs in [B.N. Breizman et al., Phys. Plasmas 10 3649 (2003)]. The high-frequency AC could be highly relevant for future ITER and fusion reactor plasmas dominated by ~ MeV energetic ions, including a significant population of passing fast ions.

Short communication: Distribution of phospholipids in parotid cancer by matrix-assisted laser desorption/ionization imaging mass spectrometry

Background Parotid cancer is relatively rare, and malignancy varies; therefore, novel markers are needed to predict prognosis. Recent advances in matrix-assisted laser desorption/ionization imaging mass spectrometry (MALDI-IMS), useful for visualization of lipid molecules, have revealed the relationship between cancer and lipid metabolism, indicating the potential of lipids as biomarkers. However, the distribution and importance of phospholipids in parotid cancer remain unclear. Objective This study aimed to use MALDI-IMS to comprehensively investigate the spatial distribution of phospholipids characteristically expressed in human parotid cancer tissues. Methods Tissue samples were surgically collected from two patients with parotid cancer (acinic cell carcinoma and mucoepidermoid carcinoma). Frozen sections of the samples were assessed using MALDI-IMS in both positive and negative ion modes, with an m/z range of 600–1000. The mass spectra obtained in the tumor and non-tumor regions were compared and analyzed. Ion images corresponding to the peak characteristics of the tumor regions were visualized. Results Several candidate phospholipids with significantly different expression levels were detected between the tumor and non-tumor regions. The number of unique lipid peaks with significantly different intensities between the tumor and non-tumor regions was 95 and 85 for Cases 1 and 2, respectively, in positive ion mode, and 99 and 97 for Cases 1 and 2, respectively, in negative ion mode. Imaging differentiated the characteristics that phospholipids were heterogeneously distributed in the tumor regions. Conclusion Phospholipid candidates that are characteristically expressed in human parotid cancer tissues were found, demonstrating the localization of their expression. These findings are notable for further investigation of alterations in lipid metabolism of parotid cancer and may have potential for the development of phospholipids as biomarkers.